The Complexity and Multiplicity of the Specific cAMP Phosphodiesterase Family: PDE4, Open New Adapted Therapeutic Approaches.

Cyclic nucleotides (cAMP, cGMP) play a major role in normal and pathologic signaling. Beyond receptors, cyclic nucleotide phosphodiesterases; (PDEs) rapidly convert the cyclic nucleotide in its respective 5'-nucleotide to control intracellular cAMP and/or cGMP levels to maintain a normal physiological state. However, in many pathologies, dysregulations of various PDEs (PDE1-PDE11) contribute mainly to organs and tissue failures related to uncontrolled phosphorylation cascade. Among these, PDE4 represents the greatest family, since it is constituted by 4 genes with multiple variants differently distributed at tissue, cellular and subcellular levels, allowing different fine-tuned regulations. Since the 1980s, pharmaceutical companies have developed PDE4 inhibitors (PDE4-I) to overcome cardiovascular diseases. Since, they have encountered many undesired problems, (emesis), they focused their research on other PDEs. Today, increases in the knowledge of complex PDE4 regulations in various tissues and pathologies, and the evolution in drug design, resulted in a renewal of PDE4-I development. The present review describes the recent PDE4-I development targeting cardiovascular diseases, obesity, diabetes, ulcerative colitis, and Crohn's disease, malignancies, fatty liver disease, osteoporosis, depression, as well as COVID-19. Today, the direct therapeutic approach of PDE4 is extended by developing allosteric inhibitors and protein/protein interactions allowing to act on the PDE interactome.

PDE4 inhibition as a therapeutic strategy for improvement of pulmonary dysfunctions in Covid-19 and cigarette smoking.

Angiotensin-converting enzyme 2 (ACE2) is the binding-site and entry-point for SARS-CoV-2 in human and highly expressed in the lung. Cigarette smoking (CS) is the leading cause of pulmonary and cardiovascular diseases. Chronic CS leads to upregulation of bronchial ACE2 inducing a high vulnerability in COVID-19 smoker patients. Interestingly, CS-induced dysregulation of pulmonary renin-angiotensin system (RAS) in part contributing into the potential pathogenesis COVID-19 pneumonia and acute respiratory distress syndrome (ARDS). Since, CS-mediated ACE2 activations is not the main pathway for increasing the risk of COVID-19, it appeared that AngII/AT1R might induce an inflammatory-burst in COVID-19 response by up-regulating cyclic nucleotide phosphodiesterase type 4 (PDE4), which hydrolyses specifically the second intracellular messenger 3', 5'-cyclic AMP (cAMP). It must be pointed out that CS might induce PDE4 up-regulation similarly to the COVID-19 inflammation, and therefore could potentiate COVID-19 inflammation opening the potential therapeutic effects of PDE4 inhibitor in both COVID-19-inflammation and CS.

Statins Therapy Improves Acute Ischemic Stroke in Patients with Cardio-metabolic Disorders Measured by Lipoprotein-Associated Phospholipase A2 (Lp-PLA2): New Focal Point.

BACKGROUND: Acute ischemic stroke (AIS) leads to neuronal and endothelial damage that activate the release of proinflammatory mediators such as lipoprotein-associated phospholipase A2 (Lp-PLA2), which lead to the development of brain edema injury. Most of statins produce differential effects on Lp-PLA2 activity and mass with a comparable reduction in low-density lipoprotein (LDL) serum levels. AIMS: The aim of this study is to evaluate the differential effect of different statins on the mass of level of Lp-PLA2 in patients with AIS. METHODS: A total of 69 patients with AIS aged 40-70 years compared with matched 39 healthy controls were involved in this case-control study. The AIS patients were divided according to the statins therapy into 39 patients on statins therapy (statins on), and 30 patients were not on the statins therapy (statins off). Anthropometric variables including weight, height, body mass index (BMI), and blood pressure profile were estimated. Besides, biochemical variables including lipid profile[total cholesterol (TC), triglyceride (TG), LDL, very low-density lipoprotein (VLDL), high-density lipoprotein (HDL)], Lp-PLA2 mass levels, high-sensitive C-reactive protein (Hs-CRP) were estimated. RESULTS: Patients with AIS had high Lp-PLA2 mass levels (P < 0.01) that positively correlated with high Hs-CRP, blood pressure, BMI, TC, TG, VLDL, LDL, and negatively correlated with HDL as compared with healthy controls. As well, statins on patients had lower Lp-PLA2 mass levels (9.82 ± 3.19 IU/mL) compared with statins off patients (16.55 ± 4.72 IU/mL), (P = 0.0001). Regarding the gender differences in the Lp-PLA2 mass level, it was higher in men patients with AIS compared with comparable females (P = 0.03). CONCLUSIONS: Lp-PLA2 mass level was higher in patients with AIS and linked with underlying poor cardio-metabolic disorders. Therefore, the Lp-PLA2 mass level is observed to be a surrogate biomarker of AIS mainly in patients with poor cardio-metabolic disorders. Statin therapy improves the Lp-PLA2 mass level and the poor cardio-metabolic profile in patients with AIS.

NCS 613, a PDE4 inhibitor, by increasing cAMP level suppresses systemic inflammation and immune complexes deposition in kidney of MRL/lpr lupus- prone mice.

Nephritis remains the most common severe manifestation of systemic lupus erythematosus in which auto-antibodies mediate chronic inflammation and kidney damage. cAMP-phosphodiesterases regulate sodium excretion and inflammation in various tissues. How cAMP elevation can reduce systemic inflammation and suppress kidney inflammation and damage remains elusive. PDE4 signaling and cAMP metabolism were investigated along immune complex depositions in target tissues and kidney damage (histology). SLE disease progression is associated with changes in kidney PDE4 activity and expression. Moreover, lupus prone mice exhibit low kidney cAMP level which is associated to induction and relocation of nuclear and cytoskeleton PDE4 isoforms. Auto-antibodies-induced kidney damage was attested by mesangial proliferation and cellular infiltration. Interestingly, we reported that NCS 613 treatment decreases systemic auto-antibody secretion and their corresponding immune complex deposition in target tissues. Furthermore, NCS 613 is able to increase cAMP levels in the kidney; hence this compound rescues kidney PDE4 alterations in treated mice. NCS 613 overcomes disease progression in lupus prone mice by improving wellbeing and decreasing inflammation in treated mice. The PDE4 inhibitor, NCS 613, is a new anti-inflammatory compound that is believed to be a leading drug candidate for the treatment of inflammatory diseases such as lupus nephritis.

La Société de Biologie de Strasbourg : 100 ans au service de la science et de la société.

Founded in 1919, the Society of Biology of Strasbourg (SBS) is a learned society whose purpose is the dissemination and promotion of scientific knowledge in biology. Subsidiary of the Society of Biology, the SBS celebrated its Centenary on Wednesday, the 16th of October 2019 on the Strasbourg University campus and at the Strasbourg City Hall. This day allowed retracing the various milestones of the SBS, through its main strengths, its difficulties and its permanent goal to meet scientific and societal challenges. The common thread of this day was the transmission of knowledge related to the past, the present, but also the future. At the start of the 21st century, the SBS must continue to reinvent itself to pursue its objective of transmitting scientific knowledge in biology and beyond. Scientific talks performed by senior scientists and former SBS thesis prizes awardees, a round table, and informal discussions reflected the history and the dynamism of the SBS association. All SBS Centennial participants have set the first milestone for the SBS Bicentennial.

TITLE: La Société de Biologie de Strasbourg : 100 ans au service de la science et de la société. ABSTRACT: Filiale de la Société de Biologie, la Société de Biologie de Strasbourg (SBS) est une société savante qui a pour objet la diffusion et la promotion du savoir scientifique en biologie et en médecine. Fondée en 1919, La SBS a célébré son Centenaire le mercredi 16 octobre 2019. Cette journée a permis de retracer les différents jalons de la SBS, à travers ses lignes de forces, ses difficultés et sa volonté permanente de mettre en exergue les défis scientifiques et sociétaux auxquels participent les recherches strasbourgeoises. Le fil rouge de cette journée a été la transmission d'un savoir en lien avec le passé, le présent, mais également le futur. En ce début du 21e siècle, la SBS se doit de continuer de se réinventer pour poursuivre son objectif de transmission des connaissances scientifiques en biologie et au-delà. L'ensemble des participants du Centenaire de la SBS a ainsi posé la première pierre du Bicentenaire de la SBS.

NCS 613, a Potent PDE4 Inhibitor, Displays Anti-Inflammatory and Anti-Proliferative Properties on A549 Lung Epithelial Cells and Human Lung Adenocarcinoma Explants.

Chronic inflammation is a deleterious process occurring in several pulmonary diseases; it is a driving force promoting tumorigenesis. By regulating local cyclic nucleotide concentration, cyclic nucleotide phosphodiesterases (PDE) govern important biological processes, including inflammation and proliferation. The aim of this study was to investigate the anti-inflammatory and anti-proliferative effects of NCS 613, a specific PDE4 inhibitor, on TNFα-treated human lung adenocarcinoma cell line (A549) and on human lung adenocarcinoma explants. PDE4 isoforms and inflammatory pathways mediated by p38 MAPK, ERK1/2, and IκBα were analyzed by Western blot and immunostainings. Proliferation were performed using [3H]-thymidine incorporation under different experimental conditions. TNFα-stimulation increased p38 MAPK phosphorylation and NF-κB translocation into the nucleus, which was abolished by NCS 613 treatment. Concomitantly, NCS 613 restores IκBα detection level in human adenocarcinoma. An IC50 value of 8.5 µM was determined for NCS 613 on anti-proliferative properties while ERK1/2 signaling was down-regulated in A549 cells and lung adenocarcinoma explants. These findings shed light on PDE4 signaling as a key regulator of chronic inflammation and cancer epithelial cell proliferation. It suggests that PDE4 inhibition by NCS 613 represent potential and interesting strategy for therapeutic intervention in tackling chronic inflammation and cell proliferation.

Hyperspectral Imaging Quantification of Mouse Limb Microcirculation Using an Ischemia Reperfusion Model with Phosphodiesterase 5 Inhibitor Preconditioning.

Background: Peripheral arterial disease has high incidence and complication rates. Vessel recanalization represents the main therapy. However, it induces reperfusion injury. Preconditioning with sildenafil has been advocated to protect against this injury. In this study, we show a real-time noninvasive quantitative assessment using hyperspectral imaging (HSI) of ischemia/reperfusion (IR) and analyzing the sildenafil effect. Materials and Methods: A one-sided hindlimb ischemia (120 minutes) followed by reperfusion (30 minutes) was created. Five mice received Sildenafil (1 mg/kg, i.p. twice before ischemia) and 5 mice served as control. The StO2 at T0, 5, 30, 60, 120 minutes after ischemia (T5, 30, 60, 120) and 5, 15, and 30 minutes after reperfusion (T125, 135, 150) were measured through HSI. Results: The control group showed a significantly lower StO2 at T120 (24.8% ± 17%) as compared with T0 (53.3% ± 7.04%) (P = .013) and T150 (76.8 ± 3.77; P = .0008). T150 showed a statistically significantly higher StO2 than T0 (P = .0134). In the sildenafil group, T120 StO2 (28.6% ± 20%) was lower than T0 (63.3% ± 8.46%; P = .0312) and T150 (73.3% ± 19.1%, P = .0075). The StO2 values did not differ statistically between sildenafil and control groups. Conclusions: HSI is a feasible tool to quantify both ischemia and reperfusion phases during lower limb IR. Preconditioning with sildenafil did not modify IR-related StO2 changes.

Cyclic nucleotide phosphodiesterases: New targets in the metabolic syndrome?

Metabolic diseases have a tremendous impact on human morbidity and mortality. Numerous targets regulating adenosine monophosphate kinase (AMPK) have been identified for treating the metabolic syndrome (MetS), and many compounds are being used or developed to increase AMPK activity. In parallel, the cyclic nucleotide phosphodiesterase families (PDEs) have emerged as new therapeutic targets in cardiovascular diseases, as well as in non-resolved pathologies. Since some PDE subfamilies inactivate cAMP into 5'-AMP, while the beneficial effects in MetS are related to 5'-AMP-dependent activation of AMPK, an analysis of the various controversial relationships between PDEs and AMPK in MetS appears interesting. The present review will describe the various PDE families, AMPK and molecular mechanisms in the MetS and discuss the PDEs/PDE modulators related to the tissues involved, thus supporting the discovery of original molecules and the design of new therapeutic approaches in MetS.

COVID-19 and Phosphodiesterase Enzyme Type 5 Inhibitors.

COVID-19 pathology is mainly associated to a pulmonary disease which sometimes might result in an uncontrollable storm related to inflammatory diseases which could be fatal. It is well known that phosphodiesterase enzyme type 5 inhibitors (PDE5Is), such as sildenafil, have been successfully developed for the treatment of pulmonary arterial hypertension; interestingly, more recently, it was shown that PDE5Is might be also anti-inflammatory. Therefore, it would be of interest to question about the use of PDE5Is to overcome the COVID-19 storm, as much as PDE5 is mainly present in the lung tissues and vessels.

The Endocrine Function of the Heart: Physiology and Involvements of Natriuretic Peptides and Cyclic Nucleotide Phosphodiesterases in Heart Failure.

Besides pumping, the heart participates in hydro-sodium homeostasis and systemic blood pressure regulation through its endocrine function mainly represented by the large family of natriuretic peptides (NPs), including essentially atrial natriuretic (ANP) and brain natriuretic peptides (BNP). Under normal conditions, these peptides are synthesized in response to atrial cardiomyocyte stretch, increase natriuresis, diuresis, and vascular permeability through binding of the second intracellular messenger's guanosine 3',5'-cyclic monophosphate (cGMP) to specific receptors. During heart failure (HF), the beneficial effects of the enhanced cardiac hormones secretion are reduced, in connection with renal resistance to NP. In addition, there is a BNP paradox characterized by a physiological inefficiency of the BNP forms assayed by current methods. In this context, it appears interesting to improve the efficiency of the cardiac natriuretic system by inhibiting cyclic nucleotide phosphodiesterases, responsible for the degradation of cGMP. Recent data support such a therapeutic approach which can improve the quality of life and the prognosis of patients with HF.

Design and synthesis of 3-aminophthalazine derivatives and structural analogues as PDE5 inhibitors: anti-allodynic effect against neuropathic pain in a mouse model.

Neuropathic pain is a chronic pain caused by a lesion or disease affecting the somatosensory nervous system. To date, no specific treatment has been developed to cure this pain. Antidepressants and anticonvulsant drugs are used, but they do not demonstrate universal efficacy, and they often cause detrimental adverse effects. Some studies highlighted the efficacy of sildenafil, a well-known inhibitor of phosphodiesterase 5 (PDE5, (IC50 = 3.3 nM)), in models of pain. Based on these results, we focused our attention on MY 5445, another known PDE5 inhibitor. Homologues, isosteres and structural analogues of MY 5445 were designed and all synthesized compounds were evaluated for their inhibitory activity toward PDE5. Selectivity profiles towards other PDE1-4 isoenzymes, water solubility and stability in acidic medium of the most potent PDE5 inhibitors were determined and the aminophthalazine 16h and its mimetic 41n (3-aminoindazole) were evaluated in comparison to MY 5445 (4b) in vivo in a model of neuropathic pain induced by sciatic nerve cuffing in mice (3 and 0.5 mg/kg, ip twice a day). Both compounds showed the same efficacy on neuropathic allodynia as MY 5445, and thus produced a significant relief of mechanical hypersensitivity after 12 days of treatment.

Effect of the Phosphodiesterase 5 Inhibitor Sildenafil on Ischemia-Reperfusion-Induced Muscle Mitochondrial Dysfunction and Oxidative Stress.

Lower-limb ischemia-reperfusion (IR) is frequent and associated with significant morbidity and mortality. Phosphodiesterase 5 inhibitors demonstrated antioxidant and beneficial effects in several organs submitted to IR, but their effects on muscle mitochondrial functions after lower-limb IR are unknown. Unilateral hindlimb IR (2 h tourniquet followed by 2 h reperfusion) without or with sildenafil (1mg/kg ip 30 minutes before ischemia) was performed in 18 mice. Maximal oxidative capacity (VMax), relative contribution of the mitochondrial respiratory chain complexes, calcium retention capacity (CRC)-a marker of apoptosis-and reactive oxygen species (ROS) production were determined using high-resolution respirometry, spectrofluorometry, and electron paramagnetic resonance in gastrocnemius muscles from both hindlimbs. IR significantly reduced mitochondrial VMax (from 11.79 ± 1.74 to 4.65 ± 1.11 pmol/s*mg wet weight (ww), p < 0.05, -50.2 ± 16.3%) and CRC (from 2.33 ± 0.41 to 0.84 ± 0.18 µmol/mg dry weight (dw), p < 0.05; -61.1 ± 6.8%). ROS tended to increase in the ischemic limb (+64.3 ± 31.9%, p = 0.08). Although tending to reduce IR-related ROS production (-42.4%), sildenafil failed to reduce muscle mitochondrial dysfunctions (-63.3 ± 9.2%, p < 0.001 and -55.2 ± 7.6% p < 0.01 for VMax, and CRC, respectively). In conclusion, lower limb IR impaired skeletal muscle mitochondrial function, but, despite tending to reduce ROS production, pharmacological preconditioning with sildenafil did not show protective effects.

Design, Synthesis, Pharmacological Evaluation and Vascular Effects of Delphinidin Analogues.

BACKGROUND: Among polyphenolic compounds suggested to prevent cardiovascular diseases (CVDs) and to explain the "French paradox", the anthocyanidin delphinidin (Dp) has been reported to support at least partly the vascular beneficial effects of dietary polyphenolic compounds including those from fruits and related products as red wine. It has also been highlighted that Dp interacts directly with the active site of estrogen receptor α (ERα), leading to activation of endothelial NO synthase (eNOS) pathway thus contributing to the prevention of endothelial dysfunction in mice aorta. However, anthocyanidins have very low bioavailability and despite a well described in vitro efficacy, the very high hydrophilicity and physicochemical instability of Dp might explain the lack of in vivo reported effects. OBJECTIVE: The aim of this study was to identify new Dp analogues with increased lipophilicity and vasorelaxation potential by a chemical modulation of its structure and to characterize the signaling pathway notably in relation with ERα signaling and nitric oxide (NO) production. METHOD: OCH3-substituted delphinidin analogues were obtained through the coupling of the corresponding acetophenones with substituted benzaldehydes. Prediction of resorption of the flavylium derivatives was performed with the calculated logP and induction of vasorelaxation was performed by myography on WT and ERαKO mice thoracic aorta rings and compared to Dp. NO production was evaluated in vitro on human primary endothelial cells. RESULTS: Eight Dp analogues were synthesized including four new flavylium derivatives. Two compounds (9 and 11) showed a strong increase of vasorelaxation potential and a theoretically increased bioavailability compared to Dp. Interestingly, 9 and 11 induced increased O2 - or NO endothelial production respectively and revealed a novel NO-dependent ERα-independent relaxation compared to Dp. We suggested that this mechanism may be at least in part supported by the inhibition of vascular cyclic nucleotide phosphodiesterase (PDEs). CONCLUSION: The current study demonstrated that pharmacomodulation of the Dp backbone by replacement of OH groups by OCH3 groups of the A and B rings led to the identification and characterization of two compounds (9 and 11) with enhanced physio-chemical properties that could be associated to higher permeability capability and pharmacological activity for the prevention of CVDs compared to Dp.

Therapeutic potentials of natural compounds acting on cyclic nucleotide phosphodiesterase families.

Intracellular cyclic AMP and/or cyclic GMP are characterized in the 1960th. These second messengers, hydrolysed specifically by cyclic nucleotide phosphodiesterase (PDE), play a major role in intracellular signalling. Natural products have been a rich source of drug discovery, Theophylline and Methylxanthine originated from tea leaves used for asthma treatment, whereas, Papaverine, a natural isoquinolein originated from Papaver somniferum traditionally used in impotency, altogether as caffeine where firstly described as PDE-inhibiting compounds. Since that time, the knowledge in PDE field has been drastically increased, allowing the design and development of new therapeutic drugs acting against different pathologies in the nanomolar range. During this period some natural compounds have been identified as PDE inhibitors and used in that context to investigate their therapeutic potential effects. The aim of this literature review is to point out the reported data and demonstrating the contribution of natural characterized molecules as PDE inhibitors in various pathologies that can open new fields of research for drug discovery, notably in epigenetic regulation.

Effects of cyclic nucleotide phosphodiesterases (PDEs) on mitochondrial skeletal muscle functions.

Mitochondria play a critical role in skeletal muscle metabolism and function, notably at the level of tissue respiration, which conduct muscle strength as well as muscle survival. Pathological conditions induce mitochondria dysfunctions notably characterized by free oxygen radical production disturbing intracellular signaling. In that way, the second messengers, cyclic AMP and cyclic GMP, control intracellular signaling at the physiological and transcription levels by governing phosphorylation cascades. Both nucleotides are specifically and selectively hydrolyzed in their respective 5'-nucleotide by cyclic nucleotide phosphodiesterases (PDEs), which constitute a multi-genic family differently tissue distributed and subcellularly compartmentalized. These PDEs are presently recognized as therapeutic targets for cardiovascular, pulmonary, and neurologic diseases. However, very few data concerning cyclic nucleotides and PDEs in skeletal muscle, specifically in mitochondria, are reported in the literature. The knowledge of PDE implication in mitochondrial signaling would be helpful for resolving critical mitochondrial dysfunctions in skeletal muscle.

Delphinidin Inhibits Tumor Growth by Acting on VEGF Signalling in Endothelial Cells.

The vasculoprotective properties of delphinidin are driven mainly by its action on endothelial cells. Moreover, delphinidin displays anti-angiogenic properties in both in vitro and in vivo angiogenesis models and thereby might prevent the development of tumors associated with excessive vascularization. This study was aimed to test the effect of delphinidin on melanoma-induced tumor growth with emphasis on its molecular mechanism on endothelial cells. Delphinidin treatment significantly decreased in vivo tumor growth induced by B16-F10 melanoma cell xenograft in mice. In vitro, delphinidin was not able to inhibit VEGFR2-mediated B16-F10 melanoma cell proliferation but it specifically reduced basal and VEGFR2-mediated endothelial cell proliferation. The anti-proliferative effect of delphinidin was reversed either by the MEK1/2 MAP kinase inhibitor, U-0126, or the PI3K inhibitor, LY-294002. VEGF-induced proliferation was reduced either by U-0126 or LY-294002. Under these conditions, delphinidin failed to decrease further endothelial cell proliferation. Delphinidin prevented VEGF-induced phosphorylation of ERK1/2 and p38 MAPK and decreased the expression of the transcription factors, CREB and ATF1. Finally, delphinidin was more potent in inhibiting in vitro cyclic nucleotide phosphodiesterases (PDEs), PDE1 and PDE2, compared to PDE3-PDE5. Altogether delphinidin reduced tumor growth of melanoma cell in vivo by acting specifically on endothelial cell proliferation. The mechanism implies an association between inhibition of VEGF-induced proliferation via VEGFR2 signalling, MAPK, PI3K and at transcription level on CREB/ATF1 factors, and the inhibition of PDE2. In conjunction with our previous studies, we demonstrate that delphinidin is a promising compound to prevent pathologies associated with generation of vascular network in tumorigenesis.

Angiotensin-converting enzyme inhibition prevents myocardial infarction-induced increase in renal cortical cGMP and cAMP phosphodiesterase activities.

We investigated whether myocardial infarction (MI) enhances renal phosphodiesterases (PDE) activities, investigating particularly the relative contribution of PDE1-5 isozymes in total PDE activity involved in both cGMP and cAMP pathways, and whether angiotensin-converting enzyme inhibition (ACEi) decreases such renal PDE hyperactivities. We also investigated whether ACEi might thereby improve atrial natriuretic peptide (ANP) efficiency. We studied renal cortical PDE1-5 isozyme activities in sham (SH)-operated, MI rats and in MI rats treated with perindopril (ACEi) 1 month after coronary artery ligation. Circulating atrial natriuretic peptide (ANP), its second intracellular messenger cyclic guanosine monophosphate (cGMP) and cGMP/ANP ratio were also determined. Cortical cGMP-PDE2 (80.3 vs. 65.1 pmol/min/mg) and cGMP-PDE1 (50.7 vs. 30.1 pmol/min/mg), and cAMP-PDE2 (161 vs. 104.1 pmol/min/mg) and cAMP-PDE4 (307.5 vs. 197.2 pmol/min/mg) activities were higher in MI than in SH rats. Despite increased ANP plasma level, ANP efficiency tended to be decreased in MI compared to SH rats. Perindopril restored PDE activities and tended to improve ANP efficiency in MI rats. One month after coronary ligation, perindopril treatment of MI rats prevents the increase in renal cortical PDE activities. This may contribute to increase renal ANP efficiency in MI rats.

Tumour growth inhibition and anti-angiogenic effects using curcumin correspond to combined PDE2 and PDE4 inhibition.

Vascular endothelial growth factor (VEGF) plays a major role in angiogenesis by stimulating endothelial cells. Increase in cyclic AMP (cAMP) level inhibits VEGF-induced endothelial cell proliferation and migration. Cyclic nucleotide phosphodiesterases (PDEs), which specifically hydrolyse cyclic nucleotides, are critical in the regulation of this signal transduction. We have previously reported that PDE2 and PDE4 up-regulations in human umbilical vein endothelial cells (HUVECs) are implicated in VEGF-induced angiogenesis and that inhibition of PDE2 and PDE4 activities prevents the development of the in vitro angiogenesis by increasing cAMP level, as well as the in vivo chicken embryo angiogenesis. We have also shown that polyphenols are able to inhibit PDEs. The curcumin having anti-cancer properties, the present study investigated whether PDE2 and PDE4 inhibitors and curcumin could have similar in vivo anti-tumour properties and whether the anti-angiogenic effects of curcumin are mediated by PDEs. Both PDE2/PDE4 inhibitor association and curcumin significantly inhibited in vivo tumour growth in C57BL/6N mice. In vitro, curcumin inhibited basal and VEGF-stimulated HUVEC proliferation and migration and delayed cell cycle progression at G0/G1, similarly to the combination of selective PDE2 and PDE4 inhibitors. cAMP levels in HUVECs were significantly increased by curcumin, similarly to rolipram (PDE4 inhibitor) and BAY-60-550 (PDE2 inhibitor) association, indicating cAMP-PDE inhibitions. Moreover, curcumin was able to inhibit VEGF-induced cAMP-PDE activity without acting on cGMP-PDE activity and to modulate PDE2 and PDE4 expressions in HUVECs. The present results suggest that curcumin exerts its in vitro anti-angiogenic and in vivo anti-tumour properties through combined PDE2 and PDE4 inhibition.

Cyclic GMP catabolism up-regulation in MRL/lpr lupus-prone mice is associated with organ remodeling.

Production of high titer of antibodies against nuclear components is a hallmark of systemic lupus erythematosus, an autoimmune disease characterized by the progressive chronic inflammation of multiple joints and organs. Organ damage and dysfunction such as renal failure are typical clinical features in lupus. Cell hypermetabolism and hypertrophy can accelerate organ dysfunction. In this study we focus on a specific murine model of lupus, the MRL/lpr strain, and investigated the role of cyclic guanosine monophosphate (cGMP) catabolism in organ remodeling of main target tissues (kidney, spleen and liver) in comparison with age-matched control mice. In MRL/lpr-prone mice, the cGMP-phosphodiesterase (PDE) activities were significantly increased in the kidney (3-fold, P<0.001), spleen (2-fold, P<0.001) and liver (1.6-fold, P<0.05). These raised activity levels were paralleled by both an increased activity of PDE1 in the kidney (associated with nephromegaly) and in the liver, and PDE2 in the spleen of lupus-prone mice. The up-regulation of PDE1 and PDE2 activities were associated with a decrease in intracellular cGMP levels. This underlines an alteration of cGMP-PDE signaling in the kidney, spleen and liver targeting different PDEs according to organs. In good agreement with these findings, a single intravenous administration to MRL/lpr mice of nimodipine (PDE1 inhibitor) but not of EHNA (PDE2 inhibitor) was able to significantly lower peripheral hypercellularity (P=0.0401), a characteristic feature of this strain of lupus-prone mice. Collectively, our findings are important for generating personalized strategies to prevent certain forms of the lupus disease as well as for understanding the role of PDEs and cGMP in the pathophysiology of lupus.

Endothelium-independent and endothelium-dependent vasorelaxation by a dichloromethane fraction from Anogeissus Leiocarpus (DC) Guill. Et Perr. (Combretaceae): possible involvement of cyclic nucleotide phosphodiesterase inhibition.

Many traditional medicinal herbs from Burkina Faso are used to treat arterial hypertension (HTA). Among them, Anogeissus leiocarpus (A. Leiocarpus) which is well known and widely used in Burkina traditional medicine. Herein we assess the effects of dichloromethane fraction from A. leiocarpus stem bark (ALF), selected as the most active on cyclic nucleotide phosphodiesterases (PDEs) and characterized its specificity towards purified vascular PDE1 to PDE5 isoenzymes and study its effects on a vascular model. ALF potently and preferentially inhibits (IC50=1.6 ± 0.6 µg/mL) the calmodulin-dependent phosphodiesterase PDE1, being mainly present in vascular smooth muscle and preferentially hydrolyses cGMP. In the same range (IC50 =2.8 ± 0.2 µg/ml) ALF inhibits PDE2, a cGMP-activated enzyme that is only present in endothelial cells and hydrolyses both cAMP and cGMP. PDE5, which specifically hydrolyses cGMP and which mainly contributes to cGMP hydrolysis is also potently inhibited by ALF (IC50=7.6 ± 3.5 µg/ml). The potencies of ALF on cAMP hydrolyzing isoenzymes was lesser, being more effective on PDE4 (IC50= 17.6 ± 3.5 µg/ml) than on PDE3 (60.9 ± 1.8 µg/ml). Since the major effect of ALF were against cGMP hydrolysis and since cGMP is implicated in endothelium-dependent relaxation, the endothelium-dependent vasorelaxation was studied on isolated porcine coronary arteries rings pre-contracted with U46619. The endothelium-dependent vasorelaxation is significantly inhibited by N(ω)-nitro-L-arginine (LNA 300 µmol/L, an inhibitor of endothelial NO synthase), but not affected by charybdotoxin (CTX, 100 nM) plus apamin (APA, 100 nM) (two inhibitors of EDHF-mediated responses). The combination of 4-aminopyridine (4-AP, 1 mmol/L, inhibitor of voltage-dependent potassium channels, Kv) plus baryum (Ba(2+), 30 µmol/L, inhibitor of the potassium channels with entering correction, Kir) plus ouabain (3 µmol/L, inhibitor of ATPase Na(+)/K(+) channels) partially inhibits endothelium-independent vasorelaxant effect. This endothelium-independent relaxant effect was also sensitive to combination of 1H-[1,2,4]-oxadiazole-[4,3-α]-quinoxalin1-one (ODQ, 10 µM, soluble guanylyl cyclase inhibitor) and N-[2-(p-Bromocinnamylamino)ethyl]-5-isoquinoline sulfonamide dihydrochloride (H89, 100 nM, Protein Kinase A inhibitor). Taken together, these results indicate that ALF is a powerful vasodilator modulated by the formation of NO from endothelium, but also act by directly relaxing the vascular smooth muscle cells, by inhibiting cGMP hydrolyzing PDEs (PDE1, PDE2 and PDE5) and to a lesser extend on cAMP degradation (PDE3 and PDE4), cAMP and cGMP being second messengers involved in vascular relaxation.