Poster Presentation Lorne Infection and Immunity 2018

Examination of the role of activin A and resident testicular cell types in fibrosis following experimental autoimmune orchitis (EAO) in mice (#136)

Christine A Kleinert 1 2 , Nour Nicolas 1 2 , Sudhanshu Bhushan 1 , Eva Wahle 1 , Kate L Loveland 2 3 , Andreas Meinhardt 1 2 , Mark P Hedger 2 , Monika Fijak 1
  1. Anatomy and Cell Biology, Justus Liebig University, Giessen, Hessen, Germany
  2. Hudson Institute of Medical Ressearch, Clayton, VIC, Australia
  3. Department of Molecular and Translational Sciences, School of Clinical Sciences, Monash University, Melbourne, Victoria, Australia

Testicular inflammation is a cause of male subfertility that commonly features severe fibrosis. Experimental autoimmune orchitis (EAO) in mice models this immunopathology, but the cells responsible for production of extracellular matrix components (ECMC), leading to fibrosis in the EAO testis, are unknown. Proliferation and activation of resident fibroblasts, the myoid peritubular cells (PTC), or monocyte-to-fibroblast transition (MFT) are all potential sources. Activins are members of the transforming growth factor-β superfamily that regulate cell growth and differentiation, as well as inflammatory and fibrotic responses in disease. They control spermatogenesis and steroidogenesis within the normal testis, but activin A levels increase substantially during the development of EAO.

 

In order to investigate the potential cellular origins and regulation of ECMC-producing cells in EAO, we measured fibrotic gene expression by qRT-PCR and Western blot in PTC from 21 day-old mice and NIH 3T3 mouse fibroblasts treated with human recombinant activin A (25 and 50 ng/ml), the activin-binding protein, follistatin-288 (FST288; 100 ng/ml), or a combination of both. Localization of the macrophage marker, F4/80, and collagen I was also investigated by immunofluorescence in adult mouse testis sections following induction of EAO.

 

Treatment with activin A increased the level of fibronectin mRNA, and production of collagen type I and fibronectin in both 3T3 fibroblasts and PTC. Activin A also increased alpha-smooth muscle actin mRNA and protein expression in 3T3 fibroblasts, while collagen type IV mRNA levels were significantly upregulated in PTC. FST288, a potent antagonist of activin A, significantly inhibited these effects of activin A. Immunofluorescence identified collagen I-positive testicular macrophages in the EAO testis, indicating the occurrence of MFT. These data indicate a role for activin A in the fibrotic response to EAO and suggest that intratesticular PTC, fibroblasts and monocytes all contribute to the development of fibrosis under activin control.