We have previously documented significant differences in orthogonal P wave morphology between patients with and without paroxysmal atrial fibrillation (PAF). However, there exists little data concerning normal P wave morphology. This study was aimed at exploring orthogonal P wave morphology and its variations in healthy subjects.120 healthy volunteers were included, evenly distributed in decades from 20–80 years of age; 60 men (age 50+/-17) and 60 women (50+/-16). Six-minute long 12-lead ECG registrations were acquired and transformed into orthogonal leads. Using a previously described P wave triggered P wave signal averaging method we were able to compare similarities and differences in P wave morphologies.Orthogonal P wave morphology in healthy individuals was predominately positive in Leads X and Y. In Lead Z, one third had negative morphology and two-thirds a biphasic one with a transition from negative to positive. The latter P wave morphology type was significantly more common after the age of 50 (P < 0.01). P wave duration (PWD) increased with age being slightly longer in subjects older than 50 . Minimal intraindividual variation of P wave morphology was observed.Changes of signal averaged orthogonal P wave morphology , earlier reported in PAF patients, are common in healthy subjects and appear predominantly after the age of 50. Subtle age-related prolongation of PWD is unlikely to be sufficient as a sole explanation of this finding that is thought to represent interatrial conduction disturbances. To serve as future reference, P wave morphology parameters of the healthy subjects are provided. Atrial fibrillation (AF) is the most common sustained arrhythmia of clinical importance. Over the last couple of decades, many important findings have contributed to the enhanced understanding of AF pathophysiology, but much still remains to be clarified. AF is normally characterized as permanent, persistent or paroxysmal (PAF), with the latter being more common in younger persons. Both variants might be asymptomatic but, unfortunately, PAF is none the less still associated with considerably increased mortality and morbidity, thus implying the need for early diagnosis and subsequent prophylactic measures .Using band-pass filtered signal averaging technique, P wave duration (PWD) has been studied in PAF patients with inconsistent findings regarding possible clinical applications -7. The mThe study population was recruited by advertising in local press. The subjects were interviewed and examined by a physician to ensure good health. A 12-lead ECG was performed and blood pressure was measured. Exclusion criteria included history of cardiovascular disease , excessive use of alcohol, or a pathological 12-lead ECG at rest, including non-sinus rhythm. A total of 120 healthy individuals were included. The study was approved by the Ethics Committee of Lund University . Written informed consent was obtained and the study complied with the Declaration of Helsinki.A standard 12-lead ECG was recorded for 6 minutes with the subjects at rest. Data was acquired at a rate of 1 kHz and with a resolution of 0.625 μV. The data acquisition board used for the recordings was supplied by Siemens-Elema AB, Solna, Sweden.® , was used for signal processing and analysis. The method has been described in detail earlier ) was defined as the difference, in time, between the P wave onset and end.In Leads X and Y, the maximum amplitude was determined  and also the distance from the onset of the P wave to the position of these maxima . In Lead Z, the maximum and minimum amplitude was determined  along with their locations . In addition to this, the location of the zero crossing in Lead Z was measured when applicable (Z zero pos). The different parameters and a schematic illustration of the method used are shown in Figure All data is presented as mean ± standard deviation unless stated otherwise. Statistic evaluation was performed using the software StatView 4.5 . The Mann-Whitney U-test and Kruskal-Wallis one-way analysis of variance were used for comparisons between populations of continuous data. For comparisons between groups of proportions the Chi-square test was used. A P-value less than 0.05 was considered statistically significant.The study group was comprised by 60 men (age 50 ± 17 years) and 60 women (50 ± 16), (n.s.). The subjects were evenly distributed in groups of ten men and women for every decade from 20 years of age to 80. All subjects presented data, i.e. 6-minute registrations, available for analysis.The males were significantly taller and heavier than the females . They also had lower heart rate  and higher systolic and diastolic blood pressure . With increasing age, a slight increase of systolic and diastolic blood pressure was also observed but still within normal limits.P wave duration (PWD) was longer in males . PWD also increased significantly with increasing age (P < 0.05). When comparing PWD between subjects aged below and above 50, a slight but significant increase was observed in the older group . Baseline data is presented in Table With the exception of higher P wave amplitude in Lead Y (Y max) for females compared to males, the only significant gender or age-differences regarding amplitudes or positions of P wave maxima or minima were seen in Lead Z. Here females had lower amplitudes of maxima and also earlier location of minima. With older age, the amplitude of the maxima in Lead Z increased while the zero-crossing was located earlier thus making the positive phase of the P wave in Lead Z greater. The numerical data and P-values of PWD as well as the amplitudes and temporal markers of X max, Y max, Z max, Z min and Z zero are shown in Table Of the 120 subjects, 118 had P wave morphology types consistent with those observed in earlier reports; 45 Type 1 , and 73 Type 2 . Two subjects  had P wave morphologies that were classified as atypical, i.e. neither Type 1, 2 or 3.When comparing the distribution of P wave morphology types in the different age groups it was found that the Type 2 morphology was more common with increasing age (P < 0.01). There was also a significant (P < 0.001) increase in Type 2 morphology in subjects older than 50 when comparing equally large groups of subjects younger and older than 50. In the younger group 26 of 60 cases of Type 2 morphology were observed in comparison to 50 of 60 in the older group. The distributions of P wave morphology types in different age groups are illustrated in Figure In 92 % of the cases only one group of P wave morphologies was observed during 6-min long recording. In the remaining cases (8%), additional groups of P waves with a cross-correlation coefficient < 0.90 were obtained. The P waves from different groups appeared to be very similar in shape and of the same morphological type Figure .In the present study, orthogonal ECG registrations from 120 healthy adults of varying age were analysed using PSAECG. Two main P wave morphologies were observed (Type 1 and 2). Type 2 morphology, associated with PAF and other arrhythmia-prone patients, was more common in healthy subjects than earlier perceived  and highIn a report on the anatomy of the human atria Ho and co-workers denoted BB, the muscular fibre band that abridges the anterior inter-atrial groove, the "superhighway for inter-atrial conduction" . HoweverFrom an electrophysiological standpoint, BB has been considered the preferential conductive route from RA to LA but other transseptal pathways, close to Koch's triangle and also in the CS area have been recognized . The funPSAECG has previously been used by our group to study orthogonal P wave morphology in patients with PAF, HCM and control patients ,21. ThreIn the reported material, Type 2 morphology is significantly more common with increasing age with a marked increase in prevalence after the age of 50. We did not observe any unexpected changes of the characteristics of the study group that would explain these findings that implicate altered RA-LA activation. For instance, only slight prolongation of mean PWD (ΔPWD = 7 ms) with age was observed and the longer PWD in men could possibly be explained by their heavier stature. As explained above, interatrial conduction can occur via several pathways. One could speculate that age-related changes in the human heart such as fibrosis that have been reported and suggested to play a role in atrial reentrant mechanisms  could afHowever, other causes of altered P wave morphology are also plausible. These include variations in the atrial pacemaker complex and sinus node morphology but also differences of conduction velocities . FurtherHowever, intraindividual variation of P wave morphology during 6-min long recordings was minimal in the present study. This could indicate that interatrial conduction propagation on an individual basis is stable and that the changes that occur are stable as well. These age-related changes in P wave morphology are therefore unlikely to be caused solely by the age-related decrease in conduction velocity reflected in marginal prolongation of PWD, but could best be explained by changes in the function of interatrial conductive pathways.Even though the selection process was aimed at including only healthy asymptomatic subjects with normal blood pressure, no known diseases and not taking any medication it cannot be ruled out that some subjects with asymptomatic AF may have been included. However, in our opinion possible impact of these single individuals on the overall results would be negligible. Furthermore, influence of atrial dimensions on P wave morphology that has been demonstrated earlier cannot be excluded since echocardiography data for the study group was not available.The presented study describes orthogonal P wave morphology of a material comprised of healthy men and women and provides a reference material for future research. Changes of signal averaged orthogonal P wave morphology , earlier reported in PAF patients, are common in healthy subjects and appear predominantly after the age of 50. Subtle age-related prolongation of PWD is unlikely to be sufficient as a sole explanation of this finding that is thought to represent interatrial conduction disturbances. However, further studies are needed to determine the electrophysiological explanation for these changes and its possible role as a substrate for arrhythmia.The author(s) declare that they have no competing interests.RH participated in the processes of study design, data acquisition and analysis. JC, AH and PP contributed to study design, data acquisition and analysis. CM and BO took part in study design and analysis. All authors participated in the process of drafting the manuscript as well as read and approved the final manuscript.The pre-publication history for this paper can be accessed here:


Heart chambers rupture in blunt trauma is uncommon and is associated with a high mortality. The determinant factors, and the incidence of isolated heart chambers rupture remains undetermined. Isolated rupture of the right atrium appendage (RAA) is very rare, with 8 cases reported in the reviewed literature. The thin wall of the RAA has been presumed to render this chamber more prone to rupture in blunt trauma.To report a case of isolated RAA rupture in blunt trauma, and to compare right atrium (RA) and RAA wall thickness in a necropsy study.The thickness of RA and RAA wall of hearts from cadavers of fatal penetrating head trauma victims was measured. Our case of isolated RAA rupture is presented. The main findings of the 8 cases reported in the literature, and the findings of our case, were organized in a table.The comparison of the data showed that wall thickness of the RAA (0.53 ± 0.33 mm) was significantly thinner than that of RA (1.11 ± 0.42 mm) (p < 0.05).In all these 9 cases of isolated RAA rupture, cardiac tamponade occurred, RAA rupture was diagnosed intraoperatively and sutured, and the patients survived. Main mechanisms hypothesized for heart chamber rupture include mechanical compression coincident with phases of cardiac cycle, leading to high hydrostatic pressure inside the chamber. Published series include numerous cases of RA rupture, and only a few cases of RAA rupture.Thus, our data suggests that wall thickness is not a determinant factor for RA or RAA rupture in blunt trauma. Traumatic rupture of the heart occurs in 0.5% blunt trauma (BT)  cases, wobjectives of this study were: 1) To compare the thickness of the RAA and RA walls of hearts of lethal head trauma victims; 2) to present a case report of isolated BT rupture of the RAA, and discuss the role of wall thickness as a possible factor.Among the diverse mechanisms proposed for traumatic rupture of heart chambers ,9,12,13,To evaluate the wall thickness of RAA and RA of trauma victims, twenty-eight hearts that had been removed during autopsy, at the Legal Medicine Institute of the USPSM, from cadavers of penetrating lethal head trauma victims aged from 14 to 84 (mean 31) years, were studied. The hearts were preserved in formaldehyde for anatomic study at the MAPD of the INCOR, CH-USPSM. The study was performed according to the norms of the Ethics Committee of the institution. With the use of a pachymeter, the RAA wall thickness was measured at the apex of this chamber. The RA wall thickness was measured at the antero-superior wall of the RA, at 1 cm from the base of the RAA. The data obtained for the RA and RAA wall thickness (mm) were expressed as mean ± SEM, and compared using Student's t test. P values < 0.05 were considered significant.The wall thickness obtained for the RA was 1.11 ± 0.42 mm, and for the RAA was 0.53 ± 0.33 mm , and the comparison of these results showed that RAA thickness was significantly smaller than that of RA (p < 0.05).20. Cardiac tamponade was suspected, and confirmed through a subxiphoid pericardial window. A thoracotomy was performed, with pericardiotomy and removal of blood clots from the pericardial cavity. A 0.5 cm bleeding tear in the apex of the RAA was found, and closed with sutures. The thoracotomy incision was closed. The patient was discharged from the hospital on the 20th postoperative day. His injury severity score (ISS) was 50, and his TRISSCAN was 0.31.A 16-year old male, unrestrained passenger was a victim of high speed frontal collision of the car with an electricity pole, and 30 min thereafter was admitted to the emergency room (ER). On admission, he was complaining of thoracic and abdominal pain, and presented free airways, normal chest expansion, BP:70 × 40 mmHg, HR: 120 bpm, RR: 18 epm, GCS: 15, RTS: 6.4, abdominal tenderness, and a closed right femur fracture. Plain X ray films suggested a widened mediastinum. After 3000 mL of warmed saline infusion, his BP was 60 × 30 mmHg, and his HR was 120 bpm. A diagnostic peritoneal lavage was positive for blood. As ultrasonography was unavailable, a laparotomy was performed, with splenectomy and cauterization of a superficial liver injury. After infusion of 6000 more mL saline, 8 units PRBC, and 9 units fresh frozen plasma, the patient's conditions worsened, with muffled heart sounds, undetectable peripheral pulses, and CVP of 20 cmHThe main data of this case are presented (see Table The 9 cases (see Table) comprised 5 men and 4 women, aged 15 to 34 (mean: 28) years, all involved in high speed MVC (8 cars and 1 motorcycle). The victims were a motorcyclist (case 1), unrestrained car drivers , or passengers (cases 7 and 9), or unspecified position (case 6). Overall, the main diagnostic findings in these 9 cases included hypotension or persistent unresponsive shock , muffled heart sounds , high CVP values , neck vein distension , thoracic imaging alterations , and traumatic ascitis (case 7). Cardiac tamponade was present in the 9 cases. All the patients underwent a thoracotomy allowing RAA injury identification and repair, and all patients survived. The length of hospital stay ranged from 8 to 20 days.Blunt trauma rupture of heart chambers is more frequent in MVC with great deceleration leading to high energy impact of drivers' thorax on the steering wheel -20. In tAlthough not supported by clinical or experimental data, diverse mechanisms were proposed for heart rupture in BT, including: sudden deceleration , the occAccording to some authors, ventricular versus atrial rupture depends on the phase of the cardiac cycle, atrium rupture being more likely to occur under a forceful compression in late systole, when the atria are most distended with venous blood, and the atrioventricular valves are closed ,24. Giveconclusion, the findings of our study indicate that the RAA wall is thinner than that of the RA. However, given the reported higher incidence of RA rupture than of RAA rupture, our findings do not suggest wall thickness as a predominant factor either for RA or RAA rupture in blunt trauma.In Concerning the present manuscript, the authors declare that there are no personal, or financial, or non-financial competing influence, interests, or conflicts involving them with other people or organizations.


Dendritic cells (DC) are important for MHC class I processing and presentation of peptide epitopes to memory CD8+ T cells, and could potentially be targeted to activate memory CD8+ T cells to a broad array of HIV-1 epitopes during ART.HIV-1 remains sequestered during antiretroviral therapy (ART) and can resume high-level replication upon cessation of ART or development of drug resistance. Reactivity of memory CD8+ T cells specific for a much broader range and magnitude of Gag and Nef epitopes than do peptides without DC. The DC also reveal novel, MHC class I restricted, Gag and Nef epitopes that are able to induce polyfunctional T cells producing various combinations of IFN gamma, interleukin 2, tumor necrosis factor alpha, macrophage inhibitory protein 1 beta and the cytotoxic de-granulation molecule CD107a.We show for the first time that HIV-1 peptide-loaded, CD40L-matured DC from HIV-1 infected persons on ART induce IFN gamma production by CD8+ T cell reactivity in persons on ART that is revealed by DC. This supports the use of DC-based immunotherapy for HIV-1 infection.There is an underlying, broad antigenic spectrum of anti-HIV-1, memory CD8 Our results show that HIV-1 peptide-loaded, mature DC induced IFNγ production to a much broader range of HIV-1 Gag and Nef epitopes than did peptides without DC. The MHC class I restricted Gag and Nef epitopes included novel ones that could activate polyfunctional T cells producing various combinations of IFNγ interleukin 2 (IL-2), TNFα, macrophage inhibitory protein 1β (MIP-1β) and the cytotoxic de-granulation molecule CD107a. This indicates that there is a broader and more robust array of memory CD8+ T cells specific for HIV-1 antigens circulating in persons on ART than has previously been appreciated, and supports use of DC-based immune therapies.Induction of broad and robust T cell reactivity could be particularly important in immunotherapy of HIV-1 infection during antiretroviral therapy (ART) This research was part of the Pittsburgh Multicenter AIDS Cohort Study (MACS), an investigation of the natural history of HIV infection, and was approved by the University of Pittsburgh Institutional Review Board. 7 HIV-1 seropositive homosexual men on ART were randomly selected for study from the Pittsburgh, PA, portion of the MACS . Four HI+ monocytes were positively selected from peripheral blood mononuclear cells (PBMC) using anti-CD14 monoclonal antibody (mAb)-coated magnetic microbeads  to a purity of >96%, cultured for 5 to 6 days in AIM V medium  containing 1000 U/ml of recombinant IL-4  and 1000 U/ml of recombinant granulocyte-monocyte colony stimulating factor (GM-CSF) . Fresh IL-4 and GM-CSF were added every other day. The DC were treated with maturation factor CD40L  for 40 h to induce DC maturation.To obtain immature DC, CD14The number of viable DC was determined by typical morphology in trypan blue dye-stained preparations. The maturation status of the DC was determined by flow cytometry as the percent positive and mean fluorescent intensity of expression of MHC class II (HLA-DR), MHC class I (HLA ABC), CD80, CD86 and CD83. Viable DC displayed a characteristic DC morphology and cell surface marker expression and responded to stimulation with CD40L.+ T Cell Epitope Database A library of HIV-1 peptides (consecutive 15mers overlapping by 11 amino acids) spanning the consensus B HIV-1 proteome was obtained through the AIDS Research and Reference Reagent Program, Division of AIDS, NIAID, NIH. These were used as singlets or in pools of consecutive peptides. Known, “A-list” epitopes were identified by the Los Alamos CTL/CD8°C. Responder cells were autologous PBMC or CD8+ T cells (96–98% pure) positively selected from PBMC using anti-CD8 mAb-coated, magnetic microbeads (StemCell). The peptide-loaded DC (stimulators) were washed to remove excess peptide and mixed with responder cells at a responder-to-stimulator [R:S] cell ratio of 10 to 1 and incubated for 18 h with peptide-loaded DC at 37°C in a 5% CO2 atmosphere. The wells were washed with PBS and treated with biotinylated anti-IFN-γ mAb  and incubated at 37°C for 3 hours. Avidin-peroxidase (100 µl/well) was added after the biotinylated antibody was decanted and the plates were washed four times with PBS-0.05% Tween 20 . Diaminobenzidine solution  was added to each well for 5 minutes at room temperature. The plates were washed and air-dried overnight. A negative control (medium without peptides), and 2 positive controls – CEF (1 µg/ml), which is a mixture of human cytomegalovirus, Epstein-Barr virus and influenza A virus 6 cells. The results were considered positive if the number of SFC in the peptide-stimulated cultures was more than 50 and above the mean plus two standard deviations of SFC in cultures with medium alone.An ELISPOT assay modified from AIDS Clinical Trials Group protocol A5181 was used to determine single cell IFNγ production In some experiments, HLA restriction of the T cell responses was confirmed by ELISPOT assay using a panel of EBV-transformed B cell lines (BLCL) matched with the effector cells at only one MHC class I allele.6/ml in RPMI-10% FCS and rested overnight at 37°C. The PBMC were then cultured with 2 µl each of T cell costimulatory mAb specific for CD28 and CD49d , monensin  and brefeldin A  to inhibit extracellular release of the immune mediator CD107a-PECy5 , and peptides or peptide pools (5 µg/ml). In some experiments, DC loaded with peptide (5 µg/ml) were used at a 1∶10 ratio with PBMC but without αCD28/49d. Negative controls (without peptides) and positive controls  were included in each assay. Cells were incubated for 6 h at 37°C and then kept at 4°C for 16 h. The cells were washed, fixed using the Cytofix/Cytoperm kit (BD PharMingen) and stained with mAb CD8-APC Cy7, CD4-APC Cy7, IL-2-APC (BD Biosciences), CD3-PE Cy7, IFNγ-FITC, MIP-1β-PE (BD PharMingen) and TNFα-PB (eBiosciences). Following staining, the cells were washed, fixed and analyzed with an LSR II flow cytometer (BD Immunocytometry Systems), with 200,000 to 1,000,000 events collected per sample. T lymphocyte subsets were analyzed by first identifying and gating the whole lymphocyte population according to light scatter properties (FSC and SSC), followed by gating T cell subsets based on the expression of the surface markers CD3 and CD8, as well as the intracellular expression of IL-2, IFNγ, TNFα, MIP-1β and CD107a compared to negative controls. All data were background-subtracted using the non-antigen stimulated control and analyzed by FlowJo  and SPICE (version 4.1.6). The expression of CD107a, IFNγ, IL-2, MIP-1β, TNFα and T cell surface markers was quantified separately and in combination.Frozen-thawed PBMC were suspended to 2×10+ T cell responses to HIV-1 peptides are stronger in the presence than in the absence of DC; (b) CD8+ T responses to HIV-1 peptides exceed PBMC responses both in the presence and in the absence of DC; and (c) relative changes in the level of response to HIV-1 peptides is highly correlated between CD8+ T cells and PBMC both in the presence and the absence of DC. The first two hypotheses were tested using the binomial sign test. In each case, the peptide data was transformed into a binary variable and tested against the null hypothesis (H0 = 0.5) using a one-sided alternative. For the first two hypotheses, we also calculated mean ratios and 95% confidence intervals in order to provide a better sense of the size of the comparative responses. For the third hypothesis, we ranked the size of the response observed to each of the HIV-1 peptides by PBMC and CD8+ T cells both in the presence and the absence of DC. Under both of the latter conditions (presence or absence of DC), we calculated Spearman rank order correlations between the PBMC and CD8+ T cell responses and the statistical significance  of each correlation. We used the Scheffe multiple comparison test and chi-square test for analysis of the polyfunctional T cell results.Statistical analyses were carried out to test three specific hypotheses: (a) PBMC and CD8+ T cell responses + T cells from 3 HIV-1 seronegative, uninfected persons, with or without DC (data not shown). Furthermore, using purified CD8+ T cells as responders, we confirmed that the broad reactivity induced by the peptide-loaded DC was mediated by CD8+ T cells, as there was a correlation between the total response of the purified CD8+ T cells and the PBMC to the HIV-1 peptide pools   for the peptides in the absence of DC, and 1.57  in the presence of DC.We first examined the effects of DC loaded with a library of overlapping 15mer peptides spanning the HIV-1 proteome arranged into 29 pools of 19–32 peptides each on T cell reactivity in HIV-1 infected persons on ART. We have previously shown that CD40L-matured DC loaded with pools of ≤32 HIV-1 peptides are optimal for stimulation of CD8P<0.001) . This en+ T cells in the PBMC cultures. These immune responses were of significantly greater magnitude and breadth across the whole HIV-1 proteome (except Vpu) compared to those induced by conventional stimulation with peptides without DC in HIV-1 infected persons on ART.These results indicate that the IFNγ production in response to DC loaded with HIV-1 15mer peptides was produced mainly by CD8+ T cell counts or viral load in the 7 HIV-1 infected subjects (data not shown).We next focused on HIV-1 specific IFNγ production in response to CD40L-treated DC that were loaded with single peptides spanning 49 consecutive HIV-1 Nef 15mers overlapping by 11 amino acids in HIV-1 infected persons on ART. As expected, the number of positive responses to Nef varied among these genetically disparate study subjects . Of the Based on MHC class I alleles of the 7 subjects, we next determined the known and potential Nef epitopes associated with the T cell responses. There were 19 known Nef epitopes matched for the MHC class I alleles of the 7 subjects within the 42 Nef 15mers that induced positive responses . Of thesTaken together, these results show that DC from HIV-1 infected subjects on ART can process 15mer peptides for stimulation of responses against a broad range of known and potential Nef epitope in greater magnitude and breadth compared to that stimulated by these Nef peptides without DC.73–87 (QVPLRPMTYKAAVDL) peptide. We found that stimulation with peptide-loaded DC, but not with peptide without DC, resulted in positive IFN-γ responses in PBMC from 3 HIV-1 infected subjects who shared HLA B*2703  stimulated the highest levels of IFNγ compared to N and C terminal extended and truncated peptides, in a concentration-dependent manner  detected by the ELISPOT assay, i.e., Nef69–83, Nef73–87 and Nef77–91, induced the greatest polyfunctional CD8+ T cell responses when presented without DC (P<0.01 compared to N flanking Nef65–79) . With DC, the Nef69–83 and Nef73–8715mers induced the greatest polyfunctional CD8+ T cell responses (P<0.05 compared to Nef65–79 and Nef77–91)  (+ T cell response to DC loaded with each of the 5 8-10mer variants (P =  ns)  , no DC. ef77–91) , DC. Amo = 0.07) , no DC. P =  ns) , DC. Mos76–84, by a conventional ELISPOT in individuals on ART that was unrecognized using direct stimulation of PBMC with peptide alone. Monofunctional and polyfunctional ICS responses supported that these overlapping Nef 15mers contained a dominant CD8+ T cell epitope. However, a range of less definitive, CD8+ T cell activity was noted against Nef76–84 as well as to the N and C terminal variants, with and without DC.These results show that DC revealed a new, HLA B*2703 epitope, NefWe next determined HIV-1 specific IFNγ production in response to CD40L-treated DC that were loaded with single peptides spanning HIV-1 Gag (122 consecutive 15mers overlapping by 11 amino acids) in HIV-1 infected persons on ART. Overall, there were responses to 80 of the 122 peptides by PBMC from the 7 subjects presented with or without DC . SubjectOf the total of 854 possible responses to the 122 Gag peptides among the 7 subjects, there were 114 (13.3%) responses to DC loaded with Gag peptides compared to only 14 (1.6%) responses to peptides without DC (P<0.001). There were 24 (2.8%) common responses to peptides with and without DC. Higher magnitude responses were observed to DC loaded with peptides compared to peptides without DC (P<0.001). DC enhanced the number of Gag peptide-responding T cells by an average of 8.5 fold (P<0.001).+ T cell counts or viral load in the 7 infected subjects (data not shown). Repeat testing of different blood samples from 2 of these subjects resulted in IFNγ production comparable to the previous responses . IFNγ production was HIV-1 immune specific, as there was little or no IFNγ production induced by the Gag peptides with or without DC in 4 HIV-1 negative controls (data not shown).Enhanced T cell responses to Gag were not associated with CD45–31, p1773–103, p245–27, p2457–95, p24117–179 and p24205–227. The breadth of responses varied across subjects  that elicited positive IFNγ responses with DC in 4 of 7 subjects, and its C terminal overlapping peptide p1721–35 (LRPGGKKKYKLKHIV), which was positive in 2 of 7 subjects using DC . Analysis using the HLA Binding Motif Scanner 23–31 (PGGKKKYKL) was a potential new epitope for HLA B*5101. ELISPOT results for the predicted epitope showed that the 10mer sequence p1722–31 (RPGGKKKYKL) stimulated IFN-γ production without DC in a concentration-dependent manner      : no DC. o no DC) : DC. Of P =  ns) : no DC. o no DC) : DC.22–31 as a novel, optimal 10mer epitope restricted by HLA B*5101. Polyfunctional CD8+ T cell responses induced by the 15mer peptide-loaded DC supported this peptide as the optimal epitope, but were less discriminatory among the 8-10mer, N and C terminal variants presented with or without DC.Thus, using DC and single cell IFN-γ production, we identified Gag p17161–175 (FRDYVDRFYKTLRAE), 5/7 to p24165–179 (VDRFYKTLRAEQASQ) and 4/7 to p24169–183 (YKTLRAEQASQEVKN), whereas there were no responses to either the N or C terminal flanking 15mers (161–169 (FRDYVDRFY) was a potential new epitope restricted by HLA A*0101, and p24173–181 (RAEQASQEV) for HLA B*5101.A cluster of positive responses resulted from stimulation with peptides 74–76 from the p24 region of Gag, with 4/7 subjects responding to p24g 15mers . 12 of 1161–169 showed that there was little or no IFN-γ production induced at any peptide concentration without DC in subject 2 , one aa longer than the predicted, optimal HLA B*5101 epitope p24173–181  (+ T cells by p24165–179 (P = 0.01 compared to p24161–175) and p24169–183 (P = 0.07 compared to p24161–175)  , no DC, 161–175) , DC.161–169 (FRDYVDRFY) peptide and its 8-10mer peptide variants showed no distinct, polyfunctional CD8+ T cell responses (P =  ns)  , no DC. P =  ns) , no DC. P =  ns) , DC.+ T cell responses were induced by the 5 peptides without DC derived from the putative optimal 9mer p24173–181 peptide, with no distinct response to a particular peptide (P =  ns)  , No DC. P =  ns) , DC. In 161–169, and a novel HLA B*5101 10mer epitope p24173–182. N and C terminal variants of these optimal peptides were able to induce appreciable levels of IFN-γ detected by ELISPOT assay only when using DC as APC. There was no clear immunodominance of these 2 Gag p24 epitopes compared to their 8-10mer variants detected by monofunctional or polyfunctional CD8+ T cell responses when presented with or without DC.Taken together, our data indicate that within this cluster of 3 overlapping Gag p24 15mers, stimulation with peptide-loaded DC revealed a novel HLA A*0101 9mer epitope p24+ T memory responses, particularly against Gag + T cells + T cells to non-HIV-1 viral infections + T cell reactivity in persons on ART. The IFNγ response induced by peptide loaded DC was mediated by CD8+ T cells, with purified CD8+ T cells exhibiting an enhanced magnitude and breadth of IFNγ responses relative to PBMC. The 15mer peptides targeted by CD8+ T cells were similar to those targeted by PBMC, but included a broader array of peptides across the proteome of HIV-1. CD8+ T cell responses were noted to DC loaded with 15mer peptides within all 9 HIV-1 proteins except Vpu. Confirming evidence that CD8+ T cells were the predominant responders to HIV-1 peptides was induction of polyfunctional immune mediator reactivity in CD8+ T cells to both Gag and Nef peptides.Control of HIV-1 infection has been related to the magnitude and breadth of HIV-1 CD8Focusing on Nef and Gag specificities, we found that T cells responded to clusters of overlapping 15mer Nef and Gag peptides that contained known, immunodominant epitopes matched to the subjects' MHC class I alleles. Notably, T cell reactivity was induced to 22% of these overlapping 49 Nef peptides by DC compared to only 4% of Nef peptides without DC. Furthermore, the magnitude of the anti-Nef responses was significantly greater for peptides with DC compared to peptides without DC. Similar to Nef, positive T cell responses to Gag 15mers containing known MHC class I epitopes were clustered in well documented, immunodominant regions of the protein that matched the MHC class I alleles of our study subjects. DC revealed T cell responses to 16% of these overlapping 122 Gag peptides compared to only 4% of Gag peptides without DC. The magnitude of the anti-Gag responses was also significantly greater for Gag peptides presented by DC compared to peptides without DC.+ T cell responses to immunodominant epitopes of Nef and Gag has important implications for assessment of T cell immunity in HIV-1 infection. Much of this research relies heavily on use of overlapping 15mer peptides in PBMC without taking into account the role of professional APC + T cell reactivity + T cells were the predominant responders in our study. Our finding implies that the conventional assessment of single cell, IFN-γ responses to libraries of overlapping HIV-1 peptides without DC is potentially missing a significant number of epitopes in persons on ART.That autologous, mature DC significantly enhance the breadth and magnitude of CD8We noted reactivity to 15mer peptides with no previously reported epitope matched for the study subjects' MHC class I alleles for 52% of the reactive Nef peptides and 55% of the reactive Gag peptides. Most of this reactivity was in response to peptides with DC, i.e., T cells specific for 86% of the reactive Nef peptides and 75% of the reactive Gag peptides were only activated by peptide-loaded DC. Analysis of these peptide sequences by predicted binding to their MHC class I motif 73–87 with DC, and the predicted binding motif of HLA B*2703, we defined a novel HLA B*2703 epitope, Nef76–84 (LRPMTYKAA). This new HLA B*2703 Nef epitope is found in 34% of circulating HIV-1 subtype B sequences, while the most common peptide is LRPMTYKgA found in 42% of circulating sequences (based on a dataset of 1184 subtype sequences). This is a variable peptide region since 77 additional peptides were identified in the 1184 circulating sequences but 73 of them were found in less than 1% of sequences. Similarly, using DC we found 3 novel epitopes within Gag that would have been missed using peptide stimulation alone. The 10mer HLA B*5101 epitope for p17  is found in 24% of circulating sequences, while RPGGKKKYrL is found in 27% of sequences. The Gag p24161–169 peptide RAEQASQEVK (HLA A*0101 restricted) is the consensus in 49% of circulating HIV-1 sequences, whereas p24173–182 FRDYVDRFY (HLA B*5101 restricted) is extremely conserved, found in 99.5% of HIV-1 sequences.An extensive analysis of a subset of these putative Nef and Gag epitopes indicated that DC indeed revealed novel epitopes for Nef and Gag. Based on single cell IFN-γ production induced by the 15mer Nef+ T cell reactivity for CD107a, IFNγ, IL-2, MIP-1β, and TNFα, and various polyfunctional combinations of these immune mediators. In particular, the novel Gag peptides presented by DC induced greater levels of polyfunctional CD8+ T cells than peptides without DC. However, with or without DC, polyfunctional T cell responses were less discriminatory for optimal epitope specificity than single cell production of IFN-γ by ELISPOT assay. We have recently noted a similar, limited discrimination by polyfunctional analysis of novel T cell epitopes of human herpesvirus 8 presented by DC We observed that the 15mer overlapping peptides encompassing the 4 novel Nef and Gag epitopes, as well as the 8-10mer epitope variants within these 15mers, induced monofunctional CD8+ T cell responses, the majority of immunodominance is based on the affinity of the peptide for its MHC class I allele on APC, forming a stable number of complexes to activate naive T cells + T cells. However, it is not clear how closely measures of binding of soluble peptides to MHC class I molecules by in vitro affinity assays reflect peptide-MHC binding in DC. Moreover, an increased antigen storage capacity of DC has been linked to their ability to activate T cells by facilitating a continuous supply of MHC class I ligands + T cells There are many factors of mature DC that could relate to their enhancing afferent T cell responses to HIV-1 epitopes. In primary CD8+ T cells by DC is also related to the relative expression of T cell receptor and CD8 molecules on memory T cells Activation of CD8+ T cell responses to HIV-1 epitopes in subjects who have suppressed HIV-1 infection on ART may be misleading. Clearly, the present results indicate that mature DC reveal a broad spectrum of T cell epitopes recognized by CD8+ T cells in persons on ART with suppressed viral load that are not detectable by conventional stimulation of PBMC with peptide alone. These include novel, MHC class I restricted, HIV-1 epitopes that induce monofunctional and polyfunctional T cells producing up to 5 immune mediators that have been linked to control of HIV-1. This suggests that DC could be potent inducers of anti-HIV-1 T cell immunity as an immunotherapy for HIV-1 infected persons on ART.In conclusion, our findings have important implications for T cell immune control of HIV-1 infection. Previously reported, low memory recall, CD8Table S1Characteristics of the HIV-1 infected subjects.(0.21 MB TIF)Click here for additional data file.Table S2MHC class I Nef epitopes detected in 7 HIV-1 infected subjects.(0.03 MB TIF)Click here for additional data file.Table S3MHC class I Gag epitopes detected in 7 HIV-1 infected subjects.(0.03 MB DOCX)Click here for additional data file.


Cryptomeria japonica being pharmacologically active substances. We investigated whether M1 and M4, end products of steroidal ginseng saponins metabolized in digestive tracts, as well as T-cadinol and calamenene can drive DC maturation from human monocytes in vitro. Human monocytes were cultured with GM-CSF and IL-4 for 6 days under standard conditions, followed by another 2 days in the presence of M1, M4, T-cadinol or calamenene. The expression levels of CD1a, CD80, CD83, CD86 and HLA-DR on M1-primed DC, M4-primed DC, T-cadinol-primed DC and calamenene-primed DC were enhanced with a concomitant decrease in endocytic activity. M1-primed DC, M4-primed DC, T-cadinol-primed DC or calamenene-primed DC enhanced the T cell stimulatory capacity in an allo MLR . Naïve T cells co-cultured with allogeneic M1-primed DC, M4-primed DC, T-cadinol-primed DC or calamenene-primed DC turned into typical Th1 cells, which produced large quantities of IFN-γ and released small amounts of IL-4 depending on IL-12 secretion. In the CTL assay (cytotoxic T-lymphocyte assay), the production of IFN-γ and 51Cr release on M4-primed DC was more augmented than of immature DC or TNF-α-primed DC. These results suggest that M1, M4, T-cadinol and calamenene appear to be a good factor to induce DC maturation, or even better in some respect, for the use in clinical DC therapy to induce strong Th1 type immune responses.Dendritic cells (DC) play a pivotal role in the initiation of T-cell-mediated immune responses, making them an attractive cellular adjuvant for use in cancer vaccines. The interaction of T cells with DC is crucial for directing T cell differentiation towards the Th1, Th2 or Th17 type, and several factors determining the direction of the T cell polarization. IL-12 plays a central role in the immune system, not only by augmenting the cytotoxic activity of T cells and NK cells and regulating IFN-γ production, but also by the capacity of IL-12 to promote the development of Th1 cells. Therefore, it is important to identify factors that might affect the differentiation, maturation and function of DC. Ginseng is a medicinal herb widely used in Asian countries, and many of its pharmacological actions are attributed to the ginsenosides. Moreover, T-cadinol and calamenene are sesquterpenes isolated from the heartwood of  The purity of isolated cells was >95% of CD4+ naïve T cells as determined by flow cytometry using FACSCalibur. Allogeneic CD4+ naïve T cells (5 × 105 cell/200 μl) at 1:5 DC/T cell ratio in 96 well U-bottomed tissue culture plates . Some cultures were supplemented with neutralizing Abs to block endogenous cytokines: anti-IL-12 . On day 5, cells were washed extensively and expanded with fresh medium containing 10 U/ml of recombinant human IL-2 (IL-2) . One hundred microliters of culture supernatant was replaced with medium of the same concentration every 3 days. On day 14, cells were washed, counted, and T cells (106/ml) were re-stimulated for 24 h on plates coated with anti-CD3  and anti-CD28 . The cell-free supernatants were collected and frozen at −20 °C until measurement of cytokines using ELISA.Irradiated (30 Gy) DC were co-cultured with naïve T cells (2.5 × 106/ml) were stimulated with phorbol myristate acetate (PMA) (10 ng/ml) and ionomycin (1 μg/ml) for 5 h at 37 °C in a water bath. Brefeldin A (10 μg/ml) was added during the last 2 h of incubation to prevent cytokine secretion. Cells were collected, fixed with 1% paraformaldehyde, permeabilized with a commercial solution (BD-Pharmingen), and stained with FITC-labelled anti-IFN-γ (IgG2a) and PE-labelled anti-IL-4 (IgG1) mAbs. The samples were analyzed on a FACS-Calibur with CellQuest software. Ten thousand cells were analyzed per sample.The intracellular cytokine concentrations of the harvested T cells were measured by FACS analysis as previously described . Briefly5/100 μl) were then added to the upper chambers of the Transwell plates. Transwell cultures were maintained for 5 h in 5% CO2 at 37 °C, and DC migrated from the upper wells were counted using a Colter Counter. The results were expressed as net migration percent calculated as:/total number of cells loaded in the upper chamber × 100.Migration of DC induced by macrophage inflammatory protein (MIP)-3β was measured using a double chamber system as previously described . Briefly2+ ([Ca2+]i) concentration was carried out as previously described i was monitored with a CAF110 spectrofluorimeter .The determination of intracellular Caescribed . Briefly6 DC using an RNeasy Mini Kit . The cDNA synthesized from total RNA by Moloney murin leukaemia virus reverse transcriptase  was subjected to RT-PCR using 30 cycles at 94 °C for 0.5 min, 57.5 °C for 1 min, and 72 °C for 1 min for CCR7 . The sense and anti-sense oligonucleotide primers for CCR7 were 5′-CGCGTCCTTCTCATCAGCAA-3′ and 5′-GTCCCGACAGGAAGACCACT-3′, respectively. PCR products were identified by electrophoresis on 2% agarose gels that were photographed.Total RNA was extracted from 1–2 × 105) cultured from monocytes of HLA-A24+ donors were loaded with 10 μM Epstein-Barr virus (EBV)-derived peptide (TYGPVFMCL) for 2 h at 37°C in 5% CO2. EBV-derived peptide can bind to HLA-A2402 as reported by Lee et al. [6 cells/ml with 10μM EBV peptide and incubated for 2 h at 37 °C in 5% CO2. Effector cells were co-cultured with target cells at effector-to-target ratios of 2:1, 10:1 and 20:1 in 96 well round-bottomed culture plates  in duplicate. After overnight incubation at 37 °C in 5% CO2, IFN-γ-concentration of the supernatant was determined by ELISA.Details of the method used in this assay have been previously described . Brieflye et al. . In 24 w51Cr labelled target cells (1 × 104) at various effecor-to-target cell ratios for 6 h, followed by supernatant harvesting for measuring radioactivity using as automatic γ counter. This assay was done in triplicate.Details of the methods used in this assay have been previously described . BrieflyStatistical analysis of the results was performed by ANOVA. Differences were considered statistically significant when p value were less than 0.05.To study the direct effects of M1, M4, T-cadinol and calamenene on the maturation of sentinel DC into effector DC, human monocytes-derived DC were cultured with M1, M4, T-cadinol or calamenene. Human monocytes were cultured with GM-CSF and IL-4 for 6 days under standard conditions, followed by another 2 days in the presence of various concentrations of M1, M4, T-cadinol or calamenene. LPS, CT and TNF-α were used as a positive control. The resulting populations of DC were analyzed by flow cytometry. As shown in Because the level of IL-12 production by DC is a major factor driving the development of Th1 cells, we studied the influence of M1, M4, T-cadinol or calamenene on IL-12 production by DC. We measured IL-12p70 production in immature DC and in DC matured for 2 days in the presence of the above factors after stimulation by CD40-L for 24 h. The production of IL-12p70 by M1-primed DC, M4-primed DC, T-cadinol-primed DC and calamenene-primed DC was more augmented than that of LPS-primed DC . In contWe observed that M1-primed DC, M4-primed DC, T-cadinol-primed DC and calamenene-primed DC expressed increased levels of Ag-presenting and the expression levels of CD1a, CD80, CD83, CD86 and HLA-DR as expressed by MFI. Therefore, we next compared the capacity of M1-primed DC, M4-primed DC, T-cadinol-primed DC, calamenene-primed DC, LPS-primed DC, CT-primed DC or TNF-α-primed DC to stimulate T cells in an allo MLR. M4-primed DC showed higher stimulatory efficiency in an allo MLR than LPS-primed DC , CT-primWe next evaluated the nature of primary allogeneic T cell responses stimulated by M1-primed DC, M4-primed DC, T-cadinol-primed DC and calamenene-primed DC. Allogeneic M1-primed DC, M4-primed DC, T-cadinol-primed DC or calamenene-primed DC co-cultured with naïve T cells at 1:5 DC/T cell ratio secreted sizeable amounts of IFN-γ , but litin vitro. T-cadinol-primed DC and calamenene-primed DC had a higher migration response to MIP-3β than LPS-primed DC (2+ mobilization to MIP-3β (2+ mobilization to MIP-3β were observed with CT-primed DC and TNF-α-primed DC (data not shown). On the other hand, migration response and intracellular Ca2+ mobilization to MIP-3β in immature DC were low or just detectable  was used as a target cell (negative control)  .in vitro. Here, we demonstrate that culture of immature DC with M1, M4, T-cadinol or calamenene increase cell surface expression of CD1a, CD80, CD83, CD86 and HLA-DR, while decreasing endocytic activity, resulted in cells with a phenotype characterized by efficient Ag-presentation and costimulatory capacity of mature DC. Functionally, M1-primed DC, M4-primed DC, T-cadinol-primed DC or calamenene-primed DC have enhanced primary allogeneic T cell stimulatory activity in an allo MLR.The present study was performed in order to investigate whether M1 and M4, end products of steroidal ginseng saponins metabolized in digestive tracts, as well as T-cadinol and calamenene can change the phenotype and function associated with DC differentiation from human monocytes in vivo anti-metastatic effect by oral administration of ginsenosides may be primarily meditated by their metabolic component M1 and M4. Moreover, Scaglione et al. [The differentiation of monocytes into DC has a critical impact on the immune response. DC interaction with naïve T cells plays a key role in primary immune responses , 3 and te et al.  have inve et al. . The core et al. . M1-primin vitro is very likely due to the lack of IL-12 production by CT-treated DC and may be independent of IL-4. We showed that the production of IL-12 by CT-primed DC that were stimulated CD40-L was low or detectable. Therefore, it suggests that the involvement of these cells in the innate responses to Th1 response-inducing pathogens during the period preceding the initiation of adaptive immunity can create an environment rich in IL-12, which promotes the production of IFN-γ and the outgrowth of Th1 cells. It has been demonstrated that immune balance controlled by cytokines such as IL-10 and IL-12 plays an important role in immune regulation, including anti-tumor immunity [In contrast, CT-primed DC induced naïve T cells showed a shift towards Th2 effector cell. CT is a powerful mucosal adjuvant that amplifies B cell and T cell responses to mucosally co-administered antigens, stimulating predominant Th2-type responses , 46. Gagimmunity . Our resin vivo. In most clinical trials using DC-based immunotherapy, immature monocytes-derived DC pulsed with tumor antigen peptides were used. However, recent observation suggests that mature DC could be a better anti-tumor adjuvant [One result of DC maturation and activation demonstrate increased motility and a potential to propensity to migrate towards lymphoid organs. During the maturation process, DC down-regulate the expression of inflammatory chemokines and their receptors and up-regulate the synthesis of constitutive chemokines and the chemokine receptor CCR7. We found that M1-primed DC, M4-primed DC, T-cadinol-primed DC and calamenene-primed DC had high migration, high calcium mobilization in response to MIP-3β and up-regulated the expression of CCR7, suggesting that M1-primed DC, M4-primed DC, T-cadinol-primed DC and calamenene-primed DC are likely to be mature DC that have the potential to migrate adjuvant . Althougin vivo.Antitumor immunity has classically been measured by the quantity of tumor-antigen-specific CD8+ T cells . Immunotin vitro. Moreover, DC differentiated with M1, M4, T-cadinol or calamenene enhance the differentiation of naïve T cells towards the Th1 type depending on IL-12 secretion. M-4-primed DC produces a stronger CTL response. Although the molecular events leading to the effects of M1, M4, T-cadinol and calamenene on DC function remain to be resolved, DC appear to be potential targets of M1, M4, T-cadinol and calamenene. Further understanding of the mechanisms by which M1, M4, T-cadinol and calamenene modulate DC function may lead to the development of effective immunotherapy for cancer.This is first study on the effect of terpenoids, e.g. ginseng saponins on human monocytes-derived DC giving new insights into the action of terpenoids. M1, M4, T-cadinol and calamenene potentially regulate differentiation of DC from human monocytes in combination with GM-CSF and IL-4


Social factors may be of importance causally and act as maintenance factors in patients with anorexia nervosa. Oxytocin is a neuromodulatory hormone involved in social emotional processing associated with attentional processes. This study aimed to examine the impact of oxytocin on attentional processes to social faces representing anger, disgust, and happiness in patients with anorexia nervosa.A double-blind, placebo-controlled within-subject crossover design was used. Intranasal oxytocin or placebo followed by a visual probe detection task with faces depicting anger, disgust, and happiness was administered to 64 female subjects: 31 patients with anorexia nervosa and 33 control students.Attentional bias to the disgust stimuli was observed in both groups under the placebo condition. The attentional bias to disgust was reduced under the oxytocin condition (a moderate effect in the patient group). Avoidance of angry faces was observed in the patient group under the placebo condition and vigilance was observed in the healthy comparison group; both of these information processing responses were moderated by oxytocin producing an increase in vigilance in the patients. Happy/smiling faces did not elicit an attentional response in controls or the patients under either the placebo or oxytocin conditions.Oxytocin attenuated attentional vigilance to disgust in patients with anorexia nervosa and healthy controls. On the other hand, oxytocin changed the response to angry faces from avoidance to vigilance in patients but reduced vigilance to anger in healthy controls. We conclude that patients with anorexia nervosa appear to use different strategies/circuits to emotionally process anger from their healthy counterparts. Anorexia nervosa (AN) is characterized by problems with eating, weight, and shape. Shape and weight concerns often involve social contagion and social evaluation The social processing problems in patients with eating disorders have been synthesized in recent systematic reviews, and impairments in emotional recognition, social hierarchy, and theory of mind have been found Oxytocin is an important neuromodulatory hormone involved in social emotional processing, particularly in association with attentional processes. Oxytocin increases the sensitivity to detect emotional expressions, which is associated with recruitment of attentional resources, as evidenced by pupil dilation It is possible that changes in oxytocin function may account for some of the anomalies in social cognition in patients with AN. A recent review synthesized evidence for abnormal oxytocin functioning in patients with AN The aim of this study was to examine whether oxytocin impacts on social attentional processes in patients with AN. Our first hypothesis, based on previous studies using the dot probe paradigm, is that patients with AN would have vigilance towards threatening cues and away from rewarding stimuli. Our second hypothesis is that oxytocin would attenuates the selective attention towards negative social cues.Sixty-four women  took part in this double-blind, placebo-controlled cross-over study. The first participant entered the study on August 16, 2012, and the last participant was examined on November 30, 2013. The patients with AN were recruited at the Eating Disorders Clinic of Seoul Paik Hospital, Seoul, South Korea. As this study aimed to examine broadly whether oxytocin might be of benefit in the short term for patients with AN, patients were recruited at various stages and phase of illness both from an outpatient clinic (n = 13) and an inpatient ward (n = 18). All patients who volunteered to participate were screened during the early phase of treatment. All of them were in an active stage of illness, and none of them were in the weight-recovered stage. The diagnosis of AN was confirmed by the Structured Clinical Interview from the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition The Korean version of the Wechsler Adult Intelligence Scale was used to measure IQ. Healthy participants were tested during the follicular phase of their menstrual cycle. None of the patients were menstruating. Compensation was provided for travelling expenses and time. This study protocol was approved by both the Korean Food and Drug Association Institutional Review Board (12061) and the Institutional Review Board of Seoul Paik Hospital (IIT-2012-096). All participants provided written informed consent prior to participating in the study. Consent was provided by both the patients and their guardians in the case of 16-year-old patients.JW Pharmaceuticals  from oxytocin powder . The solution was prepared by combining 35.2 mg oxytocin (568 U) with 300 mL 0.9% sodium chloride solution, and adjusting the pH to 4.01 with 10× diluted acetic acid. The placebo spray (pH 4.01) was formulated with 0.9% sodium chloride solution and acetic acid, but without the peptide. The filtered and sterilized solution was sealed in individual vials (1.5 mL each) and stored frozen. On the day of use, the vials were thawed and kept in a refrigerator (4°C) until required. A clinician prepared the nasal spray by transferring oxytocin or the placebo from the vial into a nebulizer.The intranasal oxytocin spray was formulated by Oxytocin and the placebo were administered intranasally 4–7 days apart 45 min before the neuropsychological tasks. We chose a dose of 40 IU of oxytocin based on a recent review reporting that cumulative evidence from clinical trials shows that short-term use of intranasal oxytocin administered to both male and female humans in dosages up to 40 IU (per dose) results in no detectable subjective adverse effects in a controlled research setting The photographic stimuli used for the dot probe task were obtained from a validated collection of the Korean Facial Expressions of Emotions (KOFEE) Our pool of target stimuli consisted of 60 images of adults showing 15 positive (happy) or 30 negative (15 angry and 15 disgust) expressions, with each adult having a corresponding picture showing a neutral expression (15 non-target stimuli). The target stimuli were paired with matched non-target stimuli.Attentional biases were assessed using a visual probe detection task As the two images disappeared from the screen, one of two probes (‘:’ or ‘..’) appeared where used to be the center of one of the two pictures. The participants were required to press one of two buttons on a keyboard (marked with white stickers) that corresponds to the probe (Q for ‘:’ and Z for ‘..’). When a response was made, the probe disappeared and the next trial started. Participants were advised to identify the probe as quickly and accurately as possible. The target stimuli positions and the probe positions were balanced across trials, so each appeared in either location  with equal frequency, following the paradigm of Mogg and Bradley Attention during the visual probe detection task was measured as the time to respond to the probe. The assumption is that the time to respond to the probe is faster if attention was already allocated to the spatial location where the probe appeared. Faster responses to congruent trials (when the probe emerges at the threatening stimulus location) compared to incongruent trials  were interpreted as vigilance to a threat.These tests were comprised of 10 trials in each of six conditions: three target stimuli  and two probe locations (the location of the target stimuli or non-target stimuli), for a total of 60 trials, repeated twice. Test trials were presented in a new random order for each participant. The dot-probe task was presented on E-prime ver. 2 .The Korean version of the clinician rating interview of the EDE The EDE-Q assesses the main behavioral features of an eating disorder over the past 28 days. The questionnaire consists of 36 items on a 7-point forced choice rating scale. It measures weight, shape, eating concerns, and dietary restraint. We used the standardized Korean version of the 12th edition of the EDE-Q, which has high internal consistency and good 2 week reliability.The STAXI is a 44-item self-report questionnaire that is divided into seven scales. It measures the current intensity of anger (State Anger) and disposition toward anger as a personality trait (Trait Anger) and the tendency to express anger when criticized (Angry Reaction). The other scales measure the frequency with which provoked anger is suppressed (Anger-Suppression), the frequency of the expression of anger toward other people or objects (Anger-Expression), and the frequency at which the expression of anger is controlled (Anger-Control). The Korean version of the STAXI was used in this study The AQ is a 50-item, self-administered questionnaire that assesses the degree to which adults with normal intelligence have autism spectrum traits. It has five sub-scales, including social skill, attention switching, attention to detail, communication, and imagination.Depression and anxiety were assessed in each subject using the standardized Korean versions of the Beck Depression Inventory (BDI) The participants were tested in a private room at Seoul Paik Hospital. They were instructed to abstain from alcohol and caffeine on the day of drug administration and food and drink (other than water) for 2 hours before drug administration. Upon arrival, the participants completed baseline measures of physical symptoms, including abdominal, neurological, dermatological, and cardiac symptoms. Subsequently, oxytocin or the placebo was self-administered by inhalation of the spray into each nostril. The order of oxytocin and placebo administration was randomized using Microsoft Excel in this double-blind procedure. The participants completed the self-report measures to assess their psychological state during the 45 minutes after administration of each spray. At the end of the study period, each participant completed follow-up measures for adverse physical symptoms.We followed the standard analysis procedure used for this task Attentional bias scores were calculated for each matched trial type  as PN − PE, where PN is the mean RT for probes replacing neutral stimuli, and PE is the mean RT for probes replacing emotional target stimuli. Therefore, positive scores suggest increased attention towards the emotional/target stimuli, whereas negative scores indicate attention away from emotional/target stimuli. A score of 0 suggests no bias towards any stimuli.t-test comparisons for each group. Greenhouse–Geisser corrections were applied on the assumption that sphericity was violated. Data are shown as mean and with the effect size (ES) if appropriate . The estimation of ES was based on Cohen's d: described as negligible ( = 0 and <0.15), small (≥0.15 and <0.40), medium (≥0.40 and <0.75), large (≥0.75 and <1.10), very large (≥1.10 and <1.45) and huge (>1.45) . We defined the drug effect as the difference in the RT score between the administration of the placebo and oxytocin (drug effect  =  RTplacebo − RToxytocin). Pearson's correlations were used to investigate the relationship between the drug effect on attentional bias and psychological variables. Analyses were performed using SPSS 19.0 , and the two-tailed p-value was 0.05.The attentional responses to shape stimuli were investigated via a series of initial 2 (group: AN and HC) ×2 (drug: oxytocin and placebo) ×3  repeated-measures analyses of variance (ANOVAs). Post-hoc analyses were conducted through paired 2) and the EDE-Q scales. The AN group also scored significantly higher on the BDI, STAI, and the Social Skill, Attention, and Communication subscales and the total score on the AQ compared to those in the HC group. The EDE interview scores of the AN group were 5.07±6.04 for the restraint subscale, 3.73±4.41 for the eating concern subscale, 4.35±8.33 for the weight concern subscale, and 3.85±3.10 for the shape concern subscale.The demographic and the group characteristics of the participants are shown in F  = 3.568, p = 0.031, Δ2η  = 0.057) was observed in the three-way group (AN vs. HC) × drug (oxytocin vs. placebo) × emotion  repeated-measures ANOVA.A significant group x drug x emotion interaction effect   = 4.988, p = 0.029, Δ2η  = 0.078).The two-way group (AN vs. HC) × drug (oxytocin vs. placebo) repeated-measures ANOVA identified a group × drug interaction effect   = 0.028, p = 0.868, Δ2η  = 0.000) nor a group × drug interaction effect in response to happy stimuli   = 0.009, p = 0.925, Δ2η  = 0.000).The two-way group (AN vs. HC) × drug (oxytocin vs. placebo) repeated-measures ANOVA showed that there was neither a main effect of drug   = 8.970, p = 0.004, Δ2η  = 0.130) in response to the angry stimuli. Post-hoc tests showed avoidance of angry faces in the AN group and vigilance in the HC group with a moderate to large difference between the AN and HC groups under the placebo condition (t(62)  = 2.510, p = 0.015, d = −0.633). Both of these information processing responses were moderated by oxytocin and produced a large increase in vigilance in the AN group (t(30)  = −2.847, p = 0.008, d = 0.785) but a tendency to decrease with a small ES in attention in the HC group (t(32)  = 1.218, p = 0.233, d = 0.219).The two-way group (AN vs. HC) × drug (oxytocin vs. placebo) repeated-measures ANOVA showed that there was a group × drug interaction effect   = 10.120, p = 0.002, Δ2η  = 0.142) with no group × drug interaction effect   = 0.375, p = 0.542, Δη2  = 0.006) in response to the disgust stimuli. Post-hoc tests showed an attentional bias to the disgust stimuli in both the AN and HC groups under the placebo condition (t(62)  = 3.208, p = 0.002, d = 0.597). Attentional bias was reduced under the oxytocin condition in both groups with a small effect in the AN group (t(30)  = 1.602, p = 0.120, d = 0.418) and a moderate/large effect in the HC group (t(32)  = 3.138, p = 0.004, d = 0.555). The attentional bias scores across conditions in each group are shown in The two-way group (AN vs. HC) × drug (oxytocin vs. placebo) repeated-measures ANOVA showed a main effect of drug  i.e., individuals with higher anger expression showed a greater reduction in vigilance with oxytocin. There was no association between STAXI subscales and the oxytocin effect on angry stimuli in patients with AN. No association was observed between the drug effect and aspects of psychopathology typically associated with eating disorders (EDE-Q), depression (BDI), state and trait anxiety (STAI), autism traits (AQ) in either the AN or HC groups. No association between BMI or weight and the oxytocin effect were observed in either the AN group  or in the HC group .The reduced vigilance to angry stimuli in the HC group was positively correlated with STAXI subscales of anger expression in the oxytocin condition  faces produced no attentional bias in patients with AN or the HC group. Our second hypothesis was confirmed, in part, as oxytocin attenuated the selective attention towards disgust in both the patients with AN and the HC group. In contrast, oxytocin converted an avoidant response to anger into vigilance in patients with AN but reduced the vigilant response to anger in the HC group. Oxytocin did not change the lack of attention to happy (smiling) faces in either the AN or HC groups.Our result that smiling faces did not produce an attentional bias in either the AN or HC groups was consistent with other findings in Caucasian patients with AN In the oxytocin condition, the attentional bias was away from disgusted faces in both groups with a larger effect in the HC group. To our knowledge no previous study has evaluated the impact of oxytocin on recognition of the disgust emotion in humans. The results are consistent with another study, which found that oxytocin decreases attention toward negative faces  in healthy people The avoidance response to anger expression in patients with AN and a vigilant response in the HC group under the placebo condition differed from a study that used “rejecting” facial expressions in which the opposite pattern was found, i.e., vigilance to the rejecting stimuli in patients with AN These findings raise the possibility that oxytocin treatment may moderate some of these social difficulties and possibly improve the outcome. Resistance to treatment is associated with less activation in social cognition circuits within the brain There are some limitations in this study that need to be considered and which may account for some of the individual variation in this study. First, a third of the patients with AN were recruited from the outpatient clinic and the others were from the inpatient ward. So the patients were at different phases of treatment. The oxytocinergic pathway may differ in the more severely affected patients undergoing refeeding, compared to that of outpatients. However, all of our participants were screened when they initially visited to the clinic and were in the active stage of the illness, regardless of their inpatient or the outpatient status. None of them were weight-recovered. This is a proof of concept study which aimed to broadly examine that oxytocin might be of benefit in the short term across different stages. Further studies should consider controlling the illness stages. Second, the doses of intranasal oxytocin were not adjusted for individual differences in BMI and therefore the underweight patients received a higher doses per weight than healthy controls. However, in our subsidiary analysis, no relationship was observed between the BMI or weight and the oxytocin effect in either the AN or healthy control groups. Further studies examining for a dose relationship may be of interest and reduce the variance. Third, three patients took fluoxetine, which may affect the oxytocin pathway, so may have increased the variance. An additional analysis found no differences in the response of this subgroup to the non medicated patients. Finally, we did not measure other individual factors that are known to impact attention bias. For example, individuals with a short 5-hydroxytryptamine allele have been found to be more vigilant towards threat In conclusion, patients with AN showed an avoidance of angry faces whereas healthy women were vigilant to anger in the placebo condition. This response in the AN group shifted from avoidance to vigilance following oxytocin administration but there was a trend for the healthy women to be less vigilant to angry faces. Patients with AN showed vigilance to faces of disgust similar to the healthy women and this was moderated with oxytocin in both groups reaching a significant effect in healthy women. Neither group had any attentional bias to happy faces under either the oxytocin or control conditions. These findings suggest that further evaluation of oxytocin nasal spray as a treatment to improve emotional processing and social communication particularly in relationship to anger may be of value in patients with AN.


In response to “Megadose Methylprednisolone (MDMP) for Hemangiomatosis” We are grateful to Dr. Özsoylu for his valuable comments on our case report concerning the treatment of hemangiomatosis with propranolol . One of Systemic corticosteroids have been the mainstay of hemangioma treatment since the 1960s. Oral or intravenous MP and prednisone are effective for shrinking hemangiomatous lesions in infants. Cushingoid appearance and delayed healing of the cervical ulcerated lesion in our case were attributed to prolonged use of systemic MP. Recent studies have reported successful treatment of hemangiomas with the β-blocking agent propranolol, even in low birth weight infants ,5. Propr


Despite all the knowledge on depression, it is still unclear whether current literature covers all the psychosocial difficulties (PSDs) important for depressed patients. The aim of the present study was to identify the gaps in the recent literature concerning PSDs and their related variables. Psychosocial difficulties were defined according to the World Health Organization International Classification of Functioning, Disability and Health (ICF). A comparative approach between a systematic literature review, a focus group, and individual interviews with depressed patients was used. Literature reported the main psychosocial difficulties almost fully, but not in the same degree of importance as patients' reports. Furthermore, the covered areas were very general and related to symptomatology. Regarding the related variables, literature focused on clinical variables and treatments above all but did not report that many psychosocial difficulties influence other PSDs. This study identified many existing research gaps in recent literature mainly in the area of related variables of PSDs. Future steps in this direction are needed. Moreover, we suggest that clinicians select interventions covering not only symptoms, but also PSDs and their modifiable related variables. Furthermore, identification of interventions for particular psychosocial difficulties and personalisation of therapies according to individuals' PSDs are necessary. Depression is a major public health issue due to its prevalence, high mortality rates , suicideThis evidence suggests that usual management strategies do not address sufficiently relevant areas of depression. One substantial dimension of depression comprises the psychosocial difficulties (PSDs) which people experience. PSDs constitute the impairment on psychological and social daily functioning of individuals, linked with their particular health condition . The broPrevious scientific literature provides different definitions of psychosocial difficulties in depression but has some limitations: the methods either focus only on specific areas or consider the PSDs as a result of depression. Recent studies –9, howevOnce this comprehensive definition of psychosocial difficulties has been extracted from the literature, it is very important to incorporate the patient perspective in clinical practice and research. The majority of studies on this topic are quantitative; however, the literature that uses qualitative methodologies—which could provide deeper, richer, and more elaborated data, exploratory analyses on patients' needs and standpoints, and thus more objective results—is sparse. Furthermore, this kind of studies can provide a general view if particular psychosocial difficulties, named by patients as important, are missing in the literature.There are few existing studies using qualitative research to identify a full list of PSDs which depressed people experience. Yet, none of them has applied a comprehensive approach in order to encompass all the psychosocial difficulties from different perspectives. An example is a study by Lasch et al. , which aAn article by Brütt et al.  based itTo date, no study has analyzed the full set of PSDs and their related variables in depression by including both literature and patient perspective. The aim of the present study is to tackle this research gap and discover whether the recent scientific literature actually reports the PSDs and their related variables that are important for depressed patients. Our objective is to obtain information about whether the latest literature extensively covers the issues that are pointed out as problematic by depressed individuals, or if there are particular areas, which should receive more attention. Moreover, a potential identification of these missing previously-ignored questions in literature will enhance the quality of future research and enable new strategies for treatment and rehabilitation in depression.In order to compare whether the psychosocial difficulties experienced by depressed patients are actually reported by recent literature in depression, we included a step by step methodology.We gathered information from three different studies.First, a systematic review to directly collect information reported in recent literature, consulting the MEDLINE and PsycINFO databases, was conducted. Search terms were adapted to each database combining the MeSH headings of “Depression,” “Depressive Disorder,” and “Depressive Disorder, Major” with “depress*” (title) in MEDLINE database and (DE=) “Major Depression,” “Recurrent Depression,” and “Depressive disorder” in PsycINFO. For psychosocial difficulties the following keywords were used: “psychosocial*,” exp Quality of life/, exp Personal satisfaction/exp Human activities/exp social support/disabilit*, homelessness, environmental factor*, exp Interpersonal relations/, exp Quality of life/, exp personal satisfaction/, exp human activities/, exp paternalism/, prejudice/, psychosocial deprivation/, social values/, exp Social Problems/, Social Adjustment/, social isolation/stereotyping/, exp Social environment/, exp emotions/, exp family/, exp socioeconomic factors/exp life style/exp Disability evaluation/, exp Communication Barriers/, “Adaptation”, exp Psychological/, exp Aggression/, exp Psychological stress/, exp community /, Sexual* or intimacy. Inclusion criteria were articles reporting information on psychosocial difficulties in people with a diagnosis of major or minor depression according to DSM-III-TR, DSM-IV, or DSM-IV-TR –16, or aOnce all the included studies were selected, we extracted information about the sample characteristics and then collected the variables, the tools, the psychosocial difficulties and their related variables. Related variables in literature refer to determinants of PSDs since only longitudinal studies were included. Determinants in literature were those variables that were longitudinal predictors for incidence or changing of psychosocial difficulties, so consequently a causal relationship with the PSDs can be hypothesized. Extraction of the information was double checked by two independent reviewers in 20% of the articles (MC and BM) (Kappa = 0.80). Kappa's coefficient was calculated according to Fleiss and Cohen rules [ Hospital Universitario de la Princesa in Madrid were invited to participate. Selection was done by their main mental health care provider (psychiatrist) taking into consideration the maximum variability of sampling in gender, work status, and clinical status , consulting the patients' clinical records. Eight patients were invited to participate and one of them did not consent to participate reporting having no time for this. Participation in this study was not mandatory and only patients with motivation to participate were included in the final sample; therefore a selection bias could have affected our results. Participation was formally agreed after signing a consent informed form. One moderator and one assistant (MC and IL), who had been previously trained, encouraged all the members to participate during the session.On the other hand, the collection of patients' perspectives was performed including information from two studies. The first one consisted of one focus group composed of seven depressed patients. We based our sample size decisions on literature recommendations, “The ideal size of a focus group for most noncommercial topics is five to eight participants”  and the Four open questions were posed: (1) which psychosocial difficulties are usually experienced due to participant's depression; (2) which ones are more relevant (ranking the five most important ones for each participant); (3) how these difficulties changed overtime; and (4) which events are responsible for the onset or change of these psychosocial difficulties overtime. All the dialogues were digitally recorded and subsequently transcribed. “I have no time to participate,” one had no interest in research studies, and one showed no interest in this particular study. During the initial interview researchers verified the fulfillment of the inclusion criteria. At that point five patients that were derived from the primary care center were excluded because they met diagnostic criteria for other different disorders: bipolar disorders (2), generalized anxiety disorder (1), substance use disorder (1), and complicated grief (n = 1) and thus satisfied one of the exclusion criteria.Finally, in order to gather other difficulties that are not usually reported during focus group sessions because of potential unwillingness of self-disclosure , we inclPatients were singly asked, among other questions, which were the five most disabling psychosocial difficulties that they experienced due to depression and which were the variables that were responsible for the onset or the change of these problems. A causal relationship with the PSDs and extraction of determinants, unlike in the literature, cannot be established, because of the nature of the focus group and the individual interviews; therefore we will refer to these variables of onset or change of PSDs according to the patient perspective as related variables.Santa Hortensia” Primary Care center of Madrid or in the outpatient psychiatric service at the Hospital Universitario de la Princesa in Madrid. In both study sites patients were chosen according to their availability and were invited to participate by their primary care doctors or psychiatrists. All participants signed an informed consent form. Both studies were independently reviewed and approved by the Hospital Ethics Committee for Clinical Research.Trained interviewers  conducted all the individual interviews. Inclusion criteria for individual interviews were patients older than 18 years with a diagnosis of depressive episode or major depression criteria according to ICD-10. All patients were collected in “After gathering information from the three different studies, we needed to establish a common language necessary to directly compare outcomes. For that purpose a list of common categories for classifying psychosocial difficulties was agreed on during one research group meeting. Participants were researchers who had been involved in data collection for different health conditions. Researchers involved in the data collection of depression  also participated. All of them were requested to share with the group the PSDs that they had identified in the different studies and the terminology they had used to name them. After each naming, the working group was asked whether they agreed with the terminology proposed. After a brief discussion, stating pros and cons for the proposal, an agreed-on term was decided and documented for each PSD. The same procedure was followed to extract the names of related variables in subsequent working group sessions with the same participants. personal factors we used the definition given by the WHO's International Classification of Functioning, Disability and Health: “Personal factorsare the particular background of an individual's life and living, and comprise features of the individual that are not part of a health condition or health states. These factors may include gender, race, age, other health conditions, fitness, lifestyle, habits, upbringing, coping styles, social background, education, profession, past and current experience (past life events and concurrent events), overall behaviour pattern and character style, individual psychological assets and other characteristics, all or any of which may play a role in disability at any level. Personal factors are not classified in ICF.” [After obtaining the list of common categories, we associated the different concepts extracted in the studies to the agreed category list. Forin ICF.” . The likin ICF.” . BM and Focusing on the information extracted from the literature, a frequency analysis was performed regarding how many different studies particularly reported the psychosocial difficulties or related variables. In the case of individual interviews, the number of times that the different participants reported the psychosocial difficulties and their related variables was calculated. Finally, for the focus group, digital recordings were analyzed in order to extract the number of times in which psychosocial difficulties and related variables had been a topic during the session . These analyses were performed with the software for qualitative research NVIVO.Tables Comparison between the literature review and the studies reporting patient's opinion showed that literature does cover almost fully the main psychosocial difficulties directly addressed by depressed individuals. There were relatively few PSDs, not covered by the recent scientific literature. Emotional functions [ energy and drive functioning [ cognitive functions [ employment [ relationship with the others [unctions –26, enerctioning –29, cognunctions –32, emplployment –35, and e others –38 were  Carrying out daily routine was pointed out by ten patients in the individual interviews (making it the fifth most important PSD), but it has been investigated less than three times in the revised literature.However, some main PSDs in the patient reports were not covered enough by the literature.Communication with others was among the most important psychosocial problems, according to the participants in the focus group, but our systematic review did not find it mentioned anywhere in the literature. Moreover, weight maintenance functions and doing housework were highlighted by the patients in the individual interviews, but literature omitted them as significant and important psychosocial difficulties in depression. In addition, there were some main categories, which did not meet the same ranking of significance when the different sources of information were compared. Pain [ sleep [ed. Pain –41 and s [ sleep , 43, forRegardless of these few above mentioned PSDs, literature in general addresses substantially the main psychosocial difficulties in depression. Therefore, we conducted an elaborate second level analysis to investigate whether the specific psychosocial difficulties (components within the main PSD categories), reported in the focus group and individual interviews, were also identified in the literature. Once more, the results demonstrated that the literature, with some exceptions, almost fully covers the range of specific psychosocial difficulties, experienced by depressed patients. Loneliness and distress  were among the most important and commonly mentioned difficulties by the patients, whereas these specific PSDs were reported less than three times in the literature. The same is valid for attention and memory (part of cognitive functions), efficiency (employment), and intimate relationships (relationships with others).However, as can be seen in  treatment [ emotional functions as a cause of psychosocial difficulties as well. Specifically, results showed that the literature did not analyse sufficiently cognitive functions, relationships with others, energy and drive, and employment problems as causes of PSDs.Additionally, we investigated whether literature sufficiently reports the most important related variables of change and onset for psychosocial problems in depression that are addressed by patients. All related variables can be seen in reatment –46 was aThe current study aimed to analyse whether the recent scientific literature reports those psychosocial difficulties and their related variables that are important for depressed patients. Our findings indicated that the literature does report almost fully the main psychosocial difficulties, experienced by patients with depression, but the degree of importance of each PSD depended on the source of information. The same refers to the specific PSDs, extracted by a second level analysis. Contrary to the main PSDs reported, however, only a few related variables of onset of PSDs were currently reported in literature. In addition, literature was specifically focused on clinical variables as related to the PSDs, whereas it ignored that some PSDs can also become related variables for other PSDs.To our knowledge, this is the first study to throw light on whether the latest scientific literature actually reflects the issues that are indicated as problematic by depressed patients. However, all the subsequent comments on the literature have to be considered with respect to the only 103 existing studies within our period of search that covered the whole spectrum of PSDs in depression and the fact that their average quality rate is not high . In this. Emotional functions stood out as an evident problem for depressed patients, as part of the outcomes of 62 research studies and being mentioned more than 100 times during the focus group and individual interviews. A review by Brockow et al. [ depressive mood and symptoms [ loneliness and distress (emphasized notably by depressed patients but narrowly explored in the literature) can be explained through the literature tendency to focus mainly on those PSDs which belong to the symptomatology spectrum. In addition, according to our results, the literature usually reported major PSD concepts. However, other more specific and smaller categories, also highly affecting the emotional functioning of individuals—like loneliness and distress—were neglected.With regard to the most common psychosocial difficulties, the literature and the patients' outcomes matched almost entirelyw et al. , based osymptoms –50 . Literature should therefore consider the positive and negative consequences of treatments on patient's PSDs and not only report the positive impact on them. This type of information would be useful to help clinicians decide among the wide range of interventions available.Regarding the related variables of PSDs, the results of our analysis revealed interesting and alarming gaps in the recent literature. Essentially, only few related variables of onset of PSDs were reported in research. cognitive functions, relationships with others, energy and drive, and employment problems as related variables of PSDs would have to be described as insufficient. These components are fundamental to the understanding of the psychosocial functioning of depressed patients. Hence, if the aim is to reach a general development in this direction, future research needs to focus on these gaps.On the other hand, current scientific literature concentrates on clinical variables and treatments as the only PSDs' related variables, whereas patients additionally highlight the relationships between different PSDs. Specifically, if the patient perspective is considered, literature onOverall, these findings reveal the fact that there is a discrepancy between the patient's and the health science researchers' perspective when analysing not only the biological and psychological factors of depression, but also the socioeconomic and environmental variables. The subject's view of his health and functioning is not usually taken into account when designing research studies in depression. In this sense, future studies may address this wider view in a more accurate way by taking into account additional sources that are primarily focused on the subject and its sociological environment. From a sociological point of view it could be claimed that the mainstream perspective from which most institutions and professionals develop their work is not reflecting the quotidian environmental stressors that interfere with an adaptive and healthy development of everyday life in people with depression. It could be hypothesized that this is a result of the current and most common definition of depression through a list of symptoms facilitated by established diagnostic manuals: the DSM and ICD. They are based on passive categorical labelling , which list a number of prototype behaviours that could occur if certain psychological problems occur. This conceptualization within the medical model does not take into account the life circumstances and biographical context, which the individual interacts with, and thus the psychological sense that provides the key to understanding how this problem has been generated and why it remains is disregarded.The ICF model has been created by WHO as a complementary tool to the diagnostic systems (DSM and ICD) to describe the day-to-day functioning of people. Within the ICF model, psychosocial difficulties are seen as a continuum and as a result of the complex interaction of environmental variables, mental functions, personal variables, activities and participation, and health status. Another option for personalization of medicine and treatment in real clinical settings might be the inclusion of evaluations of daily functional problems experienced by patients. Therefore we suggest customization of healthcare—with therapeutic decisions being tailored to the individual patient. The diagnostic testing has to be adjusted accordingly for selecting appropriate therapies. Eventually, engagement of patients in identifying specific personal problems  could provide more personalized information and change the pattern of diagnosing. For example, ecological momentary assessments of patients via different technologies, eventual self-monitoring, or even more collaborative interactions with therapists and professional carers would enhance research in this direction and give us a new insight. Recent literature based on ecological momentary assessments (EMA) gives us promising results in this line. Different studies , 57 haveEcological momentary assessments have been initially used to identify moment-to-moment patterns and mechanisms of psychopathology , but witMoreover, the patient perspective on PSDs and their related variables has also an impact on the clinical arena. Results of this study highlight the need for change and adjustment of future strategies for treatment and rehabilitation in depression. The process could be more productive, when clinicians select interventions covering PSDs and their modifiable determinants or related variables, and not only symptoms. Solving some activity problems, for example, can help in improving other areas as well. Furthermore, personalization of therapies according to individuals' PSDs will enhance the treatment process. In general, identification of specific interventions for particular psychosocial difficulties is necessary and future research should address this issue. Universitario de la Princesa in general did not allow us to have a more representative sample in terms of age. However, we decided to proceed with the focus group, because, in fact, a more homogeneous sample of participants is often preferable in terms of age, since it might increase the group comfort level when discussing sensitive topics. Therefore we could assess a wide variety of PSDs including sexual problems, relationships with the therapist, and somatic problems. Finally, in order to achieve more reliable results, we conducted individual interviews with 80 patients ranging substantially in terms of age. Thereat, in spite of this limitation, we found a high variability of PSDs and the valuable finding about the discrepancies between literature and patient reports. Further studies reaching saturation points should be done to test if there are other underexplored PSDs.The findings of this study, however, have to be interpreted assuming its limitations. First, only one focus group of seven patients was performed and saturation point—considering the quantity of information regarding the analysed PSDs—was not reached; therefore the focus group can be considered by some as nonrepresentative. As a partial solution we included a heterogeneous group through maximum sampling variation regarding sex, working status, and clinical status. The age range, however, was between 44 and 55 years, which might have caused a potential bias in the identification of PSDs. This is an important limitation, since some psychosocial difficulties, usually experienced in particular ages, like physical pain in older adults, or problems with intimate relationships or loss of life goals in younger adults, might have been omitted by the nonpresence of representatives of these age groups in our focus group. During the selection process, the psychiatrists who were responsible for the recruitment of participants took into consideration the maximum variability of sampling, but the time frame and the profile of patients in the catchment area of the HospitalSecond, a limited period of time was included for the literature search (studies between 2005 and 2010). The search was performed over papers published within the cited period of time because of temporal limitations and because we were interested in the recent literature outcomes rather than general literature. However, we made replication of the original search from 2011 to September 2013 in order to check if any new PSDs will occur in comparison with the initial search and the results did not identify new PSDs. The results of the review have to be read in light of the limitations due to the type of databases consulted. This review study was a component of a larger project that gathered psychosocial factors from several mental health and neurological conditions where only Medline and PsycINFO were included. As a consequence, an amount of studies might have not been identified. As a limitation concerning the reliability of the extraction process we have to indicate that only 20% of articles were independently double checked. Finally, by limiting our search to English literature, we might have omitted relevant papers in other languages.The present study is, to our knowledge, the first to analyse the full set of psychosocial difficulties and their related variables in depression through the literature and the patient perspective. We made an elaborate comparison between both sources of information in order to verify whether recent research literature reported all the PSDs and related variables that are important for depressed patients and to identify the existing gaps in this area. Regarding the research on depression, the results obtained within our study show the existence of many literature gaps and encourage future studies to focus on them more in depth. Concerning the clinical implications, this study emphasizes the need for change and adjustment of future strategies for treatment and rehabilitation in depression. We suggest that clinicians should select interventions that cover PSDs and their modifiable related variables and not only improve symptoms. Furthermore, eventual identification and personalization of therapies according to individuals' PSDs would potentially enhance the rehabilitation process.