Delayed apoptosis of buffalo neutrophil during mastitis and metritis
221 / 149
Authors
-
Chirag Uppal
Guru Angad Dev Veterinary and Animal Sciences University, Ludhiana
-
BV Sunil Kumar
Guru Angad Dev Veterinary and Animal Sciences University, Ludhiana
-
Dipak Deka
Guru Angad Dev Veterinary and Animal Sciences University, Ludhiana
-
Ramneek Verma
Guru Angad Dev Veterinary and Animal Sciences University, Ludhiana
-
R S Sethi
Guru Angad Dev Veterinary and Animal Sciences University, Ludhiana
http://orcid.org/0000-0003-0416-8946
Keywords:
Apoptosis, Buffalo, Caspase-3, Caspase-9, Neutrophils, ViabilityAbstract
Mastitis and metritis are the common inflammatory diseases of buffalo and result huge economic losses to the small-scale farmers. Neutrophils are recruited during inflammation and cleared from site of inflammation by apoptosis to prevent further tissue damage. However, data are lacking on this aspect in buffalo. Hence, the study was aimed to determine the viability and apoptosis of neutrophils of buffalo during mastitis and metritis. Peripheral blood samples were collected from healthy, mastitis and metritis group (n=5, each group). Neutrophils were isolated and maintained up to 48 hr in RPMI-1640. The viability of neutrophils at different time intervals (6, 12, 24, 36 and 48 hr) was determined using trypan blue dye exclusion method. DNA Laddering, light microscopy, caspase-3 and caspase-9 colorimetric assays were used to observe the apoptosis. Freshly isolated neutrophils showed ≥ 97% purity and ≥ 92% viability. Healthy group showed decrease (p<0.05) in viable neutrophils 12hr and onwards post-incubation. The number of apoptotic neutrophils was lower (p<0.05) at 36hr and onwards post-incubation in mastitis group and at 48hr post-incubation in metritis group as compared to control group. DNA fragmentation was observed after 24hr post-incubation in healthy group and was absent in mastitis and metritis groups suggesting the delayed apoptosis during mastitis and metritis. Caspase-3 and caspase-9 activity did not show any change in healthy group, however, there was significant increase at 24hr and 36hr post-incubation in mastitis and metritis group. Taken together, data suggest delayed apoptosis in mastitis and metritis.
References
Anwer AM, Asfour HAE, Gamal IM (2016) Apoptosis in somatic cells and immunological bioactive parameters of cow’s milk and their relation to subclinical mastitis. Alexandria J Vet Sci 49(2): 31-41
Appelberg R (2007) Neutrophils and intracellular pathogens: beyond phagocytosis and killing. Trends Microbiol 15(2): 87-92
Boutet P, Boulanger D, Gillet L, Vanderplasschen A, Closset R, Bureau F, Lekeux P (2004) Delayed neutrophil apoptosis in bovine subclinical mastitis. J Dairy Sci 87: 4104–4114.
Fadeel B, Ahlin A, Henter JI, Orrenius S, Hampton MB (1998) Involvement of Caspases in neutrophil apoptosis: regulation by reactive oxygen species. Blood 92(12): 4808-4818
Fadeel B, Orrenius S (2005) Apoptosis: a basic biological phenomenon with wide ranging implications in human disease. J Intern Med 258(6): 479-517
Garlichs CD, Eskafi S, Cicha I Schmeisser A, WalzogB, Raaz D, Stumpf C, Yilmaz A, Bremer J, Ludwig J, Daniel WG (2004) Delay of neutrophil apoptosis in acute coronary syndromes. J Leukoc Biol 75: 828-835
Kirschnek S, Vier J, Gautam S, Frankenberg T, Rangelova S, Eitz-Ferrer P, Grespi F, Ottina E, Villunger A, Hacker H, Hacker G (2011) Molecular analysis of neutrophil spontaneous apoptosis reveals a strong role for the pro-apoptotic BH3-only protein Noxa. Cell Death Differ 18: 1805–1814
Lee A, Whyte MK, Haslett C (1993) Inhibition of apoptosis and prolongation of neutrophil functional longevity by inflammatory mediators. J Leukoc Biol 54(4): 283-288
Liles WC, Kiener PA, Ledbetter JA, Aruffo A, Klebanoff SJ (1996) Differential expression of Fas (CD95) and Fas ligand on normal human phagocytosis: Implications for the regulations of apoptosis in neutrophils. J Exp Med 184: 429-440
Martin C, Burdon PC, Bridger G, Gutierrez-Ramos JC, Williams TJ, Rankin SM (2003) Chemokines acting via CXCR2 and CXCR4 control the release of neutrophils from the bone marrow and their return following senescence. Immunity 19(4): 583-593
Riedemann NC, Guo RF, Gao H, Sun L, Hoesel M, Hollmann TJ, Wetsel RA, Zetoune FS, Ward PA (2004) Regulatory role of C5a on macrophage migration inhibitory factor release from neutrophils. J Immunol 173:1355–59
Savill J, Fadok V (2000) Corpse clearance defines the meaning of cell death. Nature 407 (6805): 784-788
Singh U, Sethi RS, Mukhopadhyay CS, Sunil Kumar BV, Deka D, Verma R (2015) Endotoxin exposure alters viability and apoptotic potential of buffalo neutrophils. J Adv Res 3(12): 1636-1644
Sladek Z, Rysanek D (2000) Apoptosis of polymorphonuclear leukocytes of the juvenile bovine mammary glandduring induced influx. Vet Res 31: 553–563
Sladek Z, Rysanek D (2001) Apoptosis of neutrophilic granulocytes of bovine virgin mammary gland in scanning electron microscopy. Vet Med – Czech 46(7):185–189
Sladek Z, Rysanek D, Ryznarova H, Faldyna M (2006) The role of neutrophil apoptosis during experimentally induced Streptococcus uberis mastitis. Vet Med - Czech 51: 437–447
Swain DK, Kushwah MS, Kaur M, Patbandha TK, Mohanty AK, Dang AK (2014) Formation of NET, phagocytic activity, surface architecture, apoptosis and expression of toll like receptors 2 and 4 (TLR2 and TLR4) in neutrophils of mastitic cows. Vet Res Commun 38: 209–219
Taylor RC, Cullen SP, Martin SJ (2008) Apoptosis: controlled demolition at the cellular level. Nat Rev Mol Cell Biol 9(1): 231-241
Vermeulen K, Bockstaele DRV, Berneman ZN (2005) Apoptosis: mechanisms and relevance in cancer. Ann Hematol 84(10): 627-639
