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Biomedical subjects

Z Zhou

Publications and source records attributed to Z Zhou.

At least 325 records · Page 18Linked to original sources
PubMed1997

Identification and characterization of a fibroblast growth factor (FGF) binding domain in the cysteine-rich FGF receptor.

Three distinct transmembrane glycoproteins bind fibroblast growth factor (FGF) family members. These include heparan sulfate proteoglycans, the tyrosine kinase-containing FGF receptors (FGFRs), and a cysteine-rich FGF receptor (CFR). The four FGFRs are thought to mediate FGF-signaling events but require the participation of the heparan sulfate proteoglycans to bind FGFs and transduce intracellular signals. However, a number of groups have proposed that FGF action requires events independent of FGFR activation. CFR, a high affinity FGF-binding protein, was first isolated from chicken embryos. To better understand the interactions between CFR and FGFs, we have constructed a series of CFR deletion mutants and CFR fragments. Analysis of these has identified a approximately 200-amino acid domain that constitutes a CFR FGF binding site. A CFR fragment of 450 residues, CFR290-740, binds FGF-2 with an affinity indistinguishable from the full-length molecule, whereas smaller fragments display greatly reduced FGF binding. Although CFR binds heparin with high affinity, an analysis of the heparin-CFR interaction failed to identify a linear sequence containing a heparin binding site. Two types of FGF binding sites were identified: an ionic strength and heparin-independent site that represents FGF binding to CFR290-740 and an additional FGF binding site that is heparan sulfate-dependent and sensitive to high ionic strength. This latter site is likely to bind FGF indirectly via heparan sulfate binding to CFR. FGF-2 peptides that encompass a sequence implicated in FGF-2 binding to FGFRs also block FGF-2 binding to CFR. Our data suggest that binding of FGFs to CFR and FGFRs is mutually exclusive, since the CFR FGF binding site does not require heparan sulfate, and similar regions on FGF-2 interact with both FGFRs and CFR.

Animals↗
PubMed1997

Cysteine-rich FGF receptor regulates intracellular FGF-1 and FGF-2 levels.

The cysteine-rich FGF receptor (CFR) is a 150-kD membrane-associated glycoprotein that specifically binds FGFs. CFR protein is not detectable at the cell surface and immunocytochemistry with anti-CFR antibodies demonstrates that CFR is concentrated in the Golgi apparatus. These data suggest CFR does not function as a plasma membrane FGF receptor. CFR expressed in chinese hamster ovary cells reduces the intracellular accumulation of exogenously applied FGF-1 and FGF-2. A mutant CFR lacking the juxtamembrane, transmembrane and intracellular domains is unable to alter intracellular FGF levels. Mutant CFR is detected throughout the cell, indicating that the domains absent in mutant CFR are required for appropriate subcellular localization and the regulation of intracellular FGF levels. Although the activation of plasma membrane receptors is necessary for cellular responses to FGFs, a requirement for intracellular FGF has also been proposed. The subcellular localization of CFR and its ability to regulate the levels of intracellular FGFs suggests that CFR may be involved in intracellular FGF trafficking and the regulation of cellular responses to FGFs.

Amino Acid Sequence↗
PubMed1997

Identification of a splice site mutation (2789 +5 G > A) associated with small amounts of normal CFTR mRNA and mild cystic fibrosis.

A splicing mutation was identified at the +5 position of the splice donor site of exon 14b of CFTR in CF patients in a consanguineous family that is remarkable for unusually mild disease. Quantitative studies of nasal epithelial mRNA revealed that homozygotes for the spice site mutation produced approximately 4% of the normal amount of normally-spliced CFTR. We propose that this small amount of normally spliced mRNA is associated with synthesis of some normal CFTR protein, and accounts for the mild phenotype. Further characterization of epithelial function and clinical phenotype in patients bearing this form of mutation, termed a type V mutation, will be useful in determining the level of CFTR associated with amelioration of lung disease.

Adult↗
PubMed1997

Quantification of p53 in epithelial ovarian cancer.

OBJECTIVE: The perceived function of wild-type p53 is suppression of cell proliferation. An alteration in the p53 tumor suppressor gene is a common defect in human malignancies. The purpose of this study was to prospectively determine whether p53 expression, as quantified by image analysis, was related to traditional prognostic indicators as well as survival in patients epithelial ovarian cancer. METHODS: Eighty-three consecutive patients with epithelial ovarian cancer had their p53 expression studied by immunohistochemical staining and quantified by image analysis. Unless otherwise noted, p53 expression was reported as the percentage positive nuclear area staining. RESULTS: The mean follow-up was 37 months (median, 30 months; range 24-55 months). In patients with serous carcinomas of the ovary, the mean p53 expression was 29.4%, whereas in patients with other histologies, the mean was 10.5% (P < 0.001). The tumors of patients with stage III or IV tumors stained significantly higher (mean 28. 7%) than the tumors of patients with stage I or II disease (mean 8. 36%) (P < 0.001). The tumors of patients with disease which could be optimally cytoreduced stained significantly lower (mean 23.0%) than the tumors of patients whose disease was unable to be optimally cytoreduced (mean 28.6%) (P = 0.041). Utilizing survival as the endpoint for multivariate analysis, FIGO stage (P = 0.006), p53 expression (P = 0.046), and the level of cytoreduction (P < 0.001) were independent prognostic indicators. CONCLUSION: Image analysis allows quantitative measurements of p53 staining. p53 staining is significantly higher in advanced-stage, high-grade tumors which are unable to be cytoreduced than in early-stage, low-grade tumors which can be optimally cytoreduced. p53 expression is an independent prognostic indicator of survival in patients with epithelial ovarian carcinomas.

Adult↗
PubMed1997

Measurement of apolipoprotein B in various cell lines: correlation between intracellular levels and rates of secretion.

We have standardized simple but sensitive enzyme-linked immunoassays to understand a relationship between intracellular levels and secretion rates of apoB. The assays were based on commercially available antibodies and were specific to human apoB. A monoclonal antibody, 1D1, was immobilized on microtiter wells and incubated with different amounts of low density lipoproteins to obtain a standard curve. Conditioned media were added to other wells in parallel, and the amount of apoB was quantitated from a linear regression curve. To standardize conditions for the measurement of intracellular apoB, cells were homogenized and solubilized with different concentrations of taurocholate. We found that 0.5% taurocholate was sufficient to solubilize all the apoB in HepG2, Caco-2, and McA-RH7777 cells. Next, a standard curve was prepared in the presence of taurocholate and used to determine intracellular levels of apoB in different cell lines. The intracellular levels (pmol/mg cell protein) and the rates of secretion (pmol/mg/h) of apoB100 were positively correlated (r2 = 0.81, P = 0.0009) in HepG2 cells. Furthermore, a positive correlation (r2 = 0.88, P < 0.0001) was found between intracellular and secreted apoB42 in stably transfected McA-RH7777 cells. In contrast, no correlation was observed for human apoB28 and apoB18 in stably transfected cells that were secreted either partially associated or completely unassociated with lipoproteins. These studies indicated that the rate of secretion of lipid-associated apoB, but not the lipid-free apoB, was tightly controlled.

Apolipoproteins B↗
PubMed1997

Characterization of the binding of amyloid-beta peptide to cell culture-derived native apolipoprotein E2, E3, and E4 isoforms and to isoforms from human plasma.

The epsilon 4 allele of apolipoprotein E (apoE, protein; APOE, gene) is a major risk factor for Alzheimer's disease (AD). Genetically, the frequency of the epsilon 4 allele is enriched in early-onset sporadic, late-onset familial, and common late-onset sporadic AD. ApoE is found in the extracellular amyloid-beta (A beta) deposits that are characteristic features of AD. In this study, we examined the interaction between A beta and apoE isoforms. The apoE isoforms used in this study were either produced by stably transfected Chinese hamster ovary cells (CHO) or were from human plasma. We report that when similar concentrations of the apoE isoforms were used, native nonpurified apoE3 from recombinant CHO-derived sources bound A beta, but apoE4 did not. In fact, in our system, binding of recombinant apoE4 to A beta was never detectable, even after incubation for 4 days. Furthermore, using the same assay conditions, native apoE2, like apoE3, binds A beta avidly. Furthermore, when human plasma apoE isoforms are tested in A beta binding experiments, apoE3 bound A beta more avidly than apoE4, and a major apoE/A beta complex (the 40-kDa form) was observed with plasma apoE3 but not apoE4. These data extend our understanding of apoE isoform-dependent binding of A beta by associating apoE2 with efficient apoE/A beta complex formation and demonstrate that native apoE3 (whether recombinant or derived from human plasma) forms sodium dodecyl sulfate-stable apoE/A beta complexes more readily than native apoE4. The different A beta-binding properties of native apoE4 versus native apoE3 provide insight into the molecular mechanisms by which the APOE epsilon 4 allele exerts its risk factor effects in AD.

Amyloid beta-Peptides↗
PubMed1997

A surgical model for determination of true absorption and biliary excretion of manganese in conscious swine fed commercial diets.

Some trace elements, such as Mn, Cu and Zn, are absorbed and quickly resecreted into the gut through the bile. When this occurs, the unabsorbed nutrient and the absorbed and resecreted nutrient may mix in the gut, preventing quantitative calculation of either. We have developed a surgical model that prevents this complication. Pigs (20-40 kg) were fitted with cannulas in the bile duct, lumen of the duodenum, portal vein, ileocolic vein and jugular vein. After recovery for 6-8 d, pigs were given an oral dose of 9.25 mBq of 54Mn. The flow rate of blood past the portal vein was determined by infusion of P-amino hippuric acid into the ileocolic vein. Absorption was quantified by multiplying the concentration of 54Mn in the portal blood by the flow rate. Biliary excretion was determined by quantitative collection of bile, and previously collected bile was reinfused into the gut lumen. Urine and feces were also quantitatively collected. A postoperative time of 6-8 d was sufficient for pigs to recover from the effects of surgery and anesthesia, as assessed by several measures of metabolic function and food and water intake. True absorption was calculated to be 0.5%. 54Mn in the urine and bile began to increase after 4 d. When the pigs were killed after 12 d, only 0.5% of the 54Mn remained in the carcass. Results of this study show that pigs surgically modified by the described procedure can recover fully and can serve as a model to study intestinal absorption and biliary excretion of nutrients. Furthermore, initial studies using 54Mn showed that the model is applicable to studying Mn metabolism and suggest the need for a more detailed study of Mn absorption and biliary excretion.

Animals↗
PubMed1997

An essential function for the phosphate-dependent exoribonucleases RNase PH and polynucleotide phosphorylase.

Escherichia coli cells lacking both polynucleotide phosphorylase (PNPase) and RNase PH, the only known P(i)-dependent exoribonucleases, were previously shown to grow slowly at 37 degrees C and to display a dramatically reduced level of tRNA(Tyr)su3+ suppressor activity. Here we show that the RNase PH-negative, PNP-negative double-mutant strain actually displays a reversible cold-sensitive phenotype and that tRNA biosynthesis is normal. In contrast, ribosome structure and function are severely affected, particularly at lower temperatures. At 31 degrees C, the amount of 50S subunit is dramatically reduced and 23S rRNA is degraded. Moreover, cells that had been incubated at 42 degrees C immediately cease growing and synthesizing protein upon a shift to 31 degrees C, suggesting that the ribosomes synthesized at the higher temperature are defective and unable to function at the lower temperature. These data indicate that RNase PH and PNPase play an essential role that affects ribosome metabolism and that this function cannot be taken over by any of the hydrolytic exoribonucleases present in the cell.

Cold Temperature↗
PubMed1997

Effects of fluid flow on intracellular calcium in bovine articular chondrocytes.

Fluid flow-induced shear stress results in a variety of morphological and metabolic changes in cultured bovine articular chondrocytes (BAC). However, the mechanism by which the flow signal is transduced into a biological response is unknown. Therefore, we investigated the effects of fluid flow on intracellular Ca2+ concentration ([Ca2+]i) in BAC. Cells loaded with fura 2 were exposed to steady and pulsatile (0.5 Hz) flow at 9, 18, and 34 ml/min in a parallel-plate flow chamber. In response to flow, there was a significant and flow rate-dependent increase in the percentage of cells showing a rise in [Ca2+]i, but no effect on the [Ca2+]i response amplitude. There was no significant difference between the [Ca2+]i responses to steady and pulsatile flow. Mean intracellular Ca2+ response values ranged between 26.2 +/- 1.6 (9 ml/min) and 38.0 +/- 6.8 nM (34 ml/min) above basal [Ca2+]i (81.3 +/- 24.1 nM, n = 90). Removal of extracellular Ca2+ or addition of Gd3+ significantly reduced the percentage of cells responding, suggesting that influx of Ca2+, possibly through mechanosensitive channels, contributes to the rise in intracellular Ca2+. Our data suggest fluid flow-induced mobilization of intracellular Ca2+ may contribute to the mechanism by which mechanical loads are transduced by chondrocytes.

Animals↗
PubMed1997

Age-related decreases in stimulatory G protein-coupled adenylate cyclase activity in osteoblastic cells.

In this study we examined parathyroid hormone (PTH)-, forskolin (FSK)-, and cholera toxin (CTX)-stimulated adenosine 3',5'-cyclic monophosphate (cAMP) accumulation in rat osteoblastic cells (ROB) isolated from young (4 mo), mature (12 mo), and old (24-28 mo) male rats. Exposure to PTH increased cAMP accumulation in a concentration-dependent manner in all ROB cells examined. However, the maximum response in ROB from young rats was threefold greater than the maximum response in those from mature and old rats. Exposure to FSK also stimulated cAMP accumulation in a concentration-dependent manner, but there were no significant differences in responsiveness among ROB isolated from young, mature, and old rats. Exposure to CTX resulted in a dramatic concentration-dependent increase in cAMP in ROB from young rats but only a modest increase in ROB from mature and old rats. PTH binding kinetics were similar in ROB from rats in each age group. These data suggest an age-related defect in stimulatory G protein coupling to adenylate cyclase, which contributes to decreased osteoblastic responsiveness to PTH.

Adenylyl Cyclases↗
PubMed1997

Blockage of the HERG human cardiac K+ channel by the gastrointestinal prokinetic agent cisapride.

Cisapride, a gastrointestinal prokinetic agent, is known to cause long Q-T syndrome and ventricular arrhythmias. The cellular mechanism is not known. The human ether-á-go-go-related gene (HERG), which encodes the rapidly activating delayed rectifier K+ current and is important in cardiac repolarization, may serve as a target for the action of cisapride. We tested the hypothesis that cisapride blocks HERG. The whole cell patch-clamp recording technique was used to study HERG channels stably expressed heterologously in HEK293 cells. Under voltage-clamp conditions, cisapride block of HERG is dose dependent with a half-maximal inhibitory concentration of 6.5 nM at 22 degrees C (n = 25 cells). Currents rapidly recovered with drug washout. The onset of block by cisapride required channel activation indicative of open or inactivated state blockage. Block of HERG with cisapride after channel activation was voltage dependent. At -20 mV, 10 nM cisapride reduced HERG tail-current amplitude by 5%, whereas, at + 20 mV, the tail-current amplitude was reduced by 45% (n = 4 cells). At -20 and + 20 mV, 100 nM cisapride reduced tail-current amplitude by 66 and 90%, respectively. We conclude that cisapride is a potent blocker of HERG channels expressed in HEK293 cells. This effect may account for the clinical occurrence of Q-T prolongation and ventricular arrhythmias observed with cisapride.

Cation Transport Proteins↗
PubMed1997

Assembly and secretion of VLDL in nondifferentiated Caco-2 cells stably transfected with human recombinant ApoB48 cDNA.

Intestinal cells secrete apoB48-containing very low density lipoproteins (VLDLs) and chylomicrons for the transport of biliary and dietary lipids. The molecular mechanisms regulating the assembly of intestinal lipoproteins are not known due to a lack of reliable and specific cell culture models. Caco-2 (a human colon carcinoma) cells have been used to study intestinal lipid metabolism. These cells have been shown to secrete both apoB100- and apoB48-containing triglyceride (TG)-rich lipoproteins only after differentiation into enterocyte-like cells. To study lipoprotein assembly in nondifferentiated Caco-2 cells, we stably expressed human recombinant apoB48 cDNA under the control of a constitutive cytomegalovirus promoter. Pulse-chase analysis revealed that the majority (> 50%) of apoB48 synthesized was degraded intracellularly in the presence or absence of oleic acid. Transfected nondifferentiated cells secreted lipoproteins with flotation densities similar to those of plasma HDL or LDL when cultured in serum-free or serum-containing media, respectively. Incubation of cells with media containing serum and oleic acid resulted in the secretion of VLDL-like particles. Secretion of VLDL was inhibited (> 80%) by triacsin C due to > 60% inhibition of oleate-induced TG synthesis. However, inhibition of cholesteryl ester synthesis by 70% with an acyl coenzyme A:cholesterol acyltransferase inhibitor did not affect VLDL secretion. Efficient assembly of lipoproteins usually requires the microsomal TG transfer protein (MTP). The presence of MTP in transfected Caco-2 cells was investigated by measuring TG transfer activity in microsomal fractions. Microsomal fractions had 0.2% TG transfer activity per hour per microgram of protein, which corresponds to 30% to 60% of the MTP activity present in liver-derived cells. To determine whether MTP activity was required for lipoprotein assembly, transfected cells were incubated in the presence of the MTP inhibitor CP-10,447. This compound completely abolished the secretion of apoB. These data show that the transfected cell lines secrete lipoproteins of different densities under different culture conditions and that the assembly of larger VLDL particles requires active TG synthesis and MTP activity. Thus, in nondifferentiated Caco-2 cells, the amount of apoB secreted and not the MTP activity is the limiting factor for lipoprotein assembly.

Apolipoprotein B-48↗
PubMed1997

[The prevalence of hypertension, diabetes mellitus and hyperlipemia among 1369 subjects for physical examination in Changsha].

A survey of the prevalence of hypertension (HT) and diabetes mellitus (DM) and hyperlipemia (HL) was conducted among 1369 subjects for health check in Changsha. The results showed that the prevalence of HT and DM and impaired glucose tolerance (IGT) and HL were 17.2% and 4.6% and 5.9% and 27.4%, respectively. The prevalence of HT and DM and IGT increased with age. The higher the body mass index was, the higher the prevalence of HT and DM and HL was. These findings indicate that the aging and over weight are important risk factors to the prevalence of HT, DM and HL.

Adult↗
PubMed1997

[Discordance of increase in stress proteins in right and left myocardium of perfused isolated rat heart].

Discordance of functional and structural changes in right and left myocardium has been considerably discovered in various physiologically stimulated hearts and patients with cardiopathy, but the reasons for that have not yet been known. In the present study, induction of stress proteins, i.e. heat shock proteins, was analyzed and compared between the left and right myocardium of normoxic and hypoxic perfused isolated rat hearts by the methods of two dimensional electrophoresis and silver staining. The results showed that three Hsp70 isoforms (molecular weight 68, 70 and 72 k mu) with pI ranging from 6.3 to 7.3 were increased distinctly in both right and left myocardium under the conditions of perfusion as mentioned above. Moreover, the amount of the stress proteins increased in the right myocardiums was higher than that in the left myocardium, indicating that the response of right heart to the stimulus of hypoxic or normoxic perfusions is different from the reaction of left heart, and the extent of protection in left and right heart by stress proteins is unlike. In addition, the activity of catalase was found to be obviously declined in all perfused isolated hearts. It was suggested that the different increase in stress proteins may be due to different structure and status of right and left ventricle and the oxidative stress may be one of the important reasons to induce or enhance the synthesis of stress proteins.

Animals↗
PubMed1997

[Transduction of the IL-2 gene into human gastric cancer cell: an experimental study].

We evaluated the possibility of inducing a productive transfer of the IL-2 gene into human gastic cancer cells (GCC) and assessed the phenotypic and proliferative changes generated in the nude mice. The plasmid vector (BMGNeo-IL-2) carrying the human IL-2 gene was used to transduce the SGC-7901 GCC line by the lipofectin reagent. The IL-2 gene was analyzed by Southern blot and productive IL-2 release using IL-2-dependent CTLL. Cytotoxicity of LAK cells was tested by MTT colorimetry. The kinetics of in vitro growth and proliferation of parental and engineered cells were also measured. Parental and IL-2 gene transduced GCC were injected into nude mice. The tumorigenic potential of IL-2 gene-transfected GCC was evaluated by examining their in vivo growth in nude mice. The productive insertion of the IL-2 gene was achieved in SGC-7901. The amounts of IL-2 constitutively released by the engineered neoplastic cells ranged from 20 to 131 U/ml of IL-2 produced from 10(6) cells in 24 hours. Transduction of GCC with IL-2 gene did not modify the morphology and growth rate. IL-2 gene transfected cells demonstrated increased susceptibility to cell killing by LAK cells. IL-2 producing cells lost their tumorigenicity as evidenced by failure to grow in nude mice. The results demonstrate that IL-2 gene can be productively transduced into human gastric cancer cells without modifying their morphology and growth rate and this transduction leads to reduced or abrogated in vivo tumorigenic potential.

Animals↗
PubMed1997

[An experimental study of cellular response on implanted intraocular lens].

OBJECTIVE: To study the pathological changes and morphological features of cellular response on the implanted posterior chamber intraocular lens (IOL) in rabbits and discuss the mechanism of postoperative intraocular inflammatory response. METHODS: 9 adult pigmented rabbits were divided into three groups. IOLs were extracted on days 1, 7 and 14 after IOL implantation. Pathological changes on IOLs were observed with light microscopy and scanning electron microscopy, and the number of inflammatory cells on IOLs were counted. The data were analyzed by using analysis of variance of SAS software. RESULTS: There were inflammatory cells distributed over the entire IOL surface. In the mean time this study had clearly shown that morphological transformation of macrophages into spindle-shaped fibroblast-like cells took place on IOL, and phagocytic particles were in the cytoplasm of fibroblast-like cells, epithelioid cells and macrophages on the IOL. CONCLUSIONS: The phagocytic ability of fibroblast-like cells, epithelioid cells and macrophages is relatively strong, and besides macrophages and eosinophils, there were many lymphocytic clusters on the IOL, suggesting that an active immune response exist on the IOL surface.

Animals↗
PubMed1997

[A study of immunopathology on salivary gland of Sjögren's syndrome].

OBJECTIVE: To understand the mechanism and pathogenesis of Sjögrens syndrome (SS) and to provide theoretical basis for the treatment of SS. METHOD: Salivary gland lymphocytes (SGL) from 20 patients with SS were analyzed by using immunohistochemical avidin-biotin-peroxidase complex (ABC) method. RESULT: A predominance of T cells was found in SGL with the majority belonging to the CD4+(TH), and the CD8+(TS) subset was obviously decreased, while the CD4+/CD8+ ratio was considerably increased. Most lymphocytes expressed HLA-DR antigen in SS. The deficiency of Ts and the overactivity of TH would result in autoantibody production, that would contribute to the reaction of autoimmune. CONCLUSION: The immune hyperfunction at exocrine glands of SS is the direct cause of the pathogenesis of SS. SS is further proved to be an autoimmune disease.

Aged↗