Aleman, M. M.

Geddings, J.

Mackman, N.

Massberg, S.

von Brühl, M.-L.

Wolberg, A.

PubMed

Carolina Digital Repository

RECOMMENDATIONS AND GUIDELINESSStrengths and weaknesses of a new mouse model ofthrombosis induced by inferior vena cava stenosis:communication from the SSC of the ISTHJ . GEDDINGS ,* M. M. ALEMAN,* A . WOLBERG,* M. -L . VON BR €UHL ,† S . MASSBERG†and N . MACKMAN‡*Pathology and Laboratory Medicine, University of North Carolina, Chapel Hill, NC, USA; †Medizinische Klinik und Poliklinik I, LMUMunich, M€unchen, Germany; and ‡Department of Medicine, UNC McAllister Heart Institute, University of North Carolina, Chapel Hill, NC,USATo cite this article: Geddings J, Aleman MM, Wolberg A, von Br€uhl ML, Massberg S, Mackman N. Strengths and weaknesses of a new mousemodel of thrombosis induced by inferior vena cava stenosis: communication from the SSC of the ISTH. J Thromb Haemost 2014; 12: 571–3.Deep vein thrombosis (DVT) is a significant cause ofmorbidity and mortality throughout the world [1]. Riskfactors for DVT include major surgery, immobilization,trauma, and cancer, among others [1]. Many mouse mod-els of thrombosis have been used to study mechanisms ofthrombosis, including those involving small and largeveins. In this report from the Animal Models Subcommit-tee of the ISTH, we summarize the strengths and weak-nesses of the inferior vena cava (IVC) stenosis mousemodel of venous thrombosis.HistoryIn this model, the lumen volume of the IVC is reduced by~ 90% [2,3]. This model was adapted from a similar ratmodel called the ‘St Thomas Model’, involving IVC ste-nosis followed by endothelial damage with vascularclamps upstream of the stenosis site [4]. Many variants ofthe St Thomas model have been described. The IVC ste-nosis mouse model described in this article was firstreported in 2011, and does not involve a vascular damagestep [2]. Since then, several groups have used the model,with slight variations [3,5–9]. Thrombosis in this newmodel is thought to be initiated by the combination ofendothelial activation, a reduction in blood flow velocity,and disturbed blood flow upstream of the stenosis site[2,3].MethodsIn this model, a midline laparotomy is performed onanesthetized male mice that are at least 8 weeks of ageand 22 g in weight [2,3,6]. The IVC is exposed by atrau-matic blunt dissection, and the IVC is carefully separatedfrom the abdominal aorta. A spacer is placed on top ofthe exposed IVC, and a non-reactive permanent narrow-ing ligature (7.0 or 8.0 monofil polypropylene) is securedaround both the IVC and spacer directly below the renalveins. The spacer is then removed, resulting in a ~ 90%reduction in IVC lumen size at the stenosis site [2]. Injuryto the IVC wall is avoided during the procedure. Micewith any bleeding from the IVC are excluded. The surgi-cal site is then closed, and mice are allowed to recoverunder the influence of appropriate analgesia.Sources of variabilityThe spacers used for this model range from 0.26 to0.36 mm in diameter, and consist of guide wires, sutures,or blunted needles [2,3,5–9]. The most common spacersused to date are a 0.36-mm guide wire [3,9] and a blunted30-gauge needle [2,6,7]. The effect of spacer diameter andlength on thrombosis in this model has not been evalu-ated. The method of surgical anesthesia also varies, withan isoflurane/oxygen mixture being the most common[2,3,5–9]. There is some debate as to whether sidebranches should be ligated to control for IVC anatomyvariation. A concern with side branch ligation is that itmay result in endothelial damage and increase the dura-tion of surgery. One group compared the size of thrombiin mice with and without side branch ligation, and foundCorrespondence: Nigel Mackman, Pathology and Laboratory Medi-cine, UNC McAllister Heart Institute, University of North Carolina,Chapel Hill, NC, USA.Tel.: +1 919 843 3961; fax: +1 919 966 7639.E-mail: nmackman@med.unc.eduReceived 24 September 2013Manuscript handled by: S. EichingerFinal decision: P. H. Reitsma, 7 January 2013© 2014 The Authors. Journal of Thrombosis and Haemostasis published by Wiley Periodicals, Inc. on behalf of International Society on Thrombosis and HaemostasisThis is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License,which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial andno modifications or adaptations are made.Journal of Thrombosis and Haemostasis, 12: 571–573 DOI: 10.1111/jth.12510no difference between the two groups [9]. This samestudy, however, found that side branches that were< 1.5 mm from the stenosis site resulted in smallerthrombi [9].Quantification of thrombus sizeThrombus size in this model is typically quantified bymeasuring thrombus mass or size at a given time pointafter the induction of stenosis. In order to obtain thesemeasurements, mice are euthanized and the IVC is har-vested. We and others have extended the utility of thismodel by using high-frequency ultrasonography to imageblood flow and quantify IVC thrombus growth in vivo(Fig. 1A–C) [9].Strengths and weaknesses of the new modelIt has been well documented that human DVT is mostoften initiated within the valve sinuses of large veins onan intact endothelium [1]. A strength of this model is thatthrombosis is initiated in a large vein in the mouse in theabsence of major vessel damage. Furthermore, althoughthere are no valves in the IVC, the disturbed blood flowoccurring around the stenosis site may mimic changes inblood flow that occur over valves and within valve pock-ets in large veins in humans (Fig. 1D). The thrombi thatdevelop in this model are also histologically similar tohuman venous thrombi [2,3].The major concern with this model is that throm-bus development is variable. The reported incidence ofStenosissiteStenosissiteStenosis siteDirection of blood flowThrombusDirection of blood flowDirection of blood flowArea 14:A = 13.729 mm200 10 20 30Thrombus mass (mg)40 505Longitudinal area (mm2 )101520r2 = 0.96A BC DFig. 1. In vivo imaging of the inferior vena cava (IVC) stenosis model. (A, B) IVC stenosis was performed in C57Bl/6J mice, and ultrasono-graphic imaging of the stenosis site (A) before and (B) after thrombus formation was performed. (C) Longitudinal thrombus area was mea-sured in mice at 24 h, by ultrasonography. Thrombi were then immediately harvested and weighed. Linear regression analysis for longitudinalthrombus area vs. thrombus mass identified a strong correlation (r2 = 0.96). (D) Color Doppler imaging reveals a pattern of disturbed bloodflow around the stenosis site prior to thrombus development.© 2014 The Authors. Journal of Thrombosis and Haemostasis published by Wiley Periodicals, Inc. on behalf of International Society on Thrombosis and Haemostasis572 J. Geddings et althrombosis at 48 h in wild-type mice ranges from 44% to100% [2,5,8,9]. We have a lot of experience with thismodel in our laboratory, and followed the developmentand growth of thrombi in 15 C57Bl/6J mice aged 14–18 weeks over a period of 48 h. High-frequency ultraso-nography was used to measure thrombus size in eachmouse at 3, 6, 9 and 48 h after the initiation of thrombo-sis (Table S1). As expected, we found that both thrombussize at each time point and time to thrombus initiationwere variable, with thombi appearing in the IVC any-where from 3 to 48 h after stenosis (thrombus area, inci-dence: 3 h, 0–8.5 mm2, 13%; 6 h, 0–11.1 mm2, 33%; 9 h,0–10.1 mm2, 40%; 48 h, 0–11.1 mm2, 53%). Another lim-itation of this model is that it is performed in a vein thatdoes not have valves.ConclusionsLike any model, this mouse model of venous thrombosisdoes not accurately reflect all aspects of human DVT.However, as discussed above, it does have certainstrengths over other available models. Primarily, the initi-ation of thrombosis occurs without the induction ofmajor vascular damage, which more closely mimicshuman DVT than models triggered by chemical or physi-cal vessel damage. Nevertheless, the high variability inthrombus development is a significant concern that limitsits utility for detecting small differences between groups.AddendumJ. Geddings and N. Mackman wrote the manuscript, withinput from M. Aleman, A. Wolberg, M. von Br€uhl, andS. Massberg.AcknowledgementsTechnical support was provided by M. Rojas in the UNCMcAllister Heart Institute surgical models core.Disclosure of Conflict of InterestsThe authors state that they have no conflict of interest.Supporting InformationAdditional Supporting Information may be found in theonline version of this article:Table S1. Size and incidence of thrombosis in the IVCstenosis model.References1 Mackman N. New insights into the mechanisms of venous throm-bosis. J Clin Invest 2012; 122: 2331–6.2 Brill A, Fuchs TA, Chauhan AK, Yang JJ, De Meyer SF, Kol-lnberger M, Wakefield TW, Lammle B, Massberg S, WagnerDD. von Willebrand factor-mediated platelet adhesion is criticalfor deep vein thrombosis in mouse models. Blood 2011; 117:1400–7.3 von Bruhl ML, Stark K, Steinhart A, Chandraratne S, Konrad I,Lorenz M, Khandoga A, Tirniceriu A, Coletti R, Kollnberger M,Byrne RA, Laitinen I, Walch A, Brill A, Pfeiler S, Manukyan D,Braun S, Lange P, Riegger J, Ware J, et al. Monocytes, neutroph-ils, and platelets cooperate to initiate and propagate venousthrombosis in mice in vivo. J Exp Med 2012; 209: 819–35.4 McGuinness C, Humphries J, Smith A, Burnand K. A new modelof venous thrombosis. Cardiovasc Surg 1997; S5: 123.5 Brill A, Fuchs TA, Savchenko AS, Thomas GM, Martinod K, DeMeyer SF, Bhandari AA, Wagner DD. Neutrophil extracellulartraps promote deep vein thrombosis in mice. J Thromb Haemost2012; 10: 136–44.6 Wang JG, Geddings JE, Aleman MM, Cardenas JC, Chant-rathammachart P, Williams JC, Kirchhofer D, Bogdanov VY,Bach RR, Rak J, Church FC, Wolberg AS, Pawlinski R, Key NS,Yeh JJ, Mackman N. Tumor-derived tissue factor activates coagu-lation and enhances thrombosis in a mouse xenograft model ofhuman pancreatic cancer. Blood 2012; 119: 5543–52.7 Brill A, Yesilaltay A, De Meyer SF, Kisucka J, Fuchs TA,Kocher O, Krieger M, Wagner DD. Extrahepatic high-densitylipoprotein receptor SR-BI and apoA-I protect against deep veinthrombosis in mice. Arterioscler Thromb Vasc Biol 2012; 32:1841–7.8 Martinod K, Demers M, Fuchs TA, Wong SL, Brill A, GallantM, Hu J, Wang Y, Wagner DD. Neutrophil histone modificationby peptidylarginine deiminase 4 is critical for deep vein thrombo-sis in mice. Proc Natl Acad Sci USA 2013; 110: 8674–9.9 Brandt M, Schonfelder T, Schwenk M, Becker C, Jackel S, Rein-hardt C, Stark K, Massberg S, Munzel T, von Bruhl ML, WenzelP. Deep vein thrombus formation induced by flow reduction inmice is determined by venous side branches. Clin HemorheolMicrocirc 2013. [Epub ahead of print].© 2014 The Authors. Journal of Thrombosis and Haemostasis published by Wiley Periodicals, Inc. on behalf of International Society on Thrombosis and HaemostasisMouse model of venous thrombosis 573

Strengths and weaknesses of a new mouse model of thrombosis induced by inferior vena cava stenosis: communication from the SSC of the ISTH

https://cdr.lib.unc.edu/downloads/6t053p59w

Strengths and weaknesses of a new mouse model of thrombosis induced by inferior vena cava stenosis: communication from the SSC of the ISTH

Abstract

Similar works

Full text

Available Versions

Carolina Digital Repository