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M Parvinen

Publications and source records attributed to M Parvinen.

At least 37 records · Page 2Linked to original sources
PubMed1999

Function of stem cell factor as a survival factor of spermatogonia and localization of messenger ribonucleic acid in the rat seminiferous epithelium.

To address the possibility that stem cell factor (SCF) is a paracrine regulator of germ cell development in the adult rat testis, stage-specific distribution of SCF messenger RNA (mRNA) was investigated with Northern blot and in situ hybridization analyses. The highest levels of SCF mRNA were found in stages II-VI of the rat seminiferous epithelial cycle, whereas the lowest levels were in stages VII-VIII. Intermediate levels of SCF mRNA were detected in stages IX-XIV-I of the cycle. The expression of the SCF gene was found to be developmentally regulated, and the expression pattern followed the process of Sertoli cell proliferation and differentiation during postnatal life. The effect of mouse recombinant SCF on spermatogonial DNA synthesis was studied using an in vitro tissue culture system for stage-defined seminiferous tubules. A significant increase in DNA synthesis in spermatogonia could be detected when tubule segments from stage XII were cultured in the presence of 100 ng/ml SCF for 48 h (P < 0.05) and 72 h (P < 0.01). This observation was further confirmed with autoradiographic analyses; almost a 100-fold increase in thymidine incorporation in the SCF-treated (100 ng/ml) tubule segments was observed compared with that in untreated samples. The results of the present study suggest that SCF is a Sertoli cell-produced paracrine regulator and acts as a survival factor for spermatogonia in the adult rat seminiferous epithelium in a stage-specific manner.

Animals↗
PubMed1999

Constitutive expression of interleukin-1alpha messenger ribonucleic acid in rat Sertoli cells is dependent upon interaction with germ cells.

Interleukin-1 (IL-1), a proinflammatory cytokine originally isolated as a product of activated mononuclear phagocytes, consists of two distinct agonist proteins, IL-1alpha and IL-1beta, of which IL-1beta is the major inducible IL-1 protein produced by macrophages. We show here that mRNA of IL-1alpha, but not IL-1beta, is constitutively expressed by the intact rat testis and localize the transcript to Sertoli cells as confirmed by a novel squash technique. The expression is developmentally regulated and appears only after postnatal day 20 in the rat testis, corresponding to onset of puberty. IL-1alpha mRNA shows a stage-dependent expression pattern during the cycle of the seminiferous epithelium. It is low or absent in stage VII, but present in all other stages of the cycle. The same stage-dependent distribution was also observed at the protein level when bioactive IL-1 was measured in extracts of accurately defined one millimeter segments of seminiferous tubules. No IL-1alpha mRNA was detected in adult rat testes after germ cell depletion by fetal irradiation or cytostatic drug treatment. Because stage VII is the only segment of the seminiferous tubules lacking DNA replication, we propose that IL-1alpha is involved in this event during mitosis and meiosis of spermatogenesis and that its expression is dependent upon interactions between Sertoli cells and germ cells.

Aging↗
PubMed1999

Partial oxygen pressure and mitochondrial permeability transition affect germ cell apoptosis in the human testis.

During regular spermatogenesis, a number of testicular germ cells degenerate by an apoptotic process that is under hormonal control. Oxidative and mitochondrial changes have been proposed to play a role in apoptosis of many cell types. Previously, whether human germ cell survival is controlled by oxygen or by effectors of the mitochondrial permeability transition has not been investigated. In the present study, apoptosis was induced in human testicular germ cells by incubating segments of seminiferous tubules without survival factors (i.e. serum or hormones; 21% oxygen). Apoptosis was significantly suppressed in an inversely dose-dependent fashion at partial oxygen pressures below 10%, as detected by Southern blot analysis of DNA fragmentation, DNA labeling in situ, and electron microscopy. Cyclosporin A and its nonimmunosuppressive derivative N-methyl-Val4-cyclosporin A prevented cell death, suggesting a key role for the mitochondrial permeability transition in apoptosis. Apoptotic cells were identified as mainly spermatocytes and spermatids, the mitochondria of which underwent morphological changes during the apoptotic process. The present results imply that to improve germ cell viability in in vitro fertilization techniques, the partial oxygen pressure should be lowered.

Aged↗
PubMed1998

Overexpression of VEGF in testis and epididymis causes infertility in transgenic mice: evidence for nonendothelial targets for VEGF.

Vascular endothelial growth factor (VEGF) is a key regulator of endothelial growth and permeability. However, VEGF may also target nonendothelial cells, as VEGF receptors and responsiveness have been detected for example in monocytes, and high concentrations of VEGF have been reported in human semen. In this work we present evidence that overexpression of VEGF in the testis and epididymis of transgenic mice under the mouse mammary tumor virus (MMTV) LTR promoter causes infertility. The testes of the transgenic mice exhibited spermatogenic arrest and increased capillary density. The ductus epididymidis was dilated, containing areas of epithelial hyperplasia. The number of subepithelial capillaries in the epididymis was also increased and these vessels were highly permeable as judged by the detection of extravasated fibrinogen products. Intriguingly, the expression of VEGF receptor-1 (VEGFR-1) was detected in certain spermatogenic cells in addition to vascular endothelium, and both VEGFR-1 and VEGFR-2 were also found in the Leydig cells of the testis. The infertility of the MMTV-VEGF male mice could thus result from VEGF acting on both endothelial and nonendothelial cells of the male genital tract. Taken together, these findings suggest that the VEGF transgene has nonendothelial target cells in the testis and that VEGF may regulate male fertility.

Animals↗
PubMed1998

Stage-specific expression and cellular localization of the heat shock factor 2 isoforms in the rat seminiferous epithelium.

Heat shock transcription factors (HSFs) are generally known as regulators of cellular stress response. The mammalian HSF1 functions as a classical stress factor, whereas HSF2 is active during certain developmental processes, including embryogenesis and spermatogenesis. In the present study, we examined HSF2 expression at specific stages of the rat seminiferous epithelial cycle. We found that expression of the alternatively spliced HSF2-alpha and HSF2-beta isoforms is developmentally regulated in a stage-specific manner. Studies on cellular localization demonstrated that HSF2 is present in the nuclei of early pachytene spermatocytes at stages I-IV and in the nuclei of round spermatids at stages V-VIIab. In contrast a strong HSF2 immunoreactivity was detected in small distinct cytoplasmic regions from zygotene spermatocytes to maturation phase spermatids. Immunoelectron microscopic analysis revealed that these structures are mainly cytoplasmic bridges between germ cells. Our results on cellular localization of HSF2 and stage-specific expression of the HSF2 isoforms indicate that HSF2, in addition to its function as a nuclear transcription factor, may be involved in other cellular processes during spermatogenesis, possibly in the sharing process of gene products between the germ cells.

Animals↗
PubMed1998

Stage-specific apoptosis of male germ cells in the rat: mechanisms of cell death studied by supravital squash preparations.

Apoptosis has been proposed as a mechanism by which testis germ cells are removed during normal and various pathological conditions. To establish a new rapid way to detect stage-specific apoptosis in male rat germ cells, their supravital morphology was examined from carefully squashed monolayers of living cells, after several established toxic treatments, using a phase contrast microscope. The results were compared with early detection of apoptosis using annexin V and propidium iodide (PI) stainings. The apoptosis of type-A spermatogonia and round spermatids proceeded in a similar way to somatic cells, while intermediate and type-B spermatogonia, and particularly the dividing spermatocytes, possessed characteristics not entirely typical for apoptosis. Death of elongated spermatids was difficult to assess owing to their compacted chromatin. As the first phases of degeneration seemed different in various germ cell classes, the final stage (karyopycnosis) was similar for most cells. Degenerating cells also showed positive reactions for annexin V and PI. The 'living cell method' provides rapid and accurate possibilities for analysis of stage-specific apoptosis during spermatogenesis. This method is not influenced by artefacts induced by fixation, embedding and sectioning. It may be developed further for routine analyses of the accurate stage-specific effects of various physical and chemical effects on mammalian and human spermatogenesis.

Animals↗
PubMed1998

Histochemical tracing of zinc ions in the rat testis.

To detect free zinc ions in the rat testes four rats were transcardially perfused with Na2S, and the seminiferous tubules from two other rats were incubated in Na2S. Sections from the two sources were autometallographically (AMG) developed, whereby zinc sulphide crystal lattices created in the tissue by the sulphide treatment were silver enhanced. Light microscopical analysis showed zinc ions in primary spermatogonia until the zygotene primary spermatocytes (stage I), in late pachytene spermatocytes (stages XII and XIII), and in late spermatids from step 15 to step 19 (stages I-VIII). The highest intensity of AMG grains was detected in the residual bodies and tails of step 19 spermatids. Grains were occasionally found in the cytoplasm of Leydig cells. Sections from animals treated with the chelator diethyldithiocarbamate prior to sulphide treatment showed a complete lack of AMG staining. At ultrastructural levels the AMG grains were found in smooth-surfaced endoplasmic reticulum of all spermatogonial stages, and in the acrosome, midpiece, and tail of late spermatids. The presence of zinc ions in preleptotene spermatocytes and cytoplasmic lobes of late spermatids suggests a specific role of free zinc at the onset of meiosis and at spermiation.

Animals↗
PubMed1998

Differential regulation of leucine-rich primary response gene 1 (LRPR1) mRNA expression in rat testis and ovary.

In immature rat Sertoli cells, leucine-rich primary response gene 1 (LRPR1) represents a follicle stimulating hormone (FSH)-responsive gene; the function of the encoded protein is not yet known. LRPR1 mRNA expression is up-regulated very rapidly and specifically by FSH, both in cultured Sertoli cells and in vivo in regulation in more detail, in testis and ovary of fetal, immature, and adult rats. In addition, we have studied the expression of FSH receptor (FSHR) mRNA in relation to LRPR1 mRNA expression. In rat testis, LRPR1 mRNA and FSHR mRNA followed a similar expression pattern, during postnatal development and also at different stages of the spermatogenic cycle in the adult rat. Furthermore, after short-term challenge of the FSH signal transduction pathway in intact immature rats by injection with a relatively high dose of FSH, an inverse relationship between LRPR1 mRNA (up-regulation) and FSHR mRNA expression (down-regulation) was observed. Similar studies in the ovary provided completely different results. LRPR1 mRNA in the postnatal ovary is present well before expression of FSHR mRNA can be first detected. In addition, incubation of ovaries of immature rats with FSH or dibutyryl cyclic AMP (dbcAMP) did not result in up-regulation of LRPR1 mRNA expression. During fetal development, the LRPR1 mRNA expression pattern involved many more tissues, in contrast to the relatively tissue-specific expression of LRPR1 mRNA in gonads of 21 day old and adult rats. Moreover, LRPR1 mRNA expression could be detected as early as 12.5 days post-coitum, whereas FSHR mRNA is absent at this stage of fetal development. We concluded that the pronounced regulation of LRPR1 by FSH observed in the immature rat testis does not occur in the ovary. Furthermore, in the ovary LRPR1 mRNA expression does not appear to be dependent on FSH action. Finally, the LRPR1 gene product may play a general role during fetal development.

Animals↗
PubMed1998

Elevated expression of lanosterol 14alpha-demethylase (CYP51) and the synthesis of oocyte meiosis-activating sterols in postmeiotic germ cells of male rats.

Mammalian CYP51 encodes lanosterol 14alpha-demethylase (P45014DM) that is involved in the postsqualene part of cholesterol biosynthesis. This enzyme removes the 14alpha-methyl group from lanosterol and 24,25-dihydrolanosterol producing intermediates in cholesterol biosynthesis, the oocyte meiosis-activating sterols FF-MAS and MAS-412. Human and rat CYP51 messenger RNAs (mRNAs) are expressed in all tissues, with highest levels in the testis due to the presence of an additional shorter CYP51 transcript in this tissue. In situ hybridization shows the highest CYP51 mRNA levels in seminiferous tubules, with only background levels in Leydig cells. The rat testis-specific CYP51 mRNA arises from the use of an upstream polyadenylation site and is restricted to germ cells, being most abundant in elongating spermatids in stages VII-XIV, whereas somatic CYP51 transcripts are present in all cells. In contrast, the mRNA levels of squalene synthase are maximal in round spermatids, and no germ cell-specific transcript is observed. The rat male germ cell-specific CYP51 transcript is translated in vitro to two proteins of approximately 55 and 53.5 kDa. CYP51 activity is higher in protein extracts of testes and germ cells of sexually mature rats than in prepubertal animals, in which postmeiotic germ cells are not yet present. This shows increased capacity for the production of MAS sterols by male germ cells that have already completed meiosis, suggesting that they serve a role different from meiosis activation.

Animals↗
PubMed1998

N-acetyl-L-cysteine inhibits apoptosis in human male germ cells in vitro.

Antioxidant defenses play a critical role in the regulation of programmed cell death, even when death is induced by nonoxidative stimuli. During spermatogenesis, most of the testicular germ cells degenerate by an apoptotic process that is under hormonal control. However, the exact mechanisms by which hormonal signals are transduced within the cells to direct their life, and whether other effectors of the apoptotic pathway, for example antioxidants, take part in the control of human germ cell survival, are not known. In the present study, testosterone and N-acetyl-L-cysteine (NAC), which is an antioxidant, an inhibitor of apoptosis in several systems, and a survival factor in human semen, were found to suppress programmed cell death in human testicular germ cells in vitro. The samples came from adult men undergoing orchidectomy for prostate cancer. Germ cell death was induced by incubating segments of seminiferous tubules under serum-free culture conditions. This apoptosis, detected by Southern blot analysis of DNA fragmentation, by DNA labeling in situ, and by morphological analysis under the electron microscope, was significantly inhibited by testosterone at concentrations of 10(-6) and 10(-7) mol/L. NAC concentrations of 125, 100, 50, and 25 mmol/L suppressed germ cell death in a dose-dependent manner. This inhibition was effective during 4, 24, and 48 h of incubation. Apoptotic cells were identified mainly as spermatocytes and early spermatids. Programmed cell death was also demonstrated in late spermatids. We conclude that NAC, which is an antioxidant, plays an important role in germ cell survival in the human seminiferous tubules in vitro. We also suggest NAC as a possible new therapeutic factor for some men with idiopathic oligospermia.

Acetylcysteine↗
PubMed1997

The in vivo expression pattern of mouse Nek2, a NIMA-related kinase, indicates a role in both mitosis and meiosis.

The human protein kinase Nek2 is related to the NIMA cell cycle regulatory kinase of Aspergillus nidulans. Whereas NIMA has been shown to be essential for cell cycle progression into mitosis in this fungus, the function of mammalian Nek2 remains to be elucidated. In this study, we isolated a cDNA coding for a mouse ortholog of human Nek2 and analyzed the expression of this kinase in different organs. RNase protection assays performed on RNAs from mouse adult organs showed very high expression of Nek2 in testis. Lower levels of transcripts were detected in intestine, thymus, and skin, three mitotically active organs, and whole-mount in situ hybridization performed on 10.5-day embryos allowed the detection of Nek2 transcripts in the brain. In situ hybridization analysis of testis sections revealed that the transcription of Nek2 occurred in a stage-specific manner during spermatogenesis. The strongest signals were seen in cells undergoing meiosis, but Nek2 transcripts could also be detected in haploid cells (stage I and II spermatids). Extending these results, in situ hybridization performed on ovary sections revealed strong signals in meiotically active oocytes. In addition, some Nek2 transcription was observed in actively dividing follicle cells surrounding the oocytes and in the oviduct. Finally, indirect immunofluorescence staining of testis sections with Nek2-specific antibodies confirmed that this kinase is highly expressed in spermatocytes and, to a lesser extent, in early spermatids. Taken together, these results indicate that Nek2 may play an important role not only during mitosis but also during meiosis.

Amino Acid Sequence↗
PubMed1997

Apoptosis in human acute myocardial infarction.

BACKGROUND: After reopening of the infarct-related coronary artery, cardiomyocytes continue to die during reperfusion. The mechanisms of cell death have been subject to debate. We studied whether an apoptotic type of cell death occurs in human acute myocardial infarction (AMI). METHODS AND RESULTS: We studied myocardial samples of eight patients who died of AMI and had patent infarct-related arteries at autopsy. Six of the patients had received initially successful thrombolysis. Extensive formation of DNA strand breaks, the typical biochemical feature of apoptosis, was detected with the use of the in situ DNA end-labeling method. Apoptotic cardiomyocytes were observed particularly in the border zones of histologically infarcted myocardium, whereas very few apoptotic cells were present in the remote noninfarcted myocardium. Internucleosomal fragmentation was confirmed by agarose gel electrophoresis of DNA isolated from the representative myocardial areas. CONCLUSIONS: This study provides evidence that in addition to overt necrosis, a subset of myocytes undergo apoptosis during ischemia-reperfusion injury. Apoptosis may provide a new target for cardioprotection during evolving AMI in humans.

Adult↗
PubMed1997

Computer analysis of living cells: movements of the chromatoid body in early spermatids compared with its ultrastructure in snap-frozen preparations.

Quantitative analyses of cytoplasmic and nuclear organelle movements in living interphase cells at defined stages of differentiation are few. By phase contrast videomicroscopy and digital imaging techniques, we have traced the path of the chromatoid body (CB) and analysed its rapidly changing positions in relation to the nuclear envelope, Golgi complex and nuclear pale chromatin areas in living early spermatids of the rat. The CB had intimate interactions with the nuclear envelope and moved both in parallel and perpendicular fashion in relation to it. It had successive short contacts with the Golgi complex and nuclear pale chromatin areas. It was also seen to scan between two pale chromatin areas and it had pinocytosis-like transient engulfments during interactions with the pale chromatin. In ultrastructural analysis of snap-frozen preparations, the CB had a large contact area with the nuclear envelope with several intermediate organelles that may be involved in nucleocytoplasmic material transport. It is evident that quantitative image analysis of living cells is a powerful guide for ultrastructural analyses. The snap-freezing technique gives new possibilities for studies of structures that are sensitive to conventional fixation procedures.

Animals↗
PubMed1997

Stage-specific degeneration of germ cells in the seminiferous tubules of non-obese diabetic mice.

The cause of fertility problems in insulin-dependent diabetes is largely unknown. To evaluate the role of autoimmunity-associated phenomena in the testis as a possible cause of the derangement in spermatogenesis, the stage-specific apoptosis of germ cells in the insulitis phase of pre-diabetes was quantified in the testes of non-obese diabetic (NOD) mice. The seminiferous epithelium of normal BALB/c and NOD mice contained cells positive for in-situ end-labelling (ISEL) of DNA. ISEL-positive germ cells formed clusters in the seminiferous epithelium of the NOD mice in marked contrast to the seminiferous epithelium of the BALB/c mice, which contained only individual cells positive for ISEL. ISEL-positive cells were present in the basal and luminal compartments of the epithelium. Ultrastructural analysis and demonstration of externalized phosphatidyl serine confirmed that the cells were undergoing apoptosis. The ultrastructurally apoptotic cells included spermatogonia, spermatocytes and spermatids. In cytological squash preparations of segments of seminiferous tubules from NOD mice aged 17-20 weeks, the number of ISEL-positive cells/mm tubule was significantly lower in segments at stages I-II of the seminiferous epithelial wave but higher at stages III-IV in comparison to BALB/c mice. The numbers of ISEL-positive cells/mm tubule in the other stages were similar in the two strains of mice. Analysis of 32P-3'-end labelled DNA from the testes showed that the BALB/c mice had relatively more DNA fragmentation than did the NOD mice. These data suggest that autoimmune insulitis in the NOD mice is associated with increased amounts and abnormal stage distribution of apoptosis in the seminiferous epithelium, resulting in derangement of spermatogenesis.

Animals↗
PubMed1997

The transcriptional and translational control of diazepam binding inhibitor expression in rat male germ-line cells.

The diazepam binding inhibitor [DBI, also known as acyl-CoA-binding protein, (ACBP), or endozepine] is a 10-kD protein that has been suggested to be involved in the regulation of several biological processes such as acyl-CoA metabolism, steroidogenesis, insulin secretion, and gamma-aminobutyric acid type A (GABA(A))/benzodiazepine receptor modulation. DBI has been cloned from vertebrates, insects, plants, and yeasts. In mammals, DBI is expressed in almost all the tissues studied. Nevertheless, DBI expression is restricted to specific cell types. Here we have studied DBI gene expression in the germ-line cells of rat testis. The DBI gene was intensively transcribed in postmeiotic round spermatids from stages VI to VIII of the seminiferous epithelial cycle. A prominent, spermatid-specific upstream transcription initiation site was identified in addition to the multiple common transcriptional initiation sites found in the somatic tissues. However, no DBI protein was detected in round spermatids, suggesting that the DBI transcripts were translationally arrested. The DBI protein was detected in the late spermatogenic stages starting from elongating spermatids from step 18 (stage VI) onward. The DBI protein was also detected in mature spermatozoa and in ejaculated human sperms. The majority of DBI was located at the middle piece of the spermatozoons tail enriched with mitochondria. On the basis of this observation and the well-established role of DBI in acyl-CoA metabolism, we propose that DBI expression in spermatozoa reflects the usage of fatty acids as a primary energy source by spermatozoa. The biological function of DBI in spermatozoa could thus be related to the motility function of sperm.

Animals↗
PubMed1997

Testosterone regulates apoptosis in adult human seminiferous tubules in vitro.

In the present study an in vitro model was developed and characterized for evaluation of the role of apoptosis in adult human testes. The samples came from adult men undergoing orchidectomy for prostate or testicular cancer. Segments of seminiferous tubules were isolated and incubated under serum-free conditions in the absence or presence of testosterone. Apoptosis was assessed by low mol wt DNA fragmentation (185-bp multiples) by use of 3'-end-labeled DNA, in situ end labeling, and morphological detection under light and electron microscopy. During the 4-h incubation, a 15-fold increase was seen in apoptotic DNA fragmentation. The extent of low mol wt DNA showed a time-dependent increase and reached a 20-fold intensity in 24 h of incubation compared to the level at 0 h. Apoptosis was significantly suppressed by testosterone concentrations of 10(-7) and 10(-6) mol/L during the first 4 h of incubation. Apoptotic cells were identified mainly as spermatocytes and occasionally as spermatids. We conclude that apoptosis is induced in human seminiferous tubules under serum-free conditions in vitro. That this apoptosis is suppressed by testosterone indicates that testosterone in the human male is a critical germ cell survival factor. The model created in the present study provides a valuable tool for further investigation of hormonal and gene regulation of human germ cell death and survival.

Adult↗
PubMed1996

Sequential expression of nucleoproteins during rat spermiogenesis.

Transition proteins and protamines are highly basic sperm-specific nuclear proteins that serve to compact the DNA during late spermiogenesis. To understand their sequential role in this function, transition protein 1 (TP1), transition protein 2 (TP2), and protamine 1 (P1) were assayed by polyacrylamide gel electrophoresis in pools of microdissected, staged seminiferous tubule segments in the rat. The results were compared with immunocytochemical analyses of squash preparations from accurately identified stages of the epithelial cycle. TP2 was the first to appear as a faint band at stages IX-XI, followed by high levels at stages XII-XIV of the cycle. TP1 showed a low expression at stage XII of the cycle and peaked at stages XIII-I, whereas protamine 1 first appeared at stage I of the cycle and remained high throughout the rest of spermiogenesis. Immunocytochemical analyses and Western blots largely confirmed these results: TP2 in steps 9-14, TP1 in steps 12-15, and P1 from late step 11 to step 19 of spermiogenesis. We propose that TP2 is the first nucleoprotein that replaces histones from the spermatid nucleus, and its appearance is associated with the onset of nuclear elongation. TP1 shows up along with the compaction of the chromatin. The two transition proteins seem to have distinct roles during transformation of the nuclei and compaction of spermatid DNA.

Animals↗