bluuebunny/crossref_metadata_2025_split huggingface dataset:
Aged HIV-1 Tg26 Transgenic Mice display Vascular Dysfunction with No Alterations in Neurocognitive Function or Cerebral Blood Flow
Authors: Khakina Beryl , Kress Taylor C. , Rosewater Cody , Bosomtwi Asamoah , Kennard Simone , Belin de Chantemèle Eric J. 
Published: May 2024

Although combination antiretroviral therapy (cART) has markedly reduced HIV mortality and increased life expectancy, people living with HIV (PLWH) develop premature cognitive and brain aging, classified as HIV-associated neurocognitive disorder (HAND). Vascular dysfunction and expression of HIV-derived proteins have independently been associated with cognitive impairment without causality being established. Thus, we investigated the role of HIV viral proteins in endothelial dysfunction and cognitive impairment in an aged mouse model of HIV. We used 18-month-old Tg26 mice and their wild-type littermate controls. Tg26 mice express 7 out of the 9 HIV viral proteins under the control of the long-term repeat promoter and have previously been shown to exhibit signs of cognitive impairment at eight months of age. Conduction of concentration-response curves to acetylcholine in mesenteric arteries using wire myography revealed impaired endothelium-dependent relaxation in Tg26 mice compared to WT mice. Measurements of brain perfusion with arterial spin labeling magnetic resonance imaging showed no differences in the whole brain and hippocampal cerebral blood flow. In addition, there were no changes in the hippocampus volume, which has been reported to be reduced with cognitive impairments. Two methods were employed to assess the cognitive abilities of the mice: Morris water maze (MWM) and the IntelliCage system. MWM requires human handling while the IntelliCage system is fully automated enabling the testing of mouse cognitive function during their active period and independently of human handling. Tg26 and WT mice underwent ten days of MWM testing and eight days of IntelliCage testing to assess spatial learning and memory. Tg26 mice showed no significant differences in latency to platform, distance swam to platform, or swim velocity when compared to their WT littermates. The IntelliCage results showed no differences in total visits, total visits with nose pokes, or total licks during the reversal learning period. Additionally, there were no significant differences in percent side error or place error indicating that cognitive function is not impaired in Tg26 mice. The Y-maze test further showed no differences in spontaneous alteration, a measure of spatial working memory. Altogether, our data shows that although 18-month-old female Tg26 mice develop endothelial dysfunction, they display no alterations in cerebral blood flow and no cognitive impairment indicating that systemic viral protein expression and vascular dysfunction might minimally contribute to neurocognitive impairment in HIV. R01HL130301 & R01HL155265 (E.J.B). This is the full abstract presented at the American Physiology Summit 2024 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.
Don’t lose your transporters! Optimizing renal transporter recovery after kidney freezing
Authors: Hartman-Houstman Hannah , Ralph Donna , Moronge Desmond , Faulkner Jessica , Sullivan Jennifer , McDonough Alicia 
Published: May 2024

Renal ion transporters are critical to maintain fluid and electrolyte homeostasis and are primary targets for (patho)physiology studies and drug discovery. Transporter abundance and covalent modifications are determined by immunoblots. Our standard protocol is quick kidney removal and homogenization in buffer containing 0.2 mM PMSF, 0.9 ug/ml aprotinin, and Sigma P0044 phosphatase inhibitors followed by isolation of a low speed supernatant (the homogenate). Collaborations with other labs requires shipping frozen kidneys. We tested whether flash freezing (FZ) Sprague Dawley (SD) kidneys impacts ion transporter recovery compared to quick removal without freezing (QR) defined as 100% recovery; assessed by immunoblot with specific antibodies. FZ and ~ 3 d storage before homogenization in inhibitors significantly reduced recoveries of proximal tubule (PT) NHE3 and megalin to 20% and Cldn2 to 40%. Medullary NHE3 and NKCC2 remained at 100%, DCT/CD proteins were reduced ~ 10-60% after FZ vs. QR. Renal perfusion pre-FZ further reduced recovery of all transporters vs. non-perfused kidneys. To test whether Fz activates proteases not inhibited in our standard buffer, Roche protease inhibitor (minitabs) were added to homogenization buffer. Adding minitabs to FZ kidneys increased NHE3 recovery to 85% and Cldn2 recovery to 100% of QR, NKCC2 and ENaC-beta to 85% and NCC to100%. Lastly, we tested, in male and female (M,F) SD rats, whether minitab addition to QR renal cortex homogenates followed by Fz prevents proteolysis vs. minitab addition to QR homog without Fz. In F, minitab addition with Fz preserved 100% recovery of transporters: NHE3, Cldn2, AQP1, megalin, NKCC2, NCC, ENaC-beta, Cldn-7 and AQP2. In M, minitab addition with Fz preserved 100% recovery of only AQP1, villin, Cldn7 and AQP2; all other M transporters exhibited ~40% less recovery. In conclusion, flash freezing kidney tissue differentially activates proteolysis of ion transporters - up to 80% in cortex and not at all in medulla - despite inclusion of standard inhibitors in homogenizing buffers. Strong sex differences are evident: addition of minitabs prevents proteolysis of F not M cortical ion transporters. Single cell ‘omics approaches may be useful to identify tubule and sex specific proteases. NIH: DK083785, DK134695. This is the full abstract presented at the American Physiology Summit 2024 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.
Development of CRISPR/Cas9-Based Mouse Models for X-linked Alport Syndrome
Authors: Decker Stephen , Davey Crystal , Cao Guoshen , Al-Rabadi Laith , Funai Katsuhiko 
Published: May 2024

Intro: Alport Syndrome (AS) is a rare hereditary disorder characterized by progressive renal impairment and sensorineural hearing loss, resulting from mutations in the genes that encode COL4 synthesis. X-linked Alport Syndrome (XLAS) is the most common form of AS, comprising an estimated 80% of all human AS cases (the others being various forms of autosomal recessive or autosomal dominant AS), and is caused by various mutations on the COL4A5 gene. However, to date, there are no animal models of XLAS. Methods: To facilitate comprehensive preclinical investigations and therapeutic developments, we established two novel mouse models of XLAS using CRISPR/Cas9 technology. Leveraging the robust C57Bl/6J mouse strain, we precisely introduced the L1655R and C1570S mutations, which represent the predominant human variants of XLAS observed in the Western United States. Discussion: Our innovative mouse models not only serve as invaluable tools for elucidating the underlying mechanisms of XLAS pathogenesis but also provide a platform for evaluating the effcacy of potential therapeutic interventions. The establishment of these faithful genetic replicas of human XLAS marks a significant advancement in the field, offering unprecedented opportunities for in-depth mechanistic studies and the development of targeted therapeutic strategies for this debilitating renal and auditory disorder. This research is supported by the National Institutes of Health under Ruth L. Kirschstein National Research Service Award 5T32DK091317 from the National Institute of Diabetes and Digestive and Kidney Diseases and the National Kidney Foundation of Utah and Idaho. This is the full abstract presented at the American Physiology Summit 2024 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.
Tissue dependence of variation in metabolic rates of harvester ants
Authors: Harrison Jon , Perl Craig , Haas James , Johnson Meredith , Johnson Robert 
Published: May 2024

Metabolic rates vary tremendously among individuals and across species, and the causes of this variation remain controversial and challenging to identify. We utilized an Agilent Seahorse system to measure the metabolic rates of brains, flight muscles, leg muscles, jaw muscles, digestive tract, fat and reproductive organs of harvester ants of different species, sex and caste, and compared these to whole body metabolic rates. Across Pogonomyrmex species varying an order of magnitude in body size, mass-specific brain metabolic rates declined dramatically in larger species, such that the brain accounted for 43% of whole body metabolism in the smallest species and 14% in the largest. In Veromessor pergandei, males have much higher mass-specific metabolic rates than queens or workers, with queens being slightly higher than workers. The tissue specific bases to metabolic rates varied with sex and caste. Males are "flying gonads", with flight muscle, fat and gonads accounting for 44, 25 and 20%, respectively, of resting metabolic rate, and tissue-specific rates were generally higher in males than females. For queens, which have by far the largest fat content, the fat body contributes the largest fraction of the total tissue metabolic rate (38%), with flight muscle, femoral muscle, brain and gut each contributing 13 – 15% of total cost. For workers, brains are the most important contributors to total tissue metabolic rate (38%), followed by fat body (24%) and gut (19%). Together these results demonstrate that variation in whole body metabolic rate within and across closely related species is strongly affected by size, sex and caste-specific differences in tissue function, likely associated with life history variation. Supported by NSF IOS-1953419. This is the full abstract presented at the American Physiology Summit 2024 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.
IGFBP3 regulates airway basal cell stemness and airway regeneration
Authors: Murthy Shubha , Pathmanathan Aparna , Caceres Adrian , Sease Rosemary , Calvert Ben , Marconett Crystal , Ryan Amy 
Published: May 2024

Basal Cells (BCs) are multipotent progenitor cells of the upper airway and critical in airway epithelial regeneration. Impaired self-renewal and differentiation capacity of BCs is a significant cause of chronic respiratory diseases. However, mechanisms mediating these changes remain largely unexplored. We, and others, have observed attenuated growth and differentiation of BCs after repeated cell divisions in vitro. To investigate mechanisms regulating BC stemness, we performed bulk RNA-sequencing to analyze transcriptomes of early and late passage BCs maintained in culture. Our analyses reveal significant differences in the expression of genes involved in IGF- and IGFBP3-dependent pathways that are known to modulate cellular growth and ROS generation. The role of these pathways in regulating BC stemness have not been systematically investigated. We hypothesized that IGFBP3 regulates BC growth and differentiation by modulating cellular redox status and DNA damage repair. Herein, we demonstrate that passage of non-diseased human BCs over 5 passages increases IGFBP3 transcript in 3 independent growth conditions. Interestingly, continued passage decreased IGFBP3 expression and significantly altered its intracellular localization. In contrast to low passage cells where IGFBP3 is largely cytosolic, high passage cells almost exclusively express IGFBP3 in their nuclei where it is known to participate in DNA repair. Consistent with this, we observed increased DNA repair gene expression in high passage cells coupled with significant reduction in thioredoxin levels. Together, these results suggest that serial passage of BCs causes oxidative stress-mediated BC exhaustion and altered IGFBP3 expression and localization. To investigate the role of IGFBP3 in BC growth and differentiation, we knocked down IGFBP3 expression and, as hypothesized, IGFBP3 knock down of low passage cells attenuated growth, BC markers, and DNA damage repair gene expression. In addition, differentiation to secretory, ciliated and club cells at the air-liquid interphase was significantly attenuated. In conclusion, our data support an IGFBP3-dependent regulation of BC stemness by preserving growth and differentiation. This study was supported by the NIH R01HL139828 (ALR) and Cystic Fibrosis Foundation grants FIRTH17XX0 (ALR), RYAN21XX0, FIRTH21XX0 (ALR, CNM). BCs for this study were provided though the Center for Gene Therapy Cell Core funded by the NIH:NIDDK. This is the full abstract presented at the American Physiology Summit 2024 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.
Determining optimal surveillance techniques for the microbial community on human skin using RNAseq data analysis
Authors: Bhargava Disha , Prokop Jeremy 
Published: May 2024

Background: The largest organ in the human body, the skin, serves as a physical barrier to the outside world. It also houses millions of bacteria, viruses, and fungi that protect us against invading pathogens or result in altered physiology and disease. These microorganisms can contribute to normal skin health or predispose individuals to pathogenic disorders. Surveillance and observation of the microbiome can give insight into identifying common taxa that compose our skin and indications of new or changing pathogens. However, to do this effciently, it is important to understand what can be detected from skin samples and the differences we may see using various collection methods. Methods: We used publicly available NCBI SRA RNAseq bioprojects that queried “skin,” processing 132 bioprojects with 8,274 samples. Using FastQC, Salmon, KRAKEN2, and MiCXR/Immunarch, we aimed to analyze the quality, human gene transcripts, infections, and B/T- cell responses, respectively. We aimed to identify what skin sampling method provides the best data on the skin microbiome, human gene mapping, and B/T-cell immune repertoire. This project is based on observational methods, not a specific hypothesis. Results: From the KRAKEN2 data, there were 14,383 viruses and 14,691 bacteria assessed through all “skin” BioProjects. From the extensive data pool, we identified unique BioProject signatures for protest (Leishmania), bacteria (Mycoplasma), and viruses (HSV1 dose dependencies, HPV in human warts, Chinese transmitted tick viruses, Human T-cell lymphotropic virus, and the first-ever human case of equine infectious anemia virus). The use of read mapping and coverage is critical to confirming the presence of foreign species, noted through the observation of T4 DNA ligase RNA presence prior to 2018 RNA sequencing. Once pathogens were detected, cellular response mapping was developed, yielding insights into the pathophysiology of multiple viral infections of the skin and sex differences. Conclusion: Detecting a wide variety of microorganisms on the skin can give insight into dermatological pathologies and skin physiology. Using the publicly available skin datasets, we see many unique stories of microorganisms in the skin. This big data mapping strategy highlights the importance of broad observational mapping, possible with cutting-edge sequencing tools. This work was supported by the MI-SAPPHIRE program of MDHHS and the CDC. This is the full abstract presented at the American Physiology Summit 2024 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.
Quantifying Prefrontal-Hippocampal Circuit Engagement Using Scalp EEG and the Paired Associates Learning Task
Authors: Besosa Cristina , Lovett Sarah , Kennedy Jack , Maurer Andrew , Burke Sara 
Published: May 2024

The hippocampus (HPC)-prefrontal cortical (PFC) circuit, critical for affective processing and cognition, is vulnerable in mental health disorders. Although disrupted HPC-PFC functional connectivity is a common feature of anxiety, schizophrenia, and autism, how local cellular interactions within this circuit manifest as large-scale temporal coordination remains unknown. The objective of this project is to provide novel mechanistic insights into disrupted neural coordination in mental illness by examining the relationship between invasive local field potential (LFP) recordings from the HPC and PFC, and scalp electroencephalogram (EEG) during the Paired Associates Learning (PAL) task. The PAL task measures visuospatial associative memory by requiring animals to learn the stimulus-location pairing of different images on a touchscreen. We have previously shown that manipulations that disrupt HPC-PFC coordination, such as traumatic brain injury and advanced age, lead to PAL performance impairments. Importantly, EEG recordings during PAL performance in human study participants have shown associations between behavioral deficits and altered neural dynamics in schizophrenia and anxiety. We hypothesize that simultaneous LFP and EEG recordings from rats performing the PAL task will reveal highly correlated theta power across brain regions, as well as significant phase relationships among frequencies when compared to rest periods. Preliminary data show correlated increases in theta power in both the LFP and EEG from HPC as well as PFC, and higher power in the theta frequency band during PAL compared to rest periods. Future analyses will determine phase coherence between frequencies within a signal as well as brain regions. Exploring the relationship between invasive neurophysiological measures of the LFP and non-invasive surface EEG signals in the context of a translatable behavioral paradigm could provide new physiological and behavioral biomarkers that may be capable of predicting HPC-PFC coordination and aid the development of therapies for treating mental health disorders. The Evelyn F. McKnight Brain Research Foundation and the NIH/NIMH (grant R01MH126236). This is the full abstract presented at the American Physiology Summit 2024 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.
Beclin-1-Dependent Autophagy Preserves Perivascular Adipose Tissue Function and Suppresses Hypertension Development in Hyperaldosteronism
Authors: Costa Rafael , Rodrigues Daniel , Gonçalves Tiago , Alves Juliano , Awata Wanessa , Singh Shubhnita , Bruder-Nascimento Ariane 
Published: May 2024

Hyperaldosteronism (HA), characterized by the independent overproduction of aldosterone (Aldo), leads to cardiovascular damage and perivascular adipose tissue (PVAT) dysfunction. Beclin-1 (Becn1) is a key protein regulating autophagy, a recycling process of unwanted components. Whether HA disturbs autophagic flux in PVAT via Becn1 is unknown. Thus, we hypothesize that Becn1-dependent autophagic flux will improve PVAT function and inhibit the genesis of hypertension (HTN) in HA. 10-12-weeks-old male C57BL/6J (WT) or Becn1 knock-in mice (Becn1Tg) mice were infused with aldosterone for 14 days (600ug/Kg/day via osmotic mini-pump). Vascular function was studied in PVAT-intact thoracic aortae and blood pressure was analyzed by radiotelemetry. Becn1Tg mice were protected from HA-induced HTN [mean blood pressure (mmHg) − before HA: 101.2±0.5 vs. after HA: 108.2±0.9]. HA disrupted PVAT autophagic flux characterized by accumulation of LC3A/B and p62 and reduction in Becn1 expression and activity. In terms of function, presence of PVAT decreased the contractile response in WT [PVAT (-): 137.2±1.9 vs. PVAT (+): 107.4±2.7, Emax (% KCl)], while HA induced loss of PVAT anti-contractile effect [PVAT (-): 159.9±2.9 vs. PVAT (+): 164.1±3.4]. Becn1Tg mice were protected from this effect. Furthermore, HA induced PVAT oxidative stress and inflammation, characterized by elevated TNF-α, IL-6, IL-1β, IL-17, and IFN-γ mRNA expression, and elevated reactive oxygen species (ROS), measured by lucigenin chemiluminescence. All these effects were prevented in Becn1Tg mice. Pharmacological induction of Becn1 activation, via TB-peptide (16mg/Kg/day for 7 days), improved PVAT function of WT mice with HA. To evaluate the molecular mechanisms, we treated brown adipocyte cells (BAC, WT1) with Aldo (100nM), which triggered a transient autophagic flux, depicted by an initial increase in LC3A/B (12 and 24 h), followed by a decrease at 48 and 96 h. Such effects were prevented by a selective mineralocorticoid receptor antagonist (Eplerenone, 1μM). Finally, Aldo induced BAC inflammation and oxidative stress, which were blunted by TB-peptide. In summary, our findings reveal that autophagy overactivation confers protection against HA-induced PVAT inflammation and dysfunction and HTN. Therefore, activation of Becn1 could be pharmacological tool to prevent HA-associated cardiovascular deleterious effects. FAPESP (2022/06639-2), NHLBI-R00 (R00HL14013903), AHA (CDA857268 and 23DIVSUP1069230), Vascular Medicine Institute, the Hemophilia Center of Western Pennsylvania Vitalant, and Children's Hospital of Pittsburgh of the UPMC Health System. This is the full abstract presented at the American Physiology Summit 2024 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.