ABSTRACT
Objective: To evaluate the antiviral activity and phytochemicals of selected plant extracts and their effect on the mitogen-activated protein kinase (MAPK) signaling pathway modulated by hepatitis C virus (HCV) nonstructural protein 5A (NS5A). Methods: A total of ten plant extracts were initially screened for their toxicities against HepG2 cells. The non-toxic plants were tested for their inhibitory effect on the expression of HCV NS5A at both mRNA and protein levels using real-time PCR and Western blotting assays, respectively. The differential expression of the genes associated with MAPK pathway in the presence of NS5A gene and plant extract was measured through real-time PCR. Subsequently, the identification of secondary metabolites was carried out by phytochemical and HPLC analysis. Results: The phytochemical profiling of Berberis lyceum revealed the presence of alkaloids, phenols, saponins, tannins, flavonoids, carbohydrates, terpenoids, steroids, and glycosides. Similarly, quercetin, myricetin, gallic acid, caffeic acid, and ferulic acid were identified through HPLC analysis. The methanolic extract of Berberis lyceum strongly inhibited HCV RNA replication with an IC50 of 11.44 μg/mL. RT-PCR and Western blotting assays showed that the extract reduced the expression of HCV NS5A in a dosedependent manner. Berberis lyceum extract also attenuated NS5Ainduced dysregulation of the MAPK signaling pathway. Conclusions: Our findings suggest that Berberis lyceum extract strongly inhibits HCV propagation by reducing HCV NS5Ainduced perturbation of MAPK signaling.
ABSTRACT
Objective: To evaluate the antiviral activity and phytochemicals of selected plant extracts and their effect on the mitogen-activated protein kinase (MAPK) signaling pathway modulated by hepatitis C virus (HCV) nonstructural protein 5A (NS5A). Methods: A total of ten plant extracts were initially screened for their toxicities against HepG2 cells. The non-toxic plants were tested for their inhibitory effect on the expression of HCV NS5A at both mRNA and protein levels using real-time PCR and Western blotting assays, respectively. The differential expression of the genes associated with MAPK pathway in the presence of NS5A gene and plant extract was measured through real-time PCR. Subsequently, the identification of secondary metabolites was carried out by phytochemical and HPLC analysis. Results: The phytochemical profiling of Berberis lyceum revealed the presence of alkaloids, phenols, saponins, tannins, flavonoids, carbohydrates, terpenoids, steroids, and glycosides. Similarly, quercetin, myricetin, gallic acid, caffeic acid, and ferulic acid were identified through HPLC analysis. The methanolic extract of Berberis lyceum strongly inhibited HCV RNA replication with an IC50 of 11.44 μg/mL. RT-PCR and Western blotting assays showed that the extract reduced the expression of HCV NS5A in a dosedependent manner. Berberis lyceum extract also attenuated NS5Ainduced dysregulation of the MAPK signaling pathway. Conclusions: Our findings suggest that Berberis lyceum extract strongly inhibits HCV propagation by reducing HCV NS5Ainduced perturbation of MAPK signaling.
ABSTRACT
Objective: To compare the efficiency of conventional diagnostic techniques and Insertion Sequence [IS]6110 based PCR assay for M. tuberculosis in pulmonary and extra-pulmonary specimens from tertiary care chest hospital
Study Design: Observational study
Place and Duration of Study: This study was conducted at Gulab Devi Chest Hospital, Lahore from August to January 2013
Materials and Methods: A total of 1599 [1417 pulmonary and 182 extra-pulmonary] non-duplicate clinical specimens, obtained over a period of six months, were tested by conventional techniques such as Ziehl-Neelsen staining [ZN], Lowenstein Jensen [LJ] medium and Fluorescent staining. MTB was extracted through DNAzol method. Insertion Sequence [IS] 6110 based PCR assay was used for M. tuberculosis from pulmonary and extra-pulmonary specimens. Of the 1599 specimens, 781 were suspect cases while 818 were MDR [follow up] cases. Mean age of TB patient was +/- 33 years. 18% of follow-ups and 20% of suspects were <20 year in age, 52% follow-ups and 36% suspects were about 20-40 years, and 30% follow-ups and 33% suspects were >40 years ofage
Results: It was seen that, among MDR cases [follow-ups] 68% were males and 32% were females. Similarly,among TB-suspects, 58% were males and 42% were females. Of total 168 suspected pulmonary samples ZN [48.2.7%], fluorescent microscopy [79.7%], LJ culture [52.9%] and PCR [91.6%] were positive for M. tuberculosis. In total 143 suspected extra-pulmonary samples, ZN [34.95%], fluorescent microscopy [45.5%], LJ culture [39.8%] and PCR [87.4%] werepositive
Conclusion: In contrast to conventional methods of TB diagnosis, PCR is more quick, sensitive, reliable and cost effective technique