Gene expression in older rats with delayed union of femoral fractures.
BACKGROUND
Fracture-healing slows with age. While six-week-old rats regain normal bone biomechanics at four weeks after a fracture, one-year-old rats require more than twenty-six weeks. The objective of this study was to examine the possible role of altered mRNA gene expression in this delayed union.


METHODS
Closed midshaft femoral fractures were created in six-week-old and one-year-old Sprague-Dawley female rats. The animals were killed at zero-time (unfractured) or at 0.4, one, two, three, four, or six weeks after the fracture. mRNA levels were measured by reverse transcription-polymerase chain reaction in the fracture callus for twenty-seven matrix, cytokine, and cytokine-receptor genes for the seven animals per time-point per age-group.


RESULTS
The younger rats healed radiographically by four weeks after the fracture, whereas none of the older rats had healed by the sixth week. Despite the difference in healing rates, the levels of mRNA gene expression, in general, followed the same pattern in both age-groups. The mRNA expression levels increased to a peak at one to two weeks after the fracture and then decreased to very low or undetectable levels at four and six weeks after the fracture for both age-groups. Significantly lower levels of mRNA for Indian hedgehog (Ihh) and bone morphogenetic protein-2 (BMP-2) were detected in the fracture calluses of the older rats (p < 0.01 and p < 0.05, respectively).


CONCLUSIONS
All genes studied were up-regulated by the fracture in both age-groups. Thus, the failure of the older rats to heal promptly was not due to the lack of expression of any of the studied genes. The increase in mRNA for Ihh and BMP-2 in the older rats was smaller than that in the younger rats, which may contribute to slower fracture repair. The return of mRNA gene expression to baseline in the older rats prior to healing may contribute to the delayed union. The slower healing response of the older rats did not stimulate a negative-feedback increase in the mRNA expression of stimulatory cytokines.
Meyer
Ralph A
RA
Orthopaedic Research Laboratory, Department of Orthopaedic Surgery, Carolinas Medical Center, Charlotte, North Carolina 28232, USA. rmeyer@carolinas.org
Meyer
Martha H
MH
Tenholder
Mark
M
Wondracek
Sharon
S
Wasserman
Ruth
R
Garges
Patricia
P
D000367:Age Factors; D053674:Aggrecans; D000375:Aging / Q000235:genetics*; D000818:Animals; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins / Q000032:analysis / Q000235:genetics; D002146:Bony Callus / Q000000981:diagnostic imaging / Q000473:pathology* / Q000503:physiopathology; D024043:Collagen Type II / Q000032:analysis / Q000235:genetics; D016207:Cytokines / Q000032:analysis / Q000235:genetics; D004195:Disease Models, Animal; D016326:Extracellular Matrix Proteins; D005260:Female; D005264:Femoral Fractures / Q000000981:diagnostic imaging / Q000473:pathology* / Q000503:physiopathology; D017102:Fracture Healing / Q000235:genetics*; D005599:Fractures, Ununited; D015870:Gene Expression / Q000235:genetics*; D053823:Hedgehog Proteins; D037181:Lectins, C-Type; D015675:Osteocalcin / Q000032:analysis / Q000235:genetics; D011509:Proteoglycans / Q000032:analysis / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis* / Q000235:genetics*; D011859:Radiography; D051381:Rats; D017207:Rats, Sprague-Dawley; D018121:Receptors, Cytokine / Q000032:analysis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017423:Sequence Analysis, RNA; D015534:Trans-Activators / Q000032:analysis / Q000235:genetics; D016212:Transforming Growth Factor beta; D015854:Up-Regulation / Q000235:genetics
C508321:Acan protein, rat; D053674:Aggrecans; C525720:Bmp2 protein, rat; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins; D024043:Collagen Type II; D016207:Cytokines; D016326:Extracellular Matrix Proteins; D053823:Hedgehog Proteins; D037181:Lectins, C-Type; D011509:Proteoglycans; D012333:RNA, Messenger; D018121:Receptors, Cytokine; D015534:Trans-Activators; D016212:Transforming Growth Factor beta; D015675:Osteocalcin