Objective To investigate the effect of elevated cyclic stretch induced by hypertension on mitochondrial biogenesis of vascular smooth muscle cells (VSMCs), and the role of PGC1α in this process. Methods The Flexcell-5000T stretch loading system in vitro was applied to VSMCs with a frequency of 1.25 Hz and an amplitude of 5% or 15% to simulate the mechanical environment under normal physiological or pathological conditions of hypertension respectively. Western blotting and qPCR were used to detect the expression of PGC1α, citrate synthase and mitochondrial DNA (mtDNA) copy number in VSMCs under normal physiological or hypertensive pathophysiological conditions. VSMCs was treated with PGC1α specific activator ZLN005 to promote PGC1α expression or specific interfering fragment siRNA to inhibite PGC1α expression and detect the effect on citrate synthase and mtDNA copy number. Results Compared with 5% physical cyclic stretch, 15% pathological elevated cyclic stretch significantly inhibited the expression of PGC1α, citrate synthase and mtDNA copy number in VSMCs, indicating that the mitochondrial biogenesis was impaired by pathological elevated cyclic stretch. Compared with the blank control group, the protein expression of PGC1α was significantly decreased and increased respectively, when VSMCs transfected with PGC1α siRNA or incubated PGC1α activator ZLN005, the expression of citrate synthase and mtDNA copy number were also significantly downregulated and upregulated in VSMCs accordingly, the results suggested that PGC1α regulated mitochondrial biogenesis in VSMCs. Under physiological cyclic stretch conditions, the protein level of PGC1α was significantly downregulated by PGC1α siRNA, which also significantly downregulated citrate synthase expression and mtDNA copy number. Under pathological cyclic stretch conditions, the protein expression of PGC1α was significantly upregulated by ZLN005, which also enhanced the expression of citrate synthase and mtDNA copy number. These results indicated that elevated cyclic stretch affects mitochondrial biogenesis by regulating PGC1α expression. Conclusions The pathological cyclic stretch induced by hypertension significantly downregulated the expression of citrate synthase and mtDNA copy number via suppressing the expression of PGC1α, resulting in mitochondrial dysfunction of VSMCs, suggesting that PGC1α may be a potential therapeutic target molecule to alleviate the progression of hypertension.