Cementum protein 1 transfection does not lead to ultrastructural changes in nucleolar organization of human gingival fibroblasts.

BACKGROUND AND OBJECTIVE: Transfection of cementum protein 1 (CEMP1) into human gingival fibroblasts (HGFs) notably increases cell metabolism and results in overexpression of molecules related to biomineralization at transcriptional and protein levels. Therefore, HGF-CEMP1 cells are considered as putative cementoblasts. This represents a significant advance in periodontal research because cementum neoformation is a key event in periodontal regeneration. In addition, it is well known that important changes in cell metabolism and protein expression are related to nucleolar structure and the function of this organelle, which is implicated in ribosome biogenesis. The aim of this study was to determine the effect of transfecting CEMP1 gene in human HGF on the ultrastructure of the nucleolus. MATERIAL AND METHODS: Cells were processed using the conventional technique for transmission electron microscopy, fixed with glutaraldehyde, postfixed with osmium tetraoxide, and embedded in epoxy resin. Semi-thin sections were stained with Toluidine blue and observed by light microscopy. Thin sections were stained with uranyl acetate and lead citrate. For ribonucleoprotein detection, the staining method based on the regressive effect of EDTA was used. In addition, the osmium ammine technique was used for specific staining of DNA. RESULTS: The results obtained in this study suggest that transfection of CEMP1 into HGFs does not produce changes in the general nucleolar ultrastructure because the different components of the organelle are present as fibrillary centers, and dense fibrillar and granular components compared with the control. CONCLUSION: The transfection of CEMP1 into HGFs allows these cells to perform cementoblast-like functions without alteration of the ultrastructure of the nucleolus, evaluated by the presence of the different compartments of this organelle involved in ribosomal biogenesis.

FGF receptor genes and breast cancer susceptibility: results from the Breast Cancer Association Consortium.

BACKGROUND: Breast cancer is one of the most common malignancies in women. Genome-wide association studies have identified FGFR2 as a breast cancer susceptibility gene. Common variation in other fibroblast growth factor (FGF) receptors might also modify risk. We tested this hypothesis by studying genotyped single-nucleotide polymorphisms (SNPs) and imputed SNPs in FGFR1, FGFR3, FGFR4 and FGFRL1 in the Breast Cancer Association Consortium. METHODS: Data were combined from 49 studies, including 53 835 cases and 50 156 controls, of which 89 050 (46 450 cases and 42 600 controls) were of European ancestry, 12 893 (6269 cases and 6624 controls) of Asian and 2048 (1116 cases and 932 controls) of African ancestry. Associations with risk of breast cancer, overall and by disease sub-type, were assessed using unconditional logistic regression. RESULTS: Little evidence of association with breast cancer risk was observed for SNPs in the FGF receptor genes. The strongest evidence in European women was for rs743682 in FGFR3; the estimated per-allele odds ratio was 1.05 (95% confidence interval=1.02-1.09, P=0.0020), which is substantially lower than that observed for SNPs in FGFR2. CONCLUSION: Our results suggest that common variants in the other FGF receptors are not associated with risk of breast cancer to the degree observed for FGFR2.

Common variations in ERCC2 are associated with response to cisplatin chemotherapy and clinical outcome in osteosarcoma patients.

Platinum agents cause DNA cross-linking. Nucleotide excision repair genes play a key role in DNA damage repair. This study aims to investigate whether polymorphisms in these genes are associated with tumor response and survival in cisplatin-treated osteosarcoma patients. Eight single nucleotide polymorphisms in ERCC2, XPC, XPA, ERCC1, ERCC4 and ERCC5 genes were analyzed in 91 patients diagnosed with osteosarcoma and treated with cisplatin. A significant association with tumor response, after correction for multiple testing, was found for the Lys751Gln polymorphism in the ERCC2 gene. We found that only 45% of patients with at least one polymorphic G allele responded compared with 80% of patients homozygous for the common T allele (odds ratio=4.9, 95% confidence interval=1.64-14.54, adjusted P-value=0.047). In addition, carrying at least one ERCC2 Lys751GlnG allele was significantly associated with shorter event-free survival (median=184 months, compared with 240 months for TT homozygotes; hazard ratio=5.76, 95% confidence interval=1.30-25.55; P-value=0.021). Although ototoxicity was only recorded in 32 patients, we found weak evidence of an association with the CC genotype of XPC Lys939Gln (P-value= 0.042). This is the first pharmacogenetic study focused on osteosarcoma treatment providing evidence that polymorphic variants in DNA repair genes could be useful predictors of response to cisplatin chemotherapy in osteosarcoma patients.

New evidence that Lacandonia granules are ultrastructurally related to perichromatin and Balbiani ring granules.

Lacandonia granules are abundant non-typical extranucleolar ribonucleoprotein particles found in the nucleus of Lacandonia schismatica, a rare plant showing spatial inversion of sex organs. In the present study, changes in the number of Lacandonia granules during flower development, and the presence of SR proteins and poly(A)+ RNA in the nuclei of L. schismatica were analyzed by electron microscopy, immunoelectron microscopy and ultrastructural in situ hybridization. Our results show an important reduction in the number of Lacandonia granules in the nuclei of cells of opened (post-anthesis) in relation to unopened (pre-anthesis) flowers, where granules are very abundant. The SR family of splicing factors and poly(A)+ RNA are present in both perichromatin fibers and Lacandonia granules. The developmental behavior, the presence of SR proteins, recently involved in post-splicing events, poly(A)+ RNA and the reported absence of snRNPs splicing factors in Lacandonia granules, suggest that these particles are involved in postranscriptional events as storage and/or transport of mRNAs. A similar situation is present in other nuclear RNP as perichromatin granules present in mammals and Balbiani ring granules of salivary glands of Chironomus. Based on similarities in morphological, developmental behavior, immunocytochemistry and in situ hybridization results, we conclude that Lacandonia, perichromatin and Balbiani ring granules may be also functionally similar structures.

Further ultrastructural characterization of the intranuclear ring-shaped bodies of the plant Lacandonia schismatica.

Ring-shaped bodies are found in the nucleus of Lacandonia schismatica, a rare plant with the sexual organs inverted. They are 0.5-microm-diameter structures that present an electron-dense external ring surrounding a central core. Ultrastructural studies indicate that these bodies contain RNA. The external ring is labeled with antibodies against small nuclear ribonucleoproteins, suggesting that they may be involved in pre-mRNA metabolism. In the present work we further characterized these intranuclear ring-shaped structures by serial-sectioning analysis. Moreover, we tested the presence of additional molecular elements related to pre-mRNA metabolism, such as SR proteins and poly(A)(+) RNA, using immunoelectron microscopy and ultrastructural in situ hybridization. Our results show that these nuclear bodies are spherical. They contain SR proteins involved in splicing and postsplicing events and little to no poly(A)(+) RNA. We also found similar nuclear bodies in other plant and animal species. Therefore, ring-shaped bodies in L. schismatica are spherical, highly compartmentalized nuclear structures that may be involved in pre-mRNA metabolism.