Applications Of The Chick Chorioallantoic Membrane As A Novel In Vivo Model For Idiopathic Multicentric Castleman Disease

Background:Castleman disease(CD)includes a group of rare and heterogeneous disorders with characteristic lymph node histopathological abnormalities.CD can occur in a single lymph node station,which is referred to as unicentric CD(UCD).CD can also involve multicentric lymphadenopathy and inflammatory symptoms(multicentric CD,MCD).MCD includes human herpesvirus-8(HHV-8)–associated MCD,and HHV-8-/HIVMCD(idiopathic MCD,i MCD).Patients with the i MCD frequently have systemic manifestations,such as organ insufficiency,edema,effusion,low-grade fever,fatigue,and weight loss.The international consensus published in 2018 recommended anti-IL-6,Rituxiamb and corticosteroids therapy as first line for all patients with i MCD.Chick chorioallantoic membrane(CAM)is the respiratory organ of chick embryo and rich in blood vessels.The CAM is naturally immunodeficient early during hatching and therefore can tolerate the transplantation of human tumor cells.In this model,tumor tissue is inoculated on the chorionic epithelium,for highly visible tumor growth.Furthermore,the model is cost-and time-efficient.Objective:To establish the xenotransplanted tumor model of Castleman disease in CAM and drug validation was performed using methylpredisolone,Rituximab and Tocilizumab.Methods:1.Surgical specimens from patients with i MCD were transplanted on the CAM.The distribution of blood vessels around the xenotransplanted tumor was observed,and the morphology and immunology feature of the xenotransplanted tumor were evaluated.2.The xenograft model of i MCD were treated by methylprednisolone,Rituximab and Tocilizumab;then appearance changes of transplanted tumor were observed.The morphology and immunology feature of the xenotransplanted tumor was performed after the experiment.3.The density of positive cells with CD38,CD20 and CD3 were evaluated after treated with methylprednisolone,Rituximab and Tocilizumab.Results:1.On the first day after transplantation,the transplanted tumor was attached to the CAM of chicken embryo and grew with a little angiogenesis around it;On the 3rd day after transplantation,the number of blood vessels increased and the lumen diameter increased;On the 5th day after transplantation,the growth of blood vessels around the transplanted tumor reached the peak,distributed in a spoke shape,and the blood vessels became thicker and tortuous.2.HE stains revealed no follicles and germinal center structure in the transplanted tumor tissue.Plasma cells and CD38 positive cells were distributed in the transplanted tumor tissue in a patchy manner,CD3 positive cells were scattered,and only a few CD20 positive cells were observed.3.The transplanted tumors were pale pink in the 0.9%Na Cl group and the normal control group whileas the tumor tissues showed dark gray white after treatment with Rituximab,Tocilizumab and methylprednone.HE stains and immunohistochemical staining showed less distribution of various cells,and the area of coagulated necrosis increased in the drug treated groups versus control groups.4.After treatment with methylprednisolone,Tocilizumab and Rituximab,the positive cells density of CD38 and CD3 was significantly lower than that of the0.9%Na Cl group,and the positive cell density of methylprednisolone and Rituximab groups were lower than that of Tocilizumab group.Conclusions:1.The xenotransplanted tumor model of i MCD in CAM was established and maintained morphological and immunological stability,which can be used for drug screening for i MCD.2.Methylprednisolone,Tocilizumab and Rituximab inhibited the growth of CD38 and CD3 positive cells in xenograft with i MCD in the CAM,and the inhibition effect of methylprednisolone and Rituximab is superior to Tocilizumab.