Caspase 3 may participate in the anti-tumor immunity of dendritic cells

https://doi.org/10.1016/j.bbrc.2019.02.081Get rights and content

Highlights

  • Caspase 3 overexpression only slightly influenced apoptosis of DC2.4 cells.

  • Antigen uptake, maturation and T cell activation of DC2.4 cells were enhanced.

  • Finally, antitumor immunity of DC2.4 cells were enhanced.

Abstract

Background

Caspase 3 is not only involved in apoptosis, but also participates in the nonapoptotic functions. Previously, we found that caspase 3 gene knockout mice displayed decreased number of dendritic cells (DCs). However, whether caspase 3 participate in the function of DCs is unclear. Thus, the present study aims to investigate the role of caspase 3 in the maturation and antitumor function of DCs.

Methods

Caspase 3 gene was overexpressed in DC2.4 cell line through Lentivirus system. The impact of caspase 3 gene overexpression on the biological behavior of DC2.4 cells was determined by CCK-8, colony formation and apoptosis analysis. The impact of caspase 3 gene overexpression on the antigen uptake, maturation, migration, T cell activation of DC2.4 cells was analyzed with phagocytosis, transwell and mixed lymphocyte reaction assay. Tumor growth and tumor infiltrated T cells were also investigated through tumor bearing model.

Results

Caspase 3 gene overexpression could slightly increase the apoptosis of DC2.4 cells. Antigen uptake capability and maturation of DC2.4 cells were significantly promoted through caspases 3 gene overexpression. However, CXCR4 expression on DC2.4 cells and migration of DC2.4 cells were not influenced. Caspase 3 gene overexpression also enhanced the T cell activation and cytotoxicity of activated T cells. Finally, overexpression of caspase 3 gene significantly increased the tumor suppression of DC2.4 cells, accompanied by increased infiltration of CD4+ and CD8+ Cells in tumor tissue.

Conclusion

Caspase 3 gene overexpression could promote maturation and enhance antitumor capability of DC2.4 cells.

Introduction

As the sentinel of immune system and the most potent professional specific antigen presenting cells, DCs play critical roles in the immunosurveillance in vivo [1]. Although the quantity is small, DCs spread throughout the body, including tissues and circulation [2], constantly monitoring their surroundings for different antigens [3]. In the steady state, DCs are largely immature [4], which is characterized by low expression of major histocompatibility complex (MHC) II, co-stimulatory molecules (CD80 and CD86) and chemokine receptor, and high capacity of antigen capture [5]. After antigen encounter and uptake, immature DCs become mature and the co-stimulatory molecules were upregulated [6]. Then, they migrate into secondary lymphoid organs to initiate immune responses [7].

Caspases are proteolytic enzymes which mediate the programmed cell death known as apoptosis, and they are highly conserved among different species [8]. The family of caspases can be further classified as initiator (caspases 8, 9, 10) and executioner (caspases 3, 6, 7) [9]. In principle, caspases could be activated through either extrinsic or intrinsic signal transduction pathways [10]. Upon activation, the initiator cleaves the executioner. These proteases then degrade structural substrates and DNA repair enzymes, etc. Among these executioners, caspase 3 is extremely important as both intrinsic and extrinsic pathways converge at caspase 3 [11]. In addition to the classical role in apoptosis, emerging evidence indicates that caspase 3 also execute nonapoptotic functions [12]. However, the regulation of caspase 3 on the differentiation and function of DCs has not been fully elucidated yet.

Previously, we found that the number of CD11c+MHCII+ dendritic cells was significantly decreased in caspase 3 gene knockout mice [13]. Thus, we speculate that caspase 3 may participate in the regulation of maturation and antitumor function of DCs. The present study aims to investigate the role of caspase 3 on the antitumor function of DCs.

Section snippets

Mice and cells

C57BL/6 Mice (male, 7–8 weeks old) were obtained from Animal Recourses Center of the Fourth Military Medical University. All mice were maintained in the specific pathogen-free (SPF) conditions. All animal experiments were approved by the Animal Experiment Administration Commission of Fourth Military Medical University.

DC2.4 cell line [14] was purchased from Huiying biological technology (China). Mouse melanoma (B16) cell line was gift from Dr. Y. Z. Chen. All the cells were cultured in

Overexpression of caspase 3 gene slightly influenced the biological behavior of DC2.4 cells

To establish a cellular model to gain of function of Caspase 3 gene, a lentivirus infection system was employed. RT-PCR and western blotting results showed that caspase 3 mRNA levels (Fig. 1A) and protein levels (Fig. 1B and C) significantly increased, demonstrated that the overexpression of Caspase 3 gene in DC2.4 cells were successful.

Since caspase 3 plays a key role in apoptosis, we then analyzed the proliferation and apoptosis profile of DC2.4 cells after lentivirus infection. As shown in

Discussion

DCs were initially reported by Langerhans in 1868 and then identified by Steinman in 1973 [17]. Since methods for in vitro culture of DCs were established and developed, investigation about DCs were largely accelerated [18]. As DCs could initiate specific immune responses against antigens, many studies have focused on the application of DC vaccines to treat cancer. Although DC vaccination is considered to be a potent treatment for tumor immunotherapy, it has not been as effective as expected

Conflicts of interest

The authors declare no conflict of interest.

Acknowledgements

This work was supported by the grants from National Natural Science Foundation of China (No. 31570907, No.81572306).

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