Therapeutic activity and biodistribution of a nano-sized polymer-dexamethasone conjugate intended for the targeted treatment of rheumatoid arthritis.

Rheumatoid arthritis is a chronic inflammatory autoimmune disease caused by alteration of the immune system. Current therapies have several limitations and the use of nanomedicines represents a promising strategy to overcome them. By employing a mouse model of adjuvant induced arthritis, we aimed to evaluate the biodistribution and therapeutic effects of glucocorticoid dexamethasone conjugated to a nanocarrier based on biocompatible N-(2-hydroxypropyl) methacrylamide copolymers. We observed an increased accumulation of dexamethasone polymer nanomedicines in the arthritic mouse paw using non-invasive fluorescent in vivo imaging and confirmed it by the analysis of tissue homogenates. The dexamethasone conjugate exhibited a dose-dependent healing effect on arthritis and an improved therapeutic outcome compared to free dexamethasone. Particularly, significant reduction of accumulation of RA mediator RANKL was observed. Overall, our data suggest that the conjugation of dexamethasone to a polymer nanocarrier by means of stimuli-sensitive spacer is suitable strategy for improving rheumatoid arthritis therapy.

A forskolin-mediated increase in cAMP promotes T helper cell differentiation into the Th1 and Th2 subsets rather than into the Th17 subset.

The adenylyl cyclase (AC) signaling pathway is suggested to be a key regulator of immune system functions. However, specific effects of cyclic adenosine monophosphate (cAMP) on T helper (Th) cell differentiation and functions are unclear. The involvement of cAMP in the Th cell differentiation program, in particular the development of Th1, Th2, and Th17 subsets, was evaluated employing forskolin (FSK), a labdane diterpene well known as an AC activator. FSK mediated an elevation in Th1-specific markers reinforcing the Th1 cell phenotype. The Th2 differentiation was supported by FSK, though cell metabolism was negatively affected. In contrast, the Th17 immunophenotype was severely suppressed leading to the highly specific upregulation of CXCL13. The causality between FSK-elicited cAMP production and the observed reinforcement of Th2 differentiation was established by using AC inhibitor 2',5'-dideoxyadenosine, which reverted the FSK effects. Overall, an FSK-mediated cAMP increase affects Th1, Th2 and Th17 differentiation and can contribute to the identification of novel therapeutic targets for the treatment of Th cell-related pathological processes.

Pseurotin D inhibits delayed type IV hypersensitivity response.

Pseurotins, secondary metabolites of fungi, represent a group of bioactive natural products with newly recognized biological activities, including the modulation of specific immune response. However, the type of immune response affected by pseurotins and the mechanistic details underlying these effects are still not understood. Thus, the aim of the current study was to examine the effects of pseurotin D on delayed-type IV hypersensitivity (DTH) reaction induced by chicken ovalbumin in vivo and to examine the effects of pseurotin D on major types of leukocytes responsible for DTH development in vitro. Pseurotin D significantly decreased paw swelling, the major symptom of DTH, as well as the DTH-related production of pro-inflammatory cytokine IL-1ß, IL-4, IL-6, IFN-γ and anti-inflammatory cytokine IL-10 in paws tissue, spleen enlargement, and DTH-related changes in leukocyte counts in peripheral blood. In vitro, pseurotin D mediated a decrease in the proliferation and differentiation of both Th1 and Th2 cells, as was concluded on the basis of the inhibition of the gene expressions of Gata3 and Tbx21 and the production of effector cytokines IFN-γ and IL-13 in vitro. Further, pseurotin D significantly inhibited the activation and differentiation of B cells, as was documented by the significant inhibition of B cell proliferation, CD138 expression, and IgE production. In conclusion, the results show the potential of pseurotin D to inhibit DTH reaction, this phenomenon involving the inhibition of the activation and differentiation of both T cells and B cells.

Myeloperoxidase Deficiency Alters the Process of the Regulated Cell Death of Polymorphonuclear Neutrophils.

Polymorphonuclear neutrophils (PMNs) play a key role in host defense. However, their massive accumulation at the site of inflammation can delay regenerative healing processes and can initiate pathological inflammatory processes. Thus, the efficient clearance of PMNs mediated by the induction of regulated cell death is a key process preventing the development of these pathological conditions. Myeloperoxidase (MPO), a highly abundant enzyme in PMN granules, primarily connected with PMN defense machinery, is suggested to play a role in PMN-regulated cell death. However, the contribution of MPO to the mechanisms of PMN cell death remains incompletely characterized. Herein, the process of the cell death of mouse PMNs induced by three different stimuli - phorbol 12-myristate 13-acetate (PMA), opsonized streptococcus (OST), and N-formyl-met-leu-phe (fMLP) - was investigated. MPO-deficient PMNs revealed a significantly decreased rate of cell death characterized by phosphatidylserine surface exposure and cell membrane permeabilization. An inhibitor of MPO activity, 4-aminobenzoic acid hydrazide, did not exhibit a significant effect on PMA-induced cell death compared to MPO deficiency. Interestingly, only the limited activation of markers related to apoptotic cell death was observed (e.g. caspase 8 activation, Bax expression) and they mostly did not correspond to phosphatidylserine surface exposure. Furthermore, a marker characterizing autophagy, cleavage of LC3 protein, as well as histone H3 citrullination and its surface expression was observed. Collectively, the data show the ability of MPO to modulate the life span of PMNs primarily through the potentiation of cell membrane permeabilization and phosphatidylserine surface exposure.