The application of an in situ karyotyping technique for mesenchymal stromal cells: a validation and comparison study with classical G-banding

The cytogenetic analysis of mesenchymal stromal cells (MSCs) is essential for verifying the safety and stability of MSCs. An in situ technique, which uses cells grown on coverslips for karyotyping and minimizes cell manipulation, is the standard protocol for the chromosome analysis of amniotic fluids. Therefore, we applied the in situ karyotyping technique in MSCs and compared the quality of metaphases and karyotyping results with classical G-banding and chromosomal abnormalities with fluorescence in situ hybridization (FISH). Human adipose- and umbilical cord-derived MSC cell lines (American Type Culture Collection PCS-500-011, PCS-500-010) were used for evaluation. The quality of metaphases was assessed by analyzing the chromosome numbers in each metaphase, the overlaps of chromosomes and the mean length of chromosome 1. FISH was performed in the interphase nuclei of MSCs for 6q, 7q and 17q abnormalities and for the enumeration of chromosomes via oligo-FISH in adipose-derived MSCs. The number of chromosomes in each metaphase was more variable in classical G-banding. The overlap of chromosomes and the mean length of chromosome 1 as observed via in situ karyotyping were comparable to those of classical G-banding (P=0.218 and 0.674, respectively). Classical G-banding and in situ karyotyping by two personnel showed normal karyotypes for both cell lines in five passages. No numerical or structural chromosomal abnormalities were found by the interphase-FISH. In situ karyotyping showed equivalent karyotype results, and the quality of the metaphases was not inferior to classical G-banding. Thus, in situ karyotyping with minimized cell manipulation and the use of less cells would be useful for karyotyping MSCs.

Utilidad del bandeo cromosómico con la enzima ALU I para la identificación de zonas metiladas en cáncer de mama / Utility of chromosome banding with ALU I enzyme for identifying methylated areas in breast cancer

El cáncer es un conjunto de trastornos que comparten la característica común de un crecimiento celular descontrolado, teniendo la facultad de comenzar en las células, generando dos procesos sucesivos: el aumento de la proliferación celular (tumor o neoplasia) y la capacidad invasiva de estas células, proliferando y colonizando otros tejidos (metástasis). La metilación del DNA es un proceso epigenético que recurrentemente ha sido involucrado como un factor importante en la patogenia de esta enfermedad el cual participa en la regulación de la expresión génica directamente al impedir la unión de factores de trascripción, e indirectamente propiciando la estructura “cerrada” de la cromatina. El objetivo de este trabajo fue determinar regiones hipermetiladas en muestras de extendidos cromosómicos mediante la utilización de la endonucleasa de restricción Alu I y relacionar estas regiones con sitios de localización de genes supresores de tumores relacionados con el cáncer de mama. Se analizaron 60 muestras de sangre periférica de mujeres con diagnóstico de cáncer de mama a las cuales se les realizó cultivo celular; los extendidos cromosómicos fueron teñidos con Giemsa previamente digeridos con la enzima Alu I. Se observaron cromosomas con regiones centroméricas y no centroméricas teñidas en el 37% de los casos, comprobándose que en el 95,46% de los casos existen genes asociados descritos, como metilados en cáncer de mama. Ejemplo de ellos son los localizados en los cromosomas 1q, 2q, 6q, y regiones centroméricas no teñidas usualmente como en los cromosomas 3, 4, 8, 13, 14, 15, y 17. Se sugiere la importancia de esta técnica ya que permite la visualización total del genoma, pudiendo localizar genes metilados relacionados con cáncer de mama y, de esta manera dirigir la terapia de forma específica, logrando una mejor respuesta terapéutica.

Cancer is a group of disorders characterized by uncontrolled cell growth which is produced by two successive events: increased cell proliferation (tumor or neoplasia) and the invasive capacity of these cells (metastasis). DNA methylation is an epigenetic process which has been involved as an important pathogenic factor of cancer. DNA methylation participates in the regulation of gene expression, directly, by preventing the union of transcription factors, and indirectly, by promoting the “closed” structure of the chromatine. The objectives of this study were to identify hypermethyled chromosomal regions through the use of restriction Alu I endonuclease, and to relate cytogenetically these regions with tumor suppressive gene loci. Sixty peripheral blood samples of females with breast cancer were analyzed. Cell cultures were performed and cytogenetic spreads, previously digested with Alu I enzyme, were stained with Giemsa. Chromosomal centromeric and not centromeric regions were stained in 37% of cases. About 96% of stained hypermethyled chromosomal regions (1q, 2q, 6q) were linked with methylated genes associated with breast cancer. In addition, centromeric regions in chromosomes 3, 4, 8, 13, 14, 15 and 17, usually unstained, were found positive to digestion with Alu I enzime and Giemsa staining. We suggest the importance of this technique for the global visualization of the genome which can find methylated genes related to breast cancer, and thus lead to a specific therapy, and therefore a better therapeutic response.

Testes genéticos em diagnóstico pré-natal: onde estamos, para onde vamos / Prenatal genetic testing: where we are at, where we are heading to

Este texto tem como objetivo apresentar uma revisão acerca do estado da arte da citogenética convencional e molecular aplicada ao diagnóstico pré-natal, discutindo as aplicações, vantagens e desvantagens dos diferentes métodos, em suas bases teóricas e históricas. Desde 1960, a citogenética convencional, com a análise microscópica dos cromossomos em divisão, vem sendo utilizada como padrão ouro. Entretanto, mesmo adotando essa abordagem, para uma significativa parcela de casos não é possível estabelecer diagnóstico sindrômico definitivo em cerca de metade dos pacientes que apresentam cariótipo normal, na presença de malformações. Para esse grupo, as técnicas moleculares que envolvem estudos em nível genômico poderiam permitir a identificação de novos microarranjos cromossômicos possivelmente responsáveis pelo fenótipo anormal, contribuindo para a caracterização molecular e estabelecimento de um diagnóstico mais preciso, uma abordagem perinatal mais adequada e um aconselhamento genético mais detalhado. Destaca-se o advento das técnicas de FISH, SKY, CGH e array CGH como promissoras aliadas, de forma complementar ao cariótipo convencional

This paper aims at presenting a review of the state of the art of conventional and molecular cytogenetics applied to prenatal diagnosis, the applications, pros and cons of different techniques and their historical and theoretical background. Since 1960, conventional cytogenetics, based on the analysis of chromosomes has been used as a gold standard. However, for a significant proportion of cases it is not possible to establish definitive syndromic diagnosis in about half of the patients with normal karyotype in the presence of malformations. For this group, molecular techniques at the genomic level might allow the identification of new chromosomal areas potentially responsible for the abnormal phenotype, contributing to the molecular characterization and establishment of a more accurate diagnosis and the most appropriate perinatal approach, including a more detailed genetic counseling. The advent of FISH techniques, SKY, CGH and array CGH will be discussed as promising tools to complement cytogenetic diagnosis based on conventional karyotyping

Importância do estudo genético pré-natal / Importance of prenatal genetic diagnosis

O desenvolvimento de técnicas, como o cariótipo e ensaios enzimáticos em células fetais, a determinação de metabólitos no líquido amniótico e a ultrassonografia, propiciaram o diagnóstico pré-natal de desordens genéticas. A investigação genética pré-natal permite a detecção, ainda no útero, de doenças que, de outra forma, somente seriam diagnosticadas após o nascimento. Diversas técnicas são utilizadas para avaliação do estado fetal, algumas como a biópsia de vilos coriais, a amniocentese e a cordocentese. Com desenvolvimento tecnológico, novas técnicas moleculares foram desenvolvidas apresentando-se de forma mais refinada e de rápido resultado. A utilização dessas técnicas é fundamental para o desenvolvimento fetal, podendo então indicar uma conduta adequada para cada caso. Dessa forma, o conhecimento e a aplicação da genética clínica, utilizando o aconselhamento genético, trazem a certeza de um bom acompanhamento pré-natal necessário à assistência médica.

The development of techniques, such as karyotype and enzymatic assay in fetal cells, the determination of metabolites in amniotic liquid and the ultrasonography, allowed prenatal diagnosis of genetic disorders. The prenatal genetic investigation allowed the detection, in the womb, of diseases that, in other way, just could be diagnosed after birth. Many techniques are used to fetal state assessment, some of them such as villi cori biopsy, the amniocentesis and cordocentesis. Through the technological development, new molecular techniques were developed. They present a more refined and fast results. The use of these techniques is fundamental to fetal development, enabling the use of adequate conduct in each case. In this way, the knowledge and application of genetic clinic, using genetic counseling, bring the certainty of a good prenatal care, which is necessary to medic assistance.

Chromosomal abnormalities & oxidative stress in women with premature ovarian failure (POF).

Background & objectives: Premature ovarian failure (POF) is defined as the cessation of ovarian function under the age of 40 yr and is characterized by amenorrhoea, hypoestrogenism and elevated serum gonadotrophin levels. The cause of POF remains undetermined in majority of the cases. This study was aimed to investigate the type and frequency of cytogenetic abnormalities in patients with idiopathic POF and also to study the role of oxidative stress in such cases. Methods: Seventy five women with idiopathic POF were included in this study. Chromosome analysis was done in peripheral blood lymphocytes by conventional GTG banding to identify numerical or structural abnormalities. Cytogenetically normal cases were investigated for reactive oxygen species (ROS) levels in their blood by luminol-chemiluminescence assay. Results: Eighteen chromosomal anomalies were identified in POF patients (24%). Majority of the cases were found to have X-chromosome abnormalities (28%). Overall median ROS range was found to be significantly higher (P<0.01) in POF patients [50480 (120,132966) RLU/min] compared to controls [340 (120,5094) RLU/min]. Among these, 50 per cent of the POF patients had higher ROS levels, 20 per cent had medium elevation and 30 per cent were found to have normal values comparable to controls. Interpretation & conclusions: X-chromosome anomalies were found to be the major contributor of POF. Oxidative stress may be the underlying aetiology in idiopathic premature ovarian failure. Thus the results of this study highlight the role of cytogenetic abnormalities and supraphysiological levels of ROS in causation of idiopathic POF. But the role of oxidative stress needs to be confirmed by other studies on patients from different geographical areas and from different ethnicities.

Cytogenetic analysis of the 18S, 5S rDNA and B chromosome of Iheringichthys labrosus (Lütken, 1874) (Siluriformes, Pimelodidae) / Análise Citogenética de DNAr 18S, 5S e cromossomo B de Iheringichthys labrosus (Lütken, 1874) (Siluriformes, Pimelodidae)

Cytogenetic analyses of the location of 18S and 5S ribosomal DNAs, and the base composition of B chromosomes of Iheringichthys labrosus from Tibagi River, Paraná, Brazil, are provided. AgNORs were observed in the terminal position on the long arm of a subtelocentric chromosome pair. CMA3-positive staining was observed in some chromosomes, which besides being associated with NORs, were all DAPI-negative. Chromosome B showed a strong fluorescence with CMA3. The concomitant use of 18S and 5S rDNA probes using the FISH technique revealed 18S ribosomal cistrons in a pair of subtelocentric chromosomes, on the long arm in the terminal position, coinciding with the AgNOR. The 5S sites were found in another subtelocentric pair, on the long arm in the interstitial region, near the centromere. The findings of the present study suggest that, although there are some more conserved cytogenetic characteristics, populations of I. labrosus may show their own characteristics.

Foram realizadas análises citogenéticas em Iheringichthys labrosus do Rio Tibagi, Paraná, Brasil com a localização cromossômica dos DNAs ribossômicos 18S e 5S e a composição de bases de seus cromossomos B. As AgNORs foram observadas em posição terminal, no braço longo de um par de cromossomos subtelocêntricos. Marcações CMA3 positivas foram observadas em alguns cromossomos e associadas com as RONs. Porém, todas estas marcações apresentaram-se DAPI negativas. O cromossomo B mostrou-se fortemente fluorescente com CMA3. O uso concomitante das sondas de DNAr 18S e 5S, através da técnica de FISH, revelou os cístrons ribossômicos em um par de cromossomos subtelocêntricos, em posição terminal do braço longo, coincidindo com a AgNOR. Os sítios 5S foram observados em outro par subtelocêntrico, em posição intersticial do braço longo, próximo ao centrômero. Os resultados observados no presente estudo sugerem que, embora existam algumas características citogenéticas mais conservadas, as populações de I. labrosus podem mostrar suas próprias características.

Chromosomal abnormalities associated with mental retardation in female subjects.

Chromosomal abnormalities are thought to be the most common cause of mental retardation (MR). However, apart from a few selected types with typical aneuploidy, like Downs syndrome, Klinefelter syndrome, Turner syndrome, etc., the frequency of detectable chromosomal abnormalities in association with idiopathic MR is very low. In this study, we have investigated chromosomal abnormalities in female MR subjects (n = 150) by high-resolution GTG banding. Of them, 30 cases were diagnosed as Downs syndrome. Among the remaining (n = 120), chromosomal abnormalities/marked polymorphisms were detectable in only three MR cases (0.025).

Comparative cytogenetic study in Muscidae flies

The chromosome modal number in Muscoidea Diptera is 2n = 12, including five pairs of autosomes and one sex chromosome pair. Nevertheless, some species with 2n = 10 chromosomes have been described, all of them from the Muscidae family. We analyzed the karyotype of some Muscidae species from different subfamilies and compared the obtained data with the karyotypes of some species of the families Calliphoridae and Sarcophagidae. Comparisons of these species with other Muscidae species revealed a considerable variation among their sex chromosomes. This variation in the length of the sex chromosomes suggests that parts of these chromosomes were lost or fused with autosomes. The constitutive heterochromatic regions and the nucleolar organizer regions (NORs) were also analyzed and some aspects about the relationship between these regions and the sex chromosomes are discussed.

O número modal de cromossomos dos Dípteros Muscóideos é 2n = 12, incluindo cinco pares de autossomos e um par de cromossomos sexuais. No entanto, algumas espécies com 2n = 10 cromossomos já foram descritas, sendo todas pertencentes à família Muscidae. No presente trabalho, foram analisados os cariótipos de algumas espécies de Muscidae de diferentes subfamílias e os dados obtidos foram comparados com os cariótipos de algumas espécies das famílias Calliphoridae e Sarcophagidae. Comparações destas espécies com outras da família Muscidae revelaram uma considerável variação entre seus cromossomos sexuais. Esta variação no tamanho dos cromossomos sexuais sugere que parte destes cromossomos foram perdidos ou sofreram fusão com autossomos. As regiões de heterocromatina constitutiva e as regiões organizadoras de nucléolos (RONs) foram também analisadas e alguns aspectos sobre a relação destas com os cromossomos sexuais são discutidos.

Chromosome polymorphism and complements in populations of Girardia species (Platyhelminthes, Tricladida, Paludicola) from southern Brazil

The karyotypes of four species of freshwater triclads of the genus Girardia (Platyhelminthes), i.e. G. schubarti, G. tigrina, G. anderlani, and G. biapertura, from populations of different localities of the Rio Grande do Sul State, in southern Brazil, were analyzed. The karyotype of G. biapertura is presented for the first time. Three basic complements of 4, 8, and 9 chromosomes were found. Diploids, triploids, or mixoploids (2n/3n) specimens were frequently detected in these populations. The basic chromosomal complement of n = 9 was verified in two different species (G. biapertura and G. anderlani), presenting a large acrocentric chromosome which is rare in the family Dugesiidae. An intra and interspecific chromosomal variability was also detected and its evolutionary implications are discussed.

Os cariótipos de quatro espécies de planárias de água doce do gênero Girardia (Platyhelminthes), a saber, G. schubarti, G. tigrina, G. anderlani e G. biapertura, de populações ocorrentes em diferentes locais do estado do Rio Grande do Sul, na região sul do Brasil, foram analisados. O cariótipo de G. biapertura é apresentado pela primeira vez. Foram observados três complementos básicos, de 4, 8 e 9 cromossomos. Espécimes diplóides, triplóides e mixoplóides (2n/3n) foram observados freqüentemente nessas populações. O complemento cromossômico básico de n = 9 foi verificado em duas espécies (G. biapertura e G. anderlani), apresentando um grande cromossomo acrocêntrico que é raro na família Dugesiidae. Também foi observada certa variabilidade cromossômica, tanto intra- como interespecífica, cujas implicações evolutivas são discutidas.

Evaluation of hot saline solution and restriction endonuclease techniques in cytogenetic studies of Cycloneda sanguinea L. (Coccinellidae)

The goal of the present study was to determine if simple methods, especially hot saline solution (HSS) and MspI and HaeIII restriction endonucleases, which do not require special equipments, may be helpful in studies of genetic variability in the lady beetle, Cycloneda sanguinea. The HSS method extracted the heterochromatin region, suggesting that it is composed mostly of DNA rich in A-T base pairs. However, the X and y chromosomes were resistant to HSS banding. These bands facilitated the identification of each chromosome. In this study, we used the restriction endonucleases with different G-C base target sequences: MspI C/GGC and HaeIII GG/CC. The use of restriction enzyme MspI did not show an effect on the autosomal chromosomes. On the other hand, the sex pair showed a pale staining, to help in the recognition of these chromosomes. HaeIII produced characteristic bands which were identified all along the chromosomes, facilitating the identification of each chromosome. Based on these results, we can consider the heterochromatin being heterogeneous. The findings obtained here, using different chromosomal banding techniques, may be useful in the identification of intraspecific chomosome variability, specifically in Coccinellidae (Coleoptera) chromosomes, even without special equipment.

Composition of constitutive heterochromatin of Pseudonannolene strinatii Mauriès, 1974 (Diplopoda, Spirostreptida) analyzed by AT/CG specific fluorochromes

Triple staining with fluorochromes (DA/DAPI/CMA) and C-banding were used to characterize the composition of Pseudonannolene strinatii heterochromatin. C-banding showed C+ bands of different labeling intensity on chromosomes 1 and 2 in some cells. Fluorochrome staining revealed DAPI+ regions corresponding to the C-banding pattern, indicating that the heterochromatin of this species is abundant in AT-rich sequences.

First chromosome data on Steindachneridion scripta (Pisces, Siluriformes, Pimelodidae) from Brazilian rivers: Giemsa, CBG, G-, and RE banding

A cytogenetic study was performed on the large pimelodid species Steindachneridion scripta (Siluriformes, Sorubiminae) from the Paraná River basin (Brazil). Chromosome preparations were obtained avoiding sacrifice of the specimens, by means of lymphocyte culture, and several staining and chromosome banding techniques were applied. The karyotype consisted of 56 chromosomes, 24 metacentrics, 20 submetacentrics, 4 subtelocentrics, and 8 acrocentrics (fundamental number = 104). The first pair of acrocentric chromosomes (pair 25) consistently had a decondensed secondary constriction; the C-banding pattern of some chromosomes allows them to be considered cytogenetic markers (i.e., pairs 1, 3, 4, 6, 7, 9, 13, 23, and 24). G-banding and restriction enzymes provided patterns that helped improve chromosome pairing. This is the first report on a Neotropical pimelodid species of economic interest using several cytogenetic techniques and providing an integral karyotypic characterization.

Heterochromatin and chromosome evolution: a FISH probe of Cebus apella paraguayanus (Primate: Platyrrhini) developed by chromosome microdissection

Neotropical Primate karyotypes are highly variable, particularly in the heterochromatic regions, not only regarding the amount of heterochromatin, but also the composition. G and C banding and FISH techniques provide useful information to characterize interspecific relationships. We used chromosome microdissection to develop a FISH probe of the chromosome 11 heterochromatic block (11qHe+) of Cebus apella paraguayanus (CAPp). Fragments of the 11qHe+ microdissected from fibroblast cell culture were collected in a PCR tube, amplified by degenerate oligonucleotide primer-PCR and subsequently labeled. The specificity of the FISH probe was confirmed in metaphases of some Ceboidea species. Signals were located in the He+ of chromosomes 4, 11, 12, 13, and 19 of CAPp and in the He+ of chromosomes 4, 12 and 13 of C. a. nigritus (CAPn); no signals were observed when other Ceboidea species were analyzed. We propose that the heterochromatin observed in CAPp and CAPn is specific for these species. We consider this C. apella heterochromatin identity as a possible key for the interpretation of chromosomal evolution in these Ceboidea.

Karyotypic characterization of Iheringichthys labrosus (Pisces, Pimelodidae): C-, G- and restriction endonuclease banding

Various chromosomal banding techniques were utilized on the catfish, Iheringichthys labrosus, taken from the Capivara Reservoir. C-banding regions were evidenced in telomeric regions of most of the chromosomes. The B microchromosome appeared totally heterochromatic. The restriction endonuclease AluI produced a banding pattern similar to C-banding in some chromosomes; the B microchromosome, when present, was not digested by this enzyme and remained stained. G-banding was conspicuous in almost all the chromosomes, with the centromeres showing negative G-banding. When the restriction endonuclease BamHI was used, most of the telomeres remained intact, while some centromeres were weakly digested. The B chromosome was also not digested by this enzyme. The first pair of chromosomes showed a pattern of longitudinal bands, both with G-banding and BamHI; this was more evident with G-banding. This banding pattern can be considered a chromosomal marker for this population of I. labrosus.

Cytogenetics of genetic counseling patients in Pelotas, Rio Grande do Sul, Brazil

From 1986 to 2002, we examined the chromosomal composition of 916 patients attended by two genetic counseling services in the city of Pelotas, in the Brazilian State of Rio Grande do Sul, to determine the genetic causes of their disturbances. Patterns of G-banding using trypsin and Giemsa (GTG) and C-banding using barium and Giemsa (CBG) were studied using phytohemagglutinin M-stimulated lymphocytes cultured from peripheral blood. Among the patients, 110 had Down's syndrome, 7 had Edward's syndrome, 4 had Patau's syndrome, 29 had Turner's syndrome, 5 had Klinefelter's syndrome, and 3 had [quot ]cri-du-chat[quot ] syndrome. Abnormal chromosomes were observed in 29.3% of the patients. Most of these (56.3%) were numerical abnormalities, with the remaining being structural variants.

Una saga citogenética: El descubrimiento de los métodos de bandeo cromosómico. Significado y proyección bio-médica / A cytogenetic saga: The discovery of chromosome banding methods. Bio-medical significance and projection

Se exponen, desde una perspectiva histórica y en forma sucinta, los sistemas de identificación de los cromosomas humanos y varios métodos de bandeo cromosómico desarrollados en las últimas cuatro décadas. Se relatan ámbitos y hechos acontecidos -varios de los cuales protagonizó el autor- que originaron la actual moderna citogenética estructural y molecular. Se mencionan, asimismo, algunos aspectos moleculares implícitos en estas metodologías con el propósito de contribuir al esclarecimiento de los variados aspectos revelados por dichos métodos sobre las estructuras cromosómicas subyacentes. Se describen también los procedimientos analíticos desarrollados en nuestro laboratorio para el estudio de núcleos y cromosomas como así los resultados obtenidos sobre el análisis cuantitativo de la región subtelomérica y su probable relación con las aberraciones crípticas, detectadas en dicho segmento cromosómico, productoras de retardo mental y malformaciones congénitas. Finalmente, se realiza un breve relato sobre las aplicaciones del bandeo cromosómico en el proyecto del genoma humano y su posible utilidad en los experimentos de síntesis artificial de núcleos celulares eucarióticos, de particular significado en biología.

Summary Human chromosome identification systems and several chromosome banding methods, developed during the last four decades, are briefly reviewed under a historical perspective. Numerous episodes that built the basis of the present modern structural and molecular citogenetics (the author took part in some of them) are reported. Some molecular aspects related to this methodology are also mentioned, in order to contribute to the understanding of different aspects of the underlying chromosomal structures revealed by these methods. The analytical procedures developed in our laboratory for the study of nuclei and chromosomes, the results of quantitative analysis of the subtelomeric region and its probable relationship with cryptic aberrations detected in this chromosomal segment and responsible for mental retardation and congenital abnormalities, are described. Finally, we also include a brief report about the application of chromosome banding in the Human Genome Project and on its potential usefulness in the development of experiments on the artificial synthesis of eukariotic cell nuclei, of considerable significance in biology, is also included.

Résumé On montre, depuis une perspective historique, les systèmes d'identification des chromosomes humains et plusieurs méthodes de marquage chromosomique développées pendant les quatre dernières décades. On décrit des faits et des circonstances -dont l'auteur a été souvent protago-niste- qui ont été à l'origine de l'actuelle cytogénétique structurale et moléculaire. On fait aussi allusion à quelques aspects moléculaires implicites dans ces méthodologies afin de contribuer à expliquer de divers aspects des structures chromosomiques révélés par ces méthodes. On fait la description des procédés analytiques développés dans notre laboratoire pour l'étude de noyaux et de chromosomes ainsi que les résultats obtenus sur l'analyse quantitative de la région subtelomérique et son rapport avec les aberrations cryptiques repérées dans le segment chromosomique qui sont la cause de retard mental et de malformations congénitales. Finalement, on fait un bref descriptif sur les applications des bandes chromosomiques dans le projet du génome humain et sa possible utilité dans des expériments de synthèse artificielle de noyaux eucaryotes, de grande importance en biologie.

Bandeo de cromosomas humanos con extracto crudo de frutas u hojas de papaya / Human chromosome banding with raw extract of fruits or leaves of papaya

One week old human chromosome preparations were treated with filtrate from one liquefied leaf (53 g) of papaya (Carica papaya) in 100 ml of distilled water, and stained with 1.5 Giemsa (pH 6.8). Good chromosome banding was obtained after 2 min of treatment. Solutions that have been frozen even for years are effective and the method is cheaper and easier than others.

Chromosomes and C-banding of Opisthorchis viverrini.

Chromosome of Opisthorchis viverrini was observed by air-drying and C-banding techniques. The chromosome number was 2n=12 and n=6 consisting of one large-sized metacentric, one medium-sized metacentric, one small-sized metacentric, one small-sized submetacentric or subtelecentric, one small-sized subtelocentric or acrocentric and one small-sized acrocentric. The relative lengths of the chromosomes were 32.02 +/- 2.52, 23.28 +/- 1.98, 15.24 +/- 3.40, 13.39 +/- 3.11, 10.18 +/- 1.56 and 5.82 +/- 0.59% respectively. After C-banding treatment, two of the small-sized chromosomes showed a remarkable constitutive heterochromatin.

Evaluación del clon neoplásico en síndromes mielodisplásicos mediante hibridización in-situ / Fluorescence in situ hybridization to evaluate the neoplastic clon in myelodysplastic syndromes

La trisomía 8 es una de las alteraciones citogenéticas más frecuentes en neoplasias hematológicas y se asocia principalmente a los desórdenes mieloides. En este trabajo evaluamos la trisomía 8 en 20 pacientes con mielodisplasias mediante las técnicas de Bandeo G y de hibridización in situ por fluorescencia (FISH). Ambas metodologías se realizaron a partir de un cultivo a corto plazo de médula ósea. En base al estudio citogenético clasificamos a los pacientes en dos grupos: a) pacientes con trisomía 8 (6/20) y b) pacientes sin trisomía 8 (14/20). Mediante FISH, el porcentaje de pacientes con trisomía 8 se duplicó debido a que esta metodología no sólo confirmó la presencia de esta alteración en aquellos pacientes donde fue previamente identificado por Bandeo G, sino también en 3 casos con cariotipo normal, 1 caso sin células en división y 2 casos con alteraciones estructurales. Nuestros resultados demuestran que esta metodología posibilita la detección de clones presentes en baja proporción así como también estimar la real incidencia del clon neoplásico independientemente del ciclo celular de la célula tumoral.

Estudio cromosómico: experiencia del laboratorio de genética del Hospital San Juan de Dios / Chromosomal study: experience at the genetics laboratory at the San Juan de Dios Hospital

Se revisan los estudios cromosómicos realizados en el Hospital San Juan de Dios entre abril y diciembre de 1995. De total de 58 exámenes, el mayor porcentaje de alteraciones (57 por ciento) correspondió a trisomía 21 libre (síndrome de Down); en menor proporción se encontraron aneuploidia de cromosomas sexuales (síndrome de Klinefelter) 9,5 por ciento y síndrome de Turner (4,8 por ciento). Otras alteraciones correspondieron a trisomía 18, trisomía 13, trisomía 22 parcial, deleción 5p, deleción 13q en un paciente con retinobalstoma y translocación 9,22 (cromosoma Philadelfia)