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PubMed1994

Formation of nodular structures resembling mature articular cartilage in long-term primary cultures of human fetal epiphyseal chondrocytes on a hydrogel substrate.

OBJECTIVE: To establish long-term cultures of human fetal epiphyseal chondrocytes under conditions that allow the preservation of a cartilage-specific phenotype. METHODS: Chondrocytes isolated from 20-24-week human fetal epiphyseal cartilage were cultured for up to 180 days on plastic dishes previously coated with the hydrogel, poly-(2-hydroxyethyl methacrylate). Morphologic, ultrastructural, and biochemical characteristics of the cultures were examined at various intervals, and the expression of genes encoding types I, II, and IX collagen and aggrecan core protein was determined by Northern hybridizations of total cellular RNA with human-specific complementary DNAs. RESULTS: Human fetal epiphyseal chondrocytes cultured for 180 days under conditions that prevented their attachment to the underlying substratum formed nodular structures with morphologic and structural characteristics resembling mature articular cartilage. The cells in the center of the nodules remained spherical and were surrounded by an abundant cartilaginous extracellular matrix, as evidenced by histochemical and ultrastructural examinations. The cells in the periphery of the nodules acquired a discoid morphology and were surrounded by a sparse extracellular matrix. Biosynthetic studies demonstrated the maintenance of a cartilage-specific phenotype throughout the 180 days of culture, with the production of aggrecan and types II, IX, and XI collagens but not type I collagen. Northern hybridizations showed high levels of messenger RNAs (mRNAs) for aggrecan core protein, type II procollagen, and type IX collagen, but type I procollagen mRNA was not detectable even at 180 days of culture. CONCLUSION: The human chondrocyte culture system described here allows the maintenance of a chondrocyte-specific phenotype for prolonged periods (up to 180 days). The long-term chondrocyte cultures formed nodular structures that resemble mature articular cartilage morphologically, ultrastructurally, biosynthetically, and in the pattern of cartilage-specific gene expression.

Blotting, Northern↗
PubMed1995

Can quantitative 99mTc-MDP bone scans be used to predict longitudinal growth of epiphyseal plate allografts after microvascular transplantation? An experimental study.

Allograft and autograft microvascular proximal tibial epiphyseal plate transplants were performed in female New Zealand White (NZW) rabbits to quantify the growth rate and total growth potential of immunosuppressed and nonimmunosuppressed rabbits. The purpose of this experiment is to examine whether the 99mTc-MDP radionuclide uptake of the transplanted epiphyseal plate at 1 week postoperatively, done to assess anastomotic patency of the transplant, could also serve as a predictor of eventual longitudinal growth of the transplant or replant. All transplants and replants demonstrating positive 99mTc-MDP uptake in the proximal tibial epiphyseal plate at 1 week showed continued longitudinal growth. The precise amount of 99mTc-MDP uptake, however, did not correlate with the amount of growth at 3 and 5 weeks follow-up.

Animals↗
PubMed1986

The effects of different steroids on costal and epiphyseal cartilage of fetal and adult rats.

The effects of different doses of various steroids on growth, and on costal and epiphyseal chondrocytes, have been studied in prenatal, immature, and adult Long-Evans rats using histochemical techniques, and both light and electron microscopy. Both prenatal and postnatal treatments have been employed. The steroids used were cortisone (CA), betamethasome (BM), and, in the prenatal group only, dexamethasone (DM). Body weight is reduced in all treated rats (except the low dose of CA) by day 17 of gestation, with greater weight reductions occurring in rats receiving the higher dose level of each steroid. In rats treated prenatally or neonatally, and sacrificed postnatally on days 39-43 or days 116-127, body weights, and tibial and tail lengths, are less than in correspondingly aged controls, thus showing a persistence of the effects of treatment. Costal and epiphyseal cartilages in prenatal rats show cellular, synthetic, and ultrastructural alterations induced by treatment with glucocorticoids but the responses are not necessarily comparable. Except for the low dose of DM, the higher doses of each steroid are more effective in inhibiting, or altering, growth and cellular differentiation in the developing fetuses. Surprisingly, a low dose of DM has a more devastating effect on the cells and extracellular matrix of both costal and epiphyseal cartilage, than do higher dose-levels of the various steroids. Low doses of CA and BM are also effective in inhibiting or altering growth and cellular differentiation, but their effectiveness is largely limited to 17 days of gestation. The order of effect of the various doses of the different steroids on fetal cartilage, listed in decreasing order of severity, is as follows: 0.12 DM, 0.24 DM, 0.42 BM, 50 CA, with 25 CA and 0.18 BM being approximately equal and only slightly different from control cartilages. The effect of prenatal or neonatal glucocorticoid treatment on chondrocytes is minimal in the 30-43 days, or 116-127 day, postnatal groups. In immature and adult rats, cortisone affects the chondrocytes more deleteriously than does betamethasone, and a 5.0 mg dose of CA seems to affect chondrocytes, body weight, and tibial and tail lengths more than 0.2 or 7.5 mg doses.

Animals↗
PubMed1992

Epiphyseal chondroblastoma of bone. Long-term effects on skeletal growth and articular function in 15 cases treated surgically.

The authors describe the long-term sequelae of chondroblastoma in 15 patients with open growth plates, whose age at operation ranged from 8 to 15 years. At follow-up, the youngest patient was 21 and the oldest 48 years old. Upper limb length discrepancy ranging from 2 cm to 10 cm was present in the four patients who had a proximal humeral epiphyseal location; in three of them, the range of motion of the shoulder was also limited and X-rays showed marked irregularities of the humeral head. Lower limb length discrepancy ranging from 0.5 cm to 2 cm was found in five of the eight patients in whom the tumor affected lower limb epiphyses. One patient with proximal tibial epiphyseal involvement also had mild genu valgum. Radiographic osteoarthritis was present only in the trapeziometacarpal joint of a patient in whom the first metacarpal bone affected by the tumor was replaced by a free fibular graft. The abnormalities observed did not cause important functional loss in either the everyday or the working activities of any of our patients.

Adult↗
PubMed1990

Stippled epiphyses in fetal alcohol syndrome.

We report on punctate epiphyseal calcifications (stippled epiphyses) in the fetal alcohol syndrome and present the differential diagnosis of chondrodysplasia punctata. A literature survey shows that epiphyseal calcifications accompanying alcoholic embryopathy are regularly located in the lower limbs and rarely found in the upper extremities.

Calcinosis↗
PubMed1979

Identification of phosphopeptides and gamma-carboxyglutamic acid-containing peptides in epiphyseal growth plate cartilage.

Uncalcified cartilage from the epiphyseal portion of bovine scapulae, both distant and adjacent to the epiphyseal growth plate, and the calcified cartilage of the epiphyseal growth plate itself were analyzed for the presence of O-phosphoserine [Ser(P)], O-phosphothreonine [Thr(P)] and gamma-carboxyglutamic acid (G1a). Only trace amounts of these Ca2+-binding amino acids or the peptides containing them were found in the unmineralized tissues. In contrast, whole calcified cartilage, and especially the most mineralized fraction obtained by density centrifugation, contained considerable amounts of all three amino acids. Essentially all of the G1a and the majority of the Ser(P) and Thr(P) were present in non-collagenous, non-diffusible proteins extractable in EDTA at near-neutral pH.

1-Carboxyglutamic Acid↗
PubMed1995

[Behavior of artificially-induced epiphyseal groove defects. 3: Transplantation of autologous and homologous rib cartilage in Göttingen minipigs. Findings after a 16 week interval].

Drillholes of 8 mm diameter were made through the epiphyseal cartilage of the distal femora in 20 6-week-old Göttingen minipigs. The defects were filled with autologous or with homologous cartilage. This was intended to prevent epi-metaphyseal osseous bridge formation. If of appropriate size and location, the latter would lead to subsequent misgrowth. In group A (autologous costal cartilage), at all drillholes the transplanted costal cartilage can prevent ossification of the defect. An epiphyseal boundary lamella was formed over the transplant which precluded penetration of vessels into the defect. In group B (homologous costal cartilage), the transplanted costal cartilage showed a tendency to mineralization in all preparations at the 20 drillholes available. The cartilage was integrated into the primary cancellous bone developing within the defect. An epiphyseal boundary lamella had not formed in the period of the experiment (in contrast to autogenous transplantation). These investigations show that autologous costal cartilage is superior to homologous costal cartilage in the potential clinical application of costal cartilage transplants in the treatment of Brodie abscesses or post-traumatic epiphyseodeses.

Animals↗
PubMed2000

Delayed closure of epiphyseal cartilages induced by the aromatase inhibitor anastrozole. Would it help short children grow up?

Estrogens locally generated from androgen precursors due to the action of aromatase play a main role in epiphyseal cartilage fusion. Treatment with an aromatase inhibitor (anastrozole, 1 mg/day for 3 yr) in a boy previously operated on for a hamartoma causing precocious puberty and presenting with advanced bone maturation and nearly fused epiphyseal cartilages, slowed cartilage fusion consenting a higher final stature than expected (164.4 cm vs 158.4 cm). It is suggested that treatment with aromatase inhibitors, alone or in combination with rh-GH, may also be useful in children with constitutional short stature in order to delay epiphyseal closure and improve the final height.

Anastrozole↗
PubMed2000

Multiple epiphyseal dysplasia--report of two families.

Multiple epiphyseal dysplasia (MED) is a relatively uncommon inherited disorder of epiphyseal maturation. Affected individuals may have a degree of short-limbed dwarfism, short stubby digits, and stiff or painful joints. We report two families of MED and emphasize the variations of joint involvement. Ten out of 34 members in family A and 13 out of 39 members in family B were suspected of having MED by questionnaire. Radiological examination was done for 3 out of the 10 members in family A and 6 out of the 13 members in family B. In both families, the epiphyseal disturbances in the skeleton were bilaterally symmetric and involved several joints. Apparent dwarfism, short stubby digits and spinal involvement were not observed. The degree and the pattern of affected joints were different in the two families and even among members of the same family. In family A, the knee joint was commonly affected, followed by the ankle joint. The deformity of the joints was mild and caused only slight disability. No apparent hip lesion was present. In family B, the hip joint was predominantly affected, followed by the knee and ankle joints, and the deformity was severer than that in family A. These observations suggest that MED is a group of heterogeneous disorders.

Adult↗
PubMed2001

Multiple epiphyseal dysplasia: radiographic abnormalities correlated with genotype.

Multiple epiphyseal dysplasia (MED) is an osteochondrodysplasia characterized clinically by mild short stature and early-onset degenerative joint disease and radiographically by epiphyseal hypoplasia/dysplasia. MED is genetically heterogeneous, with autosomal dominant cases resulting from mutations in at least three genes: the cartilage oligomeric matrix protein (COMP) gene (EDM1) and the COL9A2 (EDM2) and COL9A3 (EDM3) genes of type IX procollagen. We present here a comparison of the radiographic phenotypes of MED patients with type IX collagen gene mutations and those with COMP gene mutations. We reviewed radiographs from two patients with MED produced by COMP mutations, two families with COL9A2 mutations, and one family with a mutation in COL9A3. The data demonstrated that the patients with type IX collagen defects had more severe joint involvement at the knees and relative hip sparing, while the patients with COMP mutations had significant involvement at the capital femoral epiphyses and irregular acetabuli. This pattern of joint involvement was consistent regardless of overall degree of severity of the phenotype.

Adolescent↗
PubMed1997

Expression of Bcl-2 protein in the epiphyseal plate cartilage and trabecular bone of growing rats.

The protooncogene protein, Bcl-2, protects cells from apoptosis and ensures their survival in vitro by inhibiting the action of the apoptosis-inducer, Bax. Its expression in proliferative and long-lived cells in vivo also indicates that it protects against cell death. The chondrocytes of the epiphyseal plate cartilage undergo a series of maturation steps and deposit mineral in the cartilage matrix before dying. The possibility that Bcl-2 helps protect chondrocytes until mineral deposition is completed was investigated by determining the distribution of Bcl-2 immunoreactivity in the epiphyseal plate cartilage of growing rats and its subcellular localization, using a specific antibody. The involvement of Bax in the triggering of chondrocyte death was checked by immunocytochemistry. Bcl-2 expression in the osteoblasts and the final result of their evolution, the osteocytes, was also examined in trabecular bone. Bcl-2 immunoreactivity was non-uniformly distributed throughout the epiphyseal cartilage. It was maximal in proliferative chondrocytes, decreased in mature chondrocytes, and low in hypertrophic chondrocytes, whereas there was Bax immunoreactivity in all chondrocytes examined. Immunolabeling was intense in osteoblasts but considerably lower in fully differentiated osteocytes. Bcl-2 immunoreactivity was mainly in the cytoplasm of chondrocytes, osteoblasts, and early osteocytes; the nuclei appeared clear. The subcellular distribution of Bcl-2 immunolabeling in chondrocytes, revealed by gold particles in the electron microscope, showed that gold particles were frequently concentrated in the mitochondria in all the cartilage zones and lay mainly within the organelles, not at their periphery. The endoplasmic reticulum contained moderate immunoreactivity and there were few gold particles in the cytoplasm and nuclei. The number of gold particles decreased in all the subcellular compartments from proliferative to hypertrophic chondrocytes. In contrast, Bax immunoreactivity changed little during chondrocyte terminal evolution, and its subcellular distribution mirrored that of Bcl-2. These immunocytochemical data indicate that Bcl-2 helps maintain chondrocytes and osteoblasts until their terminal maturation.

Animals↗
PubMed2004

Animal model of chondrocyte apoptosis in the epiphyseal cartilage of the neonatal bone.

Apoptosis is considered to be the mechanism responsible for the death of chondrocytes during endochondral bone formation. It is also claimed that apoptosis of the chondrocytes is age related and that the apoptotic index increases with age. However, a detailed analysis of the apoptotic activity of the neonatal epiphyseal cartilage is lacking. A model that evaluates apoptosis in the femoral rat epiphyseal cartilage both quantitatively and qualitatively is reported. Apoptotic incidence in the epiphyseal cartilage reached a maximum at age 6 days, but the age in our study did not significantly affect the percentile rate of apoptotic chondrocytes (P > 0.05, Kruskal-Wallis test). Apoptosis in the zone of hypertrophic cartilage played the most important role in the growth plate's homeostasis. Morphologic evidence of apoptosis was necessary in addition to positive nick end labeling of cells. Electron microscopy studies revealed atypical modes of programmed death of the growth plate chondrocytes in addition to the classical apoptotic mode.

Animals↗
PubMed2006

Intrafamilial phenotypic diversity in multiple epiphyseal dysplasia associated with a COL9A2 mutation (EDM2).

We describe a Japanese family with an autosomal dominant multiple epiphyseal dysplasia (MED EDM2) showing significant phenotypic diversity among the five affected members. Genomic analysis for COL9A2 identified an Ex3-1A>G heterozygous mutation, which has been proved to result in skipping of exon 3. The proband was a 9-year-old boy, who presented with ulnar club hands due to severe epiphyseal dysplasia in the distal ulnae. Radiological examination showed multiple epiphyseal dysplasias, predominantly involving the knee and the wrist. The hip appeared almost normal. The malalignment of the wrist was successfully treated with a limb lengthening procedure. The phenotype of the asymptomatic 12-year-old brother was similar to, but milder than, that of the proband. The asymptomatic 39-year-old mother, the 35-year-old uncle, and the 65-year-old grandmother with bilateral painful knees showed radiographically mild and severe osteoarthritis of the knee, respectively, and none of them had wrist deformity.

Base Sequence↗
PubMed1990

Growth hormone regulation of the growth hormone receptor mRNA in cultured rat epiphyseal chondrocytes.

This study was designed to investigate whether growth hormone (GH) influences the expression of its own receptor in chondrocytes. To investigate this possibility GH-receptor mRNA was measured in cultured rat epiphyseal chondrocytes in the absence or presence of GH under various experimental conditions. Chondrocytes were isolated enzymatically from epiphyseal growth plates of the proximal tibia of 20-day-old male rats and cultured in monolayer in Ham's F-12 medium supplemented with 10% calf serum and 1% of a serum substitute. The cells were seeded at various densities (100,000-1,000,000 cells per flask) and cultured for 14 days. Subsequently, the calf serum-containing medium and the cells cultured for various periods of time (0-24 h) before total nucleic acid preparation. GH-receptor mRNA was measured with a solution hybridization technique using [35S]UTP-labeled RNA growth hormone receptor cloned from rat liver cDNA. Human GH (hGH; 50 ng/ml) increased GH-receptor mRNA after 3 h and maximal levels were seen 12 h after GH addition. This effect of hGH was time and dose dependent with a significant effect of hGH at a concentration of 0.5 ng/ml and a maximal effect at 50 ng/ml. The hGH-stimulated increase of GH-receptor mRNA was completely blocked by actinomycin-C1 (1.0-0.1 micrograms/ml), while cycloheximide (10 micrograms/ml) only slightly counteracted the hGH effect. Rat and human GH were equally potent, and ovine prolactin was effective at 500 ng/ml but not 5 and 50 ng/ml. A high dose of insulin-like growth factor-I (IGF-I; 1 microgram/ml) caused a small stimulatory effect and addition of 10% calf serum caused a marked increase in GH-receptor mRNA. The level of GH receptor mRNA after 14 days of culture was inversely proportional to the cell density at the start of culture. These results show that GH specifically regulates mRNA levels for its own receptor in rat epiphyseal chondrocytes by interacting with somatogenic binding sites. These findings also suggest a transcription-dependent regulatory system between the GH-receptor and the GH-receptor gene.

Animals↗
PubMed1995

Ultrastructural studies of the epiphyseal plate of chicks fed a vitamin D-deficient and low-calcium diet.

The epiphyseal plates of rachitic chicks fed a vitamin D-deficient and low-calcium diet were examined ultrastructurally at 4, 7, 14, 18 and 21 days old. On and after 14 days, changes were observed. Chondrocytes in resting, proliferating and maturing zones commonly showed a reduction in cell organelles, suggestive of a decrease in synthetic activity of matrix components. In addition, the resting zone cells had numerous intracytoplasmic microfilaments. Mitotic figures were present but not increased in number in the proliferating zone. Autolysosome-like dense bodies in the chondrocytes and clusters of degenerative and necrotic chondrocytes were observed in the proliferating and maturing zones. In the calcifying zone of the rachitic epiphyseal plate, initial calcification was observed characterized by the deposition of apatite crystals in matrix vesicles and the formation of spherical crystal clusters. The crystal clusters were separated from each other by collagen fibrils on which only small amounts of apatite crystals had been deposited. In this zone, hypertrophic chondrocytes responsible for the initial calcification were seen, but not the stellate chondrocytes responsible for the progression of matrix calcification after its initiation. These findings suggest that accumulation of the proliferating and maturing zone cells as a characteristic lesion of the epiphyseal plate in rachitic chicks is caused by a disturbance of cell maturation. Such disturbance in the calcifying zone may explain the progression of the defect in matrix calcification.

Animals↗
PubMed1986

Premature epiphyseal closure--a complication of etretinate therapy in children.

Two children are described who developed premature epiphyseal closure while receiving etretinate for treatment of congenital hyperkeratotic disorders. The first patient was an 8 1/2-year-old boy with nonbullous ichthyosiform erythroderma who had been on treatment for 6 years, 4 months when premature fusion of the right distal tibial epiphysis was detected. Shortness of stature, thinning of long bones, and traumatic fractures were also observed in this patient. The second child was an 11-year-old girl with systematized verrucous nevi in whom symmetric fusion of both elbow epiphyses and narrowing of the femoral epiphyses bilaterally were noted following treatment with etretinate for 5 years, 5 months.

Age Determination by Skeleton↗
PubMed2003

Time course of epiphyseal growth plate fusion in rat tibiae.

Although the rat is the most common animal model used in studying osteoporosis, it is often used inappropriately. Osteoporosis is a disease that most commonly occurs in humans long after growth plate fusion with the associated cessation of longitudinal bone growth, but there has been a question as to when or to what extent the rat growth plate fuses. To investigate this question, we used microcomputed X-ray tomography, at voxel resolutions ranging from (5.7 micro m)(3) to (11 micro m)(3), to image the proximal epiphyseal growth plates of both male (n = 19) and female (n = 15) rat tibiae, ranging in age from 2 to 25 months. The three-dimensional images were used to evaluate fusion of the epiphyseal growth plate by quantitating the amount of cancellous bone that has bridged across the growth plate. The results suggest that the time course of fusion of the epiphyseal growth plate follows a sigmoidal pattern, with 10% of the maximum number of bridges having formed by 3.9 months in the male tibiae and 5.8 months in the female tibiae, 50% of the maximum number of bridges having formed by 5.6 months in the male tibiae and 5.9 months in the female tibiae, and 90% of the total maximum of bridges have formed by 7.4 months for the males and 6.5 months for the females. The total volume of bridges per tibia at the age at which the maximum number of bridges per tibia has first formed is 0.99 mm(3)/tibia for the males and 0.40 mm(3)/tibia for the females. After the maximum number of bridges (-290 for females, -360 for males) have formed the total volume of bridges per tibia continues to increase for an additional 7.0 months in the males and 17.0 months for the females until they reach maximum values (-1.5 mm(3)/tibia for the males and -2.2 mm(3)/tibia for the females).

Aging↗