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Exosomes Derived from Human Umbilical Cord Mesenchymal Stem Cells Alleviates Oxalate and Calcium Oxalate Monohydrate-Induced Epithelial-Mesenchymal Transition in HK-2 Cells
Li Dian, He Dawei*, Tang Bo, Guo Wenhao, Zhang Dan, Shen Lianju, Wei Guanghui*
Department of Urology, Children’s Hospital of Chongqing Medical University, Ministry of Education Key Laboratory of Child Development and Disorders, China International Science and Technology Cooperation Base of Child Development and Critical Disorders, Chongqing Key Laboratory of Pediatrics, Chongqing Key Laboratory of Children Urogenital Development and Tissue Engineerin, Chongqing 400014, China
Abstract: Oxalate and calcium oxalate monohydrate (COM) was used to induce epithelial-mesenchymal transition (EMT) in human proximal renal tubular epithelial cells (HK-2)， meanwhile exosomes derived from human umbilical cord mesenchymal stem cells (hucMSC-Ex) were isolated by ultracentrifugation and applied into the induced HK-2 cells to study its relieving effects on fibrosis. Then Western blot， transmission electron microscope and nanoparticle tracking analysis (NTA) were used to identify the markers and morphology of exosomes. The effect of hucMSC-Ex on cell viability was observed by MTT. Elements in TGF-β/Smad signaling pathway， including TGF-β and Smad2， epithelial markers， including ZO-1 and E-cadherin， and mesenchymal markers， including N-cadherin and α-SMA changes， were evaluated by immunofluorescence and Western blot， respectively. The results showed that， after incubated with oxalate and COM crystals， the cell morphology was changed， the expression of E-cadherin and ZO-1 was down-regulated， while the expressions of N-cadherin， α-SMA and TGF-β， Smad2 were increased. The morphology of hucMSC-Ex showed a double-layer membrane， hollow， circular or elliptical. Alix， CD63 and TSG101 were positively expressed in hucMSC-Ex， their diameters were distributed between 80 nm and 300 nm. And hucMSC-Ex pretreatment can significantly improve the viability of HK-2 cells exposed to oxalate and COM crystals. Then， the expressions of N-cadherin， α-SMA and TGF-β， Smad2 in HK-2 cells were reduced， and the expressions of E-cadherin and ZO-1 were up-regulated. We conclude that hucMSC-Ex could alleviate EMT and reduce the injury of HK-2 cells induced by oxalate and COM crystals.
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