A Multiplex Fluorescence of Loop Primer Upon Self-Dequenching Loop-Mediated Isothermal Amplification Assay for the Detection of Epstein-Barr Virus and Human Parvovirus B19 in Clinical Transplant Samples.

Viral infections are major causes of mortality in solid-organ and hematopoietic stem cell transplant recipients. Epstein-Barr virus (EBV) and Parvovirus B19 (B19V) are among the common viral infections after transplantation and were recommended for increased screening in relevant guidelines. Therefore, the development of rapid, specific, and cost-effective diagnostic methods for EBV and B19V is of paramount importance. We applied Fluorescence of Loop Primer Upon Self-Dequenching Loop-mediated Isothermal Amplification (FLOS-LAMP) for the first time to develop a novel multiplex assay for the detection of EBV and B19V; the fluorophore attached to the probe are self-quenched in unbound state. After binding to the dumbbell-shaped DNA target, the fluorophore is dequenched, resulting in fluorescence development. The novel multiplex FLOS-LAMP assay was optimized by testing various ratios of primer sets. This novel assay, with great specificity, did not cross-react with the common virus. For the detection of EBV and B19V, the limits of detection could reach 969 and 798 copies/µL, respectively, and the assay could be completed within 25 min. Applying this novel assay to detect 200 clinical transplant individuals indicated that the novel assay had high specificity and good sensitivity. We developed multiplex FLOS-LAMP assay for the detection of EBV and B19V, which has the potential to become an important tool for clinical transplant patient screening.

Corneal subbasal nerve plexus reinnervation and stromal cell morphology with different cap thicknesses in small incision lenticule extraction.

PURPOSE: The corneal cap thickness is a vital parameter designed in small incision lenticule extraction (SMILE). The purpose was to investigate the changes in corneal subbasal nerve plexus (SNP) and stromal cells with different cap thicknesses and evaluate the optimized design for the surgery. METHODS: In this prospective, comparative, non-randomized study, a total of 108 eyes of 54 patients who underwent SMILE were allocated into three groups with different corneal cap thicknesses (110 µm, 120 µm or 130 µm group). The SNP and stromal cell morphological changes obtained from in vivo corneal confocal microscopy (IVCCM) along with their refractive outcomes were collected at 1 week, 1 month, 3 months and 6 months postoperatively. One-way analysis of variance (ANOVA) was used to compare the parameters among the three groups. RESULTS: The SNPs in the three groups all decreased after surgery and revealed a gradual increasing trend during the 6-month follow-up. The values of the quantitative nerve metrics were significantly lower in the 110 µm group than in the 120 µm and 130 µm groups, especially at 1 week postoperatively. No difference was detected between the 120 µm and 130 µm groups at any time point. Both Langerhans cells and keratocytes were activated after surgery, and the activation was alleviated during the follow-up. CONCLUSIONS: The SMILE surgeries with 110 µm, 120 µm or 130 µm cap thickness design achieved good efficacy, safety, accuracy and stability for moderate to high myopic correction while the thicker corneal cap was more beneficial for corneal nerve regeneration.

A novel multiplex real-time PCR assay for the detection of cytomegalovirus, Epstein-Barr virus, herpes simplex virus 1/2 and strategies for application to blood screening.

A multiplex real-time PCR has been developed to simultaneously detect transfusion-transmissible pathogens cytomegalovirus, Epstein-Barr virus and herpes simplex virus, as well as to provide sample quality testing, for the conserved regions of the cytomegalovirus UL123 gene, the Epstein-Barr virus BKRF1 gene, and the herpes simplex virus 1/2 UL30 gene, tested on 500 blood donors and 320 transfusion recipients. The laboratory sensitivities for all 3 pathogens were 100 copies/µL. Compared to the commercial real-time PCR reference kit, the multiplex real-time PCR assay for the detection of CMV, EBV and HSV presented 100% consistency. In 820 whole blood samples, the multiplex real-time PCR assay identified 34 (4.15%) positive for CMV DNA, 15 (1.83%) positive for EBV DNA, and 6 (0.73%) positive for HSV DNA. For blood transfusions in high-risk groups, whole blood herpes virus test should be included in the spectrum of pathogen testing for blood donors and recipients.

Glutaminolysis of CD4+ T Cells: A Potential Therapeutic Target in Viral Diseases.

CD4+ T cells play a critical role in the pathogenesis of viral diseases, which are activated by the internal metabolic pathways encountering with viral antigens. Glutaminolysis converts glutamine into tricarboxylic acid (TCA) circulating metabolites by α-ketoglutaric acid, which is essential for the proliferation and differentiation of CD4+ T cells and plays a central role in providing the energy and structural components needed for viral replication after the virus hijacks the host cell. Changes in glutaminolysis in CD4+ T cells are accompanied by changes in the viral status of the host cell due to competition for glutamine between immune cells and host cells. More recently, attempts have been made to treat tumours, autoimmune diseases, and viral diseases by altering the breakdown of glutamine in T cells. In this review, we will discuss the current knowledge of glutaminolysis in the CD4+ T cell subsets from viral diseases, not only increasing our understanding of immunometabolism but also providing a new perspective for therapeutic target in viral diseases.

Expression of PD-1 mitigates phagocytic activities TAM in osteosarcoma.

The high expression of programmed death 1 (PD-1) is a hallmark of T cell exhaustion, consequently inhibiting the anti-tumor immunity, tumor-associated macrophages (TAMs) aggravate Osteosarcoma (OS) progression. However, PD-1 expression on TAMs in OS metastasis remains unclear. Here, we used scRNA-Seq of 15500 individual cells from human OS lung metastatic lesion, identified thirteen major cell clusters. Our data revealed that tumor-infiltrating lymphocytes (TILs) OS lung metastatic accompanied by accumulation of exhausted T cells and regulatory T cells (Tregs). CD3+ T cells from human OS lung metastatic exhibited lower proliferation than in primary tissue. Importantly, TAMs mainly comprise immunosuppressive M2 phenotype in OS metastasis. Mechanistically, we found that PD-1 of TAMs inhibits the phagocytic potency, further promoting the progression of OS metastasis. Therefore, the study provides a strong technical support for OS immunotherapy based on PD-1 inhibitors.

Associations of Near Work, Time Outdoors, and Sleep Duration With Myopic Regression 5 Years After SMILE and FS-LASIK: A Cross-sectional Study.

PURPOSE: To investigate the association of near work, time outdoors, and sleep duration with myopic regression 5 years after small incision lenticule extraction (SMILE) and femtosecond laser-assisted in situ keratomileusis (FS-LASIK) . METHODS: This cross-sectional study included patients who received SMILE or FS-LASIK at Beijing Tongren Hospital 5 years ago. The patients underwent ophthalmic examinations including visual acuity, intraocular pressure, subjective refraction, slit-lamp examination, keratometry, corneal topography, optical coherence tomography, and fundus examination from January 2020 to March 2023. Fluorescein break-up time was measured and the Ocular Surface Disease Index questionnaire was completed to exclude dry eye. A self-administered questionnaire was used to collect data on near work exposure, physical activities, and sleep duration. RESULTS: A total of 323 eyes were included in the analysis, with a 5-year incidence rate of myopic regression after SMILE or FSLASIK of 16.1%. After adjusted for all confounders, total near work more than 8 hours/day revealed a significant association with myopic regression (odds ratio: 2.461; 95% CI: 1.143 to 5.298, P = .021), particularly in younger adults, women, and patients with high myopia and FS-LASIK treatment. The significant association between sleep duration 8 hours/day or more and myopic regression was restricted to women and patients with FS-LASIK (near significant). However, no significant associations were observed between continuous near work or time outdoors and myopic regression. CONCLUSIONS: Excessive near work exposure was associated with greater myopic regression 5 years after SMILE or FS-LASIK. It is crucial to maintain good visual behavior and care for preventing regression after SMILE or FS-LASIK, especially for younger patients and eyes with high myopia. [J Refract Surg. 2024;40(1):e10-e19.].

A Comparison of the Decline in Glomerular Filtration Rate between Elderly Patients with Diabetes and those without Diabetes in Southwest China.

OBJECTIVE: To investigate the effect of high blood glucose on the decline in the estimated glomerular filtration rate (eGFR) in the elderly. METHODS: We compared the decline in eGFR of diabetic and non-diabetic groups in the noninterventional state and analyzed the effect of hyperglycemia on the decline in eGFR among the elderly in a retrospective analysis of 1,223 cases of elderly people aged 65 years or older with a 4-year follow-up period. RESULTS: The prevalence of diabetes in the elderly increased significantly from 12.67% in 2017 to 16.68% in 2021. The rate of decline in eGFR in patients with diabetes was higher than in the population without diabetes, at 9.29% and 5.32%, respectively (both p <0.05). CONCLUSION: The results of this study revealed that the prevalence of diabetes in the elderly increased significantly, and there is a more rapid decrease in the eGFR levels in those with diabetes than those without diabetes.

Identifying key genes for diabetic kidney disease by bioinformatics analysis.

BACKGROUND: There are no reliable molecular targets for early diagnosis and effective treatment in the clinical management of diabetic kidney disease (DKD). To identify novel gene factors underlying the progression of DKD. METHODS: The public transcriptomic datasets of the alloxan-induced DKD model and the streptozotocin-induced DKD model were retrieved to perform an integrative bioinformatic analysis of differentially expressed genes (DEGs) shared by two experimental animal models. The dominant biological processes and pathways associated with DEGs were identified through enrichment analysis. The expression changes of the key DEGs were validated in the classic db/db DKD mouse model. RESULTS: The downregulated and upregulated genes in DKD models were uncovered from GSE139317 and GSE131221 microarray datasets. Enrichment analysis revealed that metabolic process, extracellular exosomes, and hydrolase activity are shared biological processes and molecular activity is altered in the DEGs. Importantly, Hmgcs2, angptl4, and Slco1a1 displayed a consistent expression pattern across the two DKD models. In the classic db/db DKD mice, Hmgcs2 and angptl4 were also found to be upregulated while Slco1a1 was downregulated in comparison to the control animals. CONCLUSIONS: In summary, we identified the common biological processes and molecular activity being altered in two DKD experimental models, as well as the novel gene factors (Hmgcs2, Angptl4, and Slco1a1) which may be implicated in DKD. Future works are warranted to decipher the biological role of these genes in the pathogenesis of DKD.

Corneal metabolic biomarkers for moderate and high myopia in human.

This study aimed to identify the corneal metabolic biomarkers for moderate and high myopia in human. We enrolled 221 eyes from 221 subjects with myopia to perform the femtosecond laser small incision lenticule extraction (SMILE) surgery. Among these, 71 eyes of 71 subjects were enrolled in the low myopic group, 75 eyes of 75 subjects in the moderate myopic group and 75 eyes of 75 subjects in the high myopic group. The untargeted metabolomics analysis was performed to analyze the corneal tissues extracted during the SMILE surgery using an ultra-high-performance liquid chromatography (UHPLC) coupled to a quadrupole time-of-flight (Q-TOF) mass spectrometry (MS). The one-way analysis of variance (ANOVA) was used to identify the different metabolites among the three myopic groups, the orthogonal partial least-squares discriminant analysis (OPLS-DA) model was used to reveal the different metabolites between moderate myopia and low myopia, and between high myopia and low myopia. The Venn gram was used to find the overlapped metabolites of the three datasets of the different metabolites. The stepwise multiple linear regression analysis was used to determine the metabolic molecules associated with manifest refractive spherical equivalents (MRSE). The Receiver Operating Characteristics (ROC) analysis was performed to reveal the corneal biomarkers for moderate and high myopia. The hub biomarker was further selected by the networks among different metabolites created by the Cytoscape software. A total of 1594 metabolites were identified in myopic corneas. 321 metabolites were different among the three myopic groups, 106 metabolites were different between high myopic corneas and low myopic corneas, 104 metabolites were different between moderate myopic corneas and low myopic corneas, and 30 metabolic molecules overlapped among the three datasets. The multivariate linear regression analysis revealed the myopic degree was significantly influenced by the corneal levels of azelaic acid, arginine-proline (Arg-Pro), 1-stearoyl-2-myristoyl-sn-glycero-3-phosphocholine, and hypoxanthine. The ROC curve analysis showed that azelaic acid, Arg-Pro and hypoxanthine were effective in discriminating low myopia from moderate to high myopia with the area under the curve (AUC) values as 0.982, 0.991 and 0.982 for azelaic acid, Arg-Pro and hypoxanthine respectively. The network analysis suggested that Arg-Pro had the maximum connections among these three biomarkers. Thus, this study identified azelaic acid, Arg-Pro and hypoxanthine as corneal biomarkers to discriminate low myopia from moderate to high myopia, with Arg-Pro serving as the hub biomarker for moderate and high myopia.

High-Fat Mouse Model to Explore the Relationship between Abnormal Lipid Metabolism and Enolase in Pancreatic Cancer.

Malignant tumors have become a major social health problem that seriously threatens human health, among which pancreatic cancer has a high degree of malignancy, difficult diagnosis and treatment, short survival time, and high mortality. More and more attention has been paid to abnormal lipid metabolism as a momentous carcinogenesis mechanism. Here, we explored the relationship between abnormal lipid metabolism, enolase, and pancreatic cancer by clinical data analysis. A high-fat mouse model was constructed, and then, a subcutaneous tumorigenesis mouse model of carcinoma of pancreatic cells and a metastatic neoplasm mouse pattern of pancreatic carcinoma cells injected through the tail vein were constructed to explore whether abnormal lipid metabolism affects the progression of pancreatic cancer in mice. We constructed a high-lipid model of pancreatic carcinoma cell lines and knockdown and overexpressed enolase in pancreatic carcinoma cell lines and investigated whether high lipid regulates epithelial-mesenchymal transition (EMT) by upregulating enolase (ENO), thereby promoting the cells of pancreatic carcinoma to invade and migrate. Triglycerides, total cholesterol, free cholesterin, high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), and neuron-specific enolase (NSE) from pancreatic cancer patients and nonpancreatic cancer patients were tested. The differences in blood lipids between patients with and without pancreatic carcinoma were compared, and the correlation between blood lipids and neuron-specific enolase was analyzed. We confirmed that the serum triglyceride level of pancreatic cancer patients at initial diagnosis is overtopping nonpancreatic cancer patients, and the neuron-specific enolase level of patients with pancreatic carcinoma is better than nonpancreatic carcinoma sufferers. Triglyceride level is positively correlated with neuron-specific enolase level, and serum triglyceride level has predictive value for pancreatic cancer. Hyperlipidemia can promote tumor growth and increase the expression levels of ENO1, ENO2, and ENO3 in subcutaneous tumor formation of pancreatic cancer in mice. Additional hyperlipidemia promoted pancreatic carcinoma metastasis in the lung in mice injected through the tail vein, which confirmed that hyperlipidemia accelerated the process of EMT by increasing the expression of ENO1, ENO2, and ENO3, therefore promoting the pancreatic cancer cell metastasis.

Effect of 3% Diquafosol Sodium on Dry Eye After Femtosecond Laser-Assisted In Situ Keratomileusis and Small Incision Lenticule Extraction Surgery in High-Myopic Eyes.

PURPOSE: To evaluate the effect of 3% diquafosol sodium eye drop on dry eye after femtosecond laser-assisted in situ keratomileusis (FS-LASIK) and small incision lenticule extraction (SMILE) in high-myopic eyes. METHODS: Eighty-one cases with high myopia (162 eyes) who received FS-LASIK or SMILE were divided into four groups by surgical design and tear film stability: D-FS-LASIK (5s

Effects of riboflavin/ultraviolet-A(UVA) scleral crosslinking on the mechanical behavior of the scleral fibroblasts of lens-induced myopia Guinea pigs.

Myopia is becoming increasingly severe, and studies have shown that the cellular mechanics of scleral fibroblasts are altered following myopia. Scleral UVA-Riboflavin Collagen Crosslinking（sCXL） is a promising treatment for myopia prevention and control of axial growth. Understanding the mechanical properties of scleral fibroblasts is crucial, as it influences the cellular response and limits the extent of molecular deformation triggered. Thus, our study aimed to investigate the effect of mechanical properties of scleral fibroblasts in a lens-induced myopic guinea pig model following sCXL. For this purpose, we performed the 0.1% riboflavin/UVA scleral crosslinking (365 nm,3 mW/cm2,30 min) in the right eyes of guinea pigs in Group CXL. In Group LIM, the right eyes were only administrated negative lens for 6 weeks. No treatment was performed in both eyes of the guinea pigs in group Control. The scleral fibroblasts were isolated and cultured from the scleral tissue at the cross-linking area in Group CXL and the corresponding area in Group LIM and control. The curve of the length of microtubules inhaled by cells under negative pressure was measured by a microaspiration-based isolation technique, and the equilibrium Young's modulus and apparent viscosity of scleral fibroblasts were calculated by formula fitting. The equilibrium Young's modulus of scleral fibroblasts in group CXL was significantly lower than that in the LIM group (P < 0.01, two-sample t-test between pairs）, and there was no significant difference between groups CXL and control. The results show that sCXL can effectively moderate the phenomenon that scleral fibroblasts are not easy to deform after myopia. The apparent viscosity modulus in the CXL group was higher than the groups' control and LIM. Taken together, our data demonstrate the biomechanics of the scleral fibroblasts altered after Riboflavin/UVA scleral collagen cross-linking in a lens-induced myopia model.

Comparisons of the protein expressions between high myopia and moderate myopia on the anterior corneal stroma in human.

PURPOSE: To investigate the differentially expressed proteins (DEP) between high myopia and moderate myopia on the anterior corneal stroma. METHODS: Tandem mass tag (TMT) quantitative proteomics was utilized to reveal proteins. DEPs were screened by the multiple change of more than 1.2 times or less than 0.83 and the P value < 0.05. The DEPs were functional annotated by Gene Ontology (GO) terms. Proteins and protein interaction (PPI) networks were conducted with String online tool. Parallel reaction monitoring (PRM) data processing was used to verify the TMT proteomics results. RESULTS: There are 36 DEPs between high myopia and moderate myopia on the anterior corneal stroma, of which 11 proteins are upregulated, 25 proteins are downregulated. The GO analysis demonstrated keratinocyte migration and structural constituent of cytoskeleton that are significantly changed with most of the proteins decreased in high myopic corneas. Keratin 16 (KRT16) and erythrocyte membrane protein band 4.1-like protein 4B are the only two proteins involved in both functions. The PPI analysis showed keratin type II cytoskeletal 6A (KRT6A) and KRT16 that have strong connections. Immunoglobulin lambda variable 8-61(IGLV8-61) and nicotinamide phosphoribosyl transferase (NAMPT) have consistent results with the TMT. CONCLUSIONS: The high myopic corneas have 36 DEPs compared to the moderate myopic corneas on the anterior corneal stroma. Keratinocyte migrations and structural constituent of cytoskeleton are weakened in high myopic corneas, which may partly account for the lower corneal biomechanics in high myopic eyes. The lower expressed KRT16 plays important roles in high myopic corneas.

Development and clinical validation of a one-step pentaplex real-time reverse transcription PCR assay for detection of hepatitis virus B, C, E, Treponema pallidum, and a human housekeeping gene.

BACKGROUND: With the safety of blood transfusion being a major public health concern, the development of a rapid, sensitive, specific, and cost-effective multiplex PCR assay for simultaneous detection of hepatitis B virus(HBV), hepatitis C virus (HCV), hepatitis E virus (HEV), and Treponema pallidum(T. pallidum) in blood is crucial. METHODS: Five primer pairs and probes were designed towards conserved regions of target genes and used to establish a one-step pentaplex real-time reverse transcription PCR(qRT-PCR) assay for simultaneous detection of HBV, HCV, HEV, T. pallidum, and RNase P(housekeeping gene), providing sample quality check. The clinical performance of the assay was further determined with 2400 blood samples from blood donors and patients in Zhejiang province, and compared the results with commercial singleplex qPCR and serological assays. RESULTS: The 95% limit of detection(LOD) of HBV, HCV, HEV, and T. pallidum were 7.11 copies/µL, 7.65 copies/µL, 8.45 copies/µL, and 9.06 copies/µL, respectively. Moreover, the assay has good specificity and precision. Compared to the singleplex qPCR assay, the novel assay for detecting HBV, HCV, HEV, and T. pallidum presented 100% clinical sensitivity, specificity, and consistency. Several discrepant results between serological and pentaplex qRT-PCR assays were found. Of 2400 blood samples, there were 2(0.08%) HBsAg positive samples, 3(0.13%) anti-HCV positive samples, 29(1.21%) IgM anti-HEV positive samples and 6(0.25%) anti-T. pallidum positive samples proven negative in nucleic acid detection. 1(0.04%) HBV DNA positive sample and 1(0.04%) HEV RNA positive sample were detected negative by serological testing. CONCLUSIONS: The developed pentaplex qRT-PCR is the first assay on simultaneous, sensitive, specific, and reproducible detection of HBV, HCV, HEV, T. pallidum, and RNase P in a single tube. It could detect pathogens in blood during the window period of infection and is a good tool for effectively screening blood donors and early clinical diagnosis.

A deep learning model for the differential diagnosis of benign and malignant salivary gland tumors based on ultrasound imaging and clinical data.

Background: The preoperative differentiation between benign parotid gland tumors (BPGTs) and malignant parotid gland tumors (MPGTs) is of great significance for therapeutic decision-making. Deep learning (DL), an artificial intelligence algorithm based on neural networks, can help overcome inconsistencies in conventional ultrasonic (CUS) examination outcomes. Therefore, as an auxiliary diagnostic tool, DL can support accurate diagnosis using massive ultrasonic (US) images. This current study developed and validated a DL-based US diagnosis for the preoperative differentiation of BPGT from MPGT. Methods: A total of 266 patients, including 178 patients with BPGT and 88 patients with MPGT, were consecutively identified from a pathology database and enrolled in this study. Ultimately, considering the limitations of the DL model, 173 patients were selected from the 266 patients and divided into 2 groups: a training set, and a testing set. US images of the 173 patients were used to construct the training set (including 66 benign and 66 malignant PGTs) and testing set (consisting of 21 benign and 20 malignant PGTs). These were then preprocessed by normalizing the grayscale of each image and reducing noise. Processed images were imported into the DL model, which was then trained to predict the images from the testing set and evaluated for performance. Based on the training and validation datasets, the diagnostic performance of the 3 models was assessed and verified using receiver operating characteristic (ROC) curves. Ultimately, before and after combining the clinical data, we compared the area under the curve (AUC) and diagnostic accuracy of the DL model with the opinions of trained radiologists to evaluate the application value of the DL model in US diagnosis. Results: The DL model showed a significantly higher AUC value compared to doctor 1 + clinical data, doctor 2 + clinical data, and doctor 3 + clinical data (AUC =0.9583 vs. 0.6250, 0.7250, and 0.8025 respectively; all P<0.05). In addition, the sensitivity of the DL model was higher than the sensitivities of the doctors combined with clinical data (97.2% vs. 65%, 80%, and 90% for doctor 1 + clinical data, doctor 2 + clinical data, and doctor 3 + clinical data, respectively; all P<0.05). Conclusions: The DL-based US imaging diagnostic model has excellent performance in differentiating BPGT from MPGT, supporting its value as a diagnostic tool for the clinical decision-making process.

Bulk and single-cell transcriptome analyses of islet tissue unravel gene signatures associated with pyroptosis and immune infiltration in type 2 diabetes.

Introduction: Type 2 diabetes (T2D) is a common chronic heterogeneous metabolic disorder. However, the roles of pyroptosis and infiltrating immune cells in islet dysfunction of patients with T2D have yet to be explored. In this study, we aimed to explore potential crucial genes and pathways associated with pyroptosis and immune infiltration in T2D. Methods: To achieve this, we performed a conjoint analysis of three bulk RNA-seq datasets of islets to identify T2D-related differentially expressed genes (DEGs). After grouping the islet samples according to their ESTIMATE immune scores, we identified immune- and T2D-related DEGs. A clinical prediction model based on pyroptosis-related genes for T2D was constructed. Weighted gene co-expression network analysis was performed to identify genes positively correlated with pyroptosis-related pathways. A protein-protein interaction network was established to identify pyroptosis-related hub genes. We constructed miRNA and transcriptional networks based on the pyroptosis-related hub genes and performed functional analyses. Single-cell RNA-seq (scRNA-seq) was conducted using the GSE153885 dataset. Dimensionality was reduced using principal component analysis and t-distributed statistical neighbor embedding, and cells were clustered using Seurat. Different cell types were subjected to differential gene expression analysis and gene set enrichment analysis (GSEA). Cell-cell communication and pseudotime trajectory analyses were conducted using the samples from patients with T2D. Results: We identified 17 pyroptosis-related hub genes. We determined the abundance of 13 immune cell types in the merged matrix and found that these cell types were correlated with the 17 pyroptosis-related hub genes. Analysis of the scRNA-seq dataset of 1892 islet samples from patients with T2D and controls revealed 11 clusters. INS and IAPP were determined to be pyroptosis-related and candidate hub genes among the 11 clusters. GSEA of the 11 clusters demonstrated that the myc, G2M checkpoint, and E2F pathways were significantly upregulated in clusters with several differentially enriched pathways. Discussion: This study elucidates the gene signatures associated with pyroptosis and immune infiltration in T2D and provides a critical resource for understanding of islet dysfunction and T2D pathogenesis.

Relationships of adiponectin to regional adiposity, insulin sensitivity, serum lipids, and inflammatory markers in sedentary and endurance-trained Japanese young women.

Introduction: This study aims to compare the differences in circulating adiponectin levels and their relationships to regional adiposity, insulin resistance, serum lipid, and inflammatory factors in young, healthy Japanese women with different physical activity statuses. Methods: Adipokines (adiponectin and leptin), full serum lipid, and inflammatory factors [white blood cell counts, C-reactive protein, tumor necrosis factor-α, tissue plasminogen activator inhibitor-1 (PAI-1)] were measured in 101 sedentary and 100 endurance-trained healthy Japanese women (aged 18-23 years). Insulin sensitivity was obtained through a quantitative insulin-sensitivity check index (QUICKI). Regional adiposity [trunk fat mass (TFM), lower-body fat mass (LFM), and arm fat mass (AFM)] was evaluated using the dual-energy X-ray absorptiometry method. Results: No significant difference was observed between the sedentary and trained women in terms of adiponectin levels. The LFM-to-TFM ratio and the high-density lipoprotein cholesterol (HDL-C) were the strong positive determinants for adiponectin in both groups. Triglyceride in the sedentary women was closely and negatively associated with adiponectin, as well as PAI-1 in the trained women. The QUICKI level was higher in the trained than sedentary women. However, no significant correlation between adiponectin and insulin sensitivity was detected in both groups. Furthermore, LFM was associated with a favorable lipid profile against cardiovascular diseases (CVDs) in the whole study cohort, but this association became insignificant when adiponectin was taken into account. Conclusions: These findings suggest that adiponectin is primarily associated with regional adiposity and HDL-C regardless of insulin sensitivity and physical activity status in young, healthy women. The associations among adiponectin, lipid, and inflammatory factors are likely different in women with different physical activity statuses. The correlation of LFM and a favorable lipid profile against CVD and adiponectin is likely involved in this association.

miR-223-3p mediates the diabetic kidney disease progression by targeting IL6ST/STAT3 pathway.

Diabetic kidney disease (DKD), the most pervasive complication in diabetic patients, has become a major health threat to the aging population. Our previous miRNA profiling identified hsa-miR-223-3p as a dysregulated miRNA in the DKD samples, which may serve as a biomarker for DKD diagnosis. However, the specific mechanism of miR-223-3p in the pathogenesis of DKD remains to be elucidated. In this study, we first verified that miR-223-3p level was significantly decreased in the in vitro cell model and in vivo db/db DKD model, accompanied with endothelial cell damage. Importantly, inhibiting the expression of miR-223-3p exacerbated high-glucose induced damages in Human Umbilical Vein Endothelial Cells (HUVECs) and Human Renal Glomerular Endothelial Cells (HRGECs), while miR-223-3p overexpression showed the opposite effect. We further demonstrated that miR-223-3p associated with IL6T mRNA and attenuated the progression of DKD by suppressing the downstream STAT3 activation, indicative of the implication of miR-223-3p/IL6T/STAT3 axis in the pathogenesis of DKD.

Unnatural tetradeoxy echinocandins produced by gene cluster design and heterologous expression.

The hydroxyl groups in the amino acid residues of echinocandin B were related to the biological activity, the instability, and the drug resistance. The modification of hydroxyl groups was expected to obtain the new lead compounds for next generation of echinocandin drug development. In this work one method for heterologous production of the tetradeoxy echinocandin was achieved. A reconstructed biosynthetic gene cluster for tetradeoxy echinocandins composed of ecdA/I/K and htyE was designed and successfully hetero-expressed in Aspergillus nidulans. The target product of echinocandin E (1) together with one unexpected derivative echinocandin F (2), were isolated from the fermentation culture of engineered strain. Both of compounds were unreported echinocandin derivatives and the structures were identified on the basis of mass and NMR spectral data analysis. Compared with echinocandin B, echinocandin E demonstrated superior stability and comparable antifungal activity.

Effects of riboflavin/ultraviolet-A scleral collagen cross-linking on regional scleral thickness and expression of MMP-2 and MT1-MMP in myopic guinea pigs.

OBJECTIVE: To investigate the effects of scleral collagen cross-linking (SXL) using riboflavin and ultraviolet A (UVA) light on the scleral thickness of different regions and expression of matrix metalloproteinase 2 (MMP-2) and membrane-type MMP-1 (MT1-MMP) in guinea pigs with lens-induced myopia. METHODS: Forty-eight 4-week-old guinea pigs were assigned to three groups (n = 16 per group): SXL group, lens-induced myopia (LIM) group, and control group. The sclera of the right eye of the guinea pig in the SXL group was surgically exposed, riboflavin was dropped on the treatment area for 10 minutes before the 30-minute UVA irradiation. The same surgical procedure was performed in the LIM group without UVA irradiation. The -10.00 D lenses were then placed on the right eyes of guinea pigs in the SXL and LIM groups for six weeks. The control group received no treatment. The left eyes were untreated in all groups. The ocular axial length (AXL) and refraction were measured at 4 weeks and 10 weeks of age. 10-week-old guinea pigs were sacrificed, and the right eyes were enucleated and evenly divided for preparation of hematoxylin and eosin (HE) stained sections, quantitative real-time polymerase chain reaction (qPCR) and western blotting. The scleral thickness of different regions was measured on HE stained sections. The temporal half of the sclera was harvested to measure the expression of MMP-2 and MT1-MMP by qPCR and western blotting. RESULTS: The AXL was significantly shorter, and the degree of myopic refraction was significantly lower in the SXL group than those in the LIM group at 10 weeks of age. The scleral thickness of the cross-linked area was significantly greater in the SXL group than that of the corresponding area in the LIM group, while the scleral thickness of the untreated nasal side was not significantly different between the SXL group and the LIM group. The expression of MMP-2 and MT1-MMP of the cross-linked sclera was significantly downregulated compared with that of the corresponding area in the LIM group. CONCLUSION: Riboflavin/UVA SXL could slow myopia progression and thicken the cross-linked sclera in guinea pigs, which might be related to the downregulation of MMP-2 and MT1-MMP expression during the scleral remodeling process.