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At least 883 records · Page 49Linked to original sources
PubMed2005

Genistein, a dietary isoflavone, down-regulates the MDM2 oncogene at both transcriptional and posttranslational levels.

Although genistein has chemopreventive effects in several human malignancies, including cancers of the breast, colon, and prostate, the mechanisms of action are not fully understood. Herein we report novel mechanisms whereby genistein down-regulates the MDM2 oncogene, perhaps explaining some of its anticancer activities. In a dose- and time-dependent manner, genistein reduced MDM2 protein and mRNA levels in human cell lines of breast, colon, and prostate cancer; primary fibroblasts; and breast epithelial cells. The inhibitory effects were found at both transcriptional and posttranslational levels and were independent of tyrosine kinase pathways. We found that the NFAT transcription site in the region between -132 and +33 in the MDM2 P2 promoter was responsive to genistein. At the posttranslational level, genistein induced ubiquitination of MDM2, which led to its degradation. Additionally, genistein induced apoptosis and G2 arrest and inhibited proliferation in a variety of human cancer cell lines, regardless of p53 status. We further showed that MDM2 overexpression abrogated genistein-induced apoptosis in vitro and that genistein inhibited MDM2 expression and tumor growth in PC3 xenografts. In conclusion, genistein directly down-regulates the MDM2 oncogene, representing a novel mechanism of its action that may have implications for its chemopreventive and chemotherapeutic effects.

Animals↗
PubMed2006

4-nitroquinoline-1-oxide induces the formation of cellular topoisomerase I-DNA cleavage complexes.

RecQ helicase BLM-deficient cells are characteristically hypersensitive to 4-nitroquinoline-1-oxide (4NQO). We recently reported that isogenic BLM-deficient cells (PNSG13) are more sensitive than BLM-complemented cells (PNSF5) to camptothecin, which specifically traps topoisomerase I cleavage complexes (Top1cc). We now report that PNSG13 are also 3.5-fold more sensitive to 4NQO compared with PNSF5 and that 4NQO induces higher levels of Top1cc and reduced histone gamma-H2AX in PSNG13 than in PNSF5. Similarly, 4NQO induces more Top1cc in primary fibroblasts from a patient with Bloom syndrome than in normal human fibroblasts. 4NQO also induces Top1cc in colon cancer HCT116 and HT29 cells in a time- and concentration-dependent fashion. Of note, distinct from camptothecin, the Top1cc produced by 4NQO accumulate progressively after 4NQO addition and persist following 4NQO removal. The Top1cc induced by 4NQO are detectable by alkaline elution. To examine the functional relevance of the Top1cc induced by 4NQO, we used two stable topoisomerase I small interfering RNA (siRNA) cell lines derived from HCT116 and MCF7 cells. Both topoisomerase I siRNA cell lines are resistant to 4NQO, indicating that Top1cc contribute to the cellular activity of 4NQO. Collectively, these data show that 4NQO is an effective inducer of cellular Top1cc. Because 4NQO does not directly trap Top1cc in biochemical assays, we propose that active metabolites of 4NQO trap Top1cc by forming DNA adducts. Induction of Top1cc and histone gamma-H2AX by 4NQO may contribute to the cellular effects of 4NQO, including its selective activity toward RecQ helicase BLM-deficient cells.

4-Nitroquinoline-1-oxide↗
PubMed2005

Chromosomal radiosensitivity in two cell lineages derived from clinically radiosensitive cancer patients.

PURPOSE: Despite its prominent contribution to cancer cure and palliation, around 1% to 5% of cancer patients suffer serious side effects from radiotherapy. A cardinal goal in the fields of radiobiology and oncology is to predict normal tissue radiosensitivity of a cancer patient before radiotherapy. Higher tumor control rates are likely if radiotherapy individualization could be achieved by applying predictive approaches. EXPERIMENTAL DESIGN: Here, we make use of the cytokinesis block micronucleus assay to assess radiosensitivity in cell lines derived from two different cell lineages obtained from clinically radiosensitive patients. We determined the micronucleus frequency after graded doses of ionizing radiation to primary fibroblasts and lymphoblast cell lines derived from 36 highly radiosensitive cancer patients. RESULTS: Many cell lines, following exposure to ionizing radiation, from patients with severe clinical reactions to radiotherapy showed statistically significantly higher frequencies of micronuclei than those from patients who had normal reactions to radiotherapy. One individual revealed significantly higher micronucleus frequencies in both cell lineages. Interestingly, lymphoblast cell lines from one patient showed micronucleus frequencies similar to ataxia telangiectasia mutated-deficient cells. CONCLUSIONS: These results indicate that the micronucleus assay may have use for identifying predisposition to clinical radiosensitivity, at least in a subset of patients as a component of a pretreatment radiosensitivity assay for use in the clinic.

Cell Lineage↗
PubMed2006

Accurate prediction of BRCA1 and BRCA2 heterozygous genotype using expression profiling after induced DNA damage.

PURPOSE: In this study, the differential gene expression changes following radiation-induced DNA damage in healthy cells from BRCA1/BRCA1 mutation carriers have been compared with controls using high-density microarray technology. We aimed to establish if BRCA1/BRCA2 mutation carriers could be distinguished from noncarriers based on expression profiling of normal cells. EXPERIMENTAL DESIGN: Short-term primary fibroblast cultures were established from skin biopsies from 10 BRCA1 and 10 BRCA2 mutation carriers and 10 controls, all of whom had previously had breast cancer. The cells were subjected to 15 Gy ionizing irradiation to induce DNA damage. RNA was extracted from all cell cultures, preirradiation and at 1 hour postirradiation. For expression profiling, 15 K spotted cDNA microarrays manufactured by the Cancer Research UK DNA Microarray Facility were used. Statistical feature selection was used with a support vector machine (SVM) classifier to determine the best feature set for predicting BRCA1 or BRCA2 heterozygous genotype. To investigate prediction accuracy, a nonprobabilistic classifier (SVM) and a probabilistic Gaussian process classifier were used. RESULTS: In the task of distinguishing BRCA1 and BRCA2 mutation carriers from noncarriers and from each other following radiation-induced DNA damage, the SVM achieved 90%, and the Gaussian process classifier achieved 100% accuracy. This effect could not be achieved without irradiation. In addition, the SVM identified a set of BRCA genotype predictor genes. CONCLUSIONS: We conclude that after irradiation-induced DNA damage, BRCA1 and BRCA2 mutation carrier cells have a distinctive expression phenotype, and this may have a future role in predicting genotypes, with application to clinical detection and classification of mutations.

Adult↗
PubMed2000

Cloning of the murine Drm gene (Cktsf1b1) and characterization of its oncogene suppressible promoter.

The Drm gene, first identified in rat cells in our laboratory, appears to play a significant role in early embryo patterning and limb bud development. We have now isolated mouse Drm (mDrm) cDNA as well as genomic DNA clones and have mapped the Drm gene (Cktsf1b1) to murine chromosome 2. Cktsf1b1 is regulated in a tissue specific fashion and is expressed only in nontransformed mouse cells or primary fibroblasts in culture, but not in established transformed or tumor-derived mouse cell lines. The major transcription start sites map to within 69 bp upstream of the initiating ATG. A promoter was contained in the -214 to +1 bp 5' flanking region, and promoter/reporter constructs showed 10-fold higher activity than control in REF-1 (rat), A31 (mouse) and CHO (hamster) cells. The region contains a TATA sequence and multiple potential transcription factor binding sites. Promoter activity was dose-dependently inhibited by cotransfection with either ras or mos oncogenes, but oncogene inhibition was reversed and the overall activity increased when cells were treated with the MAP kinase kinase (MKK) inhibitor PD98059. An NF-1 and Yi-like site, identified in the minimal promoter region, showed different mobility shift patterns when normal and transformed cell nuclear extracts are compared. Mutation of the NF-1 site reduced Cktsf1b1 promoter activity 25%, while mutation of the Yi-like site destroyed all the activity. Our results indicate that the expression of Cktsf1b1, a gene associated with early development and cell transformation, is sensitive to MKK levels and may be regulated via multiple transcription factor complexes.

Animals↗
PubMedDate not supplied

The p16/Cdkn2a/Ink4a gene is frequently deleted in nitrosourea-induced rat glial tumors.

The present study investigates nitrosourea-induced rat (Rattus norvegicus) glioma cell lines for the functional status of the p16/Cdkn2a/Ink4a gene, which encodes the p16 cdk4 inhibitor and the alternative reading frame protein, p19ARF. We detected homozygous deletions of the p16/Cdkn2a/Ink4a gene locus in 4 of 5 glioma cell lines (C6, F98, RG2, and RGL.3), but not in the 9L gliosarcoma cell line or in a rat primary fibroblast cell line. RT-PCR demonstrated expression of the p16 and p19ARF mRNAs only in 9L cells and in rat fibroblasts. Comparative genomic in situ hybridization showed that the copy number of rat chromosome RNO5 was not altered in any of the glioma cell lines investigated, indicating that the deletions result from a discrete loss in the region of the p16/Cdkn2a/Ink4a locus. This is the first report of p16/Cdkn2a/Ink4a deletions present in nitrosourea-induced rat glioma cell lines. Since this genetic alteration is also commonly observed in human malignant glial tumors, our results validate the use of chemically induced rat glioma cell lines as an experimental model in the development of gene therapy strategies.

Animals↗
PubMed1991

Comparison of RBG-banded karyotypes of cattle, sheep, and goats.

The karyotypes of the three main domestic Bovidae species. Bos taurus, Ovis aries, and Capra hircus, have been investigated by RBG-banding. Primary fibroblast cells were cultured from fetal lung or muscle tissue and collected for chromosome preparation after double thymidine synchronization. Depending on the culture and species, BrdU and FdU were added for 8-11 h or 7-10 h, respectively, before harvest. The chromosomes of the three species were classified and numbered according to standard conventions (ISCNDA, 1989) and compared at different stages of condensation. Within the resolving power of the images obtained, the RBG-banding patterns appear to be very similar, with only minor differences existing between chromosome 9 and the sex chromosomes of cattle and the same chromosomes in the domestic sheep and the goat.

Animals↗
PubMed2002

Differences in the fibrogenic response after transfer of active transforming growth factor-beta1 gene to lungs of "fibrosis-prone" and "fibrosis-resistant" mouse strains.

Pulmonary fibrosis is characterized by excessive deposition of extracellular matrix in the interstitium, resulting in impaired lung function and respiratory failure. Investigation of the differences in individual susceptibility to the development of fibrosis may help to detect patients that are at risk to fibrosis when exposed to fibrogenic stimuli. In this study we used adenoviral gene transfer to transiently expose a fibrosis-prone (C57BL/6) and a fibrosis-resistant (Balb/c) mouse strain to high levels of active transforming growth factor (TGF)-beta1, a key profibrotic cytokine. Balb/c mice developed significantly less fibrosis compared with C57BL/6 mice in response to active TGF-beta1 despite higher levels of the transgene protein in the lung. This was not due to a general unresponsiveness of cells to TGF-beta1, because primary fibroblasts of both strains increased collagen synthesis upon stimulation with TGF-beta1 in vitro to the same degree. However, TGF-beta1 induced a strong upregulation of tissue inhibitor of metalloprotease-1 gene in pulmonary fibroblasts as well as in lungs of C57BL/6 mice, in contrast to a weak induction in Balb/c mice. These findings suggest that the differences in susceptibility to pulmonary fibrosis are downstream from TGF-beta1 and that fibrosis-prone individuals may have an altered collagen metabolism in the lungs that is balanced toward a "nondegrading" environment.

Adenoviridae↗
PubMedDate not supplied

Polymer-encapsulated Schwannoma cells expressing human nerve growth factor promote the survival of cholinergic neurons after a fimbria-fornix transection.

Many investigators have recently used genetically modified primary fibroblasts of fibroblast cell lines (e.g., 3T3, 208F, or BHK cells) to deliver recombinant nerve growth factor (NGF) into the CNS. In the current study, SCT-1 cells, a Schwannoma cell line derived from a transgenic mouse, were transfected with a human NGF (hNGF) cDNA. After selection, these cells were encased within a polymer capsule and implanted into the ventricles of fimbria-fornix lesioned rats. Encapsulated, non-transfected cells served as controls. Results demonstrated that the hNGF transgene is expressed for at least 3 weeks after implantation. Moreover, the cells did not overgrow the capsule. Recombinant hNGF was able to save > 70% of lesioned cholinergic neurons, as assessed by NGF-receptor (NGFr) and choline acetyltransferase (ChAT) immunohistochemistry, from cell death. The number of cholinergic neurons in animals that received control capsules (i.e., nontransfected SCT-1 cells) was similar to lesion only animals (i.e., approximately 27% and approximately 33% for NGFr- and ChAT-positive neurons, respectively. These results show that SCT-1 cells can be used to deliver biologically active hNGF into the lesioned rat brain.

Animals↗
PubMed2004

NF-kappaB protection against apoptosis induced by HEMA.

The cytotoxicity of dental monomers has been widely investigated, but the underlying mechanisms have not been elucidated. We studied the molecular mechanisms involved in cell death induced by HEMA. In human primary fibroblasts, HEMA induced a dose-dependent apoptosis that was confirmed by the activation of caspases-8, -9, and -3. We found an increase of reactive oxygen species (ROS) and NF-kappaB activation after HEMA exposure. Blocking of ROS production by anti-oxidants had no direct influence on apoptosis caused by HEMA, but inhibition of NF-kappaB increased the fraction of apoptotic cells. Accordingly, mouse embryonic fibroblasts (MEF) from p65-/- mice were more susceptible to HEMA-induced apoptosis than were wild-type controls. Our results indicate that exposure to HEMA triggers apoptosis and that this mechanism is not directly dependent upon redox signaling. Nevertheless, ROS induction by HEMA activates NF-kappaB, which exerts a protective role in counteracting apoptosis.

Analysis of Variance↗
PubMed1998

A novel coculture model for benign prostatic hyperplasia expressing both isoforms of 5 alpha-reductase.

We have developed a coculture system for primary fibroblast and epithelial cells derived from benign prostatic hyperplasia (BPH) that retained many of the characteristics of the intact human prostate. In contrast to separately cultured prostate fibroblast and epithelial cells, cocultures of fibroblasts and epithelial cells maintained messenger ribonucleic acid expression and functional activity for both isoenzymes of 5 alpha-reductase (type I and type II) as well as maintained expression of androgen receptors and prostate-specific antigen. Furthermore, levels of prostate-specific antigen secreted by cocultured epithelial cells were increased by treatment with androgens, mimicking the situation in the human gland. This contrasted with conventionally cultured fibroblasts or epithelial cells, which failed to express 50 alpha-reductase type II and rapidly lost expression of androgen receptors and androgen sensitivity upon being placed into culture. Electron microscopy demonstrated intracellular structures indicative of the differentiated state of the cocultured cell types, including round nuclei, tonofibrils, and microvilli in epithelial cells and elongated nuclei; large amounts of Golgi and cilia; along with immature collagen fibers in fibroblasts. The present study demonstrates that the coculture model reflects more closely the in vivo system for human BPH and is thus a far more suitable model for investigating the molecular and cellular events that underlie BPH than current in vitro systems.

3-Oxo-5-alpha-Steroid 4-Dehydrogenase↗
PubMed2003

Hyaluronan-oligosaccharide-induced transcription of metalloproteases.

Activated dendritic epidermal Langerhans cells and metastatic tumour cells share many properties. Both cell types can invade the surrounding tissue, enter the lymphatic system and travel to regional lymph nodes. We have recently shown that fragments of the extracellular matrix component hyaluronan, which are typically produced at sites of inflammation, can activate dendritic cells. Upon activation, dendritic cells upregulate expression of matrix metalloproteases (MMPs). These observations prompted us to investigate whether exposure to hyaluronan fragments also induces MMP expression in tumour cells. Here, we report that MMP-9, MMP-13 and urokinase plasminogen activator are upregulated in murine 3LL tumour cells after exposure to mixed-size hyaluronan. Similarly upregulated MMP-9 and MMP-13 expression was observed in primary fibroblasts. By using size-fractionated hyaluronan preparations, we show that the enhanced expression of MMP-9 and MMP-13 is only induced by small hyaluronan (HA) fragments. Although our data suggest that HA-fragment-induced MMP-9 and MMP-13 expression is receptor mediated, they rule out an involvement of the hyaluronan receptors CD44, RHAMM/IHAP and TLR-4. Finally, we show that HA fragment-induced MMP-9 transcription is mediated via NF-kappa B. Our results suggest that the metastasis-associated HA degradation in tumours might promote invasion by inducing MMP expression.

Animals↗
PubMed2005

Neurogenic potential of human mesenchymal stem cells revisited: analysis by immunostaining, time-lapse video and microarray.

The possibility of generating neural cells from human bone-marrow-derived mesenchymal stem cells (hMSCs) by simple in vitro treatments is appealing both conceptually and practically. However, whether phenotypic modulations observed after chemical manipulation of such stem cells truly represent a genuine trans-lineage differentiation remains to be established. We have re-evaluated the effects of a frequently reported biochemical approach, based on treatment with butylated hydroxyanisole and dimethylsulphoxide, to bring about such phenotypic conversion by monitoring the morphological changes induced by the treatment in real time, by analysing the expression of phenotype-specific protein markers and by assessing the modulation of transcriptome. Video time-lapse microscopy showed that conversion of mesenchymal stem cells to a neuron-like morphology could be reproduced in normal primary fibroblasts as well as mimicked by addition of drugs eliciting cytoskeletal collapse and disruption of focal adhesion contacts. Analysis of markers revealed that mesenchymal stem cells constitutively expressed multi-lineage traits, including several pertaining to the neural one. However, the applied ;neural induction' protocol neither significantly modulated the expression of such markers, nor induced de novo translation of other neural-specific proteins. Similarly, global expression profiling of over 21,000 genes demonstrated that gene transcription was poorly affected. Most strikingly, we found that the set of genes whose expression was altered by the inductive treatment did not match those sets of genes differentially expressed when comparing untreated mesenchymal stem cells and immature neural tissues. Conversely, by comparing these gene expression profiles with that obtained from comparisons between the same cells and an unrelated non-neural organ, such as liver, we found that the adopted neural induction protocol was no more effective in redirecting human mesenchymal stem cells toward a neural phenotype than toward an endodermal hepatic pathway.

Adult↗
PubMed1995

Delayed cyclin A and B1 degradation in non-transformed mammalian cells.

Cyclins A and B are known to exhibit significant differences in their function, cellular distribution and timing of degradation at mitosis. On the basis of observations in marine invertebrates and Xenopus, it was proposed that cyclin destruction triggers cdc2 kinase inactivation and anaphase onset. However, this model has recently been questioned, both in Xenopus and in budding yeast. In this report, we present evidence for delayed degradation of both cyclins A and B1 in non-transformed mammalian cells. Indeed, by means of indirect immunofluorescence and confocal microscopy, we show that cyclins A and B1 are present up to anaphase in REF52, Hs68, human primary fibroblasts and NRK epithelial cells. In marked contrast, cyclin A is shown to be degraded within metaphase and cyclin B just at the transition to anaphase in HeLa and two transformed cell lines, derivatives of normal NRK and REF52. These results further support the notion that cyclin destruction might be not correlated with anaphase onset in normal cells and highlight a significant difference in the fate of mitotic cyclins between transformed and non-transformed cells.

Animals↗
PubMed2000

Rapid and reciprocal regulation of tenascin-C and tenascin-Y expression by loading of skeletal muscle.

Tenascin-C and tenascin-Y are two structurally related extracellular matrix glycoproteins that in many tissues show a complementary expression pattern. Tenascin-C and the fibril-associated minor collagen XII are expressed in tissues bearing high tensile stress and are located in normal skeletal muscle, predominantly at the myotendinous junction that links muscle fibers to tendon. In contrast, tenascin-Y is strongly expressed in the endomysium surrounding single myofibers, and in the perimysial sheath around fiber bundles. We previously showed that tenascin-C and collagen XII expression in primary fibroblasts is regulated by changes in tensile stress. Here we have tested the hypothesis that the expression of tenascin-C, tenascin-Y and collagen XII in skeletal muscle connective tissue is differentially modulated by mechanical stress in vivo. Chicken anterior latissimus dorsi muscle (ALD) was mechanically stressed by applying a load to the left wing. Within 36 hours of loading, expression of tenascin-C protein was ectopically induced in the endomysium along the surface of single muscle fibers throughout the ALD, whereas tenascin-Y protein expression was barely affected. Expression of tenascin-C protein stayed elevated after 7 days of loading whereas tenascin-Y protein was reduced. Northern blot analysis revealed that tenascin-C mRNA was induced in ALD within 4 hours of loading while tenascin-Y mRNA was reduced within the same period. In situ hybridization indicated that tenascin-C mRNA induction after 4 hours of loading was uniform throughout the ALD muscle in endomysial fibroblasts. In contrast, the level of tenascin-Y mRNA expression in endomysium appeared reduced within 4 hours of loading. Tenascin-C mRNA and protein induction after 4-10 hours of loading did not correlate with signs of macrophage infiltration. Tenascin-C protein decreased again with removal of the load and nearly disappeared after 5 days. Furthermore, loading was also found to induce expression of collagen XII mRNA and protein, but to a markedly lower level, with slower kinetics and only partial reversibility. The results suggest that mechanical loading directly and reciprocally controls the expression of extracellular matrix proteins of the tenascin family in skeletal muscle.

Animals↗
PubMed1983

Bacillus thuringiensis var israelensis crystal delta-endotoxin: effects on insect and mammalian cells in vitro and in vivo.

Bacillus thuringiensis var israelensis parasporal crystal delta-endotoxin was purified by ultracentrifugation on a discontinuous sucrose gradient. Native delta-endotoxin crystals showed no detectable toxicity in the vitro and in vivo systems that are described. By contrast alkali-solubilized crystal delta-endotoxin caused rapid cytological and cytopathological changes in Aedes albopictus, Choristoneura fumiferana 63 CF1, Spodoptera frugiperda and Trichoplusia ni cell lines as observed by phase-contrast microscopy and vital staining. Mouse fibroblasts, primary pig lymphocytes and three mouse epithelial carcinoma cell types showed a similar response to the alkali-soluble crystal delta-endotoxin. In addition the soluble crystal delta-endotoxin protein caused haemolysis of rat, mouse, sheep, horse and human erythrocytes. Intravenous administration of the alkali-soluble crystal delta-endotoxin to Balb. c mice at a dose rate of 15-30 micrograms of protein per gram body weight resulted in rapid paralysis followed by death within 12h. Subcutaneous inoculation of 15-30 micrograms of protein per gram body weight resulted in death of suckling mice in 2-3 h. The alkali-solubilized crystal delta-endotoxin was not toxic however, when administered per os. A comparison is made with a similar alkali-soluble fraction from the parasporal crystal delta-endotoxin of B. thuringiensis var kurstaki. With the exception of the Lepidopteran cell line, Choristoneura fumiferana 63 CF1, this soluble crystal delta-endotoxin protein showed no in vitro or in vivo toxicity, and no haemolytic activity.

Animals↗
PubMed2002

Deviation from additivity in mixture toxicity: relevance of nonlinear dose-response relationships and cell line differences in genotoxicity assays with combinations of chemical mutagens and gamma-radiation.

Sublinear dose-response relationships are often seen in toxicity testing, particularly with bioassays for carcinogenicity. This is the result of a superimposition of various effects that modulate and contribute to the process of cancer formation. Examples are saturation of detoxification pathways or DNA repair with increasing dose, or regenerative hyperplasia and indirect DNA damage as a consequence of high-dose cytotoxicity and cell death. The response to a combination treatment can appear to be supra-additive, although it is in fact dose-additive along a sublinear dose-response curve for the single agents. Because environmental exposure of humans is usually in a low-dose range and deviation from linearity is less likely at the low-dose end, combination effects should be tested at the lowest observable effect levels (LOEL) of the components. This principle has been applied to combinations of genotoxic agents in various cellular models. For statistical analysis, all experiments were analyzed for deviation from additivity with an n-factor analysis of variance with an interaction term, n being the number of components tested in combination. Benzo[a]pyrene, benz[a]anthracene, and dibenz[a,c]anthracene were tested at the LOEL, separately and in combination, for the induction of revertants in the Ames test, using Salmonella typhimurium TA100 and rat liver S9 fraction. Combined treatment produced no deviation from additivity. The induction of micronuclei in vitro was investigated with ionizing radiation from a 137Cs source and ethyl methanesulfonate. Mouse lymphoma L5178Y cells revealed a significant 40% supra-additive combination effect in an experiment based on three independent replicates for controls and single and combination treatments. On the other hand, two human lymphoblastoid cell lines (TK6 and WTK1) as well as a pilot study with human primary fibroblasts from fetal lung did not show deviation from additivity. Data derived from one cell line should therefore not be generalized. Regarding the testing of mixtures for deviation from additive toxicity, the suggested experimental protocol is easily followed by toxicologists.

Animals↗
PubMed2003

Overexpression of Nell-1, a craniosynostosis-associated gene, induces apoptosis in osteoblasts during craniofacial development.

UNLABELLED: We studied the cellular function of Nell-1, a craniosynostosis-related gene, in craniofacial development. Nell-1 modulates calvarial osteoblast differentiation and apoptosis pathways. Nell-1 overexpression disrupts these pathways resulting in craniofacial anomalies such as premature suture closure. INTRODUCTION: Craniosynostosis (CS), one of the most common congenital craniofacial deformities, is the premature closure of cranial sutures. Previously, we reported NELL-1 as a novel molecule overexpressed during premature cranial suture closure in patients with CS. Nell-1 overexpression induced calvarial overgrowth and resulted in premature suture closure in a rodent model. On a cellular level, Nell-1 is suggested to promote osteoblast differentiation. MATERIALS AND METHODS: Different levels of Nell-1 were introduced into osteoblastic cells by viral infection and recombinant protein. Apoptosis and gene expression assays were performed. Mice overexpressing Nell-1 were examined for apoptosis. RESULTS: In this report, we further showed that overexpression of Nell-1 induced apoptosis along with modulation of apoptosis-related genes. The induction of apoptosis by Nell-1 was observed only in osteoblastic cells and not in NIH3T3 or primary fibroblasts. The CS mouse model overexpressing Nell-1 showed increased levels of apoptosis in the calvaria. CONCLUSION: We show that Nell-1 expression modulates calvarial osteoblast differentiation and apoptosis pathways. Nell-1 overexpression disrupts these pathways resulting in craniofacial anomalies such as premature suture closure.

Animals↗