Biomedical subjects

H Li

Publications and source records attributed to H Li.

At least 1,243 records · Page 69Linked to original sources

PubMed1997

Cloning and chromosomal location of a novel member of the myotonic dystrophy family of protein kinases.

We have cloned a novel serine/threonine protein kinase (PK428) which is highly related (65%) within the kinase domain to the myotonic dystrophy protein kinase (DM-PK), as well as the cyclic AMP-dependent protein kinase (33%). Northern blots demonstrate that PK428 mRNA is distributed widely among tissues and is expressed at the highest levels in pancreas, heart, and skeletal muscle, with lower levels in liver and lung. Two PK428 mRNAs 10 and 3.8 kilobase pairs in size are seen in a number of cell lines, including hematopoietic and breast cancer cells. An antibody generated to a glutathione S-transferase-PK428 fusion protein detects a 65-kDa protein in these cell lines, and a similarly sized protein when the cloned cDNA is transiently expressed in Cos 7 cells. Immunoprecipitation of the transiently expressed PK428 protein and incubation with [gamma-32P]ATP demonstrate that it is capable of autophosphorylation. In addition, immunoprecipitates of the PK428 protein kinase also phosphorylated histone H1 and a peptide encoding a cyclic AMP-dependent protein kinase substrate. The gene corresponding to the 3.8-kb PK428 mRNA, and its corresponding 65-kDa protein, was isolated by polymerase chain reaction screening of a P1 phage human genomic library. Using this P1 phage clone as a probe, the PK428 gene was located on 1q41-42, a possible location for a human senescence gene, a gene associated with Rippling muscle disease, as well as a region associated with genetically acquired mental retardation.

Amino Acid Sequence↗

PubMed1997

Molecular and functional characterization of two novel human C-C chemokines as inhibitors of two distinct classes of myeloid progenitors.

Two novel human beta-chemokines, Ck beta-8 or myeloid progenitor inhibitory factor 1 (MPIF-1), and Ck beta-6 or MPIF-2, were discovered as part of a large scale cDNA sequencing effort. The MPIF-1 and MPIF-2 cDNAs were isolated from aortic endothelium and activated monocyte libraries, respectively. Both of the cDNAs were cloned into a baculovirus vector and expressed in insect cells. The mature recombinant MPIF-1 protein consists of 99 amino acids and is most homologous to macrophage inflammatory protein (MIP)-1alpha, showing 51% identity. It displays chemotactic activity on resting T lymphocytes and monocytes, a minimal but significant activity on neutrophils, and is negative on activated T lymphocytes. MPIF-1 is also a potent suppressor of bone marrow low proliferative potential colony-forming cells, a committed progenitor that gives rise to granulocyte and monocyte lineages. The mature recombinant MPIF-2 has 93 amino acid residues and shows 39 and 42% identity with monocyte chemoattractant protein (MCP)-3 and MIP-1alpha, respectively. It displays chemotactic activity on resting T lymphocytes, a minimal activity on neutrophils, and is negative on monocytes and activated T lymphocytes. On eosinophils, MPIF-2 produces a transient rise of cytosolic Ca2+ and uses the receptor for eotaxin and MCP-4. In hematopoietic assays, MPIF-2 strongly suppressed the colony formation by the high proliferative potential colony-forming cell (HPP-CFC), which represents a multipotential hematopoietic progenitor.

Amino Acid Sequence↗

PubMed1997

Collateral projections from single neurons in the dorsal column nuclei to the inferior colliculus and the ventrobasal thalamus: a retrograde double-labeling study in the rat.

A number of single neurons in the dorsal column nuclei (DCN) and the sensory trigeminal nuclear complex were found to project to the thalamus and the inferior colliculus (IC). In the rats which were injected unilaterally with tetramethylrhodamine-dextran amine (TMR-DA) and Fluoro-Gold (FG) respectively into the posterolateral ventral thalamic nucleus and the external nucleus of IC, a number of neuronal cell bodies labeled retrogradely with both TMR-DA and FG were found throughout DCN contralateral to the injections. A few double-labeled neurons were also seen in the caudomedial part of the gracile nucleus ipsilateral to the injections. The double-labeled DCN neurons had relatively small cell bodies which are oval or fusiform in shape. When the injection sites in the thalamus extend into the posteromedial ventral thalamic nucleus, a few double-labeled neuronal cell bodies were seen contralaterally in the trigeminal sensory nuclear complex, mainly in the caudal part of the interpolar spinal trigeminal nucleus.

Animals↗

PubMed1997

Production of the novel C-C chemokine MCP-4 by airway cells and comparison of its biological activity to other C-C chemokines.

Monocyte chemotactic protein-4 (MCP-4) is a newly identified C-C chemokine with potent eosinophil chemoattractant properties. We describe studies of its biological activity in vitro to induce chemotaxis of peripheral blood eosinophils and to induce histamine release from IL-3-primed peripheral blood basophils. MCP-4 and eotaxin caused a similar rise in eosinophil intracytoplasmic Ca2+ and complete cross-desensitization. MCP-4 also abolished the eosinophil Ca2+ response to MCP-3 and partially desensitized the response to macrophage inflammatory protein-1alpha. MCP-4 activated cell migration via either CCR2b or CCR3 in mouse lymphoma cells transfected with these chemokine receptors. MCP-4 inhibited binding of 125I-eotaxin to eosinophils and CCR3-transfected cells and inhibited 125I-MCP-1 binding to CCR2b-transfectants. MCP-4 mRNA was found in cells collected in bronchoalveolar lavage of asthmatic and nonasthmatic subjects and was prominently expressed in human lung and heart. MCP-4 mRNA was expressed in several human bronchial epithelial cell lines after cytokine stimulation. Pretreatment of BEAS-2B epithelial cells with the glucocorticoid budesonide inhibited MCP-4 mRNA expression. These features make MCP-4 a candidate for playing a role in eosinophil recruitment during allergic respiratory diseases.

Amino Acid Sequence↗

PubMed1997

Patterns of hemopoietic reconstitution in nonobese diabetic mice: dichotomy of allogeneic resistance versus competitive advantage of disease-resistant marrow.

Complete replacement of the immune system via allogeneic bone marrow transplantation is sufficient to prevent diabetes in the nonobese diabetic (NOD) mouse model. In the present study we examined whether mixed allogeneic reconstitution would be sufficient to interrupt the autoimmune process with respect to occurrence of overt diabetes, as well as preexisting autoimmune insulitis. NOD mice were lethally irradiated and reconstituted with a mixture of NOD and B10.BR marrow. A relative resistance to allogeneic bone marrow engraftment was noted in NOD recipients of the mixed bone marrow inoculum, compared with disease-resistant controls. Moreover, unlike disease-resistant controls, all animals that initially repopulated as mixed donor/host chimeras became predominantly allogeneic by 4 mo, suggesting a competitive advantage for long term engraftment for disease-resistant marrow. All but one mouse in the group that engrafted with allogeneic marrow remained free of diabetes for the entire follow-up period (n = 22). Moreover, in all animals examined, virtually all islets were free of insulitis. In contrast, 74% of NOD mice that received similar conditioning and failed to engraft with donor marrow developed acute diabetes and intra-islet insulitis was present in all animals examined. These data suggest that NOD mice exhibit a relative resistance to engraftment compared with disease-resistant recipients. Conversely, animals that initially repopulated as a mixture of syngeneic and donor marrow become converted to virtually all donor by 4 mo. These data provide additional support that a defective stem cell is responsible for autoimmune diabetes in this experimental model.

Animals↗

PubMed1997

Localization of a metabotropic glutamate receptor, mGluR7, in axon terminals of presumed nociceptive, primary afferent fibers in the superficial layers of the spinal dorsal horn: an electron microscope study in the rat.

It was examined electron microscopically in the rat if a metabotropic glutamate receptor, mGluR7, might be localized in axon terminals of nociceptive, primary afferent fibers in laminae I and II of the spinal dorsal horn. Nociceptive nature of axon terminals showing mGluR7-like immunoreactivity (mGluR7-LI) was indicated by binding to the isolectin I-B4 from Griffonia simplicifolia (I-B4), or by substance P-like immunoreactivity (SP-LI). Axon terminals labeled with immunogold particles indicating mGluR7-LI were usually filled with round synaptic vesicles and were in asymmetric synaptic contact with dendritic or somatic profiles; occasionally they contained pleomorphic vesicles and were in symmetric synaptic contact with somatic profiles in lamina II. The double-labeling studies revealed that most of axon terminals with I-B4 labeling as well as a small population of axon terminals with SP-LI, showed mGluR7-LI. About one-third or much smaller population of axon terminals with mGluR7-LI in laminae I and II were labeled, respectively, with I-B4 or SP-LI; these were in asymmetric synaptic contact with dendritic profiles.

Animals↗

PubMed1997

PL48: a novel gene associated with cytotrophoblast and lineage-specific HL-60 cell differentiation.

The cDNA for a novel gene, PL48, isolated by subtractive hybridization between undifferentiated human term cytotrophoblast and differentiating cytotrophoblast, has been cloned and sequenced. PL48 contains an open reading frame coding for a 537-amino acid protein, has multiple potential PKC, casein kinase II, and cAMP/cGMP-dependent kinase phosphorylation sites, and N-linked glycosylation sites. It is not present in a wide variety of proliferating cancer cells, but PL48 mRNA shows marked expression during cytotrophoblast and granulocyte lineage-specific HL-60 promyelocytic cell differentiation induced by DMSO.

Amino Acid Sequence↗

PubMed1997

Collateral projections from single neurons in the dorsal column nucleus to both the cochlear nucleus and the ventrobasal thalamus: a retrograde double-labeling study in the rat.

In the rats which were injected with Fluoro-Gold into the cochlear nucleus and simultaneously with tetramethylrhodamine-dextran amine into the ventrobasal thalamus, a number of neurons in the gracile and the cuneate nuclei of the dorsal column nucleus (DCN) and in the interpolar and caudal subnuclei of the spinal trigeminal nucleus (STN) were double-labeled retrogradely with the both tracers. The results indicated the existence of single DCN/STN-neurons which provided axons to the cochlear nucleus, mainly ipsilaterally, and to the contralateral ventrobasal thalamus.

Animals↗

PubMed1997

Binding of the isolectin I-B4 from Griffonia simplicifolia to the general visceral afferents in the vagus nerve: a light- and electron-microscope study in the rat.

Prominent binding to the isolectin I-B4 from Griffonia simplicifolia (I-B4) was observed not only in the terminal area of general somatic afferents (lamina II of the medullary and the spinal dorsal horns), but also in the terminal area of the general visceral afferents (nucleus of the solitary tract, NST). The vast majority of neuronal cell bodies in the nodose ganglion (NG), including those with substance P-like immunoreactivity, also showed the I-B4-binding. After ganglionectomy of the NG, the I-B4-binding in the solitary tract and NST was highly reduced throughout the solitary tract and the NST on the side ipsilateral to the operation. The I-B4-binding was electron microscopically observed in many axon terminals within the NST.

Animals↗

PubMed1997

Purification, visualization, and biophysical characterization of Kv1.3 tetramers.

The voltage-gated K+ channel of T-lymphocytes, Kv1.3, was heterologously expressed in African Green Monkey kidney cells (CV-1) using a vaccinia virus/T7 hybrid expression system; each infected cell exhibited 10(4) to 5 x 10(5) functional channels on the cell surface. The protein, solubilized with detergent (3-[cholamidopropyl)dimethylammonio]-1-propanesulfonic acid or cholate), was purified to near-homogeneity by a single nickel-chelate chromatography step. The Kv1.3 protein expressed in vaccinia virus-infected cells and its purified counterpart are both modified by a approximately 2-kDa core-sugar moiety, most likely at a conserved N-glycosylation site in the external S1-S2 loop; absence of the sugar does not alter the biophysical properties of the channel nor does it affect expression levels. Purified Kv1.3 has an estimated size of approximately 64 kDa in denaturing SDS-polyacrylamide electrophoresis gels, consistent with its predicted size based on the amino acid sequence. By sucrose gradient sedimentation, purified Kv1.3 is seen primarily as a single peak with an approximate mass of 270 kDa, compatible with its being a homotetrameric complex of the approximately 64-kDa subunits. When reconstituted in the presence of lipid and visualized by negative-staining electron microscopy, the purified Kv1.3 protein forms small crystalline domains consisting of tetramers with dimensions of approximately 65 x 65 A. The center of each tetramer contains a stained depression which may represent the ion conduction pathway. Functional reconstitution of the Kv1.3 protein into lipid bilayers produces voltage-dependent K+-selective currents that can be blocked by two high affinity peptide antagonists of Kv1.3, margatoxin and stichodactylatoxin.

Animals↗

PubMed1997

Effects of delayed re-innervation on the expression of c-erbB receptors by chronically denervated rat Schwann cells in vivo.

We propose that chronically denervated Schwann cells may be less able to respond to axonal signals than their acutely denervated counterparts, and that this lack of sensitivity may be one reason why axons fail to regenerate into chronically denervated nerve stumps. To test this proposal we have used in situ hybridization, and quantitative and qualitative immunohistochemistry to compare the expression of c-erbB2 and c-erbB4 receptors in Schwann cells denervated for up to 6 months in vivo, with that seen in Schwann cells denervated for similar periods of time but then exposed to regenerating axons. The results were correlated with the extent of axonal regeneration in each experimental group as assessed from transverse sections which had been double-immunolabelled using anti S-100 and anti-beta tubulin III antibodies. Since c-erbBs are receptors for neuronally derived neuregulins we probed the appropriate axotomised DRG neurons for expression of GGF2 mRNA. When the denervated distal stumps were anastomosed to acutely transected proximal stumps, GGF expression in DRGs increased transiently during the first week: we assume that secreted GGF2 derived from regrowing axon sprouts would have been available to Schwann cells in all distal stumps. Endoneurial cell proliferation (predominantly Schwann cell proliferation); levels of expression of c-erbB receptors by Schwann cells, and the degree to which axons regenerated into the distal stumps all decreased as the period of prior denervation increased: the longer the time of denervation, the lower the expression of c-erbBs in Schwann cells, and the smaller the percentage of bands of Bungner which were re-innervated.

Animals↗

PubMed1997

Novel regions of chromosomal loss in familial neuroblastoma by comparative genomic hybridization.

Childhood neuroblastoma, an embryonal neoplasm of sympathetic nervous system progenitors, occurs in a familial form with an autosomal dominant mode of inheritance. Genetic susceptibility to this disorder is thought to arise via a germline mutation affecting a tumor suppressor gene, in accord with the two-hit model established for familial and sporadic retinoblastoma. Surprisingly, the familial neuroblastoma predisposition locus does not map to chromosome band 1p36, a genomic region likely to contain one or more neuroblastoma suppressor genes. We reasoned that inherited point mutations affecting one allele would be unmasked in many cases by somatically acquired deletions of the second allele that included the target gene in the tumor cells from these patients. Thus, to identify chromosomal regions that might contain suppressor genes important in hereditary neuroblastoma, we analyzed six familial tumors by comparative genomic hybridization. Recurrent losses of genetic material were detected on chromosome arms 3p (consensus region, 3p24-pter), 10p (consensus, 10p12-p13), 10q (consensus, 10q25-qter), 16q (consensus, 16q12-q22), and 20q (consensus, 20q13.3-qter), in addition to the regions commonly deleted in sporadic neuroblastomas (1p36 and 11q). These chromosomal sites may harbor novel tumor suppressor genes that could aid in our understanding of the predisposition to and pathogenesis of familial neuroblastoma and potentially sporadic tumors as well.

Child, Preschool↗

PubMed1997

Identifying genetic markers to assess the presence of gene-environment interactions.

We analyzed a randomly chosen replicate with the goals of locating the closest markers to the genes involved in the discrete trait and utilizing these as surrogates for the genes in assessing the presence of gene-environment interactions. We screened the markers with an association test prior to using the transmission-disequilibrium test. We performed a segregation analysis, with regressive models and including the selected markers, to understand the underlying genetic mechanism and the role of the environmental factor. We were unsuccessful in locating the relevant markers due to the absence of linkage disequilibrium. Nevertheless, some insights were gained from the methods used.

Chromosome Mapping↗

PubMed1997

Genotype relative-risks and association tests for nuclear families with missing parental data.

The development of a new method for testing the association of genetic markers with disease is presented. This approach is applicable when sampling nuclear families with one or more affected siblings and when neither, one, or both parents are missing marker genotype data. All siblings, affected and not affected, are used to probabilistically infer the missing parental marker data. A likelihood ratio statistic, which treats marker allele frequencies as nuisance parameters, is presented to test whether all marker relative risks are equal to one (i.e., no marker association). This approach offers a solution to test for marker associations when parents are difficult to obtain.

Algorithms↗

PubMed1997

GENEHUNTER: application to analysis of bipolar pedigrees and some extensions.

A newly developed nonparametric multipoint linkage analysis (NPL) by Kruglyak et al. [1996] was applied to the data set from the Johns Hopkins Bipolar Study available through the GAW10 workshop. Our results from GENEHUNTER do not support the analyses of Stine et al. [1995]. Some possible extensions to incorporate unaffecteds are also discussed.

Bipolar Disorder↗

PubMed1997

Tissue inhibitor of metalloproteinase-3 expression is upregulated during human cytotrophoblast invasion in vitro.

Surprisingly, a successful human pregnancy requires cells from the fetal portion of the placenta (cytotrophoblasts) to adopt tumor-like properties. Cytotrophoblasts attach the conceptus to the endometrium by invading the uterus, and they initiate blood flow to the placenta by breaching maternal vessels. But unlike tumor metastasis, cytotrophoblast invasion is highly regulated both spatially and temporally. Our previous work showed that matrix metalloproteinase-9 (MMP-9) expression is upregulated during cytotrophoblast differentiation along the invasive pathway, and that activity of this proteinase specifies the cells' ability to degrade extracellular matrix (ECM) substrates in vitro. Here we tested the hypothesis that invading cytotrophoblasts express an unusual tissue inhibitor of metalloproteinase (TIMP) repertoire that allows them to regulate their MMP-9 proteolytic activity. By using protease-substrate gel electrophoresis, we found that human cytotrophoblasts express primarily TIMP-3. We showed that the cells' TIMP-3 expression is regulated in accord with that of MMP-9. The highest levels of protein and mRNA for both these molecules were detected after differentiation to a fully invasive phenotype and during early gestation, when invasion peaks, rather than at term, when invasion has stopped. Our results suggest that coexpression of MMP-9 and TIMP-3 by invading cytotrophoblasts plays an important role in regulating the depth of uterine invasion.

Cell Movement↗

PubMed1997

Homology modeling using simulated annealing of restrained molecular dynamics and conformational search calculations with CONGEN: application in predicting the three-dimensional structure of murine homeodomain Msx-1.

We have developed an automatic approach for homology modeling using restrained molecular dynamics and simulated annealing procedures, together with conformational search algorithms available in the molecular mechanics program CONGEN (Bruccoleri RE, Karplus M, 1987, Biopolymers 26:137-168). The accuracy of the method is validated by "predicting" structures of two homeodomain proteins with known three-dimensional structures, and then applied to predict the three-dimensional structure of the homeodomain of the murine Msx-1 transcription factor. Regions of the unknown protein structure that are highly homologous to the known template structure are constrained by "homology distance constraints," whereas the conformations of nonhomologous regions of the unknown protein are defined only by the potential energy function. A full energy function (excluding explicit solvent) is employed to ensure that the calculated structures have good conformational energies and are physically reasonable. As in NMR structure determinations, information on the consistency of the structure prediction is obtained by superposition of the resulting family of protein structures. In this paper, our homology modeling algorithm is described and compared with related homology modeling methods using spatial constraints derived from the structures of homologous proteins. The software is then used to predict the DNA-bound structures of three homeodomain proteins from the X-ray crystal structure of the engrailed homeodomain protein (Kissinger CR et al., 1990, Cell 63:579-590). The resulting backbone and side-chain conformations of the modeled yeast Mat alpha 2 and D. melanogaster Antennapedia homeodomains are excellent matches to the corresponding published X-ray crystal (Wolberger C et al., 1991, Cell 67:517-528) and NMR (Billeter M et al., 1993, J Mol Biol 234:1084-1097) structures, respectively. Examination of these structures of Msx-1 reveals a network of highly conserved surface salt bridges that are proposed to play a role in regulating protein-protein interactions of homeodomains in transcription complexes.

Algorithms↗

PubMed1997

Ouabain-induced hypertrophy in cultured cardiac myocytes is accompanied by changes in expression of several late response genes.

Partial inhibition of cardiac Na/K-ATPase by digitalis drugs such as ouabain is the initial event leading to positive inotropy in the heart. We showed recently that exposure of rat cardiac myocytes to ouabain concentrations that produce positive inotropy, but no overt toxicity, caused inductions of some early response genes and hypertrophy of these myocytes. The aim of this work was to determine if ouabain also affects the expressions of certain late response genes that are regulated by other hypertrophic stimuli. Non-toxic concentrations of ouabain (5-100 microM) increased mRNAs of skeletal alpha-actin, atrial natriuretic factor, myosin light chain 2, and transforming growth factor beta: indicating that ouabain's effects on these marker genes are similar to those of hypertrophic stimuli that mimic the effects of pressure overload. Expression of skeletal alpha-actin was more sensitive to ouabain than that of atrial natriuretic factor, suggesting significant differences in the ouabain-specific pathways of the induction of these fetal genes. The effects of ouabain on skeletal alpha-actin gene were transcriptional, and required an increase in net influx of extracellular Ca2+. Protein kinase C and Ca(2+)-calmodulin kinases, but not protein kinase A, were involved in the signal pathways leading to the induction of skeletal alpha-actin gene. These data and our prior findings indicate that an increase in net influx of Ca2+ through partial inhibition of Na/K-ATPase initiates protein kinase-dependent pathways resulting in alterations in cardiac growth and expressions of both early and late response genes.

Actins↗