Computer-aided identification of Mycobacterium tuberculosis resuscitation-promoting factor B (RpfB) inhibitors from Gymnema sylvestre natural products.

Tuberculosis (TB), an infectious disease caused by multi-drug resistant Mycobacterium tuberculosis (Mtb), has been a global health concern. Mtb affects over a third of the world's population, causing two million deaths annually due to its dormancy and propensity to spread infection during this period. Resuscitation-promoting factor B (RpfB) plays a pivotal role in the growth of Mtb during dormant periods, making it a critical target for eliminating Mtb and curing TB. Gymnema sylvestre is a famous medicinal plant with several medicinal properties, including antimicrobial activity; however, the therapeutic potential of the various reported metabolites of this plant against Mtb has not yet been explored. The aim of this study was to explore the reported natural products of G. sylvestre against the RpfB of the Mtb. A total of 131 reported secondary metabolites of this plant were collected and virtually screened against the RpfB. We particularly targeted the Glu292 residue of RpfB as it is crucial for the catalysis of this protein. From our in-house library, 114 compounds showed a binding affinity higher than the standard drug. The binding stability of the top three lead compounds was further confirmed through MD simulation analysis. Drug likeness analyses indicated that the ten hits had zero violations of the Lipinski rule of five. In addition, analyses of pharmacokinetics, toxicity, and target prediction revealed that the top compounds are devoid of toxicity and do not affect human proteins. Additionally, they reflect multifaceted approach as anti-TB agents. Our selected hits not only exhibit molecular properties favoring physiological compatibility but also exhibit properties enhancing their potential efficacy as therapeutic candidates. The compounds investigated here are worthy of experimental validation for the discovery of novel treatments against TB. Further, this study also provides a promising avenue for research on the pharmacological potential of G. sylvestre.

Enhanced Efficacy of Direct-Acting Antivirals in Hepatitis C Patients by Coadministration of Black Cumin and Ascorbate as Antioxidant Adjuvants.

The widespread adaptation of a new generation of direct-acting antiviral agents (DAAs) unveils a superlative effect in the eradication of the hepatitis C virus (HCV). However, this therapy has been reported to exhibit vigorous side effects that pose a risk in fleet recovery. This study was conducted to investigate the efficacy of DAAs: sofosbuvir (SOF) and ribavirin (RBV), along with black cumin (BLC) and ascorbate (ASC), as adjuvants on hematological parameters; oxidative stress markers such as total antioxidant status (TAS), superoxide dismutase (SOD), reduced (GSH) and oxidized (GSSG) glutathione (GSH), gamma-glutamyl transferase (GGT), and malondialdehyde (MDA); liver function markers such as aspartate transaminase (AST), alanine aminotransferase (ALT), bilirubin, and alkaline phosphatase (ALP); and viral load with determined genotypes. HCV-infected patients (n = 30) were randomly divided into two equal groups: control group (n = 15) and treatment group (n = 15). The control group was subjected only to SOF and RBV (400 mg each/day). Synergistically, the treatment group was administered with adjuvant therapy of BLC (250 mg/day) and ASC (1000 mg/day) along with DAAs (400 mg each/day) for 8 weeks. All selected patients were subjected to sampling at pre- and posttreatment stages for the assessment of defined parameters. The data revealed that the BLC/ASC adjuvant therapy boosted the efficacy of DAAs by reducing the elevated levels of liver markers such as AST, ALT, ALP, and bilirubin in the treatment group compared with those in the control group (P > 0.05). The adjuvant therapy synchronously showed an ameliorating effect on hematological parameters. The SOF/RBV with adjuvant therapy also demonstrated an increasing effect in the activity of SOD, TAS, and GSH and a decreasing effect for GSSG, GGT, and malondialdehyde (MDA; P > 0.05) followed by curtailing a RT-PCR-quantified viral load. Our findings provide evidence that systemic administration of BLC/ASC efficiently alleviates hematological, serological, and antioxidant markers as well as the viral load in hepatitis C patients. This highlights a potentially novel role of BLC and ASC in palliating hepatitis C.

[Effects of lingqijuangan capsule medicated serum on apoptosis of activated rat hepatic stellate cells].

OBJECTIVE: To investigate the effects of LingQiJuanGan Capsule medicated serum on the apoptosis of and the expression of Bcl-2/Bax in hepatic stellate cell-T6 (HSC-T6) activated by platelet-derived growth factor (PDGF). METHODS: Preparation of drug-medicated serum. 30 SD rats were randomly divided into three groups. Serum was obtained from the SD rats administrated intragastricly with saline (10 mL/kg, group A), Fufang Biejia Ruangan Tablet solution (1.5 g/kg, group B) and LingQiJuanGan Capsule solution respectively (4.25 g/kg, group C). Detection of cell apoptosis and the expression of Bcl-2/Bax. HSC-T6 cells were activated by 10 ng/mL PDGF and incubated with drug-medicated serum at 200 mL/L. Cell apoptosis and the expression of Bcl-2/Bax were measured by flow cytometry at 0, 12, 24 and 48 h. RESULTS: Significantly higher apoptotic rates of HSC-T6 in group B and group C were detected at 12, 24, 48 h compared with that of group A (P < 0.05), wheraus no significant difference was found between group B and group C (P > 0.05). With increase of exposure time, the apoptotic rates of HSC-T6 in group B and group C increased (P < 0.05). The levels of Bcl-2 expression in HSC-T6 in group B and C were lower than that of group A (P < 0.05) at 12, 24, 48 h (No significant difference between group B and group C, P > 0.05). The expression level of Bcl-2 decreased with increased exposure time (P > 0.05) in both group B and group C. However, no such trend was observed in group A (P > 0.05). In both group B and group C, the expression level of Bax increased in a time-dependent manner. CONCLUSION: LingQiJuanGan Capsule can induce apoptosis of PDGF activated HSC-T6 and inhibit hepatic fibrosis in a time-dependent manner by influencing the expression of Bcl-2/Bax.

[Effects of Ling Qi Juan Gan capsule drug-containing serum on PDGF-induced proliferation and JAK/STAT signaling of HSC-T6 cells].

OBJECTIVE: To analyze the effects of Ling Qi Juan Gan capsule drug-containing serum at different concentrations on the platelet-derived growth factor (PDGF)-induced proliferative capabilities of and JAK2 and p-STAT3 protein expression in hepatic stellate cells (HSC) using an in vitro system. METHODS: Twenty-five Sprague-Dawley rats were randomly divided into five equal groups for intragastric administration of physiological saline (10 ml/kg; group A), Fufang Biejia Ruangan tablet solution (1.5 g/kg; group B), or Ling Qi Juan Gan capsule solution at low dose (2.125 g/kg; group C1), mid dose (4.25 g/kg; group C2), or high dose (8.5 g/kg; group C3). The post-administration serum isolated from each rat (200 ml/L) was used to treat the HSC-T6 cell line following induction by PDGF (10 ng/ml). At 24, 48 and 72 h post-exposure, the cells' proliferation was measured using the Cell Counting Kit-8 (CCK-8) colorimetric assay. In addition, at 24 h post-exposure the expression of JAK2 and p-STAT3 was measured by western blotting (expressed as grey scale intensity). Multiple group comparison of repeated measures data was made by one-way ANOVA with Student-Newman-Keuls post hoc test. RESULTS: Compared to group A, groups C2 and C3 had significantly higher inhibited proliferation at all post-exposure time points examined (24 h: A = 1.550 +/- 0.065, C2 = 1.335 +/- 0.106, C3 = 1.241 +/- 0.205; 48 h: A = 1.311 +/- 0.650, C2 = 1.090 +/- 0.106, C3 = 0.909 +/- 0.191; 72 h: A = 1.039 +/- 0.103, C2 = 0.719 +/- 0.116, C3 = 0.641 +/- 0.110, F = 36.292, all P less than 0.05); in contrast, compared to group A, group C1 showed no inhibition of proliferation at 24 h (1.522 +/- 0.128, P = 0.717) but showed significantly higher inhibition of proliferation at 48 h and 72 h (1.153 +/- 0.183 and 0.753 +/- 0.210, respectively, F = 36.292, P less than 0.05). Compared with group A, all Ling Qi Juan Gan capsule-containing serum-treated groups showed significantly lower expression of both JAK2 (A = 1.605 +/- 0.024 vs. C1 = 1.170 +/- 0.042, C2 = 0.842 +/- 0.036, C3 = 0.555 +/- 0.036, F = 43.091) and p-STAT3 (A = 1.401 +/- 0.030 vs. C1 = 1.229 +/- 0.025, C2 = 0.668 +/- 0.034, C3 = 0.630 +/- 0.026, F = 78.426, all P less than 0.01). CONCLUSION: Ling Qi Juan Gan capsule drug-containing serum can inhibit the proliferation of HSC-T6 cells in a dose-dependent manner and cause an overall decrease in the expression of JAK2 and p-STAT3 in activated HSC, thereby leading to a suppression of the JAK/STAT signal transduction pathway.

[Role and mechanism of Salvia miltiorrhiza Bunge in alleviating injury induced by hypoxia/reoxygenation in HK-2 cells].

OBJECTIVE: Salvia miltorrhiza Bunge (SMB) is a traditional Chinese herb, which is considered to promote blood flow and remove blood stasis. This study examined whether SMB can alleviate injury induced by hypoxia/reoxygenation (H/R) in human kidney proximal tubular cells-2 (HK-2 cells). METHODS: There were 3 experimental groups: control, H/R injury and SMB-treated H/R injury. H/R injury of HK-2 cells was induced by first covering the cells with and then removing liquid paraffin wax. Different concentrations of compound SMB solution (0.05%, 0.10%, 0.15% or 0.20%) were administered to the SMB-treated H/R injury group before the hypoxic injury. After 4, 12 and 24 hrs of hypoxia and 4, 12, 24 and 48 hrs of reoxygenation, morphologic changes of HK-2 cells were observed under an inverted microscope. Cell viability was measured by the MTT method. Lactate dehydrogenase (LDH) activity in the culture supernatants was assayed using biochemical methods; TNF-alpha levels were determined by radioimmunoassay (RIA). RESULTS: The number of HK-2 cells was significantly reduced in the H/R injury group after hypoxia, and reached a nadir 24 hrs after hypoxia treatment. Various concentrations of SMB-treated groups showed significantly greater number of HK-2 cells than the H/R injury group. SMB solution (0.10%) produced the best effect. The levels of LDH and TNF-alpha in the H/R injury group were significantly increased, and reached a peak between 24 hrs of hypoxia and 4 hrs of reoxygenation when compared to the control group. Pre-treating with 0.10% SMB resulted in significantly lower levels of LDH and TNF-alpha than in the untreated H/R injury group at various time points of H/R. CONCLUSIONS: SMB has protective effects against H/R injury of HK-2 cells, possibly through inhibition of inflammatory cytokines.