Study of Poly(<i>N</i>‑isopropylacrylamide-<i>co</i>-acrylic acid) (pNIPAM) Microgel Particle Induced Deformations of Tissue-Mimicking Phantom by Ultrasound Stimulation

Poly­(<i>N</i>-isopropylacrylamide) (pNIPAm) microgels (microgels) are colloidal particles that have been used extensively for biomedical applications. Typically, these particles are synthesized in the presence of an exogenous cross-linker, such as <i>N</i>,<i>N</i>′-methylenebis­(acrylamide) (BIS); however, recent studies have demonstrated that pNIPAm microgels can be synthesized in the absence of an exogenous cross-linker, resulting in the formation of ultralow cross-linked (ULC) particles, which are highly deformable. Microgel deformability has been linked in certain cases to enhanced bioactivity when ULC microgels are used for the creation of biomimetic particles. We hypothesized that ultrasound stimulation of microgels would enhance particle deformation and that the degree of enhancement would negatively correlate with the degree of particle cross-linking. Here, we demonstrate in tissue-mimicking phantoms that using ultrasound insonification causes deformations of ULC microgel particles. Furthermore, the amount of deformation depends on the ultrasound excitation frequency and amplitude and on the concentration of ULC microgel particles. We observed that the amplitude of deformation increases with increasing ULC microgel particle concentration up to 2.5 mg/100 mL, but concentrations higher than 2.5 mg/100 mL result in reduced amount of deformation. In addition, we observed that the amplitude of deformation was significantly higher at 1 MHz insonification frequency. We also report that increasing the degree of microgel cross-linking reduces the magnitude of the deformation and increases the optimal concentration required to achieve the largest amount of deformation. Stimulated ULC microgel particle deformation has numerous potential biomedical applications, including enhancement of localized drug delivery and biomimetic activity. These results demonstrate the potential of ultrasound stimulation for such applications

Complete histological evaluation of the cervix.

<p>Mucous plug (m), epithelial cells (e), mucosal chorion (c), submucosa (s).</p

Bland-Altman plots demonstrating degree of concordance between pairs of cervical elastography measurements obtained by the same (left) and by two different operators (right).

<p>Mean difference (solid line) and 95% limits of agreement (dashed lines) are shown.</p

Structures identified by histological evaluation in the cervical tissue.

<p>Mucous plug (m), epithelial cells (e), mucosal chorion (c), submucosa (s). Collagen fibres: red arrow; Macrophages: arrowhead; lymphocytes: solid arrow; fibroblasts: empty arrow.</p

Measurement of the external diameter of the cervical mucosa (green line).

<p>Measurement of the external diameter of the cervical mucosa (green line).</p

SHG zoomed images.

<p>Typical SHG zoomed images in the mucosal chorion region of (a) control and (b) treated samples, and in the submucosa region of (c) control and (d) treated sample. Scale bars: 20 <i>μ</i>m. SHG ROI images are intentionally contrasted in order to enhance the visibility of the morphological differences between the control and treated samples. Corresponding 2D FT images, thresholded at 80% of their maximal intensity, are shown in (e-h) for (e) control/mucosal chorion (maximum intensity = 971 counts, R = 0.28), (f) treated/mucosal chorion (maximum intensity = 737 counts, R = 0.39), (g) control/submucosa (maximum intensity = 581 counts, R = 0.42), (h) treated/ submucosa (maximum intensity = 412 counts, R = 0.43). Average values and standard deviations (plotted error bars) are given for 5-10 regions, measured from two different animals (* = significant at 5% level; ** = significant at 1% level).</p

Characteristics of the study population.

<p>Characteristics of the study population.</p

PGE2 and its metabolites in sheep serum just before the first elastographic exam (sample 1) and at the end of the experiment just before euthanasia (sample 2), assayed via its metabolite PGEM with Cayman’s Prostaglandin E Metabolite EIA Kit (Item No. 514531, Cayman chemical).

<p>PGE2 and its metabolites in sheep serum just before the first elastographic exam (sample 1) and at the end of the experiment just before euthanasia (sample 2), assayed via its metabolite PGEM with Cayman’s Prostaglandin E Metabolite EIA Kit (Item No. 514531, Cayman chemical).</p

Example of a ROI (white circle) of 5 mm diameter positioned in elastographic image following only anatomical criteria on the conventional B-mode image.

<p>Stiffness value estimated inside the ROI: 10.2 kPa ± 0.5 kPa [9.6—11.9] kPa.</p

SHG/2PF images.

<p>(a-c) control and (d-f) treated samples imaged in two photon microscopy, in the mucosal chorion/submucosa region of fixed slices. (a,d) Composite 2PF(grey)/SHG(green) image (scale bar: 100 <i>μ</i>m). Separated SHG/2PF are shown in (b,c) and (e,f) inside the mucosal chorion (scale bar: 100 <i>μ</i>m).</p