Abstract
Macrophages have vital roles in innate immunity by modulating the inflammatory response via their ability to alter their phenotype from pro-inflammatory (M1) to anti-inflammatory (M2). Aging increases activation of the innate immune system, and macrophage numbers increase in the aged liver. Since macrophages also produce free radical molecules, they are a potential source of age-related oxidative injury in the liver. This study evaluated macrophage phenotype in the aged liver and whether the increase in the number of macrophages with aging is associated with enhanced hepatic oxidative stress. Hepatic macrophage phenotype and oxidative stress were evaluated 2 days after a single intraperitoneal injection of saline or gadolinium chloride (GdCl3, 10 mg/kg) in young (6 months) and aged (24 months) Fischer 344 rats. GdCl3 has been shown to decrease the expression of macrophage-specific markers and impair macrophage phagocytosis in the liver. Saline-treated aged rats demonstrated greater numbers of both M1 (HO-1+/iNOS+) and M2 (HO-1+/CD163+) macrophages, without evidence of a phenotypic shift. GdCl3 did not alter levels of dihydroethidium fluorescence or malondialdehyde, suggesting that macrophages are not a major contributor to steady-state levels of oxidative stress. However, GdCl3 decreased M1 and M2 macrophage markers in both age groups, an effect that was attenuated in aged rats. In old animals, GdCl3 decreased iNOS expression to a greater extent than HO-1 or CD163. These results suggest a novel effect of aging on macrophage biology and that GdCl3 shifts hepatic macrophage polarization to the M2 phenotype in aged animals.
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References
Amanzada A, Moriconi F, Mansuroglu T, Cameron S, Ramadori G, Malik I (2013) Induction of chemokines and cytokines before neutrophils and macrophage recruitment in different regions of rat liver after TAA administration. Lab Invest 94:235–247. https://doi.org/10.1038/labinvest.2013.134
Bauer I, Wanner GA, Rensing H, Alte C, Miescher EA, Wolf B, Pannen BHJ, Clemens MG, Bauer M (1998) Expression pattern of heme oxygenase isoenzymes 1 and 2 in normal and stress-exposed rat liver. Hepatology 27:829–838
Biagi BA, Enyeart JJ (1990) Gadolinium blocks low- and high-threshold calcium currents in pituitary cells. Am J Physiol cell physiol 259(3):C515–C520. https://doi.org/10.1152/ajpcell.1990.259.3.C515
Bloomer SA, Zhang HJ, Brown KE, Kregel KC (2009) Differential regulation of hepatic heme oxygenase-1 protein with aging and heat stress. J Gerontol A Biol Sci Med Sci 64A(4):419–425. https://doi.org/10.1093/gerona/gln056
Bloomer SA, Brown KE, Kregel KC (2019) Renal iron accumulation and oxidative injury with aging: effects of treatment with an iron chelator. J Gerontol Ser A. https://doi.org/10.1093/gerona/glz055
Bogdan C, Vodovotz Y, Nathan C (1991) Macrophage deactivation by interleukin 10. J Exp Med 174(6):1549–1555. https://doi.org/10.1084/jem.174.6.1549
Boscá L, Lazo PA (1994) Induction of nitric oxide release by MRC OX-44 (anti-CD53) through a protein kinase c-dependent pathway in rat macrophages. J Exp Med 179(4):1119–1126. https://doi.org/10.1084/jem.179.4.1119
Brouwer A, Horan MA, Barelds RJ, Knook DL (1986) Cellular aging of the reticuloendothelial system. Arch Gerontol Geriatr 5(4):317–324. https://doi.org/10.1016/0167-4943(86)90034-8
Buechler C, Ritter M, Orsó E, Langmann T, Klucken J, Schmitz G (2000) Regulation of scavenger receptor CD163 expression in human monocytes and macrophages by pro- and antiinflammatory stimuli. J Leuk Biol 67(1):97–103. https://doi.org/10.1002/jlb.67.1.97
Dijkstra CD, Döpp EA, Joling P, Kraal G (1985) The heterogeneity of mononuclear phagocytes in lymphoid organs: distinct macrophage subpopulations in the rat recognized by monoclonal antibodies ED1, ED2 and ED3. Immunology 54(3):589–599
Elmore MRP, Hohsfield LA, Kramar EA, Soreq L, Lee RJ, Pham ST, Najafi AR, Spanenberg EE, Wood MA, West BL, Green KN (2018) Replacement of microglia in the aged brain reverses cognitive, synaptic, and neuronal deficits in aged mice. Aging Cell 17(6):e12832. https://doi.org/10.1111/acel.12832
Fontana L, Zhao E, Amir M, Dong H, Tanaka K, Czaja MJ (2013) Aging promotes the development of diet-induced murine steatohepatitis but not steatosis. Hepatology 57(3):995–1004. https://doi.org/10.1002/hep.26099
Fukuda M, Yokoyama H, Mizukami T, Ohgo H, Okamura Y, Kamegaya Y, Horie Y, Kato S, Ishii H (2004) Kupffer cell depletion attenuates superoxide anion release into the hepatic sinusoids after lipopolysaccharide treatment. J Gastroenterol Hepatol 19(10):1155–1162. https://doi.org/10.1111/j.1440-1746.2004.03408.x
Goda N, Suzuki K, Naito M, Takeoka S, Tsuchida E, Ishimura Y, Tamatani T, Suematsu M (1998) Distribution of heme oxygenase isoforms in rat liver topographic basis for carbon monoxide-mediated microvascular relaxation. J Clin Invest 101(3):604–612. https://doi.org/10.1172/jci1324
Haak JL, Buettner GR, Spitz DR, Kregel KC (2009) Aging augments mitochondrial susceptibility to heat stress. Am J Physiol Reg Int Comp Physiol 296(3):R812–R820. https://doi.org/10.1152/ajpregu.90708.2008
Hardonk MJ, Dijkhuis FWJ, Hulstaert CE, Koudstaal J (1992) Heterogeneity of rat liver and spleen macrophages in gadolinium chloride–induced elimination and repopulation. J Leuk Biol 52(3):296–302. https://doi.org/10.1002/jlb.52.3.296
Harrison-Findik DD, Klein E, Evans J, Gollan J (2009) Regulation of liver hepcidin expression by alcohol in vivo does not involve kupffer cell activation or TNF-α signaling. Am J Physiol-Gastrointest Liver Physiol 296(1):G112–G118. https://doi.org/10.1152/ajpgi.90550.2008
Hilmer SN, Cogger VC, Le Couteur DG (2007) Basal activity of kupffer cells increases with old age. J Gerontol A Biol Sci Med Sci 62(9):973–978
Husztik E, Lázár G, Párducz A (1980) Electron microscopic study of kupffer-cell phagocytosis blockade induced by gadolinium chloride. Br J Exp Pathol 61(6):624–630
Iimuro Y, Yamamoto M, Kohno H, Itakura J, Fujii H, Matsumoto Y (1994) Blockade of liver macrophages by gadolinium chloride reduces lethality in endotoxemic rats—analysis of mechanisms of lethality in endotoxemia. J Leuk Biol 55(6):723–728. https://doi.org/10.1002/jlb.55.6.723
Isidro RA, Isidro AA, Cruz ML, Hernandez S, Appleyard CB (2015) Double immunofluorescent staining of rat macrophages in formalin-fixed paraffin-embedded tissue using two monoclonal mouse antibodies. Histochem Cell Biol 144(6):613–621. https://doi.org/10.1007/s00418-015-1364-9
Jackaman C, Radley-Crabb HG, Soffe Z, Shavlakadze T, Grounds MD, Nelson DJ (2013) Targeting macrophages rescues age-related immune deficiencies in C57BL/6 J geriatric mice. Aging Cell 12(3):345–357. https://doi.org/10.1111/acel.12062
Jaeschke H, Farhood A (1991) Neutrophil and kupffer cell-induced oxidant stress and ischemia-reperfusion injury in rat liver. Am J Physiol-Gastrointest Liver Physiol 260(3):G355–G362. https://doi.org/10.1152/ajpgi.1991.260.3.G355
Jenkins SJ, Ruckerl D, Cook PC, Jones LH, Finkelman FD, van Rooijen N, MacDonald AS, Allen JE (2011) Local macrophage proliferation, rather than recruitment from the blood, is a signature of Th2 inflammation. Science 332(6035):1284–1288. https://doi.org/10.1126/science.1204351
Khan HA, Ibrahim KE, Khan A, Alrokayan SH, Alhomida AS, Lee YK (2016) Comparative evaluation of immunohistochemistry and real-time PCR for measuring proinflammatory cytokines gene expression in livers of rats treated with gold nanoparticles. Exp Toxicol Pathol 68(7):381–390. https://doi.org/10.1016/j.etp.2016.05.006
Klein AD, Oyarzún JE, Cortez C, Zanlungo S (2018) Gadolinium chloride rescues Niemann-Pick type C liver damage. Int J Mol Sci 19(11):3599. https://doi.org/10.3390/ijms19113599
Knook DL, Brouwer A (1989) Kupffer cells and the acute phase response: the effect of aging. Immunol Invest 18(1–4):339–350
Lee CM, Yeoh GC, Olynyk JK (2004) Differential effects of gadolinium chloride on kupffer cells in vivo and in vitro. Int J Biochem Cell Biol 36(3):481–488. https://doi.org/10.1016/j.biocel.2003.08.004
Maeso-Díaz R, Ortega-Ribera M, Fernández-Iglesias A, Hide D, Muñoz L, Hessheimer AJ, Vila S, Francés R, Fondevila C, Albillos A, Peralta C, Bosch J, Tacke F, Cogger VC, Gracia-Sancho J (2018) Effects of aging on liver microcirculatory function and sinusoidal phenotype. Aging Cell 17(6):e12829. https://doi.org/10.1111/acel.12829
Mahbub S, Deburghgraeve CR, Kovacs EJ (2011) Advanced age impairs macrophage polarization. J Interferon Cytokine Res 32(1):18–26. https://doi.org/10.1089/jir.2011.0058
Metzger Z, Hoffeld JT, Oppenheim JJ (1981) Regulation by PGE2 of the production of oxygen intermediates by LPS-activated macrophages. J Immunol 127(3):1109–1113
Murray PJ, Allen JE, Biswas SK, Fisher EA, Gilroy DW, Goerdt S, Gordon S, Hamilton JA, Ivashkiv LB, Lawrence T, Locati M, Mantovani A, Martinez FO, Mege JL, Mosser DM, Natoli G, Saeij JP, Schultze JL, Shirey KA, Sica A, Suttles J, Udalova I, van Ginderachter JA, Vogel SN, Wynn TA (2014) Macrophage activation and polarization: nomenclature and experimental guidelines. Immunity 41(1):14–20. https://doi.org/10.1016/j.immuni.2014.06.008
Oswald IP, Gazzinelli RT, Sher A, James SL (1992) Il-10 synergizes with Il-4 and transforming growth factor-beta to inhibit macrophage cytotoxic activity. J Immunol 148(11):3578–3582
Parameswaran N, Patial S (2010) Tumor necrosis factor-α signaling in macrophages. Crit Rev Eukaryot Gene Expr 20(2):87–103
Pilkington SM, Barron MJ, Watson REB, Griffiths CEM, Bulfone-Paus S (2019) Aged human skin accumulates mast cells with altered functionality that localize to macrophages and vasoactive intestinal peptide-positive nerve fibres. Bri J Dermatol 180(4):849–858. https://doi.org/10.1111/bjd.17268
Rai R, Zhang J, Clemens M, Diehl A (1996) Gadolinium chloride alters the acinar distribution of phagocytosis and balance between pro- and anti-inflammatory cytokines. Shock 6(4):243–247
Rana B, Mischoulon D, Xie Y, Bucher NL, Farmer SR (1994) Cell-extracellular matrix interactions can regulate the switch between growth and differentiation in rat hepatocytes: reciprocal expression of C/EBP alpha and immediate-early growth response transcription factors. Moll Cell Biol 14(9):5858–5869
Roland CR, Naziruddin B, Mohanakumar T, Flye MW (1996) Gadolinium chloride inhibits kupffer cell nitric oxide synthase (iNOS) induction. J Leuk Biol 60(4):487–492. https://doi.org/10.1002/jlb.60.4.487
Rüttinger D, Vollmar B, Wanner GA, Messmer K (1996) In vivo assessment of hepatic alterations following gadolinium chloride-induced kupffer cell blockade. J Hepatol 25(6):960–967. https://doi.org/10.1016/S0168-8278(96)80302-3
Sholomskas LM, Roche KL, Bloomer SA (2015) Aging impairs induction of redox factor-1 after heat stress: a potential mechanism for heat-induced liver injury. Int J Physiol Pathophysiol Pharmacol 7(1):14–26
Smallwood HS, López-Ferrer D, Squier TC (2011) Aging enhances the production of reactive oxygen species and bactericidal activity in peritoneal macrophages by upregulating classical activation pathways. Biochemistry 50(45):9911–9922. https://doi.org/10.1021/bi2011866
Stein M, Keshav S, Harris N, Gordon S (1992) Interleukin 4 potently enhances murine macrophage mannose receptor activity: a marker of alternative immunologic macrophage activation. J Exp Med 176(1):287–292. https://doi.org/10.1084/jem.176.1.287
Stout RD, Jiang C, Matta B, Tietzel I, Watkins SK, Suttles J (2005) Macrophages sequentially change their functional phenotype in response to changes in microenvironmental influences. J Immunol 175(1):342–349. https://doi.org/10.4049/jimmunol.175.1.342
Uhlén M, Fagerberg L, Hallström BM, Lindskog C, Oksvold P, Mardinoglu A, Sivertsson Å, Kampf C, Sjöstedt E, Asplund A, Olsson I, Edlund K, Lundberg E, Navani S, Szigyarto CAK, Odeberg J, Djureinovic D, Takanen JO, Hober S, Alm T, Edqvist PH, Berling H, Tegel H, Mulder J, Rockberg J, Nilsson P, Schwenk JM, Hamsten M, von Feilitzen K, Forsberg M, Persson L, Johansson F, Zwahlen M, von Heijne G, Nielsen J, Pontén F (2015) Tissue-based map of the human proteome. Science 347(6220):1260419. https://doi.org/10.1126/science.1260419
Videla LA, Tapia G, Fernández V (2001) Influence of aging on Kupffer cell respiratory activity in relation to particle phagocytosis and oxidative stress parameters in mouse liver. Redox Rep 6(3):155–159. https://doi.org/10.1179/135100001101536265
Vömel T, Platt D, Schnorr B (1984) Phagocytic activity of isolated perfused liver of rats of different ages. Biochemical and structural studies following perfusion with Cu-ceruloplasmin. Z Gerontol 17(2):69–72
Wan J, Benkdane M, Teixeira-Clerc F, Bonnafous S, Louvet A, Lafdil F, Pecker F, Tran A, Gual P, Mallat A, Lotersztajn S, Pavoine C (2014) M2 kupffer cells promote M1 kupffer cell apoptosis: a protective mechanism against alcoholic and nonalcoholic fatty liver disease. Hepatology 59(1):130–142. https://doi.org/10.1002/hep.26607
Weibel ER, Stäubli W, Gnägi HR, Hess FA (1969) Correlated morphometric and biochemical studies on the liver cell. J Cell Biol 42(1):68–91. https://doi.org/10.1083/jcb.42.1.68
Wong CK, Smith CA, Sakamoto K, Kaminski N, Koff JL, Goldstein DR (2017) Aging impairs alveolar macrophage phagocytosis and increases influenza-induced mortality in mice. J Immunol 199(3):1060–1068. https://doi.org/10.4049/jimmunol.1700397
Xie Q, Cho H, Calaycay J, Mumford R, Swiderek K, Lee T, Ding A, Troso T, Nathan C (1992) Cloning and characterization of inducible nitric oxide synthase from mouse macrophages. Science 256(5054):225–228. https://doi.org/10.1126/science.1373522
Yan BC, Gong C, Song J, Krausz T, Tretiakova M, Hyjek E, Al-Ahmadie H, Alves V, Xiao SY, Anders RA, Hart JA (2010) Arginase-1: a new immunohistochemical marker of hepatocytes and hepatocellular neoplasms. Am J Surg Pathol 34(8):1147–1154. https://doi.org/10.1097/PAS.0b013e3181e5dffa
Yu SF, Koerner TJ, Adams DO (1990) Gene regulation in macrophage activation: differential regulation of genes encoding for tumor necrosis factor, interleukin-1, JE, and KC by interferon-γ and lipopolysaccharide. J Leuk Biol 48(5):412–419. https://doi.org/10.1002/jlb.48.5.412
Zhang HJ, Xu L, Drake VJ, Oberley LW, Kregel KC (2003) Heat-induced liver injury in old rats is associated with exaggerated oxidative stress and altered transcription factor activation. FASEB J 17(15):2293–2295
Zhong S, Xu J, Li P, Tsukamoto H (2012) Caveosomal oxidative stress causes Src-p21ras activation and lysine 63 TRAF6 protein polyubiquitination in iron-induced M1 hepatic macrophage activation. J Biol Chem 287(38):32078–32084. https://doi.org/10.1074/jbc.M112.377358
Zhu F, Li X, Jiang Y, Zhu H, Zhang H, Zhang C, Zhao Y, Luo F (2015) GdCl3 suppresses the malignant potential of hepatocellular carcinoma by inhibiting the expression of CD206 in tumor-associated macrophages. Oncol Rep 34(5):2643–2655. https://doi.org/10.3892/or.2015.4268
Zwacka RM, Zhou W, Zhang Y, Darby CJ, Dudus L, Halldorson J, Oberley L, Englehardt JF (1998) Redox gene therapy for ischemia/reperfusion injury of the liver reduces AP1 and NF-kappaB activation. Nat Med 4(6):698–704
Acknowledgments
We thank Tom Moninger at the University of Iowa Central Microscopy Research Facility for assistance with confocal microscopy.
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SAB was supported by a Faculty Development Grant from Penn State Abington. EDM was supported by research funds from the Schreyer Honors College of the Pennsylvania State University, and an Erickson Summer Discovery Grant from the Office of Undergraduate Research at Penn State. This work was also supported by NIH grant AG12350 to KCK. No funding source had a role in study design, data collection, analysis or interpretation of the data, in the writing of the final report, or the decision to submit the article for publication.
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Bloomer, S.A., Moyer, E.D., Brown, K.E. et al. Aging results in accumulation of M1 and M2 hepatic macrophages and a differential response to gadolinium chloride. Histochem Cell Biol 153, 37–48 (2020). https://doi.org/10.1007/s00418-019-01827-y
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DOI: https://doi.org/10.1007/s00418-019-01827-y