Objective:To discuss the construction of a NOD-signal regulatory protein α(SIRPA)gene knock-in mouse model based on clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated protein(Cas)9 technology,and to provide the reference for the establishment of more advanced humanized mouse models.Methods:Based on CRISPR/Cas9 technology,the NOD-background SIRPA gene was knocked into the BALB/c mouse fertilized eggs,and homozygous mice with stable genotypes(SIRPA-KI mice)were obtained through expansion and breeding for experiments;PCR primers were designed,and mouse genotypes were identified by nucleic acid electrophoresis.The mice were divided into C57BL/6 group,BALB/c group,and SIRPA-KI group according to their strains,and 3 mice with similar ages were randomly selected from each group for experiments.Mature bone marrow-derived macrophages were co-incubated with human CD47-Fc fusion protein,stained with Streptavidin PE/Cy7,and the mean fluorescence intensity(MFI)of PE/Cy7 was detected by flow cytometry to compare the ability of SIRPA in the mice from various groups to bind human CD47 Fc.Each mouse was intravenously injected with 2×10? carboxyfluorescein diacetate,succinimidyl ester(CFSE)-labeled human red blood cells,and the phagocytosis of human red blood cells by macrophages in various groups was detected by in vivo phagocytosis assay.One BALB/c mouse and one SIRPA-KI mouse were randomly selected to induce mature bone marrow-derived macrophages,and the phagocytosis of human red blood cells by macrophages in various groups was detected by in vitro phagocytosis assay.Results:BLAB/c SIRPANOD/NOD homozygous mice were successfully obtained.The flow cytometry results showed that compared with C57BL/6 group,the MFI of the mice in BALB/c group was significantly increased(P<0.01);compared with C57BL/6 group and BALB/c group,the MFI of the mice in SIRPA-KI group was significantly increased(P<0.01).The in vivo phagocytosis assay results showed that the macrophages in C57BL/6 group exhibited the fastest overall clearance rate of human red blood cells;at 6 h of macrophage phagocytosis,compared with SIRPA-KI group,the residual percentage of the human red blood cells in C57BL/6 group was significantly decreased(P<0.01)and was closed to 0%;compared with SIRPA-KI group,the residual percentage of the human red blood cells in BALB/c group was significantly decreased(P<0.05).The in vitro phagocytosis assay results showed that the macrophages in BALB/c group significantly phagocytosed the human red blood cells,with a high phagocytic index of 30.7%;compared with BALB/c group,the phagocytic index of the macrophages in SIRPA-KI group was significantly decreased(P<0.01).Conclusion:The study successfully constructs a mouse model with enhanced affinity for human CD47 and significantly inhibited phagocytosis of human red blood cells by knocking the NOD-background SIRPA gene into the BALB/c strain using CRISPR/Cas9 technology,providing a superior human cell xenotransplantation efficiency.

Objective:To investigate the protective effect and its mechanism of proscillaridin A (PSD-A) on podocyte injury in lupus nephritis (LN).Methods:Molecular docking and surface plasmon resonance techniques were used to analyze the binding status of PSD-A to signal transducer and activator of transcription 1 (STAT1). The immortalized human podocyte injury model in the lupus group was induced by the serum of systemic lupus erythematosus patients, and the control and PSD-A intervention (2 nmol/L, 4 nmol/L) groups were also set up. Six female 12-week-old C57BL/6 mice were designated as the control group, and 12 female 12-week-old MRL/lpr lupus mice were randomly divided into lupus group and PSD-A intervention group by random number table method. The PSD-A intervention group was intraperitoneally injected with 5 mg/kg PSD-A, once per week for 6 consecutive weeks. While the control group and the lupus group were intraperitoneally injected with the same volume of the solvent without PSD-A. Western blotting and real-time quantitative PCR were employed to detect the relative protein and mRNA expression levels of podocin, STAT1, and interferon-induced protein with tetratricopeptide repeat 1 (IFIT1) in podocytes of each group. Enzyme-linked immunosorbent assay was used to detect the levels of serum anti-double strand DNA antibody and interferon-α in mice. Coomassie brilliant blue was used to detect the urinary protein level. HE, PAS, Masson and PASM staining and transmission electron microscopy were used to observe the pathological changes of renal tissues. Immunohistochemistry was used to examine the protein expression of podocin, STAT1 and IFIT1 in renal tissues.Results:Molecular docking and surface plasmon resonance techniques proved that PSD-A could bind to STAT1 protein and they exhibited a robust binding affinity. The podocyte experiments showed that, compared with the lupus group, the relative expression levels of podocin protein and mRNA in the PSD-A intervention group were upregulated, while the relative expression levels of STAT1 and IFIT1 protein and mRNA were downregulated (all P<0.05). The animal experiments showed that, compared with the lupus group, the serum levels of anti-double strand DNA antibody, interferon-α, and urinary protein in PSD-A intervention group were decreased, the pathological damage of renal tissues was alleviated, and the injury of renal podocytes was reduced. Immunohistochemical staining showed that the relative protein expression levels of STAT1 and IFIT1 of renal tissues in the PSD-A intervention group were lower than those in the lupus group (all P<0.05). Conclusion:PSD-A can play a protective role in podocyte injury in LN, and its mechanism may be related to the inhibition of the STAT1 signaling pathway.

Although cancer immunotherapy has made great strides in the clinic, it is still hindered by the tumor immunosuppressive microenvironment (TIME). The stimulator of interferon genes (STING) pathway which can modulate TIME effectively has emerged as a promising therapeutic recently. However, the delivery of most STING agonists, specifically cyclic dinucleotides (CDNs), is performed intratumorally due to their insufficient pharmacological properties, such as weak permeability across cell membranes and vulnerability to nuclease degradation. To expand the clinical applicability of CDNs, a novel pH-sensitive polycationic polymer-modified lipid nanoparticle (LNP-B) system was developed for intravenous delivery of CDNs. LNP-B significantly extended the circulation of CDNs and enhanced the accumulation of CDNs within the tumor, spleen, and tumor-draining lymph nodes compared with free CDNs thereby triggering the STING pathway of dendritic cells and repolarizing pro-tumor macrophages. These events subsequently gave rise to potent anti-tumor immune reactions and substantial inhibition of tumors in CT26 colon cancer-bearing mouse models. In addition, due to the acid-sensitive property of the polycationic polymer, the delivery system of LNP-B was more biocompatible and safer compared with lipid nanoparticles formulated with an indissociable cationic DOTAP (LNP-D). These findings suggest that LNP-B has great potential in the intravenous delivery of CDNs for tumor immunotherapy.

Gastrointestinal tumors （GTs）， including colorectal cancer， gastric cancer， liver cancer， pancreatic cancer， and esophageal cancer， are increasing in incidence worldwide and have become one of the major diseases threatening human health. Tumor stem cells （TSCs）， an undifferentiated subpopulation within tumor tissues， possess biological characteristics such as self-renewal， multidirectional differentiation， high tumorigenicity， and resistance to radiochemotherapy. They play an important role in the occurrence， progression， recurrence， and metastasis of GTs and have increasingly become a research hotspot in GT treatment. Although modern medicine has made remarkable progress， there remain many problems in therapeutic approaches targeting TSCs. In this context， traditional Chinese medicine （TCM）， with its favorable safety profile and multi-target mechanisms， has shown potential advantages and value in regulating TSCs. It can reduce TSC drug resistance， enhance the sensitivity of tumor cells to chemotherapeutic agents， inhibit tumor growth and metastasis， and has shown unique advantages in improving the quality of life and prolonging the survival of GT patients. Studies have found that active components of Chinese medicine， such as terpenoids， polyphenols， flavonoids， glycosides， and quinones， and Chinese medicine compound formulas， including Zuojin pills， Sijunzi decoction， Biejiajian pills， and Xuanfu Daizhe decoction， can inhibit TSCs-related signaling pathways such as the Notch signaling pathway， the Wnt/β-catenin signaling pathway， the phosphatidylinositol 3-kinase/protein kinase B （PI3K/Akt） signaling pathway， and the Hippo signaling pathway. They also reduce the expression of TSC surface markers， including sex-determining region Y-box 2 （SOX2）， sex-determining region Y-box 9 （SOX9）， octamer-binding transcription factor 4 （OCT4）， prominin-1 （CD133）， cluster of differentiation 44 （CD44）， cluster of differentiation 24 （CD24）， and thyroid transmembrane protein 1 （CD90）， thereby hindering TSC differentiation， accelerating their metabolic processes， improving the tumor microenvironment， and consequently inhibiting GT growth. This study collects and analyzes recent research on the regulation of TSCs by TCM in the treatment of GT， aiming to provide a new theoretical basis for tumor therapy with TCM， expand its application in the comprehensive treatment of GT， and offer new therapeutic ideas and methods for clinical practice.

Objective Through reviewing the herbs,medical books and classic prescriptions of the past dynasties,the herbal examination of Gypsum fibrosum and its analogs was carried out to clarify the relationship between them.The qualitative analysis was carried out by single-point Raman,infrared,near infrared and XRD techniques,the quantitative analysis of the main components was carried out by EDTA titration and Raman surface scanning technique,the elemental analysis was carried out by ICP-OES,and the differences between gypsum and argillite were observed by scanning electron microscope.Results Gypsum fibrosum is mixed with marble,feldspar,north cold-water stone,south cold-water stone and xuanjing stone and so on.Before the Ming Dynasty,there was no differentiation between soft and anhydrite,and after the Ming Dynasty,it was clear that gypsum was soft gypsum and feldspar was anhydrite;in modern times,marble is also made to be gypsum for medicinal use;Feldspar is anhydrite and is no longer made to be used for medicinal use,north cold-water stone is red gypsum,and south cold-water stone is calcite.Gypsum,south coldwater stone is calcite.The peaks of argillite appeared to be buried in the Raman spectrum compared with Gypsum fibrosum,and the original profiles of both infrared and near-infrared were basically the same,and the near-infrared model established by the preprocessing method of the first-order derivatives plus vector normalization(5-point smoothing)could effectively differentiate between Gypsum fibrosum and its analogues,but it could not differentiate between argillite and Gypsum fibrosum with high content.XRD showed that there are differences in the relative intensities of the peaks of argillite and Gypsum fibrosum,and some XRD shows that there are differences in the relative intensities of the peaks of basalt and Gypsum fibrosum,and some of the peaks of basalt are characterized by impurities such as quartz,and the contents of trace elements such as Fe,Mn,Cr,Pb,Hg and As are higher in basalt.The peak shapes of XRD,Raman spectra,infrared spectra and near-infrared spectra of Gypsum fibrosum and calcined gypsum are closer but can still be distinguished,and the Ca content of calcined gypsum is higher than that of gypsum.Commercially available south chrysocolla and stalactite source are carbonate minerals calcite calcite.Spectral detection can not be distinguished,the trace element content is basically the same,but the traits are different.North chrysocolla(red gypsum)is higher than the Fe content of commercially available white gypsum.Raman surface scanning not only can be a qualitative and quantitative determination of minerals such as gypsum and other minerals,and the results of the content of the titration is basically similar,but also to determine the state of Gypsum fibrosum and calcined gypsum calcined water loss status and the degree of calcined gypsum.The results of Raman surface scanning are similar to the titration results.Conclusion This study can provide a scientific basis for the traceability of Gypsum fibrosum,and can better guide the clinical use of medicine and the rational use of resources.

Objective Through reviewing the herbs,medical books and classic prescriptions of the past dynasties,the herbal examination of Gypsum fibrosum and its analogs was carried out to clarify the relationship between them.The qualitative analysis was carried out by single-point Raman,infrared,near infrared and XRD techniques,the quantitative analysis of the main components was carried out by EDTA titration and Raman surface scanning technique,the elemental analysis was carried out by ICP-OES,and the differences between gypsum and argillite were observed by scanning electron microscope.Results Gypsum fibrosum is mixed with marble,feldspar,north cold-water stone,south cold-water stone and xuanjing stone and so on.Before the Ming Dynasty,there was no differentiation between soft and anhydrite,and after the Ming Dynasty,it was clear that gypsum was soft gypsum and feldspar was anhydrite;in modern times,marble is also made to be gypsum for medicinal use;Feldspar is anhydrite and is no longer made to be used for medicinal use,north cold-water stone is red gypsum,and south cold-water stone is calcite.Gypsum,south coldwater stone is calcite.The peaks of argillite appeared to be buried in the Raman spectrum compared with Gypsum fibrosum,and the original profiles of both infrared and near-infrared were basically the same,and the near-infrared model established by the preprocessing method of the first-order derivatives plus vector normalization(5-point smoothing)could effectively differentiate between Gypsum fibrosum and its analogues,but it could not differentiate between argillite and Gypsum fibrosum with high content.XRD showed that there are differences in the relative intensities of the peaks of argillite and Gypsum fibrosum,and some XRD shows that there are differences in the relative intensities of the peaks of basalt and Gypsum fibrosum,and some of the peaks of basalt are characterized by impurities such as quartz,and the contents of trace elements such as Fe,Mn,Cr,Pb,Hg and As are higher in basalt.The peak shapes of XRD,Raman spectra,infrared spectra and near-infrared spectra of Gypsum fibrosum and calcined gypsum are closer but can still be distinguished,and the Ca content of calcined gypsum is higher than that of gypsum.Commercially available south chrysocolla and stalactite source are carbonate minerals calcite calcite.Spectral detection can not be distinguished,the trace element content is basically the same,but the traits are different.North chrysocolla(red gypsum)is higher than the Fe content of commercially available white gypsum.Raman surface scanning not only can be a qualitative and quantitative determination of minerals such as gypsum and other minerals,and the results of the content of the titration is basically similar,but also to determine the state of Gypsum fibrosum and calcined gypsum calcined water loss status and the degree of calcined gypsum.The results of Raman surface scanning are similar to the titration results.Conclusion This study can provide a scientific basis for the traceability of Gypsum fibrosum,and can better guide the clinical use of medicine and the rational use of resources.

Objective:To investigate the protective effect and its mechanism of proscillaridin A (PSD-A) on podocyte injury in lupus nephritis (LN).Methods:Molecular docking and surface plasmon resonance techniques were used to analyze the binding status of PSD-A to signal transducer and activator of transcription 1 (STAT1). The immortalized human podocyte injury model in the lupus group was induced by the serum of systemic lupus erythematosus patients, and the control and PSD-A intervention (2 nmol/L, 4 nmol/L) groups were also set up. Six female 12-week-old C57BL/6 mice were designated as the control group, and 12 female 12-week-old MRL/lpr lupus mice were randomly divided into lupus group and PSD-A intervention group by random number table method. The PSD-A intervention group was intraperitoneally injected with 5 mg/kg PSD-A, once per week for 6 consecutive weeks. While the control group and the lupus group were intraperitoneally injected with the same volume of the solvent without PSD-A. Western blotting and real-time quantitative PCR were employed to detect the relative protein and mRNA expression levels of podocin, STAT1, and interferon-induced protein with tetratricopeptide repeat 1 (IFIT1) in podocytes of each group. Enzyme-linked immunosorbent assay was used to detect the levels of serum anti-double strand DNA antibody and interferon-α in mice. Coomassie brilliant blue was used to detect the urinary protein level. HE, PAS, Masson and PASM staining and transmission electron microscopy were used to observe the pathological changes of renal tissues. Immunohistochemistry was used to examine the protein expression of podocin, STAT1 and IFIT1 in renal tissues.Results:Molecular docking and surface plasmon resonance techniques proved that PSD-A could bind to STAT1 protein and they exhibited a robust binding affinity. The podocyte experiments showed that, compared with the lupus group, the relative expression levels of podocin protein and mRNA in the PSD-A intervention group were upregulated, while the relative expression levels of STAT1 and IFIT1 protein and mRNA were downregulated (all P<0.05). The animal experiments showed that, compared with the lupus group, the serum levels of anti-double strand DNA antibody, interferon-α, and urinary protein in PSD-A intervention group were decreased, the pathological damage of renal tissues was alleviated, and the injury of renal podocytes was reduced. Immunohistochemical staining showed that the relative protein expression levels of STAT1 and IFIT1 of renal tissues in the PSD-A intervention group were lower than those in the lupus group (all P<0.05). Conclusion:PSD-A can play a protective role in podocyte injury in LN, and its mechanism may be related to the inhibition of the STAT1 signaling pathway.

By reviewing the historical materia medica， medical books and modern literature， this paper has systematically sorted out and verified the name， origin， quality and other aspects of Ophicalcitum. After herbal textual research， it is shown that before the Qing dynasty， the mineral medicine was mostly recorded in the name of Huarushi， but now it is called Huaruishi， and there is another mixed name Baiyunshi. The light white spots described in the historical materia medica are consistent with the characteristics of marble with sparkling star-like luster， combined with the color like sulfur， color are green， black spots and other serpentine features， it is deduced that it is serpentine marble， consistent with the present-day Ophicalcitum， and Ophicalcitum in the Song dynasty has a high content of serpentine. The main producing areas are Henan， Shaanxi， Shanxi and Sichuan， Jiangsu， Zhejiang， Hebei and other places are also available. Successive generations of materia medica on the quality evaluation of Ophicalcitum is less， the modern to neat and firm in the texture， sandwiched with yellow-green mottled for the best. Ophicalcitum is acidic， astringent and neutral in nature， belonging to the liver meridian， with the efficacy of treatment of gold sores and blood flow， internal leakage of cataracts， dropping afterbirth， now describing its efficacy as removing blood stasis and stopping bleeding. In ancient times， the earliest processing method was burning， followed by calcination by sulphur， calcination， quenching with vinegar and other methods. In modern times， it has been simplified to open calcination， processing with vinegar and the addition of water quenching. The gravimetric method and ethylenediaminetetraacetic acid titration were used to detect the contents of CO₃^2- and CaCO₃ in Ophicalcitum， respectively， and it was found that the gap in CaCO₃ content among commercially available products was wide， and the content of CaCO₃ in sample S13 and sample S18 was the same， but their compositions were different， and according to the contents of CO₃^2- and CaCO₃， the dolomite and calcite contents could be calculated， of which the higher the calcite content the more obvious the sparkling star-like luster. Raman spectroscopy and X-ray diffraction（XRD） were used to detect the physical phase composition of the powder of the samples， and Raman spectroscopy was used for the rapid non-destructive testing of the striped part， which showed that Ophicalcitum was mainly composed of dolomite， calcite， serpentine， olivine and pyroxene， with serpentine dominanting the striped part. In summary， the 2020 edition of Chinese Pharmacopoeia stipulates that the content of CaCO₃ in Ophicalcitum is not less than 40%， which is difficult to control its quality， and it is suggested to increase the detection of CO₃^2- content. This study can provide a scientific basis for the traceability of Ophicalcitum and better guide the clinical medication and rational utilization of resources.

Objective:This study aimed to reveal the distribution and course of the branches of the infraorbital nerve(IN),its communication relationship between the branches of the infraorbital nerve and facial nerve,so as to provide morphological basis for clinical implementation of accurate infraorbital nerve trunk in the infraorbital canal,regional facial anesthesia and facial surgery,so as to improve the success rate of maxillofacial surgery.Methods:25 adult cada-vers with formalin immobilized semi-face were selected.Exclude facial defect samples caused by tumor,trauma,deformity,surgery,etc.The length and diameter of the trunk of the infraorbital nerve and the length of the infraorbital canal were measured.The total number of infraorbital nerve and the number of branches were counted,and the course,distribution and communication relationship between infraorbital nerve and facial nerve were investigated.Results:The length of infraorbital nerve trunk ranged from 19.61 to 44.47 mm,with an average length of(23.33±4.95)mm.The length of infraorbital canal ranged from 9.49 to 31.21 mm,with an average length of(12.87±3.99)mm.The number of infraorbital nerve branches ranged from 5 to 12,and the average number was(7.29±2.29).The number of upper labial branches was the widest,ranging from 1 to 5,while the distribution area of eyelid branches was the narrowest.There are(were)a large number of intersections and anastomoses between the infraorbital nerve and the facial nerve,forming a complex multi-layer network structure.Conclusion:The infraorbital nerve trunk and the infraorbital canal va-ry in length.The number and distribution range of infraorbital nerve branches are not constant,and the communication relationship between infraorbital nerve and facial nerve is complicated.

OBJECTIVE: Because of the limited number of studies and small sample sizes, whether metabolic syndrome (MS) leads to the occurrence and progression of osteoporosis and the possible underlying mechanisms require further investigation. This study aimed to investigate the association between MS and osteoporosis, along with its influencing factors. METHODS: This observational cross-sectional study included 139,470 individuals aged ≥ 18 years who underwent health examinations from September 2014 to March 2022. Based on bone mineral density (BMD) screening results, the participants were categorized into a suspected osteoporosis or non-osteoporosis group (control). Participants were further divided into those who met 0 MS criteria, 1 MS criterion, 2 MS criteria, and ≥ 3 MS criteria (MS group). Participants who had undergone health examinations at least twice formed the follow-up cohort; a self-matched analysis was performed on those with follow-up periods ≥ 5 years and unchanged MS grouping. RESULTS: Several examination indicators in the suspected osteoporosis group showed statistically significant differences compared with the control group. The proportion of suspected osteoporosis in the MS group was significantly increased compared with that in the 0 MS criteria group (odds ratio [ OR]: 1.215, Z = 29.11, P < 0.001, 95% confidence interval: 1.199-1.231). After adjusting for age, sex, smoking, and alcohol consumption, the 2 MS criteria group and MS group still had OR values > 1 ( P < 0.001). In the follow-up cohort, the proportion of suspected osteoporosis increased gradually with an increase in the number of MS criteria met at baseline and during each follow-up visit ( P < 0.05), with the highest proportion observed in the MS group. However, the proportion of suspected osteoporosis did not increase significantly over time in the different MS groups ( P > 0.05). In the follow-up cohort, the proportion of individuals transitioning from normal BMD to suspected osteoporosis was higher in the MS group after ≥ 5 years of follow-up compared with the group meeting 0 MS criteria (0.08% versus 1.15%, χ ² = 10.76, P = 0.001). There was no significant difference in BMD values for the 0 MS criteria group after 5 years ( P > 0.05), whereas the other three groups experienced a significant decrease in BMD values after 5 years ( P < 0.05). CONCLUSION MS is an independent risk factor for osteoporosis, and the effect of risk factors related to MS on osteoporosis may exceed that of aging alone. The specific mechanisms warrant further investigation.
Humans ; Osteoporosis/etiology* ; Female ; Male ; Metabolic Syndrome/complications* ; Middle Aged ; Cross-Sectional Studies ; China/epidemiology* ; Aged ; Adult ; Bone Density ; Risk Factors