Measurement of serum Interleukin 34 (IL-34) and correlation with severity and pruritus scores in client-owned dogs with atopic dermatitis.

BACKGROUND: Atopic dermatitis (AD) is a common inflammatory skin disease of dogs. Interleukin (IL)-34 is a monocyte/macrophage growth factor, produced mainly by keratinocytes, that has been implicated in several human inflammatory conditions including human AD. HYPOTHESIS: Canine serum IL-34 concentrations are increased in dogs with AD and correlate with clinical lesion and pruritus scores. ANIMALS: Forty seven client-owned dogs diagnosed with AD and 25 healthy, unaffected control dogs. METHODS AND MATERIALS: A commercially available IL-34 ELISA was optimized for the measurement of IL-34 in canine serum samples. Information regarding treatment, clinical lesion scores [Canine Atopic Dermatitis Extent and Severity Index, 4th iteration (CADESI-04)] and pruritus Visual Analog Score (pVAS) were recorded for each dog at the time of serum collection. RESULTS: Dogs with AD had significantly increased serum IL-34 concentrations compared to controls. There was a significant positive correlation between IL-34 concentrations and CADESI-04 and pVAS scores. Concentrations of IL-34 remained increased in dogs with AD receiving steroids or the JAK1 inhibitor, oclacitinib, compared to unaffected control dogs. CONCLUSIONS AND CLINICAL IMPORTANCE: Serum IL-34 concentrations are increased in dogs with AD and are correlated with clinical severity and pruritus. IL-34 may be a suitable candidate therapeutic target for canine AD.

CSF1 Restores Innate Immunity After Liver Injury in Mice and Serum Levels Indicate Outcomes of Patients With Acute Liver Failure.

BACKGROUND & AIMS: Liver regeneration requires functional liver macrophages, which provide an immune barrier that is compromised after liver injury. The numbers of liver macrophages are controlled by macrophage colony-stimulating factor (CSF1). We examined the prognostic significance of the serum level of CSF1 in patients with acute liver injury and studied its effects in mice. METHODS: We measured levels of CSF1 in serum samples collected from 55 patients who underwent partial hepatectomy at the Royal Infirmary Edinburgh between December 2012 and October 2013, as well as from 78 patients with acetaminophen-induced acute liver failure admitted to the Royal Infirmary Edinburgh or the University of Kansas Medical Centre. We studied the effects of increased levels of CSF1 in uninjured mice that express wild-type CSF1 receptor or a constitutive or inducible CSF1-receptor reporter, as well as in chemokine receptor 2 (Ccr2)-/- mice; we performed fate-tracing experiments using bone marrow chimeras. We administered CSF1-Fc (fragment, crystallizable) to mice after partial hepatectomy and acetaminophen intoxication, and measured regenerative parameters and innate immunity by clearance of fluorescent microbeads and bacterial particles. RESULTS: Serum levels of CSF1 increased in patients undergoing liver surgery in proportion to the extent of liver resected. In patients with acetaminophen-induced acute liver failure, a low serum level of CSF1 was associated with increased mortality. In mice, administration of CSF1-Fc promoted hepatic macrophage accumulation via proliferation of resident macrophages and recruitment of monocytes. CSF1-Fc also promoted transdifferentiation of infiltrating monocytes into cells with a hepatic macrophage phenotype. CSF1-Fc increased innate immunity in mice after partial hepatectomy or acetaminophen-induced injury, with resident hepatic macrophage as the main effector cells. CONCLUSIONS: Serum CSF1 appears to be a prognostic marker for patients with acute liver injury. CSF1 might be developed as a therapeutic agent to restore innate immune function after liver injury.

Characterisation of a novel Fc conjugate of macrophage colony-stimulating factor.

We have produced an Fc conjugate of colony-stimulating factor (CSF) 1 with an improved circulating half-life. CSF1-Fc retained its macrophage growth-promoting activity, and did not induce proinflammatory cytokines in vitro. Treatment with CSF1-Fc did not produce adverse effects in mice or pigs. The impact of CSF1-Fc was examined using the Csf1r-enhanced green fluorescent protein (EGFP) reporter gene in MacGreen mice. Administration of CSF1-Fc to mice drove extensive infiltration of all tissues by Csf1r-EGFP positive macrophages. The main consequence was hepatosplenomegaly, associated with proliferation of hepatocytes. Expression profiles of the liver indicated that infiltrating macrophages produced candidate mediators of hepatocyte proliferation including urokinase, tumor necrosis factor, and interleukin 6. CSF1-Fc also promoted osteoclastogenesis and produced pleiotropic effects on other organ systems, notably the testis, where CSF1-dependent macrophages have been implicated in homeostasis. However, it did not affect other putative CSF1 targets, notably intestine, where Paneth cell numbers and villus architecture were unchanged. CSF1 has therapeutic potential in regenerative medicine in multiple organs. We suggest that the CSF1-Fc conjugate retains this potential, and may permit daily delivery by injection rather than continuous infusion required for the core molecule.

Cloning and expression of feline colony stimulating factor receptor (CSF-1R) and analysis of the species specificity of stimulation by colony stimulating factor-1 (CSF-1) and interleukin-34 (IL-34).

Colony stimulating factor (CSF-1) and its receptor, CSF-1R, have been previously well studied in humans and rodents to dissect the role they play in development of cells of the mononuclear phagocyte system. A second ligand for the CSF-1R, IL-34 has been described in several species. In this study, we have cloned and expressed the feline CSF-1R and examined the responsiveness to CSF-1 and IL-34 from a range of species. The results indicate that pig and human CSF-1 and human IL-34 are equally effective in cats, where both mouse CSF-1 and IL-34 are significantly less active. Recombinant human CSF-1 can be used to generate populations of feline bone marrow and monocyte derived macrophages that can be used to further dissect macrophage-specific gene expression in this species, and to compare it to data derived from mouse, human and pig. These results set the scene for therapeutic use of CSF-1 and IL-34 in cats.

Cloning and expression of porcine Colony Stimulating Factor-1 (CSF-1) and Colony Stimulating Factor-1 Receptor (CSF-1R) and analysis of the species specificity of stimulation by CSF-1 and Interleukin 34.

Macrophage Colony Stimulating Factor (CSF-1) controls the survival, differentiation and proliferation of cells of the mononuclear phagocyte system. A second ligand for the CSF-1R, Interleukin 34 (IL-34), has been described, but its physiological role is not yet known. The domestic pig provides an alternative to traditional rodent models for evaluating potential therapeutic applications of CSF-1R agonists and antagonists. To enable such studies, we cloned and expressed active pig CSF-1. To provide a bioassay, pig CSF-1R was expressed in the factor-dependent Ba/F3 cell line. On this transfected cell line, recombinant porcine CSF-1 and human CSF-1 had identical activity. Mouse CSF-1 does not interact with the human CSF-1 receptor but was active on pig. By contrast, porcine CSF-1 was active on mouse, human, cat and dog cells. IL-34 was previously shown to be species-specific, with mouse and human proteins demonstrating limited cross-species activity. The pig CSF-1R was equally responsive to both mouse and human IL-34. Based upon the published crystal structures of CSF-1/CSF-1R and IL34/CSF-1R complexes, we discuss the molecular basis for the species specificity.

Prevalence of potentially pathogenic enteric organisms in clinically healthy kittens in the UK.

Faecal samples were collected from 57 clinically healthy kittens presented for initial vaccination, in the UK. Routine bacteriological examination identified Salmonella species in one and Campylobacter species in five samples. Polymerase chain reaction (PCR) detected the presence of Campylobacter species in a further four samples. Routine parasitological examination revealed Toxocara species ova in nine (including four kittens stated to have been administered an anthelmintic) and Isospora species in four samples. No Giardia or Cryptosporidium species were detected by routine methods. A Giardia species enzyme-linked immunosorbent assay (ELISA) test kit designed for use in cats was positive in three kittens. A similar test kit designed for use in humans was negative in all samples and produced negative results even when known positive samples were tested. Potentially pathogenic enteric organisms were detected in 19 kittens by routine methods and 26 (prevalence 45%) by all methods. The high prevalence in asymptomatic kittens highlights the possibility that the detection of these organisms in kittens with gastrointestinal disease may be an incidental finding.

Pleiotropic effects of extended blockade of CSF1R signaling in adult mice.

We investigated the role of CSF1R signaling in adult mice using prolonged treatment with anti-CSF1R antibody. Mutation of the CSF1 gene in the op/op mouse produces numerous developmental abnormalities. Mutation of the CSF1R has an even more penetrant phenotype, including perinatal lethality, because of the existence of a second ligand, IL-34. These effects on development provide limited insight into functions of CSF1R signaling in adult homeostasis. The carcass weight and weight of several organs (spleen, kidney, and liver) were reduced in the treated mice, but overall body weight gain was increased. Despite the complete loss of Kupffer cells, there was no effect on liver gene expression. The treatment ablated OCL, increased bone density and trabecular volume, and prevented the decline in bone mass seen in female mice with age. The op/op mouse has a deficiency in pancreatic ß cells and in Paneth cells in the gut wall. Only the latter was reproduced by the antibody treatment and was associated with increased goblet cell number but no change in villus architecture. Male op/op mice are infertile as a result of testosterone insufficiency. Anti-CSF1R treatment ablated interstitial macrophages in the testis, but there was no sustained effect on testosterone or LH. The results indicate an ongoing requirement for CSF1R signaling in macrophage and OCL homeostasis but indicate that most effects of CSF1 and CSF1R mutations are due to effects on development.

CSF-1, IGF-1, and the control of postnatal growth and development.

Growth hormone controls somatic growth in mammals by regulating the production of IGF-1, which is predominantly made by the liver. The development of cells within the MPS is controlled by the lineage-specific growth factor M-CSF (CSF-1). In this review, we summarize the role of CSF-1-dependent macrophages in somatic growth and organogenesis. We propose that macrophages are the major extrahepatic source of IGF-1 and that a surge of CSF-1 production contributes to the control of postnatal growth and organ maturation. Accordingly, CSF-1 may be considered a part of the GH/IGF-1 axis.