MEDICA@MUSC (Medical University of South Carolina)
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Sphingosine Kinase 1 in Adipose Tissue: Roles in Metabolism, Adipogenesis, and Glucocorticoid Signaling
No full textSphingosine kinase 1 (SPHK1) has been identified as a regulator of key processes in many metabolic tissues, though its role in adipose tissue has not been well established. As SPHK1 has been shown to be elevated in obesity, we developed an adipocyte-specific knockout mouse (SK1fatKO) and conducted a high-fat diet study to investigate the role of SPHK1 in the adipocyte in the context of obesity. Alternative to the constitutive SPHK1 knockout mouse which is protected from obesity induced maladies, high-fat diet fed SK1fatKO mice were glucose intolerant, hyperinsulinemic, had liver steatosis and hypertrophic adipocytes. Interestingly, these mice were protected from adipose tissue inflammation. Additionally, RNA sequencing of the gonadal adipose tissue from SK1fatKO mice revealed decreased expression of many mature adipocyte markers. Paired with the adipocyte hypertrophy results, this indicated that SPHK1 depletion may interfere with adipogenesis. To better understand the mechanisms by which SPHK1 regulates adipogenesis, we utilized an in vitro approach using primary adipose derived stem cells. Under standard pro-adipogenic culture conditions, Sphk1 expression, along with sphingosine-1-phosphate (S1P) and SPHK activity levels, but not SPHK2 were determined to increase early in adipogenesis and decrease as the cells mature. Treatment with individual pro-adipogenic media components revealed that only dexamethasone, a synthetic glucocorticoid, induced Sphk1. This led us to investigate how SPHK1 is involved in glucocorticoid signaling in early adipogenesis. We found that expression of key genes in the adipogenesis pathway, C/EBPα and C/EBPδ were reduced in SPHK1-/- cells. Additionally, chronic treatment of SPHK1-/- mice with corticosterone in drinking water led to reduced adipose expansion compared to controls, along with altered adipogenic gene expression. These results highlight the importance of SPHK1 for adipose expansion and metabolism in the contexts of high-fat diet and glucocorticoid induced obesity
Launching a Library Career in a Fully Remote Work Environment
Get PDFObjective: This poster aims to outline one librarian\u27s experience obtaining their first professional role as a health sciences librarian during the COVID-19 pandemic.
Methods: Graduating in the middle of the COVID-19 pandemic in May 2020 provided unique challenges in the process of applying and beginning my first position as a health sciences librarian. I was applying for jobs out of state while living in Fort Lauderdale, FL as a new graduate from library school.
Results: I was hired for my first professional role as a Research and Education Informationist at the Medical University of South Carolina in October 2020 and began my position fully remote. The hiring process included a virtual interview, learning to build a rapport through Zoom, completing the onboarding process remotely, and attending an online orientation. The greatest challenges to overcome were building relationships with my team and learning about the university while living over 600 miles away. Team building in this virtual space included one-on-one meetings for introductions, virtual water-cooler sessions to facilitate conversation, and utilizing Microsoft Teams chat as a tool to keep lines of communication open. Learning about the university involved trial and error by shadowing chat shifts, reading through the library\u27s website, and being open to asking my team questions as they appeared.
Conclusions: Being hired as a librarian in a fully remote setting during the COVID-19 pandemic helped me strengthen my communication skills as I built relationships with my team solely through Microsoft Teams and e-mail communication. This experience pushed me out of my comfort zone by learning to collaborate with people I have never met in person before and increased my ability to adapt to rapidly changing situations
Bayesian Hierarchical Profile Regression for Categorical Data
Get PDFIn this research we consider problems involving discrete data which are divided into a set of hierarchical groups with each observation in each group believed to be drawn from a mixture model. Our goal is to recover the latent clustering structures for each data group, allow these discovered clusters to be shared among each of the data groups, and assess for associations between the estimated cluster memberships and a clinically relevant outcome. For the clustering model, we assume the number of mixture components within each group is unknown a priori and is to be inferred from the data. To analyze this type of data and accomplish our analytic goals we propose Bayesian Hierarchical Profile Regression (BHPR), a model which utilizes Bayesian Profile Regression (BPR) in conjunction with a Hierarchical Dirichlet Process Mixture Model (HDPMM). The utilization of the HDPMM allows for the discovery of latent clusters within each data group and allows for the identified latent clusters to be shared across the groups, while using the approach of BPR allows for the characterization of the associations between latent cluster memberships and the clinically relevant outcome. Data utilized to recapture cluster memberships within each data group can be either binary or ordinal. For binary data, one of the fundamental assumptions found in similar mixture models called Conditional Independence can either be maintained or relaxed. We demonstrate the utility of our model through applications to two patient populations: 1) a subset of data collected by the Acute Liver Failure Study Group, and 2) a cohort of patients referred for a Modified Barium Swallow Study assessed using the Modified Barium Swallow Impairment Profile. Results from these applications demonstrate that our research results can be used to inform appropriate intervention strategies and provide tools for clinicians to make better multidimensional management and treatment decisions in a variety of disease areas
RAGE Signaling as a Pharmacological Target in Amyotrophic Lateral Sclerosis
No full textAmyotrophic lateral sclerosis (ALS) is characterized by the progressive degeneration of both upper and lower motor neurons. Astrocytes are key for maintaining central nervous system homeostasis and are important determinants of motor neuron fate in ALS. Accordingly, IPSC-derived astrocytes from ALS patients or astrocytes from diverse ALS mouse models, including mice overexpressing the ALS-linked mutant hSOD1G93A, induce motor neuron death in co-culture. Previously our lab reported that motor neurons isolated from receptor for advanced glycation end products (RAGE) knockout mice are resistant to the ALS-astrocyte derived neurotoxicity. Therefore, our working hypothesis states that inhibition of RAGE signaling can suppress astrocyte-mediated neurotoxicity and has potential translational value to prevent or delay ALS disease progression. We confirmed that in a co-culture model, the motor neuron death induced by ALS-astrocytes is prevented by RAGE pharmacological inhibition (FPS-ZM1 or RAP). RAGE inhibition also prevented the motor neuron death induced by lumbar spinal cord extracts from symptomatic hSOD1G93A mice. To confirm the relevance of this neurotoxic mechanism in ALS pathology, we evaluated the therapeutic potential of FPS-ZM1 in vivo in hSOD1G93A mice. FPS-ZM1 treatment significantly improved hind-limb grip strength of hSOD1G93A mice during the progression of the disease. This was associated with improved survival of large motor neurons, reduced gliosis, and decreased ER stress in the spinal cord. Moreover, FPS-ZM1 reduced the expression of atrophy markers in the gastrocnemius muscle. However, RAGE inhibition did not alter the onset of the disease nor the survival of hSOD1G93A mice significantly. The maintenance of hind-limb grip strength was also observed in hSOD1G93A mice with genetic RAGE haploinsufficiency, further indicating the beneficial effect of RAGE inhibition on motor function. However, this beneficial effect was not observed in mice with complete RAGE ablation. Moreover, the partial or complete RAGE ablation drastically shortened the median survival of hSOD1G93A mice by 14 and 18 days, respectively. These results reveal a complex effect of RAGE inhibition in hSOD1G93A mice. Together, our data indicate that the development of therapies targeting RAGE in ALS require better understanding of its role, in a cell type- and stage-specific manner, during the progression of the disease
Targeting Transcription Factors XBP-1 and Fli-1 as Novel Translational Strategies to Control Graft-Versus-Host Disease
Get PDFHematopoietic stem-cell transplantation (HCT) is a curative procedure for hematological malignancies, but chronic graft-versus-host disease (GVHD) remains a major complication after allogeneic HCT. Because donor B cells are essential for chronic GVHD (cGVHD) development and B cells are sensitive to endoplasmic reticulum stress, we hypothesized that the IRE-1α/XBP-1 pathway is required for B-cell activation and function in cGVHD. Here, we used mice deficient of XBP-1 specifically in B cells, and recipients transplanted with grafts containing XBP-1–deficient B cells displayed reduced cGVHD compared with controls that was associated with reduced B-cell activation and production of alloreactive antibodies. Prophylactic administration of B-I09 (an IRE-1α/XBP-1 inhibitor) also reduced cGVHD without compromising GVL effect against chronic myelogenous leukemia. Although B cells play an important role in chronic GVHD development, effector T cells are still primary drivers of the disease. We investigated the T-cell specific role of a second transcription factor, Fli-1, in GVHD pathogenesis. Ablation of two critical exons of the fli-1 gene in donor-derived T cells was associated with significant reduction of disease development in allo-HCT models of cGVHD and aGVHD. This reduction was associated with increased regulatory T cells (Tregs) and decreased IFN-γ+, IL-17A+, and TFH T cells in lymphoid organs. We also demonstrated that low-dose camptothecin exhibits action as a potent Fli-1 inhibitor, both in vitro and in vivo against mouse and human Fli-1. Utilization of drugs that targeted Fli-1 was able to prevent cGVHD development and reverse already established cGVHD. Camptothecin also prevented aGVHD while preserving the GVL effect against P815 mastocytoma. Further, camptothecin reduced human T-cell proliferation both in vitro and in vivo, while also reducing GVHD in a human xenograft model. These findings in combination suggest that XBP-1 is a critical factor regulating the B-cell component of chronic GVHD development while Fli-1 is a critical factor involved in the T-cell component. Many transcription factors are also notoriously difficult to inhibit, however, we identified that both XBP-1 and Fli-1 could be targeted using pharmaceuticals, making these factors promising translational strategies for reducing GVHD in the clinical setting, while also preserving the GVL effect of the donor graft
The Impact of the ACA Medicaid Expansion on Access to Care and Hospitalization Charges for Lupus Patients
No full texthttps://medica-musc.researchcommons.org/posters/1025/thumbnail.jp
A Descriptive Case Study on Patient Flow at a Medical Center in a Resource-Limited Setting
Get PDFMedical centers operating in resource-limited settings face unique challenges associated with a scarcity of resources, expertise, and infrastructure. While inefficiencies can be identified in any organization, they can be particularly problematic for health care facilities that seek to expand the scope of their operations in resource-limited areas as resources that are already in short supply are utilized to fuel non-value-added activities. In this descriptive case study of a newly built medical center in Khorog, Tajikistan, we consider a medical center’s patient flow and the underlying systemic factors that contribute to the current state of affairs as well as interventions that could be employed to make more financial capital, human capital, and governmental support available to address the root causes of systemic challenges. In addition to exploring how countries like Tajikistan can thrive given decades of unrest, we explore how resource-limited areas can position themselves to capitalize on emerging trends in the global marketplace
Inhibition of Penicillin-Binding Protein 2: Toward New Therapeutics for Antimicrobial-Resistant Gonorrhea
Get PDFGonorrhea is the second most common sexually transmitted bacterial infection in the United States, with nearly 600,000 cases reported in 2018 by the Centers for Disease Control. Alarmingly, the causative agent Neisseria gonorrhoeae has developed resistance to a number of antimicrobials over the last century. With limited options remaining, the CDC now recommends dual therapy with ceftriaxone and azithromycin to decrease the likelihood of resistance development. However, strains with combined cephalosporin and macrolide resistance have now emerged, raising concerns of a post-antibiotic future in which untreatable gonorrhea would impose enormous human and economic cost. The discovery and development of novel antigonococcal agents is, therefore, necessary to avoid a public health crisis. The pharmacologic receptors for β-lactams are a group of transpeptidases known as penicillin-binding proteins (PBP), which catalyze the cross-linkage of peptidoglycan, an essential component of the bacterial cell wall that plays major roles in cell growth and division. N. gonorrhoeae develops chromosomally mediated β-lactam resistance via alterations of PBPs affecting drug affinity, specifically through the acquisition of mutations in the penA gene encoding PBP2. Resistant strains harbor mosaic penA alleles encoding PBP2 variants containing around 60 amino acid changes compared to wild-type. In this work, we examine inhibition of a mosaic form of N. gonorrhoeae PBP2 from the cephalosporin-resistant strain H041, seeking to understand better which features of ligand structure enhance or diminish PBP2 binding in order to develop more effective PBP2 inhibitors. First, we report structure-activity relationships (SAR) for the cephalosporin class of β-lactams against PBP2 from N. gonorrhoeae H041 with the goal of identifying or designing cephalosporins effective against resistant strains. We find that structural features of the C7 acylamino side chain (R1) correlate highly with the second-order rate of PBP2H041 acylation, including increased size, modest lipophilicity, and two ring systems separated by a single branch point. The C3 side chain (R2) makes lesser, but still important, contributions to inhibition, with electronegative elements and planarity enhancing activity. We also found that many of the features enhancing target inhibition (e.g., lipophilicity, aromaticity) diminish antimicrobial activity against the H041 strain, perhaps due to decreased accumulation in the periplasm. Finally, we identify cefoperazone as highly active against PBP2H041 and similarly active against N. gonorrhoeae H041 both in vitro and in vivo compared to ceftriaxone. Second, we report the in silico discovery of novel noncovalent PBP2 inhibitors possessing a 1,1’-biphenyl system. Arylamide JEK-42 and its isosteric sulfonamide derivative JMT-1 are capable of inhibiting PBP2 from both β-lactam-susceptible and -resistant gonococcal strains. Their cross-inhibition of P. aeruginosa PBP3, predicted binding modes showing interaction with highly conserved residues, and structural similarities to bicyclic β-lactam scaffolds indicate their potential for broader activity against class B PBPs. Using the structural similarities between JEK-42, JMT-1, and bicyclic β-lactam scaffolds (i.e., penam, carbapenem, and cephem), a three-point pharmacophore was generated that can be used to identify additional PBP-inhibitory scaffolds. Third, we report the synthesis of 127 derivatives of JMT-1, showing specific substitutions that enhance the inhibition of PBP2 derived from both β-lactam-susceptible and -resistant strains. In keeping with the cephalosporin SAR, hydrophobic substitutions enhance PBP2 inhibition, likely through increased van der Waals contact with the active site, but they can also result in diminished antimicrobial activity. Together, our efforts yielded 10 compounds that show near full inhibition of PBP2 from susceptible and resistant strains, as well as large zones of gonococcal growth inhibition in disc diffusion assays. These studies lay the groundwork for the development of several structurally diverse antigonococcal chemotypes, thereby increasing the probability of producing a successful preclinical candidate
Contributions of Aberrant Inflammatory and Immune Cell Activity in the Cochlea to Age-Related Hearing Loss
No full textAge-related hearing loss (ARHL), or presbyacusis, is a prominent chronic degenerative disorder affecting older individuals. Characterization of ARHL pathology reveals degeneration of both sensory and non-sensory cells, as well as alteration of the cochlear microenvironment. It is unclear when and to what extent the tissue homeostasis of the cochlea is perturbed during aging. The progression of age-related neurodegenerative diseases is associated with an altered microenvironment reflective of chronic inflammatory signaling. Under these conditions, resident and recruited immune cells, such as microglia and macrophages, demonstrate aberrant cellular activity that may contribute to neurodegeneration. Two key elements of the innate immune response are effector immune cells, which include macrophages, and pathways of regulation, such as the complement system. Complement molecules are key mediators of the innate immune system, and dysregulation of their expression contributes to the progression of several neurodegenerative diseases. In the inner ear, macrophages have been identified to populate in various compartments of the cochlea, with a demonstrated role in auditory nerve (AN) refinement during development. Furthermore, previous studies have documented the cochlear macrophage response to noise-induced injury and sensory cell degeneration in animal models. However, our understanding of how an altered cochlear microenvironment may impact or result from altered resident immune cell activity is limited. This dissertation investigates if the aging inner ear demonstrates enhanced inflammatory signaling and aberrant macrophage cellular activity, thereby contributing to ARHL pathology. Examination of age-related alterations in the cochlear lateral wall (CLW) and AN was performed in both human and mouse specimens. The finding of morphological distinction of macrophages in the CLW and AN may be reflective of region-specific cellular functions. Additionally, macrophage number and morphological alterations consistent with activation were shown to increase with age in the AN. Evidence of enhanced inflammatory signaling in the aged cochlea was identified via the transcriptional profiles of the AN and CLW. Significant enrichment of the complement signaling pathway was identified in the aging AN and CLW. To further investigate the origin of complement signaling molecules in the cochlea, we performed transcriptomic analysis on the samples enriched for cochlear macrophages compared to the samples obtained from non-macrophage cochlear cells. Using this novel approach, we identified expression of central complement molecule (C3) in both macrophages and non-macrophage cochlear cells. Further, cell surface receptors that are documented to mediate macrophage/microglia recruitment and activation, such as C3ar1, C5ar1, Itgam, and Itgb2, were found to be expressed by cochlear macrophages. We next tested the hypothesis that reduction of complement activation has a protective effect on age-related hearing loss. Our data suggests that deficiency of C3 reduced the incidence of cochlear macrophage activation and recruitment in the aged AN and CLW. Although we did not observe improved hearing thresholds, quantitative analysis indicates protection for spiral ganglion sub- types when comparing C3-deficient mice relative to controls. Together, our investigations indicate over-expression of central complement molecule C3 in the aging cochlea contributes to changes in macrophage activity that are associated with pathophysiological alterations of the aged cochlea
Determination of N-Linked Glycosylation Changes in Hepatocellular Carcinoma and the Associated Glycoproteins for Enhanced Biomarker Discovery and Therapeutic Targets
Get PDFWith hepatocellular carcinoma (HCC) remaining as the fifth most common cancer in the world, causing more than 700,000 deaths annually, the need for reliable, early stage diagnoses and preventive treatments is crucial. While serum glycoproteins are hepatic in origin, making them excellent targets for HCC biomarkers, they can originate from both cancerous and non-cancerous regions and direct analysis of cancerous tissue itself is lacking. To counteract this, I hypothesized that direct tissue analysis combined with proteomic analysis could be utilized to identify more potential targets specific to HCC for early detection. This was done with a primary focus on glycosylation—as most clinically approved biomarkers are glycoproteins—and examined direct tissue glycomics in conjunction with glycoproteomic techniques through two specific aims: 1) Determining patterns of N-linked glycan changes in HCC tissue using MALDI imaging mass spectrometry to compare to previously published serum changes and 2) identifying glycopeptides containing changes in observed patterns of N- linked glycans in HCC samples using a targeted glycoproteomic approach. In Aim 1, HCC tissue was examined using MALDI imaging mass spectrometry to v verify changes in glycosylation via direct tissue analysis. Here, it was found that increased branching and fucosylation were directly associated with the cancerous tissue when compared to normal or cirrhotic. To further identify changes in glycosylation, two methods (one novel and one adapted for imaging) were implemented on tissue to further classify N-linked glycan isoforms through linkage analysis, specifically for sialic acids and core fucose. Again, it was shown that core fucose is most directly related to HCC tissue, thus confirming serum findings in the literature. For Aim 2, the novel method of determining core fucosylation was used in conjunction with glycoproteomic techniques to further elucidate the core fucosylated glycoproteins of interest. With the tag left behind following the enzymatic cleavage, targeted glycoproteomics was used to determine glycoproteins of interest while eliminating some biases inherent in the method, such as low ionization efficiencies for more complex N-glycans. This work outlines the first in-depth analysis of HCC tissue specifically regarding N- glycan changes, a novel application to determine N-glycan isoforms, and the application of these methods for glycoproteomic enhancement. With these findings, new trends in glycosylation related to the disease state could be further uncovered, as well as provide new biomarker candidates or therapeutic targets for future studies