| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
a Department of Physiology, Southern Illinois University School of Medicine, Carbondale, Illinois 62901-6512
b School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104
c Department of Morphology, Institute of Biological Sciences, Federal University of Minas Gerais, Belo Horizonte, Minas Gerais, Brazil 31270-901 CP 486
 |
ABSTRACT |
|---|
 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
|
INTRODUCTION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
Questions have arisen about how germ cells introduced into the seminiferous tubules eventually produce sperm from stem cells residing on the basal aspect of the tubules. In particular, it was necessary to determine how fast and with what characteristics spermatogenesis developed. In this report, we examine the characteristics of spermatogenesis at different times after transplantation of donor germ cells into genetically sterile recipients.
|
MATERIALS AND METHODS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
Testis cells were isolated, according to the technique of Bellvé [8] as employed by Ogawa et al. [9], from C57Bl/6 mice ranging in age from 5 to 6 wk. Cells were injected into WCB6F1 W/Wv male mice, obtained from Jackson Labs (Bar Harbor, ME), at a concentration of 108 cells/ml, according to the methodological details provided by Ogawa et al. [9]. At the time of injection, trypan blue dye was mixed into the cell suspension to visualize the entry of injected cells into seminiferous tubules. From 75% to 95% of surface tubules were filled with blue dye. Recipients, chosen randomly, were killed at 10 min, 9 h, 24 h, 1 wk, 1 mo, 2 mo, and 3 mo after transplantation. Control WCB6F1 W/WV genotype mice were killed without germ cell transplantation, one group at 10 min ("start control") and one group at 3 mo ("end control").
Tissue Preparation
All recipients were killed after administration of pentobarbital, while applying the perfusion procedure to fix the testis [10]. Briefly, after i.p. administration of heparin (130 IU/kg BW), mice were anesthetized, and the abdominal and thoracic cavities were opened to expose the heart. The animal's circulation was cleared by introducing 0.9% saline via a needle in the heart, and then the tissues were perfused-fixed by gravity-fed perfusion with 5% buffered glutaraldehyde for 25–30 min. The testes and epididymides were removed, weighed, and prepared for embedding in epoxy by using standard techniques [11]. Sections of 1-µm thickness were examined by light microscopy.
A sample of the testis cells used for transplantation was prepared for plastic embedding by first gently pelleting the cell suspension. The supernatant was removed, and 5%-buffered glutaraldehyde was placed over the pellet, which was about 0.5 mm in thickness. The pellet was prepared for light microscopy.
The criteria used to stage mouse spermatogenesis were those outlined by Oakberg [12] as modified by Russell et al. [13]. Groups were considered statistically different if their least-squares means differed with p < 0.05. ANOVA was performed followed by individual t-tests. Each testis obtained was considered to represent an "n" of 1. Thus 4 mice and 8 testes were examined for each sampling time point.
|
RESULTS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
Most W/Wv (W-locus) control mice, not having received transplants, displayed small seminiferous tubules comprising Sertoli cells. Some tubules contained undifferentiated interphase and mitotic spermatogonia (14–19%; Fig. 1). Only one tubular profile (of thousands examined) proceeded to pachytene and another to round spermatids. Both of the latter tubular profiles were noted in the start control. Like the control, spermatogenesis in W-locus recipient mice did not occur in any of those tubules from the 10-min and 1-wk groups that were randomly selected for quantitation.
|
The seminiferous epithelium of W-locus mice was highly vacuolated, and the apical processes of Sertoli cells either abutted a lumen (Fig. 2) or made contact with those radially directed Sertoli cell processes elsewhere around the tubule, the latter occurring in tubules in which there was no lumen. In some tubules, spermatogonia were undergoing mitosis or degeneration (Fig. 2).
Sertoli cells not showing contact with the basal lamina were grouped in "balls" [2] within the epithelium or lumen and displayed dense cytoplasm and small nuclei with peripheral heterochromatin (Fig. 3). Their cytoplasmic boundaries were generally rounded. Sertoli cells whose nucleus was luminally positioned never demonstrated cytoplasmic processes that contacted the basal lamina. Macrophages were also found in the tubular lumen or within the epithelium.
Donor Cells
Donor cells used for transplantation contained germ cells of all types, including type A spermatogonia, as well as somatic cells (Fig. 4). Somatic cells that could be identified within the cell suspension were Sertoli cells, Leydig cells, and macrophages; some unidentified cells may have been endothelial cells or peritubular myoid cells. Cell types within the pellet are better characterized by electron microscopy, and such a characterization is currently being finalized.
Testis and Epididymis Weights
Recipient testis weights at 10 min or 9 h after injection were not different from those of start controls. Increments in testicular weight at 24 h and thereafter were significant (Fig. 5) but were not due to age-related growth as the end and start controls were similar. After transplantation, the testis progressively became heavier; by 3 mo the mean testis weight had not quite doubled.
|
With time, epididymal weights of transplanted groups generally increased compared with those of the start control group, but the data showed no clear trends (Fig. 6). Since the epididymal weight of the end control increased and was comparable to that of the 3-mo transplants, one could attribute the increase in epididymal weight as being age related.
Lumen Size and Donor Germ Cells in the Lumen
Transversely sectioned tubules were classified into those with no lumen, or a small (1 to 40 µm) or large (> 40 µm) lumen by measuring their greatest diameter. The percentage of tubules with no lumen decreased significantly until 1 wk, increased at 1 mo, and decreased again at 3-mo posttransplantation. The 3-mo-transplanted animals showed significantly fewer tubules with no lumen, a feature suggesting that active spermatogenesis resulted in the production of a lumen (Fig. 7). On the other hand, our histological observations showed that many tubules developed spermatogenesis at 1 mo and 2 mo after transplantation, and some of the tubules with active spermatogenesis did not have a lumen or showed only a small lumen (Fig. 8). Tubules with large lumina predominated in transplanted tubules from 10 min to 1 wk posttransplantation compared with the other groups (Fig. 9).
|
|
At 10 min and 9 h posttransplantation, ~42% of the tubules showed germ cells in the lumen, but the percentage was less at 1 through 3 mo (Figs. 10–11) compared with none in the start control. There was no statistical difference between the percentage of tubules containing germ cells in the lumen at the 10-min and 9-h intervals; however, the percentage of tubules with luminal germ cells decreased progressively at each time interval until 1 mo. No further significant decreases in the percentage of tubules with luminal germ cells was noted in the 2-mo and 3-mo groups.
|
Development and Characterization of Donor Spermatogenesis
Spermatogenesis was quantified in all groups as the number of transversely sectioned tubules containing germ cells that were more advanced than the type A spermatogonia (Fig. 12). In transplanted tubules, the first evidence of spermatogenesis was at 1 mo, at a time when germ cells had maximally progressed to early pachytene (Fig. 13). Spermatogenesis occurred at 1 mo in 0.8% of tubular profiles (Fig. 13), in 8.9% by 2 mo (Fig. 8), and in 28.2% by 3 mo (Fig. 14). A few step 16 spermatids and sperm were evident at 2 mo, and virtually all tubules with spermatogenesis contained elongated(ing) spermatids in the 3-mo group. No spermatogenesis was seen in the end control group. The percentage of tubules with spermatogonia did not differ in any of the groups examined (Fig. 15). Vacuolation of the epithelium was far less common in tubules with active spermatogenesis (Fig. 8), especially in the 3-mo group (Fig. 14).
|
|
|
Recognized stages were evident in most tubules with spermatogenesis from transplanted animals (Fig. 14). Seminiferous tubules showed a frequency of stages of the cycle similar to what could be expected for normal spermatogenesis, although only a preliminary determination was made. Spermatogenesis was not quantitatively normal, in that animals transplanted for 2 mo showed a smaller number of elongated spermatids compared with the number of round spermatids present in the same tubule (Fig. 8). To some degree, the paucity of elongated spermatids was similar to what is known to occur in pubertal rats during the first wave of spermatogenesis [14]. At 3 mo, a much greater number of elongated spermatids were present, although we did not verify whether the number produced was comparable to that of normal animals.
Although recognized stages were formed in most tubules from 2- to 3-mo-old transplants, a minority of tubules showed cells that were incompatible with our concept of the makeup of cell associations found in a mouse staging map [13]. For example, in a stage VII or early stage VIII tubule, occasional meiotic figures were found as well as basal compartment mitotic figures (Fig. 16). In another tubule, secondary spermatocytes, step 10 and step 6–7 spermatids were noted in relatively close juxtaposition (Fig. 17). At 2 mo, 76.5% of tubules with spermatogenesis showed compatible layers of a recognized cell association. At 3 mo, this number increased to 94.1%.
|
In a minority of tubules, an entire generation of cells (spermatocytes or spermatids) was lacking, although both the less mature and the more mature cells of that cell association were present. For example, in a stage IV tubule, all pachytene spermatocytes, an expected component of this cell association, were lacking (Fig. 18). At 2 mo and 3 mo posttransplantation only 17.6% of the tubules with spermatogenesis showed a missing cell layer.
The morphological appearance of transplanted germ cells generally was normal, except that defective elongated spermatids occurred frequently at stage VII (Fig. 19). It was not uncommon to detect phagocytosed elongated spermatids deep within the epithelium in stages VIII–XI after the expected period of normal sperm release.
Examination of seminiferous tubules at the short intervals after transplantation revealed that occasional germ cells were closely related to apical processes. Some were positioned between adjacent Sertoli cells and thus formed part of the epithelium (Fig. 20). The percentages of tubules (~200 tubules enumerated for each animal of each group) with intraepithelial adluminal germ cells at 10 min, 9 h, 24;thh, and 1 wk were 6.4%, 7.9%, 3.1%, and 2.4%, respectively. These observations indicated that the uptake of germ cells by the epithelium occurred rapidly, but in only a small proportion of the tubules, and that most cells were lost (Fig. 21) between 1 and 4 wk after transplantation.
|
|
Spermatocytes were the most abundant cell type present in the adluminal epithelium at 10 min, 9 h, and 1 day after transplantation; but at 1 wk, over twice as many spermatids as spermatocytes were present (Fig. 21). Meiotic cells (metaphase, anaphase, and telophase) were present within the epithelium at 9 h, 24 h, and 1 wk (Figs. 22 and 23), although they were present in only a small number (less than 1%) of the tubules. At 1 wk, we observed meiotic cells (Fig. 23) and apparent clones of developmentally similar spermatids or sometimes symplasts of young spermatids, not seen at earlier time periods (Fig. 24).
|
|
Germ Cell Degeneration
The number of degenerating germ cells within seminiferous tubules was expressed per tubule cross section and expressed per Sertoli cell nucleus in the same tubule in which they were quantified. The results of both methods were generally similar; only quantitative information about degenerating germ cells expressed per Sertoli cell nucleus in the same tubules is presented herein (Fig. 25). Degenerating germ cells were seen in all control and transplanted groups. By light microscopy, spermatogonia were observed to degenerate in the start control and in the end control; it was found that there were no significant differences in the numbers of degenerating cells between these two groups. Ten minutes after transplantation, the number of degenerating germ cells of all types had increased significantly, and their number remained high at the 9-h and 24-h groups (Fig. 25). A significant decrease in the number of degenerating cells occurred by 1 mo such that the 1-mo, 2-mo, and 3-mo transplanted groups showed no significant differences in cell degeneration when compared with both control groups.
|
Other Features
As reported previously [2], balls of Sertoli cells were seen in control untransplanted (Fig. 3) as well as transplanted testes. Sertoli cell balls, expressed as the percentage of tubules with balls, increased in frequency as the time after transplantation increased (Fig. 26).
Intratubular macrophages were expressed as a percentage of Sertoli cells within the same tubules in which they were enumerated (Fig. 27). By 24 h after transplantation, the number of macrophages increased over the start control and remained increased until 1 mo, when the numbers decreased to control levels in the 2-mo transplant group. The start control and the end control showed no difference in the number of intratubular macrophages. Examination of macrophages showed them to phagocytose some sperm produced at the 2-mo transplantation interval and numerous sperm at the 3-mo transplantation interval (Fig. 28). Sperm were also phagocytosed by Sertoli cells in areas of the tubules not showing spermatogenesis, in accordance with what has been shown previously [2]. Macrophages were often seen in association with Sertoli cell balls.
|
In the 10-min, 9-h, and 24-h transplant groups, germ cells were occasionally detected outside the seminiferous tubules in the intertubular compartment (Fig. 29). When all slides made for animals in these groups were searched, it was noted that 50% of the animals in the 10-min group and 37.5% of the animals in the 9-h group demonstrated extratubular germ cells. Extravasated germ cells were usually localized to only one region of the testis. By 1 wk and thereafter, none of the testis showed evidence of extratubular germ cells.
Epididymis
The caput and cauda epididymidis of each testis was examined for the presence of luminal germ cells. Occasional degenerating cells that could not be identified were present in the caput of both control and transplanted groups. Unidentified cells were always seen in the cauda of both control and transplanted animals. Of the transplanted animals, only one animal (found in the 9-h group) showed the presence of viable germ cells in the caput, and no animals showed them in the cauda. Observations made at the time of introduction of the cell suspension by two of us (T.O. and M.R.A.) indicates that the cell suspension backflows into the caput epididymidis more frequently than would be indicated by histological studies performed herein, especially those conducted at 10 min after transplantation.
|
DISCUSSION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
The positive-pressure introduction of a testis cell suspension mixture into seminiferous tubules results in occasional breaks of either the seminiferous tubules or the rete testis since some of the transplanted testes display germ cells and other cell types lying outside the tubules at early time intervals after transplantation. Although about a third of the animals showed extratubular germ cells at 24 h after transplantation, these cells must have been phagocytosed by interstitial macrophages before 1 wk after transplantation since no intertubular germ cells were seen after the 24-h group.
Microinjected germ cells were seen in a minority of the sectioned tubular profiles within the lumen of seminiferous tubules at 10 min, 9 h, and 24 h. The frequency of finding germ cells in the lumen declined substantially by 1 wk, suggesting they were virtually eliminated by this time. The few luminal germ cells present at 2 mo and 3 mo were probably from sloughing of established transplant spermatogenesis. The quantitative data on numbers of degenerating germ cells as well as their infrequent presence in the epididymal duct system verified that germ cells were eliminated primarily through uptake and phagocytosis by Sertoli cells, beginning as early as 10 min after transplantation and continuing through 24 h.
A statistical increase in tubules with large lumina was in evidence by 10 min after transplantation (Fig. 9). Although it seems obvious that microinjecting a suspension of cells to seminiferous tubules during the transplantation procedure will cause the tubules to swell to accommodate the cell suspension, it is not obvious why the lumen stays large for up to a period of 1 wk. After 1 wk, the percentage of tubules with a large lumen decreased sharply and was low compared with earlier time periods, although tubules with a large lumen were significantly greater in numbers than the end control.
At any one time period (10 min, 9 h, and 24 h), a minority of germ cells were closely related to the apical processes of Sertoli cells and were in the intercellular spaces between Sertoli cells (i.e., within the epithelium). All germ cell types could be found within the epithelium. Intraepithelial germ cells comprised a small fraction of the germ cells seen in seminiferous tubules since most appeared to be within the lumen. It did not appear that the early reaction of Sertoli cells was to attach to them and move them en mass into the epithelium. The data suggested that the slow, but progressive, uptake of germ cells and their subsequent degeneration was the main process by which transplanted germ cells were eliminated, given that few germ cells were seen within the epididymal lumen or the rete.
Although spermatogonia of the type A variety were seen within the epithelium, it was not possible to determine whether these spermatogonia were stem cells or were in the process of migrating from the lumen to the basal compartment, where they are known to reside after transplanted spermatogenesis is established [2]. Since stem cells comprise a small percentage of all type A spermatogonia and spermatogenesis is established at only a few sites in the testis (Brinster and colleagues, personal observations) and the migration may be rather rapid, it is unreasonable to expect to observe single stem cells migrating.
Confirmation of the initial establishment of donor spermatogenesis was at the 1-mo time point, when small clones of young spermatocytes (stages II–VI) were observed. Although only a few colonies were observed, all that were observed had advanced to the young spermatocyte phase of development. Had we chosen to sample at approximately 21–22 days posttransplantation (a spermatogenic cycle earlier), we believe that small clones of intermediate and B spermatogonia could have been detected as the first evidence of successful spermatogenesis. Detection of transplanted spermatogonia at earlier periods would have necessitated the use of a donor gene marker such as LacZ.
Our calculations indicated a delay in the development of spermatogenesis posttransplantation. Assuming that cells of the As variety are the "true" stem cell, as is widely believed [15, 16], and that the seminiferous epithelial cycle in the mouse follows a pathway and timing in the early transplants similar to that demonstrated for longer-term transplants [17], our calculations indicate that approximately two cycles (~16–18 days) were necessary before transplanted mouse cells could have begun division from As cells and attained the early pachytene phase that we observed as the maximal progress of spermatogenesis at the 1-mo interval. Thus there were approximately 12–14 days available for transplanted stem cells to migrate from the tubular lumen to the base of the tubule, become established, divide to replenish themselves, and later divide to form cells committed to spermatogenesis. The best-studied model of germ cell renewal from adult stem cells is that of postirradiation recovery. It is known that the re-establishment of spermatogenesis after severe depletion of germ cells with neutron irradiation is not immediate [18]. Time is needed for stem cells to form a pool of stem cells that, at a later time, can go forward to initiate spermatogenesis.
After irradiation, a disabling condition that few stem cells survive, the probability is near 100% that stem cells will divide to replenish themselves. As isolated stem cells form a larger colony of stem cells, the probability of self-renewing divisions progressively decreases [19]. The more severe the treatment, the longer the time before repopulation begins [18], with delays of 1 wk not being unusual [20]. As in the irradiation model, in which the dose of irradiation is high, at short time intervals after transplantation there are only widely separated stem cells to initiate development. The cell cycle time for As spermatogonia is probably about 4 days [21], although the exact timing of the cell cycle in the mouse is not known. We could also assume that transplanted cells probably undergo two or three self-replenishment divisions before a differentiating division.
The establishment of spermatogenesis was rapid, occupying approximately 10-fold more tubular profiles at 2 mo than at 1 mo and 2.5-fold more tubule profiles at 3 mo than at 2 mo. We rarely saw tubules at 2 mo or 3 mo like those shown in Figure 13 (1 mo) which could have represented new clones of germ cells. Thus, it is our belief that the increased number of tubular profiles in which spermatogenesis was present was due to the lateral expansion of spermatogenesis along tubules rather than to a later activation of quiescent transplanted stem cells. Recruitment of quiescent stem cells would have produced small patches of qualitatively and quantitatively incomplete spermatogenesis at 2 mo and 3 mo, a feature that was not seen.
Some regions of the testis are rich in spermatogenesis, and other regions are completely deficient. We suggest that transplanted spermatogenesis occurred only in those tubules in which type As cells were successfully transplanted, usually less than half of the tubules. Spermatogenesis expands laterally until it reaches a certain point (or the rete) and then stops. We have observed in longer-term transplants that many tubules never attain spermatogenesis [2]. It appears that if a tubule does not initially receive stem cells, that tubule will not develop spermatogenesis by migration of stem cells through the rete. Repopulation after cell transplantation presents a pattern similar to what occurs after irradiation, in which case it is hypothesized that lateral spread of spermatogonia after stem self-renewal cell divisions results in repopulation of tubules [22]. After neutron irradiation, the lateral (longitudinal) spread of spermatogenesis is hypothesized to be 27–33 µm/day [20, 22].
A statistical analysis of testis weight showed that the transplantation procedure itself did not significantly increase testis weight. Surprisingly, testis weight was increased at 24 h after transplantation and remained increased in all groups transplanted for longer periods. Since no significant spermatogenesis could have occurred in this period to contribute to the weight increase, we believe that the sterile W-locus testis began to respond to the presence of germ cells by increasing cell mass or by increasing fluid secretion. It is known that the presence of germ cells can stimulate changes in size and secretions of the Sertoli cell [23, 24] and promote the development of the smooth endoplasmic reticulum [24]. Three months after transplantation, the testis size had almost doubled from the start control, but it had not approached the weight of a normal mouse testis [25].
The apical processes of Sertoli cells will attach to and move germ cells within the seminiferous epithelium such that they come to occupy the adluminal compartment and lie between adjoining Sertoli cells. Spermatogonia were taken up in the adluminal Sertoli compartment by recipient Sertoli cells but were not seen in this position after 9 h. The fate of most transplanted spermatogonia remains unknown. Perhaps most were transported to the basal compartment, where the minority of them, stem cells, initiated spermatogenesis and/or where all other spermatogonia were phagocytosed. Good markers to differentiate spermatogonial types will facilitate our efforts to follow stem cells.
Although numerous spermatids were found in the donor cell suspension used for transplantation, the data show that the recipient Sertoli cells prefer to develop a relationship with spermatocytes. It is known that spermatocytes are preferentially bound to cultured Sertoli cells when a mixture of germ cells is available to them (reviewed in [23]). Although uptake of germ cells is the primary way Sertoli cells eliminate transplanted cells, we found that less than 10% of the sectioned tubule profiles at any of the early time intervals show evidence of uptake of germ cells.
The observations suggest to us that dividing (metaphase, anaphase, and telophase) spermatocytes undergo development within the Sertoli cell epithelium at a time when virtually all other germ cells have been phagocytosed. The presence of meiotic figures at 9 h and 24 h as well as the relative increase in round spermatids in the 1-wk group (Figs. 22–24) suggests that meiosis is one of the "favored" phases of differentiation in the nearly Sertoli-cell-only testis. Since no animals were killed between 1 wk and 1 mo, we were not able to determine whether the production of small numbers of round spermatids resulted in continued differentiation of round spermatids into sperm. It should be noted that less than 1% of tubular profiles up to 1 wk showed meiotic figures and/or clones of round spermatids. Although the differentiation phenomenon involving meiotic divisions appears to occur, it is rare.
Missing layers of germ cells is a feature that some tubules in transplanted animals have in common with tubules in which repopulation of germ cells occurs after irradiation (Meistrich, personal communication). This phenomenon may reflect two alternative actions of stem cells: division for self renewal or division to form paired cells (connected by intercellular bridges) that in turn form sperm. The former would result in the complete lack of a cell layer. At the growing edge of spermatogenesis as it expands laterally along the tubule, these alternatives—self-renewal or differentiation—are available more frequently than in areas where spermatogenesis is fully established. More stem cells would be needed to establish a stem cell population in rapidly expanding areas; thus occasionally differentiating divisions are sacrificed, resulting in a missing cell layer. Spermatogenesis in the 2-mo-transplanted animal shows more areas of missing cell layers than in 3-mo-transplanted animals, revealing that spermatogenesis becomes more like its normal counterpart as the time increases after transplantation.
Macrophages that were present within tubules of control W-locus mice appeared to increase in number after transplantation, reaching their greatest number at about 1 mo and declining to control levels thereafter. The increase in the number of macrophages was not due to injection of macrophages since there was no significant difference in their numbers 10 min after transplantation. Macrophages may either migrate into seminiferous tubules or may divide in situ. Although the number of macrophages at 3 mo was reduced compared with 2 mo, the phagocytic activity of macrophages was obvious in 3-mo transplants. Numerous sperm were engulfed by macrophages at this time. The factors responsible for engulfment of apparently histocompatible sperm are not known.
The data indicate that the formation of balls of Sertoli cells is a normal feature in the sterile mouse. Since the end control showed a greater number of Sertoli cell balls compared with the start control, their formation is age-related. An alternative explanation is that the frequency of balls in tubules with spermatogenesis is lower because the tubule is larger. The age-related increase in balls leads us to conclude that they are the result of age-related sloughing of Sertoli cells, since the latter do not proliferate in adult rodents. The cause of Sertoli cell sloughing is not known, but our observations suggest that prolonged Sertoli cell inactivity may lead to their progressive sloughing.
It should be noted that the data reported based on use of the W-locus mouse as a recipient may not be representative of transplantation under conditions in which depletion of germ cells in the recipient is produced by other means. The W-locus mice were preferable for the present study compared with busulfan-treated animals since, in the latter, some endogenous spermatogenesis returns in concert with transplanted spermatogenesis. The conditions under which transplantation success is achieved in W-locus mice reveal that Sertoli cells from adult animals that have never had germ cells, other than a few undifferentiated spermatogonia, can produce qualitatively complete spermatogenesis within a 2-mo period. Under such conditions, the growth of spermatogenesis is rapid, and the presence of endogenous, albeit defective, spermatogonia in 14–19% of the tubules does not apparently interfere with the development of apparently normal spermatogenesis in other tubules. There is considerable work to be done in identifying ways to improve the purity of donor cells and to determine the best conditions for recipients to develop optimal spermatogenesis.
|
|
ACKNOWLEDGMENTS |
|---|
|
FOOTNOTES |
|---|
Financial support from the National Institutes of Health (HD 36504 and HD35494), USDA/NRI Competitive Grants Program, Commonwealth and General Assembly of Pennsylvania, and the Robert J. Kleberg Jr. and Helen C. Kleberg Foundation as well as support of the Brazilian Research Foundation, CNPq, for Ms. Gleydes Parreira and FAPEMIG for Luiz França is gratefully acknowledged. 
2 Correspondence. FAX: 618 453 1517; lrussell{at}som.siu.edu
Accepted: July 31, 1998.
Received: May 1, 1998.
|
REFERENCES |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
This article has been cited by other articles:
|
|
 |
|
  R. Lagos-Cabre and R. D Moreno Mitotic, but not meiotic, oriented cell divisions in rat spermatogenesis Reproduction, April 1, 2008; 135(4): 471 - 478. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. J. DOYLE, K. W. BRAUN, D. J. MCLEAN, R. W. WRIGHT, M. D. GRISWOLD, and K. H. KIM Potential Functions of Retinoic Acid Receptor A in Sertoli Cells and Germ Cells During Spermatogenesis Ann. N.Y. Acad. Sci., December 1, 2007; 1120(1): 114 - 130. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. M. K. MORROW, C. E. HOSTETLER, M. D. GRISWOLD, M.-C. HOFMANN, K. M. MURPHY, P. S. COOKE, and R. A. HESS ETV5 Is Required for Continuous Spermatogenesis in Adult Mice and May Mediate Blood Testes Barrier Function and Testicular Immune Privilege Ann. N.Y. Acad. Sci., December 1, 2007; 1120(1): 144 - 151. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Kanatsu-Shinohara, K. Inoue, H. Miki, N. Ogonuki, M. Takehashi, T. Morimoto, A. Ogura, and T. Shinohara Clonal Origin of Germ Cell Colonies after Spermatogonial Transplantation in Mice Biol Reprod, July 1, 2006; 75(1): 68 - 74. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 K. Jahnukainen, J. Ehmcke, and S. Schlatt Testicular xenografts: a novel approach to study cytotoxic damage in juvenile primate testis. Cancer Res., April 1, 2006; 66(7): 3813 - 3818. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. Khaira, D. McLean, D. A. Ohl, and G. D. Smith Spermatogonial Stem Cell Isolation, Storage, and Transplantation J Androl, July 1, 2005; 26(4): 442 - 450. [Full Text] [PDF]
|
|
|
|
|
|
J. M. Oatley, J. J. Reeves, and D. J. McLean Biological Activity of Cryopreserved Bovine Spermatogonial Stem Cells During In Vitro Culture Biol Reprod, September 1, 2004; 71(3): 942 - 947. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. M. Oatley, D. M. de Avila, J. J. Reeves, and D. J. McLean Testis Tissue Explant Culture Supports Survival and Proliferation of Bovine Spermatogonial Stem Cells Biol Reprod, March 1, 2004; 70(3): 625 - 631. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. Johnston, P. J. Baker, M. Abel, H. M. Charlton, G. Jackson, L. Fleming, T. R. Kumar, and P. J. O'Shaughnessy Regulation of Sertoli Cell Number and Activity by Follicle-Stimulating Hormone and Androgen during Postnatal Development in the Mouse Endocrinology, January 1, 2004; 145(1): 318 - 329. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. Goossens, V. Frederickx, G.D. Block, A.C.V. Steirteghem, and H. Tournaye Reproductive capacity of sperm obtained after germ cell transplantation in a mouse model Hum. Reprod., September 1, 2003; 18(9): 1874 - 1880. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. C. Nagano Homing Efficiency and Proliferation Kinetics of Male Germ Line Stem Cells Following Transplantation in Mice Biol Reprod, August 1, 2003; 69(2): 701 - 707. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. Lu and C. E. Bishop Late Onset of Spermatogenesis and Gain of Fertility in POG-Deficient Mice Indicate that POG Is Not Necessary for the Proliferation of Spermatogonia Biol Reprod, July 1, 2003; 69(1): 161 - 168. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Kanatsu-Shinohara, S. Toyokuni, T. Morimoto, S. Matsui, T. Honjo, and T. Shinohara Functional Assessment of Self-Renewal Activity of Male Germline Stem Cells Following Cytotoxic Damage and Serial Transplantation Biol Reprod, May 1, 2003; 68(5): 1801 - 1807. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. R. Franca and C. L. Godinho Testis Morphometry, Seminiferous Epithelium Cycle Length, and Daily Sperm Production in Domestic Cats (Felis catus) Biol Reprod, May 1, 2003; 68(5): 1554 - 1561. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
