SARS-CoV-2 infection as a model to study the effect of cinnamaldehyde as adjuvant therapy for viral pneumonia.

BACKGROUND: The recent pandemic outbursts, due to SARS-CoV-2, have highlighted once more the central role of the inflammatory process in the propagation of viral infection. The main consequence of COVID-19 is the induction of a diffuse pro-inflammatory state, also defined as a cytokine storm, which affects different organs, but mostly the lungs. We aimed to prove the efficacy of cinnamaldehyde, the active compound of cinnamon, as an anti-inflammatory compound, able to reduce SARS-CoV-2 induced cytokine storm. RESULTS: We enrolled 53 COVID-19 patients hospitalized for respiratory failure. The cohort was composed by 39 males and 13 females, aged 65.0 ± 9.8 years. We reported that COVID-19 patients have significantly higher IL-1ß and IL-6 plasma levels compared to non-COVID-19 pneumonia patients. In addition, human mononuclear cells (PBMCs) isolated from SARS-CoV-2 infected patients are significantly more prone to release pro-inflammatory cytokines upon stimuli. We demonstrated, using in vitro cell models, that macrophages are responsible for mediating the pro-inflammatory cytokine storm while lung cells support SARS-CoV-2 replication upon viral infection. In this context, cinnamaldehyde administration significantly reduces SARS-CoV-2-related inflammation by inhibiting NLRP3 mediated IL-1ß release in both PBMCs and THP-1 macrophages, as well as viral replication in CaLu-3 epithelial cells. Lastly, aerosol-administered cinnamaldehyde was able to significantly reduce IL-1ß release in an in vivo lung-inflammatory model. CONCLUSION: The obtained results suggest the possible use of cinnamaldehyde as a co-adjuvant preventive treatment for COVID-19 disease together with vaccination, but also as a promising dietary supplement to reduce, more broadly, viral induced inflammation.

Defective endoplasmic reticulum-mitochondria contacts and bioenergetics in SEPN1-related myopathy.

SEPN1-related myopathy (SEPN1-RM) is a muscle disorder due to mutations of the SEPN1 gene, which is characterized by muscle weakness and fatigue leading to scoliosis and life-threatening respiratory failure. Core lesions, focal areas of mitochondria depletion in skeletal muscle fibers, are the most common histopathological lesion. SEPN1-RM underlying mechanisms and the precise role of SEPN1 in muscle remained incompletely understood, hindering the development of biomarkers and therapies for this untreatable disease. To investigate the pathophysiological pathways in SEPN1-RM, we performed metabolic studies, calcium and ATP measurements, super-resolution and electron microscopy on in vivo and in vitro models of SEPN1 deficiency as well as muscle biopsies from SEPN1-RM patients. Mouse models of SEPN1 deficiency showed marked alterations in mitochondrial physiology and energy metabolism, suggesting that SEPN1 controls mitochondrial bioenergetics. Moreover, we found that SEPN1 was enriched at the mitochondria-associated membranes (MAM), and was needed for calcium transients between ER and mitochondria, as well as for the integrity of ER-mitochondria contacts. Consistently, loss of SEPN1 in patients was associated with alterations in body composition which correlated with the severity of muscle weakness, and with impaired ER-mitochondria contacts and low ATP levels. Our results indicate a role of SEPN1 as a novel MAM protein involved in mitochondrial bioenergetics. They also identify a systemic bioenergetic component in SEPN1-RM and establish mitochondria as a novel therapeutic target. This role of SEPN1 contributes to explain the fatigue and core lesions in skeletal muscle as well as the body composition abnormalities identified as part of the SEPN1-RM phenotype. Finally, these results point out to an unrecognized interplay between mitochondrial bioenergetics and ER homeostasis in skeletal muscle. They could therefore pave the way to the identification of biomarkers and therapeutic drugs for SEPN1-RM and for other disorders in which muscle ER-mitochondria cross-talk are impaired.

Corrigendum: Pioglitazone Improves Mitochondrial Organization and Bioenergetics in Down Syndrome Cells.

[This corrects the article DOI: 10.3389/fgene.2019.00606.].

Pioglitazone Improves Mitochondrial Organization and Bioenergetics in Down Syndrome Cells.

Mitochondrial dysfunction plays a primary role in neurodevelopmental anomalies and neurodegeneration of Down syndrome (DS) subjects. For this reason, targeting mitochondrial key genes, such as PGC-1α/PPARGC1A, is emerging as a good therapeutic approach to attenuate cognitive disability in DS. After demonstrating the efficacy of the biguanide metformin (a PGC-1α activator) in a cell model of DS, we extended the study to other molecules that regulate the PGC-1α pathway acting on PPAR genes. We, therefore, treated trisomic fetal fibroblasts with different doses of pioglitazone (PGZ) and evaluated the effects on mitochondrial dynamics and function. Treatment with PGZ significantly increased mRNA and protein levels of PGC-1α. Mitochondrial network was fully restored by PGZ administration affecting the fission-fusion mitochondrial machinery. Specifically, optic atrophy 1 (OPA1) and mitofusin 1 (MFN1) were upregulated while dynamin-related protein 1 (DRP1) was downregulated. These effects, together with a significant increase of basal ATP content and oxygen consumption rate, and a significant decrease of reactive oxygen species (ROS) production, provide strong evidence of an overall improvement of mitochondria bioenergetics in trisomic cells. In conclusion, we demonstrate that PGZ is able to improve mitochondrial phenotype even at low concentrations (0.5 µM). We also speculate that a combination of drugs that target mitochondrial function might be advantageous, offering potentially higher efficacy and lower individual drug dosage.

Melatonin as a master regulator of cell death and inflammation: molecular mechanisms and clinical implications for newborn care.

Melatonin, more commonly known as the sleep hormone, is mainly secreted by the pineal gland in dark conditions and regulates the circadian rhythm of the organism. Its intrinsic properties, including high cell permeability, the ability to easily cross both the blood-brain and placenta barriers, and its role as an endogenous reservoir of free radical scavengers (with indirect extra activities), confer it beneficial uses as an adjuvant in the biomedical field. Melatonin can exert its effects by acting through specific cellular receptors on the plasma membrane, similar to other hormones, or through receptor-independent mechanisms that involve complex molecular cross talk with other players. There is increasing evidence regarding the extraordinary beneficial effects of melatonin, also via exogenous administration. Here, we summarize molecular pathways in which melatonin is considered a master regulator, with attention to cell death and inflammation mechanisms from basic, translational and clinical points of view in the context of newborn care.

ß-Sitosterol Reduces the Expression of Chemotactic Cytokine Genes in Cystic Fibrosis Bronchial Epithelial Cells.

Extracts from Nigella arvensis L. seeds, which are widely used as anti-inflammatory remedies in traditional medicine of Northern Africa, were able to inhibit the expression of the pro-inflammatory neutrophil chemokine Interleukin (IL)-8 in Cystic Fibrosis (CF) bronchial epithelial IB3-1 cells exposed to the Gram-negative bacterium Pseudomonas aeruginosa. The chemical composition of the extracts led to the identification of three major components, ß-sitosterol, stigmasterol, and campesterol, which are the most abundant phytosterols, cholesterol-like molecules, usually found in plants. ß-sitosterol (BSS) was the only compound that significantly reproduced the inhibition of the P. aeruginosa-dependent expression of IL-8 at nanomolar concentrations. BSS was tested in CF airway epithelial CuFi-1 cells infected with P. aeruginosa. BSS (100 nM), showed a significant and consistent inhibitory activity on expression of the P. aeruginosa-stimulated expression chemokines IL-8, GRO-α GRO-ß, which play a pivotal role in the recruitment of neutrophils in CF inflamed lungs. Preliminary mechanistic analysis showed that BSS partially inhibits the P. aeruginosa-dependent activation of Protein Kinase C isoform alpha, which is known to be involved in the transmembrane signaling activating IL-8 gene expression in bronchial epithelial cells. These data indicate BSS as a promising molecule to control excessive lung inflammation in CF patients.

Intravital imaging reveals p53-dependent cancer cell death induced by phototherapy via calcium signaling.

One challenge in biology is signal transduction monitoring in a physiological context. Intravital imaging techniques are revolutionizing our understanding of tumor and host cell behaviors in the tumor environment. However, these deep tissue imaging techniques have not yet been adopted to investigate the second messenger calcium (Ca²âº). In the present study, we established conditions that allow the in vivo detection of Ca²âº signaling in three-dimensional tumor masses in mouse models. By combining intravital imaging and a skinfold chamber technique, we determined the ability of photodynamic cancer therapy to induce an increase in intracellular Ca²âº concentrations and, consequently, an increase in cell death in a p53-dependent pathway.

The endoplasmic reticulum-mitochondria connection: one touch, multiple functions.

The endoplasmic reticulum (ER) and mitochondria are tubular organelles with a characteristic "network structure" that facilitates the formation of interorganellar connections. The ER and mitochondria join together at multiple contact sites to form specific domains, termed mitochondria-ER associated membranes (MAMs), with distinct biochemical properties and a characteristic set of proteins. The functions of these two organelles are coordinated and executed at the ER-mitochondria interface, which provides a platform for the regulation of different processes. The roles played by the ER-mitochondria interface range from the coordination of calcium transfer to the regulation of mitochondrial fission and inflammasome formation as well as the provision of membranes for autophagy. The novel and unconventional processes that occur at the ER-mitochondria interface demonstrate its multifunctional and intrinsically dynamic nature. This article is part of a Special Issue entitled: Dynamic and ultrastructure of bioenergetic membranes and their components.

Subcellular calcium measurements in mammalian cells using jellyfish photoprotein aequorin-based probes.

The jellyfish Aequorea victoria produces a 22-kDa protein named aequorin that has had an important role in the study of calcium (Ca(2+)) signaling. Aequorin reacts with Ca(2+) via oxidation of the prosthetic group, coelenterazine, which results in emission of light. This signal can be detected by using a special luminescence reader (called aequorinometer) or luminescence plate readers. Here we describe the main characteristics of aequorin as a Ca(2+) probe and how to measure Ca(2+) in different intracellular compartments of animal cells (cytosol, different mitochondrial districts, nucleus, endoplasmic reticulum (ER), Golgi apparatus, peroxisomes and subplasma-membrane cytosol), ranging from single-well analyses to high-throughput screening by transfecting animal cells using DNA vectors carrying recombinant aequorin chimeras. The use of aequorin mutants and modified versions of coelenterazione increases the range of calcium concentrations that can be recorded. Cell culture and transfection takes ∼3 d. An experiment including signal calibration and the subsequent analyses will take ∼1 d.

[A new transportable machine for the preservation of livers to be transplanted by means of hyperbaric oxygenation perfusion]. / Una nuova macchina trasportabile per la conservazione mediante perfusione iperbarica ossigenata del fegato da trapiantare.

The preservation of livers to be transplanted is currently obtained by static cold storage at 4 C degrees and flushing with UW solution. New methods of preservation are being studied that take advantage of machines for continuous hypothermic perfusion of the organ. Such machines have permitted a lengthening of preservation times and the use of livers from non-beating-heart donors. In an attempt to eliminate the damage due to hypothermia, to lengthen preservation times, and to extend the availability of livers to be transplanted, also using those subjected to short periods of warm ischaemia, we have constructed a transportable machine that produces a hyperbaric atmosphere and allows continuous perfusion of the liver. Ten pig livers from beating-heart donors were perfused with Ringer solution in hyperbaric conditions with oxygen at temperatures ranging from 10 to 25 degrees C for periods of up to 24 hours and studied by means of histopathological analysis and tests of mitochondrial activity (FAU) in order to verify cell viability. The group of livers perfused up to 15 hours yielded an FAU value of 169.40 +/- 5.5 compared to the value of the non-perfused livers (controls) established as 100 and those perfused up to 24 hours had a FAU value of 139.18 +/- 10.7 compared to the controls established as 100, thus demonstrating cell viability. The viability of the organs after preservation with our procedure in the hyperbaric oxygenation perfusion machine gives us good reason to believe that, after appropriate further confirmation of the results, it will be possible to use the machine for the transplantation both of livers subjected to warm ischaemia and of livers preserved for longer periods than is currently the case.