ABSTRACT
We were intrigued to analyze donor eyes of two individuals without retinopathy even after 40 years of type 2 diabetes mellitus. Targeted molecular factors associated with angiogenesis and the key antioxidant enzymes in retinal tissue were analyzed. Accordingly PEDF, Adiponectin and Paraoxonase 2 showed augmented mRNA expression in both the retina with no significant change in VEGF expression. Vitreous showed increased PEDF protein in donor 1 and Adiponectin in donor 2 with no change in VEGF protein. This study highlights the profile of specific molecular factors that contribute to the non-development of diabetic retinopathy changes in these individuals.
Subject(s)
Adiponectin/biosynthesis , Aryldialkylphosphatase/biosynthesis , Diabetes Mellitus, Type 2/diagnosis , Eye Proteins/biosynthesis , Gene Expression Regulation , Nerve Growth Factors/biosynthesis , Retina/pathology , Serpins/biosynthesis , Tissue Donors , Adiponectin/genetics , Aged, 80 and over , Aryldialkylphosphatase/genetics , Biomarkers/metabolism , Diabetes Mellitus, Type 2/genetics , Diabetes Mellitus, Type 2/metabolism , Eye Proteins/genetics , Female , Humans , Male , Middle Aged , Nerve Growth Factors/genetics , Oxidative Stress , RNA/genetics , Retina/metabolism , Retinal Diseases , Serpins/geneticsABSTRACT
OBJECTIVE: The study was conducted to observe the serum and vitreous levels of LXA4, BDNF and Th1/Th2 cytokines in type 2 diabetes mellitus (DM) and changes associated with diabetic retinopathy (DR). Further, the in vitro study was performed to analyze the exposure of BDNF and LXA4 on LPS-induced pro-inflammatory state in ARPE 19 cells. MATERIALS AND METHODS: Totally 114 individuals were recruited in a prospective case control study. Of these, 27 were type 2 DM cases with no complications, 30 cases were type 2 DM with non proliferative diabetic retinopathy (NPDR), 30 were type 2 DM with proliferative diabetic retinopathy (PDR), and 27 were healthy control. ELISA was done to estimate the serum and vitreous levels of BDNF, VEGF and PEDF. FACS cytometric Bead Array system was used to analyze the serum cytokines. RESULTS: The serum BDNF and LXA4 levels were significantly reduced in both NPDR and PDR cases compared to control (p=0.005, 0.01; p=0.033, 0.015). Serum IL-6 was significantly increased in the PDR group (p=0.04). BDNF showed a significant negative correlation with VEGF levels (r=-0.522, p<0.01) and positive correlation with IL-10 (r=0.67, p<0.05) in serum. A significant odds ratio for the serum BDNF (OR: 3.20, p=0.025) as well as serum IL-6 (OR: 1.244, p=0.042) indicated them as potential risk factors for progression of type 2 DM to DR. A significant decrease in both the LXA4 (p=0.013) and BDNF (p=0.0008) with increase in cytokines IL-6 and IL-10 levels were observed in the vitreous of PDR cases ((p=0.04, 0.01). In vitro studies showed that both LXA4 (10 nmol/L) and BDNF (500 pg) decreased the IL-6 levels (p=0.036, 0.0002), in LPS induced pro-inflammatory condition in ARPE 19 cells, thereby their anti-inflammatory effect. CONCLUSIONS: This study reports that low serum BDNF and higher IL-6 levels are potential risk factors for DR in type 2 DM. This study supports the role of BDNF in modulating the pro- and anti-inflammatory cytokines, and low level of BDNF is associated with development of diabetic retinopathy.
Subject(s)
Brain-Derived Neurotrophic Factor/blood , Diabetic Retinopathy/blood , Lipoxins/blood , Th1-Th2 Balance , Adult , Aged , Case-Control Studies , Cell Line , Cytokines/blood , Diabetes Mellitus, Type 2/complications , Diabetic Retinopathy/epidemiology , Female , Humans , In Vitro Techniques , Lipopolysaccharides/pharmacology , Male , Middle Aged , Prospective Studies , Retina/cytology , Risk Factors , Vascular Endothelial Growth Factor A/bloodABSTRACT
Epithelial to Mesenchymal Transition (EMT) of the retinal pigment epithelium is involved in the pathogenesis of proliferative vitreoretinopathy (PVR) that often leads to retinal detachment. In this study, Triphala, an ayurvedic formulation and two of its active ingredients, namely chebulagic acid and chebulinic acid were evaluated for anti-EMT properties based on in vitro experiments in human retinal pigment epithelial cell line (ARPE-19) under TGFß1 induced conditions. ARPE-19 cells were treated with TGFß1 alone or co-treated with various concentrations of aqueous extract (AqE) (30-300 µg/ml); alcoholic extract (AlE) (50-500 µg/ml) of triphala and the active principles chebulagic acid (CA) and chebulinic acid (CI) (CA,CI: 50-200 µM). The expression of EMT markers namely MMP-2, αSMA, vimentin and the tight junction protein ZO-1 were evaluated by qPCR, western blot and immunofluorescence. The functional implications of EMT, namely migration and proliferation of cells were assessed by proliferation assay, scratch assay and transwell migration assay. AqE, AlE, CA and CI reduced the expression and activity of MMP-2 at an ED50 value of 100 µg/ml, 50 µg/ml, 100 µM and 100 µM, respectively. At these concentrations, a significant down-regulation of the expression of αSMA, vimentin and up-regulation of the expression of ZO-1 altered by TGFß1 were observed. These concentrations also inhibited proliferation and migration of ARPE-19 cells induced by TGFß1. EMT was found to be induced in ARPE-19 cells, through SMAD-3 phosphorylation and it was inhibited by AqE, AlE, CA and CI. Further studies in experimental animals are required to attribute therapeutic potential of these extracts and their active compounds, as an adjuvant therapy in the disease management of PVR.
