Serial blood cytokine and chemokine mRNA and microRNA over 48 h are insult specific in a piglet model of inflammation-sensitized hypoxia-ischaemia

Ingran Lingam; Adnan Avdic-Belltheus; Christopher Meehan; Kathryn Martinello; Sara Ragab; Donald Peebles; Melinda Barkhuizen; Cally J Tann; Ilias Tachtsidis; Tim G A M Wolfs; Henrik Hagberg; Nigel Klein; Bobbi Fleiss; Pierre Gressens; Xavier Golay; Boris W Kramer; Nicola J Robertson

doi:10.1038/s41390-020-0986-3

Serial blood cytokine and chemokine mRNA and microRNA over 48 h are insult specific in a piglet model of inflammation-sensitized hypoxia-ischaemia

Ingran Lingam, Adnan Avdic-Belltheus, Christopher Meehan, Kathryn Martinello, Sara Ragab, Donald Peebles, Melinda Barkhuizen, Cally J Tann, Ilias Tachtsidis, Tim G A M Wolfs, Henrik Hagberg, Nigel Klein, Bobbi Fleiss, Pierre Gressens, Xavier Golay, Boris W Kramer, Nicola J Robertson

Neonatology, Institute for Women's Health, University College London, London, UK.
University of Adelaide
Maternal and Fetal Medicine Department, Institute for Women's Health, University College London, London, United Kingdom.
Department of Pediatrics, University of Maastricht, Maastricht, The Netherlands.
Maternal Adolescent, Reproductive and Child Health (MARCH) Centre, Department of Infectious Disease Epidemiology, London School of Hygiene and Tropical Medicine, London, UK.
9Department of Medical Physics and Biomedical Engineering, University College London.
Centre of Perinatal Medicine and Health, Institute of Clinical Sciences, Department of Obstetrics and Gynecology & Institute of Neuroscience and Physiology, Sahlgrenska Academy, University of Gothenburg, Sweden.
Paediatric Infectious Diseases & Immunology, Institute of Child Health, University College London, London, UK.
Centre for the Developing Brain, Kings College, St Thomas Campus, London, United Kingdom.
University of Paris VII - Denis Diderot University
Leonard Wolfson Experimental Neurology Centre and Academic Neuroradiological Unit, Department of Brain Repair and Rehabilitation, University College London Queen Square Institute of Neurology, University College London, London, UK.
Neonatology, Institute for Women's Health, University College London, London, UK. [email protected].

Research output: Contribution to journal › Article › peer-review

7 Citations (Scopus)

Abstract

Background: Exposure to inflammation exacerbates injury in neonatal encephalopathy (NE). We hypothesized that brain biomarker mRNA, cytokine mRNA and microRNA differentiate inflammation (E. coli LPS), hypoxia (Hypoxia), and inflammation-sensitized hypoxia (LPS + Hypoxia) in an NE piglet model. Methods: Sixteen piglets were randomized: (i) LPS 2 μg/kg bolus; 1 μg/kg infusion (LPS; n = 5), (ii) Saline with hypoxia (Hypoxia; n = 6), (iii) LPS commencing 4 h pre-hypoxia (LPS + Hypoxia; n = 5). Total RNA was acquired at baseline, 4 h after LPS and 1, 3, 6, 12, 24, 48 h post-insult (animals euthanized at 48 h). Quantitative PCR was performed for cytokines (IL1A, IL6, CXCL8, IL10, TNFA) and brain biomarkers (ENO2, UCHL1, S100B, GFAP, CRP, BDNF, MAPT). MicroRNA was detected using GeneChip (Affymetrix) microarrays. Fold changes from baseline were compared between groups and correlated with cell death (TUNEL) at 48 h. Results: Within 6 h post-insult, we observed increased IL1A, CXCL8, CCL2 and ENO2 mRNA in LPS + Hypoxia and LPS compared to Hypoxia. IL10 mRNA differentiated all groups. Four microRNAs differentiated LPS + Hypoxia and Hypoxia: hsa-miR-23a, 27a, 31-5p, 193-5p. Cell death correlated with TNFA (R = 0.69; p < 0.01) at 1–3 h and ENO2 (R = −0.69; p = 0.01) at 48 h. Conclusions: mRNA and miRNA differentiated hypoxia from inflammation-sensitized hypoxia within 6 h in a piglet model. This information may inform human studies to enable triage for tailored neuroprotection in NE. Impact: Early stratification of infants with neonatal encephalopathy is key to provide tailored neuroprotection.IL1A, CXCL8, IL10, CCL2 and NSE mRNA are promising biomarkers of inflammation-sensitized hypoxia.IL10 mRNA levels differentiated all three pathological states; fold changes from baseline was the highest in LPS + Hypoxia animals, followed by LPS and Hypoxia at 6 h.miR-23, -27, -31-5p and -193-5p were significantly upregulated within 6 h of a hypoxia insult.Functional analysis highlighted the diverse roles of miRNA in the cellular processes.

Original language	English
Journal	Pediatric Research
DOIs	https://doi.org/10.1038/s41390-020-0986-3
Publication status	E-pub ahead of print - 10 Jun 2020

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10.1038/s41390-020-0986-3

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Lingam, I., Avdic-Belltheus, A., Meehan, C., Martinello, K., Ragab, S., Peebles, D., Barkhuizen, M., Tann, C. J., Tachtsidis, I., Wolfs, T. G. A. M., Hagberg, H., Klein, N., Fleiss, B., Gressens, P., Golay, X., Kramer, B. W., & Robertson, N. J. (2020). Serial blood cytokine and chemokine mRNA and microRNA over 48 h are insult specific in a piglet model of inflammation-sensitized hypoxia-ischaemia. Pediatric Research. Advance online publication. https://doi.org/10.1038/s41390-020-0986-3

@article{ff0ae1c84f214724ba52d33d989af52f,

title = "Serial blood cytokine and chemokine mRNA and microRNA over 48 h are insult specific in a piglet model of inflammation-sensitized hypoxia-ischaemia",

abstract = "Background: Exposure to inflammation exacerbates injury in neonatal encephalopathy (NE). We hypothesized that brain biomarker mRNA, cytokine mRNA and microRNA differentiate inflammation (E. coli LPS), hypoxia (Hypoxia), and inflammation-sensitized hypoxia (LPS + Hypoxia) in an NE piglet model. Methods: Sixteen piglets were randomized: (i) LPS 2 μg/kg bolus; 1 μg/kg infusion (LPS; n = 5), (ii) Saline with hypoxia (Hypoxia; n = 6), (iii) LPS commencing 4 h pre-hypoxia (LPS + Hypoxia; n = 5). Total RNA was acquired at baseline, 4 h after LPS and 1, 3, 6, 12, 24, 48 h post-insult (animals euthanized at 48 h). Quantitative PCR was performed for cytokines (IL1A, IL6, CXCL8, IL10, TNFA) and brain biomarkers (ENO2, UCHL1, S100B, GFAP, CRP, BDNF, MAPT). MicroRNA was detected using GeneChip (Affymetrix) microarrays. Fold changes from baseline were compared between groups and correlated with cell death (TUNEL) at 48 h. Results: Within 6 h post-insult, we observed increased IL1A, CXCL8, CCL2 and ENO2 mRNA in LPS + Hypoxia and LPS compared to Hypoxia. IL10 mRNA differentiated all groups. Four microRNAs differentiated LPS + Hypoxia and Hypoxia: hsa-miR-23a, 27a, 31-5p, 193-5p. Cell death correlated with TNFA (R = 0.69; p < 0.01) at 1–3 h and ENO2 (R = −0.69; p = 0.01) at 48 h. Conclusions: mRNA and miRNA differentiated hypoxia from inflammation-sensitized hypoxia within 6 h in a piglet model. This information may inform human studies to enable triage for tailored neuroprotection in NE. Impact: Early stratification of infants with neonatal encephalopathy is key to provide tailored neuroprotection.IL1A, CXCL8, IL10, CCL2 and NSE mRNA are promising biomarkers of inflammation-sensitized hypoxia.IL10 mRNA levels differentiated all three pathological states; fold changes from baseline was the highest in LPS + Hypoxia animals, followed by LPS and Hypoxia at 6 h.miR-23, -27, -31-5p and -193-5p were significantly upregulated within 6 h of a hypoxia insult.Functional analysis highlighted the diverse roles of miRNA in the cellular processes.",

author = "Ingran Lingam and Adnan Avdic-Belltheus and Christopher Meehan and Kathryn Martinello and Sara Ragab and Donald Peebles and Melinda Barkhuizen and Tann, {Cally J} and Ilias Tachtsidis and Wolfs, {Tim G A M} and Henrik Hagberg and Nigel Klein and Bobbi Fleiss and Pierre Gressens and Xavier Golay and Kramer, {Boris W} and Robertson, {Nicola J}",

year = "2020",

month = jun,

day = "10",

doi = "10.1038/s41390-020-0986-3",

language = "English",

journal = "Pediatric Research",

issn = "0031-3998",

publisher = "Lippincott Williams and Wilkins",

}

Lingam, I, Avdic-Belltheus, A, Meehan, C, Martinello, K, Ragab, S, Peebles, D, Barkhuizen, M, Tann, CJ, Tachtsidis, I, Wolfs, TGAM, Hagberg, H, Klein, N, Fleiss, B , Gressens, P, Golay, X, Kramer, BW & Robertson, NJ 2020, 'Serial blood cytokine and chemokine mRNA and microRNA over 48 h are insult specific in a piglet model of inflammation-sensitized hypoxia-ischaemia', Pediatric Research. https://doi.org/10.1038/s41390-020-0986-3

TY - JOUR

T1 - Serial blood cytokine and chemokine mRNA and microRNA over 48 h are insult specific in a piglet model of inflammation-sensitized hypoxia-ischaemia

AU - Lingam, Ingran

AU - Avdic-Belltheus, Adnan

AU - Meehan, Christopher

AU - Martinello, Kathryn

AU - Ragab, Sara

AU - Peebles, Donald

AU - Barkhuizen, Melinda

AU - Tann, Cally J

AU - Tachtsidis, Ilias

AU - Wolfs, Tim G A M

AU - Hagberg, Henrik

AU - Klein, Nigel

AU - Fleiss, Bobbi

AU - Gressens, Pierre

AU - Golay, Xavier

AU - Kramer, Boris W

AU - Robertson, Nicola J

PY - 2020/6/10

Y1 - 2020/6/10

N2 - Background: Exposure to inflammation exacerbates injury in neonatal encephalopathy (NE). We hypothesized that brain biomarker mRNA, cytokine mRNA and microRNA differentiate inflammation (E. coli LPS), hypoxia (Hypoxia), and inflammation-sensitized hypoxia (LPS + Hypoxia) in an NE piglet model. Methods: Sixteen piglets were randomized: (i) LPS 2 μg/kg bolus; 1 μg/kg infusion (LPS; n = 5), (ii) Saline with hypoxia (Hypoxia; n = 6), (iii) LPS commencing 4 h pre-hypoxia (LPS + Hypoxia; n = 5). Total RNA was acquired at baseline, 4 h after LPS and 1, 3, 6, 12, 24, 48 h post-insult (animals euthanized at 48 h). Quantitative PCR was performed for cytokines (IL1A, IL6, CXCL8, IL10, TNFA) and brain biomarkers (ENO2, UCHL1, S100B, GFAP, CRP, BDNF, MAPT). MicroRNA was detected using GeneChip (Affymetrix) microarrays. Fold changes from baseline were compared between groups and correlated with cell death (TUNEL) at 48 h. Results: Within 6 h post-insult, we observed increased IL1A, CXCL8, CCL2 and ENO2 mRNA in LPS + Hypoxia and LPS compared to Hypoxia. IL10 mRNA differentiated all groups. Four microRNAs differentiated LPS + Hypoxia and Hypoxia: hsa-miR-23a, 27a, 31-5p, 193-5p. Cell death correlated with TNFA (R = 0.69; p < 0.01) at 1–3 h and ENO2 (R = −0.69; p = 0.01) at 48 h. Conclusions: mRNA and miRNA differentiated hypoxia from inflammation-sensitized hypoxia within 6 h in a piglet model. This information may inform human studies to enable triage for tailored neuroprotection in NE. Impact: Early stratification of infants with neonatal encephalopathy is key to provide tailored neuroprotection.IL1A, CXCL8, IL10, CCL2 and NSE mRNA are promising biomarkers of inflammation-sensitized hypoxia.IL10 mRNA levels differentiated all three pathological states; fold changes from baseline was the highest in LPS + Hypoxia animals, followed by LPS and Hypoxia at 6 h.miR-23, -27, -31-5p and -193-5p were significantly upregulated within 6 h of a hypoxia insult.Functional analysis highlighted the diverse roles of miRNA in the cellular processes.

AB - Background: Exposure to inflammation exacerbates injury in neonatal encephalopathy (NE). We hypothesized that brain biomarker mRNA, cytokine mRNA and microRNA differentiate inflammation (E. coli LPS), hypoxia (Hypoxia), and inflammation-sensitized hypoxia (LPS + Hypoxia) in an NE piglet model. Methods: Sixteen piglets were randomized: (i) LPS 2 μg/kg bolus; 1 μg/kg infusion (LPS; n = 5), (ii) Saline with hypoxia (Hypoxia; n = 6), (iii) LPS commencing 4 h pre-hypoxia (LPS + Hypoxia; n = 5). Total RNA was acquired at baseline, 4 h after LPS and 1, 3, 6, 12, 24, 48 h post-insult (animals euthanized at 48 h). Quantitative PCR was performed for cytokines (IL1A, IL6, CXCL8, IL10, TNFA) and brain biomarkers (ENO2, UCHL1, S100B, GFAP, CRP, BDNF, MAPT). MicroRNA was detected using GeneChip (Affymetrix) microarrays. Fold changes from baseline were compared between groups and correlated with cell death (TUNEL) at 48 h. Results: Within 6 h post-insult, we observed increased IL1A, CXCL8, CCL2 and ENO2 mRNA in LPS + Hypoxia and LPS compared to Hypoxia. IL10 mRNA differentiated all groups. Four microRNAs differentiated LPS + Hypoxia and Hypoxia: hsa-miR-23a, 27a, 31-5p, 193-5p. Cell death correlated with TNFA (R = 0.69; p < 0.01) at 1–3 h and ENO2 (R = −0.69; p = 0.01) at 48 h. Conclusions: mRNA and miRNA differentiated hypoxia from inflammation-sensitized hypoxia within 6 h in a piglet model. This information may inform human studies to enable triage for tailored neuroprotection in NE. Impact: Early stratification of infants with neonatal encephalopathy is key to provide tailored neuroprotection.IL1A, CXCL8, IL10, CCL2 and NSE mRNA are promising biomarkers of inflammation-sensitized hypoxia.IL10 mRNA levels differentiated all three pathological states; fold changes from baseline was the highest in LPS + Hypoxia animals, followed by LPS and Hypoxia at 6 h.miR-23, -27, -31-5p and -193-5p were significantly upregulated within 6 h of a hypoxia insult.Functional analysis highlighted the diverse roles of miRNA in the cellular processes.

UR - http://www.scopus.com/inward/record.url?scp=85086241759&partnerID=8YFLogxK

U2 - 10.1038/s41390-020-0986-3

DO - 10.1038/s41390-020-0986-3

M3 - Article

C2 - 32521540

SN - 0031-3998

JO - Pediatric Research

JF - Pediatric Research

ER -

Serial blood cytokine and chemokine mRNA and microRNA over 48 h are insult specific in a piglet model of inflammation-sensitized hypoxia-ischaemia

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