circMTO1/miR-30c-5p/SOCS3 axis alleviates oral submucous fibrosis through inhibiting fibroblast-myofibroblast transition.

BACKGROUND: circRNAs have been shown to participate in diverse diseases; however, their role in oral submucous fibrosis (OSF), a potentially malignant disorder, remains obscure. Our preliminary experiments detected the expression of circRNA mitochondrial translation optimization 1 homologue (circMTO1) in OSF tissues (n = 20) and normal mucosa tissues (n = 20) collected from Hunan Xiangya Stomatological Hospital, and a significant decrease of circMTO1 expression was showed in OSF tissues. Therefore, we further explored circMTO1 expression in OSF. METHODS: Target molecule expression was detected using RT-qPCR and western blotting. The migration and invasion of buccal mucosal fibroblasts (BMFs) were assessed using wound healing and Transwell assays. The interaction between miR-30c-5p, circMTO1, and SOCS3 was evaluated using dual luciferase, RNA immunoprecipitation (RIP), and RNA pull-down assays. The colocalisation of circMTO1 and miR-30c-5p was observed using fluorescence in situ hybridisation (FISH). RESULTS: circMTO1 and SOCS3 expression decreased, whereas miR-30c-5p expression increased in patients with OSF and arecoline-stimulated BMFs. Overexpression of circMTO1 effectively restrained the fibroblast-myofibroblast transition (FMT), as evidenced by the increase in expression of Coll I, α-SMA, Vimentin, and the weakened migration and invasion functions in BMFs. Mechanistic studies have shown that circMTO1 suppresses FMT by enhancing SOCS3 expression by sponging miR-30c-5p and subsequently inactivating the FAK/PI3K/AKT pathway. FMT induced by SOCS3 silencing was reversed by the FAK inhibitor TAE226 or the PI3K inhibitor LY294002. CONCLUSION: circMTO1/miR-30c-5p/SOCS3 axis regulates FMT in arecoline-treated BMFs via the FAK/PI3K/AKT pathway. Expanding the sample size and in vivo validation could further elucidate their potential as therapeutic targets for OSF.

The role of epithelial cells in fibrosis: Mechanisms and treatment.

Fibrosis is a pathological process that affects multiple organs and is considered one of the major causes of morbidity and mortality in multiple diseases, resulting in an enormous disease burden. Current studies have focused on fibroblasts and myofibroblasts, which directly lead to imbalance in generation and degradation of extracellular matrix (ECM). In recent years, an increasing number of studies have focused on the role of epithelial cells in fibrosis. In some cases, epithelial cells are first exposed to external physicochemical stimuli that may directly drive collagen accumulation in the mesenchyme. In other cases, the source of stimulation is mainly immune cells and some cytokines, and epithelial cells are similarly altered in the process. In this review, we will focus on the multiple dynamic alterations involved in epithelial cells after injury and during fibrogenesis, discuss the association among them, and summarize some therapies targeting changed epithelial cells. Especially, epithelial mesenchymal transition (EMT) is the key central step, which is closely linked to other biological behaviors. Meanwhile, we think studies on disruption of epithelial barrier, epithelial cell death and altered basal stem cell populations and stemness in fibrosis are not appreciated. We believe that therapies targeted epithelial cells can prevent the progress of fibrosis, but not reverse it. The epithelial cell targeting therapies will provide a wonderful preventive and delaying action.

Drug-loaded mucoadhesive microneedle patch for the treatment of oral submucous fibrosis.

Oral submucous fibrosis is a chronic, inflammatory and potentially malignant oral disease. Local delivery of triamcinolone to lesion site is a commonly used therapy. The existing methods for local drug delivery include topical administration and submucosal injection. However, in the wet and dynamic oral microenvironment, these methods have drawbacks such as limited drug delivery efficiency and injection pain. Therefore, it is urgently needed to develop an alternative local drug delivery system with high efficiency and painlessness. Inspired by the structure of band-aid, this study proposed a novel double-layered mucoadhesive microneedle patch for transmucosal drug delivery. The patch consisted of a mucoadhesive silk fibroin/tannic acid top-layer and a silk fibroin microneedle under-layer. When applying the annealing condition for the medium content of ß-sheets of silk fibroin, the microneedles in under-layer displayed both superior morphology and mechanical property. The mechanical strength of per needle (0.071N) was sufficient to penetrate the oral mucosa. Sequentially, the gelation efficiency of silk fibroin and tannic acid in top-layer was maximized as the weight ratio of tannic acid to silk fibroin reached 5:1. Moreover, in vitro results demonstrated the double-layered patch possessed undetectable cytotoxicity. The sustained release of triamcinolone was observed from the double-layered patch for at least 7 days. Furthermore, compared with other commercial buccal patches, the double-layered patch exhibited an enhanced wet adhesion strength of 37.74 kPa. In addition, ex vivo mucosal tissue penetration experiment confirmed that the double-layered patch could reach the lamina propria, ensuring effective drug delivery to the lesion site of oral submucous fibrosis. These results illustrate the promising potential of the drug-loaded mucoadhesive microneedle patch for the treatment of oral submucous fibrosis.

Immunomodulatory zinc-based materials for tissue regeneration.

Zinc(Zn)-based materials have contributed greatly to the rapid advancements in tissue engineering. The qualities they possess that make them so beneficial include their excellent biodegradability, biocompatibility, anti-bacterial activity, among and several others. Biomedical materials that act as a foreign body, will inevitably cause host immune response when introduced to the human body. As the osteoimmunology develops, the immunomodulatory characteristics of biomaterials have become an appealing concept to improve implant-tissue interaction and tissue restoration. Recently, Zn-based materials have also displayed immunomodulatory functions, especially macrophage polarization states. It can promote the transformation of M1 macrophages into M2 macrophages to enhance the tissue regeneration and reconstruction. This review covers mainly Zn-based materials and their characteristics, including metallic Zn alloys and Zn ceramics. We highlight the current advancements in the type of immune responses, as well as the mechanisms, that are induced by Zn-based biomaterials, most importantly the regulation of innate immunity and the mechanism of promoting tissue regeneration. To this end, we discuss their applications in biomedicine, and conclude with an outlook on future research challenges.

Administration methods of lipid-based nanoparticle delivery systems for cancer treatment.

As nano medications have developed in the recent four decades, nano-delivery systems have been applied in treating various diseases and are especially common in cancer treatment. Nano-delivery systems could target cancer-associated cells, enhance the accuracy and efficacy of treatment, and reduce systemic side effects. Among the many drugs on nano-carriers, the load system of lipid-based nanoparticles has the brightest prospect due to the high level of biocompatibility, biodegradability, loading capability, and immunogenicity. Previous reviews have comprehensively introduced their effects and progress. However, most of them did not provide great attention to practical applications. This article will focus on different intake methods, which decide the biological process of drugs. This suggests that we can modify lipid-based nano-delivery systems according to how they are capable of prolonging the half-life span and magnifying therapy effects in treating cancer. Besides, we put forth the problems that should be further studied in the future and their probable solutions.

Prognosis-related molecular subtyping in head and neck squamous cell carcinoma patients based on glycolytic/cholesterogenic gene data.

BACKGROUND: Head and neck squamous cell carcinoma (HNSCC) remains an unmet medical challenge. Metabolic reprogramming is a hallmark of diverse cancers, including HNSCC. METHODS: We investigated the metabolic profile in HNSCC by using The Cancer Genome Atlas (TCGA) (n = 481) and Gene Expression Omnibus (GEO) (n = 97) databases. The metabolic stratification of HNSCC samples was identified by using unsupervised k-means clustering. We analyzed the correlations of the metabolic subtypes in HNSCC with featured genomic alterations and known HNSCC subtypes. We further validated the metabolism-related subtypes based on features of ENO1, PFKFB3, NSDHL and SQLE expression in HNSCC by Immunohistochemistry. In addition, genomic characteristics of tumor metabolism that varied among different cancer types were confirmed. RESULTS: Based on the median expression of coexpressed cholesterogenic and glycolytic genes, HNSCC subtypes were identified, including glycolytic, cholesterogenic, quiescent and mixed subtypes. The quiescent subtype was associated with the longest survival and was distributed in stage I and G1 HNSCC. Mutation analysis of HNSCC genes indicated that TP53 has the highest mutation frequency. The CDKN2A mutation frequency has the most significant differences amongst these four subtypes. There is good overlap between our metabolic subtypes and the HNSCC subtype. CONCLUSION: The four metabolic subtypes were successfully determined in HNSCC. Compared to the quiescent subtype, glycolytic, cholesterogenic and mixed subtypes had significantly worse outcome, which might offer guidelines for developing a novel treatment strategy for HNSCC.

Opposing functions of circadian protein DBP and atypical E2F family E2F8 in anti-tumor Th9 cell differentiation.

Interleukin-9 (IL-9)-producing CD4+ T helper cells (Th9) have been implicated in allergy/asthma and anti-tumor immunity, yet molecular insights on their differentiation from activated T cells, driven by IL-4 and transforming growth factor-beta (TGF-ß), is still lacking. Here we show opposing functions of two transcription factors, D-binding protein (DBP) and E2F8, in controlling Th9 differentiation. Specifically, TGF-ß and IL-4 signaling induces phosphorylation of the serine 213 site in the linker region of the Smad3 (pSmad3L-Ser213) via phosphorylated p38, which is necessary and sufficient for Il9 gene transcription. We identify DBP and E2F8 as an activator and repressor, respectively, for Il9 transcription by pSmad3L-Ser213. Notably, Th9 cells with siRNA-mediated knockdown for Dbp or E2f8 promote and suppress tumor growth, respectively, in mouse tumor models. Importantly, DBP and E2F8 also exhibit opposing functions in regulating human TH9 differentiation in vitro. Thus, our data uncover a molecular mechanism of Smad3 linker region-mediated, opposing functions of DBP and E2F8 in Th9 differentiation.

In Vivo Generation of SSA/Ro Antigen-Specific Regulatory T Cells Improves Experimental Sjögren's Syndrome in Mice.

OBJECTIVE: Sjögren's syndrome (SS) is a systemic autoimmune disease, and T cells play an important role in the initiation and perpetuation of the disease. In this study, we developed an immunotherapy for NOD/LtJ mice with SS-like symptoms by combining a transient depletion of CD4+ T cells with the administration of autoantigen-specific peptide Ro480. METHODS: NOD/LtJ mice were treated with single anti-CD4 monoclonal antibody (mAb) followed 2 days later by a series of 6 intraperitoneal injections of Ro480-494 every other day. Salivary flow rates were determined pre- and posttreatment once a week. Mice were euthanized 6 weeks after the initial anti-CD4 mAb treatment, salivary glands (SGs) were collected for analyses of histologic disease scores and inflammatory cell infiltration, polymerase chain reaction determination of genes was conducted, and flow cytometry analysis including major histocompatibility complex class II tetramer staining of immune cells was performed. In addition, adoptive transfer of Treg cells was administrated to investigate the function of the newly generating Treg cells in vivo. RESULTS: The combination of anti-CD4 mAb with autoantigen-specific peptide Ro480 generated SSA/Ro antigen-specific Treg cells in vivo, which can suppress interferon-γ production of CD4+ T cells and inflammation infiltration in SGs and maintain the function of SGs. CONCLUSION: Our findings provide a new approach to generating antigen-specific Treg cells in vivo for SS treatment, which may have implications for potential therapy for patients with SS.

Ginsenoside Rh2 inhibits breast cancer cell growth via ERß-TNFα pathway.

Ginsenoside Rh2 is one of rare panaxidiols extracted from Panax ginseng and a potential estrogen receptor ligand that exhibits moderate estrogenic activity. However, the effect of Rh2 on growth inhibition and its underlying molecular mechanism in human breast cells are not fully understood. In this study, we tested cell viability by MTT and colony formation assays. Cell growth and cell cycle were determined to investigate the effect of ginsenoside Rh2 by flow cytometry. The expressions of estrogen receptors (ERs), TNFα, and apoptosis-related proteins were detected by qPCR and western blot analysis. The mechanisms of ERα and ERß action were determined using transfection and inhibitors. Antitumor effect of ginsenoside Rh2 against MCF-7 cells was investigated in xenograft mice. Our results showed that ginsenoside Rh2 induced apoptosis and G1/S phase arrest in MCF-7 cells. Treatment of cells with ginsenoside Rh2 down-regulated protein levels of ERα, and up-regulated mRNA and protein levels of ERß and TNFα. We also found that ginsenoside Rh2-induced TNFα over-expression is through up-regulation of ERß initiated by ginsenoside Rh2. Furthermore, ginsenoside Rh2 induced MCF-7 cell apoptosis via estrogen receptor ß-TNFα pathway in vivo. These results demonstrate that ginsenoside Rh2 promotes TNFα-induced apoptosis and G1/S phase arrest via regulation of ERß.

Circular RNA ITCH: An Emerging Multifunctional Regulator.

In the last decade, numerous circRNAs were discovered by virtue of the RNA-Seq technique. With the deepening of experimental research, circRNAs have brought to light the key biological functions and progression of human diseases. CircRNA ITCH has been demonstrated to be a tumor suppressor in numerous cancers, and recently it was found to play an important role in bone diseases, diabetes mellitus, and cardiovascular diseases. However, the functions of circ-ITCH have not been completely understood. In this review, we comprehensively provide a conceptual framework to elucidate circ-ITCH biological functions of cell proliferation, apoptosis and differentiation, and the pathological mechanisms of inflammation, drug resistance/toxicity, and tumorigenesis. Finally, we summarize its clinical applications in various diseases. This research aimed at clarifying the role of circ-ITCH, which could be a promising therapeutic target.

Aging and Mesenchymal Stem Cells: Therapeutic Opportunities and Challenges in the Older Group.

With aging, a portion of cells, including mesenchymal stem cells (MSCs), become senescent, and these senescent cells accumulate and promote various age-related diseases. Therefore, the older age group has become a major population for MSC therapy, which is aimed at improving tissue regeneration and function of the aged body. However, the application of MSC therapy is often unsatisfying in the aged group. One reasonable conjecture for this correlation is that aging microenvironment reduces the number and function of MSCs. Cellular senescence also plays an important role in MSC function impairment. Thus, it is necessary to explore the relationship between senescence and MSCs for improving the application of MSCs in the elderly. Here, we present the influence of aging on MSCs and the characteristics and functional changes of senescent MSCs. Furthermore, current therapeutic strategies for improving MSC therapy in the elderly group are also discussed.

Senolytics improve bone forming potential of bone marrow mesenchymal stem cells from aged mice.

The osteogenic potential of bone marrow mesenchymal stem cells (BMSCs) declines dramatically with aging. By using a calvarial defect model, we showed that a senolytic cocktail (dasatinib+quercetin; D + Q) improved osteogenic capacity of aged BMSC both in vitro and in vivo. The study presented a model to assess strategies to improve bone-forming potential on aged BMSCs. D + Q might hold promise for improving BMSC function in aged populations.

Naturally-occurring spinosyn A and its derivatives function as argininosuccinate synthase activator and tumor inhibitor.

Argininosuccinate synthase (ASS1) is a ubiquitous enzyme in mammals that catalyzes the formation of argininosuccinate from citrulline and aspartate. ASS1 genetic deficiency in patients leads to an autosomal recessive urea cycle disorder citrullinemia, while its somatic silence or down-regulation is very common in various human cancers. Here, we show that ASS1 functions as a tumor suppressor in breast cancer, and the pesticide spinosyn A (SPA) and its derivative LM-2I suppress breast tumor cell proliferation and growth by binding to and activating ASS1. The C13-C14 double bond in SPA and LM-2I while the Cys97 (C97) site in ASS1 are critical for the interaction between ASS1 and SPA or LM-2I. SPA and LM-2I treatment results in significant enhancement of ASS1 enzymatic activity in breast cancer cells, particularly in those cancer cells with low ASS1 expression, leading to reduced pyrimidine synthesis and consequently the inhibition of cancer cell proliferation. Thus, our results establish spinosyn A and its derivative LM-2I as potent ASS1 enzymatic activator and tumor inhibitor, which provides a therapeutic avenue for tumors with low ASS1 expression and for those non-tumor diseases caused by down-regulation of ASS1.

CircHIPK3: Key Player in Pathophysiology and Potential Diagnostic and Therapeutic Tool.

A large number of studies in China and other countries have confirmed that circularHIPK3 (circHIPK3) plays an important role in the pathophysiological processes of various diseases. Through the action of sponge miRNA (miR), circHIPK3 regulates cell proliferation, differentiation, and migration, and plays a key role in disease processes. By referring to a large number of research reports, this article explores the specific functional role of circHIPK3 in fibrotic diseases, cancer, and other diseases. This review aims to clarify the role of circHIPK3 in disease processes in order to aid further studies into the specific pathogenesis and clinical diagnosis of various diseases and provide new ideas for treatments.

Interleukin (IL)-33: an orchestrator of immunity from host defence to tissue homeostasis.

Interleukin (IL)-33, a member of the IL-1 superfamily, functions as an alarm signal, which is released upon cell injury or tissue damage to alert the immune system. It has emerged as a chief orchestrator in immunity and has a broad pleiotropic action that influences differentiation, maintenance and function of various immune cell types via the ST2 receptor. Although it has been strongly associated with immunopathology, critically, IL-33 is involved in host defence, tissue repair and homeostasis. In this review, we provide an overview of the signalling pathway of IL-33 and highlight its regulatory functions in immune cells. Furthermore, we attempt a broader discussion of the emerging functions of IL-33 in host defence, tissue repair, metabolism, inflammatory disease and cancer, suggesting potential avenues to manoeuvre IL-33/ST2 signalling as treatment options.

Human dental pulp stem cells regulate allogeneic NK cells' function via induction of anti-inflammatory purinergic signalling in activated NK cells.

OBJECTIVES: Mesenchymal stem cells (MSCs) could regulate the function of various immune cells. It remains unclear whether MSCs additionally possess immunostimulatory properties. We investigated the impact of human MSCs on the responsiveness of primary natural killer (NK) cells in terms of induction of anti-inflammatory purinergic signalling. MATERIAL AND METHODS: We obtained human bone marrow mesenchymal stem cells (BMMSCs) and dental pulp stem cells (DPSCs). NK cells were isolated from peripheral blood of healthy volunteers. Activated NK cells were cultured with MSCs. Proliferation assay, apoptosis analysis, activating or inhibitory receptor expression and degranulation assay were used to explore NK cells' function. High-performance liquid chromatography was used to investigate the purinergic signalling in activated NK cells. RESULTS: Both DPSCs and BMMSCs could impair proliferation and promote apoptosis of activated NK cells. Also, activated NK cells could cause DPSCs to lyse. Furthermore, the expression of activating NK cells' receptors was decreased, but inhibitory receptors of NK cells were elevated following co-cultivation. NK cells acquired CD73 expression, while MSCs could release ATP into the extracellular space where nucleotides were converted into adenosine (ADO) following co-culture system. Under the existence of exogenous 2-chloroadenosine (CADO), the cytotoxic capacity of NK cells was remarkably depressed in a concentration-dependent manner. CONCLUSIONS: DPSCs and BMMSCs could depress NK cells' function by hydrolysing ATP to ADO using CD39 and CD73 enzymatic activity. Our data suggested that DPSCs might represent a new strategy for treating immune-related diseases by regulating previously unrecognized functions in innate immune responses.

Mitogen-activated protein kinase signaling pathway in oral cancer.

The mitogen-activated protein kinase (MAPK) signaling pathway is associated with tumor cell proliferation, differentiation, apoptosis, angiogenesis, invasion and metastasis. The present review assesses the involvement of the MAPK signaling pathway in oral cancer progression and invasion based on analysis of individual sub-pathways and their mechanisms of action. The regulation of this pathway for targeted oral cancer therapy is explored and the challenges confronting this, as well as corresponding potential solutions, are discussed. Exploring this pathway with an emphasis on its components, subfamilies, sub-pathways, interactions with other pathways and clinical practice modes may improve oral cancer treatment.

Electroactive BaTiO3 nanoparticle-functionalized fibrous scaffolds enhance osteogenic differentiation of mesenchymal stem cells.

It has been proven that the surface topographic cues of fiber arrangement can induce osteogenic differentiation of mesenchymal stem cells. However, this effect alone is weak and insufficient to meet the needs of regenerative medicine. In this work, electroactivity concept was introduced to enhance the osteoinductivity of fibrous scaffolds. The randomly oriented and aligned electroactive fibrous scaffolds of poly-(l-lactic acid) (PLLA) with incorporation of ferroelectric ceramic BaTiO3 (BTO) nanoparticles (NPs) were fabricated by electrospinning. Physicochemical properties, including fiber morphology, microstructure, composition, thermal stability, surface roughness, and surface wettability, of these fibrous scaffolds were studied. The dielectric properties of the scaffolds were evaluated. The results showed that the randomly oriented BTO/PLLA composite fibrous scaffolds had the highest dielectric permittivity of 1.19, which is of the same order of magnitude as the natural bone. The combined effects of fiber orientation and electrical activity on the osteogenic responses of bone marrow mesenchymal stem cells (BM-MSCs) were specifically investigated. Randomly oriented composite fibrous scaffolds significantly promoted polygonal spreading and encouraged early osteogenic differentiation in BM-MSCs, whereas aligned composite fibrous scaffolds promoted cell elongation and discouraged osteogenic differentiation. These results evidenced that randomly fiber orientation and biomimetic electric activity have combining effects on osteogenic differentiation of BM-MSCs. Our findings indicate that coupling effects of multi-physical properties should be paid more attention to mimic the microenvironment for enhancing osteogenic differentiation of BM-MSCs.

Investigation of the association between miR­181b, Bcl­2 and LRIG1 in oral verrucous carcinoma.

Abnormal expression of microRNAs (miRNAs) is involved in the development of and anti­apoptotic effects in various types of human cancer. However, miRNA­mediated regulation of oral verrucous carcinoma (OVC) remains to be elucidated. The present study aimed to investigate the expression of miR­181b in OVC and oral squamous cell carcinoma (OSCC). The expression levels of miR­181b were determined using reverse transcription­quantitative polymerase chain reaction. The expression levels of B­cell lymphoma 2 (Bcl­2) and leucine rich repeats and immunoglobulin like domains 1 (LRIG1), were evaluated using immunohistochemical staining. The correlation between Bcl­2 and LRIG1 expression was determined using a Pearson correlation analysis. The expression levels of miR­181b and Bcl­2 in OVC were significantly higher compared with normal mucosal tissue (NM); however, lower compared with the OSCC. The key target of miR­181b was LRIG1 and it was significantly lower in OVC tissues compared with NM tissue; however this was higher when compared with OSCC tissue. The expression levels of Bcl­2 were correlated with expression levels of LRIG1 in OVC tissues. Therefore, LRIG1 may be associated with anti­apoptotic function in OVC tissues.

An adaptive immune response driven by mature, antigen-experienced T and B cells within the microenvironment of oral squamous cell carcinoma.

Lymphocyte infiltrates have been observed in the microenvironment of oral cancer; however, little is known about whether the immune response of the lymphocyte infiltrate affects tumor biology. For a deeper understanding of the role of the infiltrating-lymphocytes in oral squamous cell carcinoma (OSCC), we characterized the lymphocyte infiltrate repertoires and defined their features. Immunohistochemistry revealed considerable T and B cell infiltrates and lymphoid follicles with germinal center-like structures within the tumor microenvironment. Flow cytometry demonstrated that populations of antigen-experienced CD4+ and CD8+ cells were present, as well as an enrichment of regulatory T cells; and T cells expressing programmed death-1 (PD-1) and T cell Ig and mucin protein-3 (Tim-3), indicative of exhaustion, within the tumor microenvironment. Characterization of tumor-infiltrating B cells revealed clear evidence of antigen exposure, in that the cardinal features of an antigen-driven B cell response were present, including somatic mutation, clonal expansion, intraclonal variation and isotype switching. Collectively, our results point to an adaptive immune response occurring within the OSCC microenvironment, which may be sustained by the expression of specific antigens in the tumor.