Variedades de cultivo de líquido amniótico para el diagnóstico in vitro de poliploidías / Amniotic fluid culture varietys for poliploydies in vitro diagnosis

RESUMEN Fundamento: El cultivo celular permite el análisis directo de las células vivas mediante un microscopio. El estudio de las células contenidas en el líquido amniótico, mediante técnicas de cultivo, detecta anomalías en número y morfología de los cromosomas, que pueden relacionarse con enfermedades genéticas. Objetivo: Caracterizar las variedades de cultivo de líquido amniótico para el diagnóstico in vitro de poliploidías. Metodología: Se realizó un estudio descriptivo transversal, en el Centro Provincial de Genética Médica de Camagüey, en el periodo de noviembre de 2016 a abril de 2018.La población de estudio estuvo constituida por 1571 muestras útiles de líquido amniótico obtenidas por amniocentesis, en gestantes en el segundo trimestre, evaluadas en consulta multidisciplinaria con criterios clínicos de estudios cromosómicos según lo establecido en el diagnóstico prenatal citogenético, previo consentimiento informado. Se utilizaron 20 mL de líquido amniótico para la siembra de células fetales, y se aplicaron tres variantes de cultivo abierto (directo, centrifugado y expandido). Se determinó el complemento cromosómico en cada variedad. Resultados: Predominó el complemento cromosómico normal. Las tetraploidías prevalecieron en el cultivo expandido. El índice mitótico fue similar en las tres variedades de cultivo y el cultivo directo tuvo el más bajo índice de poliploidías. Conclusiones: El cariotipo normal fue predominante. Las tetrapolidías fueron las alteraciones más frecuentes y prevalecieron en el cultivo expandido. En el cultivo directo se presentó el más bajo índice de errores inducidos in vitro.

ABSTRACT Background: Cell culture allows direct analysis of live cells under a microscope. The cell study contained in amniotic fluid, by culture techniques, detects abnormalities in chromosome number and morphology, which can be related to genetic diseases. Objective: To describe amniotic fluid culture strains for the in vitro diagnosis of polyploidy. Methodology: A cross-sectional descriptive study was conducted at the Camagüey Provincial Center of Medical Genetics, from November 2016 to April 2018.The study population consisted of 1571 useful amniotic fluid samples obtained by amniocentesis, in pregnant women in the second trimester, evaluated by multidisciplinary discussion with clinical criteria for chromosomal studies as established in the cytogenetic prenatal diagnosis, prior informed consent. 20 mL of amniotic fluid were used for fetal cell seeding, and three open culture strains (direct, centrifuged and expanded) were applied. Chromosomal complement was determined in each variety. Results: Normal chromosome complement was predominant. Tetraploidy prevailed in the expanded culture. The mitotic index was similar in the three culture strains and the direct culture had the lowest polyploidy index. Conclusions: Normal karyotype was predominant. Tetraploidy were the most frequent modifications and prevailed in the expanded culture. Direct culture had the lowest rate of the in vitro induced errors.

Tetraploid/near-tetraploid acute promyelocytic leukaemia with double (15;17) translocation

@#A 57-year-old man presented with intermittent fever and bleeding following dental surgery. Peripheral smear and bone marrow aspirate exhibited unusually large and bizarre-looking abnormal cells which were found to be myeloblasts with aberrant CD56 and CD2 expression on immunophenotyping. Fluorescence in situ hybridization analysis revealed an extra RARA gene rearrangement. This finding correlated well with a near-tetraploid karyotype with double t(15;17)(q22;q21). Bcr-3 type PML/ RARA copies were identified in reverse transcriptase-polymerase chain reaction. The diagnosis of near-tetraploid acute promyelocytic leukaemia (APML) was established. The patient was treated with all-trans retinoic acid and idarubicin and six weeks later achieved complete remission. Tetraploid/ near-tetraploid APML is exceedingly rare. It is a distinct cytogenetic subgroup with unique clinical and biological features as highlighted by atypical morphology, frequent CD2 expression and association with the bcr-3 type PML/RARA fusion transcripts. Early recognition of this rare entity is essential for timely and appropriate treatment.

Chromosomal Abnormalities in Myelodysplastic Syndrome with Near-tetraploidy: A Case Report

Massive hyperdiploidy and tetraploidy are rare cytogenetic abnormalities in myelocytic malignancies, especially in myelodysplastic syndrome (MDS). These abnormalities are known to be associated with leukemogenesis, leukemic transformation and poor prognosis. We report here the first case of MDS with near-tetraploid cytogenetic abnormality in Korea. A 80-yr-old male was diagnosed with refractory anemia with excess blasts-2 (RAEB-2). Bone marrow aspiration smear showed 16% of blasts, which were large sized myeloid blasts with irregular margins and cytoplasmic vacuolation. Cytogenetic analysis of bone marrow cells revealed numercal and structural cytogenetic abnormalities including near-tetraploidy in 8 of 20 metaphases: 45,XY,add(1)(p36.1),del(10)(p11.2),del(11)(q13),-12,-16,der(17)t(11;17) (q13;q21),add(20)(q13.1),+mar[8]/85~90,idemx2[cp8]/46,XY[4]. After chemotherapy with decitabine, he showed pancytopenia during follow-up period and died of sepsis 14 months after the diagnosis.

Effects of Chromosomal Polyploidy on Survival of Colon Cancer Cells / 대한소화기학회지

BACKGROUND/AIMS: Tetraploid cells are frequently observed in the inflamed mucosal epithelial cells of the patients with Barrett's esophagus or chronic ulcerative colitis. Polyploidy often occurs during cell fusion, abortive cell cycle, and endoreplication. Most tetraploid cells are engaged to apoptotic pathway, but some remaining stable tetraploid cells consequently cause aneuploidization and chromosomal instability. We investigated whether tetraploid cells could acquire survival advantage and hold a dominant position for natural selection. METHODS: We established tetraploid cell line (HCT116GH) from parental diploid colorectal cancer cell line (HCT116) via PEG-mediated cell fusion and compared its cell viability, cell cycle response and apoptotic fractions responded to H2O2 with diploid HCT116 and p53 suppressed HCT116/H6 cell lines. RESULTS: Using MTT assay, plating efficiency and clonogenicity, we evaluated the survival of each cell line. Tetraploid cell line HCT116GH demonstrated an 83 fold greater resistance to 100 microM H2O2 than the parental diploid HCT116, and 6 fold greater than even the p53 negative diploid HCT116/E6. Cellular sensitivity, G2/M arrests, and apoptotic proportion were observed less in response to H2O2 in HCT116GH compared with HCT116 and HCT116/E6. HCT116GH expressed lower level of p53 and p21 than diploid HCT116. CONCLUSIONS: Stable tetraploid cell lines showed enhanced viability in comparison to parental diploid cell lines. The enhanced viability observed in tetraploidization surpassed that from downregulation of p53. Frequent appearance of tetraploid cells in stressful condition can be caused by natural selection owing to their enhanced viability and may consequently contribute to cancer cell transformation.

Near-tetraploidy Acute Myeloid Leukemia with RUNX1-RUNX1T1 Rearrangement Due to Cryptic t(8;21) / 대한진단검사의학회지

Tetraploidy or near-tetraploidy is a rare cytogenetic abnormality found in AML, and is divided into primary and secondary forms. The secondary tetraploidy or near-tetraploidy found in AML is known to be specifically associated with t(8;21). In this case report, FISH analysis detected RUNX1-RUNX1T1 gene rearrangement in the absence of cytogenetic abnormality of t(8;21), which suggests the presence of unvailed t(8;21). This is the first case report of tetraploidy or near-tetraploidy AML with cryptic RUNX1/RUNX1T1 in Korea. Although the prognosis of tetraploidy or near- tetraploidy with t(8;21) is known to be poor, this patient shows a relatively good clinical course compared to other reported cases.

A Case of Mosaic Tetrapaloidy

Tetraploidy of human chromosome(92, XXYY) has been described very rarely. Liveborn infant with tetraploidy was reported in only 17 cases(complete 7 cases, mosaicism 10 cases) in the world, and no cases have been reported in Korea. The diploid-tetraploid mosaicism could arise during the early mitosis of the zygote. In one blastomeric cell, the chromosomes replicate but the cytoplasrn does not divide. We report a case of live infant with mosaic tetraploidy detected in lymphocyte(30%) and amniotic cell culture(25%), who presented with multiple congenital anomalies. A brief review of the literature is included.