Enhancement of porcine in vitro embryonic development through luteolin‑mediated activation of the Nrf2/Keap1 signaling pathway

Background Oxidative stress,caused by an imbalance in the production and elimination of intracellular reactive oxygen species(ROS),has been recognized for its detrimental effects on mammalian embryonic development.Luteolin(Lut)has been documented for its protective effects against oxidative stress in various studies.However,its specific role in embryonic development remains unexplored.This study aims to investigate the influence of Lut on porcine embryonic development and to elucidate the underlying mechanism.Results After undergoing parthenogenetic activation(PA)or in vitro fertilization,embryos supplemented with 0.5μmol/L Lut displayed a significant enhancement in cleavage and blastocyst formation rates,with an increase in total cell numbers and a decrease in the apoptosis rate compared to the control.Measurements on D2 and D6 revealed that embryos with Lut supplementation had lower ROS levels and higher glutathione levels compared to the control.Moreover,Lut supplementation significantly augmented mitochondrial content and membrane potential.Intriguingly,activation of the Nrf2/Keap1 signaling pathway was observed in embryos supplemented with Lut,leading to the upregulation of antioxidant-related gene transcription levels.To further validate the relationship between the Nrf2/Keap1 signaling pathway and effects of Lut in porcine embryonic development,we cultured PA embryos in a medium supplemented with brusatol,with or without the inclusion of Lut.The positive effects of Lut on developmental competence were negated by brusatol treatment.Conclusions Our findings indicate that Lut-mediated activation of the Nrf2/Keap1 signaling pathway contributes to the enhanced production of porcine embryos with high developmental competence,and offers insight into the mechanisms regulating early embryonic development.

Effect of pestle intervention in type 2 diabetic peripheral neuropathy on Keap1/Nrf2/ARE pathway and the relationship with oxidative stress

Objective:To investigate the effect of pestle needle treatment on Nrf2 pathway and the relationship with oxidative stress in diabetic peripheral neuropathy.Methods:Patients with DPN who met the inclusion criteria were randomly divided into control and test groups with 30 patients in each group in a 1:1 allocation ratio.Both groups were given basic treatment,and the pestle group was treated with needle pestle therapy 5 times a week for a total of 4 weeks of intervention.Serum SOD and GSH PX levels were examined by colorimetry before and after intervention;Serum Keap1/Nrf2/ARE signaling pathway related factors expression levels were measured by ELISA;Keap1 and Nrf2 mRNA expression was determined by RT-PCR.Results:Compared with the control group,SOD and GSH-Px in the test group were significantly increased,Keap1 expression was decreased,Nrf2 expression was increased,Keap1 mRNA expression was significantly decreased,and Nrf2 mRNA expression was significantly increased.Conclusions:the pestle needle may enhance the body's antioxidant capacity by modulating the Keap1/Nrf2/ARE signaling pathway to enhance the production of its downstream antioxidant enzymes SOD and GSH Px,thereby protecting and repairing the damaged peripheral nerves in DPN patients.

枸杞多糖对四氯化碳致急性肝损伤小鼠的保护作用

目的探讨枸杞多糖(lycium barbarum polysaccharides,LBP)对四氯化碳(CCl4)致急性肝损伤小鼠的保护作用。方法40只ICR小鼠适应性饲养一周后随机分为正常对照组,模型组,LBP低剂量组(100 mg·kg^(-1))、中剂量组(200 mg·kg^(-1))和高剂量组(400 mg·kg^(-1)),每组8只。LBP低、中、高剂量组每天灌胃0.5 mL各对应浓度LBP溶液,正常对照组和模型组给予与各给药组等剂量的生理盐水,每天1次,持续7 d。在末次给药后1 h,除正常对照组外,其余各组小鼠一次性腹腔注射CCl_(4)花生油溶液(浓度为0.1%,10 mg·kg^(-1))诱导小鼠急性肝损伤。(1)造模后24 h眼球取血,取血前称取小鼠体质量及取血后称取肝脏重量计算肝指数变化情况;(2)HE染色观察各组小鼠肝脏的病理学改变;(3)ELISA检测血清中肝功能指标谷丙转氨酶(ALT),谷草转氨酶(AST)及炎性指标肿瘤坏死因子-α(TNF-α)、白细胞介素1β(IL-1β)和白细胞介素6(IL-6)水平;(4)qRT-PCR检测Keich样环氧氯丙烷相关蛋白(Keich-like ECH-associated protein 1,Keap1)、核因子-kβ抑制蛋白激酶(inhibitor of nuclear fac-tors-kB kinase,IKKβ)、核因子-kB(nuclear factor-kB,NF-kB)mRNA表达水平;(5)Western blot检测Keap1、IKKβ、NF-kB蛋白表达水平。结果(1)与正常对照组相比,模型组肝指数升高(P<0.01);与模型组相比,高剂量组肝指数降低(P<0.01);(2)正常对照组肝细胞形态结构正常,肝细胞索边界清楚,肝细胞以中央静脉为中心呈放射状排列;模型组有不同程度的细胞水肿;各LBP组病理学形态均有不同程度改善;(3)与正常对照组相比,模型组AST、ALT、TNF-α、IL-1β、IL-6水平均升高(P均<0.01);与模型组相比,低剂量组IL-1β降低(P<0.01),中剂量组AST、TNF-α、IL-1β、IL-6均降低(P均<0.01),高剂量组AST、ALT、TNF-α、IL-1β、IL-6均降低(P均<0.01);(4)与正常对照组相比,模型组Keap1 mRNA表达降低(P<0.01),IKKβ、NF-kB mRNA表达均升高(P均<0.01);与模型组相比,中剂量和高剂量组Keap1 mRNA表达均升高(P均<0.01),IKKβ和NF-kB mRNA表达均降低(P均<0.01);(5)与正常对照组相比,模型组Keap1蛋白表达降低(P<0.01),IKKβ、NF-kB蛋白表达升高(P<0.01或<0.05),与模型组相比,高剂量组Keap1蛋白表达升高(P<0.05),中剂量和高剂量组IKKβ、NF-kB蛋白表达均降低(P均<0.01)。结论LBP对CCl4致急性肝损伤小鼠具有保护作用,其机制可能与增强Keap1、抑制IKKβ/NF-kB抗炎信号通路有关。

Protective effects of peptide KSPLY derived from Hericium erinaceus on H_(2)O_(2)-induced oxidative damage in HepG2 cells

Reactive oxygen species(ROS)-induced oxidative damage is strongly associated with the pathogenesis of chronic diseases,and natural antioxidant peptides have good abilities of scavenging ROS.The antioxidant activity of peptide Lys-Ser-Pro-Leu-Tyr(KSPLY)derived from Hericium erinaceus remains unclear.In the present study,the antioxidant effect and mechanism of KSPLY on H_(2)O_(2)-induced oxidative damage in HepG2 cells were investigated.The results indicated that KSPLY exhibited the antioxidant capacity in H_(2)O_(2)-induced HepG2 cells by enhancing superoxide dismutase(SOD),glutathione peroxidase(GSH-Px),and catalase(CAT)activities.In comparison with the H_(2)O_(2)-treated damage group,the apoptosis rate,ROS level,and malondialdehyde(MDA)content of HepG2 cells treated with KSPLY were significantly decreased.The H.erinaceus-derived peptide KSPLY pretreatment promoted the expression of detoxification and antioxidant enzymes via the Keap1/Nrf2 signal pathway,thereby inhibiting the generation of ROS and MDA.In conclusion,the H.erinaceus-derived peptide KSPLY effectively protected HepG2 cells against H_(2)O_(2)-induced oxidative damage,and it provided a theoretical basis for the further development of new natural antioxidants.

Advance in the Study of the Mechanisms Regulated by Sphingosine-1-Phosphate

Sphingosine-1-phosphate（S1P） is a bioactive lipid messenger in the cells that regulate gene expression and NF-κB signal pathway through unknown mechanisms.Recently,Cheng Luo,associate professor of DDDC in Shanghai Institute of Materia Medica,whose project was funded by the National Natural Science Foundation of China,joined in a research team led by Professor Sarah Spiegel of Virginia Commonwealth University.The team continuously made significant breakthroughs in understanding the regulation mechanism of Sphingosine-1- Phosphate.In September 2009,in a paper published on SCIENCE magazine（Science 2009, 325：1254-7）,they firstly demonstrated that S1P is a physiologically important regulator of histone deacetylases（HDACs）,HDACs are direct intracellular targets of S1P.Furthermore,they identified the mechanism that S1P regulates gene expression through regulating the activity of HDACs.In June 24th,2010,in another paper to be published on NATURE magazine（Nature 2010,June 24th,advance online publication,（doi：10.1038/ nature09128）） which reports the regulation of NF-κB signaling pathway by S1P.They demonstrate that S1P is the missing cofactor for TRAF2（tumour-necrosis factor receptor-associated factor 2） and indicate a new paradigm for the regulation of lysine-63- linked poly-ubiquitination.The study also highlight the key role of SphK1 and its product S1P in TNF-αsignalling and the canonical NF-κB activation pathway, and then play crucial role in inflammatory,antiapoptotic and immune processes.The identification of new mechanisms fay which S1P regulates gene expression and TNF and NF-κB signaling pathway will light up the road to develop novel inhibitors that might be useful for treatment of cancer and in- flammatory diseases.