〔Abstract〕 To investigate the effect of phosphatase and tensin homolog (PTEN) overexpression on au- tophagy in mouse primary cardiac fibroblasts . Methods Neonatal mice ( 1 - 3 days old) were purchased and sub- jected to cardiac tissue harvesting. Cardiac fibroblasts were isolated through enzymatic digestion and cultured . After cellular adhesion , rapamycin (Rapa)-treated cardiac fibroblasts were used to establish an autophagy model . Fol- lowing successful model construction , cells were transfected with either PTEN-overexpressing plasmid (PTEN over- expression group) or empty vector (control group) , followed by 24 - 48 h incubation . The molecular expressions of PTEN , autophagy-related proteins ( microtubule-associated protein light chain 3 [ LC3 Ⅱ/ Ⅰ ] , Beclin1) , and fi- brotic marker collagen Ⅰ were detected by Western blot and RT-qPCR. Autophagic flux was assessed using mCher- ry-GFP-LC3 fluorescence staining to evaluate changes in autophagic activity. Finally , transmission electron micros- copy (TEM) was employed to observe autophagosome formation in cardiac fibroblasts . Results In the Rapa-in- duced cardiac fibroblast autophagy model , compared with the control group , the protein and mRNA levels of PTEN significantly decreased (P < 0. 05) , while the expression of autophagy-related proteins (LC3 Ⅱ/ Ⅰ , Beclin1) and fibrotic marker Collagen Ⅰ was upregulated at both protein and mRNA levels (P < 0. 05) . Additionally , in PTEN- overexpressing cardiac fibroblasts , the expression levels of LC3 Ⅱ/ Ⅰ , Beclin1 , and Collagen Ⅰ were markedly re- duced compared to the empty vector group (P < 0. 05) . mCherry-GFP-LC3 fluorescence staining demonstrated that autophagic activity was significantly attenuated in the PTEN overexpression group versus the empty vector group (P < 0. 05) . TEM further revealed a decreased number of autophagosomes in PTEN-overexpressing cardiac fibroblasts (P < 0. 05) . Conclusion The overexpression of PTEN significantly inhibits autophagy in cardiac fibroblasts , sug- gesting that PTEN may be a key gene involved in regulating autophagy in cardiac fibroblasts .