Paracrine Factors Released from Tonsil-Derived Mesenchymal Stem Cells Inhibit Proliferation of Hematological Cancer Cells Under Hyperthermia in Co-culture Model.

Mesenchymal stem cells (MSCs) are promising biological therapeutic candidates in cancer treatment. As a source of MSCs, palatine tonsil tissue is one of the secondary lymphoid organs that form an essential part of the immune system, and the relation between the secondary lymphoid organs and cancer progression leads us to investigate the effect of tonsil-derived MSCs (T-MSC) on cancer treatment. We aimed to determine the anti-tumoral effects of T-MSCs cultured at the febrile temperature (40 °C) on hematological cancer cell lines. The co-culture of cancer cells with T-MSCs was carried out under fever and normal culture conditions, and then the cell viability was determined by cell counting. In addition, apoptosis rate and cell cycle arrest were determined by flow cytometry. We confirmed the apoptotic effect of T-MSC co-culture at the transcriptional level by using real-time polymerase chain reaction (RT-PCR). We found that co-culture of cancer cells with T-MSCs significantly decreased the viable cell number under the febrile and normal culture conditions. Besides, the T-MSC co-culture induced apoptosis on K562 and MOLT-4 cells and induced the cell cycle arrest at the G2/M phase on MOLT-4 cells. The apoptotic effect of T-MSC co-culture under febrile stimulation was confirmed at the transcriptional level. Our study has highlighted the anti-tumoral effect of the cellular interaction between the T-MSCs and human hematological cancer cells during in vitro co-culture under hyperthermia.

Conjugated linoleic acid strengthens the apoptotic effect of cisplatin in A549 cells.

One of the chemotherapeutic agents widely used in the treatment of non-small cell lung cancer (NSCLC) is cisplatin. However, the resistance of cancer cells to cisplatin and additionally serious side effects from cisplatin limit its use. Conjugated linoleic acid (CLA) has been shown to suppress the development of carcinogenesis in vitro and in vivo studies and has antitumoral activity in many cancers. The study aimed to investigate the potential effect of using cisplatin, the first-line treatment for NSCLC, in combination with CLA to increase its efficacy in low-dose use. MTT cytotoxicity assay was performed to determine the effects of CLA in combination with cisplatin on cell viability of NSCLC cell lines. The apoptotic effect of this combination on NSCLC cell lines and cell cycle distribution was determined by flow cytometry. At the same time, apoptosis and cell cycle-related gene expression levels were determined by Real-Time PCR. Combination treatment of low-dose cisplatin with CLA resulted in a significant decrease in cell viability compared to cisplatin alone, and an increase in the rate of apoptotic cells was observed. While cisplatin caused G1 phase arrest in cancer cells, there was an increase in cell percentages in S and G2 phases after combined application with CLA. In high-dose cisplatin administration, it was observed that the efficiency of the decrease in anti-apoptotic BCL2 expression related to resistance to chemotherapeutic agents was less than that of low-dose cisplatin administration. Combined administration of high-dose cisplatin with CLA significantly recovered BCL2 downregulation.

Hyperthermia-stimulated tonsil-mesenchymal stromal cells suppress hematological cancer cells through downregulation of IL-6.

There are contradictory reports on the use of mesenchymal stromal cells (MSCs) in cancer therapy. Variable outcomes have been associated with several factors including cancer pathology, experimental procedure, MSC source tissue, and individual genetic differences. It is also known that MSCs exert their therapeutic effects with various paracrine factors released from these cells. The profiles of the factors released from MSCs are altered by heat shock, hypoxia, oxidative stress, starvation or various agents such as inflammatory cytokines, and their therapeutic potential is affected. In this study, the antitumor potential of conditioned media (CM), which contains paracrine factors, of mild hyperthermia-stimulated mesenchymal stromal cells derived from lymphoid organ tonsil tissue (T-MSC) was investigated in comparison with CM obtained from T-MSCs grew under normal culture conditions. CM was obtained from T-MSCs that were successfully isolated from palatine tonsil tissue and characterized. The cytotoxic effect of CM on the growth of hematological cancer cell lines at different concentrations (1:1 and 1:2) was demonstrated by methylthiazoldiphenyl-tetrazolium bromide analysis. In addition, the apoptotic effect of T-MSC-CM treatment was evaluated on the cancer cells using Annexin-V/PI detection method by flow cytometry. The pro/anti-apoptotic and cytokine-related gene expressions were also analyzed by real-time polymerase chain reaction post T-MSC-CM treatment. In conclusion, we demonstrated that the factors released from hyperthermia-stimulated T-MSCs induced apoptosis in hematological cancer cell lines in a dose-dependent manner. Importantly, our results at the transcriptional level support that the factors and cytokines released from hyperthermia-stimulated T-MSC may exert antitumoral effects in cancer cells by downregulation of IL-6 that promotes tumorigenesis. These findings reveal that T-MSC-CM can be a powerful cell-free therapeutical strategy for cancer therapy.

TP53 Codon 72 Polymorphism and the Risk of Colorectal Cancer in an Azerbaijani Population.

BACKGROUND: We aimed to evaluate the association between the TP53 Arg72Pro gene polymorphism and risk of colorectal cancer in an Azerbaijani population. METHODS: A total of 141 patients with colorectal cancer and 150 gender- and age-matched controls were involved in the study. The genotypes of the TP53 gene Arg72Pro polymorphism were detected by polymerase chain reaction-based restriction fragment length polymorphism analysis. RESULTS: We found that the heterozygous genotypes Arg/Pro (odds ratio, 1.128; 95% CI, 0.657-1.937) and mutant Pro/Pro (odds ratio, 1.274; 95% CI, 0.648-2.504) were more frequent in colorectal cancer patients compared to healthy controls. The frequency of the mutant Pro allele (odds ratio, 1.122; 95% CI, 0.809-1.554) was revealed in 47.5% of colorectal cancer patients and in 44.7% of healthy controls. There was no association observed between TP53 Arg72Pro polymorphism and risk of colorectal cancer in an Azerbaijani population (P > .05). CONCLUSION: Our findings indicate a lack of relationship between TP53 Arg72Pro polymorphism and risk of colorectal cancer. Furthermore, we have found no statistical differences in the frequency of genotype and allele by sex, age, histological grade, tumor stage, smoking status, and alcohol consumption in this study.

Gene editing and RNAi approaches for COVID-19 diagnostics and therapeutics.

The novel coronavirus pneumonia (COVID-19) is a highly infectious acute respiratory disease caused by Severe Acute Respiratory Syndrome-Related Coronavirus (SARS-CoV-2) (Prec Clin Med 2020;3:9-13, Lancet 2020;395:497-506, N. Engl J Med 2020a;382:1199-207, Nature 2020;579:270-3). SARS-CoV-2 surveillance is essential to controlling widespread transmission. However, there are several challenges associated with the diagnostic of the COVID-19 during the current outbreak (Liu and Li (2019), Nature 2020;579:265-9, N. Engl J Med 2020;382:727-33). Firstly, the high number of cases overwhelms diagnostic test capacity and proposes the need for a rapid solution for sample processing (Science 2018;360:444-8). Secondly, SARS-CoV-2 is closely related to other important coronavirus species and subspecies, so detection assays can give false-positive results if they are not efficiently specific to SARS-CoV-2. Thirdly, patients with suspected SARS-CoV-2 infection sometimes have a different respiratory viral infection or co-infections with SARS-CoV-2 and other respiratory viruses (MedRxiv 2020a;1-18). Confirmation of the COVID-19 is performed mainly by virus isolation followed by RT-PCR and sequencing (N. Engl J Med 2020;382:727-33, MedRxiv 2020a, Turkish J Biol 2020;44:192-202). The emergence and outbreak of the novel coronavirus highlighted the urgent need for new therapeutic technologies that are fast, precise, stable, easy to manufacture, and target-specific for surveillance and treatment. Molecular biology tools that include gene-editing approaches such as CRISPR-Cas12/13-based SHERLOCK, DETECTR, CARVER and PAC-MAN, antisense oligonucleotides, antisense peptide nucleic acids, ribozymes, aptamers, and RNAi silencing approaches produced with cutting-edge scientific advances compared to conventional diagnostic or treatment methods could be vital in COVID-19 and other future outbreaks. Thus, in this review, we will discuss potent the molecular biology approaches that can revolutionize diagnostic of viral infections and therapies to fight COVID-19 in a highly specific, stable, and efficient way.

Novel RET Proto-oncogene variants identified in Turkish patients with thyroid carcinoma.

Thyroid cancer is one of the few malignancies whose incidence is increasing in the last decades. Advances in understanding the molecular mechanisms lead to provide opportunity for prevention, effective early identification and targeted therapies for management. A total of 63 patients with participated in this study Genomic DNA samples were obtained from the samples formalin- embedded tissue and peripheral blood. Following polymerase chain reaction amplification of the 6 RET key exons (10, 11, 13, 14, 15, and 16) were applied and PCR products were subjected to next generation DNA sequencing (ABI 3730). Results revealed that; genotype frequencies were for rs1800961 (G > T) , GG 6 (%9.5), GT 17 (%27) TT40 (%63.5) for rs2472732 (G > A), GG31 (%49.2) GA29 (%46) AA3 (%4.8,) for rs1799939, (G > A) GG42 (%66.7) GA19 (%30.2) AA2 (%3.2), for rs1800962, (C > T) CC54 (%85.7) CT9 (%14.3), for rs1800863 (C > G), CC39 (%61.9) CG22 (%34.9) GG2 (%3.2), for rs3026272 (C > G) CC 13 (%20.6) CG 50 (%79.4). Additionally 15 potential novel genetic variants were identified in these key exons. Detailed information was given both known and new detected variants in supplementary table. Genetic variants distribution frequencies and new variants represented in Turkish thyroid cancer patients for RET proto-oncogene and that results would provide contribution to the literature.

Development of gene editing strategies for human ß-globin (HBB) gene mutations.

Recent developments in gene editing technology have enabled scientists to modify DNA sequence by using engineered endonucleases. These gene editing tools are promising candidates for clinical applications, especially for treatment of inherited disorders like sickle cell disease (SCD). SCD is caused by a point mutation in human ß-globin gene (HBB). Clinical strategies have demonstrated substantial success, however there is not any permanent cure for SCD available. CRISPR/Cas9 platform uses a single endonuclease and a single guide RNA (gRNA) to induce sequence-specific DNA double strand break (DSB). When this accompanies a repair template, it allows repairing the mutated gene. In this study, it was aimed to target HBB gene via CRISPR/Cas9 genome editing tool to introduce nucleotide alterations for efficient genome editing and correction of point mutations causing SCD in human cell line, by Homology Directed Repair (HDR). We have achieved to induce target specific nucleotide changes on HBB gene in the locus of mutation causing SCD. The effect of on-target activity of bone fide standard gRNA and newly developed longer gRNA were examined. It is observed that longer gRNA has higher affinity to target DNA while having the same performance for targeting and Cas9 induced DSBs. HDR mechanism was triggered by co-delivery of donor DNA repair templates in circular plasmid form. In conclusion, we have suggested methodological pipeline for efficient targeting with higher affinity to target DNA and generating desired modifications on HBB gene.

Is there any relationship between human foamy virus infections and familial Mediterranean fever?

BACKGROUND: Familial Mediterranean fever (FMF) is generally defined as an autosomal recessive disease, characterized by the automatic activation of the innate immune system in the absence of a detectable pathogenic stimulant. We hypothesize that the pathogenic factors, besides the genetic causes, may affect the development of FMF symptoms. To test this hypothesis, we examined the effects of human foamy virus (HFV) positivity on the occurrence of the clinical symptoms of FMF. MATERIALS AND METHODS: Two hundred and twenty-two FMF patients with definitive diagnosis according to Tel Hashomer criteria (study group 1 [SG1]), 205 symptomatic FMF patients who had definitive diagnosis according to the same criteria but did not carry any of the 12 most commonly occurring MEFV gene mutations (study group 2 [SG2]), and 200 healthy individuals were included as control group (study group 3 [SG3]) in the study. The genetic analysis was applied in the Molecular Genetics Laboratory of the Department of Medical Biology, Faculty of Medicine, Ondokuz Mayis University. This study was designed as a case-control study. HFV positivity was tested by amplifying the HFV bel1 gene sequence with polymerase chain reaction technique. Statistical analyses were conducted using SPSS version 23.0 software. RESULTS: HFV positivity showed significant differences between the study groups (P = 0.002). While 43 (19.02%) of the 222 SG1 patients were positive for the HFV bel1 gene sequence, 33 (16.09%) of the 205 SG2 patients were positive for the same sequence. Only 15 (7.5%) of the SG3 participants were positive for the presence of HFV bel1 gene sequence. CONCLUSION: The results of our study suggested that HFV positivity can be a stimulant pathogenic factor of natural immune system which can cause the emergence of FMF symptoms.