Fecal let-7b and miR-21 directly modulate the intestinal microbiota, driving chronic inflammation.

Inflammatory bowel diseases (IBD) etiology is multifactorial. Luminal microRNAs (miRNAs) have been suspected to play a role in the promotion of chronic inflammation, but the extent to which fecal miRNAs are interacting with the intestinal ecosystem in a way that contribute to diseases, including IBD, remains unknown. Here, fecal let-7b and miR-21 were found elevated, associated with inflammation, and correlating with multiple bacteria in IBD patients and IL-10-/- mice, model of spontaneous colitis. Using an in vitro microbiota modeling system, we revealed that these two miRNAs can directly modify the composition and function of complex human microbiota, increasing their proinflammatory potential. In vivo investigations revealed that luminal increase of let-7b drastically alters the intestinal microbiota and enhances macrophages' associated proinflammatory cytokines (TNF, IL-6, and IL-1ß). Such proinflammatory effects are resilient and dependent on the bacterial presence. Moreover, we identified that besides impairing the intestinal barrier function, miR-21 increases myeloperoxidase and antimicrobial peptides secretion, causing intestinal dysbiosis. More importantly, in vivo inhibition of let-7b and miR-21 with anti-miRNAs significantly improved the intestinal mucosal barrier function and promoted a healthier host-microbiota interaction in the intestinal lining, which altogether conferred protection against colitis. In summary, we provide evidence of the functional significance of fecal miRNAs in host-microbiota communication, highlighting their therapeutic potential in intestinal inflammation and dysbiosis-related conditions, such as IBD.

The Role of the MiR-181 Family in Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is the fourth-leading cause of cancer-related death worldwide. Due to the high mortality rate in HCC patients, discovering and developing novel systemic treatment options for HCC is a vital unmet medical need. Among the numerous molecular alterations in HCCs, microRNAs (miRNAs) have been increasingly recognised to play critical roles in hepatocarcinogenesis. We and others have recently revealed that members of the microRNA-181 (miR-181) family were up-regulated in some, though not all, human cirrhotic and HCC tissues-this up-regulation induced epithelial-mesenchymal transition (EMT) in hepatocytes and tumour cells, promoting HCC progression. MiR-181s play crucial roles in governing the fate and function of various cells, such as endothelial cells, immune cells, and tumour cells. Previous reviews have extensively covered these aspects in detail. This review aims to give some insights into miR-181s, their targets and roles in modulating signal transduction pathways, factors regulating miR-181 expression and function, and their roles in HCC.

Anti-miRNA103/107 encapsulated in transferrin-conjugated lipid nanoparticles crosses blood-brain barrier and reduces brain ischemic damage.

MicroRNA (miRNA), by post-transcriptionally regulating the expression of genes involved in stroke response, represents important effectors in stroke pathophysiology. Recently, the 103/107 miRNA family emerged as a possible therapeutic target in stroke, as it controls the expression of sodium calcium exchanger 1, a plasma membrane transporter that plays a fundamental role in stroke pathophysiology. Although the neuroprotective properties of this and other miRNAs are promising, several pharmacokinetic drawbacks remain to be faced for the development of a translatable therapy based on small RNAs in CNS diseases. In the present study, to overcome these limitations, the anti-miRNA103/107 was encapsulated in specific preparations of lipid nanoparticles (LNPs), and their effectiveness was evaluated both in an in vitro model of hypoxia represented by primary neuronal cortical cultures exposed to oxygen and glucose deprivation followed by reoxygenation, and in an in vivo model of stroke obtained in rats exposed to transient occlusion of the middle cerebral artery. The results of the present study demonstrated that the encapsulation of anti-miRNA103/107 in transferrin-conjugated PEG-stabilized LNPs allowed the blood-brain barrier crossing and significantly reduced brain ischemic damage. The present achievements pave the way for the exploitation of a systemic intravenous miRNA delivery strategy in stroke therapy.

Use of Therapeutic RNAs to Accelerate Wound Healing in Diabetic Rabbit Wounds.

Introduction: Diabetes mellitus (DM) affects over 422 million people globally. Patients with DM are subject to a myriad of complications, of which diabetic foot ulcers (DFUs) are the most common with ∼25% chance of developing these wounds throughout their lifetime. Innovation: Currently there are no therapeutic RNAs approved for use in DFUs. Use of dressings containing novel layer-by-layer (LbL)-formulated therapeutic RNAs that inhibit PHD2 and miR-210 can significantly improve diabetic wound healing. These dressings provide sustained release of therapeutic RNAs to the wounds locally without systemic side effects. Clinical Problem Addressed: Diabetic foot wounds are difficult to heal and often result in significant patient morbidity and mortality. Materials and Methods: We used the diabetic neuroischemic rabbit model of impaired wound healing. Diabetes was induced in the rabbits with alloxan, and neuroischemia was induced by ligating the central neurovascular bundle of each ear. Four 6-mm full-thickness wounds were created on each ear. A LbL technique was used to conformally coat the wound dressings with chemically modified RNAs, including an antisense oligonucleotide (antimiR) targeting microRNA-210 (miR-210), an short synthetic hairpin RNA (sshRNA) targeting PHD2, or both. Results: Wound healing was improved by the antimiR-210 but not the PHD2-sshRNA. Specific knockdown of miR-210 in tissue as measured by RT-qPCR was ∼8 Ct greater than nonspecific controls, and this apparent level of knockdown (>99%) suggests that delivery to the tissue is highly efficient at the administered dose. Discussion: Healing of ischemic/neuropathic wounds in diabetic rabbits was accelerated upon inhibition of miR-210 by LbL delivery to the wound bed. miR-210 inhibition was achieved using a chemically modified antisense RNA.

Comparison of interaction between antimiR and microRNA versus HDO-antimiR and microRNA by molecular dynamics simulation.

Recently, we found DNA/RNA heteroduplex oligonucleotide-based antimiR (HDO-antimiR) can more efficiently inhibit the target miRNA than conventional antimiR after its cellular uptake. But the mechanism of HDO-antimiR about the target-silencing is unknown. We here tried to elucidate the interaction mechanism of HDO-antimiR to miRNA using molecular dynamics (MD) simulation. When interaction of the conventional antimiR or HDO-antimiR and the target miRNA was simulated, they combined with each other in various forms. In the hydrogen bond analyses, base site of the antimiR formed hydrogen bond with miRNA. On the other hand, phosphate site of the HDO-antimiR formed hydrogen bond with miRNA. These results suggested that there were differences about the binding mechanisms between antimiR and HDO-antimiR to the target miRNA. In particular, there was a difference in the binding site between antimiR and HDO-antimiR. Additionally, it was found that guanine in the miRNA is mainly involved in the binding to the antimiR or HDO-antimiR. MD simulation method is useful in understanding the mechanism of oligonucleotide therapeutics.

Modulation of miR-29 influences myocardial compliance likely through coordinated regulation of calcium handling and extracellular matrix.

MicroRNAs (miRNAs) control the expression of diverse subsets of target mRNAs, and studies have found miRNA dysregulation in failing hearts. Expression of miR-29 is abundant in heart, increases with aging, and is altered in cardiomyopathies. Prior studies demonstrate that miR-29 reduction via genetic knockout or pharmacologic blockade can blunt cardiac hypertrophy and fibrosis in mice. Surprisingly, this depended on specifically blunting miR-29 actions in cardiomyocytes versus fibroblasts. To begin developing more translationally relevant vectors, we generated a novel transgene-encoded miR-29 inhibitor (TuD-29) that can be incorporated into a viral-mediated gene therapy for cardioprotection. Here, we corroborate that miR-29 expression and activity is higher in cardiomyocytes versus fibroblasts and demonstrate that TuD-29 effectively blunts hypertrophic responses in cultured cardiomyocytes and mouse hearts. Furthermore, we found that adeno-associated virus (AAV)-mediated miR-29 overexpression in mouse hearts induces early diastolic dysfunction, whereas AAV:TuD-29 treatment improves cardiac output by increasing end-diastolic and stroke volumes. The integration of RNA sequencing and miRNA-target interactomes reveals that miR-29 regulates genes involved in calcium handling, cell stress and hypertrophy, metabolism, ion transport, and extracellular matrix remodeling. These investigations support a likely versatile role for miR-29 in influencing myocardial compliance and relaxation, potentially providing a unique therapeutic avenue to improve diastolic function in heart failure patients.

pH-Sensitive Nanoparticles for Colonic Delivery Anti-miR-301a in Mouse Models of Inflammatory Bowel Diseases.

Though the anti-miR-301a (anti-miR) is a promising treatment strategy for inflammatory bowel disease (IBD), the degradability and the poor targeting of the intestine are a familiar issue. This study aimed to develop a multifunctional oral nanoparticle delivery system loaded with anti-miR for improving the targeting ability and the therapeutic efficacy. The HA-CS/ES100/PLGA nanoparticles (HCeP NPs) were prepared using poly (lactic-co-glycolic acid) copolymer (PLGA), enteric material Eudragit®S100 (ES100), chitosan (CS), and hyaluronic acid (HA). The toxicity of nanoparticles was investigated via the Cell Counting Kit-8, and the cellular uptake and inflammatory factors of nanoparticles were further studied. Moreover, we documented the colon targeting and pharmacodynamic properties of nanoparticles. The nanoparticles with uniform particle size exhibited pH-sensitive release, favorable gene protection, and storage stability. Cytology experiments showed that anti-miR@HCeP NPs improved the cellular uptake through HA and reduced pro-inflammatory factors. Administering anti-miR@HCeP NPs orally to IBD mice markedly reduced their pro-inflammatory factors levels and disease activity indices. We also confirmed that anti-miR@HCeP NPs mostly accumulated in the colon site, and effectively repaired the intestinal barrier, as well as relieved intestinal inflammation. The above nanoparticle is a candidate of the treatment for IBD due to its anti-inflammatory properties.

Crosstalk between Non-Coding RNAs and Wnt/ß-Catenin Signaling in Head and Neck Cancer: Identification of Novel Biomarkers and Therapeutic Agents.

Head and neck cancers (HNC) encompass a broad spectrum of neoplastic disorders characterized by significant morbidity and mortality. While contemporary therapeutic interventions offer promise, challenges persist due to tumor recurrence and metastasis. Central to HNC pathogenesis is the aberration in numerous signaling cascades. Prominently, the Wnt signaling pathway has been critically implicated in the etiology of HNC, as supported by a plethora of research. Equally important, variations in the expression of non-coding RNAs (ncRNAs) have been identified to modulate key cancer phenotypes such as cellular proliferation, epithelial-mesenchymal transition, metastatic potential, recurrence, and treatment resistance. This review aims to provide an exhaustive insight into the multifaceted influence of ncRNAs on HNC, with specific emphasis on their interactions with the Wnt/ß-catenin (WBC) signaling axis. We further delineate the effect of ncRNAs in either exacerbating or attenuating HNC progression via interference with WBC signaling. An overview of the mechanisms underlying the interplay between ncRNAs and WBC signaling is also presented. In addition, we described the potential of various ncRNAs in enhancing the efficacy of chemotherapeutic and radiotherapeutic modalities. In summary, this assessment posits the potential of ncRNAs as therapeutic agents targeting the WBC signaling pathway in HNC management.

Targeted Expression to Liver of an antimiR-33 Sponge as a Gene Therapy Strategy against Hypercholesterolemia: In Vitro Study.

Atherosclerosis is the leading cause of cardiovascular diseases in Mexico and worldwide. The membrane transporters ABCA1 and ABCG1 are involved in the reverse transport of cholesterol and stimulate the HDL synthesis in hepatocytes, therefore the deficiency of these transporters promotes the acceleration of atherosclerosis. MicroRNA-33 (miR-33) plays an important role in lipid metabolism and exerts a negative regulation on the transporters ABCA1 and ABCG1. It is known that by inhibiting the function of miR-33 with antisense RNA, HDL levels increase and atherogenic risk decreases. Therefore, in this work, a genetic construct, pPEPCK-antimiR-33-IRES2-EGFP, containing a specific antimiR-33 sponge with two binding sites for miR-33 governed under the PEPCK promoter was designed, constructed, and characterized, the identity of which was confirmed by enzymatic restriction, PCR, and sequencing. Hep G2 and Hek 293 FT cell lines, as well as a mouse hepatocyte primary cell culture were transfected with this plasmid construction showing expression specificity of the PEPCK promoter in hepatic cells. An analysis of the relative expression of miR-33 target messengers showed that the antimiR-33 sponge indirectly induces the expression of its target messengers (ABCA1 and ABCG1). This strategy could open new specific therapeutic options for hypercholesterolemia and atherosclerosis, by blocking the miR-33 specifically in hepatocytes.

Therapeutic inhibition of miR-155 attenuates liver fibrosis via STAT3 signaling.

Most chronic liver diseases progress to liver fibrosis, which, when left untreated, can lead to cirrhosis and hepatocellular carcinoma. MicroRNA (miRNA)-targeted therapeutics have become attractive approaches to treat diseases. In this study, we investigated the therapeutic effect of miR-155 inhibition in the bile duct ligation (BDL) mouse model of liver fibrosis and evaluated the role of miR-155 in chronic liver fibrosis using miR-155-deficient (miR-155 knockout [KO]) mice. We found increased hepatic miR-155 expression in patients with cirrhosis and in the BDL- and CCl4-induced mouse models of liver fibrosis. Liver fibrosis was significantly reduced in miR-155 KO mice after CCl4 administration or BDL. To assess the therapeutic potential of miR-155 inhibition, we administered an rAAV8-anti-miR-155 tough decoy in vivo that significantly reduced liver damage and fibrosis in BDL. BDL-induced protein levels of transforming growth factor ß (TGF-ß), p-SMAD2/3, and p-STAT3 were attenuated in anti-miR-155-treated compared with control mice. Hepatic stellate cells from miR-155 KO mice showed attenuation in activation and mesenchymal marker expression. In vitro, miR-155 gain- and loss-of-function studies revealed that miR-155 regulates activation of stellate cells partly via STAT3 signaling. Our study suggests that miR-155 is the key regulator of liver fibrosis and might be a potential therapeutic target to attenuate fibrosis progression.

Aptamer functionalized nucleic acid nano drug for targeted synergistic therapy for colon cancer.

Due to its complicated pathophysiology, propensity for metastasis, and poor prognosis, colon cancer is challenging to treat and must be managed with a combination of therapy. Using rolling circle transcription (RCT), this work created a nanosponge therapeutic medication system (AS1411@antimiR-21@Dox). Using the AS1411 aptamer, this approach accomplished targeted delivery to cancer cells. Furthermore, analysis of cell viability, cell apoptosis, cell cycle arrest, reactive oxygen species (ROS) content, and mitochondrial membrane potential (MMP) levels revealed that functional nucleic acid nanosponge drug (FND) can kill cancer cells. Moreover, transcriptomics uncovered a putative mechanism for the FND anti-tumor effect. These pathways, which included mitotic metaphase and anaphase as well as the SMAC-mediated dissociation of the IAP: caspase complexes, were principally linked to the cell cycle and cell death. In conclusion, by triggering cell cycle arrest and apoptosis, the nano-synergistic therapeutic system allowed for the intelligent and effective targeted administration of RNA and chemotherapeutic medicines for colon cancer treatment. The system allowed for payload efficiency while being customizable, targeted, reliable, stable, and affordable.

Combined Treatment of Cancer Cells Using Allyl Palladium Complexes Bearing Purine-Based NHC Ligands and Molecules Targeting MicroRNAs miR-221-3p and miR-222-3p: Synergistic Effects on Apoptosis.

Combined treatments employing lower concentrations of different drugs are used and studied to develop new and more effective anticancer therapeutic approaches. The combination therapy could be of great interest in the controlling of cancer. Regarding this, our research group has recently shown that peptide nucleic acids (PNAs) that target miR-221 are very effective and functional in inducing apoptosis of many tumor cells, including glioblastoma and colon cancer cells. Moreover, in a recent paper, we described a series of new palladium allyl complexes showing a strong antiproliferative activity on different tumor cell lines. The present study was aimed to analyze and validate the biological effects of the most active compounds tested, in combination with antagomiRNA molecules targeting two miRNAs, miR-221-3p and miR-222-3p. The obtained results show that a "combination therapy", produced by combining the antagomiRNAs targeting miR-221-3p, miR-222-3p and the palladium allyl complex 4d, is very effective in inducing apoptosis, supporting the concept that the combination treatment of cancer cells with antagomiRNAs targeting a specific upregulated oncomiRNAs (in this study miR-221-3p and miR-222-3p) and metal-based compounds represents a promising therapeutic strategy to increase the efficacy of the antitumor protocol, reducing side effects at the same time.

Zinc treatment reverses and anti-Zn-regulated miRs suppress esophageal carcinomas in vivo.

Esophageal squamous cell carcinoma (ESCC) is a deadly disease with few prevention or treatment options. ESCC development in humans and rodents is associated with Zn deficiency (ZD), inflammation, and overexpression of oncogenic microRNAs: miR-31 and miR-21. In a ZD-promoted ESCC rat model with upregulation of these miRs, systemic antimiR-31 suppresses the miR-31-EGLN3/STK40-NF-κB-controlled inflammatory pathway and ESCC. In this model, systemic delivery of Zn-regulated antimiR-31, followed by antimiR-21, restored expression of tumor-suppressor proteins targeted by these specific miRs: STK40/EGLN3 (miR-31), PDCD4 (miR-21), suppressing inflammation, promoting apoptosis, and inhibiting ESCC development. Moreover, ESCC-bearing Zn-deficient (ZD) rats receiving Zn medication showed a 47% decrease in ESCC incidence vs. Zn-untreated controls. Zn treatment eliminated ESCCs by affecting a spectrum of biological processes that included downregulation of expression of the two miRs and miR-31-controlled inflammatory pathway, stimulation of miR-21-PDCD4 axis apoptosis, and reversal of the ESCC metabolome: with decrease in putrescine, increase in glucose, accompanied by downregulation of metabolite enzymes ODC and HK2. Thus, Zn treatment or miR-31/21 silencing are effective therapeutic strategies for ESCC in this rodent model and should be examined in the human counterpart exhibiting the same biological processes.

Osteoclast-targeted delivery of anti-miRNA oligonucleotides by red blood cell extracellular vesicles.

Osteoporosis (OP) affects millions worldwide but currently cannot be cured. Suppressing the level of miR-214 in osteoclasts by the anti-miRNA oligonucleotide (AMO) anti-miR-214 reverses bone absorption and provides a potential treatment. Here we report a peptide-guided delivery strategy using red blood cell extracellular vesicles (RBCEVs) as the vehicle to realize osteoclast-targeted delivery of anti-miR-214. A bi-functional peptide, TBP-CP05, which binds to both the CD63 on RBCEVs and receptors on osteoclasts, acts as the guide. TBP-CP05 binds with RBCEVs through CP05, displays the TRAP-binding peptide (TBP) on the surface of EVs, and endows RBCEVs with osteoclast-targeting capability both in vitro and in vivo. Intravenous injection of the osteoclast-targeting RBCEVs (OT-RBCEVs) led to the enrichment of EVs in the bone skeleton, significant inhibition of the osteoclast activity, elevated osteoblast activity, and improved bone density in osteoporotic mice. Altogether, this work demonstrates efficient guidance of drug-loaded EVs to the targeted cells in vivo using bi-functional fusion peptides, and showcases that targeted delivery of anti-miR-214 by OT-RBCEVs may be a viable method for OP treatment. SIGNIFICANCE STATEMENT. Surface functionalization of EVs endows these nanovesicles cell-specific targeting property which guides the drug cargos to specific tissues and cells with higher accuracy, longer retention, and minimal off-target effects. Methods to functionalize EVs with minimal procedures are highly desired for clinical applications. Here we present a facile method using a bifunctional fusion peptide to guide RBCEVs to osteoclasts. A simple incubation of the bifunctional peptide and RBCEVs results in osteoclast-targeting RBCEVs (OT-RBCEVs) that effectively deliver anti-miR-214 to osteoclasts in vivo in a mouse model of osteoporosis, bringing a potential therapy to osteoporotic patients. This is, to our knowledge, the first report on peptide functionalization of RBCEVs and osteoclast-targeted delivery using RBCEVs.

MicroRNA-147b induces neuroendocrine differentiation of prostate cancer cells by targeting ribosomal protein RPS15A.

BACKGROUND: Prostate cancer (PCa) is the leading cause of cancer related deaths in men, often androgen deprivation therapy (ADT) leads to the progression of androgen independent PCa (AIPC) which further leads to Neuroendocrine PCa (NEPC). Identifying the molecular mechanisms which navigate the neuroendocrine differentiation (NED) of PCa cells is clinically relevant. It has been suggested that the micro RNAs (miRNAs) play an important role in the regulation of intrinsic mechanisms relevant to tumor progression, resistance as a result leads to poor prognosis. miR-147b has been transpiring as one of the deregulated miRNAs associated with the occurrence of multiple cancers. The present study has studied the role of miRNA-147b in inducing NEPC. METHODS: To investigate the functional role of miR-147b in NEPC, we have expressed miRNA mimics or inhibitors in PCa cells and monitored the progression of NEPC along with PCa cell proliferation and survival. The molecular mechanism miRNA-147b follows was studied using western blot and reverse transcription polymerase chain analysis. miRNA target prediction using bioinformatics tools followed by target validation using luciferase reporter assays was performed. RESULTS: In the present study, we found that miR-147b is highly expressed in AIPC cell lines in particular neuroendocrine cells NCI-H660 and NE-LNCaP derived from LNCaP. Mechanistic studies revealed that overexpression of miR-147b or miRNA mimics induced NED in LNCaP cells in in-vitro while its inhibitor reversed the NE features (increased NE markers and reduced prostate specific antigen) of PC3, NCI-H660 and NE-LNCaP cells. In addition, miR-147b reduced the proliferation rate of LNCaP cells via elevated p27kip1 and lowered cyclin D1 for promoting differentiation. In reporter assays, we have identified ribosomal protein S15A (RPS15A) is a direct target of miRNA-147b and RPS15A expression was negatively regulated by miR-147b in PCa cells. Furthermore, we also report that RPS15A is downregulated in NEPC cells and its expression is inversely correlated with NE markers. CONCLUSION: Targeting the miR-147b - RPS15A axis may overcome the progression of NEPC and serve as a novel therapeutic target to attenuate NED progression of PCa.

Inhibition of microRNA-328 Increases Ocular Mucin Expression and Conjunctival Goblet Cells.

We previously reported anti-miR-328 therapy for dry eye disease (DED). Since decreased mucin secretion is a risk factor for DED, we aimed to explore whether anti-miR-328 affects mucin expression and goblet cells. MiR-328 was increased in goblet cells when they were under desiccating stress or treated with benzalkonium chloride (BAC), both of which are risk factors for DED. Based on bioinformatics tool results, miR-328 was predicted to directly target the transcription factor CREB1 that has been known to promote the expression of mucin5AC. The inhibitory effect of miR-328 on CREB1 was confirmed by the transfection assay. A miR-328 binding site on the CREB1 gene was confirmed by the luciferase assay. Furthermore, anti-miR-328 increased CREB1 and mucin5AC in cultured goblet cells according to qPCR, Western blot, and IF staining experiments. Anti-miR-328 increased mucin5AC secretion from the cultured goblet cells based on an ELISA assay for the cultured medium. Finally, impression cytology data revealed anti-miR-328 increased conjunctival goblet cells in the DED rabbits induced by BAC. In conclusion, anti-miR-328 increases CREB1 expression leading to an increase in mucin5AC production and secretion. Furthermore, anti-miR-328 also increases conjunctival goblet cells. These results warrant the further development of anti-miR-328 therapy for DED.

Inhibition of miR-101-3p prevents human aortic valve interstitial cell calcification through regulation of CDH11/SOX9 expression.

BACKGROUND: Calcific aortic valve disease (CAVD) is the second leading cause of adult heart diseases. The purpose of this study is to investigate whether miR-101-3p plays a role in the human aortic valve interstitial cells (HAVICs) calcification and the underlying mechanisms. METHODS: Small RNA deep sequencing and qPCR analysis were used to determine changes in microRNA expression in calcified human aortic valves. RESULTS: The data showed that miR-101-3p levels were increased in the calcified human aortic valves. Using cultured primary HAVICs, we demonstrated that the miR-101-3p mimic promoted calcification and upregulated the osteogenesis pathway, while anti-miR-101-3p inhibited osteogenic differentiation and prevented calcification in HAVICs treated with the osteogenic conditioned medium. Mechanistically, miR-101-3p directly targeted cadherin-11 (CDH11) and Sry-related high-mobility-group box 9 (SOX9), key factors in the regulation of chondrogenesis and osteogenesis. Both CDH11 and SOX9 expressions were downregulated in the calcified human HAVICs. Inhibition of miR-101-3p restored expression of CDH11, SOX9 and ASPN and prevented osteogenesis in HAVICs under the calcific condition. CONCLUSION: miR-101-3p plays an important role in HAVIC calcification through regulation of CDH11/SOX9 expression. The finding is important as it reveals that miR-1013p may be a potential therapeutic target for calcific aortic valve disease.

Therapeutic potential of anti-miR29a in breast cancer patients with type 2 diabetes: an in vitro and xenograft mouse-model study.

Background: MicroRNAs (miRNAs) acting as tumour suppressors or oncogenes, known as oncomiRs, are a promising new focus in targeted therapies for cancer. Approximately 16% of breast cancer patients have pre-existing diabetes. Breast cancer with type 2 diabetes mellitus (BDM) is provided with its unique biological characteristics and clinical characteristics. This study primarily investigated the therapeutic potential and regulatory mechanism of miR-29a in patients with BDM. Methods: The significance of miR-29a in BDM was analyzed by real-time reverse transcriptase polymerase chain reaction (qRT-PCR) in breast tissues. A cell model for BDM was established by using MDA-MB-231 cells cultured in 3T3-L1 adipocytes cultured with high levels of glucose and insulin. A type 2 diabetes mellitus (T2DM) mouse model was induced in female BALB/c mice through a high-fat diet plus low doses of streptozotocin (STZ). The xenograft mouse-model for BDM was established on these T2DM mouse by using MDA-MB-231 cells. Then the biological effects of miR-29a knockdown mediated by lentivirus-shRNAs on cell proliferation, apoptosis, cell cycle, and migration were investigated. Results: Our results indicated that miR-29a was upregulated in patients with BDM, which correlated with a worse prognosis. In human breast cancer cells, miR-29a knockdown reduced cell proliferation and cell migration and invasion in BDM. In the T2DM xenograft, miR-29a knockdown suppressed MDA-MB-231 cells tumorigenesis and metastasis. We also demonstrated that miR-29a promoted BDM cell growth and metastasis by targeting Sirtuin 1 (SIRT1). Conclusions: Our findings indicated that anti-miR-29a inhibited cell proliferation and invasion in BDM by targeting SIRT1. We believe anti-miR-29a may represent a novel therapeutic approach for the management of patients with BDM.

MicroRNA inhibition using antimiRs in acute human brain tissue sections.

Antisense inhibition of microRNAs is an emerging preclinical approach to pharmacoresistant epilepsy. A leading candidate is an "antimiR" targeting microRNA-134 (ant-134), but testing to date has used rodent models. Here, we develop an antimiR testing platform in human brain tissue sections. Brain specimens were obtained from patients undergoing resective surgery to treat pharmacoresistant epilepsy. Neocortical specimens were submerged in modified artificial cerebrospinal fluid (ACSF) and dissected for clinical neuropathological examination, and unused material was transferred for sectioning. Individual sections were incubated in oxygenated ACSF, containing either ant-134 or a nontargeting control antimiR, for 24 h at room temperature. RNA integrity was assessed using BioAnalyzer processing, and individual miRNA levels were measured using quantitative reverse transcriptase polymerase chain reaction. Specimens transported in ACSF could be used for neuropathological diagnosis and had good RNA integrity. Ant-134 mediated a dose-dependent knockdown of miR-134, with approximately 75% reduction of miR-134 at 1 µmol L-1 and 90% reduction at 3 µmol L-1 . These doses did not have off-target effects on expression of a selection of three other miRNAs. This is the first demonstration of ant-134 effects in live human brain tissues. The findings lend further support to the preclinical development of a therapy that targets miR-134 and offer a flexible platform for the preclinical testing of antimiRs, and other antisense oligonucleotide therapeutics, in human brain.

Synergistic Effects of A Combined Treatment of Glioblastoma U251 Cells with An Anti-miR-10b-5p Molecule and An AntiCancer Agent Based on 1-(3',4',5'-Trimethoxyphenyl)-2-Aryl-1H-Imidazole Scaffold.

(1) Background: In the development of new and more effective anticancer approaches, combined treatments appear of great interest. Combination therapy could be of importance in the management of glioblastoma (GBM), a lethal malignancy that accounts for 42% of cancer of the central nervous system, with a median survival of 15 months. This study aimed to verify the activity on a glioblastoma cancer cell line of one of the most active compounds of a novel series of tubulin polymerization inhibitors based on the 1-(3',4',5'-trimethoxyphenyl)-2-aryl-1H-imidazole scaffold, used in combination with a miRNA inhibitor molecule targeting the oncomiRNA miR-10b-5p. This microRNA was selected in consideration of the role of miR-10b-5p on the onset and progression of glioblastoma. (2) Methods: Apoptosis was analyzed by Annexin-V and Caspase 3/7 assays, efficacy of the anti-miR-10b-5p was assessed by determining the miR-10b-5p content by RT-qPCR. (3) Results: The results obtained show that a "combination therapy" performed by combining the use of an anti-miR-10b-5p and a 1-(3',4',5'-trimethoxyphenyl)-2-aryl-1H-imidazole derivative is an encouraging strategy to boost the efficacy of anticancer therapies and at the same time to reduce side effects.