Human Health Sciences Group

The Nrf2-Keap1 regulatory pathway regulates arsenic responsive genes

Known and novel arsenic responsive Nrf2 transcripts are shown. Empty circles depict unaffected target transcripts (not differentially expressed in lymphoblastoid cells). Bold and no continuous line circles indicate the level of increased gene expression as identified within this study. Bold line is 2–5 fold, no continuous line is more than 10 fold. Bold letters depict novel Nrf2-target genes identified by microarrays. PLoS One. 2014.

The Nrf2-Keap1 regulatory pathway regulates arsenic responsive genes

iCHIP: an integrated system for testing human response

The integrated in vitro chip-based human investigational platform (iCHIP) will contain human tissue from each of the body’s major functional systems. Tissues are placed in tiny experimental wells on top of the microelectrode array.

iCHIP: an integrated system for testing human response

Distribution of RAD51 foci in irs1 xrcc2 and GT619 cell lines exposed to hydroxyurea

A and C: RAD51 foci images in DAPI-stained irs1 nuclei at 4 and 20 hr, respectively, post 3-hr hydroxyurea exposure. (B and D) RAD51 foci distributions for irs1 (grey bars) GT619 (black bars) at 4 and 20 hr, respectively, post hydroxyurea exposure. At 4 hr, among 445 GT619 nuclei scored, 0.53 had zero foci; among 505 irs1 nuclei scored, 0.56 had zero foci. At 20 hr, among 424 nuclei of GT619 scored, 0.53 had zero foci; among 429 irs1 nuclei scored, 0.78 had zero foci. Uncoupling of RAD51 focus formation and cell survival after replication fork stalling in RAD51D null CHO cells. Environmental and Molecular Mutagenesis. 2012

Distribution of RAD51 foci in irs1 xrcc2 and GT619 cell lines exposed to hydroxyurea

Molecular changes in invasive PC3 cells

A, Microarray analyses of PC3 cells co-cultured with OBSostKO, OBWT+ rhDKK1, and OBWT + rhSOST as compared to monocultures; overlay representation of upregulated (green) and downregulated (red) genes. B, representative list of up-regulated transcripts. C, analysis of the effect of CRIM1 on the invasive potential of PC3 cells. D, images of Western blots of PC3 and C4-2B cell extracts following immunoprecipitation with anti-beta-catenin antibody. E, representative images of CRIM1 protein expression modulation by rhDKK1 and rhSOST; green (CRIM1), blue (DAPI).

Molecular changes in invasive PC3 cells

SOST gene expression -- effect on developing limbs

SOST is a negative regulator of bone formation, and mutations in human SOST are responsible for sclerosteosis. In addition to high bone mass, sclerosteosis patients occasionally display hand defects, suggesting that SOST may function embryonically. We found that overexpression of SOST leads to loss of posterior structures of the zeugopod and autopod by perturbing anterior–posterior and proximal–distal signaling centers in the developing limb.

SOST gene expression -- effect on developing limbs

Identifying irradiated samples

Transcript and protein panel discriminates between 2 Gy exposure and unirradiated samples, independent of inflammation stress. In comparison to untreated sham samples, inflammation in the absence of radiation exposure upregulates CDKN1A (red) and downregulates FDXR and BBC3 (green). Samples exposed to 2 Gy radiation only exhibit increased expression of all nine biomarkers, whereas subjects exposed to 2 Gy plus inflammation stress show modified induction of CDKN1A, FDXR and BBC3 and abrogation of the phosphorylation of CHK2 protein. The arrows in the radiation and inflammation combined treatment group indicate the direction of expression relative to the radiation alone group. PLoS One. 2012.

Identifying irradiated samples

TG expression of ECR5

(A) Using multiple sequence alignment, a 255-bp element, ECR5, was identified 62 kb downstream of Sost transcriptional start site. (B) Two TG constructs were used to generate TG mice, ECR5L and 3XECR5. (C–F) ECR5L expressed LacZ in the entire mouse neonate skeleton. Here we show representative images of forelimbs (C), hindlimbs (D), ribs (E), and head (F). Lower expression was observed in the calvaria (ca) relative to the mandible (ma). Nonskeletal tissues expressing LacZ included the thoracic vasculature (red arrow) (E) and the kidney (G). At higher resolution, in the calvaria, ECR5L (H), 3XECR5 (I), and LacZ expressed from the endogenous mouse Sost locus (J), marked the osteocytes. PNAS. 2012

TG expression of ECR5

Human Health Sciences Group

The Human Health Sciences Group conducts basic science and applied research on the mechanisms of action of the effects of chemicals and drugs in humans, how gene expression is regulated, and bone metabolism and fracture repair. We also focus on understanding the damage caused by radiation exposure, developing new technology for biosurveillance of outbreaks of infectious diseases, and accelerating the development of medical countermeasures. Our studies help us to understand how people respond to drugs and chemicals, how they vary in their response, and how to prevent deleterious effects.

Publications

DNA isolation and sample preparation for quantification of adduct levels by accelerator mass spectrometry. Dingley KH, Ubick EA, Vogel JS, Ognibene TJ, Malfatti MA, Kulp K, Haack KW. Methods Mol Biol. 2014;1105:147-57. doi: 10.1007/978-1-62703-739-6_12.

Evaluation of nanolipoprotein particles (NLPs) as an in vivo delivery platform. Fischer NO, Weilhammer DR, Dunkle A, Thomas C, Hwang M, Corzett M, Lychak C, Mayer W, Urbin S, Collette N, Chiun Chang J, Loots GG, Rasley A, Blanchette CD. PLoS One. 2014;9(3):e93342. doi: 10.1371/journal.pone.0093342.

Nanocomposite Scaffold for Chondrocyte Growth and Cartilage Tissue Engineering: Effects of Carbon Nanotube Surface Functionalization. Chahine NO, Collette NM, Thomas CB, Genetos DC, Loots GG. Tissue Eng Part A. 2014 May 20. [Epub ahead of print]

Quantifying interactions of a membrane protein embedded in a lipid nanodisc using fluorescence correlation spectroscopy. Ly S, Bourguet F, Fischer NO, Lau EY, Coleman MA, Laurence TA. Biophys J. 2014 Jan 21;106(2):L05-8. doi: 10.1016/j.bpj.2013.12.014.

The NRF2-KEAP1 pathway is an early responsive gene network in arsenic exposed lymphoblastoid cells. Córdova EJ, Martínez-Hernández A, Uribe-Figueroa L, Centeno F, Morales-Marín M, Koneru H, Coleman MA, Orozco L. PLoS One. 2014 Feb 7;9(2):e88069. doi: 10.1371/journal.pone.0088069. eCollection 2014.

Controlling the diameter, monodispersity, and solubility of ApoA1 nanolipoprotein particles using telodendrimer chemistry. He W, Luo J, Bourguet F, Xing L, Yi SK, Gao T, Blanchette C, Henderson PT, Kuhn E, Malfatti M, Murphy WJ, Cheng RH, Lam KS, Coleman MA. Protein Sci. 2013 Aug;22(8):1078-86. doi: 10.1002/pro.2292. Epub 2013 Jun 27.

D-Lactate production as a function of glucose metabolism in Saccharomyces cerevisiae. Stewart BJ, Navid A, Kulp KS, Knaack JL, Bench G. Yeast. 2013 Feb;30(2):81-91. doi: 10.1002/yea.2942. Epub 2013 Jan 30.

Directly coupled high-performance liquid chromatography-accelerator mass spectrometry measurement of chemically modified protein and peptides. Thomas AT, Stewart BJ, Ognibene TJ, Turteltaub KW, Bench G. Anal Chem. 2013, 85(7):3644-50. doi: 10.1021/ac303609n.

How genomics is changing our view of cancer. Loots GG. Brief Funct Genomics. 2013 Sep;12(5):389-90. doi: 10.1093/bfgp/elt035. No abstract available.

Interrogating transcriptional regulatory sequences in Tol2-mediated Xenopus transgenics. Loots GG, Bergmann A, Hum NR, Oldenburg CE, Wills AE, Hu N, Ovcharenko I, Harland RM. PLoS One. 2013 Jul 16;8(7):e68548. doi: 10.1371/journal.pone.0068548. Print 2013.

Nanosensor dosimetry of mouse blood proteins after exposure to ionizing radiation. Kim D, Marchetti F, Chen Z, Zaric S, Wilson RJ, Hall DA, Gaster RS, Lee JR, Wang J, Osterfeld SJ, Yu H, White RM, Blakely WF, Peterson LE, Bhatnagar S, Mannion B, Tseng S, Roth K, Coleman M, Snijders AM, Wyrobek AJ, Wang SX. Sci Rep. 2013;3:2234. doi: 10.1038/srep02234.

Prostate cancer invasion and metastasis: insights from mining genomic data. Hudson BD, Kulp KS, Loots GG. Brief Funct Genomics. 2013 Sep;12(5):397-410. doi: 10.1093/bfgp/elt021. Epub 2013 Jul 22. Review.

Sost and its paralog Sostdc1 coordinate digit number in a Gli3-dependent manner. Collette NM, Yee CS, Murugesh D, Sebastian A, Taher L, Gale NW, Economides AN, Harland RM, Loots GG. Dev Biol. 2013 Nov 1;383(1):90-105. doi: 10.1016/j.ydbio.2013.08.015. Epub 2013 Aug 29.

The use of nanolipoprotein particles to enhance the immunostimulatory properties of innate immune agonists against lethal influenza challenge. Weilhammer DR, Blanchette CD, Fischer NO, Alam S, Loots GG, Corzett M, Thomas C, Lychak C, Dunkle AD, Ruitenberg JJ, Ghanekar SA, Sant AJ, Rasley A. Biomaterials. 2013;34(38):10305-18. doi: 10.1016/j.biomaterials.2013.09.038.

The use of nanolipoprotein particles to enhance the immunostimulatory properties of innate immune agonists against lethal influenza challenge. Weilhammer DR, Blanchette CD, Fischer NO, Alam S, Loots GG, Corzett M, Thomas C, Lychak C, Dunkle AD, Ruitenberg JJ, Ghanekar SA, Sant AJ, Rasley A. Biomaterials. 2013 Dec;34(38):10305-18. doi: 10.1016/j.biomaterials.2013.09.038. Epub 2013 Sep 27.

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