Objective To explore the protective mechanism of icariin on neuronal oxidative damage,providing a basic pharmacological basis for the treatment of cognitive impairment.Methods Glutamate was used to induce oxidative stress injury in HT22 cells.HT22 cells were divided into control group(normal cultured cells),model group(glutamate injury model)and experimental-L,-M,-H groups(5,10 and 20 μmol·L-1 icariin pretreatment for modeling,respectively).Cell proliferation was detected by cell counting kit-8(CCK-8)method;cytotoxicity was detected by lactate dehydrogenase(LDH)method;reactive oxygen species(ROS)levels were detected by flow cytometry;superoxide dismutase(SOD)levels were detected by biochemical kits;the expression levels of Kelch-like epichlorohydrin-related protein-1(Keap1),nuclear factor E2-related factor 2(Nrf2)were detected by Western blotting;the corresponding mRNA expression was detected by real-time fluorescence quantification polymerose chain reaction.Results The cell viability of control group,model group and experimental-L,-M,-H groups were(100.00±1.31)％,(66.38±2.44)％,(72.07±4.95)％,(82.41±3.57)％and(87.97±4.98)％;LDH release were(0.48±0.52)％,(18.82±2.09)％,(15.32±1.17)％,(10.37±1.39)％and(6.51±0.87)％;ROS level were(14.23±1.13)％,(41.74±1.60)％,(35.69±1.08)％,(33.28±1.69)％and(30.32±2.03)％;SOD levels were(54.84±1.17),(37.95±1.13),(48.02±1.28),(50.56±1.34)and(52.55±1.04)U·mg-1;Keap1 protein levels were 0.36±0.01,0.52±0.03,0.46±0.04,0.39±0.09 and 0.35±0.12;Nrf2 protein levels were 0.29±0.02,0.13±0.08,0.18±0.03,0.21±0.11 and 0.26±0.04;catalase(CAT)mRNA levels were 1.01±0.08,0.81±0.06,0.90±0.04,1.05±0.15 and 1.33±0.26;SOD mRNA levels were 1.09±0.12,0.83±0.03,0.86±0.08,0.94±0.08 and 1.09±0.16.Among the above indicators,the differences between the model group and the control group were statistically significant(all P＜0.01);the differences between the experimental-M,-H groups and the model group were statistically significant(P＜0.01,P＜0.05).Conclusion Icariin may activate the Keap1/Nrf2/antioxidant response element(ARE)signaling pathway,regulate the expression of related proteins,and reduce the level of ROS to effectively alleviate oxidative stress injury in neuronal cells.

Vesicular glutamate transporter 2(VGlut2)is expressed in the PVN of the hypothalamic paraventricular nucleus(PVNVG1ut2)as an excitatory neurotransmitter,which regulates food intake and energy metabolism and plays an important role in maintaining homeostasis.However,it is not clear that the upstream and downstream projection network of PVNVGut2 neurons hinders the anal-ysis of glutamatergic neuron circuit function.Anterograde and retrograde tracer viruses were injec-ted into the PVN of VGlut2 mice by stereotactic brain injection technique to find the input and output nuclei of PVNVGlut2 neurons.Anterograde tracing results showed that PVNVGlut2 neurons pro-jected to the downstream medial amygdala(MeAD)and arcuate nucleus(ARC).Retrograde trac-ing results showed that PVNVGlut2 received input from the prefrontal nucleus(Pr),the reticular tegmental nucleus(RtTg),and the hypoglossal nucleus(12N).In addition,VGlut2 was found to be co-expressed with neuronal nitric oxide synthase(nNOS)neurons in the PVN.The anatomical net-work of PVNVG1ut2 neurons was analyzed by virus tracking tool,which laid the anatomical founda-tion for further study on the functional regulation of PVNVGlut2.

Objective:To investigate the molecular mechanism and distribution characteristics of RhD negative phenotypes in Han population of blood donors in Wuhu city.Methods:A total of 210 RhD-samples from August 2021 to August 2022 were screened by serological test and collected from Wuhu Central Blood Station for the voluntary blood donor population.Exons 1 and 10 of the RHD gene were amplificated by PCR to determine whether the samples had the RHD gene.Exons 1-10 of the RHD gene were amplificated by PCR and zygosity analysis were performed in 82 samples containing D gene,and Sanger sequencing was performed on 55 samples containing all RHD exons to determine the genotype.Results:Among 210 RhD-specimens,128 cases(60.38％)had RHD gene deletion.27 cases had partial exons of RHD,including 2 cases with RHD*DVI.3/RHD*01N.01,24 cases with RHD*01N.04/RHD*01N.01,and 1 case with RHD-CE(2-10)/RHD*01N.01.55 cases had retained all of 10 exons,including 4 cases with RHD*01/RHD*01N.01,6 cases with RHD*15/RHD*01N.01,1 case with RHD*01W.72/RHD*01N.01,1 case with RHD*15/RHD*01EL.01,39 cases with RHD*01EL.01/RHD*01N.01,and the remaining 4 cases were determined to have no RHD gene deletion by zygosity analysis and sequencing showed the presence of 1227G＞A mutation loci.Conclusion:There is polymorphism in the molecular mechanism of RhD-D gene in Wuhu blood donor population,among which RHD*01EL.01 and RHD*15 are the main variants in this region.The results of this study provide a theoretical basis for RhD blood group identification and clinical blood transfusion in this region.

Aim To study the regulation and mechanism of phloroglucinol in bladder smooth muscle spasm. Methods In vitro the experiment used bladder muscle strip to verify the relieving effect of phloro-glucinol on bladder spasm by different drugs. At the same time,RT-qPCR and Western blot were used to detect the expression levels of genes involved in the calcium signaling pathway caused by the antispasmodic effect of phloroglucinol. Results Phloroglucinol could relieve bladder spasm, and the antispasmodic effect was enhanced with the increase of concentration, and the expression of calponin 1 and MYLK3 in tissue cells increased. The results of RT-qPCR showed that the expression of Gprc5b G,Ppp2r5a, Chptl, Prkar2b ,Abcd2 and Rasdl genes in mouse bladder tissue significantly decreased, which was consistent with the sequencing results of RNA-seq.Conclusions Phloroglucinol can relieve bladder smooth muscle spasm, and its mechanism is related to calcium signaling pathway. Meanwhile, phloroglucinol also inhibits the expression of Rasdl gene, suggesting that it may be related to cell cycle , protein phosphorylation, choline metabolism, ATP synthesis and tumor-related pathways.

Since the anatomical location of acupoints was recorded in The latest Practice of Western Acupuncture in 1915, and Lecture Notes on Advanced Acupuncture in 1931, the Japanese acupuncture works of Chinese translation version, the location of Dazhui (GV 14) (under the spinous process of the 7th cervical vertebra) and Yaoyangguan (GV 3) (under the spinous process of the 4th lumbar vertebra) had rarely been questioned for nearly a century. In order to confirm the above statement, the writers have reviewed ancient literature, combined with the modern anatomical knowledge and searched the evidences from the core arguments of the acupuncture Mingtang chart and the bronze acupuncture statue. It is believed that Dazhui （GV 14） should be positioned under the spinous process of the 1st thoracic vertebra, and Yaoyangguan（GV 3） be under the spinous process of the 5th lumbar vertebra. Accordingly, all of the other acupoints of these meridians should be moved down by 1 vertebra, i.e. those on the governor vessel from Dazhui (GV 14) to Yaoyangguan (GV 3), those on the 1st lateral line of the bladder meridian of foot-taiyang from Dazhu (BL 11) to Baihuanshu (BL 30) and those on the 2nd lateral line of the bladder meridian from Fufen (BL 41) to Zhibian (BL 54).
Acupuncture Therapy/history* ; Meridians ; Acupuncture Points ; Lumbar Vertebrae ; Thoracic Vertebrae

BACKGROUND: Patients with atrial fibrillation (AF) and prior stroke history have a high risk of cardiovascular events despite anticoagulation therapy. It is unclear whether catheter ablation (CA) has further benefits in these patients. METHODS: AF patients with a previous history of stroke or systemic embolism (SE) from the prospective Chinese Atrial Fibrillation Registry study between August 2011 and December 2020 were included in the analysis. Patients were matched in a 1:1 ratio to CA or medical treatment (MT) based on propensity score. The primary outcome was a composite of all-cause death or ischemic stroke (IS)/SE. RESULTS: During a total of 4.1 ± 2.3 years of follow-up, the primary outcome occurred in 111 patients in the CA group (3.3 per 100 person-years) and in 229 patients in the MT group (5.7 per 100 person-years). The CA group had a lower risk of the primary outcome compared to the MT group [hazard ratio (HR) = 0.59, 95% CI: 0.47-0.74, P < 0.001]. There was a significant decreasing risk of all-cause mortality (HR = 0.43, 95% CI: 0.31-0.61, P < 0.001), IS/SE (HR = 0.73, 95% CI: 0.54-0.97, P = 0.033), cardiovascular mortality (HR = 0.32, 95% CI: 0.19-0.54, P < 0.001) and AF recurrence (HR = 0.33, 95% CI: 0.30-0.37, P < 0.001) in the CA group compared to that in the MT group. Sensitivity analysis generated consistent results when adjusting for time-dependent usage of anticoagulants. CONCLUSIONS In AF patients with a prior stroke history, CA was associated with a lower combined risk of all-cause death or IS/SE. Further clinical trials are warranted to confirm the benefits of CA in these patients.

Polycystic ovary syndrome (PCOS) is a common disease caused by complex endocrine and metabolic abnormalities in women of childbearing age. Metformin is the most widely used oral hypoglycemic drug in clinic. In recent years, metformin has been used in the treatment of PCOS, but its mechanism is not clear. In this study, we aimed to investigate the effect of metformin on PCOS and its mechanism through PCOS mouse model. Female C57BL/6J mice aged 4-5 weeks were intragastrically given letrozole (1 mg/kg daily) combined with a high-fat diet (HFD) for 21 days to establish the PCOS model. After modeling, metformin (200 mg/kg daily) was intragastrically administered. One month later, the body weight and oral glucose tolerance test (OGTT) were measured. Hematoxylin eosin (H&E) staining was used to detect the pathological changes of ovary. The serum levels of anti-Mullerian hormone (AMH), follicle-stimulating hormone (FSH), luteinizing hormone (LH), E₂ and testosterone (T) were measured by ELISA. The expression of DDX4/MVH was detected by immunohistochemistry. DDX4/MVH and PCNA were co-labeled by immunofluorescence. The protein levels of DDX4/MVH, PCNA, cyclin D2, AMPK and mTOR were detected by Western blot. The results showed that after metformin treatment, the body weights of PCOS mice were gradually returned to normal, glucose tolerance was significantly improved, serum E₂ levels were increased, while AMH, LH, T levels and LH/FSH ratio were decreased. Ovarian polycystic lesions were reduced with reduced atresia follicles. Furthermore, the number of proliferative female germline stem cells (FGSCs) and levels of proliferation related proteins (PCNA, cyclin D2) were significantly increased, and the p-mTOR and p-AMPK levels were markedly up-regulated. These results suggest that metformin treatment not only improves hyperandrogenemia, glucose intolerance and polycystic ovarian lesions in PCOS, but also activates the function of FGSCs. The underlying mechanism may be related to the phosphorylation of AMPK and mTOR. These findings provide new evidence to use metformin in the treatment of PCOS and follicular development disorder.
AMP-Activated Protein Kinases ; Animals ; Cyclin D2 ; Female ; Follicle Stimulating Hormone/therapeutic use* ; Humans ; Luteinizing Hormone/therapeutic use* ; Metformin/pharmacology* ; Mice ; Mice, Inbred C57BL ; Oogonial Stem Cells/metabolism* ; Ovarian Cysts/drug therapy* ; Ovarian Neoplasms ; Polycystic Ovary Syndrome/drug therapy* ; Proliferating Cell Nuclear Antigen/therapeutic use* ; TOR Serine-Threonine Kinases

ObjectiveTo study the in vitro kinetics of Jiaojiang cataplasms and evaluate its pharmacodynamics， so as to provide a feasible basis for the development of this preparation. MethodThe improved Franz diffusion cell was used for the in vitro release in semipermeable membrane and transdermal absorption in in vitro mouse skins. The contents of hydroxy-α-sanshool， 6-gingerol， ginsenoside Rb₁ were determined by high performance liquid chromatography （HPLC）， to evaluate the in vitro release and transdermal absorption of Jiaojiang cataplasms. The mobile phase of 6-gingerol and hydroxy-α-sanshool was water-acetonitrile-methanol （2∶1∶1） with the detection wavelength of 280 nm. The mobile phase of ginsenoside Rb₁ was acetonitrile-0.1% phosphoric acid aqueous solution （31∶69） with the detection wavelength of 203 nm. A mouse intestinal paralysis model was established， and mice were randomly divided into five groups， namely sham operation group， model group， domperidone group （3.9 mg·kg^-1） and high- and low-dose groups of Jiaojiang cataplasms （6.2， 3.1 g·kg^-1， measured by crude drug dosage）， to observe the effect of this preparation on gastrointestinal propulsion function. ResultAverage release rates of hydroxy-α-sanshool， 6-gingerol and ginsenoside Rb₁ at 24 h were 16.41， 4.23， 4.15 μg∙cm^-2∙h^-1， the average transdermal rates of them at 24 h were 2.31， 0.64， 0.29 μg∙cm^-2∙h^-1， their skin retention values were 19.56， 3.59， 1.61 μg， respectively. According to the Ritger-Peppas equation， the release of hydroxy-α-sanshool， 6-gingerol， ginsenoside Rb₁ was non-Fick diffusion. The high-dose group of Jiaojiang cataplasms could improve intestinal function of model mice after small intestinal friction injury， and promote intestinal peristalsis and small intestinal propulsion rate （P<0.05）. ConclusionJiaojiang cataplasms has in vitro release and transdermal properties， the in vitro release conforms to Higuchi equation， and transdermal absorption behavior conforms to zero-order kinetic equation， which can improve the postoperative function of the small intestine and the propulsion function of small intestine. It preliminarily indicates that the preparation has certain clinical development value.

ObjectiveTo explore the mechanism of Cervi Cornu Pantotrichumin in the treatment of osteoarthritis by network pharmacology. MethodThe active ingredients and the corresponding targets of Cervi Cornu Pantotrichumin were screened out by a Bioinformatics Analysis Tool of Molecular mechANism of Traditional Chinese Medicine （BATMAN-TCM）. The targets related to osteoarthritis were obtained through GeneCards and Online Mendelian Inheritance in Man （OMIM）. The targets corresponding to the active ingredients and those related to osteoarthritis were intersected to reveal the common targets， and STRING was adopted to build a protein-protein interaction （PPI） network. DAVID was used for gene ontology （GO） annotation and Kyoto encyclopedia of genes and genomes （KEGG） pathway enrichment on the anti-osteoarthritis targets of Cervi Cornu Pantotrichumin， and R x64 3.6.3 was employed to produce the advanced bubble charts of GO terms and KEGG pathways. Cytoscape 3.7.2 was used to establish the “Chinese medicinal herb-active ingredient-target-signaling pathway” network. In vitro experiments were performed to detect the viability of RAW 264.7 cells exposed to oxidative stress and the tumor necrosis factor （TNF）-α level in RAW 264.7 cells with inflammation under the treatment by Cervi Cornu Pantotrichumin. ResultA total of 20 active ingredients of Cervi Cornu Pantotrichum were obtained， of which ceramide， 6'-O-β-D-glucosylgentiopicroside， cerebroside， oleuropein， sphingomyelin， and cholesterol ferulate did not meet the screening conditions. Therefore， a total of 14 active ingredients were finally screened out， and 303 and 3 093 targets of active ingredients and osteoarthritis were respectively obtained. The two target sets were taken to intersect， which revealed 92 common targets. GO annotation and KEGG pathway enrichment showed that the targets were mainly involved in redox process， positive regulation of RNA polymerase Ⅱ promoter transcription， inflammatory response， protein synthesis， osteoclast differentiation， TNF signaling pathway， signaling pathways in cancer， mammalian target of rapamycin （mTOR） signaling pathway， mitogen-activated protein kinase （MAPK） signaling pathway， and cyclic adenosine monophosphate （cAMP） signaling pathway. The results of in vitro experiments showed that a certain concentration of protein in Cervi Cornu Pantotrichum significantly increased the viability of RAW 264.7 cells exposed to H₂O₂-induced oxidative damage （P<0.05， P<0.01） and reduced the level of TNF-α in the RAW 264.7 cells experiencing lipopolysaccharide （LPS）-induced inflammation （P<0.05）. ConclusionBased on the network pharmacology method， the mechanism of the multi-component， multi-target and multi-pathway treatment of OA by antler antler was explained， and the anti-inflammatory and antioxidant activities of antler antler were confirmed， which provided theoretical guidance and scientific basis for further research on the treatment of OA by antler antler.

Objective: To observe the effect of lipid regulating therapy on carotid atherosclerotic plaque in diabetic patients. Methods: The REACH study, conducted between March 2009 and February 2012, enrolled asymptomatic patients with magnetic resonance imaging (MRI) confirmed carotid atherosclerotic plaque, who had never taken lipid-lowering drugs. Patients were treated with a moderate dose of rosuvastatin for 24 months. Blood lipid levels were measured and carotid MRI was performed at baseline, 3 and 24 months after treatment. The volume of carotid wall and lipid-rich necrotic core (LRNC) were measured by image analysis software. This study retrospectively analyzed patients in the REACH study. Patients were divided into diabetes group and non-diabetic group. The changes of blood lipid level and MRI parameters of carotid atherosclerotic plaque were compared between the two groups and their correlation was analyzed. Results: A total of 38 patients with carotid atherosclerotic plaque were included in this study, including 13 patients (34.2%) in the diabetic group and 25 patients (65.8%) in the non-diabetic group. Baseline parameters were comparable between the two groups, except higher HbA1c level in diabetes group (P<0.05). Compared with baseline, the total cholesterol (TC), low density lipoprotein cholesterol (LDL-C) and triglyceride (TG) levels were significantly decreased at 3 and 24 months in both two groups (P<0.05). The change of high-density lipoprotein cholesterol (HDL-C) in diabetes group was not obvious, while it was significantly increased in non-diabetic group at 24 months ((1.38±0.33) mmol/l vs. (1.26±0.26) mmol/l, P<0.05). MRI results showed that the volume and percentage of LRNC remained unchanged at 3 months, slightly decreased at 24 months (64.86 (45.37, 134.56) mm³ vs. 75.76 (48.20, 115.64) mm³, P>0.05) and (15.84% (11.47%, 24.85%) vs. 16.95% (11.64%, 22.91%), P>0.05) in diabetic group. In non-diabetic group, the volume and percentage of LRNC were significantly decreased at 3 months (63.01 (44.25, 188.64) mm³ vs. 72.49 (51.91, 199.59) mm³, P<0.05) and (13.76% (8.81%, 27.64%) vs. 16.04% (11.18%, 27.05%), P<0.05) respectively. Both parameters further decreased to (55.63 (27.18, 179.40) mm³) and (12.71% (8.39%, 24.41%)) at 24 months (both P<0.05). Wall volume, lumen volume and percent wall volume (PWV) were not affected post therapy in both two groups(P>0.05). There were no correlations between the changes of plaque parameters including volume and percentage of LRNC, wall volume, lumen volume, PWV and the changes of blood lipid parameters (TC, LDL-C, HDL-C and TG) in 3 and 24 months (P>0.05). Conclusion: Lipid-lowering therapy possesses different effects on carotid atherosclerotic plaque in diabetic and non-diabetic patients, and the LRNC improvement is more significant in non-diabetic patients as compared to diabetic patients.
Carotid Arteries/pathology* ; Carotid Artery Diseases/drug therapy* ; Cholesterol, HDL/therapeutic use* ; Cholesterol, LDL ; Diabetes Mellitus ; Humans ; Magnetic Resonance Imaging/methods* ; Necrosis/pathology* ; Plaque, Atherosclerotic/drug therapy* ; Retrospective Studies ; Rosuvastatin Calcium/therapeutic use*