Suppressor of cytokine signaling 1 regulates embryonic myelopoiesis independently of its effects on T cell development

Suppressor of cytokine signaling 1 (SOCS1) has been shown to play important roles in the immune system. It acts as a key negative regulator of signaling via receptors for IFNs and other cytokines controlling T cell development, as well as Toll receptor signaling in macrophages and other immune cells. To gain further insight into SOCS1, we have identified and characterized the zebrafish socs1 gene, which exhibited sequence and functional conservation with its mammalian counterparts. Initially maternally derived, the socs1 gene showed early zygotic expression in mesodermal structures, including the posterior intermediate cell mass, a site of primitive hematopoiesis. At later time points, expression was seen in a broad anterior domain, liver, notochord, and intersegmental vesicles. Morpholino-mediated knockdown of socs1 resulted in perturbation of specific hematopoietic populations prior to the commencement of lymphopoiesis, ruling out T cell involvement. However, socs1 knockdown also lead to a reduction in the size of the developing thymus later in embryogenesis. Zebrafish SOCS1 was shown to be able to interact with both zebrafish Jak2a and Stat5.1 in vitro and in vivo. These studies demonstrate a conserved role for SOCS1 in T cell development and suggest a novel T cell-independent function in embryonic myelopoiesis mediated, at least in part, via its effects on receptors using the Jak2-Stat5 pathway.

Aberrant cytokine signaling in ovarian cancer

The studies described in this thesis suggest that IL-6R family members have the potential to significantly impact on ovarian cancer progression. These results suggest that assessment of IL-6R family expression represents a potential prognostic tool in determining appropriate therapy for ovarian cancer patients.

Cytokine regulation of skeletal muscle fatty acid metabolism: effect of interleukin-6 and tumor necrosis factor-α

IL-6 and TNF-α have been associated with insulin resistance and type 2 diabetes. Furthermore, abnormalities in muscle fatty acid (FA) metabolism are strongly associated with the development of insulin resistance. However, few studies have directly examined the effects of either IL-6 or TNF-α on skeletal muscle FA metabolism. Here, we used a pulse-chase technique to determine the effect of IL-6 (50-5,000 pg/ml) and TNF-α (50-5,000 pg/ml) on FA metabolism in isolated rat soleus muscle. IL-6 (5,000 pg/ml) increased exogenous and endogenous FA oxidation by ≃50% (P < 0.05) but had no effect on FA uptake or incorporation of FA into endogenous lipid pools. In contrast, TNF-α had no effect on FA oxidation but increased FA incorporation into diacylglycerol (DAG) by 45% (P < 0.05). When both IL-6 (5,000 pg/ml) and insulin (10 mU/ml) were present, IL-6 attenuated insulin's suppressive effect on FA oxidation, increasing exogenous FA oxidation (+37%, P < 0.05). Furthermore, in the presence of insulin, IL-6 reduced the esterification of FA to triacylglycerol by 22% (P < 0.05). When added in combination with IL-6 or leptin (10 μg/ml), the TNF-α-induced increase in DAG synthesis was inhibited. In conclusion, the results demonstrate that IL-6 plays an important role in regulating fat metabolism in muscle, increasing rates of FA oxidation, and attenuating insulin's lipogenic effects. In contrast, TNF-α had no effect on FA oxidation but increased FA incorporation into DAG, which may be involved in the development of TNF-α-induced insulin resistance in skeletal muscle.

Effect of cell-surface components and metabolites of lactic acid bacteria and probiotic organisms on cytokine production and induction of CD25 expression in human peripheral mononuclear cells

In the current study, the relative contribution of cell-surface components (CSC) and cell-free supernatants (CFS) in the immuno-modulatory properties of 17 strains of probiotic and lactic acid bacteria (LAB) was assessed. The production of pro- and antiinflammatory cytokines including IL-2, IL-4, IL-10, IL-12 p70, IFN-γ, tumor necrosis factor-α (TNF-α), and transforming growth factor-β was measured at different time points after stimulation of buffy coat derived-peripheral blood mononuclear cells (PBMC) from healthy donors with CSC and CFS of probiotic and LAB. Results showed that CSC of probiotic and LAB strains induced production of T helper 1 and 2 type cytokines. Transforming growth factor-β was stimulated at highest concentrations, followed by IL-10 and TNF-α. The CFS of all tested bacterial strains induced PBMC for significantly high levels of IL-10 secretion compared with unstimulated cells, but the values were less than lipopolysaccharide-stimulated cells. Cytokines due to CFS stimulation showed declined concentration for IL-2, TNF-α, and IL-4, and complete disappearance of IL-12, IFN-γ, and transforming growth factor-β in the cultured medium at 96 h of incubation. Results of cytokine data demonstrate proinflammatory TNF-α immune responses are mainly directed through cell-surface structures of probiotic and LAB, but antiinflammatory immune responses are mediated both by metabolites and cell-surfaces of these bacteria. The induction of CD4(+)CD25(+) regulatory T cells after stimulation of PBMC with CSC and CFS of probiotic and LAB showed regulatory T cell activity appeared to be influenced both by the CSC and metabolites, but was principally triggered by cell surfaces of probiotic and LAB strains.

Identification of a gene for an ancient cytokine, interleukin 15-like, in mammals; interleukins 2 and 15 co-evolved with this third family member, all sharing binding motifs for IL-15Rα

Interleukins 2 and 15 (IL-2 and IL-15) are highly differentiated but related cytokines with overlapping, yet also distinct functions, and established benefits for medical drug use. The present study identified a gene for an ancient third IL-2/15 family member in reptiles and mammals, interleukin 15-like (IL-15L), which hitherto was only reported in fish. IL-15L genes with intact open reading frames (ORFs) and evidence of transcription, and a recent past of purifying selection, were found for cattle, horse, sheep, pig and rabbit. In human and mouse the IL-15L ORF is incapacitated. Although deduced IL-15L proteins share only ~21 % overall amino acid identity with IL-15, they share many of the IL-15 residues important for binding to receptor chain IL-15Rα, and recombinant bovine IL-15L was shown to interact with IL-15Rα indeed. Comparison of sequence motifs indicates that capacity for binding IL-15Rα is an ancestral characteristic of the IL-2/15/15L family, in accordance with a recent study which showed that in fish both IL-2 and IL-15 can bind IL-15Rα. Evidence reveals that the species lineage leading to mammals started out with three similar cytokines IL-2, IL-15 and IL-15L, and that later in evolution (1) IL-2 and IL-2Rα receptor chain acquired a new and specific binding mode and (2) IL-15L was lost in several but not all groups of mammals. The present study forms an important step forward in understanding this potent family of cytokines, and may help to improve future strategies for their application in veterinarian and human medicine.