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Pre-term infant transfusion support: adult or cord blood donation? A longitudinal study of CD71 expression in pre-term, cord and adult blood samples

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Pre-term infant transfusion support: adult or cord blood donation? A longitudinal study of CD71 expression in pre-term, cord and adult blood samples Dr Megan Forrester Infection, Immunity & Inflammation
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Pre-term infant transfusion support: adult or cord blood donation? A longitudinal study of CD71 expression in pre-term, cord and adult blood samples Dr Megan Forrester Infection, Immunity & Inflammation University of Aberdeen Circulating CD71+ Glycophorin A+ cells CD71 or Transferrin Receptor Imports iron Expressed during erythropoiesis Not expressed on mature red blood cells Glycophorin A or GPA Immunosuppressive CD71+ erythroid cells compromise neonatal host defence against infection Elahi et al. Nature Dec 2013 Mouse Human Necrotising Enterocolitis Necrosis of the bowel caused by inflammation and ischaemic death of the gut Pathogenesis involves immature gut mucosal barrier, abnormalities of blood flow, abnormal bacterial colonisation and immune responses Risk factors include low birthweight, prematurity and transfusion Transfusion-associated NEC - Pathogenesis Adult Blood Transfusion Cord Blood Transfusion INFLAMMATION Inflammation Aims Identify in neonatal cord blood samples the early red cell population (CD71+ Glycophorin A+) Compare to adult blood donors Establish a time course of their development with respect to gestational age Assess their potential anti-inflammatory activity. Aims Identify in neonatal cord blood samples the early red cell population (CD71+ Glycophorin A+) Compare to adult blood donors Establish a time course of their development with respect to gestational age Assess their potential anti-inflammatory activity. CD71 Expression Analysis Pre-term Cord Lower density MNC Fraction Higher density RBC Fraction Adult Glycophorin A CD71 CD71 Expression Analysis Cord Adult Examples of flow cytometry plots of CD71 vs Glycophorin A from pre-term, cord and adult samples. Pre-term Cord Adult Glycophorin A Lower Density MNC Fraction Higher Density RBC Fraction CD71 Cord Blood Has A Higher Concentration of CD71+ Glycophorin A+ Cells Than Adult Blood Lower Density MNC Fraction Higher Density RBC Fraction % CD71+ Glycophorin A % CD71+ Glycophorin A Cord Adult Sample 0 Cord Adult Sample Pre-Term Blood Has A Higher Concentration of CD71+ Glycophorin A+ Cells Than Adult Blood Lower Density MNC Fraction Higher Density RBC Fraction % CD71+ Glycophorin A % CD71+ Glycophorin A Pre-Term Full Term Adult Sample 0 Pre-Term Full Term Adult Sample Average of each Pre-Term infant Pre-Term Blood Has A Higher Concentration of CD71+ Glycophorin A+ Cells Than Adult Blood Lower Density MNC Fraction Higher Density RBC Fraction % CD71+ Glycophorin A % CD71+ Glycophorin A Pre-Term Cord Adult Sample 0 Pre-Term Cord Adult Sample All Pre-Term samples CD71+ Glycophorin A+ cell concentration in the mononuclear cell fraction does not follow a pattern with equivalent gestational age 100 Lower Density MNC Fraction % CD71+ Glycophorin A wk wk wk wk Equivalent Gestational Age Equivalent gestational age = gestation age at birth + age 37+ wk Full Term Cord CD71+ Glycophorin A+ cell concentration in the RBC fraction decreases according to equivalent gestational age until similar to full term cord 10 Higher Density RBC Fraction % CD71+ Glycophorin A wk wk wk wk Equivalent Gestational Age 37+ wk Equivalent gestational age = gestation age at birth + age Full Term Cord Aims oidentify in neonatal cord blood samples the early red cell population (CD71+ Glycophorin A+) ocompare to adult blood donors oestablish a time course of their development with respect to gestational age oassess their potential anti-inflammatory activity. Aim:- Identify in neonatal cord blood samples the early red cell population (CD71+ Glycophorin A+) Identified CD71+ Glycophorin A+ cells in neonatal cord blood and pre-term samples Glycophorin A CD71 Aim:- Compare to adult blood donors CD71+ Glycophorin A+ cells in cord and pre-term samples have a higher concentration compared to adult blood donors Lower Density MNC Fraction Higher Density RBC Fraction % CD71+ Glycophorin A % CD71+ Glycophorin A Pre-Term Cord Adult Sample 0 Pre-Term Cord Adult Sample Aim:- Establish a time course of their development with respect to gestational age CD71+ Glycophorin A+ cells in the RBC fraction in pre-term samples decreases in concentration with increasing equivalent gestational age 10 Higher Density RBC Fraction % CD71+ Glycophorin A wk wk wk wk Equivalent Gestational Age 37+ wk Full Term Cord Aims Identified CD71+ Glycophorin A+ cells in neonatal cord blood and pre-term samples CD71+ Glycophorin A+ cells in cord and pre-term samples have a higher concentration compared to adult blood donors CD71+ Glycophorin A+ cells in the RBC fraction in pre-term samples decreases in concentration with increasing equivalent gestational age oassess their potential anti-inflammatory activity. Aims Identified CD71+ Glycophorin A+ cells in neonatal cord blood and pre-term samples CD71+ Glycophorin A+ cells in cord and pre-term samples have a higher concentration compared to adult blood donors CD71+ Glycophorin A+ cells in the RBC fraction in pre-term samples decreases in concentration with increasing equivalent gestational age oassess their potential anti-inflammatory activity. Adult RBC Fraction Cord RBC Fraction Cord Glycophorin A+ MNC Fraction Cord CBMC Adult PBMC Listeria LPS Monocytes TNF-α IL-6 IL-10? Examples of flow cytometry plots of inhibition experiments CD11c CD11b No Stimuli Listeria LPS TNF-α IL-6 No difference in CBMC and PBMC Response to Listeria or LPS % IL-6+ TNFa CBMC PBMC 20 0 Unstimulated Listeria LPS Stimuli Removing the glycophorin A+ cells from the MNC fraction reduces the pro-inflammatory response to LPS by CBMC % IL-6+ TNFa ** CBMC CBMC - GPA+ MNC CBMC (Pre-isolation) CBMC GPA+ MNC (Depleted) 0 Unstimulated Listeria LPS Stimuli Removing the glycophorin A+ cells then adding them back reduces the pro-inflammatory response to LPS by CBMC % IL-6+ TNFa+ 100 * CBMC - GPA+ MNC CBMC - GPA+ MNC + GPA+ MNC 80 PBMC C - GPA+ MNC PBMC C + - GPA+ MNC + GPA+ MNC Unstimulated Listeria LPS CBMC GPA+ MNC (Depleted) GPA+ MNC (Enriched) Stimuli Adding RBC did not inhibit pro-inflammatory cytokine production % IL-6+ TN Fa+ 100 CBMC CBMC + Cord RBC 80 C BM C + A dult RBC PBMC PBMC + Cord RBC 60 PBMC + Adult RBC U nstim ulated Listeria LPS Stim uli Cord RBC Adult RBC Adding RBC after removing GPA+ cells from the MNC fraction did inhibit pro-inflammatory cytokine production by CBMC % IL-6+ TNFa * * * ns ** * ns CBMC - GPA+ MNC CBMC - GPA+ MNC + Cord RBC CBMC - GPA+ MNC + Adult RBC PBMC - GPA+ MNC PBMC - + GPA+ Cord RBC MNC + Cord RBC PBMC - + GPA+ Adult RBC MNC + Adult RBC 20 0 Unstimulated Listeria LPS Stimuli Cord RBC Adult RBC Aims Identified CD71+ Glycophorin A+ cells in neonatal cord blood and pre-term samples CD71+ Glycophorin A+ cells in cord and pre-term samples have a higher concentration compared to adult blood donors CD71+ Glycophorin A+ cells in the RBC fraction in pre-term samples decreases in concentration with increasing equivalent gestational age oassess their potential anti-inflammatory activity. Aim:- Assess their potential anti-inflammatory activity CD71+ Glycophorin A+ cells from either MNC or RBC fraction do not conclusively demonstrate antiinflammatory properties % IL-6+ TNFa CBMC CBMC + Cord RBC CBMC + Adult RBC PBMC PBMC + Cord RBC PBMC + Adult RBC 20 0 Unstimulated Listeria LPS Stimuli Aims Identified CD71+ Glycophorin A+ cells in neonatal cord blood and pre-term samples CD71+ Glycophorin A+ cells in cord and pre-term samples have a higher concentration compared to adult blood donors CD71+ Glycophorin A+ cells in the RBC fraction in pre-term samples decreases in concentration with increasing equivalent gestational age CD71+ Glycophorin A+ cells from either MNC or RBC fractions do not conclusively demonstrate antiinflammatory properties Transfusion-associated NEC - Pathogenesis Adult Blood Transfusion Cord Blood Transfusion INFLAMMATION Inflammation Conclusions INFLAMMATION Inflammation Conclusions INFLAMMATION Inflammation Conclusions Adult Blood Transfusion Cord Blood Transfusion Acknowledgements Dr Sylvia Armstrong-Fisher Aberdeen SNBTS Dr Sumaiya Mohamed Cassim NHS Grampian Dr Caitlin Dimasco - NHS Grampian Professor Rob Barker -University of Aberdeen Professor Mark Vickers University of Aberdeen and Aberdeen SNBTS
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