Miracle-dz/UniFAR-FTUs huggingface dataset:
Optimizing the transduction efficiency of human hematopoietic stem cells using capsid-modified AAV6 vectors in vitro and in a xenograft mouse model in vivo

Background—Although recombinant adeno-associated virus serotype 2 (AAV2) vectors have gained attention owing to their safety and efficacy in number of Phase I/II clinical trials, their transduction efficiency in hematopoietic stem cells (HSCs) has been reported to be low. Only a handful of additional AAV serotype vectors have been evaluated, and comparative analyses of their transduction efficiency in HSCs from different species have not been performed. Methods—Here, we evaluated the transduction efficiency of all available AAV serotype vectors (AAV1 through AAV10) in primary mouse, cynomolgus monkey, and human HSCs, respectively. The transduction efficiency of the optimized AAV vectors was also evaluated in human HSCs in a murine xenograft model in vivo. Results—We observed that although there are only six amino acid differences between AAV1 and AAV6, AAV1, but not AAV6, transduce mouse HSCs cells well, whereas AAV6, but not AAV1, transduce human HSCs well. None of the 10 serotypes transduce cynomolgus monkey HSCs in vitro. We also evaluated the transduction efficiency of AAV6 vectors containing mutations in surface-exposed tyrosine residues, and observed that tyrosine (Y) to phenylalanine (F) point mutations in residues 445, 705, and 731, led to a significant increase in transgene expression in human HSCs in vitro and in a mouse xenograft model in vivo. Discussion—These studies suggest that the tyrosine-mutant AAV6 serotype vectors are the most promising vectors for transducing human HSCs, and that it is possible to further increase the transduction efficiency of these vectors for their potential use in HSC-based gene therapy in humans.
NT-methylhistidine in urine

The claim of Millward, Bates, Grimble, Brown, Nathan & Rennie 1(1980) Biochem. J. 190, 225-2281 that muscle actomyosin contributes as little as 25% of urinary NT-methylhistidine is not consistent with other published data from that group [Bates, DeCoster, Grimble, Holloszy, Millward & Rennie (1980) J. Physiol. (London) 303, 41 P1 or with literature values. It appears likely that the turnover rate of muscle actomyosin has been considerably underestimated and that when realistic rates of protein turnover are used, muscle tissue methylhistidine in urine.
Columns Are Quality Improvement Messages Registering ?

More often than not, commentaries highlight the findings of studies with important new positive findings. Instead, we highlight three papers that we believe are important and have largely negative or null findings. We are all familiar with efforts, particularly in the last decade or so that have been undertaken to improve the quality and efficiency of medical care. Changing the behavior of physicians and other health care workers is at the crux of many of these efforts. Some progress has been made in improving quality, but three papers in this issue deal with various aspects of these efforts, and they highlight the difficulty of communicating and bringing about some of the changes and improvements that many would agree need to occur. The papers report on three commonly used approaches for changing clinical practice. Clinical pathways and computerized decision support have been advocated to improve adherence with practice guidelines, and Quality Improvement Organizations (QIOs) have been charged with assisting providers in quality improvement (QI) activities. Tierney et al. (2005) report on a randomized trial of just-in-time, tailored guideline-based suggestions via an electronicmedical record for clinicians for improving outpatient management of asthma or chronic obstructive pulmonary disease. They found that the intervention had no effect on adherence with recommendations, medication compliance, quality of life, satisfaction, or emergency room/hospital visits. Clinical pathways, structured care plans that note the essential elements of care by hospital day, have been widely used by hospitals in efforts to improve the quality and efficiency of care. In a second paper, Dy et al. (2005) report that only seven of 26 pathways used at a large academic medical center had the desired impact. They report on a qualitative analysis to describe characteristics that differentiate effective from ineffective pathways. They found thatmany of these care plans were unused. Even among the few that did appear to have an impact, their effectiveness may not have been related to their actual use. Bradley et al. (2005) in another paper surveyed hospital quality management directors about their interactions with QIOs, formerly known as Professional ReviewOrganizations (PROs), that are contracted by the Centers for Medicare & Medicaid Services (CMS) to promote quality of care in the
Choosing the Correct AED: From Animal Studies to the Clinic

Epilepsy is a chronic condition caused by an imbalance of normal excitatory and inhibitory forces in the brain. Antiepileptic drug therapy has been directed primarily toward reducing excitability through blockage of voltage-gated Na+ or Ca2+ channels, or increasing inhibition through enhancement of γ-aminobutyric acid currents. Prior to clinical studies, putative antiepileptic drugs are screened in animals, usually rodents. Maximal electrical shock, pentylenetetrazol, and kindling are typically used as non-mechanistic screens for antiseizure properties and the rotorod test for assessing acute toxicity. While antiseizure drug screening has been successful in bringing drugs to the market and improving our understanding of the pathophysiology of seizures, it should be emphasized that the vast majority of drug screening occurs in mature male rodents and involves models of seizures, not epilepsy. Effective drugs in acute seizures may not be effective in chronic models of epilepsy. Seizure type, clinical and electroencephalographic phenotype, syndrome, and etiology are often quite different in children with epilepsy than adults. Despite these age-related unique features, drugs used in children are generally the same as used in adults. As awareness of the unique features of seizures during development increases, it is anticipated that more drug screening in the immature animal will occur.
Cdc48pNpl4p/Ufd1p binds and segregates heterodimeric membrane anchored/tethered transcription factor complexes via a poly- ubiquitin signal present on the anchors in vitro

Cdc48p is an abundant and conserved member of the AAA ATPase family of molecular chaperones. Cdc48p performs ubiquitin selective functions, which are mediated by numerous ubiquitin-binding adaptors, including the Npl4p-Ufd1p complex. Previous studies suggest that Cdc48p-containing complexes carry-out many biochemical activities, including ubiquitination, deubiquitination, protein complex segregation and targeting of ubiquitinated substrates to the proteasome. The molecular mechanisms by which Cdc48p containing complexes participate in these processes remain poorly defined. We show here using physiologically relevant Cdc48p substrates (i.e. endoplasmic membrane associated/tethered dimers of Mga2p and Spt23p) and in vitro systems with purified proteins that Cdc48pNpl4p/Ufd1p binds to and promotes segregation of the tethered proteins via a poly-ubiquitin signal present on the membrane-bound proteins. Mobilization does not involve retrotranslocation of the associated anchors. These results provide biochemical evidence that Cdc48pNpl4p/Ufd1p functions as a poly-ubiquitin-selective segregase and that a poly-ubiquitin-Cdc48p-pathway modulates protein interactions at cell membranes.
Reliability and Validity of an Algorithm for the Diagnosis of Normal Cognition, MCI and Dementia: Implications for Multi-Center Research Studies

Background—The traditional consensus diagnosis (ConsDx) of normal cognition, mild cognitive impairment (MCI), and dementia relies on the reconciliation of an informant-based report of cognitive and functional impairment by a physician (PhyDx), and a neuropsychological diagnosis (NPDx). As this procedure may be labor-intensive and influenced by the philosophy and biases of a clinician, the diagnostic algorithm (AlgDx) was developed to identify individuals as cognitively normal, with MCI or dementia. Methods—The AlgDx combines the PhyDx with the NPDx, using a computational algorithm that provides cognitive diagnoses, as defined by the National Alzheimer Coordinating Center/Uniform Data Set (NACC/UDS) nomenclature. Reliability of the AlgDx was assessed in 532 communitydwelling elderly subjects by its concordance with the ConsDx, and association with two biomarkers, medial temporal atrophy (MTA) scores of brain MRI scans, and ApoE-ε4 genotype. Results—A high degree of concordance was observed between ConsDx and AlgDx with a weighted Cohen’s kappa of 0.84. Concordance of the AlgDx to the same ConsDx categories ranged from 85% to 92%. Excellent discriminative validity was observed as AlgDx, MTA scores, and APOE-ε4 allele frequencies distinguished amnestic MCI and dementia subjects from normal subjects. Conclusion—The AlgDx of normal cognition, MCI, and dementia is a valid alternative that reduces time, effort and biases associated with the ConsDx. The inherent reliability of a fixed algorithm, together with its efficiency and avoidance of individual bias, suggests the AlgDx may be used in longitudinal, multi-site clinical trials and population studies of MCI and dementia. Address Correspondence To: Ranjan Duara, M.D., Wien Center for Alzheimer’s Disease and Memory Disorders, Mt. Sinai Medical Center, 4300 Alton Rd, Miami Beach, FL 33140, Telephone: 305-674-2543; Fax: 305-532-5241, ranjan-duara@msmc.com. Disclosure: The authors report no conflicts of interest. Statistical analysis was performed by David A. Loewenstein, Ph.D., University of Miami NIH Public Access Author Manuscript Am J Geriatr Psychiatry. Author manuscript; available in PMC 2011 April 1. Published in final edited form as: Am J Geriatr Psychiatry. 2010 April ; 18(4): 363–370. N IH PA Athor M anscript N IH PA Athor M anscript N IH PA Athor M anscript
Two-Year Changes in Health-Related Quality of Life in Gastric Bypass Patients Compared to Two Severely Obese Groups

Background—Few weight loss surgery trials evaluate changes in health-related quality of life (HRQOL) relative to obese individuals not participating in weight loss interventions. Objectives—In a prospective study we evaluated two-year changes in HRQOL in gastric bypass patients compared to two separate severely obese groups not undergoing weight loss intervention. Setting—Bariatric surgery practice. Methods—421 surgery patients (GBP) were compared with 405 individuals who sought but did not have bariatric surgery (No GBP) and 319 population-based obese individuals (Pop OB) on obesity-specific (IWQOL-Lite) and general (SF-36) HRQOL at baseline and two-years. Results—Weight loss was 34.2% for GBP, 1.4% for No GBP and a gain of 0.5% for Pop OB. Both measures of HRQOL showed greater improvements for the GBP group (p<0.001), even after controlling for baseline differences. Effect sizes for changes in physical and weight-related HRQOL were very large for GBP, but small to medium for the two comparison groups. Effect sizes for changes in psychosocial aspects of HRQOL were moderate to very large for the GBP, but small for comparison groups. Ninety-seven percent of surgery patients reported meaningful improvements on IWQOL-Lite total score, versus 43% of the No GBP group and 30% of the Pop OB group (p<0.001). Conclusions—Dramatic improvements occurred in both obesity-specific and physical healthrelated quality of life for gastric bypass surgery patients two-years post-surgery compared to two severely obese groups not enrolled in weight loss intervention. Changes in the psychosocial aspects of HRQOL were medium to large and more variable across domains.
OBSTRUCTIVE SLEEP APNEA (OSA) IS CHARACTER- IZED BY REPETITIVE APNEAS AND HYPOPNEAS DUR- ING SLEEP ASSOCIATED WITH OXYGEN DESATU-

1551 OBSTRUCTIVE SLEEP APNEA (OSA) IS CHARACTERIZED BY REPETITIVE APNEAS AND HYPOPNEAS DURING SLEEP ASSOCIATED WITH OXYGEN DESATURATION, leading to sleep disruption and excessive daytime sleepiness. It has been estimated that OSA is present in 9% to 24%1 of adults, being clinically significant in 2% to 4%,1 and becoming more prevalent as the average population body weight rises.2 Excessive daytime sleepiness related to increased sleep fragmentation and impaired sleep quality clearly has an adverse impact on quality of life and health status.3 Nasal continuous positive airways pressure (CPAP) is considered to be the treatment of choice for OSA. Excessive daytime sleepiness remains the main indication for CPAP treatment in OSA,4 although cardiovascular benefits are thought to result as well. Results from previous studies, including a randomized double-blind controlled parallel trial of therapeutic versus subtherapeutic CPAP showed that CPAP has beneficial effects on excessive daytime sleepiness and on self-reported functioning and well-being, affecting quality of life of OSA patients.4-8 The costeffectiveness of this treatment has also been demonstrated.9 Despite an increasing amount of information on the efficacy of CPAP treatment in OSA patients, data from randomized double-blind controlled trials on treatment outcome measures that included quality of life assessment have been reported in only 2 previous studies.4,7 Particularly, it remains unclear which aspects of symptomatic improvement are the most sensitive to CPAP treatment and carry independent information in predicting response to CPAP. To address this uncertainty we investigated, using data from a randomized double-blind controlled trial, which outcome measures best identify the effect of real CPAP compared to nonspecific placebo effects. The overall aim of this study is to improve the current knowledge about the efficacy of CPAP treatment on symptoms and health status. This issue may be relevant in clinical practice and for future treatment trials.
Modeling of transendothelial transport,1,2

Capillary-tissue exchange of inert hydrophilic solutes in the heart occurs through aqueous channels, the clefts between endothelial cells (ECs). For adenosine (and other vasoactive agents and substrates), there is also transport across the plasmalemma of the ECs. The multiple-indicator dilution technique comparing tracer adenosine flux with that of 9-β-Darabinofuranosylhypoxanthine (an analog that is not transported by the nucleoside carrier) can be used to estimate the conductance of the facilitated transport mechanism, which is equivalent to a permeability-surface area product. Analysis by using a model of exchanges among capillary, EC, interstitium, and myocardial cells suggests that the abluminal surface of the ECs is also highly permeable to adenosine. The inference is that ECs may be an important component of a system for adenosine exchange and regulation in the heart. Now that is it well recognized that endothelial cells (ECs) cannot be regarded as passive cellophane membranes, studies of transcapillary exchange have taken on new interest. We now consider escape from the plasma space across the capillary wall to be accomplished via transport along aqueous pathways (endothelial cell [EC] clefts or possibly chains of linked vesicles traversing the ECs), and across the luminal plasmalemma of the ECs in parallel. For lipid-soluble substances this last route might simply be passive diffusional transport across the lipid bilayer, but for a large variety of hydrophilic solutes, one can expect something more complex. A short list of possibilities includes facilitated or carrier-mediated transport across the luminal endothelial surface, cotransport or counter transport with other substances, and surface binding whether followed or not by engulfment of a surface membrane fragment into the cell. A problem in estimating the rates of entry of substances into ECs is that the cells are small, occupying only about 1.0–1.5% of even a very richly vascularized organ such as the heart. This means that tracer entering ECs will reflux back into capillary plasma rapidly unless there is some mechanism either for retention within the ECs or for its transport out the other side of the ECs across the antiluminal plasmalemma, into the interstitial fluid space, followed by retention in parenchymal cells of the organ. With such small distances and volumes, the rate constants for exchange can be very high, particularly for solutes such as hormones and substrates. The situation therefore requires careful, relatively complete mathematical modeling to evaluate parameters defining rates of exchange. 1From the Symposium Capillary Endothelium: A Metabolic Barrier for Solute Transport presented by The American Physiological Society at the 68th Annual Meeting of the Federation of American Societies for Experimental Biology, St. Louis, Missouri, April 3, 1984. Accepted for publication September 13, 1984. 2Research supported by National Institutes of Health grants HL 19139 and RR 01243. © FASEB NIH Public Access
Activation of aPKCζ Toward TC10 is Regulated by High Fat Diet and Aerobic Exercise in Skeletal Muscle

We determined whether chronic aerobic exercise reverses high-fat diet-induced impairments in the CAP (c-Cbl associated protein)/c-Cbl (Casitas b-lineage lymphoma) signaling cascade in rodent skeletal muscle. Sprague-Dawley rats were placed into either control (CON, n=16) or high-fat fed (n=32) diet groups for 4 wk. During a subsequent 4 wk experimental period 16 high-fat fed rats remained sedentary (HF), 16 high-fat fed rats completed 4 wk of exercise training (HFX), and CON animals were sedentary and remained on the control diet. Following the intervention period, animals were subjected to hind limb perfusions in the presence (n=8/group) or absence (n=8/group) of insulin. In the plasma membrane fractions, neither highfat feeding nor exercise training altered APS (adaptor protein with PH and SH2 domains), c-Cbl or TC10 protein concentrations. In contrast, CAP protein concentration and insulin-stimulated plasma membrane c-Cbl tyrosine phosphorylation were reduced by high-fat feeding, but exercise training reversed these impairments. Of note was that insulinstimulated aPKC ζ (atypical protein kinase C) kinase activity toward TC10 was reduced by high-fat feeding but normalized by exercise training. We conclude that chronic 4 wk of exercise training can reverse high-fat diet-induced impairments on the CAP/c-Cbl pathway in high-fat fed rodent skeletal muscle. We also provide the first evidence that the CAP/c-Cbl insulin signaling cascade in skeletal muscle may directly interact with components of the classical (phosphoinositide 3-kinase dependent) insulin signaling cascade.
TRANSLATIONAL REVIEW Complement System in Lung Disease

In addition to its established contribution to innate immunity, recent studies have suggested novel roles for the complement system in the development of various lung diseases. Several studies have demonstrated that complement may serve as a key link between innate and adaptive immunity in a variety of pulmonary conditions. However, the specific contributions of complement to lung diseases based on innate and adaptive immunity are just beginning to emerge. Elucidating the role of complement-mediated immune regulation in these diseases will help to identify new targets for therapeutic interventions.
Topoisomerase 2β-dependent nuclear DNA damage shapes extracellular growth factor responses via dynamic AKT phosphorylation to control virus latency

The mTOR pathway integrates both extracellular and intracellular signals and serves as a central regulator of cell metabolism, growth, survival and stress responses. Neurotropic viruses, such as herpes simplex virus-1 (HSV-1), also rely on cellular AKT-mTORC1 signaling to achieve viral latency. Here, we define a novel genotoxic response whereby spatially separated signals initiated by extracellular neurotrophic factors and nuclear DNA damage are integrated by the AKTmTORC1 pathway. We demonstrate that endogenous DNA double-strand breaks (DSBs) mediated by Topoisomerase 2β-DNA cleavage complex (TOP2βcc) intermediates are required to achieve AKT-mTORC1 signaling and maintain HSV-1 latency in neurons. Suppression of host DNA repair pathways that remove TOP2βcc trigger HSV-1 reactivation. Moreover, perturbation of AKT phosphorylation dynamics by downregulating the PHLPP1 phosphatase led to AKT mislocalization and disruption of DSB-induced HSV-1 reactivation. Thus, the cellular genome integrity and environmental inputs are consolidated and co-opted by a latent virus to balance lifelong infection with transmission.
Cytoplasmic localization of p27 (CDKN1B / KIP1) in colorectal cancer: inverse correlations with nuclear p27 loss, microsatellite instability and CpG island methylator phenotype (CIMP)

Cytoplasmic mislocalization of p27 (cyclin-dependent kinase inhibitor-1B, CDKN1B/KIP1) is caused by activated AKT1, and has been associated with poor prognosis in various cancers. CpG island methylator phenotype (CIMP) in colorectal cancer is characterized by extensive promoter methylation, and is associated with microsatellite instability-high (MSI-H) and BRAF mutations. We have recently shown a positive correlation between MSI/CIMP and loss of nuclear p27. However, no study has examined cytoplasmic p27 mislocalization in relation to CIMP and MSI in colorectal cancer. Using MethyLight assays, we quantified DNA methylation in eight CIMP-specific gene promoters {CACNA1G, CDKN2A (p16), CRABP1, IGF2, MLH1, NEUROG1, RUNX3 and SOCS1} in 853 colorectal cancer samples obtained from two large prospective cohorts. We assessed expressions of nuclear and cytoplasmic p27 and nuclear p53 by immunohistochemistry. Cytoplasmic p27 expression was inversely associated with loss of nuclear p27 (p<0.0001), CIMP-high (p<0.0001), MSI-H (p<0.0001) and BRAF mutations (p<0.0001). The inverse association of cytoplasmic p27 with CIMP-high (or MSI-H) was independent of MSI (or CIMP) status. In addition, the inverse association of cytoplasmic p27 with CIMP-high was independent of KRAS/BRAF status. BRAF and CDKN2A (p16) methylation were not correlated with cytoplasmic p27 after stratification by CIMP status. The inverse associations of cytoplasmic p27 with MSI-H and CIMP-high were much more pronounced in p53-negative tumors than p53-positive tumors. In conclusion, cytoplasmic p27 expression is inversely associated with MSI-H and CIMP-high, particularly in p53-negative tumors, suggesting interplay of functional losses of p27 and p53 in the development of various molecular subtypes of colorectal cancer.
Recruitment of Tyrosine Phosphatase HCP (SHP-1) by the Killer Cell Inhibitory Receptor

Cytolysis of target cells by natural killer (NK) cells and by some cytotoxic T cells occurs unless prevented by inhibitory receptors that recognize MHC class I on target cells. Human NK cells express a p58 inhibitory receptor specific for HLA-C. We report association of the tyrosine phosphatase HCP with the p58 receptor in NK cells. HCP association was dependent on tyrosine phosphorylation of p58. Phosphotyrosyl peptides corresponding to the p58 tail bound and activated HCP in vitro. Furthermore, introduction of an inactive mutant HCP into an NK cell line prevented the p58-mediated inhibition of target cell lysis. These data imply that the inhibitory function of p58 is dependent on its tyrosine phosphorylation and on recruitment and activation of HCP.
INFECTIOUS ECTROMELIA: EXPERIMENTS ON INTERFERENCE AMND IMNIZATION

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