Adenosine A1 Receptor Deficiency Aggravates Extracellular Matrix Accumulation in Diabetic Nephropathy through Disturbance of Peritubular Microenvironment

Background. We previously observed that adenosine A1 receptor (A1AR) had a protective role in proximal tubular megalin loss associated with albuminuria in diabetic nephropathy (DN). In this study, we aimed to explore the role of A1AR in the fibrosis progression of DN. Methods. We collected DN patients’ samples and established a streptozotocin-induced diabetes model in wild-type (WT) and A1AR-deficient (A1AR-/-) mice. The location and expression of CD34, PDGFRβ, and A1AR were detected in kidney tissue samples from DN patients by immunofluorescent and immunohistochemical staining. We also analyzed the expression of TGFβ, collagen (I, III, and IV), α-SMA, and PDGFRβ using immunohistochemistry in WT and A1AR-/- mice. CD34 and podoplanin expression were analyzed by Western blotting and immunohistochemical staining in mice, respectively. Human renal proximal tubular epithelial cells (HK2) were cultured in medium containing high glucose and A1AR agonist as well as antagonist. Results. In DN patients, the expression of PDGFRβ was higher with the loss of CD34. The location of PDGFRβ and TGFβ was near to each other. The A1AR, which was colocalized with CD34 partly, was also upregulated in DN patients. In WT-DN mice, obvious albuminuria and renal pathological leisure were observed. In A1AR-/- DN mice, more severe renal tubular interstitial fibrosis and more extracellular matrix deposition were observed, with lower CD34 expression and pronounced increase of PDGFRβ. In HK2 cells, high glucose stimulated the epithelial-mesenchymal transition (EMT) process, which was inhibited by A1AR agonist. Conclusion. A1AR played a critical role in protecting the tubulointerstitial fibrosis process in DN by regulation of the peritubular microenvironment.

Clock genes Period1 and Period2 in the hippocampal CA1 mediate depression-like behaviors and rapid antidepressant response

Accumulated reports have indicated that circadian rhythm is closely related to the pathogenesis of major depressive disorder (MDD). Recently, adenosine has been identified to modulate circadian clock via adenosine A1 and A2A receptor signaling pathways. Cyclic AMP-response element binding protein (CREB) is a convergent point that plays a critical role in the pathogenesis of depression and is a downstream molecule of adenosine A1 receptor signaling pathway as an endpoint that can regulate the expression of circadian genes Period1 (Per1) and Period2 (Per2). However, whether Per mediates the development of MDD via CREB has not been elucidated. We used chronic unpredictable stress (CUS) to induce depression-like behaviors and found that it could induce decrease in p-CREB and PER1 levels in the hippocampal CA1 region in rats. Both depression-like behaviors and the decreased protein levels could be rapidly rescued by the administration of adenosine A1 receptor agonist 2-Choro-N6-cyclopentyladenosine (CCPA). Furthermore, knockdown of Per1 in hippocampal CA1 region could also induce depression-like behaviors, which could also be rescued by CCPA. Interestingly, Per2 knockdown in hippocampal CA1 region resulted in potential antidepressant-like effect. In addition, knockout of CRE sequence in the promoter regions of either Per1 or Per2 led to depression-like behaviors, which could not be rescued by CCPA. These results indicated that clock genes Per1 and Per2 play critical roles in the pathophysiology of depression and CRE sequences in the promoter regions of Per1 and Per2 may be a critical antidepressant target.

Activation of Adenosine A1 Receptor in Ischemic Stroke: Neuroprotection by Tetrahydroxy Stilbene Glycoside as an Agonist

Ischemic stroke is the main cause of death/disability, posing a great menace to human health. Though efforts to search for therapeutic drugs are ongoing, few of them have succeeded. Adenosine A1 receptor (A1R) activation could ameliorate ischemic injury, representing a very tempting target for stroke treatment. Tetrahydroxy stilbene glycoside (TSG), a potent antioxidant from the well-known Chinese herb Polygonum multiflorum Thunb., has been reported to have notable neuroprotective activities but the underlying mechanisms are elusive. This study investigated the mechanism of TSG focusing on A1R. TSG markedly decreased mortality, neurological deficit score, cerebral infarct size and brain water content of MCAO rats, and ameliorated the disorders in purine metabolism, energy metabolism and antioxidative defense system. TSG helped the survival of SH-SY5Y cells in OGD/R by alleviating oxidative stress and glutamate release, and by maintaining calcium homeostasis. TSG effects were abolished by A1R antagonist DPCPX. Docking and binding assays confirmed the binding of TSG with A1R. In addition, TSG upregulated the A1R level lowered by MCAO and OGD/R. The downstream signals of A1R activation, ERK1/2, HIF-1α and NF-κB contributed to the neuroprotection of TSG. Moreover, void of “well-known” cardiovascular side effects of classical A1R agonists, TSG showcased its great potential for stroke treatment.

Effects of Levetiracetam in Acute and Kindling Model of Epilepsy Through Adenosinergic Pathway in Mice: Possible Involvement of Adenosine A1 Receptor

Objective: The present study aimed to explore the possible levetiracetam mechanisms of action in the adenosine signaling systems using the PTZ-induced acute seizure and the PTZ-induced kindling model of epileptogenesis.Method: In acute model, male mice received caffeine (non-specific adenosine receptor antagonist), or dipropylcyclopentylxanthine (DPCPX) (specific A1 receptor antagonist) prior to levetiracetam. After 30 minutes, a convulsant dose of PTZ (100 mg/kg) was administered to determine whether caffeine or DPCPX have any antagonistic effects on anti-seizure activity of levetiracetam by analyzing the onset of first myoclonic jerk (FMJ), generalized clonic seizures (GCS) and percent mortality.The chronic PTZ-induced kindling model was set to assess the gene expression changes in adenosine A1 receptor, inwardly-rectifying potassium channel (Kir3.2) and equilibrative nucleoside transporter-1 (ENT-1) through RT-qPCR. Data were analyzed using Origin statistical software version 8.5 and represented as Mean ± SEM.Results: In acute study, we found that caffeine (100 mg/kg) and DPCPX (25 mg/kg) reversed the anti-seizures effects of the levetiracetam significantly by reversing the percent protection and shortening the delay in the FMJ and onset of GCS in animals. In kindling model of epileptogenesis, it was found that levetiracetam increased the gene expression of adenosine A1 receptor andKir3.2 in the brain. Furthermore, levetiracetam significantly reduced the gene expression of ENT-1 in the brain that supposed to enhance the extracellular adenosine in the brain.Conclusion: Based on these results, it can be concluded that in addition to its action on SV2A vesicular protein, levetiracetam also prevent epileptogenesis by acting on the adenosine pathway in the CNS.