Stemness maintenance of stem cells derived from human exfoliated deciduous teeth (SHED) in 3D spheroid formation through the TGF-ß/Smad signaling pathway.

Mesenchymal stem cells (MSCs) have shown great potential as important therapeutic tools for dental pulp tissue engineering, with the maintenance and enhancement of their stemness being crucial for successful therapeutic application in vivo and three-dimensional (3D) spheroid formation considered a reliable technique for enhancing their pluripotency. Human exfoliated deciduous tooth stem cells (SHED) were cultured in a low attachment plate to form aggregates for five days. Then, the resulting spheroids were analyzed for pluripotent marker expression, paracrine secretory function, proliferation, signaling pathways involved, and distribution of key proteins within the spheroids. The results indicated that 3D spheroid formation significantly increased the activation of the transforming growth factor beta (TGF-ß)/Smad signaling pathway and upregulated the secretion and mRNA expression levels of TGF-ß, which in turn enhanced the expression of pluripotency markers in SHED spheroids. The activation of the TGF-ß/Smad signaling pathway through 3D spheroid formation was found to preserve the stemness properties of SHED. Thus, understanding the mechanisms behind pluripotency maintenance of SHED culture through 3D spheroid formation could have implications for the therapeutic application of MSCs in regenerative medicine and tissue engineering.

Quercetin inhibits the activity and function of dendritic cells through the TLR4/IRAK4/NF-κB signalling pathway.

Introduction: To investigate the inhibitory effect of quercetin (QUE) on dendritic cells (DCs) through the toll-like receptor 4/interleukin-1 receptor-associated kinase 4/nuclear factor kappa-B (TLR4/IRAK4/NF-κB) signalling pathway. Material and methods: CCK-8 and apoptosis assays were performed to determine the optimal concentration and action time of QUE to inhibit DCs. Protein extracts from treated DCs were used for Western blotting experiments to determine the relative expression levels of TLR4, IRAK4, and NF-κB p65 proteins. Changes in the ratio of CD86 and CD11c positive cells on the DCs surface were detected using flow cytometry. The molecular docking technique was used to analyse the binding site and free energy of QUE and IRAK4. Results: CCK-8 and apoptosis assays suggested that QUE inhibited the activity and function of DCs in a time-dose-dependent manner. The results of Western blotting suggested that the relative expression levels of TLR4, IRAK4, and NF-κB p65 proteins were increased in the lipopolysaccharide (LPS) group compared with the normal control group, and the relative expression of the above proteins was decreased after treatment with QUE and IRAK4-IN-4. The results of flow cytometry suggested that LPS increased the expression of CD86 and CD11c on the surface of DCs, and QUE and IRAK4-IN-4 decreased the expression of CD86 and CD11c induced by LPS. Molecular docking results showed that the binding sites of QUE and IRAK4 were stable, with the minimum binding energies comparable to that of IRAK4-IN-4. Conclusions: Quercetin may inhibit the activity and function of DCs through the TLR4/IRAK4/NF-κB signalling pathway, and IRAK4 may be its target.

LncRNA HOXC-AS1 promotes nasopharyngeal carcinoma (NPC) progression by sponging miR-4651 and subsequently upregulating FOXO6.

The incidence rate of nasopharyngeal carcinoma (NPC) is the highest among the malignant tumors of otorhinolaryngology, posing a huge burden to public health. Long noncoding RNAs (lncRNAs) exert an important role in tumorigenesis and the progression of various cancers. The present study found that HOXC-AS1 was highly expressed in NPC and in NPC cell lines, suggesting a critical role of HOXC-AS1 in NPC progression. In addition, the abundance of HOXC-AS1 was negatively correlated with the prognosis of NPC. To molecularly dissect the mechanism of HOXC-AS1 in NPC progression, we knocked down the expression of HOXC-AS1 in HNE1 and C666-1 cells. Then, we employed CCK8, colony-formation experiment and Transwell to investigate how the cell performed when HOXC-AS1 was knocked down. It could be observed that HOXC-AS1 knockdown decreases cell proliferation, migration and invasion, but induces cell apoptosis in NPC. We found that HOXC-AS1 could sponge miR-4651 subsequently binding FOXO6 and inhibiting its expression. Therefore, HOXC-AS1/miR-4651/FOXO6 may form a competing endogenous RNA (ceRNA) network that promotes NPC progression. In conclusion, our study demonstrates that HOXC-AS1 promotes NPC progression by sponging miR-4651 and regulating FOXO6 expression, thus providing potential pharmaceutical targets for developing new NPC treatments.

SPZ1 promotes deregulation of Bim to boost apoptosis resistance in colorectal cancer.

Colorectal cancer (CRC) is the third most common malignancies in adults. Similar to other solid tumors, CRC cells show increased proliferation and suppressed apoptosis during the development and progression of the disease. Previous studies have shown that a novel tumor oncogene, spermatogenic basic helix-loop-helix transcription factor zip 1 (SPZ1), can promote proliferation. However, it is unclear whether SPZ1 plays a role in suppressing apoptosis, and the molecular mechanism behind SPZ1's suppression of apoptosis in CRC remains unclear. Here, we found that silencing endogenous SPZ1 inhibits cell growth and induces apoptosis, and overexpression of SPZ1 promotes cell growth. These findings were corroborated by in vitro and in vivo studies. Interestingly, SPZ1 overexpressing cells were resistant to 5-fluorouracil, a drug commonly used to treat cancer. Moreover, knocking down SPZ1 led to the activation of caspase through the deregulation of Bim by ERK1/2, we found that CRC tissues had significantly higher SPZ1 and lower Bim expression, and SPZ1HBimL were associated with advanced clinical stage of CRC. Collectively, our findings demonstrate that SPZ1 contributes to tumor progression by limiting apoptosis. SPZ1 reduces apoptosis by altering the stability of Bim, suggesting SPZ1 may serve as a biomarker and therapeutic target for CRC.

Allergen-specific immune response suppresses interleukin 10 expression in B cells via increasing micro-RNA-17-92 cluster.

Interleukin (IL)-10-expressing B cells play a critical role in the immune homeostasis in the body; its regulation has not been fully understood. Micro-RNA (miR)-17-92 cluster has strong regulation in the immunity. This study tests a hypothesis that miR-17-92 cluster suppresses IL-10 expression in B cells. In this study, peripheral B cells were collected from patients with allergic rhinitis (AR). The B cells were treated with specific allergens, dust mite extracts, in the culture. The expressions of miR-17-92 cluster and IL-10 in the culture were assessed by real-time quantitative reverse transcription polymerase chain reaction and enzyme-linked immunosorbent assay. The results showed that the levels of miR-19a, but not the rest of the 5 members (miR-17, miR-18a, miR-19b, miR-20a, and miR-92a), were significantly higher in peripheral B cells from AR patients as than in B cells from healthy participants. Exposure of B cells from AR patients to specific allergen, dust mite extracts, significantly increased the levels if miR-19a and suppressed the expression of IL-10 in B cells. The levels of histone deacetylase 11 and acetylated H3K9 were higher, and the RNA polymerase II and c-Maf (the IL-10 transcription factor) were lower, at the IL-10 promoter locus. In conclusion, miR-19a mediates the allergen-specific immune response-decreased IL-10 expression in B cells.

Insulin-like growth factor-2 enhances functions of antigen (Ag)-specific regulatory B cells.

Regulatory B cells (Bregs) are important in immune regulation. The factors that regulate Breg functions are less clear. Insulin-like growth factor 2 (IGF2) is capable of inducing hematopoietic stem cell differentiation. This study aimed to investigate the role of IGF2 in the development of Bregs and the enhancement of their function. In this study, the expression of IGF1 receptor (IGF1R) and IGF2R in ovalbumin (OVA)-specific B cells (OVAsBCs) was assessed by real time RT-PCR and Western blotting. The release of interleukin (IL)-10 from OVAsBCs and OVAsBC proliferation were assessed by enzyme-linked immunoassay and proliferation assay. The role of IGF2 in enhancing the function of OVAsBCs was tested with an intestinal allergic inflammation mouse model. The results showed that OVAsBCs expressed high levels of IGF2R. Exposure to both IGF2 and a specific antigen (Ag), OVA, markedly enhanced the expression of IL-10 in OVAsBCs as well as enhanced the IL-10(+) OVAsBC proliferation. The concurrent exposure to IGF2 and specific Ag markedly induced the IL-10 promoter DNA demethylation via activating the STAT5 pathway. IGF2 also enhanced both the OVAsBC proliferation in vivo and the effect of Ag-specific immunotherapy on inhibiting allergic inflammation in the intestine. We conclude that OVAsBCs express high levels of IGF2R and that IGF2 increases the expression of IL-10 in OVAsBCs and enhances OVAsBC proliferation and the inhibitory effect on allergic inflammation.

PDGFR-ß-activated ACK1-AKT signaling promotes glioma tumorigenesis.

Aberrant PDGF-PDGFR signaling and its effects on downstream effectors have been implicated in glioma development. A crucial AKT regulator, ACK1 (TNK2) has been shown to be a downstream mediator of PDGF signaling; however, the exact underlying mechanisms in gliomas remain elusive. Here, we report that in glioma cells, PDGFR-ß activation enhanced the interaction between ACK1 and AKT, resulting in AKT activation. PDGF treatment consistently promoted the formation of complexes containing PDGFR-ß and ACK1. Mutational analysis suggested that Y635 of ACK1 is a PDGFR-ß phosphorylation site and that the ACK1 Y635F mutant abrogated the sequential activation of AKT. Moreover, PDK1 interacted with ACK1 during PDGF stimulation, which is required for the binding of ACK1 to PDGFR-ß. Further mutational analysis showed that T325 of ACK1 was crucial for the ACK1 and PDK1 interaction. ACK1 Y635F or T325A mutants abolished PDGFR-ß-induced AKT activation, the subsequent nuclear translocation of ß-catenin and the expression of cyclin D1. Glioma cell cycle progression, proliferation and tumorigenesis were accordingly blocked by ACK1 Y635F or T325A. In glioblastoma multiforme samples from 51 patients, increased ACK1 tyrosine phosphorylation correlated with upregulated PDGFR-ß activity and AKT activation. Taken together, our data demonstrate that ACK1 plays a pivotal role in PDGF-PDGFR-induced AKT signaling in glioma tumorigenesis. This knowledge contributes to our understanding of glioma progression and may facilitate the identification of novel therapeutic targets for future glioma treatment.

Insulin-like growth factor 2 enhances regulatory T-cell functions and suppresses food allergy in an experimental model.

BACKGROUND: The functions of regulatory T (Treg) cells are important in immunity, and the regulatory mechanisms of Treg cell activities are not fully understood yet. OBJECTIVES: We sought to investigate the role of insulin-like growth factor (IGF) 2 in the upregulation of Treg cell function. METHODS: The expression of insulin-like growth factor 2 receptor (IGF2R) on T cells was assessed by using flow cytometry. Treg cell functions were evaluated by assessing the suppressor effect on proliferation of other effector T (Teff) cells. The effect of IGF2 on regulating Treg cell functions were evaluated with a cell-culture model and a food allergy mouse model. RESULTS: Expression of IGF2R was observed in more than 90% of murine and human Treg cells but in less than 10% of effector CD4(+) T cells. Activation of IGF2R and T-cell receptor induced marked Treg cell proliferation and release of TGF-ß from Treg cells, which enhanced Treg cell immune suppressor effects on other Teff cell activities and allergic inflammation in the intestine. CONCLUSIONS: Activation of IGF2R enhances Treg cell functions in suppressing other Teff cell activities and inhibiting allergic inflammation in the intestine.

[Progress in dendritic cell-derived exosomes in allergic rhinitis].

Allergic rhinitis (AR), a common disease in otolaryngology, is a key risk factor for poorly controlled asthma and many complications, although it is not life-threatening. The negative impact of AR on social productive forces and human health is no less than that of asthma. Dendritic cells (DCs) play an important role in AR. In addition to sharing some of DC's biological characteristics, DCs-derived exosomes (DEXs) can promote the priming and activation of T cells and the maturation and differentiation of T helper type 2 (Th2) cells. Multiple signaling pathways in AR can be modulated by DEXs, which present allergens and participate in allergic immune responses. Anti-allergic drugs can be carried by DEXs to alleviate allergic airway inflammation and treat Th2-mediated AR effectively. Therefore, DEXs are crucial in the pathogenesis and treatment of AR.

Involvement and repair of epithelial barrier dysfunction in allergic diseases.

The epithelial barrier serves as a critical defense mechanism separating the human body from the external environment, fulfilling both physical and immune functions. This barrier plays a pivotal role in shielding the body from environmental risk factors such as allergens, pathogens, and pollutants. However, since the 19th century, the escalating threats posed by environmental pollution, global warming, heightened usage of industrial chemical products, and alterations in biodiversity have contributed to a noteworthy surge in allergic disease incidences. Notably, allergic diseases frequently exhibit dysfunction in the epithelial barrier. The proposed epithelial barrier hypothesis introduces a novel avenue for the prevention and treatment of allergic diseases. Despite increased attention to the role of barrier dysfunction in allergic disease development, numerous questions persist regarding the mechanisms underlying the disruption of normal barrier function. Consequently, this review aims to provide a comprehensive overview of the epithelial barrier's role in allergic diseases, encompassing influencing factors, assessment techniques, and repair methodologies. By doing so, it seeks to present innovative strategies for the prevention and treatment of allergic diseases.

Automated hearing screening for children: A pilot study in China.

OBJECTIVE: To establish the reliability and validity of an automated hearing screening test system for children. DESIGN: Cross-sectional within a comparative study of subjects. STUDY SAMPLE: Subjects were 325 first-grade and second-grade children (6-10 years old) from primary schools in Shenzhen, China. RESULTS: Using the conventional pure-tone screening test with the pass/refer criterion set as 25 dB HL, as the 'gold standard", the sensitivity and specificity of the automated hearing screening test was 0.63 and 0.82, respectively. No specific pattern in the failure rates was observed to relate to the students' grade. There was no statistically significant age effect or gender effect. CONCLUSIONS: The results suggest that with further improvement in terms of its sensitivity and specificity, it may be feasible to use the automated hearing screening test system to conduct routine school hearing screenings.

Identification of galactosamine-(N-acetyl)-6-sulfatase (GALNS) as a novel therapeutic target in progression of nasopharyngeal carcinoma.

Nasopharyngeal carcinoma (NPC) is a commonly diagnosed malignancy in southern China and southeast Asia. Previous studies have identified galactosamine-(N-acetyl)-6-sulfatase (GALNS) as a potential biomarker for multiple cancers. However, it is unknown whether GALNS plays a role in NPC development, and the underlying mechanisms remain unclear. In this study, we found that GALNS is overexpressed in NPC cell lines and tissues compared to the normal nasopharyngeal counterparts. Knocking down GALNS expression in the NPC cells significantly decreased their proliferation in vitro, and inhibited xenograft growth in a mouse model. Mechanistically, the anti-proliferative effect of GALNS silencing was the result of autophagy induction via the inhibition of PI3K-AKT-mTOR signaling pathway. Taken together, GALNS drives the progression of NPC via PI3K-AKT-mTOR signaling-mediated autophagy, and is therefore a promising therapeutic target.

The Role of Inflammation-Associated Factors in Head and Neck Squamous Cell Carcinoma.

Head and neck squamous cell carcinoma (HNSCC), which originates in the head or neck tissues, is characterized by high rates of recurrence and metastasis. Inflammation is important in HNSCC prognosis. Inflammatory cells and their secreted factors contribute to the various stages of HNSCC development through multiple mechanisms. In this review, the mechanisms through which inflammatory factors, signaling pathways, and cells contribute to the initiation and progression of HNSCC have been discussed in detail. Furthermore, the diagnostic and therapeutic potential of targeting inflammation in HNSCC has been discussed to gain new insights into improving patient prognosis.

Identification of alpha-linolenic acid as a broad-spectrum antiviral against zika, dengue, herpes simplex, influenza virus and SARS-CoV-2 infection.

Zika virus (ZIKV) has garnered global attention due to its association with severe congenital defects including microcephaly. However, there are no licensed vaccines or drugs against ZIKV infection. Pregnant women have the greatest need for treatment, making drug safety crucial. Alpha-linolenic acid (ALA), a polyunsaturated ω-3 fatty acid, has been used as a health-care product and dietary supplement due to its potential medicinal properties. Here, we demonstrated that ALA inhibits ZIKV infection in cells without loss of cell viability. Time-of-addition assay revealed that ALA interrupts the binding, adsorption, and entry stages of ZIKV replication cycle. The mechanism is probably that ALA disrupts membrane integrity of the virions to release ZIKV RNA, inhibiting viral infectivity. Further examination revealed that ALA inhibited DENV-2, HSV-1, influenza virus and SARS-CoV-2 infection dose-dependently. ALA is a promising broad-spectrum antiviral agent.

TOM40 regulates the progression of nasopharyngeal carcinoma through ROS-mediated AKT/mTOR and p53 signaling.

Nasopharyngeal carcinoma (NPC) is a prevalent cancer in Southern China, North Africa, and Southeast Asia. The translocase of the outer membrane (TOM) 40 is a transporter of mitochondrial proteins, and is involved in ovarian cancer cell growth. However, its role in the progression of NPC is still unclear. We found that TOM40 levels were upregulated in NPC tissues and multiple NPC cell lines. In addition, high TOM40 expression in the tumor tissues was associated with poor overall survival and disease specific survival. TOM40 knockdown in the NPC cell lines inhibited their proliferation in vitro and in vivo. Furthermore, TOM40 silencing also increased intracellular production of reactive oxygen species (ROS) and decreased mitochondrial membrane potential (MMP). Mechanistically, the anti-tumor effects of TOM40 silencing were dependent on the inhibition of AKT/mTOR signaling and activation of p53 signaling. To summarize, TOM40 mediates NPC progression through ROS-mediated AKT/mTOR and p53 signaling. Our findings highlight the potential of TOM40 as a therapeutic target for NPC.

Role of dendritic cell­derived exosomes in allergic rhinitis (Review).

Allergic rhinitis (AR) is a common pathological condition in otorhinolaryngology. Its prevalence has been increasing worldwide and is becoming a major burden to the world population. Dendritic cells (DCs) are typically activated and matured after capturing, phagocytosing, and processing allergens during the immunopathogenesis of AR. In addition, the process of DC activation and maturation is accompanied by the production of exosomes, which are cell­derived extracellular vesicles (EVs) that can carry proteins, lipids, nucleic acids, and other cargoes involved in intercellular communication and material transfer. In particular, DC­derived exosomes (Dex) can participate in allergic immune responses, where the biological substances carried by them can have potentially important implications for both the pathogenesis and treatment of AR. Dex can also be exploited to carry anti­allergy agents to effectively treat AR. This provides a novel method to explore the pathogenesis of and treatment strategies for AR further. Therefore, the present review focuses on the origin, composition, function, and biological characteristics of DCs, exosomes, and Dex, in addition to the possible relationship between Dex and AR.

Tolerogenic dendritic cells and TLR4/IRAK4/NF-κB signaling pathway in allergic rhinitis.

Dendritic cells (DCs), central participants in the allergic immune response, can capture and present allergens leading to allergic inflammation in the immunopathogenesis of allergic rhinitis (AR). In addition to initiating antigen-specific immune responses, DCs induce tolerance and modulate immune homeostasis. As a special type of DCs, tolerogenic DCs (tolDCs) achieve immune tolerance mainly by suppressing effector T cell responses and inducing regulatory T cells (Tregs). TolDCs suppress allergic inflammation by modulating immune tolerance, thereby reducing symptoms of AR. Activation of the TLR4/IRAK4/NF-κB signaling pathway contributes to the release of inflammatory cytokines, and inhibitors of this signaling pathway induce the production of tolDCs to alleviate allergic inflammatory responses. This review focuses on the relationship between tolDCs and TLR4/IRAK4/NF-κB signaling pathway with AR.

B cell immunity in allergic nasal mucosa induces T helper 2 cell differentiation.

BACKGROUND: The pathogenesis of allergic diseases is to be further understood. Recent studies indicate that B cells are involved in the immune regulation. The present study aimed to investigate the role of B cells in the initiation of skewed T helper (Th)2 polarization. METHODS: The surgically removed nasal mucosal specimens from 24 patients with allergic rhinitis (AR) and 22 patients with non-AR (nAR) were collected. B cells isolated from the AR nasal mucosa were characterized. The effect of B cells on inducing naïve CD4+ T cells to differentiate into Th2 cells was evaluated with a cell culture model. RESULTS: Abundant B cells were detected in the nasal mucosa of patients with AR, which also expressed high levels of T cell immunoglobulin mucin domain (TIM)4 and costimulatory molecules. High levels of Staphylococcal enterotoxin B (SEB) were detected in the AR nasal mucosa. Expression of TIM4 could be induced in naïve B cells in the presence of SEB in culture. TIM4+ B cells could induce naïve CD4+ T cells to differentiate into Th2 cells. CONCLUSIONS: TIM4+ B cells from AR nasal mucosa can induce skewed Th2 polarization. It may be a potential therapeutic target in the treatment of AR. B cells plays an important role in the initiation of Th2 polarization. KEY MESSAGES: • High frequency of B cells exists in nasal mucosa of allergic rhinitis • These B cells express high levels of TIM4 • TIM4+ B cells can initiate the skewed Th2 polarization.

Antigen-specific activities of CD8+ T cells in the nasal mucosa of patients with nasal allergy.

BACKGROUND: The prevalence of chronic rhinitis is increasing rapidly. Its pathogenesis is not fully understood but immune inflammation is one plausible causative factor. Antigen specific CD8+ T cells play a critical role in the induction of chronic inflammation. This study aims to investigate the role of antigen specific CD8+ T cells in the pathogenesis of chronic AR. METHODS: Nasal mucosal epithelial samples obtained by the surface of the nasal mucosaof patients with AR complicated with inferior turbinate hypertrophy. Exosomes were purified from the scratching samples and examined by immune gold electron microscopy. Cell culture models were employed to evaluate the effect of exosomes on modulating CD8+ T cell activity. RESULTS: Exosomes purified from patients with chronic AR carried microbial products, Staphylococcal enterotoxin B (SEB), and airborne antigen, Derp1. Dendritic cells pulsed by SEB/Derp1-carrying exosomes showed high levels of CD80, CD86 and the major histocompatibility class I (MHCI). Exosome-pulsed dendritic cells could induce naive CD3+ T cells to differentiate into CD8+ T cells. Upon exposure to a specific antigen, the CD8+ T cells released granzyme B and perforin and more than 30% antigen specific CD8+ T cells proliferated. CONCLUSIONS: Antigen specific CD8+ T cells play an important role in the pathogenesis of chronic AR complicated with inferior turbinate hypertrophy.

Research Advances in the Treatment of Allergic Rhinitis by Probiotics.

Allergic rhinitis (AR) impairs the quality of life of patients and reduces the efficiency of social work, it is an increasingly serious public medical and economic problem in the world. Conventional anti-allergic drugs for the treatment of allergic rhinitis (AR) can cause certain side effects, which limit the quality of life of patients. Therefore, it makes sense to look for other forms of treatment. Several studies in recent years have shown that probiotics have shown anti-allergic effects in various mouse and human studies. For example, the application of certain probiotic strains can effectively relieve the typical nasal and ocular symptoms of allergic rhinitis in children and adults, thereby improving the quality of life and work efficiency. At the same time, previous studies in humans and mice have found that probiotics can produce multiple effects, such as reduction of Th2 cell inflammatory factors and/or increase of Th1 cell inflammatory factors, changes in allergy-related immunoglobulins and cell migration, regulate Th1/Th2 balance or restore intestinal microbiota disturbance. For patients with limited activity or allergic rhinitis with more attacks and longer attack duration, oral probiotics have positive effects. The efficacy of probiotics in the prevention and treatment of allergic rhinitis is remarkable, but its specific mechanism needs further study. This review summarizes the research progress of probiotics in the treatment of allergic rhinitis in recent years.