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The role of Qa-2, the functional homolog of HLA-G, in a Behcet's disease-like mouse model induced by the herpes virus simplex.

Lee M, Choi B, Kwon HJ, Shim JA, Park KS, Lee ES, Sohn S - J Inflamm (Lond) (2010)

Bottom Line: In this study, we evaluated the level of Qa-2, a murine nonclassical class I MHC molecule and possible functional homolog of HLA-G, to determine if it was associated with various symptoms of BD-like mice.The silencing of Qa-2 by intravenous injection of siRNA (500 nmol/mouse, 4 times at 3-day intervals) specifically reduced the Qa-2 levels and worsened the BD-like symptoms.Silencing Qa-2 by injecting siRNA into mice resulted in deterioration of symptoms in BD-like mice.

View Article: PubMed Central - HTML - PubMed

Affiliation: Laboratory of Cell Biology, Ajou University Institute for Medical Sciences, Suwon, Korea. sohnsh@ajou.ac.kr.

ABSTRACT

Background: It has been suggested that the HLA-G molecule is a genetic risk factor for Behcet's disease (BD). In this study, we evaluated the level of Qa-2, a murine nonclassical class I MHC molecule and possible functional homolog of HLA-G, to determine if it was associated with various symptoms of BD-like mice. In addition, we investigated siRNA (small interfering RNA) treatment to determine if it inhibited Qa-2 expression, thereby changing the symptoms of mice.

Methods: RNA interference (RNAi) and vector transfection were employed to manipulate gene expression in vivo in mice. siRNA (small interfering RNA) or Qa-2 expression vector was applied to inhibit or up-regulate Qa-2 expression, respectively.

Results: The Qa-2 levels in granulocytes were lower in BD-like mice than in normal controls. The silencing of Qa-2 by intravenous injection of siRNA (500 nmol/mouse, 4 times at 3-day intervals) specifically reduced the Qa-2 levels and worsened the BD-like symptoms.

Conclusions: Silencing Qa-2 by injecting siRNA into mice resulted in deterioration of symptoms in BD-like mice.

No MeSH data available.


Related in: MedlinePlus

Expression of pc3.1DNA Qa-2 vector in vivo in normal mice. The frequency of Qa-2 protein in splenocytes and macrophages isolated from normal mice injected with 50 μg pc3.1DNA Qa-2 vector as determined by FACS analysis. The macrophages isolated from mice injected with pc3.1DNA Qa-2 vector showed a higher frequency of Qa-2 positive cells when compared to the non-injected control (p = 0.01) and pc3.1 DNA vector-injected mice (p = 0.003). The splenocytes also showed a higher frequency of Qa-2 positive cells when compared to the non-injected control and pc3.1 DNA vector injected mice, although this difference was not significant.
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Figure 7: Expression of pc3.1DNA Qa-2 vector in vivo in normal mice. The frequency of Qa-2 protein in splenocytes and macrophages isolated from normal mice injected with 50 μg pc3.1DNA Qa-2 vector as determined by FACS analysis. The macrophages isolated from mice injected with pc3.1DNA Qa-2 vector showed a higher frequency of Qa-2 positive cells when compared to the non-injected control (p = 0.01) and pc3.1 DNA vector-injected mice (p = 0.003). The splenocytes also showed a higher frequency of Qa-2 positive cells when compared to the non-injected control and pc3.1 DNA vector injected mice, although this difference was not significant.

Mentions: To confirm if Qa-2 could influence IFNγ expression, Qa-2 vector was constructed in PC3.1 vector and then administered to normal and BD mice. Cloning of the Qa-2 gene of pGEM-Qa-2 into pcDNA3.1 vector was confirmed by digestion with EcoRI and BamHI (Figure 6A), after which the inserted sequence was confirmed by sequencing using T7 promoter (Figure 6B). The vector was intraperitoneally injected once into mice, and peritoneal macrophages and splenocytes were isolated four days later. As shown in Figure 7, the frequency of Qa-2 expressing cells in splenocytes increased to 94.53 ± 0.64% in the Qa-2 vector injected mice, while it was 89.83 ± 2.66% in control vector injected mice (p = 0.45). Additionally, their frequency in macrophages increased to 82.25 ± 5.62% in the Qa-2 vector injected mice, while it was 67.53 ± 4.66% in control vector injected mice (p = 0.003). The IFNγ levels in macrophages of Qa-2 vector-injected mice also decreased to 16.60 ± 6.11%, while they were 66.24 ± 7.28% in control mice (p < 0.001) (Figure 8). Qa-2 expression vector appeared to work in macrophages, and these effects were accompanied by a decrease in IFNγ.


The role of Qa-2, the functional homolog of HLA-G, in a Behcet's disease-like mouse model induced by the herpes virus simplex.

Lee M, Choi B, Kwon HJ, Shim JA, Park KS, Lee ES, Sohn S - J Inflamm (Lond) (2010)

Expression of pc3.1DNA Qa-2 vector in vivo in normal mice. The frequency of Qa-2 protein in splenocytes and macrophages isolated from normal mice injected with 50 μg pc3.1DNA Qa-2 vector as determined by FACS analysis. The macrophages isolated from mice injected with pc3.1DNA Qa-2 vector showed a higher frequency of Qa-2 positive cells when compared to the non-injected control (p = 0.01) and pc3.1 DNA vector-injected mice (p = 0.003). The splenocytes also showed a higher frequency of Qa-2 positive cells when compared to the non-injected control and pc3.1 DNA vector injected mice, although this difference was not significant.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2902457&req=5

Figure 7: Expression of pc3.1DNA Qa-2 vector in vivo in normal mice. The frequency of Qa-2 protein in splenocytes and macrophages isolated from normal mice injected with 50 μg pc3.1DNA Qa-2 vector as determined by FACS analysis. The macrophages isolated from mice injected with pc3.1DNA Qa-2 vector showed a higher frequency of Qa-2 positive cells when compared to the non-injected control (p = 0.01) and pc3.1 DNA vector-injected mice (p = 0.003). The splenocytes also showed a higher frequency of Qa-2 positive cells when compared to the non-injected control and pc3.1 DNA vector injected mice, although this difference was not significant.
Mentions: To confirm if Qa-2 could influence IFNγ expression, Qa-2 vector was constructed in PC3.1 vector and then administered to normal and BD mice. Cloning of the Qa-2 gene of pGEM-Qa-2 into pcDNA3.1 vector was confirmed by digestion with EcoRI and BamHI (Figure 6A), after which the inserted sequence was confirmed by sequencing using T7 promoter (Figure 6B). The vector was intraperitoneally injected once into mice, and peritoneal macrophages and splenocytes were isolated four days later. As shown in Figure 7, the frequency of Qa-2 expressing cells in splenocytes increased to 94.53 ± 0.64% in the Qa-2 vector injected mice, while it was 89.83 ± 2.66% in control vector injected mice (p = 0.45). Additionally, their frequency in macrophages increased to 82.25 ± 5.62% in the Qa-2 vector injected mice, while it was 67.53 ± 4.66% in control vector injected mice (p = 0.003). The IFNγ levels in macrophages of Qa-2 vector-injected mice also decreased to 16.60 ± 6.11%, while they were 66.24 ± 7.28% in control mice (p < 0.001) (Figure 8). Qa-2 expression vector appeared to work in macrophages, and these effects were accompanied by a decrease in IFNγ.

Bottom Line: In this study, we evaluated the level of Qa-2, a murine nonclassical class I MHC molecule and possible functional homolog of HLA-G, to determine if it was associated with various symptoms of BD-like mice.The silencing of Qa-2 by intravenous injection of siRNA (500 nmol/mouse, 4 times at 3-day intervals) specifically reduced the Qa-2 levels and worsened the BD-like symptoms.Silencing Qa-2 by injecting siRNA into mice resulted in deterioration of symptoms in BD-like mice.

View Article: PubMed Central - HTML - PubMed

Affiliation: Laboratory of Cell Biology, Ajou University Institute for Medical Sciences, Suwon, Korea. sohnsh@ajou.ac.kr.

ABSTRACT

Background: It has been suggested that the HLA-G molecule is a genetic risk factor for Behcet's disease (BD). In this study, we evaluated the level of Qa-2, a murine nonclassical class I MHC molecule and possible functional homolog of HLA-G, to determine if it was associated with various symptoms of BD-like mice. In addition, we investigated siRNA (small interfering RNA) treatment to determine if it inhibited Qa-2 expression, thereby changing the symptoms of mice.

Methods: RNA interference (RNAi) and vector transfection were employed to manipulate gene expression in vivo in mice. siRNA (small interfering RNA) or Qa-2 expression vector was applied to inhibit or up-regulate Qa-2 expression, respectively.

Results: The Qa-2 levels in granulocytes were lower in BD-like mice than in normal controls. The silencing of Qa-2 by intravenous injection of siRNA (500 nmol/mouse, 4 times at 3-day intervals) specifically reduced the Qa-2 levels and worsened the BD-like symptoms.

Conclusions: Silencing Qa-2 by injecting siRNA into mice resulted in deterioration of symptoms in BD-like mice.

No MeSH data available.


Related in: MedlinePlus