ABSTRACT
Biliverdin (BV) has long been thought to be a cytotoxic metabolic waste product. It has also been demonstrated to have important cytoprotective functions during oxidative stress. The present study aimed to examine the cytoprotective effect of BV on NRK-52E cells, a proximal tubular cell line derived from rat kidney. Cells were treated with 50 µmol/L cisplatin for 24 h (cisplatin group) or pre-treated with BV for 30 min, then with 50 µmol/L cisplatin for 24 h (cisplatin+BV group). Those given no treatment served as a control. Cell apoptosis was evaluated by flow cytometry and cell viability by Cell Counting Kit-8 (CCK-8). The protein expressions of cleaved caspase3, Bax and Bcl-2 were assessed by Western blotting. Reactive oxygen species (ROS) levels were measured using carboxydichlorodihydrofluorescein diacetate (H2DCF). The results showed that cisplatin induced the apoptosis of NRK-52E cells, decreased cell viability, and increased the formation of ROS by upregulating the expression of cleaved caspase3 and Bax and decreasing Bcl-2 protein expression. These effects could be significantly reversed by pretreatment with BV. It was concluded that BV can protect against cisplatin-induced cell apoptosis through the anti-oxidative effects.
Subject(s)
Animals , Rats , Antioxidants , Pharmacology , Apoptosis , Biliverdine , Pharmacology , Cell Line , Cisplatin , Toxicity , Epithelial Cells , Metabolism , Kidney Tubules , Cell Biology , Reactive Oxygen Species , MetabolismABSTRACT
Biliverdin (BV) has long been thought to be a cytotoxic metabolic waste product. It has also been demonstrated to have important cytoprotective functions during oxidative stress. The present study aimed to examine the cytoprotective effect of BV on NRK-52E cells, a proximal tubular cell line derived from rat kidney. Cells were treated with 50 µmol/L cisplatin for 24 h (cisplatin group) or pre-treated with BV for 30 min, then with 50 µmol/L cisplatin for 24 h (cisplatin+BV group). Those given no treatment served as a control. Cell apoptosis was evaluated by flow cytometry and cell viability by Cell Counting Kit-8 (CCK-8). The protein expressions of cleaved caspase3, Bax and Bcl-2 were assessed by Western blotting. Reactive oxygen species (ROS) levels were measured using carboxydichlorodihydrofluorescein diacetate (H2DCF). The results showed that cisplatin induced the apoptosis of NRK-52E cells, decreased cell viability, and increased the formation of ROS by upregulating the expression of cleaved caspase3 and Bax and decreasing Bcl-2 protein expression. These effects could be significantly reversed by pretreatment with BV. It was concluded that BV can protect against cisplatin-induced cell apoptosis through the anti-oxidative effects.
ABSTRACT
This study is to explore the effect of ginkgolide B (BN52021) on the production of nitric oxide (NO), interleukin (IL)-6 and regulated upon activation normal T cell expressed and secreted (RANTES) from astrocytes induced by stimulators. Primary cultured rat astrocytes were stimulated with lipopolysaccharides (LPS), the production of NO was assayed using Griess reaction; U251 cells were stimulated with IL-1 beta, the contents of IL-6 and RANTES in the supernatant were measured using ELISA. The mRNA expressions of IL-6 and RANTES were detected using RT-PCR. LPS (10 ng mL(-1) to 10 microg mL(-1)) could stimulate rat astrocytes to produce NO in a dose-dependent manner. Ginkgolide B at the concentrations of 0.1-10 micromol L(-1) were shown to decrease NO production significantly. IL-1 beta could induce the mRNA expression and protein secretion of IL-6 from U251 cells, as well as RANTES. Ginkgolide B at concentrations of 0.1-10 micromol L(-1) were shown to inhibit RANTES secretion, and to inhibit mRNA expression of IL-6 and RANTES at concentration of 10 micromol L(-1). Ginkgolide B has inhibitory effect on the production of NO, IL-6 and RANTES from astrocytes treated with inflammatory stimulators.
Subject(s)
Animals , Humans , Male , Mice , Rats , Astrocytes , Cell Biology , Metabolism , Cell Line, Tumor , Cells, Cultured , Chemokine CCL5 , Genetics , Metabolism , Dose-Response Relationship, Drug , Ginkgolides , Pharmacology , Glioblastoma , Metabolism , Pathology , Interleukin-1beta , Interleukin-6 , Genetics , Bodily Secretions , Lactones , Pharmacology , Lipopolysaccharides , Mice, Inbred C57BL , Nitric Oxide , Metabolism , Platelet Activating Factor , RNA, Messenger , Metabolism , Rats, WistarABSTRACT
<p><b>AIM</b>To investigate the inhibitory effect of ginkgolide B on angiogenesis in chronic inflammation and the possible mechanisms.</p><p><b>METHODS</b>The murine chronic granulomatous air pouch model was used to observe the anti-angiogenesis effect of ginkgolide B. The vascular index was determined by colorimetry of carminic acid, and angiogenesis was observed by histology method. The interleukin-1beta (IL-1beta) levels in mice serum and in supernatants of U937 cell culture stimulated by phorbol 12-myristate 13-acetate (PMA) were detected by radioimmunoassay (RIA). The tumor necrosis factor-alpha (TNF-alpha) levels in mice serum and in supernatant of U937 cell culture were measured by cytotoxicity bioassay. The mRNA expression of IL-1beta and TNF-alpha of U937 cell culture was investigated by RT-PCR.</p><p><b>RESULTS</b>Oral administration of ginkgolide B 25 and 100 mg x kg(-1) was shown to significantly inhibit the vascular index of murine chronic granulomatous air pouch model with the inhibitory rate of 22.52% and 25.29%, respectively. This result was supported by histological observation. Concomitantly, the IL-1beta levels in mice serums were also significantly decreased with the inhibitory rate of 50.61% and 58.66%; so were the TNF-alpha levels with the inhibitory rate of 28.91% and 52.41%. Ginkgolide B at concentration of 1 x 10(-5) to 1 x 10(-8) mol x L(-1) could also reduce both the IL-1beta and TNF-alpha contents in the supernatants of U937 cell culture stimulated by PMA, but the scopes of changes were much different. For IL-1beta the IC50 was 1.93 x 10(-8) mol x L(-1), while ginkgolide B at concentration of 1 x 10(-5) mol x L(-1) only decreased the release of TNF-alpha by 25.99%. Furthermore, ginkgolide B at concentrations of 1 x 10(-5) to 1 x 10(-7) mol x L(-1) was shown to significantly inhibit TNF-alpha mRNA expression of U937 cells; and at concentrations of 1 x 10(-5) and 1 x 10(-6) mol x L(-1) could inhibit IL-1beta mRNA expression.</p><p><b>CONCLUSION</b>Ginkgolide B was shown to significantly inhibit angiogenesis of the murine chronic granulomatous air pouch model, reduce the IL-1beta and TNF-alpha levels in mice serums, and significantly inhibit IL-1beta and TNF-alpha mRNA expression and protein secretion in supernatants of U937 cell culture. It was suggested that reduction of proangiogenic cytokines IL-1beta and TNF-alpha secretion may contribute to the anti-angiogenesis effect of ginkgolide B in the murine chronic granulomatous air pouch model.</p>