LncRNAH19 acts as a ceRNA of let-7 g to facilitate endothelial-to-mesenchymal transition in hypoxic pulmonary hypertension via regulating TGF-ß signalling pathway.

BACKGROUND: Hypoxic pulmonary hypertension (HPH) is a challenging lung arterial disorder with remarkably high incidence and mortality rates, and the efficiency of current HPH treatment strategies is unsatisfactory. Endothelial-to-mesenchymal transition (EndMT) in the pulmonary artery plays a crucial role in HPH. Previous studies have shown that lncRNA-H19 (H19) is involved in many cardiovascular diseases by regulating cell proliferation and differentiation but the role of H19 in EndMT in HPH has not been defined. METHODS: In this research, the expression of H19 was investigated in PAH human patients and rat models. Then, we established a hypoxia-induced HPH rat model to evaluate H19 function in HPH by Echocardiography and hemodynamic measurements. Moreover, luciferase reporter gene detection, and western blotting were used to explore the mechanism of H19. RESULTS: Here, we first found that the expression of H19 was significantly increased in the endodermis of pulmonary arteries and that H19 deficiency obviously ameliorated pulmonary vascular remodelling and right heart failure in HPH rats, and these effects were associated with inhibition of EndMT. Moreover, an analysis of luciferase activity indicated that microRNA-let-7 g (let-7 g) was a direct target of H19. H19 deficiency or let-7 g overexpression can markedly downregulate the expression of TGFßR1, a novel target gene of let-7 g. Furthermore, inhibition of TGFßR1 induced similar effects to H19 deficiency. CONCLUSIONS: In summary, our findings demonstrate that the H19/let-7 g/TGFßR1 axis is crucial in the pathogenesis of HPH by stimulating EndMT. Our study may provide new ideas for further research on HPH therapy in the near future.

The microRNA Let-7f Induces Senescence and Exacerbates Oxidative Stress in Retinal Pigment Epithelial Cells.

This study aims to investigate the role of microRNA let-7f in the dysfunction and degeneration of retinal pigment epithelium (RPE) cells through the induction of senescence and oxidative stress. Furthermore, we explore whether let-7f inhibition can protect these cells against sodium iodate (SI)-induced oxidative stress. Oxidative stress and let-7f expression are reciprocally regulated in retinal pigment epithelial cells. Overexpression of let-7f in ARPE-19 cells induced oxidative stress as demonstrated by increased reactive oxygen species (ROS) production as well as senescence. Inhibition of let-7f successfully protected RPE cells from the detrimental effects induced by SI. In addition, let-7f overexpression induced RPE cellular dysfunction by diminishing their migratory capabilities and reducing the phagocytosis of porcine photoreceptor outer segments (POS). Results were further confirmed in vivo by intravitreal injections of SI and let-7f antagomir in C57BL/6 mice. Our results provide strong evidence that let-7f is implicated in the dysfunction of RPE cells through the induction of senescence and oxidative injury. These findings may help to uncover novel and relevant processes in the pathogenesis of dry AMD.

Let-7b-TLR7 Signaling Axis Contributes to the Anesthesia/Surgery-Induced Cognitive Impairment.

Perioperative neurocognitive disorders (PNDs) are severe and common neurological complications among elderly patients following anesthesia and surgery. As the first line of defense of the innate immune system, Toll-like receptors (TLRs) have been found to be involved in the occurrence of neurodegenerative diseases in recent years. However, the role of TLR7 in the pathology and development of PNDs remains largely unclear. In our current study, we hypothesized that increased microRNA let-7b (let-7b) during anesthesia and surgical operation would activate TLR7 signaling pathways and mediate PNDs. Using a mouse model of PNDs, 18-20 months wild-type (WT) mice were undergoing unilateral nephrectomy, and increased TLR7 and let-7b expression levels were found in the surgery group compared with the Sham group. Of note, increased TLR7 was found to be co-localized with let-7b in the hippocampal area CA1 in the PNDs model. In addition, TLR7 and let-7b inhibition could improve hippocampus-dependent memory and attenuate the production of inflammatory cytokines. Together, our results indicated that TLR7 activation and up-regulation might be triggered by increased let-7b under stressful conditions and initiated the downstream inflammatory signaling, playing a substantial role in the development of PNDs.

Role of miR­let­7c­5p/c­myc signaling axis in the committed differentiation of leukemic THP­1 cells into monocytes/macrophages.

In a preliminary experiment, it was found that c-myc expression was decreased following the differentiation of THP-1 cells into monocytes/macrophages induced by phorbol 12-myristate 13 acetate (PMA) + lipopolysaccharide (LPS) + interferon (IFN)-γ. The expression of miR-let-7c-5p was then found to be elevated by cross-sectional analysis using TargetScan and PubMed and differential microarray analysis. The present study aimed to investigate the role of the miR-let-7c-5p/c-myc signaling axis in the committed differentiation of THP-1 leukemic cells into monocytes/macrophages induced by PMA + LPS + IFN-γ. Human THP-1 leukemic cells were induced to differentiate into monocytes/macrophages by PMA + LPS + IFN-γ. Following induction for 48 h, the growth density of the THP-1 cells was observed directly under an inverted microscope, cell proliferation was measured using Cell Counting Kit-8 assay and the cell cycle and the expression of differentiation-related antigens (CD11b and CD14) were measured using flow cytometry. The mRNA expression of miR-let-7c-5p and c-myc was detected using reverse transcription-quantitative PCR and the protein expression of c-myc was detected using western blot analysis. Dual luciferase reporter gene analysis was used to detect the targeted binding of miR-let-7c-5p on the 3'UTR of c-myc. The relative expression of miR-let-7c-5p and c-myc genes in THP-1 cells induced by PMA + LPS + IFN-γ was found to be up- and downregulated respectively, and expression of miR-let-7c-5p was negatively correlated with the expression of c-myc gene. Dual luciferase reporter gene assays confirmed that miR-let-7c-5p targeted the 3'UTR of c-myc and inhibited luciferase activity. Following transfection with miR-let-7c-5p mimics, the expression of c-myc was markedly downregulated and the proliferative ability of the THP-1 cells was decreased, while the expression rate of CD11b and CD14 was significantly increased. The rescue experiment revealed that the effects of miR-let-7c-5p mimics on the proliferation and differentiation of THP-1 cells were attenuated by transfection with c-myc overexpression vector. Together, the findings of the present study demonstrated that miR-let-7c-5p can target the 3'UTR region of c-myc and that the miR-let-7c-5p/c-myc signaling axis is one of the critical pathways involved in the directional differentiation of leukemic cells into monocytes/macrophages.

Genetic variants in the promoters of let-7 are associated with the risk and age at onset of ischemic stroke: A case control study.

PURPOSE: Let-7 family members serve as crucial regulatory molecules in the pathogenesis of ischemic stroke. We predicted that genetic variations in the let-7 family's promoters may be linked to the risk of ischemic stroke. The connection of rs10877887 and rs13293512 in the let-7 family promoters with liability to ischemic stroke was explored in this study. PATIENTS AND METHODS: Clinical data and peripheral blood samples were collected from 914 ischemic stroke patients and 836 controls in this case-control study. All statistical analyses were carried out using SPSS. RESULTS: Our analysis results reveal that the rs10877887 TC+CC genotype in the dominant model is associated with a lower risk of ischemic stroke than the TT genotype. Individuals with heterozygous TC or homozygous CC genotypes in the male population showed higher odds of ischemic stroke than those with the wild TT genotype in rs13293512 analysis. Furthermore, there existed a multiplicative interaction between the rs10877887 C allele and the rs13293512 T allele. In the presence of the rs13293512 T allele, the effect of the rs10877887 C allele on ischemic stroke risk was increased. Similarly, in the presence of the rs10877887 C allele, the outcome of the rs13293512 T allele on ischemic stroke risk was elevated. In addition, the rs13293512 CC genotype seemed to lead to an earlier onset of ischemic stroke. CONCLUSION: Our findings indicated that these two SNPs might have a joint role in IS and could potentially act as risk markers. Detecting let-7 promoter polymorphisms could raise awareness of the risk of IS, which directed individuals with risk alleles to have regular checks at an appropriate frequency to avoid developing the disease.

A Panel of Circulating Non-Coding RNAs in the Diagnosis and Monitoring of Therapy in Egyptian Patients with Breast Cancer.

Background: Non-coding RNAs (ncRNAs) have recently been identified to have a pivotal role in many diseases, including breast cancer (BC). This study aims to investigate the relative quantification of long non-coding RNA (lncRNA) H19, microRNA (miR) 675-5p, 675-3p, and miR-let 7 in breast cancer patients. Methods: The study was performed on three groups: Group 1: 30 non-intervened BC female patients about to undergo breast surgery; group 2: 30 postoperative female BC patients about to receive adjuvant anthracycline chemotherapy; and group 3: 30 apparently healthy female volunteers as the control group. Plasma samples were drawn before and after the intervention in groups 1 and 2, with a single sample drawn from group 3. The relative quantification levels were compared with healthy control subjects and were related with the clinicopathological statuses of these patients. Results: There was a statistically significant increase in H19, miR-675-5p, miR-675-3p, and miR-let 7 in the non-intervened BC patients when compared to the control group. Surgery resulted in a significant reduction in all four ncRNAs under investigation. Chemotherapy brought about a significant increase in the level of miR-let 7, with no significant effect on the remaining parameters measured. The assay discriminated normal from BC where a receiver operating characteristic for the area under the curve (ROCAUC) of miR-675-3p showed the maximal AUC of 1.000. The diagnostic sensitivity and specificity were also 100% when CA 15-3 and H19 were combined. Conclusion: The results strongly indicate that the panel of ncRNAs in this study can all potentially act as novel biomarkers whether alone or combined in the diagnosis of BC.

Green Tea Epigallocatechin Gallate Inhibits Preadipocyte Growth via the microRNA-let-7a/HMGA2 Signaling Pathway.

SCOPE: This study investigates the effect of epigallocatechin gallate (EGCG) on white and beige preadipocyte growth and explores the involvement of the miR-let-7a/HMGA2 pathway. METHODS AND RESULTS: 3T3-L1 and D12 cells are treated with EGCG. The effect of EGCG on cell proliferation and viability is evaluated, as well as microRNA (miRNA)-related signaling pathways. EGCG inhibits 3T3-L1 and D12 preadipocyte growth, upregulates miR-let-7a expression, and downregulates high-mobility group AT-hook 2 (HMGA2) mRNA and protein levels in a time- and dose-dependent manner. In addition, overexpression of miR-let-7a significantly inhibits the growth of 3T3-L1 and D12 cells and decreases HMGA2 mRNA and protein levels. MiR-let-7a inhibitor antagonizes the inhibitory effects of EGCG on the number and viability of 3T3-L1 and D12 cells. Furthermore, miR-let-7a inhibitor reverses the EGCG-induced increase in miR-let-7a expression levels and decrease in HMGA2 mRNA and protein levels. HMGA2 overexpression induces an increase in cell number and viability and antagonizes EGCG-suppressed cell growth and HMGA2 expression in 3T3-L1 and D12 preadipocytes. CONCLUSION: EGCG inhibits the growth of 3T3-L1 and D12 preadipocytes by modulating the miR-let-7a and HMGA2 pathways.

Targeting Castration-Resistant Prostate Cancer Using Mesenchymal Stem Cell Exosomes for Therapeutic MicroRNA-let-7c Delivery.

BACKGROUND: Castration-resistant prostate cancer (PCa; CRPC) has a poor response to androgen deprivation therapy and is considered an incurable disease. MicroRNA (miR)-lethal 7c (let-7c) was implied to be a tumor suppressor in PCa, and treatment with exogenous let-7c targets both cancer cells and their associated mesenchymal stem cells (MSCs) to prevent CRPC progression and metastasis. Exosomes are nanometer-sized membrane-bound vesicles which have an absolute predominance in biocompatibility for drug delivery and gene therapy by mediating cell-to-cell communication. By utilizing the intrinsic tumor-targeting property of MSCs, this study aimed to investigate the feasibility of MSC-derived exosomes as an exogenous miR delivery system to target CRPC, using miR let-7c as an example. METHODS: Bioinformatics analysis was performed to observe miR-let-7c expression in clinical samples by utilizing the GEO database. MSC-derived exosomes were collected from a human bone marrow-derived MSC cell line after cell transfection with either a pre-miR negative control or pre-miR-let-7c, and further characterized through nanoparticle tracking analysis and Western blotting. miR-let-7c expression was determined using RT-qPCR, and the phenotypic effects of both naked and MSC-exosome-encapsulated let-7c on CRPC cells (PC3 and CWR22Rv1) were determined by WST-1 cell proliferation assay and wound healing migration assay. RESULTS: miR-let-7c was downregulated in metastatic PCa and high grade group patients. miR-let-7c expression was confirmed to be downregulated in PCa cell lines, with massively decreased in most metastatic CRPC-like cells. Exogenous miR-let-7c can be successfully packaged into MSC exosomes. Treatment with either naked or MSC-exosome-encapsulated miR-let-7c resulted in significant reductions in cell proliferation and migration in CRPC-like PC3 and CWR22Rv1 cells. CONCLUSIONS: MSC-derived exosomes could serve as a therapeutic let-7c delivery system to target CRPC.

MicroRNA let-7 Suppresses Influenza A Virus Infection by Targeting RPS16 and Enhancing Type I Interferon Response.

Given the frequent emergence of drug-resistant influenza virus strains and new highly pathogenic influenza virus strains, there is an urgent need to identify new antiviral drugs and targets. We found that influenza A virus (IAV) infection caused a significant decrease of microRNA let-7 expression in host cells; that overexpression of let-7 increased interferon expression and effectively inhibit IAV infection; and that let-7 targets the 3'-untranslated region (UTR) of the ribosomal protein 16 (RPS16) gene, decreasing its expression. Knocking down the expression of RPS16 increased the expression of type I interferon and inhibited viral replication. The present study uncovered the regulatory effect of let-7b and let-7f on influenza A infection, which is a potential biomarker of IAV infection. In addition, let-7 may be a promising therapeutic agent against influenza A.

miR-let-7c-3p targeting on Egr-1 contributes to the committed differentiation of leukemia cells into monocyte/macrophages.

In preliminary experiments, it was found that the expression of early growth response-1 (Egr-1) was upregulated during the committed differentiation of leukemia cells into monocytes/macrophages. The cross-analysis of gene chip detection and database prediction indicated that Egr-1 was associated with upstream microRNA (miR)-let-7c-3p, thus the present study focused on the role of the miR-let-7c-3p/Egr-1 signaling axis in the committed differentiation of leukemia cells into monocytes/macrophages. Phorbol 12-myristate 13-acetate (PMA) was used to induce the directed differentiation of human K562 leukemia cells into monocytes/macrophages and the differentiation of K562 leukemia cells was determined by cell morphology observation and expression of differentiation antigens CD11b and CD14 by flow cytometry. The expression levels of Egr-1 and miR-let-7c-3p were detected by reverse transcription-quantitative PCR and the protein expression of Egr-1 was detected by western blotting. The effect of Egr-1 on the differentiation of K562 cells was detected by short interfering (si)RNA interference assay. A dual-luciferase reporter assay was used to detect target binding of miR-let-7c-3p on the 3'UTR of Egr-1. Cell transfection of miR-let-7c-3p mimics and inhibitors was used to modulate the expression of miR-let-7c-3p, as indicated by RT-qPCR assays. Western blotting was also used to examine the effect of miR-let-7c-3p on Egr-1 expression. The PMA-induced differentiation of K562 cells was transfected with miR-let-7c-3p and the expression of differentiation antigen was detected by flow cytometry. A differentiation model of K562 leukemia cells into monocytes/macrophages was induced by PMA, which was indicated by morphological observations and upregulation of CD11b and CD14 antigens. The gene or protein expression of Egr-1 was significantly higher compared with that of the control group, while the expression of miR-let-7c-3p was significantly lower compared with that of the control group. siRNA interference experiments showed that the expression of cell differentiation antigen CD14 in the 100 µg/ml PMA + si-Egr-1 group was significantly lower compared with that in the 100 µg/ml PMA + si-ctrl group. The dual luciferase reporter gene results showed that the luciferase activity of the co-transfected mimic and Egr-1 WT groups was significantly lower than that of the NC control group, while the luciferase activity of the co-transfected mimic and Egr-1 MUT groups was comparable to that of the NC control group. Therefore, the dual-luciferase reporter gene assay confirmed that miR-let-7c-3p can target Egr-1. Western blotting showed that the expression of Egr-1 following transfection with miR-let-7c-3p inhibitor was significantly higher compared with that of the negative control and the expression of Egr-1 after transfection with miR-let-7c-3p mimic was significantly lower than that of the negative control. Following exposure to PMA, the expressions of CD11b and CD14 in the miR-let-7c-3p inhibitor group were significantly higher than those in the miR-let-7c-3p NC group, as indicated by CD11b and CD14 respectively. In conclusion, miR-let-7c-3p could bind to the 3'UTR of Egr-1 and negatively regulated Egr-1 expression. The miR-let-7c-3p/Egr-1 signaling axis was closely associated with the committed differentiation of K562 cells from leukemia cells to monocytes/macrophages.

KDM2B mediates the Wnt/ß-catenin pathway through transcriptional activation of PKMYT1 via microRNA-let-7b-5p/EZH2 to affect the development of non-small cell lung cancer.

The significance of KDM2B in oncogenesis has been appreciated, but the mechanism behind is incompletely understood. In this work, we addressed its effects on the progression of non-small cell lung cancer (NSCLC). Overexpression of KDM2B was linked to dismal prognoses of NSCLC patients. Based on the expression levels of KDM2B in a panel of NSCLC cell lines, A549, showing lower level of expression, and SK-MES-1, showing higher levels of expression, were selected as model systems to evaluate the effect of KDM2B overexpression and KDM2B silencing, respectively. Knockdown of KDM2B hampered NSCLC cell proliferation, invasion, as well as migration, while enhanced apoptosis. Additionally, KDM2B repressed the expression of microRNA (miR)-let-7b-5p through demethylation modification of H3K36me2, thereby promoting the expression of zester homolog 2 (EZH2), the target gene of let-7b-5p in NSCLC. Moreover, EZH2 transcriptionally induced the expression of PKMYT1 to activate the Wnt/ß-catenin pathway. Sh-EZH2 and sh-PKMYT1 neutralized the supporting effects of KDM2B on cell proliferation, invasion and migration. Additionally, deletion of KDM2B reduced the xenograft volumes in nude mice. In conclusion, KDM2B induces the EZH2/PKMYT1/Wnt/ß-catenin axis by inhibiting the let-7b-5p expression, which promotes NSCLC growth. More investigations are essential to determine the oncogenic role of KDM2B in NSCLC.

Radix ranunculus temate saponins sensitizes ovarian cancer to Taxol via upregulation of miR-let-7b.

A common cause of treatment failure in ovarian cancer is acquired drug resistance. Therefore, effective novel drugs against chemoresistance need to be developed. MicroRNAs (miRNAs or miRs) serve key regulatory roles in tumorigenesis and chemoresistance. The objective of the present study was to explore the role of miR-let-7b in ovarian cancer chemoresistance, and to develop novel strategy for the treatment of drug-resistant ovarian cancer. For this purpose, reverse transcription-quantitative PCR was performed to evaluate the expression level of miR-let-7b in fresh ovarian cancer tissues and cell lines. miR-let-7b mimic was transfected into ovarian cancer cell lines. Functional experiments, cell apoptosis and cell viability assays were carried out to identify the tumor-suppressor function of miR-let-7b. The treatment effect of Radix ranunculus temate saponins (RRTS), one of the primary constituents extracted from the traditional Chinese medicine radix Ranunculi ternati, was identified in vitro and in vivo. The results revealed that miR-let-7b was downregulated significantly in chemoresistant ovarian cancer patients. miR-let-7b overexpression suppressed cell growth and invasion and enhanced sensitivity to Taxol of ovarian cancer cells. Furthermore, miR-let-7b levels in ovarian cancer tissue were inversely associated with collagen type III α1 chain (COL3A1) levels. COL3A1, a non-fibrillar collagen associated with chemoresistance, was targeted by miR-let-7b. RRTS showed cytotoxic effects on ovarian cancer cells through inducing miR-let-7b expression and decreasing COL3A1 expression. In addition, RRTS sensitized ovarian cancer to Taxol both in vitro and in vivo. In conclusion, the present results revealed synergistic cytotoxicity of RRTS and Taxol on against ovarian cancer cells via upregulating expression of miR-let-7b. Combination of Taxol and RRTS may be a novel treatment strategy for patients with TR ovarian cancer.

Combination microRNA-based cellular reprogramming with paclitaxel enhances therapeutic efficacy in a relapsed and multidrug-resistant model of epithelial ovarian cancer.

Most advanced-stage ovarian cancer patients, including those with epithelial ovarian cancer (EOC), develop recurrent disease and acquisition of resistance to chemotherapy, leading to limited treatment options. Decrease in Let7b miRNA levels in clinical ovarian cancer has been associated with chemoresistance, increased proliferation, invasion, and relapse in EOC. We have established a murine EOC relapsed model by administering paclitaxel (PTX) and stopping therapy to allow for tumor regrowth. Global microRNA profiling in the relapsed tumor showed significant downregulation of Let7b relative to untreated tumors. Here, we report the use of hyaluronic acid (HA)-based nanoparticle formulation that can deliver Let7b miRNA mimic to tumor cells and achieve cellular programming both in vitro and in vivo. We demonstrate that a therapeutic combination of Let7b miRNA and PTX leads to significant improvement in anti-tumor efficacy in the relapsed model of EOC. We further demonstrate that the combination therapy is safe for repeated administration. This novel approach of cellular reprogramming of tumor cells using clinically relevant miRNA mimic in combination with chemotherapy could enable more effective therapeutic outcomes for patients with advanced-stage relapsed EOC.

Iron Oxide Nanoparticles Conjugated to Thiosemicarbazone Reduce the Survival of Cancer Cells by Increasing the Gene Expression of MicroRNA let-7c in Lung Cancer A549 Cells.

BACKGROUND: Cancer cells have a higher demand for iron to grow and proliferate. A new complex of iron nanoparticles and thiosemicarbazones was synthesized. Confirmation tests included UV-visible, scanning electron microscopy (SEM), energy dispersive X-ray analysis (EDX), Fourier transform infrared (FTIR), X-ray diffraction (XRD) and zeta potential. METHODS: MTT assay, flow cytometry and qRT-PCR were used to investigate anti-proliferative effect, amount of apoptosis and the effect of Fe3 O4 @Glu/BTSC on changes in gene expression of microRNA let-7c (let-7c), respectively. The specifications of Fe3 O4 @ Glu/BTSC were confirmed at 5 nm. RESULTS: Fe3O4@Glu/BTSC was more effective than BTSC and Fe3 O4 on A549 cells (IC50=166.77 µg/mL) but its effect on healthy cells was smaller (CC50=189.15 µg/mL). The drug selectivity index (SI) was calculated to be 1.13. The initial apoptosis rate was 46.33% for Fe3 O4 @Glu/BTSC, 28.27% for BTSC and 26.02% for Fe3 O4 . BTSC and BTSC@Fe3 O4 inhibited the cell cycle progression in the Sub-G1 and S phases. let-7c expression was 6.9 times higher in treated cells compared to the control group. The expression rate was 2.2 with BTSC compared to the control group and 1.6 times for Fe3 O4. CONCLUSION: Fe3 O4 @Glu/BTSC has proper anti-proliferative effects against lung cancer cells by increasing the expression of let-7c and inhibiting the cell cycle with the apoptosis activation pathway.

Catalpol Inhibits Tregs-to-Th17 Cell Transdifferentiation by Up-Regulating Let-7g-5p to Reduce STAT3 Protein Levels.

PURPOSE: Rheumatoid arthritis (RA) is a chronic systemic autoimmune disease, and Th17 cells are key factors in the pathogenesis of human inflammatory conditions, such as RA. Catalpol (CAT), a component in Rehmanniae Radix (RR), has been found to regulate human immunity. However, the effects of CAT on Th17 cell differentiation and improvement of RA are not clear. MATERIALS AND METHODS: Collagen-induced arthritis (CIA) mice were constructed to detect the effects of CAT on arthritis and Th17 cells. The effect of CAT on Th17 differentiation was evaluated with let-7g-5p transfection experiments. Flow cytometry was used to detect the proportion of Th17 cells after CAT treatment. Levels of interleukin-17 and RORγt were assessed by qRT-PCR and enzyme-linked immunosorbent assay. The expression of signal transducer and activator of transcription 3 (STAT3) was determined by qRT-PCR and Western blot. RESULTS: We found that the proportion of Th17 cells was negatively associated with let-7g-5p expression in CIA mice. In in vitro experiments, CAT suppressed traditional differentiation of Th17 cells. Simultaneously, CAT significantly decreased Tregs-to-Th17 cells transdifferentiation. Our results demonstrated that CAT inhibited Tregs-to-Th17 cells transdifferentiation by up-regulating let-7g-5p and that the suppressive effect of CAT on traditional differentiation of Th17 cells is not related with let-7-5p. CONCLUSION: Our data indicate that CAT may be a potential modulator of Tregs-to-Th17 cells transdifferentiation by up-regulating let-7g-5p to reduce the expression of STAT3. These results provide new directions for research into RA treatment.

MicroRNA let-7b inhibits hepatitis C virus and induces apoptosis in human hepatoma cells.

BACKGROUND: Small non-coding RNAs have emerged as essential modulators of viral infections such as hepatitis C virus (HCV). Cellular miRNAs directly regulate the viral infectivity and indirectly by targeting virus-host factors. The current study investigates the inhibitory effect of let-7b miRNA on HCV replication in the Hepatocarcinoma cell line (Huh7.5). METHODS AND RESULTS: The algorithm-based search revealed that let-7b, a high score microRNA, has target sequences on the HCV genome. The Huh7.5 cells were stably transduced with let-7b lentiviral vectors (Huh7.5/let-7b) and mock (Huh7.5/scrambled). The expression of the let-7b level was assessed by real-time PCR assay and Red fluorescence microscope. A dual-luciferase assay was conducted to evaluate the liver-specific let-7b and HCV genome interaction. In the next step, for establishing HCVcc, Full-length HCV-RNA was transduced to naïve Huh7.5, Huh7.5/scrambled, and Huh7.5/let-7b cells. The results of in silico analysis and dual-luciferase reporter assay exhibited a specific interaction of HCV-NS5B and let-7b. Real-time PCR analysis revealed that in contrast to infected naïve Huh7.5 cells and Huh7.5/scrambled, a significant decrease in HCV-RNA load was seen in Huh7.5/let-7b cells. On the other hand, the Flow Cytometry test showed that let-7b could significantly induce the apoptosis pathway in Huh7.5/let-7b. CONCLUSIONS: The results also suggest that let-7b, as a target of the HCV genome, potentially reduces HCV replication and raises cell apoptosis rate. We suggest that let-7b directly downregulates HCV replication and may serve as a unique antiviral therapy.

MicroRNA let-7: A promising non-invasive biomarker for diagnosing and treating external genital endometriosis

Objective: To evaluate the possibility of using microrna let-7 and mir-9 as non-invasive biomarkers for the diagnosis and treatment of external genital endometriosis. Materials and Methods: We explored the samples of relatively healthy individuals and patients with endometriosis. All patients had undergone laparoscopic surgical treatment after clinical and laboratory examinations. We used RNA-GO to obtain total RNA from endometriosis samples excised by laparoscopic method. Next step involved reverse transcription for microRNA let-7 and mir-9. Correlation-regression analysis was performed using Mann-Whitney-Wilcoxon method. Subsequently, receiver operating characteristic analysis was conducted to determine the possibility of using let-7 microRNA for noninvasive detection of endometriosis. The results of the analysis in all groups were tested considering the normality of statistical distribution. Results: Mann-Whitney analysis showed that the difference in mir-9 mRNA between the groups with and without endometriosis, as well as between the groups with more clinically and histologically severe and mild endometriosis, was statistically insignificant. In addition, a significant difference was noted regarding let-7 microRNA between the groups with and without endometriosis, as well as between the groups with more clinically and histologically severe and mild endometriosis. Comparison with cancer antigen-125 (CA-125) showed that let-7 microRNA was a more specific test than CA-125. Conclusion: MicroRNA let-7 had the best parameters (sensitivity, specificity, and predictive value of positive and negative results) among the biomarkers studied. These biomarkers may be used for early and sometimes preclinical diagnosis of endometriosis.

Long non-coding RNA NEAT1 facilitates the growth, migration, and invasion of ovarian cancer cells via the let-7 g/MEST/ATGL axis.

BACKGROUND/AIM: Growing evidence indicates a significant role of long non-coding RNA (lncRNA) nuclear-enriched abundant transcript 1 (NEAT1) in ovarian cancer, a frequently occurring malignant tumor in women; however, the possible effects of an interplay of NEAT1 with microRNA (miRNA or miR) let-7 g in ovarian cancer are not known. The current study aimed to investigate the role of the NEAT1/let-7 g axis in the growth, migration, and invasion of ovarian cancer cells and explore underlying mechanisms. METHODS: NEAT1 expression levels were examined in clinical tissue samples and cell lines. The relationships between NEAT1, let-7 g, and MEST were then analyzed. Gain- or loss-of-function approaches were used to manipulate NEAT1 and let-7 g. The effects of NEAT1 on cell proliferation, migration, invasion, and apoptosis were evaluated. Mouse xenograft models of ovarian cancer cells were established to verify the function of NEAT1 in vivo. RESULTS: NEAT1 expression was elevated while let-7 g was decreased in ovarian cancer clinical tissue samples and cell lines. A negative correlation existed between NEAT1 and let-7 g, whereby NEAT1 competitively bound to let-7 g and consequently down-regulate let-7 g expression. By this mechanism, the growth, migration, and invasion of ovarian cancer cells were stimulated. In addition, let-7 g targeted mesoderm specific transcript (MEST) and inhibited its expression, leading to promotion of adipose triglyceride lipase (ATGL) expression and inhibition of ovarian cancer cell growth, migration, and invasion. However, the effect of let-7 g was abolished by overexpression of MEST. Furthermore, silencing of NEAT1 decreased the xenograft tumor growth by decreasing MEST while up-regulating let-7 g and ATGL. CONCLUSIONS: Cumulatively, the findings demonstrated that NEAT1 could promote malignant phenotypes of ovarian cancer cells by regulating the let-7 g/MEST/ATGL signaling axis. Therefore, NEAT1 can be regarded as an important molecular target and biomarker for ovarian cancer.

Long non-coding RNA NEAT1 absorbs let-7 g-5p to induce epithelial-mesenchymal transition of colon cancer cells through upregulating BACH1.

OBJECTIVE: Long noncoding RNAs (lncRNAs) are critical regulators in diverse human cancers. However, the role of lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1) in colon cancer remains to be further investigated. We aimed to verify the role of NEAT1/let-7 g-5p/BTB and CNC homology 1 (BACH1) axis in colon cancer development. METHODS: Expression of NEAT1, let-7 g-5p and BACH1 in colon cancer tissues and cells was determined. The interactions between NEAT1 and let-7 g-5p, and between let-7 g-5p and BACH1 were assessed. The colon cancer cell lines were treated with plasmids or oligonucleotides to alter NEAT1, BACH1 and let-7 g-5p expression. Then, viability, migration, invasion, and apoptosis of colon cells were evaluated, and the cell growth in vivo was observed as well. RESULTS: NEAT1 and BACH1 were upregulated while let-7 g-5p was downregulated in colon cancer tissues and cells. NEAT1/BACH1 silencing or let-7 g-5p elevation suppressed colon cancer cell growth in vivo and in vitro. The effects of silenced NEAT1 on colon cancer cells and xenografts were reversed by downregulating let-7 g-5p. Down-regulation of BACH1 reversed the effect of NEAT1 overexpression on colon cancer cells. NEAT1 directly bound to let-7 g-5p and let-7 g-5p targeted BACH1. CONCLUSION: Downregulated NEAT1 elevated let-7 g-5p to suppress EMT of colon cancer cells through inhibiting BACH1. This research may contribute to treatment of colon cancer.

MicroRNA let-7i-3p affects osteoblast differentiation in ankylosing spondylitis via targeting PDK1.

Ankylosing spondylitis (AS) is a chronic autoimmune disease in which let-7i has been studied to involved. But, whether let-7i-3p could regulate osteoblast differentiation in AS remains unclear. This research targeted to decipher the impact of let-7i-3p on AS progression by modulating pyruvate dehydrogenase kinase 1 (PDK1). The bone mineral density of femur and lumbar vertebra and the maximum loading and bending elastic modulus of tibia, tumor necrosis factor-α (TNF-α), matrix metalloproteinase (MMP)-3, osteoprotegerin (OPG) and receptor activator of nuclear factor-κB ligand (RANKL) in serum of AS mice, the pathological condition of synovial tissue were determined via let-7i-3p inhibitor and OE-PDK1 in animal experiment. Also, the cell viability and ALP activity were measured by let-7i-3p inhibitor and OE-PDK1 in cell experiments. let-7i-3p and PDK1 expression were detected. Let-7i-3p raised and PDK1 declined in AS mice. Depleted let-7i-3p and restored PDK1 increased bone mineral density and maximum loading and bending elastic modulus of tibia, reduced TNF-α, MMP-3 and RANKL contents, attenuated the pathological condition of synovial tissue and raised OPG content in AS mice. In cell experiments, up-regulating PDK1 and down-regulating let-7i-3p enhanced cell viability and ALP activity in AS mice. Low expression of let-7i-3p could enhance osteoblast differentiation in AS by up-regulating PDK1.Abbreviations: AS: Ankylosing spondylitis; PDK1: pyruvate dehydrogenase kinase 1; TNF-α: tumor necrosis factor-α MMP: matrix metalloproteinase; OPG: osteoprotegerin; RANKL: receptor activator of nuclear factor-κB ligand; miRNAs: MicroRNAs; BMD: bone mineral density; PFA: paraformaldehyde; NC: negative control; OE: overexpression; HE: Hematoxylin-eosin; PBS: phosphate-buffered saline; EDTA: ethylene diamine tetraacetic acid; DMEM: Dulbecco's Modified Eagle Medium; RT-qPCR: Reverse transcription quantitative polymerase chain reaction; GAPDH: glyceraldehyde phosphate dehydrogenase; UTR: untranslated region; WT: wild type; MUT: mutant type.