OBJECTIVES: To fabricate an injectable composite microsphere hydrogel reinforced with silk fibroin/hyaluronic acid microspheres, achieving synergistic enhance-ment of mechanical robustness and biofunctionality. METHODS: Methacrylated hyaluronic acid (HAMA) and thiolated silk fibroin (TSF) were synthesized. Monodisperse microspheres generated via microfluidics were UV-cured (420 nm) through thiol-ene click reaction. These microspheres were embedded in a TSF/HAMA matrix to form photo-cured composites. The grafting rate of TSF and HAMA was characterized by H1-NMR; particle size distribution of microsphere hydrogels in soybean oil was observed by optical microscopy; gel point of composite microsphere hydrogels was determined by advanced extensional rheometer; microscopic morphology of microsphere hydrogels was observed by scanning electron microscopy; elemental distribution of microsphere hydrogels was detected by X-ray energy dispersive spectroscopy; tunability of composite microsphere hydrogels was observed by inverted confocal microscopy; mechanical properties of composite microsphere hydrogels were tested by compression testing; swelling ratio, degradation rate and water retention rate of composite microsphere hydrogels were measured by gravimetric method. Cytotoxicity of the composite microsphere hydrogels was determined by Calcein-AM/propidium iodide dual staining and CCK-8 assay; cell migration capability was observed by scratch assay. RESULTS: The grafting rates of HAMA and TSF was 48.03% and 17.99%, respectively. Microsphere hydrogels with particle sizes of (43.3±1.2), (78.1±3.0), and (130.8±1.9) μm were prepared. The gel time of the composite microsphere hydrogels was 48-115s. The laser confocal imaging confirmed dynamic regulation characteristics of the composite microsphere hydrogels. The compressive strength of the composite microsphere hydrogels reached 22.7 kPa and maintained structural integrity at 40% strain after 20 compression cycles. The composite microsphere hydrogels exhibited differential deswelling behaviors in simulated physiological environments, and reducing microsphere particle size could significantly enhance its stability under moist conditions. The degradation rate of the composite microsphere hydrogels was (49.1±0.9)% after 200 h, and water retention rate was maintained at 40%-60% after 96 h. Biocompatibility assays confirmed >95% cell viability and unimpaired cell migration abilities. CONCLUSIONS The TSF/HAMA composite microsphere hydrogel developed in this study has characteristics of rapid fabrication, adjustable mechanical properties, enhanced environmental stability and excellent biocom-patibility, thus providing a new material solution for tissue repair and regenerative medicine.
Fibroins/chemistry* ; Hydrogels/chemistry* ; Microspheres ; Hyaluronic Acid/chemistry* ; Humans

Interfering hepatitis B virus (HBV) capsid assembly holds promise as a therapeutic approach for chronic hepatitis B (CHB). Novel anti-HBV agents are urgently needed to overcome drug resistance challenges, with targeted protein degradation (TPD) emerging as a hopeful strategy. Herein, we report the first degradation of HBV core protein (HBC), a multifunctional structural protein, using small-molecule degraders developed by hydrophobic tagging (HyT) technology. Structure-activity relationship (SAR) analysis identified compound HyT-S7, featuring an adamantyl group, exhibiting potent inhibitory activity (EC₅₀ = 0.46 μmol/L, HepAD38 cells) and degradation ability (DC₅₀ = 3.02 ± 0.54 μmol/L) in a dose- and time-dependent manner. Mechanistic studies demonstrated that the autophagy-lysosome pathway was a potential driver of HyT-S7-induced HBC degradation. Remarkably, HyT-S7 effectively degraded 11 drug-resistant mutants, including highly resistant strains P25G and T33N, to Phase III drug GLS4. Furthermore, cellular thermal shift assay, surface plasmon resonance assay, and molecular dynamics simulations revealed the precise mode of HyT-S7 binding to HBC with the adamantyl group potentially mimicking protein misfolding to facilitate HBC degradation. This first proof-of-concept study highlights the potential of HyT-mediated TPD in HBC as a promising avenue for discovering novel HBV and other antiviral agents with favorable drug resistance profiles.

Sennoside A (SA), a typical prodrug, exerts its laxative effect only after its transformation into rheinanthrone catalyzed by gut microbial hydrolases and reductases. Hydrolases have been identified, but reductases remain unknown. By linking a photoreactive group to the SA scaffold, we synthesized a photoaffinity probe to covalently label SA reductases and identified SA reductases using activity-based protein profiling (ABPP). From lysates of an active strain, Bifidobacterium pseudocatenulatum (B. pseudocatenulatum), 397 proteins were enriched and subsequently identified using mass spectrometry (MS). Among these proteins, chromate reductase/nicotinamide adenine dinucleotide (NADH) phosphate (NADPH)-dependent flavin mononucleotide (FMN) reductase/oxygen-insensitive NADPH nitroreductase (nfrA) was identified as a potent SA reductase through further bioinformatic analysis and The Universal Protein Resource (UniProt) database screening. We also determined that recombinant nfrA could reduce SA. Our study contributes to further illuminating mechanisms of SA transformation to rheinanthrone and simultaneously offers an effective method to identify gut bacterial reductases.

Natural products (NPs) are an important source of new drugs for the treatment of stroke. Identifying cellular targets for bioactive molecules is a major challenge and critical issue in the development of new drugs for stroke. Small-molecule probes play a unique role in target discovery. However, drawbacks to these probes include non-specificity, unstable activity, and difficulty in synthesis. Small-molecule probes based on NPs at least partially compensate for these shortcomings. NPs feature rich chemical and structural diversity, biocompatibility, and unique biological activities. These features could be exploited to provide new ideas and tools for target discovery. Small-molecule probes based on NPs provide a precise and direct search for interacting protein targets of NPs-active small molecules. This review explores the properties of small-molecule probes based on NPs and their applications in mechanistic studies of stroke and other diseases. We hope that this review will bring new perspectives to the mechanistic study of NPs-active small molecules and accelerate the translation of these ingredients into drug candidates for the treatment of stroke.

Objective To understand the reliability of caloric test results through the analysis of the pod shape dia-gram types of caloric test and to provide a more accurate and reliable basis for vestibular system function evaluation.Meth-ods The results of caloric test in 528 patients from July 2021 to May 2022 in vertigo diagnosis and treatment center were analyzed.According to pod pattern and unilateral weakness(UW),the patients were divided into five groups:cold and warm test symmetrical group,left and right side asymmetry group,single value significant abnormal group,bilateral hypo-plasia group and pod morphology unable to be classified group.Unreliable data can be identified by comparing,explaining,and summarizing the results of UW of different groups,so as to reduce the interference of unreliable data to clinical diagno-sis and treatment.Results Cold and warm test symmetric group(n=439)included patients with symmetry of bilateral vestibular function(n=273)and decreased vestibular function of one side(n=166).Left and right symmetrical groups in-eluded pationts with bilateral symmetric vestibular function(n=11)and unilateral reduction(n=13).Single value signifi-cant abnormal group showed cases with unilateral vestibular function reduction(n=25)and bilateral symmetric vestibular function(n=11).Bilateral hypolasion group showed cases with bilateral symmetric vestibular function(n=25)and unilat-eral reduction(n=9).Pod morphology unclassified group(n=5)suggested varions interference factors.Conduson When the function of the peripheral vestibular is symmetrical and normal,we should be alert to the existence of temperature effect,and should consider the inspection of the perfusion apparatus and the calibration of the irrigation temperature.When the function of the peripheral vestibular shows unilateral weakness,repeating the irrigation is recommended when the single value of the caloric test significantly increases or decreases.If there is no change,repeating the irrigation at the same tem-perature in the other ear or even repeating the whole caloric test are recommended.The UW value could not truly reflect the state of peripheral vestibular function when the caoric test produces bilateral weakness.Therefore,water irrigation could be used to make a correct judgment based on the medical history and other auxiliary examinations.It is of great significance to judge the effectiveness of caloric test comprehensively by combining the morphology of pod diagram with UW value.

Objective:To assess the value of optical genome mapping (OGM) for the detection of chromosomal structural abnormalities including ring chromosomes, balanced translocations, and insertional translocations.Methods:Clinical data of four patients who underwent pre-implantation genetic testing concurrently with OGM and chromosomal microarray analysis at the Center of Reproductive Medicine of the Sixth Affiliated Hospital of Sun Yat-sen University from January to October 2022 due to chromosomal structural abnormalities were selected as the study subjects. Some of the results were verified by multi-color fluorescence in situ hybridization. Results:The OGM has successfully detected a balanced translocation and fine mapped the breakpoints in a patient. Among two patients with insertional translocations, OGM has provided more refined breakpoint locations than karyotyping analysis in a patient who had chromosome 3 inserted into chromosome 6 and determined the direction of the inserted fragment. However, OGM has failed to detect the chromosomal abnormalit in a patient with chromosome 8 inserted into the Y chromosome. It has also failed to detect circular signals in a patient with ring chromosome mosaicism.Conclusion:OGM has successfully detected chromosomal structural variations in the four patients and provided assistance for their diagnosis.

Based on the discussions in the The Inner Canon of Yellow Emperor （《黄帝内经》）， it is proposed that in the course of the disease， "bind" represents the initial stage of pulmonary nodules， while "accumulation" represents the final form. In terms of the pathogenesis， "two colds interacting" represented by "body cold" and "cold fluid retention" are the prerequisites for the formation of pulmonary nodules， while "disorder of qi" represented by "fainting" is the core of the formation. The specific manifestation is the disturbance of pivot of shaoyang （少阳） or shaoyin （少阴）， resulting in a complex of cold and heat， and then phlegm and stasis are suddenly generated and further formed into nodules. Therefore， the treatment principle should be to regulate the cardinal mechanism， dissolve phlegm and blood stasis. Depending on the complex degree of cold and heat， it is suggested to use Chaihu Guizhi Decoction （柴胡桂枝汤）， Chaihu Guizhi Ganjiang Decoction （柴胡桂枝干姜汤）， or Chaihu Xianxiong Decoction （柴胡陷胸汤） for disturbance of shaoyang pivot， while for shaoyin pivot dysfunction， modified Mahuang Fuzi Xixin Decoction （麻黄附子细辛汤） or Shengjiang Powder （升降散） can be used.

Objective:Laboratory evaluation of the relationship between cerebrospinal fluid and plasma indicators and intrathecal immunoglobulin G(IgG) synthesis in patients with neurological diseases, and establishment of a new diagnostic method for intrathecal IgG synthesis.Methods:This study retrospectively analyzed the content of IgG in cerebrospinal fluid samples and blood albumin in blood samples, and other test results of 410 patients with neurological diseases who visited Beijing Tiantan Hospital from 2019 to 2022. According to the results of oligoclonal bands in cerebrospinal fluid, patients were divided into intrathecal IgG synthesis group and non-intrathecal IgG synthesis group. The Mann Whitney U test was used for inter group comparison, and a bilateral test with P<0.05 indicates a statistically significant difference. Include indicators with differences between groups in logistic regression analysis, construct a predictive model, and compare it with the established quantitative formula IgG index. Results:There were significant differences in 10 indicators, including cerebrospinal fluid leukocyte count and 24-hour intrathecal IgG synthesis rate, between the intrathecal IgG synthesis group and the non-intrathecal IgG synthesis group, with P<0.05. The area under the curve (AUC) of intrathecal IgG synthesis was higher than the IgG index (AUC=0.920, 0.809, Z=31.178, P<0.001), the sensitivity was higher than the IgG index (0.825, 0.618), and the specificity was lower than the IgG index (0.876, 0.908). Conclusion:The combination of 10 indicators such as cerebrospinal fluid white blood cell count and 24-hour intrathecal IgG synthesis rate can improve the diagnostic efficacy and sensitivity of intrathecal IgG synthesis.

Objective:To investigate the molecular epidemiological characteristics of coxsackievirus A16 (CVA16) in Fujian province from 2020 to 2023.Methods:The epidemiological characteristics of CVA16 associated hand, food and mouth disease (HFMD) in Fujian province from 2020 to 2023 was analyzed. The complete VP1 gene of CVA16 was amplified by RT-PCR and then sequenced, and genetic evolution was analyzed by MEGA X and other softwares.Results:From 2020 to 2023, there were 13 120 cases of HFMD in Fujian province, and the proportion of HFMD which caused by CVA16 was 16.5% (2 160/13 120). From 2020 to 2023, the proportion of accounted cases was 4.7% (94/2 019), 14.1% (457/3 243), 47.6% (1 521/3 199) and 1.9% (88/4 659) respectively. HFMD caused by CVA16 was mainly concentrated in children aged 1 to 5 years, and most of them were 3 years old. The genetic evolution and genotype analysis of 92 complete VP1 gene sequences obtained from 2020 to 2023 showed that the genetic distance between CVA16 strains in Fujian province and the prototype strain was far away. The CVA16 genotype in Fujian province from 2020 to 2023 has three clusters of B1a, B1b and B1c, among which the composition ratio of B1a and B1b in Fujian province in 2020 was 40% and 60% respectively. In 2021, B1a and B1b accounted for 81.8% and 18.2% respectively. Only B1a in 2022; in 2023, there were B1a, B1b and B1c, which respectively accounted for 44.4%, 7.4% and 48.2%. During the period from January to September, B1a was the main cluster. After October we observed an emergence of B1c cluster, which had never been found in Fujian province and was rare in China, was detected and became the dominant cluster.Conclusions:The evolutionary cluster of CVA16 dominant changed from B1b in 2020 to B1a in 2021-2023. After October 2023, the newly discovered B1c became the dominant cluster in Fujian province.

Objective:To compare the results of invasive prenatal diagnosis and preimplantation genetic testing (PGT) and explore the underlying mechanism.Methods:Clinical data of pregnant women undergoing PGT and invasive prenatal diagnosis at the Sixth Affiliated Hospital of Sun Yat-sen University from January 2019 to December 2022 were collected. The results of PGT and invasive prenatal diagnosis were compared, and the outcomes of pregnancies were followed up. This study has been approved by the Medical Ethics Committee of the the Sixth Affiliated Hospital of Sun Yat-sen University (No. 2022SLYEC-491).Results:A total of 172 couples were included in this study, and 26 non-targeted variants were discovered upon prenatal diagnosis, including 10 cases (38.5%) by chromosomal karyotyping, 15 (57.7%) by chromosomal microarray analysis (CMA), and 1 (3.8%) by whole exome sequencing. The 10 karyotypic anomalies had included 6 chromosomal polymorphisms, 2 chromosomal mosaicisms, 1 paternally derived translocation, and 1 missed maternal chromosomal inversion. CMA has identified 15 copy number variations (CNVs), which included 11 microdeletions and microduplications, 3 loss of heterozygosity, and 1 low-level mosaicism of paternal uniparental disomy. One CNV was classified as pathogenic, and another one was likely pathogenic, whilst the remaining 13 were classified as variants of uncertain significance. Therefore, 8.7% of CNVs was detected by invasive prenatal diagnosis after PGT. 92.3% (24/26) of the non-targeted variants have been due to technological limitations of next-generation sequencing (NGS).Conclusion:Invasive prenatal diagnosis after PGT can detect non-targeted variants, which may further reduce the incidence of birth defects.