Cytotoxicity of Local Anesthetics in Human Neuronal CellsBACKGROUND: In addition to inhibiting the excitation conduction process in peripheral nerves, local anesthetics (LAs) cause toxic effects on the central nervous system, cardiovascular system, neuromuscular junction, and cell metabolism. Different postoperative neurological complications are ascribed to the cytotoxicity of LAs, but the underlying mechanisms remain unclear. Because the clinical concentrations of LAs far exceed their EC(50) for inhibiting ion channel activity, ion channel block alone might not be sufficient to explain LA-induced cell death. However, it may contribute to cell death in combination with other actions. In this study, we compared the cytotoxicity of six frequently used LAs and will discuss the possible mechanism(s) underlying their toxicity. METHODS: In human SH-SY5Y neuroblastoma cells, viability upon exposure to six LAs (bupivacaine, ropivacaine, mepivacaine, lidocaine, procaine, and chloroprocaine) was quantitatively determined by the MTT-(3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetra-odium bromide) colorimetry assay and qualitatively confirmed by fluorescence imaging, using the LIVE/DEAD assay reagents (calcein/AM and ethidium homodimer-1). In addition, apoptotic activity was assessed by measuring the activation of caspase-3/-7 by imaging using a fluorescent caspase inhibitor (FLICA). Furthermore, LA effects on depolarization- and carbachol-stimulated intracellular Ca(2+)-responses were also evaluated. RESULTS: 1) After a 10-min treatment, all six LAs decreased cell viability in a concentration-dependent fashion. Their killing potency was procaine < or = mepivacaine < lidocaine < chloroprocaine < ropivacaine < bupivacaine (based on LD(50), the concentration at which 50% of cells were dead). Among these six LAs, only bupivacaine and lidocaine killed all cells with increasing concentration. 2) Both bupivacaine and lidocaine activated caspase-3/-7. Caspase activation required higher levels of lidocaine than bupivacaine. Moreover, the caspase activation by bupivacaine was slower than by lidocaine. Lidocaine at high concentrations caused an immediate caspase activation, but did not cause significant caspase activation at concentrations lower than 10 mM. 3) Procaine and chloroprocaine concentration-dependently inhibited the cytosolic Ca(2+)-response evoked by depolarization or receptor-activation in a similar manner as a previous observation made with bupivacaine, ropivacaine, mepivacaine, and lidocaine. None of the LAs caused a significant increase in the basal and Ca(2+)-evoked cytosolic Ca(2+)-level. CONCLUSION: LAs can cause rapid cell death, which is primarily due to necrosis. Lidocaine and bupivacaine can trigger apoptosis with either increased time of exposure or increased concentration. These effects might be related to postoperative neurologic injury. Lidocaine, linked to the highest incidence of transient neurological symptoms, was not the most toxic LA, whereas bupivacaine, a drug causing a very low incidence of transient neurological symptoms, was the most toxic LA in our cell model. This suggests that cytotoxicity-induced nerve injury might have different mechanisms for different LAs and different target(s) other than neurons.
Remodeling of Cardiolipin by Phospholipid TransacylationYang Xu, Richard I. Kelley, Thomas J. J. Blanck et al.|Journal of Biological Chemistry|2003 Mitochondrial cardiolipin (CL) contains unique fatty acid patterns, but it is not known how the characteristic molecular species of CL are formed. We found a novel reaction that transfers acyl groups from phosphatidylcholine or phosphatidylethanolamine to CL in mitochondria of rat liver and human lymphoblasts. Acyl transfer was stimulated by ADP, ATP, and ATPγS, but not by other nucleotides. Coenzyme A stimulated the reaction only in the absence of adenine nucleotides. Free fatty acids were not incorporated into CL under the same incubation condition. The transacylation required addition of exogenous CL or monolyso-CL, whereas dilyso-CL was not a substrate. Transacylase activity was decreased in lymphoblasts from patients with Barth syndrome (tafazzin deletion), and this was accompanied by drastic changes in the molecular composition of CL. In rat liver, where linoleic acid was the most abundant residue of CL, only linoleoyl groups were transferred into CL, but not oleoyl or arachidonoyl groups. We demonstrated complete remodeling of tetraoleoyl-CL to tetralinoleoyl-CL in rat liver mitochondria and identified the intermediates linoleoyl-trioleoyl-CL, dilinoleoyl-dioleoyl-CL, and trilinoleoyl-oleoyl-CL by high-performance liquid chromatography. The data suggest that CL is remodeled by acyl specific phospholipid transacylation and that tafazzin is an acyltransferase involved in this mechanism. Mitochondrial cardiolipin (CL) contains unique fatty acid patterns, but it is not known how the characteristic molecular species of CL are formed. We found a novel reaction that transfers acyl groups from phosphatidylcholine or phosphatidylethanolamine to CL in mitochondria of rat liver and human lymphoblasts. Acyl transfer was stimulated by ADP, ATP, and ATPγS, but not by other nucleotides. Coenzyme A stimulated the reaction only in the absence of adenine nucleotides. Free fatty acids were not incorporated into CL under the same incubation condition. The transacylation required addition of exogenous CL or monolyso-CL, whereas dilyso-CL was not a substrate. Transacylase activity was decreased in lymphoblasts from patients with Barth syndrome (tafazzin deletion), and this was accompanied by drastic changes in the molecular composition of CL. In rat liver, where linoleic acid was the most abundant residue of CL, only linoleoyl groups were transferred into CL, but not oleoyl or arachidonoyl groups. We demonstrated complete remodeling of tetraoleoyl-CL to tetralinoleoyl-CL in rat liver mitochondria and identified the intermediates linoleoyl-trioleoyl-CL, dilinoleoyl-dioleoyl-CL, and trilinoleoyl-oleoyl-CL by high-performance liquid chromatography. The data suggest that CL is remodeled by acyl specific phospholipid transacylation and that tafazzin is an acyltransferase involved in this mechanism. Mitochondria contain cardiolipin (CL), 1The abbreviations used are: CLcardiolipin (1,3-diphosphatidylglycerol)MLCLmonolyso-cardiolipinDLCLdilyso-cardiolipinL4tetralinoleoyl-cardiolipinL3Otrilinoleoyl-oleoyl-cardiolipinL2O2dilinoleoyl-dioleoyl-cardiolipinLO3linoleoyl-trioleoyl-cardiolipinO4tetraoleoyl-cardiolipinATPγSadenosine-5′-[γ-thio]triphosphateCoAcoenzyme AHPLChigh-performance liquid chromatographyPCphosphatidylcholinePEphosphatidylethanolamine.1The abbreviations used are: CLcardiolipin (1,3-diphosphatidylglycerol)MLCLmonolyso-cardiolipinDLCLdilyso-cardiolipinL4tetralinoleoyl-cardiolipinL3Otrilinoleoyl-oleoyl-cardiolipinL2O2dilinoleoyl-dioleoyl-cardiolipinLO3linoleoyl-trioleoyl-cardiolipinO4tetraoleoyl-cardiolipinATPγSadenosine-5′-[γ-thio]triphosphateCoAcoenzyme AHPLChigh-performance liquid chromatographyPCphosphatidylcholinePEphosphatidylethanolamine. a unique phospholipid with two phosphate groups and four fatty acids. Throughout the eukaryotic kingdom, CL is the signature lipid of the cristae membrane, suggesting it may play an essential role in mitochondrial physiology (1Hostetler K.Y. Hawthorne J.N. Ansell G.B. Phospholipids. Elsevier Press, Amsterdam1982: 215-261Google Scholar, 2Daum G. Biochim. Biophys. Acta. 1985; 822: 1-42Crossref PubMed Scopus (700) Google Scholar, 3Dowhan W. Annu. Rev. Biochem. 1997; 66: 199-232Crossref PubMed Scopus (766) Google Scholar, 4Schlame M. Rua D. Greenberg M.L. Progr. Lipid Res. 2000; 39: 257-288Crossref PubMed Scopus (651) Google Scholar). However, only recently has the availability of CL-lacking yeast strains offered a first glimpse into this role. In the absence of CL, coupling between respiration and protonmotive force is impaired, causing reduction of the mitochondrial membrane potential (Δψ) and inhibition of Δψ-dependent functions, such as oxidative phosphorylation and protein import (5Jiang F. Ryan M.T. Schlame M. Zhao M. Gu Z. Klingenberg M. Pfanner N. Greenberg M.L. J. Biol. Chem. 2000; 275: 22387-22394Abstract Full Text Full Text PDF PubMed Scopus (319) Google Scholar, 6Koshkin V. Greenberg M.L. Biochem. J. 2000; 347: 687-691Crossref PubMed Scopus (108) Google Scholar, 7Koshkin V. Greenberg M.L. Biochem. J. 2002; 364: 317-322Crossref PubMed Scopus (111) Google Scholar). CL is firmly integrated into the quarternary structure of mitochondrial protein complexes (4Schlame M. Rua D. Greenberg M.L. Progr. Lipid Res. 2000; 39: 257-288Crossref PubMed Scopus (651) Google Scholar, 8Robinson N.C. J. Bioenerg. Biomembr. 1993; 25: 153-162Crossref PubMed Scopus (223) Google Scholar). For instance, the position of CL in the crystal structure of the bc1 complex suggests that it participates directly in proton conduction (9Lange C. Nett J.H. Trumpower B.L. Hunte C. EMBO J. 2001; 20: 6591-6600Crossref PubMed Scopus (356) Google Scholar), providing a rationale for the dependence of Δψ on CL. CL also supports the functional membrane conformation of the ADP-ATP carrier (10Beyer K. Klingenberg M. Biochemistry. 1985; 24: 3821-3826Crossref PubMed Scopus (272) Google Scholar, 11Schlame M. Beyer K. Hayer-Hartl M. Klingenberg Eur. J. Biochem. 1991; 199: 459-466Crossref PubMed Scopus (59) Google Scholar, 12Beyer K. Nuscher B. Biochemistry. 1996; 35: 15784-15790Crossref PubMed Scopus (92) Google Scholar), and it is required for the assembly of mitochondrial supercomplexes (13Zhang M. Mileykovskaya E. Dowhan W. J. Biol. Chem. 2002; 277: 43553-43556Abstract Full Text Full Text PDF PubMed Scopus (488) Google Scholar). cardiolipin (1,3-diphosphatidylglycerol) monolyso-cardiolipin dilyso-cardiolipin tetralinoleoyl-cardiolipin trilinoleoyl-oleoyl-cardiolipin dilinoleoyl-dioleoyl-cardiolipin linoleoyl-trioleoyl-cardiolipin tetraoleoyl-cardiolipin adenosine-5′-[γ-thio]triphosphate coenzyme A high-performance liquid chromatography phosphatidylcholine phosphatidylethanolamine. cardiolipin (1,3-diphosphatidylglycerol) monolyso-cardiolipin dilyso-cardiolipin tetralinoleoyl-cardiolipin trilinoleoyl-oleoyl-cardiolipin dilinoleoyl-dioleoyl-cardiolipin linoleoyl-trioleoyl-cardiolipin tetraoleoyl-cardiolipin adenosine-5′-[γ-thio]triphosphate coenzyme A high-performance liquid chromatography phosphatidylcholine phosphatidylethanolamine. It is not known how the function of CL is related to its unique structure. CL is not only a dimeric phospholipid, it also has an unusual composition of fatty acids (2Daum G. Biochim. Biophys. Acta. 1985; 822: 1-42Crossref PubMed Scopus (700) Google Scholar, 4Schlame M. Rua D. Greenberg M.L. Progr. Lipid Res. 2000; 39: 257-288Crossref PubMed Scopus (651) Google Scholar, 11Schlame M. Beyer K. Hayer-Hartl M. Klingenberg Eur. J. Biochem. 1991; 199: 459-466Crossref PubMed Scopus (59) Google Scholar). In some animal and plant tissues, CL contains mostly linoleic acid (1Hostetler K.Y. Hawthorne J.N. Ansell G.B. Phospholipids. Elsevier Press, Amsterdam1982: 215-261Google Scholar, 2Daum G. Biochim. Biophys. Acta. 1985; 822: 1-42Crossref PubMed Scopus (700) Google Scholar, 4Schlame M. Rua D. Greenberg M.L. Progr. Lipid Res. 2000; 39: 257-288Crossref PubMed Scopus (651) Google Scholar). For instance, in mammalian heart, tetralinoleoyl-CL makes up 70-80% of all molecular species (14Schlame M. Towbin J.A. Heerdt P.M. Jehle R. DiMauro S. Blanck T.J.J. Ann. Neurol. 2002; 51: 634-637Crossref PubMed Scopus (217) Google Scholar). In certain marine animals CL contains almost exclusively arachidonic acid (15Kraffe E. Soudant P. Marty Y. Kervarec N. Jehan P. Lipids. 2002; 37: 507-514Crossref PubMed Scopus (31) Google Scholar), whereas in yeast the dominant acyl groups are oleic and palmitoleic acid (11Schlame M. Beyer K. Hayer-Hartl M. Klingenberg Eur. J. Biochem. 1991; 199: 459-466Crossref PubMed Scopus (59) Google Scholar). Regardless of these variations, in most cells CL maintains a distinct fatty acid pattern that is dominated by only one or two acyl species. CL is relatively resistant to dietary manipulation of its fatty acid pattern (16Wolff R.L. Reprod. Nutr. Dev. 1988; 28: 1489-1507Crossref PubMed Scopus (17) Google Scholar, 17Wolff R.L. Lipids. 1995; 30: 893-898Crossref PubMed Scopus (17) Google Scholar), suggesting that the acyl composition has functional significance. This idea was supported by the discovery of abnormal CL species in patients with Barth syndrome (14Schlame M. Towbin J.A. Heerdt P.M. Jehle R. DiMauro S. Blanck T.J.J. Ann. Neurol. 2002; 51: 634-637Crossref PubMed Scopus (217) Google Scholar, 18Valianpour F. Wanders R.J.A. Overmars H. Vreken P. van Gennip A.H. Baas F. Plecko B. Santer R. Barth P.G. J. Pediatr. 2002; 141: 729-733Abstract Full Text Full Text PDF PubMed Scopus (88) Google Scholar, 19Schlame M. Kelley R.I. Feigenbaum A. Towbin J.A. Heerdt P. Schieble T. Wanders R.J.A. DiMauro S. Blanck T.J.J. J. Am. Coll. Cardiol. 2003; 42: 1994-1999Crossref PubMed Scopus (155) Google Scholar), an X-linked disease associated with cardiomyopathy, skeletal myopathy, neutropenia, and growth retardation (20Barth P.G. Wanders R.J.A. Vreken P. Janssen E.A. Lam J. Baas F. J. Inher. Metab. Dis. 1999; 22: 555-567Crossref PubMed Scopus (109) Google Scholar). Barth syndrome is caused by mutations of tafazzin (21Bione S. D'Adamo P. Maestrini E. Gedeon A.K. Bolhuis P.A. Toniolo D. Nat. Gen. 1996; 12: 385-389Crossref PubMed Scopus (593) Google Scholar). Because tafazzin is a putative acyltransferase (22Neuwald A.F. Curr. Biol. 1997; 7: R465-R466Abstract Full Text Full Text PDF PubMed Google Scholar), the abnormal composition of CL may be a direct result of acyltransferase deficiency, and it may play a crucial role in the pathogenesis of Barth syndrome. CL species are thought to emerge from remodeling of acyl groups subsequent to de novo formation (23Schlame M. Rüstow B. Biochem. J. 1990; 272: 589-595Crossref PubMed Scopus (104) Google Scholar, 24Ma B.J. Taylor W.A. Dolinsky V.W. Hatch G.M. J. Lipid Res. 1999; 40: 1837-1845Abstract Full Text Full Text PDF PubMed Google Scholar). The remodeling idea was suggested by (i) lack of acyl specificity in the de novo pathway (25Hostetler K.Y. Galesloot J.M. Boer P. van den Bosch H. Biochim. Biophys. Acta. 1975; 380: 382-389Crossref PubMed Scopus (71) Google Scholar, 26Rüstow B. Schlame M. Rabe H. Reichmann G. Kunze D. Biochim. Biophys. Acta. 1989; 1002: 261-263Crossref PubMed Scopus (35) Google Scholar), (ii) independent turnover of acyl and glycerol moieties of CL (27Landriscina C. Megli F.M. Quagliariello E. Lipids. 1976; 11: 61-66Crossref PubMed Scopus (38) Google Scholar), and (iii) presence of lyso-CLs in mitochondria (23Schlame M. Rüstow B. Biochem. J. 1990; 272: 589-595Crossref PubMed Scopus (104) Google Scholar). The classical mechanism of phospholipid remodeling is the Lands cycle in which endogenous fatty acids are removed from phospholipids by phospholipase A2 and new fatty acids are re-attached by acyl-CoA:lysophospholipid acyltransferase. This process requires activation of fatty acids by MgATP and CoA. Acyl-CoA:MLCL acyltransferase activity has been identified in liver and heart, but it failed to show the anticipated linoleoyl specificity (24Ma B.J. Taylor W.A. Dolinsky V.W. Hatch G.M. J. Lipid Res. 1999; 40: 1837-1845Abstract Full Text Full Text PDF PubMed Google Scholar, 28Taylor W.A. Hatch G.M. J. Biol. Chem. 2003; 278: 12716-12721Abstract Full Text Full Text PDF PubMed Scopus (68) Google Scholar). In this paper we report that acyl groups can be transferred directly from phosphatidylcholine (PC) or phosphatidylethanolamine (PE) to CL. This transacylation is linoleoyl-specific in liver and it is deficient in patients with Barth syndrome. Materials—CL from bovine heart, CoA, linoleoyl-CoA, phospholipase A2 from Naja naja venom, and all nucleotides were purchased from Sigma. Tetralinoleoyl-CL (1,3-di[1′,2′-dilinoleoyl-glycero-3′-phosphoryl]-glycerol) was purified from bovine heart CL as described previously (29Schlame M. Haller I. Sammaritano L.R. Blanck T.J.J. Thromb. Haemost. 2001; 86: 1475-1482Crossref PubMed Scopus (22) Google Scholar). Tetraoleoyl-CL (1,3-di[1′,2′-dioleoyl-glycero-3′-phosphoryl]-glycerol), tetramyristoyl-CL (1,3-di[1′,2′-dimyristoyl-glycero-3′-phosphoryl]-glycerol), MLCL (1-[1′,2′-diacyl-glycero-3′-phosphoryl]-3-[1′-acyl-glycero-3′-phosphoryl]-glycerol), and DLCL (1,3-di[1′-acyl-glycero-3′-phosphoryl]-glycerol) were obtained from Avanti Polar Lipds (Alabaster, AL). Methyl arachidonyl fluorophosphonate was supplied by Cayman Chemicals (Ann Arbor, MI). 1-Palmitoyl-2-[1′-14C]linoleoyl-PC (52 Ci/mol), 1-palmitoyl-2-[1′-14C]oleoyl-PC (56 Ci/mol), 1-palmitoyl-2-[1′-14C]linoleoyl-PE (54 Ci/mol), and 1-acyl-2-[1′-14C]arachidonoyl-PE (56 Ci/mol) were purchased from Amersham Biosciences. [1-14C]Linoleic acid (51 Ci/mol) and 1-palmitoyl-2-[1′-14C]arachidonoyl-PC (48 Ci/mol) were obtained from Perkin Elmer (Boston, MA). The scintillation fluid Ecoscint was purchased from National Diagnostics (Atlanta, GA). Rat Liver Mitochondria—Mitochondria were prepared from liver of Sprague-Dawley rats (100-200 g). Animals were housed in the Central Animal Care Facility of New York University Medical Center. Treatment was approved by the institutional animal care and use committee. Rats were anesthetized by pentobarbital injection, and the liver was placed in ice-cold saline. The liver was minced, washed, and homogenized in isolation buffer (150 mm sucrose, 50 mm KCl, 20 mm Hepes, 2 mm 2-mercaptoethanol, 1 mm EDTA, pH 7.4, at 4 °C), adjusting a ratio of 10 ml/g wet weight. The homogenate was spun at 750 × g for 5 min and the supernatant was spun again at 8,000 × g for 4 min. The mitochondrial pellet was suspended in 50 ml isolation buffer followed by centrifugation at 17,000 × g for 80 s. Mitochondria were resuspended in isolation buffer and spun at 17,000 × g for 10 min. The final pellet was resuspended at a concentration of 50 mg protein per milliliter isolation buffer and stored at -80 °C. Rat liver microsomes were prepared from the first post-mitochondrial supernatant by centrifugation at 100,000 × g for 90 min. Protein concentrations of mitochondria and microsomes were determined by the method of Lowry (30Lowry O.H. Rosebrough N.J. Farr A.L. Randall R.J. J. Biol. Chem. 1951; 193: 265-275Abstract Full Text PDF PubMed Google Scholar). Human Lymphoblast Mitochondria—Lymphoblastoid cell lines were established by Epstein-Barr virus transformation of leukocytes isolated from whole blood of three patients with Barth syndrome and three control subjects. All patients with Barth syndrome had a mutation in the tafazzin gene (G4.5) that resulted in total deletion of the protein. In addition these patients met clinical criteria of Barth syndrome, i.e. they had cardiomyopathy plus two or more of the following non-cardiac symptoms: skeletal myopathy, neutropenia, growth retardation, and increased urinary excretion of 3-methylglutaconic acid. Patients or their guardians gave written informed consent. The protocol was approved by the Institutional Review Board of Johns Hopkins University. Lymphoblasts were grown in RPMI were and × cells were in growth for isolation of mitochondria 1996; PubMed Google Scholar). this cells were suspended in 4 ml isolation mm mm sucrose, 1 mm bovine and 5 mm Hepes, pH were with more of to 1996; PubMed Google Scholar). cells were in a and cells were removed by centrifugation at × g for 5 min. mitochondria were spun at × g for 20 min and they were in isolation of Mitochondria—Mitochondria mg were in ml 50 mm pH 10 mm 2-mercaptoethanol, mm EDTA, 4 mm ADP, acyl or and acyl or at of phospholipids were and were buffer was and phospholipids were by in a for s. The incubation was by addition of mitochondria and it was by addition of 2 ml was min. In some we used other nucleotides or of In other we used acid as acyl The concentration of some was to their and their were by Lipid were by the of and J. Biochem. 37: PubMed Scopus Google Scholar). The were under a of and resuspended in ml were to supplied by and by chromatography with in the first and in the were with of of were transferred into scintillation was by liquid scintillation with All data were as Treatment with incubation of rat liver and were isolated by chromatography as described were from into and in 1 ml of of ml of buffer was of phospholipase 50 mm 50 mm Hepes, pH and mm The was at were J. Biochem. 37: PubMed Scopus Google and by chromatography on with The of were identified with the of The were into and was in by liquid scintillation of was isolated from incubation by chromatography as described was in ml and into a × was at a of the composition from to in 80 min. A and 10 mm phosphate buffer at a ratio of and at a ratio of of ml were into scintillation The was and was by liquid scintillation of CL species were determined in (29Schlame M. Haller I. Sammaritano L.R. Blanck T.J.J. Thromb. Haemost. 2001; 86: 1475-1482Crossref PubMed Scopus (22) Google Scholar). of of mg were with J. Biochem. 37: PubMed Scopus Google Scholar), and CL was to as described M. S. S. T. T. DiMauro S. Blanck T.J.J. J. Lipid Res. 1999; 40: Full Text Full Text PDF PubMed Google Scholar). The CL was purified by and by with M. S. S. T. T. DiMauro S. Blanck T.J.J. J. Lipid Res. 1999; 40: Full Text Full Text PDF PubMed Google Scholar). A (150 × was The was 2 The was with for 10 min. that the concentration of was increased from to min. species of cardiolipin were identified by of fatty acids M. S. S. T. T. DiMauro S. Blanck T.J.J. J. Lipid Res. 1999; 40: Full Text Full Text PDF PubMed Google Scholar). in Rat Liver liver mitochondria were with and of decreased by this 50 was as linoleic was in CL, 10 was in was in and was in other phospholipids of groups into CL and at a that was increased by acid was the of in CL, we acid to the incubation of The of acid was to the of acid incubation with In the presence of of into CL was In was an of to CL Acyl transfer from dependence on and incubation as the transfer from not suggest that groups were transferred from or to CL by of into CL in the presence of Rat liver mitochondria were with ADP, and the for min. were and by chromatography. was in CL. are of three We the that the transacylation was a result of phospholipase A2 of phospholipase A2 by 10 arachidonyl fluorophosphonate had on the reaction in the presence of ADP, and was of activity suggesting that A2 were involved in The specific transacylation activity was in the mitochondrial in the suggesting a specific on mitochondria or We the of nucleotides on ADP, ATP, and stimulated transfer from to CL and from to whereas other nucleotides had or at a concentration of mm increased transfer from into CL and the concentration required for was However, the of in the presence of ADP, i.e. transfer with was of the transfer with transfer from to CL was on exogenous MLCL exogenous CL was of the same of transfer was The concentration required for was protein for CL and protein for In DLCL was not a of the transacylation of exogenous CL, and DLCL on transfer from to CL. Rat liver mitochondria were with ADP, and concentrations of CL, or were and by chromatography. was in of CL in Rat Liver acyl specificity of the transacylation between and CL, we rat liver mitochondria with species of and species of CL. was an acyl in the presence of or In in the transacylation of the acyl Acyl transfer from to CL also linoleoyl the specificity was i.e. transacylation from to CL was only from to CL of liver mitochondria with and tetraoleoyl-CL to complete the tetralinoleoyl-CL as as the intermediates and all of which were identified by high-performance liquid chromatography all acyl of CL were to This was suggested by of isolated with phospholipase of as linoleic acid as DLCL and as MLCL The data are with the following remodeling phospholipase A2 of as linoleic acid from the position of in Human Lymphoblast phospholipid transacylation in mitochondria from human cell was with oleic acid was the most abundant residue in CL, and of lymphoblasts. in liver, activity was stimulated by and it was independent of in the presence of not In lymphoblasts with tafazzin deficiency, from patients with Barth syndrome, transfer from to CL and to was the same were drastic changes in the molecular composition of CL in i.e. the four characteristic CL species of oleic and palmitoleic were by molecular species a more fatty acid pattern We demonstrated formation of tetralinoleoyl-CL by linoleoyl-specific phospholipid transacylation in rat liver groups were transferred directly from or (i) linoleic acid was not incorporated into CL under that of linoleoyl and (ii) linoleoyl transfer not 1 and The of a transacylation that CL is remodeled with fatty acids (23Schlame M. Rüstow B. Biochem. J. 1990; 272: 589-595Crossref PubMed Scopus (104) Google Scholar). the one the data that and stimulated on the other we found that was for transfer from to CL. had or ATPγS, and not linoleoyl transfer in the presence of adenine nucleotides. However, may be the of the transacylation it required adenine concentrations 2 mm for also between and suggesting that all three mitochondrial phospholipids can acyl groups by this mechanism. We found a reaction in human The activity was decreased in lymphoblasts with tafazzin deletion suggesting that tafazzin a role in the transacylation mechanism. deletion also resulted in composition of CL, suggesting that tafazzin is involved in CL remodeling was to be an acyltransferase on its acid (22Neuwald A.F. Curr. Biol. 1997; 7: R465-R466Abstract Full Text Full Text PDF PubMed Google Scholar). of tafazzin and the pattern is characteristic for cell and (21Bione S. D'Adamo P. Maestrini E. Gedeon A.K. Bolhuis P.A. Toniolo D. Nat. Gen. 1996; 12: 385-389Crossref PubMed Scopus (593) Google Scholar). It was suggested that tafazzin may acyl specificity (22Neuwald A.F. Curr. Biol. 1997; 7: R465-R466Abstract Full Text Full Text PDF PubMed Google Scholar). Acyl specificity of the transacylation to the molecular composition of CL in liver and In liver where CL was in linoleic we found of the such specificity was in lymphoblasts where CL mostly oleic and palmitoleic acid. the data supported the that phospholipid transacylation is the mechanism of CL The data also suggested that CL was not from the remodeling was This is more of the remodeling in liver were tetralinoleoyl-CL and trilinoleoyl-oleoyl-CL a result that be by acyl with exogenous CL and MLCL had on the transacylation suggesting that acyl groups with the putative we the presence of an complex that is in with acyl transfer This complex may acyl groups with other by transacylation the molecular species of CL of acyl groups into the complex may show acyl In of acyl groups is species of CL are of the acyl fatty acids may in CL by of and more one is involved in this mechanism. been to be in A. T. K. J. Biochem. 1997; PubMed Scopus Google Scholar). In only between and fatty acids A. T. K. J. Biochem. 1997; PubMed Scopus Google Scholar, H. Biochim. Biophys. Acta. 1991; PubMed Scopus Google Scholar). This is also for purified acyltransferase from liver mitochondria W.A. Hatch G.M. J. Biol. Chem. 2003; 278: 12716-12721Abstract Full Text Full Text PDF PubMed Scopus (68) Google Scholar). this specific of linoleoyl groups into CL, it may be involved in the remodeling For it may transfer acyl groups of CL to or other acyl