Thyroxine regulates pig Sertoli cell line proliferation and maturation through the IKK/NFκB and p38 MAPK signaling pathways.

The aim of this study was to investigate the signaling pathways involved in the proliferation and differentiation of pig Sertoli cells (SCs) mediated by thyroid hormone (T3) to provide a theoretical and practical basis for enhancing pig semen production. The effects of different concentrations of T3 on the proliferation of pig SCs were evaluated using the CCK8 assay. The impact of T3 on the proliferation and differentiation of pig SCs was further examined using RNA-seq, qPCR, and Western Blotting techniques. Additionally, the involvement of the p38 MAPK and NFκB pathways in mediating the effects of T3 on SCs proliferation and differentiation was investigated. Our findings revealed a strong correlation between the dosage of T3 and the inhibition of pig SCs proliferation and promotion of maturation. T3 regulated the activation state of the NFκB signaling pathway by upregulating IKKα, downregulating IKKß, and promoting IκB phosphorylation. Furthermore, T3 facilitated SCs maturation by upregulating AR and FSHR expression while downregulating KRT-18. In conclusion, T3 inhibits pig SCs proliferation and promote pig SCs maturation through the IKK/NFκB and p38 MAPK pathways. These findings provide valuable insights into the mechanisms by which T3 influences the proliferation and maturation of pig SCs.

Quality Evaluation of Guidelines for the Diagnosis and Treatment of Liver Failure.

OBJECTIVES: This study aimed to systematically assess the methodological quality and key recommendations of the guidelines for the diagnosis and treatment of liver failure (LF), furnishing constructive insights for guideline developers and equipping clinicians with evidence-based information to facilitate informed decision-making. DATA SOURCES: Electronic databases and manual searches from January 2011 to August 2023. STUDY SELECTION: Two reviewers independently screened titles and abstracts, then full texts for eligibility. Fourteen guidelines were included. DATA EXTRACTION AND SYNTHESIS: Two reviewers extracted data and checked by two others. Methodological quality of the guidelines was appraised using the Appraisal of Guidelines for Research and Evaluation II tool. Of the 14 guidelines, only the guidelines established by the Society of Critical Care Medicine and the American College of Gastroenterology (2023) achieved an aggregate quality score exceeding 60%, thereby meriting clinical recommendations. It emerged that there remains ample room for enhancement in the quality of the guidelines, particularly within the domains of stakeholder engagement, rigor, and applicability. Furthermore, an in-depth scrutiny of common recommendations and supporting evidence drawn from the 10 adult LF guidelines unveiled several key issues: controversy exists in the recommendation, the absence of supporting evidence and confusing use of evidence for recommendations, and a preference in evidence selection. CONCLUSIONS: There are high differences in methodological quality and recommendations among LF guidelines. Improving these existing problems and controversies will benefit existing clinical practice and will be an effective way for developers to upgrade the guidelines.

Effectiveness of Lianhua Qingwen Granule and Jingyin Gubiao Prescription in Omicron BA.2 Infection and Hospitalization: A Real-World Study of 56,244 Cases in Shanghai, China.

OBJECTIVE: To examine the effectiveness of Chinese medicine (CM) Lianhua Qingwen Granule (LHQW) and Jingyin Gubiao Prescription (JYGB) in asymptomatic or mild patients with Omicron infection in the shelter hospital. METHODS: This single-center retrospective cohort study was conducted in the largest shelter hospital in Shanghai, China, from April 10, 2022 to May 30, 2022. A total of 56,244 asymptomatic and mild Omicron cases were included and divided into 4 groups, i.e., non-administration group (23,702 cases), LHQW group (11,576 cases), JYGB group (12,112 cases), and dual combination of LHQW and JYGB group (8,854 cases). The length of stay (LOS) in the hospital was used to assess the effectiveness of LHQW and JYGB treatment on Omicron infection. RESULTS: Patients aged 41-60 years, with nadir threshold cycle (CT) value of N gene <25, or those fully vaccinated preferred to receive CM therapy. Before or after propensity score matching (PSM), the multiple linear regression showed that LHQW and JYGB treatment were independent influence factors of LOS (both P<0.001). After PSM, there were significant differences in LOS between the LHQW/JYGB combination and the other groups (P<0.01). The results of factorial design ANOVA proved that the LHQW/JYGB combination therapy synergistically shortened LOS (P=0.032). CONCLUSIONS: Patients with a nadir CT value <25 were more likely to accept CM. The LHQW/JYGB combination therapy could shorten the LOS of Omicron-infected individuals in an isolated environment.

Capsaicin pretreatment attenuates salt-sensitive hypertension by alleviating AMPK/Akt/Nrf2 pathway in hypothalamic paraventricular nucleus.

Background: Long term hypertension seriously promotes target organ damage in the brain and heart, and has increasingly become serious public health problem worldwide. The anti-hypertensive effects of capsaicin has been reported, however, the role and mechanism of capsaicin within the brain on salt-induced hypertension have yet to be elucidated. This study aimed to verify the hypothesis that capsaicin attenuates salt-induced hypertension via the AMPK/Akt/Nrf2 pathway in hypothalamic paraventricular nucleus (PVN). Methods: Dahl salt-sensitive (Dahl S) rats were used as animal model for the present study. Rats were randomly divided into four groups based on their dietary regimen (0.3% normal salt diet and 8% high salt diet) and treatment methods (infusion of vehicle or capsaicin in the PVN). Capsaicin was chronically administered in the PVN throughout the animal experiment phase of the study that lasted 6 weeks. Results: Our results demonstrated that PVN pretreatment with capsaicin can slow down raise of the blood pressure elevation and heart rate (HR) of Dahl S hypertensive rats given high salt diet. Interestingly, the cardiac hypertrophy was significantly improved. Furthermore, PVN pretreatment with capsaicin induced decrease in the expression of mRNA expression of NADPH oxidase-2 (NOX2), inducible nitric oxide synthase (iNOS), NOX4, p-IKKß and proinflammatory cytokines and increase in number of positive cell level for Nrf2 and HO-1 in the PVN of Dahl S hypertensive rats. Additionally, the protein expressions of phosphatidylinositol 3-kinase (p-PI3K) and phosphorylated protein kinase-B (p-AKT) were decreased, phosphorylated adenosine monophosphate-activated protein kinase (p-AMPK) were increased after the PVN pretreatment with capsaicin. Conclusion: Capsaicin pretreatment attenuates salt-sensitive hypertension by alleviating AMPK/Akt/iNOS pathway in the PVN.

A NAC transcription factor represses a module associated with xyloglucan content and regulates aluminum tolerance.

The transcriptional regulation of aluminum (Al) tolerance in plants is largely unknown, although Al toxicity restricts agricultural yields in acidic soils.. Here, we identified a NAM, ATAF1/2, and cup-shaped cotyledon 2 (NAC) transcription factor that participates in Al tolerance in Arabidopsis (Arabidopsis thaliana). Al substantially induced the transcript and protein levels of ANAC070, and loss-of-function anan070 mutants showed remarkably increased Al sensitivity, implying a beneficial role of ANAC070 in plant tolerance to Al toxicity. Further investigation revealed that more Al accumulated in the roots of anac070 mutants, especially in root cell walls, accompanied by a higher hemicellulose and xyloglucan level, implying a possible interaction between ANAC070 and genes that encode proteins responsible for the modification of xyloglucan, including xyloglucan endo-transglycosylases/hydrolase (XTH) or ANAC017. Yeast one hybrid analysis revealed a potential interaction between ANAC070 and ANAC017, but not for other XTHs. Furthermore, dual-luciferase reporter assay, RT-qPCR, and GUS analysis revealed that ANAC070 could directly repress the transcript levels of ANAC017, and knockout of ANAC017 in the anac070 mutant partially restored its Al sensitivity phenotype, indicating that ANAC070 contributes to Al tolerance mechanisms other than suppression of ANAC017 expression. Further analysis revealed that the core transcription factor SENSITIVE TO PROTON RHIZOTOXICITY1 (STOP1) and its target genes, which control Al tolerance in Arabidopsis, may also be involved in ANAC070-regulated Al tolerance. In summary, we identified a transcription factor, ANAC070, that represses the ANAC017-XTH31 module to regulate Al tolerance in Arabidopsis.

Microscopic Electric Rotary Grinding of Plaques Combined with Graft Repair in the Management of Peyronie's Disease.

Peyronie's Disease (PD) is clinically characterized by the development of localized fibrous plaques, primarily on the tunica albuginea, especially on the dorsal area of the penis. These plaques are the hallmark feature of this condition, resulting in penile curvature, deformity, and painful erections for affected individuals. Although various nonsurgical treatment options exist, their overall effectiveness is limited. As a result, surgical intervention has become the ultimate choice for patients with severe penile curvature deformities and associated erectile dysfunction. Our research team has successfully employed a combined approach involving microscopic electric rotary grinding of the fibrous plaques and the use of tunica vaginalis or bovine pericardium as graft materials for the repairing of the defects of tunica albuginea in the treatment of PD. This approach has consistently yielded highly satisfactory results regarding the restoration of penile shape, with excellent cosmetic results and significantly improved sexual satisfaction. This protocol aims to present a comprehensive surgical management strategy utilizing electric rotary grinding of the plaques and repairing the defects of tunica albuginea by using the tunica vaginalis, which represents an optimal surgical strategy for treating PD.

A Novel, Controllable, and Efficient Method for Building Highly Hydrophobic Aerogels.

Aerogels prepared using freeze-drying methods have the potential to be insulation materials or absorbents in the fields of industry, architecture, agriculture, etc., for their low heat conductivity, high specific area, low density, degradability, and low cost. However, their native, poor water resistance caused by the hydrophilicity of their polymer matrix limits their practical application. In this work, a novel, controllable, and efficient templating method was utilized to construct a highly hydrophobic surface for freeze-drying aerogels. The influence of templates on the macroscopic morphology and hydrophobic properties of materials was investigated in detail. This method provided the economical and rapid preparation of a water-resistant aerogel made from polyvinyl alcohol (PVA) and montmorillonite (MMT), putting forward a new direction for the research and development of new, environmentally friendly materials.

Virome diversity shaped by genetic evolution and ecological landscape of Haemaphysalis longicornis.

BACKGROUND: Haemaphysalis longicornis is drawing attentions for its geographic invasion, extending population, and emerging disease threat. However, there are still substantial gaps in our knowledge of viral composition in relation to genetic diversity of H. longicornis and ecological factors, which are important for us to understand interactions between virus and vector, as well as between vector and ecological elements. RESULTS: We conducted the meta-transcriptomic sequencing of 136 pools of H. longicornis and identified 508 RNA viruses of 48 viral species, 22 of which have never been reported. Phylogenetic analysis of mitochondrion sequences divided the ticks into two genetic clades, each of which was geographically clustered and significantly associated with ecological factors, including altitude, precipitation, and normalized difference vegetation index. The two clades showed significant difference in virome diversity and shared about one fifth number of viral species that might have evolved to "generalists." Notably, Bandavirus dabieense, the pathogen of severe fever with thrombocytopenia syndrome was only detected in ticks of clade 1, and half number of clade 2-specific viruses were aquatic-animal-associated. CONCLUSIONS: These findings highlight that the virome diversity is shaped by internal genetic evolution and external ecological landscape of H. longicornis and provide the new foundation for promoting the studies on virus-vector-ecology interaction and eventually for evaluating the risk of H. longicornis for transmitting the viruses to humans and animals. Video Abstract.

Anti-Microbial Effect of AgBr-NP@CTMAB on Streptococcus Mutans and Assessment of Surface Roughness Hardness and Flexural Strength of PMMA.

Purpose: To investigate the inhibition of Streptococcus mutans (S.mutans) and its biofilm by AgBr-nanoparticles (NP) @CTMAB (cetyltrimethyl-ammonium bromide) and evaluate the changes in Polymethyl methacrylate (PMMA)'s surface roughness (Ra), microhardness, and flexural strength during prolonged immersion in AgBr-NP@CTMAB for application in the denture cleaning industry. Patients and Methods: The antibacterial activity of AgBr-NP@CTMAB against S.mutans was measured colony formation assay, OD600 and laser confocal microscopy. Changes in the specimens' values for surface roughness, microhardness, and flexural strength (MPa) were measured after immersion solutions for 180 or 360 days. Results: The AgBr-NP@CTMAB solution exhibited a robust antibacterial effect on planktonic S. mutans, with a minimum bactericidal concentration of 5 µg/mL. The 10 µg/mL AgBr-NP@CTMAB solution efficiently inhibited S. mutans biofilm formation. (2) No significant difference in surface roughness after immersion in AgBr-NP@CTMAB (10 µg/mL and 20 µg/mL) comparing with distilled water (P > 0.05) and Polident had significantly higher than distilled water (P < 0.05). There was a significant decrease in the surface hardness of the PMMA specimens that were immersed in the Polident compared with those in distilled water (P < 0.05). While, no significant differences in surface hardness after immersion in the AgBr-NP@CTMAB (P > 0.05). The result of flexural strength suggested that there was no statistically significant difference (P < 0.05) between AgBr-NP@CTMAB as well as Polident and water. Conclusion: AgBrNP@CTMAB can efficiently inhibit the growth of plankton S.mutans and biofilm formation, without affecting the flexural strength, microhardness, or surface roughness of PMMA. Therefore, AgBrNP@CTMAB holds promise as a new denture cleaning agent.

Anti-necroptosis and anti-ferroptosis compounds from the Deep-Sea-Derived fungus Aspergillus sp. MCCC 3A00392.

Eight undescribed (1-8) and 46 known compounds (9-54) were isolated from the deep-sea-derived Aspergillus sp. MCCC 3A00392. Compounds 1-3 were three novel oxoindolo diterpenoids, 4-6 were three bisabolane sesquiterpenoids, while 7 and 8 were two monocyclic cyclopropanes. Their structures were established by exhaustive analyses of the HRESIMS, NMR, and theoretical calculations of the NMR data and ECD spectra. Compounds 10, 33, 38, and 39 were able to inhibit tumor necrosis factor (TNF)-induced necroptosis in murine L929 cell lines. Functional experiments verified that compounds 10 and 39 inhibited necroptosis by downregulating the phosphorylation of RIPK3 and MLKL. Moreover, compound 39 also reduced the phosphorylation of RIPK1. Compounds 10, 33, and 34 displayed potent inhibitory activities against RSL-3 induced ferroptosis with the EC50 value of 3.0 µM, 0.4 µM, and 0.1 µM, respectively. Compound 10 inhibited ferroptosis by the downregulation of HMOX1, while compounds 33 and 34 inhibited ferroptosis through regulation of NRF2/SLC7A11/GCLM axis. However, these compounds only showed weak effect in either the necroptosis or ferroptosis relative mouse disease models. Further studies of pharmacokinetics and pharmacodynamics might improve their in vivo bioactivities.

Changes in Cytokine Levels in Patients with Severe Fever with Thrombocytopenia Syndrome Virus.

Purpose: To characterize the cytokine profile of patients with severe fever with thrombocytopenia syndrome (SFTS) in relation to disease severity. Patients and Methods: 60 laboratory-confirmed SFTS patients and 12 healthy individuals from multi-centers in Shandong Province of China were included, and all patients were divided into fatal patients (9) and recovered patients (51) due to their final outcomes. Multiplex-microbead immunoassays were conducted to estimate levels of 27 cytokines in the sera of patients and controls. Results: The results showed that levels of IL-2, IL-4, IL-6, IL-7, IL-8, IL-15, IL-1RA, G-CSF, GM-CSF, IFN-γ, TNF-α, basic FGF, PDGF-BB, RANTES, IP-10, MIP-1α, MIP-1ß, MCP-1, and Eotaxin differed significantly among the SFTS fatal patients, recovered patients, and the healthy controls (all p<0.05). Compared to the healthy controls, the fatal patients and recovered patients had reduced levels of IL-2, IL-4, IL-7, PDGF-BB, RANTES, and Eotaxin, while the levels of PDGF-BB and RANTES were significantly lower in fatal patients compared to recovered patients. The increasing levels of IL-6, IL-8, IL-15, IL-1RA, G-CSF, GM-CSF, IFN-γ, TNF-α, basic FGF, IP-10, MIP-1α, MIP-1ß, and MCP-1 were observed in fatal patients (all p<0.05), and the levels of IL-6, IP-10, MIP-1α, and MCP-1 were significantly higher than other two groups. The Spearman correlation analysis indicated a positive correlation between platelet count and PDGF-BB levels (p<0.05), while the white blood cell count had a negative correlation with MIP-1 level (p<0.05). Conclusion: The research exhibited that the SFTS virus (SFTSV) caused an atypical manifestation of cytokines. The levels of IL-6, IP-10, MIP-1α, and MCP-1 had been observed a positive association with the severity of the illness.

Pathogenesis and Differentiated Treatment Strategies of Childhood Tic Disorders Based on WANG Xugao's “Thirty Methods of Treating the Liver” / 中医杂志

Based on WANG Xugao's “thirty methods of treating the liver”， it is believed that the occurrence and development of childhood tic disorders follow the dynamic progression from liver qi disease to liver fire disease and then liver wind disease. The basic pathogenesis of three stages are characterized by binding constraint of liver qi， liver fire hyperactivity， and internal stirring of liver wind. Moreover， liver-blood deficiency and stagnation， and malnutrition of liver yin as the main point in terms of the imbalance of liver qi， blood， yin， and yang should be considered， as well as the imbalance relationship of the five zang organs such as the involvement of other organs and the gradually reach of the other organs. Guided by the principles of “thirty methods of treating the liver”， the treatment of tic disorders in liver qi stage should focus on soothing the liver and rectifying qi， soothing the liver and unblocking the collaterals， using Xiaochaihu Decoction （小柴胡汤） and Sini Powder （四逆散）. The treatment of tic disorders in liver fire stage involves clearing， draining and resolving liver heat， using Longdan Xiegan Decoction （龙胆泻肝汤）， Xieqing Pill （泻青丸）， Danggui Longhui Pill （当归龙荟丸）， and Huagan Decoction （化肝煎）. The treatment of tic disorders in liver wind stage involves extinguishing wind and subduing yang， using Lingjiao Gouteng Decoction （羚角钩藤汤） and Liuwei Dihuang Pill （六味地黄丸）. Throughout the treatment process， attention should be paid to harmonizing the liver's qi， blood， yin， and yang， as well as addressing the pathology of other organs.

Exploring the Role of Wnt Ligands in Osteogenic Differentiation of Human Periodontal Ligament Stem Cells.

OBJECTIVES: This study aimed to investigate the functions of 19 types of Wnt ligands during the process of osteogenic differentiation in human periodontal ligament stem cells (hPDLSCs), with particular attention to WNT3A and WNT4. MATERIALS AND METHODS: The expression levels of 19 types of Wnt ligands were examined using real-time quantitative polymerase chain reaction (real-time qPCR) during hPDLSCs osteogenic differentiation at 7, 10, and 14 days. Knockdown of WNT3A and WNT4 expression was achieved using adenovirus vectors, and conditioned medium derived from WNT3A and WNT4 overexpression plasmids was employed to investigate their roles in hPDLSCs osteogenesis. Osteogenic-specific genes were analyzed using real-time qPCR. Alkaline phosphatase (ALP) and alizarin red S activities and staining were employed to assess hPDLSCs' osteogenic differentiation ability. RESULTS: During hPDLSCs osteogenic differentiation, the expression of 19 types of Wnt ligands varied, with WNT3A and WNT4 showing significant upregulation. Inhibiting WNT3A and WNT4 expression hindered hPDLSCs' osteogenic capacity. Conditioned medium of WNT3A promoted early osteogenic differentiation, while WNT4 facilitated late osteogenesis slightly. CONCLUSION: Wnt ligands, particularly WNT3A and WNT4, play an important role in hPDLSCs' osteogenic differentiation, highlighting their potential as promoters of osteogenesis. CLINICAL RELEVANCE: Given the challenging nature of alveolar bone regeneration, therapeutic strategies that target WNT3A and WNT4 signaling pathways offer promising opportunities. Additionally, innovative gene therapy approaches aimed at regulating of WNT3A and WNT4 expression hold potential for improving alveolar bone regeneration outcomes.

Clinical pharmacists' involvement in carbapenem antibiotics management at Wenzhou Integrated Hospital.

BACKGROUND: Carbapenem antibiotics are a pivotal solution for severe infections, particularly in hospital settings. The emergence of carbapenem-resistant bacteria owing to the irrational and extensive use of carbapenems underscores the need for meticulous management and rational use. Clinical pharmacists, with their specialized training and extensive knowledge, play a substantial role in ensuring the judicious use of carbapenem. This study aimed to elucidate the patterns of carbapenem use and shed light on the integral role played by clinical pharmacists in managing and promoting the rational use of carbapenem antibiotics at Wenzhou Integrated Traditional Chinese and Western Medicine Hospital. AIM: To analyze carbapenem use patterns in our hospital and role of clinical pharmacists in managing and promoting their rational use. METHODS: We performed a retrospective analysis of carbapenem use at our hospital between January 2019 and December 2021. Several key indicators, including the drug utilization index, defined daily doses (DDDs), proportion of antimicrobial drug costs to total hospitalization expenses, antibiotic utilization density, and utilization rates in different clinical departments were comprehensively analyzed. RESULTS: Between 2019 and 2021, there was a consistent decline in the consumption and sales of imipenem-cilastatin sodium, meropenem (0.3 g), and meropenem (0.5 g). Conversely, the DDDs of imipenem-cilastatin sodium for injection increased in 2020 and 2021 vs 2019, with a B/A value of 0.67, indicating a relatively higher drug cost. The DDDs of meropenem for injection (0.3 g) exhibited an overall upward trend, indicating an increasing clinical preference. However, the B/A values for 2020 and 2021 were both > 1, suggesting a relatively lower drug cost. The DDDs of meropenem for injection (0.5 g) demonstrated a progressive increase annually and consistently ranked first, indicating a high clinical preference with a B/A value of 1, signifying good alignment between economic and social benefits. CONCLUSION: Carbapenem use in our hospital was generally reasonable with a downward trend in consumption and sales over time. Clinical pharmacists play a pivotal role in promoting appropriate use of carbapenems.

Cascading effects of hypobaric hypoxia on the testis: insights from a single-cell RNA sequencing analysis.

Most mammals tolerate exposure to hypobaric hypoxia poorly as it may affect multiple regulatory mechanisms and inhibit cell proliferation, promote apoptosis, limit tissue vascularization, and disrupt the acid-base equilibrium. Here, we quantified the functional state of germ cell development and demonstrated the interaction between the germ and somatic cells via single-cell RNA sequencing (scRNA-seq). The present study elucidated the regulatory effects of hypobaric hypoxia exposure on germ cell formation and sperm differentiation by applying enrichment analysis to genomic regions. Hypobaric hypoxia downregulates the genes controlling granule secretion and organic matter biosynthesis, upregulates tektin 1 (TEKT1) and kinesin family member 2C (KIF2C), and downregulates 60S ribosomal protein 11 (RPL11) and cilia- and flagella-associated protein 206 (CFAP206). Our research indicated that prosaposin-G protein-coupled receptor 37 (PSAP-GPR37) ligands mediate the damage to supporting cells caused by hypobaric hypoxic exposure. The present work revealed that hypoxia injures peritubular myoid (PTM) cells and spermatocytes in the S phase. It also showed that elongating spermatids promote maturation toward the G2 phase and increase their functional reserve for sperm-egg binding. The results of this study provide a theoretical basis for future investigations on prophylactic and therapeutic approaches toward protecting the reproductive system against the harmful effects of hypobaric hypoxic exposure.

Chemical Constituents of the Deep-sea Gammarid Shrimp-Derived Fungus Penicillium citrinum XIA-16.

One new alkaloid, (S)-2-acetamido-4-(2-(methylamino)phenyl)-4-oxobutanoic acid (1), was isolated from the deep-sea-derived Penicillium citrinum XIA-16, together with 25 known compounds including ten polyketones (2-11), eight alkaloids (12-19), six steroids (20-25), and a fatty acid (26). Their planar and relative structures were determined by an analysis of 1D and 2D nuclear magnetic resonance (NMR) as well as high resolution electrospray ionization mass spectroscopy (HR-ESI-MS) data. The absolute configuration of 1 was determined by comparison of the experimental and calculated electronic circular dichroism (ECD) spectra. Penicitrinol B (6) significantly inhibited RSL3-induced ferroptosis (EC50 =2.0 µM) by reducing lipid peroxidation and heme oxygenase 1 (HMOX1) expression. Under the concentration of 10 µM, penicitrinol A (7) was able to inhibit cuproptosis with the cell viabilities of 68.2 % compared to the negative control (copper and elesclomol) with the cell viabilities of 14.8 %.

Multiplex gene editing reduces oxalate production in primary hyperoxaluria type 1.

Targeting key enzymes that generate oxalate precursors or substrates is an alternative strategy to eliminate primary hyperoxaluria type I (PH1), the most common and life-threatening type of primary hyperoxaluria. The compact Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) from the Prevotella and Francisella 1 (Cpf1) protein simplifies multiplex gene editing and allows for all-in-one adeno-associated virus (AAV) delivery. We hypothesized that the multiplex capabilities of the Cpf1 system could help minimize oxalate formation in PH1 by simultaneously targeting the hepatic hydroxyacid oxidase 1 ( Hao1) and lactate dehydrogenase A ( Ldha) genes. Study cohorts included treated PH1 rats ( Agxt Q84X rats injected with AAV-AsCpf1 at 7 days of age), phosphate-buffered saline (PBS)-injected PH1 rats, untreated PH1 rats, and age-matched wild-type (WT) rats. The most efficient and specific CRISPR RNA (crRNA) pairs targeting the rat Hao1 and Ldha genes were initially screened ex vivo. In vivo experiments demonstrated efficient genome editing of the Hao1 and Ldha genes, primarily resulting in small deletions. This resulted in decreased transcription and translational expression of Hao1 and Ldha. Treatment significantly reduced urine oxalate levels, reduced kidney damage, and alleviated nephrocalcinosis in rats with PH1. No liver toxicity, ex-liver genome editing, or obvious off-target effects were detected. We demonstrated the AAV-AsCpf1 system can target multiple genes and rescue the pathogenic phenotype in PH1, serving as a proof-of-concept for the development of multiplex genome editing-based gene therapy.

Systematic Appraisal of Guidelines for the Diagnosis and Treatment of Choledocholithiasis.

BACKGROUND: To systematically evaluate the methodological quality of the current up-to-date guidelines pertaining to choledocholithiasis, we conducted a comprehensive analysis of key recommendations and corresponding evidence, focusing on the heterogeneity among these guidelines. METHOD: Systematic searches across various databases were performed to identify the latest guidelines. The identified guidelines, which met the inclusion criteria, underwent evaluation using the Appraisal of Guidelines for Research and Evaluation II (AGREE II) tool. The key recommendations and evidence from the included guidelines were extracted and reclassified using the Oxford Centre for Evidence-Based Medicine (OCEBM) grading system, and the obtained results were analyzed. RESULTS: Nine guidelines related to choledocholithiasis were included in this study, out of which 4 achieved an overall standardized score of more than 60%, indicating their suitability for recommendation. Upon closer examination of the main recommendations within these guidelines, we discovered significant discrepancies concerning the utilization of similar treatment techniques for different diseases or different treatment methods under comparable conditions, and discrepancies in the recommended treatment duration. High-quality research evidence was lacking, and some recommendations either failed to provide supporting evidence or cited inappropriate and low-level evidence. CONCLUSION: The quality of guidelines pertaining to choledocholithiasis is uneven. Recommendations for the treatment of choledocholithiasis demonstrate considerable disparities among the guidelines, particularly regarding the utilization of endoscopic retrograde cholangiopancreatography as a treatment method and the management approaches for difficult stone cases. Improvements by guideline developers for these factors contributing to the heterogeneity would be a reasonable approach to further update the guidelines for cholangiolithiasis.

Antibody responses following the surge of SARS-CoV-2 Omicron infection among patients with systemic autoimmune rheumatic diseases.

Objectives: The surge of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variant Omicron infections has affected most Chinese residents at the end of 2022, including a number of patients with systemic autoimmune rheumatic diseases (SARDs). Methods: To investigate the antibody level of the Omicron variant in SARD patients after SARS-CoV-2 Omicron infection, we tested BA.5.2 and BF.7 Omicron variant IgG antibody levels using ELISA on blood samples collected from 102 SARD patients and 19 healthy controls (HCs). The type of SARD, demographics, concurrent treatment, doses of SARS-CoV-2 vaccines and outcomes were also recorded. Results: A total of 102 SARD patients (mean age: 40.3 years; 89.2% female), including 60 SLE, 32 RA and 10 other SARDs, were identified. Of these, 87 (85.3%) were infected with SARS-CoV-2. We found that the BA.5.2 and BF.7 antibody levels of infected SARD patients were lower than those of HCs (P < 0.05). Sixty-five (63.7%) patients had at least one dose of a SARS-CoV-2 vaccine. SARD patients with at least two doses of SARS-CoV-2 vaccine had a higher level of BA.5.2 and BF.7 antibodies than the unvaccinated group (P < 0.05). There was no evidence for a significant inhibitory effect of glucocorticoids (GCs) on the BA.5.2 and BF.7 Omicron variant antibody levels in SARD patients. SLE patients using biologic DMARDs had a lower BA.5.2 Omicron variant antibody level than patients using GCs and/or HCQ. Conclusion: These data suggest that patients with SARDs had a lower antibody response than HCs after Omicron infection.

Downregulated calmodulin expression contributes to endothelial cell impairment in diabetes.

Endothelial dysfunction, a central hallmark of cardiovascular pathogenesis in diabetes mellitus, is characterized by impaired endothelial nitric oxide synthase (eNOS) and NO bioavailability. However, the underlying mechanisms remain unclear. Here in this study, we aimed to identify the role of calmodulin (CaM) in diabetic eNOS dysfunction. Human umbilical vein endothelial cells and murine endothelial progenitor cells (EPCs) treated with high glucose (HG) exhibited downregulated CaM mRNA/protein and vascular endothelial growth factor (VEGF) expression with impeded eNOS phosphorylation and cell migration/tube formation. These perturbations were reduplicated in CALM1-knockdown cells but prevented in CALM1-overexpressing cells. EPCs from type 2 diabetes animals behaved similarly to HG-treated normal EPCs, which could be rescued by CALM1-gene transduction. Consistently, diabetic animals displayed impaired eNOS phosphorylation, endothelium-dependent dilation, and CaM expression in the aorta, as well as deficient physical interaction of CaM and eNOS in the gastrocnemius. Local CALM1 gene delivery into a diabetic mouse ischemic hindlimb improved the blunted limb blood perfusion and gastrocnemius angiogenesis, and foot injuries. Diabetic patients showed insufficient foot microvascular autoregulation, eNOS phosphorylation, and NO production with downregulated CaM expression in the arterial endothelium, and abnormal CALM1 transcription in genome-wide sequencing analysis. Therefore, our findings demonstrated that downregulated CaM expression is responsible for endothelium dysfunction and angiogenesis impairment in diabetes, and provided a novel mechanism and target to protect against diabetic endothelial injury.