Involvement of CFTR and CAII in bacterial killing <i>in vitro</i>.

<p>(A)When 1×10⁴ CFU of <i>E.coli</i> was inoculated to the apical compartment of the rat prostate epithelial cells for 18 h, there was no bacterial activity detected in the culture medium. 10 µM CFTR_inh-172 (A), 1∶500 CFTR antibody (B) or 50 µM acetazolamide (C) were added with 1×10⁵<i>E.coli</i> to block CFTR or CAII activity and their effect on bacterial activity 18 hours after incubation was shown. (**P<0.01, ***P<0.001).</p

LPS-induced upregulation of cytokines, CFTR and CAII expression in rat prostate epithelial cells.

<p>Primary rat prostate epithelial cells were treated with 1 µg/ml <i>E.coli</i>-LPS for 24 h. The expression levels of IL-6, IL-1β, TNF-α, CFTR and CAII were evaluated by RT-PCR (A, B) and GAPDH was used as control. Data were from three experiments. (C) <i>E.coli</i>-LPS up-regulated the protein expression of CFTR (160 kD) and CAII (29 kD)as detected by western blot, with β-tubulin (55 kD) used as the loading control. Data were from three experiments. (*P<0.05, **P<0.01, ***P<0.001).</p

Expression of CFTR and CAII in human hyperplasia prostate with inflammation.

<p>CFTR (A, B) and CAII (C, D) were detected in human hyperplasia tissues. There was lymphocytes infiltration (Yellow arrow, B, D) in the inflamed area of the clinical prostate hyperplasia samples. Note that the expression of CFTR and CAII was stronger in the area with lymphocytes infiltration (Red arrow, B, D) than those without infiltration (Red arrow, A, C). (E) Negative control. Scale bar: 50 µm.</p

Solutions for different concentration of HCO₃⁻.

<p>Solutions for different concentration of HCO₃⁻.</p

Bacterial killing effect of CFTR <i>in vivo</i> and upregulation of cytokines, CFTR and CAII in <i>E coli</i>-infected rat prostate.

<p>(A) Comparison of <i>E coli</i> bacterial activities recovered from rat prostatitis models without or with CFTR_inh-172 (10 µM). Each point indicates the bacterial CFU per gram of prostate tissue weight (***P<0.001). (B) <i>E.coli</i> up-regulated the expression of cytokine genes, CFTR and CAII in rat prostate as determined by RT-PCR. Data were from three experiments. (C) Expression of CFTR (160 kD) and CAII (29 kD) protein was significantly up-regulated in <i>E.coli</i> -infected rat prostate as determined by western blot. Data were from three experiments. (*P<0.05, **P<0.01, ***P<0.001).</p

HCO₃⁻ but not pH exhibits bactericidal capacity <i>in vitro</i>.

<p>(A). The activity of <i>E.coli</i> was inhibited by 80 mM HCO₃⁻ and 50 mM HCO₃⁻. (B) Insignificant effect on bacterial activities of varied pH (at constant 25 mM HCO₃⁻) at 7.35, 7.95, 8.14 and 8.24 which was corresponding with the pH value of different concentration of HCO₃⁻. Data were from three experiments. (**P<0.01vs 0 mM HCO₃⁻, ^$$$p<0.001 vs 25 mM HCO₃⁻,^##P<0.01vs 50 mM HCO₃⁻, ^&P<0.05 vs 25 mM HCO₃⁻).</p

Primers and RT-PCR conditions.

<p>Primers and RT-PCR conditions.</p

Expression of CFTR in rat prostate epithelial cells.

<p>(A) Immunohistochemical staining of CFTR in SD rat prostate with negative control in the absence of primary antibody. CFTR was expressed at the apical surface of rat ventral prostate epithelium. Scale bar: 100 µm. (B) CFTR transcript was detected by RT-PCR in cultured rat prostate epithelial cells with predicted amplification products at 481 bp. (C) CFTR protein was detected in rat prostate epithelial cells by Western blotting which recognizes a band at MW 160 kDa. (D) Phase contrast image (left) and immunofluorescence staining of cytokeratin 5&8 (middle, green) or CFTR (right, green) in rat prostate epithelial cells. Cell nuclei were counterstained with DAPI (blue). Scale bar: 100 µm.</p

Effect of HCO₃⁻ and pH on expression of <i>E. coli</i> initiation factors IF1, IF2 and IF3.

<p>(A) The mRNA expression of initiation factors was significantly inhibited by 80 mM HCO₃⁻. (B) Alkaline pH with a constant HCO₃⁻ of 25 mM did not suppress IF1, IF2 and IF3 gene expression. 16 s was used as control. Data were from three experiments. (*P<0.05).</p

Involvement of CFTR in mediating cAMP-stimulated bicarbonate secretion by rat prostate epithelial cells.

<p>(A) The pH_i recovered quickly after cellular alkalinization induced by removing bicarbonate/CO₂ from perfusate in the presence of Cl⁻. (B) The rate of pH_i recovery was markedly attenuated when extracellular Cl⁻ was removed from the perfusate. (C) In the absence of Cl⁻, forskolin (forsk, 10 µM) stimulated pH_i recovery. (D) The forskolin-induced pHi recovery could be blocked by NPPB (100 µM). The scales in A–D are the same. (E) Summary of pH_i recovery rates under different conditions after cellular alkalinization induced by removing bicarbonate/CO₂ from perfusate. (*P<0.05).</p