[Objective] The paper was to study molecular characteristics of gp90 gene of 14 Reticuloendotheliosis viruses isolated in China.[Method] The surface envelop gene gp90 of 14 REV strains isolated from different commerci...[Objective] The paper was to study molecular characteristics of gp90 gene of 14 Reticuloendotheliosis viruses isolated in China.[Method] The surface envelop gene gp90 of 14 REV strains isolated from different commercial layer farms in China were amplified,and their nucleotide sequences were determined.[Result] Sequence analysis showes that 14 REV strains are more identical to the subtype 3 isolates than to the early Chinese REV isolates.In addition,14 REV strains have a high identity with some REV strains in US and Taiwan.[Conclusion] The study provided necessary information for further understanding the evolution of REV.展开更多
AIM： To determine the prevalence of porcine endogenous retrovirus （PERV） in various pig breeds raised in China including Chinese experimental mini-pigs by PERV-reverse transcriptase （PERV-RT enzyme）. Moreover, th...AIM： To determine the prevalence of porcine endogenous retrovirus （PERV） in various pig breeds raised in China including Chinese experimental mini-pigs by PERV-reverse transcriptase （PERV-RT enzyme）. Moreover, the potential for infection of PERV was investigated in patients treated with a bioreactor based on porcine liver cells （n = 3）. METHODS： Pig serum, liver and muscle cell-free supernatants were collected from various Chinese pig breeds. Porcine hepatocytes were isolated with a two-step perfusion method. Three patients with acute or chronic liver failure were treated with a bioartificial liver support system （BALSS） for 8-12 h and serum samples were collected from the patients before, immediately after and 30 d after treatment. RESULTS： The activities of PERV-RT enzyme in pig liver and muscle cell-free supernatants were higher than in normal human controls. PERV-TR enzyme activity did not increase in patients before and after 1 mo of treatment. PERV-RT activities were not significantly different when compared with pre-treatment group （1.544±0.155576）, the post-treatment groups （1.501±0.053507, 1.461±0.033808 and 1.6006667±0.01963 for 0, 14 and 30 d post-treatment, respectively, P〉0.05）, and normal control group （1.440±1.0641, P〉0.05）. RT enzyme activity in Chinese experimental mini-pigs was higher than in normal human control group （1.440±1.0641 U/mL, P〈0.05）, and not significantly different （P〉0.05） when compared with the pig breeds except in the muscle supernatants. All the samples including muscle and liver cell supernatants from the Chinese mini-experimental pigs and the four domestic Chinese pig breeds contained PERVs. CONCLUSION： These results suggest that the risk of PERV infection through BALSS containing porcine liver cells without immunosuppressants may be quite low. Although there were PERVs in Chinese experimental mini-pigs and porcine liver cell culture suspensions, we did not find any evidence of persistent PERV infection in patients treated with this porcine hepatocyte-based bioartificial liver.展开更多
In order to evaluate the anti-influenza virus activity of the effective monomer from Folium Isatidis (FI) in vivo,we established mice model with viral pneumonia and divided them into 3 different dose groups,then obser...In order to evaluate the anti-influenza virus activity of the effective monomer from Folium Isatidis (FI) in vivo,we established mice model with viral pneumonia and divided them into 3 different dose groups,then observed their lung indexes,pulmonary pathological changes,pulmonary virus hemagglitination titers,living time and death rates.The results showed that the monomer could reduce the pulmonary index from 2.64 to 1.93,1.63 and 1.40 (P<0.01) and decrease the hemagglitination titer from 1.15 to 0.84,0.70 and 0.59 (P<0.01).In addition,different groups of FI could significantly lessen the mortality rate from 100% to 30%,25% and 15%,and prolong the living time from 5.1d to 6.5d,8.4d and 8.9d respectively(P<0.01).The high dose (75 mg/kg/d) has the similar effect with 100 mg/kg/d dose of virazole(P>0.05),and more effective than 200 mg/kg/d dose of antiviral liquor (P<0.05).展开更多
AIM: To investigate the killing efficiency of a recombinant plasmid containing a thymidine kinase (TK) domain insert driven by the vascular endothelial growth factor receptor 2 (VEGFR2) promoter (KDR) on vascular endo...AIM: To investigate the killing efficiency of a recombinant plasmid containing a thymidine kinase (TK) domain insert driven by the vascular endothelial growth factor receptor 2 (VEGFR2) promoter (KDR) on vascular endothelial cells.METHODS: The KDR-TK fragment was extracted from pBluescript Ⅱ KDR-TK plasmid by enzymatic digestion with Xho I and Sal I. The enhanced green fluorescence protein (EGFP) carrier was extracted from pEGFP by the same procedure. The KDR-TK was inserted into the pEGFP carrier to construct pEGFP-KDR-TK. Using ultrasound irradiation and microbubble, pEGFP-KDR-TK was transferred into human umbilical vein endothelial cells (HUVECs). The transient infection rate was estimated by green fluorescent protein (GFP) expression. Transfected HUVECs, non-transfected HUVECs, and HepG2 cells were cultured in the presence of different concentrations of ganciclovir (GCV), and the killing efficacy of HSV-TK/GCV was analyzed by 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl tetrazolium bromide (MTT) assay. RESULTS: The recombinant pEGFP-KDR-TK was successfully constructed by inserting the KDR-TK fragment into the pEGFP carrier. Transfected HUVECs showed cytoplasmic green fluorescence, and the transient transfection rate was about 20.3%. Pools of G418-resistant cells exhibited a higher sensitivity to theprodrug/GCV compared to non-transfected HUVECs or non-transfected HepG2 cells, respectively. CONCLUSION: KDR promoter and the suicide gene/prodrug system mediated by diagnostic ultrasound combined with microbubble can significantly kill HUVECs. Such therapy may present a novel and attractive approach to target gene therapy on tumor vessels.展开更多
We attempted to improve the activity of hTERT promoter by fusing the vascular endothelial growth factor （VEGF） enhancer. To determine the potential as cancer specific promoters, we measured the reporter gene transfe...We attempted to improve the activity of hTERT promoter by fusing the vascular endothelial growth factor （VEGF） enhancer. To determine the potential as cancer specific promoters, we measured the reporter gene transfection assay driven by the hTERT promoter and the VEGF enhancer in human cancer cells. We found that the hTERT promoter containing VEGF enhancer conferred strong expression of the reporter gene only in different cancer cell lines but not in normal human cells. Retrovirus vector expressing HSV-TK controlled by the hTERT promoter and the VEGF enhancer was constructed. A549 cells infected with LN-enh-hT-TK was significantly suppressed and induced to apoptosis more than those infected with LN-hT-TK. The apoptosis ratio ofA549 cell infected with two kinds of retrovirus cell with GCV in lower concentration is 20.94% and 50.7%. It suggested that there is significant differentiation between the assay groups. Our results demonstrated the possible application of hTERT promoter and the VEGF enhancer in targeted cancer gene therapy.展开更多
Objective： To determine the potential of sustained transgene expression by intratumoral injection of Ad-PTEN in the nude mouse model of endometrial carcinoma. Methods and Results： We constructed recombinant adenovir...Objective： To determine the potential of sustained transgene expression by intratumoral injection of Ad-PTEN in the nude mouse model of endometrial carcinoma. Methods and Results： We constructed recombinant adenovirus carrying the wild-type PTEN gene （Ad-PTEN）. RL95-2 cells, an endometrial carcinoma cell line lacking PTEN function, was infected with Ad-PTEN and showed increased expression of PTEN and chemosensitivity to doxorubicin, decreased proliferation rate, and elevated apoptosis and Go/G1 arrest. Furthermore, the tumorigenicity of these cells was also completely suppressed. These results indicated that gene therapy with Ad-PTEN could significantly inhibit the endometrial carcinoma xenografts growth in nude mice by intratumoral injection, induce apoptosis of tumor cells, and reduce expression of proliferating cell nuclear antigen （PCNA）. Immunohistochemistry analysis also showed that the expression of progesterone receptors （PR） in Ad-PTEN treated tumor cells were induced, while P-glycoproteins （P-gp） and estrogen receptors （ER） decreased significantly. Conclusion： PTEN may play an important role in the development of endometrial carcinoma. Our findings cast new lights for treatment ofendometrial carcinoma.展开更多
OBJECTIVE: To verify whether the extracellular domain of kinase domain region (KDR) has anti-angiogenesis activity in vivo. METHODS: cDNA was cloned into adeno-associated virus (AAV) vector pSNAV and transfected to ba...OBJECTIVE: To verify whether the extracellular domain of kinase domain region (KDR) has anti-angiogenesis activity in vivo. METHODS: cDNA was cloned into adeno-associated virus (AAV) vector pSNAV and transfected to baby hamster kidney (BHK) cells. Recombinant AAV was obtained from the cell culture supernatant after adding helper virus. Recombinant AAV-infected human bladder cancer EJ cell line (EJ cells) were injected subcutaneously into Balb-c nude mice. Tumor specimens were removed from the mice, paraffin-embedded and sliced, then stained by immunohistochemistry. Microvessel density (MVD) was determined under a microscope. RESULTS: The tumor volume developed by EJ cells transfected with the extracellular domain of KDR was significantly smaller (1.70 +/- 0.18 cm(3)) compared with that in the control (5.62 +/- 0.67 cm(3)) (P展开更多
We report the recovery of a 7068-nt viral sequence from the ＂viral fossils＂ embedded in the genome of Alhagi sparsifolia, a typical desert plant. Although the full viral genome remains to be completed, the putative ...We report the recovery of a 7068-nt viral sequence from the ＂viral fossils＂ embedded in the genome of Alhagi sparsifolia, a typical desert plant. Although the full viral genome remains to be completed, the putative genome structure, the deduced amino acids and phylogenetic analysis unambiguously demonstrate that this viral sequence represents a novel species of the genus Badnavirus. The putative virus is tentatively termed Alhagi bacilliform virus （ABV）. Southern blotting and inverse polymerase chain reaction （PCR） data indicate that the ABV-related sequence is integrated into the A. sparsifolia genome, and probably does not give rise to functional episomal virus. Molecular evidence that the ABV sequence exists widely in A. sparsifolia is also presented. To our knowledge, this is the first endogenous badnavirus identified from plants in the Gobi desert, and may provide new clues on the evolution, geo- graphical distribution as well as the host range of the badnaviruses.展开更多
Antiviral defense is one of the important roles of RNA silencing in plants. Virus-derived small interfering RNAs (vsiRNAs) are found in the infected host cells, indicating that the host RNA silencing machinery can tar...Antiviral defense is one of the important roles of RNA silencing in plants. Virus-derived small interfering RNAs (vsiRNAs) are found in the infected host cells, indicating that the host RNA silencing machinery can target viral RNAs for destruction. With the development of high-throughput sequencing of vsiRNAs, recent genetic studies have shed light on the origin and composition of vsiRNAs and their potential functions in the regulation of gene expression. Here, we briefly describe the origin and biogenesis of vsiRNAs, and review the recent discoveries regarding vsiRNA-mediated RNA silencing of viral genomes and host transcripts. This will better our understanding of virus pathogenicity and RNA silencing-related host-pathogen inter- actions in plants.展开更多
基金Supported by Earmarked Fund for Modern Agro-industry Technology Research System of China ( nycytx-42-G3-01)~~
文摘[Objective] The paper was to study molecular characteristics of gp90 gene of 14 Reticuloendotheliosis viruses isolated in China.[Method] The surface envelop gene gp90 of 14 REV strains isolated from different commercial layer farms in China were amplified,and their nucleotide sequences were determined.[Result] Sequence analysis showes that 14 REV strains are more identical to the subtype 3 isolates than to the early Chinese REV isolates.In addition,14 REV strains have a high identity with some REV strains in US and Taiwan.[Conclusion] The study provided necessary information for further understanding the evolution of REV.
文摘AIM： To determine the prevalence of porcine endogenous retrovirus （PERV） in various pig breeds raised in China including Chinese experimental mini-pigs by PERV-reverse transcriptase （PERV-RT enzyme）. Moreover, the potential for infection of PERV was investigated in patients treated with a bioreactor based on porcine liver cells （n = 3）. METHODS： Pig serum, liver and muscle cell-free supernatants were collected from various Chinese pig breeds. Porcine hepatocytes were isolated with a two-step perfusion method. Three patients with acute or chronic liver failure were treated with a bioartificial liver support system （BALSS） for 8-12 h and serum samples were collected from the patients before, immediately after and 30 d after treatment. RESULTS： The activities of PERV-RT enzyme in pig liver and muscle cell-free supernatants were higher than in normal human controls. PERV-TR enzyme activity did not increase in patients before and after 1 mo of treatment. PERV-RT activities were not significantly different when compared with pre-treatment group （1.544±0.155576）, the post-treatment groups （1.501±0.053507, 1.461±0.033808 and 1.6006667±0.01963 for 0, 14 and 30 d post-treatment, respectively, P〉0.05）, and normal control group （1.440±1.0641, P〉0.05）. RT enzyme activity in Chinese experimental mini-pigs was higher than in normal human control group （1.440±1.0641 U/mL, P〈0.05）, and not significantly different （P〉0.05） when compared with the pig breeds except in the muscle supernatants. All the samples including muscle and liver cell supernatants from the Chinese mini-experimental pigs and the four domestic Chinese pig breeds contained PERVs. CONCLUSION： These results suggest that the risk of PERV infection through BALSS containing porcine liver cells without immunosuppressants may be quite low. Although there were PERVs in Chinese experimental mini-pigs and porcine liver cell culture suspensions, we did not find any evidence of persistent PERV infection in patients treated with this porcine hepatocyte-based bioartificial liver.
基金Natural Science Foundation of South Central University for Nationalities (YZQ05011)
文摘In order to evaluate the anti-influenza virus activity of the effective monomer from Folium Isatidis (FI) in vivo,we established mice model with viral pneumonia and divided them into 3 different dose groups,then observed their lung indexes,pulmonary pathological changes,pulmonary virus hemagglitination titers,living time and death rates.The results showed that the monomer could reduce the pulmonary index from 2.64 to 1.93,1.63 and 1.40 (P<0.01) and decrease the hemagglitination titer from 1.15 to 0.84,0.70 and 0.59 (P<0.01).In addition,different groups of FI could significantly lessen the mortality rate from 100% to 30%,25% and 15%,and prolong the living time from 5.1d to 6.5d,8.4d and 8.9d respectively(P<0.01).The high dose (75 mg/kg/d) has the similar effect with 100 mg/kg/d dose of virazole(P>0.05),and more effective than 200 mg/kg/d dose of antiviral liquor (P<0.05).
基金New Century Distinguished Scholar Supporting Program of Ministry of Education (80000-3171404) The National Natural Science Foundation of China, No. 30300082, No. 30470467
文摘AIM: To investigate the killing efficiency of a recombinant plasmid containing a thymidine kinase (TK) domain insert driven by the vascular endothelial growth factor receptor 2 (VEGFR2) promoter (KDR) on vascular endothelial cells.METHODS: The KDR-TK fragment was extracted from pBluescript Ⅱ KDR-TK plasmid by enzymatic digestion with Xho I and Sal I. The enhanced green fluorescence protein (EGFP) carrier was extracted from pEGFP by the same procedure. The KDR-TK was inserted into the pEGFP carrier to construct pEGFP-KDR-TK. Using ultrasound irradiation and microbubble, pEGFP-KDR-TK was transferred into human umbilical vein endothelial cells (HUVECs). The transient infection rate was estimated by green fluorescent protein (GFP) expression. Transfected HUVECs, non-transfected HUVECs, and HepG2 cells were cultured in the presence of different concentrations of ganciclovir (GCV), and the killing efficacy of HSV-TK/GCV was analyzed by 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl tetrazolium bromide (MTT) assay. RESULTS: The recombinant pEGFP-KDR-TK was successfully constructed by inserting the KDR-TK fragment into the pEGFP carrier. Transfected HUVECs showed cytoplasmic green fluorescence, and the transient transfection rate was about 20.3%. Pools of G418-resistant cells exhibited a higher sensitivity to theprodrug/GCV compared to non-transfected HUVECs or non-transfected HepG2 cells, respectively. CONCLUSION: KDR promoter and the suicide gene/prodrug system mediated by diagnostic ultrasound combined with microbubble can significantly kill HUVECs. Such therapy may present a novel and attractive approach to target gene therapy on tumor vessels.
文摘We attempted to improve the activity of hTERT promoter by fusing the vascular endothelial growth factor （VEGF） enhancer. To determine the potential as cancer specific promoters, we measured the reporter gene transfection assay driven by the hTERT promoter and the VEGF enhancer in human cancer cells. We found that the hTERT promoter containing VEGF enhancer conferred strong expression of the reporter gene only in different cancer cell lines but not in normal human cells. Retrovirus vector expressing HSV-TK controlled by the hTERT promoter and the VEGF enhancer was constructed. A549 cells infected with LN-enh-hT-TK was significantly suppressed and induced to apoptosis more than those infected with LN-hT-TK. The apoptosis ratio ofA549 cell infected with two kinds of retrovirus cell with GCV in lower concentration is 20.94% and 50.7%. It suggested that there is significant differentiation between the assay groups. Our results demonstrated the possible application of hTERT promoter and the VEGF enhancer in targeted cancer gene therapy.
基金Supported by the National Natural Science Foundation of China(30471676)Shanghai Science and Technology Committee(04DZ19207-2)
文摘Objective： To determine the potential of sustained transgene expression by intratumoral injection of Ad-PTEN in the nude mouse model of endometrial carcinoma. Methods and Results： We constructed recombinant adenovirus carrying the wild-type PTEN gene （Ad-PTEN）. RL95-2 cells, an endometrial carcinoma cell line lacking PTEN function, was infected with Ad-PTEN and showed increased expression of PTEN and chemosensitivity to doxorubicin, decreased proliferation rate, and elevated apoptosis and Go/G1 arrest. Furthermore, the tumorigenicity of these cells was also completely suppressed. These results indicated that gene therapy with Ad-PTEN could significantly inhibit the endometrial carcinoma xenografts growth in nude mice by intratumoral injection, induce apoptosis of tumor cells, and reduce expression of proliferating cell nuclear antigen （PCNA）. Immunohistochemistry analysis also showed that the expression of progesterone receptors （PR） in Ad-PTEN treated tumor cells were induced, while P-glycoproteins （P-gp） and estrogen receptors （ER） decreased significantly. Conclusion： PTEN may play an important role in the development of endometrial carcinoma. Our findings cast new lights for treatment ofendometrial carcinoma.
文摘OBJECTIVE: To verify whether the extracellular domain of kinase domain region (KDR) has anti-angiogenesis activity in vivo. METHODS: cDNA was cloned into adeno-associated virus (AAV) vector pSNAV and transfected to baby hamster kidney (BHK) cells. Recombinant AAV was obtained from the cell culture supernatant after adding helper virus. Recombinant AAV-infected human bladder cancer EJ cell line (EJ cells) were injected subcutaneously into Balb-c nude mice. Tumor specimens were removed from the mice, paraffin-embedded and sliced, then stained by immunohistochemistry. Microvessel density (MVD) was determined under a microscope. RESULTS: The tumor volume developed by EJ cells transfected with the extracellular domain of KDR was significantly smaller (1.70 +/- 0.18 cm(3)) compared with that in the control (5.62 +/- 0.67 cm(3)) (P
基金Project supported by the National Natural Science Foundation of China(Nos.31370181 and 31570146)the Fujian Natural Science Funds for Distinguished Young Scholar(No.2014J06008),China
文摘We report the recovery of a 7068-nt viral sequence from the ＂viral fossils＂ embedded in the genome of Alhagi sparsifolia, a typical desert plant. Although the full viral genome remains to be completed, the putative genome structure, the deduced amino acids and phylogenetic analysis unambiguously demonstrate that this viral sequence represents a novel species of the genus Badnavirus. The putative virus is tentatively termed Alhagi bacilliform virus （ABV）. Southern blotting and inverse polymerase chain reaction （PCR） data indicate that the ABV-related sequence is integrated into the A. sparsifolia genome, and probably does not give rise to functional episomal virus. Molecular evidence that the ABV sequence exists widely in A. sparsifolia is also presented. To our knowledge, this is the first endogenous badnavirus identified from plants in the Gobi desert, and may provide new clues on the evolution, geo- graphical distribution as well as the host range of the badnaviruses.
文摘Antiviral defense is one of the important roles of RNA silencing in plants. Virus-derived small interfering RNAs (vsiRNAs) are found in the infected host cells, indicating that the host RNA silencing machinery can target viral RNAs for destruction. With the development of high-throughput sequencing of vsiRNAs, recent genetic studies have shed light on the origin and composition of vsiRNAs and their potential functions in the regulation of gene expression. Here, we briefly describe the origin and biogenesis of vsiRNAs, and review the recent discoveries regarding vsiRNA-mediated RNA silencing of viral genomes and host transcripts. This will better our understanding of virus pathogenicity and RNA silencing-related host-pathogen inter- actions in plants.
