Immunotherapy: Open access wants to thank the eminent researchers
On the occasion of entering into its 5th anniversary Immunotherapy: Open access wants to thank the most cited research in the journal.
The article entitled “Joint Action of the Nano-sized System of Small Non-coding RNAs with DDMCVector and Recombinant IL-7 Reprograms A-549 Lung Adenocarcinoma Cellsinto CD4+ Cells” by the researcher Oxana V. Klimenko from SID ALEX GROUP, Ltd. Kyselova 1185/2, Prague, Czech Republic.
The abstract of the article is as follows:
Small non-coding RNAs, as class of small regulatory molecules, control normal development and differentiation ofcells. Micro-RNAs and piwi-interacting-RNAs are members of epigenetic regulators family. In previous studiesresearches investigated the role of different small non-coding RNAs in pathophysiology of cancer.
Objective: In this study A-549 lung adenocarcinoma cells were firstly epigenetically reprogrammed andtransformed into CD4+ cells.
Method: I used new non-viral carrier the complex of DDMC vector with antago-miR-155 and piR-30074 for long-lasting transfection of lung cancer cells.
Results: The transformation of A-549 lung cancer cells was proved by morphological and genetic changes(AltAnalyze Platform) in dynamics. I observed CD4+ lymphocytes phenotypic marker and OCT4 marker of pluripotent cells by immunofluorescence microscopy in transformed cells.
The results of the study was the DDMC Vector® (Non-viralTransfection reagent) from Ryujyu science co. was used. The DDMCvector was toxic to cells at a concentration more than 10 μg/ml(Figures 1A and 1B). In my experiments, I used concentration ofDDMC vector 7 mg/ml. The toxicity was decreased after treatmentwith a complex of the DDMC vector and oligonucleotide (F observed ≤F crit). The transfection efficiency of the DDMC vector was lower thanthat particles of poly-N-vinylpyrrolidone (PNVP), which were used inprevious studies; however, the transfection maximum was reached11-14 days faster than that of the PNVP complexes (21 days) (Figure 2E) . DDMC vector based particles in non-toxic concentrationshad greater stability and increased cell transfection with low sncRNAand DDMC vector concentrations. The DDMC vector is stable after sterilization and is easier to use. The addition of oligonucleotidesdecreased the toxic effect of the DDMC vector compared to theaddition of pure DDMC vector in cells (Figure 1). Three weeks after a complex of sncRNAs was added, A-549 cellswere morphologically changed compared to non-treated control cells.In the microscopic photos was observed an increase in number ofsharp large cells (approximately 50 μm in diameter) with single nuclei,displaying different morphological forms. These cells had vesicles ofvarying sizes on the periphery of the cytoplasm with properties ofsuspension cells. Un-stained cells and cells full of DDMC crystals weremicroscopically detected. These cells were large (approximately 60 μmin diameter) and sharp with polymorphic nuclei that took 1/3 – 2/3 ofthe cell volume (Figures 2A-2E). Giant dendritic-like cells withmultiple pseudopodia approximately 90 μm in diameter were alsodetected (Figure 2B, 2D). Gene expression in A-549 cells was changedafter addition of DDMC vector with a-miR-155, DDMC vector with piR-30074 or DDMC vector with a-miR-155 and piR-30074 comparedto the control group (Figures 3A-3C and 4A-B). Differences wereobserved in separate genes’ expression levels. The expression of somegenes was increased on different days, whereas that of other genes wasfully inhibited after the transfection procedure on other to comparedto the control cells. In the control A-549 cells, the expression ofPIWIL1L1, ICOS1B, GITR3A, cKIT1B and KIR2DL1 genes was notdetected. However, the expression of these genes was obtained intransfected cells. It is known that these genes are expressed in T-cellsonly [15-19].The expression levels of HMOX1 and EBI3 were increased morethan two-fold compared controls. The BACH2 gene expression levels.
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