Cardioprotective mitochondrial factors in ischemia / reperfusion (IR) lesions

Delphine Prunier, University Professor - Hospital Practitioner

(delphine.prunier @ univ-angers.fr)

delphine.prunier @ univ-angers.fr (delphine.prunier @ univ-angers.fr)

Keywords : Cardioprotection, mitochondria, IR

State of the art and objectives

The Mitolab team joins the Biochemistry and Molecular Biology team, as well as Genetics and various clinical specialties.

In this perspective I have developed a research work with Professor Fabrice Prunier of the Cardiovascular Physiopathology team of our UMR on cardioprotective mitochondrial factors in ischemia / reperfusion (IR) lesions. From a pathophysiological point of view, IR lesions are the consequence of mitochondrial dysfunction represented by respiratory chain dysfunction, calcium overload, overproduction of ROS and the opening of the mitochondrial permeability transition pore (mPTP), associated with fragmentation of mitochondria. The cardioprotection methods developed are pharmacological with the use of calcium regulators, antioxidants, modulator of the opening of the mitochondrial transition pore (mPTP) ... or mechanical by the ischemic conditioning which consist in realizing short episodes of IR at level of the heart or other organ without causing irreversible injury. In our project we studied the influence of mitochondrial dynamics on cardioprotection and looked for metabolites involved in myocardial conditioning by metabolomic approaches.

Main results from the last 5 years

• Cardioprotection and metabolism of Tryptophan. We have shown through targeted and non-targeted metabolomic studies that kynurenine is a factor involved in cardioprotection during remote ischemic conditioning (RIC) (ref 8 and 10 in Appendix 5 Publications). These results were found in the conditioned rat in the paw and in the man conditioned by the swelling of a cuff in the arm. Reinjection of this metabolite into the rat confirmed its cardioprotective effect.

• Cardioprotection and mitochondrial dynamics. Discordant results on the mitochondrial fusion-fission relationship and cardioprotection led us to study murine models deficient in OPA1 fusion protein (Dr. Guy Lenaers) or fission protein DRP1 (Pr. Hiromi SESAKI, USA ) in the context of myocardial IR (ref 9 and 13 in Appendix 5, Publications).

These studies supported the idea that mice deficient in the fusion proteins (Opa1 +/- model) have increased sensitivity to IR lesions, in contrast to fission-deficient mice that are protected (Drp1 +/- model). We have reproduced these results pharmacologically by showing a decrease in the size of the necrotic area after IR in mice treated with a mitochondrial fission inhibitor (mDIVI). The mechanisms involved seem to be a modification of the calcium flux in the Opa1 +/- model and a modification of the mitopathy in our model Drp1 +/-. These abnormalities might be compensated since the double heterozygote mice Opa1 +/- and Drp1 +/- have a sensitivity to ischemia reperfusion lesions identical to the control mice. Thus, modulation of mitochondrial dynamics represents a possibility to reduce IR lesions. However, it remains to determine the modalities of this protection.

Hospital activities

• Oncogenetic endocrine

The various "cancer" plans put in place since 2004 have made possible to structure and label 11 laboratories in France for the study of gene variants involved in rare endocrine neoplasia. This network of laboratories named TENGEN set up various collaborative works under the aegis of the GTE (Working Group of Endocrine Tumors).

In this context, I coordinated a national work on RET gene variants identified in France. This study collected data from 5109 patients over 10 years (ref 1 in Appendix 5, Publications). It allowed to identify 74 variants of which 26 were not classified (VSI, variant of unknown meaning). We set up the study of the activation of intracellular signaling pathways on a NIH3T3 cell model (mouse fibroblasts) stably transfected with the wild or mutated human RET gene (collaboration with Pr Anne Barlier and Massimo Santoro). However, this approach does not allow to categorize these variants with certainty. To complete these data and to classify VSI more precisely, we chose to do a metabolomic analysis. An initial analysis by mass spectrometry (QTRAP 5500, AB Sciex) using the Biocrates kit p180 will allow the identification and analysis of about 180 endogenous molecules of the following families: acylcarnitines, amino acids, biogenic amines, glycerophospholipids, sphingolipids and sugars. This analysis could allow us to identify the metabolic pathways preferentially used by these cancer cells and to classify the RET gene variants according to their metabolism.

• Hormonal receptivity and analytical interferences

We are a reference center for rare diseases of the thyroid and hormonal receptors since 2006. The center was renewed as a coordinating reference center in the FIRENDO (rare endocrine diseases) sector in December 2017. The center is particularly involved in the pathologies of thyroid hormone receptivity. We have implemented by high-throughput sequencing the analysis of about fifty genes involved in the thyroid drive axis. This development allowed us to propose genetic analyzes in the context of congenital hypothyroidism (prevalence 1/3000 in France). This service makes it possible to adapt the genetic counseling and to propose a adapted follow-up of the children.

I set up a retrospective study then prospective on the cases of the syndromes of sensitivities reduced to the thyroid hormones. This syndrome has a biological definition: a high concentration of thyroxine (T4) and / or triiodothyronine (T3) without the expected effects and in particular the absence of a decline in TSH. The causes are 85% related to a genetic variant of the beta thyroid hormone receptor (THRB) gene. Rare cases involving other genes such as coactivators, coreceptors, alpha receptors with thyroid hormones ... have also been described. In 15% no cause is found. However, the differential diagnoses of thyroid hormone dosing artifacts or the very rare thyrotropic pituitary adenomas must be eliminated beforehand. The establishment of the study of about 50 genes by high-throughput sequencing allowed us to find variants in the genes known to be involved in hypothyroidism, in the cellular transport of thyroid hormones and especially in proteins. carriers such as albumin, transthyretin and TBG. In order to improve the detection of the artefacts of assays, we wish to set up the reference technique for the determination of thyroid hormones, ie detection by mass spectrometry. This quantification should allow us to eliminate the assay artifacts related to the immunoassay technique and to select the patients actually carrying a syndrome of reduced sensitivity to thyroid hormones.

Main publications and patents from the last 5 years

• Lebeault M, Pinson S, Guillaud-Bataille M, Gimenez-Roqueplo AP, Carrie A, Barbu V, Pigny P, Bezieau S, Rey JM, Delvincourt C, Giraud S, Veyrat-Durebex C, Saulnier P, Bouzamondo N, Chabbert M, Blin J, Mohamed A, Romanet P, Borson-Chazot, F, Rohmer V, Barlier A, Mirebeau-Prunier D. Nationwide French Study of RET Variants Detected from 2003 to 2013 Suggests a Possible Influence of Polymorphisms as Modifiers. Thyroid. 2017;27:1511-1522. ( IF5.515)
• Prunier F, Reynier P, Mirebeau-Prunier D. Letter in response to remote ischaemic conditioning provides humoral cross-species cardioprotection through glycine receptor activation. Cardiovasc Res. 2017;113:562. (IF 5.465)
• Chao de la Barca JM, Simard G, Sarzi E, Chaumette T, Rousseau G, Chupin S, Gadras C, Tessier L, Ferré M, Chevrollier A, Desquiret-Dumas V, Gueguen N, Leruez S, Verny C, Miléa D, Bonneau D, Amati-Bonneau P, Procaccio V, Hamel C, Lenaers G, Reynier P, Prunier-Mirebeau D. Targeted Metabolomics Reveals Early Dominant Optic Atrophy Signature in Optic Nerves of Opa1delTTAG/+ Mice. Invest Ophthalmol Vis Sci, 2017;58:812-820. (IF 3.427)
• Le Page S, Niro M, Fauconnier J,Cellier L, Tamareille S, Gharib A, Chevrollier A, Loufrani L, Grenier C, Kamel R, Sarzi E, Lacampagne, Ovize M, Henrion D, Reynier P, Lenaers G, Mirebeau-Prunier D, Prunier F. Increase in Cardiac Ischemia-Reperfusion Injuries in Opa1+/- Mouse Model. PLoS ONE 2016;11:e0164066. (IF 3.057)
• Chao de la Barca JM, Bakhta O, Kalakech H, Simard G, Tamareille S, Catros V, Callebert, Gadras C, Tessier L, Reynier P, Prunier F, Mirebeau-Prunier D. Metabolic Signature of Remote Ischemic Preconditioning Involving a Cocktail of Amino Acids and Biogenic Amines. J Am Heart Assoc 2016;5. (IF 5.117)
• Cellier L ; Tamareille S ; Kalakech H ; Guillou S ; Lenaers G ; Prunier F, Mirebeau-Prunier D. Remote Ischemic Conditioning Influences Mitochondrial Dynamics. Shock, 2016;45:192-7. ( IF 3.113)
• Mirebeau-Prunier D, Le Pennec S, Jacques C, et al. Estrogen-related receptor alpha modulates lactate dehydrogenase activity in thyroid tumors. PLoS One 2013;8:e58683. (IF 3.057)
• Mirebeau-Prunier D, Le Pennec S, Jacques C et al. Estrogen-related receptor alpha and PGC-1-related coactivator constitute a novel complex mediating the biogenesis of functional mitochondria. FEBS J 2010;277:713-25. (IF 3.902)
• Mirebeau D, Acquaviva C, Suciu S et al. The prognostic significance of CDKN2A, CDKN2B and MTAP inactivation in B-lineage acute lymphoblastic leukemia of childhood. Results of the EORTC studies 58881 and 58951. Haematologica 2006;91:881-5. (IF 7.702)
• Mirebeau-Prunier D, Guyétant S, Rodien P et al. Decreased expression of thyrotropin receptor gene suggests a high-risk subgroup for oncocytic adenoma. Eur J Endocrinol 2004;150:269-76. (IF 4.101)