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Biomedical subjects

M Polette

Publications and source records attributed to M Polette.

At least 55 records · Page 3Linked to original sources
PubMed1994

Epithelial-to-mesenchymal transition in HPV-33-transfected cervical keratinocytes is associated with increased invasiveness and expression of gelatinase A.

The invasive potential of a set of HPV-33- and HPV-33 + ras-transfected cervical keratinocytes was investigated. These cell lines were previously separated into 2 groups according to their behavior on collagen rafts. Cell lines from the first group reconstituted CINIII-like lesions, whereas cell lines from the second group reconstituted epithelia comparable to micro-invasive carcinomas. They were thus postulated to represent distinct stages of cervical carcinogenesis. The present results have shown that lines from group I, which have conserved an epithelial morphology in monolayer, (i) could not invade matrigel when tested in a modified Boyden chamber assay, (ii) produced solely gelatinase B and (iii) were unable to activate exogenous gelatinase A. On the other hand, lines from group II associated epithelial-to-mesenchymal transition (acquisition of elongated morphology, vimentin positivity) with high in vitro invasive potential and with the ability both to produce and to activate gelatinase A. These results strongly support the hypothesis that the epithelial-to-mesenchymal transition and the associated events might be implicated in the progression to the metastatic phenotype.

Blotting, Northern↗
PubMed1994

Expression of gelatinase A, a mediator of extracellular matrix remodeling, by tracheal gland serous cells in culture and in vivo.

Tracheal gland morphogenesis and gland hypertrophy in disease involve the penetration of epithelial cells into the submucosa, a process that requires digestion of the basal lamina and the surrounding extracellular matrix. We observed that bovine tracheal gland cells invaded collagen substrates and were inhibited from doing so in the presence of a metalloproteinase inhibitor GM6001. The gland cells, but not bovine tracheal surface epithelial cells, secreted a 72-kDa metalloproteinase. The purified enzyme could be activated with 4-aminophenylmercuric acetate and converted to an active 65-kDa form that was far more effective in degrading denatured collagen (gelatin) than nondenatured type I and IV collagens and was ineffective in degrading intact interstitial collagen fibers. At 25 degrees C, the initial rate of degradation of acid-solubilized type I collagen was approximately 50 mg of type I collagen cleaved per min per mg of enzyme, whereas acid-solubilized type IV collagen was degraded at approximately 250 mg cleaved per min per mg of enzyme. In contrast, at the same temperature, heat-denatured type I collagen was degraded 1000-fold more rapidly, while heat-denatured type IV collagen was cleaved 50-fold more rapidly. The activity of the enzyme was maximal at pH 7-8 and was completely abolished by the metalloproteinase inhibitors EDTA and 1,10-phenanthroline. In immunoblots, the enzyme was recognized by an antibody directed against human gelatinase A, the 72-kDa gelatinase. The purified enzyme disrupted the distribution pattern of type IV collagen in the gland basal lamina, as well as of interstitial collagen in the underlying stromal tissue, as shown in tissue sections by immunocytochemistry. Using an antibody directed against the purified enzyme, we also showed by immunocytochemistry that the gelatinase was present in tracheal tissue and was specifically located at the periphery of some tracheal gland acini. Northern blots showed higher concentrations of gelatinase A mRNA in glands than in epithelium microdissected from adult cow tracheas. These data indicate that gelatinase A is a specialized product of the tracheal gland epithelial cell, a cell type normally invasive as part of its developmental program; the enzyme may play an important role in normal gland development and disease-associated hypertrophy.

Animals↗
PubMed1994

Coordinate enhancement of gelatinase A mRNA and activity levels in human fibroblasts in response to breast-adenocarcinoma cells.

Gelatinases/type-IV collagenases are metalloproteinases involved in some carcinoma invasion and metastatic processes. The exact cellular source of the 72-kDa gelatinase A is controversial. We have analyzed the expression of mRNA coding for gelatinase A in vivo by in situ hybridization on breast-cancer tissues. The mRNA for gelatinase A was present in fibroblasts. We have therefore evaluated the gelatinase-A activity in vitro, in co-cultures of different breast adenocarcinoma cell lines and human fibroblasts. In monoculture, none of the tumor cells tested produced detectable amounts of gelatinase A. The gelatinase-A activity was enhanced in cultures of fibroblasts maintained in the presence of MDA-MB 231 or SKBR3 cells, or their conditioned medium. This increased enzymatic activity was evidenced both in the culture medium and in the membrane fraction and was paralleled by enhancement of the steady-state levels of mRNA. These results are an in vitro demonstration of a regulation of fibroblasts gelatinase-A production by soluble factors secreted by breast-tumor cells.

Adenocarcinoma↗
PubMed1994

Gelatinase A expression and localization in human breast cancers. An in situ hybridization study and immunohistochemical detection using confocal microscopy.

The gelatinase A (72 kDa type IV collagenase) is a matrix metallo-proteinase which degrades basement membrane collagens. Various studies emphasize its role in stromal invasion of cancers, but there is some controversy about its origin. Gelatinase A was localized by immunohistochemistry using confocal microscopy in 15 human mammary carcinomas. In addition, the cells responsible for the synthesis of this enzyme were detected by in situ hybridization. Most invasive and non-invasive tumour cells were labelled by immunohistochemistry. Of particular interest was the pattern observed in some pre-invasive areas. Gelatinase A was found in fibroblasts in close contact with pre-invasive tumour clusters. Confocal observation allowed a more precise localization of gelatinase A to the periphery of tumour clusters along the basement membranes and in peritumour fibroblasts. The malignant epithelial cells were negative by immunohistochemistry in these areas. By in situ hybridization, mRNAs encoding gelatinase A were detected only in fibroblasts in close contact with pre-invasive and well differentiated tumour clusters. These findings support the hypothesis that peritumour fibroblasts produce gelatinase A and that breast cancer cells may bind this enzyme to their cell surface and/or internalize it.

Amino Acid Sequence↗
PubMed1994

Expression of stromelysin 3 and tissue inhibitors of matrix metallo-proteinases, TIMP-1 and TIMP-2, in rheumatoid arthritis.

Rheumatoid hyperplastic synovial lining cells and sublining fibroblasts are known to produce, in vivo and in vitro, matrix metallo-proteinases which degrade extracellular matrix components of joints. We have studied by immunohistochemistry and in situ hybridization the presence of matrix metallo-proteinase stromelysin 3 and its potential inhibitors TIMP-1 and TIMP-2 in 5 cases of normal synovia, 5 cases of chronic synovitis and 12 cases of rheumatoid arthritis. Few hyperplastic synoviocytes and some sparse fibroblasts have been found to produce stromelysin 3 in all rheumatoid arthritis and 2 chronic synovitis. Stromelysin 3 seems to have a limited role in the destructive process of extracellular matrix. TIMP-1 and TIMP-2 were largely expressed principally in hyperplastic synoviocytes and in endothelial cells of all rheumatoid synovitis and 2 chronic synovitis. These findings plead for a balance between matrix metallo-proteinases and their inhibitors in these inflammatory lesions.

Arthritis, Rheumatoid↗
PubMed1994

Role of matrix, fibroblasts and type IV collagenases in tumor progression and invasion.

We have studied the role of the extracellular matrix and host cells in tumor progression and tumor invasion. Our results emphasize the importance of tumoral cell-host cell interactions during this process. Addition of human fibroblasts and/or basement membrane components to human mammary adenocarcinoma cells, when injected into athymic nude mice, results in an increase of take and growth rate of the tumors. Peritumoral extracellular matrix is remodeled through multiple mechanisms: overproduction of matrix components by fibroblasts, enhanced fibroblasts proliferation, modulation of interstitial collagenase production by fibroblasts and retraction of the matrix by tumoral cells. The degradation of basement membranes during the metastatic process is often associated with the secretion of proteolytic enzymes. The 72 kDa type IV collagenase, a metalloproteinase, can be produced by some tumoral cells. However, it appears also to be secreted by peritumoral stromal fibroblasts under the influence of tumoral cells. We have demonstrated the existence of a binding site for this enzyme on the membrane of mammary tumoral cells. These results suggest a cooperation between tumor cells and fibroblasts during basement membrane destruction.

Animals↗
PubMed1994

Expression of gelatinases A and B and their tissue inhibitors by cells of early and term human placenta and gestational endometrium.

BACKGROUND: Human placentation is mediated by fetal trophoblastic cells that invade the maternal uterine endometrium. Trophoblast invasion requires a precisely regulated secretion of specific proteolytic enzymes able to degrade the endometrial basement membrane and extracellular matrix. EXPERIMENTAL DESIGN: Several studies have documented the key roles of matrix metalloproteinases and their tissue inhibitors in the invasion of various matrices by cultured trophoblasts. In vitro studies suggest that placentation could result from a balance between the secretion of these enzymes by trophoblast cells and their inhibition by the natural tissue inhibitors (TIMPs) produced by maternal decidual cells. The precise localization and levels of expression of these proteins that account for and control invasion during human placentation in vivo however, have not been described. We have evaluated, in vivo, by immunohistochemistry, Northern blot analysis and in situ hybridization, the expression of two metalloproteinases (gelatinases A and B) and their two tissue inhibitors (TIMPs 1 and 2) in placental villi and placental beds of first and third trimesters of normal pregnancy. RESULTS: Human first trimester intermediate trophoblast produced both gelatinases A and B; these two gelatinases were respectively less and no more detected at term in these cells. We found that both TIMP1 and 2 were also expressed in maternal decidual cells with a dramatic increase of TIMP1 at the term of pregnancy. In floating villi, gelatinase A and TIMP1 were localized in the stromal compartment, whereas gelatinase B and TIMP2 were codistributed in trophoblast cells. CONCLUSIONS: The gelatinases A and B and their tissue inhibitors are thus expressed by specific cells in early and late placental beds and villi. This pattern of expression varies during pregnancy. Therefore, our morphologic study supports biologic findings suggesting that these proteins may participate in placentation.

Blotting, Northern↗
PubMed1994

[Detection of anti-gonococcal antibodies using a dot immunobinding assay in a high risk female population].

The bacteriological diagnosis of gonorrhea in women has a low efficiency. A dot immunobinding assay, using whole piliated Neisseria gonorrhoeae cells, was used to detect antigonococcal antibodies in 300 prostitutes in whom a parallel Thayer-Martin culture of endocervical secretion was performed. Twenty sera from men with bacteriologically confirmed gonorrhoea were used as positive controls and 20 sera from children without history of gonococcal infections, as negative controls. Antibodies were detected in 49.7% of women and culture was positive in only one. All men with gonorrhea has positive titers and all sera from children were negative, except one that had a cross-reaction. Test sensitivity in the female population was 1 and specificity 0.5. There were no false negative reactions. Although the test detected antibodies in high risk women, there were cross-reactions. Therefore, the technique should be improved, using a more specific antigen.

Antibodies, Bacterial↗
PubMed1994

[Breast cancer invasion: the key role of normal cells of host tissues].

Tumor progression is influenced by extracellular matrices and by soluble factors or cytokines locally produced by host tissue cells (fibroblasts, immune cells ...). Such factors may also accumulate in close association with some extracellular matrix molecules in the tumor. They may also be unmasked during breaking down of extracellular matrices. The most insidious aspect of tumors is their propensity to locally invade normal tissues of the host and to form secondary foci in organs at distant sites from the primary tumor called metastases. During this process, invasive cells come into contact with host tissue cells such as fibroblasts, endothelial cells, macrophages, lymphocytes. These cells are not the passive witnesses of the metastatic cascade but actively participate to the malignant invasion. Through soluble messages (cytokines) and through insoluble molecules of the extracellular matrix, neoplastic and normal cells mutually modulate their activities. Cancer cells regulate the biosynthetic activities of fibroblasts and alter in this way the scaffold of the tumor. Reciprocally, host cells secrete extracellular matrix proteins and cytokines which influence the growth and activities of tumor cells. They also produce at the periphery of tumor cells proteolytic enzymes which promote host tissue destruction and cancerous cells migration. Among these enzymes, matrix metalloproteinases appear to play a key role during invasion and metastasis. Tumors represent thus a complex ecosystem. Tumor cells interact with several components of the extracellular matrix and with host cells (immune cells, fibroblasts, endothelial cells). Such multiple cell-cell and cell-matrix interactions condition angiogenesis, tumor growth, destruction of host tissues, local migration of cancer cells and their metastatic dissemination. It is probable that a precise knowledge of the genes which are selectively activated in tumors under the influence of the host cells or of the tumor cells will allow to define new therapeutic strategies. These treatments will aim not at destroying the metastatic neoplastic cells but at preventing their growth by interfering with their microenvironment.

Animals↗
PubMed1993

Localization by in situ hybridization of mRNAs encoding stromelysin 3 and tissue inhibitors of metallo-proteinases TIMP-1 and TIMP-2 in human head and neck carcinomas.

The presence and distribution of mRNAs encoding a matrix metalloproteinase (MMP) stromelysin 3 and two tissue inhibitors of MMP, TIMP1 and TIMP-2 have been studied by in situ hybridization of 18 human epidermoid head and neck carcinomas and four normal tissues. We found that in 16 tumors out of 18, stromelysin 3 mRNAs were only expressed by fibroblasts which were in close contact to invasive cancer cells. Tumor cells and normal tissues were not labeled. TIMP-1 mRNAs were detected in well differentiated cancer cells and in endothelial cells in all the cancers. In 13 out of the 18 carcinomas, TIMP-2 mRNAs were localized in only a few stromal cells near well differentiated invasive cancer cells and in endothelial cells. The significant expression of TIMP-1 and TIMP-2 mRNAS may lead to less aggressive MMPs, especially in the case of stromelysin 3, in the invasive process of the stroma.

Blotting, Northern↗
PubMed1993

[Cell interactions in the extracellular matrix in tumoral invasion in mammary pathology].

Tumoral invasion in breast lesions is associated with a progressive loss of the basement membrane (BM) components. In situ hybridization reveals a loss of synthetic ability of type IV collagen by invasive cells in a murine model. The loss of BM components does not seem related to the expression of the plasmin system by cancer cells. By contrast, there is a cooperation between fibroblasts and cancer cells for the expression of matrix metallo-proteinases involved in BM degradation in breast carcinomas in vivo.

Animals↗
PubMed1993

Non radioactive single-strand conformation polymorphism (SSCP) analysis of exon 11 of the CFTR gene using the Pharmacia PhastSystem.

The authors report here the use of the PhastSystem (Pharmacia) to perform the single strand conformation polymorphism analysis of polymerase chain reaction products of the exon 11 of the CFTR gene. It provides a rapid (2 hours) safe and reliable technique for the development of carrier testing for individuals or couples with a family history of cystic fibrosis.

Cystic Fibrosis↗
PubMed1993

Detection and localization of mRNAs encoding matrix metalloproteinases and their tissue inhibitor in human breast pathology.

Matrix metalloproteinases (MMPs) are a group of enzymes thought to be responsible for both normal connective-tissue-matrix remodelling and the accelerated breakdown associated with tumor development. These MMPs and tissue inhibitor of MMPs (TIMP1) could be expressed by either the cancer or the stromal cells. Expression of mRNAs encoding interstitial collagenase (MMP1), 72-kD type IV collagenase (MMP2) and stromelysin (MMP3), which are probably involved in tumor invasion and metastasis, and of TIMP1 were studied in human mammary pathology by in situ hybridization and Northern blot analysis. Out of 6 benign lesions, 2 expressed MMP2 mRNAs. mRNAs encoding MMP1 and MMP3 were detectable in occasional stromal and tumor cells in 2 out of 17 carcinomas. Thirteen out of 17 cancers expressed MMP2 mRNA throughout the tumor in stromal cells close to noninvasive tumor clusters and well-differentiated invasive cancer cells. TIMP1 mRNA expression was detected in noninvasive and well-differentiated invasive tumor cells. These data suggest that there is a cooperation between tumor and stromal cells, in particular for the production of 72-kD type IV collagenase, involved in the disruption of basement membranes. A lack of TIMP1 expression from invasive cancer cells would also contribute to matrix destruction.

Blotting, Northern↗
PubMed1992

[Modulation of the organization of the extracellular matrix and the production of collagen by interferon gamma in three-dimensional cultures of normal and sclerodermic fibroblasts].

Recent studies have shown inhibition of collagen synthesis by interferon gamma (IFN-gamma) in monolayer human fibroblast cultures on a plastic solid phase. However, the existence of this effect in vivo has not yet been demonstrated. Three-dimensional fibroblast cultures in collagen matrices (collagen lattices or dermal equivalents) provide a more physiological model than conventional cultures and fairly closely simulate in vivo conditions. This model was used for studying effects of IFN-gamma on normal and scleroderma fibroblasts. IFN-gamma induced a dose-dependent inhibition of fibroblast-mediated retraction of collagen lattices. IFN-gamma also inhibited total protein and collagen synthesis. Scleroderma fibroblasts were especially susceptible to the inhibitory effects of IFN-gamma. Since fibrotic scleroderma lesions are associated with tissue retraction and increased production of extracellular matrix macromolecules, these data confirm the potential value of IFN-gamma for the treatment of scleroderma and other fibrotic diseases.

Adult↗
PubMed1992

DNA content measurement and in situ hybridization in condylomatous cervical lesions.

The present study reports results obtained with DNA ploidy measurement by Feulgen cytophotometry and in situ hybridization (ISH) on the same sections of 36 cervical lesions containing human papillomavirus (HPV) DNA. Fifteen of 16 low grade lesions [11 flat condylomas and five condylomatous with intraepithelial neoplasias (CIN)-1, 1 with condylomatous features] were diploid. "Oncogenic" HPV (types 16 and 33) were detected in three of these cases, while the aneuploid case expressed HPV type 11. By contrast, in the 12 CIN-2 and eight CIN-3 with condylomas, there was a good correlation between presence of an aneuploid DNA pattern and detection of "oncogenic" HPVs. Demonstration of an aneuploid profile and "oncogenic" HPV as found in all our CIN-3, may indicative of a poor prognosis in low grade lesions. Hence, the combination of the two techniques (ISH and Feulgen cytophotometry) may provide significant prognostic information for condylomas and CINs.

Aneuploidy↗
PubMed1992

A new nonisotopic detection of human papillomavirus DNA using polymerase chain reaction.

A novel technique using a two-step polymerase chain reaction (PCR) with specific primers detecting human papillomavirus (HPV) DNA of types 6/11, 16, and 18 and a final nonisotopic colorimetric detection has been developed. Sixty formalin-fixed and paraffin-embedded sections were treated with this methodology and the results compared with those obtained with in situ hybridization (ISH). Twenty cases displaying HPV DNA with ISH were positive with PCR. Seven (35%) of 20 cases negative for ISH but evocative of HPV infection with classic histology displayed HPV DNA with the two-step PCR. Only one case (5%) of 20 normal tissues and/or inflammatory lesions not evocative of HPV infection and negative upon ISH showed HPV DNA. This original technique allows rapid, highly sensitive, and specific detection of HPV DNA and is suitable for most laboratories.

Base Sequence↗
PubMed1992

Immunolocalization of matrix metallo-proteinases and their tissue inhibitor in human mammary pathology.

Matrix metallo-proteinases (MMPs) are a group of enzymes thought to be responsible for both normal connective tissue matrix remodelling and accelerated breakdown associated with tumor development. The distribution of 3 major matrix metallo-proteinases was studied in human mammary pathology: collagenase (MMP1) which degrades fibrillar interstitial collagens, a 72-kDa gelatinase (MMP2) which mainly degrades type IV collagen and denatured collagens, and stromelysin (MMP3) which has a wider range of action, degrading several matrix components including the core proteins of proteoglycans, laminin and non-helical regions of collagens. These MMPs and the MMP tissual inhibitor (TIMP1) were detected by immunohistochemistry in 30 benign and 79 malignant lesions of the breast. MMPs were detected in 1 fibroadenoma (collagenase) and 22 breast carcinomas: collagenase (9 cases), stromelysin (12 cases) and gelatinase (16 cases) with a limited distribution. Tumor cells were preferentially labelled and the localization of gelatinase and stromelysin at the periphery of some non-invasive and well-differentiated clusters supports the role of these enzymes in the breakdown of basement membranes. Only a few stromal cells (fibroblasts) were found to be immunopositive. In contrast, TIMP1 was more frequently detected, and was found in 7 benign lesions and 55 carcinomas out of 79. It was mainly localized at the periphery of the endothelial cells but was occasionally detected in cancer cells and fibroblasts.

Adult↗