A significant amount of prior research has been conducted to investigate type 2 diabetes, the most prevalent form afflicting over 90% of diabetic individuals [6]. Yet, gestational diabetes is an understudied form of diabetes that is thought to share various attributes with type 2 diabetes. It was the aim of this project to investigate a proposed mechanism of the disease, the contra-insulin effect, through a cell-culture experiment. To address the question of whether glycemic and hormonal conditions of cell-culture media affect Hs 795.Pl morphology, cellular growth, and glucose uptake, immunocytochemistry (ICC) and a glucose uptake assay was performed. It was hypothesized that higher the presence of hormones, specifically lactogen, in cell culture media will exacerbate the contra-insulin effect, decreasing the glucose uptake of the Hs 795.Pl cells and inducing abhorrent cell morphology. Qualitatively, estradiol and cortisol had a severe impact on cellular morphology indicative of stress and death. As for glucose uptake, it was decreased when the hormones were isolated compared to all together with estradiol thought to be majorly inhibitory to insulin’s proper functioning. It was concluded that cell morphology, growth, and glucose uptake were detrimentally impacted by the gestational hormones, especially those of cortisol and estrogen.

A significant amount of prior research has been conducted to investigate type 2 diabetes, the most prevalent form afflicting over 90% of diabetic individuals [6]. Yet, gestational diabetes is an understudied form of diabetes that is thought to share various attributes with type 2 diabetes. It was the aim of this project to investigate a proposed mechanism of the disease, the contra-insulin effect, through a cell-culture experiment. To address the question of whether glycemic and hormonal conditions of cell-culture media affect Hs 795.Pl morphology, cellular growth, and glucose uptake, immunocytochemistry (ICC) and a glucose uptake assay was performed. It was hypothesized that higher the presence of hormones, specifically lactogen, in cell culture media will exacerbate the contra-insulin effect, decreasing the glucose uptake of the Hs 795.Pl cells and inducing abhorrent cell morphology. Qualitatively, estradiol and cortisol had a severe impact on cellular morphology indicative of stress and death. As for glucose uptake, it was decreased when the hormones were isolated compared to all together with estradiol thought to be majorly inhibitory to insulin’s proper functioning. It was concluded that cell morphology, growth, and glucose uptake were detrimentally impacted by the gestational hormones, especially those of cortisol and estrogen.

Influenza virus A (IVA) poses a serious threat to human health, killing over 25,000 Americans in the 2022 flu season alone. In the past 10 years, vaccine efficacy has varied significantly, ranging from 20-60% each season. Because IVA is subject to high antigenic shift and strain cocirculation, more effective IVA vaccines are needed to reduce the incidence of disease. Herein we report the production of a recombinant immune complex (RIC) vaccine “4xM2e” in Nicotiana benthamiana plants using agroinfiltration for use as a potential universal IVA vaccine candidate. RICs fuse antigen to the C-terminus of an immunoglobulin heavy chain with an epitope tag cognate to the antibody, promoting immune complex formation to increase immunogenicity. IVA matrix protein 2 ectodomain (M2e) is selected to serve as vaccine antigen for its high sequence conservation, as only a small number of minor mutations have occurred since its discovery in 1981 in the human sequence. However, there is some divergence in zoonotic IVA strains, and to account for this, we designed a combination of human consensus, swine, and avian M2e variants, 4xM2e. This was fused to the C terminus of the RIC platform to improve M2e immunogenicity and IVA strain coverage. The 4xM2e RIC was produced in N. benthamiana and verified with SDS-PAGE and Western blot assays, along with an analysis of complex formation and the potential for complement activation via complement C1q ELISA. With this work, we demonstrate the potential of RICs and plant-expression systems to generate universal IVA vaccine candidates.

The burden of dementia and its primary cause, Alzheimer’s disease, continue to devastate many with no available cure although present research has delivered methods for risk calculation and models of disease development that promote preventative strategies. Presently Alzheimer’s disease affects 1 in 9 people aged 65 and older amounting to a total annual healthcare cost in 2023 in the United States of $345 billion between Alzheimer’s disease and other dementias making dementia one of the costliest conditions to society (“2023 Alzheimer’s Disease Facts and Figures,” 2023). This substantial cost can be dramatically lowered in addition to a reduction in the overall burden of dementia through the help of risk prediction models, but there is still a need for models to deliver an individual’s predicted time of onset that supplements risk prediction in hopes of improving preventative care. The aim of this study is to develop a model used to predict the age of onset for all-cause dementias and Alzheimer’s disease using demographic, comorbidity, and genetic data from a cohort sample. This study creates multiple regression models with methods of ordinary least squares (OLS) and least absolute shrinkage and selection operator (LASSO) regression methods to understand the capacity of predictor variables that estimate age of onset for all-cause dementia and Alzheimer’s disease. This study is unique in its use of a diverse cohort containing 346 participants to create a predictive model that originates from the All of Us Research Program database and seeks to represent an accurate sampling of the United States population. The regression models generated had no predictive capacity for the age of onset but outline a simplified approach for integrating public health data into a predictive model. The results from the generated models suggest a need for continued research linking risk factors that estimate time of onset.

This project uses All of Us Data to analyze how well of a predictor APOE ε4 is in the Latinx community, a high grandparent care community. APOE is used as a predictor for Alzheimer’s disease, but it is unknown, due to the lack of studies, how strong of a predictor it will be for Latinx individuals. This project aims to understand if the increased risk of Alzheimer’s disease among Hispanics is associated with a different level of ε4 gene frequency.

Memory CD8+ T cells protect against secondary viral infections. They develop and maintain exclusively in circulation (e.g. central memory - Tcm) or are excluded from re-circulation (resident memory - Trm). The extracellular ATP receptor P2RX7 promotes both Tcm and Trm generation. High (P2RX7hi) P2RX7-expressing early effector cells show survival, memory and pluripotency genes. Conversely, many terminal effector (TE) and apoptosis genes are upregulated in low (P2RX7lo) P2RX7-expressing cells. Among these genes is the zinc-finger transcriptional repressor Zeb2, which promotes TE differentiation at the expense of the memory cell pool. Given that Zeb2 was higher in P2RX7lo early effector cells, we postulated that Zeb2 ablation would allow P2RX7-deficient CD8+ T cells to skew towards memory subsets. To test this, we used RNP-based CRISPR-Cas9 to knockout Zeb2 in wild type or P2RX7-deficient P14 cells. At the memory timepoint, Zeb2 ablation led to a rescue of the ability of P2RX7-deficient cells to differentiate into the CD62L+ Tcm and CD69hiCD103hi Trm subsets, as well as increase the population of each. Our data suggest that P2RX7 imprints a pro-memory signature that is, to some extent, dependent on the negative regulation of Zeb2.

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.

Contemporary vaccine development relies less on empirical methods of vaccine construction, and now employs a powerful array of precise engineering strategies to construct immunogenic live vaccines. In this review, we will survey various engineering techniques used to create attenuated vaccines, with an emphasis on recent advances and insights. We will further explore the adaptation of attenuated strains to create multivalent vaccine platforms for immunization against multiple unrelated pathogens. These carrier vaccines are engineered to deliver sufficient levels of protective antigens to appropriate lymphoid inductive sites to elicit both carrier-specific and foreign antigen-specific immunity. Although many of these technologies were originally developed for use in Salmonella vaccines, application of the essential logic of these approaches will be extended to development of other enteric vaccines where possible. A central theme driving our discussion will stress that the ultimate success of an engineered vaccine rests on achieving the proper balance between attenuation and immunogenicity. Achieving this balance will avoid over-activation of inflammatory responses, which results in unacceptable reactogenicity, but will retain sufficient metabolic fitness to enable the live vaccine to reach deep tissue inductive sites and trigger protective immunity. The breadth of examples presented herein will clearly demonstrate that genetic engineering offers the potential for rapidly propelling vaccine development forward into novel applications and therapies which will significantly expand the role of vaccines in public health.