Testisin/Prss21 deficiency causes increased vascular permeability and a hemorrhagic phenotype during luteal angiogenesis

Abstract

Testisin (encoded by PRSS21) is a membrane anchored serine protease, which is tethered to the cell surface via a glycosylphosphatidylinositol (GPI)-anchor. While testisin is found in abundance in spermatozoa, it is also expressed in microvascular endothelial cells where its function is unknown. Here we identify testisin as a novel regulator of physiological hormoneinduced angiogenesis and microvascular endothelial permeability. Using a murine model of rapid physiological angiogenesis during corpus luteal development in the ovary, we found that mice genetically deficient in testisin (Prss21 -/-) show a substantially increased incidence of hemorrhages which are significantly more severe than in littermate control Prss21 +/+ mice. This phenotype was associated with increased vascular leakiness, demonstrated by a greater accumulation of extravasated Evans blue dye in Prss21 -/ovaries. Live cell imaging of in vitro cultured microvascular endothelial cells depleted of testisin by siRNA knockdown revealed that loss of testisin markedly impaired reorganization and tubule-like formation on Matrigel basement membranes. Moreover testisin siRNA knockdown increased the paracellular permeability to FITC-albumin across endothelial cell monolayers, which was associated with decreased expression of the adherens junction protein VE-cadherin and increased levels of phospho(Tyr658)-VE-cadherin, without affecting the levels of the tight junction proteins occludin and claudin-5, or ZO-1. Decreased expression of VE-cadherin in the neovasculature of Prss21 -/ovaries was also observed without marked differences in endothelial cell content, vascular claudin-5 expression or pericyte recruitment. Together, these data identify testisin as a novel regulator of VE-cadherin adhesions during angiogenesis and indicate a potential new target for regulating neovascular integrity and associated pathologies.

Intersection of Coagulation and Fibrinolysis by the Glycosylphosphatidylinositol (GPI)-Anchored Serine Protease Testisin

“…Testisin-deficient mice have no developmental defects and no obvious defects in hemostasis when observed under unchallenged conditions [7,12], suggesting a post-natal role in angiogenesis. In adult organisms, the vascular tree is fully developed, and physiological angiogenesis is normally limited to the female reproductive system.…”

“…Our previous studies using a limited panel of peptide substrates and recombinant testisin determined that testisin possesses a trypsin-like substrate specificity with a preference for cleavage after P 1 -Arg over P 1 -Lys [46]. Testisin is expressed by endothelial cells of various vascular beds and during capillary morphogenesis [10][11][12]. When challenged with hormones that induce rapid angiogenesis of the corpus luteum, testisin-deficient mice display defective microvascular development, resulting in increased hemorrhages, implicating testisin in protecting endothelial barrier integrity during angiogenesis.…”

“…A global microarray analysis of human endothelial cell diversity in 14 distinct vascular tissues also identified testisin as expressed consistently in both large vessels and the microvasculature [16]. In vitro siRNA knockdown studies showed that the loss of testisin markedly impairs microvascular endothelial cell migration and reorganization during Matrigel-induced angiogenesis [12]. In a recent study of the effect of testisin deletion on angiogenesis during corpus luteal development, we found that Prss21 −/− mice display a strikingly increased incidence and severity of hemorrhages, which were associated with impaired endothelial barrier formation and function [12].…”

“…The complete genetic deletion of testisin in Prss21 −/− mice causes defective epidydimal sperm maturation and reduced fertilizing ability compared to Prss21 +/+ littermate control mice [7][8][9]. Beyond sperm, testisin expression is extremely restricted in healthy individuals, being expressed at low levels in endothelial cells and some immune cells [4,[10][11][12][13], and it is overexpressed in ovarian cancers compared to normal ovary tissue [14,15]. Using a protease-gene-specific screen, we identified testisin expression in human dermal microvascular endothelial cells (HMVECs) undergoing reorganization and tubule-like formation on the basement membrane Matrigel, as well as during pre-capillary morphogenesis on fibrillar type I collagen [10].…”

scRNA-Seq and Bulk-Seq Analysis Identifies S100A9 Plasma Cells as a Potentially Effective Immunotherapeutic Agent for Multiple Myeloma

“…[55][56][57] PRSS21 encodes testosterone, which is anchored to the cell surface through glycosylphosphatidylinositol (GPI), inducing angiogenesis and regulating microvascular endothelial permeability. 58 IFI6, also known as interferon alpha-inducible protein 6, prevents virus-induced ER membrane invasion, impairs viral entry by inhibiting EGFR activation, modulates innate immune responses, and inhibits virus replication. 59 LYZ (Lysozyme) is an antibacterial protein present in various human tissues and fluids, capable of directly dissolving bacterial cell walls to exhibit antibacterial activity.…”