Abstract
Objective: Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is an effective treatment for hematological diseases, but the difficulty in finding a matched donor limits its application. Cord blood transplantation (CBT) and haploidentical hematopoietic stem cell transplantation (haplo-HSCT) serve as alternative options. Although they have their own advantages, they also face challenges such as delayed engraftment and a high incidence of graft versus host disease (GVHD). Among them, GVHD, especially chronic graft versus host disease (cGVHD), is the main cause of transplantation failure and death. The existing immunosuppressive regimens have limited efficacy and significant side effects. In recent years, studies have found that interleukin-17A (IL-17A) is closely related to the fibrotic pathology of cGVHD, and its level is positively correlated with the severity of the disease. However, the mechanism of its action in GVHD remains controversial. In this study, through the intervention of an anti-IL-17A monoclonal antibody and combined with humanized mouse models, we explored the role and therapeutic potential of the anti-IL-17A monoclonal antibody in cGVHD, aiming to provide a new perspective for optimizing the prevention and treatment strategies of GVHD.
Methods:
After the NCG mice were pretreated with X-ray total body irradiation, a certain dose of cord blood cells was infused to establish a humanized mouse model of acute graft versus host disease (aGVHD). Under the same pretreatment conditions, human peripheral blood mononuclear cells (PBMCs) with T cells removed were infused to establish a humanized mouse model of cGVHD. The clinical manifestations of the mice after transplantation were observed for GVHD scoring; HE staining was used for pathological scoring of the damage to the GVHD target organs (lungs, small intestine, skin, liver); flow cytometry was used to monitor the chimerism to evaluate the engraftment of donor cells in the mice.
Using the humanized mouse GVHD model, the recipient mice were intraperitoneally injected with the anti-IL-17A monoclonal antibody or an equal volume of normal saline at the corresponding frequency. The severity of acute and chronic GVHD in the mice of the anti-IL-17A monoclonal antibody group was compared with that in the mice of the normal saline control group to explore the effect of the anti-IL-17A monoclonal antibody on GVHD.
The cytokine microsphere multiplex assay technology was used to detect the levels of IL-2, IL-4, IL-6, IL-10, IL-17, IFN-γ, and TNF in the plasma of the mice after transplantation. Immunohistochemistry was used to analyze the expression of IL-6, IL-10, FN, Foxp3, STAT3, etc. in the target tissues to explore the mechanism of action of the anti-IL-17A monoclonal antibody in regulating cGVHD.
Results:
Humanized NCG mouse models of aGVHD and cGVHD were successfully established. aGVHD model: After total body irradiation with 2.0 Gy X-rays, the transplantation of 3×10^6 human cord blood cells could induce aGVHD. cGVHD model: After total body irradiation with 2.0 Gy X-rays, the transplantation of 0.5×10^6 human PBMCs with T cells removed could induce cGVHD. The recipient mice showed typical GVHD manifestations such as weight loss, hunchback, hair loss, diarrhea, and reduced activity. Histopathological analysis of the GVHD target organs showed inflammatory infiltration and structural damage.
Administration of the anti-IL-17A monoclonal antibody had no obvious effect on aGVHD in humanized mice. However, for cGVHD in humanized mice, the anti-IL-17A monoclonal antibody could reduce its severity, prolong the survival time of the mice, reduce the degree of weight loss in the mice, and had no effect on the engraftment of donor cells.
In the mice of the anti-IL-17A monoclonal antibody administration group after transplantation, the level of IFN-γ in the plasma was decreased, the level of IL-6 in the GVHD target tissues was decreased, the level of IL-10 was increased, the proportion of Treg cells was higher, and the expression of STAT3 was decreased.
Conclusions:
The anti-IL-17A monoclonal antibody has no obvious effect on aGVHD in humanized mice.
The anti-IL-17A monoclonal antibody can reduce cGVHD in humanized mice. Its mechanism may be to reduce fibrosis by down-regulating the levels of inflammatory factors, up-regulating the levels of anti-inflammatory factors, promoting the expansion of Treg cells, and reducing the production of STAT3.
