Letter to the Editor
To the Editor
We read the article of Osuka et al. (1) entitled “A Protective Role for Inflammasome Activation Following Injury” with great interest. However, we are concerned that the authors have not sufficiently ruled out the possibility that the major effects attributed to inflammasome inhibition were merely due to the solvent used.
The authors describe inflammasome activation in burned mice 1 day after injury as revealed by caspase 1 activation and increased interleukin 1β (IL-1β) production. Interestingly, the data suggest that inhibiting caspase 1 activity—and thereby inhibiting inflammasome activation—with the Ac-YVAD-cmk peptide did not reduce inflammation as expected. On the contrary, it caused a significantly higher mortality and increased expression of the proinflammatory cytokines IL-6 and IL-33 as compared with untreated burned mice. The authors therefore conclude that inflammasome activation might have a protective role following severe injury. Inhibition of (pro)caspase 1 activity shifts the proportion between pro–caspase 1 and mature processed caspase 1 toward the pro form. Interestingly, a potential role of enzymatically inactive pro–caspase 1 in activating nuclear factor κB (NF-κB) via receptor-interacting protein 2 (RIP2) has been described (2, 3). Interleukin 6 transcription is regulated by an NF-κB–sensitive promotor. Hence, elevated IL-6 expression after inhibition of (pro)caspase 1 could indicate NF-κB activation and might be mediated by pro–caspase 1. The authors show this correlation for the first time in an in vivo model. According to this model, inflammasome formation could reduce the amount of pro–caspase 1 available for NF-κB activation via RIP2 and thereby downregulate this proinflammatory pathway (3).
In the current study, (pro)caspase 1 activation was inhibited by injecting Ac-YVAD-cmk intraperitoneally. Ac-YVAD-cmk is a peptide inhibitor of (pro)caspase 1, which is typically dissolved in dimethyl sulfoxide (DMSO). Dimethyl sulfoxide is used in a variety of fields. For example, it is commonly applied as a cryoprotectant of cultured cells and as a solvent for hydrophobic compounds in biological studies. However, it is important to note that DMSO is biologically active (4). Although it can often be used safely as a drug vehicle there is a lot of evidence that it can have undesirable anti-inflammatory and/or proinflammatory, nonspecific, and toxic effects in vitro and in vivo (5). For instance, treatment with 1% DMSO activates NF-κB in THP-1 and HL-60 cells (6, 7), and DMSO increases lipopolysaccharide-induced IL-1β secretion in a dose dependent-manner in human peripheral blood mononuclear cells and mice (8). Furthermore, it induces neurophysiological changes in rats and leads to lymphocyte apoptosis in murine lymphoid organs (9, 10). These effects are undesirable and unpredictable when DMSO is used as solvent. Therefore, control experiments of the effects of DMSO itself always have to be performed. Unfortunately, in the study of Osuka et al., the control group was treated with saline solution. Thus, DMSO could have contributed to the enhanced expression of proinflammatory cytokines and the increased mortality in the Ac-YVAD-cmk–treated burned-mice group. Therefore, the effect of Ac-YVAD-cmk and the conclusions drawn by the authors remain unsettled.
Michael C. Heyman
Department of Pediatrics
University Medical Center Carl Gustav Carus
1. Osuka A, Hanschen M, Stoecklein V, Lederer JA: A protective role for inflammasome activation following injury. Shock 37 (1): 47–55, 2012.
2. Lamkanfi M, Kalai M, Saelens X, Declercq W, Vandenabeele P: Caspase-1 activates nuclear factor of the kappa-enhancer in B cells independently of its enzymatic activity. J Biol Chem 279 (23): 24785–24793, 2004.
3. Sarkar A, Duncan M, Hart J, Hertlein E, Guttridge DC, Wewers MD: ASC directs NFkappaB activation by regulating receptor interacting protein-2 (RIP2) caspase-1 interactions. J Immunol 176 (8): 4979–4986, 2006.
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7. Lee YR, Shim HJ, Yu HN, Song EK, Park J, Kwon KB, Park JW, Rho HW, Park BH, Han MK, et al..: Dimethylsulfoxide induces upregulation of tumor suppressor protein PTEN through nuclear factor-kappab activation in HL-60 cells. Leuk Res 29 (4): 401–405, 2005.
8. Xing L, Remick DG: Mechanisms of dimethyl sulfoxide augmentation of IL-1 beta production. J Immunol 174 (10): 6195–6202, 2005.
9. Cavaletti G, Oggioni N, Sala F, Pezzoni G, Cavalletti E, Marmiroli P, Petruccioli MG, Frattola L, Tredici G: Effect on the peripheral nervous system of systemically administered dimethylsulfoxide in the rat: a neurophysiological and pathological study. Toxicol Lett 118 (1–2): 103–107, 2000.
10. Aita K, Irie H, Tanuma Y, Toida S, Okuma Y, Mori S, Shiga J: Apoptosis in murine lymphoid organs following intraperitoneal administration of dimethyl sulfoxide (DMSO). Exp Mol Pathol 79 (3): 265–271, 2005.