Protein lysine β-hydroxybutyrylation (Kbhb) is a newly discovered post-translational modification associated with a wide range of biological processes. Identifying Kbhb sites is critical for a better understanding of its mechanism of action. However, biochemical experimental methods for probing Kbhb sites are costly and have a long cycle. Therefore, a feature embedding learning method based on the Transformer encoder was proposed to predict Kbhb sites. In this method, amino acid residues were mapped into numerical vectors according to their amino acid class and position in a learnable feature embedding method. Then the Transformer encoder was used to extract discriminating features, and the bidirectional long short-term memory network (BiLSTM) was used to capture the correlation between different features. In this paper, a benchmark dataset was constructed, and a Kbhb site predictor, AutoTF-Kbhb, was implemented based on the proposed method. Experimental results showed that the proposed feature embedding learning method could extract effective features. AutoTF-Kbhb achieved an area under curve (AUC) of 0.87 and a Matthews correlation coefficient (MCC) of 0.37 on the independent test set, significantly outperforming other methods in comparison. Therefore, AutoTF-Kbhb can be used as an auxiliary means to identify Kbhb sites.
Protein Processing, Post-Translational ; Lysine/chemistry* ; Proteins/chemistry* ; Machine Learning ; Algorithms

OBJECTIVE: To explore the mechanism of antibiotic delivery system targeting bacterial biofilm with linezolid (LZD) based on ε-poly- L-lysine (ε-PLL) and cyclodextrin (CD) (ε-PLL-CD-LZD), aiming to enhance antibiotic bioavailability, effectively penetrate and disrupt biofilm structures, and thereby improve the treatment of bone and joint infections. METHODS: ε-PLL-CD-LZD was synthesized via chemical methods. The grafting rate of CD was characterized using nuclear magnetic resonance. In vitro biocompatibility was evaluated through live/dead cell staining after co-culturing with mouse embryonic osteoblast precursor cells (MC3T3-E1), human umbilical vein endothelial cells, and mouse embryonic fibroblast cells (3T3-L1). The biofilm-enrichment capacity of ε-PLL-CD-LZD was assessed using Staphylococcus aureus biofilms through enrichment studies. Its biofilm eradication efficacy was investigated via minimum inhibitory concentration (MIC) determination, scanning electron microscopy, and live/dead bacterial staining. A bone and joint infection model in male Sprague-Dawley rats was established to validate the antibacterial effects of ε-PLL-CD-LZD. RESULTS: In ε-PLL-CD-LZD, the average grafting rate of CD reached 9.88%. The cell viability exceeded 90% after co-culturing with three types cells. The strong biofilm enrichment capability was observed with a MIC of 2 mg/L. Scanning electron microscopy observations revealed the effective disruption of biofilm structure, indicating potent biofilm eradication capacity. In vivo rat experiments demonstrated that ε-PLL-CD-LZD significantly reduced bacterial load and infection positivity rate at the lesion site ( P<0.05). CONCLUSION The ε-PLL-CD antibiotic delivery system provides a treatment strategy for bone and joint infections with high clinical translational significance. By effectively enhancing antibiotic bioavailability, penetrating, and disrupting biofilms, it demonstrated significant anti-infection effects in animal models.
Biofilms/drug effects* ; Animals ; Anti-Bacterial Agents/pharmacology* ; Polylysine/chemistry* ; Cyclodextrins/administration & dosage* ; Humans ; Linezolid/pharmacology* ; Staphylococcus aureus/physiology* ; Rats, Sprague-Dawley ; Mice ; Rats ; Male ; Drug Delivery Systems ; Staphylococcal Infections/drug therapy* ; Microbial Sensitivity Tests ; Human Umbilical Vein Endothelial Cells ; Osteoblasts/cytology*

In contrast to the conventional spermiogram, metabolomics approaches give insights into the molecular composition of semen and may provide more detailed information on the fertility status of the respective donor. Given the intra-individual variability of spermiogram parameters between two donations, this study sought to elucidate the biological variability of the seminal plasma metabolome over an average period of 8 weeks. Two time-shifted semen samples from 15 healthy donors were compared by a targeted metabolomics approach utilizing the Biocrates AbsoluteIDQ p180 kit. Next to intraclass correlation coefficients (ICC), which represent a measure of reliability, coefficients of variation within individuals (CVW) and coefficients of variation between individuals (CVB) were calculated for each metabolite to demonstrate its stability. Furthermore, men were divided into two cohorts, a similar sperm concentration (SSC) and a differing sperm concentration (DSC) cohort, based on the observed variance in sperm concentration between the two semen donations. The ICC was higher in the SSC compared to the DSC cohort. The levels of 18 metabolites, primarily acylcarnitines, varied between the initial and subsequent donations. After subdivision into subgroups, only ornithine and phosphatidylcholine 40:5 exhibited differential levels between the two donations in the SSC group, compared to 14 metabolites in the DSC group. CVB was higher than CVW but both differed between the metabolite subclasses. Biogenic amines were identified as the least reliable analytes over time, exhibiting the highest CVW, compared to sphingomyelins, which demonstrated the highest reliability with the lowest variation. CVB was the highest for ether-bound glycerophosphatidylcholines and the lowest for amino acids.
Humans ; Male ; Semen/metabolism* ; Metabolome ; Adult ; Sperm Count ; Carnitine/metabolism* ; Metabolomics ; Ornithine/metabolism* ; Semen Analysis ; Phosphatidylcholines/metabolism*

OBJECTIVES: To summarize the clinical and genetic characteristics of epilepsy with intellectual disability caused by SETD1B gene variants in children. METHODS: A retrospective analysis was conducted on the clinical data of three children with SETD1B gene variants diagnosed and treated at the Department of Pediatric Neurology of Xiangya Hospital of Central South University. Relevant literature was reviewed to summarize the clinical characteristics of this condition. RESULTS: All three children presented with symptoms during infancy or early childhood, including mild intellectual disability and myoclonic seizures, with two cases exhibiting eyelid myoclonia. After treatment with three or more antiepileptic drugs, two cases achieved seizure control or partial control, while one case remained refractory. Each of the three children was found to have a heterozygous variant in the SETD1B gene (one deletion, one frameshift, and one missense variant). To date, 54 cases with SETD1B gene variants have been reported, involving a total of 56 variants, predominantly missense variants (64%, 36/56). The main clinical manifestations included varying degrees of developmental delay (96%, 52/54) and seizures (81%, 44/54). Among the 44 patients with seizures, myoclonic (20%, 9/44) and absence seizures (34%, 15/44) were common, with eyelid myoclonia reported in six cases. Approximately one-fifth of these patients had poorly controlled seizures. CONCLUSIONS The primary phenotypes associated with SETD1B gene variants are intellectual disability and seizures, and seizures exhibit distinct characteristics. Eyelid myoclonia is not uncommon.
Humans ; Intellectual Disability/complications* ; Epilepsy/complications* ; Male ; Female ; Histone-Lysine N-Methyltransferase/genetics* ; Child, Preschool ; Child ; Retrospective Studies

OBJECTIVE: To characterize the occurrence of SRSF2 mutations in chronic myelomonocytic leukemia(CMML) patients and their correlation with other gene mutations and some clinical characteristics. METHODS: The clinical data of 43 CMML patients diagnosed in Changzhou No.2 People's Hospital and Wuxi No.2 People's Hospital were retrospectively analyzed, and gene mutations detection was performed using next-generation sequencing (NGS). RESULTS: Among the 43 CMML patients the SRSF2 mutation detection rate was 39.5%(17/43). These mutations clustered collectively at the proline 95 residue in the splicing factor SRSF2. The other genes with mutation rate greater than 15% were ASXL1 (48.8%), TET2 (41.9%), NRAS (30.2%), RUNX1 (25.6%), and SETBP1 (16.3%). Among SRSF2- mutated patients, the most common co-mutation was ASXL1, followed by TET2. The median age of SRSF2 mutant patients was significantly higher than that of the wild type (68 vs 51.5, P < 0.001), but there was not statistically significant differences in gender, peripheral leukocytes, hemoglobin, platelets, karyotype, and blast cell compared to the wild-type (all P >0.05). Notably, 4 out of the 6 SRSF2 ^mutASXL1^mut CMML patients developed leukemia transformation, and 1 out of 10 SRSF2 ^wtASXL1^wt CMML patients developed leukemia transformation, with statistically significant difference in leukemia transformation rates (66.7% vs 10%, P =0.036). CONCLUSION SRSF2 mutations have a high incidence in CMML, occurring frequently in older patients, and often coexisting with ASXL1 and TET2 mutations. Patients with CMML carrying both SRSF2^mut ASXL1^mut double mutations have a higher risk of acute leukemia transformation.
Humans ; Serine-Arginine Splicing Factors/genetics* ; Mutation ; Leukemia, Myelomonocytic, Chronic/genetics* ; Retrospective Studies ; Male ; Female ; Repressor Proteins/genetics* ; DNA-Binding Proteins/genetics* ; Dioxygenases ; Middle Aged ; Aged ; Proto-Oncogene Proteins/genetics*

OBJECTIVE: To explore the role of nuclear receptor-binding SET-domain protein 1 (NSD1) in the pathogenesis of nonobstructive azoospermia (NOA) by regulating the expressions of relevant genes. METHODS: We detected the expression of NSD1 in the testis tissue of 7 male patients with obstructive azoospermia (OA) and 18 with NOA by qPCR and immunofluorescence assay, and determined the modification level of H3K36me2 in the testes of two groups of patients by immunofluorescence staining, Western blot and immunoprecipitation (IP). We examined the difference in the enrichment of H3K36me2 in the testis tissue by chromatin IP-based sequencing (ChIP-Seq), analyzed the genomic distribution and target genes using bioinformatics, and verified the expression levels of the target genes in the testes of the two groups of patients by qPCR. RESULTS: Compared with the patients with OA, those with NOA showed dramatically decreased mRNA and protein expressions of NSD1 (P=0.000 8). The binding of NSD1 to H3K36me2 was observed in the testis tissue of both the two groups of patients, while the modification level of H3K36me2 was evidently reduced in the NOA males. H3K36me2 was distributed mainly in the intergenic region in the testes of the two groups of patients, but the enrichment of H3K36me2 was obviously decreased in the NOA group. The differentially H3K36me2-enriched genes were involved in various biological processes, including tissue development, and cell morphogenesis. Results of ChIP-Seq and qPCR showed significantly down-regulated expressions of the target genes KIT, SPO11 and ACRV1 in the testis tissue of the NOA males compared with those in the OA patients (P<0.01). CONCLUSION The levels of NSD1 and H3K36me2 are decreased in testis tissue of the NOA patient, H3K36me2 is highly enriched in the spermatogenesis-related key genes KIT, SPO11 and ACRV1, and the down-regulated expression of NSD1 impairs spermatogenesis.
Humans ; Male ; Azoospermia/genetics* ; Testis/metabolism* ; Histone-Lysine N-Methyltransferase/metabolism* ; Histones/metabolism*

OBJECTIVE: To explore the specific mechanism of histone deacetylase 2 (HDAC2) mediated histone H3 lysine 27 acetylation (H3K27ac) modification in promoting the proliferation and migration of hepatocellular carcinoma cells. METHODS: Samples of 40 cases of hepatocellular carcinoma and paracancerous tissues resected from January 2021 to January 2023 were collected. The expressions of HDAC2 and H3K27ac in hepatocellular carcinoma, paracancerous tissues and cell lines were detected by immunohistochemistry and Western blotting. The correlation between the expression levels of HDAC2 and H3K27ac and the relationship between HDAC2 expression and clinicopathological characteristics of patients with hepatocellular carcinoma were analyzed. The proliferation, migration and invasion of Hep3B and HepG2 cells were determined by MTS, clone formation, scratch and Transwell experiments. The acetylation of H3K27 mediated by HDAC2 was verified by Western blotting, real-time fluorescence quantitative PCR (qRT-PCR) and chromatin immunoprecipitation high-throughput sequencing (ChIP-seq). In vivo xenotransplantation experiment, the tumorigenicity of cells in each group was measured, and the expression of proteins related to phosphoinositide 3-kinases/phosphatase and tensin homolog deleted on chromosome ten/protein kinase B/mammalian target of rapamycin (PI3K/PTEN/AKT/mTOR) signal pathway was detected. RESULTS: High expression of HDAC2 and low expression of H3K27ac were found in hepatocellular carcinoma tissues and cell lines (P < 0.05), and there was a negative correlation between them (r=-0.477, P=0.002). The expression of HDAC2 was related to tumor size, hepatitis B virus infection, TNM stage and portal vein tumor thrombus (P < 0.05). Compared with the sh-NC group of Hep3B and HepG2 cells, the proliferation, clone formation, migration and invasion ability of sh-HDAC2 group were decreased (P < 0.05). Compared with the Empty group, the HDAC2 group exhibited increased expression levels and activity of HDAC2, as well as enhanced cell proliferation, clone formation, migration, invasion ability, tumor volume and mass in vivo, and elevated expression levels of p-PI3K, p-AKT, and p-mTOR (P < 0.05). Conversely, the enrichment and expression levels of H3K27ac, along with the expression level of PTEN, were decreased (P < 0.05). In the iHDAC2 group, the expression levels and activity of HDAC2, as well as the proliferation, clone formation, migration, invasion ability, tumor volume and mass in vivo, and expression levels of p-PI3K, p-AKT, and p-mTOR were reduced (P < 0.05). Additionally, the expression levels of H3K27ac and PTEN were increased (P < 0.05). To validate the involvement of the PI3K/PTEN/AKT/mTOR signaling pathway in HDAC2-mediated regulation of malignant behaviors in liver cancer cells through H3K27ac, the PI3K activator 740Y-P was introduced. Compared with the iHDAC2 group, the iHDAC2+740Y-P group exhibited increased proliferation, clone formation, migration, invasion ability, tumor volume and mass in vivo, and elevated expression levels of p-PI3K, p-AKT, and p-mTOR (P < 0.05). Conversely, the expression level of PTEN was decreased (P < 0.05). CONCLUSION HDAC2 initiates PI3K/PTEN/AKT/mTOR signal pathway by mediating H3K27 acetylation, which promotes the occurrence and development of hepatocellular carcinoma.
Humans ; Carcinoma, Hepatocellular/metabolism* ; Liver Neoplasms/metabolism* ; Histone Deacetylase 2/physiology* ; Cell Proliferation ; Acetylation ; Cell Movement ; Histones/metabolism* ; Animals ; Hep G2 Cells ; Male ; Female ; Mice ; Cell Line, Tumor ; Signal Transduction ; Mice, Nude ; PTEN Phosphohydrolase/metabolism* ; Lysine/metabolism* ; Middle Aged

The nod-like receptor family pyrin domain containing 3 (NLRP3) inflammasome plays a crucial role in the prognosis of subarachnoid hemorrhage (SAH). WNK1 kinase negatively regulates NLRP3 in various inflammatory conditions, but its role in early brain injury (EBI) after SAH remains unclear. In this study, we used an in vivo SAH model in rats/mice and AAV-WNK1 intraventricular injection to investigate its neuroprotective mechanisms. WNK1 expression was significantly reduced in SAH patient blood and SAH model brain tissue, correlating negatively with microglial activation. AAV-WNK1 alleviated brain edema, neuronal necrosis, behavioral deficits, and inflammation by inhibiting NLRP3 inflammasome activation. In hemin-stimulated BV-2 cells, WNK1 overexpression reduced NLRP3 activation and inflammatory cytokines. Chloride counteracted WNK1's inhibitory effects, and WNK1 suppressed P2X7R-induced NLRP3 activation. Mechanistically, WNK1 functioned via the OXSR1/STK39 pathway. These findings highlight WNK1 as a key regulator of intracellular chloride balance and neuroinflammation, presenting a potential therapeutic target for SAH treatment.
Animals ; NLR Family, Pyrin Domain-Containing 3 Protein/metabolism* ; Subarachnoid Hemorrhage/complications* ; Inflammasomes/metabolism* ; Rats ; Mice ; Neuroinflammatory Diseases/metabolism* ; WNK Lysine-Deficient Protein Kinase 1/genetics* ; Male ; Humans ; Chlorides/metabolism* ; Mice, Inbred C57BL ; Rats, Sprague-Dawley ; Brain Injuries/metabolism* ; Microglia/metabolism* ; Protein Serine-Threonine Kinases

Stem cells play a crucial role in maintaining tissue regenerative capacity and homeostasis. However, mechanisms associated with stem cell senescence require further investigation. In this study, we conducted a proteomic analysis of human dental pulp stem cells (HDPSCs) obtained from individuals of various ages. Our findings showed that the expression of NUP62 was decreased in aged HDPSCs. We discovered that NUP62 alleviated senescence-associated phenotypes and enhanced differentiation potential both in vitro and in vivo. Conversely, the knocking down of NUP62 expression aggravated the senescence-associated phenotypes and impaired the proliferation and migration capacity of HDPSCs. Through RNA-sequence and decoding the epigenomic landscapes remodeled induced by NUP62 overexpression, we found that NUP62 helps alleviate senescence in HDPSCs by enhancing the nuclear transport of the transcription factor E2F1. This, in turn, stimulates the transcription of the epigenetic enzyme NSD2. Finally, the overexpression of NUP62 influences the H3K36me2 and H3K36me3 modifications of anti-aging genes (HMGA1, HMGA2, and SIRT6). Our results demonstrated that NUP62 regulates the fate of HDPSCs via NSD2-dependent epigenetic reprogramming.
Humans ; Dental Pulp/cytology* ; Nuclear Pore Complex Proteins/genetics* ; Cellular Senescence/genetics* ; Stem Cells/metabolism* ; Epigenesis, Genetic ; Cell Proliferation ; Cell Differentiation ; Histone-Lysine N-Methyltransferase/metabolism* ; Cells, Cultured ; Cellular Reprogramming ; Cell Movement ; Proteomics

The caries-preventive effects of arginine have been attributed to its impact on biofilm composition and pH. Recent in vitro studies suggest that arginine also affects the production of biofilm matrix components that contribute to virulence, but this mechanism has not been investigated clinically. This randomized, placebo-controlled, triple-blind, split-mouth in situ trial assessed arginine's impact on the microbial composition, matrix architecture, and microscale pH of biofilms from caries-active patients (N = 10). We also examined whether individual differences in the pH response to arginine were related to biofilm composition and matrix structure. Biofilms were grown for four days on carriers attached to intraoral splints. Three times daily, the biofilms were treated extraorally with sucrose (5 min), followed by arginine or placebo (30 min), in a split-mouth design. After growth, the microscale biofilm pH response to sucrose was monitored by pH ratiometry. Microbial biofilm composition and carbohydrate matrix architecture were analyzed by 16S rRNA gene sequencing and fluorescence lectin-binding analysis, respectively. Arginine treatment significantly mitigated sucrose-induced pH drops, reduced total carbohydrate matrix production, and altered the spatial distribution of fucose- and galactose-containing carbohydrates. Both arginine- and placebo-treated biofilms were dominated by streptococci and Veillonella spp. Paired analyses showed a significant reduction in mitis/oralis group streptococci and a non-significant increase in several arginine metabolizers in arginine-treated biofilms. Individual pH responses were not significantly associated with the abundance of specific bacterial taxa or carbohydrate matrix components. In conclusion, arginine reduced the virulence of biofilms from caries-active patients through multiple mechanisms, including suppressing matrix carbohydrate production.
Biofilms/drug effects* ; Humans ; Arginine/pharmacology* ; Hydrogen-Ion Concentration ; Dental Caries/prevention & control* ; Male ; Female ; Adult ; Double-Blind Method ; Sucrose/pharmacology*