Interleukin 10 knockout frail mice develop cardiac and vascular dysfunction with increased age☆
Highlights
► IL-10 knockout frail mouse model develops vascular stiffness and hypertension. ► These mice get vascular endothelial dysfunction as a result of COX dependent vasoconstriction. ► IL-10 knockout mice have cardiac hypertrophy, remodeling and decreased function. ► Our findings are consistent with the IL-10 knockout as a good model for human frailty and cardiovascular aging.
Introduction
Aging is inevitable, yet its physiologic consequences are, to some degree, modifiable. Cardiovascular (CV) dysfunction is the final common pathway of many acquired disease states and hence the most common cause of age-related deaths in the United States (Godwin, 2005, Heron, 2011). Frailty is a geriatric syndrome of late-life vulnerability to adverse outcomes and early mortality associated with declines in multiple physiological systems, the activation of inflammatory pathways, skeletal muscle decline, and subclinical cardiovascular disease(Dato et al., 2012, Espinoza and Walston, 2005, Ko et al., 2012). Given that frailty is such an important marker of adverse outcomes, the identification of etiological pathways that influence frailty-related vulnerability will greatly facilitate the development of improved risk assessment and better preventive and treatment modalities.
Interleukin (IL) 10 was originally demonstrated to be an anti inflammatory product of T-helper 2 cells (Fiorentino et al., 1989). Genetic deletion of IL-10 in mice (Kuhn et al., 1993) leads to a series of IL-10 associated pathologies. An increased risk of developing enterocolitis and colorectal cancer (Berg et al., 1996), inflammatory bowel disease (Das et al., 2003), development of osteopenia, decreased bone formation, mechanical fragility of long bones (Dresner-Pollak et al., 2004), and exacerbation of fatigue and motor deficits (Krzyszton et al., 2008) have been demonstrated in IL-10 deficient mice. The IL-10 mouse has been proposed as a mouse model of frailty, as aging IL-10(tm/tm) mice develop increased inflammatory pathway activation, decreasing skeletal muscle strength, and declining activity as well as early mortality. This phenotype is consistent with frail older humans. (Ko et al., 2012, Walston et al., 2008).
Further studies have shown that IL-10 inhibits LDL/Ox-LDL dependent monocyte-endothelial interaction thereby inhibiting atherogenesis and hence preventing the development of atherosclerotic plaque in mice (Caligiuri et al., 2003, Mallat et al., 1999, Pinderski Oslund et al., 1999). Furthermore, plasma IL-10 levels have been shown to decrease in patients following myocardial infarction (Heeschen et al., 2003). Additionally, data demonstrate that plasma IL-10 levels are directly correlated with good prognosis and remain an independent predictor of long-term adverse cardiovascular outcomes in Acute Coronary Syndromes (Cavusoglu et al., 2011). IL-10 levels also have a strong inverse correlation with stroke mortality, as shown in the Leiden 85-Plus study (van Exel et al., 2002).
It is well established that the endothelium is critical in mediating vasorelaxation to agonists such as acetylcholine (ACH) through nitric oxide (NO) and Endothelial Derived Hyperpolarizing Factors such as hydrogen sulfide (Mustafa et al., 2011). Equally important are the Endothelium Derived Contractile Factors (EDCF). Indeed, arachidonic acid derivatives produced by endothelial COX mediate constriction or relaxation in different vascular beds (Moncada and Vane, 1978). Recent studies have reinforced the idea of endothelial and COX dependent vasoconstriction, induced by mechanical or chemical stimuli. These cholinergic or stretch-mediated stimuli lead to increased intracellular calcium concentration (Miller and Vanhoutte, 1985, Katusic et al., 1987, Katusic et al., 1988, Ihara et al., 1999, Okon et al., 2002, Yang et al., 2004). It is also known that senescence increases expression of physiologic and inflammatory isotypes of COX protein, COX-1 and inducible cyclo-oxygenase (COX-2) respectively (Heymes et al., 2000, Matz et al., 2000, Stewart et al., 2000), endothelial COX-2 mRNA (Voghel et al., 2007), and mRNA and protein expression of inducible NOS (iNOS) (Tabernero et al., 2000, Chou et al., 1998). IL-10 is known to impair production of inflammatory TNF and iNOS produced by liver CD11b1/Ly6C1 cells (Bosschaerts et al., 2011). Interestingly, iNOS binds to and S-nitrosylates COX-2, leading to activation and increased catalytic activity (Kim et al., 2005).
Given the support for the use of this mouse as a model of human frailty, and the knowledge that IL-10 as well as inflammatory mediators profoundly effect cardiovascular function, we hypothesized that the IL-10(tm/tm) mice would develop an age-related change in cardiovascular phenotype, develop endothelial dysfunction and vascular stiffness consistent with that reported in frail older adults. We explored the role of IL-10 in vasoregulation and maintenance of cardiac function in this model of aging, frailty and inflammation.
Section snippets
Animals
Age and background matched, IL-10 deficient (IL-10(tm/tm)); B6.129P2-Il10tm1Cgn/J and control mice (C57BL6; WT) were obtained from Jackson Laboratories (Bar Harbor, ME, USA). IL-10(tm/tm) mice used are homozygous for the Il10tm1Cgn targeted mutation. These mice were housed in Association for Assessment and Accreditation of Laboratory Animal Care International accredited facilities and pathogen contact prevention (prophylaxis from infections, inflammatory bowel disease and early mortality) was
Body mass
There was no significant difference in the body mass in age matched IL-10(tm/tm) and WT mice. Young IL-10(tm/tm) vs. WT mice average weight was measured to be 27 g vs. 31 g and in old IL-10(tm/tm) vs. WT mice group the average weights were 38 g vs. 36 g (Fig. 2E).
Vascular studies
In ex vivo myograph experiments, measured tension represents a balance between vasorelaxant and vasoconstrictor dependent function and mediators. In phenylephrine pre-constricted isolated mouse aorta, ACH stimulates the release of
Discussion
The role of inflammatory pathway activation and elevation of serum inflammatory cytokines in age-related disease states, frailty, and functional decline is an active area of investigation (Singh and Newman, 2011, Chen and Frangogiannis, 2010). Chronic activation of NF-kB induced inflammatory cascades, such as that induced via deletion of IL-10, influences the frailty phenotype and the associated vulnerability to multi-systemic decline in these mice, similar to that observed in frail older human
Conclusion
We conclude that the older IL-10(tm/tm) mouse model of frailty has stiffer vessels and a loss of endothelial vascular relaxation, possibly due to an increase in eicosanoids, especially due to the increase in activity of COX 2 and resultant thromboxane A2 receptor activation, but not due to the increase in prostacyclin. Our results also suggest that this model of frailty has increased heart size but also a worsening of cardiac function. Finally, further studies will be required to determine
Acknowledgments
We thank Dr. Solomon H. Snyder and Paul Scherer for their continuous collaborative efforts in helping with experiments and technology sharing.
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Grant support: NHLBI R01 grant (HL105296-02) to D.E.B., Claude D. Pepper Older Americans Independence Center (P30 AG021334) to J.D.W, AHA postdoctoral fellowship grant (10POST4010028) to G.S., American Diabetes Association Research Grant and American Heart Association Beginning Grant in Aid to L.A.B.