ObjectiveTo explore the spatial distribution patterns of flavonoid glycosides in Epimedium sagittatum and the influences of environmental factors on the accumulation of these components. MethodsThe spatial statistical analysis and GeoDetector model were used to analyze the distribution patterns of epimedin A，epimedin B，epimedin C，icariin，and total flavonoid glycosides in E. sagittatum samples from 92 different production areas in 36 cities of 13 provinces/municipalities/autonomous regions of China，as well as the effects of 28 environmental factors on the accumulation of each component. ResultsThe average content of flavonoid glycosides 64 （69.56%） producing areas and 30 （83.33%） cities met the quality standard of no less than 1.50% of total flavonoid glycosides in the 2020 edition of Chinese Pharmacopoeia.Epimedin A，epimedin B，epimedin C，icariin，and their sum showed significantly high accumulation.The hot spots regions of epimedin A and epimedin B were similar with each other，mainly located in western Hunan，eastern Hubei，eastern Guizhou，and northern Guangxi.The common hot spot areas of epimedin C and total flavonoid glycosides were in western and southwestern Hunan，southern Henan，northern Anhui，eastern Guizhou，and southern Chongqing.The hot spots areas of icariin were in southern Chongqing，western Hunan，and eastern and northeastern Guizhou.The interactions between environmental factors had stronger explanatory power for the accumulation of components than single factors.The strongest single factor and interactive factor affecting the accumulation of epimedin C were precipitation of wettest quarter （q=0.16） and its interaction with temperature seasonality （q=0.35），respectively.The strongest single factor influencing both the accumulation of icariin and total flavonoid glycosides was the precipitation of coldest quarter （q equals 0.15 and 0.22，respectively）.The strongest interactions were observed between precipitation of coldest quarter and gravel content （q=0.34），as well as between precipitation of coldest quarter and aspect （q=0.35）. ConclusionThirteen cities，including Zhumadian and Nanyang in Henan，Huaihua，Shaoyang，and Zhangjiajie in Hunan，and Zunyi，Qiandongnan，and Tongren in Guizhou，were hot spots of total flavonoid glycosides in E.sagittatum.Precipitation，gravel content，temperature seasonality，and aspect significantly influence the accumulation of flavonoid glycosides in E.sagittatum.This study provides reference for the utilization and production zoning of E.sagittatum.

Objective To explore the construction of α-1,3-galactosyltransferase (GGTA1) gene-knockout (GTKO) Diannan miniature pigs and the kidney xenotransplantation from pigs to rhesus macaques, and to assess the effectiveness of GTKO pigs. Methods The GTKO Diannan miniature pigs were constructed using the CRISPR/Cas9 gene-editing system and somatic cell cloning technology. The phenotype of GTKO pigs was verified through polymerase chain reaction, Sanger sequencing and immunofluorescence staining. Flow cytometry was used to detect antigen-antibody (IgM) binding and complement-dependent cytotoxicity. Kidney xenotransplantation was performed from GTKO pigs to rhesus macaques. The humoral immunity, cellular immunity, coagulation and physiological indicators of the recipient monkeys were monitored. The function and pathological changes of the transplanted kidneys were analyzed using ultrasonography, hematoxylin-eosin staining, immunohistochemical staining and immunofluorescence staining. Results Single-guide RNA (sgRNA) targeting exon 4 of the GGTA1 gene in Diannan miniature pigs was designed. The pGL3-GGTA1-sgRNA1-GFP vector was transfected into fetal fibroblasts of Diannan miniature pigs. After puromycin selection, two cell clones, C59# and C89#, were identified as GGTA1 gene-knockout clones. These clones were expanded to form cell lines, which were used as donor cells for somatic cell nuclear transfer. The reconstructed embryos were transferred into the oviducts of trihybrid surrogate sows, resulting in 13 fetal pigs. Among them, fetuses F04 and F11 exhibited biallelic mutations in the GGTA1 gene, and F04 had a normal karyotype. Using this GTKO fetal pig for recloning and transferring the reconstructed embryos into the oviducts of trihybrid surrogate sows, seven surviving piglets were obtained, all of which did not express α-Gal epitope. The binding of IgM from the serum of rhesus monkey 20# to GTKO pig PBMC was reduced, and the survival rate of GTKO pig PBMC in the complement-dependent cytotoxicity assay was higher than that of wild-type pig. GTKO pig kidneys were harvested and perfused until completely white. After the left kidney of the recipient monkey was removed, the pig kidney was heterotopically transplanted. Following vascular anastomosis and blood flow restoration, the pig kidney rapidly turned pink without hyperacute rejection (HAR). Urine appeared in the ureter 6 minutes later, indicating successful kidney transplantation. The right kidney of the recipient was then removed. Seven days after transplantation, the transplanted kidney had good blood flow, the recipient monkey's serum creatinine level was stable, and serum potassium and cystatin C levels were effectively controlled, although they increased 10 days after transplantation. Seven days after transplantation, the levels of white blood cells, lymphocytes, monocytes and eosinophils in the recipient monkey increased, while platelet count and fibrinogen levels decreased. The activated partial thromboplastin time, thrombin time and prothrombin time remained relatively stable but later showed an upward trend. The recipient monkey survived for 10 days. At autopsy, the transplanted kidney was found to be congested, swollen and necrotic, with a small amount of IgG deposition in the renal tissue, and a large amount of IgM, complement C3c and C4d deposition, as well as CD68⁺ macrophage infiltration. Conclusions The kidneys of GTKO Diannan miniature pigs may maintain normal renal function for a certain period in rhesus macaques and effectively overcome HAR, confirming the effectiveness of GTKO pigs for xenotransplantation.

Objective:Glaucoma is a multifactorial optic neuropathy with a high rate of irreversible visual loss,and its pathogenesis is complex and still unclear.Elevated intraocular pressure(IOP)is well recognized as the sole modifiable risk factor for the development of glaucoma in the majority of cases.This study aims to compare 2 different methods of inducing chronic ocular hypertension by circumlimbal suture or by laser burns in degree and lasting time of the IOP,different status of the retina and retinal ganglion cells(RGCs),and changes of the microstructure of neurons. Methods:The chronic ocular hypertension models were induced by 2 different ways.One kind of the models was built by unilateral circumlimbal suture(10/0)implantation(suture group),another kind of model was built by laser burns at trabecular meshwork and episcleral veins(laser group).The untreated contralateral eye served as the control group.Changes in IOP were observed and regularly monitored in the 2 groups of rats.HE staining was applied to observe the retinal and optic nerve pathology.Transmission electron microscope(TEM)was used to observe the mitochondrial morphology.RGCs were specifically labeled with Brn3b antibody and counted.The expression of caspase-3 was detected by Western blotting to clarify the apoptosis of RGCs. Results:Compared with the control group,IOP were significantly increased in the suture group and the laser group(both P<0.05).The suture group induced a 1.5-fold elevation of IOP,and sustained for 8 weeks.The laser group induced a 2-fold elevation of IOP for 12 weeks.Both methods could cause RGCs loss(both P<0.05),which were verified by pathology and immune staining of Brn3b.The expressions of caspase-3 were also increased(both P<0.05).The mitochondrial morphology became more fragment,which changed from long shape to round and small one under TEM in 2 models.For comparison,the pathology changes of retinal structure in suture group were not obviously than those in the laser group. Conclusion:Circumlimbal suture can build an effective model of chronic elevated IOP and induce glaucomatous pathologic changes similar to those in the laser photocoagulation,but the pathologic changes are milder than those in laser photocoagulation.Compare with translimbal laser photocoagulation,equipment and skill demand for circumlimbal suture is less.

Background::Pathological scars are a disorder that can lead to various cosmetic, psychological, and functional problems, and no effective assessment methods are currently available. Assessment and treatment of pathological scars are based on cutaneous manifestations. A two-photon microscope (TPM) with the potential for real-time non-invasive assessment may help determine the under-surface pathophysiological conditions in vivo. This study used a portable handheld TPM to image epidermal cells and dermal collagen structures in pathological scars and normal skin in vivo to evaluate the effectiveness of treatment in scar patients. Methods::Fifteen patients with pathological scars and three healthy controls were recruited. Imaging was performed using a portable handheld TPM. Five indexes were extracted from two dimensional (2D) and three dimensional (3D) perspectives, including collagen depth, dermo-epidermal junction (DEJ) contour ratio, thickness, orientation, and occupation (proportion of collagen fibers in the field of view) of collagen. Two depth-dependent indexes were computed through the 3D second harmonic generation image and three morphology-related indexes from the 2D images. We assessed index differences between scar and normal skin and changes before and after treatment.Results::Pathological scars and normal skin differed markedly regarding the epidermal morphological structure and the spectral characteristics of collagen fibers. Five indexes were employed to distinguish between normal skin and scar tissue. Statistically significant differences were found in average depth ( t = 9.917, P <0.001), thickness ( t = 4.037, P <0.001), occupation ( t= 2.169, P <0.050), orientation of collagen ( t = 3.669, P <0.001), and the DEJ contour ratio ( t = 5.105, P <0.001). Conclusions::Use of portable handheld TPM can distinguish collagen from skin tissues; thus, it is more suitable for scar imaging than reflectance confocal microscopy. Thus, a TPM may be an auxiliary tool for scar treatment selection and assessing treatment efficacy.

ObjectiveTo explore the interaction between bruceoside B and gut microbiota and the inhibitory activity of its metabolites on human lung cancer A549 cells， and to explore the value of bruceoside B in the treatment of non-small cell lung cancer（NSCLC）. MethodBruceoside B was co-incubated with the human gut microbiota under anoxic conditions in vitro， and ultra high performance liquid chromatography-quadrupole-time-of-flight mass spectrometry（UPLC-Q-TOF-MS） was used to analyze the metabolic transformation products. Cell counting kit-8（CCK-8） assay was performed to determine the effects of bruceoside B and its metabolites on the proliferation of human lung cancer A549 cells and the half inhibitory concentration（IC₅₀） was calculated. Five healthy male rats were gavaged with bruceoside B（2 mg·kg^-1） for 7 days after adaptive feeding. The feces of rats were collected before and after administration. 16S rRNA sequencing was used to assess gut microbiota. ResultBruceoside B was mainly metabolized to brusatol by human gut microbiota， the IC₅₀ of bruceoside B and the conversion product to A549 cells were 1 755.50， 19.57 μmol·L^-1， respectively， and the conversion product had a better activity at inhibiting A549 cells proliferation than bruceoside B. Additionally， The results of intestinal flora analysis showed no significant differences in α diversity and β diversity of gut microbiota after administration. In terms of species abundance， at the phylum level， bruceoside B decreased the relative abundance of Actinobacteriota and Proteobacteria， increased the relative abundance of Firmicutes， Patescibacteria and Cyanobacteria. At the genus level， bruceoside B decreased the relative abundance of Staphylococcus， Aerococcus and Psychrobacter， increased the relative abundance of Romboutsia， Lactobacillus， Clostridium sensu stricto 1， Norank-f-norank-o-Clostridia-UCG-014， Turicibacter， Allobaculum and Candidatus Saccharimonas. The results of functional prediction showed that the gut microbiota functional compositions were relatively stable. ConclusionBruceoside B can be deglycosylated by intestinal flora and converted into brusatol， with a significant increase in antitumor activity. The administration of bruceoside B will not cause significant changes in the structure and function of the intestinal flora， resulting in intestinal microecological balance disorders， and the administration appears to be beneficial to the intestinal flora of NSCLC patients.

Objective:To obsere the clinical efficacy of Hall technology in the treatment of primary molar caries in children with au-tism.Methods:80 children aged 4-8 years with primary molar caries,40 normal children and 40 children with autism,with a total of 153 primary molars.The normal and the autism children were respectively divided into 2 groups(n=20)randomly.The normal children were grouped into resin filling(CR)of 38 teeth and Hall technology group(CH)of 39 teeth,the autistic children were grouped into resin filling(AR)of 37 teeth and Hall technology group(AH)of 38 teeth.Corresponding treatment was given to the pa-tients in the groups.The duration of oral treatment time,Frankl scores,Houpt scores and parental satisfaction scoves were compaired among groups.The patients were followed up for 6,12,18 and 24 months and the treatment outcomes were compared among groups.Results:There was no statistically difference between CH group and AH group in terms of operating time,compliance,Houpt score and follow-up effects(P＞0.05).The satisfaction of parents in the AR group was the lowest(P＜0.05).Hall technology treatment re-mained effective rate was higher than resin filling at the 24 month follow-up(P＜0.05).Conclusion:Hall technology is more effective than traditional resin filling in the treatment of primary molars caries in children with autism.

Objective:To evaluate the healthcare accessibility and quality of diagnosis and treatment services of Internet hospitals in China.Methods:One hundred and eighty Internet hospitals in 60 cities were seleted based on the sampling of development levels in the eastern, central and western regions of China. From April to May 2023, standardized patients methodology was applied to evaluate the accessibility(including the number of Internet hospitals, functional settings, online doctor status, the doctor′s attending rate and consultation fees) and diagnosis and treatment service quality(including the diagnosis and treatment services quality, response speed and patient′s evaluation) of Internet hospitals.Results:The average opening rate of Internet hospitals in China was 52.9% (560/1 058), the average online rate of doctors was 64.2% (1 099/1 713), the average doctor′s attending rate was 33.6% (112/333), the average consultation fee was 4.85 yuan, the average score of consultation was 1.92 out of 9, the average score of diagnosis and treatment was 1.12 out of 4, the average score of the response speed was 1.70 out of 3, and patient satisfaction was 2.73 out of 3.Conclusions:The Internet hospital accessibility in China is unevenly developed, and the overall quality of diagnosis and treatment is low. It is recommended to accurately position and optimize the function of Internet hospital, establish the incentive mechanism for online consultation doctors, construct and improve the regulatory system of Internet hospital diagnosis and treatment, so as to improve the accessibility and quality of diagnosis and treatment of Internet hospitals.

Objective: This study aimed to determine the predictive performance of non-contrast CT (NCCT) signs for hemorrhagic growth after intracerebral hemorrhage (ICH) when stratified by onset-to-imaging time (OIT). Materials and Methods: 1488 supratentorial ICH within 6 h of onset were consecutively recruited from six centers between January 2018 and August 2022. NCCT signs were classified according to density (hypodensities, swirl sign, black hole sign, blend sign, fluid level, and heterogeneous density) and shape (island sign, satellite sign, and irregular shape) features. Multivariable logistic regression was used to evaluate the association between NCCT signs and three types of hemorrhagic growth: hematoma expansion (HE), intraventricular hemorrhage growth (IVHG), and revised HE (RHE). The performance of the NCCT signs was evaluated using the positive predictive value (PPV) stratified by OIT. Results: Multivariable analysis showed that hypodensities were an independent predictor of HE (adjusted odds ratio [95% confidence interval] of 7.99 [4.87–13.40]), IVHG (3.64 [2.15–6.24]), and RHE (7.90 [4.93–12.90]). Similarly, OIT (for a 1-h increase) was an independent inverse predictor of HE (0.59 [0.52–0.66]), IVHG (0.72 [0.64–0.81]), and RHE (0.61 [0.54– 0.67]). Blend and island signs were independently associated with HE and RHE (10.60 [7.36–15.30] and 10.10 [7.10–14.60], respectively, for the blend sign and 2.75 [1.64–4.67] and 2.62 [1.60–4.30], respectively, for the island sign). Hypodensities demonstrated low PPVs of 0.41 (110/269) or lower for IVHG when stratified by OIT. When OIT was ≤ 2 h, the PPVs of hypodensities, blend sign, and island sign for RHE were 0.80 (215/269), 0.90 (142/157), and 0.83 (103/124), respectively. Conclusion Hypodensities, blend sign, and island sign were the best NCCT predictors of RHE when OIT was ≤ 2 h. NCCT signs may assist in earlier recognition of the risk of hemorrhagic growth and guide early intervention to prevent neurological deterioration resulting from hemorrhagic growth.

ObjectiveTo explore the mechanism of Huangqisan （HQS） in regulating autophagy to alleviate hepatic steatosis and improve non-alcoholic fatty liver disease （NAFLD） based on adenosine 5'-monophosphate-activated protein kinase （AMPK）/mammalian target of rapamycin （mTOR） signaling pathway. MethodThe main chemical components and targets of HQS and NAFLD-related targets were collected from database and the intersection targets were used for Gene Ontology （GO） functional enrichment analysis and Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment analysis. The protein-protein interaction （PPI） network was constructed， and in vivo experimental verification was conducted. Sixty C57BL/6J male mice were randomly divided into normal control group （NCD）， model group high-fat diet （HFD）， metformin group （MET， 0.25 g·kg^-1）， low-dose Huangqisan group （HQS-L， 0.5 g·kg^-1）， and the high-dose Huangqisan group （HQS-H， 1 g·kg^-1）， with 12 mice in each group after a one-week acclimatization period. NAFLD model was induced by HFD， and intragastric administration was performed at the same time， once a day for 13 weeks. Random blood glucose， serum total cholesterol （TC）， triglyceride （TG）， non-esterified fatty acid （NEFA）， low density lipoprotein-chdesterol （LDL-C） levels， and liver TG content were determined. The liver weight was weighed， and liver index was calculated. Hematoxylin-eosin （HE） staining， oil red O staining， transmission electron microscope （TEM）， real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）， and Western blot were used to verify the effect and reveal the potential mechanism of C57BL/6J mice in vivo. ResultThrough network pharmacology analysis， combined with previous studies， it was predicted that HQS may improve NAFLD by regulating autophagy via the AMPK/mTOR signaling pathway. The result of in vivo experiment showed that， as compared with NCD group， random blood glucose， body weight， serum TC， LDL-C， NEFA， liver weight， liver index， and liver TG content of mice in the HFD groups were significantly increased （P<0.01）. HE staining showed massive lipid droplets （LDs） vacuolated， oil red O staining showed lipid accumulation in liver cells， and no obvious autophagosomes and autolysosome were observed under TEM. The relative mRNA expression of LC3A、LC3B、AMPKα1 and protein expression of AMPK， phosphory phosphorylated（p）-AMPK， and p-AMPK/AMPK were significantly down-regulated （P<0.01）， while the protein expression of microtubule-associated protein 1 light chain 3 （LC3）Ⅱ/Ⅰ and p-mTOR was significantly up-regulated （P<0.01）. As compared with HFD groups， liver weight， serum TG， and NEFA levels in HQS-L and HQS-H groups were significantly deceased （P<0.05， P<0.01）. HE staining and oil red O staining showed the improvement of liver pathological changes after HQS administration. Under TEM， a small amount of autophagosome and autolysosome were observed. Besides， liver index was significantly decreased in the HQS-L group （P<0.01）， and random blood glucose， serum TC level and liver TG content were significantly decreased in the HQS-H group （P<0.05）. The results of Western blot and Real-time PCR showed that the mRNA expression of LC3A and LC3B and the protein expression of LC3Ⅱ/Ⅰ， p-AMPK， and p-AMPK/AMPK were significantly up-regulated （P<0.01）， while the mRNA expressions of p62 and protein expression of p62 and p-mTOR were significantly down-regulated （P<0.05， P<0.01）. ConclusionHQS may promote autophagy and restore autophagy flux via the AMPK/mTOR signaling pathway to alleviate hepatic steatosis improving NAFLD.

Objective:To construct the "Internet +" peripherally inserted central catheter (PICC) home nursing service quality evaluation index system, so as to provide objective basis for standardizing the "Internet +" PICC home nursing service quality evaluation.Methods:Based on the three-dimensional quality structure model of "structure-process-outcome", combined with literature research, semi-structured interviews, group discussions and Delphi method, 20 experts who were engaged in intravenous therapy nursing and had management or practical experience in "Internet + nursing services" were selected for two rounds of consultation to determine the "Internet?+" PICC home nursing service quality evaluation index system. The positive coefficient of experts was expressed by the effective response rate of the questionnaire and the rate of expert opinion submission. The degree of expert authority was expressed by the expert authority coefficient. The coordination degree of expert opinions was represented by the Kendall harmony coefficient. The concentration degree of expert opinions was usually expressed in terms of the importance assigned to the indicator, the coefficient of variation, and the full score rate (%) .Results:The effective response rates of the two rounds of consultation questionnaires were all 100%, with expert authority coefficients of 0.793 and 0.848 respectively. The Kendall coordination coefficients of expert opinions were 0.202-0.216 and 0.222-0.270 respectively ( P<0.05). After the second round of expert consultation, the mean importance assigned to all indicators was greater than 4.0, the coefficient of variation was less than 0.2, and the full score rate was greater than 20%. The final "Internet +"PICC home nursing service quality evaluation index system included 3 first-level indicators, 12 second-level indicators and 64 third-level indicators. Conclusions:The "Internet + " PICC home nursing service quality evaluation index system constructed is scientific, reliable and practical, and reflects the specialty characteristics. It can effectively evaluate the "Internet + " PICC home nursing service quality and provide guidance for continuous improvement.