Vijay D'Souza

Vitamin D deficiency contributes directly to the acute respiratory distress syndrome (ARDS)

Rationale Vitamin D deficiency has been implicated as a pathogenic factor in sepsis and intensive therapy unit mortality but has not been assessed as a risk factor for acute respiratory distress syndrome (ARDS). Causality of these associations has never been demonstrated. Objectives To determine if ARDS is associated with vitamin D deficiency in a clinical setting and to determine if vitamin D deficiency in experimental models of ARDS influences its severity. Methods Human, murine and in vitro primary alveolar epithelial cell work were included in this study. Findings Vitamin D deficiency (plasma 25(OH)D levels <50 nmol/L) was ubiquitous in patients with ARDS and present in the vast majority of patients at risk of developing ARDS following oesophagectomy. In a murine model of intratracheal lipopolysaccharide challenge, dietary-induced vitamin D deficiency resulted in exaggerated alveolar inflammation, epithelial damage and hypoxia. In vitro, vitamin D has trophic effects on primary human alveolar epithelial cells affecting >600 genes. In a clinical setting, pharmacological repletion of vitamin D prior to oesophagectomy reduced the observed changes of in vivo measurements of alveolar capillary damage seen in deficient patients. Conclusions Vitamin D deficiency is common in people who develop ARDS. This deficiency of vitamin D appears to contribute to the development of the condition, and approaches to correct vitamin D deficiency in patients at risk of ARDS should be developed. Trial registration UKCRN ID 11994.

Vitamin D deficiency in human and murine sepsis

Objectives: Vitamin D deficiency has been implicated as a pathogenic factor in sepsis and ICU mortality but causality of these associations has not been demonstrated. To determine whether sepsis and severe sepsis are associated with vitamin D deficiency and to determine whether vitamin D deficiency influences the severity of sepsis. Design, Setting, and Patients: Sixty-one patients with sepsis and severe sepsis from two large U.K. hospitals and 20 healthy controls were recruited. Murine models of cecal ligation and puncture and intratracheal lipopolysaccharide were undertaken in normal and vitamin D deficient mice to address the issue of causality. Measurements and Main Results: Patients with severe sepsis had significantly lower concentrations of 25-hydroxyvitamin D3 than patients with either mild sepsis or age-matched healthy controls (15.7 vs 49.5 vs 66.5 nmol/L; p = 0.0001). 25-hydroxyvitamin D3 concentrations were significantly lower in patients who had positive microbiologic culture than those who were culture negative (p = 0.0023) as well as those who died within 30 days of hospital admission (p = 0.025). Vitamin D deficiency in murine sepsis was associated with increased peritoneal (p = 0.037), systemic (p = 0.019), and bronchoalveolar lavage (p = 0.011) quantitative bacterial culture. This was associated with reduced local expression of the cathelicidin-related antimicrobial peptide as well as evidence of defective macrophage phagocytosis (p = 0.029). In the intratracheal lipopolysaccharide model, 1,500 IU of intraperitoneal cholecalciferol treatment 6 hours postinjury reduced alveolar inflammation, cellular damage, and hypoxia. Conclusions: Vitamin D deficiency is common in severe sepsis. This appears to contribute to the development of the condition in clinically relevant murine models and approaches to correct vitamin D deficiency in patients with sepsis should be developed.

ResolvinD 1 stimulates epithelial wound repair and inhibits TGF- β -induced EMT whilst reducing fibroproliferation and collagen production

Acute and chronic inflammatory lung diseases are often associated with epithelial cell injury/loss and fibroproliferative responses. ResolvinD1 (RvD1) is biosynthesized during the resolution phase of inflammatory response and exerts potent anti-inflammatory and promotes resolution of inflammatory lung diseases. The aim of this study was to investigate whether RvD1 exerts protective effects on alveolar epithelial cell function/differentiation and protects against fibroproliferative stimuli. Primary human alveolar type II cells were used to model the effects of RvD1 in vitro upon wound repair, proliferation, apoptosis, transdifferentiation, and epithelial–mesenchymal transition (EMT). Effects of RvD1 upon primary human lung fibroblast proliferation, collagen production, and myofibroblast differentiation were also examined. RvD1 promoted alveolar type II (ATII) cell wound repair and proliferation. RvD1 protected ATII cells against sFas-ligand/TNF-α-induced apoptosis and inhibition on cell proliferation and viability. RvD1 promoted ATII cells transdifferentiation. Moreover, we demonstrate that RvD1 inhibited EMT in response to TGF-β. Furthermore RvD1 inhibited human lung fibroblast proliferation, collagen production, and myofibroblast differentiation induced by both TGF-β and bronchoalveolar lavage fluid from acute respiratory distress syndrome (ARDS) patients. The effects of RvD1 were PI3-kinase dependent and mediated via the resolvin receptor. RvD1 seems to promote alveolar epithelial repair by stimulating ATII cells wound repair, proliferation, reducing apoptosis, and inhibiting TGF-β-induced EMT. While RvD1 reduced fibroproliferation, collagen production, and myofibroblast differentiation. Together, these results suggest a potential new therapeutic strategy for preventing and treating chronic diseases (such as idiopathic pulmonary fibrosis) as well as the fibroproliferative phase of ARDS by targeting RvD1 actions that emphasizes natural resolution signaling pathways.

Lipoxin A4 promotes lung epithelial repair whilst inhibiting fibroblast proliferation

Therapy that promotes epithelial repair whilst protecting against fibroproliferation is critical for restoring lung function in acute and chronic respiratory diseases. Primary human alveolar type II cells were used to model the effects of lipoxin A4 in vitro upon wound repair, proliferation, apoptosis and transdifferention. Effects of lipoxin A4 upon primary human lung fibroblast proliferation, collagen production, and myofibroblast differentiation were also assessed. Lipoxin A4 promoted type II cell wound repair and proliferation, blocked the negative effects of soluble Fas ligand/tumour necrosis factor α upon cell proliferation, viability and apoptosis, and augmented the epithelial cell proliferative response to bronchoaveolar lavage fluid (BALF) from acute respiratory distress syndrome (ARDS). In contrast, Lipoxin A4 reduced fibroblast proliferation, collagen production and myofibroblast differentiation induced by transforming growth factor β and BALF from ARDS. The effects of Lipoxin A4 were phosphatidylinositol 3′-kinase dependent and mediated via the lipoxin A4 receptor. Lipoxin A4 appears to promote alveolar epithelial repair by stimulating epitheial cell wound repair, proliferation, reducing apoptosis and promoting trans-differentiation of alveolar type II cells into type I cells. Lipoxin A4 reduces fibroblast proliferation, collagen production and myofibroblast differentiation. These data suggest that targeting lipoxin actions may be a therapeutic strategy for treating the resolution phase of ARDS. Lipoxin A4 promotes epithelial repair while inhibiting fibroproliferation in vitro in human alveolar epithelial cells http://ow.ly/SxMu301cBRP

S49 Is the development of acute lung injury influenced by increased levels of IL17 as a result of Treg/TH17 imbalance?

Introduction Vitamin D is known to have profound effects on the immune system. We have shown that vitamin D is lower in patients with Acute Lung Injury (ALI) than in healthy or at risk controls and that in patients at risk of ALI post oesophagectomy, low vitamin D levels are associated with increased post-operative systemic inflammatory response and alveolar epithelial dysfunction. Studies have shown that when T cells are exposed to vitamin D, expression of IL17 decreases and regulatory capacity increases. We hypothesised that vitamin D deficiency may play a role in development of Acute Lung Injury (ALI) via changes in the balance between regulatory T cells (Treg) and pro-inflammatory Th17 cells. Methods Plasma levels of 25-OH Vitamin D (Tandem mass spectrometry) and 1,25-OH Vitamin D (ELISA)were measured in samples from patients with ALI. Normal T cells were exposed to BAL from patients with ALI with or without addition of exogenous vitamin D and determined frequencies of Treg and Th17 cells using flow cytometry. Results All samples tested had insufficient plasma levels of 25-OH vitamin D (<75 nmol/l, median 14.1 nmol/l). 1,25 vitamin D levels ranged from <20 to 176 pmol/l (reference range 43–144 pmol/l). 1,25 vitamin D levels were significantly related to both ITU survival (p=0.04) and survival at 90 days (p=0.04). Our initial findings suggest that BAL taken on day 0 of ARDS upregulated IL17 expression in normal T cells. This finding was blocked by exogenous 1,25-OH Vitamin D. By contrast, BAL taken on day 4 upregulated FoxP3 and CD25 expression, suggesting an increase in regulatory T cell activity. Discussion These results suggest that in early ARDS an imbalance in T cells favouring expression of IL-17 may play a role in the inflammatory response to injury, and this may be attenuated by adequate vitamin D levels. Later in the course of the disease, Treg cells may predominate and play a role in resolution.

S105 Does vitamin D deficiency increase risk of acute lung injury post oesophagectomy

Introduction Vitamin D has profound effects on the immune system and its deficiency has been implicated in increased risk of diseases such as tuberculosis and pneumonia. We have shown vitamin D levels to be lower in patients with Acute Lung Injury than in healthy or at risk controls. We hypothesised that vitamin D deficiency may be a risk factor for developing Acute Lung Injury (ALI) following transthoracic oesophagectomy. Methods 25-OH vitamin D (tandem mass spectrometry) and 1.25-OH vitamin D (ELISA) were measured in plasma samples taken from patients prior to oesophagectomy. IL-6, RAGE and HMGB-1 were measured by ELISA. Extravascular Lung Water (EVLW) measurements were recorded using a PiCCO catheter. Results All patients undergoing oesophagectomy had insufficient levels of 25-OH vitamin D (<75 nmol/l, median 25.5 nmol/l). 1.25-OH vitamin D levels ranged from 26 to 182 pmol/l (reference range 43–144 pmol/l). Patients who developed ALI more than 72 h post-op had lower levels of 25-Vitamin D (p=0.032). Very low levels of 25-OH vitamin D (<15 nmol/l) were significantly associated with elevated post-operative systemic inflammatory response (as demonstrated by higher plasma levels of IL-6 (p=0.006) and HMGB-1 (p=0.04)) with evidence of increased epithelial damage (elevated RAGE (p=0.03)). Levels of 25 vitamin D3<15 nmol/l were associated with greater post-operative increases in extra vascular lung water (p=0.03). Patients with severe vitamin d deficiency (<20 nmol/l) had a 40% risk of developing post-operative ALI compared to 15% in those with less severe deficiency (p=0.03). Discussion These results suggest that very low 25 vitamin D levels in oesophagectomy patients are associated with an elevated post-operative systemic inflammatory response, increased alveolar epithelial dysfunction and an increased risk of developing lung injury. These data support the rationale for clinical trials of vitamin D replacement as a preventative therapy for acute lung injury.