地塞米松和维生素B12干预下小鼠胚胎腭突Shh相关信号因子的研究

目的：　探讨地塞米松及其拮抗剂维生素B₁₂干预妊娠关键期孕鼠后,小鼠胚胎腭突上Shh信号通路重要因子、细胞周期调控因子及初级纤毛解聚相关因子的调控及三者间的变化关系。方法：　予特定孕期C57BL/6J孕鼠腹腔注射一定剂量的生理盐水（空白对照组）、地塞米松（地塞米松组）、维生素B₁₂（维生素B₁₂组）、地塞米松+维生素B₁₂（地塞米松+维生素B₁₂组）,观察妊娠17.5 d和13.5 d各组小鼠胚胎腭突融合情况;Western blot检测妊娠13.5 d各组小鼠胚胎腭突中Smo、Ptch1、Cyclin D1、Aurora A的表达变化并进行统计学分析。结果：　地塞米松延缓胚胎腭突发育而导致腭裂发生,而维生素B₁₂干预后,可部分恢复胚胎腭突发育。Western blot结果显示,与对照组相比,Ptch 1、Aurora A、Cyclin D1的表达在地塞米松组中明显降低（均P=0.000）,Smo无明显改变（P=0.695）;地塞米松+维生素B₁₂组中Ptch 1、Aurora A、Cyclin D1的表达与地塞米松组相比,均不同程度上调（均P=0.000）,Smo无明显改变（P=0.818）;维生素B₁₂组中各目的蛋白表达与对照组无统计学差异（P=0.879、0.399、0.645、0.168）。结论：　地塞米松影响小鼠胚胎腭突Shh下游信号传递和初级纤毛解聚,从而抑制Cyclin D1表达使小鼠胚胎腭突细胞增殖降低,维生素B₁₂可通过上调相关因子蛋白含量拮抗这一抑制效果。     

SHH信号通路对子痫前期患者滋养细胞凋亡和侵袭的影响及其机制

目的：探讨激活SHH信号通路对子痫前期（PE）患者滋养细胞凋亡和侵袭的影响,并阐明其作用机制。方法：将HTR8/SVneo细胞分为正常对照组、缺氧/复氧（H/R）处理组和purmorphamine+H/R处理组。Western blotting法检测PE胎盘组织和正常妊娠晚期胎盘组织及各组HTR8/SVneo细胞中SHH信号通路相关蛋白SHH、Ptch1、Smo、Gli1和Gli3蛋白表达水平,流式细胞术（FCM）检测各组HTR8/SVneo细胞的凋亡率,Transwell小室法检测各组HTR8/SVneo细胞的穿膜细胞数,Western blotting法检测各组HTR8/SVneo细胞中基质金属蛋白酶2（MMP-2）和基质金属蛋白酶9（MMP-9）蛋白表达水平。结果：PE胎盘组织HTR8/SVneo细胞中SHH、Ptch1、Smo、Gli1和Gli3蛋白表达水平均明显低于正常妊娠晚期胎盘组织（P<0.05）;H/R处理组HTR8/SVneo细胞中SHH、Ptch1、Smo、Gli1、Gli3、MMP-2和MMP-9蛋白表达水平及穿膜细胞数均明显低于正常对照组,细胞凋亡率明显高于正常对照组（P<0.05）;purmorphamine+H/R处理组HTR8/SVneo细胞中SHH、Ptch1、Smo、Gli1、Gli3、MMP-2和MMP-9蛋白表达水平及穿膜细胞数均明显高于H/R处理组（P<0.05）,细胞凋亡率明显低于H/R处理组（P<0.05）。结论：激活SHH信号通路可减少PE患者胎盘组织中HTR8/SVneo细胞凋亡,并通过上调MMP-2和MMP-9表达提高细胞的侵袭能力。     

调控Smo活性对成年大鼠神经干细胞增殖和迁移的影响

目的 探讨Shh信号通路中G蛋白偶联受体Smoothened（Smo）对成年大鼠神经干细胞（ANSCs）增殖、迁移的影响并阐明其作用机制。方法 培养ANSCs,分别添加Smo的激动剂Purmorphamine及其抑制剂Cyclopamine,细胞增殖-毒性实验（CCK8）检测其对ANSCs增殖的影响;细胞划痕实验检测其对ANSCs迁移能力的影响;实时荧光定量PCR（RT-PCR）技术检测Sonic Hedgehog（Shh）信号通路的细胞表面Patched蛋白（Ptch1）、G蛋白偶联受体Smo、转录因子（Gli1）以及神经导向因子（Slit1）和脑源性神经营养因子（BDNF）表达量的变化。结果 Smo激动剂PM对ANSCs的增殖有明显促进作用,48 h后PM组ANSCs的吸光度明显升高（P<0.01）;PM可以促进ANSCs的迁移,48 h后的PM组ANSCs划痕面积显著减小（P<0.01）;Smo抑制剂CPM对ANSCs的增殖和迁移均有显著的抑制作用;RT-PCR实验显示PM组细胞中的Ptch1、Smo、Gli1及Slit1、BDNF表达水平均明显增加（P<0.05）。结论 调控Smo的活性可以促进或抑制成年神经干细胞的增殖和迁移,其作用机制可能与调节BDNF和Slit1的表达有关。     

干预Hedgehog信号对肾小管上皮细胞增殖细胞核抗原表达的影响

目的:探讨Hedgehog(HH)信号通路的活化和抑制对肾小管上皮细胞中增殖细胞核抗原(PCNA)表达的影响。方法:根据干预措施将体外培养的大鼠肾小管上皮细胞NRK-52E分为对照组、活化组(10μg/L Shh)和干预组(10μg/L Shh加5μmol/L环杷明)。细胞培养24 h后,细胞免疫荧光染色检测HH信号关键分子Ptch1、Smo和Gli1,以及细胞增殖标志物PCNA的表达,real-time RT-PCR检测Ptch1和Smo m RNA的表达。结果:免疫荧光染色显示,对照组细胞维持一定的增殖状态,HH信号活性较低,PCNA表达水平较低;当外源性重组蛋白Shh活化NRK-52E细胞24 h后,Smo和Gli1 m RNA和蛋白表达显著升高,Ptch1的表达显著下调,这提示Shh可激活HH信号(P<0.05)。在此过程中,PCNA表达水平显著升高,NRK-52E细胞数量增加,同时伴随细胞形态的改变;在Shh的基础上加入环杷明干预后,HH信号被抑制(P<0.05),PCNA表达下降,NRK-52E细胞增殖被抑制,出现凋亡或坏死。结论:HH信号与肾小管上皮细胞的增殖密切相关。通过药物调控HH信号的活化,可影响PCNA的表达,进而影响细胞增殖效应。     

溶酶体膜蛋白Sidt2缺失导致肝脏细胞自噬受损

目的 研究溶酶体膜蛋白Sidt2缺失后对肝脏自噬的影响。方法 利用Crispr-Cas9技术构建Sidt2敲除的人肝脏（HL7702）细胞模型,对自噬关键蛋白LC3Ⅱ/Ⅰ、P62及自噬相关蛋白Atg5、Atg7、Atg12进行蛋白水平检测,同时使用免疫荧光方法对LC3B及P62进行共定位,检测自噬小体对P62货物蛋白的识别与封存以及自噬溶酶体的降解。对LC3B及LAMP1进行免疫荧光共定位,检测自噬小体与溶酶体融合过程。使用LysoTracker对酸性溶酶体进行示踪。结果 成功构建HL7702细胞Sidt2+/+组 及Sidt2-/-组,HL7702细胞Sidt2-/-组较Sidt2+/+组相比,自噬关键蛋白LC3-Ⅱ/Ⅰ及P62表达均增多（P<0.01）,免疫荧光结果显示LC3B及P62亦明显增多（P<0.001）,同时自噬相关蛋白Atg5、Atg7、Atg12表达减少（P<0.05）。LC3B与P62共定位降低,LC3B 与LAMP1共定位降低,酸性溶酶体数量减少（P<0.05）。结论 Sidt2基因的缺失导致人肝脏细胞自噬小体对P62货物蛋白的识别和封存障碍,同时酸性溶酶体数量减少、自噬小体与溶酶体融合障碍导致自噬溶酶体途径受阻,最终导致LC3B与P62发生堆积,肝脏细胞自噬受损。     

Sonic hedgehog信号通路在胃癌中的研究进展

Shh（Sonichedgehog）作为最重要的Hh同源基因产物,它的异常激活与胃癌的发生、发展关系密切成为近年研究热点。经典的Shh信号通路是由Shh配体、Ptch和Smo组成的受体复合物以及下游转录因子Gli蛋白（Glil、G1i2、Gli3）组成。研究发现相比正常组织,胃癌组织中Shh的表达都有不同程度的升高,且升高程度与胃癌的发展转移呈正相关,本文就Shh信号通路与胃癌的研究进展做一综述。     

初级纤毛介导的Shh信号对大鼠骨髓基质细胞神经元样细胞分化的影响

目的 探讨初级纤毛介导的Sonic hedgehog(Shh)信号对大鼠骨髓基质细胞(MSCs)神经元样细胞分化的影响。方法 全骨髓贴壁法分离培养大鼠MSCs。实验分为白藜芦醇正常培养组、白藜芦醇诱导组、Smoothened (Smo)激动剂SAG组、Smo抑制剂环巴明组,分别给予相应的药物处理。倒置显微镜下观察各组细胞形态; 免疫荧光法检测各组细胞初级纤毛及Shh信号通路相关蛋白\[Ac-Tu、Patched (Ptc)、Smo、Gli1\]的表达; RT-PCR检测各组细胞Smo、Gli1 mRNA的表达;蛋白质印迹法检测各组细胞Smo和Gli1蛋白的表达。结果 正常培养的大鼠MSCs不表达初级纤毛。经过预诱导或饥饿处理24 h后,MSCs表达初级纤毛及Ptc、Smo和Gli1蛋白,其中Smo和Gli1蛋白位于胞质,Ptc蛋白位于初级纤毛。正常培养时,白藜芦醇不能促使Smo移位及MSCs向神经元样细胞分化。当MSCs表达初级纤毛时,白藜芦醇及SAG可使Smo从胞质进入初级纤毛,同时伴随MSCs向神经元样细胞分化及 Smo、Gli1 mRNA和蛋白表达水平的增加(P<0.05); 加入抑制剂环巴明后,Smo仍主要表达于胞质,Smo、Gli1 mRNA和蛋白的表达水平降低(P<0.05),MSCs向神经元样细胞的分化受抑制。结论 MSCs表达初级纤毛并存在Shh信号,初级纤毛介导的Shh信号参与调控大鼠MSCs的神经元样细胞分化。     

“Sirt1-自噬”在低氧诱导小鼠成牙骨质细胞凋亡中的作用

目的：初步研究“Sirt1-自噬”在低氧诱导小鼠成牙骨质细胞OCCM-30凋亡中的作用。方法：体外培养小鼠成牙骨质细胞,随机分为对照组、低氧组、低氧+Sirt1激活剂白藜芦醇组、低氧+自噬抑制剂3-MA组、低氧+白藜芦醇+Sirt1抑制剂ex527组、低氧+白藜芦醇+3-MA组,MDC染色观察自噬溶酶体变化;LC3Ⅱ免疫荧光检测自噬体变化;透射电镜观察自噬体;Western blot检测Sirt1、LC3Ⅱ、p62等蛋白表达;流式细胞仪检测细胞凋亡情况;不同组间结果数据采用单因素方差分析,组间采用LSD-t分析。结果：低氧组较对照组自噬溶酶体含量（P=0.000）及自噬体数量（P=0.000）明显增加,LC3Ⅱ蛋白表达明显增加（P=0.001）,p62蛋白表达明显减少（P=0.000）,细胞凋亡率增加（P=0.000）;低氧+白藜芦醇组较低氧组自噬溶酶体（P=0.019）及自噬体数量（P=0.000）、LC3Ⅱ蛋白表达（P=0.049）、Sirt1蛋白表达（P=0.000）明显增加,p62蛋白表达明显减少（P=0.000）,细胞凋亡率减少（P=0.021）;低氧+白藜芦醇+ex527组...     

小鼠胚胎腭突间充质细胞的分离与体外培养方法研究

目的改进小鼠胚胎腭突取材方法，原代培养C57BL／6J小鼠胚胎腭突间充质细胞并加以纯化。方法通过改良方法在显微镜下切取胚胎腭突，然后应用Dispase酶消化离体腭突，纯化腭突胚胎间充质细胞，免疫荧光染色法鉴定腭突间充质细胞生物学特性。结果改良的取材方法使小鼠胚胎腭突取材更精确，操作更简单；纯化后的原代间充质细胞几乎不含上皮细胞；免疫荧光显示细胞HNK-1、S-100、波形蛋白标记阳性，角蛋白标记阴性。结论建立了一种小鼠胚胎腭突精确取材及纯化腭突外胚间充质细胞的方法，为下一步研究工作打下基础。     

白藜芦醇介导SIRT1干预地塞米松诱导成骨细胞线粒体自噬的研究

目的 研究白藜芦醇（RES）参与糖皮质激素诱导的成骨细胞线粒体自噬的作用机制。方法 体外培养人成骨细胞hFob1.19，地塞米松（DEX）诱导细胞线粒体自噬，使用不同浓度的RES (10-8～10-6 mol/L)处理成骨细胞5～120 min,CCK-8法检测成骨细胞的增殖，透射电镜检测成骨细胞内的自噬小体，酶联免疫吸附实验、实时荧光定量PCR和Western blotting分别检测成骨细胞内碱性磷酸酶（ALP）、caspase 3、转录沉默信息调节因子1 (SIRT1)、细胞自噬标志蛋白（LC3和Beclin1）和细胞线粒体自噬标志蛋白（TOM20和Hsp60）的活性表达。结果 DEX和RES作用成骨细胞2 h后，RES可明显减弱DEX对细胞增殖的抑制作用（P <0.05）；透射电镜显示，DEX+RES组自噬小体明显少于DEX组（P <0.05）; DEX可明显抑制成骨细胞内ALP活性，而RES明显改善了DEX对ALP蛋白活性的抑制作用（P <0.05）; RES可导致成骨细胞内caspase 3的蛋白活性明显降低（P <0.05）; RES以剂量依赖性和...     

APPLICATION OF LORNOXICAM TO PATIENT-CONTROLLED ANALGESIA IN PATIENTS UNDERGOING ABDOMINAL SURGERIES

FOR anesthesiologis s ,treatingpostoperativepainhas alwaysbeen a problem.Althoughopioidshave been provedtobe effective,theirsideeffectscouldnotbeignored.With thedevelopmentofscienceand pharmacology,many drugs with aspectsof satisfactoryanalgesicefficacyand couldbe welltoleratedby patientshave been developed.And lornoxicamisone of them, which isa non-steroidalanti-inflammatorydrug (NSAID ), with analgesic, anti-infl-ammatory,andantipyreticproperties.Itseliminationhalf-time(3 to 5 hours) isle…     

Dr.Zhang Ren''''s Experience in the Acupuncture Treatment of Different Diseases with the Same Therapeutic Principle

Dr.Zhang Ren,the chief physician,is the chairman of Shanghai Acupuncture and Moxibustion Association.Having been engaged in medicine for about 40 years,he is experienced in treating various intractable diseases.In his long years of clinical practice,he advocates taking the TCM differentiation as the basis to seek for the acupuncture method for treatment of modern intractable diseases.The author of this essay had the fortune to follow Dr.Zhang in study.The following is a summary of Dr.Zhang's experience in the acupuncture treatment for different intractable diseases with the same therapeutic principle.     

Luo Lingjie''''s Experience in Treating Chronic Nephropathy

In treating chronic nephropathy,Luo Lingjie,a chief physician,pays attention to regulating the balance between yin and yang,treating infection if present,and removing pathogenic factors.He prescribes gentle drugs and uses carefully strongly warming-tonifying ones,emphasizes the importance of persuading the patient to persist in treatment with medication and nurse one's health for recuperation,and is good at combined use of TCM and western medicine therapy and brings the merits of various therapies into full play,with obvious theraoeutic effects.     

Acupuncture Combined with Spinal Tui Na for Treatment of Primary Dysmenorrhea in 30 Cases

Objective: To observe the therapeutic effects in acupunture treatment of primary dysmenorrhea combined with spinal Tui Na, and study its mechanism. Methods: Thirty cases of the treatment group were treated by acupuncture combined with spinal Tui Na, and thirty cases in the control group were treated by routine acupuncture. Results: The total effective rate was 93.3% in the treatment group, and 73.3% in the control group, with a significant difference between the two groups (P<0.05). Conclusions: Acupuncture combined with spinal Tui Na has good prospects for treatment of primary dysmenorrhea.     

猪肺磷脂注射液联合经鼻持续气道正压通气对呼吸衰竭早产儿的临床疗效及肌酸激酶同工酶活性的影响

目的 探讨猪肺磷脂注射液联合经鼻持续气道正压通气(NCPAP)对呼吸衰竭早产儿的临床疗效及肌酸激酶同工酶活性(CK-MB)的影响.方法 选取呼吸衰竭早产儿80例,分为观察组和对照组各40例.对照组采用NCPAP给氧治疗,观察组给予NCPAP给氧联合猪肺磷脂气管内给药.观察两组患儿治疗前及治疗12h、24 h后PaO2、PaCO2、血氧饱和度(SaO2)、pH的变化情况,检测治疗前及治疗5d后血清CK-MB水平;评估两组患儿的临床治疗效果.结果 两组患儿PaO2、PaCO2、SaO2、pH比较,差异均有统计学意义(P＜0.05),其中观察组治疗后的PaO2、SaO2、pH均高于对照组,PaCO2则低于对照组.两组的PaO2、SaO2、pH均随观察时间延长而升高(P＜0.05),PaCO2均随观察时间的延长而降低(P＜0.05).观察组治疗有效率为87.5％,显著高于对照组的70.0％ (P ＜0.05).治疗5d后两组患儿血清CK-MB水平均较前降低(P＜0.05),且观察组明显低于对照组(P＜0.05).结论 猪肺磷脂注射液气管内给药联合NCPAP可以显著降低呼吸衰竭早产儿CK-MB的含量,提高治疗有效率,起到很好的呼吸循环支持作用.     

Acupuncture Treatment of Vomiting

CASE HISTORY A female patient, 46 years old, head of the foreign affairs department of a certain university in Beijing, paid her first visit on October 9, 2006, with the chief complaint of vomiting for one month. She got vomiting after meals in early September. Before that, she had discomfortable sensation in the stomach due to angry with others, but she didn't pay much attention. Later, it developed into vomiting after eating. After the vomiting, the discomfort would be relieved, but with slight hypodynamia. She was once diagnosed as having 'neurogenic vomiting'. Having taken some western and Chinese drugs, the above symptoms were a little bit improved, but she would have nausea upon eating and with regurgitation. Because of the fear for vomiting, she did not dare to have food intake, with body weight reduction of 6 kilos in one month.     

Qiu Baoguo''''s Experience in Treating Consumptive Syndromes after Radio-Chemotherapies

Radiotherapy and chemotherapy are the important modern medical therapies for malignant tumors,yet they can also bring about serious local and systemic toxic side reactions so to decrease the patient;'s life quality,manifested by a series of consumptive symptoms.Having engaged in the combined work of Chinese and western medicine for nearly 50 years,the research fellow Qiu Baoguo in Henan Provincial Academy of TCM has developed his unique views on the TCM study of consumptive syndromes.The author of this essay had once the fortune tO follow Dr.Qiu in clinic,and specially would like to introduce in the following Dr.Qiu's experience in treating consumptive syndromes after radio-chemotherapies for patients with malignant tumor.     

Insomnia Due to Deficiency of Both the Heart and Spleen Treated by Acupuncture-Moxibustion and Chinese Tuina

OBJECTIVE: To observe therapeutic effects of the comprehensive therapy of acupuncture-moxibustion and Chinese Tuina for treatment of insomnia due to deficiency of both the heart and spleen. METHODS: 92 cases were divided randomly into the treatment group (treated by acupuncture-moxibustion and Chinese Tuina) and the control group (treated by acupuncture-moxibustion). RESULTS: The therapeutic effect of the treatment group was obviously superior to that of the control group (the CHI2 test showed P < 0.01). CONCLUSIONS: The comprehensive therapy of acupuncture-moxibustion and Chinese Tuina can give marked therapeutic effects for treatment of insomnia due to deficiency of both the heart and spleen.     

Survey and practice of reporting quality of randomized controlled clinical trials on traditional Chinese medicine

Evidence obtained from randomized controlled trials （RCTs） has been generally accepted as the gold standard in the evaluation of clinical effectiveness. Readers need to understand the trial design, implementation, results, analysis and interpretation, so as to fully Jnderstand the results of RCTs. Thus, the investigators of RCTs have to report these items in a complete, accurate and clear manner. Since 1998, we have conducted several evaluations on the reporting quality of RCTs published in Chinese journals on traditional Chinese medicine （TCM） and results have shown that there is an urgent need for higher quality RCTs on TCM.