Efavirenz is a non-nucleoside reverse transcriptase inhibitor (NNRTI) and an active constituent of the highly active antiretroviral therapy regime. It has significantly contributed in control and management of human immunodeficiency virus propagation. However, EFV administration has also led to severe adverse effects, several reports highlighted the role of EFV in mitochondrial dysfunction and toxicity but the molecular mechanism has been poorly understood. In present study, human hepatoma cells Huh 7.5 were treated with clinically relevant concentrations of EFV and parameters like cytotoxicity, mitochondrial transmembrane potential, mitochondrial morphology, cytochrome c release, mitochondria-mediated apoptosis, mtDNA and mtRNA levels and EFV distribution into mitochondrial compartment were evaluated to understand sequence of events leading to cell death in EFV-treated cells. EFV at its clinically relevant concentration was significantly toxic after 48 and 72 h of treatments. EFV-mediated toxicity is initiated with the permeabilization of mitochondrial outer membrane and change in mitochondrial membrane potential (Δψm) which triggers a series of events like cytochrome c release, alteration in mitochondrial morphology and mitochondria-mediated apoptosis. Total mitochondrial content is reduced after 48 h of EFV treatment at IC concentration which is also reflected in reduced mitochondrial DNA and RNA levels. After detecting EFV in mitochondrial compartment after 12 h of incubation with EFV, we hypothesize that EFV being a lipophilic molecule is internalized into the mitochondrial compartment causing depolarization of Δψm which subsequently leads to a cascade of events causing cell death.
Hepatitis C virus is major cause of chronic liver diseases such as chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma. Presently available direct-acting antiviral drugs have improved success rate; however, high cost limits their utilization, especially in developing countries like India. In the present study, we evaluated anti-HCV potential of several siRNAs targeted against the HCV RNA-dependent RNA polymerase NS5B and cellular factors, La autoantigen, PSMA7, and human VAMP-associated protein to intercept different steps of viral life cycle. The target genes were downregulated individually as well as in combinations and their impact on viral replication was evaluated. Individual downregulation of La autoantigen, PSMA7, hVAP-A, and NS5B resulted in inhibition of HCV replication by about 67.2%, 50.7%, 39%, and 52%, respectively. However, antiviral effect was more pronounced when multiple genes were downregulated simultaneously. Combinations of siRNAs against La autoantigen with NS5B or hVAP-A resulted in greater inhibition in HCV replication. Our findings indicate that siRNA is a potential therapeutic tool for inhibiting HCV replication and simultaneously targeting multiple viral steps with the combination of siRNAs is more effective than silencing a single target.
Hepatitis C virus (HCV) is a serious global health problem affecting approximately 130-150 million individuals. Presently available direct-acting anti-HCV drugs have higher barriers to resistance and also improved success rate; however, cost concerns limit their utilization, especially in developing countries like India. Therefore, development of additional agents to combat HCV infection is needed. In the present study, we have evaluated anti-HCV potential of water, chloroform, and methanol extracts from roots of Valeriana wallichii, a traditional Indian medicinal plant. Huh-7.5 cells infected with J6/JFH chimeric HCV strain were treated with water, chloroform, and methanol extracts at different concentrations. Semi-quantitative reverse transcription polymerase chain reaction result demonstrated that methanolic extract showed reduction in HCV replication. The methanolic extract was fractionated by thin layer chromatography, and the purified fractions (F1, F2, F3, and F4) were checked for anti-HCV activity. Significant viral inhibition was noted only in F4 fraction. Further, intrinsic fluorescence assay of purified HCV RNA-dependent RNA polymerase NS5B in the presence of F4 resulted in sharp quenching of intrinsic fluorescence with increasing amount of plant extract. Our results indicated that methanolic extract of V. wallichii and its fraction (F4) inhibited HCV by binding with HCV NS5B protein. The findings would be further investigated to identify the active principle/lead molecule towards development of complementary and alternative therapeutics against HCV. Copyright © 2017 John Wiley & Sons, Ltd.
Background: Staphylococcus aureus, the most common pathogen in skin and soft tissue infections (SSTI), harbors many well-characterized virulence genes. However, the expression of many of them in SSTIs is unknown. In this study, S. aureus virulence genes expressed in SSTI were investigated. Methods: Fifty-three subjects presenting to the outpatient’s care and emergency departments with a purulent SSTI at two medical centers in Wisconsin, USA, were enrolled in the study. Total mRNA was extracted from the purulent or swab materials, made into cDNA and sequenced on MiSeq platform. The relative cDNA counts to gmk and identifications of the transcripts were carried out with respect to USA300 reference genome and using SAMTOOLS v.1.3 and BWA, respectively. Result: A significantly higher cDNA count was observed for many of the virulence and regulatory gene transcripts in the pus samples compared to the swab samples relative to the cDNA counts for gmk, a housekeeping gene. They were for lukS-PV (18.6 vs. 14.2), isaA (13.4 vs. 8.5), ssaA (4.8 vs. 3.1), hlgC (1.4 vs. 1.33), atl (17.7 vs. 8.33), clfA (3.9 vs. 0.83), eno (6.04 vs. 3.16), fnbA (5.93 vs. 0.33), saeS (6.3 vs. 1.33), saeR (5.4 vs. 3.33) and agrC (5.6 vs. 1.5). Conclusions: A relative increase in the transcripts of several toxins, adhesion and regulatory genes with respect to a gmk in purulent materials suggests their role in situ during SSTIs, perhaps in an orchestrated manner.
Different cell types and/or tissues may serve as a reservoir for HIV-1 during ART-induced viral suppression. We compared proviral pol and env sequences from CD3 + T cells and CD14 + macrophage lineage cells from brain and nonbrain tissues from two virally suppressed individuals. We found strong evidence of viral population structure among cells/tissues, which may result from distinct selective pressures across cell types and anatomic sites.
Background: Dengue is the most rapidly spreading mosquitoborne viral disease in the world with an estimated 50 million dengue infections occur annually. India is one of the seven identified countries in the South-East Asia region regularly reporting incidence of DF/DHF outbreaks and becoming major niche for dengue infection.Methods & Materials: History, clinical examination findings, laboratory data, treatment details and outcome of patients with dengue fever was collected prospectively.SD BIO LINE DENGUE DUO (STANDARD DIGNOSTICS, KOREA) kits were utilized to diagnose the dengue in fection. This is a rapid diagnostic test which can detect simultaneously NS1 and antibodies of either Types (Ig M and Ig G). Patients were categorized into Undifferentiated DF, Dengue with Warning Signs and Severe Dengue according to WHO newer grading.Results: Total 148 patients were included in the study. Among them 85(57%) were male. There were 15(10%)in Undifferentiated DF, 45(31%) in DF with warning signs, 88(59%) in severe Dengue respectively. Mean age of our study population was 32.75 ± 14.99, 33.43 ± 14.95 respectively. Mean hospital stay in study population was 6.34 ± 4.66 days. Rash was found in 47 (37.76%), hepatomegaly in 23 (15%) and splenomegaly seen in 25(16%) patients. Bleeding manifestations were seen 61(41%) patients in total study group, out of which malena was seen in 33(22%)patients, gum bleeding, hematuria, hematemesis, hemoptysis and epistaxis seen in 6(4%), 7(5%), 4(3%), 3(2%) and 3(2%) patients respectively. Intracranial bleed as bleeding manifestation was seen 2(1%) patients one SDH and one case of intracerebral bleed, both non traumatic. Mean hemoglobin (g/dl) for was 13.5 ± 2.6g/dl. Hypoalbuminemia (<3.5 g/dl) was seen in 62(42%) patients with the mean value of 2.8 ± g/dl. Chest roentgenographic abnormalities found in 38(26%) patients of the study population. Pleural effusion was found in 31(19%) patients. Acalculous cholecystitis on ultrasonogram was seen in 18(12%) patients in our study population. NS1 antigen was detected in 80 (54%). Mortality was noted in 3.3% of the study population. Conclusion:Out of 148 patients, majority consisted of severe dengue infection. Elevated transaminases were seen in 136 (92%) patients. Mortality was noted in 5(3.3)% of the study population.
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