Selective progesterone receptor modulator asoprisnil down-regulates collagen synthesis in cultured human uterine leiomyoma cells through up-regulating extracellular matrix metalloproteinase inducer

BACKGROUND: A recent clinical trial demonstrated that selective progesterone receptor modulator asoprisnil is effective in reducing uterine leiomyoma volume. We investigated the effects of asoprisnil in vitro on the expression of the extracellular matrix (ECM)-remodeling enzymes and collagens in cultured leiomyoma and matching normal myometrial cells. METHODS: The expression of extracellular matrix metalloproteinase inducer (EMMPRIN), matrix metalloproteinases (MMPs), tissue inhibitors of MMP (TIMPs) and collagens were assessed by western blot analysis. RESULTS: Untreated cultured leiomyoma cells had significantly lower EMMPRIN (P < 0.05), MMP-1 (P < 0.05) and membrane type 1-MMP (MT1-MMP) (P < 0.01) protein contents, but significantly higher TIMP-1 (P < 0.05), TIMP-2 (P < 0.01), type I (P < 0.05) and type III (P < 0.01) collagen protein contents compared with untreated cultured myometrial cells. Treatment with asoprisnil at concentrations > or =10(-7) M for 48 h significantly (P < 0.05) increased EMMPRIN, MMP-1 and MT1-MMP protein contents, and decreased TIMP-1 (P < 0.05), TIMP-2 (P < 0.01), type I (P < 0.01) and type III (P < 0.05 at 10(-7) M; P < 0.01 at 10(-6) M) collagen protein contents in cultured leiomyoma cells compared with control cultures. However, asoprisnil treatment did not affect the protein contents of ECM-remodeling enzymes and collagens in cultured myometrial cells. CONCLUSIONS: These results suggest that asoprisnil may reduce collagen deposit in the ECM of cultured leiomyoma cells through decreasing collagen synthesis and enhancing the expression of EMMPRIN, MMPs and TIMPs without comparable effects on cultured myometrial cells.     

Progesterone receptor modulator CDB-2914 down-regulates vascular endothelial growth factor, adrenomedullin and their receptors and modulates progesterone receptor content in cultured human uterine leiomyoma cells

BACKGROUND: This study was conducted to evaluate the effects of graded concentrations (10(-8), 10(-7) and 10(-6) M) of progesterone receptor (PR) modulator CDB-2914 on the protein contents of PR, of vascular endothelial growth factor (VEGF), adrenomedullin (ADM) and their receptors in cultured human uterine leiomyoma and matching myometrial cells. METHODS: PR-A, PR-B, VEGF-A, VEGF-B, VEGF receptor (VEGFR)-1, VEGFR-2, ADM and ADM receptor (ADMR) contents were assessed by Western blot analysis. RESULTS: Treatment with 100 ng/ml progesterone increased VEGF-A, VEGF-B and ADM contents in cultured leiomyoma cells and normal myometrial cells. The concomitant treatment with 10(-6) M CDB-2914 significantly decreased the progesterone-induced VEGF-A, VEGF-B and ADM contents in cultured leiomyoma cells but not in normal myometrial cells. CDB-2914 treatment alone decreased VEGFR-1, VEGFR-2 and ADMR contents in cultured leiomyoma cells but not in normal myometrial cells. CDB-2914 treatment increased PR-A and decreased PR-B contents in cultured leiomyoma cells in a dose-dependent manner compared with untreated cultures, whereas no significant changes in PR isoform contents were observed in normal myometrial cells. CONCLUSIONS: These results suggest that CDB-2914 down-regulates VEGF, ADM and their receptor contents and modulates PR isoform contents in cultured leiomyoma cells in a cell-type-specific manner.     

Luteal phase dose-response relationships of the antiprogestin CDB-2914 in normally cycling women

BACKGROUND: Progesterone receptor modulators have potential therapeutic use in progesterone-dependent conditions such as endometriosis, fibroids and induction of labour. The synthetic steroid CDB-2914 binds to the progesterone and glucocorticoid receptors. In animals it has antiprogestational activity at doses 50-fold less than those required for antiglucocorticoid effects. METHODS AND RESULTS: We evaluated the biological activity, blood levels and safety of CDB-2914 at escalating single doses, in 36 normally cycling women at mid-luteal phase. CDB-2914 at doses of 1-100 mg did not change luteal phase length, but after 200 mg, all women had early endometrial bleeding. Four women with early menses had concurrent functional luteolysis (one at 10, 50, 100 and 200 mg). There were no biochemical or clinical signs of toxicity, and no effect on urinary cortisol or circulating thyroxine, prolactin, adrenocorticotrophic hormone or renin levels. Higher serum equivalents of CDB-2914 were observed by radioimmunoassay than by high performance liquid chromatography detection, indicating a considerable contribution of metabolites. CONCLUSIONS: Mid-luteal administration of CDB-2914 antagonizes progesterone action on the endometrium, in a dose-dependent fashion, without apparent antiglucocorticoid effects. Further study of CDB-2914 is needed to determine its clinical role.     

Progesterone down-regulates insulin-like growth factor-I expression in cultured human uterine leiomyoma cells

BACKGROUND: Insulin-like growth factor-I (IGF-I) plays crucial roles in uterine leiomyoma cell growth through stimulating proliferation and inhibiting apoptosis. The present study was conducted to elucidate whether progesterone affects IGF-I and its receptor expression in cultured leiomyoma cells. METHODS: Isolated leiomyoma cells were subcultured in Phenol Red-free Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum for 120 h and then stepped down to serum-free conditions for an additional 48 h in the presence or absence of 17beta-estradiol (E(2)) (10 ng/ml) or progesterone (100 ng/ml). IGF-I and its receptor mRNA and immunoreactive IGF-I in the cultured cells were assessed by quantitative RT-PCR with Southern blot analysis and by radioimmunoassay with Seppak C18 chromatography, respectively. The presence of estrogen receptor (ER) and progesterone receptor (PR) in cultured leiomyoma cells was immunocytochemically examined. RESULTS: Both treatment with progesterone alone and treatment with E(2) and progesterone combined significantly decreased IGF-I mRNA and protein expression in cultured leiomyoma cells compared with that in untreated cultures, but treatment with E(2) alone did not. IGF-I receptor mRNA expression in those cells was not affected by treatment with either E(2) or progesterone. Immunocytochemical analysis revealed that PR protein expression in cultured leiomyoma cells maintained in a serum-free condition for 48 h whereas ER protein expression in the cells remarkably decreased after 24 h culture under the serum-free condition. CONCLUSIONS: The present study provided evidence for the first time that progesterone down-regulates IGF-I mRNA and protein expression in cultured leiomyoma cells without affecting IGF-I receptor mRNA expression.     

Concentration-dependent effects of a selective estrogen receptor modulator raloxifene on proliferation and apoptosis in human uterine leiomyoma cells cultured in vitro

BACKGROUND: This study was conducted to elucidate the effects of raloxifeneon proliferation and apoptosis in cultured human uterine leiomyomacells. METHODS: The monolayer cultures were treated with graded concentrations(10^–9, 10^–8 and 10^–7 M) of raloxifeneand 10^–7 M 17-estradiol (E₂). Cell viability, percentageof proliferating cell nuclear antigen (PCNA)-positive cells,percentage of terminal deoxynucleotidyl transferase-mediated2'-deoxyuridine 5'-triphosphate nick-end labelling (TUNEL)-positivecells and the expression of PCNA and Bcl-2 proteins were assessedby 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxylphenyl)-2-(4-sulphophenyl)-2H-tetrazolium assay, immunocytochemistry, TUNEL assay and western blotanalysis, respectively. RESULTS: Compared with untreated cultures, the number of viable culturedcells, percentage of PCNA-positive cells and PCNA protein expressionwere significantly decreased by treatment with 10^–9 Mraloxifene, but increased by treatment with either 10^–8 Mor 10^–7 M raloxifene. In contrast, the percentageof TUNEL-positive cells was significantly increased and Bcl-2protein expression was significantly decreased by treatmentwith 10^–9 M raloxifene, whereas they were not affectedby treatment with either 10^–8 or 10^–7 M raloxifene. CONCLUSIONS: In cultured leiomyoma cells, low concentration (10^–9 M)of raloxifene may inhibit the growth of leiomyoma cells, whereashigh concentrations (10^–8 M, 10^–7 M) ofraloxifene may promote their growth.     

Progesterone withdrawal causes endothelin release from cultured human uterine microvascular endothelial cells

BACKGROUND: Current theories on the physiology of menstrual bleeding in humans offers an explanation for the shedding of the endometrium as a result of a breakdown of the extracellular matrix due to an inflammatory reaction. The link between the fall in progesterone levels and these events is not clear. Neither has an explanation been presented for the vasoconstriction in the coiled arteries occurring during menses. We have hypothesized a chain of events where the fall in progesterone levels induces an upregulation of the thrombin receptor in the small uterine arteries leading to an increased thrombin response and subsequent endothelin release. METHODS: Endothelial cells from human umbilical cord (HUVECs) and from human small uterine arteries (UtMVECs) were cultured under conditions partly resembling the female hormonal cycle with progesterone withdrawal. RESULTS: Following progesterone increase and subsequent withdrawal, we found an increased production of thrombin receptor and an increased release of endothelin from UtMVECs compared with HUVECs. CONCLUSION: Endothelin release in response to progesterone withdrawal in UtMVECs can offer an explanation for the vasoconstriction seen in the coiled arteries during menses in humans.     

A novel selective progesterone receptor modulator asoprisnil (J867) down-regulates the expression of EGF, IGF-I, TGFbeta3 and their receptors in cultured uterine leiomyoma cells

BACKGROUND: This study was conducted to evaluate the effects of a novel selective progesterone receptor modulator (SPRM) asoprisnil on the expression of growth factors and their receptors and on growth factor-induced proliferation of cultured uterine leiomyoma and matching myometrial cells. METHODS: The expression of epidermal growth factor (EGF), insulin-like growth factor-I (IGF-I) and transforming growth factor (TGFbeta3) was assessed by immunocytochemistry and semi-quantitative RT-PCR. The expression of phosphorylated EGF receptor (p-EGFR), IGF-I receptor alpha subunit (IGF-IRalpha) and phosphorylated TGFbeta receptor type II (p-TGFbeta RII) was assessed by Western blot analysis. Cell proliferation was assessed by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium assay. RESULTS: Treatment with 10(-7) M asoprisnil decreased EGF, IGF-I and TGFbeta3 mRNA and protein expression as well as p-EGFR, IGF-IRalpha and p-TGFbeta RII protein expression in leiomyoma cells cultured for 72 h. EGF (100 ng/ml), IGF-I (100 ng/ml) and TGFbeta3 (10 ng/ml) increased the number of viable leiomyoma cells cultured for 72 h, whereas the concomitant treatment with 10(-7) M asoprisnil antagonized the growth factor-induced increase in leiomyoma cell proliferation. In cultured myometrial cells, however, asoprisnil affected neither the growth factor and their receptor expression nor the cell proliferation. CONCLUSION: Asoprisnil inhibits the expression of EGF, IGF-I, TGFbeta3 and their receptors in cultured leiomyoma cells without affecting their expressions in myometrial cells.     

PPARs配体对巨噬细胞、泡沫细胞细胞外基质金属蛋白酶诱导因子表达的影响

目的：观察PPAR α、γ配体对巨噬细胞、泡沫细胞细胞外基质金属蛋白酶诱导因子(EMMPRIN)表达的影响。方法：体外诱导THP-1单核细胞转化为巨噬细胞、泡沫细胞，分别加入PPAR α配体氯贝特（clofibrate）、PPARγ配体吡格列酮(pioglitazone)共同培养，应用Real-time RT-PCR和Western blotting测定巨噬细胞、泡沫细胞中EMMPRIN基因和蛋白表达，ELISA测定细胞培养上清液MMP-9浓度，Zymgraphy法测定MMP-9活性。结果：氯贝特和吡格列酮均能显著抑制巨噬细胞和泡沫细胞EMMPRIN的表达，此抑制作用与PPAR α、γ配体抑制MMP-9分泌及活性的趋势一致。结论：PPAR α、γ配体均可抑制巨噬细胞、泡沫细胞EMMPRIN的表达，下调EMMPRIN可能是PPARs配体抑制粥样斑块局部MMPs产生的机制之一。     

The influence of extracellular matrix proteins on T-cell proliferation and apoptosis in women with endometriosis or uterine leiomyoma

PROBLEM: Interactions between the extracellular matrix (ECM) and peripheral blood T cells in women with endometriosis and leiomyoma are hardly unknown. We have investigated the influence of two major ECM components, collagen IV (C-IV) and fibronectin (Fn), on T-cell proliferation and apoptosis in women with endometriosis and uterine leiomyoma. beta1 integrin expression, responsible for interactions with ECM proteins, was also studied. METHOD OF STUDY: Peripheral blood lymphocytes were obtained from 53 women (17 with uterine leiomyomas, 18 with endometriosis, and 18 from healthy donors). T cells were exposed to ECM proteins co-immobilized with monoclonal antibody anti-CD3 for 72 hr. Apoptosis and S phase of the cell cycle of the T cells were studied by DNA analysis using flow cytometry. The proliferation of T cells was evaluated by MTT assay. The percentage of CD3+ cells expressing CD29 (beta1 integrin chain) was evaluated by double-color flow cytometry. Results were analyzed statistically using the Mann-Whitney test. RESULTS AND CONCLUSIONS: (1) A general increase in the percentage of T cells in S phase could be seen in women with endometriosis and uterine leiomyoma in all culture conditions what may suggest general activation of T cells. (2) A significant increase in the percentage of cells in S phase was shown only in the case of T cells exposed to anti-CD3 + C-IV in both women with uterine leiomyoma and endometriosis. (3) However, no apoptotic cells were observed. (4) T cells from patients with uterine leiomyoma exhibited significantly increased level of proliferation after culture with anti-CD3 + C-IV. (5) More T cells expressed beta1 integrin in women with endometriosis or uterine leiomyoma than in healthy donors. Our data may suggest that increased beta1 integrin expression may enhance T-cell-ECM interactions, which may be responsible for the increased proliferation of T cells but not for apoptosis. Therefore, it is possible that interactions of T cells with ECM proteins, especially with C-IV, may contribute to the pathogenesis of endometriosis and uterine leiomyoma.     

非小细胞肺癌中EMMPRIN和HGF的表达及其对淋巴结转移和预后的影响

目的： 研究细胞外基质蛋白酶诱导因子(EMMPRIN)和肝细胞生长因子(HGF)在非小细胞肺癌（NSCLC）组织中的表达及其临床意义。方法： 免疫组织化学方法检测77例NSCLC组织中EMMPRIN与HGF的表达，分析其与患者吸烟情况、肿瘤大小、组织学类型、组织分化程度、淋巴结转移和预后的关系。 结果：77例NSCLC组织中EMMPRIN与HGF的阳性表达率分别为68%和44%。EMMPRIN与HGF的表达均与淋巴结转移呈正相关（r=0.371和0.339，P＜0.01），与患者术后生存时间呈负相关（P＜0.01）。EMMPRIN与HGF的表达均与患者吸烟、肿瘤大小、组织学类型和组织分化程度无关（P＞0.05）。EMMPRIN与HGF的表达之间呈正相关（r=0.281,P＜0.01）。结论: EMMPRIN和HGF的表达与NSCLC的淋巴结转移和预后密切相关，它们的高表达提示NSCLC患者预后不良。     

细胞外基质金属蛋白酶诱导因子和内皮素转化酶对非小细胞肺癌患者术后淋巴结转移和预后的影响

目的： 研究细胞外基质金属蛋白酶诱导因子(EMMPRIN)和内皮素转化酶(ECE)在非小细胞肺癌（NSCLC）组织中表达的临床意义。 方法： 用免疫组织化学方法检测77例NSCLC组织中EMMPRIN和ECE的表达情况，分析其与肿瘤大小、癌的组织学类型、组织分化程度、淋巴结转移和预后的关系。 结果： 77例NSCLC组织中EMMPRIN和ECE的阳性表达率分别为66%和45%。EMMPRIN和ECE的表达均与淋巴结转移呈正相关（均P＜0.01，r=0.371和0.467），两者表达阳性组术后生存期均明显短于阴性组（均P＜0.01）。NSCLC组织中EMMPRIN和ECE的表达之间呈正相关（P＜0.05，r=0.243）。EMMPRIN和ECE的表达与肿瘤大小、癌组织学类型和组织分化程度均无相关性（P>0.05）。 结论： EMMPRIN和ECE的表达与NSCLC的淋巴结转移和预后密切相关，它们的高表达提示非小细胞肺癌患者预后不良。     

Comparative effects of heparin-binding epidermal growth factor-like growth factor on the growth of cultured human uterine leiomyoma cells and myometrial cells

BACKGROUND: The objective of this study was to investigate the comparative effects of heparin-binding epidermal growth factor-like growth factor (HB-EGF) on the growth of cultured human leiomyoma cells and myometrial cells. METHODS: Isolated cells were subcultured in Phenol Red-free Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum for 120 h and then stepped down to serum-free conditions for an additional 24 and 48 h in the presence or absence of graded concentrations of HB-EGF (0.1, 1, 10 and 100 ng/ml). These cells were used for immunocytochemical analysis for Ki67, western blot analysis for proliferating cell nuclear antigen (PCNA) and human EGF receptor (HER1), and TdT (terminal deoxynucleotidyl transferase)-mediated dUDP nick-end labelling (TUNEL) assay. RESULTS: Treatment with HB-EGF at concentrations >1 ng/ml significantly increased the Ki67-positive rate of cultured leiomyoma cells and myometrial cells. Treatment with HB-EGF also resulted in a dose-dependent increase in PCNA expression in both cells compared with untreated control cultures. A significant increase in PCNA expression in cultured myometrial cells was noted following treatment with HB-EGF at concentrations >1 ng/ml, whereas an increase in PCNA expression in cultured leiomyoma cells was noted following treatment with HB-EGF at concentrations >10 ng/ml. HER1 expression was significantly higher in untreated myometrial cells than in untreated leiomyoma cells. A significant increase in HER1 expression in myometrial cells was observed when treated with HB-EGF at concentrations >10 ng/ml, whereas a significant increase in HER1 expression in leiomyoma cells was noted only by the treatment with HB-EGF at concentrations >100 ng/ml. Treatment with HB-EGF decreased the TUNEL-positive rate of those cells with no significant differences between the two cell types. CONCLUSIONS: The results obtained suggest that HB-EGF plays a role in stimulating the proliferation of leiomyoma cells and myometrial cells and in inhibiting apoptosis of those cells through augmentation of HER1 expression. Since the proliferative potential of myometrial cells responded better to HB-EGF than that of leiomyoma cells, HB-EGF may play a more vital role in myometrial growth than leiomyoma growth.     

罗格列酮对人子宫肌瘤细胞凋亡和雌激素受体表达的影响

目的:探讨罗格列酮对人子宫肌瘤细胞增殖和凋亡的影响及可能机制.方法:不同浓度的罗格列酮(10-8,10-7和10-6 mol/L)处理原代培养的人子宫肌瘤细胞24,48和72 h.用四甲基偶氮唑盐(methyl thiazolyl tetrazolium,MTT)法测定细胞生长曲线,用流式细胞术测定细胞凋亡率,采用放射...     

Clinical implications of coexpression of growth arrest-specific gene 6 and receptor tyrosine kinases Axl and Sky in human uterine leiomyoma

The expression of Gas6, the protein product of the growth arrest-specific gene 6 (gas6), a member of the vitamin K-dependent protein family, and the receptor tyrosine kinases Axl and Sky and their mRNAs in uterine leiomyoma and normal uterine myometrium tissues were investigated by competitive RT-PCR-Southern blot analysis using recombinant RNA and immuno histochemical analysis respectively. There was no significant difference between the histoscores and levels of Sky mRNA in uterine leiomyoma and normal uterine myometrium, although the levels of Gas6 and Axl mRNAs in uterine leiomyoma were significantly higher than in normal uterine myometrium in each case. It is suggested that Gas6 and Axl signal transduction is aberrantly stimulated in uterine leiomyoma, possibly related to its growth.     

Cytokines regulate matrix metalloproteinases in human uterine endometrial fibroblast cells through a mechanism that does not involve increases in extracellular matrix metalloproteinase inducer

PROBLEM: Endometriosis is the presence of ectopic uterine endometrial tissue in the peritoneal cavity. Peritoneal fluid samples of women with endometriosis show elevated interleukin-1 (IL-1)beta, tumor necrosis factor (TNF)-alpha and transforming growth factor (TGF)-beta(1) levels, indicating that an altered immune system may play an important role in the pathogenesis of endometriosis. The invasion of ectopic endometrium into peritoneal mesothelium requires matrix metalloproteinases (MMPs) for tissue remodeling. Several MMPs are differentially expressed in human uterine endometrium with menstrual endometrium showing the highest level of expression. MMPs are stimulated by cytokines and also by the protein Extracellular Matrix Metalloproteinase Inducer (EMMPRIN). METHOD OF STUDY: To determine the role of cytokines in ectopic endometrial invasion, we investigated whether cytokines could regulate MMP production by endometrial fibroblast cells and whether this stimulation occurred through an effect on EMMPRIN expression. Human uterine fibroblasts (HUF) were treated with IL-1beta, TGF-beta(1) and TNF-alpha in a dose dependent and time dependent manner (C, 0.1, 1, 10 ng/mL IL-1beta or TGF-beta(1); C, 2, 10, 50 ng/mL TNF-alpha) for 0, 6, 12, and 24 hr. Cell conditioned medium samples were collected and concentrated at each timepoint for immunoblot analysis. Cellular RNA was collected for real time PCR analysis of MMPs-1, -2, -3 and EMMPRIN mRNA levels. RESULTS: Our results showed that IL-1beta stimulated MMP-1 protein secretion and mRNA levels in a time dependent manner (P < 0.05), MMP-2 mRNA in a time dependent manner and MMP-3 in a time and dose dependent manner. TNF-alpha stimulated MMP-1 and -3 protein secretion in a time dependent manner and stimulated MMP-1, -2 and -3 mRNA levels in a time dependent manner (P < 0.05). Neither IL-1beta nor TNF-alpha treatment affected MMP-2 protein secretion. TGF-beta(1) inhibited MMP-1 and MMP-2 mRNAs at the highest treatment dose after 24 hr but there was no effect on protein secretion. TGF-beta(1) exerted no effect on MMP-3 mRNA or protein secretion (P < 0.05). Neither of the cytokines affected EMMPRIN protein or mRNA levels but the 10 ng/mL TGF-beta(1) treatment did cause a reduction in EMMPRIN mRNA levels. CONCLUSIONS: These data show that elevated cytokines may play a role in the establishment of ectopic endometrium in the peritoneal cavity by stimulating MMPs to remodel the mesothelial lining of the peritoneum thus allowing for tissue invasion. The stimulation of MMPs by cytokines occurred without any change in EMMPRIN expression whereas the inhibitory effect of TGF-beta(1) involved a reduction in EMMPRIN mRNA levels.     

Progesterone receptor expression in human decidua and fetal membranes before and after contractions: possible mechanism for functional progesterone withdrawal

In humans, progesterone levels are sustained before the onset of labour. Therefore, the mechanism for parturition that has been proposed for humans is 'functional' progesterone withdrawal. Immunohistochemical staining for the progesterone receptor (PR) was positive in the decidua with a decline after contractions began. Western blot analysis revealed a number of PR isoforms expressed in the decidua, with the PR-B form being dominant. After contractions began, all PR isoforms decreased sharply. PR-B and PR-A decreased by 85.8% +/- 6.7 and 78.2% +/- 7.1, respectively (P < 0.001). Incubation of decidua with Prostaglandin F2alpha 1.0 microg/ml decreased the expression of all forms of PR isoforms. PR-B was reduced by 64% +/- 6.09 (P < 0.01); PR-A was reduced by 77% +/- 5.9 (P < 0.05), while PR-C was reduced by 80% +/- 7.24 (P < 0.05). Progesterone (80 microg/ml) increased the PR-B, PR-C the 45 and 36 kDa isoforms to 150% +/- 7.89, 210% +/- 12.4, 270% +/- 9.7 and 216% +/- 13.5, respectively (P < 0.05). In immunohistochemical studies, the PR was not identified in the amnion or in the chorion, regardless of the presence or absence of contractions. Western blot analysis demonstrated that PR-C (60 kDa) and the 36 kDa isoforms were dominant in the amnion. After contractions began, PR-A decreased significantly by 61.9% +/- 7.1 (P < 0.001).     

细胞外基质成分及其构筑对平滑肌细胞黏着斑激酶表达的影响

目的:探讨细胞外基质纤维连接蛋白(FN)、层黏连蛋白(LN)及三维基膜Matrigel胶对平滑肌细胞黏着斑激酶(FAK)表达的影响。方法:免疫荧光组织化学技术对生长在FN、LN和Matrigel胶里的平滑肌细胞FAK的表达进行检测。结果:生长存FN、LN和Matrigel胶里的平滑肌细胞FAK的表达依次为最高、次之和最低,各组之间有统计学意义;平滑肌细胞FAK磷酸化水平和FAK的表达呈正相关,生长在FN上的平滑肌细胞FAK(py397)的表达水平最高,生长在Matrigel胶里的平滑肌表达最低。结论:细胞外基质成分及其构筑对平滑肌细胞FAK的表达具有重要调节作用。     

Increased matrix metalloproteinase-9 activity in human ovarian cancer cells cultured with conditioned medium from human peritoneal tissue

Ovarian cancer cells disseminate by attachment to the peritoneal mesothelial cell surface of the abdominal cavity. We therefore investigated the influence of conditioned medium (CM) from human peritoneal tissues and mesothelial cells on the secretion of matrix metalloproteinases (MMPs) by ovarian cancer cells. The molecular weights of MMPs stimulating factors derived from human peritoneal tissues and mesothelial cells were estimated using microconcentrators with various cut-off membranes. Human peritoneal tissues were obtained from 12 surgical patients, and mesothelial cells were isolated from three peritoneal specimens. Exposure to CM from peritoneal tissue caused a concentration-dependent increase of the MMP-2 and MMP-9 bands in CM from NOM1 ovarian cancer cells, as shown by zymography. There was a significant difference in the increase of MMP-2 and MMP-9 (2.46-fold and 7.14-fold, respectively, at 0.4mg/ml protein; P < 0.005). CM from mesothelial cells also significantly increased the secretion of MMP-9 by NOM1 cells. The molecular size of possible MMP-9-stimulating factors secreted by peritoneal tissues and mesothelial cells was above M 100000. Further, CM of peritoneal tissues and mesothelial cells also induced the invasiveness of NOM1 cells. These findings suggest that mesothelial cells may secrete some factors which predominantly induce the MMP-9 production and increase invading cell numbers.     

雌激素诱导胎儿血管平滑肌细胞基质金属蛋白酶-2分泌和活化

目的: 探讨雌激素对胎儿血管平滑肌细胞(HVSMC)基质金属蛋白酶-2(MMP-2)分泌及其活化的影响。方法: 分别以不同浓度的雌二醇与体外培养的HVSMC作用24h,明胶酶谱法检测细胞培养上清液中明胶酶活性。结果: 随着雌二醇剂量增加,MMP-2的酶原形式(pro-MMP-2)和MMP-2活性形式逐渐增强。0.1 nmol/L、1 nmol/L、10 nmol/L、100 nmol/L雌二醇组pro-MMP-2分别比对照组高9.4%±8.2%(P>0.05)、73.0%±13.7%(P<0.05)、229.2%±42.2%(P<0.01)、428.4%±72.3%(P<0.01), MMP-2活性形式分别比对照组高39.8%±12.9%(P>0.05)、127.2%±20.4%(P<0.01)、428.4%±37.6%(P<0.01)、683.4%±50.9%(P<0.01)。结论: 雌二醇以剂量依赖性方式促进pro-MMP-2分泌和活化。