Search PubMed⌕ Search

Biomedical subjects

J Hilgers

Publications and source records attributed to J Hilgers.

At least 91 records · Page 5Linked to original sources
PubMed1987

Mouse lymphotoxin and tumor necrosis factor: structural analysis of the cloned genes, physical linkage, and chromosomal position.

Lymphotoxin (LT) and tumor necrosis factor (TNF) are cytotoxic and immunoregulatory lymphokines which have similar activities but are produced by different cell types. We have cloned the murine LT and TNF genes from a lambda:mouse DNA recombinant library, using as probes synthetic oligonucleotides defined by portions of the human LT or TNF cDNA sequences. Analysis of the genomic clones indicates that the LT and TNF genes are physically linked, i.e., approximately 1.2 kb separates the 3' end of LT from the 5' end of TNF genes. By using, first, a series of recombinant inbred lines, and second, a series of H-2-recombinant congenic strains, we determined that the LT/TNF gene cluster lies on chromosome 17, closely linked to the H-2D end of the murine H-2 complex. Comparison of the primary sequence of murine and human LT revealed that the intron-exon structure of murine LT is similar in these two species. Comparison of the predicted amino acid sequences of murine and human LT indicates that the proteins are about 72% homologous with much greater sequence conservation in regions encoding the COOH-terminal portion. Comparison of the 5' flanking sequence of LT to a number of genes that are specifically expressed in activated T cells reveals a number of conserved sequences that may play a role in control of these genes.

Amino Acid Sequence↗
PubMed1987

Oncogene expression during progression of mouse mammary tumor cells; activity of a proviral enhancer and the resulting expression of int-2 is influenced by the state of differentiation.

The RAC cell lines are derived from a mammary tumor induced by the mouse mammary tumor virus (MMTV). Polygonal cells have retained many characteristics of epithelial cells and induce adenocarcinomas; cuboidal cells are poorly tumorigenic; and elongated cells produce highly malignant sarcoma-like tumors. MMTV proviral integrations, one of which has cis-activated the int-2 gene, demonstrated that the lines are clonal descendents from a single tumor cell. Polygonal cells have a high constitutive expression of MMTV and contain int-2 RNA. In contrast, the cuboidal and the elongated cells show no detectable expression of int-2, even in the presence of glucocorticoid hormone. Transcription of MMTV in these cells is also low but can be stimulated by dexamethasone, albeit to levels lower than in polygonal cells. Thus, expression of int-2 seems to be caused by an enhancing activity on the MMTV provirus which is not dependent on steroid hormone and is specific for mammary tumor cells with epithelial characteristics. Progression in this cell system does not require sustained expression of int-2.

Adenocarcinoma↗
PubMed1987

Evolutionary relationships between laboratory mice and subspecies of Mus musculus based on the genetic study of pancreatic proteinase loci, Prt-1, Prt-2, Prt-3, and Prt-6.

Various patterns of mouse pancreatic proteinase activity bands were observed on agarose gel electrophoresis. Prt-1a and Prt-1b genes control the positive (PRT-1A) and negative (PRT-1B) expression of tryptic band V, respectively; Prt-2a and Prt-2b correspond to chymotryptic bands II (PRT-2A) and III (PRT-2B); Prt-3a and Prt-3b control the low (PRT-3A) and high (PRT-3B) tryptic activities of band IV; the Prt-1 and Prt-3 loci are closely linked on the same chromosome; Prt-6a and Prt-6b correspond to tryptic bands I (PRT-6A) and I' (PRT-6B). Twenty-four laboratory strains from the United States showed the phenotype PRT-1A, PRT-3A, and PRT-2A. Of laboratory strains established in Europe, 6 showed PRT-1A, PRT-3A, and PRT-2A, and 10 had PRT-1B, PRT-3A, and PRT-2A bands. Most wild mice around the world and their descendants showed the phenotype PRT-1B, PRT-3B, and PRT-2A. Only the phenotype of M. m. brevirostris was PRT-1A, PRT-3A, and PRT-2A, which was the same as most laboratory inbred strains. PRT-2B was observed mainly in Japanese (M.m. molossinus) and Korean (M.m. yamashinai) wild mice. PRT-6B was detected only in Mus spicilegus and Mus caroli, but all other mice including wild populations and laboratory strains showed PRT-6A. New biochemical phenotypes such as PRT-2C and PRT-3C were also found in this study.

Alleles↗
PubMed1987

Production of monoclonal antibodies to thyroglobulin by in vitro immunization with a free synthetic peptide.

A synthetic peptide corresponding to amino acids 1-19 of thyroglobulin was used to test the possibility of generating protein-reactive monoclonal antibodies by immunization in vitro with a synthetic peptide as antigen. Splenocytes from non-immunized Balb/c mice were cultured in serum-free medium for 3 days in the presence of thymocyte-conditioned medium and the synthetic peptide prior to fusion with SP2/0 murine myeloma cells. The synthetic peptide was used in its free form, i.e. not coupled to a protein carrier. Hybridomas secreting monoclonal antibodies reactive with the synthetic peptide were obtained after immunization in vitro with as little as 10 ng/ml of the synthetic peptide. Between 50 and 70% of the primary clones obtained in different experiments produced monoclonal antibodies also reactive with the intact protein. Six stable hybridomas were isolated; all produced antibodies of the IgM class. We conclude that immunization in vitro with a free synthetic peptide is an efficient method for the generation of monoclonal antibodies reactive with the intact protein.

Animals↗
PubMed1987

Immunoelectron microscopic localization of mammary tumor virus (MTV) antigens in normal and cancerous mammary glands of mice.

Peroxidase-labeled Fab' fragments of rabbit antisera against gp52 (major envelope protein) and A-particles of mammary tumor virus (MTV) were prepared and used for investigation by immunoelectron microscopy of the replication cycle of MTV-specific envelope and core antigens in normal and malignant mammary gland cells of female mice. The specificity of the antisera was proven by absorption tests and lack of reactivity to MTV-free mammary tissues. Periodate-lysine-paraformaldehyde (PLP) fixation sufficiently preserved the antigenicity of gp52, while Zamboni's fixative was useful to preserve the core antigen. Saponin pretreatment was necessary to reveal the intracellular antigen of A particles but had no influence on gp52. In addition to its presence at the envelope of D particles, gp52 was clearly associated with the biomembrane system, including the nuclear membrane, endoplasmic reticulum, Golgi apparatus and plasma membrane independent of the presence of virus particles. In mammary tumors, a significant level of gp52 antigen was often expressed on the entire cell surface membrane. In contrast, it was localized only to the apical plasma membrane in normal mammary gland cells. A particle antigens were confined to the intracytoplasmic A particles, usually visible as clusters, and to the inner part of B particles. These ultrastructural findings support the available biochemical data on the morphogenesis of MTV particles.

Animals↗
PubMed1987

Classification of routinely processed anaplastic large cell tumours with a small panel of antibodies. An immunohistochemical study with clinical follow-up.

A proportion of anaplastic large cell tumours is difficult to classify on sections of routinely processed, paraffin-embedded tissue. Differentiation into large cell lymphoma, carcinoma, melanoma or sarcoma is important in order to assess prognosis and proper treatment. Although the use of immunohistochemistry has been reported in the differentiation between some of these types of neoplasms, no antibody panel, which can directly differentiate all of them, has been described. In the present study we evaluated the value of a panel of 5 antibodies for the classification of 29 anaplastic large cell tumours, which could not be classified by experienced pathologists using conventional histological and histochemical techniques. The panel, which can be used on routinely fixed paraffin-embedded tissue, consisted of 5 different antibodies directed against keratin, vimentin, the human milk-fat globule membrane antigen MAM-6, a melanoma associated antigen and common leucocyte antigen. The use of this panel directly resulted in a definite diagnosis in 95% of the cases and provided valuable information for the diagnosis in the remaining cases. The diagnosis was confirmed by additional marker studies and electron microscopy. Moreover, clinical follow-up, including treatment data, was in accordance with the diagnosis based on the panel.

Aged↗
PubMed1987

Immunohistochemical localization of the epithelial marker MAM-6 in invasive malignancies and highly dysplastic adenomas of the large intestine.

MAM-6 antigen is detectable in a large variety of formalin fixed and paraffin embedded normal and neoplastic tissues throughout the body with some exceptions. One such exception is the large intestine in which normal mucosa has been found negative for MAM-6, while colorectal carcinomas express the antigen in almost all cases. We examined the expression of MAM-6 in 147 benign epithelial tumors (adenomas) of the colorectum and compared this with the detectability of the antigen in 123 colorectal cancers and in 108 non-neoplastic mucosa specimens. The tissue samples were subjected to an indirect immunoperoxidase assay applying the MoAb 115D8 which recognizes the epitope "a" of MAM-6. A majority of the carcinomas (99.2%) and of the adenomas (61.4%) exhibited reactivity for MAM-6 "a", whereas only 8 of the 108 non-neoplastic biopsy specimens (= 7.4%) exhibited faint focal staining. In adenomas, positive reactions were only seen in highly dysplastic tissue areas, in part reflecting carcinomata in situ. The reactivity in four non-neoplastic tissue samples could be explained by shedding of the antigen from carcinomas into glandular structures of the mucosa. Two out of 32 cases exhibiting severe colitis were also focally positive. The detectability of MAM-6 "a" in routinely processed specimens from the large intestine is therefore considered as being closely associated with malignant and premalignant processes. According to a limited study with the antibodies 139H2 and 140C1, this holds true for two additional MAM-6 epitopes. MAM-6 might be considered as a marker of severe (premalignant) dysplasia in adenomas of the large intestine. Further studies are needed to clarify if there is a biological (possibly prognostic) difference between MAM-6 positive and negative high epithelial atypia in these adenomas.

Adenocarcinoma↗
PubMed1986

Monoclonal antibodies detecting different epitopes on the Forssman glycolipid hapten.

Two hybridomas, derived by fusing mouse myeloma cells with spleen cells from a rat immunized with mouse mammary tumors, have been shown to produce antibodies that recognize cell surface antigens on mesenchymal cells in a variety of tissues. Evidence presented in this report suggests that these antibodies detect overlapping epitopes on the Forssman glycolipid hapten (GalNAc alpha 1-3GalNAc beta 1-3Gal alpha 1-4Gal beta 1-4Glc beta 1-1Cer). One antibody (33B12) reacts with the terminal sugar sequence GalNAc alpha 1-3GalNAc and is specific for Forssman. The other antibody (117C9) recognizes the internal sugar sequence GalNAc beta 1-3Gal. The terminal sugar sequence GalNAc beta 1-3Gal in globoside, as well as the internal sugar sequence GalNAc beta 1-4Gal in asialo-GM1, is not recognized as an antigenic determinant by 117C9. Nevertheless, the 117C9 antibody does not react exclusively with the Forssman antigen. In a lipid extract fractionated by Folch partition of mouse mammary tumors, the antibody also detects other glycolipids.

Animals↗
PubMed1986

Production of murine monoclonal antibodies against human thyroglobulin using an in vitro immunization procedure in serum-free medium.

A serum-free in vitro immunization method for the generation of hybridomas producing specific antibodies to an antigen is described. The method was tested with human thyroglobulin as antigen. The serum-free medium used (Yssel et al., 1984) consisted of Iscove's modification of Dulbecco's modified Eagle's medium, supplemented with albumin, transferrin, insulin, ethanolamine and linoleic, oleic and palmitic acids. An optimal response was obtained when splenocytes from BALB/c mice were cultured for 3 days in the presence of 1.5 nM thyroglobulin and thymocyte-conditioned medium prior to fusion with SP2/0 myeloma cells and seeding of the fused cells in microtitre plates. The frequency of positive wells, defined as the number of wells secreting anti-(thyroglobulin) antibodies/number of viable cells used for the fusion, was 1.6 X 10(-6) +/- 0.25 X 10(-6) (mean +/- SD; n = 4). Eight stable clones producing anti-(thyroglobulin) antibodies were isolated. One clone (3D12) produced antibodies reacting only with human thyroglobulin. The antibodies produced by the other clones reacted with human, murine and porcine thyroglobulins. Seven of the clones produced antibodies of the IgM class and one clone produced IgG. The specificity of 3D12 (IgM) for human thyroglobulin and the absence of any reactivity with murine thyroglobulin provides evidence for a primary response of splenocytes in culture to the presence of an antigen.

Antibodies, Monoclonal↗
PubMed1986

Detection of surface antigens defined by monoclonal antibodies in primary mucinous breast carcinomas. Relation to prognostic factors and recurrence-free survival.

Three monoclonal antibodies, 67D11, 115H10 and 115C2, raised against human milk fat globule membranes, have been applied to 207 primary mucinous breast carcinomas. The tumours reacted positively in 18% (67D11), 54% (115H10), and 20% (115C2) of the cases. The detected epitopes (MAM-3a (67D11), MAM-3b (115H10), and MAM-3c (115C2)) have formerly been shown to be markers of differentiation in infiltrating ductal carcinomas. In the present group of mucinous breast carcinomas, statistically significant correlation to high risk factors, such as occurrence of primary lymph node metastases, large tumour size, and local invasion of the tumour into overlying skin or deep fascie, were found. Furthermore, the antigen expression was less marked in pure mucinous carcinomas as compared to carcinomas also presenting with non-mucinous tumour areas. Thus, especially the antigen MAM-3b, is more frequently present in mixed tumours, in large tumours, in tumours with local invasion, and in tumours with primary lymph node metastases. However, no association could be demonstrated between expression of MAM-3b and recurrence-free survival. Mucinous carcinomas of the breast apparently differ from other carcinomas not only with respect to morphology, but also in their pattern of antigenic expression in relation to other prognostic factors.

Adenocarcinoma, Mucinous↗
PubMed1986

Genetic mapping of Pim-1 putative oncogene to mouse chromosome 17.

Pim-1 is a putative oncogene activated in T-cell lymphomas induced by Moloney and AKR mink cell focus forming (MCF) viruses. We have determined the chromosomal localization of the Pim-1 gene in mice by Southern blot analysis of DNAs obtained from a panel of mouse-Chinese hamster somatic cell hybrids. The Pim-1 gene was localized on chromosome 17, a chromosome frequently aberrant in T-cell lymphomas. Two chromosomal regions, containing sequences homologous to regions within the Pim-1 locus, were localized on chromosome 6 and 16.

Animals↗
PubMed1986

Characterization and mapping of DNA sequence homologous to mouse U1a1 snRNA: localization on chromosome 11 near the Dlb-1 and Re loci.

A phage clone which contained a functional U1a1 snRNA gene was isolated from a mouse genomic library. A single copy fragment was isolated from the 3' flanking region of the U1a1 gene and used as a hybridization probe for Southern blotted DNAs from recombinant inbred strains of mice, mouse-hamster hybrid cells, and the offspring from backcrosses between BALB/c mice and mice which were heterozygous for the Rex (Re) marker. The results of these experiments prove that the U1a1 gene is located on chromosome 11 near the Delb-1 and Re loci.

Animals↗
PubMed1986

Development of mouse mammary gland: identification of stages in differentiation of luminal and myoepithelial cells using monoclonal antibodies and polyvalent antiserum against keratin.

The development of the mouse mammary gland was studied immunohistochemically using monoclonal antibodies against cell surface and basement membrane proteins and a polyclonal antibody against keratin. We have identified three basic cell types: basal, myoepithelial, and epithelial cells. The epithelial cells can be subdivided into three immunologically related cell types: luminal type I, luminal type II, and alveolar cells. These five cell types appear at different stages of mammary gland development and have either acquired or lost one of the antibody-defined antigens. The cytoplasmic distribution of several of these antigens varied according to the location of the cells within the mammary gland. Epithelial cells which did not line the lumen expressed antigens throughout the cytoplasm. These antigens were demonstrated on the apical site in situations where the cells lined the lumen. One antigen became increasingly basolateral as the cells became attached to the basement membrane. The basal cells synthesize laminin and deposit it at the cell base. They are present in endbuds and ducts and are probably the stem cells of the mammary gland. Transitional forms have been demonstrated which developmentally link these cells with both myoepithelial and (luminal) epithelial cells.

Animals↗
PubMed1986

Genetic marker patterns and endogenous mammary tumor virus genes in inbred mouse strains of Japan.

In order to establish the genetic relatedness of the inbred mouse strains kept in Nara, genetic marker patterns were determined in conjunction with a study on endogenous mammary tumor viral genes in these strains. Isoenzyme patterns combined with patterns of other genetic markers, show that the unrelatedness between various inbred strains of the dd stock is as high or even higher as between strains of known different origin and geneology. Based on endogenous viral gene patterns the dd stock derived mice can be subdivided into three group, DDD, DDN, DDO, KF and DD/Tbr. The DD/Tbr and its foster-nursed substrain (DD/Tbrf) have the lowest number of endogenous viral genes, i.e. two, while the other strains carry 4-6 such genes. The SLN and SHN strains, derived from a Swiss stock, have a similar pattern of viral genes different that of all other strains studied, also strains of Swiss origin from other sources, such as the NFS and the GR.

Animals↗
PubMed1986

In vitro differentiation and progression of mouse mammary tumor cells.

We have isolated clonal cell lines from a transplanted adenocarcinoma induced by the RIII strain of mouse mammary tumor virus in a BALB/c mouse. Three major morphological cell types of these lines are developmentally linked; polygonal cells give rise to cuboidal and then to elongated cells. All cell lines expressed markers that are characteristic of mammary basal cells. In addition, the polygonal lines contained cells that have cell markers and ultrastructural features of epithelial cells; in these lines an occasional cell was found with myoepithelial features. The cuboidal and elongated lines lacked many epithelial differentiation characteristics and showed no myoepithelial differentiation. The cell lines contained variable numbers of acquired mouse mammary tumor virus and ecotropic murine leukemia virus proviruses. The various subclones derived from the original cell lines contained, in addition to the acquired proviruses of the parental line, one or more unique proviruses of either mouse mammary tumor virus or ecotropic murine leukemia virus origin. These unique insertions were used as genotypic markers to demonstrate the clonal relationship of the cell lines. Both polygonal and elongated cells are tumorigenic and give rise to adenocarcinomas and sarcoma-like tumors, respectively. In contrast, the cuboidal cells are poorly tumorigenic. Since cuboidal cells are derived from the polygonal cells, this suggests that tumor progression in this system proceeds via intermediates that are either poorly or nontumorigenic.

Adenocarcinoma↗