AYUSH-64 modulates Th17/Treg balance to attenuate LPS-induced inflammation in cell-based assays and BALB/c mice model

Abstract

AYUSH-64, a polyherbal formulation, has demonstrated therapeutic utility in managing several inflammatory diseases over the past three decades. Notably, during the COVID-19 pandemic, randomized clinical trials highlighted its efficacy in treating mild to moderate disease. Strong immunity is crucial for maintaining health and providing robust protection against infections and diseases. This study aimed to adopt an evidence-based approach to evaluate the anti-inflammatory and immunomodulatory potential of AYUSH-64 through various pharmacological models. Several in vitro cell-based assays were employed to assess the effects of AYUSH-64, using lipopolysaccharide (LPS)-stimulated human peripheral blood mononuclear cells (PBMCs), differentiated THP-1 cells, and A549 adenocarcinoma cell lines. Additionally, LPS-induced lung inflammation in a BALB/c mouse model was used to evaluate its anti-inflammatory potential in vivo. The in vitro experiments revealed that LPS stimulation (1 µg/ml) significantly increased the expression of pro-inflammatory cytokines, including TNF-α, IL-6, and IL-8, in PBMCs, differentiated THP-1 cells, and A549 cells. AYUSH-64, at concentrations of 10 mg/ml, 3 mg/ml, and 1 mg/ml, reduced the expression of these cytokines in a dose-dependent manner. Furthermore, AYUSH-64 suppressed LPS-induced IL-17 levels in PBMCs, while enhancing the regulatory T cell (Treg) population and FoxP3 expression, suggesting a modulatory effect on immune response. In the in vivo model, AYUSH-64 at 100 mg/kg and 30 mg/kg significantly reduced neutrophil infiltration in BALB/c mice challenged intranasally with LPS (25 µg in 200 µL PBS per mouse). This was accompanied by improved lung pathophysiology and reduced pro-inflammatory cytokine levels in lung tissues. These findings underscore the distinct anti-inflammatory effects of AYUSH-64 across all studied models. Moreover, treatment with AYUSH-64 resulted in an increased Treg population and elevated FoxP3 levels in PBMCs, indicating its immunoregulatory potential. In conclusion, AYUSH-64 demonstrates promise as a herbal therapeutic for managing inflammation and enhancing immunity, warranting further investigation for its potential applications in human health.