Biomedical subjects

W Born

Publications and source records attributed to W Born.

At least 37 records · Page 2Linked to original sources

PubMed1998

Receptor activity modifying proteins regulate the activity of a calcitonin gene-related peptide receptor in rabbit aortic endothelial cells.

In Xenopus oocytes with an endogenous calcitonin gene-related peptide (CGRP) receptor, a receptor activity modifying protein (RAMP1) enhancing CGRP stimulated chloride currents of the cystic fibrosis transmembrane regulator was recently cloned [McLatchie, L.M. et al. (1998) Nature 393, 333-339]. Here, transient expression of RAMP1 in rabbit aortic endothelial cells (RAEC) brought about stimulation of cAMP accumulation by human (h) alphaCGRP with an EC50 of 0.41 nM. This was antagonized by a CGRP receptor antagonist alphaCGRP(8-37). Co-expression of RAMP3 together with RAMP1 reduced the maximal cAMP response to h alphaCGRP by 47% (P < 0.05). The cells also express RAMP2 encoding mRNA and an adrenomedullin (ADM) receptor coupled to stimulation of cAMP formation by hADM (EC50 0.18 nM). The latter was antagonized by an ADM receptor antagonist hADM(22-52). In conclusion, expression of a CGRP receptor in RAEC requires RAMP1. The same receptor presumably recognizes ADM making use of endogenous RAMP2. The results reveal competition between the different RAMPs in the regulation of CGRP/ADM receptor activity.

Adrenomedullin↗

PubMed1998

Early preferential stimulation of gamma delta T cells by TNF-alpha.

Although recent findings indicate that gamma delta T cells influence both early innate and Ag-specific adaptive host responses, it has remained unclear what triggers gamma delta T cell reactivity. Investigating very early T cell activation in mouse and human models of bacterial infection, we measured CD69 expression as an indicator of early cellular activation. Both murine alpha beta and gamma delta T cells responded polyclonally to systemic bacterial infections, and to LPS. However, gamma delta T cells responded more strongly to the bacteria and to LPS. In vitro LPS-stimulated human T cells showed a similar differential response pattern. We identified TNF-alpha as mediator of the early differential T cell activation, and of differential proliferative responses. The stronger response of gamma delta T cells to TNF-alpha was correlated with higher inducible expression levels of TNF-Rp75. Among unstimulated splenocytes, more gamma delta T cells than alpha beta T cells expressed CD44 at high levels. The data suggest that TNF-Rp75 determines the differential T cell reactivity, and that most gamma delta T cells in the normal spleen are present in a presensitized state. As TNF-alpha stimulates activated T cells, it may early preferentially connect gamma delta T cell functions with those of cells that produce this cytokine, including activated innate effector cells and Ag-stimulated T lymphocytes.

Animals↗

PubMed1998

Parathyroid hormone responses of cyclic AMP-, serum- and phorbol ester-responsive reporter genes in osteoblast-like UMR-106 cells.

Parathyroid hormone (PTH) and PTH-related protein interact with a G protein-coupled receptor linked to the activation of adenylyl cyclase and phospholipase C signaling pathways. Regulation by PTH of the expression of three distinct, stably transfected luciferase reporter genes responsive to cAMP (CRE-luc), serum (SRE-luc) and phorbol ester (TRE-luc) has been studied in rat osteoblast-like UMR-106 cells. Maximal 43-fold induction of CRE-luc expression occurred in response to 100 nM rat (r)PTH(1-34) (EC50=0.44 nM), but SRE-luc and TRE-luc remained unaffected. Maximal 2.8- and 3.4-fold inductions of SRE-luc by 10 ng/ml EGF and 100 nM phorbol ester (PMA) were suppressed with 100 nM rPTH(1-34) (IC50=0.04 and 0.15 nM, respectively). Similarly, 7.3-fold induction of TRE-luc by 100 nM PMA was inhibited to 50% with 100 nM rPTH(1-34) (IC50=0.5 nM). Activation of mitogen-activated protein kinase by EGF and PMA was also suppressed by rPTH(1-34). 1 mM 8-Br-cAMP and 0.1 mM forskolin mimicked all the effects of rPTH(1-34). In conclusion, the regulation of target genes by PTH in osteoblast-like UMR-106 cells is mediated by the activation of the cAMP/protein kinase A signaling pathway.

8-Bromo Cyclic Adenosine Monophosphate↗

PubMed1998

An inherited mutation associated with functional deficiency of the alpha-subunit of the guanine nucleotide-binding protein Gs in pseudo- and pseudopseudohypoparathyroidism.

Pseudohypoparathyroidism type Ia (PSP) is a disorder characterized by Albright's osteodystrophy, secondary hyperparathyroidism, lowered Gs activity, and resistance of the urinary cAMP excretion to exogenous PTH. The patients had raised basal serum levels of TSH and/or excessive TSH response to TRH. Here we have described a 38-bp deletion at the exon 1/intron 1 boundary of one Gs alpha allele in two mothers with pseudo-PSP and in six offsprings with PSP of a kindred with Albright's osteodystrophy. The deletion eliminates the splice donor site of exon 1. The pseudo-PSP patients presented decreased Gs activity, but normal urinary cAMP responses to PTH and normal TSH levels and responses to TRH. As monitored during 22 yr, they had normal serum levels of calcium and PTH. The findings demonstrate the same inherited functional defect of Gs alpha in two female patients with pseudo-PSP and in six of their offspring with PSP. The pathogenesis of clinical hypoparathyroidism remains to be clarified.

Adolescent↗

PubMed1997

Evidence that the same gamma delta T cells respond during infection-induced and autoimmune inflammation.

Inflammatory responses are induced in both testes of a mouse following injection of Listeria monocytogenes into one testis. Although the uninjected testis contains no detectable bacteria, it undergoes an autoimmune attack. Normally, the testis lacks lymphocytes, but in the infected and autoimmune state, both gamma delta and alpha beta T cells are found as infiltrates. Here, we have examined the repertoire of the infiltrating gamma delta T cells, using two different methods, and found a high frequency of V gamma 6/V delta 1 gamma delta T cells in both infected and autoimmune testes. All of these expressed the invariant V gamma 6/V delta 1 TCR previously reported. However, secondary gamma and delta transcripts present within V gamma 6/V delta 1 hybridomas indicated nonclonality. Interestingly, some of these secondary transcripts were derived from gamma gene rearrangements not previously found in this gamma delta T cell subset, implying a difference in its origin. The increase in V gamma 6/V delta 1 cells observed here in both infected and autoimmune testes, together with our previous finding of a preferential response by the same subset in Listeria-infected liver, indicates that their response is triggered by the inflammation rather than by the infectious agent or because they are already resident in the tissue. We and others have previously reported that the presence of gamma delta T cells during certain inflammatory conditions correlates with less host tissue damage. This result, together with the evidence presented here, further implies that a response by the V gamma 6/V delta 1 subset in some way exerts a controlling influence on the host inflammatory response.

Animals↗

PubMed1997

Parathyroid hormone-related protein (PTHrP) inhibits proliferation of Chinese hamster ovary cells stably transfected with a PTH/PTHrP receptor cDNA.

The effect of parathyroid hormone-related protein (PTHrP) on proliferation was examined in Chinese hamster ovary (CHO) cells transfected with a PTH/PTHrP receptor encoding cDNA. Treatment with chicken PTHrP(1-36) (chPTHrP) lowered the cell number to 49 +/- 2% of untreated controls after 6 days with a half-maximal effect at 1 nM. The effect was mimicked by human (h) PTH(1-34), Br-cAMP and forskolin, but not by the receptor antagonist hPTH(3-34). Reduction of cell number was accompanied by increased PTH/PTHrP receptor expression and persistently activated adenylyl cyclase, together with altered cell morphology from epithelial to spindle-like forms, clustered growth and increased phosphate uptake. chPTHrP increased [Ca2+]i, but failed to activate membrane bound protein kinase C (PKC). Pretreatment with chPTHrP did not affect phorbol ester stimulated PKC activity, and chPTHrP or serum evoked increases in [Ca2+]i. In conclusion, PTHrP induced inhibition of proliferation and altered cell morphology is mediated by increased adenylyl cyclase rather than by PKC-dependent mechanisms in CHO cells.

8-Bromo Cyclic Adenosine Monophosphate↗

PubMed1997

Tissue-specific mRNA expression of a calcitonin receptor-like receptor during fetal and postnatal development.

The distribution of calcitonin receptor-like receptor (CRLR) mRNA in developing rats was investigated by in situ hybridization. Signals were found in the piriform cortex, the central and basolateral amygdala and the amygdalostriatal transition area. Among peripheral organs, the CRLR was predominantly expressed in the lung. mRNA expression in blood vessels, liver, midgut, rectum and urethra was restricted to gestational days 16 and/or 20. The CRLR was thought to be a calcitonin gene-related peptide (CGRP) type 1 receptor (Aiyar et al., J. Biol. Chem., 271 (1996) 11325-11329). This contrasts with previously reported evidence that the CRLR is an orphan receptor with no identifiable interactions with CGRP and other related ligands (Flühmann et al., Biochem. Biophys. Res. Commun., 206 (1995) 341-347). In situ hybridization signals have not been detected in the cerebellum and the spleen known to present a high density of CGRP binding sites. The different regional distribution of CGRP receptor binding sites and CRLR mRNA implies the latter encoding a different CGRP receptor subtype.

Aging↗

PubMed1997

Reconstitution of SCID mice with haemopoietic precursors: a detailed analysis of gamma delta T-cell reconstitution.

A well-known characteristic of gamma delta T cells is that they are produced in waves during ontogeny, with cells expressing T-cell receptor V gamma 5 appearing early in fetal thymic ontogeny, followed by V gamma 6, then by other gamma delta T-cell types. In addition, evidence exists to suggest that the potential of haemopoietic precursors to generate different types of gamma delta T cells changes in ontogeny. We have used these observations as the basis for an extensive study of the potential for haemopoietic precursors isolated from fetal liver, neonatal spleen and adult bone marrow to reconstitute severe combined immunodeficient (SCID) mice. Mice that were reconstituted as newborns with fetal liver cells most closely resembled normal C.B-17 mice with respect to both lymphocyte numbers and subsets, while mice reconstituted with adult bone marrow had fewer cells than normal mice. This deficit spanned both T and B cells in all organs examined. Among the gamma delta T-cell subsets examined, the ability to reconstitute V gamma 4+ cells was particularly dependent on the ontogenic age of the reconstituting presursors, with fetal liver cells having the greatest capacity to generate V gamma 4+ cells, and adult bone marrow cells the least. The vast majority of the T cells produced in the reconstituted mice were of donor origin, and the level of reconstitution was found to be dependent upon some factor other than the presursor frequency.

Aging↗

PubMed1997

Biologically active human islet amyloid polypeptide/amylin in transgenic mice.

OBJECTIVE: Human islet amyloid polypeptide (hIAPP), also named amylin, is a pancreatic beta cell protein implicated in the pathogenesis of pancreatic islet amyloid formation and type 2 diabetes mellitus. To study the (patho)physiological roles of hIAPP, we have generated transgenic mice that overexpress hIAPP mRNA, in relation to endogenous mouse IAPP (mIAPP) mRNA, in pancreatic beta cells. The biological activity of human and mouse IAPP derived from pancreatic extracts was determined. METHODS: Pancreatic and plasma extracts of transgenic and control mice were analyzed by reversed-phase high-performance liquid chromatography (HPLC) and radioimmunoassay, yielding a separation of hIAPP from mIAPP. Biological activity of immunoreactive human and mouse IAPP components derived from pancreatic extracts was assessed by calcitonin receptor-mediated stimulation of cyclic AMP accumulation in T47D human breast carcinoma cells. RESULTS: The predominant immunoreactive human and mouse IAPP gene products had the retention times on HPLC analysis of the corresponding synthetic peptides. The ratio of bioactive over immunoreactive hIAPP and mIAPP was 0.93 +/- 0.18 and 1.19 +/- 0.56 respectively. In extracts of two plasma pools from 4 transgenic animals, hIAPP was 4.6- to 7-fold more abundant than mIAPP. CONCLUSION: This study has shown that correctly processed hIAPP produced in transgenic mouse pancreatic beta cells exhibits full biological activity. The results validate these transgenic mice for the study of (patho)physiological roles of hIAPP in vivo.

Amyloid↗

PubMed1997

Coexistence of novel amylin-binding sites with calcitonin receptors in human breast carcinoma MCF-7 cells.

Amylin, calcitonin (CT) and calcitonin gene-related peptide (CGRP) share limited structural homology including amino-terminal ring structures linked by a disulfide bridge and amidated carboxy-termini. Here, we have compared [125I]Bolton-Hunter-[Lys1] rat amylin ([125I]amylin) binding and the stimulation of cyclic AMP accumulation by human (h) amylin, hCT and hCGRP-I in the human breast carcinoma cell lines MCF-7 and T47D, which predominantly express hCT1a and hCT1b receptor isoforms (hCTR1a, hCTR1b) at a similar total number of hCT-binding sites. In MCF-7 cells, half-maximal inhibition (IC50) of [125I]amylin binding by human amylin was observed at 3.6 +/- 0.8 nM (n = 6). hCT and hCGRP-I displaced [125I]amylin binding with 22 and 66 times higher IC50. [125I]hCT binding was inhibited by hCT with an IC50 of 8.1 +/- 1.9 nM (n = 5), and human amylin and hCGRP-I were over 100 times less potent. In T47D cells, on the other hand, specific binding of [125I]amylin was not observed, but hCT inhibited [125I]hCT binding with an IC50 of 3.2 +/- 0.4 nM (n = 3), and human amylin and hCGRP-I had over 200 times higher IC50. In MCF-7 cells, half-maximal stimulation (EC50) of cyclic AMP accumulation by human amylin, hCT and hCGRP-I occurred at 1.4 +/- 0.2, 1.7 +/- 0.4 and 6.3 +/- 1.3 nM respectively. In T47D cells, the EC50 of hCT was 0.32 +/- 0.02 nM (n = 3), and 30- and 1900-fold higher with human amylin and hCGRP-I. In conclusion, the expression of hCTR1a and hCTR1b and [125I]hCT binding were indistinguishable in MCF-7 and T47D cells. Yet, [125I]amylin binding was only recognized in MCF-7 cells, consistent with a distinct amylin receptor.

Amyloid↗

PubMed1996

Bias in the gamma delta T cell response to Listeria monocytogenes. V delta 6.3+ cells are a major component of the gamma delta T cell response to Listeria monocytogenes.

Despite extensive research, the role that gamma delta T cells play in the immune response to infectious disease has yet to be established. Here we report the generation of a mAb specific for the V delta 6.3 TCR and investigate the gamma delta+ and V delta 6.3+ T cell responses to the intracellular bacterium Listeria monocytogenes in BALB/c mice. By infecting animals with various doses of Listeria and analyzing the components of the cellular immune response at the two primary sites of infection, the liver and spleen, we have shown that the kinetics, composition, and magnitude of the gamma delta and V delta 6.3 T cell responses are dependent upon the injected dose of bacteria and the organ in which the infection is established. At low doses of infection, the gamma delta T cell response occurs late in the disease course, while at high doses, the response is earlier and of greater magnitude, particularly in the liver. At all infectious doses and in both tissues, the V delta 6.3+ population predominates and together with V delta 4+ cells composes the bulk of the gamma delta T cell response. Changes in the morphology of gamma delta+ and V delta 6.3+ cells at the site of infection are consistent with cellular activation and suggest that these cells are active participants in the Listeria-induced immune response. The results of our study suggest that many features of the gamma delta T cell response to Listeria are dose and tissue related.

Animals↗

PubMed1996

Structure of a parathyroid hormone/parathyroid hormone-related peptide receptor of the human cerebellum and functional expression in human neuroblastoma SK-N-MC cells.

Cloning and functional expression of a cDNA from the human cerebellum revealed a parathyroid hormone/parathyroid hormone-related peptide (PTH/PTHrP) receptor protein of 593 amino acids, identical in sequence to the PTH/PTHrP receptor of the human kidney and an osteoblast-like cell line (Schipani et al., Endocrinology, 132 (1993) 2157-2165). Expression of mRNA hybridizing with the cloned cDNA, indistinguishable in size on Northern blots from a 2.3 kb transcript in kidney and liver, was detected in eight brain areas. In situ hybridization histochemistry in rat brain tissue sections revealed predominant signals in the Purkinje cell layer of the cerebellum and in the mesencephalic nucleus of the trigeminal nerve. In human neuroblastoma (SK-N-MC) cells, stably transfected with the cloned cDNA, hPTH(1-84) and hPTH(1-34) displaced binding of 125 pM [125I][Tyr36]chPTHrP(1-36) to the PTH/PTHrP receptor with IC50 values of 4.0 +/- 0.6 nM and 2.00 +/- 0.08 nM, and stimulated cyclic AMP accumulation with EC50 values of 0.19 +/- 0.06 nM and 0.09 +/- 0.01 nM, respectively. 16 out of 48 cells responded to 100 nM hPTH(1-34) with a 2-10-fold transient increase of cytosolic free calcium concentrations. In conclusion, a PTH/PTHrP receptor, identified in the human cerebellum, has the primary structure of the corresponding receptors of kidney and bone. Expression in human neuroblastoma SK-N-MC cells revealed functional properties indistinguishable from those of non-neuronal tissues. The widespread distribution of PTHrP and its receptor in brain implies biological functions remaining to be elucidated.

Amino Acid Sequence↗

PubMed1996

Cutaneous exposure to the superantigen staphylococcal enterotoxin B elicits a T-cell-dependent inflammatory response.

We analyzed the impact of superantigens secreted by skin-colonizing Staphylococci on the skin and the associated lymphoid tissue following epicutaneous application and intracutaneous injection of small amounts of staphylococcal enterotoxin B (SEB). A single intracutaneous injection of 50 ng of SEB elicited a strong inflammatory response in the skin of BALB/c mice. Three to 6 h later, we observed langerhans cell activation, mast cell degranulation, vasodilation, upregulation of ICAM-1, and induction of VCAM-1 on dermal blood vessels, with vascular adhesion of granulocytes. by 12 to 24 h, cell infiltration of the dermis increased, reaching the epidermis. Among the infiltrating leukocytes, a substantial number of eosinophils was found. After 48 h, the infiltrate was dominated by mononuclear cells. The response to SEB was dose-dependent, and signs of inflammation slowly disappeared over 5 to 7 days. Although the induction of VCAM-1 on dermal blood vessels suggested a role for interleukin-1/tumor necrosis factor-alpha in this reaction, the activation of monocytes/macrophages was not able to substitute for lymphocytes, as severe combined immunodeficiency (SCID) mice (which are lymphocyte-deficient) did not mount an inflammatory skin response to intradermal injection of SEB. The fact that nude mice (T-cell-deficient) also did not mount an inflammatory response to SEB indicated the T-cell dependency of the response. The V beta specificity of the SEB effect was demonstrated by the fact that SJL/J mice, which lack V beta 8+ T cells (the major SEB-reactive T cell population in mice), exhibited much weaker responses. Deletion or tolerization of SEB-reactive V beta T cells was not observed after a single intradermal injection of such minute amounts of SEB.

Animals↗

PubMed1995

Bacterial infection of the testis leading to autoaggressive immunity triggers apparently opposed responses of alpha beta and gamma delta T cells.

The mechanisms that lead to the breakdown of self-tolerance and testis-specific immune reactivity in the murine orchitis model are understood only in part. We investigated the histopathologic and immunologic consequences of a unilateral bacterial (Listeria monocytogenes) infection of the testis. Both infected and contralateral sides of this bilateral organ suffered severe inflammatory responses despite a conspicuous absence of bacteria in the contralateral tissue. Also, in both testicles, T cell populations increased, involving both alpha beta and gamma delta T cell subsets. Concomitant with the bilateral orchitis, testis-specific delayed type hypersensitivity and Ab responses developed. Ab depletion experiments indicated that in this orchitis model, as in others, alpha beta T cells are initiators of the autoaggressive reactivity. In contrast, Ab depletion of gamma delta T cells accelerated the inflammatory response in both testicles, suggesting a regulatory role for this type of T cells in both infection-induced and autoimmune orchitis.

Animals↗

PubMed1995

A human orphan calcitonin receptor-like structure.

A novel calcitonin receptor-like protein of 461 amino acids with seven putative transmembrane domains has been identified through molecular cloning in a cDNA library of the human cerebellum. 91% and 56% of the amino acids are identical in a rat orphan calcitonin receptor-like sequence and the human calcitonin receptor, respectively. 5.2 kb mRNA is predominantly expressed in the lung, heart and kidney. Specific binding of 125I-labeled salmon calcitonin and human calcitonin gene-related peptide-I to COS-7 cells transiently transfected with the receptor cDNA was less then 0.5%. Cellular cAMP accumulation was indistinguishable in cDNA transfected and non-transfected control COS-7 and renal tubular cells from the American opossum stimulated with human and salmon calcitonin, human calcitonin gene-related peptide-I and -II, human amylin, human adrenomedullin, lizard helodermin, salmon stanniocalcin and chicken parathyroid hormone-related protein. The receptor-like protein whose ligand remains to be discovered belongs to the family of receptors of calcitonin, parathyroid hormone, secretin, vasointestinal peptide and pituitary adenylate cyclase-activating polypeptide.

Amino Acid Sequence↗

PubMed1995

Receptors for calcitonin, calcitonin gene related peptide, amylin, and adrenomedullin.

Calcitonin, calcitonin gene related peptide, amylin, and adrenomedullin are structurally related polypeptides characterized by a six or seven amino acid ring structure linked by a disulfide bridge and an amidated C-terminus. They exhibit overlapping biological actions as a result of cross-reactivity between the different receptors. In this article, the respective receptors and G-protein-coupled postreceptor events are reviewed in relation to some of the biological actions of the peptides.

Animals↗

PubMed1995

Calcitonin, calcitonin gene-related peptide, adrenomedullin and amylin: homologous peptides, separate receptors and overlapping biological actions.

Calcitonin, calcitonin gene-related peptide, adrenomedullin and amylin are structurally related peptides with N-terminal 6-7 amino acid ring structures linked by a disulfide bridge and with amidated C-termini. Among the related bioactive peptides, the structures of the calcitonin receptor and subtypes thereof have been identified so far through molecular cloning. Cross-reaction between receptors of calcitonin, calcitonin gene-related peptide, adrenomedullin and amylin, as well as overlapping biological actions, anticipate that the respective receptors belong to a family of G-protein-coupled receptors that include those of parathyroid hormone, secretin and vasointestinal peptide.

Adrenomedullin↗

PubMed1994

Recognition of trophoblasts by gamma delta T cells.

The juxtaposition of maternal and fetal tissues in the hemochorial placenta has led to speculation that maternal recognition of fetal Ags present on trophoblasts might play an important role in reproductive biology. We report here for the first time such recognition of trophoblasts by T lymphocyte hybridomas representative of certain cells present in the maternal decidua. Trophoblast recognition is TCR dependent and is mediated by members of the V gamma 1+ subset of gamma delta T lymphocytes, a population that we previously have shown to be associated with heat shock protein-60 reactivity. Recognition occurred in experiments in which trophoblast clones or freshly prepared trophoblasts were used and requires cell-cell interaction. Although the maternal-fetal immune relationship typically has been cast in terms of an allograft, the T cell recognition of trophoblasts as described herein is not MHC-restricted, inasmuch as freshly prepared trophoblasts from beta 2-microglobulin-deficient mice were found to be stimulatory. Furthermore, the trophoblast ligand that mediates this recognition is probably a conserved mammalian molecule, because a human trophoblast cell line is also stimulatory. Our findings suggest a novel form of T cell recognition, and may provide an enhanced understanding of the maternal-fetal immune relationship.

Animals↗