bluuebunny/crossref_metadata_2025 huggingface dataset:
Study on changes of cardiac function and peripheral vascular resistance in elderly patients with hypertension
Authors: Hao Chunyan , Yang Shuang , Li Mingli , Duan Hubin
Published: May 2022

Objective

To investigate the changes of cardiac function and peripheral vascular resistance in elderly patiants with hypertention and their correlation with homocysteine (Hcy) and lipid levels.

Methods

126 hypertensive patients aged ≥60 years in the department of geriatrics, the first hospital of shanxi medical university from October 2017 to March 2019 were selected as the hypertension group, and 107 healthy patients aged ≥60 years in the same period were selected as the control group(The clinical study has passed ethical review). Serum Hcy and lipid levels in the two groups were measured, and parameters of cardiac function and peripheral vascular resistance were recorded by non‐invasive hemodynamic detector. The changes of Hcy, lipid profile, cardiac function and peripheral vascular resistance parameters in the two groups were compared, and the correlation between cardiac function and peripheral vascular resistance parameters and Hcy and lipid profile was analyzed.

Results

Hcy, total cholesterol (TC), triglyceride (TG) and low‐density lipoprotein cholesterol (LDL‐C) in the hypertension group were higher than those in the control group (P<0.05), while high‐density lipoprotein cholesterol (HDL‐C) was lower than those in the control group (P<0.05). Peripheral vascular resistance (SSVR) and systemic resistance (SVR) in the hypertensive group were higher than those in the control group (P<0.05), and cardiac function parameters (CO), SV, LSW and LVET were lower than those in the control group (P<0.05). Pearson correlation analysis showed that hypertension group CO and Hcy in cardiac output, and negatively correlated with TC, LDL‐C, stroke volume SV and negatively correlated with Hcy, left ventricular stroke work LSW and negatively correlated with Hcy, LDL‐C, left ventricular ejection time (LVET)and negatively correlated with Hcy, peripheral vascular resistance index SSVR, SVR and Hcy, TC, TG, LDL‐C were positively correlated, and negatively correlated with the SVR and HDL‐C.

Conclusion

In elderly patients with hypertension, cardiac function, including cardiac displacement and myocardial contractility, was significantly reduced, and peripheral vascular resistance was significantly increased. Moreover, such hemodynamic disorder was associated with Hcy and lipid levels.
The Effect of Curricular Reform on Gross Anatomy Laboratory Examination Performance: A Retrospective Analysis
Authors: Taylor Melissa , Nichols Richard A. , Herr Michael , Loder Danielle
Published: May 2022

Introduction

Medical education in the United States has changed dramatically in the last few decades. Medical schools have moved away from a traditional, didactic, and discipline‐based curriculum to a more integrative, active, and student‐centered curriculum. Another common change has been the shift from a graded curriculum to a pass‐fail curriculum. While much research has documented overall performance, especially on high stakes examinations, little research has been conducted on gross anatomy laboratory performance.

Aim

This study examines the effects of curricular change, including decreased contact hours and shift to a pass‐fail curriculum, on gross anatomy laboratory performance.

Methodology

Gross anatomy laboratory examinations were reviewed retrospectively in two cohorts: one cohort that was in the previous, nonintegrated curriculum with a graded (i.e., A, B, F) curriculum; and a second cohort that was part of the first integrated class with a pass‐fail curriculum. For the pass‐fail curriculum in particular, the standard for passing was notably lower than that for the graded curriculum. Each of the students’ examinations were de‐identified. Only identification questions (i.e., ones that asked “Identify the tagged structure”) were reviewed. Questions that were common between the two cohorts were used for analysis. An independent samples t‐test was performed on the data, and Cohen’s d effect size was calculated. Significance was ascribed for p<0.05.

Results

There were 69 questions that were similar between all the laboratory examinations from the pre‐curricular reform and post‐curricular reform cohorts. There was an approximately even number of questions from muscles, nerves, vessels, and additional structures on the cadaveric specimens and plastic models used on the examinations. It was found that the post‐curricular reform cohort scored significantly lower (mean = 67.05 ± 22.35) than the pre‐curricular reform cohort (mean = 79.89 ± 19.56) p<0.001, d = 0.61.

Discussion and Conclusion

The results of the study demonstrate that students within the new curriculum, with reduced contact hours and a pass‐fail curriculum, performed significantly poorer than students in the non‐integrated curriculum. Students may have felt that the gross anatomy laboratory experience was lower stakes compared to their other coursework, especially since the passing standard for the post‐curricular reform cohort was markedly lower than the that of the pre‐curricular reform cohort.
Development of highly potent and selective GRK5 inhibitors
Authors: Chen Yueyi
Published: May 2022

G protein‐coupled receptor kinases (GRKs) regulate cell signaling by triggering receptor desensitization via phosphorylation on G protein‐coupled receptors (GPCRs). (1) The seven human GRKs (GRK1−GRK7) are classified into three subfamilies: GRK1 (GRK1 and GRK7), GRK2 (GRK2 and GRK3), and GRK4 (GRK4, GRK5, and GRK6). GRK2 and GRK5 are the most abundant in cardiovascular tissue, where they are potential targets for treatment of cardiovascular disease. GRK5 also undergoes Ca2+/calmodulin‐dependent nuclear localization, where it phosphorylates histone deacetylase 5 (HDAC5), inducing an increase in transcription of cardiac hypertrophy‐related genes. In GRK5‐knockdown mice, cardiomyocytes are protected from hypertrophy; however, the specific roles of GRK5 in heart failure and hypertrophic cardiomyopathy are still unclear. Furthermore, GRK5 is required for cancer progression in various cancer types. Depletion of GRK5 has been shown to suppress prostate cancer, breast cancer, and non‐small‐cell lung cancer. (2) Therefore, targeting GRK5 can also be a chemotherapeutic strategy. Here we are testing a series of inhibitors for GRK5/6 derived from the indolinone scaffold, utilizing Cys474 residue unique in GRK5/6 to enhance selectivity by covalent capture. An immediate goal is to facilitate the development of more selective inhibitors via rational design by determining x‐ray crystal structures of inhibitor complexes with GRK5. Recently, we have crystallized a hyper‐phosphorylated GRK5, and the active site structure suggested the possibility for compound soaking. Overall, discovery of the inhibitors can significantly facilitate understanding and treatment of cardiovascular diseases and cancer.

1. Z. Yang et al., Phosphorylation of G Protein‐Coupled Receptors: From the Barcode Hypothesis to the Flute Model. Molecular Pharmacology 92, 201‐210 (2017).

2. L.‐P. Jiang et al., GRK5 functions as an oncogenic factor in non‐small‐cell lung cancer. Cell Death Dis 9, 295 (2018).
Enhanced confidence and ultrasound‐performance skills of medical students: A comparative study with and without prior videographic ultrasound exposure
Authors: Li Jeffrey J. , Kim Jane J. , Tan Shirley , Young Corey , Park Claire , Haney Megan , Nausheen Fauzia
Published: May 2022

INTRODUCTION

Point‐of‐care ultrasonography (POCUS) is commonly used in many modalities of medicine. However, there is a lack of standardized protocol for integrating POCUS into preclinical medical education. Previous literature has detailed the benefits of POCUS in clinical diagnoses and reasoning, but many medical students still lack exposure to POCUS. At California University of Science and Medicine (CUSM), medical students will be exposed to POCUS in their preclinical years. We aim to compare the confidence levels and ultrasound‐performance skills of medical students with and without prior videographic ultrasound exposure.

Hypothesis

Prior videographic ultrasound exposure will improve the confidence and ultrasound‐performance skills of medical students during a live ultrasound workshop compared to those without videographic exposure.

Methods

Participants will be divided into two groups. One group will watch a 5–10‐minute procedural video about ultrasound‐guided peripheral intravenous (IV) access prior to ultrasound training, while the other group will be ultrasound‐naive prior to ultrasound training. Ultrasound training will be led by three other medical students who are trained and approved by ultrasound physicians. After demonstrations, participants will practice ultrasound‐guided IV access on a practice pad. Instructors will grade students on the spot on their ability to perform ultrasound‐guided IV access correctly. The parameters measured will be proper positioning of the ultrasound probe, insertion of the needle, ultrasound probe movement, puncturing of the vein, and overall technique. The two study groups will then take surveys on their perceived performance and overall confidence levels.

Results

Results from participants will be scored using two measures. Firstly, their self‐reflection surveys will measure their level of confidence and perceived ultrasound‐performance skills. Secondly, on‐the‐spot grading by instructors will provide information on participants’ actual ultrasound‐performance skills. A standardized grading scheme will ensure fairness among different instructors and participants.

Conclusion

Comparison of the effectiveness of an additional videographic ultrasound component prior to live demonstration will be elucidated. This study will help create elements within existing ultrasound curriculae that will enhance the confidence and ultrasound‐performance skills of preclinical medical students.
Marsupsin Isolated from Polyherbal Product ABPA Rejuvenates Insulin Resistance in L6 Skeletal Muscle Cell Line via IRS1‐PI3K‐AKT‐GLUT4 Signaling Pathway
Authors: Balachandran Abbirami , Okechukwu Patrick N. , Gunasekaran Baskaran , Nalweyiso Jacinta , Sapra Sameer , Stanslas Johnson , Froemming Gabriele R. A. , Rajendran Salini D.
Published: May 2022

Background

Insulin resistance (IR) is one of the key characteristics of type 2 diabetes mellitus (T2DM) which impairs the insulin signal transduction pathway. It is typically caused by many factors including hyperglycemia, accumulation of advanced glycation end products (AGEs), chronic oxidative stress and hyperlipidemia. Hence, complementary medicine such as herbal medications have been widely used as a complement to allopathic medicine to facilitate its efficacy while minimizing its potential adverse effects. ABPA is a polyherbal formulation that consists several powerful antidiabetic agents such as Pterocarpus marsupium. Our previous study showed that ABPA exhibited strong antidiabetic effect and detected high expression of marsupsin through HPLC studies.

Objective

The current study aims to elucidate the effect of marsupsin in rejuvenating insulin resistance through the IRS1‐PI3K‐AKT‐GLUT4 insulin signaling pathway.

Method

L6 skeletal muscle cells were cultured and differentiated into L6 myotubes under standard conditions. The differentiated L6 myotubes were induced with high insulin to mimic insulin resistance and treated. Glucose uptake assay was also conducted to evaluate the effect of marsupsin on the rate of glucose uptake in the L6 cells. The cells were then lysed to extract the protein and RNA where the protein and differential gene expression of IRS1, PI3K, AKT and GLUT4 were analyzed using ELISA, qPCR, and Western Blot respectively.

Results

The rate of glucose uptake was higher in MAR‐treated cells as compared to the negative control. This was further confirmed in the ELISA, qPCR and Western Blot analysis where MAR‐treated cells showed an upregulation of IRS1, PI3K, AKT and GLUT4 proteins as compared to the vehicle control.

Conclusion

Marsupsin rejuvenates insulin resistance in skeletal muscle cells by increasing the rate of glucose uptake through the activation of the IRS1‐PI3K‐AKT‐GLUT4 pathway.
Aminoisoquinolines Penetrate the Blood‐Tumor Barrier and May Inhibit Metastasis of Lung Cancer to the Brain
Authors: Bakhle Kimaya , Nimishetti Naganna , Dayal Neetu , van Veldhuisen Aletea , Bai Yunfeng , Crain Christa , Sintim Herman , Goergen Craig , Lyle L. T.
Published: May 2022

Two out of five patients with non‐small cell lung cancer (NSCLC) develop brain metastases with 3‐8 month overall survival time. One of the greatest challenges for developing chemotherapeutics for brain metastases of NSCLC is the blood‐brain barrier (BBB). The BBB is composed of endothelial cells with distinct tight junctions, pericytes, a unique basement membrane and astrocyte endfeet. This unique vasculature structure opposes the movement of deleterious molecules, toxins, and drugs from the bloodstream into the central nervous system. We have previously characterized a shift from the impermeable BBB to a heterogeneously permeable blood‐tumor barrier (BTB) during the formation of brain metastases of non‐small cell lung cancer. These changes in permeability and composition may make brain tumors more accessible to therapeutics delivered through the bloodstream. One potential therapeutic is aminoisoquinolines. In vitrodata has demonstrated the efficacy of aminoisoquinolines in growth inhibition of lung cancers. They do so through inhibition of AuroraB and p70S6K kinases, two kinases critical to proliferation of lung cancer cells. Herein, we investigate the ability of aminoisoquinolines to inhibit metastasis of NSCLC to the brain through the BTB. We hypothesize that treatment of mice harboring brain‐seeking NSCLC cells with aminoisoquinolines will prevent formation of brain metastases. Nineteen athymic nude mice were injected with brain‐seeking NSCLC tumor cells, adenocarcinoma subtype (A549), via ultrasound‐guided intracardiac injection. The mice were then treated with one of two aminoisoquinolines or a vehicle control. After 10 days of treatment, brains were procured from the animals and evaluated microscopically for presence or absence of brain metastases. We identified an average of 4.25 (n=4) metastases in animals treated with the aminoisoquinoline HSN 789, with an average diameter of 15.66 micrometers. Animals treated with the aminoisoquinoline HSN 804 had an average of 18.5 metastases (n=2), with an average diameter of 23.00 micrometers. Animals treated with the vehicle control failed to form brain metastases; we are investigating cell viability in this experimental group. A complete evaluation of cell viability in all treatment groups will reveal the in vivo therapeutic efficacy of aminoisoquinolines for treatment of brain metastases of NSCLC. Preliminary data suggest that treatment with aminoisoquinolines may be sufficient to inhibit growth of brain metastases from NSCLC.
Machinery in motion: New insights into mitochondrial proteostasis
Authors: Lander Gabriel , Mindrebo Jeff
Published: May 2022

ATP‐dependent AAA+ proteases are critical regulators of mitochondrial functions, playing crucial roles in the mitochondrial quality control response system. The past years have provided much structural insight into the molecular mechanisms associated with degradation of substrates by these proteolytic machines. Recent cryo‐electron microscopy (cryo‐EM) studies have provided critical insights into a conserved, AAA+‐mediated hand‐over‐hand substrate translocation mechanism required to processively engage, unfold, and degrade proteolytic substrates. However, the underlying mechanisms regulating their various activities are not well understood. Numerous prior studies suggest that AAA+ protease have evolved numerous layers of regulation to control or tune proteolytic activity to meet cellular needs. Herein, we present compelling biochemical and structural data that support a long‐range allosteric model linking substrate binding to proteolytic activity.
Creative approaches to perform an inclusive faculty search
Authors: Gilbert Wendy
Published: May 2022

Names are powerful cues that can trigger unintended bias including ingroup favoritism, a cognitive bias that leads people to overvalue the past accomplishments and future potential of individuals who seem similar to themselves. Research shows that anonymous job searches can significantly increase the chance for applicants from underrepresented (UR) backgrounds to be invited for an interview. This result strongly suggests that in traditional (non‐anonymous) job searches, well‐qualified applicants are overlooked due to bias. I will present the design and results from an unusual tenure‐track Assistant Professor search in the department of Molecular Biophysics and Biochemistry at Yale University. Applicants submitted anonymized research statements—no names of people, places, funding agencies or journals—and a short list of diverse candidates was created based solely on de‐identified documents. Our experiment suggests that anonymous application procedures are one potentially useful tool to combat persistent racial hiring discrimination which could explain the very slow increase of UR scientists among research faculty compared to the growth of UR PhDs.
Identifying toxic metabolites and their roles in disease
Authors: Kim Dohoon
Published: May 2022

In a metabolic pathway, a series of metabolites are formed then chemically converted to the next one along the pathway. When this chain of events is broken, such as due to mutation of a pathway enzyme, a massive accumulation of a metabolite can occur and can have deleterious consequences, especially if the metabolite has toxic properties. Here, we describe our recent efforts to identify such toxic metabolites in a systemic manner, and examine their relevance in diseases. When a metabolic pathway is overactive in a cancer cell, blocking enzymes which process a toxic intermediate within that pathway can result in cancer cell selective toxic metabolite accumulation and poisoning, a highly attractive therapeutic strategy. We demonstrate as proofs of principles of this approach, manipulation of the selenocysteine biosynthesis pathway and the de novo sphingolipid biosynthesis pathway, for cancer therapy. We will also discuss how toxic metabolite accumulation can be contributing factors in other such as neurodegenerative disorders, and how metabolic pathway manipulation may be considered as a therapeutic strategy.
Transcriptional regulation of primary and specialized metabolism
Authors: Brady Siobhan
Published: May 2022

Michelle Tang1, Allison Gaudinier1, Baohua Li1, Lifang Zhang2, Christophe Liseron‐Monfils2, Joel Rodriguez‐Medina1, Anne‐Maarit Bagman1, Andrew Olsen2, Doreen Ware2,3, Mary Frank4, Bo Shen4, Daniel J. Kliebenstein1, Siobhan M. Brady1

1UC Davis, Davis, CA

2Cold Spring Harbor Laboratory, Cold Spring Harbor, NY

3USDA‐ARS, Albany, NY

4DuPont Pioneer, Johnston, IA

Regulation of plant development requires intricate communication with both primary and specialized metabolism in order to fuel growth. While transcriptional regulation of metabolism is evident from myriad whole genome‐expression analyses, our understanding of which transcriptional regulators are responsible for these changes as well as their underlying mode of action is unclear. I will highlight our efforts on systematic mapping of transcriptional regulators of primary metabolism (nitrogen and central carbon) as well as specialized metabolism (glucosinolates). Network analyses incorporating protein‐DNA interaction data, gene expression and connectivity were used to identify critical regulators, most of which were shown to regulate metabolism via developmental‐ and stress‐conditional processes that coordinate across primary and specialized metabolism. These analyses demonstrate distinct design principles responsible for metabolism between microbes and this multicellular eukaryote.
Regulation of PIP2 homeostasis at ER‐plasma membrane contacts by Nir proteins
Authors: Liou Jen
Published: May 2022

Phosphatidylinositol 4,5‐bisphosphate (PIP2) in the plasma membrane is a critical lipid molecule that regulates nearly all functional processes occurring at the cell surface. Additionally, PIP2 is hydrolyzed following receptor‐induced activation of phospholipase C to generate second messengers for signal transduction. It is crucial to quickly replenish PIP2 in the plasma membrane following receptor‐induced hydrolysis to sustain signaling outputs and to maintain PIP2‐dependent cellular functions. Replenishment of plasma membrane PIP2 in receptor‐stimulated cells involves transport of phosphatidylinositol (PI) from the endoplasmic reticulum (ER) to the plasma membrane for PIP2 re‐synthesis. ER‐plasma membrane contacts are subcellular loci characterized by the close apposition of the ER to the plasma membrane. Recent findings revealed that the PI transfer proteins, Nir2 and Nir3, dynamically localize at ER‐plasma membrane contacts to mediate replenishment of PIP2 hydrolyzed following receptor stimulation. Deficiency in Nir2 and Nir3 suppressed replenishment of PIP2 in the plasma membrane and Ca2+ signaling in receptor‐stimulated cells. A third mammalian Nir protein, Nir1, has been linked to several human diseases including retinal dystrophy. Unlike Nir2 and Nir3, Nir1 lacks a PI transfer protein domain and readily localizes at ER‐plasma membrane contacts in resting cells. The presence of Nir1 promotes Nir2 localization at ER‐plasma membrane contacts to mediate PIP2 replenishment following receptor stimulation. Together, Nir proteins maintain plasma membrane PIP2 homeostasis at ER‐plasma membrane contacts to support cell signaling and plasma membrane functions in receptor‐stimulated cells.
Roles for inter‐organelle contacts in organizing metabolism
Authors: Henne W. M.
Published: May 2022

When faced with nutrient shortage, cells adapt by remodeling their metabolic pathways and organelles. I will discuss our work using budding yeast to dissect how nutrient shortage impacts the spatial organization of metabolism and lipid droplets. We find that in response to glucose restriction, yeast remodel their mevalonate pathway by spatially compartmentalizing its rate‐limiting enzyme, HMG‐CoA Reductase (HMGCR). HMGCR spatial compartmentalization occurs at a unique inter‐organelle contact site called the nucleus‐vacuole junction (NVJ). This spatial partitioning enhances HMGCR activity, driving mevalonate synthesis to enable cellular adaptation. Our work suggests a new use for an inter‐organelle contact site in the fine‐tuning of mevalonate metabolism during nutrient stress.

Remarkably, we also find that glucose restriction drives the phase transition of lipids within lipid droplets (LDs), causing them to convert from a disordered to liquid‐crystalline phase. Mechanistically, we find that these liquid crystalline lattices (LCLs) within LDs require triglyceride lipolysis. We also find that LCL‐LDs exhibit changes to the LD surface proteome. Several known LD proteins redistribute from LDs to the ER network, whereas others remain on LCL‐LDs, suggesting phase transitions of LD lipids influences LD protein targeting. Global proteomics also reveals that triglycerides harvested from LDs fuel cellular energetics at peroxisomes and mitochondria. This indicates that glucose starvation induces inter‐organelle lipid flux while promoting lipid phase transitions within LDs.
Correlated motions in enzymes
Authors: Ando Nozomi , Meisburger Steve P. , Case David A.
Published: May 2022

Correlated motions in proteins arising from the collective movements of residues have long been proposed to be fundamentally important to key properties of proteins, from catalysis and allostery to evolvability. Recent breakthroughs in structural biology have made it possible to capture proteins undergoing complex conformational changes, yet the subtle and intrinsic fluctuations within a single conformation remain one of the least understood facets of protein structure. For many decades, the analysis of diffuse X‐ray scattering held the promise of animating crystal structures with such correlated motions. With recent advances in both X‐ray detectors, this long‐held promise can now be met. In this talk, I’ll describe our recent work in interpreting this signal and discuss how it can be used to the understand the role of correlated motions in enzymes.
Building the microtubule cytoskeleton via phase transitions
Authors: Petry Sabine
Published: May 2022

How does a cell construct its microtubule cytoskeleton? According to Feynman’s principle “what I cannot create, I do not understand”, my lab pursues this question by building the chromosome segregation machinery from scratch. I will first tell you how the microtubule framework is generated in a cell. Upon deciphering the function of the most important microtubule accessory proteins, I will present how we use those building blocks to reconstitute a spindle substructure in vitro and determine its building plan. Surprisingly, the spindle substructure relies on a protein condensate, which forms on the surface of a microtubule and arranges into a droplet pattern like a fluid.

By studying how the MT cytoskeleton is built, I hope to help explain how hundreds of proteins can self‐assemble on the nm scale into a complex molecular machine 1000‐fold larger than its constituents, a challenge for the biochemistry of the 21st century.
The role of phase separation in oncogenesis by fusion oncoproteins
Authors: Kriwacki Richard
Published: May 2022

NUP98 fusion oncoproteins (FOs) are drivers in pediatric leukemias and many transform hematopoietic cells. Most NUP98 FOs harbor an intrinsically disordered region from NUP98 that is prone to liquid‐liquid phase separation (LLPS) in vitro. A predominant class of NUP98 FOs, including NUP98‐HOXA9 (NHA9), retains a DNA‐binding homeodomain, whereas others harbor other types of DNA‐ or chromatin‐binding domains. NUP98 FOs have long been known to form puncta, but long‐standing questions are how nuclear puncta form, and how they drive leukemogenesis. We will discuss our studies of NHA9 condensates, showing that homotypic interactions and different types of heterotypic interactions are required to form nuclear puncta, which are associated with aberrant transcriptional activity and transformation of hematopoietic stem and progenitor cells. We also show that three other leukemia‐associated NUP98 FOs form nuclear puncta and transform hematopoietic cells. To extend our findings, we tested ~150 additional fusion oncoproteins associate with a wide range of human cancers for puncta formation in cells. We will discuss our observation that >50% of these formed cellular condensates, with this behavior for some linked with aberrant gene expression and cell transformation. Our findings indicate that LLPS is critical for leukemogenesis by NUP98 FOs and likely contributes to oncogenesis driven by many other fusion oncoproteins.
RNA methylation in gene expression regulation
Authors: He Chuan
Published: May 2022

Over 150 types of post‐transcriptional RNA modifications have been identified in all kingdoms of life. We have discovered two RNA demethylases, FTO and ALKBH5, which catalyze oxidative demethylation of the most prevalent modifications of mammalian messenger RNA (mRNA) and other nuclear RNA, N6‐methyladenosine (m6A). These findings indicate that reversible RNA modification could impact biological regulation analogous to the well‐known reversible DNA and histone chemical modifications. I will present recent discoveries on m6A‐mediated regulation of both transcriptional and post‐transcriptional events. We found m6A methylation of chromatin‐associated regulatory RNAs (promoter‐associated RNAs, enhancer RNAs, and repeat RNAs), or carRNAs. carRNA methylation regulates local chromatin state and downstream transcription. Regulatory pathways and underlying mechanisms will be presented.
Epigenetic Mediators of Risk for Metabolic Disease Across Generations
Authors: Patti Mary E.
Published: May 2022

Obesity and type 2 diabetes (T2D) are leading global health threats and innovative multi‐pronged approaches are needed to combat both epidemics. Many risk factors for obesity and T2D – beyond genetics – are present before birth, reflecting the influence of environmental factors on germ cells and on the developing infant. For example, parental obesity or diabetes, prenatal undernutrition, and both low and high birth weight, are linked to offspring’s obesity and T2D risk later in life. Such prenatal exposures increase obesity risk not only in the individual exposed to the stressor, but also in their offspring. The result is a vicious cycle of risk to subsequent generations.

The concept that the maternal perinatal environment shapes childhood chronic diseases is well‐recognized. However, mounting evidence demonstrates that fathers can also contribute to obesity and diabetes risk to their offspring. While genetic factors are important, additional non‐genetic impacts of paternal metabolism have been demonstrated both in rodent models and in humans. Paternal metabolism can influence epigenetic signals in sperm, including DNA methylation, histone modification, and/or noncoding RNA (ncRNA), which can impact gene expression and development in the offspring. We and others have shown that epigenetic marks are altered in sperm from rodents exposed to prenatal undernutrition, postnatal overnutrition, and other stimuli. While human data are more limited, paternal BMI at the time of conception is associated with infant birth weight and DNA methylation of infant cord blood; differential DNA methylation at some loci persisted to age 7. Our new preliminary data demonstrate that T2D also modifies the sperm methylome.

Given that sperm epigenetic marks may be modifiable, this suggests a potential novel approach and untapped opportunity for multigenerational prevention of metabolic disease. We demonstrate that interventions in male mice, including exercise and pharmacological lowering of glucose levels, result in amelioration of obesity, glucose tolerance and metabolic syndrome in offspring.
Volume electron microscopy analysis reveals a new type of membrane junction required for mixing of parental genomes after fertilization
Authors: Cohen‐Fix Orna , Rahman Mohammad , Maheshwari Richa , Onyundo Megan , Drey Seth , Narayan Kedar
Published: May 2022

Imaging cells by light or fluorescence microscopy often provides great insights into mechanisms and dynamics of cellular processes. However, to gain information regarding the cellular structures that underlie these processes, nm‐scale resolution is often required. This type of resolution can be obtain using electron microscopy (EM). Traditionally, investigators imaged slices of chemically fixed cells or tissue in order to glimpse at relevant structures in 2‐dimensions. While powerful, this approach can potentially miss important information due to the somewhat random location of the planes analyzed, especially if a small number of images is a acquired. More recently, methodologies of volume EM have been developed that allow for sampling of a continuous volume at high resolution. Here we will describe how one such methodology, Focused Ion Beam‐ Scanning Electron Microscopy (FIB‐SEM), has been used to understand the process by which parental genomes mix after fertilization in C. elegans embryos. One‐cell embryos were collected during different stages of interphase and mitosis, fixed using high‐pressure freezing followed by freeze substitution, and stained to maximize membrane contrast. Images were acquired at 9 nm lateral (x,y) and axial (z) resolution, for volumes of approximately 10 X 10 X 25 µm3. These analyses revealed that for parental chromosomes to mix, the membranes of the two pronuclei must fused via a membrane structure that we termed “3‐way sheet junction”. The steps leading to the formation of 3‐way sheet junctions, and the more recent discovery of a novel membrane structure around centrosomes using these datasets, will be discussed.
How do aerobic organisms solve the oxygen sensitivity problem of [4Fe‐4S] in radical SAM enzymes?
Authors: Lin Hening
Published: May 2022

All radical‐SAM enzymes require [4Fe‐4S] clusters for activity. It is well known the [4Fe‐4S] cluster is very air sensitive and requires strict anaerobic conditions to reconstitute activity in vitro. However, aerobic species that live under oxygen do use radical SAM enzymes. How such enzymes function in vivo in the presence of oxygen is an interesting question. My lab works on the yeast diphthamide biosynthetic noncanonical radical‐SAM enzyme Dph1‐Dph2. Consistent with the known oxygen sensitivity, we found that in vitro the [4Fe‐4S] cluster in Dph1‐Dph2 is easily degraded into a [3Fe‐4S] cluster. Remarkably, a small iron‐containing protein Dph3 can donate one Fe atom to convert the [3Fe‐4S] cluster in Dph1‐Dph2 to a functional [4Fe‐4S] cluster during the radical‐SAM enzyme catalytic cycle. We believe this is a general strategy to solve the oxygen sensitivity problem and Dph3‐like proteins may exist to keep other radical‐SAM enzymes functional in aerobic environments.
Reprogramming T cells to target glycans and overcome glycan‐mediated immunosuppression for cancer therapy
Authors: Posey Avery D.
Published: May 2022

T‐lymphocytes (or T‐cells) exhibit rapid proliferation and cytotoxicity after immune recognition of foreign or non‐self antigens. Cancerous cells can often display mutated peptides that can be recognized as non‐self by the immune system, but a frequent mechanism of tumor escape is loss of the antigen display complexes. Chimeric antigen receptor (CAR) technology allows T‐cells to be genetically engineered with a novel immune receptor (as well as additional pro‐inflammatory or effector molecules) that redirect T‐cell specificity based on the targets of monoclonal antibodies. This genetic engineering allows the manufacturing of billions of T‐cells specific for one tumor‐associated antigen.

Previously, we designed a CAR T‐cell therapy that targets the glycopeptide epitope Tn‐MUC1 and demonstrated anti‐tumor efficacy against a variety of tumor histotypes, including T‐cell lymphoma, pancreatic cancer, breast cancer, prostate cancer, and other cancer histotypes. Now, we extend the targeting of Tn‐glycopeptides to an oncofetal form of fibronectin, recognized by the antibody FDC‐6. Tn‐fibronectin‐targeting CAR T‐cells exhibit similar efficacy against PC3 prostate tumors as CAR T‐cells targeting cell surface antigens; however, the mechanism of action of cytotoxicity appears to be different, suggesting that T‐cells targeting extracellular antigens may represent a novel platform for inducing anti‐tumor efficacy. Additionally, CAR T‐cell activity against tumors can be improved through additional engineering approaches, such as the use of enzymatic activity, the generation of immunomodulatory fusion proteins, and secretion of large molecules, such as cytokines and antibodies.
Orchestrated Intragranular Restructuring of Mucins During Secretory Granule Maturation
Authors: Ten Hagen Kelly G. , Syed Zulfeqhar A. , Zhang Liping , Tran Duy T. , Bleck Christopher K. E.
Published: May 2022

Regulated secretion is an essential process where proteins are packaged into membranous secretory vesicles. However, the details of cargo packaging and secretory granule maturation are largely unknown. Here, we demonstrate that multiple distinct proteins undergo orchestrated intragranular restructuring during secretory granule maturation in vivo, to allow spatial segregation of distinct components within the same granule. Furthermore, through a combination of genetics and multimodality imaging, we demonstrate the molecular identity of each distinct intragranular structure. We further identify genes that are essential for the temporally‐ordered restructuring events, including those controlling pH (vha16.1), Cl‐ ions (Clic and ClC‐c) and Ca2+ ions (fwe). Finally, we show that altered cargo glycosylation influences dimensions of these structures, thereby affecting secretory granule morphology. This study elucidates key steps and factors involved in intragranular, rather than intergranular, segregation of cargo through regulated restructuring events during secretory granule maturation. Understanding how multiple distinct proteins are efficiently packaged into and secreted from the same secretory granule may provide insight into diseases resulting from defects in secretion.
Polyubiquitin effects on phase transitions of shuttle protein UBQLN2
Authors: Castaneda Carlos A. , Dao Thuy P. , Yang Yiran
Published: May 2022

ALS‐linked ubiquitin‐binding shuttle protein UBQLN2 mediates crosstalk between proteasomal degradation and autophagy pathways, which are thought to be controlled by interactions involving K48‐linked and K63‐linked polyubiquitin chains, respectively. We previously showed that UBQLN2 is recruited to stress granules in cells and undergoes liquid‐liquid phase separation (LLPS) in vitro. Interactions with ubiquitin or multivalent K48‐linked chains inhibit UBQLN2 LLPS. Here, we show that other multivalent chains of longer lengths, including K63‐ and M1‐linked chains and a designed tetrameric ubiquitin construct that mimics a multi‐monoubiquitinated substrate, significantly enhanced UBQLN2 LLPS over a wide range of Ub:UBQLN2 ratios. With nuclear magnetic resonance (NMR) spectroscopy and complementary biophysical techniques, we demonstrated that these opposing effects stem from differences in polyUb chain conformations, but not in affinities between polyUb chains and UBQLN2. Using microscopy and turbidity assay experiments, we obtained multi‐component phase diagrams to demonstrate that compact K11 and K48‐linked Ub4 inhibit UBQLN2 LLPS whereas extended K63 and M1‐linked Ub4 stabilize UBQLN2 LLPS. Increasing chain flexibility and accessibility to the ubiquitin binding surface enables a switch between homotypically‐driven to partially heterotypically‐driven UBQLN2 phase separation. These observations provide mechanistic insights into how UBQLN2 could differentiate between proteasomal degradation and autophagy pathways via LLPS.
Exosomes as Key Regulators of Neutrophil Chemotaxis
Authors: Parent Carole A. , Arya Subhash B. , Chen Song , Javed Fatima
Published: May 2022

Neutrophils, the most abundant leukocytes in human blood, are the first line of defense at sites of injury or infection. Upon activation by primary chemoattractants such as the formyl peptide N‐Formyl‐methionine‐leucyl‐phenylalanine (fMLF), neutrophils secrete the secondary chemoattractant leukotriene B4 (LTB4) which serves to relay primary chemotactic signals and increase the range and persistence of migration. LTB4 is synthesized from arachidonic acid (AA) through the sequential action of the 5‐lipoxygenase (5‐LO), its associated activating protein (FLAP) and leukotriene A4 hydrolase (LTA4H). We previously demonstrated that LTB4 and its synthesizing enzymes are packaged and released in extracellular vesicles called exosomes. Conventional exosome biogenesis originates from the plasma membrane. However, the LTB4 synthesizing machinery resides on the nuclear envelope (NE). We found that the localized activation of membrane curvature inducing proteins facilitates the selective sorting of the LTB4‐synthesis machinery on the NE and its packaging within NE‐derived exosomes. We show that chemoattractant stimulation leads to the generation of 5‐LO‐ and FLAP‐positive NE buds and cytosolic vesicles, which are also positive for the inner nuclear membrane protein lamin B receptor (LBR). Furthermore, we found that the generation of ceramide from sphingomyelin through the action of the neutral sphingomyelinase 1 (nSMase1) is essential for the generation of these NE buds and cytosolic vesicles by generating ordered lipid microdomains on the NE. Together, our findings show that the spatiotemporal orchestration of chemoattractant‐initiated events leads to the recruitment of the LTB4 biogenesis machinery on the NE followed by NE‐budding and subsequent release of cytosolic vesicles containing exosomes. We envision that this novel mechanism will represent an important means of exosome biogenesis for the release of NE‐associated components in other cells, including immune cells and invasive cancer cells.
Single Cell transcriptomes highlight the temporal hierarchy of fate decisions of the developmental tree from Neural Border Stages to Migration of NC Cells
Authors: Monsoro‐Burq Anne‐Helene , Alexandr Kotov , Seal Subham , Alkobtawi Mansour , Kappes Vincent
Published: May 2022

Gene regulatory networks govern the steps of embryonic development and control cell transcriptional states. The neural crest cells are an archetypal example of cellular diversification from a multipotent progenitor population. However, the full sequence of molecular choices governing the emergence of neural crest heterogeneity from the ectoderm state remains elusive. Here we combine ultra‐dense single cell transcriptomes with machine‐learning strategies and experimental validation to provide a comprehensive gene regulatory network driving vertebrate neural crest fate diversification from induction to early migration stages. Transcription factor connectome and bifurcation analyses discover that several early neural crest fates emerge at the neural plate stage, alongside an unbiased multipotent neural crest lineage persisting until after epithelial‐mesenchymal transition. We also define a new and transient neural border zone state, preceding choice between neural crest and placodes during gastrulation. Our comprehensive experimental approach combined to several ML strategies broadly applicable to single cell transcriptomics deciphers the circuits driving cranial and vagal neural crest formation and provides a general model for investigating vertebrate GRNs in development, evolution and disease
Learning the Legs When You Have None
Authors: Buckley Miracle
Published: May 2022

Living life in a wheelchair was never part of the plan, but life comes the way it comes. The speaker has endured a spinal surgery combined with a lower extremity amputation, proceeded through public schools, learning to play piano without the use of pedals, learning to drive, ultimately deciding a future in the medical field was the way to go. One of the biggest decisions that would face her would be deciding to not want a wheelchair to define her life. Which is why she always chose things that were considered for “able‐bodied” people. Throughout her education, there have always been classes that have been a bit of a struggle. However, one of the hardest was Gross Anatomy.

This session will focus on the speaker’s lived experience as a bilateral lower extremity amputee navigating life and the health professional curriculum. For most people anatomy class is a little easier because they have all their limbs and can demonstrate on their own. But for her, not having those major parts, it was a bit of a struggle until her anatomy professor helped relate the lower extremity to the upper extremity. She can ambulate on her hands when not using her power wheelchair, so this technique helped to make that topic stick. This speaker will help demonstrate how anatomy educators can facilitate success for students with disabilities through an individualized approach.
Making the Human Face: Elucidating the Role of Enhancers in Hominid Craniofacial Evolution
Authors: Mohammed Jaaved , Hansen Karissa , Claes Peter , Weinberg Seth , Selleri Licia , Swigut Tomek , Wysocka Joanna
Published: May 2022

There is increasing evidence that morphological differences among individuals of the same species and between species are driven by changes in gene expression mediated by non‐coding mutations that are enriched at enhancer elements. The identification and comparative evaluation of these cis‐regulatory variants specific to human facial morphology have been facilitated by several recent advances. Among these are access to high‐quality genome sequences, especially from archaic hominins such as the Neanderthals, by expanded catalogs of SNPs implicated in normal range human facial variation from Genome‐wide Association Studies (GWAS), and by the curation of active human and chimp Cranial Neural Crest Cell (CNCC) enhancers that are important for facial development. To identify functional variants that mediate change in enhancer activity in recent human evolution and normal range variation and to nominate their target genes we employed two methods: (1) STARR‐seq, an enhancer reporter assay that permits accurate, comparative analysis of regulatory activity across enhancer homologs and their constituent cis‐mutations, and (2) H3K27ac HiChIP, a method to characterize the three‐dimensional interactome between enhancers and their target genes.

We performed STARR‐seq and HiChIP experiments in the in vitro derived CNCCs, and early proliferative cranial chondrocytes. Comparisons of enhancer activity among the orthologs present in homo‐ or hominin species uncovered several classes of enhancers with species‐ or clade‐ restricted activity. Furthermore, for enhancers associated with facial shape GWAS signals, we identified genetic variants associated with changes in regulatory activity. By incorporating HiChIP labeled target genes, we found that biased intra‐species and GWAS enhancer homologs target important developmental transcription factors and signaling molecules. Our experiments have begun to provide insights into how evolutionarily important and functionally relevant enhancer cis‐variants can modulate gene regulatory programs at the individual‐ or species‐centric level with an outcome on facial phenotypes.
The Imperative for Scientific Societies to Change the Face of Academia: Recommendations for Immediate Action
Authors: Jerome‐Majewska Loydie A. , Carroll Melissa A. , Boynes Shawn , Topp Kimberly S.
Published: May 2022

Scientific societies have an opportunity, and a responsibility, to drive inclusion, diversity, equity, and accessibility in science in academia. The American Association for Anatomy (AAA) can serve as a model of best practice for other professional associations working to become more inclusive of individuals from historically underrepresented groups. In this presentation, we will discuss anatomy's exclusionary past, describe the current face of science in academia, and present our recommendations, including to the AAA, to accelerate change in academia. We recommend that scientific societies investigate inequities and revise practices for inclusivity; develop and empower underrepresented minority leadership; and commit resources in a sustained manner as an investment in scientists who bring diverse perspectives and lived experiences to science in academia.