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IntroductionBrain tumors (BTs) are perceived as one of the most common malignancies among children. The specific regulation of each gene can play a critical role in cancer progression. The present study aimed to determine the transcripts of the TSGA10 and GGNBP2 genes, considering the alternative 5′UTR region, and investigating the expression of these different transcripts in BTs.Material and methodsPublic data on brain tumor microarray datasets in GEO were analyzed with R software to evaluate the expression levels of TSGA10 and GGNBP2 genes (the Pheatmap package in R was also used to plot DEGs in a heat map). In addition, to validate our in-silico data analysis, RT-PCR was performed to determine the splicing variants of TSGA10 and GGNBP2 genes in testis and brain tumor samples. The expression levels of splice variants of these genes were analyzed in 30 brain tumor samples and two testicular tissue samples as a positive control.ResultsIn silico results show that the differential expression levels of TSGA10 and GGNBP2 were significant in the GEO datasets of BTs compared to normal samples (with adjusted p-value 1). This study’s experimental results showed that the TSGA10 gene produces four different transcripts with two distinct promoter regions and splicing exon 4. The relative mRNA expression of transcripts without exon 4 was higher than transcripts with exon 4 in BT samples (p-valueGGNBP2, exon 2 in the 5′UTR region and exon 6 in the coding sequence were spliced. The expression analysis results showed that the relative mRNA expression of transcript variants without exon 2 was higher than other transcript variants with exon 2 in BT samples (p-valueConclusionThe decreased expression levels of transcripts with longer 5′UTR in BT samples than in testicular or low-grade brain tumor samples may decrease their translation efficiency. Therefore, decreased amounts of TSGA10 and GGNBP2 as potential tumor suppressor proteins, especially in high-grade brain tumors, may cause cancer development by angiogenesis and metastasis.

This study highlights the cytotoxic effect of three L. casei strains on colorectal cell lines in invitro conditions. Different concentrations of live, heat killed (HK) and cell free supernatant (CFS) of three L.casei strains were subjected to CaCo2 and MRC5 cell lines. The viability of the treated and untreated cells was determined after 72 hrs by MTT assay, and IC50 estimated. Apoptosis was evaluated by Annexin V-propidium iodide method using flow cytometry. The live, HK and CFS of the L. casei strains showed cytotoxic effects on colorectal cell lines with significant differences. The cytotoxicity effects of live cells on CaCo2 cells were significantly higher (p˂0.01) than the HK cells. A dose dependent response was observed, as higher concentrations resulted in enhanced cytotoxicity effects. Live L.casei 1296-2cells inhibited 91% of CaCo2 cell growth, with IC50 of less than 108 cfu/ml. MRS medium and concentrations of CFS at above 20% v/v, were cytotoxic to the normal cell lines. Flow cytometry analyses of L. casei 1296-2 indicated that cytotoxicity effects on CaCo2 cells is related to apoptotic induction. Invitro studies indicate that Live and CFS of L. casei 1296-2 might be promising candidate for the control of colorectal cancers.

Additional file 1: Table S1. The sequences of DNA oligos. Figure S1. SDS-PAGE gel electrophoresis (12%) was applied to illustrate the efficiency of the loading, before and after adding AGu@PEG1500-PMO at different concentrations (including 30, 60, 120, and 240 nM) of Cas9. Cas9 concentrations were quantified with image j software. Figure S2. SDS-PAGE gel electrophoresis (12%) of a) RNP released from RNP@AGu@PEG1500-PMO at 0, 2, 4, 6, 12, and 24 h (lane 1-6), and known free RNP samples (lane a-d). Figure S3. SDS-PAGE (12%) of purified Cas9. Figure S4. Agarose gel electrophoresis (1%) of purified sgRNA. Figure S5. a) The fluorescence microscopy images with GFP filter, and b) bright field fluorescence microscopy images of the colony formed by expansion of single cell for 16, 48, and 120 h. Scale bar: 200 µm. Figure S6. Agarose gel electrophoresis (1%) of Cas9 activity assay using GFP-PCR product from pLenti6.3-To-V5-Dest- GFP (645 bp) as substrate, a) released-RNP from RNP@AGu@PEG1500-PMO complex digest the PCR product at 7.5 (lane 1), b) free-RNP, and c) digest the PCR product at 7.5 and 5 pH (lane 4 and lane 5, respectively).

Background: This study aimed to evaluate the diagnostic value of outer dense fiber 4 (ODF4), melanoma-associated antigen A3 (MAGEA3), and MAGEAB4 mRNAs in transitional cell carcinoma (TCC), using a small amount of cell reverse transcriptase-polymerase chain reaction (RT-PCR) on urinary exfoliated cells. Methods: We recruited a total of 105 suspected TCC patients and 54 sex- and age-matched non-TCC controls. The candidates’ genetic expression patterns were investigated with RT-PCR, while reverse transcription quantitative PCR was applied to quantify and compare each mRNA level between cases and control groups. Results: The sensitivity of ODF4, MAGEA3, and MAGEAB4 RT-PCR was 54.8%, 63%, and 53.4%, whereas the specificity was 73.7%, 86%, and 94.7%, respectively. Combining ODF4, MAGEA3, and MAGEAB4 RT-PCR offered a relatively higher sensitivity (83.6%). Conclusion: RT-PCR with ODF4, MAGEA3, and MAGEAB4 on urinary exfoliated cells could provide clinicians with a promising method to improve TCC diagnosis, especially in the case of gross hematuria and catheterization. The method used here is non-invasive, simple and convenient, and unlike cytology, it does not rely directly on expert professional opinions. These features can be of particular importance to the management of TCC patients in whom regular and lifelong surveillance is required.