bluuebunny/crossref_metadata_2025_split huggingface dataset:
Developmental Programming of Glomerular Podocyte Endowment
Authors: Bertram John , Cullen-McEwen Luise , Gazzard Sarah , Bertram michael , van der Wolde James , Walton Sarah , Moritz Karen , Haruhara Kotaro , Goncalves Gessica 
Published: April 2020

A range of perturbations to the maternal environment result in low nephron endowment in offspring, which is often linked to increased blood pressure and renal pathology in later life. Given that nephrogenesis in mammals ends shortly before or after birth, this nephron deficit is permanent. To date, very few studies have examined the morphology of nephrons in offspring born with low nephron endowment. Given the link between podocyte loss and glomerulosclerosis in adults, we examined whether maternal perturbations also result in low podocyte endowment in offspring, thereby rendering offspring at increased risk of adult disease.Three models of maternal perturbation were investigated: (1) gestational hypoxia in mice; (2) maternal low protein diet (LPD) in mice followed by postnatal hyperglycaemia; and (3) maternal low protein diet in rats followed by high fat feeding. Podocytes were counted at postnatal day 21 in whole glomeruli contained in tissue slices immunostained for specific podocyte markers, cleared and imaged with confocal microscopy.In all three models, total podocyte number per glomerulus was reduced in settings of maternal perturbation. In male mice exposed to hypoxia, podocyte endowment was 15% lower than controls (p<0.01), but no effect was observed in females; in male and female mouse LPD offspring, podocyte endowment was 15% (p<0.001) and 9% (p<0.05) lower than controls; and in male LPD rats (females were not studied), podocyte endowment was 15% (p<0.0001) lower than controls.This is the first report of developmental programming of low podocyte endowment. We have identified low podocyte endowment in two species, three models and both sexes. We are currently investigating whether this podocyte deficit is permanent, and the effects on renal function in later life.
Determining the Effects of Legionella pneumophila SidE Effector Proteins on Host Vesicular Trafficking
Authors: Favichia Michelle , Allgood Samual C. , Neunuebel M. Ramona 
Published: April 2020

Legionella pneumophila is an opportunistic intracellular pathogen responsible for Legionnaires’ disease in humans. Upon infection, L. pneumophila forms the Legionella‐containing vacuole (LCV), shielding itself from the host immune response. From the LCV, L. pneumophila uses a specialized secretion system to translocate over 350 effector proteins into the host cell. Effectors target and manipulate host pathways in order to optimize the host’s cellular environment to support L. pneumophila replication
(1). One important cellular process targeted during infection is endocytic recycling. Recycling pathways are generally categorized as slow or fast, depending on the time it takes for endocytosed cargo to return to the plasma membrane. Rab11 is a key regulator of the slow endocytic recycling pathway
(2) and it moves along microtubules partly through its interaction with Rab11‐FIP1A. It was previously shown that mutation of Rab11‐FIP1A disrupts transferrin recycling
(3). In the context of infection by L. pneumophila, our group has shown that transferrin recycling in human macrophages is inhibited early during infection. We hypothesize that the L. pneumophila effectors protein family SidE is at least partly responsible for this defect. Pull‐down assays revealed that interaction between Rab11 and Rab11‐FIP1A is disrupted in the presence of SdeC. In transiently transfected CHO cells, GFP‐Rab11‐FIP1A is found on vesicles in the perinuclear region. However, when GFP‐Rab11‐FIP1A and mCherry‐SdeC are co‐expressed in CHO cells, we observed that GFP‐Rab11‐FIP1A changed its localization to several enlarged structures. We are investigating whether SdeC’s ubiquitination activity is responsible for disrupting interaction of Rab11 with Rab11‐FIP1A. These results will help establish the effects of ubiquitination by SidE effector proteins on the slow recycling pathway.Support or Funding Information
Fibroblast Inflammasomes Promote Inflammation and Fibrosis in the Breast Tumor Microenvironment
Authors: Postma Makenzie 
Published: April 2020

Clinically, both incidence and mortality of breast cancer remain significant, despite progress in our understanding of cancer, and advances in treatments. Fibrosis is an independent prognosis marker of breast cancer, and cancer‐associated fibroblasts promote cancer growth. Whether inflammasome activities of fibroblasts critically influence inflammation and fibrosis remains unclear. Thus, we determined whether fibroblast inflammasome activation altered the secretions of pro‐inflammatory cytokines and fibrosis associated proteins. We assessed (1) the expression of the inflammasomes’ proteins and (2) the secretions of the pro‐inflammatory cytokines IL‐1ß and IL‐18 along with collagen I secretions by L929 fibroblasts in vitro. The effects of conditioned media collected from the aggressive 4T1 mammary tumor cells were also assessed. Our data indicate that L929 fibroblasts express the proteins associated with functional NLRP3 inflammasomes. Furthermore, following incubation with LPS, a classical inflammasome activator, L929 fibroblasts secreted significantly more IL‐1ß, matrix metalloproteinase 14, and collagen I. The effects of the NLRP3 inflammasome inhibitor MCC950 on L929 fibroblast secretions of pro‐inflammatory cytokines and pro‐fibrotic extracellular matrix proteins are also presented. These results (1) confirm the activity of inflammasome in fibroblasts, and (2) highlight the role of fibroblast inflammasomes in the generation of the pro‐inflammatory and pro‐fibrotic microenvironment in breast cancer.Support or Funding InformationMakenzie Postma is an Honors student enrolled in the Honors in Biological Sciences Program at UNC Charlotte. This research is supported by the University of North Carolina at Charlotte under the direction of Dr. Didier Dréau.
Type 1 Diabetes Results in Significant Purinergic Receptor Remodeling in Podocytes
Authors: Klemens Christine A. , Spires Denisha , Levchenko Vladislav , Palygin Oleg , Staruschenko Alexander 
Published: April 2020

P2 (purinergic) receptors have been proposed to be potential viable targets for the treatment of various cardiovascular and renal disorders. Previous studies in our lab have demonstrated that the predominant purinergic signaling pathway in healthy podocytes is through P2Y1 metabotropic receptors. It was also reported that expression of purinergic receptors in different tissues, including kidney, is dynamic and might be changed under pathological conditions such as diabetic nephropathy. We hypothesized that prolonged type 1 diabetes (T1D) would result in substantial changes to podocyte purinergic calcium signaling and result in a transition from predominantly metabotropic signaling through P2Y1 to increased ionotropic signaling through P2X receptors. To do this we utilized the established T1D model of streptozocin‐injected Dahl Salt Sensitive (SS) rats, and measured intracellular calcium ([Ca2+]i) signaling in podocytes from their freshly isolated glomeruli. We determined that in comparison to non‐diabetic rats, which have a rapid elevation of [Ca2+]i levels in response to ATP (40 μM) that returns to baseline within 2 min, podocytes from T1D rats had reduced peak response amplitudes (0.53 ± 0.04 vs. 0.34±0.02, p<0.001). Furthermore, podocytes of glomeruli isolated from diabetic rats had prolonged sustained elevation of [Ca2+]i level, which was not observed in non‐diabetic rats. Additionally, we used P2 specific antagonists to determine which receptors resulted in the altered calcium response curve in T1D podocytes. We found that pharmacological inhibition of P2X7 and P2X4 (10 μM A438079 and 5‐BDBD, respectively) altered the calcium curve dynamics, but did not significantly reduce the total [Ca2+]i level over a 5 min time period, as determined by area under the curve (AUC) measurements. Similarly, inhibition of P2Y1 (1 μM MRS2500) changed the [Ca2+]i curve response, but did not reduce the AUC. Combining both P2X7 and P2X4 inhibitors reduced the AUC; however, did not completely eliminate podocyte purinergic signaling (51.42 ± 5.52 vs 30.39 ± 5.00). Combined inhibition of P2X7, P2X4, and P2Y1 resulted in the near complete elimination of [Ca2+]i release in response to ATP (AUC, 51.42 ± 5.52 vs 10.57 ± 2.56, p<0.001). These results demonstrate that T1D increases ionotropic purinergic signaling in podocytes; however, does not eliminate metabotropic signaling. These studies establish the potential for targeting both P2X4 and P2X7 for therapeutic intervention in T1D.Support or Funding InformationNational Institutes of Health grants T32 HL134643 and R35 HL135749, and Department of Veteran Affairs I01 BX004024
Using VR‐Enriched Tasks (VRETs) to Promote Active Learning in Pre‐Clinical Gross Anatomy Education
Authors: Cheung Chun Chung , Cecot Tomasz Stanislaw , Tipoe George Lim , Yang Jian 
Published: April 2020

With the blooming growth of technological innovations, virtual reality (VR) is slowly being incorporated into anatomy education and has provided a brand‐new area of research interest for anatomy educators. It is paramount to make anatomy classes more active and student‐centred as that creates deeper understanding of the subject matter. The use of VR is not merely to create another atlas for students to browse in their free time. Rather, VR should be used to enhance active learning. For that, task‐based learning is believed to be an ideal approach to create an active and engaging learning environment. We aim to introduce VR into large classes of over 250 students and integrate it seamlessly into dissection classes. This pilot study looked at our students’ perception and satisfaction of a learning activity assisted with VR.Prior to the pilot session, three VR‐enriched tasks (VRETs) were written by two anatomy teachers. All tasks required participants to work in groups of minimum two people, one using the VR device while the other giving instructions. Two other anatomy teachers, who did not participate in task writing, were invited to review and perform the tasks to verify the tasks were of adequate difficulty and written with clear instructions. During the VRETs sessions, the participants were given an introductory demonstration and a warm‐up task before proceeding to the three VRETs. Each session lasted on average 60 minutes and participants were asked to fill out a 7‐point Likert scale questionnaire (7=strongly agree and 1=strongly disagree) at the end of the session.Two VRETs sessions were held with a total of 48 participants. 10 sets of VR devices were deployed and anatomy teachers and technicians were present to assist the participants. All participants were second year medical students at the University of Hong Kong and all completed and returned the questionnaire. The questionnaire was modified based on a recent publication on technology‐enabled active learning inventory. Overall, three participants reported to have previous VR experience. 4 items received an average score over 5.9, in which participants moderately agreed that VRETs gave them deeper insights to issues they did not think of; piqued their curiosity; act as extra support for learning anatomy and improved the overall learning experience. Participants also gave written feedback, which mainly focused on the steep learning curve of operating the VR devices.This pilot study showed that VRETs were well‐received by participants. However, extra training time should be given to users as many commented on the operational difficulty to novice users. Further study would be conducted to look at the impact of VRETs in learning and whether it is useful in creating an active learning environment. We would also look at how VRETs can be run in conjunction to dissection classes. IRB approval Ref: HKU/HA HKW IRB, Ref: UW 19‐277.
Sex Differences in Airway Inflammation and Structural Properties in a Mouse Model of House Dust Mite Challenge
Authors: Silveyra Patricia , Fuentes Nathalie , Montes Deborah T. , Spinelli Amy , Abraham Thomas , Purushothaman Indira , Chroneos Zissis 
Published: April 2020

BackgroundAsthma is a chronic airway disease that leads to compromised lung function and affects more than 300 million patients worldwide, leading to >180,000 deaths each year, many of which are avoidable with proper treatment and care. Despite significant advances in diagnostic and treatment of the disease, asthma continues to be a significant health problem and an economic burden. Epidemiological studies in adult populations have revealed sex differences in asthma morbidity, with higher incidence in females than males. The causes of this disparity are still unclear, but studies have shown that sex‐specific signaling pathways regulated by hormones contribute to differences in immune cell activation and airway reactivity in response to different environmental stimuli.MethodsWith the goal of understanding the mechanisms associated with the observed sex disparities in allergen‐induced asthma, we challenged both male and female adult C57BL/6J mice with a house dust mite (HDM) solution from two species, Dermatophagoides pteronyssinus and Dermatophagoides farinae. The protocol consisted of five consecutive intranasal challenges with 25μg of HDM or control (PBS) followed by two days off, repeated for five weeks. The asthma phenotype was assessed by testing airway hyperresponsiveness (AHR) to methacholine challenge, and airway inflammation was characterized by cell differential assessment of bronchoalveolar lavage fluid obtained from experimental animals. Lung tissue was harvested for inflammatory mRNA expression determinations and histology. Airway remodeling was characterized using the two‐photon imaging technique, second harmonic generation (SHG), on lung tissue sections to determine collagen content in mouse airways (Velocity, imaging quantification software).ResultsWe found that HDM promoted alterations in respiratory mechanics in challenged animals, including alterations in lung function measures that led to airway hyperresponsiveness (AHR) and increased parenchymal elastance in both males and females. While HDM‐challenged male mice showed higher AHR, airway collagen & elastin content, and BALF eosinophilia than females, challenged females displayed higher BALF neutrophilia and expression of type II pro‐inflammatory cytokines than males.ConclusionOur results indicate sex differences in both inflammatory and immune responses, as well as in airway structural changes induced by HDM exposure in mice. Characterization of the mechanisms leading to these sex differences will provide better insight into the causes of sex disparities observed in asthma prevalence and severity.Support or Funding InformationThis work was supported by NIH grants HL133520 (PS) and HL141618 (PS) and the APS Porter Fellowship (NF)
Interplay of microRNA‐503 and N‐acetylcysteine in regulating hypoxia‐reoxygenation injury in cardiomyocyte H9C2 cells
Authors: He Yanjing , Cai Yin , Irwin Michael G , Xia Zhengyuan 
Published: April 2020

microRNAs(miRs) are involved in many cardiovascular diseases including myocardial ischemia / reperfusion injury (IRI). Myocardial IRI can lead to oxidative stress, many miRs are regulated in response to oxidative stress. N‐acetylcysteine (NAC) –a known antioxidant, is reported to have protective effects against myocardial IRI. Previously, we and others observed that cardiac miR‐503 was down‐regulated during myocardial IRI in vivo in rodents or in cardiomyocyte H9C2 cells subjected to hypoxia and reoxygenation (H/R). This study is aimed to explore the interplay of miR503 and NAC in the regulation of hypoxia‐reoxygenation injury and possible molecular mechanisms in cardiomyocyte H9C2 cells. Here, H9C2 cells were subjected to hypoxia for 10 hours followed by reoxygenation 6 hours as simulated myocardial IRI. Results showed that cell viability was decreased and lactate dehydrogenase (LDH) activity was increased upon H/R; Pretreatment with NAC (1mM) to H9C2 cells for 30 hours can improve cell viability and reduce LDH activity in both normoxia and H/R states, while pretreatment with NAC (2mM) decreased cell viability and increased LDH activity. We also found that NAC (1mM) can down‐regulate miR‐503 expression level in H9C2 cells subjected to normoxia, it also down‐regulated miR‐503 expression level upon H/R. In order to inhibit endogenous miR‐503, H9C2 cells was transfected with Antagomir‐503, inhibition of miR‐503 can improve cell viability and reduce LDH activity upon H/R; Western blotting results showed that there was no significant difference among control, HR, HR+negative control of Antagomir‐503(NC) or HR+Antagomir‐503 groups for proapoptotic Bax protein, while anti‐apoptotic Bcl2 protein was increased after transfection of Antagomir‐503 compared with HR+NC. Target‐Scan software showed that Phosphoinositide‐3‐kinase, regulatory subunit 1(PI3K p85) is one of miR‐503 target genes, and P85 protein was decreased after H/R, but it was increased significantly after administration of Antagomir‐503, its down‐stream protein phospho‐AKT (Thr450) was also significantly increased. However, Antagomir‐503 and NAC (1mM) used in combination to H9C2 cells upon H/R, did not provide an additional advantage in protecting cell injury. It is concluded that administration of NAC (1mM) or Antagomir‐503 alone can alleviate cell injury, while the combination of NAC (1mM) and Antagomir‐503 has no synergistic cytoprotective effects, the possible molecular mechanisms need to be further explored.Support or Funding InformationSupported by Hong Kong RGC(17158616 M) and HMRF (05161826)grants.
RNAseq in fat pads from Alstrom syndrome 1 (ALMS1) Knockout rats support a role for ALMS1 in leptin release and fat metabolism in adipocytes.
Authors: Ortiz Victoria C. , Liao Tandong , Mendez Mariela , Ortiz Pablo A. 
Published: April 2020

Inactivating mutations in the ALMS1 gene in humans cause early onset obesity and metabolic syndrome. Deletion of ALMS1 in rodents induces early onset obesity and insulin resistance on normal chow. The mechanisms by which ALMS1 cause obesity are not clear but likely involve the development of leptin resistance since adult ALMS1 KO rats are hyperphagic and hyperleptinemic. It is not known whether ALMS1 controls leptin release and expression in adipocytes prior to development of obesity. ALMS1 has been associated to ciliary function which may play a role in adipogenesis. We hypothesized that deletion of ALMS1 would alter gene pathways involved in fat metabolism and cilia formation in young ALMS1 KO rats, which should contribute to hyperleptinemia. To study this, we obtained abdominal fat pads from young ALMS1 KO rats shortly after weaning (7 weeks) and prior to changes in body weight. We found that plasma leptin levels were higher in young ALMS1 KO rats (ALMS1: 3530±300 vs. WT: 172±16 pg/ml, p<0.05) despite no differences in body weight or baseline glucose. Leptin release (normalized to fat weight) was 120% higher (wild type: 30±8, ALMS1‐KO: 56± 15 pg/mg tissue/2h) in abdominal fat pads from ALMS1 KO rats. To study the genes involved in higher leptin release we performed RNAseq in freshly isolated
abdominal fat pads. 12,860 genes were identified and quantified. In fat pads from ALMS1 KO there were 163 upregulated and 237 significantly downregulated genes. Leptin mRNA was significantly increased (1.68 Log2 fold, p<0.001, n=4) as were several genes involved in lipogenesis (Elovl6, Scd1, Scd2, Mogat2, Acly, Lpl, Dgat2, Fasn). Ingenuity pathway analysis indicated that the top canonical pathways impacted were: oleate biosynthesis, glycolysis I, hepatic fibrosis, and clathrin‐mediated endocytosis signaling. Importantly, at least 10 ciliary genes were significantly decreased (Traf3ip1, Pard6g, Cngb1, Ak7, Ttll1, Mlf1, Ccdc162, Dnah6, Ccdc187, Foxj1) suggesting a role for ALMS1 in ciliary function. To examine this, we immunolabeled cilia in abdominal adipocytes with gamma acetylated tubulin antibodies, and examined sections by confocal microscopy. Short cilia (1–2 μm) were observed in both WT and ALMS1 KO adipocytes. We concluded that ALMS1 is likely involved in leptin release from adipocytes. It is not clear whether increased leptin expression and release is secondary to neuroendocrine alterations or alterations in cilia‐induced signaling. Our data also point to an important role for ALMS1 in regulating adipocyte function and leptin physiology.Support or Funding InformationHenry Ford Hospital Research Fund
Plasma Orexin A Level is Increased in Salt‐Sensitive Hypertension
Authors: Gao Huanjia , Bigalke Jeremy , Bloch Cassie , Chen Qinghui , Chen Bojun , Shan Zhiying 
Published: April 2020

Accumulating evidence demonstrates that the brain orexin system plays a crucial role in the control of blood pressure and cardiovascular function. In addition, hyperactivity of central orexin system function has been implicated in the pathology of salt sensitive hypertension (SSH). However, whether or not peripheral orexin system activity is altered in SSH remains unknown. In this study, we compared plasma orexin A levels and adrenal orexin receptors including orexin 1 receptor (OX1R) and orexin 2 receptor (OX2R) expression in Dahl salt‐sensitive (Dahl S) rat, an animal model of SSH, under high salt diet with their normotensive Dahl S cohorts. 7‐weekold male and age‐matched female Dahl S rats were divided into two groups of each and were fed either normal diet (NS, 0.4% NaCl) or high salt diet (HS, 4%NaCl). Six weeks following different diet treatments, all rats were euthanized and their plasma orexin A levels were measured using ELISA. Their adrenal glands were collected and subjected to real time PCR for OX1R and OX2R mRNA measurements. The results showed that HS diet increased plasma orexin A levels in both male (HS: 6.97±0.98 vs. NS 4.34±0.95 pg/ml; n=3; p=0.053); and female (HS: 6.17±0.65 vs. NS: 3.65±0.15 pg/ml; n=4; *P<0.05) rats. No significant difference was observed in orexin A level between male and female rats in both NS and HS groups. We therefore performed the rest of the experiments using female Dahl S rats. Real Time PCR results showed that OX1R mRNA level is increased by 3‐fold (n=5; *P<0.05) and OX2R mRNA level is increased by 1.9‐fold (n=5; *P<0.05) in HS diet treated rats compared to control rats. Immunohistochemistry assay showed that OX1R and OX2R are primarily expressed in the medulla of the adrenal gland and their expressions are co‐localized with tyrosine hydroxylase, an enzyme that catalyzes the first and rate limiting step in the synthesis of catecholamines including norepinephrine. This result coupled with the evidence that adrenal and plasma norepinephrine levels are increased in HS loaded Dahl S rats suggests that increased orexin system activity in the adrenal gland may increase norepinephrine production and subsequent release, therefore contributing to the development of SSH. Further studies will be performed to investigate whether blockage of the adrenal orexin receptors will decrease norepinephrine level and prevent or attenuate SSH in Dahl S rats.Support or Funding Information(Grant support: NIHR15HL129213 (Shan); Michigan Technological University and Portage Health Foundation Research Excellence Fund (Shan); Guangzhou University of Chinese Medicine, Graduate School international Schloarship (Gao)