title: Viability and Functionality of Cryopreserved Peripheral Blood Mononuclear Cells in Pediatric Dengue.
abstract: Cryopreserved peripheral blood mononuclear cells (PBMCs) are widely used in studies of dengue. In this disease, elevated frequency of apoptotic PBMCs has been described, and molecules such as soluble tumor necrosis factor (TNF)-related apoptosis-inducing ligands (sTRAIL) are involved. This effect of dengue may affect the efficiency of PBMC cryopreservation. Here, we evaluate the viability (trypan blue dye exclusion and amine-reactive dye staining) and functionality (frequency of gamma interferon [IFN-γ]-producing T cells after polyclonal stimulation) of fresh and cryopreserved PBMCs from children with dengue (in acute and convalescence phases), children with other febrile illnesses, and healthy children as controls. Plasma sTRAIL levels were also evaluated. The frequencies of nonviable PBMCs detected by the two viability assays were positively correlated (r = 0.74; P < 0.0001). Cryopreservation particularly affected the PBMCs of children with dengue, who had a higher frequency of nonviable cells than healthy children and children with other febrile illnesses (P ≤ 0.02), and PBMC viability levels were restored in the convalescent phase. In the acute phase, an increased frequency of CD3+CD8+amine-positive cells was found before cryopreservation (P = 0.01). Except for B cells in the acute phase, cryopreservation usually did not affect the relative frequencies of viable PBMC subpopulations. Dengue infection reduced the frequency of IFN-γ-producing CD3+cells after stimulation compared with healthy controls and convalescent-phase patients (P ≤ 0.003), and plasma sTRAIL correlated with this decreased frequency in dengue (rho = -0.56; P = 0.01). Natural dengue infection in children can affect the viability and functionality of cryopreserved PBMCs.
authors.affiliation: Programa de Medicina, Facultad de Salud, Universidad Surcolombiana, Neiva, Colombia.
authors.forename: Federico
authors.initials: F
authors.lastname: Perdomo-Celis
authors.affiliation: Programa de Medicina, Facultad de Salud, Universidad Surcolombiana, Neiva, Colombia.|Departamento de Pediatría, Hospital Universitario de Neiva, Neiva, Colombia.
authors.forename: Doris M
authors.initials: DM
authors.lastname: Salgado
authors.affiliation: Programa de Medicina, Facultad de Salud, Universidad Surcolombiana, Neiva, Colombia.
authors.forename: Diana M
authors.initials: DM
authors.lastname: Castañeda
authors.affiliation: Programa de Medicina, Facultad de Salud, Universidad Surcolombiana, Neiva, Colombia cfnarvaez@usco.edu.co.
authors.forename: Carlos F
authors.initials: CF
authors.lastname: Narváez
mesh_terms: D000293:Adolescent; D002470:Cell Survival; D002648:Child; D002675:Child, Preschool; D015925:Cryopreservation; D003715:Dengue / Q000276:immunology*; D005260:Female; D006801:Humans; D007223:Infant; D007371:Interferon-gamma / Q000096:biosynthesis; D007963:Leukocytes, Mononuclear / Q000276:immunology* / Q000378:metabolism* / Q000473:pathology; D008297:Male; D019595:Severe Dengue / Q000276:immunology; D013601:T-Lymphocytes / Q000276:immunology*; D014343:Trypan Blue / Q000378:metabolism
chemical_list: D007371:Interferon-gamma; D014343:Trypan Blue