ObjectiveTo explore the possible mechanisms of Wenyang Huazhuo Formula （温阳化浊方， WHF） in improving reproductive aging from the perspective of the ovarian mechanical microenvironment. MethodsThe experiment included five groups， 3-month group （20 female mice at 3 months of age）， 6-month group （20 female mice at 6 months of age）， 6-month + WHF group （20 female mice at 5 months of age treated with WHF）， 9-month group （20 female mice at 9 months of age）， and 9-month + WHF group （20 female mice at 8 months of age treated with WHF）. The 6-month + WHF group and 9-month + WHF group were orally administered WHF 41.2 g/（kg·d） once daily for 4 consecutive weeks. The other three groups received no intervention. Reproductive hormone levels were measured by ELISA. HE staining was used to count the numbers of various stages of follicles. Ovarian hyaluronic acid （HA） content and collagen fiber content were measured to evaluate the ovarian mechanical microenvironment. Superovulation was performed to observe the number of eggs obtained， as well as the number of offspring and birth weight to assess fertility. The in vitro fertilization and blastocyst culture of oocytes from female offspring in each group were observed to evaluate the effect of WHF on offspring reproductive potential. ResultsCompared with the 3-month group， the 6-month group and 9-month group showed significantly decreased serum levels of gonadotropin-releasing hormone （GnRH）， follicle-stimulating hormone （FSH）， and luteinizing hormone （LH）， decreased ovarian collagen content， and reduced numbers of primordial and secondary follicles. In contrast， the numbers of primary follicles， antral follicles， and atretic follicles increased. The levels of anti-Müllerian hormone （AMH）， ovarian HA content， and the fertilization rate， cleavage rate， and blastocyst formation rate of oocytes from offspring were significantly lower （P＜0.05）. Compared with the 6-month group， the 6-month + WHF group showed significantly reduced serum levels of GnRH， FSH， and LH， with a significant decrease in primary follicles， antral follicles， and atretic follicles as well as increase of AMH levels， ovarian HA content， number of primordial and secondary follicle， egg count， and offspring birth weight （P＜0.05）. Compared with the 9-month group， the 9-month + WHF group exhibited reduced GnRH， FSH， and collagen fiber content， as well as reduced number of primary follicles， antral follicles， and atretic follicles. However， AMH levels， ovarian HA content， number of primordial and secondary follicle， egg count， offspring numbers， birth weight， fertilization rate， cleavage rate， and blastocyst formation rate of oocytes from offspring all significantly increased （P＜0.05）. ConclusionWHF can significantly improve the ovarian reserve， fertility， and reproductive potential in offspring during reproductive mid-life and late-life stages. Its effect may be related to the remodeling of the mechanical microenvironment of aging ovaries. Moreover， the effect on the mechanical microenvironment remodeling of late-stage ovaries and the improvement of the offspring reproductive potential is more significant.

Objective: To investigate the role of calcium/calmodulin-dependent protein kinase Ⅱ delta (CaMKⅡδ) in platelet activation mediated by thrombin and thrombin receptor activator peptide (TRAP). Methods: The CaMKⅡδ-specific inhibitor Hesperadin was used to assess its inhibitory effects on platelet activation using multiple detection methods. These included: analyzing the inhibitory effect of different concentrations of Hesperadin on thrombin- and TRAP-induced platelet aggregation using a platelet aggregometer; measuring CD62P expression levels and calcium mobilization via flow cytometry; and determining dense granule release activity using a microplate reader. Results: Hesperadin at concentrations of 40 μM and 80 μM significantly inhibited thrombin- and TRAP-induced platelet aggregation (P<0.05). Within the concentration range of 20–80 μM, it significantly inhibited α-granule release and TRAP-induced calcium ion mobilization (P<0.05); at 80 μM, it markedly suppressed thrombin-induced platelet calcium ion mobilization and significantly inhibited dense granule release (P<0.05). Conclusion: The experimental results indicate that CaMKⅡδ in platelets is activated and plays a regulatory role in the process of platelet activation induced by thrombin and TRAP.

ObjectiveTo investigate the mechanism of salvianolic acid F （Sal F） in repairing the high glucose-induced injury in human kidney-2 （HK-2） cells via the B-cell lymphoma-2 （Bcl-2）-associated X protein （Bax）/cysteinyl aspartate-specific proteinase 3 （Caspase-3）/gasdermin-E （GSDME） pathway. MethodThe cell counting kit-8 （CCK-8） was used to measure the relative viability of HK-2 cells exposed to high glucose and different concentrations （2.5， 5， 10， 20 μmol·L^-1） of Sal F and the relative viability of HK-2 cells treated with Sal F for different time periods. The levels of lactate dehydrogenase （LDH） and interleukin-1β （IL-1β） in the supernatant of the cell culture were measured by the LDH assay kit and enzyme-linked immunosorbent assay （ELISA） kit， respectively. Flow cytometry combined with Annexin V-FITC/propidium iodide （PI） and Hoechst 33342/PI staining was employed to reveal the proportion of PI-positive HK-2 cells exposed to high glucose. Western blotting was employed to determine the protein levels of Bax， Bcl-2， cytochrome C， cysteinyl aspartate-specific proteinase （Caspase）-9， Caspase-3， and GSDME in the HK-2 cells exposed to high glucose and treated with Sal F. The 2，7-dichlorodihydrofluorescein diacetate fluorescence probe （DCFH-DA） and mitochondrial membrane potential assay kit （JC-1） were used to determine the production of reactive oxygen species （ROS） and the mitochondrial membrane potential in the HK-2 cells exposed to high glucose and treated with Sal F. ResultCompared with the blank group， the model group showed decreased cell viability （P<0.01）， elevated levels LDH and IL-1β， increased proportion of PI-positive cells （P<0.01）， up-regulated protein levels of Bax， cytochrome C， Caspase-9， Caspase-3， and GSDME （P<0.01）， down-regulated protein level of Bcl-2 （P<0.01）， decreased mitochondrial membrane potential， and excessive ROS accumulation. Compared with the model group， Sal F repaired the high glucose-induced injury in HK-2 cells （P<0.05）， lowered the levels of LDH and IL-1β （P<0.05， P<0.01）， and decreased the proportion of PI-positive cells （P<0.01）. In addition， Sal F down-regulated the protein levels of Bax， cytochrome C， Caspase-9， Caspase-3， and GSDME and up-regulated the protein level of Bcl-2 （P<0.05， P<0.01）， increased the mitochondrial membrane potential， and decreased the accumulation of ROS in HK-2 cells. ConclusionSal F can reduce the production of ROS， restore the balance of mitochondrial membrane potential， and inhibit pyroptosis via the Bax/Caspase-3/GSDME signaling pathway to repair the high glucose-induced injury in HK-2 cells.

ABSTRACT OBJECTIVE To explore the correlation between HPLC fingerprint and analgesic efficacy of Zhejiang vinegar Corydalis Rhizoma, and to screen out the material basis of analgesic effect of Zhejiang vinegar Corydalis Rhizoma.METHODS Established HPLC fingerprint of Zhejiang vinegar Corydalis Rhizoma and dysmenorrhea model in rats. The weight loss rate, writhing response, serum malondialdehyde(MDA), estradiol(E2), prostaglandin E2(PGE2) and prostaglandin F2α(PGF2α) levels were used as the evaluation indexes of analgesic efficacy. Combined with chemometrics, the main active components of analgesic effect of Zhejiang vinegar Corydalis Rhizoma and the important components with great contribution to the content were screened.RESULTS The fingerprints of different batches of Zhejiang vinegar Corydalis Rhizoma were established, and nine components were identified; compared with the blank group, the weight loss rate, writhing reaction, MDA, E2 and PGF2α levels in the model group were significantly increased. Compared with the model group, the above indexs in the administration group was decreased. Compared with the blank group, the PGE2 level in the model group was significantly decreased. Compared with the model group, the PGE2 level in the administration group was significantly increased. The vinegar Corydalis Rhizoma produced in Zhejiang had a good analgesic effect, and the main active components of its analgesic effect were protopine, palmatine hydrochloride, and dehydrocorydaline. The important components with greater contribution to the content were tetrahydropalmatine, protopine, palmatine hydrochloride and stylopine.CONCLUSION The efficacy of Zhejiang vinegar Corydalis Rhizoma is the result of the combined action of multiple components, and each component has a strong correlation with the pharmacodynamic indexes. This study provides a reference for the screening of analgesic material basis of Zhejiang vinegar Corydalis Rhizoma.

On May 25, 2024, Shanghai Municipal Administration for Market Regulation released Shanghai local standard Requirements for Outdoor Smoking Areas Setting Up and Management (DB 31/T 1482‒2024) (hereinafter referred to as Standard), which scheduled for official implementation from September 1, 2024. This article provided an interpretation of the key provisions in the Standard, with a particular emphasis on the scope of application, establishment and management requirements. In addition, the significance and potential difficulties and challenges during subsequent implementation of the Standard was summarized and outlined simultaneously, so as to provide a guarantee for users to fully comprehend and effectively implement the Standard.

ObjectiveTo investigate the effect of Fuhe Decoction （敷和汤） with different doses of Suanzaoren （Ziziphus jujuba） for atopic dermatitis （AD）. MethodsForty-eight female BALB/c mice were randomly divided into normal group， model group， loratadine group， and Fuhe Decoction groups with high， medium， and low doses of Fuhe Decoction （Fuhe Decoction high-， medium-， and low-dose groups）， with eight mice in each group. The AD model was prepared by continuous stimulation with 2，4-dinitrofluorobenzene （DNFB） in all groups but the normal group. After modelling， the Fuhe Decoction high-， medium- and low-dose groups were given 24， 18 and 15 g/（kg·d） of Fuhe Decoction， the loratadine group was given 0.001 g/（kg·d） of loratadine dry suspension， and the normal group and the model group were given 10 ml/（kg·d） of normal saline by gavage. All groups were gavaged for 14 days. The number of scratches within 10 min and the score of skin lesions were observed on the 7th and 14th days of modelling and on the 7th and 14th days of drug administration， respectively； serum immunoglobulin E （IgE） was detected by ELISA； the histopathological and morphological changes of the skin were observed by HE staining； and the diversity and abundance of intestinal flora were detected by 16S rRNA sequencing of fecal matter from the colon of the mice. ResultsCompared with the normal group， mice in the model group on the 7th day and the 14th day of modelling and the 7th day， the 14th day of gavage showed increased scratching within 10 min and higher skin lesion scores （P＜0.05）， with hyperkeratotic or incomplete epidermis， marked thickening of spiny cells， and a large number of inflammatory cells infiltrated in the mice after gavage； serum levels of IgE elevated （P＜0.05）， and the abundance of Bacillota decreased， that of the Bacteroidota and bacteria elevated， and relative abundance of Lactobacillus spp. and Prevotella spp. decreased， and relative abundance of Anaplasma spp. and Treponema spp. increased （P＜0.05）. Compared with the model group， the number of scratches within 10 min and the skin lesion scores of mice in the loratadine group and the Fuhe Decoction medium- and high-dose groups decreased on the 7th day and the 14th day of gavage （P＜0.05）， serum IgE reduced， and the bacteria reduced in the loratadine group， the abundance of Bacteroidesmus spp. increased and Bacteriodesmus spp. decreased in the medium-dose group of Fuhe Decoction， the abundance of Bacteriodesmus spp. decreased in the loratadine group， the abundance of Bacteriodesmus spp. decreased， and that of both Lactobacillus spp. and Prevotella spp. increased in Fuhe Decoction medium-dose group （P＜0.05）. Compared with the loratadine group， the skin lesion scores increased in Fuhe Decoction low-dose group， and the number of scratching increased in the Fuhe Decoction low- and high-dose groups on the 7th day and the 14th day of gavage； the IgE content increased in Fuhe Decoction low-dose group， the Bacillota increased and the Bacteroidota decreased， the Lactobacillus spp. and Prevotella spp. increased in Fuhe Decoction middle-dose group， and Anopheles spp. increased in Fuhe Decoction high-dose group after gavage （P＜0.05）. ConclusionFuhe Decoction can improve the clinical symptoms of AD， regulate the relative abundance of intestinal flora to correct the disorders of the bacterial flora， among which the effect of Fuhe Decoction medium-dose group is optimal and comparable to that of the loratadine group， and the reduction of serum IgE inflammatory response may be one of its mechanisms of action.

Objective To investigate the binding and carrying effects of human serum albumin(HSA)from various sources on sphingosine-1-phosphate(S1P).Methods Utilizing human plasma-derived HSA(pHSA)and recombinant HSA(rHSA)samples as the focal points of our investigation,LC-MS/MS technology was employed to meticulously compare and an-alyze the disparities in S1P content among the aforementioned samples.Subsequently,under physiological concentration condi-tions,S1P was directly introduced to HSA samples for loading processing,facilitating a comprehensive comparison of the bind-ing efficacy of HSA from different sources to S1P.Within a serum-free culture setting,HSA samples from various sources were co-cultured with HUVEC cells.The alterations in S1P content within the cell culture supernatant across different treatment groups were meticulously analyzed,allowing for a nuanced comparison of the S1P carry effects exerted by HSA from different sources on cells.The interaction between HSA and S1P molecules from different sources was analyzed and their affinity was cal-culated using surface plasmon resonance(SPR)technology.Furthermore,leveraging AutoDock Vina software and the Mol-prophet platform,the molecular docking analysis of HSA and S1P was conducted,aiming to predict the key binding pocket do-main of S1P within HSA.Results All pHSA samples exhibited detectable levels of S1P(ranging from 3.31±0.03 to 30.35±0.07 μg/L),with significant variations observed among pHSA samples from different manufacturers(P＜0.001).Conversely,S1P was undetectable in all rHSA samples.Upon load treatment,the binding affinity of HSA from diverse sources to S1P dem-onstrated significant discrepancies(P＜0.001),with rHSA exhibiting approximately double the average S1P loading compared to pHSA(ΔCrHSA=801.75±142.45 μg/L vs ΔCpHSA=461.94±85.73 μg/L;P＜0.001,t=5.006).Co-culture treatment out-comes revealed a significant elevation in S1P concentration within the supernatant after 6 hours of co-culture across all HSA sample processing groups with HUVEC cells,while no changes were observed in the supernatant of the blank control group.Notably,significant differences in supernatant S1P concentration were observed among treatment groups at 6 h,12 h,and 24 h(P＜0.001).SPR analysis unveiled a stronger affinity of pHSA for S1P compared to rHSA(KDpHSA-S1P:2.38E-06,KDrHSA-S1P:3.72E-06).Molecular docking analysis and binding pocket prediction suggested that the key binding pocket of HSA and S1P may reside in the IB subdomain of the HSA molecule.Conclusion HSA from various sources exhibits distinct binding and carrying effects on S1P,which appear to be closely associated with the IB subdomain of the HSA molecule.

Objective:To analyze the genus, drug resistance/virulence and phylogenetic characteristics of Brucella strains isolated from brucellosis surveillance sentinels in Henan Province from 2013 to 2022, and provide baseline data for the surveillance, early warning and outbreak tracing of brucellosis. Methods:Blood samples were collected from patients with Brucella infection for strain isolation, culture and species identification, drug susceptibility test, whole genome sequencing, splicing and assembly, functional/virulence/resistance gene prediction analysis and phylogenetic tree drawing based on single nucleotide polymorphism (SNP). Results:In 36 brucellosis patients, the majority were men (86.11%, 31/36), young adults aged 18-50 (88.89%, 32/36) and farmers/herdsmen (72.22%, 26/36). A total of 36 strains of Brucella melitensis were isolated, and average 1 305 functional proteins of 21 categories were predicted by strain genome; all the strains carried four main virulence factors (pmm, VirB group, BtpA/BtpB, BvrS/BvrR). The drug sensitivity rate was 100.00% to six types of antibiotics including levofloxacin, rifampicin, doxycycline, streptomycin, tetracycline and gentamicin, they showed different resistances to three antibiotics including compound trimethoprim-sulfamethoxazole, ciprofloxacin and ampicillin. The strains carried four types of resistance genes and two clusters of resistance genes, with four combinations of genotypes, the resistance mechanisms included antibiotic degradation/modification enzymes, resistant nodular cell differentiation (RND) efflux pumps, 16S/23S ribosomal rRNA binding site mutations, etc. The number of SNP differed in the genomes of 36 Brucellamelitensis strains ranged from 0 to 454 and phylogenetic tree was divided into three major branches, with relative branch distances between 0.000 0 and 0.498 6 for each strain. Conclusions:Human Brucellamelitensis strains isolated from surveillance sentinels in Henan from 2013 to 2022 carried multiple virulence and antibiotic resistance genes and had different drug resistance phenotypes. Single nucleotide polymorphism analysis and phylogenetic tree analysis showed significant differences in phylogenetic relationships among different strains.

Objective To investigate the mechanism of astragaloside Ⅰ,the active constituent of milkvetch root,in inhibiting podocyte injury and improving diabetic kidney disease.Methods According to the body weight,60 male db/db mice were randomly divided into the model group,astragaloside Ⅰ low-dose group(10 mg/kg),astragaloside Ⅰ medium-dose group(20 mg/kg),astragaloside Ⅰ high-dose group(40 mg/kg),and valsartan group(10mg/kg),with 12 mice per group.Twelve db/db littermate control db/m mice were used as the control group.The drug was administered by gavage for 8 weeks.Transmission electron microscope was used to observe the ultrastructure of the kidney;immunohistochemistry and Western blotting were used to detect the expression of nephrotic protein(nephrin),a marker of renal podocytes;enzyme-linked immunosorbent assay was used to detect the contents of interleukin-1β(IL-1β)and interleukin-18(IL-18)in the serum of mice;Western blotting was used to detect the protein expressions of NOD-like receptor thermoprotein domain-related protein 3(NLRP3),cysteinyl aspartate specific proteinase 1(Caspase-1),and Gasdermin D(GSDMD)in kidney tissue.Results Compared with the control group,the glomeruli of the model group showed obvious podocyte loss and foot process fusion;the protein expression of nephrin was decreased(P＜0.05);the contents of IL-1 β and IL-18 in serum were increased(P＜0.05);the protein expressions of NLRP3,Cleaved-Caspase-1,and GSDMD-N were increased(P＜0.05).Compared with the model group,the renal pathological damage in the astragaloside Ⅰ administration groups were alleviated;the protein expression of nephrin was increased(P＜0.05);the contents of IL-1β and IL-18 in serum were decreased(P＜0.05);the protein expressions of NLRP3,Cleaved-Caspase-1,and GSDMD-N were decreased(P＜0.05).Conclusion Astragaloside Ⅰ may play a role in intervening diabetic kidney disease by inhibiting pyroptosis and improving podocyte injury.

Localized juvenile spongiotic gingival hyperplasia(LJSGH)is a kind of gingival hyperplasia with unique pathological manifestations.Its clinical manifestations are atypical,and the etiology and pathogenesis are unclear.No case report was reported in China.The diagnosis of this disease mainly relies on pathological testing,and recurrence may occur after treatment.The best treatment method still lacks medical evidence.This paper reports a case of LJSGH in a teenager and summarizes its clinical,pathological,and treatment through literature review.This work provides a refer-ence for the diagnosis and treatment of this disease.