The immune system of patients infected by SARS-CoV-2 is severely impaired. Detailed investigation of T cells and cytokine production in patients affected by COVID-19 pneumonia are urgently required. Here we show that, compared with healthy controls, COVID-19 patients' T cell compartment displays several alterations involving naïve, central memory, effector memory and terminally differentiated cells, as well as regulatory T cells and PD1 + CD57 + exhausted T cells. Significant alterations exist also in several lineage-specifying transcription factors and chemokine receptors. Terminally differentiated T cells from patients proliferate less than those from healthy controls, whereas their mitochondria functionality is similar in CD4 + T cells from both groups. Patients display significant increases of proinflammatory or anti-inflammatory cytokines, including T helper type-1 and type-2 cytokines, chemokines and galectins; their lymphocytes produce more tumor necrosis factor (TNF), interferon-γ, interleukin (IL)-2 and IL-17, with the last observation implying that blocking IL-17 could provide a novel therapeutic strategy for COVID-19.

We have deeply investigated T cell compartment, plasma cytokines and cells producing cytokines in patients affected by Covid-19. At admission, patients were lymphopenic; in all of them SARS-CoV-2 was detected in a nasopharyngeal swab specimen by real-time RT-PCR, and pneumonia was subsequently confirmed by X-rays.Detailed 18-parameter flow cytometry was performed in 21 patients and 13 controls. Coupling polychromatic cytometry with unsupervised data analysis, we found that patients show an increased amount of CD4+ T lymphocytes that were activated, exhausted, stem memory or Treg. Similar results concerning activation and exhaustion were found in the CD8+ T cell compartment, within which the differences were even greater.Measuring plasma level of 31 cytokines linked to inflammation revealed that Covid-19 showed a dramatic increase of several molecules, such as TH1 and TH2 cytokines, chemokines, galectins, pro- and anti-inflammatory mediators, confirming the importance of a massive immune activation causing the cytokine storm. Then, intracellular staining detecting the simultaneous production of different cytokines after a para-physiologic stimulus given by anti-CD3/CD28 mAbs revealed not only a high capacity to produce a variety of molecules, including TNF-a, IFN-g and IL-2, but also a significant skewing of CD4+ T cells towards the TH17 phenotype.A therapeutic approach now exists based on the administration of drugs that block IL-6 pathway, and is now consistently improving the course of the disease. IL-17 is crucial in recruiting and activating neutrophils, cells that can migrate to the lung and are heavily involved in the pathogenesis of Covid-19. We show here that a significant skewing of activated T cells towards TH17 functional phenotype exists in Covid-19 patients. Thus, we suggest that blocking IL-17 pathway by already available biological drugs that are used to treat different pathologies could be a novel, additional strategy to improve the health of patients infected by SARS-CoV-2.

In patients infected by SARS-CoV-2 who experience an exaggerated inflammation leading to pneumonia, monocytes likely play a major role but have received poor attention. Thus, we analyzed peripheral blood monocytes from patients with COVID-19 pneumonia and found that these cells show signs of altered bioenergetics and mitochondrial dysfunction, had a reduced basal and maximal respiration, reduced spare respiratory capacity, and decreased proton leak. Basal extracellular acidification rate was also diminished, suggesting reduced capability to perform aerobic glycolysis. Although COVID-19 monocytes had a reduced ability to perform oxidative burst, they were still capable of producing TNF and IFN-c in vitro. A significantly high amount of monocytes had depolarized mitochondria and abnormal mitochondrial ultrastructure. A redistribution of monocyte subsets, with a significant expansion of intermediate/pro-inflammatory cells, and high amounts of immature monocytes were found, along with a concomitant compression of classical monocytes, and an increased expression of inhibitory checkpoints like PD-1/PD-L1. High plasma levels of several inflammatory cytokines and chemokines, including GM-CSF, IL-18, CCL2, CXCL10, and osteopontin, finally confirm the importance of monocytes in COVID-19 immunopathogenesis.

Studies on the interactions between SARS‐CoV‐2 and humoral immunity are fundamental to elaborate effective therapies including vaccines. We used polychromatic flow cytometry, coupled with unsupervised data analysis and principal component analysis (PCA), to interrogate B cells in untreated patients with COVID‐19 pneumonia. COVID‐19 patients displayed normal plasma levels of the main immunoglobulin classes, of antibodies against common antigens or against antigens present in common vaccines. However, we found a decreased number of total and naïve B cells, along with decreased percentages and numbers of memory switched and unswitched B cells. On the contrary, IgM+ and IgM− plasmablasts were significantly increased. In vitro cell activation revealed that B lymphocytes showed a normal proliferation index and number of dividing cells per cycle. PCA indicated that B‐cell number, naive and memory B cells but not plasmablasts clustered with patients who were discharged, while plasma IgM level, C‐reactive protein, D‐dimer, and SOFA score with those who died. In patients with pneumonia, the derangement of the B‐cell compartment could be one of the causes of the immunological failure to control SARS‐Cov2, have a relevant influence on several pathways, organs and systems, and must be considered to develop vaccine strategies.

Triterpenoids are natural compounds synthesized by plants through cyclization of squalene, known for their weak anti-inflammatory activity. 2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oic acid (CDDO), and its C28 modified derivative, methyl-ester (CDDO-Me, also known as bardoxolone methyl), are two synthetic derivatives of oleanolic acid, synthesized more than 20 years ago, in an attempt to enhance the anti-inflammatory behavior of the natural compound. These molecules have been extensively investigated for their strong ability to exert antiproliferative, antiangiogenic, and antimetastatic activities, and to induce apoptosis and differentiation in cancer cells. Here, we discuss the chemical properties of natural triterpenoids, the pathways of synthesis and the biological effects of CDDO and its derivative CDDO-Me. At nanomolar doses, CDDO and CDDO-Me have been shown to protect cells and tissues from oxidative stress by increasing the transcriptional activity of the nuclear factor (erythroid-derived 2)-like 2 (Nrf2). At doses higher than 100 nM, CDDO and CDDO-Me are able to modulate the differentiation of a variety of cell types, both tumor cell lines or primary culture cell, while at micromolar doses these compounds exert an anticancer effect in multiple manners; by inducing extrinsic or intrinsic apoptotic pathways, or autophagic cell death, by inhibiting telomerase activity, by disrupting mitochondrial functions through Lon protease inhibition, and by blocking the deubiquitylating enzyme USP7. CDDO-Me demonstrated its efficacy as anticancer drugs in different mouse models, and versus several types of cancer. Several clinical trials have been started in humans for evaluating CDDO-Me efficacy as anticancer and anti-inflammatory drug; despite promising results, significant increase in heart failure events represented an obstacle for the clinical use of CDDO-Me.

Cell death mechanisms are crucial to maintain an appropriate environment for the functionality of healthy cells. However, during viral infections, dysregulation of these processes can be present and can participate in the pathogenetic mechanisms of the disease. In this review, we describe some features of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and some immunopathogenic mechanisms characterizing the present coronavirus disease (COVID-19). Lymphopenia and monocytopenia are important contributors to COVID-19 immunopathogenesis. The fine mechanisms underlying these phenomena are still unknown, and several hypotheses have been raised, some of which assign a role to cell death as far as the reduction of specific types of immune cells is concerned. Thus, we discuss three major pathways such as apoptosis, necroptosis, and pyroptosis, and suggest that all of them likely occur simultaneously in COVID-19 patients. We describe that SARS-CoV-2 can have both a direct and an indirect role in inducing cell death. Indeed, on the one hand, cell death can be caused by the virus entry into cells, on the other, the excessive concentration of cytokines and chemokines, a process that is known as a COVID-19-related cytokine storm, exerts deleterious effects on circulating immune cells. However, the overall knowledge of these mechanisms is still scarce and further studies are needed to delineate new therapeutic strategies.

To better understand the mechanisms at the basis of neutrophil functions during SARS‐CoV‐2, we studied patients with severe COVID‐19 pneumonia. They had high blood proportion of degranulated neutrophils and elevated plasma levels of myeloperoxidase (MPO), elastase, and MPO‐DNA complexes, which are typical markers of neutrophil extracellular traps (NET). Their neutrophils display dysfunctional mitochondria, defective oxidative burst, increased glycolysis, glycogen accumulation in the cytoplasm, and increase glycogenolysis. Hypoxia‐inducible factor 1α (ΗΙF‐1α) is stabilized in such cells, and it controls the level of glycogen phosphorylase L (PYGL), a key enzyme in glycogenolysis. Inhibiting PYGL abolishes the ability of neutrophils to produce NET. Patients displayed significant increases of plasma levels of molecules involved in the regulation of neutrophils’ function including CCL2, CXCL10, CCL20, IL‐18, IL‐3, IL‐6, G‐CSF, GM‐CSF, IFN‐γ. Our data suggest that metabolic remodelling is vital for the formation of NET and for boosting neutrophil inflammatory response, thus, suggesting that modulating ΗΙF‐1α or PYGL could represent a novel approach for innovative therapies.

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