Abigail Durkes

Critical role of acrolein in secondary injury following ex vivo spinal cord trauma

The pathophysiology of spinal cord injury (SCI) is characterized by the initial, primary injury followed by secondary injury processes in which oxidative stress is a critical component. Secondary injury processes not only exacerbate pathology at the site of primary injury, but also result in spreading of injuries to the adjacent, otherwise healthy tissue. The lipid peroxidation byproduct acrolein has been implicated as one potential mediator of secondary injury. To further and rigorously elucidate the role of acrolein in secondary injury, a unique ex vivo model is utilized to isolate the detrimental effects of mechanical injury from toxins such as acrolein that are produced endogenously following SCI. We demonstrate that (i) acrolein–Lys adducts are capable of diffusing from compressed tissue to adjacent, otherwise uninjured tissue; (ii) secondary injury by itself produces significant membrane damage and increased superoxide production; and (iii) these injuries are significantly attenuated by the acrolein scavenger hydralazine. Furthermore, hydralazine treatment results in significantly less membrane damage 2 h following compression injury, but not immediately after. These findings support our hypothesis that, following SCI, acrolein is increased to pathologic concentrations, contributes significantly to secondary injury, and thus represents a novel target for scavenging to promote improved recovery.

Notch activation drives adipocyte dedifferentiation and tumorigenic transformation in mice

Adipocyte-specific activation of Notch signaling suppresses lipid metabolism pathways that provide ligands to Pparγ, leading to adipocyte dedifferentiation and development of liposarcomas (LPSs) resembling human dedifferentiated LPSs with complete penetrance. Pparγ ligand supplementation prevents liposarcoma development.

Murine ultrasound-guided transabdominal para-aortic injections of self-assembling type I collagen oligomers

&NA; Abdominal aortic aneurysms (AAAs) represent a potentially life‐threatening condition that predominantly affects the infrarenal aorta. Several preclinical murine models that mimic the human condition have been developed and are now widely used to investigate AAA pathogenesis. Cell‐ or pharmaceutical‐based therapeutics designed to prevent AAA expansion are currently being evaluated with these animal models, but more minimally invasive strategies for delivery could improve their clinical translation. The purpose of this study was to investigate the use of self‐assembling type I collagen oligomers as an injectable therapeutic delivery vehicle in mice. Here we show the success and reliability of a para‐aortic, ultrasound‐guided technique for injecting quickly‐polymerizing collagen oligomer solutions into mice to form a collagen‐fibril matrix at body temperature. A commonly used infrarenal mouse AAA model was used to determine the target location of these collagen injections. Ultrasound‐guided, closed‐abdominal injections supported consistent delivery of collagen to the area surrounding the infrarenal abdominal aorta halfway between the right renal artery and aortic trifurcation into the iliac and tail arteries. This minimally invasive approach yielded outcomes similar to open‐abdominal injections into the same region. Histological analysis on tissue removed on day 14 post‐operatively showed minimal in vivo degradation of the self‐assembled fibrillar collagen and the majority of implants experienced minimal inflammation and cell invasion, further confirming this materials potential as a method for delivering therapeutics. Finally, we showed that the typical length and position of this infrarenal AAA model was statistically similar to the length and targeted location of the injected collagen, increasing its feasibility as a localized therapeutic delivery vehicle. Future preclinical and clinical studies are needed to determine if specific therapeutics incorporated into the self‐assembling type I collagen matrix described here can be delivered near the aorta and locally limit AAA expansion. Graphical abstract Figure. No caption available.

In vivo investigation of acidified pepsin exposure to porcine vocal fold epithelia.

The study objective was to investigate epithelial changes in response to direct, repeated, acidified pepsin exposures in an in vivo porcine model. We hypothesized that 12 acidified pepsin applications to simulate reflux would elicit a vocal fold response characterized by inflammation, epithelial proliferation, and increased intercellular space, as well as changes in the gene expression of epithelial junctional proteins, ion transporter proteins, and proinflammatory cytokines.

Bicarbonate Availability for Vocal Fold Epithelial Defense to Acidic Challenge

Objectives: Bicarbonate is critical for acid-base tissue homeostasis. In this study we investigated the role of bicarbonate ion transport in vocal fold epithelial defense to acid challenges. Acidic insults to the larynx are common in gastric reflux, carcinogenesis and metastasis, and acute inflammation. Methods: Ion transport was measured in viable porcine vocal fold epithelium. First, 18 vocal folds were exposed to either the carbonic anhydrase antagonist acetazolamide or to vehicle. Second, 32 vocal folds were exposed to either a control buffer or a bicarbonate-free buffer on their luminal or basolateral surface or both. Third, 32 vocal folds were challenged with acid in the presence of bicarbonate-free or control buffer. Results: The vocal fold transepithelial resistance was greater than 300 Ω*cm2, suggesting robust barrier integrity. Ion transport did not change after exposure to acetazolamide (p > 0.05). Exposure to bicarbonate-free buffer did not compromise vocal fold ion transport (p > 0.05). Ion transport increased after acid challenge. This increase approached statistical significance and was the greatest for the control buffer and for the bicarbonate-free buffer applied to the basolateral surface. Conclusions: Bicarbonate secretion may contribute to vocal fold defense against acid challenge. Our data offer a potential novel role for bicarbonate as a therapeutic agent to reduce pH abnormalities in the larynx and prevent associated pathological changes.

Nondermal irritating hyperosmotic nanoemulsions reduce treatment times in a contamination model of wound healing

Increased microbial burden within the wound often complicates wound healing and may lead to subsequent infection or delayed healing. Here, we investigate a novel topical for addressing wound contamination that utilizes hyperosmotic saccharides with a cell membrane disrupting emulsion. These hyperosmotic nanoemulsions (HNE) were administered topically in a full‐thickness biopsy model of wound healing. Results show that HNE were well tolerated in noninfected animals with no indications of dermal irritation or acute toxicity. Additionally, HNE was able to reduce bacterial bioburden (Escherichia coli and Enterococcus faecalis) levels by 3 logs within 24 h when wounds were inoculated with 5 × 106 total CFU. These bactericidal values were similar to wounds treated with silver sulfadiazine. Wound closure showed HNE wounds closed in 7.6 ± 0.2 days while SSD and control required 10.2 ± 0.4 and 10.4 ± 0.3 days, respectively. HNE maintained a moist wound environment, were well debrided, and exhibited improved hemostatic response. Further histological examination revealed enhanced granulation tissue as compared to silver sulfadiazine and control cohorts. These results were corroborated with 3D topographical imprints of the wounds at day 14 which qualitatively showed a smoother surface. In contrast, silver sulfadiazine appeared to delay wound closure. Finally, dermal sensitization and irritation studies conducted in guinea pig and rabbits did not reveal any acute dermal side effects from HNE exposure. The cumulative data indicates nonantibiotic‐based HNEs may be a promising topical treatment for the management of contaminated wounds.

A Method to Administer Agents to the Larynx in an Awake Large Animal

Purpose This research note describes an adapted experimental methodology to administer an exogenous agent to the larynx and upper airway of awake animals. The exogenous agent could be a perturbation. In the current study, the agent was isotonic saline. Isotonic saline was selected because it is safe, of similar composition to extracellular fluid, and used in voice studies. The described approach allowed large animals such as pigs to be comfortably restrained without chemical sedation or anesthesia for extended periods while receiving the agent. Method Six Sinclair pigs were successfully trained with positive reinforcement to voluntarily enter and then be restrained in a Panepinto Sling. Once restrained, the pigs accepted a nose cone that delivered nebulized isotonic saline. This procedure was repeated 3 times per day for 20 days. At the end of the study, the larynx and airway tissues were excised and examined using histology and transmission electron microscopy. Results Pathology related to the procedure (i.e., nebulized inhaled isotonic saline or stress) was not identified in any examined tissues. Conclusions This methodology allowed for repeated application of exogenous agents to awake, unstressed animals. This method can be used repeatedly in the laboratory to test various therapeutics for safety, toxicity, and dosage. Future studies will specifically manipulate the type of agent to further our understanding of laryngeal pathobiology.

Cholesterol Esterification Inhibition Suppresses Prostate Cancer Metastasis by Impairing the Wnt/β-catenin Pathway

Dysregulation of cholesterol is a common characteristic of human cancers including prostate cancer. This study observed an aberrant accumulation of cholesteryl ester in metastatic lesions using Raman spectroscopic analysis of lipid droplets in human prostate cancer patient tissues. Inhibition of cholesterol esterification in prostate cancer cells significantly suppresses the development and growth of metastatic cancer lesions in both orthotopic and intracardiac injection mouse models. Gene expression profiling reveals that cholesteryl ester depletion suppresses the metastatic potential through upregulation of multiple regulators that negatively impact metastasis. In addition, Wnt/β-catenin, a vital pathway for metastasis, is downregulated upon cholesteryl ester depletion. Mechanistically, inhibition of cholesterol esterification significantly blocks secretion of Wnt3a through reduction of monounsaturated fatty acid levels, which limits Wnt3a acylation. These results collectively validate cholesterol esterification as a novel metabolic target for treating metastatic prostate cancer. Mol Cancer Res; 16(6); 974–85. ©2018 AACR.

Proton density–weighted laryngeal magnetic resonance imaging in systemically dehydrated rats

Dehydrated vocal folds are inefficient sound generators. Although systemic dehydration of the body is believed to induce vocal fold dehydration, this causative relationship has not been demonstrated in vivo. Here we investigate the feasibility of using in vivo proton density (PD)–weighted magnetic resonance imaging (MRI) to demonstrate hydration changes in vocal fold tissue following systemic dehydration in rats.

Cigarette Smoke Exposure to Pig Larynx in an Inhalation Chamber

OBJECTIVES This study investigated the effects of cigarette smoke exposure on the pig larynx using an inhalation chamber. Specifically, we compared the effects of cigarette smoke exposure from either 3 cigarettes per day (3cd) or 15 cigarettes per day (15cd) for 20days. STUDY DESIGN In vivo prospective design. METHODS Female pigs were exposed via an inhalation chamber to cigarette smoke (3R4F research cigarettes) from 3cd (n = 6) or 15cd (n = 6) for 20days. Outcomes included histopathology of vocal fold and airway tissues; gene expression of interleukins, TNF-α, and VEGF; protein levels of TNF-α and IL-6; and number of coughs recorded in the chamber. RESULTS Pigs exposed to cigarette smoke from 15cd exhibited mild vocal fold edema as compared to the 3cd group on histopathological evaluation. There was also minimal inflammation of nasal and tracheal tissue characterized by presence of more granulocytes in the 15cd group compared to the 3cd group. Cough frequency was significantly greater for the 15cd group compared to the 3cd group. CONCLUSIONS A custom-designed large animal inhalation chamber successfully challenged pigs repeatedly, to varying levels of cigarette smoke. Future studies will combine such low levels of smoke exposure with other common challenges such as acid reflux to understand the multifactorial causation of laryngeal pathologies.