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1.
目的 探讨Ghrelin联合LY294002对脂肪间充质干细胞(ADSCs)神经分化的影响,并阐明其作用机制。 方法 倒置相差显微镜下观察ADSCs形态表现,流式细胞术鉴定ADSCs表面抗体阳性表达率,将第4代ADSCs分为对照组(不进行干预)、LY294002组[给予40 μmol·L-1磷酸酰肌醇3激酶(PI3K)/蛋白激酶B(Akt)通路特异抑制剂LY294002]、Ghrelin组(给予0.1 μmol·L-1 Ghrelin)、Ghrelin+LY294002组(给予40 μmol·L-1 LY294002+0.1 μmol·L-1 Ghrelin)。倒置相差显微镜下观察各组ADSCs神经分化后的形态表现,免疫荧光染色法检测各组ADSCs中神经元特异性烯醇化酶(NSE)、巢蛋白(Nestin)和胶质纤维酸性蛋白(GFAP)阳性细胞百分率,Western blotting法检测各组ADSCs中 PI3K/Akt通路蛋白表达水平,并计算磷酸化PI3K(p-PI3K)/PI3K和磷酸化Akt(p-Akt)/Akt比值。 结果 ADSCs表面抗体CD90和CD13高表达,CD34、CD45和CD106低表达。与对照组比较,LY294002组仅部分细胞由长梭形转变为类圆形胞体,突起较短、数量减少,而Ghrelin组细胞由长梭形转变为类圆形胞体速度增快;与LY294002组比较,Ghrelin+ LY294002组细胞由长梭形转变为类圆形胞体速度增快,且突起数量增加。与对照组比较,LY294002组ADSCs中NSE和Nestin阳性细胞百分率及p-PI3K/PI3K和p-Akt/Akt比值明显降低(P<0.05),Ghrelin组ADSCs中NSE和Nestin阳性细胞百分率及p-PI3K/PI3K和p-Akt/Akt比值升高(P<0.05);与LY294002组比较,Ghrelin+LY294002组ADSCs中NSE和Nestin阳性细胞百分率及p-PI3K/PI3K和p-Akt/Akt比值升高(P<0.05);各组ADSCs中GFAP阳性细胞百分率比较差异无统计学意义(P>0.05)。 结论 Ghrelin通过激活PI3K/Akt通路促进ADSCs神经分化。  相似文献   

2.
目的 探讨胰岛素样生长因子-1(insulin-like growth factor-1,IGF-1)对脂肪间充质干细胞(adipose-derived stem cells,ADSCs)迁移能力的影响,为勃起功能障碍的治疗建立实验基础。方法 采用酶消化法分离提取SD大鼠皮下脂肪干细胞,以含有10%的胎牛血清的DMEM培养基培养。通过流式细胞仪检测ADSCs表面标志物(CD34、CD44、CD45)的表达情况。采用终浓度为20ng/ml IGF-1预处理ADSCs,观察IGF-1对ADSCs迁移能力以及CXCR4蛋白的表达情况。结果 流式细胞术显示ADSCs的表型分子CD44呈阳性,CD34、CD45呈阴性。IGF-1可上调CXCR4的表达,并且促进 ADSCs向SDF-1迁移。结论 本实验成功分离培养ADSCs,IGF-1能够提高ADSCs表面CXCR4蛋白的表达,从而促进ADSCs迁移,并为下一步将IGF-1预处理的ADSCs应用于大鼠体内勃起功能障碍的治疗提供基础。  相似文献   

3.
目的 探讨小鼠骨髓间充质干细胞(BM-MSCs)自噬与衰老的关系。方法 通过体外分离培 养年轻组小鼠(8 周龄)和老年组小鼠(18 个月龄)BM-MSCs,流式细胞术检测细胞表型,TUNEL 染色检 测细胞凋亡,ELISA 试剂盒检测BM-MSCs 分泌蛋白VEGF、bFGF、IGF-1、HGF 的表达,Western blot 检 测自噬相关蛋白LC3- Ⅰ、LC3- Ⅱ、Beclin 1、ATG12-5、p62 及信号通路蛋白p-Akt、Akt、p-mTOR、 mTOR 的表达。结果 与年轻组比较,老年组BM-MSCs 凋亡增加(P <0.05),分泌功能下降(P <0.05),自 噬相关蛋白LC3- Ⅰ、LC3- Ⅱ、Beclin 1、ATG12-5 表达水平下降(P <0.05),相反p62 蛋白表达水平升高 (P <0.05),自噬信号通路相关蛋白p-Akt、Akt、p-mTOR、mTOR 表达升高。结论 老年小鼠BM-MSCs 凋亡增加,自噬活性及旁分泌功能下降。  相似文献   

4.
目的研究肝细胞生长因子(HGF)对大鼠骨髓间充质干细胞(BMSCs)迁移的影响。方法采用Percoll密度梯度离心法分离BMSCs,进行表面抗原表达及分化鉴定。使用Dunn chamber装置研究BMSCs的定向迁移。并且研究PI3K抑制剂LY294002对HGF诱导BMSCs迁移的影响。结果分离培养的BMSCs呈CD29、CD90、CD106阳性表达,CD34、CD45阴性表达,BMSCs可诱导分化为成骨细胞及脂肪细胞。Dunn chamber迁移实验显示,外槽加入不同浓度的HGF,BMSCs迁移速率没有变化,迁移效率随着HGF浓度的增加而增加,50ng/ml与100ng/mlHGF均显著提高了细胞迁移效率;内外槽同时加入50ng/mlHGF,迁移速率与迁移效率均没有变化;经30μmol/LLY294002预处理1h后,HGF诱导的BMSCs的定向迁移受到抑制。结论 HGF能够趋化BMSCs的定向迁移,PI3K信号通路参与介导这一过程。  相似文献   

5.
目的 探讨MAD2B基因过表达对毛乳头细胞(dermal papilla cells,DPCs)生物学功能的影响.方法 构建真核表达载体pIRES2-MAD2B和pIRES2-TCF4质粒,在Lpofectamine TM 2000脂质体的介导下转染毛乳头细胞,荧光显微镜下观察转染效率,使用Western blot检测目的蛋白的表达,并验证两种蛋白的相互作用,比较细胞增殖状况,Wnt通路信号强度,毛乳头分泌胰岛素样生长因子-1(insulin-like growth factor-1,IGF-1)、血管内皮生长因子(vascular endothelial growth factor,VEGF)、肝细胞生长因子(hepatocyte growth-promoting factors,HGF)的变化情况.结果 成功构建pIRES2-MAD2B和plRES2-TCF4重组质粒,且转染后发现MAD2B和TCF4蛋白过表达于毛乳头细胞.免疫共沉淀实验结果提示在凝集生长的毛乳头细胞中存在相互作用的MAD2B和TCF4蛋白质复合物.在MAD2B过表达于毛乳头细胞之后,发现Wnt途径信号通路强度下调.CCK-8实验结果提示MAD2B过表达后细胞增殖活性下降,与对照组比较有统计学差异(P<0.05);同时实时荧光定量PCR检测结果显示,与对照组相比,IGF-1、VEGF、HGF mRNA的表达有所下降(P<0.05).结论 免疫共沉淀提示MAD2B与TCF4在毛乳头细胞中存在相互作用,MAD2B过表达后可能通过抑制Wnt信号通路而对毛乳头细胞增殖和其旁分泌因子IGF-1、VEGF、HGF产生抑制作用.  相似文献   

6.
 【目的】观察VEGF(内皮生长因子)和PlGF(胎盘生长因子)对小鼠胎肝造血干细胞髓系分化的影响,并探讨其调控机制;【方法】体外培养E13胎肝造血细胞,培养造血集落CFU-GM,观察VEGF和PlGF对胎肝造血干细胞髓系分化的作用,应用Western-Blot检测不同处理条件对胎肝造血细胞Akt蛋白以及p-Akt蛋白表达水平的影响,用MTT法来评估不同浓度的LY294002(PI3K/Akt抑制剂)对胎肝造血细胞增殖的影响,通过CFU-GM观察LY294002对VEGF和PlGF调控胎肝造血作用的影响;【结果】VEGF和PlGF均可促进胎肝造血干细胞髓系分化,集落数分别为对照组的141%(P < 0.01)和123%(P < 0.05); PlGF可上调胎肝造血细胞p-Akt蛋白表达,其表达水平为对照组的143%(P < 0.05);VEGF及其与PlGF联合应用对胎肝造血细胞p-Akt蛋白表达水平无明显影响;LY294002可抑制胎肝造血细胞的增殖,并具有明显的剂量依赖关系(r = 0.9647,P < 0.01),在LY294002存在的条件下加入VEGF或PlGF,其CFU-GM产率与单独应用LY294002相比无明显差异(P > 0.05);【结论】VEGF和PlGF均能促进胎肝造血干细胞的髓系分化;PlGF可通过上调胎肝造血细胞Akt的磷酸化水平调节胎肝造血细胞的髓系分化;LY294002可通过抑制PI3K/Akt信号通路抑制VEGF和PlGF对胎肝造血干祖细胞髓系分化的调控作用;  相似文献   

7.
Ning YJ  Zhang W  Cheng JF  Li XL  Wang MF  Lin L 《中华医学杂志》2010,90(34):2402-2406
目的 探讨胰岛素样生长因子1(IGF-1)调节胃平滑肌细胞(SMC)表达干细胞因子(SCF)的信号传导途径.方法 培养SD大鼠胃SMC,α-肌动蛋白免疫荧光鉴定,Western印迹、反转录-聚合酶链反应(RT-PCR)检测胃SMC表达SCF的变化情况:(1)IGF-1作用后;(2)IGF-1受体(IGF-1Rα)抗体阻断IGF-1R后;(3)PD-98059、LY-294002分别抑制丝裂原激活蛋白激酶的激酶(MEK)、磷脂酰激醇3激酶(PI3K)后.结果 在无血清培养条件下,SCF在大鼠胃SMC中少量表达,低剂量IGF-1(5和10μg/L)对SCF表达均无影响(均P>0.05);中、高剂量IGF-1(50、100、150 μg/L)诱导分别使SCF蛋白表达上调达2.79、5.51及5.35倍,mRNA表达上调达1.81、2.54及2.38倍(均P<0.05);且促进SCF合成的最高峰在第16小时(蛋白表达上调达2.36倍,mRNA表达上调达5.51倍,均P<0.05).IGF-1Rot抗体可抑制SCF合成,抑制作用呈浓度依赖性(均P<0.05).分别用PD-98059、LY-294002预处理SMC、再用IGF-1诱导,PD-98059能部分阻断SCF的表达(蛋白抑制率为23%,mRNA抑制率为48%,均P<0.05)、LY-294002对SCF的表达无影响(P>0.05).结论 IGF-1可通过IGF-1R以时间和剂量依赖性方式诱导胃SMC产生SCF,且IGF-1诱导SCF的表达可能依赖于ERKMAPK信号传导通路.  相似文献   

8.
目的:分离培养脂肪干细胞(ADSCs)以探讨其分泌因子对HaCaT细胞的生物学作用.方法:从人腹部脂肪组织中分离并培养ADSCs3d后,ELISA法测定ADSCs分泌血管内皮生长因子(VEGF),肝细胞生长因子(HGF)含量;收集ADSCs培养液(ADSC-CM)作用于HaCaT细胞株.采用MTT法测定HaCaT细胞的增殖;AnexinV/PI双染色法测定HaCaT细胞的凋亡情况;体外细胞划痕法测定其迁移能力.结果:培养获得ADSCs细胞3d后,培养液中VEGF、HGF含量分别为(287±47),(577±85)ng/L.MTT实验30%ADSC-CM组、50%ADSC-CM组的A490nm值分别高于对照组,差异具有统计学意义(P〈0.05或P〈0.01).实验组HaCaT细胞凋亡率明显低于对照组(P〈0.05).与对照组相比较,实验组HaCaT细胞迁移距离36,48h时间点显著增大(P〈0.05或P〈0.01).结论:ADSCs能分泌细胞因子且对HaCaT细胞具有促增殖、迁移和抑制凋亡作用.  相似文献   

9.
目的:细胞水平观察不同浓度的辛伐他汀对骨髓间充质干细胞(bone marrow-drived mesenchymal stem cells,BMSCs)的增殖及旁分泌功能的影响并探讨其可能的机制?方法:采用全骨髓贴壁培养法培养SD大鼠的骨髓间充质干细胞,取3代对数生长期细胞,无血清培养12 h,不同剂量的辛伐他汀(0?0.001?0.010?0.100?1.000 ?滋mol/L)与MSCs共培养24 h,采用CCK-8试剂盒检测各浓度组BMSCs增殖情况,通过半定量RT-PCR检测不同浓度的辛伐他汀诱导BMSCs中血管内皮生长因子(vascular endothelial growth factor,VEGF)和肝细胞生长因子(hepatocyte growth factor,HGF)的mRNA表达?采用Western blot检测PAkt?AKt的表达?结果:辛伐他汀在一定浓度范围内(0.001~0.100 ?滋mol/L)显著提高了BMSCs增殖和VEGF和HGF的mRNA表达水平(与对照组比较,P < 0.01)?辛伐他汀浓度在0.010 ?滋mol/L时对BMSCs的增殖和分泌功能的影响最为显著,随着药物浓度的加大,BMSCs的上述功能呈下降趋势?与对照组比较,1.000 ?滋mol/L辛伐他汀对BMSCs无显著的促增殖作用,但却明显上调了VEGF和HGF mRNA的表达?0.010 ?滋mol/L辛伐他汀组pAKt表达及pAKt/AKt比值显著高于对照组(P < 0.01)?结论:辛伐他汀在一定浓度范围内对骨髓间充质干细胞的增殖及分泌功能有促进作用,其机制可能与Akt信号通路有关?  相似文献   

10.
目的:观察低氧条件下,大鼠骨髓间充质干细胞分泌VEGF、TGF、IL-6、bFGF、HGF和的变化情况。方法:应用ELISA方法检测正常组和低氧组细胞上清中的VEGF、TGF、IL-6、bFGF和HGF的浓度。结果:低氧组的营养因子浓度明显高于正常组,具有统计学意义。结论:骨髓间充质干细胞在移植早期可能通过高表达营养因子而发挥促修复作用。  相似文献   

11.
Objective To observe blood pressure change with age in salt-sensitive teenagers whose salt sensitivity were determined by repeated testing.Methods Salt sensitivity was determined through intravenous infusion of normal saline combined with volume-depletion by oral diuretic furosemide in 55 teenagers. After five years, salt sensitivity was re-examined and subject blood pressure was followed up. Blood pressure changes in salt-sensitive teenagers were compared to that of non-salt sensitive teenagers over five years.Results After 5 years, the repetition rate of salt sensitivity determined by intravenous saline loading is 92.7%. In teenagers with salt sensitivity on the baseline, both the systolic blood pressure increments and increment rates were much higher than non-salt sensitive teenagers (12.7±12.1 mmHg vs. 2.8±5.2 mmHg, P< 0.01; 12.2%± 12.0% vs. 2.5% ±4.4%, P< 0.001,respectively). There was a similar trend for diastolic blood pressure (8.4 ± 6.4 mmHg vs. 3.7 ± 6.4 mmHg, P = 0.052; 13.2% ±10.6 % vs. 6.8%± 10.1%, P = 0.053, respectively).Conclusions Salt sensitivity determined by intravenous saline loading showed good reproducibility. Blood pressure increments with age were much higher in salt-sensitive teenagers than non-salt sensitive teenagers, especially in terms of systolic blood pressure.  相似文献   

12.
Objective: To evaluatel the value of D-dimers in patients with acute aortic dissection (AAD). Methods: This study consisted of 16 patients with AAD and 27 non-AAD patients. Serum D-dimets were measured by Sta-Liatest D-DI immunoturbidimetric assay. Results: D-dimer level was higher (P < 0.001) in patients with AAD(7.91 ± 5.52 μg/ml) than that in non- AAD group(1.57±1.24 μg/ml). D-dimer was positive (>0.4 μg/ml) in all patients with AAD and in 10 control group patients (37%). Among patients with acute AAD, D-dimers tended to be higher in Stanford A than in Stanford B (8.67 ± 4.31 μg/ml vs. 3.24±1.27 μg/ml, P <0.01). D-dimer values tended to be higher in more extended disease(3.84 ± 1.65 μg/ml, 8.57 ± 3.58 μg/ml and 11.87 ± 5.69 μg/ml in thoracic aorta, thoracic and abdominal aorta, thoracic and abdominal aorta and iliacal arteries, respectively, P < 0.05 for both 8.57 ± 3.58 and 11.87 ± 5.69 vs. 3.84 ± 1.65 ). Including the control group into the analysis, we found a sensitivity of 100%, a negative predictive value of 100%, and a specificity of 66% and a positive predictive value of 64% for D-dimer in diagnosis of AAD in our patients with suspected AAD. Conclusion: D-dimer was elevated in patients with AAD. A negative D-dimer test result could be useful in excluding AAD.  相似文献   

13.
Objective: To set up a simple and reliable rat model of combined liver-kidney transplantation. Methods: SD rats served as both donors and recipients. 4℃ sodium lactate Ringer's was infused from portal veins to donated livers,and from abdominal aorta to donated kidneys, respectively. Anastomosis of the portal vein and the inferior vena cava (IVC) inferior to the right kidney between the graft and the recipient was performed by a double cuff method, then the superior hepatic vena cava with suture. A patch of donated renal artery was anastomosed to the recipient abdominal aorta. The urethra and bile duct were reconstructed with a simple inside bracket. Results: Among 65 cases of combined liver-kidney transplantation, the success rate in the late 40 cases was 77.5%. The function of the grafted liver and kidney remained normal. Conclusion: This rat model of combined liver-kidney transplantation can be established in common laboratory conditions with high success rate and meet the needs of renal transplantation experiment.  相似文献   

14.
Increasing maternal age is the only etiological factor unequivocally linked to Down's syndrome in humans. The occurrence rate of newborns with Down's syndrome is about 1/220 in women over 35 years old. However, the occurrence rate in embryos fertilized in vitro, of the elder woman is unclear. Using FISH we screened the number of chromosome 21 in preimplanted embryos of 5 elderly women (average age, 38.4 years) to study the feasibility and necessity of screening trisomy 21 in embryos in patients over 35 years old at the in vitro fertilization (IVF) center.  相似文献   

15.
Real-time three-dimensional echocardiography (RT3DE)is a new ultrasound technique that enables dynamic threedimensional visualization and quantification of the heart in real time. Investigation of feasibility and methodology of RT3DE in determining left ventricular (LV) and right ventricular (RV) volumes, RT3DE was performed in 35 normal adults using Philips SONOS 7500 system with a 2-4 MHz matrix array transducer. The 60°×60° "pyramid" volume database was obtained and analyzed on a TomTec echo workstation. Both LV and RV volumes were calculated with four 3DE methods (i.e. apical 2, 4, 8, and 16-plane) through manually tracing ventricular endocardial borders in end diastole and end systole. Stroke volumes were then calculated. LV volume was also measured by 2DE Simpson's rule using GE VIVID 7 ultrasound machine.  相似文献   

16.
A clinical guideline for the therapeutic interventions of integrative medicine may be defined as a written document which states a series of recommendations on therapeutic interventions of integrative medicine for a special disease or condition. The guideline may provide assistance to medical professionals in making clinical decisions aimed at improving the clinical outcome of patients and reducing the costs of medical care(~'4~. Recommendations issued by a guideline should be based on the best available evidence in both Western and Chinese medicine. For fulfilling this purpose, the development of clinical guidelines for therapeutic interventions in the field of integrative medicine should follow scientific principles and undergo a rigorous processes.  相似文献   

17.
Objective:To probe into the influence of changes of ovarian hormones on the pathogenesis of the specific sub-type premenstrual syndrome(PMS)and reveal partial microcosmic mechanisms of adverse flow of liver-qi.Methods:Estradiol(E2)and progesterone(P)levels in serum were determined at different phases of menstrual cycle by radioimmunoassay.Results:In the group of PMS with adverse flow of liver-qi.the secretive peak value Of E2 and P at the follicular phase significantly decreased,and the secretive peak value at the luteal phase did not come into being.Conclusions:Low E2 and P secretive peak at the follicular phase and absence of secretive peak at the luteal phase is one of the microcosmic mechanisms of PMS with adverse flow of liver-qi.One of the pathophysiologic mechanisms of specific sub-type PMS is probably the continuous low level of E2and P.  相似文献   

18.
Journal of Nanjing Medical University (English Edition) JNMU, sponsored by Nanjing Medical University, was established in 1987. It is a bimonthly comprehensive English medical journal published locally and abroad.Since 2007, Journal of Nanjing Medical University (English Edition )was granted Elsevier the full publishing and distribution rights worldwide for the Electronic Edition, excluding the People's Republic of China.  相似文献   

19.
Objective To measure Derpl and Blot5 allergen levels in asthmatics' homes in Hongkong.Methods Seventy houses were enrolled for a mite indoor environment study. Dust samples were obtained from two sites of each patients' house: bed and floor. Derpl and Blot5 levels were quantified by a two-site monoclonal antibody-based ELISA technique.Results The levels of Derpl allergens found in bed (geometric mean (GM) 3.43 μg/g of dust; 95%CI, 1.89-4.96 μg/g)and on the floor (GM 1.12 μg/g of dust; 95%CI, 0.71-1.53 μg/g) indicated significant differences (P=0.005). However, the levels of Blot5 allergens found in bed (GM 19.00 μg/g of dust; 95%CI, 0.89-38.90 μg/g) and on the floor (GM 6.14 μg/g of dust; 95%CI, 0.40-11.90 μg/g) showed no statistically significant difference. In addition, in regards to the exposure index for Derpl and Blot5 allergens found in bed and on the floor, 17.6% in bed and 8.6% on the floor had levels of Blot5 ≥ 10 μg/g of dust, higher than those obtained for Derp1 (7.2% and 0% in bed and on the floor respectively, P< 0.05); higher percentages in bed and on the floor (25.0% and 35.7%) were observed for levels of Blot5 =0 μg/g of dust as compared with Derpl in bed and on the floor (4.3% and 14.5% respectively, P< 0.05).Conclusions Derpl and Blot5 are the major allergens found in this regional study, Blot5 is a more potent allergen in Hongkong, probably reflecting the high level of exposure to Blomia tropicalis (Bt). Bt and Dermatophagoides pteronyssinus (Dp) allergens should be included for precise diagnosis and effective immuno-therapeutic treatment of mite allergy in Hongkong.  相似文献   

20.
ANTI-PROLIFERATION EFFECT OF ORIDONIN ON HL-60 CELLS AND ITS MECHANISM   总被引:4,自引:0,他引:4  
Objcetive To investigate the anti-proliferation effect of oridonin on leukemic HL-60 cells and its mechanism. Methods HL-60 cells in vitro in culture medium were given different concentrations of oridonin. The inhibitory rate of cells were measured by microculture tetrazolium (MTT) assay, cell apoptotic rate was detected by flow cytometry (FCM), morphology of cell apoptosis was observed by hoechst 33258 fluorescence staining, and the activity of telomerase was detected using telomere repeat amplification protocol (TRAP) PCR-ELISA before and after apoptosis occurred. Results Oridonin could decrease telomerase activity, inhibit growth of HL-60 cells, and cause apoptosis significantly. The suppression was both in time- and dose-dependent manner. Marked morphological changes of cell apoptosis including condensation of chromatin and nuclear fragmentation were observed clearly by hoechst 33258 fluorescence staining especially after cells were treated 48-60 hours by oridonin. Conclusions Oridonin has apparent anti-proliferation and apoptotic effects on HL-60 cells in vitro, decreasing telomerase activity of HL-60 cells may be one of its most important mechanisms. These results will provide strong laboratory evidence of oridonin for clinical treatment of acute leukemia.  相似文献   

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