A luminance-based heart chip assay for assessing the efficacy of graft preservation solutions in heart transplantation in rats

Abstract

ObjectiveWe developed a novel luciferase-based viability assay for assessing the viability of hearts preserved in different solutions. We examined whether this in vitro system could predict heart damage and survival after transplantation in rats.DesignBy our novel system, preserved heart viability evaluation and transplanted heart-graft functional research study.SettingUniversity basic science laboratory.InterventionsIsolated Luciferase-transgenic Lewis (LEW) rat cardiac-tissue-chips were plated on 96-well tissue-culture plates and incubated in preservation solutions at 4°C. Viability was measured as photon intensity by using a bio-imaging system. Heart-grafts preserved in University of Wisconsin (UW), extracellular-trehalose-Kyoto (ETK), Euro-Collins (EC), histidin-tryptophan-ketoglutarat solution (HTK), lactated Ringer's (LR) or normal saline solution were transplanted cervically by using a cuff-technique or into the abdomens of syngeneic wild-type LEW rats by using conventional microsurgical suture techniques.Main outcome measuresImaging an evaluation of preservation heart-graft and functional analysis.ResultsCardiac-tissue-chips preserved with UW, HTK or ETK solution gave higher luminance than those preserved with EC, LR or normal saline (p<0.03). After 24 h of preservation of hearts in each solution at 4°C, the beating of the isolated hearts was evaluated. The success rate, evaluation of beating, of cervical heart transplants using UW and ETK solution exceeded 70%, but those using other preservation solutions were lower (UW: 100%, ETK: 75%, EC: 42.86%, HTK: 14.29%, normal saline: 0%). Histological analysis of cervical heart-grafts after 3 h preservation by myeloperoxidase (MPO), zona occludens-1(ZO-1), and caspase-3 immunostaining revealed different degrees of preservation damage in all grafts.ConclusionsOur novel assay system is simple and can test multiple solutions. It should therefore be a powerful tool for developing and improving new heart-graft preservation solutions.

Ebselen Preserves Tissue-Engineered Cell Sheets and their Stem Cells in Hypothermic Conditions

“…Therefore, this system shows good performance for screening the effect of different factors in a preservation medium by measuring their results on the luciferase activity as an index of the remaining amount of ATP. To date, this method has been used to screen several types of preservation media in various organs such as the heart16, liver17, kidney18, islets1920, and small intestine21. However, to our knowledge, a preservation medium that is ideal for tissue-engineered cell sheets has not yet been screened.…”

Hepatocyte Transplantation Attenuates the Course of Acute Liver Failure Induced by Thioacetamide in Lewis Rats

“…The reliability of BLI analyses of transplanted cells isolated from luc-Lew-Tg was described and validated in previous studies (Hakamata et al 2006, Maeda et al 2013. Therefore in our experiments the same technique was used.…”

“…A recent development of transgenic rats expressing ubiquitously the gene for luciferase derived from the firefly (Photinus pyralis) is a major technological achievement in this field because it allows bioluminescence imaging (BLI) of the transplanted cells in vivo (Hakamata et al 2006). Using this technique, the same recipient can be repeatedly examined throughout the study without the need for animals to be killed at each time point (Hakamata et al 2006, Maeda et al 2013). Moreover, it was shown that an immunological reaction or immunosuppressive therapy substantially altered the function of the transplanted cells (Kawahara et al 2010, Loukopoulos et al 2014, Wu et al 2010.…”

“…Moreover, it was shown that an immunological reaction or immunosuppressive therapy substantially altered the function of the transplanted cells (Kawahara et al 2010, Loukopoulos et al 2014, Wu et al 2010. Since the transgenic "firefly luciferase rat" was developed on the background of an inbred rat strain (Lewis rats), the studies of transplanted hepatocytes can be performed without a need for post-Tx immunosuppression (Hakamata et al 2006, Maeda et al 2013. It is also noteworthy that in our recent study (Koblihová et al 2014) we developed and characterized a highly reproducible model of thioacetamide (TAA)-induced ALF in Lewis rats which, in our opinion, is optimal for evaluation of new approaches to the therapy of ALF.…”

Activation of whole body by high levels of polyamine intake in rats

“…We previously described the use of a luciferase transgenic (Luc-Tg) rat system, together with optical imaging techniques, to better understand cell and/or tissue conditions (Hakamata et al 2006;Teratani and Kobayashi 2012). We previously described the use of a luciferase-based cell-viability assay that detects intracellular ATP levels in viable cells, which we used to assess the viability of Luc-Tg rat-derived islets and kidney, heart, small intestine, and liver samples (Doi et al 2014;Iwai et al 2012Iwai et al , 2014Kasahara et al 2013;Maeda et al 2013;Negishi et al 2011;Teratani and Kobayashi 2012).…”