The growth plate is responsible for longitudinal bone growth. The problem of repair of damaged growth plate in children has never been adequately solved. The purpose of this study is to investigate the ability of the cultured chondrocyte for the prevention of bony bridge and repairment of damaged growth plate. Chondrocytes were obtained from the new born canine epiphyseal plate and was cultured in high density. Fourteen days later they formed micromass easily removable from the culture flask. Twenty dogs were divided into two groups; in group I, the medial proximal tibial growth plate was destroyed and then cultured chondrocytes were transplanted into the defect, and in group II, the medial proximal tibial growth plate was left in destroyed state. Each left leg was remained as a control. The growth pattern was observed radiographically and histologically until 16 weeks after graft. 4 weeks after the operation, the angular deformity had been observed, and 31 degrees of angulation was noted at the 16th week in group II, while there was less than 8 degrees of angulation and nearly normal growth in most of dogs of group I (8 of 10 dogs). The other 2 dogs had shown 20 degrees angulation. In group II, there was definite bony bridge on the medical proximal growth plate. In group I, initially, the cultured chondrocyte remained as a amorphous cartilagenous mass, but as time progressed, amorphous cartilagenous mass had formed cartilagenous matrix which was proved by Safranin-O staining. Although this study showed the role of cultured chondrocyte as a method of preventation of bony bridge formation and possibility to repair of growth plate, further studies should be done to prove the reconstruction of the growth plate.
Animal ; Cartilage/*cytology ; *Cell Transplantation ; Cells, Cultured ; Dogs ; Growth Plate/injuries/*surgery ; Transplantation, Homologous

OBJECTIVETo develop an epiphyseal slide-traction plate in child, which can supply the fracture a sufficient internal fixation, and will not restrain the growth of epiphyses. Animal experiments were carried out with the plates to compare the slide-traction with traditional plate.

METHODSDevelop a slide-traction plate for the configuration of the femur condylus of children. Thirty adolescent goats in the experiment were divided into control group (12 goats) and plate group (18 goats). In plate group, right femurs of goats were fixed with common plates and the left femurs with slide-traction plates. All the goats were given X-ray examination at different time after surgery. And the goats were sacrificed at 3 and 6 month, histological method and electron microscopy were performed to evaluate the development of epiphyseal plate.

RESULTSThe both femurs of the goats in control group have no difference in evidence in length at all time we examined. And the both femurs of the goats fixed with plates have no difference in evidence in length at 1 day after surgery. However, the both femurs of the goats fixed with plates have difference in evidence in length at 1 month, 2 month, 3 month, 6 month after surgery. The increased length of the femurs at I month, 2 month, 3 month, 6 month after surgery was also compared with the length at 1 day after surgery, there was difference in evidence between the right femurs of the control group and the femurs were fixed with common plates, but no difference in evidence between the left femurs of the normal control group and the femurs were fixed with slide-traction plate (P > 0.05). More thicker epiphyseal plate were found in the left femurs than the right femurs of the group fixed with plates at 3 and 6 month after surgery (P < 0.01). In the plate group, safranine O staining showed epiphyseal plates at the left femurs had more fuscous staining than the right femurs at 3 and 6 month after surgery and electron microscopy also found that the cells of the epiphyseal plates of left femurs were more eugenic than the right femurs at 3 and 6 month after surgery.

CONCLUSIONThe epiphyseal slide-traction plate can slide with the growth of epiphyses, which is suitable for fixation of the fracture in this part.

Animals ; Bone Plates ; Female ; Femur ; cytology ; diagnostic imaging ; growth & development ; surgery ; Goats ; Growth Plate ; cytology ; diagnostic imaging ; growth & development ; surgery ; Humans ; Internal Fixators ; Male ; Orthopedic Procedures ; Radiography ; Traction

PURPOSE: To examine the effects of change in weight bearing on the growth plate metabolism, a simulated animal model of weightlessness was introduced and the chondrocytes' cellular kinetics was evaluated. MATERIALS AND METHODS: Unloading condition on the hind-limb of Sprague-Dawley rats was created by fixing a tail and lifting the hind-limb. Six rats aged 6 weeks old were assigned to each group of unloading, reloading, and control groups of unloading or reloading. Unloading was maintained for three weeks, and then reloading was applied for another one week thereafter. Histomorphometry for the assessment of vertical length of the growth plate, 5-bromo-2'-deoxyuridin immunohistochemistry for cellular kinetics, and biotin nick end labeling transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL) assay for chondrocytes apoptosis in the growth plate were performed. RESULTS: The vertical length of the growth plate and the proliferative potential of chondrocytes were decreased in the unloading group compared to those of control groups. Inter-group differences were more significant in the proliferative and hypertrophic zones. Reloading increased the length of growth plate and proliferative potential of chondrocytes. However, apoptotic changes in the growth plate were not affected by the alterations of weight bearing. CONCLUSION: Alterations in the weight bearing induced changes in the chondrocytic proliferative potential of the growth plate, however, had no effects on the apoptosis. This may explain why non-weight bearing in various clinical situations hampers normal longitudinal bone growth. Further studies on the factors for reversibility of chondrocytic proliferation upon variable mechanical stresses are needed.
Animals ; Apoptosis/physiology ; Cell Proliferation ; Chondrocytes/cytology ; Growth Plate/*cytology ; In Situ Nick-End Labeling ; Kinetics ; Rats ; Rats, Sprague-Dawley ; Weight-Bearing/*physiology

OBJECTIVETo compare the cell viability of chondrocytes between the anterior and the posterior spinal growth plates in adolescent idiopathic scoliosis (AIS) by proliferation and apoptosis labelling.

METHODSSeventeen AIS patients (4 male and 13 female, mean age 13.6 years old, ranged from 10 to 17 years old) were recruited in this study. Growth plates were harvested during anterior and posterior surgery. PCNA (proliferating cell nuclear antigen) and TUNEL (terminal deoxynucleotide transferase-mediated nick end labeling) were used for proliferation and apoptosis labeling on chondrocytes respectively.

RESULTSIn AIS, the distribution of the proliferating nests were denser and more parallel in anterior column than those in posterior under microscope observation. In the proliferative and hypertrophic zone the PCNA index and PCNA/TUNEL ratio were higher in the anterior column than those in the posterior column (P < 0.05 and P < 0.01 respectively). While in resting zone the differences were not so significant.

CONCLUSIONIn adolescent idiopathic scoliosis the growth viability of chondrocytes is more vigorous in anterior spinal column than in the posterior column.

Adolescent ; Apoptosis ; Cell Proliferation ; Cell Survival ; Child ; Chondrocytes ; cytology ; Female ; Growth Plate ; pathology ; Humans ; Immunohistochemistry ; In Situ Nick-End Labeling ; Male ; Scoliosis ; pathology ; physiopathology ; Spine ; growth & development ; pathology

OBJECTIVETo compare the histological features of the thoracic vertebral body growth plates (VBGPs) of rats at different ages and assess their proliferative capability.

METHODSThe thoracic VBGPs obtained from rats aged 1 day and 1, 4, 8, 16 and 28 weeks were identified using safranin O-fast green staining, and the height of the hypertrophic zone, proliferative zone, and resting zone were measured. The chondrocytes were isolated from these VBGPs with a modified trypsin-collagenase type II digestion method for primary culture in vitro. The expressions of proliferating cell nuclear antigen (PCNA) mRNA and protein was detected by real time-PCR and Western blotting, respectively.

RESULTSThe 1-day- and 1-week-old rats showed significantly greater hypertrophic zone and proliferative zone in the VBGPs than older rats (P<0.01); the proliferative zone was significantly greater in rats aged 4 weeks than in those aged 28 weeks (P<0.05). The resting zone was obviously greater in rats aged 1 day and 1 week than in older rats (P<0.05), and also greater in rats aged 4 weeks than in those aged 16 and 28 weeks (P<0.05). Obvious ossification in the resting zone occurred at 16 weeks, and most of the resting zone became ossified at 28 weeks. The expression of PCNA decreased at both the mRNA and protein levels as the rats grew.

CONCLUSIONThe 3 zones of VBGPs are greater in rats aged 1 day and 1 week than in older ones. Ossification in the resting zone begins at 16 weeks, and till 28 weeks, most of the resting zone is ossified. The proliferation ability of VBGP chondrocytes decreases with the increase of age of the rats.

Age Factors ; Animals ; Animals, Newborn ; Cell Proliferation ; Cells, Cultured ; Chondrocytes ; cytology ; Growth Plate ; anatomy & histology ; cytology ; Male ; Proliferating Cell Nuclear Antigen ; analysis ; RNA, Messenger ; analysis ; Rats ; Rats, Sprague-Dawley ; Thoracic Vertebrae ; growth & development

OBJECTIVETo observe the effects of Nourishing yin clearing heat ([Chinese characters: see text]) Chinese herbal medicine on vascular endothelial growth factor (VEGF) in growth retardation induced by Decamethasone and observe its mechanisms.

METHODSThirty one-month-old New Zealand white rabbits were randomly divided into normal group, dexamethasone-treated group and Nourishing yin clearing heat ([Chinese characters: see text]) Chinese herbal medicine-treated group. The rabbits in dexamethasone group and Nourishing yin clearing heat ([Chinese characters: see text]) Chinese herbal medicine-treated group received dexamethasone (5 mg/kg x d). The rabbits were sacrificed at the 6th and 12th week after administration, and then rabbit tibia articular was removed. (1) Using TUNEL stain to detect apoptotic index. (2) Using immunohistochemical stain to detect the positive index of the expression of vascular endothelial growth factor (VEGF) in the epiphyseal cartilage of growth. (3) Using fluorescent quantitative PCR to detect the expression intensity of VEGF mRNA in each group.

RESULTSAt the 6th and 12th week after administration, there were significant difference in apoptotic index and cell proliferation index between dexamethasone group and normal group (P<0.01, dexamethasone group more than normal group). Immunohistochemical stain and fluorescent quantitative PCR indicated that the expression of VEGF and VEGF mRNA in dexamethasone group was significantly decreased as compared with that in normal group (P<0.01), and also obviously lower than Chinese herbal medicine-treated group (P<0.01).

CONCLUSIONVEGF has 2. an important role during the growth retardation induced by Dexamethasone. Nourishing yin clearing heat ([Chinese characters: see text]) Chinese herbal medicine can reduce the growth retardation induced by Dexamethasone through increasing the VEGF expression in growth plate chondrocytes and then increase angiogenesis.

Animals ; Apoptosis ; Cell Proliferation ; Dexamethasone ; pharmacology ; Drugs, Chinese Herbal ; pharmacology ; Female ; Gene Expression ; drug effects ; Growth Plate ; cytology ; drug effects ; growth & development ; metabolism ; Male ; Rabbits ; Random Allocation ; Vascular Endothelial Growth Factor A ; genetics ; metabolism

PURPOSE: Pathologic changes in the growth plate remain unknown in Legg-Calve-Perthes (LCP) disease. Spontaneously hypertensive rats have proven to be a good model for studying LCP disease. This study investigated the histopathologic changes and the expression of vascular endothelial growth factor in the growth plate of spontaneously hypertensive rats (SHR). MATERIALS AND METHODS: Sixty SHR rats were divided into two groups: those showing osteonecrosis (SHR+n group: 32), and those showing normal ossification (SHR-n group: 28). Thirty Wister Kyoto rats served as a control. For histomorphological measurement, the length of each zone of the growth plate was measured. Cell kinetics was measured by 5-bromo-2'-deoxyuridin (BrdU) immunohistochemistry and transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL) assays. Vascular endothelial growth factor (VEGF) immunohistochemistry was used to identify of expression of VEGF. RESULTS: The lengths of growth plates of the SHR+n group were significantly shorter in the initial growth period than those of the other groups. The lowest proliferative rate and the highest apoptosis rate were observed in the SHR+n group at the initial growth period. The expression of VEGF in the growth plate of the SHR group was lower than the control group, and it was lower in the SHR+n group than in the SHR-n group. CONCLUSION: The growth plate of the SHR+n group was found to be affected by disease process of ischemic necrosis of the femoral head, and this might explain the relative overgrowth of the greater trochanter in the later stages of LCP disease.
Animals ; Apoptosis ; Femur Head/metabolism/*pathology ; Femur Head Necrosis/metabolism/pathology ; Growth Plate/*cytology/metabolism ; Osteogenesis/physiology ; Rats ; Rats, Inbred SHR ; Rats, Sprague-Dawley ; Vascular Endothelial Growth Factor A/metabolism

PURPOSE: The purpose of this study was to investigate the effects of transplantation of an in vitro-generated, scaffold-free, tissue-engineered cartilage tissue analogue (CTA) using a suspension chondrocyte culture in a rabbit growth-arrest model. MATERIALS AND METHODS: We harvested cartilage cells from the articular cartilage of the joints of white rabbits and made a CTA using a suspension culture of 2x107 cells/mL. An animal growth plate defect model was made on the medial side of the proximal tibial growth plate of both tibias of 6-week-old New Zealand white rabbits (n=10). The allogenic CTA was then transplanted onto the right proximal tibial defect. As a control, no implantation was performed on the left-side defect. Plain radiographs and the medial proximal tibial angle were obtained at 1-week intervals for evaluation of bone bridge formation and the degree of angular deformity until postoperative week 6. We performed a histological evaluation using hematoxylin-eosin and Alcian blue staining at postoperative weeks 4 and 6. RESULTS: Radiologic study revealed a median medial proximal tibial angle of 59.0degrees in the control group and 80.0degrees in the CTA group at 6 weeks. In the control group, statistically significant angular deformities were seen 3 weeks after transplantation (p<0.05). On histological examination, the transplanted CTA was maintained in the CTA group at 4 and 6 weeks postoperative. Bone bridge formation was observed in the control group. CONCLUSION: In this study, CTA transplantation minimized deformity in the rabbit growth plate injury model, probably via the attenuation of bone bridge formation.
Animals ; *Bone Transplantation ; Cartilage/anatomy & histology ; Cell Culture Techniques ; Cells, Cultured ; Chondrocytes/*cytology/transplantation ; Growth Plate/anatomy & histology/*surgery ; *Mesenchymal Stem Cell Transplantation ; Rabbits ; Tibia/*surgery ; Tissue Engineering ; Transplantation, Autologous/methods ; Transplantation, Homologous

OBJECTIVETo investigate the effects and the mechanisms of stanozolol (ST) on the proliferation, maturation and differentiation of in vitro cultured growth plate chondrocyte isolated from gonadotropin releasing hormone analogue (GnRHa)-treated adolescent rats, to study if ST mediates the proliferation of chondrocytes via the estrogen receptor alpha (ERalpha), androgen receptor (AR) and/or insulin-like growth factor-1 receptor (IGF-1R) and interactions of the two receptor and IGF-1R receptor signaling pathway, to investigate the mechanism of the biological effects in ST promoting bone growth/maturity at molecular level.

METHODThe rats were weaned at the end of 3 weeks and intramuscular injection of triptorelin of GnRHa preparations, qow x 2 was started. The rats were sacrificed at the end of 7 weeks, and then the tibiae growth plates were taken out with sterile procedure. The chondrocytes were obtained by two-time enzyme digestion method, and the experiments were carried out with the primary chondrocytes. Immunohistochemical staining of proliferating cell nuclear antigen (PCNA) and Western blot analysis were applied.

RESULTThe results of PCNA demonstrated that stanozolol enhanced the proliferation of the chondrocytes, time-course studies showed that the proliferation were maximally stimulated by stanozolol after 2 days of incubation and decreased again after longer periods of incubation. The expression of p-ERalpha, p-IGF-1R and p-extracellular-signal regulated kinase 1/2 (ERK1/2) increased with the incubation period of ST treatment, and reached the peak value at a certain time, and then gradually decreased. The expression of p-ERalpha, p-IGF-1R and p-ERK1/2 increased with the elevation of ST concentration, and reached the peak value at 10(-9) - 10(-8) mol/L, then gradually decreased. ST induced-p-ERalpha expression was partially blocked by ERalpha and mitogen-activated protein kinase kinase inhibitors. ST induced-p-IGF-1R expression was partially blocked by ERalpha and IGF-1R inhibitors. ST induced-p-ERK1/2 expression was partially blocked by mitogen-activated protein kinase kinase and IGF-1R inhibitors.

CONCLUSIONAs an androgen derivation, ST exerts its biological effects of promoting proliferation of the long bone growth plate chondrocytes via activating the classic ERalpha receptor pathway and mitogen-activated protein kinase pathway, and at the same time, by activation of IGF-1R. Both IGF-1R and ERalpha can promote "cross-talk" of two systems' receptor signal through mitogen-activated protein kinase signal pathway.

Androgens ; pharmacology ; Animals ; Cells, Cultured ; Chondrocytes ; cytology ; drug effects ; metabolism ; Estrogen Receptor alpha ; metabolism ; Female ; Growth Plate ; drug effects ; metabolism ; Mitogen-Activated Protein Kinase 3 ; metabolism ; Rats ; Receptor Cross-Talk ; Receptor, IGF Type 1 ; metabolism ; Signal Transduction ; drug effects ; Stanozolol ; pharmacology