Background: Nuclear ribonucleoprotein particles play a key role in RNA processing and in the gene... more Background: Nuclear ribonucleoprotein particles play a key role in RNA processing and in the gene expression pathway. Interchromatin granules (GICs) involved in the metabolism of pre-messenger RNA (pre-mRNA) were described in Allium cepa and Chiranthodendron pentadactylon . Other particles as Lacandonia granules (LGs) were found in Lacandonia schismatica as well as Ginkgo biloba and Welwitschia mirabilis . LGs are structures equivalent to perichromatin granules (PCGs) described in mammals and to Balbiani ring granules (BRGs) described in the midge Chironomus tentans . PCGs and BRGs are involved in the metabolism of messenger RNA (mRNA). Here, we analyze the extranucleolar particles from Cycas revoluta and Ceratozamia mexicana and compare them to GICs and LGs using conventional electron microscopy and atomic force microscopy. Species study: Cycas revoluta (Cycadaceae) and Ceratozamia mexicana (Zamiaceae) Hypothesis: The extranucleolar ribonucleoprotein particles in the nuclei of C. r...
et al. Cementum protein 1 transfection does not lead to ultrastructural changes in nucleolar orga... more et al. Cementum protein 1 transfection does not lead to ultrastructural changes in nucleolar organization of human gingival fibroblasts.
Light and electron microscopy have been used to study cell structure for many years, but atomic f... more Light and electron microscopy have been used to study cell structure for many years, but atomic force microscopy is a more recent technique used to analyze cells, mainly due to the absence of techniques to prepare the samples. Isolated molecules or organelles, whole cells, and to a lesser extent in situ cell structure have been observed by different atomic force microscopy imaging modes. Here, we review efforts intended to analyze in situ the cell structures using approaches involving imaging of the surface of semithin sections of samples embedded in resin and sections prepared with an ultramicrotome. The results of such studies are discussed in relation to their implications to analyze the fine structure of organelles at the nanoscale in situ at enhanced resolution compared to light microscopy.
The nucleolus is the major site of ribosome biogenesis in eukaryotes. Until recently, G. Lamblia ... more The nucleolus is the major site of ribosome biogenesis in eukaryotes. Until recently, G. Lamblia was considered the only eukaryote lacking nucleoli. Recently, light and electron microscopy cytochemical techniques have been used to demonstrate the presence of nucleoli in the interphase nucleus of G. Lamblia. Here we review the work made during the last few years on the nucleolus of G. Lamblia in interphase and cell division. This mini review indicates that nucleoli are present in G. Lamblia interphase cell nuclei. Also, the persistence of nucleoli during cell division was documented more recently. Therefore, microscopical anatomy of G. Lamblia should include the presence of authentic morphological nucleoli.
The presence of RNA in the cell nucleus is well known. However, a high resolution in situ hybridi... more The presence of RNA in the cell nucleus is well known. However, a high resolution in situ hybridization evidence for the presence of RNA in some nuclear particles is still lacking. The aim of this work is to localize RNA in subnuclear particles using a novel ultrastructural in situ hybridization procedure. In this study, biotinylated genomic mouse DNA as a probe to localize total RNA in the nuclei of mouse hepatocytes was used. The procedure is based on paraformaldehyde fixation and embedding in lowicryl resin. Thin sections are mounted in formvar-coated gold grids. Hybridization is performed on non-denatured thin sections. DNA-RNA hybrids are detected with streptavidin-10 nm gold particles complex. By controlling the time of nick-translation during incorporation of biotin into the probe, labeling in the fibrillar portions of the nucleoplasm is obtained. More digested probes generate more labeling in the granular components. Nucleoli were similarly labeled. As expected, no label was...
Different studies in ovariectomized estrogen treated animals support the idea that c-fos plays a ... more Different studies in ovariectomized estrogen treated animals support the idea that c-fos plays a role in the proliferation of uterine epithelial cells. However, these studies invite us to reassess the role played by c-fos in epithelial cell types of the endometrium during the estrous cycle. The present study was undertaken to determine the c-fos and estrogen receptor (ER) gene expression pattern in the rat uterine epithelium during the estrous cycle in which natural and cyclic changes of steroid hormones occur, and correlate these changes with the proliferation status of this cellular types. Proliferation was assessed during the estrous cycle using bromodeoxyuridine incorporation to DNA. ERalpha and beta proteins were assessed by immunohistochemistry. The regulation of c-fos gene expression in the uterus of intact animals during the estrous cycle was evaluated using both in situ hybridization and immunohistochemistry. Estradiol (E(2)) and progesterone (P(4)) plasma levels were assessed by radioimmunoassay. The results indicated that luminal (LE) and glandular epithelia (GE) presented maximal proliferation during the metestrus (M) and the diestrus (D) days. However, during the proestrus (P) day only LE presented proliferation, and during the estrus (E) day only the stromal cells proliferated. A marked immunostaining for ERalpha was detected in both LE and GE cells during the early phases of the cycle but diminished on the P and the E day. In contrast, ERbeta was undetectable in both epithelia during all stages of the cycle. The highest c-fos mRNA level was detected in both epithelia on the M day, followed by a significant reduction during the other days of the cycle. The highest protein content was observed on the M and D days, and the minimal value was detected on the E day. The c-Fos protein level in LE was increased during M and D days, presenting a high correlation with the cellular proliferation pattern of this cell type. In conclusion, the overall results indicate that c-Fos protein presented a good correlation with uterine epithelial cell proliferation of LE. In the case of GE, the same tendency was observed, although no significant correlation was found. Both in LE and GE, c-fos mRNA did not strictly correlate with its protein levels. c-fos seems to have a postranscriptional regulation in uterine epithelial cells during the rat's estrous cycle.
We have investigated the distribution of U3 snRNA and rRNA in HeLa cells and normal rat kidney ce... more We have investigated the distribution of U3 snRNA and rRNA in HeLa cells and normal rat kidney cells during interphase and mitosis. U3 snRNA, known to be involved in pre-rRNA processing, was detected in nucleoli and coiled bodies during interphase, whereas rRNA was distributed in the nucleoli and throughout the cytoplasm. By comparison, ribosomal protein S6 was detected in nucleoli, coiled bodies, and in the cytoplasm. During nucleologenesis, pre-rRNA was observed in newly forming nucleoli during late telophase but not in prenucleolar bodies (PNBs), whereas U3 snRNA was detected in forming nucleoli and PNBs. Similar findings to those reported here for the localization of U3 snRNA have been reported previously for the U3 small nuclear ribonucleoprotein fibrillarin. These results suggest that components involved in pre-rRNA processing localize to discrete PNBs at the end of mitosis. The nucleolus is formed at specific telophase domains (nucleolar organizing regions) and the PNBs, cont...
... 14 Visualizing Nuclear Structure In Situ by Atomic Force Microscopy Luis Felipe Jiménez-Garcí... more ... 14 Visualizing Nuclear Structure In Situ by Atomic Force Microscopy Luis Felipe Jiménez-García and María de Lourdes Segura-Valdez ... Mariani, T., Musio, A., Frediani, C., Sbrana, I., and Ascoli, C. (1994) An atomic force microscope for cytological and histological investigations. ...
En los machos de algunas especies del genero Peltophryne, se ha descrito un organo vestigial fren... more En los machos de algunas especies del genero Peltophryne, se ha descrito un organo vestigial frente a sus testiculos llamado organo de Bidder. A este organo se le ha atribuido funcion endocrina y la capacidad de convertirse en un ovario funcional como estrategia reproductiva de las especies de este grupo. En el presente trabajo se describe la ultraestructura del organo de Bidder en los machos de siete especies endemicas de sapos de Cuba: Peltophryne fustiger, P. peltocephala, P. taladai, P. florentinoi, P. longinasa longinasa, P l. cajalbanensis, P. l. dunni, P. gundlachi, y P. catalauciceps. Las gonadas se fijaron, en paraformaldehido al 4% para aplicar histologia clasica y DAPI y en glutaraldehido 2,5% para ser procesadas para Microscopia Electronica de Transmision. Los resultados muestran que la gonada de todos los machos estudiados tiene un organo de Bidder por encima de los testiculos en posicion craneal. La morfologia del organo Bidder muestra algunas diferencias entre especie...
Although cell structure of Entamoeba histolytica is well known, the presence of nucleolar materia... more Although cell structure of Entamoeba histolytica is well known, the presence of nucleolar material has not been described with the electron microscope. Here we use light and electron microscopy cytochemical techniques to search for evidence1 of nucleolar material in this amoeba trophozoyte. Toluidine blue for RNA and silver staining for nucleolar organizer stain peripheral intranuclear material2. With the electron microscope, a3 similar material is fibro-granular and is contrasted with techniques for ribonucleoproteins and nucleolar organizer, but it is negative for DNA. These results show ultrastructural evidence for the presence of a peripheral and ring-shaped nucleolus4 in the nucleus of E. histolytica. It is suggested that this intranuclear organelle is a general feature in eukaryotes.
En eucariontes, el mRNA se forma a partir de un transcrito primario o pre-mRNA que madura mediant... more En eucariontes, el mRNA se forma a partir de un transcrito primario o pre-mRNA que madura mediante tres pasos que son la 7-metil-guanilacion del extremo 5', la poliadenilacion del extremo 3' y el splicing o corte de intrones y ligado de los exones resultantes. Estos pasos requieren de diversos factores cuya organizacion celular observada con el microscopio de epifluorescencia corresponde a una distribucion intranuclear conocida como patron moteado (speckles), que incluye regiones de distribucion concentrada (motas) y difusa en el nucleoplasma. La morfologia de este patron cambia en funcion de la actividad transcripcional y de splicing tanto en celulas en cultivo como en tejidos de mamiferos. En este trabajo utilizamos inmunofluorescencia indirecta con anticuerpos monoclonales contra la familia de factores de splicing SR y mostramos que este patron moteado tambien se presenta en celulas de tejidos de otros vertebrados como peces, anfibios, reptiles, aves y en otros cordados c...
Resumen Se describe la morfologia del testiculo y el epididimo del lagarto Anolis sagrei usando m... more Resumen Se describe la morfologia del testiculo y el epididimo del lagarto Anolis sagrei usando microscopia optica y electronica de transmision. La arquitectura del epitelio germinal del testiculo es similar a la de otros reptiles. Se encontraron en los tubulos seminiferos espermatogonias, espermatocitos, espermatidas en diversos grados y numerosos espermatozoides en una disposicion gradual desde la membrana basal hasta el lumen preludio de la organizacion radial de los amniotas. Se describen particularidades como la intensa pigmentacion de los testiculos y del epididimo y se atribuye su presencia a estrategias de proteccion. Todos los animales tenian una fuerte actividad espermatogenica. El espermatozoide en formacion presenta el nucleo con la cromatina filamentosa, un acrosoma con dobles membranas, ausencia de cuerpos densos y 10 mitoncodrias en la pieza media. En el epididimo es notable la altura de las celulas columnares y su actividad secretora. Palabras clave: Anolis sagrei , ...
The cell nucleus contains domains related to gene expression. The nucleolus is involved in pre-rR... more The cell nucleus contains domains related to gene expression. The nucleolus is involved in pre-rRNA metabolism while interchromatin granules are involved in pre-mRNA metabolism. It was recently shown that no ultrastructural changes are detected in the nucleolar organization, but no data of the effect of transfection in speckles are known. The aim of this work is to know if the interchromatin granule clusters display ultrastructural changes when introducing cement protein 1 (CEMP1) genes into human gingival fibroblasts (HGFs), a cell line of interest since they do not mineralize and upon transfection they perform mineralization, a potential to use in treatments of periodontal disease. Here we explored ultrastructural changes in interchromatin granule clusters after transfection of CEMP1. Non transfected and transfected cells display similar nuclear particles that form clumps in the interchromatin space and contain 10 15 nm in diameter individual granules intermingled with fibers, whi...
Background: Nuclear ribonucleoprotein particles play a key role in RNA processing and in the gene... more Background: Nuclear ribonucleoprotein particles play a key role in RNA processing and in the gene expression pathway. Interchromatin granules (GICs) involved in the metabolism of pre-messenger RNA (pre-mRNA) were described in Allium cepa and Chiranthodendron pentadactylon . Other particles as Lacandonia granules (LGs) were found in Lacandonia schismatica as well as Ginkgo biloba and Welwitschia mirabilis . LGs are structures equivalent to perichromatin granules (PCGs) described in mammals and to Balbiani ring granules (BRGs) described in the midge Chironomus tentans . PCGs and BRGs are involved in the metabolism of messenger RNA (mRNA). Here, we analyze the extranucleolar particles from Cycas revoluta and Ceratozamia mexicana and compare them to GICs and LGs using conventional electron microscopy and atomic force microscopy. Species study: Cycas revoluta (Cycadaceae) and Ceratozamia mexicana (Zamiaceae) Hypothesis: The extranucleolar ribonucleoprotein particles in the nuclei of C. r...
et al. Cementum protein 1 transfection does not lead to ultrastructural changes in nucleolar orga... more et al. Cementum protein 1 transfection does not lead to ultrastructural changes in nucleolar organization of human gingival fibroblasts.
Light and electron microscopy have been used to study cell structure for many years, but atomic f... more Light and electron microscopy have been used to study cell structure for many years, but atomic force microscopy is a more recent technique used to analyze cells, mainly due to the absence of techniques to prepare the samples. Isolated molecules or organelles, whole cells, and to a lesser extent in situ cell structure have been observed by different atomic force microscopy imaging modes. Here, we review efforts intended to analyze in situ the cell structures using approaches involving imaging of the surface of semithin sections of samples embedded in resin and sections prepared with an ultramicrotome. The results of such studies are discussed in relation to their implications to analyze the fine structure of organelles at the nanoscale in situ at enhanced resolution compared to light microscopy.
The nucleolus is the major site of ribosome biogenesis in eukaryotes. Until recently, G. Lamblia ... more The nucleolus is the major site of ribosome biogenesis in eukaryotes. Until recently, G. Lamblia was considered the only eukaryote lacking nucleoli. Recently, light and electron microscopy cytochemical techniques have been used to demonstrate the presence of nucleoli in the interphase nucleus of G. Lamblia. Here we review the work made during the last few years on the nucleolus of G. Lamblia in interphase and cell division. This mini review indicates that nucleoli are present in G. Lamblia interphase cell nuclei. Also, the persistence of nucleoli during cell division was documented more recently. Therefore, microscopical anatomy of G. Lamblia should include the presence of authentic morphological nucleoli.
The presence of RNA in the cell nucleus is well known. However, a high resolution in situ hybridi... more The presence of RNA in the cell nucleus is well known. However, a high resolution in situ hybridization evidence for the presence of RNA in some nuclear particles is still lacking. The aim of this work is to localize RNA in subnuclear particles using a novel ultrastructural in situ hybridization procedure. In this study, biotinylated genomic mouse DNA as a probe to localize total RNA in the nuclei of mouse hepatocytes was used. The procedure is based on paraformaldehyde fixation and embedding in lowicryl resin. Thin sections are mounted in formvar-coated gold grids. Hybridization is performed on non-denatured thin sections. DNA-RNA hybrids are detected with streptavidin-10 nm gold particles complex. By controlling the time of nick-translation during incorporation of biotin into the probe, labeling in the fibrillar portions of the nucleoplasm is obtained. More digested probes generate more labeling in the granular components. Nucleoli were similarly labeled. As expected, no label was...
Different studies in ovariectomized estrogen treated animals support the idea that c-fos plays a ... more Different studies in ovariectomized estrogen treated animals support the idea that c-fos plays a role in the proliferation of uterine epithelial cells. However, these studies invite us to reassess the role played by c-fos in epithelial cell types of the endometrium during the estrous cycle. The present study was undertaken to determine the c-fos and estrogen receptor (ER) gene expression pattern in the rat uterine epithelium during the estrous cycle in which natural and cyclic changes of steroid hormones occur, and correlate these changes with the proliferation status of this cellular types. Proliferation was assessed during the estrous cycle using bromodeoxyuridine incorporation to DNA. ERalpha and beta proteins were assessed by immunohistochemistry. The regulation of c-fos gene expression in the uterus of intact animals during the estrous cycle was evaluated using both in situ hybridization and immunohistochemistry. Estradiol (E(2)) and progesterone (P(4)) plasma levels were assessed by radioimmunoassay. The results indicated that luminal (LE) and glandular epithelia (GE) presented maximal proliferation during the metestrus (M) and the diestrus (D) days. However, during the proestrus (P) day only LE presented proliferation, and during the estrus (E) day only the stromal cells proliferated. A marked immunostaining for ERalpha was detected in both LE and GE cells during the early phases of the cycle but diminished on the P and the E day. In contrast, ERbeta was undetectable in both epithelia during all stages of the cycle. The highest c-fos mRNA level was detected in both epithelia on the M day, followed by a significant reduction during the other days of the cycle. The highest protein content was observed on the M and D days, and the minimal value was detected on the E day. The c-Fos protein level in LE was increased during M and D days, presenting a high correlation with the cellular proliferation pattern of this cell type. In conclusion, the overall results indicate that c-Fos protein presented a good correlation with uterine epithelial cell proliferation of LE. In the case of GE, the same tendency was observed, although no significant correlation was found. Both in LE and GE, c-fos mRNA did not strictly correlate with its protein levels. c-fos seems to have a postranscriptional regulation in uterine epithelial cells during the rat's estrous cycle.
We have investigated the distribution of U3 snRNA and rRNA in HeLa cells and normal rat kidney ce... more We have investigated the distribution of U3 snRNA and rRNA in HeLa cells and normal rat kidney cells during interphase and mitosis. U3 snRNA, known to be involved in pre-rRNA processing, was detected in nucleoli and coiled bodies during interphase, whereas rRNA was distributed in the nucleoli and throughout the cytoplasm. By comparison, ribosomal protein S6 was detected in nucleoli, coiled bodies, and in the cytoplasm. During nucleologenesis, pre-rRNA was observed in newly forming nucleoli during late telophase but not in prenucleolar bodies (PNBs), whereas U3 snRNA was detected in forming nucleoli and PNBs. Similar findings to those reported here for the localization of U3 snRNA have been reported previously for the U3 small nuclear ribonucleoprotein fibrillarin. These results suggest that components involved in pre-rRNA processing localize to discrete PNBs at the end of mitosis. The nucleolus is formed at specific telophase domains (nucleolar organizing regions) and the PNBs, cont...
... 14 Visualizing Nuclear Structure In Situ by Atomic Force Microscopy Luis Felipe Jiménez-Garcí... more ... 14 Visualizing Nuclear Structure In Situ by Atomic Force Microscopy Luis Felipe Jiménez-García and María de Lourdes Segura-Valdez ... Mariani, T., Musio, A., Frediani, C., Sbrana, I., and Ascoli, C. (1994) An atomic force microscope for cytological and histological investigations. ...
En los machos de algunas especies del genero Peltophryne, se ha descrito un organo vestigial fren... more En los machos de algunas especies del genero Peltophryne, se ha descrito un organo vestigial frente a sus testiculos llamado organo de Bidder. A este organo se le ha atribuido funcion endocrina y la capacidad de convertirse en un ovario funcional como estrategia reproductiva de las especies de este grupo. En el presente trabajo se describe la ultraestructura del organo de Bidder en los machos de siete especies endemicas de sapos de Cuba: Peltophryne fustiger, P. peltocephala, P. taladai, P. florentinoi, P. longinasa longinasa, P l. cajalbanensis, P. l. dunni, P. gundlachi, y P. catalauciceps. Las gonadas se fijaron, en paraformaldehido al 4% para aplicar histologia clasica y DAPI y en glutaraldehido 2,5% para ser procesadas para Microscopia Electronica de Transmision. Los resultados muestran que la gonada de todos los machos estudiados tiene un organo de Bidder por encima de los testiculos en posicion craneal. La morfologia del organo Bidder muestra algunas diferencias entre especie...
Although cell structure of Entamoeba histolytica is well known, the presence of nucleolar materia... more Although cell structure of Entamoeba histolytica is well known, the presence of nucleolar material has not been described with the electron microscope. Here we use light and electron microscopy cytochemical techniques to search for evidence1 of nucleolar material in this amoeba trophozoyte. Toluidine blue for RNA and silver staining for nucleolar organizer stain peripheral intranuclear material2. With the electron microscope, a3 similar material is fibro-granular and is contrasted with techniques for ribonucleoproteins and nucleolar organizer, but it is negative for DNA. These results show ultrastructural evidence for the presence of a peripheral and ring-shaped nucleolus4 in the nucleus of E. histolytica. It is suggested that this intranuclear organelle is a general feature in eukaryotes.
En eucariontes, el mRNA se forma a partir de un transcrito primario o pre-mRNA que madura mediant... more En eucariontes, el mRNA se forma a partir de un transcrito primario o pre-mRNA que madura mediante tres pasos que son la 7-metil-guanilacion del extremo 5', la poliadenilacion del extremo 3' y el splicing o corte de intrones y ligado de los exones resultantes. Estos pasos requieren de diversos factores cuya organizacion celular observada con el microscopio de epifluorescencia corresponde a una distribucion intranuclear conocida como patron moteado (speckles), que incluye regiones de distribucion concentrada (motas) y difusa en el nucleoplasma. La morfologia de este patron cambia en funcion de la actividad transcripcional y de splicing tanto en celulas en cultivo como en tejidos de mamiferos. En este trabajo utilizamos inmunofluorescencia indirecta con anticuerpos monoclonales contra la familia de factores de splicing SR y mostramos que este patron moteado tambien se presenta en celulas de tejidos de otros vertebrados como peces, anfibios, reptiles, aves y en otros cordados c...
Resumen Se describe la morfologia del testiculo y el epididimo del lagarto Anolis sagrei usando m... more Resumen Se describe la morfologia del testiculo y el epididimo del lagarto Anolis sagrei usando microscopia optica y electronica de transmision. La arquitectura del epitelio germinal del testiculo es similar a la de otros reptiles. Se encontraron en los tubulos seminiferos espermatogonias, espermatocitos, espermatidas en diversos grados y numerosos espermatozoides en una disposicion gradual desde la membrana basal hasta el lumen preludio de la organizacion radial de los amniotas. Se describen particularidades como la intensa pigmentacion de los testiculos y del epididimo y se atribuye su presencia a estrategias de proteccion. Todos los animales tenian una fuerte actividad espermatogenica. El espermatozoide en formacion presenta el nucleo con la cromatina filamentosa, un acrosoma con dobles membranas, ausencia de cuerpos densos y 10 mitoncodrias en la pieza media. En el epididimo es notable la altura de las celulas columnares y su actividad secretora. Palabras clave: Anolis sagrei , ...
The cell nucleus contains domains related to gene expression. The nucleolus is involved in pre-rR... more The cell nucleus contains domains related to gene expression. The nucleolus is involved in pre-rRNA metabolism while interchromatin granules are involved in pre-mRNA metabolism. It was recently shown that no ultrastructural changes are detected in the nucleolar organization, but no data of the effect of transfection in speckles are known. The aim of this work is to know if the interchromatin granule clusters display ultrastructural changes when introducing cement protein 1 (CEMP1) genes into human gingival fibroblasts (HGFs), a cell line of interest since they do not mineralize and upon transfection they perform mineralization, a potential to use in treatments of periodontal disease. Here we explored ultrastructural changes in interchromatin granule clusters after transfection of CEMP1. Non transfected and transfected cells display similar nuclear particles that form clumps in the interchromatin space and contain 10 15 nm in diameter individual granules intermingled with fibers, whi...
Uploads
Papers by Lourdes Segura