Molecular and Cell Biologist - Scientist
Resume:
Daniel O. Espinoza, PhD
***************@*****.***
www.linkedin.com/in/espinozadaniel
Summary of Qualifications
. Broad background of knowledge and techniques, especially strong in
cell and molecular biology.
. Working on biomedical research for more than 15 years.
. Post-doctoral training at Harvard University.
. Industrial and academic work experience. Work in BSL1 and BSL2
environments, Good Manufacturing Practices (GMP) and Standard
Operating Procedures (SOP).
Grants / Awards
. Award "Carlos Monge Medrano" from the National Council of Science and
Technology of Peru - 1989.
. PhD fellowship from the New York State Office of Mental Retardation and
Developmental Disabilities, NY - 1994 - 1998.
. Postdoctoral fellowship from the National Instiutes of Health - 2004-
2006.
. Travel award - Endocrine Society's 89th Annual Meeting - Toronto, Jun 2-
5, 2007.
. Grant from the Portuguese Foundation for Science and Technology (FCT) to
study: "The molecular mechanisms of heart failure: the role of the
adipocyte as an endocrine organ" - 2010-2013.
Positions / Work Experience
Oct 2012 -Dec 2012: Visiting Scientist, Joslin Diabetes Center / Harvard
Medical School.
. Research on cell and molecular mechanisms of type 2 diabetes and
obesity.
. Genotyping and characterization of a transgenic mouse model of
diabetes.
. Supervised technician and graduate student.
. Techniques used include: culture of mouse and rat myoblasts, DNA, RNA
and protein isolation from cell cultures and mouse tissues (muscle,
liver, heart), RNA isolation from human muscle, primer design, mRNA
expression analysis, PCR, RT-PCR, DNA manipulation, restriction
analysis, electrophoresis, bacteria electroporation, prepare samples
for DNA sequencing, Western blotting, and mouse genotyping by
multiplex PCR, among others.
Jan 2010 - Date: Co-Principal Investigator - Center for Neuroscience and
Cell Biology, University of Coimbra, Portugal
. Research on cell and molecular mechanisms of type 2 diabetes and its
complications.
. Currently studying the interaction between cardiac muscle and cardiac
fat, at the molecular level, in human patients with and without
diabetes.
. Currently analyzing the expression levels of different genes involved
in inflammation, lipid and carbohydrate metabolism.
. Supervise technician and graduate students.
. Gave lectures on execise and obesity to graduate students.
. Techniques used include: DNA, RNA and protein isolation, primer
design, PCR, RT-PCR, Western blotting, cell signaling analysis among
others. In vivo studies include work with mouse and rat models of
diabetes, obesity and wound healing, tissue collection for microscopy
or protein/DNA/RNA analysis. Use of radioactive isotopes for metabolic
studies (14C-glucose).
Jul 2006 - Mar 2009: Postdoctoral fellow Joslin Diabetes Center / Beth
Israel Deaconess Medical Center / Harvard Medical School.
. Research on cell and molecular mechanisms of type 2 diabetes, obesity
and apetite control using in vitro and in vivo model systems.
. Studied the effects of bone morphogenic factors on brown adipocite
differentiation.
. Studied apetite control by neuropeptides.
. Techniques used include: tissue culture (culture of human primary pre-
adipocytes, human and mouse marrow stromal cells, and stablished cell
lines), DNA, RNA and protein isolation, cloning and subcloning of DNA
fragments, cDNA / protein overexpression, primer design, DNA
sequencing, biotinylation of DNA oligos, PCR, RT-PCR (Sybr Green,
Taqman, multiplex PCR), EMSA (electrophoresis mobilitiy shift assay),
Western blots, dual luciferase reporter assays, making of mammalian
expression vectors, lentivirus vectors, making of stables cell lines,
cell signaling, etc. In vivo studies include work with mouse models of
diabetes, obesity and colitis, drug administration, glucose and
insulin tolerance tests, tissue collection for microscopy or
protein/DNA/RNA analysis, and designing of DNA cassette to make
transgenic mice. Work with mice and hamsters.
Apr 2003 -Apr 2006: Postdoctoral fellow, Joslin Diabetes Center / Harvard
Medical School.
. Research on cell and molecular mechanisms of type 2 diabetes and
obesity.
. Characterized the effects of transcription co-activator PGC-1? on
lipid and carbohydrate metabolism.
. Found that PGC-1? upregulates both lipid synthesis and degradation
pathways.
. Techniques used include: Mammalian cell culture (myoblasts and
preadipocytes), cell metabolism assays (lipid and carbohydrate
metabolism), cell signaling networks, genomic and plasmid DNA and RNA
isolation, cloning and subcloning of DNA fragments, primer design, DNA
sequencing, PCR, RT-PCR (Sybr Green and Taqman), making of adenovirus
vectors, cDNA overexpression, expression knockdown by siRNA, basic
protein chemistry, immunoprecipitations, SDS-PAGE, Western blots;
microscopy (light, fluorescence, confocal), glucose and insulin
tolerance test, genomics and metabolomics. In vivo work: insulin and
glucose tolerance test in mice, blood and tissue collection for
protein, DNA and RNA analysis. Safe use of radioactive isotopes for
metabolic studies (14C, 3H, 32P).
Mar 2001 - Mar 2003: Postdoctoral Fellow - Center for Neurologic Diseases,
Brigham and Women's Hospital / Harvard Medical School.
. Research on cellular and molecular mechanisms of Alzheimer's disease.
. Used yeast screening system to identify and characterize human
Alzheimer's APP proteases candidates.
. Developed a mammalian cell system in HEK293 cells to screen and
characterize human Alzheimer's APP proteases candidates.
. Use of a variety of biochemical, cell and molecular biology
techniques, including: mammalian cell culture, bacteria and yeast
culture, DNA and RNA isolation, cloning and subcloning of DNA
fragments, primer design, DNA sequencing, DNA mutagenesis, PCR, cDNA
overexpression in mammalian cell, yeast and bacteria, basic protein
chemistry, immunoprecipitations, SDS-PAGE, Western blots; microscopy,
protein expression and characterization, making of stable cell lines,
basic zebra fish care, etc.
1994 - 2001: PhD Student, Biology, City University of New York, New York,
NY.
. Research on cell and molecular mechanisms of Alzheimer's disease.
. Developed a yeast in vivo screening system to identify and
characterize human Alzheimer's APP protease candidates.
. Department of Molecular Biology - New York State for Basic Reasearch
on Developmental Disabilities (Staten Island, NY). Research on
Alzheimer's disease. Developed a yeast system to screen for APP
proteases. Use of a variety of biochemical, cell and molecular biology
techniques, including: mammalian cell culture, bacteria and yeast
culture, genomic and plasmid DNA and RNA isolation, cloning and
subcloning of DNA fragments, primer design, DNA sequencing, DNA
mutagenesis, PCR, cDNA overexpression in mammalian cell, yeast and
bacteria, basic protein chemistry, immunoprecipitations, SDS-PAGE,
Western blots; protein expression and characterization, making of
stable cell lines, yeast genetics, protein expression and
characterization, affinity chromatography purification of HIS-tag
proteins, FPLC, HPLC, confocal microscopy, making of stable cell
lines, use of biomek robot liquid handler, etc.
1991 - 1994: Research Associate IV, Dept. of Microbiology and Molecular
Genetics, University of California, Los Angeles, CA.
. Research on molecular engineering of antibody molecules,
. Cloned different immunoglobulin variable regions
. Carried out DNA mutations and modification of different regions of
antibody molecules to increase affinity, target different tissues,
target cancer cells, etc.
. Techniques used include: mammalian cell and bacteria culture, genomic
and plasmid DNA isolation, RNA purification (from cell cultures or
blood), cloning and subcloning of antibody molecules, primer design,
DNA sequencing (using 35S), PCR, plasmid purification by cesium
gradients, protein purification and characterization by FPLC and HPLC,
protein concentration, enzymatic analysis, cell survival, etc. Safe
use of radiative isotopes (35S, 125I, 32P).
1990 - 1991: Research Assistant II, Section of Exploratory Research,
Genelabs Inc, Redwood City, CA.
. Screening and characterization of compounds with anti-cancer and anti-
viral properties.
. Characterization of new cytokines in mammalian cell lines.
. Sequenced DNA from alpha and beta globin chains of the northern
elephant seal, Mirounga angustirostris, (GenBank Acc. #: M73996 and
M73997).
. Techniques used include: mammalian cell and bacteria culture, DNA
isolation, RNA purification (from cell cultures or blood), cloning and
subcloning of antibody molecules, primer design, DNA sequencing (using
35S), PCR, plasmid purification by cesium gradients, protein
purification and characterization by affinity chromatography, FPLC and
HPLC, protein concentration, enzymatic analysis, cell survival, etc.
1988 - 1989: Fellowship from the Peruvian National Council of Science and
Technology.
1988 - 1989: Graduate Student, Physiology, Universidad Peruana Cayetano
Heredia, Lima - Peru.
. Research on adaptation of animals to high altitude using mice,
chickens and coots as model systems.
. Focus on respiratory physiology parameters.
. Found a DNA mutation in globin beta chain of chickens that lived for
several generations in the highlands of Peru (Beta chain thr 69 -->
ser; GenBank acc. #: M73995).
1981 - 1988: Student, Biology, Universidad Peruana Cayetano Heredia, Lima -
Peru.
. Research on the thermoregulatory behavior of sea lions,
. Research on the effects of cold and hypoxia on oxygen radical
production (using rat as a model system)
. Research on adaptation of animals to high altitude using mice,
chickens and coots as model systems.
Education
BS in Biology, Universidad Peruana Cayetano Heredia, 1988.
MPhil, City University of New York, 1998
Ph.D. in Biology, City University of New York, 2001
Summary of Skills
Molecular Biology
More than 20 years experience working with molecular biology techniques,
including: DNA and RNA isolation from different sources (tissue, culture
cells, yeast, bacteria), Southern blots, Northern blots, PCR, RT-PCR (Sybr
Green and Taqman - 96 and 384 wells), screening of genomic and cDNA
libraries, restriction analysis, DNA sequencing, sequence analysis, cloning
and expression of human and yeast genes, cDNA overexpression, expression
knockdown by siRNA, DNA mutagenesis, manipulation of plasmids, making of
adenovirus and lentivirus vectors, viral transduction, gel electrophoresis,
DNA oligo labeling (32P and biotin), making and characterization of
transgenic mouse models, use of DNA / RNA / protein analysis software and
databases (Vector NTI, DNA Star, BLAST and other NCBI databases, among
others), etc.
Cell Biology
More than 20 years experience doing cell culture of mammalian cell lines
(including: HK293, 3T3-L1, CHO, HELA, COS7, muscle cells, and colonocites
among others), culture of primary human and mouse cells (including
myoblasts, pre-adipocytes, and marrow stromal cells), transient and stable
expression of recombinant proteins, transfections, proliferation assays,
cell fractionation, protein localization, immunocytochemistry, organelle
staining, cell counting, fluorescent and confocal microscopy, etc.
Microbiology
More than 15 years experience working with bacteria (E. coli) and almost 10
years working with yeast (Saccharomyces cerevisiae), growth in solid and
liquid selective media, transformation (calcium chloride and
electroporation) with foreign DNA, protein expression and characterization,
and mutagenesis, among others.
Biochemistry
More than 15 years working on protein expression and characterization using
mammalian, yeast and bacterial systems, Western blotting,
immunoprecipitation, phosphatase assays, immunocytochemistry, ELISA, SDS-
PAGE, EMSA (electrophoresis mobilitiy shift assay, gel shift assay), dual
luciferase reporter assays, triglyceride determination from plasma and
tissues, lyophilization, etc. Some experience with protein purification,
affinity chromatography, FPLC and HPLC systems, enzymatic assays, etc.
Computer skills
Proficient knowledge of MS Word, Excel, PowerPoint, Adobe Photoshop, Adobe
illustrator, DNA analysis software (Vector NTI, DNA Star, BLAST), and
database searches, among others. Some knowledge of computer programming
languages (BASIC and Pascal) and use of PeopleSoft.
Animal / in vivo experience
Many years of experience working with animals, including mice, rats,
rabbits, hamsters, birds, and sea lions. Laboratory animal husbandry,
breeding, basic surgery, perfusion, bleeding, and tissue collection for
RNA/DNA/protein extraction. Work with mouse models of diabetes, obesity and
colitis. Kwnoledge of basic anatomy and physiology of laboratory animals,
drug administration by multiple routes (subcutaneous, intravenous, oral
gavage, intra-peritoneal, intramuscular), measure of basic blood parameters
(hematocryte, glucose, blood cell count), insulin and glucose tolerance
tests, adipocite isolation, etc. Implantation of osmotic pumps in rats.
Some experience making transgenic mice (making of fat and muscle tissue
specific expression cassettes and genotyping).
Respiratory Physiology
Making of gas mixtures, determining O2 and CO2 content in gas mixtures,
measure of blood gases, pH, hemoglobin concentration and hemoglobin oxygen
affinity.
Use of radioactivity/radioisotopes
Work safely with radioactive isotopes for almost 15 years (including 3H,
14C, 35S, and 32P) for metabolic studies of lipids, carbohydrates,
proteins, DNA sequencing and making of DNA probes.
Assay development
Developed a yeast (Saccharomyces cerevisiae) system to screen for protease
candidates involved in Alzheimer's disease. System is based on the cleavage
of a phosphatase fusion protein and the release of the phosphatase to the
conditioned medium if protease is present. Phosphatase activity was
detected by colorimetric assay in the presence of a substrate.
Use of highthroughput equipment
Use of Biomek robot liquid handler and multiplex plate readers. Use of ABI
7900 for quantitative PCR.
Languages
Fluent in written and spoken English and Spanish. Basic French and
Portuguese.
Other
Work in BSL1 and BSL2 environments, Good Manufacturing Practices (GMP) and
Standard Operating Procedures (SOP).
Publications / Abstracts
Townsend KL, Suzuki R, Huang TL, Jing E, Schulz TJ, Lee K, Taniguchi CM,
Espinoza DO, McDougall LE, Zhang H, He TC, Kokkotou E, Tseng YH. 2012. Bone
morphogenetic protein 7 (BMP7) reverses obesity and regulates appetite
through a central mTOR pathway. FASEB J. 26(5):2187-96
Cypess AM, Zhang H, Schulz TJ, Huang TL, Espinoza DO, Kristiansen K,
Unterman TG, Tseng YH. 2011. Insulin/IGF-I regulation of necdin and brown
adipocyte differentiation via CREB- and FoxO1-associated pathways.
Endocrinology 152(10):3680-9.
Macotela Y, Emanuelli B, B ng AM, Espinoza DO, Boucher J, Beebe K, Gall W,
Kahn CR. 2011. Dietary leucine - an environmental modifier of insulin
resistance acting on multiple levels of metabolism. PLoS One. 6(6):e21187.
Espinoza DO, Boros L, Crunkhorn S, Gami H, and Patti ME. 2010. Dual
modulation of both lipid oxidation and synthesis by PGC-1( and ( in
cultured myotubes. FASEB J. 24(4):1003-14.
Zhang H, Schulz TJ, Espinoza DO, Huang TL, Emanuelli B, Kristiansen K,
Tseng YH. 2010. Cross talk between insulin and bone morphogenetic protein
signaling systems in brown adipogenesis. Mol Cell Biol. 30(17):4224-33.
Kuizon S, DiMaiuta K, Walus M, Jenkins EC, Kuizon M, Kida E, Golabek AA,
Espinoza DO, Pullarkat RK, and Junaid MA. 2010. A critical Tryptophan and
Ca2+ in activation and catalysis of TPPI, the enzyme deficient in classic
late-infantile neuronal ceroid lipofuscinosis. PLoS One. 5(8):e11929.
Kokkotou E, Espinoza DO, Torres D, Karagiannides I, Kosteletos S, Savidge
T, O'Brien M, and Pothoulakis C. 2009. Melanin-Concentrating Hormone (MCH)
modulates C.difficile toxin A-mediated enteritis in mice. Gut 58(1):34-40.
Tseng YH, Kokkotou E, Huang TL, Taniguchi CM, Tran TT, Suzuki R, Schulz TJ,
Espinoza DO, Winnay JN, Yamamoto Y, Ahrens MJ, Dudley AT, Norris AW,
Kulkarni RN, and Kahn CR. 2008. Novel role of bone morphogenetic protein-7
in brown adipogenesis and energy expenditure. Nature 454(7207):1000-4.
Kokkotou E, Moss AC, Michos A, Espinoza D, Cloud JW, Mustafa N, O'Brien M,
Pothoulakis C, Kelly CP. 2008 Efficacy of rifaximin vs. vancomycin for
treatment of Clostridium difficile-associated diarrhea and prevention of
disease recurrence in hamsters. Antimicrob Agents Chemother. 52(3):1121-6.
Karagiannides I, Torres D, Tseng Y-H, Bowe1 C, Carvalho E, Espinoza D,
Pothoulakis C, and Kokkotou E. 2008. Substance P as a novel anti-obesity
target. J of Gastroenterology 134(3):747-55.
Crunkhorn S, Dearie F, Mantzoros C, Gami H, da Silva WS, Espinoza D,
Faucette R, Barry K, Bianco AC, and Patti ME. 2007. PGC-1 expression is
reduced in obesity: Potential pathogenic role of saturated fatty acids and
P38 MAP kinase activation. J Biol Chem. 282(21):15439-50.
Espinoza DO. 2001. The characterization of the secretases involved in beta-
APP processing in yeast and mammals [Dissertation]. New York, New York:
City University of New York.
Zhang W, Espinoza D, Hines V, Innis M, Mehta P, Miller DL. 1997.
Characterization of beta-amyloid peptide precursor processing by the yeast
Yap3 and Mkc7 proteases. Biochim Biophys Acta, 1359(2):110-122.
Leon-Velarde F, Mejia, O, Espinoza D, and Monge C. 1995. La hemoglobina, es
un marcador genetico invariable? Hallazgos en la gallina criolla andina
[Hemoglobin, is it an invariable genetic marker? Findings in Andean native
chicken]. Acta Andina, 4:89-94
Morrison SL, Coloma MJ, Espinoza D, Hastings A, Shin S-U, Wims L, Wright A.
1995. Vectors and approaches for the eukaryotic expression of antibodies
and antibody fusion proteins. Chapter 9 in Antibody Engineering, 2nd Ed.,
by Carl Borrebaek (editor), Oxford Univ. Press.
Carey C, Dunin-Borkowski O, Leon-Velarde F, Espinoza D, and Monge-C C.
1994. Gas exchange and blood gases of puna-teal (Anas versicolor-puna)
embryos in the Peruvian Andes. Journal of comparative physiology B-
Biochemical systematic and environmental physiology, 163(8):649-656.
Carey C, Dunin-Borkowski O, Leon-Velarde F, Espinoza D, and Monge C. 1993.
Blood gases, pH and hematology of montane and low land coot embryos.
Respiration Physiology, 93(2):151-163.
Coloma MJ, Espinoza DO, Fry KE, and Larrick JW. 1993. Antibody engineering
and immunotherapy. Chapter in Molecular Diagnostics. Ed by Rapley & Walker.
Blackwell Scientific Publications Ltd. Oxford - England.
Leon-Velarde F, Espinoza D, Monge C, DeMuizon C. 1991. A genetic response
to high altitude hypoxia - high hemoglobin-oxygen affinity in chicken
(Gallus gallus) from the Peruvian Andes. Comptes Rendus de l academie des
sciences serie III - Sciences de la vie, 313(9):401-406.
Carey C, Leon-Velarde F, Dunin-Borkowski O, Bucher TL, Gomez de la Torre G,
Espinoza D, Monge-C C. 1989. Variation in eggshell characteristics and gas
exchange of montane and low land coot eggs. Journal of comparative
physiology B-Biochemical systematic and environmental physiology,
159(4):389-400.
Abstracts:
Macotela Y, Emanuelli B, B ng A, Espinoza D, Kahn CR. 2009. Dietary Leucine
Improves Insulin Signaling And Reduces HFD-Induced Inflammation In Visceral
Fat Without Changes In Food Intake or Fat Mass. American Diabetes
Association, 69th Scientific Sessions.
Zhang H, Schulz TJ, Espinoza DO, Huang TL, Li J, Lai Y-C, Russell1 SJ,
Emanuelli B, Kristiansen K, and Tseng Y-H. 2009. Crosstalk between Insulin
and BMP Signaling Systems in Brown Adipogenesis. Diabetes Association, 69th
Scientific Sessions.
Espinoza DO, Huang TL, Giorgadze N, Tchkonia T, Kirkland JL, Tseng Y-H.
2008. Induction of Adipocyte Differentiation and Expression of Brown Fat
Markers by Bone Morphogenetic Protein-7 in Human Primary Preadipocytes.
American Diabetes Association, 68th Scientific Sessions.
Suzuki R, Kokkotou E, Jing E, Espinoza DO, Schulz TJ, Huang TL, Khan CR,
Tseng Y-H. 2008. Anorectic Effect of Bone Morphogenetic Protein-7 Mimics
Leptin's Action in the Hypothalamus at the Level of Signaling and Food
Intake. American Diabetes Association, 68th Scientific Sessions.
Schulz TJ, Espinoza DO, Cerletti M, Hettmer S, Shadrach J, Huang TL, Wagers
AJ, Tseng Y-H. Bone Morphogenetic Protein 7 Drives Brown Adipogenesis in
Muscle Resident Progenitor Cells. American Diabetes Association, 68th
Scientific Sessions.
Espinoza DO, Boros L and Patti ME. 2007. PGC1( and ( modulate lipid
metabolism in cultured myotubes. Abstract presented at The Endocrine
Society's 89th Annual Meeting.
Crunkhorn S, Dearie F, Bianco A, Kim B, Espinoza D, and Patti ME. 2007.
Thyroid hormone action is reduced with lipid exposure in C2C12 myotubes.
Abstract to be presented at the American Diabetes Association's 67th
Scientific Sessions.
Karagiannides I, Espinoza DO, Torres D, Carvalho E, Tseng Y-H, Pothoulakis
C, and Kokkotou E. 2007. Role of Substance P in feeding behavior in mice.
Presented at: NEUROPEPTIDES 2007. Functions, Dysfunctions & Thetapeutic
Options. Santorini, Greece
Espinoza DO, Patti, ME 2005. PGC1 expression alters lipid metabolism and
partitioning in skeletal muscle cells: potential links to insulin
resistance susceptibility. Abstract presented at the American Diabetes
Association's 65th Scientific Sessions. Diabetes 54(supplement 1): A360
Espinoza DO, Gami H, and Patti ME. 2004. Modulation of PGC1 Expression in
Cultured Myotubes Enhances Lipid Oxidation. Abstract presented at the
American Diabetes Association's 64th Scientific Sessions. Diabetes
53(supplement 2):A336
Espinoza DO, Miller DL, Xia W. 2001. Presenilin 1 interacts with C99 in
yeast. Abstract presented at the Annual Meeting of the Society for
Neuroscience.
Xia W, Iskandar MK, Espinoza DO, Miller DL, Zhang J and Ye W. 2001. (-
secretase can cleave the APP C-terminal fragment in a sequence-sensitive
fashion. Abstract presented at the Annual Meeting of the Society for
Neuroscience.
Espinoza DO, Potempska A, Merz G, Mack K, Mehta P, Currie JR, and Miller
DL. 2000. Roles of (-amyloid converting enzymes BACE-1 and BACE-2 in (-
amyloid secretion. Abstract presented at the World Alzheimer Congress.
Espinoza DO, Mack K, Potempska A, Currie JR, Merz G, and Miller DL. 2000.
Intracellular localization of BACE and ASP21 (BACE2) in transiently
transfected COS7 cells. Abstract presented at the Annual Meeting of the
Society for Neuroscience.
Espinoza DO, Potempska A, Tezapsidis N, and Miller DL. 1999. Expression of
Presenilin 1 in the yeast Saccharomyces cerevisiae. Abstract presented at
the Annual Meeting of the Society for Neuroscience.
Espinoza DO and Miller DL. 1998. Construction of a yeast reporter/selection
system to identify and characterize (-amyloid peptide precursor (APP)
processing enzymes. Abstract presented at the American Society for Cell
Biology Annual Meeting.
Frackowiak, J, Mazur-Kolecka, B, Carrol, RT, Espinoza DO, and Wisniewski,
HM. 1995. The effects of heavy metal ions on the metabolism of (-amyloid
precursor protein and (-amyloid in cultured smooth muscle cells. Abstract
presented at the Annual meeting of the Society for Neuroscience.
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