Angiogenesis and airway reactivity in asthmatic Brown Norway rats.

Expanded and aberrant bronchial vascularity, a prominent feature of the chronic asthmatic airway, might explain persistent airway wall edema and sustained leukocyte recruitment. Since it is well established that there are causal relationships between exposure to house dust mite (HDM) and the development of asthma, determining the effects of HDM in rats, mammals with a bronchial vasculature similar to humans, provides an opportunity to study the effects of bronchial angiogenesis on airway function directly. We studied rats exposed bi-weekly to HDM (Der p 1; 50 µg/challenge by intranasal aspiration, 1, 2, 3 weeks) and measured the time course of appearance of increased blood vessels within the airway wall. Results demonstrated that within 3 weeks of HDM exposure, the number of vessels counted within airway walls of bronchial airways (0.5-3 mm perimeter) increased significantly. These vascular changes were accompanied by increased airway responsiveness to methacholine. A shorter exposure regimen (2 weeks of bi-weekly exposure) was insufficient to cause a significant increase in functional vessels or reactivity. Yet, 19F/1H MR imaging at 3T following αvß3-targeted perfluorocarbon nanoparticle infusion revealed a significant increase in 19F signal in rat airways after 2 weeks of bi-weekly HDM, suggesting earlier activation of the process of neovascularization. Although many antigen-induced mouse models exist, mice lack a bronchial vasculature and consequently lack the requisite human parallels to study bronchial edema. Overall, our results provide an important new model to study the impact of bronchial angiogenesis on chronic inflammation and airways hyperreactivity.

Molecular photoacoustic imaging of angiogenesis with integrin-targeted gold nanobeacons.

Photoacoustic tomography (PAT) combines optical and acoustic imaging to generate high-resolution images of microvasculature. Inherent sensitivity to hemoglobin permits PAT to image blood vessels but precludes discriminating neovascular from maturing microvasculature. α(v)ß(3)-Gold nanobeacons (α(v)ß(3)-GNBs) for neovascular molecular PAT were developed, characterized, and demonstrated in vivo using a mouse Matrigel-plug model of angiogenesis. PAT results were microscopically corroborated with fluorescent α(v)ß(3)-GNB localization and supporting immunohistology in Rag1(tm1Mom) Tg(Tie-2-lacZ)182-Sato mice. α(v)ß(3)-GNBs (154 nm) had 10-fold greater contrast than blood on an equivolume basis when imaged at 740 nm to 810 nm in blood. The lowest detectable concentration in buffer was 290 nM at 780 nm. Noninvasive PAT of angiogenesis using a 10-MHz ultrasound receiver with α(v)ß(3)-GNBs produced a 600% increase in signal in a Matrigel-plug mouse model relative to the inherent hemoglobin contrast pretreatment. In addition to increasing the contrast of neovessels detected at baseline, α(v)ß(3)-GNBs allowed visualization of numerous angiogenic sprouts and bridges that were undetectable before contrast injection. Competitive inhibition of α(v)ß(3)-GNBs with α(v)ß(3)-NBs (no gold particles) almost completely blocked contrast enhancement to pretreatment levels, similar to the signal from animals receiving saline only. Consistent with other studies, nontargeted GNBs passively accumulated in the tortuous neovascular but provided less than half of the contrast enhancement of the targeted agent. Microscopic studies revealed that the vascular constrained, rhodamine-labeled α(v)ß(3)-GNBs homed specifically to immature neovasculature (PECAM(+), Tie-2(-)) along the immediate tumor periphery, but not to nearby mature microvasculature (PECAM(+), Tie-2(+)). The combination of PAT and α(v)ß(3)-GNBs offered sensitive and specific discrimination and quantification of angiogenesis in vivo, which may be clinically applicable to a variety of highly prevalent diseases, including cancer and cardiovascular disease.

Detecting vascular biosignatures with a colloidal, radio-opaque polymeric nanoparticle.

A synthetic methodology for developing a polymeric nanoparticle for targeted computed tomographic (CT) imaging is revealed in this manuscript. The work describes a new class of soft type, vascularly constrained, stable colloidal radio-opaque metal-entrapped polymeric nanoparticle using organically soluble radio-opaque elements encapsulated by synthetic amphiphile. This agent offers several-fold CT signal enhancement in vitro and in vivo demonstrating detection sensitivity reaching to the low nanomolar particulate concentration range.

Subharmonic Response of Polymer Contrast Agents Based on the Empirical Mode Decomposition.

The subharmonic threshold for ultrasound contrast agents has been defined as a 20-25 dB difference between the fundamental and subharmonic (2/1) spectral components of the backscatter signal. However, this Fourier-based criterion assumes a linear time-invariant signal. A more appropriate criterion for short cycle and frequency-modulated waveforms is proposed with an adaptive signal-processing approach based on the empirical mode decomposition (EMD) method. The signal is decomposed into an orthogonal basis known as intrinsic mode functions (IMFs) and a subharmonic threshold is defined with respect to the energy ratio of the subharmonic IMF component to that of the incident signal. The method is applied to backscatter data acquired from two polymer-shelled contrast agents, Philips (#38, mean diameter 2.0 [Formula: see text]) and Point Biomedical (#12027, mean diameter 3.9 [Formula: see text]). The acoustic backscatter signals are investigated for a single contrast agent subjected to monofrequency (20 MHz, 20 cycles) and chirp (15-25 MHz, 20 cycles) forcing for incident pressures ranging from 0.5 to 2.4 MPa. In comparison to the spectral peak difference (20 dB) criterion, the EMD method is more sensitive in determining subharmonic signals.

Correlation of rupture dynamics to the nonlinear backscatter response from polymer-shelled ultrasound contrast agents.

Polymer-shelled ultrasound contrast agents (UCAs) may expel their encapsulated gas subject to ultrasound-induced shell buckling or rupture. Nonlinear oscillations of this gas bubble can produce a subharmonic component in the ultrasound backscatter. This study investigated the relationship between this gas-release mechanism and shell-thickness-to-radius ratios (STRRs) of polymer-shelled UCAs. Three types of polylactide-shelled UCAs with STRRs of 7.5, 40, and 100 nm/µm were studied. Each UCA population had a nominal mean diameter of 2 µm. UCAs were subjected to increasing static overpressure ranging from 2 to 330 kPa over a duration of 2 h in a custom-designed test chamber while being imaged using a 200× magnification video microscope at a frame rate of 5 frames/s. Digitized video images were binarized and processed to obtain the cross-sectional area of individual UCAs. Integration of the normalized cross-sectional area over normalized time, defined as buckling factor (Bf), provided a dimensionless parameter for quantifying and comparing the degree of pre-rupture buckling exhibited by the UCAs of different STRRs in response to overpressure. The UCAs with an STRR of 7.5 nm/µm exhibited a distinct shell-buckling phase before shell rupture (Bf < 1), whereas the UCAs with higher STRRs (40 and 100 nm/µm) did not undergo significant prerupture buckling (Bf ≈ 1). The difference in the overpressure response was correlated with the subharmonic response produced by these UCAs. When excited using 20-MHz ultrasound, individual UCAs (N = 3000) in populations that did not exhibit a buckling phase produced a subharmonic response that was an order of magnitude greater than the UCA population with a prominent pre-rupture buckling phase. These results indicate the mechanism of gas expulsion from these UCAs might be a relevant factor in determining the level of subharmonic response in response to high-frequency ultrasound.

Circadian regulation of cortisol after hippocampal damage in humans.

BACKGROUND: There is substantial evidence that the hippocampus (HC) regulates the activity of the hypothalamic-pituitary-adrenocortical (HPA) axis. Damage to the HC in animals produces a transient alteration in diurnal and stress-related HPA activity. This study was designed to examine the effects of HC damage on basal cortisol secretion in humans. METHODS: Salivary cortisol was measured in 22 patients with HC damage (12 with bilateral damage and 10 with unilateral damage), 7 brain-damaged comparison participants, 10 healthy, age-matched comparison participants, and 6 of the patients' caregivers. Salivary cortisol samples were taken immediately after awakening, 30 min after awakening, at 8:00 am, 11:00 am, 3:00 pm, 6:00 pm, and at bedtime on a single day. Brain-injured patients underwent a structural magnetic resonance imaging scan to examine quantitative volumes of the HC. RESULTS: Both bilateral and unilateral HC damage abolished the cortisol response to awakening documented in the comparison groups. Caregivers of bilateral HC patients showed a reduced response to awakening. The remainder of the circadian pattern was not affected in the HC patients; all groups showed a significant diurnal variation. There was no association between HC volume and cortisol secretion. CONCLUSIONS: Hippocampal damage in humans abolishes the cortisol response to awakening, whereas the remainder of the diurnal cycle is unaffected in these patients. These data suggest a unique role of the HC in the control of basal cortisol secretion.

Dynamics and fragmentation of thick-shelled microbubbles.

Localized delivery could decrease the systemic side effects of toxic chemotherapy drugs. The unique delivery agents we examine consist of microbubbles with an outer lipid coating, an oil layer, and a perfluorobutane gas core. These structures are 0.5-12 microm in radius at rest. Oil layers of these acoustically active lipospheres (AALs) range from 0.3-1.5 microm in thickness and thus the agents can carry a large payload compared to nano-scale drug delivery systems. We show that triacetin-based drug-delivery vehicles can be fragmented using ultrasound. Compared with a lipid-shelled contrast agent, the expansion of the drug-delivery vehicle within the first cycle is similar, and a subharmonic component is demonstrated at an equivalent radius, frequency, and driving pressure. For the experimental conditions explored here, the pulse length required for destruction of the drug-delivery vehicle is significantly greater, with at least five cycles required, compared with one cycle for the contrast agent. For the drug-delivery vehicle, the observed destruction mechanism varies with the initial radius, with microbubbles smaller than resonance size undergoing a symmetric collapse and producing a set of small, equal-sized fragments. Between resonance size and twice resonance size, surface waves become visible, and the oscillations become asymmetrical. For agents larger than twice the resonance radius, the destruction mechanism changes to a pinch-off, with one fragment containing a large fraction of the original volume.

Excitation of polymer-shelled contrast agents with high-frequency ultrasound.

Few experimental and complementary theoretical studies have investigated high-frequency (>20 MHz) nonlinear responses from polymer-shelled ultrasound contrast agents. Three polymer agents with different shell properties were examined for their single-bubble backscatter when excited with a 40 MHz tone burst. Higher-order harmonic responses were observed for the three agents; however, their occurrence was at least partly due to nonlinear propagation. Only one of the agents (1.1 microm mean diameter) showed a subharmonic response for longer excitations (approximately 10-15 cycles) and midlevel pressure excitations ( 2.5 MPa). Theoretical calculations of the backscattered spectrum revealed behavior similar to the experimental results in specific parameter regimes.