Opposing Effects of Adenosine and Inosine in Human Subcutaneous Fibroblasts May Be Regulated by Third Party ADA Cell Providers

Abstract

Human subcutaneous fibroblasts (HSCF) challenged with inflammatory mediators release huge amounts of ATP, which rapidly generates adenosine. Given the nucleoside’s putative relevance in wound healing, dermal fibrosis, and myofascial pain, we investigated the role of its precursor, AMP, and of its metabolite, inosine, in HSCF cells growth and collagen production. AMP (30 µM) was rapidly (t½ 3 ± 1 min) dephosphorylated into adenosine by CD73/ecto-5′-nucleotidase. Adenosine accumulation (t½ 158 ± 17 min) in the extracellular fluid reflected very low cellular adenosine deaminase (ADA) activity. HSCF stained positively against A2A and A3 receptors but were A1 and A2B negative. AMP and the A2A receptor agonist, CGS21680C, increased collagen production without affecting cells growth. The A2A receptor antagonist, SCH442416, prevented the effects of AMP and CGS21680C. Inosine and the A3 receptor agonist, 2Cl-IB-MECA, decreased HSCF growth and collagen production in a MRS1191-sensitive manner, implicating the A3 receptor in the anti-proliferative action of inosine. Incubation with ADA reproduced the inosine effect. In conclusion, adenosine originated from extracellular ATP hydrolysis favors normal collagen production by HSCF via A2A receptors. Inhibition of unpredicted inosine formation by third party ADA cell providers (e.g., inflammatory cells) may be a novel therapeutic target to prevent inappropriate dermal remodeling via A3 receptors activation.

Therapeutic Perspectives of Adenosine Deaminase Inhibition in Cardiovascular Diseases

“…While from the other, the opposed effects of inosine and adenosine have been demonstrated. In the study of Herman-de-Sousa et al [ 32 ] adenosine favored normal collagen production by human subcutaneous fibroblasts via A 2A R, whereas, inosine via A 3 R stimulated inappropriate dermal remodeling, providing ADA inhibition as a therapeutic target to prevent connective tissue disorganization. It is worth noting that inosine at micromolar concentrations stimulates the receptors with high affinity for adenosine, which may be a compensatory anti-inflammatory mechanism in the case of increased eADA activity.…”

Pharmacological Tuning of Adenosine Signal Nuances Underlying Heart Failure With Preserved Ejection Fraction

“…One may, therefore, hypothesize that the anti-fibrotic effect of adenosine via A 2A AR activation is operated either, directly, by inhibiting growth and/or differentiation of cardiac fibroblast, or, indirectly, through nucleoside-induced immunosuppressive effects, thereby preventing pro-fibrotic cells chemotaxis and the release of inflammatory mediators. In addition to these possibilities, adenosine deamination to inosine by third-party ADA cell providers (e.g., inflammatory cells) may also play a role in the anti-fibrotic effect of adenosine, as we recently demonstrated in human subcutaneous fibroblasts (Herman- de-Sousa et al, 2020). Thus, clarification of the mechanism and receptor involved in the anti-fibrotic role of adenosine in myocardial fibrosis is warranted given to the fact it may exert pro-fibrotic effects in other organs, such as the liver (Chan et al, 2006) and the skin (Herman- de-Sousa et al, 2020), while the opposite is verified with the A 2B AR which inhibits fibrosis in the heart (see below) and promotes fibrosis of the lung (reviewed in (Bessa-Goncalves et al, 2018)).…”

“…In addition to these possibilities, adenosine deamination to inosine by third-party ADA cell providers (e.g., inflammatory cells) may also play a role in the anti-fibrotic effect of adenosine, as we recently demonstrated in human subcutaneous fibroblasts (Herman- de-Sousa et al, 2020). Thus, clarification of the mechanism and receptor involved in the anti-fibrotic role of adenosine in myocardial fibrosis is warranted given to the fact it may exert pro-fibrotic effects in other organs, such as the liver (Chan et al, 2006) and the skin (Herman- de-Sousa et al, 2020), while the opposite is verified with the A 2B AR which inhibits fibrosis in the heart (see below) and promotes fibrosis of the lung (reviewed in (Bessa-Goncalves et al, 2018)).…”

“…Confirmation of the protective role of A 3 AR blockage on the inflammatory component attenuating pressure overload cardiac remodelling requires the use of more potent and selective drugs. It is also worth noting that the adenosine deamination metabolite, inosine, may exert opposite effects to adenosine in the heart via A 3 AR receptors, which activation is strengthened when tissues become infiltrated by ADA-bearing inflammatory cells ( Herman-de-Sousa et al, 2020 ). This concept raises new lines of research using multi-target drugs designed to increase local adenosine levels while decreasing inosine formation (e.g., ADA inhibitors with blocking properties of adenosine cellular uptake, like the lymphocytotoxic drug 2′-deoxycoformycin or pentostatin) applied in combination with enzymatically stable adenosine receptor agonists and/or antagonists ( Herman-de-Sousa et al, 2020 ).…”

“…Major benefits of adenosine in this endeavor result from A 1 AR and A 2A AR activation (Hamad et al, 2012;Quast et al, 2017;Zhou et al, 2020) associated or not with A 3 AR antagonism (Lu et al, 2008), which may also ameliorate symptoms and prognosis of HFpEF caused by cardiac pressure overload. Adenosine levels in the pulmonary circulation are pathologically low in patients with WHO group 1 pulmonary arterial hypertension (PAH) as a consequence of local endothelial dysfunction and increased inactivation by ADA, possibly indicating that adenosine A 1 AR and A 2A AR activation deficits unbalanced by inosine-mediated A 3 AR tone (Herman-de-Sousa et al, 2020) may contribute to maladaptive lung disease (Saadjian et al, 1999). In contrast to other vascular beds where adenosine has potent vasodilatory actions, in the pulmonary circulation, the nucleoside exerts dual opposing effects depending on the vascular tone (Cheng et al, 1996).…”