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Low-magnitude high-frequency loading, by whole-body vibration, accelerates early implant osseointegration in ovariectomized rats

Identifieur interne : 001265 ( Ncbi/Merge ); précédent : 001264; suivant : 001266

Low-magnitude high-frequency loading, by whole-body vibration, accelerates early implant osseointegration in ovariectomized rats

Auteurs : Yong-Qiang Liang [République populaire de Chine] ; Meng-Chun Qi [République populaire de Chine] ; Jiang Xu [République populaire de Chine] ; Juan Xu [République populaire de Chine] ; Hua-Wei Liu [République populaire de Chine] ; Wei Dong [République populaire de Chine] ; Jin-Yuan Li [République populaire de Chine] ; Min Hu [République populaire de Chine]

Source :

RBID : PMC:4227418

Abstract

Osteoporosis deteriorates jaw bone quality and may compromise early implant osseointegration and early implant loading. The influence of low-magnitude, high-frequency (LMHF) vibration on peri-implant bone healing and implant integration in osteoporotic bones remains poorly understood. LMHF loading via whole-body vibration (WBV) for 8 weeks has previously been demonstrated to significantly enhance bone-to-implant contact, peri-implant bone fraction and implant mechanical properties in osteoporotic rats. In the present study, LMHF loading by WBV was performed in osteoporotic rats, with a loading duration of 4 weeks during the early stages of bone healing. The results indicated that 4-week LMHF loading by WBV partly reversed the negative effects of osteoporosis and accelerated early peri-implant osseointegration in ovariectomized rats.


Url:
DOI: 10.3892/mmr.2014.2597
PubMed: 25270245
PubMed Central: 4227418

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PMC:4227418

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<p>Osteoporosis deteriorates jaw bone quality and may compromise early implant osseointegration and early implant loading. The influence of low-magnitude, high-frequency (LMHF) vibration on peri-implant bone healing and implant integration in osteoporotic bones remains poorly understood. LMHF loading via whole-body vibration (WBV) for 8 weeks has previously been demonstrated to significantly enhance bone-to-implant contact, peri-implant bone fraction and implant mechanical properties in osteoporotic rats. In the present study, LMHF loading by WBV was performed in osteoporotic rats, with a loading duration of 4 weeks during the early stages of bone healing. The results indicated that 4-week LMHF loading by WBV partly reversed the negative effects of osteoporosis and accelerated early peri-implant osseointegration in ovariectomized rats.</p>
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</TEI>
<pmc article-type="research-article">
<pmc-dir>properties open_access</pmc-dir>
<front>
<journal-meta>
<journal-id journal-id-type="nlm-ta">Mol Med Rep</journal-id>
<journal-id journal-id-type="iso-abbrev">Mol Med Rep</journal-id>
<journal-title-group>
<journal-title>Molecular Medicine Reports</journal-title>
</journal-title-group>
<issn pub-type="ppub">1791-2997</issn>
<issn pub-type="epub">1791-3004</issn>
<publisher>
<publisher-name>D.A. Spandidos</publisher-name>
</publisher>
</journal-meta>
<article-meta>
<article-id pub-id-type="pmid">25270245</article-id>
<article-id pub-id-type="pmc">4227418</article-id>
<article-id pub-id-type="doi">10.3892/mmr.2014.2597</article-id>
<article-id pub-id-type="publisher-id">mmr-10-06-2835</article-id>
<article-categories>
<subj-group subj-group-type="heading">
<subject>Articles</subject>
</subj-group>
</article-categories>
<title-group>
<article-title>Low-magnitude high-frequency loading, by whole-body vibration, accelerates early implant osseointegration in ovariectomized rats</article-title>
</title-group>
<contrib-group>
<contrib contrib-type="author">
<name>
<surname>LIANG</surname>
<given-names>YONG-QIANG</given-names>
</name>
<xref ref-type="aff" rid="af1-mmr-10-06-2835">1</xref>
<xref ref-type="aff" rid="af2-mmr-10-06-2835">2</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>QI</surname>
<given-names>MENG-CHUN</given-names>
</name>
<xref ref-type="aff" rid="af2-mmr-10-06-2835">2</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>XU</surname>
<given-names>JIANG</given-names>
</name>
<xref ref-type="aff" rid="af3-mmr-10-06-2835">3</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>XU</surname>
<given-names>JUAN</given-names>
</name>
<xref ref-type="aff" rid="af1-mmr-10-06-2835">1</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>LIU</surname>
<given-names>HUA-WEI</given-names>
</name>
<xref ref-type="aff" rid="af1-mmr-10-06-2835">1</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>DONG</surname>
<given-names>WEI</given-names>
</name>
<xref ref-type="aff" rid="af2-mmr-10-06-2835">2</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>LI</surname>
<given-names>JIN-YUAN</given-names>
</name>
<xref ref-type="aff" rid="af2-mmr-10-06-2835">2</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>HU</surname>
<given-names>MIN</given-names>
</name>
<xref ref-type="aff" rid="af1-mmr-10-06-2835">1</xref>
<xref ref-type="corresp" rid="c1-mmr-10-06-2835"></xref>
</contrib>
</contrib-group>
<aff id="af1-mmr-10-06-2835">
<label>1</label>
Department of Stomatology, Chinese PLA General Hospital, Beijing 100853, P.R. China</aff>
<aff id="af2-mmr-10-06-2835">
<label>2</label>
College of Stomatology, Hebei United University, Tangshan, Hebei 063000, P.R. China</aff>
<aff id="af3-mmr-10-06-2835">
<label>3</label>
Department of Stomatology, Tongchuan City People’s Hospital, Tongchuan, Shaanxi 727100, P.R. China</aff>
<author-notes>
<corresp id="c1-mmr-10-06-2835">Correspondence to: Professor Min Hu, Department of Stomatology, Chinese PLA General Hospital, 28 Fuxing Road, Haidian, Beijing 100853, P.R. China, E-mail:
<email>humin301@gmail.com</email>
</corresp>
</author-notes>
<pub-date pub-type="ppub">
<month>12</month>
<year>2014</year>
</pub-date>
<pub-date pub-type="epub">
<day>23</day>
<month>9</month>
<year>2014</year>
</pub-date>
<pub-date pub-type="pmc-release">
<day>23</day>
<month>9</month>
<year>2014</year>
</pub-date>
<pmc-comment> PMC Release delay is 0 months and 0 days and was based on the . </pmc-comment>
<volume>10</volume>
<issue>6</issue>
<fpage>2835</fpage>
<lpage>2842</lpage>
<history>
<date date-type="received">
<day>30</day>
<month>11</month>
<year>2013</year>
</date>
<date date-type="accepted">
<day>04</day>
<month>7</month>
<year>2014</year>
</date>
</history>
<permissions>
<copyright-statement>Copyright © 2014, Spandidos Publications</copyright-statement>
<copyright-year>2014</copyright-year>
<license license-type="open-access" xlink:href="http://creativecommons.org/licenses/by/3.0">
<license-p>This is an open-access article licensed under a Creative Commons Attribution-NonCommercial 3.0 Unported License. The article may be redistributed, reproduced, and reused for non-commercial purposes, provided the original source is properly cited.</license-p>
</license>
</permissions>
<abstract>
<p>Osteoporosis deteriorates jaw bone quality and may compromise early implant osseointegration and early implant loading. The influence of low-magnitude, high-frequency (LMHF) vibration on peri-implant bone healing and implant integration in osteoporotic bones remains poorly understood. LMHF loading via whole-body vibration (WBV) for 8 weeks has previously been demonstrated to significantly enhance bone-to-implant contact, peri-implant bone fraction and implant mechanical properties in osteoporotic rats. In the present study, LMHF loading by WBV was performed in osteoporotic rats, with a loading duration of 4 weeks during the early stages of bone healing. The results indicated that 4-week LMHF loading by WBV partly reversed the negative effects of osteoporosis and accelerated early peri-implant osseointegration in ovariectomized rats.</p>
</abstract>
<kwd-group>
<kwd>low-magnitude high-frequency loading</kwd>
<kwd>whole-body vibration</kwd>
<kwd>osteoporosis</kwd>
<kwd>dental implants</kwd>
<kwd>osseointegration</kwd>
<kwd>rat</kwd>
</kwd-group>
</article-meta>
</front>
<floats-group>
<fig id="f1-mmr-10-06-2835" orientation="portrait" position="float">
<label>Figure 1</label>
<caption>
<p>(A) X-ray demonstrating insertion of titanium implant in the proximal metaphysis of the tibia and (B) the vibrating device.</p>
</caption>
<graphic xlink:href="MMR-10-06-2835-g00"></graphic>
</fig>
<fig id="f2-mmr-10-06-2835" orientation="portrait" position="float">
<label>Figure 2</label>
<caption>
<p>Region of interest for evaluation of BV/TV. The area is defined as a rectangular region within 2 mm from the axis of the implant.</p>
</caption>
<graphic xlink:href="MMR-10-06-2835-g01"></graphic>
</fig>
<fig id="f3-mmr-10-06-2835" orientation="portrait" position="float">
<label>Figure 3</label>
<caption>
<p>Histological examination of the distal metaphyses of the femurs using Toluidine blue in (A) sham, (B) OVX and (C) OVX-V groups at four weeks following vibration. The trabeculae near the proximal metaphysis were sparse and discontinuous in the OVX group. This condition was ameliorated in the OVX-V group, although not to the level of the sham group. Magnification, ×100. Sham, sham operation; OVX, bilateral ovariectomy; OVX-V, bilateral ovariectomy with vibration treatment.</p>
</caption>
<graphic xlink:href="MMR-10-06-2835-g02"></graphic>
</fig>
<fig id="f4-mmr-10-06-2835" orientation="portrait" position="float">
<label>Figure 4</label>
<caption>
<p>Histological images of the proximal tibiae with implants using Toluidine blue stain. (A) sham, (B) OVX and (C) OVX-V groups at four weeks following vibration (magnification, ×40) and (D) sham, (E) OVX and (F) OVX-V groups at 10 weeks (magnification, ×200). Differences were observed in the trabecular structure and implant-bone contact around the implants between the three groups. The OVX group presented the poorest implant-bone integration and the least calcified trabecular structure, while vibration treatment markedly increased the presence of implant-bone integration and calcified trabecular structure in the OVX-V group, as compared with the OVX group. Sham, sham operation; OVX, bilateral ovariectomy; OVX-V, bilateral ovariectomy with vibration treatment.</p>
</caption>
<graphic xlink:href="MMR-10-06-2835-g03"></graphic>
</fig>
<fig id="f5-mmr-10-06-2835" orientation="portrait" position="float">
<label>Figure 5</label>
<caption>
<p>Observation of bone lamellae in direct contact with implant by fluorescence analysis. Fluorescent imaging of the (A) sham, (B) OVX and (C) OVX-V groups. Normal microscopy imaging without fluorescence in the (D) sham, (E) OVX and (F) OVX-V groups. A significant difference in fluorescence intensity, thickness and width of the double-labeled fluorescent lines was observed between the three groups. The OVX group exhibited the poorest fluorescent labeling, in which the fluorescence intensity was weak, fluorescent lines were thin and discontinuous and the distance of the double-labeled line was narrow. The vibration was markedly increased in the fluorescent labeling of the interface bone lamellae in the OVX-V group. The intensity of the fluorescence was stronger in the OVX-V group as compared with the OVX group. Magnification, ×600. Sham, sham operation; OVX, bilateral ovariectomy; OVX-V, bilateral ovariectomy with vibration treatment.</p>
</caption>
<graphic xlink:href="MMR-10-06-2835-g04"></graphic>
</fig>
<fig id="f6-mmr-10-06-2835" orientation="portrait" position="float">
<label>Figure 6</label>
<caption>
<p>Comparison of the removal torque (N.cm) between the three groups. One-way analysis of variance indicated a P<0.01 between the three groups, and post-hoc tests indicated a P<0.01 between any two groups. Sham, sham operation; OVX, bilateral ovariectomy; OVX-V, bilateral ovariectomy with vibration treatment.</p>
</caption>
<graphic xlink:href="MMR-10-06-2835-g05"></graphic>
</fig>
<table-wrap id="tI-mmr-10-06-2835" orientation="portrait" position="float">
<label>Table I</label>
<caption>
<p>Analysis of the body weight of rats and the bone mineral density of the femur.</p>
</caption>
<table frame="hsides" rules="groups">
<thead>
<tr>
<th valign="bottom" align="left" rowspan="1" colspan="1">Parameters</th>
<th valign="bottom" align="center" rowspan="1" colspan="1">Time</th>
<th valign="bottom" align="center" rowspan="1" colspan="1">Sham</th>
<th valign="bottom" align="center" rowspan="1" colspan="1">OVX</th>
<th valign="bottom" align="center" rowspan="1" colspan="1">OVX-V</th>
</tr>
</thead>
<tbody>
<tr>
<td valign="top" align="left" rowspan="1" colspan="1">Body weight (g)</td>
<td valign="top" align="left" rowspan="1" colspan="1">Pre-exp</td>
<td valign="top" align="left" rowspan="1" colspan="1">247.50±10.76</td>
<td valign="top" align="left" rowspan="1" colspan="1">246.33±7.59</td>
<td valign="top" align="left" rowspan="1" colspan="1">244.77±7.03</td>
</tr>
<tr>
<td valign="top" align="left" rowspan="1" colspan="1"></td>
<td valign="top" align="left" rowspan="1" colspan="1">Post-exp</td>
<td valign="top" align="left" rowspan="1" colspan="1">288.40±5.73</td>
<td valign="top" align="left" rowspan="1" colspan="1">322.18±7.78
<xref rid="tfn1-mmr-10-06-2835" ref-type="table-fn">a</xref>
</td>
<td valign="top" align="left" rowspan="1" colspan="1">314.10±5.14
<xref rid="tfn1-mmr-10-06-2835" ref-type="table-fn">a</xref>
,
<xref rid="tfn3-mmr-10-06-2835" ref-type="table-fn">c</xref>
</td>
</tr>
<tr>
<td valign="top" align="left" rowspan="1" colspan="1">Bone mineral density (g/cm
<sup>2</sup>
)</td>
<td valign="top" align="left" rowspan="1" colspan="1">Pre-exp</td>
<td valign="top" align="left" rowspan="1" colspan="1">0.2149±0.0119</td>
<td valign="top" align="left" rowspan="1" colspan="1">0.2182±0.0115</td>
<td valign="top" align="left" rowspan="1" colspan="1">0.2186±0.0107</td>
</tr>
<tr>
<td valign="top" align="left" rowspan="1" colspan="1"></td>
<td valign="top" align="left" rowspan="1" colspan="1">Post-exp</td>
<td valign="top" align="left" rowspan="1" colspan="1">0.2238±0.0204</td>
<td valign="top" align="left" rowspan="1" colspan="1">0.1806±0.0163
<xref rid="tfn1-mmr-10-06-2835" ref-type="table-fn">a</xref>
</td>
<td valign="top" align="left" rowspan="1" colspan="1">0.2005±0.0185
<xref rid="tfn2-mmr-10-06-2835" ref-type="table-fn">b</xref>
,
<xref rid="tfn3-mmr-10-06-2835" ref-type="table-fn">c</xref>
</td>
</tr>
</tbody>
</table>
<table-wrap-foot>
<fn id="tfn1-mmr-10-06-2835">
<label>a</label>
<p>P<0.01 vs. sham group;</p>
</fn>
<fn id="tfn2-mmr-10-06-2835">
<label>b</label>
<p>P<0.05 vs. sham group;</p>
</fn>
<fn id="tfn3-mmr-10-06-2835">
<label>c</label>
<p>P<0.05 vs. OVX group; n=12.</p>
</fn>
<fn id="tfn4-mmr-10-06-2835">
<p>Sham, sham operated group; OVX, bilateral ovariectomy group; OVX-V, bilateral ovariectomy with vibration treatment.</p>
</fn>
</table-wrap-foot>
</table-wrap>
<table-wrap id="tII-mmr-10-06-2835" orientation="portrait" position="float">
<label>Table II</label>
<caption>
<p>Histomorphometric analysis of bone indices.</p>
</caption>
<table frame="hsides" rules="groups">
<thead>
<tr>
<th valign="bottom" align="left" rowspan="1" colspan="1">Indices</th>
<th valign="bottom" align="center" rowspan="1" colspan="1">Sham</th>
<th valign="bottom" align="center" rowspan="1" colspan="1">OVX</th>
<th valign="bottom" align="center" rowspan="1" colspan="1">OVX-V</th>
</tr>
</thead>
<tbody>
<tr>
<td valign="top" align="left" rowspan="1" colspan="1">BIC</td>
<td valign="top" align="center" rowspan="1" colspan="1">63.64±4.74</td>
<td valign="top" align="center" rowspan="1" colspan="1">37.66±4.63
<xref rid="tfn5-mmr-10-06-2835" ref-type="table-fn">a</xref>
</td>
<td valign="top" align="left" rowspan="1" colspan="1">58.09±4.56
<xref rid="tfn6-mmr-10-06-2835" ref-type="table-fn">b</xref>
</td>
</tr>
<tr>
<td valign="top" align="left" rowspan="1" colspan="1">BF/TV</td>
<td valign="top" align="center" rowspan="1" colspan="1">56.74±2.81</td>
<td valign="top" align="center" rowspan="1" colspan="1">40.39±7.08
<xref rid="tfn5-mmr-10-06-2835" ref-type="table-fn">a</xref>
</td>
<td valign="top" align="left" rowspan="1" colspan="1">48.05±4.18
<xref rid="tfn5-mmr-10-06-2835" ref-type="table-fn">a</xref>
,
<xref rid="tfn7-mmr-10-06-2835" ref-type="table-fn">c</xref>
</td>
</tr>
<tr>
<td valign="top" align="left" rowspan="1" colspan="1">TBL</td>
<td valign="top" align="center" rowspan="1" colspan="1">65.34±6.90</td>
<td valign="top" align="center" rowspan="1" colspan="1">47.42±7.06
<xref rid="tfn5-mmr-10-06-2835" ref-type="table-fn">a</xref>
</td>
<td valign="top" align="left" rowspan="1" colspan="1">55.60±3.45
<xref rid="tfn5-mmr-10-06-2835" ref-type="table-fn">a</xref>
,
<xref rid="tfn8-mmr-10-06-2835" ref-type="table-fn">d</xref>
</td>
</tr>
</tbody>
</table>
<table-wrap-foot>
<fn id="tfn5-mmr-10-06-2835">
<label>a</label>
<p>P<0.01 vs. sham group;</p>
</fn>
<fn id="tfn6-mmr-10-06-2835">
<label>b</label>
<p>P<0.05 vs. sham group;</p>
</fn>
<fn id="tfn7-mmr-10-06-2835">
<label>c</label>
<p>P<0.05 vs. OVX group;</p>
</fn>
<fn id="tfn8-mmr-10-06-2835">
<label>d</label>
<p>P<0.01 vs. OVX group; n=12 in each group.</p>
</fn>
<fn id="tfn9-mmr-10-06-2835">
<p>Sham, sham operation; OVX, bilateral ovariectomy; OVX-V, bilateral ovariectomy with vibration treatment; TBL, thickness of bone lamellae.</p>
</fn>
</table-wrap-foot>
</table-wrap>
<table-wrap id="tIII-mmr-10-06-2835" orientation="portrait" position="float">
<label>Table III</label>
<caption>
<p>Dynamic indexes of bone metabolism (n=12).</p>
</caption>
<table frame="hsides" rules="groups">
<thead>
<tr>
<th valign="bottom" align="left" rowspan="1" colspan="1">Indices</th>
<th valign="bottom" align="center" rowspan="1" colspan="1">Sham</th>
<th valign="bottom" align="center" rowspan="1" colspan="1">OVX</th>
<th valign="bottom" align="center" rowspan="1" colspan="1">OVX-V</th>
</tr>
</thead>
<tbody>
<tr>
<td valign="top" align="left" rowspan="1" colspan="1">MAR</td>
<td valign="top" align="center" rowspan="1" colspan="1">2.63±0.25</td>
<td valign="top" align="center" rowspan="1" colspan="1">1.64±0.32
<xref rid="tfn10-mmr-10-06-2835" ref-type="table-fn">a</xref>
</td>
<td valign="top" align="center" rowspan="1" colspan="1">2.27±0.34
<xref rid="tfn11-mmr-10-06-2835" ref-type="table-fn">b</xref>
,
<xref rid="tfn13-mmr-10-06-2835" ref-type="table-fn">d</xref>
</td>
</tr>
<tr>
<td valign="top" align="left" rowspan="1" colspan="1">MS/BS</td>
<td valign="top" align="center" rowspan="1" colspan="1">0.24±0.06</td>
<td valign="top" align="center" rowspan="1" colspan="1">0.15±0.02
<xref rid="tfn10-mmr-10-06-2835" ref-type="table-fn">a</xref>
</td>
<td valign="top" align="center" rowspan="1" colspan="1">0.18±0.03
<xref rid="tfn11-mmr-10-06-2835" ref-type="table-fn">b</xref>
,
<xref rid="tfn12-mmr-10-06-2835" ref-type="table-fn">c</xref>
</td>
</tr>
<tr>
<td valign="top" align="left" rowspan="1" colspan="1">BFR/BS</td>
<td valign="top" align="center" rowspan="1" colspan="1">0.61±0.12</td>
<td valign="top" align="center" rowspan="1" colspan="1">0.25±0.08
<xref rid="tfn10-mmr-10-06-2835" ref-type="table-fn">a</xref>
</td>
<td valign="top" align="center" rowspan="1" colspan="1">0.43±0.11
<xref rid="tfn10-mmr-10-06-2835" ref-type="table-fn">a</xref>
,
<xref rid="tfn13-mmr-10-06-2835" ref-type="table-fn">d</xref>
</td>
</tr>
</tbody>
</table>
<table-wrap-foot>
<fn id="tfn10-mmr-10-06-2835">
<label>a</label>
<p>P<0.01 vs. sham group;</p>
</fn>
<fn id="tfn11-mmr-10-06-2835">
<label>b</label>
<p>P<0.05 vs. sham group;</p>
</fn>
<fn id="tfn12-mmr-10-06-2835">
<label>c</label>
<p>P<0.05 vs. OVX group;</p>
</fn>
<fn id="tfn13-mmr-10-06-2835">
<label>d</label>
<p>P<0.01 vs. OVX group; n=12 in each group.</p>
</fn>
<fn id="tfn14-mmr-10-06-2835">
<p>Sham, sham operation; OVX, bilateral ovariectomy; OVX-V, bilateral ovariectomy with vibration treatment; MAR, mineral apposition rate; MS/BS, ratio of mineralising surface to bone surface; BFR/BS, bone formation rate per unit of bone surface.</p>
</fn>
</table-wrap-foot>
</table-wrap>
</floats-group>
</pmc>
<affiliations>
<list>
<country>
<li>République populaire de Chine</li>
</country>
<settlement>
<li>Pékin</li>
</settlement>
</list>
<tree>
<country name="République populaire de Chine">
<noRegion>
<name sortKey="Liang, Yong Qiang" sort="Liang, Yong Qiang" uniqKey="Liang Y" first="Yong-Qiang" last="Liang">Yong-Qiang Liang</name>
</noRegion>
<name sortKey="Dong, Wei" sort="Dong, Wei" uniqKey="Dong W" first="Wei" last="Dong">Wei Dong</name>
<name sortKey="Hu, Min" sort="Hu, Min" uniqKey="Hu M" first="Min" last="Hu">Min Hu</name>
<name sortKey="Li, Jin Yuan" sort="Li, Jin Yuan" uniqKey="Li J" first="Jin-Yuan" last="Li">Jin-Yuan Li</name>
<name sortKey="Liang, Yong Qiang" sort="Liang, Yong Qiang" uniqKey="Liang Y" first="Yong-Qiang" last="Liang">Yong-Qiang Liang</name>
<name sortKey="Liu, Hua Wei" sort="Liu, Hua Wei" uniqKey="Liu H" first="Hua-Wei" last="Liu">Hua-Wei Liu</name>
<name sortKey="Qi, Meng Chun" sort="Qi, Meng Chun" uniqKey="Qi M" first="Meng-Chun" last="Qi">Meng-Chun Qi</name>
<name sortKey="Xu, Jiang" sort="Xu, Jiang" uniqKey="Xu J" first="Jiang" last="Xu">Jiang Xu</name>
<name sortKey="Xu, Juan" sort="Xu, Juan" uniqKey="Xu J" first="Juan" last="Xu">Juan Xu</name>
</country>
</tree>
</affiliations>
</record>

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