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Biomedical subjects

Q X Zhang

Publications and source records attributed to Q X Zhang.

At least 37 records · Page 2Linked to original sources
PubMed1997

An estrogen receptor mutant with strong hormone-independent activity from a metastatic breast cancer.

Thirty tumors from metastatic breast cancer patients were screened for mutations in the estrogen receptor (ER) gene using single-strand conformation polymorphism and sequence analysis. Three missense mutations, Ser47Thr, Lys531Glu, and Tyr537Asn, were identified in these lesions. To investigate these mutated ERs or altered transcriptional activation function, expression vectors containing wild-type (wt) and mutant ERs were constructed and cotransfected with different estrogen response element reporter gene constructs into HeLa cells and MDA-MB-231 human breast cancer cells. The first two ER mutants were similar to wt ER. However, the Tyr537Asn ER mutant possessed a potent, estradiol-independent transcriptional activity, as compared to wt ER. Moreover, the constitutive activity of the Tyr537Asn ER mutant was virtually unaffected by estradiol, tamoxifen, or the pure antiestrogen ICI 164,384. Tyr537 is located at the beginning of exon 8 in the COOH-terminal portion of the hormone-binding domain of the ER, to which dimerization and transcription activation functions have also been ascribed. It has been identified as a phosphorylation site implicated in hormone binding, dimerization, and hormone-dependent transcriptional activity. Our results suggest that the Tyr537Asn substitution induces conformational changes in the ER that might mimic hormone binding, not affecting the ability of the receptor to dimerize, but conferring a constitutive transactivation function to the receptor. If present in other metastatic breast tumors, this naturally occurring ER mutant may contribute to breast cancer progression and/or hormone resistance.

Breast Neoplasms↗
PubMed1997

Vascular endothelial growth factor is the major angiogenic factor in omentum: mechanism of the omentum-mediated angiogenesis.

Omentum has been used clinically to promote wound healing and to stimulate the revascularization of ischemic tissues. The biologic mechanism responsible for these effects has, however, not yet been defined. A number of polypeptide growth factors that possess potent angiogenic properties have recently been identified, and we therefore sought to determine whether one of these growth factors might be responsible for the angiogenic properties of the omentum. The levels of vascular endothelial growth factor (VEGF) protein in a number of rat tissues and organs were analyzed by Western and enzyme immunoassay analysis. Because omentum was found to have the greatest VEGF concentrations of the tissues examined, antibody neutralization, transcription inhibition assays, and Northern blot analysis were performed under hypoxic and normoxic conditions on tissues extractions and primary tissue cultures of omentum to further characterize the functional significance of VEGF expression in these tissues. The omentum demonstrated the highest VEGF secretion rate as well as the highest concentration of VEGF protein of the various rat tissues and organs examined. Fractionation studies of the omentum furthermore demonstrated that omental adipocytes, rather than the stromal-vascular cells, were the primary source of VEGF protein. An endothelial cell mitogenic assay showed that a major portion of the mitogenic activity of heparin-binding proteins and conditioned media derived from omentum was abolished by VEGF antibody. Additional studies with the transcription inhibitor actinomycin-D furthermore demonstrated that the VEGF gene was continuously transcribed in the rat omental adipocytes. Incubation of the omental adipocytes under hypoxic conditions induced approximately a 1.7-fold increase in VEGF protein expression, which was abolished by actinomycin-D. Northern blot analysis demonstrated that hypoxia resulted in upregulation of the VEGF mRNA in the hypoxia-cultured omental adipocytes, suggesting that the augmentation of VEGF expression in omental adipocytes by hypoxia occurs at the transcriptional level. These data suggest that VEGF is the major angiogenic factor produced by omentum and possibly underlies the mechanism of omentum-induced angiogenesis. Augmented expression of VEGF by omental cells under hypoxic conditions may furthermore reflect the mechanism responsible for enhancing the angiogenic activity of omentum in the setting of ischemia.

Adipose Tissue↗
PubMed1997

Therapeutic angiogenesis: a comparative study of the angiogenic potential of acidic fibroblast growth factor and heparin.

PURPOSE: Acidic fibroblast growth factor (aFGF) is a potent mitogen for vascular and other mesenchymal cells in vitro that can induce angiogenesis in vivo. Although heparin has no mitogenic potential of its own, it is an important aFGF cofactor in vitro and may also be capable of stimulating angiogenesis. Because the development of a collateral vasculature in response to ischemia appears to be dependent on angiogenesis, we compared the abilities of aFGF with or without heparin and heparin alone to accelerate angiogenesis in a rat hind limb ischemia model. METHODS: Daily subcutaneous injections of saline solution (1 ml), heparin (0.05 mg), or human recombinant aFGF with or without heparin (1 microgram aFGF, 0.05 mg heparin) were administered into the hind limb region distal to the point of unilateral femoral artery ligation in the rat for the 10 days immediately after vascular occlusion. Angiogenicity was determined by histologic assessment of treatment outcomes. RESULTS: Histologic assessment of the number of vessels per microscopic field 10 days after vascular ligation in the fibrofatty tissues distal to the ligation point had the following results: saline solution, 10 +/- 4 vessels; heparin, 13 +/- 4 vessels (p < 0.05 vs saline solution); aFGF, 26 +/- 8 vessels; and aFGF/heparin 36 +/- 8 vessels (aFGF, aFGF/ heparin, p < 0.001 vs saline solution). Similar increases in vascularization were also noted in the skeletal muscle tissues distal to the vascular ligation point. Immunohistochemical analysis for the presence of proliferating cell nuclear antigen, a marker for mitogenic activity, demonstrated corresponding increases in proliferating cell nuclear antigen labeling for each of the treatment groups, expressed as a percentage of total vascular cell nuclei, as follows: saline solution, 7% +/- 2%; heparin, 21% +/- 8% (p < 0.05 vs saline solution); aFGF, 67% +/- 9%; and aFGF/heparin, 83% +/- 5% (aFGF, aFGF/heparin, p < 0.001 vs saline solution). CONCLUSIONS: The increased vascularization and mitogenic activity demonstrated by these respective studies suggest that angiogenesis is significantly accelerated by the administration of heparin alone and is accelerated to a greater extent by the administration of aFGF with or without heparin. The aFGF/heparin regimen may represent an optimal means of augmenting collateral vessel growth to relieve ischemia in the clinical setting.

Animals↗
PubMed1996

Controlled overexpression of selected domains of the P85 subunit of phosphatidylinositol 3-kinase reverts v-Ha-Ras transformation.

Selected domains of the regulatory p85 subunit of phosphatidylinositol 3-kinase have been expressed under the control of the tetracycline transactivator in NIH 3T3 fibroblasts transformed by the v-Ha-Ras oncogene. The domains expressed were the SH3 domain, the BCR homology domain, the region between the two SH2 domains which contains the p110 binding site (the inter SH2 (IS) domain), and the C-terminal (CT) domain (containing both SH2 domains and the IS domain). The levels of IS or SH3 domain expressed in the presence of tetracycline were sufficient to reverse the transforming effects of v-Ha-Ras, and no further inhibition of proliferation was observed when expression was increased 7-fold by removal of tetracycline. In contrast inhibition of proliferation by the CT domain was observed only when the level of expression was increased 5-fold by removal of tetracycline. Overexpression of the BCR domain of p85 had no effect on v-Ha-Ras transformation. Expression of the IS domain disrupted the interaction of the p85 regulatory subunit with the p110 catalytic subunit. These results indicate that the association of the p85 subunit of PI 3-kinase with the p110 subunit is necessary for v-Ha-Ras-induced transformation in NIH 3T3 cells.

3T3 Cells↗
PubMed1996

Multiple splicing variants of the estrogen receptor are present in individual human breast tumors.

Transcript variants of the estrogen receptor (ER) were investigated in 109 primary breast tumors using reverse transcription-polymerase chain reaction (RT-PCR) and primers allowing analysis of each internal exon. A high incidence of different ER variants was observed, each individual tumor often manifesting multiple variants coexisting with corresponding wildtype (wt) ER. These variants, by sequence analysis confirmed to represent exon splicing deletions, included ER deltaE2 (found in 41% of the tumors examined), ER deltaE3 (74%), ER deltaE4 (72%), ER deltaE5 (66%), and ER deltaE7 (88%). No evidence of transcripts lacking exon 6 was found, although a ER deltaE5,7 variant manifesting simultaneous deletion of exons 5 and 7 was observed. The presence of specific ER variants was not significantly correlated to the status of ER and progesterone receptor (PgR) protein expression, as assessed by routine analysis, although a trend towards a higher incidence of ER deltaE3 and increased expression of ER deltaE7 in ER+/PgR- tumors was observed, suggesting a dominant inhibitory effect on normal ER function to be involved. Moreover, ER deltaE4 was more common in ER+ tumors, possibly due to a cytoplasmic sequestring of this variant lacking a nuclear localization sequence. The presence of ER variants was not associated to clinicopathological variables, and equally frequent in tumors from patients having recurred or remained recurrence-free during adjuvant tamoxifen therapy. In conclusion, although ER splicing variants are abundant in breast cancer, the present study provides no evidence for a direct role of these ER variants in tumor development and tamoxifen resistance. It remains possible, however, that minor cell clones within the tumor, undetected by analysis of tumor homogenates, displaying extreme difference in content of ER variants, could be selected for during therapy or metastasis.

Adult↗
PubMed1996

Calcium binds dynamin I and inhibits its GTPase activity.

Synaptic vesicle recycling is a neuronal specialization of endocytosis that requires the GTPase activity of dynamin I and is triggered by membrane depolarization and Ca2+ entry. To establish the relationship between dynamin I GTPase activity and Ca2+, we used purified dynamin I and analyzed its interaction with Ca2+ in vitro. We report that Ca2+ bound to dynamin I and this was abolished by deletion of dynamin's C-terminal tail. Phosphorylation of dynamin I by protein kinase C promoted formation of a dynamin I tetramer and increased Ca2+ binding to the protein. Moreover, Ca2+ inhibited dynamin I GTPase activity after stimulation by phosphorylation or by phospholipids but not after stimulation with a GST-SH3 fusion protein containing the SH3 domain of phosphoinositide 3-kinase. These results suggest that in resting nerve terminals, phosphorylation of dynamin I by protein kinase C converts it to a tetramer that functions as a Ca(2+)-sensing protein. By binding to Ca2+, dynamin I GTPase activity is specifically decreased, possibly to regulate synaptic vesicle recycling.

Animals↗
PubMed1995

Reduction in platelet-derived growth factor receptor mRNA in v-src-transformed fibroblasts.

The status of the platelet-derived growth factor (PDGF) receptor in normal rat kidney (NRK) fibroblasts and in NRK fibroblasts transformed by the v-src oncogene or the polyoma middle T (pmt) antigen has been compared. v-src-NRK cells have 7-fold fewer surface binding sites for PDGF than NRK cells, but the affinity of the residual receptors for PDGF is reduced only 2-fold. Levels of the PDGF receptor measured by Western blotting or in an autophosphorylation assay in vitro are 8- and 4-fold lower respectively in v-src-NRK cells than in NRK cells. No PDGF-induced phosphorylation of the PDGF receptor is apparent after 32P-labelling of intact v-src-NRK cells, implying that the reduction in PDGF receptor levels is not a consequence of production of autocrine PDGF. A 10-fold reduction in the amount of mRNA for the PDGF receptor is also observed in v-src-NRK cells. No decrease in PDGF receptor protein or mRNA levels is observed in pmt-NRK cells. We conclude that levels of the PDGF receptor in v-src-transformed NRK fibroblasts are modulated by reduction in the level of PDGF receptor mRNA.

Animals↗
PubMed1995

Effect of tumor necrosis factor-alpha on a cell line transformed by a secreted form of human fibroblast growth factor-1 gene and on its parental cell line.

In the present study we characterized the responses of the cell line HST-alpha, stably transformed by a secreted form of human fibroblast growth factor-1 (acidic FGF) gene, and its parental NIH 3T3 cell line to recombinant murine (rMu) TNF-alpha. Treatment of HST-alpha cells with rMu TNF-alpha can significantly reduce the number of foci formed in the in vitro transformation assay. In the presence of the RNA polymerase inhibitor actinomycin-D, the transformed HST-alpha cell is more susceptible to the cytotoxicity of rMu TNF-alpha than is its parental NIH 3T3 cell. The median lethal concentrations (LC50) of rMu TNF-alpha to HST-alpha cell and NIH 3T3 cell were 0.35 ng/ml and 4.56 ng/ml respectively. These results demonstrated that the introduction of a single oncogene or a growth factor gene to a cell can change the cell's response to cytokines.

3T3 Cells↗
PubMed1994

Structures of the human cDNA and gene encoding the 78 kDa gastrin-binding protein and of a related pseudogene.

The nucleotide sequence encoding the human 78 kDa gastrin binding protein (GBP) has been deduced from overlapping fragments generated by the polymerase chain reaction with oligonucleotides based on the sequence of the porcine GBP (Mantamadiotis, T. et al. (1993) Biochim. Biophys. Acta 1170, 211-215) and cDNA from the colonic carcinoma cell line LIM 1215 as template. The mature human GBP is 90% identical to the porcine GBP. Clones encoding the human GBP gene, which contains 19 exons, have been isolated from human genomic libraries. The positions of the exon/intron junctions are completely different from the junctions in the gene encoding the related peroxisomal trifunctional enzyme. Clones encoding a related pseudogene have also been isolated and sequenced.

Amino Acid Sequence↗
PubMed1994

Platelet-derived growth factor induces phosphatidylinositol 3-kinase release from the middle T-pp60c-src complex and association with the platelet-derived growth factor receptor.

Both platelet-derived growth factor (PDGF) and epidermal growth factor (EGF) induce mitogenesis in normal rat kidney (NRK) fibroblasts transformed by the polyoma virus middle T (pmt) oncogene. In unstimulated pmt-NRK cells phosphatidylinositol 3-kinase forms a complex with the middle T protein and pp60c-src. PDGF treatment causes a release of phosphatidylinositol 3-kinase activity from the complex and a simultaneous increase in activity associated with the PDGF receptor. In contrast after treatment with EGF the majority of phosphatidylinositol 3-kinase activity remains associated with the middle T-pp60c-src complex. Proliferation of NRK fibroblasts transformed by the v-src oncogene is already maximal, and no further stimulation is observed with either PDGF or EGF. Neither growth factor induces dissociation of the complex between phosphatidylinositol 3-kinase and pp60v-src. These observations suggest that the complex between phosphatidylinositol 3-kinase, the middle T protein and pp60c-src is dissociable, and that phosphatidylinositol 3-kinase plays different roles in mitogenic signal transduction by the PDGF and EGF receptors.

Animals↗
PubMed1994

[A study on the radionuclide method for assessment of right ventricular function in COPD with cor pulmonale].

Right ventricular ejection fraction (RVEF), right ventricular peak filling rate (RVPFR) and right atrial early diastolic emptying rate (RAER) were measured with radionuclide gated blood pool scintigraphy in 19 healthy subjects and 15 cases of COPD with cor pulmonale and right heart catheterization was performed in the latter group. It was shown that RVEF in the group of cor pulmonale patients with pulmonary arterial hypertension (PAH) was significantly lower than that of the group of healthy subjects and cor pulmonale patients without PAH (P < 0.001, P < 0.001), no difference in RVEF was found between cor pulmonale patients without PAH and healthy subjects. As pulmonary arterial pressure increased, RAER and RVPFR decreased gradually, and the reduction of RAER and RVPFR occurred earlier than that of RVEF. It is suggested that right ventricular diastolic function may be impaired before right ventricular systolic function.

Adult↗
PubMed1993

An exon 5 deletion variant of the estrogen receptor frequently coexpressed with wild-type estrogen receptor in human breast cancer.

Recent evidence suggests that the expression of estrogen receptor (ER) variants in breast cancer may interfere with wild-type (wt) ER function and be related to tumor progression and resistance to hormone treatment. One of these variants, ER delta E5, lacking that part of the hormone-binding domain encoded by exon 5, has previously been identified in breast tumors with the unusual estrogen receptor negative (ER-) and progesterone receptor positive (PgR+) phenotype and found to possess constitutive and hormone-independent transcriptional activity. Using a ribonuclease protection assay, we analyzed 27 breast tumors and 4 breast cell lines for the presence of this variant. We found the ER delta E5 variant to be expressed, not only in all of three ER-/PgR+ tumors but also in 19 of 20 ER+/PgR+ or ER+/PgR- tumors. Moreover, the variant was always coexpressed with and often in excess of wtER. ER delta E5 was also found in three breast cancer cell lines (MCF7, T47D, and ZR75-1), although to a lesser extent than wtER. A complete absence of both ER delta E5 and wtER was noted in four ER-/PgR- tumors and one normal breast cell line (HBL-100). Thus, our data suggest that the occurrence of ER delta E5 in breast cancer may represent a critical stage in tumor progression to autonomy.

Breast Neoplasms↗
PubMed1993

Human rheumatoid factor cross-idiotypes. IV. Studies on WA XId-positive IgM without rheumatoid factor activity provide evidence that the WA XId is not unique to rheumatoid factors and is distinct from the 17.109 and G6 XIds.

The WA cross-idiotype (XId) is the major XId among human monoclonal rheumatoid factors (mRF) and is almost always associated with the light (L) chain XId, 17.109, and the heavy (H) chain XId, G6. A cell line, 35G6, was cloned that bears the WA XId, but shows no reactivity with immunoglobulin G (IgG) and is negative for the 17.109 and G6 XIds. The 35G6 L chain appears to be derived from the same VKIII-JKI genes as most WA mRFs L chains. In contrast to the WA mRFs H chains in which VH1 genes are used, the 35G6 IgM expresses a VH3 gene. Sequence comparisons with other WA XId-positive mRF suggested several common structural features that may be related to the WA XId and differences that may relate to lack of IgG reactivity. Cells similar to 35G6 have previously been described in pokeweed mitogen-stimulated cell lines of peripheral blood lymphocytes from normal individuals and patients with rheumatoid arthritis and type II mixed cryoglobulinemia. These observations were confirmed, and in addition, it was shown that the majority of WA XId-positive cells in these cultures were negative for the 17.109 and G6 XIds. The presence of the WA XId in the absence of IgG reactivity suggests that the WA XId is more directly associated with an antigen specificity other than IgG, and its association with RF activity may be incidental. It is postulated that these WA XId-positive RF-negative antibodies may serve a physiologic role as natural antibodies to a pervasive pathogen, and that IgG reactivity is a consequence of somatic diversification accompanying proliferation of the WA XId-positive RF-negative cell.

Amino Acid Sequence↗
PubMed1993

Responses of human fetal splenocytes and thymocytes to interleukin-2 and comparison with adult peripheral blood lymphocytes.

By means of cytotoxicity and thymidine uptake assays, the effect of human recombinant interleukin-2 (rIL-2) on the induction of lymphokine-activated killer (LAK) cell activity and cellular proliferation in splenocytes and thymocytes from human fetuses (18-22 weeks) was studied and compared with that in mononuclear cells from adult peripheral blood. It was shown that the fetal splenocytes and thymocytes incubated with low doses of rIL-2 (10-100 U/ml) developed a broad spectrum of antitumor activity (LAK cytotoxicity), although the kinetics and magnitudes of the responses were different. It was also suggested that LAK precursors were present in fetal spleen and thymus. Furthermore, rIL-2 induced a stronger proliferative and cytotoxic response in splenocytes than in thymocytes. The human fetal LAK cells isolated from the spleen were found to be 10-20 times more potent than those from adult peripheral blood with regard to cell proliferation and cytotoxicity in vitro.

Adult↗