Detection of somatic TP53 mutations and 17p deletions in patients with chronic lymphocytic leukemia: a review of the current methods

ABSTRACT Chronic lymphocytic leukemia is the most common hematologic malignancy among adults in Western countries. Several studies show that somatic mutations in the TP53 gene are present in up to 50% of patients with relapsed or refractory chronic lymphocytic leukemia. This study aims to review and compare the methods used to detect somatic TP53 mutations and/or 17p deletions and analyze their importance in the chronic lymphocytic leukemia diagnosis and follow-up. In chronic lymphocytic leukemia patients with refractory or recurrent disease, the probability of clonal expansion of cells with the TP53 mutation and/or 17p deletion is very high. The studies assessed showed several methodologies able to detect these changes. For the 17p deletion, the chromosome G-banding (karyotype) and interphase fluorescence in situ hybridization are the most sensitive. For somatic mutations involving the TP53 gene, moderate or high-coverage read next-generation sequencing and Sanger sequencing are the most recommended ones. The TP53 gene mutations represent a strong adverse prognostic factor for patient survival and treatment resistance in chronic lymphocytic leukemia. Patients carrying low-proportion TP53 mutation (less than 20-25% of all alleles) remain a challenge to these tests. Thus, for any of the methods employed, it is essential that the laboratory conduct its analytical validation, documenting its accuracy, precision and sensitivity/limit of detection.

Genetic Aberrations in Caspase Family of Genes and Their Possible Association with HNSCC

The cell suicide pathway of apoptosis is a necessary event in the life of multicellular organisms. It is involved in many biological processes ranging from development to the immune response. Over expression of interleukin-1β-converting enzyme (later renamed caspase-1) was shown to be sufficient to induce apoptosis in mammalian cells. The present study aims to assess the gene alterations in the Caspase family of cytochromes so as to derive an association with HNSCC. Earlier eleven genes were found in the human genome to encode 11 human caspases, caspase-1 to caspase-10 and caspase-14, which is now populated to 13, whereas 10 genes were found in the mouse genome to encode 10 murine caspases including caspase-1, 2, 3, 6, 7, 8, 9, 11, 12 and 14 Caspases share a number of features distinguishable from other proteases. The analysis follows an observational study design, employing several computational tools to identify and predict the possible outcomes of gene alterations identified in HNSCC patients. cBioportal server was used to identify the gene alterations which was further analyzed using tools such as PROVEAN, I-Mutant and gnomAD. Several reported polymorphic variants were also identified. The pathogenicity and protein stability of gene alterations documented in the present study were identified at standard biological conditions. Further experimental studies would provide concrete evidence on the association of the observed genetic abnormalities with HNSCC especially in individuals exposed to habitual carcinogens

Secondary manifestations of mitochondrial disorders / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Mitochondrial disorders (MIDs) are a heterogeneous group of genetic metabolic diseases due to mutations in the mitochondrial DNA (mtDNA) or in the nuclear DNA (nDNA) (Rahman and Rahman, 2018). Some affected genes encode proteins with various functions, or structural RNAs such as transfer RNAs (tRNAs) and ribosomal RNAs (rRNAs). MIDs may also be caused by mutations in non-coding regions (e.g., D-loop of mtDNA) (Rahman and Rahman, 2018). Proteins involved in MIDs include enzymes, assembling factors, transport proteins, signaling proteins, pore proteins, and fusion/fission proteins (Gorman et al., 2016). The pathways most frequently affected by mutations in "mitochondrial genes" are the respiratory chain and the oxidative phosphorylation. Dysfunction of many other pathways (e.g., β-oxidation, pyruvate-dehydrogenase complex, and heme synthesis) may also manifest as MIDs (Hu et al., 2019). The estimated prevalence of MIDs is at least 1:5000 (Ng and Turnbull, 2016).

Targeted sequencing of the DMD locus: A comprehensive diagnostic tool for all mutations

Background & objectives: Duchenne muscular dystrophy (DMD) is an X-linked recessive disorder and is caused mainly by deletion, duplication and point mutations in the DMD gene. Diagnosis of DMD has been a challenge as the mutations in the DMD gene are heterogeneous and require more than one diagnostic strategy for the validation of the mutation. This study was planned to evaluate the targeted next-generation sequencing (NGS) as a single platform to detect all types of mutations in the DMD gene, thereby reducing the time and costs compared to conventional sequential testing and also provide precise genetic information for emerging gene therapies. Methods: The study included 20 unrelated families and 22 patients from an Indian population who were screened for DMD based on phenotypes such as scoliosis, toe walking and loss of ambulation. Peripheral blood DNA was isolated and subjected to multiplex ligation-dependent probe amplification (MLPA) and targeted NGS of the DMD gene to identify the nature of the mutation. Results: In the study patients, 77 per cent of large deletion mutations and 23 per cent single-nucleotide variations (SNVs) were identified. Novel mutations were also identified along with reported deletions, point mutations and partial deletions within the exon of the DMD gene. Interpretation & conclusions: Our findings showed the importance of NGS in the routine diagnostic practice in the identification of DMD mutations over sequential testing. It may be used as a single-point diagnostic strategy irrespective of the mutation type, thereby reducing the turnaround time and cost for multiple diagnostic tests such as MLPA and Sanger sequencing. Though MLPA is a sensitive technique and is the first line of a diagnostic test, the targeted NGS of the DMD gene may have an advantage of having a single diagnostic test. A study on a larger number of patients is needed to highlight NGS as a single, comprehensive platform for the diagnosis of DMD.

Intervención multidisciplinaria en paciente diagnosticado con el síndrome de Prader-Will / Interdisciplinary intervention in a pacient diagnosed with Prader-Willi síndrome

Introducción: El síndrome de Prader-Willi es una enfermedad genética, causada por deleciones de novo en la región 15q11q13 en el cromosoma paterno. Se caracteriza por falta de saciedad que conduce a obesidad mórbida, trastornos del comportamiento, discapacidad intelectual, baja estatura e hipogonadismo. Objetivo: Describir los resultados obtenidos del análisis e intervención multidisciplinar realizados en paciente diagnosticado con el síndrome de Prader-Willi. Presentación del caso: Análisis de caso clínico en menor de 8 años, sexo masculino, diagnosticado con síndrome de Prader-Willi, a través de intervención multidisciplinaria realizado en tres momentos: evaluación, diagnóstico e intervención con enfoque cognitivo conductual. Conclusiones: Las estrategias adoptadas generaron cambios significativos en el contexto familiar y social, entre ellas, apropiación de las recomendaciones suministradas, adopción de factores protectores, identificación de roles y optimización en la adherencia farmacológica. La atención a estas consideraciones proporciona mejoras, que apuntan a la calidad de vida y clínica del paciente(AU)

Introduction: Prader-Willi syndrome is a genetic disease caused by de novo deletions in the 15q11q13 region in the paternal chromosome. It is characterized by lack of satiety leading to morbid obesity, behavioral disorders, intellectual disability, short height and hypogonadism. Objective: To describe the results obtained from the multidisciplinary analysis and intervention performed in a patient diagnosed with Prader-Willi syndrome. Presentation of the clinical case: Clinical case analysis in an 8 years old child, male sex, diagnosed with Prader-Willi syndrome through a multidisciplinary intervention performed in three moments: assessment, diagnosis and intervention with cognitive behavioral approach. Conclusions: The strategies adopted generated significant changes in the social and family context, family´s appropriation of the recommendations provided, adoption of protective factors, roles identification and improving of adherence to treatment. By taking into account this considerations, improvements lead to clinic and life quality of the patient(AU)

A descriptive pilot study of mitochondrial mutations & clinical phenotype in fibromyalgia syndrome

Background & objectives: Fibromyalgia syndrome (FMS) is one of the most common chronic pain conditions of unknown aetiology. Mitochondrial dysfunction has been reported in FMS with some studies reporting the presence of mitochondrial mutation namely A3243G, which also causes mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes. This pilot study was conducted to assess this mutation and also detect large deletions in mitochondrial DNA (mtDNA) in patients with FMS. Methods: Thirty female patients with FMS participated and 30 matched controls were included. Genomic DNA was subjected to polymerase chain reaction (PCR) amplification using specific primers followed by restriction digestion with Apa I enzyme to detect the specific A3243G mtDNA mutation. Long-range PCR was done in two sets to detect the large deletions in the mtDNA. Biochemical parameters including thyroid-stimulating hormone and vitamin D levels were also looked at. Results: None of the patients were found to carry the common mutation or large deletions. Low vitamin D level was a common finding. Hypothyroidism was found in a few patients. Interpretation & conclusions: Although the common mutation or large mtDNA deletions were not detected in blood mtDNA in the FMS patients, mutations in the muscle and sequence variation in mtDNA remained a possibility. Future studies in both blood and muscle tissue including mtDNA sequencing are warranted in such patients to determine if a subset of FMS patients have mitochondrial myopathy.

Large-single scale mitochondrial DNA deletions in different tissues in Kearns-Sayre syndrome / 中华实用儿科临床杂志

Objective To investigate the clinical significance of different samples (the peripheral blood,urine and skeletal muscle) that could be detected the large-scale single deletions directly by using next-generation sequencing in the diagnosis of Kearns-Sayre syndrome (KSS) by concluding the clinical and genetic features of KSS,in order to explore a non-invasive method for diagnosis.Methods The clinical data,skeletal muscle's pathology and enzymology and genetic results of individuals with KSS,who were hospitalized from October 2016 to October 2017 in Department of Neurology,Beijing Children's Hospital,Capital Medical University,were collected.The gene tests were performed by using next generation sequencing technology and long-PCR technology of mitochondrial DNA(mtDNA) and the whole exon in the peripheral blood,urine and skeletal muscle.Results Four patients were all consistent with the diagnosis criteria of KSS,among whom the age of onset was 8.2 years old on average,and the initial symptoms were statue,ptosis,headache and vomiting,and visual impairment.The common symptoms of the 4 cases were ophthalmoplegia,exercise intolerance,development delay,loss of appetite,hypotonia,muscle weakness,with cerebrospinal fluid protein concentration over 1 000 mg/L,the cerebral magnetic resonance imaging showed that abnormal signals in the brainstem,in addition,white matter,thalamus,basal ganglia,cerebrum and cerebellum atrophy could be found.Moreover,3 cases had cardiac conduction block.Two cases had maternal family history.Molecular analysis of the 4 cases revealed the large-scale single deletions of mtDNA from the peripheral blood,the urine,the skeletal muscle through the nextgeneration sequencing,which were m.6460-15590(9 131 bp del),m.8482-13446(4 964 bp del),m.6831-14981 (8 151 bp del),m.7983-15495 (7 513 bp del),respectively.Among 3 cases who did pedigree analysis,only the mother of case 4 was detected with the same variation of the proband.Conclusions KSS is a rare mitochondrial disease,which could be detected with the single large scale mtDNA deletions in the peripheral blood,urine and skeletal muscle.With the development of the methodology,the diagnosis of KSS maybe no longer than depends on the muscle biopsy with the next-generation sequencing.And the possibility to get the positive results in the peripheral blood and urine by the non-invasive method could improve the molecular diagnosis of KSS.

Large-single scale mitochondrial DNA deletions in different tissues in Kearns-Sayre syndrome / 中华实用儿科临床杂志

Objective@#To investigate the clinical significance of different samples (the peripheral blood, urine and skeletal muscle) that could be detected the large-scale single deletions directly by using next-generation sequencing in the diagnosis of Kearns-Sayre syndrome (KSS) by concluding the clinical and genetic features of KSS, in order to explore a non-invasive method for diagnosis.@*Methods@#The clinical data, skeletal muscle′s pathology and enzymology and genetic results of individuals with KSS, who were hospitalized from October 2016 to October 2017 in Department of Neurology, Beijing Children′s Hospital, Capital Medical University, were collected.The gene tests were performed by using next generation sequencing technology and long-PCR technology of mitochondrial DNA(mtDNA) and the whole exon in the peripheral blood, urine and skeletal muscle.@*Results@#Four patients were all consistent with the diagnosis criteria of KSS, among whom the age of onset was 8.2 years old on average, and the initial symptoms were statue, ptosis, headache and vomiting, and visual impairment.The common symptoms of the 4 cases were ophthalmoplegia, exercise intolerance, development delay, loss of appetite, hypotonia, muscle weakness, with cerebrospinal fluid protein concentration over 1 000 mg/L, the cerebral magnetic resonance imaging showed that abnormal signals in the brainstem, in addition, white matter, thalamus, basal ganglia, cerebrum and cerebellum atrophy could be found.Moreover, 3 cases had cardiac conduction block.Two cases had maternal family history.Molecular analysis of the 4 cases revealed the large-scale single deletions of mtDNA from the peripheral blood, the urine, the skeletal muscle through the next-generation sequencing, which were m. 6460-15590(9 131 bp del), m.8482-13446(4 964 bp del), m.6831-14981(8 151 bp del), m.7983-15495(7 513 bp del), respectively.Among 3 cases who did pedigree analysis, only the mother of case 4 was detected with the same variation of the proband.@*Conclusions@#KSS is a rare mitochondrial disease, which could be detected with the single large scale mtDNA deletions in the peripheral blood, urine and skeletal muscle.With the development of the methodology, the diagnosis of KSS maybe no longer than depends on the muscle biopsy with the next-generation sequencing.And the possibility to get the positive results in the peripheral blood and urine by the non-invasive method could improve the molecular diagnosis of KSS.

Caracterización de la variabilidad genética de cepas de campo de Brucella canis aisladas en Antioquia / Characterization of the genetic variability of field strains of Brucella canis isolated in Antioquia

Brucella canis, un patógeno intracelular facultativo, es responsable de la brucelosis canina, una enfermedad zoonótica que afecta a los caninos y al hombre. En los primeros causa abortos y fallas reproductivas; en el ser humano genera síntomas inespecíficos. En el año 2005 se demostró la presencia de B. canis en Antioquia (Colombia). Las cepas halladas se identificaron como tipo 2. La secuenciación del genoma completo de una cepa de campo denominada Brucella canis str. Oliveri mostró indels específicos de especie; a partir de estos se buscó conocer características genómicas de las cepas de B. canis aisladas y establecer relaciones filogenéticas, así como el tiempo de divergencia de la cepa Oliveri. Se realizó PCR convencional y secuenciación de 30 cepas de campo, se identificaron 5 indels reconocidos en B. canis str. Oliveri, se empleó ADN de Brucella suis, Brucella melitensis y cepas vacunales de Brucella abortus como controles. Se determinó que las cepas de campo estudiadas comparten 4 de los 5 indels de la cepa Oliveri, lo que indica la presencia de más de una cepa de B. canis circulando en la región. El análisis filogenético se realizó con 24 cepas de Brucella mediante secuencias concatenadas de genes marcadores de especie. Se probó la hipótesis del reloj molecular y adicionalmente se realizó test de tasas relativas de Tajima. De esta manera se demostró que la cepa Oliveri, al igual que las otras cepas de B. canis analizadas, divergen de B. suis. Se rechazó la hipótesis del reloj molecular entre las especies de Brucella y se demostró una tasa de evolución y una distancia genética similar entre las cepas de B. canis.

Brucella canis is a facultative intracellular pathogen responsible for canine brucellosis, a zoonotic disease that affects canines, causing abortions and reproductive failure; and the production of non-specific symptoms in humans. In 2005 the presence of B. canis in Antioquia was demonstrated and the strains were identified as type 2. The sequencing of the genome of a field strain denoted Brucella canis str. Oliveri, showed species-specific indel events, which led us to investigate the genomic characteristics of the B. canis strain isolated and to establish the phylogenetic relationships and the divergence time of B. canis str. Oliveri. Conventional PCR sequencing was performed in 30 field strains identifying 5 indel events recognized in B. canis str. Oliveri. ADN from Brucella suis, Brucella melitensis and vaccine strains from Brucella abortus were used as control, and it was determined that all of the studied field strains shared 4 out of the 5 indels of the sequenced Oliveri strain, indicating the presence of more than one strain circulating in the region. Phylogenetic analysis was performed with 24 strains of Brucella using concatenated sequences of genetic markers for species differentiation. The molecular clock hypothesis and Tajima's relative rate test were tested, showing that the Oliveri strain, similarly to other canis species, diverged from B. suis. The molecular clock hypothesis between Brucella species was rejected and an evolution rate and a similar genetic distance between the B. canis were demonstrated.

Insertion-deletions burden in copy number polymorphisms of the Tibetan population.

BACKGROUND: Many studies have been conducted to identify either insertions-deletions (inDels) or copy number variations (CNVs) in humans, but few studies have been conducted to identify both of these forms coexisting in the same region. AIMS AND OBJECTIVES: To map the functionally significant sites within human genes that are likely to influence human traits and diseases. MATERIALS AND METHODS: In this report, we describe an inDel map in the 1051 Tibetan CNV regions obtained through CNV genotyping using Affymetrix Genome-wide single nucleotide polymorphism 6.0 chip. InDel polymorphisms in these copy number polymorphism regions were identified with a computational approach using the 2500 deoxyribonucleic acid sequences obtained from the 1000 Genome Project. RESULTS: The study identified a total of 95935 inDels that range from 1 bp to several bps in length which were found scattered across regulatory regions, exons and in introns of genes underlying the CNVs. A study on the distribution of inDels revealed that the majority of inDels were found in coding regions of the genome than the noncoding, while within the genes, inDels in intron regions were more followed by exonic regions and finally the regulatory regions. CONCLUSION: Study of inDels in CNV regions contribute to the enhanced understanding of the role played by the two variations and their collective influence on the genome. Further, a collection of these inDel genetic markers will aid in genetic mapping, further understanding of the phenotypic variability, identification of disease genes and in detecting novel CNVs.

Lipid peroxidation and large-scale deletions of mitochondrial DNA in asthenoteratozoospermic patients.

The correlation between malondialdehyde (MDA) an index of lipid peroxidation (LPO) with large-scale deletion mitochondrial DNA (mtDNA) was investigated in a case-control study with a total of 50 semen samples from infertile men, including 25 normozoospermic donor as the control group and 25 asthenoteratozoospermic (AT) patients as the case group. Routine semen analysis was performed according to World Health Organization (WHO, 1999) guidelines. MDA levels of the seminal plasma and spermatozoa were measured by TBARS method. A long-range polymerase chain reaction (PCR) method was used for the analysis of multiple large-scale mtDNA deletions based in two areas of mtDNA. The results showed that mean concentration of MDA in seminal plasma (nmol/ml) and spermatozoa (nmol/10 × 106 sperm) of AT men was higher than in normozoospermic patients, but the differences were not statistically significant. The products of PCR analysis showed multiple deletions of ~4.7, 4.8, 7.2, 7.3 and 7.4-kb in mtDNA of the spermatozoa in both AT and control groups. Multiple deletions were also observed in 64% of AT patients and 44% of the control group. Moreover, MDA level of the spermatozoa in deleted mtDNA samples group was significantly higher than in non-deleted mtDNA group (p = 0.01). Our findings indicated a positive correlation between increased MDA levels and large-scale mtDNA deletions in human spermatozoa. It is suggested that LPO or other oxidative stress factors might be causative elements in mtDNA damage, effect on sperm motility and morphology, resulting in decline of fertility in men.

Multiplex ligation-dependant probe amplification study of children with idiopathic mental retardation in South India.

BACKGROUND: Mental retardation (MR) is a heterogeneous dysfunction of the central nervous system exhibiting complex phenotypes and has an estimated prevalence of 1-3% in the general population. However, in about 50% of the children diagnosed with any form of intellectual disability or developmental delay the cause goes undetected contributing to idiopathic intellectual disability. MATERIALS AND METHODS: A total of 122 children with developmental delay/MR were studied to identify the microscopic and submicroscopic chromosome rearrangements by using the conventional cytogenetics and multiplex ligation dependent probe amplification (MLPA) analysis using SALSA MLPA kits from Microbiology Research Centre Holland [MRC] Holland. RESULTS: All the recruited children were selected for this study, after thorough clinical assessment and metaphases prepared were analyzed by using automated karyotyping system. None was found to have chromosomal abnormality; MLPA analysis was carried out in all subjects and identified in 11 (9%) patients. CONCLUSION: Karyotype analysis in combination with MLPA assays for submicroscopic micro-deletions may be recommended for children with idiopathic MR.

A novel deletion cluster at 13q14.2-q21.33 in an 80-year man with late onset leukemia: Clinical and molecular findings.

Chromosomal deletions are among the most common genetic events observed in hematologic malignancies; loss of genetic material is regarded as a hallmark of putative tumor suppressor gene localization. We have identified an unusual cluster of deletions at 13q14.2-13q21.33 in an 80-year-old father of a monozygotic twin pair discordant for schizophrenia, who developed chronic leukemia (CLL) at age 69. MATERIALS AND METHODS: The breakpoints for individual deletions in this cluster was identified by Affymetrix Human Array 6.0 screening. RESULTS: The deleted segments harbours a number of genes, most associated with cancer as well as a high concentration of LINEs, SINEs and related repeats. The derived chromosome represents an intra-chromosomal re-arrangement that quickly overtook blood progenitor cells probably before age 69 as a cause of CLL. CONCLUSIONS: The study highlights the role of ongoing de novo changes at susceptible sites, such as repeat rich regions, in the human genome. Also, it argues for the involvement of genes/deletions in the 13q(14.2-21.33) region in the development of CCL.

Prenatal Diagnosis of Chromosome 22q11.2 Deletions: Experiences in a Single Institution

PURPOSE: This study was designed to determine the frequency and echocardiographic findings of 22q11.2 deletions in fetuses with cardiac defects on fetal ultrasound or familial backgrounds of 22q11.2 deletions. MATERIALS AND METHODS: We retrospectively reviewed the medical and ultrasonographic records of 170 fetuses that underwent fluorescence in situ hybridization (FISH) analysis for chromosome 22q11.2 deletions between February 2001 and April 2013. RESULTS: Among 145 fetuses with cardiac defects, six (4.1%) had 22q11.2 deletions. Deletions of 22q11.2 were detected in 6 (5%) of the 120 fetuses with conotruncal defects: 5 (8.9%) of 56 with tetralogy of Fallot (TOF) and 1 (5.9%) of 17 with double outlet right ventricle (DORV). No deletions were found in cases of pulmonary atresia, truncus arteriosus, right aortic arch, or transposition of the great arteries. No 22q11.2 deletions were found in non-conotruncal cardiac malformations. Among 25 fetuses with familial backgrounds of 22q11.2 deletions, one (4%) had a maternally inherited 22q11.2 deletion with no cardiac findings. CONCLUSION: Knowledge of the frequency and echocardiographic findings of 22q11.2 deletions might be helpful for prenatal genetic counseling. It is advisable to perform FISH analysis for 22q11.2 deletions in pregnancies exhibiting conotruncal cardiac defects such as TOF or DORV.

Time-and dose-effect of mitochondrial DNA deletions in γ-ray irradiated human peripheral blood / 中华放射医学与防护杂志

Objective To study the time-and dose-effect of mitochondrial DNA (mtDNA) 4934 bp and 4977 bp deletions in the human peripheral blood irradiated by137 Cs γ-rays,and to evaluate its implication in biological dosimetry.Methods The peripheral blood from five healthy adults was collected and irradiated with γ-rays.The peripheral blood of one healthy adult was irradiated with 5 Gy and cultured for 2,24,48 and 72 h after irradiation.The peripheral blood from the other four healthy adults was cultured for 2 h after 0,0.5,1,2,5 and 10 Gy irradiation.The peripheral blood mtDNA 4934 bp and 4977 bp deletions were detected by real-time polymerase chain reaction and gel electrophoresis.The doseeffect curves were fitted using Curve Expert 1.4 Software.Results mtDNA 4934 bp and 4977 bp deletions were induced at 2 h post-irradiation and the mtDNA 4934 bp deletion had relative high levels at 2 h and 48h after radiation (t =10.782 and 8.966,P ＜ 0.05),and mtDNA 4977 bp deletion reached the highest level at 48 h after radiation (t =7.433,P ＜0.05).mtDNA 4934 bp (t =2.895-8.105,P ＜0.05) and 4977 bp deletion (t =3.006-7.715,P ＜0.05) irradiated at 0.5-10 Gy increased with a dosedependent manner.The incidence of mtDNA 4977 bp deletion was higher than that of 4934 bp deletion for those samples exposed with same dose of irradiation,especially at 10 Gy (t =2.919,P ＜ 0.05),which suggested that 4977 bp deletion might be more sensitive than 4934 bp deletion at high dose.But larger individual differences were found in 4977 bp deletion compared with 4934 bp deletion.The dose-effect equations for 4934 bp deletion and 4977 bp deletion were Y1 =1.178 + 0.1219D (R2 =0.9269) and Y2 =1.2578 +0.1933D (R2 =0.9016),respectively.Conclusions The induction of mtDNA deletion was correlated with radiation dose,and thus it may be a available method for biological dose estimation and prognostic evaluation.

Mutational Analysis of Mitochondria DNA in Children with IgA Nephropathy

PURPOSE: The association of mitochondrial DNA (mtDNA) mutations, deletions and copy number with progressive changes in patients with some glomerular disease and end-stage renal disease have been reported. In this study, we performed mtDNA mutation analysis in children with IgA nephropathy to investigate its role in progressive clinical course. METHODS: Seven children with IgA nephropathy were involved in this study. MtDNA isolated from platelet was amplified by PCR and sequenced entirely. RESULTS: The mean age at renal biopsy was 11.5+/-2.2 year and the mean age at latest evaluation was 17.9+/-3.2 year. The mean follow-up period were 7.8+/-3.1 years. Patients was divided into 2 groups according to the amount of proteinuria at presenting manifestation. Group 2 patients were nephrotic syndrome. Renal function reveals within normal range in all patients. In group 2 patients, the mean serum albumin level was significantly lower than those of group 1 (3.7+/-0.6 g/dL vs. 4.7+/-0.2 g/dL, P=0.0241) and the mean total cholesterol level was significantly higher than those of group 1 (222.7+/-35.7 mg/dL vs. 148.3+/-29.1 mg/dL, P=0.0283). In Group 2 patients, total amount of protein of 24 hour collected urine also significantly higher than those of group 1 (1,466.0+/-742.5 mg vs. 122.5+/-48.1 mg, P=0.0135). Pr/Cr ratio in random urine sample was also higher in group 2 than those of group 1 but the statistical significance was not noted (1.8+/-1.6 vs. 0.2+/-0.2, P=0.0961). Deletion of mtDNA nt 8272-8281 were observed in two patients, one patient in each groups, respectively. This is non-coding lesion. No patients demonstrated the mtDNA mutations. CONCLUSIONS: We have identified a deletion of mtDNA nt 8272-8281 in two children with IgA nephropathy. Further studies are needed to clarify the role of mitochondrial function in the progressive change of IgA nephropathy.

Identificación ee factores genéticos en la etiología de la esquizofrenia / Identification of Genetic Factors in the Etiology of Schizophrenia

Esquizofrenia es un trastorno mental que afecta aproximadamente 1% de la población mundial. Está caracterizada por episodios psicóticos, en los cuales los individuos presentan alucinaciones y/o delirios. A su vez, este trastorno involucra un fuerte componente de disfunción social, falta de motivación y deficiencias cognitivas profundas. Las causas de este trastorno se desconocen a ciencia cierta, aunque la evidencia acumulada indica que surge por alteraciones en el desarrollo del sistema nervioso central. Entre los factores que incrementan el riesgo a desarrollar este trastorno se encuentran varios elementos del ambiente incluyendo, infecciones y malnutrición prenatal, así como complicaciones durante el parto. Sin embargo, estudios detallados han corroborado la existencia de factores genéticos involucrados en el desarrollo de esquizofrenia y señalan a estos como los factores más importantes que parecen determinar la aparición de la enfermedad. A pesar de esto, la identificación de genes involucrados en el desarrollo de esta enfermedad ha resultado ser una de las tareas más difíciles que enfrentan la genética y la genómica. El desarrollo de técnicas modernas para el estudio del genoma humano ha permitido estudiar de forma sistemática las variaciones en la secuencia y estructura del genoma que dan lugar a esquizofrenia, permitiendo la identificación de cientos de genes, que pueden estar involucrados en el desarrollo de la enfermedad. Además, se ha sugerido que muchos de estos genes están involucrados en varias enfermedades mentales que en la actualidad se diagnostican como trastornos diferentes, pero cuyo substrato biológico pudiera ser similar.

Schizophrenia is a mental disorder that affects approximately 1% of the worldwide population. It is characterized by psychotic episodes in which individuals have hallucinations or delusions. This disorder also involves a strong element of social dysfunction, lack of motivation and profound cognitive deficits. The causes of this disorder remain largely unknown, but evidence indicates that arises from changes in the development of the central nervous system. Among the identified risk factors for this disorder are several environmental events, including prenatal infections and malnutrition, and complications during childbirth. However, the most important factor seems to be genetics. Despite this, the identification of genes involved in the development of this disorder has emerged as one of the most difficult tasks facing modern genetics and genomics. The development of techniques for studying the human genome has allowed a more systematic approach to determine variations in the genome sequence and structure that area casually involved in schizophrenia. These studies suggest the participation of hundreds of genes in schizophrenia development. In addition, it has been suggested that many of these genes are involved in various mental illnesses that today are diagnosed as separate entities, but whose biological substrate may be shared.

Deletion of ABL/BCR on der(9) associated with severe basophilia.

Chronic basophilic leukemia is a rare form in chronic myeloid leukemia patients. Only limited number of reports are available. Herein, we describe a patient who presented with fatigue, weight loss, leucocytosis, prominent basophilia, and mild eosinophilia. On biopsy, bone marrow was hypercellular with marked basophils. The immunophenotype showed abnormal expression of CD7, which is suggestive of basophilic maturation. Chromosomal analysis from GTG-banded metaphases revealed Ph positivity, and fluorescence in situ hybridization (FISH) with BCR/ABL dual color, dual fusion probe showed single fusion on the der(22) chromosome and ABL/BCR fusion was deleted on the der(9) chromosome. The deletion (ABL/BCR) on der(9) may be associated with basophilia which may be also indicative of the transformation of CML to acute myeloid leukemia.

Estimating genomic instability mediated by Alu retroelements in breast cancer

Alu-PCR is a relatively simple technique that can be used to investigate genomic instability in cancer. This technique allows identification of the loss, gain or amplification of gene sequences based on the analysis of segments between two Alu elements coupled with quantitative and qualitative analyses of the profiles obtained from tumor samples, surgical margins and blood. In this work, we used Alu-PCR to identify gene alterations in ten patients with invasive ductal breast cancer. Several deletions and insertions were identified, indicating genomic instability in the tumor and adjacent normal tissue. Although not associated with specific genes, the alterations, which involved chromosomal bands 1p36.23, 1q41, 11q14.3, 13q14.2, occurred in areas of well-known genomic instability in breast and other types of cancer. These results indicate the potential usefulness of Alu-PCR in identifying altered gene sequences in breast cancer. However, caution is required in its application since the Alu primer can produce non-specific amplification.

Detection of deletions and duplications in the Duchenne muscular dystrophy gene by the molecular method MLPA in the first Argentine affected families

Deletions/duplications in the Duchenne muscular dystrophy (DMD) gene account for 60 to 70% of all alterations. A new technique, multiplex ligation-dependent probe amplification (MLPA), has been described that allows the detection of large genetic rearrangements by simultaneous amplification of up to 45 target sequences. The present article is based on the diagnosis of the first Argentine affected families by the application of MLPA. DNA samples from patients with and without a previous diagnosis were included. MLPA assays were performed according to manufacturer recommendations. Polymerase chain reaction and direct sequencing were performed when a single-exon deletion was detected. Results were analyzed using the Gene Marker v1.6 and Sequencing Analysis v5.2 software. In the samples with a previous diagnosis (as identified by short tandem repeat-polymerase chain reaction analysis), MLPA confirmed in some samples the same deletion and detected in others a larger deleted fragment. This enabled the prediction of the expected male phenotype. One deletion and one duplication were detected in patients without previous diagnosis. In this study, we investigated the applicability of MLPA in our country. Our results showed a 100% confirmation of the deleted fragments detected by short tandem repeat segregation analysis. Moreover, in some cases, the MLPA assay was able to refine the breakpoints involved. In addition, MLPA identified deletions/duplications in samples without previous diagnosis. In comparison to the available diagnosis strategies in Argentina, MLPA is less time-consuming, and spans the complete coding region of DMD. The application of MLPA will improve the genetic diagnosis of DMD/Becker muscular dystrophy in our country.