Biomedical subjects

Chen-Ou Zhang

Publications and source records attributed to Chen-Ou Zhang.

8 recordsLinked to original sources

PubMed2006

CKAP4/p63 is a receptor for the frizzled-8 protein-related antiproliferative factor from interstitial cystitis patients.

Antiproliferative factor (APF) is a low molecular weight sialoglycopeptide that is secreted by bladder cells from interstitial cystitis patients and is a potent inhibitor of both normal bladder epithelial and bladder carcinoma cell proliferation. We hypothesized that APF may produce its antiproliferative effects by binding to a transmembrane receptor. This study demonstrates that cytoskeleton-associated protein 4/p63 (CKAP4/p63), a type II transmembrane receptor, binds with high affinity to APF. The antiproliferative activity of APF is effectively inhibited by preincubation with anti-CKAP4/p63-specific antibodies, as well as by short interfering RNA knockdown of CKAP4/p63. Immunofluorescent confocal microscopy showed co-localization of anti-CKAP4/p63 and rhodamine-labeled synthetic APF binding in both cell membrane and perinuclear areas. APF also inhibits the proliferation of HeLa cervical carcinoma cells that are known to express CKAP4/p63. These data indicate that CKAP4/p63 is an important epithelial cell receptor for APF.

Amino Acid Sequence↗

PubMed2005

APF, HB-EGF, and EGF biomarkers in patients with ulcerative vs. non-ulcerative interstitial cystitis.

BACKGROUND: Interstitial cystitis (IC) is a chronic bladder disorder, with symptoms including pelvic and or perineal pain, urinary frequency, and urgency. The etiology of IC is unknown, but sensitive and specific biomarkers have been described, including antiproliferative factor (APF), heparin-binding epidermal growth factor-like growth factor (HB-EGF), and epidermal growth factor (EGF). However, the relative sensitivity of these biomarkers in ulcerative vs. nonulcerative IC is unknown, and these markers have yet to be validated in another laboratory. We therefore measured these markers in urine from patients with or without Hunner's ulcer, as well as normal controls, patients with bladder cancer, and patients with bacterial cystitis, at the First Hospital of China Medical University. METHODS: Urine specimens were collected from two groups of Chinese IC patients (38 IC patients with Hunner's ulcers, 26 IC patients without Hunner's ulcers), 30 normal controls, 10 bacterial cystitis patients and 10 bladder cancer patients. APF activity was determined by measuring 3H-thymidine incorporation in vitro, and HB-EGF and EGF levels were determined by ELISA. RESULTS: APF activity (inhibition of thymidine incorporation) was significantly greater in all IC patient urine specimens than in normal control specimens or in specimens from patients with bacterial cystitis or bladder cancer (p < 0.0001 for each comparison). Urine HB-EGF levels were also significantly lower and EGF levels significantly higher in both groups of IC patients than in the three control groups (p < 0.0001 for each comparison). Although APF and HB-EGF levels were similar in ulcerative and nonulcerative IC patients, EGF levels were significantly higher in IC patients with vs. without ulcers (p < 0.004). CONCLUSION: These findings indicate that APF, HB-EGF and EGF are good biomarkers for both ulcerative and nonulcerative IC and validate their measurement as biomarkers for IC in Chinese patients.

Adult↗

PubMed2005

Regulation of tight junction proteins and bladder epithelial paracellular permeability by an antiproliferative factor from patients with interstitial cystitis.

PURPOSE: Previous reports have suggested that the bladder epithelial barrier may be compromised in interstitial cystitis (IC). Antiproliferative factor (APF) is a small glycoprotein made specifically by bladder epithelial cells in patients with IC that induces changes in expression of certain epithelial cell proteins and profoundly inhibits cell growth. Therefore, we confirmed the increased permeability and decreased tight junction formation of bladder epithelial cell monolayers grown from biopsies in patients with IC compared to cells from normal controls. We then determined the effect of APF on the permeability of normal bladder epithelial cell monolayers and the expression of tight junction proteins. MATERIALS AND METHODS: Permeability was determined by measuring the C-mannitol and H-inulin flux between cells in confluent monolayers on Transwell culture plates (Corning, Corning, New York). Tight junction formation was assessed by immunofluorescence microscopy and the expression of specific proteins was determined by Western blot. RESULTS: APF treatment caused significant increases in the paracellular permeability of normal bladder epithelial cell monolayers and the attenuation of tight junctions compared to mock APF, similar to changes seen in IC cells. APF treatment also decreased expression of the tight junction proteins zonula occludens-1 and occludin. CONCLUSIONS: Because of its apparent effects on bladder epithelial cell tight junctions and paracellular permeability in vitro, APF may contribute to the leakiness of the bladder epithelial barrier seen in IC.

Adolescent↗

PubMed2004

An antiproliferative factor from interstitial cystitis patients is a frizzled 8 protein-related sialoglycopeptide.

Approximately 1 million people in the United States suffer from interstitial cystitis, a chronic painful urinary bladder disorder characterized by thinning or ulceration of the bladder epithelial lining; its etiology is unknown. We have identified a glycosylated frizzled-related peptide inhibitor of cell proliferation that is secreted specifically by bladder epithelial cells from patients with this disorder. This antiproliferative factor (APF) profoundly inhibits bladder cell proliferation by means of regulation of cell adhesion protein and growth factor production. The structure of APF was deduced by using ion trap mass spectrometry (MS), enzymatic digestion, lectin affinity chromatography, and total synthesis, and confirmed by coelution of native and synthetic APF derivatives on microcapillary reversed-phase liquid chromatography (microRPLC)/MS. APF was determined to be an acidic, heat-stable sialoglycopeptide whose peptide chain has 100% homology to the putative sixth transmembrane domain of frizzled 8. Both synthetic and native APF had identical biological activity in normal bladder epithelial cells and T24 bladder cancer cells. Northern blot analysis indicated binding of a probe containing the sequence for the frizzled 8 segment with mRNA extracted from cells of patients with interstitial cystitis but not controls. APF is therefore a frizzled-related peptide growth inhibitor shown to contain exclusively a transmembrane segment of a frizzled protein and is a potential biomarker for interstitial cystitis.

Antineoplastic Agents↗

PubMed2004

Interstitial cystitis antiproliferative factor (APF) as a cell-cycle modulator.

BACKGROUND: Interstitial cystitis (IC) is a chronic bladder disorder of unknown etiology. Antiproliferative factor (APF), a peptide found in the urine of IC patients, has previously been shown to decrease incorporation of thymidine by normal bladder epithelial cells. This study was performed to determine the effect of APF on the cell cycle of bladder epithelial cells so as to better understand its antiproliferative activity. METHODS: Explant cultures from normal bladder biopsy specimens were exposed to APF or mock control. DNA cytometry was performed using an automated image analysis system. Cell cycle phase fractions were calculated from the DNA frequency distributions and compared by two-way analysis of variance (ANOVA). RESULTS: APF exposure produced statistically significant increases in the proportion of tetraploid and hypertetraploid cells compared to mock control preparations, suggesting a G2 and/or M phase cell cycle block and the production of polyploidy. CONCLUSIONS: APF has a specific effect on cell cycle distributions. The presence of a peptide with this activity may contribute to the pathogenesis of interstitial cystitis through disruption of normal urothelial proliferation and repair processes.

Adult↗

PubMed2004

Antiproliferative factor, heparin-binding epidermal growth factor-like growth factor, and epidermal growth factor in men with interstitial cystitis versus chronic pelvic pain syndrome.

OBJECTIVES: To determine whether men with chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS) have urine markers previously described for patients with interstitial cystitis (IC; presence of antiproliferative factor [APF] activity, decreased levels of heparin-binding epidermal growth factor-like growth factor [HB-EGF], and increased levels of epidermal growth factor). METHODS: Clean catch urine specimens were collected from 41 symptomatic patients with CP/CPPS, 36 asymptomatic men without bladder disease who served as the control group, and 24 men with IC. APF activity was determined by (3)H-thymidine incorporation into primary normal adult human bladder epithelial cells. HB-EGF and epidermal growth factor levels were determined by enzyme-linked immunosorbent assay. RESULTS: Men with CP/CPPS did not differ significantly from asymptomatic controls for any of the three markers tested (P >0.49). In contrast, APF activity was present significantly more often and HB-EGF levels were significantly lower in the urine specimens from men with IC than in the specimens from controls or patients with CP/CPPS (P <0.00001 for all four comparisons). Although the epidermal growth factor levels also tended to be higher in the urine from patients with IC than in the urine from controls, the difference did not reach statistical significance (P = 0.06). CONCLUSIONS: These findings indicate that at least two of the urine biomarkers previously identified in women with IC (presence of APF activity and decreased levels of HB-EGF) are also found in men with IC, but not in men with CP/CPPS. This finding suggests that IC and CP/CPPS may be two different disorders with distinct pathophysiologies. It also confirms the utility of the presence of APF activity and HB-EGF levels as markers for IC in men, as well as in women, with this disorder.

Adolescent↗

PubMed2003

Changes in human bladder epithelial cell gene expression associated with interstitial cystitis or antiproliferative factor treatment.

Explanted bladder epithelial cells from patients with interstitial cystitis (IC) have been shown to differ from explanted control cells in several ways, including production of an antiproliferative factor (APF), altered production of certain epithelial growth factors, and rate of proliferation. To better understand the role of the APF in abnormal bladder epithelial cell proliferation in IC, we studied gene expression patterns in normal bladder epithelial cells treated with APF vs. mock APF and compared them to expression patterns in IC vs. normal cells using microarray analysis. Oligo-dT-primed total cellular RNA was labeled with [(33)P]dCTP and hybridized to GeneFilter GF211 microarray membranes (Research Genetics) containing cDNA for 3,964 human genes. Thirteen genes that function in epithelial cell proliferation or differentiation were consistently differentially expressed in both IC (compared with control) and APF-treated (compared with mock APF-treated) normal bladder epithelial cells. The general pattern of gene expression in IC and APF-treated cells suggested a less proliferative phenotype, with increased expression of E-cadherin, phosphoribosylpyrophosphate synthetase-associated protein 39, and SWI/SNF complex 170-kDa subunit, and decreased expression of vimentin, alpha2-integrin, alpha1-catenin, cyclin D1, and jun N-terminal kinase 1; these findings were confirmed for the structural gene products (E-cadherin, vimentin, alpha2-integrin, and alpha-catenin) by immunohistochemistry. These results are compatible with the previously noted decreased proliferation rate of IC and APF-treated normal cells, and indicate that the mechanism whereby APF inhibits cell proliferation may involve both downregulation of genes that stimulate cell proliferation along with upregulation of genes that inhibit cell growth.

Adolescent↗

PubMed2003

Decreased in vitro proliferation of bladder epithelial cells from patients with interstitial cystitis.

OBJECTIVES: To determine whether explanted bladder epithelial cells from patients with interstitial cystitis (IC) display intrinsically decreased rates of proliferation in vitro, and to compare the growth rates of untreated IC and normal bladder cells with the rates of normal cells treated with a purified antiproliferative factor (APF) at levels found in urine from patients with IC. METHODS: Epithelial cell explants were prepared from the bladder biopsies of 4 patients with IC and 2 asymptomatic controls. Cell proliferation was determined by serial counting of trypan blue-negative cells. APF and mock APF were purified chromatographically, and activity was determined by (3)H-thymidine incorporation into primary normal bladder epithelial cells. Heparin-binding epidermal growth factor-like growth factor and epidermal growth factor were measured by enzyme-linked immunosorbent assay. RESULTS: Bladder epithelial cells from patients with IC proliferated significantly less than did control cells by day 2 after serum starvation (P = 0.02). Similar inhibition of the proliferation rate was seen in control cells treated with APF; APF-induced changes in heparin-binding epidermal growth factor-like growth factor, but not epidermal growth factor, production by cells were associated with changes in growth rates. CONCLUSIONS: The proliferation rate of explanted bladder epithelial cells from patients with IC in serum-free medium was significantly less than that of control cells, indicating an intrinsic abnormality in IC cell proliferation. This abnormality may be caused by APF, which induces reversible inhibition of heparin-binding epidermal growth factor-like growth factor production and normal bladder epithelial cell proliferation.

Adult↗