Hemorrhagic shock is a leading cause of morbidity and mortality worldwide. Significant
blood loss may lead to decreased blood pressure and inadequate tissue perfusion with
resultant organ failure and death, even after replacement of lost blood volume. One reason
for this high acuity is that the fundamental mechanisms of shock are poorly understood.
Proteomic and metabolomic approaches have been used to investigate the molecular events
occurring in hemorrhagic shock but, to our knowledge, a systematic analysis of the
transcriptomic profile is missing. Therefore, a pilot analysis using paired-end RNA
sequencing was used to identify changes that occur in the blood transcriptome of rats
subjected to hemorrhagic shock after blood reinfusion. Hemorrhagic shock was induced using
a Wigger’s shock model. The transcriptome of whole blood from shocked animals shows
modulation of genes related to inflammation and immune response (Tlr13, Il1b, Ccl6,
Lgals3), antioxidant functions (Mt2A, Mt1), tissue injury and repair pathways (Gpnmb,
Trim72) and lipid mediators (Alox5ap, Ltb4r, Ptger2) compared with control animals. These
findings are congruent with results obtained in hemorrhagic shock analysis by other
authors using metabolomics and proteomics. The analysis of blood transcriptome may be a
valuable tool to understand the biological changes occurring in hemorrhagic shock and a
promising approach for the identification of novel biomarkers and therapeutic targets.
Impact statement
This study provides the first pilot analysis of the changes occurring in transcriptome
expression of whole blood in hemorrhagic shock (HS) rats. We showed that the analysis of
blood transcriptome is a useful approach to investigate pathways and functional
alterations in this disease condition. This pilot study encourages the possible
application of transcriptome analysis in the clinical setting, for the molecular
profiling of whole blood in HS patients.
KauvarDSWadeCE. The epidemiology and modern management of
traumatic hemorrhage: US and international perspectives. Crit
Care2005; 9(Suppl 5):
S1–S9.
2.
KochanekKDMurphySLXuJ. Deaths: final data for 2011.
Natl Vital Stat Rep2015; 63:
1–120.
3.
MongardonNDysonASingerM. Is MOF an outcome parameter or a transient,
adaptive state in critical illness?Curr Opin Crit Care2009; 15:
431–6.
4.
CardenDLGrangerDN. Pathophysiology of ischaemia-reperfusion
injury. J Pathol2000; 190:
255–66.
5.
KvietysPRGrangerDN. Endothelial cell monolayers as a tool for
studying microvascular pathophysiology. Am J Physiol1997; 273:
G1189–99.
6.
RahbarECardenasJCBaimukanovaGUsadiBBruhnRPatiSOstrowskiSRJohanssonPIHolcombJBWadeCE. Endothelial glycocalyx shedding and vascular
permeability in severely injured trauma patients. J Transl
Med2015; 13:
1–7.
7.
GrangerDNKvietysPR. Reperfusion injury and reactive oxygen species:
the evolution of a concept. Redox Biol2015; 6:
524–51.
8.
LiewCCMaJTangHCZhengRDempseyAA. The peripheral blood transcriptome dynamically
reflects system wide biology: a potential diagnostic tool. J
Lab Clin Med2006; 147:
126–32.
9.
AlettiFMaffioliENegriASantamariaMHDeLanoFAKistlerEBSchmid-SchönbeinGWTedeschiG. Peptidomic analysis of rat plasma: proteolysis in
hemorrhagic shock. Shock2015; 45:
540–54.
LiaoYSmythGKShiW. FeatureCounts: an efficient general purpose
program for assigning sequence reads to genomic features.
Bioinformatics2014; 30:
923–30.
12.
LoveMIHuberWAndersS. Moderated estimation of fold change and
dispersion for RNA-seq data with DESeq2. Genome Biol2014; 15:
550–550.
13.
RobinsonMDMcCarthyDJSmythGK. edgeR: a Bioconductor package for differential
expression analysis of digital gene expression data.
Bioinformatics2010; 26:
139–40.
14.
BindeaGMlecnikBHacklHCharoentongPTosoliniMKirilovskyAFridmanWHPagèsFTrajanoskiZGalonJ. ClueGO: a Cytoscape plug-in to decipher
functionally grouped gene ontology and pathway annotation networks.
Bioinformatics2009; 25:
1091–3.
15.
FülöpATurócziZGarbaiszDHarsányiLSzijártóA. Experimental models of hemorrhagic shock: a
review. Eur Surg Res2013; 50:
57–70.
MiraJCSzpilaBENacionalesDCLopezMGentileLFMathiasBJVanzantELUngaroRHoldenDRosenthalMDRinconJVerdugoPTLarsonSDMooreFABrakenridgeSCMohrAMBakerHVMoldawerLLEfronPA. Patterns of gene expression among murine models
of hemorrhagic shock/trauma and sepsis. Physiol
Genomics2016; 6:
135–44.
18.
LundJMAlexopoulouLSatoAKarowMAdamsNCGaleNWIwasakiAFlavellRA. Recognition of single-stranded RNA viruses by
Toll-like receptor 7. Proc Natl Acad Sci USA2004; 101:
5598–603.
19.
DivanovicSTrompetteAAtabaniSFMadanRGolenbockDTVisintinAFinbergRWTarakhovskyAVogelSNBelkaidYKurt-JonesEAKarpCL. Negative regulation of Toll-like receptor 4
signaling by the Toll-like receptor homolog RP105. Nat
Immunol2005; 6:
571–8.
20.
LafleurAMLukacsNWKunkelSLMatsukawaAArborAStudiesM. Role of CC chemokine CCL6/C10 as a monocyte
chemoattractant in a murine acute peritonitis. Mediators
Inflamm2004; 13:
349–55.
21.
HendersonNCSethiT. The regulation of inflammation by
galectin-3. Immunol Rev2009; 230:
160–71.
22.
FilipeMDMeijersWCRogier van der VeldeAde BoerRA. Galectin-3 and heart failure: prognosis,
prediction & clinical utility. Clin Chim Acta2015; 443:
48–56.
23.
JakobSMPickRBrechtefeldDNussbaumCKieferFSperandioMWalzogB. Hematopoietic progenitor kinase 1 (HPK1) is
required for LFA-1-mediated neutrophil recruitment during the acute inflammatory
response. Blood2013; 121:
4184–94.
24.
BradfieldPFNoursharghSAurrand-LionsMImhofBA. JAM family and related proteins in leukocyte
migration (Vestweber series). Arterioscler Thromb Vasc
Biol2007; 27:
2104–12.
25.
AlvarezJIKebirHCheslowLChabaratiMLarochelleCIvJPratA. JAML mediates monocyte and CD8 T cell migration
across the brain endothelium. Ann Clin Transl Neurol2015; 2:
1032–7.
26.
WitherdenDAVerdinoPRiederSEGarijoORobynETeytonLFischerWHWilsonIAHavranWL. The adhesion molecule JAML is a costimulatory
receptor for epithelial gammadelta T cell activation.
Science2010; 329:
1205–10.
27.
Eon KuekLLefflerMMackayGAHulettMD. The MS4A family: counting past 1, 2 and
3. Immunol Cell Biol2016; 94:
11–23.
28.
HotchkissRSMonneretGPayenD. Sepsis-induced immunosuppression: from cellular
dysfunctions to immunotherapy. Nat Rev Immunol2013; 13:
862–74.
29.
SerhanCN. Pro-resolving lipid mediators are leads for
resolution physiology. Nature2014; 510:
92–101.
30.
EunJCMooreEEMauchleyDCJohnsonCAMengXBanerjeeAWohlauerMVZariniSGijónMAMurphyRC. The 5-lipoxygenase pathway is required for acute
lung injury following hemorrhagic shock. Shock2012; 37:
599–604.
31.
D’AlessandroAMooreHBMooreEEWitherMNemkovTGonzalezESlaughterAFragosoMHansenKCSillimanCCBanerjeeA. Early hemorrhage triggers metabolic responses
that build up during prolonged shock. Am J Physiol – Regul
Integr Comp Physiol2015; 308:
1034–44.
32.
SzebeniJBaranyiLSavaySGötzeOAlvingCRBüngerRMonganPD. Complement activation during hemorrhagic shock
and resuscitation in swine. Shock2003; 20:
347–55.
D’AlessandroADzieciatkowskaMPeltzEDMooreEEJordanJRSillimanCCBanerjeeAHansenKC. Dynamic changes in rat mesenteric lymph proteins
following trauma using label-free mass spectrometry.
Shock2014; 42:
509–17.
35.
MittalAMiddleditchMRuggieroKLovedayBDelahuntBJülligMCooperGJSWindsorJAPhillipsARJ. Changes in the mesenteric lymph proteome induced
by hemorrhagic shock. Shock2010; 34:
140–9.
36.
DzieciatkowskaMAlessandroADMooreEEWohlauerMBanerjeeASillimanCCHansenKC. Lymph is not a plasma ultrafiltrate: a proteomic
analysis of injured patients. Shock2015; 42:
485–98.
37.
TarbellJMCancelLM. The glycocalyx and its significance in human
medicine. J Intern Med2016; 280:
97–113.
38.
WeinbaumSTarbellJMDamianoER. The structure and function of the endothelial
glycocalyx layer. Annu Rev Biomed Eng2007; 9:
121–67.
39.
ScheibnerKALutzMABoodooSFentonMJPowellJDHortonMR. Hyaluronan fragments act as an endogenous danger
signal by engaging TLR2. J Immunol2006; 177:
1272–81.
40.
De la MotteCNigroJVasanjiARhoHKesslerSBandyopadhyaySDaneseSFiocchiCSternR. Platelet-derived hyaluronidase 2 cleaves
hyaluronan into fragments that trigger monocyte-mediated production of proinflammatory
cytokines. Am J Pathol2009; 174:
2254–64.
41.
BenarafaC. The serpinB1 knockout mouse: a model for studying
neutrophil protease regulation in homeostasis and inflammation.
Methods Enzymol2011; 499:
135–48.
42.
BaumannMPhamCTNBenarafaC. SerpinB1 is critical for neutrophil survival
through cell-autonomous inhibition of cathepsin G.
Blood2013; 121:
3900–8.
SiegelDGustafsonDLDehnDLHanJYBoonchoongPBerlinerLJRossD. NAD(P)H:quinone oxidoreductase 1: role as a
superoxide scavenger. Mol Pharmacol2004; 65:
1238–47.
45.
ChiaveriniNDe LeyM. Protective effect of metallothionein on oxidative
stress-induced DNA damage. Free Radic Res2010; 44:
605–13.
46.
WakidaKShimazawaMHozumiISatohMNagaseHInuzukaTHaraH. Neuroprotective effect of erythropoietin, and
role of metallothionein-1 and -2, in permanent focal cerebral ischemia.
Neuroscience2007; 148:
105–14.
47.
QuallsJESubramanianCRafiWSmithAMBalouzianLDefreitasAAShireyKAReuttererBKernbauerEStockingerSDeckerTMiyairiIVogelSNSalgamePRockCOMurrayPJ. Sustained generation of nitric oxide and control
of mycobacterial infection requires argininosuccinate synthase 1.
Cell Host Microbe2012; 12:
313–23.
48.
HierholzerCHarbrechtBMenezesJMKaneJMacMickingJNathanCFPeitzmanABBilliarTRTweardyDJ. Essential role of induced nitric oxide in the
initiation of the inflammatory response after hemorrhagic shock.
J Exp Med1998; 187:
917–28.
49.
MarchantDJBoydJHLinDCGranvilleDJGarmaroudiFSMcManusBM. Inflammation in myocardial
diseases. Circ Res2012; 110:
126–44.
50.
LiBCastanoAPHudsonTENowlinBTLinS-LBonventreJV.SwansonKDDuffieldJS. The melanoma-associated transmembrane
glycoprotein Gpnmb controls trafficking of cellular debris for degradation and is
essential for tissue repair. FASEB J2010; 24:
4767–81.
51.
NakanoYSuzukiYTakagiTKitashojiAOnoYTsurumaKYoshimuraSShimazawaMIwamaTHaraH. Glycoprotein nonmetastatic melanoma protein B
(GPNMB) as a novel neuroprotective factor in cerebral ischemia-reperfusion
injury. Neuroscience2014; 277:
123–31.
WangXXieWZhangYLinPHanLHanPWangYChenZJiGZhengMWeislederNXiaoRPTakeshimaHMaJChengH. Cardioprotection of Ischemia/Reperfusion injury
by cholesterol-dependent MG53-mediated membrane repair. Circ
Res2010; 107:
76–83.
54.
FangZCuiX. Design and validation issues in RNA-seq
experiments. Brief Bioinform2011; 12:
280–7.
55.
SuZŁabajPPLiSSThierry-MiegJThierry-MiegDShiWWangCSchrothGPSetterquistRAThompsonJFJonesWDXiaoWXuWJensenRVKellyRXuJConesaAFurlanelloCGaoHHongHJafariNLetovskySLiaoYLuFOakeleyEJPengZPraulCASantoyo-LopezJSchererAShiTSmythGKStaedtlerFSykacekPTanXXThompsonEAVandesompeleJWangMDWangJWolfingerRDZavadilJAuerbachSSBaoWBinderHBlomquistTBrilliantMHBushelPRCaiWCatalanoJGChangCWChenTChenGChenRChiericiMChuTMClevertDADengYDertiADevanarayanVDongZDopazoJDuTFangHFangYFasoldMFernandezAFischerMFurió-TariPFuscoeJCCaimetFGajSGandaraJGaoHGeWGondoYGongBGongMGongZGreenBGuoCGuoLGuoLWHadfieldJHellemansJHochreiterSJiaMJianMJohnsonCDKaySKleinjansJLababidiSLevySLiQZLiLLiLLiPLiYLiHLiJLiSLinSMLópezFJLuXLuoHMaXMeehanJMegherbiDBMeiNMuBNingBPandeyAPérez-FloridoJPerkinsRGPetersRPhanJHPiroozniaMQianFQingTRainbowLRocca-SerraPSambourgLSansoneSASchwartzSShahRShenJSmithTMStegleOStralis-PaveseNStupkaESuzukiYSzkotnickiLTTinningMTuBvan DelftJVela-BozaAVenturiniEWalkerSJWanLWangWWangJWangJWiebenEDWilleyJCWuPYXuanJYangYYeZYinYYuYYuanYCZhangJZhangKKZhangWZhangWZhangYZhaoCZhengYZhouYZumboPTongWKreilDPMasonCEShiL. A comprehensive assessment of RNA-seq accuracy,
reproducibility and information content by the Sequencing Quality Control
Consortium. Nat Biotechnol2014; 32:
903–14.
Supplementary Material
Please find the following supplemental material available below.
For Open Access articles published under a Creative Commons License, all supplemental material carries the same license as the article it is associated with.
For non-Open Access articles published, all supplemental material carries a non-exclusive license, and permission requests for re-use of supplemental material or any part of supplemental material shall be sent directly to the copyright owner as specified in the copyright notice associated with the article.
