Background: The pathogenesis of peritoneal dialysis (PD)-related peritoneal fibrosis (PF) is not clearly understood, and current treatment options are limited. Methods: In this study, the effect of PD-related PF on mitochondrial biogenesis was investigated, and the effect of activation of the adenosine monophosphate-activated protein kinase (AMPK)-PGC-1 alpha (peroxisome proliferator-activated receptor gamma coactivator-1 alpha) pathway on PF was evaluated in mice. Results: In a mouse model of PD-related PF, AMPK-PGC-1 alpha signaling (phospho-AMPK, PGC-1 alpha, NRF-1, NRF-2 and TFAM expression) was downregulated, mitochondrial DNA (mtDNA) levels were reduced, and mitochondrial structure was damaged in the peritoneum. In addition, TdT-mediated dUTP nick-end labeling (TUNEL) staining showed typical apoptosis characteristics in peritoneal mesothelial cells (PMCs). Activation of the AMPK-PGC-1 alpha pathway (PGC-1 alpha overexpression or metformin, which is an agonist of AMPK) upregulated phospho-AMPK, PGC-1 alpha, nuclear respiratory factors 1 (NRF-1) and 2 (NRF-2), and mitochondrial transcription factor A (TFAM) expression and mtDNA content, improved mitochondrial morphological manifestations, inhibited apoptosis of PMCs and alleviated PF. Conclusion: Our study may suggest that activation of the AMPK-PGC-1 alpha pathway ameliorates PD-related PF by enhancing mitochondrial biogenesis.