| Background and objective:Allogeneic hematopoietic stem cell transplantation(allo-HSCT)is an important method for the treatment of malignant hematological diseases,and has been widely used in clinical practice.Infection,disease recurrence,graft-versus-host disease(GVHD)and other complications may occur after transplantation.With the deepening of understanding of the disease and early intervention,secondary poor graft function(sPGF)after allo-HSCT has become one of the major complications.The main manifestation is that severe hemoglobin,leukocyte and thrombocytopenia occur again after allo-HSCT hematopoietic reconstruction,leading to severe anemia,severe infection and hemorrhage.It is a fatal and serious complication.Its pathogenesis,disease evolution and prognosis are still unclear.Poor graft function after allogeneic hematopoietic stem cell transplantation has attracted much attention,but there is no uniform definition at home and abroad.People have defined SPGF according to different populations and their own clinical experience.The incidence of SPGF is as high as 5%-27%.Bone marrow hematopoietic microenvironment,also known as niche,can be divided into osteoblastic niche and vascular niche,which play different roles in the proliferation,differentiation,homing and mobilization of HSC,that is,osteoblastic niche is conducive to HSC's resting,and vascular niche is the place of HSC's proliferation and differentiation.Bone marrow stromal cells are important components of hematopoietic microenvironment.They can regulate HSC by secreting cytokines,chemokines and adhesion molecules.Recently,some scholars attempted to prevent poor graft function after allogeneic hematopoietic stem cell transplantation by co-transplantation of endothelial progenitor cell(EPC)/ hematopoietic stem cell(HSC),mesenchymal stem cell(MSC)/ hematopoietic stem cell(HSC)and some achievements have been achieved.Therefore,we speculate that the pathogenesis of poor graft function after allogeneic hematopoietic stem cell transplantation may be related to the disorder of bone marrow stromal cells' distribution,adhesion and secretion ability.At present,the main research methods of stromal cells in sPGF patients after allo-HSCT include direct or cell culture,flow cytometry,PCR and animal experiments.The advantages of these methods are high throughput,many detectable indicators,but can not directly reflect the true state of stromal cells in human bone marrow.Bone marrow pathology can be observed by routine HE staining and immunohistochemical staining of allo-HSCT bone marrow pathological sections of patients with sPGF,to understand the myeloproliferative status,cell distribution,the number of immunohistochemical antibody-labeled cells,expression intensity,distribution status and so on.Therefore,bone marrow pathology has some advantages in the study of stromal cells in sPGF patients after allo-HSCT.The aim of this study was to establish the clinical standard of sPGF after allo-HSCT in our center by retrospective analysis of clinical data,and to detect the molecular CD29,CD44,ICAM-1,VCAM-1 involved in cell adhesion and HSC homing in bone marrow biopsy specimens of allo-HSCT patients before transplantation,and vascular endothelial cell labeling CD34,CD105,connexins43 and other indicators,reveal the role of bone marrow hematopoietic microenvironment in sPGF and related mechanisms,for clinical prevention of allo-HSCT after sPGF to find new targets.Methods:1.The clinical data of 118 patients who underwent allo-HSCT in our department in 2016 were analyzed retrospectively.According to whether peripheral blood cells decreased again after hematopoietic reconstitution,the patients were divided into two groups: sPGF group and good graft function(GGF)group.The age,sex,primary disease,stem cell source,HLA matching,sex matching,ABO matching,CMV infection after transplantation,remission before transplantation,degree of bone marrow hyperplasia before transplantation,number of peripheral blood cells decreased after transplantation were compared between the two groups.After statistical analysis,the clinical standard of sPGF after allo-HSCT was established.2.According to the above criteria,the patients were divided into two groups: poor graft function group and good graft function group.28 patients with poor graft function were selected as experimental group and 27 patients with good graft function as control group.Bone marrow biopsy specimens were taken before transplantation for immunohistochemically staining.The antibodies were CD44,CD105,CD29,ICAM-1,VCAM-1 and Cx43,the value of integrated optical density of the above markers was compared.3.According to the results of IOD comparison,CD34 immunohistochemical staining was used to compare the IOD values of CD34 and CD105 staining,and the microvascular density(MVD)and microvascular morphology of the two groups were observed.Results:1.There were no significant differences in age,sex,primary disease,stem cell source,HLA matching,sex matching,ABO matching,remission before transplantation,degree of myelodysplastic before transplantation between Allo-HSCT group and GGF group(P > 0.05);the two groups of patients compared with CMV infection after transplantation,the difference was statistically significant(P<0.05);the recurrence rate of GGF group was lower than that of sPGF group,and the difference was statistically significant(P < 0.05).There was no significant difference in recurrence rate among patients with 1,2 and 3 line hematocytopenia(P > 0.05).The mortality of GGF group was lower than that of sPGF group,and the difference was statistically significant(P < 0.05),and there was no significant difference in mortality between the 1,2,and 3 line patients with hemocyte depletion(P>0.05).2.The results of immunohistochemical staining showed that the CD105 IOD value [M(Q)] was 32 569.7(26 824.9)in the experimental group and 14 091.1(5 694.1)in the control group,which was significantly higher in the experimental group than in the control group(P < 0.05).Pearson correlation analysis showed that CD105 IOD value was negatively correlated with grouping(r=-0.349,P=0.009).Cx43 IOD value [M(Q)] was 29085.2(39361.9)in the experimental group and 69069.6(118486.45)in the control group,which was significantly lower in the experimental group than in the control group(P < 0.05).Pearson correlation analysis showed that Cx43 IOD was positively correlated with grouping(r=0.314,P=0.000).The values of CD44,CD29,ICAM-1 and VCAM-1 IOD [M(Q)] were 11 476.2(4770.5)to 6 499.0(3 276.1),9 450.5(6 666.8)to 7 284.8(4 159.5),1 061.6(1014.2)to 469.7(372.9),69 258.3(46 892.7)to 51 024.7(38 621.5),respectively,with no significant difference(P > 0.05).3.Immunohistochemical staining showed that the CD105 IOD value [M(Q)] was 32 569.7(26 824.9)in the experimental group and 14 091.1(5 694.1)in the control group,which was significantly higher in the experimental group than in the control group(P < 0.05).Pearson correlation analysis showed that CD105 IOD value was negatively correlated with grouping(r=-0.349,P=0.009).CD34 IOD value [M(Q)] was 4155.6(3986.1)in the experimental group and 4146.85(5964.53)in the control group.There was no significant difference between the two groups(P > 0.05).The results of morphological observation showed that the MVD value of CD105 was 26.6 ± 15.8 in the experimental group and 12.9 ± 11.7 in the control group,which was significantly higher in the experimental group than in the control group(P < 0.05).Pearson correlation analysis showed that CD105 MVD value was negatively correlated with grouping(r=-0.446,P=0.001).The MVD value of CD34 was 7.2(± 2.7)in the experimental group and 5.6(± 2.3)in the control group.There was no significant difference between the experimental group and the control group(P > 0.05).The positive sites of CD105 were located in the cytoplasm and membrane of spindle cells.The distribution of CD105 in the experimental group was disordered,most of them were isolated,and less lumen-like structure was formed.In the control group,the distribution of CD105-positive cells was disordered,but the number of CD105-positive cells was less than the former.The positive sites of CD34 immunohistochemical staining were located in the cytoplasm and membrane of endothelial cells.There were many lumen-like structures in both experimental group and control group.The positive expression of CD34 in vascular endothelial cells was clear.Conclusions:1.The presence of poor graft function after allogeneic hematopoietic stem cell transplantation is an independent risk factor for survival.The presence of at least one line of cytopenia in patients with allo-HSCT is considered to be associated with SPGF.2.Increased expression of CD105 and decreased expression of Cx43 in bone marrow stromal cells before transplantation may be responsible for the occurrence of poor graft function after transplantation.3.Poor graft function may be associated with increased tumor associated endothelial cells in bone marrow before transplantation. |
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