冈田酸诱导SH-SY5Y细胞后tau蛋白磷酸化与PTEN蛋白水平的变化

目的 研究冈田酸（OA）诱导SH—SY5Y细胞后tau蛋白磷酸化程度和PTEN蛋白水平的变化。方法 40nmol／LOA孵育SH—SY5Y细胞不同时间（0，3，6，12，18，24，36，48h）后，倒置显微镜观察细胞的形态变化，免疫印迹检测各时间点tau蛋白ser^396。位点的磷酸化及PTEN蛋白水平的变化。结果 SH-SY5Y细胞随OA孵育时间的延长逐渐出现细胞胞体变圆，突起回缩，悬浮生长至死亡。细胞tau蛋白ser^396位点磷酸化水平于OA作用3h开始升高，18h达高峰，随后逐渐下降，48h低于基础水平；而PTEN蛋白水平于OA作用3h开始升高，6h达高峰，之后逐渐下降，18h开始低于基础水平。结论 OA诱导SH—SY5Y细胞后tau蛋白磷酸化水平和PTEN蛋白水平均呈先上升后降低的变化趋势。     

褪黑素对冈田酸致NG108-15细胞tau蛋白过度磷酸化的影响

目的 观察褪黑素（MLT）对冈田酸（OA）致大鼠神经母细胞和小鼠胶质细胞瘤细胞的杂交细胞系NG108—15细胞tau蛋白过度磷酸化的影响。方法 10nmol／LOA作用于NG108—15细胞0，3，6，12及24h，免疫细胞化学法和免疫印迹法（Western blot法）观察Ser-396位点磷酸化tau蛋白，观察MLT对细胞内Ser-396位点磷酸化tau蛋白的影响。结果 在OA作用后3h增高，6h达到峰值，之后逐渐降低。选取OA作用6h，低、高剂量MLT较模型组Ser-396位点磷酸化tau蛋白水平明显降低。结论MLT 可对抗tau蛋白过度磷酸化，可能是其发挥抗阿尔茨海默病作用的机制之一。     

冈田酸诱导SH-SY5Y细胞后tau蛋白磷酸化与PTEN蛋白水平的变化

目的 研究冈田酸(OA)诱导SH-SY5Y细胞后tau蛋白磷酸化程度和PTEN蛋白水平的变化.方法 40nmol/L OA孵育SH-SY5Y细胞不同时间(0,3,6,12,18,24,36,48h)后,倒置显微镜观察细胞的形态变化,免疫印迹检测各时间点tau蛋白ser396位点的磷酸化及PTEN蛋白水平的变化.结果 SH-SY5Y细胞随OA孵育时间的延长逐渐出现细胞胞体变圆,突起回缩,悬浮生长至死亡.细胞tau蛋白ser396位点磷酸化水平于OA作用3h开始升高,18h达高峰,随后逐渐下降,48h低于基础水平;而PTEN蛋白水平于OA作用3h开始升高,6h达高峰,之后逐渐下降,18h开始低于基础水平.结论 OA诱导SH-SY5Y细胞后tau蛋白磷酸化水平和PTEN蛋白水平均呈先上升后降低的变化趋势.     

补体C3/C3aR 在冈田酸诱导的SH-SY5Y 细胞tau 蛋白过度磷酸化中的作用及机制研究

目的：tau 蛋白过度磷酸化与AD 的发病机制直接相关,本实验通过冈田酸（okadaic acid,OA）诱导SH-SY5Y 细胞tau 蛋白过度磷酸化实验模型,研究补体C3/C3aR 在 OA 致SH-SY5Y 细胞 tau 蛋白过度磷酸化的作用及机制。通过对以上问题的阐明将为研究针对C3/C3aR 这一靶点开发新型药物保护Tau 蛋白病尤其是 AD 提供理论基础。方法：①采用OA 诱导 SH-SY5Y 细胞,以 tau 蛋白过度磷酸化和细胞活性的变化为观察指标,建立用于 AD 研究的细胞模型。②CCK8 法检测细胞活性。③Western blot 测定 SH-SY5Y 细胞 Ser396,Thr231,Thr205 位点磷酸化tau 蛋白水平。④Western blot 法测定tau 蛋白磷酸化关键调节酶GSK3β,Cdk5,PP2A 的蛋白表达水平。结果：①OA 处理SH-SY5Y 细胞,CCK8 法检测显示细胞活性显著降低,作用随着 OA 浓度的增高和作用时间的延长而增强;40 nmol·L-1 OA 作用 SH-SY5Y 细胞不同时间,可见细胞中 Ser396,Thr231,Thr205 位点磷酸化 tau 蛋白水平于作用后 3 h 开始升高,24 h 达高峰,随后开始下降,48 h 的水平仍高于基础值,提示 OA 可诱导 SH-SY5Y 细胞活性下降和tau 蛋白过度磷酸化。②C3a 受体拮抗剂SB290157 预处理24 h 后,再给予40 nmol·L-1 OA 处理 SH-SY5Y 细胞12 h,检测细胞中Ser396,Thr231,Thr205 位点磷酸化 tau 蛋白水平,可见 Ser396,Thr231,Thr205 位点磷酸化tau 蛋白水平变化同时下调,显著低于单独OA 处理组,提示在 OA 诱导SH-SY5Y 细胞tau 蛋白过度磷酸化过程中,补体C3/C3aR 可能参与其中,C3aR 受体拮抗剂可显著抑制tau 蛋白过度磷酸化。③C3a 受体拮抗剂SB290157 预处理,western blot 结果显示SB290157 可抑制GSK-3β和Cdk5 的蛋白表达,但可上调蛋白磷酸酶PP-2A 的蛋白表达。④通过转染C3aR-siRNA,同样发现OA 不能诱导tau 的过度磷酸化及其调节蛋白的改变。结论：①OA 可诱导SH-SY5Y 细胞活性下降和tau 蛋白过度磷酸化,该细胞模型可用于AD 研究。②在OA 诱导SH-SY5Y 细胞tau 蛋白过度磷酸化过程中,C3a受体拮抗剂SB290157 可抑制SH-SY5Y 细胞 Ser396,Thr231,Thr205 位点磷酸化 tau 蛋白水平。③C3a 受体拮抗剂对过磷酸化tau 蛋白水平的抑制,可能是通过抑制GSK-3β 和 Cdk5 ,同时上调蛋白磷酸酶PP-2A 实现的。     

PTEN与磷酸化tau蛋白水平在Aβ25-35诱导的AD样细胞模型中的变化

目的探讨在Aβ25-35诱导人神经母细胞瘤细胞系SH-SY5Y细胞和大鼠胎鼠皮层原代神经元细胞的过程中PTEN及磷酸化tau(Ser396)蛋白水平的变化。方法 40μmol/L和20μmol/L的Aβ25-35分别孵育SH-SY5Y细胞和大鼠原代皮层神经元,孵育后的不同时间点MTT法测定细胞活力,显微镜观察细胞形态的变化,免疫印迹法观测各时间点磷酸化tau(Ser396)蛋白水平来确定模型的建立;同时观察在此过程中PTEN蛋白水平的变化。结果随着Aβ25-35孵育时间的延长,SH-SY5Y细胞和大鼠皮层原代神经元细胞逐渐出现细胞活力下降,细胞胞体变圆,突起回缩甚至消失,细胞间连接断裂等现象,细胞磷酸化tau(Ser396)蛋白水平于诱导后6h升高,12～24h达到高峰;PTEN蛋白水平于3h左右开始升高,6～12h达到高峰,后逐渐下降,48h甚至低于正常对照组,且PTEN蛋白水平的升高先于磷酸化tau(Ser396)蛋白水平的升高。结论 PTEN与Aβ25-35诱导的阿尔茨海默病样细胞模型的发生发展可能具有一定的关联性,在其发生早期PTEN表达可能增强。     

2,3-吲哚醌对人神经母细胞瘤SH-SY5Y细胞增殖影响

目的 研究2，3-吲哚醌对人神经母细胞瘤SH-SY5Y细胞增殖的影响。方法 应用MTT比色法观察2，3-吲哚醌对SH—SY5Y细胞生长的抑制情况；流式细胞仪检测2，3-吲哚醌对SH—SY5Y细胞周期分布的影响；Western blot方法观察其对细胞信号传导通路蛋白有丝分裂原激活的蛋白激酶（ERK）的作用，同时结合免疫细胞化学方法检测2，3-吲哚醌对SH-SY5Y细胞caspase-3的影响。结果 2，3-吲哚醌能显著抑制SH—SY5Y细胞增殖，并使细胞周期阻滞于G0／G1期；caspase-3蛋白表达随着药物浓度的升高逐渐增加；加2，3-吲哚醌后磷酸化的有丝分裂原激活的蛋白激酶蛋白（ERK1／2）相对表达量降低。结论 2，3-吲哚醌对SH—SY5Y细胞具有抗增殖作用，其作用可能是通过诱导细胞凋亡、改变细胞周期以及作用于细胞通路而实现的。     

冈田酸所致微管相关蛋白tau Ser396位点过度磷酸化抑制细胞凋亡

目的 探讨磷酸酯酶活性降低致tau过度磷酸化与细胞凋亡的关系.方法 稳定表达人tau eDNA的鼠成神经瘤细胞(N2a/tau)分3组.对照组不处理;十字孢碱(staurosporine,STP,凋亡诱导剂)处理6 h为STP组;用磷酸酯酶PP-2A的抑制剂冈田酸(okadaic acid,OA)处理4 h,再用STP处理6 h为OA+STP组.流式细胞仪检测凋亡率,免疫印迹检测Ser396位点磷酸化tau(Ser396 p-tau)和总tau的表达,免疫荧光双标检测Ser396 p-tau的阳性产物与活化的半胱氨酸蛋白酶(Cleaved Caspase-3)阳性产物的共定位关系.结果 ①凋亡率:对照组为(4.37±1.47)%,STP组为(19.84±3.25)%,OA+STP组为(12.12±2.46)%;②免疫印迹:OA+STP组Ser396 p-tau的表达明显高于另两组,总tau表达组间无差异;③免疫荧光双标:进一步印证免疫印迹结果,且3组均显示活化的Caspase-3和Ser396 p-tau阳性产物大多不共定位于相同细胞.结论 PP-2A活性降低所致tau Ser396位点的过度磷酸化可抑制细胞凋亡,结合前期的实验进一步明确特殊位点过度磷酸化的tau可抑制细胞进入快速的凋亡程序.     

雷公藤红素对β淀粉样蛋白诱导SH-SY5Y细胞Tau蛋白异常磷酸化影响的研究

观察雷公藤红素对β淀粉样蛋白142导致SHSY5Y细胞Tau蛋白异常磷酸化的影响及其可能机制。〖HT5”H〗方法〖HT5”K〗用 Aβ142刺激SHSY5Y细胞,建立Tau蛋白异常磷酸化的细胞模型。加入雷公藤红素干预,Western印迹法检测Aβ142导致Tau蛋白异常磷酸化的变化,同时检测pNFSymbolkA@B表达情况;siRNA法下调TLR4表达,Western印迹法检测Aβ142导致的Tau蛋白异常磷酸化的变化情况;同时观察下调TLR4后,Aβ142刺激对pNFSymbolkA@B表达的影响。〖HT5”H〗结果〖HT5”K〗Aβ142刺激SHSY5Y细胞,导致Tau蛋白Ser199/202、Ser396位点的磷酸化水平增加,而雷公藤红素能降低这两个位点磷酸化水平;Aβ142导致细胞pNFSymbolkA@B表达增加,雷公藤红素干预后pNF SymbolkA@B表达下调,且NFSymbolkA@B抑制剂BAY117082同样能降低Aβ142导致的Tau蛋白异常磷酸化;下调TLR4表达,Aβ142导致的Tau蛋白异常磷酸化及pNFSymbolkA@B表达下降。〖HT5”H〗结论〖HT5”K〗雷公藤红素降低Aβ142导致的SH SY5Y细胞Tau蛋白异常磷酸化可能与其抑制TLR4/NF SymbolkA@ B通路活性有关。     

反义P38cDNA抑制胆红素诱导的人神经母细胞瘤SH-SY5Y细胞凋亡

目的：观察P38激酶特异性抑制剂SB203580及反义P38激酶cDNA对胆红素诱导人神经母细囊瘤SH—SY5Y细胞凋亡的影响，探讨P38激酶信号转导通路在胆红素诱导SH—SY5Y细胞凋亡中的作用及高胆红素血症损伤中枢神经细胞的分子机制及在听神经病发病中的作用。方法：分别经SB203580和二甲基亚砜预处理的SH—SY5Y细胞在胆红素作用3h和5h后，在倒置光显微镜下观察细胞形态的变化及存活情况；流式细胞仪检测凋亡细胞百分率。采用脂质体法将含人反义P38cDNA的真核表达载体pcDNA3．O—antiP38导入SH—SYSY细胞，建立稳定低表达P38蛋白的细胞株，经Western blotting鉴定后命名为SH—SY5Y—antiP38，而转染空载体pcDNA3．0的SH—SY5Y细胞株则命名为SH—SY5Y细胞株则命名为SH-SY5Y-pcDNA3.0；SH—SY5Y—antiP38细胞和SH-SY5Y-pcDNA3．O细胞分别经胆红素作用3h和5h，在倒置光显微镜下观察细胞形态的变化及存活情况；流式细胞仪检测凋亡细胞百分率。结果：在0．02g／L胆红素作用下，流式细胞仪显示与对照相比SB203580预处理的SH—SY5Y细胞在胆红素作用3h后，其凋亡细胞由19．4％下降到12．1％，5h后由66．2％降到39．3％；与SH—SY5Y—pcDNA3．O细胞相比，SH—SY5Y—antiP38细胞在胆红素作用3h后，其凋亡细胞由11．1％降到8．02％，5h后67％降到23．4％。结论：P38激酶参与了胆红素诱导SH—SY5Y细胞凋亡的信号转导。P38通路的激活可能在高胆红素血症导致的神经细胞损伤中(包括听神经细胞)发挥重要作用。     

Aβ25-35诱导SH-SY5Y细胞Tau蛋白的过度磷酸化

目的采用凝聚态Aβ25-35建立Tau蛋白过度磷酸化的阿尔茨海默病样细胞模型.方法用不同浓度凝聚态Aβ25-35作用于SH-SY5Y细胞24,48,72 h,通过四唑盐比色法及乳酸脱氢酶释放实验观察Aβ25-35对SH-SY5Y细胞存活的影响;选择对细胞存活影响较小的适宜浓度和作用时间处理细胞,以蛋白免疫印迹法研究Tau蛋白磷酸化水平的变化.结果 Aβ25-35对细胞存活的影响随着作用时间的延长或作用浓度的增加而增强.10 μmol/L Aβ25-35作用于细胞72 h或20 μmol/L Aβ25-35作用于细胞≥48 h时,MTT代谢率均明显下降(P<0.05).Aβ25-35作用浓度>20 μmol/L作用72 h后,细胞LDH释放率明显增加(P<0.05).选用细胞毒性小的10,20 μmol/L Aβ25-35作用于SH-SY5Y细胞0,3,6,12,24,48 h,蛋白免疫印迹示Tau蛋白在Ser396、Ser199/202位点的磷酸化水平增高,于6 h达到最高峰.结论凝聚态Aβ25-35可诱导建立阿尔茨海默病样Tau蛋白过度磷酸化的细胞模型.     

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…     

Value of D-Dimers in patients with acute aortic dissection

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.     

Research of combined liver-kidney transplantation model in rats

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.     

Development on Adjustable Calibration Marker for Shock Wave Focus

Shock wave lithotripsy （SWL） is a treatment of choice for upper urinary stones. However, this procedure is inappropriate for obese patients because the focus is often unable to reach the target owing to the limited focal distance in shock wave source. Although treating such patients in a blast path may increase the application length of shock wave source, it＇s difficult to find this path on the lithotripter monitor. For this reason, we invented an adjustable calibration marker in order to set an effective focus in the shock wave hath.     

Study on the TCM Pathogenesis of Angiopathic Complications of Type II Diabetes

Excess production of reactive oxygen species(ROS)of mitochondrion mediated by hyperglycemia is the common pathogenesis of angiopathic complications of diabetes.TCM holds that the damp from the dysfunction of spleen.kidney and liver is the causative factor of complications of diabetes.This is similar to the mechanism of Ros resulting in angiopathic complications of diabetes.When the angiopathic complications of type II diabetes mellitus(T2DM)are difierentiated as caused by turbid damp in TCM can be explained as ROS.Since the obstruction of pathogenic damp in channels and collaterals is said to be the main pathogenesis,the treating principle should be dissolving the damp to remove the obstruction.     

Leptin, Ghrelin and TNF-α before and after Hypo-caloric Traditional Chinese Diet and Auricular Acupuncture

INTRODUCTION Obesity is a complex emergent problem, which can be possibly solved not only by the diet but also by the life style and promotion of a constant physical exercise. 1, 2 No doubt careful attentions must be given to the nutritional condition of obese people, the dietary habits, the somatic build (i.e. distribution of fat mass) and the organic functions linked to formation of the fat mass. All the parameters should be constantly monitored before, during and after a diet treatment. 3, 4, 5     

BLOOD PRESSURE CHANGE WITH AGE IN SALT-SENSITIVE TEENAGERS

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.     

Relationship between Dysglycemia and Carotid Atherosclerosis in Tibetan Population

People with dysglycemia are at high risk for atherosclerotic diseases. This study aims at investigating the atherosclerotic vascular damage in dysglycemia and its metabolic origin in Tibetan population.     

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.     

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.