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Biomedical subjects

M M Matzuk

Publications and source records attributed to M M Matzuk.

At least 19 recordsLinked to original sources
PubMed2006

Spermatogenesis does not require the local production of follistatin.

It has been proposed that follistatin can modulate the actions of activins and/or other members of the transforming growth factor-beta superfamily of proteins on testicular function, since mice overexpressing follistatin showed spermatogenic disruption. However, since mice with targeted disruption of the follistatin gene die soon after birth, it is not feasible to determine the effect of the absence of follistatin on testicular function using this model. To further understand the role of follistatin on the development and maintenance of spermatogenesis, fetal testes, collected by Caesarean section at day 18 of gestation from follistatin null mice, were transplanted to the external ear of castrated recombination activating gene 1 immunocompromised male mice. The testicular grafts were then analysed 7-8 weeks after transplantation and showed that full spermatogenesis developed in both the testes of wild-type and follistatin null mice. This study indicates that, if follistatin is required to modulate spermatogenic development, it is not supplied by local testicular production but by circulating follistatin from the host mouse.

Animals↗
PubMed2004

Human cumulus granulosa cell gene expression: a predictor of fertilization and embryo selection in women undergoing IVF.

BACKGROUND: A biochemical marker for embryo development would increase the chance of a successful pregnancy with IVF by optimizing oocyte and embryo selection, and allow fewer embryos to be transferred. In this study, we correlated cumulus granulosa cell gene expression of hyaluronic acid synthase 2 (HAS2), cyclooxygenase 2 (COX2; PTGS2) and gremlin (GREM1) with subsequent embryo development in search of a parameter for embryo selection. METHODS: Cumulus cell gene expression was determined prospectively on eight consecutive patients undergoing IVF with ICSI. Immediately following oocyte retrieval, the cumulus was stripped from the oocyte, and cumulus gene expression for PTGS2, HAS2 and GREM1 was assessed using a one-step real-time quantitative RT-PCR assay. Oocyte quality, fertilization and embryo morphology were correlated to relative gene expression. RESULTS: Gene expression data were available on cumulus cells from 108 oocytes that developed into 70 embryos (64.8% fertilization rate). Cumulus PTGS2, HAS2 and GREM1 expression was higher from oocytes that developed into higher quality embryos (grades 3, 4 and 5) compared with lower quality embryos (grades 1 and 2) (P<0.05, P<0.001 and P<0.001, respectively). HAS2 and GREM1 expression was also higher from the cumulus surrounding oocytes that gave rise to higher grade embryos (P<0.001). The expression of PTGS2 and HAS2 was 6-fold higher, and that of GREM1 was 15-fold higher in cumulus yielding higher grade embryos versus lower grade embryos. CONCLUSION: PTGS2, HAS2 and GREM1 gene expression correlates to morphological and physiological characteristics and provides a novel approach to predict human embryo development. Ultimately, with better predictors of follicular and embryonic health, higher quality embryos can be selected and transferred, reducing higher order pregnancy rates.

Adult↗
PubMed2001

Studying TGF-beta superfamily signaling by knockouts and knockins.

The transforming growth factor beta (TGF-beta) superfamily has profound effects on many aspects of animal development. In the last decade, our laboratory and others have performed in vivo functional studies on multiple components of the TGF-beta superfamily signal transduction pathway, including upstream ligands, transmembrane receptors, receptor-associated proteins and downstream Smad proteins. We have taken gene knockout approaches to generate null alleles of the genes of interest, as well as a gene knockin approach to replace the mature region of one TGF-beta superfamily ligand with another. We found that activin betaB, expressed in the spatiotemporal pattern of activin betaA, can function as a hypomorphic allele of activin betaA and rescue the craniofacial defects and neonatal lethal phenotype of activin betaA-deficient mice. With the knockout approach, we have shown that the expression pattern of a component in the TGF-beta superfamily signal transduction cascade does not necessarily predict its in vivo function. Two liver-specific activins, activin betaC and activin betaE are dispensable for liver development, regeneration and function, whereas ubiquitously expressed Smad5 has specific roles in the development of multiple embryonic and extraembryonic tissues.

Activins↗
PubMed2001

Continuous fatty acid oxidation and reduced fat storage in mice lacking acetyl-CoA carboxylase 2.

Malonyl-coenzyme A (malonyl-CoA), generated by acetyl-CoA carboxylases ACC1 and ACC2, is a key metabolite in the regulation of energy homeostasis. Here, we show that Acc2-/- mutant mice have a normal life span, a higher fatty acid oxidation rate, and lower amounts of fat. In comparison to the wild type, Acc2-deficient mice had 10- and 30-fold lower levels of malonyl-CoA in heart and muscle, respectively. The fatty acid oxidation rate in the soleus muscle of the Acc2-/- mice was 30% higher than that of wild-type mice and was not affected by addition of insulin; however, addition of insulin to the wild-type muscle reduced fatty acid oxidation by 45%. The mutant mice accumulated 50% less fat in their adipose tissue than did wild-type mice. These results raise the possibility that pharmacological manipulation of ACC2 may lead to loss of body fat in the context of normal caloric intake.

3-Hydroxybutyric Acid↗
PubMed2001

Oosp1 encodes a novel mouse oocyte-secreted protein.

Oocyte-somatic cell communication is necessary for normal ovarian function. However, the identities of the majority of oocyte-secreted proteins remain unknown. A novel cDNA encoding mouse oocyte-secreted protein 1 (OOSP1) was identified using a modified subtractive hybridization screen. The Oosp1 cDNA encodes a 202-amino acid protein that contains a 21-amino acid signal peptide sequence, 5 putative N-linked glycosylation consensus sequences, and 6 cysteines that are predicted to form 3 disulfide bonds. OOSP1 shares amino acid identity with placental-specific protein 1 (PLAC1), a secreted protein expressed in the placenta and the ectoplacental cone. The Oosp1 mRNA is approximately 1.0 kb and is present at high levels in the oocytes of adult ovaries and at lower levels in the spleen. The mouse Oosp1 gene is 5 exons, spans greater than 16.4 kb, and localizes to chromosome 19 at a position that shares synteny with human chromosome 11q12-11q13. The identification of OOSP1 as a new oocyte-secreted protein permits future in vitro and in vivo functional analyses to define its role in ovarian folliculogenesis.

Amino Acid Sequence↗
PubMed2001

Smad5 is required for mouse primordial germ cell development.

Smad5, together with Smad1 and Smad8, have been implicated as downstream signal mediators for several bone morphogenetic proteins (BMPs). Recent studies have shown that primordial germ cells (PGCs) are absent or greatly reduced in Bmp4 or Bmp8b mutant mice. To define the role of Smad5 in PGC development, we examined PGC number in Smad5 mutant mice by Oct4 whole-mount in situ hybridization and alkaline phosphatase staining. We found ectopic PGC-like cells in the amnion of some Smad5 mutant mice, however, the total number of PGCs was greatly reduced or completely absent in Smad5 mutant embryos, similar to Bmp4 or Bmp8b mutant embryos. Therefore, Smad5 is an important factor involved in PGC generation and localization.

Alkaline Phosphatase↗
PubMed2001

Eggs in the balance.

Polycyclic aromatic hydrocarbons (PAH), found in cigarette smoke and air pollution, interact with the aryl hydrocarbon receptor (Ahr) to cause reproductive defects. Mice lacking either Ahr or the pro-apoptotic protein Bax have an increased number of primordial follicles, and these mutant oocytes are resistant to PAH toxicity. A new study shows that the Bax promoter contains two core Ahr response elements, which are required for PAH stimulation of Bax promoter activity in oocytes. Thus, the toxic effects of PAH in oocytes are mediated directly by Ahr induction of the Bax pathway.

Animals↗
PubMed2001

Disruption of gamma-glutamyl leukotrienase results in disruption of leukotriene D(4) synthesis in vivo and attenuation of the acute inflammatory response.

To study the function of gamma-glutamyl leukotrienase (GGL), a newly identified member of the gamma-glutamyl transpeptidase (GGT) family, we generated null mutations in GGL (GGL(tm1)) and in both GGL and GGT (GGL(tm1)-GGT(tm1)) by a serial targeting strategy using embryonic stem cells. Mice homozygous for GGL(tm1) show no obvious phenotypic changes. Mice deficient in both GGT and GGL have a phenotype similar to the GGT-deficient mice, but approximately 70% of these mice die before 4 weeks of age, at least 2 months earlier than mice deficient only in GGT. These double-mutant mice are unable to cleave leukotriene C(4) (LTC(4)) to LTD(4), indicating that this conversion is completely dependent on the two enzymes, and in some organs (spleen and uterus) deletion of GGL alone abolished more than 90% of this activity. In an experimental model of peritonitis, GGL alone is responsible for the generation of peritoneal LTD(4). Further, during the development of peritonitis, GGL-deficient mice show an attenuation in neutrophil recruitment but not of plasma protein influx. These findings demonstrate an important role for GGL in the inflammatory response and suggest that LTC(4) and LTD(4) have distinctly different functions in the inflammatory process.

Animals↗
PubMed2001

Blockage of the rete testis and efferent ductules by ectopic Sertoli and Leydig cells causes infertility in Dax1-deficient male mice.

DAX-1, an X-linked member of the orphan nuclear receptor superfamily of transcription factors, plays a key role in sex determination and gonadal differentiation. Dax1-deficient male mice are infertile and have small testes despite normal serum levels of T and gonadotropins. Examination of Dax1-deficient testes reveals dilated seminiferous tubules and abnormal parameters of sperm fertilizing capability consistent with a possible obstruction in the testis. To test this hypothesis, we performed a comprehensive evaluation of the male reproductive tract in Dax1-deficient mice. Light and electron microscopic examination revealed the rete testis is blocked by aberrantly located Sertoli cells, creating a tailback of necrosing sperm in the testis. Sertoli cells also obstruct the proximal and middle efferent ductules, and this is accompanied by an overgrowth of the efferent duct epithelium. Seminiferous tubules close to the rete testis contain ectopic Leydig cells, distinct from the hyperplastic Leydig cells present in the interstitial space. The peritubular tissue surrounding these tubules is frequently abnormal, containing relatively undifferentiated myoid cells and no basement membrane between the myoid cells and Sertoli cells. A third of aged (>1-yr-old) Dax1-deficient male mice develop sex cord-stromal tumors, derived from cells of the Sertoli/granulosa cell or Leydig cell lineages. Combined, these observations reveal abnormal differentiation and proliferation of Leydig cells and Sertoli cells in Dax1-deficient male mice, leading to obstruction of the rete testis and infertility.

Animals↗
PubMed2001

Anti-Müllerian hormone attenuates the effects of FSH on follicle development in the mouse ovary.

Although ovarian follicle growth is under the influence of many growth factors and hormones of which FSH remains one of the most prominent regulators. Therefore, factors affecting the sensitivity of ovarian follicles to FSH are also important for follicle growth. The aim of the present study was to investigate whether anti-Müllerian hormone (AMH) has an inhibitory effect on follicle growth by decreasing the sensitivity of ovarian follicles to FSH. Furthermore, the combined action of AMH and FSH on ovarian follicle development was examined. Three different experiments were performed. Using an in vitro follicle culture system it was shown that FSH-stimulated preantral follicle growth is attenuated in the presence of AMH. This observation was confirmed by an in vivo experiment showing that in immature AMH-deficient females, more follicles start to grow under the influence of exogenous FSH than in their wild-type littermates. In a third experiment, examination of the follicle population of 4-month-old wild-type, FSH beta-, AMH-, and AMH-/FSH beta-deficient females revealed that loss of FSH expression has no impact on the number of primordial and preantral follicles, but the loss of inhibitory action of AMH on the recruitment of primordial follicles in AMH-deficient mice is increased in the absence of FSH. In conclusion, these studies show that AMH inhibits FSH-stimulated follicle growth in the mouse, suggesting that AMH is one of the factors determining the sensitivity of ovarian follicles for FSH and that AMH is a dominant regulator of early follicle growth.

Animals↗
PubMed2001

Analysis of the testicular phenotype of the follicle-stimulating hormone beta-subunit knockout and the activin type II receptor knockout mice by stereological analysis.

This study evaluated the role of FSH and activin A on testicular function using quantitative stereological analysis of testicular cell types in mice with targeted disruption of genes encoding the FSH beta-subunit and the activin type IIA receptor (ActRIIA). Using the optical dissector technique, the numbers of Sertoli cells and germ cells per testis were determined. Testis weights in homozygous males lacking the FSHbeta gene or the ActRIIA gene were decreased approximately 60% compared with wild-type or respective heterozygotes. Sertoli cell numbers decreased in both homozygous mice by 30-39%, and there was a comparable decline in germ cell numbers in both models. The degree of germ cell attrition increased in the later stages of spermatogenesis from a 46% reduction of spermatogonia to a 60% decrease in round spermatids. As the FSH levels are decreased in both models, the cellular lesion in both is most likely due to the FSH deficiency. Although the decrease in the Sertoli cell complement represents one cause of lower germ cell numbers, the ability of Sertoli cells to nurture germ cells is compromised by the lower FSH levels, as shown by a decrease in the round spermatid to Sertoli cell ratios in both homozygous models. We conclude that the defects in FSH beta-subunit gene knockout and ActRIIA knockout mice are related to diminished FSH action on both Sertoli cell proliferation and the capacity of Sertoli cells to nurture germ cells.

Activin Receptors, Type II↗
PubMed2001

Analysis of ovarian gene expression in follicle-stimulating hormone beta knockout mice.

FSH is a heterodimeric glycoprotein hormone that is produced in the gonadotroph cells of the anterior pituitary. It acts on Sertoli cells of the testis and granulosa cells of the ovary. We previously demonstrated that FSHbeta knockout female mice are infertile due to a block in folliculogenesis preceding antral stage development. To investigate aberrations of ovarian gene regulation in the absence of FSH, we analyzed the expression of several important marker genes using Northern blot and in situ hybridization techniques. Key findings are as follows: 1) Follicles of FSHbeta knockout mice develop a well organized thecal layer, which is positive for P450 17alpha-hydroxylase and LH receptor messenger RNAs (mRNAs). This indicates that theca recruitment is completed autonomously with respect to FSH. 2) Granulosa cells in FSH-deficient mice demonstrate an increase in FSH receptor mRNA, and decreases in P450 aromatase, serum/glucocorticoid-induced kinase, and inhibin/activin subunit mRNAs. These data support studies that implicate FSH signaling cascades in the expression of these genes. 3) In contrast to the thecal layer, granulosa cell populations in FSHbeta knockout mice do not accumulate LH receptor mRNA. This suggests that although the granulosa cells have a block in proliferation at the antral follicle stage in the absence of FSH, they do not initiate programs of terminal differentiation as seen in luteinizing cells of wild-type ovaries. 4) Ovaries of FSH-deficient mice demonstrate a modest decrease in cyclin D2 mRNA, without up-regulation of cell cycle inhibitor mRNAs associated with luteinization (i.e. p15, p27, and p21). Although components of the FSH null phenotype may be caused by partial cyclin D2 loss of function, these findings indicate that the mechanisms of granulosa cell cycle arrest in FSHbeta knockout mice are distinct from those of cycle withdrawal at luteinization. Underscoring the usefulness of the FSH-deficient mouse model, this study clarifies aspects of gonadotropin-dependent folliculogenesis, thecal layer development, cycle control in granulosa cells, and luteinization.

Animals↗
PubMed2001

Male reproductive phenotypes in double mutant mice lacking both FSHbeta and activin receptor IIA.

Activins are known to signal through two serine/threonine kinase type II receptors. Activin receptor IIA is widely expressed in the male reproductive axis, including the pituitary and testis. Our previous studies using gene knockout mice have confirmed the essential in vivo role of activin receptor IIA in FSH homeostasis. Activin receptor IIA-null male mice are fertile, have suppressed pituitary and serum FSH levels, and demonstrate a decrease in testis size as a result of reduced Sertoli cells and germ cells. Similarly, FSHbeta null male mice are fertile despite reduced testis size and Sertoli cell number. To define the direct roles of activin receptor IIA signaling locally in the testis, independent of its effects on FSH homeostasis, we generated double mutant mice lacking both activin receptor IIA and FSH by a genetic intercross and analyzed the male reproductive phenotypes. The double mutant male mice lacking both FSH and activin receptor IIA are fertile, demonstrate no significant reduction in testis size, and produce small litters compared with mice lacking either FSH or activin receptor IIA alone. Histological analyses of the testes from double mutant mice revealed the presence of normal stages of spermatogenesis. However, there was a significant reduction in the epididymal sperm number compared with that of the individual mutants. Northern blot analyses of total RNA from testes of double mutants did not reveal transcriptional up-regulation of activin receptor IIB, the other activin type II receptor. Although RNA expression profiles of many testis cell-specific markers are unaltered, stereological analysis of the testes from double mutants indicates that there was a reduction in type A and I spermatogonial number compared with that observed in individual mutants. Our results provide in vivo genetic evidence to demonstrate that activin receptor IIA signaling plays an important local role within the testis, independent of its actions via FSH homeostasis in the pituitary.

Activin Receptors, Type II↗
PubMed2001

Inhibin and p27 interact to regulate gonadal tumorigenesis.

Tumor suppressors function as antiproliferative signaling proteins, and defects in these genes lead to uncontrolled cell proliferation and cancer. For example, absence of the tumor suppressor p27(Kip1), a cyclin-dependent kinase inhibitor (CKI), results in increased body size, hyperplasia of several organs including the testes, and cancer in mice. Similarly, lack of inhibins, alpha/beta heterodimeric members of the transforming growth factor-beta (TGFbeta) superfamily, causes testicular and ovarian tumors of the granulosa/Sertoli cell lineage beginning at 4 weeks of age and adrenal tumors in gonadectomized mice. Neither the cell cycle alterations in the absence of inhibin nor the cause of the increased testis size in the p27 knockout mice is known. To study the molecular (cell cycle) changes that result from absence of inhibins, we analyzed the regulation of cell cycle proteins in gonadal tumors derived from inhibin alpha knockout mice (Inha(-/-)). Northern blot analyses demonstrate that cyclin-dependent kinase 4 (Cdk4) and cyclin D2 mRNA levels are elevated, and immunohistochemistry shows that p27 protein levels are decreased in both ovarian and testicular tumors from Inha(-/-) mice. These findings suggest that increased Cdk4/cyclin D2 (positive) activity and decreased p27 (negative) activity is causal for gonadal tumor formation. To test this hypothesis, we generated double mutant mice lacking both p27 and inhibin alpha to determine whether the tumor suppressors p27 and inhibin have additive suppressor activity in the gonads. Like Inha(-/-) mice, p27(-/-)Inha(-/-) mice demonstrate elevated serum activin levels, ovarian and testicular tumors, and a resultant lethal cachexia-like syndrome. However, whereas 95% of the Inha(-/-) female mice die by 18 weeks of age, 100% of the p27(-/-)Inha(-/-) female mice are dead by 8 weeks. Similarly, 95% of the Inha(-/-) single mutant males die by 13 weeks while 100% of the p27(-/-)Inha(-/-) male mice die by 10 weeks. Moreover, tumor foci in p27(-/-)Inha(-/-) mice can be observed as early as 2 weeks of age in males and as early as 4 weeks in females. These findings demonstrate that absence of both inhibin and p27 in mice causes earlier development of ovarian and testicular tumors and earlier death compared with absence of inhibin alone.

Activins↗
PubMed2001

Synergistic roles of bone morphogenetic protein 15 and growth differentiation factor 9 in ovarian function.

Knockout mouse technology has been used over the last decade to define the essential roles of ovarian-expressed genes and uncover genetic interactions. In particular, we have used this technology to study the function of multiple members of the transforming growth factor-beta superfamily including inhibins, activins, and growth differentiation factor 9 (GDF-9 or Gdf9). Knockout mice lacking GDF-9 are infertile due to a block in folliculogenesis at the primary follicle stage. In addition, recombinant GDF-9 regulates multiple cumulus granulosa cell functions in the periovulatory period including hyaluronic acid synthesis and cumulus expansion. We have also cloned an oocyte-specific homolog of GDF-9 from mice and humans, which is termed bone morphogenetic protein 15 (BMP-15 or Bmp15). To define the function of BMP-15 in mice, we generated embryonic stem cells and knockout mice, which have a null mutation in this X-linked gene. Male chimeric and Bmp15 null mice are normal and fertile. In contrast to Bmp15 null males and Gdf9 knockout females, Bmp15 null females (Bmp15(-/-)) are subfertile and usually have minimal ovarian histopathological defects, but demonstrate decreased ovulation and fertilization rates. To further decipher possible direct or indirect genetic interactions between GDF-9 and BMP-15, we have generated double mutant mice lacking one or both alleles of these related homologs. Double homozygote females (Bmp15(-/-)Gdf9(-/-)) display oocyte loss and cysts and resemble Gdf9(-/-) mutants. In contrast, Bmp15(-/-)Gdf9(+/-) female mice have more severe fertility defects than Bmp15(-/-) females, which appear to be due to abnormalities in ovarian folliculogenesis, cumulus cell physiology, and fertilization. Thus, the dosage of intact Bmp15 and Gdf9 alleles directly influences the destiny of the oocyte during folliculogenesis and in the periovulatory period. These studies have important implications for human fertility control and the maintenance of fertility and normal ovarian physiology.

Animals↗
PubMed2000

Growth differentiation factor-9 stimulates progesterone synthesis in granulosa cells via a prostaglandin E2/EP2 receptor pathway.

Growth differentiation factor-9 (GDF-9), an oocyte-secreted member of the transforming growth factor beta superfamily, progesterone receptor, cyclooxygenase 2 (Cox2; Ptgs2), and the EP2 prostaglandin E(2) (PGE(2)) receptor (EP2; Ptgerep2) are required for fertility in female but not male mice. To define the interrelationship of these factors, we used a preovulatory granulosa cell culture system in which we added recombinant GDF-9, prostaglandins, prostaglandin receptor agonists, or cyclooxygenase inhibitors. GDF-9 stimulated Cox2 mRNA within 2 h, and PGE(2) within 6 h; however, progesterone was not increased until 12 h after addition of GDF-9. This suggested that Cox2 is a direct downstream target of GDF-9 but that progesterone synthesis required an intermediate. To determine whether prostaglandin synthesis was required for progesterone production, we analyzed the effects of PGE(2) and cyclooxygenase inhibitors on this process. PGE(2) can stimulate progesterone synthesis by itself, although less effectively than GDF-9 (3-fold vs. 6-fold increase over 24 h, respectively). Furthermore, indomethacin or NS-398, inhibitors of Cox2, block basal and GDF-9-stimulated progesterone synthesis. However, addition of PGE(2) to cultures containing both GDF-9 and NS-398 overrides the NS-398 block in progesterone synthesis. To further define the PGE(2)-dependent pathway, we show that butaprost, a specific EP2 agonist, stimulates progesterone synthesis and overrides the NS-398 block. In addition, GDF-9 stimulates EP2 mRNA synthesis by a prostaglandin- and progesterone-independent pathway. Thus, GDF-9 induces an EP2 signal transduction pathway which appears to be required for progesterone synthesis in cumulus granulosa cells. These studies further demonstrate the importance of oocyte-somatic cell interactions in female reproduction.

Alprostadil↗
PubMed2000

Disruption of gastrulation and heparan sulfate biosynthesis in EXT1-deficient mice.

Mutations in the EXT1 gene are responsible for human hereditary multiple exostosis type 1. The Drosophila EXT1 homologue, tout-velu, regulates Hedgehog diffusion and signaling, which play an important role in tissue patterning during both invertebrate and vertebrate development. The EXT1 protein is also required for the biosynthesis of heparan sulfate glycosaminoglycans that bind Hedgehog. In this study, we generated EXT1-deficient mice by gene targeting. EXT1 homozygous mutants fail to gastrulate and generally lack organized mesoderm and extraembryonic tissues, resulting in smaller embryos compared to normal littermates. RT-PCR analysis of markers for visceral endoderm and mesoderm development indicates the delayed and abnormal development of both of these tissues. Immunohistochemical staining revealed a visceral endoderm pattern of Indian hedgehog (Ihh) in wild-type E6.5 embryos. However, in both EXT1-deficient embryos and wild-type embryos treated with heparitinase I, Ihh failed to associate with the cells. The effect of the EXT1 deletion on heparan sulfate formation was tested by HPLC and cellular glycosyltransferase activity assays. Heparan sulfate synthesis was abolished in EXT1 -/- ES cells and decreased to less than 50% in +/- cell lines. These results indicate that EXT1 is essential for both gastrulation and heparan sulfate biosynthesis in early embryonic development.

Animals↗