BMC Cancer 2005, 5:104 http://www.biomedcentral.com/1471-2407/5/104 cEFmexiaglplurlSiernAseesEsi1ocnelolsf, GnoIRr Kin1aidnesnoomcaerSciCnLoCmaceClllalirnaecsebllupthneontoitnypneor- Expression of GIRK1 in some SCLC cell lines but not in normal SAE cells, nor in adenocarcinoma Clara cell phenotype cell lines. Top panel: GIRK1 is expressed in WBA, H69, and H146 SCLC cell lines but not in H187, H209 and H526 SCLC cell lines. GIRK1 is also not expressed in normal SAE cells, nor is it expressed in H322 or H441 adenocarcinoma Clara cell lines. Bottom panel: Cyclophilin, used as a positive reaction control was seen in all samples. For all gene expression experiments, negative control reac- tions were performed and found to be negative. The bands on the agarose gels were consistent with the expected sizes: GIRK1-441 bp; cyclophilin-216 bp; M-100 bp. Table 1: Expression of GIRK2 or GIRK4 in small cell lung cancer cell lines. Expression of either GIRK2 or GIRK4 was determined by real time PCR. Threshold values (CT) values are listed below. Cell Line WBA H69 H146 H187 H209 H526 GIRK2 19.82 23.54 NF NF 24.53 27.55 GIRK4 30.56 32.18 40.54 31.60 24.64 23.21 NF – No CT value calculated due to lack of expression. Gene expression was determined by one sample. No comparisons can be made between CT values for the samples. H2170, H226, and H520 (Figure 4). GIRK2 is only expressed in the H520 cell line, but not in either H2170 or H226 cell lines (Figure 4). Although the predominant GIRK heterotetramers seem to be GIRK1/2 and GIRK1/4 [reviewed in [30]], GIRK3 expression may also be important in lung cancer. GIRK3 expression in normal cells, SCLC cell lines and NSCLC cell LFaigckuroef G2IRK1 and 2 expression in hamster PNEC cells Lack of GIRK1 and 2 expression in hamster PNEC cells. Top panel: Pulmonary neuroendocrine cells were iso- lated from fetal hamsters on the 15th day of gestation. Nei- ther GIRK1 nor GIRK2 was expressed in normal PNEC. Bottom panel: Tissue was isolated from adult hamster brain to determine if the lack of expression in PNEC was not due to species differences in PCR primers. Both GIRK1 and GIRK2 were expressed in hamster brain. Cyclophilin, used as a positive reaction control was seen in the samples. For all gene expression experiments, negative control reactions were performed and found to be negative. The bands on the agarose gels were consistent with the expected sizes: GIRK1-441 bp; GIRK2-438 bp; cyclophilin-216 bp, M-100 bp. lines was examined. GIRK3 expression was seen in all six SCLC cell lines (Figure 5). GIRK3 was expressed in normal cells (SAEC, PNEC) and in the five NSCLC cell lines (A549, H727, H2170, H226, H520) (Figure 6). A sum- mary of all the above experiments indicating gene expres- sion data for the cell lines and normal cells is shown in Table 2. We also wanted to determine if GIRK1 was expressed in fresh surgical tissue samples. A limited number of samples of pulmonary adenocarcinomas (three) or pulmonary squamous carcinomas (two) were Page 4 of 10 (page number not for citation purposes)