Biomedical subjects

Michel Samson

Publications and source records attributed to Michel Samson.

17 recordsLinked to original sources

PubMed2006

Dynamic distribution of Spatial during mouse spermatogenesis and its interaction with the kinesin KIF17b.

The Spatial gene is expressed in highly polarized cell types, such as epithelial cells in the thymus, neurons in the brain and germ cells in the testis. In this study, we report the characterization and distribution of Spatial proteins during mouse spermatogenesis. Besides Spatial-epsilon and -delta, we show that the newly described short isoform Spatial-beta is expressed specifically in round spermatids. Using indirect immunofluorescence, we detected Spatial in the cytosol of the early round spermatid. By the end stages of round spermatids, Spatial is concentrated at the opposite face of the acrosome near the nascent flagellum and in the manchette during the elongation process. Finally in mature sperm, Spatial persists in the principal piece of the tail. Moreover, we found that Spatial colocalizes with KIF17b, a testis-specific isoform of the brain kinesin-2 motor KIF17. This colocalization is restricted to the manchette and the principal piece of the sperm tail. Further, coimmunoprecipitation experiments of native proteins from testis lysates confirmed Spatial-KIF17b association through the long Spatial-epsilon isoform. Together, these findings imply a function of Spatial in spermatid differentiation as a new cargo of kinesin KIF17b, in a microtubule-dependent mechanism specific to the manchette and the principal piece of the sperm tail.

Animals↗

PubMed2006

Protein profiling of rat ventral prostate following chronic finasteride administration: identification and localization of a novel putative androgen-regulated protein.

To better understand the effects of antiandrogens on the prostate, we investigated the changes in the proteome of rat ventral prostate (VP) following treatment with a well characterized 5alpha-reductase inhibitor, finasteride. Sprague-Dawley rats were treated daily by gavage with finasteride at 0, 1, 5, 25, and 125 mg/kg/day. Changes in plasma hormone levels as well as the weight and histology of sex accessory tissues were determined after 28 days of treatment and showed a dose-related decrease of VP weights together with a marked atrophy of the tissue visible at the macroscopic and microscopic levels. In addition, significant reductions in seminal vesicle and epididymis weights were noted. VP proteins were analyzed by two-dimensional gel electrophoresis: 37 proteins, mainly involved in protein synthesis, processing, and cellular trafficking and in metabolism, detoxification, and oxidative stress, were identified as modulated by finasteride. The prominent feature of this study is the demonstration of finasteride dose-dependent up-regulation of a protein similar to l-amino-acid oxidase 1 (Lao1). An up-regulation of this protein was also observed with the antiandrogen flutamide. Lao1 expression occurred as early as 48 h after antiandrogen administration and persisted throughout the treatment duration. Immunohistochemistry showed that this protein was only detectable in epithelial cells and secretory vesicles. Altogether these data point to a potential use of Lao1 to reveal antiandrogen-induced prostate injury.

5-alpha Reductase Inhibitors↗

PubMed2006

Robotic harvest of internal mammary vessels in breast reconstruction.

The authors have robotically harvested the internal mammary vessels to provide the recipient pedicle for free-flap breast reconstruction in 22 consecutive cases. Through minimal-access incisions, the robot facilitated the harvest of a pedicle up to 10 cm in length. The pedicle was brought out through the second intercostal space into the mastectomy wound, allowing the anastomosis to be performed on the skin surface with loupes or microscope. This paper illustrates their experience and offers an innovative approach to free-flap breast reconstruction.

Abdominal Muscles↗

PubMed2006

RANKL treatment releases the negative regulation of the poly(ADP-ribose) polymerase-1 on Tcirg1 gene expression during osteoclastogenesis.

UNLABELLED: The Tcirg1 gene encodes the osteoclast-specific a3 isoform of the V-ATPase a subunit. Using the mouse osteoclastic model RAW264.7 cells, we studied Tcirg1 gene expression, and we identified PARP-1 as a transcriptional repressor negatively regulated by RANKL during osteoclastogenesis. INTRODUCTION: The TCIRG1 gene encodes the a3 isoform of the V-ATPase a subunit, and mutations at this locus account for approximately 60% of infantile malignant osteopetrosis cases. Using RAW264.7 cells as an osteoclastic differentiation model, we undertook a transcriptional study of the mouse Tcirg1 gene focused on the 4-kb region upstream of the transcription starting point. MATERIALS AND METHODS: The promoter activity of serial-deletion fragments of the Tcirg1 gene promoter was monitored throughout the RAW264.7 cell differentiation process. We next performed EMSA, UV cross-linking, affinity purification, mass spectrometry analysis, gel supershift, and siRNA transfection experiments to identify the factor(s) interacting with the promoter. RESULTS: The -3946/+113 region of the mouse Tcirg1 gene displayed a high basal promoter activity, which was enhanced by RANKL treatment of RAW264.7 cells. Constructs deleted up to -1589 retained this response to RANKL. A deletion up to -1402 induced a 3-fold enhancement of the basal activity, whereas RANKL response was not affected. EMSA experiments led us to identify within the -1589/-1402 region, a 10-nucleotide sequence, which bound a nuclear protein present in nondifferentiated RAW264.7 cells. This interaction was lost using nuclear extracts derived from RANKL-treated cells. Affinity purification followed by mass spectrometry analysis and gel supershift assay allowed the identification of poly(ADP-ribose) polymerase-1 (PARP-1) as this transcriptional repressor, whereas Western blot experiments revealed the cleavage of the DNA-binding domain of PARP-1 on RANKL treatment. Finally, both PARP-1 depletion after siRNA transfection and RAW264.7 cell treatment by an inhibitor of PARP-1 activity induced an increase of a3 mRNA expression. CONCLUSIONS: We provide evidence that the basal transcription activity of the Tcirg1 gene is negatively regulated by the binding of PARP-1 protein to its promoter region in mouse pre-osteoclast. On RANKL treatment, PARP-1 protein is cleaved and loses its repression effect, allowing an increase of Tcirg1 gene expression that is critical for osteoclast function.

Amino Acid Sequence↗

PubMed2005

Molecular cloning of several rat ABC transporters including a new ABC transporter, Abcb8, and their expression in rat testis.

Several members of the ABC transporter superfamily play an important role in testicular physiology and defence against anticancer drugs. Using a reverse transcription-polymerase chain reaction strategy with degenerate primers and rat testis RNA as template, we have looked for the presence of other members of this superfamily. Of the six partial cDNA found, five corresponded to ABC transporters already known -Mdr1b, Mrp1, Tapl/Abcb9, Umat/Abcb6 and Sur2/Abcc9- and one presented a strong homology with mouse and human ABCB8. Using a 5' and 3' RACE approach, we cloned the full-length cDNA and found that the predicted protein presented 92% and 80% homology with the mouse and human proteins respectively. Strong expression of rat abcb8 was found in heart, brain and testis when compared with liver, lung and spleen. In the testis, rat abcb8 was expressed both in the somatic Sertoli cells and peritubular cells and in the germline (spermatogonia and pachytene spermatocytes). Furthermore, Umat/Abcb6 was very highly expressed in the testis (high amounts in meiotic pachytene spermatocytes and low amount in post-meiotic early spermatids). In conclusion, we confirm the presence of several ABC transporters in the testis and also provide evidence of the presence of Abcb8 in the organ.

ATP-Binding Cassette Transporters↗

PubMed2004

Dominant negative effect of connexin33 on gap junctional communication is mediated by connexin43 sequestration.

Gap junctional intercellular communication is involved in the control of cell proliferation and differentiation. Connexin33, a member of the multi-gene family of gap junction proteins, exerts an inhibitory effect on intercellular communication when injected into Xenopus oocytes. However, the molecular mechanisms involved remain to be elucidated. Our results show that connexin33 was only expressed within the seminiferous tubules in the testis. In contrast to the majority of connexins, connexin33 was unphosphorylated. Immunoprecipitation experiments revealed that connexin33 physically interacted with connexin43, mainly with the phosphorylated P1 isoform of connexin43 but not with connexin26 and connexin32, two other connexins expressed in the tubular compartment. In Sertoli cells and COS-7 cells, connexin43 was located at the plasma membrane, whereas in connexin33 transfected cells, the specific association of connexin33/43 was sequestered in the intracellular compartment. High-resolution fluorescent deconvolution microscopy indicated that the connexin33/43 complex was mainly found within early endosomes. Sequestration of connexin33/43 complex was associated with a complete inhibition of the gap junctional coupling between adjacent cells. These findings provide the first evidence of a new mechanistic model by which a native connexin, exerting a dominant negative effect, can inhibit gap junctional intercellular communication. In the testis, connexin33 could exert a specific role on germ cell proliferation by suppressing the regulatory effect of connexin43.

Animals↗

PubMed2004

Human endocrine gland-derived vascular endothelial growth factor: expression early in development and in Leydig cell tumors suggests roles in normal and pathological testis angiogenesis.

Angiogenesis is essential for tumor growth and metastasis. A new human angiogenic mitogen, endocrine gland-derived vascular endothelial growth factor (EG-VEGF), has been recently identified; its expression pattern is restricted to endocrine glands, with the highest expression in testis. We used in situ hybridization and newly generated monoclonal antibodies to investigate the expression of EG-VEGF in normal human prenatal and adult testis and in 48 human testicular tumors of different subtypes. We found that EG-VEGF was expressed from 14 wk until birth in human fetal testis. In the adult testis, EG-VEGF was strongly expressed only in Leydig cells. In testicular tumors, EG-VEGF was expressed specifically in Leydig cell tumors, whereas germ cell-derived neoplasms, including carcinoma in situ, seminoma, and nonseminomatous germ cell tumors, were negative for this antigen. In contrast, VEGF, another powerful angiogenic factor, was expressed in seminoma, but very weakly in Leydig cell tumors. Interestingly, we found that Leydig cell tumors presented vessel surface density 3.2-fold higher than seminoma. These findings argue that human EG-VEGF may play a role in angiogenesis both during the early endocrine development of testis and in the adult testis as well as in Leydig cell tumor growth.

Adult↗

PubMed2003

Aberrant Connexin 43 endocytosis by the carcinogen lindane involves activation of the ERK/mitogen-activated protein kinase pathway.

Although worldwide concerns have emerged about environmental factors that display carcinogenic and reprotoxic effects, little is known about the mechanism(s) by which these chemicals alter testicular function. Using the 42GPA9 Sertoli cell line, we recently reported that one widely used lipid-soluble pesticide, Lindane impairs gap junctional intercellular communication by promoting the intracellular localization of Connexin 43 (Cx43), a tumor suppressor. We showed here that this chemical triggered the accumulation of Cx43 within Rab5 positive endosomes. Interestingly, evidence is provided that Lindane-induced Cx43 endocytosis did not stem on alteration of Cx43 partition in lipid rafts. Lindane induced concomitantly Cx43 phosphorylation and activation of extracellular signal-regulated kinases (ERK) but not of JNK and p38 mitogen- activated protein kinases. Inhibition of ERK pathway by PD98059, a MEK1-specific inhibitor, prevented Lindane-induced Cx43 phosphorylation, restored Cx43 membranous localization and gap junction coupling. Altogether, these findings provide the first evidence that Lindane-altered Cx43 endocytosis requires ERK activation. Such inappropriate activation of the mitogenic MAPK pathway and inactivation of the tumor suppressor Cx43 by Lindane may participate in the promotion of neoplastic cell growth.

Animals↗

PubMed2003

CD47 and the 19 kDa interacting protein-3 (BNIP3) in T cell apoptosis.

CD47 is a surface receptor that induces either coactivation or apoptosis in lymphocytes, depending on the ligand(s) bound. Interestingly, the apoptotic pathway is independent of caspase activation and cytochrome c release and is accompanied by early mitochondrial dysfunction with suppression of mitochondrial membrane potential (Deltapsim). Using CD47 as bait in a yeast two-hybrid system, we identified the Bcl-2 homology 3 (BH3)-only protein 19 kDa interacting protein-3 (BNIP3), a pro-apoptotic member of the Bcl-2 family, as a novel partner. Interaction between CD47 and the BH3-only protein was confirmed by immunoprecipitation analysis, and CD47-induced apoptosis was inhibited by attenuating BNIP3 expression with antisense oligonucleotides. Finally, we showed that the C-terminal domain of thrombospondin-1 (TSP-1), but not signal-regulatory protein (SIRPalpha1), is the ligand for CD47 involved in inducing cell death. Immunofluorescence analysis of CD47 and BNIP3 revealed a partial colocalization of both molecules under basal conditions. After T cell stimulation via CD47, BNIP3 translocates to the mitochondria to induce apoptosis. These results show that the BH3-dependent apoptotic pathways, previously shown to be activated by intracellular pro-apoptotic events, can also be turned on by surface receptors. This new pathway results in a fast induction of cell death resembling necrosis, which is likely to play an important role in lymphocyte regulation at inflammatory sites and/or in the vicinity of thrombosis.

Antigens, CD↗

PubMed2003

Spatial, a new nuclear factor tightly regulated during mouse spermatogenesis.

In order to clarify general mechanisms of T cell development, we used array technology to identify genes differentially expressed between wild type and mutant mice thymuses. This quantitative differential screening leads to the isolation of Spatial, a putative transcription factor, expressed at high level in thymic cortical stromal cells. We report here, by RT-PCR on 48 distinct tissues that Spatial is also highly expressed in testis. Interestingly, the testicular expression is developmentally regulated, since it only appears in adult mice around 7-8 weeks of age. Analysis of purified testicular cell types show that Spatial expression is restricted to haploid round spermatids during spermiogenesis and the expression sites were further localized by in situ hybridization to step 2-10 spermatids. Thus, this recently described nuclear factor constitutes a new marker of the round spermatid stage that may play a key role in the control of male germ cell development.

Animals↗

PubMed2003

The emerging phenotype of the testicular carcinoma in situ germ cell.

This review summarises the existing knowledge on the phenotype of the carcinoma in situ (CIS) cell. CIS is a common pre-invasive precursor of testicular germ cell tumours of adolescents and young adults. These tumours display a variety of histological forms. Classical seminoma proliferates along the germ cell lineage, whereas embryonal carcinoma retains embryonic features and readily differentiates into teratomas that resemble various somatic cell lineages. A thorough review of the gene expression in CIS cells in comparison to normal testicular germ cells and overt tumours supports the view that CIS is a common precursor for both tumour types. Impaired cell differentiation resulting in a partial retention of the embryonic features, associated with an increasing genomic instability may be responsible for a remarkable phenotypic heterogeneity of CIS cells. Depending on the degree of differentiation and pluripotency, CIS cells found in adult patients seem to be predestined for further malignant progression into one or the other of the two main types of overt tumours. A new concept of phenotypic continuity of differentiation of germ cells along germinal lineage with a gradual loss of embryonic features based on the analysis of gene expression in all types of germ cells during their ontogeny is presented in this review. The data point out that despite the phenotypic continuum of gene expression, there are two periods of rapid changes of gene expression: first at the transition from primordial germ cells to pre-spermatogonia, and later during the pubertal switch from the mitotic to meiotic cell division. The persistent expression of embryonic traits in CIS cells, and the high expression of the cell cycle regulators that are typical of mitotic germ cells support our long-standing hypothesis that CIS cells originate from primordial germ cells or gonocytes and not from germ cells in the adult testis.

Adolescent↗

PubMed2003

Expression of fractalkine in the rat testis: molecular cloning of a novel alternative transcript of its gene that is differentially regulated by pro-inflammatory cytokines.

The testis is a very complex organ in which cellular communications and interactions are central to spermatogenesis and where inflammatory processes can lead to sterility. Fractalkine (CX3CL1) is a chemokine involved in cell-cell interactions and in leukocyte chemoattraction. It has been reported to be expressed in testis, but its cellular expression and function in this organ has not been described. In this study we report constitutive expression of fractalkine in the testis. Expression is higher in Leydig cells than Sertoli cells, spermatogonia, pachytene spermatocytes and elongated spermatids. In both, Sertoli cells stimulated by interleukin-1beta and tumour necrosis factor alpha, and in Leydig cells, two forms of fractalkine mRNA were observed: the previously described transcript of 3.7 kb and a novel transcript of 4.2 kb. The 4.2 kb transcript has a 5' elongation and is differentially regulated. To investigate fractalkine function in testis, we abolished Leydig cell expression of fractalkine by specific destruction of this cell type using ethylene dimethane sulphonate. The absence of fractalkine expression in Leydig cells did not seem to affect the fractalkine expression by other testicular cells. In addition, the destruction of testicular macrophages by Cl2MDP (chlodronate) did not seem to affect Leydig cell expression of fractalkine. We conclude that Leydig cell expression of fractalkine could be preferentially involved in inflammation in interstitial space whereas fractalkine expressed by germ cells may participate in the cellular interactions between germ cells and other seminiferous tubule cell types.

Alternative Splicing↗

PubMed2003

Mumps virus decreases testosterone production and gamma interferon-induced protein 10 secretion by human leydig cells.

Mumps virus is responsible for sterility. Here, we show that the mumps virus infects Leydig cells in vitro and totally inhibits testosterone secretion and that ribavirin in mumps virus-infected Leydig cell cultures completely restores testosterone production. Moreover, we show that gamma interferon-induced protein 10 (IP-10) is highly expressed by mumps virus-infected Leydig cells and that ribavirin does not block IP-10 production.

Animals↗

PubMed2002

Study of the HIV-1 receptors CD4, CXCR4, CCR5 and CCR3 in the human and rat testis.

Sexual transmission of HIV is one of the main routes of transmission of AIDS. Despite the fact that the virus has been found in the semen and germ cells of patients with HIV, little is known about how the virus infects the cells of the genital tract. We studied the cellular distribution of CD4, a receptor necessary for HIV infection, and the major HIV co-receptors CCR3, CCR5 and CXCR4 in the rat and human testis. We used RT-PCR, Northern blotting and immunohistochemistry to demonstrate that CCR3 is absent from the testes of both species, whereas CCR5 and CXCR4 are present on the resident testicular macrophages in the interstitial space but not in the germ cell line. All of the human testicular macrophages expressed the markers CD45 and MAC387 and most also expressed CD4. Thus, our data suggest that macrophages in the testis may be infected by HIV and that these macrophages may be a site of early viral localization and a potential HIV reservoir. This may in turn alter the activity of Leydig cells and subsequently affect spermatogenesis.

Animals↗

PubMed2002

The cancer-testis gene, NY-ESO-1, is expressed in normal fetal and adult testes and in spermatocytic seminomas and testicular carcinoma in situ.

Cancer/testis genes are potential targets for therapeutic genetic and immunologic approaches, and are highly expressed in a large variety of human cancers. However, they are not expressed in normal tissues, with the exception of the testis. The NY-ESO-1 gene is the most recently identified member of the cancer/testis family and its product is one of the most immunogenic tumor antigens. We used immunohistochemistry to investigate the expression of NY-ESO-1 in healthy human prenatal and adult testes and in 59 human testicular tumors of different subtypes. We found that NY-ESO-1 was expressed from 18 weeks until birth in human fetal testes. In the adult testis, NY-ESO-1 was strongly expressed in spermatogonia and in primary spermatocytes, but not in post-meiotic cells or in testicular somatic cells. NY-ESO-1 was not expressed in the Sertoli cells, Leydig cells, classical seminomas, or nonseminomatous germ cells in the 59 testicular tumors. In contrast, NY-ESO-1 was expressed both in carcinomas in situ, which are the earliest stage of testicular tumors (7 of 15 cases), and in spermatocytic seminomas, which are believed to be derived from spermatogonia or primary spermatocytes (8 of 16 cases). We conclude that NY-ESO-1 is a marker that can be used to follow the early progression of testicular tumorigenesis when the tumors present a similar pattern of expression to the cells from which they originated, although the later tumors cease to express NY-ESO-1.

Antigens, Neoplasm↗

PubMed2002

Production of the chemokines monocyte chemotactic protein-1, regulated on activation normal T cell expressed and secreted protein, growth-related oncogene, and interferon-gamma-inducible protein-10 is induced by the Sendai virus in human and rat testicular cells.

Several viruses infect the testis, inducing inflammation, which may lead to infertility. In this study we investigated the production in rat and human testicular cells exposed to the Sendai virus of several chemokines that play a major role in inflammatory processes. Exposure of rat testicular macrophages and Sertoli, Leydig, and peritubular cells to the Sendai virus led to the production of mRNA and protein for monocyte chemotactic protein-1 (MCP-1), regulated on activation normal T cell expressed and secreted protein, growth-related oncogene-alpha, and interferon-gamma-inducible protein-10. In rat peritubular cells exposed to the Sendai virus, MCP-1 production was time and dose dependent. In contrast, rat germ cells did not produce these chemokines. Chemokine synthesis was detected in human Leydig cells exposed to the Sendai virus, but not in human total germ cells, suggesting that rats and humans display similar responses in terms of chemokine production. MCP-1, regulated on activation normal T cell expressed and secreted protein, growth-related oncogene-alpha, and interferon-gamma-inducible protein-10 have been reported to be chemoattractants for a large variety of leukocytes. The ability of the Sendai virus to induce chemokine production in somatic cells (mostly peritubular and Leydig cells) may therefore increase the recruitment of leukocytes to sites of infection.

Animals↗

PubMed2002

[Mumps virus and orchitis: towards a physiopathologic approach].

Mumps orchitis is a dreaded complication of mumps is pubescent men. The literature on this subject includes epidemiological, clinical, histological and endocrine findings, indicating a marked variability of the clinical features from one patient to another, an alteration of endocrine function that can persist in the long term and finally post-mumps infertility, which is exceptional. On the other hand, relatively few studies have investigated the pathophysiological mechanisms, and suggest replication of the mumps virus in the testis. Based on these data from the literature, this article reviews or proposes various hypotheses concerning the various pathophysiological aspects of this disease and discusses the lines of research that could advance the current knowledge in order to improve the therapeutic management of patients, while also providing a better knowledge of the mumps virus and basic testicular physiology.

Humans↗