Kaneez Fatima

Oxidative stress and hepatitis C virus

The disproportionate imbalance between the systemic manifestation of reactive oxygen species and body’s ability to detoxify the reactive intermediates is referred to as oxidative stress. Several biological processes as well as infectious agents, physiological or environmental stress, and perturbed antioxidant response can promote oxidative stress. Oxidative stress usually happens when cells are exposed to more electrically charged reactive oxygen species (ROS) such as H2O2 or O2-. The cells’ ability to handle such pro-oxidant species is impeded by viral infections particularly within liver that plays an important role in metabolism and detoxification of harmful substances. During liver diseases (such as hepatocellular or cholestatic problems), the produced ROS are involved in transcriptional activation of a large number of cytokines and growth factors, and continued production of ROS and Reactive Nitrogen Species (RNS) feed into the vicious cycle. Many human viruses like HCV are evolved to manipulate this delicate pro- and antioxidant balance; thus generating the sustainable oxidative stress that not only causes hepatic damage but also stimulates the processes to reduce treatment of damage. In this review article, the oxidant and antioxidant pathways that are perturbed by HCV genes are discussed. In the first line of risk, the pathways of lipid metabolism present a clear danger in accumulation of viral induced ROS. Viral infection leads to decrease in cellular concentrations of glutathione (GSH) resulting in oxidation of important components of cells such as proteins, DNA and lipids as well as double strand breakage of DNA. These disorders have the tendency to lead the cells toward cirrhosis and hepatocellular carcinoma in adults due to constant insult. We have highlighted the importance of such pathways and revealed differences in the extent of oxidative stress caused by HCV infection.

Hepatitis B virus, HBx mutants and their role in hepatocellular carcinoma

Hepatocellular carcinoma (HCC) is one of the leading causes of death induced by cancer in the modern world and majority of the cases are related to chronic hepatitis B virus (HBV) infection. HBV-encoded X protein (HBx) is known to play a pivotal role in the pathogenesis of viral induced HCC. HBx is a multifunctional protein of 17 kDa which modulates several cellular processes by direct or indirect interaction with a repertoire of host factors resulting in HCC. HBX might interfere with several cellular processes such as oxidative stress, DNA repair, signal transduction, transcription, protein degradation, cell cycle progression and apoptosis. A number of reports have indicated that HBx is one of the most common viral ORFs that is often integrated into the host genome and its sequence variants play a crucial role in HCC. By mutational or deletion analysis it was shown that carboxy terminal of HBx has a likely role in protein-protein interactions, transcriptional transactivation, DNA repair, cell, signaling and pathogenesis of HCC. The accumulated evidence thus far suggests that it is difficult to understand the mechanistic nature of HBx associated HCC, and HBx mediated transcriptional transactivation and signaling pathways may be a major determinant. This article addresses the role of HBx in the development of HCC with particular emphasis on HBx mutants and their putative targets.

Hepatitis C Virus Infection: Molecular Pathways to Insulin resistance

Chronic Hepatitis C virus has the potential of inducing insulin resistance and type 2 Diabetes Mellitus in vitro as well as in vivo . Structural and non-structural proteins of HCV modulate cellular gene expression in such a way that insulin signaling is hampered, concomitantly leads toward diabetes mellitus. A number of mechanisms have been proposed in regard to the HCV induced insulin resistance involving the upregulation of Inflammatory cytokine TNF-α, hypophosphorylation of IRS-1 and IRS-2, phosphorylation of Akt, up-regulation of gluconeogenic genes, accumulation of lipids and targeting lipid storage organelles. This review provides an insight of molecular mechanisms by which HCV structural and non-structural proteins can induce insulin resistance.

Distribution of hepatitis C virus genotypes, hepatic steatosis and their correlation with clinical and virological factors in Pakistan

Abstract Background: Due to the inherently unstable nature of HCV, various genotypes have been identified. Steatosis is a histological feature in the progression of HCV-associated liver disease and has been shown to alter the host lipid metabolism. Objective: Assess the distribution of HCV genotypes in the two provinces of Pakistan, and determine the association of hepatic steatosis with altered clinical and virological factors in chronic HCV patients. Methods: One hundred twenty six chronic HCV patients (steatosis in 49 patients) were enrolled for qualitative analysis by PCR. Out of 126 ELISA and PCR positive samples, 119 (48 with hepatic steatosis) chronic HCV patients (mean age 42.0±13.3 years, mean body mass index (BMI) 24.2±4.1) were proved positive after PCR-based detection. Biochemical and virological factors such as HCV genotype, or glucose, in 119 CHC patients were determined and compared between patients with and without hepatic steatosis. Results: Out of 126 samples, 119 were HCV positive, where 58 (48.7%) were genotype 3a, 24 (20.2%) were 3b, 12 (10.1%) were 1a, eight (6.7%) were 2a, six (5.0%) were 1b, and one (0.8%) was 4. Furthermore, seven (5.9%) had a co-infection and three (2.5%) were untypable. BMI (p=0.004), genotype 3a (p<0.001), and triglycerides (p=0.002) were significantly associated with steatosis. It is noteworthy that cholesterol (p=0.281), glucose (p=0.305), lowdensity lipoprotein (p=0.101), high-density lipoprotein (p=0.129), alanine amino transferase (p=0.099), aspartate transaminase (p=0.177), bilirubin (p= 0.882), and age (p=0.846) showed non-significant association. Conclusion: Genotype 3a is the predominant genotype in Pakistan. Hepatic steatosis is quite frequent feature in HCV patients and strongly correlates with BMI, genotype 3a, and triglyceride contents in patients infected with HCV.

Hepatitis C virus and neurological damage.

Chronic hepatitis C virus (HCV) infection exhibits a wide range of extrahepatic complications, affecting various organs in the human body. Numerous HCV patients suffer neurological manifestations, ranging from cognitive impairment to peripheral neuropathy. Overexpression of the host immune response leads to the production of immune complexes, cryoglobulins, as well as autoantibodies, which is a major pathogenic mechanism responsible for nervous system dysfunction. Alternatively circulating inflammatory cytokines and chemokines and HCV replication in neurons is another factor that severely affects the nervous system. Furthermore, HCV infection causes both sensory and motor peripheral neuropathy in the mixed cryoglobulinemia as well as known as an important risk aspect for stroke. These extrahepatic manifestations are the reason behind underlying hepatic encephalopathy and chronic liver disease. The brain is an apt location for HCV replication, where the HCV virus may directly wield neurotoxicity. Other mechanisms that takes place by chronic HCV infection due the pathogenesis of neuropsychiatric disorders includes derangement of metabolic pathways of infected cells, autoimmune disorders, systemic or cerebral inflammation and alterations in neurotransmitter circuits. HCV and its pathogenic role is suggested by enhancement of psychiatric and neurological symptoms in patients attaining a sustained virologic response followed by treatment with interferon; however, further studies are required to fully assess the impact of HCV infection and its specific antiviral targets associated with neuropsychiatric disorders.

Biomarkers for virus-induced hepatocellular carcinoma (HCC)

Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide, and is advanced by severe viral hepatitis B or C (HBV or HCV) as well as alcoholic liver disease. Many patients with early disease are asymptomatic therefore HCC is frequently diagnosed late requiring costly surgical resection or transplantation. The available non-invasive detections systems are based on the clinical utility of alpha fetoprotein (AFP) measurement, together with ultrasound and other more sensitive imaging techniques. The hallmark of liver disease and its propensity to develop into fully blown HCC is depended on several factors including the host genetic make-up and immune responses. While common symptoms involve diarrhea, bone pain, dyspnea, intraperitoneal bleeding, obstructive jaundice, and paraneoplastic syndrome, the evolution of cell and immune markers is important to understand viral induced liver cancers in humans. The circulating miRNA, cell and immune based HCC biomarkers are imperative candidates to successfully develop strategies to restrain liver injury. The current molecular genetics and proteomic analysis have lead to the identification of number of key biomarkers for HCC for earlier diagnosis and more effective treatment of HCC patients. In this review article, we provide latest updates on the biomarkers of HBV or HCV-associated HCC and their co-evolutionary relationship with liver cancer.

An overview: In vitro models of HCV replication in different cell cultures

Although much of productive research has been conducted in the field of molecular virology of Hepatitis C virus (HCV) regarding its genes, gene functions and proteins, development of an efficient cell culture model for its replication remained a focused area. Focus has been directed to establish HCV in vitro replication system. This replication system should mimic its intrahepatic pathogenesis so that antivirals should be screened and in vitro gene profiling of HCV induced pathogenesis should be worked out. Since 1990 various experimental approaches and strategies have been utilized in phase of development of a robust replication model for HCV, and success has been reported for a few genotypes. Still the work is going on to have more success in availing such robust replication models for all the genotypes. This will help to have a common antiviral strategy against HCV induced pathogenesis involving any genotype or subtype.

Identification of ionotrophic purinergic receptors in Huh-7 cells and their response towards structural proteins of HCV genotype 3a

Hepatitis C virus (HCV) is a major health problem in developing countries including Pakistan. Chronic HCV infection results in progressive liver disease including fibrosis, cirrhosis, insulin resistance and eventually hepatocellular carcinoma (HCC). Ionotrophic purinergic (P2X) receptors are identified to involve in a spectrum of physiological and pathophysiological processes. However, the role of P2X receptors in HCV liver associated diseases still remains to be investigated. The current study was designed to identify the presence of P2X receptors in human liver cells. Furthermore, it investigates the response of P2X receptors towards HCV structural proteins (E1E2). To determine that how many isoforms of P2X receptors are expressed in human liver cells, human hepatoma cell line (Huh-7) was used. Transcripts (mRNA) of five different isoforms of P2X receptors were identified in Huh-7 cells. To examine the gene expression of identified isoforms of P2X receptors in presence of HCV structural proteins E1E2, Huh-7/E1E2 cell line (stably expressing HCV structural proteins E1E2) was used. The results showed significant increase (6.2 fold) in gene expression of P2X4 receptors in Huh-7/E1E2 cells as compared to control Huh-7 cells. The findings of present study confirmed the presence of transcripts of five different isoforms of P2X receptors in human liver cells and suggest that P2X4 receptors could be represented an important component of the purinergic signaling complex in HCV induced liver pathogenesis.

Potential mechanisms of hepatitis B virus induced liver injury

Chronic active hepatitis (CAH) is acknowledged as an imperative risk factor for the development of liver injury and hepatocellular carcinoma. The histological end points of CAH are chronic inflammation, fibrosis and cirrhosis which are coupled with increased DNA synthesis in cirrhotic vs healthy normal livers. The potential mechanism involved in CAH includes a combination of processes leading to liver cell necrosis, inflammation and cytokine production and liver scaring (fibrosis). The severity of liver damage is regulated by Hepatitis B virus genotypes and viral components. The viral and cellular factors that contribute to liver injury are discussed in this article. Liver injury caused by the viral infection affects many cellular processes such as cell signaling, apoptosis, transcription, DNA repair which in turn induce radical effects on cell survival, growth, transformation and maintenance. The consequence of such perturbations is resulted in the alteration of bile secretion, gluconeogenesis, glycolysis, detoxification and metabolism of carbohydrates, proteins, fat and balance of nutrients. The identification and elucidation of the molecular pathways perturbed by the viral proteins are important in order to design effective strategy to minimize and/or restore the hepatocytes injury.

Docking Studies of Pakistani HCV NS3 Helicase: A Possible Antiviral Drug Target

The nonstructural protein 3 (NS3) of hepatitis C virus (HCV) helicase is believed to be essential for viral replication and has become an attractive target for the development of antiviral drugs. The study of helicase is useful for elucidating its involvement in positive sense single-stranded RNA virus replication and to serve as templates for the design of novel antiviral drugs. In recent years, several models have been proposed on the conformational change leading to protein movement and RNA unwinding. Some compounds have been recently reported to inhibit the helicase and these include small molecules, RNA aptamers and antibodies. The current study is designed to help gain insights for the consideration of potential inhibitors for Pakistani HCV NS3 helicase protein. We have cloned, expressed and purified HCV NS3 helicase from Pakistani HCV serum samples and determined its 3D structure and employed it further in computational docking analysis to identify inhibitors against HCV genotype 3a (GT3a),including six antiviral key molecules such as quercetin, beta-carotene, resveratrol, catechins, lycopene and lutein. The conformation obtained after docking showed good hydrogen bond (HBond) interactions with best docking energy for quercetin and catechins followed by resveratrol and lutein. These anti-helicase key molecules will offer an alternative attraction to target the viral helicase, due to the current limitation with the interferon resistance treatment and presences of high rate of resistance in anti-protease inhibitor classes.