Syndecan-4 is required for early-stage repair responses during zebrafish heart regeneration.

BACKGROUND: The healing process after a myocardial infarction (MI) in humans involves complex events that replace damaged tissue with a fibrotic scar. The affected cardiac tissue may lose its function permanently. In contrast, zebrafish display a remarkable capacity for scar-free heart regeneration. Previous studies have revealed that syndecan-4 (SDC4) regulates inflammatory response and fibroblast activity following cardiac injury in higher vertebrates. However, whether and how Sdc4 regulates heart regeneration in highly regenerative zebrafish remains unknown. METHODS AND RESULTS: This study showed that sdc4 expression was differentially regulated during zebrafish heart regeneration by transcriptional analysis. Specifically, sdc4 expression increased rapidly and transiently in the early regeneration phase upon ventricular cryoinjury. Moreover, the knockdown of sdc4 led to a significant reduction in extracellular matrix protein deposition, immune cell accumulation, and cell proliferation at the lesion site. The expression of tgfb1a and col1a1a, as well as the protein expression of Fibronectin, were all down-regulated under sdc4 knockdown. In addition, we verified that sdc4 expression was required for cardiac repair in zebrafish via in vivo electrocardiogram analysis. Loss of sdc4 expression caused an apparent pathological Q wave and ST elevation, which are signs of human MI patients. CONCLUSIONS: Our findings support that Sdc4 is required to mediate pleiotropic repair responses in the early stage of zebrafish heart regeneration.

The biological age model for evaluating the degree of aging in centenarians.

BACKGROUND: Biological age (BA) has been used to assess individuals' aging conditions. However, few studies have evaluated BA models' applicability in centenarians. METHODS: Important organ function examinations were performed in 1798 cases of the longevity population (80∼115 years old) in Hainan, China. Eighty indicators were selected that responded to nutritional status, cardiovascular function, liver and kidney function, bone metabolic function, endocrine system, hematological system, and immune system. BA models were constructed using multiple linear regression (MLR), principal component analysis (PCA), Klemera and Doubal method (KDM), random forest (RF), support vector machine (SVM), extreme gradient boosting (XGBoost), and light gradient boosting machine (lightGBM) methods. A tenfold crossover validated the efficacy of models. RESULTS: A total of 1398 participants were enrolled, of whom centenarians accounted for 49.21%. Seven aging markers were obtained, including estimated glomerular filtration rate, albumin, pulse pressure, calf circumference, body surface area, fructosamine, and complement 4. Eight BA models were successfully constructed, namely MLR, PCA, KDM1, KDM2, RF, SVM, XGBoost and lightGBM, which had the worst R2 of 0.45 and the best R2 of 0.92. The best R2 for cross-validation was KDM2 (0.89), followed by PCA (0.62). CONCLUSION: In this study, we successfully applied eight methods, including traditional methods and machine learning, to construct models of biological age, and the performance varied among the models.

Peritoneal Dialysis Care in Mainland China: Nationwide Survey.

BACKGROUND: Peritoneal dialysis (PD) care in mainland China has been progressing in the past 10 years. OBJECTIVE: To complement information from the dialysis registry, a large-scale nationwide survey was conducted to investigate the current infrastructure and management of PD care at hospitals of different tiers. METHODS: A web-based multiple-choice questionnaire was distributed through the National Center for Nephrology Medical Quality Management and Control to PD centers of secondary and tertiary hospitals in October 2020. The 2-part survey collected the information of PD centers and the clinical management of patients on PD. A total of 788 effective surveys from 746 hospitals were voluntarily returned, and data were extracted and analyzed. RESULTS: The effective survey data covered 101,537 patients on PD, with 95% (96,460/101,537) in the tertiary hospitals. The median number of patients per PD center was 60 (IQR 21-152); this number was 32 (IQR 8-65) and 70 (IQR 27-192) for secondary and tertiary hospitals, respectively. There was a discrepancy in the availability of designated physical areas for different functions of PD care between the secondary and tertiary hospitals. The proportion of tertiary hospitals with PD training (P=.01), storage (P=.09), and procedure area (P<.001) was higher compared to secondary hospitals. PD catheter placement was performed in 96% (608/631) of the PD centers in tertiary hospitals, which was significantly higher compared to 86% (99/115) in secondary hospitals (P<.001). Automated PD was available in 55% (347/631) of the tertiary hospitals, which was significantly higher than that in secondary hospitals (37/115, 32%) according to the survey (P<.001). The most commonly performed PD module was continuous ambulatory peritoneal dialysis (772/788, 98%), followed by intermittent peritoneal dialysis (543/788, 69%). The overall reported nocturnal intermittent peritoneal dialysis was 31% (244/788); it was 28% (220/788) for continuous cycling peritoneal dialysis and 15% (118/788) for tidal peritoneal dialysis. Comparisons between the secondary and tertiary hospitals revealed no significant differences in prophylactic antibiotic use for PD catheter placement and therapeutic use for peritonitis. The first peritoneal equilibrium test was conducted in 58% (454/788) of patients at 4-6 weeks after initiation of PD, and 91% (718/788) reported at least one peritoneal equilibrium test per year. Overall, 79% (570/722) and 65% (469/722) of PD centers performed assessment for dialysis adequacy and residual kidney function, respectively; and 87% (685/788) of patients on PD were followed every 1 to 3 months for laboratory and auxiliary examinations. CONCLUSIONS: This national survey reflects the current status and disparities of PD center management in mainland China. The study results suggest that the PD care needs to be more conveniently accessible in secondary hospitals, and quality management and staff training in secondary hospitals are still in high demand.

Evolving peritoneal dialysis care in Chinese mainland from 2010 to 2020: Comparison data from two surveys.

INTRODUCTION: Along with the peritoneal dialysis (PD)-favored policy in China and the implementation of more comprehensive PD management, PD has evolved in Chinese mainland over the last decade. Despite the existence of national registries and several provincial epidemiological descriptive studies, there was almost no national research on the changing trajectory in PD population. A comparison study, based on two national surveys that were 10 years apart, was conducted to reveal the evolvement of PD care in Chinese mainland. METHODS: Two national surveys have been done respectively in 2010 and 2020 to capture the epidemiological status, application of different modalities, management of perioperative infection, and long-term complications among PD patients. RESULTS: In the study with 730 participating hospitals (n = 14,912 PD patients) in 2010 and 746 hospitals (n = 101,537) in 2020, prevalent PD patients have increased in the past 10 years with increased numbers of PD patients in both secondary (average 5 ± 16 vs. 43 ± 41, p < 0.01) and tertiary hospitals (32 ± 53 vs. 153 ± 215, p < 0.01). Automated PD has been accessible in 0.4% of all hospitals, only in tertiary centers in 2010 and its application increased to 51% in 2020. PD centers have become more engaged in PD catheter placement, treated properly for the PD-related infection, and carried out the follow-up in compliance with the national protocols. CONCLUSIONS: Our study indicates that over the past decade, the prevalent PD population has quickly expanded with increased APD availability in Chinese mainland. The management of PD patients has become better conforming to the guidelines and long-term follow-up of patients have remained stable. Further studies are warranted to evaluate whether the rapidly changing paradigm of PD could translate into the socio-economic benefits in the society.

Associations of Serum 25(OH)D, PTH, and ß-CTX Levels with All-Cause Mortality in Chinese Community-Dwelling Centenarians.

This longitudinal cohort study explored the associations of 25-hydroxyvitamin D [25(OH)D], parathyroid hormone (PTH), and ß-C-terminal telopeptide of type 1 collagen (ß-CTX) levels with all-cause mortality in centenarians. The study included 952 centenarians (81.4% female). During a median follow-up of 32 months, 752 (78.9%) centenarians died. The estimated 1-year, 3-year, and 5-year survival rates were 80.0%, 45.7%, and 23.6%, respectively. The association of mortality with 25(OH)D was linear, whereas the associations with PTH and ß-CTX were J-shaped, with a lower risk below the median levels. Compared with 25(OH)D of ≥30 ng/mL, 25(OH)D < 30 ng/mL was associated with increased mortality (HR 1.52, 95% CI 1.24−1.86, p < 0.001). Compared with PTH of ≤65 pg/mL, PTH > 65 pg/mL was associated with increased mortality (HR 1.30, 95% CI 1.08−1.56, p = 0.005). Compared with ß-CTX of <0.55 ng/mL, ß-CTX ≥ 0.55 ng/mL was associated with increased mortality (HR 1.30, 95% CI 1.10−1.54, p = 0.002). A higher ß-CTX level (even in the clinical reference range of 0.55−1.01 ng/mL) was associated with increased mortality (HR 1.23, 95% CI 1.04−1.47, p = 0.018). Centenarians with 25(OH)D < 30 ng/mL, PTH > 65 pg/mL, and ß-CTX ≥ 0.55 ng/mL had a 2.77-fold (95% CI 1.99−3.85, p < 0.001) increased risk of mortality when compared with those with 25(OH)D of >30 ng/mL, PTH < 65 pg/mL, and ß-CTX < 0.55 ng/mL. Lower serum 25(OH)D and higher PTH and ß-CTX were independently correlated with increased all-cause mortality in Chinese community-dwelling centenarians.

Multi-Task Learning-Based Immunofluorescence Classification of Kidney Disease.

Chronic kidney disease is one of the most important causes of mortality worldwide, but a shortage of nephrology pathologists has led to delays or errors in its diagnosis and treatment. Immunofluorescence (IF) images of patients with IgA nephropathy (IgAN), membranous nephropathy (MN), diabetic nephropathy (DN), and lupus nephritis (LN) were obtained from the General Hospital of Chinese PLA. The data were divided into training and test data. To simulate the inaccurate focus of the fluorescence microscope, the Gaussian method was employed to blur the IF images. We proposed a novel multi-task learning (MTL) method for image quality assessment, de-blurring, and disease classification tasks. A total of 1608 patients' IF images were included-1289 in the training set and 319 in the test set. For non-blurred IF images, the classification accuracy of the test set was 0.97, with an AUC of 1.000. For blurred IF images, the proposed MTL method had a higher accuracy (0.94 vs. 0.93, p < 0.01) and higher AUC (0.993 vs. 0.986) than the common MTL method. The novel MTL method not only diagnosed four types of kidney diseases through blurred IF images but also showed good performance in two auxiliary tasks: image quality assessment and de-blurring.

Establishment of a New Zealand White Rabbit Model for Lethal Toxin (LT) Challenge and Efficacy of Monoclonal Antibody 5E11 in the LT-Challenged Rabbit Model.

Anthrax caused by Bacillus anthracis is a lethal infectious disease, especially when inhaled, and the mortality rate approaches 100% without treatment. The anthrax antitoxin monoclonal antibody (MAb) 5E11 is a humanized antibody that targets the anthrax protective antigen (PA). The efficacy of 5E11 needs proper animal models. However, anthrax spores are extremely dangerous, so experiments must be conducted under Biosafety Level 3 conditions. Considering the critical effects of lethal toxin (LT) on hosts during infection, we report the establishment of a LT-challenged rabbit model, which caused 100% mortality with a dose of 2 mg PA + 1 mg LF, while a 4 mg PA + 2 mg LF challenge could limit death to within three days. Then, we evaluated 5E11 efficacy against LT. A prophylactic study showed that the i.v. administration of 40 mg/kg 5E11 four days before lethal dose LT challenge could lead to 100% survival. In therapeutic studies, the i.v. administration of 40 mg/kg 5E11 10 min after lethal dose LT challenge could provide complete protection. Overall, we developed a new LT-challenged rabbit model, and our results indicate that 5E11 shows potential for the clinical application in anthrax treatment.

Identification and characterization of a neutralizing monoclonal antibody that provides complete protection against Yersinia pestis.

Yersinia pestis (Y. pestis) has caused an alarming number of deaths throughout recorded human history, and novel prophylactics and therapeutics are necessary given its potential as a bioweapon. Only one monoclonal antibody has been identified to date that provides complete protection against Y. pestis. Here, we describe a second novel murine monoclonal antibody (F2H5) that provided complete protection against Y. pestis 141 infection when administered prophylactically to Balb/c mice (100 µg intravenously). We humanized F2H5, characterized its ability to bind to the Y. pestis F1 protein and further characterized the neutralizing epitope using computational and experimental approaches. While Western blot results suggested a linear epitope, peptide mapping using ELISA failed to identify an epitope, suggesting a conformational epitope instead. We adopted a computational approach based on Residue Contact Frequency to predict the site of antigen-antibody interaction and defined the F2H5/F1 binding site computationally. Based on computational approach, we determined that residues G104E105N106 in F1 were critical to F2H5 binding and that CDRH2 and CDRH3 of F2H5 interacted with F1. Our results show that combining computational approach and experimental approach can effectively identify epitopes.

[Screening of full human anthrax lethal factor neutralizing antibody in transgenic mice].

Anthrax is a highly lethal infectious disease caused by the spore-forming bacterium Bacillus anthracis. The major virulence factor of B. anthracis consists of protective antigen (PA), lethal factor (LF) and edema factor (EF). PA binds with LF to form lethal toxin (LT), and PA binds with EF to form edema toxin (ET). Antibiotics is hard to work in advanced anthrax infections, because injuries and deaths of the infected are mainly caused by lethal toxin (LT). Thus, the therapeutic neutralizing antibody is the most effective treatment of anthrax. Currently most of the anthrax toxin antibodies are monoclonal antibodies (MAbs) for PA and US FDA has approved ABTHRAX humanized PA monoclonal antibody for the treatment of inhalational anthrax. Once B. anthracis was artificially reconstructed or PA had mutations within recognized neutralization epitopes, anti-PA MAbs would no longer be effective. Therefore, anti-LF MAbs is an important supplement for anthrax treatment. Most of the anti-LF antibodies are murine or chimeric antibodies. By contrast, fully human MAbs can avoid the high immunogenicity of murine antibodies. First, we used LF to immunize the transgenic mice and used fluorescent cell sorting to get antigen-specific memory B cells from transgenic mice spleen lymphocytes. By single cell PCR method, we quickly found two strains of anti-LF MAbs with binding activity, 1D7 and 2B9. Transiently transfected Expi 293F cells to obtain MAbs protein after purification. Both 1D7 and 2B9 efficiently neutralized LT in vitro, and had good synergistic effect when mixed with anti-PA MAbs. In summary, combining the advantages of transgenic mice, fluorescent cell sorting and single-cell PCR methods, this study shows new ideas and methods for the rapid screening of fully human monoclonal antibodies.

Generation and Characterization of Human Monoclonal Antibodies Targeting Anthrax Protective Antigen following Vaccination with a Recombinant Protective Antigen Vaccine.

The anthrax protective antigen (PA) is the central component of the three-part anthrax toxin, and it is the primary immunogenic component in the approved AVA anthrax vaccine and the "next-generation" recombinant PA (rPA) anthrax vaccines. Animal models have indicated that PA-specific antibodies (AB) are sufficient to protect against infection with Bacillus anthracis. In this study, we investigated the PA domain specificity, affinity, mechanisms of neutralization, and synergistic effects of PA-specific antibodies from a single donor following vaccination with the rPA vaccine. Antibody-secreting cells were isolated 7 days after the donor received a boost vaccination, and 34 fully human monoclonal antibodies (hMAb) were identified. Clones 8H6, 4A3, and 22F1 were able to neutralize lethal toxin (LeTx) both in vitro and in vivo. Clone 8H6 neutralized LeTx by preventing furin cleavage of PA in a dose-dependent manner. Clone 4A3 enhanced degradation of nicked PA, thereby interfering with PA oligomerization. The mechanism of 22F1 is still unclear. A fourth clone, 2A6, that was protective only in vitro was found to be neutralizing in vivo in combination with a toxin-enhancing antibody, 8A7, which binds to domain 3 of PA and PA oligomers. These results provide novel insights into the antibody response elicited by the rPA vaccine and may be useful for PA-based vaccine and immunotherapeutic cocktail design.