Miracle-dz/UniFAR-FTUs huggingface dataset:
FoxO Transcription Factors Promote Autophagy in Cardiomyocytes*

In the heart, autophagy is required for normal cardiac function and also has been implicated in cardiovascular disease. FoxO transcription factors promote autophagy in skeletal muscle and have additional roles in regulation of cell size, proliferation, and metabolism. Here we investigate the role of FoxO transcription factors in regulating autophagy and cell size in cardiomyocytes. In cultured rat neonatal cardiomyocytes, glucose deprivation leads to decreased cell size and induction of autophagy pathway genes LC3, Gabarapl1, and Atg12. Likewise, overexpression of either FoxO1 or FoxO3 reduces cardiomyocyte cell size and induces expression of autophagy pathway genes. Moreover, inhibition of FoxO activity by dominant negative FoxO1 (Δ256) blocks cardiomyocyte cell size reduction upon starvation, suggesting the necessity of FoxO function in cardiomyocyte cell size regulation. Under starvation conditions, endogenous FoxO1 and FoxO3 are localized to the nucleus and bind to promoter sequences of Gabarapl1 and Atg12. In vivo studies show that cellular stress, such as starvation or ischemia/reperfusion in mice, results in induction of autophagy in the heart with concomitant dephosphorylation of FoxO, consistent with increased activity of nuclear FoxO transcription factors. Together these results provide evidence for an important role for FoxO1 and FoxO3 in regulating autophagy and cell size in cardiomyocytes.
Dependence of Leucine-rich Repeat Kinase 2 (LRRK2) Kinase Activity on Dimerization*

Dominant missense mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are the most common known genetic cause of Parkinson disease. LRRK2 encodes a serine/threonine protein kinase, and pathogenic mutations may increase kinase activity. Intrinsic GTP binding in the GTPase domain may govern kinase activity through an internal signal transduction cascade. As with many protein kinases, LRRK2 self-interacts through mechanisms that may regulate enzymatic activity. We find that the disruption of either GTPase or kinase activity enhances the formation of high molecular weight oligomers and prevents the formation of LRRK2 dimer structures. In addition, brief application of the broad spectrum kinase inhibitor staurosporine ablates LRRK2 dimers and promotes LRRK2 high molecular weight oligomers. LRRK2 interactions with other proteins in cell lines are kinase-independent and include chaperones and cell cytoskeleton components, suggesting that LRRK2 self-assembly principally dictates complex size. To further explore the mechanics of kinase activation, we separate soluble LRRK2 protein that encodes the pathogenic G2019S mutation into high molecular weight oligomers, dimers, and monomers and find that kinase activity resides with dimeric LRRK2. Some PD-associated mutations that increase kinase activity in vitro significantly increase the proportion of dimer structures relative to total LRRK2 protein, providing additional insight into how pathogenic mutations may alter normal enzymatic regulation. Targeting and tracking LRRK2 dimerization may provide a clear way to observe LRRK2 kinase activity in living cells, and disruption of dimeric LRRK2 through kinase inhibition or other means may attenuate pathogenic increases in LRRK2 enzymatic output.
Urologic diseases in America Project: analytical methods and principal findings.

PURPOSE The burden of urological diseases on the American public is immense in human and financial terms but it has been under studied. We undertook a project, Urologic Diseases in America, to quantify the burden of urological diseases on the American public. MATERIALS AND METHODS We identified public and private data sources that contain population based data on resource utilization by patients with benign and malignant urological conditions. Sources included the Centers for Medicare and Medicaid Services, National Center for Health Statistics, Medical Expenditure Panel Survey, National Health and Nutrition Examination Survey, Department of Veterans Affairs, National Association of Children's Hospitals and Related Institutions, and private data sets maintained by MarketScan Health and Productivity Management (MarketScan, Chichester, United Kingdom), Ingenix (Ingenix, Salt Lake City, Utah) and Center for Health Care Policy and Evaluation. Using diagnosis and procedure codes we described trends in the utilization of urological services. RESULTS In 2000 urinary tract infections accounted for more than 6.8 million office visits and 1.3 million emergency room visits, and 245,000 hospitalizations in women with an annual cost of more than 2.4 billion dollars. Urinary tract infections accounted for more than 1.4 million office visits, 424,000 emergency room visits and 121,000 hospitalizations in men with an annual cost of more than 1 billion dollars. Benign prostatic hyperplasia was the primary diagnosis in more than 4.4 million office visits, 117,000 emergency room visits and 105,000 hospitalizations, accounting for 1.1 billion dollars in expenditures that year. Urolithiasis was the primary diagnosis for almost 2 million office visits, more than 600,000 emergency room visits, and more than 177,000 hospitalizations, totaling more than 2 billion dollars in annual expenditures. Urinary incontinence in women was the primary cause for more than 1.1 million office visits in 2000 and 452 million dollars in aggregate primary cause for more than 1.1 million office visits in 2000 and 452 million dollars in aggregate annual expenditures. Other manuscripts in this series present further detail for specific urologic conditions. CONCLUSIONS Recent trends in epidemiology, practice patterns, resource utilization and costs for urological diseases have broad implications for quality of health care, access to care and the equitable allocation of scarce resources for clinical care and research.
Peptidoglycan Induces Cyclooxygenase-2 Expression in Macrophages by Activating the Neutral Sphingomyelinase-Ceramide Pathway*

The sphingomyelin signal transduction pathway is known to play a role in mediating the action of various cytokines. Herein, we examined the role of neutral sphingomyelinase (nSMase)/ceramide in peptidoglycan (PGN)-induced NF-κB activation and cyclooxygenase-2 (COX-2) expression in macrophages. PGN-induced COX-2 expression was attenuated by an nSMase inhibitor (3-O-methyl-sphingomyeline, 3-OMS) and ceramidase, but not by an acidic SMase inhibitor (imipramine). C2-ceramide, bacterial SMase (which mimics cellular SMase activity), and a ceramidase inhibitor (N-oleoyl-ethanolamine) individually had no effect on COX-2 expression; however, they markedly enhanced PGN-induced COX-2 expression. PGN activated nSMase, but not acidic SMase, resulting in increased ceramide generation. PGN-induced nSMase activation and ceramide formation were inhibited by 3-OMS, but not by imipramine. PGN-induced COX-2 expression was inhibited by a p38 MAPK inhibitor (SB 203580) and dominant negative mutants of MAPK kinase (MKK) 3, MKK6, and p38 MAPKα. 3-OMS selectively inhibited PGN-induced p38 MAPK and MKK3/6 activation, but not JNK or ERK1/2. C2-ceramide, bacterial SMase, and N-oleoyl-ethanolamine all induced p38 MAPK or MKK3/6 activation. The PGN-mediated increases in κB-luciferase activity were also inhibited by 3-OMS and the p38 MAPKαDN, but not by imipramine. Furthermore, C2-ceramide caused an increase in κB-luciferase activity. Our data demonstrate for the first time that PGN activates the nSMase/ceramide pathway to induce MKK3/6/p38 MAPK activation, which in turn initiates NF-κB activation and ultimately induces COX-2 expression in macrophages. The nSMase/ceramide pathway is required but might not be sufficient for COX-2 expression induced by PGN.
DRAGON, a GPI‐anchored membrane protein, inhibits BMP signaling in C2C12 myoblasts

Bone morphogenetic proteins (BMPs) induce osteoblastic differentiation of myoblasts via binding to cell surface receptors. Repulsive guidance molecules (RGMs) have been identified as BMP co‐receptors. We report here that DRAGON/RGMb, a member of the RGM family, suppressed BMP signaling in C2C12 myoblasts via a novel mechanism. All RGMs were expressed in C2C12 cells that were differentiated into myocytes and osteoblastic cells, but RGMc was not detected in immature cells. In C2C12 cells, only DRAGON suppressed ALP and Id1 promoter activities induced by BMP‐4 or by constitutively activated BMP type I receptors. This inhibition by DRAGON was dependent on the secretory form of the von Willbrand factor type D domain. DRAGON even suppressed BMP signaling induced by constitutively activated Smad1. Over‐expression of neogenin did not alter the inhibitory capacity of DRAGON. Taken together, these findings indicate that DRAGON may be an inhibitor of BMP signaling in C2C12 myoblasts. We also suggest that a novel molecule(s) expressed on the cell membrane may mediate the signal transduction of DRAGON in order to suppress BMP signaling in C2C12 myoblasts.
P-glycoprotein inhibition as a strategy to increase drug delivery across the blood-brain barrier: focus on antidepressants

XIII CHAPTER 1: GENERAL INTRODUCTION . 1 1.1. CENTRAL NERVOUS SYSTEM BARRIERS . 2 1.2. THE BLOOD-BRAIN BARRIER: A MAJOR OBSTACLE IN THE DELIVERY OF DRUGS TO THE BRAIN . 4 1.2. 1. Structure and function of the BBB . 5 1.2. 2. Routes of transport across the BBB . 9 1.2. 3. Drug efflux transporters at the BBB . 13 1.2. 4. Approaches to overcome the BBB to achieve drug delivery to the brain . 14 1.2. 4.1. Invasive approaches . 15 1.2. 4.2. Non-invasive approaches . 15 1.2. 5. Methods to predict or measure drug transport across the BBB . 16 1.2. 5.1. In silico . 16 1.2. 5.2. In vitro. 18 1.2. 5.3. In vivo and in situ approaches . 22 1.2. 5.4. Intracerebral microdialysis . 24 1.3. P-GLYCOPROTEIN . 30 1.3.1. Structure and function of P-glycoprotein . 31 1.3. 2. P-gp substrates and inhibitors . 32 1.3. 3. Regulation of P-gp expression and activity . 35 1.3. 4. Single nucleotide polymorphisms in ABCB1 . 36 1.3. 5. The contribution of P-gp efflux to treatment failure . 38 1.3. 6. P-gp inhibition as a strategy to augment the therapeutic effects of P-gp substrate drugs 40 1.3. 7. Methods to assess drug interactions with P-gp at the BBB . 41 1.3. 7.1. In silico approaches . 41 1.3. 7.2. In vitro studies . 42 1.3. 7.3. In vivo studies . 45 1.3. 7.4. Human studies . 45 1.4. DEPRESSION . 46 1.4.1. Neurobiology of depression . 47 1.4.2. Treatment of depression . 50 1.4. 3. Side-effects of antidepressant drugs . 54 1.4. 3.1. Serotonin syndrome . 55
Probing HIV-1 Membrane Liquid Order by Laurdan Staining Reveals Producer Cell-dependent Differences*

Viruses acquire their envelope by budding from a host cell membrane, but viral lipid composition may differ from that of the budding membrane. We have previously reported that the HIV-1 membrane is highly enriched in cholesterol, sphingolipids, and other raft lipids, suggesting that the virus may bud from pre-existing or virus-induced lipid rafts. Here, we employed the environmentally sensitive fluorescent dye Laurdan to study the membrane lateral structure of HIV-1 derived from different cell lines. Differences in viral membrane order detected by Laurdan staining were shown by mass spectrometry to be due to differences in lipid composition. Isogenic viruses from two different cell lines were both strongly enriched in raft lipids and displayed a liquid-ordered membrane, but these effects were significantly more pronounced for HIV-1 from the T-cell line MT-4 compared with virus from 293T cells. Host-dependent differences in the lipidomes predominantly affected the ratio of sphingomyelins (including dihydrosphingomyelin) to phosphatidylcholine, whereas cholesterol contents were similar. Accordingly, treatment of infectious HIV-1 with the sphingomyelin-binding toxins Equinatoxin-II or lysenin showed differential inhibition of infectivity. Liposomes consisting of lipids that had been extracted from viral particles exhibited slightly less liquid order than the respective viral membranes, which is likely to be due to absence of membrane proteins and to loss of lipid asymmetry. Synthetic liposomes consisting of a quaternary lipid mixture emulating the viral lipids showed a liquid order similar to liposomes derived from virion lipids. Thus, Laurdan staining represents a rapid and quantitative method to probe viral membrane liquid order and may prove useful in the search for lipid active drugs.
Effective management of accelerated phase myelofibrosis with low‐dose splenic radiotherapy

The main cause of hepatosplenomegaly in primary (PMF), post polycythemia vera (post-PV MF), and post essential thrombocythemia (post-ET MF) myelofibrosis (MF) is extramedullary hematopoiesis (EMH). Drug-refractory symptomatic splenomegaly in MF is usually managed by splenectomy or involved-field radiotherapy. The latter is most effective in the treatment of MF-associated bone pain and pulmonary hypertension.Our previous experience with hepatosplenic radiotherapy in MF showed efficacy in the majority of treated patients but its utility was limited by the transient nature of its benefit and the occurrence of treatment-related pancytopenia. In an effort to address these issues,we have adopted an induction-maintenance treatment strategy that utilizes lower radiation doses-induction with 100 cGy total in four daily doses of 25 cGy and maintenance with either the same or lower intensity regimen. Herein, we report our most recent experience using this treatment plan in two cases, who in addition to their expected response from the standpoint of splenomegaly, also unexpectedly showed a marked response of their underlying accelerated phase disease,including clearance of circulating blasts and basophilia.
Biosimilar epoetins and other “follow‐on” biologics: Update on the European experiences

After the patents of biopharmaceuticals have expired, based on specific regulatory approval pathways copied products (“biosimilars” or “follow‐on biologics”) have been launched in the EU. This article summarizes experiences with hematopoietic medicines, namely the epoetins (two biosimilars traded under five different brand names) and the filgrastims (two biosimilars, six brand names). Physicians and pharmacists should be familiar with the legal and pharmacological specialities of biosimilars: The production process can differ from that of the original, clinical indications can be extrapolated, glycoproteins contain varying isoforms, the formulation may differ from the original, and biopharmaceuticals are potentially immunogenic. Only on proof of quality, efficacy and safety, biosimilars are a viable option because of their lower costs. Am. J. Hematol. 2010. © 2010 Wiley‐Liss, Inc.
Assessing the performance of the Slovak and the Czech health systems : a case study examining the double transition and beyond

Measuring health system performance is essential for improving health and quality of care. It is relevant in any context, but especially in countries whose health care systems have undergone major changes. The 1989 transition from communism to democracy in Czechoslovakia followed by the 1993 split into two independent countries (the Czech Republic and Slovakia) have been studied extensively but little research has addressed the effects of these events on health and the quality of care provided. The overarching objective of this thesis is to examine pre- and post-transition health system performance at three levels: i) overall health and well-being, ii) quality of the health care system, and iii) quality of outpatient care. This is a policy piece intended to demonstrate the usefulness of various performance indicators, while applying a range of quantitative methods from different disciplines to unique datasets. The macro level findings suggest that the transition was not detrimental to overall health and wellbeing in neither of the two countries as demonstrated by a small continued height increase. Slovakia showed a larger capacity to benefit from the transition. The overall quality of the health care systems is measured by ‘avoidable’ mortality and also shows improvements. For some ‘avoidable’ mortality conditions Slovakia continues to lag behind the Czech Republic, while for others it outperforms its neighbour. The thesis also provides evidence on the absence of a significant relationship between health care inputs and ‘avoidable’ mortality. Finally, the assessment of the quality of outpatient care in Slovakia, using preventable hospitalisations and selected processes of care, shows that inappropriate care may be provided for asthma and diabetes. The findings also indicate a link between appropriate and inappropriate care and preventable hospitalisations. Overall, the results of this thesis provide the basis for policy makers to better understand the changes in health outcomes and quality of care in these two settings but also to inform future quality improvement efforts.
Heterodimerization of the Sialidase NEU1 with the Chaperone Protective Protein/Cathepsin A Prevents Its Premature Oligomerization*

Lysosomal neuraminidase-1 (NEU1) forms a multienzyme complex with β-galactosidase and protective protein/cathepsin A (PPCA). Because of its association with PPCA, which acts as a molecular chaperone, NEU1 is transported to the lysosomal compartment, catalytically activated, and stabilized. However, the mode(s) of association between these two proteins both en route to the lysosome and in the multienzyme complex has remained elusive. Here, we have analyzed the hydrodynamic properties of PPCA, NEU1, and a complex of the two proteins and identified multiple binding sites on both proteins. One of these sites on NEU1 that is involved in binding to PPCA can also bind to other NEU1 molecules, albeit with lower affinity. Therefore, in the absence of PPCA, as in the lysosomal storage disease galactosialidosis, NEU1 self-associates into chain-like oligomers. Binding of PPCA can reverse self-association of NEU1 by causing the disassembly of NEU1-oligomers and the formation of a PPCA-NEU1 heterodimeric complex. The identification of binding sites between the two proteins allowed us to create innovative structural models of the NEU1 oligomer and the PPCA-NEU1 heterodimeric complex. The proposed mechanism of interaction between NEU1 and its accessory protein PPCA provides a rationale for the secondary deficiency of NEU1 in galactosialidosis.
Genome‐wide DNA methylation profile of tissue‐dependent and differentially methylated regions (T‐DMRs) residing in mouse pluripotent stem cells

DNA methylation profile, consisting of tissue‐dependent and differentially methylated regions (T‐DMRs), has elucidated tissue‐specific gene function in mouse tissues. Here, we identified and profiled thousands of T‐DMRs in embryonic stem cells (ESCs), embryonic germ cells (EGCs) and induced pluripotent stem cells (iPSCs). T‐DMRs of ESCs compared with somatic tissues well illustrated gene function of ESCs, by hypomethylation at genes associated with CpG islands and nuclear events including transcriptional regulation network of ESCs, and by hypermethylation at genes for tissue‐specific function. These T‐DMRs in EGCs and iPSCs showed DNA methylation similar to ESCs. iPSCs, however, showed hypomethylation at a considerable number of T‐DMRs that were hypermethylated in ESCs, suggesting existence of traceable progenitor epigenetic information. Thus, DNA methylation profile of T‐DMRs contributes to the mechanism of pluripotency, and can be a feasible solution for identification and evaluation of the pluripotent cells.
Classical Hodgkin's lymphoma arising in different host's conditions: Pathobiology parameters, therapeutic options, and outcome

Epidemiologic and molecular findings suggest that classical Hodgkin's lymphoma (CHL) is not a single disease but consists of more than one entity and may occur in different clinical settings. This review analyzes similarities and disparities among CHL entities arising in different host's conditions with respect to pathobiology parameters, therapeutic options, and outcome. For the purpose of this analysis, CHL entities have been subdivided according to the immune status of the host. In nonimmunosuppressed hosts, according to the age, CHL include pediatric, adult, and elderly forms, whereas, in immunosuppressed hosts, according to the type of immunosuppression, CHL include human immunodeficiency virus (HIV)‐associated, iatrogenic, and post‐transplant types. CHL entities in different settings are similar in morphology of neoplastic cells, expression of activation markers, and aberrations/activation of NFKB, JAK/STAT, and P13K/AKT pathways, but differ in the association with Epstein‐Barr virus (EBV) infection, persistent B‐cell phenotype, and cellular background composition. Large B‐cell lymphomas resembling CHL may also be observed in the same clinical settings. These lesions, however, do not fulfill the diagnostic criteria of CHL and clinically display a very aggressive behavior. In this article, current treatment options for the CHL entities, especially for elderly CHL and HIV‐associated CHL, are specifically reviewed. ABVD remains the gold standard both in nonimmunosuppressed or immunosuppressed hosts even if there are several data suggesting a possible improvement in outcome using the aggressive BEACOPP regimen in advanced stages. Refractory CHL, a clinical condition that may occur throughout the entire spectrum of CHL, is discussed separately. Am. J. Hematol. , 2011. © 2010 Wiley‐Liss, Inc.
Transcriptional regulation of the CADM1 gene by retinoic acid during the neural differentiation of murine embryonal carcinoma P19 cells

CADM1 is a multifunctional cell adhesion molecule expressed predominantly in the nerve system, testis and lung. The expression of the Cadm1 gene is induced during the neural differentiation of murine embryonal carcinoma P19 cells by treatment with retinoic acid (RA). Here, we show that the suppression of CADM1 expression using RNAi interfered with P19 cell aggregation and reduced cell populations expressing MAP2 after RA treatment. Nonaggregated P19 cells were not differentiated into neurons, suggesting that CADM1 participates in the aggregate formation and neuronal differentiation of P19 in vitro. A luciferase assay of a series of deletion mutants of the CADM1 promoter localized an RA‐responsive cis‐acting element to an approximately 90‐bp fragment upstream of the translational start site. This element contains a putative binding site for transcription factor Sp1, named Sp1‐binding site‐1 (Sp1BS‐1). Sp1BS‐1 and adjacent Sp1‐binding sites (Sp1BS‐2 and Sp1BS‐3) showed enhanced transcriptional activity by RA. Moreover, a chromatin immunoprecipitation showed that RA receptor (RAR)α was associated with a DNA fragment containing Sp1BS‐1, whereas suppression of RARα expression using siRNA reduced the responsiveness of the CADM1 promoter to RA. These results suggest that Sp1 plays a critical role in RA‐induced CADM1 expression through possible interaction with RARα in the neural differentiation of P19.