RESUMEN

Matrix metalloproteinases, in particular the gelatinases MMP-2 and MMP-9, have received great attention in recent years as putative tumour markers for clinical applications. The main reason for the observed interest is their easy detection in body fluids. Moreover, recent evidence has shown multiple functions of MMPs, rather than simply degrading ECM, which include the mobilisation of growth factors and processing of surface molecules. Several authors have reported increased levels of MMPs in a number of cancers, but clinical correlations in breast cancer are still fragmentary. Thus, the aim of the present research was to investigate the activity levels of circulating gelatinases in the sera of breast cancer patients by means of zymographic analysis, and correlate data with clinicopathological parameters. In all, 80 patients and 22 healthy volunteers were involved in this study. Sera were obtained prior to surgery. The clinical variables were: grading of tumours, tumour size, lymph node involvement, tumour staging, oestrogen and progesterone receptor levels (76 out of 80 cases), and c-erbB-2 levels (46 cases). The densitometric measures of MMP-2 and MMP-9 activity levels indicated that the average values of both gelatinase activities were significantly higher in breast cancers than in control sera (P<0.0001). In addition, our analysis showed for the first time that elevated activity levels of both gelatinases correlated only with c-erbB-2 overexpression (P=0.0273 for MMP-2 and P=0.0075 for MMP-9). An inverse correlation was observed with regard to oestrogen receptor expression (P=0.0075 for MMP-2 and P=0.0273 for MMP-9). Moreover, a borderline inverse correlation was observed between the activity levels of both enzymes and nuclear grade (P=0.0511 for MMP-2 and P=0.0794 for MMP-9). In conclusion, the present data suggest that serum measures of MMP's activity may have diagnostic value for discriminating subgroups of breast cancer patients and support the hypothesis that ERBB2 amplification and/or overexpression enhance signalling pathways that may lead to increased production of gelatinases in c-erbB-2 positive breast cancers with higher metastatic potentialities.

Asunto(s)

Neoplasias de la Mama/enzimología , Metaloproteinasa 2 de la Matriz/sangre , Metaloproteinasa 9 de la Matriz/sangre , Biomarcadores de Tumor , Neoplasias de la Mama/patología , Femenino , Humanos , Metástasis Linfática , Metaloproteinasa 2 de la Matriz/metabolismo , Metaloproteinasa 9 de la Matriz/metabolismo , Pronóstico , Receptor ErbB-2/metabolismo , Receptores de Estrógenos/metabolismo , Receptores de Progesterona/metabolismo

RESUMEN

Gelatinase A (MMP-2) and gelatinase B (MMP-9) play a key role in the proteolytic cascade leading to ECM degradation during invasion and metastasis. The enzyme activity is regulated both at the intra- and extra-cellular level. Extracellular regulation is achieved mainly through the balance between proenzyme activation and inhibition, which appears to be altered in cancer patients. One of the mechanisms of MMP inhibition is the binding of the enzymes to appropriate tissue inhibitors (TIMP). In the recent literature, it has been suggested that MMP-2 and/or MMP-9 are indeed over-produced in many carcinomas, while the identity of the various enzymatic forms (latent, activated and enzyme/inhibitor complexes) remains to be elucidated. In this study we have analyzed the circulating forms of MMP-9 and MMP-2 in serum samples of patients with colon carcinoma, as well as the enzymatic activities present in tissue extracts from surgical fragments (primary tumor and its paired healthy tissue). Proteins were separated by means of mono-dimensional or bidimensional electrophoresis, and the enzymes detected by gelatin zymography and immunological assays. The results of densitometric analyses demonstrate that proMMP-9, but not proMMP-2, is significantly higher in the oncologic sera vs. the normal sera. In addition, several oligomeric circulating and tissue forms of MMP-9 are preferentially found in the oncologic samples, both in mono- and second-dimension zymograms. The activated forms of MMP-2 and MMP-9 are uniquely present in the primary tumor extracts, thus confirming the involvement of the tissue microenvironment in gelatinase activation and function.

Asunto(s)

Neoplasias del Colon/enzimología , Metaloproteinasa 2 de la Matriz/metabolismo , Metaloproteinasa 9 de la Matriz/metabolismo , Adulto , Neoplasias del Colon/sangre , Neoplasias del Colon/patología , Densitometría , Electroforesis en Gel Bidimensional/métodos , Electroforesis en Gel de Poliacrilamida/métodos , Precursores Enzimáticos/sangre , Precursores Enzimáticos/aislamiento & purificación , Precursores Enzimáticos/metabolismo , Humanos , Metaloproteinasa 2 de la Matriz/sangre , Metaloproteinasa 2 de la Matriz/aislamiento & purificación , Metaloproteinasa 9 de la Matriz/sangre , Metaloproteinasa 9 de la Matriz/aislamiento & purificación

RESUMEN

The extracellular matrix (ECM) is known to play an active role in numerous biological processes such as differentiation, apoptosis and cancer. Extensive alterations of epithelial basement membranes and of interstitial ECM are known to occur during the progression of most invasive carcinomas. Collagen, which represents the major component of the interstitial ECM, is primarily involved in the stromal changes at the site of tumor cell invasion. We have previously described the occurrence in breast and colon cancer ECM of an oncofetal form of collagen, characterized by an acidic chain distinct from those of type I and III collagen. In the present paper, we bring evidence that alpha2(I) collagen chains in colon cancer tissues expressing the acidic chains, are either overmodified or absent, both as protein and as regular mRNA transcripts. The results obtained strongly suggest that: i) the disorganisation of the collagen architecture and the phenomenon of fibril dispersion, which accompanies the lysis of basement membrane, is not only due to the enzymatic degradation of the collagen fibres, but presumably also to changes of the collagen molecules deposited in the stroma; ii) the neosynthesis of collagen occurring at tumor-host interface is deeply deregulated, and therefore to be considered the result of altered collagen gene expression correlated with the tumor progression, rather than as a mere defensive reaction of the host cells.

Asunto(s)

Colágeno/metabolismo , Neoplasias Colorrectales/metabolismo , Secuencia de Aminoácidos , Biopsia , Neoplasias Colorrectales/patología , Electroforesis en Gel Bidimensional , Humanos , Microscopía Electrónica , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Homología de Secuencia de Aminoácido

RESUMEN

We previously produced evidence that the human mammary-carcinoma cell line 8701-BC expresses several metalloproteinases (MMP-1, -2, -9, and -10) and their tissue inhibitors). In order to obtain a better understanding of the environmental control over gelatinolytic activities, we have tested the enzyme production of 8701-BC cells, at time intervals after plating on different collagen substrates, i.e., types I, III, IV, V and OF/LB, used as films in culture dishes. Proteinase activities, released in the conditioned culture media, were tested by zymography on SDS-PAGE, and by quantificative analyses, using 14C carboxymethylated transferrin as substrate in a liquid incubation medium. Enzymatic activities varied with time and were inversely related to cell densities, with minimum values at cell confluence. The enzymatic activity was positively supported by collagen substrates, with a maximal increase in activity when OF/LB collagen was used. In addition to the known MMPs, we found a proteinase with an M(r) of about 20 kDa, which displayed higher activity at 48 hr after cell plating and gradually decreased with cell increment. In contrast to the other MMPs, this proteinase is inhibited by soybean trypsin inhibitor, but it does not display a complete identity with trypsin, since it does not digest casein and is not inhibited by other serine proteinase inhibitors.

Asunto(s)

Neoplasias de la Mama/enzimología , Neoplasias de la Mama/patología , Comunicación Celular/fisiología , Colágeno/farmacología , Gelatinasas/biosíntesis , División Celular/fisiología , Medios de Cultivo , Endopeptidasas/aislamiento & purificación , Endopeptidasas/metabolismo , Humanos , Peso Molecular , Células Tumorales Cultivadas

RESUMEN

We have recently identified an oncofetal-laminin binding collagen (OF/LB) composed of three alpha chains, with the apparent molecular mass of about 100 kDa each, but bearing different pI. One of the chains appears markedly acidic in a bidimensional electrophoretic system, where the NEPHGE is used as first dimension separating gel, while the two more basic chains have similar migration as alpha 1(III) and alpha 1(I) collagen chains, respectively. Sequence analyses have been performed on CNBr-peptides, derived from pepsinized triple helical molecules and on tryptic fragments obtained after in gel digestion of the acidic band. The research of sequence homology with computerized databases indicated that the acidic chain represents a gene product distinct from either type I, type III and other known collagen chains, while the identity of the other two chains remains to be fully determined.

Asunto(s)

Colágeno/química , Neoplasias del Colon/química , Secuencia de Aminoácidos , Biopsia , Cromatografía Líquida de Alta Presión , Colágeno/aislamiento & purificación , Neoplasias del Colon/patología , Neoplasias del Colon/cirugía , Bromuro de Cianógeno , Electroforesis en Gel Bidimensional , Humanos , Sustancias Macromoleculares , Datos de Secuencia Molecular , Fragmentos de Péptidos/química , Fragmentos de Péptidos/aislamiento & purificación , Homología de Secuencia de Aminoácido , Tripsina

RESUMEN

OF/LB collagen is a recently described once-fetal form of collagen, with laminin-binding properties, composed of three alpha(1)(I)-sized chains, one of which displaying an unusually acidic pI. This collagen appears able to direct the migration of breast cancer cells through Matrigel, and of injury-activated epithelial cells into the underlying granulation stromal tissue. The effect exerted by OF/LB collagen in vitro appears preferentially linked to its acidic chain. The data reported strongly support the hypothesis that the presence and accumulation of OF/LB collagen in cancer may play a fundamental role in the invasive growth.

RESUMEN

8701-BC is a recently characterized cell line isolated from a primary ductal infiltrating carcinoma of the breast (d.i.c.), showing some pleomorphism in cell microanatomy at an ultrastructural level. We have obtained different sublines of 8701-BC cells by cloning in soft agar at different concentrations (0.3% and 0.6%), and we have characterized the cloned lines by some morphological and growth parameters. 8701-BC cells and clones have been submitted to analysis by reverse transcriptase-linked polymerase chain reaction to detect mRNAs of various cytokines (transforming growth factor-beta s, tumour necrosis factors, interleukin 1s, interleukin 6, parathyroid hormone-related peptide, gamma interferon) and of urokinase, which are bioactive molecules commonly involved in cell-cell and cell-stroma interactions at primary and/or secondary sites of invasion. The aims of the present investigation were to determine: (a) if the corresponding genes are active in 8701-BC cell line and (b) if the sublines tested exhibit transcriptional heterogeneity. The results obtained show that 8701-BC cells express transcripts of transforming growth factor-beta s, urokinase and parathyroid hormone-related peptide (PTHrP), the latter product being responsible for the cancer-associated humoral hypercalcemic syndrome. Moreover, while the first two mRNAs are detectable in all the sublines tested, PTHrP is expressed almost uniquely by the clones isolated in 0.6% agar which exhibit a peculiar morphological appearance, a higher growth rate and a more active invasive behaviour in vitro.(ABSTRACT TRUNCATED AT 250 WORDS)

Asunto(s)

Neoplasias de la Mama/química , Neoplasias de la Mama/patología , Carcinoma Ductal de Mama/química , Carcinoma Ductal de Mama/patología , Proteínas/análisis , Factor de Crecimiento Transformador beta/análisis , Activador de Plasminógeno de Tipo Uroquinasa/análisis , Secuencia de Bases , Biomarcadores de Tumor/análisis , Neoplasias de la Mama/genética , Carcinoma Ductal de Mama/genética , Células Clonales , Humanos , Interleucina-1/análisis , Interleucina-1/genética , Interleucina-6/análisis , Interleucina-6/genética , Datos de Secuencia Molecular , Proteína Relacionada con la Hormona Paratiroidea , Fenotipo , Reacción en Cadena de la Polimerasa , Proteínas/genética , ARN Mensajero/análisis , ARN Mensajero/genética , Factor de Crecimiento Transformador beta/genética , Células Tumorales Cultivadas , Factor de Necrosis Tumoral alfa/análisis , Factor de Necrosis Tumoral alfa/genética , Activador de Plasminógeno de Tipo Uroquinasa/genética

RESUMEN

It is widely accepted that collagenolytic enzymes are required to facilitate the invasion and spread of tumour cells into host tissues. Immunohistochemical, zymographic and PCR analyses have produced evidence that the recently established human mammary carcinoma cell line, 8701-BC, expresses several metalloproteinases (MMP-1, -2, -9 and -10) and their tissue inhibitors (TIMP-1 and -2). Application of these different techniques has led to several observations, both complementary and dissimilar. Whereas PCR analysis showed that mRNA was detected for each of the proteins, the immunolocalization study demonstrated that MMP-1, MMP-2, MMP-9 and TIMP-1 production was restricted to only a proportion of the tumour cells, with no evidence of MMP-3 or TIMP-2 synthesis. Such observations suggested phenotypic heterogeneity within the cell line, which was further examined by use of the tumour cell clones BC-3A and BC-61 derived from the parental 8701-BC line. Comparative studies using zymography and PCR analysis demonstrated differences in MMP-2 and MMP-10 expression between the 3 cultures. The data indicate that the 8701-BC cell line retains an inherent capacity for metalloproteinase and TIMP expression, with the production of both interstitial collagenase (MMP-1) and the 2 basement-membrane-degrading enzymes (MMP-2 and MMP-9) representing an aggressive collagenolytic phenotype. The concomitant production of TIMP-1 by these cell cultures, and the apparent phenotypic heterogeneity displayed by these lines, suggest that metalloproteinase dysregulation may represent an important feature of clonal heterogeneity. Although the 8701-BC and BC-61 cells were much more invasive than those of the BC-3A clone, as judged by the penetration of "Matrigel", it has not yet been possible to relate this invasive potential to the metalloproteinase and TIMP profiles reported here for each cell line.

Asunto(s)

Neoplasias de la Mama/metabolismo , Carcinoma/metabolismo , Glicoproteínas/biosíntesis , Inhibidores de la Metaloproteinasa de la Matriz , Metaloendopeptidasas/biosíntesis , Neoplasias de la Mama/patología , Carcinoma/patología , División Celular , Quimiotaxis , Electroforesis en Gel de Poliacrilamida , Técnica del Anticuerpo Fluorescente , Humanos , Reacción en Cadena de la Polimerasa , Inhibidores Tisulares de Metaloproteinasas , Células Tumorales Cultivadas

RESUMEN

Human breast and colon carcinoma tissues contain a form of collagen, not described before, composed of alpha 1 chains of similar size (approximately 100 kDa) but different charge. The three constitutive chains, separated by two-dimensional electrophoresis, are a unique acidic component, undetectable in other collagen types, with an apparent isoelectric point of 4-5, and two more basic components displaying the same electrophoretic behavior as alpha 1(III) and alpha 1(I), respectively. The acidic chain is structurally distinct from alpha 1(I) and displays a cyanogen bromide-derived fragment of similar size to CB5(III). This collagen in its native state is resistant to trypsin and metalloproteinase 3, while it is fully degraded by metalloproteinases 1 and 9. Moreover, this collagen appears able to bind to laminin, as tested by affinity chromatography. The biological significance of our data is related to the finding of this collagen form not only in the tumor tissue tested but also in embryonic-fetal tissues (bovine skin and intestine and human umbilical cord). For its peculiar laminin-binding ability and occurrence in tumoral and embryonic-fetal tissues, we propose to temporarily term this new collagen form OF/LB collagen (onco-fetal, laminin-binding collagen). The presence of OF/LB collagen during development and cancer, and its absence in normal adult tissues, make this protein a potential stromal marker of malignancy.

Asunto(s)

Neoplasias de la Mama/química , Colágeno/metabolismo , Neoplasias del Colon/química , Feto/metabolismo , Laminina/metabolismo , Animales , Bovinos , Colágeno/química , Colágeno/ultraestructura , Bromuro de Cianógeno , Electroquímica , Electroforesis en Gel de Poliacrilamida , Humanos , Intestinos/química , Intestinos/embriología , Punto Isoeléctrico , Metaloendopeptidasas/metabolismo , Microscopía Electrónica , Fragmentos de Péptidos/química , Fragmentos de Péptidos/metabolismo , Piel/química , Piel/embriología , Tripsina/metabolismo , Cordón Umbilical/química

RESUMEN

Ductal infiltration carcinomas (d.i.c.) of the breast are potentially highly metastatic tumours, associated with drastic alterations of the architecture and molecular composition of the extracellular matrix at the tumour-host interface. 8701-BC, a recently characterized cell line, isolated from primary d.i.c., was used to study different aspects of tumor cell-substratum interactions. Since type V collagen deposition is augmented in d.i.c. we have examined the ability of 8701-BC cells to interact with this collagen species. We have found that cell binding to type V collagen was mediated by protein homologous to the 67 kDa laminin receptor (67-R). This conclusion is substantiated by the following observations: (a) a major band having an apparent molecular mass of 67 kDa and immunoreactive to the anti-67 R antibody was detectable by SDS-PAGE of the membrane proteins; (b) the antibody inhibited cellular adhesion to type V collagen in a dose-dependent way; (c) membrane proteins purified by affinity chromatography on type V collagen were immunoreactive to anti-67 R antibody, but not to anti-VLA1, VLA2 and VLA3 integrin antibodies. This receptor appears to have prominent carbohydrate-binding properties, since lactose competes with cell adhesion to type V collagen.

Asunto(s)

Adhesión Celular , Colágeno/metabolismo , Neoplasias Mamarias Experimentales/metabolismo , Receptores de Superficie Celular/biosíntesis , Animales , Cromatografía de Afinidad , Lactosa/fisiología , Receptores de Colágeno , Células Tumorales Cultivadas

RESUMEN

Ductal infiltrating carcinoma (d.i.c.) of human breast is a highly invasive neoplasm characterized by enhanced deposition of collagen. Paradoxically, enhanced collagen deposition is not correlated with inhibition of the migration of tumour cells into the host tissue. d.i.c. is characterized by the reappearance of 'embryonic' type I-trimer collagen and an increase in type V collagen content in the matrix. The effects of these two collagen types were compared with type I collagen as culture substrata on the spreading pattern, cytoskeletal organization and motile behaviour of 8701-BC breast carcinoma cells using rhodamine-phalloidin staining, a DNAase I-competition assay, scanning electron microscopy and time-lapse video-microscopy. Cells grown on type I collagen were stationary, showing a well-spread morphology and an extensive stress fibre pattern. Cells grown on type V collagen were also stationary, but displayed a poorly spread and elongated morphology. In contrast, cells grown on trimer collagen were motile and displayed a compact morphology and a reduced content of stress fibres. Both single-cell and group motility were detectable on trimer collagen substratum. These data are consistent with the existence of two opposite local signals, type I-trimer and type V collagens, which may confer a more or a less metastatic phenotype on breast carcinoma cells. Moreover, the synthesis of trimer collagen in d.i.c. is conceivably instrumental in providing new stromal pathways permitting tumour cells to infiltrate the host tissue.

Asunto(s)

Neoplasias de la Mama/patología , Carcinoma Intraductal no Infiltrante/patología , Colágeno/fisiología , Metástasis de la Neoplasia , Actinas/metabolismo , Movimiento Celular , Medios de Cultivo , Matriz Extracelular/fisiología , Humanos , Fotomicrografía , Células Tumorales Cultivadas , Grabación de Cinta de Video

RESUMEN

Type V collagen is one of the minor components of the extracellular matrix (ECM) whose content is increased in cases of ductal infiltrating carcinomas of the breast. In order to clarify its biological role, we have investigated the effect of this molecule, both as substrate and as soluble factor, on the behaviour of a breast carcinoma cell line (8701-BC) grown in vitro. Cell-collagen adhesion was monitored for 24 h from plating in the absence or presence of serum. The influence of type V collagen on cell growth was followed during 9 days of culture, and the actin-vinculin arrangement was studied by simultaneous fluorescent immuno-staining. The results indicate that type V collagen is not a permissive substrate for neoplastic cell proliferation and dissemination in vitro.

Asunto(s)

Neoplasias de la Mama/patología , Carcinoma Intraductal no Infiltrante/patología , Colágeno/fisiología , Proteínas del Citoesqueleto/ultraestructura , Neoplasias de la Mama/ultraestructura , Carcinoma Intraductal no Infiltrante/ultraestructura , Adhesión Celular , División Celular , Humanos , Células Tumorales Cultivadas/patología , Células Tumorales Cultivadas/ultraestructura

RESUMEN

A continuous cell line of neoplastic cells derived from ductal infiltrating carcinoma of the human breast (8701-BC), was assayed for its ability to adhere to collagen substrates. The collagens used were regular type I and type I homotrimer isolated from primary breast carcinomas. Comparative studies were performed using an embryonic epithelial cell line derived from human intestine (Int. 407). The neoplastic cells adhere equally well to both collagens, while the embryonic epithelial cells recognized only the homotrimer. Some receptor diversity was recognized in the adhesion of the two cell lines to homotrimer collagen. The data demonstrate a functional difference between type I and homotrimer collagen with regard to cellular recognition and attachment. In addition, the data suggest that oncogenic transformation of breast epithelial cells promotes their adhesive properties to interstitial collagens and that this may be relevant to their increased potential to invade host tissue.

Asunto(s)

Neoplasias de la Mama/metabolismo , Carcinoma Intraductal no Infiltrante/metabolismo , Adhesión Celular , Colágeno/metabolismo , Neoplasias de la Mama/patología , Carcinoma Intraductal no Infiltrante/patología , Línea Celular , Transformación Celular Neoplásica/metabolismo , Colágeno/inmunología , Femenino , Fibronectinas/fisiología , Humanos , Inmunoglobulina G , Células Tumorales Cultivadas

RESUMEN

A cell line, designated 8701-BC, was established in culture from tissue fragments of primary ductal infiltrating carcinoma of human breast. The cell cultures after the sixth passage were devoid of contaminating fibroblasts as judged by the positive staining of all cells with the specific epithelial cell markers carcinoembryonic antigen (CEA), tissue polypeptide antigen (TPA) and cytokeratin 8. The epithelial nature of these cells was confirmed by ultrastructural analyses which demonstrated the retention of specific structural properties characteristic of the original tumour. The cells possessed an abnormal karyotype with 55-60 chromosomes per cell with numerous rearrangements. They do not express HLA antigens and the c-myc gene was not amplified. The 8701-BC cells have a doubling time of approx. 29h and have been maintained in culture for more than 100 passages. These properties suggest the establishment of a human neoplastic cell line which, with its ability to produce homotrimer collagen in vitro, will provide a useful model system for the study of tumour cell:stromal matrix interactions.

Asunto(s)

Neoplasias de la Mama/patología , Carcinoma Intraductal no Infiltrante/patología , Células Tumorales Cultivadas/patología , Neoplasias de la Mama/ultraestructura , Carcinoma Intraductal no Infiltrante/ultraestructura , Línea Celular , Humanos , Cariotipificación , Células Tumorales Cultivadas/ultraestructura

RESUMEN

The present ultrastructural study has been undertaken in order to contribute to the problem of morphological heterogeneity in the ductal infiltrating carcinoma of the human breast. In spite of the well known topographical variability of scirrhous breast cancers, the comparative analysis of 12 primary tumours has brought to light some basic phenotypical expressions of the neoplastic cell population. The major observation is the occurrence of two main categories of cells, which are interpretable as the transformed counterparts of the dark and light lumenal cells of the normal mammary epithelium. The phenotypical identity of the two categories has been assessed by in vitro cultivation (parallel paper). Many aberrant morphological variants, attributable to the two cell types, were observed at the tumour-stroma interface. We have therefore suggested that the high level of morphological heterogeneity may, at least in part, be the result of stromal influences on the gene expression of the transformed cells.

Asunto(s)

Neoplasias de la Mama/ultraestructura , Mama/patología , Carcinoma Intraductal no Infiltrante/ultraestructura , Anciano , Biopsia , Transformación Celular Neoplásica , Femenino , Humanos , Microscopía Electrónica , Persona de Mediana Edad

RESUMEN

The ultrastructural characterization of the present continuous cell line, derived from a primary ductal infiltrating carcinoma (d.i.c.) of the human breast, has brought to light a remarkable morphological similarity with the original neoplastic cell population. A major parallelism is the permanent presence in culture of two categories of cells, exhibiting a strong isomorphism with the in vivo counterparts. The presence of well defined ultrastructural features, as duct-like structures, microvillous projections, junctional complexes and intracytoplasmic crypts, is a further confirmation of the breast cancer origin of this line. The significance and perspective of these findings are discussed.

Asunto(s)

Neoplasias de la Mama/ultraestructura , Carcinoma Intraductal no Infiltrante/ultraestructura , Femenino , Humanos , Microscopía Electrónica , Fenotipo , Células Tumorales Cultivadas/ultraestructura

RESUMEN

Type I-trimer collagen, isolated from biopsy fragments of ductal infiltrating carcinomas, was used as a substrate for human breast carcinoma cells in long-term culture to monitor growth rate, morphological appearance and actin organization in comparison with normal type I collagen and plain plastic. After 11 days of culture, type I-trimer collagen exerts a more pronounced effect on cell proliferation, leading to a final increment of cell population of 35% versus regular type I substrate. Furthermore, type I-trimer collagen induces cell motility, as testified by morphological appearance and actin immunofluorescence test. On the basis of the in vitro results, it is postulated that in vivo the stromal areas containing trimer collagen, rather than repressing invasive growth, may provide a more suitable environment for tumor proliferation and spreading-out with respect to regular type I.

Asunto(s)

Actinas/fisiología , Neoplasias de la Mama/patología , Carcinoma Intraductal no Infiltrante/patología , Colágeno/farmacología , Actinas/análisis , Neoplasias de la Mama/metabolismo , Carcinoma Intraductal no Infiltrante/metabolismo , Transformación Celular Neoplásica/efectos de los fármacos , Colágeno/metabolismo , Citoesqueleto/análisis , Citoesqueleto/efectos de los fármacos , Femenino , Humanos , Células Tumorales Cultivadas/metabolismo , Células Tumorales Cultivadas/patología

RESUMEN

Collagen biosynthesis was assayed in tissue fragments and in cultured neoplastic cells derived from primary ductal infiltrating carcinoma of the human breast. Neoplastic cells "in vitro" produce 3-4% of collagen with respect to the high molecular weight protein fraction. The neosynthesized collagen is mainly composed of alpha 1 (I) chains, which may be assembled as homotrimer molecules, as indicated by their resistance to pepsin digestion. In tissue fragments, (where neoplastic and host stromal cells coexist), the collagen percentage increases up to 15-20% and more than one polypeptide chain is produced. Present data suggest that neoplastic cells "in vivo" contribute to the deposition of collagen components, actively synthesizing a certain amount of the type I-trimer, which is a significant component of the "scirrhous" stroma (Minafra et al.1984; Pucci Minafra et al, 1985). This phenomenon is interpreted as one of the numerous interrelationships occurring at the cell-matrix interface during the malignant growth.

Asunto(s)

Neoplasias de la Mama/metabolismo , Colágeno/biosíntesis , Células Tumorales Cultivadas/metabolismo , Biopsia , Línea Celular , Colágeno/análisis , Femenino , Humanos

RESUMEN

The tumour stroma in cases of ductal infiltrating carcinoma of the mammary gland contains substantial amounts of collagen type I trimer, besides the regular collagen types. Reconstituted collagen trimer consists of fibrils that are significantly thinner than reconstituted fibrils of type I collagen. The axial periodicity is somewhat longer due to widening of the c-d and d-e regions. Transmission EM of the tumours shows characteristic phenomena at the stromal-tumour cell junctions: frequent absence of a basal lamina and thin disordered collagen fibrils that show frequent direct contacts with tumour cells. On the basis of literature data concerning the interaction between stroma and epithelia under physiological and pathological conditions it is hypothesized that the interaction of collagen type I trimer fibrils with tumour cells is instrumental in augmenting tumour cell progression and that the trimer may provide contact guidance for invasive growth.

Asunto(s)

Adenocarcinoma Escirroso/ultraestructura , Neoplasias de la Mama/ultraestructura , Carcinoma Intraductal no Infiltrante/ultraestructura , Colágeno/análisis , Femenino , Humanos , Sustancias Macromoleculares , Microscopía Electrónica

RESUMEN

Further analyses about collagen present in ductal infiltrating carcinoma of human mammary gland indicate that a large amount of it is represented by type I omotrimer that has been separated from the two other present species, type I eterotrimer and type III, by means of fractionated saline precipitation. Quantitative determinations of the three types, extracted by mild pepsin digestion, are also reported.

Asunto(s)

Neoplasias de la Mama/análisis , Colágeno/análisis , Humanos , Masculino