bluuebunny/crossref_metadata_2025 huggingface dataset:
The possible role of O‐GlcNAc modification in the pathogenesis of depression disorder
Authors: Remigante Alessia , Morabito Rossana , Nagy Tamas , Spinelli Sara , Cordaro Marika , Marino Angela , Sarikas Antonio , Dossena Silvia
Published: April 2020

Diabetes mellitus leads to a number of complications involving brain function, including cognitive decline and depression. Depression is linked to impaired adult neurogenesis in the gyrus dentatus of the hippocampus, but the underlying molecular mechanisms are incompletely understood. The process of neurogenesis consists of the proliferation of neural progenitors and differentiation of new neurons, and the chloride conductance IClswell, activated after anisosmotic cell swelling during cell volume regulation, is thought to be essential in cell proliferation and differentiation. Diabetes also leads to increase in the post‐translational O‐GlcNAcylation (O‐GlcNAc) of cellular proteins. Recently, the protein ICln, which is crucial in the activation of the current IClswell, has been found to be O‐GlcNacylated. In the present study, we investigated the characteristic of the current IClswell following O‐GlcNAc elevation in neuronal‐like SH‐SY5Y and HEK 293 Phoenix cells, with focus on the role of ICln. In parallel to cell viability tests, IClswell was measured by patch‐clamp in native cells or cells overexpressing either the wild type or non‐glycosilable forms of ICln (IClnT223A, IClnS193X and IClnS67A). O‐GlcNAc elevation significantly reduced the metabolic activity of SH‐SY5Y and HEK 293 Phoenix cells. In SH‐SY5Y cells, O‐GlcNAc elevation significantly suppressed the basally activated IClswell current. In HEK 293 Phoenix cells, O‐GlcNAc elevation inhibited the ICln‐induced IClswell current, with no effect on the endogenous IClswell current, thus indicating that O‐GlcNAc impairs the ICln function. The IClnT223A‐induced current was similar to IClnWT‐induced current and was similarly suppressed after O‐GlcNAc elevation. IClnS193X lost most of its activity, though the residual current was sensitive to O‐GlcNAc elevation. Finally, the IClnS67A‐induced current is similar to the IClnWT‐induced current, but insensitive to O‐GlcNAc elevation. The present work underscores the essential role of O‐GlcNAcylation in governing the IClswell current through modification of the ICln protein. The results indicate that the O‐GlcNAc modification site responsible for the suppression of the ICln‐induced current should be located upstream the amino acid 193, most likely on Serine 67. We conclude that O‐GlcNAcylation of the IClswell activator ICln in the context of hyperglycaemic conditions may lead to IClswell inhibition in brain, which may result in reduced adult neurogenesis and eventually in the depression disorder. We suggest that the protein ICln may represent a novel target in the prevention or treatment of pathological states characterized by chronically elevated O‐GlcNAcylation of cellular proteins, including mood disorders linked to hyperglycaemia
Leukemia Inhibitory Factor Regulates the Post Stroke Immune Cell Infiltration in Aged Male and Female Rats
Authors: Appleton Malik
Published: April 2020

Ischemic stroke is one of the leading causes of death in the world. There currently only two FDA approved treatments for ischemic stroke, tissue plasminogen activator (tPA), and mechanical thrombectomy. Leukemia inhibitory factor or LIF, has been shown to improve functional recovery in young rats put under ischemic stroke conditions, but neither the responses of older rats or the gendered rat differences in immune responses have yet been tested with LIF. The purpose of this study is to identify the differences in immune cell infiltration of aged male and female rats subjected to MCAO and treated with LIF. For his study, the rats were subjected to either MCAO surgery or sham surgery, and given an injection of either LIF or PBS at 6, 24, and 48 hour time points after surgery. At the 72h time point the rats had MRI scans done on their brains immediately before euthanisia, and brains, blood, and spleens of the rats were collected for flow cytometry analysis. The spleen, blood, and brain samples were stained and analyzed with a flow cytometer for counts of Macrophages and Monocytes, B cells, and CD4 and CD8 T cells, and the resulting data was compiled using FlowJo and visualized using GraphPad. Results show that while there are some potential differences in the immune cell infiltrations of male vs female rats and PBS vs LIF rats, the deviation in sample levels of immune cells is on high for statistically significant conclusions to be made. A larger sample size for each group (currently n≤4 per treatment group) would be needed in a future study to investigate his further.

Support or Funding Information

KY‐WV Louis Stoke Alliance For Minority Participation (NSF #1826763), ASPET SURF Program at the University of Kentucky
Dual NMDAR signaling: flux‐independent pH sensor and flux‐dependent regulator of mitochondrial membrane potential (mΔψ) and cell membrane‐mitochondria bridging.
Authors: de Oca Pavel Montes , Hernandez-Cruz Arturo
Published: April 2020

Glutamate N‐methyl‐D‐aspartate (NMDA) receptor (NMDAR) is critical for neurotransmission as a Ca2+ channel. Nonetheless, several reports have demonstrated also flux‐independent signaling. Astrocytes express NMDAR distinct from its neuronal counterpart, but cultured astrocytes have no electrophysiological response and controversial findings have questioned NMDAR function. We recently demonstrated that in cultured astrocytes NMDA at pH6 (NMDA/pH6) elicits flux‐independent Ca2+ release from the Endoplasmic Reticulum (ER) and depletes mitochondrial membrane potential (mΔψ). Here we show that flux‐independent Ca2+ release is mainly due to pH6 and NMDAR, whereas mΔψ depletion requires both pH6 and flux‐dependent NMDAR signaling. Immunofluorescence exhibited that plasma membrane (PM) NMDAR is apposed or surrounds ER and mitochondria. Moreover, NMDA/pH6 treatment generated ER stress, increased endocytosis, mitochondria‐ER and ‐nuclear contacts and strikingly, PM invaginations near mitochondria along with electrodense structures referred here as PM‐mitochondrial bridges (PM‐m‐br). Our data and earlier observations strongly suggest PM‐mitochondria communication, as a proof of concept of this notion, NMDA/pH6 provoked mitochondria labeling by the PM dye FM‐4‐64FX. Finally, we analyzed by WB NMDAR subunit GluN1 to explore putative causes of NMDAR dual function, we found fragments with M.W. consistent with previously identified cleavage sites. Accordingly, GluN1 intracellular and extracellular domains presented little colocalization. Our findings demonstrate that NMDAR plays a dual function: a flux‐independent pH sensor and a flux‐dependent regulator of mΔψ. More importantly, mΔψ depletion seems to be mediated by PM‐mitochondria communication. Finally, we found different GluN1 fragments that could be involved in NMDAR dual signaling, although causality awaits demonstration (submitted to JBC).

Support or Funding Information

Supported by CONACyT project #132706 and PAPIIT‐UNAM AG200119.
Blood Flow Control of the Microcirculation by KATP Channels in Ventricular Myocytes
Authors: Zhao Guiling , Joca Humberto C. , Lederer W. Jonathan
Published: April 2020

The metabolic demands of the working myocardium underlies the control of blood flow to the cardiac myocytes. However, little is known about the details of the signals that are generated by metabolic need and how these signals are linked into a rapid‐response negative feedback control of blood flow. Here, using the state‐of‐the‐art confocal microscopy and whole mount immunostaining techniques and other fluorescent labels, we have investigated both the anatomic organization of the vasculature and the control mechanisms of blood flow. Our results show the key cellular elements of the system are connected to each other electrically by means of gap junctions (Connexin 43). They connect ventricular myocytes to capillary endothelial cells which are connected to pericytes and the small artery endothelium. Gap junctions thus allow the time‐averaged hyperpolarization of ventricular myocytes (due to the opening of ATP‐sensitive K+ channels (KATP) following metabolic need) to produce hyperpolarization in the rest of the system. By this means relaxation of the pericytes and the smooth muscle cells could produce an increase in blood flow with O2 and nutrition to the ventricular myocytes. Initial results showing this physiological feedback control system that appear to manage blood flow control in heart were tested in a perfused mouse cardiac papillary muscle preparation. The functional changes of arterioles and capillaries in the ex vivo heart were monitored using high‐speed confocal microscopes that include a combination of XY (Nipkow disk) and point‐scanner confocals. Our results suggest that KATP opener pinacidil caused perfusion pressure decrease (blood flow increase) while KATP blocker glibenclamide causes perfusion pressure increase in paced or quiescent heart, indicating the functional involvement of KATP channels in local blood flow control. Pinacidil‐induced perfusion pressure reduction does not differ on smooth muscle specific Kir6.1 mutated mice, suggesting the least role of smooth muscle KATP in the local blood flow regulation. An additional contribution to blood flow elevation may be attributed to extracellular K+ ([K+]o) increases (from ventricular myocyte KATP and other K+ channels), which leads to the hyperpolarization of capillary endothelial cells through the activation of inward rectifier K+ channels (Kir). Thus, the activation of KATP in cardiomyocytes caused by metabolic increase links to the local increase in blood flow. Taken together, these data provide compelling evidence that blood flow control in the mammalian heart is a classic negative‐feedback signaling mechanism that links the metabolism of ventricular myocytes to locally increased blood flow through an electrically controlled signaling system.
Crystal structure of a cross‐species reactive nanobody in complex with human serum albumin
Authors: Jian Mike , Brooks Cory L.
Published: April 2020

Nanobodies are single‐domain antibodies derived from the unique heavy‐chain antibodies found in camelid species. Nanobodies are attractive therapeutics as they are easy to produce, are highly stable, and can potentially bind targets inaccessible to traditional antibody formats. However, due to their small molecular weight, one disadvantage associated with nanobodies is rapid renal clearance resulting in a short serum half‐life. One innovative solution to increasing the serum half‐life of nanobodies is to make use of bispecific nanobodies. In this format, a therapeutic nanobody is fused to a second nanobody specific for the highly abundant plasma protein, human serum albumin. Such bispecific nanobody constructs targeting serum albumins have been shown to increase the half‐life of therapeutic nanobodies. The use of an albumin specific nanobody that cross reacts with albumin found in commonly used research animals is particularly desirable as the bispecific nanobody could be used pre‐clinical animal trials. Previously, we have generated a panel nanobodies which cross‐react with the serum albumins of humans, macaques, mouse, and rat, through immunization of a llama with human serum albumin. To determine the basis of the cross‐species albumin reactivity of the nanobodies we have determined the X‐ray structure of the nanobody M79 in complex with human serum albumin. M79 is cross reactive with serum albumins from macaques, mouse, and rat. The structure revealed of the three nanobody binding loops, only two of the loops (CDR2 and CDR3) interacted with human serum albumin. The M79 nanobody bound to the apex of subdomain IIB on human serum albumin, specifically forming polar contacts with residues D308, D314, N318, and E321. Comparison of the serum albumin sequences from human, macaques, mouse, and rat show that the residues in serum albumin that bound M79 are highly conserved amongst the species explaining the nanobodies’ cross‐reactivity. These results give insights on the cross‐species specificity of M79 towards serum albumin, and can bolster the use of M79 in animal model experiments.
Molecular Characterization of Fungi Associated with Pharmaceutical Products
Authors: Oyebanji Elizabeth Olawumi , Adekunle Adeyinka Adedotun
Published: April 2020

Pharmaceuticals are medicinal products used in the prevention, treatment, and diagnosis of diseases. As such, the presence of microorganisms’ especially fungal toxins can reduce or eliminate the product’s therapeutic activity and constitute a potential danger to patient health.

This study attempts to determine the fungal loads of liquid preparation used as medication in Lagos State and its environs and identify the fungal isolates. 252 different types of oral liquid drugs (200syrups and 52 suspensions) which included paracetamol syrup, cough syrup and antibiotics suspension manufactured in five different pharmaceutical industries in Nigeria were methodically sampled and analyzed for fungi contamination using standardized method. The isolated fungi were identified using morphological characterization as well as 16SrDNA sequencing. 13% of the sampled syrups were found to be contaminated with fungi where the colony forming units for paracetamol syrup, cough syrup and antibiotics suspension ranges from 2.0×101 – 9.6×104, 1.0×102–7.5×104 and 1.0×101–8.8×104cfu/ml respectively while 87% yielded no growth. Thirteen (13) morphologically different species of fungi were identified which included Aspergillus niger strains, A. tamari strains, A. japonicas, A. flavus, A. awamari, A. ellipticus, A. tubingensis, Meyerozyma caaribbica, Candida carpophila and Eurotiomycetes spp. The presence of microorganisms in oral liquid samples might explain the treatment complicacy of the diseased children. Therefore, microbiological quality of such drugs is thus suggested.
1. Modeling ovarian cancer for dendritic cells‐derived exosomes treatment
Authors: Pisano Simone , Gonzalez Deyarina , Ferrari Mauro , Conlan Steven , Corradetti Bruna
Published: April 2020

Ovarian cancer (OC) is the fifth leading cause of death from cancer among women in the United States and high‐grade serous carcinoma (HGSC) is the most common subtype. Patients often present metastatic disease with ascites buildup, which further complicates the treatment landscape. Although immunotherapy is a promising cancer treatment method, it fails to have an impact on HGSC patients due to the immunosuppressive and heterogeneous micro‐environment and impaired lymphatic drainage created by cancer cells within the peritoneal cavity. We are interested in boosting the anti‐tumor immunity by employing exosomes derived from dendritic cells (DC)‐based vaccines (Dex) for their capacity of overcoming the transport barriers, bypassing the immunosuppressive environment, prompting T cell infiltration in target organs, and activating them. Dex were collected and characterized in terms of yield, size distribution, and their biomolecular features as compared to parental cells, proving the expression of MHC‐II and co‐stimulatory markers (CD80 and CD86). When DC were pulsed with tumor antigen lysates from an ovalbumin‐overexpressing cancer cell line (ID8‐OVA), Dex presented OVA on their surface and activated the proliferation of OVA‐specific T cells (OT‐I). Intra‐peritoneal injection of Dex into a metastatic OC mouse model revealed a preferential accumulation within the peritoneal metastatic nodules, in tumors and in the liver. Preliminary data suggest Dex as a potential complement to the standard of care treatments for metastatic OC. We are currently testing their ability to induce an anti‐tumor immune response and increase survival when coupled with standard chemotherapeutics, with the potential of being effective in several metastatic diseases with peritoneal invasion.
Evidence for Neurohormonal Triggers of Cardiomyocyte Polyploidization and Cardiac Regenerative Potential Loss During Endothermy Acquisition
Authors: Huang Guo
Published: April 2020

Tissue regenerative potential displays striking divergence across phylogeny and ontogeny, but the underlying mechanisms remain enigmatic. Loss of mammalian cardiac regenerative potential correlates with cardiomyocyte cell‐cycle arrest and polyploidization as well as the development of postnatal endothermy. Genetic evidences in both mice and zebrafish support that cardiomyocyte polyploidization represents a major barrier limiting cardiac regenerative capacity. Through phylogenetic analyses of 41 vertebrate species, we uncover that certain monotreme, edentate, cetacean, chiropteran species have unusually high percentages of diploid cardiomyocytes in the adult heart. Furthermore, diploid cardiomyocyte abundance conforms to Kleiber’s law—the 3/4‐power law scaling of metabolism with bodyweight—and inversely correlates with standard metabolic rate, body temperature, and serum thyroxine level. Inactivation of thyroid hormone signaling reduces mouse cardiomyocyte polyploidization, delays cell‐cycle exit, and retains cardiac regenerative potential in adults. Conversely, exogenous thyroid hormones inhibit zebrafish heart regeneration. Notably, inhibition of thyroid hormone synthesis in mice after birth results in low body temperatures.

Recently, we discover the nervous system as a secondary major regulator of mammalian cardiomyocyte polyploidization, cell cycle arrest, and body temperature increase. Moreover, inhibitions of neuronal activity and thyroid hormone signaling in neonates have additive effects, yielding mice with the highest percentage of diploid cardiomyocytes and lowest body temperature to our knowledge. Thus, our results support a model where loss of cardiomyocyte regenerative capacity, driven by increasing neurohormonal activities, may be a tradeoff for the acquisition of endothermy during animal evolution and postnatal development.

Support or Funding Information

NIH
Effect of Levosimendan, an Inodilator, on Streptozotocin‐Induced Diabetic Nephropathy in Rats
Authors: Ali Badreldin H. , Al-Salam Suhail , Al Sulemani Yousuf M. , Abdelrahman Aly M.
Published: April 2020

Levosimendan, an inodilator, was shown to have a renal protective effect in different models of kidney dysfunction in experimental animals and in patients with heart failure. The aim of this study was to examine the effect of levosimendan on streptozotocin‐induced early diabetic nephropathy. Male Sprague Dawley rats (n = 24, 320–400 g) were randomly distributed into four groups and treated for 6 weeks. Groups 1 and 3 received either vehicle or levosimendan (1 mg/kg/day) for the last three weeks, respectively. Groups 2 and 4 were rendered diabetic by a single i.p. injection of streptozotocin (60 mg/kg) and were treated as in groups 1 and 3, respectively. At the end of the experiment, rats were placed in metabolic cages and urine was collected for biochemical analysis. Rats were then anesthetized and blood samples collected for biochemical analysis. Kidneys were removed for histopathology. In the untreated diabetic group, there was a significant decrease in body weight, polyuria and increased relative kidney weight. There was increased urinary albumin/creatinine ratio (UACR), 24 hour (h) albumin level and N‐acetyl‐β‐D‐glucosaminidase (NAG)/creatinine ratio (UNCR). Streptozotocin did not cause a significant effect on creatinine clearance when adjusted to body weight. In addition, diabetes was associated with increased oxidative stress [reduced plasma total antioxidant capacity (TAC) and catalase activities]. Kidneys from streptozotocin‐treated rats showed focal clear renal tubular cells affecting proximal convoluted tubules and focal mild interstitial fibrosis at the cortico‐medullary junction. Levosimendan significantly attenuated the reduction in body weight, polyuria and the increase in relative kidney weight. In addition, it significantly attenuated the increase in UACR, urinary 24 h albumin level and UNCR in diabetic rats. It also significantly mitigated the decrease in plasma TAC and catalase activities in diabetic rats. Rats in the diabetic group treated with levosimendan showed less clear cell changes but similar interstitial fibrosis when compared with the untreated diabetic group. This study shows that levosimendan ameliorated some of the changes seen in streptozotocin‐induced early diabetic nephropathy in rats. This could be partly due to its antioxidative effects.

Support or Funding Information

Supported by an internal grant from Sultan Qaboos University (IG/MED/PHAR/18/01)
Mesenchymal Stem Cell Derived Extracellular Vesicles Enhance Myelin Plasticity in Rhesus Monkeys
Authors: Go Veronica , Sarikaya Deniz , Zhao Yuxin , Bowley Bethany GE , Medalla Maria , Rosene Douglas L. , Buller Benjamin , Moore Tara L.
Published: April 2020

Cortical injury, such as injury from stroke, results in a cascade of events that includes cell death, inflammation and disruption of myelin. To date, there are no highly effective treatments for reducing the deficits that occur after injury. Recently, we have demonstrated that extracellular vesicles (EVs) harvested from rhesus monkey bone marrow derived cells when given 1 day and 14 days following injury facilitate recovery of function in aged rhesus monkeys within the first 3–5 weeks after cortical injury. Based on these findings and current proteomic literature of MSC‐EVs, we hypothesized that MSC‐EVs enhance myelin plasticity by limiting damage to oligodendrocytes and stimulating remyelination.

To assess general myelin integrity after injury, we used Spectral Confocal Reflectance Microscopy (SCoRe) to image myelinated axons and found an increase in the density of myelinated axons in the EV group (p < 0.05). To assess whether the difference was due to reduced damage or remyelination, in sublesional white matter we assessed immunohistochemical labeling of Olig2, a general oligodendrocyte marker, and 8OHdG, a marker for DNA damage. We found reduced densities of Olig2 colocalized with 8OHdG in the EV group (p<0.05). As a marker of active demyelination and myelin debris clearance, we measured Myelin Basic Protein (MBP) concentrations in CSF and found a longitudinal reduction in the EV animals. To assess remyelination, we measured expression of MBP, a gene for myelination in mature oligodendrocytes, Myelin Regulatory Factor (MyRF), a gene for oligodendrocyte differentiation and maintenance, and Breast Carcinoma Amplified Sequence 1 (BCAS1), a gene for newly myelinating oligodendrocytes. Interestingly, we found a 4 fold increase in MyRF expression, and a 1.5 fold increase in MBP and BCAS1 in the EV animals relative to the vehicle control animals in perilesional brain tissue. Consistent with these gene expression differences associated with re‐myelination, we found that the densities of newly‐myelinating oligodendrocytes immune‐labeled with BCAS1, as well as mature oligodendrocytes expressing CC1, exhibited a trend towards an increase in the EV group (p = 0.09). These results suggest that EV treatment reduces myelin damage, while also stimulating myelin repair. Finally, these results correlated with enhanced motor recovery, suggesting that EV‐mediated white matter plasticity is a critical component for recovery after cortical injury.

Support or Funding Information

Supported by NIH grant R21NS102991‐02.
Zika virus infection induces endoplasmic reticulum stress and apoptosis in placental trophoblasts
Authors: Muthuraj Philma Glora , Kraus Madison N , Sahoo Prakash Kumar , Bruett Taylor , Pattnaik Aryamav , Annamalai Arun Saravanakumar , Pattnaik Asit , Byrareddy Siddappa , Natarajan Sathish Kumar
Published: April 2020

Zika virus (ZIKV) infection in pregnant woman can be vertically transmitted to the fetus via the placenta leading to Congenital Zika syndrome (CZS). CZS is characterized by microcephaly, retinal defects and intrauterine growth retardation. ZIKV also causes placental pathology leading to severe compromise in the growth and development of the fetus. Further, the molecular mechanism behind ZIKV‐induced apoptosis in placental trophoblasts is unknown. We hypothesize that accumulation of viral proteins in the endoplasmic reticulum could lead to sustained endoplasmic reticulum stress (ER stress) and trigger apoptotic events.

Methods

HTR‐8, a human normal immortalized trophoblast cell and human choriocarcinoma derived cell lines (JEG‐3 and JAR) were infected with 0.1–1MOI ZIKV. Apoptosis was assessed by characteristic nuclear morphology staining with DAPI and caspase 3/7 activity.

Results

We observed an increase in the mRNA levels of CHOP and the spliced form of XBP1 gene, 16–24h post infection in trophoblast. We also observed an increase in the levels of ER stress markers such as p‐IRE1α, p‐eif2α, and activation of c‐Jun N‐terminal Kinase (JNK) and p38 mitogen‐activated protein kinase (MAPK) after 16–24h of ZIKV infection in
trophoblast. As prolonged ER stress can cause apoptosis, we observed a dramatic increase in trophoblast apoptosis 48h post infection. Mechanistically, inhibition of JNK by SP600125 or pan caspase by Z‐VAD‐FMK significantly blocked ZIKV‐induced apoptosis in trophoblast. In conclusion, the mechanism of ZIKV‐induced placental trophoblast apoptosis involves the activation of ER stress and MAPK activation.

Support or Funding Information

NIH: P20GM104320, Layman Seed award, UNL, Food for Health Research Initiative, UNL
Roles of Mitochondrial Dysfunction in Maleic Acid‐induced Acute Kidney Injury
Authors: Lin Hugo You-Hsien , Liang Chan-Jung
Published: April 2020

Background

Acute kidney injury (AKI) is a growing public health burden globally. Mitochondrial function plays the key role in the progress of AKI. Maleic acid (MA) has been proofed to induce various renal dysfunctions include AKI. In our previous study, there were MA affinity proteins, including ATP synthase subunits, mitochondrial glyceraldehydes‐3‐phosphate, been identified in human kidney proximal tubular cells (HK2 cells)
(1). The goal of this study is investigating mitochondria roles in AKI using our MA‐induced mitochondrial dysfunction AKI model

Methods

For the in vitro study, HK2 cells were induced with MA with 2mM and 5mM for 24 hours separately. For the in vivo study, we induced AKI in c57BL/6 mice by intraperitoneal MA injection with 400mg/kg body weight and sacrificed after 24 hours of injection.

Results

We found that the MA treatment induces HK2 cell death with dosage responsive in MA 2mM (p<0.01) and MA 5mM (p<0.001). Serum BUN and creatinine were all increased (p<0.05) in the MA‐treated mice when compared to the vehicle treated mice. The histological analysis showed increased tubule injury index Jablonski score (p<0.001), tubular dilatation areas (p<0.001) and greater casting area (p<0.001) in MA‐treated as compared to vehicle‐treated mice. AKI marker, Kidney Injury Molecule‐1 was increased (p<0.001) in MA‐treated as compared to vehicle‐treated mice. The flow cytometry analysis indicated increased apoptosis (p<0.001), cell reactive oxidative stress (ROS) (p<0.001), mitochondrial ROS (p<0.001), and decreased mitochondria membrane potential (p<0.001) in MA‐treated compared to vehicle‐treated in vitro. In O2 respiration, MA‐treated HK2 cells decreased the capacity of electro transfer system activity (p<0.001) and nonphosphorylating LEAK respiration (electron flow coupled to proton pumping to compensate for proton leaks) (p<0.001), compared to vehicle‐treated. There was also significantly decreased ATP production in MA‐treated compared to vehicle‐treated in vitro. In mitochondria structure, the cristae of mitochondria were lost in MA‐treated compared to vehicle‐treated under transmission electron miscroscope examination. The gene expression was confirmed with the significant lower of ATPase6 in MA‐treated compared to vehicle‐treated. Moreover, renal mitochondrial protein of ATP subunit C (p<0.05) were significantly decreased in MA‐treated compared to vehicle treated in vivo.

Conclusion

There results demonstrated that MA‐induced AKI through targeting renal proximal tubular mitochondrial ATP Synthase. Mitochondria may represent a novel therapeutic target for MA‐induced AKI.

Support or Funding Information
Unregulated mTORC1 activation in the liver: hepatic loss of Depdc5 and Tsc1 synergistically produces liver failure and injury
Authors: Lee Jun Hee , Cho Chun-Seok , Ho Allison , Namkoong Sim , Kim Myungjin
Published: April 2020

mTORC1 is an important metabolic regulator that responds to insulin/growth factors, nutrients and cellular stresses. DEPDC5 in the GATOR1 complex suppresses mTORC1 in conditions of limited nutrients, while TSC1 in the TSC1/TSC2 complex inhibits mTORC1 during insulin/growth factor deprivation and energetic stresses. Genetic variations in the DEPDC5 gene are associated with increased severity of liver pathologies during chronic hepatitis B and C virus (HBV/HCV) infection, which downregulates TSC1/TSC2. Using mice with liver‐specific Depdc5 deficiency, we examined how Depdc5 affects liver homeostasis, pathophysiology, and its interaction with Tsc1. Depdc5‐deficient mice exhibited enlargement of zone 3 hepatocytes, which progressed into liver damage, inflammation and fibrosis at later age. Double deletion of both Tsc1 and Depdc5 (DKO) produced synergistic mTORC1 activation, which was much more than what was observed from single Tsc1 or Depdc5 knockout mice. DKO mice experienced pronounced and widespread hepatocyte damage, and exhibited externally visible liver failure phenotypes, such as jaundice and systemic growth defects. Through systemic transcriptome analysis, we identified superoxide accumulation and oxidative damage as the main pathogenetic features distinguishing these mice from other mTORC1 models. Indeed, mTORC1 inhibition, as well as superoxide reduction, were sufficient to eliminate most pathological phenotypes we observed in mTORC1‐hyperactivated liver tissues. The current findings provide the genetic evidence supporting that growth factor/insulin‐regulated (Tsc1) and nutrient‐regulated (Depdc5) pathways genetically interact to coordinate mTORC1 activation. In addition, this multi‐pathway method for mTORC1 hyperactivation explains how DEPDC5 perturbations in humans aggravate HBV/HCV pathologies, which inhibit TSC1 and activate mTORC1.

Support or Funding Information

R01DK102850, R01DK111465 and R01DK114131 to JHL, F31DK117610 to AH, K01AG061236 to MK, T32AG000114, T32GM008322, P30AG024824, P30AR069620, P30DK034933, P30DK089503 and P30CA046592
Tissue‐engineered Vascular Grafts for Congenital Heart Surgery
Authors: Breuer Christopher
Published: April 2020

Objective

We developed a tissue engineered vascular graft (TEVG) formed by seeding autologous bone marrow‐derived mononuclear cells onto a biodegradable tubular scaffold fabricated from a knitted polyglycolic acid core and coated with a 50:50 copolymer of poly lactic acid and polycaprolactone. Once implanted, the scaffold degrades and neotissue forms giving rise to neovessel which is structurally and functionally similar to the native vessel into which it was implanted. Herein we will describe the natural history of the evolution of the mechanical properties of the TEVG after implantation and evaluate the impact of the mechanical properties of the TEVG on graft performance.

Methods

We utilized an ovine intrathoracic inferior vena cava model in order to evaluate the evolving mechanical properties of the TEVG and determine their impact on graft performance. We implanted TEVG into 24 juvenile lambs and serially assessed graft morphometry and function using serial angiography and intravascular ultrasound (IVUS) over a 6 ‐month to 1‐year period. We serially measured changes in graft size using both angiography and IVUS. We also assessed the graft hemodynamics at angiography. We measured graft displacement over both the cardiac and respiratory cycle. Finally, a subset of TEVGs were harvested and underwent biaxial mechanical testing.

Results

The TEVG undergoes significant remodeling when after implantation. As the scaffold degrades, the TEVG transforms from a relatively stiff conduit to a highly compliant neovessel with growth capacity.

Conclusions

The transformation of the TEVG from a relatively stiff vascular conduit to a more compliant neovessel enables the graft to function as a capacitance vessel improving its performance when used in the venous circulation.

Support or Funding Information

NIH R01HL139796
Developmental Basis and Consequences of a Key Innovation in Lungless Salamanders
Authors: Hanken James , Lewis Zachary
Published: April 2020

A longstanding goal of evolutionary developmental biology is to document the precise mechanisms that underlie formation of, and differences among, basic body plans, organ systems and physiological functions. Another, less‐studied approach is to reveal how those mechanisms are perturbed to enable, or even mediate, the origin of novel adaptations that are critical for the evolutionary success of individual clades. I will report results of our recent studies of evolutionary lung loss in plethodontid salamanders and of the correlated evolution of cutaneous respiration, which is the principal mode of respiratory gas exchange in all 478 species in this, the largest family of extant urodeles. Surprisingly, plethodontids are not entirely lungless: a lung rudiment forms initially in embryos but is resorbed before hatching. Some embryonic cells express lung‐associated molecular markers, such as the gene Nkx2.1, although at lower levels than in lunged species. Moreover, elevated levels of TGF‐ß signaling, which is known to mediate Nkx2.1 expression during normal lung development, may be part of the genetic mechanism responsible for lung loss. In contrast, we have discovered in salamanders a novel paralog of the gene surfactant‐associated protein C (SFTPC), which is a critical component of pulmonary surfactant expressed exclusively in the lung in other vertebrates. In plethodontids, however, and only in plethodontids, this gene is expressed in novel sites of respiration outside the lungs and at different sites in larvae vs. adults that correlate with gas exchange. These and other changes to the ancestral urodele ontogeny may have contributed to the remarkable evolutionary radiation of the lungless salamanders.
Assessing and Evaluating Anatomy Instruction: Strategies for Anatomy Educators
Authors: Linse Angela
Published: April 2020

Effective instruction is crucial to student learning in anatomy education. While anatomy teaching contexts have expanded and accreditation standards have become more focused on student learning, teaching evaluation practices have not changed. Across the US, higher education institutions have come to rely primarily or solely on students’ ratings of instruction to evaluate teaching. While more than 80 years of research has demonstrated that end‐of‐term student ratings provide results that accurately and reliably reflect students’ collective perceptions of the learning environments created by their instructors, researchers have long recommended that students’ perception data not be the only source of evidence used in the evaluation of teaching effectiveness. Students’ ratings are important and should not be excluded, but they should never be the only source of information used to evaluate teaching. Anatomy faculty do have options. In this session, we will discuss actions faculty can take to use student ratings data for formative assessment (i.e., improvement before evaluation by administrators or colleagues), as well as strategies that can be used to corroborate (or contradict) student ratings used in promotion or reappointment processes. Faculty participants in this session will learn about strategies for controlling the narrative about their teaching through systematic interpretation, targeted responses to ratings, and collection of additional evidence of teaching effectiveness from other sources. Administrators and peers will realize that they have the power to to ensure that student ratings data are used appropriately in evaluative contexts.
Students as Evaluators of their Learning Experience
Authors: Rosenberg Martina
Published: April 2020

Student evaluations of teaching (SET) have been an evergreen topic of passionate debates in higher education. Concerns have been raised about use student feedback in the form of SETs for administrative purposes. Nevertheless, they remain a reality at most institutions and are frequently the sole or key metric on which teaching excellence is evaluated.

This session will provide an overview of national trends with the struggle to define teaching excellence in context and the current discussion on what aspects of the teaching and learning can be evaluated with different SET formats. Viewed through the lens of academic development, participants will examine why SETs are so pervasive, how to mitigate the emotional charge that is a barrier to individual growth of instructors within their professional setting, and how students can act as productive partners in institutional learning.
Weaving Histology into the Fabric of Medical Education: Histology as a Foundation for Integration and Application
Authors: Goldman Haviva M
Published: April 2020

Over the past two decades, histology education in the health sciences has undergone a dramatic shift both in method (use of virtual microscopy) and curricular focus (decrease in hours; increased integration). While this new landscape introduced challenges for histology teaching, histology’s strengths as a conceptual link between molecular/cellular level mechanisms and organismal structure and function make it an ideal springboard for integrating multiple disciplines in creative ways, allowing the discipline to play a key role in building both foundational knowledge and clinical diagnostic skills.

At Drexel University College of Medicine, histology is taught as part of an integrated curriculum that takes a sequenced normal (yr 1)/abnormal (yr 2) approach. Content delivery utilizes a flipped‐classroom approach, and histology is integrated within four different types of delivery methods including integrated online ‘Independent Learning Modules’; Histology focused (but integrated) ‘Laboratory Reviews’ including scavenger hunt and mystery slide exercises; multidisciplinary ‘Think‐Pair‐Share’ sessions; and multidisciplinary ‘Team Based Learning’ sessions that focus on clinical application. Specific examples of each of these integrated modalities will be presented with a focus on highlighting the challenges and opportunities these approaches afford from the perspective of the discipline, the educator, the student, and ultimately the clinician.
Early Signals in Wound Healing
Authors: Niethammer Philipp
Published: April 2020

Reactive oxygens species (ROS) and polyunsaturated fatty acids (PUFAs) are conserved wound signals throughout phylae. In zebrafish larvae, ROS gradients and osmotically released arachidonic acid mediate rapid wound detection by afar leukocytes. But it is unclear, whether these cues initiate de novo leukocyte recruitment like alarmins or rather amplify responses to other initiating signals. This talk will summarize our recent progress on this question, outlining a novel role for lipid peroxidation gradients as physiological integrators of ROS‐ and PUFA‐mediated early wound cues.

Support or Funding Information

NIH‐R21AI139986NIH‐R01GM099970NIH‐R01GM127356
Developing Novel Means to Inhibit Metastasis Through Targeting Tumor Heterogeneity and EMT
Authors: Ford Heide L. , Kong Deguang , Neelakantan Deepika , Zhou Hengbo , Hughes Connor , Lewis Michael , Marugan Juan , Patnaik Samarijit , Zhao Rui
Published: April 2020

It is well established that a subset of cells within primary breast cancers can undergo an epithelial to mesenchymal transition (EMT), although the role of EMT in metastasis remains controversial. We previously demonstrated a novel mechanism by which EMT contributes to metastasis, via mediating tumor cell crosstalk. In this seminar, I will discuss how breast cancer cells that have undergone an oncogenic EMT can increase metastasis of neighboring epithelial cancer cells via non‐canonical paracrine‐mediated activation of GLI activity that is dependent on Six1 expression in the EMT cancer cells. I will discuss how VEGF‐C, secreted by breast cancer cells that have undergone an EMT, promotes paracrine‐mediated increases in proliferation, migration, invasion, and metastasis of epithelial breast cancer cells, via non‐canonical activation of GLI‐signaling. Importantly, the aggressive phenotypes imparted by EMT cells on adjacent epithelial cancer cells can be disrupted by either inhibiting VEGF‐C in EMT cells or by knocking down NRP2, which interacts with VEGF‐C, in neighboring epithelial cancer cells both in vitro and in vivo. Interrogation of TCGA and GEO public datasets supports the relevance of this pathway in human breast cancer, as VEGF‐C and GLI pathway genes significantly positively correlate. Our study suggests that the VEGF‐C/NRP2/GLI axis may be a novel and conserved paracrine means by which EMT cells enhance metastasis, and provides potential targets for therapeutic intervention in this heterogeneous disease. In addition, because SIX1 activates VEGF‐C expression which leads to enhancement of metastasis via cell autonomous and non‐cell autonomous means, we have developed an inhibitor that targets SIX1 action and thereby inhibits metastatic disease. We describe results using this inhibitor in cell culture, where it inhibits SIX1 function and EMT, and in animals, where it inhibits metastasis without an effect on primary tumor growth.

Support or Funding Information

Colorado Cancer Translational Research Accelerator (CCTRA) PI Heide L. Ford, Co‐I Rui ZhaoNCI R01 CA224867 PIs Heide L. Ford and Michael T. Lewis
The Development of Appendicular joint cartilages in Alligator mississippiensis: Evolutionary and Biomechanical Implications for Archosauria
Authors: Tsai Henry P.
Published: April 2020

Crocodylians and birds are the two surviving lineages within Archosauria, a clade of morphologically and ecologically diverse vertebrates that evolved highly disparate locomotor postures. The appendicular joints of crocodylians and birds differ substantially, thus complicating inferences of ancestral morphology and evolutionary transitions. Recent studies have indicated that the American alligator, an extant crocodylian, use a wide spectrum of cartilaginous and other connective tissues to construct the hip joint. However, little is known about the distribution and use of cartilage throughout other appendicular joints, as well the ontogenetic transition of the crocodilian chondro‐osseous interface during early development. In particular, the hyaline cartilage on the proximal femur of late embryonic alligators possesses a convex, cone‐like extension that inserts deep into the metaphysis. This morphology is not seen in juvenile and adults, but is superficially similar to the cartilage cone‐metaphyseal trough articulation inferred in early members of multiple archosaurian groups. The method by which alligators and other archosaurs reduced the cartilage cone during ontogeny and evolution, as well as the functional significance of the cartilage cone itself, are poorly understood.

This study examined the ontogenetic transitions of proximal appendicular joints in the American alligator, spanning from in ovo stages to the first year of life. We used dissection, histology, and imaging techniques to assess the articular soft tissue anatomy at the chondro‐osseous junction of the proximal‐ and distal epiphyses of the femur and humerus. Additionally, osteological correlates for joint cartilage were identified on fully skeletonized limb bone elements and compared with bony morphology seen in fossil archosauromorphs.

At the onset of hatching, the alligator did not reduce the cartilage cone via decreasing the “depth” of cartilaginous extension into the metaphysis, as inferred for early archosaurs. Instead, neonatal alligators regress the cartilage cone via multiple invasion fronts of trabecular bones, such that the subchondral growth plate of hatchlings presents a convex surface perforated by numerous cartilage canals, similar to those found in young birds. These results suggest that the ontogenetic loss of cartilage cone in the alligator is different from the evolutionary loss of cartilage cone hypothesized for early archosaurs. Instead, the role of cartilage thinning, fibrocartilage sleeve development and calcified epiphyseal cartilage may be highly associated within different sauropsid lineages in response to evolutionary changes in joint loading regimes.

Support or Funding Information

This research is supported by the Missouri State University Faculty Research Grant
Incorporating Trace Fossils when Exploring the Interplay of Form and Function in the Crocodyliform Feeding Apparatus
Authors: Drumheller Stephanie K.
Published: April 2020

Many crocodylians, both living and extinct, fill the semi‐aquatic ambush predator niche, and their prey preferences roughly can be predicted based on a combination of snout shape, tooth morphology, and body size. Among extant crocodylians, slender‐snouted taxa such as Gavialis gangeticus and Tomistoma schlegelii generally prefer smaller‐bodied prey, whereas the broader‐snouted taxa divide into generalists like Crocodylus niloticus and macro‐generalists like Alligator mississippiensis, who will eat a diversity of prey items including animals that are as big or even bigger than themselves. Large‐bodied members of these generalist ecomorphotypes generate the highest bite forces observed in modern taxa and are often apex predators in their ecosystems. The availability of a robust phylogenetic hypothesis for crocodylians makes them an excellent clade with which to explore these types of ecomorphological hypotheses, which link form and function in the feeding apparatus within an evolutionary framework. The resulting synthesis of extant and extinct datasets has provided both exciting new insights, but also some complications. The addition of extinct taxa into studies of morphological disparity, both within the crown group and deeper in the crocodyliform evolutionary tree, introduce a diverse array of snout shapes and associated niches that are not represented in the surviving members of the clade (e.g. fully marine, fully terrestrial, durophagous, omnivorous, and even possibly herbivorous forms). Though hypotheses regarding diet in these diverse groups are based on craniodental morphology, indirect evidence, in the form of trace fossils, provides an independent means of testing those predictions. Gut contents are extremely rare in this clade because of the highly destructive digestion of crocodyliforms, and coprolites, though comparatively common, contain remains that are so degraded as to be largely unidentifiable and uninformative. Isotopic and dental microwear studies can provide broad patterns of diet. However, for detailed, direct evidence of crocodyliform feeding, bite marks are a particularly useful source of direct evidence with which to explore patterns of diet in this clade. A recent, concentrated effort to characterize modern crocodylian bite marks has resulted in the identification of novel patterns and diagnostic traces associated with feeding behavior in that clade. These trace fossils provide critical insights into diet, feeding strategy, and even intraspecific competition in crocodyliforms and their extinct relatives. By integrating these ichnological data with traditional dietary hypotheses based on morphology and phylogeny of extinct groups, a more complete, and oftentimes more complex and realistic, concept of past ecosystems and trophic structures can be explored.

Support or Funding Information

Funding provided by the National Science Foundation, the Leakey Foundation, and National Geographic.