BACKGROUND:Ferroptosis is a mode of programmed cell death distinct from apoptosis,necrosis,and other novel cellular deaths,which occurs mainly due to accumulated lipid peroxidation.Ferroptosis has been shown to be involved in the pathological process following subarachnoid hemorrhage.Baicalein,serving as an adept sequestered of iron,evinces its prowess by quelling lipid peroxidative cascades.Nonetheless,the enigma lingers as to whether baicalein possesses the capacity to ameliorate neuronal ferroptosis,elicited in the wake of early brain injury after subarachnoid hemorrhage. OBJECTIVE:To investigate the effect and mechanism of baicalein on neuronal ferroptosis after subarachnoid hemorrhage. METHODS:Primary neuronal cells were extracted from C57BL/6L fetal mice at 16-17 days of gestation.Hemoglobin was used to stimulate primary neuronal cells to simulate an in vitro subarachnoid hemorrhage model.The viability of primary neuronal cells treated with baicalein at concentrations of 5,15,25,50,and 100 μmol/L for 24 hours was detected by CCK-8 assay to determine the optimal concentration of baicalein.Primary neuronal cells were divided into control group,hemoglobin group,and hemoglobin+baicalein group.The levels of reactive oxygen species and malondialdehyde in cells were detected by kits.The mRNA expressions of ferroptosis-related markers PTGS2,SLC7A11,and glutathione peroxidase 4 were detected by RT-PCR.The primary neuronal cells were further divided into control group,SLC7A11 inhibitor Erastin group,hemoglobin group,hemoglobin+baicalein group,and hemoglobin+baicalein+Erastin group.The expression of the ferroptosis related markers SLC7A11 and glutathione peroxidase 4 was detected by western blot assay. RESULTS AND CONCLUSION:(1)Baicalein(25 μmol/L)was selected as the following experimental concentration.(2)Compared with the hemoglobin group,the level of malondialdehyde and the level of reactive oxygen species were significantly decreased(P<0.05)in the hemoglobin+baicalein group.(3)Compared with the hemoglobin group,the mRNA expression of PTGS2 significantly decreased,and the mRNA expression of SLC7A11 and glutathione peroxidase 4 significantly increased(P<0.000 1)in the hemoglobin+baicalein group.(4)SLC7A11 inhibitor Erastin could reverse the baicalin-improved ferroptosis effect to a certain extent(P<0.05).(5)The results showed that baicalein could alleviate the ferroptosis of neuronal cells after subarachnoid hemorrhage through the SLC7A11/GPX4 pathway.

ObjectiveThis study leverages structural data from antigen-antibody complexes of the influenza A virus neuraminidase (NA) protein to investigate the spatial recognition relationship between the antigenic epitopes and antibody paratopes. MethodsStructural data on NA protein antigen-antibody complexes were comprehensively collected from the SAbDab database, and processed to obtain the amino acid sequences and spatial distribution information on antigenic epitopes and corresponding antibody paratopes. Statistical analysis was conducted on the antibody sequences, frequency of use of genes, amino acid preferences, and the lengths of complementarity determining regions (CDR). Epitope hotspots for antibody binding were analyzed, and the spatial structural similarity of antibody paratopes was calculated and subjected to clustering, which allowed for a comprehensively exploration of the spatial recognition relationship between antigenic epitopes and antibodies. The specificity of antibodies targeting different antigenic epitope clusters was further validated through bio-layer interferometry (BLI) experiments. ResultsThe collected data revealed that the antigen-antibody complex structure data of influenza A virus NA protein in SAbDab database were mainly from H3N2, H7N9 and H1N1 subtypes. The hotspot regions of antigen epitopes were primarily located around the catalytic active site. The antibodies used for structural analysis were primarily derived from human and murine sources. Among murine antibodies, the most frequently used V-J gene combination was IGHV1-12*01/IGHJ2*01, while for human antibodies, the most common combination was IGHV1-69*01/IGHJ6*01. There were significant differences in the lengths and usage preferences of heavy chain CDR amino acids between antibodies that bind within the catalytic active site and those that bind to regions outside the catalytic active site. The results revealed that structurally similar antibodies could recognize the same epitopes, indicating a specific spatial recognition between antibody and antigen epitopes. Structural overlap in the binding regions was observed for antibodies with similar paratope structures, and the competitive binding of these antibodies to the epitope was confirmed through BLI experiments. ConclusionThe antigen epitopes of NA protein mainly ditributed around the catalytic active site and its surrounding loops. Spatial complementarity and electrostatic interactions play crucial roles in the recognition and binding of antibodies to antigenic epitopes in the catalytic region. There existed a spatial recognition relationship between antigens and antibodies that was independent of the uniqueness of antibody sequences, which means that antibodies with different sequences could potentially form similar local spatial structures and recognize the same epitopes.

In recent years, polysaccharides have received much attention because of their high safety and good immunological activity. The study of polysaccharide in vivo process is a key scientific problem that needs to be solved for polysaccharide drug development. Some progress has been made in the field of polysaccharide pharmacokinetics and immunomodulation. However, due to the lack of both chromogenic and light-absorbing groups and the complex molecular structure of polysaccharides, the in vivo processes and immunomodulatory mechanisms of polysaccharides have been slow to be investigated. The effective combination of multiple techniques can break the bottleneck of difficult tracing and unknown immunomodulatory mechanism of polysaccharides in vivo, and promote the development and utilization of polysaccharides. In this paper, we systematically summarize the key techniques in the study of polysaccharide in vivo processes and immunomodulatory mechanisms in order to provide technical references and research ideas for the study of polysaccharide in vivo processes and immunomodulatory mechanisms.

"MS/MS spectrum to structure"plays a critical role in the confirmative identification of complicated matrices and is currently regarded as an extremely challenging endeavor.MS/MS information provides vital clues to structural identification.In this study,a strategy was proposed to facilitate unambiguous identification through matching MS3 with MS2 spectra.Initially,MS3 spectra of the featured ions(c-and y-type ions)generated by the decomposition of ester functional group in esters and the MS2 spectrum of the structural unit([M-H]-)were all captured on the Qtrap-MS platform equipped with two tandem-in-space collision cells,including the second quadrupole cell(q2)and linear ion trap(LIT)chambers(actually the third quadrupole unit).Subsequently,the MS/MS spectrum matching between MS3 spectra of the ester compound and MS2 spectra of the structural unit(s)were achieved.As a result,the findings corresponding to MS3 and MS2 spectra matching were summarized.Finally,based on HR-MS/MS information of total salvianolic acid derivatives(TSA),36 kinds of compounds were preliminarily identified through matching with literature information and database retrieval.The applicability of MS3 and MS2 spectra matching strategy was further justified by the confirmative identification of phenolic acid compounds(Rosmarinic acid and salvianolic acid B)in TSA.Above all,MS3 and MS2 spectra matching strategy was quite meaningful towards advancing"MS/MS spectrum to structure"analysis through recognizing and identifying featured fragment ions,and also provided inspiration and new insights for the structural characterization.

Objective To compare the clinical efficacy of anti-vascular endothelial growth factor(VEGF)combined with pars plana vitrectomy(PPV)treatment for patients with proliferative diabetic retinopathy(PDR)of different traditional Chinese medicine(TCM)syndrome types in the real world.Methods A prospective real-world study was performed in the 36 patients(involving 42 eyes)with PDR treated by anti-VEGF combined with PPV in the Department of Ophthalmology of the First Affiliated Hospital of Guangzhou University of Chinese Medicine from March 2019 to December 2019.According to the TCM syndrome manifestations,the patients were differentiated as qi-yin deficiency complicated with blood stasis obstructing collaterals type(15 cases,involving 18 eyes;shorten as qi-yin deficiency type),liver-kidney deficiency and ocular collaterals failing in the nourishment type(14 cases,involving 17 eyes;shorten as liver-kidney deficiency type),and yin-yang deficiency complicated with blood stasis and phlegm coagulation type(7 cases,involving 7 eyes;shorten as yin-yang deficiency type).The patients were treated with anti-VEGF therapy first and then received PPV after 5-7 days.Aqueous humor was sampled during anti-VEGF therapy and PPV.After treatment,the efficacy of PDR patients with different TCM syndromes was compared.Moreover,the patients were observed in the best corrected visual acuity(BCVA)of the affected eyes before surgery and 3 months after surgery,levels of cytokines in the aqueous humor before and after anti-VEGF treatment,macular central retinal thickness(CRT),area of the foveal avascular zone(FAZ),the blood density of macular center,inner ring,outer ring and intact macula 3 months after surgery,and the postoperative complications.Results(1)The difference of the therapeutic efficacy of PDR patients with various TCM syndrome types was statistically significant(P＜0.05).Among 3 syndrome types,the best efficacy was found in the qi-yin deficiency type,followed by liver-kidney deficiency type,and then yin-yang deficiency type,with the total efficacy rate being 88.89%(16/18),52.94%(9/17),and 42.86%(3/7),respectively.(2)Three months after surgery,the logarithmic value of minimum angle of resolution(LogMAR)for BCVA of patients with qi-yin deficiency type was significantly superior to that of patients with yin-yang deficiency type,with the difference being statistically significant(P＜0.05).After the anti-VEGF treatment,the levels of cytokines in the aqueous humor of the patients varied in the 3 syndrome types:vascular endothelial growth factor A(VEGF-A)level in the patients with the 3 syndrome types was significantly lower,placental growth factor(PLGF)and angiopoietin-like protein 4(ANGPTL4)levels in the patients with qi-yin deficiency type were higher,and interleukin 8(IL-8)level in the patients with liver-kidney deficiency type was higher than those before treatment,and the differences were statistically significant(P＜0.05 or P＜0.01).The blood density of macular outer ring and intact macula in the patients with qi-yin deficiency type and liver-kidney deficiency type was larger than that in the patients with yin-yang deficiency type,and the differences were statistically significant(P＜0.05).However,the differences of CRT,FAZ area,and blood density of macular center and inner ring among the 3 syndrome types were not statistically significant(P＞0.05).(3)The incidence of postoperative complications in the patients with yin-yang deficiency type was relatively high,but the difference among the 3 syndrome types was not statistically significant(P＞0.05).Conclusion In the real world,the best efficacy of anti-VEGF combined with PPV treatment in PDR patients with different TCM syndrome types can be achieved in the patients differentiated as qi-yin deficiency type,followed by liver-kidney deficiency type,and then yin-yang deficiency type.After anti-VEGF treatment,the levels of cytokines in the aqueous humor of the patients vary in the 3 syndrome types.Three months after the operation,the patients with qi-yin deficiency type and liver-kidney deficiency type have larger blood density of macular outer ring and intact macula,and exert good prognosis.

Objective To explore the eradication rate of human papillomavirus(HPV)and gestational outcome of patients with high-grade squamous intraepithelial disease of the cervix(HSIL)after loop electrosurgical excision procedure(LEEP)by transvaginal dissection of the vesicorectal form the cervix.Methods A total of 53 patients treated with LEEP by transvaginal dissection of the vesicorectal form the cervix in Obstetrics and Gynecology Hospital,Fudan University from Jan to Dec,2019 were investigated.Clinical information of cervical cytological examination,HPV test and cervical biopsy under colposcopy were followed up for 6,12 and 24 months post-LEEP were collected.HPV infection in these 53 patients were compared before and after LEEP surgery.The rate of successful fertility of the cohort,the HPV conversion rate of patients with hysterectomy and LEEP done were compared.The association between the pathological type and positive surgical margin and the association between HPV infection type and positive surgical margin were analyzed.Results HPV infection rate of was 94.3%(50/53)and the proportion of HPV16 and/or 18 infection was 75.5%(40/53).Mono-HPV infection rate(69.8%,37/53)was significantly higher than mixed HPV infection rate(22.7%,13/53).Thirty-eight patients(71.7%)were found with positive surgical margin in previous LEEP operation.Fifteen patients had recurrence(28.3%)and 40 patients(75.5%)successfully delivered baby after surgery.Postoperative pathology was mainly HSIL,accounting for 66%(30/53),and 28.3%patients(15/53)had no pathological change.Forty cases had satisfying fertility-conservative operation outcome with negative surgical margin,and 38 patients eradicated HPV infection after LEEP,which took up 95%of patients with satisfying fertility-conservative operation.There was no significant difference of positive resection margin rate in between groups of HPV16/18 infection and other types.Five cases had successful delivery(12.5%,5/40)with 1 case of vaginal delivery and 4 cases of cesarean section.Among these 5 cases,3 cases undertook preventive cervical cerclage,with 1 case of vaginal delivery and 2 cases of cesarean sections.Conclusion HPV eradication rate and surgical outcome could be significantly improved by LEEP with transvaginal dissection of the vesicorectal from the cervix,which satisfied the fertility preservation of females at reproductive age.

Objective This study aimed to investigate the effect of stage Ⅰ comprehensive cardiac rehabili-tation in patients with acute ST elevation myocardial infarction(STEMI)after emergency percutaneous coronary intervention(PCI).Methods A total of 72 patients with acute ST-segment elevation myocardial infarction combined with PCI admitted to the Department of Cardiovascular Medicine of Beijing Electric Power Hospital of State Grid Corporation from June 2021 to June 2022,which were selected as the research objectsand divided into control group and observation group randomly(36 cases in each group).The control group was treated with routine nursing and health education,and the observation group with stage Ⅰ comprehensive cardiac rehabilitation,including initial assessment(cardiovascular comprehensive assessment),exercise training(exercise training and breathing train-ing),daily activity suggestions and health education,discharge assessment(six-minute walking test and Barthel index assessment).The score of Barthel index(BI)at discharge,the 6-minute walking test distance(6MWD)at discharge,the incidence of major adverse cardiovascular event(MACE)during hospitalization and within one month of discharge,and the length of stay were compared between the two groups.Results After intervention,the six-minute walking test distance(6MWD)and Barthel index(BI)score in the observation group were better than those in the control group,the difference was statistically significant(P<0.05).The incidence of major adverse cardiovascular events(MACE)during hospitalization and one month after discharge was lower in the observation group than in the control group,and the difference was statistically significant(P<0.05).The length of hospital-ization in observation group was lower than that in control groupbut there was no statistical difference(P>0.05).Conclusion The application of phase Ⅰ comprehensive cardiac rehabilitation training in patients with acute ST-segment elevation myocardial infarction combined with emergency PCI could improve the patients'exercise ability,improve their ability of daily activity,reduce the incidence of major adverse cardiovascular events(MACE)in the early stage of the disease,facilitate the patients to return to their families and society as soon as possible,and improve their quality of life.It has high clinical application value.

BACKGROUND:The pituitary gland is an important endocrine organ in the body.Certain diseases can cause damage to the pituitary gland,such as pituitary adenoma and abnormal hormone secretion.Pituitary stem cells,due to their self-renewal and multi-directional differentiation potential,are expected to become a new therapeutic approach for repairing damaged pituitary glands. OBJECTIVE:To isolate and culture pituitary stem cells using the suspension cell ball culture method and identify their proliferation and differentiation ability. METHODS:Pituitary stem cells were isolated and cultured from the pituitary gland of newborn New Zealand white rabbits using the suspension cell ball culture method,and their morphological characteristics were observed.Immunofluorescence cytochemistry was used to detect the expression of pituitary stem cell markers SOX2 and Nestin.EdU labeling method was utilized to detect the proliferative ability of pituitary stem cells.After adherent and induced differentiation,the hormone levels in the culture medium were detected by ELISA. RESULTS AND CONCLUSION:Pituitary stem cell spheres could be successfully isolated by the suspension cell ball culture method,with strong proliferative ability.Positive expression of stem cell-specific markers SOX2 and Nestin was found in the cultured cells.After induction and differentiation,adrenocorticotropic hormone,thyroid hormone,growth hormone,luteinizing hormone,follicle-stimulating hormone,and prolactin levels significantly increased in the medium(P<0.001),with strong differentiation ability.

Objective To determine the acetylation level of nucleophosmin(NPM)in female breast cancer and to discuss its function through mutation of modified lysine sites.To construct positive and negative NPM mutants on its acetylated lysine sites and to express them in breast cancer cells.Methods Acetylation level and acetylated lysine sites of NPM in three breast cancer tissues and para-carcinoma tissues were detected by acetylome technology;NPM mutants were constructed by site-directed mutagenesis PCR,specific PCR products were digested by DpnI and transformed into Escherichia coli(E.coli)to obtain specific plasmids for mutants;The accuracy of mutants were verified by double restriction enzyme digestion and sequencing;The mutants were expressed in BT-549 cells by transient transfection and verified by RT-PCR method.Protein expression and acetylation level of NPM were validated by Western blotting;Function of NPM acetylation was analyzed by proteomic detection and bioinformatic analysis.Results The 27th and 32nd lysine of NPM were highly acetylated in breast cancer tissues,which were 2.76 and 2.22 times higher than those in adjacent normal tissues,respectively;The NPM mutants showed the same molecular weight as that of wild type NPM and contained expected mutation sites;Corresponding NPM mRNA levels of BT-549 cells transfected with NPM mutants were significantly increased.With the increase of wild type NPM expression level,NPM acetylation level increased,while decreased after 27th lysine underwent negative mutation.NPM acetylation can significantly change the expression levels of 101 proteins in BT-549 cells,which are enriched in regulation of cellular macromolecule biosynthesis,DNA-template transcription,RNA biosynthesis and RNA metabolism process.Conclusion NPM is highly acetylated in breast cancer and can play a key role in cellular macromolecule biosynthesis,DNA-templated transcription,RNA biosynthesis and RNA metabolism process.

Objective To investigate the clinical efficacy of transjugular intrahepatic portosystemic shunt(TIPS)combined with indwelling catheter-directed thrombolysis for the treatment of portal vein thrombosis(PVT).Methods The clinical efficacy of 307 patients with portal hypertension complicated by PVT,who received successful TIPS combined with indwelling catheter-directed thrombolysis at the Affiliated Beijing Shijitan Hospital of Capital Medical University of China between January 2016 and December 2019,were retrospectively analyzed.Before and after TIPS,the inferior vena cava pressure(IVCP)and portal vein pressure(PVP)were measured,and the pre-TIPS,post-TIPS(before thrombolysis),and post-thrombolysis portal pressure gradient(PPG,PPG=PVP-IVCP)was separately calculated.Reexamination of portal venography DSA was performed to determine the degree of PVT disappearance and whether the shunt was unobstructed.All patients were followed up for one year.Results The pre-TIPS,post-TIPS(before thrombolysis),and post-thrombolysis mean PPG was(24.50±6.91)mmHg,(18.51±5.11)mmHg,and(10.17±3.97)mmHg,respectively.The post-thrombolysis mean PPG was strikingly lower than the pre-thrombolysis values,the differences were statistically significant(P＜0.001).Among the 307 patients,complete disappearance of PVT was observed in 221(72.3%),remarkable reduction of PVT in 86(27.7%),and no invalid result was seen.The patients having complete patency of the shunt flow accounted for 85.7%of the 307 patients(261/307),and the patients having partial patency of the shunt flow accounted for 14.3%of the 307 patients(46/307).Forty-two patients developed complications,and no death occurred.All patients were followed up for one year,and the main clinical symptoms were improved or completely disappeared.Among the 307 patients,an increase in thrombus volume was found in 17(5.5%)when compared to their postoperative values,which returned to the first-time postoperative level after local treatment of the thrombus via the TIPS shunt combined with catheter-directed thrombolysis.Within one year after TIPS and thrombolysis,overt hepatic encephalopathy(OHE)occurred in 54 patients(17.6%,54/307).One patient died of hepatic failure 9 months after TIPS,another patient died of cerebral hemorrhage 11 months after TIPS,and all the remaining patients were alive.Conclusion For patients with portal hypertension complicated by PVT,TIPS combined with indwelling catheter-directed thrombolysis is clinically safe and effective.The standardized,systematic management of the whole therapeutic process should be strengthened.(J Intervent Radiol,2024,32:22-27)