Effect of polystyrene micro/nanoplastics on mesenchymal phenotypic transformation in testicular Sertoli cells

To investigate the effects and potential mechanisms of polystyrene micro/nanoplastics (PS-MNPs) on testicular Sertoli cells.

Sixty male C57BL/6N mice (8 weeks old) were randomly divided into a control group (deionized water), a PS-NPs group [particle size of 20 nm, 2. 5 mg/(kg·d)], and a PS-MPs group [particle size of 5 μm, 2. 5 mg/(kg·d)], with 20 mice in each group. The corresponding agents were gavaged once daily for 6 months. HE staining was used to observe the histopathological and morphological changes in the testicular tissues. Immunohistochemistry of marker proteins was employed to evaluate the changes in the number of Sertoli cells. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were performed to identify functions and signaling pathways enriched in the testicular transcriptome. Mouse testicular Sertoli cell line TM4 was divided into a control group (deionized water), a 2. 5NPs group (2. 5 μg/mL), and a 2. 5MPs group (2. 5 μg/mL). All groups received continuous exposure through 130 cell passages. Cell viability and proliferative capacity were evaluated using CCK-8 assay and EdU incorporation, while cell migration was assessed using transwell and cell scratch assays. RT-qPCR and Western blotting were used to detect the changes in the expression of key molecules regulating mesenchymal phenotypic transformation (MPT) at mRNA and protein levels.

Pathological analysis revealed that, when compared to the control group, PS-NPs and PS-MPs treatment resulted in extended spaces between testicular seminiferous tubules, loosely arranged spermatogenic cells, and enhanced vacuolization. Immunohistochemical analysis of marker proteins indicated a decreasing trend in the number of testicular Sertoli cells in the PS-NPs and PS-MPs groups than the control group, with the PS-NPs group having statistical significance (P<0. 01). GO and KEGG enrichment analyses revealed that PS-MNPs exposure-related altered genes were significantly enriched in cell adhesion signaling pathways (P<0. 05). PS-MPs exposure significantly inhibited the growth and migration ability of TM4 cells (P<0. 05), but PS-NPs exposure had no such effect on cell growth but notably enhanced cell migration ability. PS-NPs exposure inhibited the expression levels of E-cadherin and ZO-1 (P<0. 01) and up-regulated the expression of N-cadherin and vimentin (P<0. 01), and PS-MPs exposure led to significant up-regulation of vimentin (P<0. 01) and down-regulation of E-cadherin, N-cadherin, and ZO-1 (P<0. 05). Both PS-MPs and PS-NPs exposure up-regulated the mRNA levels of Snail2, Twist1, and Zeb2 (P<0. 01).

Exposure of PS-MNPs leads to abnormal proliferation and migration of TM4 cells, induces decreases in cell-cell contacts among Sertoli cells and spermatogenic cells at all levels possibly through MPT, and thus results in testicular damage.