Prevalence and characteristics of hepatitis delta virus infection in patients with hepatitis B in the United States: An analysis of the All-Payer Claims Database.

BACKGROUND AND AIMS: HDV leads to the most severe form of viral hepatitis; however, the prevalence of HDV is not well understood. Using real-world data from the All-Payer Claims Database, this study estimates the prevalence of HBV/HDV infection among the chronic HBV population and describes patient/clinical characteristics for adults with HBV/HDV infection in the United States. APPROACH AND RESULTS: Adults (≥18 years) with ≥1 inpatient claim or ≥2 outpatient claims for HDV infection or HBV in the All-Payer Claims Database from January 1, 2014, to December 31, 2020, were identified. HDV prevalence was calculated as the proportion of patients with HBV/HDV infection among total patients with HBV infection. Patient characteristics, socioeconomic status, advanced liver complications (eg, cirrhosis, HCC), and comorbidities were assessed. A total of 6719 patients were diagnosed with HBV/HDV among 144,975 with HBV and 12 months of continuous data, for a prevalence of 4.6%. At diagnosis, 31.7% of patients with HBV/HDV had advanced liver complications, including compensated cirrhosis (16.3%) and decompensated cirrhosis (10.4%). Diabetes (50.5%), hypertension (49.8%), and HIV infection (30.9%) were the top 3 comorbidities. CONCLUSIONS: In a large database capturing approximately 80% of the US-insured population, HBV/HDV infection prevalence was 4.6% among adults infected with HBV. Patients infected with HDV had high rates of baseline liver complications and other comorbidities at the time of diagnosis, suggesting potentially delayed diagnosis and/or treatment. Earlier identification of HBV/HDV infection among the population with HBV may provide opportunities to improve linkage to care and treatment, thereby reducing the risk of liver-related morbidity and mortality.

Association of hepatitis delta virus with liver morbidity and mortality: A systematic literature review and meta-analysis.

BACKGROUND AND AIMS: Studies have suggested that patients with chronic hepatitis B, either co- or superinfected, have more aggressive liver disease progression than those with the HDV. This systematic literature review and meta-analysis examined whether HDV RNA status is associated with increased risk of advanced liver disease events in patients who are HBsAg and HDV antibody positive. APPROACH AND RESULTS: A total of 12 publications were included. Relative rates of progression to advanced liver disease event for HDV RNA+/detectable versus HDV RNA-/undetectable were extracted for analysis. Reported OR and HRs with 95% CI were pooled using the Hartung-Knapp-Sidik-Jonkman method for random-effects models. The presence of HDV RNA+ was associated with an increased risk of any advanced liver disease event [random effect (95% CI): risk ratio: 1.48 (0.93, 2.33); HR: 2.62 (1.55, 4.44)]. When compared to the patients with HDV RNA- status, HDV RNA+ was associated with a significantly higher risk of progressing to compensated cirrhosis [risk ratio: 1.74 (1.24, 2.45)] decompensated cirrhosis [HR: 3.82 (1.60, 9.10)], HCC [HR: 2.97 (1.87, 4.70)], liver transplantation [HR: 7.07 (1.61, 30.99)], and liver-related mortality [HR: 3.78 (2.18, 6.56)]. CONCLUSIONS: The patients with HDV RNA+ status have a significantly greater risk of liver disease progression than the patients who are HDV RNA-. These findings highlight the need for improved HDV screening and linkage to treatment to reduce the risk of liver-related morbidity and mortality.

A systematic literature review and meta-analysis of primary evidence reporting health state preference values in chronic Hepatitis B, C, and D.

OBJECTIVES: Chronic viral hepatitis is associated with severe impairment and reduction in patient health-related quality of life (HRQoL), due to substantial morbidity associated with advanced liver disease. The aim of this study was to identify and synthesize utilities for chronic hepatitis B (cHBV), C (cHCV), and D (cHDV) through a systematic literature review (SLR) and meta-analyses. METHODS: Electronic databases were searched from inception to May 2023 to identify primary studies reporting health state utilities in English in patients aged 18 years and over, with cHBV, cHCV or cHDV in the United States, the United Kingdom, Europe, Canada, Australia or New Zealand. Meta-analyses were conducted for studies reporting a measure of uncertainty; model selection (fixed and random) was based on the observed levels of heterogeneity among studies. RESULTS: A total of 24 studies met the inclusion criteria and were included in the meta-analyses. More studies meeting the inclusion criteria reported utilities for cHCV (n=20) than for cHBV (n=8); no studies reported utility values for cHDV. While for any given health state, mean utilities were higher for cHBV than for cHCV, utilities decreased with disease progression towards cirrhosis health states. Meta-analyses in cHCV found a utility decrement of 0.1 and 0.03, based on progression from non-cirrhosis to compensated cirrhosis and for decompensation in established cirrhosis, respectively. CONCLUSIONS: Chronic viral hepatitis is associated with considerable impairment in HRQoL. Despite our findings, there is a need for more evidence on the lived experience in patients living with chronic heptatitis, notably in cHBV and cHDV.

Patient experiences in ulcerative colitis: conceptual model and review of patient-reported outcome measures.

PURPOSE: To identify symptoms and their impacts on daily functioning and health-related quality of life (HRQoL) experienced by adult patients with ulcerative colitis (UC) and evaluate patient-reported outcome (PRO) measures for UC clinical studies. METHODS: A conceptual model of symptoms and impacts of UC were developed from a literature review. PRO measures were identified from the literature, clinical trials databases, health technology assessment submissions, and regulatory label claims, and were selected for conceptual analysis based on disease specificity and use across information sources. PRO measures covering the most concepts when mapped against the conceptual model were assessed for gaps in psychometric properties using Food and Drug Administration (FDA) guidance and consensus-based standards for the selection of health measurement instruments (COSMIN) criteria. RESULTS: The conceptual model grouped the 52 symptom concepts and 72 proximal and distal impacts into eight, two, and five dimensions, respectively. Of 65 PRO measures identified, eight underwent conceptual analysis. Measures covering the most concepts and assessed for psychometric properties were the Inflammatory Bowel Disease Questionnaire, Symptoms and Impacts Questionnaire for UC, UC-PRO symptoms modules, UC-PRO impact modules, and Crohn's and UC Questionnaire; all had good or excellent support for content validity. The UC-PRO Signs and Symptoms fully met FDA guidance and COSMIN criteria for content validity and most psychometric properties. CONCLUSION: Existing PRO measures assess concepts relevant to patients with UC, but all PRO measures reviewed require further psychometric evaluation to demonstrate they are fit for purpose.

Characteristics associated with receipt of treatment among patients diagnosed with chronic hepatitis C virus.

Although current guidelines recommend that nearly all patients with chronic hepatitis C virus (HCV) infection receive treatment, a substantial proportion remain untreated. We conducted an administrative claims analysis to provide real-world data on treatment patterns and characteristics of treated versus untreated patients among individuals with HCV in the United States. Adults with an HCV diagnosis from 01 July 2016 through 30 September 2020 and continuous health plan enrolment for 12 months before and ≥1 month after the diagnosis date were identified in the Optum Research Database. Descriptive and multivariable analyses were conducted to evaluate the association between patient characteristics and the rate of treatment. Of 24,374 patients identified with HCV, only 30% initiated treatment during follow-up. Factors associated with increased rate of treatment included younger age versus age 75+ (hazard ratio [HR] 1.50-1.83 depending on age group), commercial versus Medicare insurance (HR 1.32), and diagnosis by a specialist versus a primary care physician (HR 2.56 and 2.62 for gastroenterology and infectious disease or hepatology, respectively) (p < .01 for all). Several baseline comorbidities were associated with decreased rate of treatment, including psychiatric disorders (HR 0.87), drug use disorders (HR 0.85) and cirrhosis (HR 0.42) (p < .01 for all). These findings highlight existing HCV treatment inequities, particularly among older patients and those with psychiatric disorders, substance use disorders or chronic comorbidities. Targeted efforts to increase treatment uptake in these populations could mitigate a considerable future burden of HCV-related morbidity, mortality and healthcare costs.

Stem Cells in the Mammalian Gonads.

Besides spermatogonial stem cells (SSCs) and ovarian stem cells (OSCs), a novel population of pluripotent stem cells termed very small embryonic-like stem cells (VSELs) has been reported in both adult mouse and human testes and ovaries. VSELs and SSCs/OSCs are developmentally linked to each other. VSELs are relatively quiescent, small-sized stem cells that undergo asymmetrical cell divisions (ACDs) whereby they self-renew and give rise to the slightly bigger SSCs/OSCs which in turn undergo symmetrical cell divisions (SCDs) and clonal expansion to form germ cell chains/nests before further differentiation into gametes. Comparison of VSELs and SSCs/OSCs for their potential to differentiate into sperm/oocytes is irrelevant since VSELs only undergo ACD to give rise to SSCs/OSCs that further differentiate into gametes. Being relatively quiescent, VSELs survive oncotherapy and can be manipulated to regenerate nonfunctional gonads of cancer survivors, and thus there is possibly no need to bank testicular/ovarian tissue prior to oncotherapy. Being developmentally linked to the primordial germ cells (PGCs) which are the natural precursors to the gametes, VSELs differentiate into haploid sperm/oocyte-like structures in vitro when cultured on appropriate feeder support, in the absence of a cocktail of growth factors. VSELs express receptors for pituitary and sex hormones (FSHR, ER) and thus get directly stimulated/affected by their circulating levels. Excessive self-renewal of VSELs in the gonads may initiate testicular and ovarian cancers. To conclude, VSELs can be targeted to regenerate the gonads of patients with gonadal insufficiency including cancer survivors and are excellent candidates to differentiate into gametes in vitro.

High strength distillery wastewater treatment by a PAC-MBR with low PAC dosage.

Augmentation of membrane bioreactors (MBRs) with activated carbon is established to offer several operational advantages. This work investigates the influence of low dosing (2 g/L) of powdered activated carbons (PACs) with different characteristics on the performance of MBR treating high strength molasses distillery wastewater containing difficult-to-biodegrade recalcitrant components. Two MBRs, augmented with different PACs, were operated in parallel over a period of 240 days and their performance monitored in terms of biomass growth, reduction in chemical oxygen demand (COD), sludge properties like extracellular polymeric substances content, filterability, and morphology. Removal of organics and coloring matter by adsorption, biodegradation and membrane filtration was estimated. Although adsorptive removal of color and COD is influenced by the properties of the PAC used, the performance of the PAC-MBRs was independent of PAC properties. Both PACs preferentially adsorbed the low molecular weight components in distillery wastewater. Retention by the membrane filter with the secondary cake layer contributed to reduction in color and COD of treated effluent. The findings indicate that low dosing with PAC adsorbing low molecular weight organics has a limited role in PAC-MBR treating distillery wastewater.

Integrated treatment of molasses distillery wastewater using microfiltration (MF).

To achieve zero-liquid discharge, high pressure reverse osmosis (RO) of effluent is being employed by molasses based alcohol distilleries. Low pressure and thus less energy intensive microfiltration (MF) is well established for particulate separation but is not suitable for removal of dissolved organics and color. This work investigates two schemes incorporating MF for molasses distillery wastewater (a) chemical coagulation followed by treatment in a membrane bioreactor (MBR) using MF and (b) electrocoagulation followed by MF. The performance was assessed in terms of COD and color reduction; the conversion of the generated sludge into a zeolite desiccant was also examined. A comparison of the schemes indicates electrocoagulation followed by MF through a 0.1 µm membrane to be most effective. By hydrothermal treatment, electrocoagulated sludge can be transformed into a porous NaX zeolite with a surface area of 86 m(2)/g, which is comparable to commercial desiccants.

The Efficiency of Increased HCV Testing and Treatment Strategies in Spain to Achieve Elimination Goals.

BACKGROUND: In 2015, Spain launched a national eradication strategy for hepatitis C virus (HCV), resulting in the highest treatment rate in Europe and substantial reductions in HCV prevalence. However, to achieve the goal of HCV elimination, it is necessary to scale-up the diagnosis, treatment, and management of HCV infection. OBJECTIVE: Our aim was to assess the prevalence, incidence, and cost effectiveness of scaling-up compared with status quo scenarios. METHODS: A compartmental dynamic transmission model was developed comprising of a cascade of care and a liver progression module. Cost and quality-of-life inputs were sourced from the literature. Key outcomes were the prevalence and incidence of HCV and the incremental cost per quality-adjusted life-year (QALY) and per life-year (LY). Outcomes for a hypothetical elimination strategy were compared with the status quo. RESULTS: The base-case analysis found that scaling-up testing and treatment reduced both the prevalence and incidence of HCV over time, resulting in incremental costs per QALY and LY of €13,291 and €12,285 respectively, compared with the status quo. The main drivers of the cost-effectiveness results included cost of diagnosis, cost of treatment, proportion of people who are unaware, percentage of population who inject drugs, and calibration parameters related to HCV infection prevalence. CONCLUSIONS: This analysis demonstrated that scaling-up testing and treatment with direct-acting antivirals may be an efficient strategy for reducing the incidence and prevalence of HCV and may help achieve HCV elimination goals in Spain.

Understanding the Natural History of Chronic Hepatitis D: Proposal of a Model for Cost-Effectiveness Studies.

BACKGROUND: As new therapeutic options become available, better understanding the potential impact of emerging therapies on clinical outcomes of hepatits D virus (HDV) is critical. OBJECTIVE: The aim of this study was to develop a natural history model for patients with hepatitis D virus. METHODS: We developed a model (decision tree followed by a Markov cohort model) in adults with chronic HDV infection to assess the natural history and impact of novel treatments on disease progression versus best supportive care (BSC). The model time horizon was over a lifetime (up to 100 years of age); state transitions and health states were defined by responder status. Patients in fibrosis stages 0 through 4 received treatment; decompensated patients were not treated. Response was defined as the combined response endpoint of achievement of HDV-RNA undetectability/≥2-log10 decline and alanine aminotransferase normalization; response rates of 50% and 75% were explored. Health events associated with advanced liver disease were modeled as the number of events per 10,000 patients. Scenario analyses of early treatment, alternate treatment response, and no fibrosis regression for treatment responders were also explored. RESULTS: The model was able to reflect disease progression similarly to published natural history studies for patients with HBV/HDV infection. In a hypothetical cohort of patients reflecting a population enrolled in a recent clinical trial, fewer advanced liver disease events were observed with a novel HDV treatment versus BSC. Fewer liver-related deaths were observed under 50% and 75% response (900 and 1,358 fewer deaths, respectively, per 10,000 patients). Scenario analyses showed consistently fewer advanced liver disease events with HDV treatment compared with BSC, with greater reductions observed with earlier treatment. CONCLUSION: This HDV disease progression model replicated findings from natural history studies. Furthermore, it found that a hypothetical HDV treatment results in better clinical outcomes for patients versus BSC, with greater benefit observed when starting treatment early. This validated natural history model for HBV/HDV infection can serve as a foundation for future clinical and economic analyses of novel HDV treatments that can support healthcare stakeholders in the management of patients with chronic HDV.

Preventing/Reversing Adverse Effects of Endocrine Disruption on Mouse Testes by Normalizing Tissue Resident VSELs.

Reproductive health of men is declining in today's world due to increased developmental exposure to endocrine-disrupting chemicals (EDCs). We earlier reported that neonatal exposure to endocrine disruption resulted in reduced numbers of seminiferous tubules in Stage VIII, decreased sperm count, and infertility along with testicular tumors in 65% of diethylstilbestrol (DES) treated mice. Epigenetic changes due to EDCs, pushed the VSELs out of a quiescent state to enter cell cycle and undergo excessive self-renewal while transition of c-KIT- stem cells into c-KIT + germ cells was blocked due to altered MMR axis (Np95, Pcna, Dnmts), global hypomethylation (reduced expression of 5-methylcytosine) and loss of imprinting at Igf2-H19 and Dlk1-Meg3 loci. The present study was undertaken to firstly show similar defects in FACS sorted VSELs from DES treated testis and to further explore the reversal of these testicular pathologies by (i) oral administration of XAR (a nano-formulation of resveratrol) or (ii) inter-tubular transplantation of mesenchymal stromal cells (MSCs). Similar defects as reported earlier in the testes were evident, based on RNAseq data, on FACS sorted VSELs from DES treated mice. Both strategies were found effective, improved spermatogenesis, increased number of tubules in Stage VIII, normalized numbers of VSELs and c-KIT + cells, improved epigenetic status of VSELs to restore quiescent state, and reduced cancer incidence from 65% after DES to 13.33% and 20% after XAR treatment or MSCs transplantation respectively. Results provide a basis for initiating clinical studies and the study falls under the umbrella of United Nations Sustainable Development Goal 3 to ensure healthy lives and well-being for all of all ages.

Real-world value of direct-acting antivirals for hepatitis C at Kaiser Permanente Southern California.

OBJECTIVES: Direct-acting antivirals (DAAs) for chronic hepatitis C virus (HCV) lead to cure in more than 95% of recipients; however, payers may limit access to these lifesaving drugs due to high initial cost. Here, the cost-effectiveness of treating HCV with DAAs vs no treatment over a lifetime horizon is evaluated from the perspective of Kaiser Permanente Southern California (KPSC). STUDY DESIGN: A hybrid decision-tree Markov model. METHODS: The model simulated the health and economic outcomes for a real cohort of patients with HCV treated with either ledipasvir-sofosbuvir or sofosbuvir-velpatasvir between November 1, 2014, and October 31, 2019, at KPSC. Patients entered the model at different stages of liver disease and received either active treatment with DAAs or no treatment. Patients who did not achieve sustained virological response experienced disease progression; those who achieved sustained virological response experienced either significantly slower or no disease progression depending on the stage of fibrosis at model start. Demographics, treatment experience, genotype, baseline fibrosis stage, treatment rates, and treatment efficacy were sourced from KPSC real-world data. Costs and utilities were sourced from published literature. RESULTS: A total of 7255 patients with a mean age of 59 years were treated during the study period. Over a lifetime horizon, DAAs resulted in significant reduction in advanced liver disease events and a total cost savings of $1 billion compared with no treatment based on a hybrid decision-tree Markov state-transition model. Cost savings were achieved after only 3 years. DAA intervention dominated no treatment on a per-patient and cohort basis. CONCLUSIONS: DAA treatment at KPSC is predicted to significantly reduce HCV-related morbidity and mortality, providing an anticipated return on investment in drug costs after 3 years of treatment.

Testicular cancer in mice: interplay between stem cells and endocrine insults.

BACKGROUND: Incidence of type II germ cell tumors (T2GCT) has increased in young men possibly due to fetal/perinatal exposure to estrogenic compounds. Three-fold increased incidence of T2GCT was reported in men exposed in utero to diethylstilbestrol (DES). T2GCT is a development-related disease arising due to blocked differentiation of gonocytes into spermatogonia in fetal testes which survive as germ cell neoplasia in situ (GCNIS) and initiate T2GCT. In our earlier study, T2GCT-like features were observed in 9 out of 10 adult, 100-day-old mice testes upon neonatal exposure to DES (2 µg/pup/day on days 1-5). Neonatal DES exposure affected testicular very small embryonic-like stem cells (VSELs) and spermatogonial stem cells and resulted in infertility, reduced sperm counts and tumor-like changes leading to our postulate that testicular dysgenesis syndrome possibly has a stem cell basis. The present study was undertaken to further characterize testicular tumor in mice testes. METHODS: DES-exposed mice pups (n = 70) were studied on D100 and after 12 months to understand how T2GCT progresses. Besides histological studies, a carefully selected panel of markers were studied by immuno-fluorescence and qRT-PCR. RESULTS: DES resulted in either atrophied or highly vascularized, big-sized testes and extra-testicular growth was also observed. GCNIS-like cells with big, vacuolated cytoplasm and increased expression of OCT-4, SSEA-1, SCA-1 and CD166 (cancer stem cells marker) along with reduced c-KIT, MVH and PTEN were evident. Global hypomethylation was found associated with altered expression of Dnmts, Igf2-H19 and Dlk-Meg3 imprinted genes along with reduced expression of Ezh2, cell cycle regulator p57KIP2 and Meg3; however, Pten remained unaltered. Increased expression of PCNA and Ki67 was observed in concert with complete lack of SOX-9 suggesting Sertoli cells independent proliferation. CONCLUSIONS: Mouse model for T2GCT is described which will have immense potential to understand cancer initiation, cancer stem cells and also to develop effective therapies in future. T2GCT initiates from tissue-resident, pluripotent VSELs due to their altered epigenome. Neonatal exposure to DES blocks differentiation (spermatogenesis) and VSELs get transformed into CD166 positive cancer stem cells that undergo excessive self-renewal and initiate cancer in adult life challenging existing concept of fetal origin of T2GCT.

Very small embryonic-like stem cells (VSELs) regenerate whereas mesenchymal stromal cells (MSCs) rejuvenate diseased reproductive tissues.

Compared to embryonic and induced pluripotent stem cells, mesenchymal stem/stromal cells (MSCs) have made their presence felt with good therapeutic promise and safety profile. Transplanting MSCs has successfully helped to reverse infertility and resulted in live births in animal models and also in humans. But the underlying mechanism for their therapeutic potential is not yet clear. MSCs are not pluripotent and hence lack plasticity to differentiate into multiple adult cell types. They rather act as 'paracrine providers' to the tissue-resident stem cells since similar beneficial effects are also observed when their secretome (microvesicles or exosomes) is transplanted. Cytokines, growth factors, signaling lipids, mRNAs, and miRNAs secreted by MSCs enables tissue-resident stem cells to undergo differentiation into specific cell types. Tissue-resident stem cells include pluripotent, very small embryonic-like stem cells (VSELs) and progenitors [spermatogonial (SSCs), ovarian (OSCs) and endometrial (EnSCs) stem cells in testes, ovary and uterus respectively] which function in a subtle manner to maintain life-long tissue homeostasis and regenerate damaged (non-functional) reproductive tissues by differentiating into sperm, oocytes and endometrial epithelial cells respectively. Similar to restoring spermatogenesis, primordial follicles numbers are increased upon transplanting MSCs. Published literature suggests that MSCs do not differentiate into epithelial cells in the endometrium. Nuclear OCT-4 positive VSELs and cytoplasmic OCT-4, AXIN2 and KERATIN-19 positive epithelial progenitors have a greater role during endometrial regeneration. We propose, transplantation of MSCs simply provides growth factors/cytokines essential for the tissue-resident stem/progenitor cells to undergo differentiation into sperm, eggs and endometrial epithelial cells in the reproductive tissues.

GFP Tagged VSELs Help Delineate Novel Stem Cells Biology in Multiple Adult Tissues.

Various types of stem cells are being researched upon to exploit their potential for regenerative medicine including pluripotent human embryonic stem (hES) cells derived from spare human embryos, induced pluripotent stem (iPS) cells by reprogramming somatic cells to a pluripotent state and multipotent mesenchymal stem/stromal cells (MSCs) obtained in vitro from multiple tissues. More than 50 independent groups have reported another novel population of pluripotent stem cells in adult tissues termed very small embryonic-like stem cells (VSELs). VSELs are developmentally linked to primordial germ cells, which rather than giving rise to the germ cells and later ceasing to exist, survive throughout life in multiple organs along with tissue-specific adult stem cells better described as lineage-restricted, tissue-committed progenitors with limited plasticity. VSELs survive total body irradiation in bone marrow, oncotherapy in the gonads, bilateral ovariectomy in the uterus and partial pancreatectomy in the pancreas of mice and participate in the regeneration of multiple organs under normal physiological conditions. VSELs and tissue-specific progenitor cells work together in a subtle manner, maintain life-long tissue homeostasis and their dysfunction leads to various pathologies including cancer. However, due to their quiescent state, VSELs have invariably eluded lineage-tracing studies reported so far. Present article reviews novel insights into VSELs biology and how VSELs enriched from GFP (green fluorescent protein) mice have enabled to delineate their role in various biological processes in vivo. VSELs biology needs to be understood in-depth as this alone will help evolve the field of regenerative medicine and win the war against cancer.

Cost-effectiveness of direct-acting antivirals for chronic hepatitis C virus in the United States from a payer perspective.

BACKGROUND: Direct-acting antivirals (DAAs) have been a breakthrough therapeutic innovation in the treatment of chronic hepatitis C virus (HCV) with significantly improved efficacy, safety, and tolerability. OBJECTIVE: To evaluate the cost-effectiveness of treating patients with HCV with DAAs compared with pre-DAAs or no treatment over a lifetime horizon from the perspective of the US Veterans Affairs (VA) health care system. METHODS: A hybrid decision-tree and Markov model simulated the health outcomes of a cohort of 142,147 patients with HCV with an average age of 63 years. Demographic data, treatment rates and distribution, treatment efficacy by subpopulation, and health state costs were sourced from VA data. Treatment costs and utility values were sourced from publicly available databases and prior publications for older regimens. RESULTS: Over a lifetime horizon, the use of DAAs results in a significant reduction in advanced liver disease events compared with pre-DAA and no treatment. Total cost savings of $7 and $9 billion over a lifetime horizon (50 years) were predicted for patients who received DAA treatments compared with patients treated with pre-DAA treatments and those who were untreated, respectively. Cost savings were achieved quickly after treatment, with DAAs being inexpensive when compared with both the pre-DAA and untreated scenarios within 5 years. The DAA intervention dominated (ie, more effective and less costly) for both the pre-DAA and untreated strategies on both a per-patient and cohort basis. CONCLUSIONS: The use of DAA-based treatments in patients with HCV in the VA system significantly reduced long-term HCV-related morbidity and mortality, while providing cost savings within only 5 years of treatment. DISCLOSURES: This work was supported by Gilead Inc. Health Economic Outcomes Research group, grant number GS-US-18-HCV003. Drs Yehoshua and Kaushik are employees of Gilead in the Health Economic Outcome Research group. These individuals reviewed the manuscript but did not contribute to input or output of the Markov model. Maple Health Group (Dr El-Moustaid, Ms Raad, and Dr Smith) are consultants hired by Gilead for Markov modeling expertise. The model used in this study was previously published and peer reviewed. Data inputted into the model related to patient demographic, treatment outcomes, clinical outcomes, and costs were completely independent in derivation by Drs Kaplan, Serper, and Durkin and were not influenced by the funding sponsor. Dr Kaplan reports grants from Gilead Inc. during the conduct of the study and grants from Gilead Inc., other from Glycotest Inc., other from AstraZeneca, other from Exact Sciences, and other from Bayer outside the submitted work.

Testicular Stem Cell Dysfunction Due to Environmental Insults Could Be Responsible for Deteriorating Reproductive Health of Men.

Reproductive health of men has declined over time including reduced semen quality specifically sperm count, increased incidence of infertility, and testicular cancers. Our recent findings suggest that these disease states possibly arise as a result of disruption of testicular stem cells biology by perinatal insults including exposure to endocrine disrupting chemicals. Testicular stem cells include relatively quiescent, very small embryonic-like stem cells (VSELs), and actively dividing spermatogonial stem cells (SSCs). Both VSELs and SSCs express estrogen receptors and are directly vulnerable to endocrine disruption. Exposing mice pups to estradiol (20 µg/pup/day on days 5-7) or diethylstilbestrol (2 µg/pup/day on days 1-5) affected spermatogenesis during adult life with reduced numbers of tubules in stage VIII, tetraploid cells and sperm. These mice were infertile and majority of diethylstilbestrol treated mice revealed testicular cancer-like changes. An increase in VSEL numbers, observed by both flow cytometry and qRT-PCR, was associated with marked reduction of c-KIT positive spermatogonial cells. VSELs undergo epigenetic changes due to endocrine disruption that results in blocked differentiation (impaired spermatogenesis) leading to reduced sperm count and infertility, and their excessive self-renewal initiates cancer-like changes in adult life. Thus, testicular dysgenesis syndrome (TDS) has a stem cell rather than a genetic basis.

Endogenous, tissue-resident stem/progenitor cells in gonads and bone marrow express FSHR and respond to FSH via FSHR-3.

Follicle stimulating hormone (FSH) is secreted by the anterior pituitary and acts on the germ cells indirectly through Granulosa cells in ovaries and Sertoli cells in the testes. Extragonadal action of FSH has been reported but is still debated. Adult tissues harbor two populations of stem cells including a reserve population of primitive, small-sized, pluripotent very small embryonic-like stem cells (VSELs) and slightly bigger, tissue-specific progenitors which include ovarian stem cells (OSCs) in ovaries, spermatogonial stem cells (SSCs) in testes, endometrial stem cells (EnSCs) in uterus and hematopoietic stem cells (HSCs) in the bone marrow. Data has accumulated in animal models showing FSHR expression on both VSELs and progenitors in ovaries, testes, uterus and bone marrow and eventually gets lost as the cells differentiate further. FSH exerts a direct action on the stem/progenitor cells via alternatively spliced FSHR-3 rather than the canonical FSHR-1. FSH stimulates VSELs to undergo asymmetrical cell divisions to self-renew and give rise to the progenitors that in turn undergo symmetrical cell divisions and clonal expansions followed by differentiation into specific cell types. Excessive self-renewal of VSELs results in cancer and this explains ubiquitous expression of embryonic markers including nuclear OCT-4 along with FSHR in cancerous tissues. Focus of this review is to compile published data to support this concept. FSHR expression in stem/progenitor cells was confirmed by immuno-fluorescence, Western blotting, in situ hybridization and by quantitative RT-PCR. Two different commercially available antibodies (Abcam, Santacruz) were used to confirm specificity of FSHR expression along with omission of primary antibody and pre-incubation of antibody with immunizing peptide as negative controls. Western blotting allowed detection of alternatively spliced FSHR isoforms. Oligoprobes and primers specific for Fshr-1 and Fshr-3 were used to study these alternately-sliced isoforms by in situ hybridization and their differential expression upon FSH treatment by qRT-PCR. To conclude, stem/progenitor cells in adult tissues express FSHR and directly respond to FSH via FSHR-3. These findings change the field of FSH-FSHR biology, call for paradigm shift, explain FSHR expression on cancer cells in multiple organs and provide straightforward explanations for various existing conundrums including extragonadal expression of FSHR.

Additional Evidence to Establish Existence of Two Stem Cell Populations Including VSELs and SSCs in Adult Mouse Testes.

Present study aims to describe a simple and robust protocol to delineate the presence of pluripotent, very small embryonic-like stem cells (VSELs) in addition to spermatogonial stem cells (SSCs) in adult mouse testes. Testicular seminiferous tubules were subjected to enzymatic dissociation to obtain single cells suspension. Stem cells were enriched by spinning at different speeds wherein majority of somatic cells were pelleted at 1000 rpm (250 g, Pellet A) and putative stem cells by spinning the supernatant (obtained after separating Pellet A) at 3000 rpm (1000 g, Pellet B). Viable (7AAD-ve), 2-6 µm, LIN-CD45-SCA-1+ VSELs were studied after doublets exclusion by flow cytometry in both Pellets A & B. Almost ten-fold enrichment of VSELs was obtained in Pellet B (0.27 + 0.05%) compared to Pellet A (0.03 + 0.003%). SCA-1 expressing SSCs (>6 µm, 0.18 + 0.06%) were clearly distinguished from VSELs (2-6 µm, 0.07 + 0.003%) by flow cytometry studies on total testicular cells suspension collected by spinning at 3000 rpm. Enriched stem cells from Pellet B were used to study expression of OCT-4, NANOG, SCA-1, SSEA-1, LIFR, GFRa, c-KIT, ERα and ERß. Cells in Pellet B were also subjected to RT-PCR to study pluripotent (Oct-4a, Sox2, Nanog), primordial germ cells (Stella, Fragilis), SSCs (Oct-4) and estrogen receptors (ERα and ERß) specific transcripts. qRT-PCR analysis showed >2 folds up-regulation of stem cell markers in Pellet B (Oct-4A, Oct-4, Sox2, Nanog) compared to Pellet A. To conclude, spinning at higher speed led to successful enrichment of pluripotent VSELs from testes which have remained ignored till now. Expression of ERα & ß on VSELs/SSCs makes them vulnerable to endocrine disruption.

Altered Biology of Testicular VSELs and SSCs by Neonatal Endocrine Disruption Results in Defective Spermatogenesis, Reduced Fertility and Tumor Initiation in Adult Mice.

Reproductive health of men has declined in recent past with reduced sperm count and increased incidence of infertility and testicular cancers mainly attributed to endocrine disruption in early life. Present study aims to evaluate whether testicular stem cells including very small embryonic-like stem cells (VSELs) and spermatogonial stem cells (SSCs) get affected by endocrine disruption and result in pathologies in adult life. Effect of treatment on mice pups with estradiol (20 µg on days 5-7) and diethylstilbestrol (DES, 2 µg on days 1-5) was studied on VSELs, SSCs and spermatogonial cells in adult life. Treatment affected spermatogenesis, tubules in Stage VIII & sperm count were reduced along with reduction of meiotic (4n) cells and markers (Prohibitin, Scp3, Protamine). Enumeration of VSELs by flow cytometry (2-6 µm, 7AAD-, LIN-CD45-SCA-1+) and qRT-PCR using specific transcripts for VSELs (Oct-4a, Sox-2, Nanog, Stella, Fragilis), SSCs (tOct-4, Gfra-1, Gpr-125) and early germ cells (Mvh, Dazl) showed several-fold increase but transition from c-Kit negative to c-Kit positive spermatogonial cells was blocked on D100 after treatment. Transcripts specific for apoptosis (Bcl2, Bax) remained unaffected but tumor suppressor (p53) and epigenetic regulator (NP95) transcripts showed marked disruption. 9 of 10 mice exposed to DES showed tumor-like changes. To conclude, endocrine disruption resulted in a tilt towards excessive self-renewal of VSELs (leading to testicular cancer after DES treatment) and blocked differentiation (reduced numbers of c-Kit positive cells, meiosis, sperm count and fertility). Understanding the underlying basis for infertility and cancer initiation from endogenous stem cells through murine modelling will hopefully improve human therapies in future.