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Objective:To evaluate the role of signal transducer and activator of transcription 3 (STAT3)/nuclear receptor coactivator 4 (NCOA4)-mediated ferritinophagy in salidroside-induced attenuation of intestinal ischemia-reperfusion (I/R) injury in mice and the relationship with ferroptosis.Methods:Thirty-six SPF-grade healthy male C57BL mice, aged 6-8 weeks, weighing 20-25 g, were divided into 6 groups ( n=6 each) by a random number table method: sham operation group (S group), sham operation+ salidroside group (SS group), intestinal I/R group (IR group), intestinal I/R+ salidroside group (IS group), intestinal I/R+ salidroside+ autophagy activator rapamycin group (ISR group) and intestinal I/R+ salidroside+ STAT3 activator colivelin group (ISC group). The intestinal I/R injury model was established by clamping the superior mesenteric artery for 45 min followed by 30-min reperfusion in IR, IS, ISR and ISC groups, while the superior mesenteric artery was only isolated without clipping in S and SS groups. At 1 week before developing the model, salidroside 40 mg/kg was intraperitoneally injected once a day for 7 consecutive days in SS, IS, ISC and ISR groups, rapamycin 4 mg/kg was intraperitoneally injected once a day for 7 consecutive days in group ISR, colivelin 1 mg/kg was intraperitoneally injected once a day for 7 consecutive days in group ISC, while the equal volume of normal saline was given instead in the rest two groups. The mice were sacrificed at 30 min of reperfusion, and intestinal tissues were obtained for examination of the pathological changes after HE staining (with a optical microscope) which were scored according to Chiu and for determination of contents of malondialdehyde (MDA), Fe 2+, glutathione (GSH) and reactive oxygen species (ROS), activity of superoxide dismutase (SOD) and expression of p-STAT3, STAT3, glutathione peroxidase 4 (GPX4), NCOA4 and ferritin heavy chain 1 (FTH1) in intestinal tissues (by Western blot). Results:Compared with group S, the Chiu′s score and contents of MDA, Fe 2+ and ROS were significantly increased, the content of GSH was decreased, the activity of SOD was decreased, the expression of p-STAT3 and NCOA4 was up-regulated, the expression of GPX4 and FTH1 was down-regulated, the p-STAT3/STAT3 ratio was increased ( P<0.05), pathological injury was found in intestinal tissues, and no significant change was found in the aforementioned indexes in group IR( P>0.05). Compared with group IR, the Chiu′s score and contents of MDA, Fe 2+ and ROS were significantly decreased, GSH content was increased, SOD activity was increased, the expression of p-STAT3 and NCOA4 was down-regulated, the expression of GPX4 and FTH1 was up-regulated, p-STAT3/STAT3 ratio was decreased ( P<0.05), and the pathological injury was significantly alleviated in intestinal tissues in group IS. Compared with group IS, the Chiu′s score and contents of MDA, Fe 2+ and ROS were significantly increased, GSH content was decreased, SOD activity was decreased, the expression of p-STAT3 and NCOA4 was up-regulated, the expression of GPX4 and FTH1 was down-regulated, p-STAT3/STAT3 ratio was increased ( P<0.05), and the pathological injury was aggravated in intestinal tissues in ISR and ISC groups. There was no statistically significant difference in the expression of STAT3 among the five groups ( P>0.05). Conclusions:STAT3/NCOA4-mediated ferritinophagy is involved in the process of salidroside-induced reduction of intestinal I/R injury in mice, which may be related to inhibiting ferroptosis.
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Objective:To evaluate the role of endoplasmic reticulum stress-mediated apoptosis in proanthocyanidins-induced attenuation of intestinal ischemia-reperfusion (I/R) injury in mice.Methods:Thirty SPF healthy adult male C57BL/6 mice, aged 8-10 weeks, weighing 20-25 g, were divided into 5 groups ( n=6 each) by a random number table method: sham operation group (S group), sham operation + proanthocyanidins group (S+ PC group), intestinal I/R group (I/R group), intestinal I/R + proanthocyanidins group (I/R+ PC group) and intestinal I/R + proanthocyanidins + tunicamycin group (I/R+ PC+ TM group). The superior mesenteric artery was clamped for 60 min and reperfused for 120 min to establish a mouse intestinal I/R injury model.Proanthocyanidin 100 mg/kg was given by intragastric gavage every day 1 week before ischemia in S+ PC group, I/R+ PC group and I/R+ PC+ TM group, and the equal volume of normal saline was given for 7 consecutive days in S group and I/R group, and endoplasmic reticulum stress agonist tunicamycin 1 mg/kg was intraperitoneally injected at 24 h before ischemia in I/R+ PC+ TM group.The mice were sacrificed at 120 min of reperfusion, and the small intestinal tissues were taken for microscopic examination of the histopathological changes (using light microscopy) and for determination of the level of diamine oxidase (DAO) (by enzyme-linked immunosorbent assay), cell apoptosis (by TUNEL method), glucose regulatory protein 78 (GRP78), C/EBP-homologous protein (CHOP) and cleaved caspase-3, Bax and Bcl-2 (by Western blot). Intestinal damage was assessed and scored according to Chiu, and the apoptosis index (AI) and Bcl-2/Bax ratio were calculated. Results:Compared with S group, the Chiu′s score, level of DAO and AI were significantly increased, the expression of GRP78, CHOP, cleaved caspase-3 and Bax was up-regulated, the expression of Bcl-2 was down-regulated, the ratio of Bcl-2/Bax was decreased( P<0.05), and the pathological damage occurred in the small intestinal tissue in I/R group, and no significant change was found in the aforementioned indexes in S+ PC group ( P>0.05). Compared with I/R group, the Chiu′s score, DAO level and AI were significantly decreased, the expression of GRP78, CHOP, cleaved caspase-3 and Bax was down-regulated, the expression of Bcl-2 was up-regulated, the ratio of Bcl-2/Bax was increased( P<0.05), and the pathological injury to the small intestinal tissue was significantly reduced in I/R+ PC group. Compared with I/R+ PC group, the Chiu′s score, level of DAO and AI were significantly increased, the expression of GRP78, CHOP, cleaved caspase-3 and Bax was up-regulated, the expression of Bcl-2 was down-regulated, and the ratio of Bcl-2/Bax was decreased( P<0.05), and the pathological damage to the small intestinal tissue was aggravated in I/R+ PC+ TM group. Conclusions:Proanthocyanidins can alleviate intestinal I/R injury by inhibiting endoplasmic reticulum stress-mediated cell apoptosis in mice.
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Objective:To evaluate the role of silencing information regulatory factor 1/nuclear factor E2 related factor 2 (SIRT1/Nrf2) signaling pathway in berberine preconditioning-induced reduction of intestinal ischemia-reperfusion (I/R) injury in mice and the relationship with ferroptosis.Methods:Thirty-six SPF-grade healthy male C57BL/6 mice, aged 8-10 weeks, weighing 22-25 g, were divided into 6 groups ( n=6 each) by a random number table method: sham operation group (S group), sham operation + berberine preconditioning group (SB group), intestinal I/R group (IR group), intestinal I/R + berberine preconditioning group (B group), intestinal I/R + berberine preconditioning + SIRT1 inhibitor EX527 group (BE group) and berberine preconditioning + intestinal I/R + ferroptosis inducer RSL3 group (BR group). The model of intestinal I/R injury was prepared by clamping the superior mesenteric artery for 45 min followed by 30-min reperfusion in IR group, B group, BE group and BR group, while the superior mesenteric artery was only isolated without ligation in S group and SB group. Berberine 50 mg/kg was administered by intragastric gavage once a day starting from 7 days before developing the model in SB group, B group, BE group and BR group. EX527 5 mg/kg and RSL3 5 mg/kg were intraperitoneally injected once a day at 3 days before surgery in BE group and BR group, respectively. The equal volume of normal saline was given in the other groups. The mice were sacrificed at 30 min of reperfusion, and the intestinal tissues were taken for microscopic examination of the pathological changes of intestinal mucosa (with a light microscope) which was scored according to Chiu and for determination of the contents of Fe 2+ and malondialdehyde (MDA) and superoxide dismutase (SOD) activity (by colorimetry), glutathione (GSH) content (by enzyme-linked immunosorbent assay), reactive oxygen species (ROS) content (by fluorescence staining), and expression of glutathione peroxidase 4 (GPX4), SIRT1 and Nrf2 (by Western blot). Results:Compared with S group, Chiu′s score was significantly increased, the contents of Fe 2+, MDA and ROS were increased, the content of GSH and activity of SOD were decreased, the expression of GPX4 was down-regulated, and the expression of SIRT1 and Nrf2 was up-regulated in IR group ( P<0.05), and no significant change was found in Chiu′s score in SB group ( P>0.05). Compared with IR group, Chiu′s score was significantly decreased, the contents of Fe 2+, MDA and ROS were decreased, the content of GSH and activity of SOD were increased, the expression of GPX4 was up-regulated, and the expression of SIRT1 and Nrf2 was up-regulated in B group ( P<0.05). Compared with B group, Chiu′s score was significantly increased, the contents of Fe 2+, MDA and ROS were increased, the content of GSH and activity of SOD were decreased, and the expression of GPX4 was down-regulated in BE and BR groups, and the expression of SIRT1 and Nrf2 was down-regulated in BE group( P<0.05). Conclusions:The mechanism by which berberine preconditioning reduces intestinal I/R injury may be associated with activation of SIRT1/Nrf2 signaling pathway, thus inhibiting ferroptosis in mice.
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Background@#MicroRNAs (miRNAs) exert an essential contribution to obesity and type 2 diabetes mellitus (T2DM). This study aimed to investigate the differences of miRNAs in the presence and absence of T2DM in patients with obesity, as well as before and after bariatric surgery in T2DM patients with obesity. Characterization of the common changes in both was further analyzed. @*Methods@#We enrolled 15 patients with obesity but without T2DM and 15 patients with both obesity and T2DM. Their preoperative clinical data and serum samples were collected, as well as 1 month after bariatric surgery. The serum samples were analyzed by miRNA sequencing, and the miRNAs profiles and target genes characteristics were compared. @*Results@#Patients with T2DM had 16 up-regulated and 32 down-regulated miRNAs compared to patients without T2DM. Improvement in metabolic metrics after bariatric surgery of T2DM patients with obesity was correlated with changes in miRNAs, as evidenced by the upregulation of 20 miRNAs and the downregulation of 30 miRNAs. Analysis of the two miRNAs profiles identified seven intersecting miRNAs that showed opposite changes. The target genes of these seven miRNAs were substantially enriched in terms or pathways associated with T2DM. @*Conclusion@#We determined the expression profiles of miRNAs in the obese population, with and without diabetes, before and after bariatric surgery. The miRNAs that intersected in the two comparisons were discovered. Both the miRNAs discovered and their target genes were closely associated with T2DM, demonstrating that they might be potential targets for the regulation of T2DM.
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Although the development of COVID-19 vaccines has been a remarkable success, the heterogeneous individual antibody generation and decline over time are unknown and still hard to predict. In this study, blood samples were collected from 163 participants who next received two doses of an inactivated COVID-19 vaccine (CoronaVac®) at a 28-day interval. Using TMT-based proteomics, we identified 1,715 serum and 7,342 peripheral blood mononuclear cells (PBMCs) proteins. We proposed two sets of potential biomarkers (seven from serum, five from PBMCs) at baseline using machine learning, and predicted the individual seropositivity 57 days after vaccination (AUC = 0.87). Based on the four PBMC's potential biomarkers, we predicted the antibody persistence until 180 days after vaccination (AUC = 0.79). Our data highlighted characteristic hematological host responses, including altered lymphocyte migration regulation, neutrophil degranulation, and humoral immune response. This study proposed potential blood-derived protein biomarkers before vaccination for predicting heterogeneous antibody generation and decline after COVID-19 vaccination, shedding light on immunization mechanisms and individual booster shot planning.
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Humans , COVID-19 Vaccines , Leukocytes, Mononuclear , Proteomics , COVID-19/prevention & control , Vaccination , Antibodies , Antibodies, Viral , Antibodies, NeutralizingABSTRACT
OBJECTIVE@#To screen for Jk(a-b-) phenotype among blood donors from Jining area and explore its molecular basis to enrich the rare blood group bank for the region.@*METHODS@#The population who donated blood gratuitously at Jining Blood Center from July 2019 to January 2021 were selected as the study subjects. The Jk(a-b-) phenotype was screened with the 2 mol/L urea lysis method, and the result was confirmed by using classical serological methods. Exons 3 to 10 of the SLC14A1 gene and its flanking regions were subjected to Sanger sequencing.@*RESULTS@#Among 95 500 donors, urea hemolysis test has identified three without hemolysis, which was verified by serological method as the Jk(a-b-) phenotype and demonstrated no anti-Jk3 antibody. The frequency of the Jk(a-b-) phenotype in Jining area is therefore 0.0031%. Gene sequencing and haplotype analysis showed that the genotypes of the three samples were JK*02N.01/JK*02N.01, JK*02N.01/JK-02-230A and JK*02N.20/JK-02-230A, respectively.@*CONCLUSION@#The splicing variant of c.342-1G>A in intron 4, missense variants of c.230G>A in exon 4, and c.647_ 648delAC in exon 6 probably underlay the Jk(a-b-) phenotype in the local population, which is different from other regions in China. The c.230G>A variant was unreported previously.
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Humans , Phenotype , Blood Donors , Hemolysis , Kidd Blood-Group System/genetics , Urea , Molecular BiologyABSTRACT
OBJECTIVE： To analyze the formulation regularity of Chinese patent medicine for knee osteoarthritis （KOA）， and to provide reference for the clinical standard use of Chinese patent medicine for KOA and the research and development of new drugs. METHODS： Chinese Pharmacopoeia （2015 edition， part Ⅰ），National Drug Reimbursement List （2017 edition）， National Essential Drug List （2017 edition）， Chinese Materia Medica Preparation （1992 version）， Compilation of National Standard for Chinese Patent Medicines （2002 edition）， Handbook of Rational Application of Chinese Patent Medicines in Surgery and Orthopedics （2010 edition） were searched to collect the type and formulation of Chinese patent medicines for “KOA”， “osteoarthritis”， “Bi syndrome”， “promoting blood circulation and removing blood stasis， dispelling wind and removing dampness， tonifying liver and kidney”. Supplementary the type and formulations of Chinese patent medicines for KOA by questionaire survey of clinial experts. The types， properties， meridian tropism， frequency and combination of medicinal materials used in Chinese patent medicine formulations were counted by using TCM inheritance auxiliary platform software V 2.5. The association rules and entropy clustering method were used to analyze the formulation regularity. RESULTS： A total of 190 Chinese patent medicines were collected， involving 289 TCM. With the top 10 used frequency being Angelica sinensis （75 times）， Boswellia carterii （55 times）， Carthamus tinctorius （53 times）， Commiphora myrrha （51 times）， Achyranthes bidentata （49 times）， Notopterygium incisum （47 times）， Angelica pubescens （45 times）， Saposhnikovia divaricata （45 times）， Angelica dahurica （39 times）， Ligusticum chuanxiong （39 times）. Medicinal material were mainly Xinwen in properties field and mainly liver meridian and spleen meridian in meridian entry field. Top 5 frequency of medicinal material combinations were C. myrrha-B. carterii， B. carterii-A. sinensis， A. sinensis-N. incisum， A. bidentata-A. sinensis， L. chuanxiong-A. sinensis. 14 core medicinal material combinations and 7 new developed formulations were concluded. CONCLUSIONS： This study analyzed the formulation regularity of Chinese patent medicines for KOA with the help of TCM inheritance auxiliary platform software V 2.5， which can provide reference for clinical differentiation of symptoms and signs and research and development of related new medicines related to KOA.
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Objective Currently, there are few articles about kinesiophobia in patients with total knee arthroplasty (TKA) in China.This study aims to investigate the incidence of kinesiophobia and its influencing factors in TKA patient, and provide evidence for the intervention of kinesiophobia.Methods A total of 298 TKA patients from our hospital were investigated by general information questionnaire, Tampa Scale of Kinesiophobia (TSK), Knee Self-Efficacy Scales, Simplified Coping Style Questionnaire and Social Support Rating Scale.Single-factor analysis and logistic regression analysis were conducted to explore influencing factors.Results The score of TSK was 38.50±13.52, and 31.88% of TKA patients reported kinesiophobia.Logistic regression analysis showed that duration of pain (OR=5.546, 95%CI: 2.143-14.353), education level (OR=0.145, 95%CI: 0.067-0.314), self efficacy(OR=0.606, 95%CI: 0.470-0.780), positive response (OR=0.784, 95%CI: 0.683-0.900), objective support (OR=0.807, 95%CI: 0.691-0.943) and utilization of social support (OR=0.507, 95%CI: 0.461-0.705) were factors influencing kinesiophobia in TKA patients.Conclusion Attention should be paid to the kinesiophobia in TKA patients, especially those influencing factors including duration of pain, education level, and objective support.Health care providers should encourage early stage rehabilitation exercise to improve the postoperative knee function of TKA patients.
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BACKGROUND:Early rehabilitation after total knee arthroplasty (TKA) can help the rapid functional recovery of knee,but some patients are fear of moving and refuse to do exercising because of kinesiophobia,thereby delaying functional recovery.Foreign study has confirmed that patients with cognitive behavioral therapy (CBT) can effectively alleviate kinesiophobia,but no related studies have been reported in China.OBJECTIVE:To explore the effectiveness of CBT on patients with kinesiophobia after TKA.METHODS:108 patients with kinesiophobia after TKA were recruited from Department of Orthopedics,the First Affiliated Hospital of Zhengzhou University,China between March 2014 and January 2015,and were then equivalently randomized into experimental and control groups using random number table.Both groups were subjected to conventional treatment,while the experimental group received extra CBT.The scores of Tampa Scale of Kinesiophobia and American Knee Society were evaluated immediately,1,3,and 6 months after hospital discharge;the hospitalization time and total hospitalization expenditure were compared between two groups immediately after discharge.RESULTS AND CONCLUSION:The scores of Tampa Scale of Kinesiophobia in the experimental group were significantly lower than those in the control group at different time points postoperatively (P ＜ 0.05).The scores of American Knee Society in the experimental group were significantly higher than those in the control group at 3 and 6 months after discharge and kept on rising (P ＜ 0.01),but showed no significant difference at discharge and 1 month after discharge (P ＞ 0.05).(2) The hospitalization time and total hospitalization expenditure in the experimental group were significantly lower than those in the control group (P ＜ 0.01).(3) To conclude,CBT cannot only shorten the hospitalization time and total hospitalization expenditure,but also relieve kinesiophobia,which is conductive for functional recovery of the knee.
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<p><b>OBJECTIVE</b>To explore the serological characteristics and molecular basis for an individual with para-Bombay phenotype.</p><p><b>METHODS</b>Blood type of the proband was determined with routine serological methods. Exons 6 and 7 of the ABO gene and coding regions of the FUT1 and FUT2 genes were amplified by PCR and sequenced.</p><p><b>RESULTS</b>The para-Bombay phenotype was confirmed to be of Ah-secretion type. The genotype of the individual was determined as A102/O01. Position 328 of the FUT1 gene was mutated from A to G, resulting in replacement of Alanine (Ala) at position 110 by Threonine (Thr).</p><p><b>CONCLUSION</b>The G to A mutation of nt328 of the FUT1 gene probably underlies the para-Bombay phenotype in this individual.</p>
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Adult , Female , Humans , ABO Blood-Group System , Genetics , Alleles , Base Sequence , Exons , Genotype , Molecular Sequence Data , Mutation , Point MutationABSTRACT
Objective To translate the English version of an abbreviated eight-item measure of Penn State Worry Questionnaire (PSWQ-A) into Chinese and to test the reliability and validity of the Chinese version of PSWQ-A in patients after total Hip Replacement.Methods Totally 218 patients after total hip replacement were recruited and were investigated by the Chinese version of PSWQ-A.Results The internal consistency coefficient of the Chinese version of PSWQ-A were 0.885.Factor analysis got two factors,which explain 68.428％ of the total variance.The content validity index was 0.924.Using SAS as an external criterion,The correlation coefficients of the PSWQ-A and SAS scores were 0.891,P＜0.05.Conclusions The Chinese version of PSWQ-A has been proved to be reliable and valid.It can be used as a valid tool for the measurement of self-efficacy in patients after total hip replacement.
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Objective To explore the effect of the combination of video feedback and telephone intervention on postdischarge rehabilitation in patients receiving artificial total hip replacement.Methods A total of 60 patients receiving artificial total hip replacement were divided into intervention group and control group with 30 cases each by random digits table method.The combination of video feedback and telephone intervention was conducted in intervention group,and conventional discharge guidance and telephone follow-up on a regular basis in control group.In intervention group,the patients or their family member were asked to record the video of the exercises and daily living when patients were at home according instructions and researchers assessed the condition of patient's joint and muscle function and provided individual education and instructions to the patients on the basis of the feedback video through telephone.Harris hip function rating scale and Functional Independence Measure (FIM) were investigated at discharge and 1,3,6 months after discharge.Results There were no significant differences in Harris hip function rating scale and FIM score at discharge between 2 groups,P＞0.05.Harris hip function rating scale score were (72.70± 11.65),(83.11±5.12),(94.83±3.08) scores at 1,3,6 months after discharge in intervention group,and (63.20±14.96),(77.00±11.62),(87.50±7.94) scores in control group,there were significant differences,P＜0.05 or ＜0.01.FIM score were (96.30±4.63),(108.76±3.28),(125.79±1.19) scores at 1,3,6 months after discharge in intervention group,and (78.23±5.09),(86.43±6.30),(96.54±5.35) scores in control group,there were significant differences,P＜0.01.Conclusion The combination of video feedback and telephone intervention could help to promote the postdischarge hip function for patients receiving artificial total hip replacement.
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Background The pathological basis of human primary open angle glaucoma mainly is the degeneration of trabecular meshwork and Schlemm canal.As the outflow pathway of aqueous humor, the tissue origin and characteristics of trabecular meshwork remain less clear.Studying the characteristics of trabecular meshwork may contribute a new thought to the treatment of primary open angle glaucoma.Objective This study was to explore the histological property of trabecular meshwork by detecting the expression of D2-40, a lymphatic biomarker,and CD31,CD34 and smooth muscle actin (SMA), some vascular endothelial biomarkers.Methods Twenty specimens of adult eyeballs were collected from Zhongshan Ophthalmic Center of Sun Yat-sen University,including l0 eyeballs from accident death donors,3 enucleated eyeballs due to posterior segment of choroidal malignant melanoma and 7 orbital exenterated eyeballs.The series ocular meridian sections with the thickness of 4 μm were prepared for the haematoxylin-eosin staining.Immunohistochemistry was employed to detect the expression of D2-40,CD31 ,CD34 and SMA in trabecular meshwork and Schlemm canal.Results Intact structure of chamber angle was observed under the optical microscope by haematoxylin-eosin staining.Human trabecular meshwork tissue was presented with meshlike structure,while Schlemm canal showed the lumen structure.No abnormal substance and periphery synechia were found.D2-40 was strongly expressed in the trabecular meshwork endothelial cells with staining score for 3 points, but CD31, CD34 and SMA were negatively expressed.In contrast, CD31, CD34 and SMA rather than D2-40 were positively expressed in the Schlemm canal.In addition, CD31 and CD34 were also positively expressed in the vascular endothelial cells around the trabecular meshwork.Conclusions The trabecular meshwork expresses lymphatic biomarker rather than vascular biomarkers.This result indicates that trabecular meshwork of human eyes is lymphatic vessel-like tissue.
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Inflammation is the defense response of the living tissues possessing vascular system to stimulations of various injury factors ,which plays a vital role in the initiation and progression of many major diseases .Drugs used to treat inflammation in the clinical mainly include non‐steroidal anti‐inflammatory drugs (NSAIDs) ,steroidal anti‐inflammatory drugs (SAIDs) and traditional Chinese medicine .As synthetic anti‐inflammatory drugs used in clinical currently have obvious adverse reactions , more and more attention were paid to seek anti‐inflammatory drugs from natural medicines .Reviews reported before mainly fo‐cus on anti‐inflammation mechanism of natural medicine ,however ,there are few reports on the summary of anti‐inflammatory natural products .Active natural products which were reported to possess anti‐inflammatory effects in recent years were summa‐rized in order to provide information for further study of anti‐inflammatory drugs research .
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This review summarized the research status on DNA molecular identification of plants in the genus Chrysanthemum. Some case studies on representative DNA molecular identification techniques, which included ge-nomic in situ hybridization (GISH), DNA molecular markers and DNA barcoding, were described. Simultaneously, the merits, demerits and development of the techniques were discussed. The above work provides evidence for the identi-fication and resource utilization of plants in the genus Chrysanthemum.
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This study was aimed to show the extraction research of cpDNA in Gentiana straminea Maxim. and G. crassicaulis Duthie ex Burk., which will increase the knowledge about the chloroplast genome sequence characteris-tics, understand its genetic diversity and improve the efficiency of breeding schemes. G. straminea Maxim. and G. crassicaulis Duthie ex Burk. were taken as study objects. The improved sucrose-gradient centrifugation and purifica-tion methods were used to obtain the cpDNA, measure the concentration, detect the OD value, and amplify the ITS2 sequence to verify the nucleus pollution. The results showed that the high quality and purity cpDNA (0.1 - 0.4 μg/10 g) was demonstrated without other pollution after ITS2 sequence amplification, which met to the subsequent effi-cient sequencing of chloroplast genomes. The purity and mass have an important influence on the accuracy and cred-ibility of the cp genome sequencing. It was concluded that this study improved the sucrose-gradient centrifugation and purification with great effect of the purity and mass of cpDNA, and guaranteed the obtaining of complete cpDNA of Gentiana.
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Chloroplast genome sequences have comprehensive application prospects in DNA barcoding and chloroplast engineering in traditional Chinese medicine. The complete chloroplast genome of Magnolia officinalis sequenced by high-throughput pyrosequencing and a sequencing procedure was established. Fourteen contigs were obtained after de nove assembly. The sequencing percent of coverage was 99.99%. The chloroplast genome is 160 183 bp in size, and has a typical quadripartite structure with the large (LSC, 88 210 bp) and small copy (SSC, 18 843 bp) regions separated by two copies of an inverted repeat (IRs, 26 565 bp each). chloroplast genes were successfully annotated, of which 17 genes located in each IR region. The chloroplast genome features in Magnolia officinalis are nearly identical to those from other Magnoliid chloroplast genomes. Phylogenetic analyses were performed based on 81 shared coding-genes for a total of 9 Magnolia samples of 5 closely related species. Results showed that distinguishing among species was generally straightforward at the species and population level. This study confirmed the effectiveness of our chloroplast genome sequencing procedure. The chloroplast genome can provide distinguishing differences to help identify Magnolia officinalis and its closely related plants.
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AIM: To examine the effect of compound salvae-dropping-pill (CSDP) on intracellular free calcium in cultured rat myocardial cells subjected to hypoxia and reoxygenation.METHODS: The Fluo- 3/AM was applied to probe intracellular calcium concentration and the fluorescent intensity was detected using laser confocal microscopy technique.RESULTS: Fluorescent intensity in hypoxia plus CSDP group was significantly lower (1 217 78?312 07) than that of hypoxia group (1 509?508 48), and the Fluorescent intensity of hypoxia/reoxygenation plus CSDP group was also markedly lower (1 567.91?577 61) than that of hypoxia/reoxygenation group (1 617.60?477.53).CONCLUSION: The cultured rat myocardial cells could be effectively protected by administration of CSDP in case of hypoxia and reoxygenation through decreasing the intracellular calcium concentration. [