NIH Program Project Grant (P01 DK 43785): Ischemia-Reperfusion Injury
Founded: 1991 with 5-year grant for $4.7 million
Renewed: 1996 with 5-year grant ($6.4 million); renewed 2003 with 5 year grant ($8.0 million)
Full Time: 23
Part Time: 0
Budget: $19.1 million

Director: Dr. Neil Granger, Ph.D
Boyd Professor & Head, Department of Molecular and Cellular Physiology
LSU Health Sciences Center
1501 Kings Highway, P.O. Box 33932
Shreveport, LA 71130

Telephone: 318/675-6011 FAX: 318/675-6005 E-Mail: dgrang@lsuhsc.edu

Principal Investigators (Projects): D. Neil Granger, Ph.D.; Ronald Korthuis, Ph.D.; Steven Alexander, Ph.D.; Tak Yee Aw, Ph.D.; Matthew Grisham, Ph.D.

Principal Investigators (Cores): D. Neil Granger, Ph.D.;Tak Yee Aw, Ph.D.; Lynn Harrison, Ph.D.; Hugh V. Price, Ph.D.

Research Areas:

The inflammatory process is essential for survival, playing an important role in both health and disease. In recent years, there has been an explosion of knowledge concerning the immune system and its involvement in the inflammatory process. The identity, biological actions, and underlying molecular mechanisms of several classes of inflammatory mediators have been defined, and the contributions of different circulating, vascular and extravascular cell populations to the inflammatory response are now well appreciated. A major conceptual outgrowth of the intensive effort to define the inflammatory process is the realization that vascular endothelial cells play a pivotal role in the coordination and regulation of immune responses that are elicited in inflamed tissue. This critical role for the endothelial cell reflects both its ability to respond to inflammatory mediators (eg, cytokines, oxidants) as well as its ability, when activated by these mediators, to orchestrate the recruitment of circulating leukocytes into inflamed tissue by creating a favorable environment for their trafficking, capture, and extravasation. Other consequences of the endothelial cell activation that accompanies an inflammatory response include an impaired ability of arterioles to regulate blood flow, increased fluid filtration across capillaries, and enhanced plasma protein extravasation in postcapillary venules. Collectively, these microvascular responses sustain the leukocyte infiltration, hyperemia and interstitial edema that are considered to be the cardinal features of inflammation.

Inflammatory processes have been implicated in a variety of human diseases, including the more traditionally defined chronic inflammatory diseases, such as arthritis and inflammatory bowel disease, as well as systemic cardiovascular diseases (atherosclerosis) and disorders (circulatory shock) for which the involvement of an inflammatory response has only recently been recognized. The diversity of the list of human diseases that involve inflammation is matched by an equally diverse list of models that are used to mimic the inflammatory process in experimental animals. Chemical irritants, immune activators, cytokines, and other exogenous agents have been widely used to elicit acute and/or chronic inflammatory responses in animal models. Other models have exploited the intrinsic ability of tissues to generate inflammatory mediators following a period of oxidative, ischemic or hypoxic stress, thereby negating the need to expose tissues to exogenous agents.

An experimental insult that has been widely employed to elicit and study the mechanisms underlying the microvascular and tissue responses to inflammation is ischemia and reperfusion (I/R). While tissue hypoxia and acidosis are generally regarded as the major factors that mediate the pathological alterations associated with ischemia per se, an inflammatory process has been invoked to explain the vascular dysfunction and tissue injury produced by I/R. The intensity of this inflammatory reaction in some postischemic tissues (eg, intestine) can be so severe that the reperfusion-related response is also manifested in distant organs. The remote effects of I/R are most frequently observed in the lung, liver, and cardiovascular system, and can result in the development of the systemic inflammatory response syndrome (SIRS) and multiple organ dysfunction syndrome (MODS). Although I/R induced inflammatory responses and the accompanying microvascular dysfunction have been described in most organ systems, the splanchnic circulation appears to be particularly vulnerable to the deleterious effects of such an insult. Hence, it is not surprising that the literature is replete with reports that implicate I/R in the pathogenesis of a variety of digestive diseases (eg, duodenal ulcers, inflammatory bowel disease) and conditions (liver transplantation) that are characterized by profound changes in tissue perfusion and/or an intense inflammatory response. While it remains unclear whether splanchnic I/R injury is amenable to therapeutic intervention, this physiologically relevant stimulus continues to offer unique insights into the intrinsic ability of tissues to initiate and sustain an inflammatory response. Hence, a better understanding of the processes involved in I/R injury may therefore ultimately provide us with information about how to protect tissues against the deleterious consequences of ischemic episodes as well as acute inflammatory events not associated with an ischemic insult. The extensive experimental characterization of the contribution of different adhesion molecules to leukocyte recruitment as well as the role of reactive oxygen metabolites in mediating the increased vascular permeability in intestinal I/R are two examples of the novel insights that have emerged from this model of inflammation. Over the past 10 years, the investigators participating in this Program Project Grant (PPG) have contributed significantly to the existing body of knowledge in the area of I/R injury and have taken a leadership role in proposing and testing novel concepts that relate to the molecular mechanisms that underlie the inflammatory and microvascular responses elicited by I/R.

Reactive metabolites of both oxygen (eg, superoxide) and nitrogen (eg, nitric oxide) have been implicated in the pathogenesis of most forms of inflammation. These reactive nitrogen oxide species (RNOS) appear to contribute to the inflammatory process by: 1) serving as signalling molecules, 2) activating nuclear transcription factors for both pro- and anti-inflammatory proteins, and 3) mediating cell necrosis and apoptosis. While nitric oxide (NO) and superoxide per se are often ascribed anti- and pro-inflammatory roles, respectively, the products of their chemical interaction (RNOS) can yield either phenotype (anti- or pro-inflammatory), depending on whether there is net oxidation or nitrosation of specific molecular targets that regulate the inflammatory response. These observations have led to the concept that a critical determinant of the inflammatory (pro- or anti-) phenotype assumed by tissues is the existing balance between reactive oxygen species (ROS) and NO. In no inflammatory state has this NO/ROS balance concept received more attention than in I/R, wherein an imbalance between NO and ROS is believed to produce the pro-inflammatory phenotype that is characteristic of postischemic tissues. Despite the existing body of evidence that supports a role for NO and ROS in the pathogenesis of I/R injury, no effort has been made to systematically assess the relative importance of ROS and NO in I/R-induced inflammatory responses and to determine the biochemical and molecular basis for this NO/ROS-mediated response.

The overall objectives of this PPG are: 1) to define the biochemical and cellular responses that ultimately lead to the organ dysfunction that occurs following reperfusion of ischemic tissues, and 2) to elucidate the molecular mechanisms underlying these responses. This PPG includes a spectrum of projects that will focus on the critical role of endothelial cells and inflammatory cells in the pathogenesis of I/R injury in intestine and liver. A multidisciplinary approach will be used to elucidate the biochemical, molecular, structural, and physiological responses of the microvasculature to ischemia and reperfusion. The work outlined in this PPG is a natural outgrowth of the research interests and ongoing collaborative efforts of eleven investigators with active programs in the areas of endothelial cell biology, nitric oxide/oxygen radical biochemistry, inflammation, microcirculation, and gastroenteric biology. The 5 highly focused and strongly inter-related projects in this Program rely on 4 core units (1 administrative & 3 scientific) which provide state-of-the art technical support that will help to ensure that the scientific objectives of each project are met.

Areas of Expertise:

This Program formalized and extended existing collaborative research efforts among 11 highly productive scientists at LSUHSC-S with active, well-funded programs in the areas of endothelial cell biology, inflammation, microcirculation and gastroenteric biology. The funding of this PPG allows investigators with diverse backgrounds and expertise to focus on multiple aspects of a single problem, the pathophysiology of intestinal I/R, thereby facilitating the formulation and testing of new hypotheses relative to this question. This coordinated effort involves a multidisciplinary approach aims at elucidating the mechanisms responsible for intestinal ischemia/reperfusion injury at the molecular, cellular, single microvessel and organ levels. The ultimate goal of the PPG is to obtain the fundamental knowledge that is needed to diagnose and treat ischemic disorders of the bowel.

Special Capabilities and Facilities:

Of the over 400 PPGs funded by the National Institutes of Health, this is the only Program which focuses on the mechanisms of ischemia/reperfusion injury. Another unique aspect of this Program relates to the multidisciplinary, interdepartmental approach that brings to bear the scientific expertise of a group of investigators with diverse backgrounds, research techniques and experience to focus on a single problem. The Program also supports 4 core facilities including the Administrative, Cell Culture and Imaging, Biochemistry and Molecular Biology and an Induced Mutant Mouse Core that function to provide support for, as well as to integrate and standardize the activities of the Program.

Research Equipment:

A wide variety of state-of-the-art equipment is available for studying the cellular and molecular mechanisms of ischemic disorders. Of particular note are the Confocal Imaging Microscope and Ratio Fluorescence Spectrometer facilities.

Keywords:

Ischemia/reperfusion
Endothelial cells
Cardiovascular Disease
Cell culture
Molecular biology