Suppressing high mobility group box-1 release alleviates morphine tolerance via the adenosine5'-monophosphate-activated protein kinase/heme oxygenase-1 pathway

Opioids,such as morphine,are the most potent drugs used to treat pain.Long-term use results in high tolerance to morphine.High mobility group box-1(HMGB1) has been shown to participate in neuropathic or inflammatory pain,but its role in morphine tolerance is unclear.In this study,we established rat and mouse models of morphine tolerance by intrathecal injection of morphine for 7 consecutive days.We found that morphine induced rat spinal cord neurons to release a large amount of HMGB1.HMGB1 regulated nuclear factor κB p65 phosphorylation and interleukin-1β production by increasing Toll-like receptor 4receptor expression in microglia,thereby inducing morphine tolerance.Glycyrrhizin,an HMGB1 inhibito r,markedly attenuated chronic morphine tole rance in the mouse model.Finally,compound C(adenosine 5’-monophosphate-activated protein kinase inhibitor) and zinc protoporphyrin(heme oxygenase-1 inhibitor)alleviated the morphine-induced release of HMGB1 and reduced nuclear factor κB p65 phosphorylation and interleukin-1β production in a mouse model of morphine tolerance and an SH-SY5Y cell model of morphine tole rance,and alleviated morphine tolerance in the mouse model.These findings suggest that morphine induces HMGB1 release via the adenosine 5’-monophosphate-activated protein kinase/heme oxygenase-1 signaling pathway,and that inhibiting this signaling pathway can effectively reduce morphine tole rance.

CBP基因沉默对大鼠颈动脉球囊损伤后内膜增生的影响

目的观察小干扰RNA(siRNA)重组慢病毒介导的CREB结合蛋白(CBP)基因沉默对大鼠颈动脉球囊损伤后新生内膜增生的影响,并探讨其作用机制。方法雄性SD大鼠48只,随机分为4组:假手术组、PBS对照组、慢病毒介导CBP基因siRNA转染组(CBP-siRNA-Lenti组)及慢病毒介导非CBP同源序列siRNA转染组(NC-siRNA-Lenti组),建立大鼠颈动脉球囊损伤模型,术后28天处死动物。分别用实时定量PCR、Western Blot检测大鼠颈动脉CBP和乙酰化核因子κB p65(NF-κB p65)的表达水平;病理组织学观察血管内膜增生情况;免疫组织化学染色对损伤血管壁增殖细胞核抗原(PCNA)的表达进行评估。结果术后28天,与PBS对照组和NC-siRNA-Lenti组比较,CBP-siRNA-Lenti组CBP mRNA和蛋白的表达显著下调(P均<0.05),CBP沉默能明显抑制新生内膜面积(0.108±0.008 mm2比0.238±0.022 mm2、0.252±0.016 mm2,P<0.05)、内膜与中膜面积比(0.706±0.062比1.483±0.136、1.497±0.137,P<0.05)的增加,及下调血管壁乙酰化NF-κB p65和PCNA的表达水平(P均<0.05)。结论慢病毒介导的CBP基因沉默能有效地抑制颈动脉球囊损伤后新生内膜的形成,其机制可能与抑制NF-κB p65的过度乙酰化有关。

Losartan reduced connexin43 expression in left ventricular myocardium of spontaneously hypertensive rats

Objective:To assess the effect of angiotensin II type 1(AT1)receptor antagonist losartan on myocardium con- nexin43(Cx43)gap junction(GJ)expression in spontaneously hypertensive rats(SHRs)and investigate possible mechanisms. Methods:Sixteen 9-week-old male SHRs and 8 age-matched male Wistar-Kyoto(WKY)rats were included in this study.SHRs were randomly divided into two groups to receive losartan at 30 mg/(kg·d)by oral gavage once daily for 8 weeks(SHR-L)or vehicle(0.9%saline)to act as controls(SHR-V);WKY rats receiving vehicle for 8 weeks served as normotensive controls.At the end of the experiment,rats were sacrificed and the hearts were removed.Expressions of Cx43 and nuclear factor-kappaB p65 (NF-κB p65)proteins in all three groups were observed and further investigations on the effect of angiotensin II type 1 receptor antagonist losartan(30 mg/(kg·d),8 weeks)on Cx43 expression were conducted with Western blot and immunohistochemistry. NF-κB p65 protein in nuclear extracts was determined by Western blot.Results:Left ventricular(LV)hypertrophy was prominent in SHRs,Cx43 and NF-κB p65 protein expressions were obviously upregulated and Cx43 distribution was dispersed over the cell surface.Treatment with losarton reduced the over-expressions of Cx43 and NF-κB p65 in LV myocardium.The distribution of Cx43 gap junction also became much regular and confined to intercalated disk after losartan treatment.Conclu- sion:Cx43 level was upregulated in LV myocardium of SHR during early stage of hypertrophy.Angiotensin II type 1 receptor antagonist losartan prevented Cx43 gap junction remodeling in hypertrophied left ventricles,possibly through the NF-κB pathway.

Protective effect of Jiaweibugan decoction against diabetic peripheral neuropathy

Oxygen free radical damage is regarded as a direct or indirect common pathway associated with diabetic neuropathy and is the main cause of complications in peripheral neuropathies. We speculate that Jiaweibugan decoction has a significant effect in treating diabetic peripheral neuropathy through an anti-oxidative stress pathway. In this study, a diabetic rat model was established by intraperitoneal injection of streptozotocin. Rats were treated with Jiaweibugan decoction via intragastric administration. The levels of malondialdehyde and glutathione, which are indirect indexes of oxidative stress, in serum were determined using a colorimetric method. The expression levels of nuclear factor kappa B p65 mRNA and p38 mitogen-activated protein kinase, which are oxidative stress associated factors, in the dorsal root ganglion of spinal $4-6 segments were evaluated by reverse-transcriptase polymerase chain reaction and immunohistochemistry. Results showed that, Jiaweibugan decoction significantly ameliorated motor nerve conduction velocity in diabetic rats, effectively decreased malondialdehyde levels in serum and the expression of nuclear factor kappa B p65 mRNA and p38 mitogen-activated protein kinase mRNA in the dorsa root ganglion, and increased glutathione levels in serum. Therefore, our experimental findings indicate that Jiaweibugan decoction plays an anti-oxidative stress role in the diabetic peripheral neuropathy process, which has a protective effect on peripheral nerve injury.

细菌脂多糖联合高体积分数氧对未成熟大鼠脑发育的影响

目的探讨细菌脂多糖（LPS）、高体积分数氧（高氧）单独或联合应用对新生大鼠脑发育的影响及其相关机制。方法2日龄（P2）新生sD大鼠120只随机分为4组：空气组、LPS组、高氧组、LPS＋高氧组，分别观察各组大鼠的一般情况，记录其每日体质量。7日龄（P7）时，免疫组织化学法检测各组大鼠脑组织中半胱氨酸天冬氨酸蛋白酶3（Caspase-3）、核转录因子P65（NF—KBP65）的表达情况，ELISA法检测各组大鼠脑组织中IL-6、8-异．前列腺素F2a（8-iso．PGF20L）水平；12日龄（P12）时，免疫组织化学法检测各组大鼠脑组织碱性髓鞘蛋白（MBP）的表达。结果不同处理组中Caspase一3及NF—KBP65表达水平高低依次为LPS＋高氧组、LPS组／高氧组、空气组，前3组与空气组相比差异均有统计学意义（P均〈0．05），且LPS＋高氧组与高氧组、LPS组相比差异亦均有统计学意义（P均〈0．01）；MBP的表达水平高低依次为空气组、高氧组、LPS组、LPS＋高氧组，后3组与空气组相比差异均有统计学意义（P均〈0，05），且LPS＋高氧组与高氧组、LPS组相比差异亦均有统计学意义（P均〈0．01）。各组新生大鼠IL．6表达水平高低依次为LPS＋高氧组、LPS组、高氧组、空气组；8-iso-PGF20L表达水平高低依次为LPS＋高氧组、高氧组、LPS组、空气组，而且各组之间比较差异均有统计学意义（P均〈0．05）。结论感染及高氧均可致新生大鼠脑损伤，导致神经细胞凋亡和MBP表达减少，且感染与高氧同时存在时能加重二者单独作用时脑损伤的程度。其机制可能为炎性反应及氧化应激通过Toll样受体后发生协同作用，激活核转录因子NF-κB P65，并通过Caspase-3介导神经细胞凋亡及脑白质损伤。

人源性脂肪干细胞对30%体表总面积Ⅲ度烫伤大鼠肺组织炎症反应的影响及其机制初步探讨

目的研究人源性脂肪干细胞(h-ADSCs)对30%体表总面积(TBSA)Ⅲ度烫伤大鼠肺组织炎症反应的作用及机制。方法采用随机数字表法将72只雄性Sprague-Dawley大鼠分为单纯烫伤组和h-ADSCs组,每组各36只。采用沸水水浴法制备30%TBSAⅢ度烫伤模型,烫伤后即刻,h-ADSCs组予尾静脉注射含600万h-ADSCs(总注射量约1 mL),单纯烫伤组予尾静脉注射等体积的等渗NaCl溶液。比较两组大鼠烫伤8 h、1周、2周后肺组织肿瘤坏死因子α(TNF-α)、白细胞介素6(IL-6)、α7烟碱型乙酰胆碱受体(α7nAChR)信使RNA(mRNA)及其蛋白、核因子κB(NF-κB)P65蛋白、苏木素-伊红(HE)染色评分及肺损伤评分,并在普通光镜下观察肺组织病理变化情况。结果h-ADSCs组大鼠烫伤8 h、1周、2周后肺组织TNF-α(t=2.421、2.626、2.356,P=0.035、0.041、0.032)、IL-6(t=2.514、2.751、2.756,P=0.029、0.031、0.039)、HE染色评分(t=2.869、2.631、2.813,P=0.028、0.031、0.042)和肺损伤评分(t=2.172、3.017、2.698,P=0.033、0.032、0.029)和烫伤1周、2周后NF-κB P65蛋白(t=2.639、2.491,P=0.028、0.037)均明显低于单纯烫伤组同时间点;而烫伤1周、2周后α7nAChR mRNA(t=2.912、2.663,P=0.032、0.046)及其蛋白(t=2.814、2.638,P=0.033、0.032)表达水平均明显高于单纯烫伤组同时间点。伤后8 h,h-ADSC组大鼠肺泡隔增宽,间质血管充血,病变轻于单纯烫伤组。伤后1周,h-ADSCs组大鼠镜下可见肺间隔明显增厚,肺泡腔变窄,伴有炎症细胞浸润。伤后2周,h-ADSCs组大鼠镜下可见肺间隔明显增厚,肺泡腔变窄,伴有大量炎症细胞浸润。结论h-ADSCs能减轻30%TBSAⅢ度烫伤休克大鼠肺组织炎症水平,其机制可能与h-ADSCs能增加α7nAChR的表达和抑制肺组织NF-κB P65的表达有关。