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Biomedical subjects

M Zhong

Publications and source records attributed to M Zhong.

At least 55 records · Page 3Linked to original sources
PubMed1999

Mammalian subtilisin/kexin isozyme SKI-1: A widely expressed proprotein convertase with a unique cleavage specificity and cellular localization.

Using reverse transcriptase-PCR and degenerate oligonucleotides derived from the active-site residues of subtilisin/kexin-like serine proteinases, we have identified a highly conserved and phylogenetically ancestral human, rat, and mouse type I membrane-bound proteinase called subtilisin/kexin-isozyme-1 (SKI-1). Computer databank searches reveal that human SKI-1 was cloned previously but with no identified function. In situ hybridization demonstrates that SKI-1 mRNA is present in most tissues and cells. Cleavage specificity studies show that SKI-1 generates a 28-kDa product from the 32-kDa brain-derived neurotrophic factor precursor, cleaving at an RGLT downward arrowSL bond. In the endoplasmic reticulum of either LoVo or HK293 cells, proSKI-1 is processed into two membrane-bound forms of SKI-1 (120 and 106 kDa) differing by the nature of their N-glycosylation. Late along the secretory pathway some of the membrane-bound enzyme is shed into the medium as a 98-kDa form. Immunocytochemical analysis of stably transfected HK293 cells shows that SKI-1 is present in the Golgi apparatus and within small punctate structures reminiscent of endosomes. In vitro studies suggest that SKI-1 is a Ca2+-dependent serine proteinase exhibiting a wide pH optimum for cleavage of pro-brain-derived neurotrophic factor.

Amino Acid Sequence↗
PubMed1999

Sensitivity of Drosophila heat shock transcription factor to low pH.

The heat shock transcription factor (HSF) mediates the induction of heat shock gene expression. The activation of HSF involves heat shock-induced trimerization, binding to its cognate DNA sites, and the acquisition of transcriptional competence. In this study, the oligomeric properties of Drosophila HSF were analyzed by equilibrium analytical ultracentrifugation and gel filtration chromatography. Previous findings showed that trimerization of purified Drosophila HSF was directly sensitive to heat and oxidation (1). Here we report that low pH, in the physiological range, also directly induces HSF trimerization and DNA binding in vitro. Furthermore, the induction of HSF trimerization by low pH is synergistic with the actions of heat and oxidation. Since heat or chemical stress leads to a moderate decrease of intracellular pH, we suggest that intracellular acidification may contribute to activating the heat shock response in vivo.

Animals↗
PubMed1999

Tubular-wire dual electrode for detection of thiols and disulfides by capillary electrophoresis/electrochemistry.

A new dual-electrode detector for capillary electrophoresis is described. The detector consists of an integrated gold tubular electrode as the generator electrode and a gold wire electrode for detection. The detector configuration, including electrode size and position, has been optimized in terms of detection sensitivity and separation efficiency. After amalgamation of the dual electrode with mercury, the capillary electrophoresis/electrochemistry system was employed for simultaneous detection of thiols and disulfides. The response of cystine was found to be linear from 1 microM to 1 mM with a LOD of 0.5 microM (S/N = 3) and sensitivity of 60 pA/microM. The detection limits represent 200-fold improvement over previously reported dual-electrode designs for the detection of disulfides. The use of this detector for identification of thiol- and disulfide-containing peptides was demonstrated with a tryptic digest of ribonuclease A.

Disulfides↗
PubMed1999

Inhibitory effect of resveratrol on interleukin 6 release by stimulated peritoneal macrophages of mice.

In the present investigation, interleukin 6 (IL-6) activity in the supernatant of cultured mouse peritoneal macrophages was monitored using a sensitive bioassay involving the IL-6-dependent murine hybridoma B9 cell line. The effects of resveratrol on Il-6 release by mouse peritoneal macrophages stimulated with calcium ionophore A23187 and fMLP were explored. Resveratrol, at a concentration range from 5 x 10(-6) to 4 x 10(-5) mol.l-1, was found to dose-dependently inhibit IL-6 release by cultured macrophages induced by A23187 and fMLP, and showed no direct cytotoxic effect, but induced proliferation of cultured mouse thymus cells. Resveratrol, at a concentration range from 10(-8) to 10(-5) mol.l-1, was shown to dose-dependently inhibit calcium ion influx into the cells with the stimulation of fMLP (10(-6) mol.l-1). These results suggest that the blocking of calcium ion influx into cells by reveratrol is one of the possible mechanisms of the IL-6 biosynthesis inhibitory action of resveratrol.

Animals↗
PubMed1999

Antagonistic effects of protein kinase C alpha and delta on both transformation and phospholipase D activity mediated by the epidermal growth factor receptor.

Downregulation of protein kinase C delta (PKC delta) by treatment with the tumor-promoting phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) transforms cells that overexpress the non-receptor class tyrosine kinase c-Src (Z. Lu et al., Mol. Cell. Biol. 17:3418-3428, 1997). We extended these studies to cells overexpressing a receptor class tyrosine kinase, the epidermal growth factor (EGF) receptor (EGFR cells); like c-Src, the EGF receptor is overexpressed in several human tumors. In contrast with expectations, downregulation of PKC isoforms with TPA did not transform the EGFR cells; however, treatment with EGF did transform these cells. Since TPA downregulates all phorbol ester-responsive PKC isoforms, we examined the effects of PKC delta- and PKC alpha-specific inhibitors and the expression of dominant negative mutants for both PKC delta and alpha. Consistent with a tumor-suppressing function for PKC delta, the PKC delta-specific inhibitor rottlerin and a dominant negative PKC delta mutant transformed the EGFR cells in the absence of EGF. In contrast, the PKC alpha-specific inhibitor Go6976 and expression of a dominant negative PKC alpha mutant blocked the transformed phenotype induced by both EGF and PKC delta inhibition. Interestingly, both rottlerin and EGF induced substantial increases in phospholipase D (PLD) activity, which is commonly elevated in response to mitogenic stimuli. The elevation of PLD activity in response to inhibiting PKC delta, like transformation, was dependent upon PKC alpha and restricted to the EGFR cells. These data demonstrate that PKC isoforms alpha and delta have antagonistic effects on both transformation and PLD activity and further support a tumor suppressor role for PKC delta that may be mediated by suppression of tyrosine kinase-dependent increases in PLD activity.

Acetophenones↗
PubMed1999

Detection of neuroendocrine feature about salivary adenoid cystic carcinoma.

OBJECTIVE: To study the possibility of neuroendocrine feature of salivary adenoid cystic carcinoma (SACC). METHODS: Immunohistochemical staining was operated in SACC with S-100 protein, neurospecific enolase (NSE), chromogranin A, and synaptophysin, which are specific and correlative with neuroendocrine cells and neoplasias. RESULTS: The positive expression of S-100, NSE, chromogranin, and synaptophysin were 42, 32, 8, and 7, respectively, in 50 cases of SACC. CONCLUSION: Neuroendocrine feature in some SACC cells is possibly indicated.

Adolescent↗
PubMed1999

[Gamma delta T cells and their receptor Vdelta gene rearrangement in transitional mucosa of rectal carcinoma].

OBJECTIVE: To observe gamma delta T cells and their receptors Vdelta gene rearrangement expression in the transitional mucosa of rectal carcinoma, and to investigate their relations with local tumor recurrence. METHODS: In 13 patients with rectal carcinoma, gamma delta T cells in transitional mucosa were measured with two-color immunofluorescent flow cytometric analysis and polymerase chain reaction technique. RESULTS: The volume of gamma delta T cells in transitional mucosal lymphocytes (TML) were significantly higher than that in tumor infiltrating lymphocytes (TIL), and significantly lower than that in normal intestinal epithelial lymphocytes (IEL) (P < 0.05). The gamma delta T cell receptors delta chain V region (gamma delta-TCR Vdelta1-Vdelta6) gene rearrangement was all expressed in TIL, TML and IEL; but the Vdelta 6 gene expression in both TML and TIL were higher than that in IEL (P < 0.05). CONCLUSIONS: The decreased volume of gamma delta T cells in TML resulted in decreased local immune function and may be responsible for the local tumor recurrence after resection. The high expression of Vdelta 6 gene rearrangement in both TIL and TML suggests the skewing of gamma delta-TCR receptors and the abtitumor specificity of TIL and TML in local epithelium.

Adult↗
PubMed1999

[Expression of WT1 gene mRNA in leukemia].

UNLABELLED: WT1 gene expression was determined in acute leukemic patients by RT-PCR. RESULTS: 19 patients were found to have positive results among 28 patients, the positive rate is 67.9%, and in 16 ANLL patients, 11 cases got positive results, in 12 ALL patients, 8 cases got positive results. It suggested that WT1 was highly expressed in acute leukemia. It is a new and significant detective marker for monitoring the effect of chemotherapy, predicting the prognosis and in the research of minimal residual disease (MRD).

Acute Disease↗
PubMed1999

[Detection of neuroendocrine feature about salivary adenoid cystic carcinoma].

OBJECTIVE: To study the possibility of neuroendocrine feature of salivary adenoid cystic carcinoma (SACC). METHODS: Immunohistochemical staining was operated in SACC with S-100 protein, Neuron-specific enolase (NSE), chromagranin A and synaptophysin which are specific and correlative with neuroendocrine and neoplasias. RESULTS: The positive expression of S-100, NSE, chromogranin and synaptophysin were respectively 42, 32, 8 and 7 in 50 cases of SACC. CONCLUSION: There is possibility of neuroendocrine feature in some SACC cells.

Adult↗
PubMed1998

Direct sensing of heat and oxidation by Drosophila heat shock transcription factor.

The heat shock transcription factor HSF activates expression of its target genes in response to elevated temperatures and chemical or physiological stress. A key step in the activation process involves the formation of HSF homotrimers, leading to high-affinity DNA binding. The mechanism by which HSF trimerization and DNA binding is regulated by stress signals has remained elusive. Here, we report that trimerization and DNA binding of purified Drosophila HSF can be directly and reversibly induced in vitro by heat shock temperatures in the physiological range and by an oxidant, hydrogen peroxide. Other inducers of the heat shock response, including salicylate, dinitrophenol, ethanol, and arsenite, have no effect on HSF trimerization in vitro, indicating that these inducers act by indirect mechanisms.

Adaptation, Physiological↗
PubMed1998

[Changes of serum myocardial enzymes in patients with malignant hematologic diseases].

Myocardial enzymes including aspartate aminotransferase (AST), lactate dehydrogenase (LDH), HBD (LDH1 and LDH2), and creatine kinase (CK) and its isoenzyme were monitored in 106 cases of malignant hematologic diseases. The findings were that average values of LDH and HBD increased. There were 82.4% myocardial enzyme levels of 51 patients with leukemia, lymphoma, and myelodysplastic syndrome (MDS) returning to normal or making an obvious reduction after chemotherapy associated with drugs of heart toxicity, while there were increases of the myocardial enzyme levels before chemotherapy. Patients with the increasing of enzyme levels were only 3.9% after chemotherapy. After several courses of chemotherapy, the positive rates of the increasings of CK and CK-MB were higher than that of pretreatment. The results suggest that the injuries of myocardium are possible.

Anthracyclines↗
PubMed1998

[A clinical observation on the leukopenia treated with shengbaikuai decoction].

OBJECTIVE: To study the effective orally taken medicine in treating the leukopenia. METHODS: Ninety cases of leukopenia induced by chemotherapy were divided into 3 groups. They were test group treated with Shengbaikuai Decoction (SBK), control group and blank group. Their efficacies were compared with each other, 25 cases with nonchemotherapy were also observed. RESULTS: WBC count and absolute neutrophil count (ANC) in test group were higher than those of other groups immediately after treatment and 2 weeks later (P < 0.01). The marked effective rate and total effective rate of test group after 2 weeks were 60.0% and 80.0% (P < 0.001) respectively, which were significantly higher than those of other groups. WBC and ANC count after treatment and 3 weeks later were higher than those before treatment in nonchemotherapy group (P < 0.001). The marked effective rate and total effective rate 3 weeks later were 64.0% and 80.0% respectively. CONCLUSIONS: SBK could increase WBC count rapidly and definitely without apparent side effect.

Adolescent↗
PubMed1998

[Effects of isorhapotigenin and resveratrol on function of peripheral blood polymorphonuclear leukocytes from rabbits].

In effects of isorhapotigenin and resveratrol on chemotaxis and release of beta-glucuronidase from rabbit peripheral blood polymorphonuclear leukocytes stimulated with formylmethionyl-leucyl-phenyl-phenylalanine (fMLPP) were studied. The optimal concentration of fMLPP for the induction of chemotaxis of polymorphonuclear leukocytes was approximately at 5 x 10(-10) mol.L-1. fMLPP, at the concentration range of 10(-10)-10(-6) mol.L-1, dose-dependently increased the release of beta-glucuronidase from polymorphonuclear leukocytes. Isorhapotigenin and resveratrol both inhibited the chemotaxis of polymorphonuclear leukocytes stimulated with fMLPP(5 x 10(-10) mol.L-1). Isorhapotigenin, at the concentration range of 10(-5)-10(-4) mol.L-1, dose-dependently inhibited the release of beta-glucuronidase from polymorphonuclear leukocytes stimulated with fMLPP(10(-8) mol.L-1). Resveratrol, at the concentrations of 1 x 10(-5) and 3 x 10(-5) mol.L-1, was shown to dose-dependently inhibit beta-glucuronidase release from polymorphonuclear leukocytes stimulated with fMLPP(10(-8) mol.L-1).

Animals↗
PubMed1998

[Effects of leukotrienes on production of interleukin 6 from mouse peritoneal macrophages].

In the present study, effects of leukotrienes on IL-6 production by mouse resident peritoneal macrophages were explored with a bioassay method involving IL-6 dependent murine hybridoma B9 cell line. The results showed that LTB4, LTC4 and LTD4 can enhance the production of IL-6 from cultured peritoneal macrophages of mice. The optimal concentractions for LTB4, LTC4 and LTD4 were found to be 1.45 x 10(-6), 6.9 x 10(-8) and 8.05 x 10(-8) mol.L-1, respectively. The results suggest that peptide leukotrienes might play important role on IL-6 production in the local milieu of inflammation.

Animals↗
PubMed1997

Identification of a flexible loop region (297-313) of urokinase-type plasminogen activator, which helps determine its catalytic activity.

Pro-urokinase has a much higher intrinsic catalytic activity than other zymogens of the serine protease family. Lys300(c143) in an apparent "flexible loop" region (297-313) was previously shown to be an important determinant of this intrinsic catalytic activity. This was related to the loop allowing the positive charge of Lys300(c143) to transiently interact with Asp355(c194), thereby inducing an active conformation of the protease domain (Liu, J. N., Tang, W., Sun, Z., Kung, W., Pannell, R., Sarmientos, P., and Gurewich, V. (1996) Biochemistry 35, 14070-14076). To further test this hypothesis, the charge at position 300(c143) and the flexibility of the loop were altered using site-directed mutagenesis designed according to a computer model to affect the interaction between Lys300(c143) and Asp355(c194). When the charge at Lys300(c143) but not Lys313(c156) was reduced, a significant reduction in the intrinsic catalytic activity occurred. Similarly, when the flexibility (wobbliness) of the loop was enhanced reducing the size of side chain, the intrinsic catalytic activity was also reduced. By contrast, when the loop was made less flexible, the intrinsic catalytic activity was increased. These findings were consistent with the hypothesis. The effects of these mutations on two-chain activity were less and often discordant with the intrinsic catalytic activity, indicating that they can be modulated independently. This structure-function disparity can be exploited to create a more zymogenic pro-urokinase (lower intrinsic catalytic activity) with a high catalytic activity, as exemplified by two of the mutants. The changes in intrinsic catalytic activity and two-chain activity induced by the mutations were due to changes in kcat rather than Km. Some significant structure-function differences between pro-urokinase and its highly homologous counterpart, tissue plasminogen activator, were also found.

Catalysis↗
PubMed1997

In vitro characterization of the novel proprotein convertase PC7.

Biochemical and enzymatic characterization of the novel proprotein convertase rat PC7 (rPC7) was carried out using vaccinia virus recombinants overexpressed in mammalian BSC40 cells. Pro-PC7 is synthesized as a glycosylated zymogen (101 kDa) and processed into mature rPC7 (89 kDa) in the endoplasmic reticulum. No endogenously produced soluble forms of this membrane-anchored protein were detected. A deletion mutant (65 kDa), truncated well beyond the expected C-terminal boundary of the P-domain, produced soluble rPC7 in the culture medium. Enzymatic activity assays of rPC7 using fluorogenic peptidyl substrates indicated that the pH optimum, Ca2+ dependence, and cleavage specificity of this enzyme are largely similar to those of furin. However, with some substrates, cleavage specificity more closely resembled that of yeast kexin, suggesting differential processing of proprotein substrates by this novel convertase. We examined the rPC7- and human furin-mediated cleavage of synthetic peptides containing the processing sites of three proteins known to colocalize in situ with rPC7. Whereas both enzymes correctly processed the pro-parathyroid hormone tridecapeptide and the pro-PC4 heptadecapeptide, neither enzyme cleaved a pro-epidermal growth factor hexadecapeptide. Thus, this study establishes that rPC7 is an enzymatically functional subtilisin/kexin-like serine proteinase with a cleavage specificity resembling that of hfurin. In addition, we have demonstrated that rPC7 can correctly process peptide precursors that contain the processing sites of at least two potential physiological substrates.

Amino Acid Sequence↗