Chromosomal aberrations in childhood acute lymphoblastic leukemia: diagnostic and prognostic implications

Cytogenetic analysis in ALL is often hampered by poor chromosome morphology, few malignant metaphases, undetectable chromosomal rearrangements due to regions of a similar size and banding pattern and sometimes only normal metaphases derived from normal cells are found after cell culture. Structural as well as numerical aberrations may therefore remain undetected using conventional G-banding. The application of modern molecular cytogenetic techniques including a broad set of fluorescence in situ hybridization [FISH] has greatly improved the detection rate of genetic changes in ALL. The present study was designed to estimate the incidences of different genetic subgroups in childhood ALL with abnormalities involving BCR/ABL, MLL, TEL/AML1 rearrangements, and p16 deletions using FISH technique and conventional cytogenetic analysis. We tried to demonstrate the usefulness of FISH technique. This study was conducted on BM and/or BP from 48 patients with childhood ALL. Their age range from 2-13 years mean age was 6.7 years. Patients were followed-up for 18 months [range 14-28 months]. Morphological, cytochemical, immunophenotyping, cytogenetic and FISH analysis were performed for every patient. FISH was performed with probes for BCR/ABL, MLL, TEL/AML1 rearrangements, and p16 deletions for each case of childhood ALL. Numerical and/or structural aberrations were identified in 52.1% of all cases by conventional G-banding alone. Numerical and/or structural aberrations were identified in 75% of all cases by the combination of conventional G-banding and interphase FISH. Gene rearrangements were disclosed by FISH in 11 [47.8%] of 23 patients who showed a normal banded karyotype or no mitotic cell in G-banding. The most common gene rearrangement was p16 deletion [21.27%] and the incidences of others were 15.9% for TEL/AML1, 12.1% for MLL, and 5% for BCR/ABL rearrangement. p16 homozygous deletions were observed in sex cases [12.7%] and hemizygous deletions in four cases [8.5%]. One case had both in two different cell populations. p16 deletions were significantly more common among T-lineage ALL [T-ALL] patients than among precursor-B ALL patients. TEL/AML1 translocations were found in seven [7/44] [15.9%]. Three out of the seven cases show culture failure and none of the remaining cases showed t [12; 21] in G-banding analysis. All those seven patients were pre-B cell lineage according to standard immunophenotyping. One patient showed the loss of one AML1 signal in addition to the TEL/AML1 fusion. MLL rearrangements [11q23 abnormalities] was detected in 5/41 [12.1%] by combined conventional cytogenetic analysis and by FISH. Two different types of MLL gene rearrangements were observed in FISH analysis; translocation and deletion. One had split signal of the MLL gene caused by a translocation between chromosome 6 and 11 t [6; 11], detected by conventional cytogenetics. Amplification of MLL gene was observed in one case [2.27%] Four of five cases with MLL translocations showed no chromosome abnormality involving 11q23 in G-banding analysis. All cases with MLL gene rearrangement were pre-B cell lineage according to standard immunophenotyping. BCR/ABL rearrangement: t [9; 22] [q34; q11] was detected by conventional cytogenetic and by FISH in one case. Another one displayed BCR/ABL1 fusion signal by FISH only. FISH can overcome some limitations of conventional cytogenetic and molecular-genetic analyses and due to high sensitivity specific chromosomal aberrations in mitoses and/or interphase nuclei can be detected. FISH analysis using DNA probes specific for p16 deletion, TEL/AML1, MLL, and BCR/ABL gene rearrangements is a powerful tool for leukemia diagnosis and risk stratification and it should be used as a routine procedure for all patients with newly diagnosed ALL as well as for monitoring of treatment effect in children with ALL

Airway major mucins in asthmatic children

Mucus is a protective coating secreted in the healthy airway. Structurally, mucins are complex glycoconjugates: their protein backbones are products of mucin [MUC] genes. Twenty mucin genes have been reported. MUC5AC and MUC5B are major gel-forming mucins in normal or pathologic airway secretions. Sputum production is a common symptom in asthma, especially during asthma exacerbations contributing to: airway hyperresponsiveness, airways obstruction, decreasing FEV[1] and fatal attacks of asthma. The aim of this study was to evaluate the expression and distribution of MUC5AC and MUC5B in the sputum and bronchial biopsies of mild and moderate asthmatic children. 2 9/12 years prospective study. Chest unit, Pediatric Department, ENT Department, Tanta University Hospital. 25 asthmatic children during and after acute attack, admitted and treated in chest unit;16 males and 9 females, aged 6-13 years [mean 9.4 +/- 3.6 yr.] On admission, the severity of the asthmatic paroxysm was mild persistent asthma [MiPA] in 14 patients and moderate Persistent asthma [MoPA] in 11. All were treated in chest ward. The study involved [1] sputum induction with RNAs extraction and direct Quantification of MUC5AC and MUC5B mucins of the sputum. [2] Bronchoscopy with mucosal biopsies from each subject for RNA extraction and immunohistochemical analysis. Semiquantitative reverse-transcription polymerase chain reaction [RT-PCR] was performed for MUC5AC and MUC5B to investigate their expression. On RT-PCR examination of the sputum sample, MUC5AC was significantly detected in 80%, 92% and MUC5B in 52%, 68% in MiPA and MoPA respectively compared with controls. MUC5AC and MUC5B mRNAs were amplified weekly in endobronchial mucosa of the controls, whereas they showed two- and threefold mRNA upregulation, in MiPA and MoPA respectively. In sputum samples there were significantly more mucins in MiPA and MoPA than in controls. In addition, there were significantly more mucins in MoPA than in MiPA. Also there was significantly more MUC5AC than MUC5B in each group. MUC5AC immunoreactivity in asthmatics was abundant in goblet cells with no staining of submucosal glandular cells. While immunoreactivity for MUC5B in asthmatics showed abundant signaling in submucosal glandular cells and moderately positive staining in epithelial goblet cells. Goblet cell number was significantly increased in MiPA and MoPA in comparison with controls with no difference between MiPA and MoPA. This study was designed to characterize mucin gene expression in tracheobronchial mucosa and sputum in asthmatic children. The present results suggest that upregulation of MUC5AC and MUC5B with the associated goblet cell hyperplasia [GCH] may play important role in the pathophysiology of asthma. We found that even mild asthma was associated with GCH and increased stored mucin in the airway epithelium. Moderate asthma has even more increased levels. Further elucidation of the regulation of specific airway mucin genes by relevant mediators and identification of the mechanisms that result in GCH is needed

Chromosomal aberrations in childhood acute lymphoblastic leukemia: diagnostic and prognostic implications

Cytogenetic analysis in ALL is often hampered by poor chromosome morphology, few malignant metaphases, undetectable chromosomal rearrangements due to regions of a similar size and banding pattern and sometimes only normal metaphases derived from normal cells are found after cell culture. Structural as well as numerical aberrations may therefore remain undetected using conventional G-banding. The application of modem molecular cytogenetic techniques including a broad set of fluorescence in situ hybridization [FISH] has greatly improved the detection rate of genetic changes in ALL. The present study was designed to estimate the incidences of different genetic subgroups in childhood ALL with abnormalities involving BCR/ABL, MLL, TEL/AML1 rearrangements, and p16 deletions using FISH technique and conventional cytogenetic analysis. We tried to demonstrate the usefulness of FISH technique. This study was conducted on BM and/or BP from 48 patients with childhood ALL. Their age range from 2-13 years mean age was 6.7 years. Patients were followed-up for 18 months [range 14-28 months]. Morphological, cytochemical, immunophenotyping, cytogenetic and FISH analysis were performed for every patient. FISH was performed with probes for BCR/ABL, MLL, TEL/AML1 rearrangements, and p16 deletions for each case of childhood ALL. Numerical and/or structural aberrations were identified in 52.1% of all cases by conventional G-banding alone. Numerical and/or structural aberrations were identified in 75% of all cases by the combination of conventional G-banding and interphase FISH. Gene rearrangements were disclosed by FISH in 11 [47.8%] of 23 patients who showed a normal banded karyotype or no mitotic cell in G-banding. The most common gene rearrangement was p16 deletion [21.27%] and the incidences of others were 15.9% for TEL/AML1, 12.1% for MLL, and 5% for BCR/ABL rearrangement. p16 homozygous deletions were observed in six cases [12.7%] and hemizygous deletions in four cases [8.5%]. One case had both in two different cell populations. p16 deletions were significantly more common among T-lineage ALL [T-ALL] patients than among precursor-B ALL patients. TEL/AML1 translocations were found in seven [7/44] [15.9%]. Three out of the seven cases show culture failure and none of the remaining cases showed t[12;21] in G-banding analysis. All those seven patients were pre-B cell lineage according to standard immunophenotyping. One patient showed the loss of one AML1 signal in addition to the TEL/AML1 fusion. MLL rearrangements [11q23 abnormalities] was detected in 5/41 [12.1%] by combined conventional cytogenetic analysis and by FISH. Two different types of MLL gene rearrangements were observed in FISH analysis; translation and deletion. One had split signal of the MLL gene caused by a translation between chromosome 6 and 11 t[6;11], detected by conventional cytogenetics. Amplification of MLL gene was observed in one case [2.27%]. Four of five cases with MLL translocations showed no chromosome abnormality involving 11q23 in G-banding analysis. All cases with MLL gene rearrangement were pre-B cell lineage according to standard immunophenotyping. BCR/ABL rearrangement: t[9;22][q34;q11] was detected by conventional cytogenetic and by FISH in one case. Another one displayed BCR/ABL1 fusion signal by FISH only. FISH can overcome some limitations of conventional cytogenetic and molecular-genetic analyses and due to high sensitivity specific chromosomal aberrations in mitoses and/or interphase nuclei can be detected. FISH analysis using DNA probes specific for p16 deletion, TEL/AML1, MLL, and BCR/ABL gene rearrangements is a powerful tool for leukemia diagnosis and risk stratification and it should be used as a routine procedure for all patients with newly diagnosed ALL as well as for monitoring of treatment effect in children with ALL

role of helicobacter pylori in recurrent aphthous stomatitis in children

Helicobacter pylori [HP] organisms are spiral, microaerophilic, gram-negative bacteria affecting 70-90% of the population in developing countries. Infection is acquired before the age of 10 years. HP causes the majority of gastric and duodenal ulcers. Transmission may be by; ingestion, fecal-oral and oral-oral routes. In recurrent aphthous stomatitis [RAS], various microorganisms have been suspected but, the histological similarities between RAS and peptic ulcers, and the response of RAS to the broad-spectrum antibiotics suggested that HP may has a probable role in RAS development. To determine the presence of Helicobacter pylori and, if detected, its potential prevalence in causing recurrent aphthous ulcers confined to mucosa-associated lymphoid tissues [MALT] of the pharynx. 17-months prospective, controlled study. Pediatric Department, Tanta University Hospitals, Tanta, Egypt. A total of 80 patients with recurrent multiple aphthous ulcers of the oral cavity and pharynx were assigned to group 1 [n=32] [6-12 years; mean age, 8 +/- 2 years; 14 male and 18 female], in whom the ulcers were strictly limited to the mucosa-associated lymphoid tissues, or group 2 [n=48] [7-13 years; mean age, 9 +/- 3 years; 22 male and 26 female], in whom the ulcers were randomly distributed in the oral cavity and pharynx. 20 sex- and age-matched children served as normal control. Helicobacter pylori DNA was extracted from 3mm diameter tissue samples and polymerase chain reaction [PCR] amplifications were performed for the16S ribosomal RNA gene. HP DNA was detected in 24 patients [75%] in group [I]; in group [II], 6 patients [12.5%] were shown to be PCR positive. HP DNA was not detected in any of the control samples. There is a possible causative role for HP in recurrent aphthous ulcerations with a characteristic distribution and affinity to MALT of the pharynx. Hence; RAS and the risk of HP-associated gastrointestinal complications can be decreased with therapies for eradicating HP. It is recommended that tonsillectomy and adenoidectomy for MALT affected by RAS, improving oral hygiene may protect the host against HP infection and re-infection

Relation between cytotoxin producing Helicobacter pylori and Helicobacter pylorielated upper gastrointestinal disease

Helicobacter pylori [H. pylori], a major aetiological agent in gastritis, peptic ulcer and gastric cancer, is estimated to infect more than 50% of the world's population. However, only about 10% will develop peptic ulcer disease and 1 - 2% gastric malignancy. Virulent strains carrying the cag A gene and vac A s1 genotype and capable of cytotoxin production have been proposed to be associating with the severer forms or disease, although this was not universal. We were interested in studying the relation between cytotoxin-producing H. pylori strains and the H. pylori-related upper gastrointestinal disease in our community. Sixty patients were allocated into 3 predefined groups according to their endoscopic picture: gastroesophageal reflux disease [GERD], peptic ulcer disease, and gastritis groups. Gastric biopsies from the patients were examined for the presence of H. pylori by urease test and culture. The isolated H. pylori strains were subjected to cytotoxic assay to detect cytotoxin-producing strains. Forty-one patients [68.3%] were H. pylori positive, of them 19 [46.3%] were positive for cytotoxin production. Cytotoxin-producing H. pylori strains significantly associated peptic ulcer disease where 73.3% of peptic ulcer patients were infected with cytotoxin-producing strains. GERD was significantly associated with absence of H. pylori infection [66.7% of GERD patients were free of H. pylori infection]. The presence of gastritis did not correlate with the H. pylori status, however, there was a significant association between cytotoxin-producing strains and atrophic gastritis. Cytotoxin-producing H. pylori strains are associated with severer H. pylori-related upper gastrointestinal diseases such as atrophic gastritis and peptic ulcer disease. Our findings support the hypothesis of cag A [+] H. pylori being protective against GERD. Determination of cag A status of H. pylori strains bears importance in clinical practice in detecting patients at increased risk for developing gastric cancer and in helping planning treatment strategies