Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 8 de 8
Filter
Add more filters










Database
Language
Publication year range
1.
Environ Sci Pollut Res Int ; 28(21): 26606-26616, 2021 Jun.
Article in English | MEDLINE | ID: mdl-33495953

ABSTRACT

Many toxicants are present in water as a mixture. Male infertility is one of the environmental impacts in developed countries. Using our rat seminiferous tubule culture model, we evaluated the effects of waters of different origins, on several parameters of the seminiferous epithelium. Concentrated culture medium was diluted with the waters to be tested (final concentrations of the tested waters were between 8 and 80%). The integrity of the blood-testis barrier was assessed by the trans-epithelial electric resistance (TEER). The levels of mRNAs specific of Sertoli cells, of cellular junctions, of each population of germ cells, of androgen receptor, of estrogen receptor α, and of aromatase were also studied. We report, here, the results obtained with ten waters, some of them possessing a negative effect on spermatogenesis. The results showed that, according to the tested waters, their effects on the parameters studied might be quite different indicating many different mechanisms of toxicity, including some endocrine-disrupting effects. It has been reported that men with impaired semen parameters have an increased mortality rate suggesting semen quality may provide a fundamental biomarker of overall male health. Hence, we have developed a relevant in vitro bioassay allowing the evaluation of the potential toxicity of different types of waters on male fertility and to assess some aspects of their mechanism of action. In addition to the TEER measure, the number and/or the identity of the studied mRNAs can be largely increased and/or modified, thus enhancing the possibility of using this model as a "warning system."


Subject(s)
Semen Analysis , Spermatogenesis , Animals , Biological Assay , Blood-Testis Barrier , Humans , Male , Pilot Projects , Rats , Sertoli Cells , Testis
2.
Toxicol In Vitro ; 62: 104699, 2020 Feb.
Article in English | MEDLINE | ID: mdl-31689476

ABSTRACT

Atrazine (ATZ), a widely used agricultural pesticide and benzo[a]pyrene (BaP), a ubiquitous environmental human carcinogen can induce alterations of spermatogenesis. In the present study, we showed first that our seminiferous tubule culture model, in bicameral chambers, allowed the settlement of the blood-testis barrier (BTB) in 8-day-old male rat cultures and the differentiation of spermatogonia into round spermatids.The effect of a mixture of 1 µg/L of ATZ and 1 µg/L of BaP was then investigated either during or after the establishment of the BTB by using 8- or 20-22-day-old rats. Cultures were performed over a 3-week period. Our results show that claudin-11 and connexin 43 two proteins of the BTB, were impaired by the mixture which also reduced the number of round spermatids (the direct precursors of spermatozoa), by targeting the middle to late pachytene spermatocytes. These effects were observed in 8- and 20-22-day -old rat seminiferous tubule cultures. However, the decrease of the number of round spermatids was faster and more marked in the 8-day- than in the 20-22-day -old rat seminiferous tubule cultures. Our study emphasizes the possible influence of the age of an individual on the effect of (a) toxicant(s) on spermatogenesis.


Subject(s)
Atrazine/toxicity , Benzo(a)pyrene/toxicity , Blood-Testis Barrier/drug effects , Environmental Pollutants/toxicity , Herbicides/toxicity , Seminiferous Epithelium/drug effects , Seminiferous Tubules/drug effects , Animals , Cells, Cultured , Male , Organ Culture Techniques , Rats , Rats, Sprague-Dawley , Spermatids/drug effects , Spermatogenesis , Spermatogonia/drug effects
3.
Toxicol In Vitro ; 45(Pt 3): 366-373, 2017 Dec.
Article in English | MEDLINE | ID: mdl-28576679

ABSTRACT

It has been shown that non-cytotoxic doses of Carbendazim (CBZ), a broad-spectrum benzimidazole fungicide, possess endocrine-disrupting (androgen-like) actions, ex vivo, on the pubertal rat seminiferous epithelium. Iprodione (IPR), a dicarboximide fungicide, is also known to be an endocrine-disrupter (anti-androgen). The effect of a mixture of these two pesticides was investigated in the validated rat seminiferous tubule culture model. Cultures were performed in the absence or presence of CBZ 50nM or IPR 50nM either alone or in mixture (Mix), over a 3-week period. Mix exerted a dramatic effect on two proteins (Connexin 43 and Claudin-11) of the blood-testis barrier and possessed similar effects to IPR on some germ cell populations. The presence of IPR together with CBZ (Mix) cancelled the effect of CBZ on the increase of the androgen-dependent TP1 and TP2 mRNAs and on the decrease of ERα, ERß mRNAs. Nevertheless, CBZ alone or IPR alone or Mix induced toxicity on spermatogenesis resulting in a decrease of round spermatids (the precursors of spermatozoa). These results strongly suggest that, even at these low concentrations, the effects of IPR and of CBZ are not solely dependent on their respective anti-androgenic and androgen-like effects and should involve several mechanisms of action.


Subject(s)
Aminoimidazole Carboxamide/analogs & derivatives , Benzimidazoles/toxicity , Carbamates/toxicity , Endocrine Disruptors/toxicity , Fungicides, Industrial/toxicity , Hydantoins/toxicity , Seminiferous Epithelium/drug effects , Aminoimidazole Carboxamide/toxicity , Animals , Blood-Testis Barrier/drug effects , Cells, Cultured , Claudins/biosynthesis , Claudins/genetics , Connexin 43/biosynthesis , Connexin 43/genetics , Gene Expression Regulation/drug effects , Male , Rats , Rats, Sprague-Dawley , Sexual Maturation , Spermatocytes/drug effects , Spermatogenesis/drug effects
4.
Biol Reprod ; 95(4): 89, 2016 10.
Article in English | MEDLINE | ID: mdl-27580986

ABSTRACT

Until now, complete ex vivo spermatogenesis has been reported only in the mouse. In this species, the duration of spermatogenesis is 35 days, whereas it is 54 days in the rat and 74 days in humans. We performed long-term (until 60 days) cultures of fresh or frozen rat or human seminiferous tubule segments in a bioreactor made of a hollow cylinder of chitosan hydrogel. Testicular tissues were obtained from 8- or 20-day-old male rats or from adult human subjects who had undergone hormone treatments leading to a nearly complete regression of their spermatogenesis before bilateral orchiectomy for gender reassignment. The progression of spermatogenesis was assessed by cytological analyses of the cultures; it was related to a dramatic increase in the levels of the mRNAs specifically expressed by round spermatids, Transition protein 1, Transition protein 2, and Protamine 3 in rat cultures. From 2% to 3.8% of cells were found to be haploid cells by fluorescence in situ hybridization analysis of human cultures. In this bioreactor, long-term cultures of seminiferous tubule segments from prepubertal rats or from adult men allowed completion of the spermatogenic process leading to morphologically mature spermatozoa. Further studies will need to address the way of optimizing the yield of every step of spermatogenesis by adjusting the composition of the culture medium, the geometry, and the material properties of the chitosan hydrogel bioreactors. Another essential requirement is to assess the quality of the gametes produced ex vivo by showing their ability to produce normal offspring (rat) or their biochemical normality (human).


Subject(s)
Spermatogenesis/physiology , Testis/cytology , Adult , Animals , Bioreactors , Chitosan , Cryopreservation/methods , Culture Media , Humans , Hydrogels , In Situ Hybridization, Fluorescence , In Vitro Techniques , Male , RNA, Messenger/genetics , RNA, Messenger/metabolism , Rats , Rats, Sprague-Dawley , Seminiferous Tubules/cytology , Seminiferous Tubules/metabolism , Species Specificity , Spermatids/cytology , Spermatids/metabolism , Spermatogenesis/genetics , Testis/metabolism
5.
Obesity (Silver Spring) ; 16(8): 1763-9, 2008 Aug.
Article in English | MEDLINE | ID: mdl-18551122

ABSTRACT

The objectives of this study were to identify potential alterations in gene expression of melanocortin-4 receptor (MC4-R), proopiomelanocortin (POMC), and Agouti-related protein (AgRP) in mouse hypothalamus under a chronic peripheral infusion of leptin or at early (8 weeks) and advanced (16 weeks) phases of diet-induced obesity. Control or diet-induced obesity mice (8 or 16 weeks of high-fat diet) were either treated or not treated with leptin. Metabolic features were analyzed and expression of the genes of interest was measured by quantitative reverse transcriptase-PCR (RT-qPCR) and western blot. We reported that in control mice, but not in obese mice, leptin infusion induced an increase in POMC mRNA level as well as in MC4-R mRNA level suggesting that leptin could act directly and/or through alpha-melanocyte-stimulating hormone (alpha-MSH). This hypothesis was reinforced after in vitro studies, using the mouse hypothalamic GT1-7 cell line, since both leptin and Norleucine(4), D-Phenylalanine(7)-alpha-MSH (NDP-alpha-MSH) treatments increased MC4-R expression. After 8 weeks of high-fat diet, nondiabetic obese mice became resistant to the central action of leptin and their hypothalamic content of POMC and AgRP mRNA were decreased without modification of MC4-R mRNA level. After 16 weeks of high-fat diet, mice exhibited more severe metabolic disorders with type 2 diabetes. Moreover, hypothalamic expression of MC4-R was highly increased. In conclusion, several alterations of the melanocortin system were found in obese mice that are probably consecutive to their central resistance to leptin. Moreover, when the metabolic status is highly degraded (with all characteristics of a type 2 diabetes), other regulatory mechanisms (independent of leptin) can also take place.


Subject(s)
Hypothalamus/drug effects , Hypothalamus/metabolism , Leptin/physiology , Melanocortins/metabolism , Obesity/metabolism , Agouti-Related Protein/metabolism , Animals , Cell Line , Diabetes Mellitus, Type 2/metabolism , Disease Models, Animal , Hypothalamus/cytology , Infusions, Parenteral , Leptin/administration & dosage , Male , Mice , Mice, Inbred C57BL , Norleucine/pharmacology , Pro-Opiomelanocortin/metabolism , RNA, Messenger/metabolism , Receptor, Melanocortin, Type 4/metabolism , alpha-MSH/analogs & derivatives , alpha-MSH/pharmacology
6.
Peptides ; 26(10): 1842-7, 2005 Oct.
Article in English | MEDLINE | ID: mdl-15982783

ABSTRACT

The human melanocortin-2 receptor (hMC2R) is mainly present in the adrenal cortex and has been difficult to express in heterologous cells. The hMC2R fused to the EGFP at its C-terminus has been stably transfected in the murine M3 melanoma and HEK293 cells. In the M3 cells, the hMC2R-EGFP was well-addressed to the cell membrane and functional whereas in the HEK293 cells, the hMC2R-EGFP was retained intracellularly. These results suggest that some specific factors, missing in cells, which do not express any melanocortin receptor, are involved in the correct addressing of the hMC2R to the cell membrane.


Subject(s)
Receptor, Melanocortin, Type 2/biosynthesis , Receptor, Melanocortin, Type 2/genetics , Animals , Cell Line, Tumor , Cell Membrane/genetics , Cell Membrane/metabolism , Cells, Cultured , Humans , Mice , Microscopy, Confocal , Microscopy, Fluorescence , RNA, Messenger/metabolism , Receptor, Melanocortin, Type 2/metabolism , Receptor, Melanocortin, Type 3/biosynthesis , Receptor, Melanocortin, Type 3/genetics , Receptor, Melanocortin, Type 3/metabolism , Receptor, Melanocortin, Type 4/biosynthesis , Receptor, Melanocortin, Type 4/genetics , Receptor, Melanocortin, Type 4/metabolism , Recombinant Fusion Proteins/biosynthesis , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolism , Transfection
7.
J Biochem ; 135(4): 541-6, 2004 Apr.
Article in English | MEDLINE | ID: mdl-15115780

ABSTRACT

The melanocortin receptor type 4 (MC4-R) is involved in food intake and represents a potential target for the treatment of some forms of obesity. The fluorescent protein EGFP was fused to the wild-type or mutated coding sequence of the human MC4-R. After transfection in HEK 293, clones stably expressing hMC4-R-EGFP were selected. Wild-type chimeric hMC4-R was well addressed to the cell membrane as demonstrated using confocal microscopy and displayed the same pharmacological characteristics as native hMC4R. NDP-alpha MSH induced a time-dependent internalization of MC4-R that was partially prevented by AgRP. The two mutated chimeric receptors studied here (CTCT-deleted and C271A) showed a high alteration of their response to ligand and were retained inside the cells. In conclusion, we have developed a model of clones stably expressing EGFP-tagged-hMC4-R. This is the only such model available to date and it provides a useful tool to follow the trafficking of MC4-R inside living cells.


Subject(s)
Green Fluorescent Proteins/genetics , Receptor, Melanocortin, Type 4/genetics , alpha-MSH/analogs & derivatives , Agouti-Related Protein , Binding, Competitive , Cell Line , Cell Membrane/metabolism , Cyclic AMP/metabolism , Cytoplasmic Vesicles/metabolism , Dose-Response Relationship, Drug , Genetic Vectors/genetics , Green Fluorescent Proteins/metabolism , Humans , Microscopy, Fluorescence , Mutation , Peptide Fragments/pharmacology , Polymerase Chain Reaction , Protein Binding , Protein Transport/drug effects , Protein Transport/genetics , Protein Transport/physiology , Receptor, Melanocortin, Type 4/metabolism , Receptors, Melanocortin/agonists , Receptors, Melanocortin/antagonists & inhibitors , Receptors, Melanocortin/metabolism , Recombinant Fusion Proteins/agonists , Recombinant Fusion Proteins/antagonists & inhibitors , Recombinant Fusion Proteins/metabolism , Transfection , alpha-MSH/metabolism , alpha-MSH/pharmacology
8.
Endocrinology ; 145(2): 541-7, 2004 Feb.
Article in English | MEDLINE | ID: mdl-14592963

ABSTRACT

Agouti-related protein (Agrp), primarily expressed in the hypothalamus, is an endogenous antagonist of alphaMSH at the level of melanocortin 3 receptor (MC3-R) and MC4-R, but the adrenal gland represents the second major Agrp-expressing tissue. In adrenal fasciculata cells, the glucocorticoid secretion is under the control of ACTH, which binds specifically MC2-R, the only functional melanocortin receptor described in these cells to date. Nevertheless, using cultured bovine fasciculata adrenal cells, we report that Agrp has no antagonistic properties against ACTH at the level of MC2-R. In our studies, (Nle4, d-Phe7)-alphaMSH (NDP-alphaMSH) stimulated the production of cortisol in a dose-dependent manner, and these effects were abolished by Agrp or SHU9119, a synthetic antagonist of MC3-R and MC4-R. Using a more specific antagonist (JKC-363) and RT-PCR analysis, we can postulate that the effects of NDP-alphaMSH were mediated via MC4-R. These results are suggestive that adrenal glucocorticoid production could be regulated through MC4-R that may have some relevance in the physiology of adrenal cells. Moreover, Agrp might exert an autocrine control on adrenal cells because a protein with biological Agrp-like activity is secreted by these cells. This peptide could then modulate locally the functions of some peripheral tissues such as adrenals.


Subject(s)
Adrenal Glands/physiology , Glucocorticoids/biosynthesis , Proteins/pharmacology , Receptor, Melanocortin, Type 4/antagonists & inhibitors , alpha-MSH/analogs & derivatives , alpha-MSH/pharmacology , Adrenal Cortex/chemistry , Adrenal Cortex/physiology , Adrenocorticotropic Hormone/pharmacology , Agouti-Related Protein , Animals , Autocrine Communication , Cattle , Cell Line , Cells, Cultured , Culture Media, Conditioned , Gene Expression , Green Fluorescent Proteins , Humans , Hydrocortisone/biosynthesis , Hydrocortisone/metabolism , Intercellular Signaling Peptides and Proteins , Luminescent Proteins/genetics , Melanocyte-Stimulating Hormones/pharmacology , Mice , Proteins/metabolism , RNA, Messenger/analysis , Receptor, Melanocortin, Type 4/genetics , Receptor, Melanocortin, Type 4/physiology , Recombinant Fusion Proteins , Reverse Transcriptase Polymerase Chain Reaction , Zona Fasciculata/physiology
SELECTION OF CITATIONS
SEARCH DETAIL
...