MiR-155-5p Differentially Regulates IL-13Rα1 and IL-13Rα2 Expression and Signalling, Driving Abnormal Lung Epithelial Cell Phenotype in Severe Asthma.

MiR-155-5p is known to increase in innate and adaptive immune cells in response to IL-13 and is associated with asthma severity. However, little is known about its role in airway structural cells. BECs isolated from healthy donors and severe asthma patients were stimulated with IL-13. MiR-155-5p expression and release were measured by RT-PCR in BECs and in their derived exosomes. Modulation of miR-155-5p in BECs was performed using transfection of miR-155-5p inhibitor and mimic. IL-13Rα1, IL-13Rα2, MUC5AC, IL-8 and Eotaxin-1 expression were measured by RT-PCR and western blot. BECs repair process was assessed by wound healing assay. IL-13Rα1 and IL-13Rα2 expression and downstream pathways were evaluated by western blot. Dual Luciferase assay was used to determine miR-155-5p target genes associated to IL-13 receptors signaling. BECs from severe asthma showed an increased expression and exosomal release of miR-155-5p at baseline that was amplified by IL-13 stimulation. BECs from asthmatics expressed more IL-13Rα1 and less IL-13Rα2 than healthy donors and IL-13Rα1 but not IL-13Rα2 induced miR-155-5p expression under IL-13 stimulation. MiR-155-5p overexpression favored MUC5AC, IL-8 and Eotaxin-1 through IL-13Rα1/SOCS1/STAT6 pathway to the detriment of a delayed repair process with a downregulated IL-13Rα2/MAPK14/c-Jun/c-Fos signaling. Dual Luciferase assay confirmed that miR-155-5p modulates both IL-13 receptors pathways by directly targeting SOCS1, c-Fos and MAPK14. MiR-155-5p is overexpressed in severe asthma BECs and regulates IL-13Rα1 and IL-13Rα2 expression and signaling, favoring expression of mucin and eosinophils related genes to detriment of airway repair. These results show that miR-155-5p may contribute to airway epithelial cell dysfunction in severe asthma.

Exploring of miR-155-5p, miR-181b-5p, and miR-454-3p Expressions in Circulating Cell-Free RNA: Insights from Peripheral Blood of Uveal Malignant Melanoma Patients.

The identification of novel non-invasive biomarkers is imperative for the early diagnosis and monitoring of malignant melanoma. The objective of this study is to examine the expression levels of miR-155-5p, miR-181b-5p, and miR-454-3p in circulating cell-free RNA obtained from plasma samples of the 72 uveal malignant melanoma patients and to compare these levels with those of 72 healthy controls. The analysis showed that the expression level of the miR-181b-5p has increased 9.25 fold, and expression level of miR-155-5p has increased 6.67 fold, and miR-454-3p expression level has increased 4.14 fold in the patient group compared with the levels in the healthy control group (p = 0.005). It was found that the high expression levels of the three miRNAs were statistically significant in patients compared with in the healthy control group. The statistical evaluations between miRNA expression levels and clinical data showed that miR-155-5p had significant association with radiation therapy (p = 0.040), and miR-454-3p showed a significant association with smoking and alcohol use respectively (p = 0.009, and p = 0.026). The significantly elevated expression levels of miR-181b-5p, miR-155-5p, and miR-454-3p in the circulating cell-free RNA of plasma from uveal melanoma patients, in comparison to those in the healthy control group, suggest the potential usefulness of these biomarkers for both early diagnosis and disease monitoring. However, more extensive and future studies are needed to use these molecules in early diagnosis and disease monitoring.

circ_0000592 facilitates the progression of esophageal squamous cell carcinoma via miR-155-5p/FZD5 axis.

Esophageal squamous cell carcinoma (ESCC) is one of the gastrointestinal malignancies with high prevalence and poor prognosis. Previous reports suggested that circular ribose nucleic acids might exert regulatory functions in ESCC. This study aims to explore the role of circ_0000592 in ESCC progression, providing novel insights into the diagnosis and therapeutic avenues for ESCC. The GSE131969 data set was utilized to assess circ_0000592 expression in ESCC. The validation was performed in the tumorous tissues of ESCC patients (n = 80) and human-immortalized ESCC cell lines. The correlation between circ_0000592 expression and prognosis was analyzed. The impact of circ_0000592 on ESCC cell activity was evaluated through downregulating circ_0000592, as well as encompassing cell viability, migration, and invasion abilities. The downstream pathway of circ_0000592 was explored by binding site prediction from the TargetScan database, followed by in vitro and in vivo experiments. An in vivo xenograft tumor model was established to highlight the role of circ_0000592 in ESCC. Patients with ESCC exhibited higher circ_0000592 expression levels compared to noncancerous patients, which were associated with reduced survival time, higher TNM stage, and increased lymph node metastasis. The circ_0000592 downregulation suppressed cell viability, migration, and invasion abilities in vitro. Mechanistically, circ_0000592 countered the inhibitory effects on the target gene Frizzled 5 (FZD5) through interactions with miR-155-5p. The overexpression of miR-155-5p curtailed the luciferase activity of circ_0000592 in ESCC cells, inhibiting downstream molecule FZD5 protein expression and subsequently mitigating the detrimental consequences of escalated circ_0000592 expression in ESCC cells. Consistently, circ_0000592 downregulation curbed proliferation and metastasis of ESCC tumors in vivo. In summary, circ_0000592 promoted the progress of ESCC by counteracting the inhibitory impact on FZD5 through its interaction with miR-155-5p. Together, our findings highlighted circ_0000592 as a prospective therapeutic target for ESCC.

Alternations in miR-155 and miR-200 serum levels can serve as biomarkers for COVID-19 in the post-mass vaccination era.

BACKGROUND: Mass vaccination and natural immunity reduced the severity of COVID-19 cases. SARS-CoV-2 ongoing genome variations imply the use of confirmatory serologic biomarkers besides PCR for reliable diagnosis. MicroRNA molecules are intrinsic components of the innate immune system. The expression of miR155-5p and miR200c-3p was previously correlated with SARS-CoV-2 pathogenesis. This case-control study was conducted during the third peak of the COVID-19 pandemic in Egypt and aimed to calculate the accuracy of miR155-5p and miR200c-3p as biomarkers for COVID-19. METHODS AND RESULTS: Thirty out of 400 COVID-19 patients at a main University hospital in Alexandria were included in the study along with 20 age-matched healthy controls. Plasma samples were collected for total and differential CBC. Relative quantitation of miR155-5p and miR200c-3p expression from WBCs was done by RT-qPCR. The expression of miR155-5p and miR200c-3p was positively correlated and was significantly downregulated in COVID-19 patients compared to the healthy control group (p ˂ 0.005). Both miR155-5p and miR200c-3p were of 76% and 74% accuracy as diagnostic biomarkers of COVID-19, respectively. Regarding the differentiation between mild and moderate cases, their accuracy was 80% and 70%, respectively. CONCLUSIONS: miR155-5p and miR200c-3p expression can be used to confirm the diagnosis of COVID-19 and discriminate between mild and moderate cases, with a moderate degree of accuracy.

Analysis of the differential expression of serum miR-21-5p, miR-135-5p, and miR-155-5p by Bifidobacterium triplex viable capsules during the perioperative stage of colorectal cancer.

OBJECTIVE: In this study, we investigated the impact of perioperative administration of Bifidobacterium triplex viable capsules on the serum levels of circulating miR-21-5p, miR-135-5p, and miR-155-5p in patients with colorectal cancer (CRC). The purpose of this study is to provide a foundation for future research on the use of Bifidobacterium triplex viable capsules to enhance postoperative recovery in patients with CRC. METHODS: A total of 60 patients with primary CRC admitted to the Department of General Surgery at Shanxi Bethune Hospital between June 2020 and December 2020 were selected and randomly divided into two groups: 20 cases in the control group and 40 cases in the experimental group. The experimental group was administered oral Bifidobacterium triplex viable capsules during the perioperative period, while the control group was administered oral placebo. Before and after the perioperative period, the expression levels of miR-21-5p, miR-135-5p, and miR-155-5p were compared in the serum of both groups of patients. Furthermore, we established the prognostic value of these three miRNAs in CRC patients. RESULTS: After surgery, the expression levels of miR-21-5p, miR-135-5p, and miR-155-5p decreased in both groups of patients (P < 0.05). Significantly greater differences were observed between miR-21-5p and miR-135-5p (P < 0.001). Expression levels of serum miR-21-5p (P = 0.020) and miR-135-5p (P = 0.023) decreased significantly more in the experimental group than in the control group. The levels of the above three miRNAs after surgery did not correlate with 3-year OS (HR = 4.21; 95% CI 0.37-47.48; log-rank P = 0.20) or 3-year DFS (HR = 1.57; 95% CI 0.32-7.66; log-rank P = 0.55) in two groups. CONCLUSION: Radical surgery reduces the levels of serum miR-21-5p, miR-135-5p, and miR-155-5p expression in patients with CRC. The use of Bifidobacterium triplex viable capsules assists in achieving quicker perioperative recovery from radical surgery in CRC patients, and this underlying mechanism may be associated with the regulation of serum miR-21-5p, miR-135-5p, and miR-155-5p expression levels.

ORF3a of SARS-CoV-2 modulates PI3K/AKT signaling in human lung epithelial cells via hsa-miR-155-5p.

SARS-CoV-2 infection results in cytokine burst, leading to proinflammatory responses in lungs of COVID-19 patients. SARS-CoV-2 ORF3a triggers the generation of proinflammatory cytokines. However, the underlying mechanism of dysregulation of proinflammatory responses is not well understood. We studied the role of microRNA in the generation of proinflammatory responses as a bystander effect of SARS-CoV-2 ORF3a in human lung epithelial cells. We observed upregulation of hsa-miR-155-5p in SARS-CoV-2 ORF3a transfected human lung epithelial cells, which led to the reduced expression of SHIP1. This resulted in phosphorylation of AKT and NF-κB, which further led to the increased expression of the proinflammatory cytokines IL-6 and TNF-α. Additionally, overexpression and knockdown studies of hsa-miR-155-5p were performed to confirm the role of hsa-miR-155-5p in the regulation of the SHIP1. We demonstrated that hsa-miR-155-5p modulates the proinflammatory response by activating the PI3K/AKT pathway through the inhibition of SHIP1 in SARS-CoV-2 ORF3a transfected human lung epithelial cells.

MicroRNA-155 mediates multiple gene regulations pertinent to the role of human adipose-derived mesenchymal stem cells in skin regeneration.

Introduction: The role of Adipose-derived mesenchymal stem cells (AD-MSCs) in skin wound healing remains to be fully characterized. This study aims to evaluate the regenerative potential of autologous AD-MSCs in a non-healing porcine wound model, in addition to elucidate key miRNA-mediated epigenetic regulations that underlie the regenerative potential of AD-MSCs in wounds. Methods: The regenerative potential of autologous AD-MSCs was evaluated in porcine model using histopathology and spatial frequency domain imaging. Then, the correlations between miRNAs and proteins of AD-MSCs were evaluated using an integration analysis in primary human AD-MSCs in comparison to primary human keratinocytes. Transfection study of AD-MSCs was conducted to validate the bioinformatics data. Results: Autologous porcine AD-MSCs improved wound epithelialization and skin properties in comparison to control wounds. We identified 26 proteins upregulated in human AD-MSCs, including growth and angiogenic factors, chemokines and inflammatory cytokines. Pathway enrichment analysis highlighted cell signalling-associated pathways and immunomodulatory pathways. miRNA-target modelling revealed regulations related to genes encoding for 16 upregulated proteins. miR-155-5p was predicted to regulate Fibroblast growth factor 2 and 7, C-C motif chemokine ligand 2 and Vascular cell adhesion molecule 1. Transfecting human AD-MSCs cell line with anti-miR-155 showed transient gene silencing of the four proteins at 24 h post-transfection. Discussion: This study proposes a positive miR-155-mediated gene regulation of key factors involved in wound healing. The study represents a promising approach for miRNA-based and cell-free regenerative treatment for difficult-to-heal wounds. The therapeutic potential of miR-155 and its identified targets should be further explored in-vivo.

lncRNA NORAD alleviates dysfunction of renal proximal tubular epithelial cells during the sepsis-associated acute kidney injury by modulating the miR-155-5p-PDK1 axis.

The sepsis-associated acute kidney injury (Sa-AKI) is closely related to high mortality rates worldwide. Injury to the renal proximal tubular epithelial cells (RPTECs), caused by pathological conditions, is a major cause of acute kidney injury (AKI). The lncRNA NORAD has been reported to be positively associated with kidney cancers. However, the biological roles and underlying mechanisms of NORAD in RPTECs during AKI are still unclear. In this study, we found that NORAD was significantly downregulated in RPTECs from AKI tissues. Overexpression of NORAD alleviated RPTECs injury induced by lipopolysaccharide (LPS). Additionally, glucose metabolism was significantly impaired during AKI, and LPS treatment inhibited glucose metabolism in RPTECs. We demonstrated that NORAD rescued the LPS-induced inhibition of glucose metabolism in RPTECs. Furthermore, miRNA-155-5p was significantly upregulated in RPTECs from AKI. Through bioinformatics analysis, RNA pull-down, RNA IP, and luciferase assays, we showed that NORAD directly associated with miR-155-5p to downregulate its expression. Moreover, overexpression of miR-155-5p inhibited glucose metabolism by directly targeting the 3'UTR of the glucose metabolism enzyme, pyruvate dehydrogenase kinase 1 (PDK1). Finally, rescue experiments validated that NORAD's protective effect on RPTECs injury was mediated through modulation of the miR-155-5p-PDK1-glucose metabolism pathway. In summary, these results reveal that lncRNA NORAD can alleviate RPTECs dysfunction by targeting the miR-155-5p-PDK1 axis, suggesting that NORAD has the potential to contribute to the development of therapeutic approaches against Sa-AKI.

The PIEZO1/miR-155-5p/GDF6/SMAD2/3 signaling axis is involved in inducing the occurrence and progression of osteoarthritis under excessive mechanical stress.

OBJECTIVE: To elucidate the molecular mechanism of overloading-induced osteoarthritis (OA) and to find a novel therapeutic target. METHODS: We utilized human cartilage specimens, mouse chondrocytes, a destabilization of the medial meniscus (DMM) mouse model, and a mouse hindlimb weight-bearing model to validate the role of overloading on chondrocyte senescence and OA development. Then, we observed the effect of PIEZO1-miR-155-5p-GDF6-SMAD2/3 signaling axis on the preservation of joint metabolic homeostasis under overloading in vivo, in vitro and ex vivo by qPCR, Western blot, enzyme-linked immunosorbent assay (ELISA), immunohistochemistry, immunofluorescence, SA-ß-gal staining, CCK8 assay, et al. Finally, we verified the therapeutic effects of intra-articular injection of miR-155-5p inhibitor or recombinant GDF6 on the murine overloading-induced OA models. RESULTS: Chondrocytes sensesed the mechanical overloading through PIEZO1 and up-regulated miR-155-5p expression. MiR-155-5p mimics could copy the effects of overloading-induced chondrocyte senescence and OA. Additionally, miR-155-5p could suppress the mRNA expression of Gdf6-Smad2/3 in various tissues within the joint. Overloading could disrupt joint metabolic homeostasis by downregulating the expression of anabolism indicators and upregulating the expression of catabolism indicators in the chondrocytes and synoviocytes, while miR-155-5p inhibition or GDF6 supplementation could exert an antagonistic effect by preserving the joint homeostasis. Finally, in the in vivo overloading models, intra-articular injection of miR-155-5p inhibitor or recombinant GDF6 could significantly mitigate the severity of impending OA and lessened the progression of existing OA. CONCLUSION: GDF6 overexpression or miR-155-5p inhibition could attenuate overloading-induced chondrocyte senescence and OA through the PIEZO1-miR-155-5p-GDF6-SMAD2/3 signaling pathway. Our study provides a new therapeutic target for the treatment of overloading-induced OA.

Dapagliflozin prevents kidney podocytes pyroptosis via miR-155-5p/HO-1/NLRP3 axis modulation.

Diabetic nephropathy (DN) is a significant clinical microvascular complication associated with diabetes mellitus (DM), and end-stage diabetes giving rise to kidney failure is developing into the major etiological factor of chronic kidney failure. Dapagliflozin is reported to limit podocyte damage in DM, which has proven to protect against renal failure. Mounting evidence has demonstrated that pyroptosis is associated with DM progression. Nevertheless, whether pyroptosis causes DN and the underlying molecular pathways remain obscure. In this study, we aimed to explore the antipyroptotic attributes of dapagliflozin and elucidate the underlying mechanisms of kidney damage in diabetes. In vivo, experiments were conducted in streptozotocin (STZ)-induced type 2 diabetic mice, which were administered dapagliflozin via gavage for 6 weeks. Subsequently, the specific organizational characteristics and expression of pyroptosis-related genes were evaluated. Intragastric dapagliflozin administration markedly reduced renal tissue injury. Meanwhile, dapagliflozin also attenuated the expression level of pyroptosis associated genes, including ASC, cleaved Caspase-1, GSDMD N-termini, NLRP3, IL-18, and IL-1ß in renal tissue of dapagliflozin-treated animals. Similar antipyroptotic effects were observed in palmitic acid (PA)-treated mouse podocytes. We also found that heme oxygenase 1 (HO-1) enhanced the protection of mouse podocyte clone 5 cells (MPC5). Moreover, miR-155-5p inhibition increased pyroptosis in PA-treated MPC5 cells, suggesting that miR-155-5p acts as an endogenous stimulator that increases HO-1 expression and reduces pyroptosis. Hence, our findings imply that dapagliflozin inhibits podocyte pyroptosis via the miR-155-5p/HO-1/NLRP3 axis in DM. Furthermore, dapagliflozin substitution may be regarded as an effective strategy for preventing pyroptosis in the kidney, including a therapeutic option for treating pyroptosis-related DN.

Exploring the Impact of TLR-2 Signaling on miRNA Dysregulation in Intervertebral Disc Degeneration.

Toll-like receptors (TLRs) are key mediators of inflammation in intervertebral disc (IVD) degeneration. TLR-2 activation contributes to the degenerative process by increasing the expression of extracellular matrix-degrading enzymes, pro-inflammatory cytokines, and neurotrophins. As potent post-transcriptional regulators, microRNAs can modulate intracellular mechanisms, and their dysregulation is known to contribute to numerous pathologies. This study aims to investigate the impact of TLR-2 signaling on miRNA dysregulation in the context of IVD degeneration. Small-RNA sequencing of degenerated IVD cells shows the dysregulation of ten miRNAs following TLR-2 activation by PAM2CSK4. The miR-155-5p is most significantly upregulated in degenerated and non-degenerated annulus fibrosus and nucleus pulposus cells. Sequence-based target and pathway prediction shows the involvement of miR-155-5p in inflammation- and cell fate-related pathways and TLR-2-induced miR-155-5p expression leads to the downregulation of its target c-FOS. Furthermore, changes specific to the activation of TLR-2 through fragmented fibronectin are seen in miR-484 and miR-487. Lastly, miR-100-3p, miR-320b, and miR-181a-3p expression exhibit degeneration-dependent changes. These results show that TLR-2 signaling leads to the dysregulation of miRNAs in IVD cells as well as their possible downstream effects on inflammation and degeneration. The identified miRNAs provide important opportunities as potential therapeutic targets for IVD degeneration and low back pain.

The effect of c-Fos on the prognosis, proliferation, and invasion of oral squamous cell carcinoma.

OBJECTIVE: This study aimed to determine the role of c-Fos in growth and invasion of oral squamous cell carcinoma (OSCC). METHODS: Immunohistochemistry was used to assess c-Fos expression in 94 OSCC tissues and 30 adjacent normal tissues, the correlation between c-Fos expression and clinicopathological characteristics was examined, and Kaplan-Meier and Cox analysis were used to investigate the role of c-Fos in predicting the prognosis of OSCC patients. The effects of c-Fos on the growth and invasion of OSCC were disclosed by overexpression and knockdown of c-Fos. Furthermore, based on bioinformatics prediction, the effect of miR-155-5p on c-Fos expression was examined, and dual-luciferase reporter assay system was used to determine whether miR-155-5p regulated the transcriptional activity of c-Fos in OSCC. RESULTS: c-Fos was markedly increased in OSCC tissues and cells. c-Fos upregulation indicates a poor prognosis in OSCC patients, and c-Fos promotes cell proliferation, migration, and invasion in OSCC. miR-155-5p could regulate the expression and the transcriptional activity of c-Fos by directly targeting the c-Fos 3'-UTR. CONCLUSION: This study demonstrated that c-Fos contributed to the progression of OSCC and may act as a potential target for OSCC therapy, and a potential prognostic biomarker of OSCC.

MiR-155-5p improves the insulin sensitivity of trophoblasts by targeting CEBPB in gestational diabetes mellitus.

INTRODUCTION: Gestational diabetes mellitus (GDM) is a prevalent pregnancy complication featuring impaired insulin sensitivity. MiR-155-5p is associated with various metabolic diseases. However, its specific role in GDM remains unclear. CCAAT enhancer binding protein beta (CEBPB), a critical role in regulating glucolipid metabolism, has been identified as a potential target of miR-155-5p. This study aims to investigate the impact of miR-155-5p and CEBPB on insulin sensitivity of trophoblasts in GDM. METHODS: Placental tissues were obtained from GDM and normal pregnant women; miR-155-5p expression was then evaluated by RT‒qPCR and CEBPB expression by western blot and immunohistochemical staining. To investigate the impact of miR-155-5p on insulin sensitivity and CEBPB expression, HTR-8/SVneo cells were transfected with either miR-155-5p mimic or inhibitor under basal and insulin-stimulated conditions. Cellular glucose uptake consumption was quantified using a glucose assay kit. Furthermore, the targeting relationship between miR-155-5p and CEBPB was validated using a dual luciferase reporter assay. RESULTS: Reduced miR-155-5p expression and elevated CEBPB expression were observed in GDM placentas and high glucose treated HTR8/SVneo cells. The overexpression of miR-155-5p significantly enhanced insulin signaling and glucose uptake in trophoblasts. Conversely, inhibiting miR-155-5p induced the opposite effects. Additionally, CEBPB was directly targeted and negatively regulated by miR-155-5p in HTR8/SVneo cells. Silencing CEBPB effectively restored the inhibitory effect of miR-155-5p downregulation on insulin sensitivity in trophoblasts. DISCUSSION: These findings suggest that miR-155-5p could enhance insulin sensitivity in trophoblasts by targeting CEBPB, highlighting the potential of miR-155-5p as a therapeutic target for improving the intrauterine hyperglycemic environment in GDM.

SOCS5, targeted by miR-155-5p, plays a negative regulatory role in pulmonary hypertension through inhibiting JAK2/STAT3 signaling pathway.

Pulmonary hypertension (PH) is a chronic pulmonary vascular disease and causes massive deaths. Here, we intended to investigate the function and mechanism of SOCS5 in PH. We engineered a hypoxia-induced PH model in mice. HE staining were implemented to evaluate pathological alterations in the lung tissues. The potential mechanism of SOCS5 in regulating hypoxia-induced pulmonary artery smooth muscle cell (PASMC) function was explored in vitro. RT-qPCR and western blot revealed that the level of SOCS5 was decreased both in PH mice and hypoxia-induced HPASMCs. Functional assays were performed for confirming the role of SOCS5 in modulating the cell phenotype and JAK2/STAT3 pathway in HPASMCs. Results revealed that overexpression of SOCS5 suppressed proliferation, migration and contraction of HPASMCs and negatively regulated the JAK2/STAT3 signaling pathway in HPASMCs under hypoxia in vitro, while knockdown of SOCS5 accelerated it. As evidenced by mechanism studies, SOCS5 was targeted and regulated by miR-155-5p, hence affecting on HPASMC proliferation, migration and contraction. These outcomes indicated that the decreased level of SOCS5 in hypoxia-induced HPASMCs promoted the cell proliferation, cell migration, and cell contraction through activating JAK2/STAT3 signaling pathway. Moreover, SOCS5 was targeted by miR-155-5p. All in all, our work hinted that miR-155-5p/SOCS5/JAK2/STAT3 axis played a crucial part in PH.

Gestational Hypoxia Impaired Endothelial Nitric Oxide Synthesis Via miR-155-5p/NADPH Oxidase/Reactive Oxygen Species Axis in Male Offspring Vessels.

BACKGROUND: Nitric oxide (NO) is the most important vasodilator secreted by vascular endothelial cells, and its abnormal synthesis is involved in the development of cardiovascular disease. The prenatal period is a critical time for development and largely determines lifelong vascular health in offspring. Given the high incidence and severity of gestational hypoxia in mid-late pregnancy, it is urgent to further explore whether it affects the long-term synthesis of NO in offspring vascular endothelial cells. METHODS AND RESULTS: Pregnant Sprague-Dawley rats were housed in a normoxic or hypoxic (10.5% O2) chamber from gestation days 10 to 20. The thoracic aortas of fetal and adult male offspring were isolated for experiments. Gestational hypoxia significantly reduces the NO-dependent vasodilation mediated by acetylcholine in both the fetal and adult offspring thoracic aorta rings. Meanwhile, acetylcholine-induced NO synthesis is impaired in vascular endothelial cells from hypoxic offspring thoracic aortas. We demonstrate that gestational hypoxic offspring exhibit a reduced endothelial NO synthesis capacity, primarily due to increased expression of NADPH oxidase 2 and enhanced reactive oxygen species. Additionally, gestational hypoxic offspring show elevated levels of miR-155-5p in vascular endothelial cells, which is associated with increased expression of NADPH oxidase 2 and reactive oxygen species generation, as well as impaired NO synthesis. CONCLUSIONS: The present study is the first to demonstrate that gestational hypoxia impairs endothelial NO synthesis via the miR-155-5p/NADPH oxidase 2/reactive oxygen species axis in offspring vessels. These novel findings indicate that the detrimental effects of gestational hypoxia on fetal vascular function can persist into adulthood, providing new insights into the development of vascular diseases.

miR-155-5p promotes hepatic steatosis via PICALM-mediated autophagy in aging hepatocytes.

BACKGROUND: Hepatic steatosis, a lipid disorder characterized by the accumulation of intrahepatic fat, is more prevalent in the elderly population. This study investigates the role of miR-155-5p in the autophagy dysregulation of aging hepatic steatosis. METHODS: We established an aging mouse model in vivo and a hepatocellular senescence model induced by low serum and palmitic acid in vitro. The fluctuations of microRNAs were derived from RNA-seq data and confirmed by qPCR in 4- and 18-month-old mouse liver tissues. Hematoxylin-eosin (H&E) staining observed pathological changes. Markers of senescence, autophagy, and lipolysis genes were analyzed using Western blot and qPCR. Bioinformatics analysis predicted miR-155-5p's target gene PICALM, confirmed by dual luciferase reporter assay and transfection of miR-155-5p mimic/inhibitor into senescent hepatocytes. RESULTS: Senescent markers (p21, p16, and p-P53) and miR-155-5p were up-regulated in aging liver tissues and senescent hepatocytes. Bioinformatics analysis identified PICALM as a target gene of miR-155-5p, a finding further supported by dual luciferase reporter assays. Inhibition of miR-155-5p reduced expression of senescent marker genes (p16, p21, p-P53), improved autophagy (evidenced by increased LC3B-II and ATG5, and decreased P62), and enhanced lipolysis (indicated by increased ATGL and p-HSL) in senescent hepatocytes. Oil red O staining confirmed that miR-155-5p inhibition significantly reduced lipid accumulation in these cells. CONCLUSIONS: This study suggests a potential new therapeutic approach for age-related hepatic steatosis through the inhibition of miR-155-5p to enhance autophagy.

Role of LINC00240 on T-helper 9 differentiation in allergic rhinitis through influencing DNMT1-dependent methylation of PU.1.

BACKGROUND: Allergic rhinitis (AR) is a common allergic disease with increasing prevalence globally. However, the molecular mechanism underlying AR pathogenesis remains largely undefined. METHODS: Peripheral blood and nasal mucosa samples obtained from patients with AR (n = 22), and ovalbumin-induced AR mouse model (n = 8 per group) were prepared for subsequent detection. qRT-PCR and western blot were used to detect the expression of LINC00240, miR-155-5p, PU.1 and other key molecules. ELISA assay and flow cytometry were employed to evaluate the secretion of IL-9 and T-helper 9 (Th9) cell ratio, respectively. Bioinformatics analysis, RNA immunoprecipitation (RIP), chromatin immunoprecipitation (ChIP) and luciferase reporter assays were employed to further elucidate the regulatory network of LINC00240/miR-155-5p/DNMT1. The methylation of PU.1 promoter was assessed by methylation-specific PCR (MSP). This signaling axis was further validated in the mouse model of AR. RESULTS: LINC00240 was downregulated, while miR-155-5p and PU.1 were upregulated in the peripheral blood and nasal mucosa of AR patients, as well as in AR mice. This was accompanied with the increased ratio of Th9 cells and elevated IL-9 secretion. Mechanistically, LINC00240 served as a miR-155-5p sponge, and DNMT1 was a target of miR-155-5p. In addition, DNMT1 mediated the methylation of PU.1 promoter. In vivo studies verified that LINC00240 mitigated AR progression, possibly via miR-155-5p/DNMT1/PU.1-dependent Th9 differentiation. CONCLUSION: The involvement of LINC00240 in AR pathogenesis is closely associated with Th9 differentiation through modulating DNMT1-dependent methylation of PU.1 by sponging miR-155-5p.

miR-155-5p/Bmal1 Modulates the Senescence and Osteogenic Differentiation of Mouse BMSCs through the Hippo Signaling Pathway.

BACKGROUND: The core clock gene brain and muscle ARNT like-1 (Bmal1) is involved in the regulation of bone tissue aging. However, current studies are mostly limited to the establishment of the association between Bmal1 and bone senescence, without in-depth exploration of its main upstream and downstream regulatory mechanisms. METHODS: The luciferase reporter assay, RT-qPCR and Western blotting were performed to detect the interaction between miR-155-5p and Bmal1. The effects of miR-155-5p and Bmal1 on the aging and osteogenic differentiation ability of mouse bone marrow mesenchymal stem cells (BMSCs) were investigated by cell counting kit-8 (CCK-8) assay, flow cytometry, ß-gal staining, alkaline phosphatase quantitative assay and alizarin red staining in vitro. The potential molecular mechanism was identified by ChIP-Seq, RNA-seq database analysis and immunofluorescence staining. RESULTS: The expression of Bmal1 declined with age, while the miR-155-5p was increased. miR-155-5p and Bmal1 repressed each other's expression, and miR-155-5p targeted the Bmal1. Besides, miR-155-5p inhibited the proliferation and osteogenic differentiation of BMSCs, promoted cell apoptosis and senescence, inhibited the expression and nuclear translocation of YAP and TAZ. However, Bmal1 facilitated the osteogenic differentiation and suppressed the aging of BMSCs, meanwhile inactivated the Hippo pathway. Moreover, YAP inhibitors abrogated the positive regulation of aging and osteogenic differentiation in BMSCs by miR-155-5p and Bmal1. CONCLUSION: In mouse BMSCs, miR-155-5p and Bmal1 regulated the aging and osteogenic differentiation ability of BMSCs mainly through the Hippo signaling pathway. Our findings provide new insights for the interventions in bone aging.

Exosomal miR-155-5p drives widespread macrophage M1 polarization in hypervirulent Klebsiella pneumoniae-induced acute lung injury via the MSK1/p38-MAPK axis.

BACKGROUND: Hypervirulent Klebsiella pneumoniae (hvKp) infection-induced sepsis-associated acute lung injury (ALI) has emerged as a significant clinical challenge. Increasing evidence suggests that activated inflammatory macrophages contribute to tissue damage in sepsis. However, the underlying causes of widespread macrophage activation remain unclear. METHODS: BALB/c mice were intravenously injected with inactivated hvKp (iHvKp) to observe lung tissue damage, inflammation, and M1 macrophage polarization. In vitro, activated RAW264.7 macrophage-derived exosomes (iHvKp-exo) were isolated and their role in ALI formation was investigated. RT-PCR was conducted to identify changes in exosomal miRNA. Bioinformatics analysis and dual-luciferase reporter assays were performed to validate MSK1 as a direct target of miR-155-5p. Further in vivo and in vitro experiments were conducted to explore the specific mechanisms involved. RESULTS: iHvKp successfully induced ALI in vivo and upregulated the expression of miR-155-5p. In vivo, injection of iHvKp-exo induced inflammatory tissue damage and macrophage M1 polarization. In vitro, iHvKp-exo was found to promote macrophage inflammatory response and M1 polarization through the activation of the p38-MAPK pathway. RT-PCR revealed exposure time-dependent increased levels of miR-155-5p in iHvKp-exo. Dual-luciferase reporter assays confirmed the functional role of miR-155-5p in mediating iHvKp-exo effects by targeting MSK1. Additionally, inhibition of miR-155-5p reduced M1 polarization of lung macrophages in vivo, resulting in decreased lung injury and inflammation induced by iHvKp-exo or iHvKp. CONCLUSIONS: The aforementioned results indicate that exosomal miR-155-5p drives widespread macrophage inflammation and M1 polarization in hvKp-induced ALI through the MSK1/p38-MAPK Axis.

Herbal medicine formula Huazhuo Tiaozhi granule ameliorates dyslipidaemia via regulating histone lactylation and miR-155-5p biogenesis.

BACKGROUND: Huazhuo Tiaozhi granule (HTG) is a herbal medicine formula widely used in clinical practice for hypolipidaemic effects. However, the molecular mechanisms underlying dyslipidaemia treatment have not been well elucidated. RESULTS: A significant reduction in the levels of total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) was observed in the serum of patients with dyslipidaemia after HTG treatment, without disruption in the levels of aspartate transaminase (AST), alanine transaminase (ALT), urea nitrogen (BUN), and creatinine (Cr). The dyslipidaemia rat model was induced by a high-fat diet and treated with Xuezhikang (0.14 g/kg/d) or HTG (9.33 g crude herb/kg/day) by gavage for 8 weeks. Body weight and liver index were markedly decreased in dyslipidaemic rats after treatment with Xuezhikang or HTG. HTG administration markedly ameliorated hyperlipidaemia by decreasing the levels of TC and LDL-C in serum and hepatic lipid accumulation. In vitro, lipid accumulation in LO2 and HepG2 cells was alleviated by serum treatment with HTG. High lactylation was observed in 198 proteins, including lactylation of histone H2B (K6), H4 (K80). Deep sequencing of microRNAs showed that miR-155-5p was significantly downregulated. CONCLUSIONS: This study demonstrates that HTG is an effective and safe formula for treating dyslipidaemia, which promotes lactylation in hepatocytes, and the retardation of miR-155-5p biogenesis.