七氟醚对小鼠肺表面活性蛋白的影响

目的 探讨七氟醚对小鼠肺表面活性蛋白的影响。方法 将雄性昆明小鼠随机分为4组,吸入空气组、吸入1.0MAC七氟醚2h组、吸入2.0MAC七氟醚2h组、吸入3.0MAC七氟醚2h组;RT-PCR方法检测了吸入不同浓度七氟醚2h小鼠肺内表面活性蛋白A和B(SP-A、SP-B)mRNA的表达;免疫组化方法检测吸入不同浓度小鼠肺内SP-A、SP-B蛋白的表达。结果 吸入七氟醚2h能降低小鼠肺内SP-A和SP-B表达,而且随吸入浓度升高,SP-A、SP-B表达呈负相关。结论 长时间吸入高浓度七氟醚能够降低小鼠肺内SP-A和SP-B表达。     

小鼠E-W核神经元中磷酸化ERK1/2在异氟醚麻醉-苏醒过程中的表达变化

为了观察小鼠脑内E-W核神经元中磷酸化的ERK1/2(pERK1/2)在异氟醚吸入麻醉-苏醒过程中的表达变化,为探讨E-W核在麻醉效应产生机制中的作用提供形态学证据。我们将36只8周龄雄性BALB/c小鼠随机分为6组。第1组为清醒对照组(Con);第2、3组为异氟醚麻醉组(Iso-1、Iso-2:分别吸入1.0MAC异氟醚5min和1h);4～6组为异氟醚麻醉苏醒组(W-1、W-2:吸入1.0MAC异氟醚5min后停药2min、30min;W-3:吸入1.0MAC异氟醚1h后停药30min)。用免疫组织化学方法(ABC法)观察各时间点E-W核内pERK1/2阳性细胞并计数;用荧光双重标记法进一步明确pERK1/2阳性神经元的性质。结果显示:正常清醒小鼠E-W核内pERK1/2阳性细胞数量很少(2.2±1.5);异氟醚麻醉过程中pERK1/2表达显著增高(阳性细胞计数,Iso-1∶40.9±8.1;Iso-2∶40.2±9.6,与清醒对照相比,P<0.001);苏醒30min时pERK1/2表达降至正常对照组水平(W-2∶2.1±2.2;W-3∶0.75±1.2)。pERK1/2阳性神经元部分呈促肾上腺皮质激素释放激素(CRF)阳性,部分是乙酰胆碱(ChAT)阳性。上述结果提示,异氟醚麻醉过程中小鼠脑内E-W核神经元被激活,ERK1/2信号通路可能通过兴奋CRF能和Ach能神经元对瞳孔反射及麻醉应激效应进行调控。     

吡格列酮对异氟醚麻醉引起的老年小鼠认知功能损伤的影响*

 目的：研究高选择性的过氧化物酶体增殖物激活受体γ（PPARγ）激动剂吡格列酮（Pi）对异氟醚引起的老年小鼠认知功能障碍和脑内炎症细胞因子的影响。方法：136只11个月大的雄性C57BL/6J 小鼠随机分成5组：对照组（Con）、异氟醚组（Iso）、吡格列酮（10 mg/kg）+异氟醚组（Pi10+Iso）、吡格列酮（20 mg/kg）+异氟醚组（Pi20+Iso）和单纯吡格列酮（20 mg/kg）组（Pi20）。各异氟醚处理组的小鼠吸入混合1.4%异氟醚的氧气2 h;Con和Pi20组的小鼠仅吸入氧气2 h。吡格列酮溶于1%的羧甲基纤维素钠（CMC）,在小鼠吸入异氟醚或单纯氧气前2 h,按10 mg/kg或20 mg/kg灌胃,Con组和Iso组小鼠仅给予相同容量的1% CMC。在异氟醚处理结束后48 h行恐惧记忆实验以检测小鼠的学习记忆功能;6 h时取部分小鼠分离皮质和海马行Western blotting检测PPARγ蛋白及ELISA法检测脑组织IL-1β和TNF-α水平。结果：与Con组比较,Iso组的僵住行为减少（P<0.05）,海马IL-1β水平增加 （P<0.05）;与Iso组比较,Pi10+Iso组的僵住行为和PPARγ蛋白表达无明显变化（P>0.05）,而Pi20+Iso组的僵住行为和PPARγ蛋白表达均明显增加（P<0.05）且海马中IL-1β水平降低（P<0.05）。各组海马和皮质TNF-α水平以及皮质中IL-1β水平无明显差异（P>0.05）。结论：吡格列酮可以减轻异氟醚引起的老年小鼠的认知功能障碍,并可以缓解异氟醚引起的小鼠海马IL-1β含量的增加。     

安氟醚和异氟醚预处理对实验小鼠肝组织Bcl-2/Bax mRNA和蛋白表达的影响

目的:探讨吸入异氟醚和安氟醚预处理对实验小鼠肝组织Bcl-2和Bax基因和蛋白表达的影响。方法:120只C57BL/6小鼠随机分成安氟醚预处理组(E组)和异氟醚预处理组(I组),每组60只;各组又分为麻醉前(T0)、麻醉即刻(T1)、麻醉2h(T2)、麻醉4h(T3)、麻醉6h(T4)和麻醉终止后2h(T5)。取两组各时相肝组织检测Bcl-2和Bax mRNA及蛋白表达水平。结果:两组Bcl-2 mRNA及Bcl-2和Bax蛋白水平在T1表达均有所下调,但差异无统计学意义(P>0.05);在T2～T5表达上调(P<0.05或P<0.01),并随麻醉时间延长而逐渐增加,即T4时上调最明显(P<0.01);麻醉停止后(T5)开始恢复。两组Blc-2蛋白水平在T2～T5均较T0时增加,而Bax蛋白在T3～T5时增加(P<0.05或P<0.01);两组Bcl-2/Bax蛋白比值与T0时相比,E组在T1～T3、I组在T1～T5均明显升高(P<0.05)。结论:吸入安氟醚、异氟醚后肝组织Bcl-2和Bax表达增加,以Bcl-2更为明显;安氟醚、异氟醚预处理保护肝脏的作用可能与其抑制肝细胞凋亡有关。     

异氟醚麻醉对脑内ERK1/2和PKCγ磷酸化水平的影响

目的:探讨异氟醚麻醉对脑内ERK1/2和PKCγ磷酸化水平的影响.方法:采用雄性BALB/c小鼠, 随机分为5组, 即对照组(Con):未麻醉;异氟醚麻醉5 min组(Iso-1);异氟醚麻醉1 h组(Iso-2);麻醉1 h后停药2 min组(E-1):异氟醚麻醉1 h后, 小鼠脱离麻醉环境2 min;麻醉1 h后停药1 h组(E-2):异氟醚麻醉1 h后, 小鼠脱离麻醉环境1 h.用异氟醚进行麻醉后进行Western blot.以β-Actin为内参, pERK A/Actin A为ERK表达水平指标, pPKCγ A/Actin A为pPKCγ表达水平指标.结果:在Con组, pERK1/2和pPKCγ呈现高表达状态, 给予异氟醚麻醉处理组(Iso-1和Iso-2组), pERK1/2和pPKCγ表达减弱(P＜0.05), 当异氟醚麻醉停止后(E-1和E-2组), pERK1/2和pPKCγ表达逐渐恢复, 与Con组相比无统计学差异, 与异氟醚处理组相比, 有统计学差异(P＜0.05).结论:异氟醚麻醉-苏醒过程中, 脑中部分核团pERK1/2和pPKCγ水平发生显著变化.     

异氟醚和七氟醚对新生大鼠皮质凋亡以及JNK和p38表达的不同影响

目的: 研究同等剂量的异氟醚和七氟醚对新生大鼠皮质神经元凋亡的影响以及对JNK和p38蛋白表达的不同影响。方法: 55只出生后7 d(P7)的新生大鼠(共5窝,每窝取11只),随机均分为异氟醚组(Ⅰ组)、七氟醚组(S组)和对照组(C组),各组分别吸入1.1%异氟醚、1.8%七氟醚和空气4 h。在麻醉结束后2 h每窝每组各取1只幼鼠灌注取脑,免疫组织化学法检测皮质压部后区caspase-3表达(n=5);另外,每窝C组在麻醉处理0 h,Ⅰ组和S组分别在麻醉2 h、4 h取新鲜脑皮质,Western blotting检测磷酸化SAPK/JNK、磷酸化p38,以及SAPK/JNK、p38表达的变化(n=5)。结果: Ⅰ组和S组caspase-3表达分别较对照组增加441%(P<0.01)和151%(P<0.01),Ⅰ组比S组增加115%(P<0.05);Ⅰ组磷酸化SAPK/JNK表达在麻醉2 h和4 h较对照组分别增加219%(P<0.05)和181%(P<0.05),S组在2 h和4 h均与对照组无显著差异;Ⅰ组磷酸化p38表达在麻醉2 h和4 h较对照组分别增加38.9%(P<0.05)和36.9%(P<0.05),S组在2 h和4 h较对照组分别增加32.6%(P<0.05)和128.0%(P<0.01)。结论: 0.5最低肺泡有效浓度(MAC)异氟醚比七氟醚诱导更多新生大鼠大脑皮质神经元凋亡,异氟醚诱导凋亡可能与激活SAPK/JNK磷酸化有关,而七氟醚诱导凋亡可能与激活p38磷酸化有关。     

不同浓度七氟醚对大鼠海马去甲肾上腺素、5-羟色胺水平 及P2X3表达的影响

目的：探讨不同浓度七氟醚对大鼠海马去甲肾上腺素（NE）、5- 羟色胺（5-HT）水平及嘌呤能受体P2X3 表达的影响。方法：选取健康大鼠,分为对照组（ 不吸入七氟醚气体）,低、中、高七氟醚组。干预后,对大鼠 记忆行为进行周期性测定;采用高效液相检测NE、5-HT 含量;免疫印迹及免疫荧光检测P2X3 表达;用H-E 染 色观察海马神经元形态;用TUNEL 法检测海马神经元凋亡。结果：对照组大鼠在苏醒1 d 逃避潜伏期明显低于中、 低七氟醚组;与对照组大鼠比较,七氟醚3 组NE水平及5-HT 水平明显降低;对照组P2X3 蛋白表达显著高于七 氟醚高、中、低组,与低七氟醚组比较,中、高七氟醚组P2X3 蛋白表达逐渐降低;对照组、七氟醚低、中、高 组大鼠海马神经元细胞凋亡率分别为36.22%±3.62%、24.10%±2.66%、18.42%±2.21% 及12.55%±1.69%,差 异具有统计学意义。结论：七氟醚随着其浓度的增加会降低海马NE、5-HT 水平,增强对P2X3 表达的抑制作用, 并增加海马区细胞凋亡,降低大鼠记忆行为能力。     

基于NR2B蛋白表达探究异氟醚对妊娠大鼠疼痛水平及幼鼠神经细胞活性的作用机制

目的 基于NR2B蛋白表达探究异氟醚对妊娠大鼠疼痛水平及幼鼠神经细胞活性的作用机制。方法 选取40只SPF级SD孕15天雌性大鼠，随机分为正常（N）组，低异氟醚（LI）组，高异氟醚（HI）组，七氟醚（S）组，每组10只，对LI、HI组分别给予不同体积分数异氟醚并空氧混合气体吸入，对S组给予七氟醚并空氧混合气体吸入，N组同期给予同体积空氧混合气体吸入，测痛仪检测大鼠疼痛阈值，水迷宫法检测幼鼠认识功能，高尔基体染色法检测幼鼠神经细胞突触棘数量，TUNEL法检测幼鼠神经细胞凋亡，免疫组化法检测NR2B蛋白表达；另取培养后的神经细胞进行体外实验，将其分为两组，一组加入0.3 mmol/L的乳化异氟醚1 mL（观察组）；另一组加入1 mL 0.3 mmol/L的NR2B激动剂（对照组）。结果 与N组比较，LI组、HI组、S组0、6、12、18、24 h的体表疼痛阈值明显升高（P<0.05），幼鼠逃逸潜伏期、神经细胞凋亡、NR2B蛋白表达明显升高（P<0.05），穿越平台次数及平台象限停留时间、神经细胞突触棘数量明显降低（P<0.05），且HI组比LI组变化明显（P<0.0...     

全麻气管插管后七氟醚和异氟醚吸入对吸烟患者呼吸力学的影响

目的 比较观察全麻气管插管后七氟醚和异氟醚吸入对吸烟和非吸烟患者气道阻力、肺顺应性和气道峰压的影响.方法 选择既往有和无吸烟史择期手术的普通外科患者80例[美国麻醉医师协会(ASA)Ⅰ～Ⅱ级,既往有或无吸烟史患者各40例],随机分为4组(n=20):有吸烟史患者吸入七氟醚全麻组(SS组)和吸入异氟醚全麻组(SI组),无吸烟史患者吸入七氟醚全麻组(NS组)和吸入异氟醚全麻组(NI组).使用多功能麻醉气体监护仪监测患者吸入麻醉剂浓度达到肺泡最低有效浓度(1MAC)后4、8、12、16min的气道峰压、肺顺应性,同时用无创心功能测定仪监测气道阻力,记录各组患者在吸入麻醉剂期间各项指标的变化情况.结果 与吸入前相比,所有接受全麻气管插管的患者在使用七氟醚和异氟醚吸入维持4、8、12、16 min后均出现气道阻力和气道峰压的明显下降(均P＜0.05),其中SS组和NS组8min后下降趋于稳定[气道阻力:SS组(10.38±1.12)cmH2O·L-1·s-1,NS组(9.65±1.04)cm H2O·L-1·s-1;气道峰压:SS组(13.52±1.01)cm H2O,NS组(12.86±0.94)cm H2O,1 cm H2O=0.098kPa],SI组和NI组则于12 min后下降趋于稳定[气道阻力:SI组(10.30±0.98)cm H2O·L-1·s-1,NI组(11.00±0.73)cm H2O·L-1·s-1;气道峰压:SI组(13.47±0.88)cm H2O,NI组(12.85±0.65)cm H2O],同时间点的非吸烟组下降幅度高于吸烟组(均P＜0.05).4组患者在使用七氟醚和异氟醚吸入维持后其肺顺应性较吸入前均无明显变化(均P＞0.05),同时间点的非吸烟组与吸烟组相比肺顺应性差异也没有统计学意义(均P＞0.05).结论 全麻气管插管后七氟醚和异氟醚吸入使患者的气道阻力和气道峰压出现明显下降,吸烟者比非吸烟者下降程度低.     

七氟醚处理对老龄小鼠脑内神经生长因子表达的影响

目的:探讨七氟醚对老年小鼠脑内神经生长因子(NGF)表达的影响。方法:老年C57小鼠吸入七氟醚建立全身麻醉动物模型,利用ELISA检测基底前脑、额叶、颞叶及顶叶NGF的含量,real time RT-PCR检测额叶、颞叶及顶叶NGF mRNA水平,Western Blot检测额叶皮层NGF合成代谢通路中相关蛋白纤溶酶、基质金属蛋白酶-9(MMP-9)、基质金属蛋白酶组织抑制因子-1(TIMP-1)、丝氨酸或半胱氨酸蛋白水解酶抑制蛋白1(neuroserpin)的表达量。结果:与对照组相比,七氟醚组小鼠基底前脑、额叶、颞叶及顶叶中NGF蛋白含量均显著降低(P 0. 05),但七氟醚组小鼠额叶、颞叶及顶叶中NGF mRNA水平无显著变化。进一步研究发现,七氟醚组小鼠额叶皮层NGF前体蛋白含量增加(P 0. 05),参与NGF翻译后加工的纤溶酶表达显著降低(P 0. 05),neuroserpin表达显著增加(P 0. 05),而参与NGF降解的MMP-9与TIMP-1含量无显著变化。结论:七氟醚可通过抑制NGF翻译后加工过程而降低老年小鼠脑内神经生长因子的含量。     

Norepinephrine and vasopressin counteract anti-inflammatory effects of isoflurane in endotoxemic rats

Volatile anesthetics such as isoflurane have been shown to offer anti-inflammatory effects during experimental endotoxemia whereas the alpha-adrenergic vasopressor norepinephrine exhibits proinflammatory properties on systemic cytokine release under the same conditions. However, during major surgery and in patients with systemic inflammatory response syndrome or sepsis both agents are frequently administered concurrently. We therefore aimed to investigate the influence of preexisting i.v. administration of noradrenaline or vasopressin on the anti-inflammatory effects of isoflurane during experimental endotoxemia. Anesthetized, ventilated Sprague-Dawley rats (n=7 per group) were randomly treated. In the LPS-only group, animals received lipopolysaccharide (LPS, 5 mg/kg, i.v.) with no further specific treatment. In the LPS-isoflurane group, isoflurane inhalation at 1 MAC was initiated simultaneously with induction of endotoxemia (LPS 5 mg/kg, i.v.). Animals in the LPS-isoflurane-norepinephrine group received norepinephrine infusion at 50 microg/kg/h 10 min prior to injection of LPS and inhalation of isoflurane. In the LPS-isoflurane-vasopressin group, vasopressin was administered at 0.5 IE/kg/h 10 min prior to LPS and isoflurane. In the LPS-norepinephrine and the LPS-vasopressin groups the infusion of each vasopressor was started prior to LPS injection without any application of isoflurane. A Sham group served as the control. After 4 h of endotoxemia, plasma levels of TNFalpha, IL-1beta and IL-10 were measured. Alveolar macrophages (AM) were cultured ex vivo for nitrite assay. Induction of endotoxemia resulted in a significant rise in measured plasma cytokines and nitrite production from cultured AM. Inhalation of isoflurane significantly attenuated plasma levels of TNFalpha (-65%) and IL-1beta (-53%) compared to the LPS-only group whereas it had no effect on nitrite production from cultured AM. Preexisting infusions of norepinephrine or vasopressin abolished the anti-inflammatory effects of isoflurane. The data demonstrate that the administration of norepinephrine or vasopressin both counteracted the anti-inflammatory effects of inhaled isoflurane on proinflammatory cytokine release during experimental endotoxemia in rats.     

Genetic reduction of GABAA receptor γ2 subunit expression potentiates the immobilizing action of isoflurane

Potentiation of inhibitory γ-aminobutyric acid subtype A (GABA_A) receptor function is involved in the mechanisms of anesthetic action. The present study examined the immobilizing action of the volatile anesthetic isoflurane in mice with double knockout (DKO) of phospholipase C-related inactive protein (PRIP)-1 and -2. Both of these proteins play important roles in the expression of GABA_A receptors containing the γ2 subunit on the neuronal cell surface. Immunohistochemistry for GABA_A receptor subunits demonstrated reduced expression of γ2 subunits in the spinal cord of the DKO mice. Immunohistochemistry also revealed up-regulation of the α1 and β3 subunits even though there were no apparent differences in the immunoreactivities for the β2 subunits between wild-type and DKO mice. The tail-clamp method was used to evaluate the anesthetic/immobilizing effect of isoflurane and the minimum alveolar concentration (MAC) was significantly lower in DKO mice compared with wild-type controls (1.07 ± 0.01% versus 1.36 ± 0.04% atm), indicating an increased sensitivity to isoflurane in DKO mice. These immunohistochemical and pharmacological findings suggest that reduced expression of the GABA_A receptor γ2 subunit affects the composition and function of spinal GABA_A receptors and potentiates the immobilizing action of isoflurane.     

Surfactant protein A enhances Mycobacterium avium ingestion but not killing by rat macrophages

Mycobacterium avium complex (MAC) is a significant cause of opportunistic infection in patients with acquired immunodeficiency syndrome. Although the major route of entry of MAC is via the gastrointestinal tract, MAC can infect humans through the respiratory tract and eventually encounter alveolar macrophages within the lung. Once in the lung, MAC can potentially interact with surfactant protein A (SP-A), an important component of the pulmonary innate-immune response. Previous work on other pulmonary pathogens including Mycobacterium bovis Bacillus Calmette-Guerin (BCG) suggests that SP-A participates in promoting efficient clearance of these organisms by alveolar macrophages. In the present study, we investigated the role of SP-A in clearance of MAC by cultured rat macrophages. SP-A bound to MAC organisms and enhanced the ingestion of the mycobacteria by macrophages. Infection of macrophages with SP-A-MAC complexes induced the production of nitric oxide (NO) and tumor necrosis factor-alpha. However, intracellular survival of MAC was not altered by preopsonization with SP-A. In addition, inhibitors of inducible NO synthase did not alter MAC clearance. These results suggest that SP-A can bind to and enhance the uptake of MAC by alveolar macrophages, similar to previous findings with BCG and Mycobacterium tuberculosis.However, unlike BCG and other pulmonary pathogens that are cleared effectively in the presence of SP-A via a NO-dependent pathway, macrophage-mediated clearance of MAC is not enhanced by SP-A.     

Role of surfactant protein-A in nitric oxide production and mycoplasma killing in congenic C57BL/6 mice

We generated congenic surfactant protein A (SP-A)-deficient (SP-A[-/-]) mice on the mycoplasma resistant C57BL/6 background (B6.SP-A[-/-]) and characterized their response to mycoplasma infection in comparison to C57BL/6 (B6) mice. B6.SP-A(-/-) mice infected with 10(6) colony-forming units (cfu) of Mycoplasma pulmonis had significantly higher bacterial lung loads than B6 mice at 72 h postinfection (p.i.). At the higher infection dose of 10(7), B6.SP-A(-/-) mice had significantly higher lung cfu at 24 h; however, no difference in mycoplasma cfu was observed between B6 and B6.SP-A(-/-) mice at 48 and 72 h p.i. We found that uninfected B6 mice had lower bronchoalveolar lavage nitrite (NO(2)(-)) and nitrate (NO(3)(-)) levels as compared with B6.SP-A(-/-) mice. On the other hand, infection of B6 mice with mycoplasmas resulted in significantly higher bronchoalveolar lavage NO(2)(-) and NO(3)(-) as compared with B6.SP-A(-/-) mice. These data indicate that SP-A may help regulate NO production in response to a specific stimulus, i.e., suppression of NO in the absence of bacteria and increased NO in the presence of bacteria. These data indicate that the contribution of SP-A to mycoplasma killing may be limited to lower doses of pathogens.