c-Cbl induced podocin ubiquitination contributes to the podocytes injury in diabetic nephropathy.

The ubiquitination function in diabetic nephropathy (DN) has attracted much attention, but there is a lack of information on its ubiquitylome profile. To examine the differences in protein content and ubiquitination in the kidney between db/db mice and db/m mice, we deployed liquid chromatography-mass spectrometry (LC-MS/MS) to conduct analysis. We determined 145 sites in 86 upregulated modified proteins and 66 sites in 49 downregulated modified proteins at the ubiquitinated level. Moreover, 347 sites among the 319 modified proteins were present only in the db/db mouse kidneys, while 213 sites among the 199 modified proteins were present only in the db/m mouse kidneys. The subcellular localization study indicated that the cytoplasm had the highest proportion of ubiquitinated proteins (31.87%), followed by the nucleus (30.24%) and the plasma membrane (20.33%). The enrichment analysis revealed that the ubiquitinated proteins are mostly linked to tight junctions, oxidative phosphorylation, and thermogenesis. Podocin, as a typical protein of slit diaphragm, whose loss is a crucial cause of proteinuria in DN. Consistent with the results of ubiquitination omics, the K261R mutant of podocin induced the weakest ubiquitination compared with the K301R and K370R mutants. As an E3 ligase, c-Cbl binds to podocin, and the regulation of c-Cbl can impact the ubiquitination of podocin. In conclusion, in DN, podocin ubiquitination contributes to podocyte injury, and K261R is the most significant site. c-Cbl participates in podocin ubiquitination and may be a direct target for preserving the integrity of the slit diaphragm structure, hence reducing proteinuria in DN.

ACSF2 and lysine lactylation contribute to renal tubule injury in diabetes.

AIMS: Lactate accumulation is reported to be a biomarker for diabetic nephropathy progression. Lactate drives lysine lactylation, a newly discovered post-translational modification that is involved in the pathogenesis of cancers and metabolic and inflammatory disease. Here, we aimed to determine whether lysine lactylation is involved in the pathogenesis of diabetic nephropathy. METHODS: Renal biopsy samples from individuals with diabetic nephropathy (n=22) and control samples from individuals without diabetes and kidney disease (n=9) were obtained from the First Affiliated Hospital of Zhengzhou University for immunohistochemical staining. In addition, we carried out global lactylome profiling of kidney tissues from db/m and db/db mice using LC-MS/MS. Furthermore, we assessed the role of lysine lactylation and acyl-CoA synthetase family member 2 (ACSF2) in mitochondrial function in human proximal tubular epithelial cells (HK-2). RESULTS: The expression level of lysine lactylation was significantly increased in the kidneys of individuals with diabetes as well as in kidneys from db/db mice. Integrative lactylome analysis of the kidneys of db/db and db/m mice identified 165 upregulated proteins and 17 downregulated proteins, with an increase in 356 lysine lactylation sites and a decrease in 22 lysine lactylation sites decreased. Subcellular localisation analysis revealed that most lactylated proteins were found in the mitochondria (115 proteins, 269 sites). We further found that lactylation of the K182 site in ACSF2 contributes to mitochondrial dysfunction. Finally, the expression of ACSF2 was notably increased in the kidneys of db/db mice and individuals with diabetic nephropathy. CONCLUSIONS: Our study strongly suggests that lysine lactylation and ACSF2 are mediators of mitochondrial dysfunction and may contribute to the progression of diabetic nephropathy. DATA AVAILABILITY: The LC-MS/MS proteomics data have been deposited in the ProteomeXchange Consortium database ( https://proteomecentral.proteomexchange.org ) via the iProX partner repository with the dataset identifier PXD050070.

Anti-scaling performances of different aged landfill leachates on PPR and ABS pipe materials.

Landfill leachate, because of its complex components, may cause pipe clogging during its collection and transportation, and pose a threat to the environment. This study considers two typical drainage pipe materials, i.e., polypropylene-random (PPR) and acrylonitrile-butadiene-styrene (ABS), to discriminate their anti-scaling performances through a landfill leachate immersion experiment. The results show that both PPR and ABS pipe materials immersed in the younger-aged leachate are prone to scaling. The mass concentrations of Ca2+ in the leachate play a key role in scale formation, followed by Mg2+, pH, oxidation-reduction potential (ORP), dissolved oxygen (DO), and Cl-. In particular, Ca2+, pH, and DO show a positive relationship in scaling, while Mg2+ acts in a negative relationship. Calcium carbonate is the major constituent of the scales, accounting for 72.3% of the total. The scaling on the surface of the PPR pipe material is more serious than that of the ABS pipe material, revealed by the corresponding weight increment that is 33% higher than that related to ABS during the experimental period. It is thus indicated that ABS has better anti-scaling performance, which may be an optional material selected for the system design of leachate collection and transportation.

Dynamic Predictive Models with Visualized Machine Learning for Assessing the Risk of Lung Metastasis in Kidney Cancer Patients.

Objective: To establish and verify the clinical prediction model of lung metastasis in renal cancer patients. Method: Kidney cancer patients from January 1, 2010, to December 31, 2017, in the SEER database were enrolled in this study. In the first section, LASSO method was adopted to select variables. Independent influencing factors were identified after multivariate logistic regression analysis. In the second section, machine learning (ML) algorithms were implemented to establish models and 10-foldcross-validation was used to train the models. Finally, receiver operating characteristic curves, probability density functions, and clinical utility curve were applied to estimate model's performance. The final model was shown by a website calculator. Result: Lung metastasis was confirmed in 7.43% (3171 out of 42650) of study population. In multivariate logistic regression, bone metastasis, brain metastasis, grade, liver metastasis, N stage, T stage, and tumor size were independent risk factors of lung metastasis in renal cancer patients. Primary site and sequence number were independent protection factors of LM in renal cancer patients. The above 9 impact factors were used to develop the prediction models, which included random forest (RF), naive Bayes classifier (NBC), decision tree (DT), xgboost (XGB), gradient boosting machine (GBM), and logistic regression (LR). In 10-foldcross-validation, the average area under curve (AUC) ranked from 0.907 to 0.934. In ROC curve analysis, AUC ranged from 0.879-0.922. We found that the XGB model performed best, and a Web-based calculator was done according to XGB model. Conclusion: This study provided preliminary evidence that the ML algorithm can be used to predict lung metastases in patients with kidney cancer. This low cost, noninvasive and easy to implement diagnostic method is useful for clinical work. Of course this model still needs to undergo more real-world validation.

Pediatric esophagogastroduodenoscopy in china: indications, diagnostic yield, and factors associated with findings.

BACKGROUND: Large-scale data on esophagogastroduodenoscopy (EGD) in China are scarce. This study aimed to assess the indications and diagnostic yield of EGD in children and the relationship between factors (such as age, sex, and indications) and diagnostic yield. METHODS: We performed a prospective cross-sectional observational study involving patients aged < 18 years who underwent diagnostic EGD. The study was conducted in five children's hospitals, each in a different city. Demographic features, indications for endoscopy, and endoscopic and histopathological findings were collected. Univariable and multivariable ordinal logistic regression analyses of the relationship between the factors and diagnostic yield were performed. RESULTS: The study included 2268 patients (male/female ratio, 1.3:1) with a median age of 8.68 years. Among the 2268 children, the most frequent indications were abdominal pain in 1954 (86.2%), recurrent vomiting in 706 (31.1%), weight loss in 343 (15.1%), and others. The endoscopic yield was 62.5% and was the highest in patients with dysphagia (90.9%). The histologic yield was 30.4% and was the highest in patients with unexplained anemia (45.5%). On multivariable regression analysis, the endoscopic yield was associated with dysphagia, gastrointestinal (GI) bleeding, and recurrent vomiting, and the histologic yield was associated with age. Different groups of patients with abdominal pain had variable probabilities of abnormal endoscopic findings. CONCLUSIONS: The most frequent indication of pediatric EGD is abdominal pain, with variable probabilities of abnormal endoscopic findings in different groups. Endoscopic yield and histologic yield are associated with certain alarming features. TRIAL REGISTRATION: The trial registration number (ClinicalTrials. gov): NCT03603093 (The study was registered on 27/07/2018).

Genomic insights into genetic diploidization in the homosporous fern Adiantum nelumboides.

Whole genome duplication has been recognized as a major process in speciation of land plants, especially in ferns. Whereas genome downsizing contributes greatly to the post-genome shock responses of polyploid flowering plants, diploidization of polyploid ferns diverges by maintaining most of the duplicated DNA and is thus expected to be dominated by genic processes. As a consequence, fern genomes provide excellent opportunities to study ecological speciation enforced by expansion of protein families via polyploidy. To test the key predictions of this hypothesis, we reported the de novo genome sequence of Adiantum nelumboides, a tetraploid homosporous fern. The obtained draft genome had a size of 6.27 Gb assembled into 11,767 scaffolds with the contig N50 of 1.37 Mb. Repetitive DNA sequences contributed with about 81.7%, a remarkably high proportion of the genome. With 69,568 the number of predicted protein-coding genes exceeded those reported in most other land plant genomes. Intragenomic synteny analyses recovered 443 blocks with the average block size of 1.29 Mb and the average gene content of 16 genes. The results are consistent with the hypothesis of high ancestral chromosome number, lack of substantial genome downsizing, and dominance of genic diploidization. As expected in the calciphilous plants, a notable number of detected genes were involved in calcium uptake and transport. In summary, the genome sequence of a tetraploid homosporous fern not only provides access to a genomic resource of a derived fern, but also supports the hypothesis of maintenance of high chromosome numbers and duplicated DNA in young polyploid ferns.

Changes in Retinal Vessel Flow after Small Incision Lenticule Extraction.

Objective: In order to analyze changes in retinal vessel flow after small incision lenticule extraction (SMILE). Methods: A total of 32 patients (62 eyes) who underwent SMILE were enrolled in this prospective study. Optical parameters, including vessel density (VD), and perfusion density (PD) of foveal, parafoveal, and perifoveal regions, respectively, were measured before surgery and at 1 day, 1 week, 1 month, and 3 months postoperation. Preoperative parameters and surgical parameters were recorded. Results: Significant decreases in VD and PD on postoperative day 1 were detected in all quadrants, both in 3 mm and in 6 mm regions (P < 0.001). One month after surgery, VD returned to preoperative levels. None of the preoperative and surgical parameters were significantly correlated with the VD and PD fluctuations (all P > 0.05). Conclusion. VD may decrease significantly with regional disparity 1 day after SMILE while recovering at 1 month. Elevation of intraocular pressure due to suction may account for such changes.

Application of Immune Checkpoint Inhibitors in the Treatment of Cholangiocarcinoma.

Cholangiocarcinoma is a general term for intrahepatic and extrahepatic malignant tumors deriving in the biliary system. According to the location, it is divided into intrahepatic cholangiocarcinoma, hilar cholangiocarcinoma, and distal cholangiocarcinoma. Progressive cholangiocarcinoma yields poor outcomes with radiotherapy; therefore, there is an urgent need for new therapeutic breakthroughs. Immune checkpoint inhibitor (ICI) therapy brings the treatment for cancer into a new field, with the use of drugs targeting PD-1/PD-L1 and CTLA-4 considerably extending the survival of patients with melanoma, lung cancer, and other solid tumors. The FDA has approved the application of pembrolizumab for solid tumors with high microsatellite instability and defective mismatch repair, including cholangiocarcinoma. Moreover, the combination of ICIs with chemotherapy and radiation therapy showed good promise. The aim of the present study was to review the application of ICIs in the treatment of cholangiocarcinoma and to summarize the reported individualized immunotherapy-based protocols and ongoing clinical trials for clinical reference.

Inhibitory Effect of Delphinidin on Oxidative Stress Induced by H2O2 in HepG2 Cells.

Chronic liver diseases (CLDs) are correlated with oxidative stress induced by the accumulation of intracellular reactive oxygen species (ROS). In this study, we employed HepG2, a human liver carcinoma cell line containing many antioxidant enzymes, to explore the function of delphinidin against oxidative stress induced by H2O2 and to provide scientific data of the molecular mechanism. Cells were pretreated with different concentrations of delphinidin (10 µmol/L, 20 µmol/L, and 40 µmol/L) for 2 h before treatment with 750 µM H2O2 for 1 h. The results showed that H2O2 decreased the survival rate of HepG2 cells and increased the level of ROS, but delphinidin pretreatment could possess the opposite result. At the same time, the expression of Nrf2 was enhanced by the delphinidin pretreatment. This was because delphinidin promoted Nrf2 nuclear translocation and inhibited its degradation, which led to the increase expression of antioxidant protein HO-1 (Nrf2-related phase II enzyme heme oxygenase-1). Besides, we found that delphinidin could significantly alleviate the reduction of Nrf2 protein levels and the accumulation of intracellular ROS levels in Nrf2 knockdown HepG2 cells. In conclusion, our study suggested that delphinidin, as an effective antioxidant, protected HepG2 cells from oxidative stress by regulating the expression of Nrf2/HO-1.

Preoperative Ultra-Short-Course Chemotherapy Combined with Surgery for the Treatment of Chest Wall Tuberculosis.

OBJECTIVE: To investigate the safety and efficacy of preoperative ultra-short-course chemotherapy, combined with surgical treatment for chest wall tuberculosis and summarize our experience in this regard, to provide a reference for national and international clinicians. METHODS: A retrospective analysis was conducted of the clinical data, preoperative anti-tuberculosis duration, and postoperative recurrence rate in 263 patients with chest wall tuberculosis spanning 5 years. RESULTS: Overall, 263 patients were treated with anti-tuberculosis drugs for about ± 12.49 days during the preoperative period. Simple chest wall tuberculosis was treated for ± 5.87 days and composite chest wall tuberculosis for ± 5.11 days. The postoperative recurrence rate of chest wall tuberculosis was 3.80%, which was close to or lower than the recurrence rate of routine preoperative anti-tuberculous therapy in patients subjected to ultra-short-range anti-tuberculosis treatment before surgery. CONCLUSION: Preoperative ultra-short-course chemotherapy combined with surgical treatment of chest wall tuberculosis did not increase the recurrence rate of chest wall tuberculosis; moreover, it could effectively shorten hospitalization time and improve patient compliance. Full-line anti-tuberculosis treatment and complete resolution of tuberculosis infections are crucial to curing chest wall tuberculosis.

Liposomal N-acylethanolamine-hydrolyzing acid amidase (NAAA) inhibitor F96 as a new therapy for colitis.

Despite numerous advances in the pathological mechanism of inflammatory bowel disease (IBDs), the ideal therapy is still missing. N-Acylethanolamine-hydrolyzing acid amidase (NAAA), a cysteine hydrolase that deactivates fatty acid ethanolamides, has been recognized as a new therapeutic target for IBDs. Herein, we proposed liposomal F96, a selective and potent NAAA inhibitor, as a new therapy for IBDs. F96, with an IC50 of 270 nM for NAAA, was encapsulated into anionic liposome and the anti-inflammatory activity was evaluated in dextran sulfate sodium (DSS) induced colitis mice. The anionic liposomes showed significantly higher accumulation in the colon compared with the small intestine and cecum at 6 and 10 h after administration in DSS induced colitis mice. DSS induction significantly increased myeloperoxidase (MPO) activities and shortened the colon length, while free F96 significantly lowered tissue MPO activity and restored the colon length. Anionic liposome encapsulation significantly enhanced the therapeutic efficacy of F96, as liposomal F96 resulted in lower MPO activity and better colon length restoration effects compared with those treated with free F96. This study offers a new treatment option for colitis, which may pave the way for new therapies for other IBDs.

Combination of Capsaicin and Capsiate Induces Browning in 3T3-L1 White Adipocytes via Activation of the Peroxisome Proliferator-Activated Receptor γ/ß3-Adrenergic Receptor Signaling Pathways.

This study investigated the effects and molecular mechanism of a combination of capsaicin and capsiate on promoting lipid metabolism and inducing browning in 3T3-L1 white adipocytes. The combination significantly suppressed lipid accumulation in adipocytes ( p = 0.019) and robustly improved lipid metabolic profiles, including decreased triacylglycerol (0.6703 ± 0.0385 versus 0.2849 ± 0.0188 mmol/g of protein; p < 0.001), total cholesterol (0.1282 ± 0.0241 versus 0.0651 ± 0.0178 mmol/g of protein; p = 0.003), and low-density lipoprotein cholesterol (0.0021 ± 0.0017 versus 0.0005 ± 0.0002 mmol/g of protein; p = 0.024) and increased high-density lipoprotein cholesterol (0.0162 ± 0.0141 versus 0.1002 ± 0.0167 mmol/g of protein; p = 0.012). Furthermore, this combination markedly upgraded the protein levels of cluster of differentiation 36 ( p = 0.007) and adipose triglyceride lipase ( p = 0.013) and phosphorylation of hormone-sensitive lipase at Ser660, Ser565, and Ser563 ( p < 0.001, p = 0.027, and p = 0.002, respectively), indicating increases of fatty acid transport and lipolysis. The levels of lipid metabolism regulators, phosphorylation of adenosine-monophosphate-activated protein kinases α and ß ( p = 0.011, and p < 0.001, respectively), sirtuin 1 ( p = 0.004), and vanilloid transient receptor subtype I ( p = 0.014) were also increased by the combination. Moreover, the combination greatly activated the browning program in adipocytes, as demonstrated by increases in beige-specific gene and protein. Further research found that the protein levels of peroxisome proliferator-activated receptor γ (PPARγ; p = 0.001) and ß3-adrenergic receptor (ß3-AR; p = 0.026) were elevated by the combination, and most of the beige-specific markers were abolished by pretreatment of antagonists of PPARγ or ß3-AR. In conclusion, these results indicated that a combination of capsaicin and capsiate could induce browning in white adipocytes via activation of the PPARγ/ß3-AR signaling pathway, and this combination might be worth investigating as a potential cure for obesity.

Sestrin2 and sestrin3 suppress NK-92 cell-mediated cytotoxic activity on ovarian cancer cells through AMPK and mTORC1 signaling.

Ovarian cancer is one of the major cancer types. NK-92 cell line, which has consistently and reproducibly high anti-tumor cytotoxicity, may be used for immunotherapy against ovarian cancer. Understanding the mechanisms that regulate the anti-tumor activity of NK-92 cells is important for developing novel therapeutic strategies. In the current study, using an ovarian cancer xenograft mouse model, we identified the up-regulation of sestrin2 (SESN2) and sestrin3 (SESN3) in intratumoral NK-92 cells. Lentivirus-transduced NK-92 cells, which overexpressed SESN2 or SESN3 after doxycycline treatment, exhibited less expression of activating receptors, perforin and granzyme B. Overexpression of SESN2 and SESN3 impaired tumoricidal effect of NK-92 cells both in vitro and in vivo. Furthermore, overexpression of SESN2 and SESN3 inhibited mTORC1 signaling while promoting AMPK signaling in NK-92 cells. Taken together, our data highlights the crucial effects of SESN2 and SESN3 on NK-92 cell-mediated anti-ovarian cancer activity. This study might be valuable for designing a novel therapeutic strategy for ovarian cancer.

Clonorchis sinensis enolase: identification and biochemical characterization of a glycolytic enzyme from excretory/secretory products.

Enolase plays a key role in energy metabolism and development of most organisms. We isolated a gene encoding enolase from Clonorchis sinensis (C. sinensis) adult cDNA library and expressed the recombinant protein in Escherichia coli. C. sinensis enolase (Csenolase) was identified as both an excretory/secretory product and a tegumental component of C. sinensis by western blot analysis. The transcriptional level of Csenolase was examined at adult worm, metacercaria, cercaria and egg of C. sinensis, and results showed that Csenolase is transcribed at the four life stages of C. sinensis while showing a significant higher expression level at the stage of adult worm. Immunohistochemical localization indicated that Csenolase was specifically deposited on the tegument of adult worm and cyst wall of metacercaria. Ligand blot assay revealed a specific characteristic of dose-dependent plasminogen-binding activity of Csenolase and kinetic parameters were explored using 2-phospho-D-glycerate (2-PGA) as the primary substrate by monitoring the conversion of nicotinamide-adenine dinucleotide (NADH) into nicotinamide adenine dinucleotide (NAD). In addition, Csenolase exhibited active enzyme activity in catalytic reactions while the anti-Csenolase serum inhibited the enzyme activity. In vitro incubation experiments revealed that Csenolase might play key roles in the growth of the parasites. In conclusion, Csenolase is an important glycolytic enzyme required for the development of C. sinensis, and may be a potential vaccine candidate and drug target against C. sinensis infection.

Expression and immune response analysis of Schistosoma japonicum VAL-1, a homologue of vespid venom allergens.

Many parasites such as trematodes and nematodes have been found to express members of a gene family variously termed as venom allergen-like protein (VAL) or sperm-coating protein (SCP)-like protein. The molecular functions of these proteins remain unclear. We isolated the corresponding gene from Schistosoma japonicum, which we designated as Sj-VAL-1. The cDNA of Sj-VAL-1 contains an open reading frame encoding 221 amino acids, the first 25 residues being a putative secretion signal. RT-PCR analysis confirmed that Sj-VAL-1 was transcribed mainly in cercariae and eggs stages. Western blot analysis indicated that Sj-VAL-1 protein was an egg excretory-secretory products (ES products). Immunofluorescence showed it was secreted by eggs, head gland, and penetration glands of cercariae. In S. japonicum-infected mice, Sj-VAL-1-specific antibody significantly increased 6 weeks after infection and a higher level of IgG1 antibody contrast to IgG2a antibody indicated that a polarized Th2 immune response could be induced by Sj-VAL-1. These findings suggest that Sj-VAL-1 must play a role in the interaction between parasite and host. Its role as a potential modulator of immune function or as a pathogenic factor of granuloma formation in schistosomiasis needs further study.

[Financial burden of hepatitis B-related diseases and factors influencing the costs in Shenzhen, China].

OBJECTIVE: To investigate the direct, indirect and intangible costs due to hepatitis B-related diseases and to explore main factors associated with the costs in Shenzhen. METHODS: Cluster sampling for cases collected consecutively during the study period was administrated. Subjects were selected from eligible hepatitis B-related patients. By pre-trained professional investigators, health economics-related information was collected, using a structured questionnaire. Hospitalization expenses were obtained through hospital records after the patients were discharged from hospital. Total economic burden of hepatitis B-related patients would involve direct, indirect and intangible costs. Direct costs were further divided into direct medical costs and direct nonmedical costs. Human Capital Approach was employed to measure the indirect costs both on patients and the caregivers in 1-year time span. Willing to pay method was used to estimate the intangible costs. Multiple linear stepwise regression models were conducted to determine the factors linked to the economic burden. RESULTS: On average, the total annual cost of per patient with hepatitis B-related diseases was 81 590.23 RMB Yuan. Among which, direct, indirect and intangible costs were 30 914.79 Yuan (account for 37.9%), 15 258.01 Yuan (18.7%), 35 417.43 Yuan (43.4%), respectively. The total annual costs per patient for hepatocellular carcinoma, severe hepatitis B, decompensated cirrhosis, compensated cirrhosis, chronic hepatitis B and acute hepatitis B were 194 858.40 Yuan, 144 549.20 Yuan, 120 333.60 Yuan, 79 528.81 Yuan, 66 282.46 Yuan and 39 286.81 Yuan, respectively. The ratio of direct to indirect costs based on the base-case estimation foot add to 2.0:1, increased from hepato-cellular carcinoma (0.7:1) to compensated cirrhosis (3.5:1), followed by acute hepatitis B (3.3:1), severe hepatitis B (2.8:1), decompensate cirrhosis (2.3:1) and chronic hepatitis B (2.2:1). Direct medical costs were more than direct nonmedical. Ratio between the sum total was 16:1. The proportions of total annual cost per patient with hepatitis B-related diseases accounted for annual patient income were 285.3%, and 75.4% for annual household income. Furthermore, proportions of direct costs accounted for annual patient income and annual household income were 108.1% and 28.6%. The total annual indirect cost per person was 8123.38 Yuan for patients of all hepatitis B-related diseases, while 7134.63 Yuan for caregivers. Corresponding work-loss days were 55.74 days for patients and 19.83 days for caregivers. Based on multiple linear stepwise regression analysis, age of patients was a common influencing factor to all kinds of costs. Other factors were as follows: complicated with other diseases, antiviral medication, monthly household income and self-medications. CONCLUSION: The economic burden of hepatitis B-related diseases was substantial for patients and their families. All costs tended to increase with the severity of disease. The direct costs were larger than the indirect costs. And the direct medical costs were more than the direct ones. Indirect costs based on patients were larger than the ones of caregivers.