Mary Elizabeth Sunday

Background/Objectives: Cripto-1 (CR1) is a plurifunctional embryonic protein required for implantation and re-expressed in the adult during wound repair, inflammation, and tumorigenesis. CR1 and its predicted CR1 pseudogene product Cripto-3/CR3 are highly homologous proteins, and given this physical attribute, commercially available antibodies cannot discriminate between CR1 and CR3. Methods: A series of mouse monoclonal antibodies [MoAbs] were developed with a high-affinity binding that can differentiate human CR1/CR3 proteins and showed no measurable cross-reactivity. Results: Using these reagents, we confirm that CR3 is a bona fide translated protein found in human tumor tissue, cancer cell lysates, and in normal/cancer patient donor sera. We also reveal that CR1 and CR3 compete for binding to signal transduction protein Nodal, glucose-regulated protein 78Da (GRP78), and activin receptor-like kinase 4 (Alk4). Our discriminatory MoAbs provide new reagents to help clarify current CR1/CR3 protein expression vagaries in the Cripto field of study, challenging established CR1 conventions. In addition, our data validate CR3 involvement in human carcinogenesis and cell signaling pathways, with potential clinical relevance in determining cancer patient prognosis and disease severity.

Ozone and obesity both increase IL-17A in the lungs. In mice, obesity augments the airway hyperresponsiveness and neutrophil recruitment induced by acute ozone exposure. Therefore, we examined the role of IL-17A in obesity-related increases in the response to ozone observed in obese mice. Lean wild-type and obese db/db mice were pretreated with IL-17A–blocking or isotype antibodies, exposed to air or ozone (2 ppm for 3 h), and evaluated 24 hours later. Microarray analysis of lung tissue gene expression was used to examine the mechanistic basis for effects of anti–IL-17A. Compared with lean mice, ozone-exposed obese mice had greater concentrations of BAL IL-17A and greater numbers of pulmonary IL-17A+ cells. Ozone-induced increases in BAL IL-23 and CCL20, cytokines important for IL-17A+ cell recruitment and activation, were also greater in obese mice. Anti–IL-17A treatment reduced ozone-induced airway hyperresponsiveness toward levels observed in lean mice. Anti–IL-17A treatment also reduced BAL neutrophils in both lean and obese mice, possibly because of reductions in CXCL1. Microarray analysis identified gastrin-releasing peptide (GRP) receptor (Grpr) among those genes that were both elevated in the lungs of obese mice after ozone exposure and reduced after anti–IL-17A treatment. Furthermore, ozone exposure increased BAL GRP to a greater extent in obese than in lean mice, and GRP-neutralizing antibody treatment reduced obesity-related increases in ozone-induced airway hyperresponsiveness and neutrophil recruitment. Our data indicate that IL-17A contributes to augmented responses to ozone in db/db mice. Furthermore, IL-17A appears to act at least in part by inducing expression of Grpr.

Summary: Bronchopulmonary dysplasia (BPD) is an inflammatory lung disorder common in premature infants who undergo mechanical ventilation with supplemental oxygen. Inhaled nitric oxide (iNO) has been used to prevent experimental and clinical BPD. Earlier studies showed that NO effects in alveolar epithelial cells (AEC) are mediated by S‐nitrosothiol uptake via L‐type amino acid transporter‐1 (LAT1). Because LAT1 expression could influence the efficacy of iNO therapy, we sought to determine whether pulmonary LAT1 expression is altered in preterm baboons with experimental BPD and in human newborns susceptible to developing BPD. Using fixed lung obtained from 125 d to 140 d gestation baboon models of BPD, LAT1 immunostaining was measured in control and BPD animals. In adult controls and in 140 d gestational controls (GC), LAT1 was expressed in both type I and type II AECs. In 140 d BPD lungs, LAT1 expression density in alveolar tissue was decreased. In 125 d GC baboons, LAT1 immunostaining was largely confined to cuboidal AECs, whereas animals given 14 d of mechanical ventilation exhibited diminished alveolar septal LAT1 Labeling. The pattern in adult human donor lung was comparable to that observed in adult baboons. LAT1 was expressed in lungs obtained from some but not all very premature newborns at autopsy. In human and baboon lung, adult and newborn, pulmonary vascular cells expressed LAT1. In summary, LAT1 is expressed in AECs and pulmonary vascular cells in baboons and humans. Experimental BPD in premature baboons decreases pulmonary LAT1 expression and alters its spatial localization. Heterogeneity of functional LAT1 could affect S‐nitrosothiol importation, which could impair iNO therapy. Pediatr Pulmonol. 2016;51:1048–1056.

Elastin synthesis and degradation in the airway and lung parenchyma contribute to airway mechanics, including airway patency and elastic recoil. IL-13 mediates many features of asthma pathobiology, including airway remodeling, but the effects of IL-13 on elastin architecture in the airway wall are not known. We hypothesized that IL-13 modulates elastin expression in airway fibroblasts from subjects with allergic asthma. Twenty-five subjects with mild asthma (FEV1, 89 ± 3% predicted) and 30 normal control subjects (FEV1, 102 ± 2% predicted) underwent bronchoscopy with endobronchial biopsy. Elastic fibers were visualized in airway biopsy specimens using Weigert’s resorcin-fuchsin elastic stain. Airway fibroblasts were exposed to IL-13; a pan-matrix metalloproteinase (MMP) inhibitor (GM6001); specific inhibitors to MMP-1, -2, -3, and -8; and combinations of IL-13 with MMP inhibitors in separate conditions in serum-free media for 48 hours. Elastin (ELN) expression as well as MMP secretion and activity were quantified. Results of this study show that elastic fiber staining of airway biopsy tissue was significantly associated with methacholine PC20 (i.e., the provocative concentration of methacholine resulting in a 20% fall in FEV1 levels) in patients with asthma. IL-13 significantly suppressed ELN expression in asthmatic airway fibroblasts as compared with normal control fibroblasts. The effect of IL-13 on ELN expression was significantly correlated with postbronchodilator FEV1/FVC in patients with asthma. MMP inhibition significantly stimulated ELN expression in patients with asthma as compared with normal control subjects. Specific inhibition of MMP-1 and MMP-2, but not MMP-3 or MMP-8, reversed the IL-13–induced suppression of ELN expression. In asthma, MMP-1 and MMP-2 mediate IL-13–induced suppression of ELN expression in airway fibroblasts.

Ledford, J. G., H. Goto, E. N. Potts, S. Degan, H. W. Chu, D. R. Voelker, M. E. Sunday, G. J. Cianciolo, W. M. Foster, M. Kraft, and J. R. Wright. 2009. SP-A preserves airway homeostasis during Mycoplasma pneumoniae infection in mice. J. Immunol. 182: [7818–7827][1]. We have recently become aware

A corrigendum on Oxygen, gastrin-releasing peptide, and pediatric lung disease: life in the balance by Sunday ME. Front Pediatr (2014) 2:72. doi: 10.3389/fped.2014.00072 Reference # 7 needs to be corrected to: Mary Sunday and Barbara Theriot, unpublished data. We have recently determined that most of the ovalbumin data in the article is unreliable. However, we have successfully validated the ozone data in additional experiments independent of those included in the article, which is the basis for the citation. The author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. Keywords: correction, ozone, gastrin-releasing peptide, lung, injury Citation: Sunday ME (2014) Corrigendum: Oxygen, gastrin-releasing peptide, and pediatric lung disease: life in the balance. Front. Pediatr. 2:102. doi: 10.3389/fped.2014.00102 Received: 13 August 2014; Accepted: 07 September 2014; Published online: 02 October 2014. Edited and reviewed by: Heber C. Nielsen, Tufts Medical Center and Tufts School of Medicine, USA Copyright: © 2014 Sunday. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. *Correspondence: mary.sunday@duke.edu

Excessive oxygen (O2) causes tissue injury, scarring, aging, and even death. Our laboratory is studying O2-sensing pulmonary neuroendocrine cells (PNECs) and the PNEC-derived product gastrin-releasing peptide (GRP). Reactive oxygen species (ROS) generated from exposure to hyperoxia, ozone, or ionizing radiation (RT) can induce PNEC degranulation and GRP secretion. PNEC degranulation can also be induced by hypoxia, and effects of hypoxia are also mediated by free radicals. We have determined that excessive GRP mediates lung injury with acute and chronic inflammation, leading to pulmonary fibrosis (PF), triggered via ROS exposure or by directly treating mice with exogenous GRP. In animal models, GRP blockade abrogates lung injury, inflammation, and fibrosis. The optimal time frame for GRP blockade and the key target cell types remain to be determined. The concept of GRP as a mediator of ROS-induced tissue damage represents a paradigm shift about how oxygen can cause injury, inflammation, and fibrosis. The host PNEC response in vivo may depend on individual ROS sensing mechanisms and subsequent GRP secretion. Ongoing scientific and clinical investigations promise to further clarify the molecular pathways and clinical relevance of GRP in the pathogenesis of diverse pediatric lung diseases.

Dr. Mary Ellen Avery, affectionately known as “Mel,” was a woman of courage, with a sense of humor and deep humanity. She was an intellectual powerhouse, highly creative, and productive, similar to many leaders. Yet she became the most beloved mentor for many. Here, I share some personal memories hoping to inspire others juggling life and career responsibilities. I first met Mel in 1977 when she taught our Harvard Medical School (HMS) class about respiratory distress syndrome (RDS). She held a critical audience entirely captive, listening with rapt attention. In 1979, while I was doing a medical school rotation in obstetrics at St. Thomas’s Hospital in London, Mel visited as Grand Rounds speaker. A gracious guest, she cited recent work by St. Thomas’s faculty. Always do your homework! she later said. She was also hospitable. Mel hosted Mother Teresa when the sister received an honorary degree at Harvard in 1982. Later, at Children’s Hospital (CHMC), she hosted Mildred Stahlman, who was the first to ventilate a baby. Originally competitors, Mel and Millie had become good friends. In 1980, Mel spoke to the women MD-PhD students at HMS, recalling her father’s loving support. She shared some personal challenges, especially convalescing from TB during internship. “I had a lot of time to think about the lung – and I did!” she laughed, closing with: “don’t simply do well in your field. Create a new field. For example, we could really use a field of Information Technology!” Mel often had innovative ideas (1). Starting my laboratory at Brigham and Women’s Hospital (BWH), seeking to learn methods of studying lung development, I collaborated with John Torday, whom Mel had brought from McGill to Harvard. When I asked for Mel’s feedback on our first manuscript (2), suggesting a meeting in her office, she replied, “Absolutely not! We’re going out to lunch!” Thus began a great friendship. When I first presented at an American Thoracic Society meeting, Mel sat beside me, commenting, “Interesting data, but you have to change your slides from black-on-white, which doesn’t project well. Use yellow-on-blue.” Very constructive – very Mel! Writing a grant with Jackie Coalson about bronchopulmonary dysplasia (BPD), I called Mel. At lunch, she patted my arm, “Mary, BPD isn’t a problem anymore.” By the 1990s, with surfactant therapy for RDS, BPD had become milder than originally described (3). Regardless, Mel knew BPD remained a challenge, and even collaborated with Jackie herself (4). Mel encouraged countless young professionals, especially women, but also men. Her magic stemmed from unconditional faith in others. Lewis First, then Assistant Professor and previously Mel’s intern, was asked by Mel to co-edit a new Pediatrics textbook with her (5). Lewis is now a Pediatrics Chairman (6). Mel thought highly of Mary Williams from Boston University, who had done a sabbatical in CHMC Neonatology, and often sought Mary’s valuable critique. When Stella Kourembanas became a Neonatology SCCOR Program Director at CHMC, Mel was delighted and attended monthly meetings, closely following our progress with interest. During those years, it seemed that Mel was passing the baton to Stella, one of her closest protegés. Now, Stella is Director of Newborn Medicine at CHMC. Mel’s enthusiasm was infectious. Once she asked me to tour the new Beth Israel NICU with her. The facilities and care were impressive, yet the highlight was one infant who would only drink mother’s milk, but not from a bottle – so they tried feeding her milk from a cup. Breakthrough! Mel was excited because many developing countries have insufficient bottles. This simple success could save lives. Another day, Mel whisked me to lunch with another pediatrician who was returning to clinical work after years of disability. By dessert, we were energized, embracing the future with Mel. When someone asked Mel how she felt about not having any children, Mel smiled, “What do you mean? I’ve had thousands of them!” There were no problems, only challenges; no regrets, only opportunities. Working with UNICEF deepened her awareness of global health needs, often simple yet unattainable. In India, she saw three babies in one NICU incubator and asked if they were triplets. No, they said, it was their only incubator. There weren’t sufficient resources to save all babies. She spread the word. Mel rejoiced when her trainees had children. When I was 8 months pregnant with my third child, I invited Mel for dinner at our home. She brought Maine blueberry jam, happy to meet my family. My 2-year-old son delighted her by ooh-ing and ahh-ing over the dessert, hoping to skip dinner. She later sent a warm thank you note, including 20 unusual baby names she’d collected from the NICU. In 1994, Mel was elected into the National Academy of Sciences for her discovery that newborns require surfactant to breathe (7, 8). Her comment? “Imagine that! I never even published in that journal!” Later she ran for AAAS President (9) opposite an engineer whose essay detailed his leadership experience. Mel’s essay simply addressed many challenges facing science: needs for improved rice production, vaccinations, clean water, recognizing children as our most precious resource. Mel won the election. Mel was nobody’s fool. She would tell her new ideas to over 14 people so everyone knew the ideas were hers; then she would publish quickly. She was neither offensive nor defensive. In Japan (10), Mel said: “I close with a quotation attributed to a famous German pathologist in the last century (Virchow): all new knowledge goes through three phases: (1) it is ignored; (2) it evokes hostility; (3) haven’t we known this all along?” Mel lived a full life. She often traveled with a companion. Her niece, Sue, wrote: “everyone in our family had trips with Mel including my folks. Moreover, she often took friends/colleagues with her on trips to various parts of the world.” CHMC Neonatology held a surprise 70th birthday party for her, with ~30 people telling Mel stories. When she received the Howland Award, Pediatric’s greatest award, her previous intern, Margaret Hostetter presented it (6). Her post-award celebration included Lewis First’s musical rendition of “Mel,” which she enjoyed singing at home. She was delighted to stop and smell the roses. After moving to Duke in 2004, I visited Mel every year around Thanksgiving. Once, I drove her to pick up a complete turkey dinner for her family. Mel had never learned to cook, but that never stopped her from having a party. Another time at her home, Mel showed me her nametag collection from the meetings she had attended. In 2007, Mel was animated about the BBC coming to interview her. Then, Mel became increasingly forgetful and stopped coming to CHMC. She had live-in nursing care, thanks to her family: Sue, Bill, Jennifer, and Carl Smith. At home, Mel held tightly to her biography, written by Bojan Jennings, her chemistry professor from Wheaton College (11). She was clinging to memories, precious jewels slipping away. The last time I saw Mel was 5 days before she died, at a nursing home near her childhood home. Sue and Bill had filled her room with her awards, nametags, and family photographs: a lifetime of love and accomplishment. At that moment, all I did was sit beside her, holding her hand. She was awake and comfortable, gazing into the distance and speaking in an unknown language to someone only she could see. On December 4, 2011, with Sue and Bill beside her, Mel became a free spirit. Her funeral was at the church she’d attended as a child. Like Mel, it was unpretentious and heartfelt, with family and a few friends Linda van Marter and I went together. Fred Lovejoy extolled her influence at CHMC. Sue spoke a universe of love. Bill introduced Mel as The Personal Physician for the Smith Family: “take 2 aspirins and call me in the morning.” How marvelous that Mel’s family has the same sense of humor! Returning home from Mel’s funeral, I gazed out at our brown December garden and a rose bush that had been dead for over a year, surprised to discover – a piece of paper? No, it was a perfect full-blown pink rose (Figure 1). “Mel did it!” I thought. She could move mountains – of course it was Mel! That single rose lasted over 3 weeks in winter weather. Mel, you were right: we only have to do one thing well. Love is the key: love of scientific discovery and humanity. Giving everything to help children, Mel transcended departmental expectations by founding neonatology. She transcended academics by making the world her institution through UNICEF. Ultimately, she transcended time by living on in the hearts of all. Mel’s greatest legacy was her inspiration of so many to believe in themselves and what they can do to make the world a better place, symbolized by a perfect rose. Figure 1. The Rose sent by Mel. The Redouté Rose was named after Joseph-Pierre Redouté, a Belgian artist who painted roses for Marie Antoinette, then for Josephine Bonaparte, “the rarest and most beautiful plants obtainable.” He was immortalized through his timeless inspiration of others. Just like Mel. 1. Greenes RA, Shortliffe EH. Medical informatics. An emerging academic discipline and institutional priority. JAMA (1990) 263:1114–20. doi:10.1001/jama.1990.03440080092030 CrossRef Full Text 2. Sunday ME, Hua J, Dai HB, Nusrat A, Torday JS. Bombesin increases fetal lung growth and maturation in utero and in organ culture. Am J Respir Cell Mol Biol (1990) 3:199–205. doi:10.1165/ajrcmb/3.3.199 Pubmed Abstract | Pubmed Full Text | CrossRef Full Text 3. Northway WH. Bronchopulmonary dysplasia: twenty-five years later. Pediatrics (1992) 89:969–73. 4. Berger TM, Frei B, Rifai N, Avery ME, Suh J, Yoder BA, et al. Early high dose antioxidant vitamins do not prevent bronchopulmonary dysplasia in premature baboons exposed to prolonged hyperoxia: a pilot study. Pediatr Res (1998) 43:719–26. doi:10.1203/00006450-199806000-00002 Pubmed Abstract | Pubmed Full Text | CrossRef Full Text 5. Avery ME, First LR. Pediatric Medicine. Baltimore: Williams & Wilkins (1989). 6. Hostetter MK. Introduction of the American pediatric society’s 2005 John Howland award recipient, Mary Ellen Avery, M.D. Pediatr Res (2005) 58:1311–6. doi:10.1203/01.pdr.0000189312.67186.d8 CrossRef Full Text 7. Avery ME, Mead J. Surface properties in relation to atelectasis and hyaline membrane disease. AMA J Dis Child (1959) 97:517–23. 8. Avery ME. Surfactant deficiency in hyaline membrane disease: the story of discovery. Am J Respir Crit Care Med (2000) 161:1074–5. doi:10.1164/ajrccm.161.4.16142 CrossRef Full Text 9. Avery ME. Presidential address. What is good for children is good for mankind: the role of imagination in discovery. Science (2004) 306:2212–3. doi:10.1126/science.306.5705.2212 CrossRef Full Text 10. Philip AG. Historical perspectives the underpinnings of neonatal/perinatal medicine: surfactant deficiency to surfactant use. Neoreviews (2002) 3:e239–42. doi:10.1542/neo.3-11-e227 CrossRef Full Text 11. Jennings BH. Mel: A Biography of Dr. Mary Ellen Avery. North Charleston, SC: CreateSpace (2009). Keywords: twentieth century, historical perspective, neonatology, translational medical research, basic science research, mentoring Citation: Sunday ME (2014) Mel Avery: mentor, role model, friend, mother of us all. Front. Pediatr. 2:18. doi: 10.3389/fped.2014.00018 Received: 05 March 2014; Accepted: 06 March 2014; Published online: 31 March 2014. Edited and reviewed by: John Steven Torday, University of California Los Angeles, USA Copyright: © 2014 Sunday. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. *Correspondence: mary.sunday@duke.edu