Inhibitory receptors of plasmacytoid dendritic cells as possible targets for checkpoint blockade in cancer.

Plasmacytoid dendritic cells (pDCs) are the major producers of type I interferons (IFNs), which are essential to mount antiviral and antitumoral immune responses. To avoid exaggerated levels of type I IFNs, which pave the way to immune dysregulation and autoimmunity, pDC activation is strictly regulated by a variety of inhibitory receptors (IRs). In tumors, pDCs display an exhausted phenotype and correlate with an unfavorable prognosis, which largely depends on the accumulation of immunosuppressive cytokines and oncometabolites. This review explores the hypothesis that tumor microenvironment may reduce the release of type I IFNs also by a more pDC-specific mechanism, namely the engagement of IRs. Literature shows that many cancer types express de novo, or overexpress, IR ligands (such as BST2, PCNA, CAECAM-1 and modified surface carbohydrates) which often represent a strong predictor of poor outcome and metastasis. In line with this, tumor cells expressing ligands engaging IRs such as BDCA-2, ILT7, TIM3 and CD44 block pDC activation, while this blocking is prevented when IR engagement or signaling is inhibited. Based on this evidence, we propose that the regulation of IFN secretion by IRs may be regarded as an "innate checkpoint", reminiscent of the function of "classical" adaptive immune checkpoints, like PD1 expressed in CD8+ T cells, which restrain autoimmunity and immunopathology but favor chronic infections and tumors. However, we also point out that further work is needed to fully unravel the biology of tumor-associated pDCs, the neat contribution of pDC exhaustion in tumor growth following the engagement of IRs, especially those expressed also by other leukocytes, and their therapeutic potential as targets of combined immune checkpoint blockade in cancer immunotherapy.

Extracellular microRNAs induce dendritic cell-dependent joint inflammation and potentiate osteoclast differentiation via TLR7/8 engagement.

OBJECTIVES: Monocyte-derived dendritic cells (DCs) are key players in the induction of inflammation, autoreactive T cell activation and loss of tolerance in rheumatoid arthritis (RA), but the precise mechanisms underlying their activation remain elusive. Here, we hypothesized that extracellular microRNAs released in RA synovial fluids may represent a novel, physiological stimulus triggering unwanted immune response via TLR8-expressing DC stimulation. METHODS: Human monocyte-derived DCs were stimulated with a mixture of GU-rich miRNAs upregulated in RA tissues and released in synovial fluids (Ex-miRNAs). Activation of DCs was assessed in terms of NF-κB activation by Western blot, cytokine production by ELISA, T cell proliferation and polarization by allogeneic mixed lymphocyte reaction. DC differentiation into osteoclasts was evaluated in terms of tartrate-resistant acid phosphatase production and formation of resorption pits in dentine slices. Induction of joint inflammation in vivo was evaluated using a murine model of DC-induced arthritis. TLR7/8 involvement was assessed by specific inhibitors. RESULTS: Ex-miRNAs activate DCs to secrete TNFα, induce joint inflammation, start an early autoimmune response and potentiate the differentiation of DCs into aggressive osteoclasts. CONCLUSIONS: This work represents a proof of concept that the pool of extracellular miRNAs overexpressed in RA joints can act as a physiological activator of inflammation via the stimulation of TLR8 expressed by human DCs, which in turn exert arthritogenic functions. In this scenario, pharmacological inhibition of TLR8 might offer a new therapeutic option to reduce inflammation and osteoclast-mediated bone destruction in RA.

Modulation of Human Dendritic Cell Functions by Phosphodiesterase-4 Inhibitors: Potential Relevance for the Treatment of Respiratory Diseases.

Inhibitors of phosphodiesterase-4 (PDE4) are small-molecule drugs that, by increasing the intracellular levels of cAMP in immune cells, elicit a broad spectrum of anti-inflammatory effects. As such, PDE4 inhibitors are actively studied as therapeutic options in a variety of human diseases characterized by an underlying inflammatory pathogenesis. Dendritic cells (DCs) are checkpoints of the inflammatory and immune responses, being responsible for both activation and dampening depending on their activation status. This review shows evidence that PDE4 inhibitors modulate inflammatory DC activation by decreasing the secretion of inflammatory and Th1/Th17-polarizing cytokines, although preserving the expression of costimulatory molecules and the CD4+ T cell-activating potential. In addition, DCs activated in the presence of PDE4 inhibitors induce a preferential Th2 skewing of effector T cells, retain the secretion of Th2-attracting chemokines and increase the production of T cell regulatory mediators, such as IDO1, TSP-1, VEGF-A and Amphiregulin. Finally, PDE4 inhibitors selectively induce the expression of the surface molecule CD141/Thrombomodulin/BDCA-3. The result of such fine-tuning is immunomodulatory DCs that are distinct from those induced by classical anti-inflammatory drugs, such as corticosteroids. The possible implications for the treatment of respiratory disorders (such as COPD, asthma and COVID-19) by PDE4 inhibitors will be discussed.

CCRL2 Expression by Specialized Lung Capillary Endothelial Cells Controls NK-cell Homing in Lung Cancer.

Patterns of receptors for chemotactic factors regulate the homing of leukocytes to tissues. Here we report that the CCRL2/chemerin/CMKLR1 axis represents a selective pathway for the homing of natural killer (NK) cells to the lung. C-C motif chemokine receptor-like 2 (CCRL2) is a nonsignaling seven-transmembrane domain receptor able to control lung tumor growth. CCRL2 constitutive or conditional endothelial cell targeted ablation, or deletion of its ligand chemerin, were found to promote tumor progression in a Kras/p53Flox lung cancer cell model. This phenotype was dependent on the reduced recruitment of CD27- CD11b+ mature NK cells. Other chemotactic receptors identified in lung-infiltrating NK cells by single-cell RNA sequencing (scRNA-seq), such as Cxcr3, Cx3cr1, and S1pr5, were found to be dispensable in the regulation of NK-cell infiltration of the lung and lung tumor growth. scRNA-seq identified CCRL2 as the hallmark of general alveolar lung capillary endothelial cells. CCRL2 expression was epigenetically regulated in lung endothelium and it was upregulated by the demethylating agent 5-aza-2'-deoxycytidine (5-Aza). In vivo administration of low doses of 5-Aza induced CCRL2 upregulation, increased recruitment of NK cells, and reduced lung tumor growth. These results identify CCRL2 as an NK-cell lung homing molecule that has the potential to be exploited to promote NK cell-mediated lung immune surveillance.

SARS-CoV-2-Associated ssRNAs Activate Human Neutrophils in a TLR8-Dependent Fashion.

COVID-19 disease is characterized by a dysregulation of the innate arm of the immune system. However, the mechanisms whereby innate immune cells, including neutrophils, become activated in patients are not completely understood. Recently, we showed that GU-rich RNA sequences from the SARS-CoV-2 genome (i.e., SCV2-RNA1 and SCV2-RNA2) activate dendritic cells. To clarify whether human neutrophils may also represent targets of SCV2-RNAs, neutrophils were treated with either SCV2-RNAs or, as a control, R848 (a TLR7/8 ligand), and were then analyzed for several functional assays and also subjected to RNA-seq experiments. Results highlight a remarkable response of neutrophils to SCV2-RNAs in terms of TNFα, IL-1ra, CXCL8 production, apoptosis delay, modulation of CD11b and CD62L expression, and release of neutrophil extracellular traps. By RNA-seq experiments, we observed that SCV2-RNA2 promotes a transcriptional reprogramming of neutrophils, characterized by the induction of thousands of proinflammatory genes, similar to that promoted by R848. Furthermore, by using CU-CPT9a, a TLR8-specific inhibitor, we found that SCV2-RNA2 stimulates neutrophils exclusively via TLR8-dependent pathways. In sum, our study proves that single-strand RNAs from the SARS-CoV-2 genome potently activate human neutrophils via TLR8, thus uncovering a potential mechanism whereby neutrophils may contribute to the pathogenesis of severe COVID-19 disease.

Inhibition of Class I Histone Deacetylase Activity Blocks the Induction of TNFAIP3 Both Directly and Indirectly via the Suppression of Endogenous TNF-α.

Histone deacetylase inhibitors (HDIs) are promising drugs for the treatment of inflammatory diseases. However, their therapeutical exploitation is slowed down by severe adverse manifestations that can hardly be foreseen, mainly due to incomplete knowledge of how HDIs impact the delicate balance of inflammatory mediators. In this work, we characterized the effects of the HDI trichostatin A (TSA) on the expression of TNFAIP3, which is a crucial inhibitor of the classical NF-kB pathway and an LPS-induced negative feedback regulator. The accumulation of TNFAIP3 mRNA after LPS stimulation showed biphasic behavior, with one wave within the first hour of stimulation and a second wave several hours later, which were both reduced by TSA. By using inhibition and knockdown approaches, we identified two temporally and mechanistically distinct modes of action. The first wave of TNAIP3 accumulation was directly blunted by the histone deacetylase (HDAC) blockade. By contrast, the second wave was decreased mainly because of the lack of endogenous TNF-α induction, which, in turn, depended on the intact HDAC activity. In both cases, class I HDACs appeared to play a nonredundant role, with HDAC3 required, but not sufficient, for TNF-α and TNFAIP3 induction. In addition to TNFAIP3, TNF-α is known to induce many response genes that orchestrate the inflammatory cascade. Thus, suppression of TNF-α may represent a general mechanism through which HDIs regulate a selected set of target genes.

The PDE4 inhibitor tanimilast shows distinct immunomodulatory properties associated with a type 2 endotype and CD141 upregulation.

BACKGROUND: Tanimilast is a novel and selective inhaled inhibitor of phosphodiesterase-4 in advanced clinical development for chronic obstructive pulmonary disease (COPD). Tanimilast is known to exert prominent anti-inflammatory activity when tested in preclinical experimental models as well as in human clinical studies. Recently, we have demonstrated that it also finely tunes, rather than suppressing, the cytokine network secreted by activated dendritic cells (DCs). This study was designed to characterize the effects of tanimilast on T-cell polarizing properties of DCs and to investigate additional functional and phenotypical features induced by tanimilast. METHODS: DCs at day 6 of culture were stimulated with LPS in the presence or absence of tanimilast or the control drug budesonide. After 24 h, DCs were analyzed for the expression of surface markers of maturation and activation by flow cytometry and cocultured with T cells to investigate cell proliferation and activation/polarization. The regulation of type 2-skewing mediators was investigated by real-time PCR in DCs and compared to results obtained in vivo in a randomized placebo-controlled trial on COPD patients treated with tanimilast. RESULTS: Our results show that both tanimilast and budesonide reduced the production of the immunostimulatory cytokine IFN-γ by CD4+ T cells. However, the two drugs acted at different levels since budesonide mainly blocked T cell proliferation, while tanimilast skewed T cells towards a Th2 phenotype without affecting cell proliferation. In addition, only DCs matured in the presence of tanimilast displayed increased CD86/CD80 ratio and CD141 expression, which correlated with Th2 T cell induction and dead cell uptake respectively. These cells also upregulated cAMP-dependent immunosuppressive molecules such as IDO1, TSP1, VEGF-A and Amphiregulin. Notably, the translational value of these data was confirmed by the finding that these same genes were upregulated also in sputum cells of COPD patients treated with tanimilast as add-on to inhaled glucocorticoids and bronchodilators. CONCLUSION: Taken together, these findings demonstrate distinct immunomodulatory properties of tanimilast associated with a type 2 endotype and CD141 upregulation in DCs and provide a mechanistic rationale for the administration of tanimilast on top of inhaled corticosteroids.

The PDE4 Inhibitor Tanimilast Restrains the Tissue-Damaging Properties of Human Neutrophils.

Neutrophils, the most abundant subset of leukocytes in the blood, play a pivotal role in host response against invading pathogens. However, in respiratory diseases, excessive infiltration and activation of neutrophils can lead to tissue damage. Tanimilast-international non-proprietary name of CHF6001-is a novel inhaled phosphodiesterase 4 (PDE4) inhibitor in advanced clinical development for the treatment of chronic obstructive pulmonary disease (COPD), a chronic inflammatory lung disease where neutrophilic inflammation plays a key pathological role. Human neutrophils from healthy donors were exposed to pro-inflammatory stimuli in the presence or absence of tanimilast and budesonide-a typical inhaled corticosteroid drug-to investigate the modulation of effector functions including adherence to endothelial cells, granule protein exocytosis, release of extracellular DNA traps, cytokine secretion, and cell survival. Tanimilast significantly decreased neutrophil-endothelium adhesion, degranulation, extracellular DNA traps casting, and cytokine secretion. In contrast, it promoted neutrophil survival by decreasing both spontaneous apoptosis and cell death in the presence of pro-survival factors. The present work suggests that tanimilast can alleviate the severe tissue damage caused by massive recruitment and activation of neutrophils in inflammatory diseases such as COPD.

Z-alpha1-antitrypsin polymers and small airways disease: a new paradigm in alfa-1 anti-trypsin deficiency-related COPD development?

The presence of Alpha1-Antitrypsin (AAT) polymers, known to promote a sustained pro-inflammatory activity, has been previously demonstrated in bronchial biopsies of subjects with Z-AAT deficiency (AATD) suggesting a possible role in the development of COPD through a small airway disease impairment. The study aimed to assess the presence of small airways dysfunction and the potential correlation with the presence of Z-AAT polymers obtained by Exhaled Breath Condensate (EBC) collection in PiZZ subjects, as compared with matched healthy PiMM subjects. We enrolled 19 asymptomatic, never smoker subjects: 9 PiZZ and 10 PiMM as controls, without obstructive ventilatory defect (i.e., normal FEV1/VC% ratio). All subjects underwent complete pulmonary function tests (PFT). EBC was collected in all subjects. ELISA test was applied to search for Z-AAT polymers. The PiZZ subjects showed normal lung volumes and DLCO values. However, in comparison with PiMM subjects, the single breath test N2 wash-out revealed significant differences regarding the phase III slope (1.45±0.35 N2/L vs. 0.96±0.40 N2/L) (p<0.02) in the PiZZ subjects, while the closing volume/vital capacity ratio (14.3±4.5 % vs. 11.3±6.3 %) was not significantly increased. The ELISA test detected the presence of Z-AAT polymers in 44% of PiZZ patients. Asymptomatic, never smoker PiZZ subjects with normal spirometry and lung diffusion capacity showed airways impairment when compared to PiMM subjects. Although Z-AAT polymers were found only in 44% of PiZZ subjects, these findings suggest the possibility that chronic bronchiolitis can develop as a result of the long-term pro-inflammatory activity of Z-AAT polymers in subjects with Z-related AATD.

Investigating the Link between Alpha-1 Antitrypsin Deficiency and Abdominal Aortic Aneurysms.

BACKGROUND: Alpha-1-Antitrypsin (AAT) is one of the major plasmatic protease inhibitors. In the last decade, an association between Alpha-1-Antitrypsin Deficiency (AATD) and Abdominal Aortic Aneurysms (AAA) has been hypothesized. Multiple factors may be involved in AAA's etiopathogenesis, and an underlying structural defect of the extracellular matrix (ECM) is always present. AATD could be a reasonable risk factor for AAA because it is related to protease/antiprotease imbalance and enhanced ECM degradation of the vessel wall. METHODS: We performed genotyping of 138 patients hospitalized in the Vascular Surgery Division of the ASST-Spedali Civili di Brescia, Italy, for nontraumatic rupture of AAA. The second purpose was to observe the distribution of main nongenetic risk factors for AAA between patients with and without AATD. RESULTS: Out of 138 patients, 22 were found with AATD: 16 MS, 1 SS, 3 MZ, and 2 with a new rare AAT variant. When compared to the general Italian population, our cohort's frequency of deficient S allele was significantly higher (7.8 vs. 2.2% respectively, P < 0.01), whereas the deficient Z allele was similar (1.1 vs. 1.3% respectively, P > 0.05). Although we found no differences in age, gender, hypertension, diabetes, and smoke habits between AAA patients with and without AATD, hyperlipidemia was significantly less frequent in patients with AATD (46.4 vs. 12.5% respectively, P < 0.05). CONCLUSIONS: In our AAA patients' cohort, the S allele frequency was higher than in the general Italian population. Our results support the hypothesis that AATD might be a risk factor for AAA.

SARS-CoV-2-associated ssRNAs activate inflammation and immunity via TLR7/8.

The inflammatory and IFN pathways of innate immunity play a key role in the resistance and pathogenesis of coronavirus disease 2019 (COVID-19). Innate sensors and SARS-CoV-2-associated molecular patterns (SAMPs) remain to be completely defined. Here, we identified single-stranded RNA (ssRNA) fragments from the SARS-CoV-2 genome as direct activators of endosomal TLR7/8 and MyD88 pathway. The same sequences induced human DC activation in terms of phenotype and function, such as IFN and cytokine production and Th1 polarization. A bioinformatic scan of the viral genome identified several hundreds of fragments potentially activating TLR7/8, suggesting that products of virus endosomal processing potently activate the IFN and inflammatory responses downstream of these receptors. In vivo, SAMPs induced MyD88-dependent lung inflammation characterized by accumulation of proinflammatory and cytotoxic mediators and immune cell infiltration, as well as splenic DC phenotypical maturation. These results identified TLR7/8 as a crucial cellular sensor of ssRNAs encoded by SARS-CoV-2 involved in host resistance and the disease pathogenesis of COVID-19.

The PDE4 Inhibitor Tanimilast Blunts Proinflammatory Dendritic Cell Activation by SARS-CoV-2 ssRNAs.

Phosphodiesterase 4 (PDE4) inhibitors are immunomodulatory drugs approved to treat diseases associated with chronic inflammatory conditions, such as COPD, psoriasis and atopic dermatitis. Tanimilast (international non-proprietary name of CHF6001) is a novel, potent and selective inhaled PDE4 inhibitor in advanced clinical development for the treatment of COPD. To begin testing its potential in limiting hyperinflammation and immune dysregulation associated to SARS-CoV-2 infection, we took advantage of an in vitro model of dendritic cell (DC) activation by SARS-CoV-2 genomic ssRNA (SCV2-RNA). In this context, Tanimilast decreased the release of pro-inflammatory cytokines (TNF-α and IL-6), chemokines (CCL3, CXCL9, and CXCL10) and of Th1-polarizing cytokines (IL-12, type I IFNs). In contrast to ß-methasone, a reference steroid anti-inflammatory drug, Tanimilast did not impair the acquisition of the maturation markers CD83, CD86 and MHC-II, nor that of the lymph node homing receptor CCR7. Consistent with this, Tanimilast did not reduce the capability of SCV2-RNA-stimulated DCs to activate CD4+ T cells but skewed their polarization towards a Th2 phenotype. Both Tanimilast and ß-methasone blocked the increase of MHC-I molecules in SCV2-RNA-activated DCs and restrained the proliferation and activation of cytotoxic CD8+ T cells. Our results indicate that Tanimilast can modulate the SCV2-RNA-induced pro-inflammatory and Th1-polarizing potential of DCs, crucial regulators of both the inflammatory and immune response. Given also the remarkable safety demonstrated by Tanimilast, up to now, in clinical studies, we propose this inhaled PDE4 inhibitor as a promising immunomodulatory drug in the scenario of COVID-19.

Functional Role of Dendritic Cell Subsets in Cancer Progression and Clinical Implications.

Dendritic cells (DCs) constitute a complex network of cell subsets with common functions but also with many divergent aspects. All dendritic cell subsets share the ability to prime T cell response and to undergo a complex trafficking program related to their stage of maturation and function. For these reasons, dendritic cells are implicated in a large variety of both protective and detrimental immune responses, including a crucial role in promoting anti-tumor responses. Although cDC1s are the most potent subset in tumor antigen cross-presentation, they are not sufficient to induce full-strength anti-tumor cytotoxic T cell response and need close interaction and cooperativity with the other dendritic cell subsets, namely cDC2s and pDCs. This review will take into consideration different aspects of DC biology, including the functional role of dendritic cell subsets in both fostering and suppressing tumor growth, the mechanisms underlying their recruitment into the tumor microenvironment, as well as the prognostic value and the potentiality of dendritic cell therapeutic targeting. Understanding the specificity of dendritic cell subsets will allow to gain insights on role of these cells in pathological conditions and to design new selective promising therapeutic approaches.

Cytokine Targeting by miRNAs in Autoimmune Diseases.

Persistent and excessive cytokine production is a hallmark of autoimmune diseases and may play a role in disease pathogenesis and amplification. Therefore, cytokine neutralization is a useful therapeutic strategy to treat immune-mediated conditions. MicroRNAs (miRNAs) are small non-coding RNA molecules that regulate gene expression in diverse biological processes. Altered miRNA levels are observed in most autoimmune diseases and are recognized to influence autoimmunity through different mechanisms. Here, we review the impact of altered miRNA levels on the expression of cytokines that play a relevant pathogenic role in autoimmunity, namely primary pro-inflammatory cytokines, the IL-17/IL-23 axis, type I interferons and IL-10. Regulation can be either "direct" on the target cytokine, or "indirect," meaning that one given miRNA post-transcriptionally regulates the expression of a protein that in turn influences the level of the cytokine. In addition, miRNAs associated with extracellular vesicles can regulate cytokine production in neighboring cells, either post-transcriptionally or via the stimulation of innate immune RNA-sensors, such as Toll-like receptors. Because of their tremendous potential as physiological and pathological regulators, miRNAs are in the limelight as promising future biopharmaceuticals. Thus, these studies may lead in the near future to the design and testing of therapeutic miRNAs as next generation drugs to target pathogenic cytokines in autoimmunity.

Chemokine and chemotactic signals in dendritic cell migration.

Dendritic cells (DCs) are professional antigen-presenting cells responsible for the activation of specific T-cell responses and for the development of immune tolerance. Immature DCs reside in peripheral tissues and specialize in antigen capture, whereas mature DCs reside mostly in the secondary lymphoid organs where they act as antigen-presenting cells. The correct localization of DCs is strictly regulated by a large variety of chemotactic and nonchemotactic signals that include bacterial products, DAMPs (danger-associated molecular patterns), complement proteins, lipids, and chemokines. These signals function both individually and in concert, generating a complex regulatory network. This network is regulated at multiple levels through different strategies, such as synergistic interactions, proteolytic processing, and the actions of atypical chemokine receptors. Understanding this complex scenario will help to clarify the role of DCs in different pathological conditions, such as autoimmune diseases and cancers and will uncover new molecular targets for therapeutic interventions.

The Atypical Receptor CCRL2 (C-C Chemokine Receptor-Like 2) Does Not Act As a Decoy Receptor in Endothelial Cells.

C-C chemokine receptor-like 2 (CCRL2) is a non-signaling seven-transmembrane domain (7-TMD) receptor related to the atypical chemokine receptor (ACKR) family. ACKRs bind chemokines but do not activate G protein-dependent signaling or cell functions. ACKRs were shown to regulate immune functions in vivo by their ability to scavenge chemokines from the local environment. This study was performed to investigate whether CCRL2 shares two of the main characteristics of ACKRs, namely the ability to internalize and scavenge the ligands. Cell membrane analysis of CCRL2-transfected cells revealed a weak, constitutive, ligand-independent internalization, and recycling of CCRL2, with a kinetics that was slower than those observed with ACKR3, a prototypic ACKR, or other chemotactic signaling receptors [i.e., chemokine-like receptor 1 and C-X-C motif chemokine receptor 2]. Intracellularly, CCRL2 colocalized with early endosome antigen 1-positive and Rab5-positive vesicles and with recycling compartments mainly characterized by Rab11-positive vesicles. CCRL2-transfected cells and activated mouse blood endothelial cells, that endogenously express CCRL2, were used to investigate the scavenging ability of CCRL2. These experiments confirmed the ability of CCRL2 to bind chemerin, the only recognized ligand, but excluded the ability of CCRL2 to perform scavenging. Collectively, these results identify unique functional properties for this member of the non-signaling 7-TMD receptor family.

The atypical receptor CCRL2 is required for CXCR2-dependent neutrophil recruitment and tissue damage.

CCRL2 is a 7-transmembrane domain receptor that shares structural and functional similarities with the family of atypical chemokine receptors (ACKRs). CCRL2 is upregulated by inflammatory signals and, unlike other ACKRs, it is not a chemoattractant-scavenging receptor, does not activate ß-arrestins, and is widely expressed by many leukocyte subsets. Therefore, the biological role of CCRL2 in immunity is still unclear. We report that CCRL2-deficient mice have a defect in neutrophil recruitment and are protected in 2 models of inflammatory arthritis. In vitro, CCRL2 was found to constitutively form homodimers and heterodimers with CXCR2, a main neutrophil chemotactic receptor. By heterodimerization, CCRL2 could regulate membrane expression and promote CXCR2 functions, including the activation of ß2-integrins. Therefore, upregulation of CCRL2 observed under inflammatory conditions is functional to finely tune CXCR2-mediated neutrophil recruitment at sites of inflammation.

Leukocyte trafficking in tumor microenvironment.

The tumor microenvironment consists of both malignant and non-malignant cells and a plethora of soluble mediators. Different types of tumors have specific tumor microenvironments characterized by distinct chemokines and chemotactic factors that influence leukocyte recruitment. The immune cell infiltrate continuously interacts with stroma cells and influence tumor growth. Emerging evidence suggests that the regulation of the composition and the metabolic state of tumor-associated leukocytes may represent a new promising intervention strategy. Here we summarize the current knowledge on the role of tumor-associated immune cells in tumor growth and dissemination, with a specific focus on the nature of the chemotactic factors responsible for their accumulation and activation in tumors.

Central airways remodeling in COPD patients.

BACKGROUND: The contribution to airflow obstruction by the remodeling of the peripheral airways in chronic obstructive pulmonary disease (COPD) patients has been well documented, but less is known about the role played by the large airways. Few studies have investigated the presence of histopathological changes due to remodeling in the large airways of COPD patients. OBJECTIVES: The aim of this study was to verify the presence of airway remodeling in the central airways of COPD patients, quantifying the airway smooth muscle (ASM) area and the extracellular matrix (ECM) protein deposition, both in the subepithelial region and in the ASM, and to verify the possible contribution to airflow obstruction by the above mentioned histopathological changes. METHODS: Biopsies of segmental bronchi spurs were performed in COPD patients and control smoker subjects and immunostained for collagen type I, versican, decorin, biglycan, and alpha-smooth muscle actin. ECM protein deposition was measured at both subepithelial, and ASM layers. RESULTS: The staining for collagen I and versican was greater in the subepithelial layer of COPD patients than in control subjects. An inverse correlation was found between collagen I in the subepithelial layer and both forced expiratory volume in 1 second and ratio between forced expiratory volume in 1 second and forced vital capacity. A statistically significant increase of the ASM area was observed in the central airways of COPD patients versus controls. CONCLUSION: These findings indicate that airway remodeling also affects the large airways in COPD patients who have greater deposition of ECM proteins in the subepithelial layer and a larger smooth muscle area than control smoker subjects. These changes may contribute to chronic airflow obstruction in COPD patients.

The role of bronchial epithelial cells in the pathogenesis of COPD in Z-alpha-1 antitrypsin deficiency.

BACKGROUND: Alpha-1 antitrypsin is the main inhibitor of neutrophil elastase in the lung. Although it is principally synthesized by hepatocytes, alpha-1 antitrypsin is also secreted by bronchial epithelial cells. Gene mutations can lead to alpha-1 antitrypsin deficiency, with the Z variant being the most clinically relevant due to its propensity to polymerize. The ability of bronchial epithelial cells to produce Z-variant protein and its polymers is unknown. METHODS: Experiments using a conformation-specific antibody were carried out on M- and Z-variant-transfected 16HBE cells and on bronchial biopsies and ex vivo bronchial epithelial cells from Z and M homozygous patients. In addition, the effect of an inflammatory stimulus on Z-variant polymer formation, elicited by Oncostatin M, was investigated. Comparisons of groups were performed using t-test or ANOVA. Non-normally distributed data were assessed by Mann-Whitney U test or the Kruskal-Wallis test, where appropriate. A P value of < 0.05 was considered to be significant. RESULTS: Alpha-1 antitrypsin polymers were found at a higher concentration in the culture medium of ex vivo bronchial epithelial cells from Z-variant homozygotes, compared with M-variant homozygotes (P < 0.01), and detected in the bronchial epithelial cells and submucosa of patient biopsies. Oncostatin M significantly increased the expression of alpha-1 antitrypsin mRNA and protein (P < 0.05), and the presence of Z-variant polymers in ex vivo cells (P < 0.01). CONCLUSIONS: Polymers of Z-alpha-1 antitrypsin form in bronchial epithelial cells, suggesting that these cells may be involved in the pathogenesis of lung emphysema and in bronchial epithelial cell dysfunction.