An Insight Into Mitochondrial Dysfunction and its Implications in Neurological Diseases.

In the last few years, a massive increase in research has been observed that focuses on investigating the role of mitochondria in the pathogenesis of several neurodegenerative disorders. Mitochondria are vital cell organelles having important roles in different cellular processes, including energy production, calcium signaling, Reactive Oxygen Species (ROS) generation, apoptosis, etc. Therefore, healthy mitochondria are necessary for cell survival and functioning. It would seem feasible that mitochondrial dysfunction will have implications in various pathological conditions. A large body of evidence indicates the role of the mitochondrion as a potential key player in the loss or dysfunction of neurons in various neurodegenerative disorders. In this review, we provide an insight into mitochondrial dysfunction and its involvement in the pathology of several neurological diseases such as Alzheimer's disease, Parkinson's disease, Amyotrophic Lateral Sclerosis, Hypoxic- Ischemic Brain Injury, and many more.

Identification and full-length molecular characterization of rare hepatitis C virus genotype 5a from India.

This report presents a molecular characterization of the complete genome of a rare hepatitis C virus (HCV) genotype (GT5a) from India. Sequence homology of full genome revealed that the strain belonged to HCV GT5a. To trace the origin of this virus and to understand its evolutionary pattern, a phylogenetic reconstruction was carried out on full HCV genome sequences using Bayesian coalescent methods. The phylogenetic tree reconstruction revealed genotypic divergence, with formation of distinct clades. This analysis revealed that HCV genotype 5 might have originated from HCV genotype 3, as they have a recent common ancestor.

Identification of Indian sub-continent as hotspot for HCV genotype 3a origin by Bayesian evolutionary reconstruction.

BACKGROUND: Recent emphasis in Hepatitis C virus (HCV) evolutionary biology has focused on analysis using Core, E1/E2 and/or NS5b regions, with limited appreciation of full length genome. While HCV genotypes have been described as endemic in the Indian subcontinent, there has been no confirmation at the molecular evolutionary level of these genotypes. We have attempted here to determine the status of Indian HCV genotype 3a sequences in relation to similar genotypes from other parts of the world. METHODS: Cloning, sequencing and molecular characterization was performed on 9 Indian sequences and comparative analyses were performed with 46 sequences from other countries. Evolutionary-rate and molecular-clock hypothesis testing was addressed by Bayesian MCMC. RESULTS: Genetic analysis of full length genome revealed two hypervariable regions (HVR) in E2 region - HVR496 and HVR576, with a variable 5-8 amino-acid insertion sequence and a putative N-glycosylation site. Phylogenetic analysis revealed a divergence resulting in 2 distinct clades: clade-1 represented by HCV 3a subtype and clade-2 represented by other 3 subtype genomes. Clade-2 shows earlier divergence than clade-1. Analysis revealed that genotype 3a genomes from India roots out first (∼99 years ago) in clade1. Bayesian skyline plot analysis revealed an increase in effective number of infections from 1940s to 1990s, followed by a gradual decrease after 2000. CONCLUSIONS: Genotype 3a sequences appear to have originated in India and later dispersed to United Kingdom around mid 1940s, most likely around the time of Indian independence and World War II.

Assessment of arsenic-induced DNA damage in goldfish by a polymerase chain reaction-based technique using random amplified polymorphic DNA markers.

Arsenic is a groundwater contaminant of global concern. It is a potent human carcinogen, and its marked genotoxic effects have been reported in several human and animal studies. The present work investigates the applicability of the random amplified polymorphic DNA (RAPD) assay to study the DNA-damaging effects of arsenic at low-level exposure in goldfish Carassius auratus. Four experimental groups of fish, A, B, C and D, were exposed to 0, 10, 50, and 1,000 µg L(-1) of arsenic, respectively, in aquaria water for 15 consecutive days. Genomic DNA extraction was followed by RAPD-polymerase chain reaction amplification for each fish separately. One arbitrary decamer primer (PUZ-19) of 33 primers used appeared as the most informative and was capable of exhibiting marked alterations in RAPD profiles between arsenic-exposed and unexposed (control) samples. Different sets of 11 loci were amplified in various experimental groups with four clear polymorphic bands by the primer PUZ-19. The X and XIII amplification loci, which were prominent in the unexposed group, failed to appear in the arsenic-exposed groups. In contrast, the I and XI RAPD bands appeared as new amplification loci in all of the exposed groups. Such alterations in genomic DNA, however, did not exhibit a clear dose-dependent tendency. The RAPD assay, because of its efficacy to unmask alterations in genomic DNA induced by arsenic at low exposure level of 10 µg L(-1), appears to be a sensitive and potential tool for detecting arsenic genotoxicity.

Genetic polymorphisms of CYP2E1 and DNA repair genes HOGG1 and XRCC1: association with hepatitis B related advanced liver disease and cancer.

A population based case-control study was designed to explore the genetic risk factors for hepatitis B virus (HBV) related liver disease susceptibility. A total of 424 subjects comprising 210 controls, 50 acute HBV (AVH), 84 chronic HBV (CHBV), 25 HBV related cirrhosis and 55 HBV related hepatocellular carcinoma (HCC) cases were included in the study. PCR-RFLP was used for the genotyping of Cyp2E1*5B, hOGG1 codon 326 and XRCC1 codon 399. Compared to controls, Cyp2E1 rsaI variant c2 genotype increased the risk of HBV related liver disease severity by 2.68 fold, the highest for HCC cases (3.981 folds, p=0.106); and was associated with higher histology activity index (HAI) (p<0.001) in CHBV patients. Cyp2E1 and hOGG1 variants were independently associated with a significantly higher fibrosis score in CHBV group. Analysis of gene-gene interaction studies showed an increased risk of HCC, cirrhosis and CHBV in a Cyp2E1 variant+XRCC1 variant combination (p<0.001); and hOGG1 variants+XRCC1 variants. A mutually independent heterozygous hOGG1 and XRCC1 combination resulted in a decreased risk of HBV related liver disease. On the other hand, a wild-type hOGG1 and XRCC1 combination was associated with a significantly higher risk of AVH (p=0.010) but a lower risk of CHBV (p=0.032) and HCC (p=0.006). The gene-gene interactions were also associated with a significant increase in HAI and fibrosis score in CHBV patients. Cyp2E1, hOGG1 and XRCC1 genotypes significantly alter the risk of HBV related liver disease susceptibility and severity, independently or through gene-gene interaction.

Basal core promoter, precore region mutations of HBV and their association with e antigen, genotype, and severity of liver disease in patients with chronic hepatitis B in India.

Spontaneous mutations of hepatitis B virus (HBV) could influence the severity of liver disease. Since the basal core promoter (BCP) and the precore (Pc) regions are important for viral replication, these regions were examined for naturally occurring mutations and were correlated with the genotype, e antigen status, and severity of liver disease. In 82 patients with histologically confirmed chronic hepatitis B, the BCP and Pc regions were sequenced and aligned with known wild-type sequences. Sequence based HBV genotyping was done and HBV DNA was quantified. Thirty-three (40%) patients had decompensated chronic liver disease and the remaining patients had chronic hepatitis B. Forty-six (56%) patients were HBeAg positive. HBV genotype A was found in 28%, D in 65%, and B/C in 7.3%. The Pc G1896A mutation was more common in HBeAg-negative (33% vs. 2%, P < 0.01) patients and was genotype D specific. The Pc G1862T mutation was detected more often in HBeAg-positive than HBeAg-negative (37% vs. 11%, P < 0.01) patients and was genotype A specific (P < 0.01). BCP mutations at the 1,762/64 nucleotide positions were common in HBeAg negative than positive (36% vs. 13%, P < 0.05) and were equally common in different genotypes. TA 1-3 region mutations of the BCP were significantly higher in HBeAg-negative as compared to HBeAg-positive patients (78% vs. 26%, P < 0.01). BCP mutations had significantly higher HBV DNA levels. It is concluded that Pc G1862T mutant is Genotype A-specific but is not always associated with e antigen. The TA 1-3 rich mutations of BCP region are also associated with the absence of e antigen in Indian patients.

Transmission of G145R mutant of HBV to an unrelated contact.

Household contacts of HBV-related chronic liver disease patients constitute a high-risk group for acquisition of HBV infection. Some of the HBsAg mutants are associated with liver disease and some are reported to be transmitted vertically. There is limited information on the horizontal transmission of Gly 145 Arg (G145R) mutant to related contacts. Its possible transmission to an unrelated third degree contact is reported in the present study. An HBV related chronic liver disease patient; the index patient, and his 11 household contacts were studied. This included four 1 degrees, three 2 degrees, one 3 degrees, and a sexual contact. Surface gene sequencing including the "a" determinant region was carried out in HBV DNA+ve subjects. The sequences were aligned and compared for the homology. HBV DNA was found to be positive in one 1 degrees, three 2 degrees, and one 3 degrees contact, besides the index patient. Histopathological studies revealed evidence of chronic hepatitis in all these contacts. Mutation T118V was present in all the six subjects. Mutant G145R along with T118V and T143M was identified in three subjects who included one 1 degrees, one 2 degrees, and one 3 degrees contact. Presence of T118V and T143M mutations along with G145R mutation in these subjects provides an indirect evidence for the possible horizontal transmission of G145R HBV variant to a 3 degrees unrelated contact. Of these three contacts with G145R mutation, only one 1 degrees contact was found to be HBsAg-ve. The data also reaffirms the earlier finding of HBsAg positivity in presence of G145R mutation of the S-gene. HBV exists as quasi-species and mixed population in subjects with chronic HBV infection.

Association of TNF-beta polymorphism with disease severity among patients infected with hepatitis C virus.

The pathogenesis of chronic hepatitis C virus (HCV) infection remains unclear. Tumour necrosis factor alpha (TNF-alpha) is alleged to contribute in the pathogenesis of chronic HCV infection. Single nucleotide polymorphism in TNF-alpha and -beta genes could influence the outcome of HCV infection. The aim was to study single nucleotide polymorphism in TNF-alpha promoter region and Nco I polymorphisms in the TNF-beta gene in patients with chronic hepatitis C. Fifty-two patients with histologically proven chronic hepatitis, who had raised ALT levels (>1.5 x ULN) and were HCV RNA positive, were studied. Genotyping of -308 promoter variant of TNF-alpha was performed by PCR with primers that incorporated an Nco I restriction site. For PCR typing of the TNF-beta Nco I restriction fragment length polymorphism, sequence specific primers were used. Polymorphism in the TNF-alpha G/G, G/A and A/A allele was not different between HCV patients and healthy controls. TNF-beta A/A allele was significantly more common (P = 0.02) in patients (28.8%) as compared to controls (12.8%), whereas no significant difference was observed for TNF-beta G/A and G/G alleles [corrected]. Nco I TNF-beta A/A was strongly associated with -308 TNF-alpha G/G (RR of HCV persistence = 4.9), indicating possible linkage between TNF-beta A/A and TNF-alpha G/G allele. Patients with severe hepatic fibrosis more frequently had the TNF-beta A/A allele as compared to patients with mild disease (P = 0.04). Immunogenetic factors, such as single nucleotide polymorphisms in TNF-beta (A/A allele), may affect the natural course of HCV infection, in particular, the disease progression. Larger studies including cytokine expression profiles are needed to fully understand the contribution of the polymorphisms described in the pathogenesis of chronic hepatitis C.

Prevalence and profile of mutations associated with lamivudine therapy in Indian patients with chronic hepatitis B in the surface and polymerase genes of hepatitis B virus.

Lamivudine results in the selection of resistant hepatitis B virus (HBV) variants. Because the surface gene of HBV overlaps completely the polymerase gene, the incidence and profile of surface and polymerase gene mutations were investigated prospectively in chronic HBV patients who were on lamivudine therapy. Twenty-six patients with chronic liver disease confirmed histologically were included in this study. Extracted HBV DNA from sera samples were subjected to PCR amplification for the mutation prone regions of the surface and polymerase genes of the HBV genome. The emergence of mutant forms and biochemical derangements were studied carefully during the course of the therapy. In six of 26 (23%) patients, mutations emerged on lamivudine therapy. YM552I/VDD resistant mutants were observed in one (6%) and five (29%) patients at Month 12 and 18, respectively, out of 17 patients, who had completed more than 9 months of therapy. The mean time of emergence of resistance was 16.4 +/- 6.8 months. In three of the five patients, emergence of YM552I/VDD mutation was accompanied with a rise in HBV DNA levels. In two patients, mutations were noticed at the end of the viral breakthrough; when the DNA level went down to undetectable levels (<0.5 pg/mL). In two patients, normal ALT levels were found at the time of emergence of the YMDD mutation. YM552I/VDD mutations were observed in 43% of HBeAg positive and 20% of anti-HBe positive patients (P = ns). Although the 'a'-determinant region was found to be unaffected; in one patient, a novel pattern due to emergence of YIDD mutant was observed; the corresponding aa in the S-ORF turned to a stop codon. In summary, the frequency of emergence of YM552I/VDD mutations was 29% at Month 18 in the Indian patients. The presence of normal ALT and low levels of HBV DNA do not exclude the existence of resistant mutants. Novel mutations in the S-ORF, which lead to premature surface gene termination might affect the production of HBsAg and need further study.

Vertical transmission of hepatitis B virus despite maternal lamivudine therapy.

Lamivudine given during the last weeks of pregnancy in women with chronic hepatitis B has been reported to be safe. We report a case of chronic hepatitis B virus (HBV) infection in a newborn, despite suppression of HBV DNA to undetectable levels in the mother by prolonged lamivudine therapy. The newborn had raised alanine aminotransferase concentrations and was positive for HBV DNA at birth which persisted until 9 months of age, despite neonatal vaccination, treatment with hepatitis B immune globulin, and high concentrations of anti-HBs. On HBV DNA sequencing, complete sequence homology and a similar precore mutation was found in the mother and child, indicating vertical transmission. Lamivudine therapy might not prevent perinatal transmission of HBV infection in every newborn.