Biomedical subjects

C Gilles

Publications and source records attributed to C Gilles.

At least 37 records · Page 2Linked to original sources

PubMed1997

Induction of membrane-type matrix metalloproteinase 1 (MT1-MMP) expression in human fibroblasts by breast adenocarcinoma cells.

Membrane-type matrix metalloproteinase 1 (MT1-MMP) has been recently described as an activator of proMMP-2 (MMP-2) which is involved in tumor invasion. We have shown by in situ hybridization that MT1-MMP is produced by stromal cells in close contact to preinvasive and invasive tumor cells of breast carcinomas. Of particular interest was the observation that some fibroblasts express this enzyme in focal areas in preinvasive lesions, suggesting that particular tumor cells may stimulate fibroblasts to produce MT1-MMP. We have therefore compared the ability of two different breast cancer cell lines, one non-invasive (MCF7) and one invasive (MDA-MB-231) to stimulate MT1-MMP production in human fibroblasts with consequent proMMP-2-activation. The MDA-MB-231 conditioned medium induced MT1-MMP mRNAs in human fibroblasts and a parallel activation of proMMP-2 whereas MCF7 conditioned medium did not have any effect. These results suggest the existence of soluble factor(s) secreted by invasive or some preinvasive breast tumor cells which stimulate fibroblasts to produce and activate MMPs, and emphasize the cooperation between cancer and stromal cells in tumor invasion.

Blotting, Northern↗

PubMed1997

Expression of c-ets-1 mRNA is associated with an invasive, EMT-derived phenotype in breast carcinoma cell lines.

We have previously observed in vitro that some stromal proteinases (MMP-2, MT1-MMP) were expressed or activated by invasive carcinoma cell lines exhibiting mesenchymal features, presumably acquired through an epithelial to mesenchymal transition (EMT). To examine the potential contribution of c-ets-1 to this phenotype, we have compared here the expression of c-ets-1 with invasiveness in vitro and expression of vimentin, E-cadherin, uPA, MMP-1 and MMP-3 in a panel of human breast cancer cell lines. Our results clearly demonstrate an association between c-ets-1 expression and the invasive, EMT-derived phenotype, which is typified by the expression of vimentin and the lack of E-cadherin. While absent from the two non-invasive, vimentin-negative cell lines, c-ets-1 was abundantly expressed in all the four vimentin-positive lines. However, we could not find a clear quantitative or qualitative relationship between the expression of c-ets-1 and the three proteinases known to be regulated by c-ets-1, except that when they were expressed, it was only in the invasive c-ets-1-positive lines. UPA mRNAs were found in three of the four vimentin-positive lines, MMP-1 in two of the four, and MMP-3 could not be detected in any of the cell lines. Intriguingly, MDA-MB-435 cells, which exhibit the highest metastatic potential of these cell lines in nude mice, expressed vimentin and c-ets-1, but lacked expression of these three proteinases, at least under the culture conditions employed. Taken together, our results show that c-ets-1 expression is associated with an invasive, EMT-derived phenotype in breast cancer cells, although it is apparently not sufficient to ensure the expression of uPA, MMP-1 or MMP-3, in the vimentin-positive cells. Such proteases regulation is undoubtedly qualified by the cellular context. This study therefore advances our understanding of the molecular regulation of invasiveness in EMT-associated carcinoma progression, and suggests that c-ets-1 may contribute to the invasive phenotype in carcinoma cells.

Breast Neoplasms↗

PubMed1997

Tumor collagenase stimulatory factor (TCSF) expression and localization in human lung and breast cancers.

Tumor cell-derived collagenase stimulatory factor (TCSF) stimulates in vitro the biosynthesis of various matrix metalloproteinases involved in tumor invasion, such as interstitial collagenase, gelatinase A, and stromelysin 1. The expression of TCSF mRNAs was studied in vivo, using in situ hybridization and Northern blotting analysis, in seven normal tissues and in 22 squamous cell carcinomas of the lung, and in seven benign proliferations and in 22 ductal carcinomas of the mammary gland. By in situ hybridization, TCSF mRNAs were detected in 40 of 44 carcinomas, in pre-invasive and invasive cancer cells of both lung and breast cancers. TCSF mRNAs and gelatinase A mRNAs were both visualized in the same areas in serial sections in breast cancers, and were expressed by different cells, tumor cells, and fibroblasts. The histological results were confirmed by Northern blot analysis, which showed a higher expression of TCSF mRNAs in cancers than in benign and normal tissues. These observations support the hypothesis that TCSF is an important factor in lung and breast tumor progression.

Antigens, CD↗

PubMed1997

Implication of collagen type I-induced membrane-type 1-matrix metalloproteinase expression and matrix metalloproteinase-2 activation in the metastatic progression of breast carcinoma.

We have previously demonstrated that fibroblasts and invasive human breast carcinoma (HBC) cells specifically activate matrix metalloproteinase-2 (MMP-2) when cultured on 3-dimensional gels of type I collagen but not a range of other substrates. We show here the constitutive expression of membrane-type 1 (MT1)-MMP in both fibroblasts, and invasive HBC cell lines, that have fibroblastic attributes presumably acquired through an epithelial-to-mesenchymal transition (EMT). Treatment with collagen type I increased the steady-state MT1-MMP mRNA levels in these cells but did not induce either MT1-MMP expression or MMP-2 activation in noninvasive breast carcinoma cell lines, which retain epithelial features. Basal MT3-MMP mRNA expression had a pattern similar to that of MT1-MMP but was not up-regulated by collagen. MT4-MMP mRNA was seen in both invasive and noninvasive HBC cell lines and was also not collagen-regulated, and MT2-MMP mRNA was not detected in any of the HBC cell lines tested. These data support a role for MT1-MMP in the collagen-induced MMP-2-activation seen in these cells. In situ hybridization analysis of archival breast cancer specimens revealed a close parallel in expression of both collagen type I and MT1-MMP mRNA in peritumoral fibroblasts, which was correlated with aggressiveness of the lesion. Relatively high levels of expression of both mRNA species were seen in fibroblasts close to invasive tumor nests and, although only focally, in certain areas close to preinvasive tumors. These foci may represent hot spots for local degradation and invasive progression. Collectively, these results implicate MT1-MMP in collagen-stimulated MMP-2 activation and suggest that this mechanism may be employed in vivo by both tumor-associated fibroblasts and EMT-derived carcinoma cells to facilitate increased invasion and/or metastasis.

Breast Neoplasms↗

PubMed1997

Emerging roles for proteinases in cancer.

Metalloproteinases and serine proteinases have been associated with tumor invasion and formation of metastasis which represent the major obstacles to cancer cure. The contribution of proteinases in these processes was initially thought to be the destruction of extracellular matrices. However, recent evidence suggests that they mainly affect tumor growth rather than invasion. Proteinases can indeed generate active matrix protein fragments, influence the release, the activation and the bioavailability of growth factors, and consequently modulate tumor cell growth, apoptosis and angiogenesis. Additionally, proteinases, their receptors and/or inhibitors can be directly involved in cell migration and in the processing or shedding of cell surface proteins. Further elucidation of the functions of proteinases is essential for the development of novel anticancer strategies.

Animals↗

PubMed1996

Crystal structure of pig pancreatic alpha-amylase isoenzyme II, in complex with the carbohydrate inhibitor acarbose.

Two different crystal forms of pig pancreatic alpha-amylase isoenzyme II (PPAII), free and complexed to a carbohydrate inhibitor (acarbose), have been compared together and to previously reported structures of PPAI. A crystal form obtained at 4 degrees C, containing nearly 72% solvent, made it possible to obtain a new complex with acarbose, different from a previous one obtained at 20 degrees C [Qian, M., Buisson, G., Duée, E., Haser, H. & Payan, F. (1994) Biochemistry 33, 6284-6294]. In the present form, six contiguous subsites of the enzyme active site are occupied by the carbohydrate ligand; the structural data indicate that the binding site is capable of holding more than the five glucose units of the scheme proposed through kinetic studies. A monosaccharide ring bridging two protein molecules related by the crystal packing is located on the surface, at a distance of 2.0 nm from the reducing end of the inhibitor ligand; the symmetry-related glucose ring in the crystal lattice is found 1.5 nm away from the non-reducing end of the inhibitor ligand.

Acarbose↗

PubMed1996

Crystallization and preliminary X-ray analysis of pig pancreatic alpha-amylase in complex with a bean lectin-like inhibitor.

Pig pancreatic alpha-amylase (PPA, E.C. 3.2. 1. 17, 496 amino-acid residues) has been crystallized as a complex with a lectin-like inhibitor from bean Phaseolus vulgaris (224 amino-acid residues for the inhibitor monomer). The hanging-drop vapour-diffusion method was used to grow crystals from solutions containing 2-methyl-2,4-pentanediol as precipitant. The crystals belong to monoclinic space group C2 with a = 152.5, b = 80.3, c = 68.8 A, beta = 91.4 and diffract to 2.9 A resolution. A molecular-replacement solution of the structure has been obtained using the refined PPA and LoLl (Lathyrus ochrus isolectin I) atomic coordinates as starting models. Low-resolution refinement of the model is underway. The analysis reveals that the functional inhibitor molecule is dimeric and interacts with two molecules of enzyme.

Journal Article↗

PubMed1996

High level of MT-MMP expression is associated with invasiveness of cervical cancer cells.

MMP-2 (gelatinase A) has been associated with the invasive potential of many cancer cells both in vitro and in vivo. It is now becoming clear that the activation of this enzyme might be a key step in tumor invasion. This activation process has been shown to be a membrane-associated pathway inducible by various agents such as collagen type I, concanavalin A or TGF-beta, but its physiological regulation is still largely unresolved. MT-MMP was recently discovered and described as a potential gelatinase-A activator. In the present study, we investigated the expression of MT-MMP (membrane-type metalloproteinase) in cervical cancer cells both in vitro and in vivo. Comparing several in vitro-transformed cervical cell lines, previously shown to display different invasive potentials, our results showed that the ability of cells to overexpress MT-MMP mRNA following ConA induction correlated with their ability to activate gelatinase A and with a highly invasive behavior. Moreover, using immunohistochemistry and in situ hybridization, we found a higher level of MT-MMP expression in invasive cervical carcinoma and lymph node metastases compared to its expression in non-invasive CIN III lesions. Our in vivo observations also clearly demonstrated a cooperation between stromal and tumor cells for the production of MT-MMP. Taken together, our results clearly correlated high level MT-MMP expression with invasiveness, and thus suggested that MT-MMP might play a crucial role in cervical tumor invasion.

Base Sequence↗

PubMed1996

Vimentin expression in cervical carcinomas: association with invasive and migratory potential.

Vimentin is an intermediate filament protein normally expressed in mesenchymal cells, but evidence is accumulating in the literature which suggests that the aberrant expression of vimentin in epithelial cancer cells might be related to local invasiveness and metastatic potential. Vimentin expression has previously been associated with invasive properties in an in vitro model consisting of a set of HPV-33-transformed cervical keratinocyte cell lines. In the present study, in order to emphasize those in vitro findings, the expression of vimentin has been investigated in cervical neoplasms of different grades, using immunohistochemistry. A clear association is reported between vimentin expression and metastatic progression, since vimentin was detected in all invasive carcinomas and lymph node metastases, but not in CIN III lesions. These in vivo results are compared with present and previous data obtained in vitro on cervical keratinocyte cell lines, where vimentin expression also correlated with in vitro invasiveness.

Animals↗

PubMed1996

MT-MMP expression and localisation in human lung and breast cancers.

Thirteen primary pulmonary squamous cell carcinomas, 4 specimens of normal lung from around tumours, 4 benign proliferations of the mammary gland and 16 breast carcinomas were analysed by in situ hybridisation. Northern blot and immunohistochemistry for the expression of a recently described metalloproteinase (MMP), the MT-MMP (membrane-type matrix metalloproteinase). This MT-MMP can activate gelatinase A, involved in the degradation of basement membranes. In situ hybridisation revealed MT-MMP transcripts distributed in both tumour and stromal cells in squamous cell lung cancers, whereas these mRNAs were principally detected in stromal cells in close contact to tumour clusters in breast carcinomas and in lung adenocarcinomas. Northern blot analysis showed a parallel expression of MT-MMP and gelatinase A transcripts in both lung and breast cancers. Immunohistochemistry displayed a more extensive distribution of MT-MMP in pulmonary and mammary carcinomas with numerous labelled preinvasive and infiltrating cancer cells and stromal cells near the tumour cells. The large degree of expression of MT-MMP in these cancers indicates a potential role of this enzyme in tumour progression. The finding of MT-MMP transcripts in stromal cells in the vicinity of lung and breast tumour cells emphasises the cooperation between these cells and cancer cells for the expression of MT-MMP and in tumour invasion in vivo.

Breast Neoplasms↗

PubMed1996

Viral integration sites in human papilloma virus-33-immortalized cervical keratinocyte cell lines.

The viral organization of HPV-33 was determined by Southern blotting in 2 HPV-33-immortalized cervical cell lines (CK11 and CK12) and compared to our previous results obtained on 10 other already characterized HPV-33-immortalized cell lines (CK1 to CK10). As observed in CK1 to CK10, the viral DNA was found integrated in the cellular genome of CK11 and CK12. However, in CK11 and CK12, the integrated viral genome was deleted and mostly limited to the URR and the E6-E7 ORFs, stressing the importance of those sequences in the immortalization process. Furthermore, CK11 and CK12 showed a unique and identical integration site, as observed in CK1 to CK10, which also harbored HPV-33 integrated at a unique and identical site (which was however different from the one evidenced in CK11 and CK12). Indeed, in situ hybridizations on chromosomes allowed the precise localization of the viral DNA on chromosome 13q33-34 in CK1 to CK10 whereas it was mapped to chromosome 9p13 in CK11 and CK12. We discuss the possibility that integration of HPV-33 at those two particular sites has conferred some growth advantages to the cells and could have thus played a crucial role in the immortalization.

Cell Line, Transformed↗

PubMed1996

Wound repair-induced expression of a stromelysins is associated with the acquisition of a mesenchymal phenotype in human respiratory epithelial cells.

Wound repair involves many processes including cell migration, provisional matrix deposition, and remodeling. All of these processes are likely to be affected by matrix-modifying enzymes. Members of the matrix metalloproteinases family are physiologic mediators of the extracellular matrix degradation. Within this matrix metalloproteinases family, stromelysins can degrade many components of the extracellular matrix. We therefore tested the hypothesis that stromelysins could be produced by human surface respiratory epithelial (HSRE) cells repairing a wound. Experimental wounds were created in vitro in HSRE cell cultures and in situ in human bronchial mucosa maintained in organ culture. Stromelysin production was measured by casein-gel zymography in cellular protein extracts derived from repairing migratory and nonrepairing stationary cells of wounded HSRE cell cultures. Stromelysin-producing cells present in cell and tissue cultures were localized and characterized using immunofluorescence techniques. Zymographic and immunofluorescence techniques showed that stromelysins were produced exclusively by the migratory HSRE cells. Zymogram analysis showed that stromelysins were overexpressed and overactivated during the wound repair process, with the maximal production observed at wound closure. Using an anti-cytokeratin 14 antibody, we identified stromelysin-3-producing cells as basal epithelial cells. Moreover, most stromelysin-3-producing cells expressed the mesenchymal marker vimentin. Similar to stromelysins localization, vimentin-positive HSRE cells were exclusively located in the wounded area, and they were also positive to cytokeratin 14. In conclusion, stromelysins are suggested to be involved in HSRE cell migration and extracellular matrix remodeling during wound repair. Furthermore, stromelysin production by repairing HSRE cells is linked to the acquisition of a mesenchymal phenotype. HSRE cell migration may then be associated with the shift from an epithelial to a mesenchymal phenotype.

Bronchi↗

PubMed1996

[Poly-medication and drug interaction in geriatrics].

Firstly the authors remind the reader of the geriatric paradoxe consisting in an overprescription of drugs despite the fact that the side effects are more frequent and also the common inadequacy of the polymedications resulting from time to time in hospitalisations which otherwise would not have occurred. Secondly, the authors focus on 9 families of drugs potentially dangerous to the health and 5 interactions with deadly consequences. Finally, they underline the importance of some preliminary evaluation before any prescription and of various recommendations.

Aged↗

PubMed1996

[Antigenic retrieval using a pressure cooker].

This technical note describes antigen retrieval procedures using a pressure cooker. These procedures are perfectly adapted to routine immunohistochemistry and improve the overall quality of immunostaining.

Antigens↗

PubMed1995

Neurofibrillary degeneration in amyotrophic lateral sclerosis/parkinsonism-dementia complex of Guam. Immunochemical characterization of tau proteins.

Neurofibrillary tangles are observed in several neurodegenerative disorders including Alzheimer's disease, progressive supranuclear palsy, and amyotrophic lateral sclerosis/parkinsonism-dementia complex of Guam. The major components of neurofibrillary tangles are hyperphosphorylated tau proteins that can be directly detected in brain homogenates, using immunoblotting with specific immunological probes. To investigate whether tau proteins differ biochemically among various neurodegenerative disorders, we analyzed a series of brain samples from Guamanian patients in comparison with Alzheimer's disease, progressive supranuclear palsy, and normal aging. In Alzheimer's disease, these hyperphosphorylated tau proteins are composed of a triplet referred to as tau 55, 64, and 69, whereas in progressive supranuclear palsy, neurofibrillary degeneration is characterized by a tau doublet (tau 64 and 69). In the present study, characterization of tau proteins was performed by immunoblotting, on different cortical and subcortical regions of postmortem brain specimens from Guamanian natives. In all of the cases, biochemical data were always consistent with neuropathological findings. In contrast to Alzheimer's disease patients where the tau triplet is found mostly in cortical regions, a similar triplet was strongly detected in both cortical and subcortical areas in Guamanian patients. The tau profile differed quantitatively from case to case demonstrating that the Alzheimer's disease-related tau triplet had a heterogeneous regional distribution. These data suggest that the tau triplet found in amyotrophic lateral sclerosis/parkinsonism-dementia complex of Guam is similar to that observed in Alzheimer's disease, and the regional distribution of tau proteins differs in these disorders.

Adult↗

PubMed1995

Inhibition of growth of normal and human papillomavirus-transformed keratinocytes in monolayer and organotypic cultures by interferon-gamma and tumor necrosis factor-alpha.

The growth response of normal and human papillomavirus (HPV)-transformed cervical keratinocytes to interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha was investigated in monolayer and organotypic raft cultures. The proliferation rates of monolayer cultures were assessed by [3H]TdR incorporation and fluorimetric DNA titration. The growth of keratinocytes in organotypic cultures was estimated by their ability to stratify on collagen rafts and by immunohistochemistry for Ki67 antigen expression. IFN-gamma reduced the DNA synthesis of normal and HPV-transformed keratinocytes in monolayer cultures and exerted a marked growth inhibitory effect in organotypic raft cultures. In control raft cultures, normal keratinocytes produced an epithelial sheet of approximately 10 cells in thickness that closely resembled normal cervical epithelium and was characterized by sparse Ki67 antigen-positive cells whereas HPV-transformed keratinocytes produced up to 15 poorly differentiated epithelial layers that were reminiscent of high grade cervical lesions seen in vivo and exhibited a full thickness Ki67 antigen expression. When normal and HPV-transformed keratinocytes were maintained in the presence of IFN-gamma, the epithelial sheet was reduced to a few cells in thickness and the density of Ki67 antigen-positive cells was decreased. A more pronounced growth inhibitory effect in monolayer and organotypic cultures was observed when IFN-gamma was associated with tumor necrosis factor-alpha Tumor necrosis factor-alpha alone reduced the DNA synthesis of normal keratinocytes but was significantly less effective than IFN-gamma to inhibit the growth of HPV-transformed keratinocytes. These results suggest that similar responses in vivo to regulatory molecules may play a role in the development of HPV-related lesions.

Cell Division↗

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

Differentiation ability and oncogenic potential of HPV-33- and HPV-33 + ras-transfected keratinocytes.

Five HPV-33-immortalized and 5 HPV-33 + ras-transfected cell lines were characterized in terms of growth in soft agar, tumorigenic potential in nude mice, p21 expression, morphology and expression of differentiation markers in organotypic cultures. No striking differences were observed between the HPV-33-immortalized cell lines and their corresponding ras-transfected counterparts as regards their tumorigenicity in nude mice (only one cell line was able to develop tumors in nude mice) or their behavior on lifted collagen gels. However, all the ras-transfected cell lines gave rise to colonies in soft agar while only 2 HPV-33-transfected lines (CK1 and CK4) displayed this property. The 10 cell lines could be divided into 2 groups with respect to their phenotype in monolayer and in organotypic cultures. Lines from group I (CK2, 3, 5 and their ras-transfected homologous lines) shared a typical epithelial phenotype in monolayer and the ability (a) to form an epithelium similar to a CIN-III lesion and (b) to strongly express keratins K1-K10 and involucrin in organotypic cultures. On the other hand, for the lines from group II (CK1, CK4, CK1EJ7 and CK4EJ5), there was a correlation between an elongated phenotype in monolayer and the property (a) to form a structure similar to a microinvasive carcinoma and (b) to express vimentin and keratins K8-K18. These cell lines, exhibiting various transformation-associated alterations, can be considered as an in vitro model representing various stages of HPV-33-associated cervical carcinogenesis.

Animals↗