三种不同培养条件下兔关节软骨细胞增殖速率的差别

目的:观察兔关节软骨细胞在3种不同培养条件下增殖速率的差别。方法:实验于2004-10/2006-10在上海长海医院血管外科实验室完成。取4周龄新西兰雄性兔6只,利用机械与酶消化的方法获得均一性兔关节软骨细胞,在单层培养条件下增殖至第2代;将第2代细胞在高密度条件下转入藻酸盐凝胶24孔培养介质,进行三维培养。根据实验目的,将24孔培养板分为单层培养对照组,三维培养组(培养液不含胰岛素样生长因子Ⅰ),含胰岛素样生长因子Ⅰ三维培养组(培养液含50μg/L胰岛素样生长因子Ⅰ),每日在倒置相差显微镜下观察软骨细胞的生长状态,进行细胞计数1周,绘制细胞生长曲线,比较3种不同培养条件下兔关节软骨细胞的增殖速率。结果:①单层培养细胞24h后开始贴壁生长,细胞形态呈多角形、星形、圆形,72h后,倒置显微镜下观察细胞增殖数目明显增多,细胞增殖呈片状;三维培养条件下的软骨细胞,刚接种至藻酸钠凝胶时,细胞形态为圆形;三维培养(含胰岛素样生长因子Ⅰ)体系中的软骨细胞从转入三维培养的第3天起,发现细胞出现分裂增殖,局部有细胞团族形成,第7天,凝胶中开始有大量的细胞团族形成。在培养过程中,三维培养软骨细胞形态始终保持圆形。②1周内,单层培养条件下的软骨细胞增殖速率要明显高于三维培养组;含胰岛素样生长因子Ⅰ三维培养组的软骨细胞增殖速率明显高于不含胰岛素样生长因子Ⅰ三维培养组。结论:单层培养方法短期内(1周)对软骨细胞的增殖作用要优于三维培养方法;胰岛素样生长因子Ⅰ对三维培养条件下关节软骨细胞具有促进增殖的作用。     

SD大鼠骨代谢与非洛地平的干预

目的:流行病学报告提示治疗心血管疾病的重要药物钙离子通道阻滞剂影响骨代谢,但这种作用与药物的剂量和作用时间的关系还不明确。观察不同剂量、不同时间的非洛地平对SD大鼠骨代谢的作用。方法:实验于2006-05/2007-04在西安交通大学动物实验中心完成。①实验分组:132只6月龄SD大鼠,随机分为4组:正常对照组,非洛地平0.15,0.45,1.25mg/kg组,每组33只。②实验方法:非洛地平各组每天按0.15mg/kg,0.45mg/kg,1.25mg/kg剂量灌胃非洛地平,正常对照组灌服等量生理盐水。③实验评估:在实验第3,4,5,6个月时每组随机取8只大鼠,对其股骨和腰椎骨密度、身长、体质量和血清游离钙、磷、碱性磷酸酶、胆固醇、三酰甘油含量进行测定。结果:进入结果分析的SD大鼠共有127只,正常对照组31只,非洛地平0.15mg/kg组31只,非洛地平0.45mg/kg组32只,非洛地平1.25mg/kg组33只。①各组大鼠股骨和腰椎骨密度:给药后6个月后非洛地平0.15mg/kg组大鼠的股骨和腰椎骨密度高于正常对照组(P<0.01),非洛地平0.45,1.25mg/kg组股骨骨密度则出现降低(P<0.01),非洛地平1.25mg/kg组降低更明显(P<0.01)。②各组大鼠的身长、体质量和血清钙、磷、碱性磷酸酶、胆固醇、三酰甘油含量:不同剂量的非洛地平组与正常对照组比较,差异无显著性。各时间段比较差异也无显著性。结论:①短期应用非洛地平对大鼠骨代谢无明显影响。②长期应用低剂量非洛地平对大鼠股骨和腰椎的骨代谢产生正性的影响。③长期应用中、高剂量非洛地平对大鼠负重骨(股骨)的骨代谢产生负性的影响,剂量越大,负性影响越明显;而对非负重骨(腰椎)的骨代谢无明显影响。     

关节软骨破坏与尿激酶型纤溶酶原激活物的影响

目的:观察尿激酶型纤溶酶原激活物对关节软骨的影响。方法:实验于2004-06/2006-06在石河子大学医学院第一附属医院中心实验室完成。实验选择健康新西兰大白兔42只并随机分为实验组24只,对照组18只;实验组在右膝关节内注射2mg/L尿激酶型纤溶酶原激活物0.2mL,对照组注射等容量的生理盐水;注射完成后在4,8,12周分批取兔软骨进行组织学观察、电镜检查及葡糖氨基聚糖测定。关节软骨退行性变采用组织-组化评分标准评定。结果:42只新西兰大白兔全部进入结果分析。①两组兔关节软骨大体观察结果:随着时间的延长,实验组关节软骨退行性变程度加重;对照组兔关节软骨外观无裂纹及软化。②两组兔关节软骨光镜组织学检查及评分:实验组关节软骨表面不规则、缺损及结构破坏,软骨细胞数目减少、纤维增生及异染性降低,潮线的完整性破坏;对照组关节软骨4层结构清晰可辨,软骨细胞排列整齐,潮线完整。③两组兔关节软骨超微结构观察结果:透射电镜下实验组软骨细胞明显固缩且外形不规则,胞膜上突起消失,核膜不完整,内质网扩张明显,细胞器数目减少,可见脂滴存在,细胞坏死、溶解多见;对照组可见软骨细胞呈椭圆形,细胞及胞膜完整,表面可见均匀分布的微绒毛突起,胞浆内可见丰富细胞器,基质中胶原纤维排列整齐。④两组关节软骨退行性变Mankin’s评分结果:实验组4,8,12周评分均高于对照组(P均<0.05)。⑤两组样品中葡糖氨基聚糖测定结果:对照组蛋白多糖含量无明显变化,实验组在不同时期的葡糖氨基聚糖含量均低于对照组(P<0.05);实验组8周时低于4周(P<0.01),12周虽低于8周,但差异无显著性。表明在不同时期,葡糖氨基聚糖的总含量均减少,这些改变随时间的延长而渐不明显。结论:尿激酶型纤溶酶原激活物能降解关节软骨蛋白多糖,进而导致关节软骨的破坏,尿激酶型纤溶酶原激活物可能是导致关节软骨破坏的相关因素之一。     

Engagement of the adhesion receptor CD22 triggers a potent stimulatory signal for B cells and blocking CD22/CD22L interactions impairs T-cell proliferation

The B-lymphocyte-restricted adhesion protein CD22 mediates sialic acid- dependent cell-cell interactions. Engagement of CD22 on B lymphocytes with a CD22 monoclonal antibody (MoAb) HB22.7 that blocks the binding of CD22 to its ligand(s) directly stimulated B-cell proliferation. In addition, the HB22.7 MoAb costimulated B-cell proliferation with either anti-IgM, interleukin-2 (IL-2), IL-4, or CD40 and triggered predominantly B-cell IgG secretion with IL-2. Even more striking levels of B-cell proliferation occurred with HB22.7 MoAb under culture conditions that enhanced B-B-cell interactions. In contrast, a nonblocking CD22 MoAb (CD22.5) poorly costimulated in similar experiments. The functional differences between the two antibodies likely result from differing abilities to trigger downstream signaling events as significant differences in CD22 tyrosine phosphorylation and the recruitment of the tyrosine kinase p53/56lyn and the tyrosine phosphatase SH-PTP1C were found. Besides their role in B-cell stimulation, CD22/CD22L interactions may also assist in regulating T- cell proliferation because inhibition of CD22-CD22L engagement with the HB22.7 MoAb impaired T-cell proliferation in a costimulatory assay. Thus, CD22/CD22L interactions result in stimulatory signals for both B and T lymphocytes.     

Mitochondrial function in diaphragm of emphysematous hamsters after treatment with nandrolone

Respiratory failure in patients with COPD may be caused by insufficient force production or insufficient endurance capacity of the respiratory muscles. Anabolic steroids may improve respiratory muscle function in COPD. The effect of anabolic steroids on mitochondrial function in the diaphragm in emphysema is unknown. In an emphysematous male hamster model, we investigated whether administration of the anabolic steroid nandrolone decanoate (ND) altered the activity of mitochondrial respiratory chain complexes in the diaphragm. The bodyweight of hamsters treated with ND was decreased after treatment compared with initial values, and serum testosterone levels were significantly lower in hamsters treated with ND than in control hamsters. No difference in the activity of mitochondrial respiratory chain complexes in the diaphragm between normal and emphysematous hamsters was observed. Treatment with ND did not change the activity of mitochondrial respiratory chain complexes in the diaphragm of both normal and emphysematous hamsters. In emphysematous hamsters, administration of ND decreased the activity of succinate:cytochrome c oxidoreductase compared with ND treatment in normal hamsters. We conclude that anabolic steroids have negative effects on the activity of succinate:cytochrome c oxidoreductase and anabolic status in this emphysematous hamster model.     

Preclinical assessment of purging with VP-16-213: key role for long- term marrow cultures

An evaluation of the effects of VP-16 on normal human marrow cells and representative lymphoma-leukemia cell lines was performed to assess this agent's applicability to ex vivo marrow purging. Tumoricidal dose curves were defined using malignant lymphoid (SK-DHL2 and Reh) and myeloid (HL-60) cells admixed with a 20-fold excess of irradiated marrow cells to simulate a borderline remission marrow. One-hour treatments yielded ID50 of less than 5 mumol/L of VP-16 for clonogenic units from each cell line; rare-to-zero clonogenic units survived exposure to 50 to 100 mumol/L. CFU-Mix, BFU-E, and CFU-GM were equal in their sensitivity to VP-16 (ID50s25 to 30 mumol/L). Marrows treated with 75 mumol/L were completely depleted of these colony-forming cells but produced CFU-GM in one-stage long-term marrow cultures (LTMCs). This dose had little adverse effect on the proliferative capacity of marrow stromal progenitors, as measured by CFU-F (ID50 271 mumol/L) and by the unperturbed development of adherent layers in LTMCs. Furthermore, these stromal layers were able to support hematopoiesis as well as controls in co-culture experiments with autologous marrow cells (two-stage LTMCs). In conclusion, doses of VP-16 that cleanse marrow of lymphoma-leukemia cells spare hematopoietic and stromal progenitors as demonstrated by LTMCs. These data favor the use of VP-16 in the clinical autotransplant setting.     

腹腔镜在直肠癌手术治疗中的临床应用

目的:探讨腹腔镜手术治疗直肠癌的方法和可行性．方法:2005-02／12共开展腹腔镜直肠癌手术31 例,其中Dixon手术20例,Miles手术9例,乙状结肠袢式造口术2例．结果:手术均获成功,无中转开腹,手术时间平均200 min,术中平均出血量80 mL,术后肠功能恢复时间平均30 h,术后1例Dixon手术患者出现吻合口瘘,经保守治疗痊愈．结论:腹腔镜直肠癌手术具有创伤小、手术出血少、术后痛苦小、肠道功能恢复快等特点,是可行的、安全有效的手术方法．     

Interleukin-2 induces activation of coagulation and fibrinolysis: resemblance to the changes seen during experimental endotoxaemia

The administration of Interleukin-2 (IL-2) causes the release or generation of other cytokines such as tumour necrosis factor (TNF) which, by disturbing the anticoagulant properties of the endothelium, may induce a procoagulant state in patients receiving this drug. We therefore evaluated the effects of IL-2 on coagulation and fibrinolysis in 14 patients receiving 12 or 18 x 10(6) IU/m2/d of IL-2 given as a 15 min infusion for 5 d. Blood samples were drawn at short intervals after the first IL-2 infusion. The parameters were analysed by way of analysis for repeated measures (F tests rather than t tests). During the first day, thrombin-antithrombin (TAT) complexes started to increase 2 h after the IL-2 infusion, reaching peak levels at 4 h (n = 14; 11.2 +/- 6.4 micrograms/l v 49.8 +/- 49.2 micrograms/l, P < 0.01). Plasma alpha 2 antiplasmin (PAP) complexes showed a similar pattern rising from a mean baseline value of 17.5 +/- 7.6 nmol/l to 66.8 +/- 47.7 nmol at 4 h (P < 0.01). In four patients the peak of PAP preceeded that of TAT. Tissue plasminogen activator (tPA) rose from a mean baseline value of 4.9 +/- 3.7 micrograms/l to 26.3 +/- 13.5 micrograms/l at 4 h (P < 0.01). Plasminogen-activator-inhibitor-1 (PAI-1) levels increased from 59 +/- 35 micrograms/l to 113 +/- 39 micrograms/l at 6 h (P < 0.01). tPA PAI-1 complexes increased from 0.15 +/- 0.07 to 0.69 +/- 0.21 nmol/l at 6 h (P < 0.01). Our study indicates that IL-2 activates the coagulation and fibrinolytic systems in vivo. The changes resemble the perturbations observed after endotoxin/TNF administration. These abnormalities may play a role in the side-effects induced by IL-2 therapy.