Identification of IL11RA and MELK amplification in gastric cancer by comprehensive genomic profiling of gastric cancer cell lines

AIM To identify common copy number alterations on gastric cancer cell lines.METHODS Four gastric cancer cell lines(ACP02, ACP03, AGP01 and PG100) underwent chromosomal comparative genome hybridization and array comparative genome hybridization. We also confirmed the results by fluorescence in situ hybridization analysis using the bacterial artificial chromosome clone and quantitative real time PCR analysis.RESULTS The amplification of 9p13.3 was detected in all cell lines by both methodologies. An increase in the copy number of 9p13.3 was also confirmed by fluorescence in situ hybridization analysis. Moreover, the interleukin 11 receptor alpha(IL11RA) and maternal embryonic leucine zipper kinase(MELK) genes, which are present in the 9p13.3 amplicon, revealed gains of the MELK gene in all the cell lines studied. Additionally, a gain in the copy number of IL11 RA and MELK was observed in 19.1%(13/68) and 55.9%(38/68) of primary gastric adenocarcinoma samples, respectively. CONCLUSION The characterization of a small gain region at 9p13.3 in gastric cancer cell lines and primary gastric adenocarcinoma samples has revealed MELK as a candidate target gene that is possibly related to the development of gastric cancer.

Reference genes for quantitative RT-PCR data in gastric tissues and cell lines

AIM:To evaluate the suitability of reference genes in gastric tissue samples and cell lines.METHODS:The suitability of genes ACTB,B2M,GAPDH,RPL29,and 18S rRNA was assessed in21 matched pairs of neoplastic and adjacent nonneoplastic gastric tissues from patients with gastric adenocarcinoma,27 normal gastric tissues from patients without cancer,and 4 cell lines using reverse transcription quantitative real-time polymerase chain reaction(RT-qPCR).The ranking of the best single and combination of reference genes was determined by NormFinder,geNorm,BestKeeper,and DataAssist.In addition,GenEx software was used to determine the optimal number of reference genes.To validate the results,the mRNA expression of a target gene,DNMT1,was quantified using the different reference gene combinations suggested by the various software packages for normalization.RESULTS:ACTB was the best reference gene for all gastric tissues,cell lines and all gastric tissues plus cell lines.GAPDH+B2M or ACTB+B2M was the best combination of reference genes for all the gastric tissues.On the other hand,ACTB+B2M was the best combination for all the cell lines tested and was also the best combination for analyses involving all the gastric tissues plus cell lines.According to the GenEx software,2 or 3 genes were the optimal number of references genes for all the gastric tissues.The relative quantification of DNMT1 showed similar patterns when normalized by each combination of reference genes.The level of expression of DNMT1 in neoplastic,adjacent non-neoplastic and normal gastric tissues did not differ when these samples were normalized using GAPDH+B2M(P=0.32),ACTB+B2M(P=0.61),or GAPDH+B2M+ACTB(P=0.44).CONCLUSION:GAPDH+B2M or ACTB+B2M is the best combination of reference gene for all the gastric tissues,and ACTB+B2M is the best combination for the cell lines tested.

Expanding the horizons of clinical applications of proximal humerus locking plates in the lower extremities:A technical note

Pre-contoured anatomical locking plates were designed to address the clinical need of fixing small epiphyseal segments with a larger number of screws.Those plates match the contour and shape of a variety of bones allowing for optimal buttress properties.The aim of this manuscript is to highlight the benefits of applying proximal humerus locking plates in the fixation of lower extremity bones.Although designed for the proximal humerus,the low-profile plate shape and anatomic contour also provides versatile use in certain areas of the lower extremity.This technical narrative highlights the versatile and reliable use of this plate for other anatomical areas than the one to which it has been originally conceived.