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Effects of Heibu Ointment on the growth and mitochondrial apoptosis of keloid fibroblasts under hypoxia
Ma Hui, Li Linchang, Zhang Tianlei, Chen Weiwen, Zheng Haiyun
2026, 23(7):
775-782.
DOI: 10.19803/j.1672-8629.20250922
Objective To investigate the effects of Heibu ointment on the proliferation, migration, apoptosis and expressions of hypoxia-inducible factor-1α (HIF-1α), and key proteins of the mitochondrial apoptotic pathway in keloid fibroblasts (KFs) under hypoxic conditions. Methods A human KFs cell line was cultured and divided into the following groups: normoxia, hypoxia, and hypoxia treated with Heibu ointment at concentrations of 500, 250, 125, 62.5, and 31.25 μg·mL-1. After 24 hours of culture, the MTT assay was performed to evaluate the proliferation of KFs and to select the optimal concentration for subsequent experiments. The KFs were then regrouped into normoxia, normoxia + 125 μg·mL-1 extract, hypoxia, and hypoxia + 125 μg·mL-1 extract. MTT assay, flow cytometry, scratch wound assay, immunofluorescence, and Western blot were used to evaluate the effects of the extract on the proliferation, apoptosis, migration and HIF-1α expression of KFs, and on the protein levels of key mitochondrial apoptosis factors (Bax, Bcl-2, Caspase-3). Results Compared with normoxic conditions, hypoxia significantly enhanced KFs’ proliferation (P<0.01), suppressed apoptosis (P<0.01), and promoted cell migration (P<0.01) while upregulating both the fluorescence intensity and protein expression of HIF-1α (P<0.01). Within the mitochondrial apoptotic pathway, hypoxia downregulated the expression levels of Bax and Caspase-3 (P<0.01) while upregulating Bcl-2 expressions (P<0.01). Treatment with Heibu ointment inhibited KFs’ proliferation (P<0.01), increased the apoptosis rate (P<0.01), and reduced migratory capacity P<0.01) compared to the hypoxia alone group. Concurrently, this ointment decreased both HIF-1α fluorescence intensity and protein expressions (P<0.01), elevated the expression levels of Bax and Caspase-3, but lowered Bcl-2 expression (P<0.01). Conclusion Our findings indicate that Heibu ointment can exert anti-scarring effects by inhibiting proliferation/migration, inducing apoptosis, downregulating HIF-1α, and activating the mitochondrial apoptotic pathway in hypoxic KFs.
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