Blockage of the adenosine A2B receptor prevents cardiac fibroblasts overgrowth in rats with pulmonary arterial hypertension.

Sustained pressure overload and fibrosis of the right ventricle (RV) are the leading causes of mortality in pulmonary arterial hypertension (PAH). Although the role of adenosine in PAH has been attributed to the control of pulmonary vascular tone, cardiac reserve, and inflammatory processes, the involvement of the nucleoside in RV remodelling remains poorly understood. Conflicting results exist on targeting the low-affinity adenosine A2B receptor (A2BAR) for the treatment of PAH mostly because it displays dual roles in acute vs. chronic lung diseases. Herein, we investigated the role of the A2BAR in the viability/proliferation and collagen production by cardiac fibroblasts (CFs) isolated from RVs of rats with monocrotaline (MCT)-induced PAH. CFs from MCT-treated rats display higher cell viability/proliferation capacity and overexpress A2BAR compared to the cells from healthy littermates. The enzymatically stable adenosine analogue, 5'-N-ethylcarboxamidoadenosine (NECA, 1-30 µM), concentration-dependently increased growth, and type I collagen production by CFs originated from control and PAH rats, but its effects were more prominent in cells from rats with PAH. Blockage of the A2BAR with PSB603 (100 nM), but not of the A2AAR with SCH442416 (100 nM), attenuated the proliferative effect of NECA in CFs from PAH rats. The A2AAR agonist, CGS21680 (3 and 10 nM), was virtually devoid of effect. Overall, data suggest that adenosine signalling via A2BAR may contribute to RV overgrowth secondary to PAH. Therefore, blockage of the A2AAR may be a valuable therapeutic alternative to mitigate cardiac remodelling and prevent right heart failure in PAH patients.

Controlling Macrophage Polarization to Modulate Inflammatory Cues Using Immune-Switch Nanoparticles.

The persistence of inflammatory mediators in tissue niches significantly impacts regenerative outcomes and contributes to chronic diseases. Interleukin-4 (IL4) boosts pro-healing phenotypes in macrophages (Mφ) and triggers the activation of signal transducer and activator of transcription 6 (STAT6). Since the IL4/STAT6 pathway reduces Mφ responsiveness to inflammation in a targeted and precise manner, IL4 delivery offers personalized possibilities to overcome inflammatory events. Despite its therapeutic potential, the limited success of IL4-targeted delivery is hampered by inefficient vehicles. Magnetically assisted technologies offer precise and tunable nanodevices for the delivery of cytokines by combining contactless modulation, high tissue penetration, imaging features, and low interference with the biological environment. Although superparamagnetic iron oxide nanoparticles (SPION) have shown clinical applicability in imaging, SPION-based approaches have rarely been explored for targeted delivery and cell programming. Herein, we hypothesized that SPION-based carriers assist in efficient IL4 delivery to Mφ, favoring a pro-regenerative phenotype (M2φ). Our results confirmed the efficiency of SPION-IL4 and Mφ responsiveness to SPION-IL4 with evidence of STAT6-mediated polarization. SPION-IL4-treated Mφ showed increased expression of M2φ associated-mediators (IL10, ARG1, CCL2, IL1Ra) when compared to the well-established soluble IL4. The ability of SPION-IL4 to direct Mφ polarization using sophisticated magnetic nanotools is valuable for resolving inflammation and assisting innovative strategies for chronic inflammatory conditions.

Magnetic Stimulation Drives Macrophage Polarization in Cell to-Cell Communication with IL-1ß Primed Tendon Cells.

Inflammation is part of the natural healing response, but it has been simultaneously associated with tendon disorders, as persistent inflammatory events contribute to physiological changes that compromise tendon functions. The cellular interactions within a niche are extremely important for healing. While human tendon cells (hTDCs) are responsible for the maintenance of tendon matrix and turnover, macrophages regulate healing switching their functional phenotype to environmental stimuli. Thus, insights on the hTDCs and macrophages interactions can provide fundamental contributions on tendon repair mechanisms and on the inflammatory inputs in tendon disorders. We explored the crosstalk between macrophages and hTDCs using co-culture approaches in which hTDCs were previously stimulated with IL-1ß. The potential modulatory effect of the pulsed electromagnetic field (PEMF) in macrophage-hTDCs communication was also investigated using the magnetic parameters identified in a previous work. The PEMF influences a macrophage pro-regenerative phenotype and favors the synthesis of anti-inflammatory mediators. These outcomes observed in cell contact co-cultures may be mediated by FAK signaling. The impact of the PEMF overcomes the effect of IL-1ß-treated-hTDCs, supporting PEMF immunomodulatory actions on macrophages. This work highlights the relevance of intercellular communication in tendon healing and the beneficial role of the PEMF in guiding inflammatory responses toward regenerative strategies.

Pulsed Electromagnetic Field Modulates Tendon Cells Response in IL-1ß-Conditioned Environment.

Strategies aiming at controlling and modulating inflammatory cues may offer therapeutic solutions for improving tendon regeneration. This study aims to investigate the modulatory effect of pulsed electromagnetic field (PEMF) on the inflammatory profile of human tendon-derived cells (hTDCs) after supplementation with interleukin-1ß (IL-1ß). IL-1ß was used to artificially induce inflammatory cues associated with injured tendon environments. The PEMF effect was investigated varying the frequency (5 or 17 Hz), intensity (1.5, 4, or 5 mT), and duty-cycle (10% or 50%) parameters to which IL-1ß-treated hTDCs were exposed to. A PEMF actuation with 4 mT, 5 Hz and a 50% duty cycle decreased the production of IL-6 and tumor necrosis factor-α (TNF-α), as well as the expression of TNFα, IL-6, IL-8, COX-2, MMP-1, MMP-2, and MMP-3, while IL-4, IL-10, and TIMP-1 expression increased. These results suggest that PEMF stimulation can modulate hTDCs response in an inflammatory environment holding therapeutic potential for tendon regenerative strategies. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:160-172, 2020.

ADP-Induced Ca2+ Signaling and Proliferation of Rat Ventricular Myofibroblasts Depend on Phospholipase C-Linked TRP Channels Activation Within Lipid Rafts.

Nucleotides released during heart injury affect myocardium electrophysiology and remodeling through P2 purinoceptors activation in cardiac myofibroblasts. ATP and UTP endorse [Ca2+ ]i accumulation and growth of DDR-2/α-SMA-expressing myofibroblasts from adult rat ventricles via P2Y4 and P2Y2 receptors activation, respectively. Ventricular myofibroblasts also express ADP-sensitive P2Y1 , P2Y12 , and P2Y13 receptors as demonstrated by immunofluorescence confocal microscopy and western blot analysis, but little information exists on ADP effects in these cells. ADP (0.003-3 mM) and its stable analogue, ADPßS (100 µM), caused fast [Ca2+ ]i transients originated from thapsigargin-sensitive internal stores, which partially declined to a plateau sustained by capacitative Ca2+ entry through transient receptor potential (TRP) channels inhibited by 2-APB (50 µM) and flufenamic acid (100 µM). Hydrophobic interactions between Gq/11 -coupled P2Y purinoceptors and TRP channels were suggested by prevention of the ADP-induced [Ca2+ ]i plateau following PIP2 depletion with LiCl (10 mM) and cholesterol removal from lipid rafts with methyl-ß-cyclodextrin (2 mM). ADP [Ca2+ ]i transients were insensitive to P2Y1 , P2Y12 , and P2Y13 receptor antagonists, MRS2179 (10µM), AR-C66096 (0.1 µM), and MRS2211 (10µM), respectively, but were attenuated by suramin and reactive blue-2 (100 µM) which also blocked P2Y4 receptors activation by UTP. Cardiac myofibroblasts growth and type I collagen production were favored upon activation of MRS2179-sensitive P2Y1 receptors with ADP or ADPßS (30 µM). In conclusion, ADP exerts a dual role on ventricular myofibroblasts: [Ca2+ ]i transients are mediated by fast-desensitizing P2Y4 receptors, whereas the pro-fibrotic effect of ADP involves the P2Y1 receptor activation. Data also show that ADP-induced capacitative Ca2+ influx depends on phospholipase C-linked TRP channels opening in lipid raft microdomains. J. Cell. Physiol. 232: 1511-1526, 2017. © 2016 Wiley Periodicals, Inc.