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Background: Although low-level laser therapy (LLLT) is a widely used noninvasive treatment because of photobiomodulation effects, its application for xerostomia remained uncertain. Tight junctions (TJs), mainly composed of claudins, occludin, and ZO family members, are crucial structures that determine material transport through paracellular pathway in salivary gland epithelial cells. This work aimed to investigate whether LLLT affected salivary secretion through epithelial TJs. Methods: Transepithelial electrical resistance (TER) measurement and paracellular permeability assay were applied to evaluate paracellular permeability in submandibular gland (SMG)-C6 cells after irradiation with 540 nm green light. Immunofluorescence and western blot were used to detect the expression of TJ proteins. Quantitative phosphoproteomics were performed to explore possible intracellular signals. Results: We found that irradiation with 540 nm green light significantly decreased TER values while increased paracellular transport in SMG-C6 cells. 540 nm green light-induced redistribution of claudin-1, -3, and -4, but not occludin or ZO-1. Moreover, above phenomena were abolished by preincubation with capsazepine, an antagonist of transient receptor potential vanilloid subtype 1. Notably, irradiation with 540 nm green light on the skin covering the whole submandibular gland regions promoted salivary secretion and attenuated lymphocytic infiltration in 21-week-old non-obese diabetic mice (n = 5 per group), a xerostomia animal model for Sjögren's syndrome. Through in-depth bioinformatics analysis and expression verification, ERK1/2 and EphA2 served as potential canonical and noncanonical signals underlying 540 nm green light. Conclusions: Our findings uncovered the novel therapeutic effects of 540 nm green light on xerostomia through regulation on the expression and distribution of TJs.
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Terapia com Luz de Baixa Intensidade , Animais , Camundongos , Glândula Submandibular/efeitos da radiação , Glândula Submandibular/metabolismo , Saliva/metabolismo , Xerostomia/etiologia , Junções Íntimas/efeitos da radiação , Junções Íntimas/metabolismo , Ratos , Luz VerdeRESUMO
BACKGROUND AND AIM: Primary liver cancer, particularly hepatocellular carcinoma (HCC), represents a substantial global health challenge. Although immune checkpoint inhibitors are effective in HCC treatment, several patients still experience disease progression. Interleukin-1 (IL-1) regulates immunity and inflammation. We investigate the role of IL-1 in HCC development and progression and determine the potential therapeutic impact of gemcitabine in treating HCC. METHODS: Hydrodynamics-based transfection, employing the sleeping beauty transposase system, delivered surrogate tumor antigens, NRAS (NRAS proto-oncogene, GTPase), ShP53, and SB100 to C57BL/6 mice. A basic HCC mouse model was established. Pathogen-free animals were tested for serum and hepatotoxicity. The HCC prognosis was monitored using alanine aminotransferase and aspartate aminotransferase levels. Liver histology immunohistochemistry and mouse splenocyte/intra-hepatic immune cell flow cytometry were conducted. IL-1ß levels in human and mouse serum were assessed. RESULTS: Interleukin-1ß levels were elevated in patients with HCC compared with those in non-HCC controls. Hepatic IL-1ß levels were higher in HCC mouse models than those in non-HCC mice, suggesting localized hepatic inflammation. IL-1 receptor type 1 (IL-1R1) knockout (IL-1R1-/-) mice exhibited less severe HCC progression than that in wild-type mice, despite the high intra-hepatic IL-1ß concentration. IL-1R1-/- mice exhibited increased hepatic levels of myeloid-derived suppressor cells and regulatory T cells, which may exacerbate HCC. Gemcitabine significantly reduced the HCC tumor burden, improved liver conditions, and increased survival rates in HCC mouse models. Gemcitabine reduced the hepatic levels of myeloid-derived suppressor cells and regulatory T cells, potentially alleviating immune suppression in the liver. CONCLUSIONS: Targeting IL-1 or combining gemcitabine with immunotherapy is a promising approach for treating advanced-stage HCC.
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This study investigates sex-specific effects in a gain-of-function model to evaluate Nfil3 function in relation to high-fat diet (HFD)-induced metabolic dysfunction-associated steatotic liver disease (MASLD) and gut microbiota (GM)-induced alterations in the bile acid (BA) profile. MASLD is induced in both wild type and Nfil3-deficient (NKO) C57BL/6 J mice through an HFD. The hepatic immune response is evaluated using flow cytometry, revealing that NKO mice exhibit lower body weight, serum triglyceride (TG) levels, tissue injury, inflammation, and fat accumulation. The Nfil3 deletion reduces macrophage counts in fibrotic liver tissues, decreases proinflammatory gene and protein expression, and diminishes gut barrier function. Alpha and beta diversity analysis reveal increased GM alpha diversity across different sexes. The Nfil3 gene deletion modifies the BA profile, suggesting that negative feedback through the Nfil3-FXR-FGF15 axis facilitates BA recycling from the liver via enterohepatic circulation. Therefore, inhibiting Nfil3 in the liver offers a viable treatment approach for MASLD.
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Dieta Hiperlipídica , Camundongos Endogâmicos C57BL , Camundongos Knockout , Animais , Camundongos , Masculino , Feminino , Dieta Hiperlipídica/efeitos adversos , Microbioma Gastrointestinal , Ácidos e Sais Biliares/metabolismo , Fígado/metabolismo , Fígado/patologia , Hepatopatia Gordurosa não Alcoólica/metabolismo , Hepatopatia Gordurosa não Alcoólica/etiologia , Hepatopatia Gordurosa não Alcoólica/genética , Hepatopatia Gordurosa não Alcoólica/patologia , Modelos Animais de Doenças , Fígado Gorduroso/metabolismo , Fígado Gorduroso/genética , Fígado Gorduroso/patologia , Fígado Gorduroso/etiologia , Fatores de Transcrição de Zíper de Leucina BásicaRESUMO
BACKGROUND: Circular RNAs (circRNAs) are a novel kind of non-coding RNAs proved to play crucial roles in the development of multiple diabetic complications. However, their expression and function in diabetes mellitus (DM)-impaired salivary glands are unknown. RESULTS: By using microarray technology, 663 upregulated and 999 downregulated circRNAs companied with 813 upregulated and 525 downregulated mRNAs were identified in the parotid glands (PGs) of type2 DM mice under a 2-fold change and P < 0.05 cutoff criteria. Gene ontology (GO) and kyoto encyclopedia of genes and genomes (KEGG) analysis of upregulated mRNAs showed enrichments in immune system process and peroxisome proliferator-activated receptor (PPAR) signaling pathway. Infiltration of inflammatory cells and increased inflammatory cytokines were observed in diabetic PGs. Seven differently expressed circRNAs validated by qRT-PCR were selected for coding-non-coding gene co-expression (CNC) and competing endogenous RNA (ceRNA) networks analysis. PPAR signaling pathway was primarily enriched through analysis of circRNA-mRNA networks. Moreover, the circRNA-miRNA-mRNA networks highlighted an enrichment in the regulation of actin cytoskeleton. CONCLUSION: The inflammatory response is elevated in diabetic PGs. The selected seven distinct circRNAs may attribute to the injury of diabetic PG by modulating inflammatory response through PPAR signaling pathway and actin cytoskeleton in diabetic PGs.
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Diabetes Mellitus Tipo 2 , Perfilação da Expressão Gênica , Redes Reguladoras de Genes , Glândula Parótida , RNA Circular , Animais , RNA Circular/genética , Camundongos , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/metabolismo , Glândula Parótida/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores Ativados por Proliferador de Peroxissomo/metabolismo , Receptores Ativados por Proliferador de Peroxissomo/genética , Transcriptoma , Ontologia Genética , Masculino , Transdução de Sinais , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/metabolismoRESUMO
Pancreatic ductal adenocarcinoma (PDAC) has an extremely devastating nature with poor prognosis and increasing incidence, making it a formidable challenge in the global fight against cancer-related mortality. In this innovative preclinical investigation, the VCP/p97 inhibitor CB-5083 (CB), miR-142, a PD-L1 inhibitor, and immunoadjuvant resiquimod (R848; R) were synergistically encapsulated in solid lipid nanoparticles (SLNs). These SLNs demonstrated features of peptides targeting PD-L1, EGFR, and the endoplasmic reticulum, enclosed in a pH-responsive polyglutamic (PGA)-polyethylene glycol (PEG) shell. The homogeneous size and zeta potential of the nanoparticles were stable for 28 days at 4°C. The study substantiated the concurrent modulation of key pathways by the CB, miR, and R-loaded nanoformulation, prominently affecting VCP/Bip/ATF6, PD-L1/TGF-ß/IL-4, -8, -10, and TNF-α/IFN-γ/IL-1, -12/GM-CSF/CCL4 pathways. This adaptable nanoformulation induced durable antitumor immune responses and inhibited Panc-02 tumor growth by enhancing T cell infiltration, dendritic cell maturation, and suppressing Tregs and TAMs in mice bearing Panc-02 tumors. Furthermore, tissue distribution studies, biochemical assays, and histological examinations highlighted enhanced safety with PGA and peptide-modified nanoformulations for CB, miR, and/or R in Panc-02-bearing mice. This versatile nanoformulation allows tailored adjustment of the tumor microenvironment, thereby optimizing the localized delivery of combined therapy. These compelling findings advocate the potential development of a pH-sensitive, three-in-one PGA-PEG nanoformulation that combines a VCP inhibitor, a PD-L1 inhibitor, and an immunoadjuvant for cancer treatment via combinatorial chemo-immunotherapy.
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Imunoterapia , Nanopartículas , Neoplasias Pancreáticas , Microambiente Tumoral , Animais , Microambiente Tumoral/efeitos dos fármacos , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/imunologia , Neoplasias Pancreáticas/patologia , Humanos , Imunoterapia/métodos , Camundongos , Linhagem Celular Tumoral , Carcinoma Ductal Pancreático/tratamento farmacológico , Carcinoma Ductal Pancreático/imunologia , Carcinoma Ductal Pancreático/patologia , Antígeno B7-H1/antagonistas & inibidores , Sistemas de Liberação de Fármacos por Nanopartículas/química , Feminino , Polietilenoglicóis/química , Inibidores de Checkpoint Imunológico/farmacologia , LipossomosRESUMO
The development of alcohol-associated diseases is multifactorial, mechanism of which involves metabolic alteration, dysregulated immune response, and a perturbed intestinal host-environment interface. Emerging evidence has pinpointed the critical role of the intestinal host-microbiota interaction in alcohol-induced injuries, suggesting its contribution to disease initiation and development. To maintain homeostasis in the gut, the intestinal mucosa serves as the first-line defense against exogenous factors in the gastrointestinal tract, including dietary contents and the commensal microbiota. The gut-epithelial barrier comprises a physical barrier lined with a single layer of intestinal epithelial cells and a chemical barrier with mucus trapping host regulatory factors and gut commensal bacteria. In this article, we review recent studies pertaining to the disrupted gut-epithelial barrier upon alcohol exposure and examine how alcohol and its metabolism can affect the regulatory ability of intestinal epithelium.
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Etanol , Microbioma Gastrointestinal , Mucosa Intestinal , Mucosa Intestinal/microbiologia , Mucosa Intestinal/metabolismo , Humanos , Microbioma Gastrointestinal/efeitos dos fármacos , Microbioma Gastrointestinal/fisiologia , Animais , Homeostase , Interações entre Hospedeiro e Microrganismos , Consumo de Bebidas Alcoólicas/efeitos adversosRESUMO
The impact of stress on mental and digestive health has been extensively studied, with chronic stress being associated with various disorders. However, age-related differences in the response to acute stress, both behaviorally and physiologically, remain poorly understood. Therefore, this study aimed to develop a model to detect transient stress in mice of different ages. The stressor employed in our experiments was a restraint stress procedure, where mice were subjected to brief periods of immobilization to induce an acute stress response. Male C3H/HeN mice aged 3, 6, 12, and 30 weeks were subjected to acute restrain stress (ARS) by being placed in a 50 ml conical centrifuge tube for 15 min. Subsequently, their behavior, organ tissues, hematological parameters, cortisol concentration, and immune responses were assessed. Following ARS, the increased in time and entries into the center by the 12-week-old mice following stress. In comparison to mice of other ages, those aged 6 weeks demonstrated notable elevations in erythrocytes, platelets, hemoglobin, and hematocrit, all of which were influenced by the time-dependent changes and the recovery process of ARS. Blood corticosterone levels were substantially elevated in all age groups after ARS. Furthermore, ARS induced a notable increase in leukocytes, basophils, residential macrophages, and CD4+ T cells in all age groups except for 3-week-old mice. However, the number of monocyte-derived macrophages and CD8+ T cells did not change significantly. Additionally, mice aged 3 and 6 weeks demonstrated an increase in GFAP+ cells following ARS, whereas NeuN+ cells decreased across all ages. These results suggest that ARS has varying effects on the behavior, cortisol concentration, and quantity of blood cells as well as hepatic immune cells in mice of different ages. These age-dependent responses shed light on the complex interplay between stress and physiological systems and contribute to the broader understanding of stress-related diseases.
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Linfócitos T CD8-Positivos , Hidrocortisona , Camundongos , Masculino , Animais , Camundongos Endogâmicos C3H , Leucócitos , Corticosterona , Estresse Psicológico , Restrição FísicaRESUMO
BACKGROUND & AIMS: A long immune-tolerant (IT) phase lasting for decades and delayed HBeAg seroconversion (HBe-SC) in patients with chronic hepatitis B (CHB) increase the risk of liver diseases. Early entry into the immune-active (IA) phase and HBe-SC confers a favorable clinical outcome with an unknown mechanism. We aimed to identify factor(s) triggering IA entry and HBe-SC in the natural history of CHB. METHODS: To study the relevance of gut microbiota evolution in the risk of CHB activity, fecal samples were collected from CHB patients (n = 102) in different disease phases. A hepatitis B virus (HBV)-hydrodynamic injection (HDI) mouse model was therefore established in several mouse strains and germ-free mice, and multiplatform metabolomic and bacteriologic assays were performed. RESULTS: Ruminococcus gnavus was the most abundant species in CHB patients in the IT phase, whereas Akkermansia muciniphila was predominantly enriched in IA patients and associated with alanine aminotransferase flares, HBeAg loss, and early HBe-SC. HBV-HDI mouse models recapitulated this human finding. Increased cholesterol-to-bile acids (BAs) metabolism was found in IT patients because R gnavus encodes bile salt hydrolase to deconjugate primary BAs and augment BAs total pool for facilitating HBV persistence and prolonging the IT course. A muciniphila counteracted this activity through the direct removal of cholesterol. The secretome metabolites of A muciniphila, which contained small molecules structurally similar to apigenin, lovastatin, ribavirin, etc., inhibited the growth and the function of R gnavus to allow HBV elimination. CONCLUSIONS: R gnavus and A muciniphila play opposite roles in HBV infection. A muciniphila metabolites, which benefit the elimination of HBV, may contribute to future anti-HBV strategies.
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Clostridiales , Hepatite B Crônica , Animais , Humanos , Camundongos , Akkermansia , Colesterol , Antígenos E da Hepatite B , Microbioma GastrointestinalRESUMO
Tight junctions (TJs) are cell-cell interactions that localize at the most apical portion of epithelial/endothelial cells. One of the predominant functions of TJs is to regulate material transport through paracellular pathway, which serves as a selective barrier. In recent years, the expression and function of TJs in salivary glands has attracted great interest. The characteristics of multiple salivary gland TJ proteins have been identified. During salivation, the activation of muscarinic acetylcholine receptor and transient receptor potential vanilloid subtype 1, as well as other stimuli, promote the opening of acinar TJs by inducing internalization of TJs, thereby contributing to increased paracellular permeability. Besides, endothelial TJs are also redistributed with leakage of blood vessels in cholinergic-stimulated submandibular glands. Furthermore, under pathological conditions, such as Sjögren's syndrome, diabetes mellitus, immunoglobulin G4-related sialadenitis, and autotransplantation, the integrity and barrier function of TJ complex are impaired and may contribute to hyposalivation. Moreover, in submandibular glands of Sjögren's syndrome mouse model and patients, the endothelial barrier is disrupted and involved in hyposecretion and lymphocytic infiltration. These findings enrich our understanding of the secretory mechanisms that link the importance of epithelial and endothelial TJ functions to salivation under both physiological and pathophysiological conditions.
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Sialorreia , Síndrome de Sjogren , Camundongos , Animais , Humanos , Junções Íntimas/metabolismo , Junções Íntimas/patologia , Síndrome de Sjogren/patologia , Células Endoteliais , Glândulas Salivares/patologia , Saliva/metabolismo , Glândula Submandibular/metabolismoRESUMO
There are numerous established techniques for isolating hepatic myeloid cells; however, preserving their phenotypic and functional characteristics can be challenging. We present a straightforward and efficient method to isolate hepatic myeloid cells, including Kupffer cells and lymphocyte antigen 6 complex, locus C+ (Ly6C+) monocytes/macrophages. The procedure involves perfusion of the liver with collagenase and purification with immunomagnetic particles. This protocol ensures the isolation of large quantities of purified, viable, and functional cells without influencing their physiological characteristics. For complete details on the use and execution of this protocol, please refer to Wu et al. (2019).1.
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Hepatócitos , Fígado , Camundongos , Animais , Citometria de Fluxo , Células de KupfferRESUMO
Abscondita cerata is the most abundant and widely distributed endemic firefly species in Taiwan and is considered a key environmental and ecological indicator organism. In this study, we report the first long-read genome sequencing of Abs. cerata sequenced by Nanopore technology. The draft genome size, 967 Mb, was measured through a hybrid approach that consisted of assembling using 11.25-Gb Nanopore long reads and polishing using 9.47-Gb BGI PE100 short reads. The drafted genome was assembled into 4,855 contigs, with the N50 reaching 325.269 kb length. The assembled genome was predicted to possess 55,206 protein-coding genes, of which 20,862 (37.78%) were functionally annotated with public databases. 47.11% of the genome sequences consisted of repeat elements; among them DNA transposons accounted for the largest proportion (26.79%). A BUSCO (Benchmarking Universal Single Copy Orthologs) evaluation demonstrated that the genome and gene completeness were 84.8% and 79%, respectively. The phylogeny constructed using 1,792 single copy genes was consistent with previous studies. The comparative transcriptome between adult male head and lantern tissues revealed (1) the vision of Abs. cerata is primarily UV-sensitive to environmental twilight, which determines when it begins its nocturnal activity, (2) the major expressed OR56d receptor may be correlated to suitable humidity sensing, and (3) Luc1-type luciferase is responsible for Abs. cerata's luminescent spectrum.
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OBJECTIVES: Tight junctions (TJs) are involved in the regulation of salivary secretion via paracellular pathway. Botulinum toxin type A (BTXA) is widely used for the treatment of hypersecretion diseases such as sialorrhea. This study aimed to investigate the role of TJs in BTXA-inhibited secretion of the submandibular gland (SMG). MATERIALS AND METHODS: BTXA was injected into the SMGs of rats, and the same amount of saline was injected as a control. Western blot, real-time PCR, and immunofluorescence staining were used to detect the expression and distribution of TJ proteins. Paracellular permeability was evaluated using the transepithelial electrical resistance (TER) measurements and fluorescent tracer detection in BTXA-stimulated SMG-C6 cells. RESULTS: BTXA injection into the SMGs of rats led to increased expression of claudin (Cldn) -1 and Cldn3. Immunofluorescence staining showed no significant changes in the distribution of TJ proteins. In vitro, BTXA increased the TER values and significantly reduced the permeability of fluorescent tracer, suggesting that BTXA decreased the paracellular permeability. The expression levels of Cldn1, Cldn3, and Cldn4 were upregulated after BTXA treatment. CONCLUSION: The expression of TJ proteins changed in both animal models and SMG-C6 cells after BTXA treatment, which may contribute to the inhibition of salivary secretion.
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Toxinas Botulínicas Tipo A , Junções Íntimas , Ratos , Animais , Junções Íntimas/fisiologia , Toxinas Botulínicas Tipo A/farmacologia , Toxinas Botulínicas Tipo A/metabolismo , Salivação , Glândula Submandibular/metabolismoRESUMO
AIMS: Inhibitor of soluble epoxide hydrolase (t-AUCB) has been used in the experimental therapy of hypertension. This study aimed to investigate whether the secretion of submandibular glands (SMGs) altered in renal hypertensive rats, and to explore whether t-AUCB could improve the salivary secretion. MAIN METHODS: 2-kidney 1-clip Sprague-Dawley rats were used as renal hypertensive animals. t-AUCB treatment was given for 1 week after 8 weeks modeling. Blood pressure, blood perfusion and the secretion of SMGs, and endothelium-dependent relaxation of external maxillary artery were measured to investigate the effects of t-AUCB on the vascular tone and the secretion of SMGs in renal hypertensive rats. SMGs were collected for histological evaluation and the internal arteries were dissected for primary endothelial cells culture. KEY FINDINGS: The blood perfusion and flow rate of SMGs in the renal hypertensive rats were significantly lower than those in the controls. Endothelium-dependent relaxation of the external maxillary artery and AMPK/Akt/eNOS signaling was impaired in hypertensive rats. The glandular morphology and the concentration of salivary ions did not change obviously. t-AUCB treatment ameliorated the secretion of SMGs, the blood perfusion, and the dysfunction of endothelium-dependent relaxation of the external maxillary artery by activating the AMPK/Akt/eNOS pathway in hypertensive rats. SIGNIFICANCE: t-AUCB increases the blood perfusion through ameliorating dysfunction of endothelium-dependent relaxation of SMGs arteries and thus improves the hyposecretion of SMGs in hypertensive rats.
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Epóxido Hidrolases , Hipertensão , Proteínas Quinases Ativadas por AMP , Animais , Benzoatos , Células Endoteliais/metabolismo , Endotélio/metabolismo , Inibidores Enzimáticos/farmacologia , Epóxido Hidrolases/metabolismo , Hipertensão/tratamento farmacológico , Hipertensão/metabolismo , Proteínas Proto-Oncogênicas c-akt , Ratos , Ratos Sprague-Dawley , Glândulas Salivares/metabolismo , Ureia/análogos & derivadosRESUMO
PURPOSE: Cardiac fibrosis is characterized by the excessive deposition of extracellular matrix (ECM) proteins and leads to the maladaptive changes in myocardium. Endothelial cells (ECs) undergoing mesenchymal transition contributes to the occurrence and development of cardiac fibrosis. CD146 is an adhesion molecule highly expressed in ECs. The present study was performed to explore the role of CD146 in modulating endothelial to mesenchymal transition (EndMT). METHODS: C57BL/6 mice were subjected to subcutaneous implantation of osmotic minipump infused with angiotensin II (Ang â ¡). Adenovirus carrying CD146 short hairpin RNA (shRNA) or CD146 encoding sequence were infected into cultured human umbilical vein endothelial cells (HUVECs) followed by stimulation with Ang II or transforming growth factor-ß1 (TGF-ß1). Differentially expressed genes were revealed by RNA-sequencing (RNA-Seq) analysis. Gene expression was measured by quantitative real-time PCR, and protein expression and distribution were determined by Western blot and immunofluorescence staining, respectively. RESULTS: CD146 was predominantly expressed by ECs in normal mouse hearts. CD146 was upregulated in ECs but not fibroblasts and myocytes in hearts of Ang II-infused mice and in HUVECs stimulated with Ang â ¡. RNA-Seq analysis revealed the differentially expressed genes related to EndMT and Wnt/ß-catenin signaling pathway. CD146 knockdown and overexpression facilitated and attenuated, respectively, EndMT induced by Ang II or TGF-ß1. CD146 knockdown upregulated Wnt pathway-related genes including Wnt4, LEF1, HNF4A, FOXA1, SOX6, and CCND3, and increased the protein level and nuclear translocation of ß-catenin. CONCLUSIONS: Knockdown of CD146 exerts promotional effects on EndMT via activating Wnt/ß-catenin pathway and the upregulation of CD146 might play a protective role against EndMT and cardiac fibrosis.
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Fator de Crescimento Transformador beta1 , beta Catenina , Animais , Antígeno CD146/genética , Antígeno CD146/imunologia , Antígeno CD146/metabolismo , Células Cultivadas , Transição Epitelial-Mesenquimal , Fibrose , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Fator de Crescimento Transformador beta1/metabolismo , Via de Sinalização Wnt , beta Catenina/metabolismoRESUMO
BACKGROUND: Hyposalivation is one of the common symptoms of diabetes. Although long non-coding RNAs (lncRNAs) have recently been reported to play important roles in the pathogenesis of diabetes, the role of lncRNAs in diabetes-induced hyposalivation remains unknown. METHODS: The present study aimed to explore the function of lncRNA-microRNA-mRNA regulatory network in the submandibular gland (SMGs) under the context of diabetes. LncRNA expression profile of the SMGs was analyzed using microarray technology. Differentially expressed lncRNAs were confirmed using real-time quantitative PCR. Bioinformatics analyses were performed, and Coding-non-coding gene co-expression (CNC) and competing endogenous RNA (ceRNA) networks were constructed to explore the potential mechanisms of diabetes-induced hyposalivation. RESULTS: A total of 1273 differentially expressed lncRNAs (536 up-regulated and 737 downregulated) were identified in the SMGs tissues of db/db mice. CNC and ceRNA network analyses were performed based on five differentially expressed lncRNAs validated by real-time quantitative PCR. Gene Ontology analysis of target genes of CNC network revealed that "calcium ion binding" was a highly enriched molecular function. Kyoto Encyclopedia of Genes and Genomes pathway analysis of target genes of ceRNA network revealed that the "mammalian target of rapamycin signaling pathway" was significantly enriched. CONCLUSIONS: On the whole, the findings of the present study may provide insight into the possible mechanism of diabetes-induced hyposalivation.
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Diabetes Mellitus Experimental , MicroRNAs , RNA Longo não Codificante , Xerostomia , Animais , Diabetes Mellitus Experimental/genética , Perfilação da Expressão Gênica , Redes Reguladoras de Genes , Humanos , Mamíferos/genética , Mamíferos/metabolismo , Camundongos , MicroRNAs/genética , MicroRNAs/metabolismo , RNA Longo não Codificante/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Glândula Submandibular/metabolismoRESUMO
OBJECTIVE: IgG4-related sialadenitis (IgG4-RS) is a chronic fibroinflammatory disease characterized by glandular fibrosis and hyposalivation. This study was undertaken to explore the role of cellular senescence in the pathogenesis of IgG4-RS-related fibrosis. METHODS: The expression of senescence markers and proinflammatory cytokines in the submandibular glands (SMGs) of IgG4-RS patients (n = 18) and controls (n = 14) was determined by proteomics, real-time polymerase chain reaction, Western blotting, and immunohistochemistry. After interleukin-4 (IL-4) treatment, high-throughput RNA sequencing was performed to identify the differentially expressed genes in SMG-C6 cells. A glandular fibrosis model was established by the intraglandular injection of IL-4 into mouse SMGs (n = 8 per group). RESULTS: Salivary acinar and ductal epithelial cells underwent senescence in IgG4-RS patients, as indicated by the elevated activity of senescence-associated ß-galactosidase, lipofuscin accumulation, enhanced expression of senescence markers (p53 and p16INK4A ), and up-regulation of senescence-associated secretory phenotype factors. Moreover, there was a significant increase in IL-4 levels in SMGs from IgG4-RS patients (P < 0.01), which positively correlated with p16INK4A expression and the fibrosis score. Incubation with IL-4 exacerbated salivary epithelial cell senescence by increasing the expression of p16INK4A through the reactive oxygen species (ROS)/p38 MAPK pathway. Supernatant collected from IL-4-induced senescent SMG-C6 cells enhanced fibroblast activation and matrix protein production (P < 0.05). Furthermore, injecting mice with IL-4 promoted fibrosis and senescence phenotypes in SMGs in vivo. CONCLUSION: The cellular senescence induced by IL-4 through the ROS/p38 MAPK-p16INK4A pathway promotes fibrogenesis in IgG4-RS. Our data suggest that cellular senescence could serve as a novel therapeutic target for treating IgG4-RS.
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Interleucina-4 , Sialadenite , Animais , Senescência Celular , Inibidor p16 de Quinase Dependente de Ciclina , Células Epiteliais/metabolismo , Fibrose , Humanos , Imunoglobulina G/metabolismo , Camundongos , Espécies Reativas de Oxigênio , Proteínas Quinases p38 Ativadas por MitógenoRESUMO
OBJECTIVE: Obesity contributes to the dysfunction of salivary gland. To explore the specific underlying mechanism for obesity-induced hyposalivation, a model for high-fat diet-induced obese (DIO) mice were constructed to analyze long non-coding RNAs (lncRNAs) and messenger RNAs (mRNAs) expression profiles. METHODS: The DIO group and control group were fed a diet containing 60 kcal% fat and a normal chow diet for 16 weeks respectively. Microarray analyses were performed to detect the expression profiles of lncRNA and mRNA in submandibular gland tissues from control group mice and DIO mice. Gene ontology, kyoto encyclopedia of genes and genomes, protein-protein interaction, coding-non-coding gene co-expression, transcription factors and competing endogenous RNA analyses were performed to examine the function of differentially expressed genes. RESULTS: Microarray analyses identified that 624 lncRNAs, along with 297 mRNAs were differentially expressed. Bioinformatic analyses revealed that "complement and coagulation cascades," "glutathione metabolism," "cysteine and methionine metabolism," and "estrogen signaling pathway" were significantly associated with candidate lncRNAs. Transcription factors analysis on candidate lncRNAs revealed several genes such as tribbles pseudokinase 3 may play regulatory roles. CONCLUSIONS: Our results revealed the expression profiles of lncRNAs and mRNAs and provided new insights into the mechanism of obesity-induced hyposalivation using bioinformatic analyses.
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RNA Longo não Codificante , Xerostomia , Animais , Dieta Hiperlipídica , Camundongos , Camundongos Obesos , Obesidade , RNA Longo não Codificante/genética , RNA Mensageiro/metabolismo , Glândula Submandibular/metabolismo , Fatores de Transcrição/genéticaRESUMO
Tight junctions (TJs) play an important role in water, ion, and solute transport through the paracellular pathway of epithelial cells; however, their role in diabetes-induced salivary gland dysfunction remains unknown. Here, we found that the TJ proteins claudin-1 and claudin-3 were significantly increased in the submandibular glands (SMGs) of db/db mice and high glucose (HG)-treated human SMGs. HG decreased paracellular permeability and increased claudin-1 and claudin-3 expression in SMG-C6 cells. Knockdown of claudin-1 or claudin-3 reversed the HG-induced decrease in paracellular permeability. MiR-22-3p was significantly downregulated in diabetic SMGs and HG-treated SMG-C6 cells. A miR-22-3p mimic suppressed claudin-1 and claudin-3 expression and abolished the HG-induced increases in claudin-1 and claudin-3 levels in SMG-C6 cells, whereas a miR-22-3p inhibitor produced the opposite effects. Specificity protein-1 (Sp1) was enhanced in diabetic SMGs and HG-treated SMG-C6 cells, which promoted claudin-1 and claudin-3 transcription through binding to the corresponding promoters. A luciferase reporter assay confirmed that miR-22-3p repressed Sp1 by directly targeting the Sp1 mRNA 3'-untranslated region (3'-UTR). Consistently, the miR-22-3p mimic suppressed, whereas the miR-22-3p inhibitor enhanced, the effects of HG on Sp1 expression. Taken together, our results demonstrate a new regulatory pathway through which HG decreases the paracellular permeability of SMG cells by inhibiting miR-22-3p/Sp1-mediated claudin-1 and claudin-3 expression.
Assuntos
Permeabilidade da Membrana Celular , Claudinas/metabolismo , Epitélio/metabolismo , Glucose/toxicidade , MicroRNAs/metabolismo , Transdução de Sinais , Fator de Transcrição Sp1/metabolismo , Glândula Submandibular/metabolismo , Animais , Sequência de Bases , Linhagem Celular , Permeabilidade da Membrana Celular/efeitos dos fármacos , Claudinas/genética , Diabetes Mellitus Experimental/patologia , Regulação para Baixo/efeitos dos fármacos , Epitélio/efeitos dos fármacos , Epitélio/ultraestrutura , Humanos , Masculino , Camundongos , MicroRNAs/genética , Regiões Promotoras Genéticas/genética , Ligação Proteica , Ratos , Transdução de Sinais/efeitos dos fármacos , Glândula Submandibular/ultraestrutura , Técnicas de Cultura de Tecidos , Transcrição Gênica , Regulação para Cima/efeitos dos fármacosRESUMO
BACKGROUND: Long noncoding RNAs (lncRNAs) are important regulators in tumor progression. However, their biological functions and underlying mechanisms in hypoxia adaptation remain largely unclear. RESULTS: Here, we established a correlation between a Chr3q29-derived lncRNA gene and tongue squamous carcinoma (TSCC) by genome-wide analyses. Using RACE, we determined that two novel variants of this lncRNA gene are generated in TSCC, namely LINC00887_TSCC_short (887S) and LINC00887_TSCC_long (887L). RNA-sequencing in 887S or 887L loss-of-function cells identified their common downstream target as Carbonic Anhydrase IX (CA9), a gene known to be upregulated by hypoxia during tumor progression. Mechanistically, our results showed that the hypoxia-augmented 887S and constitutively expressed 887L functioned in opposite directions on tumor progression through the common target CA9. Upon normoxia, 887S and 887L interacted. Upon hypoxia, the two variants were separated. Each RNA recognized and bound to their responsive DNA cis-acting elements on CA9 promoter: 887L activated CA9's transcription through recruiting HIF1α, while 887S suppressed CA9 through DNMT1-mediated DNA methylation. CONCLUSIONS: We provided hypoxia-permitted functions of two antagonistic lncRNA variants to fine control the hypoxia adaptation through CA9.
Assuntos
Carcinoma de Células Escamosas , Neoplasias da Língua , Anidrase Carbônica IX/genética , Carcinoma de Células Escamosas/genética , Linhagem Celular Tumoral , Estudo de Associação Genômica Ampla , Humanos , Hipóxia/genética , RNA Longo não Codificante/genética , Língua , Neoplasias da Língua/genéticaRESUMO
Hepatitis B virus (HBV) chronically infects 257 million people and is one of the most important liver diseases worldwide. A unique feature of HBV infection in humans is that viral clearance heavily depends on the age at exposure. Recent studies demonstrated that the virus takes advantage of immature innate immunity, especially hepatic macrophages, and not-yet-stabilized gut microbiota in early life to establish a chronic infection. The liver contains resident and infiltrating myeloid cells involved in immune responses to pathogens. They influence both innate and adaptive sectors of the immune system and their interplay with HBV has only been noticed recently. Here, we discuss how interactions between gut microbiota and hepatic macrophages influence the outcomes of HBV infection. Understanding the underlying mechanism would pave the way for the treatment of chronic HBV infection.