Effect of Leukapheresis on Gene Expression Profiles of Donor's Peripheral Blood Mononuclear Cells / 대한진단검사의학회지

BACKGROUND: Leukapheresis has commonly been used to obtain the cell products intended for clinical cell therapy. Hypocalcemia related to citrate toxicity and some circulatory effects such as hypovolemia and hypotension are well-known complications of leukapheresis. In this study, we analyzed the gene expression profiles of peripheral blood mononuclear cells (PBMCs) obtained before and after leukapheresis to determine if the hemodynamic changes can affect the gene expression profiles of leukocytes. METHODS: PBMCs were isolated from EDTA blood from 5 healthy donors collected before and immediately after apheresis. RNA was isolated, amplified, and analyzed using a cDNA microarray with 17,500 genes. Hierarchical clustering analysis was performed to evaluate the differences of gene expression profiling. RESULTS: Hierarchical clustering separated PBMCs from different donors with each other, but did not separate PBMCs collected before and after leukapheresis. Comparison of gene expression by PBMCs collected before and after leukapheresis found only 25 genes were differentially expressed (15 were up-regulated and 10 were down-regulated after leukapheresis) (F-test, P<0.005). Stress induced apoptosis-related genes, ANXA3, DEDD, and ATXN2L, and cytokine-related genes, IL13RA1 and IK, which were also related to stress, were up-regulated after leukapheresis. Genes involved in DNA and protein binding, such as CLSTN3, LRBA, SATB2, and HSPA8, were down-regulated. CONCLUSIONS: Leukapheresis had little effect on gene expression of PBMCs. Some genes showing differences between before and after leukapheresis were mainly involved in stress-related reactions.

Cell Processing: Current Status and Future Directions

Specialized clinical cell processing began in the Department of Transfusion Medicine at the National Institutes of Health in 1984. The number and complexity of procedures performed increased quickly and in 1997 a highly specialized cell processing laboratory was opened. The laboratory has approximately 3,000 square feet, specialized air handing, a highly trained staff, and written laboratory procedures. In addition to standard laboratory equipment, the laboratory has numerous cell isolation instruments, flow cytometers, and automated cell counting instruments. The laboratory supports blood and bone marrow transplant protocols by isolating CD34+ stem cells, removing T lymphocytes, culturing lymphocytes to eliminate donor lymphocytes that are reactive with recipient alloantigens, and stimulating lymphocytes to induce Th2 type cells to reduce graft versus host disease. The laboratory has also been preparing dendritic cells to support protocols using immune therapy to treat cancer. In addition, pancreatic islet cells are isolated from organ donors for transplantation to treat type I diabetes mellitus.

Adoptive Immunotherapy for Cytomegalovirus (CMV) Disease in Immunocompromised Patients

Cytomegalovirus (CMV) reactivation in immune compromised patients such as those undergoing hematopoietic progenitor cell transplantation (HPCT) and those with HIV infections can cause severe morbidity and mortality despite treatment with appropriate antiviral agents. The recovery of Cytomegalovirus (CMV) specific cytotoxic T lymphocytes (CTL) plays an important role in the reconstitution of CMV specific immunity in immunocompromised patients. Recent studies have reported that CMV reactivation can be successfully treated by adoptive transfer of CMV-specific T cell clones from CMV seropositive donors expanded in vitro with CMV infected fibroblasts or lysates of CMV infected cells. Other studies have used immune dominant CMV proteins or peptides to expand CMV-specific cytotoxic T lymphocytes. This review describes the clinical manifestations of CMV disease in immunocompromised patients, recent advances of antiviral therapy for CMV disease, the principals of the induction of cellular immune response to CMV, and the clinical application of CMV immunotherapy.