Open Access Kent State (OAKS)

Chronic wound infections are typically polymicrobial, consisting of multispecies bacteria including both Gram negative and Gram positive strains. Although planktonic view of infection could explain most acute infections, it has not been adequate for understanding the pathogenesis of chronic infections. Pseudomonas aeruginosa and Staphylococcus aureus are the two most common causes of chronic wound infections and are frequently found together. Studies suggest that dual infection by P. aeruginosa and S. aureus are more virulent and/or result in worse outcomes than infection by single species bacteria alone. Although they are frequently found together in human infections, P. aeruginosa has shown to quickly kill S. aureus when the two are grown together in planktonic co-cultures in vitro, which makes it difficult to develop an appropriate model that closely reflects polymicrobial infection in human chronic wounds. The objective of this study is to establish an in vitro model for polymicrobial biofilms consisting of both S. aureus and P. aeruginosa. This was achieved by inoculating them at different ratios and determining the optimal condition that results in the growth of both bacteria.

The vascular complications following hyperglycemia and hyperlipidemia are detrimental to individuals that suffer from diabetes mellitus as it accelerates atherosclerosis of the coronary arteries, and the fibrosis of the myocardium. Hyperglycemia causes abnormalities in insulin action and secretion as well as has an effect on biochemical pathways. This leads to alterations of signal transduction pathways and protein function. Prolonged exposure to hyperglycemia can cause glycation of proteins. Changes in the function of lipoprotein metabolism due to glycation, is one of the main issues increasing the risk for developing atherosclerosis. It impairs cholesterol metabolism, and excess cholesterol builds up in arteries and is no longer able to be removed via the reverse cholesterol transport pathway. The purpose of this study is to understand why diabetes contributes to cardiac disease. This study helps to prove the hypothesis that the stability of HDL proteins is smaller and their synthesis rates much faster in hyperglycemic conditions. Thus, impairing their effectiveness in functioning. To study HDL metabolism in T2DM and how to apply ²H₂O metabolism labeling, the lab developed an algorithm that allows for the calculation of the enrichment of intracellular amino acid based on body water enrichment analysis. The results of this experimental design showed very strong correlation between hyperglycemia and increased clearance of HDL where HDL is not able to remove cholesterol adequately in the reverse cholesterol transport.

Down syndrome is a complex developmental disorder that affects over 400,000 people in the United States and results from the triplication of human chromosome 21. Down syndrome is the most prevalent genetic cause of intellectual disability. While Down syndrome has been modeled in mice, the genes orthologous to human chromosome 21 are spread between mouse chromosomes 10, 16, and 17. This means replicating this disorder in mice has been difficult. Thus, the creation of a human model of this disorder would greatly improve our understanding of Down syndrome. Here, we used direct reprogramming to convert human fibroblasts into induced neurons (iNs). We introduced four transcription factors (Brn2, Ascl1, Myt1l, and NeuroD1 (BAMN)) into human Down syndrome fibroblasts via lentiviral mediated gene transfer. Expression of these factors, in combination with specific media and growth factors, resulted in the reprogramming of human fibroblasts into neuronal cells. As a control, we also reprogrammed fibroblasts from a healthy human subject that was age, sex, and race matched to the individual with Down syndrome. We found that Down syndrome fibroblasts can be converted into induced neurons. We are currently in the process of confirming this process using neuronal-specific markers and karyotyping. This is a novel discovery as it was not previously known if Down syndrome fibroblasts could successfully be converted to neurons using direct reprogramming. Using this new human model, we can gain a better understanding into the mechanisms underlying Down syndrome, as well as gain insight into treatments for this disorder.

The phytocannabinoids tetrahydrocannabinol (Δ⁹-THC) and cannabidiol (CBD) and the endocannabinoids 2-arachidonoylethanolamine (anandamide) and 2-arachidonoylglycerol (2-AG) exhibit antiproliferative effect on cancer cells derived from diiferent organs, including thyroid, brain, prostate and breast. THC and the endocannabinoids are known agonists at the G-protein coupled receptors (CB1 and CB2), which appear to mediate the antiproliferative effect on some of the cancer cells. CBD’s antiproliferative effect, on the other hand, is mediated via CB receptor-independent mechanism. In this study, I hypothesized that ovarian cancer cell proliferation will be also inhibited by these cannabinoids. I used the SKOV3 adenocarcinoma cancer line, which exhibits both rapid proliferative and highly invasive properties, for my study. The cells were plated at subconfluent density in DMEM-F12 medium containing 10% fetal bovine serum (FBS). After overnight cell attachment to the culture dish, the cells were incubated for 24 hours in serum-free medium. Subsequently, the cells were exposed to various concentrations of the cannabinoids in serum-free medium for 48 hours. Within 24 hours of incubation with phytocannabinoids at approximately 10 µM, both THC and CBD induced apoptosis and swelling of the cells. Incubation of these cells with a DNA-binding fluorescent dye revealed fragmented nuclei. R-1 methanandamide, a metabolically stable analog of the endocannabinoid anandamide, also induced cell death but at a higher concentration (~30 µM). On the other hand, the endocannabinoid 2-AG increased the proliferation of the cells. The observed differential effects of cannabinoids on SKOV3 cell proliferation support my hypothesis in part.

Background: Transient Receptor Potential (TRP) channel stimulation induces the up-regulation of several downstream mediators that are well understood to be cardioprotective in response to ischemic injury in the heart. However, since the discovery of this cardioprotective pathway is completely novel in nature, the extent to which it is modified in the setting of diabetes presents an intriguing translational interest. We hypothesized that the TRP-induced upregulation of this cardioprotective axis is compromised in diabetes.

Methods: CMs isolated from normal and diabetic mice were treated with the TRPA1 agonist, AITC, or the TRPV1 agonist, capsaicin, in the presence or absence of TRPA1, TRPV1, Akt, eNOS or PKCε inhibitors. Nitric oxide (NO) and nitrite levels were observed utilizing confocal microscopy and the Griess assay method, respectively.

Results: AITC induced NO production through a pathway dependent on Akt and eNOS whereas capsaicin elicited NO formation through a pathway dependent upon the presence of PKCε. Furthermore, the formation of nitric oxide in the diabetic model was diminished to a significant extent. TRPA1 and TRPV1 activation stimulated nitric oxide production in diabetic CMs, but to a significantly lesser degree.

Conclusions: The extent to which TRPA1 and TRPV1 activation turns on the cardioprotective axis is severely compromised in diabetic CMs. This may, in part, contribute to the significant worsening of cardiac function observed in diabetes following ischemic events in the heart.