Biomedical subjects

F Garrido

Publications and source records attributed to F Garrido.

At least 19 recordsLinked to original sources

PubMed1992

Effect of MHC class-I transfection on local tumor growth and metastasis in an H-2-negative clone derived from a chemically induced fibrosarcoma.

GR9 is a chemically induced fibrosarcoma composed of clones with different H-2 class-I expression. These clones differ with respect to local growth and spontaneous metastasis. The B9 clone (H-2 negative) is highly tumorigenic (local growth) but of low metastatic potential (spontaneous metastasis assay). We have analyzed the effect that transfection of H-2Dd and H-2Kd genes on this clone have upon local growth, lung colonization after i.v. injection and ability to form spontaneous metastases. The results showed that the effect on local growth of transfection of the Kd-gene was stronger than that of the Dd gene. In addition, B9 co-transfected with H-2Kd and Dd genes showed the highest immunogenic properties in syngeneic BALB/c mice. Interestingly, the pSV2-neo transfected clone gave almost the same result as that obtained with Dd transfection. Lung colonization after i.v. injection of the different clones (experimental metastasis), paralleled the results obtained for local growth: the number of lung nodules followed the cadence KdDd less than Kd less than Dd less than pSV2. Survival of mice was always inversely correlated with local growth, e.g., all mice injected with 5 x 10(5) B9 H-2KdDd transfected cells survived. In contrast, no mice injected with the B9 control did. These differences were abrogated in irradiated and nude BALB/c mice. Finally, all transfected clones remained non-metastatic in a spontaneous metastasis assay, behaving as the control, non-transfected B9 cells.

Animals↗

PubMed1992

Expression of alpha-tropomyosin during cardiac development in the chick embryo.

A new monoclonal antibody (mAb) that recognizes alpha-tropomyosin in cardiac muscle cells was used in a qualitative (polyacrylamide gel electrophoresis and indirect immunofluorescence) and quantitative (fluorescence-activated cell sorting) study of the expression of this protein during heart development. alpha-Tropomyosin expression was weak in early stages of chick embryo development (Hamburger and Hamilton stage 18), and increased steadily until Hamburger Hamilton stage 40. In early stages, the protein was found mainly in cytoplasm, whereas by the final stages, it was more abundant in the cytoskeletal compartment. The mAb cross-reacted with alpha-tropomyosin in smooth and striated muscle cells from chickens, mice, and humans, but did not cross-react with nonmuscle tropomyosin.

Animals↗

PubMed1992

HLA molecules in basal cell carcinoma of the skin.

Fifty basal cell carcinomas (BCC) and 8 samples of healthy skin were studied for HLA class I and class II antigen expression and for the presence of mutations in codon 12 of the K-ras and H-ras genes. All samples of healthy skin and of epithelium near the tumor showed high levels of class I molecules, whereas 38% of the tumors showed complete absence. Sixty-two percent of the tumors presented positive class I expression with heterogeneous staining. These losses were due to the simultaneous lack of heavy chain and beta 2-microglobulin. Selective losses of HLA-A or HLA-B antigens were not detected. Class II antigens were absent in most of the tumors, only two tumors showing a few weakly positive cells with anti-HLA-DR mAb. The loss of class I expression correlated significantly with the degree of histological differentiation and aggressiveness. We were unable to correlate class I expression with clinical size, depth of invasion or the extent of leukocytic infiltrate surrounding the tumor. Analysis by PCR amplification of codon 12 of the K-ras and H-ras oncogenes detected H-ras mutations in 1 out of 50 cases, and no K-ras mutations in any of the tumors studied. Thus, a positive relationship between K-ras and H-ras mutations and BCC tumorigenesis or MHC alterations seems unlikely in this neoplasia.

Antigens, Neoplasm↗

PubMed1992

Tracking genetically engineered bacteria: monoclonal antibodies against surface determinants of the soil bacterium Pseudomonas putida 2440.

Assessment of potential risks involved in the release of genetically engineered microorganisms is facilitated by the availability of monoclonal antibodies (MAbs), a tool potentially able to monitor specific organisms. We raised a bank of MAbs against the soil bacterium Pseudomonas putida 2440, which is a host for modified TOL plasmids and other recombinant plasmids. Three MAbs, 7.3B, 7.4D, and 7.5D, were highly specific and recognized only P. putida bacteria. Furthermore, we developed a semiquantitative dot blot assay that allowed us to detect as few as 100 cells per spot. A 40-kDa cell surface protein was the target for MAbs 7.4D and 7.5D. Detection of the cell antigen depended on the bacterial growth phase and culture medium. The O antigen of lipopolysaccharide seems to be the target for MAb 7.3B, and its in vivo detection was independent of the bacterial growth phase and culture medium. MAb 7.3B was used successfully to track P. putida (pWW0) released in unsterile lake mesocosms.

Antibodies, Monoclonal↗

PubMed1992

[Diurnal variations in circulating leukocytes and subsets: relation to plasma cortisol and ACTH].

Surface cellular antigens of leukocytes, lymphocytes and corresponding subpopulations have been analysed by using monoclonal antibodies marked with fluorescein (PITC), parallel to those marked with phycoerythrin (PE). Cortisol and ACTH plasmatics have also been determined through RIA, on two samples at 8am. and 8pm. During this twelve hour evolution, a highly significant dependence of the leukocytes, T, T4 and T8 lymphocytes on the circulating ACTH has also been found. In general during the experiment time leukocytes, lymphocytes and subpopulations, have experimented an increase which is significantly related to the pituitary hormone secretion. The existence of this significant correlation establishes the presence of a possible mechanism that connects the cellular immunity to determined hypothalamus hormones.

Adrenocorticotropic Hormone↗

PubMed1991

Differential mRNA levels of c-myc, c-fos and MHC class I in several clones of a murine fibrosarcoma.

We have evaluated the relationship between MHC class-I, c-myc and c-fos proto-oncogene expression in several clones of a methylcholanthrene-induced fibrosarcoma (GR9) which originated in a BALB/c mouse. These clones represent a heterogeneous population and differ markedly with regard to H-2 class-I cell-surface expression, local tumor growth, NK sensitivity and metastatic potential. In the present study we show that cell-surface expression of MHC class-I antigens correlates inversely with levels of c-myc mRNA transcripts. On the other hand, mRNA levels of c-fos are correlated directly with the amount of mRNA of MHC class I. Treatment of the B9 clone with gamma interferon increased mRNA transcription and surface expression of H-2 class-I antigens, while c-myc transcription was simultaneously down-regulated. In contrast, c-fos mRNA levels remained unaltered.

Animals↗

PubMed1991

Expression of HLA class I and II antigens in bronchogenic carcinomas: its relationship to cellular DNA content and clinical-pathological parameters.

We studied the presence of HLA class I antigens in 115 samples of bronchogenic carcinomas (66 frozen and 49 formalin-fixed and paraffin-embedded specimens) by the immunophosphatase alkaline and immunoperoxidase methods with antibodies against major histocompatibility complex antigens. We also studied HLA class II antigens on the 66 frozen tumor samples. Nonneoplastic lung tissue was also analyzed for purposes of comparison. Pneumocytes and epithelial respiratory cells expressed HLA class I and II antigens. The expression of class I antigens was totally lost in 29 tumors (25%). The defect in HLA gene expression affected both heavy chain and beta 2-microglobulin, as demonstrated by the null reactivity with specific antibodies. In 2 cases of 66 studied in cryostatic section, the selective loss of A locus was observed, and in three cases selective loss of B locus was detected. The expression of class I antigens was compared with clinical-pathological parameters such as histological type, degree of differentiation, and tumor stage, as well as tumoral ploidy. The absence of expression of HLA class I molecules was significantly associated with poorly differentiated and undifferentiated tumors (P less than 0.0001) and with aneuploid tumors (P less than 0.001), suggesting that some lung tumors may escape immune surveillance and become biologically more aggressive. Class II antigens were expressed in 13 cases of 66 studied (18%) in frozen specimens, and a clear relationship was observed with well-differentiated tumors (P less than 0.05).

Adenocarcinoma↗

PubMed1991

Altered HLA class I expression in non-small cell lung cancer is independent of c-myc activation.

We studied the expression of major histocompatibility complex class I antigens in 59 bronchogenic carcinomas, as well as in pneumocytes and epithelial respiratory cells distant from the tumor. We observed in all cases that normal lung tissue expressed major histocompatibility complex class I antigens, while this expression was completely lost in 16 tumors (27%). The defect in HLA gene expression affected both heavy chain and beta 2-microglobulin, as demonstrated by the null reactivity with the monoclonal antibodies GRH1, W6/32, and HC10. Selective underexpression was detected in 1 tumor for HLA-A locus antigens and in 3 tumors for HLA-B locus antigens. Southern blot analyses of major histocompatibility complex class I genes were performed in 20 tumor tissue specimens and 6 cell lines. No class I gene rearrangements were detected using HLA coding and locus specific noncoding probes. We also used the Southern blot method to investigate the possible relationship between c-myc amplification and HLA class I antigens in non-small cell lung cancers and detected no apparent amplification in 20 tumor tissue specimens (5 negative for HLA class I antigens) and 6 cell lines (3 with decreased expression). Northern blot analysis revealed no relationship between c-myc mRNA levels and specific mRNA for HLA-A and HLA-B antigens in cell lines with imbalanced HLA-A or HLA-B expression.

Carcinoma, Bronchogenic↗

PubMed1991

Heterogeneity of MHC-class-I antigens in clones of methylcholanthrene-induced tumors. Implications for local growth and metastasis.

We analyzed the H-2 class-I expression of different clones obtained from 4 different methylcholanthrene-induced tumors in BALB/c (H-2d) mice. The results clearly indicated high intra-tumor heterogeneity in all 4 fibrosarcomas with regard to H-2 K, D and L expression. Clones were found to be H-2-negative, H-2-positive, or to present intermediate expression. Southern blot analysis of class-I genes showed RFLPs equal to those obtained from normal BALB/c DNA, ruling out rearrangements or gross deletions in the class-I genes of different tumors. However, Northern blot studies showed a straightforward relationship between class-I mRNA levels and H-2 expression. In vivo experiments demonstrated an inverse relationship between local growth and spontaneous metastasis, e.g., H-2-positive class-I clones produced high numbers of lung colonies but very poor local growth, and vice-versa. These results paralleled the NK sensitivity or resistance of the different clones. Cross-protection experiments showed that only clones coming from the same tumor were able to protect against challenge with clones of the same neoplasia but not with clones from different chemically induced fibrosarcomas, indicating that a clone of a given tumor probably contained the same TATA. Finally, we compared the H-2, K, D and L expression and class-I mRNA levels of various metastatic colonies. Interestingly, another degree of heterogeneity was found: an H-2-negative clone (GR9.B9) gave rise to H-2-negative (B9MP6) and H-2-positive (B9MP2) metastatic colonies.

Animals↗

PubMed1991

Molecular analysis of MHC-class-I alterations in human tumor cell lines.

Molecular characterization of HLA-class-I expression was investigated in human tumor cell lines at the protein and mRNA levels using locus-specific monoclonal antibodies (MAbs) and probes. Some cell lines exhibited a differential expression of HLA-A and HLA-B products and also showed differences in the inducibility of HLA-class-I genes by gamma-IFN. Thus, gamma-IFN stimulation induced predominantly HLA-B mRNA in the HeP-2 cell line, which showed imbalances in basal levels of HLA-A and HLA-B expression. This unequal inducibility of HLA genes may imply that locus-specific regulatory mechanisms are involved in the expression of individual HLA products. The specific mechanism controlling the differential expression of HLA subsets appears to be independent of c-myc activity. Northern blot analysis found no relationship between c-myc mRNA levels and specific mRNA for HLA-A and HLA-B antigens.

Antibodies, Monoclonal↗

PubMed1991

Can the HLA phenotype be used as a prognostic factor in breast carcinomas?

There is evidence indicating that change in the expression of HLA-ABC antigens can modulate the biological behavior and metastatic potential of certain neoplasms. We studied 15 samples of normal breast epithelium, 94 breast carcinomas and 24 benign and pre-malignant lesions of the mammary gland. All cases of normal breast epithelium and non-malignant lesions presented high levels of expression of class-I antigens. In contrast, 22 out of 94 carcinomas showed a reduction in the level of expression and a heterogeneous pattern. In addition, 31 tumors were considered negative for HLA-class-I expression, and 6 cases showed selective loss of HLA-ABC: 2 tumors for HLA locus A, and 4 for HLA locus B. We found a direct relationship between patient survival and HLA-negative phenotype (p less than 0.001), as well as between histological grade of malignancy and the level of expression of class-I antigens (p less than 0.0005). Moreover, the presence of class-I molecules was significantly related to tumor ploidy (p less than 0.005). Our results lead us to conclude that HLA-ABC-negative tumors have a higher metastatic potential and greater clinical aggressiveness: patients with carcinomas exhibiting low HLA expression have more lymph-node metastases (p less than 0.02) and achieve shorter survival times (p less than 0.001).

Adult↗

PubMed1991

MHC antigens on human tumors.

MHC class I antigens on tumor cells are expected to play an important role because they regulate the sensitivity to antitumoral immunological mechanisms. Overall or selective qualitative or quantitative changes in MHC molecules may modify the recognition of tumor cells by components of the immune system. It seems clear that MHC antigen expression on tumor cells is important in triggering the immune response by autologous lymphocytes. A deficiency in or lack of MHC class I antigens may have profound effects on T and NK cell activity. In experimental models, variation in the expression of MHC class I antigens has been shown to exert a decisive influence on local tumor growth and metastasis. However, there is little information about the influence of selective loss of individual locus products on the behavior of human tumor cells. Total and selective HLA losses have been found in a large variety of tumors, and different mechanisms have been shown to be responsible for these changes. In some examples, HLA losses are associated with a poor degree of tissue differentiation and poor prognosis. In other tumors, however, no such association has been found. We do not know whether HLA class II expression in neoplastic cells plays an immunological role, although, with the exception of melanoma, HLA class II expression is more frequently observed in tumors with a more favorable prognosis. Finally, there is no doubt that we need to learn more about how to manipulate the expression of MHC class I and II antigens in human tumors, in order to stimulate immune response.(ABSTRACT TRUNCATED AT 250 WORDS)

Antigens, Neoplasm↗

PubMed1991

Characterization of CD44 antigen during lymphoid ontogeny.

We studied immunohistological and biochemical aspects of the CD44 molecule with a mAb produced in our lab: GRHL-1. The characteristic expression of this antigen in cells of B lineage was analyzed. This mAb showed identical immunohistological patterns of reactivity to other mAbs included in CD44 cluster, on a variety lymphoid and nonlymphoid human tissues, and demonstrated similar bands on SDS-PAGE of 125I labeled lymphocyte lysates. This antigen is limited to cells of mature phenotype, and disappears in proliferating B cells in the germinal centers of the lymphoid follicles. CD44 is absent in pre-B and Burkitt cell lines. PKC activation mediate in vitro differentiation of pre-B cell lines. However, it is not involved in up-regulation of CD44 antigen expression.

Antibodies, Monoclonal↗

PubMed1991

HLA class I and II expression in rhabdomyosarcomas.

The expression of class I and II histocompatibility antigens was studied in 43 specimens of skeletal and in 36 specimens of cardiac muscle. Expression was also analyzed in the tumoral counterpart of striated muscle tissue, in 21 rhabdomyosarcomas (13 embryonic, 2 alveolar and 6 pleomorphic). Normal striated muscle showed very weak HLA class I antigen expression and no class II expression. The rhabdomyosarcomas showed increased class I expression, possibly related to malignant transformation of this type of tissue and with the degree of cellular differentiation. In our series of rhabdomyosarcomas, we observed class II neoexpression only in some HLA class I positive specimens, which was unrelated to the degree of cellular differentiation in these tumors.

Cell Differentiation↗

PubMed1991

Loss of HLA heavy chain and beta 2-microglobulin in HLA negative tumours.

Expression of HLA class I antigens on the cell surface requires the association of heavy chain, endogenous peptide and beta 2-microglobulin (beta 2m). We have studied the expression of free class I heavy chain and beta 2m in cryostatic sections of 51 HLA class I negative human tumours of different origin. Selective low expression of beta 2m was observed only on the 9 colorectal adenocarcinomas. In contrast, simultaneous under-expression of heavy chain and beta 2m was detected in the rest of the tumours studied. Our results suggest that the accumulation in the cytoplasm of HLA heavy chain in HLA negative colorectal carcinomas is a particular mechanism that cannot be applied to all HLA negative tumours.

Adenocarcinoma↗

PubMed1991

DNA-RFLP analysis and genotyping of HLA-DR alleles in the south of Spain.

42 healthy individuals previously HLA-DR typed by serology, were HLA class II typed using RFLPs. The pattern of hybridization revealed allele specific DNA fragments for some DR specificities. Three fragments highly associated with HLA-DR antigens: a 7 kb RFLP with DR1; a 10 kb RFLP with DR-3/DR-5 and a 13 kb band with DR4/DR7.

Alleles↗

PubMed1991

MHC expression on human tumors--its relevance for local tumor growth and metastasis.

MHC class I and II molecules play an important role in specific interactions with cells of the immune system. Endogenous or exogenous antigens are presented to the clonotypic receptor of T cells as small peptides associated to MHC molecules. Qualitative or quantitative variation in the expression of these molecules in the surface of tumor cells could have important implications in anti-tumor immune responses. We have analysed 344 human tumors for HLA class I and II expression and found that 10-30% of tumors present a total loss of HLA ABC molecules. In addition, HLA-A or -B locus-specific losses were also detected. These alterations have been correlated with tumor aggressiveness in breast and laryngeal carcinomas. We also have observed that the expression of HLA ABC molecules in autologous metastasis did not always correspond with the expression detected in the primary tumor. In laryngeal carcinomas HLA-DR expression was associated with an excellent prognosis. We have observed in most tumors that the absence of class I molecules usually corresponds with a simultaneous loss of heavy chain and beta 2 microglobulin expression and with a low level of the mRNA specific for class I genes. Nevertheless, a variety of mechanisms are involved since in colon tumors the absence of expression is caused by beta 2 microglobulin down regulation. Also post-transcriptional mechanisms may be involved in the differential expression of HLA-A and -B locus products. There is no doubt that a more exact knowledge of the mechanisms that produce alteration in the expression of these antigens will help to manipulate MHC gene expression in human tumors and to induce a more efficient immune response.

Gene Expression Regulation, Neoplastic↗

PubMed1991

Tumor aggressiveness and MHC class I and II antigens in laryngeal and breast cancer.

HLA class I and II molecules play an important role in the immune system. Thus, alterations in their expression could affect the immunosurveillance against tumors. The expression of these molecules may therefore be one of the factors responsible for oncogenicity, due to their role as restriction elements in T cell recognition. A series of 78 primary laryngeal and 94 breast tumors were evaluated for the expression of MHC class I and II antigens, using monoclonal antibodies. We found a number of cases presenting total loss of HLA-ABC and selective losses of HLA-A and HLA-B antigens. These losses were statistically associated with clinical and pathological parameters such as T stage, degree of differentiation, scores according to the malignancy grading systems and degree of leukocytic infiltration. On the other hand, the de novo expression of DR antigens in laryngeal cancer was found in a group of neoplasms having a peculiar clinical behavior and excellent prognosis. Expression of class II molecules in breast tumors was also related to a better differentiation and prognosis. Thus, HLA class I and II antigen expression is closely related to aggressiveness and prognosis of laryngeal and breast cancer.

Adenocarcinoma↗