After more than one year since the COVID-19 outbreak, patients with severe disease still constitute the bottleneck of the pandemic management. Aberrant inflammatory responses, ranging from cytokine storm to immune-suppression, were described in COVID-19 and no treatment was demonstrated to change the prognosis significantly. Therefore, there is an urgent need for understanding the underlying pathogenic mechanisms to guide therapeutic interventions. This study was designed to assess myeloid cell activation and phenotype leading to recovery in patients surviving severe COVID-19. We evaluated longitudinally patients with COVID-19 related respiratory insufficiency, stratified according to the need of intensive care unit admission (ICU, n = 11, and No-ICU, n = 9), and age and sex matched healthy controls (HCs, n = 11), by flow cytometry and a wide array of serum inflammatory/immune-regulatory mediators. All patients featured systemic immune-regulatory myeloid cell phenotype as assessed by both unsupervised and supervised analysis of circulating monocyte and dendritic cell subsets. Specifically, we observed a reduction of CD14lowCD16+ monocytes, and reduced expression of CD80, CD86, and Slan. Moreover, mDCs, pDCs, and basophils were significantly reduced, in comparison to healthy subjects. Contemporaneously, both monocytes and DCs showed increased expression of CD163, CD204, CD206, and PD-L1 immune-regulatory markers. The expansion of M2-like monocytes was significantly higher at admission in patients featuring detectable SARS-CoV-2 plasma viral load and it was positively correlated with the levels of specific antibodies. In No-ICU patients, we observed a peak of the alterations at admission and a progressive regression to a phenotype similar to HCs at discharge. Interestingly, in ICU patients, the expression of immuno-suppressive markers progressively increased until discharge. Notably, an increase of M2-like HLA-DRhighPD-L1+ cells in CD14++CD16- monocytes and in dendritic cell subsets was observed at ICU discharge. Furthermore, IFN-γ and IL-12p40 showed a decline over time in ICU patients, while high values of IL1RA and IL-10 were maintained. In conclusion, these results support that timely acquisition of a myeloid cell immune-regulatory phenotype might contribute to recovery in severe systemic SARS-CoV-2 infection and suggest that therapeutic agents favoring an innate immune system regulatory shift may represent the best strategy to be implemented at this stage.

T helper-17 (Th17) cells are associated with inflammatory disorders and cancer. We report that environmental conditions resulting in cellular stress, such as low oxygen, glucose, and isotonic stress, particularly enhance the generation of Th17 cells. Pharmacological inhibition of cell stress reduces Th17 cell differentiation while stress inducers enhance the development of Th17 cells. The cellular stress response results in Th17 cell development via sustained cytoplasmic calcium levels and, in part, XBP1 activity. Furthermore, in an inflammatory environment, conditions resulting in cell stress can bring about de novo Th17 cell differentiation, even in the absence of transforming growth factor β (TGF-β) signaling. In vivo, cell stress inhibition enhances resistance to Th17-mediated autoimmunity while stress-exposed T cells enhance disease severity. Adverse metabolic environments during inflammation provide a link between adaptive immunity and inflammation and may represent a risk factor for the development of chronic inflammatory conditions by facilitating Th17 cell differentiation.

Tumor immune escape mechanisms are being regarded as suitable targets for tumor therapy. Among these, tryptophan catabolism plays a central role in creating an immunosuppressive environment, leading to tolerance to potentially immunogenic tumor antigens. Tryptophan catabolism is initiated by either indoleamine 2,3-dioxygenase (IDO-1/-2) or tryptophan 2,3-dioxygenase 2 (TDO2), resulting in biostatic tryptophan starvation and l-kynurenine production, which participates in shaping the dynamic relationship of the host's immune system with tumor cells. Current immunotherapy strategies include blockade of IDO-1/-2 or TDO2, to restore efficient antitumor responses. Patients who might benefit from this approach are currently identified based on expression analyses of IDO-1/-2 or TDO2 in tumor tissue and/or enzymatic activity assessed by kynurenine/tryptophan ratios in the serum. We developed a monoclonal antibody targeting l-kynurenine as an in situ biomarker of IDO-1/-2 or TDO2 activity. Using Tissue Micro Array technology and immunostaining, colorectal and breast cancer patients were phenotyped based on l-kynurenine production. In colorectal cancer l-kynurenine was not unequivocally associated with IDO-1 expression, suggesting that the mere expression of tryptophan catabolic enzymes is not sufficiently informative for optimal immunotherapy.

To investigate the importance of OX40 signals for physiological CD4(+) T-cell responses, an endogenous antigen-specific population of CD4(+) T cells that recognise the 2W1S peptide was assessed and temporal control of OX40 signals was achieved using blocking or agonistic antibodies (Abs) in vivo. Following infection with Listeria monocytogenes expressing 2W1S peptide, OX40 was briefly expressed by the responding 2W1S-specific CD4(+) T cells, but only on a subset that co-expressed effector cell markers. This population was specifically expanded by Ab-ligation of OX40 during priming, which also caused skewing of the memory response towards effector memory cells. Strikingly, this greatly enhanced effector response was accompanied by the loss of T follicular helper (TFH) cells and germinal centres. Mice deficient in OX40 and CD30 showed normal generation of TFH cells but impaired numbers of 2W1S-specific effector cells. OX40 was not expressed by 2W1S-specific memory cells, although it was rapidly up-regulated upon challenge whereupon Ab-ligation of OX40 specifically affected the effector subset. In summary, these data indicate that for CD4(+) T cells, OX40 signals are important for generation of effector T cells rather than TFH cells in this response to acute bacterial infection.

The recently discovered histone post-translational modification crotonylation connects cellular metabolism to gene regulation. Its regulation and tissue-specific functions are poorly understood. We characterize histone crotonylation in intestinal epithelia and find that histone H3 crotonylation at lysine 18 is a surprisingly abundant modification in the small intestine crypt and colon, and is linked to gene regulation. We show that this modification is highly dynamic and regulated during the cell cycle. We identify class I histone deacetylases, HDAC1, HDAC2, and HDAC3, as major executors of histone decrotonylation. We show that known HDAC inhibitors, including the gut microbiota-derived butyrate, affect histone decrotonylation. Consistent with this, we find that depletion of the gut microbiota leads to a global change in histone crotonylation in the colon. Our results suggest that histone crotonylation connects chromatin to the gut microbiota, at least in part, via short-chain fatty acids and HDACs.

AhR is a ligand-activated transcription factor that plays an important role in the innate and adaptive immune responses. In infection models, it has been associated with host responses that promote or inhibit disease progression. In pulmonary paracoccidioidomycosis, a primary fungal infection endemic in Latin America, immune protection is mediated by Th1/Th17 cells and disease severity with predominant Th2/Th9/Treg responses. Because of its important role at epithelial barriers, we evaluate the role of AhR in the outcome of a pulmonary model of paracoccidioidomycosis. AhR-/- mice show increased fungal burdens, enhanced tissue pathology and mortality. During the infection, AhR-/- mice have more pulmonary myeloid cells with activated phenotype and reduced numbers expressing indoleamine 2,3 dioxygenase 1. AhR-deficient lungs have altered production of cytokines and reduced numbers of innate lymphoid cells (NK, ILC3 and NCR IL-22). The lungs of AhR-/- mice showed increased presence Th17 cells concomitant with reduced numbers of Th1, Th22 and Foxp3+ Treg cells. Furthermore, treatment of infected WT mice with an AhR-specific antagonist (CH223191) reproduced the main findings obtained in AhR-/- mice. Collectively our data demonstrate that in pulmonary paracoccidioidomycosis AhR controls fungal burden and excessive tissue inflammation and is a possible target for antifungal therapy.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected patients with cancer show worse outcomes compared with patients without cancer. The humoral immune response (HIR) of patients with cancer against SARS-CoV-2 is not well characterized. To better understand it, we conducted a serological study of hospitalized patients with cancer infected with SARS-CoV-2.