ObjectiveTo study the effects of chemotherapy on growth development and endocrine function of long-survived children with acute lymphoblastic leukemia (ALL). Methods30 ALL patients who were received standard chemotherapy and survived more than five years were enrolled in this study. Their growth and development data and endocrine function examination were investigated. ResultsThrough testing,except two cases of height more than two standard deviation above,the others were all within the normal range; BMI exceeded bid in 1, and the rest were in the normal range;The results of sex hormones examination were consistent with age and Tanner installment, the girls appeared secondary sex characteristics in 9 years old or so,menstruation in 13 years old. Boys appeared sec-ondary sex characteristics around 10 years old;Cortisol and promote adrenal cortical hormone with 2 cases of obese children were in the normal range,but c-peptide and insulin were elevatory;The results of IGF-1 were in the normal range. ConclusionChemotherapy had no significant effect on growth development and endocrine function for patients with ALL.

OBJECTIVE:To evaluate the therapeutic effect of Xuesaitong soft capsule on symptoms of TCM in patients with stroke and cerebral infraction. METHODS:112 patients were divided into 2 groups,84 cases for experimental group and 28 cases for control group using CDAS 2.0 software. Experimental group were given Xuesaitong soft capsule and stimulation agent of Yinxingye soft capsule 2 pill at a time for 28 days,tid. RESULTS:100 patients including 75 cases for experimental group and 25 cases for control group had been completed experiment. There were statistical significance in difference between 2 groups in respect of improving half paralyzed symptoms,speech lost,dizziness and quarrel skew. Above aspects of experimental group were better than control group. CONCLUSION:Xuesaitong soft capsule have sound effect on blood stasis in stroke patients in recovery period.

AIM: To transfer 4 full-length WT1 isoforms cDNA into the leukemia cell line NB_4 so as to provide a cell model for studying the WT-1 gene function. METHODS: The eukaryotic expression recombinant vectors for WT1 isoforms (pCB6+/WT1) were introduced into the leukemia cell line NB_4 by electroporation. The positive cell clones were screened by G418 culture. The integration of WT1 gene isoforms in NB_4 cells as confirmed by PCR. The mRNA and protein of WT1 were detected by RT-PCR and Western blotting. RESULTS: WT1 gene isoforms were successfully transferred into NB_4 cells. WT1 mRNA and protein expression in the G418-selected cells increased remarkably compared with the control. CONCLUSION: WT1 gene isoforms were effectively transferred into NB_4 cells by electroporation and stably expressed in the transfected cells.

Objective To explore the gene sequencing and prenatal diagnosis of Glanzmann thrombasthenia (GT). Methods The blood samples were drawn from one case of phenotype GT pediatric patient, patient’s parents, and one normal control. The amniotic lfuid and cord blood from the fetus of patient’s mother were collected. When the fetus was born 2 days, the blood was drawn. The coagulation routine test and platelet aggregation test were performed. The expression of platelet membrane glycoprotein (GP) IIb and GPIIIa were tested by lfow cytometry. Microsatellite technology is used to determine whether fetal cord blood is contaminated with maternal cells. The expressed region and the junctional zone between exon and introns of GPIIb and GPIIIa were ampliifed by PCR technology from blood sample of patient, patient’s parents, and fetus’s cord and 2 days after birth. The PCR products were then subjected to DNA sequencing. Results Adenosine diphosphate (ADP) cannot induce the platelet aggregation in the patient. The max rate of the platelet aggregation in the fetus’s cord blood was half of the normal. However, the max aggregation rate induced by ADP in the blood sample of parents and fetus 2 days after birth were equal to normal. The mean lfuorescence intensity (MnX) of platelet membrane GPIIb and GPIIIa in the patient were 10%and nearly zero of the normal control, respectively, while those in the parents, the fetus’s cord blood and 2 days after birth were more than 90%and 30%to 50%of the normal control. The cast-off cells in amniotic lfuid and the DNA in cord blood analysis by microsatellite technology conifrmed that the amniotic lfuid and cord blood not contaminated by maternal cells. Gene analysis showed the heterozygosis mutation in exon6 A3829→C and exon9 G42186→A of the patient’s GPIIIa led to the amino acid heterozygosis mutation in GPIIIaHis281→Tyr and Cys400→Pro. These two mutations came from the father and the mother separately. However, there was only one heterozygosis mutation in exon9 G42186→A in the cast-off cells in amniotic lfuid, the fetus’s cord and blood 2 days after birth. Conclusion This GT patient have double heterozygosis mutation. The fetus has heterozygosis mutation conifrmed after birth.

Objective To discuss the mode of onset,clinical characteristics,treatment and prognosis of children with granulocyte sarcoma (GS),in order to provide guidance for early diagnosis and effective treatment of GS.Methods Six cases of children with GS diagnosed at the Department of Hematology,Children's Hospital Affiliated to Soochow University between June 2009 and June 2014 were analyzed,the data including the mode of onset,clinical manifestation,diagnosis,treatment and outcome.Results There were 2 cases with a painless mass onset (1 case was 2 years old,characterized by right waist mass,about 10 cm × 5 cm;the other case was 6 years old,characterized by axillary lump,about 2 cm × 3 cm),and both of them received surgical removal of the tumor,then the postoperative tumor was examined by pathologic and immunohistochemical method,and at last the primary granulocyte sarcoma was diagnosed.The third case was a 7 years old girl,she was onset characterized by scalp lump,about 2 cm × 3 cm,and was diagnosed by the pathologic and immunohistochemical method,and changes in hematological system appeared a month later and acute myeloid leukemia(AML) was confirmed by bone marrow examination.The onset ages of other 3 cases were in 10 months,1 year and 7 months,13 years and 3 months old respectively,characterized by scalp lump (about 2 cm × 3 cm),spinal canal tumor (about 1.0 cm × 1.5 cm),intracranial tumors (6.0 cm × 4.9 cm),with AML occurring at the same time,which was confirmed by surgical pathology,immunohistochemistry and bone marrow cell morphology,immune classification,chromosome,and fusion gene diagnosis.Four cases were hematopoietic malignancies by pathology,2 cases of then belonging to small round cell tumor.The immune pathology showed 5 cases of myeloperoxidase positive,CD68-positive,3 cases of CD43-positive,CD123-positive.All children CD3,CD20 levels in all children were negative.Four cases underwent surgery combined with chemotherapy,other 2 cases received surgery and then gave up treatment,1 case discontinued follow-up 3 months later,and the other case died of intracranial hemorrhage after 3 months,which induced by thrombocytopenia.The treated 4 cases were followed up 3 to 58 months,and all had disease-free survival.Conclusions Children with GS have low incidence and non-specific diagnostic criteria,its diagnosis depends on immune pathology,and the treatment is mainly in accordance with AML program for high-dose chemotherapy.The systematic chemotherapy helps to prolong overall survival;at the same time,the hematopoietic stem cell transplantation with bone marrow may help to improve the prognosis.

Objective To investigate the clinical features, brain imaging significance and the possible pathogenesis of posterior reversible encephalopathy syndrome (PRES) in childhood acute lymphoblastic leukemia (ALL) followed by chemotherapy induction.Methods The diagnosis and treatment of ALL were performed according to the guidelines of the Pediatric Association of Chinese Medical Association.There were 11 cases of pediatric ALL who developed PRES after chemotherapy induction.The clinical presentations, initial and follow-up radiologic features, and the neurologic outcomes of these 11 cases were investigated for one-year follow-up.All patients were reexamined 1,3,6, and 12 months after first imaging.Results Headache (10/1 1 cases), epileptic seizure (7/11 cases), high blood pressure (4/11 cases) ,visual impairment (6/11 cases) ,disturbance of consciousness (5/11 cases) and walking instability (2/11 cases) were the most common symptoms of these ALL patients with PRES.Magnetic resonance imaging (MRI) scanning revealed that lesions were mainly distributed in occipital lobe (9/11 cases), parietal lobe (8/11 cases), frontal lobe (5/11 cases) ,temporal lobe (3/11 cases), the deep white matter of bilateral periventricular and centrum semiovale (2/11 cases) and hemisphaerium cerebelli (1/11 cases).The radiological findings indicated that lesions had multifocal,symmetrical and posteriorly distributed characteristics in the cerebral hemispheres.After the diagnosis of PRES,patients stopped chemotherapy courses promptly and received symptomatic treatment, and then the clinical and imaging symptoms of most cases gradually disappeared.After 1-year follow-up,9 patients had good prognosis and no sequelae, 1 patient had symptomatic epilepsy (brain magnetic resonance imaging scan showed lesions in the left temporal lobe) ,and 1 patient had slight visual impairment.After the craniocerebral symptoms disappeared clinically ALL chemotherapy continued in all patients and no recurrent PRES was observed.Conclusions Although the clinical and imaging features of PRES may be diverse ,PRES should be recognized as a possible important complication of ALL when neurological symptoms appear.However, PRES is reversible when the patients are diagnosed and treated at an early stage.Thus,the occurrence of PRES should be considered and investigated to optimize the early induction schemes for ALL treatment.

Objective To research the expression,methylated regulation and clinical significances of miR-34b in chidren with acute leukemia(AL).Methods The methylation status of miR-34b promoter CpG islands were detected with methylation-specific polymerase chain reaction (MSP) in patients with AL.Then the expression of miR-34b was compared,which was detected by Taqman real-time fluorescence quantitative polymerase chain reaction (qRT-PCR),between the AL patients and normal group,in order to analyze their relationship with the clinical indicators.Resuits In 8 leukemia cell lines (U937,HL-60,MV4-11,M2R,K562,Raji,CCRF,DAMI) showed methylation,the positive rate of the methyl was 100％.Thirty-one acute lymphocytic leukemia(ALL) pediatric patients were newly diagnosed,and 24 cases showed methylation,the methylation positive rate 77.42％ (24/31).Nineteen acute myeloid leukemia(AML) patients were newly diagnosed,and 8 cases showed methylation,the methylation positive rate was 42.11％ (8/19 cases).There was no methylation in the 23 cases of normal children.The relative expression levels of miR-34b in the normal group,the group of leukemia cell lines,the group of ALL pediatric patients with newly diagnosed,the AML group and the group with mixed lineage leukemia gene rearrangement MLL+ were 5.22 ± 1.15,0.03 ± 0.03,1.65 ± 0.69,0.18 ± 0.06,0.64 ± 0.34,respectively.The findings indicated that there were significant differences in the relative expression levels of miR-34b between the normal group and the group of leukemia cell lines,the ALL group,the AML group,and the MLL+ group.The relative expression and methylated level of miR-34b had no statistically difference in gender,age at diagnosis,WBC count,chromosome,fusion gene,MLL gene rearrangement,and the minimal residual disease(LDH) levels in newly diagnosed AML patients(all P ＞ 0.05).And the relative expression and methylated level of miR-34b had no statistically difference in gender,age at diagnosis,WBC count,immunophenotype,chromosome,fusion gene,MLL gene rearrangement,TEL/AML1 gene,risk stratification,the minimal residual disease (MRD) in thirty-three days and the LDH levels in newly diagnosed ALL patients(all P ＞ 0.05).But as for the response to prednisone experiment,there was a significant difference between the sensitive group and the non sensitive group(P ＜ 0.05).Conclusions The expression level of miR-34b in AL was significantly lower and it was regulated by methylation mechanism,which implies that miR-34b may play a role of a tumor suppressor gene in the pathogenesis of leukemia.MiR-34b may affect the early treatment response of ALL patients,and it may be an indicator of risk stratification and poor prognosis in pediatric ALL.

Objective Multidrug resistance-associated protein 1 (MRP1) has been reported with a close correlation with tumor multi-drug resistance. Real-time quantitative PCR (QRT-PCR) was performed to detect the MRP1 gene expression in childhood acute lymphoblastic leukemia (ALL) and its clinical signiifcance was analyzed. Methods Sixty-seven denovo ALL patients and 10 healthier children as bone marrow donor were studied. The chemotherapy was given according to CCLG-2008 protocol. SPSS software was employed to analyze the data and p-value below 0.05 was regarded as statistic signiifcance. Results MRP1 expression level showed a close correlation with ALL risk, the median of MRP1 expression was 4.28 (2.75~6.12), 5.62 (4.99~8.60) and 7.56 (3.66~11.13) for standard-risk group (SR), intermediate-risk group (IR) and high-risk group (HR), respectively. MRP1 mRNA expression in T-ALL group was 7.71 (6.49~14.35), which is higher than that of B-ALL (5.18(3.89~8.46)) (P<0.01). The rate of leukemia cells’ sensitivity to prednisone on 7th day was 70.6%in high expression group (n=34), which was signiifcantly lower than that in low expression group (n=33, 90.9%, P=0.035). The complete remission rateon 33th day was 64.7%in high expression group, and 87.9%in low expression group, which showed a signiifcant difference between them (P=0.026). Conclusions In children ALL, the expression of MRP1 is closely related with immunophenotyping, treatment response, hazard level and disease relapse.

Objective To investigate the differential expression of miR-155 in newly diagnosed pediatric acute myeloid leukemia(AML) and its clinical significances.Methods Fifty-two AML children and 30 non-malignant disease matched children were recruited as the controls.The preliminary AML children were divided into favorable group,moderate group and poor group according to the National Comprehensive Cancer Network(NCCN) 2013.Real-time quantitative polymerase chain reaction was applied to validate the expressions of miR-155 in bone marrow samples (the data presented by 2-△△Ct).Results By comparing expressions of miR-155 between AML patients and controls,the miR-155 expressions were significantly higher in the AML children than those in the controls (Z =-5.391,P ＜ 0.001).There were significant differences among different prognostic groups,with a significantly lower level in the favorable group compared with others (x2 =12.586,P =0.002).It was also found that differential expressions existed not only in kinds of mutation cohort,with the highest level in FLT3-ITD and the lowest one in FLT3-TKD mutation group (x2 =11.216,P =0.024),but also among fusion gene subgroups (x2 =12.254,P =0.016),with the highest level in AML-ETO group and the lowest level in PML-RARa group:meanwhile,the expressions of miR-155 were statistic different according to French-America-British (FAB) subtypes (x2 =17.814,P =0.013),which was lower in M3 patients than non-M3 patients (Z =-3.291,P =0.001).Conclusions It indicates that the expressions of miR-155 may increase sharply in preliminary AML children,and the lower expression of miR-155 is closely related to favorable prognosis.

Objective To report two childhood acute myeloid leukemia (AML) patients with t(8;20)(q22;q13) and t(1;8;21)(q32;q22;q22) respectively,as variant t(8；21).Methods Chromosome preparation of bone marrow cells were made using short-term culture and karyotypic analysis was carried out using R and G-banding techniques.Interphase-fluorescence in situ hybridization (I-FISH) and metaphase-FISH (M-FISH) were performed using dual color,dual fusion AML1-ETO probe to detect the AML1-ETO fusion gene.Multiplex RT-PCR was used to demonstrate the expression of AML1-ETO fusion transcript.Results The karyotype of bone marrow cells for these two childhood AML patients were 45,X,-Y,t(8;20)(q22;q13)[12]/46,XY[3](case 1) and 46,XX,t(1;8;21)(q32；q22;q22)[18]/46,XX[2](case 2),respectively.I-FISH and M-FISH confirmed that they all had the AML1-ETO fusion gene and variant t(8;21).The AML1-ETO fusion transcript in both patients was detected by RT-PCR.Conclusion t (8;20)(q22;q13) and t (1;8;21)(q32;q22;q22) are variant t (8;21) in nature.It is important to combine the conventional karyotypic analysis with D-FISH and multiplex RT-PCR to determine the nature and prognosis of AML patients with variant t(8;21).