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J Toppari

Publications and source records attributed to J Toppari.

At least 73 records · Page 4Linked to original sources
PubMed1994

Etoposide (VP-16) is a potent inducer of micronuclei in male rat meiosis: spermatid micronucleus test and DNA flow cytometry after etoposide treatment.

The genotoxic and cytotoxic effects of etoposide (VP-16), a topoisomerase II inhibitor, on male rat spermatogenic cells were studied by analysing induction of micronuclei during meiosis. Micronuclei (MN) were scored in early spermatids after different time intervals corresponding to exposure of different stages of meiotic prophase. Etoposide had a strong effect on diplotene-diakinesis I cells harvested 1 day after exposure, and a significant effect also on late pachytene cells harvested 3 days after exposure. The effect at 18 days corresponding to exposure of preleptotene stage of meiosis (S-phase) was weaker but also statistically significant. Adriamycin was used as a positive control in this study. The results indicate a different mechanism of action of etoposide compared with adriamycin and other chemicals studied previously with the spermatid micronucleus test. DNA flow cytometry was carried out to assess cytotoxic damage at the same time intervals (1, 3, and 18 days after treatment) at stages I and VII of the seminiferous epithelial cycle allowing a study of cytotoxicity to different spermatogenic cell stages. Damage of differentiating spermatogonia was observed by a decrease in the cell numbers of the 2C peak 1 and 3 days after treatment and by a reduction of the number of 4C cells (primary spermatocytes) 18 d after etoposide treatment. Adriamycin also killed differentiating spermatogonia. Since the cell population which showed a high induction of MN by etoposide was not reduced in number, the genotoxic effect is remarkable. We conclude that etoposide is a potent inducer of genotoxicity and patients treated with this agent during cancer chemotherapy are at a risk of genetic damage.

Animals↗
PubMed1994

Localization of urokinase- and tissue-type plasminogen activator mRNAs in rat testes.

The expressions of urokinase (uPA) and tissue-type plasminogen activators (tPA) in different stages of the rat seminiferous epithelial cycle were analyzed by in situ and Northern hybridizations combined with zymographic analysis. Irradiated rat testes were used to assess the cell localization. Both of the plasminogen activators were expressed in a strictly stage specific manner. Maximal expression of uPA mRNA was seen in Sertoli cells during stages VII-VIII of the cycle. The same expression in the basal compartment of the tubules was detected at 7 days post-irradiation (p-i), during a selective reduction of spermatogonia and preleptotene spermatocytes. Levels of tPA mRNA started to accumulate in Sertoli cells at stage VIII and were high during stages IX-XII and detectable during stages XIII-XIV. At 26 days p-i, reduction of pachytene spermatocytes, which are shown to be immunoreactive for tPA, did not have an effect on tPA mRNA expression. Catalytic activities of uPA and tPA changed concomitantly to their RNA levels in different stages of the cycle. However, at 7 days p-i, uPA activity was decreased at stages VII-VIII of the cycle suggesting that germ cell Sertoli cell interaction is important for uPA activity.

Animals↗
PubMed1994

Sex difference in the action of activin-A on cell proliferation of differentiating rat gonad.

The effect of recombinant bovine activin-A on DNA synthesis in fetal rat gonads and mesonephroi was studied in vitro by using [3H]thymidine autoradiography to evaluate the role of activin in the regulation of gonadal development. mRNA levels of inhibin-beta A and -beta B, activin receptor II and IIB (ActRII and ActRIIB), and follistatin were studied by Northern hybridization in the fetal rat gonads and mesonephroi. Activin stimulated thymidine incorporation in ovaries and female mesonephroi on days 14 and 15 postcoitum (pc) in a dose-dependent manner, as assessed by autoradiography. In contrast, activin inhibited thymidine incorporation in testes and male mesonephroi on day 14 pc in a dose-dependent way. On day 18 pc, the stimulatory effect of activin in ovary was nearly lost, whereas that in mesonephros was still pronounced. Activin had no effect on thymidine incorporation in testes and male mesonephroi on day 15 or 18 pc. Inhibin-beta A mRNA was seen in testes and mesonephroi in the male and in mesonephroi in the female of 15 and 18 day pc embryos. Inhibin-beta A mRNA expression was also seen in the testes and epididymes of newborn male rats and in the ovaries of 11-day-old postnatal rats. Inhibin-beta B mRNA was present in both testes and ovaries from 15 day pc onward. ActRIIB mRNA was most abundant in the testes, ovaries, and mesonephroi of 15 day pc embryos. The expression of ActRIIB message diminished during aging. ActRII mRNA was ubiquitously expressed during development. Follistatin mRNA was seen in the ovaries from 15 day pc onward and in the oviducts at 11 days postnatally. The present novel findings of activin subunit and receptor mRNA expression and activin action on gonadal and mesonephric cell proliferation suggest an important, sexually dimorphic role for this substance in fetal gonadal differentiation.

Activin Receptors↗
PubMed1994

Follicle-stimulating hormone regulation of inhibin alpha-subunit mRNA in staged rat seminiferous tubules.

Steady-state levels of inhibin-alpha and -beta B mRNAs are higher in stages II-VI of the seminiferous epithelial cycle than in stages VII-VIII. We investigated follicle-stimulating hormone (FSH) regulation of inhibin alpha-subunit mRNA in stages II-VI and VII-VIII to study whether the stage specificity is due to differential hormonal regulation by FSH. Follicle-stimulating hormone caused a significant increase of inhibin-alpha mRNA levels in both stages during a 20-h incubation. The mechanism of the FSH effect was studied further in stages VII-VIII. Maximal stimulation of the inhibin-alpha mRNA level was achieved with 100 micrograms/l FSH, dibutyryl-3',5'-cyclic adenosine monophosphate (db-cAMP, 0.2 mmol/l) and Sp-adenosine-3',5'-monophosphothionate (Sp-cAMPS, 10 mumol/l) (a cAMP agonist). The presence of Rp-cAMPS (200 mumol/l) (a cAMP antagonist) abolished the stimulation, Rp-cAMPS alone had no effect. Inhibin-beta B mRNA levels in stages VII-VIII were not affected by FSH, db-cAMP, Sp-cAMPS or Rp-cAMPS. Phorbol 12-myristate 13-acetate (100 nmol/l) had no effect on inhibin-alpha or -beta B mRNA levels. Actinomycin D abolished the stimulatory effect of FSH on inhibin-alpha mRNA expression. In conclusion, FSH stimulated inhibin-alpha mRNA expression similarly both in stages II-VI and VII-VIII of the seminiferous epithelial cycle and the stimulation in stages VII-VIII was cAMP-mediated.

Animals↗
PubMed1994

Effects of recombinant human FSH in immature hypophysectomized male rats: evidence for Leydig cell-mediated action on spermatogenesis.

The mode of FSH actions within the testis was studied in immature hypophysectomized male rats by treatment with recombinant human FSH (recFSH, Org 32489). To elucidate the involvement of Leydig cells and androgens in the maintenance of spermatogenesis in FSH-treated hypophysectomized rats further, the recFSH treatment was given both alone and after destruction of Leydig cells with ethane-1,2-dimethane sulphonate (EDS). Three days after hypophysectomy (at 31 days of age) the rats were given one i.p. injection of vehicle or EDS and, 4 days later, they were implanted with osmotic minipumps releasing either 0.9% (w/v) NaCI or 1 IU recFSH/day. Recombinant FSH alone increased testicular weights 2.5-fold in 7 days (P < 0.01). The effect of FSH was similar in EDS-pretreated rats (P < 0.01). Testicular testosterone increased from 6.5 +/- 1.6 to 16.9 +/- 5.3 (S.E.M.) pmol/g tissue (P < 0.05) and serum testosterone from 0.12 +/- 0.02 to 0.22 +/- 0.03 nmol/l (P < 0.05) when the rats were treated with recFSH. EDS alone did not affect testicular testosterone but, when combined with recFSH, it totally abolished the stimulatory effect of FSH on testosterone. Testicular binding of 125I-labelled iodo human chorionic gonadotrophin (hCG) and 125I-labelled iodo recFSH was increased 2.5- and 2.1-fold respectively with recFSH treatment (P < 0.01). EDS, either alone or with FSH, abolished specific testicular hCG binding (P < 0.01), but had no effect on that of recFSH. However, FSH increased its own receptors only in animals not treated with EDS.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Expression of inhibin alpha, beta A and beta B messenger ribonucleic acids in the normal human ovary and in polycystic ovarian syndrome.

We studied the cellular distribution of inhibin alpha, beta A and beta B mRNAs in the normal human ovary and in polycystic ovarian syndrome (PCOS) by in situ hybridization. Our results show that human granulosa cells express inhibin alpha, beta A and beta B subunit mRNAs, and theca cells express inhibin alpha and beta A subunit mRNAs. The co-localization of alpha and beta A mRNAs in theca cells supports the hypothesis that inhibin also has an autocrine function in these cells. We did not detect any inhibin subunit mRNA in the granulosa cells of atretic follicles, while theca cells also expressed alpha subunit mRNA in those follicles. The present findings suggest that the expression of inhibin subunits is regulated differently in human follicular granulosa and theca cells. It has been speculated that inhibin may be involved in the development of PCOS. Our results show that the cellular localization of inhibin subunit mRNAs is not disturbed in PCOS ovaries.

Female↗
PubMed1993

Localization of activin receptor (ActR-IIB2) mRNA in the rat seminiferous epithelium.

In situ hybridization was used to localize the mRNA expression of the high affinity activin receptor (ActR-IIB2) in the rat seminiferous epithelium. ActR-IIB2 mRNA was expressed maximally in stages IX-XI of the seminiferous epithelial cycle. The mRNA signal was detected basally in the epithelium in type A1 and A2 spermatogonia and in Sertoli cells. In the pubertal rat testis the expression was localized in Sertoli cells around primary spermatocytes and around meiotically dividing cells. The localization of ActR-IIB2 mRNA in spermatogonia lends support to the hypothesis that activin is a spermatogonial growth factor. The expression of activin receptor mRNA in pubertal rat testis suggests that activin may have a function during meiotic maturation.

Activin Receptors↗
PubMed1993

Polyamines and regulation of spermatogenesis: selective stimulation of late spermatogonia in transgenic mice overexpressing the human ornithine decarboxylase gene.

Polyamines are believed to participate in the induction of cell growth, differentiation, and proliferation, but their role in spermatogenesis has remained obscure. Two transgenic mouse lines (K2 and K15) that overexpress the human ornithine decarboxylase (ODC) gene coding for a rate-controlling enzyme in polyamine biosynthesis and, hence, contain high levels of tissue putrescine have been used to study the stage-specific role of ODC in spermatogenesis. In K2 mice with 30-fold testicular ODC overexpression, [3H]thymidine incorporation at stages I-VI of the cycle of the seminiferous epithelium was significantly above the control level. This may reflect a specific stimulation of DNA synthesis in type A4, intermediate, and type B spermatogonia. The K15 mice that have about 70-fold ODC overexpression showed an elevation of DNA synthesis only at stage V of the cycle, suggesting a specific dependence of type B spermatogonia on putrescine. In K15 mice, [3H]thymidine incorporation of stage VIII tubule segments was decreased, suggesting that excess amounts of putrescine selectively inhibit meiotic DNA synthesis. We propose that putrescine has strictly selective local stimulatory and inhibitory actions during spermatogenic DNA synthesis, and that its excess amounts ultimately may lead to decreased fertility.

Animals↗
PubMed1992

Expression of beta-nerve growth factor and its receptor in rat seminiferous epithelium: specific function at the onset of meiosis.

beta-Nerve growth factor (NGF) is expressed in spermatogenic cells and has testosterone-downregulated low-affinity receptors on Sertoli cells suggesting a paracrine role in the regulation of spermatogenesis. An analysis of the stage-specific expression of NGF and its low affinity receptor during the cycle of the seminiferous epithelium in the rat revealed NGF mRNA and protein at all stages of the cycle. Tyrosine kinase receptor (trk) mRNA encoding an essential component of the high-affinity NGF receptor was also present at all stages. In contrast, expression of low affinity NGF receptor mRNA was only found in stages VIIcd and VIII of the cycle, the sites of onset of meiosis. The low-affinity NGF receptor protein was present in the plasma membrane of the apical Sertoli cell processes as well as in the basal plasma membrane of these cells at stages VIIcd to XI. NGF was shown to stimulate in vitro DNA synthesis of seminiferous tubule segments with preleptotene spermatocytes at the onset of meiosis while other segments remained nonresponsive. We conclude that NGF is a meiotic growth factor that acts through Sertoli cells.

Animals↗
PubMed1992

Expression of inhibin beta A and beta B, follistatin and activin-A receptor messenger ribonucleic acids in the rat seminiferous epithelium.

The expression of inhibin beta A and beta B subunits, follistatin, and activin-A receptor messenger RNA (mRNAs) in different stages of rat seminiferous epithelial cycle was analyzed by in situ hybridization in order to understand their role in the regulation of spermatogenesis. Inhibin beta A mRNA was expressed in Sertoli cells in a highly stage-specific manner. The mRNA levels started to accumulate in Sertoli cells at stage VIII of the cycle and were highly expressed during stages IX-XI. Follistatin mRNA expression was identical to that of inhibin beta A, while inhibin beta B mRNA was maximally expressed in Sertoli cells at stages XIII-III. Low expression was found in stages VII-VIII. Activin-A receptor mRNA was localized mainly in spermatogenic cells. Maximal expression was seen in late primary spermatocytes at stages XIII-XIV and in early round spermatids at stages I-IV. A low even expression by Sertoli cells was also seen. Inhibin beta A and follistatin mRNAs were coexpressed in stage IX-XI Sertoli cells, suggesting close interplay between these molecules. The pattern of inhibin beta B mRNA expression was similar to that of inhibin alpha-mRNA. Localization of activin-A receptor mRNA in spermatogenic cells suggests that activin may influence meiotic divisions and early spermiogenesis.

Activin Receptors↗
PubMed1991

Immunocytochemical localization of follicle regulatory-protein (FRP) in testis.

Follicle regulatory protein (FRP) can exert paracrine control over follicular development. It is synthesized by the granulosa cells of the developing follicles and was localized in the cytoplasm of the mural cells by immunocytochemistry. When administered to male dogs and rats, FRP causes impairment of spermatogenesis. In the intact male rat, it has been postulated that FRP manifests its effects at a stage prior to the conversion of testosterone to dihydrotestosterone. Sertoli cells of the seminiferous tubules are implicated in testosterone metabolism. Furthermore, Sertoli cells in male gonads are regarded as the counterpart of granulosa cells in ovaries. The exact source of FRP in the male is not known. Therefore, it was of interest to study the localization of FRP in the male gonads. Testicular sections of the pig, dog, cat, rat, mouse, monkey, and man were immunocytochemically stained with monoclonal antibody to porcine FRP of ovarian origin. Sections of pig ovaries were used as controls throughout the study. Specificity of immunocytochemical localization was established by preabsorption. FRP antibody predominantly localized to the interstitial compartment of the pig testis. In the seminiferous tubules, FRP localization was limited to basal spermatogonia and Sertoli cells of tubules at few specific stages of spermatogenesis. The study also showed that the monoclonal antibody against porcine FRP is species-specific. Antibody binding was found only in pig testis, whereas tissues from the cat, dog, mouse, rat, monkey, and man did not display any immunocytochemical reaction.(ABSTRACT TRUNCATED AT 250 WORDS)

Aged↗
PubMed1991

Stage- and cell-specific gene expression and hormone regulation of the seminiferous epithelium.

The regulation of spermatogenesis seems to involve complex cell interactions in the testis. Little is known about these cellular communication events. Advances in molecular technology and cell or cell group separation methods have made it possible to analyze function of defined spermatogenic and Sertoli cells, thereby giving some insights into the paracrine regulation of spermatogenesis. In this review we will describe how seminiferous tubule segments with distinct cell associations can be rapidly isolated and how the cell composition can be modified by high-energy X-irradiation. Results of the recent studies performed using these techniques will be briefly summarized. Spermatogenic cells at defined stages of their development can be isolated in living condition for morphological and biochemical studies by the transillumination technique. For accurate identification of the stages of the seminiferous epithelial cycle, phase contrast microscopy of live cell squashes has been used. The criteria described by Leblond and Clermont (Am. NY Acad. Sci., 55:548-573, 1952) can be used for accurate recognition of most of the stages of the cycle. However, stages I and II and substages of VII that are important in several studies are difficult to distinguish. Therefore, in addition to the morphology of early spermatids, development of the flagella at step 16 of spermiogenesis and the changing morphology of the cytoplasmic lobes (residual bodies) at stage VII of the cycle were used as criteria for rapid identification and isolation (preparative) of the seminiferous tubule segments. Expression of nucleoprotein and heat shock protein 70-related protein genes was analyzed with Northern blot, slot blot, and in situ hybridization techniques in accurately staged seminiferous tubules. Accurate stage-dependent timing of the onset of transcription, followed by storage and disappearance of the messages was demonstrated. The chromatoid body (cb) has been proposed to have a specific function in storage of the long-lived mRNAs in the spermatids. It is an actively moving cytoplasmic organelle that interacts with Golgi complex during formation of the acrosomic system. The chromatoid body is apparently also dependent on cytoplasmic microtubules, since its movements are inhibited and its structure becomes abnormal in the presence of vincristin, an inhibitor of tubulin polymerization. Follicle-stimulating hormone (FSH) is an important regulator of Sertoli cell function. Since both basal and FSH-dependent cyclic AMP (cAMP) production by seminiferous tubules showed marked stage dependency, Sertoli cells are apparently influenced by spermatogenic cells. Thus, Sertoli cell function varies cyclically depending on the stage of the seminiferous epithelial cycle to provide an optimal microenvironment for spermatogenesis.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1991

Stage-specific cellular regulation of inhibin alpha-subunit mRNA expression in the rat seminiferous epithelium.

To find out the local regulation of inhibin production and its possible paracrine role in the seminiferous epithelium, inhibin alpha mRNA levels were measured in sequential 1 mm segments of rat seminiferous tubules accurately staged by transillumination technique. Highest levels were found at stages XIV-I-IV of the cycle, and lowest at stages VI-VIIb of the cycle. When dividing spermatogonia were selectively destroyed by 3 Gy of high-energy X-irradiation, stage-specific inhibin alpha mRNA levels remained unchanged until 26 and 38 days after irradiation when stages VII and VIII of the cycle showed 6- and 4-fold increases during a selective reduction of pachytene spermatocyte and round spermatid numbers, respectively. The results suggest that these cells at a strictly stage-specific fashion have a paracrine inhibitory effect on Sertoli cell inhibin alpha gene expression. Inhibin alpha mRNA level also correlates closely to the follicle-stimulating hormone-stimulated cAMP production during the cycle of the seminiferous epithelium, but does not seem to have a correlation to spermatogonial DNA synthesis.

Animals↗
PubMed1991

Production and secretion of an interleukin-1-like factor is stage-dependent and correlates with spermatogonial DNA synthesis in the rat seminiferous epithelium.

It has been shown previously that the intact adult rat testis produces large amounts of an interleukin-1 (IL-1)-like growth factor. The present study has investigated whether this testicular IL-1-like factor (tIL-1) is produced and secreted differentially by the fourteen stages of the seminiferous epithelial cycle in the rat testis. Seminiferous tubule segments representing defined stages were identified by transillumination-assisted microscopy and isolated by microdissection. Pooled segments were either homogenized and extracted with aqueous buffer or incubated for 24 h to produce conditioned media (CM). The recovered material was then analysed for IL-1 bioactivity in a sensitive murine thymocyte proliferation assay. When divided into four stage groups, extracts of stages II-VI, IX-XII and XIII-I showed equally high IL-1 activity whereas stage group VII-VIII showed much lower activity. More detailed analysis with 10 different stage groups showed that tIL-1 activity was undetectable in extracts of substages VIIab and VIIcd. The same pattern was seen when CM from cultured tubular segments were analysed. Labelling of seminiferous tubules with tritiated thymidine in vitro and analysis by autoradiography revealed that DNA-synthesizing spermatogonia were absent in substages VIIb and VIIc and sparse in substages VIIa and VIId. The results show that tIL-1 activity is produced in a stage-dependent manner and suggest that tIL-1 might be involved in the regulation of spermatogonial proliferation in vivo.

Animals↗
PubMed1990

Stage- and cell-specific expression of the ornithine decarboxylase gene during rat and mouse spermatogenesis.

Ornithine decarboxylase (ODC) is an enzyme that has been shown to be induced in the growth, differentiation and proliferation of cells. We have used a cDNA probe to determine ODC mRNA levels in different stages of the cycle of rat and mouse seminiferous epithelium. For Northern and slot-blot hybridizations, RNA was isolated from microdissected staged seminiferous tubules. Cell-specific localization of ODC mRNA was studied by in situ hybridization. In the rat, in situ hybridization showed increasing mRNA levels during prophase of meiosis with the highest mRNA levels seen in late pachytene spermatocytes and step 3-5 spermatids. In the mouse, the mRNA levels increased in a similar fashion and the highest mRNA levels were found in step 1-8 spermatids. In the rat, Northern blot hybridizations revealed three molecular sizes of ODC mRNA: 2.2, 2.7 and 1.6 kb. The levels of all molecular sizes were highest in stages VII-VIII, and the lowest mRNA levels were seen in stage I of the seminiferous epithelial cycle. The level of the 2.2 kb transcript was low during stages XIII-I. In the mouse, the Northern blot hybridizations also showed three molecular sizes of ODC mRNA: 2.2 and 2.7 kb and very low levels of 1.6 kb transcript. The levels of the transcripts were steady throughout the cycle. In the mouse, the 2.2 kb transcript was more abundant than the 2.7 kb transcript indicating a species difference between rat and mouse in the usage of the two polyadenylation signals within the ODC gene.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

Cellular regulation of basal and FSH-stimulated cyclic AMP production in irradiated rat testes.

Basal and follicle-stimulating hormone (FSH)-stimulated cyclic AMP (cAMP) productions by seminiferous tubular segments from irradiated adult rats were investigated at defined stages of the epithelial cycle when specific spermatogenic cells were low in number. Seven days post-irradiation, depletion of spermatogonia did not influence the basal cAMP production, but FSH response increased in stages II-VIII. Seventeen days post-irradiation when spermatocytes were low in number, there was a small increase in basal cAMP level in stages VII-VIII and FSH-stimulated cAMP production increased in stages VII-XII and XIII-I. At 38 days when pachytene spermatocytes and round spermatids (steps 1-6) were low in number, a decreased basal cAMP production was measured in stages II-VI and IX-XII. FSH-stimulated cAMP output increased in stages VII-XII but decreased in stages II-VI. At 52 days when all spermatids were low in number, basal cAMP levels decreased in all stages of the cycle, whereas FSH response was elevated only in stages VII-XII. All spermatogenic cell types seem to have an effect on cAMP production by the seminiferous tubule in a stage-specific fashion. Germ cells appear to regulate Sertoli cell FSH response in a paracrine way, and a part of cAMP may originate from spermatids stimulated by an unknown FSH-dependent Sertoli cell factor. The FSH-dependent functions may control such phenomena as spermatogonial proliferation, final maturation of spermatids, and onset of meiosis.

Animals↗