bluuebunny/crossref_metadata_2025 huggingface dataset:
Cardiac Gene Expression and Histology in a Rat Model of Fat Embolism
Authors: Wacker Michael J. , Patel Shaan , Vallejo Julian , Colson Jordan , Edegbe Joy , Lin Jane , Salzman Gary , Poisner Alan , Monaghan A. Paula , Molteni Agostino
Published: April 2020

Introduction

Fat Embolism consequent to Triolein (T) intravenous injection in rats induces severe histopathologic pulmonary damage at 48 hours mostly related to vasculitis, peribronchial and septal inflammation with an increase in mast cells, and eventually, organ fibrosis. We previously reported that the hearts of the same animals at 48 hours did not show vasculitis of coronary arteries branches, inflammation or an increase in mast cells. The objective of this study was to determine if T treatment by 10 weeks induced changes in cardiac gene expression or histopathology related to fat embolism.

Materials and Methods

24 Sprague Dawley Rats (250–330g) were divided into two groups, 48 hour treatment and 10 week treatment (N=12 each). Six rats of each group were intravenously injected with 0.2 ml of T, and the other 6, 0.2 ml of normal saline. The rats were euthanized under general anesthesia 48 hours or 10 weeks after injection. A cut was made in hearts along the transverse plane; the top half of hearts were snap frozen in liquid nitrogen for gene expression studies and the bottom half was fixed in formaldehyde for histopathological evaluation similar to lung preparations. After RNA isolation, real time RT‐PCR was performed on genes associated with pathological cardiac hypertrophy or fibrosis (beta myosin heavy chain, alpha myosin heavy chain, skeletal muscle alpha actin, atrial natriuretic peptide, brain natriuretic peptide, vimentin, and collagen 1a1).

Results

Despite significant damage to the lungs, there was no significant changes in expression of any of these genes in the hearts of the rats treated with T, versus those of the saline treated group, whether sacrificed at 48 hours or 10 weeks post injection (P>0.05 ANOVA). Histopathological evaluation of the hearts at 10 weeks also did not show any damage in the myocardium in sections stained by H and E and no vasculitis, mast cell increase, or ventricular enlargements. Trichrome staining did not show any fibrotic increase in hearts of T injected animals, thus confirming the previous observations and gene expression analysis.

Conclusion

There was a marked contrast in organ damage between hearts and lungs in this study in rats with T treatment. Hearts demonstrated no significant change in gene expression or histology demonstrating a lack of fibroblast infiltration, hypertrophy, or remodeling due to T treatment. Kidney, liver, brains, and aortas also showed varying degrees of histopathological damage compared to the severity of the damage of the lungs. The renin angiotensin system plays a key role in the pathogenesis of the lung damage with fat embolism and this system needs to be further explored for organ differences to fat embolism induced by T treatment.

Support or Funding Information

Supported by the Catherine T. Geldmacher Foundation, St. Louis, MO
Quantitative 3D Reconstructions of the Human Superior Olive Complex are Consistent with Auditory Dysfunction in Autism Spectrum Disorder
Authors: Kulesza Randy , Mansour Yusra
Published: April 2020

Beyond the characteristic social, communication and behavioral aberrations associated with autism spectrum disorder (ASD), the vast majority of subjects with this condition have some degree of auditory dysfunction. The auditory deficits range from complete deafness to hypersensitivity but typically include difficulty listening to speech in the presence of background noise. Clinical testing of brainstem evoked potentials and stapedial reflexes in children and adults with ASD reveal lower thresholds but slower conduction times and right‐left ear asymmetries. Previous studies on post‐mortem brain specimens from our lab revealed drastic and consistent hypoplasia and dysmorphology in the auditory brainstem. The most drastic alterations were found in the medial superior olive (MSO), the largest and most prominent component of the superior olivary complex (SOC). Our morphological studies have shown that in subjects with ASD, there are significantly fewer neurons in the MSO and surviving neurons are smaller, more round and have an abnormal orientation. Based on these results, we hypothesized that the SOC in subjects with ASD not only includes fewer neurons but that these nuclei occupy significantly less brain volume and demonstrate abnormal nuclear contours. We investigated this hypothesis by obtaining donated brain tissue from an age‐matched series of subjects with ASD or no known neurological diagnosis (neurotypical; NT) through Autism BrainNet. Free‐floating tissue sections were mounted onto glass slides, stained for Nissl substance with Giemsa and sealed under coverslips. 3D volume renderings of the SOC nuclei were created in Amira. Subjects in this study include two NT subjects (ages 4 and 8, both female) and four age‐matched subjects with ASD (ages 2–11 years of age, all male). At autopsy, there was no difference in total brain weight between NT and ASD subjects (p=.42). Our 3D reconstructions reveal a drastic reduction in the total volume of the SOC in subjects with ASD (control = 6.8 mm3, ASD = 2.23 mm3; p<.005). In fact, all SOC nuclei were significantly smaller in ASD subjects expect for the ventral nucleus of the trapezoid body (p=.922). Together, these results support the brainstem as a core site of pathology in ASD and are consistent with previous clinical evaluations of hearing in subjects with ASD.

Support or Funding Information

Autism BrainNet
Lipocalin 2 Induces Neuroinflammation through Liver‐Brain Axis in Nonalcoholic Fatty Liver Disease
Authors: MONDAL AYAN , SETH RATANESH , SARKAR SUTAPA , BOSE DIPRO , SAHA PUNNAG , KIMONO DIANA , ALBADRANI MUAYAD , NAGARKATTI MITZI , NAGARKATTI PRAKASH , CHATTERJEE SAURABH
Published: April 2020

Nonalcoholic fatty liver disease (NAFLD) is chronic liver disorder characterized by accumulation of excess fat in liver due to consumption of high fat/lipid diets. A multiple hit paradigm that include lipotoxicity, environmental and genetic causes for prolonged period often leads to nonalcoholic steatohepatitis (NASH) with scarring and inflammation in liver that consequently causes irreversible hepatic damage. Both clinical and basic research implicated a strong correlation between NAFLD/NASH phenotypes with several ectopic manifestations like type 2 diabetes, cardiovascular disease, chronic kidney disease and more recently neuroinflammation and neurodegeneration. Notably, mechanisms of neuroinflammation in underlying NAFLD are poorly understood. In this report we identified adipokine Lipocalin 2 (Lcn2) induced neuroinflammation by acting through liver‐brain axis in diet induced murine NAFLD/NASH model. We found that Lcn2 is overexpressed in liver and in portal circulation that activated 24p3R (Lipocalin2 receptor) in brain and induced the release of high mobility group box 1 (HMGB1) from brain cells. Released HMGB1 acted as a preferential ligand to Toll like receptor 4 (TLR 4) and induced oxidative stress by activation of NOX‐2 signaling. NOX2 downstream signaling activated NF‐kB by phosphorylation of p65 protein. Activation of p65 and NOX‐2 led to NLRP‐3 inflammasome activation which subsequently released proinflammatory cytokines IL‐6 and IL‐1β from brain cells. Furthermore, we show that elevated Lcn2 level from neurons in underlying NAFLD caused a non disruptive blood brain barrier dysfunction by over production of TNFα, IL‐1β and MMP 9 without alteration of tight junction proteins Claudin 5 and Occludin. Based on the above, the above findings identify Lcn2 as a potent molecule inducing neuroinflammation in underlying NAFLD.

Support or Funding Information

NIH#P01ES028942‐01 to Dr. Saurabh Chatterjee
Nocodazole‐induced growth inhibition of human leukemia cells is linked to downregulation of CAK molecules
Authors: Almeida Adrian , Baquier Jocelyn , Hu Xiaotang
Published: April 2020

A cytoskeleton is present in the cytoplasm of all living cells. The cytoskeleton is composed of three different types of fibers: microtubules, microfilaments, and intermediate. The microtubules play an important role in a variety of cellular processes including maintaining cell structure and pulling chromosomes apart during anaphase. Nocodazole is an inhibitor of microtubule polymerization. Inhibition of microtubule polymerization by Nocodazole often lead to arrest of cells in G2 or M phase. However, it is unclear if any cell cycle regulatory molecules are involved in Nocodazole‐mediated cell growth. In this study we focus on CDK activating kinase (CAK). The CAK is composed of three subunits, CDK7, cyclin H, and MAT1. The cyclin H is regulatory subunit of CDK7 whereas MAT1 determines the specificity of CDK7. Addition of Nocodazole significantly inhibited the growth of MV4‐11 and TF‐1a cells in culture. The inhibitions were dose‐dependent with maximal inhibition being observed at a concentration of 10uM of Nocodazole. Less than 1 μM had no significant inhibitory effect on the growth of the cells, measured by XTT proliferation assay. Paralleled to the changes in the cell proliferation, expression of CDK7 and cyclin H were marked decreased when Nocodazole were added to TF‐1a and MV4‐11 cells at concentrations of 5 and 10μM. Similarly, Nocodazole at concentration of 10μM downregulated the phosphorylation levels of CDK7 and the expression of RNA polymerase II in TF‐1a cells. However, Nocodazole had no effect on the expression of MAT1, suggesting that Nocodazole did not interfere with specificity of CDK7. We also found that Nocodazole had no effect on the expression of β‐actin, a non‐muscle cytoskeleton actin.

Support or Funding Information

Barry University
Infarct macrophage secretome coordinates neutrophil degranulation
Authors: Daseke Michael , Kalusche Will , Flynn Elizabeth , Konfrst Shelby , Lindsey Merry
Published: April 2020

Following myocardial infarction (MI), the left ventricle undergoes wound healing that begins with an intense inflammatory response to recruit leukocytes to the infarcted region. Infiltrated neutrophils degranulate, releasing a series of proteases including matrix metalloproteinase (MMP)‐9 into the extracellular space. Macrophages also infiltrate into the infarct region, and we hypothesized that the macrophage may secrete factors that regulate neutrophil degranulation. Stimulating bone marrow derived neutrophils from C57BL/6J mice (3–6 month old male) with phorbol myristate acetate (PMA) induced degranulation, as evidenced by release of MMP‐9. Co‐stimulation with MI day 1 macrophage secretome (10% by volume) reduced PMA‐induced degranulation of MMP‐9 by 8‐fold (p<0.0014). Transcriptomics analysis of day 1 MI macrophages revealed galectin‐3, vimentin, fibronectin, and murinoglobulin‐1 were highly overexpressed. Examination of the day 1 MI macrophage secretome also showed high expression of vimentin and fibronectin. To further unmask signaling connections, neutrophils were co‐stimulated with PMA and day 1 MI macrophage secretome, along with blocking antibodies for the 4 proteins individually. Galectin‐3 or fibronectin inhibition promoted while murinoglobulin‐1 inhibition reversed the macrophage secretome effect. Neutrophil stimulation with recombinant galectin‐3 or fibronectin showed an additive effect with PMA on MMP‐9 release. Vimentin inhibition or stimulation had no effect on neutrophil degranulation. Overall, our results uncovered a role for galectin‐3 and fibronectin in inducing neutrophil degranulation while murinoglobulin‐1 may be a potent inhibitor of degranulation. The day 1 MI macrophage, therefore, secretes both factors that promote as well as factors that temper neutrophil degranulation. The balance of these factors determines the net effect of the macrophage secretome on neutrophil degranulation.

Support or Funding Information

NIH HL075360, HL129823, HL13731, VA 5I01BX000505.
X‐ray Crystal Structure of Agmatinase from Escherichia coli
Authors: Ahmed Syed Fardin , Chitrakar Iva , Torelli Andrew T. , French Jarrod B.
Published: April 2020

Agmatinase acts upon the cellular metabolite agmatine to produce putrescine, an important precursor for higher‐order polyamines that are required for cell proliferation, differentiation, and migration. The activity of agmatinase and the role of agmatine have garnered attention due to the importance of agmatine in various medical contexts including: neurological disorders, a possible role for agmatine as a neurotransmitter/neuromodulator, treatment of depression, and even the proposal that the family of metallohydrolases that include agmatinase may find novel biotechnological applications. Here we report the crystal structure of Escherichia coli agmatinase. The structure of E. coli agmatinase exhibits global fold and active site architecture typical of the metal‐dependent ureohydrolase protein superfamily. Moreover, the arrangement of conserved metal ligands and electron density at the active site indicate the presence of two metal ions, which is consistent with reported literature finding E. coli agmatinase to require two Mn2+ ions. Comparisons with the structures of related proteins will be described, along with functional characterization of agmatinase in terms of both enzymatic activity, as well as the requirement and identity of metal ions.
Comparing Cortactin Expression in MDA‐231 and A549 Cancer Cells
Authors: Adofo Emily , Kruchten Anne , Talaga Melanie L.
Published: April 2020

Cortactin is an actin‐binding protein that is associated with the progression of cancer. It is over‐expressed in many cancers including melanoma, colorectal cancer, breast cancer and glioblastoma. Cortactin is an important biomarker for aggressive cancers. Past research has focused on the post‐translational modifications (PTMs), acetylation and phosphorylation of expressed cortactin in prokaryotic cells. Studies also show that cortacin is overexpressed in many prokaryotic cells altering cell migration, invasion and metastasis. The structural differences of cortactin expressed in eukaryotic cells is a blank canvas. In addition, previous research regarding the role of cortactin in cell migration has remained inconsistent. To gain a better understanding of cortactin, a confluent culture of MDA‐231 human breast cancer cells and A549 lung cancer cells was used. Cortactin was purified from the cell lysate using a combination of size exclusion and ion exchange chromatography. Cortactin produced by cancer cells was visualized on SDS‐PAGE and Western Blot. Our current research focuses on gaining a better understanding of the diverse properties of cortactin when purified from cancer cells. The expression of cortactin in cancer cells enables our research to focus on studying the PTM, glycosylation to gain a better understanding of cortactin and its role in aggressive cancers.
Bacterial cell wall modification by glycolipid probes
Authors: Marando Victoria M. , Calabretta Phillip J. , Hodges Heather L. , Kraft Matthew B. , Kim Daria E. , Kiessling Laura L.
Published: April 2020

Despite the ubiquity and importance of glycans in biology, methods to probe their structures in cells are limited. The structures of mammalian glycans can be perturbed through metabolic incorporation, in which non‐natural sugars are taken up by cells, converted to nucleotide‐sugar intermediates, and incorporated via biosynthetic pathways. Applications of metabolic incorporation in bacteria are complicated by the diversity of pathways bacteria use to acquire, metabolize, and catabolize monosaccharide intermediates. We developed a strategy to alter glycan structure that avoids intracellular processing using lipid‐linked glycans, intermediates directly used in glycan biosynthesis. We generated synthetic arabinofuranosyl phospholipids to test this strategy in Corynebacterium glutamicum and Mycobacterium smegmatis, organisms that serve as models of Mycobacterium tuberculosis. Using a C. glutamicum mutant that lacks arabinan, we identified glycolipid donors whose addition restores cell wall arabinan, demonstrating that non‐natural glycolipids can serve as biosynthetic intermediates and function in chemical complementation. Our lipid‐based probe could rescue wild type cells treated with an inhibitor of cell wall biosynthesis. We then focused on incorporating derivatives with chemical handles for labeling such that bacterial glycans can be captured and visualized. To this end, we have generated the first probe to label arabinose‐containing structures in mycobacteria and corynebacteria.

Support or Funding Information

NIH: RO1‐AI063596, NSERC PGS‐D
Anti‐inflammatory effects of vagus nerve simulation in a newborn rat model of acute inflammation
Authors: Cacho Bradley , Abdala Jonathan , Wilson Christoper G.
Published: April 2020

Neonatal sepsis and inflammation pose a potential source of morbidity and mortality in newborns, specifically in the first weeks of life. We have previously shown that vagus nerve stimulation modulates pro‐inflammatory cytokine expression in the Central Nervous System (CNS). The optimal vagus nerve stimulation (VNS) parameters to reduce inflammation have been unknown.

In this study, we tested the hypothesis that vagus nerve stimulation reduces the expression of pro‐inflammatory cytokines in a lipopolysaccharide (LPS) model of airway inflammation. To test our hypothesis, we used intratracheal injections of LPS (0.5 mg/kg in 10 μL of saline) in neonatal Sprague Dawley rats (postnatal days 10 to 12) to induce pro‐inflammatory cytokine production. The P10–12 pups were anesthetized with isoflurane (2 to 4%) and then LPS was injected into the trachea to mimic airway inflammation. The experimental group was subjected to VNS at pre‐selected frequencies (ranging logarithmically from 10 to 10000 Hz) continuously for 30 minutes immediately after the LPS injection. Stimulation current was titrated down as needed to account for irregular breathing or apneas (typically approximately 0.5 mA). After recovery, each rat’s brainstem was collected and processed for cryosectioning and immunohistochemistry. We stained for and measured the expression of IL‐1 beta, TNF alpha, and HMGB1 in subjects with or without VNS (the positive control group received LPS without VNS). We then used ImageJ software and the particle analysis plug‐in to quantify the number of cells that expressed cytokines in the nucleus tractus solitarius (NTS).

Using one‐way ANOVA analysis, our data showed a significant decrease (p < 0.05) in expression of IL‐1 beta, TNF alpha, and HMGB1 in LPS‐injected rats that underwent VNS compared to LPS‐injected rats that did not receive VNS. Frequencies of 10, 100, 1000, and 10000 Hz all resulted in significant decreases in cytokine expression. Cytokine expression in the LPS + VNS group was similar when compared to sham rats (no LPS injection or VNS). Our study showed that VNS significantly decreased the expression of the early inflammatory cytokines in LPS‐induced inflammation in neonatal rats. Blocking the early cytokine cascade may be associated with decreased inflammation and a concomitant reduction in associated symptoms in patients with sepsis. VNS used either alone or in conjunction with antibiotics may lead to improved outcomes in neonatal patients suspected to be affected by infection.

Support or Funding Information

This study was supported via a grant funded by the Pediatric Research Initiative at Loma Linda University.
Rice bran supplementation improves glucose tolerance and innate immunity in high‐fat diet induced obese ovariectomized mice
Authors: Chen Li Hsin , Yeh Chiu Li
Published: April 2020

Objectives

Menopause is a state of low estrogen production which is associated with chronic low‐grade inflammation and higher incidence of overweight. Rice bran (RB) is a nutrient‐dense product of rice milling process that is considered a health improving supplement. The aim of this study was to investigate the influences of RB on the innate immunity of high‐fat diet induce ovariectomized (OVX) obese mice.

Methods

8 months‐old female mice were randomly divided into OVX sham (SH) group and OVX group. After surgery for one week, mice were randomly assigned to one of the three groups (n=12) and fed the respective diet for four months : SH group and OVX‐control (ONC) group were provided with AIN‐95 control diet containing 10% energy as fat, OVX‐high fat diet (OHF) group was treated with high fat diet (HFD) containing 40% fat, mice in the OVX‐high fat diet with RB (OHFR) group were fed with HFD containing RB (10% wt/wt).

Results

Mice in the OHF group showed significantly higher body weight gained, plasma glucose and triglyceride levels than those in the SH and other OVX groups. Compare to the SH and ONC groups, the OHF group had lower splenic natural killer (NK) cells cytotoxic capacity and peripheral blood phagocytes phagocytic ability. The chemotactic ability of leukocytes were also lower in the OHF group. As regard to the OVX group with high‐fat diet, RB supplementation increased NK cells cytotoxic function and phagocytosis ability of phagocytes. Also, the chemotactic ability of neutrophils and macrophage as measured by migration speed and directional persistence were improved in the OHFR group.

Conclusion

The findings of this study suggest that RB supplementation improved glucose tolerance and prevented the decline of innate immunity in high‐fat diet induced obese mice with OVX.

Support or Funding Information

This study was supported by research grant MOST 107‐2320‐B‐038‐014 from the Ministry of Science and Technology (Taipei, Taiwan).
Reduction of Liver Heparan Sulfate Proteoglycans Promotes Endocrine Benefits of Therapeutic FGF1
Authors: Vi Quach Anthony , Pessentheiner Ariane , Gordts Philip
Published: April 2020

Type‐2 diabetes, a is a global epidemic that currently drives an enormous health and economic burden. The disease is chronic condition in which blood glucose levels are deregulated due to insulin resistant in metabolic tissues, such as liver, muscle and adipose tissue. Recently, fibroblast growth factor 1 (FGF1) has been implicated in promoting insulin‐depended glucose uptake in adipose tissue. Therefore, FGF1 is being considered for therapeutic use in type‐2 diabetes patients.

Typically, both FGF and FGF Receptors (FGFR) bind heparan sulfate proteoglycans (HSPGs).This multimetric FGF‐FGFR interaction with HSPGs promotes stable complex formation allowing robust intracellular signaling. Our preliminary results show that the degree of HSPG sulfation in adipose tissue is critical for FGF1 mediated glucose lowering. The liver expresses significant amounts of FGFRs and highly sulfated cell surface HSPGs. Therefore, we hypothesize that the liver is a prime scavenger of therapeutic FGF1 reducing its partitioning to FGFR in adipose tissue and consequently its therapeutic window.

To address this hypothesis, we made use of wildtype and genetically modified mice with reduced liver HSPG sulfation introduced by liver specific inactivation of one of the biosynthetic enzymes N‐deacetylase and N‐sulfotransferase 1 (NDST1; Ndst1f/fAlbCre+). Both mice were fed a 60% kcal high fat diet (HFD) to initiate diet‐induced obesity and the associated peripheral insulin resistance. Throughout the HFD treatment, we monitored the mice’s glucose and body weight on a biweekly basis and found no phenotypic changes between the wildtype Ndst1f/fAlbCre+ mice. After 10 weeks on the HFD, we tested each mice group’s insulin and glucose tolerance via injections of the corresponding substrates and saw no significant differences in glucose homeostasis. By week 15 of HFD feeding, we injected a single dose of recombinant FGF1 protein (0.5 mg/kg) and recorded the FGF1‐induced glucose lowering in periodic intervals following the injection. Ndst1f/fAlbCre+ mice presented with a greater reduction in FGF1‐induced glucose lowering as well as longer lasting reduction effect.

Overall, the results from this research will provide an enhanced understanding of FGF1 and HSPGs that could contribute to future studies and treatments of type‐2 diabetes and overall help increase the therapeutic window of FGF1 treatment in type‐2 diabetes.

Support or Funding Information

Have been awarded Undergraduate Research Scholarship grant for working as a full‐time researcher during a 10‐week period.
Heat shock proteins in embryonic and adult submandibular salivary glands in healthy and tumorigenic tissues
Authors: Basset Charbel , Cappello Francesco , Rappa Francesca , Florena Ada Maria , Lentini Vincenzo Luca , de Macario Everly Conway , Macario Alberto J. L. , Jurjus Abdo , Jurjus Rosalyn , Leone Angelo
Published: April 2020

Introduction

Heat Shock Proteins (HSPs) role in development and function of human salivary glands have not yet been measured or mapped in detail, in particular, HSP60 role is unknown. They can be potential biomarkers for glandular diseases and malignancies. We present our initial findings on HSP10, HSP27, HSP60, HSP70 and HSP90 distribution in human submandibular salivary glands (SMG) at various stages of development and tumorigenesis.

Methods

Adult human SMG (normal and tumoral) and embryonic head tissue samples were processed by standard methods for routine histological analysis and immunohistochemical staining using antibodies against the five HSP’s. All procedures were in accordance with the Helsinki Declaration. In addition, proteins and RNAs were extracted from normal SMG of FVB‐NJ adult mice. Protein and gene expression of HSPs were assessed by western blot and RT‐PCR respectively.

Results

HSP27, HSP60 and HSP90 were present in the acini and ducts of embryonic salivary glands but had a different distribution pattern in adult glands: HSP27, HSP60 and HSP10 occurred only in the ducts whereas HSP90 and HSP70 were totally absent in the adult gland. The results were supported by our western blot and RT‐PCR as HSP10, HSP27 and HSP60 protein and mRNA were expressed in control adult mice SMG. HSP70 and HSP90 protein and mRNA were not expressed in adult mice SMG, thus matching our immunohistochemical results. In contrast, all 5 HSP’s levels were high in Pleomorphic Adenoma, Warthin’s tumor and Adenoid Cystic Adenoma.

Conclusions

The HSP27, HSP60 and HSP90 participate actively in the developmental process. HSP’s are also involved in neoplasm formation and progression. These results support the diagnostic and prognostic potentials of HSP’s.

Significance

Salivary glands tumors incidence is increasing, which makes it necessary to develop diagnostic tools and biometrics to understand the molecular mechanisms involved. Since molecular chaperones show quantitative variations and contribute to the carcinogenic mechanism in a variety of cancers, we aimed to investigate if a similar situation occurs in salivary glands and characterize the functions of the chaperoning system in physiology and pathology in these glands. Our findings exhibited quantitative patterns of the HSP’s, which can distinguish between normal and tumor tissues of the SMG and have potential in diagnosis and patient monitoring.
Effect of Strength Training and Green Tea on Memory Impairments Associated with &[Beta]‐Amyloid Peptide
Authors: Carpes Felipe P. , Schimidt Helen Lidiane , Carrazoni Guilherme , Garcia Alexandre , Izquierdo Iván , Mello-Carpes Pâmela Billig
Published: April 2020

Antioxidant supplementation and physical exercise have been discussed as strategies to minimize neurodegeneration in Alzheimer’s disease (AD). We investigated the neuroprotective effects of strength exercise (StrEx) and green tea (GT) supplementation either combined or isolated on memory impairments induced by a β‐amyloid characterizing an AD‐like condition. Wistar male rats were submitted to 8 weeks of StrEx or GT supplementation or its combination. AD was induced by injection of Aβ (25–35) in the hippocampus. We evaluate object recognition (OR) and social recognition (SR) memory, and then hippocampus was removed for biochemical analysis. StrEx improved OR and SR memory. StrEx combined with GT improved OR memory and did not improve SR memory. GT alone decreased antioxidant capacity and improved acetylcholinesterase activity. Both strength exercise or green tea are effective against cognitive impairments resultant of β‐amyloid but benefits do not add up when the two interventions are associated.

Support or Funding Information

National Council for Scientific and Technological Development of Brazil
Beyond academic learning: how can human cadaver dissection promote medical students’ professional identity development?
Authors: Eckert Tabitha , Topping Daniel , Abrams Matthew , Daly Katherine
Published: April 2020

INTRODUCTION

Growing evidence supports the potential activities such as memorial services and reflective writing centered around the human cadaver dissection (HCD) experience to support elements of medical student wellness. These activities promote psychological and professional development, increase resilience, protect against burnout and foster a sense of connection and community. This study is a qualitative analysis of a reflective writing exercise utilizing grounded theory to answer the question: “What is the impact of the first‐year HCD on medical students’ personal and professional development?”

METHODS

This qualitative cross‐sectional study was conducted at a medical school in the Southeastern United States. The study population consisted of 117 first year medical (M1) students, 70 males and 47 females, concluding their HCD module. A narrative reflection tool consisting of four reflection questions was designed to address the emotional elements of students’ HCD and promote introspection. The questionnaire was provided to twenty groups of 5–6 first‐year students. Grounded theory analysis was used to explore themes that arose in the students’ responses.

RESULTS

Participants exhibited several common initial reactions to HCD. Reflections commonly included themes of gratitude, awareness of one’s own mortality, appreciation for health and lifestyle, and recognition of the benefits of tangibly learning intricate anatomical details. After analysis of all team responses, a total of 266 unique open codes were identified for all four interview questions. These open codes were sorted into ten distinct broader axial codes, some of which had 2–3 sub‐themes.

CONCLUSION

Our study’s timely findings may inform anatomy program stakeholders as they face increasing pressure to decrease or eliminate HCD as part of curricular reform. HCD offers medical students holistic growth opportunities that are less impactful when encountered through alternative technologies. Institutions utilizing HCD should maximize this high‐yield, easy‐to‐implement opportunity to foster medical students’ professional identity development.

Primary Themes and Subthemes

Theme

Subtheme

1. Gratitude

No subthemes

2. Patient’s Story

No subthemes

3. Learning Anatomy

3.1 Lessons from my Donor
3.2 Tangible/Tactile Learning
3.3 Disease Process
3.4 Visual Learning

4. Diagnosis

4.1 Critical Thinking about Dx
4.2 Diligence
4.3 Creativity

5. Humanity’ and Compassion

No subthemes

6. Reflection

No subthemes

7. Career Development as a Physician

No subthemes

8. Lifestyle and Prevention

8.1 Body Appreciation
8.2 Preventing Disease

9. Empathy

No subthemes

10. Mortality

No subthemes
CaMKIIδ Is Necessary for Endothelial Cell Migration and Regulates Spouting Angiogenesis through Modification of MEF2‐dependent Gene Expression
Authors: O’Brien Brendan J. , Ginnan Roman G. , Singer Harold A.
Published: April 2020

Angiogenesis is a key homeostatic process of the endothelium that is essential for development and nourishment of hypoxic parenchyma. It is, however, also implicated as a contributor to a variety of pathologies such as rheumatoid arthritis, cancer metastasis, endometriosis, and multiple forms of retinopathy. Multifunctional Ca2+/Calmodulin–dependent protein kinase II (CaMKII) is a ubiquitous serine/threonine kinase with tissue‐specific expression and function. Data from our lab shows that CaMKII regulates VSM migration and proliferation in an isoform‐specific manner in the context of vascular remodeling following injury. We have also shown that CaMKII regulates gene expression downstream of MEF2, a key regulator of differentiation and angiogenesis, via phosphorylation of class II HDACs. Endothelial cell migration is a critical process that determines efficiency of vessel outgrowth and accurate directional commitment toward chemotactic and haptotactic cues. The goal of this study is the classify the CaMKII isoforms and their splice variants present in the endothelium and test their function in cell migration and angiogenesis. Knockdown of CaMKIIδ using RNAi attenuated both IL‐6 and VEGF‐dependent endothelial cell migration as measured by velocity and Euclidean distance in a single cell time lapse assay as well as a scratch wound. Additionally, in an electrode‐based quantitative wounding assay, cells treated with siCaMKIIδ healed standardized wounds significantly slower than non‐targeting siRNA controls. To investigate CaMKIIδ’s role in angiogenesis in vivo, we utilized the neonatal mouse retina as our model. Transgenic mice with an inducible VE‐Cadherin Cre were used to knock out CaMKIIδ and γ independently or simultaneously in an endothelial‐specific manner. In this model, we found that CaMKIIδ knockout animals have a 45% reduction of vascular outgrowth at postnatal day 6, fewer sprouts, and fewer anastomoses. These animals also display venous hypertrophy and delayed remodeling of the primary vascular plexus at postnatal day 10, consistent with a delayed angiogenic phenotype. Additionally, the knockout animals display hyperbranching, increased vessel coverage, and decreased lacunarity when allowed to grow to adulthood. The CaMKIIδ/γ double knockout phenocopies the δ knockout, suggesting that the observed phenotype is largely a δ effect. MEF2‐dependent gene expression in the endothelium was attenuated when CaMKIIδ was knocked down using siRNA. These data suggest that CaMKIIδ is necessary for endothelial cell migration and regulates sprouting angiogenesis, potentially through modification of MEF2‐dependent gene expression.
The Reflex Sympathetic Activation Attenuates Systemic Inflammation in Unanesthetized Rats
Authors: Brognara-Dias Fernanda , Castania Jaci Airton , Dias Daniel Penteado Martins , Kanashiro Alexandre , Salgado Helio Cesar
Published: April 2020

The literature reports that electrical activation or surgical denervation of the sympathetic nerves elicits anti‐inflammatory effects. Our previous study (American Heart Association’s Council on Hypertension 2016 Scientific Sessions: AP305) also indicated that sympathetic activation, by bilateral carotid occlusion (BCO) in unanesthetized rats, attenuates the release of pro‐inflammatory cytokines in an experimental model of sepsis. However, to expand the knowledge about the anti‐inflammatory potential of this arm of the autonomic nervous system, the objective of the present study was to investigate the role of global reflex sympathetic activation by BCO in a systemic inflammation induced by a low dose of lipopolysaccharide (LPS). For this, femoral artery and vein catheters were inserted into anesthetized Wistar Hannover male rats, for arterial pressure recording and LPS administration, respectively. Moreover, the subjects were implanted with pneumatic cuffs around the common carotid arteries for performing the BCO. On the next day, under unanesthetized rats, the arterial pressure was recorded continuously during 7 hours, including the baseline period. After baseline parameters recording, the BCO was performed for 20 s; and immediately after, LPS (60 μg/kg, i.v.) or saline was administrated. Blood samples were collected under different moments (90, 180, 270 and 360 min), after LPS or saline injection, for analyzing plasma cytokines (TNF, IL‐6, IL‐1β, and IL‐10) levels over time. As expected, BCO was effective to increase the arterial pressure, confirming the reflex sympathetic activation. Over time, no difference was observed among all groups concerning the arterial pressure; however, the groups which received LPS showed an increase in heart rate at 180, 270 and 360 min, compared with the saline group. However, the BCO did not change this response. Concerning the plasma cytokine levels, 90 min after LPS, BCO attenuated TNF and IL‐1β levels, as compared to LPS treated rats [TNF: 1831 ± 290 (n = 8) vs. 1326 ± 180 pg/mL (n = 8); IL‐1β: 212 ± 74 (n = 8) vs. 104 ± 39 pg/mL, (n = 8)]. BCO also increased the IL‐10 levels at 90 min [410 ± 55 (n = 8) vs. 730 ± 66 pg/mL (n = 8)]. Nevertheless, BCO did not change IL‐6 levels [IL‐6: 8389 ± 1480 (n = 8) vs. 8546 ± 1560 pg/mL (n = 8)]. No differences in plasma cytokine levels were observed at 180, 270 or 360 min after LPS, comparing the LPS group with the LPS+BCO group. In conclusion, these results suggest that the reflex sympathetic activation, in unanesthetized rats, attenuates plasma pro‐inflammatory cytokines and increase the anti‐inflammatory cytokine release in a model of systemic inflammation. These data provide support to previous evidence concerning the anti‐inflammatory effects of the sympathetic arm of the autonomic nervous system.

Support or Funding Information

FAPESP (2017/05163‐6; 2013/20549‐7) and CNPq (402076/2016‐8).
The Effect of Zinc on the Marker Expression in Cell Transitional Activities
Authors: Hu Yuli , Li Yang V.
Published: April 2020

Zinc is a trace element that participates in protein structure, enzyme activity, and gene regulation. Zinc homeostasis helps to maintain strong immunity, bone health, and the levels of various hormones, and it is also involved in the regulation of oxidative stress, cell death, and aging processes. We are interested in studying the effect of zinc homeostasis on the marker expression in cell transitional activities, including markers for epithelial‐to‐mesenchymal transition (EMT) and stem cells. In my preliminary experiments, the dissociation of zinc from its binding sites has been achieved by acidic treatment because the binding between zinc and protein is tight at basic pH and loose at acidic pH. We hypothesize that the acid‐induced dissociation of zinc from its binding sites contributes to the cell transitional activities, from one type of cell to a different kind of cells during EMT and stem cell development. In my preliminary results, acid treatment resulted in the accumulation of spindle‐shaped cells, similar to the morphology of EMT cells suggested by the literature. It increased the expression of vimentin (a marker labeling EMT), CDK2 (a marker labeling G1 and S phases), and SSEA‐4 (a marker labeling stem cells). This acidic treatment also decreased the doubling time of mouse gastric cells during long‐term cell culture, suggesting that the acid‐induced zinc dissociation promoted cellular growth perhaps through the activation of EMT, the facilitation of cell cycle, or the development of stem cells. Moreover, the chelation of zinc alleviated the spindle cell accumulation, vimentin expression, and SSEA‐4 expression induced by the acid exposure, confirming the importance of zinc dissociation from its binding sites in the cell transitional activities. In conclusion, zinc functions in cell transitional activities through regulating the expression levels of marker proteins involved in EMT, cell cycle, and stem cell development.

Support or Funding Information

Funding is provided partially by NIH (# NS081629) to YVL. Thanks to the Osteopathic Heritage Foundations, Graduate Assistantship Program at Ohio University Heritage College of Osteopathic Medicine for supporting graduate student YH.
Docosahexaenoic Acid (DHA) but not Eicosapentaenoic Acid (EPA) induces the trans‐differentiation of C2C12 into white‐like adipocytes phenotype
Authors: Ghnaimawi Saeed , Huang Yan , Baum Jamie
Published: April 2020

Muscle derived stem cells (MDSCs) and myoblasts play an important role in the regeneration of myotubes when muscle tissue is injured. However, these cells can be induced to differentiate to other lineages during embryonic development such as adipocytes, osteocytes, and chondrocytes in response to external stimuli like maternal diet enriched with EPA and DHA that are highly suggested during pregnancy because of their neuronal development benefits and positive role in improving retinal and immune functions. The objective of this study is to explore the effect of isolated EPA and DHA on myoblasts undergoing differentiation into brown and white adipocytes. C2C12 myoblasts were cultured to confluency and then treated with white and brown adipogenic differentiation medium with and without 50μM EPA and 50μM DHA separately. DHA treated group but not EPA reprogrammed C2C12 into white like adipocyte phenotype by down‐regulation the expression (P<0.05) of genes involved in regulation the terminal differentiation of myoblasts into myotubes such as MyoD, MyoG, and Mrf4 and up‐regulation the expression of white adipogenesis specific markers including PPARγ, C/EBPα, C/EBPβ, aP2, BMP4, FAT, ZFP423, and Il‐6 (P<0.05). Furthermore, DHA reduced the thermogenic capacity of brown adipocytes derived from C2C12 independent of UCP1 by significantly suppressing (P<0.05) the expression of PGC1α and DIO2 genes and mitochondrial biogenesis by down‐regulation the expression of PGC1α and TFAM followed by decreasing mDNA copy number (P<0.05). Accordingly, DHA treated group showed an increased accumulation of large monocular lipid droplets, suppressed respiration and spare capacity. EPA, on the other hand, reduced the myogenesis process at the terminal stage by inhibiting the expression of MyoD, MyoG, and Mrf4 (P<0.05), but no significant differences were observed in adipogenesis regulating genes such as PPARγ, C/EBPα, C/EBPβ, and aP2 (P>0.05). Likewise, EPA treatment suppressed the expression of WAT signature genes BMP4, FAT, and ZFP423 (P<0.05) indicating its antagonist effect to DHA. Although PGC1α and TFAM expression were reduced in the EPA treated group, no significant changes in mDNA copy number and mitochondrial function were observed. Unlike DHA treatment, EPA supplementation has no effect on the differential route of C2C12 into brown adipocytes. It is to be included that EPA and DHA may similarly affect the integrity of muscle tissue, but DHA is a potent adipogenic and lipogenic factor that can change the metabolic profile of the body by increasing intramuscular fat.
A novel series of nitrofuran derivatives produces an anti‐tumor effect via a p53‐dependent mechanism
Authors: Al Koussa Houssam Khaled , Abrahamian Carla F. , Elzahhar Perihan M. , Serie Marwa Abu , Belal Ahmed , El-Yazbi Ahmed F.
Published: April 2020

Nitrofurantoin (NF) is an antibacterial that acts by inducing DNA damage. Increased selectivity towards bacterial DNA is conferred through the rapid activation by prokaryotic reductases. In cancer therapy, radiation and anti‐neoplastic drugs induce DNA damage to activate the p53‐dependent pathway, provoking apoptosis in cancer cells. In this study, a novel series of nitrofuran derivatives was designed with a presumed enhanced eukaryotic activity. The safety and the anti‐tumor efficacy of this series was assessed. Anticancer effects were assessed on colorectal cancer (Caco‐2), breast cancer (MCF‐7), cervical cancer (Hela), and liver cancer cells (HepG‐2). Cytotoxicity was estimated by MTT assays. Three compounds, NF‐5, NPO, and NS‐1 were shown to have the highest cytotoxic activity against cancer cells at IC50 values that are several‐fold lower that those observed for non‐malignant lung fibroblasts. The abilities to induce oxidative stress and apoptosis were tested by flow cytometry to measure dihydrodichlorofluorescein fluorescence and nuclear annexin‐V staining, and compared to that of NF and the reference chemotherapeutic drug (5‐FU). Other apoptosis markers measured include caspase3/7 activity and Bax/Bcl2 ratio. NF‐5, NPO, and NS‐1 induced oxidative DNA damage in cancer cells in vitro with subsequent cellular apoptosis. Furthermore, the three derivatives induced more than 50% increase in reactive oxygen species when compared to 5‐FU. Cells treated with these drugs showed an increased Bax/Bcl2 ratio and caspase 3/7 activity compared to the traditional chemotherapeutic drug. A positive linear correlation was shown between the oxidative load produced in the cells and the increase in apoptotic markers. Interestingly, despite equivalent induction of DNA damage demonstrated by similar pATM nuclear shuttling, NF‐5, NPO, and NS‐1 produced more cytotoxicity in MCF‐7 cells compared to the p53 mutant breast cancer cell line MDA‐MB‐231. Moreover, the effect of these derivatives on the p53 interaction with its negative modulator Murine Double Minute 2 (MDM2) was assessed using a multi‐spot sandwich ELISA kit. NF‐5, NPO, and NS‐1 promoted p53 release from MDM2 at concentrations that are 2–3 orders of magnitude lower than their cytotoxic IC50 values. These findings were further confirmed by in silico docking studies demonstrating possible binding sites on the protein. Further experiments are underway to assess the ability of lower, non‐cytotoxic concentrations of these drugs to improve the radio‐sensitivity of cancer cells via activation of p53‐dependnet pathways.

Support or Funding Information

Funded by: AUB‐FM MPP grant #320148
Defining the role of microglia in breast‐to‐brain metastasis
Authors: Thompson Kaitlyn Koenig , Tsirka Stella E.
Published: April 2020

A diagnosis of brain metastasis is devastating to cancer patients, with survival measured in only months. Due to prolonged life expectancy of cancer patients, the frequency of brain metastasis is increasing, with the highest incidences occurring when the primary site is the lung, breast, and melanoma. Treatment of brain metastatic lesions is challenging; differences in the brain microenvironment and the encompassing blood‐brain barrier are two major obstacles to successful systemic therapy. A unique player in the brain tumor microenvironment (TME) are microglia, the resident innate immune cells of the central nervous system. How microglia influence metastatic tumor progression in the brain, and their therapeutic potential in this setting remain to be elucidated. To first characterize the role of microglia in a model of breast‐to‐brain metastasis, we deplete them using the orally available colony stimulating factor 1 receptor (CSF1R) inhibitor, PLX5622, which readily crosses the blood‐brain barrier. Brain metastatic development of a C57BL/6 syngeneic breast cancer line is monitored, and the resulting TME characterized histologically. Further, we utilize in vitro assays to investigate direct communication between microglia and neoplastic cells, measuring tumor cell migration and invasion, and microglial polarization. We also employ PLX5622 therapeutically, after the development of brain metastasis to understand whether dynamic elimination of microglia may be a viable therapy. Understanding microglia’s role in the development of metastatic disease and how these cells may be targeted therapeutically is critical to improving the treatment of brain metastasis.

Support or Funding Information

This work is supported by NIH IRACDA K12GM102779.