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Journal of the Anatomical Society of India

Regional Differences Of Growth Plate At The Distal End Of Radius In Albino Rat

Author(s): Abhaya A., Khatri K., Pradhan S. and Prakash R.

Vol. 51, No. 2 (2002-07 - 2002-12)

Department of Anatomy, University College of Medical Sciences Shahdra, DELHI

Abstract

Growth plate is important for the longitudinal growth of bone and the sequence of cellular events result in proliferation of chondrocytes, hypertrophy and maturation of cells. Histomorphometric differences in various zones of the growth plate in both central and lateral region at the distal end of radius in 20-day-old albino rat were studied. 6m thick longitudinal sections of decalcified radius were stained with Haematoxylin and Eosin and Masson's Trichrome. The growth plate was differentiated into resting, proliferative, upper hypertrophic and lower hypertrophic zone.

The width (horizontal thickness) of the growth plate increased gradually from diaphyseal to epiphyseal side. It was 1581.00mm ± 54.75 mm at the proximal, 1621.47mm ± 43.55 mm in the middle and 1791.53 mm ± 87.11 at its distal end. The height (vertical thickness) was higher in the lateral (523.76 mm ± 30.14) as compared to central region (474.67 mm ± 52.29 mm). The individual zonal thickness in the lateral region was more in proliferative (265.07 mm ± 41.61 mm) and upper hypertrophic zone (105.60 mm ± 26.56) while it was less in lower hypertrophic zone (153.00 mm ± 24.42 mm) as compared to the central region. The cell population of individual zones was also more in the lateral than the central region.

Key words: Rat , Radius, Growth Plate

Introduction:

Growth plate of a long bone is present between metaphysis and secondary ossification centre of epiphysis and is differentiated into resting, proliferative and hypertrophic zones demonstrating varied cell morphology (Tonna, 1961; Hert, 1972; Walker and Kember, 1972; Kember and Sissons, 1976; Seinsheimer and Sledge, 1981; Buckwalter et al., 1985). It is surrounded by a perichondral ring and persists until the postnatal longitudinal growth of bone is completed. Growth plate increases in width solely as a result of interstitial growth of its marginal parts by the differentiation of perichondral cells into chondrocytes (Tonna, 1961 and Hert, 1972). Regional differences in various zones of growth plate have been established recently by Flores and Baeza (1990) while studying the growth plate of proximal tibia in Albino rats.

The changes in cell size, shape and orientation defining different zones and related cellular functions of the growth plate have been studied in the past extensively however regional differences of this highly specialized and organized structure has received less attention. Therefore the present study was undertaken to observe the histomorphometric changes in different zones in both central and lateral regions of the growth plate at the distal end of radius of 20-day-old albino rat.

Material and Method:

The Albino rats of Wistar stain were maintained under controlled conditions of light and dark cycle (12/12h), temperature and humidity. The mothers were fed with a standard solid chow and both food and water were available ad libitum.

On day 20th, five pups selected at random from different cages, each from separate mother were anaesthetized with ether and sacrificed by decapitation. The distal end of radius including epiphyseal plate was dissected out after removal of all soft tissues and was fixed in 10% formalin saline. The radius was decalcified using neutral EDTA and the end point of decalcification was assessed by using ammonia solution. 6m thick longitudinal paraffin sections were stained with Haematoxylin Eosin and Masson's Trichrome.

The growth plate was divided into the resting zone, proliferative zone and the hypertrophic zone. The hypertrophic zone was further divided into upper hypertrophic zone and lower hypertrophic zone towards metaphysis on the basis of cell morphology as described by Brighten (1984). The central region was in the middle of the growth plate limited by secondary ossification centre of the epiphysis and metaphysis while the lateral region was towards the margin of growth plate near the perichondral ring and contiguous to the marginal germinative zone as distinguished by Flores and Baeza (1990).

The following parameters were recorded.

  1. Width (horizontal thickness) of growth plate - at three different levels (proximal, middle, distal) excluding periosteum.
  2. Height (vertical thickness) of growth plate was the sum of heights of proliferative, upper hypertrophic and lower hypertrophic zone, measured in central and lateral region.
  3. Individual zonal thickness - of proliferative, upper hypertrophic and lower hypertrophic zone in both central and lateral region.
  4. Zonal cell population - in central and lateral region.
  5. Cell columns - in upper hypertrophic zone and lower hypertrophic zone in both central and lateral region.

The results obtained were statistically evaluated.

Observations:

Growth plate of the distal end of radius was seen as a continuous plate not interrupted at any place limited on sides by perichondral ring. It was narrower towards the diaphysis and steadily its width increased distally towards its epiphyseal end (fig. 1). The width was 1581.00mm ± 54.75mm at proximal, 1621.47 mm ± 43.55 mm in the middle and 1791.53 mm ± 87.11 mm at the distal end of growth plate. The proximal margin of growth plate towards the metaphysis was more or less uniform in comparison to its distal margin that varied greatly in shape but whatsoever the shape of its distal margin, it was noted that the height of lateral regions was always more than the central region (Fig. 1). The height was 523.73 mm ± 30.14 mm in the lateral region while it was 474.67 mm ± 52.29 mm in the central region of growth plate (table 1). On the outer surface of perichondrium, dense layer of cells was seen which was continuous with the perichondrium covering the cartilaginous epiphysis.

The growth plate consisted of four successive zones merging with each other. From epiphysis to diaphysis these were (I) resting zone (ii) proliferative zone (iii) upper hypertrophic zone (iv) lower hypertrophic zone (fig. 1, 3).

The resting zone was the most distal zone towards epiphysis. The height of the resting zone appeared less in central region than the lateral region. The reserve cell layer of growth plate formed a wedge like accumulation of cartilage cells near the surface with broader end towards the perichondrium forming the marginal germinative zone in which the chondrocytes did not show any particular orientation (fig. 2). The chondrocytes of the resting zone were small, rounded or spindle shaped seen both within and without lacuna and were distributed singly, double or in small groups. The cell groups were more in the lateral than in the central region.

Next to the resting zone was the proliferative zone, in which the discoidal or cuneiform chondrocytes lying in lacunae were slightly increased in size and arranged in longitudinal columns (fig. 3). In the central region, the longitudinal columns of cells were straight and were lying close to each other and comprised of more number of cells in each column (fig. 3) as compared to lateral region where the columns were disposed obliquely and therefore number of columns in this zone were not recorded (fig. 2). Although the cell population was marginally increased in lateral region (61.17 ± 2.99) as compared to central region (59.00 ± 3.58), but the zonal thickness was remarkably higher in lateral region (265.07 mm ± 41.61 mm) in comparison to central region (199.27 mm ± 42.22 mm). table (I, II).

Table I:

Parameters Central region Lateral region
1. Height of Growth Plate (m) 474.67 ± 52.29 523.73 ± 30.14
2. Individual Zonal Thickness (m)
(i) Proliferative zone 199.27 ± 42.22 265.07 ± 41.61
(ii) Upper hypertrophic zone 93.00 ± 13.52 105.60 ± 26.56
(iii) Lower hypertrophic zone 182.20 ± 23.54 153.00 ± 24.42

Table - II

Parameters Central region Lateral region
1. Zonal Cell Population
(i) Proliferative zone 59.00 ± 3.58 61.17 ± 2.99
(ii) Upper hypertrophic zone 34.83 ± 4.12 39.33 ± 5.13
(iii) Lower hypertrophic zone 23.67 ± 3.39 28.33 ± 3.06
2. Cell Columns    
(i) Upper hypertrophic zone 6.50 ± 1.05 7.00 ± 1.10
(ii) Lower hypertrophic zone 5.17 ± 1.17 5.00 ± 1.26

In upper hypertrophic zone, there was gradual increase in the size of the lacunae around the chondrocytes arranged in columns. The cell outline of the chondrocytes was slightly distorted and nuclei became swollen and compressed at places (fig. 3) The zonal thickness was 105.60 mm ± 26.56 mm in the lateral region and 93.00 mm ± 13.52 mm in the central region (table I). The cell population in lateral region was also more (39.33 ± 5.13) while it was less in central region (34.83 ± 4.12) (table II). The number of cell columns was also slightly higher in lateral egion (7.00 ± 1.10) than in central region (6.50 ± 1.05) (table II).

The upper hypertrophic zone was followed by lower hypertrophic zone where the lacunae had greatly increased in size with irregular margins and acquired a round to polygonal shape enclosing a chondrocyte, which had reduced in size. The cytoplasm became vacuolated and nuclei became pyknotic. Some lacunae showed complete degeneration of chondrocytes. The lacunae were separated from each other by thin septa and from other columns of hypertrophic cells by a thin bar of matrix (fig. 3). The columnar arrangement was still maintained. The zonal thickness was less in lateral region (153.00 mm ± 24.42 mm) while it was 182.20 mm ± 23.54 mm in the central region of growth plate (table I). The cell population in the lateral region (28.33 ± 3.06) was slightly higher than the central (23.67 ± 3.39) region and the number of cell columns also showed a marginal variation (table II).

The matrix appeared homogenous throughout the growth plate except near the epiphyseal end, and perichondrium where it was stained more deeply. The matrix of resting zone also stained more deeply in comparison to other zones of growth plate. The staining was deeper in central region than in the lateral region.

Discussion:

Histologically, the growth plate was seen as a continuous plate and was not interrupted at any point as described earlier by Dawson (1925). It consisted of four successive zones i.e. resting, proliferative, upper hypertrophic and lower hypertrophic zone, merging with one another from epiphysis to diaphysis.

The height of growth plate was found more in lateral region as compared to the central region. The individual zonal thickness of lateral region showed a marked increase in the thickness of proliferative and upper hypertrophic zone but this difference was mainly contributed by the proliferative zone (table I). Flores and Baeza (1990) had also observed increase in the height of lateral region which was only contributed by the proliferative zone. The explanation for their observation may be that they had not subdivided the hypertrophic zone as has been done in the present study. In the lateral region, the height of upper hypertrophic zone had increased while that of the lower hypertrophic zone had decreased (table I), the sum total height of both these hypertrophic zones of the lateral region may be having marginal difference in comparison to the corresponding sum total height of the central region.

The cell population was higher in all the zones of the lateral region as compared to the central region and this difference was more marked in upper hypertrophic zone (table II). The maximum cell density was observed in the proliferative zone in both the regions and similar observations have been reported by Buckwalter et al (1985), Walker and Kember (1972) and Flores and Baeza (1990). Flores and Baeza (1990) suggested that the presence of more cells in lateral region especially in proliferative zone could be attributed to their smaller size and absence of secondary ossification centre at this level. Marginal difference was observed in the number of cell columns of the hypertrophic zones in central and lateral region whereas Flores and Baeza (1990) have reported a definite increase in cell columns of proliferative and hypertrophic zones in the lateral region.

In the present study it appeared that although the height of proliferative zone in lateral region had increased but the increase in number of cells was not so significant whereas in the upper hypertrophic zone in which the height was only marginally increased, the cell population showed a marked rise. Since the size of cells in different zones and different regions had not been recorded so it was difficult to comment whether the size of cells in these two zones had altered or the matrix cell volume had changed.

The present observation shows that the central and the lateral regions of the growth plate are quantitatively different and suggest that in histomorphometric analysis of the growth plate it may be necessary to specify which of these two regions are undertaken for study.

References:

  1. Brighton C.T. (1984): The growth plate. Orthopaedics Clinics of North America Journal. 57: 595.
  2. Buckwalter J.A. et al. (1985): Growth plate chondrocyte profile and their orientation. Journal of Bone & Joint Surgery 67A: 942-955.
  3. Dawson A.B. (1925): The age order of epiphyseal union in the long bones of the albino rat Anatomical Record 31: 1-18.
  4. Flores C.M. and Baeza E.D. (1990): Histomorphometric differences between the lateral region and central region of the growth plate in fifteen-day-old rats. Acta Anatomica 139: 209-213.
  5. Hert J. (1972): Growth of the epiphyseal plate in circumference. Acta Anatomica 82: 420-436.
  6. Kember N.F. and Sissons H.A. (1976): Quantitative histology of the human growth plate. Journal of Bone & Joint Surgery 58B: 426-435.
  7. Seinsheimer F. and Sledge C.B. (1981): Parameters of longitudinal growth rate in rabbit epiphyseal growth plates. Journal of Bone & Joint Surgery 63A: 627-630.
  8. Tonna E.A. (1961): The cellular complement of the skeletal system studied autoradiographically with tritiated thymidine (H3 TDR) during growth and aging. Journal of Biophysics, Biomedicine & Cytology. 9 : 813
  9. Walker K.V.R. and Kember N.F. (1972): Cell kinetics of growth cartilage in the rat tibia II Measurements during aging. Cell and Tissue Kinetics. 5: 409-419.

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Fig. 1: Photomicrograph of growth plate showing the central region (C), lateral region (L), marginal germinative zone (MGZ), epiphyseal end (EP) and metaphyseal end (MP). Growth plate is divided into resting (R) proliferative (P) and hypertrophic zone (HT). Haematoxylin Eosin X 63.

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Fig. 2: Photomicrograph of growth plate showing the obliquely placed cell columns of proliferative zone (P) in the lateral region. The resting zone (R) continuing with the marginal germinative zone (MGZ). Masson's Trichrome X100.

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Fig. 3: Photomicrograph showing the subdivisions of hypertrophic zone into upper hypertrophic zone (HT1) and lower hypertrophic zone (HT-II). Masson's Trichrome X100.

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