Objective: To investigate the proteins expression difference after upregulation of human CD99 in Hodgkin Lymphoma cell line, L428 cell, and verify the function of differential proteins. Methods: The differential proteins were detected by two-dimensional fluorescence difference gel electrophoresis and mass spectrometry analysis, cluster analysis was done by GOfact. Results: There were 38 proteins screened out, of which 21 proteins were positively associated with CD99, while 17 proteins were negative. Among the 38 proteins, 32 proteins participated in biological process, and 35 proteins were involved in the composition and construction. And 28 proteins participated in multifaceted biological activities including antioxidation, protein binding, catalytic activity, regulation of enzyme, signal transduction, molecular structure, regulation of translation and ion transport. Conclusions: The changes of the differential proteins, correlated with cytoskeleton, cell differentiation, signal pathway and regulating gene expression, are closely relevant to the translation between Hodgkin/Reed-Sternberg and B lymphocyte cell.
12E7 Antigen ; Cell Line, Tumor ; Hodgkin Disease ; Humans ; Proteomics ; Up-Regulation

OBJECTIVETo explore the effect of CD99 overexpression on the morphology and differentiation-related phenotypes of classical Hodgkin's lymphoma cell line L428 and investigate the role of CD99 gene in Hodgkin/Reed-Sternberg (HRS) cell generation and transformation.

METHODSThe effect of CD99 overexpression on the cell morphology was detected by HE staining and phalloidin staining. Differentiation-related protein expressions were detected by immunocytochemistry and flow cytometry after stable transfection of CD99 gene in L428 cells.

RESULTSCD99 overexpression caused a decrease of the cell size and reorganization of the actin cytoskeleton in L428 cells. Upregulation of CD99 led to the loss of classical Hodgkin's lymphoma diagnosis marker CD30 and CD15 and the restoration of the B-cell makers of PAX5, CD19, CD79α, BCL-6, and CD10.

CONCLUSIONCD99 overexpression leads to redifferentiation of L428 cells towards B cells, suggesting that the loss of B-cell phenotype in classical Hodgkin's lymphoma is very likely a result of down-regulated CD99 expression.

12E7 Antigen ; Antigens, CD ; genetics ; B-Lymphocytes ; cytology ; Cell Adhesion Molecules ; genetics ; Cell Differentiation ; Cell Line, Tumor ; Gene Expression Regulation, Neoplastic ; Hodgkin Disease ; genetics ; pathology ; Humans

12E7 Antigen ; Antigens, CD ; metabolism ; Cell Adhesion Molecules ; metabolism ; Diagnosis, Differential ; Humans ; Kidney Neoplasms ; metabolism ; pathology ; surgery ; Male ; Middle Aged ; Nephrectomy ; Osteosarcoma ; metabolism ; pathology ; surgery ; Vimentin ; metabolism

12E7 Antigen ; Antigens, CD ; metabolism ; Antigens, CD34 ; metabolism ; Cell Adhesion Molecules ; metabolism ; Diagnosis, Differential ; Head and Neck Neoplasms ; metabolism ; pathology ; Hemangioma ; pathology ; Humans ; Lipoma ; metabolism ; pathology ; Liposarcoma, Myxoid ; pathology ; Male ; Middle Aged ; S100 Proteins ; metabolism ; Soft Tissue Neoplasms ; metabolism ; pathology

12E7 Antigen ; Aged ; Antigens, CD ; metabolism ; Carcinoma, Transitional Cell ; pathology ; Carcinosarcoma ; pathology ; Cell Adhesion Molecules ; metabolism ; Cystectomy ; methods ; Diagnosis, Differential ; Follow-Up Studies ; Humans ; Male ; Nephrectomy ; Osteosarcoma ; metabolism ; pathology ; surgery ; Ureter ; surgery ; Ureteral Neoplasms ; metabolism ; pathology ; surgery ; Vimentin ; metabolism

12E7 Antigen ; Adult ; Antigens, CD ; metabolism ; Cell Adhesion Molecules ; metabolism ; Diagnosis, Differential ; Humans ; Immunohistochemistry ; Kidney ; metabolism ; pathology ; Kidney Neoplasms ; metabolism ; pathology ; surgery ; Male ; Nephrectomy ; Neuroectodermal Tumors, Primitive ; metabolism ; pathology ; surgery ; Vimentin ; metabolism ; Wilms Tumor ; pathology

12E7 Antigen ; Adult ; Antigens, CD ; metabolism ; Cell Adhesion Molecules ; metabolism ; Chondrosarcoma, Mesenchymal ; metabolism ; pathology ; surgery ; Diagnosis, Differential ; Hemangiopericytoma ; pathology ; Humans ; Lymphoma ; pathology ; Male ; Nasal Cavity ; Neuroectodermal Tumors, Primitive ; pathology ; Nose Neoplasms ; metabolism ; pathology ; surgery ; Vimentin ; metabolism ; Young Adult

12E7 Antigen ; Antigens, CD ; metabolism ; Bone Neoplasms ; metabolism ; pathology ; surgery ; Cell Adhesion Molecules ; metabolism ; Chondrosarcoma, Mesenchymal ; metabolism ; pathology ; surgery ; Diagnosis, Differential ; Hemangiopericytoma ; pathology ; Humans ; Lung Neoplasms ; metabolism ; pathology ; surgery ; Male ; Middle Aged ; Pneumonectomy ; methods

12E7 Antigen ; Antigens, CD ; metabolism ; CD57 Antigens ; metabolism ; Cell Adhesion Molecules ; metabolism ; Female ; Humans ; Immunohistochemistry ; Neuroectodermal Tumors, Primitive, Peripheral ; metabolism ; pathology ; therapy ; Pancreas ; chemistry ; pathology ; surgery ; Pancreatectomy ; methods ; Pancreatic Neoplasms ; metabolism ; pathology ; surgery ; Phosphopyruvate Hydratase ; metabolism ; Young Adult

OBJECTIVETo construct a eukaryotic expression vector of CD99 gene for transfection into Hodgkin lymphoma L428 cells.

METHODSThe full-length cDNA of CD99 gene was amplified from Jurkat cells by RT-PCR and cloned into the pcDNA3.1(+) vector and transfected into L428 cell line using Lipofextamine 2000. The sequence of CD99 mRNA in the transfected cells was confirmed by restriction endonuclease digestion and DNA sequencing, and the expression of CD99 protein was identified using immunocytochemistry.

RESULTSA gene fragment of 558 bp was amplified from the transfected cells and the sequence was verified by DNA sequencing. Immunocytochemistry identified the presence of CD99 expression in the transfected cells.

CONCLUSIONA eukaryotic expression vector pcDNA3.1(+)-CD99 is successfully constructed and stably expressed in L428 cell line.

12E7 Antigen ; Antigens, CD ; biosynthesis ; genetics ; Base Sequence ; Cell Adhesion Molecules ; biosynthesis ; genetics ; Cell Line, Tumor ; Cloning, Molecular ; Genetic Vectors ; genetics ; Hodgkin Disease ; metabolism ; pathology ; Humans ; Jurkat Cells ; Molecular Sequence Data ; Transfection