Hydroxysteroid Dehydrogenase activity and the vitamin D axis in acute lung injury - mechanistic and functional importance.

Patients who get infection may develop an exaggerated response that results in damage to organs including the lung. In the lung this is known as acute lung injury. Acute lung injury can occur due to a variety of insults including smoke inhalation, trauma as well as bacterial and viral infection. It causes the lu8ngs to fill up with water and this means that patients' breathing becomes very laboured. These patients therefore need care in the intensive care unit including support with their breathing (mechanical ventilation). The death rate associated with this happening is about 35-45%. Even those who survive acute lung injury have considerable recuperation periods and reduced quality of life 12 months afterwards.

In this application we present extensive novel research findings that identify two endocrine abnormalities in patients with ALI- namely defective 11-beta hydroxysteroid dehydrogenase type 1 (HSD-1) activity within the lungs and severe vitamin D deficiency. By using animal models of lung injury we have found that HSD-1 deficiency in genetically modified mice results in exaggerated and persistent inflammatory lung damage. Intriguingly, the HSD-1 deficient mice are very vitamin D deficient which may account for some of the exaggerated lung damage. The link between the activity of HSD-1 (which generates active steroid hormones within tissues such as the lung) and vitamin D is previously unrecognised.

The aims of this research are therefore to study why vitamin d levels are low in animals with the HSD-1 gene knocked out. We will establish if this vitamin D deficiency is functionally important by studying the levels of calcium and bone density in these animals over 6 months. We will ascertain through a series of experiments the mechanism of these mice becoming vitamin D deficient. In addition, we will test the ability of HSD-1 KO mice to resist infectious models of pneumonia and gut infection (peritonitis) as clinically relevant models. We will replace the vitamin D deficiency in the HSD-1 KO mice to establish if this can reduce lung injury. In order to translate our findings in mice we will establish whether there is a link between urinary measures of HSD-1 in a cohort of patients with or at risk of acute lung injury (expected low HSD-1 activity markers in urine).

This research should therefore expand our knowledge about the interactions between the local tissue production of active steroid cortisol and vitamin D. If our hypotheses are correct this research would provide a rationale for treating patients with acute lung injury with vitamin D or a therapy perhaps cell based to boost HSD-1 activity in inflamed tissues.

Objectives: We study why vitamin D levels are low in HSD-1 KO mice and its functional importance in terms of calcium metabolism, bone density and innate immunity. We test the ability of HSD-1 KO mice to resist infectious models of pneumonia and peritonitis following caecal ligation and puncture as clinically relevant models of infection/sepsis. We will replace the vitamin D deficiency in the HSD-1 KO mice to establish if this can both attentuate lung and systemic injury injury. We will establish whether there is a link between urinary measures of HSD-1 activity in a cohort with or at risk of ALI (expected low HSD-1 activity markers in urine).
Methodology: We have an existing colony of HSD-1 KO mice. We have experience of manipulating vitamin D levels in wild type mice by using cholecalciferol liquid (Vigantol) or making mice deficient using diet lacking vitamin D. We have expertise in the animal models of pneumonia (Mitchell) and caecal ligation and puncture (Thickett / Lax). We will utilise standard markers of lung injury and quantitative microbiology culture to assess the impact of these models. Sample and effect size calculations are based on robust data about from these models in wild type animals. For the observational clinical studies we will utilise cohorts of patients that are currently being recruited or archived samples, as well as recruiting ALI patients in a prospective manner.
Exploitation: data generated will be published in high citation peer reviewed journals. We will use the data about vitamin D replacement to plan further clinical trials in humans with ALI .

This research will benefit investigators examining the pathophysiology of acute lung injury, pneumonia and sepsis. Together these diseases represent a very significant healthcare burden and there has been little novel pharmacotherapy for these conditions for many years. In our animal experiments we will determine the mechanism and functional importance of HSD-1 KO induced vitamin D deficiency. Using clinically relevant models of sepsis related direct and remote lung injury we will determine to what extent abnormalities seen in HSD-1 KO mice are related to the vitamin D deficiency that they have. This research will also be of interest to researchers in other fields who are studying the role of local HSD-1 activity as a regulator of inflammation and fibrosis.

In the commercial sector, companies involved in the development of HSD-1 inhibitors may benefit from this research as it will potentially flag up an unwanted side effect of HSD-1 inhibition, namely vitamin D deficiency. It will be important for them to consider this research in their clinical trials design and to ensure patients taking HSD-1 inhibitors are monitored for the development of vitamin D deficiency.

Patients may also benefit from this work. Our intention with this application is to define the interaction between HSD-1 activity and vitamin D deficiency in patients with or at risk of acute lung injury. We will demonstrate the degree to which vitamin D deficiency contributes to lung inflammation in models of septic lung injury and whether treatment with vitamin D can reduce lung damage and inflammation. We will subsequently be in a position to develop clinical trials of vitamin D as therapy for acute lung injury or cell based therapies to deliver cells that over express HSD-1 to sites of inflammation to promote resolution of inflammation. It is not unrealistic to suggest that vitamin D therapy for lung injury could be evaluated clinically within 5 years of the end of this grant. Cell based therapy to promote local HSD-1 activity as a novel anti-inflammatory would likely take 10 years to deliver.
This research will also contribute to the reputation of the University of Birmingham as a centre for cutting edge translational research. The research will take place in the Centre for Translational Inflammation Research which was opened in June 2011. This is a purpose built £8 million facility located within the new £450 million Queen Elizabeth Hospital Birmingham. The centre has all the major core facilities (FACS, cell culture facilities and Q-PCR) necessary for this project and has laboratory and office accommodation for 100 clinical and biomedical researchers. It acts as a hub for researchers in critical care and respiratory medicine who have an extensive collaborative history. The success of the centre has attracted academic industry contacts with both big pharma (GSK, AZ) as well as SMEs (e.g. Creabilis, Biopta). Further MRC funding should therefore enhance the reputation of the unit and draw more collaborators

During the tenure of this grant, Dr Lax the proposed research fellow , will learn new techniques such as the mouse pneumonia model and GCMS measurements of urinary metabolites. Through her involvement in project design, analysis and writing papers for publication as well as presentation of her work on national and international conferences will enable her successful entry into the scientific community as an independent researcher. The development of these skills will be integral for her future career as an independent research leader.