OJVM  Vol.2 No.4 , December 2012
Assessment of Molecular Signalling Mechanisms for Eosinophilia in Rottweilers
Abstract: Rottweilers are predisposed to eosinophilic diseases, including hypereosinophilic syndrome. The immunopathogenesis of idiopathic eosinophilia is poorly characterised in dogs and man. Studies in people have suggested cytokines, particularly interleukin (IL)-5, play a role in instigating and perpetuating eosinophilia. This study sought to establish whether differences in gene expression, and concentration of selected, cytokines and chemokines were associated with eosinophilia in Rottweilers. Quantitative real-time reverse transcriptase polymerase chain reaction (qRT-PCR) assays were used to quantify messenger ribonucleic acid (mRNA) encoding cytokines IL-4, -5, -10, -12p19, -12p35, -12p40, -18, interferon gamma (IFN-γ) and chemokines eotaxin-2 and -3 from peripheral blood mononuclear cell (PMBC) samples obtained from healthy dogs (breeds other than Rottweiler) with normal eosinophil blood counts (n = 5) and Rottweilers with normal (n = 6), mildly increased (n = 7) and high (n = 3) eosinophil blood counts. Quantification of plasma IFN-γ and IL-5 was performed using commercially available canine-specific enzyme-linked immunosorbent assays ELISAs. Cytokine mRNA was measurable in all samples, although eotaxin-2 and -3 were not detected. No significant differences in gene expression of any cytokine were found between groups (based on eosinophil count or breed). No significant difference in plasma IL-5 or IFN-γ concentration was present between groups. In conclusion, there were no significant differences in cytokine mRNA profiles or plasma IL-5 and IFN-γ levels between Rottweilers with increased eosinophil counts and Rottweiler and non-Rottweiler dogs with normal eosinophil counts.
Cite this paper: N. Prakash, M. J. Day, P. Stumbles, I. R. Peters, M. Sharman, A. Paul and C. Mansfield, "Assessment of Molecular Signalling Mechanisms for Eosinophilia in Rottweilers," Open Journal of Veterinary Medicine, Vol. 2 No. 4, 2012, pp. 170-177. doi: 10.4236/ojvm.2012.24027.

[1]   I. R. Tizard, “Veterinary Immunology: An Introduction,” 7th Edition, Saunders, 2004.

[2]   F. Brito-Babapulle, “Review: The Eosinophilias, Including the Idiopathic Hypereosinophilic Syndrome,” British Journal of Haematology, Vol. 121, No. 2, 2003, pp. 203-223. doi:10.1046/j.1365-2141.2003.04195.x

[3]   I. LillieHook and H. Tvedten, “Investigation of Hypereosinophilia and Potential Treatments,” Veterinary Clinics of North America: Small Animal Practice, Vol. 33, No. 6, 2003, pp.1359-1378. doi:10.1016/S0195-5616(03)00097-4

[4]   I. LillieHook, L. Gunnarsson, G. Zakrisson and H. Tvedten, “Diseases Associated With Pronounced Eosinophilia: A Study of 105 Dogs in Sweden,” Journal of Small Animal Practice, Vol. 41, No. 6, 2000, pp. 248-253. doi:10.1111/j.1748-5827.2000.tb03934.x

[5]   F. Roufosse and P. F. Weller, “Practical Approach to the Patient with Hypereosinophilia,” Journal of Clinical Immunology, Vol. 126, No. 1, 2010, pp. 39-44. doi:10.1016/j.jaci.2010.04.011

[6]   C. Clercx, D. Peeters, F. Snaps, P. Hansen, K. McEntee, J. Detilleux, M. Henroteaux and M. J. Day, “Eosinophilic Bronchopneumopathy in Dogs,” Journal of Veterinary Internal Medicine, Vol. 14, No. 3, 2000, pp. 282-291. doi:10.1111/j.1939-1676.2000.tb01168.x

[7]   I. Aroch, S. Perl and A. Markovics, “Disseminated Eosinophilic Disease Resembling Idiopathic Hypereosinophilic Syndrome in a Dog,” The Veterinary Record, Vol. 149, No. 13, 2001, pp. 386-389. doi:10.1136/vr.149.13.386

[8]   M. C. Perkins and A. D. J. Watson, “Successful Treatment of Hypereosinophilic Syndrome in a Dog,” Australian Veterinary Journal, Vol. 79, No. 10, 2001, pp. 686-689. doi:10.1111/j.1751-0813.2001.tb10671.x

[9]   A. J. German, D. J. Holden, E. J. Hall and M. J. Day, “Eosinophilic Diseases in Two Cavelier King Charles Spaniels,” Journal of Small Animal Practice, Vol. 43, No. 12, 2002, pp. 533-538. doi:10.1111/j.1748-5827.2002.tb00026.x

[10]   G. D. Bellou, S. Kleinschmidt, F. Meneses, I. Nolte and M. Hewicker-Trautwein, “Eosinophilic Granulomatous Gastroenterocolitis and Hepatitis in a 1-Year-Old Male Siberian Husky,” Veterinary Pathology, Vol. 43, No. 6, 2006, pp. 1022-1025. doi:10.1354/vp.43-6-1022

[11]   F. E. James and C. S. Mansfield, “Clinical Remission of Idiopathic Hypereosinophilic Syndrome in a Rottweiler,” Australian Veterinary Journal, Vol. 87, No. 8, 2009, pp. 330-333. doi:10.1111/j.1751-0813.2009.00461.x

[12]   J. E. Sykes, D. J. Weiss, L. C. Buoen, M. M. Blauvelt, and D. W. Hayden, “Idiopathic Hypereosinophilic Syndrome in 3 Rottweilers,” Journal of Veterinary Internal Medicine, Vol. 15, No. 2, 2001, pp. 162-166. doi:10.1111/j.1939-1676.2001.tb01251.x

[13]   A. D. Klion, B. S. Bochner, G. J. Gleich, T. B. Nutman, M. E. Rothenberg, H. U. Simon, M. E. Wechsler and P. F. Weller: The Hypereosinophilic Syndromes Working Group, “Approaches to the Treatment of Hypereosinophilic Syndromes: A Workshop Summary Report,” Journal of Allergy and Clinical Immunology, Vol. 117, No. 6, 2009, pp. 1292-1301. doi:10.1016/j.jaci.2006.02.042

[14]   W. W. Busse, J. Ring, J. Huss-Marp and J. E. Kahn, “A review of treatment with mepolizumab, an anti-IL-5 mAb, in hypereosinophilic syndromes and asthma,” Journal of Allergy and Clinical Immunology, Vol. 125, No. 4, 2010, pp. 803-813. doi:10.1016/j.jaci.2009.11.048

[15]   J. Elsner, H. Petering, C. Kluthe, D. Kimmig, R. Smolarski, P. Ponath and A. Kap, “Eotaxin-2 Activates Chemotaxis-Related Events and Release of Reactive Oxygen Species via Pertussis Toxin-Sensitive G Proteins in Human Eosinophils,” European Journal of Immunology, Vol. 28, No. 7, 1998, pp. 2152-2158. doi:10.1002/(SICI)1521-4141(199807)28:07<2152::AID-IMMU2152>3.0.CO;2-G

[16]   A. Shinkai, H. Yoshisue, M. Koike, E. Shoji, S. Nakagawa, A. Saito, T. Takeda, S. Imabeppu, Y. Kato, N. Hanai, H. Amazawa, T. Kuga and T. Nishi, “A Novel Human CC Chemokine, Eotaxin-3, Which is Expressed in IL-4-Stimulated Vascular Endothelial Cells, Exhibits Potent Activity Toward Eosinophils,” Journal of Immunology, Vol. 163, No. 3, 1999, pp. 1602-1610.

[17]   N. Zimmermann, S. P. Hogan, A. Mishra, E. B. Brandt, T. R. Bodette, S. M. Pope, F. D. Finkelman and M. E. Rothenberg, “Murine Eotaxin-2: A Constitutive Eosinophil Chemokine Induced by Allergen Challenge and IL-4 Overexpression,” Journal of Immunology, Vol. 165, No. 10, 2000, pp. 5839-5846.

[18]   S. Rozen and H. Skaletsky, “Primer 3 on the www for General Users and for Biologist Programmers,” Methods in Molecular Biology, Vol. 132, 2000, pp. 365-386.

[19]   I. R. Peters, C. R. Helps, E. J. Hall and M. J. Day, “Real-time RT-PCR: Considerations for Efficient and Sensitive Assay Design,” Journal of Immunological Methods, Vol. 286, No. 1-2, 2004, pp. 203-217. doi:10.1016/j.jim.2004.01.003

[20]   I. R. Peters, C. R. Helps, E. L. Calvert, E. J. Hall and M. J. Day, “Cytokine mRNA Quantification in Histologically Normal Canine Duodenal Mucosa by Real-time RT- PCR,” Veterinary Immunology and Immunopathology, Vol. 103, No. 1-2, 2005, pp. 101-111. doi:10.1016/j.vetimm.2004.08.020

[21]   D. Peeters, I. R. Peters, F. Farnir, C. Clercx, and M. J. Day, “Real-Time RT-PCR Quantification of mRNA Encoding Cytokines and Chemokines in Histologically Normal Canine Nasal, Bronchial and Pulmonary Tissue,” Veterinary Immunology and Immunopathology, Vol. 104, No. 3-4, 2005, pp. 195-204. doi:10.1016/j.vetimm.2004.11.007

[22]   D. Peeters, I. R. Peters, C. Clerex and M. J. Day, “Real-Time RT-PCR Quantification of mRNA Encoding Cytokines, CC Chemokines and CCR3 in Bronchial Biopsies from Dogs with Eosinophilic Bronchopneumopathy,” Veterinary Immunology and Immunopathology, Vol. 110, No. 1-2, 2006, pp. 65-77. doi:10.1016/j.vetimm.2005.09.004

[23]   D. Peeters, I. R. Peters, C. R. Helps, A. Gabriel, M. J. Day and C. Clercx, “Distinct Tissue Cytokine and Chemokine mRNA Expression in Canine Sino-Nasal Aspergillosis and Idiopathic Lymphoplasmacytic Rhinitis,” Veterinary Immunology and Immunopathology, Vol. 117, No. 1-2, 2007, pp. 95-105. doi:10.1016/j.vetimm.2007.01.018

[24]   I. R. Peters, D. Peeters, C. R. Helps and M. J. Day, “Development and Application of Multiple Internal Reference (Housekeeper) Gene Assays for Accurate Normalisation of Canine Gene Expression Studies,” Veterinary Immunology and Immunopathology, Vol. 117, No. 1-2, 2007, pp. 55-66. doi:10.1016/j.vetimm.2007.01.011

[25]   J. Vandesompele, K. De Preter, F. Pattyn, B. Poppe, N. Van Roy, A. De Paepe and F. Speleman, “Accurate Normalization of Real-Time Quantitative RT-PCR Data by Geometric Averaging of Multiple Internal Control Genes,” Genome Biology, Vol. 3, No. 7, 2002, Article ID: RESEARCH0034.

[26]   A. Meager, “The Interferons: Characterization and Application,” Wiley-VCH, Verlag Gmbh and Co. KGaA, Weinheim, 2006.

[27]   A. D. Klion, M. A. Law, P. Noel, Y. J. Kim, T. P. Haverty and T. B. Nutman, “Safety and Efficacy of the Monoclonal Anti-Interleukin-5 Antibody SCH55700 in the Treatment of Patients with Hypereosinophilic Syndrome,” Blood, Vol. 103, No. 8, 2004, pp. 2939-2941. doi:10.1182/blood-2003-10-3620

[28]   A. N. Matthews, D. S. Friend, N. Zimmermann, M. N. Sarafi, A. D. Luster, E. Perlman, S. E. Wert and M. E. Rothenberg, “Eotaxin is Required for the Baseline Level of Tissue Eosinophils,” Proceedings of the National Academy of Sciences of the United States of America, Vol. 95, No. 11, 1998, pp. 6273-6278. doi:10.1073/pnas.95.11.6273

[29]   A. Mir, M. Minguez, J. Tatay, I. Pascual, A. Pena, V. Sanchiz, P. Almela, F. Mora and A. Benages, “Elevated Serum Eotaxin Levels in Patients with Inflammatory Bowel Disease,” American Journal of Gastroenterology, Vol. 97, No. 6, 2002, pp.1452-1457. doi:10.1111/j.1572-0241.2002.05687.x

[30]   A. Komiya, H. Nagase, H. Yamada, T. Sekiya, M. Yamaguchi, Y. Sano, N. Hanai, A. Furuya, K. Ohta, K. Matsushima, O. Yoshie, K. Yamamoto and K. Harai, “Concerted Expression of Eotaxin-1, Eotaxin-2, and Eotaxin-3 in Human Bronchial Epithelial Cells,” Cellular Immunology, Vol. 225, No. 2, 2003, pp. 91-100. doi:10.1016/j.cellimm.2003.10.001

[31]   T. Targowski, K. Jahnz-Rózyk, T. Plusa and E. Glodzinska-Wyszogrodzka, “Influence of Age and Gender on Serum Eotaxin Concentration in Healthy and Allergic People,” Journal of Investigational Allergology and Clinical Immunology, Vol. 15, No. 4, 2005, pp. 277-282.

[32]   K. Polzer, T. Karonitsch, T. Neumann, G. Eger, C. Haberler, A. Soleiman, B. Hellmich, E. Csernok, J. Distler, B. Manger, K. Redlich, G. Schett and J. Zwerina, “Eotaxin-3 is Involved in Churg-Strauss Syndrome—A Serum Marker Closely Correlating with Disease Activity,” Rheumatology, Vol. 47, No. 6, 2008, pp. 804-808. doi:10.1093/rheumatology/ken033

[33]   B. Vogel, A. Klinder, D. Sittig and G. Aust, “Bovine Eotaxin (CCL11)—An Unusual Member of the Eotaxin Group—Attracts Eosinophils in Vitro but is Not Responsible for Eosinophilia in the Ovary,” Veterinary Immunology and Immunopathology, Vol. 107, No. 1-2, 2005, pp. 67-77. doi:10.1016/j.vetimm.2005.03.013