BACKGROUND: Severe acute respiratory syndrome (SARS) is caused by a genetically novel coronavirus that is caused by acute infectious disease. It is not yet clear for the immunology function of SARS patients in their convalescent stage.OBJECTIVE: To study the effects on T lymphocyte, and the titer profiling of 24 T cell receptor (TCR) V β subfamilies expressions in SARS convalescent patients.DESIGN: A self-control observation.SETTING: Central Laboratory, Guangdong Provincial Hospital of Traditional Chinese Medicine.PARTICIPANTS: Seventy-six cured SARS patients who received treatment in the Second Hospital Affiliated to Guangzhou University of Traditional Chinese Medicine between January and April 2003. All the patients corresponded to "clinical diagnostic criteria of atypical pneumonia", " diagnostic criteria of severe atypical pneumonia and discharge criteria" and "clinical diagnostic criteria and discharge criteria of severe acute respiratory syndrome". The involved patients, 30 male and 46 female, averaged (32±11 (years old. Another 10 subjects who simultaneously received health examination in the same hospital, 5 male and 5 female, aged (32±7(years, were involved in the study. Informed consents of detected items were obtained from all the subjects.METHODS：①Detecting the expression of 24 T cell receptor(TCR)V β subfamilies in SARS convalescent patients：Peripheral blood(2 mL) was collected from the healthy convalescent subjects,and EDTA-K2 was used as anticoagulant．In the flow cytometry delection tubes．10 μL various fluorescein-labeled mAb，such as anti-CD3,anti-CD4，anti-CD8,anti-CD25，anti-CD28，anti-HLA-DR，anti-CD3mAb conjugated with PC5，TCR Vβ1(PE+FITC)．Vβ2(PE+FITC)。Vβ3 (FITC)，Vβ4(PE+FITC)，Vβ5．1(PE+FITC),Vβ5．2(PE),Vβ5．3(PE)，Vβ7．1(PE+FITC)，Vβ7．2(FITC),Vβ8(FITC),Vβ9 (PE)，Vβ11(PE)，Vβ12(FITC)，Vβ13．1(PE),Vβ13．2(PE),Vβ13．6(PE+FITC)，Vβ14(FITC),Vβ16(FITC),Vβ17 (PE+FITC)，Vβ18(PE)，Vβ20(FITC)，Vβ21．3(FITC),Vβ22(PE+FITC)and Vβ23(PE)，was added in special flow tubes,and then 50 μL whole blood was added．The mixed solution was incubated away from light for 15 minutes．After erythrocytolysin being added，mixed solution was washed．Finally．cell deposit was dissolved in 300 μl phosphate buffer solution (PBS)．Coulter ESP flow cytometer was used for detection．For the analysis of TCR expression,an electronic gate was set on these cells and at least 5000 events per sample were collected．Three-color cytofluodmetric analysis was performed using a Coulter ESP flow cytometer．②Detecting the T cell subset,activated T and B cells,and the percentage of Ts and Tc cells：5000 cells were collected and used to calculate the expression of T cells (CD3,CD4 and CD8),the activated T and B cells(CD3+／CD25+,CD3+/HLA-DR+ and CD3-／HLA-DR+),as well as the percentage of Ts and Tc cells by Coulter ESP flow cytometer and its software．MAIN OUTCOME MEASURES：①The change of T cell subset(CD3，CD4，and CD8)from SARS convalescent patients．②The change of activated T and B cells(CD3+／CD25+，CD3+／HLA-DR+ and CD3-／HLA-DR+)．③The percentage of Ts and Tc cells(CD8+／CD28+，CD8+/CD28-)in convalescent patients．④Analysis of the 24 TCR V β subfamilies from SARS patients in convalescence．RESULTS：All data were explored to analyze the expression profiling of 24 TCR Vβ subfamilies,the data from 74 SARSpatients and 10 healthy controls were explored to other result analysis．①The detecting results of T celI subset：The percentage of CD4+T cell mean value was lower than the reference value[(33．33±6．64)％ vs．(43±9)％，P＜0．01]．The percentage of CD8+T cell mean value was higher than the refefence value[(34．07±6．40)％ vs．(30±9)％,P＜0．01]．② The expression of activated T and B cells：Percentage of HLA-DR+ T and B cell was Increased while the percentage of CD25+ T-cell was decreased compared with reference values．In 53 out of 74 patients,the percentage of CD25+ T cells was lower than the reference value,and 64 patients had a lower percentage in CD3+/CD25+ T cells．The percentages of CD3+/HLA-DR+ and CD3-／HLA-DR+ cells were higher than the normal reference value．T cells expressing higher CD3+／HLA-DR+ were found in 36 patients，and T cells expressing higher CD3-/HLA-DR+ were found in 30 patients．③The ratios of Ts and Tc cells：The percentage of Ts cells which expressed CD8+／CD28- was increased compared with reference value [(28．75±7．31)％ vs．(15．99±5．1)％，P＜0．01],while the percentage of Tc cells which expressed CD8+／CD28+ was decreased [(5．99±3．60)％ vs．(13．2±4．1)％，P＜0．01]．Thirty-nine patients were found to possess the lower Tc cells and forty-eight patients were found to possess the higher Ts cells．The ratios of both CD4+ and CD8+ T cells were in the normal reference value．④24 TCR Vβ subfamilies expressions in T cells：It was noteworthy that Vβ14 had a highest percentage in all 24 Subfamilies,and followed by Vβ 5．3,and Vβ 23 in the convalescent patients．The percentage of Vβ 14 was the highest in the normal controls,which was consistent with the results of SARS patients．But the other subfamilies expression patterns were different．There were significant differences between Vβ1,Vβ5．2,Vβ5．3,Vβ7．2，Vβ9，Vβ11,Vβ13．1,Vv13．2，Vβ17,Vβ18，Vβ22 and Vβ23．In the convalescent period，each TCR Vβ expression of SARS patients was higher than that of controls(P＜0．05-0．01)．CONCLUSION:In SARS convalescent patients,the increased CD8+CD28- T cell may elevate CD8+ T cell number;Meanwhile.the reduced CD3+ and CD4+ T cell number may be corresponding to the increased Ts cell number．For some inhibiting factor secreted by Ts cell was also increased．The usage pattern of 24 TCR Vβ subfamilies in SARS patients is different from that of control group．The increase of percentage of CD3+／HLA-DR+ and CD3-／HLA-DR+ T cell may be related to the late response of activated T and B cells.

This study examined the effect of P85 (a pluronic block copolymer) and microbubble (MB) ultrasound contrast agents under ultrasound irradiation on gene transfection and expression. The pEGFP plasmids that can encode enhanced green fluorescent protein (pEGFP) served as a report gene and were mixed with different concentrations of MB/0.05% (w/v) P85. Then the plasmids were transfected into human hepatoma G2 (HepG2) cells. The HepG2 cells treated with MB/P85 or without treatment were exposed to ultrasound (US parameters: 1 MHz, 1.0 W/cm(2), 20 s, 20% duty cycle). Twenty-four hours later, the transfection efficiency was assessed by fluorescence microscopy and fluorescence activated cell sorting (FACS) analysis. The cell viability was evaluated by Trypan blue exclusion test. The results showed that the gene transfection efficiency in HepG2 cells under ultrasound irradiation was significantly higher than that without ultrasound irradiation. HepG2 cells in the MB or P85 group in the absence of ultrasound expressed less amount of green fluorescent protein. The expression efficiency reached (22.14±3.06)% and the survival rate was as high as (55.73±3.32)% in the 30% MB plus P85 group. It was concluded that MB and P85 in the presence of ultrasound can enhance gene transfection and expression.

Objective To detect the strain features of regional wall abnormalities in patients with coronary artery disease during each diastolic period, and its possibility to evaluate regional left ventricular cardiocyte viability and diastolic function.Methods 54 patients with anterior myocardial infarction (MI group) and 78 normal subjects (NOR group) underwent Doppler tissue imaging, which were performed in 2-chamber-view by strain curves synchronously.Results In the NOR group, strain value of 66 cases (84.62%) showed an gradually increasing negative value from the apex to base to middle of left ventricle, while in the MI group, there were 9 cases (16.67%) with such a trend. IR phase: in the NOR group, 564 segments (90.38%) were upward wave bands, but in the MI group, there were 123 segments (28.47%) having such waves ( P<0.05). RF phase: in the NOR group, 576 segments (92.31%) were upward and steep wave bands, but in the MI group, the number of upward waves were obviously less (102 segments,23.61%) ( P<0.01). SF period: compared with the NOR group, which was horizontal, the MI group had upward wave bands ( P<0.05).Conclusion Regional myocardial ischemia and infarction can cause significant regional diastolic wall abnormalities of strain value in active diastolic phase. Regional diastolic wall motion abnormalities can be evaluated quantitatively and synchronously with high sensitivity by strain curve which has the potential value in cardiocyte viability and diastolic function.

The marine biological source of mineral drugs recorded in Chinese Pharmacopoeia (2010 version) mainly including pearl, nacre, clam shell, common oyster shell, ark shell, cuttle bone, and sea-ear shell are widely used in clinical. Calcium carbonate and a small amount of protein are the main components in this type of drugs. In this paper, a systematical and comparable study were carried out by determination of calcium carbonate by EDTA titration method, the crystal of calcium carbonate by X-Ray powder diffraction and the total amino acids (TAAs) of the hydrolyzed samples by ultraviolet spectrophotometry method. As a result, the crystal structure is calcite for common oyster shell, mixture of calcite and aragonite for nacre and sea-ear shell, aragonite for the other drugs. The content of calcium carbonate ranged from 86% to 96%. Cuttle bone has the highest amount of TAAs among the seven drugs which reached 1.7% while clam shell has the lowest content of 0.16% on average. In conclusion, an effective method was developed for the quality control of marine mineral drugs by comprehensive analysis of calcium carbonate and TAAs in the seven marine mineral drugs.
Amino Acids ; analysis ; chemistry ; Animal Shells ; chemistry ; Animals ; Calcium Carbonate ; analysis ; chemistry ; Crystallization ; Edetic Acid ; chemistry ; Mollusca ; chemistry ; classification ; Pharmaceutical Preparations ; analysis ; chemistry ; standards ; Quality Control ; Reproducibility of Results ; Seawater ; Species Specificity ; Spectrophotometry, Ultraviolet ; X-Ray Diffraction

OBJECTIVEThe present study was undertaken to design antisense oligodeoxynucleotides (AS-ODNs) of glial glutamate transporter-la (GLT-1a) and to evaluate the effectiveness of the designed AS-ODNs on the expression of GLT-1a.

METHODSFive sequences of GLT-1a AS-ODNs were designed according to the C terminus specific sequences of GLT-1a mRNA using antisense design software of IDT Com- pany. Western blot analysis was used to evaluate the inhibition effects of the five GLT-1a AS-ODNs on the expression of GLT-la.

RESULTSThe sequence of GLT-1a AS-ODNs with sequence of 5'-GGTTCTTCCTCAACACTGCA-3' could specifically inhibit the expression of GLT-1a in the hippocampal CA1 subfield of rats, while it had no effect on the expression of GLT-1b. This sequence showed similar inhibition on the expression of GLT-la in sham and ceftriaxone (Cef)-treated rats. It could also significantly inhibit the cerebral ischemic preconditioning (CIP)-induced up-regulation in the expression of GLT-1a. The magnitude of the inhibition in sham, Cef- or CIP-treated rats was similar by more than 60%.

CONCLUSIONFrom the designed five sequences of GLT-1a AS-ODNs, we obtained an effective sequence which can specifically inhibit the expression of GLT-1a.

Animals ; CA1 Region, Hippocampal ; metabolism ; Excitatory Amino Acid Transporter 2 ; antagonists & inhibitors ; metabolism ; Ischemic Preconditioning ; Oligonucleotides, Antisense ; genetics ; RNA, Messenger ; Rats ; Up-Regulation

Objective To investigate the role of motility of Oddi's sphincter in pigment gallbladder stone formation in the model of guinea pigs.Methods Thirty-four general adult male Hartley guinea pigs were divided randomly into two groups.Control group (10 guinea pigs)and pigment stone groups (24 guinea pigs,which were divided into 4 subgroups with 6 guinea pigs each according to the time they were sacrificed),fed with pigment lithogenic diet.After 3,6,9 and 12 weeks later,Oddi's sphincter manometry and myoelectric activity record were obtained by multifunctional physiograph at each stage.Results The incidence of pigment gallstone formation was 0,0,16.7％ and 66.7％ in 3w,6w,9w and 12w group,respectively.The frequency of myoelectric activity decreased apparently in the 3w group(P ＜ 0.05).The amplitude of myoelectric activity had the tendency of decreasing but not statistically significant.The motility frequency of Oddi's sphincter decreased greatly in the 9w group (P ＜ 0.05).The basal pressure of Oddi's sphincter and common bile duct increased markedly in the 12w group [from (25 ± 8) mmHg (1 mmHg =0.133 kPa) to (41 ± 12) mmHg and from (22 ± 8) mmHg to (39 ± 12) mmHg,P ＜ 0.05)].Conclusions Pigment lithogenic diet may induce dysfunction of Oddi's sphincter.The disturbance of Oddi's sphincter motility may play a role in pigment gallstone formation.

To prepare the RNAse-resistant virus particles containing the partial gene fragments of avian influenza virus H5N1 for use as RNA standard and control in RNA virus detection, the genes coding the coat protein and maturase of E.coli bacteriophage MS2 were amplified by PCR and then cloned into prokaryotic expression vector pET32a to construct the intermediate vector pET32a-MS2. In addition, the gene sequences coding hemagglutinin (HA), neuraminidase(NA) and M protein of the H5N1 virus were also cloned separately to the down-stream of plasmid pET32a-MS2, thus constructing the prokaryotic expression vectors pET32a-NS2-HA, pET32a-MS2-NA and pET32a-MS2-M. These recombinant plasmids were then transformed separately to E.coli BL21(DE3) with induction by IPTG. to express the virus-like particles. The virus-like particles observed under electron microscopy were identified by RT-PCR ,while their stability was confirmed by real-time RT-PCR. In this way, the virus-like particles were successively constructed and identified through PCR amplification, enzymolysis identification and sequencing analysis. These virus-like particles observed under electron microscopy appeared to be circular in shape with a diameter of about 50 nm. Their stability was proved to be rather good. From these observations, it is apparent that these virus-like particles can be used as RNA standard and quality control in the detection of avian influenza virus H5N1.

AlM: To investigate the relationship between the anterior ischemic optic neuropathy ( AlON ) and the carotid artery change using doppler ultrasound.METHODS:Fifty-four cases of AlON patients and 54 cases of healthy control were observed, atherosclerotic spots were detected by the application of color ultrasound.RESULTS:ln AlON group of 54 patients, 38 cases appeared carotid atherosclerosis, accounting for 70%. The number of cases with hard plaque, soft plaque and mixed plaques were 18, 13, and 7 respectively, accounting for 33%, 24% and 13%. ln the control group, 20 cases were detected atherosclerotic change, accounting for 37%. And the number of cases with hard plaque, soft plaque and mixed plaques were 12, 5 and 3 respectively, accounting for 22%, 9%, 6%. Significant stenosis and velocity change were showed in neither AlON group nor control group. Compared with the control group, AlON group had more cases of atherosclerotic plaque, the difference was statistically significant (χ2=12. 836, P=0. 005)CONCLUSlON: The incidence of AlON is correlated with carotid atherosclerosis, and carotid ultrasonography is significantly valuable for AlON etiology and diagnosis.

To explore a new method for identification of Mongolian patent medicine (MPM) by PCR amplification of specific alleles. Eight kinds of MPM were used to study the identification of "Digeda" raw materials. The total DNA of Lomatogonium rotatum and Corydalis bungeana samples were extracted through modified CTAB method, psbA-trnH sequence was amplified by PCR and sequenced directionally. Specific primer was designed. The DNA of 8 kinds of MPM also was extracted and purified by the commercial DNA purification kits. The rbcL and two pair of specific primers sequences were amplified. The specific amplified products were sequenced in forward directions. All specific sequences were aligned and were analyzed. The results indicated that L rotatum can be identified by specific primers from Digeda-4 Tang, Digeda-8 San, Digeda-4 San, and C. bungeana medicinal materials can be identified by specific primers from Li Dan Ba Wei San, Yi He Ha Ri-12 and A Ga Ri-35. PCR amplification of specific alleles can stably and accurately distinguish raw medicinal materials in MPM.
Alleles ; DNA Primers ; genetics ; DNA, Plant ; genetics ; Medicine, Mongolian Traditional ; Molecular Sequence Data ; Plants, Medicinal ; classification ; genetics ; Polymerase Chain Reaction ; methods

ObjectiveTo investigate the difference of pathogenicity between the two genotypes of human metapneumovirus(hMPV) for the further research.MethodsAt various time after hMPV infection in BALB/c mice,viral titers of lung tissue were measured by real-time RT-PCR,pathology was assessed by a histopathological scoring system,airway responsiveness was assayed by animal lung function monitoring equipment.Pathogenicity was then measured by detailed evaluation through the results above.Results There is no significant difference in weight of mice between control group and experimental group through dynamic monitoring ; though the difference was exists in airway responsiveness and pathological changes of mice between control group and experimental group,the differences were not statistically in airway reaction,pathological changes and virus drops among the three groups of experimental group.ConclusionThere is no difference in pathogenicity between the two subtypes of hMPV in infection of BALB/c mice,viral genotype do not appear to be associated with pathogenicity.