TGF-β1、Smad4和Smad7与子宫内膜异位症关系的研究

目的探讨转化生长因子-β1（transforming growthfactor-β1,TGF-β1）及其信号通路Smad4和Smad7与子宫内膜异位症发病的关系。方法选择2007年8月至2008年3月郑州大学第三附属医院子宫内膜异位症患者30例为研究对象,分别取其异位内膜和在位内膜。同期子宫畸形患者30例为对照组,取其正常子宫内膜。采用免疫组织化学SP法检测组织中TGF-β1、Smad4和Smad7的表达。结果TGF-β1在异位内膜（181．71±4．90）和在位内膜组（179．05±1．26）的表达与对照组（92．45±3．26）比较,表达升高（P〈O．05）;Smad4和Smad7在异位内膜（95．58±9．05和98．83±10．48）和在位内膜组（95．09±2．16和93．51±4．49）的表达与对照组（185．37±6．39和184.56±231）比较,表达降低（P〈0．05）。三者异位内膜和在位内膜组的表达比较,差异均无统计学意义（P〉O．05）;异位内膜组TGF-β1和Smad4的表达呈负相关（r=-0．349,P〈0．05）,TGF-β1和Smad7的表达呈负相关（r=-0．247,P〈0．05）;TGF-β1、Smad4和Smad7在在位内膜组的增殖期和分泌期比较,差异无统计学意义（P〉O．05）。结论TGF-β1、Smad4和Smad7在子宫内膜异位症的发生中可能起到一定的作用。     

趋化因子CXCL12及其受体CXCR4对卵巢上皮性癌细胞增殖、迁移和侵袭能力的影响

目的 探讨趋化因子CXCL12及其受体CXCR4对卵巢上皮性癌（卵巢癌）细胞增殖、迁移和侵袭能力的影响。方法采用RT—PCR技术和免疫细胞化学染色法检测卵巢癌细胞株CAOV3细胞中CXCL12、CXCR4mRNA和蛋白的表达,以及CXCL12（100ng／m1）作用后CAOV3细胞中整合素B1和血管内皮生长因子C（VEGF—C）mRNA的表达。实验分为6组：对照组（未加药）,实验组1（加100ng／ml的CXCL12）,实验组2（加10ng／ml的CXCL12）,实验组3（加100ng／ml的CXCL12和10斗g／ml的CXCR4抗体）,实验组4（加100ng／ml的CXCL12和1pg／ml的CXCR4拮抗剂——AMD3100）,实验组5（加10μg／ml的CXCR4抗体或卵巢癌腹水）。采用四甲基偶氮唑蓝比色法检测CXCL12对CAOV3细胞增殖的影响。以穿膜小室为模型,采用重组细胞基底膜迁移、侵袭实验检测CXCL12和卵巢癌腹水对CAOV3细胞迁移、侵袭的影响。结果 CAOV3细胞中,CXCR4蛋白呈棕黄色强阳性表达,CXCR4mRNA的表达水平为0．70±0．10,经100ng／ml的CXCL12作用24h时CXCR4mRNA的表达水平（1．24±0．14）显著增加（t=-7．1088,P=0．0021）;CXCL12蛋白和mRNA均无表达。100ng／ml的CXCL12作用3h时整合素131mRNA表达水平即显著增加（作用前后分别为0．53±0．10、1．53±0．16,P〈0．01）,24h时VEGF—CmRNA表达水平显著增加（作用前后分别为0．52±0．09、1．11±0．15,P〈0．01）。对CAOV3细胞的增殖作用,实验组1分别与对照组和实验组2比较均明显增强（3组分别为0．428±0．051、0．325±0．045、0．328±0．039,P〈0．05）;实验组1分别与实验组3和实验组4比较均明显被抑制（后两组分别为0．356±0．031、0．373±0．029,P〈0．05）;而实验组5（0．349±0．038）与对照组比较,差异则无统计学意义（P〉0．05）。对CAOV3细胞的迁移和侵袭作用,实验组1分别与对照组和实验组2比较均明显增强[迁移细胞数分别为（523．3±25．2）、（108．0±7．2）、（211．7±24．7）个,侵袭细胞数分别为（39．3±4．0）、（4．0±1．0）、（15．7±3．1）个;P均〈0．01];实验组1分别与实验组3和实验组4比较均明显被抑制（P〈0．05）;但实验组5[即加入卵巢癌腹水组;迁移细胞数为（706．6±30．6）个,侵袭细胞数为（61．7±7．6）个]显著多于实验组1（P〈0．05）。结论 CXCL12可促进卵巢癌细胞增殖、迁移和侵袭,并上调整合素B1和VEGF—CmRNA的表达,此作用可被CXCR4抗体抑制,表明CXCL12在卵巢癌的生长和转移中发挥一定作用。     

叉头框F2在宫腔粘连中的表达及意义

目的:建立稳定的宫腔粘连细胞模型,检测叉头框F2(Fox F2)表达。方法:原代培养人子宫内膜基质细胞(HESCs),免疫荧光法进行细胞鉴定。用0、2.5、5和10ng/ml转化生长因子β1(TGF-β1)作用于HESCs 48h,10ng/ml TGF-β1作用于HESCs 12、24、48h和72h,PCR及Western blot法检测α-平滑肌肌动蛋白(α-SMA)、胶原I(COLI)及Fox F2 m RNA和蛋白表达。结果:与0ng/ml组比较,2.5、5ng/ml和10ng/ml组α-SMA、COLI及Fox F2的m RNA和蛋白表达均逐渐增加。随着10ng/ml TGF-β1作用时间的延长,与12h组相比,24、48和72h组α-SMA、COLI和Fox F2的m RNA和蛋白表达逐渐增加。结论:TGF-β1可诱导HESCs发生纤维化改变,建立宫腔粘连细胞模型。随着TGF-β1作用时间的延长,作用浓度的增加,Fox F2在宫腔粘连中表达增加。     

PinlsiRNA对宫颈癌细胞SiHa增殖和凋亡的影响

目的：通过小干扰RNA（siRNA）栽体介导抑制肽基脯氨酰同分异构酶（Pinl）表达,探讨Pinl对宫颈癌细胞SiHa增殖与凋亡的影响。方法：构建pGPU6／GFP／Neo—PinlsiRNA表达载体,脂质体介导将其转染至宫颈癌细胞SiHa中;实验设3组：实验组（SiHa／pGPU6-PinlsiRNA组）、阴性对照组（SiHa／pGPU6-con组）和空白对照组（Si-Ha）。采用RT—PCR、Western blot法检测Pinl的表达;M1Tr法检测细胞增殖活性;原位末端标记法（TUNEL）和流式细胞术（FCM）检测细胞凋亡。结果：转染SiHa／pGPU6．PinlsiRNA的SiHa细胞中,PinlmRNA和蛋白表达分别下调68．1％和54．8％,细胞增殖显著抑制。TUNEL显示典型细胞凋亡特征,SiHa／pGPU6-PinlsiRNA组的凋亡指数（AI）为（29．6±14．58）％,显著高于SiHa／pGPU6-con组[（8．44±5．65）％]和空白对照组[（5．34±4．47）％]（P〈O．05）。SiHa／pGPU6．PinlsiRNA组的细胞凋亡率为（30．95±16．47）％,显著高于SiHa／pGPU6-COIl组[（6．18±6．10）％]和空白对照组[（5．95±4．99）％]（P〈0．05）。结论：RNA干扰技术能特异高效的抑制Pinl基因的表达,Pinl基因的下调能抑制宫颈癌细胞增殖并促进细胞凋亡,提示Pinl可能成为治疗宫颈癌的新靶点。     

表皮生长因子卵丘细胞对卵母细胞体外成熟的影响

目的：探讨表皮生长因子（EGF）、卵丘细胞人类生殖泡（germinal vesicle ,GV)期不成熟卵母细胞体外成熟的影响，方法：将EGF和促性激素（gonadotropin,Gn）联合应用于不成熟卵母细胞的体外培养，并同单纯应用Gn相对照观察EGF的影响。同时将卵母细胞分为卵丘卵母细胞复合物（CEO组）及裸卵（DO组）进行培养，了解卵丘细胞在体外培养中的作用。结果：EGF可以提高DO组的MⅡ期率（P＜0．01），对于CEO组应用EGF后可以促进胞质的成熟，提高受精能力（P＜0．05）。关于卵丘细胞的作用，CEO组的MⅡ期率为68．8％，明显高于DO组的46．7％（P＜0．01），CEO组的卵裂率为72．7％，高于DO组的35．7％（P＜0．05）。结论：EGF可以直接促进不成熟裸卵体外成熟，提高其MⅡ期率，与Gn联合应用后可进一步促进卵丘卵母细胞复合物胞质的成熟，提高受精率，保留卵丘细胞利于卵母细胞质的发育，提高不成熟卵母细胞的体外成熟能力，使MⅡ期率，卵裂率升高。     

妊娠期高血压疾病孕妇血及脐血中抑制素及表皮生长因子水平的变化

目的探讨妊娠期高血压疾病孕妇血及其分娩的新生儿脐血血浆中抑制素(INH)和表皮生长因子(EGF)的水平变化及其对妊娠期高血压疾病发病及胎儿生长发育的影响。方法采用酶联免疫吸附试验(ELISA),检测65例妊娠期高血压疾病患者(观察组)及21例正常妊娠妇女(对照组)及其分娩的新生儿脐血血浆中INH、EGF水平。结果观察组孕妇血浆INH水平为(499±52)ng／L,对照组为(421±36)ng／L,两组比较,差异有统计学意义(P<0．01)。观察组新生儿脐血INH水平为(274±53)ng／L,对照组为(252±28)ng／L,两组比较,差异无统计学意义(P>0．05)。观察组EGF水平为(408±60)ng／L,对照组为(463±87)ng／L,两组比较,差异有统计学意义(P< 0．05)。观察组新生儿脐血中EGF水平为(232±99)ng／L,对照组为(380±97)ng／L,两组比较,差异也有统计学意义(P<0．01)。脐血中EGF水平与新生儿体重呈正相关关系(r=0．326,P<0．05),与胎盘重量呈正相关关系(r=0．451,P<0．05)。结论孕妇血浆中INH、EGF水平与妊娠期高血压疾病的发病有关,脐血中EGF水平对胎儿及胎盘的生长发育有影响。     

胰岛素对子宫内膜癌细胞增殖和凋亡的影响

目的 探讨胰岛素对子宫内膜癌细胞系Ishikawa3-H-12细胞增殖、凋亡和细胞周期的影响。方法 应用免疫细胞化学方法和RT-PCR技术检测Ishikawa3-H-12细胞胰岛素受体（INSR）蛋白和mRNA的表达。以不同浓度胰岛素作用Ishikawa3-H-12细胞不同时间,采用四甲基偶氮唑蓝比色法、流式细胞仪检测细胞增殖、凋亡和细胞周期。结果 （1）Ishikawa3-H-12细胞INSR蛋白呈棕黄色阳性表达,并可见INSR基因的表达。（2）胰岛素以浓度和时间依赖的方式促进子宫内膜癌细胞增殖,1×10^-4mol／L胰岛素作用48h时促增殖作用最显著,增殖率为（340．2±15．9）％,与对照细胞（以胰岛素浓度为0作为对照,为100％）比较,差异有统计学意义（P〈0．05）。（3）胰岛素以浓度和时间依赖的方式使Ishikawa3-H-12细胞中G0／G1期细胞比例减少,S期细胞比例增加,1×10^-4mol／L胰岛素作用72h时最显著,G0／G1期细胞比例为（27．7±2．5）％,S期细胞为（55．2±1．4）％,分别与对照细胞[分别为（67．6±1．5）％、（15．7±1．0）％]比较,差异均有统计学意义（P〈0．05）;胰岛素对G2／M期细胞无影响（P〉0．05）。（4）随着胰岛素浓度的增加,Ishikawa3-H-12细胞凋亡率逐渐下降。1×10^-4mol／L胰岛素作用最显著,作用24、48、72、96h时的细胞凋亡率分别为（1．76±0．16）％、（1．70±0．15）％、（1．56±0．20）％、（1．31±0．24）％,分别与对照细[分别为（9．81±0．61）％、（9．93±1．44）％、（9．10±0．66）％、（10．30±1．20）％]比较,差异均有统计学意义（P〈0．05）。结论 胰岛素对子宫内膜癌Ishikawa3-H-12细胞具有促进增殖、抑制凋亡的作用。     

表皮生长因子(EGF)和胰岛素样生长因子(IGF-Ⅱ)对小鼠1-细胞胚胎体外发育的影响

目的:研究表皮生长因子(epidermalgrowthfactor,EGF)在小鼠早期胚胎体外发育中的作用,以及EGF联合胰岛素样生长因子-Ⅱ(insulin-likegrowthfactor-II,IGF-Ⅱ)对小鼠早期胚胎体外发育是否有协同促进作用。方法:①在mKSOM培养液中添加0ng/ml(对照组)、0.1ng/ml、1ng/ml、10ng/ml、100ng/ml的EGF、10ng/mlIGF-Ⅱ、1ng/mlEGF+1ng/mlIGF-Ⅱ、1ng/mlEGF+10ng/mlIGF-Ⅱ、10ng/mlEGF+1ng/mlIGF-Ⅱ、10ng/mlEGF+10ng/mlIGF-Ⅱ培养小鼠1-细胞胚胎,每组胚胎在培养箱中连续培养120h,每24h观察胚胎发育情况,分别计算2-细胞率、4-细胞率、桑椹胚率、囊胚率和孵化率,并进行囊胚细胞计数。结果:添加1ng/ml、10ng/mlEGF组的囊胚率及孵化率显著增加;EGF+IGF-Ⅱ组较单一添加组、对照组囊胚率及孵化率显著增加(P<0.05),且EGF+IGF-Ⅱ组囊胚细胞计数显著高于其他组(P<0.05),其中10ng/mlEGF+10ng/mlIGF-Ⅱ组的孵化率及囊胚细胞数最高(P<0.05)。结论:EGF浓度在1￣10ng/ml范围内可以提高体外培养鼠胚的囊胚率及孵化率;EGF及IGF-Ⅱ对体外培养鼠胚的发育有协同促进作用;本实验范围内10ng/mlEGF+10ng/mlIGF-Ⅱ为最优组合。     

胰岛素受体α、β及胰岛素受体底物-1在子宫内膜癌的表达及其临床意义

目的 探讨胰岛素受体α（IRα）、β（IRβ）及胰岛素受体底物-1（IRS-1）在子宫内膜癌的表达及其临床意义。方法 用免疫组织化学法检测10例正常子宫内膜：18例子宫内膜增生及57例子宫内膜癌组织中IRα、IRβ及IRS-1蛋白的表达。结果 IRα、IRβ、IRS-1的表达在10正常子宫内膜表达分别为80．00％（8／10）、60．00％（6／10）及70．00％（7／10）；在18例子宫内膜增生组织中分别为77．78％（14／18）、61．11％（11／18）及77．78％（14／18）；在子宫内膜癌组织中分别为47．37％（27／57）、54．39％（31／57）及63．16％（36／57）。IRα的表达在子宫内膜癌组织与正常子宫内膜组及子宫内膜增生比较差异有显著性（P〈0．05），其余差异无统计学意义（P〉0．05）；Ⅰ期子宫内膜癌组织中IRα的阳性表达率明显低于Ⅱ～Ⅳ期者（P〉0．05）；IRβ及IRS-1各组间的阳性表达率与Ⅱ～Ⅳ期者相近（P〉0．05）；IRS-1的阳性表达率Ⅰ期明显低于Ⅱ～Ⅳ期者（P〉0．05）；IRα、β、IRS-1蛋白的表达在子宫内膜癌G1—2与G3差异有显著性（P〈0．05），与肌层浸润深度及淋巴结转移与否差异无显著性（P〉0．05）。结论 IRα及IRS-1的表达与子宫内膜癌的期别有关，提示其表达的改变可能是子宫内膜腺癌发生的早期事件。     

子痫前期患者胎盘组织中转化生长因子β1、血管细胞黏附分子1及E选择素表达的研究

目的探讨子痫前期患者胎盘组织中转化生长因子β1(TGF-β1)、血管细胞黏附分子1(VCAM-1)和E选择素(E-selectin)的表达变化及其意义。方法用免疫组化链霉菌抗生物素蛋白-过氧化物酶连接(SP)法,对20例正常孕妇(对照组)和40例子痫前期患者(子痫前期组,其中轻度16例、重度24例)的胎盘组织进行TGF-β1、VCAM-1和E-selectin定位,并用计算机图像分析系统进行定量分析比较。结果(1)1FGF-β1在胎盘绒毛合体滋养细胞的表达,子痫前期组为70．7±0．5,对照组为70．3±0．6,两组比较,差异有统计学意义(P<0．05)。VCAM-1的表达,子痫前期组为82．5±0．5,对照组为82．8±0．3;E-selectin的表达,子痫前期组为53．5±0．5,对照组为53．8±0．4;两组分别比较,差异均有统计学意义(P<0．05)。TGF-β1、VCAM-1、E-selectin在轻度子痫前期患者胎盘组织合体滋养细胞中的表达分别为70．6±0．6、82．4±0．6、53．4±0．5,在重度子痫前期患者胎盘组织中的表达分别为70．8±0．4、82．6±0．5、53．6±0．5,轻度与重度子痫前期患者TGF-β1、VCAM-1、E- selectin表达水平比较,差异均无统计学意义(P>0．05)。(2)E-selectin在胎盘绒毛毛细血管内皮细胞的表达,子痫前期组为63．0±0．5,对照组为62．6±0．4,两组比较,差异有统计学意义(P<0．05);轻度子痫前期患者为63．2±0．4、重度子痫前期为62．9±0．5,二者比较,差异无统计学意义(P> 0．05)。结论TGF-β1、VCAM-1和E-selectin在胎盘组织中的表达变化,在子痫前期发病中有重要作用。     

bFGF and VEGF improve the quality of vitrified-thawed human ovarian tissues after xenotransplantation to SCID mice

Purpose

The aim of this research is to study whether basic fibroblast growth factor (bFGF) alone or in combination with vascular endothelial growth factor (VEGF) could improve the quality of vitrified-thawed human ovarian tissue xenotransplanted to severe combined immune deficiency (SCID) mice.

Methods

After collection and cryopreservation, thawed human ovarian tissue were cultured in vitro for 2 days and then xenografted to severe combined immune deficiency (SCID) mice for 7 days. The in vitro culture medium was separated into six groups, including (A) the blank control group, (B) the human recombinant bFGF (150 ng/ml) group, (C) the bFGF (150 ng/ml)+human recombinant VEGF (25 ng/ml) group, (D) bFGF (150 ng/ml)+VEGF (50 ng/ml) group, (E) bFGF (150 ng/ml)+ VEGF (75 ng/ml) group and (F) bFGF (150 ng/ml) + VEGF (100 ng/ml) group. In addition, eight pieces of thawed ovarian tissue were transplanted without in vitro culture, which serve as the fresh control group. The effect of transplantation was assessed by histological analysis, immunohistochemical staining for CD34, Ki-67, and AC-3 expression, and microvessel density (MVD).

Results

There was no significant difference between the fresh and blank control group. Compared to the blank control group, the number of follicles, MVD, and rate of Ki-67-positive cells increased significantly in groups B, C, D, E, and F, while apoptosis decreased significantly. Compared to the bFGF treatment group, no significant difference appeared in group C, D, E, and F.

Conclusions

The administration of bFGF alone or in combination with VEGF improved the quality of postgraft human ovarian tissue, though VEGF, regardless of different concentrations, did not influence effect of bFGF.

     

Concentrations of MMP-1, TIMP-1, MMP-1/TIMP-1 and I CTP complexes in follicular fluid as related to fertilization rate in women treated with in-vitro fertilization

OBJECTIVES: Metalloproteinases and their inhibitors appear to control connective tissue remodelling during follicular rupture. The aim of the study was to establish if oocytes fertilisation rate after ovulation induction depends on the concentrations of MMP-1, its inhibitor TIMP-1, MMP-1/TIMP-1 complex and I CTP in follicular fluid (FF). MATERIAL AND METHODS: FF were collected from 37 infertile patients undergoing ovulation induction using either short or long protocol. FF was obtained 36 hours after administration of hCG (Pregnyl). The level of MMP-1, TIMP-1, MMP-1/TIMP-1 complex were measured using ELISA kits and I CTP, E2, FSH, LH, using RIA assay kits. RESULTS: Statistically significant (p < 0.05) difference was found in TIMP-1, E2, and FSH concentration, being higher in the group with more than 75% fertilisation rate: TIMP-1 728.8 + 100.1 ng/ml vs 666.3 + 94.5 ng/ml; E2 477.3 +/- 160.0 ng/ml vs 368.0 +/- 190.0 ng/ml and FSH 7.27 +/- 1.45 mIU/ml vs 6.24 +/- 1.6 mIU/ml. CONCLUSIONS: Statistically significant increase in TIMP-1 concentration observed among patients with fertilisation rate above 75% indicates an important role of this substance in ovulation process.     

The effect of oxygen tension on intracellular survival signalling in primary villous trophoblasts

Villous trophoblasts undergo increased apoptosis and experience a wider gradient of oxygen tensions (pO(2)) in pregnancies complicated by intrauterine growth restriction. We hypothesize that pO(2)affects trophoblast apoptosis by altering survival signalling through the phosphatidylinositol-3 (PI-3)-kinase and mitogen activated protein kinase (MAPK) pathways. Cytotrophoblasts were cultured at pO(2)from <10 to approximately 140 mmHg with Epidermal Growth Factor (EGF), Vascular Endothelial Growth Factor (VEGF) and basic Fibroblast Growth Factor (bFGF) at concentrations of 0.1 to 10 ng/ml for 1 to 12 h, then assessed for apoptosis (TUNEL) and specific protein expressions (Western blot analysis). Spontaneous apoptosis was highest at <10 mmHg and lowest approximately 15 mmHg. Only EGF activated either signalling pathway at any pO(2). Inhibition of both pathways was required to inhibit EGF-stimulated survival. Maximal EGF activation of either pathway was insensitive to pO(2). At lower oxygen tensions, MAPK phosphorylation was maximal at 1 ng/ml EGF compared with 10 ng/ml for the PI-3-kinase path. The EGF receptor was spontaneously phosphorylated with increasing culture times at lower oxygen levels, an effect reflected down-stream by PI-3-kinase and Akt phosphorylation. We conclude that strong survival signalling in trophoblasts requires both PI-3- and MAP-kinase pathways, is rather insensitive to pO(2)changes and is spontaneously activated with increasing hypoxic exposure.     

不同培养条件对孕中期羊水干细胞分离及扩增能力的影响

目的 探讨不同培养条件对孕中期羊水干细胞分离及扩增能力的影响.方法 收集2007年9月-2008年6月在解放军总医院妇产科因医疗指征引产的15例孕16～24周产妇.经腹部穿刺抽取羊水10～20 ml,收集细胞并根据培养条件不同分为4组:(1)低糖Dulbecco改良Eagle培养基(LD培养基)+10%胎牛血清(10%血清组);(2)LD培养基+20%胎牛血清(20%血清组);(3)LD培养基+15%胎牛血清+碱性成纤维生长因子(bFGF,bFGF组);(4)LD培养基+10%胎牛血清+0.1%明胶铺板(明胶组).比较各组培养的原代细胞集落数和成纤维样细胞集落数、细胞形态、可传代次数等;采用流式细胞仪、RT-PCR技术对各组干细胞特异性标志物进行鉴定;诱导各组向脂肪细胞分化;并行染色体核型分析.结果 (1)10%血清组、20%血清组、bFGF组及明胶组的羊水干细胞扩增培养的成功率分别为60%、73%、73%及60%,10%血清组及明胶组分别与20%血清组及bFGF组比较,差异均无统计学意义(P>0.05);各组细胞集落数分别为(0.9±0.5)、(2.6±1.5)、(2.9±1.5)、(1.1±0.8)个,10%血清组及明胶组分别与20%血清组及bFGF组比较,差异均有统计学意义(P<0.01);各组成纤维样细胞集落数比例分别为46%、49%、64%及44%,各组分别比较,差异均无统计学意义(P>0.05).(2)各组第2代细胞经诱导均可分化为脂肪样细胞.(3)bFGF组有5例持续传代5次以上,细胞形态稳定,细胞数多达1 ×107个以上,与其他各组比较,差异均有统计学意义(P<0.01).(4)染色体核型分析各组均正常.(5)流式细胞仪及RT-PCR技术检测显示,bFGF组传代的细胞均能表达干细胞的标志物阶段特异性胚胎抗原4(SSEA-4)、Oct-4及Nanog基因,而其他各组在传代中细胞出现分化或停止生长.结论 孕中期羊水中可成功分离获得具有干细胞特性的细胞群,适当的血清浓度及添加bFGF能增加羊水干细胞扩增培养的效率.     

孕酮和表皮生长因子在体外培养新生大鼠卵巢原始卵泡生长启动中的作用

目的:探讨孕酮(P_4)和表皮生长因子(epidermal growth factor,EGF)在原始卵泡生长启动中的作用。方法:取出生2d的SD大鼠卵巢,分别在含不同浓度P_4、EGF和P_4+EGF的Waymouth培养体系中培养,以单纯Waymouth培养体系培养的大鼠卵巢为对照,于培养8 d后石蜡包埋切片, HE染色,观察各级卵泡形态,并统计分析各级正常卵泡数。结果:P_4各浓度组的原始卵泡数明显多于对照组(P<0.01),50 ng/ml EGF组的初级卵泡和次级卵泡数明显多于对照组(P<0.01),但1 000 ng/ml EGF组的初级卵泡数和次级卵泡数均明显少于对照组(P<0.01);50 ng/ml EGF+10~(-5) mol/L P_4与相同剂量单独作用的P_4和EGF比较各级卵泡数有统计学差异(P<0.05),与对照组比较无明显差异(P<0.05)。结论:在Waymouth培养体系中孕酮对原始卵泡生长有抑制作用,一定浓度的EGF能促进原始卵泡生长,而同时加入P_4和EGF,两者的作用有部分抵消。     

Clinical significance of amphiregulin and epidermal growth factor in colostrum

Background

Colostrum contains a wide variety of crucial nutritional elements including growth factors for newborn infants to adapt to the extrauterine environment.

Objective

To investigate the clinical significance of epidermal growth factor receptor ligands in milk during the first month of lactation.

Methods

The concentrations of epidermal growth factor (EGF), amphiregulin (AR) and transforming growth factor-?? (TGF-??) in milk sampled from a total of 31 normal mothers at days 1?C3, 5, and 30 postpartum were examined using ELISA.

Results

At days 1?C3, the concentration of EGF was extremely high [131.6?±?20.4 (mean?±?SEM) ng/ml] compared to that of AR (4,197.2?±?1,055.2?pg/ml) or TGF-?? (261.7?±?33.6?pg/ml), while the concentration of AR was significantly elevated compared to that of TGF-??. At days 5 and 30, the concentration of EGF was significantly elevated compared to that of AR or TGF-??. In 16 mothers among the same 31 subjects, samples were longitudinally obtained on days 1, 2, 5, and 30 postpartum. Concentrations of AR were higher on days 1 and 2 and rapidly declined to below 1?ng/ml on day 5, and were maintained at lower levels on day 30. Concentrations of EGF were high on day 1 (greater than 10?ng/ml) but gradually declined by days 2, 5, and 30. Concentrations of TGF-?? remained at lower levels of below 1?ng/ml throughout the lactation period from days 1 to 30.

Conclusion

These results suggested that EGF and amphiregulin in colostrum might contribute to the early stage of development of neonatal gastrointestinal function.     

Studies on transfer of vitamin K into human breast milk

We studied whether the administration of vitamin K to mothers could increase the concentration of vitamin K in breast milk and prevent idiopathic vitamin K deficient bleeding in breast-feeding infants. Sixty puerperal women were divided into three groups, the control group, Menaquinone-4 (MK-4) administered group and vitamin K1 administered group. We measured the concentrations of vitamin K1, MK-4 and MK-7 in maternal plasma and breast milk on the fourth day after delivery. In the MK-4 group, the concentrations of MK-4(2.13 ng/ml in plasma, 49.3 ng/ml in milk) were significantly higher than in the control group (0.28 ng/ml, 1.51 ng/ml). In the vitamin K1 group, the concentrations of vitamin K1 (49.0 ng/ml in plasma, 71.6 ng/ml in milk) were significantly higher than in the control group (1.17 ng/ml, 2.41 ng/ml). The concentration rates (milk/plasma ratio) of vitamin K1, MK-4 and MK-7 were 2.52, 5.43 and 0.52 in the control group, 1.60, 40.2 and 0.67 in the MK-4 group and 1.65, 10.8 and 0.71 in the vitamin K1 group, respectively. The concentration rate of MK-4 was higher than that of vitamin K1 and was increased by MK-4 administration. After delivery, the daily concentration of MK-4 in milk was increased from 1.69 ng/ml on the first day to 49.3 ng/ml on the fourth day in the MK-4 group. These results indicate that MK-4 is accumulated and concentrated into breast milk, and continuous MK-4 administration can increase the concentration of vitamin K in milk, preventing idiopathic vitamin K deficient bleeding in infants.