miR-499通过Drp1介导线粒体自噬保护缺氧/复氧心肌细胞

目的探究miR-499对心肌缺血/再灌注损伤的保护作用,并从线粒体自噬方面探究其可能机制。方法缺血/再灌注(I/R)诱导心肌细胞H9c2(2-1),建立缺氧/复氧(H/R)心肌细胞模型,使用miR-499 mimics和(或)P110处理细胞。将细胞分为BC组、I/R组、I/R+mi组、I/R+NC组、I/R+mi+P110组、I/R+NC+P110组。使用CCK8法检测细胞增殖,流式细胞术检测ROS、线粒体膜电位和细胞凋亡,试剂盒检测MDA、SOD、ATP含量,qRT-PCR和Western blot检测线粒体融合、分裂、自噬相关基因Fis1、Mfn1、Parkin、LC3-Ⅱ、p62和Drp1的mRNA和蛋白表达水平,透射电镜观察线粒体自噬。结果miR-499 mimics和/或P110可使I/R诱导的心肌细胞增殖抑制率和凋亡率下降;线粒体膜电位下降、线粒体自噬减少、ATP含量上升;Fis1、Parkin、LC3-Ⅱ和Drp1的基因和蛋白表达水平下降,Mfn1和p62的基因和蛋白表达水平上升;细胞内ROS和MDA含量减少,SOD含量增加。结论miR-499可通过降低Drp1介导的线粒体自噬减少氧化应激的发生,从而保护I/R诱导的心肌细胞。

黄芩茎叶总黄酮预处理对大鼠心肌缺血再灌注损伤的保护作用

目的探讨黄芩茎叶总黄酮(Scutellaria baiculensis stem-leaf total flavonoid,SSTF)对大鼠心肌缺血再灌注损伤的保护作用。方法40只SD大鼠随机分为假手术组、缺血再灌注组和SSTF预处理组。SSTF组大鼠于术前1周分别灌胃不同剂量的SSTF〔(17.5、35、70mg/(kg.d)〕,另两组给等量生理盐水。采用结扎大鼠左冠状动脉前降支方法制备心肌缺血再灌注模型。缺血30min、再灌注2h后,观察大鼠心电图的变化;并用免疫组织化学法检测心肌组织中Bax和Bcl-2蛋白表达情况。结果SSTF抑制了缺血再灌注损伤大鼠心电图T波升高,降低了心律失常发生率。缺血再灌注组Bcl-2蛋白表达阳性细胞率明显比假手术组低(P<0.05),Bax蛋白表达阳性细胞率明显较假手术组高,而SSTF预处理组Bcl-2蛋白表达阳性细胞率明显高于缺血再灌注组(P<0.05),Bax蛋白表达阳性细胞率明显较缺血再灌注组为低(P<0.05)。结论SSTF预处理对缺血再灌注的心肌细胞具有保护作用,其机制可能与上调Bcl-2和下调Bax蛋白表达进而抑制心肌细胞凋亡有关。

氢溴酸樟柳碱对急性脑缺血再灌注损伤模型大鼠脑组织细胞凋亡及ERK1/2磷酸化水平的影响

目的:研究氢溴酸樟柳碱对急性脑缺血再灌注损伤模型大鼠脑组织细胞凋亡及细胞外信号调节蛋白激酶1/2(ERK1/2)磷酸化(p-ERK1/2)水平的影响。方法:将大鼠随机分为假手术组、模型组、阳性对照组(尼莫地平1.0 mg/kg)和氢溴酸樟柳碱高、中、低、极低剂量组(1.2、0.6、0.3、0.15 mg/kg),每组8只,采用线栓法建立大鼠急性脑缺血再灌注损伤模型。分别于脑缺血2 h和再灌注6 h时对各组大鼠尾iv给药1次,再灌注22 h后检测各组大鼠脑组织三磷酸腺苷(ATP)酶活性、Ca^(2+)含量、细胞凋亡情况、脑组织中p-ERK1/2蛋白表达和p-ERK1/2/总ERK1/2(t-ERK1/2)比例。结果:与假手术组比较,模型组大鼠脑组织ATP酶活性明显降低、Ca^(2+)含量明显增加、凋亡细胞密度明显增加,以上差异均有统计学意义(P<0.01)。与模型组比较,各给药组大鼠脑组织凋亡细胞密度均明显减小,阳性对照组和氢溴酸樟柳碱高、低剂量组大鼠脑组织Ca^(2+)含量均明显降低,氢溴酸樟柳碱高、低、极低剂量组大鼠脑组织中p-ERK1/2/t-ERK1/2比例均明显增加,以上差异均有统计学意义(P<0.05或P<0.01);其余差异均无统计学意义(P>0.05)。结论:氢溴酸樟柳碱能抑制急性脑缺血再灌注损伤模型大鼠脑组织细胞凋亡,其作用机制可能与激活ERK1/2信号通路和调节ATP酶活性,进而降低脑组织Ca^(2+)含量有关。

SAFE通路介导的炎症因子在大鼠心肌缺血再灌注致脑损伤中的作用

目的探讨生存活化因子增强通路(SAFE)介导的炎症因子在大鼠心肌缺血再灌注致脑损伤中的作用。方法 24只雄性SD大鼠按随机数表法分为三组(n=8):假手术组(Sham)、心肌缺血/再灌注组(IR)、心肌缺血/再灌注+AG490组(IR+AG490)。IR组及IR+AG490组通过结扎大鼠冠状动脉左前降支30 min,再灌注120 min建立大鼠心肌缺血/再灌注模型。取大鼠脑组织,Western blot检测Janus激酶2(Jak2)、信号转导子和转录激活子3(STAT3)蛋白表达及炎症因子白介素1(IL-1),白介素6(IL-6),白介素8(IL-8),TUNEL法检测细胞凋亡,比色法检测含半胱氨酸的天冬氨酸蛋白水解酶3(Caspase3)活性。结果与Sham组比较,IR组p-Jak2[(1.8±0.13)vs(1.0±0)]、p-STAT3[(1.6±0.08)vs(1.0±0)]表达增高,炎症因子IL-1[(3.3±0.16)vs(1.0±0)]、IL-6[(3.3±0.16)vs(1.0±0)]、IL-8[(2.8±0.28)vs(1.0±0)]表达增多,TUNEL[(18.8±1.29)%vs(4.2±0.44)%]表达增多,Caspase3活性[(2.6±0.24)vs(1.0±0)]升高,差异均有统计学意义(P<0.05);与IR组比较,IR+AG490组p-Jak2[(1.3±0.09)vs(1.8±0.13)],pSTAT3[(1.1±0.11)vs(1.6±0.08)]表达减少,炎症因子IL-1[(2.1±0.17)vs(3.3±0.16)]、IL-6[(1.9±0.22)vs(3.3±0.16)]、IL-8[(2.2±0.19)vs(2.8±0.28)表达降低,TUNEL[(13.2±1.03)%vs(18.8±1.29)%]表达降低,Caspase3活性[(2.1±0.21)vs(2.6±0.24)]降低,差异均有统计学意义(P<0.05)。结论抑制SAFE通路的激活可以降低其下游炎症因子的级联表达,从而减少细胞凋亡,减轻大鼠心肌缺血再灌注引起的脑损伤。

Sulforaphane protects liver injury induced by intestinal ischemia reperfusion through Nrf2-ARE pathway

AIM: To investigate the effect of sulforaphane (SFN) on regulation of NF-E2-related factor-2 (Nrf2)-antiox-idant response element (ARE) pathway in liver injury induced by intestinal ischemia/reperfusion (I/R). METHODS: Rats were divided randomly into four ex-perimental groups: control, SFN control, intestinal I/R and SFN pretreatment groups (n = 8 in each group). The intestinal I/R model was established by clamping the superior mesenteric artery for 1 h and 2 h reperfu-sion. In the SFN pretreatment group, surgery was performed as in the intestinal I/R group, with intraperitoneal administration of 3 mg/kg SFN 1 h before the op-eration. Intestine and liver histology was investigated. Serum levels of aspartate aminotransferase (AST), and alanine aminotransferase (ALT) were measured. Liver tissue superoxide dismutase (SOD), myeloperoxidase (MPO), glutathione (GSH) and glutathione peroxidase (GSH-Px) activity were assayed. The liver transcription factor Nrf2 and heme oxygenase-1 (HO-1) were determined by immunohistochemical analysis and Western blotting analysis.RESULTS: Intestinal I/R induced intestinal and liver injury, characterized by histological changes as well as a signif icant increase in serum AST and ALT levels (AST: 260.13 ± 40.17 U/L vs 186.00 ± 24.21 U/L, P < 0.01; ALT: 139.63 ± 11.35 U/L vs 48.38 ± 10.73 U/L, P < 0.01), all of which were reduced by pretreatment with SFN, respectively (AST: 260.13 ± 40.17 U/L vs 216.63 ± 22.65 U/L, P < 0.05; ALT: 139.63 ± 11.35 U/L vs 97.63 ± 15.56 U/L, P < 0.01). The activity of SOD in the liver tissue decreased after intestinal I/R (P < 0.01), which was enhanced by SFN pretreatment (P < 0.05). In ad-dition, compared with the control group, SFN markedly reduced liver tissue MPO activity (P < 0.05) and elevat-ed liver tissue GSH and GSH-Px activity (P < 0.05, P < 0.05), which was in parallel with the increased level of liver Nrf2 and HO-1 expression.CONCLUSION: SFN pretreatment attenuates liver injury induced by intestinal I/R in rats, attributable to the antioxidant effect through Nrf2-ARE pathway.

巴曲酶对围产期缺氧缺血性脑损伤神经保护作用的研究

目的 制备宫内缺氧缺血性脑损伤 (HIBD)动物模型 ,应用巴曲酶研究其对 HIBD的神经保护作用。方法 宫内窒息 2 0 min为实验组 ,并设立正常对照组 ,实验组随机分为两组 ,1组于生后 3 0 min、2 4h、48h各给予腹腔注射巴曲酶 (8BU/ kg) 1次 ,共 3次 (巴曲酶干预组 ) ,另 1组 (缺氧缺血组 )及正常对照组于相同时间给予腹腔注射等量生理盐水。3组仔鼠生后 72 h、7d、14 d、2 1d断头处死 ,测量脑组织含水量及重量 ,并应用 TU NEL法观察凋亡细胞数。结果 缺氧缺血组 72 h脑含水量明显高于巴曲酶干预组及正常对照组 (P<0 .0 5) ,其余时间点 3组脑含水量无差异。缺氧缺血组 72 h脑重量高于巴曲酶干预组及正常对照组 (P<0 .0 5) ,14 d、2 1d脑重量显著低于巴曲酶干预组及正常对照组 (P<0 .0 5) ;巴曲酶干预组脑重量 14 d、2 1d低于正常对照组 ,差异显著 (P<0 .0 5)。缺氧缺血组72 h～ 14 d各时间点阳性细胞数显著高于巴曲酶干预组及正常对照组 (P<0 .0 5) ,2 1d 3组阳性细胞数无显著差异 ;巴曲酶干预组 72 h～ 7d凋亡细胞数高于正常对照组 (P<0 .0 5) ,14 d降至正常 ;正常对照组各时间点偶见阳性细胞。结论 巴曲酶可明显减轻 HIBD急性期脑水肿及恢复期脑萎缩 ,抑制 HIBD神经细胞凋亡 。

左旋卡尼汀对兔缺血/再灌注心肌细胞死亡及心脏功能的影响

目的 探讨左旋卡尼汀对心肌缺血 再灌注 (MI R)后心肌细胞死亡 (坏死和凋亡 )及心脏功能的影响。方法 制备家兔MI R模型 ,缺血 45min ,再灌注 3h。 2 5只家兔随机分为 3组 :对照组 (Ⅰ组 ,n =5) ,丝线穿过冠状动脉左室支但不结扎 ;MI R组 (Ⅱ组 ,n =10 ) ,MI后 5min静脉滴注生理盐水至实验结束 ;左旋卡尼汀治疗组 (Ⅲ组 ,n =10 ) ,在缺血后 5min静脉滴注左旋卡尼汀 (1.5ml·kg- 1·h- 1)加生理盐水至实验结束。观察心率、左室压等的变化 ,再灌注结束后检测心肌梗死或用原位末端TUNEL法检测细胞凋亡指数 ,心肌组织游离脂肪酸 (FFA)含量。结果 MI R造成明显的心脏功能障碍 ,心肌梗死和缺血区细胞凋亡 ,组织FFA含量增多。与Ⅱ组比较 ,Ⅲ组的梗死面积明显缩小 (P <0 .0 5) ,凋亡指数减少 (P <0 .0 5) ,FFA减少 (P <0 .0 5)及促进再灌注后心脏收缩舒张功能恢复。结论 左旋卡尼汀可减少心肌梗死范围 。

Coexistence of hyperlipidemia and acute cerebral ischemia/reperfusion induces severe liver damage in a rat model

AIM:To investigate the correlation of hyperlipemia(HL) and acute cerebral ischemia/reperfusion(I/R) injury on liver damage and its mechanism.METHODS:Rats were divided into 4 groups:control,HL,I/R and HL+I/R.After the induction of HL via a high-fat diet for 18 wk,middle cerebral artery occlusion was followed by 24 h of reperfusion to capture I/R.Serum alanine transaminase(ALT) and aspartate aminotransferase(AST) were analyzed as part of liver function tests and liver damage was further assessed by histological examination.Hepatocyte apoptosis was evaluated by terminal deoxynucleotidyl transferase dUTP nick-end labeling(TUNEL) assay.The expression of genes related to apoptosis(caspase-3,bcl-2) was assayed by immunohistochemistry and Western blotting.Serum tumor necrosis factor-(TNF-),interleukin-1(IL-1) and liver mitochondrial superoxide dismutase(SOD),glutathione peroxidase(GSH-Px),malondialdehyde(MDA) and Ca 2+ levels were measured to determine inflammatory and oxidative/antioxidative status respectively.Microsomal hydroxylase activity of the cytochrome P450 2E1(CYP2E1)-containing enzyme was measured with aniline as the substrate,and CYP2E1 expression in the liver tissue and microsome was determined by immunohistochemistry and Western blotting respectively.RESULTS:HL alone induced by high-fat diet for 18 wk resulted in liver damage,indicated by histopathological analysis,and a considerable increase in serum ALT(25.13 ± 16.90 vs 9.56 ± 1.99,P < 0.01) and AST levels(18.01 ± 10.00 vs 11.33 ± 4.17,P < 0.05) compared with control.Moreover,HL alone induced hepatocyte apoptosis,which was determined by increased TUNEL-positive cells(4.47 ± 0.45 vs 1.5 ± 0.22,P < 0.01),higher caspase-3 and lower bcl-2 expression.Interestingly,compared with those in control,HL or I/R groups,massive increases of serum ALT(93.62 ± 24.00 vs 9.56 ± 1.99,25.13 ± 16.90 or 12.93 ± 6.14,P < 0.01) and AST(82.32 ± 26.92 vs 11.33 ± 4.17,18.01 ± 10.00 or 14.00 ± 6.19,P < 0.01) levels in HL+I/R group were observed suggesting severe liver damage,which was confirmed by liver histology.In addition,HL combined with I/R also caused significantly increased hepatocyte apoptosis,as evidenced by increased TUNEL-positive cells(6.20 ± 0.29 vs 1.5 ± 0.22,4.47 ± 0.45 or 1.97 ± 0.47,P < 0.01),elevated expression of caspase-3 and lower expression of bcl-2.Furthermore,when compared to HL or I/R alone,HL plus I/R enhanced serum TNF-,IL-1,liver mitochondrial MDA and Ca 2+ levels,suppressed SOD and GSH-Px in liver mitochondria,and markedly up-regulated the activity(11.76 ± 2.36 vs 4.77 ± 2.31 or 3.11 ± 1.35,P < 0.01) and expression(3.24 ± 0.38 vs 1.98 ± 0.88 or 1.72 ± 0.58,P < 0.01) of CYP2E1 in liver.CONCLUSION:The coexistence of HL and acute cerebral I/R induces severe liver damage,suggesting that cerebral ischemic stroke would exaggerate the damage of liver caused by HL.This effect is possibly due to en-hanced CYP2E1 induction which further promotes oxidative damage,inflammation and hepatocyte apoptosis.

Diagnosis and management of splanchnic ischemia

Splanchnic or gastrointestinal ischemia is rare and randomized studies are absent. This review focuses on new developments in clinical presentation, diagnostic approaches, and treatments. Splanchnic ischemia can be caused by occlusions of arteries or veins and by physiological vasoconstriction during low-flow states. The prevalence of significant splanchnic arterial stenoses is high, but it remains mostly asymptomatic due to abundant collateral circulation. This is known as chronic splanchnic disease (CSD). Chronic splanchnic syndrome (CSS) occurs when ischemic symptoms develop. Ischemic symptoms are characterized by postprandial pain, fear of eating and weight loss. CSS is diagnosed by a test for actual ischemia. Recently, gastro-intestinal tonometry has been validated as a diagnostic test to detect splanchnic ischemia and to guide treatment. In single- vessel CSD, the complication rate is very low, but some patients have ischemic complaints, and can be treated successfully. In multi-vessel stenoses, the complication rate is considerable, while most have CSS and treatment should be strongly considered. CT and MR-based angiographic reconstruction techniques have emerged as alternatives for digital subtraction angiography for imaging of splanchnic vessels. Duplex ultrasound is still the first choice for screening purposes. The strengths and weaknesses of each modality will be discussed. CSS may be treated by minimally invasive endoscopic treatment of the celiac axis compression syndrome, endovascular antegrade stenting, or laparotomy-assisted retrograde endovascular recanalization and stenting.The treatment plan is highly individualized and is mainly based on precise vessel anatomy, body weight, co- morbidity and severity of ischemia.

全息影像在机器人辅助腹腔镜肾部分切除术中的优势

目的分析将全息影像技术应用于机器人辅助腹腔镜肾部分切除术导航中的有效性和安全性。方法回顾性分析了2019年11月-2021年1月由中国人民解放军中部战区总医院同一术者完成机器人辅助腹腔镜肾部分切除术的44例难治性肾肿瘤患者(RENAL评分≥7分)的临床资料,根据术中是否使用全息影像实时导航分为全身影像实时术中导航(HRLPN)组和非全身影像实时术中导航(RLPN)组。记录两组患者的术中及术后3个月的随访情况。结果两组患者手术均顺利完成,HRLPN组较RLPN组的手术时间[(110.8±8.18)min vs.(135.8±8.66)min]、血管分离时间[(62.85±4.61)min vs.(77.88±4.43)min]、热缺血时间[(26.13±0.95)min vs.(30.23±0.72)min]短,差异有统计学意义(P<0.05)。两组均未发生术中并发症,无术中转开放手术或根治性肾切除术,术后所有患者的病理切缘均为阴性。HRLPN组与RLPN组的肿瘤切除时间、术中失血量、术后3个月的血肌酐和预估肾小球滤过率差异均无统计学意义(P>0.05)。结论机器人辅助腹腔镜肾部分切除术应用全息影像实时术中导航可缩短手术时间、热缺血时间和血管分离时间,减少术中血管及正常肾组织的损伤。

丁咯地尔治疗椎-基底动脉缺血临床疗效评价

目的 :探讨丁咯地尔对椎基底动脉缺血的治疗作用。方法 :将 92例椎基底动脉缺血患者随机分为丁咯地尔组(4 6例)和盐酸培他啶组 (4 6例 ) ,逐日观察疗效。结果 :丁咯地尔组临床效果显著 ,用药治疗 72h后总有效率 96.6%。结论 :丁咯地尔治疗椎基底动脉缺血疗效好 ,起效快 ,具有临床应用价值。

Effect of electroacpuncture of Neiguan(PC 6)and Tianquan(PC 2)on skin temperature,blood perfusion,and adrenoceptor response in rats with acute myocardial ischemia

OBJECTIVE: To investigate the effect of electroacupuncture(EA) of acupoints Neiguan(PC 6) and Tianquan(PC 2) on the skin temperature, blood perfusion, and alpha1-and beta2-adrenoreceptor(α1-AR and β2-AR) protein and m RNA level in rats with acute myocardial ischemia.METHODS: Thirty male adult Wistar rats [(230 ±20) g] were randomly assigned into five groups(n = 6 each): a control group, sham operation group, model group, model group treated with EA at low frequency(L-EA, 2 Hz, 1 m A) and model group treated with EA at high frequency(H-EA, 100 Hz, 1 m A). The rat model was prepared by ligating the left anterior descending coronary artery. Electroacupuncture was performed at the left Neiguan(PC 6) for 20 min daily for 3 d. After the 3 rd time of the treatment, measurements of skin blood perfusion and temperature in Neiguan(PC 6) and Tianquan(PC 2) in all groups were made by laser speckle contrast imaging and infrared thermal image instrument. Real-time PCR and ELISA were used to measure m RNA level and protein level of α1-AR andβ2-AR in the skin tissues of Neiguan(PC 6) and Tianquan(PC 2), respectively. Serum levels of c Tn T and electrocardiogram were used to identify the state of myocardium.RESULTS: In the group receiving electroacupunture at Neiguan(PC 6), compared with control group,the skin temperature, blood perfusion, and β2-AR m RNA and protein level of model group significantly decreased(P < 0.05), α1-AR m RNA and protein level of model group significantly increased(P < 0.05);compared with model group, the skin temperature,blood perfusion, and β2-AR m RNA and protein level of L-EA significantly increased(P < 0.05), α1-AR m RNA and protein level of L-EA and H-EA significantly decreased(P < 0.05). The same trend has taken place in the former four groups of the Tianquan(PC 2).CONCLUSION: Low-or high-frequency electroacupuncture can improve the skin temperature and blood perfusion which may be induced by decreasingthelevelofα1-ARandincreasingthelevelofβ2-AR of the Neiguan(PC 6) and Tianquan(PC 2) in the rat with acute myocaidial ischemia.