This study investigated the volume-regulated anion channel (VRAC) of human cervical cancer SiHa cells under various culture conditions, testing the hypothesis that the progression of the cell cycle is accompanied by differential expression of VRAC activity. Exponentially growing SiHa cells expressed VRACs, as indicated by the presence of large outwardly rectifying currents activated by hypotonic stress with the anion permeability sequence I- > Br- > Cl-. VRACs were potently inhibited by tamoxifen with an IC50 of 4.6 [mu]M. Fluorescence-activated cell sorting (FACS) experiments showed that 59 +/- 0.5, 5 +/- 0.5 and 36 +/- 1.1% of unsynchronized, exponentially growing cervical cancer SiHa cells were in G0/G1, S and G2/M stage, respectively. Treatment with aphidicolin (5 [mu]M) arrested 88 +/- 1.4% of cells at the G0/G1 stage. Arrest of cell growth in the G0/G1 phase was accompanied by a significant decrease of VRAC activity. The normalized hypotonicity-induced current decreased from 48 +/- 5.2 pA pF-1 at +100 mV in unsynchronized cells to 15 +/- 2.6 pA pF-1 at +100 mV in aphidicolin-treated cells. After removal of aphidicolin, culturing in medium containing 10% fetal calf serum triggered a rapid re-entry into the cell cycle and a concomitant recovery of VRAC density. Pharmacological blockade of VRACs by tamoxifen or NPPB caused proliferating cervical cancer cells to arrest in the G0/G1 stage, suggesting that activity of this channel is critical for G1/S checkpoint progression. This study provides new information on the functional significance of VRACs in the cell cycle clock of human cervical cancer cells.
The stiffness sensing ability is required to respond to the stiffness of the matrix. Here we determined whether normal cells and cancer cells display distinct mechanical phenotypes. Cancer cells were softer than their normal counterparts, regardless of the type of cancer (breast, bladder, cervix, pancreas, or Ha-RasV12-transformed cells). When cultured on matrices of varying stiffness, low stiffness decreased proliferation in normal cells, while cancer cells and transformed cells lost this response. Thus, cancer cells undergo a change in their mechanical phenotype that includes cell softening and loss of stiffness sensing. Caveolin-1, which is suppressed in many tumor cells and in oncogene-transformed cells, regulates the mechanical phenotype. Caveolin-1-upregulated RhoA activity and Y397FAK phosphorylation directed actin cap formation, which was positively correlated with cell elasticity and stiffness sensing in fibroblasts. Ha-RasV12-induced transformation and changes in the mechanical phenotypes were reversed by re-expression of caveolin-1 and mimicked by the suppression of caveolin-1 in normal fibroblasts. This is the first study to describe this novel role for caveolin-1, linking mechanical phenotype to cell transformation. Furthermore, mechanical characteristics may serve as biomarkers for cell transformation.
This study was aimed to identify the expression and the correlation of insulin-like growth factor-1 (IGF-1) system and their prognostic impacts in cervical cancer. Seventy-two patients with early-stage cervical cancer were eligible. We obtained the serum levels of total IGF-1 and IGF binding protein-3 (IGFBP-3) by enzyme-linked immunosorbent assay and the expression of IGF-1 receptor (IGF-1R) in cancerous tissue by immuno-fluorescent (IF) stains. The 5-year recurrence-free and overall survival rates were significantly lower (P ¼ 0.003 and P ¼ 0.01, respectively) among patients with high-grade expression of tissue IGF-1R, compared with those with low-grade expression. After adjustment for other factors, preoperative serum total IGF-1 or IGFBP-3 levels failed to predict cancer death and recurrence. High-grade expression of IGF-1R and elevated preoperative squamous cell carcinoma antigen level were independent predictors of both death and recurrence, and combination of both factors could further help identify the subgroup of patients at higher death risk. The IF staining indicates the colocalisation of IGF-1 and IGF-1R in the cancerous tissues, whereas the IGF-1R expression is not correlated with circulating levels of IGF-1 or IGFBP-3. In early-stage cervical cancer, IGF-1 system may have a paracrine or autocrine function and the adverse impacts on prognosis by IGF-1R overexpression are implicated.
The in vitro effects of four antiepileptic drugs (AEDs) on human sperm motility were studied with a transmembrane migration method. Sperm motility of epileptic patients receiving chronic AED therapy was also investigated. Sperm motility was measured immediately after semen had been mixed with AED and after a 2-h preincubation at 37 degrees C. Both in vitro and in vivo studies demonstrated that AEDs inhibited sperm motility. When the drug effect was evaluated after the semen-AED mixture had been preincubated for 2 h, sperm motility was inhibited to 50% of control at concentrations of 1.59, 4.23, and 5.00 mM for phenytoin, carbamazepine, and valproate, respectively. Both with and without preincubation, phenobarbital, even up to 12.92 mM, did not inhibit the motility to less than 50% of the control. In the in vivo study, poor sperm motility was noted in epileptic patients with long-term AED therapy despite serum levels within the therapeutic range. Shorter duration of activity of spermatozoa was also observed in these patients. Interference with sperm membrane function by AEDs may be the underlying mechanism.
Squamous cell carcinoma antigen (SCCA) is a tumor marker for patients with squamous cell carcinoma of uterine cervix, lung, and esophagus. It was encoded by two highly homologous genes, SCCA1 and SCCA2. However, the relevance of SCCA genes to squamous cell carcinogenesis and patient outcome remains far from clear. In this study, by using laser microdissection and real-time quantitative polymerase chain reaction procedures, the messenger RNA (mRNA) expression of the SCCA1 and SCCA2 genes in normal, dysplastic, and malignant squamous epithelia from uterine cervical tissues were analyzed and correlated with outcome of cancer patients. We found that the SCCA2/A1 mRNA ratios were progressively increased from normal, dysplastic, to cancer cells, and the mean ratio was significantly higher in cancer tissues than that in normal epithelium (P= 0.02). The SCCA2/A1 mRNA ratios were not significantly associated with types of human papillomavirus infection (P > 0.05). High SCCA2/SCCA1 mRNA ratios (ratio >1) were an independent predictor of disease recurrence (relative risk: 3.58; P= 0.003). Of the 38 patients with cervical cancer, 12 patients with high SCCA2/SCCA1 mRNA ratios had a significant lower 2-year disease-free survival of only 50%, while it was 92% in those with low SCCA2/SCCA1 mRNA ratios (P < 0.001). In conclusion, our study indicated that the ratios of SCCA2 to SCCA1 RNA were increased during the process of cervical carcinogenesis, and patients with elevated SCCA2/A1 ratio carried a higher risk for recurrence in early-stage uterine cervical cancer.
The clinical significance of STIM proteins and Orai Ca2+ channels in tumor progression has been demonstrated in different types of cancers. Podosomes are dynamic actin-rich cellular protrusions that facilitate cancer cell invasiveness by degrading extracellular matrix. Whether STIM1-dependent Ca2+ signaling facilitates cancer cell invasion through affecting podosome formation remains unclear. Here we show that the invasive fronts of cancer tissues overexpress STIM1, accompanied by active store-operated Ca2+ entry (SOCE). Interfering SOCE activity by SOCE inhibitors and STIM1 or Orai1 knockdown remarkably affects podosome rosettes formation. Mechanistically, STIM1-silencing significantly alters the podosome rosettes dynamics, shortens the maintenance phase of podosome rosettes and reduces cell invasiveness. The subsequently transient expression of STIM1 cDNA in STIM1-null (STIM1−/−) mouse embryo fibroblasts rescues the suppression of podosome formation, suggesting that STIM1-mediated SOCE activation directly regulates podosome formation. This study uncovers SOCE-mediated Ca2+ microdomain that is the molecular basis for Ca2+ sensitivity controlling podosome formation.
Hypotonicity activates volume-sensitive Cl- currents, which are implicated in the regulatory volume decrease (RVD) responses and transport of taurine in human cervical cancer HT-3 cells. In this study, the role of cytoskeleton in the regulation of volume-sensitive Cl- channels and RVD responses in HT-3 cells was studied. Cells were incubated with various compounds, which depolymerized or polymerized cytoskeletal elements, i.e. actin filaments and microtubules. The hypotonicity-induced changes in Cl- conductance and in cell volume were measured by whole-cell voltage clamping and cell size monitoring, respectively. Our results show that in HT-3 cells hypotonicity activated an outward rectified Cl- current that was abrogated by Cl- channel blockers. Cytochalasin B, an actin-depolymerizing compound, induced a substantial increase in Cl- conductance under isotonic condition and potentiated the expression of Cl- currents in hypotonic stress. Phorbol 12-myristate 13-acetate (PMA) significantly inhibited the cytochalasin B-induced activation of Cl- conductance under isotonic condition. On the other hand, treatment with cytochalasin B significantly prolonged the RVD responses. Phalloidin, a stabilizer of actin polymerization, did not change the basal currents under isotonic condition, but completely abolished the increase in whole-cell Cl- conductance elicited by hypotonicity and retarded the cell volume recovery. Colchicine, a microtubule-assembly inhibitor, had no effect on either basal Cl- conductance or volume-sensitive Cl- current and was unable to inhibit the RVD responses. Taxol, a microtubule-stabilizing compound, did not alter the basal Cl- conductance, but inhibited the activation of volume-sensitive Cl- channels as well as the process of RVD in a dose-dependent manner. These data support the notion that functional integrity of actin filaments and microtubules plays critical roles in maintaining the RVD responses and activation of Cl- channels in human cervical cancer HT-3 cells.
Objectives To characterize intratumoral vascularization in early-stage cervical cancer by three-dimensional (3D) power Doppler ultrasound.Methods One hundred and forty-one patients with carcinoma of the uterine cervix and 30 normal controls were studied by transvaginal 3D power Doppler ultrasound. The tumor volume of the cervical cancer was determined. VI, FI and VFI: localized, peripheral, scattered and single-vessel The blood flow within the tumor or normal cervix was measured and expressed as the vascularization index (VI), flow index (FI) and vascularization flow index (VFI). Results
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