bluuebunny/crossref_metadata_2025 huggingface dataset:
Assessment of thymic output in common variable immunodeficiency patients by evaluation of T cell receptor excision circles
Authors: GUAZZI V , AIUTI F , MEZZAROMA I , MAZZETTA F , ANDOLFI G , Mortellaro A , Pierdominici M , FANTINI R , MARZIALI M , AIUTI A
Published: March 2003

Common variable immunodeficiency (CVID) is a heterogeneous syndrome characterized by repeated infections and hypogammaglobulinaemia. Additionally, T-cell abnormalities including lymphopenia, decreased proliferation to mitogens and antigens, and the reduced production and expression of cytokines, have also been observed. In this study we have investigated the expression of naive, memory and activation markers in T-cell subpopulations in 17 CVID patients in comparison to age-matched normal controls. The numbers of CD4+ T cells, including CD45RA+CD62L+ and, to a lesser extent, CD45RA–CD62L+/RA+CD62L– were significantly reduced in patients, whereas CD8+ T cells were within normal range. In contrast, HLA-DR+ cells were increased both in CD4+ and CD8+ T cells. To assess the thymic output, we analysed the presence of T-cell receptor excision circles (TRECs) in CD4+ and CD8+ T cells by quantitative PCR. TRECs were decreased significantly in patients and the rate of TREC loss was higher with increasing age. TRECs correlated with naive CD4+ T cells, whereas there was an inverse relationship between TRECs and CD8+HLA–DR+ and CD8+CD45RA–CD62L+/RA+CD62L– T cells. Our results suggest the presence of a defect in the naive T cell compartment with origin at the thymic level in CVID, and indicate that TREC may be a useful marker to monitor thymic function in this primary immunodeficiency.
The genotype of mice influences the autoimmune response to spliceosome proteins induced by cytomegalovirus gB immunization
Authors: LIPES J , SKAMENE E , NEWKIRK M M
Published: March 2003

In previous studies we have established a link between cytomegalovirus (CMV) infection and an autoimmune response to the U1–70 k protein of the spliceosome in man. This autoimmune response, generally referred to as the anti-RNP (ribonucleoprotein) antibodies, is observed in about 30% of patients with systemic lupus erythematosus (SLE). We have also found that the CMV glycoprotein B (CMV gB) when expressed in a adenovirus vector (Ad) could induce a significant anti-U1–70 k antibody response in several strains of mice, such as C3H, MRL and BALB/c. In the present study we examined the autoimmune response induced by immunization with Ad-gB in A/J and C57BL/6 (B6) mice and determined whether there was any autoimmune phenotype similar to that observed in patients with SLE. Thus groups of A/J and B6 mice were immunized with Ad/gB or with Ad alone and then observed for possible skin or kidney disease. In addition the autoantibody response to the spliceosome was measured, and the target antigens identified by immunoblot techniques. All of the A/J mice mounted a very high IgG response primarily to the U1–70 k protein of the spliceosome, with evidence of a rapid spreading of the autoantibody response to other components of the complex. In contrast, B6 mice mounted only a very low titre autoantibody response and failed to show signs or symptoms of autoimmunity. The A/J but not the B6 mice were found to have deposits of IgG in their kidneys, which were consistent with abnormal levels of blood urea nitrogen in the A/J but not B6 mice. This study demonstrates the importance of the genetic background in the susceptibility to autoimmunity.
Phenotypic and functional analysis of human T lymphocytes in early second- and third-trimester fetuses
Authors: ZHAO Y , DAI Z-P , LV P , GAO X-M
Published: March 2003

This study was undertaken to investigate the phenotypic and functional status of T lymphocytes of human fetuses from early second- to third-trimester. Cord blood samples were obtained from 19 healthy human fetuses (gestation weeks: 18–36), by cordocentesis, and 16 term newborns (gestation weeks 37–42). Maternal and unrelated male blood samples were also taken as controls. Percentage of lymphocytes in fetal white blood cells was 79·3%, reducing to 40% by term birth, much higher than that of adults. Cord blood mononuclear cells (CBMC), prepared by density gradient centrifugation followed by lysis of erythrocytes, were stained using PE- or FITC-labelled monoclonal Abs and analysed by flow cytometry. The frequencies of CD3+ T cells in fetal (40·1%) and neonatal (42·4%) CBMC were significantly lower than that of men (59·6%) and pregnant women (53·6%). Proportions of CD8+ T cells (9·5%), γδ-T cells (0·5%) and NK cells (4·8%) in fetal CBMC were also lower than that of neonates (except γδ-T cells) and adults. A negative linear correlation (r = −0·609) between the ratio of CD4+/CD8+ T cells in fetal blood and gestation age could also be established. Fetal CBMC showed vigorous spontaneous proliferation but failed to respond to mitogen (PHA) or allogeneic stimulation in vitro. The fetal mononuclear cells were unable to produce IL-2, IL-4 or IFN-γ, but spontaneously secreted IL-10, IL-6 and TNF-αin vitro. Stimulation with PHA up-regulated the production of IL-10, IL-6 and TNF-α substantially.
Immunosuppressive properties of human amniotic membrane for mixed lymphocyte reaction
Authors: UETA M , KWEON M-N , SANO Y , SOTOZONO C , YAMADA J , KOIZUMI N , KIYONO H , KINOSHITA S
Published: March 2003

The combination of allograft limbal transplantation (ALT) and amniotic membrane transplantation (AMT) has been applied in the treatment of severe ocular surface diseases. The beneficial effect of this combination has been thought to result from possible immunosuppressive ability of amniotic membrane (AM). However, the mechanisms of any such ability remain unknown. In this study, we investigated whether human AM has the ability to suppress allo-reactive T cell responses in vitro. For mixed lymphocyte reaction (MLR), lymphocytes isolated from lymph nodes of C57BL/6 mice (Mls1b, Vβ6+) were cultured with irradiated splenocytes from DBA/2 mice (Mls1a, Vβ6−) with or without human AM. For carboxyfluorescein diacetate succinimidyl ester (CFSE) experiments, responder lymph node cells were labelled with a stable intracellular fluorescent dye and cultured with irradiated stimulator cells. The ratio of responder Vβ6+ T cells was then determined by FACS analysis, and the division profiles of responder Vβ6+ T cells were analysed by CFSE content. Furthermore, Th1 and Th2 cytokine synthesis by allo-reactive T cells in MLR culture supernatants was determined by enzyme-linked immunosorbent assay (ELISA). Addition of AM to the MLR culture resulted in the significant inhibition of thymidine incorporation compared with control culture lacking AM. The population of responder CD4+Vβ6+ T cells was significantly reduced in the AM-treated culture in comparison to control. CFSE analysis revealed less division and lower proliferation of responder CD4+Vβ6+ T cells in cultures with AM than without. In addition, allo-rective T cell synthesis of both Th1 (IL-2 and IFNγ) and Th2 (IL-6 and IL-10) type cytokine was significantly decreased in the presence of AM. These results indicate that human AM has the ability to suppress allo-reactive T cells in vitro. This inhibitory effect likely contributes to the success of the ALT-AMT combination.
Preclinical efficacy and safety of pascolizumab (SB 240683): a humanized anti-interleukin-4 antibody with therapeutic potential in asthma
Authors: HART T K , BLACKBURN M N , BRIGHAM-BURKE M , DEDE K , AL-MAHDI N , ZIA-AMIRHOSSEINI P , COOK R M
Published: March 2003

The type 2 helper T cell (TH2) cytokine interleukin (IL)-4 is thought to play a central role in the early stages of asthma. In an effort to develop an antibody treatment for asthma that neutralizes the effects of IL-4, a murine monoclonal antibody, 3B9, was generated with specificity for human IL-4. In vitro studies demonstrated that 3B9 inhibited IL-4-dependent events including IL-5 synthesis, TH2 cell activation and up-regulation of immunoglobulin E expression. 3B9 was then humanized (pascolizumab, SB 240683) to reduce immunogenicity in humans. SB 240683 demonstrated species specificity for both monkey and human IL-4 with no reactivity to mouse, rat, cow, goat or horse IL-4. Pascolizumab inhibited the response of human and monkey T cells to monkey IL-4 and effectively neutralized IL-4 bioactivity when tested against several IL-4-responsive human cell lines. Affinity studies demonstrated rapid IL-4 binding by pascolizumab with a slow dissociation rate. In vivo pharmacokinetic and chronic safety testing in cynomolgus monkeys demonstrated that pascolizumab was well tolerated, and no adverse clinical responses occurred after up to 9 months of treatment. Three monkeys developed an anti-idiotypic response that resulted in rapid pascolizumab clearance. However, in the chronic dosing study the antibody response was transient and not associated with clinical events. In conclusion, pascolizumab is a humanized anti-IL-4 monoclonal antibody that can inhibit upstream and downstream events associated with asthma, including TH2 cell activation and immunoglobulin E production. Clinical trials are under way to test the clinical efficacy of pascolizumab for asthma.
Elevated production of interleukin-18 is associated with renal disease in patients with systemic lupus erythematosus
Authors: WONG C K , HO C Y , LI E K , Tam L S , LAM C W K
Published: March 2003

To investigate the production mechanism and proinflammatory role of the cytokine interleukin (IL-18) in lupus nephritis, we investigated the plasma concentrations of IL-18 and nitric oxide (NO) and the release of IL-18 and NO from mitogen-activated peripheral blood monomuclear cells (PBMC), in 35 SLE patients with renal disease (RSLE), 37 patients without renal disease (SLE) and 28 sex- and age-matched healthy control subjects (NC). IL-18 and NO concentrations were measured by ELISA and colourimetric non-enzymatic assay, respectively. Gene expressions of IL-18 and IL-18 receptor were analysed by RT-PCR. Plasma IL-18 and NO concentrations were significantly higher in RSLE than NC (both P < 0·01). Elevation of plasma IL-18 in RSLE correlated positively and significantly with SLE ­disease activity index and plasma NO concentration (r = 0·623, P < 0·0001 and r = 0·455, P = 0·017, respectively), and the latter also showed a positive and significant correlation with plasma creatinine (r = 0·410, P = 0·034) and urea (r = 0·685, P < 0·0001). There was no significant difference in gene expressions of IL-18 and IL-18 receptor in PBMC among RSLE, SLE and NC. Percentage increase in culture supernatant IL-18 concentration was significantly higher in RSLE than SLE and NC (both P < 0·05). The basal NO release was significantly higher in RSLE than that in SLE and NC (both P < 0·005). IL-18 is therefore suggested to play a crucial role in the inflammatory processes of renal disease in SLE.
T cell vaccination in multiple sclerosis patients with autologous CSF-derived activated T cells: results from a pilot study
Authors: VAN DER AA A , HELLINGS N , MEDAER R , GELIN G , PALMERS Y , RAUS J , STINISSEN P
Published: March 2003

Myelin-reactive T cells are considered to play an essential role in the pathogenesis of multiple sclerosis (MS), an autoimmune disease of the central nervous system. We have previously studied the effects of T cell vaccination (TCV), a procedure by which MS patients are immunized with attenuated autologous myelin basic protein (MBP)-reactive T cell clones. Because several myelin antigens are described as potential autoantigens for MS, T cell vaccines incorporating a broad panel of antimyelin reactivities may have therapeutic effects. Previous reports have shown an accumulation of activated T cells recognizing multiple myelin antigens in the cerebrospinal fluid (CSF) of MS patients. We conducted a pilot clinical trial of TCV with activated CD4+ T cells derived from CSF in five MS patients (four RR, one CP) to study safety, feasibility and immune effects of TCV. CSF lymphocytes were cultured in the presence of rIL-2 and depleted for CD8 cells. After 5–8 weeks CSF T cell lines (TCL) were almost pure TCRαβ+CD4+ cells of the Th1/Th0 type. The TCL showed reactivity to MBP, MOG and/or PLP as tested by Elispot and had a restricted clonality. Three immunizations with irradiated CSF vaccines (10 million cells) were administered with an interval of 2 months. The vaccinations were tolerated well and no toxicity or adverse effects were reported. The data from this small open-label study cannot be used to support efficacy. However, all patients remained clinically stable or had reduced EDSS with no relapses during or after the treatment. Proliferative responses against the CSF vaccine were observed in 3/5 patients. Anti-ergotypic responses were observed in all patients. Anti-MBP/PLP/MOG reactivities remained low or were reduced in all patients. Based on these encouraging results, we recently initiated a double-blind placebo-controlled trial with 60 MS patients to study the effects of TCV with CSF-derived vaccines in early RR MS patients.
T cell activation by soluble C1q-bearing immune complexes: implications for the pathogenesis of rheumatoid arthritis
Authors: JIANG K , CHEN Y , XU C-S , JARVIS J N
Published: March 2003

Both innate and adaptive immune systems are thought to participate in the pathogenesis of rheumatoid arthritis in adults and children. The experiments reported here were undertaken to examine how immune complexes, potent stimulators of inflammation, may regulate cells of the adaptive immune system. Human T cells were prepared from peripheral blood by negative selection and incubated with bovine serum albumin (BSA)–anti-BSA immune complexes that were formed in the presence or absence of human C1q. C1q-bearing immune complexes, but not unopsonized complexes, elicited both TNF-α and IFN-γ secretion from human T cells. Secretion of both cytokines was time- and dose-dependent. Cross-linking C1q on the cell surface of T cells produced the same results. Cytokine secretion was not inhibited by blocking the C3b receptor (CR1, CD35) on T cells prior to incubation with immune complexes. Reverse transcriptase polymerase chain reaction (RT-PCR) of immune complex-stimulated cells revealed accumulation of both TNF-α and IFN-γ mRNA within 2 h post-stimulation. IL-2 was not detected in cell culture supernatants, but IL-2 receptor α chain (CD25) was detected in low density on a small proportion of T cells activated by C1q-bearing immune complexes. Secretion of both cytokines was inhibited partially, but not completely, by IL-10. These experiments show that immune complexes, potent inflammatory mediators, may activate T cells through a novel mechanism. These findings have implications for chronic inflammatory diseases in humans.
Human IgG Fc-binding phage antibodies constructed from synovial fluid CD38+ B cells of patients with rheumatoid arthritis show the imprints of an antigen-dependent process of somatic hypermutation and clonal selection
Authors: VAN ESCH W J E , REPARON-SCHUIJT C C , HAMSTRA H J , VAN KOOTEN C , LOGTENBERG T , BREEDVELD F C , VERWEIJ C L
Published: March 2003

The persistent presence of rheumatoid factors (RFs) in the circulation is a characteristic phenomenon in patients with rheumatoid arthritis (RA). Recent data indicate that RFs associated with seropositive RA are derived from terminally differentiated CD20–, CD38+ plasma cells (PCs) present in synovial fluids of the inflamed joints. These cells were shown to secrete RFs actively and are thought to originate from germinal centre (GC)-like structures present in the inflamed synovium. To obtain a representative image of the structural properties of IgM and IgG RFs associated with RA, phage antibody display libraries were constructed from CD38+ PCs isolated from the inflamed joints of RF-seropositive patients with RA. Subsequently, human IgG Fc-binding monoclonal phage antibodies were selected and analysed. The data suggest that RA-associated RFs are encoded by a diverse set of VL and a more restricted set of VH regions. VH gene family usage of PC-derived IgM- and IgG-RFs was found to be restricted to the VH1 and 3 gene families, with a preference for VH3, and many different VL genes were shown to contribute to RF specificity. Clonally related VH as well as VL sequences were identified, based on the presence of identical CDR3 regions and shared somatic mutations. In this B cell selection process base-pair substitutions as well as deletions of triplets in CDR regions, leaving the transcripts in frame, were involved. Together, these data provide further evidence for an Ag-driven immune response in the terminal differentiation into RF-producing PCs in patients with RA, including expansion of clonally related B cells, selection and isotype switching, all hallmarks of a GC reaction.
Molecular characterization of homozygous hereditary factor I deficiency
Authors: BARACHO G V , NUDELMAN V , ISAAC L
Published: March 2003

We have studied the molecular basis of factor I (fI) deficiency in two Brazilian sisters from a consanguineous family. By reverse transcription-polymerase chain reaction we observed that all fI cDNA amplified products from one sister had the same size as those of normal cDNA, however, they were significantly less intense. Sequencing analysis of subcloned cDNA revealed a dinucleotide insertion (AT) between positions 1204 and 1205 in the 11th exon that creates a stop codon 13 bp downstream of the insertion site. Genomic DNA sequencing and heteroduplex analysis confirmed that both probands are homozygous for this mutation, whereas their parents are heterozygous. The stop codon and the diminished amounts of fI cDNA could indicate increased fI mRNA instability, perhaps due to a mechanism of nonsense-mediated decay. This hypothesis is consistent with our observation that treatment with the translation inhibitor cycloheximide stabilized fI mRNA expression in proband's fibroblasts.
Phenotypic characterization of human decidual macrophages
Authors: HEIKKINEN J , MÖTTÖNEN M , KOMI J , ALANEN A , LASSILA O
Published: March 2003

Pregnancy is a challenge to the immune system, which not only has to protect the mother and the fetus from invading pathogens but to also maintain immunological tolerance against the fetus. However, the mechanisms inhibiting local immune responses in the maternal decidual tissue are poorly understood. We have studied decidual CD14+ macrophages, which may be important in the maintenance of a tolerance against the developing fetus. Decidual macrophages expressed HLA-DR, but lower levels of costimulatory molecule CD86 than peripheral blood CD14+ monocytes from pregnant and non-pregnant women. Decidual macrophages produced spontaneously high levels of interleukin-10. Our findings suggest that decidual macrophages could represent an inhibitory type of APCs. Supporting this conclusion indoleamine 2,3-dioxygenase (IDO), suggested to have an immunosuppressive role in pregnancy, was expressed in decidual macrophages. Furthermore, decidual macrophages were not able to differentiate into dendritic cells under the influence of IL-4 + GM-CSF. These results suggest an immunoinhibitory function of decidual macrophages at the maternal–fetal interface.
Silica accelerated systemic autoimmune disease in lupus-prone New Zealand mixed mice
Authors: BROWN J M , ARCHER A J , PFAU J C , HOLIAN A
Published: March 2003

The genetic backgrounds of lupus-prone murine models are a valuable resource for studying the influence of environmental exposure on autoimmune diseases in sensitive populations. Epidemiological studies have shown associations between silica exposure and several autoimmune diseases, including scleroderma and systemic lupus erythematosus. To determine whether silica exposure can exacerbate systemic autoimmunity in genetically predisposed animals, New Zealand mixed mice were intranasally instilled twice with saline or saline suspensions of 1 mg silica or 500 µg TiO2, a dose equivalent in surface area, and were evaluated with respect to health and immune status. Survival in silica exposed NZM mice was decreased compared to saline and TiO2 exposed mice. Proteinuria levels were elevated in silica exposed mice. Levels of circulating immune complexes, autoantibodies to nuclear antigen (ANA), histone, and double stranded DNA were measured every two weeks by ELISA. Circulating immune complexes showed a trend towards an increased acceleration in levels in the silica exposed mice compared to saline and TiO2 exposed mice. ANA levels were significantly higher in silica exposed animals compared to saline and TiO2 exposed animals (0·237 ± 0·03 versus 0·140 ± 0·029 and 0·125 ± 0·03, P < 0·05) 16 weeks postexposure. Autoantibodies to histone were also significantly elevated after 16 weeks in silica exposed animals compared to saline and TiO2 exposed animals (0·227 ± 0·03 versus 0·073 ± 0·015 and 0·05 ± 0·03, P < 0·05). In contrast, serum IgG levels were decreased in silica exposed NZM mice compared to the saline controls, however, IgM levels were unaffected. Lungs of the silica-exposed mice had increased inflammatory infiltrates as well as fibrotic lesions characterized by excess collagen deposition. Therefore, although NZM mice are susceptible to SLE, silica exposure significantly exacerbated the course of disease.
Corticotroph axis sensitivity after exercise in endurance‐trained athletes
Authors: Duclos  , Corcuff  , Arsac  , Moreau‐Gaudry  , Rashedi  , Roger  , Tabarin  , Manier
Published: March 2003

OBJECTIVE

The present study was conducted in order to describe human hypothalamo‐pituitary adrenal (HPA) axis adaptation in a model of repeated physical stress (endurance training) that causes a moderate increase in cortisol levels.

SUBJECTS

We performed the same stimulation tests (adrenal stimulation with ACTH or pituitary stimulation with combined CRH/LVP) in a population of 8 endurance‐trained athletes in two distinct situations: resting (baseline cortisol values) and 2 h after the end of strenuous exercise (increased cortisol values) to evaluate the HPA axis sensitivity to endogenous sustained increases in cortisol concentrations.

MEASUREMENTS

During these tests, saliva and plasma cortisol (Fs and Fp, respectively) were assessed and compared.

RESULT

Cortisol values in both plasma and saliva at the end of 2 h of exercise were significantly higher than in rested controls: Fs 11.5 ± 1.3 vs 6.5 ± 0.8 nmol.l−1 and Fp 428 ± 36 vs 279 ± 27 nmol.l−1 (post exercise vs post rest sessions, respectively, P < 0.001 for both). After either hormone test (CRH/LVP or ACTH), cortisol levels in plasma and saliva increased similarly when rest was compared to post exercise.  Saliva variations (Δ%) under exogenous hormone stimulation were dramatically greater than plasma variations. For example, under ACTH stimulation, the relative increments in cortisol were on control day: ΔFs 980 ± 139 vs ΔFp 218 ± 43% (saliva vs plasma, respectively, P < 0.05) and on exercise day: ΔFs 605 ± 89 vs ΔFp 102 ± 14% (saliva vs plasma, respectively, P < 0.05).

CONCLUSIONS

In endurance‐trained athletes, displaying a moderate but sustained endogenous cortisol increase: (1) ACTH responses following pituitary stimulation are not blunted, (2) cortisol responses following maximal adrenal stimulation are not blunted. Our results favour the hypothesis of a decreased pituitary sensitivity to cortisol negative feedback whereas the hypothesis of a major decreased adrenal sensitivity to ACTH was discarded. The greater ability of saliva assays to detect a cortisol increase strongly supports its use in the study of HPA physiology, whether under basal or dynamic conditions.
Urinary glycosylated, free and total pyridinoline and free and total deoxypyridinoline in diabetes mellitus
Authors: Cloos  , Wahl  , Hasslacher  , Traber  , Kistner  , Jurkuhn  , Schmidt‐Gayk
Published: March 2003

OBJECTIVE

Published data on bone metabolism in diabetes mellitus are conflicting. We have measured pyridinium crosslinks, biochemical markers of bone resorption, in order to evaluate bone resorption in diabetes mellitus. We also wished to investigate whether, as a consequence of chronic hyperglycaemia, pyridinoline is glycosylated to a greater extent in patients with diabetes mellitus.

DESIGN AND PATIENTS

This cross sectional study included 142 patients (64 males, 78 females) with insulin dependent and non‐insulin dependent diabetes mellitus (IDDM and NIDDM). These patients were compared to a healthy control group of 99 individuals (39 males and 60 females).

MEASUREMENTS

Pyridinium crosslinks, glycosylated, free and total pyridinoline (gPYD, fPYD, tPYD) and free and total deoxypyridinoline (fDPD, tDPD) were measured in a spot urine sample by high performance liquid chromatography (HPLC). Urinary creatinine, albumin and glucose were also measured.

RESULTS

In the diabetic group, values of urinary gPYD and tDPD were significantly lower than in controls. gPYD excretion was lowest in patients with severe glycosuria. Free pyridinium crosslinks, both fPYD and fDPD, were excreted to a significantly lower extent. The molar ratio of tPYD to tDPD was significantly increased in diabetes mellitus.

CONCLUSIONS

Decreased excretion of tDPD suggests low bone resorption in IDDM and NIDDM. Pyridinoline is not glycosylated to a greater extent in diabetes mellitus and tends to be decreased in proportion to the degree of glycosuria. Excretion of gPYD, fPYD and fDPD is depressed in severe glycosuria. Diminished degradation to the final products, fPYD and fDPD, might represent increased resistance to enzymatic activity or diminished enzymatic activity. The increased molar ratio tPYD/tDPD in urine suggests an increased ratio in bone collagen in diabetes mellitus.
Serum substances that interfere with thyroid hormone assays in patients with chronic renal failure
Authors: Iitaka Makoto , Kawasaki Satomi , Sakurai Shin‐ichi , Hara Yoshihito , Kuriyama Renjiro , Yamanaka Kayo , Kitahama Shinji , Miura Shiro , Kawakami Yoshito , Katayama Shigehiro
Published: March 2003

OBJECTIVE

Serum thyroid hormone concen_trations in patients with chronic renal failure are usually low, despite normal serum TSH levels. We investigated the effect on thyroid hormone assays of serum dialysable organic acids that are elevated in uraemic patients.

PATIENTS

Serum samples from 42 patients with chronic renal failure who were receiving haemodialysis and 37 sex‐ and age‐matched healthy subjects were examined.

DESIGN AND MEASUREMENTS

Serum thyroid hormone concen_trations were measured with an analogue radioimmunoassay (RIA), a labelled antibody assay, and an equilibrium dialysis/RIA method. Serum concen_trations of organic acids were determined with high performance liquid chromatography.

RESULTS

Serum thyroid hormone levels determined by an analogue RIA and a labelled antibody assay in uraemic patients increased, and serum concen_trations of organic acids decreased following haemodialysis. A significant association existed between serum free T3 (FT3) levels determined by an analogue RIA and serum concen_trations of indoxyl sulphate (IS) prior to dialysis. There was also a significant association between serum free T4 (FT4) levels determined by an analogue RIA and serum concen_tration of IS and hippuric acid (HA) prior to dialysis. There was a significant association between the changes of serum concen_trations of indole acetic acid (IAA) and FT4 concen_trations prior to and following haemodialysis when determined by an analogue RIA. Serum FT3 and FT4 levels significantly decreased after the addition of IS to serum from healthy subjects when determined by an analogue RIA but not by a labelled antibody assay. Serum FT4 levels, but not FT3 levels, decreased after addition of IAA when determined by an analogue RIA. Serum FT4 concen_trations determined by an equilibrium dialysis/RIA were significantly higher than those determined by the other two methods. The addition of IS, IAA, and HA to serum samples from healthy subjects significantly increased FT4 concen_trations when determined by an equilibrium dialysis/RIA method.

CONCLUSIONS

Increased serum levels of indoxyl sulphate, indole acetic acid and hippuric acid in sera of uraemic patients may interfere with thyroid hormone measurements when an analogue radioimmunoassay is used. In contrast, there was little interference with a labelled antibody assay. Dialysable organic acids may also interfere with thyroid hormone assays determined by an equilibrium dialysis/radioimmunoassay method.
A population‐based study of Graves' disease in Danish twins
Authors: Heiberg Brix Thomas , Christensen Kaare , Holm Niels V. , Harvald Bent , Hegedüs Laszlo
Published: March 2003

OBJECTIVE

The aetiology of Graves' disease (GD) is generally thought to fit a multi‐factorial pattern of inheritance in which clinical disease develops on the basis of genetic susceptibility interacting with environmental and endogenous factors. In previous twin studies the probandwise concordance rates for hyperthyroidism were as high as 0.86 in monozygotic twins and 0.20 in dizygotic twins, indicating a very strong genetic influence. In these studies, however, no effort was made to distinguish between GD and non‐autoimmune hyperthyroidism, and one study also included patients with simple non‐toxic goitre, hampering if not invalidating any conclusions. The aim of the present study was to determine whether there is a genetic contribution in the aetiology of GD.

DESIGN

Historical cohort study of pairs of same‐sex twins, with information on GD being gathered by questionnaire surveys in the 1950s and 1960s. All available hospital material was sought to veriify the diagnosis, which was assigned on the basis of clinical and histopathological evidence. The healthy co‐twins were followed through middle age by questionnaire surveys in the 1970s and 1980s.

PATIENTS

Same‐sex twin individuals born between 1870–1920, included in a population‐based nationwide register. A total of 118 subjects indicated hospitalization due to GD. A hospital record was available in 76 subjects. Of these, 55 (46 females and 9 males) could be classified as having GD.

MEASUREMENTS

Pairwise and probandwise concordance rates for GD in monozygotic and dizygotic twin pairs.

RESULTS

The probandwise concordance rates were 0.36 for monozygotic pairs and 0 for dizygotic pairs. The pairwise concordance rates were 0.22 and 0 for monozygotic and dizygotic pairs, respectively. The concordance rates were significantly (P = 0.012) higher in monozygotic than in dizygotic pairs.

CONCLUSIONS

These results confirm that genetic factors play an important role in the aetiology of Graves' disease. However, they may not be as powerful as previously thought.
Effectiveness of chronic treatment with alendronate in the osteoporosis of Cushing's disease
Authors: Di Somma Carolina , Colao Annamaria , Pivonello Rosario , Klain Michele , Faggiano Antongiulio , Tripodi Francesca S. , Merola Bartolomeo , Salvatore Marco , Lombardi Gaetano
Published: March 2003

BACKGROUND

Osteoporosis is common in patients with Cushing's disease and is likely due to an imbalance between bone formation and resorption. Alendronate is an aminobisphosphonate that is able to increase bone mass mainly by inhibiting bone resorption.

OBJECTIVE

We have evaluated the effect of chronic treatment with alendronate on bone mineral density (BMD) in patients with Cushing's disease.

PATIENTS

39 patients with Cushing's disease entered this study. 39 age‐, sex‐ and BMI‐matched normals served as controls for baseline evaluation. The 39 patients were divided into four groups: 1) 10 patients with active disease treated with alendronate and ketoconazole; 2) 11 patients with inactive disease treated with alendronate; 3) 8 patients with active disease treated with ketoconazole alone, 4) 10 patients with inactive disease received no treatment.

TREATMENT PROTOCOL

Alendronate was given for 12 months in a dose of 10 mg orally once daily after fasting at 0800 h in the morning. Ketoconazole was given in a dose of 200–600 mg orally daily, when pituitary surgery was unsuccessful.

STUDY DESIGN

Lumbar spine (L1–L4) and femoral neck BMD, serum osteocalcin (OC), urinary cross‐linked N‐telopeptides of type I collagen (Ntx) levels were evaluated at study entry, in patients and controls, and were repeated after 6 and 12 months in the 39 patients.

RESULTS

BMD values were lower in patients with Cushing's disease than in controls at both L1–L4 (0.72 ± 0.4 vs. 1.01 ± 0.6 g/cm2, P < 0.05) and femoral neck (0.69 ± 0.3 vs. 0.96 ± 0.6 g/cm2, P < 0.05). In the 39 patients with Cushing's disease considered as a whole, serum OC levels were lower (1.1 ± 0.1 vs 1.5 ± 0.1 nmol/l, P < 0.01), while Ntx values were higher than in controls (168 ± 25 vs. 61 ± 31 nmol BCE/mmol creatinine, P < 0.01). In the alendronate‐treated groups, serum OC levels increased, while Ntx levels significantly decreased after 6 and 12 months of treatment without any significant difference between the two groups. BMD values measured at L1–L4 and femoral neck significantly increased after 12 months of therapy. In patients of group 4, a significant increase of serum OC levels and a significant decrease of Ntx levels were observed together with a slight increase of BMD values after 12 months. No significant change in either biochemical markers or BMD values was found in patients of group 3.

CONCLUSIONS

Patients with Cushing's disease have osteoporosis which needs to be rapidly reversed to limit the risk of fracture. The results of the present study show that a 12 month treatment period with alendronate induced an improvement in bone mineral density greater than in untreated patients.
The GnRH receptor gene is preferentially expressed in functioning gonadotroph adenomas and displays a Mae III polymorphism site
Authors: Kottler Marie‐Laure , Seret‐Bégué Dominique , Lahlou Najiba , Assayag Michel , Carré Marie‐Claude , Lagarde Jean‐Pierre , Ajzenberg Christiane , Christin‐Maitre Sophie , Bouchard Philippe , Mikol Jacqueline , Counis Raymond , Warnet André
Published: March 2003

OBJECTIVE

Given the central role of the GnRH receptor (GnRHR) in the regulation of the gonadotrophin secretion, it might be implicated directly or indirectly in the pathogenesis of gonadotroph tumours.

DESIGN

We determined if GnRHR mRNA was expressed in gonadotroph tumours using RT‐PCR and analysed the GnRHR gene for the presence of mutations in its coding region, using direct sequencing of PCR products. Results were analysed according to the pattern of expression of α, β‐FSH and β‐LH subunit (SU) genes

SUBJECTS

RNA was extracted from 20 gonadotroph tumours identified by immunohistochemistry ( >10% of stained cells): 9 adenomas were functioning (high serum gonadotrophin levels), 3 were associated with high α‐SU levels and 8 were nonfunctioning. Genomic DNA was extracted from 64 normal subjects.

RESULTS

We found GnRHR mRNA in 12 tumours (60%): 8/9 functioning (88%), 1/3 α‐secreting (33%) and 3/8 nonfunctioning (37.5%) gonadotroph adenomas. There was a significant association between GnRHR expression and immunostaining for β‐FSH (P = 0.014). The nucleotide sequence of the amplified products was identical to that of human pituitary except for the presence, in 3 functioning adenomas, of a silent C to T transition at nucleotide 453 encoding for the serine residue situated in the second intracellular loop at position 151. Heterozygosity provided evidence that both alleles were transcribed in these tumours. This substitution creates a Mae III restriction site. Genomic DNA from normal subjects were then tested for the presence of this new polymorphism. The frequency of the heterozygosity (18.7%) was not significantly different from that found in gonadotroph tumours (25%) and this new Mae III polymorphism site cannot be used as a tumoural marker.

CONCLUSION

The GnRHR gene is preferentially expressed in functioning rather than in nonfunctioning gonadotroph adenomas, but no mutations altering the coding region of the gene were found to further substantiate its role in the pathogenesis of gonadotroph tumours.
Serum leptin and insulin concentrations in prepubertal lean, obese and insulin‐dependent diabetes mellitus children
Authors: Kamoda Tomohiro , Saitoh Hisako , Nakahara Satoko , Izumi Ishou , Hirano Takeki , Matsui Akira
Published: March 2003

OBJECTIVE

To evaluate the relationship between serum levels of leptin and insulin in prepubertal lean, obese and insulin‐dependent diabetes mellitus (IDDM) children.

SUBJECTS AND MEASUREMENTS

Prepubertal children, 16 lean, 17 obese and 16 IDDM were included in the study. Fastang serum leptin and insulin concentrations were measured by radioimmunoassays.

RESULTS

The serum level of leptin was significantly higher in obese children than in lean and IDDM children (P < 0.0001 and P < 0.0001, respectively), and showed a positive correlation with body mass index (BMI) for the combined group (lean, obese and IDDM; r = 0.77, P < 0.0001). In addition, the serum leptin level was higher in IDDM children than in lean controls (P < 0.01), whereas no difference was found in BMI between the two groups. The mean fasting serum levels of insulin were significantly elevated in IDDM children as compared with lean controls (P < 0.01). A significant positive correlation was found between serum insulin and leptin levels for the combined group (r = 0.37, P < 0.01). When a multiple regression analysis for all subjects was performed, the total contribution of all parameters, including gender, BMI and log insulin, accounted for 75% of the leptin variation. BMI (57.8%), log insulin (14.0%) and gender (3.2%) contributed significantly to this variation.

CONCLUSIONS

The elevated concentration of leptin in insulin‐dependent diabetic children, independent of body mass index, was probably caused by chronically increased serum insulin levels. We demonstrated that not only body mass index but also insulin was a significant independent predictor of serum leptin concentrations. It is therefore suggested that insulin might play an important role in regulating serum leptin concentrations independent of adiposity.
Defining the normal cortisol response to the short Synacthen test: implications for the investigation of hypothalamic‐pituitary disorders
Authors: Clark  , Neylon  , Raggatt  , Sheppard  , Stewart
Published: March 2003

OBJECTIVE

To define the normal cortisol response to the Short Synacthen Test using four different cortisol immunoassays and to assess the implications for the investigation of hypothalamic‐pituitary disorders.

DESIGN AND PATIENTS

The cortisol response to 250 μg im ACTH1–24 (Synacthen, Ciba Geigy) in 100 healthy volunteers using four different cortisol immunoassays has been measured. In 44 newly diagnosed and untreated patients with pituitary disease, basal and 30 minute post‐ACTH cortisol results were also determined using the four immunoassays.

RESULTS

The distribution of cortisol results at all time points and for all methods were non‐Gaussian and significant differences in the absolute values of the 5th–95th percentiles were found between methods (P < 0.01). At 30 min post‐Synacthen in normals the 5th percentile of the cortisol response ranged from 510 to 626 nmol/l with the different methods. Similarly the relationship between assay results differed at different time points. No effect of age on the cortisol response was found but for stimulated cortisol values and the incremental responses females showed significantly higher responses than males (P < 0.05) for most methods. Although there was a significant positive linear correlation (P < 0.001) between stimulated and basal cortisol values for all methods, no significant relationship was found between the incremental response and basal cortisol values. In the pituitary disease patients basal and 30 minute post‐ACTH cortisol results were significantly lower (P < 0.05 and < 0.001) than the control group using the same cortisol assay. When the results were compared to the 5th percentile of the gender and assay specific control group 33.3% of male and 17.4% of female patients failed the Synacthen test at 30 min.

CONCLUSIONS

The definition of the ‘normal’ response to Synacthen should be both gender and method related at all time points. The data suggest that up to one‐third of untreated patients with pituitary disease may have subtle defects in the hypothalamic‐pituitary‐adrenal axis.
Growth in children with craniopharyngioma following surgery
Authors: Tiulpakov A. N. , Mazerkina N. A. , Brook C. G. D. , Hindmarsh P. C. , Peterkova V. A. , Gorelyshev S. K.
Published: March 2003

OBJECTIVE

Hypopituitarism, including severe GH deficiency, is an almost inevitable outcome of craniopharyngioma. However, some GH deficient children with this tumour may grow normally or even have accelerated growth postoperatively. To study this phenomenon we have investigated the endocrine status, including IGF‐1 and its binding proteins IGFBP‐1 and IGFBP3, in children referred for follow‐up at various time intervals after surgery.

PATIENTS

Twenty‐five patients (14 boys and 11 girls, aged 3.8–18.9 years), were studied on 34 occasions between 0.5 to 10.8 years after surgery. The tumour was intrasellar in 11 cases and suprasellar in 14.

METHODS

Height and height velocity were recorded as SDS values. Body mass index (BMI) was calculated as weight/height2. GH secretory status was evaluated by stimulation both with oral clonidine and with GH releasing hormone. Serum insulin (INS) levels were studied following an oral glucose load. IGF‐1, IGFBP‐3, IGFBP‐1 and prolactin serum concentrations were evaluated in fasting samples. All hormones were measured by radioimmunoassay.

RESULTS

All patients had growth hormone (GH) deficiency. Height (HtSDSCA) and height velocity SDS for chronological age (HVSDSCA) decreased progressively after surgery (r = –0.47, P = 0.005, and r = –0.4, P = 0.032, respectively) but four patients had normal HtSDSCA 6.1 to 10.8 years following their first surgery.

There was a significant correlation between BMI and HtSDSCA (r = 0.37, P = 0.03). BMI in children with suprasellar craniopharyngioma was significantly greater than that in patients with intrasellar tumour (23.3 ± 7.0 vs. 17.3 ± 1.4 kg/m2; P = 0.001). In 13 of 33 cases oral glucose load was accompanied by hyperinsulinaemia with serum INS levels greater than 50 mU/l. The mean area under the curve (AUC) of INS after glucose load (INSAUC) in the suprasellar group was greater than in the patients with intrasellar lesion (6945.5 ± 4411.8 vs. 2495.5 ± 1768.8 mU/l. min P = 0.001). The log INSAUC correlated significantly with HtSDSCA (0.37, P = 0.03). Fasting serum IGF‐1 and IGFBP3 levels were normal in 8 and 12 of the 31 measurements, respectively. Both IGF‐1SDS and IGFBP‐3SDS correlated significantly with HtSDSCA (r = 0.77, P = 0.0002 and r = 0.65, P = 0.0001, respectively) and the log INSAUC (r = 0.39, P = 0.035, r = 0.56, P = 0.002, respectively). As determined by forward stepwise regression analysis, IGF‐1SDS was the single most important predictor of HtSDSCA (R2 = 0.33, P = 0.001) in the subset mode.

CONCLUSION

A few children with craniopharyngioma grew normally after surgery, in spite of being GH‐deficient. This growth phenomenon, which is usually accompanied by obesity, was more common in patients with suprasellar tumour and is likely to be associated with the effect of IGF‐1 bioavailability, which, in turn, may be modulated largely by insulin.
Comparison of the diagnostic utility of the simplified and standard i.m. glucagon stimulation test (IMGST)
Authors: Orme Stephen M. , Price Alun , Weetman Anthony P. , M. Ross Richard J.
Published: March 2003

OBJECTIVE

To compare the peak GH and cortisol responses to the simplified and standard i.m. glucagon stimulation test (IMGST) in order to determine whether the use of a simplified form of this test results in a loss of diagnostic utility.

PATIENT AND DESIGN

A retrospective study of 35 consecutive patients with pituitary disease who underwent a standard IMGST to measure GH (n = 34) or cortisol (n = 21) reserve at the programmed investigation unit of the Northern General Hospital, Sheffield.

MEASUREMENTS

Cortisol, glucose and GH levels were measured at baseline and 30. 60, 90, 120, 150, 180, 210 and 240 min during the standard IMGST. We used the hormone levels at baseline, 150 and 180 min for the simplified IMGST and compared the cortisol and GH responses achieved.

RESULTS

The median (range) peak GH and cortisol responses to simplified and standard IMGST were comparable 4.2 (0.5–47.5) vs 5.4 (0.5–47.5) mU/l (P = 0.75) and 635 (306–1669) vs 647 (306–1669) nmol/l (P = 0.801), respectively. Using the standard ‘cut‐off’ points to assess GH (9 mU/l) or cortisol deficiency (580 nmol/l) there was no significant loss of diagnostic utility (P = 0.98 and P = 1.00, respectively).

CONCLUSIONS

The simplified and standard IMGST have a similar diagnostic utility. The simplified IMGST could replace the standard IMGST as a screening test for GH and cortisol reserve in pituitary disease. The insulin‐induced hypoglycaemia stress test can be reserved for cases in which a second assessment of pituitary reserve is clinically indicated or because of an indeterminate biochemical result.
An in vivo study of the cortisol–cortisone shuttle in subcutaneous abdominal adipose tissue
Authors: Katz J. R. , Mohamed‐Ali V. , Wood P. J. , Yudkin J. S. , Coppack S. W.
Published: March 2003

OBJECTIVE

Previous in vitro studies have demonstrated significant 11‐beta hydroxysteroid dehydrogenase (11 β‐HSD) oxo‐reductase activity in visceral, but not subcutaneous adipose stromal cells. We have conducted an in vivo study of the cortisol–cortisone shuttle in subcutaneous abdominal adipose tissue.

DESIGN

We measured arteriovenous (A‐V) differences in serum cortisol and cortisone across subcutaneous abdominal adipose tissue and forearm muscle in a heterogeneous group of subjects.

PATIENTS

We studied 34 subjects (male:female = 12 : 22), age median (interquartile range) 45 (19–65) years, body mass index 32.7 (20.4–77.1) kg m−2, total body fat 34.4 (5.6–119.1) kg.

MEASUREMENTS

Serum cortisol and cortisone were measured in serum samples from a radial artery, superficial epigastric vein and deep forearm vein. Abdominal adipose and forearm blood flow rates were measured by 133Xenon washout and plethysmography, respectively.

RESULTS

For cortisone, there was significant (P < 0.001) clearance by adipose tissue, with an A‐V difference of 4 (1–7) nmol/l. For cortisol there was a trend for arterial concentrations (203 (142–292) nmol/l) to be lower than venous (225 (152–263) nmol/l), but this was not significant. The adipose tissue cortisone clearance rate correlated with total body fat (r = 0.35, P = 0.05).

CONCLUSIONS

We have demonstrated 11 β‐HSD oxo‐reductase activity in subcutaneous abdominal adipose tissue, which may be increased in obesity.