Pro-inflammatory cytokines are directly implicated in the pathogenesis of Rheumatoid arthritis (RA). Variable clinical response to cytokine targeted therapies as TNFalpha and IL-6, strongly highlights the heterogeneity of inflammatory process in RA. Another cytokine, IL-15 has also been related to the inflammatory process in RA. Recently we described for the first time, the presence of its specific receptor, IL-15Ralpha, in synovial fluid (SF). The aim of this work was to compare the expression profile of IL-15Ralpha, its ligand IL-15, TNFalpha and IL-6 and how these cytokines are correlated in SF from RA patients taking as a reference Osteoarthritis (OA), an articular but not autoinmmune disease. Synovial fluids were obtained from the knee joints of 60 patients, 30 with confirmed diagnosis of RA and 30 with OA diagnosis. The levels of TNFalpha, IL-6, IL-15 and IL-15Ralpha were measured by ELISA. A statistical analysis was performed with GraphPad Prism v5.0 using the Mann–Whitney U test and Spearman’s rank correlation. A cluster analysis was run in MeV software v4.9.0 and differences across clusters were evaluated by an ANOVA including post-test analysis. We found higher and significant levels of TNFalpha, IL-6 and IL-15Ralpha but not of IL-15 in RA compared with the OA group. Additionally, a high inter-individual variability in the levels of these 4 cytokines was observed in RA, although we identified 4 patients’ subgroups by cluster analysis of cytokines concentration in SF. We also found a positive correlation between IL-15Ralpha-IL-6 and IL-15Ralpha-IL-15, but not for other pairs of cytokines in RA. In addition we found correlation between the value of IL-15Ralpha in SF and disease activity score, DAS28. In our current work we found a high inter-individual variability in the levels of TNFalpha, IL-6, IL-15 and IL-15Ralpha in SF of RA patients and were identified four principal clusters of cytokines concentration in SF, suggesting the importance of identifying disease subset of patients for personalized treatment. Finally, we found a correlation between IL-15Ralpha-IL-6, IL-15Ralpha-IL-15, but we did not find any correlation between other pairs of studied cytokines in SF.

Interleukin-15 is an immunostimulatory cytokine overexpressed in several autoimmune and inflammatory diseases such as Rheumatoid Arthritis, psoriasis and ulcerative colitis; thus, inhibition of IL-15-induced signaling could be clinically beneficial in these disorders. Our approach to neutralize IL-15 consisted in active immunization with structurally modified human IL-15 (mhIL-15) with the aim to induce neutralizing antibodies against native IL-15. In the present study, we characterized the antibody response in Macaca fascicularis, non-human primates that were immunized with a vaccine candidate containing mhIL-15 in Aluminum hydroxide (Alum), Montanide and Incomplete Freund’s Adjuvant. Immunization with mhIL-15 elicited a specific antibodies response that neutralized native IL-15-dependent biologic activity in a CTLL-2 cell proliferation assay. The highest neutralizing response was obtained in macaques immunized with mhIL-15 adjuvanted in Alum. This response, which was shown to be transient, also inhibited the activity of simian IL-15 and did not affect the human IL-2-induced proliferation of CTLL-2 cells. Also, in a pool of synovial fluid cells from two Rheumatoid Arthritis patients, the immune sera slightly inhibited TNF-α secretion. Finally, it was observed that this vaccine candidate neither affect animal behavior, clinical status, blood biochemistry nor the percentage of IL-15-dependent cell populations, specifically CD56+ NK and CD8+ T cells. Our results indicate that vaccination with mhIL-15 induced neutralizing antibodies to native IL-15 in non-human primates. Based on this fact, we propose that this vaccine candidate could be potentially beneficial for treatment of diseases where IL-15 overexpression is associated with their pathogenesis.

Recombinant simian IL-15 (siIL-15) was obtained for the preclinical assessment of an anti-human IL-15 vaccine. For this purpose, the cDNA from peripheral blood mononuclear cells of a Macaca fascicularis monkey was cloned into a pIL-2 vector. The siIL-15 was expressed in Escherichia coli strain W3110 as an insoluble protein which accounted for 13% of the total cellular proteins. Inclusion bodies were solubilized in an 8 M urea solution, which was purified by ion exchange and reverse phase chromatography up to 92% purity. The protein identity was validated by electrospray ionization-mass spectrometry, confirming the presence of the amino acids which distinguish the siIL-15 from human IL-15. The purified siIL-15 stimulates the proliferation of cytotoxic T-lymphocytes line (CTLL)-2 and Kit 225 cells with EC₅₀ values of 3.1 and 32.5 ng/mL, respectively. Antisera from modified human IL-15-immunized macaques were reactive to human and simian IL-15 in enzyme-linked immunosorbent assays. Moreover, the anti-human IL-15 antibodies from immune sera inhibited siIL-15 activity in CTLL-2 and Kit 225 cells, supporting the activity and purity of recombinant siIL-15. These results indicate that the recombinant siIL-15 is biologically active in two IL-15-dependent cell lines, and it is also suitable for the preclinical evaluation of an IL-15-based therapeutic vaccine.

Phycocyanobilin (PCB), the covalently bound chromophore of the cyanobacterial protein C-phycocyanin (CPC), is recognized as a bioactive molecule with neuroprotective and anti-inflammatory properties. PCB and CPC, frequently coexisting in Spirulina extracts or experimental formulations, have demonstrated beneficial effects in preclinical models of multiple sclerosis, ischemic stroke, and Alzheimer’s disease. Reported mechanisms include attenuation of oxidative stress, reduction of neuroinflammation, and preservation of mitochondrial function, thereby contributing to a reparative microenvironment within the central nervous system. PCB can be obtained through two complementary approaches: Extraction from cyanobacterial biomass, where it remains covalently bound to CPC, and heterologous biosynthesis in Escherichia coli (E. coli), which enables production of free PCB as a high-purity, scalable linear tetrapyrrole suitable for translational applications. This mini-review summarizes current evidence on the neuroprotective actions of PCB and CPC, highlights their molecular targets, and discusses biotechnological advances that support their potential role in remyelination. By bridging natural pigment pharmacology with recombinant production strategies, PCB is positioned as a multitarget candidate of growing interest for the development of future neuroprotective and neurorepair therapies.