Triple Negative Breast Cancer (TNBC), indicated by the absence of estrogen, progesterone and human epidermal growth factor receptor 2 (HER2), is the most aggressive form of breast cancer characterized by high rates of metastasis and low survival. Among those diagnosed with TNBC, 34% contain Inhibitor of Growth 4 (ING4) deletion that is associated with poor patient outcomes. We previously showed that ING4 negatively regulates NF-B in breast cancer. Previous studies show parthenolide, a compound found in feverfew (Tanacetum parthenium) to inhibit NF-B in cervical and gastric cancer. We hypothesized that parthenolide inhibits cytokine-induced activation of NF-B in ING4 deficient TNBC cells. To test the hypothesis, previously established vectors, v2, ING4 wildtype and v2h1, ING4-deleted were synthesized in MDA-MB 231, a TNBC cell line, using a CRISPR/Cas9 system. Inflammatory cytokines, IL-1 and TNF, were tested in ING4 wildtype or ING4 deleted cells for elicited phosphorylation of NF-B, proliferation, and migration in the presence or absence of parthenolide. The results showed that TNF or IL-1 induced translocation phosphorylation of NF-B regardless of ING4 deletion. ING4 inhibited proinflammatory cytokine induced pp65, consistent with previous studies demonstrating the negative regulation of NF-B in ING4-sufficent cell lines. We found the optimal working dose of parthenolide, 100nM, had no effect on cell proliferation in the presence or absence of IL-1. Parthenolide inhibited IL-1induced phosphorylation of NF-B regardless of ING4 deletion. Parthenolide inhibited TNF-induced phosphorylation of NF-B in ING4-deleted cell lines. Moreover, parthenolide induced migration of TNBC cells regardless of ING4 presence of absence. TNF and parthenolide treated samples in ING4-deleted cell lines were found to inhibit cell migration to basal level. These results demonstrate the difference in inhibitory mechanism of parthenolide in induced phosphorylation of NF-B through proinflammatory cytokines TNF or IL-1This is demonstrated by the exclusivity of parthenolide inhibition of TNF induced phosphorylation of NF-B in ING4-deleted TNBC cell line. In contrast, parthenolide inhibition of IL-1 induced phosphorylation of NF-B occurred regardless of ING4 deletion. These results may inhibit parthenolide as an alternative to those with ING4-deleted TNBC due to its role in inducing cancer phenotype cell migration.

Stress is a necessary and functional part of human physiology. From responding to life-threatening situations to getting people out of bed in the morning, stress serves a major purpose in human survival. However, when consistent and high levels of stress are experienced, it can pose a threat to human health. One of the major mediators of physiological stress is a hormone called cortisol. Cortisol is a well-defined substance and its function in normal physiology is well understood. Scientific research indicates that consistent and high levels of this hormone may be an aid in cancer’s ability to evade the human immune response. Despite this, there is not much known about its relationship with cancer. I used immunofluorescence to determine cell-to-cell variability of vimentin expression and DNA content for cells that were exposed to cortisol at consistent and frequent doses overtime and those not exposed to cortisol to determine if cortisol altered the variability of vimentin expression and DNA content. I observed no change in the variability in vimentin expression across both cell conditions. I did observe variability in DNA content across both cell conditions, with more variability in the population affected by cortisol. These results suggest that there might be a relationship between the stress induced by cortisol, taking place at the genomic level but may have no impact on specific protein expression. Potential implications of the research conducted are looks to preventative medicine in the context of stress experienced by members of marginalized groups as a way of preventing cancer development.

Human protein diversity arises as a result of alternative splicing, single nucleotide polymorphisms (SNPs) and posttranslational modifications. Because of these processes, each protein can exists as multiple variants in vivo. Tailored strategies are needed to study these protein variants and understand their role in health and disease. In this work we utilized quantitative mass spectrometric immunoassays to determine the protein variants concentration of beta-2-microglobulin, cystatin C, retinol binding protein, and transthyretin, in a population of 500 healthy individuals. Additionally, we determined the longitudinal concentration changes for the protein variants from four individuals over a 6 month period. Along with the native forms of the four proteins, 13 posttranslationally modified variants and 7 SNP-derived variants were detected and their concentration determined. Correlations of the variants concentration with geographical origin, gender, and age of the individuals were also examined. This work represents an important step toward building a catalog of protein variants concentrations and examining their longitudinal changes.

Bacterial lipopolysaccharides (LPS) are structural components of the outer membranes of Gram-negative bacteria and also are potent inducers of inflammation in mammals. Higher vertebrates are extremely sensitive to LPS, but lower vertebrates, like fish, are resistant to their systemic toxic effects. However, the effects of LPS on the fish intestinal mucosa remain unknown. Edwardsiella ictaluri is a primitive member of the Enterobacteriaceae family that causes enteric septicemia in channel catfish (Ictalurus punctatus). E. ictaluri infects and colonizes deep lymphoid tissues upon oral or immersion infection. Both gut and olfactory organs are the primary sites of invasion. At the systemic level, E. ictaluri pathogenesis is relatively well characterized, but our knowledge about E. ictaluri intestinal interaction is limited. Recently, we observed that E. ictaluri oligo-polysaccharide (O-PS) LPS mutants have differential effects on the intestinal epithelia of orally inoculated catfish. Here we evaluate the effects of E. ictaluri O-PS LPS mutants by using a novel catfish intestinal loop model and compare it to the rabbit ileal loop model inoculated with Salmonella enterica serovar Typhimurium LPS. We found evident differences in rabbit ileal loop and catfish ileal loop responses to E. ictaluri and S. Typhimurium LPS. We determined that catfish respond to E. ictaluri LPS but not to S. Typhimurium LPS. We also determined that E. ictaluri inhibits cytokine production and induces disruption of the intestinal fish epithelia in an O-PS-dependent fashion. The E. ictaluri wild type and ΔwibT LPS mutant caused intestinal tissue damage and inhibited proinflammatory cytokine synthesis, in contrast to E. ictaluri Δgne and Δugd LPS mutants. We concluded that the E. ictaluri O-PS subunits play a major role during pathogenesis, since they influence the recognition of the LPS by the intestinal mucosal immune system of the catfish. The LPS structure of E. ictaluri mutants is needed to understand the mechanism of interaction.