Grant Program Research Articles

Protective effects of Heat-killed Lactobacillus sakeiCVL-001 on DSS-induced ulcerative colitis in mice

Abstract Recently, the incidence of Inflammatory Bowel Disease (IBD) is increasing due to westernized lifestyle and eating habits. IBD is a representative gastrointestinal immune disease, and casued by an excessive immune response in the gastrointestinal tract. In recent studies, lactic acid bacteria which known as beneficial intestinal microbiota have protective effects on IBD by various mechanisms. Here, we evaluated the protective effect of Lactobacillus sakei CVL-001 isolated from korean tradtional food kimchi against dextran sulfate sodium-induced colitis in mice. First, administration of Heat-killed (HK) CVL-001 showed protective effects against DSS-induced colitis. Also, colonic mRNA expression levels showed that pro-inflammatory cytokine was downlregulated and anti-inflammatory cytokine was upregulated. Nucleotide-oligomerization domain 2 (NOD2) is well known as innate immune receptor and deeply related to IBD. We found that HK CVL-001 induced a NOD2-dependent regulatory phenotype in mouse bone marrow derived dendritic cell (BMDCs). Especially, we demonstrated that the expression of CD103, which is known as a marker of tolerogenic DC, was expressed mediated by NOD2. In vivo studies, colon administered with HK CVL-001 favoured the expansion of mucosal CD103 DCs and regulatory T cells. However, protective effect was abolished in NOD2 knockout mice. This results indicated that Heat killed L. sakei CVL-001 might be used therapeutic agent and strategy in IBD. This research was supported by the Mid-Career Researcher Programs (Grant No. 2022R1A2C2012287) of the National Research Foundation of Korea funded by the Ministry of Science and ICT (Information and Communication Technologies).

Complete tolerogenic response adjuvant stimulates Treg response to immunization

Abstract We identified the lowest composition necessary to generate a vaccination adjuvant that promotes a Treg response following immunization in mice and named it “complete tolerogenic adjuvant.” This novel adjuvant may allow us to utilize the well-established “antigen plus adjuvant” method of vaccination to induce Treg cell-mediated antigen-specific immunosuppression. The minimal composition is dexamethasone, rapamycin, and monophosphoryl lipid A with the following mass ratios: 8:20:3. We have demonstrated why immunosuppressive and immunogenic substances are both required for the formation of genuine adjuvants for Treg cells by dissecting the roles of each of these components during vaccination. Currently, research is ongoing in our group to begin to assess the usefulness of this adjuvant for antigen immunization in murine models of human autoimmune/inflammatory diseases. Specifically, we are evaluating the tolerogenic efficacy of this adjuvant for vaccination regimen targeting the apolipoprotein B antigen, using the ApoE−/− mouse model of atherosclerosis. These studies are expected to help drive the development of further, perhaps more effective, complete tolerogenic adjuvants that could be used to create a plethora of new, novel vaccines for treating immunological illnesses. Master program and NIH grant

Adjuvant skewing of host immune response to vaccination and subsequent infection with influenza virus

Abstract Despite being a vaccine-preventable disease, seasonal influenza is responsible for thousands of hospitalizations and deaths yearly. Currently licensed quadrivalent inactivated influenza virus vaccines (QIV) aim to induce neutralizing antibodies but are poor inducers of T-cell responses, which could be potentially overcome with adjuvants. We evaluated adjuvant skewing of host-immune responses after vaccination and its persistence after infection. Hereto, we compared two squalene-based adjuvants (AddaVax, and AddaS03) and IMDQ-PC, a lymph node-draining TLR7/8 agonist. All adjuvants resulted in enhanced antibody titers and protection against challenge with H1N1 influenza virus. Lung cytokine/chemokine profiling at 5 days post-infection showed that AddaVax results in higher levels of type 2 immunity-stimulating cytokines IL-4, -5, and -13 in the lungs, which is also reflected by a higher IgG1:IgG2a ratio. Meanwhile, IMDQ-PC showed higher serological IgG2a titers after vaccination and higher levels of IFNγ in the lungs after challenge. To investigate the role of the myeloid compartment in adjuvant skewing of host responses, we examined adjuvant-mediated skewing of bone-derived macrophages in-vitro using Arginase I and iNOS as M2 or M1 markers, respectively. We also pre-licensed macrophages with IL-4 and IFNγ to evaluate adjuvant-intrinsic skewing effects. Licensing was required to induce M1 or M2 phenotypes by squalene-based adjuvants, whereas IMDQ-PC skewed towards M1 phenotype irrespective of licensing. Correlating with the observed type I immune host response, IMDQ-PC induced iNOS more robustly with IFNγ than the other adjuvants. And, AddaS03 induced the highest levels of Arginase I in the IL-4 group. This study was also partly funded by CRIPT (Center for Research on Inuenza Pathogenesis and Transmission), a NIH NIAID funded Center of Excellence for Influenza Research and Response (CEIRR, contract number 75N93021C00014), by the NIAID funded SEM-CIVIC consortium to improve inuenza vaccines (contract number 75N93019C00051), to AG-S. By NIH/NIAID HHSN272201400008C (L.C.), and NIH/NIAID R21AI151229 and R01AI160706 (M.S.). B.G.D.G. acknowledges funding from the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation program (grant N 817938).

The role of purine nucleotide metabolism in cardiac failure

Normal cardiac function requires ATP to be synthesized at a sufficient rate to drive the molecular processes underlying electromechanical coupling. Myocardial metabolic dysfunction observed during heart failure leads to reductions in the chemical potential at which ATP is synthesized. We have observed reductions in oxidative capacity, and total adenine nucleotide (TAN) levels in a rat aortic constriction (TAC) induced heart failure model that parallel changes seen in large mammal models and humans with heart failure. Computer simulations of cardiac mechano-energetics also predict a marked increase in inorganic phosphate levels in diseased hearts compared to healthy control hearts. Furthermore, simulations predict that restoring the impaired energetic dysfunction of TAC rats improves systolic function. Metabolomic and proteomic analysis reveal increases in purine degradation by-products and AMP deaminase 3 (AMPD3) levels in failing versus control hearts. AMPD activity has been hypothesized to play a role in skeletal muscle in type 2 diabetes, however, there is still a need to decern the role, if any, of AMPD3 in the heart and in failure. A cardiac ventricle specific AMPD3 conditional knock out (cKO) rat model was developed to test the hypothesis that decreasing AMPD3 activity in the failing heart will lead to a preservation of TAN and improvement in cardiac mechanical function. Cardiac hypertrophy and decomposition were induced by placing a clip (TAC) in three-week old female and male rats. In the cKO model, AMPD3 exon 6 is flanked by loxP sites, and are either heterozygous for cre recombinase driven by the cardiac myosin light chain promoter (cre+/-) or lack the cre enzyme (cre-/-). Tamoxifen was administered to all rats both pre-surgery and post-surgery to induce AMPD3 knock down. Echocardiography was performed at 2- and 18-weeks post-surgery. At 18 weeks post-surgery, hearts were harvested for mitochondrial capacity, metabolite measurements, AMPD activity, and protein and mRNA measurements. Knock down efficiency in cre +/- animals compared to cre -/- was assessed based on enzyme activity, mRNA expression, and protein abundance. There is a 45% decrease in total AMPD activity in the AMPD3 cKO rats, where measured AMPD activity reflects the sum activity of all present AMPD isoforms. Additionally, there is 52% decrease in protein abundance and a 72% reduction in AMPD3 mRNA expression. Preliminary observations reveal significant differences in oxidative capacity and in cardiac function between TAC and sham rats, consistent with the hypothesis that knock down of AMDP3, an enzyme that is upregulated during TAC induced heart failure in rodents, results in preservation of myocardial nucleotide pools and improve cardiac mechanical function. Research is supported by the NIH grants HL144657 and F31-HL154605 (RL). We are grateful for support from the post-baccalaureate research program (2R25GM086262-09), the Systems and Integrative Biology (T32-GM008322) training grant and the Cardiovascular Research and Entrepreneurship training grant (T32-HL125242) and the University of Michigan Rackham Merit Fellowship. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Cryptic MHC-E epitope from influenza elicits a potent cytolytic T cell response

Abstract Non-classically-restricted T cells have shown remarkable therapeutic activity in a variety of disease settings; most notably, CD8 T cells restricted to MHC-E (Qa-1 in mice; HLA-E in humans) have proven capable of eliminating a primate lentivirus infection. Here, we used mass spectrometry to identify a Qa-1-restricted 9-mer peptide, named M-SL9, that elicits a strong CD8 T cell response in mice infected with influenza A virus. The M-SL9 specificity accounted for roughly 5–10% of all lung CD8 T cells, approaching the frequency of classically restricted CD8 T cell responses to flu. A broad investigation of the M-SL9-specific T cells showed that they are generally similar in phenotype to classically restricted CD8 T cells, including strong co-expression of type 1-associated cytokines (polyfunctionality) and cytolytic markers; however, M-SL9-specific T cells were unique in that they were anatomically restricted to the lungs and were not present in blood and spleen like other specificities. Intriguingly, the M-SL9 epitope derives from a previously unknown translation product encoded by the +1 alternative reading frame of the matrix 1 (M1) protein-coding sequence. Presentation and immunogenicity of M-SL9 were completely dependent on ribosomal initiation at the second AUG codon of the matrix gene segment. Analysis of potential precursors suggests that M-SL9 is processed from a novel 16-mer peptide that is not required for viral replication in mice. These discoveries imply that unconventional epitopes may drive a large fraction of physiologically relevant T cell responses, and M-SL9 presents a highly tractable system in which to dissect the mechanisms governing MHC-E-restricted T cell responses and their protective effects. Supported by Cancer Research Institute Irvington Fellowship, Roy and Diana Vagelos Molecular Life Sciences Program and College Alumni Society Research Grant from the Univ. of Pennsylvania, and NIH grants R01 AI146101, R01 AI153064, R01 AI046709-18S1, R01 AI046709, R01 AI125524, and R21 AI153978.

Multi-omic analysis of the cardiac cellulome defines a vascular contribution to cardiac diastolic dysfunction in obesity

Coronary microvascular dysfunction (CMD) is associated with cardiac dysfunction and predictive of cardiac mortality in obesity, especially in females. Emerging evidence suggests development of heart failure with preserved ejection fraction in females with CMD and that mineralocorticoid receptor (MR) antagonism may be more efficacious in obese female, versus male, HFpEF patients. Accordingly, we examined the hypothesis that smooth muscle cell (SMC)-specific MR deletion prevents obesity-associated coronary and cardiac diastolic dysfunction in females. Obesity was induced in female mice via western diet (WD) feeding alongside littermates fed standard diet. Initial studies revealed that global MR blockade with spironolactone prevented impaired coronary vasodilation and diastolic dysfunction in obese females. Importantly, specific deletion of SMC-MR similarly prevented obesity-associated coronary and cardiac dysfunction. Cardiac gene expression profiling suggested reduced cardiac inflammation in WD-fed mice with SMC-MR deletion independent of blood pressure, aortic stiffening, and cardiac hypertrophy. Further mechanistic studies utilizing single-cell RNA sequencing of non-cardiomyocyte cell populations revealed novel impacts of SMC-MR deletion on the cardiac cellulome in obese mice. Specifically, WD feeding induced inflammatory gene signatures in multiple non-myocyte populations (B/T cells, macrophages, and endothelium), independent of cardiac fibrosis, that was prevented by SMC-MR deletion. Further, SMC-MR deletion induced a basal reduction in cardiac mast cells and prevented WD-induced cardiac pro-inflammatory chemokine expression and leukocyte recruitment. These data reveal a central role for SMC-MR signaling broadly impacting the cardiac cellulome and underlying obesity-associated coronary and cardiac dysfunction thus supporting the emerging paradigm of a vascular origin of cardiac dysfunction in obesity. Supported by NIH R01 HL136386 (Bender), NIH R01 HL119290 (Jaffe), the University of Missouri Research Core Facilities Grant Program (Bender), NHMRC Ideas Grant GNT1188503 (Pinto), VA Merit I01 BX004220 and RCS IK2 6004016 (Chandrasekar). The work was also supported by resources and the use of facilities at the Harry S. Truman Memorial Veterans Hospital in Columbia, MO. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

The LSU AgCenter Healthy Communities Initiative: Community-Participatory Policy, Systems, and Environmental Change

The LSU AgCenter Healthy Communities Initiative: Community-Participatory Policy, Systems, and Environmental Change

Population Health Improvement Awards: Supporting Community and Academic Capacity to Partner in Research and Improve Population Health.

The Duke Clinical and Translational Science Institute Community Engaged Research Initiative began its Population Health Improvement Awards grant program in 2017. This program builds community-engaged research capacity by promoting the formation of community-academic research teams, educating researchers about equitable partnerships, and empowering community members and organizations to access academic research resources. With a focus on community-identified priorities, the program purposefully engages local communities in an enterprise that has traditionally labeled community members as "participants" rather than "partners." Key elements of the program include innovation, relationship building, and power sharing; education and research system navigation; iterative adaptation using the Plan-Do-Study-Act framework; and continual program improvements based on applicant feedback to ensure that the program becomes a national leader in funding local community-engaged research partnerships.

ACADEMIC MOBILITY: EUROPEAN AND ORIENTAL VECTORS OF COOPERATION

The article deals with the issues related to the international academic mobility. Some key notions within the academic sphere have been specified (academic mobility, academic integration, academic normative documents). The author outlines the European and Oriental vectors of academic cooperation (involvement of the academic staff and students into the programmes of international advanced training and / or internship; joint participation in scientific grant programmes; international co-supervision of Bachelor, Master, PhD research).

Special Issue Introduction: Demystifying Academic Understandings of Critical Race Theory to Transform Discourse With Policy Makers

In this introduction to the special issue, we describe the socio-political context that spurred our efforts to disrupt the anti-critical race theory (CRT) that has seeped into popular and political conversations about the US educational system. Noting the lack of preparation educational researchers have for sharing their nuanced, academic CRT understandings to policy-making audiences, we endeavored to address this issue. With use of funding support provided by the Spencer Foundation’s Conference Grants Program and in conjunction with Penn State University’s Research-to-Policy Collaboration, we identified a panel of CRT experts, facilitated their training in translating research into policy factsheets, and hosted a Day on the Hill conference during which the factsheets were shared with policy-making professionals. This introduction provides additional details about our process and about the resulting special issue which pairs together two written products covering the same CRT topic from each author, one written in the format of a traditional academic essay and the other as a two-page factsheet. Taken together, we hope that the paired publications serve as an example for how education researchers might demistify academic understandings of CRT in order to transform discourse with policy makers.

Effect of Rebreathing-Induced Intermittent Hypoxia on Glucose Tolerance in Adults with Type 2 Diabetes

Hypoxia stimulates glucose uptake through an insulin-independent pathway. The aim of this study was to examine the acute effect of rebreathing-induced hypoxia on plasma glucose and insulin levels in adults with type 2 diabetes. We hypothesized that rebreathing-induced hypoxia would attenuate the increase in glucose and insulin levels during an oral glucose tolerance test. Nine individuals with type 2 diabetes (5 men, age: 53±10 years, body mass index: 35±7 kg/m2, HbA1c: 7.1±0.5%) visited the laboratory on two occasions. On both visits, a 2-hour, 75 g oral glucose tolerance test was conducted while simultaneously performing a rebreathing-induced hypoxia protocol (RIH) or breathing room air (Norm). Venous blood samples were collected 0, 30, 60, 90, and 120 min following ingestion of the high-glucose drink to measure plasma glucose and insulin levels. The rebreathing-induced hypoxia protocol consisted of two series of five 2-min rebreathing bouts in a low-volume, closed circuit system interspersed with two minutes of breathing room air. The first and second series of rebreathing bouts were performed within the first 30 min and 30-60 min after ingestion of the high-glucose drink, respectively. Rebreathing-induced hypoxia resulted in a nadir oxygen saturation of 88±4 % and a nadir fraction of inspired oxygen of 12±5%. Plasma glucose responses to the oral glucose tolerance test were not different between conditions, however, insulin levels were lower during rebreathing-induced hypoxia than normoxia (RIH vs. Norm: 0: 17±13 vs. 21 ±14; 30: 37 ±28 vs. 42±26; 60: 57±32 vs. 77±52; 90: 88±67 vs. 116±81; and 120: 114±96 vs. 136 ±111 ulU/ml, p=0.03), suggesting an acutely improved insulin sensitivity. Accordingly, the insulin area under the curve was lower during rebreathing-induced hypoxia than normoxia (124±84 vs. 157±102, p=0.02). In conclusion, exposure to short and intermittent rebreathing-induced hypoxia following the ingestion of a high-glucose drink acutely improves insulin sensitivity in adults with type 2 diabetes. Rebreathing-induced hypoxia could therefore represent a novel and simple strategy to improve glycemic control in individuals living with type 2 diabetes. College of Education Small Grants Program This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Investigating the metabolic changes that accompany skeletal muscle maturation

Skeletal muscle plasticity is imperative for our overall health but diminishes with inactivity and age. As we reach maturity, a shift in nutrient allocation from supporting skeletal muscle hypertrophy to adipogenesis occurs. This pivotal intersection in tissue growth marks the start of a metabolic decline in skeletal muscle and an accumulation of adipose tissue in inactive individuals. If left uncorrected, a loss of skeletal muscle metabolic flexibility can lead to a myriad of metabolic diseases. To define the metabolic changes in skeletal muscle that accompany maturation, samples were collected from the longissimus dorsi (LD, glycolytic muscle), latissimus dorsi (LAT, mixed muscle), and masseter (MS, oxidative muscle) at 20, 53, 87, 120, and 180 days of age from DNA 600 x 241 (DNA Genetics) castrated male pigs that weighed an average of 5.7, 20.8, 42.2, 83.4, and 130.5 kg, respectively. These ages were selected to correspond to tissue growth phases characterized by vital organ growth, intensive muscle hypertrophy, cessation of muscle hypertrophy, and the beginning of unrestricted adipose deposition. Samples were analyzed to determine the abundance of key metabolic enzymes through Western blotting and assess mitochondrial content through real-time PCR. Glucose-6-phosphate dehydrogenase decreased at 87, 120, and 180 d in MS compared to LAT and LD ( P < 0.01), which suggested glycolytic muscles increased nutrient allocation to the pentose phosphate pathway at this time whereas oxidative muscles did not. Moreover, pyruvate dehydrogenase ( P < 0.01) and citrate synthase ( P < 0.01) increased at 120 d, which indicated an increase in metabolites entering oxidative pathways. This increase was accompanied by an increase in mitochondrial DNA at 87 and 120 d ( P < 0.01). Further, glutamic-oxaloacetic transaminase 2 increased between 20 d to 120 d ( P < 0.01) indicating amino acid synthesis peaks at 120 d. These findings have established a metabolic fingerprint associated with the termination of muscle growth and the beginning of adipose tissue growth, which provides the framework for uncovering the regulatory mechanism that modulates nutrient allocation between muscle and fat. Elucidating this mechanism will aid in the discovery of metabolic interventions to combat metabolic diseases. Kansas State University Global Food Systems Seed Grant Program This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

0933 Sleep Specialists in Federally Qualified Health Centers: A Gap in Sleep Health Care Potentially Contributing to Sleep Disparities

Abstract Introduction Poor sleep is associated with adverse cardiometabolic health outcomes and increased risk of mortality. Although primary care providers recognize the importance of sleep, they report low comfort levels in diagnosing, treating, and managing sleep disorders; lack of clinical knowledge and skills is the most common barrier to recognizing insomnia in family practice. Furthermore, minoritized racial/ethnic groups are disproportionately affected by poor sleep health, likely due in part to structural and other forms of racism that result in limited access to sleep healthcare. Health Resources & Services Administration (HRSA) has a variety of awards that support scholarships and loan repayment programs at colleges and universities to meet critical workforce shortages and diversify the health professions. We assessed the distribution of sleep specialists among clinicians who participated in a HRSA grant program because they tend to work for health care organizations that serve persons from lower socioeconomic and diverse racial-ethnic backgrounds. Methods Publicly available data were obtained from the HRSA data warehouse. Data were available for 44,199 clinicians (49% MD/DO and 16% Nurse Practitioner) who participated in a HRSA grant program between 2011 and 2021. We report data for those clinicians who endorsed sleep medicine as their subspecialty. Results Of the 44,199 clinicians, 27 (0.06%) physicians trained in a medical subspecialty of sleep medicine, and the majority (52%) were in Michigan. The remainder of sleep medicine specialists completed their program in Pennsylvania (33%), Ohio (7%), Minnesota (4%), and California (4%). Only the participants in the Children's Hospital Graduate Medical Education program designated their subspecialty as sleep medicine. None of the sleep medicine specialists currently practice in Health Professional Shortage Areas (HPSAs) or Medically Underserved Areas/Populations (MUA/P), and 1 physician currently practices in a rural area. Conclusion A marked paucity of trained sleep specialists in HRSA grant programs and an absence of such practitioners in HSPAs underscores the disparities in access to sleep specialty care. Initiatives are needed to incentivize sleep specialists to serve in under-resourced communities as well as to implement programs training primary care practitioners working in these areas to deliver sleep health care, supported by sleep specialists. Support (if any) Divisions of Intramural Research: NIMHD/NIEHS, NIH

Maine’s Clean Water Infrastructure: Transformative Power and Ongoing Needs

"Maine’s clean water infrastructure, particularly municipal and quasi-municipal wastewater treatment facilities, was largely built after the establishment of the CWA when federal grant funding via the Environmental Protection Agency (EPA) Construction Grants Program was made available on a large scale. Clean water infrastructure is often out of sight and out of mind for most people, but the maintenance and upgrading of this essential infrastructure is critical to protect water quality and public health and to support economic growth."

Direct measurement of human skeletal muscle specific tension and fiber length

Skeletal muscle’s isometric contractile properties are one of the classic structure-function relationships in all of physiology allowing for extrapolation of single fiber mechanical properties to whole muscle properties based on the muscle’s optimal fiber length and physiological cross-sectional area (PCSA). However, this relationship has only been validated in small animals and then extrapolated to human muscles which are much larger in terms of length and PCSA. We leveraged a unique surgical technique in which an intact, living, human gracilis muscle is transferred from the thigh to the arm, restoring elbow flexion after brachial plexus injury. During this surgery we directly measured subject specific gracilis muscle force-length relationship in situ and properties ex vivo. We hypothesized that the human gracilis would have a functional fiber length of about 23 cm, based on the anatomical literature, and a specific tension of 225 kPa, based on the physiological literature. Data are provided as mean±standard deviation of the mean. Twelve subjects (7 males, average age 54±12 years). In situ passive and active tension were directly measured using a buckle force transducer (BFT) placed on the distal gracilis tendon. Passive tension was measured in each of four joint configurations (JC) prior to muscle stimulation, and, to produce active tension, the anterior branch of the obturator nerve was stimulated. The muscle was progressively lengthened by extending the knee and abducting the hip. Since we could not measure optimal fiber length directly in our subjects during isometric testing we used a functional surrogate for optimal fiber length, the full-width at half-maximum (FWHM) defined from their force vs. length relationship. The FWHM is the width of the active force vs. muscle length curve at half the maximum force. A linear relationship between FWHM and optimal fiber length has been established across several mammalian species and muscles. Thus, to calculate FWHM, we fit each patient’s calculated fully active force vs. muscle length data to a quadratic curve and then calculated width at half the calculated maximum force from the equation. Optimal fiber length was then calculated from the previously established linear relationship between FWHM and optimal fiber length. Each subject’s PCSA was calculated from their muscle volume and optimal fiber length.From these experimental data we established a human muscle fiber-specific tension of 171±84 kPa (n=12). We also determined the average gracilis optimal fiber length was 12.9±3.3 cm (n=12). Surprisingly, these fiber lengths were about half of the previously reported optimal fascicle lengths of 23 cm. Thus, the long gracilis muscle appears to be composed of relatively short fibers acting in parallel that may not have been appreciated based on traditional anatomical methods. We thus suggest that the value of 171 kPa be considered the best current estimate of human muscle specific tension. This work was supported the Department of Veterans Affairs grant 1 I01 RX002462 and, in part, by Research Career Scientist Award Number IK6 RX003351 from the United States (U.S.) Department of Veterans Affairs Rehabilitation R&D (Rehab RD) Service. Pilot projects leading to this protocol were supported by the Shirley Ryan AbilityLab Catalyst Grant Program, National Science Foundation Graduate Research Fellowship under Grant No. 1255833 and the Mayo Clinic Graduate School. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

High fat diet feeding induces brain mitochondrial dysfunction in female Ossabaw swine

Study objective: Emerging evidence now suggest reduced mitochondrial function as a key initiator in Alzheimer’s disease (AD)-like pathology increasing the need for models of AD that display mitochondrial dysfunction. Dyslipidemia is a common aliment from overconsumption of a high fat diet (HFD) that increases the risk of AD and may contribute to mitochondrial dysfunction. Rodent studies utilize HFDs to induce brain pathology resembling that of AD. Unsurprisingly, rodent studies are rarely recapitulated in humans, which leads to a very low clinical success rate and highlights the need for more translatable models to study AD-like pathology. Swine seem to share many characteristics of humans and might offer a more translatable model to study HFD induced pathology. Thus, the objective of this study was to determine if HFD feeding in swine could offer a more translatable model to study mitochondrial dysfunction during neurodegeneration. Hypothesis: 6-months of HFD feeding will decrease indices of mitochondrial function and biogenesis in female Ossabaw swine. Methods: To characterize the effects of a HFD on brain mitochondrial health in a highly translatable swine model, high resolution respirometry measurements were taken from isolated brain mitochondria from in female Ossabaw swine fed a HFD for 6-months. Additionally, indices of mitochondrial biogenesis in whole tissue was measured (transcriptional markers of biogenesis and citrate synthase activity). Results: HFD feeding significantly increased body weight (p<0.05) with trending increases in blood glucose (p<0.10). Maximal uncoupled mitochondrial respiration was significantly decreased (p<0.05) with state 3-complex I and state 3 complex I & II trending down in the dorsal hippocampus (DH) after HFD diet feeding (p<0.10). Additionally, state 2 respiration was significantly decreased in the prefrontal cortex (PFC) after HFD feeding (p<0.05). HFD feeding increased transcriptional markers of mitochondrial biogenesis in the DH and PFC (p<0.05), without altering whole homogenate citrate synthase activity in either brain region. Conclusions: These data indicate HFD feeding in Ossabaw pigs could offer a potential model to study pathologies relating to mitochondrial dysfunction during neurodegenerative process. Additionally, the DH was found to be more vulnerable to HFD-induced perturbations, evident by the exacerbated deficits in mitochondrial respiration in this brain region. Finally, these data suggest HFD-induced increase in transcriptional markers of mitochondrial biogenesis may be a compensatory response to HFD-induced mitochondrial respiration deficits. However, citrate synthase activity was not different between groups indicating this upregulation of transcriptional markers was unable to translate to an increase in mitochondrial content in the DH and PFC. The project was supported by the MU Office of Postdoctoral Education Research Grant Program. Additional funding was also generously provided Scott Rector. Images in figure 2 adapted and modified from “Krebs cycle template”, by BioRender.com (2022). Retrieved from https://app.biorender.com/biorender-templates . This work was also supported with resources and the use of facilities at the University of Missouri and Harry S. Truman Memorial Veterans Hospital in Columbia, Missouri. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

SIGHPC Travel Grantees Broaden Horizons at SC22

For SC22, the SIGHPC Student Travel Grants Program helped seven students attend the conference in person. Each grantee received travel reimbursement ($600 for North American recipients, $1,200 for those studying elsewhere) and complimentary registration. In addition, the program connects each student with a mentor while attending the conference.

Impaired Erythropoietin Response to Intermittent Hypoxia in Patients with Type 2 Diabetes

Patients with type 2 diabetes (T2D) exhibit, on average, a 20% decline in maximal oxygen consumption when compared to healthy adults. Hemoglobin mass strongly correlates to maximal oxygen consumption. A reduced total blood volume has been observed in patients with T2D, suggesting that a reduced hemoglobin mass contributes to the decreased maximal oxygen consumption in this population. Hypoxia stimulates the release of erythropoietin (EPO), the hormone regulating red blood cell production. We previously showed that intermittent hypoxia, consisting of alternating short bouts of breathing hypoxic and normoxic air, increases EPO levels. Thus, the objective of this study was to determine the effect of a single session of intermittent hypoxia on serum EPO levels and hemoglobin mass in patients with T2D. We hypothesized that a single session of intermittent hypoxia would raise serum EPO levels and lead to an increase in hemoglobin mass in patients with T2D. Ten patients with T2D (4 women, age: 53 ± 10 years, body mass index: 36.2 ± 8.5 kg/m2, HbA1c: 7.2 ± 1.2%) were exposed to an intermittent hypoxia protocol consisting of eight 4-min cycles at a targeted oxygen saturation of 80% interspersed with normoxic cycles to resaturation. Air was made hypoxic by titrating nitrogen into a breathing circuit. Pulmonary gas exchange, oxygen saturation, and hemodynamics were continuously measured throughout the protocol. EPO levels were measured before and 4.5 hours after the beginning of the protocol. Hemoglobin mass was assessed via carbon monoxide rebreathing before and seven days following intermittent hypoxia. Intermittent hypoxia lowered oxygen saturation (­­97 ±­ 2 to 81 ± 2%, p<0.01), which resulted in a lower fraction of inspired oxygen (20.8 ±­ 0.1 to 11.1 ± 1.0%, p<0.01). There was no significant change in EPO levels following exposure to intermittent hypoxia (11.9 ± 5.3­ to 12.1 ± 4.3 mU/ml, p=0.83). There was also no change in hemoglobin mass in response to intermittent hypoxia (864 ± 152­ to 850 ± 150 g, p=0.64). Intermittent hypoxia did not affect mean arterial pressure (94 ± 5 to 97 ± 7 mmHg, p=0.18) but increased cardiac output (9.1 ± 2.7 to 9.8 ± 2.8 L/min, p=0.03) due to an increase in heart rate (78 ± 9 to 84 ± 10 bpm, p<0.01). In conclusion, a single session of intermittent hypoxia did not increase serum EPO levels or hemoglobin mass in patients with T2D. These findings suggest an impaired EPO response to decreased oxygen levels in patients with T2D, which may contribute to the reduced hemoglobin mass and total blood volume observed in this population. College of Education Small Grants Program This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

β-catenin inhibition using XAV939 nanoparticle promotes immunogenic cell death and suppresses tumor progression

Abstract Although the incidence of conjunctival melanoma is rare, it is a highly metastatic tumor with high recurrence and mortality rates. The current conjunctival melanoma treatment strategies, including surgical resection combined with chemo- or cryotherapy, have limited usefulness because of systemic side effects, recurrence, and metastases. Tumor-intrinsic Wnt/β-catenin pathway mediates immune exclusion in the tumor microenvironment (TME) and promotes tumorigenesis. Previous studies have shown that XAV939, a potent tankyrase inhibitor, can be used for the pharmacological inhibition of β-catenin. In this study, using tocopheryl polyethylene glycol 1000 succinate (TPGS)-based nanoparticle formulation for XAV939, we show that β-catenin inhibition suppresses melanoma cell proliferation and migration. Additionally, XAV939 nanoparticles promoted immunogenic cell death (ICD) of tumor cells with a significant extracellular release of ICD molecules, including HMGB1, calreticulin, and ATP. Further, using a mouse model of conjunctival melanoma, we show that β-catenin inhibition significantly suppresses conjunctival melanoma progression. Collectively, the local delivery of β-catenin represents a novel and safe approach to promoting ICD and activating anti-tumor immunity for combinatorial immune checkpoint inhibition therapies against vision and life-threatening ocular melanomas. This study was supported by research funding from the Intramural Grants Program, Auburn University Research Initiative in Cancer (AURIC), and start-up funds from the Department of Pathobiology, College of Veterinary Medicine, Auburn University.

Single-cell atlas of the liver myeloid compartment with cure of chronic viral hepatitis

Abstract Background/Aims: Chronic viral infections present serious public health challenges. Direct-acting antivirals (DAAs) are now able to cure nearly all patients infected with the hepatitis C virus (HCV), representing the only cure of a human chronic viral infection to date. DAAs provide a valuable opportunity to study immune pathways in the reversal of chronic immune failures in humans. Methods: To leverage this opportunity, we used plate-based single-cell RNA-seq (scRNA-seq) to deeply profile myeloid cells from liver fine needle aspirates in HCV patients before and after DAA treatment. We characterized liver neutrophils, eosinophils, mast cells, conventional dendritic cells (cDCs), plasmacytoid DCs, classical/non-classical monocytes, and macrophages, and defined fine-grained subpopulations of several cell types. Results: We discovered cell-type-specific changes post-cure, including an increase in MCM7+STMN1+proliferating CD1C+ cDCs, which may support restoration from chronic exhaustion. We observed an unexpected inverse relationship between pre-treatment viral load and post-cure ISG expression in each cell type, revealing a link between viral loads and sustained modifications of the host’s immune system. We found an upregulation of PD-L1 expression in ISG-high neutrophils, pinpointing cell subpopulations crucial for immune regulation. We identified three recurring gene programs shared by multiple cell types, distilling core functions of the myeloid compartment. Conclusion: This comprehensive scRNA-seq atlas of human liver myeloid cells in response to a cure of chronic viral infections reveals principles of liver immunity and provides immunotherapeutic insights. The single-cell sorting was done at the MGH Flow and Mass Cytometry Research Core, which was supported by NIH Shared Instrumentation program grants 1S10OD012027-01A1, 1S10OD016372-01, 1S10RR020936-01, and 1S10RR023440-01A1. Additional sorting was done at the Immunology Core Facility of the Ragon Institute, which is part of the Harvard University Center for AIDS Research (CFAR), an NIH funded program (P30 AI060354).