Search PubMed⌕ Search

Biomedical subjects

R E Hammer

Publications and source records attributed to R E Hammer.

At least 73 records · Page 4Linked to original sources
PubMed1996

Normal luminal bacteria, especially Bacteroides species, mediate chronic colitis, gastritis, and arthritis in HLA-B27/human beta2 microglobulin transgenic rats.

Genetic and environmental factors are important in the pathogenesis of clinical and experimental chronic intestinal inflammation. We investigated the influence of normal luminal bacteria and several groups of selected bacterial strains on spontaneous gastrointestinal and systemic inflammation in HLA-B27 transgenic rats. Rats maintained germfree for 3-9 mo were compared with littermates conventionalized with specific pathogen-free bacteria. Subsequently, germfree transgenic rats were colonized with groups of five to eight bacteria that were either facultative or strictly anaerobic. Transgenic germfree rats had no gastroduodenitis, colitis, or arthritis, but developed epididymitis and dermatitis to the same degree as conventionalized rats. Colonic proinflammatory cytokine expression was increased in transgenic conventionalized rats but was undetectable in germfree and nontransgenic rats. Colitis progressively increased over the first 4 wk of bacterial exposure, then plateaued. Only transgenic rats colonized with defined bacterial cocktails which contained Bacteroides spp. had colitis and gastritis. Normal luminal bacteria predictably and uniformly induce chronic colonic, gastric and systemic inflammation in B27 transgenic F344 rats, but all bacterial species do not have equal activities.

Animals↗
PubMed1996

Defective forebrain development in mice lacking gp330/megalin.

gp330/megalin, a member of the low density lipoprotein (LDL) receptor gene family, is expressed on the apical surfaces of epithelial tissues, including the neuroepithelium, where it mediates the endocytic uptake of diverse macromolecules, such as cholesterol-carrying lipoproteins, proteases, and antiproteinases. Megalin knockout mice manifest abnormalities in epithelial tissues including lung and kidney that normally express the protein and they die perinatally from respiratory insufficiency. In brain, impaired proliferation of neuroepithelium produces a holoprosencephalic syndrome, characterized by lack of olfactory bulbs, forebrain fusion, and a common ventricular system. Similar syndromes in humans and animals are caused by insufficient supply of cholesterol during development. Because megalin can bind lipoproteins, we propose that the receptor is part of the maternal-fetal lipoprotein transport system and mediates the endocytic uptake of essential nutrients in the postgastrulation stage.

Animals↗
PubMed1996

RAP, a specialized chaperone, prevents ligand-induced ER retention and degradation of LDL receptor-related endocytic receptors.

The multifunctional low density lipoprotein (LDL) receptor-related protein (LRP) forms a complex with a receptor-associated protein (RAP) within the secretory pathway. RAP inhibits ligand binding to LRP and is required for normal functional expression of LRP in vivo, suggesting a physiological function as a specialized chaperone. We have used RAP-deficient mice, generated by gene targeting, to investigate the role of RAP in the biosynthesis and biological activity of LRP and other members of the LDL receptor gene family in various organs and in embryonic fibroblasts. Our results demonstrate that RAP is required for the proper folding and export of the receptors from the endoplasmic reticulum (ER) by preventing the premature binding of co-expressed ligands. Overexpression of apolipoprotein E (apoE), a high affinity ligand for LRP, results in dramatically reduced cellular LRP expression, an effect that is prevented by co-expression of RAP. RAP thus defines a novel class of molecular chaperones that selectively protect endocytic receptors by binding to newly synthesized receptor polypeptides, thereby preventing ligand-induced aggregation and subsequent degradation in the ER.

Animals↗
PubMed1996

Rat spermatogenesis in mouse testis.

Recently, transplantation of mouse donor spermatogonial stem cells from a fertile testis to an infertile recipient mouse testis was described. The donor cells established spermatogenesis in the seminiferous tubules of the host, and normal spermatozoa were produced. In the most successful transplants, the recipient mice were fertile and sired up to 80 per cent of progeny from donor cells. Here we examine the feasibility of transplanting spermatogonial stem cells from other species to the mouse seminiferous tubule to generate spermatogenesis. Marked testis cells from transgenic rats were transplanted to the testes of immunodeficient mice, and in all of 10 recipient mice (in 19 of 20 testes), rat spermatogenesis occurred. Epididymides of eight mice were examined, and the three from mice with the longest transplants (> or = 110 days) contained rat spermatozoa with normal morphology. The generation of rat spermatogenesis in mouse testes suggests that spermatogonial stem cells of many species could be transplanted, and opens the possibility of xenogeneic spermatogenesis for other species.

Animals↗
PubMed1996

Synaptophysin, a major synaptic vesicle protein, is not essential for neurotransmitter release.

Synaptophysin (syp I) is a synaptic vesicle membrane protein that constitutes approximately 7% of the total vesicle protein. Multiple lines of evidence implicate syp I in a number of nerve terminal functions. To test these, we have disrupted the murine Syp I gene. Mutant mice lacking syp I were viable and fertile. No changes in the structure and protein composition of the mutant brains were observed except for a decrease in synaptobrevin/VAMP II. Synaptic transmission was normal with no detectable changes in synaptic plasticity or the probability of release. Our data demonstrate that one of the major synaptic vesicle membrane proteins is not essential for synaptic transmission, suggesting that its function is either redundant or that it has a more subtle function not apparent in the assays used.

Animals↗
PubMed1996

Rat MHC-linked peptide transporter alleles strongly influence peptide binding by HLA-B27 but not B27-associated inflammatory disease.

Rats transgenic for the human MHC molecule HLA-B27 were used to study the effect of two alleles, cima and cimb, which are associated with peptide transport by the MHC-encoded Tap2 transporter, on the function of HLA-B27 as a restriction element for CTL recognition of the male H-Y minor H Ag and on the multisystem inflammatory disease characteristic of B27 transgenic rats. Anti-H-Y CTL generated in cima B27 transgenic rats lysed male B27 cimb/b targets significantly less well than cima/a or cima/b targets. Addition of exogenous H-Y peptides to male B27 cimb/b targets increased susceptibility to lysis to the level of cima/a targets. Male B27 cimb/b cells were less efficient than cima/a cells in competitively inhibiting CTL lysis of female B27 cima/a targets sensitized with exogenous H-Y peptides. 3H-Labeled peptides eluted from B27 molecules of lymphoblasts from rats of two cimb and three cima RT1 haplotypes showed that the cimb peptide pool favors comparatively longer and/or more hydrophobic peptides. These results indicate that RT1-linked Tap2 polymorphism in the rat strongly influences peptide loading of HLA-B27. Nonetheless, the prevalence and severity of multisystem inflammatory lesions were comparable in backcross rats bearing either cima/b or cimb/b. It thus appears either that binding of specific peptides to B27 is unimportant in the pathogenesis of B27-associated disease or that the critical peptides, unlike H-Y and many others, are not influenced by Tap transporter polymorphism.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1996

T cells, but not thymic exposure to HLA-B27, are required for the inflammatory disease of HLA-B27 transgenic rats.

Rats transgenic for the human MHC class I gene HLA-B27 are susceptible to a spontaneous multisystem inflammatory disease that resembles human B27-associated disease. This disease requires a high level of expression of the B27 transgene product in cells of hemopoietic origin and can be adoptively transferred to B27 transgenic or nontransgenic rats by transplantation of bone marrow (BM) or fetal liver (FL) cells. To investigate the role played by T cells and the thymus in the disease process, we produced congenitally athymic rnu/rnu F344 rats carrying the disease-prone B27 transgenic locus of the 33-3 line. Transgenic nude rats were protected from disease manifestations. This protection was abated by reconstitution with T cells from euthymic donors of the 33-3 line, with CD4 T cells being more efficient than CD8 T cells in transferring disease. Lethally irradiated, adult-thymectomized (ATX), nontransgenic recipients reconstituted with intact BM, T cell-depleted BM, FL, or nude BM from syngeneic disease-prone lines all developed disease. Pretreatment of the ATX nontransgenic recipients to deplete T cells enhanced the subsequent transferred disease. The inflammatory disease of B27 transgenic rats is thus T cell-dependent. The relevant T cells do not need to encounter B27 in the thymus, and residual radioresistant and/or extrathymically derived host T cells are sufficient to mediate the adoptively transferred disease. The data are most consistent with a model of B27-mediated disease arising from a failure of tolerance and requiring a population of CD4 T cells.

Animals↗
PubMed1996

Experimental spondyloarthropathy in HLA-B27 transgenic rats.

Rats of the LEW, F344, and PVG strains bearing high copy numbers of transgenes for HLA-B27 and human beta 2-microglobulin develop a multisystem disorder, many features of which resemble aspects of the human spondyloarthropathies, most dramatically axial and peripheral arthropathy. The disease is not seen in rats with comparable expression of HLA-B7, and so is relatively specific for HLA-B27. It requires T cells and another bone marrow derived cell, as well as an intact gut flora. Characteristic cytokine profiles distinguish the inflammatory infiltrates in the gut and the joint, and the earliest T cell infiltrates at these sites are polyclonal. Polymorphism at the Tap-2 peptide transporter locus has no significant effect on disease, whereas the DA strain background abrogates disease. The molecular role of the B27 molecule in disease remains to be elucidated, but there is good reason to believe that it will eventually yield to continued investigation of this animal model.

Animals↗
PubMed1996

Sustained somatic gene inactivation by viral transfer of Cre recombinase.

Transgenic and knockout mice have proven invaluable tools for analyzing physiologically relavant functions of numerous genes. In some cases, however, pleiotropic effects that result from a variable requirement for a particular gene in different tissues, cell types, or stages of embryonic development may complicate the analysis due to a complex phenotype or embryonic lethality. The loxP/Cre-mediated recombination system, which allows tissue-specific gene targeting in the mouse, can be used to overcome these problems. A limitation of current methods is that a mouse carrying a loxP-tagged gene must be crossed with a transgenic mouse expressing the Cre recombinase in an appropriate tissue to obtain the desired gene rearrangement. We have used recombinant adenovirus carrying the Cre recombinase to induce virtually quantitative somatic cell gene disruption in the liver. The targeted gene was the multifunctional low-density lipoprotein receptor-related protein (LRP), a cell surface receptor for alpha 2-macroglobulin and other ligands. Transient expression of Cre following adenoviral infection produced the predicted gene rearrangement, functionally inactivating LRP in the liver. Rearrangement occurred within 6 days after infection and remained stable for at least 28 days. The results demonstrate the suitability of adenoviral Cre gene transfer to induce long-term, quantitative, and temporally controlled gene disruption in the mouse.

Adenoviridae↗
PubMed1995

Mutagenesis of the human apolipoprotein B gene in a yeast artificial chromosome reveals the site of attachment for apolipoprotein(a).

Lipoprotein(a) [Lp(a)] is a lipoprotein formed by the disulfide linkage of apolipoprotein (apo) B100 of a low density lipoprotein particle to apolipoprotein(a). Prior studies have suggested that one of the C-terminal Cys residues of apo-B100 is involved in the disulfide linkage of apo-B100 to apo(a). To identify the apo-B100 Cys residue involved in the formation of Lp(a), we constructed a yeast artificial chromosome (YAC) spanning the human apo-B gene and used gene-targeting techniques to change Cys-4326 to Gly. The mutated YAC DNA was used to generate transgenic mice expressing the mutant human apo-B100 (Cys4326Gly). Unlike the wild-type human apo-B100, the mutant human apo-B100 completely lacked the ability to bind to apo(a) and form Lp(a). This study demonstrates that apo-B100 Cys-4326 is required for the assembly of Lp(a) and shows that gene targeting in YACs, followed by the generation of transgenic mice, is a useful approach for analyzing the structure of large proteins coded for by large genes.

Animals↗
PubMed1995

A transgene coding for a human insulin analog has a mitogenic effect on murine embryonic beta cells.

We have investigated the mitogenic effect of three mutant forms of human insulin on insulin-producing beta cells of the developing pancreas. We examined transgenic embryonic and adult mice expressing (i) human [AspB10]-proinsulin/insulin ([AspB10]ProIN/IN), produced by replacement of histidine by aspartic acid at position 10 of the B chain and characterized by an increased affinity for the insulin receptor; (ii) human [LeuA3]insulin, produced by the substitution of leucine for valine in position 3 of the A chain, which exhibits decreased receptor binding affinity; and (iii) human [LeuA3, AspB10]insulin "double" mutation. During development, beta cells of AspB10 embryos were twice as abundant and had a 3 times higher rate of proliferation compared with beta cells of littermate controls. The mitogenic effect of [AspB10]ProIN/IN was specific for embryonic beta cells because the rate of proliferation of beta cells of adults and of glucagon (alpha) cells and adrenal chromaffin cells of embryos was similar in AspB10 mice and controls. In contrast to AspB10 embryos, the number of beta cells in the LeuA3 and "double" mutant lines was similar to the number in controls. These findings indicate that the [AspB10]ProIN/IN analog increased the rate of fetal beta-cell proliferation. The mechanism or mechanisms that mediate this mitogenic effect remain to be determined.

Amino Acid Sequence↗
PubMed1995

Essential functions of synapsins I and II in synaptic vesicle regulation.

Synaptic vesicles are coated by synapsins, phosphoproteins that account for 9% of the vesicle protein. To analyse the functions of these proteins, we have studied knockout mice lacking either synapsin I, synapsin II, or both. Mice lacking synapsins are viable and fertile with no gross anatomical abnormalities, but experience seizures with a frequency proportional to the number of mutant alleles. Synapsin-II and double knockouts, but not synapsin-I knockouts, exhibit decreased post-tetanic potentiation and severe synaptic depression upon repetitive stimulation. Intrinsic synaptic-vesicle membrane proteins, but not peripheral membrane proteins or other synaptic proteins, are slightly decreased in individual knockouts and more severely reduced in double knockouts, as is the number of synaptic vesicles. Thus synapsins are not required for neurite outgrowth, synaptogenesis or the basic mechanics of synaptic vesicle traffic, but are essential for accelerating this traffic during repetitive stimulation. The phenotype of the synapsin knockouts could be explained either by deficient recruitment of synaptic vesicles to the active zone, or by impaired maturation of vesicles at the active zone, both of which could lead to a secondary destabilization of synaptic vesicles.

Animals↗
PubMed1995

Functional expression of low density lipoprotein receptor-related protein is controlled by receptor-associated protein in vivo.

The 39-kDa receptor-associated protein (RAP) associates with the multifunctional low density lipoprotein (LDL) receptor-related protein (LRP) and thereby prevents the binding of all known ligands, including alpha 2-macroglobulin and chylomicron remnants. RAP is predominantly localized in the endoplasmic reticulum, raising the possibility that it functions as a chaperone or escort protein in the biosynthesis or intracellular transport of LRP. Here we have used gene targeting to show that RAP promotes the expression of functional LRP in vivo. The amount of mature, processed LRP is reduced in liver and brain of RAP-deficient mice. As a result, hepatic clearance of alpha 2-macroglobulin is impaired and remnant lipoproteins accumulate in the plasma of RAP-deficient mice that also lack functional LDL receptors. These results are consistent with the hypothesis that RAP stabilizes LRP within the secretory pathway. They also suggest a further mechanism by which the activity of an endocytic receptor may be modulated in vivo.

Animals↗
PubMed1995

E6 and E7 expression from the HPV 18 LCR: development of genital hyperplasia and neoplasia in transgenic mice.

Human papillomavirus type 18 infection is highly associated with malignant tumors of the genital tract. To investigate the tissue specificity of the HPV long control region (LCR) and the transforming ability of the E6-E7 oncoproteins, an HPV-18 transgene containing the viral LCR and E6 and E7 genes was introduced into mice. Three founder males exhibited enlarged seminal vesicles and preputial glands by 50 weeks of age. A line of transgenic mice was established by in vitro fertilization, and subsequent generations of transgenic males and females were monitored for lesions. Approximately 80% of hemizygous transgenic males exhibited enlarged seminal vesicles and preputial glands as early as 12 weeks of age. Histological examination indicated that this enlargement was due to distension by fluid, along with polyploid hyperplasia of the lining secretory epithelium. E6 and E7 transcripts were limited to affected organs and kidney. Approximately 41% of transgenic females developed cervical neoplasms between 1-2 years of age. Histologically, tumors were mesenchymal rather than epithelial in origin. E6 and E7 transcripts were restricted to cervical tumor tissue and kidney. These findings suggest that the HPV-18 LCR has an element(s) which directs expression specifically to the urogenital tract in transgenic mice.

Animals↗
PubMed1995

Cooperation between elements of an organ-specific transcriptional enhancer in animals.

The elastase I gene enhancer that specifies high levels of pancreatic transcription comprises three functional elements (A, B, and C). When assayed individually in transgenic mice, homomultimers of A are acinar cell specific, those of B are islet specific, and those of C are inactive. To determine how the elements interact in the elastase I enhancer and to investigate further the role of the C element, we have examined the activity of the three possible combinations of synthetic double elements in transgenic animals. Combining the A and B elements reconstitutes the exocrine plus endocrine specificity of the intact enhancer with an increased activity in acinar cells compared with that in the A homomultimer. The B element therefore plays a dual role: in islet cells it is capable of activating transcription, whereas in acinar cells it is inactive alone but greatly augments the activity specified by the A element. The C element augments the activity of either the A or B element without affecting their pancreatic cell type specificity. The roles of each element were verified by examining the effects of mutational inactivation of each element within the context of the elastase I enhancer. These results demonstrated that when tested in animals, the individual enhancer elements can perform discrete, separable functions that combine additively for cell type specificity and cooperatively for the overall strength of a multielement stage- and site-specific transcriptional enhancer.

Animals↗
PubMed1995

Relative contributions of apolipoprotein(a) and apolipoprotein-B to the development of fatty lesions in the proximal aorta of mice.

Transgenic mice expressing transgenes for both human apolipoprotein B-100 (h-apoB) and apolipoprotein(a) [apo(a)] were fed a high-fat, atherogenic diet for 14 weeks to examine the effect of lipoprotein(a) [Lp(a)]on the development of aortic fatty lesions. The extent of lesions in the proximal region of the aorta of Lp(a) mice was measured by use of a computer-assisted image analysis of 20 sections per animal and compared with that of nontransgenic mice as well as mice expressing either the apo(a) or h-apoB transgene. The control (n = 23) and apo(a) (n = 22) transgenic mice had very small mean lesions areas (607 versus 128 microns2 per section). The h-apoB-expressing mice (n = 20) had significantly higher mean lesion areas (3288 microns2 per section) than either the control or apo(a) transgenic animals. Coexpression of apo(a) and h-apoB transgenes resulted in only a modest increase in lesion area (4678 microns2 per section, n = 19). Thus, the expression of human apo(a) in C57BL/6/SJL hybrid mice fed an atherogenic diet failed to significantly potentiate the development of aortic fatty lesions in the absence or presence of high levels of h-apoB.

Animals↗