Search PubMed⌕ Search

Biomedical subjects

M Wang

Publications and source records attributed to M Wang.

At least 145 records · Page 8Linked to original sources
PubMed2001

The small ubiquitin-like modifier-1 (SUMO-1) consensus sequence mediates Ubc9 binding and is essential for SUMO-1 modification.

SUMO-1 is an ubiquitin-related protein that is covalently conjugated to a diverse assortment of proteins. The consequences of SUMO-1 modification include the regulation of protein-protein interactions, protein-DNA interactions, and protein subcellular localization. At present, very little is understood about the specific mechanisms that govern the recognition of proteins as substrates for SUMO-1 modification. However, many of the proteins that are modified by SUMO-1 interact directly with the SUMO-1 conjugating enzyme, Ubc9. These interactions suggest that Ubc9 binding may play an important role in substrate recognition as well as in substrate modification. The SUMO-1 consensus sequence (SUMO-1-CS) is a motif of conserved residues surrounding the modified lysine residue of most SUMO-1 substrates. This motif conforms to the sequence "PsiKXE," where Psi is a large hydrophobic residue, K is the lysine to which SUMO-1 is conjugated, X is any amino acid, and E is glutamic acid. In this study, we demonstrate that the SUMO-1-CS is a major determinant of Ubc9 binding and SUMO-1 modification. Mutating residues in the SUMO-1-CS abolishes both Ubc9 binding and substrate modification. These findings have important implications for how SUMO-1 substrates are recognized and for how SUMO-1 is ultimately transferred to specific lysine residues on these substrates.

Amino Acid Sequence↗
PubMed2001

Stimulation of mammary tumorigenesis by systemic tissue inhibitor of matrix metalloproteinase 4 gene delivery.

Tissue inhibitors of matrix metalloproteinase (TIMPs) are multifunctional proteins with both matrix metalloproteinase (MMP) inhibitory effects and growth-regulatory activity. TIMPs inhibit MMP activity, suggesting a use for cancer gene therapy. However, here we report that systemic administration of human TIMP-4 by electroporation-mediated i.m. injection of naked TIMP-4 DNA stimulates tumorigenesis of human breast cancer cells in nude mice. Consistent with tumor stimulation, TIMP-4 up-regulates Bcl-2 and Bcl-X(L) protein. TIMP-4 also inhibits apoptosis in human breast cancer cells in vitro and mammary tumors in vivo. A synthetic MMP inhibitor BB-94 did not have such antiapoptotic effect. Analysis of TIMP-4 expression in human mammary specimens indicates that TIMP-4 protein is increased in mammary carcinoma cells compared with normal mammary epithelial cells. These data indicate an antiapoptotic activity in breast cancer cells and a tumor-stimulating effect of TIMP-4 when administrated systemically.

Animals↗
PubMed2001

Gene silencing: fleshing out the bones.

Genetic studies are revealing the pathway for RNA-mediated gene silencing. Short RNA molecules are the key, giving sequence specificity for RNA degradation and mediating communication within and between cells; these short RNAs are common to transcriptional and post-transcriptional silencing pathways.

Animals↗
PubMed2001

Dipeptidyl peptidase IV is a target for covalent adduct formation with the acyl glucuronide metabolite of the anti-inflammatory drug zomepirac.

The nonsteroidal anti-inflammatory drug zomepirac (ZP) is metabolised to a chemically reactive acyl glucuronide conjugate (ZAG) which can form covalent adducts with proteins. In vivo, such adducts could initiate immune or toxic responses. In rats given ZP, the major band detected in liver homogenates by immunoblotting with a polyclonal ZP antiserum was at 110 kDa. This adduct was identified as ZP-modified dipeptidyl peptidase IV (DPP IV) by immunoblotting using the polyclonal ZP antiserum and monoclonal DPP IV antibodies OX-61 and 236.3. In vitro, ZAG, but not ZP itself, covalently modified recombinant human and rat DPP IV. Both monoclonal antibodies recognized DPP IV in livers from ZP- and vehicle-dosed rats. Confirmation that the 110 kDa bands which were immunoreactive with the ZP and DPP IV antibodies represented the same molecule was obtained from a rat liver extract reciprocally immunodepleted of antigens reactive with these two antibodies. Furthermore, immunoprecipitations with OX-61 antibody followed by immunolotting with ZP antiserum, and the reciprocal experiment, showed that both these antibodies recognised the same 110 kDa molecule in extracts of ZP-dosed rat liver. The results verify that DPP IV is one of the protein targets for covalent modification during hepatic transport and biliary excretion of ZAG in rats.

Animals↗
PubMed2001

Brain-derived neurotrophic factor-enriched collagen tubule as a substitute for autologous nerve grafts.

BACKGROUND: Autologous nerve interposition grafts are frequently harvested by head and neck surgeons. The sacrifice of these donor nerves guarantees some degree of morbidity, including sensory loss, additional incision sites with associated potential complications, and prolonged operative time. An alternative to autologous nerve grafting is, therefore, desirable. OBJECTIVE: To determine if a collagen tubule (CT) filled with either a plain collagen gel or a brain-derived neurotrophic factor (BDNF)-enriched collagen gel could be used to achieve functional and histologic outcomes equivalent to an autologous nerve graft in bridging a 15-mm nerve gap in the rabbit facial nerve. DESIGN: A prospective, randomized, blinded animal study with a control group. METHODS: Thirty rabbit facial nerves were resected (15-mm segments) to create nerve gaps. The gaps were bridged using 1 of 3 methods, assigned randomly: a reversed facial nerve (control), a collagen gel-filled CT, or a BDNF-enriched collagen gel-filled CT. The animals were evaluated after 6 weeks in a blinded fashion for functional nerve recovery, axon count, and axonal diameter. RESULTS: There were no significant differences between the autologous nerve graft group, the collagen gel-filled CT group, or the BDNF-enriched collagen gel-filled CT group (n = 10 for each group) for functional nerve recovery (P =.94). The mean axon count and the mean axonal diameter were highest in the BDNF-enriched collagen gel-filled CT group, but these differences failed to reach statistical significance (P =.18 and.96, respectively). CONCLUSIONS: Collagen tubules filled with BDNF-enriched collagen gel appear to be at least as good as autologous nerve grafts for bridging short facial nerve gaps. Larger experimental studies are warranted to determine if clinical trials are justified.

Animals↗
PubMed2001

A regulatory region rearranged BK virus is associated with tubulointerstitial nephritis in a rejected renal allograft.

A renal allograft transplant patient with high serum creatinine presented clinical symptoms of rejection. Sections of renal biopsy tissue showed mononuclear leukocyte infiltration in the tubulointerstitium and nuclear enlargement with inclusions in the tubular epithelium. The morphological characteristics resembled polyomavirus-induced interstitial nephritis. Electron microscopy of the nuclear inclusions showed paracrystalline arrays of naked viral particles with a diameter of 45 nm. Molecular studies revealed that a new variant of BK virus (BKV) with rearrangement at the regulatory region was involved in the nephritis. The BKV regulatory region contained a tandem repeat from the P-block to the Q-block causing duplication of several important transcriptional elements or transcriptional factor binding motifs. This is the first report to show a naturally occurring BKV variant with regulatory region rearrangement associated with tubulointerstitial nephritis.

Adult↗
PubMed2001

Identification of a DNA encapsidation sequence for human polyomavirus pseudovirion formation.

Human polyomavirus is a naked capsid virus containing a closed circular double-stranded DNA genome. The mechanism of DNA encapsidation for the viral progeny formation is not fully understood. In this study, DNA encapsidation domain of the major capsid protein, VP1, of the human polyomavirus JCV was investigated. When the first 12 amino acids were deleted, the E. coli expressed VP1 (Delta N12VP1) failed to encapsidate the host DNA although the integrity of the capsid-like structure was maintained. In addition, capsid-like particles of Delta N12VP1 did not package exogenous DNA in vitro, which is in contrast to that of the full-length VP1 protein. These findings suggest that the N-terminal of the first 12 amino acids of VP1 were responsible for DNA encapsidation. The importance of amino acids in the DNA encapsidation domain was determined further using site-directed mutagenesis. All of the positively charged amino acids at the N-terminal region of VP1 were essential for DNA encapsidation. The results indicate that the N-terminal region of the human polyomavirus major capsid protein VP1 may be involved in viral genome encapsidation during progeny maturation.

Amino Acid Sequence↗
PubMed2001

In silico mining of EST databases for novel pre-implantation embryo-specific zinc finger protein genes.

Progress in the understanding of early mammalian embryo development has been severely hampered by scarcity of study materials. To circumvent such a constraint, we have developed a strategy that involves a combination of in silico mining of new genes from expressed sequence tags (EST) databases and rapid determination of expression profiles of the dbEST-derived genes using a PCR-based assay and a panel of cDNA libraries derived from different developmental stages and somatic tissues. We demonstrate that in a random sample of 49 independent dbEST-derived zinc finger protein genes mined from a mouse embryonic 2-cell cDNA library, more than three-quarters of these genes are novel. Examination of characteristics of the human orthologues derived from these mouse genes reveals that many of them are associated with human malignancies. Expression studies have further led to the identification of three novel genes that are exclusively expressed in mouse embryos before or up to the 8-cell stage. Two of the genes, designated 2czf45 and 2czf48 (2czf for 2-cell zinc finger), are zinc finger protein genes coding for a RBCC protein with a RFP domain and a protein with three C2H2 fingers, respectively. The third gene, designated 2cpoz56, codes for a protein with a POZ domain that is often associated with zinc finger proteins. These three genes are candidate genes for regulatory or other functions in early embryogenesis. The strategy described in this report should generally be applicable to rapid and large-scale mining of other classes of rare genes involved in other biological and pathological processes. Mol. Reprod. Dev. 59:249-255, 2001.

Animals↗
PubMed2001

Blocking CTL-based cytotoxic pathways reduces apoptosis of transplanted hepatocytes.

BACKGROUND: A major obstacle in allogenic hepatocyte transplantation is increased apoptosis of grafted cells due to CTL-based cytotoxicity. However, whether blockade of Fas- and granzyme-mediated pathways of CTL-based cytotoxicity may provide immune protection to transplanted hepatocytes is poorly defined. Our study aimed to reduce apoptosis of allogenic transplanted hepatocytes by inhibiting granzyme B (GraB) activity and blocking Fas-FasL interaction. MATERIALS AND METHODS: Hepatocyte transplantation was performed by inoculating isolated liver cells from ACI rats (allogenic) or Lewis rats (syngenic) into the spleens of Lewis rats. Recipients were treated with FLIM58, an inhibitory anti-FasL mAb, and GraB inhibitor I alone or a combination of the two drugs for 5 days after transplantation, and were sacrificed at Day 7. Apoptosis of transplanted hepatocytes was detected in situ by TUNEL assay and M30 immunostaining. Glutamate dehydrogenase (GLDH) activity in recipient spleens was examined to evaluate survival of transplanted cells. Recipient spleens were assayed for FasL level with Western blotting and for GraB activity by hydrolysis of GraB substrate. RESULTS: FLIM58 or GraB inhibitor I significantly reduced the percentage of TUNEL-positive and M30-positive hepatocytes and markedly increased GLDH levels in allogenic, but not syngenic, recipient spleens. These effects were more pronounced when the two drugs were used in combination (P < 0.05). Additionally, elevation of FasL and GraB levels in allogenic recipient spleens can be significantly reduced by FLIM58 and GraB inhibitor I, respectively. CONCLUSIONS: Inhibition of GraB activity and blockade of Fas-FasL interaction reduce the apoptosis of allogenic transplanted hepatocytes, and thus improve their survival.

Animals↗
PubMed2001

High levels of nitric oxide production decrease early but increase late survival of Brucella abortus in macrophages.

Nitric oxide (NO), produced by the iNOS protein, is known as a defense mechanism against various pathogens and an apoptotic inducer of cells. Apoptosis can also be a host protective mechanism against intracellular bacteria. The intracellular survival of Brucella abortus in RAW264.7 macrophages was examined under conditions of the apoptotic inducer, NO. Since B. abortus does not induce high output of NO, Escherichia coli LPS and IFN-gamma, as potential therapeutic modalities, were added to increase the expression of iNOS, and thus NO. Using 10 ng/ml E. coli LPS and 25 U/ml IFN-gamma, nitrite production was as high as 140 microM by 72 h. However, when macrophages were infected with B. abortus, the nitrite concentration was 60 microM after 72 h post infection, greater than a two-fold decrease. The number of surviving bacteria decreased, from 6 to 24 h, in the presence of nitrite accumulation. In the absence of B. abortus there was an increase in apoptotic cells at 72 h with high nitrite accumulation. In contrast, the number of macrophage apoptotic bodies decreased in the presence of B. abortus. The data suggest that: (i) NO accelerates the killing of intracellular B. abortus, but not to completion during the first 24 h of infection; (ii) B. abortus can prevent apoptosis as an advantage for bacterial survival inside macrophages and (iii) surviving intracellular bacteria then replicate steadily after 24 h. B. abortus probably expresses genes that counteract the effect of a high NO environment or activates genes to utilize NO as a nitrogen source, as the Brucella genome codes for nitric and nitrous oxide reductase genes.

Animals↗
PubMed2001

An adenovirus with enhanced infectivity mediates molecular chemotherapy of ovarian cancer cells and allows imaging of gene expression.

The adenovirus (Ad) is a useful vector for cancer gene therapy due to its unparalleled gene transfer efficiency to dividing and quiescent cells. Primary cancer cells, however, often have highly variable or low levels of the requisite coxsackie-adenovirus receptor (CAR). Also, assessment of gene transfer and vector persistence has been logistically difficult in human clinical trials. We describe here two novel bicistronic adenoviral (Ad) vectors, AdTKSSTR and RGDTKSSTR, which contain the herpes simplex virus thymidine kinase gene (TK) for molecular chemotherapy and bystander effect. In addition, the viruses contain the human somatostatin receptor subtype-2 gene (SSTR2), the expression of which can be noninvasively imaged. We enhanced the infectivity of RGDTKSSTR by genetically incorporating the RGD-4C motif into the HI-loop of the fiber. This allows the virus to circumvent CAR deficiency by binding to alpha(v)beta(3) and alpha(v)beta(5) integrins, which are highly expressed on most ovarian cancers. The expanded tropism of RGDTKSSTR results in increased infectivity of purified primary ovarian cancer cells and allows enhanced gene transfer in the presence of malignant ascites containing anti-Ad antibodies. RGDTKSSTR may be a useful agent for treating ovarian cancer in clinical trials.

Adenoviridae↗
PubMed2001

The gene for slow Wallerian degeneration (Wld(s)) is also protective against vincristine neuropathy.

Neurological diseases are frequently associated with axonal degeneration, which leads to dysfunction though separation of neurons from their targets. The mechanisms of axonal degeneration are largely unknown and in many cases are independent of those occurring within cell bodies in neurodegenerative disorders. The Wld(s) mouse mutant demonstrates the unique phenotype of resistance to axonal degeneration after axotomy (slow Wallerian degeneration), making it a powerful tool for studying mechanisms of axonal degeneration. We asked whether the Wld(s) mutation also provides resistance to axonal degeneration in a slowly progressing neuropathy. Using cultured dorsal root ganglion neurons we compared the course of axonal degeneration in response to exposure to the neurotoxin vincristine and found that Wld(s) neurites were relatively resistant to vincristine neuropathy. These findings suggest common pathophysiologic mechanisms between axotomy-induced Wallerian degeneration and toxic neuropathy. The implications are wide-ranging and are relevant to the pathophysiology of axonal degeneration seen in a wide spectrum of neurological diseases ranging from stroke and head trauma to spinal cord injury and peripheral neuropathy.

Animals↗
PubMed2001

Experimental study on AT1-receptor-peptide-induced myocardial immune damage in rat.

In order to investigate the immunological damage in rat immunized with AT1-receptor peptide, 18 male Wistar rats were divided into two groups: immunized-group (n = 12), each rat was immunized with 150 micrograms AT1-receptor peptide coupled to bovine serum albumin, together with Freund's adjuvant. Control group (n = 6), sham-immunized, "immunized liquid" was same as immunized-group except AT1-receptor peptide. Systolic blood pressure (SBP) was measured by using the tail-cuff technique, antibody against AT1-receptor peptide detected by using ELISA method, and left ventricular myocardium and renal cortex sections were observed under light and electron microscopy. There was no significant difference in SBP and light microscopic observation of the tissue sections between the immunized-group and control group. The O.D. value of anti-AT1-receptor peptide antiserum was significantly higher in the immunized-group than in the rats before immunization and control group (P < 0.01). Positive rate in the immunized-group was 100%, while 0% in the control group. Ultramicroscopic morphology showed potential myocardial injury, including: increase in number of mitochondria, swelling of many mitochondria with reduction in number or absence of their cristae and cristolysis, disorder of the cardiac myofibrils, and myofibrillar disruption and myocytolysis. And lysosomes were increased in renal tubular epithelia. The AT1-receptor peptide could induce to generate the antibody against AT1-receptor peptide and lead to myocardial and renal damage in rats.

Animals↗
PubMed2001

Changes in serum leptin levels during r-hGH treatment in growth hormone-deficient children.

To observe the effect of growth hormone on serum leptin levels, serum leptin concentrations were measured by enzyme immunoassay in 12 prebuttal children with growth hormone deficiency 1, 3 and 6 months before and after the treatment with recombinant human growth hormone (r-hGH). For comparison, 34 normal prepubertal children were also investigated. Relationship between leptin levels and body mass index (BMI) was observed at the same time. Our results showed that serum leptin level in normal prepubertal children was 1.22 +/- 0.34 ng/ml; the pretreatment serum leptin levels in GHD children was 3.08 +/- 2.41 ng/ml, which was significantly different from those 1, 3 and 6 months after GH treatment (i.e. 1.64 +/- 1.37 ng/ml, 1.57 +/- 1.40 ng/ml and 1.35 +/- 0.89 ng/ml respectively) (all P < 0.001). Our results suggested that r-hGH has a suppressive effect on leptin expression.

Adolescent↗
PubMed2001

Mouse histamine N-methyltransferase: cDNA cloning, expression, gene cloning and chromosomal localization.

OBJECTIVE: Histamine N-methyltransferase (HNMT) catalyzes the Ntau-methylation of histamine. We set out to clone a mouse liver HNMT cDNA and the mouse HNMT gene as steps toward characterizing molecular genetic mechanisms involved in the regulation of this important histamine-metabolizing enzyme. DESIGN: A PCR-based strategy was used to clone both the mouse HNMT cDNA and the gene encoding that cDNA, Hnmt. The cDNA was used both to express recombinant mouse HNMT and to determine the chromosomal localization of Hnmt. RESULTS: The mouse liver HNMT cDNA was 1657 bp in length with an 888 bp open reading frame (ORF) that encoded a 296 amino acid protein with a predicted Mr value of approximately 32.5 kDa. The amino acid sequence of the encoded protein was 84% identical to that of human kidney HNMT. Mouse HNMT was expressed in COS-1 cells, and its apparent Km values for histamine and S-adenosyl-L-methionine (Ado-Met), the two cosubstrates for the reaction, were 5.3 and 5.8 microM, respectively. The mouse HNMT gene, Hnmt, spanned approximately 25 kb and had 7 exons. Its structure differed from that of the human gene primarily by the presence of an additional exon at the 5'-terminus. Hnmt mapped to mouse chromosome 2 in an area of conserved synteny to human chromosome 2q, the location of the human gene (2q22) on the basis of fluorescence in situ hybridization. CONCLUSIONS: Cloning and functional characterization of the mouse HNMT cDNA and gene will now make it possible to study in the mouse molecular genetic mechanisms involved the regulation of this important histamine-metabolizing enzyme.

Amino Acid Sequence↗