RESUMO
Epstein-Barr virus (EBV) encodes a G protein-coupled receptor (GPCR) termed BILF1 that is essential for EBV-mediated immunosuppression and oncogenesis. BILF1 couples with inhibitory G protein (Gi), the major intracellular signaling effector for human chemokine receptors, and exhibits constitutive signaling activity; the ligand(s) for BILF1 are unknown. We studied the origins of BILF1's constitutive activity through structure determination of BILF1 bound to the inhibitory G protein (Gi) heterotrimer. The 3.2-Å resolution cryo-electron microscopy structure revealed an extracellular loop within BILF1 that blocked the typical chemokine binding site, suggesting ligand-autonomous receptor activation. Rather, amino acid substitutions within BILF1 transmembrane regions at hallmark ligand-activated class A GPCR "microswitches" stabilized a constitutively active BILF1 conformation for Gi coupling in a ligand-independent fashion. Thus, the constitutive activity of BILF1 promotes immunosuppression and virulence independent of ligand availability, with implications for the function of GPCRs encoded by related viruses and for therapeutic targeting of EBV.
Assuntos
Infecções por Vírus Epstein-Barr/imunologia , Herpesvirus Humano 4/imunologia , Fatores Imunológicos/imunologia , Receptores Acoplados a Proteínas G/imunologia , Proteínas Virais/imunologia , Animais , Sítios de Ligação/imunologia , Linhagem Celular , Quimiocinas/imunologia , Microscopia Crioeletrônica/métodos , Infecções por Vírus Epstein-Barr/virologia , Células HEK293 , Humanos , Ligação Proteica/imunologia , Células Sf9 , Transdução de Sinais/imunologiaRESUMO
The biological and functional heterogeneity between tumors-both across and within cancer types-poses a challenge for immunotherapy. To understand the factors underlying tumor immune heterogeneity and immunotherapy sensitivity, we established a library of congenic tumor cell clones from an autochthonous mouse model of pancreatic adenocarcinoma. These clones generated tumors that recapitulated T cell-inflamed and non-T-cell-inflamed tumor microenvironments upon implantation in immunocompetent mice, with distinct patterns of infiltration by immune cell subsets. Co-injecting tumor cell clones revealed the non-T-cell-inflamed phenotype is dominant and that both quantitative and qualitative features of intratumoral CD8+ T cells determine response to therapy. Transcriptomic and epigenetic analyses revealed tumor-cell-intrinsic production of the chemokine CXCL1 as a determinant of the non-T-cell-inflamed microenvironment, and ablation of CXCL1 promoted T cell infiltration and sensitivity to a combination immunotherapy regimen. Thus, tumor cell-intrinsic factors shape the tumor immune microenvironment and influence the outcome of immunotherapy.
Assuntos
Adenocarcinoma/terapia , Fatores Imunológicos/imunologia , Imunoterapia , Subpopulações de Linfócitos/imunologia , Linfócitos do Interstício Tumoral/imunologia , Neoplasias Pancreáticas/terapia , Microambiente Tumoral/imunologia , Adenocarcinoma/imunologia , Adenocarcinoma/patologia , Idoso , Idoso de 80 Anos ou mais , Animais , Linfócitos T CD8-Positivos/imunologia , Epigenômica , Feminino , Perfilação da Expressão Gênica , Humanos , Fatores Imunológicos/genética , Masculino , Camundongos , Pessoa de Meia-Idade , Neoplasias Pancreáticas/imunologia , Neoplasias Pancreáticas/patologia , Cultura Primária de Células , Neoplasias PancreáticasRESUMO
The efficacy of the anti-cancer immunomodulatory agent cyclophosphamide (CTX) relies on intestinal bacteria. How and which relevant bacterial species are involved in tumor immunosurveillance, and their mechanism of action are unclear. Here, we identified two bacterial species, Enterococcus hirae and Barnesiella intestinihominis that are involved during CTX therapy. Whereas E. hirae translocated from the small intestine to secondary lymphoid organs and increased the intratumoral CD8/Treg ratio, B. intestinihominis accumulated in the colon and promoted the infiltration of IFN-γ-producing γδT cells in cancer lesions. The immune sensor, NOD2, limited CTX-induced cancer immunosurveillance and the bioactivity of these microbes. Finally, E. hirae and B. intestinihominis specific-memory Th1 cell immune responses selectively predicted longer progression-free survival in advanced lung and ovarian cancer patients treated with chemo-immunotherapy. Altogether, E. hirae and B. intestinihominis represent valuable "oncomicrobiotics" ameliorating the efficacy of the most common alkylating immunomodulatory compound.
Assuntos
Ciclofosfamida/farmacologia , Streptococcus faecium ATCC 9790/imunologia , Fatores Imunológicos/imunologia , Neoplasias/tratamento farmacológico , Neoplasias/imunologia , Animais , Colo/imunologia , Colo/microbiologia , Memória Imunológica/imunologia , Imunoterapia/métodos , Interferon gama/imunologia , Intestino Delgado/imunologia , Intestino Delgado/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Monitorização Imunológica , Proteína Adaptadora de Sinalização NOD2/imunologia , Células Th1/imunologiaRESUMO
The majority of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected individuals remain paucisymptomatic, contrasting with a minority of infected individuals in danger of death. Here, we speculate that the robust disease resistance of most individuals is due to a swift production of type I interferon (IFNα/ß), presumably sufficient to lower the viremia. A minority of infected individuals with a preexisting chronic inflammatory state fail to mount this early efficient response, leading to a delayed harmful inflammatory response. To improve the epidemiological scenario, we propose combining: (i) the development of efficient antivirals administered early enough to assist in the production of endogenous IFNα/ß; (ii) potentiating early IFN responses; (iii) administering anti-inflammatory treatments when needed, but not too early to interfere with endogenous antiviral responses.
Assuntos
Antivirais/imunologia , COVID-19/imunologia , Fatores Imunológicos/imunologia , Interferon Tipo I/imunologia , SARS-CoV-2/imunologia , Enzima de Conversão de Angiotensina 2/imunologia , Enzima de Conversão de Angiotensina 2/metabolismo , Antivirais/metabolismo , Antivirais/uso terapêutico , COVID-19/virologia , Citocinas/imunologia , Citocinas/metabolismo , Humanos , Fatores Imunológicos/metabolismo , Fatores Imunológicos/uso terapêutico , Interferon Tipo I/metabolismo , SARS-CoV-2/efeitos dos fármacos , SARS-CoV-2/metabolismo , Serina Endopeptidases/imunologia , Serina Endopeptidases/metabolismo , Replicação Viral/efeitos dos fármacos , Replicação Viral/imunologia , Tratamento Farmacológico da COVID-19RESUMO
Immune recognition of pathogen-associated molecular patterns (PAMPs) by pattern recognition receptors often activates proinflammatory NF-κB signalling1. Recent studies indicate that the bacterial metabolite D-glycero-ß-D-manno-heptose 1,7-bisphosphate (HBP) can activate NF-κB signalling in host cytosol2-4, but it is unclear whether HBP is a genuine PAMP and the cognate pattern recognition receptor has not been identified. Here we combined a transposon screen in Yersinia pseudotuberculosis with biochemical analyses and identified ADP-ß-D-manno-heptose (ADP-Hep), which mediates type III secretion system-dependent NF-κB activation and cytokine expression. ADP-Hep, but not other heptose metabolites, could enter host cytosol to activate NF-κB. A CRISPR-Cas9 screen showed that activation of NF-κB by ADP-Hep involves an ALPK1 (alpha-kinase 1)-TIFA (TRAF-interacting protein with forkhead-associated domain) axis. ADP-Hep directly binds the N-terminal domain of ALPK1, stimulating its kinase domain to phosphorylate and activate TIFA. The crystal structure of the N-terminal domain of ALPK1 and ADP-Hep in complex revealed the atomic mechanism of this ligand-receptor recognition process. HBP was transformed by host adenylyltransferases into ADP-heptose 7-P, which could activate ALPK1 to a lesser extent than ADP-Hep. ADP-Hep (but not HBP) alone or during bacterial infection induced Alpk1-dependent inflammation in mice. Our findings identify ALPK1 and ADP-Hep as a pattern recognition receptor and an effective immunomodulator, respectively.
Assuntos
Açúcares de Adenosina Difosfato/imunologia , Burkholderia cenocepacia , Citosol , Imunidade Inata , Moléculas com Motivos Associados a Patógenos/imunologia , Proteínas Quinases/metabolismo , Yersinia pseudotuberculosis , Açúcares de Adenosina Difosfato/metabolismo , Animais , Infecções por Burkholderia/enzimologia , Infecções por Burkholderia/imunologia , Infecções por Burkholderia/patologia , Burkholderia cenocepacia/genética , Burkholderia cenocepacia/imunologia , Burkholderia cenocepacia/metabolismo , Sistemas CRISPR-Cas , Cristalografia por Raios X , Citocinas/biossíntese , Citosol/enzimologia , Citosol/imunologia , Dissacarídeos/metabolismo , Ativação Enzimática , Feminino , Edição de Genes , Fatores Imunológicos/imunologia , Fatores Imunológicos/metabolismo , Imunomodulação , Inflamação/enzimologia , Inflamação/imunologia , Inflamação/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Modelos Moleculares , NF-kappa B/metabolismo , Moléculas com Motivos Associados a Patógenos/metabolismo , Yersinia pseudotuberculosis/genética , Yersinia pseudotuberculosis/imunologia , Yersinia pseudotuberculosis/metabolismoRESUMO
Schistosomes are long lived, intravascular parasitic platyhelminths that infect >200 million people globally. The molecular mechanisms used by these blood flukes to dampen host immune responses are described in this review. Adult worms express a collection of host-interactive tegumental ectoenzymes that can cleave host signaling molecules such as the "alarmin" ATP (cleaved by SmATPDase1), the platelet activator ADP (SmATPDase1, SmNPP5), and can convert AMP into the anti-inflammatory mediator adenosine (SmAP). SmAP can additionally cleave the lipid immunomodulator sphingosine-1-phosphate and the proinflammatory anionic polymer, polyP. In addition, the worms release a barrage of proteins (e.g., SmCB1, SjHSP70, cyclophilin A) that can impinge on immune cell function. Parasite eggs also release their own immunoregulatory proteins (e.g., IPSE/α1, omega1, SmCKBP) as do invasive cercariae (e.g., Sm16, Sj16). Some schistosome glycans (e.g., LNFPIII, LNnT) and lipids (e.g., Lyso-PS, LPC), produced by several life stages, likewise affect immune cell responses. The parasites not only produce eicosanoids (e.g., PGE2, PGD2-that can be anti-inflammatory) but can also induce host cells to release these metabolites. Finally, the worms release extracellular vesicles (EVs) containing microRNAs, and these too have been shown to skew host cell metabolism. Thus, schistosomes employ an array of biomolecules-protein, lipid, glycan, nucleic acid, and more, to bend host biochemistry to their liking. Many of the listed molecules have been individually shown capable of inducing aspects of the polarized Th2 response seen following infection (with the generation of regulatory T cells (Tregs), regulatory B cells (Bregs) and anti-inflammatory, alternatively activated (M2) macrophages). Precisely how host cells integrate the impact of these myriad parasite products following natural infection is not known. Several of the schistosome immunomodulators described here are in development as novel therapeutics against autoimmune, inflammatory, and other, nonparasitic, diseases.
Assuntos
Proteínas de Helminto/imunologia , Fatores Imunológicos/imunologia , Schistosoma/imunologia , Esquistossomose/imunologia , Animais , HumanosRESUMO
The extent to which immune cell phenotypes in the peripheral blood reflect within-tumor immune activity prior to and early in cancer therapy is unclear. To address this question, we studied the population dynamics of tumor and immune cells, and immune phenotypic changes, using clinical tumor and immune cell measurements and single-cell genomic analyses. These samples were serially obtained from a cohort of advanced gastrointestinal cancer patients enrolled in a trial with chemotherapy and immunotherapy. Using an ecological population model, fitted to clinical tumor burden and immune cell abundance data from each patient, we find evidence of a strong tumor-circulating immune cell interaction in responder patients but not in those patients that progress on treatment. Upon initiation of therapy, immune cell abundance increased rapidly in responsive patients, and once the peak level is reached tumor burden decreases, similar to models of predator-prey interactions; these dynamic patterns were absent in nonresponder patients. To interrogate phenotype dynamics of circulating immune cells, we performed single-cell RNA sequencing at serial time points during treatment. These data show that peripheral immune cell phenotypes were linked to the increased strength of patients' tumor-immune cell interaction, including increased cytotoxic differentiation and strong activation of interferon signaling in peripheral T cells in responder patients. Joint modeling of clinical and genomic data highlights the interactions between tumor and immune cell populations and reveals how variation in patient responsiveness can be explained by differences in peripheral immune cell signaling and differentiation soon after the initiation of immunotherapy.
Assuntos
Comunicação Celular/imunologia , Imunoterapia/métodos , Neoplasias/imunologia , Neoplasias/terapia , Fenótipo , Microambiente Tumoral/imunologia , Regulação da Expressão Gênica , Humanos , Fatores Imunológicos/genética , Fatores Imunológicos/imunologia , Monócitos/imunologia , Análise de Sequência de RNA , Análise de Célula Única , Linfócitos T/imunologiaRESUMO
Hemocyanins are used as immunomodulators in clinical applications because they induce a strong Th1-biased cell-mediated immunity, which has beneficial effects. They are multiligand glycosylated molecules with abundant and complex mannose-rich structures. It remains unclear whether these structures influence hemocyanin-induced immunostimulatory processes in human APCs. We have previously shown that hemocyanin glycans from Concholepas concholepas (CCH), Fissurella latimarginata (FLH), and Megathura crenulata (KLH), participate in their immune recognition and immunogenicity in mice, interacting with murine C-type lectin receptors (CLRs). Here, we studied the interactions of these hemocyanins with two major mannose-binding CLRs on monocyte-derived human DCs: MR (mannose receptor) and DC-SIGN (DC-specific ICAM-3-grabbing nonintegrin). Diverse analyses showed that hemocyanins are internalized by a mannose-sensitive mechanism. This process was calcium dependent. Moreover, hemocyanins colocalized with MR and DC-SIGN, and were partly internalized through clathrin-mediated endocytosis. The hemocyanin-mediated proinflammatory cytokine response was impaired when using deglycosylated FLH and KLH compared to CCH. We further showed that hemocyanins bind to human MR and DC-SIGN in a carbohydrate-dependent manner with affinity constants in the physiological concentration range. Overall, we showed that these three clinically valuable hemocyanins interact with human mannose-sensitive CLRs, initiating an immune response and promoting a Th1 cell-driving potential.
Assuntos
Moléculas de Adesão Celular/imunologia , Células Dendríticas/imunologia , Hemocianinas/imunologia , Fatores Imunológicos/imunologia , Lectinas Tipo C/imunologia , Lectinas de Ligação a Manose/imunologia , Receptores de Superfície Celular/imunologia , Animais , Células CHO , Linhagem Celular Tumoral , Células Cultivadas , Cricetulus , Humanos , Imunidade Celular/imunologia , Imunização/métodos , Receptor de Manose , Monócitos/imunologia , Células U937RESUMO
During inflammatory processes, tissue environmental cues are influencing the immunoregulatory properties of tissue-resident mesenchymal stem/stromal cells (MSC). In this study, we elucidated one of the molecular and cellular responses of human MSC exposed to combinations of inflammatory cytokines. We showed that during multi-cytokine priming by TNF-α, IL-1ß, and IFN-γ, IL-1ß further augmented the well-established immunoregulatory activity induced by TNF-α/IFN-γ. On the molecular level, TNF-α and IL-1ß enhanced the expression of IFN-γ receptor (IFN-γR) via NF 'kappa-light-chain-enhancer' of activated B-cells (NF-κΒ) signaling. In turn, enhanced responsiveness to IFN-γ stimulation activated STAT5 and p38-MAPK signaling. This molecular feedback resulted in an increased IL-8 release and augmented recruitment of polymorphonuclear granulocytes (PMN). Our study suggests the possibility that responses of MSC to multi-cytokine priming regimens may be exploited therapeutically to fine-tune inflammatory activity in tissues. This study elucidates molecular mechanisms underlying the immunological priming of mesenchymal stromal cells (MSC) and their interaction with neutrophils.
Assuntos
Interferon gama/imunologia , Interleucina-1beta/imunologia , Células-Tronco Mesenquimais/imunologia , Neutrófilos/imunologia , Transdução de Sinais/imunologia , Fator de Necrose Tumoral alfa/imunologia , Adulto , Idoso , Células Cultivadas , Feminino , Humanos , Fatores Imunológicos/imunologia , Sistema de Sinalização das MAP Quinases/imunologia , Masculino , Pessoa de Meia-Idade , Infiltração de Neutrófilos/imunologiaRESUMO
Infiltrating and tissue-resident myeloid cells are essential regulators of innate and adaptive immunity. During inflammation, and in response to microbial products, these cells can adapt to microenvironmental conditions and acquire specialized functions, including phagocytosis and the production of proinflammatory cytokines. Such myeloid plasticity is driven, in part, by epigenetic dynamics that can sustain stable phenotypes after activation, and which may lead to maladaptive cell polarization states associated with inflammation and autoimmunity. Here, we review recent reports describing epigenetic mechanisms linked to such polarization states and innate immune memory (tolerance and training) in monocyte and macrophage lineages. We discuss how these mechanisms might be targeted to develop putative immunomodulatory tools that might be used to treat a variety of immune-mediated diseases.
Assuntos
Epigênese Genética/efeitos dos fármacos , Imunidade Inata/efeitos dos fármacos , Fatores Imunológicos/uso terapêutico , Memória Imunológica/efeitos dos fármacos , Células Mieloides/efeitos dos fármacos , Animais , Epigênese Genética/genética , Epigênese Genética/imunologia , Humanos , Imunidade Inata/imunologia , Fatores Imunológicos/imunologia , Memória Imunológica/imunologia , Células Mieloides/imunologiaRESUMO
As a cytokine, gamma-interferon (IFN-γ) is considered a key player in the fine-tuned orchestration of immune responses. The extreme cellular sensitivity to cytokines is attested by the fact that very few of these bioactive molecules per cell are enough to trigger cellular functions. These findings can, at least partially, explain how/why homeopathically-prepared cytokines, and especially micro-immunotherapy (MI) medicines, are able to drive cellular responses. We focused our fundamental research on a unitary MI preparation of IFN-γ, specifically employed at 4 CH, manufactured and impregnated onto sucrose-lactose pillules as all other MI medicines. We assessed the IFN-γ concentration in the medium after dilution of the IFN-γ (4 CH)-bearing pillules and we evaluated in vitro drug responses in a wide range of immune cells, and in endothelial cells. Our results showed that IFN-γ (4 CH) stimulated the proliferation, the activation and the phagocytic capabilities of primary immune cells, as well as modulated their cytokine-secretion and immunity-related markers' expression in a trend that is quite comparable with the well-recognized biological effects induced by IFN-γ. Altogether, these data provide novel and additional evidences on MI medicines, and specifically when active substances are prepared at 4 CH, thus suggesting the need for more investigations.
Assuntos
Imunomodulação/imunologia , Interferon gama/imunologia , Linhagem Celular Tumoral , Células Cultivadas , Células Endoteliais da Veia Umbilical Humana/imunologia , Humanos , Imunidade/imunologia , Fatores Imunológicos/imunologia , Imunoterapia/métodos , Leucócitos Mononucleares/imunologia , Células THP-1RESUMO
In glioblastoma, non-classical human leucocyte antigen E (HLA-E) and HLA-G are frequently overexpressed. HLA-E loaded with peptides derived from HLA class I and from HLA-G contributes to inhibition of natural killer (NK) cells with expression of the inhibitory receptor CD94/NKG2A. We investigated whether NK cells expressing the activating CD94/NKG2C receptor counterpart were able to exert anti-glioma effects. NKG2C+ subsets were preferentially expanded by a feeder cell line engineered to express an artificial disulfide-stabilized trimeric HLA-E ligand (HLA-E*spG). NK cells expanded by a feeder cell line, which facilitates outgrowth of conventional NKG2A+, and fresh NK cells, were included for comparison. Expansion via the HLA-E*spG feeder cells selectively increased the fraction of NKG2C+ NK cells, which displayed a higher frequency of KIR2DL2/L3/S2 and CD16 when compared to expanded NKG2A+ NK cells. NKG2C+ NK cells exhibited increased cytotoxicity against K562 and KIR:HLA-matched and -mismatched primary glioblastoma multiforme (GBM) cells when compared to NKG2A+ NK cells and corresponding fresh NK cells. Cytotoxic responses of NKG2C+ NK cells were even more pronounced when utilizing target cells engineered with HLA-E*spG. These findings support the notion that NKG2C+ NK cells have potential therapeutic value for treating gliomas.
Assuntos
Neoplasias Encefálicas , Glioblastoma , Imunoterapia Adotiva , Células Matadoras Naturais , Subfamília C de Receptores Semelhantes a Lectina de Células NK , Neoplasias Encefálicas/imunologia , Neoplasias Encefálicas/terapia , Glioblastoma/metabolismo , Glioblastoma/terapia , Antígenos HLA-G/imunologia , Humanos , Fatores Imunológicos/imunologia , Fatores Imunológicos/metabolismo , Células K562 , Células Matadoras Naturais/imunologia , Subfamília C de Receptores Semelhantes a Lectina de Células NK/imunologiaRESUMO
Colorectal cancer (CRC), and breast, ovarian, pancreatic and prostate cancers are generally considered as low immune-reactive cancers that represent either limited infiltration of immune cells or extensive infiltration of immunosuppressive T cells. Interaction between programmed death ligand 1 (PD-L1) with programmed death-1 receptor (PD-1) is important for immune evasion. Tumors positive for PD-L1 generally show higher responses to the immune checkpoint inhibition (ICI); however, the high presence of PD-L1 in a tumor is a predictor of poor prognosis. Triple negative breast cancer (TNBC) is the most aggressive subtype of breast cancer, but responses to the ICI is meaningful. It seems that in a tumor both the PD-L1 expression and TIL infiltration is required for improving responses to the anti-PD-1/PD-L1 immunotherapy. Combination of anti-PD-1/PD-L1 with immune modulatory drugs, such as C-X-C chemokine receptor type 4 (CXCR4), poly (ADP-ribose) polymerase (PARP) or transforming growth factor (TGF)-ß inhibitors has shown meaningful clinical benefits.
Assuntos
Antígeno B7-H1/imunologia , Inibidores de Checkpoint Imunológico/imunologia , Neoplasias/imunologia , Neoplasias/terapia , Receptor de Morte Celular Programada 1/imunologia , Humanos , Fatores Imunológicos/imunologia , Imunoterapia/métodos , Linfócitos do Interstício Tumoral/imunologia , Linfócitos T/imunologiaRESUMO
Dissecting the different steps of the processing and presentation of tumor-associated antigens is a key aspect of immunotherapies enabling to tackle the immune response evasion attempts of cancer cells. The immunodominant glycoprotein gp100209-217 epitope, which is liberated from the melanoma differentiation antigen gp100PMEL17 , is part of immunotherapy trials. By analyzing different human melanoma cell lines, we here demonstrate that a pool of N-terminal extended peptides sharing the common minimal epitope is generated by melanoma proteasome subtypes. In vitro and in cellulo experiments indicate that ER-resident aminopeptidase 1 (ERAP1)-but not ERAP2-defines the processing of this peptide pool thereby modulating the T-cell recognition of melanoma cells. By combining the outcomes of our studies and others, we can sketch the complex processing and endogenous presentation pathway of the gp100209-217 -containing epitope/peptides, which are produced by proteasomes and are translocated to the vesicular compartment through different pathways, where the precursor peptides that reach the endoplasmic reticulum are further processed by ERAP1. The latter step enhances the activation of epitope-specific T lymphocytes, which might be a target to improve the efficiency of anti-melanoma immunotherapy.
Assuntos
Aminopeptidases/imunologia , Apresentação de Antígeno/imunologia , Retículo Endoplasmático/imunologia , Epitopos de Linfócito T/imunologia , Melanoma/imunologia , Melanoma/terapia , Antígenos de Histocompatibilidade Menor/imunologia , Antígenos de Neoplasias , Linhagem Celular Tumoral , Células HeLa , Humanos , Fatores Imunológicos/imunologia , Imunoterapia/métodos , Peptídeos/imunologia , Complexo de Endopeptidases do Proteassoma/imunologia , Linfócitos T/imunologiaRESUMO
Human mesenchymal stem cells (MSCs) are known to have anti-inflammatory and immunomodulatory functions; thus, several MSC products have been applied as cell therapy in clinical trials worldwide. Recent studies have demonstrated that MSC spheroids have superior anti-inflammatory and immunomodulatory functions to a single cell suspension. Current methods to prepare MSC spheroids include hanging drop, concave microwell aggregation, spinner flask, and gravity circulation. However, all these methods have limitations such as low scalability, easy cell clumping, low viability, and irregular size distribution. Here, we present a nano-patterned culture plasticware named PAMcell™ 3D plate to overcome these limitations. Nano-sized silica particles (700 nm) coated with RGD peptide were arrayed into fusiform onto the PLGA film. This uniform array enabled the seeded MSCs to grow only on the silica particles, forming uniform-sized semi-spheroids within 48 h. These MSC spheroids have been shown to have enhanced stemness, anti-inflammatory, and immunomodulatory functions, as revealed by the increased expression of stem cell markers (Oct4, Sox2, and Nanog), anti-inflammatory (IL-10, TSG6, and IDO), and immunomodulatory molecules (HGF, VEGF, CXCR4) both at mRNA and protein expression levels. Furthermore, these MSC spheroids demonstrated an increased palliative effect on glycemic control in a multiple low-dose streptozotocin-induced diabetes model compared with the same number of MSC single cell suspensions. Taken together, this study presents a new method to produce uniform-sized MSC spheroids with enhanced anti-inflammatory and immunomodulatory functions in vitro and in vivo.
Assuntos
Tecido Adiposo/citologia , Anti-Inflamatórios/imunologia , Técnicas de Cultura de Células/métodos , Fatores Imunológicos/imunologia , Células-Tronco Mesenquimais/imunologia , Esferoides Celulares/imunologia , Animais , Técnicas de Cultura de Células/instrumentação , Células Cultivadas , Diabetes Mellitus Experimental/imunologia , Diabetes Mellitus Experimental/terapia , Expressão Gênica/imunologia , Humanos , Masculino , Transplante de Células-Tronco Mesenquimais/métodos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Camundongos Endogâmicos C57BL , Esferoides Celulares/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/imunologia , Fatores de Transcrição/metabolismoRESUMO
INTRODUCTION: Endometriosis is a risk factor for low-grade serous, clear cell, and endometroid ovarian carcinoma. In both endometriosis and ovarian carcinoma, immunological factors are associated with clinical outcome. Chronic inflammation in endometriosis may be linked to tumorigenesis, but exact processes contributing to endometriosis-associated ovarian carcinoma remain unknown. This review aims to describe potential immunological factors involved in the malignant transformation of endometriosis into ovarian carcinoma. METHODS: PubMed and Embase were searched from inception up to October 2020 for studies comparing immunological processes in endometriosis and endometriosis-associated ovarian carcinoma. RESULTS: Detailed analysis of immune components in the malignant transformation of endometriosis into endometriosis-associated ovarian carcinoma is lacking. Altered levels of chemokines and cytokines as IL-6, IL-8, IL-10, and TNF-α are reported and the function, number and polarization of NK cells, dendritic cells, and monocytes differ between endometriosis and associated ovarian carcinoma compared to healthy tissue. In addition, altered inflammasome and complement systems, indicate a role for the immune system in the carcinogenesis of endometriosis. CONCLUSION: Chronic inflammation in endometriosis may potentially drive inflammation-induced carcinogenesis in endometriosis-associated ovarian carcinoma. Exact immunological pathways and cellular processes remain unknown and require more thorough investigation.
Assuntos
Transformação Celular Neoplásica/patologia , Endometriose/complicações , Fatores Imunológicos/imunologia , Neoplasias Ovarianas/patologia , Animais , Transformação Celular Neoplásica/imunologia , Endometriose/imunologia , Feminino , Humanos , Neoplasias Ovarianas/etiologiaRESUMO
One of the major hurdles for the advancement of cancer immunotherapy is lack of robust, accessible experimental models. We aimed to produce an ex-vivo organ culture (EVOC) model of immunotherapy for non-small cell lung cancer (NSCLC). Freshly resected early stage tumors were collected from the operating room, fragmented to clusters < 450 µm and cultured with fetal calf serum and human autologous serum. The resulting EVOC includes cancer epithelial cells within tumor tissue clusters and immune cells. Original tissue features are reflected in the EVOCs. The response to immune checkpoint inhibitors (ICI) was assessed by IFNγ gene induction. Interestingly, IFNγ EVOC induction was numerically higher when anti-CTLA4 was added to anti-PD-L1 treatment, supporting the notion that anti-CTLA4 impacts cancer partly through tumor-resident immune cells. In parallel, immunohistochemistry (IHC) for key immune-related proteins was performed on the formalin-fixed paraffin embedded (FFPE) corresponding tumors. EVOC IFNγ induction by ICI correlated with basal non-induced IFNγ, CD8, CD4 and FOXP3 mRNA levels within EVOCs and with tumor-FFPE-IHC for CD8 and granzyme B. A weaker correlation was seen with tumor-FFPE-IHC for CD3, CD4, CD68, FOXP3 and tumor-PD-L1. Tertiary lymphoid structure density was also correlated with the ICI response. Our study provides novel data about biomarkers that correlate with ICI-induced response of early stage NSCLC. Retention of the microenvironment and minimal addition of exogenous factors suggest this model to reliably represent the original tumor. The cluster-based EVOC model we describe can provide a valuable, yet simple and widely applicable tool for the study of immunotherapy in NSCLC.
Assuntos
Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/terapia , Antígeno B7-H1/imunologia , Biomarcadores Tumorais/imunologia , Antígenos CD4/imunologia , Antígenos CD8/imunologia , Antígeno CTLA-4/imunologia , Carcinoma Pulmonar de Células não Pequenas/imunologia , Carcinoma Pulmonar de Células não Pequenas/terapia , Fatores de Transcrição Forkhead/imunologia , Humanos , Imuno-Histoquímica/métodos , Fatores Imunológicos/imunologia , Imunoterapia/métodos , Interferon gama/imunologia , Técnicas de Cultura de Órgãos/métodos , Microambiente Tumoral/imunologiaRESUMO
INTRODUCTION: CD73 is a membrane-bound enzyme crucial in adenosine generation. The adenosinergic pathway plays a critical role in immunosuppression and in anti-tumor effects of immune checkpoint inhibitors (ICI). Here, we interrogated CD73 expression in a richly annotated cohort of human lung adenocarcinoma (LUAD) and its association with clinicopathological, immune, and molecular features to better understand the role of this immune marker in LUAD pathobiology. MATERIALS AND METHODS: Protein expression of CD73 was evaluated by immunohistochemistry in 106 archived LUADs from patients that underwent surgical treatment without neoadjuvant therapy. Total CD73 (T +) was calculated as the average of luminal (L +) and basolateral (BL +) percentage membrane expression scores for each LUAD and was used to classify tumors into three groups based on the extent of T CD73 expression (high, low, and negative). RESULTS: CD73 expression was significantly and progressively increased across normal-appearing lung tissue, adenomatous atypical hyperplasia, adenocarcinoma in situ, minimally invasive adenocarcinoma, and LUAD. In LUAD, BL CD73 expression was associated with an increase in PD-L1 expression in tumor cells and increase of tumor-associated immune cells. Stratification of LUADs based on T CD73 extent also revealed that tumors with high expression of this enzyme overall exhibited significantly elevated immune infiltration and PD-L1 protein expression. Immune profiling demonstrated that T-cell inflammation and adenosine signatures were significantly higher in CD73-expressing lung adenocarcinomas relative to those lacking CD73. CONCLUSION: Our study suggests that higher CD73 expression is associated with an overall augmented host immune response, suggesting potential implications in the immune pathobiology of early stage lung adenocarcinoma. Our findings warrant further studies to explore the role of CD73 in immunotherapeutic response of LUAD.
Assuntos
5'-Nucleotidase/metabolismo , Adenocarcinoma de Pulmão/patologia , Biomarcadores Tumorais/análise , Carcinoma Pulmonar de Células não Pequenas/patologia , Fatores Imunológicos/imunologia , Neoplasias Pulmonares/patologia , Adenocarcinoma de Pulmão/imunologia , Adenocarcinoma de Pulmão/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma Pulmonar de Células não Pequenas/imunologia , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Feminino , Seguimentos , Proteínas Ligadas por GPI/metabolismo , Humanos , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/metabolismo , Masculino , Pessoa de Meia-Idade , Prognóstico , Estudos RetrospectivosRESUMO
Despite undeniable improvement in the management of rheumatoid arthritis (RA), the discovery of more effective, less toxic and, ideally, less immune suppressive drugs are much needed. In the current study, we set to explore the potential anti-rheumatic activity of the non-toxic, tellurium-based immunomodulator, AS101 in an experimental animal model of RA. The effect of AS101 was assessed on adjuvant-induced arthritis (AIA) rats. Clinical signs of arthritis were assessed. Histopathological examination was used to assess inflammation, synovial changes and tissue lesions. Very late antigen-4 (VLA-4)+ cellular infiltration was detected using immunohistochemical staining. Enzyme-linked immunosorbent assay (ELISA) was used to measure circulating anti-cyclic citrullinated-peptide autoantibody (ACPA) and real-time polymerase chain reaction (PCR) was used to measure the in-vitro effect of AS101 on interleukin (IL)-6 and IL-1ß expression in activated primary human fibroblasts. Prophylactic treatment with intraperitoneal AS101 reduced clinical arthritis scores in AIA rats (P < 0·01). AS101 abrogated the migration of active chronic inflammatory immune cells, particularly VLA-4+ cells, into joint cartilage and synovium, reduced the extent of joint damage and preserved joint architecture. Compared to phosphate-buffered saline (PBS)-treated AIA rats, histopathological inflammatory scores were significantly reduced (P < 0·05). Furthermore, AS101 resulted in a marked reduction of circulating ACPA in comparison to PBS-treated rats (P < 0·05). Importantly, AS101 significantly reduced mRNA levels of proinflammatory mediators such as IL-6 (P < 0·05) and IL-1ß (P < 0·01) in activated primary human fibroblasts. Taken together, we report the first demonstration of the anti-rheumatic/inflammatory activity of AS101 in experimental RA model, thereby supporting an alternative early therapeutic intervention and identifying a promising agent for therapeutic intervention.
Assuntos
Artrite Experimental/imunologia , Artrite Reumatoide/imunologia , Etilenos/imunologia , Telúrio/imunologia , Adjuvantes Imunológicos/farmacologia , Animais , Artrite Experimental/metabolismo , Artrite Experimental/prevenção & controle , Artrite Reumatoide/metabolismo , Artrite Reumatoide/prevenção & controle , Células Cultivadas , Etilenos/farmacologia , Feminino , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Expressão Gênica/efeitos dos fármacos , Humanos , Fatores Imunológicos/imunologia , Fatores Imunológicos/farmacologia , Integrina alfa4beta1/imunologia , Integrina alfa4beta1/metabolismo , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Interleucina-6/genética , Interleucina-6/metabolismo , Ratos Endogâmicos Lew , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Telúrio/farmacologiaRESUMO
Recent evidence suggested that neurological manifestations occur in patients with a severe form of coronavirus disease (COVID-19). On the basis of this issue, neurologists are very concerned about patients with neurological disorders, especially multiple sclerosis (MS), as consumers of immunosuppressive or immune-modulating drugs. Therefore, the administration of proper disease-modifying therapies (DMTs) in MS patients is critical during the pandemic status. On the one hand, both the autoimmune diseases and immunosuppressive drugs increase the risk of infection due to impairment in the immune system, and on the other hand, postponing of MS treatment has serious consequences on the central nervous system. In the present study, we discussed recent literature about the effect of DMTs administration on the severity of COVID-19 in the MS patients. Overall, it seems that DMTs do not provoke the COVID-19 infection in the MS patients by declining immune responses and cytokine storm. However, as a precaution, the supervision of a neurologist is highly recommended.