Assistant Research Scientist, Department of Orthopaedic Surgery and Unit for Laboratory Animal Medicine

Skeletal tissues are relatively acellular and contain an abundant collagenous and non-collagenous extracellular matrix (ECM). Traditionally, the matrix was thought to serve primarily a structural role; however, it is now recognized that the matrical elements are dynamically involved in regulating the function of associated mesenchymal cells. My research program is focused on studying the influence of the ECM on skeletal cells, and determining how the ECM modulates skeletal growth, remodeling, and healing.

Prior to arriving in Ann Arbor, I spent five years at the University of Washington studying the ECM component, thromobospondin-2 (TSP2). TSP2 is a 450 kDa multi-domain, trimeric protein expressed ubiquitously in the skeleton. I have found that TSP2 knockout mice (TSP2-null) have an increase in endosteal bone formation and that TSP2-null mice have an increase in the number of marrow stromal cells (MSC). As well, I have demonstrated that MSC show a dose-dependent decrease in proliferation when exposed to TSP2, but that a lack of TSP2 leads to a delay in osteogenesis and enhanced adipogenesis.

MSC are undifferentiated mesenchymal stem cells sequestered in the marrow. MSC proliferate and differentiate to become endosteal osteoblasts. They exist in an ECM compartment composed primarily of loose collagen fibrils and non-collagenous ECM proteins, including TSP2. In addition to becoming osteoblasts, MSC are capable of differentiating to adipocytes and chondrocytes. Control mechanisms that regulate MSC proliferation and differentiation involve an inter-play of autocrine, paracrine, and systemic factors; however, the complex interactions that occur in vivo are poorly understood. Presumably, regulatory factors are turned-on, and –off, in response to a variety of stimuli, including tissue injury, weight deprivation, and mechanical loading.

Current studies are directed at discerning two primary questions. First, how does TSP2 regulate the proliferation and differentiation of osteoprogenitors? Second, what is the importance of the multi-domain structure of TSP2 in the protein's function? The first question is being pursued primarily in vitro. Various recombinant TSP2 protein constructs are being used to identify interactions between TSP2 and other extracellular proteins and cell surface receptors. Proliferation, mineralization, and adipogenesis are being examined in cells +/- the addition of TSP2. The second main objective is being pursued using genetically engineered mice. Functional TSP2 domains are being eliminated from the TSP2 gene using novel knock-in technology (Cre-Lox), leaving the gene intact except for the elimination of the coding region for the domain of interest.

Focus areas for translational research include, (1) studying the distribution and survival of MSC when transplanted in vivo systemically or locally in various carriers, (2) studying the activation of MSC in fracture healing and determining whether modulating TSP2 levels can affect therapeutic outcomes, (3) examining the significance of MSC differentiating to adipocytes rather than osteoblasts in the development of osteoporosis, and (4) utilizing tissue engineering to modulate the ECM and enhance healing. From the perspective of tissue engineering, if a gene or protein can be delivered in conjunction with an engineered construct, and maintained locally without dissemination (such as an ECM protein), the proliferation and differentiation of endogenous mesenchymal cells that enter the delivered matrix can be modulated to promote bone, ligament, or tendon repair.

CURRICULUM VITAE

EDUCATION

1986-88 University of Illinois, College of Liberal Arts and Sciences, Champaign, IL.

1988-92 University of Illinois, College of Veterinary Medicine, Champaign, IL, BS, DVM.

1994-97 Purdue University, School of Veterinary Medicine, Department of Basic Medical Sciences, West Lafayette, IN, MS.

1997-01 University of Washington, School of Medicine, Department of Biochemistry, Seattle, WA, PhD.

POSTDOCTORAL TRAINING

1997-01 University of Washington, School of Medicine, Department of Biochemistry, Seattle, WA.

WORK EXPERIENCE

1989-92 Research Associate, University of Illinois College of Veterinary Medicine, Department of Biosciences, Champaign, IL

1992-93 Veterinary Intern, Lake Country Equine Center, Oconomowoc, WI

1993-94 Associate Veterinarian, Lake Country Equine Center, Oconomowoc, WI

1994-97 Graduate Instructor, Department of Basic Medical Sciences, School of Veterinary Medicine, Purdue University, West Lafayette, IN

1997-01 Senior Fellow, Department of Biochemistry, School of Medicine, University of Washington, Seattle WA

2002-present Assistant Research Scientist, Department of Orthopaedic Surgery and Unit for Laboratory Animal Medicine, School of Medicine, University of Michigan, Ann Arbor MI

CONSULTING POSITIONS

None

SCIENTIFIC ACTIVITIES

1998-present Ad-hoc Reviewer for Hong Kong Research Council

2000-present Ad-hoc Reviewer for journal Bone

2001-present Ad-hoc Reviewer for Journal of Bone and Mineral Research

2002 Ad-hoc Reviewer for Veterans Administration

2002 Moderator, ASBMR Annual Meeting, Concurrent Oral Session 26 (Bone, Cartilage, and Connective Tissue Matrix III)

GRANT SUPPORT

Recent

National Institute of Dental and Craniofacial Research-NIH (T32-DE07063-24), “Periodontics Training Grant”, 7/1/97-6/30/01.

Current

National Center for Research Resources-NIH Special Emphasis Research Career Award (K01-RR00161-1A), “Bone Formation in Mice Lacking Thrombospondin-2”, $480,000 (TDC), 2/15/01-2/14/06.

Biomedical Research Council-University of Michigan “Acquisition of a Fluorescence Based Real-Time Thermocycler”, $30,000 (TDC), 4/04/02. Co-investigator (SA Goldstein, PI)

National Institute of Arthritis and Musculoskletal Diseases-NIH (P30AR046024)

Regulation of MSC differentiation by Thrombospodin-2, $17,000 (TDC), 6/01/02-5/31/03

Department of Energy. “In Vivo Radionuclide Imaging of Mesenchymal Stem Cells”, Co-investigator (MR Kilbourn, PI) $1,139,31 (TDC), 8/1/02-7/31/05

Pending

National Institute of Arthritis and Musculoskeletal Diseases-NIH (R01), “Matricellular Proteins and Mesenchymal Stem Cells”, $750,000 (TDC), 10/1/02 – 9/30/07.

National Center for Research Resources-NIH Shared Instrumentation Grant “Fluorescence, Phospho-Chemiluminescence Imager”, $137,866 (TDC), 4/1/03-3/31/04

CERTIFICATION AND LICENSURE

Veterinary Licensure - Wisconsin

MILITARY SERVICE

NO

HONORS AND AWARDS

1986-90 Dole Scholar, University of Illinois

1986 Illinois State Scholar, University of Illinois

1987 Biology Honors, University of Illinois

1987-90 Brehm Scholar, University of Illinois

1988 Phi Eta Sigma, University of Illinois

1986-92 Dean's List, University of Illinois

1992 Equine Veterinary Internship, Lake Country Equine Center

1996 Physiology Summer Course, Marine Biological Laboratory

1997 Postdoctoral Fellowship, NIH-NRSA

1999 Young Investigator Award Semi-finalist, Orthopaedic Research Society

2001 Special Emphasis Research Career Award, NCRR-NIH

2002 Young Investigator Award Semi-finalist, Orthopaedic Research Society

2002 Young Investigator Award, American Society for Bone and Mineral Research

MEMBERSHIPS AND OFFICES IN PROFESSIONAL SOCIETIES

American Association of Veterinary Anatomists (AAVA)

American Society for Matrix Biology (ASMB)

American Society for Bone and Mineral Research (ASBMR)

American Veterinary Medical Association (AVMA)

International Society for Matrix Biology (ISMB)

Orthopaedic Research Society (ORS)

TEACHING ACTIVITIES

Aug. 1994 - June 1997 Graduate Assistant

Purdue University. School of Veterinary Medicine, Department of Basic Medical Sciences.

Sept. 1998 - Dec. 1998 Teaching Assistant

University of Washington. School of Medicine. Department of Biochemistry

February 2002 Guest Lecturer

Quantitative Cell Biology. BME 418. Course Director, Alan J. Hunt, Ph.D. University of Michigan.

EXTRAMURAL INVITED PRESENTATIONS

December 1999 Purdue University, Department of Basic Medical Sciences

June 2000 Chiroscience-Celltech Inc. Bothell WA

July 2000 Hope-Heart Institute. Seattle WA

June 2001 GlaxoSmithKline. King of Prussia PA

November 2002 University of Notre Dame, Center for the Study of Biocomplexity.

COMMITTEE AND ADMINISTRATIVE SERVICE

2002 University of Michigan Bone Center Education Committee

BIBLIOGRAPHY

Peer Reviewed Publications

1. Hankenson K and Schaeffer DJ (1991) Microtox Assay of Trinitrotoluene, Diaminonitrotoluene, and Dinitromethylalanine Mixtures. Bull Environ Contam Toxicol 46:550-553.
2. Schaeffer DJ, Goehner M, Grebe E, Hansen LG, Hankenson K, Herricks E, Matheus G, Miz A, Reddy R, Trommater K (1991) Evaluation of the "Reference Toxicant" Addition Procedure for Testing the Toxicity of Environmental Samples. Bull Environ Contam Toxicol 47:540-546.
3. Hankenson KD and Turek JJ (1999) Porcine anterior cruciate ligament fibroblasts are similar to cells derived from the ligamentum teres, another non-healing intra-articular ligament. Connect Tis Res 40:13-21.
4. Hankenson KD, Turek JJ, Watkins BA, Schoenlein IA (2000) Omega-3 Fatty Acids Enhance Ligament Fibroblast Collagen Formation in Association with Changes in Interleukin-6 Production. Proc Soc Exp Biol Med 223:88-95.
5. Hankenson KD, Bain SD, Kyriakides TR, Smith E, Goldstein SA, Bornstein P (2000) Increased marrow-derived osteoprogenitor cells and endosteal bone formation in mice lacking thrombospondin 2. J Bone Min Res 15: 851-862.
6. Bornstein P, Armstrong LC, Hankenson KD, Kyriakides TR, Yang Z (2000) Thrombospondin 2, a matricellular protein with diverse functions. Matrix Biology. 19:557-68.
7. Hankenson KD and Bornstein P (2002) The secreted protein thrombospondin-2 is an autocrine inhibitor of marrow stromal cell proliferation. J Bone Min Res 17: 415-425
8. Ekholm E, Hankenson KD, Uusitalo H, Hiltunen A, Gardner H, Heino J, Penttinen R (2002) Diminished callus size and cartilage synthesis in a1b1 integrin deficient mice during bone fracture healing. Am J Pathol 160: 1779-1785.
9. Armstrong LA, Bjorkblom B, Hankenson KD, Siadek A, Bornstein P (2002) Paracrine regulation of angiogenesis by fibroblasts is mediated by thrombospondin-2. Mol Biol Cell 13: 1893-1905.

Articles in Review

1. Kyriakides TR, Rojnuckarin P, Reidy MA, Hankenson KD, Papayannopoulou T, Kaushansky K, Bornstein P (2001) Megakaryocytes require thrombospondin2 for normal platelet formation and function. in press to Blood

Manuscripts in Preparation

1. Hankenson KD, Stroup G, Apone S, Blake S, Liang X, James IE, Lark M, Bornstein P (2002) Increased osteoblastogenesis and decreased bone resorption protects against ovariectomy-induced bone loss in thrombospondin-2-null mice. in preparation for Bone.
2. Hankenson KD, Gross TS, Bornstein P, Bain S, and Srinivasan S. Mice lacking the extracellular matrix protein, thrombospondin-2, show an altered response to mechanical loading of the tibia. in preparation for Bone.

Book Chapter in press

1. Hankenson KD, Cavaliere C, Frank RP. Animal models of skeletal diseases. In Handbook of Laboratory Animal Science, Vol II. Animal Models, GL Van Hoosier and J Hau, eds., CRC press, Cambridge, MA, 2002.

Abstracts

1. Hankenson KD and Turek JJ. An in vitro comparison of the porcine ligament of the head of the femur to the anterior cruciate and medial collateral ligaments. FASEB 1997. New Orleans LA (poster)
2. Hankenson KD, Watkins BA, Schoenlein IA, Allen KGD, Turek JJ. Arachidonic acid and eicosapentaenoic (EPA) acids improve medial collateral ligament fibroblast wound healing but only EPA enhances collagen formation and alters interleukin-6 production. FASEB 1999. Washington DC (poster)
3. Hankenson KD, Bain SD, Kyriakides TK, Goldstein SA, Bornstein P. Disruption of the murin thrombospondin-2 gene (THBS2) results in increased endosteal bone formation with a concomitant increase in bone density and cortical thickness. ORS 1999. Anaheim CA (NIRA poster)
4. Hankenson KD, Bain SD, Kyriakides TK, Goldstein SA, Smith EA, Bornstein P. The matricellular protein, thrombospondin-2, regulates marrow-derived osteoprogenitor proliferation. Thrombospondin 2000. Madison WI. (poster)
5. Hankenson KD, Bornstein P. Thrombospondin-2 is an inhibitor of marrow stromal cell proliferation. ORS 2001. San Francisco CA (poster)
6. Hankenson KD, Stroup G, Apone S, Blake SM, Liang X, James IE, Lark M, Bornstein P. Increased osteoblastogenesis in thrombospondin-2-null mice protects against ovariectomy-induced bone loss. ASBMR 2001. Phoenix AZ. (plenary poster)
7. Hankenson KD, Srinivasan S, Bornstein P, Gross T. Mice lacking the extracellular matrix protein, thrombospondin-2, show an altered response to mechanical loading of the tibia. ORS 2002. Dallas, TX. (NIRA poster)
8. Hankenson KD. The matricellular protein, thrombospondin-2, has opposing effects on osteogenic and adipogenic differentiation of marrow-derived mesenchymal stem cells. ASBMR 2002. San Antonio, TX. (Young Investigator Award)