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At least 505 records · Page 28Linked to original sources
PubMed2003

Treatment of displaced, proximal, humeral, epiphyseal fractures with a two-prong splint.

To evaluate the use of a two-prong splint for displaced, proximal, humerus, epiphyseal fractures, seven children with Neer-Horwitz Classification Type 3-4 fractures of the proximal humerus were retrospectively investigated. After closed reduction, fracture stabilization was accomplished with the use of a two-prong splint, and at an average follow-up of 54 months (range 48-62 months), all children were evaluated radiographically and functionally. There were no complications, and all patients had full painless range of shoulder and elbow motion, without malunion, joint incongruity, nonunion, avascular necrosis, and limb-length discrepancy. The results did not change over time. Treatment of displaced, proximal, humeral, epiphyseal fractures with a two-prong splint gives satisfactory results while allowing immediate motion and gradual reduction of the fragments without anesthesia, and it may be the treatment of choice for such injuries in children.

Adolescent↗
PubMed1984

Proximal tibial epiphyseal fracture in an infant.

Proximal tibial epiphyseal fractures are rare injuries occurring predominantly in older children and adolescents. They have not previously been reported in infants. The present case is that of a 7-month-old female infant with a displaced Salter-Harris type II fracture secondary to child abuse. This represents the youngest documented case of a proximal tibial epiphyseal fracture. Open reduction and internal fixation were performed because of failure to obtain an anatomic alignment by closed reduction. The anatomic reason for unsuccessful closed reduction intraoperatively was found to be an infolding of the pes anserinus and periosteum. Tibia valga occurred postoperatively because of asymmetrical physeal overgrowth and required corrective osteotomy.

Child Abuse↗
PubMed1984

Sternoclavicular epiphyseal separation with adjacent clavicular fracture.

An unusual case of a double fracture of the medial left clavicle in a 15-year-old male hockey player is presented. One fracture was an epiphyseal separation through the medial growth plate and the second a transverse fracture of the medial one-third of the same clavicle with displacement of the fragment into the chest. Radiographic and computed tomographic descriptions are presented. The injury was managed with open reduction and fixation. A discussion of sternoclavicular epiphyseal separations and intrathoracic displacement is given, with a review of the literature.

Adolescent↗
PubMed1997

Multiple epiphyseal dysplasia and pseudoachondroplasia due to novel mutations in the calmodulin-like repeats of cartilage oligomeric matrix protein.

A child with a mild form of pseudoachondroplasia was heterozygous for a deletion of 12 nucleotides from exon 10 of the cartilage oligomeric matrix protein (COMP) gene. It resulted in the deletion of valine 513 to lysine 516 from the eighth calmodulin-like repeat of COMP monomers. A child with the Fairbank's type of multiple epiphyseal dysplasia was also heterozygous for a COMP mutation. It substituted cysteine 371 by serine in the fourth calmodulin-like repeat. Both mutations were likely to alter the conformation and calcium binding of the mutant COMP protein chains. These findings support the proposal that deletions and insertions within the calmodulin-like domain produce pseudoachondroplasia, while amino acid substitutions with this domain may produce either pseudoachondroplasia or multiple epiphyseal dysplasia.

Achondroplasia↗
PubMed1994

Radiological aspects on the course of development of porcine epiphyseal osteochondrosis (OCD) from 42 up to 147 days of age.

The articular-epiphyseal (A-E) cartilage complex of the distal humeral and femoral epiphyseal condyles of eight intact pigs (4 male & 4 female) of the same age were radiologically examined every 3 weeks, beginning at 42 days up to 147 days of age; to determine the age of onset, the course and trend of development of osteochondrosis (OCD). The earliest Dyschondroplasia (Osteochondrotic) lesions were demonstrated radiologically in the A-E complexes of the humeral condyles of 42-day-old pigs and in the femoral condyles at 63 days of age. Thus the radiographic examination of live animals to demonstrate subchondral radiolucency typical for OCD lesions in the condylar A-E complexes of pigs is not of limited value until the animals were > 100 days old as indicated in earlier reports. Also the course of further development of OCD lesions associated with A-E complexes was followed. The radiolucency was seen to develop to a certain extent, and then either to regress, and stabilize or even to progress as the animal grows. Some lesions regressed completely. Also some of the regressed lesions may become active again and become progressive. However, the course of development of femoral condyle A-E complexes OCD lesions was seen to be progressive continuously, or progressive then stable and then progressive again. The regressive course and trend of osteochondrotic lesions was not observed in the femoral condyle A-E complexes up to day 147 of age. Therefore, the course and trend of development of the A-E complexes OCD is not constantly the same.

Animals↗
PubMed2007

X-linked spondylo-epiphyseal dysplasia tarda in the Danish-Swedish farm hound.

In two litters from the same parents, three out of four males had an abnormally short leg and body length. Affected dogs showed signs of pain when moving, which could be eliminated by analgesia. On radiography, these animals had widened, radiolucent, irregularly bordered intervertebral disc spaces. When examined at seven months of age, the epiphyses appeared widened and irregular in shape and outline. General bone opacity in the vertebral column was lower in the affected male dogs than in the normal littermate. The affected dogs developed spondylosis and arthrosis of the larger limb joints. All affected dogs were euthanased on humane grounds, the eldest at the age of two years nine months. Based on the clinical and radiographic evidence, the condition seen in the male dogs described here resembles X-linked spondylo-epiphyseal dysplasia tarda caused by a collagenopathy due to malformation of COL2A1 as seen in human beings.

Animals↗
PubMed1977

Perthes' disease and multiple epiphyseal dysplasia.

Five atypical cases were observed amongst ninety children with Perthes' disease, ten of whom had bilateral hip joint involvement. All five were boys, four being under 4 years of age. Four had bilateral hip joint disease, four presented with hip pain, three showing some degree of retardation of bone growth. In one case the hip disorder was familial, and in four there were bony abnormalities elsewhere. Despite the absence of the classic signs of multiple epiphyseal dysplasia, a mild form of this condition is a possible alternative diagnosis for these children. Racial and familial differences are known in the prevalence of Perthes' disease which itself may represent a dysplasia. The pathogenesis of Perthes' disease is still uncertain, although some abnormality of the blood supply to the proximal femoral epiphysis is postulated. That such a vascular defect may be engrafted on to multiple epiphyseal dysplasia is possible, with subsequent joint degeneration which may come to resemble Perthes' disease either clinically or radiologically. A plea is made for the closer study of young children presenting with what may seem to be atypical Perthes' disease.

Child↗
PubMed1996

Biomechanical reasons for the divergent morphology of the knee joint and the distal epiphyseal suture in hominoids.

The obliquity of the femoral diaphysis accounts for the valgus position of the human knee joint and reduces bending moments in the frontal plane. A high angle of obliquity is considered a hallmark of hominid bipedality, but its functional importance has rarely been identified correctly. A biostatic investigation of the knee joint in various realistic positions unveils resultant joint forces which do not deviate greatly from the long axis of the femoral shaft. This is due to the length of the femur and to the shortness of the human foot. The flat epiphyseal suture is more or less perpendicular to these joint forces, and the equal size of the femoral condyles reflects the even distribution of forces between them. In great apes the resultant forces acting in the knee joint vary considerably in dependence on the degree of flexion and rotation of the knee joint. The resultant joint force may be line with the femur shaft or diverge. The epiphyseal surfaces offer facets to all joint forces found in the course of the study. Due to the pronounced varus position of the knee joint, the joint itself and the adjacent part of the femur are under medially concave bending moments, which lead to higher compressive forces at the medial than at the lateral condyle. The enlarged medial condyle allows the distribution of medially displaced joint forces over a relatively large area, and the elliptic cross-section yields high bending resistance in the frontal plane. A human-like angle of obliquity is present in the early australopithecines, the values being mostly within the range of variation of children. The valgus position of the australopithecine knee joint is considered to be a functional, and epigenetic consequence of habitual bipedality. It is particularly pronounced because of the short length of the femur and the great bitrochanteric width.

Animals↗
PubMed1995

Ultrastructural localization of collagen types II, IX, and XI in the growth plate of human rib and fetal bovine epiphyseal cartilage: type XI collagen is restricted to thin fibrils.

The collagen fibrils of hyaline cartilage vary in diameter depending on developmental stage and location within the tissue. In general, growth plates and fetal epiphyseal cartilages contain fibrils with diameters of less than approximately 25 nm, whereas the permanent cartilage of adult tissues contains fibrils of approximately 30-200 nm. The interstitial collagen fibrils of fetal cartilage are complex, having at least three collagen types as integral components. Type XI, a member of the fibrillar collagen class, has been proposed to limit fibril diameter. To test this proposition we sought to determine if Type XI collagen was preferentially associated with fibrils of smaller diameter. We focused our study on human juvenile rib growth plate, which has thin fibrils in the hypertrophic zone, thick fibrils in the resting zone or permanent cartilage, and a mixture of thin and thick fibrils in the proliferative zone. Tissues were examined by immunoelectron microscopy with antipeptide antibodies to the carboxyl telopeptide and to the amino terminal non-triple-helical domains of alpha 1 (XI). These studies showed that (a) both epitopes of Type XI collagen were readily accessible to antibodies at the fibrillar surface, (b) Type XI collagen was associated predominantly with fibrils < 25 nm in diameter, (c) Type XI collagen was not found in thick fibrils even after disruption with chaotropic agents, and (d) collagen Types II and IX were associated with fibrils of all sizes. These studies were extended to human newborn epiphyseal cartilage and to fetal calf cartilage, with the same result.

Amino Acid Sequence↗
PubMed1987

Growth hormone in vivo potentiates the stimulatory effect of insulin-like growth factor-1 in vitro on colony formation of epiphyseal chondrocytes isolated from hypophysectomized rats.

The effect of GH pretreatment in vivo on the colony formation of epiphyseal chondrocytes from hypophysectomized rats and the subsequent responsiveness to insulin-like growth factor (IGF-I) was studied in vitro. Chondrocytes from epiphyseal growth plates of the proximal tibia of 36-day-old hypophysectomized rats were enzymatically isolated and cultured in suspension, stabilized with agarose (0.5%) in Ham's F-12 medium and serum supplement. After 14 days the cultures were terminated and screened for cloning efficiency (number of colonies with a diameter greater than 56 microns/1000 seeded cells) and for distribution of cloning efficiency as a function of colony size. Pretreatment with human GH in vivo for 24 h (10 micrograms X 3) increased the cloning efficiency during the subsequent culture period (control, 1.5 +/- 0.1; human GH, 4.4 +/- 0.3). Addition of IGF-I to the chondrocyte cultures from control rats caused a slight increase in cloning efficiency (control, 1.5 +/- 0.1; IGF-I, 2.2 +/- 0.3) but caused a marked increase in chondrocyte cultures from GH-pretreated rats (control, 4.4 +/- 0.4; IGF-I, 8.2 +/- 0.9). The cloning efficiency was increased 12 and 24 h, but not 4 h, after start of GH-treatment in vivo. The increased responsiveness to IGF-I in vivo showed a similar time course after GH pretreatment. The distribution of cloning efficiency was altered in cultures of chondrocytes isolated from the GH-pretreated rats; large colonies were overrepresented in the GH-treated group. Colonies with a diameter exceeding 180 microns were only seen in cultures of chondrocytes isolated from the GH-pretreated animals. Addition of IGF-I in vitro did not alter the distribution of cloning efficiency, but increased the mean colony size of all colonies. Pretreatment of the rats with two different doses of IGF-I in vivo for 24 h (5 micrograms X 3 or 50 micrograms X 3) had a slight stimulatory effect on subsequent colony formation, but no potentiation of IGF-I in vitro was demonstrated. The results of the present study show that pretreatment of hypophysectomized rats with GH, but not with IGF-I, promotes the formation of chondrocyte colonies and make the chondrocytes susceptible to IGF-I in vitro. The results suggest that GH induces colony formation by IGF-I-independent mechanisms and that IGF-I is a second effector in GH action as previously shown for cultured 3T3-preadipose cells.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1988

The effect of parathyroid hormone and atrial natriuretic peptide on cyclic nucleotides production and proliferation of avian epiphyseal growth plate chondroprogenitor cells.

Cells derived from avian tibia epiphyseal growth plate were cultured in vitro. The cells which exhibited a polygonal phenotype and are termed chondroprogenitor cells, developed in culture as a monolayer with a doubling time of 40-48 h in 5% fetal calf serum. Production of cAMP by the chondroprogenitor cells was stimulated by human and bovine native (1-84) PTH. The effect of PTH on cAMP production could be blocked by the (3-34) PTH analog, suggesting interaction with specific receptors. cAMP production by avian chondroprogenitor cells was also stimulated by cholera toxin, forskolin, and prostaglandin E2 but not by ACTH or prostaglandin F2 alpha. PTH, cholera toxin, and forskolin also stimulated proliferation of the chondroprogenitor cells. In contrast, neither cAMP production nor proliferation of avian skin fibroblasts was affected by PTH. Human (1-28) and rat (5-28) atrial natriuretic peptide stimulated cGMP production by avian chondroprogenitor cells and also by skin fibroblasts. Atrial natriuretic peptide inhibited the basal and PTH-stimulated [3H]thymidine incorporation into DNA of chondroprogenitor cells, but did not affect avian skin fibroblast proliferation. These results suggest that the proliferation of avian epiphyseal growth plate chondroprogenitor cells is modulated by opposing mechanisms induced by PTH and ANP, probably mediated by cAMP and cGMP, respectively.

1-Methyl-3-isobutylxanthine↗
PubMed1990

Insulin is a mitogen for isolated epiphyseal growth plate chondrocytes from the fetal lamb.

Insulin acts directly as a mitogen on isolated embryonic tissues but has been shown to be inactive at physiological concentrations on several fetal cell types. Since insulin availability is obligatory for optimal fetal growth, we have investigated its mitogenic actions on chondrocytes isolated from the epiphyseal growth plates of fetal lambs. Chondrocytes were isolated from the proximal tibial growth plate of lamb fetuses between 40 and 130 days gestation using collagenase and were cultured in monolayer before use between passages 2 and 6. The synthesis of DNA was assessed from the incorporation of [3H]thymidine after incubation in medium supplemented with glucose (0.7 mM-25 mM) with or without insulin (0.08 nM-167 nM). Increase in cell number was assessed after incubation with test medium for up to 8 days. Insulin substantially increased both DNA synthesis, and cell number, compared to control incubations with a biphasic dose response; an initial 3- to 5-fold increase in DNA synthesis occurring at approximately 1 nM insulin with a second response seen at approximately 50 nM. Within the physiological range of concentrations insulin was only 50% as active as insulin-like growth factor I (IGF I), but was 15 times more active than IGF II. Similar effects of insulin were observed throughout the fetal age range, although the DNA synthetic rate in basal medium declined with both fetal age and cell passage number. The mitogenic actions of insulin were glucose-dependent and were maximal in the presence of 2.7 mM glucose. Insulin did not cause any change in chondrocyte cell cycle duration. Chondrocytes released immunoreactive IGF II but no detectable IGF I. While exposure to insulin concentrations of approximately 50 nM or greater resulted in a statistically significant increase in IGF II release from chondrocytes, no change in IGF II release was seen in response to physiological insulin concentrations. However, exposure of cells to a blocking monoclonal antibody against human IGF I, McAb SM 1.2, which also negates the mitogenic actions of IGF II, consistently reduced insulin-stimulated DNA synthesis suggesting that IGF II presence may be necessary for optimal insulin action. Combination experiments using maximal concentrations of IGF I (13.3 nM) and increasing amounts of insulin (0.16 nM-1.67 nM) showed additive effects on DNA synthesis, suggesting that each hormone was acting through distinct receptor populations. We conclude that insulin, at physiological concentrations, may exert direct growth-promoting actions at the epiphyseal growth plates of the fetal lamb throughout gestation.

Animals↗
PubMed1990

Biological effects of androgens and identification of specific dihydrotestosterone-binding sites in cultured human fetal epiphyseal chondrocytes.

The biological effects of dihydrotestosterone (DHT) and testosterone (T) on cultured human fetal epiphyseal chondrocytes were assessed by studying the ability of these androgens to promote DNA synthesis. DNA synthesis was evaluated by measuring [3H]thymidine incorporation into DNA. After 48-h incubation in Ham's F-12 serum-free medium, chrondrocytes were incubated with or without DHT (10(-11)-10(-8) M) or T (10(-11)-10(-8) M) in MCDB-104 serum-free medium for a further 48 h, with the addition of [3H]thymidine (5 microCi/mL) for the last 24 h. In chondrocytes from five male fetuses (12-40 weeks' gestation) DHT and T significantly stimulated DNA synthesis. The maximum stimulatory effect was obtained for DHT at 10(-10) M (P less than 0.01) and for T at 10(-6) M (P less than 0.02). In chondrocytes from four female fetuses the stimulatory effect was significant only for DHT and was maximum at 10(-10) M (P less than 0.02), whereas no effect was observed for T. Cultured chondrocytes from both male and female fetuses show the presence of proteins with high affinity and limited binding capacity (Bmax) for DHT (male fetuses: Bmax, 4.9 +/- 1.9 x 10(-15) M/mg protein; Kd, 0.43 +/- 0.24 x 10(-9) M; female fetuses: Bmax, 4.8 +/- 1.6 x 10(-15) M/mg protein; Kd, 0.63 +/- 0.19 x 10(-9) M) with no significant differences between sexes. In conclusion, our results show that androgens elicit a biological response in cultured human fetal epiphyseal chondrocytes and that DHT-binding sites are present in these cells. DHT, rather than T, seems to be the active androgen. A sex difference in the degree of androgen action is also documented.

Androgen-Binding Protein↗
PubMed2003

HIF-1alpha controls extracellular matrix synthesis by epiphyseal chondrocytes.

The transcription factor HIF-1alpha plays a crucial role in modifying gene expression during low oxygen tension. In a previous study, we demonstrated that HIF-1alpha is essential for chondrocyte growth arrest and survival in vivo. To explore further the role of HIF-1alpha in cartilage biology, we undertook studies with primary epiphyseal chondrocytes with a targeted deletion of HIF-1alpha. In this study, we show that HIF-1alpha is necessary for regulating glycolysis under aerobic and anaerobic conditions. HIF-1alpha-null chondrocytes were unable to maintain ATP levels in hypoxic microenvironments, indicating a fundamental requirement for this factor for the regulation of chondrocyte metabolism. Synthesis of the angiogenic factor vascular endothelial growth factor was also significantly induced by hypoxia, and this increase is lost in HIF-1alpha-null mutant cells. Under hypoxic conditions, aggrecan mRNA and protein levels were significantly reduced in chondrocytes lacking the HIF-1alpha transcription factor. Interestingly, strongly increased type-II collagen protein levels were detected in wild-type cells after 44 hours of hypoxia. In addition, type-II collagen mRNA and protein levels were strongly decreased under low oxygen in chondrocytes lacking HIF-1alpha. In summary, our results clearly demonstrate the importance of HIF-1alpha in maintenance of anaerobic glycolysis, and thereby extracellular matrix synthesis, of epiphyseal chondrocytes.

Adenosine Triphosphate↗
PubMed1998

Local disappearance of epiphyseal growth plates in rats with hypervitaminosis A.

Epiphyseal growth plates of proximal tibiae in rats with high doses of vitamin A (V-A) were observed. Of 4 groups, each consisting of 5 rats, three groups were given V-A at doses, IU/100 g by body weight/day, of 50,000, 100,000, and 150,000, respectively. The other group rats were given no V-A (control). Rats were administered V-A for the 5 days from 4 weeks after birth and sacrificed at 12 weeks after birth. Three rats of the 150,000 IU group died during the period of observation. The decalcified sections were stained with hematoxylin-eosin or toluidine blue. In the ground sections, microradiography, backscattered electron imaging, and energy-dispersive X-ray microanalysis were performed. These observations suggest that the local disappearance of epiphyseal growth plates under high doses of V-A goes in the order of the increased doses through the process of (1) calcified cartilage areae appearing in the resting cell zone, (2) some of the calcified areae extending in the growth plate towards the diaphysial side, (3) bone tissue replacing the calcified areae, and (4) the local disappearing of the growth plate. Such a local disappearance may be formed in the stressed proximal regions of tibiae.

Animals↗
PubMed1991

The shoulder in multiple epiphyseal dysplasia.

One hundred shoulders were assessed in 50 patients with multiple epiphyseal dysplasia, of which one-third were found to have bilateral symptoms. Two distinct clinical and radiological groups emerged. Those with minor epiphyseal abnormalities developed painful osteoarthritis in middle age, but retained shoulder movement until the secondary changes were advanced. Those with 'hatchet head' shoulders likewise became painful in the fifth and sixth decades but had minimal glenohumeral movement from an early stage.

Adolescent↗
PubMed1987

Effects of growth hormone and insulin-like growth factor-I on colony formation of rabbit epiphyseal chondrocytes at different stages of maturation.

The effect of human GH (hGH) and insulin-like growth factor-I (IGF-I) on colony formation of rabbit epiphyseal tibial chondrocytes at different stages of maturation was studied in suspension culture. The epiphyseal growth plate from the proximal tibia of 8-week-old male rabbits was dissected and divided into three different (proximal, intermediate and distal) zones, each zone representing an enrichment of chondrocytes from the germinative, proliferative and hypertrophic cell layers respectively. Chondrocytes from these three zones were isolated by collagenase digestion and cultured in the presence of 10% (v/v) newborn calf serum in suspension stabilized with 0.5% (w/v) agarose for 14 days. The colonies were classified according to diameter and the number of colonies was estimated as a function of colony size (distribution of cloning efficiency). Cell clusters with a diameter of 56 micron or more were classified as colonies. The cloning efficiency (number of colonies formed per 1000 seeded cells) was 10.1 +/- 0.7 and 6.0 +/- 0.8 for chondrocytes isolated from the proximal and intermediate zones respectively. Insulin-like growth factor-I (25-200 ng/ml) increased the number of colonies in chondrocytes isolated from the proximal zone (122 +/- 9.0-156 +/- 8.4%; control value, 100%) and from the intermediate zone (136 +/- 14.0-191 +/- 29%). Low concentrations of hGH (10-40 ng/ml) stimulated colony formation in chondrocytes isolated from the proximal zone (125 +/- 6.4-137 +/- 7.9%) whereas a high concentration of hGH (160 ng/ml) was ineffective. Chondrocytes isolated from the intermediate zone showed no response to low concentrations of hGH (10-20 ng/ml).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Effect of growth hormone and insulin-like growth factor-I on DNA synthesis and matrix production in rat epiphyseal chondrocytes in monolayer culture.

The influence of various culture conditions was studied on the effect of GH and insulin-like growth factor-I (IGF-I) on DNA and matrix synthesis in epiphyseal rat chondrocytes in monolayer culture. Chondrocytes from enzymatically digested rat tibia epiphyseal growth plates were seeded in 48-well culture plates and precultured for 10 days in Ham's F-12 medium supplemented with 1% (v/v) newborn calf serum and 1% (v/v) of a serum substitute. After preculture, the medium was changed to Ham's F-12 medium supplemented with 1% serum from hypophysectomized rats, and the effect of GH and IGF-I on DNA synthesis ([3H]thymidine incorporation) and matrix production ([35S]sulphate uptake) was studied during an additional 96-h culture period. Isotopes were present during the last 24 h of culture. Both hGH and IGF-I stimulated DNA synthesis in a dose-dependent manner. A maximal effect of GH was seen at a concentration of 25 micrograms/l (60 +/- 11% stimulation over control) and for IGF-I at 10 micrograms/l (162 +/- 12%). The stimulatory effects of the same concentrations of human GH (hGH) and IGF-I on [35S]sulphate uptake were 135 +/- 25 and 320 +/- 42% respectively. In-vitro pulse labelling revealed that GH did not produce a response during the first 3 days of culture (after addition of GH) but was effective during days 4 and 5 of culture. In contrast, IGF-I was effective throughout the culture period. Pretreatment of cells with GH or IGF-I for 2.5 days showed that GH but not IGF-I produced a sustained effect on [3H]thymidine uptake. In order to study the influence of cell density on the effect of GH and IGF-I on DNA synthesis, the effect of added peptides was evaluated after different preculture periods (5-15 days). A maximal stimulatory effect of hGH was seen at a cell density of 150,000-300,000 cells/cm2. GH had no significant effect at a low (less than 100,000 cells/cm2) or a high (greater than 400,000 cells/cm2) cell density. The magnitude of the stimulatory effect of IGF-I was the same at densities between 10,000 and 250,000 cells/cm2, but was reduced at higher cell densities (over 250,000 cells/cm2). Chondrogenic properties of cells that had been cultured for 15 days were verified in vitro by positive alcian blue staining and identification of type II collagen, and in vivo by development of cartilage nodules in nude mice.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗