The regulatory role of m6A modification in the maintenance and differentiation of embryonic stem cells.

As the most prevalent and reversible internal epigenetic modification in eukaryotic mRNAs, N 6-methyladenosine (m6A) post-transcriptionally regulates the processing and metabolism of mRNAs involved in diverse biological processes. m6A modification is regulated by m6A writers, erasers, and readers. Emerging evidence suggests that m6A modification plays essential roles in modulating the cell-fate transition of embryonic stem cells. Mechanistic investigation of embryonic stem cell maintenance and differentiation is critical for understanding early embryonic development, which is also the premise for the application of embryonic stem cells in regenerative medicine. This review highlights the current knowledge of m6A modification and its essential regulatory contribution to the cell fate transition of mouse and human embryonic stem cells.

Characteristics of China's coal mine methane emission sources at national and provincial levels.

Methane (CH4) is the second most abundant greenhouse gas. China is the largest CH4 emitter in the world, with coal mine methane (CMM) being one of the main anthropogenic contributions. Thus, there is an urgent need for comprehensive estimates and strategies for reducing CMM emissions in China. However, the development of effective strategies is currently challenged by a lack of information on temporal variations in the contributions of different CMM sources and the absence of provincial spatial analysis. Here, considering five sources and utilization, we build a comprehensive inventory of China's CMM emissions from 1980 to 2022 and quantify the contributions of individual sources to the overall CMM emissions at the national and provincial levels. Our results highlight a significant shift in the source contributions of CMM emissions, with the largest contributor, underground mining, decreasing from 89% in 1980 to 69% in 2022. Underground abandoned coal mines, which were ignored or underestimated in past inventories, have become the second source of CMM emissions since 1999. From 2011 to 2022, we identified Shanxi, Guizhou, and Shaanxi as the three largest CMM-emitting provinces, while the Emissions Database for Global Atmospheric Research (EDGAR) v8 overestimated emissions from Inner Mongolia, ranking it third. Notably, we observed a substantial decrease (exceeding 1 Mt) in CMM emissions in Sichuan, Henan, Liaoning, and Hunan between 2011 and 2022, which was not captured by EDGAR v8. To develop targeted CMM emission reduction strategies at the provincial level, we classified 31 provinces into four groups based on their CMM emission structures. In 2022, the number of provinces with CMM emissions mainly from abandoned coal mines has exceeded that of provinces with mainly underground mines, which requires attention. This study reveals the characteristics of the source of CMM emissions in China and provides emission reduction directions for four groups of provinces.

Deletion of Bmal1 in aggrecan-expressing cells leads to mouse temporomandibular joint osteoarthritis.

INTRODUCTION: Articular cartilage is the major affected tissue during the development of osteoarthritis (OA) in temporomandibular joint (TMJ). The core circadian rhythm molecule Bmal1 regulates chondrocyte proliferation, differentiation and apoptosis; however, its roles in condylar cartilage function and in TMJ OA have not been fully elucidated. MATERIALS AND METHODS: TMJ OA mouse model was induced by unilateral anterior crossbite (UAC) and Bmal1 protein expression in condylar cartilage were examined by western blot analysis. To determine the role of Bmal1 in TMJ OA, we generated cartilage-specific Bmal1 conditional knockout (cKO) mice (Bmal1Agc1CreER mice) and hematoxylin and eosin staining, toluidine blue and Safranin O/fast green, immunohistochemistry, TUNEL assay, real-time PCR analysis and Western blot assay were followed. RESULTS: Bmal1 expression was reduced in condylar cartilage in a TMJ OA mouse model induced by UAC. The Bmal1 cKO mice displayed decreased cartilage matrix synthesis, reduced chondrocyte proliferation, increased chondrocyte hypertrophy and apoptosis as well as the upregulation of YAP expression in TMJ condylar cartilage. CONCLUSIONS: We demonstrated that Bmal1 was essential for TMJ tissue homeostasis and loss-of-function of Bmal1 in chondrocytes leads to the development of TMJ OA.

Widely Targeted Metabolomic Analysis Reveals the Improvement in Panax notoginseng Triterpenoids Triggered by Arbuscular Mycorrhizal Fungi via UPLC-ESI-MS/MS.

Panax notoginseng is a highly valued perennial medicinal herb in China and is widely used in clinical treatments. The main purpose of this study was to elucidate the changes in the composition of P. notoginseng saponins (PNSs), which are the main bioactive substances, triggered by arbuscular mycorrhizal fungi (AMF) via ultrahigh-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UPLC-ESI-MS/MS). A total of 202 putative terpenoid metabolites were detected, of which 150 triterpene glycosides were identified, accounting for 74.26% of the total. Correlation analysis, principal component analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS-DA) of the metabolites revealed that the samples treated with AMF (group Ce) could be clearly separated from the CK samples. In total, 49 differential terpene metabolites were identified between the Ce and CK groups, of which 38 and 11 metabolites were upregulated and downregulated, respectively, and most of the upregulated differentially abundant metabolites were mainly triterpene glycosides. The relative abundances of the two major notoginsenosides (MNs), ginsenosides Rd and Re, and 13 rare notoginsenosides (RNs), significantly increased. The differential saponins, especially RNs, were more easily clustered into one branch and had a high positive correlation. It could be concluded that the biosynthesis and accumulation of some RNs share the same pathways as those triggered by AMF. This study provides a new way to obtain more notoginsenoside resources, particularly RNs, and sheds new light on the scientization and rationalization of the use of AMF agents in the ecological planting of medicinal plants.

Pipette-tip solid-phase extraction coupled with matrix-assisted laser desorption/ionization mass spectrometry enables rapid and high-throughput analysis of antidepressants in rat serum.

Therapeutic drug monitoring is essential for ensuring the efficacy and safety of medications. This study introduces a streamlined approach that combines pipette-tip solid-phase extraction (PT-SPE) with matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS), facilitating rapid and high-throughput monitoring of drug concentrations. As a demonstration, this method was applied to the extraction and quantification of antidepressants in serum. Utilizing Zip-Tip C18, the method enabled the extraction of antidepressants from complex biological matrices in less than 2 min, with the subsequent MALDI-MS analysis yielding results in just 1 min. Optimal extraction recoveries were achieved using a sampling solution at pH 9.0 and a 10 µL ethanol desorption solution containing 0.1% phosphoric acid. For MALDI analysis, 2,5-dihydroxybenzoic acid was identified as the most effective matrix for producing the highest signal intensity. The quantification strategy exhibited robust linearities (R2 ≥ 0.997) and satisfactory limits of quantification, ranging from 0.05 to 0.5 µg/mL for a suite of antidepressants. The application for monitoring dynamic concentration changes of antidepressants in rat serum emphasized the method's efficacy. This strategy offers the advantages of high throughput, minimal sample usage, environmental sustainability, and simplicity, providing ideas and a reference basis for the subsequent development of methods for therapeutic drug monitoring.

Effect of a systemic intervention combined with a psychological intervention in stroke patients with oropharyngeal dysfunction.

BACKGROUND: Stroke frequently results in oropharyngeal dysfunction (OD), leading to difficulties in swallowing and eating, as well as triggering negative emotions, malnutrition, and aspiration pneumonia, which can be detrimental to patients. However, routine nursing interventions often fail to address these issues adequately. Systemic and psychological interventions can improve dysphagia symptoms, relieve negative emotions, and improve quality of life. However, there are few clinical reports of systemic interventions combined with psychological interventions for stroke patients with OD. AIM: To explore the effects of combining systemic and psychological interventions in stroke patients with OD. METHODS: This retrospective study included 90 stroke patients with OD, admitted to the Second Affiliated Hospital of Qiqihar Medical College (January 2022-December 2023), who were divided into two groups: regular and coalition. Swallowing function grading (using a water swallow test), swallowing function [using the standardized swallowing assessment (SSA)], negative emotions [using the self-rating anxiety scale (SAS) and self-rating depression scale (SDS)], and quality of life (SWAL-QOL) were compared between groups before and after the intervention; aspiration pneumonia incidence was recorded. RESULTS: Post-intervention, the coalition group had a greater number of patients with grade 1 swallowing function compared to the regular group, while the number of patients with grade 5 swallowing function was lower than that in the regular group (P < 0.05). Post-intervention, the SSA, SAS, and SDS scores of both groups decreased, with a more significant decrease observed in the coalition group (P < 0.05). Additionally, the total SWAL-QOL score in both groups increased, with a more significant increase observed in the coalition group (P < 0.05). During the intervention period, the total incidence of aspiration and aspiration pneumonia in the coalition group was lower than that in the control group (4.44% vs 20.00%; P < 0.05). CONCLUSION: Systemic intervention combined with psychological intervention can improve dysphagia symptoms, alleviate negative emotions, enhance quality of life, and reduce the incidence of aspiration pneumonia in patients with OD.

Association of the hemoglobin, albumin, lymphocyte, and platelet score with the risk of Erectile dysfunction: a cross-sectional study.

Erectile dysfunction (ED) is related to nutritional and inflammatory factors. The hemoglobin, albumin, lymphocyte, and platelet score (HALP), a new index reflecting the nutritional and inflammatory status, has been associated with a higher risk of diabetic retinopathy, particularly at lower values (≤ 42.9). However, studies focusing on the relationship between HALP and ED risk are scarce. Hence, this study aimed to investigate the association between HALP and ED. Data were extracted from the National Health and Nutrition Examination Survey (NHANES) conducted between 2001 and 2004. Based on self-reported data, participants were classified into either the ED group or the non-ED group. Next, the HALP score was categorized into four quartiles (Q1-4). Weighted multivariate regression analysis was performed to assess the relationship between categorical HALP and ED risk. Additionally, restricted cubic spline (RCS) analysis was conducted to examine the association between continuous HALP scores and ED risk. Furthermore, subgroup analyses were conducted to examine the association between categorical HALP and the risk of ED based on age, as well as the status of hypertension, diabetes and cardiovascular disease. Finally, a mediation analysis was carried out to investigate the mediating effect of HALP and related parameters on the association between urinary cobalt levels and ED. Initially, the data of 21,161 participants were collected. After implementing the inclusion and exclusion criteria, 3406 participants were included in the final analyses. Weighted multivariate regression analysis demonstrated that the Q4 HALP group was associated with a lower risk of ED (OR 0.96, 95% confidence intervals 0.92-1.00, P = 0.037). Meanwhile, RCS analysis showed that HALP was nonlinearly associated with the risk of ED. In addition, subgroup analyses demonstrated that participants in the Q3/4 HALP group had a significantly lower ED risk than those in the Q1 group among patients aged ≥ 50 years, as well as those with hypertension and diabetes. Lastly, mediation analysis revealed that HALP and its associated parameters had a marginal average causal mediation effect on the relationship between urinary cobalt levels and ED risk (P > 0.05). In US adults, high HALP scores were correlated with a lower risk of ED. The relationship was more pronounced in participants aged ≥ 50 years with hypertension and diabetes. Furthermore, HALP and its parameters may not mediate the association between urinary cobalt levels and ED risk.

Disruption of cyclin D1 degradation leads to the development of mantle cell lymphoma.

Cyclin D1 has been recognized as an oncogene due to its abnormal upregulation in different types of cancers. Here, we demonstrated that cyclin D1 is SUMOylated, and we identified Itch as a specific E3 ligase recognizing SUMOylated cyclin D1 and mediating SUMO-induced ubiquitination and proteasome degradation of cyclin D1. We generated cyclin D1 mutant mice with mutations in the SUMOylation site, phosphorylation site, or both sites of cyclin D1, and found that double mutant mice developed a Mantle cell lymphoma (MCL)-like phenotype. We showed that arsenic trioxide (ATO) enhances cyclin D1 SUMOylation-mediated degradation through inhibition of cyclin D1 deSUMOylation enzymes, leading to MCL cell apoptosis. Treatment of severe combined immunodeficiency (SCID) mice grafted with MCL cells with ATO resulted in a significant reduction in tumor growth. In this study, we provide novel insights into the mechanisms of MCL tumor development and cyclin D1 regulation and discover a new strategy for MCL treatment.

Assessment of Arctic sea ice simulations in cGENIE model and projections under RCP scenarios.

Simulating and predicting Arctic sea ice accurately remains an academic focus due to the complex and unclear mechanisms of Arctic sea ice variability and model biases. Meanwhile, the relevant forecasting and monitoring authorities are searching for models to meet practical needs. Given the previous ideal performance of cGENIE model in other fields and notable features, we evaluated the model's skill in simulating Arctic sea ice using multiple methods and it demonstrates great potential and combined advantages. On this basis, we examined the direct drivers of sea-ice variability and predicted the future spatio-temporal changes of Arctic sea ice using the model under different Representative Concentration Pathways (RCP) scenarios. Further studies also found that Arctic sea ice concentration shows large regional differences under RCP 8.5, while the magnitude of the reduction in Arctic sea ice thickness is generally greater compared to concentration, showing a more uniform consistency of change.

[Advances in research on organ-at-risk protection in pelvic tumor external beam radiotherapy].

The simultaneous objectives of destroying tumor cells while protecting normal pelvic organs present a dual clinical and technical challenge within the realm of pelvic tumor radiotherapy. This article reviews the latest literatures, focusing on technological innovations in key aspects of radiotherapy such as positioning, planning, and delivery. These include positioning fixation techniques, organ-at-risk avoidance irradiation, non-coplanar irradiation techniques, as well as organ displacement protection and image-guided adaptive techniques. It summarizes and discusses the research progress made in the protection of critical organs during pelvic tumor radiotherapy. The paper emphasizes technological advancements in the protection of critical organs throughout the processes of radiotherapy positioning, planning, and implementation, aiming to provide references for further research on the protection of critical organs in the external irradiation treatment of pelvic tumors.

Simultaneous effervescence-assisted microextraction and magnetic adsorbent generation for rapid and cost-effective organochlorine pesticides analysis.

This study introduced an innovative magnetic effervescence-assisted microextraction method, streamlining the preparation of effervescent tablets through a one-pot method that blends a CO2 donor (Na2CO3) and an H+ donor (NaH2PO4) with bare magnetic particles (Fe3O4) and an adsorbent (hydroxylated multi-walled carbon nanotubes), followed by pressing. During the extraction process, the bare magnetic particles and adsorbent undergo in-situ self-assembly to create a magnetic adsorbent. The effervescence generates bubbles that enhance effective extraction and magnetism facilitates the easy separation of the magnetic adsorbent from the sample solution, completing the process within 4 min. Applied to organochlorine pesticide analysis in fruit juices and herbal extracts, the method exhibits excellent linearity (R2 > 0.993), sensitivity (detection limits: 0.010-0.125 ng/mL), accuracy (recoveries: 85.8-99.9%), and precision (RSDs < 9.7%) with GC-ECD. Overall, this approach stands out for its simplicity, cost-effectiveness, and suitability for on-site analysis, owing to its operational ease and independence from specialized equipment.

Association between weight-adjusted waist index and kidney stones: a propensity score matching study.

Objective: This study aimed to investigate the association between weight-adjusted waist index (WWI), a novel adiposity index, and kidney stone (KS). Methods: Data were obtained from the National Health and Nutrition Examination Survey 2007-2018. According to the history of KS, participants were divided into the non-stone group and the stone group. Weighted multivariable logistic regression analyses were used to evaluate the correlation between WWI and KS in unadjusted, partially adjusted, and all-adjusted models. A restricted cubic spline (RCS) analysis assessed the association between continuous WWI and KS risk and obtained the risk function inflection point. Then, subgroup analysis based on the risk function inflection point was conducted to dissect the association in specific subgroups. In addition, the above analyst methods were repeatedly performed in populations after propensity score matching (PSM). The receiver operating characteristic (ROC) curve was applied to compare the ability to predict KS occurrence among WWI, visceral adiposity index (VAI), and body mass index (BMI). Results: Weighted multivariable logistic regression analyses found a positive association between continuous WWI and KS risk in the all-adjusted model [odds ratio (OR) = 1.03; 95% confidence interval (CI), 1.02-1.04; P < 0.001]. In further analysis, the Q4 WWI group was linked to the highest KS risk when compared to the Q1-Q3 group (OR = 1.06; 95% CI, 1.05-1.08, P < 0.001). RCS analysis found a linear significant correlation between continuous WWI and KS risk, and the risk function inflection point is 11.08 cm/√kg. Subgroup analysis confirmed that WWI was associated with KS risk in different groups. After PSM, increased WWI was still related to a high risk of KS. Moreover, the ROC curve demonstrated that WWI has a higher predictive ability of KS occurrence than VAI and BMI (area under curve, 0.612 vs. 0.581 vs. 0.569). Conclusion: In the US adult population, elevated WWI value was associated with an increased risk of KS. Furthermore, WWI was a better predictor of KS occurrence than VAI and BMI.

Application of novel CRISPR tools in brain therapy.

In recent years, the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-based genome editing toolkit has been widely used to modify the genome sequence of organisms. As the CRISPR toolbox continues to grow and new CRISPR-associated (Cas) proteins are discovered, its applications have expanded beyond conventional genome editing. This now encompass epigenetic editing, gene expression control, and various other functions. Notably, these advancements are finding practical application in the treatment of brain diseases. Furthermore, the amalgamation of CRISPR and Chimeric Antigen Receptor T-cell (CAR-T) technologies has emerged as a potential approach for disease treatment. With this in mind, this review commences by offering a comprehensive overview of recent advancements in CRISPR gene editing tools. This encompasses an exploration of various Cas proteins, gene expression control, epigenetic editing, base editing and primer editing. Additionally, we present an in-depth examination of the manifold applications of these innovative CRISPR tools in the realms of brain therapeutics, such as neurodegenerative diseases, neurological syndromes and genetic disorders, epileptic disorders, and brain tumors, also explore the pathogenesis of these diseases. This includes their utilization in modeling, gene screening, therapeutic gene editing, as well as their emerging synergy with CAR-T technology. Finally, we discuss the remaining technical challenges that need to be addressed for effective utilization of CRISPR tools in disease treatment.

The phosphorylation-deubiquitination positive feedback loop of the CHK2-USP7 axis stabilizes p53 under oxidative stress.

p53 regulates multiple signaling pathways and maintains cell homeostasis under conditions of DNA damage and oxidative stress. Although USP7 has been shown to promote p53 stability via deubiquitination, the USP7-p53 activation mechanism has remained unclear. Here, we propose that DNA damage induces reactive oxygen species (ROS) production and activates ATM-CHK2, and CHK2 then phosphorylates USP7 at S168 and T231. USP7 phosphorylation is essential for its deubiquitination activity toward p53. USP7 also deubiquitinates CHK2 at K119 and K131, increasing CHK2 stability and creating a positive feedback loop between CHK2 and USP7. Compared to peri-tumor tissues, thyroid cancer and colon cancer tissues show higher CHK2 and phosphorylated USP7 (S168, T231) levels, and these levels are positively correlated. Collectively, our results uncover a phosphorylation-deubiquitination positive feedback loop involving the CHK2-USP7 axis that supports the stabilization of p53 and the maintenance of cell homeostasis.

Mesenchymal stem cell-derived apoptotic vesicles ameliorate impaired ovarian folliculogenesis in polycystic ovary syndrome and ovarian aging by targeting WNT signaling.

Rationale: It has been emergingly recognized that apoptosis generates plenty of heterogeneous apoptotic vesicles (apoVs), which play a pivotal role in the maintenance of organ and tissue homeostasis. However, it is unknown whether apoVs influence postnatal ovarian folliculogenesis. Methods: Apoptotic pathway deficient mice including Fas mutant (Fasmut ) and Fas ligand mutant (FasLmut ) mice were used with apoV replenishment to evaluate the biological function of apoVs during ovarian folliculogenesis. Ovarian function was characterized by morphological analysis, biochemical examination and cellular assays. Mechanistical studies were assessed by combinations of transcriptomic and proteomic analysis as well as molecular assays. CYP17A1-Cre; Axin1fl /fl mice was established to verify the role of WNT signaling during ovarian folliculogenesis. Polycystic ovarian syndrome (PCOS) mice and 15-month-old mice were used with apoV replenishment to further validate the therapeutic effects of apoVs based on WNT signaling regulation. Results: We show that systemic administration of mesenchymal stem cell (MSC)-derived apoptotic vesicles (MSC-apoVs) can ameliorate impaired ovarian folliculogenesis, PCOS phenotype, and reduced birth rate in Fasmut and FasLmut mice. Mechanistically, transcriptome analysis results revealed that MSC-apoVs downregulated a number of aberrant gene expression in Fasmut mice, which were enriched by kyoto encyclopedia of genes and genomes (KEGG) pathway analysis in WNT signaling and sex hormone biosynthesis. Furthermore, we found that apoptotic deficiency resulted in aberrant WNT/ß-catenin activation in theca and mural granulosa cells, leading to responsive action of dickkopf1 (DKK1) in the cumulus cell and oocyte zone, which downregulated WNT/ß-catenin expression in oocytes and, therefore, impaired ovarian folliculogenesis via NPPC/cGMP/PDE3A/cAMP cascade. When WNT/ß-catenin was specially activated in theca cells of CYP17A1-Cre; Axin1fl /fl mice, the same ovarian impairment phenotypes observed in apoptosis-deficient mice were established, confirming that aberrant activation of WNT/ß-catenin in theca cells caused the impairment of ovarian folliculogenesis. We firstly revealed that apoVs delivered WNT membrane receptor inhibitor protein RNF43 to ovarian theca cells to balance follicle homeostasis through vesicle-cell membrane integration. Systemically infused RNF43-apoVs down-regulated aberrantly activated WNT/ß-catenin signaling in theca cells, contributing to ovarian functional maintenance. Since aging mice have down-regulated expression of WNT/ß-catenin in oocytes, we used MSC-apoVs to treat 15-month-old mice and found that MSC-apoVs effectively ameliorated the ovarian function and fertility capacity of these aging mice through rescuing WNT/ß-catenin expression in oocytes. Conclusion: Our studies reveal a previously unknown association between apoVs and ovarian folliculogenesis and suggest an apoV-based therapeutic approach to improve oocyte function and birth rates in PCOS and aging.

Inhibition of a cyclic nucleotide-gated channel on neuronal cilia activates unfolded protein response in intestinal cells to promote longevity.

Ciliary defects are linked to ciliopathies, but impairments in the sensory cilia of Caenorhabditis elegans neurons extend lifespan, a phenomenon with previously unclear mechanisms. Our study reveals that neuronal cilia defects trigger the unfolded protein response of the endoplasmic reticulum (UPRER) within intestinal cells, a process dependent on the insulin/insulin-like growth factor 1 (IGF-1) signaling transcription factor and the release of neuronal signaling molecules. While inhibiting UPRER doesn't alter the lifespan of wild-type worms, it normalizes the extended lifespan of ciliary mutants. Notably, deactivating the cyclic nucleotide-gated (CNG) channel TAX-4 on the ciliary membrane promotes lifespan extension through a UPRER-dependent mechanism. Conversely, constitutive activation of TAX-4 attenuates intestinal UPRER in ciliary mutants. Administering a CNG channel blocker to worm larvae activates intestinal UPRER and increases adult longevity. These findings suggest that ciliary dysfunction in sensory neurons triggers intestinal UPRER, contributing to lifespan extension and implying that transiently inhibiting ciliary channel activity may effectively prolong lifespan.

Protein phosphatase SCP4 regulates cartilage development and endochondral osteogenesis via FoxO3a dephosphorylation.

The regulatory mechanisms involved in embryonic development are complex and yet remain unclear. SCP4 represents a novel nucleus-resident phosphatase identified in our previous study. The primary aim of this study was to elucidate the function of SCP4 in the progress of cartilage development and endochondral osteogenesis. SCP4-/- and SCP4Col2ER mice were constructed to assess differences in bone formation using whole skeleton staining. ABH/OG staining was used to compare chondrocyte differentiation and cartilage development. Relevant biological functions were analysed using RNA-sequencing and GO enrichment, further validated by immunohistochemical staining, Co-IP and Western Blot. Global SCP4 knockout led to abnormal embryonic development in SCP4-/- mice, along with delayed endochondral osteogenesis. In parallel, chondrocyte-specific removal of SCP4 yielded more severe embryonic deformities in SCP4Col2ER mice, including limb shortening, reduced chondrocyte number in the growth plate, disorganisation and cell enlargement. Moreover, RNA-sequencing analysis showed an association between SCP4 and chondrocyte apoptosis. Notably, Tunnel-positive cells were indeed increased in the growth plates of SCP4Col2ER mice. The deficiency of SCP4 up-regulated the expression levels of pro-apoptotic proteins both in vivo and in vitro. Additionally, phosphorylation of FoxO3a (pFoxO3a), a substrate of SCP4, was heightened in chondrocytes of SCP4Col2ER mice growth plate, and the direct interaction between SCP4 and pFoxO3a was further validated in chondrocytes. Our findings underscore the critical role of SCP4 in regulating cartilage development and endochondral osteogenesis during embryonic development partially via inhibition of chondrocytes apoptosis regulated by FoxO3a dephosphorylation.

First report of Lasiodipilodia theobromae Causing leaf spot of Agave sisalana in GuangXi, China.

Agave sisalana, as an excellent fiber producing plant, is mainly planted in Guangxi Province, China. In November 2023, a foliar disease occured on A. sisalana at Liangjiang Town (108.3593 W, 23.4723 N), Wuming District, Nanning in GuangXi, China. Approximately 50 to 60% of the plants (n=200) had obvious leaf spots on more than 70% of the leaves. On the leaves of sisal, circular or irregularly shaped yellow brown spots can be seen, sunken, with no halo on the edges. As time goes on, the lesion gradually expands to the entire blade of the sword (Figure 1A, 1B). To identify the disease etiology, ten agave leaves were collected from GuangXi. Symptomatic midribs were cut into 3×3 mm pieces, surface sterilized with 75 % ethanol for 20 s, rinsed with sterilized distilled water three times, air dried on sterile filter paper, plated on photo dextrose agar (PDA) medium, and incubated at 28 ℃ in the dark. Five isolates (JM01, JM02, JM03, JM05, JM06) with similar morphology were obtained. Colonies on PDA medium were white to grayish-white with atrial mycelia growing initially upward and then forming clusters (Figure 1E). After five days, mycelia turned grayish black. Immature conidia were initially hyaline, aseptate, and ellipsoid. Mature conidia were dark brown, one septate, longitudinal striate, and 22.1 to 26.3×10.2 to 14.9 µm (Figure 1F). Morphologically , the isolates were identified as Lasiodiplodia theobromae (Alves et al. 2008). For molecular identification, genome DNA of five representative isolate was extracted using the Fungi Genomic DNA Purification kit. The internal transcribed spacer (ITS) region of rDNA and translation elongation factor 1-alpha (TEF-1α) and ß-tublin (TUB) gene were amplified with primer pairs ITS1/ITS4 (White et al. 1990), EF1-728F/EF1-986R (Carbone and Kohn 1999), and Bt2a/Bt2b (Glass and Donaldson 1995), respectively, and sequenced. The ITS (PP209594), TEF-1α (PP234629), and TUB (PP234628) sequences of representative isolate JM01 were deposited in GeneBank. BLAST searches showed >99% nucleotide identity to sequences of L. theobromae (ITS, 99.26% to NR111174; TEF-1α, 99.69% to MM840490; TUB, 98.92% to MN172230). Phylogenetic analysis using maximum likelihood based on the combined ITS, TEF-1α, and TUB sequences of the isolates and reference sequences of Lasiodiplodias spp. from GenBank indicated the isolates obtained in this study formed a clade strongly supported based on bootstrap values to the ex-type isolate CBS164.96 sequences of L.theobromae (Figure 2). To test pathogenicity, JM01 was tested by inoculation leaves of one year old agave plants, the epidermis at the inoculation site, 10, 15 and 20 cm below to the crown, was wiped with a 75% alcohol cotton ball, washed three times with sterile water, and punctured (5 mm diameter) with a sterile inoculation needle. A 5 mm block of each isolate cultured on PDA for 3 days was attached to the inoculation site. Controls were inoculated with sterile PDA. The inoculation area was covered with plastic wrap. All plants were kept in a controlled greenhouse at 27℃, 80% relative humidity, and natural daylight, and watered weekly. Each treatment was repeated three times. Remove the block one day later. Three days after inoculation, all inoculated had typical symptoms,but control were healthy (Figure 1C, 1D). Fungal isolates were only recovered from symptomatic stems and were morphologically identical to L. theobromae, completing Koch's postulates. L. Theobromae has been reported as the cause of leaf rot on A. angustifolia in Mexico (Reyes-García et al. 2023). To our knowledge, this is the first report of L. theobromae causing leaf spot on A. sisalana in GuangXi, China. L. theobromae is primarily a plant pathogen that causes rotting and dieback in fruits and plants in tropical and subtropical regions (Puttanna 1967). This study is useful to focus on management strategies for leaf rot disease by L. theobromae of A. sisalana.