miR-155 Inhibitor Research Articles

MicroRNAs affect GPCR and Ion channel genes needed for influenza replication.

Influenza virus causes seasonal epidemics and sporadic pandemics resulting in morbidity, mortality, and economic losses worldwide. Understanding how to regulate influenza virus replication is important for developing vaccine and therapeutic strategies. Identifying microRNAs (miRs) that affect host genes used by influenza virus for replication can support an antiviral strategy. In this study, G-protein coupled receptor (GPCR) and ion channel (IC) host genes in human alveolar epithelial (A549) cells used by influenza virus for replication (Orr-Burks et al., 2021) were examined as miR target genes following A/CA/04/09- or B/Yamagata/16/1988 replication. Thirty-three miRs were predicted to target GPCR or IC genes and their miR mimics were evaluated for their ability to decrease influenza virus replication. Paired miR inhibitors were used as an ancillary measure to confirm or not the antiviral effects of a miR mimic. Fifteen miRs lowered influenza virus replication and four miRs were found to reduce replication irrespective of virus strain and type differences. These findings provide evidence for novel miR disease intervention strategies for influenza viruses.

MiR-17 acts as a tumor suppressor by negatively regulating the miR-17-92 cluster

Anaplastic thyroid cancer (ATC) is an aggressive, highly metastatic cancer that expresses high levels of the microRNA (miR)-17-92 cluster. We employ an miR inhibitor system to study the function of the different miRs within the miR-17-92 cluster based on seed sequence homology in the ATC SW579 cell line. While three of the four miR-17-92 families were oncogenic, we uncovered a novel role for miR-17 as a tumor suppressor in vitro and in vivo. Surprisingly, miR-17 inhibition increased expression of the miR-17-92 cluster and significantly increased the levels of the miR-18a and miR-19a mature miRs. miR-17 inhibition increased expression of the cell cycle activator CCND2, associated with increased cell proliferation and tumor growth in transplanted SW579 cells in xenograft mice. miR-17 regulates MYCN and c-MYC expression in SW579 cells, and the inhibition of miR-17 increased MYCN and c-MYC expression, which increased pri-miR-17-92 transcripts. Thus, inhibition of miR-17 activated the expression of the oncogenic miRs, miR-18a and miR-19a. While many cancers express high levels of miR-17, linking it with tumorigenesis, we demonstrate that miR-17 inhibition does not inhibit thyroid tumor growth in SW579 and MDA-T32 ATC cells but increases expression of the other miR-17-92 family members and genes to induce cancer progression.

MiR-153 Enriched in Bone Marrow Mesenchymal Stem Cell-Derived Exosomes Promotes Chemotherapy Sensitivity of Papillary Thyroid Carcinoma

Our study assesses the effect of bone marrow mesenchymal stem cells (BMSCs) exosomes miR-153 on papillary thyroid carcinoma (PTC). Adipogenesis and osteogenic induction of MSCs was performed and labeled with Cy5 labeled miR inhibitor. Cells were transfected followed by analysis of miR-153 level by real-time PCR, P-gp level by immunoblotting, and cell viability. MSCs are non-hematopoietic bone marrow-derived cells and symmetrical fibroblasts have the same characteristics as MSCs. MSCs have the potential for adipogenesis and osteogenic differentiation; miR-Cy5 can only enter PTC cells through vesicle transfer. TMZ treatment upregulated miR-153 in exosomes; MSC-derived exosomes can be directly transferred to PTC cells. miR-153-inhibitor-Cy5 can effectively inhibit miR-153 transcription and expression of resistance-related proteins. miR-153-inhibitor can promote TMZ’s effect and lead to cell death as demonstrated by increased level of active caspase-3. Inhibiting the endogenous transcription of miR-153 by miR-153 inhibitor can significantly down-regulate cell resistance protein, thereby promoting cell apoptosis under the action of TMZ.

MicroRNA‐210 repression facilitates advanced glycation end‐product (AGE)‐induced cardiac mitochondrial dysfunction and apoptosis via JNK activation

Hyperglycemia results in the formation of reactive oxygen species which in turn causes advanced glycation end products (AGEs) formation, leading to diabetic cardiomyopathy. Our previous study showed that AGE-induced reactive oxygen species-dependent apoptosis is mediated via protein kinase C delta (PKCδ)-enhanced mitochondrial damage in cardiomyocytes. By using microRNA (miRNA) database, miRNA-210 was predicted to target c-Jun N-terminal kinase (JNK), which were previously identified as downstream of PKCδ in regulating mitochondrial function. Therefore, we hypothesized that miR-210 mediates PKCδ-dependent upregulation of JNK to cause cardiac mitochondrial damage and apoptosis following AGE exposure. AGE-exposed cells showed activated cardiac JNK, PKCδ, and apoptosis, which were reversed by treatment with a JNK inhibitor and PKCδ-KD (deficient kinase). Cardiac miR-210 and mitochondrial function were downregulated following AGE exposure. Furthermore, JNK was upregulated and involved in AGE-induced mitochondrial damage. Interestingly, luciferase activity of the miR-210 mimic plus JNK WT-3'-untranslated region overexpressed group was significantly lower than that of miR-210 mimic plus JNK MT-3'UTR group, indicating that JNK is a target of miR-210. Moreover, JNK activation induced by AGEs was reduced by treatment with the miR-210 mimic and reversed by treatment with the miR-210 inhibitor, indicating the regulatory function of miR-210 in JNK activation following AGE exposure. Additionally, JNK-dependent mitochondrial dysfunction and apoptosis were reversed following treatment with the miR-210 mimic, while the miR-210 inhibitor showed no effect on JNK-induced mitochondrial dysfunction and apoptosis in AGE-exposed cardiac cells. Taken together, our study showed that PKCδ-enhanced JNK-dependent mitochondrial damage is mediated through the reduction of miR-210 in cardiomyocytes following AGE exposure.

MicroRNA-155 inhibitor ameliorates collagen-induced arthritis by modulating the phenotype of pro-inflammatory macrophages in a mouse model

Background: The present study aims to investigate the roles of microRNA-155 in collagen-induced arthritis (CIA) and its underlying mechanisms.
 Methods: CIA mouse model was established and miR155 inhibitor was intravenously injected. In in vitro studies, bone marrow-derived macrophages (BMDMs) were induced into M1 macrophages followed by the treatment of miR155 inhibitor. Quantitative reverse transcription PCR (RT-qPCR) was applied to determine the mRNA expressions. Flow cytometry was applied to determine the frequency of M1 or M2 macrophages. Western blotting was determined to detect protein expressions. Enzyme-linked immunosorbent assay (ELISA) was applied to determine the levels of inflammatory cytokines and anti-collagen antibody. 
 Results: The levels of miR155 were increased in macrophages from rheumatoid arthritis (RA) patients and M1 macrophages. The treatment of miR155 inhibitor decreased inflammatory cytokines in M1 macrophages. Besides, treatment of miR155 inhibitors promoted the differentiation of M0 macrophages into M2 macrophages. In vivo studies demonstrated that the treatment of miR155 inhibitors ameliorated the RA symptoms by decreasing inflammatory cytokines in the CIA mouse model. Treatment of miR155 also resulted in a decrease of M1 macrophage biomarker and an increase of M2 macrophage biomarker.
 Conclusion: microRNA-155 inhibitor ameliorates RA symptoms in part by regulating macrophage phenotypes.

MiR221 regulates TGF-β1-induced HSC activation through inhibiting autophagy by directly targeting LAMP2.

Liver fibrosis is a serious threat to human life and health. Activated hepatic stellate cells (HSCs) play a key role in the occurrence and development of liver fibrosis. Studies have reported that microRNAs (miRNAs/miRs) are involved in the pathological process of fibrosis, as well as its relevance in clinical diagnosis. However, the role of miR221 in hepatic fibrosis remains controversial. Remarkably, transforming growth factor-β (TGF-β1) caused HSC dysfunction in autophagic activation, characterized by an increase in P62 aggregation and LC3II expression. The present study aimed to determine whether autophagy regulates hepatic fibrosis by mediating HSC activation and explore the potential targets leading to the sequence of events associated with miR221. The expression of miR221 was quantified in a liver fibrosis model in vivo and in vitro, and its specific target gene lysosome-associated membrane glycoprotein 2 (LAMP2) was predicted by bioinformatics. The results showed that the expression levels of collagen-I (COL-I) and α-smooth muscle actin (α-SMA) were increased in miR221-overexpressing LX2 cells, while the autophagy inducer rapamycin reversed the inhibition of autophagic flux induced by miR221. Additionally, the overexpression of LAMP2 could significantly inhibit TGF-β1-induced COL-I and α-SMA expression, which was similar to the effect of the miR221 inhibitor on the regulation of TGF-β1-induced HSC activation. These results indicated that miR221 may regulate TGF-β1-induced HSC activation through inhibiting autolysosome function by directly targeting LAMP2. The molecular mechanism of miR221 in regulating TGF-β1-induced HSC activation may provide novel insight into therapies to ameliorate the pathological progression of liver fibrosis.

Mechanisms of chemotherapy‐induced muscle wasting in mice with cancer cachexia

AbstractBackgroundCachexia is a debilitating complication of cancer characterized by progressive wasting and weakness of skeletal muscles that reduces quality of life and can compromise survival. Many anticancer treatments, such as chemotherapy, also cause muscle wasting, which impairs the response to treatment. Given that many cancer patients present with cachexia at the initiation of treatment, we investigated whether cachectic mice were susceptible to chemotherapy‐induced muscle wasting and to investigate contributing mechanisms, including the dysregulation of microRNAs (miRs).MethodsCachectic colon‐26 (C‐26) tumour‐bearing mice were given 5‐fluourouracil (5‐FU) chemotherapy or vehicle treatment and analysed for muscle mass, fibre size and composition, and miR expression. Mechanisms were validated in vitro using C2C12 cell culture and miR mimics and inhibitors and were confirmed in vivo by injecting muscles of 5‐FU‐treated cachectic mice with recombinant adeno‐associated viral (rAAV) vectors encoding a miR sponge.ResultsIn cachectic tumour‐bearing mice, 5‐FU chemotherapy exacerbated the loss of skeletal muscle mass compared with vehicle treatment (by −12% to −20%, P < 0.05). miR expression profiling, quantitative real‐time PCR, and in vitro analyses revealed contributing mechanisms including miR‐351‐3p‐dependent ERK2 inhibition. Intramuscular injection of rAAV vectors encoding a sponge to reduce miR‐351‐3p expression in 5‐FU‐treated cachectic mice enhanced ERK phosphorylation (+18%, P < 0.05) and increased muscle fibre size (+15%, P < 0.01). Hsa‐miR‐125a‐3p shares similar predicted gene targets as mmu‐miR‐351‐3p, and its inhibition in human muscle cells in vitro prevented 5‐FU‐induced atrophy (P < 0.001) and increased ERK phosphorylation (P < 0.001).ConclusionsThe findings implicate miR‐351‐3p‐mediated ERK2 inhibition as a contributing mechanism in chemotherapy‐induced muscle wasting in mice with cancer cachexia and that its inhibition is a promising adjunct therapy for preserving muscles during cancer treatment.

Calcium phosphate-polymeric nanoparticle system for co-delivery of microRNA-21 inhibitor and doxorubicin

Targeted combination therapy has shown promise to achieve maximum therapeutic efficacy by overcoming drug resistance. MicroRNA-21 (miR-21) is frequently overexpressed in various cancer types including breast and non-small cell lung cancer and its functions can be inhibited by miR inhibitor (miR-21i). A combination of miR-21i and a chemo drug, doxorubicin (Dox), can provide synergistic effects. Here, we developed a calcium phosphate (CaP)-coated nanoparticle (NP) formulation to co-deliver miR-21i along with Dox. This NP design can be used to deliver the two agents with different physiochemical properties. The NP formulation was optimized for particle size, polydispersity, Dox loading, and miR-21i loading. The NP formulation was confirmed to downregulate miR-21 levels and upregulate tumor suppressor gene levels. The cytotoxic efficacy of the combined miR-21i and Dox-containing NPs was found to be higher than that of Dox. Therefore, the CaP-coated hybrid lipid-polymeric NPs hold potential for the delivery of miR-21i and Dox.

Exosomes Derived from Hydroquinone-transformed Human Bronchial Epithelial Cells Inhibited Recipient Cell Apoptosis by transferring miR-221

ObjectiveAlthough benzene is a confirmed environmental carcinogen, the mechanism of its carcinogenicity remains largely unclear. The suggested oncogene, miR-221, is elevated and plays important roles in various tumors, but its role in benzene-induced carcinogenesis remains unknown. MethodsIn the present study, we constructed hydroquinone (HQ, a representative metabolite of benzene with biological activity)-transformed malignant cell line (16HBE-t) and analyzed the level of miR-221 in it with qRT-PCR. Exosomes from 16HBE-t cells incubated with or without an miR-221 inhibitor were isolated by ultracentrifugation, characterized by transmission electron microscopy and laser scanning confocal microscope, and then transfected into 16HBE cells. The effects of exosomal miR-221 on apoptosis induced by HQ in recipient cells were determined using flow cytometry. ResultsThe amount of miR-221 in 16HBE-t was significantly increased compared with controls. When recipient cells ingested exosomes derived from 16HBE-t, miR-221 was increased, and apoptosis induced by HQ was inhibited. Blocking miR-221 in 16HBE-t using an inhibitor did not significantly alter miR-221 or apoptosis in recipient cells. ConclusionExosomal miR-221 secreted by 16HBE-t inhibits apoptosis induced by HQ in normal recipient cells.

Role of MicroRNA-155 in Triptolide-induced hepatotoxicity via the Nrf2-Dependent pathway

Ethnopharmacological relevanceTriptolide (TP), the main bioactive and toxic ingredient of Tripterygium wilfordii Hook F, causes severe toxicity, particularly for hepatotoxicity. However, the underlying mechanisms for its hepatotoxicity are not entirely clear. Aim of the studyThe purpose of the study was to explore the role of miR-155, a microRNA closely related to various liver injuries and a regulator of the nuclear factor erythroid 2-related factor 2 (Nrf2) antioxidant pathway, in TP-induced liver injury in vitro and in vivo. Materials and methodsFirst, in vitro L02 cells were treated with different concentrations of TP. The protein levels of Nrf2 and its downstream genes Heme oxygenase1 (HO-1) were determined by Western blot. The mRNA expression of miR-155, Nrf2, NAD(P)H: quinone oxidoreductase 1 (NQO1) and HO-1 were measured using qRT-PCR. And we transfected miR-155 inhibitor and miminc before TP treatment to determine the mRNA and/or protein levels of miR-155, Nrf2 and HO-1. Then, we further confirmed the interaction between miR-155 and Nrf2 pathway in TP-induced hepatic injury in BALB/C mice. The degree of liver injury was determined by HE staining and serum biochemical. The mRNA expression of miR-155 was examined with qRT-PCR and Nrf2 and HO-1 gene expression in liver were evaluated by immunohistochemistry and/or Western blot. ResultsThe results showed that TP significantly induced the expression of miR-155 both in L02 cells and in rodents liver tissue, and the inhibition of miR-155 could mitigate the hepatic damages caused by TP. Further experiments demonstrated that the inhibition of miR-155 reversed the down-regulation of Nrf2 and HO-1 by TP, while the miR-155 mimic enhanced the effects of TP. Animal experiments also showed that the inhibition of miR-155 by miR-155 antagomir reversed the decrease of Nrf2 induced by TP administration. ConclusionsThese results indicated that miR-155 played an important role in TP-induced hepatotoxicity by regulating the Nrf2 signaling pathway.

Downregulation of miR-24 Enhances Cisplatin Sensitivity in Breast Cancer Cells

GSK-3β is a tumor suppressor gene in multiple cancers by phosphorylated degrading β-catenin. Several studies showed association of miR-24 with breast cancer. Bioinformatics analysis showed a relationship of miR-24 with GSK-3β. Our study assessed miR-24’s role in GSK-3β/β-catenin siganling and breast cancer cell cisplatin resistance. MiR-24, GSK-3β, β-catenin, and Bcl-2 expressions in MDA-MB-231 and MDA-MB-231/DDP cells were detected along cell proliferation and apoptosis. DDP resistance cells were assigned into miR-NC, miR-24 inhibitor, pIRES-blank, pIRES-GSK-3β, and miR-24 inhibitor+pIRES-GSK-3β groups and cell proliferation was determined. MiR-24 inhibited GSK-3β level. GSK-3β and cell apoptosis significantly downregulated, while miR-24, β-catenin, Bcl-2, and cell proliferation significantly elevated in DDP resistance cells. MiR-24 inhibitor and/or pIRES-GSK-3β significantly increased GSK-3β level, declined β-catenin and Bcl-2 expressions, attenuated cell proliferation, enhanced cell apoptosis, and weakened cisplatin resistance. MiR-24 upregulation was related to breast cancer cell cisplatin resistance. Inhibition of miR-24 upregulated GSK-3β, restrained Wnt/β-catenin signaling and cisplatin resistance in breast cancer cells.

Polycyclic aromatic hydrocarbons induce endothelial injury through miR‐155 to promote atherosclerosis

Polycyclic aromatic hydrocarbons (PAHs) are considered as an external factor that induces atherosclerotic cardiovascular disease. Although miR-155 is known to be involved in cardiovascular disease, whether it is involved in PAH-induced arteriosclerosis remains unclear. We evaluated the effects of PAHs on vascularization, permeability, and miR-155 expression in HUVECs. We found that PAHs-induced sclerosis of HUVECs was characterized by increasing permeability, decreasing proliferation, and vascular lumen number. The expression of miR-155 was upregulated by PAHs treatment, and transfection with miR-155 inhibitor could reverse above effect of PAHs-induced sclerosis. Meanwhile, transcriptome sequencing revealed that 63 genes were downregulated in the group of PAHs treatment alone, and were then upregulated in the miR-155 inhibitor group. These genes were mainly involved in complement and coagulation cascades, cytokine-cytokine receptor interaction, TNF signaling pathway, and NF-kappa B signaling pathway. Among these 63 genes, SERPIND1 was directly targeted and regulated by miR-155. Further in vivo experiments in ApoE-/- mice confirmed that PAH accelerates the development of arteriosclerosis by promoting the expression of miR-155 to downregulate the SERPIND1. Therefore, PAH exaggerates atherosclerosis by activating miR-155-dependent endothelial injury. This study provides a fundamental insight on the miR-155 mechanism for PAHs enhancing atherosclerosis and miR-155 potentially serving as a novel drug target.

Mechanism of MIR-210 mediated NF-κB pathway on cardiac ischemia-reperfusion injury.

In this study, MicroRNA-210 (miR-210), which was previously proved to be a potential immunomodulator in various disease, attenuated mouse myocardium ischemia/reperfusion (I/R) injury. miR-210 was increased in cardiomyocytes exposed to hypoxia/reoxygenation (H/R). The expression of IL-6 and TNF-α in both serum and supernatant were reduced in miR-210 mimics groups. Mice were randomly divided into four groups, which were pre-treated with saline (sham and ischemia/reperfusion group), miR-210 mimics and miR-210 inhibitor treatments. Three days later, the mouse IR model was established by ischemia for 30 min, followed by reperfusion for 3 h. Myocardium and plasma were harvested and assessed. The myocardium histopathological changes were reduced in miR-210 mimics groups, and serum levels of Creatine kinase isoenzyme (CK-MB) and Lactate dehydrogenase (LDH) were significantly decreased compared with I/R groups. The protein expression of proinflammatory factor interleukin (IL)-1β and IL-6 were suppressed by the up-regulation of miR-210. The expression of miR-210 was negatively correlated with the expression of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB). In conclusion, our study indicates that miR-210 protects heart from myocardium I/R injury via suppressing NF-κB signal pathway.

MiR-155 regulates Th9 differentiation in children with methicillin-resistant Staphylococcus aureus pneumonia by targeting SIRT1

Th9 is a subset of CD4+ T cells that mainly secrete IL-9. Th9/IL-9 participates in immune response during Staphylococcus aureus and methicillin-resistant Staphylococcus aureus pneumonia (MRSA) infection. Here, we collected bronchoalveolar lavage fluid (BALF) from 30 children with MRSA pneumonia (MRSA group) and 10 children with bronchial foreign bodies (Control group). RT-PCR, ELISA and flow cytometry were used to detect the expression of miR-155 and IL-9 in BALF and the number of Th9 cells. CD4+ T cells isolated from BALF of MRSA and Control group were transfected with miR-155 mimic or inhibitor, and then induced Th9 cell differentiation. The results showed that the expression of miR-155 and IL-9 were significantly increased in BALF and Th9 cell of MRSA group, as well as the number of Th9 cells. miR-155 mimic upregulated IL-9 mRNA expression, IL-9 secretion and increased number of Th9 cells. On the contrary, miR-155 inhibitor inhibited IL-9 mRNA expression, IL-9 secretion and decreased number of Th9 cells. The dual luciferase assays demonstrated miR-155 can target binding to SIRT1 3′UTR. Moreover, overexpression of SIRT1 could reverse the effect of miR-155 mimic on IL-9 expression level, Th9 cell number and transcription factors PU.1 and IRF4 expression. In conclusion, miR-155 regulates Th9 differentiation in children with MRSA by targeting SIRT1.

Overexpression of miR-122 Impairs Intestinal Barrier Function and Aggravates Acute Pancreatitis by Downregulating Occludin Expression.

Acute pancreatitis (AP) causes intestinal barrier damage, resulting in systemic inflammatory response syndrome (SIRS) or multiple organ dysfunction syndrome (MODS), which are important factors affecting AP severity and mortality. Here, we studied the mechanism of miR-122 in regulating intestinal barrier function in AP. AP rat model was constructed via intraperitoneal injection of ketamine, and primary intestinal epithelial cells were isolated from rats for in vitro studies. HE staining was used to assess pathological alterations of pancreas and intestines tissues. Inflammatory factors were detected by ELISA assay. qRT-PCR and WB were used to detect the expressions of miR-122 and occluding, respectively. Then dual-luciferase reporter assay, intestinal permeability test, and cell permeability were performed in vivo and in vitro to probe the molecular mechanism of miR-122 in regulating intestinal barrier function in AP. The expression of miR-122 was upregulated in AP rats, while the expression of occludin was downregulated, and the intestinal permeability was increased in AP rats and primary intestinal epithelial cells isolated from rats. Inhibition of miR-122 regulated intestinal barrier function through mediating occludin expression. miR-122 regulated intestinal barrier function to affect AP through mediating occludin expression in vivo. These results provided evidence that miR-122 overexpression impaired intestinal barrier function via regulation of occludin expression, thus promoting AP progression.

MiR-221/222 sponge abrogates tamoxifen resistance in ER-positive breast cancer cells through restoring the expression of ER\u03b1

Tamoxifen resistance (TamR) prevents ER-positive breast cancer patients from benefitting from endocrine therapy, and miR-221 or miR-222 plays vital roles in inducing TamR. In this study, we designed synthetic sponges to reverse TamR by targeting these two miRs. First, we established a tamoxifen resistant breast cancer cell line (MCF-7TamR), we verified the high expressing level of these two miRs in TamR cells. miR-221 or miR-222 inhibitors rendered MCF-7TamR cells responsive to tamoxifen. Next, we designed a miR-221/222 sponge, which contains total 8 multi-antisense binding sites (MBSs) for these two onco-miRs, and inserted it into CMV promoter- or hTERT promoter-driven expressing vectors. After transfected miR-221/222 sponge expressing vectors into MCF-7TamR cells, we identified a strong interaction between miR-221/222 sponge and endogenous miR-221 or miR-222 by RNA pulldown assay. We also found that miR-221/222 sponge restored the expression of ERα and PTEN, arrested cells in G1 phase, and finally resulted in reduced cell growth and cell migration. Notably, miR-221/222 sponge expressing cells abrogates tamoxifen resistance through restoring the expression of ERα, suggesting that miR-221/222 sponge gene therapy especially driven by tumor specific promoter could provide an effective therapeutic approach against TamR in breast cancer.

Inhibition of miR-222 by Oncolytic Adenovirus-Encoded miRNA Sponges Promotes Viral Oncolysis and Elicits Antitumor Effects in Pancreatic Cancer Models.

Simple SummaryOncolytic adenoviruses are replication-competent viruses engineered for use in cancer treatment. However, the transformation of cancer cells defines a scenario in which the viruses are not adapted to replicate. During tumorigenesis, a large number of miRNAs are deregulated, modulating the expression of genes with essential roles in carcinogenesis and with potential effects on adenoviral propagation. Here we show how OncomiRs can impact adenoviral activity, identifying miR-222 as a limiting factor. Of notice, by genetically engineering a therapeutic adenovirus with miR-222 sponges (AdNuPAR-E-miR222-S), we reduced the latter miRNA content, while enhancing adenoviral fitness, increasing cytotoxicity, and sustaining tumor growth control in xenografts.Oncolytic adenoviruses (OA) are envisioned as a therapeutic option for patients with cancer, designed to preferentially replicate in cancer cells. However, the high number of genetic alterations in tumors can generate a context in which adenoviruses have difficulties replicating. Abnormal miRNAs expression is a trademark of pancreatic cancer, with several oncogenic miRNAs playing essential roles in cancer-associated pathways. The perturbed miRNome induces reprogramming of gene expression in host cells that can impact the complex interplay between cellular processes and viral replication. We have studied the effects of overexpressed miRNAs on oncolytic adenoviral activity and identified miRNAs modulators of adenoviral oncolysis in pancreatic cancer cells. Inhibition of the highly upregulated miR-222 sensitized cancer cells to oncolysis. To provide a therapeutic application to this insight, we engineered the oncolytic adenovirus AdNuPARmE1A with miR-222 binding sites, working as sponges to withdraw the miRNA from the cellular environment. AdNuPAR-E-miR222-S mediated-decrease of miR-222 expression in pancreatic cancer cells strongly improved the viral yield and enhanced the adenoviral cytotoxic effects. Antitumoral studies confirmed a high activity for AdNuPARmE1A-miR222-S in vivo, controlling tumor progression more effectively than the scrambled control virus in xenografts. We demonstrated that the increased antitumor potency of the novel oncolytic virus resulted from the combinatory effects of miR-222 oncomiR inhibition and the restoration of miR-222 target genes activity enhancing viral fitness.

MiR-155 Inhibitor-Laden Exosomes Reverse Resistance to Cisplatin in a 3D Tumor Spheroid and Xenograft Model of Oral Cancer.

Cisplatin resistance is one of the major concerns in the treatment of oral squamous cell carcinoma (OSCC). Accumulating evidence suggests microRNA (miRNA) dysregulation as one of the mediators of chemoresistance. Toward this, our previous study revealed the role of exosomal microRNA-155 (miR-155) in cisplatin resistance via downregulation of FOXO3a, a direct target of miR-155, and induction of epithelial-to-mesenchymal transition in OSCC. In the present study, we demonstrate the therapeutic potential of miR-155 inhibitor-laden exosomes in the sensitization of a cisplatin-resistant (cisRes) OSCC 3D tumor spheroid and xenograft mouse model. The cisRes OSSC 3D tumor spheroid model recapitulated the hallmarks of solid tumors such as enhanced hypoxia, reactive oxygen species, and secretory vascular endothelial growth factor. Further treatment with miR-155 inhibitor-loaded exosomes showed the upregulation of FOXO3a and induction of the mesenchymal-to-epithelial transition with improved sensitization to cisplatin in cisRes tumor spheroids and xenograft mouse model. Moreover, the exosomal miR-155 inhibitor suppressed the stem-cell-like property as well as drug efflux transporter protein expression in cisplatin-resistant tumors. Taken together, our findings, for the first time, established that the miR-155 inhibitor-loaded exosomes reverse chemoresistance in oral cancer, thereby providing an alternative therapeutic strategy for the management of refractory oral cancer patients.

MiRNA Changes in Retinal Ganglion Cells after Optic Nerve Crush and Glaucomatous Damage.

The purpose of this study was to characterize the miRNA profile of purified retinal ganglion cells (RGC) from healthy and diseased rat retina. Diseased retina includes those after a traumatic optic nerve crush (ONC), and after ocular hypertension/glaucoma. Rats were separated into four groups: healthy/intact, 7 days after laser-induced ocular hypertension, 2 days after traumatic ONC, and 7 days after ONC. RGC were purified from rat retina using microbeads conjugated to CD90.1/Thy1. RNA were sequenced using Next Generation Sequencing. Over 100 miRNA were identified that were significantly different in diseased retina compared to healthy retina. Considerable differences were seen in the miRNA expression of RGC 7 days after ONC, whereas after 2 days, few changes were seen. The miRNA profiles of RGC 7 days after ONC and 7 days after ocular hypertension were similar, but discrete miRNA differences were still seen. Candidate mRNA showing different levels of expression after retinal injury were manipulated in RGC cultures using mimics/AntagomiRs. Of the five candidate miRNA identified and subsequently tested for therapeutic efficacy, miR-194 inhibitor and miR-664-2 inhibitor elicited significant RGC neuroprotection, whereas miR-181a mimic and miR-181d-5p mimic elicited significant RGC neuritogenesis.

Involvement of microRNA-155 in the mechanism of electroacupuncture treatment effects on experimental autoimmune encephalomyelitis

Multiple sclerosis (MS) is a neurodegenerative and demyelinating autoimmune disease mediated by autoreactive T cells that affects the central nervous system (CNS). Electroacupuncture (EA) has emerged as an alternative or supplemental treatment for MS, but the mechanism by which EA may alleviate MS symptoms is unresolved. Here, we examined the effects of EA at the Zusanli (ST36) acupoint on mice with experimental autoimmune encephalomyelitis (EAE), the predominant animal model of MS. The effects of EA on EAE emergence, inflammatory cell levels, proinflammatory cytokines, and spinal cord pathology were examined. EA treatment attenuated the EAE clinical score and associated spinal cord demyelination, while reducing the presence of proinflammatory cytokines in mononuclear cells (MNCs), downregulating microRNA (miR)-155, and upregulating the opioid peptide precursor proopiomelanocortin (POMC) in the CNS. Experiments in which cultured neurons were transfected with a miR-155 mimic or a miR-155 inhibitor further showed that the direct modulation of miR-155 levels could regulate POMC levels in neurons. In conclusion, the alleviation of EAE by EA is characterized by reduced proportions of Th1/Th17 cells and increased proportions of Th2 cells, POMC upregulation, and miR-155 downregulation, while miR-155 itself can suppress POMC expression. These results, support the hypothesis that the effects of EA on EAE may involve the downregulation of miR-155.