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Harnessing the Osteogenic Capability of Neonatal Harnessing the Osteogenic Capability of Neonatal Dura to Heal Critical Sized DefectsDura to Heal Critical Sized Defects

Walter Sweeney B.S., Brendan Alleyne B.S., Adam Cash M.D., Christy Gliniak B.S., Gregory Cooper PhD., Arun Gosain M.D.

Case Western Reserve University, Department of Plastic & Reconstructive Surgery, Cleveland, Ohio

Presenter: Brendan Alleyne B.S.

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Introduction

Critical Sized Defects Defects incapable of healing in the animals lifetime

Problematic for adults and children because vital structures are unprotected

Interestingly, children < 1 year of age can heal large cranial defects

Healing is due to osteogenic potency of dura but potency decreases with age

Can the dural osteogenic potency present in children be harnessed to heal critical sized defects?

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Disclosure

Nothing to disclose

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Hypothesis

Rat neonatal dura transplanted into adult rat critical sized defects would retain the ability to induce osteogenesis and augment healing.

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Materials and Methods

24 Adult Rats24 Neonatal Rats

Craniotomy with dural harvest from parietal aspect of animals 8, 17, and 45 days PN

8 days PN (n=6) 17 days PN

(n=6)

45 days PN (n=6)

60 day oldadult rats with 2.3mm defect

60 day oldadult rats with 2.3mm defect

60 day oldAdult rats with 2.3mm defect

60 day oldAdult rats

with 2.3mm defect

Untreated

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Materials and Methods

8,17,or 45 day old postnatal rat

Craniotomy Dura

Inlayed

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AnalysisMicro-CT images performed 5 weeks post-operatively

Image J was used to assess pixel density as a surrogate for bone formation

Pixel density was compared between the cranial defect and intact cranial bone

At 5 weeks only untreated defects and defects treated with 8 day old neonatal dura were evaluated

Radiographic images performed 8 weeks post-operatively Image J was used to measure the Bone Gap

Bone gap was defined as the distance between bone formation and edge of the defect

At 8 weeks the bone gap was measured for all animals

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Results

Untreated Treated w/ 8 day old dura

Micro-CT 5 weeks

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Results

Untreated Treated w/ 8 day old dura

Radiograph 8 wks

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Results of Animals Evaluated with Micro-CT at 5 Weeks

Defects treated with 8 day old dura achieved pixel density equivalent to intact cranial bone

Pixel density of untreated defects was decreased 3 fold as compared to intact cranial bone

Increased Pixel Density = Increased Bone Formation

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Results of Animals Evaluated with Radiographs at 8 Weeks

Decreased Bone Gap = Increased Bone Formation Defects treated with 8 day old dura had the smallest bone gap and most bone formation

Bone gap was largest in untreated defects

Supporting evidence that dura becomes less osteogenic with age, the bone formation decreased as the age of the dura increased

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Conclusion

First study to demonstrate the neonatal dura retains osteogenic potency following transplantation

This study supports that the osteogenic capability of dura decreases with age

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Significance of the findings…

Dura can be harnessed to augment CSD healing

Future studies will focus on identifying growth factors responsible for cranial regeneration

Such findings will facilitate the engineering of biocompatible grafts capable of mimicking dural induced osteogenesis

Additionally, current studies aim to determine if fetal dura has an even greater degree of osteogenic potential than that of perinatal dura

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Sincere thanks to…

Dr. Gosain

Walter Sweeney

Christy Gliniak

Davood Varghai

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Harnessing the Osteogenic Capability of Neonatal Dura to Heal Critical Sized Defects

Walter Sweeney B.S., Brendan Alleyne B.S., Davood Varghai M.D., Adam Cash M.D., Christy Gliniak B.S., Gregory Cooper PhD., Arun Gosain M.D.

Case Western Reserve University, Department of Plastic & Reconstructive Surgery, Cleveland, Ohio

Introduction ConclusionsResults

Methods Future Investigation

Moreira-Gonzalez, A., Jackson, I. T., Miyawaki, T., Barakat, K. & DiNick, V. Clinical outcome in cranioplasty:critical review in long-term follow-up. J. Craniofac. Surg. 14, 144-153 (2003).

Chim, H. & Gosain, A. K. Biomaterials in craniofacial surgery: experimental studies and clinical application. J. Craniofac. Surg. 20, 29-33 (2009).

Cho, Y. R. & Gosain, A. K. Biomaterials in craniofacial reconstruction. Clin. Plast. Surg. 31, 377-85, v (2004).

David, L., Argenta, L. & Fisher, D. Hydroxyapatite cement in pediatric craniofacial reconstruction. J. Craniofac. Surg. 16, 129-133 (2005).

David, D. J. & Cooter, R. D. Craniofacial infection in 10 years of transcranial surgery. Plast. Reconstr. Surg. 80, 213-225 (1987).

Fearon, J. A. et al. Infections in craniofacial surgery: a combined report of 567 procedures from two centers. Plast. Reconstr. Surg. 100, 862-868 (1997).

Hobar, P. C., Masson, J. A., Wilson, R. & Zerwekh, J. The importance of the dura in craniofacial surgery. Plast. Reconstr. Surg. 98, 217-225 (1996).

Li, S., Quarto, N. & Longaker, M. T. Dura mater-derived FGF-2 mediates mitogenic signaling in calvarial osteoblasts. Am. J. Physiol. Cell. Physiol. 293, C1834-42 (2007).

Gosain, A. K. et al. Quantitative assessment of cranial defect healing and correlation with the expression of TGF-beta. J. Craniofac. Surg. 12, 401-404 (2001).

Analysis References

This study suggests that the in vivo osteogenic properties of dura are preserved with transplantation and can be harnessed to augment CSD healing. Future studies will focus on identifying factors responsible for cranial regeneration and engineering biocompatible grafts capable of mimicking dural induced osteogenesis. Additionally, future studies will aim at seeing if fetal dura has an even greater degree of osteogenic potential.

Increased Pixel Density : Increased Bone Formation

•Defects treated with 8 day old dura achieved pixel density equivalent to intact cranial bone• Pixel density of untreated defects was decreased 3 fold as compared to intact cranial bone

Decreased Bone Gap : Increased Bone Formation

• Defects treated with 8 day old dura had the smallest bone gap and most bone formation• Bone gap was largest in untreated defects• Supporting evidence that dura becomes less osteogenic with age, the bone formation decreased as the age of the dura increased

This is the first study to demonstrate that neonatal dura retains osteogenic potency following transplantation. This study supports that the osteogenic capability of dura decreases as the animal ages.

Craniotomies were performed and 5x5 mm of parietal dura harvested from twenty-four Sprague Dawley rats at 8, 17, and 45 days postnatal. A 2.3 mm CSD was created in the parietal bone of twenty-four 60 day old Sprague Dawley rats. Animals were randomized into four experimental groups: A. 8-day old dura inlayed into CSD; B. 17-day old dura inlayed into CSD; C. 45 day old dura inlayed into CSD; or D. Untreated CSD. Four animals treated with 8-day old dura were evaluated five weeks post-operatively using micro-CT and the remaining animals were evaluated 8 weeks post-operatively with radiographs. Micro-CT images were analyzed for pixel density within the CSD. Radiographic images were evaluated by measuring the distance between new bone formation within the CSD and the edge of the cranial defect; we defined this as the bone gap.

(1) 8,17,or 45 day old postnatal rat

(3) Isolated dura mater (4) Inlayed dura mater

(2) Isolated calvaria

itical Sized Defects

Critical Sized Defects are defects incapable of healing in the animals lifetime

These are problematic for adults and children because vital structures are left unprotected Interestingly, children < 1 year of age retain the ability to heal large cranial defects

Healing is due to the osteogenic potency of dura, however, potency decreases with age

Can the dural osteogenic potency present in children be harnessed to heal critical sized defects?

While the osteogenic properties of dura are thoroughly documented, the ability of dura to augment critical sized cranial defect (CSD) healing is less established

In the present study we evaluated whether transplanted dura retained sufficient osteogenic potential to aid in CSD repair

Untreated Treated w/ 8 day old dura

Untreated Treated w/ 8 day old dura