IGFBP7 and CCT2 are novel lactylation-driven mediators of endothelial-to-mesenchymal transition in idiopathic pulmonary fibrosis.

BackgroundInterstitial lung diseases (ILDs) are a group of heterogeneous lung disorders with variable prognosis. Idiopathic pulmonary fibrosis (IPF) is the main idiopathic interstitial pneumonia and rheumatoid arthritis (RA) is the...

Idiopathic pulmonary fibrosis (IPF) and chronic obstructive pulmonary disease (COPD) are both debilitating lung diseases which can lead to hypoxemia and pulmonary hypertension (PH). Nuclear Factor of Activated T-cells (NFAT) is a transcription factor implicated in the etiology of vascular remodeling in hypoxic PH. We have previously shown that mice lacking the ability to generate Vasoactive Intestinal Peptide (VIP) develop spontaneous PH, pulmonary arterial remodeling and lung inflammation. Inhibition of NFAT attenuated PH in these mice suggesting a connection between NFAT and VIP. To test the hypotheses that: 1) VIP inhibits NFAT isoform c3 (NFATc3) activity in pulmonary vascular smooth muscle cells; 2) lung NFATc3 activation is associated with disease severity in IPF and COPD patients, and 3) VIP and NFATc3 expression correlate in lung tissue from IPF and COPD patients. NFAT activity was determined in isolated pulmonary arteries from NFAT-luciferase reporter mice. The % of nuclei with NFAT nuclear accumulation was determined in primary human pulmonary artery smooth muscle cell (PASMC) cultures; in lung airway epithelia and smooth muscle and pulmonary endothelia and smooth muscle from IPF and COPD patients; and in PASMC from mouse lung sections by fluorescence microscopy. Both NFAT and VIP mRNA levels were measured in lungs from IPF and COPD patients. Empirical strategies applied to test hypotheses regarding VIP, NFATc3 expression and activity, and disease type and severity. This study shows a significant negative correlation between NFAT isoform c3 protein expression levels in PASMC, activity of NFATc3 in pulmonary endothelial cells, expression and activity of NFATc3 in bronchial epithelial cells and lung function in IPF patients, supporting the concept that NFATc3 is activated in the early stages of IPF. We further show that there is a significant positive correlation between NFATc3 mRNA expression and VIP RNA expression only in lungs from IPF patients. In addition, we found that VIP inhibits NFAT nuclear translocation in primary human pulmonary artery smooth muscle cells (PASMC). Early activation of NFATc3 in IPF patients may contribute to disease progression and the increase in VIP expression could be a protective compensatory mechanism.

Simple SummaryEvaluation of the prognostic significance of serial measurements of serum concentrations of Krebs von den Lungen-6 (KL-6) showed that an annual increment in KL-6 exceeding a threshold amount was an independent risk factor for progressive disease and poor prognosis. No literature data are available on long-term KL-6 measurements in the follow-up of IPF patients treated with Nintedanib. The aim of this study is to serially analyze KL-6 in idiopathic pulmonary fibrosis (IPF) patients after 24 months of Nintedanib and to investigate its biomarker potential in patients with IPF and lung cancer with respect to fibrotic hypersensitivity pneumonitis patients, pulmonary fibrosis associated with autoimmune diseases group and healthy controls.Background: Krebs von den Lungen-6 (KL-6) was suggested as ILD biomarker including idiopathic pulmonary fibrosis (IPF). Lung cancer is one of the most severe comorbidity of IPF patients. This study aims to serially analyze KL-6 in IPF patients after 24 months of Nintedanib and to first investigate the biomarker behavior in IPF associated with adenocarcinoma. Materials and methods: One hundred and forty-two ILD patients (median (IQR), 69 (63–75) years; 86 males) were retrospectively enrolled. Serial serum samples were collected from IPF patients before starting antifibrotic therapy and after 12 months. Serum KL-6 levels were measured by KL-6 reagent assay (Fujirebio Europe, UK). Results: Increased KL-6 concentrations were identified in IPF-LC patients than IPF, fibrotic hypersensitivity pneumonitis, and pulmonary fibrosis associated with autoimmune disease groups. A cut-off value was calculated to distinguish IPF and IPF-LC patients. IPF patients monitored for 24 months with Nintedanib showed persisted increased levels of KL-6 with a progressive decline of FVC percentages. Conclusion: This preliminary study offers a first demonstration that very high serum concentrations of KL-6 in IPF-LC patients are associated with poor prognosis. Moreover, serial evaluation of serum KL-6 in IPF patients over 24 months of Nintedanib treatment revealed that most patients experienced a stabilization of lung function parameters and of serum concentrations of KL-6.

ObjectivesMatrix metalloproteinase-8 (MMP-8) promotes lung fibrotic responses to bleomycin in mice. Although prior studies reported that MMP-8 levels are increased in plasma and bronchoalveolar lavage fluid (BALF) samples from IPF patients, neither the bioactive forms nor the cellular sources of MMP-8 in idiopathic pulmonary fibrosis (IPF) patients have been identified. It is not known whether MMP-8 expression is dys-regulated in IPF leukocytes or whether MMP-8 plasma levels correlate with IPF outcomes. Our goal was to address these knowledge gaps.MethodsWe measured MMP-8 levels and forms in blood and lung samples from IPF patients versus controls using ELISAs, western blotting, and qPCR, and assessed whether MMP-8 plasma levels in 73 IPF patients correlate with rate of lung function decline and mortality. We used immunostaining to localize MMP-8 expression in IPF lungs. We quantified MMP-8 levels and forms in blood leukocytes from IPF patients versus controls.ResultsIPF patients have increased BALF, whole lung, and plasma levels of soluble MMP-8 protein. Active MMP-8 is the main form elevated in IPF lungs. MMP-8 mRNA levels are increased in monocytes from IPF patients, but IPF patients and controls have similar levels of MMP-8 in PMNs. Surprisingly, macrophages and airway epithelial cells are the main cells expressing MMP-8 in IPF lungs. Plasma and BALF MMP-8 levels do not correlate with decline in lung function and/or mortality in IPF patients.ConclusionBlood and lung MMP-8 levels are increased in IPF patients. Active MMP-8 is the main form elevated in IPF lungs. Surprisingly, blood monocytes, lung macrophages, and airway epithelial cells are the main cells in which MMP-8 is upregulated in IPF patients. Plasma and BALF MMP-8 levels are unlikely to serve as a prognostic biomarker for IPF patients. These results provide new information about the expression patterns of MMP-8 in IPF patients.

BackgroundNintedanib is an inhibitor of receptor tyrosine kinases, including vascular endothelial growth factor receptor, but its effects on pulmonary hypertension (PH) in idiopathic pulmonary fibrosis (IPF) patients with chronic hypoxia were unclear.MethodsThis study included a nintedanib prospective study and historical control study. In the nintedanib prospective study, pulmonary artery systolic pressure (PASP) measured using transthoracic echocardiography was evaluated at six points during 48 weeks in 16 IPF patients in whom nintedanib was started. In the historical control study, adjusted annual change in PASP was compared between patients treated with (n = 16) and without (n = 15) nintedanib.ResultsIn the nintedanib prospective study, the mean PASP at 48 weeks after starting nintedanib was significantly higher compared to that at baseline. When IPF patients were divided into two groups, IPF patients with or without long-term oxygen treatment (LTOT), mean PASP at 48 weeks was significantly higher than that at baseline only in IPF patients receiving LTOT (P = 0.001). In the historical control study, adjusted annual change in PASP in IPF patients treated with nintedanib was significantly lower than that in patients treated with no antifibrotic agents when considering patients without LTOT (0.26 mmHg vs 7.05 mmHg; P = 0.011).ConclusionsWe found differential effects of nintedanib on PH between IPF patients with or without LTOT. Nintedanib may have a disadvantageous effect on PH in IPF patients with LTOT. Conversely, nintedanib treatment may be beneficial to PH in IPF patients without LTOT.

Familial pulmonary fibrosis is the strongest risk factor for idiopathic pulmonary fibrosis

Serum and BALF YKL-40 levels are predictors of survival in idiopathic pulmonary fibrosis

<b>Introduction:</b> Fibroblast Growth Factors (FGF) are reactivated during Idiopathic Pulmonary Fibrosis (IPF). FGF21, an endocrine FGF that acts through the FGFR1/ beta-klotho (KLB) pathway was shown to prevent liver fibrosis in mice. We asked whether FGF21 was involved in the lung fibrotic process. <b>Methods:</b> We assessed FGF21 and KLB plasma concentration in control (n=11) and IPF patients (n=21) by ELISA. FGF21, FGFR1 and KLB expression was measured by qPCR in total lung samples of IPF and control patients. In vitro, we assessed the effect of FGF21 ± KLB on apoptosis induced by staurosporin and tunicamycin on a murine lung type 2 alveolar epithelial cell line (MLE15 cells) and on primary Human Lung Fibroblasts (HLF) from IPF or control patients. The effect of FGF21 on TGF- ß-induced myofibroblast differentiation, proliferation and migration was assessed on primary HLF. <b>Results:</b> FGF21 median concentration in plasma was increased in IPF patients as compared to controls (39.9 vs 22,9 pg/ml, p=0.001) whereas KLB was decreased (1968.3 vs 2853.8 pg/ml, p=0.001). FGFR1 was expressed similarly in total lungs of IPF and control patients at mRNA level, whereas KLB was decreased in IPF lungs. FGF21 was not expressed. In vitro, FGF21 ± KLB did not influence fibroblast proliferation and migration, and did not modify TGF- ß-induced fibroblast to myofibroblast differentiation. FGF21 associated with KLB decreased MLE15 cells apoptosis, with a decrease in cleaved-PARP expression by western blot, but did not prevent HLF apoptosis. <b>Conclusion:</b> Plasma FGF21 concentration is increased in IPF patients. In vitro, FGF21 inhibited apoptosis of MLE15 cells in vitro when associated to KLB. These data indicate a possible antifibrotic effect of FGF21 in the lung.

Background Idiopathic pulmonary fibrosis (IPF) can combine with emphysema, a condition termed as IPF with emphysema (IPFE). We compared the clinical, radiologic, and physiologic features of IPF and IPFE. Research design and methods Newly diagnosed IPF and IPFE patients were recruited between January 2018 and September 2020. Symptoms, high resolution computed tomography (HRCT), pulmonary function test (PFT) data, composite physiologic index (CPI), gender-age-physiology (GAP) scores, and follow-up data were obtained. Results The IPFE group had greater proportion of male smokers, and of lung cancer cases. The IPFE group had higher VC, FVC FEV1, and lower FEV1/FVC and DLCO and lower percent fibrosis on HRCT. Both groups had similar symptoms and mortality. Mortality rate was associated with inability to perform PFT, CPI, GAP scores, percent fibrosis, VC, FVC, FEV1, and DLCO, serum SCC-Ag and CA125, and anti-fibrotic therapy (≥12 months) in IPF, while it was associated with inability to perform PFT, CPI, percent fibrosis, DLCO, serum CEA, CYFRA21-1 and CA125, and anti-fibrotic therapy (≥12 months) in IPFE. Conclusion IPF and IPFE patients are different in smoking history, physiologic indices, HRCT patterns and prognostic factors, however, they have similar mortality. Anti-fibrotic therapy could improve the survival rate in both IPF and IPFE.

Fibroblast Growth Factors (FGF) are reactivated during Idiopathic Pulmonary Fibrosis (IPF). FGF21, an endocrine FGF that acts through the FGFR1/KLB pathway, was shown to prevent liver fibrosis in mice. We asked whether FGF21 was involved in the lung fibrotic process.&nbsp;Methods :&nbsp;We assessed FGF21 and KLB concentrations in plasma from control (n=11) and IPF patients (n=21) by ELISA. FGF21, FGFR1 and KLB expression was measured by qPCR in total lung samples of IPF and control patients. In vitro, we assessed the effect of FGF21 ± KLB on apoptosis induced by staurosporin, tunicamycin or thapsigargin on a mouse lung type 2 alveolar epithelial cell line (MLE15 cells) and on primary Human Lung Fibroblasts (HLF) from IPF or control patients. The effect of FGF21 on TGF- ß-induced myofibroblast differentiation, proliferation and migration was assessed on primary HLF.&nbsp;Results :&nbsp;FGF21 median concentration in plasma was increased in IPF patients as compared to controls (39.9 vs 22,9 pg/ml, p=0.001) whereas the co-receptor KLB was decreased (1968.3 vs 2853.8 pg/ml, p=0.001). FGF21 and KLB concentrations in plasma were negatively correlated. KLB and FGFR1 were expressed in total lungs of IPF and control patients at mRNA level, whereas FGF21 was not. In vitro, FGF21,&nbsp;with or without&nbsp;KLB, did not modify TGF-ß-induced myofibroblast differentiation, and had no effect on proliferation and migration on HLF. FGF21, alone or associated with KLB, decreased MLE15 cells apoptosis, with a decrease in cPARP expression evaluated by western blot.&nbsp;Conclusion :&nbsp;Plasma FGF21 concentration is increased in IPF patients. In vitro, FGF21 exerts a protective effect on apoptosis on MLE15 cells in vitro. These data indicate a possible antifibrotic effect of FGF21 in the lung.

Elevated BALF concentrations of α- and β-defensins in patients with pulmonary alveolar proteinosis

Idiopathic pulmonary fibrosis (IPF) is well known to be associated with lung cancer. It is important to clarify the clinical and pathological features of lung cancer with IPF in understanding the pathogenesis of lung cancer in IPF patients. We compared clinicopathological factors of lung cancer in patients with and without IPF. A retrospective study was conducted in 711 surgically resected lung cancer patients. Medical records were compared of IPF and non-IPF patients. Of the 711 patients, 53 (7.5%) were IPF patients. Lung cancer in IPF patients was more frequent in elderly male smokers. Most lung cancers in IPF (79%) arose in peripheral areas involving fibrosis (P < 0.01). The incidence of squamous cell carcinoma in the IPF patients (46%) was significantly higher than that in non-IPF patients (22%) (P < 0.01). The incidence of multiple lung cancer in IPF cases (17%) was also significantly higher. These results suggest that IPF has the potential to develop into lung cancer, especially peripheral squamous cell carcinoma. Further molecular analyses are necessary to clarify the relationship between IPF and lung cancer.

Year in review 2016: Interstitial lung disease, pulmonary vascular disease, pulmonary function, paediatric lung disease, cystic fibrosis and sleep.

Background: Evidence for an important heterogeneity in the clinical behaviour of Idiopathic Pulmonary Fibrosis (IPF) is emerging, depending on specific genetic, radiologic and clinical aspects. Despite the observation that IPF incidence is increased in patients exposed to specific organic inhalants (birds and moulds), their impact on prognosis is unknown. Aims and objectives: To compare the prognosis of IPF patients exposed to specific organic inhalants with IPF patients without such exposure and with CHP patients. Methods: we analysed exposure and survival data in IPF and CHP patients recorded in our ILD database. We investigated the impact of specific organic exposure (moulds and birds) and diagnosis on survival using a Cox proportional hazard model. Results: 293 patients were included. From the 244 patients diagnosed with IPF, 73 were exposed. 49 patients were diagnosed with CHP. Median survival (MS) of IPF patients exposed to specific organic exposure was 84 months, which was significantly different from both not-exposed IPF patients (MS = 43 months, p = 0.002) and CHP patients (MS = 157 months, p = 0.037). Conclusion: Exposure to specific organic inhalants was associated with a better survival in IPF patients. This observation strengthens the emerging evidence of an important clinical heterogeneity in IPF.

Background:Connective tissue disease (CTD) may be observed during the course of idiopathic pulmonary fibrosis (IPF). However, clinical factors associated with the development of CTD in patients with IPF have not yet been identified. These factors might be valuable clues for determining the pathogenesis of pulmonary fibrosis in patients with CTD. We hypothesize that some IPF patients have a clinically significant association with autoimmunity, and that autoantibodies are important biomarkers for identifying these patients.Objectives:Based on this hypothesis, we investigated whether the serology criteria (anti-neutrophil cytoplasmic antibody (ANCA) or autoantibodies that met the serology criteria for interstitial pneumonitis with autoimmune features (IPAF)) were associated with the development of CTD during the clinical course of IPF in the patients from our previous study(1), with a particular focus on which antibodies have a significant association with the development of CTD.Methods:We retrospectively reviewed the records of 527 patients with a first diagnosis of IPF between January 2007 and March 2014, and investigated the length of time from first visit to the clinic for IPF diagnosis (baseline) to CTD diagnosis by an expert rheumatologist in patients with IPF. Multivariable Cox proportional-hazards models with backward elimination were used to investigate the risk factors for the development of CTD.Results:CTD developed in 15 patients at a median of 2.1 years (range 1.2 to 4.8) after IPF diagnosis. All these patients had ANCA or autoantibodies that met the serology criteria for IPAF. A significant number of IPF patients with high titers of RF, ACPA or MPO-ANCA tested at first visit to the clinic progressed to CTD(Figure 1). Survival duration for IPF patients with progression to CTD was 5.3 [3.8; 6.7] years, which was significantly longer than for the IPF patients without progression to CTD (2.9 [1.7; 4.8], p = 0.001). Independent risk factors for development of CTD in IPF patients included female gender (adjusted hazard ratio (HR) 5.319, p = 0.0082), titer of rheumatoid factor (RF) (adjusted HR 1.006, p = 0.022), titer of anti-citrullinated protein antibody (ACPA) (adjusted HR 1.009, p = 0.0011), and titer of myeloperoxidase (MPO) ANCA (adjusted HR 1.02, p &lt; 0.0001).Figure 1.Connective tissue disease development in each autoantibody positive IPF patient. ACPA = anti–citrullinated protein antibody; ANA = antinuclear antibody; CTD = connective tissue disease; MPA = microscopic polyangiitis; PAN = polyarteritis nodosa; RA = rheumatoid arthritis; RF = rheumatoid factor; UCTD = Undifferentiated connective tissue disease; SjS = Sjögren’s syndrome.Conclusion:We observed development of CTD in IPF patients with ANCA or autoantibodies that met the IPAF serology criteria. Among these autoantibodies, RF, ACPA, and MPO-ANCA were significantly associated with the development of CTD in IPF patients. Progression to CTD is uncommon in IPF patients, but a significant number of IPF patients with high titers of RF, ACPA or MPO-ANCA progressed to connective tissue disease. IPF with high titers of RF, ACPA or MPO-ANCA might be the initial clinical manifestation of connective tissue disease. Further studies are needed to investigate the role of RF, ACPA, and MPO-ANCA in development of pulmonary fibrosis.