Looking Beneath the Surface—Setting the Stage for Skin Ulceration

Peripheral vascular disease (PVD) produces hypoxic conditions in the lower limb that can lead to ulceration and amputation. Studies of the cutaneous sequelae associated with this problem are much needed since demographic data predict a rising number of patients with PVD due to an increase in the geriatric population. In addition, the obesity epidemic in the US is evoking a rapid escalation in the number of cases of type II diabetes in an increasingly younger patient population. Such patient groups will predictably develop cardiovascular perturbations leading to hypoxic conditions in the leg and foot region. Breakthroughs in treatment modalities are sorely needed to combat the impact of ulcers and amputations on quality-of-life issues ranging from financial, social, and psychological implications to matters of morbidity and mortality. In this issue, Dalton and coworkers have begun to forge a pathway toward a greater molecular understanding of the events that lead to skin breakdown when the lower extremity is afflicted with severe peripheral vascular disease. Their team recognized that documentation of events/conditions in ischemic lower extremities is a requisite step that must precede the design of effective therapeutics to combat non-healing wounds in this patient population. Their approach is unique in that they chose not to analyze the wound bed after the fact. Instead, they analyzed unmistakably hypoxic skin prior to its ulceration and development into a chronic wound. Their report provides evidence of abnormal extracellular matrix metabolism in chronically ischemic skin. They hypothesize that these abnormalities may be causal factors leading to skin breakdown. Traditionally, evidence-based medicine in the chronic wound field has been hampered by the apparent unavailability of tissue samples. Institutional Review Boards have been loath to approve excisional biopsies in patients who are at highest risk of developing non-healing wounds that could easily progress to amputation. Fortunately, Dalton and associates remained undaunted by problems of sample collection and patient recruitment. Instead, they devised a clever plan to retrieve intact hypoxic skin tissues from amputated limbs that were previously relegated to the scrap heap in the biohazard waste. Their report focuses on both the synthesis and degradation aspects of the equilibrium in the extracellular matrix (ECM) in chronically ischemic leg tissues. The result is a foundational analysis of major ECM dynamics in the skin complete with site-matched controls compared against normal samples from patients having knee replacements or elective removal of superficial veins. Their data derived from chronically hypoxic yet unwounded skin augment information in the field of wound repair. PVD produces an obvious deficiency in oxygen and other nutrients necessary for tissues to heal and thrive. The role of oxygen in wound healing has been deemed to be important for years (Hunt, 2004). Recent articles on this molecule of interest have suggested that oxygen is more than just a nutrient (Tandara and Mustoe, 2004). Although the wide range of oxygen tensions found in tissues makes it difficult to establish a standardized numerical value to define hypoxia, scientists are in agreement that hypoxia is defined as a condition when oxygen delivery does not meet the metabolic demands of the tissue. As cells sense hypoxia, they mount adaptive cellular reactions that lead to multiple transcriptional as well as post-transcriptional events (Papandreou et al, 2005). Within the ECM, oxygen is essential at an enzymatic level since prolyl hydroxylase is required for the hydroxylation of proline, a requisite step if procollagen molecules are to form stable triple-helical conformation in the ECM (Tandara and Mustoe, 2004). Since hypoxic, normoxic, and hyperoxic conditions are easily reproduced in the culture dish, much of the current hypoxic-based literature is derived from such a setting. Chronic hypoxic conditions in vitro are well known to decrease the proliferative activities of fibroblasts and to decrease transforming growth factor-β (TGF-β)1 mRNA and its TGF-β receptor values, and the converse is true under normoxic or hyperbaric conditions (Mogford et al, 2002, 2004). Newer molecules such as hypoxia-inducible factor 1 (HIF-1) have also been implicated in acute and chronic hypoxic settings such as those surveyed by Dalton and coworkers. HIF-1 is one such nuclear factor that seems to serve as a prime regulator of gene expression when the microenvironment becomes hypoxic (Semenza, 1999). Oxygen-dependent regulation of gene expression in culture and in wounds has also been reported for inducible nitric oxide synthase, arginase I, vascular endothelial cell growth factor (VEGF), tumor necrosis factor-α (TNF-α), TGF-β, interleukin (IL)-6, IL-8, cyclooxygenase (COX)-2, among others (Albina and Reichner, 2003). Although the acute effects of hypoxia on cells is known to result in apoptosis or cellular death, the pathophysiologic impact of a severe oxygen deficit over a chronic time interval has remained unknown. Practically, every one of these hypoxic experiments has been performed within the wound-healing milieu; yet, none was performed in chronically ischemic tissues BEFORE development into a non-healing wound. Thus, the natural history of hypoxic events in a limb afflicted with PVD has been largely ignored until now. From time to time, promising new therapeutic approaches for treatment of chronically ischemic skin have appeared on the horizon. A recent example was the therapeutic angiogenesis approach that sought to improve perfusion by the growth of new blood vessels. Unfortunately, the phase II randomized, double-blind study with intramuscular adenoviral delivery of VEGF in patients with disabling claudication did not fulfil its expectations and did not show sustained efficacy in improving even intramuscular blood flow, much less cutaneous blood flow in the affected limb (Rajagopalan et al, 2003). A subsequent study using an ischemic hindlimb model performed a careful immunohistological analysis and provided a possible explanation by showing that angiogenesis was only transient in nature (Gounis et al, 2005). Nevertheless, optimism about future developments remains high since other creative avenues remain open, and there remain a host of candidate molecules that may be deficient or elevated in the chronically ischemic skin setting. Work with tumor cells and their blood supply has suggested that hypoxic tissues respond by halting the cell cycle, thereby blocking the addition of new cells into the tissue and/or by the induction of programmed cell death or apoptosis (Papandreou et al, 2005). Whether such mechanisms factor in the hypoxic circumstances within chronically ischemic skin tissues in vivo was not directly addressed by Dalton and colleagues in this issue but should definitely be evaluated in future wound-healing studies. Cellular responses that evolve in vivo over a chronic period are obviously very complicated and multifactorial; however, tissue collection as was performed in the Dalton study in this issue provides an excellent sampling tool to open the window to our understanding of the perturbations that lead to skin breakdown. In summary, Dalton et al have done a fine job of providing the scientific community with a valuable characterization of several major molecules that govern the state of the ECM in skin exposed to chronic hypoxia. Although the “tissue vulture strategy” in the wake of limb amputation is certainly without glamor and glitz, this group has taken the scraps from the biohazard waste and put forth information that points toward the pathway of understanding that may one day lead to advancements that make a difference. Events that precede the development of skin ulceration are far from settled.