Changes in the expression of CD169 on peripheral blood CD14 +monocytes of rhesus monkeys after SIV infection / 中国比较医学杂志

Objective To investigate the changes of CD169 expression on the surface of peripheral blood monocytes and different subsets of monocytes in normal rhesus monkeys after SIVmac239 infection and the possible reasons.Methods Normal rhesus monkeys were infected with SIVmac239 through intravenous injection, and changes in the percentage of peripheral blood monocytes and the expression of CD169 before and after SIVmac239 infection were detected by flow cytometry. The peripheral blood CD14 +monocytes of normal rhesus monkeys sorted by flow cytometry were directly infected by SIVmac239 and stimulated by different cytokines,and changes in the expression of CD169 on the cell surface and the cytokine IFN-α were detected by flow cytometry. Results After SIVmac239 infection, the percentage of CD14 +monocytes of the normal rhesus monkeys was decreased and the expression of CD169 on their surface was increased. Meanwhile,the expression of CD169 on the surface of different subsets of peripheral blood monocytes was significantly increased,and the expression of CD169 on the CD14 +CD16 + +monocytes was increased more obviously. CD169 was not expressed on the surface of peripheral blood CD14 +monocytes of the normal rhesus monkeys after stimulated by the cytokines M-CSF, IL-4 and IL-13. However, CD169 was highly expressed after the monocytes were stimulated by the cytokine IFN-α. The expression of CD169 on the surface of CD14 +monocytes and the intracellular cytokine IFN-α was not significantly changed after the monocytes were directly infected with SIVmac239. Conclusions SIVmac239 infection can lead to the increase of CD169 expression on the surface of peripheral blood monocytes in rhesus monkeys. Its expression is not associated with the direct infection of virus,but is related to the cytokine IFN-α secreted by other cells of the monkeys in vivo.

Establishment and application of a new immune-comb assay for detection of serum antibody against simian immunodeficiency virus / 中国实验动物学报

Objective SIV30 protein of simian immunodeficiency virus(SIV)was prepared by genetic engineering technique as an antigen diagnostic reagent, to establish an immune comb method for the specific detection of anti SIV IgG in monkey serum. Methods Recombinant expression plasmid of SIV SIV30 gene was constructed by prokaryotic expression vector pGEX-4T-1, and expressed in the competent BL21 cells. The recombinant protein was purified as a diagnostic antigen, and a standardized procedure for the detection of immune comb was established and applied for clinical detection. Results The optimum coating amount of antigen was 0.02 mg/mL. The prepared IC was able to specifically detect the positive serum of SIV. There was no cross reaction between the sera of other viruses. It showed a high specificity of the detection method. Sensitivity analysis showed that the SIV30 protein was able to detect 1:400 times diluted SIV positive sera. The result of stability and repeatability test(the same sample was repeated 3 times) showed that the coefficient of variation(CV)was less than 10%. The serum samples of 10 suspicious monkeys were detected by this method, showing a consistent rate of comb method and ELISA test result of 100%, Kappa =1.000. Conclusions SIV30 protein is expressed in prokaryotic cells. The immune comb is prepared,and is successfullyl applied in clinical examination. It shows that the method has a high sensitivity, strong specificity, good reproducibility and practicability.

Dynamic monitoring of virus antibody during the establishment of SPF cynomolgus monkey population / 中国实验动物学报

Objective To investigate the positive and suspicious rates of BV, SRV, SIV and STLV-1 antibody of the cynomolgus monkeys introduced from Laos, dynamic monitoring of virus antibody during the establishment of SPF cyno-molgus monkey population, and then comparing the positive rate of virus antibody in the baby monkeys between the common with SPF populations.Methods Continuous monitoring four kinds of virus in the monkeys was carried out using special reagent kits and analyzed the data.Results In the introduced cynomolgus monkeys (n=1998), the positive rate of BV antibody was 52.35%, suspicious rate of BV antibody was 8.31%, and negative rate of BV antibody was only 39.34%, the positive rates of SRV and STLV-1 antibody were 7.45%and 8.56%, respectively, and positive or suspicious SIV anti-body in the cynomolgus monkeys was not detected.After screening and establishment of the SPF population, the positive rates of BV, SRV and STLV-1 antibody were 5.24%, 1.01%and 0.4%, respectively, in the year 2010.Then after five years of screening and elimination, by the end of 2014, the positive rates of BV, SRV and STLV-1 antibodies were 0.82%, 0.27%and 0.27%, respectively, and positive or suspicious SIV antibody in the cynomolgus monkeys was not de-tected.The BV antibody positive rate of the baby monkeys was 9.71%and the suspicious rate was 1.85%in the common population, but only 0.22%of the baby monkeys in the SPF population.Conclusions Continuous monitoring virus anti-bodies and eliminating positive and suspicious animals have important significance in the establishment of a SPF cynomolgus monkey population.

Comparative analysis of Gp120 sequences of enterotropic and neurotropic SIV strains / 中国比较医学杂志

Objective To study the mutations of Env sequence of SIVmac239 after infection of Chinese rhesus monkeys, and compare the differences and characteristics of Gp120 sequences of enterotropic and neurotropic SIV strains. Methods Six strains of simian immunodeficiency virus were analyzed in this study: four separated from peripheral blood mononuclear cells of SIVmac239-infected monkeys and two neurotropic SIVmac251 strains.Isolated and cultured monoclonal virus was obtained by limiting dilution assay.Gp120 sequences were amplified after the RNA extraction and phylogenetic analysis was processed thereafter.So did the Gp120 amino acid sequence and N-glycosylation sites analysis of the enterotropic and neurotropic strains.Results SIVmac239 had different mutations in four rhesus monkeys.The diversity in amino acid sequences of the enterotropic and neurotropic strains concentrated in the V1 and V4 regions of Gp120.The enterotropic strains had an addition of glycosylation site in V4 but the glycosylation site changes of neurotropic strains were located in the conservative regions of C1, C2 and C3.Conclusions The addition of one glycosylation site in V4 region of GP120 and loss of one glycosylation site in C1 region are associated with enhanced enterotropism and neurotropism.The differences between the enterotropic and neurotropic strains are not dipicted in Gp120 V3 region which is closely related with the tropism of strains.

Comparison of bio-medical parameters in SIV infected Chinese rhesus monkeys with diverse progression correlated to the pathogenesis of simian AIDS / 中华微生物学和免疫学杂志

Objective To compare the bio-medical parameters in SIV infected Chinese rhesus monkeys with diverse disease progression,by which the pathogenesis of simian AIDS were to be investigated.Methods Sixteen Chinese rhesus monkeys were inoculated intravenously with SIVmac239 and followed-up for 18 months.Based on their progression patterns and plasma viral loads,animals were divided into 3 groups,including 1 rapid progressor( RP),13 normal progressors(NP),and 2 elite controllor(EC).Their parameters of haematology,virology,immunology and pathology were examined and compared. Results Compared with other animals,RM449(RP) showed higher viral load,unresponsive humoral immunity,and higher level of auto-antibodies against lymph node,thymus,and spleen.Additionally,its effector memory CD4 count was lower,with the transformation progress being blocked-like from naive/central memory subsets to effector memory subset,as the flow-cytometry assay showed.Notable decrease in its peripheral B cell was also observed,especially to the sub-population of tissue-like memory B cells and activated memory B cells.Pathological examination showed the depletion of lymphoid tissue,atrophy of spleen and loss of thymus.Moreover,most of these parameters of RM450 and RM453 (EC) changed opposite to that of RP.Conclusion The hallmarks of RM449 were higher viraemia and lower SIV specific IgG level,which may due to the disturbance of T cells and B cells development and differentiation.Moreover,destructions of organs of the immune system may contribute to the disturbance.Our study suggest that the change of micro-environments of thymus induced by SIV infection,which is necessary in T cell and B cell development and differentiation,may contribute at least partially to the AIDS pathogenesis.

The Comparison of Genetic Variation in the Envelope Protein Between Various Immunodeficiency Viruses and Equine Infectious Anemia Virus / 中国病毒学·英文版

The envelope protein(Env) of lentiviruses such as HIV,SIV,FIV and EIAV is larger than that of other retroviruses.The Chinese EIAV attenuated vaccine is based on Env and has helped to successfully control this virus,demonstrating that envelope is crucial for vaccine.We compared Env variation of the four kinds of lentiviruses.Phylogenetic analysis showed that the evolutionary relationship of Env between HIV and SIV was the closest and they appeared to descend from a common ancestor,and the relationship of HIV and EIAV was the furthest.EIAV had the shortest Env length and the least number of potential N-linked glycosylation sites(PNGS) as well as glycosylation density compared to various immunodeficiency viruses.However,HIV had the longest Env length and the most PNGS.Moreover,the alignment of HIV and SIV showed that PNGS were primarily distributed within extracellular membrane protein gp120 rather than transmembrane gp41.It implies that the size difference among these viruses is associated with a lentivirus specific function and also the diversity of env.There are low levels of modification of glycosylation sites of Env and selection of optimal protective epitopes might be useful for development of an effective vaccine against HIV/AIDS.

Influence of NO2 and metal ions on oxidation of aqueous-phase S(IV) in atmospheric concentrations

An investigation was made of the influence of atmospheric concentrations (15 or 130 ppbv) of NO2 on the aqueous-phase oxidation rate of S(IV) in the presence and absence of Fe(III), Mn(II) and Cr(VI) metal ions under controlled experimental conditions (pH, T, concentration of reactants, etc.). The reaction rate in the presence of the NO2 flow was slower than the reaction rate using only clean air with an initial S(IV) concentration of 10-4 mol/L. NO2 appears to react with S(IV), producing a kind of inhibitor that slows down the reaction. Conversely, tenfold lower concentrations of S(IV) ([S(IV)]º = 10-5 mol/L) caused a faster reaction in the presence of NO2 than the reaction using purified air. Under these conditions, therefore, the equilibrium shifts to sulfate formation. With the addition of Fe(III), Mn(II) or Cr(VI) in the presence of a NO2 flow, the reaction occurred faster under all the conditions in which S(IV) oxidation was investigated.

A reação de oxidação de S(IV) em fase aquosa foi estudada em laboratório em presença de NO2 dos íons metálicos Fe(III), Mn(II), e Cr(VI) sob condições experimentais controladas (pH, T, concentração dos reagentes, etc.). Na presença de corrente de ar com NO2 (15 ou 130 ppbv) a reação de oxidação de S(IV) ocorreu mais lentamente do que na presença de ar purificado, para uma concentração inicial de S(IV) de 10-4 mol/L. Ao contrário, para concentração inicial de S(IV) dez vezes menor ([S(IV)]° = 10-5 mol/L) a reação ocorreu mais rapidamente na presença de NO2. A explicação está relacionada com o equilíbrio envolvendo a formação de espécies intermediárias de longa vida, que impedem o prosseguimento da reação, porém a depender das concentrações relativas de S(IV) e NO2, essas espécies se decompõem deslocando o equilíbrio no sentido de formação de sulfato. A adição dos íons Fe(III), Mn(II) ou Cr(VI) em presença de corrente de ar com NO2 indicou atividade catalítica para esses íons, em todas as condições nas quais a oxidação de S(IV) foi investigada.

The hope for an HIV vaccine based on induction of CD8+ T lymphocytes: a review

The only long-term and cost-effective solution to the human immunodeficiency virus (HIV) epidemic in the developing world is a vaccine that prevents individuals from becoming infected or, once infected, from passing the virus on to others. There is currently little hope for an AIDS vaccine. Conventional attempts to induce protective antibody and CD8+ lymphocyte responses against HIV and simian immunodeficiency virus (SIV) have failed. The enormous diversity of the virus has only recently been appreciated by vaccinologists, and our assays to determine CD8+ lymphocyte antiviral efficacy are inadequate. The central hypothesis of a CTL-based vaccine is that particularly effective CD8+ lymphocytes directed against at least five epitopes that are derived from regions under functional and structural constraints will control replication of pathogenic SIV. This would be somewhat analogous to control of virus replication by triple drug therapy or neutralizing antibodies.

Quantification of Simian Immunodeficiency Virus by SYBR Green RT-PCR Technique / 中国病毒学·英文版

Plasma viral RNA load is widely accepted as the most relevant parameter to assess the status and progression of Simian immunodeficiency virus (SIV) infections. To accurately measure RNA levels of the virus, a one-step fluorescent quantitative assay was established based on the SYBR green Real-time reverse transcription-polymerase chain reaction (RT-PCR). The lower detection limit of the assay was 10 copies per reaction for the virus. This method was successfully applied to quantify SIVmac251 and SIVmac239 viruses produced in CEM×174 cells. Additionally, the performance of the SYBR green RT-PCR was assessed in a SIVmac251 infected rhesus macaque. The result demonstrated that the method could detect as little as 215 copies per milliliter of plasma and the dynamic pattern of viral load was highly consistent with previous results. With regard to convenience, sensitivity and accuracy our assay represents a realistic alternative to both branched-chain DNA (b-DNA) assays or real-time PCR assays based on TaqMan probes.

Establishment of AIDS Animal Model with SIVmac239 Infected Chinese Rhesus Monkey / 中国病毒学·英文版

In the present research, two Chinese rhesus monkeys were inoculated intravenously with 5000 TCID50 of SIVmac239. The changes in the numbers of CD4+T lymphocyte in peripheral blood,plasma viral loads, proviral DNA and humoral antibodies against virus were periodically monitored during 121 days. At the early stage of infection, proviral DNA had been detected in PBMCs, and infectious SIVmac239 virus had been isolated from PBMCs. At the same period, the numbers of CD4+T lymphocytes were significantly decreased, and maintained at low level during the 121-day period of infection. Plasma viral loads reached the peak at week 2 post-inoculation and kept at a steady state subsequently. Moreover, antibodies against viral proteins were detected from plasma. All the results showed that the two Chinese rhesus monkeys had been infected with SIVmac239 successfully. This animal model can be applied for further AIDS researches.