Cell Journal (Yakhteh)
Volume:22 Issue: 3, Autumn 2020

Acute myeloid leukemia (AML) is a clonal disorder of hemopoietic progenitor cells. The Raf serine/threonine (Ser/Thr) protein kinase isoforms including B-Raf and RAF1, are the upstream in the MAPK cascade that play essential functions in regulating cellular proliferation and survival. Activated autophagy-related genes have a dual role in both cell death and cell survival in cancer cells. The cytotoxic activities of arsenic trioxide (ATO) were widely assessed in many cancers. Sorafenib is known as a multikinase inhibitor which acts through suppression of Ser/Thr kinase Raf that was reported to have a key role in tumor cell signaling, proliferation, and angiogenesis. In this study, we examined the combination effect of ATO and sorafenib in AML cell lines.

In this experimental study, we studied in vitro effects of ATO and sorafenib on human leukemia cell lines. The effective concentrations of compounds were determined by MTT assay in both single and combination treatments. Apoptosis was evaluated by annexin-V FITC staining. Finally, mRNA levels of apoptotic and autophagy genes were evaluated using real-time polymerase chain reaction (PCR).

Data demonstrated that sorafenib, ATO, and their combination significantly increase the number of apoptotic cells. We found that the combination of ATO and sorafenib significantly reduces the viability of U937 and KG-1 cells. The expression level of selective autophagy genes, ULK1 and Beclin1 decreased but LC3-II increased in U937.

The expression levels of apoptotic and autophagy activator genes were increased in response to treatment. The crosstalk between apoptosis and autophagy is a complicated mechanism and further investigations seem to be necessary.

Glioblastoma (GBM) is one of the devastating types of primary brain tumors with a negligible response to standard therapy. Repurposing drugs, such as disulfiram (DSF) and metformin (Met) have shown antitumor properties in different cell lines, including GBM. In the present study, we focused on the combinatory effect of Met and DSF-Cu on the induction of apoptosis in U87-MG cells exposed to 6-MV X-ray beams.

In this experimental study, the MTT assay was performed to evaluate the cytotoxicity of each drug, along with the combinatory use of both. After irradiation, the apoptotic cells were assessed using the flow cytometry, western blot, and real-time polymerase chain reaction (RT-PCR) to analyze the expression of some cell death markers such as BAX and BCL-2.

The synergistic application of both Met and DSF had cytotoxic impacts on the U87-MG cell line and made them sensitized to irradiation. The combinatory usage of both drugs significantly decreased the cells growth, induced apoptosis, and caused the upregulation of BAX, P53, CASPASE-3, and it also markedly downregulated the expression of the anti-apoptotic protein BCL-2 at the gene and protein levels.

It seems that the synergistic application of both Met and DSF with the support of irradiation can remarkably restrict the growth of the U87-MG cell line. This may trigger apoptosis via the stimulation of the intrinsic pathway. The combinatory use of Met and DSF in the presence of irradiation could be applied for patients afflicted with GBM.

Bone morphogenetic protein 4 (BMP4) and basic fibroblast growth factor (bFGF) play important roles in embryonic heart development. Also, two epigenetic modifying molecules, 5ˊ-azacytidine (5ˊ-Aza) and valproic acid (VPA) induce cardiomyogenesis in the infarcted heart. In this study, we first evaluated the role of BMP4 and bFGF in cardiac trans-differentiation and then the effectiveness of 5´-Aza and VPA in reprogramming and cardiac differentiation of human adipose tissue-derived stem cells (ADSCs).

In this experimental study, human ADSCs were isolated by collagenase I digestion. For cardiac differentiation, third to fifth-passaged ADSCs were treated with BMP4 alone or a combination of BMP4 and bFGF with or without 5ˊ-Aza and VPA pre-treatment. After 21 days, the expression of cardiac-specific markers was evaluated by reverse transcription polymerase chain reaction (RT-PCR), quantitative real-time PCR, immunocytochemistry, flow cytometry and western blot analyses.

BMP4 and more prominently a combination of BMP4 and bFGF induced cardiac differentiation of human ADSCs. Epigenetic modification of the ADSCs by 5ˊ-Aza and VPA significantly upregulated the expression of OCT4A, SOX2, NANOG, Brachyury/T and GATA4 but downregulated GSC and NES mRNAs. Furthermore, pre-treatment with 5ˊ-Aza and VPA upregulated the expression of TBX5, ANF, CX43 and CXCR4 mRNAs in three-week differentiated ADSCs but downregulated the expression of some cardiac-specific genes and decreased the population of cardiac troponin I-expressing cells.

Our findings demonstrated the inductive role of BMP4 and especially BMP4 and bFGF combination in cardiac trans-differentiation of human ADSCs. Treatment with 5ˊ-Aza and VPA reprogrammed ADSCs toward a more pluripotent state and increased tendency of the ADSCs for mesodermal differentiation. Although pre-treatment with 5ˊ-Aza and VPA counteracted the cardiogenic effects of BMP4 and bFGF, it may be in favor of migration, engraftment and survival of the ADSCs after transplantation.

Currently, application of oncolytic-virus in cancer treatment of clinical trials are growing. Oncolytic-reovirus is an attractive anti-cancer therapeutic agent for clinical testing. Many studies used mesenchymal stem cells (MSCs) as a carrier cell to enhance the delivery and quality of treatment with oncolytic-virotherapy. But, biosynthetic capacity and behavior of cells in response to viral infections are different. The infecting process of reoviruses takes from two-hours to one-week, depends on host cell and the duration of different stages of virus replication cycle. The latter includes the binding of virus particle, entry, uncoating, assembly and release of progeny-viruses. We evaluated the timing and infection cycle of reovirus type-3 strain Dearing (T3D), using one-step replication experiment by molecular and conventional methods in MSCs and L929 cell as control.

In this experimental study, L929 and adipose-derived MSCs were infected with different multiplicities of infection (MOI) of reovirus T3D. At different time points, the quantity of progeny viruses has been measured using virus titration assay and quantitative real-time polymerase chain reaction (qRT-PCR) to investigate the ability of these cells to support the reovirus replication. One-step growth cycle were examined by 50% cell culture infectious dose (CCID50) and qRT-PCR.

The growth curve of reovirus in cells shows that MOI: 1 might be optimal for virus production compared to higher and lower MOIs. The maximum quantity of virus production using MOI: 1 was achieved at 48-hours post-infection. The infectious virus titer became stationary at 72-hours post-infection and then gradually decreased. The virus cytopathic effect was obvious in MSCs and this cells were susceptible to reovirus infection and support the virus replication.

Our data highlights the timing schedule for reovirus replication, kinetics models and burst size. Further investigation is recommended to better understanding of the challenges and opportunities, for using MSCs loaded with reovirus in cancer-therapy.

This study investigated whether short stimulation (30 minutes) of human adipose stem cells (hASCs) with 1,25-dihydroxyvitamin D3 (calcitriol or 1,25-(OH)2VitD3), fitting within the surgical procedure time frame, suffices to induce osteogenic differentiation, and compared this with continuous treatment with 1,25-(OH)2VitD3.

In this experimental study, hASCs were pretreated with/without 10 nM calcitriol for 30 minutes, seeded on biphasic calcium phosphate (BCP), and cultured for 3 weeks with/without 1,25-(OH)2VitD3. Cell attachment was determined 30 minutes after cell seeding. AlamarBlue assay, alkaline phosphatase (ALP) assay, ALP staining, real-time polymerase chain reaction (PCR), and protein assay were used to evaluate the effect of short calcitriol pretreatment on proliferation and osteogenic differentiation of hASCs up to 3 weeks.

Pretreatment with 1,25-(OH)2VitD3 enhanced the attachment of hASCs to BCP by 1.5-fold compared to nontreated cells and increased the proliferation by 3.5-fold at day 14, and 2.6-fold at day 21. In contrast, continuous treatment increased the proliferation by 1.7-fold only at day 14. After 2 weeks, ALP activity was increased by 18.5-fold when hASCs were pretreated with 1,25-(OH)2VitD3 for 30 minutes but increased only 2.6-fold when compared with its continuous counterpart. Moreover, after 14 days, pretreatment resulted in significant upregulation of the osteogenic markers RUNX2 and SPARC by 3.6-fold and 2.2-fold, respectively, while this was not observed upon continuous treatment. Finally, 30 minutes pretreatment of hASCs with 1,25-(OH)2VitD3 increased VEGF189 expression, which may contribute to the process of angiogenesis.

This study is the first research showing that 30 minutes pretreatment of hASCs with 1,25-(OH)2VitD3, not only enhanced cell attachment to the scaffold at seeding time, but also promoted the proliferation and osteogenic differentiation of hASCs more strongly than continuous treatment, suggesting that short pre-treatment with 1,25-(OH)2VitD3 is a promising approach for the regeneration of bones in a one-step surgical procedure.

Despite the effective role of chemotherapy in cancer treatment, several side effects have been reported to date. For instance, Cyclophosphamide (CP) induces deleterious effects on both cancer and normal cells. Royal jelly (RJ) has a lot of beneficial properties, such as anti-oxidant and anti-inflammatory activities. The aim of the present study was to examine the protective effect of RJ against CP-induced thrombocytopenia, as well as bone marrow, spleen, and testicular damages in rats.

In this experimental study, 48 male Wistar rats were divided into six groups (n=8/group); control, CP, RJ (100 mg/kg), RJ (200 mg/kg), RJ (100 mg/kg)+CP, and RJ (200 mg/kg)+CP groups. RJ was administered orally for 14 days. Then, CP at concentrations of 100, 50, and 50 mg/kg was intraperitoneally injected at day 15, 16, 17, respectively. The animals were sacrificed three days after the last injection of CP. Hematological parameters, serum levels of platelet factor 4 (PF4), nitric oxide (NO), and ferric reducing antioxidant power (FRAP) were measured. Also, the pathological analysis of bone marrow, spleen, and testicles was assessed.

CP caused a significant decrease in the number of platelets, white and red blood cells (P<0.001), as well as the levels of FRAP (P<0.01), whereas the serum levels of PF4 and NO were significantly increased. These detrimental alterations were significantly reversed to the baseline upon pretreatment of rats with RJ in the RJ100+CP and RJ200+CP groups (P<0.05). CP caused histological changes in bone marrow, spleen, and testes. Pretreatment with RJ showed noticeable protection against these harmful effects.

RJ prevented CP-induced biochemical and histological damages.

Bioresorbable and titanium plates/screws are considered as a standard treatment for fixation of the bone segments of craniofacial area and paying attention to their biocompatibility is an important issue along with other aspects of application. The purpose of the study was to evaluate the cell viability of two types of plate and screw used in maxillofacial surgeries in contact with gingival fibroblasts and bone marrow stem cells.

In this experimental study after extraction and cultivation of cells from healthy human gingival tissue and alveolar bone of jaw, cytotoxicity of device was evaluated. In direct contact method, samples had near vicinity contact with the both cell lines and in indirect contact method, by-products released, like ions, from samples after 8 weeks were used to assess cytotoxicity. Then cytotoxicity was evaluated on the 2nd, 4th and 6th day with MTS tests and microscopy. The data were analyzed by one-way ANOVA and independent t tests.

There was a statistically significant difference between the German plate and screw and all the samples studied on day 6 (P<0.05). Furthermore, a statistically significant difference was observed between both metal samples and both bio-absorbable samples on day 6 and both cell lines (P<0.05). Comparisons between the two groups with each other for both cell lines on the 6th day were statistically significant (P<0.05).

Our results suggest that that cytotoxicity of biomaterial, from different brands, were not similar and some of the biomaterial showed lower degree of toxicity compared to others and specialist using these products showed be aware of this differences. Our investigation indicates more biocompatibility of bioresorbable plates and screws compared to titanium. In addition our results suggest that biomaterials were not completely neutral.

Health-related studies have been recently at the heart attention of the media. Social media, such as Twitter, has become a valuable online tool to describe the early detection of various adverse drug reactions (ADRs). Different medications have adverse effects on various cells and tissues, sometimes more than one cell population would be adversely affected. These types of side effect are occasionally associated with the direct or indirect influence of prescribed drugs but do not have general unfavorable mutagenic consequences on patients. This study aimed to demonstrate a quick and accurate method to collect and classify information based on the distribution of approved data on Twitter.

In this classification method, we selected "ask a patient" dataset and combination of Twitter "Ask a Patient" datasets that comprised of 6,623, 26,934, and 11,623 reviews. We used deep learning methods with the word2vec to classify ADR comments posted by the users and present an architecture by HAN, FastText, and CNN.

Natural language processing (NLP) deep learning is able to address more advanced peculiarity in learning information compared to other types of machine learning. Moreover, the current study highlighted the advantage of incorporating various semantic features, including topics and concepts.

Our approach predicts drug safety with the accuracy of 93% (the combination of Twitter and "Ask a Patient" datasets) in a binary manner. Despite the apparent benefit of various conventional classifiers, deep learningbased text classification methods seem to be precise and influential tools to detect ADR.

This study aimed to evaluate the specific roles of polyinosinic:polycytidylic acid (polyI:C) in macrophage chemotaxis and reveal the potential regulatory mechanisms related to chemokine receptor 5 (CCR5).

In this experimental study, THP-1-derived macrophages (THP1-Mφs) induced from THP- 1 monocytes were treated with 25 μg/mL polyI:C. Toll-like receptor 3 (TLR3), Jumonji domain-containing protein (JMJD)1A, and JMJD1C small interfering RNA (siRNAs) were transfected into THP1-Mφs. Quantitative real-time reverse transcriptase polymerase chain reaction (qRT-PCR) was used to detect the expression levels of TLR3, CCR5, 23 Jumonji C domain-containing histone demethylase family members, JMJD1A, and JMJD1C in THP1-Mφs with different siRNAs transfections. Western blot was performed to detect JMJD1A, JMJD1C, H3K9me2, and H3K9me3 expressions. A transwell migration assay was conducted to detect THP1-Mφ chemotaxis toward chemokine ligand 3 (CCL3). A chromatin immunoprecipitation (ChIP) assay was performed to detect H3K9me2-CCR5 complexes in THP1- Mφs.

PolyI:C significantly upregulated CCR5 in THP1-Mφs and promoted chemotaxis toward CCL3 (P<0.05); these effects were significantly inhibited by TLR3 siRNA (P<0.01). JMJD1A and JMJD1C expression was significantly upregulated in polyI:C-stimulated THP1-Mφs, while only JMJD1A siRNA decreased CCR5 expression (P<0.05). JMJD1A siRNA significantly increased H3K9me2 expression in THP1-Mφs but not in polyI:C-stimulated THP1-Mφs. The ChIP result revealed that polyI:C significantly downregulated H3K9me2 in the promoter region of CCR5 in THP1- Mφs.

PolyI:C can enhance THP1-Mφ chemotaxis toward CCL3 regulated by TLR3/JMJD1A signalling and activate CCR5 expression by reducing H3K9me2 in the promoter region of CCR5.

Brain ischemia is the most common disease in the world caused by the disruption of the blood supply of brain tissue. Cell therapy is one of the new and effective strategies used for the prevention of brain damages. Sertoli cells (SCs) can hide from the host immune system and secrete trophic factors. So, these cells have attracted the attention of researchers as a therapeutic option for the treatment of neurodegenerative diseases. Also, memantine, as a reducer of glutamate and intracellular calcium, is a suitable candidate for the treatment of cerebral ischemia. The principal target of this research was to examine the effect of SC transplantation along with memantine on ischemic injuries.

In this experimental research, male rats were classified into five groups: sham, control, SC transplant recipient, memantine-treated, and SCs- and memantine-treated groups. SCs were taken from another rat tissue and injected into the right striatum region. A week after stereotaxic surgery and SCs transplantation, memantine was injected. Administered doses were 1 mg/kg and 20 mg/kg at a 12-hour interval. One hour after the final injection, the surgical procedures for the induction of cerebral ischemia were performed. After 24 hours, some regions of the brain including the cortex, striatum, and Piriform cortex-amygdala (Pir-Amy) were isolated for the evaluation of neurological deficits, infarction volume, blood-brain barrier (BBB) permeability, and cerebral edema.

This study shows that a combination of SCs and memantine caused a significant decrease in neurological defects, infarction volume, the permeability of the blood-brain barrier, and edema in comparison with the control group.

Probably, memantine and SCs transplantation reduce the damage of cerebral ischemia, through the secretion of growth factors, anti-inflammatory cytokines, and antioxidant factors.

The gastrointestinal tract (GI) is colonized by a complex microbial community of gut microbiota. Bacteroides spp. have significant roles in gut microbiota and they host interactions by various mechanisms, including outer membrane vesicle (OMVs) production. In the present study, we extracted and assessed Bacteroides fragilis (B. fragilis) and Bacteroides thetaiotaomicron (B. thetaiotaomicron) OMVs in order to evaluate their possible utility for in vivo studies.

In this experimental study, OMVs extraction was performed using multiple centrifugations and tris-ethylenediaminetetraacetic acid (EDTA)-sodium deoxycholate buffers. Morphology, diameter, protein content, profile, and lipopolysaccharide (LPS) concentrations of the OMVs were assessed by scanning electron microscopy (SEM), nanodrop, Bradford assay, sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE), and the Limulus Amoebocyte Lysate (LAL) test, respectively. Zeta potential (ζ-P) was also assessed. The viability effect of OMVs was assessed by the 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay in Caco-2 cells.

Spherical OMVs with diameters of 30-110 nm were produced. The OMVs had different protein profiles. The LPS concentrations of the B. fragilis and B. thetaiotaomicron OMVs were 1.80 and 1.68 EU/mL, respectively. ζ-P of the B. fragilis OMVs was -34.2 mV and, for B. thetaiotaomicron. it was -44.7 mV. The viability of Caco-2 cells treated with OMVs was more than 95%.

The endotoxin concentrations of the spherical OMVs from B. fragilis and B. thetaiotaomicron were within the safe limits. Both OMVs had suitable stability in sucrose solution and did not have any cytotoxic effects on human intestinal cells. Based on our results and previous studies, further molecular evaluations can be undertaken to design OMVs as possible agents that promote health properties.

Autograft transplantation of vitrified cortical ovarian tissue is an acceptable clinical technique for fertility preservation in women. Xenograft transplantation into animal models could be useful for evaluating the safety of human vitrified ovarian tissue. This study targeted to evaluate impact of vitrification on expression of the genes associated with folliculogenesis after xenograft transplantation of human vitrified ovarian tissue to γ-irradiated mice.

In this experimental study, ovarian biopsies were gathered from six transsexual persons. The cortical section of ovarian biopsies was separated and chopped into small pieces. These pieces were randomly divided into vitrified and non-vitrified groups. In each group some pieces were considered as non-transplanted tissues and the others were transplanted to γ-irradiated female National Medical Research Institute (NMRI) mice. Before and after two weeks of xenograft transplantation, histological assessment and evaluation of the expression of folliculogenesisassociated genes (FIGLA, GDF-9, KL and FSHR) were performed in both vitrified and non-vitrified groups.

Percentage of the normal follicles and expression of the all examined genes from transplanted and nontransplanted tissue were similar in both vitrified and non-vitrified groups (P>0.05). After transplantation, the normal follicle rate was significantly decreased and among the folliculogenesis-associated genes, expression of GDF-9 gene was significantly increased, rather than before transplantation in vitrified and non-vitrified tissues (P<0.05).

The vitrification method using dimethyl solphoxide and ethylene glycol (EG) had no remarkable effect on the normal follicular rate and expression of folliculogenesis-associated genes after two weeks human ovarian tissue xenografting. In addition, transplantation process can cause a significant decrease in normal follicular rate and expression of GDF-9 gene.

The aim of the present study was to evaluate the effects of lysophosphatidic acid (LPA) supplementation of human ovarian tissue culture media on tissue survival, follicular development and expression of apoptotic genes following xenotransplantation.

In this experimental study, human ovarian tissue was collected from eight normal female to male transsexual individuals and cut into small fragments. These fragments were vitrified-warmed and cultured for 24 hours in the presence or absence of LPA, then xenografted into back muscles of γ-irradiated mice. Two weeks post-transplantation the morphology of the recovered tissues were evaluated by hematoxylin and eosin staining. The expression of genes related to apoptosis (BAX and BCL2) were analyzed by real time revers transcription polymerase chain reaction (RT-PCR) and detection of BAX protein was done by immunohistochemical staining.

The percent of normal and growing follicles were significantly increased in both grafted groups in comparison to the non-grafted groups, however, these rates were higher in the LPA-treated group than the non-treated group (P<0.05). There was a higher expression of the anti-apoptotic gene, BCL2, but a lower expression of the pro-apoptotic gene, BAX, and a significant lower BAX/ BCL2 ratio in the LPA-treated group in comparison with non-treated control group (P<0.05). No immunostaining positive cells for BAX were observed in the follicles and oocytes in both transplanted ovarian groups.

Supplementation of human ovarian tissue culture medium with LPA improves follicular survival and development by promoting an anti-apoptotic balance in transcription of BCL2 and BAX genes.

The aim of this study was to screen the potential of human embryos to develop into expanding blastocysts following in vitro embryo splitting and then assess the quality of the generated blastocysts based on chromosomal characteristics and using morphokinetics.

In this experimental study, a total of 82 good quality cleavage-stage donated embryos (8- 14 cells) were used (24 embryos were cultured to the blastocyst stage as controls and 58 embryos underwent in vitro splitting). After in vitro splitting, the blastomere donor and blastomere recipient embryos were named twin A and twin B, respectively. Morphokinetics and morphological parameters were evaluated using a time-lapse system in the blastocysts developed from twin embryos. Aneuploidy of chromosomes 13, 15, 16, 18, 21, 22, X and Y were analyzed in the twin blastocysts.

Following in vitro splitting, of the 116 resulting twin embryos, 80 (69%) developed to the expanded blastocyst (EBL) stage compared to 21 (87.5%) embryos in the control group (P>0.05). The morphokinetics analysis suggested that the developmental time-points were influenced by the in vitro splitting. Moreover, the blastocysts developed from A and B twins had impaired morphology compared to controls. Regarding chromosome abnormalities, there was no significant difference in the rate of aneuploidy or mosaicism between the different groups.

This study showed that while no chromosomal abnormalities were seen, in vitro embryo splitting may affect the embryo morphokinetics.

Accumulating evidences indicate that long non-coding RNAs (lncRNAs) play key roles in cancer. This study aims to clarify role of the metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) in non-small cell lung cancer (NSCLC) and uncover the underlying mechanisms.

In this experimental study, MALAT1 and miR-202 expression in tissues and cell lines were detected using quantitative real time polymerase chain reaction (qRT-PCR) assay. Cell transfection was conducted using Lipofectamine 3000. Cell proliferation was determined with CCK-8 assay. MMP2 and MMP9 expressions were measured with Western blot. Cell invasive ability was evaluated by Transwell assay. Starbase 2.0 tool was used to predict targets of MALAT1. Dual luciferase reporter assay, RNA-binding protein immunoprecipitation assay and RNA pull-down assay were conducted to confirm the potential direct interaction between MALAT1 and miR-202.

MALAT1 was overexpressed in NSCLC samples and cell lines. High expression of MALAT1 was related to large tumor size (>3 cm), poor histological grade, advanced cancer and tumor metastasis in NSCLC. In vitro assays exhibited that knockdown of MALAT1 remarkably decreased A549 cell growth and invasion capacity, while overexpression of MALAT1 significantly enhanced NCI-H292 cell proliferation and invasion ability. Next, we verified that MALAT1 could act as a competing endogenous RNA (ceRNA) by sponging miR-202 in NSCLC and there is a negative correlation between MALAT1 and miR-202. Besides, overexpression of miR-202 inhibited cell proliferation and invasive ability in MALAT1-overexpressed cells.

This study demonstrated that lncRNA-MALAT1 gets involved in NSCLC progression by targeting miR- 202, indicating that MALAT1 may serve as a novel therapeutic target for NSCLC treatment.

We aimed to explore potential molecular mechanisms of clear cell renal cell carcinoma (ccRCC) and provide candidate target genes for ccRCC gene therapy.

This is a bioinformatics-based study. Microarray datasets of GSE6344, GSE781 and GSE53000 were downloaded from Gene Expression Omnibus database. Using meta-analysis, differentially expressed genes (DEGs) were identified between ccRCC and normal samples, followed by Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway and Gene Ontology (GO) function analyses. Then, protein-protein interaction (PPI) networks and modules were investigated. Furthermore, miRNAs-target gene regulatory network was constructed.

Total of 511 up-regulated and 444 down-regulated DEGs were determined in the present gene expression microarray data meta-analysis. These DEGs were enriched in functions like immune system process and pathways like Toll-like receptor signaling pathway. PPI network and eight modules were further constructed. A total of 10 outstanding DEGs including TYRO protein tyrosine kinase binding protein (TYROBP), interferon regulatory factor 7 (IRF7) and PPARG co-activator 1 alpha (PPARGC1A) were detected in PPI network. Furthermore, the miRNAs-target gene regulation analyses showed that miR-412 and miR-199b respectively targeted IRF7 and PPARGC1A to regulate the immune response in ccRCC.

TYROBP, IRF7 and PPARGC1A might play important roles in ccRCC via taking part in the immune system process.

Endoplasmic reticulum (ER) stress causes an adaptive response initiated by protein kinase RNA-like ER kinase (PERK), Ire1 and ATF6. It has been reported that these upstream regulators induce microRNAs. The current study was designed to find a novel microRNA that mediates ER stress components and finally contributes to cell fate decision.

In this experimental study, miR-706 levels were checked under different conditions of ER stress induced by Thapsigargin, Tunicamycin or low glucose media. PERK and ATF4 were knocked-down by administration of lentivirus-mediated short hairpin RNA to explore the effect of ER stress related proteins on miR-706 expression. The effect of miR-706 on caspase activity and apoptosis inhibitor 1 (CAAP1) levels were examined by using mimic-miR-706. The role of CAAP1 in inhibiting cell death (measured by Annexin V staining) and contributing to patient overall survival (measured by Kaplan-Meier estimate) were further confirmed by antimiR- 706 and CAAP1 knock-down.

We showed that Thapsigargin or Tunicamycin triggered ER stress leading to the induction of miR-706. miR-706 induction is dependent on PERK and its downstream regulator ATF4, as knocking-down of PERK and ATF4 suppressed miR-706 induction in response to ER stress. Knocking-down of miR-706 reduces cell death triggered by ER stress, indicating that miR-706 is pro-cell death microRNA. We further identified CAAP1 as a miR-706 target in regulating ER stress initiated cell death.

Collectively, our results pointed to an ER signaling network consisting of proteins, microRNA and novel target.