Missense mutation of ISL1 (E283D) is associated with the development of type 2 diabetes.

AIMS/HYPOTHESIS: Mutations in Isl1, encoding the insulin enhancer-binding protein islet-1 (ISL1), may contribute to attenuated insulin secretion in type 2 diabetes mellitus. We made an Isl1E283D mouse model to investigate the disease-causing mechanism of diabetes mellitus. METHODS: The ISL1E283D mutation (c. 849A>T) was identified by whole exome sequencing on an early-onset type 2 diabetes family and then the Isl1E283D knockin (KI) mouse model was created and an IPGTT and IPITT were conducted. Glucose-stimulated insulin secretion (GSIS), expression of Ins2 and other ISL1 target genes and interacting proteins were evaluated in isolated pancreas islets. Transcriptional activity of Isl1E283D was evaluated by cell-based luciferase reporter assay and electrophoretic mobility shift assay, and the expression levels of Ins2 driven by Isl1 wild-type (Isl1WT) and Isl1E283D mutation in rat INS-1 cells were determined by RT-PCR and western blotting. RESULTS: Impaired GSIS and elevated glucose level were observed in Isl1E283D KI mice while expression of Ins2 and other ISL1 target genes Mafa, Pdx1, Slc2a2 and the interacting protein NeuroD1 were downregulated in isolated islets. Transcriptional activity of the Isl1E283D mutation for Ins2 was reduced by 59.3%, and resulted in a marked downregulation of Ins2 expression when it was overexpressed in INS-1 cells, while overexpression of Isl1WT led to an upregulation of Ins2 expression. CONCLUSIONS/INTERPRETATION: Isl1E283D mutation reduces insulin expression and secretion by regulating insulin and other target genes, as well as its interacting proteins such as NeuroD1, leading to the development of glucose intolerance in the KI mice, which recapitulated the human diabetic phenotype. This study identified and highlighted the Isl1E283D mutation as a novel causative factor for type 2 diabetes, and suggested that targeting transcription factor ISL1 could offer an innovative avenue for the precise treatment of human type 2 diabetes.

Conductive Metal-Organic Framework Grown on the Nickel-Based Hydroxide to Realize High-Performance Electrochemical Glucose Sensing.

Glucose holds significant importance in disease diagnosis as well as beverage quality monitoring. The high-efficiency electrochemical sensor plays a crucial role in the electrochemical conversion technology. Ni(OH)2 nanosheets are provided with high specific surface area and redox activity that are widely used in electrochemistry. Conductive metal-organic frameworks (cMOFs) perfectly combine the structural controllability of organic materials with the long-range ordering of inorganic materials that possess the characteristic of high electron mobility. Based on the above considerations, the combination of Ni(OH)2 and Ni-HHTP (HHTP=2,3,6,7,10,11-hexahydroxytriphenylene) as an electrode modification material is designed to enhance electrochemical performance. In this work, to improve glucose detection, a sequence of Ni(OH)2@NiCo-HHTP and NiM-LDH@Ni-HHTP (M=Co2+, Mn2+, Cu2+, LDH=layered double hydroxide) are successfully synthesised by doping metals into Ni-HHTP and Ni(OH)2, respectively. As a result, NiCu-LDH@Ni-HHTP showed the best excellent glucose detection sensitivity.

Hf-Doped CoP Hollow Nanocubes as High-Performance Electrocatalyst for Oxygen Evolution Reaction.

Designing and synthesizing hollow frame structures with unique three-dimensional open structures in electrocatalysis remain a challenge. Etching is an effective method to synthesize metal-organic frameworks (MOFs) with a hollow structure and rich function. Herein, we report the design and synthesis of Hf-doped CoP hollow nanocubes by selective etching and ion exchange. Different from the traditional etching method, we used acid xylenol orange solution to etch typically the (211) crystal face of ZIF-67, obtaining the unique bell-like structure, named XO-ZIF-67. Subsequently, Hf-doped CoP hollow nanocubes were formed by Hf4+ doping and simple phosphating treatment. Electrochemical tests showed that the overpotential of the obtained catalyst is only 291 mV at the current density of 10 mA cm-2 when applied in catalyzing the oxygen evolution reaction (OER). Furthermore, the catalyst shows excellent stability when running in 1 M KOH solution for 25 h.

Successful In Situ Growth of Conductive MOFs on 2D Cobalt-Based Compounds and Their Electrochemical Performance.

Conductive metal-organic frameworks (cMOFs), as a kind of porous material, are considered to be highly promising materials in the field of electrochemistry due to their excellent conductivity. However, due to the low specific capacitance of pure cMOFs, their application in supercapacitors is limited. By virtue of the high theoretical capacity and excellent chemical stability of Co-based compounds, in this work, cMOFs' M-HHTP (M = Ni, Co, NiCo, HHTP = 2,3,6,7,10,11-hexahydroxytriphenylene) are grown in situ on Co(OH)2, CoP, and Co3O4 nanosheets, resulting in a series of electroactive compounds as electrode materials used in supercapacitors. Among all of the compounds, Ni-HHTP@Co(OH)2 shows the most excellent energy storage performance and outstanding cyclic stability in the application of aqueous asymmetric supercapacitors.

Research on the dentification of Panax ginseng and Panax quinquefolium using catalysed hairpin assembly technology.

INTRODUCTION: Panax ginseng and Panax quinquefolium are traditional Chinese herb medicines and similar in morphology and some chemical components but differ in drug properties, so they cannot be mixed. However, the processed products of them are often sold in the form of slices, powder, and capsules, which are difficult to identify by traditional morphological methods. Furthermore, an accurate evaluation of P. ginseng, P. quinquefolium and the processed products have not been conducted. OBJECTIVE: This study aimed to establish a catalysed hairpin assembly (CHA) identification method for authenticating products made from P. ginseng and P. quinquefolium based on single nucleotide polymorphism (SNP) differences. METHOD: By analysing the differences of SNP in internal transcribed spacer 2 (ITS2) in P. ginseng and P. quinquefolium to design CHA-specific hairpins. Establish a sensitive and efficient CHA method that can identify P. ginseng and P. quinquefolium, use the sequencing technology to verify the accuracy of this method in identifying Panax products, and compare this method with high-resolution melting (HRM). RESULTS: The reaction conditions of CHA were as follows: the ratio of forward and reverse primers, 20:1; hairpin concentration, 5 ng/µL. Compared with capillary electrophoresis, this method had good specificity and the limit of detection was 0.5 ng/µL. The result of Panax product identification with CHA method were coincidence with that of the sequencing method; the positive rate of CHA reaction was 100%. CONCLUSION: This research presents an effective identification method for authenticating P. ginseng and P. quinquefolium products, which is helpful to improve the quality of Panax products.

Spatial heterogeneity of tumor microenvironment influences the prognosis of clear cell renal cell carcinoma.

BACKGROUND: Clear cell renal cell carcinoma (ccRCC) is an immunologically and histologically diverse tumor. However, how the structural heterogeneity of tumor microenvironment (TME) affects cancer progression and treatment response remains unclear. Hence, we characterized the TME architectures of ccRCC tissues using imaging mass cytometry (IMC) and explored their associations with clinical outcome and therapeutic response. METHODS: Using IMC, we profiled the TME landscape of ccRCC and paracancerous tissue by measuring 17 markers involved in tissue architecture, immune cell and immune activation. In the ccRCC tissue, we identified distinct immune architectures of ccRCC tissue based on the mix score and performed cellular neighborhood (CN) analysis to subdivide TME phenotypes. Moreover, we assessed the relationship between the different TME phenotypes and ccRCC patient survival, clinical features and treatment response. RESULTS: We found that ccRCC tissues had higher levels of CD8+ T cells, CD163- macrophages, Treg cells, endothelial cells, and fibroblasts than paracancerous tissues. Immune infiltrates in ccRCC tissues distinctly showed clustered and scattered patterns. Within the clustered pattern, we identified two subtypes with different clinical outcomes based on CN analysis. The TLS-like phenotype had cell communities resembling tertiary lymphoid structures, characterized by cell-cell interactions of CD8+ T cells-B cells and GZMB+CD8+ T cells-B cells, which exhibited anti-tumor features and favorable outcomes, while the Macrophage/T-clustered phenotype with macrophage- or T cell-dominated cell communities had a poor prognosis. Patients with scattered immune architecture could be further divided into scattered-CN-hot and scattered-CN-cold phenotypes based on the presence or absence of immune CNs, but both had a better prognosis than the macrophage/T-clustered phenotype. We further analyzed the relationship between the TME phenotypes and treatment response in five metastatic ccRCC patients treated with sunitinib, and found that all three responders were scattered-CN-hot phenotype while both non-responders were macrophage/T-clustered phenotype. CONCLUSION: Our study revealed the structural heterogeneity of TME in ccRCC and its impact on clinical outcome and personalized treatment. These findings highlight the potential of IMC and CN analysis for characterizing TME structural units in cancer research.

Development and application of a visualization method for identification of Panax species with LAMP and a DNAzyme.

Panax ginseng and Panax quinquefolium are two valuable Chinese herbal medicines that should not be mixed because they differ in drug properties and efficacy. The traditional identification method is easily affected by subjective factors and cannot effectively distinguish between ginseng products. This study aimed to develop a new chemical analysis method to visually identify P. ginseng and P. quinquefolium. In this method, a large number of sequences containing G-quadruplex were generated by loop-mediated isothermal amplification, and the combination of G-quadruplex and hemin was used to form deoxyribozyme, which catalyzed the color change of H2O2. Artificial simulation of adulteration experiments revealed that this method could detect more than 20% adulterated P. quinquefolium. Compared with the traditional identification methods, this technology was simpler and more efficient, providing a reference for developing rapid visual identification methods and reagents for P. ginseng and P. quinquefolium.

Advances in ligase-based nucleic acid amplification technology for detecting gene mutations: a review.

Gene mutation has been a concern for researchers because it results in genetic variations with base changes in molecular structure. Researchers continue to explore methods to detect gene mutations, which may help in disease diagnosis, medication guidance, and so on. Currently, the detection methods, such as whole-genome sequencing and polymerase chain reaction, have some limitations in terms of cost and sensitivity. Ligase (an enzyme) can recognize base mismatch as a commonly used tool in genetic engineering. Therefore, the ligase-related nucleic acid amplification technology for detecting gene mutations has become a research hotspot. In this study, the main techniques explored for detecting gene mutations included the ligase detection reaction, ligase chain reaction, rolling circle amplification reaction, enzyme-assisted polymerase chain reaction, and loop-mediated isothermal amplification reaction. This review aimed to analyze the aforementioned techniques and mainly present their advantages and disadvantages, sensitivity, specificity, cost, detection time, applications, and so on. The findings may help develop sufficient grounds for further studies on detecting gene mutations.

Berberine inhibits gluconeogenesis in spontaneous diabetic rats by regulating the AKT/MAPK/NO/cGMP/PKG signaling pathway.

This work was aimed to investigate the action mechanism of berberine (BBR) on gluconeogenesis. The effects of BBR were examined in rat primary hepatocytes and confirmed in vivo in spontaneous diabetic rats. Protein levels were assessed by Western blot. Immunofluorescence staining was utilized for visualizing protein expression, while qRT-PCR helped for the determination of gene expression at the mRNA level. Besides, cGMP concentration was measured using ELISA, whereas NO level was assessed by spectrophotometry. BBR inhibited gluconeogenesis by downregulating G6Pase and PEPCK via inhibition of CREB phosphorylation. Moreover, BBR enhanced NO and cGMP concentrations, leading to the activation of the NO/cGMP/PKG signaling via activating AKT1/MAPK axis. The in vivo experiments were consistent with the findings obtained in vitro. Hence, BBR represents a drug candidate for diabetic patients and its mechanism of action may be driven via the AKT/MAPK/NO/cGMP/PKG pathway.

Sitagliptin Ameliorates Creb5/lncRNA ENSMUST00000213271-Mediated Vascular Endothelial Dysfunction in Obese Mice.

PURPOSE: Obesity is mediated by the changes in dyslipidemia, oxidative stress, and inflammation, leading to vascular endothelial dysfunction. Glucagon-like peptide-1 (GLP-1) analogues and dipeptidyl peptidase-4 inhibitors prevent the development of endothelial dysfunction. However, the underlying mechanism still remains largely unclear. Long non-coding RNAs (lncRNAs), one class of non-coding small RNAs, have been shown to exert a regulatory impact on the endothelial function in obesity. This study aimed to investigate whether the elevation of GLP-1 by a DPP-4 inhibitor sitagliptin improved vascular endothelial function by modulating lncRNAs in obese mice and to clarify the underlying molecular mechanism. METHODS: Male C57BL/6J mice were fed a high-fat diet for 4 months to induce obesity and some obese mice were treated with sitagliptin for the last 1 month. Levels of total cholesterol (TC), high-density lipoprotein (HDL), low-density lipoprotein (LDL), and glucagon-like peptide-1 (GLP-1) in plasma were detected by ELISA. LncRNA expression profile was analyzed via microarray. Aortic relaxations were examined by myograph. Protein expressions and phosphorylations were determined using western blot. The differentially expressed lncRNAs were validated using qRT-PCR. RESULTS: Obese mice exhibited increased levels of TC and LDL, decreased concentrations of HDL and GLP-1 in plasma, and impaired aortic endothelium-dependent relaxations; such effects could be reversed by sitagliptin. Moreover, the altered expression profile of lncRNAs in the obese mouse aortae could be modulated by sitagliptin. Consistent with microarray analysis, qRT-PCR also revealed that lncRNA ENSMUST00000213271 was up-regulated in obese mouse aortae and aortic endothelial cells (ECs), which could be down-regulated by sitagliptin. Creb5 silencing reduced lncRNA ENSMUST00000213271 in obese mouse ECs. Knockdown of either Creb5 or lncRNA ENSMUST00000213271 restored the activation of AMPK/eNOS in obese mouse ECs. Furthermore, sitagliptin also suppressed Creb5 and lncRNA ENSMUST00000213271 and increased the phosphorylations of AMPK and eNOS in obese mice. CONCLUSION: Creb5/lncRNA ENSMUST00000213271 mediated vascular endothelial dysfunction through inhibiting AMPK/eNOS cascade in obesity. Elevation of GLP-1 by sitagliptin possibly improved endothelial function by suppressing Creb5/lncRNA ENSMUST00000213271 and subsequently restoring AMPK/eNOS activation in obese mice. This study will provide new evidence for the benefits of GLP-1 against vasculopathy in obesity.

[Research Progress in the Regulation of Follicle Development by Melatonin].

Melatonin,an endocrine hormone synthesized by the pineal gland,plays an important role in the reproduction.The growth and development of follicles is the basis of female mammalian fertility.Follicles have a high concentration of melatonin.Melatonin receptors exist on ovarian granulosa cells,follicle cells,and oocytes.It regulates the growth and development of these cells and the maturation and atresia of follicles,affecting female fertility.This paper reviews the protective effects and regulatory mechanisms of melatonin on the development of ovarian follicles,granulosa cells,and oocytes and makes an outlook on the therapeutic potential of melatonin for ovarian injury,underpinning the clinical application of melatonin in the future.

Identification and management of GCK-MODY complicating pregnancy in Chinese patients with gestational diabetes.

Precise differentiation of glucokinase (GCK) monogenic diabetes from gestational diabetes mellitus (GDM) is critical for accurate management of the pregnancy outcome. We screened GCK-MODY complicating pregnancies in Chinese GDM patients, explored the pathogenesis of novel GCK mutations, and evaluated the patients' pregnancy outcome and management. The GCK gene from 411 GDM patients was screened with PCR-direct sequencing and multiplex ligation-dependent probe amplification (MLPA) and 15 GCK mutations were identified. We also retrospectively analyzed a total of 65 pregnancies from 21 GCK-MODY families, wherein 41 were from 15 maternal families and 24 were from six paternal families. Bioinformatic analysis and biochemical functional study were conducted to identify novel GCK mutations. In total, we identified 21 GCK mutations: 15 from the 411 GDM patients and six from 24 fathers. Of th Asp78Asn (GAC → AAC), Met87Arg (ATG → AGG), Leu451Val (CTT → GTT), Leu451Pro (CTG → CCG) and 1019 + 20G > A e mutations, five, i.e., were novel and deleterious, with markedly decreased enzyme activity and thermal stability. The unaffected offspring of GCK mutation-affected mothers were heavier than affected offspring (p < 0.001). Of 21 insulin-treated affected mothers, 10 had maternal hypoglycemia (47.6%) and seven had perinatal complications (33.3%), and the affected offspring of the insulin-treated affected mothers had significantly lower birth weights than that of the 20 diet-control affected mothers (p = 0.031). In this study, the prevalence of GCK-MODY complicating pregnancy in Chinese GDM patients was 3.6% (15/411). The defective GCK may contribute to the hyperglycemia in GCK-MODY. Insulin therapy is not beneficial for GCK-MODY complicating pregnancy and therefore should not be recommended.

MicroRNA expression is deregulated by aberrant methylation in B-cell acute lymphoblastic leukemia mouse model.

BACKGROUND: The expression of microRNAs (miRNAs) in the serum of B-cell acute lymphoblastic leukemia (B-ALL) patients is abnormal. Nevertheless, the underlying mechanism remains unclear. Recent studies indicate that the methylation state of circulating cell-free DNA (cfDNA) is different between cancer patients and healthy individuals. Therefore, we speculate that abnormal expression of miRNA may be associated with cfDNA methylation. METHODS: A green fluorescent protein (GFP) labeled B-ALL transplantation animal model was established to explore the relationship between the miRNA expression and cfDNA methylation of the related gene. Quantitative real-time PCR (qRT-PCR) was used to detect the expression levels of miRNAs. Further, cfDNA methylation levels of the related genes were evaluated through bisulfite sequencing polymerase chain reaction (BSP). RESULTS: The expression levels of miR-196b, miR-203, miR-34a-5p, miR-335-3p, miR-34b-5p, miR-615, miR-375-3p and miR-193b-5p in the serum of the model mice were significantly lower than those of the control group (P < 0.05). The methylation level of miR-196b promoter in cfDNA of the model group was significantly lower than that of the control group (P < 0.05), whereas no significant difference was noted in miR-203 promoter. The methylation levels of miR-196b and miR-203 coding region in cfDNA of the model group were significantly higher than those of the control group (P < 0.05). CONCLUSIONS: These results showed that CpG island hypermethylation in the miRNA coding region of cfDNA is related to the low expression of miR-196b and miR-203.

Profiling the lncRNA-miRNA-mRNA interaction network in the submandibular gland of diabetic mice.

BACKGROUND: Hyposalivation is one of the common symptoms of diabetes. Although long non-coding RNAs (lncRNAs) have recently been reported to play important roles in the pathogenesis of diabetes, the role of lncRNAs in diabetes-induced hyposalivation remains unknown. METHODS: The present study aimed to explore the function of lncRNA-microRNA-mRNA regulatory network in the submandibular gland (SMGs) under the context of diabetes. LncRNA expression profile of the SMGs was analyzed using microarray technology. Differentially expressed lncRNAs were confirmed using real-time quantitative PCR. Bioinformatics analyses were performed, and Coding-non-coding gene co-expression (CNC) and competing endogenous RNA (ceRNA) networks were constructed to explore the potential mechanisms of diabetes-induced hyposalivation. RESULTS: A total of 1273 differentially expressed lncRNAs (536 up-regulated and 737 downregulated) were identified in the SMGs tissues of db/db mice. CNC and ceRNA network analyses were performed based on five differentially expressed lncRNAs validated by real-time quantitative PCR. Gene Ontology analysis of target genes of CNC network revealed that "calcium ion binding" was a highly enriched molecular function. Kyoto Encyclopedia of Genes and Genomes pathway analysis of target genes of ceRNA network revealed that the "mammalian target of rapamycin signaling pathway" was significantly enriched. CONCLUSIONS: On the whole, the findings of the present study may provide insight into the possible mechanism of diabetes-induced hyposalivation.

Activation of the CaR-CSE/H2S pathway confers cardioprotection against ischemia-reperfusion injury.

Ischemia-reperfusion (I/R) injury is a multifactorial process triggered when an organ is subjected to transiently reduced blood supply. The result is a cascade of pathological complications and organ damage due to the production of reactive oxygen species following reperfusion. The present study aims to evaluate the role of activated calcium-sensing receptor (CaR)-cystathionine γ-lyase (CSE)/hydrogen sulfide (H2S) pathway in I/R injury. Firstly, an I/R rat model with CSE knockout was constructed. Transthoracic echocardiography, TTC and HE staining were performed to determine the cardiac function of rats following I/R Injury, followed by TUNEL staining observation on apoptosis. Besides, with the attempt to better elucidate how CaR-CSE/H2S affects I/R, in-vitro culture of human coronary artery endothelial cells (HCAECs) was conducted with gadolinium chloride (GdCl3, a CaR agonist), H2O2, siRNA against CSE (siCSE), or W7 (a CaM inhibitor). The interaction between CSE and CaM was subsequently detected. Plasma oxidative stress indexes, H2S and CSE, and apoptosis-related proteins were all analyzed following cell apoptosis. We found that H2S elevation led to the improvement whereas CSE knockdown decreased cardiac function in rats with I/R injury. Moreover, oxidative stress injury in I/R rats with CSE knockout was aggravated, while the increased expression of H2S and CSE in the aortic tissues resulted in alleviated the oxidative stress injury. Moreover, increased H2S and CSE levels were found to inhibit cell apoptotic ability in the aortic tissues after I/R injury, thus attenuating oxidative stress injury, accompanied by inhibited expression of apoptosis-related proteins. In HCAECs following oxidative stress treatment, siCSE and CaM inhibitor were observed to reverse the protection of CaR agonist. Coimmunoprecipitation assay revealed the interaction between CSE and CaM. Taken together, all above-mentioned data provides evidence that activation of the CaR-CSE/H2S pathway may confer a potent protective effect in cardiac I/R injury.

Assessment of Meibomian Gland (MD) Impairment Among Seasonal Allergic Conjunctivitis (SAC) Patients.

BACKGROUND Allergic conjunctivitis, one of the frequently occurring ocular surface diseases, can cause mucus discharge, itchy sensation, conjunctival hyperemia, and papillary formation. Seasonal allergic conjunctivitis (SAC) is associated with xerophthalmia and instability of tear film. Meibomian gland (MG) can secrete lipids to avoid xerophthalmia. However, there have been few reports on MG morphological alterations of SAC patients. This study aimed to examine the morphological alterations of MG among SAC patients. MATERIAL AND METHODS Our study included 89 eyes from 89 patients with SAC and 112 eyes of healthy volunteers. The symptoms were assessed by ocular surface disease index (OSDI) questionnaire. Then, the tests shown below were carried out, including tear evaporation rate from the ocular surface (TEROS), slit-lamp examination, break-up time (BUT) of tear film, Schirmer test I, vital staining, meibography, and meibum expression grading. MG was examined with laser scanning confocal microscopy (LSCM). RESULTS Relative to the control group, the OSDI was significantly higher in the SAC group. TEROS values, BUT, vital staining, MG expression, MG distortion rates, and MG dropout grades were significantly worse in the SAC group compared with the control group. As suggested by LSCM, SAC patients had markedly worse averages of parameters compared with controls. CONCLUSIONS The patients with SAC have more significant morphological and cytological changes in the MG. The Keratograph 5M system and LSCM are effective methods for evaluating MG status and ocular surface diseases.

Integrated analysis of lncRNA and mRNA expression profiles in the submandibular glands of DIO mice.

OBJECTIVE: Obesity contributes to the dysfunction of salivary gland. To explore the specific underlying mechanism for obesity-induced hyposalivation, a model for high-fat diet-induced obese (DIO) mice were constructed to analyze long non-coding RNAs (lncRNAs) and messenger RNAs (mRNAs) expression profiles. METHODS: The DIO group and control group were fed a diet containing 60 kcal% fat and a normal chow diet for 16 weeks respectively. Microarray analyses were performed to detect the expression profiles of lncRNA and mRNA in submandibular gland tissues from control group mice and DIO mice. Gene ontology, kyoto encyclopedia of genes and genomes, protein-protein interaction, coding-non-coding gene co-expression, transcription factors and competing endogenous RNA analyses were performed to examine the function of differentially expressed genes. RESULTS: Microarray analyses identified that 624 lncRNAs, along with 297 mRNAs were differentially expressed. Bioinformatic analyses revealed that "complement and coagulation cascades," "glutathione metabolism," "cysteine and methionine metabolism," and "estrogen signaling pathway" were significantly associated with candidate lncRNAs. Transcription factors analysis on candidate lncRNAs revealed several genes such as tribbles pseudokinase 3 may play regulatory roles. CONCLUSIONS: Our results revealed the expression profiles of lncRNAs and mRNAs and provided new insights into the mechanism of obesity-induced hyposalivation using bioinformatic analyses.

Prevalence and prognosis of otorhinolaryngological symptoms in patients with COVID-19: a systematic review and meta-analysis.

OBJECTIVE: A systematic review and meta-analysis were performed to evaluate the prevalence and prognosis of otorhinolaryngological symptoms in patients with the diagnosed coronavirus disease 2019 (COVID-19). METHODS: A systematic search of PubMed, Embase, Web of Science, and Google Scholar databases was performed up to August 19, 2020.We included studies that reported infections with COVID-19 and symptoms of otolaryngology. The retrieved data from the respective studies were evaluated and summarized. The study's immediate result was to assess the combined prevalence of otorhinolaryngological symptoms in patients with COVID-19. However, the secondary result was to determine the exacerbation of COVID-19 infection in patients with otorhinolaryngological symptoms. RESULTS: Fifty-four studies with 16,478 patients were included. Olfactory dysfunction, sneezing and sputum production were the 3 most prevalent otorhinolaryngological symptoms in patients with COVID-19. The pooled prevalence amongst the prevalent symptoms was 47% (95% CI 29-65; range 0-98; I2 = 99.58%), 27% (95% CI 11-48; range 12-40; I2 = 93.34%), and 22% (95% CI 16-30; range 2-56; I2 = 97.60%), respectively. The proportion of severely ill patients with sputum production and shortness of breath was significantly higher among patients with COVID-19 infections (OR 1.66 [95% CI 1.08-2.54]; P = 0.02, I2 = 51% and 3.29 [95% CI 1.57-6.90]; P = 0.002, I2 = 49%, respectively). Subgroup analysis showed no statistically significant differences between the incidence of otolaryngology symptoms in severely ill patients and non-severely ill patients (OR 1.43 [95% CI 1.12-1.82]; P = 0.07 I2 = 53.1%). In contrast, the incidence of shortness of breath in severely ill patients was significantly increased (3.29 [1.57-6.90]; P = 0.002, I2 = 49%). CONCLUSION: Our research shows that otorhinolaryngology symptoms in patients with COVID-19 are not uncommon, which should attract otorhinolaryngologists' attention.

Combination of puerarin and tanshinone IIA alleviates ischaemic stroke injury in rats via activating the Nrf2/ARE signalling pathway.

CONTEXT: Puerarin (Pue) and tanshinone IIA (Tan IIA) are often used in combination in the treatment of cerebrovascular diseases. OBJECTIVE: To investigate the neuroprotective effect and synergic mechanism of Pue-Tan IIA on the treatment of ischaemic stroke (IS). MATERIALS AND METHODS: IS was induced in rats by middle cerebral artery occlusion (MCAO). Rats were intraperitoneally injected with Pue (36 mg/kg), Tan IIA (7.2 mg/kg), or Pue-Tan IIA (36 and 7.2 mg/kg) for five times [30 min before ischaemia, immediately after reperfusion (0 h), 24, 48, and 72 h after reperfusion]. After administration, neurological function assessment and histological changes in the brain were performed. S-100ß and NSE levels were measured to determine the severity of brain injury. Oxidative stress parameters and inflammatory mediators were measured. The proteins involved in Nrf2/ARE signalling pathway were determined by qRT-PCR and western blot. RESULTS: After administration, the neurological function scores, infarct volume, S-100ß, and NSE levels were significantly reduced in MCAO rats, especially with Pue-Tan IIA treatment (p < 0.05). All treatments increased T-AOC, CAT, SOD, and GSH activities and reduced GSSG activity and MDA, TNF-α, IL-6, ICAM-1, and COX-2 levels in MCAO rats. Pue-Tan IIA significantly increased Nrf2 expression in the nucleus (1.81-fold) and decreased its expression in the cytoplasm (0.60-fold). Pue-Tan IIA significantly increased the expressions of HO-1 (1.87-fold) and NQO1 (1.76-fold) and decreased Keap1 expression (0.39-fold). DISCUSSION AND CONCLUSIONS: The combination of Pue and Tan IIA could alleviate ischaemic brain injury by activating Nrf2/ARE signalling pathway, providing an experimental basis for clinical applications.

Disruption of SIRT7 Increases the Efficacy of Checkpoint Inhibitor via MEF2D Regulation of Programmed Cell Death 1 Ligand 1 in Hepatocellular Carcinoma Cells.

BACKGROUND & AIMS: Immune checkpoint inhibitors have some efficacy in the treatment of hepatocellular carcinoma (HCC). Programmed cell death 1 ligand 1 (PD-L1), expressed on some cancer cells, binds to the receptor programmed cell death 1 (PDCD1, also called PD1) on T cells to prevent their proliferation and reduce the antigen-tumor immune response. Immune cells that infiltrate some types of HCCs secrete interferon gamma (IFNG). Some HCC cells express myocyte enhancer factor 2D (MEF2D), which has been associated with shorter survival times of patients. We studied whether HCC cell expression of MEF2D regulates expression of PD-L1 in response to IFNG. METHODS: We analyzed immune cells from 20 fresh HCC tissues by flow cytometry. We analyzed 225 fixed HCC tissues (from 2 cohorts) from patients in China by immunohistochemistry and obtained survival data. We created mice with liver-specific knockout of MEF2D (MEF2DLPC-KO mice). We knocked out or knocked down MEF2D, E1A binding protein p300 (p300), or sirtuin 7 (SIRT7) in SMMC-7721, Huh7, H22, and Hepa1-6 HCC cell lines, some incubated with IFNG. We analyzed liver tissues from mice and cell lines by RNA sequencing, immunoblot, dual luciferase reporter, and chromatin precipitation assays. MEF2D protein acetylation and proteins that interact with MEF2D were identified by coimmunoprecipitation and pull-down assays. H22 cells, with MEF2D knockout or without (controls), were transplanted into BALB/c mice, and some mice were given antibodies to deplete T cells. Mice bearing orthotopic tumors grown from HCC cells, with or without knockout of SIRT7, were given injections of an antibody against PD1. Growth of tumors was measured, and tumors were analyzed by immunohistochemistry and flow cytometry. RESULTS: In human HCC specimens, we found an inverse correlation between level of MEF2D and numbers of CD4+ and CD8+ T cells; level of MEF2D correlated with percentages of PD1-positive or TIM3-positive CD8+ T cells. Knockout of MEF2D from H22 cells reduced their growth as allograft tumors in immune-competent mice but not in immune-deficient mice or mice with depletion of CD8+ T cells. When MEF2D-knockout cells were injected into immune-competent mice, they formed smaller tumors that had increased infiltration and activation of T cells compared with control HCC cells. In human and mouse HCC cells, MEF2D knockdown or knockout reduced expression of PD-L1. MEF2D bound the promoter region of the CD274 gene (encodes PD-L1) and activated its transcription. Overexpression of p300 in HCC cells, or knockout of SIRT7, promoted acetylation of MEF2D and increased its binding, along with acetylated histones, to the promoter region of CD274. Exposure of HCC cells to IFNG induced expression of p300 and its binding MEF2D, which reduced the interaction between MEF2D and SIRT7. MEF2D-induced expression of PD-L1 upon IFNG exposure was independent of interferon-regulatory factors 1 or 9. In HCC cells not exposed to IFNG, SIRT7 formed a complex with MEF2D that attenuated expression of PD-L1. Knockout of SIRT7 reduced proliferation of HCC cells and growth of tumors in immune-deficient mice. Compared with allograft tumors grown from control HCC cells, in immune-competent mice, tumors grown from SIRT7-knockout HCC cells expressed higher levels of PD-L1 and had reduced infiltration and activation of T cells. In immune-competent mice given antibodies to PD1, allograft tumors grew more slowly from SIRT7-knockout HCC cells than from control HCC cells. CONCLUSIONS: Expression of MEF2D by HCC cells increases their expression of PD-L1, which prevents CD8+ T-cell-mediated antitumor immunity. When HCC cells are exposed to IFNG, p300 acetylates MEF2D, causing it to bind the CD274 gene promoter and up-regulate PD-L1 expression. In addition to promoting HCC cell proliferation, SIRT7 reduced acetylation of MEF2D and expression of PD-L1 in HCC cells not exposed to IFNG. Strategies to manipulate this pathway might increase the efficacy of immune therapies for HCC.