Oxidative stress increases MMP2 expression and activity in INS-1 cells.

<p>INS-1 cells were treated with GA (200 mg/L) or BSA (200 mg/L) for 24 h. Antioxidant NAC was dissolved in sterile double-distilled water and used in different final concentrations with GA (200 mg/L). The antioxidant effect of NAC was measured by DCFH-DA through flow cytometer. The concentration with better antioxidant effect was used for following experiments. (a, b) The level of cytosolic ROS in different groups. The mean fluorescence intensity (MFI) of DCF is used to measure cellular ROS levels. BSA–treated cells without DCFH-DA incubation serve as negative control. (c) Apoptotic cells in different groups. Annexin V positive cells (including Annexin V single positive population and the Annexin V and PI double positive population) representing apoptotic cells were mainly located in the right upper quadrant and right lower quadrant. (d) Average apoptotic rate in the three groups. (e) Insulin secretion of INS-1 cells stimulated with 3.3 mmol/L glucose or 16.7 mmol/L glucose in the different groups. *<i>P<0.05 vs</i> the corresponding control group; ^#<i>P<0.05 vs</i> the corresponding GA group. (f) Insulin release index (IRI) of each group. The IRI was adopted as an index to estimate the insulin secretion function of INS-1 cells. IRI = insulin concentration of high glucose stimulation (16.7 mmol/L)/insulin concentration of low glucose stimulation (3.3 mmol/L). (g, h) Protein expression of MMP2 in different groups, including both pro-MMP2(∼72 kD) and active MMP2(∼62 kD). (i, j) The gelatinolytic activity of MMP2 in different groups. The white band in gray background was the location of MMP2, band intensity was used to represent the activity of MMP2 in each group. Data are shown as means ± SD, <i>P<0.05</i> indicates a statistically significant difference. *<i>P<0.05 vs</i> the control group; ^#<i>P<0.05 vs</i> the GA group.