Systemic bone loss and induction of coronary vessel disease in a rat model of chronic inflammation
Introduction
A significant body of literature has accumulated documenting the relationship between osteopenia or osteoporosis and chronic inflammation. Bone loss is a common complication in patients with HIV [1], [2], chronic periodontitis [3], inflammatory bowel disease [4], chronic pancreatitis [5], [6], as well as rheumatoid arthritis and lupus erythematosus [7], [8]. The prevalence of osteopenia has been reported to range from as high as 71% of HIV-infected patients [9] to 34% of adult patients with cystic fibrosis [10], while 31% of female patients with rheumatoid arthritis are considered osteoporotic [11]. Although therapeutic agents commonly used in the treatment of these conditions such as glucocorticoids [12], [13] or highly active antiretroviral therapy (HAART) [14], [15] can accelerate the rate of bone loss, osteopenia also develops independent of treatment and may be mediated via host responses to the underlying medical conditions [3].
A number of growth factors, cytokines, hormones, and adhesion molecules regulate normal bone cell differentiation, growth, and function [16]. Chronic elevation of proinflammatory mediators such as tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and prostaglandin E2 (PGE2) can disrupt normal bone remodeling and ultimately lead to bone loss [7], [17], [18], [19], [20]. In general, these mediators act both directly and indirectly to increase osteoclastogenesis, prevent osteoclast apoptosis [21], [22], and/or inhibit osteoblast activity [23]. The interrelationship between bone metabolism and the innate immune responses likely explains why medical conditions typified by chronic inflammation are associated with bone loss characterized by decreased bone mineral density (BMD) and deterioration of trabecular bone microarchitecture [1], [7], [8], [24], [25], [26], [27].
Due to the influence of inflammatory mediators on bone metabolism, in vivo models are needed for mechanistic studies of inflammation-induced bone loss, as well as for evaluation of potential therapeutic interventions. Some of the most commonly used animal models of inflammation and bone loss include the adjuvant [28] and collagen-induced arthritis models [29] and the rat colitis model [30]. The arthritis models experience significant localized and systemic osteopenia, but diminished mobility due to inflamed knee joints or paws may alter weight-bearing activity and confound findings related to bone [31], [32]. The rat colitis model, used in the study of inflammatory bowel disease [30], induces remarkable loss of trabecular bone, but interpretation of this findings may be complicated by gastrointestinal involvement altering nutrient absorption. Therefore, generalizing results from these models to chronic inflammation-induced bone loss may be problematic due to alterations in basic animal behavior or nutrient status.
Recently, Jarvelainen and colleagues [33] introduced a rat model of chronic endotoxemia in which subcutaneously implanted osmotic pumps released lipopolysaccharide (LPS; 0.1 mg/kg/day), the primary cell wall component of Gram-negative bacteria, over 30 days. Persistently elevated hepatic production of proinflammatory and anti-inflammatory cytokines was observed without the animals developing tolerance [33]. Even though the primary objective of the study [33] was not focused on the skeletal response to endotoxin, we hypothesized that comparable doses of LPS may be used to induce a chronic inflammatory state utilizing time-release pellets. We anticipated that time-release pellets would provide a means of delivering a controlled dose of LPS with minimal animal stress and may result in an in vivo model that could be maintained for a period of time sufficient to study skeletal alterations. Therefore, the purpose of this study was to determine whether administration of LPS via time-release pellets produced similar alterations in bone mass, bone microarchitecture, and biomechanical properties as those observed in chronic inflammatory conditions.
Section snippets
Materials and methods
We designed a dose response study to determine the effects of chronic LPS administration using time-release pellets on bone. Twenty-four 3-month-old male Sprague–Dawley rats (Harlan, Indianapolis, IN) were housed in an environmentally controlled animal care facility and allowed to acclimate for 5 days. Animals were randomly assigned to one of three groups (n = 8/group): Low dose LPS (3.3 μg LPS/day), High dose LPS (33.3 μg LPS/day), or Placebo (pellet containing matrix only). LPS (E. coli
Body weight and inflammatory indicator
Over the course of the 90-day study, no statistically significant differences in body weight were observed between the LPS-treated and placebo groups (Fig. 1). A trend, however, toward reduced body weight (P = 0.0578) was evident in the group receiving the High dose LPS at the final time point. No noticeable alterations in animal behavior were observed in terms of grooming, food consumption, and physical activity with the low doses of endotoxin utilized in this study. Neutrophil counts were
Discussion
Although LPS has been one of the most commonly used agents to investigate the relationship between innate immune responses and bone metabolism, few studies have focused on the chronic LPS response [18], [36], [37], [38]. Animal models of periodontitis involving repeated LPS injections into the gingiva over the course of 7–12 days have been used to determine localized effects on alveolar bone [36], [37], [38]. Additionally, Miyaura et al. [18] evaluated the systemic response of young growing
Conclusion
These findings provide evidence that chronic LPS delivered via time-release pellets induces alterations in bone and coronary vessels. Bone loss was demonstrated by decreases in bone mineral content, density, and trabecular bone microarchitectural properties. The presence of fibrous areas around the arterioles and disruption of the intima border suggest that early changes consistent with arteriosclerosis may be occurring. Furthermore, COX-2, TNF-α, and IL-1β continued to be elevated in the
Acknowledgments
This work was supported in part by the Department of Veterans Affairs Administration and Department of Surgery and Department of Medicine Research Funds. We also wish to express our appreciation for the support of the OUHSC NIH General Clinical Research Center in the collection of this preclinical data.
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