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Andrew T C Tsin, Brandi S Betts-Obregon, Robert Mortiz, Richard LeBaron; Novel Mechanism which Promotes Diabetic Complications in Renal and Ocular Systems. Invest. Ophthalmol. Vis. Sci. 2017;58(8):4036. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
To determine the roles of TGFB1 and TGFB2 receptors, as well as Tranforming Growth Factor Beta-Induced (TGFBI or BIGH3), in the deaths of renal and ocular cells in culture.
BIGH3 was introduced in to cell medium of cultured RPTEC, REC, and HRPs. Macrophages and macrophage derived TGF-B1/TGFB2 receptor blockers were also added at various concentrations. Cell viability was determined via TUNEL assay per manufacturers instructions.Renal cells (RPTEC) and Human Retinal Pericytes (HRP's) were purchased from Clonetics/Lonza (Walkersville, MD). Retinal Endothelial Cells (REC) were purchased from ATCC (Manassas, VA). Monocyte-derived Macrophage (MΦ) were prepared in accordance with established methodology (Wintergerst et al., 1998). Polyclonal anti-BIG-H3 was generated in house.
Our study here indicates that macrophages (MΦ) and MΦ-derived TGF-β1 promote BIGH3 Mediated Apoptosis (BMA), thus advancing nephrology and diabetic retinopathy diseases. In addition, we show that in diabetic conditions BMA targets three different cell types. In the retina BMA targets retinal pericytes (HRP) and retinal endothelial cells (RhREC), and in the kidney BMA targets renal proximal tubule epithelial cells (RPTEC). The BMA mechanistic pathway that kills these cells involves MΦ-derived TGF-β1, BIGH3 protein, BIGH3 C-terminal cleavage, C-terminal-derived integrin ligand peptides, and specific integrins expressed on renal, endothelial, and pericyte cell surfaces. After BIGH3 is secreted into the cells’ milieu, a crucial step in the BMA mechanism is cleavage of BIGH3’s C-terminus which can be accomplished by the peptidases plasmin, matrix metalloproteinase 9, and serine protease high-temperature requirement A1. BIGH3 C-terminal cleavage generates integrin-ligand peptides that manipulate targeted integrin signaling pathways to evoke BMA. We also found that cells in a pre-diabetic environment synthesize BIGH3 protein, indicating BIGH3 is a biomarker signifying a pre-diabetic state that left untreated will likely progress to complications including diabetic retinopathy and nephrology.
Identification of the MΦ source of BIGH3 in diabetic conditions and the sequence of the integrin-ligand peptides derived from BIGH3, as well as the integrins involved in BMA, is expected to offer novel therapeutic targets for interventions to block development and progression of diabetic complications.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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