College of Sciences Faculty Research

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    Development of Aptamers for RNase Inactivation in Xtract-Free™ Sample Collection and Transport Medium
    (MDPI, 2024-06-07) Daum, Luke T.; Rodriguez, John D.; Chambers, James P.
    There is a significant need to develop new environmentally friendly, extraction-free sample collection mediums that can effectively preserve and protect genetic material for point-of-care and/or self-collection, home-collection, and mail-back testing. Systematic evolution of ligands by exponential enrichment (SELEX) was used to create anti-ribonuclease (RNase) deoxyribonucleic acid (DNA) aptamers against purified RNase A conjugated to paramagnetic carboxylated beads. Following eight rounds of SELEX carried out under various stringency conditions, e.g., selection using Xtract-Free™ (XF) specimen collection medium and elevated ambient temperature of 28 °C, a panel of five aptamers was chosen following bioinformatic analysis using next-generation sequencing. The efficacy of aptamer inactivation of RNase was assessed by monitoring ribonucleic acid (RNA) integrity via fluorometric and real-time RT-PCR analysis. Inclusion of aptamers in reaction incubations resulted in an 8800- to 11,200-fold reduction in RNase activity, i.e., digestion of viral RNA compared to control. Thus, anti-RNase aptamers integrated into XF collection medium as well as other commercial reagents and kits have great potential for ensuring quality intact RNA for subsequent genomic analyses.
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    Current Progress in the Science of Novel Adjuvant Nano-Vaccine-Induced Protective Immune Responses
    (MDPI, 2024-05-23) Saleemi, Mansab Ali; Zhang, Yan; Zhang, Guoquan
    Vaccinations are vital as they protect us from various illness-causing agents. Despite all the advancements in vaccine-related research, developing improved and safer vaccines against devastating infectious diseases including Ebola, tuberculosis and acquired immune deficiency syndrome (AIDS) remains a significant challenge. In addition, some of the current human vaccines can cause adverse reactions in some individuals, which limits their use for massive vaccination program. Therefore, it is necessary to design optimal vaccine candidates that can elicit appropriate immune responses but do not induce side effects. Subunit vaccines are relatively safe for the vaccination of humans, but they are unable to trigger an optimal protective immune response without an adjuvant. Although different types of adjuvants have been used for the formulation of vaccines to fight pathogens that have high antigenic diversity, due to the toxicity and safety issues associated with human-specific adjuvants, there are only a few adjuvants that have been approved for the formulation of human vaccines. Recently, nanoparticles (NPs) have gain specific attention and are commonly used as adjuvants for vaccine development as well as for drug delivery due to their excellent immune modulation properties. This review will focus on the current state of adjuvants in vaccine development, the mechanisms of human-compatible adjuvants and future research directions. We hope this review will provide valuable information to discovery novel adjuvants and drug delivery systems for developing novel vaccines and treatments.
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    Characterization of Excited-State Electronic Structure in Diblock π-Conjugated Oligomers with Adjustable Linker Electronic Coupling
    (MDPI, 2024-06-05) Gobeze, Habtom B.; Younus, Muhammed; Turlington, Michael D.; Ahmed, Sohel; Schanze, Kirk S.
    Diblock conjugated oligomers are π-conjugated molecules that contain two segments having distinct frontier orbital energies and HOMO-LUMO gap offsets. These oligomers are of fundamental interest to understand how the distinct π-conjugated segments interact and modify their excited state properties. The current paper reports a study of two series of diblock oligomers that contain oligothiophene (Tn) and 4,7-bis(2-thienyl)-2,1,3-benzothiadiazole (TBT) segments that are coupled by either ethynyl (-C≡C-) or trans-(-C≡C-)2Pt(II)(PBu3)2 acetylide linkers. In these structures, the Tn segment is electron rich (donor), and the TBT is electron poor (acceptor). The diblock oligomers are characterized by steady-state and time-resolved spectroscopy, including UV-visible absorption, fluorescence, fluorescence lifetimes, and ultrafast transient absorption spectroscopy. Studies are compared in several solvents of different polarity and with different excitation wavelengths. The results reveal that the (-C≡C-) linked oligomers feature a delocalized excited state that takes on a charge transfer (CT) character in more polar media. In the (-C≡C-)2Pt(II)(PBu3)2-linked oligomers, there is weak coupling between the Tn and TBT segments. Consequently, short wavelength excitation selectively excites the Tn segment, which then undergoes ultrafast energy transfer (~1 ps) to afford a TBT-localized excited state.
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    A Standardized Nomenclature Design for Systematic Referencing and Identification of Animal Cellular Material
    (MDPI, 2024-05-23) Schrade, Lisa; Mah, Nancy; Bandrowski, Anita; Chen, Ying; Dewender, Johannes; Diecke, Sebastian; Hiepen, Christian; Lancaster, Madeline A.; Marques-Bonet, Tomas; Martinez, Sira; Mueller, Sabine C.; Navara, Christopher; Prigione, Alessandro; Seltmann, Stefanie; Sochacki, Jaroslaw; Sutcliffe, Magdalena A.; Zywitza, Vera; Hildebrandt, Thomas B.; Kurtz, Andreas
    The documentation, preservation and rescue of biological diversity increasingly uses living biological samples. Persistent associations between species, biosamples, such as tissues and cell lines, and the accompanying data are indispensable for using, exchanging and benefiting from these valuable materials. Explicit authentication of such biosamples by assigning unique and robust identifiers is therefore required to allow for unambiguous referencing, avoid identification conflicts and maintain reproducibility in research. A predefined nomenclature based on uniform rules would facilitate this process. However, such a nomenclature is currently lacking for animal biological material. We here present a first, standardized, human-readable nomenclature design, which is sufficient to generate unique and stable identifying names for animal cellular material with a focus on wildlife species. A species-specific human- and machine-readable syntax is included in the proposed standard naming scheme, allowing for the traceability of donated material and cultured cells, as well as data FAIRification. Only when it is consistently applied in the public domain, as publications and inter-institutional samples and data are exchanged, distributed and stored centrally, can the risks of misidentification and loss of traceability be mitigated. This innovative globally applicable identification system provides a standard for a sustainable structure for the long-term storage of animal bio-samples in cryobanks and hence facilitates current as well as future species conservation and biomedical research.
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    Improving the Concrete Crack Detection Process via a Hybrid Visual Transformer Algorithm
    (MDPI, 2024-05-20) Shahin, Mohammad; Chen, F. Frank; Maghanaki, Mazdak; Hosseinzadeh, Ali; Zand, Neda; Khodadadi Koodiani, Hamid
    Inspections of concrete bridges across the United States represent a significant commitment of resources, given their biannual mandate for many structures. With a notable number of aging bridges, there is an imperative need to enhance the efficiency of these inspections. This study harnessed the power of computer vision to streamline the inspection process. Our experiment examined the efficacy of a state-of-the-art Visual Transformer (ViT) model combined with distinct image enhancement detector algorithms. We benchmarked against a deep learning Convolutional Neural Network (CNN) model. These models were applied to over 20,000 high-quality images from the Concrete Images for Classification dataset. Traditional crack detection methods often fall short due to their heavy reliance on time and resources. This research pioneers bridge inspection by integrating ViT with diverse image enhancement detectors, significantly improving concrete crack detection accuracy. Notably, a custom-built CNN achieves over 99% accuracy with substantially lower training time than ViT, making it an efficient solution for enhancing safety and resource conservation in infrastructure management. These advancements enhance safety by enabling reliable detection and timely maintenance, but they also align with Industry 4.0 objectives, automating manual inspections, reducing costs, and advancing technological integration in public infrastructure management.
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    The Dolan Fire of Central Coastal California: Burn Severity Estimates from Remote Sensing and Associations with Environmental Factors
    (MDPI, 2024-05-10) Oseghae, Iyare; Bhaganagar, Kiran; Mestas-Nuñez, Alberto M.
    In 2020, wildfires scarred over 4,000,000 hectares in the western United States, devastating urban populations and ecosystems alike. The significant impact that wildfires have on plants, animals, and human environments makes wildfire adaptation, management, and mitigation strategies a critical task. This study uses satellite imagery from Landsat to calculate burn severity and map the fire progression for the Dolan Fire of central Coastal California which occurred in August 2020. Several environmental factors, such as temperature, humidity, fuel type, topography, surface conditions, and wind velocity, are known to affect wildfire spread and burn severity. The aim of this study is the investigation of the relationship between these environmental factors, estimates of burn severity, and fire spread patterns. Burn severity is calculated and classified using the Difference in Normalized Burn Ratio (dNBR) before being displayed as a time series of maps. The Dolan Fire had a moderate severity burn with an average dNBR of 0.292. The ignition site location, when paired with the patterns of fire spread, is consistent with wind speed and direction data, suggesting fire movement to the southeast of the fire ignition site. Patterns of increased burn severity are compared with both topography (slope and aspect) and fuel type. Locations that were found to be more susceptible to high burn severity featured Long Needle Timber Litter and Mature Timber fuels, intermediate slope angles between 15 and 35°, and north- and east-facing slopes. This study has implications for the future predictive modeling of wildfires that may serve to develop wildfire mitigation strategies, manage climate change impacts, and protect human lives.
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    Role of Type 4B Secretion System Protein, IcmE, in the Pathogenesis of Coxiella burnetii
    (MDPI, 2024-05-14) Palanisamy, Rajesh; Zhang, Yan; Zhang, Guoquan
    Coxiella burnetii is an obligate intracellular Gram-negative bacterium that causes Q fever, a life-threatening zoonotic disease. C. burnetii replicates within an acidified parasitophorous vacuole derived from the host lysosome. The ability of C. burnetii to replicate and achieve successful intracellular life in the cell cytosol is vastly dependent on the Dot/Icm type 4B secretion system (T4SSB). Although several T4SSB effector proteins have been shown to be important for C. burnetii virulence and intracellular replication, the role of the icmE protein in the host–C. burnetii interaction has not been investigated. In this study, we generated a C. burnetii Nine Mile Phase II (NMII) mutant library and identified 146 transposon mutants with a single transposon insertion. Transposon mutagenesis screening revealed that disruption of icmE gene resulted in the attenuation of C. burnetii NMII virulence in SCID mice. ELISA analysis indicated that the levels of pro-inflammatory cytokines, including interleukin-1β, IFN-γ, TNF-α, and IL-12p70, in serum from Tn::icmE mutant-infected SCID mice were significantly lower than those in serum from wild-type (WT) NMII-infected mice. Additionally, Tn::icmE mutant bacteria were unable to replicate in mouse bone marrow-derived macrophages (MBMDM) and human macrophage-like cells (THP-1). Immunoblotting results showed that the Tn::icmE mutant failed to activate inflammasome components such as IL-1β, caspase 1, and gasdermin-D in THP-1 macrophages. Collectively, these results suggest that the icmE protein may play a vital role in C. burnetii virulence, intracellular replication, and activation of inflammasome mediators during NMII infection.
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    On the Controllability of Coupled Nonlocal Partial Integrodifferential Equations Using Fractional Power Operators
    (MDPI, 2024-04-30) Litimein, Hamida; Huang, Zhen-You; Ouahab, Abdelghani; Stamova, Ivanka; Souid, Mohammed Said
    In this research paper, we investigate the controllability in the α-norm of a coupled system of integrodifferential equations with state-dependent nonlocal conditions in generalized Banach spaces. We establish sufficient conditions for the system’s controllability using resolvent operator theory introduced by Grimmer, fractional power operators, and fixed-point theorems associated with generalized measures of noncompactness for condensing operators in vector Banach spaces. Finally, we present an application example to validate the proposed methodology in this research.
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    Astrogliosis in the GFAP-CreERT2:Rosa26iDTR Mouse Model Does Not Exacerbate Retinal Microglia Activation or Müller Cell Gliosis under Hypoxic Conditions
    (MDPI, 2024-05-10) Rorex, Colin; Cardona, Sandra M.; Church, Kaira A.; Rodriguez, Derek; Vanegas, Difernando; Saldivar, Reina; Faz, Brianna; Cardona, Astrid E.
    Diabetic retinopathy (DR) affects over 140 million people globally. The mechanisms that lead to blindness are still enigmatic but there is evidence that sustained inflammation and hypoxia contribute to vascular damage. Despite efforts to understand the role of inflammation and microglia in DR’s pathology, the contribution of astrocytes to hypoxic responses is less clear. To investigate the role of astrocytes in hypoxia-induced retinopathy, we utilized a 7-day systemic hypoxia model using the GFAP-CreERT2:Rosa26iDTR transgenic mouse line. This allows for the induction of inflammatory reactive astrogliosis following tamoxifen and diphtheria toxin administration. We hypothesize that DTx-induced astrogliosis is neuroprotective during hypoxia-induced retinopathy. Glial, neuronal, and vascular responses were quantified using immunostaining, with antibodies against GFAP, vimentin, IBA-1, NeuN, fibrinogen, and CD31. Cytokine responses were measured in both the brain and serum. We report that while both DTx and hypoxia induced a phenotype of reduced microglia morphological activation, DTx, but not hypoxia, induced an increase in the Müller glia marker vimentin. We did not observe that the combination of DTx and hypoxic treatments exacerbated the signs of reactive glial cells, nor did we observe a significant change in the expression immunomodulatory mediators IL-1β, IL2, IL-4, IL-5, IL-6, IL-10, IL-18, CCL17, TGF-β1, GM-CSF, TNF-α, and IFN-γ. Overall, our results suggest that, in this hypoxia model, reactive astrogliosis does not alter the inflammatory responses or cause vascular damage in the retina.
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    On Solutions of Two Post-Quantum Fractional Generalized Sequential Navier Problems: An Application on the Elastic Beam
    (MDPI, 2024-04-17) Etemad, Sina; Ntouyas, Sotiris K.; Stamova, Ivanka; Tariboon, Jessada
    Fractional calculus provides some fractional operators for us to model different real-world phenomena mathematically. One of these important study fields is the mathematical model of the elastic beam changes. More precisely, in this paper, based on the behavior patterns of an elastic beam, we consider the generalized sequential boundary value problems of the Navier difference equations by using the post-quantum fractional derivatives of the Caputo-like type. We discuss on the existence theory for solutions of the mentioned (p;q)-difference Navier problems in two single-valued and set-valued versions. We use the main properties of the (p;q)-operators in this regard. Application of the fixed points of the ρ-θ-contractions along with the endpoints of the multi-valued functions play a fundamental role to prove the existence results. Finally in two examples, we validate our models and theoretical results by giving numerical models of the generalized sequential (p;q)-difference Navier problems.
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    Pathogenomes of Shiga Toxin Positive and Negative Escherichia coli O157:H7 Strains TT12A and TT12B: Comprehensive Phylogenomic Analysis Using Closed Genomes
    (MDPI, 2024-03-29) Kalalah, Anwar A.; Koenig, Sara S. K.; Feng, Peter; Bosilevac, Joseph M.; Bono, James L.; Eppinger, Mark
    Shiga toxin-producing Escherichia coli are zoonotic pathogens that cause food-borne human disease. Among these, the O157:H7 serotype has evolved from an enteropathogenic O55:H7 ancestor through the displacement of the somatic gene cluster and recurrent toxigenic conversion by Shiga toxin-converting bacteriophages. However, atypical strains that lack the Shiga toxin, the characteristic virulence hallmark, are circulating in this lineage. For this study, we analyzed the pathogenome and virulence inventories of the stx+ strain, TT12A, isolated from a patient with hemorrhagic colitis, and its respective co-isolated stx− strain, TT12B. Sequencing the genomes to closure proved critical to the cataloguing of subtle strain differentiating sequence and structural polymorphisms at a high-level of phylogenetic accuracy and resolution. Phylogenomic profiling revealed SNP and MLST profiles similar to the near clonal outbreak isolates. Their prophage inventories, however, were notably different. The attenuated atypical non-shigatoxigenic status of TT12B is explained by the absence of both the ΦStx1a- and ΦStx2a-prophages carried by TT12A, and we also recorded further alterations in the non-Stx prophage complement. Phenotypic characterization indicated that culture growth was directly impacted by the strains’ distinct lytic phage complement. Altogether, our phylogenomic and phenotypic analyses show that these intimately related isogenic strains are on divergent Stx(+/stx−) evolutionary paths.
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    Development and Evaluation of an Immunoinformatics-Based Multi-Peptide Vaccine against Acinetobacter baumannii Infection
    (MDPI, 2024-03-27) Jeffreys, Sean; Tompkins, Megan P.; Aki, Jadelynn; Papp, Sara B.; Chambers, James P.; Guentzel, M. Neal; Hung, Chiung-Yu; Yu, Jieh-Juen; Arulanandam, Bernard P.
    Multi-drug-resistant (MDR) Acinetobacter baumannii is an opportunistic pathogen associated with hospital-acquired infections. Due to its environmental persistence, virulence, and limited treatment options, this organism causes both increased patient mortality and incurred healthcare costs. Thus, prophylactic vaccination could be ideal for intervention against MDR Acinetobacter infection in susceptible populations. In this study, we employed immunoinformatics to identify peptides containing both putative B- and T-cell epitopes from proteins associated with A. baumannii pathogenesis. A novel Acinetobacter Multi-Epitope Vaccine (AMEV2) was constructed using an A. baumannii thioredoxin A (TrxA) leading protein sequence followed by five identified peptide antigens. Antisera from A. baumannii infected mice demonstrated reactivity to rAMEV2, and subcutaneous immunization of mice with rAMEV2 produced high antibody titer against the construct as well as peptide components. Immunization results in increased frequency of IL-4-secreting splenocytes indicative of a Th2 response. AMEV2-immunized mice were protected against intranasal challenge with a hypervirulent strain of A. baumannii and demonstrated reduced bacterial burden at 48 h. In contrast, all mock vaccinated mice succumbed to infection within 3 days. Results presented here provide insight into the effectiveness of immunoinformatic-based vaccine design and its potential as an effective strategy to combat the rise of MDR pathogens.
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    Fabrication and Characterization of Quad-Component Bioinspired Hydrogels to Model Elevated Fibrin Levels in Central Nervous Tissue Scaffolds
    (MDPI, 2024-03-17) Diaz-Lasprilla, Ana M.; McKee, Meagan; Jimenez-Vergara, Andrea C.; Ravi, Swathisri; Bellamy, Devon; Ortega, Wendy; Crosby, Cody O.; Steele, Jennifer; Plascencia-Villa, Germán; Perry, George; Munoz-Pinto, Dany J.
    Multicomponent interpenetrating polymer network (mIPN) hydrogels are promising tissue-engineering scaffolds that could closely resemble key characteristics of native tissues. The mechanical and biochemical properties of mIPNs can be finely controlled to mimic key features of target cellular microenvironments, regulating cell-matrix interactions. In this work, we fabricated hydrogels made of collagen type I (Col I), fibrin, hyaluronic acid (HA), and poly (ethylene glycol) diacrylate (PEGDA) using a network-by-network fabrication approach. With these mIPNs, we aimed to develop a biomaterial platform that supports the in vitro culture of human astrocytes and potentially serves to assess the effects of the abnormal deposition of fibrin in cortex tissue and simulate key aspects in the progression of neuroinflammation typically found in human pathologies such as Alzheimer's disease (AD), Parkinson's disease (PD), and tissue trauma. Our resulting hydrogels closely resembled the complex modulus of AD human brain cortex tissue (~7.35 kPa), promoting cell spreading while allowing for the modulation of fibrin and hyaluronic acid levels. The individual networks and their microarchitecture were evaluated using confocal laser scanning microscopy (CLSM) and scanning electron microscopy (SEM). Human astrocytes were encapsulated in mIPNs, and negligible cytotoxicity was observed 24 h after the cell encapsulation.
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    A Numerical Study of Quantum Entropy and Information in the Wigner–Fokker–Planck Equation for Open Quantum Systems
    (MDPI, 2024-03-14) Edrisi, Arash; Patwa, Hamza; Morales Escalante, Jose A.
    Kinetic theory provides modeling of open quantum systems subject to Markovian noise via the Wigner–Fokker–Planck equation, which is an alternate of the Lindblad master equation setting, having the advantage of great physical intuition as it is the quantum equivalent of the classical phase space description. We perform a numerical inspection of the Wehrl entropy for the benchmark problem of a harmonic potential, since the existence of a steady state and its analytical formula have been proven theoretically in this case. When there is friction in the noise terms, no theoretical results on the monotonicity of absolute entropy are available. We provide numerical results of the time evolution of the entropy in the case with friction using a stochastic (Euler–Maruyama-based Monte Carlo) numerical solver. For all the chosen initial conditions studied (all of them Gaussian states), up to the inherent numerical error of the method, one cannot disregard the possibility of monotonic behavior even in the case under study, where the noise includes friction terms.
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    The Host Response to Coccidioidomycosis
    (MDPI, 2024-02-25) Kirkland, Theo N.; Hung, Chiung-Yu; Shubitz, Lisa F.; Beyhan, Sinem; Fierer, Joshua
    Coccidioidomycosis is an important fungal disease that is found in many desert regions of the western hemisphere. The inhaled organisms are highly pathogenic, but only half of infected, immunologically intact people develop symptomatic pneumonia; most symptomatic infections resolve spontaneously, although some resolve very slowly. Furthermore, second infections are very rare and natural immunity after infection is robust. Therefore, the host response to this organism is very effective at resolving the infection in most cases and immunizing to prevent second infections. People who are immunocompromised are much more likely to develop disseminated infection. This is a comprehensive review of the innate and acquired immune responses to Coccidioides spp., the genetics of resistance to severe infection, and the search for an effective vaccine.
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    Galleria mellonella Model of Coccidioidomycosis for Drug Susceptibility Tests and Virulence Factor Identification
    (MDPI, 2024-02-05) Mendoza Barker, Matthew; Saeger, Sarah; Campuzano, Althea; Yu, Jieh-Juen; Hung, Chiung-Yu
    Coccidioidomycosis (CM) can manifest as respiratory and disseminated diseases that are caused by dimorphic fungal pathogens, such as Coccidioides species. The inhaled arthroconidia generated during the saprobic growth phase convert into multinucleated spherules in the lungs to complete the parasitic lifecycle. Research on coccidioidal virulence and pathogenesis primarily employs murine models typically associated with low lethal doses (LD100 < 100 spores). However, the Galleria model has recently garnered attention due to its immune system bearing both structural and functional similarities to the innate system of mammals. Our findings indicate that Coccidioides posadasii can convert and complete the parasitic cycle within the hemocoel of the Galleria larva. In Galleria, the LD100 is between 0.5 and 1.0 × 106 viable spores for the clinical isolate Coccidioides posadasii C735. Furthermore, we demonstrated the suitability of this model for in vivo antifungal susceptibility tests to validate the bioreactivity of newly discovered antifungals against Coccidioides. Additionally, we utilized this larva model to screen a Coccidioides posadasii mutant library showing attenuated virulence. Similarly, the identified attenuated coccidioidal mutants displayed a loss of virulence in a commonly used murine model of coccidioidomycosis. In this study, we demonstrated that Galleria larvae can be applied as a model for studying Coccidioides infection.
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    Therapeutic Delivery of Soluble Fractalkine Ameliorates Vascular Dysfunction in the Diabetic Retina
    (MDPI, 2024-01-31) Rodriguez, Derek; Church, Kaira A.; Smith, Chelsea T.; Vanegas, Difernando; Cardona, Sandra M.; Muzzio, Isabel A.; Nash, Kevin R.; Cardona, Astrid E.
    Diabetic retinopathy (DR)-associated vision loss is a devastating disease affecting the working-age population. Retinal pathology is due to leakage of serum components into retinal tissues, activation of resident phagocytes (microglia), and vascular and neuronal damage. While short-term interventions are available, they do not revert visual function or halt disease progression. The impact of microglial inflammatory responses on the neurovascular unit remains unknown. In this study, we characterized microglia–vascular interactions in an experimental model of DR. Early diabetes presents activated retinal microglia, vascular permeability, and vascular abnormalities coupled with vascular tortuosity and diminished astrocyte and endothelial cell-associated tight-junction (TJ) and gap-junction (GJ) proteins. Microglia exclusively bind to the neuronal-derived chemokine fractalkine (FKN) via the CX3CR1 receptor to ameliorate microglial activation. Using neuron-specific recombinant adeno-associated viruses (rAAVs), we therapeutically overexpressed soluble (sFKN) or membrane-bound (mFKN) FKN using intra-vitreal delivery at the onset of diabetes. This study highlights the neuroprotective role of rAAV-sFKN, reducing microglial activation, vascular tortuosity, fibrin(ogen) deposition, and astrogliosis and supporting the maintenance of the GJ connexin-43 (Cx43) and TJ zonula occludens-1 (ZO-1) molecules. The results also show that microglia–vascular interactions influence the vascular width upon administration of rAAV-sFKN and rAAV-mFKN. Administration of rAAV-sFKN improved visual function without affecting peripheral immune responses. These findings suggest that overexpression of rAAV-sFKN can mitigate vascular abnormalities by promoting glia–neural signaling. sFKN gene therapy is a promising translational approach to reverse vision loss driven by vascular dysfunction.
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    Dietary Supplementation with 23-Hydroxy Ursolic Acid Reduces the Severity and Incidence of Acute Experimental Autoimmune Encephalomyelitis (EAE) in a Murine Model of Multiple Sclerosis
    (MDPI, 2024-01-25) Asmis, Reto; Medrano, Megan T.; Chase Huizar, Carol; Griffith, Wendell P.; Forsthuber, Thomas G.
    23-Hydroxy ursolic acid (23-OH UA) is a potent atheroprotective and anti-obesogenic phytochemical, with anti-inflammatory and inflammation-resolving properties. In this study, we examined whether dietary 23-OH UA protects mice against the acute onset and progression of experimental autoimmune encephalomyelitis (EAE), a mouse model of multiple sclerosis (MS). Female C57BL/6 mice were fed either a defined low-calorie maintenance diet (MD) or an MD supplemented with 0.2% wgt/wgt 23-OH UA for 5 weeks prior to actively inducing EAE and during the 30 days post-immunization. We observed no difference in the onset of EAE between the groups of mice, but ataxia and EAE disease severity were suppressed by 52% and 48%, respectively, and disease incidence was reduced by over 49% in mice that received 23-OH UA in their diet. Furthermore, disease-associated weight loss was strikingly ameliorated in 23-OH UA-fed mice. ELISPOT analysis showed no significant differences in frequencies of T cells producing IL-17 or IFN-γ between 23-OH UA-fed mice and control mice, suggesting that 23-OH UA does not appear to regulate peripheral T cell responses. In summary, our findings in EAE mice strongly suggest that dietary 23-OH UA may represent an effective oral adjunct therapy for the prevention and treatment of relapsing–remitting MS.
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    A Recombinant Multivalent Vaccine (rCpa1) Induces Protection for C57BL/6 and HLA Transgenic Mice against Pulmonary Infection with Both Species of Coccidioides
    (MDPI, 2024-01-09) Campuzano, Althea; Pentakota, Komali Devi; Liao, Yu-Rou; Zhang, Hao; Wiederhold, Nathan P.; Ostroff, Gary R.; Hung, Chiung-Yu
    Coccidioidomycosis is caused by Coccidioides posadasii (Cp) and Coccidioides immitis (Ci), which have a 4–5% difference in their genomic sequences. There is an urgent need to develop a human vaccine against both species. A previously created recombinant antigen (rCpa1) that contains multiple peptides derived from Cp isolate C735 is protective against the autologous isolate. The focus of this study is to evaluate cross-protective efficacy and immune correlates by the rCpa1-based vaccine against both species of Coccidioides. DNA sequence analyses of the homologous genes for the rCpa1 antigen were conducted for 39 and 17 clinical isolates of Cp and Ci, respectively. Protective efficacy and vaccine-induced immunity were evaluated for both C57BL/6 and human HLA-DR4 transgenic mice against five highly virulent isolates of Cp and Ci. There are total of seven amino acid substitutions in the rCpa1 antigen between Cp and Ci. Both C57BL/6 and HLA-DR4 mice that were vaccinated with an rCpa1 vaccine had a significant reduction of fungal burden and increased numbers of IFN-γ- and IL-17-producing CD4+ T cells in the first 2 weeks post challenge. These data suggest that rCpa1 has cross-protection activity against Cp and Ci pulmonary infection through activation of early Th1 and Th17 responses.
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    Irradiation of ZnPPIX Complexed with Bovine β-Lactoglobulin Causes Chemical Modifications and Conformational Changes of the Protein
    (MDPI, 2023-11-29) Albalawi, Abdullah; Castillo, Omar; Denton, Michael L.; Rickman, John Michael; Noojin, Gary D.; Brancaleon, Lorenzo
    Photosensitization of proteins mediated by chromophores is a mechanism commonly employed by nature and mimicked in a broad array of laboratory research and applications. Nature has evolved specialized complexes of proteins and photosensitizers (PS) that assemble to form photoreceptor proteins (PRP). These are used by many organisms in diverse processes, such as energy conversion, protection against photodamage, etc. The same concept has been used in laboratory settings for many applications, such as the stimulation of neurons or the selective depletion of proteins in a signaling pathway. A key issue in laboratory settings has been the relationship between the photooxidation of proteins and conformational changes in host proteins. For several years, we have been interested in creating non-native PRP using porphyrin PS. In this study, we investigated the self-assembled complex between zinc protoporphyrin IX (ZnPPIX) and bovine β-lactoglobulin (BLG) as a model of non-native PRP. Since BLG undergoes a significant conformational transition near physiological pH, the study was carried out at acidic (pH 5) and alkaline (pH 9) conditions where the two conformations are respectively prevalent. We employed a series of steady-state and time-resolved optical spectroscopies as well as gel electrophoresis to experimentally characterize the photosensitization mechanisms and their effect on the host protein. Our results show that ZnPPIX prompts light-dependent modifications of BLG, which appear to be much more significant at alkaline pH. The modifications seem to be driven by photooxidation of amino acid residues that do not lead to the formation of cross-links or protein fragmentation.