Effect on bone healing by the application of low intensity pulsed ultrasound after injection of adipose tissue-derived stem cells at the implantation of titanium implant in the tibia of diabetes-induced rat

Tae-Young Jung; Sang-Jun Park; Dae-Suk Hwang; Yong-Deok Kim; Soo-Woon Lee; Uk-Kyu Kim

doi:10.5125/jkaoms.2011.37.4.301

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Jung, Park, Hwang, Kim, Lee, and Kim: Effect on bone healing by the application of low intensity pulsed ultrasound after injection of adipose tissue-derived stem cells at the implantation of titanium implant in the tibia of diabetes-induced rat

Original Article

J Korean Assoc Oral Maxillofac Surg 2011;37(4):301-311.

Published online: 7 January 2011

DOI: https://doi.org/10.5125/jkaoms.2011.37.4.301

Effect on bone healing by the application of low intensity pulsed ultrasound after injection of adipose tissue-derived stem cells at the implantation of titanium implant in the tibia of diabetes-induced rat

Tae-Young Jung¹, Sang-Jun Park¹, Dae-Suk Hwang², Yong-Deok Kim², Soo-Woon Lee³, Uk-Kyu Kim²

¹Department of Oral and Maxillofacial Surgery, Inje University Pusan Paik Hospital, Busan

²Department of Oral and Maxillofacial Surgery, Pusan National University School of Dentistry, Yangsan

³Department of Oral and Maxillofacial Surgery, Inje University Haeundae Paik Hospital, Busan, Korea

^∗ This work was supported by the research grant of Inje University in 2008.

Uk-Kyu Kim Department of Oral and Maxillofacial Surgery, School of Dentistry, Pusan National University Beomeo-ri, Mulgeum-eup, Yangsan, 626-870, Korea TEL: +82-55-360-5100 FAX: +82-55-360-5104 E-mail: kuksjs@pusan.ac.kr

Received 8 March 201122 July 2011 Accepted 25 July 2011

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Introduction

This study examined the effect of the application of low intensity pulsed ultrasound on bone healing after an injection of adipose tissue-derived stem cells (ADSCs) during the implantation of a titanium implant in the tibia of diabetes-induced rats.

Materials and Methods

Twelve Sprague-Dawely rats were used. After inducing diabetes, the ADSCs were injected into the hole for the implant. Customized screw type implants, 2.0 mm in diameter and 3.5 ㎜ in length, were implanted in both the tibia of the diabetes-induced rats. After implantation, LIPUS was applied with parameters of 3 MHz, 40 mW/cm², and 10 minutes for 7 days to the left tibiae (experimental group) of the diabetes-induced rats. The right tibiae in each rat were used in the control group. At 1, 2 and 4 week rats were sacrificed, and the bone tissues of both tibia were harvested. The bone tissues of the three rats in each week were used for bone-to-implant contact (BIC) and bone area (BA) analyses and the bone tissues of one rat were used to make sagittal serial sections.

Results

In histomorphometric analyses, the BIC in the experimental and control group were respectively, 39.00±18.17% and 42.87±9.27% at 1 week, 43.74±6.83% and 32.27±6.00% at 2 weeks, and 32.62±11.02% and 47.10±9.77% at 4 weeks. The BA in experimental and control group were respectively, 37.28±3.68% and 31.90±2.84% at 1 week, 20.62±2.47% and 15.64±2.69% at 2 weeks, and 11.37±4.54% and 17.69±8.77% at 4 weeks. In immunohistochemistry analyses, Osteoprotegerin expression was strong at 1 and 2 weeks in the experimental group than the control group. Receptor activator of nuclear factor kB ligand expression showed similar staining at each week in the experimental and control group.

Conclusion

These results suggest that the application of low intensity pulsed ultrasound after an injection of adipose tissue-derived stem cells during the implantation of titanium implants in the tibia of diabetes-induced rats provided some positive effect on bone regeneration at the early stage after implantation. On the other hand, this method is unable to increase the level of osseointegration and bone regeneration of the implant in an uncontrolled diabetic patient.

Keywords: Diabetes, Implant, Low intensity pulsed ultrasound, ADSCs

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Fig. 1.

Cultivated adipose tissue-derived stem cells.

Fig. 2.

Non-decalcified section: histological aspects observed within the limits of the thread.(Villanueva stain, ×100)

Fig. 3.

Mean bone-to-implant contact (BIC) in experimental and control group.

Fig. 4.

Mean bone area (BA) in experimental and control group.

Fig. 5.

Microphotographs.(H-E stain, ×100) A. 1 week after implantation in experimental rat, B. 1 week after implantation in control rat, C. 2 weeks after implantation in experimental rat, D. 2 weeks after implantation in control rat, E. 4 weeks after implantation in experimental rat, F. 4 weeks after implantation in control rat.

Fig. 6.

Microphotographs. (Masson's trichrome stain, ×100) A. 1 week after implantation in experimental rat, B. 1 week after implantation in control rat, C. 2 weeks after implantation in experimental rat, D. 2 weeks after implantation in control rat, E. 4 weeks after implantation in experimental rat, F. 4 weeks after implantation in control rat.

Fig. 7.

Osteoprotegerin antibody reactions. (×100) A. 1 week after implantation in experimental rat, B. 1 week after implantation in control rat, C. 2 weeks after implantation in experimental rat, D. 2 weeks after implantation in control rat, E. 4 weeks after implantation in experimental rat, F. 4 weeks after implantation in control rat.

Fig. 8.

Receptor activator of nuclear factor kB ligand antibody reactions.(×100) A. 1 week after implantation in experimental rat, B. 1 week after implantation in control rat, C. 2 weeks after implantation in experimental rat, D. 2 weeks after implantation in control rat, E. 4 weeks after implantation in experimental rat, F. 4 weeks after implantation in control rat.

Table 1.

Mean bone-to-implant contact in experimental and control group

Weeks after implantation	Experimental (%)	Control (%)	P value
1	39.00±18.17	42.87±9.27	.572
2	43.74±6.83	32.27±6.00	.190
4	32.62±11.02	47.10±9.77	.168

Data is presented by mean±SD.

Table 2.

Mean bone area in experimental and controlgroup

Weeks after implantation	Experimental (%)	Control (%)	P value
1	37.28±3.68	31.90±2.84	.080
2	20.62±2.47	15.64±2.69	.139
4	11.37±4.54	17.69±8.77	.405

Data is presented by mean±SD.

Table 3.

OPG expressions in experimental and control group

Weeks after implantation	Experimental	Control
1	+++	++/+
2	+++	++/+
4	+	+

(−: no immunoreactivity, +: weak but visible staining, ++: moderate staining, +++: strong staining intensity, OPG: Osteoprotegerin)

Table 4.

RANKL expressions in experimental and control group

Weeks after implantation	Experimental	Control
1	+	+
2	+	+
4	+	+

(−: no immunoreactivity, +: weak but visible staining, ++: moderate staining, +++: strong staining intensity, RANKL: Receptor activator of nuclear factor kB ligand)

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