OBJECTIVETo study the relationship between late mRNA and the cytopathic effect(CPE) and ultrastructural features after human cytomegalovirus (HCMV) infection in vitro.

METHODSHuman embryo fibroblast cells(HEL) were infected with HCMV AD169 strain. The expression of the HCMV late mRNA was measured by semiquantitative RT-PCR, the cytopathic effect (CPE) and the cell ultrastructure were observed by means of light microscopy and electron microscopy, respectively.

RESULTSThe HCMV late mRNA could be detected 12 hours postinfection and increased gradually, but the CPE appeared 48 hours postinfection in HEL cells. The HCMV infected cells exhibited significant mitochondrial enlargement and the number of mitochondrial ridge deletion, the cisternae lumen of endoplasmic reticulum (ER) dilation and vacuolization (at the end age). The mature nucleocapsid could be observed 96 hours postinfection.

CONCLUSIONThe ultrastructural changes have an intimate correlation with the expression of HCMV late mRNA and play an important role in the life circle of the virus. HCMV late mRNA may serve as a indicator of the clinical effect of treatment in active HCMV infection.

Cytomegalovirus ; genetics ; physiology ; Cytopathogenic Effect, Viral ; Embryo, Mammalian ; Endoplasmic Reticulum ; pathology ; virology ; Fibroblasts ; metabolism ; ultrastructure ; virology ; Humans ; Inclusion Bodies ; pathology ; virology ; Mitochondrial Swelling ; RNA, Messenger ; metabolism

The glycoprotein 3 (GP3) of type II porcine reproductive and respiratory syndrome virus has the characteristic domains of a membrane protein. However, this protein has been reported to be retained in the endoplasmic reticulum (ER) rather than transported to the plasma membrane of the cell. In this study, we performed confocal laser scanning microscopy analysis of variants of GP3 and foundthat the signal sequence of the GP3 led to confinement of GP3 in the ER, while the functional ortransmembrane domain did not affect its localization. Based on these results, we concludedthat the signal sequence of GP3 contains the ER retention signal, which might play an important role in assembly of viral proteins.
Animals ; Cell Line ; Cell Membrane/*metabolism/virology ; Cricetinae ; Endoplasmic Reticulum/*metabolism/virology ; Microscopy, Confocal/veterinary ; Plasmids/genetics/metabolism ; Porcine respiratory and reproductive syndrome virus/*genetics/metabolism ; *Protein Sorting Signals ; Sequence Analysis, Protein/veterinary ; Viral Envelope Proteins/chemistry/*genetics/metabolism

OBJECTIVETo investigate the effect of total flavonoids of astragalus on the expression of endoplasmic reticulum chaperone, calumenin and connecxin 43 (CX43) in suckling mouse myocardium with myocarditis caused by coxsackievirus B3 (CVB3).

METHODSThe primary culture of suckling mouse myocardium cells were randomly divided into control group, CVB3 infected group and total flavonoids of astragalus group. Firstly, to confirm the identity of the suckling mouse myocardium, α-SMA was monitored by immunohistochemistry method. Then the protein expression changes of endoplasmic reticulum chaperone-glucose regulatory protein 78 ( GRP78), calumenin and CX43 were detected by Western blot.

RESULTS(1) Compared with that of the control group, the GRP78 expression level in CVB3 infected group was improved, the expression levels of calumenin and CX43 were all reduced. (2) Compared with that of CVB3 infected group, GRP78 expression level was decreased, and the expression levels of calumenin and CX43 were increased in total flavonoids of astragalus group.

CONCLUSIONCVB3 infection may cause endoplasmic reticulum stress of rat myocardium cells by increasing the expression of GRP78 and decreasing the expression of calumenin and CX43. On the other hand, total flavonoids of astragalus can reduce the expression of GRP78 and increase the expression of calumenin and CX43.The results of this experiment may be closely related to the effects of anti-arrhythmia with viral myocarditis caused by CVB3.

Animals ; Astragalus Plant ; chemistry ; Blotting, Western ; Calcium-Binding Proteins ; metabolism ; Cells, Cultured ; Connexin 43 ; metabolism ; Coxsackievirus Infections ; drug therapy ; Endoplasmic Reticulum ; metabolism ; Endoplasmic Reticulum Stress ; drug effects ; Flavonoids ; pharmacology ; Heat-Shock Proteins ; metabolism ; Mice ; Myocarditis ; drug therapy ; virology ; Myocardium ; cytology ; Myocytes, Cardiac ; drug effects ; virology ; Rats

OBJECTIVETo explore the apoptotic pathway mediated by endoplasmic reticulum stress in the mouse myocardium with heart failure induced by acute viral myocarditis caused by B-3 Coxsackie virus.

METHODSForty BALB/c male mice were randomly divided into 2 groups (n = 20): the control group and the virus infection group. The BALB/c mouse myocarditis was induced by B-3 Coxsackie virus and the mouse behavior was observed conventionally. All the mice were sacrificed on day 7 and the changes of left ventricular pressure (LVP) and the rate of change of left ventricular pressure (LV dp/dt) were measured. The cardiomyocytic apoptosis was analyzed by TUNEL method and the mRNA expression level of endoplasmic reticulum haperones glucose-regulated protein (GRP)78 and GRP94 was detected by RT-PCR.

RESULTS(1) Compared with those of control group, the parameters of cardiac hemodynamics in the virus infection group were significantly decreased (P < 0.01); (2) Compared with that of control group, myocardial apoptosis was significantly increased in the myocardial cells from mice with heart failure induced by acute viral myocarditis (P < 0.01); (3) The mRNA expression level of GRP78 and GRP94 were increased significantly in the virus infection group compared with the control group.

CONCLUSIONThese findings suggest the endoplasmic reticulum stress may mediate the apoptosis of myocardial cells in the mice myocardium of heart failure induced by acute viral myocarditis caused by B-3 Coxsackie virus.

Animals ; Apoptosis ; Coxsackievirus Infections ; physiopathology ; Endoplasmic Reticulum Stress ; Heart ; physiopathology ; Heart Failure ; physiopathology ; virology ; Heat-Shock Proteins ; metabolism ; Male ; Membrane Glycoproteins ; metabolism ; Mice ; Mice, Inbred BALB C ; Myocarditis ; physiopathology ; virology ; Myocardium ; pathology ; Myocytes, Cardiac ; cytology

OBJECTIVETo study the death mode of human hepatocellular carcinoma Hep-3B cells and human lung adenocarcinoma A549 cells induced by bluetongue virus strain HbC3 (BTV-HbC3) and the mechanism of its action.

METHODSBTV-HbC3 was used to infect the tumor cells, and the cytopathic effects (CPE) was observed. TUNEL staining was used to detect the apoptosis of tumor cells induced by BTV-HbC3. The changes of endoplasmic reticulum and nuclei treated with BTV-HbC3 were further examined by laser scanning confocal microscopy. The activities of caspase-3/7, caspase-8 and caspase-9 were determined by fluorescence analysis.

RESULTSHep-3B cells were sensitive to BTV-HbC3. Lots of early apoptotic cells were found by TUNEL staining. The laser scanning confocal microscopic examination showed characteristics of apoptosis, such as pyknotic nuclei, margination of nuclear chromatin and vacuolization of endoplasmic reticulumin in Hep-3B cells exposed to BTV-HbC3. The activity of caspase-3/7 was increased, but the activity changes of caspase-8 and caspase-9 were not found. A549 cells were sensitive to BTV-HbC3 too. But no apoptotic cells were observed by TUNEL staining. The results of laser scanning confocal microscopy showed marked vacuolization of endoplasmic reticulum, but chromatin margination was not found after A549 cells was exposed to BTV-HbC3. The activity of caspase-3/7 and caspase-9 was increased, but the activity of caspase-8 was not changed.

CONCLUSIONBTV-HbC3 induces apoptosis of Hep-3B tumor cells mainly through endoplasmic reticulum signal transduction pathway, and the features of cell death in A549 cells could be described as paraptosis.

Adenocarcinoma ; metabolism ; pathology ; virology ; Apoptosis ; Bluetongue virus ; pathogenicity ; physiology ; Carcinoma, Hepatocellular ; metabolism ; pathology ; virology ; Caspase 3 ; metabolism ; Caspase 8 ; metabolism ; Caspase 9 ; metabolism ; Cell Line, Tumor ; Cell Nucleus ; pathology ; Endoplasmic Reticulum ; pathology ; Humans ; Liver Neoplasms ; metabolism ; pathology ; virology ; Lung Neoplasms ; metabolism ; pathology ; virology ; Oncolytic Viruses ; pathogenicity ; physiology ; Signal Transduction

OBJECTIVE: To investigate the effects of total flavonids of astragalus(TFA) on arrhythmia, endoplasmic reticulum stress and connexcin in mice with viral myocarditis and to clarify the mechanisms of TFA against viral myocarditis complicated with arrhythmia. METHODS: Thirty-six male Balb/c mice were randomly divided into control group, viral myocarditis group and total flavonoids group (=12). The mice of viral myocarditis were intraperitonealy injected with 0.1 ml/day 10-950 TCID CVB3 for 3 days. The mice of TFA group were intraperitoneal injected with 0.1 ml/day 10-950 TCID CVB3 for 3 days and treated with 0.1ml, 20 mg/L TFA by tail vein injection. At the end of the experiment, arrhythmia was detected by electrocardiogram, the heart of mice were stained by HE, the expressions of glucose-regulated protein 78(GRP78), endoplasmic reticulum stress signaling pathway factor activating transcription factor 4(ATF4) and connexcin 43(Cx43) were detected by Western blot. RESULTS: The expressions of GRP78 and ATF4 were increased and the expression of Cx43 was decreased in viral myocarditis, while TFA inhibited these effect of viral myocarditis in heart of mice. CONCLUSIONS The antiarrhythmic effect of TFA may be related to the alleviation of endoplasmic reticulum stress and the increase of Cx43 expression.
Activating Transcription Factor 4 ; metabolism ; Animals ; Arrhythmias, Cardiac ; drug therapy ; Astragalus Plant ; chemistry ; Connexin 43 ; metabolism ; Coxsackievirus Infections ; drug therapy ; Drugs, Chinese Herbal ; pharmacology ; Endoplasmic Reticulum Stress ; drug effects ; Flavonoids ; pharmacology ; Heat-Shock Proteins ; metabolism ; Male ; Mice ; Mice, Inbred BALB C ; Myocarditis ; drug therapy ; virology ; Myocardium