Nanoparticle inhalation augments particle-dependent systemic microvascular dysfunction

Abstract

Background: We have shown that pulmonary exposure to fine particulate matter (PM) impairs endothelium dependent dilation in systemic arterioles. Ultrafine PM has been suggested to be inherently more toxic by virtue of its increased surface area. The purpose of this study was to determine if ultrafine PM (or nanoparticle) inhalation produces greater microvascular dysfunction than fine PM. Rats were exposed to fine or ultrafine TiO2 aerosols (primary particle diameters of ~1 mum and ~21 nm, respectively) at concentrations which do not alter bronchoalveolar lavage markers of pulmonary inflammation or lung damage.

Results: By histopathologic evaluation, no significant inflammatory changes were seen in the lung. However, particle-containing macrophages were frequently seen in intimate contact with the alveolar wall. The spinotrapezius muscle was prepared for in vivo microscopy 24 hours after inhalation exposures. Intraluminal infusion of the Ca2+ ionophore A23187 was used to evaluate endothelium-dependent arteriolar dilation. In control rats, A23187 infusion produced dose-dependent arteriolar dilations. In rats exposed to fine TiO2, A23187 infusion elicited vasodilations that were blunted in proportion to pulmonary particle deposition. In rats exposed to ultrafine TiO2, A23187 infusion produced arteriolar constrictions or significantly impaired vasodilator responses as compared to the responses observed in control rats or those exposed to a similar pulmonary load of fine particles.

Conclusion: These observations suggest that at equivalent pulmonary loads, as compared to fine TiO2, ultrafine TiO2 inhalation produces greater remote microvascular dysfunction.

Figure 1

Schematic diagram of the TiO₂ inhalation exposure system. Data-logging Real-time Aerosol Monitor (DataRAM). Aerodynamic Particle Sizer (APS). Scanning Mobility Particle Sizer (SMPS). Micro-Orifice Uniform Deposit Impactors (MOUDI).

Figure 2

Particle size distributions (Geometric Diameter, D_g) of ultrafine TiO₂ (A) and fine TiO₂ (B) aerosols generated by the inhalation exposure system. The ultrafine TiO₂ aerosol has a primary mode at 100 nm and a secondary mode at 400 nm. The fine TiO₂ aerosol has a primary mode at 710 nm and a secondary mode at 120 nm. These number-based size distributions were determined by combining the data from SMPS and APS.

Figure 3

Histopathologic alterations were subtle in the lungs of rats inhaling fine TiO₂. These changes principally consisted of particle accumulation. Particles were difficult to see using transmitted light (A) but were birefringent when visualized using cross-polarized light, enhancing their detection (B). The particle accumulation included apparently free TiO₂ particles (open arrow), TiO₂ particles within morphologically normal alveolar macrophages (dashed arrows), and TiO₂ particles within macrophages without nuclei (anuclear macrophages, solid arrows). Both morphologically normal and anuclear macrophages were frequently in contact with the alveolar wall. H&E section. Reference bar = 20 μm.

Figure 4

Histopathologic alterations were particularly subtle in the lungs of rats inhaling ultrafine TiO₂. Ultrafine particle accumulation in alveolar macrophages (arrows) was difficult to see with transmitted light (A). Ultrafine particle accumulation within alveolar macrophages was more easily visualized as birefringent intracytoplasmic material using cross-polarized light (B). Macrophages containing ultrafine TiO₂ particles were frequently in contact with the alveolar wall. H&E section. Reference bar = 20 μm.

Figure 5

Fine TiO₂ inhalation impairs systemic arteriolar dilation 24 hours after exposure in a dose-dependent manner. Sham/Control, n = 8; 90 μg, n = 8; 67 μg, n = 8; 36 μg, n = 8; 20 μg, n = 7; 8 μg, n = 12. Values are means ± SE. *, P < 0.05 vs. all groups. †, P < 0.05 vs. Sham/Control group. ‡, P < 0.05 vs. 8–36 μg groups. Adenosine (ADO).

Figure 6

Ultrafine TiO₂ inhalation impairs systemic arteriolar dilation 24 hours after exposure in a dose-dependent manner. Sham/Control, n = 8; 38 μg, n = 9; 19 μg, n = 11; 10 μg, n = 8; 6 μg, n = 7; 4 μg, n = 9. Values are means ± SE. *, P < 0.05 vs. 19 μg group. †, P < 0.05 vs. 10 μg group. ‡, P < 0.05 vs. 6 μg group. Adenosine (ADO).

Figure 7

Manipulation of ultrafine TiO₂ inhalation exposure time or aerosol concentration does not alter the impairment of systemic arteriolar dilation. 2 hr@12 mg/m³, n = 10; 4 hr@6 mg/m³, n = 8; 8 hr@3 mg/m³, n = 10. Values are means ± SE. *, P < 0.05 vs. Sham/Control group. Adenosine (ADO).

Figure 8

Systemic arteriolar dilation dose-response relationships at various inhalation exposure burdens of ultrafine vs. fine TiO₂. Open bars in all panels, Sham/Control group. Panel A, grey bars = 8 μg fine TiO₂; black bars = 10 μg ultrafine TiO₂. Panel B, grey bars = 20 μg fine TiO₂; black bars = 19 μg ultrafine TiO₂. Panel C, grey bars = 36 μg fine TiO₂; black bars = 38 μg ultrafine TiO₂. *, P < 0.05 vs. Sham/Control and fine TiO₂ at the same ejection pressure. Values are means ± SE.