Human herpesvirus 8(HHV-8) is thought to be essential for the development of all forms of Kaposi's sarcoma(KS).HHV-8 DNA is present virtually in all KS tumor biopsy samples.Genes at both ends of the HHV-8 genome h...Human herpesvirus 8(HHV-8) is thought to be essential for the development of all forms of Kaposi's sarcoma(KS).HHV-8 DNA is present virtually in all KS tumor biopsy samples.Genes at both ends of the HHV-8 genome have been shown to vary considerably.Seven major molecular subtypes of HHV-8 were defined based on the amino acid sequence of the open reading frame K1(ORF-K1),generally known as A,B,C,D,E,F,and Z.Most strains collected worldwide were clustered into two subtypes(A and C).Here,the K1/VR1 region of HHV-8 was amplified by nested PCR in 22(81.48%) of 27 cases from Xinjiang Uygur Autonomous Region,a province in northwestern China.Phylogenetic analysis on the basis of the K1/VR1 amino acid sequence indicated that the majority of these KS patients were infected by subtype C HHV-8(n=18,including 15 belonging to the C2 group),and several by subtype A(n=4,including 3 being the A1 group).This is the first report of subtype A HHV-8 in China.Furthermore,the correlations between different forms and lesions of KS and different subtypes of HHV-8 were analyzed.The findings showed that subtype A HHV-8 resulted in significantly more frequent mucosal KS lesions than subtype C.However,there was no obvious correlation between different forms of KS and different subtypes of HHV-8.展开更多
Objective:To study and confirm that recombinant cytokines similar to those produced by HIV-1 infected T cells induced lytic cycle replication of human pervirus 8(HHV-8) in BC-3 cells,another cell line from primary eff...Objective:To study and confirm that recombinant cytokines similar to those produced by HIV-1 infected T cells induced lytic cycle replication of human pervirus 8(HHV-8) in BC-3 cells,another cell line from primary effusion lymphoma(PEL).Methods:The persistent stimulation of BC-3 was conducted by several cytokines known to be produced by HIV-1-infected T cells and important in grouth and proliferation of Kaposi's sarcoma(KS) cells in vitro,such as the inter feron-γ（IFN-γ）,the hepatocyte grouth factor /scatter factor (HGF/SF),the Oncostain M(OSM),and the tumor necrosis factor-α（TNF-α）which is not produced by HIV-1-infeted T cells.Treated and antrented BC-3 cells were collected at the 3rd and 7th day of persistent stimulation,respeclively.Immunohistochemical(IHC) staining. Northern blot,quantitative PCR(real-time PCR) and electron microscopy(EM) were carried out to detect the expression of immumogenic protein ORF59,messenger RNA(mRNA) of minor capsid protein ORF26,and the presence of viral particles of HHV-8 from treated and untreated BC-3 cells.Results:It showed that IFN-γ,HGF/SF/OSM,and TNF-α were found to induce an increase in mRNA expression of ORF26 when added individually to BC-3 cells.Particularly,ORF26 expression stimulated with IFN-γ and TNF-α respectively,increased 6.1 and 2.5-fold(from,real-time PCR results) at the 7th day when compared with untreated BC-3 cells.Meanwhile ,about 20% of IFN-γ stimulated BC-3 cells expressed ORF59 at the 7th day as compared with 1.5% of untreated BC-3 cells when IHC staining was employed.In addition,viral particles of HHV-8 were readily idelified in BC-3 cells stimulated with IFN-γ at the 7th day with EM analysis.Conclusion:TNF-α and recombinant cytokines being similar to those produced by HIV-1 infected T Cells could really induce HHV-8 lytic cycle replication in BC-3 cells,another cell line of PEL.展开更多
Objective: To investigate the effect of activated protein C (APC) on inflammatory responses in human umbilical vein endothelial cells (HUVEC) stimulated with lipopolysaccharide (LPS). Methods: The second passage of co...Objective: To investigate the effect of activated protein C (APC) on inflammatory responses in human umbilical vein endothelial cells (HUVEC) stimulated with lipopolysaccharide (LPS). Methods: The second passage of collagenase digested HUVEC was divided into the following groups: serum free medium control group (SFM control), phosphate buffer solution control group (PBS control), LPS group with final concentration of 1 μg/ml (LPS group), APC group with final concentration of 7 μg/ml, Pre-APC group (APC pretreatment for 30 min prior to LPS challenge), and Post-APC group (APC administration 30 min after LPS challenge). Supernatant was harvested at 0, 4, 8, 12 and 24 h after LPS challenge. Interleukin-6 (IL-6) and Interleukin-8 (IL-8) levels were analyzed with ELISA. Cells were harvested at 24 h after LPS challenge, and total RNA was extracted. Mes-senger RNA levels for IL-6 and IL-8 were semi-quantitatively determined by RT-PCR. Results: Compared with control group, IL-6 and IL-8 levels steadily increased 4 to 24 h after LPS stimulation. APC treatment could increase LPS-induced IL-6 and IL-8 production. The mRNA levels of IL-6 and IL-8 exhibited a similar change. Conclusion: APC can further increase the level of IL-6 and IL-8 induced by LPS. The effect of these elevated cytokines is still under investigation.展开更多
Although Kaposi sarcoma(KS) has been more traditionally considered an AIDS-defining illness,it may also be seen in individuals on immunosuppresive therapy.We report a case of a patient who presented to the hospital in...Although Kaposi sarcoma(KS) has been more traditionally considered an AIDS-defining illness,it may also be seen in individuals on immunosuppresive therapy.We report a case of a patient who presented to the hospital in the setting of increasingly refractory ulcerative colitis.Computed tomography scan of the abdomen was consistent with sigmoid diverticulititis and blood cultures were positive for Klebsiella.After a course of antibiotics with resolution of infection,a colonoscopy was performed to evaluate his diverticulitis and incidentally revealed a new rectal tumor.Immunohistochemistry showed the tumor was consistent with KS,with cells staining strongly positive for human herpesvirus-8.This case not only illustrates a rare case of KS found in an HIV-negative individual,but it also highlights the importance of considering an alternative diagnosis in a patient refractory to medical treatment.We discuss the management and care of an ulcerative colitis patient diagnosed with KS on immunosuppressive therapy.展开更多
AIM: To investigate the prevalence of human leukocyte antigen (HLA) DQ2/8 alleles in Southern Italians with liver and gastrointestinal (GI) diseases outside of celiac disease. METHODS: HLA DQ2/8 status was assessed in...AIM: To investigate the prevalence of human leukocyte antigen (HLA) DQ2/8 alleles in Southern Italians with liver and gastrointestinal (GI) diseases outside of celiac disease. METHODS: HLA DQ2/8 status was assessed in 443 patients from three ambulatory gastroenterology clinics in Southern Italy (University of Federico Ⅱ, Naples, Loreto Crispi Hospital, Ruggi D'Aragona Hospital, Salerno). Patients were grouped based on disease status [pre-post transplant liver disease, esophageal/gastric organic and functional diseases, irritable bowel syndrome (IBS) and inflammatory bowel disease (IBD)] and DQ2/8 alleles, which correspond to a celiac disease genetic risk gradient. Subject allele frequencies were compared to healthy Italian controls. RESULTS: One hundred and ninety-six out of four hundred and forty-three (44.2%) subjects, median age 56 years and 42.6% female, were DQ2/8 positive. When stratifying by disease we found that 86/188 (45.7%) patients with liver disease were HLA DQ2/8 positive, 39/73 (53.4%) with functional upper GI diseases and 19/41 (46.3%) with organic upper GI diseases were positive. Furthermore, 38/105 (36.2%) patients with IBS and 14/36 (38.9%) with IBD were HLA DQ2/8 positive (P = 0.21). Compared to healthy controls those with functional upper GI diseases disorders had a 1.8 times higher odds of DQ2/8 positivity. Those with liver disease had 1.3 times the odds, albeit not statistically significant, ofDQ2/8 positivity. Both those with IBS and IBD had a lower odds of DQ2/8 positivity compared to healthy controls. CONCLUSION: The proportion of individuals HLA DQ2/8 positive is higher in those with liver/upper functional GI disease and lower in IBS/IBD as compared to general population estimates.展开更多
To establish a sensitive and specific method for seroepidermiological detection of human her- pes virus 8(HHV-8)infection,three potent antigenic proteins encoded by open reading frames(ORFs) K8.1,65 and 73C in genome ...To establish a sensitive and specific method for seroepidermiological detection of human her- pes virus 8(HHV-8)infection,three potent antigenic proteins encoded by open reading frames(ORFs) K8.1,65 and 73C in genome of HHV-8 were produced as glutathione S-transferase fusion protein in the prokaryotic expression system and was used as antigen for testing.The recombinant fusion protein ex- pressed in the prokaryotic expression vector E.coli BL21 was purified by glutathione Sepharose 4B affin- ity chromatography and was quantitated with SDS-PAGE.All these 3 fusion proteins produced in the pro- karyotic expression system showed good immunogenicity as demonstrated by Western blotting and could be recognized by mixed sera of patients with Kaposi′s sarcoma(KS).The immuno-reactivities of the single or compound fusion protein were determined by means of ELISA and compared with the traditional immu- nofluorescence assay(IFA)to determine their sensitivity and specificity of the test.It was demonstrated that the sensitivity of mixed-antigen ELISA method was significantly higher than that of IFA(81.8% vs 34.4%),while the specificity of the former was demonstrated to be 97.9%.The coincidence of the de- tection rate between these two methods was considerably high,approaching up to 90.0%.These results suggest that the mixed antigen ELISA assay appears to be a sensitive and specific method for sero-epide- miological detection of human herpesvirus 8 infection.展开更多
Despite the progress has been reached with Human herpesvirus 8 (HHV-8) research, there are gaps in the knowledge of viral induced oncogenesis. The aim of the present study was to identify possible associations between...Despite the progress has been reached with Human herpesvirus 8 (HHV-8) research, there are gaps in the knowledge of viral induced oncogenesis. The aim of the present study was to identify possible associations between HHV-8 subtypes, HHV-8 loads and clinical manifestations of HIV infected patients diagnosed with different malignancies associated with HHV-8 infection. Forty six HIV-1 infected individuals diagnosed with different HHV-8 associated diseases were studied [37 epidemic Kaposi’s sarcoma (KS), 3 pleural effusion lymphoma (PEL);5 peripheral lymphadenopathies (PL);1 Hodgkin’s lymphoma (HL);1 non Hodgkin’s lymphoma (NHL)]. HHV-8 loads were determined by quantitative real time PCR (qRT-PCR) whilst HHV-8 subtypes were determined by open-reading frame (ORF)-K1 gen genotyping. HHV-8 subtypes B, A, C, A5 and E were exhibited by 31.8%, 23.4%, 19.1%, 17% and 8.5% of the studied patients, respectively. The median HHV-8 viral load did not differ between subtypes (p > 0.05) but HHV-8 viral loads were significantly higher in PEL than in epidemic KS lesion or lymph nodes (p = 0.04). Subtype B was detected in 60% of patients with B cell lymphoma (NHL, PEL and HL) whereas subtype E was only detected in patients with epidemic KS diagnosis. Our data suggest that HHV-8 DNA quantification instead of subtype identification could be used as a surrogate marker for monitoring its infection, not only in epidemic KS patients but also in HIV infected individuals with lymphoproliferative disorders.展开更多
文摘Human herpesvirus 8(HHV-8) is thought to be essential for the development of all forms of Kaposi's sarcoma(KS).HHV-8 DNA is present virtually in all KS tumor biopsy samples.Genes at both ends of the HHV-8 genome have been shown to vary considerably.Seven major molecular subtypes of HHV-8 were defined based on the amino acid sequence of the open reading frame K1(ORF-K1),generally known as A,B,C,D,E,F,and Z.Most strains collected worldwide were clustered into two subtypes(A and C).Here,the K1/VR1 region of HHV-8 was amplified by nested PCR in 22(81.48%) of 27 cases from Xinjiang Uygur Autonomous Region,a province in northwestern China.Phylogenetic analysis on the basis of the K1/VR1 amino acid sequence indicated that the majority of these KS patients were infected by subtype C HHV-8(n=18,including 15 belonging to the C2 group),and several by subtype A(n=4,including 3 being the A1 group).This is the first report of subtype A HHV-8 in China.Furthermore,the correlations between different forms and lesions of KS and different subtypes of HHV-8 were analyzed.The findings showed that subtype A HHV-8 resulted in significantly more frequent mucosal KS lesions than subtype C.However,there was no obvious correlation between different forms of KS and different subtypes of HHV-8.
基金Supported by Grant from the National Natural Science Foundation of China(30100160,30271179)
文摘Objective:To study and confirm that recombinant cytokines similar to those produced by HIV-1 infected T cells induced lytic cycle replication of human pervirus 8(HHV-8) in BC-3 cells,another cell line from primary effusion lymphoma(PEL).Methods:The persistent stimulation of BC-3 was conducted by several cytokines known to be produced by HIV-1-infected T cells and important in grouth and proliferation of Kaposi's sarcoma(KS) cells in vitro,such as the inter feron-γ（IFN-γ）,the hepatocyte grouth factor /scatter factor (HGF/SF),the Oncostain M(OSM),and the tumor necrosis factor-α（TNF-α）which is not produced by HIV-1-infeted T cells.Treated and antrented BC-3 cells were collected at the 3rd and 7th day of persistent stimulation,respeclively.Immunohistochemical(IHC) staining. Northern blot,quantitative PCR(real-time PCR) and electron microscopy(EM) were carried out to detect the expression of immumogenic protein ORF59,messenger RNA(mRNA) of minor capsid protein ORF26,and the presence of viral particles of HHV-8 from treated and untreated BC-3 cells.Results:It showed that IFN-γ,HGF/SF/OSM,and TNF-α were found to induce an increase in mRNA expression of ORF26 when added individually to BC-3 cells.Particularly,ORF26 expression stimulated with IFN-γ and TNF-α respectively,increased 6.1 and 2.5-fold(from,real-time PCR results) at the 7th day when compared with untreated BC-3 cells.Meanwhile ,about 20% of IFN-γ stimulated BC-3 cells expressed ORF59 at the 7th day as compared with 1.5% of untreated BC-3 cells when IHC staining was employed.In addition,viral particles of HHV-8 were readily idelified in BC-3 cells stimulated with IFN-γ at the 7th day with EM analysis.Conclusion:TNF-α and recombinant cytokines being similar to those produced by HIV-1 infected T Cells could really induce HHV-8 lytic cycle replication in BC-3 cells,another cell line of PEL.
文摘Objective: To investigate the effect of activated protein C (APC) on inflammatory responses in human umbilical vein endothelial cells (HUVEC) stimulated with lipopolysaccharide (LPS). Methods: The second passage of collagenase digested HUVEC was divided into the following groups: serum free medium control group (SFM control), phosphate buffer solution control group (PBS control), LPS group with final concentration of 1 μg/ml (LPS group), APC group with final concentration of 7 μg/ml, Pre-APC group (APC pretreatment for 30 min prior to LPS challenge), and Post-APC group (APC administration 30 min after LPS challenge). Supernatant was harvested at 0, 4, 8, 12 and 24 h after LPS challenge. Interleukin-6 (IL-6) and Interleukin-8 (IL-8) levels were analyzed with ELISA. Cells were harvested at 24 h after LPS challenge, and total RNA was extracted. Mes-senger RNA levels for IL-6 and IL-8 were semi-quantitatively determined by RT-PCR. Results: Compared with control group, IL-6 and IL-8 levels steadily increased 4 to 24 h after LPS stimulation. APC treatment could increase LPS-induced IL-6 and IL-8 production. The mRNA levels of IL-6 and IL-8 exhibited a similar change. Conclusion: APC can further increase the level of IL-6 and IL-8 induced by LPS. The effect of these elevated cytokines is still under investigation.
文摘Although Kaposi sarcoma(KS) has been more traditionally considered an AIDS-defining illness,it may also be seen in individuals on immunosuppresive therapy.We report a case of a patient who presented to the hospital in the setting of increasingly refractory ulcerative colitis.Computed tomography scan of the abdomen was consistent with sigmoid diverticulititis and blood cultures were positive for Klebsiella.After a course of antibiotics with resolution of infection,a colonoscopy was performed to evaluate his diverticulitis and incidentally revealed a new rectal tumor.Immunohistochemistry showed the tumor was consistent with KS,with cells staining strongly positive for human herpesvirus-8.This case not only illustrates a rare case of KS found in an HIV-negative individual,but it also highlights the importance of considering an alternative diagnosis in a patient refractory to medical treatment.We discuss the management and care of an ulcerative colitis patient diagnosed with KS on immunosuppressive therapy.
文摘AIM: To investigate the prevalence of human leukocyte antigen (HLA) DQ2/8 alleles in Southern Italians with liver and gastrointestinal (GI) diseases outside of celiac disease. METHODS: HLA DQ2/8 status was assessed in 443 patients from three ambulatory gastroenterology clinics in Southern Italy (University of Federico Ⅱ, Naples, Loreto Crispi Hospital, Ruggi D'Aragona Hospital, Salerno). Patients were grouped based on disease status [pre-post transplant liver disease, esophageal/gastric organic and functional diseases, irritable bowel syndrome (IBS) and inflammatory bowel disease (IBD)] and DQ2/8 alleles, which correspond to a celiac disease genetic risk gradient. Subject allele frequencies were compared to healthy Italian controls. RESULTS: One hundred and ninety-six out of four hundred and forty-three (44.2%) subjects, median age 56 years and 42.6% female, were DQ2/8 positive. When stratifying by disease we found that 86/188 (45.7%) patients with liver disease were HLA DQ2/8 positive, 39/73 (53.4%) with functional upper GI diseases and 19/41 (46.3%) with organic upper GI diseases were positive. Furthermore, 38/105 (36.2%) patients with IBS and 14/36 (38.9%) with IBD were HLA DQ2/8 positive (P = 0.21). Compared to healthy controls those with functional upper GI diseases disorders had a 1.8 times higher odds of DQ2/8 positivity. Those with liver disease had 1.3 times the odds, albeit not statistically significant, ofDQ2/8 positivity. Both those with IBS and IBD had a lower odds of DQ2/8 positivity compared to healthy controls. CONCLUSION: The proportion of individuals HLA DQ2/8 positive is higher in those with liver/upper functional GI disease and lower in IBS/IBD as compared to general population estimates.
基金This work was supported by Nature and Science Foundation Grant of Xinjiang Uygur Autonomous Region(200421122)Collage Grant for Innovate Research Group of Xinjiang Uygur autonomous Region(XJEDU2004G10).
文摘To establish a sensitive and specific method for seroepidermiological detection of human her- pes virus 8(HHV-8)infection,three potent antigenic proteins encoded by open reading frames(ORFs) K8.1,65 and 73C in genome of HHV-8 were produced as glutathione S-transferase fusion protein in the prokaryotic expression system and was used as antigen for testing.The recombinant fusion protein ex- pressed in the prokaryotic expression vector E.coli BL21 was purified by glutathione Sepharose 4B affin- ity chromatography and was quantitated with SDS-PAGE.All these 3 fusion proteins produced in the pro- karyotic expression system showed good immunogenicity as demonstrated by Western blotting and could be recognized by mixed sera of patients with Kaposi′s sarcoma(KS).The immuno-reactivities of the single or compound fusion protein were determined by means of ELISA and compared with the traditional immu- nofluorescence assay(IFA)to determine their sensitivity and specificity of the test.It was demonstrated that the sensitivity of mixed-antigen ELISA method was significantly higher than that of IFA(81.8% vs 34.4%),while the specificity of the former was demonstrated to be 97.9%.The coincidence of the de- tection rate between these two methods was considerably high,approaching up to 90.0%.These results suggest that the mixed antigen ELISA assay appears to be a sensitive and specific method for sero-epide- miological detection of human herpesvirus 8 infection.
文摘Despite the progress has been reached with Human herpesvirus 8 (HHV-8) research, there are gaps in the knowledge of viral induced oncogenesis. The aim of the present study was to identify possible associations between HHV-8 subtypes, HHV-8 loads and clinical manifestations of HIV infected patients diagnosed with different malignancies associated with HHV-8 infection. Forty six HIV-1 infected individuals diagnosed with different HHV-8 associated diseases were studied [37 epidemic Kaposi’s sarcoma (KS), 3 pleural effusion lymphoma (PEL);5 peripheral lymphadenopathies (PL);1 Hodgkin’s lymphoma (HL);1 non Hodgkin’s lymphoma (NHL)]. HHV-8 loads were determined by quantitative real time PCR (qRT-PCR) whilst HHV-8 subtypes were determined by open-reading frame (ORF)-K1 gen genotyping. HHV-8 subtypes B, A, C, A5 and E were exhibited by 31.8%, 23.4%, 19.1%, 17% and 8.5% of the studied patients, respectively. The median HHV-8 viral load did not differ between subtypes (p > 0.05) but HHV-8 viral loads were significantly higher in PEL than in epidemic KS lesion or lymph nodes (p = 0.04). Subtype B was detected in 60% of patients with B cell lymphoma (NHL, PEL and HL) whereas subtype E was only detected in patients with epidemic KS diagnosis. Our data suggest that HHV-8 DNA quantification instead of subtype identification could be used as a surrogate marker for monitoring its infection, not only in epidemic KS patients but also in HIV infected individuals with lymphoproliferative disorders.
