Memory impairment was ameliorated by corticolimbic microinjections of arachidonylcyclopropylamide (ACPA) and miRNA-regulated lentiviral particles in a streptozotocin-induced Alzheimer's rat model.

The present study aimed to investigate the effect of corticolimbic cannabinoid CB1 receptors activity on memory impairment in the intracerebroventricular (ICV)-streptozotocin (STZ) animal model of Alzheimer's like-disease. This study also assessed whether the corticolimbic overexpression of miRNA-137 or -let-7a could increase the endocannabinoids by inhibiting the monoglyceride lipase (MAGL) to ameliorate STZ response. The results showed that ICV microinjection of STZ (3 mg/kg/10 µl) impaired passive avoidance memory retrieval. The chronic microinjection of arachidonylcyclopropylamide (ACPA; 10 ng/0.5 µl), a selective cannabinoid CB1 receptor agonist, into the hippocampal CA1 region, the central amygdala (CeA) or the medial prefrontal cortex (mPFC) ameliorated the amnesic effect of ICV-STZ. Intra-CA1 or -CeA microinjection of ACPA alone did not affect memory retrieval, while its microinjection into the mPFC impaired memory formation. Based on bioinformatics analysis and verification of the MAGL gene, miRNA-137 and -let-7a were chosen to target the expression levels of MAGL in the corticolimbic regions. The chronic corticolimbic microinjection of lentiviral particles containing miRNA-137 or -let-7a ameliorated ICV-STZ-induced memory impairment. The high transfection efficiency was determined for each virus using comparing fluorescent and conventional vision. Corticolimbic overexpression of miRNA-137 or -let-7a decreased the MAGL gene expression that encodes the MAGL enzyme to increase the endocannabinoids. Thus, among the molecular mechanisms and signaling pathways involved in the pathophysiology of Alzheimer's disease (AD), it is worth mentioning the role of endocannabinoids in the corticolimbic regions. CB1 receptor agonists, miRNA-137 or -let-7a, may be potential therapeutic targets against cognitive decline in AD.

Distinct power of bone marrow microRNA signatures and tumor suppressor genes for early detection of acute leukemia

BackgroundAcute leukemia involving lymphocytic and myeloid cells is cancer with a high mortality rate. Swift and timely diagnosis might be a potential approach to improving patient prognosis and survival. The microRNA (miRNA) signatures are emerging nowadays for their promising diagnostic potential. MiRNA levels from bone marrow can be used as prognostic biomarkers.MethodsThe current study was designed to evaluate if the microRNAs and tumor suppressor genes (TSGs) profiling of hematopoietic bone marrow could help in acute leukemia early detection. Also, we assessed the DNA methyltransferase 3A (DNMT3A) expression and its possible epigenetic effects on miRNAs plus TSGs expression levels. The expression levels of ten miRNAs and four TSGs involved in acute lymphocytic leukemia (ALL) as well as acute myeloid leukemia (AML) were quantified in 43 and 40 bone marrow samples of ALL and AML patients in comparison with cancer-free subjects via real-time quantitative PCR (RT-qPCR). The receiver-operating-characteristic (ROC) analysis of miRNAs was performed in the study groups. Further, the correlation between the DNMT3A and TSGs was calculated.ResultsSignificant differences were detected in the bone marrow expression of miRNAs and TSGs (P < 0.05) between acute leukemia patients and healthy group. ROC analysis confirmed the ability of miR-30a, miR-101, miR-132, miR-129, miR-124, and miR-143 to discriminate both ALL and AML patients with an area under the ROC curve of ≥ 0.80 (P < 0.001) and high accuracy. The correlation between DNMT3A and P15/P16 TSGs revealed that DNMT3A plays a vital role in epigenetic control of TSGs expression. Our findings indicated that the downregulation of bone marrow miRNAs and TSGs was accompanied by acute leukemia development.ConclusionsThe authors conclude that this study could contribute to introducing useful biomarkers for acute leukemia diagnosis.

Distinct power of bone marrow microRNA signatures and tumor suppressor genes for early detection of acute leukemia.

BACKGROUND: Acute leukemia involving lymphocytic and myeloid cells is cancer with a high mortality rate. Swift and timely diagnosis might be a potential approach to improving patient prognosis and survival. The microRNA (miRNA) signatures are emerging nowadays for their promising diagnostic potential. MiRNA levels from bone marrow can be used as prognostic biomarkers. METHODS: The current study was designed to evaluate if the microRNAs and tumor suppressor genes (TSGs) profiling of hematopoietic bone marrow could help in acute leukemia early detection. Also, we assessed the DNA methyltransferase 3A (DNMT3A) expression and its possible epigenetic effects on miRNAs plus TSGs expression levels. The expression levels of ten miRNAs and four TSGs involved in acute lymphocytic leukemia (ALL) as well as acute myeloid leukemia (AML) were quantified in 43 and 40 bone marrow samples of ALL and AML patients in comparison with cancer-free subjects via real-time quantitative PCR (RT-qPCR). The receiver-operating-characteristic (ROC) analysis of miRNAs was performed in the study groups. Further, the correlation between the DNMT3A and TSGs was calculated. RESULTS: Significant differences were detected in the bone marrow expression of miRNAs and TSGs (P < 0.05) between acute leukemia patients and healthy group. ROC analysis confirmed the ability of miR-30a, miR-101, miR-132, miR-129, miR-124, and miR-143 to discriminate both ALL and AML patients with an area under the ROC curve of ≥ 0.80 (P < 0.001) and high accuracy. The correlation between DNMT3A and P15/P16 TSGs revealed that DNMT3A plays a vital role in epigenetic control of TSGs expression. Our findings indicated that the downregulation of bone marrow miRNAs and TSGs was accompanied by acute leukemia development. CONCLUSIONS: The authors conclude that this study could contribute to introducing useful biomarkers for acute leukemia diagnosis.

Development of Inactivated FAKHRAVAC® Vaccine against SARS-CoV-2 Virus: Preclinical Study in Animal Models.

The recent viral infection disease pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has resulted in a global public health crisis. Iran, as one of the countries that reported over five million infected cases by September 2021, has been concerned with the urgent development of effective vaccines against SARS-CoV-2. In this paper, we report the results of a study on potency and safety of an inactivated SARS-CoV-2 vaccine candidate (FAKHRAVAC) in a preclinical study so as to confirm its potential for further clinical evaluation. Here, we developed a pilot-scale production of FAKHRAVAC, a purified inactivated SARS-CoV-2 virus vaccine candidate that induces neutralizing antibodies in Balb/c mice, guinea pigs, rabbits, and non-human primates (Rhesus macaques-RM). After obtaining ethical code of IR.IUMS.REC.1399.566, immunizations of animals were conducted by using either of three different vaccine dilutions; High (H): 10 µg/dose, Medium (M): 5 µg/dose, and Low (L): 1 µg/dose, respectively. In the process of screening for viral seeds, viral strains that resulted in the most severe clinical manifestation in patients have been isolated for vaccine development. The viral seed produced the optimal immunity against SARS-CoV-2 virus, which suggests a possible broader neutralizing ability against SARS-CoV-2 strains. The seroconversion rate at the H-, M-, and L-dose groups of all tested animals reached 100% by 28 days after immunization. These data support the eligibility of FAKHRAVAC vaccine candidate for further evaluation in a clinical trial.

PANC-1 cancer stem-like cell death with silybin encapsulated in polymersomes and deregulation of stemness-related miRNAs and their potential targets.

OBJECTIVES: Cancer stem cells (CSCs) have powerful self-renewal ability and tumor recurrence. Pancreatic ductal adenocarcinoma is a malignancy with high mortality rate and ˃5% survival. Silybin has anticancer and hepatoprotective properties. We loaded silybin in PEG400-OA (SPNs) and evaluated its cytotoxic effects on PANC-1 cells and PANC-1 CSCs. MATERIALS AND METHODS: Spheroids from PANC-1 cells were obtained by the hanging drop method. Anti-proliferative and apoptotic functions of SPNs were evaluated in spheroids and non-spheroids with MTT, DNA fragmentation, PI and PI/AnnexinV assays. The expression of CD markers was assessed with flow cytometry. QRT-PCR was used to evaluate the expression of some miRNAs and targets. RESULTS: IC50 of SPNs was identified to be 50 µg/ml, 45 µg/ml, and 42µg/ml, respectively after 24 hr, 48 hr, and 72 hr in PANC-1 treated cells. PI staining and PI/AnnexinV assay showed that ~20%, ~60%, and ~80%, of cells treated with 30, 50, and 60 µg/ml of SPNs were in sub-G1 and apoptosis phase, respectively. DNA degradation was confirmed after SPNS stimulation. CD24, CD44, and CD133 expression decreased after SPNs treatment both in PANC-1 spheroid cells and PANC-1 cancer cell line. Under-expression of onco-miRs (miR-21, miR-155, and miR-221), over-expression of several apoptotic potential targets of oncomiRs (Bax, Casp-9, and P53), over-expression of tumor suppressive-miRs (let-7b, miR-34a, and miR-126), and under-expression of Bcl-2 was found in SPNs-treated cells. CONCLUSION: We suggest that silybin encapsulated in polymersomes (SPNs) may be useful as a complementary agent for destroying both pancreatic cancer cells and pancreatic CSCs along with chemotherapeutic agents.

Expression analysis of NF-κB-associated long noncoding RNAs in peripheral blood mononuclear cells from relapsing-remitting multiple sclerosis patients.

Long noncoding RNAs (lncRNAs) as potential disease biomarkers might be related to severe course of multiple sclerosis (MS). We evaluated expression levels of NF-κB-associated lncRNAs including HOTAIR, THRIL, H19, NKILA, and ANRIL; as well as expression of IL-6, TNF-α and MMP9, in peripheral blood mononuclear cells (PBMCs) from 60 relapse-remitting MS (RRMS) patients. At relapse phase of RRMS, up-regulation of ANRIL and H19 was positively correlated with the overexpression of IL-6; high levels of THRIL and HOTAIR was positively correlated with increased levels of TNF-α and MMP9, respectively; however, the NKILA expression was negatively correlated with the expression of TNF-α.

Effects of increasing lipopolysaccharide concentrations on in vitro developmental competence of ovine oocytes.

Although a considerable number of studies have investigated the effects of lipopolysaccharide (LPS) on the reproductive performance of dairy cows, the response of ovine oocytes to LPS during their in vitro maturation and development is not well defined yet. Ewe's ovaries were obtained from a slaughterhouse, the oocytes were collected and matured in the presence of increasing concentrations (0, 0.01, 0.1, 1 and 10 µg/mL) of LPS in order to evaluate the meiotic maturation by measuring the proportion of oocytes reaching the MII stage. The concentration of intracellular glutathione (GSH) was measured in oocytes following maturation in vitro. In addition, concentrations of selected metabolites including glucose, pyruvate, lactate and glutamine were quantified in the medium following maturation. A number of treated matured oocytes along with the control group were subsequently fertilized using frozen semen and assessed for the rate of cleavage and for the proportion reaching the blastocyst stage. The number of oocytes in MII stage was significantly reduced in response to the increasing concentrations of LPS (77.83%, 70.64%, 68.86%, 66.32%, respectively, in case of 0.01, 0.1, 1 and 10 µg/mL LPS when compared to the control group, 76.34%; P<0.05). There were no differences neither in the intracellular concentration of GSH in the oocytes nor in case of the metabolites in the maturation medium. Although the rate of cleaved oocytes was not affected by increasing levels of LPS, the blastocyst rate was reduced in a dose dependent manner (36.69%, 34.21%, 30.35%, 17.27% and 14.03% for the control, 0.01, 0.1, 1 and 10 µg/mL LPS, respectively (P<0.05). These results demonstrate that the developmental competence of ovine oocytes may be affected detrimentally by LPS and such deleterious effects could be related to the maturation process.

Involved microRNAs in alternative polyadenylation intervene in breast cancer via regulation of cleavage factor "CFIm25".

Cleavage factor "CFIm25", as a key repressor at proximal poly (A) site, negatively correlates to cell proliferation and tumorigenicity in various cancers. Hence, understanding CFIm25 mechanism of action in breast cancer would be a great benefit. To this aim four steps were designed. First, potential miRNAs that target 3'-UTR of CFIm25 mRNA, retrieved from Targetscan web server. Second, screened miRNAs were profiled in 100 breast cancer and 100 normal adjacent samples. Third, miRNAs that their expression was inversely correlated to the CFIm25, overexpressed in MDA-MB-231 cell line, and their effect on proliferation and migration monitored via MTT and wound healing assays, respectively. Fourth, interaction of miRNAs of interest with 3'-UTR of CFIm25 confirmed via luciferase assay and western blot. Our results indicate that CFIm25 considerably down-regulates in human breast cancer tissue. qRT-PCR assay, luciferase test, and western blotting confirm that CFIm25 itself could be directly regulated by oncomiRs such as miR-23, -24, -27, -135, -182 and -374. Besides, according to MTT and wound healing assays of cell lines, CFIm25 knockdown intensifies cell growth, proliferation and migration. Our results also confirm indirect impact of CFIm25 on regulation of mRNA's 3'-UTR length, which then control corresponding miRNAs' action. miRNAs directly control CFIm25 expression level, which then tunes expression of the oncogenes and tumor proliferation. Therefore, regulation of CFIm25 expression level via miRNAs is expected to improve treatment responses in breast cancer.

Apoptosis induction and proliferation inhibition by silibinin encapsulated in nanoparticles in MIA PaCa-2 cancer cells and deregulation of some miRNAs.

OBJECTIVES: Silibinin, as an herbal compound, has anti-cancer activity. Because of low solubility of silibinin in water and body fluids, it was encapsulated in polymersome nanoparticles and its effects were evaluated on pancreatic cancer cells and cancer stem cells. MATERIALS AND METHODS: MIA PaCa-2 pancreatic cancer cells were treated with different doses of silibinin encapsulated in polymersome nanoparticles (SPNs). Stemness of MIA PaCa-2 cells was evaluated by hanging drop technique and CD133, CD24, and CD44 staining. The effects of SPNs on cell cycle, apoptosis and the expression of several genes and miRNAs were investigated. RESULTS: IC50 of SPNs was determined to be 40 µg/ml after 24 hr. Our analysis showed that >98% of MIA PaCa-2 cells expressed three stem cell markers. FACS analysis showed a decrease in these markers in SPNs-treated cells. PI/AnnexinV staining revealed that 40 µg/ml and 50 µg/ml of SPNs increased apoptosis up to ~40% and >80% of treated cells, respectively. Upregulation of miR-34a, miR-126, and miR-let7b and downregulation of miR-155, miR-222 and miR-21 was observed in SPNs-treated cells. In addition, downregulation of some genes involved in proliferation or migration such as AKT3, MASPINE, and SERPINEA12, and upregulation of apoptotic genes were observed in treated cells. CONCLUSION: Our results suggested that SPNs induced apoptosis and inhibited migration and proliferation in pancreatic cells and cancer stem cells through suppression of some onco-miRs and induction of some tumor suppressive miRs, as well as their targets.

Combination of low intensity electromagnetic field with chondrogenic agent induces chondrogenesis in mesenchymal stem cells with minimal hypertrophic side effects.

Background: There are different methods to develop in vitro neo-chondral tissues from adipose-derived stem cells (ADSCs). Application of electromagnetic field (EMF) on ADSCs is one of popular approaches, which results in chondrogenesis. If chondrogenic impact of EMF on ADSCs is supposed to be generalized as a protocol in translational medicine field, possible emergence of early or late hypertrophic maturation, mineralization and inflammatory side effects in chondrogenically differentiating ADSCs should be considered.Methods: The advent of chondrogenic and hypertrophic markers by differentiated cells under standard, platelet-rich plasma (PRP)-based or EMF treatments were monitored. Along with monitoring the expressions of chondrogenic markers, inflammatory and hypertrophic markers, VEGF/TNFα secretion, calcium deposition and ALP activity were evaluated.Results: Accordingly, treatment with %5 PRP results in higher GAG production, enhanced SOX9 transcription, lowered TNFα and VEGF secretions compared to other treatments. Although PRP up-regulates miR-146a and miR-199a in early and late stages of chondrogenesis, respectively, application of EMF + PRP down regulates miR-101 and -145 while up-regulates miR-140 and SOX9 expression.Conclusion: Comparing our results with previous reports suggests that presented EMF-ELF in this study with f = 50 Hz, EMF intensity of less than 30 mT, and 5% PRP (v/v), would facilitate chondrogenesis via mesenchymal stem cells with minor inflammation and hypertrophic maturation.Abbreviations: MSCs: mesenchymal stem cells; TGFß: transforming growth factor-beta; PRP: platelet-rich plasma; ELF-EMF: extremely low-frequency electromagnetic fields; GAGs: glycosaminoglycans; ADSCs: adipose-derived stem cells; VEGF: vascular endothelial growth factor; TNFα: tumor necrosis factor alpha; ALP: alkaline phosphatase.

Comparative impact of platelet rich plasma and transforming growth factor-ß on chondrogenic differentiation of human adipose derived stem cells.

Introduction: Transforming growth factor-beta (TGF-ß) is known as standard chondrogenic differentiation agent, even though it comes with undesirable side effects such as early hypertrophic maturation, mineralization, and secretion of inflammatory/angiogenic factors. On the other hand, platelet-rich plasma (PRP) is found to have a chondrogenic impact on mesenchymal stem cell proliferation and differentiation, with no considerable side effects. Therefore, we compared chondrogenic impact of TGF-ß and PRP on adipose-derived stem cells (ADSCs), to see if PRP could be introduced as an alternative to TGF-ß. Methods: Differentiation of ADSCs was monitored using a couple of methods including glycosaminoglycan production, miRNAs expression, vascular endothelial growth factor (VEGF)/tumor necrosis factor alpha (TNFα) secretion, alkaline phosphatase (ALP) and calcium content assays. Results: Accordingly, the treatment of differentiating cells with 5% (v/v) PRP resulted in higher glycosaminoglycan production, enhanced SOX9 transcription, and lowered TNFα and VEGF secretion compared to the control and TGF-ß groups. Besides, the application of PRP to the media up-regulated miR-146a and miR-199a in early and late stages of chondrogenesis, respectively. Conclusion: PRP induces in vitro chondrogenesis, as well as TGF-ß with lesser inflammatory and hypertrophic side effects.

Maternal supplementation with fish oil modulates inflammation-related MicroRNAs and genes in suckling lambs.

Dietary n-3 long-chain fatty acids (n-3 LCFA) have been shown to modify lipid metabolism and immune function. The objective of this study was to evaluate the effect of periparturient fish oil (FO) supplementation on the inflammation and metabolic health of ewes and their lambs at a molecular level. Prepartum ewes were fed control diet (CON, n = 12) or CON supplemented with 2% DM of calcium soap of FO (n = 12) from 28 days before until 21 days after parturition. The ewes were evaluated for plasma metabolites and milk composition. The experiment was followed by analyzing the relative transcript abundance of circulating microRNAs (miRNAs) in plasma and targeted miRNA/mRNA expression in peripheral blood mononuclear cells (PBMCs) in both ewes and lambs. FO treatment decreased prepartum feed intake (1812 ± 35 vs 1674 ± 33 g/day, P < 0.01), whereas the influence on plasma metabolites was negligible. Dietary FO supplementation decreased milk fat percentage (8.82 ± 0.49 vs 7.03 ± 0.45, P = 0.02) and reduced milk n-6/n-3 (P < 0.05). Also, it altered the expression of plasma-circulating miRNAs in both ewe and lamb (P < 0.05). Furthermore, maternal nutrition of FO downregulated the relative expression of miR-33a and miR-146b and transcript abundance of genes IL-1ß (0.41-fold) and NF-κB (0.25-fold) in lambs' PBMC. In conclusion, results showed that FO supplementation starting antepartum affects milk composition and circulating miRNA in dams and the inflammatory markers in lambs delivered by the supplemented ewes. These may provide a strategy to maintain immune balance during gestation and develop the immune system in lambs.

The effect of miR-579 on the PI3K/AKT pathway in human glioblastoma PTEN mutant cell lines.

Glioblastoma multiform (GBM) is a type of aggressive brain cancer with limited success in standard treatment. MicroRNAs are one of the most beneficial tools for diagnosis, prognosis, and treatment of cancer. This study aimed to investigate the effect of miR-579 on cellular behaviors and expression of PI3K/AKT signaling pathway in GBM cell lines. In the present study, miR-579 was overexpressed in U251 and A-172 cell lines by using lentil vector, and its effect on cellular behavior such as proliferation and migration was investigated by the cell cycle, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), Annexin V, colony formation, Transwell and wound healing assays. MiR-579 predicted target genes (AKT1, Rheb, PDK1, and a few others) were also evaluated by real-time polymerase chain reaction or luciferase assay and Western blot analysis. Our results represented that overexpression of miR-579 could inhibit proliferation, migration, cell cycle and also promoted the apoptosis of GBM cell lines. The luciferase reporter assay showed miR-579 directly targets the 3 UTR of mTOR, Rheb, and PDK1 and repressed their expressions. Furthermore, the Western blot analysis showed that miR-579 could downregulate the AKT1 and Rheb protein expression. Overall, our findings propose that miR-579 functions as a novel tumor suppressor gene in GBM by regulating the PI3K/AKT signaling pathway and may serve as a therapeutic target for clinical therapy of glioblastoma multiform.

Simultaneous regulation of miR-451 and miR-191 led to erythroid fate decision of mouse embryonic stem cell.

OBJECTIVES: Various microRNAs (miRNAs) are expressed during development of mammalian cells, when they aid in modulating gene expression by mediating mRNA transcript cleavage and/or regulation of translation rate. miR-191 and miR-451 have been shown to be critical regulators of hematopoiesis and have important roles in the induction of erythroid fate decision. So, the aim of this study is investigation of the miR-191 and miR-451 roles in the controlling mouse embryonic stem cell (mESC) differentiation toward the erythroid lineage. MATERIALS AND METHODS: mESCs were infected with either pCDH-miR-Off-191 viruses in pCDH-miR-Off-191 group or simultaneously with pCDH-miR-Off-191 and pCDH-miR-451 lentiviruses in simultaneous group. Then, the expression profiles of erythroid specific transcription factors and globin genes were analyzed using QRT-PCR on day 14 and 21 of differentiation. Flow cytometry analysis was used to evaluate of TER119 and CD235a erythroid specific surface markers. RESULTS: Gata-1, Klf-1, Epor and globin chains were found to be expressed in pCDH-miR-Off-191 and in simultaneous groups. The majority of globin chains showed changes in their expression levels with progression of differentiation from day 14 to day 21. Flow cytometry results showed that miR-451 up- regulation and miR-191 down-regulation is associated with the expression of TER119 and CD235a. Of these two groups analyzed, simultaneous group was most significantly potent in stimulation of erythroid fate decision of mESCs. CONCLUSION: Together, present data demonstrate that down-regulation of miR-191 alone can enhance the differentiation of mESCs. However, the simultaneous effect of miR-451up-regulation and miR-191 down-regulation is much stronger and can have more practical use in artificial blood production.

A critical role for miR-184 in the fate determination of oligodendrocytes.

BACKGROUND: New insights on cellular and molecular aspects of both oligodendrocyte (OL) differentiation and myelin synthesis pathways are potential avenues for developing a cell-based therapy for demyelinating disorders comprising multiple sclerosis. MicroRNAs (miRNA) have broad implications in all aspects of cell biology including OL differentiation. MiR-184 has been identified as one of the most highly enriched miRNAs in oligodendrocyte progenitor cells (OPCs). However, the exact molecular mechanism of miR-184 in OL differentiation is yet to be elucidated. METHODS AND RESULTS: Based on immunochemistry assays, qRT-PCR, and western blotting findings, we hypothesized that overexpression of miR-184 in either neural progenitor cells (NPCs) or embryonic mouse cortex stimulated the differentiation of OL lineage efficiently through regulating crucial developmental genes. Luciferase assays demonstrated that miR-184 directly represses positive regulators of neural and astrocyte differentiation, i.e., SOX1 and BCL2L1, respectively, including the negative regulator of myelination, LINGO1. Moreover, blocking the function of miR-184 reduced the number of committed cells to an OL lineage. CONCLUSIONS: Our data highlighted that miR-184 could promote OL differentiation even in the absence of exogenous growth factors and propose a novel strategy to improve the efficacy of OL differentiation, with potential applications in cell therapy for neurodegenerative diseases.

miR-455-5p downregulation promotes inflammation pathways in the relapse phase of relapsing-remitting multiple sclerosis disease.

MicroRNA-455-5p (miR-455-5p) seems to have an anti-inflammatory role in the immune system since its expression is induced by IL-10 cytokine. Multiple sclerosis (MS) is a chronic demyelinating neurodegenerative disease of the central nervous system that is caused by an autoimmune inflammatory attack against the myelin insulation of neurons. The expression level of miR-455-5p and its role in MS pathogenesis has yet to be elucidated. We found that miR-455-5p expression was highly correlated with disease severity in MS patients. miR-455-5p expression inversely correlates with its inflammatory-predicted targets (MyD88 and REL) in relapse- and remitting-phase patients. Luciferase assays confirm that MyD88 and REL are direct targets of miR-455-5p. This study represents the first report of the miR-455-5p acts as an anti-inflammatory role in MS, at least partially through targeting MyD88 and REL. This study may provide important information for the use of miR-455-5p as a novel strategy to improve the severity of disease and control inflammation and attack in MS patients.

Increased expression of mir-301a in PBMCs of patients with relapsing-remitting multiple sclerosis is associated with reduced NKRF and PIAS3 expression levels and disease activity.

Most of the multiple sclerosis (MS) patients are initially diagnosed with relapsing remitting multiple sclerosis (RRMS). Th17 cells and macrophages have been shown to play critical roles in pathogenesis of MS and initiation of CNS tissue damage. MiR-301a have recently been exposed as an activator of STAT3 in Th17 cells as well as an activator of NF-κB in macrophages by targeting PIAS3 and NKRF correspondingly. However, the possible role of miR-301a in RRMS has not yet been elucidated. Herewith, for the first time, we have studied the expression of miR-301a, NKRF and PIAS3 by quantitative real-time PCR and western blotting method in peripheral blood mononuclear cells (PBMCs) of 71 RRMS patients, including two groups of patients in relapse phase (n = 44) and a group of remitting phase patients (n = 28) in comparison to healthy volunteers (n = 28). In this work, we demonstrate a significant upregulation of miR-301a in relapse phase of MS patients compared to healthy controls and remitting phase patients (P < .05). Our findings also showed a striking decrease of NKRF and PIAS3 expression in relapse phase patients, in contrast to miR-301a and, NF-κB and STAT3 downstream genes (SKA2 and RORc) (P < .05). Subsequently, using luciferase reporter system we confirmed that miR-301a directly targets the mRNA encoding PIAS3 and NKRF proteins. We also showed that miR-301a increasing expression is correlated with down-regulation of PIAS3 and NKRF expression in RRMS patients. Our findings suggest that miR-301a, PIAS3 and NKRF play crucial roles in RRMS and could be considered as promising therapeutic targets.

Overexpression of miR-133 decrease primary endothelial cells proliferation and migration via FGFR1 targeting.

Angiogenesis is one of the essential hallmarks of cancer that is controlled by the balance between positive and negative regulators. FGFR1 signaling is crucial for the execution of bFGF-induced proliferation, migration, and tube formation of endothelial cells (ECs) and onset of angiogenesis on tumors. The purpose of this study is to identify whether or not miR-133 regulates FGFR1 expression and accordingly hypothesize if it plays a crucial role in modulating bFGF/FGFR1 activity in ECs and blocking tumor angiogenesis through targeting FGFR1. The influences of miR-133 overexpression on bFGF stimulated endothelial cells were assessed by cell growth curve, MTT assaying, tube formation, and migration assays. Forced expression of miR-133 caused significant reductions in bFGF-induced proliferation and migratory ability of ECs. MiR-133 Expression was negatively correlated with both mRNA and protein levels of FGFR1 in the transfected ECs isolated from peripheral blood. Moreover, overexpression of miR-133 drastically reduced the rate of cell division and disturbed capillary network formation of transfected ECs. These findings suggest that miR-133 plays an important function in bFGF-induced angiogenesis processes in ECs and provides a rationale for new therapeutic approaches to suppress tumor angiogenesis and cancer.

Targeting histone demethylases KDM5A and KDM5B in AML cancer cells: A comparative view.

Epigenetic modifications play an important role in initiation and progression of cancers including acute myeloid leukemia. Among different epigenetic modifiers, lysine specific demethylases have been noticed as potential therapeutic targets. KDM5 family of histone demethylases which removes methyl marks from lysine residues of H3, are frequently found in the promoter region of transcriptionally active genes resulting in repression of expression. Here we have compared the effects of KDM5A and KDM5B downregulation on HL-60 cell line behavior. KDM5A/5B knockdown resulted in lower viability of HL-60 cells in addition to modified cell cycle distribution and sub-G1 accumulation. Induction of apoptosis was observed in both knockdown cells. But in spite of similarity in their role, downregulation of KDM5A showed more efficient anti-leukemic effects in comparison to KDM5B. Cells showed higher accumulation in sub-G1 and apoptosis occurred significantly higher and also earlier after KDM5A reduction. Expression analysis confirmed almost 5 and 4 fold increased expression for bax and caspase-3 after downregulation of KDM5A in comparison to KDM5B. Due to the present study we propose KDM5A as a potential target for therapeutic aspects of acute myeloid leukemia although further investigations are needed.

Functional SNP in microRNA-491-5p binding site of MMP9 3'-UTR affects cancer susceptibility.

MicroRNAs (miRNA) are small RNA molecules that negatively regulate gene expression through base pairing interactions between 3'-UTR of the target mRNAs and seed sequence of miRNA. Any changes in the recognition site could destroy binding sites or modify binding affinity, resulting in evasion from miRNA regulation. A putative binding site for miR-491-5p resides in 3'-UTR of MMP9, and a genetic variant (rs1056628 A → C) is present in this region. The role of MMP9 over expression well marked in various cancers. However, whether rs1056628 SNP in miR-491-5p binding site of MMP9 3'-UTR could abrogate its post-transcriptional regulation and affect cancer susceptibility remains largely unknown. To test this, the rs1056628 SNP was genotyped in 300 cases of lung, gastric and breast cancers and 200 age- and sex-matched healthy controls. The results showed that compared with the AA genotype, C was a risk genotype for all three cancers development and was also associated with gastric and breast cancers metastasis and invasion. Based on the base pairing analysis and secondary structure evaluation of MMP9 mRNA and miR-491-5p, we found that miR-491-5p had a higher binding affinity for A genotype than the C genotype. The Luciferase activity of MMP9 3'-UTR indicates differential regulation of two genetic variations of MMP9. Overexpression of miR-491-5p decreased MMP9 mRNA level in cell lines of gastric, breast and lung cancers and thus leads to decreasing of the invasion ability. Therefore, for the first time we imply that the C variant of MMP9 contributes to the likelihood of gastric, breast and lung cancers susceptibility via a novel mechanism of subtle gene regulation through miRNA binding capacity.