Alterations in coronary microvessel reactivity after pulmonary nanoparticle exposure have not been previously identified. When the reactivity of these resistance vessels become dysfunctional, increased oxygen demands can not be met and overall cardiac metabolic activity is limited and/or decreased. Such alterations in cardiac physiology are consistent with ischemic events (ACSM 2005
). Events such as these were found to be increased after PM exposure (Goldberg et al. 2000
; Schwartz and Morris 1995
). Because coronary dysfunction in healthy people is unlikely to lead to observation or realization of clinical symptoms, it is more likely that PM-induced dysfunction contributes to ongoing disease or exacerbates events by diminishing collateral reserve. The present study showed that coronary arterioles from rats exposed to nano-TiO2
demonstrated reduced vasoreactivity to shear stress, ACh, and A23187, while smooth muscle responsiveness to NO remained unaltered. All of these changes are consistent with endothelial dysfunction. However, it is important to indicate that each of these stimuli operates through distinctly different cellular mechanisms. Because this is the first study to examine the relationship between nanoparticle exposure and coronary microvessel reactivity; a thorough investigation of these mechanisms need to be performed in subsequent studies.
While certain inhaled particles may be capable of exiting the lung in a limited context, there is currently not a large body of evidence that supports the translocation of particles en masse into the systemic circulation. Further, the likelihood of direct interaction of particles and endothelial cells is not high due to a relatively low pulmonary deposition (10 μg) in relation to the overall size of the rats used in the study (approximately 300 g). Despite this disproportionate relationship, the current study did not identify if nanoparticles are on or in the endothelial cells of the preparation. As such, it is not possible to distinguish between direct effects of nanoparticles and more global systemic response. However, if this possibility is entertained in the hypothetical context of an immediate, 100% particle migration; dilution in the systemic circulation would be marked, and subsequent tissue-particle interaction would be further opposed by the vast endothelial surface area of the collective vascular endothelium, thus rendering a direct effect between migrated particles and the systemic endothelium unlikely. Because this is an in vitro preparation, the influence of nerves on microvascular function has most likely been removed. While it is not possible to discount the possibility that the nerves have primed cells in the microvascular wall and altered arteriolar function that persists in vitro, there is currently no evidence to support this possibility after particle exposure. Therefore, it appears an inflammatory effect is likely mediating the resultant dysfunction in coronary arterioles that follows pulmonary nanoparticle exposure.
Increases in intraluminal flow produce profound changes in the caliber of coronary resistance arterioles via endothelial transduction of shear stress (Kuo et al. 1990
); thus, flow-induced dilation is critically important in regulating coronary vascular resistance. This is the first study to show that under similar levels of longitudinal shear stress (), coronary arterioles from nanoparticle-exposed animals exhibited impaired vasodilation as compared to those from sham-control rats (). Coronary microvessels of similar size to those used in the present study were found to actively respond to a range of shear stress (0-10 dyn/cm2
) (Kuo et al. 1995
), indicating that the flow rates correspond to physiologically significant levels of shear stress. In studies with humans, it was shown that air pollution exposure diminished brachial blood flow in response to endothelium-dependent agonists (O'Neill et al. 2005
; Tornqvist et al. 2007
). Further, endothelial cell dysfunction was suspected to be the primary cause of reduced dilation to flow (as defined by ischemic responses) after exposure to urban air pollution in humans (Briet et al. 2007
). Similarly, occupational exposure to PM carries the same risk as environmental exposure, but perhaps at higher deposition rates. Since occlusion-based studies in conduit vessels correlate poorly with myocardial microvascular perfusion (Bottcher et al. 2001
), it is difficult to extrapolate findings with conduits in the arm to direct coronary microvascular effects. Flow-mediated arteriolar dilation is heavily dependent upon endothelial cell integrity in the coronary microcirculation (Kuo et al. 1990
); therefore, the current results support the link between nanoparticle exposure and endothelial cell dysfunction via direct experiments with the target tissue in question.
In the present study, endothelium-dependent dilations to ACh and A23187 were decreased in coronary arterioles from nano-TiO2
exposed rats. This is in accordance with previous reports from our lab which showed an impaired dilation to A23187 in the spinotrapezius microcirculation after exposure to fine combustion particles (Nurkiewicz et al. 2006
) or nano-TiO2
(Nurkiewicz et al. 2008
). This is also consistent with other reports that indicate endothelium-dependent vasoreactivity is impaired after exposure to PM. Cozzi et al. (2006) showed a reduction in ACh-induced vasodilation in aortas after intratracheal (IT) instillation with UF PM. Similarly, IT instillation of larger particles (PM10
) produced an impairment in ACh-induced dilation in rabbit carotid arteries (Tamagawa et al. 2008
). Other pollutants such as diesel exhaust or its related gaseous components exert comparable deleterious effects as evidenced by decreased ACh-induced vasodilation (Hansen et al. 2007
) or enhanced vasoconstriction (Campen et al. 2005
). However, this is the first investigation to show that exposure to relatively non-inflammatory levels of a nanoparticle (TiO2
) negatively impacted endothelium-dependent vasodilation in the coronary microcirculation. Because overt pulmonary inflammation was not present at the 10 μg deposition dose used herein or with higher doses used in other studies (Nurkiewicz et al. 2006
; Sager et al. 2008
), yet considerable coronary dysfunction was, it was postulated that inflammatory mechanisms reside in the microvascular wall and continue to exert their effects in vitro
. Therefore, future studies need to characterize this inflammation in the microvascular wall and assess its intensity relative to the pulmonary exposure. Given the association between the function of this critical level of circulation and cardiac function, the potential link between pulmonary exposure and ischemic events is apparent.
Under conditions of reduced perfusion pressure in the heart, autoregulatory adjustments through vasodilation contribute to maintenance of blood flow to the myocardium. Autoregulatory adjustments are crucial in helping to prevent edema and microvascular damage by preserving capillary hydrostatic pressure when coronary perfusion pressure swells by shielding capillaries from excessive systemic pressures during diastole. Besides vasodilation to metabolic stimuli, another autoregulatory mechanism that participates in controlling coronary blood flow is the myogenic response, or the rapid and maintained constriction of a blood vessel in response to pressure elevation. Consistent with a previous study in mesenteric arteries (Knuckles et al. 2008
), the present results showed no differences in myogenic responsiveness after nanoparticle exposure. Since a functional endothelium was reported to influence arteriolar myogenic responsiveness (Kuo et al. 1988
; Nurkiewicz and Boegehold 1999
), and resting spontaneous tone was increased after exposure (), it was anticipated that arterioles from rats exposed to nanoparticles exhibit heightened myogenic responsiveness. However, myogenic responsiveness was not altered after nanoparticle exposure (). Because the isolated vessel technique removes the influence of nerves and circulating vasoactive agents, it can not be concluded that myogenic responsiveness is not altered by nanoparticle exposure. Therefore, continued investigation at a more mechanistic level is indicated.
Even though smooth muscle contractile function (assessed via myogenic responsiveness) was unaffected by nano-TiO2
exposure, coronary arterioles from nano-TiO2
rats displayed an enhanced spontaneous tone prior to the start of experiments. This is in contrast to previous reports from our lab evaluating microvessels of the spinotrapezius muscle, which showed similar spontaneous tone after exposure to either fine or nano-TiO2
(Nurkiewicz et al. 2006
). Obvious points of disparity between our previous findings and the present study include a different microvascular bed (spinotrapezius muscle), younger rats (by 3-4 weeks), and use of an in vivo
An unexpected result from the present study was the observed increase in left ventricular and heart weight in rats exposed to nano-TiO2
().. The acute rise in heart and left ventricle weight after particle exposure may be due to altered balance of fluids in the heart (i.e. an increase in inter- or intracellular fluid volume). Clearly, hydration status effects the weight of any tissue in the body (Wallace et al. 1970
). However, alterations in microvascular permeability are consistent with the functional changes in reactivity reported herein (He et al. 2006
). Nurkiewicz et al (2008)
observed potential breaches in endothelial integrity after nanoparticle exposure. In these experiments, luminal infusion of A23187 in animals exposed to nano-TiO2
produced arteriolar constriction (rather than dilation); which is highly consistent with the Ca2+
ionophore interacting with vascular smooth muscle (rather than the endothelium alone). It is important to note that the current findings in the heart are preliminary. This specific hypothesis requires continued, more rigorous experiments in order to confirm the present results.
In regards to the question of relevant human exposures, it is important to relate the current exposure paradigm to those encountered in an occupational setting. NIOSH has recently proposed recommended exposure limits (REL) of 0.1 mg/m3
). Unreported levels of nanoparticles have been measured to be as high as 1.4 mg/m3
in the occupational workplace (NIOSH Field Team, personal communication). Therefore, a worker exposed at this level could achieve a pulmonary burden equivalent to 10 μg in the rat, as used in the present study, within 5 years [normalized for alveolar epithelial surface area (Stone et al. 1992
) and taking human pulmonary deposition of a 100 nm particle to be 45% (Kreyling 2003