MEDICA@MUSC (Medical University of South Carolina)
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MALDI-MSI Identification of Tissue-Level N-Glycomic and Proteomic Molecular Biomarkers of Aggressive Prostate Cancer
No full textProstate Cancer (PCa) poses a significant clinical challenge, characterized by debates on screening efficacy and continuous issues with distinguishing aggressive from non-aggressive tumors. Despite treatment advancements, poor patient outcome at late stages and clinical overtreatment of indolent disease emphasizes the urgent need for markers of PCa aggression. Aberrant glycosylation and extracellular matrix (ECM) remodeling are two complex molecular processes necessary for PCa progression, highlighting a novel area for biomarker discovery. Leveraging formalin-fixed paraffin-embedded tissues, tissue microarrays (TMAs), and matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI), distinct N-glycan and ECM profiles are identified. Elevated bisecting, multi-antennary, and fucosylated N-glycans indicate early signs of increased risk for PCa metastasis in a novel patient outcome TMA cohort. Serial enzymatic digests of tissues revealed samples rich in N-glycans with core-fucose modifications, specifically, exhibited greater risk for eventual metastasis. Further analysis of large tissue samples from which TMAs were derived revealed a unique spatial distribution of these N-glycans of interest throughout both tumor and tumor-adjacent microenvironments, providing novel vi insight into the roles of these structures in disease progression. In further pursuit of increased fucosylation as a marker for PCa aggression, analysis of a TMA constructed from tissues with heightened risk of neuroendocrine differentiation and castrate-resistance revealed a greater abundance of fucosylated N-glycans. Investigating differences in ECM composition between tissue samples from PCa that eventually metastasized and PCa that never recurred sheds light on potential prognostic markers that have potential to improve patient outcome. Proteomic analysis of these tissues revealed several protein species exclusive to PCa at risk for metastasis. These biomarkers hold transformative implications for tailoring patient-centric treatment strategies and foster a comprehensive understanding of disease dynamics at a molecular level
The Role of Sphingolipids in T-Cell Mitophagy, Metabolism, and Signaling
No full textSphingolipids are bioactive molecules involved in several cellular processes, from maintaining cellular and subcellular structural integrity and barrier function to regulating and transducing signaling cascades in various cellular contexts. In this dissertation, I describe the involvement of two key sphingolipids – ceramide and sphingosine-1-phosphate (S1P) – in T-cell metabolism, function, and signaling.
The first part of the dissertation builds upon the recently discovered role of ceramide synthase 6 (CerS6)-generated C14/16-ceramide in inducing mitophagy in aging T-cells, which in turn attenuates T-cell anti-tumor function. This ultimately contributes to the reduced cancer resistance in the aging population. Interestingly, epidemiological studies have shown that cancer incidence is significantly less in aging Alzheimer’s Disease (AD) patients without much mechanistic insight. I aimed to understand this cancer resistance mechanism linked to AD and exploit it for broader cancer therapies. A hallmark of AD is the accumulation of amyloid precursor protein (APP) in neuronal mitochondria, inhibiting mitophagy, the clearance of dysfunctional mitochondria, and contributing to AD pathology. Unlike neurons, attenuation of ceramide-dependent mitophagy would play a favorable role in aging T-cells, as it would enhance their anti-tumor function. Given that APP is also processed in T-cells, I hypothesized that APP accumulates aberrantly in aging Alzheimer’s T-cells, inhibiting mitophagy and enhancing T-cell anti-tumor activity. Employing techniques of lipidomics, metabolomics, imaging, molecular biology, and genetic models, I show that AD T-cells from a mouse model and human patients are protected against aging-associated ceramide-dependent mitophagy, which enhanced their anti-tumor functions. The excessive localization of APP to the T-cell mitochondria, where it inhibits the accumulation of CerS6 and C14/C16-ceramides, attenuated the levels of mitophagy and protected the Alzheimer’s T-cells from metabolic defects observed in aging, namely protecting against the exhaustion of fumarate pools. Both in vitro and in vivo re-supplementation of fumarate to aging WT mouse T-cells effectively rescued their tumor killing capacity both in vitro and in adoptive T-cell transfers to mice with melanoma, functionally mimicking the AD phenotype. Moreover, I show that Alzheimer’s mice are protected against the growth of both carcinogen-induced and allografted tumors. Finally, transfer of mitochondria from AD T-cells to aging WT T-cells improved their metabolic fitness and anti-cancer functions. In summary, I show that AD-associated APP improves anti-tumor immunity of aging T-cells by inhibiting ceramide-mediated lethal mitophagy and restoring mitochondrial fumarate metabolism. These data provide a novel mechanism to explain reduced cancer incidences in AD patients.
In the second part of this dissertation, I turn my attention to another molecule at the center of sphingolipid metabolism: S1P, a bioactive lysophospholipid generated by two isoenzymes of the sphingosine kinase protein (Sphk1 and Sphk2), which plays an important role in T-cell function from activation and differentiation to egress and migration. While the role of Sphk1-generated S1P has been studied before, the role of Sphk2 in modulating T-cell functionality remains unclear and poorly defined. I show here that Sphk2 interacts with the Zeta Chain of T Cell Receptor Associated Protein Kinase 70 (ZAP70), a kinase essential for the initiation of canonical biochemical signaling pathways upon activation via T-cell antigen receptor (TCR) stimulation. ZAP70 activity is strictly regulated by phosphorylation and dephosphorylation events which act as an on-off switch that keeps the downstream signaling cascades in check. This is especially interesting as Sphk2-derived S1P has been shown to mimic protein phosphorylation in other proteins, and I hypothesized that it might be playing a similar role upon interacting with ZAP70 in T-cells. Using both pharmacological inhibition of Sphk2 as well as Sphk2 knock-out mice, I show a reduction in the time and intensity of ZAP70 phosphorylation and downstream signal propagation upon TCR activation in the absence of Sphk2. Molecular docking predictions and mutational studies identified the amino acid R160 of ZAP70 to be interacting with Sphk2-derived S1P, which in turn supports ZAP70 activity and functionality by mimicking its phosphorylation at T130 or Y126 residues upon T-cell receptor activation. This work gives new insights into the previously unexplored role of endogenous S1P, generated by SphK2, in the regulation of T-cell activation via regulating ZAP70 signaling, which plays key roles in inflammation and autoimmune diseases
The Impact of Health Insurance Coverage on Short-Term and Long-Term Readmissions to the Emergency Department for Pediatric Patients with Asthma: A Retrospective Regression Analysis
Get PDFInadequate health insurance coverage may hinder the management of asthma in children in recurrent emergency department (ED) visits in comparison to children with health care insurance. Children with asthma who do not have health insurance are at risk of not having access to primary care physicians. They may live in poorer communities, where they are more likely to encounter higher concentrations of asthma-inducing agents or factors contributing to increased healthcare utilization, resulting in increased expenditures and reduced patient outcomes
Oral Feeding of Infants Who Require Noninvasive Respiratory Support: Retrospective Investigations
Get PDFThere is a dearth of evidence supporting the safety and efficacy of orally feeding infants who require noninvasive respiratory support (NIRS). This dissertation investigates clinical practices surrounding orally feeding infants on NIRS. Manuscript 1 systematically reviews parameters impacting study of this practice. Findings emphasize the dearth of objective data surrounding this practice, and that additional research must be conducted before safety and efficacy outcomes of this practice can be established. Manuscript 2 retrospectively uses electronic health record data to describe which infant populations are orally fed during NIRS use, amount of NIRS used during infant oral feeding, changes in practice over time, and predictors for oral feeding during NIRS use, length of stay, and time on NIRS. Findings support that practice is variable, that patient-level factors can predict which infants are more likely to be fed on NIRS, and that orally feeding during NIRS use can impact length of stay and time on NIRS. The third paper qualitatively investigates clinical practices specific to feeding expert assessment and intervention in this population. Missing data in clinical notes limit ability to interpret safety and quality measures of this practice, and we used themes from documentation to recommend documentation practices
H2S-Prdx4 Axis Mitigates Golgi Stress to Bolster Tumor-Reactive T Cell Immunotherapeutic Response
No full textNew approaches to improve adoptive cell therapy (ACT) protocols are needed to enhance persistence of adoptively transferred T cells and overcome tumor-induced immunosuppression and cellular stress. In the first part of this dissertation, we show that exogenous hydrogen sulfide (H2S) can be used to promote the anti-tumor immune response. T cells treated ex vivo with an H2S donor (GYY4137) or overexpressing cystathionine β-synthase (Cbs), which is a key H2S-producing enzyme) display an increase in stemness and antioxidant capacity, enhanced production of cytolytic cytokines and protein translation, and superior tumor control upon ACT using in vivo models of melanoma and lymphoma. Global proteomics analysis revealed that H2S promotes thiolation at key cysteine residues on proteins involved in regulating ER and Golgi function. In the second part of this dissertation, we show that tumor microenvironment-mediated disruption of Golgi architecture and function, termed Golgi stress, can be mitigated by treating anti-tumor T cells with H2S or over-expressing Cbs. We further show that the H2S-induced increase in antioxidant capacity and protein translation is mediated in part by ER-Golgi shuttling of Peroxiredoxin-4. Lastly, we identify that T cells possessing high Golgi content (Golgihi T cells) exhibit unique metabolic and glycation signatures with enhanced anti-tumor capacity. These data demonstrate that strategies to mitigate Golgi network stress or using Golgihi tumor-reactive T cells can improve tumor control upon adoptive transfer
Financial Implications Associated with Laboratories Performing Laboratory Developed Tests For Medicare and Commercial Patients
Get PDFObjective: To quantify missed reimbursements associated with specific CPT codes for Laboratory Developed Tests (LDTs) via polymerase chain reaction (PCR) in the United States during 2021.
Study Design: Quantitative, retrospective observational study to identify the dollar amount of missed reimbursements associated with 19 specific CPT codes billed for Medicare patients who had LDTs performed via PCR in the United States in 2021.
Data Sources: 19 specific CPT codes from Medicare and Commercial patients with LDT PCR testing performed using 2021 Meridian MarketScan data.
Key Results: Medicare represented 1.7% of the sample size with a potential lost revenue of $1M where Medicare reimbursed higher per CPT code when claims were paid, but had a higher zero dollar payment rate when compared to Commercial claims for the same 19 CPT codes
Pediatric Burn Injuries: Addressing Psychosocial Concerns During the Acute Care Phase of Treatment
Get PDFNeural Control of Swallowing in Infants: White Matter Tracts Associated with Swallow Function
No full textThis dissertation investigates innovative approaches to improve feeding and swallowing outcomes in infants, particularly those born prematurely or with brain injury. It focuses on understanding the neural networks involved in sucking and swallowing by utilizing diffusion kurtosis imaging (DKI), a more sensitive neuroimaging technique than traditional MRI, and the BaByVFSSImP, a standardized tool for assessing swallow function. Additionally, the research explores the use of transcutaneous auricular vagus nerve stimulation (taVNS) as a neuromodulation technique to enhance oral feeding skills. The study found no adverse effects from taVNS on airway protection or swallowing physiology. Furthermore, there were some possible associations between specific white matter tracts and components of sucking and swallowing identified. Although these findings are preliminary and should be interpreted with caution. Larger studies are needed to confirm these results and to further explore the neurophysiological underpinnings of infant feeding and swallowing skills. The work provides a foundation for larger, future studies investigating the role of white matter tracts in infant feeding and swallowing
Role of Centrosomal P4.1 Associated Protein (CPAP) in Tumor Suppression
No full textIn eukaryotic cells, centrosomes are membrane-less organelles that function as the primary microtubules organizing center with a crucial role in cell division. For accurate bipolar spindle assembly and genetic transmission between generations, fidelity in centriolar duplication is critical. Disruption in this process can cause genomic instability, which could then lead to conditions like aneuploidy, a common feature of various cancers. Among the numerous proteins associated with mammalian centrioles, the centrosomal P4.1 associated protein (CPAP) stands out for its essential role in regulating centriole duplication. Dysfunctions in CPAP have been linked to abnormalities in centriole length that can contribute to conditions ranging from ciliopathies, immune dysfunction, and cancer.
Recent discoveries in our lab have shed light on CPAP\u27s involvement in endocytic vesicular transport and are investigating its implications in cancer control. Specifically, defective CPAP function in oral cancer cell lines has been associated with constitutive signaling through the epidermal growth factor receptor, heightened epithelial-mesenchymal transition (EMT), and enhanced spontaneous tumorigenic properties, suggesting a potential role for CPAP as a tumor suppressor protein.
Building on these findings, my research aimed to investigate the impact of CPAP dysfunction on tumorigenesis, by utilizing a hypomorphic mouse model. We evaluated the EMT features of both, primary and immortalized, lung as well as tongue-derived fibroblasts from these mice. In addition, we have also assessed the CPAP-deficient mice for their susceptibility towards spontaneous and induced tumors. Through my investigation, we seek to deepen our understanding of CPAP\u27s role as a tumor suppressor and its broader implications for cellular health
Readmission Rates of Mothers Experiencing Preeclampsia or Complications After Delivery in Underserved Populations
Get PDFHospital readmissions during pregnancy nationwide impacts maternal health outcomes elevating the risk of maternal deaths after discharge. The accessibility and affordability of necessary prenatal care is integral to the field of health across the lifespan. Socioeconomic barriers within rural locations place expecting mothers at an adverse risk of developing conditions including preeclampsia and other complications that may result in hospital readmission. Economic barriers including hospital closures, shortages of specialty healthcare providers, unstable internet access and, ease of care access on further contribute to this disparity among rural women.
A retrospective analysis was conducted using data from Health Care Cost and Utilization Project for the state of Florida in 2019, comparing rural versus non rural zip codes. This analysis was conducted to assess the rate of admission for mothers experiencing preeclampsia and pregnancy-related complications who lived in rural settings within the state. As Public Health Maternal and Child Health graduate and former employee of REACHUP, Incorporated, I have observed many barriers and socio-economic factors impacting rural maternal health. The provision of multifaceted support, emphasizing patient advocacy, and monitoring efforts to address medical concerns for women experiencing medical emergencies and additional barriers to care is necessary in addressing this disparity