Emine ATAKİŞİ, ALİ HAYDAR KIRMIZIGÜL, ONUR ATAKİŞİ, EBRU KARADAĞ SARI, METİN ÖĞÜN, ŞABAN MARAŞLI, Ayla ÖZCAN, Mahmut KARAPEHLİVAN, FATİH BÜYÜK, ÖZGÜR ÇELEBİ

Leptospirozlu sığırlarda plazma nitrik oksit (NO) ve Tümör Nekrozis Faktör-α (TNF-α) düzeyleri ile adenozin deaminaz (ADA), gama glutamil transferaz (GGT) aktiviteleri ve perifer kan lökositlerinde alfa naftil asetat esteraz (ANAE) yöntemiyle lenfosit oranlarının belirlenmesi

Yapılan bu çalışmada plazmada nitrik oksit (NO), Tümör Nekrozis Faktör-α (TNF-α) düzeyleri, adenozin deaminaz (ADA), gama glutamil transferaz (GGT) aktiviteleri ve alfa naftil asetat esteraz (ANAE) yöntemi ile lenfosit oranları belirlenerek, sığır leptospirozunun immunpatogenezinin ve biyokimyasal mekanizmalarının araştırılması amaçlanmıştır. Kars civarındaki köylerde yetiştirilen 20 baş leptospirozlu ve 20 baş sağlıklı danaların vena jugularislerinden heparinli tüplere kan örnekleri alındı ve histolojik analizler için frotiler alındıktan sonra, 3.000 rpm’de 15 dak. santrifüj edilerek plazmaları elde edildi. Plazmada NO düzeyi ile ADA aktivitesi kimyasal yöntemle, TNF-α düzeyi ile GGT aktivitesi ticari test kitleri kullanılarak kolorimetrik yöntemle analiz edildi. T/B lenfosit oranları ise perifer kan lökositlerinde ANAE yöntemiyle belirlendi. Leptospirozlu hayvanların plazma TNF-α ve NO düzeyleri kontrol grubuna göre sırasıyla P

Plasma nitric oxide (NO) and tumor necrosis factor-α (TNF-α) levels, adenosine deaminase (ADA), gamma glutamyl transferase (GGT) activities and to determine the rate of lymphocytes in the peripheral Blood leukocytes alpha naphthyl acetate esterase (ANAE) in cattle with leptospirosis

In this study, by determining in plasma the nitric oxide (NO), tumour necrosis factor-α (TNF-α) levels, adenosine deaminase (ADA), gamma glutamyl transferase (GGT) activities and percentage of T lymphocytes with alpha naphthyl acetate esterase method (ANAE), it was aimed to investigate the immunopathogenesis and biochemical mechanism of bovine leptospirosis. Blood samples of twenty healthy calves and twenty calves with leptospirosis which grown in the villages around Kars were collected into heparinised tubes from jugular vein. After taking the blood smear, blood samples were centrifuged at 3.000 rpm for 15 min. Plasma NO levels and ADA activities were determined with chemical method. TNF-α levels and GGT activities were determined with commercial kits. T/B lymphocyte ratios were determined at peripheral blood leukocytes with ANAE method. Plasma TNF-α and NO levels, ADA and GGT activities in animals with leptospirosis were found to be high P<0.001, P<0.05, P<0.05, P<0.05 respectively compared to healthy animals. ANAE positive T lymphocytes ratio were determined as 43%. As a result, TNF-α as a cytokine can be determined in early stage of infection and, accordingly, significantly increased levels of NO, ADA and GGT activities and, on the basis of the increased in the percentage of B lymphocytes, it is proposed to these parameters could be used for determining in explain the pathogenesis of the disease. It is tought that, it must be done more studies on biochemical mechanism of leptospirosis besides native infected cattle.

PDF

___

1. Lowanitchapat A, Payungporn S, Sereemaspun A, Ekpo P, Phulsuksombati D, Poovorawan Y, Chirathaworn C: Expression of TNF-α, TGF-β, IP- 10 and IL-10 mRNA in kidneys of hamsters infected with pathogenic Leptospira. Comp Immunol Microbiol Infect Dis, 33, 423-434, 2010.
2. Merien F, Baranton G, Perolat P: Invasion of vero cells and induction of apoptosis in macrophages by pathogenic Leptospira interrogans are correlated with virulence. Infect Immun, 65 (2): 729-738, 1997.
3. Baganha MF, Pego A, Lima MA, Gaspar EU, Pharma B: Serum and pleural adenosine deaminase correlation with lymphocytic populations. Chest, 97 (3): 605-610, 1990.
4. Henson SE, Nichols, TC Holers, VM Karp DR: The ectoenzyme γ- glutamyl transpeptidase regulates antiproliferative effects of S- nitrosoglutathione on human T and B lymphocytes. J Immunol, 163, 1845- 1852, 1999.
5. Garthwaite J: Glutamate, nitric oxide and cell-cell signalling in the nervous system. Trends Neurosci, 14 (2): 60-67, 1991.
6. Stuehr DJ: Mammalian nitric oxide syntheses. Biochim Biophy Acta, 1411, 217-230, 1999.
7. Tangpong J, Sompol P, Vore M, St Clair W, Butterfield DA, St Clair DK: Tumor necrosis factor alpha–mediated nitric oxide production enhances manganese superoxide dismutase nitration and mitochondrial dysfunction in primary neurons: An insight into the role of glial cells. Neuroscience, 151, 622-629, 2008.
8. Kontas T, Salmanoglu B: Tumour necrosis factor-α, adenosine deaminase and nitric oxide levels in cattle babesiosis before and after treatment. Bull Vet Inst Pulawy, 50 (4): 485-48, 2006.
9. Knowles DM, Hoffman T, Ferrarini M, Kunkel HG: The demonstration of acid alpha naphthyl acetate esterase activity in human lymphocytes usefulness as a T cell marker. Cell Immun, 35, 112-123, 1978.
10. Mueller J, Brun del Re G, Buerki H, Keller HU, Hess MW, Cottier H: Nonspesific acid esterase activity: A criterion for differentiation of T and B lymphocytes in mouse lymph nodes. Eur J Immunol, 5, 270-274, 1975.
11. Giusti G, Galanti B: Methods of Enzymatic Analysis, s. 315-323, Verlac Chemie, Weinheim, 1984.
12. Miranda KM, Espey MG, Wink DA: A rapid, simple spectrophotometric method for simultaneous detection of nitrate and nitrite. Nitric Oxide, 5 (1): 62-71, 2001.
13. SPSS: SPSS for windows release 16.0.2, SPSS Inc., Chicago, April, 2008.
14. Adler B, Moctezuma AP: Leptospira and Leptospirosis. Vet Microbiol, 140, 287-296, 2010.
15. Rodostits OM, Blood DC, Gay CC: Veterinary Medicine: A Text Book of the Diseases of Cattle, Horses, Sheep, Pigs and Goats. 8th ed., 884-908, Baillaire Tindall, London, 1994.
16. Erdogan HM, Karapehlivan M, Citil M, Atakisi O, Uzlu E, Unver A: Serum sialic acid and oxidative stres parameters changes in cattle with leptospirosis. Vet Res Com, 32, 333-339, 2008.
17. Quin PJ, Carter ME, Markey B, Carter GR: Clinical Veterinary Microbiology. 1st ed., 292-298, Wolfe Publishing, London, 1994.
18. Urbanik-Sypniewska T, Choma A, Kutkowska J, Kaminska T, Kandefer-Szerszen M, Russa R, Dolecka J: Cytokine inducing activities of rhizobial and mesorhizobial lipopolysaccharides of different lethal toxicity. Immunobiology, 202, 408-420, 2000.
19. Ergün E, Ergün L, Özen A, Aştı RN: Determination of alpha naphtyl acetate esterase activity in the peripheral blood leukocytes of ostrich (Struthio camelus masaicus). Revue Med Vet, 81, 147-150, 2004.
20. Marinho M, Silva C, Lima VMF, Machado GF, Peiro JR, Perri SHV: Production of TNF-α and IL-6, antibody response, and bacterial recovery, during leptospirosis infection. Vet Immunol Immunopathol, 128, 211-347, 2009.
21. Marinho M, Silva C, Lima VMF, Peiró JR, Perri SHV: Cytokine and antibody production during murine Leptospirosis. J Venom Anim Toxins incl Trop Dis, 12 (4): 595-603, 2006.
22. Elves AP Maciel, Daniel A Athanazio, Eliana AG Reis, Fernando Q Cunha, Adriano Queiroz, Deusdelia Almeida, Alan JA McBride, Albert I Ko, Mitermayer G Reis: High serum nitric oxide levels in patiens with severe leptospirosis. Acta Tropica, 100, 256-260, 2006.
23. Lalenti A, Moncada S, Di-Rosa M: Modulation of adjuvant arthritis by endogenous nitric oxide. Br J Pharmacol, 110, 701-706, 1993.
24. Stefanovic-Racic M, Meyers K, Meschter C, Coffey JW, Hoffman RA, Evans CH: N-Monomethyl arginine. An inhibitor of nitric oxide synthase. Suppresses the devolopment of adjuvant arthritis in rats. Arthritis Reum, 37 (7): 1062-1069, 1994.
25. Kajikawa DVM, Koyama H, Yushikawa T, Tsubaki S: Use of alpha- naphthyl acetate esterase staining to identify T lymphocytes in cattle. Am J Vet Res, 44, 1549-1552, 1983.
26. Fehr J, Scherer O: Nonspecific esterase as a marker for human T- lymphocytes sequential studies during states of transient blood lympho- cyte redistributions. Ann Hematol, 44, 201-109, 1981.
27. Kulenkampff J, Janossy G, Greaves JR: Acid esterase in human lymphoid cells and leukaemic blasts: A marker for T lymphocytes. Br J Haematol, 36, 231-240, 1977.
28. Gancheva G, Atanasova M, Lukanov T, Ilieva P: Flow cytometry in leptospirosis. JIMAB, 1, 3-97, 2008.
29. Tonin AA, Pimentel VC, Da Silva AS, De Azevedo MI, Souza VC, Wolkmer P, Rezer JF, Badke MR, Leal DB, Schetinger MR, Monteiro SG, Lopes ST: Adenosine deaminase activity in serum, erthrocytes and lymphocytes of rats infected with Leptospira icterohaemorrhagiae. Res Vet Sci, 92 (2): 197-201, 2012.