Adenosine signalling pathway as modulator of the inflammatory response

Adenosine is an endogenous nucleoside that has been recognised to be a molecule with autocrine/paracrine functions, acting as a signal molecule to preserve host defence and tissue integrity during inflammation and trauma in addition to its important role as homeostatic regulator. The physiological activities of adenosine involve its interactions with four types of receptors, designed as A1, A2A, A2B and A3. Adenosine mediates its anti-inflammatory activity primarily through the A2A receptor (A2AAR). The ecto-5’nucleotidase/CD73 degrades AMP to adenosine and represents a key enzyme for adenosine accumulation at the site of injury. The aim of this research work was to explore different aspects of adenosine signalling pathway in inflammation. Several findings implicate the adenosine signalling pathway as an innate mechanism to attenuate excessive tissue damage and identify CD73 as critical control points for endogenous adenosine generation. It has been shown that CD73 plays an important role in regulating vascular permeability and leukocyte trafficking in inflammatory disease; and a crucial role in the regulation of immune/inflammatory cell function. A better understanding of the role of CD73 enzyme in the development of inflammatory processes can help to identify new therapeutic strategies aimed at strengthening the endogenous anti-inflammatory mechanisms. For this reason we sought to investigate on the role of CD73, the key enzyme in “switching on” adenosine signalling, in the development of inflammation through its pharmacological blockade by using the selective inhibitor, adenosine 5'-(α,β-methylene) diphosphate (APCP; 400 μg/site), in an in vivo model of acute inflammation represented by carrageenan-induced pleurisy in rats. We found that local inhibition of CD73 significantly increased cell accumulation, exudate formation and pro-inflammatory cytokine content into the pleural cavity in the acute phase of inflammation with no differences in the sub-acute phase. The in vivo treatment with APCP induced cells recruited into the pleural cavity to change in a phenotype with increased ability to migrate in vitro either in the presence or in the absence of a chemotactic stimulus. In parallel, these cells showed a reduced CD73 expression and activity compared to cells collected from control group. In addition, APCP, in vitro, strongly increased the ability of cells from control groups to migrate in the presence of a chemotactic stimulus. Local inhibition of CD73 increased also the infiltration of the lung with polymorphonuclear leukocytes (PMNs) and the degree of lung injury 4 hours following carrageenan injection. The interest to explore the role of adenosine signalling pathway in the control of inflammation has been growing further following evidence that adenosine signalling is also involved in the mechanism of action of some well-known anti-inflammatory drugs. With regard to this, we focused our interest on the possible involvement of adenosine signalling in the anti-inflammatory mechanism of nimesulide, in vivo (rat paw oedema) and in vitro (J774A.1cell line); indeed, there is evidence that nimesulide anti-inflammatory effect is the consequence of regulation of the production and actions of a wide range of inflammatory mediators, independently from the sole cyclooxygenase-2 (COX-2) enzyme inhibition. To date, the molecular mechanisms at the basis of nimesulide peculiar pharmacological effects are still unclear. In vivo, in the model of carrageenan-induced rat paw oedema, we found that the anti-inflammatory effect of nimesulide (5 mg/kg i.p.) was inhibited by pre-treatment with the adenosine A2A receptor antagonist, ZM 241385 (3 mg/kg i.p.), and by local administration of the CD73 inhibitor, APCP (400 μg/paw). Furthermore, we observed increased activity of 5'-nucleotidase/CD73 in plasma and paws of nimesulide-treated rats, 4 h following oedema induction that represented the inflammatory peaking point. In vitro, the inhibitory effect of nimesulide on nitrite and prostaglandin (PG)E2 (PGE2) production by lipopolysaccharide (LPS)-activated J774 macrophage cell line was again reverted by ZM 241385 and APCP. Furthermore, nimesulide increased CD73 activity in J774 macrophages while it did not inhibit nitrite accumulation by LPS-activated small interfering RNA (SiRNA) CD73 silenced J774 macrophages. Our data demonstrate that the anti-inflammatory effect of nimesulide is, in part, mediated by CD73-derived adenosine acting on A2A receptors. There is evidence that A2AAR activation beside anti-inflammatory effects promotes wound healing and extracellular matrix production; given that extracellular matrix and fibroblasts take an active part in the modulation of inflammation beside wound healing, we investigated whether and how extracellular matrix was involved in the anti-inflammatory effect of A2A receptor. Specifically, we evaluated changes in tissue fibroblast growth factor-2 (FGF-2), an important growth factor for fibroblasts that has been shown to facilitate not only tissue regeneration but also to dampen inflammation, following systemic administration of the A2A agonist, CGS 21680, in a rat model of acute inflammation (paw oedema). We observed that CGS 21680 prevented oedema development and inflammation, confirming an anti-inflammatory effect of A2AR. The effect of CGS 21680 was specific, through A2A adenosine receptor stimulation, as revealed by co-administration with ZM 241385 that reverted CGS 21680 inhibitory effect. On the basis of histological analysis showing an increased matrix deposition following rat treatment with CGS 21680, we evaluated whether the beneficial effect of A2A agonist, CGS 21680, was paralleled by changes in FGF-2 expression. Interestingly, we found that the expression of FGF-2 in rat paws, evaluated at each hour following carrageenan injection, was increased following rat treatment with CGS 21680. Immunofluorescence analysis confirmed data obtained by western blot and also showed spots of co-localization between A2AR and FGF-2. In conclusion, in this research work we demonstrate that CD73 regulates cell migration in the acute phase of inflammation and that the anti-inflammatory effects mediated by A2AR activation are paralleled by changes in extracellular matrix morphology. These findings suggest the important role of CD73/adenosine/A2A signalling in the control of the acute phase of inflammation, characterised by PMNs infiltration, but also in the control of a late phase, characterised by re-arrangement of extracellular matrix. In addition, we also demonstrate that CD73/adenosine/A2A axis is involved in the mode of action of nimesulide. Our study may open a path to re-evaluate the mechanism of action of nimesulide and to identify new therapeutic opportunities in COX-2 inhibitors which display a more potent activity on adenosine signalling. Furthermore, these results may give the cue to project an innovative anti-inflammatory strategy based on the manipulation of endogenous anti-inflammatory pathways

Thrombo-Inflammation: A Focus on NTPDase1/CD39

There is increasing evidence for a link between inflammation and thrombosis. Following tissue injury, vascular endothelium becomes activated, losing its antithrombotic properties whereas inflammatory mediators build up a prothrombotic environment. Platelets are the first elements to be activated following endothelial damage; they participate in physiological haemostasis, but also in inflammatory and thrombotic events occurring in an injured tissue. While physiological haemostasis develops rapidly to prevent excessive blood loss in the endothelium activated by inflammation, hypoxia or by altered blood flow, thrombosis develops slowly. Activated platelets release the content of their granules, including ATP and ADP released from their dense granules. Ectonucleoside triphosphate diphosphohydrolase-1 (NTPDase1)/CD39 dephosphorylates ATP to ADP and to AMP, which in turn, is hydrolysed to adenosine by ecto-50 -nucleotidase (CD73). NTPDase1/CD39 has emerged has an important molecule in the vasculature and on platelet surfaces; it limits thrombotic events and contributes to maintain the antithrombotic properties of endothelium. The aim of the present review is to provide an overview of platelets as cellular elements interfacing haemostasis and inflammation, with a particular focus on the emerging role of NTPDase1/CD39 in controlling both processes

Editorial: Emerging mechanisms in cardiovascular disease

Editorial on the Research Topic Emerging mechanisms in cardiovascular disease The leading cause of worldwide mortality is cardiovascular disease (CVD) (Aboukhater et al., 2023). Despite many significant advances in the field, CVD continues to claim more lives than all cancers combined (Sawma et al., 2022). There is then an urgent need for more efficacious treatment modalities or therapeutics that could aid in the management of CVD (Badran et al., 2019; Maaliki et al., 2019; El-Hachem et al., 2021). For such potential new drugs to be determined, a better understanding of the underlying mechanisms and the potential targets is critical. This Research Topic seeks to highlight a few of these emerging mechanisms and targets that could be employed for a better treatment of CVD. Myocardial injury continues to be a major contributor to CVD-associated mortality. In this thematic issue, Liu et al. discuss how they established a model for coronary microembolization (CME) in rats, and report that ferroptosis and inflammation are two key players in CME-induced myocardial injury. The authors then show that suppressing ferroptosis attenuates myocardial injury and inflammation following CME. It appears that Ptgs2, a core factor in ferroptosis, and Hif1a are the two mediators of this suppressed ferroptosis. Importantly, the authors further report that by inhibiting the Hif1a/Ptgs2 axis, atorvastatin was able to suppress ferroptosis-dependent CME-precipitated myocardial injury and inflammation (Liu et al.)

Why do some asthma patients respond poorly to glucocorticoid therapy?

Glucocorticosteroids are the first-line therapy for controlling airway inflammation in asthma. They bind intracellular glucocorticoid receptors to trigger increased expression of anti-inflammatory genes and suppression of pro-inflammatory gene activation in asthmatic airways. In the majority of asthma patients, inhaled glucocorticoids are clinically efficacious, improving lung function and preventing exacerbations. However, 5–10 % of the asthmatic population respond poorly to high dose inhaled and then systemic glucocorticoids. These patients form a category of severe asthma associated with poor quality of life, increased morbidity and mortality, and constitutes a major societal and health care burden. Inadequate therapeutic responses to glucocorticoid treatment is also reported in other inflammatory conditions such as rheumatoid arthritis and inflammatory bowel disease; however, asthma represents the most studied steroid-refractory disease. Several cellular and molecular events underlying glucocorticoid resistance in asthma have been identified involving abnormalities of glucocorticoid receptor signaling pathways. These events have been strongly related to immunological dysregulation, genetic, and environmental factors such as cigarette smoking or respiratory infections. A better understanding of the multiple mechanisms associated with glucocorticoid insensitivity in asthma phenotypes could improve quality of life for people with asthma but would also provide transferrable knowledge for other inflammatory diseases. In this review, we provide an update on the molecular mechanisms behind steroid-refractory asthma. Additionally, we discuss some therapeutic options for treating those asthmatic patients who respond poorly to glucocorticoid therapy

Lack of Ecto-5'-Nucleotidase Protects Sensitized Mice against Allergen Challenge

Ecto-5'-nucleotidase (CD73), the ectoenzyme that together with CD39 is responsible for extracellular ATP hydrolysis and adenosine accumulation, regulates immune/inflammatory processes by controlling innate and acquired immunity cell functions. We previously demonstrated that CD73 is required for the assessment of a controlled allergic sensitization, in mice. Here, we evaluated the response to aerosolized allergen of female-sensitized mice lacking CD73 in comparison with their wild type counterpart. Results obtained show, in mice lacking CD73, the absence of airway hyperreactivity in response to an allergen challenge, paralleled by reduced airway CD23+B cells and IL4+T cells pulmonary accumulation together with reduced mast cells accumulation and degranulation. Our findings indicate CD73 as a potential therapeutic target for allergic asthma

Salvinorin A Inhibits Airway Hyperreactivity Induced by Ovalbumin Sensitization

Salvinorin A, a neoclerodane diterpene isolated from Salvia divinorum, exerts a number of pharmacological actions which are not solely limited to the central nervous system. Recently it has been demonstrated that Salvinorin A inhibits acute inflammatory response affecting leukotriene (LT) production. Since LTs are potent lipid mediators implicated in allergic diseases, we evaluated the effect of Salvinorin A on allergic inflammation and on airways following sensitization in the mouse. Mice were sensitized with s.c. injection of ovalbumin (OVA) on days 1 and 8. Sensitized mice received on days 9 and 12 on the shaved dorsal surface air administration to induce the development of the air-pouches. On day 15 animals were challenged by injection of OVA into the air-pouch. Salvinorin A, administered (10 mg/kg) before each allergen exposure, significantly reduced OVA-induced LT increase in the air pouch. This effect was coupled to a reduction in cell recruitment and Th2 cytokine production. In another set of experiments, mice were sensitized with OVA and both bronchial reactivity and pulmonary inflammation were assessed. Salvinorin A abrogated bronchial hyperreactivity and interleukin (IL)-13 production, without effect on pulmonary inflammation. Indeed cell infiltration and peribronchial edema were still present following diterpenoid treatment. Similarly, pulmonary IL-4 and plasmatic IgE levels were not modulated. Conversely, Salvinorin A significantly reduced LTC4 production in the lung of sensitized mice. Finally mast cell activity was evaluated by means of toluidine blue staining. Data obtained evidenced that Salvinorin A significantly inhibited mast cell degranulation in the lung. Our study demonstrates that Salvinorin A inhibits airway hyperreactivity induced by sensitization by inhibition of LT production and mast cell degranulation. In conclusion Salvinorin A could represent a promising candidate for drug development in allergic diseases such as asthma

Impairment of T cell development and acute inflammatory response in HIV-1 Tat transgenic mice

Immune activation and chronic inflammation are hallmark features of HIV infection causing T-cell depletion and cellular immune dysfunction in AIDS. Here, we addressed the issue whether HIV-1 Tat could affect T cell development and acute inflammatory response by generating a transgenic mouse expressing Tat in lymphoid tissue. Tat-Tg mice showed thymus atrophy and the maturation block from DN4 to DP thymic subpopulations, resulting in CD4(+) and CD8(+) T cells depletion in peripheral blood. In Tat-positive thymus, we observed the increased p65/NF-κB activity and deregulated expression of cytokines/chemokines and microRNA-181a-1, which are involved in T-lymphopoiesis. Upon LPS intraperitoneal injection, Tat-Tg mice developed an abnormal acute inflammatory response, which was characterized by enhanced lethality and production of inflammatory cytokines. Based on these findings, Tat-Tg mouse could represent an animal model for testing adjunctive therapies of HIV-1-associated inflammation and immune deregulation

Sex-specific relationships of inflammatory biomarkers with blood pressure in older adults

Emerging evidence indicates an association between blood pressure and inflammation, yet this relationship remains unclear in older adults, despite the elevated prevalence of hypertension. We investigated the association between blood pressure, high sensitivity C-reactive protein (hs-CRP), interleukin-6 (IL-6), and white blood cell (WBC) count in a cohort of 3571 older adults aged 65 and above, and 587 middle-aged participants (55–59 years old). In women aged 65 and above, the relationship between inflammatory markers and blood pressure was consistent, with hs-CRP and WBC emerging as predictors of high blood pressure. For hs-CRP, the adjusted odds ratio (OR) was 1.5 (95% CI, 1.07 to 2.10, P = 0.02), and for WBC, the adjusted OR was 1.41 (95% CI, 1.02 to 1.94, P = 0.04), comparing the highest to the lowest quartiles. In men, only the WBC count was significantly associated with an increased OR for high BP (adjusted OR 1.49, 95% CI, 1.09 to 2.02, P = 0.01) across quartiles. Across the entire study population, in a fully adjusted model, all inflammatory markers were modestly associated with blood pressure levels, while the effect of being over 65 years was the most significant predictor of high blood pressure (OR 1.84, 95% CI, 1.50 to 2.25, P < 0.001). The link between key inflammation markers and blood pressure in older adults varies by sex and biomarker type and may differ from the relationship observed in younger individuals. These relationships are likely to be affected by factors linked to age

Circulating cytokines and risk of developing hypertension: a systematic review and meta-analysis

Background: Immune responses play a significant role in hypertension, though the importance of key inflammatory mediators remains to be defined. We used a systematic literature review and meta-analysis to study the associations between key cytokines and incident hypertension. Methods: We performed a systematic search of Pubmed/Medline, Embase, Web of Science, and the Cochrane Central Register of Controlled Trials (CENTRAL), for peer-reviewed studies published up to August 2022. Incident hypertension was defined as systolic blood pressure ≥140 mmHg or diastolic blood pressure ≥90 mmHg and/or the use of antihypertensive medications. Random effects meta-analyses were used to calculate pooled hazard ratios (HRs)/risk ratios (RRs) and 95% confidence intervals by cytokine levels (highest vs. lowest quartile). Results: Only IL-6 and IL-1β levels have evidence allowing for quantitative evaluation concerning the onset of hypertension. Six studies (10,406 participants, 2,932 incident cases) examined the association of IL-6 with incident hypertension. The highest versus lowest quartile of circulating IL-6 was associated with a significant HR/RR of hypertension (1.61, 95% CI: 1.00 to 2.60; I2=87%). After adjusting for potential confounders, including body mass index (BMI), HR/RR was no longer significant (HR/RR: 1.24; 95% CI, 0.96 to 1.61; I2= 56%). About IL-1β, neither the crude (HR/RR: 1.03; 95% CI, 0.60 to 1.76; n=2) nor multivariate analysis (HR/RR: 0.97, 95% CI, 0.60 to 1.56; n=2) suggested a significant association with the risk of developing hypertension. Conclusions: A limited number of studies suggest that higher IL-6, but not IL-1β, might be associated with the development of hypertension

Acute Delta Hepatitis in Italy spanning three decades (1991–2019): Evidence for the effectiveness of the hepatitis B vaccination campaign

Updated incidence data of acute Delta virus hepatitis (HDV) are lacking worldwide. Our aim was to evaluate incidence of and risk factors for acute HDV in Italy after the introduction of the compulsory vaccination against hepatitis B virus (HBV) in 1991. Data were obtained from the National Surveillance System of acute viral hepatitis (SEIEVA). Independent predictors of HDV were assessed by logistic-regression analysis. The incidence of acute HDV per 1-million population declined from 3.2 cases in 1987 to 0.04 in 2019, parallel to that of acute HBV per 100,000 from 10.0 to 0.39 cases during the same period. The median age of cases increased from 27 years in the decade 1991-1999 to 44 years in the decade 2010-2019 (p < .001). Over the same period, the male/female ratio decreased from 3.8 to 2.1, the proportion of coinfections increased from 55% to 75% (p = .003) and that of HBsAg positive acute hepatitis tested for by IgM anti-HDV linearly decreased from 50.1% to 34.1% (p < .001). People born abroad accounted for 24.6% of cases in 2004-2010 and 32.1% in 2011-2019. In the period 2010-2019, risky sexual behaviour (O.R. 4.2; 95%CI: 1.4-12.8) was the sole independent predictor of acute HDV; conversely intravenous drug use was no longer associated (O.R. 1.25; 95%CI: 0.15-10.22) with this. In conclusion, HBV vaccination was an effective measure to control acute HDV. Intravenous drug use is no longer an efficient mode of HDV spread. Testing for IgM-anti HDV is a grey area requiring alert. Acute HDV in foreigners should be monitored in the years to come