P2X7 is a purinergic receptor involved in important physiological functions and pathological processes, such as inflammation, neurodegeneration, and pain. Despite its relevance, there is no selective antagonist useful in the treatment of diseases related to this receptor. In this context, research for a selective, safe, and potent antagonist compound that can be used in clinical therapy has been growing. In this work, we evaluated the potential antagonistic activity of three fungal extracts, namely, Vishniacozyma victoriae, Metschnikowia australis, and Ascomycota sp., which were discovered in a high‐throughput screening campaign to search for new antagonists for P2X7R from natural products. First, the IC₅₀ values of these fungal extracts were determined in J774.G8 (murine macrophage cell line) and U937 (human monocyte cell line) cells through dye uptake assays. The IC₅₀ values of V. victoriae were 2.6 and 0.92 μg/mL, M. australis has IC₅₀ values of 3.8 and 1.5 μg/mL, and Ascomycota sp. showed values of 2.1 and 0.67 μg/mL in J774.G8 and U937 cells, respectively. These extracts also significantly inhibited propidium iodide and Lucifer yellow uptake via P2X7R pore, P2X7R currents in electrophysiology, IL‐1β release, and the production of oxide nitric and reactive oxygen species. The extracts did not cause cytotoxicity within a period of 24 h. The results showed the promising antagonistic activity of these extracts toward P2X7R, thereby indicating that they can be future candidates for phytomedicines with potential clinical applicability.

P2X7 is a purinergic receptor physiologically activated by extracellular ATP. Its activation induces proinflammatory responses, including cytokine release, reactive oxygen species formation, and cell death. Previous in vivo experimental models demonstrated that P2X7 blockade has anti-inflammatory effects; however, there are no drugs used in clinical therapy that act on the P2X7 receptor. In the context of inflammatory diseases, nonsteroidal anti-inflammatory drugs (NSAIDs) are widely used as the first-line treatment; however, their major side effects include stomach ulcer formation, which increases patient morbidity and mortality. Here, we analyzed for the first time the analgesic and gastroprotective activities of three fungal extracts that showed antagonistic effects on P2X7 in vitro. The Antarctic fungal extracts obtained from Vishniacozyma victoriae, Metschnikowia australis, and Ascomycota sp. were tested via in vivo models of acute pain and ethanol-induced ulceration. These three extracts reduced paw licking by approximately 50 %, which is related to pain behavior, and reduced the number of stomach ulcers 3-7 times compared with the control (70 % ethanol), making them more efficient than the lansoprazole, an NSAID drug, and Brilliant Blue G (BBG), a known P2X7 antagonist, which only halves the number of ulcers. Furthermore, the extracts also protected the gastric mucosa and significantly reduced the levels of liver and renal enzymes compared with those in the ethanol group. Taken together, the fungal extracts presented both analgesic and possibly anti-inflammatory activities and had a protective effect on the gastric epithelium.

BACKGROUND Zika virus (ZIKV) was first identified in the Americas in 2015, with Brazil experiencing a major epidemic marked by teratogenic effects during early pregnancy. Ex vivo studies of microglia from fatal ZIKV-induced microcephaly cases revealed elevated levels of inducible nitric oxide synthase (iNOS) and nitric oxide (NO). Direct-acting antivirals (DAAs) have been proposed as potential ZIKV treatments. OBJECTIVES This study aimed to investigate the effects of two DAAs, simeprevir and sofosbuvir, on iNOS expression and NO production during in vitro ZIKV infection. METHODS Molecular docking analyses were performed to assess the interaction between DAAs and the iNOS enzyme. ZIKV-infected Vero CCL-81 cells were treated with simeprevir or sofosbuvir (10 µmol/L), and iNOS expression, NO production, and cytotoxicity were evaluated. FINDINGS Both DAAs demonstrated strong binding affinity to the iNOS active site. Compared with the untreated infected controls, treatment with simeprevir and sofosbuvir significantly upregulated iNOS expression and increased NO production by approximately 60%. No cytotoxic effects were observed under the experimental conditions. MAIN CONCLUSIONS Simeprevir and sofosbuvir not only exhibit antiviral properties but also may enhance the host antiviral response via modulation of the NO pathway, suggesting a dual mechanism of action against ZIKV.