Enterovirus A, Human ; pathogenicity ; Enterovirus Infections ; complications ; Humans ; Pulmonary Edema ; etiology ; virology

Objective To establish a method for determining cinnamaldehyde in Zegui Longshuang Capsules. Methods A C18 column was used with Acetonitrile-Water (30∶70) as the mobile phase,and the detection wavelength was at 290 nm. Results The calibration curve of cinnamaldehyde was linear from 0.010 ?g to 0.507 ?g,and r=0.999 9. The average recovery was 98.55 %with RSD being 2.06 %. Conclusion This method is proved to be accurate and reliable,and can be used to control the quality of Zegui Longshuang Capsules.

Objective To establish the murine congenital infection model by MCMV and observe the pathological changes and infection status of brain tissue.Methods After anesthesia,mice who were pregnant 11-13.5 days (E11-13.5 d) were intra-amniotic injected one uterus by one with virus (MCMV K181 suspension,1 μl,1×103 PFU).The control group of the same period was intra-anmiotic injected with culture medium DMEM (1 μl).Carefully reset the uteruses and close the abdomen.After 5 days of separated feeding,kill the pregnant mice,take the fetus out of the uterus,anesthetize and kill them.Make frozen sections of these fetal brains.Some sections were stained using conventional HE method,to observe the pathological changes under the light microscope.Detect MCMV early antigen in the brain tissue by immunohistochemistry staining and immunofluorescence assay.Results The survival rates of the infected group were 71.9％.Compared with the control group,intra-amniotic inoculation of MCMV does not affect the rate of fetal survival,fetal absorption,fetal death and the average weight of the heads,but decrease their average weight of the bodies.The pathological changes are found in the brain tissue of the mouse in the infection group.Through enzyme immunohistochemistry assay,there are many MCMV infected cells in brain-ventricular zone,brain subependymal zone,cerebral cortex and hippocampus area in the infection group.Similar findings were observed by immunofluorescence method.Conclusion By intra-amniotic injection of MCMV suspension,murine model of MCMV congenital infection can be successfully established.This model could be used to study the mechanisms of encephalodysplasia caused by congenital CMV infection in vivo.

Amphotericin B ; administration & dosage ; therapeutic use ; Antifungal Agents ; administration & dosage ; therapeutic use ; Biomarkers ; blood ; Child ; Cough ; diagnosis ; drug therapy ; etiology ; Cryptococcosis ; Fever ; diagnosis ; drug therapy ; etiology ; Fluconazole ; administration & dosage ; therapeutic use ; Humans ; Lung ; diagnostic imaging ; pathology ; Lung Diseases, Fungal ; complications ; diagnosis ; drug therapy ; Male ; Radiography, Thoracic ; Tomography, X-Ray Computed

Objective To investigate the nature of Th17 cells in murine cytomegalovirus(MCMV)infection during the acute stage,we characterized the frequency of IL-17A-producing CD4 T cells and the level of Th17 cytokines,IL-17A,in MCMV-infected mice.Methods BALB/c mice were randomly divided into two groups.One was infected with MCMV Smith for establishing disseminative infection; the other was sham-inoculated control.On day 3,7,14 and 28 of the experiment,three mice of each group were randomly chosen to be killed separately.Real-time PCR was used to detect MCMV loads in organs of MCMV-infected mice,the pathology of spleen was observed by HE staining.The frequency of CD4+IL-17A+ T cells in total splenocytes of mice was detected by flow cytometry.The level of IL-17A in culture supernatants of splenocytes was measured by double antibody sandwich ELISA.Results MCMV loads in salivary gland reached the peak on day 14 after MCMV infection,the most severe spleen injury was also shown on day 14,the frequencies of CD4+IL-17A+ T cells in total splenocytes increased significantly( all P＜0.01 ) in MCMV-infected mice than those in controls,and reached the peak on day 14 ( 1.14％ ±0.09％ vs 0.19％ ±0.04％,t =17.551,P=0.000).The levels of MCMV-specific IL-17A in culture supernatants were increased dramatically in MCMV-infected mice than those in controls on day 14 [ (81.98± 12.37) pg/ml vs (44.96±8.44)pg/ml,t=4.281,P=0.006].In MCMV-infected mice,correlation was positive between the levels of MCMV-specific IL-17A in culture supernatants and MCMV loads in salivary gland tissues (r=0.54,P＜0.05 ),the levels of IL-17 A in culture supernatants were higher in more severe spleen injury.Conclusion Thl7 cells and IL-17A were involved in the immunity response during acute MCMV infection.They may correlate with the persistence of MCMV and the pathology of spleen in infected mice.

Objective To explore the correlation between the expression of IL-17A and the degree of spleen damage in acute mouse cytomegalovirus(MCMV) disseminated infection in vivo and to understand the mechanism about how IL-17A involved in the pathological damage of the spleen in MCMV infection.Methods An acute disseminated MCMV infection model was established in mice.BALB/c mice were randomly divided into two groups.Mice in group one were infected with MCMV Smith to establish disseminated infection.Mice in another group were sham-infected control.Three mice from each group were randomly chosen to be sacrificed on days 3,7,14 and 28 after the infection.Viral titers in spleen tissues were determined using a standard plaque assay.The expression of IL-17A mRNA and MCMV mRNA in the splenocytes were measured by RT-PCR.The expression of IL-17A in spleen tissues was observed by immunohistochemical staining.The pathology of the infected mice was assessed by histological examination of H&E stained spleen sections.Results Viral titers and MCMV mRNA in the spleen peaked on day 3,but quickly diminished on day 7.Virus was no longer detectable in the spleen on day 14 after the infection.The expression of IL-17A mRNA was significantly increased during the acute infection and reached the highest level on day 14,then decreased on day 28.It is significantly higher than that of the mock infection group.Immunohistochemistry assay also indicated that the expression of IL-17A in spleen tissue gradually increased to climax on day 14,then decreased on day 28.Accordingly,the pathological damages of spleen tissue in the infected mice deteriorated until day 14,then showed signs of recovery on day 28.The most severe pathological injury of spleen tissue and the highest expression of IL-17A appeared in the same period of time.Conclusion Our results showed a close correlation between IL-17A and the pathological damage in spleen.Thus,IL-17A may contribute to the spleen pathological damage during the acute disseminated MCMV infection.

ObjectiveTo observe the changes of AIM2 ( absent in melanoma 2) inflammasome during early murine cytomegalovirus (MCMV) infection.MethodsBALB/c mice were randomly divided into two groups.One was infected with MCMV Smith for establishing disseminated infection,the other was sham-inoculated control.On days 1,3,5 and 7 of the experiment,three mice of each group were randomly chosen to be killed separately.The expression of AIM2,ASC and caspase-1 in splenic macrophages was detected by Western blot,the levels of IL-1β and IL-18 in sera were measured by double antibody sandwich ELISA,and the viral titers in salivary gland tissues were quantified by a standard plaque assay.Results The MCMV titers in salivary gland tissues were gradually increased in MCMV-infected mice on days 3,5 and 7,while the expressions of AIM2 in macrophages were began to increase on day 1 and significantly increased and reached the highest level on day 3 but gradually decreased afterwards.The relative intensity of AIM2 on day 3 differed significantly between the MCMV-infected mice and the controls (1.121±0.243 vs 0.240±0.046,P＜0.01,t test),as did ASC ( 1.318±0.333 vs 0.248±0.090,P＜0.01 ) and caspase-1 ( 1.085±0.243 vs 0.247±0.064,P＜0.01 ).Meanwhile,the levels of IL-1β and IL-18 in MCMV-infected mice were (112.72±5.20) pg/ml and (42.74±4.23) pg/ml,and the levels were significantly higher (P＜0.01 ) than those in controls [ (47.86±4.35) pg/ml and (22.60±2.82) pg/ml].ConclusionThese results demonstrate that AIM2 inflammasome is activated in macrophages during early MCMV infection and could be as a therapeutic target for CMV-induced diseases.

Murine cytomegalovirus (MCMV) IE3 protein is a multifunctional viral protein that interacts with several target proteins of both viral and host cellular origin. To investigate the biological function of IE3 in the pathogenesis of the brain disorders caused by CMV, a screening for host cellular proteins that could interact with IE3 was performed. By yeast two-hybrid screening, ankyrin repeats domain 17 (Ankrd17, also known as Gtar) was identified as a host factor that could interact with IE3. This interaction was verified by yeast two-hybrid assay and chemiluminescent co-immunoprecipitaion. Mapping analysis suggested that the 1-148 residues of IE3 were responsible for the interaction. These results suggested that the interaction between Ankrd17 and IE3 may play a key role in the pathogenesis of MCMV-associated disease.

OBJECTIVE To surveillance invasive fungal infection rate in SICU,in order to direct intervention to prevent invasive fungal infection.METHODS The samples collected from SICU patients in our hospital between Jan 2003-Nov 2008 were cultured.RESULTS According to the diagnosis standard of nosocomial infections,75 case of 3699 patients were isolated fungi.During 6-years invasive fungal infection rate is 2.027%,(1.05%-2.63%).Totally 86 fungi strains were isolated,the majority of them being Candida albicans,accounting for 46.51%;Candida glabrata 22.09%;Candida tropicalis 13.95%.CONCLUSIONS During 6-years,invasive fungal infection rate and incidence density do not increase.Candida are the major pathogens of fungal infections in SICU.

Objective To investigate the influence of murine cytomegalovirus on the expansion of CD+CD25+ Foxp3+ regulatory T cell (Treg) and the activation of CD4+ CD25+Foxp3 - effector T cell (TE) in vivo. Methods Forty-two BALB/c mice were intraperitoneally inoculated with appropriate amount ofMCMV Smith strain for establishing the model of infection, another 42 mice served as mocked-infected con-trois. Day 28 post MCMV infection was determined to be the demarcation point of the acute and chronic in- fection based on the viral load of major visceral organs. On day 1,3, 7, 14, 28, 45 and 60 post infection, splenocytes were prepared by means of routine method. The proportions of CD4+CD25+ Foxp3+Treg and CD4+CD25+Foxp3- activated TE in T lymphocyte were measured by flow cytometry. Results The propor- tion of CD+CD25+ Foxp3+ T cells in T lymphocytes was persistently suppressed since day 7 post infection, and fell to the lowest level on day 28 post infection (P <0.01), then zoomed and reached the peak value on day 60 post infection (P < 0.05). CD4+CD25+ Foxp3 - TE proportion was up to the highest on day 3 post infection(P < 0. 01), then suppressed and in significantly lower level since day 45 post infection (P < 0.05). Treg/CD+TE ratio was in lower level on day 3 to 14 post infection(P <0.05) ; but on day 45 and day 60 post infection Treg/CD+ TE ratio was markedly increased (P < 0.05). Conclusion MCMV infec- tion can increase the CD+CD25+Foxp3+ Treg proportion, and inhibit CD4+T cells activation in chronic in- fection phase, which is likely to suppress the function of antiviral immunity in the infected host to cause a persistent latent infection.