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. 2019 Jun 2;15(7):1472–1487. doi: 10.7150/ijbs.33817

Figure 3.

Figure 3

Hypericin protects INS-1 cells from glucotoxicity and lipotoxicity via Erk signaling and maintianing PDX1 expression. (A) Effects of hypericin on the PDX1 mRNA level in INS-1 cells under glucotoxicity. INS-1 cells were treated with high (33 mM) glucose, 200 nM hypericin or a combination of the two for 72 h. Total RNA was extracted, and PDX1 mRNA was amplified by conventional SYBR Green real-time PCR analysis. Data are presented as the mean ± S.D. (n = 3). (B) Effects of hypericin on the protein levels of PDX1, Erk1/2 and p-Erk in INS-1 cells under glucotoxicity. INS-1 cells were treated as in (A). Cell lysates were prepared and subjected to Western blots using the indicated antibodies. GAPDH was used as a loading control. The density ratios of PDX1 to GAPDH or p-Erk to Erk were measured by ImageJ as shown in the right panel. The experiment was repeated three times. *** p<0.001 versus the 33 mM glucose-treated group. (C) Blockade of hypericin-mediated effects by U0126 in INS-1 cells under glucotoxicity. INS-1 cells were treated with different combinations of high (33 mM) glucose, 200 nM hypericin and U0126 as indicated for 72 h. Then, target proteins were detected by Western blot using the indicated antibodies. GAPDH was used as a loading control. The density ratios of PDX1 to GAPDH, p-Erk to Erk or CC3 to GAPDH were measured by ImageJ as shown in the right-hand panel. The experiment was repeated three times. **p<0.01, ***p<0.001 versus the 33 mM glucose-treated group; ##p<0.01, ###p<0.001 versus the 33 mM glucose+200 nM hypericin-treated group. (D) Effects of hypericin on the Erk pathway in INS-1 cells under lipotoxicity. INS-1 cells were treated with 200 μM PA, 200 nM hypericin or a combination of the two for 24 h. Cell lysates were prepared and subjected to Western blots using anti-Erk and anti-p-Erk antibodies. GAPDH was used as a loading control. The density ratios of p-Erk to Erk were measured by ImageJ as shown in the right panel. The experiment was repeated three times. **p<0.01, *** p<0.001 versus the 200 μM PA-treated group. (E) Blockade of hypericin-mediated effects by U0126 in INS-1 cells under lipotoxicity. INS-1 cells were treated with different combinations of 200 μM PA, 200 nM hypericin and U0126 as indicated for 24 h. Then, target proteins were detected and analysed as in (c). *** p<0.001 versus the 200 μM PA-treated group; ## p<0.01 versus the 200 μM PA+200 nM hypericin-treated group.