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

Overexpression of STK15 and chromosomal instability in hepatocellular Carcinoma

This study aimed to examine the potential role of STK15 in Heptaocellular tumohgenesis. We analyzed the STK15 [protein and mRNA] expression and try to correlate STK15 expression with chromosome ploidy, various clinicopathological and molecular features of HCC. We aimed to evaluate chromosomal instability [CIN] by evaluating chromosome ploidy analyses, and correlate polsomy with various molecular features of HCC. Primary histologically proved HCC and adjacent nontumorous liver tissues from 29 patients were selected and surgically resected from Tanta University Hospital and Menophyia University Hospital. The tumor tissue and paired adjacent noncancerous liver tissue were obtained independently. We analyzedjhe STK15 expression by evaluating its mRNA and protein expression status using RT-PCR and Western blot and analysis. We try to correlate STK15 expression with chromosomae ploidy, various clinicopathological and molecular features. For chromosome ploidy analyses, FISH was carried out with direct fluorescence-labeled -satellite probes for chromosome 3,7,17 and 18 in both HCC and in adjacent noncancerous liver tissue. In the present study we demonstrated overexpression of STK15 protein in HCC. STK15 protein expression was highly elevated in 20 of the 29 tumors [69%] examined with 8 tumors displaying a >10-fold increase. It appeared that the extent of STK 15 protein expression, was more frequently associated with high grade and higher siage of the tumors. STK 15 mRNA was overexpressed in 20 tumors [69%]. In the nontumorous liver, STK15 mRNA was overexpressed in 2 cases [7%]. STK15 overexpression in HOC did not correlate with age, gender chronic HCV infection, and chronic HBV infection, but was associated with serum -fetoprotein elevation [P = 0.04]. Histologically, STK15-mRNA overexpression was more frequent in grade II-IIIB [P = 0.006]. STK15-mRNA overexpression was associated with portal vein tumor invasion [stage 1 1 IB HCC; P = 0.033], regardless of tumor size. As regard chromosomal ploidy all examined chromosomes were found to be aneuploid to a similar extent. The modal copy number of each chromosome ranged from two copies [disomy] to six copies [hexasomy], and the percentage of cells that did not carry the modal copy number was quite variable. Nine tumors [31%] showing numerical heterogeneity in all four examined chromosomes, were judged to be carrying CIN. Possible associations between the presence of CIN and potential prognostic factors [gender, age, tumor size, tumor stage, and differentiation grade] were examined by the X2 test, and none were identified. In summary, we have demonstrated that STK15 is highly expressed in HCC. Overexpression of STK15 may have important pathogenetic and prognostic significance in HCC. Also we can conclude that, CIN can be detectable in primary HCC using FISH technique. The STK15/BTAK/Aurora. A pathway play roles in hepatocellualr carcinogenesis and, therefore, could prove to be relevant novel disease markers and therapeutic targets

Modulation of immune activation and lipoproteins in chronic heart failure

Heart failure is a major problem of public health associated with poor outcome in the advanced stages, thus justifying its prevention based on the prevention and treatment of its principle etiological factors. In chronic heart failure mesenteric venous congestion leads to increased bowel permeability, bacterial translocation and thereby endotoxin release. The increased endotoxine challenge is hypothesized to cause immune activation, which might have an important role in the progression of heart failure. This study is designed to examine the degree of immune activation, to study the degree of endotoxin, interaction and to evaluate the beneficial immune regulatory effects of high density lipoprotein, [HDL] as an endotoxin, binding protein in CHF. We studied thirty five patients with CHF [Age 50+9 Years NYHA 2.7 +/- 0. 4, LVEF 28.0 +/- 6 all mean +/- SEM and eight healthy control subjects matched for age and sex. Serum levels of cholesterol, HDL-C, LDL-C, soluble tumor necrosis factor-alpha [sTNE-alpha], soluble tumor necrosis factor reteptor-l [sTNFl]., sCDI4, kidney function test, and liver function test were assessed. Patients with CHF had increased concentration of sTNF-Rl [P=0.005], TNF- alpha <0.05. whereas HDL levels were decreased [P=0.007]. sCDl4 [indicative of endotoxine cell interaction] was significantly increased, and correlated positively with TNF-alpha [r=0.68, P<0.001] and inversely with MDL. TNF-alpha as a measure of immune activation was inversely correlated with HDL [r=0.36, P=0.045]. From the present study we could concluded that in CHF patients increased concentrations of sCDI4 are related to lower HDL levels and increased concentration of TNF-alpha. This may support the role of HDL in the modulation of immune activation and cytokine and cytokine release in CHF