Medicine and Health Sciences Commons^™

Mdm2 Is Required For Survival And Growth Of P53-Deficient Cancer Cells., Kyle P Feeley, Clare M. Adams, Ramkrishna Mitra, Christine M. Eischen

Department of Cancer Biology Faculty Papers

p53 deletion prevents the embryonic lethality of normal tissues lacking Mdm2, suggesting that cells can survive without Mdm2 if p53 is also absent. Here we report evidence challenging this view, with implications for therapeutically targeting Mdm2. Deletion of Mdm2 in T-cell lymphomas or sarcomas lacking p53 induced apoptosis and G₂ cell-cycle arrest, prolonging survival of mice with these tumors. p53^-/- fibroblasts showed similar results, indicating that the effects of Mdm2 loss extend to pre-malignant cells. Mdm2 deletion in p53^-/- cells upregulated p53 transcriptional target genes that induce apoptosis and cell-cycle arrest. Mdm2 deletion also increased levels of …

Δfosb Regulates Gene Expression And Cognitive Dysfunction In A Mouse Model Of Alzheimer's Disease., Brian F Corbett, Jason C You, Xiaohong Zhang, Mark S Pyfer, Umberto Tosi, Daniel M Iascone, Iraklis Petrof, Anupam Hazra, Chia-Hsuan Fu, Gabriel S Stephens, Annie Ashok, Suzan Aschmies, Lijuan Zhao, Eric J Nestler, Jeannie Chin

Department of Neuroscience Faculty Papers

Alzheimer's disease (AD) is characterized by cognitive decline and 5- to 10-fold increased seizure incidence. How seizures contribute to cognitive decline in AD or other disorders is unclear. We show that spontaneous seizures increase expression of ΔFosB, a highly stable Fos-family transcription factor, in the hippocampus of an AD mouse model. ΔFosB suppressed expression of the immediate early gene c-Fos, which is critical for plasticity and cognition, by binding its promoter and triggering histone deacetylation. Acute histone deacetylase (HDAC) inhibition or inhibition of ΔFosB activity restored c-Fos induction and improved cognition in AD mice. Administration of seizure-inducing agents to nontransgenic …

Corrosion And Tribology Of Materials Used In A Novel Reverse Hip Replacement, Linda Braddon, Zafer Termanini, Steven Macdonald, Jay Parvizi, Jay Lieberman, Victor Frankel, Joseph Zuckerman

Rothman Institute Papers

Total hip arthroplasty has been utilized for the past 50 years as an effective treatment for degenerative, inflammatory and traumatic disorders of the hip. The design of these implants has generally followed the anatomy of the hip as a ball and socket joint with the femoral head representing the ball and the acetabulum representing the socket. We describe a novel hip arthroplasty design in which the "ball" is located on the acetabular side and the "socket" is located on the femoral side. The results of extensive biomechanical testing are described and document wear and corrosion characteristics that are at least …

A Word From The Writing Center (July 2017), Jennifer Wilson, Ms, Els

A Word From the Writing Team (Newsletter)

This issue includes:

• Grammar tip: affect vs effect
• Opportunity for online writing workshop for scientists

New Interleukin-15 Superagonist (Il-15sa) Significantly Enhances Graft-Versus-Tumor Activity., Cavan P Bailey, Tulin Budak-Alpdogan, Christopher T Sauter, Michelle M Panis, Cihangir Buyukgoz, Emily K Jeng, Hing C Wong, Neal Flomenberg, Md, Onder Alpdogan

Department of Medical Oncology Faculty Papers

Interleukin-15 (IL-15) is a potent cytokine that increases CD8+ T and NK cell numbers and function in experimental models. However, obstacles remain in using IL-15 therapeutically, specifically its low potency and short in vivo half-life. To help overcome this, a new IL-15 superagonist complex comprised of an IL-15N72D mutation and IL-15RαSu/Fc fusion (IL-15SA, also known as ALT-803) was developed. IL-15SA exhibits a significantly longer serum half-life and increased in vivo activity against various tumors. Herein, we evaluated the effects of IL-15SA in recipients of allogeneic hematopoietic stem cell transplantation. Weekly administration of IL-15SA to transplant recipients significantly increased the number …

Jefferson Digital Commons Quarterly Report (April-June 2017) Now Available, Daniel G. Kipnis, Msi

Jefferson Digital Commons News

The Jefferson Digital Commons quarterly report for April-June 2017 is now available. Users from 7,500 institutions in 196 countries located materials in the JDC this quarter. View the report to see what assets were added this quarter.

Lack Of Evidence For Involvement Of Tonebp And Hyperosmotic Stimulus In Induction Of Autophagy In The Nucleus Pulposus., Chao Liu, Hyowon Choi, Zariel I. Johnson, Jiwei Tian, Irving Shapiro, Makarand V Risbud

Department of Orthopaedic Surgery Faculty Papers

Nucleus pulposus (NP) cells reside in a physiologically hyperosmotic environment within the intervertebral disc. TonEBP/NFAT5 is an osmo-sensitive transcription factor that controls expression of genes critical for cell survival under hyperosmotic conditions. A recent report on NP and studies of other cell types have shown that hyperosmolarity triggers autophagy. However, little is known whether such autophagy induction occurs through TonEBP. The goal of this study was to investigate the role of TonEBP in hyperosmolarity-dependent autophagy in NP. Loss-of-function studies showed that autophagy in NP cells was not TonEBP-dependent; hyperosmolarity did not upregulate autophagy as previously reported. NP tissue of haploinsufficient …

Atrophy Of Gluteus Maximus Among Women With A History Of Chronic Low Back Pain., Amy H. Amabile, John H. Bolte, Saskia D. Richter

Department of Physical Therapy Faculty Papers

BACKGROUND: Although the relationship between low back pain (LBP) and the size of certain trunk muscles has been extensively studied, the relationship between gluteus maximus (GM) size and LBP has been only minimally examined. Determining whether such a relationship exists would help improve our understanding of the etiology of LBP, and possibly provide a rationale for the use of therapeutic exercise interventions targeting GM with LBP patients. The objective of this study was to compare gluteus maximus cross-sectional area in individuals with chronic LBP, and in a group of individuals without LBP. Our hypothesis was that individuals with LBP would …

Jefferson Digital Commons Quarterly Report: April-June 2017, Daniel G. Kipnis, Msi, Kelsey Duinkerken, Msi, James Copeland, Libt

Jefferson Digital Commons Reports

For Full-Text access download report.

Jefferson Digital Commons Quarterly Report: April-June 2017

The Jefferson Digital Commons had another very busy quarter with assets being deposited from across the enterprise.

• Total number of assets deposited this quarter: 371
• Total downloads for this quarter: 113,261(April 10, 2017 highest download count: 1,763)
• 7,500 different institutions accessed content in the JDC Asset with the most downloads this quarter: 2017 issue of Inside Out
• Countries visiting the JDC this quarter: 196

Below are highlighted assets that were added to the inventory along with more feedback from researchers that continue to arrive from around the globe …

Cyclin D1 Restrains Oncogene-Induced Autophagy By Regulating The Ampk-Lkb1 Signaling Axis., Mathew C. Casimiro, Gabriele Disante, Agnese Di Rocco, Emanuele Loro, Claudia Pupo, Timothy G. Pestell, Sara Bisetto, Marco A. Velasco-Velázquez, Xuanmao Jiao, Zhiping Li, Christine M. Kusminski, Erin L. Seifert, Chenguang Wang, Daniel Ly, Bin Zheng, Che-Hung Shen, Philipp E. Scherer, Richard Pestell

Department of Cancer Biology Faculty Papers

Autophagy activated after DNA damage or other stresses mitigates cellular damage by removing damaged proteins, lipids, and organelles. Activation of the master metabolic kinase AMPK enhances autophagy. Here we report that cyclin D1 restrains autophagy by modulating the activation of AMPK. In cell models of human breast cancer or in a cyclin D1-deficient model, we observed a cyclin D1-mediated reduction in AMPK activation. Mechanistic investigations showed that cyclin D1 inhibited mitochondrial function, promoted glycolysis, and reduced activation of AMPK (pT172), possibly through a mechanism that involves cyclin D1-Cdk4/Cdk6 phosphorylation of LKB1. Our findings suggest how AMPK activation by cyclin D1 …

Metabolite Profiling Of Infection-Associated Metabolic Markers Of Onchocerciasis., Sasisekhar Bennuru, Sara Lustigman, David Abraham, Thomas B. Nutman

Department of Microbiology and Immunology Faculty Papers

The global efforts for onchocerciasis elimination may require additional tools (safe micro and macrofilaricidal drugs, vaccines and biomarkers) as elimination efforts move toward the "end game". Efforts toward the identification of suitable biomarkers have focused on specific protein(s) and/or nucleic acids, but metabolites present an alternative option as they have limited half-lives and are the result of combinatorial effects. In comparison to previously used methodology of LC-MS for metabolomic approaches, we used a non-targeted capillary electrophoresis time-of-flight mass spectrometry (CE-TOFMS) to analyze the serum metabolic profiles of Ov-infected and -uninfected individuals (n=20). We identified 286 known metabolites (167 in the …

Biochemical Effects Of Exercise On A Fasciocutaneous Flap In A Rat Model., Edita Aksamitiene, Adam L. Baker, Sudeep Roy, Salini Hota, Li-Hui Zhang, Julianna Rodin, Kealan Hobelmann, Jan B. Hoek, Edmund A. Pribitkin

Department of Otolaryngology - Head and Neck Surgery Faculty Papers

Importance: An overwhelming amount of data suggest that cardiovascular exercise has a positive effect on the mind and body, although the precise mechanism is not always clear.

Objective: To assess the clinical and biochemical effects of voluntary cardiovascular exercise on pedicled flaps in a rodent model.

Design, Setting, and Participants: Eighteen adult Sprague-Dawley male rats were randomized into a resting animal group (RAG) (n=9) and an exercise animal group (EAG) (n=9) for 14 days (July 23, 2013, through July 30, 2013). A pedicled transposition flap was performed on the ventral surface of the rat, and biopsy specimens were taken from …

Ixazomib Enhances Parathyroid Hormone-Induced Β-Catenin/T-Cell Factor Signaling By Dissociating Β-Catenin From The Parathyroid Hormone Receptor., Yanmei Yang, Hong Lei, Ya-Wei Qiang, Bin Wang

Center for Translational Medicine Faculty Papers

The anabolic action of PTH in bone is mostly mediated by cAMP/PKA and Wnt-independent activation of β-catenin/T-cell factor (TCF) signaling. β-Catenin switches the PTH receptor (PTHR) signaling from cAMP/PKA to PLC/PKC activation by binding to the PTHR. Ixazomib (Izb) was recently approved as the first orally administered proteasome inhibitor for the treatment of multiple myeloma; it acts in part by inhibition of pathological bone destruction. Proteasome inhibitors were reported to stabilize β-catenin by the ubiquitin-proteasome pathway. However, how Izb affects PTHR activation to regulate β-catenin/TCF signaling is poorly understood. In the present study, using CRISPR/Cas9 genome-editing technology, we show that …

Cesarean In The Second Stage: A Possible Risk Factor For Subsequent Spontaneous Preterm Birth., Vincenzo Berghella, Alexis C. Gimovsky, Lisa D. Levine, Joy Vink

Department of Obstetrics and Gynecology Faculty Papers

No abstract provided.

Lung Rest During Extracorporeal Membrane Oxygenation For Neonatal Respiratory Failure-Practice Variations And Outcomes., Deepthi Alapati, Zubair H. Aghai, Jobayer Hossain, Daniel R Dirnberger, Mark T. Ogino, Thomas H. Shaffer

Department of Pediatrics Faculty Papers

OBJECTIVE: Describe practice variations in ventilator strategies used for lung rest during extracorporeal membrane oxygenation for respiratory failure in neonates, and assess the potential impact of various lung rest strategies on the duration of extracorporeal membrane oxygenation and the duration of mechanical ventilation after decannulation.

DATA SOURCES: Retrospective cohort analysis from the Extracorporeal Life Support Organization registry database during the years 2008-2013.

STUDY SELECTION: All extracorporeal membrane oxygenation runs for infants less than or equal to 30 days of life for pulmonary reasons were included.

DATA EXTRACTION: Ventilator type and ventilator settings used for lung rest at 24 hours after …

Intravitreal Melphalan For Treatment Of Primary Vitreoretinal Lymphoma: A New Indication For An Old Drug., Carol L. Shields, Kareem Sioufi, Arman Mashayekhi, Jerry A. Shields

Wills Eye Hospital Papers

No abstract provided.

Non-Adherence To Eye Care In People With Diabetes, Ann P. Murchison, Lisa A. Hark, Laura T. Pizzi, Yang Dai, Eileen Mayro, Philip P. Storey, Benjamin E. Leiby, Julia A. Haller

Wills Eye Hospital Papers

Objective Evaluate individual factors that impact adherence to eye care follow-up in patients with diabetes. Design and methods A 4-year retrospective chart review was conducted for 1968 patients with diabetes over age 40 from an urban academic center. Data collected included demographics, insurance, visual acuity, smoking status, medications, dates of dilated fundus examinations (DFE), and reported hemoglobin A1C and blood glucose levels. The primary outcome was timely DFE follow-up adherence following the initial eye exam visit. Results Overall, 41.6% of patients adhered to initial follow-up eye care recommendations. Multivariable analysis demonstrated that patients with severe diabetic retinopathy (DR) were more …

Autoimmune Retinopathy Associated With Systemic Lupus Erythematosus: A Diagnostic Dilemma., Wadakarn Wuthisiri, Yu-Hung Lai, Jenina Capasso, Martin Blidner, David Salz, Erik Kruger, Alex V. Levin

Wills Eye Hospital Papers

Visual loss in systemic lupus erythematosus (SLE) due to autoimmune retinopathy (AIR) is rare and easily misdiagnosed as hydroxychloroquine retinopathy. We report the rare clinical presentation of severe visual loss in a patient with SLE due to nonparaneoplastic AIR as differentiated from hydroxychloroquine toxicity. A 70-year-old female diagnosed and treated for lupus for 17 years and had been taking hydroxychloroquine for 15 years. Over the past 2 years, she developed progressive peripheral visual loss oculus uterque which rapidly advanced in the latter 6 months. Hydroxychloroquine toxicity was initially suspected, but diagnostic testing revealed a retinal degeneration. Antiretinal autoantibody testing using …

Adjuvant Radiation Therapy, Androgen Deprivation, And Docetaxel For High-Risk Prostate Cancer Postprostatectomy: Results Of Nrg Oncology/Rtog Study 0621., Mark D. Hurwitz, Jonathan Harris, Oliver Sartor, Ying Xiao, Bobby Shayegan, Paul W. Sperduto, Kasra R. Badiozamani, Colleen A.F. Lawton, Eric M. Horwitz, Jeff M. Michalski, Kevin Roof, David C. Beyer, Qiang Zhang, Howard M. Sandler

Department of Radiation Oncology Faculty Papers

BACKGROUND: Phase 3 trials have demonstrated a benefit from adjuvant radiation therapy (ART) for men who have adverse factors at radical prostatectomy (RP). However, some patients have a high risk of progression despite ART. The role of systemic therapy with ART in this high-risk group remains to be defined.

METHODS: Patients who had either a post-RP prostate-specific antigen (PSA) nadir > 0.2 ng/mL and a Gleason score ≥7 or a PSA nadir ≤0.2 ng/mL, a Gleason score ≥8, and a pathologic tumor (pT) classification ≥ pT3 received 6 months of androgen-deprivation therapy (ADT) plus radiotherapy and 6 cycles of docetaxel. The …

Functional Selectivity Of Gpcr-Directed Drug Action Through Location Bias., Roshanak Irannejad, Veronica Pessino, Delphine Mika, Bo Huang, Philip B. Wedegaertner, Marco Conti, Mark Von Zastrow

Department of Biochemistry and Molecular Biology Faculty Papers

G-protein-coupled receptors (GPCRs) are increasingly recognized to operate from intracellular membranes as well as the plasma membrane. The β 2 -adrenergic GPCR can activate G s -linked cyclic AMP (G s -cAMP) signaling from endosomes. We show here that the homologous human β 1 -adrenergic receptor initiates an internal G s -cAMP signal from the Golgi apparatus. By developing a chemical method to acutely squelch G-protein coupling at defined membrane locations, we demonstrate that Golgi activation contributes significantly to the overall cellular cAMP response. Golgi signaling utilizes a preexisting receptor pool rather than receptors delivered from the cell surface, requiring …

Evaluating The Relationship Between Participation In Student-Run Free Clinics And Changes In Empathy In Medical Students., Anita Modi, Michele Fascelli, Zachary Daitch, Mohammadreza Hojat Professor

Department of Medicine Faculty Papers

PURPOSE: We explored differences in changes in medical student empathy in the third year of medical school between volunteers at JeffHOPE, a multisite medical student-run free clinic of Sidney Kimmel Medical College (SKMC), and nonvolunteers.

METHOD: Volunteerism and leadership experience at JeffHOPE were documented for medical students in the Class of 2015 (n = 272) across their medical educations. Students completed the Jefferson Scale of Empathy at the beginning of medical school and at the end of the third year. Students who reported participation in other Jefferson-affiliated clinics (n = 44) were excluded from this study. Complete data were available …

Resolution Of Occlusive Carotid Artery Thrombus Treated With Anticoagulation As Demonstrated On Duplex Ultrasonography., Laura Stein, John W. Liang, Jesse Weinberger

Department of Neurology Faculty Papers

BACKGROUND AND PURPOSE: While the majority of cerebral ischemic events due to carotid occlusive disease result from atherosclerotic plaque rupture, intraluminal carotid artery thrombus occasionally occurs in patients without preexisting carotid atherosclerosis. Identification of nonatherosclerotic thrombus as the cause of the carotid occlusive disease can obviate the need for an interventional procedure, and resolution of thrombus can be monitored with B-mode duplex ultrasonography.

METHODS: We reviewed 3 patients treated on The Mount Sinai Hospital Stroke Unit with anticoagulation for nonatherosclerotic carotid thrombi and followed with serial Doppler ultrasonogrpahy for resolution of thrombus.

RESULTS: Occlusive carotid thrombus was successfully treated in …

Connecting The Brain To Itself Through An Emulation., Mijail D. Serruya

Department of Neurology Faculty Papers

Pilot clinical trials of human patients implanted with devices that can chronically record and stimulate ensembles of hundreds to thousands of individual neurons offer the possibility of expanding the substrate of cognition. Parallel trains of firing rate activity can be delivered in real-time to an array of intermediate external modules that in turn can trigger parallel trains of stimulation back into the brain. These modules may be built in software, VLSI firmware, or biological tissue as in vitro culture preparations or in vivo ectopic construct organoids. Arrays of modules can be constructed as early stage whole brain emulators, following canonical …

A Phase Ii Randomized Trial Of Observation Versus Stereotactic Ablative Radiation For Oligometastatic Prostate Cancer (Oriole)., Noura Radwan, Ryan Phillips, Ashley Ross, Steven P. Rowe, Michael A. Gorin, Emmanuel S. Antonarakis, Curtiland Deville, Stephen Greco, Samuel Denmeade, Channing Paller, Daniel Y. Song, Maximilian Diehn, Hao Wang, Michael Carducci, Kenneth J. Pienta, Martin G. Pomper, Theodore L. Deweese, Adam Dicker, Mario Eisenberger, Phuoc T. Tran

Department of Radiation Oncology Faculty Papers

BACKGROUND: We describe a randomized, non-blinded Phase II interventional study to assess the safety and efficacy of stereotactic ablative radiotherapy (SABR) for hormone-sensitive oligometastatic prostate adenocarcinoma, and to describe the biology of the oligometastatic state using immunologic, cellular, molecular, and functional imaging correlates. 54 men with oligometastatic prostate adenocarcinoma will be accrued. The primary clinical endpoint will be progression at 6 months from randomization with the hypothesis that SABR to all metastases will forestall progression by disrupting the metastatic process. Secondary clinical endpoints will include local control at 6 months post-SABR, toxicity and quality of life, and androgen deprivation therapy …

The Effects Of C5-Substituted 2,4-Diaminoquinazolines On Selected Transcript Expression In Spinal Muscular Atrophy Cells., Cinsley Gentillon, Andrew J Connell, Ryan W Kirk, Matthew E R Butchbach

Department of Pediatrics Faculty Papers

C5-substituted 2,4-diaminoquinazolines (2,4-DAQs) ameliorate disease severity in SMA mice. It is uncertain, however, that these compounds increase SMN protein levels in vivo even though they were identified as activators of the SMN2 promoter. These compounds also regulate the expression of other transcripts in neuroblastoma cells. In this study, we investigate the mechanism by which the 2,4-DAQs regulate the expression of SMN2 as well as other targets. D156844, D158872, D157161 and D157495 (RG3039) increased SMN2 promoter-driven reporter gene activity by at least 3-fold in NSC-34 cells. These compounds, however, did not significantly increase SMN2 mRNA levels in type II SMA fibroblasts …

Dach1 Suppresses Breast Cancer As A Negative Regulator Of Cd44., Hanxiao Xu, Shengnan Yu, Xun Yuan, Jing Xiong, Dong Kuang, Richard Pestell, Kongming Wu

Department of Cancer Biology Faculty Papers

Dachshund homolog 1 (DACH1), a key cell fate determination factor, contributes to tumorigenesis, invasion, metastasis of human breast neoplasm. However, the exact molecular mechanisms for the anti-tumor roles of DACH1 in breast carcinoma are still lack of extensive understanding. Herein, we utilized immunohistochemistry (IHC) staining and public microarray data analysis showing that DACH1 was higher in normal breast, low-grade and luminal-type cancer in comparison with breast carcinoma, high-grade and basal-like tumors respectively. Additionally, both correlation analysis of public databases of human breast carcinoma and IHC analysis of mice xenograft tumors demonstrated that DACH1 inversely related to cancer stem cells (CSCs) …

Resident Reflection: Mrs. C, Rachel Nash, Md

The Medicine Forum

“Make friends with those critical care nurses in the ICU,” my friend from high school and general floor nurse whispered in my ear before the start of my MICU rotation. “They can really save your butt.” This was one of my many goals I set out to achieve during my first month in the intensive care unit as an intern. I wanted to become more comfortable managing very sick patients, get some procedures under my belt, and mostly “survive” the emotionally and physically taxing weeks that were sure to come.

The sign out I received on my 3-patient load suddenly …

Grand Rounds Panel Discussion: Provider Payment Reform: Past, Present, And Future, Neha Bansal Etherington, Md, Shuwei Wang, Md, Debbie Chen, Md, Anita Modi, Md, Jad Al Danaf, Md

The Medicine Forum

Panelists:

Gregory C. Kane, MD, FACP The Jane and Leonard Korman, Professor and Chairman, Department of Medicine

David B. Nash, MD, MBA The Dr Raymond C and Doris N Grandon Professor, and Dean of the Jefferson College of Population Health

Rhea E. Powell, MD, MPH Assistant Professor of Medicine

Rachel B. Sorokin, MD, FACP Associate Professor of Medicine and Medical Director of Safety and Quality

Lawrence D. Ward, MD, MPH Associate Professor of Medicine and Vice Chairman for Clinical Practice and Quality

Introduction:

The impetus for provider payment reform in healthcare grew out of the recognition that growth in healthcare …

Fatal Rituximab-Induced Nonspecific Interstitial Pneumonia: Case Report And Review Of The Literature, Rino Sato, Md, Jeremy Molligan, Md, Sameh Gaballa, Md

The Medicine Forum

Introduction:

Rituximab is a chimeric anti-CD20 monoclonal antibody that is used to treat some hematological malignancies such as B-cell lymphomas, and various autoimmune diseases including immune thrombocytopenic purpura, systemic lupus erythematous, rheumatoid arthritis, and autoimmune hemolytic anemia. The most common side effects include fever, chills, and rigors. Respiratory complications such as cough, bronchospasm, sinusitis, and rhinitis have also been reported in 30% of patients in clinical trials.1 Rituximab-induced lung injury is a very rare but potentially fatal complication. We report a case of fatal single agent rituximab-induced nonspecific interstitial pneumonia to increase awareness about this serious side effect and review …

Brugada Syndrome: Diagnosis And Management, Amit Vira, Md, Ehsanur Rahman, Md

The Medicine Forum

Introduction:

The vast majority of cases of sudden cardiac arrest (SCA) are caused by ventricular tachyarrhythmias (VT), with most cases associated with structural heart disease. SCA in a structurally normal heart is an uncommon occurrence, most often occurring in the third decade of life and accounting for 5-15% of total SCA cases1-2. Common causes include Brugada syndrome, congenital long QT syndrome, catecholaminergic polymorphic ventricular tachycardia, arrhythmogenic right ventricular cardiomyopathy, and idiopathic ventricular tachycardia1-3. The majority of these patients with a structurally normal heart are undiagnosed until presenting with sudden cardiac death. It is believed that cardiac arrest is the initial …