ObjectiveTo establish the specific chromatogram and quantitative analysis of multi-components by single-marker（QAMS） based on linear calibration using two reference substances（LCTRS）， explore the consistency between Hedyotis Herba dispensing granules and standard decoction， and evaluate the quality of the dispensing granules. MethodsHigh performance liquid chromatography（HPLC） specific chromatogram was established based on 15 batches of Hedyotis Herba standard decoction and 10 batches of the dispensing granules， and LCTRS was used to locate chromatographic peaks. The actual retention times of 7 characteristic peaks in the specific chromatogram was measured on 24 different types of C₁₈ columns， taking deacetyl asperulosidic acid and asperulosidic acid as the dual standard compounds， the retention times of the other 5 characteristic peaks were predicted and validated. Based on this， QAMS was developed to determine the contents of four components（deacetyl asperulosidic acid， deacetyl asperulosidic acid methyl ester， asperulosidic acid， and p-coumaric acid）. Then， the relative correction factors of deacetyl asperulosidic acid， deacetyl asperulosidic acid methyl ester and p-coumaric acid were calculated using the reference peak of asperulosidic acid in the dual standard compounds， and each component was quantified accordingly. Finally， the consistency between the dispensing granules and standard decoction was assessed by taking extract rate of the standard decoction， consistency of the specific chromatograms， contents and transfer rates of the indicator components as indexes， and the quality of the dispensing granules was evaluated. ResultsThere were 7 common peaks in the characteristic chromatogram of samples of Hedyotis Herba standard decoction and the dispensing granules， and four of them were identified by reference standards， namely deacetyl asperulosidic acid（peak 1）， deacetyl asperulosidic acid methyl ester（peak 3）， asperulosidic acid（peak 6） and p-coumaric acid（peak 7）. The similarity between the dispensing granules and the standard decoction was >0.9. The absolute deviation in the predicted retention time for each component by LCTRS was lower than that of the relative retention time method. The extract rate of the 15 batches of Hedyotis Herba standard decoction ranged from 7.89% to 14.60%， the contents of deacetyl asperulosidic acid， deacetyl asperulosidic acid methyl ester， asperulosidic acid and p-coumaric acid were 6.62-19.70， 3.83-17.99， 1.57-6.69， 1.62-4.52 mg·g^-1， and the transfer rates of these components from decoction pieces to the standard decoction were 22.89%-39.60%， 34.03%-62.24%， 24.25%-43.70%， and 40.58%-73.71%， respectively. The extract rate， index component contents and transfer rates from decoction pieces to the three batches of Hedyotis Herba dispensing granules（P1-P3）， produced by manufacturer A， were similar to those of the standard decoction prepared from the same batch of decoction pieces， and all fell within the specified range. The contents of the 4 indicator components in 7 batches of the dispensing granules（P4-P10） from manufacturers B-E were all within the range of the content converted from the standard decoction based on the quantity of the dispensing granules. ConclusionThe established specific chromatogram and QAMS based on LCTRS are reasonable and reliable. Based on the evaluation indicators of standard decoction yield， consistency of specific chromatograms， contents and transfer rates of the four index components， the 10 batches of Hedyotis Herba dispensing granules from various manufacturers have exhibited good consistency with the standard decoction， indicating that the current production process is relatively reasonable.

ObjectiveTo establish the specific chromatogram of Chuanxiong Rhizoma dispensing granules（CRdg）， and to evaluate its quality by chemometrics and two reference substances for determination of multiple components（TRSDMC）. MethodsHigh performance liquid chromatography（HPLC） specific chromatograms were established using 13 batches of CRdg from 7 manufacturers， and preliminary quality evaluation was performed by similarity evaluation and chemometrics analysis. Eight characteristic peaks in the specific chromatogram of CRdg were measured on 22 different types of C₁₈ columns， and the actual retention times were recorded. Taking chlorogenic acid（peak 1） and senkyunolide A（peak 8） as double standard compounds， the retention times of the eight characteristic peaks were predicted by linear calibration using two reference substances（LCTRS）， and the method was validated on three other columns of different brands. Taking chlorogenic acid as reference peak， the relative correction factor method（RCFM） was used to quantify cryptochlorogenic acid， caffeic acid， ferulic acid， senkyunolide I and senkyunolide A， and the results were compared with the external standard method（ESM）. ResultsThe similarities of specific chromatograms of 13 batches of CRdg were all >0.90， and a total of 8 characteristic peaks were calibrated， and six of them were identified， including chlorogenic acid（peak 1）， cryptochlorogenic acid（peak 2）， caffeic acid（peak 3）， ferulic acid（peak 5）， senkyunolide I（peak 6） and senkyunolide A（peak 8）. Through chemometric analysis， it was found that ferulic acid， chlorogenic acid， senkyunolide I and cryptochlorogenic acid were the main components causing quality difference in CRdg， and the accuracy of LCTRS in predicting the retention time of 8 characteristic peaks was superior to that of the relative retention time method（RRT）. Further comparison of the results obtained from RCFM and ESM showed that there was no statistically significant difference between the two methods. ConclusionA quality evaluation method for CRdg based on HPLC specific chromatogram and TRSDMC is established， its qualitative accuracy is better than that of RRT， the quantitative accuracy is similar to that of ESM， and 4 quality-differentiated components among different manufacturers are found. This method is stable and reliable， and has reference value for the quality evaluation of other dispensing granules.

ObjectiveTo establish the specific chromatogram of Chuanxiong Rhizoma dispensing granules（CRdg）， and to evaluate its quality by chemometrics and two reference substances for determination of multiple components（TRSDMC）. MethodsHigh performance liquid chromatography（HPLC） specific chromatograms were established using 13 batches of CRdg from 7 manufacturers， and preliminary quality evaluation was performed by similarity evaluation and chemometrics analysis. Eight characteristic peaks in the specific chromatogram of CRdg were measured on 22 different types of C₁₈ columns， and the actual retention times were recorded. Taking chlorogenic acid（peak 1） and senkyunolide A（peak 8） as double standard compounds， the retention times of the eight characteristic peaks were predicted by linear calibration using two reference substances（LCTRS）， and the method was validated on three other columns of different brands. Taking chlorogenic acid as reference peak， the relative correction factor method（RCFM） was used to quantify cryptochlorogenic acid， caffeic acid， ferulic acid， senkyunolide I and senkyunolide A， and the results were compared with the external standard method（ESM）. ResultsThe similarities of specific chromatograms of 13 batches of CRdg were all >0.90， and a total of 8 characteristic peaks were calibrated， and six of them were identified， including chlorogenic acid（peak 1）， cryptochlorogenic acid（peak 2）， caffeic acid（peak 3）， ferulic acid（peak 5）， senkyunolide I（peak 6） and senkyunolide A（peak 8）. Through chemometric analysis， it was found that ferulic acid， chlorogenic acid， senkyunolide I and cryptochlorogenic acid were the main components causing quality difference in CRdg， and the accuracy of LCTRS in predicting the retention time of 8 characteristic peaks was superior to that of the relative retention time method（RRT）. Further comparison of the results obtained from RCFM and ESM showed that there was no statistically significant difference between the two methods. ConclusionA quality evaluation method for CRdg based on HPLC specific chromatogram and TRSDMC is established， its qualitative accuracy is better than that of RRT， the quantitative accuracy is similar to that of ESM， and 4 quality-differentiated components among different manufacturers are found. This method is stable and reliable， and has reference value for the quality evaluation of other dispensing granules.

High-quality standard drafting is the foundation for ensuring the effective implementation of radiological health standards. Based on the current standard drafting requirements and the practices in drafting and reviewing radiological health standards in recent years, this article analyzed and discussed the connotations of three principles of standardized document expression, including consistency, coordination, and usability, as well as their application in standard drafting. This article summarized the important considerations during drafting and provided reasonable suggestions, which can be used as a reference for standardization professionals.

ObjectiveTo establish the quality standards for Sennae Folium（Cassia angustifolia） dispensing granules based on standard decoctions. MethodsHigh performance liquid chromatography（HPLC） specific chromatograms were established for 15 batches of Sennae Folium（C. angustifolia） standard decoctions and 10 of Sennae Folium（C. angustifolia） dispensing granules from different manufacturers， and the similarity evaluation， hierarchical cluster analysis（HCA） and principal component analysis（PCA） were performed. Linear calibration with two reference substances（LCTRS） and quantitative analysis of multi-components by single-marker（QAMS） were established for the common peaks in the specific chromatograms to determine the contents of main components in the decoction pieces， standard decoctions and dispensing granules， and to calculate their transfer rates from decoction pieces to standard decoctions and dispensing granules. ResultsThe similarities of specific chromatograms of 15 batches of Sennae Folium（C. angustifolia） standard decoctions and 10 batches of Sennae Folium（C. angustifolia） dispensing granules were all greater than 0.95， and a total of 8 characteristic peaks were calibrated， and five of them were identified， including kaempferol-3，7-O-diglucoside， apigenin-6，8-di-C-glucoside， quercetin-3-O-gentianoside， sennoside B and sennoside A. HCA and PCA results showed that there were certain differences in the composition of different batches of standard decoctions， but no clustering was observed in the production area. As the standard decoctions， the extract rate of 15 batches of samples was 26.54%-45.38%， the contents of kaempferol-3，7-O-diglucoside， apigenin-6，8-di-C-glucoside， quercetin-3-O-gentianoside， sennoside B and sennoside A were 12.16-19.26， 2.57-4.94， 3.27-5.11， 6.75-11.39， 4.69-7.79 mg·g^-1， and their transfer rates from decoction pieces to standard decoctions were 45.41%-79.02%， 29.12%-55.07%， 40.52%-67.90%， 24.72%-49.12%， 27.54%-49.34%， respectively. The extract rates of Sennae Folium（C. angustifolia） dispensing granules（C8-C10） were 38.10%-39.50%， the transfer rates of the above five components from decoction pieces to dispensing granules were 72.85%-73.58%， 53.43%-53.94%， 40.19%-40.74%， 24.62%-25.00%， 28.65%-29.11%， respectively， which were generally consistent with the transfer rates from decoction pieces to standard decoctions. ConclusionCompared with the relative retention time method， LCTRS has higher prediction accuracy and is more suitable for chromatographic columns. The established quality control standard of Sennae Folium（C. angustifolia） dispensing granules based on standard decoction is reasonable and reliable， and all indicators of samples from different manufacturers are within the range specified based on the standard decoction， which can provide reference for the quality control and process research of this dispensing granules.

ObjectiveTo develop a nomogram-based risk prediction model for chronic obstructive pulmonary disease (COPD) incidence among the community-dwelling population aged 40 years old and above, so as to provide targeted references for the screening and prevention of COPD. MethodsBased on a natural population cohort in suburban Shanghai, a total of 3 381 randomly selected participants aged ≥40 years underwent pulmonary function tests between July and October 2021. Cox stepwise regression analysis was used to develop overall and gender-specific risk prediction models, along with the construction of corresponding risk nomograms. Model predictive performance was evaluated using the C-indice, area under the curve (AUC) values, and Brier score. Stability was assessed through 10-fold cross-validation and sensitivity analysis. ResultsA total of 3 019 participants were included, with a median follow-up duration of 4.6 years. The COPD incidence density was 17.22 per 1 000 person-years, significantly higher in males (32.04/1 000 person-years) than that in females (7.38/1 000 person-years) (P<0.001). The overall risk prediction model included the variables such as gender, age, education level, BMI, smoking, passive smoking, and respiratory comorbidities. The male-specific model incorporated the variables such as age, BMI, respiratory comorbidities, and smoking, while the female-specific model included age, marital status, respiratory comorbidities, and pulmonary tuberculosis history. The C-indices for the overall, male-specific, and female-specific models were 0.829, 0.749, and 0.807, respectively. The 5-year AUC values were 0.785, 0.658, and 0.811, with Brier scores of 0.103, 0.176, and 0.059, respectively. Both 10-fold cross-validated C-indices and sensitivity analysis (excluding participants with a follow-up duration of <6 months) yielded C-indices were above 0.740. ConclusionThis study developed concise and practical overall and gender-specific COPD risk prediction models and corresponding nomograms. The models demonstrated robust performance in predicting COPD incidence, providing a valuable reference for identifying high-risk populations and formulating targeted screening and personalized management strategies.

Background::Heterotaxy (HTX) is a thoracoabdominal organ anomaly syndrome and commonly accompanied by congenital heart disease (CHD). The aim of this study was to analyze rare copy number variations (CNVs) in a HTX/CHD cohort and to examine the potential mechanisms contributing to HTX/CHD.Methods::Chromosome microarray analysis was used to identify rare CNVs in a cohort of 120 unrelated HTX/CHD patients, and available samples from parents were used to confirm the inheritance pattern. Potential candidate genes in CNVs region were prioritized via the DECIPHER database, and PNPLA4 was identified as the leading candidate gene. To validate, we generated PNPLA4-overexpressing human induced pluripotent stem cell lines as well as pnpla4-overexpressing zebrafish model, followed by a series of transcriptomic, biochemical and cellular analyses. Results::Seventeen rare CNVs were identified in 15 of the 120 HTX/CHD patients (12.5%). Xp22.31 duplication was one of the inherited CNVs identified in this HTX/CHD cohort, and PNPLA4 in the Xp22.31 was a candidate gene associated with HTX/CHD. PNPLA4 is expressed in the lateral plate mesoderm, which is known to be critical for left/right embryonic patterning as well as cardiomyocyte differentiation, and in the neural crest cell lineage. Through a series of in vivo and in vitro analyses at the molecular and cellular levels, we revealed that the biological function of PNPLA4 is importantly involved in the primary cilia formation and function via its regulation of energy metabolism and mitochondria-mediated ATP production. Conclusions::Our findings demonstrated a significant association between CNVs and HTX/CHD. Our data strongly suggested that an increased genetic dose of PNPLA4 due to Xp22.31 duplication is a disease-causing risk factor for HTX/CHD.

Objective To summarize the diagnosis and treatment experience of surgical site infection(SSI)after percutaneous endoscopic lumbar discectomy(PELD).Methods A retrospective analysis was made on 11 cases of postoperative SSI after PELD from January 2016 to December 2022.After PELD surgery,severe lower back or lower limb pain occurred in all the patients,with a Visual Analog Scale(VAS)score of 7-9(average,8.1).The erythrocyte sedimentation rate(ESR)was 17-114 mm/h(average,54.4 mm/h),and the level of C-reactive protein(CRP)was 8-151 mg/L(average,56.5 mg/L).The MRI showed changes of inflammatory signals in the intervertebral space of the surgical area.The SSI was diagnosed at 2-17 d(average,9.5 d)after PELD surgery.All the cases were treated with antibiotics(4 cases with empiric antibiotic therapy and 7 cases with sensitive antibiotic treatment),8 of which underwent combined surgery,including percutaneous endoscopic lesion removal,catheter irrigation and drainage in 4 cases,posterior lesion removal in 2 cases,and posterior lesion removal combined with internal fixation in 2 cases.Results The patients were followed up for 14-75 months(average,36.8 months).According to the modified MacNab criteria,5 cases were excellent,4 cases were good,and 2 cases were fair.Conclusions SSI after PELD is characterized by acute onset,recurrent severe lower back or lower limb pain,elevated inflammatory markers,and characteristic imaging features.Standardized antibiotic treatment combined with surgery can achieve good therapeutic effects.

Objective We systematically analyze the changes in the count and function of platelet at the early sepsis based on clinical study and single cell sequencing.Methods Clinical data of sepsis patients at the early stage were collected and had been compared between different prognostic groups in the prospective case-control study.The independent risk factors of death were analyzed by logistic regression,and the predictive efficacy of clini-cal indicators was evaluated by receiver operating characteristic(ROC)curve.The healthy volunteers and sepsis patients were recruited.Clinical researchers collected peripheral venous blood samples for sorting cell samples to carry out single-cell RNA sequencing(sc-RNA seq).Through bioinformatics techniques,we analyzed the changes in platelet count,the significantly differential-expressed genes and its enriched functional signaling pathways in the early stages of sepsis.Results(1)A total of 224 patients were enrolled,with a 90 day survival rate of 70.5%.Compared with the survival group,the count of platelet and MAP in the death group at the early stage of sepsis were significantly lower,but the plasma lactate content and SOFA score were significantly higher.(2)Based on single cell sequencing technology,cells are annotated as six groups.The proportion of innate immune cells(neutrophils,monocytes,and dendritic cells)was significantly increased in the early stage of sepsis compared to the healthy volun-teers(2.15∶1),while platelets significantly decreased(0.31∶1).(3)Through bioinformatics technology,CD41/CD42a/CD61 was identified as platelet specific molecules,with significantly increased expression levels in sepsis.Three molecules can distinguish platelets together.(4)771 genes were significantly upregulated and 1101 genes were significantly downregulated in platelets of patients with sepsis,including core molecules involved in physiological functions such as cell adhesion,chemotaxis,and immune response.Functional analysis suggests that differentially expressed genes are enriched in coagulation,immune functions and cell death,participating in oxidative phosphory-lation,leukocyte chemotaxis,iron death,and NOD like receptor signaling pathways.Conclusion Reduced platelet count is associated with poor prognosis in the early stage of sepsis.The specific high expression molecules CD41/CD42a/CD61 that are significantly upregulated in platelets can serve as biomarkers for platelets.Platelets not only mediate cell adhesion and coagulation cascade,but also participate in functional changes such as immune cell chemotaxis,inflammatory response,and the pathological death of inflammatory cells.

Objective To construct a recombinant poxvirus vector vaccine, rVTTδTK-RBD, and to evaluate its safety and immunogenicity. Methods The receptor-binding domain (RBD) gene was synthesized with reference to the gene sequence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and was inserted into the polyclonal site of the self-constructed recombinant plasmid pSTKE, to construct the recombinant poxvirus shuttle vector pSTKE-RBD. This was then transfected into BHK-21 cells pre-infected with the vaccinia virus Tiantan strain (VTT). The recombinant poxvirus rVTTδTK-RBD was successfully obtained after several rounds of fluorescence phage screening. The effect of rVTTδTK-RBD on the body mass of BALB/c mice was detected after immunizing mice by intra-nasal vaccination. The levels of specific and neutralizing antibodies produced by rVTTδTK-RBD on BALB/c mice were analyzed after immunizing mice intramuscularly. The effect of rVTTδTK-RBD on T cell subsets in BALB/c mice was detected by flow cytometry. Results Through homologous recombination, enhanced green fluorescent protein (EGFP) screening marker, and multiple rounds of fluorescent phosphorescence phage screening, a recombinant poxvirus rVTTδTK-RBD, expressing RBD with deletions in the thymidine kinase (TK) gene, was successfully obtained, which was validated by PCR. The in vivo experiments on BALB/c mice showed that rVTTδTK-RBD was highly immunogenic against SARS-CoV-2 and significantly reduced toxicity to the body compared to the parental strain VTT. Conclusion The recombinant poxvirus vaccine rVTTδTK-RBD against SARS-CoV-2 is successfully constructed and obtained, with its safety and immunogenicity confirmed through various experiments.
Animals ; Mice ; SARS-CoV-2/genetics* ; COVID-19 ; Vaccines, Synthetic/genetics* ; Genes, Reporter ; Bacteriophages ; Mice, Inbred BALB C