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Innate Immunity Group, IdiPAZ, La Paz University Hospital, Madrid, SpainTumor Immunology Lab, IdiPAZ, La Paz University Hospital, Madrid, SpainCenter for Biomedical Research Network, CIBEres, Spain
Innate Immunity Group, IdiPAZ, La Paz University Hospital, Madrid, SpainTumor Immunology Lab, IdiPAZ, La Paz University Hospital, Madrid, SpainCenter for Biomedical Research Network, CIBEres, Spain
Center for Biomedical Research Network, CIBEres, SpainRespiratory Diseases Group, Respiratory Service, La Paz University Hospital, IdiPAZ, Madrid, Spain
Center for Biomedical Research Network, CIBEres, SpainRespiratory Diseases Group, Respiratory Service, La Paz University Hospital, IdiPAZ, Madrid, Spain
Innate Immunity Group, IdiPAZ, La Paz University Hospital, Madrid, SpainTumor Immunology Lab, IdiPAZ, La Paz University Hospital, Madrid, SpainCenter for Biomedical Research Network, CIBEres, Spain
P. aeruginosa is associated with PD-L1 overexpression in patients with CF.
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Endotoxin Tolerance is a CF hallmark in patients colonized by P. aeruginosa.
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Translocation of P. aeruginosa LPS induces PD-L1 expression and Endotoxin Tolerance.
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Colistimethate ameliorates the effects of LPS translocation
Abstract
Background
Cystic fibrosis (CF) is an endotoxin tolerance (ET)-related disease. Given that increased PD-L1 has been reported in ET, its expression and physiological effects on cystic fibrosis monocytes should be studied.
Methods
We analyzed the phenotype and ex vivo response of immune system cells in 32 patients with CF, 19 of them colonized by Pseudomonas aeruginosa. An in vitro model was developed of Pseudomonas aeruginosa colonization using purified lipopolysaccharides (LPS) from one of the most prevalent strains in patients with CF (a CF-adapted Pseudomonas aeruginosa ST395 clone). Changes in the immune response, including cytokine production and T-lymphocyte proliferation, as well as expression of PD-L1, were evaluated.
Results
PD-L1 was overexpressed in the monocytes of patients with CF compared with healthy volunteers, and levels of this immune checkpoint were associated with Pseudomonas aeruginosa colonization. In addition, patients with Pseudomonas aeruginosa colonization showed a patent ET status, including poor inflammatory response, reduced HLA-DR expression and T-lymphocyte proliferation impairment. PD-L1/PD-1 blocking assays reverted the impaired adaptive response. Ultimately, monocytes from healthy volunteers cultured in the presence of the clinically relevant strain of Pseudomonas aeruginosa or serum collected from patients with CF colonized by Pseudomonas aeruginosa reproduced the previous observed features.
Conclusions
Pseudomonas aeruginosa colonization in patients with CF was associated with PD-L1 overexpression and impaired T cell response, and LPS from this pathogen induced the observed phenotype. Our findings open new avenues for the use of anti-PD-1/PD-L1 immunotherapy in patients with CF who are colonized by Pseudomonas aeruginosa.
]. Patients with CF have high infection rates compared with the normal population. Infections in patients with CF are associated with disease complications, and the mortality of these patients is usually the result of chronic lower airway bacterial colonization [
]. According to a number of studies, Pseudomonas aeruginosa is the most prevalent Gram-negative bacterial pathogen in patients with CF, reaching rates of up to 49.56% [
]. In fact, current efforts are focused on the development of vaccines against Pseudomonas aeruginosa. Thus, several vaccines dedicated to patients who have not yet been colonized or who have intermittent colonization have been evaluated without success [
Johansen HK, Gotzsche PC. Vaccines for preventing infection with Pseudomonas aeruginosa in cystic fibrosis. The cochrane database of systematic reviews. 2015:Cd001399.
Pulmonary and systemic pharmacokinetics of inhaled and intravenous colistin methanesulfonate in cystic fibrosis patients: targeting advantage of inhalational administration.
]. Therefore, identifying those patients with a suppressed immune system would be advantageous in terms of choosing treatment and preventing infection-derived complications. Although most studies have been focused on immune system changes at a local pulmonary level [
], we have previously reported the role of peripheral cells in patients with CF. Due to the translocation of bacterial products, mainly lipopolysaccharides (LPS) [
Potent phagocytic activity with impaired antigen presentation identifying lipopolysaccharide-tolerant human monocytes: demonstration in isolated monocytes from cystic fibrosis patients.
Potent phagocytic activity with impaired antigen presentation identifying lipopolysaccharide-tolerant human monocytes: demonstration in isolated monocytes from cystic fibrosis patients.
]. In fact, a clinical trial has been approved to examine the potential of circulating blood cell monocytes as a predictive biomarker of osteoporosis in patients with CF (NCT03492567).
In recent years, crosstalk between programmed cell death ligand 1 (PD-L1) and its receptor, PD-1, has emerged as an important mechanism in the immune response, not only in cancer but also in infectious diseases. Several studies have indicated the PD-L1/PD-1 axis as a promising therapy in an increasing number of clinical contexts [
]. In addition, treatment with anti-PD-L1 antibodies protects against infection due to increased interferon-γ secretion in CD8+ T cells and wound-draining lymph nodes after burn wound sepsis [
]. We have demonstrated that PD-L1 is a robust ET marker, and its control appears to be mediated by hypoxia-inducible factor-1α in a cohort of patients with sepsis [
]. However, PD-L1 expression in the context of CF has not been explored, especially in those patients colonized by Pseudomonas aeruginosa.
Herein, we have examined the PD-L1 and PD-1 levels on monocytes and lymphocytes, respectively, in 32 patients with CF. We have also analyzed the potential association between the expression of PD-L1 and the clinical parameters available, such as Pseudomonas aeruginosa colonization and type of infection. Finally, we have corroborated the results obtained in patients using an in vitro model.
2. Materials and methods
2.1 Study design
The study was conducted in accordance with the ethical guidelines of the 1975 Declaration of Helsinki and was approved by the Committee for Human Subjects of La Paz University Hospital. All the participants provided written consent for the study. Patients with CF were prospectively enrolled from the Pneumology Department of La Paz University Hospital, Madrid, Spain. Patients were nonsmoking adults who fulfilled the diagnostic criteria for CF (clinical phenotype, sweat testing, and CFTR genotyping [
]). Exclusion criteria included a history of chronic obstructive pulmonary disease, asthma, or other active lung disease, mental or physical handicap, or other significant diseases such as diabetes mellitus, congestive heart failure, ischemic or valvular cardiopathy, or neuromuscular disease. None of the patients had experienced a lung exacerbation of respiratory tract infection within the previous 4 weeks, and none had had received oral corticosteroid therapy for at least 3 months before the study. The clinical data on the patients included in the study are summarized in Supplementary Table S1. Age and sex-matched healthy volunteers (HVs) were also included as controls.
2.2 Reagents
Roswell Park Memorial Institute (RPMI) medium (Invitrogen) was used for the cell cultures. The LPS, from Escherichia coli O111:B4, was obtained from Sigma-Aldrich. Carboxyfluorescein succinimidyl ester (CFSE) for the proliferation assays was purchased from Thermo Fisher. The CF-adapted ST395 P. aeruginosa clone was isolated from a patient who had been colonized during 13 years. LPS was obtained as previously described [
]. Colistimethate Sodium (polymyxin E) was purchased from Genéricos Españoles (GES) and dissolved in RPMI medium. The lymphocyte stimulus pokeweed (PWD) was purchased from Sigma-Aldrich. To inhibit PD-L1/PD-1 interaction, a fully human IgG4 (S228P) anti-PD-1 receptor-blocking monoclonal antibody was used (Bristol-Myers Squibb). All the reagents used for the cell cultures were endotoxin-free, as assayed with the Limulus amebocyte lysate test (Cambrex).
2.3 Cytokine production in ex vivo LPS stimulation
Fresh blood from patients with CF and paired HV were diluted in RPMI medium in a 1:1 proportion. The cells were then stimulated with 10 ng/mL of LPS. After 3 and 6 h, the cultures were centrifuged at 18,000 g for 10 min and supernatants were collected for their analysis by cytometric bead array (CBA). Tumor necrosis factor alpha (TNFα), interleukin (IL)10, and IL6 protein levels in the culture supernatants were determined using the Human Inflammatory CBA kit (BD Biosciences), following the manufacturer's protocol. Supernatants were analyzed by flow cytometry using a BD FACSCalibur flow cytometer (BD Biosciences).
2.4 Peripheral blood mononuclear cell isolation and flow cytometry analysis
The peripheral blood mononuclear cells (PBMCs) from patients with CF and the controls were isolated using Ficoll-Plus gradient (GE Healthcare Bio-Sciences) [
Potent phagocytic activity with impaired antigen presentation identifying lipopolysaccharide-tolerant human monocytes: demonstration in isolated monocytes from cystic fibrosis patients.
]. The cells were labeled with allophycocyanin (APC)-conjugated anti-human CD14, fluorescein isothiocyanate-conjugated anti-human HLA-DR, APC-conjugated anti-human CD3 (all from Immunostep, Spain); and phycoerythrin-conjugated anti-human PD-L1 (Miltenyi Biotec, USA). Matched isotype antibodies were used as negative controls. The cells were incubated for 30 min at 4 °C in the dark. The data were acquired by flow cytometry using a BD FACSCalibur flow cytometer (BD Biosciences) and analyzed with FlowJo vX.0.7 software (FlowJo, LLC). Gating strategy is show on Supplementary Fig. S1.
2.5 T cell proliferation assays
Proliferation was analyzed by flow cytometry of CFSE labeled cells as reported previously [
Potent phagocytic activity with impaired antigen presentation identifying lipopolysaccharide-tolerant human monocytes: demonstration in isolated monocytes from cystic fibrosis patients.
]. The PBMCs were then labeled and cultured in a 96-well plate with fresh RPMI medium and were stimulated or not with PWD (2.5 μg/mL) and treated with 5 μg/mL of a fully human IgG4 (S228P) anti-PD-1 receptor-blocking monoclonal antibody (Bristol-Myers Squibb).
2.6 Statistical analysis
The number of experiments analyzed is indicated in each figure. In summary, statistical significance was calculated using an unpaired Mann–Whitney U test or a paired Wilcoxon t-test, as appropriate, depending on the specific assay. The statistical significance was set at p < .05, and the statistical analyses were conducted using Prism 6.0 (GraphPad) and SPSS version 23 (IBM) software.
3. Results
3.1 PD-L1 on monocytes and PD-1 on lymphocytes are overexpressed in patients with cystic fibrosis
Given that circulating monocytes from patients with CF have traditionally been associated with an ET phenotype [
Potent phagocytic activity with impaired antigen presentation identifying lipopolysaccharide-tolerant human monocytes: demonstration in isolated monocytes from cystic fibrosis patients.
], we evaluated the expression of this immune checkpoint ligand. We found PD-L1 overexpression on CF monocytes as well as high levels of sPD-L1 in plasma from CF patients (Fig. 1A ). PD-1 was also overexpressed in both CD4 and CD8 lymphocytes from patients (Fig. 1B). In full agreement, T lymphocyte (CD3) proliferation was significantly reduced in cells from patients with CF compared with HVs (Fig. 1C). This effect was negated when PD-L1/PD-1 crosstalk was blocked using an anti-PD-1 antibody (Fig. 1C).
Fig. 1PD-L1 on monocytes and PD-1 on T lymphocytes are overexpressed in patients with CF, corresponding to an impaired T-cell response.
(A) PD-L1 expression on monocytes from patients with CF and HVs was measured by flow cytometry. Percentage of PD-L1+ on CD14+ gated cells is shown on left panel. Soluble PD-L1 (sPD-L1) in sera from patients with CF and HVs was measured by ELISA and summarized on right panel. (n = 32 patients with CF; n = 15 HVs). (B) PD-1 expression on T lymphocytes from patients with CF and HVs were measured by flow cytometry. Percentages of PD-1+ on both CD4+ and CD8+ gated cells are shown (n = 32 patients with CF; n = 15 HVs). (C) T cell proliferation in patients with CF and HVs was measured with CFSE proliferation assay by flow cytometry. Percentage of proliferation (defined as CFSEdim) on gated CD3+ cells is shown. (n = 16 patients with CF randomly selected; n = 7 HVs) *, p < .05; **, p < .01; ***, p < .001 vs. HVs using an unpaired t-test. Data are expressed as the mean ± SD.
3.2 PD-L1 overexpression on monocytes and T cell proliferation impairment is associated with Pseudomonas aeruginosa colonization in patients with CF
Patients were classified in subgroups according to relevant parameters: sex, CFTR mutation, age at diagnosis, number of infections in the last year, airflow limitation, body mass index and microbiological data. We then evaluated the influence of these parameters on the PD-L1 expression on their monocytes. No correlations were found with sex, mutations, age at diagnosis, airflow limitation, number of infections in the last year, and body mass index (Supplementary Fig. S2). In contrast, colonization by Pseudomonas aeruginosa correlated not only with PD-L1 overexpression on monocytes (Fig. 2A) but also for sPD-L1 levels in serum (Fig. 2B). Note that patients with Staphylococcus aureus colonization but not Pseudomonas aeruginosa did not exhibit PD-L1 overexpression (Supplementary Fig. S3). In agreement, T cell proliferation impairment was greater in cells from patients with Pseudomonas aeruginosa than in patients free from this pathogen. As expected, an anti-PD-1 antibody reverted the adaptive immune response impairment (Fig. 2C).
3.3 Monocytes from patients with CF colonized by Pseudomonas aeruginosa exhibited a patent ET phenotype
Previously, we reported that the monocytes of patients with CF are locked into a refractory state in which they are unable to respond to an endotoxin stimulus [
Potent phagocytic activity with impaired antigen presentation identifying lipopolysaccharide-tolerant human monocytes: demonstration in isolated monocytes from cystic fibrosis patients.
]. Here, we have explored the main ET hallmarks in monocytes from patients with CF classified according to the presence of Pseudomonas aeruginosa colonization. Although TNFα and IL6 showed a patent downregulation after an ex vivo LPS challenge of these cells isolated from colonized patients (Fig. 3A and B), IL10 exhibited significant activation (Fig. 3C). Analysis of HLA-DR levels revealed a diminished expression in colonized patients (Fig. 3D). In order to corroborate whether the ET phenotype and PD-L1 overexpression occurs in the same cell population, we assessed a Spearman correlation test between basal PD-L1 and HLA-DR increment after stimulation. As Supplementary Fig. S4 illustrates, we have observed a negative correlation between these two parameters, in agreement to the observed in sepsis [
]. Note that no differences in plasma cytokines were found between groups, ruling out the possibility of chronic basal inflammation in patients (Supplementary Fig. S5). Collectively, these data suggest a deeper refractory status in those patients with CF colonized by Pseudomonas aeruginosa. Note that none of the other clinical parameters have been shown to have a clear association with cytokine production (data not shown).
3.4 LPS isolated from a clinically relevant Pseudomonas aeruginosa strain and sera from patients colonized by this pathogen induced PD-L1 overexpression on naïve monocytes
To evaluate the role of LPS in the observed phenomenon, we isolated LPS from a clinically relevant Pseudomonas aeruginosa strain. Next, human monocytes from healthy donors were challenged with the isolated LPS and PD-L1 overexpression was verified (Fig. 4A ). Similar results were obtained in the presence of sera from patients colonized by Pseudomonas aeruginosa (Fig. 4B, left panel). The effect was negated when colistimethate (polymyxin E; PmE, an LPS scavenger [
]) was added to the culture (Fig. 4B, right panel). In line, sPD-L1 levels detected in sera from patients colonized by Pseudomonas aeruginosa were lower in those under inhaled colistimethate sodium (Fig. 5) suggesting a role for colistimethate in the Pseudomonas aeruginosa LPS translocation to peripheral blood.
Fig. 2Pseudomonas aeruginosa colonization in patients with CF is associated with higher PD-L1 overexpression.
(A) PD-L1 expression on monocytes from patients with CF by flow cytometry; patients were classified according the presence of Pseudomonas aeruginosa in their sputum. Percentage of PD-L1+ on CD14+ gated cells is shown (n = 32 patients with CF; n = 19 CF patients with Pseudomonas aeruginosa; n = 15 HVs). (B) Soluble PD-L1 (sPD-L1) in sera from patients with CF and HVs was measured by ELISA (n = 32 patients with CF; n = 19 CF patients with Pseudomonas aeruginosa; n = 15 HVs). (C) T cell proliferation in patients with CF was measured with CFSE proliferation assay. Percentage of proliferation (defined as CFSEdim) on gated CD3+ cells is shown (n = 16 patients with CF randomly selected and n = 7 HVs). “+”, patients with Pseudomonas aeruginosa colonization; “-”, patients without Pseudomonas aeruginosa colonization. *, p < .05 vs. noncolonized patients, using an unpaired t-test. Data are expressed as the mean ± SD.
Whole blood from patients with CF was stimulated with 10 ng/mL of LPS for 3 h. Inflammatory cytokine production was measured by CBA. HLA-DR expression was analyzed by flow cytometry. Concentrations of TNFα (A), IL-6 (B) and IL-10 (C) supernatants are shown. HLA-DR expression data as fold (ratio between mean intensity of fluorescence (MIF) on LPS stimulated cells divided by MIF on unstimulated cells) are show (D). “+”, patients with Pseudomonas aeruginosa colonization (n = 19); “-”, patients without Pseudomonas aeruginosa colonization (n = 13). *, p < .05 vs. noncolonized patients, using an unpaired t-test. Data are expressed as the mean ± SD.
Fig. 4LPS and sera from patients with CF colonized by Pseudomonas aeruginosa induce PD-L1 expression in naïve monocytes, which is reverted by polymyxin E.
(A) Monocytes from HVs were stimulated with 100 ng/mL of LPS from a Pseudomonas aeruginosa ST395 clone, and PD-L1 expression was analyzed by flow cytometry. Percentage of PD-L1+ on gated CD14+ cells is shown. (B) Monocytes from HVs were stimulated with 10% patient sera pretreated or not with polymyxin E (150 IU/mL); PD-L1 expressions were analyzed by flow cytometry. Percentage of PD-L1+ on gated CD14+ cells is shown. “+”, treated with sera from patients with Pseudomonas aeruginosa colonization (n = 5); “-” treated with sera from patients without Pseudomonas aeruginosa colonization (n = 5). *, p < .05; ***, p < .001 using a paired t-test. The data are pooled from four independent experiments and are expressed as the mean ± SEM.
Soluble PD-L1 in sera from patients with CF classified according to Pseudomonas aeruginosa colonization and inhaled colistimethate sodium treatment was measured by ELISA (n = 32 patients with CF; n = 15 HVs). **: p < .01 versus noncolonized patients, using an unpaired t-test.
]. Thus, identifying those patients with immunosuppression could confer an advantage in choosing optimal treatment and preventing infection-derived complications. Although basal chronic inflammation and neutrophilia in the lungs of CF patients clearly leads to a decline in lung function [
Reynolds CJ, Quigley K, Cheng X, Suresh A, Tahir S, Ahmed-Jushuf F, et al. Lung defense through interleukin-8 carries a cost of chronic lung remodeling and impaired function. Am J Respir Cell Mol Biol[0:null].
Here, we have observed no basal inflammatory differences comparing the sera of patients and HVs regarding IL6 and TNFα cytokine levels in sera, which is in accordance with others' observations [
]. However, to evaluate immune status properly, we must study not only basal immunological status but also the capability to generate an appropriate immune response against an insult. We have previously demonstrated that the monocytes of patients with CF are in an ET status in which they were unable to respond properly against LPS stimulus [
Potent phagocytic activity with impaired antigen presentation identifying lipopolysaccharide-tolerant human monocytes: demonstration in isolated monocytes from cystic fibrosis patients.
]. According to our findings, PD-L1 overexpression on monocytes appears to be a novel characteristic of the CF circulating immune system. Moreover, the data regarding T-cell proliferation with anti PD-1 monoclonal antibodies not only confirms the functional implication of this observation, but also opens the possibility of evaluating this novel therapeutic option for treating chronic infections in some patients with CF, increasing their clinical management. Ingersoll's et al. observed how a PD-L1 upregulation on CF neutrophils did not produce T cell suppression [
]. On the contrary, our data illustrated a crucial role of PD-L1 in T cell suppression. This inconsistency took place when the cohort was divided into two groups, according to Pseudomonas aeruginosa colonization (please compare Fig. 2C and Fig. 1C). Moreover, monocytes, but not neutrophils, were able to express antigen presentation molecules such as HLA-DR, which we have shown to be deregulated in this pathology. Additionally, it has been reported that PWD-induced T cell proliferation requires monocytes activation [
]. Altogether, these data indicate that not all patients, but rather those with P. aeruginosa colonization would be beneficiaries of anti-PD-1 therapy. Along these lines, PD-L1 levels on monocytes could be better predictors for immunotherapy efficacy than PD-L1 expression on neutrophils, which is in line with what is observed in cancer [
In a previous study with a smaller cohort including all patients with Gram-negative colonization, we have proposed that LPS translocation to the bloodstream is a putative explanation for the ET observed in CF [
]. Herein, we have a larger cohort including a wider variety of colonizing bacteria, which has allowed us to classify patients with CF into various subgroups according to the microorganism in their sputum.
On one hand, our results showed that only Pseudomonas aeruginosa colonization, and no other clinical parameters, exhibited a strong association with PD-L1 levels in CF. Moreover, the patients with Pseudomonas aeruginosa colonization showed a greater ET. Although the immune system effects of some intrinsic features of CF such CFTR mutations cannot be totally ruled out [
], our data indicated the relevance of the Pseudomonas aeruginosa colonization in this context. In agreement, other groups have reported altered adaptive responses in P. aeruginosa + patients mainly characterized by disrupted balance of Th17 and Th2 responses [
]. On the other hand, our in vitro model reinforced the idea of LPS translocation as an explanation of the features found in patients. Monocytes from HVs stimulated with isolated P. aeruginosa LPS increased PD-L1 expression in a time-dependent manner. Therefore, sera from patients with Pseudomonas aeruginosa were shown to induce more PD-L1 expression in HV monocytes than in sera from patients without colonization; this induction was reduced when the samples were pretreated with the LPS scavenger polymyxin E. Accordingly, we detected lower levels of sPD-L1 sera from patients treated with inhaled sodium colistimethate (polymyxin E) than from those treated with other antibiotics. This last datum suggests that colistimethate could reduce the effects of LPS translocation in patients with CF who have Pseudomonas aeruginosa colonization, although a prospective, randomized, double-blind clinical trial must be performed to confirm this hypothesis.
The results presented here support the hypothesis that translocated LPS on Pseudomonas aeruginosa-colonized patients with CF causes both PD-L1 overexpression and an ET phenotype. Moreover, the antibody-mediated blockade of PD-L1 and PD-1 in these patients reversed their impaired T cell response. Altogether, our results suggest the possibility of studying antibiotics and antiPD-1/PD-L1 antibody combination as a new potential treatment to provide clinical benefits to patients with P. aeruginosa colonization.
Funding
This work was supported by grants from the “Instituto de Salud Carlos III” (ISCIII), “Fondos de Investigación Sanitarias” (FIS) and FEDER [PI14/01234 and PIE15/00065] to ELC, and a grant from “Comunidad de Madrid” [PEJ15/BIO/AI-0021] to JAO.
The following are the supplementary data related to this article.
PD-L1 expression on monocytes from CF patients and HV was measured by flow cytometry. Percentages of PD-L1+ on CD14+ gated cells from patients classified according their sex (A), type of CFTR mutation (B), age at diagnosis (C), airflow limitation (D), number of infections during last year (E) and body mass index (F) are shown (n = 32 CF patients; n = 15 HV). Airflow limitation was defined as FE1V/FVC ratio < 70. Data are expressed as the mean ± SD.
PD-L1 expression on monocytes from patients with CF according to their bacterial colonization.
PD-L1 expression on monocytes from CF patients and HV was measured by flow cytometry. Patients were classified in four groups according the presence of Staphylococcus aureus and Pseudomonas aeruginosa in their sputum. “None”, CF patients without bacteria colonization; “SA”, patients with Staphylococcus aureus but not Pseudomonas aeruginosa colonization; “PA”, patients with Pseudomonas aeruginosa but not Staphylococcus aureus colonization; “Both”, patients with both Pseudomonas aeruginosa and Staphylococcus aureus colonization. *: p < .05; ***, p < .001 and n. s., no significant using an unpaired t-test. Data are expressed as the mean ± SD.
PD-L1 expression on monocytes from patients with CF correlates with endotoxin tolerance phenotype.
Percentages of PD-L1+ cells and HLA-DR expression on gated CD14+ cells from patients with CF (n = 32) were analyzed by FACS. The PD-L1 data is basal in non-stimulated cells. HLA-DR expression data are shown as fold (ratio between mean intensity of fluorescence (MIF) on LPS stimulated cells divided by MIF on unstimulated cells). The correlation between PD-L1 and Fold of HLA-DR is shown.
Basal cytokines levels on plasma from patients with CF.
Cytokines levels on plasma from CF patients were measured. Concentrations of TNFα (A), IL-6 (B) and IL-10 (C) are shown (n = 32 patients with CF; n = 19 CF patients with Pseudomonas aeruginosa; n = 15 HVs). “+”, patients with Pseudomonas aeruginosa colonization; “-”, patients without Pseudomonas aeruginosa colonization. Data are expressed as the mean ± SD.
Summary of clinical data in all patients included in the study. M, Male; F, Female; −, not included on medical history; MSSA, methicillin-sensitive Staphylococcus aureus; MRSA, methicillin-resistent Staphylococcus aureus.
Conflict of interest statement
The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
Acknowledgements
We thank the blood donor service from La Paz University Hospital; Aurora Muñoz for technical assistance; Ana Sierra from Cytostatic Service for her collaboration; and ServingMed.com for the editing of the manuscript.
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Translocated LPS might cause endotoxin tolerance in circulating monocytes of cystic fibrosis patients.
Johansen HK, Gotzsche PC. Vaccines for preventing infection with Pseudomonas aeruginosa in cystic fibrosis. The cochrane database of systematic reviews. 2015:Cd001399.
Pulmonary and systemic pharmacokinetics of inhaled and intravenous colistin methanesulfonate in cystic fibrosis patients: targeting advantage of inhalational administration.
Potent phagocytic activity with impaired antigen presentation identifying lipopolysaccharide-tolerant human monocytes: demonstration in isolated monocytes from cystic fibrosis patients.
Reynolds CJ, Quigley K, Cheng X, Suresh A, Tahir S, Ahmed-Jushuf F, et al. Lung defense through interleukin-8 carries a cost of chronic lung remodeling and impaired function. Am J Respir Cell Mol Biol[0:null].
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