The dynamic impact of hydrodynamic gene transfer on the immune system

Abstract

Hydrodynamic gene transfer (HGT) has been used as an effective and convenient way to achieve gene expressions in vivo. However, its time-dependent impact on the immune system is unknown. The aim of the current study is to investigate the dynamic changes of the immune parameters after HGT. Plasmids were delivered to BALB/c mice by HGT. Each group of mice was sacrificed on day 1, 2, 3, 4 and 5 after HGT. The immune cell subsets from spleens and livers were analyzed by flow cytometry. IFN-γ, IL-6 and TNF-α in the serum were quantitated by cytometric bead array. The mice without HGT injection were used as control group on day 0. Compared to the normal mice (day 0), the T lymphocyte infiltrations in the spleen and liver were increased starting from day 1 after HGT. T cells. NK cells and myeloid cells such as dendritic cells, neutrophils and macrophages were also significantly expanded and peaked around day 2-3. Both T cells and NK cells were greatly activated. Serum levels of IFN-γ and IL-6 increased and peaked on day 1 after HGT. Most of the increased immune parameters returned to normal levels after day 4. However, the activated T cells remained at a high level, especially in the liver. In conclusion, HGT significantly increased the immune cell infiltration in the spleen and liver and activated T cells and NK cells. The immune response induced by HGT should be taken into consideration when evaluating the functions of the over-expressed genes using this strategy.

Keywords: Hydrodynamic gene transfer; NK cell; T cell; cytokine.

Figure 1

Histology analysis of different organs and T cell percents and numbers in the spleen and liver after HGT. A. HE staining were performed with sections of spleen, liver, lung and kidney (original magnification 200×). B. The percentages of total T, CD4⁺T, CD8⁺T and B cells in both spleen and liver. C. The numbers of total T, CD4⁺T, CD8⁺T and B cells in both spleen and liver. Values are presented as means ± SD. The data shown are the representative of three experiments. *P<0.05, **P<0.01, ***P<0.001.

Figure 2

The CD69 expression on T, CD4⁺T and CD8⁺T cells after HGT. A. The percentages of CD69⁺T, CD69+CD4⁺T, and CD69+CD8⁺T cells in both spleen and liver. B. The absolute numbers of CD69⁺T, CD69+CD4⁺T, and CD69+CD8⁺T cells in both spleen and liver. Values are presented as means ± SD. The data shown are the representative of three experiments. *P<0.05, **P<0.01, ***P<0.001.

Figure 3

The effects of HGT on effector and memory T cell percentages and numbers in both spleen and liver. A. The percentages of spleen and liver effector (CD62L^-CD44^hi) and memory (CD62L⁺CD44^hi) T cells. B. The absolute numbers of spleen and liver effector (CD62L^-CD44^hi) and memory (CD62L⁺CD44^hi) T cells. Values are presented as means ± SD. The data shown are the representative of three experiments. *P<0.05, **P<0.01, ***P<0.001.

Figure 4

The effect of HGT on NK cells in both spleen and liver. A. The percentages and absolute numbers of spleen and liver NK (CD3^-DX5⁺) cells. B. The percentages NKG2D⁺NK, NKP46⁺NK, and CD43+NK in both spleen and liver. C. The absolute numbers of NKG2D⁺NK, NKP46⁺NK, and CD43+NK in both spleen and liver. Values are presented as means ± SD. The data shown are the representative of three experiments. *P<0.05, **P<0.01.

Figure 5

The effect of HGT on myeloid cells in both spleen and liver. A. The percentages of DCs, macrophages and neutrophils in spleen and liver. B. The absolute numbers of DCs, macrophages and neutrophils in spleen and liver. Values are presented as means ± SD. The data shown are the representative of three experiments. *P<0.05, **P<0.01.

Figure 6

The effect of HGT on serum cytokine levels. Serum (A) IFN-γ, (B) IL-6 and (C) TNF-α concentrations were de tected by Flowcytomix kit on different time points after HGT. Values are presented as means ± SD. The data shown are the representative of three experiments. *P<0.05, **P<0.01.