• Examining Behavioral Flexibility in Pavlovian Processes through the lens of Endocannabinoid Regulation and Sex Differences

  Stapf, Catherine; Calu, Donna (2024)

  Behavioral flexibility is an adaptive process where one alters behavior due to a change in the internal or external environment. Prior research in the field shows that both the dorsolateral striatum (DLS) and dorsomedial striatum (DMS) contribute to the regulation of behavioral flexibility in instrumental conditioning. I begin in Chapter 1 by presenting background literature on behavioral flexibility, comparing instrumental and Pavlovian paradigms, and considering sex differences in either context. In Chapter 2, I focus on how male and female Long Evans rats differ in the expression of a Pavlovian conditioned response and sensitivity to outcome devaluation and how DMS cannabinoid-1 receptors (CB1Rs) mediate these processes. I find that females show more lever-directed behavior and are insensitive to outcome devaluation. I also find that DMS CB1Rs are required for the devaluation sensitivity of males. In Chapter 3, I use slice electrophysiology to assess whether DMS inhibitory tone at baseline differs between males and females and how CB1R activation affects inhibitory synaptic transmission. I find that cells recorded from male rats show reduced frequency of inhibitory events and that CB1R activation reduces inhibitory tone in both males and females. In Chapter 4, I investigate the relationship between social reward and sign-tracking. I find that sign-tracking behavior persists under extinction and in the presence of an alternative reward choice. Collectively, this research establishes a role for DMS CB1R in Pavlovian outcome devaluation and supports previously established sex differences in behavioral flexibility.
• The CCCH-type Zinc Finger Tristetraprolin and Post-translational Modification by Metals and Hydrogen Sulfide: Implications on Zinc Finger Function during Inflammation

  Stoltzfus, Andrew; Michel, Sarah L.J. (2024)

  Zn(II) is an essential transition metal that cofactors ~10% of the human proteome to play key roles in regulating immune function and inflammation. A large proportion of this Zn(II) proteome are Zinc Finger (ZF)-containing proteins which coordinate Zn(II) with 4 Cys/His ligands to achieve secondary structure and function. ZFs are, however, subject to adventitious metal binding and post-translational modification (PTM), both of which may affect protein structure and function. The nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB) pathway is a focal point for inflammation and immunity and is regulated by numerous ZFs. Tristetraprolin (TTP) is one such ZF that negatively regulates the NFκB pathway by degrading cytokine mRNAs. I first set out to understand TTP’s interactions with the toxic metal, lead (Pb(II)), by expressing and purifying an apo-peptide construct of TTP’s two CCCH ZF domains (TTP-2D). I utilized an array of optical spectroscopies which revealed that Pb(II) binds apo-TTP-2D with an affinity comparable to Zn(II) and perturbs TTP-2D’s structure. Furthermore, Pb2-TTP-2D had a higher affinity for a biologically relevant mRNA target, suggesting a change in protein function with the non-native metal. To elucidate ZF persulfidation, a PTM of Cys-SH to Cys-SSH, I used TTP-2D as a template for amino acid mutation and testing of various ZF domains for their reactivity with hydrogen sulfide (H2S), the endogenously produced signaling molecule associated with persulfidation. I designed a series of mutants of TTP’s first ZF domain (TTP-1D) to include all iterations of Cys ligands for binding (i.e. CHHH, CCHH, and CCCC), representing other biological ZF domains. I found that all domains reacted with H2S, showing a minimum requirement of one Cys residue for persulfidation. Finally, I am connecting ZF persulfidation with the inflammatory response of the NFκB pathway during TNFα stimulation in ongoing cell studies. To discern the requirements for H2S and persulfidation in the inflammatory response, we have validated a knockout cell line which significantly reduces persulfdation. We have prepared chemoselective proteomics samples which will identify ZF-persulfides during inflammation to compare against the resting state. Continued studies will investigate the functional impact of persulfidation on novel ZF-persulfides identified from these data.
• MYBPC1 E248K Myotrem: an investigation into disease progression and pathogenic mechanisms

  Mariano, Jennifer; Kontrogianni-Konstantopoulos, Aikaterini (2024)

  Myosin binding protein-C (MyBP-C) comprises a family of muscle accessory proteins that function in organizing and maintaining sarcomeric structure and regulating contractile function and actomyosin crossbridge cycling. Mutations in MYBPC1, the gene encoding the slow skeletal isoform (sMyBP-C), lead to a novel dominant form of myopathy termed Myotrem (OMIM: 318524) characterized by muscle weakness, congenital hypotonia, skeletal deformities, and characteristic tremor affecting the extremities and tongue. Due to the restricted expression of MYBPC1 to skeletal muscle, as well as the absence of neuropathy in these patients, this tremor is believed to be a novel entity originating within the muscle itself. To further investigate Myotrem, our group has developed a murine model harboring a knock-in (KI) MYBPC1 E248K mutation that faithfully recapitulates disease phenotype. Subsequent studies in KI muscle revealed that structural alterations of the sarcomere underlie the myopathy and are integral to pathogenesis. In human patients, Myotrem manifestations progress through childhood until adolescence, at which the phenotype stabilizes with no further deterioration. However, the clinical manifestation of Myotrem after mid-adulthood in response to the biological processes of aging is not known. In these studies, we used the murine KI model to perform in vivo contractile force measurements and quantitative structural analysis of immunofluorescence-stained muscle to extensively characterize muscle structure and function in mid- and late-adulthood. We show that this stabilization period expires in late adulthood as KI mice show a decline in muscle structure and function in a sex- and muscle-specific manner where males and larger muscle muscles are more affected. Furthermore, although Myotrem tremorgenesis is considered to originate intrinsically in the muscle, the molecular mechanism is largely unknown. Notably, we found that permeabilized soleus fibers from KI mice exhibited unsolicited pulse activity with unprecedented regularity and persistence. Immunoblot and mechanical step-stretch experiments reveal that these pulsing KI fibers are slow-twitch in nature and display enhanced stretch activation properties. We posit that these pulsing fibers function as a myogenic pacemaker for tremor. Collectively, these studies provide essential information to inform patients afflicted with Myotrem of disease development and provide putative mechanisms for pathologic myogenic tremorgenesis.
• The role of Penicillin-Binding Protein-4 (PBP4) in β-lactam resistance of Staphylococcus aureus

  Satishkumar, Nidhi; Chatterjee, Som (2024)

  The Gram-positive bacterium Staphylococcus aureus, a frequent colonizer of humans, can also cause infections such as bacteremia, osteomyelitis or sepsis. Along with its pathogenicity, S. aureus also has the ability to be resistant to antibiotics such as β-lactams, making it one of the leading causes of global mortality. Classically, β-lactam resistance is attributed to mecA (or mecC), as seen in methicillin resistant S. aureus (MRSA). Prior studies identified PBP4 (Penicillin Binding Protein-4), a protein involved in cell wall synthesis, as a non-classical mediator of resistance. Mutations associated with the regulatory region of the gene (Ppbp4) led to increased expression of PBP4 that subsequently caused increased cell wall cross-linking, while missense mutations in the pbp4 gene led to structural alterations, both of which resulted in β-lactam resistance. Using techniques in microbiology and molecular genetics, this study substantiated the potential of PBP4 as a novel mediator of β-lactam resistance by first demonstrating the clinical relevance of pbp4-associated mutations and their corresponding phenotypic alterations. The Ppbp4 region is shared with a divergently transcribed gene, abcA, which encodes a multidrug exporter. This study confirmed that Ppbp4 mutations caused an upregulation of pbp4 and downregulation of abcA, confirming that the resistant phenotype was attributed to PBP4 overexpression. In prior experiments, pbp4-associated mutations in resistant strains were largely accompanied by loss-of-function mutations of gdpP, a phosphodiesterase. This study demonstrated that alterations of PBP4 and GdpP led to synergistic, broad-spectrum resistance to β-lactams at levels comparable to that seen by MRSA. Lastly, the effect of PBP4 on bacterial virulence was inspected using C. elegans, the results of which indicated that β-lactam resistance mediated via PBP4-overexpression likely came at the cost of virulence. This study emphasizes the relevance of pbp4-associated mutations as a clinically significant cause of β-lactam resistance, not only by itself but also in synergy with altered GdpP. The experimental findings put forth the suggestion of considering PBP4 as an important target not only for development of therapeutics, but also during diagnosis of S. aureus infections.
• The Association Between Pre-Hip Fracture Depression and Antidepressant Use and Post-Fracture Days at Home, Length of Hospital, and Mortality in Medicare Beneficiaries and Related Sex Differences

  Mehta, Rhea; Orwig, Denise L.; Falvey, Jason (2024)

  Hip fracture and depression are significant public health issues for older adults, and antidepressants are the primary treatment for depression in this population. While it is known that post-hip fracture depression impedes hip fracture recovery, and antidepressants are widely used among older adults, no studies have assessed the influence of pre-fracture depression and pre-fracture antidepressant use on hip fracture outcomes among older adults, or related sex differences. Thus, Aim 1 assessed the association between pre-fracture depression and days at home (DAH) after fracture, and related sex differences; Aim 2 examined pre-fracture antidepressant use and hospital length of stay (LOS); and Aim 3 explored the association between pre-fracture antidepressant use and mortality among hip fracture survivors with depression, and related sex differences. Medicare fee-for-service claims data was used and included community-dwelling beneficiaries with a hospitalization claim for hip fracture surgery between 2010 and 2017. Poisson regression models assessed the association between pre-fracture depression and days at home (DAH) (Aim 1), while ordinal logistic regression estimated the association between pre-fracture antidepressant use hospital LOS (Aim 2), and Cox proportional hazards models evaluated pre-fracture antidepressant use and mortality (Aim 3). A sex-by-exposure interaction term was added to adjusted models to examine sex differences. Beneficiaries with depression and prior antidepressant use were 79 years of age, on average, and most were White females. Those with depression spent 11 fewer average DAH compared to counterparts without depression (incidence rate ratio [IRR]=0.92; 95% CI=0.91, 0.93; p<.0001), but this associated attenuated after adjusting for comorbidities. Beneficiaries with depression who used antidepressants before fracture had 8% higher odds of a shorter hospital LOS compared to non-users (odds ratio [OR]= 1.08; 95% CI=1.02, 1.14; p=0.01), and had a lower risk of 90-day (hazard ratio [HR]=0.84, 95% CI=0.76, 0.93, p=0.001) mortality compared to antidepressant non-users. No significant differences by sex were found in any analyses. Results demonstrate the importance of recognizing the intersection of mental and physical health in post-hip fracture care, and considering patients’ entire, individual health profiles when making treatment decisions after fracture among older hip fracture survivors with depression.
• Vestibulotoxicity in Pediatric Oncology: Screening, Assessment, and Relationships with Audiometric and Balance Function

  McCarthy, Emily; Marchese, Victoria G. (2024)

  Background: Balance is an important gross motor function which allows children to safely participate in recreational and sport activities. Balance requires integration of sensory input from three main systems, the somatosensory system, visual system, and vestibular system. Childhood cancers and the medical treatments used in the management of these diseases can cause toxicity or damage to any one of these systems. Furthermore, children with a diagnosis of cancer are known to experience balance deficits both during and after cancer treatment. There is limited knowledge, however, of toxicity to the vestibular system specifically, in children with a diagnosis of cancer. In addition, relationships between vestibular function, audiometric function, and balance performance in this population have not been well established. Methods: Vestibular screening failures (Pediatric Vestibular Symptom Questionnaire, Modified Clinical Test of Sensory Interaction on Balance, dynamic visual acuity), vestibular function (video head impulse test, cervical and ocular vestibular evoked myogenic potentials), audiometric function (tympanometry, audiogram), balance performance (Bruininks-Oseretsky Test of Motor Proficiency, 2nd Edition, Balance subtest), and sensory integration (Modified Clinical Test of Sensory Interaction on Balance) were measured in children with a diagnosis of cancer and non-cancer age-matched controls. Results: Children with a diagnosis of cancer presented with a greater prevalence of vestibular screening failures as compared to non-cancer age-matched controls. Children with cancer had a greater prevalence of vestibular dysfunction as compared to controls, and the severity of vestibular loss was also greater in this group. Relationships between vestibular dysfunction and hearing loss were not identified. Children with a diagnosis of cancer had impaired balance performance and sensory integration as compared to non-cancer age-matched controls, and significant relationships between vestibular loss severity and balance performance were identified in children with a diagnosis of cancer. Conclusions: Children with a diagnosis of cancer demonstrate vestibular dysfunction that is related to reduced balance performance and impaired sensory integration.
• Precision in vivo genome editing for the study of rare genetic disease variants

  Robertson, Colin; Poulopoulos, Alexandros (2024)

  The medical community is on the cusp of revolutionary curative treatments through therapeutic genome editing. However, technical limitations, such as unintentional and unpredictable gene silencing, along with lengthy development and regulatory approval processes severely restrict the scope of these therapies. Recent advancements that we and others have made in the field of precision genome editing offer potential solutions for overcoming barriers to translating genome editors into genome therapeutics. In this work I demonstrate novel applications of precision genome editing tools that could provide more immediate benefit to ailing populations. I explore multiple gene editing modalities and describe a flexible approach for rapidly producing gene editors as ribonucleoproteins. Using prime editing, I introduce epilepsy variants from individual patients into wild type mice. We validate the precision of in vivo genome editing using both Next-Generation Sequencing and electrophysiology. Notably, a portion of wild type mice edited with these ultra-rare epilepsy variants display spontaneous motor seizures, reproducing a key clinical symptom of the modeled patient. I further demonstrate how these mice could be used as surrogate models to test pharmacotherapies in the context of an individual’s specific genetic variants, providing direct evidence for new applications of prime editing in precision medicine. This approach is robust across multiple genomic loci and produces reliable models of genetic epilepsies much faster than traditional disease modeling. Overall, these results are among the first to show successful application of prime editing in the brain, among the first to demonstrate the production of prime editing ribonucleoproteins, and demonstrates a novel application of these technologies that can directly benefit patients with genetic epilepsies.
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  Determining Intercondylar Distance Using Cone-beam Computed Tomography (CBCT)

  Alqabandei, Omar M; Masri, Radi RM (2024)

  Purpose: To measure the intercondylar distance (ICD) from cone-beam computed tomography (CBCT) scans of a large population of subjects to determine the average ICD in male and female subject and to correlate ICD measurement obtained with sex and ethnicity of the subjects. Materials and Methods: This cross-sectional study analyzed consecutive patients who had received maxillofacial field of view CBCT radiographic examinations at the University of Maryland School of Dentistry (UMSOD) between January 20th, 2016, and July 5th, 2023. Inclusion criteria comprised individuals over 18 years old who had maxillofacial CBCT scans on file in the UMSOD INFINITT (INFINITT NA, Phillipsburg, NJ) PACS system. Exclusion criteria included patients with prosthetic condyles, bone growth-affecting diseases, incomplete scans, poor image quality, or missing sex and age information. Of the initially included 459 patients, 25 were excluded. Data collected from patient charts included age, sex, presence of bone growth-affecting diseases, and measurements of left and right condyles, as well as ICD. The primary investigator reviewed scans, conducted chart reviews, and made measurements on the 434 included files. Results: Four hundred and thirty-four images of subjects were analyzed. The sample consisted of an even number of male and female subjects (217). The median ICD value was 102.9 mm (min 86.2 mm – max 118.2 mm) for male subjects and 98.4 mm (min 81.5 – max 117.2) mm for female subjects. The mean ICD value of male and female subjects combined (total 434 subjects) was 100.92 mm (min 81.5 mm – max 118.2 mm) and the median value was 100.5 mm. There was statistically significant difference between ICD values of male and female subjects (P<0.0001), but not between Caucasian and African American subjects (P=0.69). There was no significant difference in the interaction between sex and ethnicity (P=0.84). Conclusions: The ICD influences the radius of movement and the arcs traveled by the cusps during lateral mandibular movements in the horizontal plane. Although canine disocclusion can mitigate inaccuracies arising from an average ICD, it may not fully compensate for individual variations in all patients. For patients restored with group function occlusion, ICD setting may be more critical. Articulators with an adjustable ICD would provide a more anatomically correct tooth form for the treatment of full mouth rehabilitation cases.
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  The metabolic and peroxisomal role of ubiquitin E3 ligase MARCH5 in a mitochondria-reliant bioenergetic model

  Verhoeven, Nicolas; Karbowski, Mariusz (2024)

  Peroxisome biogenesis requires up-to-now unidentified mitochondrial proteins. We show that the outer mitochondrial membrane (OMM) associated E3 Ub ligase MARCH5 is critical for early steps of generating mitochondria-derived peroxisomes. MARCH5 deficiency leads to an accumulation of immature peroxisomes, as well as lower expression of several peroxisomal proteins. When peroxisome biogenesis is induced through exposure of cells to fatty acids, MARCH5 is found in newly formed peroxisomes; however, peroxisome biogenesis is lost in MARCH5 deficient cells. Additionally, cells deficient in PEX3 and PEX14 peroxisome biogenesis factors exhibit impaired ability to generate peroxisomes and fail to indicate peroxisome biogenesis rescue if MARCH5 is simultaneously knocked out. WT MARCH5 re-expression pre-peroxisome formation in PEX14/MARCH5 DKO cells. PEX3 re-expression only reintroduces peroxisome formation in PEX3 KO, but not PEX3/MARCH5 DKO cells. Furthermore, our data also show reduced cellular ATP in a mitochondria-reliant energy generation model in MARCH5 KO cells. Our data suggest MARCH5 is essential for mitochondria dependent peroxisome biogenesis and may play a direct role in cellular energetics when cells are reliant on β-oxidation for energy generation.
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  The Role of Relationships for Adults Who Served Long Prison Sentences that Began in Childhood: A Mixed Methods Approach

  Franke, Nancy; Greif, Geoffrey L.; Bright, Charlotte Lyn (2023)

  The US is the only country in the world to sentence children to die in prison. As of 2016, there were over 12,000 people in US prisons who had, starting between the ages of 12 and 17, been sentenced to spend the rest of their lives behind bars. Several Supreme Court rulings have opened the door for reconsideration of some of these sentences, resulting in the release of more than 980 “juvenile lifers.” Trauma histories, depleted social networks, diminished well-being and complex barriers to reentry are well-documented realities for people leaving prison after long sentences that began in adulthood. However, the post-release lives of those who served life and long sentences that began during childhood remain almost completely unexplored. This dissertation study, believed to be the first national study of returned juvenile lifers, used mixed methods with a social network component to explore the post-release lives of this population. Findings are reported from a quantitative sample of 78 juvenile lifers from 24 US states and Washington, DC, with social network analytic tools considering the type, quality, and social support of the sample’s 555 social network members. Data from in-depth semi-structured interviews with 46 of the 78 participants were compared and integrated into the quantitative data. Quantitative analyses included multivariate regression and multi-level modeling, while qualitative analyses used thematic analysis. Respondents report high quality of life, overall positive relationships, high amounts of social support, and very little undermining. In a multiple regression model, attending religious services, spending more than 20 years in prison, and having fewer unmet reentry needs were all associated with higher well-being. Themes were developed regarding how respondents navigated different types of relationships (with family members, romantic partners, friends), life in prison, barriers to reentry, and parole, as well as their well-being. Positive relationship assessments, supportive relationships, number of types of support, and proportion of friends in a network were all associated with higher quality of life scores in social networks. Social network age, race, employment, and incarceration history were also associated with altered levels of well-being. Research, policy, and practice implications are discussed.
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  A Cadaver-based Comparison of Sleeve-guided Implant-drill and Dynamic Navigation Osteotomy and Root-end Resections

  Rollor, Corey; Martinho, Frederico C. (2024)

  The purpose of this study was to compare the accuracy and efficiency of fully guided static and dynamic computer-assisted surgical navigation methods for osteotomy and root-end resection (RER). Fifty root specimens from cadaver heads were divided into two groups: fully guided static computer-assisted endodontic microsurgery (FG sCAEMS) and dynamic computer-assisted endodontic microsurgery (dCAEMS) (n=25). Cone-beam computed tomography (CBCT) scans were conducted before and after the surgery. Osteotomy and RER were virtually planned in the preoperative CBCT scan. Endodontic microsurgery was executed with the guidance of 3D-printed surgical guides in FG sCAEMS and a 3D-dynamic navigation system in dCAEMS. Analysis involved calculating 2D and 3D deviations as well as angular deflection. Additionally, osteotomy volume, resected root length, and recording osteotomy and RER time were measured. The number of procedural errors were recorded. FG sCAEMS demonstrated comparable accuracy to dCAEMS, with no significant difference in 2D and 3D deviation values or angular deflection (P > .05). FG sCAEMS led to reduced osteotomy and RER times (P < .05). However, FG sCAEMS exhibited a higher incidence of incomplete RERs compared to dCAEMS. Osteotomy volume, RER angle, and resected root length were similar in both groups (P > .05). Overall, both FG sCAEMS and dCAEMS proved to be viable options for performing osteotomy and RER. Within the constraints of this study based on cadaver samples, FG sCAEMS demonstrated comparable accuracy to dCAEMS and both methods were efficient for performing osteotomy and RER.
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  Navigating the Unprecedented: A Study on How Senior Administrators in Higher Education Learned and Made Decisions During the COVID-19 Pandemic

  Edwards, Hillary; Kulo, Violet A. (2024)

  The purpose of this qualitative multi-case research study was to explore how senior administrators at four higher education institutions learned and made decisions during the first two years of the COVID-19 pandemic. Bandura’s social cognitive theory guided this study as it addressed how people learn enactively and vicariously as well as the triadic reciprocal causation between environmental, personal, and behavioral factors. The study included 13 senior administrators and data were collected through semi-structured interviews. The data analysis followed Creswell and Guetterman’s six-step process for analyzing and interpreting qualitative data. Data analysis involved both inductive and deductive coding and categorizing codes into sub-themes and themes. Trustworthiness was ensured through researcher reflexivity, peer debriefing, member checking, and thick description. All participants engaged in some form of observational learning, predominantly from within their own institution or across the university system. Perceived self-efficacy had a significant influence on leadership learning and decision-making. Participants shared their values of teamwork and collaboration and consistent communication in service to keeping their communities safe and continuing their institutional educational missions. The results of this study demonstrated the importance of observational learning on how leaders learned during the COVID-19 crisis, as well as the importance of self-efficacy including coping efficacy and its influence on a leader’s confidence, flexibility, and resolution during times of significant and rapid change. Practical implications include recommendations for crisis management planning, teambuilding, and communications strategies to improve academic leadership self-efficacy.
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  Genotypic and Phenotypic Characterization of Shigella flexneri serotype 6.

  Gabor, Caitlin; Barry, Eileen M.; Rasko, David A. (2024)

  Shigella is among the leading bacterial pathogens responsible for diarrheal disease causing > 200,000 deaths per year. Shigella flexneri, one of four Shigella species with 15 defined serotypes/subgroups, is the predominant Shigella species recovered from pediatric shigellosis in low- and middle- income countries (LMICs). Despite significant public health efforts, Shigella continues to be a major health concern. Shigella vaccine development has been hindered largely by the antigenic and genomic diversity. A quadrivalent Shigella vaccine including S. flexneri 2a (Sf2a), S. flexneri 3a (Sf3a), S. flexneri 6 (Sf6), and S. sonnei would provide 84.7% coverage across LMICS, providing protection for children in LMICs as well as travelers and military personnel. Of these three S. flexneri serotypes, Sf6 is the most phylogenomically and phenotypically distinct compared to other S. flexneri serotypes. We hypothesize that the distinct genomic content of Sf6 confers serotype-specific host-pathogen interactions. Preliminary results comparing historical clinical (archetype strains) Sf2a strain 2457T, Sf3a strain J17B, and Sf6 strain CCH060 demonstrated that CCH060 contains the greatest amount of unique genomic content, as well as lacking several previously described Shigella virulence factors. Phenotypic analysis revealed that there is variation in the expression and production of the virulence Ipa proteins and reduced invasion by CCH060 in in vitro assays. Utilizing comparative genomics on a larger collection of Sf6 genomes from diverse geographic locations, we identified similarity among Sf6 strains, despite disparate collection time frames and global locations. The unique genetic properties shared among Sf6, included potential virulence factors (Type II Secretion System) and genomic variation in the Type III Secretion System. We confirmed that geographically representative Sf6 strains from Africa and Asia demonstrated reduced invasion phenotype and Ipa effector secretion. Overexpression of Ipa proteins did not rescue the invasion phenotype. Through preliminary genomic and phenotypic comparisons of Sf6 to S. boydii serotypes 2, 4, and 14, we identified several Sf6 unique genes absent from non-Sf6 S. flexneri (Type II Secretion System and intrinsic antimicrobial resistance genes) and the reduced invasion phenotype was conserved in S. boydii. Together these data highlight highly conserved and unique Sf6 genotypic and phenotypic features, not found within other S. flexneri serotypes.
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  Weathering the Storms of Racial Capitalism: Examining the Impact of Racial Capitalism on Well-Being across the Black American Life Course

  Brown, Jocelyn; Wallace, Brandy (2024)

  This dissertation explores the pervasive effects of racial capitalism on the well-being of Black Americans across different age groups, investigating the mechanisms by which racial capitalism impacts their physical, mental, and socioeconomic health over the life course. Racial capitalism is defined as the interlocking systems of capitalism and racism, where capitalist economies and racial hierarchies converge to perpetuate and exploit racial disparities. Using constructivist grounded theory, this study engaged 27 Black Americans in interviews to understand their perspectives on the intersection of racism and capitalism, coping mechanisms, and views on reparations. The findings reveal a generational divide in the conceptualization of racial capitalism and its effects. Older Black Americans often view their experiences through lenses of personal resilience and historical progress, tending to disconnect the explicit links between racism and capitalism. In contrast, middle-aged Black Americans perceive a persistent social hierarchy that has adapted rather than diminished, recognizing ongoing systemic barriers despite social gains. Younger Black Americans express a critical awareness of racial capitalism, identifying it as an evolving and interlocking system that perpetuates socio-economic disparities through covert means. The study introduces the "Racial Capitalism Life Course Theory," positing that racism and capitalism function as dual systems that cyclically reinforce socio-economic inequalities across the life course and generations. This theory aligns with established theories of life course and cumulative disadvantage, but provides critical interdisciplinary lens to examine the intersecting impacts of racism and economic systems over time. Policy implications of this research are vast and underscore the need for targeted interventions that address the specific challenges faced by each age cohort while considering the broader historical and socio-political context. The dissertation advocates for a grassroots, community-focused approach to reparations and policy reforms, aiming to address both historical injustices and current disparities, with a significant emphasis on educational, healthcare, and economic reforms to dismantle the structural underpinnings of racial capitalism. This comprehensive exploration contributes to the fields of sociology, gerontology, and public health by integrating age-specific impacts of racial capitalism into broader societal and policy contexts, offering a framework for addressing the compounded effects of racism and economic exploitation that Black Americans face throughout their lives.
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  Subject-Specific Off-axis Assessment and Training Strategies for Medial Knee Osteoarthritis Rehabilitation

  baghi, raziyeh; Zhang, Li-Qun (2024)

  Excessive loading of the medial compartment of the knee and functional instability in the off-axis planes (frontal and transverse planes) are related to the development, progression, and severity of knee osteoarthritis (KOA). Gait modification strategies such as walking with a modified foot progression angle (FPA) and wider step-width can reduce peak knee adduction moment (pKAM), a surrogate measure of medial knee loading. Specifically, subject-specific FPAs are shown to be more effective in lowering KAM than a generalized FPA. However, detecting the minimum alteration in the kinematic gait parameters that reduce pKAM by a specific amount without causing an unnatural walking pattern has yet to be investigated. Another strategy to reduce pKAM is the electrically induced contraction of the primary knee muscles, such as the long head of the biceps femoris and lateral gastrocnemius, which provide resistance to pKAM. However, studies are scarce on the immediate effect of the enhanced activation of these muscles on pKAM during functional tasks such as elliptical stepping, an exercise recommended for KOA rehabilitation. Firstly, this work explored the feasibility of determining the subject-specific FPA and step-width during the footplates' motorized rotation and linear transition of a robotic elliptical stepping system. Study findings demonstrated a linear relationship between pKAM and FPA and pKAM and step-width during stepping. Using the characteristics of these relationships, we could estimate the minimum change in the FPA and step-width to induce a desired pKAM reduction. Secondly, this work explored the effect of functional electrical stimulation (FES) of the lateral knee muscles on pKAM during the stepping task. Participants reduced their pKAM with FES of lateral gastrocnemius and biceps femoris, which indicated the feasibility of using combined FES and elliptical stepping for pKAM reduction. Thirdly, the last study used a subject-specific training program using the robotic elliptical stepping system to assess the training’s effect on pKAM, functional instability, and clinical outcome measures in participants with KOA. Participants reduced their pKAM and functional instability and improved their stepping speed, gait velocity, and symptoms. These findings demonstrate the feasibility of subject-specific training with the robotic elliptical stepping system for KOA rehabilitation.
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  Clinical Investigation of the Impact of Endodontic Disinfection on the Bacteriome of Root Canal Infection Using Next-Generation Sequencing on the Illumina MiSeq Platform

  Alquria, Theeb Abdullah; Martinho, Frederico C. (2024)

  The primary cause of root canal infection is bacteria and their by-products, making disinfection of the root canal system a key goal in endodontic therapy. However, the complex anatomy of root canal systems, particularly the isthmus and its ramifications, poses challenges for effective disinfection. Currently, no disinfection protocol can eliminate all bacterial contents from root canal infections, driving the ongoing search for an optimal disinfection approach. Recently, next-generation sequencing (NGS), particularly the Illumina MiSeq platform, has been widely explored in endodontic infections due to its low sequencing error rates, cost-effectiveness, and high-quality reads. Leveraging advanced sequencing techniques to reveal the bacteriome of root canal infections and assess the impact of current disinfection methods could enable the development of more targeted and effective disinfection protocols. This dissertation presents an interventional clinical study aiming to investigate the diversity and composition of the bacteriome in primary endodontic infection (PEI) with apical periodontitis (AP) and evaluate the impact of root canal disinfection on the endodontic bacteriome using NGS on the Illumina MiSeq Platform. First, we characterized the bacteriome in PEI with AP, identified core and rare bacteriome species, and analyzed community diversity metrics using the Illumina MiSeq platform. Our results showed that Bacteroidetes, Firmicutes, Synergistetes, Fusobacteria, and Actinobacteria were the most abundant bacterial phyla. We identified 113 genera and 215 species. Analysis revealed differences in abundant taxa among distinct age, gender, symptomatology, and lesion size groups. These findings suggest that the bacteriome in PEI with AP is complex and has high microbial heterogeneity among patients. Moreover, age, gender, symptomatology, and lesion size might play a role in the abundant taxa present in PEI with AP. Second, we determined quantitatively and qualitatively the impact of chemomechanical preparation (CMP) using 2.5% sodium hypochlorite (NaOCl) on the bacteriome found in PEI with AP using the Illumina MiSeq platform. Our findings demonstrated a distinct community composition and increased alpha diversity after CMP using 2.5% NaOCl, despite a significant decrease in bacterial abundance. We observed differential enrichment of specific taxa, including Stenotrophomonas_unclassified, Enterococcus_unclassified, and Actinomyces_unclassified, suggesting lower effectiveness of CMP using 2.5% NaOCl against these taxa. Findings from this dissertation highlight the complexity and heterogeneity of the bacteriome in PEI with AP, emphasizing the influence of patient-related factors on microbial diversity. The research highlighted the limited effectiveness of current endodontic disinfection protocols, specifically the use of 2.5% NaOCl, in reducing bacterial abundance while revealing limitations against certain taxa. These insights provide a foundation for developing more targeted and effective disinfection strategies, potentially leading to improved outcomes in endodontic therapy.
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  Roles of Cytoskeleton Networks and Insulin metabolic pathway on Drosophila nephrocyte function and slit diaphragm dynamics

  Delaney, Megan; Han, Zhe (2024)

  Changes within the podocyte slit diaphragm, actin cytoskeleton, and tubulin cytoskeleton structures lead to varying forms of Nephrotic syndrome, a renal disease that affects the filtration function in every 16 of 100,000 individuals, a third of those being children. Using Drosophila melanogaster nephrocytes, podocyte-equivalent cells, I elucidated the roles of individual actin and tubulin genes, examining how they work together to form the sophisticated structures in the nephrocytes. I further investigate how the cytoskeleton crosslinking protein GAS2L1 (Pickled eggs, Pigs) affects nephrocyte structure and function. Diabetes can also affect these podocyte structural components, resulting in diabetic nephropathy. I examined the environmental factor via high sugar diets and genetic manipulation of the insulin metabolic pathway to model diabetes, revealing the slit diaphragm and cytoskeleton structural defects in diabetic nephropathy. Furthermore, I show metformin, an antidiabetic drug, can be used to rescue nephrocyte function and structure using slit diaphragm, actin cytoskeleton, tubulin cytoskeleton, and mitochondria markers. This study provides essential insight to understand the filtration structure and sets up a platform to test new anti-diabetic medications using the nephrocyte model. The outlined markers can then be used to understand the medication’s mechanisms of action, allowing the clinical trial process to be streamlined through the reduction of time and resources necessary by utilizing Drosophila nephrocytes as the initial screening model.
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  A Biopsychosocial Model to Study Interindividual Differences in Placebo Effects: Translational Approaches for Acute and Chronic Pain Management

  White, Jewel; Colloca, Luana (2024)

  To date, chronic pain is among the most prevalent and economically burdensome chronic diseases, affecting nearly one in five individuals worldwide and costing $635 billion per year in the United States alone. Current treatments for chronic pain are limited or mildly efficacious, and some may produce adverse side effects and conditions such as drug-induced hyperalgesia or substance use disorder. Limitations of pharmacological treatments urge the development of nonpharmacological therapies for pain. Placebo effects engage the descending modulatory neural systems to induce analgesia and present a promising opportunity to study the effectiveness of nonpharmacological and noninvasive interventions for improving pain symptoms. However, not everyone will respond to a placebo treatment. Biopsychosocial factors that vary between individuals, including culture, environment, sex, and neural characteristics, may influence individual response to placebo. Therefore, understanding the mechanisms underlying the effects of biopsychosocial factors on interindividual differences in placebo effects is critical for the development of translational approaches for pain management. Here, two well-established social learning and classical conditioning paradigms were used to test the hypothesis that social, sex, and neural traits would contribute to the generation of placebo effects. The main findings of this work are threefold. First, cognitive state empathy was greater for a human demonstrator in pain compared to an avatar, and this socially induced empathy mediated subsequently induced placebo effects. Second, women exhibited larger expectations and placebo effects compared to men, and gonadal hormone levels were associated with conditioning strength, expectations, and placebo hypoalgesia. Testosterone levels were negatively associated with chronic pain severity in women, and estradiol levels in individuals with chronic pain and pain-free individuals mediated the impact of sex on expectations assessed after conditioning. These expectations mediated the impact of sex on placebo effects in individuals with chronic pain. Third, cortical morphology was positively correlated with placebo response in individuals with chronic pain. This work provides support for the careful consideration of the effects of biopsychosocial factors in the analysis and interpretation of placebo hypoalgesia studies. Importantly, these findings suggest that empathy, sex hormone, and morphological measurements can be important factors to leverage for translational and clinical approaches for chronic pain management.
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  Impact of SARS-CoV-2 Salivary Gland Infection on the Production of the Antifungal Peptide Histatin-5, Candida Colonization, and the oral Microbiota.

  Alfaifi, Areej; Jabra-Rizk, Mary Ann (2024)

  Despite the myriad oral manifestations during COVID-19 and presence of SARS-CoV-2 in saliva, the oral cavity remains an underappreciated site. However recent findings have shown that SARS-CoV-2 can replicate in salivary gland cells, leading to inflammation and tissue destruction. Saliva contains antimicrobial peptides considered integral components of innate immunity crucial for oral health, with the most notable being histatin-5. This peptide is exclusively produced in salivary glands and exhibits unique potent antifungal activity against Candida albicans. In this dissertation, we discovered that destruction of salivary gland by SARS-CoV-2 can compromise histatin-5 production, predisposing patients to oral candidiasis and dysbiosis in the oral microbiome. First, we utilized our novel customized immunoassay to measure salivary histatin-5 levels in a prospective study involving stratified COVID-19 cohorts. Our data indicates a trend showing a decrease in salivary histatin-5 and increase in Candida during COVID-19, persisting post-COVID-19 recovery, potentially contributing to the long COVID-19 syndrome. To provide lacking mechanistic insights into the pathophysiology of salivary gland dysfunction during COVID-19, we performed in situ hybridization coupled with immunofluorescence to co-localize SARS-C0V-2 and histatin-5, respectively, in salivary gland tissue from deceased COVID-19 patients. Our findings indicated diminished or absent histatin presence in salivary gland acini with proliferating SARS-CoV-2 providing the first direct evidence associating SARS-CoV-2 with histatin-5 production. Next, we conducted a comprehensive metagenomic analysis on clinical oral samples and identified potential COVID-19 associated pathologic dysbiotic shifts in the oral microbiome. Lastly, we conducted in-vitro experiments coupled with scanning electron microscopy and confocal imaging to identify the effect of histatin-5 on candida cells and biofilm concluding the antifungal effect of histatin-5 on candida albicans. This clinical study clearly shows the effect of SARS-CoV-2 on oral microbiota, highlighting the importance of understanding and managing the complex dynamics within the oral cavity of COVID-19 patients.
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  From Zinc Fingers to Fe-S Clusters to Iron Nanoparticle Drugs: Understanding the Impact of Metals on Biological Proteins and Formulations

  Hursey, Matthew; Michel, Sarah L.J. (2024)

  Iron and zinc co-factored metalloproteins perform a variety of functions including providing structural integrity, aiding in transport and storage, and engaging in enzymatic activity. One important group of metalloproteins are zinc finger (ZF) proteins. In this thesis, I investigate two RNA-binding ZFs to understand both structural and functional aspects and aim to highlight their importance in biology. The cleavage and polyadenylation specificity factor 30 (CPSF30) is a non-classical ZF containing five CCCH-type and one CCHC-type ZF domains that binds RNA targets. One of CPSF30’s CCCH ZF domains can bind a 2Fe-2S cluster; however, the role of this cluster is poorly understood. RNA binding assays on CPSF30 determined that the CCCH domains bind AU-rich RNA and the CCHC domain binds U-rich RNA. Metal-catalyzed oxidation – mass spectrometry (MCO-MS) identified the site of the 2Fe-2S cluster as the second CCCH domain. Additional EPR and Mössbauer spectroscopies demonstrated the 2Fe-2S cluster is redox active, however, the redox activity doesn’t affect RNA binding. I characterized individual CCCH-type maquettes of CPSF30 to determine if Fe-S cluster binding can occur in the other domains. Not only can each individual domain load an Fe-S cluster as confirmed by UV-Vis and XAS, but the clusters are redox active, as confirmed by EPR. Another non-classical ZF is the Ran-binding domain containing protein 2 (ZRANB2). ZRANB2 contains two CCCC-type ZF domains and functions to bind RNA, interact with other proteins, and participate in alternate splicing. I determined the effect of zinc binding on ZRANB2 conformation and analyzed protein dynamics with RNA utilizing UV-Vis, CD, fluorescence assays, and HDX-MS. ZRANB2 is found to be persulfidated in a variety of cell lines when measured by persulfide specific proteomics. I demonstrate that isolated ZRANB2 is persulfidated by H2S in a Zn and O2 dependent manner via an in situ dimedone switch tagging method. Superoxide was determined to be an intermediate of the persulfidation reaction and persulfidation abrogated RNA binding. I proposed that this modification is linked to regulation of the spliceosome. Lastly at the end of my thesis, through a variety of techniques, I analyze the physicochemical properties of FDA-approved iron nanoparticle drug, Monoferric.

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