目的 探讨壮通饮对脑缺血再灌注小鼠神经细胞损伤的影响及其作用机制.方法 采用线栓大脑中动脉栓塞法诱导建立小鼠脑缺血再灌注模型,造模成功后的 6～7 周龄雄性C57BL6J小鼠随机分为模型组、壮通饮组、阿司匹林组、壮通饮联合阿司匹林组,各组再随机分为 1、3、7 d时点组,另设不造模的空白组和假手术组,每组 10 只,按照分组灌胃给药至相应时点.记录各组每日体质量及Zea Longa评分,采用TTC染色法测定脑缺血小鼠的脑梗死体积百分比;HE染色观测缺血病理改变情况,TUNEL染色比较神经细胞凋亡情况,透射电镜观测神经细胞线粒体结构改变及自噬情况,Western blot以及RT-PCR技术分析P62、LC3Ⅱ/LC3Ⅰ、Bc1-2/腺病毒E1B19kD相互作用蛋白 3(Bc1-2/adenovirusE1B 19kDa interacting protein3,BNIP3)、β-actin蛋白或基因表达情况.结果 与空白组、假手术组相比,模型组体质量明显下降(P＜0.05),TTC染色可见脑组织缺血梗死,出现神经细胞凋亡及线粒体损伤.与模型组相比,各给药组体质量升高(P＜0.05),行为学评分降低(P＜0.05),梗死体积降低,且持续减小(P＜0.05);神经细胞凋亡率降低(P＜0.05).模型组线粒体结构损伤明显较重,在 1d时各给药组即可观测到较多线粒体自噬小体,而模型组在 3d时才可观测到,在 7d时各给药组大多数线粒体结构完整,可见被溶酶体消化完成的脂滴,而模型组可见自噬小体与自噬溶酶体同时存在.与模型组比较,各给药组P62蛋白表达在1d时降低,3、7 d时升高(P＜0.05);LC3Ⅱ/LC3Ⅰ的蛋白比值以及P62、BNIP3的基因表达水平在1d时升高,7 d时降低(P＜0.05).在各给药组中,壮通饮联合阿司匹林组各项指标均优于其他给药组及模型组(P＜0.05).结论 壮通饮能够减轻脑缺血再灌注小鼠的神经细胞损伤,这可能与促进神经细胞发生线粒体自噬,维持细胞内线粒体稳态有关.

目的:观察壮通饮(ZTY)对缺氧/复氧诱导的人脐静脉血管内皮细胞(HUVEC)和人星形细胞瘤U251细胞损伤模型的作用,探讨壮药保护血脑屏障的相关机制.方法:将8只SD大鼠随机分为4组:空白组(蒸馏水)及低、中、高剂量(3.74、7.48和14.96 g/kg)ZTY含药血清组,每天1次,连续7 d灌胃饲养,获取含药血清.将HUVEC和U251细胞共培养,构建缺氧/复氧诱导的体外细胞屏障损伤模型.将细胞分为5组:空白组、模型组及低、中、高剂量ZTY含药血清组.RT-qPCR定量检测各组HUVEC中基质金属蛋白酶2(MMP2)、MMP3、MMP9和内皮素1(ET-1)的mRNA表达水平;Western blot检测各组HUVEC中紧密连接蛋白(TJPs)和ET-1蛋白的表达水平.结果:与模型组比较,给药组的跨内皮细胞电阻显著增大(P<0.05),MMP2、MMP3、MMP9和ET-1的mRNA表达水平显著降低(P<0.05),TJPs的表达水平显著升高(P<0.05),ET-1蛋白表达水平显著降低(P<0.05).结论:ZTY通过上调TJPs、下调MMPs和ET-1表达而减轻缺氧/复氧对HUVEC/U251细胞屏障的损伤.

目的:观察壮通饮对脑缺血再灌注损伤模型大鼠脑缺血损伤的保护作用,并从时钟基因Bmal1调控昼夜节律途径初步探讨其保护作用的机制.方法:取SD大鼠60只,按6、12、18、24时四个时间点随机分为4组,每组大鼠随机分为:假手术组、模型组、壮通饮组.观察昼夜节律对脑缺血模型大鼠神经功能缺损评分,脑梗死体积,脑含水量,Bmal1蛋白表达水平的影响以及壮通饮干预后每组大鼠各指标的变化情况.结果:各组大鼠均有明显的神经功能缺损症状,其中6:00组大鼠神经缺损症状明显重于24:00组,24:00组大鼠脑梗死体积明显小于6:00组,18:00、24:00组大鼠脑含水量明显小于6:00组,18:00、24:00组大鼠脑组织Bmal1表达量明显高于6:00组.壮通饮干预后,24:00组大鼠神经缺损症状较6:00组大鼠明显减轻,18:00、24:00组大鼠脑梗死体积,脑含水量较6:00,12:00组明显下降,24:00组大鼠脑组织Bmal1表达水平较6:00组明显增高.结论:壮通饮可通过调控时钟基因Bmal1活性来调控昼夜节律发挥脑缺血损伤保护作用.

Objective] To explore the effect of modified Simiao Yong’an decoction on collagen induced arthritis about arthritis index (AI), the joint tissue inflammatory pathological manifestations, and effect on protein expression of IL-6,IL-17,TNF-αin rats. [Methods] The successful CIA models of SD rats were randomly divided into model group, positive group (leflunomide), and high, medium, low dose group of Chinese medicine, and normal control group, 6 rats in each group. The intervention and treatment period was 12 weeks. AI val-ues of each rat were evaluated every week. At 12 weeks, rats were executed and right ankle joint of each group of rats was taken. Joint pathlological changes of each right ankle were observed. At the same time levels of IL-6, IL-17 and TNF-αin serum were examined by ELISA, mRNA transcription levels of IL-6, IL-17and TNF-αin synovial were detected by RT-PCR. [Results] AI scores of the three Chi-nese medicine groups decreased significantly (P﹤0.05 or P﹤0.01) in 6~12 weeks compared with the model group, and in the 8~10 weeks AI scores were even improved significantly than positive group (P﹤0.05 or P﹤0.01). Histological examination showed that the ankle joint cartilage pathological had an improvement in the medium dose of modified Simiao Yong’an decoction group compared with the control model group. Compared with the model group, mRNA and protein expression of IL-6, IL-17and TNF-αwere down regulated in all treat-ment groups significantly (P﹤0.01). [Conclusion] The anti-inflammatory action mechanism on CIA rats of modified Simiao Yong’an de-coction might achieve by reducing expression of IL-6, IL-17 and TNF-α.

目的:观察秦苓液对尿酸性肾损害大鼠肾功能及肾组织病理改变的影响,探讨其对尿酸性肾病(UN)肾脏损伤的保护作用及作用机制.方法:采用腺嘌呤灌胃伴酵母饲料喂养的方法造模,将成模大鼠随机分为正常组,模型组,别嘌呤醇组(23.33 mg· kg-·d-1),秦苓液大、中、小剂量组(36.4,18.2,9.1g·kg-·d-1),连续干预8周.分别于0,2,4,6,8周检测各组大鼠24 h尿蛋白(uPRO/24 h),血尿酸(sUA),血肌酐(sCR)和血尿素氮(BUN);干预6周和8周后分别处死各组大鼠1/2量,取肾脏进行苏木素-伊红和过碘酸雪夫氏染色,光镜下观察.结果:与正常组比较,模型组大鼠uPRO/24 h,sUA,sCR,BUN明显升高,肾组织出现较明显的病理改变;与模型组比较,秦苓液各剂量组大鼠uPRO/24 h,sUA,sCR,BUN有不同程度的降低,肾组织病理损伤明显减轻.结论:秦苓液能显著改善UN模型大鼠肾功能,减轻肾组织病理损伤,保护肾脏.

Objective To observe the protective effect of Tangshenqing Formula II ( TSQF) on the renal function of mice with type 2 diabetes mellitus ( DM) . Methods KK-Ay mice, a spontaneous DM mod-el, were fed with fat-enriched diet for three weeks. Based on their serum glucose ( Glu) and body weight ( BW) , the mice were divided into model group, valsartan group, high, middle, and low dose of TSQF ( high-, mid-, low- dose ) group ( n =6 each ) and were treated for consecutive 12 weeks. And six C57BL/6J mice were included into control group. At the timepoint of Week 0, 4, 8, 12, Glu, urine al-bumin ( Alb) , urine creatine ( Cr) , Alb/Cr( ACR) and 24-h urine protein ( pr/24 h) were detected. At the end of Week 12, all the mice were sacrificed and their serum samples were collected. And the serum levels of TG, TC, Scr, BUN and β2-MG were measured. Results Compared with the control group, the levels of pr/24 h, ACR, Scr, TG and TC in the model group significantly increased ( P<0. 05 or P<0. 01), While BUN,β2-MG and ratio of kidney weight to body weight showed no statistcal differences (P>0. 05). Compared to the model group, the levels of Scr, TG and ACR in valsartan group and three TSQF groups significantly decreased (P<0. 05 or P<0. 01), and those of TC, Glu, and pr/24 h de-creased at different degree. Conclusion TSQF could reduce the serum levels of Glu, Scr and TG of kk-Ay mice, and alleviate the kidney injuries to reduce the progression of diabetic nephropathy.

OBJECTIVE:To explore the molecular mechanism of exocrine immune inflammatory injury of Sjögren's Syndrome and the intervention of Banxia Qinlian Decoction (BQD).METHODS:Totally 18 female NOD mice were randomly divided into the model group, the positive drug group, and the BQD group, 6 in each group. Six female BALB/c mice were recruited as a blank control group. Mice in the blank control group and the model group were gavaged with deionized water at the daily dose of 0.1 mL/10 g body weight. Tripterygium Tablet was administered by gastrogavage to mice in the positive group at the daily dose of 10 mg/kg. BQD was administered by gastrogavage to mice in the BQD group at the daily dose of 60 g crude drugs/kg. After 12 weeks of medication, mice were sacrificed. Their eyeballs were excised and blood collected. Tissues of bilateral parotids and submandibular glands were kept. mRNA transcriptional levels of IL-17, IL-6, type 3 muscarinic acetylcholine receptors (M3R), aquaporin protein-5 (AQP5) were detected by RT-PCR. Expression levels of M3R and AQP5 protein were detected by Western blot. Protein expression levels of IL-17 and IL-6 were detected by ELISA.RESULTS:Compared with the normal group, mRNA transcriptional levels and protein expression levels of IL-17, IL-6, M3R, and AQP5 were significantly up-regulated in the model group (P < 0.01). Compared with the model group, mRNA transcriptional levels and protein expression levels of IL-17, IL-6, M3R, and AQP5 were significantly down-regulated in the positive drug group and the BQD group with statistical difference (P < 0.01, P < 0.05). Compared with the BQD group, mRNA-transcriptional levels of IL-17, IL-6, and M3R, as well as M3R and AQP5 protein expression levels were significantly down-regulated in the positive drug group (all P < 0.01).CONCLUSION:The molecular mechanism of BQD in inhibiting SS exocrine neurotoxic injury might be possibly related to regulating Th17/IL-17 immune inflammatory way.

Objective To explore the effects of Chinese herbal compound, Tengmei decoction, on type II collagen-induced arthritis ( CIA) in rats, and to examine the changes of arthritis index ( AI) , limb swelling, joint tissue inflammatory infiltration, and the effects on immune-inflammatory factors.Methods Sprague-Dawley rat models of arthritis were successfully established by intradermal injection of type II collagen and Freund’ s complete adjuvant.The model rats were randomly divided into model group, positive drug group, and high-and low-dose Chinese medicine groups, 6 rats in each group.The intervention and treatment period was 12 weeks.To measure weekly the anteroposterior and transverse diameters of the rear ankles and wrists, the transverse diameter of the claw foot palm pad, the thickness and the highest point width of hind limb plantar joint swelling, and to evaluate the integrated scores of joints and limbs swelling using a vernier caliper.Results ①Compared with the normal group, the total arthritis scores and hind limbs AI scores of the model group were significantly increased ( P <0.05 or P <0.01 ) .The left forelimb AI scores were significantly increased during 10 -12 weeks ( P <0.05 ) .The anteroposterior and transverse diameters of the left hind limb, the thickness of the highest point measurement of the left hind foot pad metatarsal were significantly increased ( P<0.05 or P<0.01) in different time periods between 1-12 weeks.Compared with the model group, the total scores and the left hind limb joints AI scores of the high-and low-dose drug groups were decreased after 6 weeks (P<0.05).②Compared with the normal control group, levels of mRNA transcription and protein expression of IL-6 and TNF-αwere significantly up-regulated ( P<0.01 ) in the model group.Compared with the model group, the levels of mRNA transcription and the expression of IL-6 and TNF-αproteins were significantly down-regulated in the positive group and Chinese medicine groups ( P <0.01 ) .③ Histological examination showed that the low-dose TCM significantly improved the CIA synovial hyperplasia and inflammatory cell infiltration.Conclusions The molecular mechanism of Chinese herbal compound Tengmei decotion in improving joint pathological injury of CIA rat models may be related to its inhibitory effect on the high expression of immune-inflammatory factors in the synovial tissue of CIA rats.

Objective: To investigate the intervention effects of Wenyang Huoxue Prescription(WYHXF) on 5/6 nephrectomized rats.Methods: One hundred and twenty SD rats were used to establish the 5/6 nephrectomized models,and then randomized into 5 groups: sham-operation group,model group,WYHXF group,Losartan group and combination(Losartan combined WYHXF) group.After the treatment,eight rats in each group were sacrificed at 4th,8th and 12th weeks,and the indexes of renal failure were detected.Results: Renal pathology showed glomerulosclerosis,tubular atrophy or disappearance,interstitial fibrosis in model group.Pathological changes in three treatment groups with different degrees of improvement,glomerulosclerosis was milder in WYHXF group and Losartan group than that in combination group.Compared with sham-operation group,the BUN,Scr,Cys-C,24-hour urinary protein and urinary N-acetyl-β-D-glucosaminidase in model group were significantly increased(P0.01,P0.05);the BUN,Scr,Cys-C,24-hour urinary protein and urinary N-acetyl-β-D-glucosaminidase in three treatment groups were lower than those in model group(P0.01).Conclusion: Wenyang Huoxue Prescription has protective effects on renal function by reducing the levels of BUN,Scr,Cys-C,24-hour urinary protein and urinary N-acetyl-β-D-glucosaminidase in rats with chronic renal failure.

Background: To investigate the renal microvascular injury in chronic aristolochic acid nephropathy (AAN) and the protective effects of Cozaar. Methods: Male Sprague-Dawley rats were randomized into three groups. Rats in the model group received Caulis Aristolochiae manshuriensis (CAM) decoction by gavage (10 mL/kg/day); those in the Cozaar group were gavaged with CAM and Cozaar (33.3 mg/kg/day); and those in the control group only received an equal daily volume of saline solution by gavage. Kidney tissues were observed under a light and electron microscope. CD34, caspase-3, and bone morphogenetic protein-7 (BMP-7) were determined by immunohistochemistry, and expressions of angiopoietin (Ang) 1 and 2, Tie-2, BMP-7, and vascular endothelial growth factor (VEGF) mRNA were monitored via real-time polymerase chain reaction (PCR). Results: (1) The kidney tissue injury in the chronic AAN model group was apparent, compared to the normal structure in the normal control group, and the Cozaar group showed relieved injury. (2) The expression of caspase-3 in the model group was elevated, while expressions of BMP-7 and CD34 were decreased (p < 0.05). Cozaar lessened caspase-3 expression (p < 0.05) and promoted BMP-7 and CD34 expressions (p < 0.05). (3) Real-time PCR demonstrated a downregulation of Ang-1, Tie-2, BMP-7, and VEGF mRNA (p < 0.05) and an upregulation of Ang-2 mRNA (p < 0.01) in the renocortex, while Cozaar upregulated the expression of Ang-1, Tie-2, BMP-7, and VEGF mRNA (p < 0.05). Conclusion: Renal microvascular injury was observed in chronic AAN, which was hypothetically correlated with a lack in the expressions of Ang-1, BMP-7, Tie-2, and VEGF and an excess in caspase-3 and Ang-2. Cozaar can significantly ameliorate the renal microvascular injury and protect renal function.

Objective: To observe the damage effects of aristolochic acid(AA) on proximal tubular epithelial cells(NRK-52E) of rats in vitro.Methods: NRK-52E were cultured in vitro with different concentrations of AA-Na stimulating these cells for 24 hours and 6 duplicated pores were set in each group.The ultrastructure of NRK-52E was observed by transmission electron microscope.The proliferation and apoptosis of NRK-52E were tested by MTT assay and flow cytometry respectively.The expression of α-smooth muscle actin(α-SMA) of NRK-52E was detected by immunocyto-chemical method.The expressions of transforming growth factor-β1(TGF-β1),endothelin-1(ET-1) mRNA and collagenⅠof NRK-52E were measured in different periods(12,24,48 h).Results: ① The AA-Na had no significant effect on proliferation of NRK-52E below the concentration of 10 μg/ml and the AA-Na inhibited the proliferation of NRK-52E obviously at the concentration of 20～40 μg/ml.② The AA-Na with high concentration(40 μg/ml) could stimulate the apoptosis of NRK-52E.③ The AA-Na could stimulate the expression of α-SMA of NRK-52E at the concentrations of 5,10,20 and 40 μg/ml especially at 10 μg/ml.④ The expressions of TGF-β1 mRNA and ET-1 mRNA of NRK-52E were highest after 24 hours stimulated by AA-Na and the expressions decreased after 48 hours,while the expression of collagenⅠincreased with time.Conclusion: AA-Na can directly damage renal tubular epithelial cells and promote the expression of α-SMA to promote transdifferentiation,and the damage effects of AA-Na on NRK-52E have dose and time dependent manner.

Objective:To investigate the effect of transdifferentiation for the aristolochic acid (Aristolochic Acid,AA) on rat proximal tubular epithelial cells (NRK-52E),and antagonistic role of Wenyanghuoxue for transdifferentiation.Methods:(1)To observe the proliferation of NRK-52E by MTT assay under the intention of different concentrations of Wenyanghuoxuefang containing serum.(2) To observe the expression of α-smooth muscle actin protein for NRK-52E under intention of different concentration AA (0,5,10,20,40 μg/ml) by immunocytochemical method.(3) to assay the expression of Col-Ⅰ and regulatory role of Wenyanghuoxuefang by ELISA assay.(4) to assay the expression of TGF-β1,VEGF,ET-1for NRK-52E under Wenyanghuoxuefang containing serum treatment.Results:(1) AA 5 μg/ml,AA 10 μg/ml,AA 20 μg/ml,AA 40 μg/ml can stimulate the NRK-52E expressing α-smooth muscle actin,the group of AA 10 μg/ml have the strongest role of stimulation.(2) After AA (10 μg/ml) stimulating cells at different time points(12、24、48 h),the expression of Col-Ⅰ increasedand showed time-depended trend.(3) After AA stimulating 24 hours,the expression of TGF-β1 mRNA was increased (P0.05),VEGF m RNA was significantly lower (P0.01),ET-1 mRNA was significantly raised (P0.01),Col-Ⅰ was significantly higher (P0.01),for the group of Wenyanghuoxuefang,the expression of TGF-β1 mRNA decreased (P0.05),the expression of VEGF mRNA was significantly raised (P0.01),ET-1 mRNA showed downward trend,the expression of Col-Ⅰ was significantly reduced.Conclusion:(1)Aristolochic acid can promote the transdifferentiation of tubular epithelial cells.(2)Wenyanghuoxuefang have antagonistic role in renal tubular epithelial cells.

近年来,有关中草药肾毒性的报道逐年增多,其中马兜铃酸肾病成为中草药不良反应的典型受到了广泛关注.该病起病隐匿,进展较快,预后不良,治疗尚无成熟方案.本文从临床与病理、发病机制及疾病的防治等方面时马兜铃酸肾病进行了论述.重点介绍了该病发生发展过程中的关键细胞因子和生长因子重要作用的研究。

AIM: To investigate the clinical effect of the treatment of the method of purging liver and the heart fire or nourishing yin and moistering arid, and that of both of them being used together in treating type 2 diabetes mellius, at the same time to evaluate differences of them of improving insulin resistance one by one. METHODS: 120 patients with type 2 diabetes were randomly divided into four groups with 30 cases each. They were treated with the method of purging liver and the heart fire (group 2)、with the methods of nourishing yin and moistering arid (group 3)、both of the two methods (group 1)、the drugs of rosiglitazone 4 mg a day (group 4), each for 1 month. The serum level of resistin、adiponectin、Leptin、TNF-α、CRP、IL-6 were measured of the four groups before and after the treatment by ELISA method, and blood pressure、blood sugar、lipid as well as insulin resistance levels were measured before and after treatment too. RESULTS: After treatment, Four groups could reduce blood sugar, no significant difference was observed between the four groups (P>0.05). group 1 and group 4 could significantly reduce the level of lipid、FINS, increase the level of IAI (P<0.01), but the group 2 and group 3 affection were not significant (P>0.05). After treatment, the clinical symptom score dropped in group 1、2、3 (P<0.01 or P< 0.05 ), no significant difference was observed in group 4, and it was the lowest in the group 1 in the four groups (P<0.01). At the same time, serum levels of CRP decreased in four groups. serum levels of Leptin、TNF-α、IL-6 decreased in group 1、2、4 (P<0.01 or P<0.05), no difference was observed in group 3 (P>0.05); serum levels of resistin decreased and adiponectin increased in group 1、4 (P<0.01 or P<0.05), no difference was observed in group 2、3 (P>0.05). More significant difference was observed in group 1、4 between the four groups about the serum level of resistin、adiponectin、Leptin、TNF-α、CRP、IL-6 (P<0.01). And between the group 2 and group 3, the serum level of CRP、Leptin、TNF-α、IL-6 in group 2 were reduced more than group 3, there was difference was observed between the two groups (P<0.01 or P<0.05). CONCLUSION: The method of purging liver and the heart fire, nourishing yin and moistering arid can improve insulin resistance and alleviate clinical symptoms of Type 2 diabetes mellitus patients, the mechanism may be related to its actions in regulating the production of inflammatory cytokines and inhibiting inflammatory reaction.