bluuebunny/crossref_metadata_2025 huggingface dataset:
Neutrophil interactions with apical epithelial ICAM‐1 contribute to resolution of inflammation by promoting intestinal epithelial wound repair.
Authors: Sumagin Ronen , Nava Porfirio , Nusrat Asma , Parkos Charles A
Published: June 2021

A prominent feature of neutrophil (PMN) infiltration of the intestinal mucosa is transepithelial migration (TEM) that is associated with epithelial injury, barrier dysfunction and disease symptoms. While PMN TEM has been extensively characterized, the functional significance of PMN interactions with the luminal (apical) epithelial surface on intestinal epithelial cell (IEC) homeostasis is not clear. Here we show that under inflammatory conditions, PMN adhesion to upregulated ICAM‐1 on the apical epithelial membrane may initiate wound repair through increased cell proliferation. Specifically, we report that PMN adhesion to IEC ICAM‐1 results in activation of Akt and β‐catenin pathways leading to nuclear translocation of β‐catenin, and initiation of proliferative responses, as determined by incorporation of Edu. Such responses were ICAM‐1 dependent, as experiments modeling neutrophil engagement of epithelial ICAM‐1 by antibody‐mediated crosslinking yielded similar results. Furthermore, antibody‐crosslinking of ICAM‐1 enhanced epithelial wound repair in scratch wound assays. These findings suggest that while acute PMN TEM results in epithelial wounds, subsequent upregulation and engagement of ICAM‐1 by PMN on the luminal surface may mediate reparative proliferative responses.
NLRP3 Inflammasome is terminated by PGE2‐cAMP‐PKA signaling
Authors: Mortimer Leanne Marie , Moreau France , Chadee Kris
Published: June 2021

Inflammatory responses are measured and tightly controlled. Inflammasome activation has emerged as a major pathway of inflammation: these are intracellular complexes activated by pathogens and cellular stresses that recruit and activate caspase‐1, which successively activates IL‐1β. To date, we know little on how inflammasome activation is terminated. Macrophages highly express inflammasomes and are primary producers of IL‐1β. PGE2 is a potent lipid mediator liberally produced at sites of inflammation and can alter many innate immune functions of macrophages. Here we show NLRP3 inflammasome is rapidly dampened by concurrent PGE2‐cAMP‐PKA signaling. Bonemarrow‐ derived or THP‐1 macrophages were stimulated with LPS followed by nigericin or ATP to activate the NLRP3 inflammasome. Caspase‐1 and IL‐1β activation were detected by Western blot and ELISA. Co‐application of PGE2 or highly selective EP2 and EP4 agonists diminished NLRP3 activation. NLRP3 inhibition was reproduced by the cAMP agonist forskolin, cAMP analogue db‐cAMP and selective PKA activation. Likewise, PKA inhibition reversed NLRP3 inhibition by PGE2 and agonists of EP2, EP4 and cAMP. This is the first study to show PGE2 signaling arrests activation of the NLRP3 inflammasome. This work is funded by NSERC.
Cytosolic K+ and extracellular Na+ as regulators of NLRP3 inflammasome activation and the IL‐1β secretion response of macrophages to crystalline stimuli
Authors: Katsnelson Michael , Dubyak George
Published: June 2021

The crystalline particulate alum has widely been used as an adjuvant in vaccines. Alum and other crystalline particulates also stimulate NLRP3 (Nod‐like receptor protein 3)/caspase‐1 inflammasome assembly and secretion of mature IL‐1β via poorly understood mechanisms that involve destabilization of lysosomes and decreases in cytosolic [K+]. This study directly measures changes in cytosolic [K+] and characterizes mechanisms for K+ efflux in LPS‐primed murine bone marrow‐derived macrophages (BMDM) stimulated with either Imject Alum or the lysosomal destabilizing agent leu‐leu‐OMe. IL‐1β secretion in response to either alum or leu‐leu‐OMe was inhibited by increased extracellular [K+] or decreased extracellular [Na+]. Atomic absorbance spectroscopy was used to directly demonstrate reduced intracellular [K+] upon stimulation with alum. Alum‐stimulated K+ efflux and IL‐1β secretion was insensitive to several Trp channel antagonists. In contrast, pannexin1−/− BMDM demonstrated reduced IL‐1β release upon stimulation with alum. These observations indicate that combined efflux of intracellular K+ and influx of extracellular Na+ are necessary for optimal NLRP3 inflammasome activation in response to alum and lysosomal destabilization. Pannexin1 hemichannels may mediate in part the alum‐induced cation fluxes linked to NLRP3 inflammasome activation. (Supported by T32‐HL‐105338 and R01‐GM36387)
Resolvin D1 and Resolvin D5 Lower Antibiotic Doses in Infection
Authors: Chiang Nan , Fredman Gabrielle , Oh Sungwhan , Serhan Charles N
Published: June 2021

Underlying mechanisms for how bacterial infections contribute to active resolution of acute inflammation are unknown. Here, we carried out LM‐metabololipidomics following Escherichia coli (E. coli) infections in mice for temporal identification of pro‐inflammatory (prostaglandins and leukotrienes) and specialized pro‐resolving mediators (SPM). In self‐resolving E. coli exudates (105 CFU), the dominant SPM were resolvin (Rv) D5 and protectin D1 (PD1), which at 12 h were significantly greater than levels in exudates from higher titer E. coli (107 CFU) challenged mice. RvD1 and RvD5 (100 ng) each reduced bacterial titers, prevented hypothermia and increased survival, demonstrating the first actions of RvD5. With human macrophages, RvD1 and RvD5 each enhanced phagocytosis of E. coli via human G protein‐coupled receptor (GPR) 32, and countered a panel of pro‐inflammatory genes, including NF‐κB and TNF‐α. During E. coli infections, SPM (RvD1, RvD5, PD1) and the antibiotic ciprofloxacin accelerated resolution and heightened host antimicrobial responses. In skin infections, SPM enhanced vancomycin clearance of S. aureus. Therefore, specific SPM are temporally and differentially regulated during infections. They are anti‐phlogistic, enhance containment and lower antibiotic requirements for bacterial clearance. These results illustrate new opportunities to address antibiotic resistance.

[This work was supported in part by NIH grants P01GM095467 and R01GM38765 (CNS).]
A Biomechanical Study of Patterning During Gut Development
Authors: Shyer Amy Elizabeth
Published: June 2021

In order to efficiently pack the lengthy small intestine into the body cavity, the developing vertebrate gut tube forms a reproducible looped pattern as it grows. We previously determined that gut looping morphogenesis is driven by the forces that arise from the differential growth between the gut tube and the anchoring dorsal mesenteric sheet. While the looping pattern provides an artful mechanism for fitting a longer tube into the body, undoubtedly contributing to the luminal surface area necessary for optimal nutrient absorption, one can imagine alternative mechanisms of increasing surface area such as modifying the morphology of the luminal tissue. The lumen of the intestine is covered with finger‐like projections termed villi which serve to dramatically increase the surface area of the intestine. Although very little is known about the mechanism driving villi morphogenesis, the formation of villi has previously been explored at a descriptive level in chick. During this process in chick, the intestine undergoes multiple phases of morphogenesis, beginning with the formation of longitudinally oriented ridges, which appear as an inward buckling of the luminal surface of the tube. These ridges become kinked with regular and uniform periodicity appearing as raised zigzags. These zigzags then enlarge and flatten at which point definitive villi become visible in a specific, predetermined pattern along the zigzags. We hypothesize that tissue‐level forces from the smooth muscle layers lead to these striking patterns seen in the compressed mesenchyme and attached endoderm. These same forces appear to instigate patterns of signaling necessary for outgrowth of definitive villi from the zigzag pattern. Additionally, these signaling environments delineated by tissue‐level forces may be essential for directing the location of adult intestinal stem cell markers during the morphogenesis of villi and crypts.
Localization and distribution of CNC and VSM cells in a mouse model of aortopathy: a novel 3D approach
Authors: Riley Ashlie Evans , Maddy Kelsey , Junor Lorain , Biechler Stefanie , Fuseler John , Krishnamurthy Varun , Hinton Robert , Goodwin Richard
Published: June 2021

Aortopathy is a subclinical disease characterized by aortic dilation, which can result in aortic dissection and/or aortic aneurysm. Hallmarks of aortopathy include vascular smooth muscle cell (VSMC) and elastin fiber deposition misregulation. Elastin hemizygous (Eln+/−) mice have a thickened and altered arterial wall structure, specifically, aortic root distension and an increase in VSMC layers along the ascending aorta. We hypothesize that aortopathy has a congenital basis with the associated defects originating in the cardiac neural crest (CNC) regions of the aorta. Aortic tissues from adult wild type (WT) and Eln+/− mice were examined to detect anatomic and/or developmental variation. Sonic Hedgehog (a marker of CNC derived cells), SM22 (a marker of VSMCs), and Heart's stain (a stain for elastin architecture) were utilized in a novel 3D reconstructive manner to discern spatial, temporal localization in WT vs. Eln +/− hearts. Quantification of these findings demonstrate that Shh is co‐localized to the observed anomalies with VSMC deposition being increase post sinotubular junction suggesting that aortopathic disruption in structure originates in the cardiac neural crest region with symptoms not manifesting until adulthood.
The Myth of Cryopreservation of Heart Valves
Authors: Stock Ulrich A. , Lisy Milan , Kalender Guenay , Schenke‐Layland Katja , Brockbank Kelvin GM
Published: June 2021

Allografts represent almost perfect heart valve substitutes with optimal hemodynamic characteristics and resistance to infections. Disadvantages are limited availability, inability to grow and long‐term failure. Since 1968 frozen cryopreservation (FC) is employed including storage in vapour phase liquid nitrogen. For decades it was assumed that this preservation protects cells and extracellular matrix (ECM). However as learnt from explantations latest 1 year after implantation no more viable cells can be detected. Application of multiphoton imaging analysis (MIA) enables three‐dimensional visualization of ECM by induction of autofluorescence allowing detection of freezing induced destruction of elastic and collagenous matrix in FC valves due to ice crystals. The disruptive interstitial ice damage can be avoided by promoting vitrification instead of ice formation. As the preservation and warming is accomplished without any tissue destruction vitrification enabled for the first time complete preservation of extracellular matrix in heart valves. In order to simplify the required infrastructure, we have further developed ice free cryopreservation (IFC) permiting storage above the solution's glass transition temperature at −80°C. We applied IFC in a sheep and could demonstrate that it is anti‐inflammatory preventing maintains ECM resulting in better valve function and diminished degeneration.
The Visible Interactive Dinosaur project: fleshing out dinosaur head anatomy and function with advanced 3D imaging
Authors: Witmer Lawrence M
Published: June 2021

The American Association of Anatomists’ first President in 1888 was Joseph Leidy, who was not only a medical anatomist but also the preeminent American dinosaur paleontologist of his time. The link between dinosaurs, medical anatomy, and AAA remains strong, but, 125 year later, has a distinctly 21st‐Century character. The Visible Interactive Dinosaur (VID) project restores dinosaur head anatomy by reconstructing soft‐tissue systems within a 3D digital environment. Hypotheses regarding dinosaur soft tissues are tested by reference to extant sister groups (birds, crocs, etc.), drawing on the osteological correlates of the soft tissues (e.g., muscle scars, neurovascular grooves) that can be surveyed in the fossils. CT scanning brings the anatomy into the digital realm where 3D analytical and modeling software allows restored dinosaur skulls to be “fleshed out” in a testable manner. Packing constraints—getting the soft tissues to fit and function within the confined space of the skull—emerge as a key test of restored anatomy. Example dinosaur taxa (the herbivorous sauropod Diplodocus and the predatory theropod Majungasaurus) show the utility of VID restorations to test higher‐order functional hypotheses pertaining to thermal physiology, nasal airflow, and feeding biomechanics. Moreover, VID restorations are outreach vehicles to educate the public about complex issues pertaining to science and evolution.
The evolution of human anatomy and locomotion
Authors: Ward Carol V
Published: June 2021

The interplay between anatomy and evolution has been at the heart of the American Association of Anatomists since its origin in 1888 with its first president, Joseph Leidy, who was a paleontologist, biologist and anatomist. Our current conception of the basic trajectory of human evolution is even younger, dating back only to the 1925 discovery of Australopithecus. Since then, an impressive array of hominin fossils have been discovered, resulting in a more complex and nuanced view of how humans evolved than Leidy could have imagined. These fossils show us that the transition to upright, bipedal locomotion is the hallmark of the human lineage, and that the specialized anatomy associated with bipedality was present in our earliest ancestors. An appreciation of the functional demands on the human skeleton imposed by upright posture, and the resulting anatomy and variation that remains within humans, is at the forefront of medicine and contributing to advances in management and treatment of modern human disorders. Reviewing some of the most recent hominin fossil discoveries highlights that it is consideration of human and comparative anatomy that informs our interpretations of these fossils, but also that understanding the sequence of morphological changes that shaped our lineage gives us important new perspectives on modern human anatomy, function and dysfunction.
Reprogramming of Vascular Smooth Muscle Cells into Calcifying Cells: What Can We Learn from the Embryo?
Authors: Kempf Hervé
Published: June 2021

Vascular calcifications are widespread in aged individuals and patients with atherosclerosis, diabetes or chronic renal failure. These calcifications of the blood vessels are mostly due to a phenotypic switch of vascular smooth muscle cells (VSMCs) that reprogram into calcifying cells through mechanisms and gene regulations that mimic embryonic bone formation.

Over the years, a regiment of systemic factors has been shown to induce VSMCs to trans‐differentiate into cartilage or bone cells and eventually calcify. However, in vivo, vascular calcifications do not form in a randomized pattern, suggesting that intrinsic properties of VSMCs happen to play a role in the occurrence of the calcified lesions. As VSMCs derive from various independent origins during development, evidence will be put forward to prove that VSMCs are differentially prone to calcify within the vascular wall upon their heterogeneous embryological origin.
Teaching Histology in Integrated Curricula: Challenges and Opportunities
Authors: Amerongen Helen
Published: June 2021

Approximately half of AAMC medical schools now teach histology as part of an integrated program in which individual basic science disciplines have no distinct identity. Challenges of this approach include that histology content can be too scattered and too superficial to support mastery, and the true integration rather than simple juxtaposition that is required is time consuming and therefore costly. Also, because it is a difficult but fractional part of their work, students can choose to selectively ignore histology and still pass exams. Opportunities arising out of the integrative approach include that the necessary collaboration of faculty across discipline boundaries can foster collaboration in other arenas such as research. In addition, melding histology with bordering disciplines of anatomy, physiology and pathology gives tremendous scope for building a clinically‐relevant context that is more motivating for students and may also be more effective in supporting transfer of knowledge to clinical settings. Moreover, interdisciplinary content is ideally suited for case‐based student‐centered learning formats including problem‐based learning, team‐based learning, audience participation via response systems, online case study, etc. Specific examples illustrating these challenges and opportunities including incorporation of histology into a variety of learning formats will be presented.
The Pitx2:miR‐200 Family Axis Regulates WNT and BMP Signaling During Tooth Morphogenesis and Renewal
Authors: Li Xiao , Cao Huojun , Jheon Andrew , Sun Zhao , Wang Jianbo , Klein Ophir , Amendt Brad A
Published: June 2021

The mouse incisor is a remarkable tooth that grows throughout the animal's lifetime. This continuous renewal is fueled by epithelial stem cells and little is known about the function of specific miRNAs in this process. We describe a novel Pitx2:miR‐200c/141:Noggin regulatory pathway in dental epithelial cell differentiation. miR‐200c represses Noggin, an antagonist of Bmp signaling. Pitx2 expression caused an up‐regulation of miR‐200c and ChIP assays reveal endogenous Pitx2 binding to the miR‐200c/141 promoter. A positive feedback loop was discovered between miR‐200c and Bmp signaling. miR‐200c/141 induced expression of E‐cadherin and the dental epithelial cell differentiation marker, amelogenin. miR‐200c/141 knockout mice show defects in enamel formation and decreases in E‐cadherin and amelogenin. Wnt/β‐catenin signaling is regulated by miR‐200a,b/429 under the control of Pitx2. A negative feedback loop was identified between miR‐200a and Pitx2 and includes β‐ catenin. Our in vivo and in vitro studies reveal a multistep transcriptional program involving the Pitx2:miR‐200c/141:Noggin regulatory pathway and a Pitx2:miR‐200a,b/429 Wnt/β‐catenin pathway that is important in tooth development. Support for this research was provided from grants DE13941 and DE18885 from the National Institute of Dental and Craniofacial Research.

Grant Funding Source: National Institute of Dental and Craniofacial Research
Intrinsic pluripotent stem cells, Muse cells, are a primary source of iPS cells in human fibroblasts
Authors: Dezawa Mari
Published: June 2021

We found a novel type of pluripotent stem cells Multilineage differentiating Stress Enduring (Muse) cells (Kuroda et al., PNAS, 2010). They reside in adult mesenchymal tissues or cultured fibroblasts and can be isolated as cells double positive for mesenchymal marker CD105 and human ES cell marker SSEA‐3. Muse cells express pluripotency markers, self‐renew and exhibit triploblastic differentiation from a single cell but are not immortal in culture or tumorigenic in vivo. Recently, we found that Muse cells are a primary source of iPS cells in human fibroblasts (Wakao et al., PNAS, 2011). When human fibroblasts were separated into Muse and non‐Muse cells and transduced with Oct3/4, Sox2, Klf4, and c‐Myc, iPS cells were generated exclusively from Muse cells, but never from non‐Muse cells. Epigenetic alterations were not seen in non‐Muse cells and some of the major pluripotency markers were not expressed for the entire period of generation, suggesting that Muse cells that are already pluripotent selectively become iPS cells by acquiring tumorigenic proliferative activity, whereas the remaining cells make no contribution to the generation of iPS cells.
Uncovering cranial anatomy variation in fossil tarsiiform crania using microCT
Authors: Rosenberger Alfred L , Boyer Doug
Published: June 2021

Our microCT study of two differentially damaged ~50 mya crania from central Wyoming, identified as the omomyid primate Shoshonius, reveals new morphology and clarifies prior observations done without the benefit of CT. Omomyids were a successful group during the Eocene Epoch but their functional anatomy and adaptive diversity remains poorly documented, and their affinities are debatable. They may be most closely related to the living tarsiiform genus Tarsius, but some believe tarsiers are most closely related to the anthropoids, to the exclusion of the omomyids. We reinterpret these specimens as two distinct morphs, potentially warranting generic separation, which may explain prior conflicting morphological assessments. Both have extra‐large orbits similar to tarsiers but the CM60494 basicranium resembles the highly derived Tarsius in the auditory region, including the centralized or medial position of the posterior carotid foramen, while CM31366 is more primitive and plesiadapid‐like, exhibiting a highly inflated auditory bulla with a reduced posterior carotid foramen located postero‐laterally behind the auditory meatus. Among the implications: an early split of the Tarsius lineage, a further challenge to the tarsier‐anthropoid hypothesis, and indications that eyeball hypertrophy and nocturnal visual predation was an early, formative adaptation of the tarsiiform group.
Mechanism of Eukaryotic Helicase Loading and Activation
Authors: Bell Stephen P , Kang Sukhyun , Ticau Simina , Chan Bena Sze Ham
Published: June 2021

The Mcm2–7 helicase is the only eukaryotic DNA replication protein that participates in all steps of this essential process. Loading of the Mcm2–7 complex at origin DNA licenses replication origins during the G1 phase. Assembly of the Cdc45 and GINS helicase‐activating proteins onto the Mcm2–7 helicase is a central event during the replication origin activation. The resulting Cdc45/Mcm2–7/GINS (CMG) complex is the replicative DNA helicase during replication elongation.

We are using a combination of genetic screens and biochemical assays that recapitulate the events of replication initiation to investigate the mechanism of Mcm2–7 loading and activation. Contrary to previous studies, we find that the majority of mutations predicted to inhibit Mcm2–7 helicase function (e.g. mutations in the ATP binding and hydrolysis motifs) are defective for helicase loading. This has led us to hypothesize that the process of Mcm2–7 helicase loading requires one or more activities that are also required for helicase activity of this complex. We will report on our latest studies investigating this connection. We have identified new lethal mutations in proteins predicted to activate the Mcm2–7 helicase. We will report on what we have learned from studying the corresponding mutant proteins using biochemical‐complementation assays for helicase activation and replication initiation.
Spatiotemporal Control of Endocytosis by Phosphatidylinositol 3,4‐Bisphosphate
Authors: Haucke Volker , Posor York
Published: June 2021

Phosphoinositides (PIs) serve crucial roles in cell physiology ranging from cell signalling to intracellular membrane traffic. While the function of many PIs has been studied in detail little is known about the role of phosphatidylinositol‐3,4‐bisphosphate [PI(3,4)P2]. Here, we show that formation of PI(3,4)P2 by class II phosphatidylinositol 3‐kinase C2α (PI3K C2α) spatiotemporally controls clathrin‐mediated endocytosis. Depletion of PI(3,4)P2 or PI3K C2α impairs the maturation of clathrin‐coated pits at a late stage preceding fission. Timed formation of PI(3,4)P2 by PI3K C2α is required for the selective enrichment of the BAR domain protein SNX9 at late‐stage endocytic intermediates, in accordance with mathematical modelling. These findings provide a mechanistic framework for the role of PI(3,4)P2 in endocytosis and unravel a novel discrete function of PI(3,4)P2 in a central cell physiological process.
Separation of time‐scales in the functional biophysics of BAR domain proteins.
Authors: Baumgart Tobias
Published: June 2021

Membrane curvature has recently emerged at the forefront of membrane biophysics. Numerous proteins involved in membrane curvature sensing and membrane curvature generation have recently been discovered, including proteins containing the crescent‐shaped BAR domain as membrane binding and shaping module. Accordingly, the structure determination of these proteins and their multimeric complexes is increasingly well‐understood. Substantially less understood, however, are thermodynamic and kinetic aspects and the detailed mechanisms of how these proteins interact with membranes in a curvature‐dependent manner. New experimental approaches need to be combined with established techniques to be able to fill in these missing details. Here we use model membrane systems in combination with a variety of biophysical techniques to characterize mechanistic aspects of BAR domain protein function. We first establish kinetic and thermodynamic aspects of BAR protein dimerization in solution, and then investigate kinetic aspects of membrane binding. We identify significantly different time scales for protein dimerization, membrane association, membrane insertion, and oligomerization. This separation of time scales likely is relevant for BAR protein function in‐vivo.
The Presidential Postdoctoral Program at Novartis Institutes for BioMedical Research
Authors: Pond Leslie
Published: June 2021

The Presidential Postdoctoral Program provides scientists with a unique opportunity to perform innovative, basic research in a pharmaceutical company, while supporting flexibility in their future career options in academia or industry. Postdoctoral scholars design their own research plan, receive guidance from two mentors (a Principal Investigator at Novartis who serves as the primary mentor, and an academic faculty member), and are expected to present their work at major research conferences and publish their results in leading peer‐reviewed journals. Postdocs also have regular opportunities to present their work within Novartis and interact with scientific leaders. We encourage applications from candidates who wish to pursue a career as academic faculty as well as candidates interested in industry careers. Postdocs conduct research at one of our global sites. Our postdocs go on to lead research groups in industry and academia or work in a variety of science‐based, non‐research positions.
Breastmilk is a novel source of stem cells with multi‐lineage differentiation potential
Authors: Hassiotou Foteini , Filgueira Luis , Hartmann Peter E.
Published: June 2021

The breast undergoes extensive remodelling at each pregnancy/lactation, based on mammary stem cell (MaSC) expansion and differentiation. Most of our knowledge on MaSCs comes from the resting breast, where these cells are quiescent and in low numbers. In our laboratory we examine the lactating breast that contains higher numbers of MaSCs, which we access non‐invasively via breastmilk. Here, we describe the presence of MaSCs (CD49f+/CD29+) in breastmilk, which were able to self‐renew and differentiate into mammary cells. In addition, a cell population was identified expressing pluripotency markers (OCT4, SOX2, NANOG) and being able to self‐renew and differentiate into various lineages, including: mammary cells, neuron‐like and glia‐like cells, hepatocyte‐like cells expressing albumin, pancreatic beta‐like cells expressing insulin, osteoblasts, chondrocytes, adipocytes and cardiomyocytes. These findings strongly suggest the existence of a cellular hierarchy in the human lactating breast, at the top of which sits a cell population with potential pluripotency, which can be easily accessed via breastmilk.

Grant Funding Source: Women and Infants Research Foundation; and Medela AG (Switzerland)
Genetic dissection of Treacher Collins Syndrome: Polr1c and Polr1d
Authors: Achilleos Annita , Watt Kristin , Trainor Paul
Published: June 2021

Treacher Collins Syndrome (TCS) is a congenital disorder characterized by craniofacial deformities including external and middle ear defects, cleft palate and hypoplasia of facial bones. TCOF1 underlies TCS in humans and a mouse model was generated to investigate its developmental causes. Tcof1+/− mice show phenotypes similar to human TCS caused by a decrease in cranial neural crest cells (CNCC) that give rise to the majority of bone in the head and face. Tcof1 encodes the nucleolar phosphoprotein Treacle which functions in ribosomal DNA (rDNA) transcription. Tcof1+/− mice exhibit death of CNCC precursors due to a reduction in mature ribosomes. Recently, two more genes involved in rDNA transcription have been identified in TCS patients: POLR1C and POLR1D. To investigate the role of these genes in TCS we turned to zebrafish and mouse. Disruption of polr1c or polr1d in zebrafish results in specific craniofacial defects including loss of facial structures such as the ceratobranchials, hypoplasia of the cranial skeleton and deformed ethmoid plate. Similar to tcof mice, these defects arise from a reduction in CNCC caused by cell death. Further, neural crest specific knock out of Polr1c in mice leads to craniofacial defects similar to human TCS, including hypoplasia of the mandible and severe defects in the inner and middle ear bones. Our analysis suggests that both animal models can be used to dissect out the etiology of TCS.

Grant Funding Source: AAA Postdoctoral Fellowship
Post‐initiation roles for the sigma subunit of bacterial RNA polymerase
Authors: Hochschild Ann , Deighan Padraig , Nair Nikhil , Berry Katie , Goldman Seth , Nickels Bryce
Published: June 2021

Promoter recognition in bacteria depends on the σ subunit of RNA polymerase. However, σ can also function during the elongation phase of transcription by recognizing promoter‐like DNA sequence elements in the initial transcribed region that induce early elongation pausing (Roberts et al., CSH Symp Quant Biol 63: 319–325; 1998). In previous work, we demonstrated that such promoter‐like pause‐inducing elements function to inhibit σ70 loss during the earliest stage of elongation, increasing the σ70 content of elongation complexes throughout the transcription unit. This effect of a σ70‐dependent early elongation pause element can be detected as enhanced pausing at downstream locations. Current efforts are directed at investigating the implications of these findings for the control of transcription elongation. We are also investigating the possibility that σ70 can extend its reach by gaining access to elongation complexes on transcription units that are controlled by non σ70‐dependent promoters.
Roles for Glycans in Mammalian Development and Spermatogenesis
Authors: Stanley Pamela , Batista Frank , Huang Hung‐Hsiang
Published: June 2021

N‐ and O‐glycans of mammalian glycoproteins play numerous roles within the secretory pathway and at the cell surface. Thus, deletion of a glycosyltransferase responsible for the synthesis of major classes of glycans usually leads to embryonic death. To identify roles for specific glycans in spermatogenesis, conditional deletion in spermatogonia was performed using males transgenic for Stra8‐iCre recombinase. All male germ cells derive from spermatogonia via differentiation in close association with Sertoli cells to sperm. Of three glycosyltransferase genes inactivated, only conditional deletion of Mgat1, encoding the glycosyltransferase that initiates complex N‐glycan synthesis, disrupted spermatogenesis. A block at the post‐meiotic spermatid stage resulted in male infertility. Interestingly, at an earlier stage of spermatogenesis, Mgat1 gene expression is downregulated in spermatocytes, and expression of a physiological inhibitor of MGAT1 transferase activity, termed GnT1IP‐L, is upregulated. We are testing the hypothesis that high mannose N‐glycans expressed on germ cell glycoproteins when Mgat1 is downregulated and the GnT1IP‐L gene is expressed, are required for spermatocyte/Sertoli cell interactions; while complex N‐glycans, generated when Mgat1 is upregulated and GnT1IP‐L is low, are required for spermatids to differentiate into spermatozoa.
The ESCRT Pathway in HIV Budding and Cytokinesis
Authors: Sundquist Wesley I.
Published: June 2021

The Endosomal Sorting Complexes Required for Transport (ESCRT) pathway mediates intraluminal endosomal vesicle formation, budding of HIV‐1 and other enveloped viruses, and the final abscission step of cytokinesis in mammals and archaea. I will review our current understanding of the roles of different ESCRT factors in HIV budding and abscission. In particular, I will describe our experiments aimed at understanding how the filament‐forming ESCRT‐III subunits and the VPS4 ATPase assemble and function in membrane fission.
Disrupting Bacterial Communication to Suppress Multidrug Resistant Phenotypes
Authors: Melander Christian
Published: June 2021

Bacteria have evolved an intricate communication network to sense and react to environmental stressors, including protection from antibiotic threats. A key bacterial sensing system is a two component system that typically involves a membrane‐bound histidine kinase (HK) sensor and cytoplasmic protein called a response regulator (RR). Upon sensing the environmental input, the HK will phosphorylate the RR and induce changes in gene transcription to help the bacteria adapt and survive to a diverse array of environmental conditions. Small molecules based upon the 2‐aminoimidazole (2‐AI) scaffold have been developed that potentiate this system and we have demonstrated that these small molecules will effectively restore susceptibility of mutldrug resistant bacteria to the action of antibiotics they are resistant to.
Signals from the Surface to Control Cell Fate Decisions
Authors: Kiessling Laura L
Published: June 2021

Human pluripotent stem cells (human embryonic stem cells and induced pluripotent stem cells) have the ability to self‐renew indefinitely and differentiate into all cell types. The repertoire of signals that have been used to date to direct their propagation or differentiation is limited. Additionally, undefined culture conditions are typically used, which complicates determining the critical signaling pathways for expansion or differentiation. Our goal is to identify chemically defined conditions to control signaling pathways to elicit human pluripotent stem (hPS) cell selfrenewal or differentiation. In physiological settings, the microenvironment of the niche transmits information through three types of components: soluble molecules, neighboring cells, and the extracellular matrix. While the focus has been on identifying soluble components that influence cell decisions, insoluble components are likely to have critical roles in orchestrating signaling. To mine this molecular space, we devised arrays of chemically defined surfaces composed of self‐assembled monolayers. From these arrays, we identified surfaces with surprising assets: They not only permit human pluripotent stem cells to differentiate to specific lineages but even instruct them to do so. Our findings highlight the dramatic effects of insoluble cues on stem cell pluripotency and differentiation. Moreover, they underscore the utility of defined surfaces to dissect critical signaling pathways. This research was supported by the NIH (R01‐ GM049975 and R01‐AI055258).
Educating Students for an Interdisciplinary World
Authors: Kuhn Leslie A.
Published: June 2021

There is a lot of buzz about interdisciplinary training these days. What really works? The interdisciplinary graduate training program in Quantitative Biology at Michigan State University (biomodel.msu.edu) was developed a dozen years ago, with the goal of potentiating creative, groundbreaking work between the life sciences and physical, computational, and mathematical sciences. The key element has been joint research mentoring of our students in the context of collaboration, with one major advisor from the biosciences and one from the physical/computational/math disciplines. How do we keep the course load reasonable? Is it possible to become too interdisciplinary (neither fish nor fowl)? What have our graduates gone on to do? How has their work impacted the labs in which they trained? Can this approach work at the undergraduate level, too? We will share our successes and lessons learned, and look forward to learning from the audience's successes, too!
Gene and nutrient interaction in obesity: trans fatty acid intake and genetic risk score interaction in a multiethnic sample of children
Authors: Brown Michelle M Bohan , Rodriguez‐Cruz Maricela , Fernandez Jose R.
Published: June 2021

Associations of trans fatty acids (TFA) consumption with obesity may be mediated by individual's genetic background. We hypothesized that adiposity in children may be explained by the interaction of TFA intake and obesity genetic risk score. Fat mass index (FMI = fat mass/height2) and BMI were calculated for 279 children of European‐, African‐ and Hispanic‐American background (EA, AA, HA, respectively). Fat mass was determined using dual x‐ray absorptiometry, TFA intake was obtained from 24 hour dietary recall. Genetic risk score (GRS) was calculated from 16 single nucleotide polymorphisms by counting the number of obesity‐risk alleles. Association models evaluating the interaction of GRS and TFA were adjusted by standard covariates and genetic admixture. Significant GRS × TFA interactions were obtained for BMI and FMI (p= 0.0203 and 0.0055 respectively) in the total sample. In EA children, interaction of GRS and TFA intake was significantly associated with BMI and FMI (p=0.0433 and 0.0215 respectively); however, the interaction was not significant in AA or HA children. In the entire sample, the interaction model explained more of the variability for FMI and BMI (2.54% and 1.9% respectively) than a model with GRS alone. Our results support that dietary contributions to adiposity may be dependent on genetic background, a relevant finding for obesity prevention strategies.

Grant Funding Source: NIDDK DK067426 and T32DK062710
Silencing of the Menkes Copper‐Transporting ATPase (Atp7a) in Intestinal Epithelial Cells Impairs Cell Growth and Cell Cycle Control
Authors: Gulec Sukru , Collins James F.
Published: June 2021

The Menkes copper‐transporting ATPase (Atp7a) is induced in the rodent duodenum during iron deficiency, suggesting that copper influences iron homeostasis. To understand the role of Atp7a in intestinal iron & copper metabolism, we generated an Atp7a knockdown (KD) IEC‐6 cell line using shRNA technology. Atp7a protein was reduced >;70% and KD cells accumulated copper, consistent with defective copper efflux mediated by Atp7a. Unexpectedly, Atp7a KD impaired growth rate as compared to negative control shRNA‐transfected cells (~40% at 96 hours; p<0.0001). Further experiments were thus designed to elucidate the role of Atp7a in cell growth. Western blot analysis demonstrated that cyclin D1 (CD1) expression increased in KD cells while PCNA expression was unchanged. CD1 induces cell cycle arrest while PCNA promotes cell division. This suggested that Atp7a KD perturbed progression through the cell cycle. CD1 expression was not further enhanced by copper loading, implicating Atp7a in regulation of cell growth independent of its documented role in maintaining cellular copper homeostasis. Subsequent experiments showed that expression of additional markers of cell cycle control (p53, p21 & Ki67) was consistent with decreased cell growth. Overall, these novel findings support the hypothesis that Atp7a function is important for regulation of intestinal epithelial cell proliferation and differentiation.

Grant Funding Source: NIH
Status of Selected Iron‐Status Biomarkers in Juvenile End‐ Stage Renal Disease in a Guatemalan Context: Comparative Aspects by Mode of Dialysis
Authors: Casimiro Juliana , Lou‐Meda Randall , Olbert Marion , Weiss Günter , Swinkels Dorine W , Tjalsma Harold , Solomons Noel W , Schümann Klaus
Published: June 2021

Background

Anemia is common in end‐stage renal disease (ESRD) with both inflammation and reduced erythropoietin exerting an important role.

Objective

To assess selected iron (Fe) biomarkers in Guatemalan low‐income patients on hemodialysis (HD) and peritoneal dialysis (PD).

Methods

Iron biomarkers were determined in 27 juveniles after HD and 27 on PD therapy (7–20 y).

Results

Hb mean was 9.0±1.9 g/dL in HD and 9.9±2.7 g/dL in PD (p=0.17). Respective ranges for serum Fe were 24–148 and 30–175 μg/dL (p=0.29); for transferrin were 7–63 and 12–103 ng/mL (p=0.23); for sTfR were 1.8–6.1 and 0.3–7.1 mg/L (p=0.14); for ferritin 193–4,113 and 17–1,506 ng/mL (p=0.01) and hepcidin‐25 (hep‐25) median values were 36 and 43 nM (p=0.68). Ferritin correlated significantly with hep‐25 in PD (r=0.72; p<0.001), but not in HD (r=0.04; p=0.82).

Conclusions

Higher ferritin status and its low correlation with hep‐25 in HD may in part be due to high parenteral Fe vs. oral Fe administration in PD. Overall levels of ferritin and hep‐25 in juvenile, Guatemalan ESRD patients were 4–5 times higher than Dutch adults (Peters et al, Nephrol Dial Transplant, 2010; 25: 848–53). This is possibly related to an inflammatory overlay of the low‐income setting that leads to high hep‐25 associated and to macrophage iron withholding and Fe‐restricted erythropoiesis.

Funded by Hildegard Grunow Foundation, Munich and MBR Optical Systems, Wuppertal
Epigallocatechin gallate (EGCG) ‐ loaded nanoparticles decrease cholesterol content in THP‐1 derived macrophages
Authors: Zhang Jia , Nie Shufang , Wang Shu
Published: June 2021

The aim of this study is to synthesize EGCG‐loaded nanoparticles (Nano‐E) to improve EGCG stability, cellular content, decrease cholesterol accumulation and inflammatory factor production in THP‐1 derived macrophages.

Biocompatible and biodegradable Nano–E was prepared by a phase inversion method. The particle size and zeta potential of Nano‐E were measured using Brookhaven analyzer. Encapsulation efficiency (EE), loading content (LC), EGCG release, stability of Nano‐E in different temperatures and pH, cellular EGCG and cholesterol content were measured using a HPLC system. Cellular uptake of fluorescent‐labeled Nano‐E was measured using a fluorescent microscope. Cell viability was measured using a MTT method.

Nano‐E were about 45–55nm in diameter. The EE and LC were 90% and 3%, respectively. Nano‐E dramatically improved EGCG stability and increased its uptake by macrophages. The Nano‐E increased cellular EGCG content by 5‐fold compared with native EGCG at 100 μM at 37°C. As compared to 10 μM of native EGCG, Nano‐E at the same concentration significantly decreased cellular total and esterified cholesterol content in macrophages. Both Nano‐E and native EGCG at 10μM didn't decrease macrophage viability.

Nano‐E significantly increased EGCG stability, cellular EGCG content, and decreased cholesterol accumulation in THP‐1 derived macrophages.

Grant Funding Source: Grant Funding Source: NIH 1R15AT007013–01
Differential effects of whole and refined grains on abdominal adipose tissue depots in an exploratory subgroup analysis of the GRAIN Study
Authors: Harris Kristina , Lemieux Susan , Jonnalagadda Satya , West Sheila , Heuvel Jack Vanden , Kris‐Etherton Penny
Published: June 2021

Abdominal obesity is associated with insulin resistance, specifically having a higher amount of adipose tissue (AT) in visceral (VAT) rather than abdominal subcutaneous (ASAT) depots. We explored the relationship between ASAT, VAT and metabolic and anthropometric variables in a subgroup (n =28/50) of participants from a 12‐week, randomized, parallel arm, weight‐loss study in overweight/obese individuals comparing whole (WG) and refined grains (RG). Abdominal MRI was performed at 0 and 12 weeks. Axial scans at lumbar vertebrae L4–5 to L3 were manually traced to separate the AT depots. Baseline ASAT correlated with BMI, waist circumference, total and central AT (from DXA), C‐reactive protein, interleukin 6, and leptin (Rs>;0.55, P<0.05); VAT correlated with central AT, insulin and HOMA‐IR (Rs>;0.35, P<0.05). VAT % change correlated inversely with alkylresorcinol % change, a biomarker of whole grain intake (L3 only, R=−0.51, P<0.01). While the between group changes in ASAT and VAT were not significant, VAT decreased significantly from baseline in only the WG group (RG: −14 ± 20 g, P=0.08; WG: −18 ± 20 g, P=0.001). These findings suggest that VAT and insulin are strongly associated, whereas ASAT is more highly correlated with body fatness and inflammatory markers. Consumption of WG in the place of RG during weight loss may elicit a greater reduction in VAT, but more research is warranted.

Grant Funding Source: General Mills Bell Institute of Health and Nutrition
Validity of the National Cancer Institute's Automated Self‐ Administered 24‐hour Recall (ASA24): Results of a Feeding Study
Authors: Kirkpatrick Sharon I , Thompson Frances E , Subar Amy F , Douglass Deirdre , Zimmerman Thea P , Kahle Lisa L , George Stephanie M , Potischman Nancy
Published: June 2021

The Automated Self‐administered 24‐hour dietary recall (ASA24) is a web‐based tool that enables the collection of multiple recalls in large studies at minimal cost. To assess the validity of ASA24, we studied 83 adults, aged 20–70 years, who ate three meals at a study center. True intake was ascertained by inconspicuously weighing foods and beverages offered at a buffet before and after each participant served him/herself, as well as plate waste. Participants were randomly assigned to complete ASA24 or an interviewer‐administered recall using the Automated Multiple‐Pass Method (AMPM) on the following day, and the degree to which reported and true intake matched was examined. Data for one ASA24 participant was excluded because reported intake did not correspond to the previous day. For study center meals, average true energy intake of ASA24 participants (n=40) was 2329 calories compared to an average reported intake of 2138 calories (−191). On average, exact or close matches were reported for 72% of items consumed. Among AMPM participants (n=42), average true energy intake was 2268 calories compared to an average reported intake of 2425 calories (+157). On average, exact or close matches were reported for 78% of items consumed. The average number of items consumed but not reported was 7 for ASA24 and 6 for AMPM. The results support the use of ASA24 as a cost‐effective means of collecting high‐quality dietary data.

Grant Funding Source: n/a
Evaluation of the Healthy Eating Index‐2010 (HEI‐2010)
Authors: Guenther Patricia M. , Kirkpatrick Sharon I. , Krebs‐Smith Susan M. , Reedy Jill , Buckman Dennis W. , Dodd Kevin W. , Carroll Raymond J.
Published: June 2021

The Healthy Eating Index, a measure of diet quality, was recently updated to reflect the 2010 Dietary Guidelines for Americans. To assess the validity and reliability of the HEI‐2010, 24‐hour dietary recall data from individuals aged 2 years and older (n=8262) from the 2003–2004 National Health and Nutrition Examination Survey were used to estimate usual HEI‐2010 scores and to assess whether the HEI‐2010 measures diet quality independently of energy intake, distinguishes between groups with known differences in diet quality, and has more than one underlying dimension. In addition to these population‐level assessments, exemplary menus expected to have high diet quality were scored with the HEI‐2010. Low correlations were observed between total and component scores with energy intake (|r|<0.21), demonstrating that the HEI‐2010 uncouples diet quality from quantity. Diet quality was lower among smokers (mean total HEI‐2010 score=45.7) compared to nonsmokers (53.3) (P<0.01), and exemplary menus received scores at or near the maximum levels, indicating construct validity. Cronbach's alpha was 0.68, supporting the reliability of the total score as an indicator of overall diet quality. Nonetheless, principal components analysis indicated multiple underlying factors, highlighting the importance of the component scores. Overall, the HEI‐2010 is a valid and reliable measure of diet quality. Support: NCI/CNPP.