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We aim to evaluate the clinical efficacy and safety of using laser and light combined with topical minoxidil for alopecia areata. We searched PubMed, Embase, Web of Science, the Cochrane Library, China National Knowledge Infrastructure (CNKI), China Biomedical Literature Database (CBM), VIP database, and Wanfang Data from their inception to September 18, 2022. The risk of bias of the included RCTs was assessed by the Cochrane Collaboration tool. RevMan 5.3 software and Stata 14.0 software were used to perform the statistical analysis. The GRADE system assessed the quality of evidence. Ten studies were enrolled finally. The results of the meta-analysis showed that compared with topical minoxidil alone, the 308-nm excimer laser/light or He-Ne laser combined with topical minoxidil could reduce the SALT (Severity of Alopecia Tool) score (MD= -5.88, 95% CI [-9.79, -1.98], P=0.003). Whether fractional CO2 laser (RR=1.29, 95% CI [1.14, 1.46], P<0.0001), 308-nm excimer laser/light (RR=1.32, 95% CI [1.12, 1.55], P=0.001), He-Ne laser (RR=1.69, 95% CI [1.07, 2.69], P=0.03), or NB-UVB (RR=1.35, 95% CI [1.07,1.70], P=0.01) combined with topical minoxidil may improve the treatment response rate, comparing with topical minoxidil only. The recurrence rate of laser and light combined with topical minoxidil was lower than that of the minoxidil alone group (RR=0.54, 95% CI [0.31, 0.93], P=0.03) when follow-up time was 1 year. In addition, the incidence of adverse events including irritant contact dermatitis, erythema, desquamation, pain, and pruritus was no significant difference between the two groups (RR=1.50, 95% CI [0.95, 2.36], P=0.08). The level of evidence for outcomes was classified as very low to moderate. Based on the available evidence, laser and light combined with topical minoxidil therapy may be effective and safe for alopecia areata. However, more high-quality trials are required for comprehensive analysis and further verification.
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Alopecia em Áreas , Minoxidil , Humanos , Minoxidil/uso terapêutico , Alopecia em Áreas/tratamento farmacológico , Alopecia em Áreas/radioterapia , Ensaios Clínicos Controlados Aleatórios como Assunto , Fototerapia , LasersRESUMO
The present work aimed to systematically identify the efficacy and safety of fractional carbon dioxide (CO2) laser plus hyaluronic acid (HA) dressing in dealing with facial atrophic acne scars. Randomized controlled trials (RCTs) concerning fractional CO2 laser in combination with HA dressing for treating atrophic acne scars were screened in 8 electronic databases (containing PubMed, Embase, the Cochrane Library, Web of Science, China National Knowledge Internet, Wanfang, Sinomed as well as VIP). Besides, for the purpose of evaluating the risk of bias of the enrolled RCTs, the Cochrane Collaboration tool was adopted. Statistical analysis was completed using Revman5.3 software and Stata 14.0 software. Meanwhile, the quality of evidence was assessed by the GRADE system. Finally, 6 studies involving 623 patients were enrolled. According to the findings in this study, compared with fractional CO2 laser alone, fractional CO2 laser therapy combined with HA dressing reduced the scores of ECCA (échelle d'évaluation clinique des cicatrices d'acné) grading scale (MD=-3.37,95% CI [-5.03, -1.70], P<0.0001), shortened the time of crust formation (MD=-0.42,95% CI [-0.80, -0.04], P=0.03) and the time of crust removal(MD=-1.31,95% CI [-1.67, -0.95], P<0.00001), enhanced patient satisfaction (RR=1.85, 95% CI [1.44, 2.38], P<0.00001). All the reported adverse events including hyperpigmentation, erythema, edema, mild itching, and slight burning pain were controllable. In addition, fractional CO2 laser combined with HA dressing therapy had a lower incidence of hyperpigmentation than fractional CO2 laser alone (RR=0.37, 95% CI [0.23, 0.61], P<0.0001). The level of evidence for outcomes was classified to be low to moderate. According to our findings, fractional CO2 laser combined with HA dressing is efficacious and safe option for facial atrophic acne scars. Nevertheless, more high-quality trials are required for further verification in the future.
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Acne Vulgar , Terapia a Laser , Humanos , Cicatriz/etiologia , Cicatriz/radioterapia , Ácido Hialurônico , Dióxido de Carbono , Ensaios Clínicos Controlados Aleatórios como Assunto , Bandagens , Acne Vulgar/complicações , Acne Vulgar/radioterapiaRESUMO
Sporothrix schenckii (S. schenckii), a ubiquitous thermally dimorphic fungus, is the etiological agent of sporotrichosis, affecting immunocompromised and immunocompetent individuals. Despite current antifungal regimens, sporotrichosis results in prolonged treatment and significant mortality rates in the immunosuppressed population. The innate immune system forms the host's first and primary line of defense against S. schenckii, which has a bi-layered cell wall structure. Many components act as pathogen-associated molecular patterns (PAMPs) in pathogen-host interactions. PAMPs are recognized by pattern recognition receptors (PRRs) such as toll-like receptors, C-type lectin receptors, and complement receptors, triggering innate immune cells such as neutrophils, macrophages, and dendritic cells to phagocytize or produce mediators, contributing to S. schenckii elimination. The ultrastructure of S. schenckii and pathogen-host interactions, including PRRs and innate immune cells, are summarized in this review, promoting a better understanding of the innate immune response to S. schenckii and aiding in the development of protective and therapeutic strategies to combat sporotrichosis.
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Sporothrix , Esporotricose , Humanos , Esporotricose/microbiologia , Moléculas com Motivos Associados a Patógenos , Imunidade Inata , MacrófagosRESUMO
Objective: The aim of this research was to determine the correction coefficients of different spot urinary iodine concentrations (UICs) in a day to predict the early morning UIC, to make the different spot UICs of a day comparable in assessing iodine status in the same population. Methods: In total, 424 participants aged 18 to 28 years were recruited from Tianjin, China. Three spot urine samples were collected from each participant during three periods of the day (6:30-7:00, 10:00-10:30, and 16:00-16:30). A total of 1272 urine samples were collected. A 24-hour dietary record was reported by each participant for 3 consecutive days. Results: Both the UICs at 10:00-10:30 and 16:00-16:30 were higher than that at 6:30-7:00 (181.75 or 198.15µg/L vs 157.69 µg/L; all p < 0.05). Bland-Altman plot showed no good agreements between the other two spot UICs and that at 6:30-7:00 with both Bland-Altman indexes of 7.1%. Correction coefficients used to predict UIC at 6:30-7:00 from the UIC at 10:00-10:30 and 16:00-16:30 were 0.9231 and 0.8592, respectively. The predicted UICs at 6:30-7:00 by using the UIC at 10:00-10:30 and 16:00-16:30 both had no statistically significant difference with the actual UIC at 6:30-7:00 (all p > 0.05). Bland-Altman plot showed good agreements between the predicted UICs and actual UICs at 6:30-7:00 with both Bland-Altman indexes of 4.5%. Conclusions: It was reliable to predict the early morning UIC using correction coefficients of other spot UIC of the day, which gave a new idea for standardizing the methodology of surveys assessing iodine status of a population.
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Povo Asiático , Iodo/deficiência , Iodo/urina , Estado Nutricional , Adolescente , Adulto , China , Feminino , Humanos , Masculino , Adulto JovemRESUMO
Anoctamin 6 (ANO6/TMEM16F) is a recently identified membrane protein that has both phospholipid scramblase activity and anion channel function activated by relatively high [Ca2+]i. In addition to the low sensitivity to Ca2+, the activation of ANO6 Cl- conductance is very slow (>3-5â¯min to reach peak level at 10⯵M [Ca2+]i), with subsequent inactivation. In a whole-cell patch clamp recording of ANO6 current (IANO6,w-c), disruption of the actin cytoskeleton with cytochalasin-D (cytoD) significantly accelerated the activation kinetics, while actin filament-stabilizing agents (phalloidin and jasplakinolide) commonly inhibited IANO6,w-c. Inside-out patch clamp recording of ANO6 (IANO6,i-o) showed immediate activation by raising [Ca2+]i. We also found that intracellular ATP (3â¯mM MgATP in pipette solution) decelerated the activation of IANO6,w-c, and also prevented the inactivation of IANO6,w-c. However, the addition of cytoD still accelerated both activation and inactivation of IANO6,w-c. We conclude that the actin cytoskeleton and intracellular ATP play major roles in the Ca2+-dependent activation and inactivation of IANO6,w-c, respectively.
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Citoesqueleto de Actina/metabolismo , Trifosfato de Adenosina/metabolismo , Anoctaminas/metabolismo , Cálcio/metabolismo , Proteínas de Transferência de Fosfolipídeos/metabolismo , Células HEK293 , Humanos , Técnicas de Patch-ClampRESUMO
OBJECTIVE: High salt intake among lactating women can increase the risk of hypertension and cardiovascular disease in infants/offspring. However, considering the limited salt intake data in lactating women, the aims of this study were to compare the salt intake assessed by modified food weighted records (FWR) with that estimated by 24-h urinary sodium excretion and to investigate the salt intake of lactating women. METHODS: In total, 30 lactating women aged 20-39 years who were 2 to 4 months postpartum were recruited from the cities of Tianjin and Luoyang in China. The household salt intakes of the lactating women were collected by modified FWR for 3 days. Information on the gender, age, eating behaviours and labour intensity of the family members and guests dining at home during the 3 days was recorded. Meanwhile, 24-h urine samples of lactating women were collected. RESULTS: The salt intakes of the lactating women estimated by modified FWR and 24-h urinary sodium excretion were 8.50 ± 5.32 g/d and 9.34±3.74 g/d (t=-1.29, P=0.207), respectively, which exceeded the WHO recommendation of 5 g/d. There was a significant correlation (r=0.628, P < 0.001) between the salt intakes assessed by the two methods. A Bland-Altman plot showed no significant mean difference between the two methods (salt intake measured by 24-h urinary sodium excretion-salt intake assessed by modified FWR=0.46 g/d, P=0.207). CONCLUSIONS: The modified FWR is a reliable tool to assess the salt intake of lactating women. The salt intake of lactating women in China remains higher than the WHO recommendation and should be restricted through further efforts.
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Lactação , Cloreto de Sódio na Dieta/administração & dosagem , Adulto , China , Registros de Dieta , Feminino , Humanos , Recomendações Nutricionais , Adulto JovemRESUMO
LPSs are widely used to stimulate TLR4, but their effects on ion channels in immune cells are poorly known. In THP-1 cells and human blood monocytes treated with LPS, inwardly rectifying K(+) channel current (IKir,LPS) newly emerged at 1 h, peaked at 4 h (-119 ± 8.6 pA/pF), and decayed afterward (-32 ± 6.7 pA/pF at 24 h). Whereas both the Kir2.1 and Kir2.2 mRNAs and proteins were observed, single-channel conductance (38 pS) of IKir,LPS and small interfering RNA-induced knockdown commonly indicated Kir2.2 than Kir2.1. LPS-induced cytokine release and store-operated Ca(2+) entry were commonly decreased by ML-133, a Kir2 inhibitor. Immunoblot, confocal microscopy, and the effects of vesicular trafficking inhibitors commonly suggested plasma membrane translocation of Kir2.2 by LPS. Both IKir,LPS and membrane translocation of Kir2.2 were inhibited by GF109203X (protein kinase C [PKC] inhibitor) or by transfection with small interfering RNA-specific PKCε. Interestingly, pharmacological activation of PKC by PMA induced both Kir2.1 and Kir2.2 currents. The spontaneously decayed IKir,LPS at 24 h was recovered by PI3K inhibitors but further suppressed by an inhibitor of phosphatidylinositol(3,4,5)-trisphosphate (PIP3) phosphatase (phosphatase and tensin homolog). However, IKir,LPS at 24 h was not affected by Akt inhibitors, suggesting that the decreased phosphatidylinositol(4,5)-bisphosphate availability, that is, conversion into PIP3 by PI3K, per se accounts for the decay of IKir,LPS. Taken together, to our knowledge these data are the first demonstrations that IKir is newly induced by TLR4 stimulation via PKC-dependent membrane trafficking of Kir2.2, and that conversion of phosphatidylinositol(4,5)-bisphosphate to PIP3 modulates Kir2.2. The augmentation of Ca(2+) influx and cytokine release suggests a physiological role for Kir2.2 in TLR4-stimulated monocytes.
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Monócitos/imunologia , Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Proteína Quinase C-épsilon/metabolismo , Receptor 4 Toll-Like/metabolismo , Cálcio/metabolismo , Linhagem Celular , Humanos , Imidazóis/farmacologia , Indóis/farmacologia , Lipopolissacarídeos , Maleimidas/farmacologia , Técnicas de Patch-Clamp , Fenantrolinas/farmacologia , Fosfatidilinositol 3-Quinases/metabolismo , Fosfatidilinositol 4,5-Difosfato/metabolismo , Fosfoinositídeo Fosfolipase C/metabolismo , Inibidores de Fosfoinositídeo-3 Quinase , Monoéster Fosfórico Hidrolases/antagonistas & inibidores , Canais de Potássio Corretores do Fluxo de Internalização/antagonistas & inibidores , Canais de Potássio Corretores do Fluxo de Internalização/genética , Proteína Quinase C-épsilon/antagonistas & inibidores , Proteína Quinase C-épsilon/genética , Transporte Proteico , Interferência de RNA , RNA Mensageiro/genética , RNA Interferente Pequeno , Transdução de SinaisRESUMO
The general consensus is that immune cells are exposed to physiological hypoxia in vivo (PhyO2, 2-5% P(O2)). However, functional studies of B cells in hypoxic conditions are sparse. Recently, we reported the expression in mouse B cells of TASK-2, a member of pH-sensitive two-pore domain K(+) channels with background activity. In this study, we investigated the response of K(+) channels to sustained PhyO2 (sustained hypoxia [SH], 3% P(O2) for 24 h) in WEHI-231 mouse B cells. SH induced voltage-independent background K(+) conductance (SH-K(bg)) and hyperpolarized the membrane potential. The pH sensitivity and the single-channel conductance of SH-K(bg) were consistent with those of TASK-2. Immunoblotting assay results showed that SH significantly increased plasma membrane expressions of TASK-2. Conversely, SH failed to induce any current following small interfering (si)TASK-2 transfection. Similar hypoxic upregulation of TASK-2 was also observed in splenic primary B cells. Mechanistically, upregulation of TASK-2 by SH was prevented by si hypoxia-inducible factor-1α (HIF-1α) transfection or by YC-1, a pharmacological HIF-1α inhibitor. In addition, TASK-2 current was increased in WEHI-231 cells overexpressed with O2-resistant HIF-1α. Importantly, [Ca(2+)]c increment in response to BCR stimulation was significantly higher in SH-exposed B cells, which was abolished by high K(+)-induced depolarization or by siTASK-2 transfection. The data demonstrate that TASK-2 is upregulated under hypoxia via HIF-1α-dependent manner in B cells. This is functionally important in maintaining the negative membrane potential and providing electrical driving force to control Ca(2+) influx.
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Linfócitos B/metabolismo , Sinalização do Cálcio , Cálcio/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Oxigênio/farmacologia , Canais de Potássio de Domínios Poros em Tandem/metabolismo , Animais , Linfócitos B/citologia , Linfócitos B/efeitos dos fármacos , Hipóxia Celular , Linhagem Celular , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Regulação da Expressão Gênica , Subunidade alfa do Fator 1 Induzível por Hipóxia/antagonistas & inibidores , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Indazóis/farmacologia , Potenciais da Membrana/efeitos dos fármacos , Camundongos , Canais de Potássio de Domínios Poros em Tandem/antagonistas & inibidores , Canais de Potássio de Domínios Poros em Tandem/genética , Cultura Primária de Células , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismoRESUMO
BACKGROUND: Biologics are essential in treating psoriasis. In recent years, the pathogenesis exploration and development of new target drugs have provided a more complete evidence-based foundation for the biological treatment of psoriasis. This study aims to use bibliometrics to analyze the research status and development trends of biologics in psoriasis. METHODS: The bibliometric analysis of publications related to biologics in psoriasis from 2004 to 2023 was conducted using the Web of Science Core Collection (WoSCC) database as the search data source. To perform the bibliometric analysis and create visual knowledge graphs, CiteSpace, the Bibliometrix R package, and VOSviewers were utilized. RESULTS: The study included a total of 3800 articles. The United States had the highest number of publications. The leading authors and institutions were Steven R. Feldman and the University of Manchester, respectively, in the global partnership. The cluster plot divided all keywords into 11 categories. Currently, Secukinumab and Guselkumab are representative biological agents being studied due to their considerable efficacy and long-term safety. CONCLUSIONS: Targeted therapy has emerged as a significant trend in the current treatment of psoriasis. Early and active use of biologics can effectively control disease progression, prevent or delay the occurrence of comorbidities, and may even alter the natural course of psoriasis. However, further investigation is required to fully understand the specific mechanisms of psoriasis and the use of biological agents.
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Bibliometria , Psoríase , Psoríase/tratamento farmacológico , Humanos , Produtos Biológicos/uso terapêutico , Anticorpos Monoclonais Humanizados/uso terapêutico , Fármacos Dermatológicos/uso terapêutico , Pesquisa BiomédicaRESUMO
Background: It is now understood that T cells play a key role in the occurrence and development of psoriasis. Herein, a bibliometric analysis was conducted to summarize the content and trends of T cell-related research in psoriasis. Methods: A bibliometric analysis was conducted on publications pertaining to T cells in psoriasis between 2003 and 2022 retrieved from the Web of Science Core Collection (WoSCC) database using tools such as CiteSpace, the Bibliometrix R package, and VOSviewer. Results: The study included a total of 3595 articles authored by 14,188 individuals, including all coauthors in article bylines. The Laboratory for Investigative Dermatology at Rockefeller University, led by James G Krueger, has made significant contributions to this field through focusing on the pathogenesis of psoriasis and exploring the potential of using biological agents to treat psoriasis. Furthermore, targeted inhibitors have significantly impacted the treatment of psoriasis, with researchers focusing on small-molecule targeted drugs as a new area of research that could potentially replace biological agents. Conclusions: Research has established the efficacy and long-term safety of targeted inhibition of T cell-related targets. Deucravacitinib, a psoriasis treatment drug targeting TYK2 as an allosteric inhibitor, has attracted significant attention and raised high expectations.
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Objective: To explore the validation of a disease-free survival (DFS) model for predicting disease progression based on the combination of ubiquitin-conjugating enzyme E2 C (UBE2C) levels and clinical indicators in breast cancer patients. Methods: We enrolled 121 patients with breast cancer, collected their baseline characteristics and follow-up data, and analyzed the UBE2C levels in tumor tissues. We studied the relationship between UBE2C expression in tumor tissues and disease progression events of patients. We used the Kaplan-Meier method for identifying the disease-free survival rate of patients, and the multivariate Cox regression analysis to study the risk factors affecting the prognosis of patients. We sought to develop and validate a model for predicting disease progression. Results: We found that the level of expression of UBE2C could effectively distinguish the prognosis of patients. In the Receiver Operating Characteristic (ROC) curve analysis, the Area under the ROC Curve (AUC) = 0.826 (0.714-0.938) indicating that high levels of UBE2C was a high-risk factor for poor prognosis. After evaluating different models using the ROC curve, Concordance index (C-index), calibration curve, Net Reclassification Index (NRI), Integrated Discrimination Improvement Index (IDI), and other methods, we finally developed a model for the expression of Tumor-Node (TN) staging using Ki-67 and UBE2C, which had an AUC=0.870, 95% CI of 0.786-0.953. The traditional TN model had an AUC=0.717, and 95% CI of 0.581-0.853. Decision Curve Analysis (DCA) and Clinical Impact Curve (CIC) analysis indicated that the model had good clinical benefits and it was relatively simple to use. Conclusion: We found that high levels of UBE2C was a high-risk factor for poor prognosis. The use of UBE2C in addition to other breast cancer-related indicators effectively predicted the possible disease progression, thus providing a reliable basis for clinical decision-making.
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BACKGROUND: Several prognostic algorithms were specifically or nonspecifically used for papillary renal cell carcinoma (PRCC). No consensus was reached upon their efficacy of discrimination. We aim to compare the stratifying ability of current models or systems in predicting the risk of recurrence of PRCC. METHODS: A PRCC cohort consisting of 308 patients from our institution and 279 patients from The Cancer Genome Atlas (TCGA) was generated. With ISUP grade, TNM classification, UCLA Integrated Staging System (UISS), STAGE, SIZE, GRADE AND NECROSIS (SSIGN), Leibovich model and VENUSS system, recurrence-free survival (RFS), disease-specific survival (DSS) and overall survival (OS) were studied using Kaplan-Meier method and concordance index (c-index) was compared. Differences between risk groups in gene mutation and infiltration of inhibitory immune cells were studied with TCGA database. RESULTS: All the algorithms were able to stratify patients in RFS as well as DSS and OS (all P < 0.001). VENUSS score and risk group generally had the highest and balanced c-index (0.815 and 0.797 for RFS). ISUP grade, TNM stage and Leibovich model had the lowest c-indexes in all analysis. Among the 25 most frequently mutated genes in PRCC, eight had different mutation frequency between VENUSS low- and intermediate-/high-risk patients and mutated KMT2D and PBRM1 resulted in worsened RFS (Pâ¯=â¯0.053 and Pâ¯=â¯0.007). Increased Treg cells in tumors of intermediate-/high- risk patients were also identified. CONCLUSIONS: VENUSS system showed better predictive accuracy in RFS, DSS and OS compared with SSIGN, UISS and Leibovich risk models. VENUSS intermediate-/high-risk patients had increased frequency of mutation in KMT2D and PBRM1 and increased infiltration of Treg cells.
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Carcinoma de Células Renais , Neoplasias Renais , Humanos , Carcinoma de Células Renais/patologia , Neoplasias Renais/patologia , Prognóstico , Estadiamento de Neoplasias , Fatores de Risco , Estudos RetrospectivosRESUMO
Cells are exposed to various pathological stimulus within the cardiovascular system that challenge cells to adapt and survive. Several of these pathological stimulus alter the normal function of the endoplasmic reticulum (ER), leading to the accumulation of unfolded and misfolded proteins, thus triggering the unfolded protein response (UPR) to cope with the stress or trigger apoptosis of damaged cells. Downstream components of the UPR regulate transcription and translation reprogramming to ensure selective gene expression in response to pathological stimulus, including the expression of non-coding RNAs (ncRNAs). The ncRNAs play crucial roles in regulating transcription and translation, and their aberrant expression is associated with the development of cardiovascular disease (CVD). Notably, ncRNAs and ER stress can modulate each other and synergistically affect the development of CVD. Therefore, studying the interaction between ER stress and ncRNAs is necessary for effective prevention and treatment of CVD. In this review, we discuss the UPR signaling pathway and ncRNAs followed by the interplay regulation of ER stress and ncRNAs in CVD, which provides further insights into the understanding of the pathogenesis of CVD and therapeutic strategies. This article is categorized under: RNA in Disease and Development > RNA in Disease.
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Doenças Cardiovasculares , Humanos , Doenças Cardiovasculares/metabolismo , Estresse do Retículo Endoplasmático/genética , Resposta a Proteínas não Dobradas , Retículo Endoplasmático/metabolismo , Retículo Endoplasmático/patologia , RNA não Traduzido/genética , RNA não Traduzido/metabolismoRESUMO
Background: Astragalus mongholicus polysaccharides (APS) have anti-inflammatory, antioxidant and immunomodulatory effects. Recent studies have demonstrated the epigenetic regulation of N6-methyladenosine (m6A) in the development of inflammation. However, the effect of APS on m6A modification is unclear. Here, for the first time, we investigate the mechanism of m6A modification in APS regulation of THP-1 macrophage inflammation. Methods: We treated LPS-induced THP-1 macrophages with APS at different concentrations and times, and detected IL-6 mRNA and protein levels by quantitative real-time PCR (qRT-PCR) and western blot, respectively. The m6A modification level was detected by m6A quantification kit. The proteins that regulate m6A modification were screened by western blot. Wilms' tumor 1-associating protein (WTAP) was overexpressed in APS-treated THP-1 macrophages and the m6A modification level and IL-6 expressions were detected. Results: These findings confirmed that APS significantly abolished LPS-induced IL-6 levels in THP-1 macrophages. Meanwhile, APS reduced m6A modification levels and WTAP gene expression in THP-1 macrophages. Further overexpression of WTAP can significantly reverse APS-induced m6A modification level and IL-6 expression. Mechanistically, APS regulates IL-6 expression through WTAP-mediated p65 nuclear translocation. Conclusion: Overall, our study suggested that WTAP mediates the anti-inflammatory effect of APS by regulating m6A modification levels in THP-1 macrophages. This study reveals a new dimension of APS regulation of inflammation at the epigenetic level.
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Outer hair cell (OHC) degeneration is a major cause of progressive hearing loss and presbycusis. Despite the high prevalence of these disorders, targeted therapy is currently not available. Methods: We generated a mouse model harboring Kcnq4W276S/+ to recapitulate DFNA2, a common genetic form of progressive hearing loss accompanied by OHC degeneration. After comprehensive optimization of guide RNAs, Cas9s, vehicles, and delivery routes, we applied in vivo gene editing strategy to disrupt the dominant-negative allele in Kcnq4 and prevent progressive hearing loss. Results:In vivo gene editing using a dual adeno-associated virus package targeting OHCs significantly improved auditory thresholds in auditory brainstem response and distortion-product otoacoustic emission. In addition, we developed a new live-cell imaging technique using thallium ions to investigate the membrane potential of OHCs and successfully demonstrated that mutant allele disruption resulted in more hyperpolarized OHCs, indicating elevated KCNQ4 channel activity. Conclusion: These findings can facilitate the development of targeted therapies for DFNA2 and support the use of CRISPR-based gene therapy to rectify defects in OHCs.
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Edição de Genes , Perda Auditiva , Animais , Modelos Animais de Doenças , Células Ciliadas Auditivas Externas/metabolismo , Perda Auditiva/genética , Perda Auditiva/metabolismo , Perda Auditiva/terapia , Canais de Potássio KCNQ/genética , Canais de Potássio KCNQ/metabolismo , Potenciais da Membrana , CamundongosRESUMO
Anoctamin-6 (ANO6) belongs to a family of calcium (Ca2+)-activated chloride channels (CaCCs), with three splicing variants (V1, V2, and V5) showing plasma membrane expression. Unlike other CaCCs, ANO6 requires a non-physiological intracellular free calcium concentration ([Ca2+]i > 1 µM) and several minutes for full activation under a whole-cell patch clamp. Therefore, its physiological role as an ion channel is uncertain and it is more commonly considered a Ca2+-dependent phospholipid scramblase. Here, we demonstrate that physiological temperature (37 °C) increases ANO6 Ca2+ sensitivity under a whole-cell patch clamp; V1 was activated by 1 µM [Ca2+]i, whereas V2 and V5 were activated by 300 nM [Ca2+]i. Increasing the temperature to 42 °C led to activation of all ANO6 variants by 100 nM [Ca2+]i. The delay time for activation of the three variants was significantly shortened at 37 °C. Notably, the temperature-dependent Ca2+-sensitisation of ANO6 became insignificant under inside-out patch clamp, suggesting critical roles of unknown cytosolic factors. Unlike channel activity, 27 °C but not 37 °C (physiological temperature) induced the scramblase activity of ANO6 at submicromolar [Ca2+]i (300 nM), irrespective of variant type. Our results reveal a physiological ion conducting property of ANO6 at 37 °C and suggest that ANO6 channel function acts separately from its scramblase activity.
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Anoctaminas/metabolismo , Cálcio/metabolismo , Proteínas de Transferência de Fosfolipídeos/metabolismo , Anoctaminas/genética , Cálcio/farmacologia , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Células HEK293 , Humanos , Técnicas de Patch-Clamp , Fosfolipases A2/metabolismo , Proteínas de Transferência de Fosfolipídeos/genética , TemperaturaRESUMO
KCNQ4 is frequently mutated in autosomal dominant non-syndromic hearing loss (NSHL), a typically late-onset, initially high-frequency loss that progresses over time (DFNA2). Most KCNQ4 mutations linked to hearing loss are clustered around the pore region of the protein and lead to loss of KCNQ4-mediated potassium currents. To understand the contribution of KCNQ4 variants to NSHL, we surveyed public databases and found 17 loss-of-function and six missense KCNQ4 variants affecting amino acids around the pore region. The missense variants have not been reported as pathogenic and are present at a low frequency (minor allele frequency < 0.0005) in the population. We examined the functional impact of these variants, which, interestingly, induced a reduction in potassium channel activity without altering expression or trafficking of the channel protein, being functionally similar to DFNA2-associated KCNQ4 mutations. Therefore, these variants may be risk factors for late-onset hearing loss, and individuals harboring any one of these variants may develop hearing loss during adulthood. Reduced channel activity could be rescued by KCNQ activators, suggesting the possibility of medical intervention. These findings indicate that KCNQ4 variants may contribute more to late-onset NSHL than expected, and therefore, genetic screening for this gene is important for the prevention and treatment of NSHL.
Assuntos
Bases de Dados Genéticas , Perda Auditiva/genética , Ativação do Canal Iônico/genética , Canais de Potássio KCNQ/genética , Mutação , Animais , Células CHO , Cricetinae , Cricetulus , Surdez/genética , Surdez/fisiopatologia , Frequência do Gene , Células HEK293 , Audição/genética , Perda Auditiva/fisiopatologia , Humanos , Ativação do Canal Iônico/fisiologia , Setor PúblicoRESUMO
MK801 and ketamine, which are phencyclidine (PCP) derivative N-methyl-d-aspartate receptor (NMDAr) blockers, reportedly enhance the function of 5-hydroxytryptamine (HT)-2A receptors (5-HT2ARs). Both are believed to directly affect the pathogenesis of schizophrenia, as well as hypertension. 5-HT2AR signaling involves the inhibition of Kv conductance. This study investigated the interaction of these drugs with Kv1.5, which plays important roles in 5-HT2AR signaling and in regulating the excitability of the cardiovascular and nervous system, and the potential role of this interaction in the enhancement of the 5-HT2AR-mediated response. Using isometric organ bath experiments with arterial rings and conventional whole-cell patch-clamp recording of Chinese hamster ovary (CHO) cells ectopically overexpressing Kv1.5, we examined the effect of ketamine and MK801 on 5-HT2AR-mediated vasocontraction and Kv1.5 channels. Both ketamine and MK801 potentiated 5-HT2AR-mediated vasocontraction. This potentiation of 5-HT2AR function occurred in a membrane potential-dependent manner, indicating the involvement of ion channel(s). Both ketamine and MK801 rapidly and directly inhibited Kv1.5 channels from the extracellular side independently of NMDArs. The potencies of MK801 in facilitating the 5-HT2AR-mediated response and blocking Kv1.5 were higher than those of ketamine. Our data demonstrated the direct inhibition of Kv1.5 channels by MK801/ketamine and indicated that this inhibition may potentiate the functions of 5-HT2ARs. We suggest that 5-HT2AR-Kv1.5 may serve as a receptor-effector module in response to 5-HT and is a promising target in the pathogenesis of MK801-/ketamine-induced disease states such as hypertension and schizophrenia.
Assuntos
Maleato de Dizocilpina/farmacologia , Ketamina/farmacologia , Canal de Potássio Kv1.5/antagonistas & inibidores , Bloqueadores dos Canais de Potássio/farmacologia , Agonistas do Receptor 5-HT2 de Serotonina/farmacologia , Animais , Células CHO , Cricetulus , Hipertensão/metabolismo , Canal de Potássio Kv1.5/metabolismo , Masculino , Técnicas de Patch-Clamp , Ratos Sprague-Dawley , Receptor 5-HT2A de Serotonina/metabolismo , Esquizofrenia/metabolismo , Vasoconstritores/farmacologiaRESUMO
Arterioles and small arteries change their tone in response to transmural pressure changes, called myogenic tone (MT). In comparison to the branches of cerebral arteries (CA) showing prominent MT, the third branches of mesenteric arteries (MA) with similar diameters show weaker MT Here, we aimed to analyze the electrophysiological differences responsible for the weaker MT in MA (MTMA) than MT in CA (MTCA). We measured ionic current using patch clamp in isolated MA smooth muscle cells (MASMCs) and CA smooth muscle cells (CASMCs) of rats. MT was analyzed using video analysis of pressurized small arteries. Quantitative-PCR (q-PCR) and immunofluorescence confocal microscopy were used to compare the mRNA and protein expression level of big-conductance Ca2+-activated K+ channel (BKCa) subunits (Slo1α and Sloß1). Whole-cell patch clamp study revealed higher density of voltage-operated Ca2+ channel current (ICaV) in the MASMCs than in CASMCs. Although voltage-gated K+ channel current (IKv) was also higher in MASMCs, treatment with Kv inhibitor (4-aminopyridine) did not affect MTMA Interestingly, BKCa current density and the frequency of spontaneous transient outward currents (STOCs) were consistently higher in MASMCs than in CASMCs. Inside-out patch clamp showed that the Ca2+-sensitivity of BKCa is higher in MASMCs than in CASMCs. Iberiotoxin, a selective BKCa inhibitor, augmented MTMA by a larger extent than MTCA Although q-PCR analysis did not reveal a significant difference of mRNAs for Slo1α and Sloß1, immunofluorescence image suggested higher expression of Slo1α in MASMCs than in CASMCs. Despite the large ICaV density, the high activities of BKCa including the more frequent STOCs in MASMCs veils the potentially strong MTMA.
Assuntos
Canais de Potássio Ativados por Cálcio de Condutância Alta/metabolismo , Artérias Mesentéricas/metabolismo , Tono Muscular , Miócitos de Músculo Liso/metabolismo , Potenciais de Ação , Animais , Células Cultivadas , Artérias Cerebrais/citologia , Artérias Cerebrais/metabolismo , Artérias Cerebrais/fisiologia , Canais de Potássio Ativados por Cálcio de Condutância Alta/antagonistas & inibidores , Canais de Potássio Ativados por Cálcio de Condutância Alta/genética , Masculino , Artérias Mesentéricas/citologia , Artérias Mesentéricas/fisiologia , Miócitos de Músculo Liso/fisiologia , Bloqueadores dos Canais de Potássio/farmacologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-DawleyRESUMO
The voltage-gated potassium channel, Kv1.3, and the Ca2+-activated potassium channel, KCa3.1, regulate membrane potentials in T cells, thereby controlling T cell activation and cytokine production. However, little is known about the expression and function of potassium channels in human effector memory (EM) CD8+ T cells that can be further divided into functionally distinct subsets based on the expression of the interleukin (IL)-7 receptor alpha (IL-7Rα) chain. Herein, we investigated the functional expression and roles of Kv1.3 and KCa3.1 in EM CD8+ T cells that express high or low levels of the IL-7 receptor alpha chain (IL-7Rαhigh and IL-7Rαlow, respectively). In contrast to the significant activity of Kv1.3 and KCa3.1 in IL-7Rαhigh EM CD8+ T cells, IL-7Rαlow EM CD8+ T cells showed lower expression of Kv1.3 and insignificant expression of KCa3.1. Kv1.3 was involved in the modulation of cell proliferation and IL-2 production, whereas KCa3.1 affected the motility of EM CD8+ T cells. The lower motility of IL-7Rαlow EM CD8+ T cells was demonstrated using transendothelial migration and motility assays with intercellular adhesion molecule 1- and/or chemokine stromal cell-derived factor-1α-coated surfaces. Consistent with the lower migration property, IL-7Rαlow EM CD8+ T cells were found less frequently in human skin. Stimulating IL-7Rαlow EM CD8+ T cells with IL-2 or IL-15 increased their motility and recovery of KCa3.1 activity. Our findings demonstrate that Kv1.3 and KCa3.1 are differentially involved in the functions of EM CD8+ T cells. The weak expression of potassium channels in IL-7Rαlow EM CD8+ T cells can be revived by stimulation with IL-2 or IL-15, which restores the associated functions. This study suggests that IL-7Rαhigh EM CD8+ T cells with functional potassium channels may serve as a reservoir for effector CD8+ T cells during peripheral inflammation.