SARS-CoV-2 and its ORF3a, E and M viroporins activate inflammasome in human macrophages and induce of IL-1α in pulmonary epithelial and endothelial cells.

Inflammasome assembly is a potent mechanism responsible for the host protection against pathogens, including viruses. When compromised, it can allow viral replication, while when disrupted, it can perpetuate pathological responses by IL-1 signaling and pyroptotic cell death. SARS-CoV-2 infection was shown to activate inflammasome in the lungs of COVID-19 patients, however, potential mechanisms responsible for this response are not fully elucidated. In this study, we investigated the effects of ORF3a, E and M SARS-CoV-2 viroporins in the inflammasome activation in major populations of alveolar sentinel cells: macrophages, epithelial and endothelial cells. We demonstrated that each viroporin is capable of activation of the inflammasome in macrophages to trigger pyroptosis-like cell death and IL-1α release from epithelial and endothelial cells. Small molecule NLRP3 inflammasome inhibitors reduced IL-1 release but weakly affected the pyroptosis. Importantly, we discovered that while SARS-CoV-2 could not infect the pulmonary microvascular endothelial cells it induced IL-1α and IL-33 release. Together, these findings highlight the essential role of macrophages as the major inflammasome-activating cell population in the lungs and point to endothelial cell expressed IL-1α as a potential novel component driving the pulmonary immunothromobosis in COVID-19.

SARS-CoV-2 virus-like particle variants alpha and delta mimic the native viruses in their differential inflammasome activating potential.

The emerging SARS-CoV-2 variants are evolving to evade human immunity and differ in their pathogenicity. While evasion of the variants from adaptive immunity is widely investigated, there is a paucity of knowledge about their interactions with innate immunity. Inflammasome assembly is one of the most potent mechanisms of the early innate response to viruses, but when it is inappropriate, it can perpetuate tissue damage. In this study, we focused on the capacity of SARS-CoV-2 Alpha and Delta variants to activate the NLRP3 inflammasome. We compared the macrophage activation, particularly the inflammasome formation, using Alpha- and Delta-spike virus-like particles (VLPs). We found that VLPs of both variants activated the inflammasome even without a priming step. Delta-spike VLPs had a significantly stronger effect on triggering pyroptosis and inflammasome assembly in THP-1 macrophages than did Alfa-spike VLPs. Cells treated with Delta VLPs showed greater cleavage of caspase-1 and IL-1ß release. Furthermore, Delta VLPs induced stronger cytokine secretion from macrophages and caused essential impairment of mitochondrial respiration in comparison to Alpha VLPs. Additionally, infection of primary human monocyte-derived macrophages with the SARS-CoV-2 variants confirmed the observations in VLPs. Collectively, we revealed that SARS-CoV-2 Delta had a greater impact on the inflammasome activation, cell death and mitochondrial respiration in macrophages than did the Alpha variant. Importantly, the differential response to the SARS-CoV-2 variants can influence the efficacy of therapies targeting the host's innate immunity.

Anti-inflammatory and antioxidant actions of extracts from Rheum rhaponticum and Rheum rhabarbarum in human blood plasma and cells in vitro.

Rheum rhaponticum L. (rhapontic rhubarb) and Rheum rhabarbarum L. (garden rhubarb) are edible and medicinal rhubarb species used for many centuries in traditional medicine. This work is focused on the biological activity of extracts from petioles and roots of R. rhaponticum and R. rhabarbarum as well as rhapontigenin and rhaponticin, typical stilbenes present in these rhubarbs, in a context of their effects on blood physiology and cardiovascular health. Anti-inflammatory properties of the examined substances were evaluated in human peripheral blood mononuclear cells (PBMCs) and THP1-ASC-GFP inflammasome reporter cells. Due to the coexistence of inflammation and oxidative stress in cardiovascular diseases, the study design included also antioxidant assays. This part of the work involved the assessment of the protective efficiency of the examined substances against the peroxynitrite-triggered damage to human blood plasma components, including fibrinogen, a protein of critical importance for blood clotting and maintaining the haemostatic balance. Pre-incubation of PBMCs with the examined substances (1-50 µg/mL) considerably decreased the synthesis of prostaglandin E2 as well as the release of pro-inflammatory cytokines (IL-2 and TNF-α) and metalloproteinase-9. A reduced level of secreted apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) specks in the THP-1-ASC-GFP cells was also observed. The examined substances significantly diminished the extent of ONOO‾induced oxidative modifications of blood plasma proteins and lipids and normalized, or even strengthened blood plasma antioxidant capacity. Furthermore, a reduction of oxidative damage to fibrinogen, including modifications of tyrosine and tryptophan residues along with the formation of protein aggregates was found.

Identification of prion protein-derived peptides of potential use in Alzheimer's disease therapy.

Soluble form of the prion protein (PrP) has been previously shown to interact with amyloid-ß (Aß) peptides, suppressing their fibrillization as well as toxicity, which indicates that this protein may play a protective role in Alzheimer's disease (AD). The shortest known PrP fragment retaining all of these properties corresponds to physiologically generated proteolytic polypeptide PrP23-110/111, called N1. Here we have identified two N1-derived synthetic peptides, encompassing residues 23-50 (PrP23-50) and 90-112 (PrP90-112), which bind to Aß1-42 protofibrillar oligomers as well as amyloid fibrils. We found that, akin to N1, the abovementioned synthetic peptides not only reduce the initial rate of Aß fibrillization, but also alter the aggregation pathway of Aß, inhibiting formation of protofibrillar oligomers and facilitating amorphous aggregation. Furthermore, our data show that N1, PrP23-50 and PrP90-112 protect cultured hippocampal neurons from neurotoxic effects of Aß oligomers, preventing oligomers-induced retraction of neurites and loss of cell membrane integrity. The above PrP fragments can also attenuate neuronal intake of Aß. Our results strongly suggest that synthetic peptides such as PrP23-50 and PrP90-112 can be useful in designing a novel class of therapeutics in AD.

Effect of extended olanzapine administration on POMC and neuropeptide Y mRNA levels in the male rat amygdala and hippocampus.

BACKGROUND: Neuropeptides play an important role in various neural pathways, being able to control a wide spectrum of physiological responses. Neuropeptide Y (NPY) and proopiomelanocortin (POMC) functions are quite well studied, however little is known about their action at the level of limbic structures. The present work was focused on the expression of the aforementioned peptides in this brain structure of rats treated with olanzapine, a second generation neuroleptic drug. The detailed purpose of this experiment was the evaluation of potential relationships between chronic olanzapine administration and NPY and POMC mRNA expression in the amygdala and hippocampal formation. METHODS: The studies were carried out on adult, male Sprague-Dawley rats that were divided into 2 groups: control and experimental animals treated with olanzapine (28 day-long intraperitoneal injection). All individuals were sacrificed under anaesthesia, then the amygdaloid complexes and hippocampi were excised. Total mRNA was isolated from homogenized samples of both structures and the RT-PCR method was used for estimation of NPY and POMC gene relative expression. RESULTS: Prolonged olanzapine administration is reflected in qualitatively different changes in expression of NPY and POMC mRNA in the rat amygdala and hippocampus. Interestingly enough, olanzapine did not affect NPY expression, but significantly increased the POMC level in both examined regions. CONCLUSIONS: Olanzapine can affect amygdalar and hippocampal neuronal populations by the modulation of neuropeptide activity. Importantly, it may suggest the existence of an alternative mode of its action. Undoubtedly this hypothetic regulatory mechanism requires further pharmacological and neurostructural study.