Regulation of inflammation during tissue repair and regeneration

Tissue repair and regeneration require dramatic and coordinated changes in cell behaviour in both wound-resident cells at the site of injury and in distant cells that respond to and are recruited to the injured tissue. In the last decade, the influence of inflammatory cells on wound healing has been shown to be highly significant, as they can function to promote or inhibit wound healing. Discovering the underlying mechanisms controlling the behaviour of inflammatory cells is key to controlling these cells for therapeutic benefit. It is often very instructive to compare normal processes with diseased processes in order to understand how that process is regulated.
Diabetic patients and animal models have severely impaired wound healing and often develop chronic wounds. By comparing factors in diabetic wounds with normal wounds, we can begin to understand what is important for efficient wound healing and how to promote impaired wound healing. Inflammatory cells from diabetic patients and animal models are dysfunctional and inhibit wound healing. However, this process is poorly understood and the key mediators that control these cells are not known. Many pro-inflammatory factors are over-expressed in diabetic chronic wounds compared to normal wounds, but whether they are causative or a consequence of the dysfunctional inflammatory cells is not known.
Diabetes causes changes to cells that seem to be permanent. This is due to complex marks on DNA and proteins called histones that associate with DNA. They control which genes are on or off. Researchers are now learning that these marks may be reversible, but not much is known about them. However, inflammation may be significantly influenced by these marks.
We have found a factor called Hoxa3 that can suppress inflammation during wound healing. Preliminary data suggests that Hoxa3 controls inflammation by regulating the enzymes that control marking the DNA and proteins, as well as a well-known critical regulator of inflammation called NF-kB. We are proposing to carry out a work programme to test this hypothesis in inflammatory cells from healthy and diabetic patients and mice. Our objectives are to (1) determine if there are differences in the marks on the histones, as well as the enzymes that confer those marks, between diabetic and healthy white blood cells, (2) compare NF-kB activation in white blood cells isolated from diabetic and healthy controls, and (3) elucidate the mechanisms by which Hoxa3 suppresses inflammation with regards to the histone marks (via control of the enzymes that confer those marks), NF-kB activation (suppressing the pro-inflammatory function of this factor), and whether these functions of Hoxa3 can bring about pro-wound healing behaviours in inflammatory cells. The results of this study will be important in future therapeutic development.

Dysregulated inflammation leading to wound retention of inflammatory cells is a major contributing factor to chronic wounds, the leading cause of non-traumatic amputations in the developed world today. In acute wounds, inflammation resolves naturally, following a switch from pro-inflammatory to anti-inflammatory gene expression in myeloid cells, although the precise regulatory mechanisms controlling this switch are unknown. We have identified a factor, Hoxa3, which is normally upregulated in wounds during this switch, but is repressed in diabetic wounds. Our recent published studies have shown enforced expression of Hoxa3 in diabetic wounds represses genes such as Traf and Irak that are required for TNF-mediated NF-kB activation. In addition, Hoxa3 upregulates Kdm3b, a histone demethylase that suppresses myeloid cell differentiation/activation. These changes are associated with a 50% decrease in the wound retention of myeloid cells and significantly improve healing. Our most recent data demonstrates that Hoxa3 protein transduction of myeloid cells can induce similar changes. Altogether, our work points to novel functions of Hox protein control of myeloid cell development and inflammation that must be pursued.
The work proposed here aims to link the deregulated inflammatory response in diabetic wounds to chromatin misregulation and aberrant NF-kB modulation. Our central hypothesis is that hyperactivation of pro-inflammatory genes is due to changes in chromatin accessibility and NF-kB hyperactivation, but that these changes can be reversed by Hoxa3 gain of function via protein transduction. The work proposed here will shed light on the underlying causes of chronic inflammation in diabetic wounds and provide insight into Hox protein function in the haematopoietic system that will inform development of urgently needed therapies for chronic wounds.

Beneficiaries:
Over 220 million people suffer from diabetes worldwide, a condition that often leads to the development of chronic wounds, and ultimately limb amputation. Currently it is estimated that in the UK alone there are over 100 amputations per week due to chronic wounds. Current treatments are ineffective due to a poor understanding of the underlying mechanisms causing chronic wounds, leading to a significant burden on the health care system. High morbidity and mortality in chronic wound sufferers causes significant stress on their families and carers. We believe our research will ultimately benefit all members of the public, as it will reduce the cost burden to the NHS of a significant number of patients with chronic wounds. Development of cost-effective, efficacious treatments that significantly promote wound healing would reduce hospitalization time and stress of the many diabetic and elderly patients who suffer from chronic wounds, as well as their families and care-givers. This study will provide much-needed data that will contribute to development of novel therapies for chronic wounds. The lead-time for clinical translation of this work is approximately 5 years.

Communications and engagement:
Given the potential for clinical application, we will disseminate our findings as widely as possible in order to maximize the impact of our research. We will communicate our findings to the scientific community and the public through online and print media, such as websites and the Faculty Research brochure, national and international scientific conferences, and open access, peer-reviewed publications. The Faculty of Life Sciences is highly supportive of innovative and open dissemination of research, and provides funds to help meet costs of doing this. We will also communicate our research to lay audiences through public engagement events, such as science days for the public at the University and with The Healing Foundation Charity, who actively engage in disseminating our research findings throughout the UK. We will also continue to support the Healing Foundation fundraising efforts through their organized public engagement events.

Education:
Our lab has hosted A-level students from the local Manchester area to provide them with work experience and encourage their pursuing science degrees at university, and will continue to do so each summer. In addition Dr Mace and Dr Torbica participate regularly in the Manchester Museum's A-level Stem Cell Day, aimed at educating lower and upper 6th Form students about embryonic and adult stem cell biology as well as the 'hope and hype' of stem cell therapies. Our goal is to foster a desire to participate in scientific research and to promote a healthy society.

Exploitation and Application:
We envisage the results of this study providing us with significant data from which to design a clinical trial. Intellectual property rights will be pursued where applicable via the university's IP team, but it is also possible that existing drugs, e.g., small molecule inhibitors approved for use in other clinical applications, may prove to be appropriate. Our goal is to develop effective therapeutics as soon as possible.

Training:
The postdoctoral research associate and technical staff associated with this project will develop their knowledge regarding an important area of health and welfare in addition to technical skills required for the work. They will contribute to a highly interactive environment within the Healing Foundation Centre and in the wider Faculty of Life Sciences, developing transferable skills, such as collaboration and presentation effectiveness, which will aid them in pursuing a life-long career in science.