SciELO - Scientific Electronic Library Online

 
vol.29 número4Morfología del Arco Maxilar y Mandibular en Niños de Ascendencia Mapuche y no MapucheAlgoritmo Matemático para Cuantificar Directamente el Ángulo de Convergencia en Preparaciones de Dientes Artificiales índice de autoresíndice de materiabúsqueda de artículos
Home Pagelista alfabética de revistas  

Servicios Personalizados

Revista

Articulo

Indicadores

Links relacionados

Compartir


International Journal of Morphology

versión On-line ISSN 0717-9502

Int. J. Morphol. vol.29 no.4 Temuco dic. 2011

http://dx.doi.org/10.4067/S0717-95022011000400006 

Int. J. Morphol., 29(4):1109-1114, 2011.

 

Histogenesis of Testicular Parenchyma During Prenatal Life in Buffalo

 

Histogénesis del Parénquima Testicular Durante la Vida Intrauterina en el Búfalo

 

Manjinder Kaur; Neelam Bansal & Varinder Uppal

Department of Veterinary Anatomy & Histology, Guru Angad Dev Veterinary and Animal Sciences University, Ludhiana, Punjab, India.

Correspondence to:


SUMMARY: The study was conducted on the testes of 18 buffalo foetii to reveal histogenesis and differentiation of different cells of testicular parenchyma. At 8.0 cm CVR (65 days) the seminiferous tubules were present at gonadal periphery and a network of polygonal mesenchymal cells was seen in the centre of testis. These tubules were surrounded by a distinct basement membrane and a single layer of peritubular cells at 10 cm CVR (74 days), which became double layered at 88.0 cm CVR (272 days). The testicular parenchyma at 12.0 cm CVR had two zones; outer zone having longitudinal tubules and inner zone having rounded tubules. But a reverse pattern of their arrangement was observed at 14.0 cm CVR (92 days). The pre-Sertoli cells were first observed in buffalo foetii of 8.0 cm CVR (65 days) in the periphery of seminiferous tubular epithelium whereas the gonocytes were demonstrable in the centre of tubules at 10.6 cm CVR (76 days). The fetal Leydig cells were also reported at 8.0 cm CVR (65 days) but at 14.0 cm CVR (92 days), the interstitium had considerably expanded due to the differentiation of mesenchymal cells into the Leydig cells.

KEY WORDS: Histogenesis; Testicular parenchyma; Prenatal life; Buffalo.


RESUMEN: El estudio fue realizado en los testículos de 18 fetos de búfalos, para revelar la histogénesis y diferenciación de las diferentes células de parénquima testicular. A los 8,0 cm de longitud corona-rabadilla (LCR) (65 días) los túbulos seminíferos estuvieron presentes en la periferia de la gónada y una red poligonal de células mesenquimales se observó en el centro del testículo. Estos túbulos estaban rodeados por una membrana basal y una sola capa de células peritubular a los 10 cm LCR (74 días), la cual se convirtió en una doble capa a los 88,0 cm LCR (272 días). El parénquima testicular a 12,0 cm LCR tenía dos zonas, zona exterior con túbulos longitudinales y zona interior con los túbulos redondeados transversalmente. Sin embargo, un patrón inverso en su disposición se observó a los 14,0 cm LCR (92 días). Las células pre-Sertoli se observaron primero en fetos de búfalos de 8,0 cm LCR (65 días) en la periferia del epitelio seminífero tubular, mientras que los gonocitos fueron visibles en el centro de los túbulos a 10,6 cm LCR (76 días). Las células de Leydig fetales también se observaron a los 8,0 cm LCR (65 días), pero a los 14,0 cm LCR (92 días), el intersticio tuvo una considerable expansión debido a la diferenciación de células mesenquimales en células de Leydig.

PALABRAS CLAVE: Histogénesis; Parénquima testicular; Vida prenatal; Buffalo.


 

INTRODUCTION

The differentiation of testes is a complex process and involves a number of mechanisms at various stages of development. Any disturbance in the development of the organ during critical period of organization can lead to low reproductive efficiency during the adult life. In literature reports are available on the histogenesis and morphogenesis of testes in sheep (Hochereau de Riviers et al., 1995), goat (Tekade, 2001), cattle (Wrobel, 2000; Abd-Elmaksoud, 2005) but scanty information is available in the buffalo. So the present study was conducted to understand the normal developmental process of testes and to evaluate the unrecognized defects in the histogenesis of the organ which can hamper the reproductive efficiency during adult life and data obtained can be used to improve the production and reproduction in buffalo.

MATERIAL AND METHOD

The present study was conducted on testes of 18 Indian buffalo foetii of different ages obtained from abattoir and Veterinary clinics. The foetal body length was measured as a curved line in centimeters with a calibrated inelastic thread along the vertebral column between the most anterior part of frontal bone to the rump at ischiatic tuberosity and designated as curved crown rump length (CVR) (Edward, 1965). The approximate age of the foetii was calculated by using the following formula given by Soliman (1975).

Y = 28.66 + 4.496 X (CVR < 20 cm)

Y = 73.544 + 2.256 X (CVR 20 cm)

Where Y is the age in days and X is the CVR in centimeters. Based on CVR the samples were divided into three groups viz:

Group I: Foetal samples of CVR between 0 cm to 20 cm

Group II: Foetal samples of CVR between 20 cm to 40 cm

Group III: Foetal samples of CVR > 40 cm

Both the right and left testes were collected and fixed in Bouin's and 10% neutral buffered formalin (NBF) fixatives. The tissues were processed for paraffin blocks preparation by acetone-benzene schedule (Luna, 1968) and sections of 5-7mm thickness were cut with rotary microtome. These paraffin sections were stained with hematoxylin and eosin and Masson's trichrome stains to study the histomorphological details. The micrometrical observations on diameters of seminiferous tubules, Sertoli cells, gonocytes, Leydig cells and their nuclei were recorded on hematoxylin and eosin stained sections with the help of filar and ocular micrometer duly calibrated with stage micrometer.

RESULTS AND DISCUSSION

The micrometrical data on the longitudinal and rounded seminiferous tubules, sertoli cells, Leydig cells and gonocytes of right and left testes in group I, II and III have been summarized in Table I and the values have been statistically analyzed in Table II.

Seminiferous tubules. The testicular parenchyma comprised of seminiferous tubules and intertubular tissue. The seminiferous tubules were observed first in buffalo foetii of 8.0 cm CVR (65 days) at the gonadal periphery whereas the inner part of parenchyma consisted of network of polygonal mesenchymal cells and thin walled blood vessels. At 12.0 cm CVR (83 days), the testicular parenchyma was further subdivided into two zones. The outer zone contained the longitudinal/elongated/oval tubules whereas the inner zone had rounded tubules. The number of longitudinal tubules was more than the rounded tubules at this age group but the reverse pattern was observed at 14.0 cm CVR (92 days) (Fig.1). With the advancing age of the foetus the central zone disappeared and was replaced by mediastinum testes at 18.2 cm CVR (110 days) whereas the outer zone was responsible for the formation of permanent seminiferous tubules. These seminiferous tubules developed from pre existing clusters of epitheloid cells. The present findings are in agreement with the earlier reports on the testicular parenchyma of dog (Kawakami et al., 1993) and bovine fetuses (Wrobel, 2000). At 39.5 cm CVR (163 days) most of the seminiferous tubules were rounded in shape and only a few were longitudinal whereas in group III the number of longitudinal tubules were more than the rounded ones as observed earlier by Wrobel (2000).


Fig. 1. 14.0 cm CVR buffalo foetus showing blood vessels (BV)
and seminiferous tubules (S) in outer zone. Hematoxylin and Eosin stain x 175.


Fig. 2 12.0 cm CVR buffalo foetus showing the formation of fetal Leydig cells
(LC) and fibroblasts (F) in the interstitium. Hematoxylin and Eosin stain x 875.


Fig. 3 Differentiation of pre-Sertoli cells (Ps), gonocytes (G), fibroblasts (F),
peritubular cells (PC) and Leydig cells (LC) in the buffalo
foetus of 14.0 cm CVR. Hematoxylin and Eosin stain x 875.


Fig. 4 Testicular parenchyma of left testis in 75.0 cm CVR buffalo foetus showing well
defined basement membrane (B), peritubular cells (PC), pre-Sertoli cells (Ps), gonocytes (G)
and Leydig cells (LC) could be seen. Hematoxylin and Eosin stain x 875.

The seminiferous tubules were surrounded by a distinct basement membrane and a layer of peritubular cells at 8.0 cm CVR (65 days) buffalo foetii. These peritubular cells were suggested to be originating from the tunica albuginea and then settled around the seminiferous tubules. These cells are responsible for maintaining the structural integrity of seminiferous tubules. It has been suggested that the basement membrane underlying the seminiferous tubular epithelium in post natal life is a shared product of Sertoli and peritubular cells (Skinner et al., 1985). These peritubular cells were elongated in shape with a rounded nucleus. The single layer of these cells became double layer at 88.0 cm CVR (272 days) (Fig.5).

Fig. 5 88.0 cm CVR buffalo foetus showing more differentiation of pre-Sertoli cells (Ps),
gonocytes (G), Leydig cells (LC) and peritubular cells (PC). Masson's Trichrome stain x 350.

 

The number of seminiferous tubules increased with the advancing age of the foetii. From the data it is inferred that there was no significant difference between the diameter of longitudinal seminiferous tubules of right and left testes. A progressive increase in the diameter of seminiferous tubules was noticed with the increase in age of buffalo foetii. The present findings are in agreement with the findings of Singh et al. in goat fetuses.


The correlation in the diameters of the seminiferous tubules in right and left testes with respect to the age of buffalo foetii was statistically significant for rounded tubules (p = 0.05) and non significant for longitudinal tubules. No significant difference was observed regarding various parameters between right and left testes within the individual age groups except the diameter of longitudinal seminiferous tubules.

Pre Sertoli cell. The pre Sertoli cells were first observed in buffalo foetii of 8.0 cm CVR (65 days) at the periphery of the seminiferous tubular. Similar organization of pre Sertoli cells was also observed by Pelliniemi (1975) and Dang & Fouquet (1979) in pig and Macaca fascicularis fetuses, respectively.

The pre Sertoli cells were large in number and polygonal or conical in shape with irregular dark nuclei and pale cytoplasm (Fig.3). The shape of these cells changed to columnar with an elongated nuclei in the testis of buffalo foetii at 88.0 cm CVR and they were lying perpendicular to the basement membrane at this age. Similar observations were made in guinea pig and human fetal testes by Black & Christensen (1969) and Rath & Anand (1992), respectively. These cells were found to be closely arranged adjacent to the basement membrane of seminiferous tubule. The basement membrane act as a uniting media between the pre Sertoli cells and peritubular cells which are responsible for the formation of blood-testicular barrier (Byskov & Hoyer, 1994).

The maximum number of pre Sertoli cells was found to be 16-20 per cross-sectional area and their number considerably increased with the progression of fetal life (Fig.4). Similar observations were also made by Abd-Elmaksoud in the bovine fetal testis. It was interpreted that there was no significant difference in the diameter of pre-Sertoli cells in left and right testes but it increased progressively with the advancing age of the buffalo foetii. Similar pattern was also observed with the nuclear diameter of these cells. The statistical analysis revealed a positive correlation between the size of pre Sertoli cells with respect to age of buffalo foetii, which was found to be significant at 1% level (p=0.01) in both the testes. The comparison of pre- Sertoli cell and its nuclear diameter in right and left testes showed that it was more in group I as compared to group III.

Gonocytes. The gonocytes were demonstrable in the centre of seminiferous tubules surrounded by pre Sertoli cells at 10.0 cm CVR (74 days) of buffalo foetii. These cells were rounded in shape with a prominent spherical nucleus at 14.0 cm CVR (92 days). The gonocytes were also located at the periphery and centre of seminiferous tubules (Fig.4) but these cells rarely come in contact with the basement membrane of seminiferous tubules. It is clear from the data that the size of gonocyte and their nucleus did not vary significantly with respect to age and testis of the buffalo foetii. This might be due to the fact that these cells continue to undergo mitotic replication and do not enter meiosis in the male fetuses (Mc-Carrey, 1993). The above findings are in agreement with the observations made on the fetal testes of pig (Black & Christensen; Black, 1971), goat (Singh et al., 1979) and dog (Kawakami et al.). The correlation of coefficient between the diameters of gonocytes and their nuclear with respect to CVR of buffalo foetii was found to be non significant in both the testes.

Interstitial tissue. The interstitial tissue contained numerous blood vessels, lymph vessels, Leydig cells, loose network of mesenchymal cells, connective tissue cells and 1-2 layers of peritubular cells surrounding testicular tubules. These cells were present in close proximity to the blood vessels indicating their endocrine nature. At 8.0 cm CVR (65 days) the interstitium was comparatively less developed and showed the presence of darkly stained fetal Leydig cells. These cells were found to be polygonal in shape with rounded nuclei which is darker than the pre-Sertoli cells, the gonocytes and their cytoplasm was acidophilic. In addition to fetal Leydig cells, few fibroblasts were also observed at various stages of differentiation at 12.0 cm CVR (83 days) (Fig. 2) which may form the adult Leydig cells. Our findings are in agreement with the findings of Wrobel (2000). At 14.0 cm CVR (92 days), the interstitium significantly expanded due to the differentiation of mesenchymal cells into fetal Leydig cells and constituted the larger cell population of all the other components of interstitium which was well presented at 36.5 cm CVR (156 days). This concept is supported by the observation of Orth (1993) who found that the number of mesenchymal cells decreased with the appearance of Leydig cells in pig testes. The number of fetal Leydig cells was found to be more as compared to the gonocytes and pre-Sertoli cells but a decrease in the number of Leydig cells with the advancement of fetal age could also be demonstrated. This could be due to degeneration of earlier Leydig cells or due to the proliferation of intertubular tissue during the prenatal development of testes (Wrobel, 1990). Our findings are also supported by Huhtaniemi & Pelliniemi (1992) and Singh (1996) who also reported a decrease in number of Leydig cells with increase in the age of the foetii. The diameter of fetal Leydig cells remained same in both the right and left testes but there was an increase in the diameter of fetal Leydig cells from group I to group III. Their nuclear diameter did not vary significantly with respect to age of the foetii and left and right testes. Our results correspond well with the findings of Singh et al. (1979) and Pachpande et al. (2006). There was a positive correlation between the diameters of Leydig cells and their nuclei with respect to age of foetus, which was found to be significant at 1% level (p=0.01) for Leydig cells and non significant for their nuclei.

 

REFERENCES

Abd-Elmaksoud, A. Morphological, glycohistochemical and immunohistochemical studies on the embryonic and adult bovine testis. PhD. Thesis, Ludwig Maximilians Universitat Munich, Germany, 2005.         [ Links ]

Black, V. H. Gonocytes in fetal guinea pig testes: phagocytosis of degenerating gonocytes by Sertoli cells. Am. J. Anat., 131(4):415-26, 1971.         [ Links ]

Black, V. H. & Christensen, A. K. Differentiation of interstitial cells and Sertoli cells in fetal guinea pig testes. Am. J. Anat., 124(2):211-37, 1969.         [ Links ]

Byskov, A. G. & Hoyer, P. E. Embryology of mammalian gonads and ducts. In: Knobil, E. & Neill, J. D. (eds). The Physiology of Reproduction. 2nd Ed. New York, Reven Press, 1994. pp 487-540.         [ Links ]

Dang, D. C. & Fouquet, J. P. Differentiation of the fetal gonad of Macaca fascicularis with special reference to the testis. Ann. Biol. Anim. Bioch. Biophys., 19(4B):1197-209, 1979.         [ Links ]

Edward, M. J. Observations on the anatomy of reproductive organs of cows with special reference to those features sought during examination per rectum. N. Z. Vet. J., 13(2):25-37, 1965.         [ Links ]

Hochereau de Reviers, M. T.; Perreau, C.; Pisselet, C.; Locatelli, A. & Bosc, M. Ontogensis of somatic and germ cells in sheep fetal testis. J. Reprod. Fertil., 103(1):41-6,1995.         [ Links ]

Huhtaniemi, I. & Pelliniemi, L. J. Fetal Leydig cells: Cellular origin, morphology, life span and special functional features. Proc. Soc. Exp. Biol. Med., 201(2):125-40, 1992.         [ Links ]

Kawakami, E.; Yamada, Y.; Tsutsui, T.; Ogasa, A. & Yamauchi, M. Changes in plasma androgen levels and testicular histology with descent of the testis in the dog. J. Vet. Med. Sci., 55(6):931-5, 1993.         [ Links ]

Luna, L. G. Manual of Histological Staining Methods of Armed Forces Institute of Pathology. 3rd Ed. New York, McGraw Hill Book Company, 1968. pp.38-196.         [ Links ]

Mc-Carrey, J. R. Development of the germ cell. In: Desjardins, C. & Ewing, L. L. (eds). Cell and Molecular Biology of the Testis. Oxford, Oxford University Press, 1993.         [ Links ]

Orth, J. M. Cell biology of testicular development in the fetus and neonate. In: Desjardins, C. & Ewing, L. L. (eds). Cell and Molecular Biology of the Testis. Oxford, Oxford University Press, 1993.         [ Links ]

Pachpande, A. M.; Raut, S. S.; Patil, A. D.; Dhande, P. L.; Gaikwad, S. A. & Yadav, G. B. Histological and Histochemical study of Testis and epididymis during prenatal development in sheep. Proc. XX Convention IAVA, Jabalpur, India, 2006. p.65.         [ Links ]

Pelliniemi, L. J. Ultrastructure of the early ovary and testis in pig embryos. Am. J. Anat., 144(1):89-111, 1975.         [ Links ]

Rath, G. & Anand, C. Differentiation of cells of seminiferous tubules: A study on human fetal testis. J. Anat. Soc. India, 41(1):35-41, 1992.         [ Links ]

Singh, P. Histological, histochemical and histoenzymic studies on the post natal development and differentiation of seminiferous epithelium and Leydig cells in buffalo testis. M.V.Sc. Thesis, Punjab Agricultural University, Ludhiana, India 1996.         [ Links ]

Singh, Y.; Sharma, D. N. & Dhingra, L. D. Morphogenesis of the testis in goat. Indian J. Anim. Sci., 49:925-31, 1979.         [ Links ]

Skinner, M. K.; Tung, P. S. & Fritz, I. B. Cooperativity between Sertoli cells and testicular peritubular cells in the production and deposition of extracellular matrix components. J. Cell. Biol., 100(6):1941-7, 1985.         [ Links ]

Soliman, M. K. Studies on the physiological chemistry of the allantoic and amniotic fluids of buffaloes at various periods of pregnancy. Indian Vet. J., 52:111-7, 1975.         [ Links ]

Tekade, J. S. Studies on prenatal development of gonads in goat (Capra hircus). M. Sc. Thesis, Nagpur, Maharashtra, India, 2001.         [ Links ]

Wrobel, K. H. The postnatal development of the bovine Leydig cell population. Reprod. Dom. Anim., 25(2):51-60, 1990.         [ Links ]

Wrobel, K. H. Morphogenesis of the bovine rete testis: the intratesticular rete and its connection to the seminiferous tubules. Anat. Embryol., 202(6):475-90, 2000.         [ Links ]


 

Correspondence to:

Manjinder Kaur

Professor-cum-Head
Department of Veterinary Anatomy & Histology
Guru Angad Dev Veterinary and Animal Sciences University
Ludhiana
Punjab
INDIA
Phone No. +91-161-2414028
Fax No. +91-161-2400822

Email: bansal.neelam@rediffmail.com

Received: 12-11-2008
Accepted: 16-04-2009

Creative Commons License Todo el contenido de esta revista, excepto dónde está identificado, está bajo una Licencia Creative Commons