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Biomedical subjects

H Ding

Publications and source records attributed to H Ding.

At least 145 records · Page 8Linked to original sources
PubMed1996

Impaired actions of insulin-like growth factor 1 on protein Synthesis and degradation in skeletal muscle of rats with chronic renal failure. Evidence for a postreceptor defect.

The actions of insulin-like growth factor 1 (IGF-1) on protein turnover and of the IGF-1 receptor (IGF-1R) were examined in skeletal muscle of rats with chronic renal failure (CRF) and sham operated (SO), pair-fed controls. Acidemia was prevented in CRF rats with NaHCO3. Serum IGF-1 and skeletal muscle IGF-1 and IGF-1 mRNA were reduced in CRF rats. Dose-response studies revealed impaired stimulation of protein synthesis and suppressed inhibition of protein degradation by IGF-1 in epitrochlearis muscle of CRF rats. Neither IGF-1 analogues with low affinity to IGF binding proteins nor proteinase inhibitors obliterated the IGF-1 resistance. In CRF rats, skeletal muscle IGF-1R mRNA was increased; displacement ligand binding studies and affinity labeling of the IGF-1R alpha subunit indicated increased total skeletal muscle IGF-1R number with normal affinity. However, both autophosphorylation of the IGF-1R beta subunit (i.e., IGF-1R tyrosine kinase) and the IGF-1R tyrosine kinase activity towards exogenous insulin receptor substrate-1, a natural substrate for IGF-1R tyrosine kinase, were reduced in CRF fats. These data indicate that in skeletal muscle of CRF rats there is resistance to the IGF-1 effects on protein synthesis and degradation and decreased IGF-1 and IGF-1 mRNA levels; IGF-1R mRNA and number are increased; but activity of IGF-1R tyrosine kinase is impaired. This postreceptor defect may be a cause of the skeletal muscle resistance to IGF-1 in CRF.

Animals↗
PubMed1996

Characterization of a helicase-like transcription factor involved in the expression of the human plasminogen activator inhibitor-1 gene.

A 5.4-kb cDNA encoding the protein that binds to the B Box of the plasminogen activator inhibitor-1 (PAI-1) gene was isolated and sequenced. The protein, named helicase-like transcription factor (HLTF), contains a DNA-binding domain, a RING finger domain, and seven helicase domains and is homologous to SWI/SNF proteins. Two HLTF mRNAs of 5.5 and 4.5 kb were detected in most human tissues, a single gene was located on chromosome 3q24-25, and the protein was located in the nucleoplasm. Two HLTF proteins differing in translation start site (Met-1 or Met-123) were obtained by in vitro translation in reticulocyte lysate or by immunoprecipitation from HeLa cell nuclear extracts. In vitro transcription from the PAI-1 promoter in HeLa cell extracts was inhibited by HLTF antibodies and by the HLTF DNA binding domain. Over-expression of HLTF or HLTFMet123 produced a three-fold induction of PAI-1-LUC transient expression in HeLa cells. Mutation of the PAI-1 B Box led to an eight-fold reduction of basal PAI-1-LUC expression in these cell lines, but did not affect the four- to six-fold induction by phorbol esters.

Amino Acid Sequence↗
PubMed1996

Purification and further characterization of a haemolysin of Actinomyces pyogenes.

A haemolysin produced by Actinomyces pyogenes ATCC 8164 was purified from culture supernatant by ammonium sulphate and polyethylene glycol precipitation, ion-exchange chromatography on DEAE-Sephacel, and fast-protein-liquid-chromatography on Superose 12 prep grade. The purified haemolysin, designated as pyolysin, displayed a single band on poly-acrylamide gel electrophoresis, indicating a molecular weight of 55000. Additionally, using gel filtration, the same molecular weight was estimated. Further studies of the eluate of ion-exchange chromatography using isoelectric focusing also revealed a single protein band at pH 9.38 with haemolytic activity. A specific antiserum produced against pyolysin inhibited the haemolytic activity. The purity of the isolated protein was also determined by Western Blot analysis with antiserum obtained from a cow inoculated with culture supernatant from A. pyogenes and Peptococcus indolicus. The isolated pyolysin appeared to be heat-labile and displayed cytotoxic effects on poly-morphonuclear leucocytes and on pTK2 kidney cells.

Actinomyces↗
PubMed1996

A human colon carcinoma cell line exhibits adhesive interactions with P-selectin under fluid flow via a PSGL-1-independent mechanism.

It has been postulated that endothelial cell adhesion molecules involved in leukocyte recruitment play a role in metastasis. Using an in vitro flow model, we studied the adhesion of the human colon carcinoma cell line KM12-L4 to P-selectin, an inducible endothelial-expressed adhesion molecule involved in leukocyte recruitment. Recombinant forms of P-selectin and Chinese hamster ovary cells stably expressing P-selectin supported attachment and rolling of KM12-L4 cells at 1 to 2 dynes/cm2. The adhesive interactions to P-selectin were abolished by pretreatment of the KM12-L4 cells with neuraminidase but were unaltered by pretreatment of the KM12-L4 cells with O-sialoglycoprotein endopeptidase, an enzyme that cleaves mucin type glycoproteins such as P-selectin glycoprotein ligand-1 (PSGL-1). PSGL-1 is the only counter-receptor for P-selectin known to mediate myeloid cell adhesion to P-selectin under flow. Flow cytometric and Northern blot analyses revealed that KM12-L4 cells did not express PSGL-1 and monoclonal antibody PL1, a function-blocking monoclonal antibody to PSGL-1, had no inhibitory effect on KM12-L4 adhesion to P-selectin under flow. Compared with HL-60 cells, which express PSGL-1, the KM12-L4 cells exhibited a slightly lower rate of attachment to P-selectin and rolled at a significantly higher velocity. In summary, KM12-L4 human colon carcinoma cells interact with P-selectin, under flow, through a PSGL-1-independent adhesion pathway.

Animals↗
PubMed1996

[The expression of bcl-2, P53 and c-erbB-2 onco-proteins in breast cancer and their clinicopathological significance].

In order to study the interrelation between bcl-2, P53 and c-erbB-2 protein, their expression in 145 breast carcinomas were observed by immunohistochemical LSAB method. 19 cases were negative, 12 were positive for the three oncoproteins. 19 cases were positive for c-erbB-2 only. 20 positive for P53 only and 20 positive for bcl-2 only. 26 cases were positive for both P53 and c-erbB-2, 17 for both c-erbB-2 and bcl-2, 12 for both P53 and bcl-2. Strong inverse correlation was found between the expression of P53 and bcl-2 (P < 0.001). The P53 expression or the expression of both P53 and c-erbB-2 correlated with low histological grade, while the bcl-2 expression correlated with high histological grade (P < 0.05). The necrosis in tumors with only bcl-2 expression was less than those with c-erbB-2 or both c-erbB-2 and P53 expression (P < 0.02). Mitosis count was lower in tumors with only bcl-2 expression than those with expression of both c-erbB-2 and P53 (P < 0.05). The expression of c-erbB-2 and P53 proteins are associated with poor prognosis (P < 0.02, P < 0.05) but not affected by bcl-2. The results suggest that bcl-2, P53 and c-erbB-2 are involved in tumorigenesis and development of breast carcinoma with different biological effects.

Adult↗
PubMed1996

A molecular pathologic study on apoptosis in retinoblastoma and the mechanism of spontaneous regression in retinoblastoma.

OBJECTIVE: The present study was designed to prove the existence of apoptosis in retinoblastoma (Rb) and to determine the pathogenic mechanism of spontaneous regression of Rb as well as the relationship between them. METHODS: Qualitative morphological study on Rb was performed by means of light microscope, electron microscope and TdT mediated biotin-dUTP nick-end labeling (TUNEL). Quantitative study was performed by automatic image analysis technology (AIAT) stained with Feulgen reaction. RESULTS: The characteristic regressed area occurred in all 47 cases of Rb. Morphological changes observed within Rb closely resembled the apoptotic cell described by Kerr et al in 1972. Under electron microscope, details of apoptosis were observed in 7 cases of Rb: the morphological sequence of events occurred in and around the cell nucleus. The morphology of the TUNEL labeling Rb cell was various. TUNEL labeling showed more positive cells in regressed area, while fewer in advanced area. AIAT revealed that apoptosis index (AI) in regressed areas was higher than that in other areas of Rb, DNA average ploidy (DP) consisted with the histology grade of Rb, the degree of hyperdiploid (DH) in metastatic area was distinctly higher than that in other areas (P < 0.05). CONCLUSIONS 1. Morphologic evidence proved the existence of apoptosis, especially more in regressed area of Rb. 2. Apoptosis may participate in the spontaneous regression of Rb. Apoptosis contributed to the spontaneous regression of Rb. 3. Tumor growth parameters of Rb (AI, DP, DH) obtained by AIAT may be used as quantitative index for pathologic classification, the selection of clinical treatment and the prognostic evaluation.

Apoptosis↗
PubMed1995

Ubiquinone pair in the Qo site central to the primary energy conversion reactions of cytochrome bc1 complex.

The mechanistic heart of the ubihydroquinone-cytochrome c oxidoreductase (cyt bc1 complex) is the catalytic oxidation of ubihydroquinone (QH2) at the Qo site. QH2 oxidation is initiated by ferri-cyt c, mediated by the cyt c1 and [2Fe-2S] cluster of the cytochrome bc1 complex. QH2 oxidation in turn drives transmembrane electronic charge separation through two b-type hemes to another ubiquinone (Q) at the Qi site. In earlier studies, residues F144 and G158 of the b-heme containing polypeptide of the Rhodobacter capsulatus cyt bc1 complex were shown to be influential in Qo site function. In the present study, F144 and G158 have each been singly substituted by neutral residues and the dissociation constants measured for both Q and QH2 at each of the strong and weak binding Qo site domains (Qos and Qow). Various substitutions at F144 or G158 were found to weaken the affinities for Q and QH2 at both the Qos and Qow domains variably from zero to beyond 10(3)-fold. This produced a family of Qo sites with Qos and Qow domain occupancies ranging from nearly full to nearly empty at the prevailing approximately 3 x 10(-2) M concentration of the membrane ubiquinone pool (Qpool). In each mutant, the affinity of the Qos domain remained typically 10-20-fold higher than that of the Qow domain, as is found for wild type, thereby indicating that the single mutations caused comparable extents of the weakening at each domain. Moreover, the substitutions were found to cause similar decreases of the affinities of both Q and QH2 in each domain, thereby maintaining the Q/QH2 redox midpoint potentials (Em7) of the Qo site at values similar to that of the wild type. Measurement of the yield and rate of QH2 oxidation generated by single turnover flashes in the family of mutants suggests that the Qos and Qow domains serve different roles for the catalytic process. The yield of the QH2 oxidation correlates linearly with Qos domain occupancy (QH2 or Q), suggesting that the Qos domain exchanges Q or QH2 with the Qpool at a rate which is much slower than the time scale of turnover.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗
PubMed1995

Ion pair formation between basic residues at 144 of the Cyt b polypeptide and the ubiquinones at the Qo site of the Cyt bc1 complex.

Loci of spontaneous Qo site inhibitor resistant mutants in the cyt bc1 complex of the photosynthetic bacterium Rhodobacter capsulatus are M140, F144, G152, G158, and T163 of the cyt b polypeptide. In this report, we have studied the effects of arginine (R) substitution at these positions with a view to test for specific interactions with the [2Fe-2S] cluster, cyt bL with Qo site ubiquinone (Q), or hydroquinone (QH2). All the arginine mutants displayed severely or completely impeded photosynthetic growth resulting from dysfunctional cyt bc1 complexes. The source of dysfunction in G158R and T163R was identified by a > 1000-fold decrease in the Qo site affinity for QH2 and Q, sufficient to empty the site in the presence of the 30 mM ubiquinone pool of the chromatophore membrane; they appear similar to the class of mutants described in the preceding paper [Ding, H., Moser, C. C., Robertson, D. E., Tokito, M., Daldal, F., & Dutton, P. L (1995) Biochemistry 34, 15979-15996]. The source(s) of dysfunction of M140R and G152R is not so apparent since they possess Qo sites with normal QH2/Q affinity; they appear to be members to the class of mutants identified and characterized in the following paper [Saribaş, S., Ding, H., Dutton, P. L., & Daldal, F. (1995) Biochemistry 34, 16004-16012]. The present paper focuses on the unique affects of F144R. Redox potential and EPR spectral properties of the Qo site of F144R showed that arginine forms an ion pair with the head group of an anionic ubiquinone, tentatively suggested to be a ubihydroquinone anion (QH-) in the Qos domain.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1995

Tyrosine 147 of cytochrome b is required for efficient electron transfer at the ubihydroquinone oxidase site (Qo) of the cytochrome bc1 complex.

In Rhodobacter capsulatus, tyrosine (Y) 147 is a highly conserved residue of the cyt b subunit of the bc1 complex. It is located in the vicinity of residues altered in spontaneous inhibitor resistant mutants that affect the ubihydroquinone oxidase (Qo) site of this enzyme. In this work, Y147 was substituted with phenylalanine (F), valine (V), serine (S), and alanine (A) using site-directed mutagenesis in an effort to investigate its specific role in the Qo site. Of the four mutants obtained, Y147S and Y147A exhibited very low ubihydroquinone:cyt c reductase activities and were unable to support photosynthetic growth (Ps) while Y147F and Y147V were Ps+. In all mutants, no changes in the redox midpoint potentials (Em7) of the cyt bH and cyt bL, the occupancy of the Qo site by Q/QH2, and the flash-induced reverse electron transfer kinetics from Qi to cyt bH were observed. On the other hand, rates of electron transfer from Qo to cyt bH were mildly reduced (2-3-fold) in Y147F and V but dramatically decreased (about 20-fold) in Y147A and S, localizing the defect to the Qo site. Thus, Y147A and S are members of a novel class of Qo site mutants that affect the Qo site catalysis without perturbing the accessibility or binding of the substrate. Additional insight to the role of Y147 on ubihydroquinone oxidation was gained by analyzing the Ps+ revertants of these mutants. Two pseudorevertants contained a second mutation [isoleucine (I) or valine (V)] at the highly conserved M154 position, six residues away from Y147.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1995

Arsenic speciation by micellar liquid chromatography with inductively coupled plasma mass spectrometric detection.

Four environmentally and biologically important arsenic species, dimethylarsenic acid (DMA), monomethylarsonic acid (MMA), As(III) and As(V) are separated by micellar liquid chromatography. Linear dynamic ranges for the four species are three orders of magnitude and detection limits are in the picogram range with inductively coupled plasma mass spectrometric (ICP-MS) detection. This paper discussed in detail the development of the chromatographic conditions. The micellar mobile phase, which consisted of 0.05 M cetyltrimethylammonium bromide, 10% propanol and 0.02 M borate buffer, showed good compatibility with ICP-MS. This method allowed direct injection of urine samples onto the chromatographic system without extensive pretreatment and presented no interference from chlorine in the matrix. Detection limits are comparable with other LC-ICP-MS studies. An SRM urine sample was used to demonstrate the applicability of this technique to "real-life" situations. Results indicated that DMA, MMA and As(V) were present in the urine sample.

Arsenicals↗