OBJECTIVETo assess the long-term therapeutic effect of Spatholobus Composite (SSC) for treatment of aplastic anemia (AA).

METHODSA follow-up study was carried on 110 AA patients, who were successfully treated by SSC and testosterone propionate from 1976 to 1994, by correspondence, the information regarding blood cell count, condition of health and labor, as well as incidence of relapse and complications were collected, and the data from 66 patients response to the correspondence were compared with those of 36 healthy persons and 13 recently visited AA patients.

RESULTSIn the 66 AA patients, the average blood cell counts were WBC: 4.7 x 10(9)/L, hemoglobin 121.2 g/L, platelet 109.5 x 10(9)/L, neutrophilic granulocytes 2.5 x 10(9)/L, and their mean hemopoietic tissue in bone marrow 72.7%, all were significantly different to those in the recent visitors (P < 0.01); SF-36 scoring showed insignificant difference between the patients and the healthy person (P > 0.05). Out of the 66 patients, 17 of the 30 school-aged (56.7%) have schooling; the family consummative rate was 87.9% (58/66), including 24 got married after curing, 26 had already married before illness and 8 at school students; 43 (65.2%) have got job; 4 died. The incidence of recurrence was 10.6% (7/66), and the complication occurred in 7.6% (5/66).

CONCLUSIONSThe AA patients could be cured by SSC and testosterone propionate, which is a remedy with safety. Cured patients could go into society harmoniously and have their own regular life and job.

Adolescent ; Adult ; Aged ; Anemia, Aplastic ; drug therapy ; Case-Control Studies ; Drugs, Chinese Herbal ; therapeutic use ; Female ; Follow-Up Studies ; Humans ; Male ; Middle Aged ; Phytotherapy ; Treatment Outcome ; Young Adult

OBJECTIVETo investigate the effect of p65 gene inhibited by siRNA on neuronic differentiation in the marrow mesenchymal stem cells (MSCs).

METHODSThe MSCs were transfected with Rn-p65-siRNA. Fasudil hydrochloride induced MSCs differentiating into neurons. The non-transfected group and negative control group (transfected with negative control siRNA marked by Cy3) were used as controls. The fluorescence expressed by transfected MSCs were observed under inverted fluorescence microscope at 24 h,48 h and 72 h after transfected with negative control siRNA. The viability of MSCs was detected by MTT at 24 h, 48 h and 72 h after transfected with Rn-p65-siRNA. The expressions of p65 mRNA and protein in MSCs were detected by RT-PCR and Western blot respectively. The expressions of p65 protein, NSE, MAP-2 and glial fibrillary acidic protein (GFAP) were detected by immunocytochemical method after transfection for 6 h.

RESULTSThe fluorescence of MSCs was mostly displayed after transfection of 72 hours and the efficiency of transfection was up to 83.3% ± 3.8%. Meanwhile, the p65 mRNA and p65 protein expressed by MSCs of transfected group were significantly decreased (P < 0.05); MTT displayed that the viability of MSCs was also significantly reduced (P < 0.05). The best efficiency of induction was observed in the transfected group. There were higher expressions of NSE and MAP-2 than the other group (P < 0.05).

CONCLUSIONThe p65 gene inhibited by siRNA can promote the marrow mesenchymal stem cells to differentiate into neurons.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine ; analogs & derivatives ; Animals ; Cell Differentiation ; Glial Fibrillary Acidic Protein ; metabolism ; Mesenchymal Stromal Cells ; cytology ; Neurons ; cytology ; RNA, Messenger ; RNA, Small Interfering ; Rats ; Transcription Factor RelA ; antagonists & inhibitors ; metabolism ; Transfection

OBJECTIVETo determine whether the skilled reaching test is an objective method for evaluating long-term neurological deficits after focal cerebral ischemia in mice.

METHODSIn a reaching box, mice were trained to reach food pellets with their left forelimb through a 0.5 cm slit for 3 weeks. Then focal cerebral ischemia was induced by occluding the right middle cerebral artery, and the percentage of success in obtaining food was observed for 4 weeks. In comparison, the neurological deficit score, the holding angle in an inclined board test, and right turns in a corner test were simultaneously performed. At the end of the experiments, brain infarcts and neuron densities were determined.

RESULTAfter focal cerebral ischemia, the percentage of success in the reaching test was reduced, the right turns in the corner test were increased, the neurological deficit score was increased, and the holding angle in the inclined board test was reduced as well. The holding angle recovered 5 d after ischemia, whereas other 3 indicators remained abnormal until 4 weeks. At the end of the experiments, the brain infarct volumes were increased, and the neuron densities in the cortex, hippocampal CA1 region and striatum were reduced in ischemic mice.

CONCLUSIONThe skill reaching test is an objective and stable method for evaluating long-term neurological deficits after focal cerebral ischemia in mice.

Animals ; Behavior, Animal ; physiology ; Brain ; pathology ; physiopathology ; Brain Ischemia ; complications ; physiopathology ; Cell Count ; Male ; Mice ; Mice, Inbred ICR ; Movement Disorders ; etiology ; physiopathology ; Neurologic Examination ; methods ; Neurons ; pathology ; Psychomotor Performance ; physiology

OBJECTIVETo study the changes of microRNA expression in cortex tissues in neonatal rats with hypoxic-ischemic brain damage (HIBD)and the possible roles of microRNA in the pathogenesis of HIBD. METHODS Rat HIBD model was prepared. The cortex tissues were obtained 14 days after the HIBD event. The microRNA expression profiles were measured using microRNA microarray. Expression of 9 microRNAs (miR-126,-26a,-674-5p,-21,-25,-290, miR-124,-125b-5p and microRNA-9a) was determined by quantitative real-time PCR.

RESULTShe results of microRNA expression profiles indicated that 27 pieces of microRNA were up-regulated more than 2 folds and 60 pieces were down-regulated more than 2 folds compared with the normal control group. The results of the 9 microRNAs detected by quantitative real-time PCR were consistent with those detected by microRNA microarray.

CONCLUSIONSHIBD rats have significant changes in microRNA expression, suggesting that microRNA expression may play important roles in the pathogenesis of HIBD.

Animals ; Animals, Newborn ; Apoptosis ; Cell Cycle ; Hypoxia-Ischemia, Brain ; etiology ; genetics ; MicroRNAs ; genetics ; physiology ; Oligonucleotide Array Sequence Analysis ; Polymerase Chain Reaction ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo study the effects of Down syndrome cellular adhesion molecule (DSCAM) on differentiation of rat marrow mesenchymal stem cells (MSCs) into neurons in vitro.

METHODSMSCs from Sprague-Dawley rats were induced into neurons by baicalin. The expression of DSCAM before and after induction was evaluated by immunocytochemical staining and Western blot assay. After knockdown of DSCAM by siRNA transfection, the differentiation rate of neurons derived from MSCs was measured.

RESULTSBefore induction, the expression of DSCAM was not detectable in MSCs. After bFGF preinduction for 24 hrs, DSCAM was slightly expressed in MSCs (1.71+/- 0.67%). The DSCAM expression increased 6 hrs after baicalin induction (15.79+/- 4.24%), reached a peak at 3 days (53.16+/- 5.94%) and then decreased gradually. The DSCAM expression 6 days after baicalin induction (28.99+/- 6.72%) was significantly lower than that at 3 days (P<0.01). However, after DSCAM-siRNA transfection, the DSCAM expression in MSCs was significantly reduced. MSCs did not express neuron-specific beta-III-tubulin before induction. After baicalin induction for 6 hrs, 3 days and 6 days, the expression of beta-III-tubulin was 1.40+/- 0.79%, 41.59+/- 3.17% and 59.11+/- 4.76% respectively. But the beta-III-tubulin expression significantly decreased 3 and 6 days after DSCAM-siRNA transfection (28.57+/- 2.91% and 43.90+/- 12.31% respectively).

CONCLUSIONSDSCAM may play an important role in MSCs differentiation into neural cells.

Animals ; Bone Marrow Cells ; cytology ; Cell Adhesion Molecules ; physiology ; Cell Differentiation ; Mesenchymal Stromal Cells ; cytology ; Neurons ; cytology ; RNA, Small Interfering ; genetics ; Rats ; Rats, Sprague-Dawley ; Transfection

Shigella flexneri is an infectious pathogen that causes dysentery to human, which remains a serious threat to public health, particularly in developing countries. In this study, the global protein expression patterns of S. flexneri during transition from exponential growth to stationary phase in vitro were analyzed by using 2-D PAGE combined with MALDI-TOF MS. In a time-course experiment with five time points, the relative abundance of 49 protein spots varied significantly. Interestingly, a putative outer membrane protein YciD (OmpW) was almost not detected in the exponential growth phase but became one of the most abundant proteins in the whole stationary-phase proteome. Some proteins regulated by the global regulator FNR were also significantly induced (such as AnsB, AspA, FrdAB, and KatG) or repressed (such as AceEF, OmpX, SodA, and SucAB) during the growth phase transition. These proteins may be the key effectors of the bacterial cell cycle or play important roles in the cellular maintenance and stress responses. Our expression profile data provide valuable information for the study of bacterial physiology and form the basis for future proteomic analyses of this pathogen.
Bacterial Proteins ; analysis ; Computational Biology ; Electrophoresis, Gel, Two-Dimensional ; Gene Expression Profiling ; methods ; Kinetics ; Peptide Mapping ; Proteome ; analysis ; Proteomics ; methods ; Shigella flexneri ; growth & development ; metabolism ; pathogenicity ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; Temperature ; Trypsin ; pharmacology ; Virulence

The effect of Fructus Mume formula and its separated prescription extract on insulin resistance in type 2 diabetic rats was investigated. The rat model of type 2 diabetes was established by feeding on a high-fat diet for 8 weeks and by subsequently intravenous injection of small doses of streptozotocin. Rats in treatment groups, including the Fructus Mume formula treatment group (FM), the cold property herbs of Fructus Mume formula treatment group (CFM), the warm property herbs of Fructus Mume formula treatment group (WFM), were administrated with Fructus Mume formula and its separated prescription extract by gavage, while the rats in diabetic model group (DM) and metformin group (MET) were given by gavage with normal saline and metformin correspondingly. The body weight before and after treatment was measured, and the oral glucose tolerance test (OGTT) and the insulin release test (IRT) were performed. The homeostasis model assessment-insulin resistance index (HOMA-IR) was calculated. The protein and mRNA expression levels of Insr, β-arrestin-2, Irs-1 and Glut-4 in the liver, skeletal muscle and fat tissues were detected by using Western blotting and RT-PCR respectively. The results demonstrated that, as compared with DM group, OGTT, IRT (0 h, 1 h) levels and HOMR-IR in treatment groups were all reduced, meanwhile their protein and mRNA expression levels of Insr, Irs-1 and Glut-4 in the liver, skeletal muscle and fat tissues were obviously increased, and their protein and mRNA expression levels of β-arrestin-2 in the liver and skeletal muscle tissues were also markedly increased. It was suggested that the Fructus Mume formula and its separated prescription extracts could effectively improve insulin resistance in type 2 diabetic rats, which might be related to the up-regulated expression of Insr, Irs-1 and Glut-4 in the liver, skeletal muscle and fat tissues, and β-arrestin-2 in the liver and skeletal muscle tissues.
Adipose Tissue ; drug effects ; metabolism ; Animals ; Arrestins ; genetics ; metabolism ; Diabetes Mellitus, Experimental ; drug therapy ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Glucose Intolerance ; drug therapy ; Glucose Transporter Type 4 ; genetics ; metabolism ; Hypoglycemic Agents ; pharmacology ; therapeutic use ; Insulin Receptor Substrate Proteins ; genetics ; metabolism ; Insulin Resistance ; Liver ; drug effects ; metabolism ; Male ; Muscle, Skeletal ; drug effects ; metabolism ; RNA, Messenger ; genetics ; metabolism ; Rats ; Rats, Wistar ; Receptor, Insulin ; genetics ; metabolism ; beta-Arrestin 2 ; beta-Arrestins

OBJECTIVETo investigate the effect of notch signaling on differentiation of rat bone marrow mesenchymal stem cells (MSCs) into neurons induced by fasudil hydrochloride.

METHODSThe experiments were divided into non-transfected group, transfected group (transfected with Rn-Notch1-siRNA), positive control group (transfected with Rn-MAPK-1 Control siRNA) and negative control group (transfected with negative control siRNA). Fasudil hydrochloride induced MSCs differentiating into neurons. The fluorescence expressed by transfected MSCs were observed under inverted fluorescence microscope. The expression of notch1 mRNA, Hes1 mRNA and MAPK1 mRNA in MSCs was detected by RT-PCR. The expression of Notch1 protein, NSE, neurofilament M (NF-M) and glial fibrillary acidic protein(GFAP)was detected by immunocytochemical method. The viability of MSCs was detected by MTT.

RESULTS(1) The fluorescence of MSCs was mostly displayed after transfection for 72 h and the efficiency of transfection was up to 91.3% +/- 4.2%. Meanwhile, the notch1 mRNA and Hes1 mRNA expressed by MSCs of transfected group were significantly decreased (P < 0.05) and MTT displayed that the viability of MSCs was also significantly reduced (P < 0.05). (2) Fasudil hydrochloride could induce MSCs differentiate into neurons and the best efficiency of induction was observed in the transfected group. There was higher expression of NSE and neurofilament-M (NF-M) than the other groups (P < 0.05).

CONCLUSIONThere may be notch1 signaling and Rho/Rho GTPase signaling synergy on differentiation of rat bone marrow stromal cell into neurons induced by fasudil hydrochloride and they jointly promote the differentiation of MSCs into neurons.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine ; analogs & derivatives ; pharmacology ; Animals ; Bone Marrow Cells ; cytology ; drug effects ; Cell Differentiation ; drug effects ; Cells, Cultured ; Mesenchymal Stromal Cells ; cytology ; drug effects ; Neurons ; cytology ; Rats ; Rats, Wistar ; Receptor, Notch1 ; metabolism ; Signal Transduction

Objective:To construct the recombinant DNA E.coli ghosts (EBGs) expressing Treponema pallidum adhesin Tp0751 (pcD/Tp0751-BG) and determine its immunocompetence in immunized mice,and provide a potential novel method for syphilis vaccine developing.Methods:The recombinant eukaryotic expression plasmid pcDNA3.1(+)/Tp0751 was constructed and loaded into empty EBGs to create pcD/Tp0751-BG.The loading rate was determined accordingly.Macrophages cell line RAW264.7 was transfected with pcD/Tp0751-BG,and the expression of recombinant Tp0751(rTp0751) protein was detected by Western blot(WB).For immuno-competence in mice,the female BALB/c mice were randomly divided into 6 groups,including three control groups,A (PBS),B (EBG),C (empty pcDNA),and experimental group D(naked pcD/Tp0751),E (pcD/Tp0751-BG) and F (pcD/Tp0751-BG+rTp0751).All the mice were immunized as indicated for three times by intramuscular injection at two weeks intervals.The levels of specific IgG in sera and SIgA in genital tract lavage fluid were measured by ELISA.Levels of lymphocyte proliferation and IFN-γ secretion in spleen cells were measured by CCK-8 Cell Counting Kit and ELISA as well,respectively.Results:The loading rate of pcD/Tp0751 to EBGs was 76.1%.WB showed that the target recombinant protein pcD/Tp0751 expressed in RAW264.7 cells was active with Tp-infected rabbit sera.The titers of specific IgG and SIgA in group D,E,F gradually increased to significantly higher level as compared to the control groups (P<0. 01),which reached its peak at wk 8 after last immunization(the titers of IgG and SIgA were 1 :102 400 and 1 :12 800 in group F,respectively). Higher levels of specific IgG and SIgA were observed in groups E and F as compared to group D after first boost (P<0. 01),with groups F higher than group E after last boost(P<0. 01). At wk 8 after the last boost,the stimulation index (SI) and levels of IFN-γ in group D,E,F were all significant higher than the control groups (P<0. 01), with group E and F higher than group D (P<0. 01),and group E higher than group F (P<0. 05). Conclusion: The recombinant DNA EBGs of T. pallidum adhesin Tp0751 (pcD/ Tp0751-BG) possesses the immunocompetence to induce not only strong mucosal and systemic humoral immune response but also systemic cellular immune response in BALB/ c mice. The heterologous boost can be more efficient than homologous boost during immunization process.

Objective To establish the three-dimensional finite element model of human lower cervical spine C3-7 motion segments after anterior cervical corpectomy and fusion (ACCF) surgery with titanium mesh and bone graft,and to analyze the stability of cervical spine and stress distribution of internal fixation devices after ACCF surgery.Methods The finite element model of cervical spine C3-7 segments after ACCF of C5 segment with titanium mesh,bone graft,plate and screw fixation was established,and C3-7 segment intact model of cervical vertebra was also constructed.The torque moment of 0.5,1.0,1.5,2.0 N · m was applied to the ACCF surgery model.The ROM,maximum stress in facet joint and stress distributions on internal fixation devices under flexion,extension,lateral bending and axial rotation movement were analyzed.Results ROM of reconstructed C5 segment increased with the torque moment increasing after ACCF surgery.In the case of 1.0 N · m tomue moment and 50 N preload,the ROM of reconstructed C5,C3-4,C6-7 and C3-7 segment was reduced by 81％,62％,58％ and 80％ compared with the intact model.The maximum stress in facet joint of reconstructed C5 segment reduced and the stress in adjacent segments significantly increased.The stress of titanium mesh was mainly distributed on the compression side of movement,and high stress was located in the roots of screws.Conclusions ACCF surgery can promote the stability of cewical spine,decrease the stress in facet joint of operation segment,and has better treatment effect on easing compression from spinal cord caused by cervical spondylotic myelopathy.The research results will provide some theoretical basis for clinical application of ACCF surgery.