Skeletal Regeneration and Repair

Craniofacial Bone Repair and Suture Biology

The craniofacial group at the Longaker lab focuses on two main areas:

1.     The first area of investigation focuses on calvarial bones of different embryonic tissue origin and how these different tissue origins impact the osteogenic potential and skeletal repair of these bones. The current research focuses on the neural crest-derived frontal bone and paraxial mesoderm-derived parietal bone. This research has unveiled significant and substantial differences between frontal and parietal bones highlighting some of the molecular mechanism(s) responsible for the different osteogenic capacity and tissue repair potential observed between the two bones. The results gained from this investigation represent an important potential for translational implications to skeletal repair/regeneration.

Several papers have been published; describing the bio-molecular mechanism(s) governing the higher osteogenic potential and skeletal repair of the neural crest-derived frontal bone and its relationship to FGF, Wnt, and BMP signaling pathways.

2.     The second area of research is the developmental biology of cranial sutures, specifically, the posterior frontal (PF) suture (metopic in humans), which is a fusing suture, and the non-closing sagittal (Sag) and coronal (Cor) sutures. We have identified the timing and process through which the PF suture closes, showing that this suture fuses through an “autonomous” endochondral ossification process between embryonic days 14-17. Furthermore, we have identified canonical Wnt signaling as a major player in controlling the patency of cranial sutures. Our research extends also to pathological conditions such as craniosynostosis, a premature closure of cranial sutures.

Current studies are aimed towards establishing the presence of and characterization of stem cell population(s), to profile their molecular signature, and to determine how these stem cells may contribute to the different patterning/fate of the cranial sutures.

Highlights

1.     Li S, Meyer NP, Quarto N, Longaker MT. Integration of multiple signaling regulates through apoptosis the differential osteogenic potential of neural crest-derived and mesoderm-derived Osteoblasts. Plos One. 2013;8(3):e58610. doi: 10.1371/journal.pone.0058610. PubMed PMID: 23536803; PubMed Central PMCID: PMC3607600.

2.     Quarto N, Wan DC, Kwan MD, Panetta NJ Origin matters: differences in embryonic tissue origin and Wnt signaling determine the osteogenic potential and healing capacity of frontal and parietal calvarial bones. J Bone Miner Res. 2010; 25 (7): 1680-94.

3.     Behr B, Panetta NJ, Longaker MT, Quarto N. Different endogenous threshold levels of Fibroblast Growth Factor-ligands determine the healing potential of frontal and parietal bones. Bone. 2010; 47 (2): 281-94.

4.     Li S, Quarto N, Longaker MT. Activation of FGF signaling mediates proliferative and osteogenic differences between neural crest derived frontal and mesoderm parietal derived bone. PLoS One. 2010; 5(11): e14033.

Skeletal Stem Cell and Microenvironment (Niche)

Injury and disease of skeletal tissue including bone and cartilage is an enormous, and growing medical burden. Our group has recently identified a self-renewing skeletal stem cell (mSSC) in mice that generates bone, cartilage and hematopoietic-supportive niche stromal cells (Cell, 2015). We then molecularly characterized the stem cell micro-environment (niche) of mSSC and identified specific signaling pathways guiding SSC expansion and differentiation towards bone or cartilage lineages. Through the lens of SSC biology, we observe that normal SSC activity is essential for normal skeletal homeostasis and regeneration (PNAS, 2015) while diminished or defective SSC activity underlies osteoporosis in aging and poor fracture healing in Type2 Diabetes Mellitus (STM, submitted). These studies define a pressing clinical need for identifying new methods to amplify SSC numbers and SSC activity in treating injuries or diseases of the skeletal system. To speed clinical translation for our findings on mSSC, we have now succeeded in isolating and purifying human skeletal stem cells (hSSC) that are the functional equivalent of mouse SSC. As we demonstrated in mice, we have also identified conditions for inducing human SSC and human bone and cartilage formation from human adipose tissue. Inducing SSC formation in situ with soluble factors and subsequently regulating the SSC niche to specify its differentiation towards bone, cartilage, or stromal cells would represent a paradigm shift in the therapeutic regeneration of skeletal tissues. Conversely, it is important to characterize the biology of SSC inducible cells and their potential role in pathological heterotropic ossification. This research seeks to address key questions regarding the identify of the SSC inducible cells present in human adipose tissue and the genetic mechanisms underlying plasticity in their cell fates and the process of re-specification into induced SSCs (iSSCs). We are also examining the biological differences between iSSCs and endogenous SSCs isolated from skeletal tissues in response to signaling that promote expansion and lineage commitment of SSC.

Highlights:

1.     Marecic O, Tevlin R, McArdle A, Seo EY, Wearda T, Duldulao C, Walmsley GG, Nguyen A, Weissman IL, Chan CK, Longaker MT. Identification and characterization of an injury-induced skeletal progenitor. Proc Natl Acad Sci U S A. 2015 Aug 11;112(32):9920-5. doi: 10.1073/pnas.1513066112. Epub 2015 Jul 27. PMID: 26216955

2.     Chan CK, Seo EY, Chen JY, Lo D, McArdle A, Sinha R, Tevlin R, Seita J, Vincent-Tompkins J, Wearda T, Lu WJ, Senarath-Yapa K, Chung MT, Marecic O, Tran M, Yan KS, Upton R, Walmsley GG, Lee AS, Sahoo D, Kuo CJ, Weissman IL, Longaker MT. Identification and specification of the mouse skeletal stem cell. Cell. 2015 Jan 15;160(1-2):285-98. doi: 10.1016/j.cell.2014.12.002.

Distraction Osteogenesis

Distraction osteogenesis (DO) is a powerful method of endogenous bone tissue engineering that has been applied to the craniofacial skeleton with great success. We have previously established a model of mandibular distraction osteogenesis in mice and current efforts are aimed to study the underlying biology governing the DO process. A more thorough understanding of the molecular and cellular responses during this process may allow for new potential strategies of bone formation that could be applied to several other clinical scenarios involving bone deficiency.

Wound Healing

Approximately six million Americans suffer from poor cutaneous wound healing (e.g. diabetic wounds, pressure wounds) annually, with 1.1 to 1.8 million new cases arising each year. It is estimated that we spend over $20 billion annually in wound care for these patients. Of patients who undergo abdominal surgery, 93% will develop abdominal adhesions, or bands of fibrous tissue that form between organs and the abdominal wall. From 1998 to 2002, 18.1% of hospitalizations were related to abdominal adhesions, resulting in an estimated cost of $1.18 billion annually. Unfortunately, there are limited means to prevent or treat these scars, adhesions, and their sequelae.

We hope to characterize and develop therapies to minimize and even prevent the development scar tissue that develops in the skin and the abdomen, thereby addressing the clinical and financial burdens of poor wound healing and scar formation.

We currently have projects in the five following areas, and collaborate with other laboratories to develop and test novel small molecule therapies, three-dimensional hydrogels to deliver cells and therapies, and disease models:

1.     The effect of tumors on cutaneous wound repair and regeneration.

2.     Pathophysiology of abdominal adhesion formation.

3.     Macrophage-mediated cutaneous wound repair and regeneration.

4.     Identification and characterization of dermal lineages critical for fibrosis and scar formation in dorsal and ventral skin.

5.     The effect of small molecule inhibitors on cutaneous wound repair and regeneration.

Highlights:

1.     Rinkevich Y, Walmsley GG, Hu MS, Maan ZN, Newman AM, Drukker M, Januszyk M, Krampitz GW, Gurtner GC, Lorenz HP, Weissman IL, Longaker MT. Skin fibrosis. Identification and isolation of a dermal lineage with intrinsic fibrogenic potential. Science. 2015 Apr 17;348(6232):aaa2151. doi: 10.1126/science.aaa2151. PMID: 25883361

2.     Walmsley GG, Hu MS, Hong WX, Maan ZN, Lorenz HP, Longaker MT. A mouse fetal skin model of scarless wound repair. J Vis Exp. 2015 Jan 16;(95):52297. doi: 10.3791/52297. PMID: 25650841

3.     Walmsley GG, Rinkevich Y, Hu MS, Montoro DT, Lo DD, McArdle A, Maan ZN, Morrison SD, Duscher D, Whittam AJ, Wong VW, Weissman IL, Gurtner GC, Longaker MT. Live fibroblast harvest reveals surface marker shift in vitro. Tissue Eng Part C Methods. 2015 Mar;21(3):314-21. doi: 10.1089/ten.TEC.2014.0118. Epub 2014 Dec 17. PMID: 25275778

Autologous Fat Grafting

In collaboration with Dr. Derrick Wan’s laboratory, the Longaker group also investigates strategies to enhance volume retention in autologous fat grafting. With long term retention rates of 10-90% reported in the literature, optimization of all steps of fat grafting - from procurement, to processing, to placement – are necessary in order to ensure reliable outcomes.

Highlights:

1.     Paik KJ, Zielins ER, Atashroo DA, Maan ZN, Duscher D, Luan A, Walmsley GG, Momeni A, Vistnes S, Gurtner GC, Longaker MT, Wan DC. Studies in Fat Grafting: Part V. Cell-Assisted Lipotransfer to Enhance Fat Graft Retention Is Dose Dependent. Plast Reconstr Surg. 2015 Jul;136(1):67-75. doi: 10.1097/PRS.0000000000001367. PMID: 25829158

2.     Garza RM, Rennert RC, Paik KJ, Atashroo D, Chung MT, Duscher D, Januszyk M, Gurtner GC, Longaker MT, Wan DC. Studies in fat grafting: Part IV. Adipose-derived stromal cell gene expression in cell-assisted lipotransfer. Plast Reconstr Surg. 2015 Apr;135(4):1045-55. doi: 10.1097/PRS.0000000000001104. PMID: 25502860

3.     Garza RM, Paik KJ, Chung MT, Duscher D, Gurtner GC, Longaker MT, Wan DC. Studies in fat grafting: Part III. Fat grafting irradiated tissue--improved skin quality and decreased fat graft retention. Plast Reconstr Surg. 2014 Aug;134(2):249-57. doi: 10.1097/PRS.0000000000000326. PMID: 25068325

4.     Atashroo D, Raphel J, Chung MT, Paik KJ, Parisi-Amon A, McArdle A, Senarath-Yapa K, Zielins ER, Tevlin R, Duldulao C, Walmsley GG, Hu MS, Momeni A, Domecus B, Rimsa JR, Greenberg L, Gurtner GC, Longaker MT, Wan DC. Studies in fat grafting: Part II. Effects of injection mechanics on material properties of fat. Plast Reconstr Surg. 2014 Jul;134(1):39-46. doi: 10.1097/PRS.0000000000000289. PMID: 25028817

5.     Chung MT, Paik KJ, Atashroo DA, Hyun JS, McArdle A, Senarath-Yapa K, Zielins ER, Tevlin R, Duldulao C, Hu MS, Walmsley GG, Parisi-Amon A, Momeni A, Rimsa JR, Commons GW, Gurtner GC, Wan DC, Longaker MT. Studies in fat grafting: Part I. Effects of injection technique on in vitro fat viability and in vivo volume retention. Plast Reconstr Surg. 2014 Jul;134(1):29-38. doi: 10.1097/PRS.0000000000000290. PMID: 24622574