Dendritic cells (DCs) are antigen presenting cells (APCs) that belongs a diversify group of cells, being still the main cells that can presents the antigen and activate T lymphocytes by inducing different kinds of cytokine production, which can influence the development of different profiles of lymphocytes. The production of IL-12p70 leads to the Th1 profile differentiation, furthermore the IL-23 production can be important for the establishment and maintenance of Th17 profile. Thus, IL-12 and IL-23 play an important role in the induction or establishment of the adaptive immune response. The aim of the study was characterize the DC AP284 cell line by checking its surface markers, TLRs expression and capacity of response to different TLRs agonists. Besides of that, the production of IL-12p40, IL-12p70, IL-23 and the ability to induce Th1 or Th17 profile were evaluated. AP284 cells have expressed surface molecules compatible with DCs including class II MHC, CD11c and 33D1. After stimulation using different TLRs agonists these cells produced higher amounts of de IL-12p40 and IL-23, but no IL-12p70 production was observed. The IFN-γ addition was not able to induce IL-12p70 production, instead of that an inhibition of the IL-12p40 and IL-23 production were observed. The results have also shown that AP284 cell line is related with the Th17 profile because higher amounts of IL-12p40 and IL-23, but not IL-12p70 were observed, besides of that the results showed an increase in IL-17 levels produced by mice lymph node cells after immunization with Freund’s adjuvant and BCG. Thus, AP284 cells can be important to study the mechanisms of induction and regulation related with the production of IL-12p40 and IL-23 working as a possible tool to study the establishment and maintenance of Th17 cells profile. Moreover, AP284 cells have some similar characteristics among the other immortalized murine DCs, but the AP284 line produces a much higher amount of IL-12p40 than all described cells and is also differentiated among them by the production of IL-23.

Dendritic cells (DCs) are a heterogeneous group of cells and the major activators of naive T lymphocytes. During antigen presentation, different types of cytokines are produced and they will interfere with the immune response profile generated. IL-12p70 promotes the differentiation of Th1 phenotype and IL-23 p enhances the Th17 response. Given the major role of IL-12 and IL-23 on the stabilization of the acquired immune response, the production of these cytokines is highly controlled and it is an issue for several studies. The objective of this work was to characterize a new lineage of DC, described as AP284, based on their surface markers and evaluate the ability of these cells to produce IL-12p40, IL-12p70 and IL-23 after different stimulus in vitro. Additionally it will be evaluate their phagocytic capacity and their ability to induce a Th1 and Th17 response in vivo. The expression of the markers was analyzed by flow cytometry and the quantitation of IL-12 and IL23 was performed by ELISA after stimulation of cells AP284 in vitro with lipopolysaccharide (LPS) and Escherichia coli. AP284 cells express MHC class II, CD11c and 33D1 on its surface, which are characteristic markers of DCs. After stimulus with LPS or E.coli, AP284 cells produce large amounts of IL-12p40 and IL-23 but do not produce IL-12p70. The production of IL-12p70 was not detected even when IFN-γ is added to prime cultures, moreover, priming the cells with IFN-γ promoted inhibition of IL-12p40 and IL-23. Our data sugest that the AP284 is a DC17 lineage. Thus, AP284 can be an important tool to study the mechanisms of induction or regulation of IL-23 production and a possible tool to study generation and maintenance of a Th17 profile.

Dendritic cells (DCs) are specialized antigen-presenting cells that play a critical role in the immune response against human papillomavirus (HPV) infection, and represent a therapeutic target in cancer. Objective: To identify and quantify DCs in tonsillar squamous cell carcinoma (TSCC) under the influence of HPV infection. Methodology: CD1a and CD83 antibodies were used to identify immature dendritic cells and mature dendritic cells by immunohistochemistry in 33 primary TSCC and 10 normal tonsils (NTs), respectively. For the TSCC samples, the number of DCs per area was evaluated in the intra- and peritumoral compartments. For the NTs, the quantification of DCs was evaluated in the intra- and peritonsillar compartments. HPV detection methods were determined according to the ASCO Clinical Practice Guidelines from the College of American Pathologists Guideline (2018). Results: There were fewer intratumoral CD1a+ DCs in the HPV-positive and HPV-negative TSCC groups than in the NT group (p<0.05). In the peritumoral compartment, there were fewer CD83+ DCs in the HPV-positive and HPV-negative TSCC groups than in the NT group (p<0.001). The quantification of DCs subtypes showed no statistical differences between HPV-positive and HPV-negative TSCC groups (p>0.137). Patients with HPV-positive TSCC had significantly better overall survival rate than those with HPV-negative TSCC (p=0.004). Conclusion: Tumor activity contributes to DC depletion regardless of intralesional HPV positivity. An improved prognosis has been reported in patients with HPV-positive TSCC.

Plasmacytoid dendritic cells (pDCs) play a key role in systemic sclerosis (SSc) pathophysiology. However, despite the recognised importance of metabolic reprogramming for pDC function, their metabolic profile in SSc remains to be elucidated. Thus, our study aimed to explore the metabolic profile of pDCs in SSc and their potential contribution to the disease.

Immune cells detect specific microbes or damage to tissue integrity in order to initiate efficient immune responses. Abnormal accumulation of proteins in the endoplasmic reticulum (ER) can be seen as a sign of cellular malfunction and stress that triggers a collection of conserved emergency rescue programs. These different signaling cascades, which favor ER proteostasis and promote cell survival, are collectively known as the unfolded protein response (UPR). In recent years, a synergy between the UPR and inflammatory cytokine production has been unraveled, with different branches of the UPR entering in a cross-talk with specialized microbe sensing pathways, which turns on or amplify inflammatory cytokines production. Complementary to this synergetic activity, UPR induction alone, can itself be seen as a danger signal, and triggers directly or indirectly inflammation in different cellular and pathological models, this independently of the presence of pathogens. Here, we discuss recent advances on the nature of these cross-talks and how innate immunity, metabolism dysregulation, and ER-signaling pathways intersect in specialized immune cells, such as dendritic cells (DCs), and contribute to the pathogenesis of inflammatory diseases.

Mounting evidence has accumulated on the critical role of the different myeloid cells in the regulation of the cancerous process, and in particular in the modulation of the immune reaction to cancer. Myeloid cells are a major component of host cells infiltrating tumors, interacting with each other, with tumor cells and other stromal cells, and demonstrating a prominent plasticity. We describe here various myeloid regulatory cells (MRCs) in mice and human as well as their relevant therapeutic targets. We first address the role of the monocytes and macrophages that can contribute to angiogenesis, immunosuppression and metastatic dissemination. Next, we discuss the differential role of neutrophil subsets in tumor development, enhancing the dual and sometimes contradicting role of these cells. A heterogeneous population of immature myeloid cells, MDSCs, was shown to be generated and accumulated during tumor progression as well as to be an important player in cancer-related immune suppression. Lastly, we discuss the role of myeloid DCs, which can either contribute to effective anti-tumor responses or play a more regulatory role. We believe that MRCs play a critical role in cancer-related immune regulation and suggest that future anti-cancer therapies will focus on these abundant cells.

BACKGROUND: Blastic plasmacytoid dendritic cell neoplasm (BPDCN) is a rare hematopoietic tumor that was once believed to be derived from natural killer cells and is now recognized as originating from precursors of plasmacytoid dendritic cells. It generally involves the skin and has an aggressive clinical course. Due to its highly malignant behavior, a fast and accurate diagnosis of this condition is of the utmost importance. METHODS: Six cytology specimens from 5 patients diagnosed with BPDCN were reviewed as well as their clinical records. Findings were compared with 3 previously published cases. RESULTS: Two exfoliative cytology specimens (cerebrospinal fluid) and 4 fine-needle aspiration specimens (3 lymph node specimens and 1 cutaneous specimen) were reviewed. The cytomorphological aspects were similar in all cases. The smears were hypercellular with a monotonous population of intermediate-sized cells, dispersed singly or arranged in loose aggregates. The cells had round to oval nuclei, with fine chromatin and prominent nucleoli; the cytoplasm was generally scant, without visible granules. Intracytoplasmic microvacuoles were found in the majority of cases. Four cases were also characterized by flow cytometry, which revealed expression of CD 123, CD56 and/or CD4. Fine-needle aspiration was used in 1 case for primary diagnosis. Histological confirmation was available in all cases. CONCLUSIONS: BPDCN is a highly malignant neoplasm with a poor outcome. Cytology, in association with flow cytometry immunophenotyping and clinical history, is a reliable method with which to establish the diagnosis.

Introdução: O Lúpus Eritematoso Sistémico (LES) é uma doença auto-imune caracterizada por uma desregulação do sistema imune, que leva a uma produção aumentada de anticorpos e formação de imuno-complexos, resultando na produção de citocinas pró-inflamatórias e dano celular de diferentes órgãos. Objectivos: Este trabalho teve como objectivos a avaliação do efeito estimulatório provocado pelo soro de doentes com LES em monócitos e células dendríticas (DCs) do sangue periférico de controlos saudáveis e a quantificação de TNF-α no soro de doentes com LES e de um grupo controlo. Material e Métodos: Foram incluídos neste estudo 21 doentes com LES, agrupados de acordo com o índice de actividade da doença, em doentes com doença activa (LESA) - 11 e inactiva (LESI) - 11, bem como 10 indivíduos saudáveis que constituíram o grupo controlo. A expressão de TNF-α e IL-6 em monócitos, mDCs e DCs CD16+CD14-/low obtidas do sangue periférico de indivíduos saudáveis foi avaliada por citometria de fluxo, após estimulação in vitro com LPS e na presença de soro de doentes com LES e de um grupo controlo. O estudo da expressão génica de TNF-α foi realizado após separação das diferentes populações celulares em estudo por qRT-PCR. A concentração sérica de TNF-α foi determinada em 64 doentes com LES e 27 controlos, quantificada através da MILLIPLEX MAP Human TH17 Magnetic Bead Panel Premixed. Resultados: Observou-se que o soro de doentes com LES, particularmente aqueles com um maior título de ac. anti-dsDNA, provocava um aumento da expressão de TNF-α, nos monócitos e principalmente nas DCs CD16+CD14-/low. Também se observou um aumento da expressão de mRNA de TNF-α nesta última população celular, bem como um aumento da concentração sérica de TNF-α nos doentes com LES. Conclusão: Este estudo enfatiza o papel do ambiente inflamatório a nível periférico nos doentes com LES e a sua importância na activação das células circulantes.

Dendritic cells (DCs) are professional antigen-presenting cells and are extremely important in activating acquired immunity, as they can modulate various specific T and B cell responses. There are several subtypes of dendritic cells, which allow the body to generate different responses to each type of invading agent, through different activation cascades depending on the type of response that is intended to be generated, by the presentation of antigens through the major histocompatibility complex 1, which specializes in activating cytotoxic CD8+ T ,cells, normally, by the presentation of intracelular antigens, or by major histocompatibility complex 2, which activates effector CD4+ T cells by presenting antigens captured externally by antigen presenting cells.In addition, dendritic cells present the function to trigger a response to eliminate tumor cells, mainly by stimulating cytotoxic T lymphocytes with exogenous antigens with the ability to use crosspresentation. This ability to activate anti-tumor responses has enabled the development of immunotherapies using dendritic cells. This scientific review thesis aims to address the immunobiology of dendritic cells and their different subtypes and therapies involving these cells, with a special focus on in vivo therapy targeting DCs using monoclonal antibodies. This targeting strategy has been developed more recently and shows promising results in the fight against cancer while presenting advantages over the ex vivo use of dendritic cells.

Dendritic cells (DCs) are a heterogenous family of professional antigen presenting cells that specialize in the uptake, processing and presentation of antigens to T cells to promote downstream immune responses, therefore acting as orchestrators of immunity. Conventional type 1 DCs (cDC1) are cross-presenting cells, cDC type 2 (cDC2) cells govern type 1, 2 and 3 immunity, and plasmacytoid DCs (pDCs) are critical in type-I IFN (IFN-I) secretion for antiviral responses. However, the intricacy of DC diversity has become more apparent with recent studies highlighting the range and redundancy of some DC functions previously thought as restricted. In the context of cancer, the effectiveness of classical immunotherapy approaches, such as immune checkpoint blockade, has been associated with cDC1, but also with the presence of cDC2 and pDCs in the tumor microenvironment. Understanding how DC diversity is generated is crucial to predict and promote immune responses. However, the transcription factors (TFs) driving DC subset identity and functional divergence remain unclear. By allowing the direct conversion of a somatic cell type into another, direct cell reprogramming offers not only a strategy to generate cells for regenerative medicine or immunotherapy, but also a platform to dissect the TF codes underlying heterogeneous cell lineages. The minimal network composed of PU.1, IRF8, and BATF3 was shown to be sufficient to induce cDC1s from fibroblasts, opening opportunities to understand DC diversity with reprogramming. Here, I applied direct cell reprogramming to uncover the TF networks underlying the diversity of DCs, while also generating diverse DC subsets for cancer immunotherapy. Using an additive screening strategy, I identified two TF triads that induce cDC2 or pDC identities. PU.1, IRF4, and PRDM1 (PIP) induced cDC2s that acquired a pro-inflammatory cDC2B transcriptional program with an interferon gene signature and the ability to present antigens to T cells. SPIB, IRF8, and IKZF2 (SII) induced pDCs that displayed an immature pDC pro-inflammatory profile which could be primed to IFN-β secretion by the addition of IRF4. We further elucidated TF engagement at the onset of the reprogramming process, highlighting the need for cooperative action between an ETS and an IRF factor as the base program for DC induction, further aided by a third factor to establish subset-specific identities. The injection of all induced DCs into tumors promoted anti-tumor immunity in mice, solidifying their immunotherapeutic potential. In vivo reprogramming of two different tumor models revealed a differential response to cDC1 and cDC2 reprogramming, highlighting the specificity of each DC-reprogramming combination in driving tumor-protective responses. Additionally, I developed a combinatorial barcoded TF-based single-cell screening platform to further identify TF combinations for DC reprogramming. A set of 48 DC-specific barcoded TFs allowed the simultaneous induction of multiple DCs and other immune cell types and the construction of a TF hierarchy map to inform DC reprogramming. Collectively, these findings contribute to better understanding TF dynamics in DC specification, heterogeneity, and function, paving the way for advancing precision cancer immunotherapies based on DC reprogramming. Additionally, I developed a multiplexed technology that opens opportunities to harness immune cell reprogramming for broader applications, ranging from cancer to autoimmunity and beyond.