ObjectiveIn the realm of forensic science, dust is a valuable type of trace evidence with immense potential for intricate investigations. With the development of DNA sequencing technologies, there is a heightened interest among researchers in unraveling the complex tapestry of microbial communities found within dust samples. Furthermore, striking disparities in the microbial community composition have been noted among dust samples from diverse geographical regions, heralding new possibilities for geographical inference based on microbial DNA analysis. The pivotal role of microbial community data from dust in geographical inference is significant, underscoring its critical importance within the field of forensic science. This study aims to delve deeply into the nuances of fungal community composition across the urban landscapes of Beijing, Fuzhou, Kunming, and Urumqi in China. It evaluates the accuracy of biogeographic inference facilitated by the internal transcribed spacer 2 (ITS2) fungal sequencing while concurrently laying a robust foundation for the operational integration of environmental DNA into geographical inference mechanisms. MethodsITS2 region of the fungal genomes was amplified using universal primers known as 5.8S-Fun/ITS4-Fun, and the resulting DNA fragments were sequenced on the Illumina MiSeq FGx platform. Non-metric multidimensional scaling analysis (NMDS) was employed to visually represent the differences between samples, while analysis of similarities (ANOSIM) and permutational multivariate analysis of variance (PERMANOVA) were utilized to statistically evaluate the dissimilarities in community composition across samples. Furthermore, using Linear Discriminant Analysis Effect Size (LEfSe) analysis to identify and filter out species that exhibit significant differences between various cities. In addition, we leveraged SourceTracker to predict the geographic origins of the dust samples. ResultsAmong the four cities of Beijing, Fuzhou, Kunming and Urumqi, Beijing has the highest species richness. The results of species annotation showed that there were significant differences in the species composition and relative abundance of fungal communities in the four cities. NMDS analysis revealed distinct clustering patterns of samples based on their biogeographic origins in multidimensional space. Samples from the same city exhibited clear clustering, while samples from different cities showed separation along the first axis. The results from ANOSIM and PERMANOVA confirmed the significant differences in fungal community composition between the four cities, with the most pronounced distinctions observed between Fuzhou and Urumqi. Notably, the biogeographic origins of all known dust samples were successfully predicted. ConclusionSignificant differences are observed in the fungal species composition and relative abundance among the cities of Beijing, Fuzhou, Kunming, and Urumqi. Employing fungal ITS2 sequencing on dust samples from these urban areas enables accurate inference of biogeographical locations. The high feasibility of utilizing fungal community data in dust for biogeographical inferences holds particular promise in the field of forensic science.

Objective: To analyze CD36 gene by PacBio Sequel Ⅱ the third-generation sequencing technology (TGS), including non-coding sequence, and to investigate the molecular mechanism of CD36 deficiency. Methods: Flow cytometry was performed in the southern Chinese population to detect the CD36 phenotype. Among them, 15 cases of CD36 type I deficiency, 15 cases of CD36 type Ⅱ deficiency, and 10 positive samples were selected. The TGS of the CD36 gene was performed and statistical analysis was conducted. Results: 40 samples (including 15 cases of type I deficiency, 15 cases of type Ⅱ deficiency, and 10 positive samples) were subjected by TGS of CD36 full-length sequences (except part of intron1). A total of 180 polymorphic loci were identified. Among them, 13 kinds were in the coding region, the rest were in non-coding region, with most mutations located in regulatory regions such as the 5′-UTR and 3′-UTR. Conclusion: The high polymorphism of CD36 non-coding regions, particularly in regulatory sequences, provides mechanistic insights into type Ⅱ CD36 deficiency.

ObjectiveTo optimize the processing technology of honey bran-fried Rosae Laevigatae Fructus（h-RLF）， formulate relevant quality standards， and explore its improving effect and mechanism on mice with ulcerative colitis（UC） induced by dextran sodium sulfate（DSS）. MethodsTaking the content of polysaccharides and water-soluble extract as the indexes， L₉（3⁴） orthogonal test was used to optimize parameters of the amount of honey bran， frying time and frying temperature. The quality of 15 batches of h-RLF decoction pieces was evaluated according to the optimized process， and the inspection limit standard was preliminarily drawn up. Eighty SPF male Kunming mice were randomly divided into 8 groups， including the blank group， model group， mesalazine group（0.13 g·kg^-1）， RLF group（3.77 g·kg^-1）， bran-fried RLF group（3.77 g·kg^-1）， h-RLF low， medium and high dose groups（1.89， 3.77， 7.54 g·kg^-1）， with 10 mice in each group. The mice in the blank group were free to drink pure water， and the other groups were free to drink 3% DSS solution for 7 days to prepare UC mouse model. Each treatment group was given corresponding drugs by intragastric administration， and the blank and model groups were given equal volume of normal saline. The body weight of mice was recorded daily and the disease activity index（DAI） was calculated. After the administration， the colon tissues of mice were collected to observe the pathological changes by hematoxylin-eosin（HE） staining. The levels of tumor necrosis factor（TNF）-α， interleukin（IL）-1β， IL-6 and IL-10 in the colon of mice were detected by enzyme-linked immunosorbent assay（ELISA）. Western blot was used to detect the expression levels of phosphorylation nuclear transcription factor-κB p65（p-NF-κB p65）， Toll-like receptor 4（TLR4）， p-p38 mitogen-activated protein kinase（p-p38 MAPK）， p-extracellular signal-regulated kinase（p-ERK） and p-c-Jun N-terminal kinase（p-JNK） proteins in colon tissues. ResultsThe optimum processing technology of h-RLF was 20 g honey bran per 100 g RLF， and stir-frying at 200 ℃ for 8 min. The limit standard under the examination of h-RLF was preliminarily formulated as follows：the polysaccharide content should not be less than 25% based on anhydrous glucose（C₆H₁₂O₆）， the content of water-soluble extract should not be less than 38%， the moisture content should not be more than 12.0%， the total ash content should not be more than 5.0%， and the acid-insoluble ash content should not be more than 1.0%. The cluster heat map analysis showed that the quality of RLF from Huanggang， Hubei province was better. Animal experiments showed that compared with the blank group， the DAI score of the model group was significantly increased， the levels of TNF-α， IL-1β and IL-6 in the colon tissue were significantly increased， the IL-10 level was significantly decreased， the colonic mucosa was seriously damaged， accompanied by a large number of inflammatory cell infiltration， tissue congestion and a significant reduction in glands， and the expression levels of p-NF-κB p65， TLR4， p-p38 MAPK， p-ERK and p-JNK proteins were significantly increased（P<0.01）. Compared with the model group， each administration group could alleviate the symptoms of colonic ulcer， the structure of colonic crypt was basically intact， and the glands were arranged in an orderly manner. Among them， the high-dose group of h-RLF had a better effect， which could significantly reduce the DAI score and the levels of TNF-α， IL-1β and IL-6 in colon tissue（P<0.01）， and significantly increase the level of IL-10（P<0.01）， alleviate the colonic mucosal injury， and effectively inhibit the expression levels of p-NF-κB p65， TLR4， p-p38 MAPK， p-ERK and p-JNK proteins（P<0.01）. ConclusionThe key parameters of the processing technology of h-RLF are determined， and the optimized technology is stable and feasible. The established quality standard is simple and reliable， and can be used for the quality control. h-RLF can effectively alleviate DSS-induced UC， and its mechanism may be related to inhibiting the activation of NF-κB/TLR4/MAPK pathway.

Objective: To explore the role of the c.598G>A mutation of the ITGB3 gene in the occurrence of fetal and neonatal alloimmune thrombocytopenia (FNAIT) through its expression in vitro. Methods: The platelet antibodies in the sera of the affected neonate and her mother were detected using commercial enzyme-linked immunosorbent assay (ELISA), solid-phase agglutination, flow cytometry and the gold standard monoclonal antibody-specific immobilization of platelet antigens (MAIPA). The common human platelet antigen (HPA) genotypes of the neonate and her parents were obtained using the HPA-SSP method. The presence of mutations was analyzed by sequencing the exons of the ITGB3 and ITGA2B genes. The target gene of ITGB3 was obtained by PCR amplification using the existing human platelet cDNA. The wild-type ITGB3 eukaryotic expression vector was constructed by TA cloning technology. The 598G>A mutant ITGB3 eukaryotic expression vector was obtained by point mutation, and the plasmid DNA was co-transfected with that of ITGA2B (αⅡb) into HEK293 cells. The transfected cells stably expressing GP Ⅱb/Ⅲa were screened and obtained. The expression of GP Ⅱb/Ⅲa in 598G>A mutant transfected cells and the presence of antibodies against this mutation in the serum of mother were detected by flow cytometry and MAIPA. Results: Antibodies against HLA-class Ⅰ and GP Ⅱb/Ⅲa glycoproteins were detected in the serum of the neonate's mother, and subsequent HLA antibody-specific testing confirmed the presence of antibodies against HLA-B 57∶01 and A 02∶05. ITGB3 sequencing showed that the neonate and her father carried the c.598G>A point mutation, which results in the change of glutamate to lysine at position 200. Antibodies against GP Ⅱb/Ⅲa glycoproteins were not detected using constructed c.598G>A mutant transfected cells reacted with the maternal serum. Conclusion: The in vitro expression and analysis of the ITGB3 c.598G>A mutation did not support a role for this mutation in the pathogenesis of FNAIT. The establishment of this method facilitates the discovery of new platelet low-frequency antigens, and provides a theoretical foundation for the detection of antibodies against platelet antigens associated with patients with adverse pregnancy and childbirth histories.

ObjectiveTo investigate the mechanism of the modified of Bazhentang combined with Guishentang in improving pregnancy outcomes in mouse models of embryo implantation dysfunction by regulating T helper 1/T helper 2 （Th1/Th2） immune balance. MethodsEighty ICR female mice were randomly divided into four groups （n=20 per group） on gestational day 1 （GD1）： control， model， western medicine， and traditional Chinese medicine （TCM） groups. Except for the control group， all mice received mifepristone solution （0.2 mg/mouse） via oral gavage on GD4 to induce embryo implantation dysfunction. The TCM group received a water decoction of the modified of Bazhentang combined with Guishentang （20.8 g·kg^-1）， with the western medicine group administered dydrogesterone （3.9 mg·kg^-1）， and the control/model groups given equal volumes of saline. All treatments were administered once daily from GD1 until one day before sample collection. Outcomes included implantation site counts （macroscopic observation）， pregnancy rates， body weight， endometrial histopathology （hematoxylin-eosin staining）， uterine expression of T-box expressed in T cells （T-bet）， GATA-binding protein 3 （GATA3）， interferon gamma （IFN-γ）， and interleukin-4 （IL-4） at protein （Western blot） and mRNA （real-time polymerase chain reaction， Real-time PCR） levels， serum IFN-γ and IL-4 levels （enzyme-linked immunosorbent assay， ELISA）， and Th1/Th2 immune balance evaluated by calculating T-bet/GATA3 and IFN-γ/IL-4 ratios. ResultsCompared to the control group， the model group showed no significant change in pregnancy rate but exhibited a marked reduction in average implantation sites and body weight （P<0.01）. Histopathological analysis revealed endometrial abnormalities， including decreased glandular density， stromal compaction， and absence of nucleolar vacuoles. At the molecular level， uterine tissue in the model group demonstrated significantly upregulated expression of T-bet and IFN-γ （P<0.05， P<0.01）， alongside markedly downregulated GATA3 and IL-4 expression （P<0.05， P<0.01）. Serum analysis confirmed markedly elevated IFN-γ （P<0.01） and reduced IL-4 levels （P<0.01）， resulting in significantly increased T-bet/GATA3 and IFN-γ/IL-4 ratios （P<0.01）. Compared to the model group， pregnancy rates in all treatment groups showed no significant change. Implantation sites and body weight increased substantially （P<0.01）， with restored endometrial morphology characterized by enhanced glandular density， stromal edema， and reappearance of nucleolar vacuoles. Significant downregulation of T-bet and IFN-γ （P<0.01） and upregulation of GATA3 and IL-4 （P<0.05， P<0.01） in uterine tissue were observed. Serum IFN-γ levels were significantly reduced （P<0.05， P<0.01）， while IL-4 levels were significantly elevated （P<0.05）. The Th1/Th2 ratios were significantly decreased （P<0.01）. ConclusionThe modified of Bazhentang combined with Guishentang significantly enhances the number of embryo implantation sites in mice with embryo implantation dysfunction， potentially through modulating T-bet/GATA3 expression， restoring Th1/Th2 immune balance， and improving endometrial receptivity.

Objective: To explore the role of the c.598G>A mutation of the ITGB3 gene in the occurrence of fetal and neonatal alloimmune thrombocytopenia (FNAIT) through its expression in vitro. Methods: The platelet antibodies in the sera of the affected neonate and her mother were detected using commercial enzyme-linked immunosorbent assay (ELISA), solid-phase agglutination, flow cytometry and the gold standard monoclonal antibody-specific immobilization of platelet antigens (MAIPA). The common human platelet antigen (HPA) genotypes of the neonate and her parents were obtained using the HPA-SSP method. The presence of mutations was analyzed by sequencing the exons of the ITGB3 and ITGA2B genes. The target gene of ITGB3 was obtained by PCR amplification using the existing human platelet cDNA. The wild-type ITGB3 eukaryotic expression vector was constructed by TA cloning technology. The 598G>A mutant ITGB3 eukaryotic expression vector was obtained by point mutation, and the plasmid DNA was co-transfected with that of ITGA2B (αⅡb) into HEK293 cells. The transfected cells stably expressing GP Ⅱb/Ⅲa were screened and obtained. The expression of GP Ⅱb/Ⅲa in 598G>A mutant transfected cells and the presence of antibodies against this mutation in the serum of mother were detected by flow cytometry and MAIPA. Results: Antibodies against HLA-class Ⅰ and GP Ⅱb/Ⅲa glycoproteins were detected in the serum of the neonate's mother, and subsequent HLA antibody-specific testing confirmed the presence of antibodies against HLA-B 57∶01 and A 02∶05. ITGB3 sequencing showed that the neonate and her father carried the c.598G>A point mutation, which results in the change of glutamate to lysine at position 200. Antibodies against GP Ⅱb/Ⅲa glycoproteins were not detected using constructed c.598G>A mutant transfected cells reacted with the maternal serum. Conclusion: The in vitro expression and analysis of the ITGB3 c.598G>A mutation did not support a role for this mutation in the pathogenesis of FNAIT. The establishment of this method facilitates the discovery of new platelet low-frequency antigens, and provides a theoretical foundation for the detection of antibodies against platelet antigens associated with patients with adverse pregnancy and childbirth histories.

Objective: To investigate the health-related quality of life and its influencing factors in elderly patients with metabolic syndrome (MS), so as to provide the evidence for improving health-related quality of life in older adults with chronic diseases. Methods: In 2021, elderly MS patients aged ≥60 years from four districts in Nanjing City were selected as the study subjects using a multi-stage random sampling method. Data on social demographic information, lifestyle, disease history and blood biochemical indicators were collected through questionnaire surveys, physical examination and laboratory tests. Health utility value and EuroQol Visual Analog Scale (EQ-VAS) score were assessed using the EuroQol 5-dimension 3-level questionnaire. Factors affecting health-related quality of life were identified with the Tobit regression model and multiple linear regression model. Results: A total of 3 378 elderly MS patients were included, with a median age of 67.00 (interquartile range, 7.00) years. There were 1 558 males (46.12%) and 1 820 females (53.88%). The median (interquartile range) of health utility value and EQ-VAS score were 1.00 (0.03) and 80.00 (15.00). Tobit regression and multiple linear regression analysis showed that gender (female, β=-0.034), education level (middle school, β=0.024; junior college and above, β=0.046), marital status (married, β=0.014), physical activity (sufficient, β=0.013), vegetable intake (meet standard, β=-0.009) and fruit intake (meet standard, β=0.016) were the influencing factors of health utility value. Residence (urban area, β=1.933) and alcohol consumption (yes, β=1.761) were influencing factors of EQ-VAS score. Age, cardiovascular and cerebrovascular diseases, malignant tumors and chronic respiratory diseases were the influencing factors of health utility value and EQ-VAS score. Conclusion Age, sex, marital status, residence, lifestyle and disease are mainly associatied with the health-related quality of life in elderly MS patients.

OBJECTIVE To provide empirical basis for promoting the dynamic adjustment of the pharmaceutical care catalogue and the formulation of policies such as hierarchical payment of medical insurance. METHODS A multicenter cross- sectional survey method was adopted to conduct a questionnaire survey among 424 inpatients in 22 tertiary medical institutions in 12 prefecture-level cities of Hubei Province to evaluate their demand for pharmaceutical care， willingness to pay and preference for service forms. Combined with univariate and multivariate Logistic regression analysis， the influencing factors and key factors that affect patients’ willingness to pay for pharmaceutical care were identified. RESULTS Only 39.86% of the patients were aware of pharmaceutical care or pharmacists， and 89.62% of the patients hope to receive pharmaceutical care. Among the 16 types of pharmaceutical care， the patients surveyed had a relatively high recognition rate for guidance on drug usage and dosage， notification of medication precautions， and the identification， prevention and handling of adverse drug reactions. 96.70%， 95.30%， and 94.12% respectively expressed strong approval and approval. The demand for services such as insurance-related policy consultation， popular science on the mechanism of drug action， and assessment of the combined use of traditional Chinese and Western medicines was relatively low， with 61.65%， 68.47%， and 68.47% expressing strong approval and approval respectively. The positive influencing factors of willingness to pay were household monthly income ＞ 5 000 yuan （OR＝1.742）， awareness of pharmaceutical care or pharmacists （OR＝3.620）， and the desire to receive pharmaceutical care （OR＝4.686） （P＜0.05）， while self-rating health as “good” （OR＝0.390） was a negative influencing factor （P＜0.05）. Cardiovascular and cerebrovascular diseases （54.48%） and antihypertensive drugs （45.05%） were the service scenarios that the surveyed patients most hope to be covered. 85.14% of the patients preferred “service when xiaochnye@126.com needed”， with a single service duration of less than 10 minutes being appropriate （84.43%）， and the willingness to pay within 20 yuan being the main type （85.38%）. CONCLUSIONS Based on the characteristics of patients’ needs and payment behaviors， it is suggested that our country could consider establishing a hierarchical payment mechanism for pharmaceutical care， and focus on differentiated design in combination with diseases and medication situations. At the same time， the rights， responsibilities and service standards of resident pharmacists in the links such as medication reorganization and medical order review should be further clarified to comprehensively enhance the clinical value and policy operability of pharmaceutical care.

Lymphatic metastasis is the main metastatic route for colorectal cancer, which increases the risk of cancer recurrence and distant metastasis. The properties of the lymph node metastatic colorectal cancer (LNM-CRC) cells are poorly understood, and effective therapies are still lacking. Here, we found that hypoxia-induced fibroblast activation protein alpha (FAPα) expression in LNM-CRC cells. Gain- or loss-function experiments demonstrated that FAPα enhanced tumor cell migration, invasion, epithelial-mesenchymal transition, stemness, and lymphangiogenesis via activation of the STAT3 pathway. In addition, FAPα in tumor cells induced extracellular matrix remodeling and established an immunosuppressive environment via recruiting regulatory T cells, to promote colorectal cancer lymph node metastasis (CRCLNM). Z-GP-DAVLBH, a FAPα-activated prodrug, inhibited CRCLNM by targeting FAPα-positive LNM-CRC cells. Our study highlights the role of FAPα in tumor cells in CRCLNM and provides a potential therapeutic target and promising strategy for CRCLNM.

It has become an industry consensus that self-assembled nanoparticles (SAN) are formed by molecular recognition of chemical components in traditional Chinese medicine during the decoction process. The insoluble components in the decoction are mostly in the form of nanoparticles, which can improve the problem of poor water solubility. However, the transfer rate of these insoluble components in the decoction is still very low, which limits the efficacy of the drug. This study aimed to refine the traditional decoction self-assembly phenomenon. The self-assembled nanoparticles were constructed by micro-precipitation method (MP-SAN), and characterized by particle size, zeta potential, stability index and morphology. The formation of MP-SAN and alterations in related physicochemical properties were evaluated using modern spectroscopic and thermal analysis techniques. The quality value transmitting pattern of lignan components within the MP-SAN was assessed via high performance liquid chromatography (HPLC). The MP-SAN showed sphere-like structure with uniform morphology, particle size of (245.3 ± 3.2) nm, polydispersity index (PDI) of (0.13 ± 0.03), zeta potential of (-48.9 ± 5.9) mV and stability index (SI) of (86.05% ± 2.27%). Comprehensive analyses using ultraviolet visible spectroscopy, Fourier transform infrared spectroscopy, differential scanning calorimetry, and other techniques confirmed molecular recognition between the decoction and ethanol extraction, leading to electron rearrangement under the influence of non-covalent bonding. This resulted in the formation of nanoparticles possessing superior thermal stability. As determined by HPLC, the encapsulation rates of the index components in the MP-SAN were all greater than 75% (dehydrodiconiferyl alcohol: 77.00%; herpetolide A: 78.57%; herpetrione: 94.53%), and the transfer rates were all higher than 65% (dehydrodiconiferyl alcohol: 96.01%; herpetolide A: 67.86%; herpetrione: 65.55%), which were 1.34, 1.38 and 4.81 times compared with those of the traditional decoction. In summary, this study successfully constructed the MP-SAN based on micro-precipitation method to achieve high transfer rate and high encapsulation rate of insoluble components in docoction, which provides a pharmaceutics idea for the efficient utilization of pharmacodynamic substance basis of traditional Chinese medicine.