Cx31.1 expression is modulated in HaCaT cells exposed to UV-induced damage and scrape-wounding.

Connexin31.1 (Cx31.1) is a gap junction protein associated with apoptosis. In the skin, apoptosis is modulated by diabetes. A HaCaT skin model investigated whether normal (NGI) and high glucose and insulin (HGI; diabetic) conditions altered Cx31.1 expression, and if these were apoptosis linked. Cx31.1 was found in HaCaT and HeLa Ohio cells, with HaCaT Cx31.1 protein increased in HGI conditions, and around apoptotic cells. HeLa Cx31.1 channels were noncommunicative. Post scrape-wounding, Cx31.1 increased at wound edges. Caspase 3/7 in scrape-wounds media (containing cells) elevated in HGI. UV exposure raised Cx31.1, and caspase 3/7, in NGI and HGI. UV reduced cell viability in NGI cells, although not significantly in HGI. Cx31.1 is modulated during HaCaT cell wound closure, and associated with 'diabetic' conditions. Cx31.1 expression matched apoptosis levels, higher in HGI cultures. Cx31.1 is noncommunicating, modulated after wounding, linked to apoptosis, and may be associated with tissue turn-over around diabetic wounds.

Narrow Spectrum Kinase Inhibitors Demonstrate Promise for the Treatment of Dry Eye Disease and Other Ocular Inflammatory Disorders.

Purpose: The purpose of this study is to determine the potential of narrow spectrum kinase inhibitors (NSKIs) to treat inflammatory eye disorders. Methods: Human conjunctival epithelial (HCE) cells were retrieved from subjects via impression cytology. Real-time quantitative PCR (qPCR) was performed on HCE cells to determine gene expression of NSKI kinase targets and proinflammatory cytokines in dry eye disease (DED) patients versus healthy controls. qPCR also assessed p38α expression in hyperosmolar-treated Chang conjunctival epithelial cells. Interaction of NSKI TOP1362 with the kinases was evaluated in ATP-dependent Z-LYTE and competition binding assays. Anti-inflammatory activity was assessed in human peripheral blood mononuclear cells and primary macrophages. In an endotoxin-induced uveitis (EIU) study, lipopolysaccharide (LPS) was administered intravitreally to Lewis rats. TOP1362, dexamethasone, or vehicle was administered topically, and inflammatory cytokine levels were measured 6 hours after LPS injection. Results: HCE cells from DED patients showed significantly increased expression of p38α, spleen tyrosine kinase (Syk), Src, lymphocyte-specific protein tyrosine kinase (Lck), interleukin one beta (IL-1ß), interleukin eight (IL-8), monocyte chemotactic protein-1 (MCP-1), and matrix metalloproteinase-9 (MMP-9). TOP1362 strongly inhibited the kinase targets p38α, Syk, Src, and Lck, blocked the rise in p38α expression in hyperosmolar Chang cells, and potently reduced inflammatory cytokine release in cellular models of innate and adaptive immunities. In the EIU model, TOP1362 dose-dependently attenuated the LPS-induced rise in inflammatory cell infiltration and ocular cytokine levels with efficacy comparable to that of dexamethasone. Conclusions: TOP1362 is a potent inhibitor of kinases upregulated in DED and markedly attenuates proinflammatory cytokine release in vitro and in vivo, highlighting the therapeutic potential of NSKIs for treating ocular inflammation, such as that observed in DED.