An optimized multi-parameter flow cytometry protocol for human T regulatory cell analysis on fresh and viably frozen cells, correlation with epigenetic analysis, and comparison of cord and adult blood

Nettenstrom, Lauren; Alderson, Kory L.; Raschke, Eva; Evans, Michael D.; Sondel, Paul M.; Olek, Sven; Seroogy, Christine M.

doi:10.1016/j.jim.2012.09.014

Cited by 33 publications

(47 citation statements)

References 24 publications

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“…However, in our hands, Treg retain their suppressive capacity and phenotype after cryopreservation (data not shown). Similar findings have been described by other authors [21–25]. We quantified peripheral Treg frequency in CBMC from the different groups of infants and PBMC from adults as an external control group.…”

Section: Resultssupporting

confidence: 64%

Effect of chorioamnionitis on regulatory T cells in moderate/late preterm neonates

et al. 2015

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Regulatory T-cells (Treg) have a protective role for the control of immune activation and tissue damage. The effects of chorioamnionitis (chorio) on Treg in moderate/late preterm newborns are not known. We hypothesized that infants exposed to chorio would have decreased Treg frequency and/or function. We isolated mononuclear cells from adult peripheral blood and cord blood from term and moderate/late preterm infants who were classified for severity of chorio exposure. Mononuclear cells were analyzed by flow cytometry for Treg frequency and phenotype. Treg suppression of activation of conventional T-cells (Tcon) was also quantified. Treg frequencies were similar in all groups of neonates, but lower than that found in adults. Newborn Treg had a naïve phenotype, with decreased levels of CD45RO, HLA-DR, CD39 and TIGIT compared to adult Treg and chorio did not affect the phenotype. Treg from preterm newborns exposed to severe chorio had higher expression of Ki67 compared to the other groups. Treg from preterm newborns were less suppressive than Treg from adults or term, and the level of suppression was reduced with severe chorio. Relative to term, Treg frequency and phenotype were not affected by prematurity and chorio but their functionality was decreased. Lower Treg activity may contribute to inflammation in newborns that is often associated with chorioamnionitis.

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Section: Resultssupporting

confidence: 64%

Effect of chorioamnionitis on regulatory T cells in moderate/late preterm neonates

et al. 2015

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“…In this study, CD4 + CD25 + CD127lo/-T cells were used to define Treg cells. Recent work from our group utilizing multi-parameter flow cytometry alongside the demethylation status of a region of the Foxp3 promoter (Treg specific demethylated region,TSDR) found the strongest positive relationship using CD4 + CD25 + CD127lo/-T cells as the Treg cell defining phenotype when compared to CD4 + CD25 + CD127lo/-Foxp3+ T cells [36]. This finding suggests Foxp3 protein levels may not be constantly detectable using flow cytometry in bona fide Treg cells.…”

Section: Discussionmentioning

confidence: 95%

Inhaled corticosteroid use is associated with increased circulating T regulatory cells in children with asthma

et al. 2013

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BackgroundT regulatory (Treg) cells are important in balancing immune responses and dysregulation of Treg cells has been implicated in the pathogenesis of multiple disease states including asthma. In this study, our primary aim was to determine Treg cell frequency in the peripheral blood of children with and without asthma. The secondary aim was to explore the association between Treg cell frequency with allergen sensitization, disease severity and medication use.MethodsPeripheral blood mononuclear cells from healthy control subjects (N = 93) and asthmatic children of varying disease severity (N = 66) were characterized by multi-parameter flow cytometry.ResultsOur findings demonstrate that children with asthma had a significantly increased frequency of Treg cells compared to children without asthma. Using a multivariate model, increased Treg cell frequency in children with asthma was most directly associated with inhaled corticosteroid use, and not asthma severity, allergic sensitization, or atopic status of the asthma.ConclusionWe conclude that low dose, local airway administration of corticosteroids is sufficient to impact the frequency of Treg cells in the peripheral blood. These data highlight the importance of considering medication exposure when studying Treg cells and suggest inhaled corticosteroid use in asthmatics may improve disease control through increased Treg cell frequency.

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“…In human neonates, the percentage of Tregs in peripheral blood increases significantly in the first 5 days of life, reaching adult levels at that time 35 . Recent studies suggest that there is a significantly greater number of Tregs in cord blood compared with adult Tregs 36 . Furthermore, cord blood Tregs are highly functional and can suppress T-cell proliferation and T H 1 IFN-γ production, similar to adult Treg function 37 …”

Section: Adaptive Cellular Immunity: T-cell Subsetsmentioning

confidence: 99%

What Causes Biliary Atresia? Unique Aspects of the Neonatal Immune System Provide Clues to Disease Pathogenesis

Mack

2015

Cellular and Molecular Gastroenterology and Hepatology

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Biliary atresia (BA) is the most frequent identifiable cause of neonatal cholestasis, and the majority of patients will need liver transplantation for survival. Despite surgical intervention with the Kasai portoenterostomy, significant fibrosis and cirrhosis develop early in life. An increased understanding of what causes this inflammatory fibrosing cholangiopathy will lead to therapies aimed at protecting the intrahepatic biliary system from immune-mediated damage. This review focuses on studies pertaining to the role of the adaptive immune response in bile duct injury in BA, including cellular and humoral immunity. The neonatal presentation of BA prompts the question of what potential modifications of unique aspects of the neonatal immune system set the stage for the progressive biliary disease. This review also discusses the characteristics of neonatal immune response and the theories on how alterations of this response could contribute to the pathogenesis of BA. These include aberrant type 1 helper T-cell (TH1) and TH17 responses, deficiencies in regulatory T cells, activation of humoral immunity, and autoimmunity. To advance our understanding of the etiology of BA, future studies should focus on the unique aspects of the neonatal immune system that have gone awry.

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An optimized multi-parameter flow cytometry protocol for human T regulatory cell analysis on fresh and viably frozen cells, correlation with epigenetic analysis, and comparison of cord and adult blood

Cited by 33 publications

References 24 publications

Effect of chorioamnionitis on regulatory T cells in moderate/late preterm neonates

Effect of chorioamnionitis on regulatory T cells in moderate/late preterm neonates

Inhaled corticosteroid use is associated with increased circulating T regulatory cells in children with asthma

What Causes Biliary Atresia? Unique Aspects of the Neonatal Immune System Provide Clues to Disease Pathogenesis

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