Search

Abstract We investigated the potential correlation between miR‐223 and NAcHT, LRR, and PYd domain‐containing protein 3 (NLRP3) in the context of renal ischemia‐reperfusion injury (RIRI), which is a leading cause of acute renal failure with significant mortality rates. Additionally, miR‐223 has been implicated in renal inflammation, further highlighting its relevance to this study. C57BL/6 male mice were used as RIRI models. After successful modeling, pathological examinations and serum creatinine and miR‐223 levels were tested. Pro‐inflammatory cytokine (IL‐1β, IL‐6, IL‐8, NLPR3, TLR4) expression was detected in mice by western blot (kidney tissue) and enzyme‐linked immunosorbent assay (serum). HK‐2 cells were used for in vitro experiments. A hypoxia/reoxygenation (H/R) model was used, and miR‐223 and pro‐inflammatory cytokine levels were detected using PCR and western blot assays, respectively. A dual‐luciferase reporter assay was conducted to confirm the binding of miR‐223 to NLPR3. Next, NLRP3 was knocked down to determine whether the anti‐inflammatory function of miR‐223 is dependent on NLRP3. MiR‐223 expression was lower in RIRI mice than in the sham operation group. The level of miR‐223 negatively correlated with serum creatinine levels and the severity of tubule injury. Increased proinflammatory cytokine levels in RIRI mice were observed. In vitro, miR‐223 alleviated the inflammatory response in H/R treated cells by inhibiting proinflammatory cytokines. Dual‐luciferase reporter and western blot assays confirmed the binding of miR‐223 to NLRP3. NLRP3 knockdown reversed the anti‐inflammatory effects of miR‐223 in HK‐2 cells. MiR‐223 plays an anti‐inflammatory role in RIRI by targeting NLRP3 to repress pro‐inflammatory factors.

Abstract Background Spinal cord injury (SCI) is characterized by extensive demyelination and inflammatory responses. Facilitating the clearance of lipid droplets (LDs) within microglia contributes to creating a microenvironment that favors neural recovery and provides essential materials for subsequent remyelination. Therefore, investigating MicroRNAs (miRNAs) that regulate lipid homeostasis after SCI and elucidating their potential mechanisms in promoting LDs clearance in microglia have become focal points of SCI research. Methods We established a subacute C5 hemicontusion SCI model in mice and performed transcriptomic sequencing on the injury epicenter to identify differentially expressed genes and associated pathways. Confocal imaging was employed to observe LDs accumulation. Multi-omics analyses were conducted to identify differentially expressed mRNA and miRNA post-SCI. Pathway enrichment analysis and protein-protein interaction network construction were performed using bioinformatics methods, revealing miR-223-Abca1 as a crucial miRNA-mRNA pair in lipid metabolism regulation. BV2 microglia cell lines overexpressing miR-223 were engineered, and immunofluorescence staining, western blot, and other techniques were employed to assess LDs accumulation, relevant targets, and inflammatory factor expression, confirming its role in regulating lipid homeostasis in microglia. Results Histopathological results of our hemicontusion SCI model confirmed LDs aggregation at the injury epicenter, predominantly within microglia. Our transcriptomic analysis during the subacute phase of SCI in mice implicated ATP-binding cassette transporter A1 (Abca1) as a pivotal gene in lipid homeostasis, cholesterol efflux and microglial activation. Integrative mRNA-miRNA multi-omics analysis highlighted the crucial role of miR-223 in the neuroinflammation process following SCI, potentially through the regulation of lipid metabolism via Abca1. In vitro experiments using BV2 cells overexpressing miR-223 demonstrated that elevated levels of miR-223 enhance ABCA1 expression in myelin debris and LPS-induced BV2 cells. This promotes myelin debris degradation and LDs clearance, and induces a shift toward an anti-inflammatory M2 phenotype. Conclusions In summary, our study unveils the critical regulatory role of miR-223 in lipid homeostasis following SCI. The mechanism by which this occurs involves the upregulation of ABCA1 expression, which facilitates LDs clearance and myelin debris degradation, consequently alleviating the lipid burden, and inhibiting inflammatory polarization of microglia. These findings suggest that strategies to enhance miR-223 expression and target ABCA1, thereby augmenting LDs clearance, may emerge as appealing new clinical targets for SCI treatment.

Abstract Background Lipid droplet (LD)-laden microglia is a key pathological hallmark of multiple sclerosis. The recent discovery of this novel microglial subtype, lipid-droplet-accumulating microglia (LDAM), is notable for increased inflammatory factor secretion and diminished phagocytic capability. Lipophagy, the autophagy-mediated selective degradation of LDs, plays a critical role in this context. This study investigated the involvement of microRNAs (miRNAs) in lipophagy during demyelinating diseases, assessed their capacity to modulate LDAM subtypes, and elucidated the potential underlying mechanisms involved. Methods C57BL/6 mice were used for in vivo experiments. Two weeks post demyelination induction at cervical level 4 (C4), histological assessments and confocal imaging were performed to examine LD accumulation in microglia within the lesion site. Autophagic changes were observed using transmission electron microscopy. miRNA and mRNA multi-omics analyses identified differentially expressed miRNAs and mRNAs under demyelinating conditions and the related autophagy target genes. The role of miR-223 in lipophagy under these conditions was specifically explored. In vitro studies, including miR-223 upregulation in BV2 cells via lentiviral infection, validated the bioinformatics findings. Immunofluorescence staining was used to measure LD accumulation, autophagy levels, target gene expression, and inflammatory mediator levels to elucidate the mechanisms of action of miR-223 in LDAM. Results Oil Red O staining and confocal imaging revealed substantial LD accumulation in the demyelinated spinal cord. Transmission electron microscopy revealed increased numbers of autophagic vacuoles at the injury site. Multi-omics analysis revealed miR-223 as a crucial regulatory gene in lipophagy during demyelination. It was identified that cathepsin B (CTSB) targets miR-223 in autophagy to integrate miRNA, mRNA, and autophagy gene databases. In vitro, miR-223 upregulation suppressed CTSB expression in BV2 cells, augmented autophagy, alleviated LD accumulation, and decreased the expression of the inflammatory mediator IL-1β. Conclusion These findings indicate that miR-223 plays a pivotal role in lipophagy under demyelinating conditions. By inhibiting CTSB, miR-223 promotes selective LD degradation, thereby reducing the lipid burden and inflammatory phenotype in LDAM. This study broadens the understanding of the molecular mechanisms of lipophagy and proposes lipophagy induction as a potential therapeutic approach to mitigate inflammatory responses in demyelinating diseases.

Abstract Background Neuroinflammation is a characteristic pathological change of Alzheimer’s Diseases (AD). Microglia have been reported to participate in inflammatory responses within the central nervous system. However, the mechanism of microglia released exosome (EXO) contribute to communication within AD microenvironment remains obscure. Methods The interaction between microglia and AD was investigated in vitro and in vivo. RNA-binding protein immunoprecipitation (RIP) was used to investigate the mechanisms of miR-223 and YB-1. The association between microglia derived exosomal YB-1/miR-223 axis and nerve cell damage were assessed using Western blot, immunofluorescence, RT-PCR, ELISA and wound healing assay. Results Here, we reported AD model was responsible for the M1-like (pro-inflammatory) polarization of microglia which in turn induced nerve cell damage. While M2-like (anti-inflammatory) microglia could release miR-223-enriched EXO which reduced neuroinflammation and ameliorated nerve damage in AD model in vivo and in vitro. Moreover, YB-1 directly interacted with miR-223 both in cell and EXO, and participated in microglia exosomal miR-223 loading. Conclusion These results indicate that anti-inflammatory microglia-mediated neuroprotection form inflammatory damage involves exporting miR-223 via EXO sorted by YB-1. Consequently, YB-1-mediated microglia exosomal sorting of miR-223 improved the nerve cell damage repair, representing a promising therapeutic target for AD.

ABSTRACT: Within the poultry industry, hens' reproductive performance is of great economic significance. The development and growth of follicles is a key aspect of hen egg production, and ovarian follicle growth and development are closely associated with granulosa cells (GCs) proliferation and the synthesis of steroid hormones. It has been confirmed by numerous studies that microRNAs (miRNAs) play important roles in the steroid hormone synthesis and proliferation of GCs. In this study, we examined the main miRNAs influencing hens' ability to reproduce, identified the miR-223 that is mainly expressed in atretic follicles based on sequencing, and investigated its role in GCs. Then, we used miR-223 mimic and inhibitor to knockdown or overexpress miR-223 expression. The result showed that miR-223 significantly inhibits both the steroid hormone synthesis and the proliferation of GCs. Subsequently, the results of the dual luciferase reporter experiment and bioinformatics prediction demonstrated that cysteine rich transmembrane BMP regulator 1 (CRIM1) was a downstream target gene of miR-223, and overexpression of miR-223 prevented CRIM1 expression. The function of CRIM1 was further investigated, and we observed a significant reduction in the synthesis of steroid hormones and the proliferation of GCs after transfection with CRIM1 siRNA. The opposite function of miR-223 was observed for CRIM1 in our study. Additionally, we demonstrated the involvement of the miR-223/CRIM1 axis in GCs through modulation of the AKT signaling pathway. Our data demonstrate the pivotal role of the miR-223 in the proliferation and steroid hormone synthesis of chicken GCs, which helps to explain how non-coding RNA (ncRNA) affects chicken reproductive function.

Objective To investigate the effects and potential mechanisms of osteoblast-derived exosomes on the biological properties of prostate cancer (PC) cells. Methods Human bone marrow mesenchymal stem cells was induced to differentiate into osteoblasts, and then the exosomes were extracted from the supernatant of cell culture medium by ultracentrifugation and identified. Clone formation assay was employed to determine the effect of derived exosomes on the proliferation of PC cells, and PCR microarray was used to analyze miRNA changes after exosome treatment. Fluorescence labeling was applied to validate the transfer of miRNA between osteoblasts and tumor cells. The impact of key miRNA on the biological characteristics of PC cells was studied at the cellular and animal levels. Bioinformatics analysis was conducted to explore the target genes of key miRNA and then further validated. Results Osteoblast-derived exosomes were successfully extracted and then identified. These exosomes significantly promoted the clonogenic ability of PC cells when compared with the cells without treatment (P < 0.05). PCR microarray showed that miR-223 might be a key molecule in this process. Membrane exchange experiments demonstrated the presence of membrane exchange between osteoblasts and PC cells, and miRNA FAM labeling displayed that miR-223 was transferred within above cells. Functional experiments indicated that overexpression of miR-223 enhanced the proliferation, anti-apoptosis, and migration/invasion ability of PC cells (P < 0.05). Bioinformatics analysis indicated that APC, an inhibitor of WNT signaling pathway, may be a target gene of miR-223, which was further confirmed that overexpressing miR-223 down-regulated APC expression. Rescue experiment showed that overexpression of APC reversed the promoting effect of miR-223 on the proliferation, migration and invasion of PC cells. Conclusion Osteoblast-derived exosomes are pro-carcinogenic factors in PC, and miR-223 may be a key molecule in exosomes, which plays a oncogenic role by regulating APC in PC cells.

Abstract Background Acute ischemic stroke (AIS) is the second leading cause of disability and death worldwide. Micro-RNA (miRNA)-223 was first identified as a regulator of hematopoietic lineage differentiation. Later, its diverse roles were discovered in a wide spectrum of pathological conditions. The present study aimed to assess the clinical and prognostic significance of miR-223 in patients with acute ischemic stroke (AIS). The study included 93 patients with AIS diagnosed on the basis of clinical and radiological findings. In addition, there were 50 healthy subjects who served as controls. Patients were classified into two categories: Those with favorable functional outcome (modified Rankin Scale (mRS): 0–2) and others with unfavorable functional outcome (mRS: 3–6) at 6 months post-stroke. Results The present prospective longitudinal study included 93 patients with AIS. They included 60 males (64.5%) and 33 females (35.5%) with an age of 64.5 ± 12.4 years. At the end of 6-month follow up, 44 patients (47.3%) had favorable outcome while the remainder 49 patients (52.7%) had unfavorable outcome. Patients with favorable outcome had significantly lower baseline miR-223 levels [median (IQR): 4.4 (2.0–6.3) versus 8.4 (4.5–14.9), p

Abstract Objectives To evaluate the diagnostic value of plasma exosomal miR-223 and its combination with CA125 for the diagnosis of early-stage epithelial ovarian cancer (EOC). Patients and methods Exosomes derived from the plasma of 78 EOC patients, 40 patients with epithelial benign ovarian tumors, and 52 healthy participants were isolated using the ultracentrifugation method and identified by transmission electron microscopy (TEM) and western blot. Results The expression of exosomal miR-223 was significantly upregulated in the plasma of EOC patients compared to that in healthy subjects and patients with benign diseases. The combination of exosomal miR-223 and CA125 from plasma had an equivalent area under the ROC curve (AUC) to CA125 alone for discriminating between EOC and non-EOC cases, including healthy subjects and benign ovarian tumors. However, the AUC value of the combination was 0.944 (95% CI: 0.899–0.990) for differentially diagnosing early-stage EOC from healthy subjects, slightly higher than that of CA125 alone (0.928, 95% CI: 0.875–0.981), with a sensitivity and specificity of 0.9784 and 0.885, respectively. Conclusion Our data suggest that plasma exosomal miR-223 can be used as a complement to CA125 to increase the diagnostic power for differentiating early-stage EOC from healthy subjects.

Background This study aimed to analyse the expression of microRNA-223 (miR-223) in embryo culture medium and its correlation with pregnancy outcomes.Methods Two hundred and two patients undergoing in vitro fertilisation/intracytoplasmic sperm injection (IVF/ICSI) were divided into clinical pregnancy group (n = 101) and non-pregnant group (n = 101). The baseline data, clinical indicators, and the expression level of miR-223 in the embryo medium were compared between the two groups. Logistic regression analysis was used to analyse the relationship between each index and the pregnancy outcome. Receiver operator characteristic curve was carried out to evaluate the differential ability of miR-223 in pregnancy status. Bioinformatics methods were used to identify the target genes of miR-223 and elucidate their functions.Results Compared with pregnancy group, the non-pregnancy group exhibited a reduction in miR-223 expression (p

Objective To investigate the potential effects of long non-coding RNA (lncRNA) MINCR on lipopolysaccharide (LPS)-induced human alveolar epithelial cell line A549 damage and inflammation through targeted regulation of miR-223. Methods A549 cells were randomly divided into control group, LPS group, LPS+transfection group(Si NC group, Si MINCR group, miR-223 NC group, miR-223 mimic group, Si NC+miR-223 NC group, Si NC+miR-223 antagomir group, Si MINCR+miR-223 NC group, and Si MINCR+miR-223 antagomir group). RT-qPCR was performed to determine the expression levels of MINCR and miR-223; Flow cytometry was performed to determine the rate of apoptosis; ELISA was performed to determine the levels of serum tumor necrosis factor-α(TNF-α) and interleukin-6 (IL-6); Bioinformatics prediction and dual luciferase reporter gene assay were performed to verify the targeting relationship of MINCR and miR-223; Western blot was performed to test the expression levels of activated caspase-3 (cleaved caspase-3) and B-cell lymphoma-2(Bcl-2) proteins in the cells. Results There was a targeting relationship between MINCR and miR-223. Compared with the control group, the apoptosis rate, the level of TNF-α and IL-6, and the expression of MINCR, cleaved caspase-3 proteins in the LPS group were significantly increased (PPPPPPPConclusions Silencing the expression of lncRNA MINCR may target the activation of miR-223 expression, inhibit cell apoptosis and inflammation, and reduce LPS-induced damage of alveolar epithelial cells.

Psoriasis is a chronic inflammatory skin disease with keratinocyte hyperproliferation and T cells as key mediators of lesional and systemic inflammatory changes. To date, no suitable differential biomarkers are available for the disease diagnosis. More recently, microRNAs have been identified as critical regulators of lesional and systemic immune changes in psoriasis with diagnostic potential. We have performed expression profiling of T cell-specific miRNAs in 38 plasma samples from psoriasis vulgaris patients and an equal number of age- and gender-matched healthy subjects. Our findings have identified a panel of five blood-based circulatory miRNAs with a significant change in their expression levels, comprising miR-215, miR-148a, miR-125b-5p, miR-223, and miR-142-3p, which can differentiate psoriasis vulgaris patients from healthy individuals. The receiver operating characteristic (ROC) curves for all five miRNAs individually and in combination exhibited a significant disease discriminatory area under the curve with an AUC of 0.762 and a p < 0.0001 for all the miRNAs together. Statistically, all five miRNAs in combination depicted the best-fit model in relation to disease severity (PASI) compared with individual miRNAs, with the highest R2 value of 0.94 and the lowest AIC score of 131.8. Each of the miRNAs also exhibited a significant association with at least one of the other miRNAs in the panel. Importantly, the five miRNAs in the panel regulate one or more immune-inflammation pathways based on target prediction, pathway network analysis, and validated roles in the literature. The miRNA panel provides a rationalized combination of biomarkers that can be tested further on an expanded cohort of patients for their diagnostic value.

Small noncoding RNAs, known as microRNAs (miRNAs), are vital for the regulation of diverse biological processes. miR-223, an evolutionarily conserved anti-inflammatory miRNA expressed in cells of the myeloid lineage, has been implicated in the regulation of monocyte–macrophage differentiation, proinflammatory responses, and the recruitment of neutrophils. The biological functions of this gene are regulated by its expression levels in cells or tissues. In this review, we first outline the regulatory role of miR-223 in granulocytes, macrophages, endothelial cells, epithelial cells and dendritic cells (DCs). Then, we summarize the possible role of miR-223 in chronic obstructive pulmonary disease (COPD), acute lung injury (ALI), coronavirus disease 2019 (COVID-19) and other pulmonary inflammatory diseases to better understand the molecular regulatory networks in pulmonary inflammatory diseases.

Objective To explore the effect of miR-223 on pulmonary lesions and IFN-γ in rats with pulmonary tuberculosis(PT) mechanism of protein expression. Methods Rats were divided into normal group (NC group), pulmonary tuberculosis rat model group (PT group) and pulmonary tuberculosis rat model transfected with miR-223 mimics group (miR-223 group) with 10 in each group. T cells in peripheral blood was quantified by flow cytometry, IFN-γ in serum was detected by ELISA and IL-18 levels, and the number of Mycobacterium tuberculosis colonies was counted; the pathological changes of lung tissue were micros copied by HE staining; IFN-γ in lung tissue was detected by immunohistochemistry method; RT-qPCR was used to detect miR-223 mRNA and IFN-γ in lung tissue MRNA expression. Results In PT group, the peripheral blood T cell subsets were disordered, the level of CD3+,CD4+% significantly decreased, and CD8+% was increased (P＜0.05); CD3+ and CD4+ percentage in miR-223 group were higher than those in PT group, while the percentage of CD8+ was lower(P＜0.05). Serum IFN-γ and IL-18 in PT group increased(P＜0.05); the number of Mycobacterium tuberculosis colonies in PT group was more than that in NC group (P＜0.05); the colony number of Mycobacterium tuberculosis in miR-223 group was significantly less than that in PT group (P＜0.05). Total IFN-γ level in PT group was higher than that in NC group(P＜0.05) but was significantly inhibited in miR-223 group(P＜0.05). Conclusions miR-223 can inhibit the pulmonary inflammatory response caused by Mycobacterium tuberculosis, and miR-223 showed an effect of anti-tuberculosis bacteria and may regulate host immunity, so and can effectively inhibit the expression of IFN-γ in lung tissue and thus alleviates severity of tuberculosis.

Yiming Yang, Di Wang, Kaixiong Tao, Guobin Wang Department of Gastrointestinal Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430033, People’s Republic of ChinaCorrespondence: Guobin Wang, Email wgb@hust.edu.cnPurpose: Colorectal cancer (CRC) is one of the most common carcinomas worldwide with a high mortality rate. Numerous studies suggest that circular RNA (circRNA) plays a crucial role in the progression of various carcinomas, including CRC. The present work focused on exploring the role and underlying molecular mechanism of action of the circRNA circLRCH3 in CRC.Methods: Real-time reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was conducted to detect expression levels of circLRCH3, miR-233, and lipoma preferred partner (LPP). The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to measure the proliferation of CRC cells and the transwell assay was used to evaluate cell migration and invasion capacity. A flow cytometry assay was used to analyze the effect of circLRCH3 on the distribution of the cell cycle and apoptosis of CRC cells. The expression of LPP was analyzed using Western blotting or an RT-qPCR assay. The relationship between miR-223 and circLRCH3, and that between miR-223 and LPP, was predicted and examined using bioinformatics analysis and luciferase reporter gene experiments. A xenograft tumor formation assay was also performed.Results: We found that the expression level of circLRCH3 was downregulated in CRC cells and negatively correlated with miR-223. The overexpression of circLRCH3 or silencing of miR-223 inhibited the growth, invasion, and migration of CRC cells, but promoted their apoptosis. In contrast, overexpression of miR-223 and depletion of LPP severally abrogated the tumor suppressive roles of circLRCH3 and miR-223 knockdown in CRC cells in vitro. The xenograft experiments in nude mice also proved the antitumor effect of circLRCH3.Conclusion: These results suggested that the circLRCH3/miR-223/LPP axis likely plays a critical role in CRC.Keywords: circLRCH3, miR-223, LPP, colorectal cancer, TCGA, GEO

Abstract Background Sevoflurane is considered as a lung-protective factor in acute lung injury (ALI), but the underlying molecular mechanism remains largely unknown. The present study identified for the first time that sevoflurane ameliorated lipopolysaccharide (LPS)-induced ALI through regulating a novel long non-coding RNA LINC00839, and uncovered its regulatory mechanism. Methods LPS-induced ALI models were established in mice or mouse pulmonary microvascular endothelial cells (MPVECs), and they were administered with sevoflurane. Real-Time quantitative PCR, western blot and bioinformatics analysis were performed to screen the aberrantly expressed long non-coding RNA and the downstream molecules in sevoflurane-treated ALI models, and their roles in the protection effect of sevoflurane were verified by functional recovery experiments. Results Sevoflurane relieved LPS-induced lung injury, cell pyroptosis and inflammation in vitro and in vivo. LINC00839 was significantly suppressed by sevoflurane, and overexpression of LINC00839 abrogated the protective effects of sevoflurane on LPS-treated MPVECs. Mechanismly, LINC00839 positively regulated NOD-like receptor protein 3 (NLRP3) via sequestering miR-223. MiR-223 inhibitor reversed the inhibitory effects of LINC00839 knockdown on NLRP3-mediated pyroptosis in LPS-treated MPVECs. Furthermore, both miR-223 ablation and NLRP3 overexpression abrogated the protective effects of sevoflurane on LPS-treated MPVECs. Conclusion In general, our work illustrates that sevoflurane regulates the LINC00839/miR-223/NLRP3 axis to ameliorate LPS-induced ALI, which might provide a novel promising candidate for the prevention of ALI.

Background/Purpose: MicroRNA-223 (miR-223) is involved in several inflammatory diseases, including gestational diabetes mellitus (GDM) and periodontitis. We first described a procedure for purifying miR-223 from gingival crevicular blood (GCB) of pregnant women with or without GDM and periodontitis. This study aimed to determine whether GDM and/or periodontitis modifies miR-223 expression in pregnant women and to analyze miR-223-targeted messenger RNA (mRNA) expression levels in GCB compared to peripheral blood (PB). Materials and methods: Pregnant women were allocated to 4 groups: 10 women with GDM and periodontitis (GDM/P), 10 women with GDM without periodontitis (GDM/NP), 9 women with periodontitis and without GDM (NGDM/P) and 10 women without either condition (NGDM/NP). Clinical parameters of GDM and periodontal status were examined. GCB and PB were collected to assess miR-223, ICAM-1, IL-1β and β1-integrin gene expression by quantitative real-time polymerase chain reaction. Results: The GDM/P group demonstrated the highest miR-223 expression levels among the 4 groups in GCB. A significant difference was found between GDM/P and GDM/NP group (P = 0.04). In contrast, the GDM/P showed the lowest miR-223 expression level in PB among the 4 groups. Moreover, ICAM-1 and IL-1β mRNA expression exhibited the opposite trend of miRNA-223, indicating that miRNA-223 might regulate the mRNA function of those genes by epigenetic events. Conclusion: The upregulation of miR-223 expression in GCB but downregulation in PB, ICAM-1 and IL-1β genes expression in women with GDM and periodontitis suggest a promising role of miR-223 in the association between GDM and periodontitis.

As skeletal muscle is one of the largest organs in the body, its damage can directly reflect a decline in somatic function, thus, further affecting daily life and health. Inflammation is a prerequisite for the repair of injured skeletal muscles. Chronic inflammation induced by inadequate repair in skeletal muscle aggravates tissue injury. Exosomes regulate inflammatory responses to facilitate the repair of skeletal muscle injury. Moreover, exosomal miR‐223 with high specificity is the most abundant miRNA in peripheral blood and regarded as biomarkers for inflammation post skeletal muscle injury, which warrants further investigation. Available studies have demonstrated that exosomal miR‐223 negatively correlates with TNF‐α levels in serum and regulates the canonical inflammatory NF‐κB signaling pathway. miR‐223 is a negative feedback regulator with great potential for adjusting inflammatory imbalance and promoting skeletal muscle repair. The research on the regulation of negative feedback factors in the inflammatory signaling pathway is essential in biology and medicine. Therefore, this review mainly elaborates the formation, heterogeneity and markers of exosomes and points out exosomal miR‐223 as a beneficial role in chronic skeletal muscle inflammation and can be expected to be a potential therapeutic target for skeletal muscle damage.

Abstract Background Excessive autophagic activity in alveolar epithelial cells is one of the main causes of acute lung injury (ALI), but the underlying molecular mechanism has not been fully elucidated. Previous studies have shown that microRNAs (miRs) are involved in regulating autophagy in several diseases. This study aimed to determine the role of miR-223 in excessive autophagic activity in alveolar epithelial cells and the underlying mechanism to identify a novel therapeutic targets for the development of new drugs to treat acute respiratory distress syndrome (ARDS). Methods A549 cells were treated with lipopolysaccharide (LPS) to establish an ALI in vitro model. The expression of miR-223 and its role of miR-223 in regulating oxidative stress and autophagy in the LPS-treated A549 cells, were examined using RT-PCR, flow cytometry and ELISA. A luciferase reporter assay was performed to verify the interaction between miR-223 and the high-mobility group box 2 (HMGB2) protein. Results The results showed that the LPS treatment downregulated miR-223 expression in alveolar epithelial cells. We further proved that miR-223 directly targeted the 3-untranslated region of the HMGB2 gene and the downregulation of miR-223 increased HMGB2 protein level, which activated the JNK signalling pathway and thus induced oxidative stress and autophagy in LPS-treated alveolar epithelial cells. Knockdown of HMGB2 protein deactivated the JNK signalling pathway and inhibited autophagy and oxidative stress in alveolar epithelial cells. Conclusions The results of this study suggest that miR-223 regulates oxidative stress and autophagy in alveolar epithelial cells by targeting HMGB2 via the JNK signalling pathway.

Inflammatory bowel disease (IBD) is a common inflammation-related intestinal disease. Studies have shown that excessive pyroptosis of intestinal cells is involved in the development of IBD. However, the regulatory mechanism of pyroptosis in IBD remains unclear. Here, our study purposed to clarify the underlying regulatory mechanism of miR-223 to promote pyroptosis in IBD. MiR-223 and Smad Nuclear Interacting Protein 1 (SNIP1) expression in colon tissues collected from IBD patients and healthy volunteers were evaluated using qRT-PCR. Cell viability and pyroptosis were evaluated by CCK8 and flow cytometry assay, respectively. Pyroptosis-related proteins and nuclear factor κB (NF-κB) signals were determined by WB. Dual-luciferase reporter gene assay was employed to investigate the binding relationship between miR-223 and SNIP1. MiR-223 was significantly upregulated in IBD colon tissues and cell models, while SNIP1 was significantly decreased. Silence of miR-223 markedly enhanced cell viability and inhibited pyroptosis in the IBD cell model. MiR-223 could bind to 3’-UTR of SNIP1 and SNIP1 could activate NF-κB signalling pathway. Further rescued experiment found that knockdown of SNIP1 dramatically abolished the bio-effects mediated by miR-223 silence on the cell viability and pyroptosis of the IBD cell model. Likewise, the inactivation of NF-κB signalling markedly weakened the regulatory roles of SNIP1 downregulation in the IBD cell model. Besides, inhibition of NF-κB signalling attenuated the pyroptosis-promoting effect of overexpressing miR-223. Our data suggested that miR-223 activated the NF-κB pathway via targeting SNIP1, thus promoting the process of cell pyroptosis, and ultimately participating in the pathogenesis of IBD.

We previously demonstrated that microRNA(miR)‐223 is overexpressed in intestinal tissue of infants with necrotizing enterocolitis (NEC). The objective of the current study was to identify the target gene of miR‐223 and to investigate the role of the miR‐223/nuclear factor I‐A (NFIA) axis in cellular functions that underpin the pathophysiology of NEC. The target gene of miR‐223 was identified by in silico target prediction bioinformatics, luciferase assay, and western blotting. We investigated downstream signals of miR‐223 and cellular functions by overexpressing the miRNA in Caco‐2 and FHs74 cells stimulated with lipopolysaccharide or lipoteichoic acid (LTA). NFIA was identified as a target gene of miR‐223. Overexpression of miR‐223 significantly induced MYOM1 and inhibited NFIA and RGN in Caco‐2 cells, while costimulation with LTA decreased expression of GNA11, MYLK, and PRKCZ. Expression levels of GNA11, MYLK, IL‐6, and IL‐8 were increased, and levels of NFIA and RGN were decreased in FHs74 cells. These potential downstream genes were significantly correlated with levels of miR‐223 or NFIA in primary NEC tissues. Overexpression of miR‐223 significantly increased apoptosis of Caco‐2 and FHs74 cells, while proliferation of FHs74 was inhibited. These results suggest that upon binding with NFIA, miR‐223 regulates functional effectors in pathways of apoptosis, cell proliferation, G protein signaling, inflammation, and smooth muscle contraction. The miR‐223/NFIA axis may play an important role in the pathophysiology of NEC by enhancing inflammation and tissue damage.

BackgroundKawasaki disease (KD) is an acute vasculitis that may result in permanent coronary artery damage with unknown etiology. Endothelial cell (EC) dysfunction and platelet hyperactivity are the hallmarks of KD. Platelets are involved in the development of endothelial dysfunction. MiR-223 transferred by platelet microparticles (PMPs) has been found to involve in the functional regulation of endothelial cells in sepsis. However, the role of platelet-derived miR-223 in endothelial dysfunction has not yet been investigated in KD.ObjectivesWe seek to investigate the role of platelet-derived miR-223 in endothelial dysfunction of KD vasculopathy.Methods and resultsForty-five acute KD patients and 45 matched controls were randomly recruited in the study. When co-cultured with human coronary artery endothelial cells (HCAECs), KD platelets with higher levels of miR-223 were incorporated into HCAECs, resulting in the horizontal transfer of miR-223. Using KD platelets, PMPs, and platelet-releasate from the same amount of blood co-cultured with HCAECs, we found the increased expression of miR-223 in HCAECs was primarily derived from KD platelets, rather than PMPs or free miRNAs from platelet- releasate. KD platelet-derived miR-223 attenuated TNF-α induced intercellular cell adhesion molecule-1 (ICAM-1) expression in HCAECs. KD platelet-derived miR-223 also suppressed the monocyte adhesion to HCAECs. In vivo, platelet-specific miR-223 knockout (PF4-cre: miR-223flox/flox) C57BL/6 mice and miR-223flox/flox C57BL/6 mice were used. Using Lactobacillus casei cell wall extract (LCWE) to establish KD murine model, we showed that in LCWE-injected PF4-cre: miR-223flox/flox mice, deficiency of platelet-miR-223 exacerbates the medial thickening of the abdominal aorta, increased ICAM-1 expression with concomitant CD45+ inflammatory cells infiltration into the endothelium compared to LCWE-injected miR-223flox/flox mice.ConclusionsThe horizontal transfer of platelet-derived miR-223 suppresses the expression of ICAM-1 in HCAECs, which at least in part attenuates leukocyte adhesion, thereby reducing endothelial damage in KD vasculitis

BackgroundMicroRNAs (miRNA, miR) have an undeniable physiological and pathophysiological significance and act as promising novel biomarkers. The aim of the study was to investigate blood-derived miRNAs and their association with long-term all-cause mortality in patients with multivessel disease (MVD) suffering from acute coronary syndrome (ACS).Materials and MethodsThis study was an observational prospective study, which included 90 patients with MVD and ACS. Expression of miR-125a, miR-125b, and miR-223 was analysed by polymerase chain reaction (PCR). Patients were followed-up for a median of 7.5 years. All-cause mortality was considered as the primary endpoint. Adjusted Cox-regression analysis was performed for prediction of events.ResultsElevated expression of miR-125b (>4.6) at the time-point of ACS was associated with increased long-term all-cause mortality (adjusted [adj.] hazard ratio [HR] = 11.26, 95% confidence interval [95% CI]: 1.15–110.38; p = 0.038). The receiver operating characteristic (ROC) analysis showed a satisfactory c-statistics for miR-125b for the prediction of long-term all-cause mortality (area under the curve [AUC] = 0.76, 95% CI: 0.61–0.91; p = 0.034; the negative predictive value of 98%). Kaplan–Meier time to event analysis confirmed an early separation of the survival curves between patients with high vs low expression of miR-125b (p = 0.003). An increased expression of miR-125a and miR-223 was found in patients with non-ST-segment elevation ACS (NSTE-ACS) as compared to those with ST-segment elevation myocardial infarction (STEMI) (p = 0.043 and p = 0.049, respectively) with no difference in the expression of miR-125b between the type of ACS.ConclusionIn this hypothesis generating study, lower values of miR-125b were related to improved long-term survival in patients with ACS and MVD. Larger studies are needed to investigate whether miR-125b can be used as a suitable predictor for long-term all-cause mortality.

Qi-Bin Yang,1,2 Ling-Qin Li,1 Quan-Bo Zhang,2,3 Yong-Long He,1 Qing-Sheng Mi,2 Jing-Guo Zhou4 1Department of Rheumatology and Immunology, Affiliated Hospital of North Sichuan Medical College, Nanchong, 637000, Sichuan Province, People’s Republic of China; 2Henry Ford Immunology Program, Departments of Dermatology and Internal Medicine, Henry Ford Health System, Detroit, MI, 48202, USA; 3Department of Gerontology, Affiliated Hospital of North Sichuan Medical College, Nanchong, 637000, Sichuan Province, People’s Republic of China; 4Department of Rheumatology and Immunology, Clinical Medical College and The First Affiliated Hospital of Chengdu Medical College, Chengdu, 610500, Sichuan Province, People’s Republic of ChinaCorrespondence: Qing-Sheng MiHenry Ford Immunology Program, Departments of Dermatology and Internal Medicine, Henry Ford Health System, 1 Ford Place, Detroit, MI, 48202, USATel +1-313-876-1017Email qmi1@hfhs.orgJing-Guo ZhouDepartment of Rheumatology and Immunology, Clinical Medical College and the First Affiliated Hospital of Chengdu Medical College, Chengdu, 610500, Sichuan Province, People’s Republic of ChinaTel +86-28-8301-6078Email jgzhou@cmc.edu.cnObjective: MicroRNAs were identified as master-switch molecules limiting acute inflammatory response. This study investigated the potential role of microRNA (miR)-223 in the mechanism of gout.Methods: Wild-type (WT) and miR-223 knock-out (KO) mice were used to evaluate the phenotypes of gout models. Inflammatory cytokines were measured in air pouch and peritoneal cavity lavage fluid. In addition to miR-223 level in gout patients, miR-223 and pro-inflammatory genes were examined in bone marrow-derived macrophages (BMDMs) from mice as well as peripheral blood mononuclear cells from healthy controls (HC) treated with monosodium urate (MSU) crystals in vitro.Results: MiR-223 was up-regulated in the early phase in BMDMs from WT mice after MSU challenge and decreased rapidly, and this was not observed in miR-223 KO mice in vitro. In addition, miR-223 was required for macrophages homeostasis. In comparison with WT mice in vivo, miR-223 deficiency exacerbated swelling index of MSU-induced inflammation in foot pad and ankle joint models. MiR-223 deficiency also markedly aggravated inflammatory cells infiltration and cytokines release including interleukin (IL)-1β, IL-6 and monocyte chemotactic protein-1 (MCP-1) in the air pouch and peritonitis models. In the in vitro experiments, miR-223 deficiency promoted the inflammatory response by targeting NLR family pyrin domain containing protein 3 (NLRP3). Besides, miR-223 level was down-regulated in gout patients and in HC exposed to MSU in vitro.Conclusion: MiR-223 was down-regulated in gout patients and miR-223 deficiency exacerbated inflammatory response in diverse murine models, suggesting that up-regulation of miR-223 could be a potential therapeutic strategy for alleviating gouty inflammation.Keywords: miR-223, gout, animal models, cytokines, monosodium urate

Abstract Introduction The pathogenesis of inflammatory bowel disease (IBD) has not yet been clarified and is closely related to several pro‐inflammatory factors. MicroRNA‐233 (miR‐223) might be involved in the development of IBD; however, the mechanism underlying its pathogenesis is unclear. In this study, we attempted to determine the role of miR‐223 in dextran sodium sulfate (DSS)‐induced colitis and explore the involvement of the IL‐6/STAT3 pathway in the development of intestinal mucosal inflammation. Materials and Methods Except control (WT) group, male C57BL/6 mice were provided DSS, then treated for with miR‐223 agomir or antagomir including DSS group, DSS + miR‐223 agomir (DSS + A) group, and DSS + miR‐223 antagomir (DSS + AN) group. The colitis symptoms were observed, the disease activity index (DAI) score were recorded daily, and colonic inflammation was evaluated by histopathological scoring. The expression of myeloperoxidase (MPO), cytokines and IL‐6/STAT3 pathway‐related proteins were measured. Results miR‐223 expression in the terminal ileum and colon was increased in the DSS group compared with the WT group. Colitis symptoms were significantly alleviated in the DSS + A group and exacerbated in the DSS + AN group after administration of the miR‐223 agomir and antagomir, respectively. MPO, tumor necrosis factor‐α, IL‐6, and IL‐17 were decreased and IL‐10 was increased in the DSS + A group, but these changes were reversed in the DSS + AN group. Gp130, p‐STAT3, Bcl‐2, and Bcl‐xl in the colon declined in the DSS + A group, but these levels increased in the DSS + AN group. Conclusions The upregulation of miR‐223 by agomir administration alleviated colonic inflammation in a DSS‐induced colitis model, which was likely mediated by inhibiting the production of pro‐inflammatory cytokines via the IL‐6/STAT3 signaling pathway. These findings provide evidence that miR‐223 might have potential therapeutic implications in IBD.

Abstract Background Mesenchymal stem cells (MSCs) have been utilized in treating acute graft-versus-host disease (aGvHD) as they show strong immunosuppressive capacity through the release of various mediators, including immunosuppressive molecules, growth factors, chemokines, and exosomes. MicroRNAs (miRNAs) derived from MSC exosomes (MSCs-Exo) play a critical role in the regulation of immune responses. However, the function of miRNAs in treating aGvHD remains unknown. Here, we performed expression profiling of exosome-miRNAs from human umbilical cord MSCs (huc-MSCs) and murine compact bone MSCs (mb-MSCs) to investigate their immunoregulation effects in aGvHD. Methods Huc-MSCs-Exo and mb-MSCs-Exo were isolated and constructed MSCs-Exo-derived miRNA expression profiling using high-throughput sequencing. High expression of miR-223 was identified in both kinds of MSCs-Exo by bioinformatics analysis and quantitative real-time PCR (qPCR). In vitro cell crawling assay, transmigration assay and adhesion assay were subsequently applied to investigate the regulation of miR-223 on T cells. MiR-223 target gene was analyzed by western blot, luciferase analysis, and qPCR. Moreover, murine aGvHD model was established by infusing splenocytes and bone marrow nuclear cells from C57BL/6j mice (H-2Kb) into BALB/c recipient mice (H-2Kd). For therapeutic effect, MSCs or miR-223 Agomir were injected via tail vein. The general conditions of the mice in each group were monitored. Hematoxylin-eosin (H&E) staining was used to detect pathological changes of mice spleen, liver, and intestine. Mechanistically, immunofluorescence and flow cytometry were used to evaluate donor T cell migration, and enzyme-linked immunosorbent assay (ELISA) was used to detect the expression of serum inflammatory cytokines IFN-γ, TNF-α, and IL-17. Results High-throughput sequencing revealed high expression of miR-223 in huc-MSCs-Exo and mb-MSCs-Exo. MiR-223 could restrain adhesion and migration of T cells by inhibiting ICAM-1 expression in mouse lymphatic endothelial cells. MiR-223Agomir infusion attenuated aGvHD clinical symptoms, reduced the donor T cell infiltration into the spleen, liver, and intestine, and decreased inflammatory cytokines IFN-γ, TNF-α, and IL-17. Conclusion MSCs-Exo-derived miR-223 could attenuate aGvHD in mice through decreasing donor T cell migration. Our results unveil a new role of MSCs-Exo containing miR-223 in the treatment of aGvHD.

Ibnu Purwanto,1 Didik Setyo Heriyanto,2 Irianiwati Widodo,2 Mohammad Hakimi,3 Mardiah Suci Hardianti,1 Teguh Aryandono,4 Sofia Mubarika Haryana5 1Division of Hematology and Medical Oncology, Department of Internal Medicine, Faculty of Medicine, Public Health, and Nursing, Gadjah Mada University/Dr Sardjito Hospital, Yogyakarta, Indonesia; 2Department of Anatomical Pathology, Faculty of Medicine, Public Health, and Nursing, Gadjah Mada University/Dr Sardjito Hospital, Yogyakarta, Indonesia; 3Department of Clinical Epidemiology and Biostatistics Unit, Faculty of Medicine, Public Health, and Nursing, Gadjah Mada University/Dr Sardjito Hospital, Yogyakarta, Indonesia; 4Department of Surgical Oncology, Faculty of Medicine, Public Health, and Nursing, Gadjah Mada University/Dr Sardjito Hospital, Yogyakarta, Indonesia; 5Department of Histology and Cell Biology, Faculty of Medicine, Public Health, and Nursing, Gadjah Mada University/Dr Sardjito Hospital, Yogyakarta, IndonesiaCorrespondence: Ibnu PurwantoDivision of Hematology and Medical Oncology, Department of Internal Medicine, Faculty of Medicine, Public Health, and Nursing, Gadjah Mada University/Dr Sardjito Hospital, Jalan Farmako, Sekip Utara, Yogyakarta 55281, IndonesiaTel/Fax (+62 274) 560300Email ibnupurwanto@ugm.ac.idPurpose: Determining the optimal strategy to implement systemic treatment modalities has been challenging in triple-negative breast cancer (TNBC). We aim to investigate the role of microRNA-223 (miR-223) as prognostic factor and predictor of response toward chemotherapy in TNBC.Patients and Methods: We retrospectively analyzed the association of pretreatment miR-223 expression with clinicopathologic characteristics and 36-month overall survival (OS) of 53 all stages TNBC patients. Tumor level of miR-223 was measured using real-time quantitative polymerase chain reaction (expressed in fold change). Cutoff value for miR-223 was determined by using receiver operating curve (ROC). Kaplan–Meier curve was used to perform survival analysis.Results: The optimum cutoff value for miR-223 was 23.435 (AUC: 0.706, 95%CI: 0.565– 0.848; p:0.01; sensitivity: 78.6%; specificity: 56%) and was used to categorize mir-223 expression into over- and underexpressed group. Overexpression of miR-223 was associated with increased expression of EGFR (69.7% vs 35%, p: 0.022) and lower 36-month OS (33.3% vs 70%; median OS±SE (months): 25.66± 1.58 vs 30.23± 1.99; log rank p< 0.05). Worse survival is observed in miR-223 overexpressed group receiving platinum-based chemotherapy compared to miR-223 underexpressed group (mean OS (95%CI) months: 24.7 (20.3– 29.1) vs 34.3 (31.2– 37.4); p< 0.01), while no significant difference observed in non-platinum containing regimen. No significant association was observed between miR-223 expression with other clinicopathologic characteristics.Conclusion: Overexpression of miR-223 is associated with increased expression of EGFR, worse prognosis, and resistance toward platinum-based chemotherapy in Indonesian TNBC patients.Keywords: miR-223, chemotherapy, prognosis, EGFR, TNBC

Inflammatory bowel diseases (IBD) are characterized by chronic inflammation and damage of colonocytes with etiology of genetic, epigenetic and environmental factors. MicroRNA-223 (miR-223) has been found to be increased in both IBD patients and animal colitis models. However, contentious opinions relevant to the roles of miR-223 in IBD have been reported. Notwithstading that most studies have described that miR-223 has anti-inflammatory effects, several reports have progressed a pro-inflammatory view. In this review, we summarise both the anti-inflammatory and pro-inflammatory effects of miR-223 on key molecules in inflammatory responses in both animal models and in patients diagnosed with IBD and objectively discuss the possible basis for the discrepancies.

Huijie Gao,1 Jincheng Ma,1 Yanhui Cheng,1 Peiming Zheng2 1Department of Oncology, The First Affiliated Hospital of Henan University, Kaifeng, Henan 475000, People’s Republic of China; 2Department of Clinical Laboratory, Henan Provincial People’s Hospital, People’s Hospital of Zhengzhou University, People’s Hospital of Henan University, Zhengzhou, Henan 450003, People’s Republic of ChinaCorrespondence: Peiming Zheng Email zpm8266@163.comPurpose: Macrophages are a major component of the tumour microenvironment and play an important role in chemoresistance of cancer. However, how exosomal microRNAs (miRNAs) derived from macrophages contribute to the development of doxorubicin resistance in gastric cancer (GC) are not clearly defined. The aim of this study was to investigate whether macrophage-derived exosomes mediate doxorubicin resistance in GC.Methods: Exosomes isolated from macrophage culture medium were characterized and co-cultured with GC cells and the miR-223 level was detected using real-time quantitative PCR (RT-qPCR). The internalization of exosomes and transfer of miR-223 were observed via immunofluorescence. Macrophages were transfected with an miR-223 inhibitor or negative control. Cell Counting Kit-8 and flow cytometry were employed to explore the effect of macrophage-derived exosomes on the doxorubicin resistance of GC cells. Western blot and RT-qPCR assay were also performed to explore the regulation of GC chemotherapy resistance by exosomal miR-223.Results: Here, the macrophages and macrophage-derived exosomes promoted doxorubicin resistance in GC cells. MiR-223 was enriched in macrophage-derived exosomes and they could be transferred to co-cultivated GC cells. The miR-223 knockdown in macrophages could reduce the effects of exosomes on GC cells. Functional studies revealed that exosomal miR-223 derived from macrophages promoted doxorubicin resistance in GC cells by inhibiting F-box and WD repeat domain-containing 7 (FBXW7). Clinically, the expression of miR-223 significantly increased in GC tissues and high expression of plasma exosomal miR-223 was highly linked with doxorubicin resistance in GC patients.Conclusion: The exosomal transfer of macrophage-derived miR-223 conferred doxorubicin resistance in GC and targeting exosome communication may be a promising new therapeutic strategy for GC patients.Keywords: exosome, miR-223, macrophage, doxorubicin resistance, gastric cancer

Benzo(a)pyrene [B(a)P], a ubiquitous environmental pollutant, causes lung inflammatory damage. Pyroptosis，a new inflammation-dependent programmed cell death, happened when pyroptosis-related GSDMD is activated mediated by NLRP3 inflammasome. microRNA-223 (miRNA-223) is involved in inflammatory diseases by regulating NLRP3. However, whether GSDMD regulate NLRP3 inflammasome through miR-223 in B(a)P induced lung inflammatory injury remain unknown. In this study, alveolar epithelial cells (A549) were stimulated with 0, 2, 4, 8 μM B(a)P for 12 h or 24 h. The inflammatory injury and pyroptosis were determined. And the activation of NLRP3 inflammasomes and the level of miRNA-223 were detected. Then, the change of inflammatory injury and activation of NLRP3 inflammasomes in B(a)P-induced A549 cells were detected after inhibiting of GSDMD or miR-223 using siRNA-GSDMD (siGSDMD) or miR-223 inhibitor, respectively. Our results indicated that after B(a)P exposure, TNF-α and IL-6 in the supernatant were increased. Transmission electron microscope (TEM) results showed that A549 cells were obviously swollen and the cell membrane ruptured. Hoechest33342/PI staining showed that pyroptosis occurred. NLRP3, IL-1β, IL-18, GSDMD, GSDMD-N, pro caspase-1 and cleaved caspase-1 were significantly increased. Additionally, after transfecting with siGSDMD in B(a)P-induced A549 cells, the expression level of miR-223 was significantly increased. But IL-6 and TNF-α in the supernatant, the expression of NLRP3, IL-1β, IL-18 and cleaved caspase-1 protein were also decreased. And after inhibiting miR-223 in B(a)P-induced A549 cells, the expression of TNF-α and IL-6 in the supernatant, the protein expression of NLRP3, IL-1β, IL-18 and cleaved caspase-1 were increased. In conclusion, GSDMD may regulate NLRP3 inflammasome through miR-223, which is involved in B(a)P induced inflammatory damage in A549 cells.

BackgroundF-box and WD repeat domain-containing 7 (Fbw7) is well known as a tumor suppressor and ubiquitin ligase which targets a variety of oncogenic proteins for proteolysis. We previously reported that Fbw7 promotes apoptosis in diffuse large B-cell lymphoma (DLBCL) through Fbw7-mediated ubiquitination of Stat3. This study aimed to identify the mechanism of Fbw7-mediated aerobic glycolysis reprogramming in DLBCL.MethodsExpression levels of Fbw7 and Lactate Dehydrogenase A (LDHA) in human DLBCL samples were evaluated by immunohistochemistry. Crosstalk between Fbw7 and LDHA signaling was analyzed by co-immunoprecipitation, ubiquitination assay, western blotting and mRNA quanlitative analyses. In vitro and in vivo experiments were used to assess the effect of the Fbw7-mediated LDHA/lactate/miR-223 axis on DLBCL cells growth.ResultsFbw7 could interact with LDHA to trigger its ubiquitination and degradation. Inversely, lactate negatively regulated Fbw7 via trigging the expression of miR-223, which targeted Fbw7 3’-UTR to inhibit its expression. In vivo and in vitro experiments revealed that miR-223 promoted tumor growth and that the effects of miR-223 on tumor growth were primarily related to the inhibition of Fbw7-mediated LDHA’s ubiquitination.ConclusionsWe demonstrated that the ubiquitin-ligase Fbw7 played a key role in LDHA-related aerobic glycolysis reprogramming in DLBCL. Our study uncovers a negative functional loop consisting of a Fbw7-mediated LDHA/lactate/miR-223 axis, which may support the future ABC-DLBCL therapy by targeting LDHA-related inhibition.

MicroRNAs (miRNAs) are endogenous non-coding single-stranded small molecule RNAs consisting of 20–24 nucleotides that are highly conserved in species evolution. Expression of miRNAs is strictly tissue-specific, and it is chronological in fungi and plants, as well as in animals. MiR-223 has been shown to play a key role in innate immunity, and dysregulation of its expression contributes to the pathogenesis of multiple inflammatory diseases, and cancers. In this article the biosynthesis and functions of miR-223 in innate immunity are reviewed, and the role of miR-223 in liver physiopathology and therapeutic prospects are highlighted.

Cervical cancer is induced by persistent infections with high‐risk human papillomaviruses (HPVs), which produce the early protein 6 of HPVs (E6)/E7 protein that is involved in cell transformation by interacting with several oncoproteins or tumor suppressors. However, the role of noncoding RNA in mediating the pathogenesis of cervical cancer remains unclear. Here, we report that the novel signal transducer and activator of transcription 3 (STAT3)‐microRNA‐223‐3p (miR‐223)‐TGFBR3/HMGCS1 axis regulated by the E6 protein controls cervical carcinogenesis. miR‐223 was highly expressed in cervical tumor tissues, whereas TGFBR3 or HMGCS1 was significantly downregulated. miR‐223 targeted the 3′‐UTRs of TGFBR3 and HMGCS1 and suppressed their expression, leading to increased anchorage‐independent growth and cervical squamous cell carcinoma (CSCC) tumor growth in vitro and in vivo. The increased expression of miR‐223 was mediated by the transcription factor STAT3, whose activity was enhanced by E6 in the context of interleukin (IL)‐6 stimulation. In addition, exosomal miR‐223 derived from CSCC cells induced IL‐6 secretion by monocyte/macrophage in a coculture system in vitro, and IL‐6 secretion, in turn, led to enhanced STAT3 activity in CSSC cells, forming a positive feedback loop. Furthermore, modified miR‐223 inhibitor effectively suppressed tumor growth in cell line‐derived xenograft model, suggesting that miR‐223 is a potential promising therapeutic target in CSCC. In conclusion, our results demonstrate that the STAT3‐miR‐223‐HMGCS1/TGFBR3 axis functions as a key signaling pathway in cervical cancer progression and provides a new therapeutic target.

Abstract Background Activation of hepatic stellate cells (HSCs) is a prominent driver of liver fibrosis. We previously demonstrated that exosomes derived from natural killer (NK) cells (NK-Exo) attenuated TGF-β1-induced HSC activation. Herein, this study was designed to investigate the mechanism underlying the action of NK-Exo. Methods NK-Exo was isolated from NK-92MI cells and then administered into TGF-β1-treated LX-2 (human HSC line) cells. MiR-223 expression in NK-Exo was downregulated by transfecting NK-92MI cells with miR-223 inhibitor followed by exosome isolation. The HSC activation was evaluated by determining cell proliferation using CCK-8 assay and measuring the protein levels of α-SMA and CoL1A1 using western blot in LX-2 cells. The expression of miR-223 was detected by qRT-PCR. The interaction between miR-223 and ATG7 was analyzed by a dual-luciferase activity assay. The autophagy was evaluated by measuring the autophagy-related proteins using western blot. Results miR-223 was highly expressed in NK-Exo and inhibition of miR-223 expression in NK-Exo abrogated the inhibitory effect of NK-Exo on TGF-β-induced HSC activation. ATG7 was confirmed as a direct target of miR-223. Furthermore, treatment with the autophagy activator rapamycin and ATG7 overexpression in LX-2 cells abolished the HSC activation-suppressive effect of NK-Exo. Conclusion NK-Exo attenuated TGF-β-induced HSC activation by transferring miR-223 that inhibited autophagy via targeting ATG7.

Abstract Background Cisplatin is widely used as a first-line treatment for non-small cell lung cancer (NSCLC), but chemoresistance remains a major clinical obstacle for efficient use. As a microRNA, miR-223 was reported to promote the doxorubicin resistance of NSCLC. However, whether miR-223 is also involved in cisplatin resistance of NSCLC and the mechanism miR-223 involved in drug resistance is unclear. Accumulated evidence has shown that abnormal autophagy is associated with tumor chemoresistance. The study aimed to study the role of miR-223 on cisplatin sensitivity in NSCLC and uncover the potential mechanisms. Methods NSCLC cells transfected with mimic or inhibitor for miR-223 was assayed for chemoresistance in vitro. MiR-223 expression was assessed by quantitative real-time PCR (qRT-PCR). Western blot were used to study the expression level of F-box/WD repeat-containing protein 7 (FBXW7) and autophagy-related protein. The effect of miR-223 on cisplatin sensitivity was examined by using CCK-8, EdU assays and Autophagic flux assay. Luciferase assays, EdU assays and small interfering RNA were performed to identify the targets of miR-223 and the mechanism by which it promotes treatment resistance. Xenograft models were established to investigate the effect of mir-223 on cisplatin sensitivity. Results In the present study, we found that the level of miR-223 was significantly positively correlated with cisplatin resistance. MiR-223 overexpression made NSCLC cells resistant to cisplatin treatment. We further found that autophagy mediated miR-223-mediated cisplatin resistance in NSCLC cells. Further mechanistic research demonstrated that miR-223 directly targeted FBXW7. The overexpression of miR-223 could inhibit the level of FBXW7 protein expression, thus promoting autophagy and making NSCLC cells resistant to cisplatin. Finally, we confirmed the increased effect of cisplatin sensitivity by miR-223 Antagomir in xenograft models of NSCLC. Conclusions Our results demonstrate that miR-223 could enhance autophagy by targeting FBXW7 in NSCLC cells. Inhibition of autophagy by miR-223 knockdown provides a novel treatment strategy to alleviate cisplatin resistance in NSCLC.

Anli Hou,1 Yali Zhang,2 Yujuan Fan,1 Yi Zheng,3 Xiuying Zhou,1 Huilan Liu1 1Department of Gynaecology, University of Chinese Academy of Sciences Shenzhen Hospital, Shenzhen City, Guangdong Province 518106, People’s Republic of China; 2Department of Gynaecology, Affiliated Hospital of Chengde Medical University, Chengde, Hebei 067000, People’s Republic of China; 3Department of Central Lab, University of Chinese Academy of Sciences Shenzhen Hospital, Shenzhen City, Guangdong Province 518106, People’s Republic of ChinaCorrespondence: Anli HouDepartment of Gynaecology, University of Chinese Academy of Sciences Shenzhen Hospital, No. 4253 Songbai Road, Matian Street, Guangming District, Shenzhen City, Guangdong Province 518106, People’s Republic of ChinaTel +86-755-27548704Email idmbze9672487918@126.comIntroduction: The incidence of cervical squamous cell carcinoma (CSCC) has expanded in recent years. However, the function of long non-coding RNA (lncRNA) MAGI2-AS3 in the occurrence and progression of CSCC remains unclear. Therefore, the role of lncRNA MAGI2-AS3 in cervical squamous cell carcinoma (CSCC) was investigated in our study.Methods: We used qRT-PCR analysis to identify the level of MAGI2-AS3 mRNA expression in CSCC clinical samples and cell lines. We investigated cell migration and invasion of CSCC cells transfected with MAGI2-AS3, miR-233 mimic, or EPB41L3 with transwell assays. Bioinformatics analysis and a luciferase reporter assay were employed to predict the interaction between MAGI2-AS3 and miR-233.Results: We found that MAGI2-AS3 and EPB41L3 were both downregulated in CSCC and the expression of this two was positively correlated. Bioinformatics analysis showed that MAGI2-AS3 might bind to miR-233, which could directly target EPB41L3. In CSCC cells, overexpression of MAGI2-AS3 led to upregulated, while overexpression of miRNA-233 led to downregulated expression of EPB41L3. However, MAGI2-AS3 and miR-233 did not affect the expression of each other. In addition, overexpression of MAGI2-AS3 and EPB41L3 led to inhibited cancer cell invasion and migration, while overexpression of miR-233 played an opposite role and attenuated the effects of overexpressing MAGI2-AS3.Conclusion: MAGI2-AS3 may sponge miR-233 to upregulate EPB41L3, thereby inhibiting CSCC cell invasion and migration.Keywords: lncRNA MAGI2-AS3, cervical squamous cell carcinoma, EPB41L3, miR-223

Abstract Background Neutrophilic subtype asthma occurs in approximately 15–25% of the asthma cases and is associated with severe airflow obstruction, corticosteroid resistance. MicroRNA plays a vital role in regulating many immune processes, but how miRNA circuits coordinate airway inflammation during neutrophilic asthma is unclear. Methods To investigate the molecular mechanism of miR-223 in regulation of neutrophilic airway inflammation, miR-223 knockout mice were used to the OVA/CFA-induced neutrophilic asthma or treated with NLRP3 inhibitor and IL-1β receptor antagonist. Based on the results obtained, wide-type mice were subsequently treated with miR-223 agomirs or negative control agomirs, and the effects on airway inflammation were assessed using morphometric techniques, quantitative RT-PCR, western blot, ELISA and other molecular approaches. Results The expression of miR-223 was upregulated in lung tissues of experimental mice model. Furthermore, miR-223−/− mice led to aggravated neutrophilic airway inflammation with heightened histopathological, inflammatory cells and cytokines readouts. Moreover, miR-223−/− mice also presented with enhanced NLRP3 inflammasome level with elevated IL-1β. Blocking NLRP3 or IL-1β diminished this phenotype. Finally, overexpression of miR-223 via treatment with miR-223 agomirs attenuated airway inflammation, NLRP3 levels and IL-1β release. Conclusions The findings of this study revealed a crucial role for miR-223 in regulating the immunoinflammatory responses by depressing the NLRP3/ IL-1β axis in neutrophilic asthma.

Abstract Background Mast cells (MCs) have been found to play a critical role during development of inflammatory bowel disease (IBD) that characterized by dysregulation of inflammation and impaired intestinal barrier function. However, the function of MCs in IBD remains to be fully elucidated. Results In our study, we used exosomes isolated from human mast cells-1 (HMCs-1) to culture with NCM460, HT-29 or CaCO2 of intestinal epithelial cells (IECs) to investigate the communication between MCs and IECs. We found that MCs-derived exosomes significantly increased intestinal epithelial permeability and destroyed intestinal barrier function, which is attributed to exosome-mediated functional miRNAs were transferred from HMCs-1 into IECs, leading to inhibit tight junction-related proteins expression, including tight junction proteins 1 (TJP1, ZO-1), Occludin (OCLN), Claudin 8 (CLDN8). Microarray and bioinformatic analysis have further revealed that a panel of miRNAs target different tight junction-related proteins. Interestingly, miR-223 is enriched in mast cell-derived exosome, which inhibit CLDN8 expression in IECs, while treatment with miR-223 inhibitor in HT-29 cells significantly reversed the inhibitory effect of HMCs-1-derived exosomes on CLDN 8 expression. Most importantly, enrichment of MCs accumulation in intestinal mucosa of patients with IBD compared with those healthy control. Conclusions These results indicated that enrichment of exosomal miR-223 from HMCs-1 inhibited CLDN8 expression, leading to destroy intestinal barrier function. These finding provided a novel insight of MCs as a new target for therapeutic treatment of IBD.

MicroRNAs (miRNAs) are diminutive noncoding RNAs that can influence disease development and progression by post-transcriptionally regulating gene expression. The anti-inflammatory miRNA, miR-223, was first identified as a regulator of myelopoietic differentiation in 2003. This miR-223 exhibits multiple regulatory functions in the immune response, and abnormal expression of miR-223 is shown to be associated with multiple infectious diseases, including viral hepatitis, human immunodeficiency virus type 1 (HIV-1), and tuberculosis (TB) by influencing neutrophil infiltration, macrophage function, dendritic cell (DC) maturation and inflammasome activation. This review summarizes the current understanding of miR-223 physiopathology and highlights the molecular mechanism by which miR-223 regulates immune responses to infectious diseases and how it may be targeted for diagnosis and treatment.

Dendritic cells (DCs) are key mediators of transplant rejection. Numerous factors have been identified that regulate transplant immunopathology by modulating the function of DCs. Among these, microRNAs (miRNAs), small non-coding RNA molecules, have received much attention. The miRNA miR-223 is very highly expressed and tightly regulated in hematopoietic cells. It plays an important role in modulating the immune response by regulating neutrophils and macrophages, and its dysregulation contributes to multiple types of immune diseases. However, the role of miR-223 in immune rejection is unclear. Here, we observed expression of miR-223 in patients and mice who had undergone heart transplantation and found that it increased in the serum of both, and also in DCs from the spleens of recipient mice, although it was unchanged in splenic T cells. We also found that miR-223 expression decreased in lipopolysaccharide-stimulated DCs. Increasing the level of miR-223 in DCs promoted polarization of DCs toward a tolerogenic phenotype, which indicates that miR-223 can attenuate activation and maturation of DCs. MiR-223 effectively induced regulatory T cells (Tregs) by inhibiting the function of antigen-presenting DCs. In addition, we identified Irak1 as a miR-223 target gene and an essential regulator of DC maturation. In mouse allogeneic heterotopic heart transplantation models, grafts survived longer and suffered less immune cell infiltration in mice with miR-223-overexpressing immature (im)DCs. In the miR-223-overexpressing imDC recipients, T cells from spleen differentiated into Tregs, and the level of IL-10 in heart grafts was markedly higher than that in the control group. In conclusion, miR-223 regulates the function of DCs via Irak1, differentiation of T cells into Tregs, and secretion of IL-10, thereby suppressing allogeneic heart graft rejection.

This study aimed to explore whether microRNA (miR) 223 affects microglial autophagy by targeting autophagy-related 16-like 1 (ATG16L1) in the kainic acid (KA) model of temporal lobe epilepsy (TLE). The miRNA and mRNA expression levels were quantified using quantitative real-time polymerase chain reaction (qRT-PCR), and the protein expression was investigated using western blotting. A dual-luciferase reporter assay was used to test the direct interaction between miR 223 and ATG16L1. In situ hybridization was performed to measure the hippocampal expression of miR 223. We used immunofluorescence staining to assess the expression of ATG16L1 and microtubule-associated protein light chain 3 (LC3) in the murine hippocampal microglia. Inhibitor of miR 223 was utilized to investigate the role of miR 223 in TLE, and the epileptic activity was assessed using electroencephalography (EEG). The autophagosomes were observed by transmission electron microscopy. In patients with TLE, the murine KA model of TLE, and the KA-stimulated BV2 cells, miR 223, and sequestosome 1 (SQSTM1/P62) expressions were remarkably increased, whereas ATG16L1 and LC3 levels were significantly decreased. Using a dual-luciferase reporter assay, ATG16L1 was determined as a direct target of miR 223. Treatment with antagomir 223 alleviated epilepsy, prevented abnormalities in EEG recordings and increased the ATG16L1 and LC3 levels in KA-treated mice. Inhibition of miR 223 induced increased autophagy in BV2 cells upon Rapamycin stimulation. These findings show that miR 223 affects microglial autophagy via ATG16L1 in the KA model of TLE. The miR 223/ATG16L1 pathway may offer a new treatment option for TLE.

Patients diagnosed with prostate cancer often have a poor prognosis and limited treatment options, as the specific pathogenesis remains to be elucidated. Circular RNA (circRNA) is a type of non-coding RNA that interacts with microRNA (miRNA/miR) and transcription factors to regulate gene expression. However, little is known about specific circRNAs that serve roles in the pathogenesis of prostate cancer. Findings of the present study confirmed that circRNA G protein subunit γ 4 (circGNG4) was upregulated in prostate cancer tissues and cell lines. Knockdown of circGNG4 inhibited the malignant behavior of prostate cancer cells. Furthermore, bioinformatics were used to predict targeting interactions between circGNG4 or miR-223 and EYA transcriptional coactivator and phosphatase 3 (EYA3)/c-Myc mRNA. miR-223 inhibited the malignant behavior of prostate cancer cells, while EYA3/c-Myc had the opposite effect. circGNG4 enhanced the expression of EYA3/c-Myc by sponging miR-223 to promote the growth of prostate cancer tumors in vivo. In conclusion, the circGNG4/miR-223/EYA3/c-Myc regulatory pathway promoted the malignant progression of prostate cancer. The results of the present study may provide potential new targets for the diagnosis or treatment of prostate cancer.

MiR-223 is a miRNA with important functions in apoptosis, carcinogenesis, and inflammation, and it was demonstrated to be over-expressed in brain tissue after traumatic brain injury (TBI). However, few studies have focused on its role in protecting brain microvascular endothelial cells (BMECs). This study evaluated the protective effect of miR-223 on BMECs after stretch injury (SI). bEnd.3 cells (BMECs of mouse) were transfected with overexpressing and blocking lentivirus of miR-223, then were subjected to SI. After immunofluorescence assay, it was demonstrated that miR-223 overexpression significantly rescued the SI-induced loss of ZO-1 (Zonula Occludens 1, tight junction protein) (p < 0.01), while miR-223 blocking exacerbated the loss of ZO-1 (p < 0.05). Flow cytometry confirmed a significant increase in the proportion of apoptotic bEnd.3 cells after SI, and miR-223 overexpression reduced this proportion (p < 0.001). The result of Western blot revealed that miR-223 overexpression significantly reduced the expression of cleaved caspase-3 (cl-caspase 3) (p < 0.05) and RhoB (p < 0.01), while miR-223 blocking increased the expression of these proteins (p < 0.05, p < 0.001). Additionally, knockdown of RhoB significantly reduced the expression of cl-caspase 3 (p < 0.001). These findings suggested that miR-223 can alleviate SI-induced apoptosis of BMECs, and this anti-apoptotic effect is at least partially achieved by inhibiting the expression of RhoB. Moreover, miR-223 may play a role in maintaining the integrity of BBB during TBI.

MicroRNAs (miRNAs) are small, non-coding RNAs that negatively regulate target mRNA expression, and altered expression of miRNAs is associated with liver pathological conditions. Recent studies in animal models have shown neutrophil/myeloid-specific microRNA-223 (miR-223) as a key regulator in the development of various liver diseases including fibrosis, where hepatic stellate cells (HSCs) are the key player in pathogenesis. However, the precise roles of miR-223 in human HSCs and its therapeutic potential to control fibrosis remain largely unexplored. Using primary human HSCs, we demonstrated that miR-223 suppressed the fibrogenic program and cellular proliferation while promoting features of quiescent HSCs including lipid re-accumulation and retinol storage. Furthermore, induction of miR-223 in HSCs decreased cellular motility and contraction. Mechanistically, miR-223 negatively regulated expression of smooth muscle α-actin (α-SMA) and thus reduced cytoskeletal activity, which is known to promote amplification of fibrogenic signals. Restoration of α-SMA in miR-223-overexpressing HSCs alleviated the antifibrotic effects of miR-223. Finally, to explore the therapeutic potential of miR-233 in liver fibrosis, we generated co-cultured organoids of HSCs with Huh7 hepatoma cells and challenged them with acetaminophen (APAP) or palmitic acid (PA) to induce hepatotoxicity. We showed that ectopic expression of miR-223 in HSCs attenuated fibrogenesis in the two human organoid models of liver injury, suggesting its potential application in antifibrotic therapy.

The health of mammary gland is essential for lactation. Epidermal growth factor (EGF) is reported to play an important role in lactation initiation and miR-223 is a conserved microRNA in anti-inflammation. In this study, EGF was found to induce a higher expression of miR-223 in goat mammary epithelial cell (gMEC). The downstream genes of miR-223 were screened by RNA sequencing, including Interferon-stimulated gene product 15 (ISG15), a pivotal immune responder, which was detected to be downregulated by EGF and miR-223. Due to the correlation between inflammation and apoptosis, the gMEC apoptosis modulated by EGF, miR-223, and ISG15 was investigated, and the protein expressions of Bcl-2/Bax, Caspase 3 and p53 were examined to evaluate the apoptosis of gMEC. The protein expressions of p-STAT3/STAT3, PR, FOXC1, and HOXA10, which had been shown to be related to inflammation, were detected to assess the inflammation of gMEC. This study provided a regulation axis, EGF/miR-223/ISG15, and illustrated its regulation to gMEC apoptosis and inflammation.

Kawasaki disease (KD), an acute febrile childhood illness and systemic vasculitis of unknown etiology, is the leading cause of acquired heart disease among children. Experimental data from murine models of KD vasculitis and transcriptomics data generated from whole blood of KD patients indicate the involvement of the NLRP3 inflammasome and interleukin-1 (IL-1) signaling in KD pathogenesis. MicroRNA-223 (miR-223) is a negative regulator of NLRP3 activity and IL-1β production, and its expression has been reported to be upregulated during acute human KD; however, the specific role of miR-223 during KD vasculitis remains unknown. Here, using the Lactobacillus casei cell wall extract (LCWE) murine model of KD vasculitis, we demonstrate increased miR-223 expression in LCWE-induced cardiovascular lesions. Compared with control WT mice, LCWE-injected miR-223-deficient mice (miR223−/y) developed more severe coronary arteritis and aortitis, as well as more pronounced abdominal aorta aneurysms and dilations. The enhanced cardiovascular lesions and KD vasculitis observed in LCWE-injected miR223−/y mice correlated with increased NLRP3 inflammasome activity and elevated IL-1β production, indicating that miR-223 limits cardiovascular lesion development by downmodulating NLRP3 inflammasome activity. Collectively, our data reveal a previously unappreciated role of miR-223 in regulating innate immune responses and in limiting KD vasculitis and its cardiovascular lesions by constraining the NLRP3 inflammasome and the IL-1β pathway. These data also suggest that miR-223 expression may be used as a marker for KD vasculitis pathogenesis and provide a novel therapeutic target.

In the process of sepsis, activated platelets shed microvesicles containing microRNAs (miRNAs), which can be internalized by distinct recipient cells in circulation, consequently eliciting a potent capability to regulate their cellular functions in different diseases. In the present study, activated human platelets transferring miR-223 into endothelial cells via platelet-derived microparticles (PMPs) was investigated in vitro during septic conditions with a proposed mechanism involving in downregulation of the enhanced expression of intercellular adhesion molecule-1 (ICAM-1). The uptake of PMPs encasing miR-223 and the adhesion of peripheral blood mononuclear cells (PBMCs) on human coronary artery endothelial cells (HCAECs) were observed by immunofluorescence microscopy upon co-culture with PMPs isolated from sepsis or control plasma. The expression of miR-223-3p and its gene target ICAM1 in HCAECs were quantified by RT-qPCR and ELISA after the cells were incubated with septic or control PMPs, whose levels were induced with thrombin-receptor activating peptide (TRAP). Leukocyte-depleted platelets (LDPs) from septic patients showed a decreased miR-223 level, while septic plasma and PMPs revealed an elevated miRNA level compared to control samples. Similarly, TRAP-activated LDPs demonstrated a reduced intracellular miR-223 expression, while increased levels in the supernatant and PMP isolates were observed vs. untreated samples. Furthermore, TNF-α alone resulted in decreased miR-223 and elevated ICAM1 levels in HCAECs, while PMPs raised the miRNA level that was associated with downregulated ICAM1 expression at both mRNA and protein levels under TNF-α treatment. Importantly, miR-223 was turned out not to be newly synthesized as shown in unchanged pre-miR-223 level, and mature miR-223 expression was also elevated in the presence of PMPs in HCAECs after transfection with Dicer1 siRNA. In addition, septic PMPs containing miR-223 decreased ICAM1 with a reduction of PBMC binding to HCAECs. In conclusion, septic platelets released PMPs carrying functional miR-223 lower ICAM1 expression in endothelial cells, which may be a protective role against excessive sepsis-induced vascular inflammation.

The data on studies of the expression of microRNA regulatory molecules miR-155 and miR-223 in bone marrow and peripheral blood cells in patients with chronic lymphoid leukemia (CLL) at various stages of the disease progression are presented. The microRNAs were determined by the method of highly sensitive PCR in various peripheral blood substrates (plasma, lymphocytes, extracellular vesicles) and bone marrow in patients with CLL. The expression rates correlated with clinical data and the effectiveness of standard therapy. It is noted that an increase in the miR-155 expression and decrease in the miR-223 expression are associated with poor prognosis, resistance to therapy and high rate of the tumors progression of chronic lymphocytic leukemia. MiR-155 overexpression in all studied blood substrates (plasma, lymphocytes, extracellular vesicles) and bone marrow and decrease in miR-223 expression in lymphocytes, bone marrow and plasma were significantly associated with the stages of chronic lymphocytic leukemia according to J. Binet. The patients with CLL with the miR-223 low level in the bone marrow, extracellular vesicles and plasma and the miR-155 high level before treatment initiation did not achieve an optimal response to the therapy.

As a promising new target, miR-233 may regulate oxidative stress by targeting keap1-Nrf2 system to affect the pathological process of liver injury in T2DM. Ellagic acid (EA) is versatile for protecting oxidative stress damage and metabolic disorders. In the present study, we investigated the effect of EA on oxidative stress and insulin resistance in high glucose-induced T2DM HepG2 cells and examined the role of miR-223/keap1-Nrf2 pathway in system. HepG2 cells were incubated in 30 mM of glucose, with or without EA (15 and 30 μM) or metformin (Met, 150 μM) for 12 h. Glucose consumption, phosphorylation of IRS1, Akt and ERK under insulin stimulation, ROS and O2·− production, MDA level, SOD activity and miR-223 expression, as well as protein levels of keap1, Nrf2, HO-1, SOD1 and SOD2 were analyzed. Furthermore, dual luciferase reporter assay, miR-223 mimic and inhibitor were implemented in cellular studies to explore the possible mechanism. EA upregulated glucose consumption, IRS1, Akt and ERK phosphorylation under insulin stimulation, reduced ROS and O2·− production and MDA level, and increased SOD activity in high glucose-exposed HepG2 cells. In addition, EA elevated miR-223 expression level, downregulated mRNA and protein levels of keap1, and upregulated Nrf2, HO-1, SOD1 and SOD2 protein levels in this cell model. What’s more, dual luciferase reporter assay, miR-223 mimic and inhibitor transfection confirmed that EA activated keap1-Nrf2 system via elevating miR-223. The miR-223, a negative regulator of keap1, represents an attractive therapeutic target in hepatic injury in T2DM. EA ameliorates oxidative stress and insulin resistance via miR-223-mediated keap1-Nrf2 activation in high glucose-induced T2DM HepG2 cells.

Abstract Background How exosomal microRNAs (miRNAs) derived from macrophages contribute to the development of drug resistance in the context of the hypoxic tumor microenvironment in epithelial ovarian cancer (EOC) remains poorly understood. Methods The miRNA levels were detected by qRT-PCR. Protein levels of HIF-1α, CD163 and PTEN-PI3K/AKT pathway were assessed by Western blot (WB) and Immunohistochemistry (IHC). Exosomes were isolated, and then confirmed by Transmission electron microscopy (TEM), Nanoparticle Tracking Analysis (NTA) and WB. Internalization of macrophages-secreted exosomes in EOC cells was detected by Confocal microscope. Subsequently, Dual-luciferase reporter assay verified PTEN was the target of miR-223. Gain- and loss-of-function experiments, rescue experiments, and SKOV3 xenograft models were performed to uncover the underlying mechanisms of miR-223 and PTEN-PI3K/AKT pathway, as well as the exosomal miR-223 in inducing multidrug resistance in vitro and in vivo. Results Here, we showed hypoxic EOC cells triggered macrophages recruitment and induced macrophages into a tumor-associated macrophage (TAM)-like phenotype; exosomes derived from hypoxic macrophages enhanced the malignant phenotype of EOC cells, miR-223 was enriched in exosomes released from macrophages under hypoxia, which could be transferred to the co-cultivated EOC cells, accompanied by enhanced drug resistant of EOC cells. Besides, results from a functional assay revealed that exosomal miR-223 derived from macrophages promoted the drug resistance of EOC cells via the PTEN-PI3K/AKT pathway both in vivo and in vitro. Furthermore, patients with high HIF-1a expression had statistically higher CD163+ cell infiltration and intertumoral levels of miR-223. Finally, circulating exosomal miR-223 levels were closely related to the recurrence of EOC. Conclusions These data indicate a unique role of exosomal miR-223 in the cross-talk between macrophages and EOC cells in chemotherapy resistance, through a novel exosomal miR-223/PTEN-PI3K/AKT signaling pathway.