Berberine reverses LPS-induced repression of CYP7A1 through an anti-inflammatory effect

Objective: To evaluate the anti-inflammatory effect of berberine(BBR) on the lipopolysaccharide(LPS)-induced acute phase response and its modulation of the altered bile acid metabolism induced by LPS treatment.Methods: An acute phase response was induced by intraperitoneal injection of LPS(5 mg/kg, ip) in C57 BL/6 J mice, and the BBR treatment group was orally administered with BBR(200 mg/ky, ig). The levels of TNFα, IL-1β and IL-6 in the serum were measured using an ELISA kit, and their expression levels in the liver were measured using q RT-PCR. The bile acid pool was measured using a commercial bile acid kit, and the expression levels of enzymes involved in bile acid metabolism were measured by q RTPCR. The expression levels of CYP7 A1, p65 NF-κ B and the MAPK signaling pathway was measured using Western blotting.Results: LPS treatment suppressed the expression of CYP7 A1 and CYP8 B1, and the total bile acid pool was also reduced. Pretreatment with BBR inhibited the pro-inflammatory biomarkers TNFα and IL-1β in the serum, as well as the expression of TNFα, IL-1β and i NOS mRNA in the liver. BBR treatment did not affect the reduction in the bile acid pool size induced by LPS, but significantly increased the concentration of bile acids in the liver, which was consistent with the upregulated expression of CYP7 A1 and CYP8 B1. The MAPK signaling pathway was activated by BBR treatment, while the p65 NF-κ B signaling pathway was inhibited.Conclusion: BBR can offer an anti-inflammatory effect and reverse the inhibition of CYP7 A1 and CYP8 B1 expression caused by LPS treatment, as well as induce the production of bile acids in liver, probably via MAPK signaling; However, treatment with BBR had no effect on the size of total bile acid pool.     

艾灸对克罗恩病大鼠结肠NF-κB p65及TNF-α、IL-1β调节作用的研究

目的：通过观察隔药灸对克罗恩病大鼠结肠NF-κB p65及炎症因子TNF-α、IL-1β表达的影响,探讨艾灸治疗克罗恩病的效应机制。方法：清洁级雄性SD大鼠随机分为4组,分别为正常组、模型组、隔药灸组和西药组,采用三硝基苯磺酸（TN-BS）合乙醇溶液灌肠制备克罗恩病大鼠模型,模型制备成功后,隔药灸组取天枢穴（双）、气海穴进行隔药灸治疗,西药组采用美沙拉秦灌胃治疗。治疗结束后,采用HE染色,光镜下观察各组大鼠结肠组织病理学变化,应用免疫组织化学技术观察各组大鼠结肠NF-κB p65与TNF-α、IL-1β的表达,并进行图像分析。结果：与正常组比较,模型组大鼠结肠组织呈现裂隙状溃疡及炎症表现,部分大鼠结肠在黏膜下层可见肉芽肿形成,结肠NF-κB p65、TNF-α、IL-1β的表达均显著增高（均P〈0.01）;与模型组比较,隔药灸组、西药组大鼠结肠形态结构改善,肠道炎症减轻,结肠NF-κB p65与TNF-α、IL-1β的表达均显著降低（均P〈0.01）;相关性分析结果显示,TNF-α、IL-1β的表达与NF-κB p65的表达均呈正相关性。结论：隔药灸能下调克罗恩病大鼠结肠NF-κB p65、TNF-α、IL-1β的表达;艾灸（隔药灸）可能通过抑制克罗恩病大鼠结肠NF-κB p65的表达,进而减少其下游炎症因子TNF-α、IL-1β的表达,从而减轻肠道炎症,改善结肠组织形态结构,发挥治疗作用。     

电针耳甲区对内毒素血症模型大鼠的抗炎保护作用

目的:观察耳甲区电针对脂多糖致内毒素血症模型大鼠血清炎性反应因子水平与肺组织核因子κB(NF-κB)表达的影响,探讨耳甲刺激对炎性反应的保护作用机制。方法:SD大鼠随机分为正常对照组、模型组、单纯耳甲电针组、耳甲电针组、迷走神经刺激组、后三里组,每组12只。尾静脉注射脂多糖(5 mg/kg)复制内毒素血症模型。耳甲电针取双侧耳甲区,迷走神经刺激给予左侧颈部迷走神经电刺激,后三里组行双侧"后三里"电针,均为20 min。采用酶联免疫吸附法测定各组大鼠血清肿瘤坏死因子-α(TNF-α)和白介素-6(IL-6)水平,采用免疫印迹法测定各组肺组织NF-κB p 65蛋白表达。结果:与正常对照组相比,模型组TNF-α、IL-6水平明显升高(P<0.01),NF-κB p 65表达明显上调(P<0.01);单纯耳甲电针组NF-κB p 65表达明显上调(P<0.01)。与模型组相比,耳甲电针组和迷走神经刺激组TNF-α、IL-6水平明显下降(P<0.01),NF-κB p65表达明显下调(P<0.01);后三里组TNF-α水平明显下降(P<0.05)。与迷走神经刺激组相比,耳甲电针组IL-6水平显著升高(P<0.01),后三里组TNF-α、IL-6水平、NF-κB p 65表达显著升高(P<0.05,P<0.01)。与耳甲电针组相比,后三里组NF-κB p65表达显著升高(P<0.05)。结论:耳甲刺激能降低内毒素血症模型大鼠致炎因子水平,下调NF-κB蛋白表达,其效应与直接刺激迷走神经相似,说明耳甲刺激可能激活了胆碱能抗炎通路,从而启动抗炎效应。     

电针“肠病方”腧穴对溃疡性结肠炎大鼠结肠组织核因子-κB p65及血清白介素-4的影响

目的:探讨电针"肠病方"腧穴治疗溃疡性结肠炎的作用机制。方法:将SD大鼠随机分为空白组、模型组、肠病方组,每组10只。采用冰乙酸灌肠法复制溃疡性结肠炎模型。肠病方组电针"中脘"天枢"和"上巨虚"15min,分别在造模后即刻、24h和48h进行治疗。采用酶联免疫吸附法检测血清白介素-4(IL-4)含量,免疫组化、蛋白质印迹法检测肠组织核因子-κB p65(NF-κB p65)蛋白表达。结果:模型组NF-κBp65蛋白表达明显高于空白组(P<0.01,P<0.05),肠病方组低于模型组(P<0.01,P<0.05);模型组血清IL-4含量较空白组升高(P<0.05),经电针"肠病方"治疗后,IL-4含量进一步升高(P<0.01)。结论:腧穴"肠病方"对乙酸所致肠黏膜损伤可通过调节炎性反应因子起到修复作用。     

清热排毒胶囊对急性痛风性关节炎模型大鼠核因子κB和白细胞介素-1β的影响

目的：观察清热排毒胶囊对尿酸钠晶体（MSU）诱导的急性痛风性关节炎模型大鼠核因子κB（NF-κB）和白细胞介素-1β（IL-1β）的影响,探讨其防治急性痛风性关节炎的作用机理。方法60只SD雄性大鼠随机分为空白组、模型组、双氯芬酸钠缓释片组（戴芬组）及清热排毒胶囊高、中、低剂量组,各治疗组予相应药物灌胃7 d,于第5日灌胃给药1 h后采用MSU溶液踝关节腔注射制备大鼠急性痛风性关节炎模型。各组大鼠于造模后48 h处死并取材。光镜下观察各组大鼠踝关节滑膜组织的病理形态学改变,ELISA测定各组大鼠踝关节浸出液中NF-κB和IL-1β的表达水平。结果与模型组比较,清热排毒胶囊各剂量组和戴芬组大鼠踝关节浸出液中 NF-κB 和 IL-1β的表达水平均显著降低（P＜0.05,P＜0.01）。病理观察结果显示,清热排毒胶囊可减轻模型大鼠踝关节滑膜组织充血、水肿及炎性细胞浸润,并改善滑膜增生等病理改变。结论清热排毒胶囊通过抑制大鼠踝关节组织中NF-κB和IL-1β的表达来发挥抗炎作用。     

清胰Ⅱ号颗粒剂对大鼠重症急性胰腺炎肝损伤的保护作用

目的探讨NF-κB在重症急性胰腺炎(SAP)合并肝损伤过程中的作用,并观察清胰Ⅱ号颗粒剂对重症急性胰腺炎肝损伤的保护作用。方法 24只SD大鼠随机分为3组:假手术组(A组)、胰腺炎组(B组)、清胰Ⅱ号治疗组(C组)。A组仅开腹不制作SAP模型,B、C组以3%牛磺胆酸钠逆行注入大鼠胆胰管制作SAP模型,C组给予清胰Ⅱ号治疗,造模后于24 h取材。取肝组织采用电泳迁移率试验(EMSA)法检测NF-κB活性,ELISA法检测血清白细胞介素(IL)-1、IL-6、IL-8,同时观察血淀粉酶、肝功能;余肝组织浸入10%甲醛溶液以便病理检查。结果 A组肝组织中几乎测不到NF-κB活性,与A组比较B组肝组织NF-κB活性明显增高(P<0.05),B组血清IL-1、IL-6、IL-8浓度也异常升高(P均<0.05);血清AST、ALT、LDH也显著升高(P均<0.01);C组与B组相比,肝组织NF-κB活性显著下降(P<0.01);同时血清IL-1、IL-6、IL-8浓度也显著下降(P均<0.01),血清AST、ALT、LDH显著降低(P均<0.01)。C组肝组织病理损害较B组减轻。结论肝组织NF-κB活化参与了大鼠SAP肝损伤的发病过程,清胰Ⅱ号颗粒剂可以减轻SAP肝损害,其机制可能为抑制NF-κB活性,减少炎性递质释放。     

百事乐加味方对脑卒中后抑郁症大鼠海马-前额叶皮层环路炎症平衡的影响

目的研究百事乐加味方(MBD)对脑卒中后抑郁症(Post-stroke depression,PSD)大鼠炎症因子IL-1β、IL-6、TNF-α含量及海马(HP)、前额叶皮层(PFC)组织NF-κB表达水平的影响,从炎症平衡角度探讨复方中药防治该病可能机制。方法采用MCAO+CUMS+孤养法复合因素建立PSD模型,记录大鼠每周体重;采用ELISA方法,检测PSD大鼠血清炎症因子IL-1β、IL-6和TNF-α的含量;采用免疫组化技术检测HP、PFC组织NF-κB的表达水平。结果对体质量影响,与PSD大鼠比较,MBD组在应激第14、21、28天能够恢复模型大鼠体重,差异具有统计学意义(P<0.01~0.05);对炎症相关指标影响,与PSD大鼠比较,MBD高剂量组明显降低大鼠血清中IL-1β,IL-6、TNF-α含量,明显降低大鼠HP、PFC组织NF-κB水平,差异有统计学意义(P<0.01~0.05);MBD低剂量组明显降低大鼠血清中IL-6,TNF-α含量及大鼠HP、PFC组织NF-κB表达水平,差异有统计学意义(P<0.05)。与卒中组比较,MBD高剂量组明显降低大鼠血清中IL-1β、IL-6含量,同时降低HP、PFC组织NF-κB的表达水平,差异有统计学意义(P<0.05);低剂量治疗组HP、PFC组织NF-κB水平明显降低,差异有统计学意义(P<0.01~0.05)。与抑郁组比较,MBD高剂量组明显降低大鼠血清中IL-1β含量,HP、PFC组织NF-κB的表达水平明显降低,差异有统计学意义(P<0.01~0.05);MBD低剂量组HP、PFC组织的NF-κB表达水平略降低,但差异不具有统计学意义(P>0.05)。结论 MBD能有效抑制PSD大鼠相关炎症反应因子,同时降低HP、PFC组织NF-κB表达水平,改善PSD大鼠脑神经功能,从而发挥治疗作用,是其抗脑卒中后抑郁重要机制之一。     

芒针对脊髓损伤大鼠运动功能和炎症水平的影响

目的探究芒针深刺秩边穴对大鼠脊髓损伤后运动功能的影响及可能作用机制。方法选择健康雄性Wister大鼠81只,随机分为正常组、模型组和芒针组(正常组9只,其余两组各36只),采用改良Allen's造模法制备大鼠脊髓中度损伤模型,模型组不做特殊处理,芒针组采用芒针深刺秩边穴,每日1次,每次30 min。分别于术后1 d、3 d、5 d、7 d行BBB(Basso-Beattie-Bresnahan)运动功能评分;术后1 d、3 d、5 d、7 d取受损段脊髓组织行酶联免疫吸附测定(ELISA)、实时荧光定量PCR(RT-qPCR)和苏木素-伊红染色(HE染色)。结果术后5 d和7 d,芒针组大鼠BBB评分高于模型组,差异有统计学意义(P<0.01);脊髓损伤后,模型组和芒针组大鼠脊髓组织中高迁移率族蛋白B1(HMGB1)、核转录因子kB(NF-kB)、白介素-6(IL-6)含量及HMGB1mRNA、NF-kBmRNA、IL-6mRNA水平显著升高(P<0.05);受损的脊髓组织松散,灰质中有许多空洞形成,伴有炎性细胞浸润。芒针治疗后,芒针组大鼠脊髓组织中HMGB1、NF-kB、IL-6含量及HMGB1mRNA、NF-kBmRNA、IL-6mRNA水平较模型组降低,且在3 d、5 d、7 d差异有统计学意义(P<0.05);受损部位的空洞及炎性细胞逐渐减少。结论脊髓损伤后,炎症因子的大量聚集引起级联性炎症反应,影响大鼠运动功能的恢复。芒针的抗炎机制可能包括抑制HMGB1的表达,降低NF-kB信号通路的传导,下调促炎因子IL-6的分泌。     

白术内酯Ⅰ对慢性萎缩性胃炎大鼠胃黏膜保护作用

目的:探讨白术内酯Ⅰ对慢性萎缩性胃炎(CAG)大鼠胃黏膜保护机制。方法:实验动物随机分为空白对照组、模型组、白术内酯Ⅰ组(10、20、40 mg/kg),采用脱氧胆酸钠和热盐水交替灌胃、结合饥饱失常方法制备CAG大鼠模型,治疗8周后,采用ELISA法检测白细胞介素-8(IL-8)含量、Western blot法检测热休克蛋白70(HSP 70)、核转录因子-kB(NF-kB)和环氧合酶-2(COX-2)蛋白表达。结果:与空白对照组对比,模型组HSP70蛋白表达降低(P0.01),IL-8含量(P0.01)增加、NF-κB和COX-2蛋白表达升高(P0.01);与模型组对比,白术内酯Ⅰ高、中、低3个剂量组提高HSP70蛋白表达,降低IL-8含量、NF-κB和COX-2蛋白表达(P0.05),以白术内酯Ⅰ组(40 mg/kg)作用显著。结论:白术内酯Ⅰ对CAG大鼠的保护作用是通过抗炎作用。     

不同灸量对溃疡性结肠炎大鼠结肠上皮形态学及血清中炎性细胞因子、结肠组织中炎性细胞信号转导通路的影响

目的:从形态学、免疫学、分子生物学角度探讨不同灸量治疗溃疡性结肠炎(UC)效果的异同。方法:32只SD大鼠随机分为空白组6只、模型复制组26只。采用三硝基苯甲酸/葡聚糖硫酸钠制备UC大鼠模型。模型复制成功后的大鼠按随机数字表分为模型组、3壮组、6壮组、9壮组,每组各6只。各治疗组所取穴位为"天枢""大横",艾炷直接灸法,每次分别施灸3壮(3min)、6壮(6min)、9壮(9min),共治疗14次。观察大鼠治疗前后疾病活动指数(DAI),电镜、光镜观察结肠组织形态学改变,酶联免疫法检测大鼠血清中白介素-8(IL-8)、白介素-10(IL-10)含量,Western blot法检测大鼠结肠中Toll样受体9(TLR-9)和核转录因子-κB(NF-κB)p 65表达。结果:灸法可明显降低大鼠DAI的评分(与模型组比较均P0.05);光镜与电镜结果显示,灸量越大,结肠组织腺体排列越规则。模型组血清IL-8含量升高,IL-10含量降低;与模型组比较,各治疗组IL-8降低,IL-10增高,其中9壮组和6壮组的变化较3壮组更明显(均P0.05)。模型组结肠组织中TLR-9、NF-κB p 65大量表达;与模型组比较,各治疗组TLR-9、NF-κB p 65表达均降低,且以9壮组的变化最为明显(均P0.05)。结论:艾灸可修复UC大鼠受损黏膜上皮,抑制血清中IL-8含量,提高血清中IL-10含量,通过抑制结肠组织中NF-κB p 65转录而下调TLR-9表达。灸量越大治疗效果越明显。     

养阴清热化瘀方对急性放射性肺炎大鼠TLR-4/NF-κB信号通路的影响

目的:探讨养阴清热化瘀方对大鼠急性放射性肺炎的防护作用及可能机制。方法:采用随机数字表法将48只雌性Wistar大鼠随机分为空白对照组(NC)、模型组(MD)、养阴清热化瘀方提前干预组(YHF-EI)及养阴清热化瘀方组(YHF-ST),分别于照射后4周时采集血清及取出右肺。观察大鼠的一般情况,应用HE染色观察肺组织病理改变,ELISA法检测血清中含量, RT-PCR和Western-blotting法分别检测肺组织内TLR4、NF-κB p65 mRNA和蛋白的水平。结果:养阴清热化瘀方能显著减轻大鼠放射性肺炎的炎症表现; 与空白对照组比较,血清中TNF-α、IL-6、TGF-β的含量升高,TLR4和NF-κB p65 mRNA转录活性增强,肺组织中TLR4和NF-κB p65的蛋白表达水平升高; 与模型组比较,两组养阴清热化瘀方组血清中TNF-α、IL-6、TGF-β的含量降低,TLR4和NF-κB p65 mRNA转录活性下降,肺组织中TLR4和NF-κB p65的蛋白表达水平降低。结论:养阴清热化瘀方可能通过抑制“TLR4/NF-κB”信号通路,从而抑制TNF-α、IL-6、TGF-β的产生,起到抵抗放射性肺炎的发生,从而发挥放射防护作用。     

白细胞介素-23及核因子-κB在大鼠脑梗死模型中的表达

目的探讨白细胞介素-23(IL-23)及核因子-κB(NF-κB)在脑梗死模型大鼠脑梗死区域炎性损伤组织中的表达情况。方法健康成年雄性SD大鼠50只,随机分为梗死组和假手术组,每组25只。梗死组用线栓法制作左侧大脑中动脉闭塞模型,2组均于手术后6 h、24 h、48 h、72 h、7 d随机取5只大鼠取材,采用免疫组化染色,观察脑梗死区域光镜下形态学变化及IL-23、NF-κB的表达。比较2组大鼠在同一时间点脑组织形态学变化及IL-23、NF-κB表达的异同。结果梗死组IL-23、NF-κB阳性细胞数各时间点均高于假手术组(P均<0.01)。梗死组IL-23在梗死后6 h表达增高,并且在梗死后48 h达到高峰,3d后阳性细胞数表达逐渐减少,各时间点比较具有显著性差异(P均<0.01)。梗死组NF-κB在梗死后48 h达到高峰,3 d后阳性细胞数表达逐渐减少,各时间点比较具有显著性差异(P均<0.01)。结论 IL-23和NF-κB在大鼠脑梗死区域表达增高,参与了梗死区域的炎性反应。     

小钻木脂素类化合物戈米辛R的体外抗炎机制研究

目的 通过脂多糖(LPS)诱导活化小鼠单核巨噬细胞系RAW264.7建立炎症模型,探讨小钻木脂素类化合物戈米辛R对炎症细胞增殖及炎症因子分泌的影响,评价其抗炎活性及机制。方法 采用MTT比色法检测经LPS诱导的RAW264.7细胞存活率并计算半数抑制浓度(IC₅₀);采用ELISA法检测细胞上清液TNF-α、IL-1β、IL-6等炎症因子含量;利用RT-PCR法检测细胞TNF-α、IL-1β、IL-6、IκB-α、NF-κB p65的mRNA表达量;Western blot法检测细胞IκB-α及NF-κB p65蛋白表达量。结果 与LPS模型组比较,戈米辛R可显著抑制LPS诱导的RAW264.7细胞增殖;降低细胞分泌TNF-α、IL-1β及IL-6的含量;升高LPS所致降低的IκB-α mRNA的表达量同时降低LPS引起升高的TNF-α、IL-1β、IL-6、NF-κB p65的mRNA表达量;升高IκB-α并降低NF-κB p65的蛋白表达量,显示出良好的剂量依赖性,具有统计学差异(P<0.05)。结论 戈米辛R可通过抑制TNF-α、IL-1β、IL-6、NF-κB p65的mRNA和NF-κB p65蛋白的表达,增加IκB-α mRNA和IκB-α蛋白的表达,减少致炎因子TNF-α、IL-1β、IL-6的释放,从而发挥显著抗炎活性。     

温肾活血化湿方对肾间质纤维化大鼠肾组织NF-κB表达的影响

目的:探讨温肾活血化湿方对单侧输尿管梗阻(unilateral ureteral obstruction,UUO)法致肾间质纤维化大鼠肾组织核转录因子-κB(NF-κB)表达的影响,初步揭示该方防治肾间质纤维化的作用机制。方法:采用UUO建立大鼠肾间质纤维化模型,将雄性SD大鼠80只随机分为假手术组、UUO组、氯沙坦组和温肾活血化湿方组,每组20只;于造模后第2天开始灌胃,假手术组、UUO组给予等量生理盐水,氯沙坦组灌胃氯沙坦[10 mg/(kg·d)],温肾活血化湿方组灌温肾活血化湿方汤剂[9.8 mg/(kg·d)]。于造模后第7、14、21天留取梗阻侧肾组织行HE、Masson染色,观察肾组织病理变化,免疫组化法检测肾组织NF-κB的表达。结果:第7、14、21天UUO组中NF-κB表达较假手术组高(P0.05),治疗组在第7天较UUO组低(P0.05)。结论:温肾活血化湿方可能通过抑制NF-κB的过度表达从而延缓肾间质纤维化的进程。     

电针颈夹脊穴对神经根型颈椎病神经病理性疼痛模型大鼠脊髓背角GFAP、NF-κB及炎性细胞因子表达的影响

目的:通过观察电针颈夹脊穴对神经根型颈椎病(CSR)模型大鼠脊髓背角神经胶质纤维酸性蛋白(GFAP)、核转录因子B(NF-κB)及炎性细胞因子表达的影响,探讨电针颈夹脊穴治疗神经根型颈椎病所致神经病理性疼痛潜在的镇痛机制。方法:将62只SD大鼠随机分为空白组、假手术组、模型组、电针组和星形胶质细胞抑制剂L-α-氨基己二酸(LAA)组。除空白组外,所有大鼠均进行CSR造模和鞘内置管处理,假手术组只暴露神经根而不结扎。空白组无任何干预,常规饲养;假手术组和模型组仅在干预时同电针组进行捆绑操作,无其他干预;电针组采取电针双侧颈夹脊穴干预,20 min/次;LAA组采取鞘内置管LAA干预。4组均从造模后第7天开始干预,1次/d。采用免疫荧光法观察脊髓背角NF-κB、GFAP的表达;采用real-time PCR技术观察脊髓背角组织中炎性细胞因子IL-1β、IL-6、IL-18、TNF-αmRNA的表达。结果:空白组与假手术组NF-κB、GFAP和IL-1β、IL-6、IL-18及TNF-αmRNA的表达差异无统计学意义(P>0.05)。与空白组比较,模型组NF-κB、GFAP和IL-1β、IL-6、IL-18及TNF-αmRNA的表达显著升高(P<0.05)。与模型组比较,电针组和LAA组NF-κB、GFAP和IL-1β、IL-6、IL-18及TNF-αmRNA的表达明显降低(P<0.05)。电针组与LAA组比较,大鼠脊髓背角NF-κB、GFAP和IL-1β、IL-6及IL-18 mRNA的表达差异有统计学意义(P<0.05),TNF-αmRNA的表达差异无统计学意义(P>0.05)。结论:电针颈夹脊穴治疗神经根型颈椎病所致神经病理性疼痛的镇痛机制可能与抑制星形胶质细胞活化,下调NF-κB、IL-1β、IL-6、IL-18及TNF-α的表达有关。     

Electroacupuncture at “Zusanli(ST36)” alleviates 5-fluorouracil-induced renal injury in colorectal cancer-bearing mice by exerting effects on oxidative stress,inflammatory responses,and cell apoptosis: 电针“足三里”减轻大肠癌荷瘤小鼠5-FU化疗后肾损伤及其氧化应激、炎症反应和细胞凋亡的研究

ObjectiveTo observe the effect of electroacupuncture at “Zusanli (ST36)” on 5-fluorouracil (5-FU)-induced renal injury in colorectal cancer-bearing mice, and to further investigate its effects on oxidative stress, inflammatory responses, and cell apoptosis in mouse renal tissue.MethodsThirty-five male BALB/c mice were randomly divided into five groups of seven mice each, namely the control, CT26, 5-FU, sham point (SP), and ST36 (which received EA at the “ST36”) groups. With the exception of the control group, each group was subjected to establishment of a subcutaneous implantation tumor model using the murine CT26 colorectal cancer cell line. Once the models were successfully established, the 5-FU, SP, and ST36 groups received 5-FU injection solution intraperitoneally at a dose of 5 mg/mL once every three days over a 21-day period. Mice in the SP and ST36 groups additionally received an EA intervention after each intraperitoneal 5-FU injection. EA were performed on mice of the SP group at bilateral sham acupoints and on mice of the ST36 group at the bilateral “ST36” using the continuous wave mode at a frequency of 2 Hz for a duration of 5 min, intervention was administered once every two days for a duration of 21 days. Samples were collected from the mice at the end of the experiment. The pathological morphology of the renal tissue was observed using hematoxylin and eosin (HE) staining; the contents of creatine (Cre), blood urea nitrogen (BUN) and malondialdehyde (MDA), and superoxide dismutase (SOD) activity were measured using biochemical assays; the expression and nuclear translocation of nuclear factor kappa B p65 subunit (NF-κB p65) were measured by immunofluorescence; the expression levels of tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6), and interleukin-1β (IL-1β) in serum were measured by ELISA; cell apoptosis in renal tissue was detected using the TUNEL assay; and the expression levels of the anti-B-cell lymphoma/leukemia 2 (Bcl-2), Bcl-2-associated X protein (Bax), cleaved caspase-3, cleaved caspase-9, and cytochrome C (cyt c) in renal tissue were measured by Western blotting.ResultsCompared with the control group, mice of the CT26 group showed a significant increase in serum Cre content (P<0.01), but the difference in BUN content was not statistically significant (P>0.05). HE staining of renal tissue revealed clear structures with normal glomerular and renal tubular morphology. SOD activity was decreased (P<0.01); MDA content was increased, but the increase was not statistically significant (P>0.05). Differences in NF-κB p65 protein expression in the cytoplasm of renal tissue and serum levels of TNF-α, IL-6, and IL-1β were not statistically significant (all P>0.05). Results of immunofluorescent TUNEL staining indicated an absence of significant cell apoptosis. In the renal tissue, Bcl-2 protein expression was slightly increased (P<0.05), and the expression levels of Bax (P<0.01), cleaved caspase-3 (P>0.05), cleaved caspase-9 (P<0.01), and cyt c (P>0.05) were increased. Compared with the CT26 group, mice of the 5-FU group exhibited an increase in Cre (P<0.01) and BUN (P<0.05) content. HE staining of renal tissue revealed the presence of glomerular atrophy and dilated Bowman's capsular spaces. SOD activity was significantly decreased (P<0.01), while the increase of MDA content was not significant (P>0.05). The expression of NF-κB p65 in the nucleus and serum TNF-α, IL-6, and IL-1β levels showed significant increases (P<0.05). The cell apoptosis level was significantly increased. The protein expression of Bcl-2 (P<0.05), Bax (P<0.01), cleaved caspase-9 (P<0.01), and cyt c (P<0.05) was significantly increased. Compared with the 5-FU group, the ST36 group showed decreased serum Cre and BUN levels (both P<0.01). HE staining of renal tissue showed less renal tissue injury and less dilation of renal capsular cavities. SOD activity was significantly higher (P<0.01), while MDA content was lower (P<0.05). Nuclear expression of NF-κB p65 and serum TNF-α (P<0.05), IL-6 (P<0.05), and IL-1β (P<0.01) levels were lower. The cell apoptosis level was decreased relative to the 5-FU group. Bcl-2 protein expression was significantly increased (P<0.01), while the protein expression levels of Bax (P<0.01), cleaved caspase-3 (P<0.01), cleaved caspase-9 (P<0.01), and cyt c (P<0.05) also were reduced.ConclusionEA at “ST36” attenuated 5-FU-induced renal injury in colorectal cancer-bearing mice. The mechanism may be related to the regulation of renal oxidative stress, alleviation of inflammatory responses, and inhibition of cell apoptosis.     

Protective effect of hydroxysafflor yellow A on MSCs against senescence induced by D-galactose

Objective To examine the protective effects of hydroxysafflor yellow A (HSYA) against the senescence of mesenchymal stem cells (MSCs) induced by D-galactose (D-gal) in vitro, and investigate the potential mechanism involved. Methods Grouping experiment, Normal control (NC) group: conventional culture with complete medium; Senescence group: MSCs were cultured for 48 h with complete medium containing 10 g/L D-gal; HSYA group: on the basis of senescence induction, HSYA with the suitable concentration was used to protect MSCs. The key experimental indices associated with oxidative stress, inflammatory response, cell senescence, proliferation and apoptosis were measured through chemical colorimetry, β-galactosidase staining, EdU incorporation and flow cytometry, respectively. The relative quantity (RQ) of proteins related closely to cell proliferation, apoptosis, and NF-κB signaling were measured by Western blotting. Results As compared with Senescence group, treatment with HSYA (120 mg/L) effectively ameliorated the adverse situation of MSCs. Oxidation stress and inflammation along with D-Gal induction was dramatically alleviated in MSCs; The β-Gal-positive staining indicated that MSC senescence was significantly mitigated; The proliferative capability of MSCs was significantly increased by up-regulating PCNA and inhibiting p16 expression; The anti-apoptotic effect on MSCs was exerted by down-regulating the RQ of cleaved Caspase-3 and Bax; The activity of NF-κB signaling in MSCs was notably suppressed through inhibiting phosphorylation of IKKβ and p65. Conclusion HSYA (120 mg/L) significantly delayed the D-Gal-induced senescence process in MSCs through attenuating inflammatory reaction and oxidative stress, and suppressing the activity of NF-κB signaling.     

基于TLR4/NF-κB信号通路探讨加味升降散对膜性肾病大鼠肾组织纤维化的影响

目的:观察加味升降散对膜性肾病(MN)大鼠肾组织纤维化的影响,探讨其防治MN及肾纤维化的作用机制。方法:大鼠尾静脉注射阳离子化牛血清白蛋白(C-BSA)复制MN大鼠模型,将造模成功的MN大鼠随机分为模型组、加味升降散组(27.3 g·kg^-1)和贝那普利组(10 mg·kg^-1),分别给予相应剂量药物灌胃,1次/d,连续干预4周。给药结束后,采用马松(Masson)染色,碘酸六胺银(PASM)染色,免疫荧光和透射电镜(TEM)等方法观察大鼠肾脏病理学变化;酶联免疫吸附测定(ELISA)检测大鼠血清中白细胞介素-1β(IL-1β),白细胞介素-6(IL-6),肿瘤坏死因子-α(TNF-α)含量;实时荧光定量聚合酶链式反应(Real-time PCR)和免疫组化分别检测大鼠肾组织中单核细胞趋化蛋白-1(MCP-1),细胞间黏附分子-1(ICAM-1),核转录因子-κB(NF-κB),Toll样受体4(TLR4),纤溶酶原激活物抑制剂-1(PAI-1),转化生长因子-β(1TGF-β1),α-平滑肌肌动蛋白(α-SMΑ)和Ⅳ型胶原(CollagenⅣ)mRNA和蛋白表达水平。结果:与正常组比较,模型组大鼠肾组织纤维化明显,血清中IL-1β,IL-6,TNF-α含量明显升高(P<0.05),肾组织中MCP-1,ICAM-1,NF-κB,TLR4,PAI-1,TGF-β1,α-SMA和CollagenⅣmRNA及蛋白表达量均明显升高(P<0.05);与模型组比较,加味升降散和贝那普利可明显改善大鼠肾脏纤维化,明显降低MN大鼠血清中IL-1β,IL-6,TNF-α含量(P<0.05),明显下调肾组织中MCP-1,ICAM-1,NF-κB,TLR4,PAI-1,TGF-β1,α-SMA和CollagenⅣmRNA及蛋白表达量(P<0.05)。结论:加味升降散可通过下调TLR4/NF-κB信号通路,减少炎症因子的释放和表达,抑制肾纤维化,减轻MN大鼠肾损伤。     

豨莶草醇提物调控TLRs/NF-κB通路及NLRP3炎症小体干预痛风性关节炎的机制研究

目的探讨豨莶草醇提物通过Toll样受体(TLRs)/核因子κB(NF-κB)信号通路及NOD样受体蛋白3(NLRP3)干预痛风性关节炎(GA)的作用机制。方法采用脂多糖(LPS)联合尿酸钠结晶(MSU)刺激急性单核细胞白血病细胞(THP-1)源性巨噬细胞,建立GA炎症细胞模型;以豨莶草醇提物300、200、100μg·mL^-1不同浓度进行干预。采用CCK-8法检测细胞活力;ELISA法检测细胞上清中白细胞介素-1β(IL-1β)的水平;qRT-PCR法检测TLRs/NF-κB信号通路相关基因(TLR2、TLR4、MYD88、IRAK-1、TRAF-6、TAK-1、NF-κB、IL-1β)及NLRP3 m RNA的表达;Western Blot法检测NF-κB、NLRP3、IL-1β蛋白的表达。结果与对照组比较,豨莶草醇提物300、200、100μg·mL^-1浓度对THP-1巨噬细胞活力均无明显影响(P>0.05);模型组的IL-1β水平明显升高(P<0.05),TLRs/NF-κB信号通路相关基因及NLRP3 mRNA表达水平均明显上调(P<0.05),NF-κB、NLRP3、IL-1β蛋白表达水平明显上调(P<0.05)。与模型组比较,豨莶草醇提物300、200、100μg·mL^-1浓度组的细胞IL-1β水平明显降低(P<0.05),TLRs/NF-κB信号通路相关基因及NLRP3 mRNA表达水平均不同程度下调(P<0.05),NF-κB、NLRP3及IL-1β蛋白表达水平亦不同程度下调(P<0.05)。结论豨莶草醇提物可能通过调控TLRs/NF-κB信号通路及抑制NLRP3炎症小体活化,减少炎症因子的产生与释放,进而减轻痛风性炎症反应。