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Transcriptionally silent HIV proviruses form the major obstacle to eradicating HIV. Many studies of HIV latency have focused on the cellular mechanisms that maintain silencing of proviral DNA. Here we show that viral sequence variation affecting replicative ability leads to variable rates of silencing and ability to reactivate. We studied naturally occurring and engineered polymorphisms in a recently identified exonic splice enhancer (ESEtat) that regulates tat mRNA splicing and constructed viruses with increased (strain M1), reduced (strain M2), or completely absent (strain ERK) binding of splicing factors essential for optimal production of tat mRNA resulting in a corresponding change in Tat activity. The mutations affected viral replication, with M1 having wild-type (WT) kinetics, M2 exhibiting reduced kinetics, and ERK showing completely abrogated replication. Using single-round infection with green fluorescent protein (GFP)-expressing viruses to study proviral gene expression, we observed progressively greater rates of silencing relating to the degree of ESEtat disruption, with the WT strain at 53%, strain M2 at 69%, and strain ERK at 94%. By stimulating infected cells with a latency reversal agent (phorbol myristate acetate [PMA], panobinostat, or JQ1), we observed that the dose required to achieve 50% of the maximum signal was lowest in the WT, intermediate in M2, and highest in ERK, indicating progressively higher thresholds for reactivation. These results suggest that the ability of silent proviruses to reactivate from latency is variable and that minor differences in the viral sequence can alter the proportion of silenced viruses as well as the threshold required to induce silenced viruses to reactivate and express. IMPORTANCE A reservoir of infected cells in which the HIV genome is transcriptionally silent is acknowledged to be the principal barrier to eradicating the virus from an infected person. A number of cellular processes are implicated in this silencing; however, the viral factors that may contribute remain underexplored. Here we examined mutations altering the correct splicing of HIV gene products as a model to study whether differences in viral sequence can affect either the proportion of viruses that are active or silent or their ability to reactivate. We found that some naturally occurring variations result in viruses that are silenced at a higher rate and require a proportionally increased stimulus for reactivation from latency. These data suggest that the silencing and reactivation behavior of HIV exists in a spectrum, influenced by factors intrinsic to the virus.

Viral hepatitis is a major cause of morbidity and mortality worldwide and a rising cause for concern in Asian countries. Weather it is blood borne or water/food borne hepatotropic virus, increasing burden is alarming for Asian countries. In this review we have evaluated the existing data to estimate the burden of viral hepatitis in populations of all age groups nationwide, along with an assessment of the risk factors and preventive and management strategies currently employed in Pakistan. The aim of our work is to consolidate and supplement the present knowledge regarding viral hepatitis in light of past and present trends and to provide future direction to the existing health policies.How to cite this articleButt AS, Sharif F. Viral Hepatitis in Pakistan: Past, Present, and Future. Euroasian J Hepato-Gastroenterol 2016;6(1):70-81.

Aim: Prevalence of obesity and type 2 diabetes mellitus (T2DM), both of which represent are related to nonalcoholic fatty liver disease (NAFLD), is an increasing trend among Asian people. The study aimed to assess the prevalence of NAFLD in T2DM with their risk factors in the Southern part of Pakistan. Materials and methods: A cross-sectional study was accomplished during 2008-2013 at The Aga Khan University Hospital, Karachi, Pakistan. Adult patients diagnosed with T2DM during last 6 months were enrolled in this study. NAFLD was identified using ultrasound of the liver. Clinical and biochemical relevant measurements were accomplished. Results: Out of a total of 203 patients with T2DM, NAFLD was detected in 146 patients (71.9%). Multivariate analysis revealed that NAFLD was significantly associated with dyslipidemia (OR 2.38, 95% CI 1.06-5.34, p = 0.035), higher LDL (OR 1.02, 95%CI 1.01-1.03, p = 0.003), HbA1c (OR1.27, 95% CI 0.97-1.68, p = 0.045) and diastolic blood pressure (OR 1.05, 95% CI 1.01-1.10, p = 0.009). The highest odds of 10.8 for NAFLD (95% CI 4.9-24, p = 0.001) was found for the combination of hypertension, dyslipidemia, body mass index (BMI), waist circumference, lack of physical inactivity, triglycerides, lower HDL, LDL, HbA1c, and ALT (multiplicative analysis). Conclusion: High incidence of NAFLD with the association of different lifestyle-related factors has been analyzed. It unmasks the need for screening for NAFLD in newly diagnosed DM patients in Pakistan with the assessment of parameters of risk factors.

Hepatocellular carcinoma (HCC) is a major cause of morbidity and mortality worldwide. Chronic hepatitis C virus infection (HCV) is the most common cause of HCC in many European countries, Japan and Pakistan. Introduction of the new direct acting antivirals (DAAs) has revolutionized the management of HCV worldwide, with high rates of sustained virologic response in patients who could not have tolerated the previous interferon based treatments. However, recently there have been reports raising caution about the long term effects of DAAs, particularly a possible increased risk of HCC. Therefore this review explores the current molecular studies as well as clinical data that investigate the impact of DAAs on occurrence and recurrence of HCC.