[Study on Toll-like receptors expression and cytokine production induced by bacillus Calmette-Guerin in human bladder cancer cell].
OBJECTIVE
To investigate the effect of bacillus Calmette-Guerin (BCG) on Toll-like receptors (TLRs) expression and cytokine production in human bladder cancer cell line T24.


METHODS
Human transitional carcinoma cell line T24 was cultured in RPMI1640 medium with 10% FBS, BCG was added into the cell culture with various doses in bacteria-cell ratio. After T24 cells were stimulated by BCG for 1 hour, total RNA of cells was extracted. RT-PCR procedure was conducted with the primer of TLR-2 and TLR-4, and the products were analyzed with agarose gels electrophoresis. Then the expression of TLR-2 and TLR-4 in T24 cells was assessed by the analyzing system of gel imaging. After the stimulation culture for 12 hours, the supernatant of cells was collected. The levels of IL-4 and IL-12 in each sample were assayed by ELISA method.


RESULTS
The expression level of either TLR-2 or TLR-4 was increased by the stimulation of BCG in a dose-dependent manner, the effects reached the maximal level at the dose of BCG as 10 bacilli per cell. The production of IL-12 in T24 cells was also increased gradually by the stimulation of BCG in a dose-dependent manner, and the dose of BCG obtained maximal effect was 10 bacilli per cell, which is coincident with the results observed in the expression of TLRs. There was no difference showed on the production of IL-4 between T24 cells stimulated with BCG and control.


CONCLUSIONS
The stimulation of BCG not only up-regulated the expression of TLR-2 and TLR-4, but also increased the production of IL-12 in bladder cancer T24 cell line. The expression of TLRs and the production of cytokine in bladder cancer cells may be related to the BCG-induced immunol response to human bladder cancer.
Li
Xiang
X
Department of Urology, Huaxi Hospital of Sichuan University, Chengdu 610041, China.
Gong
Zhi-yong
ZY
Li
Hong
H
Wei
Qiang
Q
Shi
Ming
M
Yang
Yu-ru
YR
D001500:BCG Vaccine / Q000494:pharmacology*; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D004587:Electrophoresis, Agar Gel; D004797:Enzyme-Linked Immunosorbent Assay; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D007376:Interleukin-2 / Q000032:analysis; D015847:Interleukin-4 / Q000032:analysis; D008562:Membrane Glycoproteins / Q000235:genetics*; D011956:Receptors, Cell Surface / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4; D051193:Toll-Like Receptors; D001749:Urinary Bladder Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology
D001500:BCG Vaccine; D007376:Interleukin-2; D008562:Membrane Glycoproteins; D011956:Receptors, Cell Surface; C495342:TLR2 protein, human; C495345:TLR4 protein, human; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4; D051193:Toll-Like Receptors; D015847:Interleukin-4