Biomedical subjects

A Dierich

Publications and source records attributed to A Dierich.

At least 37 records · Page 2Linked to original sources

PubMed1999

Cell autonomous and non-cell autonomous functions of Otx2 in patterning the rostral brain.

Previous studies have shown that the homeobox gene Otx2 is required first in the visceral endoderm for induction of forebrain and midbrain, and subsequently in the neurectoderm for its regional specification. Here, we demonstrate that Otx2 functions both cell autonomously and non-cell autonomously in neurectoderm cells of the forebrain and midbrain to regulate expression of region-specific homeobox and cell adhesion genes. Using chimeras containing both Otx2 mutant and wild-type cells in the brain, we observe a reduction or loss of expression of Rpx/Hesx1, Wnt1, R-cadherin and ephrin-A2 in mutant cells, whereas expression of En2 and Six3 is rescued by surrounding wild-type cells. Forebrain Otx2 mutant cells subsequently undergo apoptosis. Altogether, this study demonstrates that Otx2 is an important regulator of brain patterning and morphogenesis, through its regulation of candidate target genes such as Rpx/Hesx1, Wnt1, R-cadherin and ephrin-A2.

Animals↗

PubMed1998

Visualization of CD4/CD8 T cell commitment.

A system to innocuously visualize T cell lineage commitment is described. Using a "knock-in" approach, we have generated mice expressing a beta-galactosidase reporter in place of CD4; expression of beta-galactosidase in these animals appears to be an accurate and early indicator of CD4 gene transcription. We have exploited this knock-in line to trace CD4/CD8 lineage commitment in the thymus, avoiding important pitfalls of past experimental approaches. Our results argue in favor of a selective model of thymocyte commitment, demonstrating a fundamentally symmetrical process: engagement of either class of major histocompatibility complex (MHC) molecule by a differentiating CD4(+)CD8(+) cell can give rise to T cell antigen receptor (TCR)hi thymocytes of either lineage. Key findings include (a) direct demonstration of a substantial number of CD4-committed, receptor/coreceptor-mismatched cells in MHC class II- deficient mice, a critical prediction of the selective model; (b) highly efficient rescue of such "mismatched" intermediates by forced expression of CD8 in a TCR transgenic line, and an explanation of why previous experiments of this nature were less successful-a major past criticism of the selective model; (c) direct demonstration of an analogous, though smaller, population of CD8-committed mismatched intermediates in class I-deficient animals. Finally, we found no evidence of a CD4 default pathway.

Animals↗

PubMed1998

Impairing follicle-stimulating hormone (FSH) signaling in vivo: targeted disruption of the FSH receptor leads to aberrant gametogenesis and hormonal imbalance.

Pituitary gonadotropins follicle-stimulating hormone (FSH) and luteinizing hormone stimulate the gonads by regulating germ cell proliferation and differentiation. FSH receptors (FSH-Rs) are localized to testicular Sertoli cells and ovarian granulosa cells and are coupled to activation of the adenylyl cyclase and other signaling pathways. Activation of FSH-Rs is considered essential for folliculogenesis in the female and spermatogenesis in the male. We have generated mice lacking FSH-R by homologous recombination. FSH-R-deficient males are fertile but display small testes and partial spermatogenic failure. Thus, although FSH signaling is not essential for initiating spermatogenesis, it appears to be required for adequate viability and motility of the sperms. FSH-R-deficient females display thin uteri and small ovaries and are sterile because of a block in folliculogenesis before antral follicle formation. Although the expression of marker genes is only moderately altered in FSH-R -/- mice, drastic sex-specific changes are observed in the levels of various hormones. The anterior lobe of the pituitary gland in females is enlarged and reveals a larger number of FSH- and thyroid-stimulating hormone (TSH)-positive cells. The phenotype of FSH-R -/- mice is reminiscent of human hypergonadotropic ovarian dysgenesis and infertility.

Animals↗

PubMed1998

Contribution of retinoic acid receptor beta isoforms to the formation of the conotruncal septum of the embryonic heart.

To investigate the relative contribution of retinoic acid receptor (RAR)beta isoforms in conotruncal septation, RAR beta 1 and beta 3 were inactivated in the mouse. Mice lacking RAR beta 1 and beta 3 appear normal. Disruption of these isoforms in RAR alpha or RAR gamma null genetic backgrounds results in a high postpartum lethality. However, except for ocular defects found in RAR beta 1-3/RAR gamma compound mutants, the double null mutants display only abnormalities seen in single null mutants. This probably reflects a functional redundancy with other RARs, most notably with RAR beta 2 which is five- to sixfold more abundant than RAR beta 1 and beta 3 and whose domain of expression is largely overlapping. The conotruncal ridges form normally in retinoid X receptor (RXR)alpha/RAR beta compound mutants but fail to fuse, apparently as a result of excessive apoptosis of mesenchymal cells. Additionally, many cardiomyocytes in the conotruncal wall of these mutants appear necrotic. Although RAR beta 1 and beta 3 are expressed specifically in the conotruncal ridges, failure of fusion of these structures is not more frequent in RXR alpha/RAR beta 1-3 double null mutants than in RXR alpha single null mutants. Similarly, the disruption of the sole RAR beta 2 isoform in a RXR alpha null genetic background does not result in an increase of the frequency of conotruncal septum agenesis. However, this agenesis is fully penetrant in RXR alpha/RAR beta +/- mutants, which reflects distinct role of RXR alpha:RAR beta 1 (and beta 3) and RXR alpha:RAR beta 2 heterodimers in promoting the survival of conotruncal mesenchymal cells. Unexpectedly, we discovered that, in wild-type embryos, the conotruncal mesenchyme is a major site of morphogenetic cell death and that conotruncal myocytes are occasionally necrotic. Thus, excessive cell death in the conotruncus is a potential cause of ventricular septal defects in humans.

Animals↗

PubMed1998

In vivo evidence that the stromelysin-3 metalloproteinase contributes in a paracrine manner to epithelial cell malignancy.

Stromelysin-3 (ST3; Basset, P., J.P. Bellocq, C. Wolf, I. Stoll, P. Hutin, J.M. Limacher, O.L. Podhajcer, M.P. Chenard, M.C. Rio, P. Chambon. 1990. Nature. 348:699-704) is a matrix metalloproteinase (MMP) expressed in mesenchymal cells located close to epithelial cells, during physiological and pathological tissue remodeling processes. In human carcinomas, high ST3 levels are associated with a poor clinical outcome, suggesting that ST3 plays a role during malignant processes. In this study we report the ST3 gene inactivation by homologous recombination. Although ST3 null mice (ST3-/-) were fertile and did not exhibit obvious alterations in appearance and behavior, the lack of ST3 altered malignant processes. Thus, the suppression of ST3 results in a decreased 7, 12-dimethylbenzanthracene-induced tumorigenesis in ST3-/- mice. Moreover, ST3-/- fibroblasts have lost the capacity to promote implantation of MCF7 human malignant epithelial cells in nude mice (P < 0.008). Finally, we show that this ST3 paracrine function requires extracellular matrix (ECM)-associated growth factors. Altogether, these findings give evidence that ST3 promotes, in a paracrine manner, homing of malignant epithelial cells, a key process for both primary tumors and metastases. Therefore, ST3 represents an appropriate target for specific MMP inhibitor(s) in future therapeutical approaches directed against the stromal compartment of human carcinomas.

9,10-Dimethyl-1,2-benzanthracene↗

PubMed1998

Disruption of the kappa-opioid receptor gene in mice enhances sensitivity to chemical visceral pain, impairs pharmacological actions of the selective kappa-agonist U-50,488H and attenuates morphine withdrawal.

***micro***-, delta- and kappa-opioid receptors are widely expressed in the central nervous system where they mediate the strong analgesic and mood-altering actions of opioids, and modulate numerous endogenous functions. To investigate the contribution of the kappa-opioid receptor (KOR) to opioid function in vivo, we have generated KOR-deficient mice by gene targeting. We show that absence of KOR does not modify expression of the other components of the opioid system, and behavioural tests indicate that spontaneous activity is not altered in mutant mice. The analysis of responses to various nociceptive stimuli suggests that the KOR gene product is implicated in the perception of visceral chemical pain. We further demonstrate that KOR is critical to mediate the hypolocomotor, analgesic and aversive actions of the prototypic kappa-agonist U-50, 488H. Finally, our results indicate that this receptor does not contribute to morphine analgesia and reward, but participates in the expression of morphine abstinence. Together, our data demonstrate that the KOR-encoded receptor plays a modulatory role in specific aspects of opioid function.

3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cycloh↗

PubMed1998

The bHLH protein NEUROGENIN 2 is a determination factor for epibranchial placode-derived sensory neurons.

neurogenin2 encodes a neural-specific basic helix-loop-helix (bHLH) transcription factor related to the Drosophila proneural factor atonal. We show here that the murine ngn2 gene is essential for development of the epibranchial placode-derived cranial sensory ganglia. An ngn2 null mutation blocks the delamination of neuronal precursors from the placodes, the first morphological sign of differentiation in these lineages. Mutant placodal cells fail to express downstream bHLH differentiation factors and the Notch ligand Delta-like 1. These data suggest that ngn2 functions like the Drosophila proneural genes in the determination of neuronal fate in distal cranial ganglia. Interestingly, the homeobox gene Phox2a is activated independently of ngn2 in epibranchial placodes, suggesting that neuronal fate and neuronal subtype identity may be specified independently in cranial sensory ganglia.

Animals↗

PubMed1998

The RXRalpha ligand-dependent activation function 2 (AF-2) is important for mouse development.

We have engineered a mouse mutation that specifically deletes the C-terminal 18 amino acid sequence of the RXRalpha protein. This deletion corresponds to the last helical alpha structure (H12) of the ligand-binding domain (LBD), and includes the core of the Activating Domain of the Activation Function 2 (AF-2 AD core) that is thought to be crucial in mediating ligand-dependent transactivation by RXRalpha. The homozygous mutants (RXRalpha af2(o)), which die during the late fetal period or at birth, exhibit a subset of the abnormalities previously observed in RXRalpha -/- mutants, often with incomplete penetrance. In marked contrast, RXRalpha af2(o)/RXRbeta -/- and RXRalpha af2(o)/RXRbeta -/- /RXRgamma -/- compound mutants display a large array of malformations, which nearly recapitulate the full spectrum of the defects that characterize the fetal vitamin A-deficiency (VAD) syndrome and were previously found in RAR single and compound mutants, as well as in RXRalpha/RAR(alpha, beta or gamma) compound mutants. Analysis of RXRalpha af2(o)/RAR(alpha, beta or gamma) compound mutants also revealed that they exhibit many of the defects observed in the corresponding RXR alpha/RAR compound mutants. Together, these results demonstrate the importance of the integrity of RXR AF-2 for the developmental functions mediated by RAR/RXR heterodimers, and hence suggest that RXR ligand-dependent transactivation is instrumental in retinoid signalling during development.

Abnormalities, Multiple↗

PubMed1998

Sequential roles for Otx2 in visceral endoderm and neuroectoderm for forebrain and midbrain induction and specification.

The homeobox gene Otx2 is a mouse cognate of the Drosophila orthodenticle gene, which is required for development of the brain, rostral to rhombomere three. We have investigated the mechanisms involved in this neural function and specifically the requirement for Otx2 in the visceral endoderm and the neuroectoderm using chimeric analysis in mice and explant recombination assay. Analyses of chimeric embryos composed of more than 90% of Otx2-/- ES cells identified an essential function for Otx2 in the visceral endoderm for induction of the forebrain and midbrain. The chimeric studies also demonstrated that an anterior neural plate can form without expressing Otx2. However, in the absence of Otx2, expression of important regulatory genes, such as Hesx1/Rpx, Six3, Pax2, Wnt1 and En, fail to be initiated or maintained in the neural plate. Using explant-recombination assay, we could further demonstrate that Otx2 is required in the neuroectodem for expression of En. Altogether, these results demonstrate that Otx2 is first required in the visceral endoderm for the induction, and subsequently in the neuroectoderm for the specification of forebrain and midbrain territories.

Animals↗

PubMed1997

Requirement of poly(ADP-ribose) polymerase in recovery from DNA damage in mice and in cells.

Poly(ADP-ribose) polymerase [PARP; NAD+ ADP-ribosyltransferase; NAD+: poly(adenosine-diphosphate-D-ribosyl)-acceptor ADP-D-ribosyltransferase, EC 2.4.2.30] is a zinc-finger DNA-binding protein that detects specifically DNA strand breaks generated by genotoxic agents. To determine its biological function, we have inactivated both alleles by gene targeting in mice. Treatment of PARP-/- mice either by the alkylating agent N-methyl-N-nitrosourea (MNU) or by gamma-irradiation revealed an extreme sensitivity and a high genomic instability to both agents. Following whole body gamma-irradiation (8 Gy) mutant mice died rapidly from acute radiation toxicity to the small intestine. Mice-derived PARP-/- cells displayed a high sensitivity to MNU exposure: a G2/M arrest in mouse embryonic fibroblasts and a rapid apoptotic response and a p53 accumulation were observed in splenocytes. Altogether these results demonstrate that PARP is a survival factor playing an essential and positive role during DNA damage recovery.

Alleles↗

PubMed1997

Limited specificity and large overlap of the functions of the mouse RAR gamma 1 and RAR gamma 2 isoforms.

The RAR gamma gene generates two major isoforms, RAR gamma 1 and RAR gamma 2, which originate from two distinct promoters. We report here the engineering of mice lacking RAR gamma 1, but in which RAR gamma 2 is normally expressed. The effect of this null mutation has been compared with those previously described for RAR gamma 2 and all RAR gamma isoforms (total RAR gamma gene inactivation), both in single mutants and in double mutants bearing additional null mutations in their RAR alpha, RAR beta or RXR alpha genes. RAR gamma 1 mutants, but not RAR gamma 2 mutants, displayed a subset of the abnormalities exhibited by total RAR gamma null mutants (growth deficiency, abnormal cricoid cartilage and occasional cervical vertebra defects), suggesting that RAR gamma 1 is the main isoform mediating the corresponding RAR gamma functions. Interestingly, cricoid cartilage defects were also found in a fraction of heterozygote animals for the RAR gamma 1, RAR gamma or RAR alpha mutations, indicating that wild type levels of RARs are required for the normal morphogenesis of this structure. Compound RAR alpha/RAR gamma 1 and RAR alpha/RAR gamma 2 double null mutants exhibited only a small fraction of the defects found in RAR alpha/RAR gamma double null mutants. Moreover, these defects were often partially penetrant, or corresponded to a less severe form. However, they occurred preferentially in certain compound mutants, demonstrating that given isoforms mediate specific functions of RAR gamma in the context of a RAR alpha null background. In a RXR alpha null background, both RAR gamma 1 and gamma 2 isoform mutations resulted in increased severity of the RXR alpha null ocular phenotype. Together, the present observations indicate that the functions of the two RAR gamma isoforms overlap to a large extent, but also that each of these isoforms exhibits a limited functional specificity. Furthermore, the occurrence of morphological defects in heterozygote mutants for a single RAR isoform provides a basis for explaining the strong conservation of these isoforms during vertebrate evolution.

Animals↗

PubMed1997

In vivo functional analysis of the Hoxa-1 3' retinoic acid response element (3'RARE).

Retinoids are essential for normal development and both deficiency and excess of retinoic acid (RA) are teratogenic. Retinoic acid response elements (RAREs) have been identified in Hox gene promoters suggesting that endogenous retinoids may be involved in the direct control of Hox gene patterning functions. In order to test this hypothesis, we have mutated the Hoxa-1 3'RARE using the Cre-loxP targeting strategy, and studied its functional role during mouse development. We find that this enhancer plays an important role in the early establishment of the Hoxa-1 anterior expression boundary in the neural plate. This early disturbance in Hoxa-1 activation results in rhombomere and cranial nerve abnormalities reminiscent of those obtained in the Hoxa-1 total knockout, although their severity and penetrance are lower, thus providing strong evidence for direct control of Hox gene function by retinoids during normal development. Interestingly, we also find that the Hoxa-1 expression response to RA treatment is not entirely controlled by the RARE, suggesting the existence of other retinoid-induced factors mediating the Hoxa-1 response to RA and/or the presence of additional RAREs. Interestingly, although the RARE is not required for the spatiotemporal control of colinear expression of the Hoxa genes, it is absolutely required for correct Hoxa-2 expression in rhombomere 5.

Animals↗

PubMed1997

Role of the retinoic acid receptor beta (RARbeta) during mouse development.

Homozygous RAR beta mutants are growth-deficient, but are fertile and have a normal longevity. They display homeotic transformations and malformations of cervical vertebrae and a retrolenticular membrane. This latter abnormality arises from the persistence and hyperplasia of the primary vitreous body. In contrast, we found that abnormalities of cranial nerves IX and X which were previously proposed to be specific features of the RAR beta mutant phenotype (Luo et al., Mech. Dev. 53: 61-71, 1995) occur with the same low penetrance in wildtype littermates. Although the RAR beta protein is expressed at high levels in the striatum and interdigital mesenchyme, the brain and limbs of RAR beta mutants appear morphologically normal. RAR alpha/RAR beta double mutants display numerous visceral abnormalities, most of which are incompatible with post-natal life. The majority of these abnormalities was previously detected in RAR alpha/RAR beta2 mutants with the notable exceptions of agenesis of the stapedial (2nd aortic arch-derived) artery, thymic and spleen agenesis and abnormal inferior vena cava. RAR beta/RAR gamma double mutants show major ocular defects including a shortening of the ventral retina and pre-natal retinal dysplasia, both of which represent the only abnormalities of the fetal vitamin-A deficiency (VAD) syndrome not previously detected in RAR beta2/RAR gamma compound mutants. In addition, RAR beta is apparently functionally redundant with either RAR alpha or RAR gamma for the formation of a small subset of craniofacial skeletal elements, as well as for eyelid development and digit separation. We also provide evidence that, at least in some instances, this phenomenon of functional redundancy between RARs may be an artifactual consequence of gene knock-out.

Animals↗

PubMed1996

Loss of morphine-induced analgesia, reward effect and withdrawal symptoms in mice lacking the mu-opioid-receptor gene.

Despite tremendous efforts in the search for safe, efficacious and non-addictive opioids for pain treatment, morphine remains the most valuable painkiller in contemporary medicine. Opioids exert their pharmacological actions through three opioid-receptor classes, mu, delta and kappa, whose genes have been cloned. Genetic approaches are now available to delineate the contribution of each receptor in opioid function in vivo. Here we disrupt the mu-opioid-receptor gene in mice by homologous recombination and find that there are no overt behavioural abnormalities or major compensatory changes within the opioid system in these animals. Investigation of the behavioural effects of morphine reveals that a lack of mu receptors abolishes the analgesic effect of morphine, as well as place-preference activity and physical dependence. We observed no behavioural responses related to delta- or kappa-receptor activation with morphine, although these receptors are present and bind opioid ligands. We conclude that the mu-opioid-receptor gene product is the molecular target of morphine in vivo and that it is a mandatory component of the opioid system for morphine action.

Analgesics↗

PubMed1996

Gastric mucosa abnormalities and tumorigenesis in mice lacking the pS2 trefoil protein.

To determine the function of the pS2 trefoil protein, which is normally expressed in the gastric mucosa, the mouse pS2 (mpS2) gene was inactivated. The antral and pyloric gastric mucosa of mpS2-null mice was dysfunctional and exhibited severe hyperplasia and dysplasia. All homozygous mutant mice developed antropyloric adenoma, and 30 percent developed multifocal intraepithelial or intramucosal carcinomas. The small intestine was characterized by enlarged villi and an abnormal infiltrate of lymphoid cells. These results indicate that mpS2 is essential for normal differentiation of the antral and pyloric gastric mucosa and may function as a gastric-specific tumor suppressor gene.

Adenoma↗

PubMed1996

RXR gamma null mice are apparently normal and compound RXR alpha +/-/RXR beta -/-/RXR gamma -/- mutant mice are viable.

The RXR gamma (RXR, retinoid X receptor) gene was disrupted in the mouse. Homozygous mutant mice developed normally and were indistinguishable from their RXR gamma +/- or wild-type littermates with respect to growth, fertility, viability, and apparent behavior in the animal facility. Moreover, RXR alpha -/-/RXR gamma -/- and RXR beta -/-/RXR gamma -/- mutant phenotypes were indistinguishable from those of RXR alpha -/- and RXR beta -/- mutants, respectively. Strikingly, RXR alpha +/-/RXR beta -/-/RXR gamma -/- triple mutants were viable. Thus, it appears that RXR gamma does not exert any essential function that cannot be performed by RXR alpha or RXR beta, and one copy of RXR alpha is sufficient to perform most of the functions of the RXRs.

Animals↗

PubMed1996

Independent modes of natural killing distinguished in mice lacking Lag3.

The LAG3 protein has several features in common with CD4, suggesting that it may be important in controlling T cell reactivity. However, mice with a Lag3 null mutation have now been shown to exhibit a defect in the natural killer cell, rather than the T cell, compartment. Killing of certain tumor targets by natural killer cells from these mice was inhibited or even abolished, whereas lysis of cells displaying major histocompatibility complex class I disparities remained intact. It appears that LAG3 is a receptor or coreceptor that defines different modes of natural killing.

Animals↗

PubMed1996

Spermiogenesis deficiency and germ-cell apoptosis in CREM-mutant mice.

Spermiogenesis is a complex process by which postmeiotic male germ cells differentiate into mature spermatozoa. This process involves remarkable structural and biochemical changes including nuclear DNA compaction and acrosome formation. Transcription activator CREM (cyclic AMP-responsive element modulator) is highly expressed in postmeiotic cells, and CREM may be responsible for the activation of several haploid germ cell-specific genes involved in the structuring of the spermatozoon. The specific role of CREM in spermiogenesis was addressed using CREM-mutant mice generated by homologous recombination. Analysis of the seminiferous epithelium in mutant male mice reveals postmeiotic arrest at the first step of spermiogenesis. Late spermatids are completely absent, and there is a significant increase in apoptotic germ cells. We show that CREM deficiency results in the lack of postmeiotic cell-specific gene expression. The complete lack of spermatozoa in the mutant mice is reminiscent of cases of human infertility.

Animals↗