信号传导通路抑制剂PD98059和LY294002对子宫内膜癌细胞和裸鼠移植瘤的抑制作用

目的 观察有丝分裂原活化蛋白激酶(MAPK)信号传导通路抑制剂PD98059和磷脂酰肌醇3激酶(PI3K)信号传导通路抑制剂LY294002对子宫内膜癌细胞株Ishikawa细胞和子宫内膜癌裸鼠移植瘤的抑制作用.方法 (1)体外实验:实验分为4组,即PD组(加入浓度分别为0、1、50、100 μmol/L的PD98059)、LY组(加入浓度分别为0、1、50、100 μmol/L的LY294002)、PD+LY组(加入浓度均为50 μmol/L的PD98059和LY294002)、对照组[仅加入二甲基亚砜(DMSO)],分别培养24、48和72 h.应用四甲基偶氮唑蓝(MTT)比色法检测各组Ishikawa细胞的增殖情况,流式细胞仪检测各组Ishikawa细胞的细胞周期比例和细胞凋亡率.(2)体内实验:建立子宫内膜癌裸鼠移植瘤模型,随机分为4组(每组6只):PD组(注射PD98059 50 mg/kg)、LY组(注射LY294002 50 mg/kg)、PD+LY组(注射PD98059 50 mg/kg和LY294002 50 mg/kg)、对照组(注射等量的生理盐水),每周2次,共3周;观察移植瘤的生长情况,并计算抑瘤率;用末端脱氧核苷酸转移酶介导的脱氧尿苷三磷酸标记(TUNEL)法检测移植瘤组织的细胞凋亡情况,以细胞凋亡指数表示;免疫组化法检测移植瘤组织内磷酸化细胞外信号调节激酶(p-ERK)和磷酸化蛋白激酶B(p-Akt)蛋白的表达.结果 (1)PD98059和(或)LY294002处理后,PD组、LY组Ishikawa细胞增殖的抑制作用均呈明显的时间和浓度依赖性(P＜0.05),且PD+LY组细胞增殖的这种抑制作用明显高于PD组、LY组(P＜0.05).LY组细胞S期、G0/G1期比例的变化呈明显的时间和浓度依赖性(P＜0.05);PD组细胞各期比例的变化均无时间依赖性(P＞0.05),但G0/G1期、S期比例的变化呈明显的浓度依赖性(P＜0.05);PD+LY组与PD组、LY组比较,G0/G1期比例明显增加、S期比例明显减少(P＜0.05).PD+LY组细胞凋亡率为(63.3±0.5)%,明显高于PD组、LY组[分别为(30.7±20.1)%和(40.8±1.3)%,P＜0.01].(2)注射PD98059和(或)LY294002后,随着时间的延长,PD组、LY组、PD+LY组裸鼠移植瘤体积增长相对缓慢,对照组移植瘤体积增长明显,PD组、LY组、PD+LY组分别与对照组比较,差异均有统计学意义(P＜0.05);PD+LY组分别与PD组、LY组比较,差异也均有统计学意义(P＜0.05),而PD组与LY组比较,差异无统计学意义(P＞0.05).LY组、PD组、PD+LY组抑瘤率分别为(32±16)%、(38±17)%、(68±9)%,PD+LY组明显高于LY组、PD组(P＜0.05).LY组、PD组、PD+LY组细胞凋亡指数分别为(13.7±1.5)%、(14.1±1.2)%、(29.0±1.8)%,PD+LY组明显高于LY组、PD组(P＜0.01).LY组、PD组、LY+PD组裸鼠移植瘤组织中p-ERK和p-Akt蛋白的表达强度均明显弱于对照组.结论 信号传导通路抑制剂PD98059、LY294002能够抑制体外及体内子宫内膜癌细胞的生长,并促进其凋亡.

Abstract：

Objective To investigate the effects of signal pathway inhibitors PD98059 and LY294002 on cell proliferation, apoptosis, expressions of phosphorylated extracellular signal-regulared kinase (p-ERK) and phosphorylated protein kinase B ( p-Akt) in endometrial carcinoma xenografts. Methods Human endometrial carcinoma Ishikawa cells were cultured in vitro. The effects of PD98059 and LY294002 on proliferation, apoptosis, and cell cycle distribution of endometrial cancer cells were detected by monotetrazolium ( MTT) assay and fluorescence-activated cell sorting technique. The models of xenografted tumor were established by the subcutaneous inoculation in 24 nude mice, and then they were randomly divided into 4 groups ( n = 6) , normal saline group, PD98059 group (PD group) , LY294002 group ( LY group) or PD98059 + LY294002 group ( PD + LY group) by intraperitoneal injections, respectively. The anti-tumor efficacy was evaluated by measuring tumor volume and tumor growth status. The histopathological change of tumor specimens was observed using HE staining and terminal deoxynucleotidyl transferasemediated dUTP-digoxigen in nick and labeling method (TUNEL) testing and the expression levels of p-ERK and p-Akt were detected by immunohistochemistry method. Results ( 1) The proliferation of Ishikawa cells were suppressed after treated by PD98059 and ( or) Y294002, in which A570 values of cells decreased showing both time-dependent and concentration-dependent manner ( LY294002: Fgroup = 9. 801, P = 0. 002; Ftime = 10. 398, P = 0. 001. PD98059: Fgroup= 8. 213, P = 0. 015; Ftime = 6. 839, P = 0. 036). Cell cycle distribution analysis revealed that percentage of Ishikawa cells at G0/G1 phase(Ftime =35.049, P= 0.004; Fgroup = 32. 024, P ＜0. 01) increased and percentage of S phase cells (Ftime = 7. 789, P = 0. 049; Fgroup = 30. 132, P ＜0. 01) decreased significantly. The percentage of apoptotic cells increased significantly among PD group, LY group and PD + LY group, in which there were significant difference [(63. 3 ±0.5)% vs (30. 7 ± 20. 1) % vs(40. 8 ± 1. 3) % ; F = 621. 059, P ＜ 0. 01]. (2) Compared with the control group, the increasing of transplanting tumor volume in the treated groups were obviously ( F = 23. 545 , P ＜ 0. 01) , and the inhibited rate of the tumor was higher in PD + LY group than that in PD group or LY group [(68 ± 9 ) % vs ( 32 ± 16 ) % or ( 38 ± 17 ) % ; F = 10. 283 , P ＜ 0. 05]. ( 3 ) HE staining shown that there were different degrees of necrosis for endometrial carcinoma cell in different groups. The apoptosis of tumor cells were significantly increased in treated groups by TUNEL testing [(13. 7 ± 1. 5)% , ( 14. 1 ± 1. 2)% , (29. 0 ± 1. 8 ) % ; F = 320. 344, P ＜ 0. 01]. Immunohistochemistry results demonstrated that the expressions of p-ERK and p-Akt in treated groups were lower than that in control group, of which LY + PD group was the lowest one. Conclusion The signal pathway inhibitors PD98059 and LY294002 could inhibit the growth of human endometrial carcinoma in vivo and in vitro, in which may induce cell apoptosis.     

高胆红素血症对新生儿T淋巴细胞亚群、血清可溶性白细胞介素-2受体变化趋势的影响和意义

目的 探讨新生儿高胆红素血症(简称高胆)时T淋巴细胞亚群和血清可溶性白细胞介素-2受体(soluble interleukin-2 receptor,sIL-2R)水平的变化趋势及其临床意义.方法 选择2006年12月1日至2007年1月31日住院的31例高胆新生儿作为高胆组,再根据黄疸程度分为重度黄疸组和轻度黄疸组;将其中16例随访病例按照病程分为黄疸高峰期与黄疸恢复期.选取同期与高胆组日龄相匹配的32例健康足月新生儿(无黄疸或血清胆红素水平≤204.0 μmol/L)作为与高胆组相对应的对照组(对照组Ⅰ);选取同期与黄疸恢复期病例日龄相匹配的26例健康足月新生儿(日龄＞7 d)作为与随访病例相对应的对照组(对照组Ⅱ).采用方差分析及两两检验比较各组血清胆红素、T淋巴细胞亚群、sIL-2R水平,并分析其间的相关性.结果 高胆组新生儿的CD3、CD4、CD4/CD8比值分别为(54.0±5.1)%、(26.8±5.0)%和0.8±0.1,较对照组Ⅰ[(62.0±4.7)%、(43.0±4.7)%和1.4±0.2]降低(P＜0.01);而黄疸恢复期较黄疸高峰期增高[(62.4±3.3)%和(55.1±4.2)%、(43.6±2.5)%和(26.1±4.4)%、1.4±0.1和0.8±0.1](P＜0.01);黄疸高峰期血清sIL-2R水平[(319.4±185.2)kU/L]高于黄疸恢复期[(129.7±99.3)kU/L]和对照组Ⅱ[(171.9±102.2)kU/L](P＜0.01).总体的血清胆红素水平与CD4/CD8比值呈负相关(r=-0.99,P＜0.01),与sIL-2R水平呈正相关(r=0.95,P＜0.05),sIL-2R水平与CD4/CD8比值呈负相关(r=-0.92,P＜0.05).结论 新生儿高胆时存在细胞免疫功能抑制状态,该抑制状态有随着黄疸消退而逐渐减轻的趋势.

Abstract：

Objective To investigate the dynamic changes and the clinical significance of T-cell subsets and serum soluble interleukin-2 receptor (sIL-2R)in neonates with hyperbilirubinemia.Methods Thirty-one neonates with hyperbilirubinemia, admitted to the hospital from Decembr 1,2006 to January 31, 2007, were enrolled and divided into two subgroups: severe jaundice group and mild jaundice group according to the bilirubin level. Thirty-two age-mached healty newborns were as controls(control group Ⅰ). The T-cell subsets and sIL-2R of peripheral venous blood samples from these neonates were measured and compared. Sixteen of these 31 neonates with hyperbilirubinemiawere followed up and another twenty-six age-mached healty newborns were as controls(control group Ⅱ ). The level of serum bilirubin in convalescence of sixteen hyperbilirubinemia neonates and control group Ⅱ were tested and analyzed also. Results The levels of CD3, CD4, CD4/CD8 in the neonates with hyperbilirubinemia were lower compared with those of control group Ⅰ [(54.0±5.1)% vs (62.0±4.7)%, (26.8±5.0)% vs (43.0±4.7)%, 0.8±0.1 vs 1.4±0.2] (P＜0.01), but was higher in convalescence than in peak phase[ (62.4±3.3)% vs (55.1±4.2)%, (43.6±2.5)% vs (26.1±4.4)%, 1.4 ± 0.1 vs 0.8±0.1] (P＜0.01). The peak level of sIL-2R in the hyperbilirubinemia group was (319.4± 185.2) kU/L, higher than that in the convalescence [(129.7±99.3) kU/L] and in the control group Ⅱ [(171.9±102.2) kU/L] (P＜0.01). The serum bilirubin level showed negative correlation with CD4/CD8 ( r = -0.99, P ＜ 0.01 ) and positive correlation with sIL-2R (r=0.95, P＜0.05). The sIL-2R level was negatively correlated with CD4/CD8 (r=-0.92, P＜0.05). Conclusions Neonates, when suffering from hyperbilirubinemia, are immunosuppressed which may recover with the alleviation of jaundice.     

妊娠期糖尿病母鼠血清晚期糖基化终产物与子鼠心脏发育异常的关系

Objective To explore the effect of advanced glycation end product(AGE) in serum of maternal rats with gestational diabetes mellitus (GDM) on the heart development of their offsprings. Methods Fifty-four SD rats were randomly assigned into control group (n= 24) and GDM group (n=30) which were established by administration of streptozotocin intra-abdominally. On the gestational age of 13, 16, 19 days, all rats underwent hysterectomy to obtain the fetal heart tissues. Serum level of AGE and blood glucose level of maternal rats were tested. The expression of receptor AGE (RAGE) in fetal cardiac tissue were detected by immunohistoehemistry. Results The incidence of fetal heart defect in GDM group was significantly higher than the control group at each time point (P<0.01). Rats in GDM group had higher blood glucose level at each time point (P<0.01). The AGE levels of GDM group on gestational age of 13, 16 and 19 day [(5.72±0.68) U/mgpr, (7.31±0.29) U/mgpr and (7.77±0.39) U/mgpr] were significantly higher than those of the control group [(4.45±0.27) U/mgpr, (4.71±0. 35) U/mgpr and (4. 37±0. 44) U/rngpr] (t=6. 142, 16. 295, 0. 399,P<0. 01). The number of heart malformation in fetal rats (r=0.994,P=0. 000) and blood glucose (r=0. 717,P=0. 000) had the positive relationship with the maternal serum AGE level. The expression of RAGE in fetal heart was positively related with the number of fetal heart malformation (r= 0. 638,P= 0. 004). Conclusions The increased maternal serum AGE level in GDM rats may be an important factor in fetal heart dysplasia.     

妊娠期糖尿病母鼠血清晚期糖基化终产物与子鼠心脏发育异常的关系

Objective To explore the effect of advanced glycation end product(AGE) in serum of maternal rats with gestational diabetes mellitus (GDM) on the heart development of their offsprings. Methods Fifty-four SD rats were randomly assigned into control group (n= 24) and GDM group (n=30) which were established by administration of streptozotocin intra-abdominally. On the gestational age of 13, 16, 19 days, all rats underwent hysterectomy to obtain the fetal heart tissues. Serum level of AGE and blood glucose level of maternal rats were tested. The expression of receptor AGE (RAGE) in fetal cardiac tissue were detected by immunohistoehemistry. Results The incidence of fetal heart defect in GDM group was significantly higher than the control group at each time point (P<0.01). Rats in GDM group had higher blood glucose level at each time point (P<0.01). The AGE levels of GDM group on gestational age of 13, 16 and 19 day [(5.72±0.68) U/mgpr, (7.31±0.29) U/mgpr and (7.77±0.39) U/mgpr] were significantly higher than those of the control group [(4.45±0.27) U/mgpr, (4.71±0. 35) U/mgpr and (4. 37±0. 44) U/rngpr] (t=6. 142, 16. 295, 0. 399,P<0. 01). The number of heart malformation in fetal rats (r=0.994,P=0. 000) and blood glucose (r=0. 717,P=0. 000) had the positive relationship with the maternal serum AGE level. The expression of RAGE in fetal heart was positively related with the number of fetal heart malformation (r= 0. 638,P= 0. 004). Conclusions The increased maternal serum AGE level in GDM rats may be an important factor in fetal heart dysplasia.     

妊娠期糖尿病母鼠血清晚期糖基化终产物与子鼠心脏发育异常的关系

Objective To explore the effect of advanced glycation end product(AGE) in serum of maternal rats with gestational diabetes mellitus (GDM) on the heart development of their offsprings. Methods Fifty-four SD rats were randomly assigned into control group (n= 24) and GDM group (n=30) which were established by administration of streptozotocin intra-abdominally. On the gestational age of 13, 16, 19 days, all rats underwent hysterectomy to obtain the fetal heart tissues. Serum level of AGE and blood glucose level of maternal rats were tested. The expression of receptor AGE (RAGE) in fetal cardiac tissue were detected by immunohistoehemistry. Results The incidence of fetal heart defect in GDM group was significantly higher than the control group at each time point (P<0.01). Rats in GDM group had higher blood glucose level at each time point (P<0.01). The AGE levels of GDM group on gestational age of 13, 16 and 19 day [(5.72±0.68) U/mgpr, (7.31±0.29) U/mgpr and (7.77±0.39) U/mgpr] were significantly higher than those of the control group [(4.45±0.27) U/mgpr, (4.71±0. 35) U/mgpr and (4. 37±0. 44) U/rngpr] (t=6. 142, 16. 295, 0. 399,P<0. 01). The number of heart malformation in fetal rats (r=0.994,P=0. 000) and blood glucose (r=0. 717,P=0. 000) had the positive relationship with the maternal serum AGE level. The expression of RAGE in fetal heart was positively related with the number of fetal heart malformation (r= 0. 638,P= 0. 004). Conclusions The increased maternal serum AGE level in GDM rats may be an important factor in fetal heart dysplasia.     

妊娠期糖尿病母鼠血清晚期糖基化终产物与子鼠心脏发育异常的关系

Objective To explore the effect of advanced glycation end product(AGE) in serum of maternal rats with gestational diabetes mellitus (GDM) on the heart development of their offsprings. Methods Fifty-four SD rats were randomly assigned into control group (n= 24) and GDM group (n=30) which were established by administration of streptozotocin intra-abdominally. On the gestational age of 13, 16, 19 days, all rats underwent hysterectomy to obtain the fetal heart tissues. Serum level of AGE and blood glucose level of maternal rats were tested. The expression of receptor AGE (RAGE) in fetal cardiac tissue were detected by immunohistoehemistry. Results The incidence of fetal heart defect in GDM group was significantly higher than the control group at each time point (P<0.01). Rats in GDM group had higher blood glucose level at each time point (P<0.01). The AGE levels of GDM group on gestational age of 13, 16 and 19 day [(5.72±0.68) U/mgpr, (7.31±0.29) U/mgpr and (7.77±0.39) U/mgpr] were significantly higher than those of the control group [(4.45±0.27) U/mgpr, (4.71±0. 35) U/mgpr and (4. 37±0. 44) U/rngpr] (t=6. 142, 16. 295, 0. 399,P<0. 01). The number of heart malformation in fetal rats (r=0.994,P=0. 000) and blood glucose (r=0. 717,P=0. 000) had the positive relationship with the maternal serum AGE level. The expression of RAGE in fetal heart was positively related with the number of fetal heart malformation (r= 0. 638,P= 0. 004). Conclusions The increased maternal serum AGE level in GDM rats may be an important factor in fetal heart dysplasia.     

妊娠期糖尿病母鼠血清晚期糖基化终产物与子鼠心脏发育异常的关系

Objective To explore the effect of advanced glycation end product(AGE) in serum of maternal rats with gestational diabetes mellitus (GDM) on the heart development of their offsprings. Methods Fifty-four SD rats were randomly assigned into control group (n= 24) and GDM group (n=30) which were established by administration of streptozotocin intra-abdominally. On the gestational age of 13, 16, 19 days, all rats underwent hysterectomy to obtain the fetal heart tissues. Serum level of AGE and blood glucose level of maternal rats were tested. The expression of receptor AGE (RAGE) in fetal cardiac tissue were detected by immunohistoehemistry. Results The incidence of fetal heart defect in GDM group was significantly higher than the control group at each time point (P<0.01). Rats in GDM group had higher blood glucose level at each time point (P<0.01). The AGE levels of GDM group on gestational age of 13, 16 and 19 day [(5.72±0.68) U/mgpr, (7.31±0.29) U/mgpr and (7.77±0.39) U/mgpr] were significantly higher than those of the control group [(4.45±0.27) U/mgpr, (4.71±0. 35) U/mgpr and (4. 37±0. 44) U/rngpr] (t=6. 142, 16. 295, 0. 399,P<0. 01). The number of heart malformation in fetal rats (r=0.994,P=0. 000) and blood glucose (r=0. 717,P=0. 000) had the positive relationship with the maternal serum AGE level. The expression of RAGE in fetal heart was positively related with the number of fetal heart malformation (r= 0. 638,P= 0. 004). Conclusions The increased maternal serum AGE level in GDM rats may be an important factor in fetal heart dysplasia.     

血小板生长因子对缺氧缺血性脑损伤新生鼠脑细胞凋亡率和血清神经元特异性烯醇化酶的影响

目的 研究血小板源性生长因子(platelet derived growth factor,PDGF)对缺氧缺血性脑损伤(hypoxic-ischemic brain damage,HIBD)的新生鼠神经细胞凋亡率及血清神经元特异性烯醇化酶(neuron-specific enolase,NSE)浓度的影响,进而探讨其对HIBD的神经保护作用. 方法 7日龄新生Wistar大鼠48只制备HIBD模型,并分为PDGF治疗组和生理盐水对照组,每组各24只.另取24只为假手术组.治疗组在缺氧缺血后即刻给PDGF-BB 50 ng/kg腹腔注射.对照组和假手术组腹腔注射等体积的生理盐水.每组于处置后12、24和72 h随机取8只处死,留血清标本,酶联免疫吸附法检测大鼠血清标本NSE浓度;取右侧大脑组织制备脑细胞悬液,双染法流式细胞仪检测脑细胞凋亡率.采用单因素方差分析及q检验进行统计学分析. 结果 (1)脑细胞凋亡率:治疗组[(6.09±0.70)％、(9.67±1.52)％和(14.15±1.52)％]和对照组[(8.00±1.10)％、(11.45±2.42)％和(22.90±2.03)％]3个时点的脑细胞凋亡率均较假手术组(2.11±0.54)％、(2.34±0.46)％和(2.21±0.49)％]显著增加(P均＜0.01或＜0.05),治疗组较对照组各时点脑细胞凋亡率均明显降低(P均＜0.01或＜0.05),3组大鼠在12、24、72 h时的组间比较差异均有统计学意义(F=39.01、66.60、194.20,P均＜0.01).(2)血清NSE浓度:各时点对照组[(10.04±0.19) μg/L、(9.33±0.15)μg/L和(8.36±0.16)μg/L]和治疗组[(8.43±0.17)μg/L、(6.73±0.16) μg/L和(6.12±0.13)μg/L]较假手术组[(4.22±0.53)μg/L、(3.96±0.60) μg/L和(3.59±0.55) μg/L]NSE浓度增加(P均＜0.01),治疗组较对照组各时点NSE浓度降低(P均＜0.01),3组大鼠在12、24、72h组间比较差异均有统计学意义(F=371.25、245.61、236.22,P均＜0.01). 结论 PDGF能抑制新生大鼠HIBD后神经细胞凋亡及降低血清NSE浓度,对HIBD新生大鼠有神经保护作用.     

中药补肾调冲方对卵巢早衰大鼠激素水平和卵巢Bcl-2/Bax表达的影响

目的:探讨补肾调冲方对雷公藤多苷片(GTW)致卵巢早衰(POF)的治疗作用。方法:雌性SD大鼠42只,随机分为正常组、模型组、结合雌激素片(雌激素组)和补肾调冲方治疗高、中、低剂量组。运用酶联免疫吸附试验(ELISA)法检测血清中雌二醇(E_2)、卵泡刺激素(FSH)、抑制素B(INHB)水平。Western blotting法和RT-PCR法检测卵巢组织中Bcl-2、Bax蛋白及mRNA的表达水平。结果:正常组和各给药组E_2的含量均高于模型组(P0.01)。正常组和各给药组FSH含量均低于模型组(P0.01);正常组和各给药组INHB含量均高于模型组(P0.01)。低剂量组INHB的含量与雌激素组比较,差异有统计学意义(P0.05)。模型组大鼠卵巢中Bcl-2蛋白和mRNA水平的表达显著低于正常组(P0.05);各给药组Bcl-2蛋白和mRNA水平的表达显著高于模型组(P0.01)。模型组大鼠卵巢中Bax蛋白和m RNA水平的表达显著高于正常组(P0.05);各给药组Bax蛋白和mRNA水平的表达显著低于模型组(P0.01)。低剂量组Bax蛋白的表达与雌激素组比较,差异有统计学意义(P0.05)。结论:中药补肾调冲方对卵巢性激素水平具有调节作用,能提高卵巢对性激素的敏感性,促进卵巢排卵;通过上调Bcl-2和下调Bax的表达,抑制卵巢中颗粒细胞的过度凋亡,减少卵泡闭锁,促进卵巢功能的恢复。     

细胞外信号调节激酶在多囊卵巢综合征患者子宫内膜中的活化及意义

目的 通过测定子宫内膜中胰岛素的丝裂原活化蛋白激酶(MAPK)/细胞外信号调节激酶(ERK)信号通路中ERK1/2(表示ERK1和ERK2,下同)的表达及其活化程度,探讨其在多囊卵巢综合征(PCOS)子宫内膜增生及癌变形成中的作用及意义,以及影响其表达及活化程度的因素.方法 选择2007年1月-2008年6月天津医科大学总医院行诊断性刮宫的PCOS患者52例(PCOS组),相匹配的非PCOS患者(良性卵巢肿瘤)32例为对照组.测定所有观察对象的空腹血糖及胰岛素水平;所有观察对象均行子宫内膜病理检查,按子宫内膜病理结果将PCOS组患者分为子宫内膜增生及癌变组(19例)和正常子宫内膜组(33例);根据是否存在胰岛素抵抗将PCOS组患者分为胰岛素抵抗组(38例)和非胰岛素抵抗组(14例).蛋白印迹法(western blot)检测子宫内膜中ERK1/2及其活化形式--磷酸化ERK1/2(p-ERK1/2)蛋白的表达水平.结果 (1)PCOS组患者子宫内膜中p-ERK1/2蛋白的表达水平[(61±13)%]高于对照组[(44±10)%],两组比较,差异有统计学意义(P＜0.01).(2)子宫内膜增生及癌变组子宫内膜中p-ERK1/2蛋白的表达水平[(70±11)%]高于正常子宫内膜组[(55±10)%],两组比较,差异有统计学意义(P＜0.01);胰岛素抵抗组子宫内膜中p-ERK1/2蛋白的表达水平[(63±13)%]高于非胰岛素抵抗组[(55±7)%],两组比较,差异有统计学意义(P＜0.01).(3)空腹胰岛素水平、胰岛素曲线下面积、体质指数与PCOS组患者子宫内膜中p-ERK蛋白的表达水平相关,相关系数分别为0.447、0.456、0.381(P均＜0.01).结论 PCOS患者子宫内膜存在MAPK/ERK信号通路的过度活化,与子宫内膜增生及癌变的发生有关;胰岛素抵抗及高胰岛素血症有促进ERK活化的作用.     

宫内窘迫胎儿出生后心肌损伤情况的监测

目的 探讨无窒息的宫内窘迫胎儿出生后是否合并有心肌损伤,以及检测心肌损伤的生化敏感指标.方法 2009年7月至12月,随机选择中山大学附属第一医院分娩的、有宫内窘迫史的53例新生儿为宫内窘迫组,新生儿出生后Apgar评分1 min及5 min均＞7分.同期分娩的无宫内窘迫史的新生儿36例作为对照组,新生儿出生后Apgar评分1 min及5 min均为10分.胎儿娩出后立即抽取脐动脉血进行血气分析和生化指标测定.结果 (1)宫内窘迫组新生儿脐动脉血pH值及剩余碱分别为7.23±0.07及(-4.8±3.0)mmol/L,明显低于对照组的7.31±0.03及(-2.1±1.5)mmol/L,两组比较,差异有统计学意义(P＜0.05).宫内窘迫组新生儿脐动脉血乳酸水平为(5.2±2.3)mmol/L,明显高于对照组的(2.3±1.1)mmol/L,两组比较,差异有统计学意义(P＜0.01).宫内窘迫组新生儿脐动脉血氧分压及二氧化碳分压水平分别为(16.2±7.9)及(54.0±11.2)mm Hg(1mm Hg=0.133 kPa),对照组分别为(17.5±6.7)及(48.5±5.4)mm Hg,两组分别比较,差异均无统计学意义(P＞0.05).(2)宫内窘迫组新生儿脐动脉血肌酸激酶同工酶MB(CK-MB)水平为(48±59)U/L,对照组为(36±27)U/L,两组比较,差异有统计学意义(P＜0.05);宫内窘迫组新生儿脐动脉血肌酸激酶(CK)及脑钠肽(BNP)水平分别为(194±73)U/L及(519±309)ng/L,对照组分别为(162±95)及(481±216)ng/L,两组比较,差异无统计学意义(P＞0.05).(3)新生儿脐动脉血CKMB水平与脐动脉血pH值、剩余碱呈负相关性(r=-0.296及-0.318,P均＜0.05);BNP与乳酸水平呈正相关(r=0.278,P＜0.05);其余各变量间均无相关性(P＞0.05).结论 宫内窘迫的胎儿即使出生时无新生儿窒息的表现,也存在着不同程度的心肌损伤;脐血CK-MB水平变化可作为监测心肌损伤的敏感指标,心肌损伤的程度与胎儿酸中毒的程度有关.

Abstract：

Objective To investigate whether no asphyxia neonates with intrauterine distress are complicated with myocardial injury and determine the sensitive biochemical diagnostic parameters. Methods A total of 89 neonates born in the First Affiliated Hospital of Sun Yat-sen University from July 2009 to December 2009 were enrolled. Fifty-three fetal distress cases with Apgar score ＞ 7 at 1 and 5 minites were enrolled in the study group; while the rest 36 healthy neonates, whose Apgar score = 10 at 1 and 5 minites, were the control group. Umbilical artery blood samples of all cases were collected for blood gas analysis and biochemical measurement. Results(1)pH(7.23±0.07) and BE [(-4.8±3.0)mmol/L] in the study group were significantly lower than pH (7.31 ±0.03) and BE [(-2.1±1.5)mmol/L] in the control group (P＜0.05).The lactic acid of study group [(5.2±2.3)mmol/L] was higher than that of the control group [(2.3±1.1)mmol/L], and the difference was significant (P＜0.01). However, there was no significant difference between the two groups in PaO2[(16.2±7.9)mm Hg(1 mm Hg=0.133 kPa) vs. (17.5±6.7)mm Hg] and PaCO2[(54.0±11.2)mm Hg vs. (48.5±5.4) mm Hg; P＞0. 05]. (2) The level of CK-MB in neonates with fetal distress[(48 ±59) U/L] was significantly higher than that of healthy neonates [(36±27)U/L]. However, no significant difference was found in CK [(194±73)U/L vs. (162±95) U/L]and BNP levels[(519±309)ng/L vs.(481±216)ng/L;P ＞ 0.05]. (3) Spearman rank correlation analysis showed that CK-MB level was negatively correlated with pH(r=-0.296, P＜0.05) and BE (r=-0.318,P＜0.05) of umbilical artery blood,while BNP level was positively correlated with umbilical lactic acid (r=0.278, P＜0.05). No correlation was found between other parameters (P＞0.05).Conclusions Intrauterine distress without neonatal asphyxia had effect on fetal myocardial injury. CK-MB can be used as a sensitive parameter for monitoring the development of myocardial injury. The severity of myocardial injury was related to fetal acidosis.     

细胞外信号调节激酶信号通路对蛋白酶体抑制剂MG262诱导卵巢上皮性癌细胞凋亡的调控作用

目的 探讨细胞外信号调节激酶(ERK)信号通路对蛋白酶体抑制剂MG262诱导卵巢上皮性癌(卵巢癌)细胞凋亡的调控作用.方法 不同浓度(1、10、20、40、60、80 nmol/L)的MG262分别处理卵巢癌细胞株SKOV3细胞24、48 h后,四甲基偶氮唑蓝(MTT)比色法检测SKOV3细胞的存活率.MG262及ERK抑制剂PD98059分别及联合处理24 h后,流式细胞仪检测SKOV3细胞及胚肾上皮细胞株293T细胞的凋亡率;酶联免疫吸附试验(ELISA)及蛋白印迹法分别检测SKOV3细胞培养上清液中血管内皮生长因子(VEGF)的浓度及细胞内VEGF蛋白的表达.MG262处理不同时间(3、6,9、12 h)后,蛋白印迹法检测SKOV3细胞内磷酸化ERK(p-ERK)及野生型p53蛋白的表达.以上实验以未加药物者为对照.结果 MTT比色法检测发现,随着MG262浓度的增加,SKOV3细胞的存活率明显下降(P＜0.05).流式细胞仪检测发现,MG262、PD98059分别或联合处理SKOV3细胞后,其细胞凋亡率分别为(30.7±4.3)%、(26.8±8.6)%、(50.3±10.6)%,与对照细胞的(7.9±1.9)%比较,差异均有统计学意义(P＜0.05);两药单独处理者分别与联合处理者比较,差异也均有统计学意义(P＜0.01).而两药分别或联合处理293T细胞后,其细胞凋亡率分别为(14.5±5.3)%、(16.2±7.5)%、(10.8±7.3)%,与对照细胞的(12.2±6.3)%比较,差异均无统计学意义(P＞0.05).ELISA及蛋白印迹法检测显示,MG262处理后SKOV3细胞培养上清液中VEGF的浓度及细胞内VEGF蛋白的表达水平均明显下降(P＜0.05).MG262处理不同时间后,SKOV3细胞内p-ERK蛋白的表达水平明显下降(P＜0.05);但SKOV3细胞内野生型p53蛋白在MG262处理前、后均无表达.结论 MG262可通过ERK信号通路诱导卵巢癌细胞凋亡,抑制卵巢癌细胞的增殖及血管生成.     

促性腺激素释放激素激动剂联合反向添加疗法治疗子宫内膜异位症的效果及安全性

目的 探讨促性腺激素释放激素激动剂(GnRH-a)联合反向添加疗法(经皮雌激素及口服醋酸甲羟孕酮)治疗子宫内膜异位症(内异症)的疗效及安伞性.方法 选择2007年1月1日-7月31日于复旦大学附属妇产科医院住院接受治疗、经腹腔镜或开腹手术确诊的内异症患者28例,随机分为A、B两组,每组各14例.A组患者于月经周期第2天起给予戈舍瑞林3.6 mg皮下注射,每4周注射1次,共12周;B组患者在A组治疗方案基础上同时加用半水合雌二醇贴剂,每周1/2片贴于腹部皮肤,并每晚口服醋酸甲羟孕酮6 mg,共12周.比较两组患者治疗前、治疗后(疗程第12周内)及月经恢复后的血清激素及骨钙素水平、阴道脱落细胞百分比、疼痛症状视觉模拟评分(VAS)、腰椎骨密度及骨量丢失率、绝经症状严重程度(以Kupperman评分表示)等.结果 (1)两组患者治疗后的m清卵泡刺激素(FSH)及雌二醇水平,A组分别为(5.0±2.6)U/L和(29±17)pmol/L,B组分别为(3.0±1.5)U/L和(87±53)pmol/L,均分别低于治疗前水平[A组分别为(17.0±12.2)U/L和(184±194)pmol/L.,B组分别为(15.3±13.6)U/L和(281±242)pmol/L],差异均有统计学意义(P＜0.01);治疗后B组的雌二醇水平高于A组,FSH水平低于A组,差异也均有统计学意义(P＜0.01).(2)治疗后的阴道脱落细胞中,A组的底层细胞百分比[(66.2±29.0)%]高于B组[(11.8±28.0)%],中层细胞[(29.1±23.1)%]、表层细胞[(4.0±5.5)%]和伊红染色细胞百分比[(2.3±2.6)%]则分别低于B组[分别为(73.0±25.2)%、(15.2±10.9)%、(10.8±7.9)%],差异均有统计学意义(P＜0.01).(3)两组患者治疗前的疼痛症状总分及盆腔痛、痛经、性交痛评分,A组分别为(7.43±3.20)、(2.35±1.82)、(4.93±1.98)和(0.14±0.53)分,B组分别为(7.71±2.02)、(2.57±1.60)、(4.86±1.56)和(0.29±1.07)分;治疗后,两组患者的VAS总分及各项评分[A组分别为(0.14±0.36)、(0.07±0.27)、(0.07±0.27)和0分,B组分别为(0.36±0.50)、(0.29±0.47)、(0.07±0.27)和0分]均低于治疗前,差异均有统计学意义(P＜0.01);月经恢复后,两组的总分及各项评分[A组分别为(0.21±0.43)、(0.07±0.27)、(0.14±0.36)和0分,B组分别为(0.50±0.65)、(0.29±0.47)、(0.2l±0.43)和0分]也均低于治疗前,差异也均有统计学意义(P＜0.01);两组之间各项评分比较,差异均无统计学意义(P＞0.05).(4)治疗前A、B两组的骨密度分别为(0.99±0.06)g/cm2和(0.99±0.10)g/cm2,治疗后A组的骨密度为(0.96±0.06)g/cm2,低于治疗前水平,差异有统计学意义(P＜0.01),B组的骨密度为(0.98±0.09)g/cm2,也低于治疗前水平,但差异无统计学意义(P=0.201);治疗前、后两组间骨密度分别比较,筹异均无统计学意义(P＞0.05).A、B两组的骨量丢失率分别为(-2.77±1.97)%和(-0.93±2.86)%,两组之间比较,差异无统计学意义(P=0.058).治疗前A、B两组外周血骨钙素水平分别为(13±3)μg/L和(13±6)μg/L,治疗后A组外周血骨钙素水平为(17±6)μg/L,高于治疗前水平,差异有统计学意义(P＜0.01),B组为(16±6)o,g/L,也高于治疗前水平,但差异无统计学意义(P=0.053);治疗前、后两组问骨钙素水平分别比较,差异均无统计学意义(P＞0.05).(5)A、B两组患者治疗后的改良Kupperman评分总分分别为(15±7)分和(11±6)分,两组间比较,差异无统计学意义(P＞0.05).A、B两组患者中潮热出汗的发生率分别为93%(13/14)和57%(8/14),两组间比较,差异有统计学意义(P＜0.01).结论 经皮雌激素加订服醋酸甲羟孕酮治疗内异症是一种安全有效的反向添加疗法.