Cardiac hypertrophy is a common response to injury and hemodynamic stress and an important harbinger of heart failure and death. Herein, we identify the Kruppel-like factor 15 (KLF15) as an inhibitor of cardiac hypertrophy. Myocardial expression of KLF15 is reduced in rodent models of hypertrophy and in biopsy samples from patients with pressure-overload induced by chronic valvular aortic stenosis. Overexpression of KLF15 in neonatal rat ventricular cardiomyocytes inhibits cell size, protein synthesis and hypertrophic gene expression. KLF15-null mice are viable but, in response to pressure overload, develop an eccentric form of cardiac hypertrophy characterized by increased heart weight, exaggerated expression of hypertrophic genes, left ventricular cavity dilatation with increased myocyte size, and reduced left ventricular systolic function. Mechanistically, a combination of promoter analyses and gel-shift studies suggest that KLF15 can inhibit GATA4 and myocyte enhancer factor 2 function. These studies identify KLF15 as part of a heretofore unrecognized pathway regulating the cardiac response to hemodynamic stress.

Mammalian cell invasion by the protozoan parasite, Trypanosoma cruzi, is facilitated by the activation of host cell phosphatidylinositol 3 (PI 3)-kinases. We demonstrate that the well-characterized Ca2+-regulated lysosome-mediated parasite entry pathway is abolished by wortmannin pretreatment. In addition, we have characterized a novel route of T. cruzi invasion unexpectedly revealed in the course of this study. For over a decade, targeted exocytosis of lysosomes at the host cell plasma membrane was considered as the primary mechanism for T. cruzi entry into non-professional phagocytic cells. We now provide evidence that a significant fraction (50% or greater) of invading T. cruzi trypomastigotes exploit an alternate actin-independent entry pathway that involves formation of a tightly associated host cell plasma membrane-derived vacuole enriched in the lipid products of class I PI 3-kinases, PtdInsP3/PtdIns(3,4)P2. Initially devoid of lysosomal markers, the resultant parasite-containing vacuoles gradually acquire lysosome associated membrane protein 1 (lamp-1) and fluid phase endocytic tracer from the lysosomal compartment. In striking contrast to latex bead phagosomes, few T. cruzi vacuoles associate with the early endosomal marker, EEA1 and the 'maturation' process becomes refractory to PI 3-kinase inhibition immediately following parasite internalization. Jointly, these data provide a new paradigm for T. cruzi invasion of non-professional phagocytic cells and reveal a novel vacuole maturation process that appears to bypass the requirement for EEA1.

SUMMARYOver 77 million dogs and 93 million cats share our households in the United States. Multiple studies have demonstrated the importance of pets in their owners' physical and mental health. Given the large number of companion animals in the United States and the proximity and bond of these animals with their owners, understanding and preventing the diseases that these companions bring with them are of paramount importance. Zoonotic protozoal parasites, including toxoplasmosis, Chagas' disease, babesiosis, giardiasis, and leishmaniasis, can cause insidious infections, with asymptomatic animals being capable of transmitting disease.GiardiaandToxoplasma gondii, endemic to the United States, have high prevalences in companion animals.LeishmaniaandTrypanosoma cruziare found regionally within the United States. These diseases have lower prevalences but are significant sources of human disease globally and are expanding their companion animal distribution. Thankfully, healthy individuals in the United States are protected by intact immune systems and bolstered by good nutrition, sanitation, and hygiene. Immunocompromised individuals, including the growing number of obese and/or diabetic people, are at a much higher risk of developing zoonoses. Awareness of these often neglected diseases in all health communities is important for protecting pets and owners. To provide this awareness, this review is focused on zoonotic protozoal mechanisms of virulence, epidemiology, and the transmission of pathogens of consequence to pet owners in the United States.

Control of Leishmania infantum infection is dependent upon Th1 CD4+ T cells to promote macrophage intracellular clearance of parasites. Deficient CD4+ T cell effector responses during clinical visceral leishmaniasis (VL) are associated with elevated production of IL-10. In the primary domestic reservoir of VL, dogs, we define occurrence of both CD4+ and CD8+ T cell exhaustion as a significant stepwise loss of antigen-specific proliferation and IFNγ production, corresponding to increasing VL symptomatology. Exhaustion was associated with a fourfold increase in the population of T cells with surface expression of Programmed Death 1 (PD-1) between control and symptomatic populations. Importantly, exhausted populations of CD8+ T cells and to a lesser extent CD4+ T cells were present prior to onset of clinical VL. VL exhausted T cells did not undergo significant apoptosis ex vivo after antigen stimulation. Antibody block of PD-1 ligand, B7.H1, promoted return of CD4+ and CD8+ T cell function and dramatically increased reactive oxygen species production in co-cultured monocyte-derived phagocytes. As a result, these phagocytes had decreased parasite load. We demonstrate for the first time that pan-T cell, PD-1-mediated, exhaustion during VL influenced macrophage reactive oxygen intermediate production. Blockade of the PD-1 pathway improved the ability of phagocytes isolated from dogs presenting with clinical VL to clear intracellular parasites. T cell exhaustion during symptomatic canine leishmaniasis has implications for the response to vaccination and therapeutic strategies for control of Leishmania infantum in this important reservoir species.

BackgroundDogs are the predominant domestic reservoir for human L. infantum infection. Zoonotic visceral leishmaniasis (ZVL) is an emerging problem in some U.S. dog breeds, with an annual quantitative PCR prevalence of greater than 20% within an at-risk Foxhound population. Although classically Leishmania is transmitted by infected sand flies and phlebotomine sand flies exist in the United States, means of ongoing L. infantum transmission in U.S. dogs is currently unknown. Possibilities include vertical (transplacental/transmammary) and horizontal/venereal transmission. Several reports have indicated that endemic ZVL may be transmitted vertically.AimsOur aims for this present study were to establish whether vertical/transplacental transmission was occurring in this population of Leishmania-infected US dogs and determine the effect that this means of transmission has on immune recognition of Leishmania.MethodologyA pregnant L. infantum-infected dam donated to Iowa State University gave birth in-house to 12 pups. Eight pups humanely euthanized at the time of birth and four pups and the dam humanely euthanized three months post-partum were studied via L. infantum-kinetoplast specific quantitative PCR (kqPCR), gross and histopathological assessment and CD4+ T cell proliferation assay.Key ResultsThis novel report describes disseminated L. infantum parasites as identified by kqPCR in 8 day old pups born to a naturally-infected, seropositive U.S. dog with no travel history. This is the first report of vertical transmission of L. infantum in naturally-infected dogs in North America, emphasizing that this novel means of transmission could possibly sustain infection within populations.Major ConclusionsEvidence that vertical transmission of ZVL may be a driving force for ongoing disease in an otherwise non-endemic region has significant implications on current control strategies for ZVL, as at present parasite elimination efforts in endemic areas are largely focused on vector-borne transmission between canines and people. Determining frequency of vertical transmission and incorporating canine sterilization with vector control may have a more significant impact on ZVL transmission to people in endemic areas than current control efforts.

In both dogs and humans Leishmania infantum infection is more prevalent than disease, as infection often does not equate with clinical disease. Previous studies additively indicate that advanced clinical visceral leishmaniasis is characterized by increased production of anti-Leishmania antibodies, Leishmania-specific lymphoproliferative unresponsiveness, and decreased production of gamma interferon (IFN-␥) with a concomitant increase of interleukin-10 (IL-10). In order to differentiate infection versus progressive disease for better disease prognostication, we temporally evaluated humoral and cellular immunologic parameters of naturally infected dogs. The work presented here describes for the first time the temporal immune response to natural autochthonous L. infantum infection in foxhounds within the United States. Several key changes in immunological parameters should be considered when differentiating infection versus clinical disease, including a dramatic rise in IgG production, progressive increases in antigen-specific peripheral blood mononuclear cell proliferation, and IFN-␥ production. Polysymptomatic disease is precluded by increased IL-10 production and consistent detection of parasite kinetoplast DNA in whole blood. This clinical presentation and the immuno-dysregulation mirror those observed in human patients, indicating that this animal model will be very useful for testing immunomodulatory anti-IL-10 and other therapies.

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