Targeting TGF-β1/MMP-9/α-SMA Signaling Axes Using Piperine and Galangin in Bleomycin Induced Pulmonary Fibrosis Model
K. VANITHA SREE *
Administrative Office, PVNRTVU, Hyderabad, India.
M. USHA RANI
CVSc, Mamnoor, India.
A. GOPALA REDDY
CVSc, Rajendra Nagar, India.
M. LAKSHMAN
CVSc, Rajendra Nagar, India.
B. RAJENDER
CVSc, Rajendra Nagar, India.
D. DURGA VEERA HANUMAN
CVSc, Rajendra Nagar, India.
K V VENKATA RAO
CVSc, Garividi, India.
DIPALI P. PATIL
COVAS, Parbhani, M S., India.
*Author to whom correspondence should be addressed.
Abstract
Aims: To evaluate and compare the anti-fibrotic efficacy of galangin and piperine alone and in combination against bleomycin-induced pulmonary fibrosis in C57BL/6 mice, using biochemical (hydroxyproline, TGF-β1), histological (Masson’s trichrome), and immunohistochemical (MMP-9, α-SMA) endpoints.
Study Design: Controlled, randomized, experimental animal study with seven parallel groups.
Place and Duration of Study: Department of Veterinary Pharmacology and Toxicology, College of Veterinary Science, Hyderabad, India. Experiments were conducted under IAEC approval using a 21-day protocol.
Methodology: Fifty male C57BL/6 mice were allocated into seven groups, including control, bleomycin, galangin, piperine, and their combinations. After 21 days of treatment, lung tissues were analyzed for hydroxyproline, TGF-β1, collagen (Masson’s trichrome), and MMP-9 and α-SMA (IHC). Data were statistically evaluated using one-way ANOVA followed by Tukey’s test (P < 0.05).
Results: BLM significantly increased hydroxyproline vs control (171.41±6.46 vs 27.35±2.57 pg/mg protein; P<0.05) and TGF-β1 (129.24±2.87 vs 16.61±2.09 pg/mg; P<0.05). Treatment reduced both biomarkers vs BLM: GA (hydroxyproline 110.87±2.84; TGF-β1 64.17±2.87), PIP (97.68±3.22; 58.73±2.56), and combination (65.82±5.22; 42.15±2.41), with the combination superior to monotherapies (all P<0.05). Per se GA and PIP (groups 3–4) did not differ from control for either marker (P>0.05). Masson’s trichrome showed marked collagen deposition in BLM lungs, attenuated to mild (GA, PIP) and very mild (combination) fibrosis. IHC demonstrated strong MMP-9 and α-SMA expression in BLM lungs, reduced with GA or PIP and minimal with combination therapy.
Conclusion: Galangin and piperine mitigate bleomycin-induced lung fibrosis, with combination therapy yielding the greatest improvement across biochemical, histological, and immunohistochemical endpoints. Benefits likely arise from dampening profibrotic signaling (TGF-β1/α-SMA/MMP-9) alongside antioxidant/anti-inflammatory actions, supporting these phytochemicals particularly in combination as promising candidates for anti-fibrotic intervention.
Keywords: Bleomycin, piperine, galangin, hydroxyproline, fibrosis