脂肪来源干细胞体外成骨和成脂及成神经的诱导分化研究

目的 探讨脂肪组织来源干细胞 (adipose stem cell,ADSC) 的表面表型及向成骨、成脂和成神经多向分化的生物学特性.方法 取SD大鼠的腹股沟、附睾垫和腹膜后脂肪,Ⅰ型胶原酶消化、分离培养ADSC,检测第3代ADSC的CD29、CD44和CD45的表达情况.将ADSC加入成骨诱导剂,通过茜素红S染色鉴定成骨细胞;成脂诱导剂利用油红O染色鉴定脂肪细胞;神经干培养基诱导,免疫荧光染色鉴定诱导的神经元.结果 ADSC表达CD29(99.07±0.12)%、CD44(95.82±0.68)%和CD45(0.11±0.12)%.其经成骨诱导4周,茜素红S染色显示聚集的细胞团中央形成红色钙化结节,碱性磷酸酶染色可见到细胞的胞质内有紫红色颗粒;成脂诱导1周,油红O染色可见细胞质内有橙红色脂滴;经过神经干培养基诱导6 d后,免疫荧光染色显示诱导的细胞Nestin、NF-200及GFAP阳性表达.结论 ADSC表达间质干细胞的表面表型,体外经诱导培养后可向成骨细胞、脂肪细胞和神经元多向分化,表明 ADSC具有干细胞的多向分化潜能.     

绿色荧光蛋白转基因小鼠脂肪间充质干细胞的多向分化潜能

目的:研究绿色荧光蛋白(GFP)转基因小鼠脂肪间充质干细胞(ADSC)体外定向诱导下的多向分化潜能.方法:取出生后4 d的GFP转基因小鼠腹股沟的皮下脂肪组织,经过胶原酶消化,加入脂肪组织来源干细胞条件培养基中培养.将生长状态良好的第5代GFP-ADSCs分别加入成骨及成脂条件培养液进行体外多向分化诱导研究.并分别采用茜素红钙盐染色和油红O染色进行鉴定.结果:成功获得了稳定表达GFP的ADSCs,GFP-ADSC经成骨诱导20 d后茜素红染色可见有橘红色钙盐沉积;经成脂诱导10 d后油红O染色阳性.结论:GFP作为报告基因在ADSCs分化过程中没有被关闭,亦未影响到ASCs的多向分化潜能,是ADSC在体多向分化研究的一个有效示踪工具.     

大鼠脂肪基质干细胞的体外培养及其成骨细胞分化特性的研究

目的研究脂肪基质干细胞（ADSCs）分离培养的方法,探讨大鼠ADSCs在体外向成骨细胞分化的能力。方法从成年SD大鼠腹股沟处获取脂肪组织后进行体外培养,并通过骨诱导培养基使其分化成成骨细胞,通过碱性磷酸酶（ALP）v、on Kossa染色鉴定向成骨细胞分化能力。采取逆转录-聚合酶链反应（RT-PCR）技术检测成骨细胞的标志基因。结果大鼠脂肪组织能够分离培养出ADSCs;向成骨细胞诱导,ALP、von Kossa染色阳性,RT-PCR检测有ALP、osteocalcin及osteopontin表达。结论成功从大鼠脂肪组织分离培养出ADSCs,其生物学特性与骨髓基质干细胞（MSCs）相似,能够向成骨细胞分化,有希望成为组织工程理想的种子细胞来源。     

脂肪组织分泌物中蛋白浓度对诱导脂肪干细胞成脂的影响

目的 观察胎牛血清(FBS)对体外培养获取的脂肪组织分泌物(ATE)诱导大鼠脂肪干细胞(ADSCs)成脂能力有无影响,并探讨ATE中不同蛋白浓度对诱导大鼠ADSCs成脂、细胞增殖和迁移能力的影响。 方法 培养大鼠ADSCs并传代,对ADSCs成骨和成神经诱导,采用茜素红染色和神经丝蛋白(NF)免疫荧光检测鉴定。含有FBS和无FBS的培养基对脂肪组织块进行培养并收集ATE,对第4代ADSCs进行诱导,第7 d时油红染色鉴定并计算成脂率;收集无FBS培养基获取的ATE,分为不同蛋白质量浓度组(100 μg/mL、250 μg/mL、500 μg/mL组),对第4代ADSCs进行成脂诱导,观察各组成脂率;并观察不同蛋白质量浓度组对ADSCs细胞增殖和迁移的影响。 结果 培养的ADSCs成骨和成神经诱导后茜素红染色和NF免疫荧光染色阳性;获取ATE时添加或不添加FBS,在第3 d时均能诱导ADSCs成脂,第7 d时的成脂率差异无统计学意义;ATE 500 μg/mL组较100 μg/mL、250 μg/mL组成脂率高(P均<0.05),细胞培养2 d后,3个不同蛋白质量浓度组均抑制细胞增殖,不同蛋白质量浓度间细胞抑制率差异无统计学意义(P均>0.05);ATE中不同蛋白质量浓度对细胞的迁移无影响。 结论 无FBS的培养基获取的ATE,短期内对ADSCs成脂能力没有影响;ATE蛋白浓度与ADSCs成脂率有关。     

培养的人骨髓间充质干细胞向成骨细胞、脂肪细胞分化

目的探讨体外培养的人骨髓间充质干细胞(MSCs)向成骨细胞、脂肪细胞分化的方法,以及钙结节形成、Ⅰ型胶原表达等情况. 方法分离人骨髓间充质干细胞,经过原代培养和传代培养,分别加入成骨诱导、脂肪诱导培养体系,经过倒置显微镜观测与HE染色了解诱导后细胞形态变化,并通过von Kossa染色、Ⅰ型胶原免疫组化染色、苏丹黑染色与油红O法染色鉴定细胞的性质.另取2代细胞体外培养,加入成骨诱导分化试剂,以碱性磷酸酶(ALP)为检测指标计数细胞阳性率,比较成骨诱导1、2、3、4周时细胞碱性磷酸酶的变化.结果体外培养的MSCs在合适的条件下能够向成骨细胞、脂肪细胞分化.细胞经成骨诱导2～3周时细胞碱性磷酸酶阳性率最高,达到85%.结论 MSCs体外培养在一定条件下向成骨细胞、软骨细胞与脂肪细胞分化,体外培养时形成钙结节,并表达Ⅰ型胶原.培养2～3周后细胞经成骨诱导分化细胞ALP阳性率较高.     

两种培养基诱导人和大鼠脂肪干细胞向脂肪细胞分化的比较

目的比较两种培养基诱导人和大鼠脂肪干细胞(adipose-derived stem cell,ADSC)的成脂分化能力,探寻更为适宜的ADSC成脂诱导方案。方法提取3例临床患者的脂肪干细胞(h ADSCs)及出生6周大鼠的脂肪干细胞(r ADSCs),将h ADSCs、r ADSCs分别接种于六孔板,分为未诱导组、成脂诱导I组和II组,分别使用普通培养基、成脂诱导培养基I和II,培养10 d后以油红O染色,镜下观察及检测490 nm吸光度值(A490),比较脂滴形成情况。结果未诱导的h ADSCs和r ADSCs均呈成纤维细胞样生长,成脂诱导培养3 d后出现脂滴;油红O染色显示,成脂诱导组脂滴呈橘红色,h ADSCs和r ADSCs的成脂诱导II组形成的脂滴均明显大于诱导I组,统计学分析显示h ADSCs的成脂诱导II组测得的A490明显高于诱导I组(P<0.01);而两组的r ADSCs的A490无明显差异(P>0.05)。结论 h ADSCs和r ADSCs在两种诱导培养基中均可成脂分化,但成脂诱导II组优于成脂诱导I组。     

大鼠脂肪干细胞体外定向诱导分化为脂肪细胞和成骨细胞的研究

目的研究大鼠脂肪干细胞的基本生物学特性及其向脂肪细胞和成骨细胞诱导分化潜能。方法取6周龄SD大鼠腹股沟处脂肪组织,胶原酶消化法分离出脂肪干细胞,体外培养。取第3代细胞,免疫细胞化学测定其CD44、CD34抗原表达,分别用成脂和成骨诱导培养液诱导其向脂肪细胞及成骨细胞分化。油红染色、碱性磷酸酶染色和vonKossa染色鉴定其分化能力。结果大鼠脂肪干细胞生长能力旺盛,表达CD44,而不表达CD34;定向诱导后,油红染色、碱性磷酸酶染色和vonKossa染色阳性。结论大鼠脂肪组织可分离培养出脂肪干细胞,能向脂肪细胞和成骨细胞诱导分化,有可能成为组织工程理想的种子细胞来源之一。     

人脂肪干细胞体外培养及向软骨细胞诱导分化的实验研究

目的:观察人脂肪基质干细胞(ADSCs)体外分离培养及成脂、成软骨诱导分化的生物学性状,探讨其作为软骨组织工程种子细胞的可行性.方法:(1)人脂肪组织来源于健康成年女性腹部吸脂术,酶消化法分离出ADSCs,体外培养扩增.(2)取第3代ADSCs,测细胞生长曲线,流式细胞仪测原代细胞表面标记;成脂和成软骨诱导分化.(3)倒置显微镜观察油红O、阿尔辛蓝(AB-PSA)染色和Ⅱ型胶原免疫细胞化学染色检测分化情况;RT-PCR检测相关标志基因表达.结果:(1)体外培养的ADSCs细胞形态均一,传代稳定.(2)经成脂诱导,细胞内出现空泡,油红O染色呈红色,RT-PCR检测到有Leptin、PPAR-γ表达;经成软骨诱导,AB-PSA染色呈紫红色,Ⅱ型胶原免疫细胞化学染色胞浆呈棕黄色,RT-PCR检测COLLⅡ、SOX9和aggrecan表达.结论:脂肪干细胞能向软骨细胞方向诱导分化,可作为软骨组织工程种子细胞.     

飞机辅助动力装置引气特性计算方法

目的 观察胎牛血清(FBS)对体外培养获取的脂肪组织分泌物(ATE)诱导大鼠脂肪干细胞(ADSCs)成脂能力有无影响,并探讨ATE中不同蛋白浓度对诱导大鼠ADSCs成脂、细胞增殖和迁移能力的影响.方法 培养大鼠ADSCs并传代,对ADSCs成骨和成神经诱导,采用茜素红染色和神经丝蛋白(NF)免疫荧光检测鉴定.含有FBS和无FBS的培养基对脂肪组织块进行培养并收集ATE,对第4代ADSCs进行诱导,第7d时油红染色鉴定并计算成脂率;收集无FBS培养基获取的ATE,分为不同蛋白质量浓度组(100 μg/mL、250 μg/mL、500μg/mL组),对第4代ADSCs进行成脂诱导,观察各组成脂率;并观察不同蛋白质量浓度组对ADSCs细胞增殖和迁移的影响.结果 培养的ADSCs成骨和成神经诱导后茜素红染色和NF免疫荧光染色阳性;获取ATE时添加或不添加FBS,在第3d时均能诱导ADSCs成脂,第7d时的成脂率差异无统计学意义;ATE 500 μg/mL组较100 μg/mL、250 μg/mL组成脂率高(P均＜0.05),细胞培养2d后,3个不同蛋白质量浓度组均抑制细胞增殖,不同蛋白质量浓度间细胞抑制率差异无统计学意义(P均＞0.05);ATE中不同蛋白质量浓度对细胞的迁移无影响.结论 无FBS的培养基获取的ATE,短期内对ADSCs成脂能力没有影响;ATE蛋白浓度与ADSCs成脂率有关.     

人骨髓间质干细胞成骨向脂肪转分化的研究

目的建立人骨髓来源的间质干细胞成骨向脂肪转分化的体外培养条件,为了解成骨向脂肪转分化的分子机制和骨质疏松的发病机制提供基础。方法运用Haynesworth法分离出间质干细胞,经向成骨细胞和脂肪细胞诱导分化后,将分化后的成骨细胞向脂肪细胞转分化诱导,碱性磷酸酶（ALP）、油红O染色后,RT-PCR检测转分化效果。结果转分化后的脂肪细胞强表达脂蛋白脂肪酶（LPL）和过氧物酶体增殖物激活受体y2（PPARy2）,弱表达ALP,但完全不表达骨钙素（OC）;转分化后的脂肪细胞与正常脂肪细胞相似,油红0染色显示脂滴在胞质内积聚成小泡。结论本试验建立了hMSCs成骨向脂肪转分化的体外培养体系,为了解骨质疏松的发病机制奠定了基础;hMSCs向脂肪定向分化优于向成骨分化。     

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.     

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.     

TCM Treatment of Ankylosing Spondylitis by Tonifying the Kidney and Strengthening the Governor Vessel

Ankylosing spondylitis is a chronic and progressive disorder with inflammation mainly involving the central axis joints. It mainly affects the cervical spine and the lumbosacral area, with the pathogenesis closely related to the kidney and the Governor Vessel (GV). TCM holds that the syndrome is deficiency in origin and excess in superficiality, which is due to insufficiency of the kidney, deficiency of GV, and blocking of the channels with the invasion of exogenous evil, leading to poor circulation of qi and blood and malnutrition of the bones, muscles and joints. The TCM method of tonifying the kidney and strengthening GV to regulate circulation of qi and blood and check the arthralgia pain should be adopted, with the Kidney-Tonifying and GV Strengthening Decoction (益肾强督汤) prescribed.     

IMPROVING P-gp EXPRESSION IN HUMAN MONONUCLEAR CELLS IN VITRO TRANSFECTED BY MULTIDRUG RESISTANCE-1 mRNA

CHEMOTHERAPY playsa greatrolein the treat- ment of malignanttumors,especiallyingynecolo- gicalones.But inanticancerchemotherapy,leuko-cytopeniaisfrequentlytheprimarydose-limitingsideeffect factor.Moreover,cancersarefrequentlychemoresistantbe-causeof overexpressionof P-glycoprotein(P-gp), which isencodedby multidrugresistancegene (MDR1 ) and detectableinup to50% ofhuman cancersand renderscellsresistancetoanticancerdrugs.The safetyand potentialtherapeuticbenefitof mdr1 gene transferredto h…