热休克蛋白在子宫内膜异位症中的表达及意义

目的 探讨热休克蛋白27（HSP27）和HSP70在子宫内膜异位症发病中的作用。方法 采用免疫组化链霉菌抗生素蛋白－过氧化物酶染色法（SP法），检测子宫内膜异位症中56例异位内膜与在位内膜（30例）中HSP27和HSP70的表达。采用原位杂交，检测HSP70mRNA的水平，并与正常子宫内膜相比较。结果 免疫组化结果显示，异位内膜组织中HSP27与HSP70均呈高表达，与正常内膜组的表达差异显著（P＜0．01），而且失去其在正常内膜组织中表达的周期性变化。卵巢子宫内膜异位症组（OEM）与子宫腺肌症组（AM）组中，HSP27和HSP70的表达无显著差异（P＞0．05）。原位杂交结果显示，HSP70 mRNA（分别为P＜0．01和P＜0．05）。HSP27与HSP70的表达水平，同OEM的严重性无关。结论 异位内膜组织中的HSP70基因被激活，转录增加。HSP27和HSP70呈高表达，可能在EM的发病中起重要作用。     

Localization and variable expression of G alpha(i2) in human endometrium and Fallopian tubes

BACKGROUND: Heterotrimeric G proteins take part in membrane-mediated cell signalling and have a role in hormonal regulation. This study clarifies the expression and localization of the G protein subunit G alpha(i2) in the human endometrium and Fallopian tube and changes in G alpha(i2) expression in human endometrium during the menstrual cycle. METHODS: The expression of G alpha(i2) was identified by Polymerase chain reaction (PCR), and localization confirmed by immunostaining. Cyclic changes in G alpha(i2) expression during the menstrual cycle were evaluated by quantitative real-time PCR. RESULTS: We found G alpha(i2) to be expressed in human endometrium, Fallopian tube tissue and in primary cultures of Fallopian tube epithelial cells. Our studies revealed enriched localization of G alpha(i2) in Fallopian tube cilia and in endometrial glands. We showed that G alpha(i2) expression in human endometrium changes significantly during the menstrual cycle, with a higher level in the secretory versus proliferative and menstrual phases (P < 0.05). CONCLUSIONS: G alpha(i2) is specifically localized in human Fallopian tube epithelial cells, particularly in the cilia, and is likely to have a cilia-specific role in reproduction. Significantly variable expression of G alpha(i2) during the menstrual cycle suggests G alpha(i2) might be under hormonal regulation in the female reproductive tract in vivo.     

Progressive rise in the expression of interleukin-6 in human endometrium during menstrual cycle is initiated during the implantation window

In order to be prepared for implantation, human endometriumundergoes a predictable series of proliferative and secretorychanges. Cytokines play an important role in regulation of thesechanges. Therefore, in this study, we immunolocalized the cytokine,interleukin-6 (IL-6), its receptor and the signal transducergp130 in human endometrium throughout the menstrual cycle. Duringthe entire menstrual cycle, the IL-6 receptor and gp130 werefound primarily in the endometrial glands and to a lesser extentin the stroma. The immunoreactivity of these proteins did notchange in endometrial cells during the entire menstrual cyclewith an exception of reduced immunoreactivity of gp130 in endometrialglands during menstrual phase. Immunostaining showed that immunoreactiveIL-6 was weakly expressed in human endometrium during the proliferativephase. Strong immunoreactivity for IL-6 appeared in endometriumduring the putative 'implantation window'. Expression was byfar most pronounced both in the glandular and surface epithelialcells. The amount of immunoreactive IL-6 in the epithelium progressivelyincreased during the secretory/menstrual phases. During thelate secretory phase, only stromal cells in the upper functionalisexhibited immunoreactivity for IL-6. Western blot analysis corroboratedthe immunohistochemical data. Human endometrial IL-6 consistedof a protein with an apparent mobility of 26 kDa. The immunoreactiveband of IL-6 was weak in the proliferative phase. The expressionof this protein increased progressively during the secretory/menstrualphases. The findings show a cell-specific pattern of distributionfor immunoreactive IL-6 in human endometrium. The menstrualcycle-dependent expression of IL-6 suggests that this cytokinemay play a role in changes in endometrium that prepare thistissue for implantation and menstrual shedding.     

The effect of hormone replacement therapy on the immunoreactive concentrations in the endometrium of oestrogen and progesterone receptor, heat shock protein 27, and human beta-lactoglobulin

We determined the expression of oestrogen receptor (ER), progesterone receptor (PR), heat shock protein 27 (HSP27) and human beta-lactoglobulin in the endometrium under hormone replacement therapy (HRT). The immunohistochemical expression during the late progestogenic phase of sequential HRT was compared semi-quantitatively and using image analysis, to the early, mid-, and late luteal phase of the physiological cycle. Under sequential HRT, smaller glands were positive for the ER but larger glands with more advanced secretory features were negative. ER expression was lower in the stroma under HRT, and the difference was statistically significant compared with the early luteal phase (P < 0.05). Expression of HSP27 under HRT was lower in the epithelium but higher in the stroma compared with the physiological luteal phase. Epithelial PR expression was lower under HRT compared with the early, but not the mid- or the late luteal phase. The number of PR-positive stromal cells under HRT was lower compared with the physiological cycle, and the difference was statistically significant in comparison with the early luteal phase (P < 0.05). The glandular area expressing human beta-lactoglobulin during the late progestogenic phase was statistically significantly higher compared with the early, but lower in comparison with the mid- or the late luteal phase (P < 0.05). The study demonstrates a sub-physiological progestogenic response superimposed on evidence of a hypo-oestrogenism, and a differential response in the epithelium and stroma.     

Expression and regulation of vascular endothelial growth factor ligands and receptors during menstruation and post-menstrual repair of human endometrium

Regeneration and growth of the human endometrium after shedding of the functional layer during menstruation depends on an adequate angiogenic response. We analysed the mRNA expression levels of all known vascular endothelial growth factor (VEGF) ligands and receptors in human endometrium collected in the menstrual and proliferative phases of the menstrual cycle. In addition, we evaluated the expression of VEGF-A, VEGF-R2 and NRP-1 at the protein level. Two periods of elevated mRNA expression of ligands and receptors were observed, separated by a distinct drop at cycle days (CDs) 9 and 10. Immunohistochemical staining showed that VEGF and VEGF-R2 were expressed in epithelial, stromal and endothelial cells. NRP-1 was mainly confined to stroma and blood vessels; only in late-proliferative endometrium, epithelial staining was also observed. Except for endothelial VEGF-R2 expression in CDs 6-8, there were no significant differences in the expression of VEGF, VEGF-R2 or NRP-1 in any of the cell compartments. In contrast, VEGF release by cultured human endometrium explants decreased during the proliferative phase. This output was significantly reduced in menstrual and early-proliferative endometrium by estradiol (E2) treatment. Western blot analysis indicated that part of the VEGF-A was trapped in the extracellular matrix (ECM). Changes in VEGF ligands and receptors were associated with elevated expression of the hypoxia markers HIF1alpha and CA-IX in the menstrual and early proliferative phases. HIF1alpha was also detected in late-proliferative phase endometrium. Our findings indicate that VEGF-A exerts its actions mostly during the first half of the proliferative phase. Furthermore, VEGF-A production appears to be triggered by hypoxia in the menstrual phase and subsequently suppressed by estrogen during the late proliferative phase.     

热休克蛋白70、90在子宫内膜癌中的表达

目的:探讨子宫内膜癌组织中热休克蛋白(HSP)70、90的表达及意义。方法:用免疫组化Envision法及图象分析仪,检测 30例正常子宫内膜、30例增生过长子宫内膜和53例子宫内膜癌中HSP70、HSP90的表达。结果:子宫内膜癌中HSP70表达的灰度值为(209.06±5.36),明显高于正常内膜[(145.21±4.09),P<0.01]和增生过长内膜[(148.59±4.23),P<0.01];子宫内膜癌中HSP90表达的灰度值为(166.98±5.71),明显低于正常子宫内膜[(208.57±31.14),P<0.05]和增生过长子宫内膜[(249.73±4.94),P<0.01]。子宫内膜癌中HSP70的表达随肿瘤病理分级的增加而增强(P<0.01),非内膜样癌(229.90±3.77)较内膜样癌表达强[(198.37±3.15),P<0.01];子宫内膜癌中HSP90表达随肿瘤病理分级的升高而表达减弱,非内膜样癌(140.21±3.22)较内膜样癌表达减弱[(176.59±2.79),P<0.01]。子宫内膜癌中HSP70、90的表达与肌层浸润深度、临床分期及淋巴结转移未见显著相关性(P>0.05)。结论:HSP70、90可能与子宫内膜癌的发生及预后有关。     

妊娠早期小鼠子宫内膜热休克蛋白70的免疫组化研究

目的 :研究妊娠早期小鼠子宫内膜热休克蛋白 70的表达。方法 :采用免疫组织化学方法。结果 :热休克蛋白 70主要存在于子宫内膜固有层的基质细胞及蜕膜细胞 ,内膜上皮、腺上皮中未见表达。与未孕小鼠相比 ,孕鼠热休克蛋白 70免疫反应阳性细胞显著增多 (P <0 .0 1 )且随妊娠日龄的增加而增加 (P <0 .0 1 )。结论 :热休克蛋白70可能参与了子宫内膜蜕膜反应中基质细胞的增殖 ,与蜕膜反应密切相关     

Chaperones in the parotid gland: localization of heat shock proteins in human adult salivary glands

Heat shock proteins (HSPs) are expressed or increased in response to various biological stresses. Moreover, these 'stress proteins' seem to be expressed by some cells living in physiological conditions. From then on, they could play an important physiological role in normal cell functioning. The best-known physiological role of these HSP proteins is to act as 'molecular chaperones'. In this context, we have investigated the immunohistochemical expression of HSP27, HSP70, HSP90 and HSP110 in 10 human adult salivary glands. To highlight the presence of RNAm encoding HSP70, an in situ hybridization was performed. In our material, HSP27 was strongly expressed in the cytoplasm of striated duct cells and in some myoepithelial cells. The same localization was less stained for HSP70 and HSP90. The immunocytochemical reaction was weak or negative for HSP110 in striated ducts. HSPs were not expressed in acinic cells. In situ hybridization gave a positive signal in striated ducts with a probe encoding HSP70. Epithelial cells of the striated ducts and myoepithelial cells expressed HSP27, HSP70 and HSP90. These HSPs probably act in part as molecular chaperones for protein synthesis, transport and for several interactions between HSPs and different proteins.     

Immunohistochemical localization of androgen receptor in the human endometrium, decidua, placenta and pathological conditions of the endometrium.

The immunohistochemical localization of the androgen receptor in the human endometrium at various stages of the menstrual cycle and post-menopausal period, in decidua and placenta of early pregnancy, and in several pathological conditions of the endometrium has been investigated. At any phase of the menstrual cycle, both endometrial glandular cells and endometrial stromal cells showed positive nuclear staining. Endometrial stromal cells of the functional layer showed stronger staining than those of the basal layer, but endometrial glandular cells of both layers showed the same staining intensity. There was little staining in myometrium. Even after menopause, endometrial glandular and stromal cells showed the same staining pattern as the basal layer of pre-menopausal endometrium and the staining intensity of endometrial stromal cells was weak. In decidua and placenta of early pregnancy, decidual and trophoblastic cells showed positive staining and there was no staining in the stromal cells of placenta. The expression of the androgen receptor was also detected in adenomyosis, endometriosis and endometrial carcinoma. Although the proliferation and differentiation of endometrium are mediated mainly by oestrogen and progesterone receptors, the androgen receptor may play some role in modulating these changes. These results suggest that it may be involved in both physiological and pathological changes of the endometrium.     

Inhibition by prostaglandin PGA1 on the multiplication of influenza virus is a dose-dependent effect

Cyclopentenone prostaglandins (PGs), are strong inhibitors of the multiplicative cycle of a wide variety of enveloped RNA and DNA viruses. Their antiviral activity is generally associated with alterations in the synthesis or maturation of specific virus proteins. In this report, we describe the effect of cyclopentenone PGA1 on the replication of influenza A virus Ulster 73 in LLC-MK2 cells. PGA1 was found to inhibit viral replication in a dose-dependent fashion and virus particle yield was reduced at a PGA1 concentration, which did not suppress protein synthesis in mock-infected cells. The kinetic of late viral protein synthesis was delayed in PGA1-treated cells till 10 h post-infection; after that period, viral polypeptide synthesis appeared to be similar in PGA1-treated as well as untreated cells both infected by Ulster 73 virus. This finding suggests that PGA1 might interfere with one or more events in the viral multiplicative cycle such as protein synthesis and assembly, correct insertion of virus polypeptides into the cell membrane and, or maturation of Ulster 73 virion particles. In particular, inhibition of viral replication in LLC-MK2 cells by PGA1 is accompanied by the induction of a cellular polypeptide of 70K molecular weight. We identified this cell protein as a heat shock protein (HSP) related to the inducible isoform of HSP 70, a polypeptide of 72K molecular weight. Induction of this polypeptide by PGA1 was found to be dose-dependent and a substantial accumulation could be seen at a PGA1 concentration that did not inhibit cell protein synthesis in uninfected cells. HSP 70 synthesis started after the beginning of PGA1 treatment and remained at the same level for at least 10 h, leading us to hypothesize that the delay of production of late Ulster 73 proteins could be the consequence of HSP 70 synthesis. These results suggest that HSP 70 could play a role in the antiviral activity of cyclopentenone PGA1 in LLC-MK2 cells.     

Immunohistochemical Localization of C9 Neoantigen and the Terminal Complement Inhibitory Protein CD59 in Human Endometrium

PROBLEM : Human endometrium expresses complement components, receptors, and regulatory proteins, many of which appear to be expressed in a hormone-dependent manner. Whether terminal complement components are also present in the endometrium is unknown. CD59, a broadly expressed protein that blocks association of C9 with C8 in the membrane attack complex, is localized in reproductive tissue to human spermatozoa, seminal plasma, amniotic fluid, and placenta. The present study examines human endometrium for the presence of CD59 and terminal complement proteins. METHOD : Endometrial biopsies were obtained from six normal women from various phases of the menstrual cycle and analyzed by immunohistochemistry, using MEM-43 anti-human CD59 and anti-human SC5b-9 murine monoclonal antibodies and the immunoperoxidase technique. RESULTS : Both CD59 protein and SC5b-9 (C9 neoantigen) were demonstrated to be present in endometrial glandular epithelium throughout the menstrual cycle. No specific staining was demonstrated in the stromal compartment. CONCLUSION : CD59 protein and terminal complement proteins are expressed in glandular epithelial cells of normal human endometrium, in both proliferative and luteal phases, suggesting that expression is not hormonally dependent. These analyses further support the presence of a functionally active complement system in normal human endometrium.     

Stressed stem cells: Temperature response in aged mesenchymal stem cells

Mesenchymal stem cells (MSCs) derived from young (6 week) and aged (56 week) Wistar rats were cultured at standard (37 degrees C) and reduced (32 degrees C) temperature and compared for age markers and stress levels. (ROS, NO, TBARS, carbonyls, lipofuscin, SOD, GPx, apoptosis, proteasome activity) and heat shock proteins (HSP27, -60, -70, -90). Aged MSCs display many of the stress markers associated with aging in other cell types, but results vary across marker categories and are temperature dependant. In young MSCs, culturing at reduced temperature had a generally beneficial effect: the anti-apoptotic heat shock proteins HSP 27, HSP70, and HSP90 were up-regulated; pro-apoptotic HSP60 was downregulated; SOD, GPx increased; and levels in ROS, NO, TBARS, carbonyl, and lipofuscin were diminished. Apoptosis was reduced, but also proteasome activity. In contrast, in aged MSCs, culturing at reduced temperature generally produced no 'beneficial' changes in these parameters, and can even have detrimental effects. Implications for tissue engineering and for stem cell gerontology are discussed. The results suggest that a 'hormesis' theory of stress response can be extended to MSCs, but that cooling cultivation temperature stress produces positive effects in young cells only.     

HSP70在早期妊娠大鼠子宫中的表达及己烯雌酚的诱导作用

目的探讨在早期妊娠大鼠子宫中热休克蛋白70(hearshockprotelns,HSP70)的表达变化以及己烯雌酚(dithylstilbestrol,DES)对子宫HSP70表达的诱导作用。方法采用免疫组织化学和图像分析技术研究正常大鼠未孕及已孕的子宫。结果①孕鼠子宫HSP70的阳性表达较未孕鼠明显增强,差异有显著性(P<0.01);②小剂量(1.25滋g/50g)大剂量(5滋g/50g)DES均可诱导大鼠子宫HSP70免疫反应阳性细胞数量显著增加(P<0.01)。结论热休克蛋白70在早期妊娠大鼠子宫内膜固有层的基质细胞及蜕膜内蜕膜细胞的表达明显增高,DES对子宫HSP70。表达具有明显诱导作用。     

Thioredoxin expression in the human endometrium during the menstrual cycle

Human thioredoxin, a cellular disulphide reducing protein, is known to be secreted by some types of cells and to display unique extracellular activities including modulation of cytokine actions and protection of the cell against damage from oxidative stress. This study has been undertaken to investigate the pattern of expression and tissue distribution of thioredoxin in human endometrium during the menstrual cycle. Immunohistochemical studies showed increased thioredoxin immunoreactivity in the glands of the secretory phase compared to those of the proliferative phase. Although the staining of thioredoxin was relatively intense in predecidual stromal cells, the most prominent staining of thioredoxin was present in both glands and stroma of the endometrium in the early secretory phase of the menstrual cycle. Northern hybridization analyses revealed that expression of thioredoxin mRNA in the endometrium of the early secretory phase was approximately 3-fold compared to the other phases of the menstrual cycle, consistent with the results of the immunohistochemical studies. These results suggest that both protein and gene expression of thioredoxin in the endometrium are menstrual cycle phase-specific and highly active in the phase of endometrial differentiation which occurs in preparation for implantation (early secretory phase of the menstrual cycle). Thioredoxin expressed in the early secretory phase of the menstrual cycle may be advantageous for blastocyst implantation.      

Dietary selenium supplementation alleviates immune toxicity in the hearts of chickens with lead-added drinking water

Lead (Pb) is an environmental pollutant and can damage organisms. Selenium (Se) can alleviate Pb poisoning. The present study aimed to investigate the alleviative effect of Se on Pb-induced immune toxicity in chicken hearts. One-hundred-and-eighty Hy-line male chickens were randomly divided into four groups at 7 days of age. The control group was offered a standard commercial diet (SD) and drinking water (DW); the Se group was offered SD supplemented with sodium selenite (SeSD) and DW; the Pb?+?Se group was offered SeSD and DW supplemented with lead acetate (PbDW); and the Pb group was offered SD and PbDW. Relative mRNA expression of inducible nitric oxide synthase (iNOS), interleukins (IL-2, IL-4, IL-6, IL-12β, IL-17 and IFN-γ), and heat shock proteins (HSP27, HSP40, HSP60, HSP70, and HSP90) were determined by means of quantitative real-time PCR. Relative protein expression of iNOS, HSP60, HSP70, and HSP90 was assessed, as well as nitric oxide (NO) content and iNOS activity in heart tissue. The results indicated a down-regulation of interleukin (IL)-2 and IFN-γ and an up-regulation of NO, iNOS, interleukins (IL-4, IL-6, IL-12β, IL-17), and heat shock proteins (HSP27, HSP40, HSP60, HSP70, and HSP90) in Pb-damaged hearts. Se alleviated all of the above Pb-induced changes. There were time-dependent effects on NO content, iNOS activity, and mRNA levels of iNOS, IL-2, IL-4, IL-6, IL-17, HSP27, HSP40, HSP60, HSP70, and HSP90 after Pb treatment in the chicken hearts. Se alleviated Pb-induced immune toxicity in the chicken hearts.     

Expression of heat shock proteins in osteosarcoma and its relationship to prognosis

The prognosis of osteosarcoma has been improved by chemotherapy. Heat shock proteins (HSPs) assist in folding proteins at posttranslation and degeneration under stress. We investigated the effect of HSPs on survival in osteosarcoma. Conventional osteosarcomas of the extremities from 70 patients aged 30 years or younger were used. Preoperational chemotherapy was performed in all cases. Tissues at surgery and biopsy were immunohistochemically stained with anti-HSP27, HSP47, HSP60, HSP70, HSP90alpha, HSP90beta, and p53 antibodies. We classified the cases in which more than 10% of tumor cells were positive into the overexpressing group. Overall survival was compared between the groups either overexpressing HSPs or not using Wilcoxon's test and Cox's proportional hazard model. The overexpression rate at biopsy was 22% (HSP27), 88% (HSP47), 66% (HSP60), 48% (HSP70(, 47% (HSP90alpha), 31% (HSP90beta), and 17% (p53), respectively. The rate at surgery was 33% (HSP27), 94% (HSP47), 60% (HSP60), 49% (HSP70), 28% (HSP90alpha), 40% (HSP90beta), and 17% (p53), respectively. HSP27 and p53 overexpression at biopsy had a negative prognostic value. HSP27 showed the strongest negative prognostic value in osteosarcoma. It is therefore important to investigate further its function in cellular regulation and drug resistance.     

Heat shock proteins, endothelin, and peripheral neuronal injury

Heat shock proteins (HSP) are important in neuroprotection after a variety of stresses or injuries. Both heat shock proteins and endothelin are upregulated after peripheral nerve injury, but HSP regulation after injury has not been systematically studied in peripheral nerve. The purpose of this study was to examine the regulation of small and large heat shock proteins after injury to rat sciatic nerve. Secondly, using a parallel tissue culture model for the sciatic nerve (PC12 cells), we examined potential regulation of heat shock proteins by endothelin. Western analysis of constricted, mobilized or unperturbed nerve was used to examine HSP abundance after injury. Semiquantitative PCR was used to examine heat shock message levels after nerve injury in the dorsal root ganglia. Cultured undifferentiated and differentiated PC12 cells were treated with endothelin, then western analysis of cytosol- and membrane-enriched fractions of these cells was used to examine heat shock protein regulation after endothelin treatment. Heat shock proteins are expressed at very low levels in unperturbed sciatic nerve. Constriction injury of the nerve results in increased expression of small and large heat shock proteins, but no upregulation of HSP message in corresponding dorsal root ganglia. Endothelin treatment of PC12 does not cause upregulation of heat shock proteins. Together these data show that a broad range of HSP is involved in endogenous response to peripheral nerve injury and deserve further study as potential neuroprotectants. Regulation of heat shock proteins after nerve injury is not likely due to endothelin signaling.