Claudia Dos Santos
MD, MSc, FRCPC
Dr. dos Santos is a Scientist in the Keenan Research Center of the Li Ka Shing Knowledge Institute of St. Michael's Hospital and an Assistant Professor of Medicine, Department of Medicine, Division of Respirology and Interdepartmental Division of Critical Care at the University of Toronto.
She did her BSc Hons in Biology at Trent University, her MSc degree at the Hospital for Sick Children in Molecular and Medical Genetics where she worked with Dr. Manuel Buchwald on Fanconi’s Anemia and DNA repair and her Medical degree, Internal Medicine and Critical Care training at the University of Toronto. She continued in Toronto for her post-doc in the lab of Dr. Arthur Slutsky, a world renowned expert on mechanical ventilation and Dr. Mingyao Liu an expert on mechanotransduction.
Dr. dos Santos now holds an appointment at the Institute of Medical Sciences and Collaborative Program in Genome Biology and Bioinformatics. Her clinical specialty is Critical Care Medicine. She held a Fellowship from the Parker B. Francis Foundation, a Phase I Clinician-Scientist from the Heart and Stroke Foundation, and now holds an Early Research Award from the Ministry of Research and Innovation and a New Investigator Award from the Canadian Institutes of Health Research (CIHR).
- My laboratory focuses on developing integrated systems biology and functional genomics approaches to: understanding molecular mechanisms of acute multiple organ failure (HEART and LUNG) in the critically ill (sepsis and acute respiratory distress syndrome ARDS)
- developing an “informed” approach to the discovery of novel molecular targets for therapy including stem cell and gene therapy.
My lab is dedicated to applying novel whole throughput and computational strategies to identify - using a “candidate” gene approach - putative genes/molecules, pathways and networks involved in the pathophysiology, diagnosis, prognosis and management of critical illness: specifically I am interested in mechanisms of HEART and LUNG failure in critically ill patients.
Acute and Ventilator Induced Lung Injury –Bioinformatics was used to determine cyclic stretch is associated with a specific gene expression profile that underlies cellular remodeling in the setting of injury (dos Santos et al. CCM 2008). (ii) Response to injury is a complex, multigenic event that involves multiple factors acting at different times. I have shown that in part, the coordinated response of lung cells to cyclic injury is regulated by the stretch sensitive transcription factor activating transcription factor 3 (ATF3). To establish the biological importance of this finding, I used an ATF3 knock out mouse to demonstrate that ATF3 is a negative transcriptional regulator that acts to “counter-balance” the hyperinflammatory response (Akram et al. AJRCCM 2010 Appendix 2).
Combining microarray data using gene-gene meta-analysis has yielded a profile that has the potential to differentiate clinically relevant ALI (P. Hu et al. Plos One, 2011). We have recently developed ATF3 chimera to demonstrate the relative contribution of ATF3 expression in different cells to ALI/VILI (Shan et al. under revision, Appendix 4)
Mesenchymal Stem Cells (MSCs) in Multiorgan Injury: (iii) More recently my focus has shifted towards stem cells, which have an immunomodulatory and reparative potential. In collaboration we have shown that MSCs significantly decreased mortality in septic mice. Analysis of expression profiles from treated and untreated mice demonstrated an overall downregulation of inflammation-related genes, and a shift towards upregulation of genes involved in bacterial killing. These findings strongly suggest that the beneficial effects of MSCs extend beyond suppression of inflammation (Mei et al. AJRCCM 2010).
Global expression analysis points to a fundamental role of mitochondria related genes in conferring the beneficial effects of MSCs (dos Santos et al. Am J of Pathol, 2011 Appendix 1). Work from my lab has also shown that we can generate MSCs and that these can be used to attenuate fibroproliferative changes in the lung that occur secondary to dysregulated repair of lung injury (Maron_gutierrez et al. in press, Appendix 5)
Sepsis-induced Myocardial Depression: Myocardial Depression – (iv) I compared the transcriptional profile of wild type mice sensitive to sepsis induced myocardial depression with nitric oxide synthase deficient mice that are resistant. Sepsis-induced myocardial depression results in down regulation of peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC1a), involved in mitochondrial biogenesis and substrate bioenergy utilization.
Decrease in PGC1a was not present in iNOS deficient animals (dos Santos et al. CCM 2010). (iv) Treatment of wild type septic mice with Resveratrol – an antioxidant that increases PGC1a – protected septic animals from myocardial depression, suggesting that mitochondrial “resuscitation”, via preservation of PGC1a represents a potential treatment strategy for myocardial depression (dos Santos C et al. CCM 2012 Appendix 3)
Anti-cholinergic Anti-inflammatory Reflex in Critical Illness – (v) We have shown that stimulation of the anti-inflammatory cholinergic pathway mitigates mechanical injury caused by repetitive cyclic stretch associated with ventilator-induced lung injury.
In vivo (animal) and in vitro (human cellular) models, cyclic stretch injury is exacerbated by inhibition of the alpha 7 nicotinic receptor for acetylcholine, and that stimulation of this receptor results in down regulation of critical intracellular mediators of cyclic stretch injury. Moreover, in vivo electrical or pharmacological stimulation of the efferent vagus nerve protects animals from severe lung injury by reducing both inflammation and cellular apoptosis (dos Santos et al. AJRCCM 2010). This work opens up an entirely new line of therapeutic options in the field of acute lung injury.
The Li Ka Shing Knowledge Institute (LKSKI): The Building
All of the research will be conducted in my laboratory which is located at the Keenan Research Center inside the LKSKI building. The LKSKI is a 25,204-square-metre facility accommodates top-tier researchers from multidisciplinary fields. The building is designed to promote collaboration and innovation.
The Keenan Research Centre is a programmatically oriented, state of the art Basic Science lab facility working to accelerating the Translation of Research Based Discoveries to Frontline Healthcare. It includes 3 basic science floors (66,000sf) all designated as level 2 Bio-containment.
The RCF is intended to provide technical and expert support to investigators with a view to enhancing the productivity, interaction and collaboration between scientists, their staff and students. This self-financed business provides the very latest equipment and techniques in addition to professional training and management for basic scientists at St. Michael’s Hospital. As part of the research infrastructure of the LKSKI there are three research coordinators, one for each floor. The coordinators’ provide:
- technical expertise
- maintenance of equipment.
Research Core Facility:
Existing within the wet bench floors (26,000sf). Shared facilities includes both common and specialized resources such as: Molecular Biology, and Histology.
The BioIMAGING Centre provides state of the art imaging support including veterinary imaging (CT scans, US and MIRs). Vivarium (leading edge design occupies 1 floor [22,000sf]). Large and small animal housing. OR Suites for large and small animal surgery. Suites for: transgenic barrier and quarantine. We also have a large Aquatics Facility (Zebrafish Centre for Advanced Drug Discovery), and Neurobehavioral Testing.
Analytical room: Fume hood, Millipore water system, acids and flammables storage units, ultracentrifuges, high speed centrifuges, plate readers (absorbance, fluorescence and luminescence) spectrophotometers, Nanodrop nano-spectrophotometers (for quantitation of small quantities of nucleic acid and protein), and 2D SDS-PAGE system (4th floor only), Gentle MACS tissue dissociator and sonicators (4th floor only) and an analytical balance (2ug lower limit; 4th floor only).
Radioisotope Facilities (for both high and low level radiation)
Biobank: Liquid nitrogen and biobank.
Flow Cytometry: BD FACS Calibur 1 & 2 analyzers (HeNe and Argon Lasers, 4 colour capability); BD FACS Sort cytometer analyzer (Argon Laser with 3 colour capability); Miltenyi MACS Quant analyzer (Violet, blue-green and red solid state CW lasers, 7 colour capability with autoloaders for stains, 24 tube rack and 96 well plate loader) and analysis stations with FCS Express, Miltenyi MACS Quantify and FlowJo software.
Molecular Devices: SpectraMax monochromator based spectrophotometer with absorbance capabilities. BTX ECM 830 electroporator for gene delivery to cultured and primary cells
Virus lab: designed for the in-house production/packaging of lentivirus and mengovirus
BSC, 2 CO2 incubators, centrifuge, autoclave, white light microscope
Histology lab: A complete facility for the preparation and processing of tissues and cultured cells for cytological techniques such as immunohistochemistry and immunofluorescence.
Bacteria lab: Biological safety cabinet, CO2 incubators, Bacteria incubator, Shake incubator, Autoclave
Analytical lab: Waters HPLC system including Waters 1525 binary HPLC pump, 2487 Dual absorbance detector, 2475 multi fluorescence detector, 2465 electrochemical detector and Bio-Rad 2110 faction collector.
Is located on the 6th floor of the LKSKI. It includes designated areas:
- small rodent ICU. We have 3 surgical stations, and 2 mechanical ventilators and equipment for monitoring and resuscitating. We house our animals in the vivarium and for more chronic models post-ops are conducted in the vivarium.
- Wet bench facilities. We have a separate wet bench area for protein, DNA and RNA work where we conduct the bulk of the molecular biology work.
- Cell culture. We currently work in 2 separate cell culture rooms on the same floor as our lab – this allows us to separate cell line from primary cell work. Containement facilities, bacterial, virus and other biohazard work is conducted in separate tissue culture rooms.
- Dry bench. We have a 3 terminal dry bench laboratory that allows us to perform the computational work. A lot of the bioinformatic experiments are also conducted in the computational facilities of the Center for Applied Genomics where we have access to critical large data management and bioinformatics software. We also have access to the Center for Phenogenomics (TCP) of the University of Toronto. The TCP is an innovative, scientific collaboration between four research hospitals, including St. Mike’s, to operate a centralized, state-of-the-art research-enabling mouse facility. The TCP conducts and supports genetic research involving generation of mutant mice, physiological phenotyping, behavioural analysis, imaging, pathology and cryopreservation for storage and distribution.
Effects of mesenchymal stem cell therapy on the time course of pulmonary remodeling depend on the etiology of lung injury in mice. Maron-Gutierrez T, Silva JD, Asensi KD, Bakker-Abreu I, Shan Y, Diaz BL, Goldenberg RC, Mei SH, Stewart DJ, Morales MM, Rocco PR, Dos Santos CC. Crit Care Med. 2013 Nov;41(11):e319-33. doi: 10.1097/CCM.0b013e31828a663e.
Network analysis of transcriptional responses induced by mesenchymal stem cell treatment of experimental sepsis. dos Santos CC, Murthy S, Hu P, Shan Y, Haitsma JJ, Mei SH, Stewart DJ, Liles WC. Am J Pathol. 2012 Nov;181(5):1681-92. doi: 10.1016/j.ajpath.2012.08.009.
Salutary effect of resveratrol on sepsis-induced myocardial depression. Smeding L, Leong-Poi H, Hu P, Shan Y, Haitsma JJ, Horvath E, Furmli S, Masoom H, Kuiper JW, Slutsky AS, Parker TG, Plötz FB, dos Santos CC. Crit Care Med. 2012 Jun;40(6):1896-907. doi: 10.1097/CCM.0b013e31824e1370
Neuroimmune regulation of ventilator-induced lung injury. dos Santos CC, Shan Y, Akram A, Slutsky AS, Haitsma JJ. Am J Respir Crit Care Med. 2011 Feb 15;183(4):471-82. doi: 10.1164/rccm.201002-0314OC. Epub 2010 Sep 24. Activating transcription factor 3 confers protection against ventilator-induced lung injury.