Two cases of female-restricted X-linked syndromic mental retardation-99 caused by USP9X gene variations / 中华实用儿科临床杂志

Objective:To summarize and analyze the clinical and genotype features of female-restricted X-linked syndromic mental retardation-99(MRXS99F, OMIM: 300968)caused by USP9 X gene mutation, and to improve the clinicians′ understanding of the disease. Methods:Clinical data and genotypes of 2 children with MRXS99F treated in the Children′s Hospital of Nanjing Medical University in March 2020 (case 1) and June 2020 (case 2) were analyzed, and the relevant databases at home and abroad were reviewed to summarize the clinical characteristics and gene variation characteristics of the disease.Results:The 2 cases were 6 months old (case 1) and 5 years old (case 2), both showed psychomotor retardation.Case 1 presented a short stature, pigment abnormality, characteristic facial features, hypotonia, recurrent respiratory tract infections, laryngeal cartilage hypoplasia, atrial septal defect, feeding difficulty, hearing loss and brain hypoplasia.Case 2 had abnormal electroencephalogram.As confirmed by whole-exome sequencing, two children carried c. 6972+ 1G>A, c.6437C>T of USP9 X, respectively.Neither of the 2 variations was previously reported.Twenty-two cases of MRXS99F caused by USP9 X gene mutation were reported in 4 literatures globally, and 24 cases were combined with this study.The clinical manifestations of 20/22 children had special faces.All of them accompanied mental retardation combined with motor and language retardation, and carried neonatal variation. Conclusions:This is the first case report of MRXS99F induced by USP9 X gene variation in China.MRXS99F caused by functional deletion and variation of USP9 X gene is mainly characterized by psychomotor retardation, language disorder, special face and multiple congenital malformations.For children with unexplained growth retardation, special face and multiple congenital malformations, genetic testing like high-throughput sequencing should be carried out as early as possible to determine the etiology.

Practice of multidisciplinary discussion on case coding of cerebral infarction / 中华医院管理杂志

It is an important measure to establish an effective communication mechanism for filling in the front page of medical records to ensure the good operation of diagnosis-related groups. Taking cerebral infarction as an example, the authors carried out the pilot work of multidisciplinary cooperation mode based on disease types around its coding axis. The multi-disciplinary assistance model could provide a good platform for communication and learning among multiple disciplines, break the barriers between disciplines, improve the quality of the front page and medical record writing of clinicians, and improve the quality and efficiency of coders′ coding.

Network correlation of circRNA-miRNA and regulatory mechanisms of female major depressive disorder / 中华行为医学与脑科学杂志

Objective:To obtain differential expression profiles of circRNAs and miRNAs in peripheral blood of female patients with major depressive disorder based on high-throughput sequencing technology, and then construct an interaction network.Based on the outcome, it made a further exploration of the possible occurrence and development mechanisms of female major depressive disorder through functional annotation and pathway enrichment analysis.Methods:According to the ceRNA theory, circRNA-miRNA network was constructed by TargetScan software via predicting the binding sites.Subsequently, the GO functional enrichment analysis was performed, and KEGG pathway enrichment analysis were utlized to illustrate the target genes of co-expressed miRNAs.Thereby, the key genes related to major depressive disorder could be screened out.Results:A total of 724 differential circRNAs and 26 differential miRNAs were detected in female patients with major depressive disorder.And hsa_circ_0086092 and hsa-miR-146a-3p were the most co-expressed.Go functional annotations pointed out that it involved the regulation of nucleobase-containing compound metabolic process, regulation of RNA splicing, regulation of cell communication, amino acid transfer, regulation of RNA metabolic process, regulation of signaling and other biological processes.KEGG pathway analysis showed that target genes were mainly enriched in neurotrophin signaling pathway, Rap1 signaling pathway, FoxO signaling pathway, AMPK signaling pathway, cocaine addiction, mTOR signaling pathway, Jak-STAT signaling pathway, cAMP signaling pathway, etcetera.Among the predicted target genes, BDNF, FGF2, MAPK14, GRIN2A, GRIN2B, GRM2 and PDE4 have the highest correlation with major depressive disorder.Conclusion:Hsa_circ_0086092 may be involved in the occurrence and development of female major depressive disorder through interaction with hsa-miR-146a-3p.

Bioinformatics analysis of peripheral blood circRNAs in female patients with major depressive disorder / 中华行为医学与脑科学杂志

Objective:To screen the circRNAs with differential expression between female patients with major depressive disorder and healthy women, and to explore the circRNAs that might be associated with depressive disorder through bioinformatics analysis.Methods:Using high-throughput sequencing screen differentially expressed circRNAs of female major depressive disorder patients, |log 2FC|≥1 and FDR<0.05 were used to determine whether circRNA had a difference in expression.According to the miRNA sponges function of circRNA, the online databases were used to predict miRNAs which may be targeted by circRNAs, and miRNAs related target genes were also predicted by the five most different circRNA.GO and KEGG pathway enrichment analysis were used to predict biological processes for the target genes and signaling pathways.Based on the results of high-throughput sequencing, biological processes and signaling pathways related to depression, circRNAs related to depression were screened. Results:Thirteen targeted miRNAs were predicted by the five most different circRNAs(hsa_circ_0020959, hsa_circ_0005959, hsa_circ_0033064, hsa_circ_0006862, hsa_circ_0027732), and multiple biological processes and signaling pathways related to depressive disorders were predicted by the target genes, such as glucocorticoid receptor signaling pathway, interleukin-7 response, nervous system development, Wnt signaling pathway, FoxO signaling pathway, thyroid hormone signaling pathway, neurotrophin signaling pathway, and so on.Conclusion:All the 5 circRNAs enriched biological processes or signaling pathways related to depressive disorder, among which hsa_circ_0005959, hsa_circ_0033064, hsa_circ_0006862 and hsa_circ_0027732 may be more closely related to female major depressive disorder.

Epigenetic strategies synergize with PD-L1/PD-1 targeted cancer immunotherapies to enhance antitumor responses

Immunotherapy strategies targeting the programmed cell death ligand 1 (PD-L1)/programmed cell death 1 (PD-1) pathway in clinical treatments have achieved remarkable success in treating multiple types of cancer. However, owing to the heterogeneity of tumors and individual immune systems, PD-L1/PD-1 blockade still shows slow response rates in controlling malignancies in many patients. Accumulating evidence has shown that an effective response to anti-PD-L1/anti-PD-1 therapy requires establishing an integrated immune cycle. Damage in any step of the immune cycle is one of the most important causes of immunotherapy failure. Impairments in the immune cycle can be restored by epigenetic modification, including reprogramming the environment of tumor-associated immunity, eliciting an immune response by increasing the presentation of tumor antigens, and by regulating T cell trafficking and reactivation. Thus, a rational combination of PD-L1/PD-1 blockade and epigenetic agents may offer great potential to retrain the immune system and to improve clinical outcomes of checkpoint blockade therapy.

The research of nursing human resource optimization in emergency department / 中华急诊医学杂志

Objective To explore the way to optimize nursing human resources and analysis the effects of setting up the nurse part-time position in emergency department. Methods The nurses were recruited in the hospital of other departments, and these nurses worked in the rescue unite during the rush hour, the Practice Environment Scale of the Nursing Work Index (PES-NWI) was used to measure the effects of the alleviation nurse shortage, and the statistical methods was t test. Results The Part-time nurse is significantly to alleviate the pressure of the nurse shortage in emergency department, especially the cooperation ability between doctors and nurses were significantly improved. Conclusions Part-time nurses as a way to alleviate the shortage of nurse human resources is acceptable,which also could help promote the care quality.

Comparison of the Preventive and Therapeutic Effect of Different Dosage Forms of Vinegar Testudinis Car-apax on Mice with Hemopenia or Osteoporosis / 中国药房

OBJECTIVE:To compare the preventive and therapeutic effect of Vinegar testudinis carapax (VTC) water decoc-tion and powder on mice with hemopenia or osteoporosis. METHODS:Mice were randomly divided into normal group (1%CMC-Na),model group (1% CMC-Na),positive group (Compound e-jiao slurry 20 mL/kg or Gusongbao capsule 1.98 g/kg), VTC water decoction and powder low-dose,medium-dose,high-dose groups(1.95,3.90,7.80 g/kg),10 in each group. In hemo-penia mice model experiment,except for the normal group,mice in other groups were intraperitoneally injected cyclophosphamide 0.1 g/kg for modeling,and intragastrically given medicines from the first day(positive drug was Compound e-jiao slurry),once a day,for 8 d. After administration,peripheral blood levels of white blood cells (WBC),red blood cells (RBC),hemoglobin (HG),hematocrit(HCT),platelet(PLT)and bone marrow nucleated cells(BMNC)numbers in femur of mice were determined. In osteoporosis mice model experiment,except for the normal group,mice in other groups were intragastrically given retinoic acid 0.105 g/kg for modeling,and intragastrically given relevant medicines from the first day(positive drug was Gusongbao capsule), once a day,for 30 d. After administration,alkaline phosphatase(ALP)level,organ(thymus,liver,spleen,ovary)coefficients, dry masses and degreasing,ash quality of the femur,and contents of femur calcium and phosphorus in serum of mice were detect-ed. RESULTS:In hemopenia mice model experiment,compared with normal group,WBC,RBC,HG,HCT,PLT levels in pe-ripheral blood and BMNC numbers in femur of mice in model group were decreased (P0.05). Compared with model group,ALP level in serum and dry mass,phosphorus content of femur of mice in VTC water decoction high-dose group were significantly in-creased (P<0.05);ALP level in serum and dry mass,phosphorus content of femur of mice in VTC,powder medium-dose, high-dose group,the ash mass,ash mass/dry mass of degreasing,contents of femur calcium and phosphorus in VTC powder medi-um-dose group were significantly increased(P<0.05). CONCLUSIONS:VTC water decoction shows better preventive and thera-peutic effect on hemopenia mice;while VTC powder shows better preventive and therapeutic effect on osteoporosis mice.

Correlation studies on tuberculosis patient′s self-efficacy and coping style / 中国实用护理杂志

Objective To describe the status of the self-efficacy and coping styles of tuberculosis patients,analysis the relationship between self-efficacy and coping styles. Methods With the method of convenient sampling, used the general information questionnaire self-designed, General Self-Efficacy Scale (GSES) and Medical Coping Modes Questionnaire (MCMQ) to take questionnaire survey on 132 patients with pulmonary tuberculosis.Applicated SPSS19.0 software for the statistics, and analyzed the relationship between self-efficacy and coping styles on patients with pulmonary tuberculosis. Results The mean score of self-efficacy in 132 pulmonary tuberculosis patients was 24.90 ± 7.04,which was in the medium to low level. The proportion of mediun,lower,higher level of self-efficacy was 44.70% (59/132), 39.40%(52/132),15.90%(21/132).Scores of the coping styles on face,avoid, yield were 21.16±9.23,16.55± 2.83,9.68 ± 4.12.Patients self-efficacy level was positively correlated with face coping style (r=0.242, P<0.05), avoid coping style (r=0.150, P<0.05),and negatively correlated with the yield response (r=-0.008, P<0.05). Conclusions The general self-efficacy level of tuberculosis patients is at a lower level. Tuberculosis patients should have more choice to handle it.

Value of texture feature analysis of MRI dynamic contrast enhancement in diagnosis of benign and malignant breast nodules / 中国医学影像技术

Objective To assess the diagnostic value of texture analysis of MRI in differential dignosis of benign and ma lignant breast nodules.Methods The MRI data of 78 patients (80 breast nodules) identified by surgical pathology were retrospectively studied.Sixty-three texture parameters were obtained from each nodule.ROC curve of texture parameters in differential diagnosis of benign and malignant breast nodules were performed.Results In all of the 63 texture parameters,the run length nonuniformity (RLN) had the highest AUC value (0.836) and accuracy,the diagnostic sensitivity,specificity,accuracy,positive predictive value and negative predictive value in differentiation of breast nodules were 82.93％ (34/41),94.87％ (37/39),88.75％ (71/80),94.44％ (34/36) and 84.09％ (37/44).The sensitivity,specificity,accuracy,positive predictive value,and negative predictive value of breast imaging reporting and data system (BIRADS) were 95.12％ (39/41),87.18％ (34/39),91.25％ (73/80),88.63％ (39/44),and 94.44％ (34/36).The difference of diagnostic accuracy between texture parameter and BI-RADS had no statistical significance (P =0.11).BIRADS combined texture parameter improved specificity significantly (P＜0.001).Conclusion The texture analysis could be complementary to improve the accuracy of BI-RADS-MRI in breast nodules.

Study on effect of curcumin in inhibiting proliferation of multidrug resistant liver cancer HepG2/ADM cells and its mechanism / 重庆医学

Objective To observe the inhibitory effect of curcumin on the proliferation of multidrug resistance liver cancer line HepG2/ADM cells and to explore its mechanisms.Methods HepG2/ADM cells were prepared and cultured in vitro,and treated by different concentrations (5,10,20,40 μmol/L) of curcumin for 24,48,72 h respectively.The effect of curcumin on proliferation of HepG2/ADM cells was measured by CCK-8 reagent;the concentration of intracellular rhodamine-123 (Rh-123) and adriamycin (ADM) were determined by flow cytometry;the level change of intracellular mdr-1 mRNA in each group was determined by RT-PCR,the P-gp protein level was detected by Western blot.Results Compared with the blank control and DMSO group,curucmin had more obvious inhibitory effect on HepG2/ADM cells proliferation (P＜0.05),and could more remarkably inhibit the intracellular Rh-123 excretion(P＜0.05).The RT-PCR and Western blot results showed that curcumm more significantly decrease the mdr-1 mRNA and P-gp protein levels in dose-time dependent manner (P＜0.05).Conclusion Curcumin could significantly inhibit the proliferation of multidrug-resistant HepG2/ADM cells,and its mechanism may be related with inhibiting mdr-1 gene expression and its encoded P-gp protein level,which are closely related with MDR.

Application of allergen specific immunotherapy in children with allergic diseases / 国际儿科学杂志

Allergen specific immunotherapy(AIT) is a desensitization treatment for IgE-mediated allergic diseases.Combined with pharmacological therapy,AIT is used to treat allergic asthma,allergic rhinitis and atopic dermatitis caused by allergens that cannot be avoided.AIT is the only treatment modality that can alter the natural course of allergic disease.However,because of the lack of studies,there still exists controversy about the application of immunotherapy in children with allergic diseases.The mechanism ofAIT,its clinical efficacy and safety in children with allergic diseases are reviewed.

Optimization of Water Extraction Technology for Shenqi Yiqi Capsules with Multi-index Comprehensive Score-orthogonal Test / 中国药房

OBJECTIVE:To optimize water extraction technology of Shenqi yiqi capsules. METHODS:The water extraction technology of Shenqi yiqi capsules was optimized by L9(34)orthogonal test with amount of water added,decoction times and decoc-tion time as factors using the extraction rate of astragaloside,calyosin-7-O-β-D-glycoside,icariin and bergenin as index,and the verification test was made. RESULTS:The best extraction technology was as follows as 8-fold water,decocting for 3 times,2 h each time;comprehensive score of 3 batches of samples in validation test were 98.95,99.71 and 99.98(RSD<2%,n=3). CON-CLUSIONS:The water extraction technology optimized by multi-index comprehensive score-orthogonal test is stable,scientific and reasonable.

Comparison study on sampling methods of Oncomelania hupensis snail sur-vey in marshland schistosomiasis epidemic areas in China / 中国血吸虫病防治杂志

Objective To optimize and simplify the survey method of Oncomelania hupensis snail in marshland endemic re?gion of schistosomiasis and increase the precision,efficiency and economy of the snail survey. Methods A quadrate experimen?tal field was selected as the subject of 50 m×50 m size in Chayegang marshland near Henghu farm in the Poyang Lake region and a whole?covered method was adopted to survey the snails. The simple random sampling,systematic sampling and stratified ran?dom sampling methods were applied to calculate the minimum sample size,relative sampling error and absolute sampling error. Results The minimum sample sizes of the simple random sampling,systematic sampling and stratified random sampling meth?ods were 300,300 and 225,respectively. The relative sampling errors of three methods were all less than 15%. The absolute sampling errors were 0.221 7,0.302 4 and 0.047 8,respectively. Conclusion The spatial stratified sampling with altitude as the stratum variable is an efficient approach of lower cost and higher precision for the snail survey.

Systematic review of the effects of education on medical students’ knowledge and attitudes toward organ donation and transplantation / 中国组织工程研究

BACKGROUND:Medical stuff have significant impacts on the development of organ donation and transplantation, however, medical students present insufficient knowledge and ambivalent attitude towards organ donation and transplantation. OBJECTIVE:To perform a systemic review on the effectiveness of educational program intervention on medical students’ knowledge and attitudes toward organ donation and transplantation. METHODS:PubMed, WILEY ONLINE LIBRARY, ProQuest Health, Medline, EMbase, CNKI, Wanfang and VIP databases were ful y searched for the published literature of medical students receiving an educational program intervention on organ donation and transplantation. According to the inclusion criteria, randomized control ed trials and quasi-experimental study were evaluated with the standard of Joanna Briggs Institute. The GRADE (Grading of Recommendations Assessment, Development and Evaluation) framework was used to guide the assessment of the methodological quality of the studies. The outcomes were mainly medical students’ knowledge and attitudes toward organ donation and transplantation. RESULTS AND CONCLUSION:Eight studies from three countries were summarized. The methods were employed in two studies of randomized control ed trials and quasi-experiment received a high quality rating, five studies received middle quality rating and one study received low quality rating. Educational program increased the knowledge in six studies, and changed the attitudes in six studies. Educational program intervention is a significant strategy to increase medical student’s knowledge about organ donation and transplantation. The majority of involved study are quasi-experiment study, so more high-quality randomized control ed studies are needed.

Reflection on the training mode of PhD candidates majoring in medical science / 国际中医中药杂志

Medical education of PhD candidates is of great importance to medical personnel training in our country. With the development of society, the progress of science and technology and renewal of knowledge, cultivation of innovative talents and improvement of graduate education quality is priority nowadays. This paper reflects on the progress we’ve made on medical education of PhD candidates. Though efforts have been made to improve the innovative ability of talents in our country gradually, we still need to be more innovative, more consistent and more scientific in the future. According to the state Degree Requirements and our goal of talent training, we should combine courses with scientific research systematically, pay more attention to scientific methodology and academic training, finally form a better training mode of PhD candidates majoring in medical science.

Clinical research of external application of sihuang-honey for aseptic inflammation after total knee arthroplasty / 国际中医中药杂志

ObjectiveTo observe the clinical efficacy of external application of sihuang-honey for aseptic inflammation after total knee arthroplasty.Methods 30 cases of total knee arthroplasty patients were recruited into two groups,of 15 cases in each group.The treatment group was treated with sihuang-honey (made of rhubarb,scutellaria,phellodendron,coptis according to the same proportion of powder with honey ),which was placed on a sterile plastic sheet,0.5 cm thickness.The sheet was applied on the knee of the highest skin temperature for 6 hours,removed it for 2 hours interval,and reapplied on it,for altogether 24 hours before changing another sheet.The control group were treated by continued covering ice on the knee every day,with the ice completely surrounded the knee.Both groups were treated for 2 weeks.Clinical efficacy was observed before the treatment,and 3 d,7 d,14 d after the treatment.ResultsAfter the treatment,compared with the control group,the treatment group was effective in terms of VAS score (4.12± 1.08) points in 3d,(3.34±0.61)points in the first 7 days; skin temperature (37.56±0.40) ℃,activity (98.56±3.24) ° in 3d,and the difference was statistically significant (P＜0.05).The degree of swelling in 3 d (5.80±2.00) ％,7 d (9.33±2.55)％,14 d (12.13 ± 2.70)％ in the treatment group were better than the control group (P＜0.05); The rate of pain relief of treatment group was 93.33％ and satisfaction was 66.67％,although better than 86.67％ and 46.67％ of the control group,the difference was not statistically significant (x2 were 1.33,3.04,P＞0.05).Conclusion Sihuang-honey was effective for the treatment of aseptic inflammation after the surgery of total knee arthroplasty.

Comparison of epidemiological situation in chronic kidney disease between urban and rural areas in Guangxi province / 中华肾脏病杂志

ObjectiveTo investigate and compare the prevalence,awareness and risk factors of chronic kidney disease (CKD) between urban and rural population in Guangxi province in order to provide information for prevention and treatment of CKD.MethodsBy a stratified multistage random sampling method,18 to 74 years old residents in Guangxi province were surveyed. They wereevaluatedbyquestionnaire,urinaryalbumin/creatinineratio, hematuria (microscopic examination of centrifuged urine sediment),kidney B-mode ultrasound,and abnormal results were reviewed 3 months later.Estimated glomerular filtration rate(eGFR) was calculated with the simplified MDRD equation modified by a Chinese coefficient.The risk factors associated with CKD were also investigated.Results There were no significant differences between urban and rural residents in the prevalence of albuminuria (5.22％ vs 5.47％) and hematuria (1.07％ vs 1.11％)(all P＞0.05).The prevalence of renal lithiasis in rural residents was significantly higher than that in the town(10.54％ vs 6.95％)(P＜0.05).The decreased renal function between urban and rural residents(3.87％ vs 4.04％,P＞0.05) had no significant difference.The prevalence of CKD was 9.58％ in urban and 9.42％ in rural(P＞0.05).The prevalence of albuminuria according to the age distribution was different between urban and rural,which increased along with the age in urban but showed two peaks(30-40 years old and 60-74 years old) in rural.Based on logistic regression analysis,the risk factors for albuminuria were diabetes,hyperuricemia,the history of cardiovascular disease,chronic tonsillitis and HBsAg positive.The risk factors for kidney function decline were age,hyperuricemia,hypertension,diabetes,renal lithiasis and history of cardiovascular disease.The awareness rate of CKD in urban was significantly higher than that in rural (14.45％ vs 6.27％,P＜0.05).Conclusions The prevalenceof CKD has no significant difference between urban and rural in Guangxi province.The awareness rate of CKD in urban is significantly higher than that in rural.It is needed to enhance the prevention and treatment of CKD in rural.

Evidence of pterygial fibrous tissues derived from mesenchymal stem cells / 中国组织工程研究

BACKGROUND: Pterygium is associated with local chronic inflammatory responses and chronic stimulation from external factors such as. sunlight and wind dust. Presently. there are various theories concerning the onset mechanism of pterygium, but these theories are not generally accepted.OBJECTIVE: To investigate histopathological chractefistics of pterygium and analyze the mulfipotent stem cell effects on the onset of pterygium DESIGN: An open experiment.SETTING: Zhongshan Ophthalmic Center of Sun Yat-sen University.MATERIALS: Experiments were performed at the State Key Laboratory of Ophthalmology, Sun Yat-sen University from September 2006 to January 2007. 218 pterygial paraffin specimens following clinical and pathological diagnosis were obtained from Pathology Lab of Zhongshan Ophthalmic Center, Sun Yat-sea University.METHODS: Pterygial specimens harvested from clinical operations received morphology, immunohistochemistry and immunofluorescence under a confocal microscope.MAIN OUTCOME MEASURES: Morphology of ptorygium and expressions of CD34, vimentin (VIM), smooth muscle acfin (SMA), S- 100 in pterygium.RESULTS: Changes in morphology: Fibroplasia and neovascularization were the main changes in pterygium. Fibroplasia wasdiverse in different regions, and two main phenomena were observed. First, the tissues arranged tightly like the scleral fiber.Secondly, in some loose region, some of spindle-shape, polygonal, asteroid fibroblast-like cells, arranging loosely, could be seen only. No apparent collagen fibers were identified between them. Immunohistochemistry were positive for CD34 in some region where the fibroblast actively proliferated, whereas fibrocytes in mature fibrous tissue were negatively stained.Immunohistochemisu'y was positive for VIM in a large fraction of fibrocytes, vascular endothelial cells, vessel wall and perithelial cells. SMA staining was positive in basophilous small blocks, spindle or irregular cell cluster. Of the 218 cases, 56 cases had smooth muscle. S-100 staining demonstrated that neurofilament protein and adipocytes were positive. Of the 218 cases, 44 cases had adipose tissue, Immunofluorescence showed that proliferative active cells were positive and stained green under a confocal microscope.CONCLUSION: The fibrous tissues in pterygium originate from mesenchymal stem cells, and can differentiate into smooth muscle and adipose tissue.

Biological characteristics of bone marrow-derived mesenchymal stem cells cultured by density gradient centrifugation combined with adherence in adult rats / 中国组织工程研究

BACKGROUND: It is important to study the methods of culturing bone marrow-derived mesenchymal stem cells (MSCs) to obtain a great amount of high purity MSCs for applying ocular tissues constructed by tissue engineering technique to treat eye diseases.OBJECTTVE: To separate and culture in vitro MSCs from bone marrow of the adult rats by density gradient centrifugation combined with adherence culture, and observe the growing characteristics and the possibility of mass multiplication.DESIGN: A completely randomized grouping design/repetitive measuring experiment.SETTING: Pathological laboratory, Zhongshan Ophthalmic Center, Sun Yat-sen University.MATERIALS: Four six-week-old SD rats about 250 g, grade Ⅱ of cleaning, were provided by the Animal Center of Sun Yat-sen University [certificate number: SCSK(Yue)2004/0011], about 250 g each rat and there was no limit to the sex. The main reagents and instruments included low sugar Dulbecco modified Eagle culture medium (DMEM/F12, American Gibco Corporation), trypsin (fetal bovine serum (FBS, Hangzhou Sijiqing Bio-Engineering Material Research Institute), American Gibco Corporation), disodium edetate, lymphocyte separating medium, fibronectin, CD44, CD34, CD31 monoclonal antibodies, two-step-method kit for immunohistochemistry (Beijing Zhong shan Biotechnology Corporation).METHODS: This experiment was conducted at the Key laboratory of Ophthalmology (Sun Yat-sen University), Ministry of ethanol (750 g/L) for 10 minutes. Under aseptic condition, the medullaris cavitas was exposed, the syringe containing application m edium was directly punctured into the femoral cavity, the cells in the medullaris cavitas were washed out with the culture medium containing heparin and taken as the cell suspension. The bone marrow-derived MSCs were separated and purified by density gradient centrifugation combined with adherence culture, and the growing conditions of the wells. When the cells had generally connected with each other, they were fixed with methanol or dimethy ketone in situ for 10 minutes, and then hematoxylin eosin (HE) staining or immunohistochemical staining. Antibodies against fiinoculated to 96-well culture plate by a cell density of 4.25×107/L, with 200 μL every well; then put the culture plate into culture box. Then from the next day to the sixth day, 5 g/L MTT solution was added into two rows (20 μL every well) every day, continuously cultured for 4 hours, then the supernatant was removed, and 200 μL DMSO was added to each well, agitated for 5 minutes, then detected the absorbance (A) values at 570 nm wave length, and a growth curve was drawn.branes were clear and the cell bodies were lucent. Being cultured for 2 days, there appeared adherent cells. 3 days later,most of the adherent cells extended and appeared to be in polygon, star or long-shuttle shapes. 4 days later, the cells showed to be in division growth stage; and about 12 days later, the cell clones were connected to each other, appearing curve of subcultured MSCs was in S shape. Cells began growing fast on the 2nd day after being passaged, and they entered the growth period for 3-4 days followed by saturation period, and then cells stopped growing.CONCLUSION: It is a simple and practical method to separate MSCs from the bone marrow of adult rats by means of density gradient centrifugation and adherence. MSCs can greatly proliferate in vitro and offer seed cells for the application of tissue engineering technique to treat eye diseases.