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Biomedical subjects

T Zhang

Publications and source records attributed to T Zhang.

At least 127 records · Page 7Linked to original sources
PubMed2000

Huntingtin is required for normal hematopoiesis.

Huntington's disease (HD) is a neurodegenerative disease associated with polyglutamine expansion in huntingtin, a widely expressed protein. The function of huntingtin is unknown although huntingtin plays a fundamental role in development since gene targeted HD (-) (/-)mouse embryos die shortly after gastrulation. Expression of huntingtin is detected in spleen and thymus but its role in hematopoiesis has not been examined. To determine the function of huntingtin and to provide insight into potential pathologic mechanisms in HD, we analyzed the role of huntingtin in hematopoietic development. Expression of huntingtin was analyzed in a variety of hematopoietic cell types, and in vitro hematopoiesis was assessed using an HD ( +/-)and several HD( -) (/-)embryonic stem (ES) cell lines. Although wild-type, HD ( +/-)and HD( -) (/-)ES cell lines formed primary embryoid bodies (EBs) with similar efficiency, the numbers of hematopoietic progenitors detected at various stages of the in vitro differentiation were reduced in HD ( +/-)and HD( -/-)() ()ES cell lines examined. Expression analyses of hematopoietic markers within the EBs revealed that primitive and definitive hematopoiesis occurs in the absence of huntingtin. However, further analysis using a suspension culture in the presence of hematopoietic cytokines demonstrated a highly significant gene dosage-dependent decrease in proliferation and/or survival of HD ( +/-)and HD( -) (/-)cells. Enrichment for the CD34(+)cells within the EB confirmed that the impairment is intrinsic to the hematopoietic cells. These obser- vations suggest that huntingtin expression is required for the generation and expansion of hematopoietic cells and provides an alternative system in which to assess the function of huntingtin.

Animals↗
PubMed2000

Helicobacter pylori alters gastric epithelial cell cycle events and gastrin secretion in Mongolian gerbils.

BACKGROUND & AIMS: Human colonization with Helicobacter pylori increases the risk for distal gastric adenocarcinoma, possibly by altering gastric epithelial cell cycle events and/or gastrin secretion. This study aimed to determine whether H. pylori virulence-related characteristics affect apoptosis, proliferation, and gastrin levels in a rodent model of gastric adenocarcinoma. METHODS: Mongolian gerbils were challenged with H. pylori wild-type or isogenic cagA(-) and vacA(-) mutants, and apoptotic and proliferating cells were identified by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling and proliferating cell nuclear antigen immunohistochemistry, respectively. Serum gastrin levels were determined by radioimmunoassay. RESULTS: Gastric epithelial cell turnover was no different after infection with the wild-type, cagA(-), or vacA(-) strains. H. pylori infection significantly increased antral apoptosis 2-4 weeks after challenge, before apoptotic indices decreased to baseline. In contrast, antral proliferation rates were significantly higher 16-20 weeks after inoculation, but then decreased by 40 weeks. Antral proliferation was significantly related to serum gastrin levels, whereas antral apoptosis was inversely related to acute inflammation and lymphoid follicles. CONCLUSIONS: In H. pylori-infected gerbils, enhanced antral apoptosis is an early and transient cell cycle event. Epithelial cell proliferation peaks later and is significantly related to increased gastrin levels, suggesting that epithelial cell growth in H. pylori-colonized mucosa may be mediated by gastrin-dependent mechanisms.

Animals↗
PubMed2000

Expression of membrane-type 1 matrix metalloproteinase in coronary vessels of allotransplanted primate hearts.

BACKGROUND: The mechanisms of intimal thickening in cardiac allograft vasculopathy (CAV) remain controversial after heart transplantation. Matrix metalloproteinase-2 (MMP-2) plays a crucial role in degrading extracellular matrix (ECM) during neointimal formation. Recently, it has been revealed that MMP-2 is activated by membrane-type 1 matrix metalloproteinase (MT1-MMP). This process involves tissue inhibitor of MMP-2 (TIMP-2), forming an MT1-MMP/TIMP-2/pro-MMP-2 complex. In this study, we hypothesize that these components contribute to the pathogenesis of CAV. METHODS: Heterotopic cardiac allografting was performed in randomly paired Japanese monkeys with an immunosuppressive regimen of intravenous administration of antihuman CD18 monoclonal antibody. The donor hearts were harvested at Days 22, 28, 40, 41, and 95 posttransplantation. We examined expression of MMP-2, MT1-MMP, and TIMP-2 of graft vessels using immunohistochemistry and protein level by western blot analysis. RESULTS: Pathologically, various degrees of neointimal formation were observed. In the allografts harvested at Days 22, 28, 40, and 41, MT1-MMP was expressed in the endothelial cells and smooth muscle cells (SMCs) in media of some arteries without histological change, accompanied by expression of MMP-2 and TIMP-2. In the severely thickened neointima of the allograft harvested at Day 95, MMP-2 and faint MT1-MMP were expressed in SMCs of severely thickened neointima and media; TIMP-2 expression was seen only in noncollagenous tissue of severely thickened neointima. MMP-2 protein was more intensely expressed in the allograft harvested at Day 95 than in the allograft harvest at Day 41, while TIMP-2 protein level was almost same in the 2 samples. CONCLUSION: We observed the simultaneous expression of MMP-2, MT1-MMP, and TIMP-2. Thus, ECM degradation triggered by MT1-MMP/TIMP-2/pro-MMP-2 complex could be a novel mechanism of CAV.

Animals↗
PubMed2000

Different cytokines are required for induction and maintenance of the Th2 type response in DBA/2 mice resistant to infection with Leishmania major.

Experimental cutaneous leishmaniasis is a useful model in studying the mechanism regulating immune responses between T helper type 1 (Th1) and Th2. Mice susceptible to Leishmania major infection such as BALB/c (H-2(d)) are associated with the induction of the disease-promoting Th2 response, while the resistant mice such as DBA/2 (H-2(d)) develop the protective Th1 response. To understand the induction mechanism of Th1 and Th2 responses, it is necessary to establish an immunization scheme by which the induction of each Th response can be easily and experimentally controlled. Adjuvants are known to enhance the immune responses through the combined effect of several factors: prolonged release of antigen, migration of cells, mitogenic effect and so forth. When the genetically resistant DBA/2 mice were immunized twice with soluble leishmanial antigen (SLA), emulsified in incomplete Freund's adjuvant (IFA) before L. major inoculation, these mice mounted a Th2 cell response and suffered from progressive infection. While IL-4 and IL-13 were upregulated early after the infection in both healer and non-healer groups of mice, IL-5 and IL-10 were upregulated only in non-healer mice. From these results, IL-5 and IL-10 appear to have an important role, at least in the early phases of the infection, rather than IL-4 and IL-13 in establishing the disease-promoting Th2 response in leishmaniasis. Further, IL-9 was found to be expressed in both BALB/c and DBA/2 mice immunized with IFA/SLA. This cytokine may support the establishment of a Th2 response in these mice. Therefore it is suggested that Th2 cytokines play different roles between priming and maintaining the Th2 immune response after the infection.

Animals↗
PubMed2000

Unusual inheritance of the AxinFu mutation in mice is associated with widespread rearrangements in the proximal region of chromosome 17.

AxinFu is a mutation in mice that causes fused tails and other developmental abnormalities as a result of insertion of an intracisternal-A particle (IAP), a murine retrotransposon, into intron 6. In a small percentage of offspring we found that the mutant allele reverts to wild-type through loss of the insertion with concomitant disappearance of the mutant phenotype. Investigation of a series of microsatellite loci in the proximal region of chromosome 17 revealed novel alleles which arise simultaneously with disappearance of IAP from AxinFu. These novel microsatellite variants are distinct from the parental alleles and those so far discovered are organized into two haplotypes. Both haplotypes demonstrate stable Mendelian inheritance. Results show that these rearrangements, which are involved in the production of the new haplotypes, exceed millions of base pairs.

Animals↗
PubMed2000

Pathologic, cytogenetic and molecular assessment of acute promyelocytic leukemia patients treated with arsenic trioxide (As2O3).

Arsenic trioxide (As2O3) shows great promise as an effective therapy for patients with all-trans retinoic acid (ATRA)-resistant acute promyelocytic leukemia (APL). Little data is available addressing the pathology of As2O3 treated APL and whether the antileukemic mechanism of As2O3 is primarily cytolysis or through stimulation of cell differentiation. In this report, we made a morphologic, cytogenetic, and molecular evaluation of five ATRA-refractory APL patients who were treated with As2O3. Four of the five patients had morphologic responses after one or two cycles of As2O3 treatment. Of the four responders based on bone marrow morphology, two achieved molecular remission (negative RT-PCR for PML- RAR alpha fusion transcripts) by the end of the second and third cycles of As2O3 therapy. Two patients exhibited marked leukocytosis during the first cycle of As2O3, and at that time point the APL cells were largely replaced by the cells showing partial differentiation towards myelocytes with co-expression of CD11b and CD33. Nevertheless, these "myelocyte-like" cells that showed the t(15;17) translocation eventually disappeared with continuous As2O3 therapy. As2O3 treatment appears to be effective therapy for the patients with relapsed APL after the failure of conventional chemotherapy and ATRA therapy. The pathologic findings in these five cases suggest that at low doses As2O3 primarily induces differentiation of the APL cells, generating abnormal myelocytes resembling APL cells treated with ATRA, whereas at higher doses AS2O3 induces marrow necrosis.

Adult↗
PubMed2000

Lysosomal cathepsin B plays an important role in antigen processing, while cathepsin D is involved in degradation of the invariant chain inovalbumin-immunized mice.

We previously reported that CA074, a specific inhibitor of cathepsin B, modulates specific immune responses from the T helper 2 (Th2) type to Th1 type in BALB/c mice infected with Leishmania major. In the present study, we found that a similar type of immune deviation was also induced in mice immunized with ovalbumin (OVA). However, treatment of mice with pepstatin A, a specific cathepsin D inhibitor, suppressed the OVA-specific proliferation of lymphocytes and blocked the development of both Th1 and Th2 cellular responses. These inhibitors did not appear to have any direct influence in vitro on functions of naive lymphocytes. OVA antigen (47 000 MW) was digested mainly into 40 000 MW protein in vitro by lysosomal proteases from naive BALB/c mice, and its digestion was markedly inhibited by the addition of CA074, but not by addition of pepstatin A, during incubation. However, pepstatin A strongly suppressed the degradation of the major histocompatibility complex class II-associated invariant chain (Ii) molecule in vivo and in vitro. Thus, cathepsin B appears to process antigens directed to preferential activation of Th2 cells, while cathepsin D may be responsible for the degradation of Ii, the processing of which is essential in initiating the antigen-specific activation of Th1 and Th2 CD4+ T cells. These lysosomal proteases may have different functions in regulating immune responses.

Animals↗
PubMed2000

Biochemical characterization of S100A2 in human keratinocytes: subcellular localization, dimerization, and oxidative cross-linking.

S100A2 is a calmodulin-like protein of unknown function, whose transcription is positively regulated in response to ErbB and p53 signaling. Expression of S100A2 is markedly increased in the context of ErbB-driven reactive epidermal hyperplasia, and decreased in the context of hypofunctional p53 mutations in carcinoma cell lines and tumors. This bimodal pattern of regulation suggests an important function for S100A2 in keratinocyte differentiation and carcinogenesis. Taking the biochemical approach to the determination of S100A2 function, we have characterized its physical state and subcellular localization in normal human keratinocytes. S100A2 in hypotonic lysates remained soluble after centrifugation at 100 000 x g, indicating that it is not associated with cell membranes. Permeabilization experiments confirmed the lack of membrane association and revealed a digitonin-insoluble nuclear fraction of S100A2, which was confirmed by immunofluorescence microscopy. Pulldown assays of epitope-tagged S100A2 and yeast two-hybrid screening revealed that S100A2 displays a strong propensity to homodimerize. Naturally expressed S100A2 dimers in normal human keratinocytes readily underwent intermolecular disulfide cross-linking unless a strong denaturant was present during cell lysis. Treatment of intact normal human keratinocytes with hydrogen peroxide strongly promoted S100A2 cross-linking. These results demonstrate that native S100A2 is a homodimer that does not depend on disulfide cross-linking for stability, but undergoes intermolecular cross-linking at cysteine residues in response to oxidative stress. Based on these findings, we propose that S100A2 may protect normal keratinocytes against carcinogens by participating in the cellular proof-reading response to oxidative stress.

Antibodies, Monoclonal↗
PubMed2000

CD4(+) v(alpha)14 NKT cells play a crucial role in an early stage of protective immunity against infection with Leishmania major.

The roles of gamma delta T, NK and NKT cells in an early stage of protective immunity against infection with Leishmania major were investigated. Further, the contribution of these innate cells to the expression of 65 kDa heat shock protein (HSP65) in host macrophages was examined, since we found previously that this expression prevents apoptotic death of infected macrophages and is a crucial step in the acquisition of protective immunity against infection with various obligate intracellular protozoa including L. major. C57BL/6 and DBA/2 mice were found to be resistant against the infection on the basis of the parasite burden in their regional lymph nodes, and to strongly express HSP65 in their macrophages, whereas BALB/c mice were susceptible and barely expressed the HSP65. In those resistant mice, CD4(+) NKT cells prominently increased in their regional lymph node and were the main effector cells at least for an early stage of the protective immunity and for the HSP65 expression, whereas this subset did not increase in susceptible BALB/c mice. Further, neither gamma delta T nor NK cells in resistant mice contributed to those protective immune responses. The NKT cell subset bore CD3, CD4, TCR alpha beta, IL-2R beta and NK1.1 but scarcely asialo-GM(1). Moreover, this effector subset was confirmed to be V(alpha)14 NKT cells by using J(alpha)281(-/-) mice.

Animals↗
PubMed2000

Two N-ortho-nitrobenzenesulfonyl alpha,alpha-disubstituted amino acid adducts.

The carboxy group of 2-methyl-N-[(2-nitrophenyl)sulfonyl]alanine, C(10)H(12)N(2)O(6)S, forms centrosymmetric hydrogen-bonded dimers with an O.O distance of 2.629 (2) A and an intramolecular N-H. O(nitro) hydrogen bond N.O distance of 2.823 (2) A. 1-[(2-Nitrophenyl)sulfonylamino]cyclohexanecarboxylic acid, C(13)H(16)N(2)O(6)S, has Z' = 2 and forms similar interactions.

Alanine↗
PubMed2000

Involvement of endogenously synthesized interleukin (IL)-18 in the protective effects of IL-12 against pulmonary infection with Cryptococcus neoformans in mice.

We previously demonstrated that interleukin (IL)-12 protected mice against fatal pulmonary infection with a highly virulent strain of Cryptococcus neoformans, which correlated well with the production of interferon (IFN)-gamma as well as IL-18 in the primary infected site. In the present study, we examined the role of endogenously synthesized IL-18 in IL-12-induced host resistance to this pathogen. There was little or no production of IFN-gamma and IL-18 both at mRNA and protein levels in lungs of mice infected with C. neoformans, while treatment with IL-12 induced a marked production of these cytokines. Caspase-1 mRNA was expressed in infected mice even without IL-12 treatment. Administration of neutralizing anti-IFN-gamma monoclonal antibody (mAb) clearly inhibited production of IFN-gamma and IL-18 induced by IL-12, while control IgG did not show such an effect. However, administration of IFN-gamma did not induce the production of both cytokines in infected mice, although tumor necrosis factor (TNF)-alpha and IFN-gamma-inducible protein (IP)-10 were synthesized by the same treatment. Finally, neutralizing anti-IL-18 antibody (Ab) significantly interfered with the production of IFN-gamma and elimination of the microorganism from the lung induced by IL-12 treatment. Furthermore, both IFN-gamma synthesis and host protection caused by IL-12 were profoundly diminished in IL-18 gene-disrupted mice. Considered collectively, our results indicated that host protection against C. neoformans induced by IL-12 involved endogenously synthesized IL-18 and that the production of IL-18 was mediated at least in part by endogenous IFN-gamma.

Animals↗
PubMed2000

Role of the cyclic AMP response element binding complex and activation of mitogen-activated protein kinases in synergistic activation of the glycoprotein hormone alpha subunit gene by epidermal growth factor and forskolin.

The aim of these studies was to elucidate a role for epidermal growth factor (EGF) signaling in the transcriptional regulation of the glycoprotein hormone alpha subunit gene, a subunit of chorionic gonadotropin. Studies examined the effects of EGF and the adenylate cyclase activator forskolin on the expression of a transfected alpha subunit reporter gene in a human choriocarcinoma cell line (JEG3). At maximal doses, administration of EGF resulted in a 50% increase in a subunit reporter activity; forskolin administration induced a fivefold activation; the combined actions of EGF and forskolin resulted in synergistic activation (greater than eightfold) of the alpha subunit reporter. Mutagenesis studies revealed that the cyclic AMP response elements (CRE) were required and sufficient to mediate EGF-forskolin-induced synergistic activation. The combined actions of EGF and forskolin resulted in potentiated activation of extracellular signal-regulated kinase (ERK) enzyme activity compared with EGF alone. Specific blockade of ERK activation was sufficient to block EGF-forskolin-induced synergistic activation of the alpha subunit reporter. Pretreatment of JEG3 cells with a p38 mitogen-activated protein kinase inhibitor did not influence activation of the alpha reporter. However, overexpression of c-Jun N-terminal kinase (JNK)-interacting protein 1 as a dominant interfering molecule abolished the synergistic effects of EGF and forskolin on the alpha subunit reporter. CRE binding studies suggested that the CRE complex consisted of CRE binding protein and EGF-ERK-dependent recruitment of c-Jun-c-Fos (AP-1) to the CRE. A dominant negative form of c-Fos (A-Fos) that specifically disrupts c-Jun-c-Fos DNA binding inhibited synergistic activation of the alpha subunit. Thus, synergistic activation of the alpha subunit gene induced by EGF-forskolin requires the ERK and JNK cascades and the recruitment of AP-1 to the CRE binding complex.

Choriocarcinoma↗
PubMed2000

The coiled-coil domain of Stat3 is essential for its SH2 domain-mediated receptor binding and subsequent activation induced by epidermal growth factor and interleukin-6.

STAT proteins are a family of latent transcription factors that mediate the response to various cytokines and growth factors. Upon stimulation by cytokines, STAT proteins are recruited to the receptors via their SH2 domains, phosphorylated on a specific tyrosine, dimerized, and translocated into the nucleus, where they bind specific DNA sequences and activate the target gene transcription. STATs share highly conserved structures, including an N-domain, a coiled-coil domain, a DNA-binding domain, a linker domain, and an SH2 domain. To investigate the role of the coiled-coil domain, we performed a systematic deletion analysis of the N-domain and each of the alpha-helices and mutagenesis of conserved residues in the coiled-coil region of Stat3. Our results indicate that the coiled-coil domain is essential for Stat3 recruitment to the receptor and the subsequent tyrosine phosphorylation and tyrosine phosphorylation-dependent activities, such as dimer formation, nuclear translocation, and DNA binding, stimulated by epidermal growth factor (EGF) or interleukin-6 (IL-6). Single mutation of Asp170 or, to a lesser extent, Lys177 in alpha-helix 1 diminishes both receptor binding and tyrosine phosphorylation. Furthermore, the Asp170 mutant retains its ability to bind to DNA when phosphorylated on Tyr705 by Src kinase in vitro, implying a functional SH2 domain. Finally, we demonstrate a direct binding of Stat3 to the receptor. Taken together, our data reveal a novel role for the coiled-coil domain that regulates the early events in Stat3 activation and function.

Animals↗