AIM To investigate the effects of Changpu Yujin Decoction(CPYJD)on striatal mitophagy and PINK1/Parkin signaling pathway in a rat model of Tourette syndrome(TS).METHODS Thirty-six SPF male SD rats were randomly assigned to the control group(n=9)and the TS modeling group(n=27).Rats in the modeling group received daily intraperitoneal injections of 3,3'-iminodipropionitrile(IDPN)(300 mg/kg)for 7 consecutive days to establish the TS model.Post-modeling,successfully induced TS rats were re-randomized into model group(no treatment),tiapride group(47.91 mg/kg)and CPYJD group(77.28 g/kg).All groups received their respective interventions via intragastric administration daily for 28 days.Following drug administration,behavioral scores were assessed in each group.Pathological alterations in the striatum were examined using HE staining,while ultrastructural changes were evaluated by transmission electron microscopy(TEM).Neuronal apoptosis was quantified via TUNEL staining,and ROS levels in striatum were measured by ELISA.Co-localization of PINK1 and LC3B was assessed using immunofluorescence(IF).Finally,mRNA and protein expressions of PINK1,Parkin,Beclin-1,P62 and LC3B(LC3B-Ⅱ/Ⅰ ratio)were analyzed by RT-qPCR and Western blot.RESULTS Compared to the control group,the model group demonstrated significantly increased behavioral scores(P＜0.01),elevated neuronal apoptosis rate and higher ROS levels in the striatum(P＜0.01);severe neuronal and mitochondrial damage in the striatum;significantly reduced mRNA and protein expressions of PINK1,Parkin,Beclin-1 and LC3B(LC3B-Ⅱ/Ⅰ ratio)in the striatum(P＜0.01);markedly upregulated P62 mRNA and protein expressions(P＜0.01).Compared to the model group,both the tiapride and CPYJD intervention groups exhibited significantly reduced behavioral scores(P＜0.01);decreased neuronal apoptosis rate and lower ROS levels(P＜0.01);improved pathological alterations in the striatal neurons and mitochondria;increased mRNA and protein expressions of PINK1,Parkin and Beclin-1 in the striatum(P＜0.05,P＜0.01);and decreased P62 mRNA and protein expressions(P＜0.01).Furthermore,the rats in the CPYJD group specifically showed elevated LC3B mRNA level and LC3B-Ⅱ/Ⅰ protein ratio in striatum(P＜0.05,P＜0.01).CONCLUSION The effect of CPYJD intervention in TS rats may involve activation of mitophagy through regulation of the PINK1/Parkin signaling pathway,improving mitochondrial function,reducing ROS levels,and thereby protecting neurons.

Objective To explore the therapeutic effect of cornuside on diabetic retinopathy(DR)in rats and ana-lyze the acting mechanism of the reactive oxygen species(ROS)/thioredoxin interacting protein(TXNIP)/NOD-like recep-tor thermal protein domain associated protein 3(NLRP3)signaling pathway in this process.Methods A total of 56 suc-cessfully modeled DR rats were randomly divided into the model group,the cornuside 20 mg·kg-1 group,the cornuside 40 mg·kg-1 group,the calcium dobesilate 5.8 mg·kg-1 group,with 14 rats in each group;while,meanwhile another 14 healthy rats were selected as the control group.After the corresponding intervention of rats in each group,retinal tissue in-flammation,oxidative stress indicators,fasting blood glucose(FBG),and angiogenic factor levels were detected by the en-zyme-linked immunosorbent assay(ELISA).The hematoxylin-eosin(HE)staining and terminal deoxynucleotidyl transfer-ase dUTP nick end labeling(TUNEL)staining were used to observe retinal histopathology and retinal cell apoptosis,re-spectively.The mRNA expression of ROS,TXNIP,and NLRP3 in the retinal tissue was detected by the quantitative fluores-cence polymerase chain reaction(PCR).The apoptosis of retinal cells and the protein expression of the ROS/TXNIP/NL-RP3 signaling pathway were detected by Western blotting.Results Compared with the control group,the model group showed disordered arrangement of retinal cells and a significant decrease in the cell number,accompanied by nuclear con-densation and edema;interleukin-6(IL-6),tumor necrosis factor-α(TNF-α),malondialdehyde(MDA),FBG,angiopoie-tin-1(Ang-1),vascular endothelial growth factor(VEGF),retinal cell apoptosis rate,ROS,and the mRNA and protein ex-pression levels of TXNIP and NLRP3 increased significantly,while superoxide dismutase(SOD)and B cell lymphoma-2(Bcl-2)protein expression levels decreased significantly(all P＜0.05).Compared with the model group,the arrangement of retinal cells in the cornuside 20 mg·kg-1 group,the cornuside 40 mg·kg-1 group,and the calcium dobesilate 5.8 mg·kg-1 group gradually became normal,the number of retinal cells increased,and the nuclear condensation and edema were relieved;IL-6,TNF-α,MDA,FBG,Ang-1,VEGF,retinal cell apoptosis rate,ROS,and the mRNA and protein expression level of TXNIP and NLRP3 decreased significantly,while the protein expression level of SOD and Bcl-2 increased signifi-cantly(all P＜0.05).In the intergroup comparison of the pathological damage of retinal tissue and the improvement degree of the above quantitative indexes in DR rats,the cornuside 40 mg·kg-1group was superior to the cornusin 20 mg·kg-1 group(all P＜0.05);the calcium dobesilate 5.8 mg·kg-1 group was superior to the cornusin 20 mg·kg-1 group,but infe-rior to the cornuside 40 mg·kg-1 group(all P＜0.05).Conclusion Cornuside can mitigate retinal inflammation,oxi-dative stress,and pathological damage in DR rats and inhibit blood glucose,retinal angiogenesis,and cell apoptosis.The acting mechanism of cornuside may be related to the inhibition of the ROS/TXNIP/NLRP3 signaling pathway.

Objective To explore the therapeutic effect of cornuside on diabetic retinopathy(DR)in rats and ana-lyze the acting mechanism of the reactive oxygen species(ROS)/thioredoxin interacting protein(TXNIP)/NOD-like recep-tor thermal protein domain associated protein 3(NLRP3)signaling pathway in this process.Methods A total of 56 suc-cessfully modeled DR rats were randomly divided into the model group,the cornuside 20 mg·kg-1 group,the cornuside 40 mg·kg-1 group,the calcium dobesilate 5.8 mg·kg-1 group,with 14 rats in each group;while,meanwhile another 14 healthy rats were selected as the control group.After the corresponding intervention of rats in each group,retinal tissue in-flammation,oxidative stress indicators,fasting blood glucose(FBG),and angiogenic factor levels were detected by the en-zyme-linked immunosorbent assay(ELISA).The hematoxylin-eosin(HE)staining and terminal deoxynucleotidyl transfer-ase dUTP nick end labeling(TUNEL)staining were used to observe retinal histopathology and retinal cell apoptosis,re-spectively.The mRNA expression of ROS,TXNIP,and NLRP3 in the retinal tissue was detected by the quantitative fluores-cence polymerase chain reaction(PCR).The apoptosis of retinal cells and the protein expression of the ROS/TXNIP/NL-RP3 signaling pathway were detected by Western blotting.Results Compared with the control group,the model group showed disordered arrangement of retinal cells and a significant decrease in the cell number,accompanied by nuclear con-densation and edema;interleukin-6(IL-6),tumor necrosis factor-α(TNF-α),malondialdehyde(MDA),FBG,angiopoie-tin-1(Ang-1),vascular endothelial growth factor(VEGF),retinal cell apoptosis rate,ROS,and the mRNA and protein ex-pression levels of TXNIP and NLRP3 increased significantly,while superoxide dismutase(SOD)and B cell lymphoma-2(Bcl-2)protein expression levels decreased significantly(all P＜0.05).Compared with the model group,the arrangement of retinal cells in the cornuside 20 mg·kg-1 group,the cornuside 40 mg·kg-1 group,and the calcium dobesilate 5.8 mg·kg-1 group gradually became normal,the number of retinal cells increased,and the nuclear condensation and edema were relieved;IL-6,TNF-α,MDA,FBG,Ang-1,VEGF,retinal cell apoptosis rate,ROS,and the mRNA and protein expression level of TXNIP and NLRP3 decreased significantly,while the protein expression level of SOD and Bcl-2 increased signifi-cantly(all P＜0.05).In the intergroup comparison of the pathological damage of retinal tissue and the improvement degree of the above quantitative indexes in DR rats,the cornuside 40 mg·kg-1group was superior to the cornusin 20 mg·kg-1 group(all P＜0.05);the calcium dobesilate 5.8 mg·kg-1 group was superior to the cornusin 20 mg·kg-1 group,but infe-rior to the cornuside 40 mg·kg-1 group(all P＜0.05).Conclusion Cornuside can mitigate retinal inflammation,oxi-dative stress,and pathological damage in DR rats and inhibit blood glucose,retinal angiogenesis,and cell apoptosis.The acting mechanism of cornuside may be related to the inhibition of the ROS/TXNIP/NLRP3 signaling pathway.

Objective:To establish a stable rat model of sequelae of pelvic inflammatory disease(SPID)with clinical characteristics,and to provide a reliable experimental model for the study of the pharmcological effect and mechanism of SPID.Methods:Twenty-four 7-week-old SD rats were divided into sham operation group,model-A(108 cfu/mL mixed bacterial solution,0.2 mL),model-B(109 cfu/mL mixed bacterial solution 0.2 mL),and model-C(108 cfu/mL E.coli 0.2 mL).The weight of the rat's uterine was weighed and the uterine index was calculated.The automatic hematology analyzer was used to detect the blood routine;hematoxylin-eosin staining(HE)and masson staining were used to detect uterine pathlogical changes in rats.Enzyme-linked immunosorbent assay(ELISA)was used to detect interleukin-1β(IL-1β),interleukin-6(IL-6)and tumor necrosis factor-α(TNF-α)in rat uterine tissue homogenates.Western blot was used to detect the expression of proteins related to NF-κB signaling pathway.Results:Compared with the sham operation group,the uterine index of model-A,model-B,and model-C were significantly increased(P＜0.05,P＜0.01).The levels of WBC and NE in the model-A increased significantly(P＜0.01).The level of LY in model-B decreased significantly(P＜0.01).The levels of IL-1β,TNF-α in model-A,model-B,and model-C were significantly increased(P＜0.01).The levels of IL-6 in model-A and model-B were significantly increased(P＜0.05,P＜0.01).The collagen volume fraction of model-A and model-B were significantly increased(P＜0.01).Mechanism study indicates that the expression levels of p-IKKβ/IKKβ,p-IκBα/IκBα and p-p65/p65 in model-A were significantly increased(P＜0.01),and the expression levels of IκBα/β-actin were significantly decreased(P＜0.01).The expression level of p-IKKβ/IKKβ in model-B was significantly increased(P＜0.01).Conclusions:A stable rat model of SPID that conforms to clinical characteristics can be successfully constructed by combining 0.2 mL of mixed bacterial solution with a concentration of 108 cfu/mL and mechanical injury.This modeling method intervened in the expression of the NF-κB inflammatory signaling pathway.

A method was developed for determination of dilauryl thiodipropionate(DLTDP)in fried foods by coupling solid-phase extraction(SPE)pretreatment with reverse-phase liquid chromatography-tandem mass spectrometry(RPLC-MS/MS)detection.Samples were extracted with n-hexane as the solvent,purified using a neutral alumina SPE cartridge,and finally analyzed by RPLC-MS/MS.Quantitative analysis was performed using matrix-matched calibration curves combined with an external standard method under optimal experimental conditions.The results showed that DLTDP exhibited good linearity in the range of 2.0-50.0 μg/L,with a correlation coefficient(R2)≥0.999.The limit of detection(LOD)and the limit of quantification(LOQ)of the method were 0.15 mg/kg and 0.5 mg/kg,respectively.The mean recoveries at three fortification levels(0.5,1.0,and 200 mg/kg)in different samples ranged from 84.8%to 96.8%,with the relative standard deviations(RSDs)all less than 8.0%.The developed method was highly sensitive,accurate and reliable,and easy to operate,making it well suited for the routine quantitative analysis of DLTDP in fried foods.

Objective:To investigate the association between serum 25-hydroxyvitamin D [25(OH)D] levels and 24-h urinary calcium excretion (24-h UCaE) and hypercalciuria in Chinese adults.Methods:This cross-sectional study was conducted from March 2022 to March 2023 in nine cities in China and included 1 239 residents. Demographic characteristics were collected through questionnaires and physical examinations, fasting blood samples were assessed for bone metabolism indicators, and 24-h urine samples were used to determine the 24-h UCaE. Multiple linear regression analysis was used to explore the relationship between serum 25(OH)D and 24-h UCaE and bone metabolism indexes. The relationship between serum 25(OH)D and hypercalciuria was analyzed using a multiple logistic regression model combined with restricted cubic spline modeling.Results:The mean participant age was (47.9±18.1) years, of which 453 (36.6%) were male. The percentages of vitamin D sufficiency, insufficiency, and deficiency were 7.6% (94/1 239), 29.0% (359/1 239), and 63.4% (786/1 239), respectively. The multiple linear regression model showed that after adjusting for the covariates the 24-h UCaE gradually increased with higher levels of 25(OH)D ( P overall <0.001, P nonlinear <0.001). The logistic regression analysis revealed that compared with the vitamin D deficient group, the OR for the prevalence of hypercalciuria in the vitamin D sufficient and vitamin D insufficient groups were 3.290 (95% CI 1.745 to 6.202) and 3.742 (95% CI 2.458 to 5.697), respectively. The results of the restricted cubic spline modeling showed a positive nonlinear relationship between 25(OH)D and the prevalence of hypercalciuria ( P overall <0.001, P nonlinear <0.001). The prevalence of hypercalciuria increased when 25(OH)D was >17.00 μg/L and peaked at 26.71 μg/L, after which there was a decreasing trend in the prevalence of hypercalciuria with increasing 25(OH)D. Conclusion:Associations between serum 25(OH)D levels and urinary calcium excretion and the prevalence of hypercalciuria were observed in the Chinese adult population.

AIM To investigate the effects of Changpu Yujin Decoction(CPYJD)on striatal mitophagy and PINK1/Parkin signaling pathway in a rat model of Tourette syndrome(TS).METHODS Thirty-six SPF male SD rats were randomly assigned to the control group(n=9)and the TS modeling group(n=27).Rats in the modeling group received daily intraperitoneal injections of 3,3'-iminodipropionitrile(IDPN)(300 mg/kg)for 7 consecutive days to establish the TS model.Post-modeling,successfully induced TS rats were re-randomized into model group(no treatment),tiapride group(47.91 mg/kg)and CPYJD group(77.28 g/kg).All groups received their respective interventions via intragastric administration daily for 28 days.Following drug administration,behavioral scores were assessed in each group.Pathological alterations in the striatum were examined using HE staining,while ultrastructural changes were evaluated by transmission electron microscopy(TEM).Neuronal apoptosis was quantified via TUNEL staining,and ROS levels in striatum were measured by ELISA.Co-localization of PINK1 and LC3B was assessed using immunofluorescence(IF).Finally,mRNA and protein expressions of PINK1,Parkin,Beclin-1,P62 and LC3B(LC3B-Ⅱ/Ⅰ ratio)were analyzed by RT-qPCR and Western blot.RESULTS Compared to the control group,the model group demonstrated significantly increased behavioral scores(P＜0.01),elevated neuronal apoptosis rate and higher ROS levels in the striatum(P＜0.01);severe neuronal and mitochondrial damage in the striatum;significantly reduced mRNA and protein expressions of PINK1,Parkin,Beclin-1 and LC3B(LC3B-Ⅱ/Ⅰ ratio)in the striatum(P＜0.01);markedly upregulated P62 mRNA and protein expressions(P＜0.01).Compared to the model group,both the tiapride and CPYJD intervention groups exhibited significantly reduced behavioral scores(P＜0.01);decreased neuronal apoptosis rate and lower ROS levels(P＜0.01);improved pathological alterations in the striatal neurons and mitochondria;increased mRNA and protein expressions of PINK1,Parkin and Beclin-1 in the striatum(P＜0.05,P＜0.01);and decreased P62 mRNA and protein expressions(P＜0.01).Furthermore,the rats in the CPYJD group specifically showed elevated LC3B mRNA level and LC3B-Ⅱ/Ⅰ protein ratio in striatum(P＜0.05,P＜0.01).CONCLUSION The effect of CPYJD intervention in TS rats may involve activation of mitophagy through regulation of the PINK1/Parkin signaling pathway,improving mitochondrial function,reducing ROS levels,and thereby protecting neurons.

OBJECTIVE To improve the quality standard of Zhuang medicine Yingbupu （Aralia armata）. METHODS A total of 23 batches of Yingbupu （A. armata） were studied. Their macroscopic characteristics and powder microscopic features were observed. TLC was employed for the qualitative identification of oleanolic acid and araloside A. Items such as water content， total ash， acid-insoluble ash， and ethanol-soluble extract were determined according to the methods specified in the 2020 edition of the Chinese Pharmacopoeia （part Ⅳ）. UPLC fingerprint was established for 23 batches of samples by using Similarity Evaluation System for Chromatographic Fingerprints of Traditional Chinese Medicine （2012 edition）， and the contents of oleanolic acid and araloside A were determined. RESULTS The powder microscopic characteristics of the medicinal material were distinctive. Oleanolic acid and araloside A were detected by TLC in all 23 batches. Among the 23 batches of samples， the content ranges of moisture， total ash， acid-insoluble ash， and ethanol-soluble extract were 6.9% to 10.4%， 1.8% to 6.8%， 0.1% to 1.9%， and 2.8% to 8.4%， respectively. Based on the UPLC fingerprint， a total of 15 common peaks were obtained， and 9 of these common peaks were identified. The content ranges of oleanolic acid and araloside A in the 23 batches of samples were 0.86% to 2.69% and 0.16% to 1.10%， respectively. CONCLUSIONS This study has added items such as moisture and total ash content fingerprint， TLC identification. A preliminary quality standard has been established for the medicinal material of Yingbupu （A. armata）， stipulating that the moisture content should not exceed 11.0%， the total ash content should not exceed 5.0%， the acid-insoluble ash content should not exceed 2.5%， the ethanol-soluble extract（No. content should not be less than 4.0%， and the contents of zyyzdxk-2023165） oleanolic acid and araloside A should not be less than 1.00% and 0.45%（ calculated by a dried basis）， respectively.

Objective To establish a rapid,sensitive,and specific multiplex PCR detection method for the simultaneous detection of Cryptosporidium,Eimeria,and bovine parvovirus.Methods Specific primers targeting the SSU rRNA genes of Cryptosporidium and Eimeria,as well as the VP2 gene of bovine parvovirus were designed and the corresponding recombinant plasmid standards were constructed.To establish the multiplex PCR method,the reaction conditions were optimized using temperature gradient PCR and single-variable control methods.The sensitivity,specificity,reproducibility,and clinical application of the protocol were evaluated.Results The optimal annealing temperature was found to be 60.5℃,and the forward and reverse primer concentrations were determined to be 0.2 μmol/L for Eimeria,and 0.4 μmol/L for Cryptosporidium and bovine parvovirus.The assay demonstrated high sensitivity,with detection limits of 243,260,and 3 110 copies for the recombinant plasmid standards of Cryptosporidium,Eimeria,and bovine parvovirus,respectively.Specificity testing showed no cross-reactivity with ten common bovine pathogens,including Salmonella,bovine viral diarrhea virus,and bovine rotavirus.Consistent intra-and inter-batch results confirmed the strong reproducibility of the method.Clinical application to 81 diarrhea samples from various regions in the Ganzi Prefecture,Sichuan,revealed positivity rates of 18.52%(15/81)for Cryptosporidium,34.57%(28/81)for Eimeria,and 18.52%(15/81)forbovineparvovirus,withamixedinfectionrateof3.7%(3/81).Conclusions Themultiplex PCR method established in this study offers a reliable tool for differential diagnosis and epidemiological investigation of the three common diarrheal pathogens in yaks.

In-depth study of the components of polymyxins is the key to controlling the quality of this class of antibiotics.Similarities and variations of components present significant analytical challenges.A two-dimensional(2D)liquid chromatography-mass spectrometry(LC-MS)method was established for screening and comprehensive profiling of compositions of the antibiotic colistimethate sodium(CMS).A high concentration of phosphate buffer mobile phase was used in the first-dimensional LC system to get the components well separated.For efficient and high-accuracy screening of CMS,a targeted method based on a self-constructed high resolution(HR)mass spectrum database of CMS components was established.The database was built based on the commercial MassHunter Personal Compound Database and Library(PCDL)software and its accuracy of the compound matching result was verified with six known components before being applied to genuine sample screening.On this basis,the unknown peaks in the CMS chromatograms were deduced and assigned.The molecular formula,group composition,and origins of a total of 99 compounds,of which the combined area percentage accounted for more than 95％of CMS components,were deduced by this 2D-LC-MS method combined with the MassHunter PCDL.This profiling method was highly efficient and could distinguish hundreds of components within 3 h,providing reliable results for quality control of this kind of complex drugs.