Immunohistological localisation of vascular endothelial growth factor in human endometrium.

Several polypeptide growth factors regulate epithelial and stromal development in endometrium under the influence of estrogen and progesterone, and thereby regulate growth and differentiation of endometrium during menstrual cycle. However, little is known about the angiogenic growth factors that may affect endometrial vasculature throughout each menstrual cycle. Vascular endothelial growth factor (VEGF) is suggestively an important angiogenic growth factor in the female reproductive tract. The aim of the present study was to immunolocalize and assess semi-quantitatively VEGF immunostaining in cells of proliferative phase (n = 3), secretory phase (n = 6) and hyperplastic (n = 6) human endometrial samples. VEGF concentrations were significantly higher in glandular (P < 0.001) and stromal (P < 0.01) compartments of proliferative stage endometrium compared with those in secretory stage and hyperplastic endometrial samples, with no difference in the scores for glandular and stromal compartments between secretory stage and hyperplastic endometrial samples. Generally, glandular expression of VEGF was higher as compared to stromal compartment. Thus, it appears that endometrial VEGF expression and concentration are enhanced by estrogen, and may be correlated with neovascularization and increased vascular permeability during late proliferative period. Additionally, there was no enhancement in VEGF expression in hyperplastic glands, suggesting that regulation of glandular growth and that of angiogenesis in human endometrium operate through different mechanisms.     

In vitro action of leukemia inhibitory factor (LIF) on mid-secretory stage endometrial stromal cells collected from hormone-simulated, ovariectomized monkey and maintained in three-dimensional primary culture

Leukemia inhibitory factor (LIF) is a pleiotropic cytokine that is known to play an important role in blastocyst implantation. The putative action of LIF in the regulation of uterine function has been examined using mid-secretory stage monkey endometrial stromal cells cultured on rat-tail collagen type I and treated with recombinant human LIF (rhLIF) or immunoneutralized LIF (in LIF) under serum-free condition. Long-term ovariectomized rhesus monkeys (n=8) underwent simulation of their menstrual cycles with steroid hormones and endometrial tissue samples were collected on cycle day 18; stromal cells were isolated and grown in primary culture on three-dimensional collagen matrix. Significant decline in cellular protein synthesis (P < 0.01) and cell proliferation index (P < 0.05) was observed in cells with increasing doses (0-1000 ng/ml) of rhLIF under serum-free in vitro condition. JAK1 expression in cultured cells increased (P < 0.01) in response to rhLIF as revealed from Western blot and confocal laser scanning microscopic examination, STAT1 and STAT2 expressions were unchanged, while pSTAT3 expression increased (P < 0.01) with increased concentration of rhLIF in culture medium. Autophosphorylation of JAK1 in endometrial stromal cells showed no change with increasing concentration (0.01 to 100 ng/ml) of rhLIF in vitro, but significant (P < 0.05) increase was observed with the time of exposure to rhLIF. Immunoneutralization of LIF or no addition of rhLIF to cultured cells led to significant (P < 0.01) increase in stromal cell proliferation index and significant (P < 0.01) decrease in the level of JAK1 and its autophosphorylation as compared to cells exposed to rhLIF alone. From the present set of experiments we conclude that rhLIF affects the physiological behaviour of monkey mid secretory stage endometrial stromal cells in vitro via the JAK-STAT signaling pathway.     

Effect of early luteal phase administration of a single dose mifepristone on immunohistochemical distribution of interleukin 1 alpha (IL-1 alpha) and transforming growth factor beta 1 (TGF-beta 1) in mid-luteal phase ovary of the rhesus monkey

A single low dose administration of a high affinity anti-progestin agent like mifepristone during the early luteal phase inhibits blastocyst implantation in human and non-human primates. Though it has been observed that luteal phase serum concentrations of estradiol and progesterone were not affected by the application of anti-nidatory dose of early luteal phase mifepristone suggesting that ovarian steroidogenic function is not compromised, it is nevertheless possible that ovarian physiology at the local tissue level is affected in this treatment schedule. In the present study, healthy, mature, proven fertile female rhesus monkeys were divided into two groups. Group 2 animals were treated with a single dose of mifepristone (2 mg/kg body weight), while group 1 animals were injected with vehicle (1:4 benzoyl benzoate: olive oil, v/v, s.c.) on day 2 post-ovulation. The morphological examination including that of vascularity, as well as, histometric determination of profiles of immunopositivity for IL-1alpha and TGF-beta1 in stromal, follicular and luteal compartments of mid-luteal phase ovaries from animals with or without a single, anti-nidatory dose of mifepristone applied on day 2 after ovulation failed to reveal any significant change between the two groups. Thus, it appears that early luteal phase administration of a single antinidatory dose of mifepristone does not affect the ovarian physiology in the treatment cycle.     

Effect of human chorionic gonadotropin (hCG) on expression of vascular endothelial growth factor a (VEGF-a) in human mid-secretory endometrial cells in three-dimensional primary culture

Several lines of evidence suggest that human uterine endometrial cells can bind human chorionic gonadotropin (hCG) which, in turn, influences the physiology of implantation stage endometrium. Vascular endothelial growth factor (VEGF) appears to be a candidate mediator in this process. However, our knowledge about hCG action on VEGF in human endometrial cells is very thin. In the present study, we have examined microscopically hCG binding to dissociated human endometrial cells collected from mid-luteal phase and maintained in three-dimensional primary co-culture on rat-tail collagen type I biomatrix and examined the effect of different concentrations (0, 1, 10, 100 and 1000 IU/ML) of hCG on VEGF expression and secretion by endometrial cells maintained in the above system. We report that both cytokeratin positive epithelial cells as well as vimetin positive stromal cells from human mid luteal phase endometrium could bind hCG and that their number increased (P < 0.01) steadily with time. Administration of hCG enhanced (P < 0.05) immunoreactive VEGF protein expression in dose dependent manner in endometrial cells retrieved from mid-luteal phase of cycle, and co-cultured in a three-dimensional cell culture system, but with no marked change in VEGF secretion. Collectively, it appears that hCG influences VEGF protein synthesis in human midluteal phase endometrial cells, but has little effect on post-translational regulation and secretion. From physiological homeostasis point of view, it is likely that synthesis and secretion of VEGF exhibits a modular and factorial regulation to achieve a fine tuning of this potent vasotropic agent in receptive stage endometrium.     

白血病抑制因子在原发性不孕患者黄体中期子宫内膜的表达及意义

目的检测白血病抑制因子(LIF)在原发性不孕、正常早孕蜕膜和正常子宫内膜中的表达,探讨其与原发性不孕及妊娠的关系。方法应用免疫组织化学S-P法检测LIF在30例原发性不孕患者的黄体中期子宫内膜、20例早孕流产蜕膜组织和15名正常黄体中期子宫内膜中的表达。结果 LIF在原发性不孕子宫内膜表达水平显著降低,与正常子宫内膜及宫内早孕蜕膜组织相比差异均有统计学意义(P<0.05)。结论 LIF在原发性不孕子宫内膜中低表达可能会导致子宫内膜接受性的降低,使着床失败引起不孕。     

沙利度胺对子宫内膜异位症的内膜间质细胞体外表达VEGF的影响

目的:观察沙利度胺(ThD)对子宫内膜异位症病人内膜间质细胞的增殖及其血管内皮生长因子(VEGF)表达的影响。方法:刮取子宫内膜异位症病人的子宫内膜组织,经体外分离、培养,用波形蛋白单抗进行免疫组化染色鉴定,应用四唑盐(MTS)分析肿瘤坏死因子α(TNF-α)和ThD对间质细胞生长的影响,以实时荧光定量PCR、Western Blot和定量ELISA分析在存在外源TNF-α的情况下,不同浓度ThD对间质细胞表达VEGF的影响。结果:TNF-α能促进子宫内膜间质细胞的增殖和分泌VEGF,而ThD能剂量依赖性抑制内膜间质细胞的增殖,并逆转TNF-α对VEGF的上调作用。结论:ThD可抑制体外培养的子宫内膜异位症内膜间质细胞的增殖及其VEGF的表达和分泌,提示ThD对于治疗子宫内膜异位症可能具有一定的作用。     

935MHz微波电磁辐射对小鼠着床期内膜细胞LIF蛋白表达的影响

目的:研究935MHz微波电磁辐射对小鼠着床期内膜细胞LIF蛋白表达的影响。方法:将受精后小鼠暴露于不同时间、不同强度的935MHz辐射波连续辐射3天,于受孕第4天取孕鼠子宫内膜,运用Western blot方法检测细胞内LIF蛋白的表达。结果:在辐射时间相同的情况下,高强度4h/d组和中强度4h/d组子宫内膜腺上皮细胞内膜LIF表达量明显下降;辐射强度相同时,中强度4h/d组与中强度2h/d组组比较、高强度4h/d组组与高强度2h/d组比较,LIF表达量也有明显减弱,显示辐射时间的长短可能影响LIF的表达,且微波对LIF表达的抑制作用具有蓄积效应。结论:935MHz微波电磁辐射通过影响孕鼠胚胎着床期子宫内膜细胞内LIF蛋白的表达,导致子宫内膜容受性发生改变,进而影响胚胎着床过程。     

同种异体骨髓基质干细胞移植对兔激素性股骨头缺血性坏死血清血管内皮生长因子的影响

目的研究同种异体骨髓基质干细胞（MSC）移植对兔激素性股骨头缺血性坏死（SAN—FH）血清血管内皮生长因子（VEGF）的影响。方法将45只健康新西兰兔随机分为3组：空白组、模型组和治疗组,每组各15只。模型组和治疗组,第1、3周马血清10ml／kg耳缘静脉注射,于第5周腹腔注射醋酸泼尼松龙7．5mg／kg,1次／d,连续3d;空白组,第1、3周生理盐水10ml／kg兔耳缘静脉注射,于第5周腹腔注射生理盐水1ml,1次／d,连续3d。治疗组于第6周行MSC移植。第10周测血清中VEGF水平。结果治疗组兔血清VEGF水平较模型组明显偏高（P〈0．01）。结论MSC移植使兔SANFH模型血清VEGF水平升高。     

Quantitative analysis of expression level of estrogen and progesterone receptors and VEGF genes in human endometrial stromal cells after treatment with nicotine

Cigarette smoke is a complex mixture of toxic chemicals, including nicotine, carbon monoxide, and several recognized carcinogens and mutagens. Nicotine has a direct disturbing influence on steroid hormones (estrogen and progesterone), which are essential components of the female reproductive system, but the effect of nicotine on the hormone receptors is not yet clear. The aim of this study was to elucidate the effect of nicotine on the expression of estrogen receptor (ER), progesterone receptor (PR), and vascular endothelial growth factor (VEGF) in endometrial stromal cells. Expression levels of PR, ER, and VEGF in human endometrial stromal primary cells treated with nicotine (0, 10^?11, 10^?8, and 10^?6?μM) for 24?h were measured by quantitative real-time PCR. MTT assay demonstrated that nicotine decreased cell viability in a dose-dependent manner. Real-time PCR data showed that despite decrease in ER expression in the nicotine-treated groups compared with the control, nicotine exerted an increased inhibitory effect on PR expression compared to that on ER expression. VEGF mRNA expression in nicotine-treated endometrial stromal cells was increased. The results from this study provide novel evidence for inhibitory effects of nicotine on steroid hormones receptor expression in human primary endometrial cells. Also, our data suggest that nicotine might have angiogenesis effects on these cells.     

猴脑选择性超深低温断血流复苏对碱性成纤维细胞生长因子血管内皮生长因子表达水平的影响

目的 观察猴脑选择性超深低温断血流复苏后碱性成纤维细胞生长因子(bFGF)、血管内皮生长因子(VEGF)表达变化,探索超深低温脑保护的可能机制.方法 成功建立脑选择性超深低温断血流复苏实验模型7只恒河猴,饲养到术后12周处死开颅取脑.低温组与常温组分别对大脑皮质脑组织切片用bFGF、VEGF抗体进行免疫组织化学染色,图像分析系统对切片进行分析,得出每张切片的平均灰度值,用SPSS 11.5软件进行统计学分析.结果 超深低温能使神经元和胶质细胞bFGF表达增高,抑制VEGF在神经元和胶质细胞中的表达.结论 超深低温能够抑制脑缺血后神经细胞的增生及肥大,增加bFGF的表达,抑制VEGF的表达,从而减少脑缺血对神经细胞的破坏作用.     

Macaque cytochromes P450: nomenclature, transcript, gene, genomic structure, and function

Monkeys, especially macaques, including cynomolgus (Macaca fascicularis) and rhesus monkeys (Macaca mulatta), are frequently used in drug metabolism studies due to their evolutionary closeness to humans. Recently, numerous cytochrome P450 (P450 or CYP) cDNAs have been identified and characterized in cynomolgus and rhesus monkeys and were named by the P450 Nomenclature Committee. However, recent advances in genome analysis of cynomolgus and rhesus monkeys revealed that some monkey P450s are apparently orthologous to human P450s and thus need to be renamed corresponding to their human orthologs. In this review, we focus on the P450s identified in cynomolgus and rhesus monkeys and present an overview of the identity and functional characteristics of each P450 cDNA in the CYP1-4 families. Information on the Japanese monkey (Macaca fuscata), African green monkey (Cercopithecus aethiops), and marmoset (Callithrix jacchus), primate species used in some drug metabolism studies, are also included. We compared the genomic structure of the macaque P450 genes to those of human and rat P450 genes in the CYP1-4 families. Based on sequence identity, phylogeny, and genomic organization of monkey P450s, we determined orthologous relationships of monkey P450s and, in this article, propose a revised nomenclature: CYP2B17/CYP2B30 to CYP2B6, CYP2C20/CYP2C74 to CYP2C8, CYP2C43/CYP2C83 to CYP2C9, CYP2C75 to CYP2C19, CYP2F6 to CYP2F1, CYP3A8/CYP3A21/CYP3A64 to CYP3A4, CYP3A66 to CYP3A5, and CYP4F45 to CYP4F2. The information presented in this review is expected to promote a better understanding of monkey P450 genes through comparative genomics and thereby make it more feasible to use monkeys in drug metabolism studies.     

香虎清妇炎颗粒对大鼠子宫内膜异位症ER、PR、VEGF的影响

目的 研究香虎清妇炎颗粒对实验性子宫内膜异位症大鼠ER、PR、VEGF的影响.方法 用大鼠子宫组织块自体腹壁移植的方法复制EMT模型.模型复制成功后,随机分为香虎清妇炎颗粒高、中、低剂量组(分别给予香虎清妇炎颗粒1.125 g·kg-1、0.75 g·kg-1及0.375 g·kg-1),阳性对照组(给予达那唑0.2 g·kg-1)及模型组;另取大鼠,切除同侧子宫组织,作为假手术对照组.各组大鼠每天用药1次,连续4周.用免疫组化方法检测香虎清妇炎颗粒对异位子宫内膜雌激素受体(ER)、孕激素受体(PR)、血管内皮生长因子(VEGF)表达的影响.结果 与模型组大鼠比较,香虎清妇炎颗粒各个剂量组的异位子宫内膜ER、PR(P<0.05)和VEGF(P<0.01)的表达明显降低.结论 香虎清妇炎颗粒可通过降低异位内膜中ER、PR、VEGF的表达水平,使异位内膜对E2、P的反应性下降,抑制异位内膜血管形成,从而抑制异位内膜的生长.     

IL-6、IL-10对骨髓基质细胞白血病抑制因子(LIF)的调节

目的 研究白细胞介素—6(IL-6)、白细胞介素—10(IL-10)对管髓基质细胞白血病抑制因子(LIF)表达的调节。方法 用酶联免疫吸附试验(ELISA)测定IL-6和IL-10作用下的LIF的表达水平，用逆转录—聚合酶使反应(PT—PCR)测定IL-6和IL-10作用下的LIF转逆子水平。结果 在IL-6作用下LIF水平明显降低(P＜0．05)；而IL-10对LIF水平没有明显影响。结论 IL-6能下调骨髓基质细胞LIF的表达，而IL-10对骨髓基质细胞LIF的表达无明显调控作用。     

MK、VEGF在子宫内膜癌中的表达及意义

目的 探讨子宫内膜癌组织中中期因子（MK）及血管内皮生长因子（VEGF）的表达,为子宫内膜癌的化学防治提供实验依据.方法 采用免疫组织化学S-P法检测49例子宫内膜癌和30例正常子宫内膜组织中MK和VEGF蛋的表达,并结合临床病理参数进行分析.结果 子宫内膜癌组织中MK和VEGF蛋白的表达均明显高于正常子宫内膜,MK蛋白的高表达与子宫内膜癌的临床分期及组织学分级相关性显著（r=1.55,P〈0.01;r=1.11,P〈0.05）,MK与VEGF 蛋白表达阳性者VEGF明显高于阴性者,二者呈正相关（r=1.76,P〈0.05）.结论 MK蛋白过表达可能参与了子宫内膜癌的发生发展,并与VEGF相关.     

血管内皮生长因子在乳腺癌中的表达及意义

目的研究血管内皮生长因子(VEGF)在乳腺癌中的表达情况及其与乳腺癌预后的关系,探讨其在乳腺癌中的预后价值。方法应用免疫组织化学法检测64例乳腺癌组织VEGF。结果VEGF的阳性表达率为63%。VEGF与肿瘤的大小、TNM分期、组织学分级、腋淋巴结转移呈正相关。结论VEGF为判断乳腺癌预后的有效指标。     

珊瑚羟基磷灰石结合血管内皮细胞生长因子修复兔骨缺损的实验研究

目的探讨珊瑚羟基磷灰石(CHA)结合血管内皮细胞生长因子(VEGF)修复骨缺损的效果,为其临床、科研应用提供实验依据。方法以CHA作为VEGF的可吸附性载体,制备成复合人工骨,将其植入兔尺骨中段10mm骨缺损处,以单纯CHA组,自体骨移植组和空白组作为对照,在术后2、4、8、12周,进行大体解剖、X线摄片、病理组织切片、生物力学测试等方法观察,研究各组骨愈合,血管化情况及力学强度。结果在2、4、8周病理组织切片及X线摄片显示骨缺损修复程度CHA/VEGF组明显优于自体移植骨组优于单纯CHA组,而空白组骨缺损处被纤维组织及肌组织等填充。生物力学测试显示术后12周CHA/VEGF组抗扭转强度明显优于自体骨移植组。结论CHA与VEGF结合有明显促进骨缺损修复的作用,强于自体骨移植及单纯CHA移植。二者协同,可用于骨缺损修复。     

Compartmentalized uterotrophic effects of tamoxifen, toremifene, and estradiol in the ovariectomized Wistar (Han) rat.

The comparative uterotrophic responses of ovariectomized Wistar (Han) rats to tamoxifen, toremifene, and 17beta-estradiol have been determined over a period of 72 h. Uterine wet weight; luminal epithelial cell hypertrophy; and BrdU labeling index in the different tissue compartments of the uterus, and the immunohistochemical expression of nuclear estrogen receptor alpha (nERalpha), and nuclear progesterone receptor (nPR) were examined. Luminal epithelial cell hypertrophy was produced by all three compounds to a similar degree. 17beta-Estradiol produced an increase in uterine wet weight due to fluid imbibition over the 3-day period, and an increase in DNA synthesis in the endometrial stromal and myometrial compartments of the uterus, as measured by increased BrdU incorporation. Estradiol increased the expression of nERalpha and nPR in the myometrium with time and decreased nERalpha levels from the overexpressed levels in control ovariectomized rat luminal epithelial cells. Tamoxifen and toremifene caused a smaller increase in uterine weight and the BrdU labeling index in the endometrial stroma and myometrium than did estradiol, and they increased the expression of nERalpha and nPR in the myometrium. Tamoxifen and toremifene differed from estradiol in that they did not decrease the expression of nERalpha in the luminal epithelial cells of the uterus. The response of PR expression was the same for tamoxifen, toremifene, and estradiol, and was therefore considered to be the most reliable indication of an estrogen-agonist effect in this study. The ability to distinguish differential, compartmentalized effects for agonists of estrogen action in the uterus will allow a better risk assessment for new pharmaceuticals that are used as breast cancer chemotherapeutic agents, especially where their use may also be associated with an increased risk of uterine cancers, in particular.     

活血消异方对子宫内膜异位症模型小鼠子宫内膜容受性的影响研究

目的 探讨子宫内膜异位症对子宫内膜容受性的影响机制以及活血消异方对子宫内膜异位症模型小鼠子宫内膜容受性的作用.方法 通过"皮下+腹腔"同系异体子宫内膜注射的方法建立子宫内膜异位症妊娠功能低下小鼠模型,假手术予等量0 Z.9%氯化钠溶液注射.建模成功后,随机分为中药组和模型组,中药组予活血消异方干预治疗,模型组和假手术组予等量0.5%羧甲基纤维素钠(CMC)灌胃.连续治疗3个周期后,3组分别与不育雄鼠合笼,次日清晨8:00观察阴栓,见栓者定为假孕第1天,于假孕第4天晚上10:30取材,运用Western blot法检测小鼠子宫内膜整合素αvβ3和LIF的蛋白表达.结果 模型组小鼠种植窗口期子宫内膜整合素αvβ3和LIF的蛋白表达量较假手术组显著降低,差异有统计学意义(P<0.05);中药组小鼠种植窗口期子宫内膜整合素αvβ3和LIF的蛋白表达量较模型组显著升高,差异有统计学意义(P<0.05),但均低于假手术组(P<0.05).结论 子宫内膜异位症对子宫内膜容受性存在不良影响.活血消异方能够显著提高小鼠围着床期子宫内膜着床因子整合素αvβ3和LIF的蛋白表达量,从而改善子宫内膜异位症模型小鼠子宫内膜容受性.     

三七总皂苷对流产大鼠子宫出血的影响及子宫内膜 修复作用机制的研究

目的 探究三七总皂苷对流产大鼠子宫出血的影响及可能的作用机制。方法 取未怀孕雌性大鼠10只 为空白对照组,受孕的雌性大鼠采用米非司酮（8.3 mg/kg）和米索前列醇（100 μg/kg）诱导建立异常子宫出血（AUB）模 型,随机分为模型组、阳性组（断血流片,0.45 g/kg）、三七总皂苷低（25 mg/kg）、中（50 mg/kg）、高（100 mg/kg）剂量组, 每组10只。阳性组、三七总皂苷各剂量组分别给予相应剂量的药物灌胃,1次/d,共7 d。紫外分光光度法测定大鼠 子宫出血量;酶联免疫吸附试验（ELISA）检测血浆肿瘤坏死因子（TNF）-α、白细胞介素（IL）-6、雌二醇（E2）、孕酮 （P）、6-酮前列腺素F1α（6-keto-PGF1α）以及血栓素B2（TXB2）水平;HE、Masson染色观察子宫内膜的病理损伤;免 疫荧光染色检测子宫内膜组织中微血管密度（MVD）;实时荧光定量PCR（qPCR）检测子宫组织血管内皮生长因子 （VEGF）、基质金属蛋白酶（MMP）-2、MMP-9 mRNA 的表达。结果 与空白对照组比较,模型组大鼠血浆 6-ketoPGF1α、TNF-α、IL-6水平、子宫内膜组织的纤维化面积比、MMP-2和MMP-9 mRNA表达增加,TXB2、E2、P水平、子 宫内膜厚度、腺体数量、子宫内膜MVD、VEGF mRNA表达降低（P＜0.05）;与模型组比较,三七总皂苷高剂量组大鼠 子宫出血量、血浆6-keto-PGF1α、TNF-α、IL-6水平、子宫内膜组织的纤维化面积比、MMP-2和MMP-9 mRNA降低, TXB2、E2水平、子宫内膜厚度、腺体数量、子宫内膜MVD、VEGF mRNA表达增加（P＜0.05）。结论 高剂量三七总皂 苷可治疗流产大鼠子宫出血,其作用机制可能与抑制过度纤维化和炎症,上调VEGF的表达,促进血管重塑,进而促 进子宫内膜修复有关。     

C型子宫内膜种植窗期组织形态学特征

目的 观察C型子宫内膜种植窗期形态学的特征.方法 应用组织形态学方法和图像分析技术,观察、分析C型子宫内膜10例与A型子宫内膜10例(对照组)的组织形态变化.结果 C型子宫内膜发育不良,其腺体面积、腺体周长、腺体最大直径、腺腔面积、腺腔周长、腺腔最大直径、腺上皮细胞总面积均小于A型子宫内膜,C型子宫内膜组织结构构成比包括:腺体面积、腺腔面积、腺上皮细胞总面积和腺/间比值均低于对照组,而间质面积大于对照组.结论 (1)种植窗期子宫内膜腺体发育不良可能是C型子宫内膜患者种植率低下的重要原因.(2)黑白B超测量C型子宫内膜厚度预测其容受性有一定局限性.