RESUMO
We investigated the gene expression of matrix metalloproteinases-9 (MMP-9) and tissue inhibitors of matrix metalloproteinases-1 (TIMP-1) in peripheral blood cells from infected cattle with Mycobacterium avium subsp. paratuberculosis (Map) in the ELISA-negative subclinical stage compared with uninfected control cattle. Significant decreased MMP-9 expression and increased TIMP-1 expression were found in peripheral blood cells from Map-infected cattle after stimulation with Map lysate and Map purified protein derivative (PPD) than in control cattle by real-time RT-PCR analysis. In contrast to the uninfected controls, the activity of MMP-9 was also decreased in peripheral blood cell culture supernatants from Map-infected cattle at 24 hr after Map lysate and MapPPD stimulation by gelatin zymography analysis. As a result, the MMP-9 may play an important role in the development of Mycobacterium avium subsp. paratuberculosis disease.
Assuntos
Doenças dos Bovinos/genética , Expressão Gênica , Metaloproteinase 9 da Matriz/genética , Mycobacterium avium subsp. paratuberculosis , Paratuberculose/genética , Inibidor Tecidual de Metaloproteinase-1/genética , Animais , BovinosRESUMO
Urocortin (UCN) is a new neuropeptide of the corticotrophin-releasing hormone (CRH) family which plays an important role in immune responses. Mycobacterium avium subspecies paratuberculosis (Map) is the etiological agent of paratuberculosis (Johne's disease). The role of UCN or CRH in the pathogenesis of Map-infection is unknown. In the present study, we first cloned the bovine UCN gene and demonstrated the profile of UCN or CRH expression in peripheral blood cells from Map-infected cattle and uninfected controls by real-time reverse transcription-polymerase chain reaction (RT-PCR) and ELISA analysis. These data are the first observations of the characteristic kinetics of these neuropeptides in Map-infection. UCN or CRH expression in non-stimulated blood samples from infected cattle was higher than that in similarly treated samples from uninfected controls; however, exposure to Map lysate and live Map resulted in down-regulated expression of UCN in infected cattle compared to their counterparts from uninfected controls. These results have provided a direction in understanding the pathogenesis of paratuberculosis and improving diagnostic methods for Map-infection.
Assuntos
Doenças dos Bovinos/sangue , Hormônio Liberador da Corticotropina/biossíntese , Leucócitos Mononucleares/metabolismo , Paratuberculose/sangue , Urocortinas/biossíntese , Sequência de Aminoácidos , Animais , Sequência de Bases , Bovinos , Doenças dos Bovinos/genética , Doenças dos Bovinos/microbiologia , Clonagem Molecular , Hormônio Liberador da Corticotropina/sangue , Hormônio Liberador da Corticotropina/genética , Ensaio de Imunoadsorção Enzimática/métodos , Interleucina-10/biossíntese , Interleucina-10/genética , Masculino , Dados de Sequência Molecular , Mycobacterium avium subsp. paratuberculosis/crescimento & desenvolvimento , Paratuberculose/genética , Paratuberculose/microbiologia , RNA Mensageiro/biossíntese , RNA Mensageiro/sangue , RNA Mensageiro/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Urocortinas/sangue , Urocortinas/genéticaRESUMO
Paratuberculosis (Ptb), caused by Mycobacterium avium subsp. paratuberculosis (Map), is a chronic enteritis that affects many ruminants and other wild animals worldwide. Ptb is a great concern in animal health and in etiology of human Crohn's disease (CD). In the present study, we detected Map-specific insertion sequence IS900 of DNA in tissue sections surgically removed from lesions of patients with CD (29 samples), ulcerative colitis (UC) (17 samples), and non-inflammatory bowel disease (IBD) (20 samples). We then compared the histopathological findings of 29 CD and 17 UC cases with those of 35 cases of bovine Ptb, since few comparative pathological studies of human IBD and Ptb have been conducted. The QPCR examination indicated positive results in 13.37% of CD cases, 3.57% of UC cases, and 10% of non-IBD cases. Human CD tissues typically exhibited destructive full thickness enteritis with severe lympho-plasma infiltration and scattered additional granulomas; UC lesions exhibited much less inflammation than CD lesions. Non-IBD control samples did not exhibit pathological changes. Human CD and UC lesions were very different from Ptb lesions that are characterized by predominant granuloma formation. Immunohistochemistry for Map antigen and acid-fast staining were negative in all human IBD cases but were always positive in Ptb cases. Our present comparative study strongly suggests that we reconsider the previous hypothesis that "Map infection" causes CD, even though human intestines were considered to have been exposed to the Map antigen containing the DNA.