Abstract
C-reactive protein (CRP), haptoglobin (Hp) and fibrinogen (Fbgn) are acute phase reactants (APRs), the blood levels of which increase during acute inflammation. However, although the levels of these APRs are used to monitor inflammation in man, their usefulness and sensitivity as markers of inflammation in rodents are less clear. We therefore wished to evaluate, in a comparative fashion, a prototype immunoassay for serum CRP, a commercial assay for serum Hp, and an automated assay for Fbgn, using a model of acute inflammation in the rat. Additionally, pro-inflammatory cytokines and serum protein fractions were also measured. The model of inflammation used was the intraperitoneal injection of Freund's complete adjuvant (FCA). In a concluding experiment, findings with Hp in the FCA rat model were validated in a toxicologically relevant study involving the induction of acute hepatic inflammation using the model hepatotoxicant carbon tetrachloride (CCl4). Female Wistar Han rats were treated with a single injection of FCA in a dose–response study (1.25–10.0 ml/kg, sampling at 36 h) and two time-course studies (over 40 h and 21 days). In a final experiment, rats were dosed with CCl4 at 0.8 ml/kg and sampled over a 17-day period. In FCA and CCl4 experiments, serum/plasma was prepared and tissues taken at autopsy for histological assessment (CCl4 study only). In the dose–response study, serum CRP, Hp and plasma Fbgn were increased at all FCA dose levels at 36 h post-dosing. Serum α2 and β1 globulin fractions were also increased, while albumin levels were decreased. In the 40-h time-course study, CRP levels peaked at 25–40 h post-dosing, to approximately 120% of control (as 100%). Hp levels increased to a maximum at 25 and 40 h post-dosing with values greater than 400% of control, and α2 and β1 globulin fractions peaked at 30 and 40 h post-dosing to 221 and 187% of control, respectively. Increased serum interleukin-6 (IL-6) and interleukin-1β (IL-1β) levels peaked at 20 h (11-fold) and 25 h (19-fold), respectively. In a 21-day time-course study, no increased CRP levels were measured despite elevated levels of Hp, which peaked at 36 h (approximately 7-fold above control), and remained elevated up to 21 days. IL-6 and IL-1β levels peaked at 12 h (19-fold) and 24 h (28-fold), respectively. Liver histopathology of animals treated with CCl4 showed centrilobular hepatocellular degeneration and necrosis (most significant at 36 h) with an inflammatory response (most significant at 48 h). Resolution of the lesion was complete by 4 days post-dosing. Serum alanine aminotransferase, aspartate aminotransferase and glutamate dehydrogenase levels peaked at 36 h post-dosing. Hp levels increased maximally at 48 h (426% of control). We conclude that serum CRP is a poor marker of acute inflammation in the rat in comparison with serum Hp and plasma Fbgn. Between Hp and Fbgn, serum Hp is shown to be the most sensitive and useful marker of acute inflammation.
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References
Baumann H, Gauldie J (1994) The acute phase response. Immunol Today 15:74–80
Baumann H, Morella KK, Jahreis GP, Marinkovic S (1990) Distinct regulation of the interleukin-1 and interleukin-6 response elements of the rat haptoglobin gene in rat and human hepatoma cells. Mol Cell Biol 10:5967–5976
Bürger W, Schade R, Hirschelmann R (1987) The rat C-reactive protein — isolation and response to experimental inflammation and tissue damage. AgentsActions 21:93–97
Bürger W, Ewald C, Fenner, E-M (1998) Increase in C-reactive protein in the serum of piglets (pCRP) following ACTH or corticosteroid administration. J Vet Med 45:1–6
Clauss A (1957) Rapid physiological coagulation method for the determination of fibrinogen. Acta Haematol 17:237–246
Connolly KM, Stecher VJ, Kent L (1988) Examination of interleukin-1 activity, the acute phase response, and leukocyte subpopulations in rats with adjuvant-induced arthritis. J Lab Clin Med 111:341–347
Crockson RA, Payne CJ, Ratcliff AP, Soothill JF (1966) Time sequence of acute phase reactive proteins following surgical trauma. Clin Chim Acta 14:435–441
DeForge LE, Takeuchi E, Nguyen DT, Remick DG (1994) Immunological priming attenuates the in vivo pathophysiological response to lipopolysaccharide. Am J Pathol 144:599–611
Ebersole JL, Machen RL, Steffen MJ, Willmann DE (1997) Systemic acute-phase reactants, C-reactive protein and haptoglobin in adult periodontitis. Clin Exp Immunol 107:347–352
Fouad FM, Mamer OA, Shahidi F (1996) Acute-phase response in rat to carbon tetrachloride-azathioprine induced cirrhosis and partial hepatectomy of cirrhotic liver. J Toxicol Environ Health 47:601–615
Gardner SY, Lehmann JR, Costa DL (2000) Oil fly ash-induced elevation of plasma fibrinogen in rats. Toxicol Sci 56:175–180
Geisterfer M, Gauldie J (1996) Regulation of signal transducer, GP130 and the LIF receptor in acute inflammation in vivo. Cytokine 8:283–287
Giffen PS, Andrews CM, Barret P, Clarke CJ, Fung K-W, Munday M, Roman IF, Smyth R, Turton J, Walshe K, York MJ (2002) Markers of experimental acute inflammation in the Wistar Han rat, with particular reference to haptoglobin and C-reactive protein. Comp Clin Path 11:186–187
Gilbertsen RB (1986) Rat haptoglobin: method of quantification and response to antiarthritic therapy in collagen arthritis. Immunopharmacology 11:69–77
Heegaard PMH, Klausen J, Nielsen JP, González-Ramón N, Piñeiro M, Lampreave F, Alava MA (1998) The porcine acute phase response to infection with Actinobacillus pleuropneumoniae. Haptoglobin, C-reactive protein, major acute phase protein and serum amyloid A protein are sensitive indicators of infection. Comp Biochem Physiol 119B:365–373
Jinbo T, Sakamoto T, Yamamoto S (2002) Serum α2-macroglobulin and cytokine measurements in an acute inflammation model in rats. Lab Anim 36:153–157
Kerr R, Stirling D, Ludlam CA (2001) Interleukin 6 and haemostasis. Br J Haematol 115:3–12
Koj A (1970) Acute-phase reactants and lysosomal enzymes in the blood of rats with experimental inflammation or radiation injury. Folia Biol 18:275–286
Koj A (1985) Biological functions of acute-phase proteins. In: Gordon AH, Koj A (eds) The acute-phase response to injury and infection. Elsevier, London, pp 145–159
Kushner I (1982) The phenomenon of the acute phase response. Ann NY Acad Sci 389:39–48
Larsson A, Bjork J, Lundberg C (1997) Nephelometric determination of rat fibrinogen as a marker of inflammatory response. Vet Immunol Immunopathol 59:163–169
Liao WSL, Jefferson LS, Taylor JM (1986) Changes in plasma albumin concentration, synthetic rate, and mRNA level during acute inflammation. Am J Physiol 251:C928–C934
Luster MI, Simeonova PP, Galluci RM, Matheson JM, Yucesoy B (2000) Immunotoxicology: role of inflammation in chemical-induced hepatotoxicity. Int J Immunopharmacol 22:1143–1147
Myers MA, Fleck A (1988) Observations on the delay in onset of the acute phase protein response. Br J Exp Pathol 69:169–176
Olivier E, Soury E, Risler J-L, Smith F, Schneider K, Lochner K, Jonzeau J-Y, Fey GH, Salier J-P (1999) A novel set of hepatic mRNAs preferentially expressed during an acute inflammation in the rat represents mostly intracellular proteins. Genomics 57:352–364
Simeonova PP, Gallucci RM, Hulderman T, Wilson R, Kommineni C, Rao M, Luster MI (2001) The role of tumor necrosis factor-α in liver toxicity, inflammation and fibrosis induced by carbon tetrachloride. Toxicol Appl Pharmacol 177:112–120
Steel DM, Whitehead AS (1994) The major acute phase reactants: C-reactive protein, serum amyloid P component and serum amyloid A protein. Immunol Today 15:81–88
Sternberg M, Peyroux J, Engler R, Grochulski A, Lesaget C, Lagrue G, Jayle MF (1974) Orosomucoid, seromucoid and haptoglobin in serum during adjuvant arthritis of the rat. Experientia 30:193–195
Tanahashi M, Koike J, Kawabe N, Nakadate-Matsushita T (1998) Inhibitory effect of TRK-530 on inflammatory cytokines in bone marrow of rats with adjuvant arthritis. Pharmacology 56:237–241
Volkova ZI (1976) The zonal distribution of the haptoglobin and ceruloplasmin synthesis in liver tissue. Vopr Med Khim 22:171–176
Whicher JT, Westacott CI (1992) The acute-phase response. In: Whicher JT, Evans SW (eds) Biochemistry of inflammation. Kluwer, London, pp 243–269
Whicher JT, Banks RE, Thompson D, Evans SW (1993) The measurement of acute-phase proteins as disease markers. In: Baumann H, Kushner I, Mackiewicz A (eds) Acute phase proteins, molecular biology, biochemistry and clinical applications. CRC Press, London, pp 633–650
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The authors would like to acknowledge the assistance of the technical staff at the School of Pharmacy for the care of the animals, and the co-operation and technical assistance supplied by the Department of Pre-Clinical Safety Sciences at GlaxoSmithKline, UK.
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Giffen, P.S., Turton, J., Andrews, C.M. et al. Markers of experimental acute inflammation in the Wistar Han rat with particular reference to haptoglobin and C-reactive protein. Arch Toxicol 77, 392–402 (2003). https://doi.org/10.1007/s00204-003-0458-7
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DOI: https://doi.org/10.1007/s00204-003-0458-7