Reversion from basal histone H4 hypoacetylation at the replication fork increases DNA damage in FANCA deficient cells.

The FA/BRCA pathway safeguards DNA replication by repairing interstrand crosslinks (ICL) and maintaining replication fork stability. Chromatin structure, which is in part regulated by histones posttranslational modifications (PTMs), has a role in maintaining genomic integrity through stabilization of the DNA replication fork and promotion of DNA repair. An appropriate balance of PTMs, especially acetylation of histones H4 in nascent chromatin, is required to preserve a stable DNA replication fork. To evaluate the acetylation status of histone H4 at the replication fork of FANCA deficient cells, we compared histone acetylation status at the DNA replication fork of isogenic FANCA deficient and FANCA proficient cell lines by using accelerated native immunoprecipitation of nascent DNA (aniPOND) and in situ protein interactions in the replication fork (SIRF) assays. We found basal hypoacetylation of multiple residues of histone H4 in FA replication forks, together with increased levels of Histone Deacetylase 1 (HDAC1). Interestingly, high-dose short-term treatment with mitomycin C (MMC) had no effect over H4 acetylation abundance at the replication fork. However, chemical inhibition of histone deacetylases (HDAC) with Suberoylanilide hydroxamic acid (SAHA) induced acetylation of the FANCA deficient DNA replication forks to levels comparable to their isogenic control counterparts. This forced permanence of acetylation impacted FA cells homeostasis by inducing DNA damage and promoting G2 cell cycle arrest. Altogether, this caused reduced RAD51 foci formation and increased markers of replication stress, including phospho-RPA-S33. Hypoacetylation of the FANCA deficient replication fork, is part of the cellular phenotype, the perturbation of this feature by agents that prevent deacetylation, such as SAHA, have a deleterious effect over the delicate equilibrium they have reached to perdure despite a defective FA/BRCA pathway.

Fanconi Anemia Patients from an Indigenous Community in Mexico Carry a New Founder Pathogenic Variant in FANCG.

Fanconi anemia (FA) is a rare genetic disorder caused by pathogenic variants (PV) in at least 22 genes, which cooperate in the Fanconi anemia/Breast Cancer (FA/BRCA) pathway to maintain genome stability. PV in FANCA, FANCC, and FANCG account for most cases (~90%). This study evaluated the chromosomal, molecular, and physical phenotypic findings of a novel founder FANCG PV, identified in three patients with FA from the Mixe community of Oaxaca, Mexico. All patients presented chromosomal instability and a homozygous PV, FANCG: c.511-3_511-2delCA, identified by next-generation sequencing analysis. Bioinformatic predictions suggest that this deletion disrupts a splice acceptor site promoting the exon 5 skipping. Analysis of Cytoscan 750 K arrays for haplotyping and global ancestry supported the Mexican origin and founder effect of the variant, reaffirming the high frequency of founder PV in FANCG. The degree of bone marrow failure and physical findings (described through the acronyms VACTERL-H and PHENOS) were used to depict the phenotype of the patients. Despite having a similar frequency of chromosomal aberrations and genetic constitution, the phenotype showed a wide spectrum of severity. The identification of a founder PV could help for a systematic and accurate genetic screening of patients with FA suspicion in this population.

MYC Promotes Bone Marrow Stem Cell Dysfunction in Fanconi Anemia.

Bone marrow failure (BMF) in Fanconi anemia (FA) patients results from dysfunctional hematopoietic stem and progenitor cells (HSPCs). To identify determinants of BMF, we performed single-cell transcriptome profiling of primary HSPCs from FA patients. In addition to overexpression of p53 and TGF-ß pathway genes, we identified high levels of MYC expression. We correspondingly observed coexistence of distinct HSPC subpopulations expressing high levels of TP53 or MYC in FA bone marrow (BM). Inhibiting MYC expression with the BET bromodomain inhibitor (+)-JQ1 reduced the clonogenic potential of FA patient HSPCs but rescued physiological and genotoxic stress in HSPCs from FA mice, showing that MYC promotes proliferation while increasing DNA damage. MYC-high HSPCs showed significant downregulation of cell adhesion genes, consistent with enhanced egress of FA HSPCs from bone marrow to peripheral blood. We speculate that MYC overexpression impairs HSPC function in FA patients and contributes to exhaustion in FA bone marrow.

FANCC Dutch founder mutation in a Mennonite family from Tamaulipas, México.

BACKGROUND: Fanconi anemia (FA) (OMIM #227650) is a rare hereditary disease characterized by genomic instability. The clinical phenotype involves malformations, bone marrow failure, and cancer predisposition. Genetic heterogeneity is a remarkable feature of FA; at least 22 FANC genes are known to cooperate in a unique FA/BRCA repair pathway. A common rule on the mutations found in these genes is allelic heterogeneity, except for mutations known to have arisen from a founder effect like the FANCC c.67delG in the Dutch Mennonite Community. Here, we present an 11-year-old male patient, member of the Mennonite Community of Tamaulipas México, with a clinical and cytogenetic diagnosis of FA. METHOD: Chromosome fragility test was performed in all siblings. Genomic DNA was obtained from peripheral blood samples. Sanger sequencing was used to identify the FANCC c.67delG mutation (NC_000009.11(NM_000136.2):c.67delG p.(Asp23IlefsTer23)) and its accompanying haplotype. RESULTS: The FANCC c.67delG mutation in 13 members of his family confirmed a FA diagnosis in two of his siblings and identified heterozygous carriers. Haplotype analysis supports that in this family, FA is caused by the founder mutation that initially appeared in Mennonite Dutch and followed this population's migrations through Canada and further to Mexico. CONCLUSION: The identification of the FANCC c.67delG mutation in this family not only allows proper genetic counseling, but it also grants the possibility to raise awareness of FA risk among the Mennonite community living in Mexico.

Nonclonal Chromosome Aberrations and Genome Chaos in Somatic and Germ Cells from Patients and Survivors of Hodgkin Lymphoma.

Anticancer regimens for Hodgkin lymphoma (HL) patients include highly genotoxic drugs that have been very successful in killing tumor cells and providing a 90% disease-free survival at five years. However, some of these treatments do not have a specific cell target, damaging both cancerous and normal cells. Thus, HL survivors have a high risk of developing new primary cancers, both hematologic and solid tumors, which have been related to treatment. Several studies have shown that after treatment, HL patients and survivors present persistent chromosomal instability, including nonclonal chromosomal aberrations. The frequency and type of chromosomal abnormalities appear to depend on the type of therapy and the cell type examined. For example, MOPP chemotherapy affects hematopoietic and germ stem cells leading to long-term genotoxic effects and azoospermia, while ABVD chemotherapy affects transiently sperm cells, with most of the patients showing recovery of spermatogenesis. Both regimens have long-term effects in somatic cells, presenting nonclonal chromosomal aberrations and genomic chaos in a fraction of noncancerous cells. This is a source of karyotypic heterogeneity that could eventually generate a more stable population acquiring clonal chromosomal aberrations and leading towards the development of a new cancer.

Genomic chaos in peripheral blood lymphocytes of Hodgkin's lymphoma patients one year after ABVD chemotherapy/radiotherapy.

Hodgkin's lymphoma (HL) is a lymphoid malignancy representing 5% of all cancers in children, 16% in adolescents, and 30-40% of all malignant lymphomas and has a survival rate of ~95% at 10 years. One of the most common treatment schemes uses a cocktail of genotoxic agents including adriamycin, bleomycin, vinblastine, and dacarbazine (ABVD) with or without radiotherapy. We investigated the occurrence of chromosomal damage in peripheral blood lymphocytes from five patients diagnosed with HL who provided samples before (BT), during chemotherapy (DT) and ~1 year after ABVD chemotherapy/radiotherapy (AT). Five healthy subjects served as controls. Chromosomal abnormalities were evaluated by multicolor fluorescence in situ hybridization. The average frequencies of structural chromosomal aberrations in HL samples were 0.11, 0.22, and 0.96 per cell in BT, DT, and AT samples, respectively. These frequencies were significantly different (P < 0.0001) with respect to control subjects (0.02 per cell). Interestingly, the highest frequency of structural damage, including genomic chaos and nonclonal abnormalities, was observed in the AT samples indicating that new aberrations were continuously produced. Rejoined structural chromosomal aberrations were the most common type of aberrations, although aneuploidies were also significantly increased. Finally, we found several chromosomal abnormalities linked to cancer secondary to treatment in all five HL patients. Our results show that ABVD chemotherapy plus radiotherapy is inducing genomic chaos in vivo; moreover, the persistence of genomic instability in the hematopoietic stem cells from HL patients may play a role in the occurrence of secondary cancer that is observed in 5-20% of HL patients. Environ. Mol. Mutagen. 59:755-768, 2018. © 2018 Wiley Periodicals, Inc.

TCNQ molecular semiconductor of the Cu(II)TAAB macrocycle: Optical and electrical properties.

The present study reports the doping of a semiconducting molecular material through the formation of hydrogen bonds between the macrocycle Cu(II)(TAAB) and the electronic acceptor TCNQ. According to density functional theory (DFT) calculations and electron paramagnetic resonance (EPR) analysis, the doped compound has the shape of a distorted square pyramid, with four nitrogen atoms in the equatorial position and the apical oxygen atom from the water ligands. These water molecules can generate strong hydrogen bonds with TCNQ and the TAAB metallic complex. Thin films of copper molecular material were obtained through high vacuum evaporation and were structurally characterized by IR spectroscopy, EPR and scanning electron microscopy (SEM). Additionally, the absorption coefficient (α) and photon energy (hν) were calculated from UV-vis spectroscopy and used to determine the optical activation energy in each film, from which its semiconducting behavior was established. An important aspect to consider is that the presence of hydrogen bonds is essential to establish the semiconducting nature of these species; this chemical behavior, as well as the resulting electronic mobility, have been studied by DFT theoretical calculations, which reinforce the experimental conclusion of a relationship between Cu(II)TAAB and TCNQ moieties generated by a weak bond. Finally, I-V characteristics have been obtained from a glass/ITO/doped molecular semiconductor/Ag device using Ag and ITO electrodes. Results for the copper-based device show that, at low voltages, the conduction process is of an ohmic nature while, at higher voltages, space-charge-limited current (SCLC) is found. It is highly probable that the doping effect in TCNQ favors electronic transport due to the formation of conduction channels caused by dopant-favored anisotropy.

7p15 deletion as the cause of hand-foot-genital syndrome: a case report, literature review and proposal of a minimum region for this phenotype.

BACKGROUND: Hand-foot-genital syndrome (HFGS) is a rare condition characterized by congenital malformations in the limbs and genitourinary tract. Generally, this syndrome occurs due to point mutations that cause loss of function of the HOXA13 gene, which is located on 7p15; however, there are some patients with HFGS caused by interstitial deletions in this region. CASE PRESENTATION: We describe a pediatric Mexican patient who came to the Medical Genetics Department at the National Institute of Pediatrics because he presented with genital, hand and feet anomalies, facial dysmorphisms, and learning difficulties. Array CGH reported a 12.7 Mb deletion that includes HOXA13. CONCLUSIONS: We compared our patient with cases of HFGS reported in the literature caused by a microdeletion; we found a minimum shared region in 7p15.2. By analyzing the phenotype in these patients, we suggest that microdeletions in this region should be investigated in all patients with clinical characteristics of HFGS who also present with dysplastic ears, mainly low-set implantation with a prominent antihelix, as well as a low nasal bridge and long philtrum.

NFE2L2 Gene Variants and Arsenic Susceptibility: A Lymphoblastoid Model.

Arsenic (As) exposure is a major risk for several types of cancer and metabolic diseases such as diabetes. The transcription factor nuclear factor erythroid 2-related factor (Nrf2) is a key mediator in the cellular defense against As-induced adverse effects. The -653G/A and -617C/A gene variants modulate expression levels of the Nrf2 coding gene (NFE2L2) and are postulated to be associated with several illnesses. In this study the functional effect of these polymorphisms was investigated in the cellular sensitivity to As-mediated effects. Using quantitative real-time polymerase chain reaction (qRT-PCR) the basal levels of NFE2L2 mRNA and the induced levels of NFE2L2 and its target gene NQO1 were measured in lymphoblastoid cells carrying different genotypes for -653G/A and -617C/A polymorphisms following As exposure. The effects of different NFE2L2 gene genotypes on cell proliferation were also explored after chronic metal exposure. A tendency toward reduction in basal levels of NFE2L2 mRNA was noted in the heterozygous (GA/CA) and risk homozygous (AA/AA) genotypes of both polymorphisms in immortalized lymphoblastoid cells. Although the expression of NFE2L2 and NQO1 after acute acute iAs exposure was not markedly influenced by -653G/A and -617C/A genotype, it was found that these single-nucleotide polymorphisms (SNPs) were correlated with a differential sensitivity to chronic exposure to the metalloid. Further studies are needed to completely understand the role of -653G/A and -617C/A SNPs in regulation of the NFE2L2 gene.

Hydroxyurea induces chromosomal damage in G2 and enhances the clastogenic effect of mitomycin C in Fanconi anemia cells.

Fanconi's anemia (FA) is a recessive disease; 16 genes are currently recognized in FA. FA proteins participate in the FA/BRCA pathway that plays a crucial role in the repair of DNA damage induced by crosslinking compounds. Hydroxyurea (HU) is an agent that induces replicative stress by inhibiting ribonucleotide reductase (RNR), which synthesizes deoxyribonucleotide triphosphates (dNTPs) necessary for DNA replication and repair. HU is known to activate the FA pathway; however, its clastogenic effects are not well characterized. We have investigated the effects of HU treatment alone or in sequential combination with mitomycin-C (MMC) on FA patient-derived lymphoblastoid cell lines from groups FA-A, B, C, D1/BRCA2, and E and on lymphocytes from two unclassified FA patients. All FA cells showed a significant increase (P < 0.05) in chromosomal aberrations following treatment with HU during the last 3 h before mitosis. Furthermore, when FA cells previously exposed to MMC were treated with HU, we observed an increase of MMC-induced DNA damage that was characterized by high occurrence of DNA breaks and a reduction in rejoined chromosomal aberrations. These findings show that exposure to HU during G2 induces chromosomal aberrations by a mechanism that is independent of its well-known role in replication fork stalling during S-phase and that HU interfered mainly with the rejoining process of DNA damage. We suggest that impaired oxidative stress response, lack of an adequate amount of dNTPs for DNA repair due to RNR inhibition, and interference with cell cycle control checkpoints underlie the clastogenic activity of HU in FA cells. Environ. Mol. Mutagen. 56:457-467, 2015. © 2015 Wiley Periodicals, Inc.

A Boolean network model of the FA/BRCA pathway.

MOTIVATION: Fanconi anemia (FA) is a chromosomal instability syndrome originated by inherited mutations that impair the Fanconi Anemia/Breast Cancer (FA/BRCA) pathway, which is committed to the repair of DNA interstrand cross-links (ICLs). The disease displays increased spontaneous chromosomal aberrations and hypersensitivity to agents that create DNA interstrand cross-links. In spite of DNA damage, FA/BRCA-deficient cells are able to progress throughout the cell cycle, probably due to the activity of alternative DNA repair pathways, or due to defects in the checkpoints that monitor DNA integrity. RESULTS: We propose a Boolean network model of the FA/BRCA pathway, Checkpoint proteins and some alternative DNA repair pathways. To our knowledge, this is the largest network model incorporating a DNA repair pathway. Our model is able to simulate the ICL repair process mediated by the FA/BRCA pathway, the activation of Checkpoint proteins observed by recurrent DNA damage, as well as the repair of DNA double-strand breaks and DNA adducts. We generated a series of simulations for mutants, some of which have never been reported and thus constitute predictions about the function of the FA/BRCA pathway. Finally, our model suggests alternative DNA repair pathways that become active whenever the FA/BRCA pathway is defective.

Detection of short-term chromosomal damage due to therapeutic 131I exposure in patients with thyroid cancer.

We evaluated the chromosomal aberration (CA) frequencies in the peripheral blood lymphocytes of ten female patients, age average 43.7 +/- 12.9, with thyroid cancer (TC) who had been exposed to 100-200 mCi therapeutic doses of 131I. The blood samples were obtained before-treatment and at 2 and 24 h after-treatment. Radiation was measured in the samples by means of dysprosium-activated calcium sulfate thermoluminescent dosimetry. The maximum radiation levels were detected in the samples taken 2 h after treatment. A positive correlation was found between the sample-emitted radiation values and the frequencies of CAs (r = 0.495; p < 0.01). The average baseline frequency of aberrations found in the ten studied patients was 0.009 per cell. Upon application of the 131I therapeutic dose, this frequency increased to 0.04 and 0.02 CAs/cell at 2 and 24 h after-treatment, respectively (p < 0.05). Break-type aberrations experienced a peak at 2 h after-treatment, whereas rejoined aberrations, such as dicentrics, rings, and radial figures, increased with sampling time. Seven patients with metastases had high amounts of CAs at 2 and 24 h after-treatment, in comparison to three patients without metastases who had a lower frequency of CAs at 24h aftertreatment. This difference could be due to the fact that circulating lymphocytes were exposed to a greater cancerous tissue mass, which retains 131I during the diagnostic and therapeutic processes. These results demonstrate the importance of detecting and surgically removing the largest possible amount of thyroid tissue in order to diminish the exposure of normal cells to radiation.

Differential expression of TP53 associated genes in Fanconi anemia cells after mitomycin C and hydroxyurea treatment.

Fanconi anemia (FA) is a rare, heritable chromosomal instability disease characterized by several congenital defects and cancer predisposition. Functional interactions between specific FA proteins and DNA damage response and repair activities have been reported, but the interplay between these mechanisms for maintaining genomic stability are not well understood. Many DNA damage response proteins are transcriptionally regulated by the tumor suppressor protein p53 (TP53), suggesting an important regulatory role for the DNA damage and stress response pathway. To better understand the association between FA and the DNA damage stress response we analyzed the levels of chromosomal damage and damage mediated gene transcription responses in lymphoblastoid cells derived from normal individuals and patients carrying the most common FA complementation group (FA-A). Chromosomal aberrations were first measured after exposure to mitomicyn C (MMC) or hydroxyurea (HU). Aliquots of the same cell were than assayed for the transcriptional response of 21 DNA damage and stress response genes using quantitative real-time PCR. The FA-A lymphoblastoid cells showed significant increases in the frequency of chromosome aberrations relative to non-FA-A lymphoblastoid lines after MMC treatment. The MMC induced damage was correlated with a general increase in expression of TP53-modulated DNA damage stress response genes involved in processes such as DNA repair, cell cycle progression, and apoptosis. Following HU treatment FA cells showed a decreased induction of CAs with much less transcriptional differences between targeted genes. Overall, the differences between the normal and FA-A cells after genotoxic treatments imply an increased activation and reliance of FA cells on the down-stream activities of TP53 for prevention of cell killing and chromosome damage from interstrand crosslinks but not for general replication arrest and double strand breaks. Furthermore, these results imply a regulatory connection between the FA pathway and activation of TP53 for responding to DNA damage. Alterations in the regulation of the DNA damage response may be related to the complex phenotypes seen in FA patients.

Duplication 2p and monosomy 8p in mosaicism: clinical, molecular cytogenetic and molecular markers of a unique case.

We report on a female patient, with a de novo mosaicism for a structural rearrangement producing trisomy 2p21-->pter and monosomy 8p21-pter. GTG bands and fluorescence in situ hybridization (FISH) in lymphocytes identified: mos 46,XX,der(8)(8qter-->8p21::p21::2p21-->pter),9qh +[52]/ 46,XX,9qh+[82]. Fibroblasts showed the same cell lines in 15 and 12 cells respectively. DNA profiling with fourteen autosomal STR markers, did not reveal a chimerism status in our patient. She did not present the classical phenotype described for trisomy 2p and for monosomy 8p probably due to approximately 60% of the patient's cells being normal. The abnormality probably arose in a very early stage of development during the first post-fertilization divisions with a non-sister chromatid exchange event between chromosomes 2 and 8 producing three cellular clones: a normal clone, one with trisomy 2p and monosomy 8p and a third with monosomy 2p and trisomy 8p. Only the first two cell lines were found in both lymphocytes and fibroblasts of hypopigmented skin; the third may have been lost or limited to other tissues.

DEB test for Fanconi anemia detection in patients with atypical phenotypes.

Pancytopenia, hyperpigmentation, small stature, congenital abnormalities, and predisposition to neoplasia characterize Fanconi anemia (FA). The clinical phenotype is extremely variable, therefore the diagnosis is frequently delayed until the pancytopenia appears, making diagnosis difficult on the basis of clinical manifestations alone. Hypersensitivity of FA cells to the clastogenic effect of diepoxybutane (DEB) provides a unique marker for the diagnosis before the beginning of hematological manifestations. Our aim in this study was to detect FA in children with atypical manifestations to define which conditions should be routinely included in the DEB test screening. We performed the chromosomal breakage test in 34 patients with probable FA and 83 patients with clinical conditions that could suggest FA, but are not usually screened by the DEB test: 20 patients with aplastic anemia, 20 patients with VACTERL association, 20 with radial ray abnormalities, 7 with tracheo-esophageal fistulae, 12 with anal atresia, and 4 with myelodysplastic syndrome. We found 18 DEB-positive patients: 12 were in the group of probable FA and 6 in the other groups. Among the last ones: three were included because of aplastic anemia, without any other sign of FA, however when re-examined, other anomalies were detected. The third patient had anal atresia, renal hypoplasia, pre-axial polydactyly, and normal blood cell counts and was diagnosed as having VACTERL association. The other two patients lacking physical or hematological signs were identified among the group of radial ray abnormalities. Thus, our results highlight the need to increase the number of abnormalities indicating need for a DEB test. Delay in the diagnosis of FA may have serious consequences for the patients and their family members.

Práctia de amamantamiento en madres trabajadoras y no trabajadoras que recibieron orientación sobre lactancia materna exclusiva / Breast feeding practice in working and non working mothers who received guidance about exclusive maternal nursing period

Objetivo: Verificar la práctia de amamantamiento por madres trabajadoras (MT) y no trabajadoras (MNT) que fueron capacitadas en lactancia materna exclusiva (LME). Método: Encuesta comparativa que incluyó 16 MT y 16 MNT con antecedente de embarazo de bajo riesgo y capacitadas en LME. Se excluyeron madres con productos que presentaban malformaciones congénitas bucodentomaxilares. La encuesta exploró capacitación sobre LME y se verificaron técnicas y procedimientos para amamantamiento en el domicilio de las pacientes. Resultados: Sólo 75 por ciento de las entrevistadas mantuvieron LME en los primeros 30 días del puerperio; posterior a este periodo, 38 por ciento de MNT y 63 por ciento de MT refirieron uso de medios alternos de lactancia. Las MT mencionaron LME a libre demanda en 19 por ciento más que las MNT. Conclusiones: La LME aún no forma parte contundente de la cultura en alimentación de lactantes, independientemente del ejercicio o no de trabajo por parte de la madre.

Detección de aneuploidías por hibridación in situ en células de mucosa oral / Detection of aneuploides in bucal mucosa by in situ hibridization

La técnica de hibridación in situ con fluorescencia (FISH) permite identificar alteraciones cromosómicas sin necesidad de cultivo en células en interfase. En este trabajo se realizó el estudio de FISH en 15 pacientes con diagnóstico citogenético de trisomía 21 regular, cinco pacientes cuyo estudio citogenético en sangre había mostrado la presencia de mosaicismo, y uno en el cual se identificó un cromosoma marcador sin evidencia de mosaicismo. Cinco sujetos con cariotipo normal sirvieron como testigo. De cada paciente se obtuvieron células de descamación de la mucosa oral; para facilitar la penetración de la sonda, se hizo pretratamiento mediante digestión con pepsina durante 20 minutos. En los 15 pacientes con síndrome de Down, la técnica de FISH con sonda Ó-satélite 13/21 mostró, en todas las células, cinco señales fluorescentes de las cuales tres correspondía al cromosoma 21 y dos al cromosoma 13, mientras que en los normales hubo cuatro señales. En un caso con fenotipo de síndrome de Turner, el estudio citogenético en 50 metafases había mostrado un cariotipo 46,X+mar mientras que el análisis de 1000 núcleos con la técnica de FISH permitió reconocer que el cromosoma maracador provenía de un cromosoma X y la presencia de un mosaicismo 45,X/46,X,r(X). En los otros casos de mosaicismo, el porcentaje de células con FISH pero estas diferencias no fueron significativas. En conclusión, el método de FISH en células de la mucosa bucal proporcionó resultaos confiables y al ser rápido y no invasivo, puede utilizarse para el diagnóstico de aberraciones cromosómicas, principalmente cuando se dificulta el estudio citogenético en sangre periférica

Efecto de la adición de medios condicionados sobre la respuesta de los linfocitos de anemia de Fanconi a la mitomicina-C / Effects of the addition of conditioned media on the response of Fanconi anemia lymphocytes to mitomicyn C

Se ha demostrado que la adicción de plasma normal a los cultivos de linfocitos de pacientes con anemia de fanconi (AF) disminuye significativamente la frecuencia de aberraciones cromosómicas inducidas por la mitocina C (MMC). Esto ha sugerido que el plasma normal contiene un factor de corrección (FC), capaz de complementar parcialmente a los linfocitos de la AF. Por otra parte, existe evidencia de que la alteración en la AF es a nivel de la reparación postreplicativa, y se puede suponer que el FC pertenece al sistema de reparación deficiente en la AF y que es de tipo inducible. En el presente estudio, los linfocitos AF previamente tratados con MMC crecieron durante las últimas 24 horas de cultivo, en medios condiconados obtenidos de linfocitos normales tratados y no tratados con MMC. El propósito fue investigar si las células normales dañadas con MMC eran inducidas para producir el FC in vitro. Los resultados no demostraron una complementación constante cuando los linfocitos AF se cultivaron en medios condicionados por linfocitos normales dañados con MMC (experimentos 2 y 3). Sin embargo, cuando los medios condicionados se suplementaron con plasma normal (experimentos 1 y 4) se observó una disminución significativa en la frecuencia de aberracioens cromosómicas en las células AF. Los resultados corroboraron la presencia del FC en el plasma normal; los posibles mecanismos de acción son que o bien participa en la reparación del ADN dañado por la MMC en las células AF o menos dañadas para que entren en mitosis.

Estudio de heterogeneidad genética en anemia de Fanconi por medio de la adición del plasma / Genetic heterogenicity study in Fanconis anemia by the addition of plasma

Con el objetivo de investigar heterogeneidad genética en la variedad infantil de anemia de Fanconi (AF) se realizaron cultivos de linfocitos de pacientes con Af con plasma autólogo, con plasma de otros pacientes con AF y con plasma normal. Se sembraron además linfocitos de un sujeto normal con plasma autólogo y con plasma de cada uno de los pacientes con AF. Para cada tipo de cultivo se cuantificó la freuencia de aberraciones cromossómicas inducidas por MMC. Los linfocitos de pacientes con AF cultivados con plasma autólogo mostraron hipersensibilidad a la MMC y una disminución significativa de la frecuencia de aberraciones inducidas en los cultivos con plasma normal. Sin embargo, en presencia del plasma de los otros pacientes la gran variabilidad en la respuesta de los linfocitos de los pacientes con AF a la MMC no permitió demostrar heterogeneidad genética. Esto pudiera deberse a que a pesar de existir heterogeneidad genética la metodología empleada no pudiera evidenciarla o bien, a que nuestros pacientes correspondiean ala mismo tipo de AF