Abstract
Veterinaria México OA ISSN: 2448-6760Cite this as:Landeros Sánchez B, Gutierrez Pabello JA, Medina Basulto GE, Renteria Evangelista TB, Díaz Aparicio E, Oshima S. Mycobacterium avium subsp. paratuberculosis down-regulates mRNA expression of iron-induced macrophage Ferroportin 1. Veterinaria México OA. 2016;3(1). doi: 10.21753/vmoa.3.1.361Mycobacterium avium subsp. paratuberculosis (MAP) is the causative agent of Johne’s disease. The mechanisms by which MAP is able to adapt to the innate host response are still unclear. We examined Ferroportin 1 (FPN1) mRNA expression levels via real-time PCR of the mouse macrophage cell line J774 that was incubated in the presence of Mycobacterium avium subsp. paratuberculosis (MAP) or MAP crude protein extract. Infection with live MAP decreased FPN1 mRNA levels in a multiplicity of infection (MOI)-dependent fashion. Macrophages infected with MOIs of 20:1 and 15:1 did not show any change in FPN1 gene expression, whereas MOIs of 10:1 and 5:1 induced a decrease of 50 and 80%, respectively. Macrophages treated with 50, 100, 150 and 200 µg/mL of MAP crude extract (ATCC19698) decreased FPN1 mRNA expression by 25%. Additionally, up-regulation of FPN1 mRNA by an iron overload treatment of 400 µM of ferric nitrilotriacetate (FeNTA) was abrogated by live MAP (MOI 20:1) by approximately 70%. Our data revealed an inhibitory effect of MAP on FPN1 mRNA and suggested a bacterial mechanism that may play a role in host iron regulation.Figure 1. Infection by Mycobacterium avium subsp. paratuberculosis (MAP) down-regulates macrophage fpn1 gene expression. Murine macrophages, J774, were infected with MAP at different multiplicities of infection (MOIs) for 6 hours. Total RNA was extracted and gene expression was measured by real-time PCR. Results were normalized to GAPDH and expressed as units of relative expression (URE). Results are the mean ± standard deviation of 3 independent experiments with 3 replicates each. Statistical significance, P = 0.05.
Highlights
El Mycobacterium avium subesp. paratuberculosis disminuye la regulación de la expresión del ARNm de la ferroportina 1 en los macrófagos inducidos con hierro
El regulador transcripcional de map, el gen ldeR, reconoce secuencias consenso en regiones llamadas “cajas de hierro”, localizadas en las regiones del promotor de genes encargadas de regular la expresión de genes implicados en la adquisición y almacenamiento de hierro (Janagama et al, 2009, 2010)
Transcriptional analysis of diverse strains Mycobacterium avium subspecies paratuberculosis in primary bovine monocyte derived macrophages
Summary
A través de la pcr en tiempo real, se examinaron los niveles de expresión del arn de la ferroportina 1 (fpn1) en células de macrófagos de ratón J774 infectados con Mycobacterium avium subsp. Se infectaron los macrófagos con map vivo, con o sin hierro y con el extracto de proteína cruda de map, lo que permitió identificar la inhibición de la expresión del arnm de fpn[1] como efecto directo de map en macrófagos tratados con hierro. Para determinar el efecto de map en la infección de macrófagos con una sobrecarga de hierro, los macrófagos se infectaron con una moi 20:1 o se les cultivó con 200 μg/mL del extracto de proteína cruda de map y 400 μM de fenta (fenta- map). Los resultados de la expresión del gen fpn[1] de los macrófagos son el promedio de tres experimentos independientes, cada uno con tres repeticiones
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