Effect of recombinant human erythropoietin expressions of apoptosis genes in rats following traumatic brain injury

Purpose: To explore the effect of recombinant human erythropoietin (r-HuEPO) on apoptosis in rats after traumatic brain injury. Methods: A total of 48 traumatic brain-injured Sprague Dawley (SD) rats were obtained by improved Feeney’s traumatic brain injury model, and were randomly divided into four groups: normal salinetreated rats (control) and rats treated with r-HuEPO at doses of 1000 U/kg, 3000 U/kg and 5000 U/kg. Brain tissues were collected on the 7th day after trauma surgery. Apoptotic cells, and NF-kappa B (NFĸB)-, c-myc-, and Fas/Fasl-positive cells were identified in brain tissues by immunohistochemical assay. Results: After treatment with r-HuEPO (3000 and 5000 U/kg), expression of NF-κB and Fas/Fasl were significantly decreased (p < 0.05) compared to control rats, especially at the 5000 U/kg dose (p < 0.01). However, for c-myc, no significant difference was observed between r-HuEPO treatment and control groups (p > 0.05). Compared to the 1000 U/kg r-HuEPO group, Fas/Fasl expression levels were significantly lower in the 3000 and 5000 U/kg r-HuEPO groups (p < 0.05). Additionally, expression of NF-κB and Fasl in the 5000 U/kg r-HuEPO group was significantly lower than that in the 3000 U/kg r- HuEPO group (p < 0.05). Moreover, the number of apoptotic cells in the r-HuEPO group (5000 U/kg) was significantly lower than in the control group (p < 0.05). Conclusion: Thus, r-HuEPO may be beneficial for treating traumatic brain injury via inhibition of NFkappa B and Fas/Fasl expressions.

Effect of Lycii fructus polysaccharides on ovulation failure in rats

Purpose: To investigate the effect of Lycii fructus polysaccharides (LFPS) on ovulation failure. Methods: A rat model of ovulation failure was established by intragastric administration of hydroxyurea (300 mg/kg). Rats with ovulation failure then received LFPS via oral administration at doses of 100, 200, or 400 mg/kg. The body, uterus and ovary of each rat were weighed using electronic scales. The hypothalamic-pituitary-ovarian (HPO) axis hormones, including estradiol (E2) level, follicle-stimulating hormone (FSH) activity, and luteinizing hormone (LH) activity in the serum of each rat were determined by enzyme-linked immunosorbent assay (ELISA). The levels of pro-apoptotic proteins (Fas, FasL, FADD, c-caspase-8, c-caspase-10, c-caspase-3, c-caspase-6, and c-caspase-7) in the ovarian tissue of each rat were detected by western blot. Results: Hydroxyurea reduced significantly (p < 0.01) uterus and ovary indices (uterus or ovary weight/body weight) (0.119 and 0.026 %), E2 level (3.42 pmol/L), and FSH and LH activities (2.28 and 2.76 U/L), compared with those in the normal group (0.169 and 0.039 %; 6.72 pmol/L; 2.76 and 3.75 U/L). Hydroxyurea increased significantly (p < 0.01) the levels of the above-mentioned pro-apoptotic proteins relative to those in the normal group. LFPS (100, 200, or 400 mg/kg) reversed significantly (p < 0.05 or 0.01) the effect of hydroxyurea on all of the above indices. Conclusion: LFPS exhibits a protective effect on hydroxyurea-induced ovulation failure by regulating the HPO axis hormones and death receptor-mediated apoptotic pathway.

Isolation and identification of two galangin metabolites from rat urine and determination of their in vitro hypolipidemic activity

Purpose: To investigate the lipid-lowering activity of two metabolites of galangin, namely, galangin-3-Oβ-D-glucuronic acid (GG-1) and galangin-7-O-β-D-glucuronic acid (GG-2). Methods: Female Sprague-Dawley rats were orally administered with galangin. The two metabolites of galangin were isolated from urine sample and purified using Sephadex LH-20 and semi-preparative high performance liquid chromatography (HPLC). The structures of the metabolites were identified by analyzing spectroscopic data. Hypolipidemic activity was evaluated in HepG2 cells. The down- or upregulation of lipogenic genes was detected using real-time quantitative polymerase chain reaction (qPCR). Results: Both metabolites of galangin showed hypolipidemic activity. These activities are closely associated with the down-regulation of lipogenic genes such as SREBP-1a, SREBP-1c, and SREBP-2 transcription factors, and the downstream genes such as FAS, ACC, and HMGR were revealed by realtime qPCR data. Conclusion: The results show that both metabolites possess better lipid-lowering activities than galangin. These hypolipidemic activities are closely associated with inhibiting key genes or proteins that regulated the biosynthesis of both cholesterol and triglycerides.

Withaferin A promotes proliferation and migration of brain endothelial cells

Purpose: To investigate the effect of withaferin A (WFA) on the proliferation and migration of brain endothelial cells. Methods: BALB-5023 mouse microvascular cells were treated with a range of withaferin A (WFA) concentrations from 10 to 100 ng/mL. Dojindo’s CCK-8 cell proliferation kit was used for the analysis of cell proliferation. Transwell cell culture inserts were used to determine the migration potential of WFAtreated endothelial cells. Absorbance was measured at 450 nm on an enzyme-linked immunosorbent (ELISA) reader. Results: The results revealed a significant increase in the proliferation and migration of endothelial cells following treatment with a low concentration (30 ng/mL) of WFA compared with the higher concentration (> 10 ng/mL). The effect was further enhanced when WFA was used in combination with soluble Fas ligand (sFasL). Autocrine signaling of vascular endothelial growth factor (VEGF) by endothelial cells was significantly increased following treatment with WFA or in combination with sFasL. WFA increased the expression of Fas on endothelial cells, suggesting the involvement of sFasL in the proliferation and migration of brain endothelial cells. Conclusion: Thus, WFA promotes the proliferation and migration of endothelial cells through increase in the expression of Fas and secretion of VEGF.