This study was purposed to investigate the mutational status of DNA methyltransferase (DNMT3a) gene and the clinical features of AML patients with DNMT3a mutations. Using PCR combined with directly sequencing, the somatic mutations of DNMT3a involving residue of amino acid 882 were detected in 77 AML patients. Furthermore, the clinical features of these patients were also studied. The results showed that the DNMT3a mutation were detected in 7 out of 59 patients with de novo AML (11.9%), which included 4 patients with DNMT3a R882C, 2 patients with DNMT3a R882H and 1 patient with DNMT3a Y874C. Morphology examination indicated that 2 patients were M(2), 1 patient was M(4) and 4 patients were M(5). Cytogenetic analysis revealed that karyotype in 5 out of 7 patients with DNMT3a mutation were normal. In total of 27 patients with normal karyotype 5 patients (22.7%) were found harboring DNMT3a mutation, while no DNMT3a mutation was found in 21 patients with abnormal karyotype. The mutation rate in patients with positive CEBPA was obviously higher than that in patients with negative CEBPA (p = 0.002). Immunophenotype analysis showed that 4 patients (4/7, 57.1%) with DNMT3a mutation expressed lymphoid antigens including CD4 or/and CD7. There were no statistical significance in age, gender, blast cells of bone marrow, white blood cell and platelet counts, hemoglobin level, ratio of CR, mutations of FLT3-ITD, NPM1 and c-kit between patients with DNMT3a mutation and patients with wild DNMT3a (p > 0.05). It is concluded that the DNMT3a mutations are more prevalent in AML patients with normal karyotype accompanying with positive NPM1 and/or CEBPA mutation, the role of DNMT3a mutation in AML prognosis needs to be further studied.
Adolescent ; Adult ; Aged ; Aged, 80 and over ; CCAAT-Enhancer-Binding Proteins ; genetics ; Child ; DNA (Cytosine-5-)-Methyltransferases ; genetics ; Female ; Humans ; Leukemia, Myeloid, Acute ; genetics ; Male ; Middle Aged ; Mutation ; Nuclear Proteins ; genetics ; Young Adult

OBJECTIVETo evaluate the clinical significance of the application of fluorescence in situ hybridization (FISH) in detecting chronic myeloid leukemia (CML).

METHODSChromosome preparation was made by using 24-hour culture. FISH technique using dual color dual fusion (DC-DF) BCR/ABL probe was performed in all 158 cases and R-banding was also employed for karyotyping in some patients.

RESULTSAmong the 158 cases, 98 cases were Ph positive, of which 69 cases (70.4%) were typical FISH pattern (1R1G2F), the other 29 cases (29.6%) showed 12 different types of atypical FISH pattern. The most frequent atypical patterns found were 1R1G1F in 7 cases (7.1%), 2R1G1F in 5 cases (5.1%), 1R1G2F and 1R1G3F in 4 cases (4.1%), 2R2G1F in 3 cases (3.1%). Karyotype analysis on 18 CML cases with atypical FISH patterns demonstrated that the atypical FISH patterns were due to variant translocation in 3 cases; the additional third signal was because of a supernumerary Ph chromosome. The karyotyping results did not conform to FISH results in four cases suggesting the conceivable mistakes in karyotyping. The 1R1G1F signal pattern seen in 3 cases with classical t(9;22) resulted from the deletion of derivative chromosome 9. The 1R1G2F signal pattern detected in 40% to 64% of interphase cells of 3 cases without Ph chromosome by conventional cytogenetic analysis suggested a submicroscopic translocation. Three cases treated with Glivec or bone marrow transplantation showed normal karyotypes with a small amount of BCR/ABL positive cells by FISH detection.

CONCLUSIONFISH technique is of great value for the diagnosis of CML and confirmation of variant translocation, occult Ph translocation, derivative chromosome 9 deletion, therapeutic effect of interferon and Glivec as well as detection of minimal residual disease after bone marrow transplantation.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Biomarkers, Tumor ; analysis ; Chromosome Banding ; Chromosome Deletion ; Chromosomes, Human, Pair 22 ; Chromosomes, Human, Pair 9 ; Female ; Fusion Proteins, bcr-abl ; Gene Deletion ; Humans ; In Situ Hybridization, Fluorescence ; methods ; Karyotyping ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; diagnosis ; genetics ; Male ; Middle Aged ; Translocation, Genetic ; Young Adult

OBJECTIVETo evaluate the genetic constitution and mutation status of the immunoglobulin variable heavy chain region (IGVH) gene expression in Chinese patients with chronic lymphocytic leukemia (CLL).

METHODSThe IGVH mutation was detected by multiplex PCR and direct sequencing of the purified PCR products from 64 CLL patients. The segments of VH, DH and JH family and mutations were analyzed by IMGT/V-QUEST and IGBlast.

RESULTSIn the 64 patients, the most common usage was VH3 (31/64, 48%), followed by VH4 (26/64, 41%), VH1 (4/64, 6%), VH2 (2/64, 3%) and VH7 (1/64, 2%). The results also showed that 44 patients (69%) had mutated VH, 6 cases (9%) had identical germline sequences. Among the 64 sequences of DH segments, DH3 gene family was used most frequently (25/64, 39%), among which 11 cases had unmutated VH. The most frequent usage of the JH segments was JH6.

CONCLUSIONThere is significant difference in the frequency of the IGVH gene family in Chinese CLL patients compared to Western patients, suggesting the involvement of antigen selection in different ethnic and/or environmental factors in CLL disease initiation, and its prognostic significance needs further investigation.

Adult ; Aged ; Aged, 80 and over ; Asian Continental Ancestry Group ; genetics ; Female ; Gene Expression ; Humans ; Immunoglobulin Class Switching ; genetics ; Immunoglobulin Heavy Chains ; genetics ; Immunoglobulin Variable Region ; genetics ; Leukemia, Lymphocytic, Chronic, B-Cell ; genetics ; immunology ; Male ; Middle Aged ; Mutation

OBJECTIVETo investigate the incidence of Philadelphia chromosome (Ph) in adult B-lineage acute lymphoblastic leukemia (B-ALL).

METHODSOne hundred and twelve adult patients with previously untreated B-ALL were prospectively investigated by interphase dual-color dual-fusion fluorescence in situ hybridization (DD-FISH) with two-color break apart probe BCR-ABL and the results were compared with that of conventional cytogenetics (CC).

RESULTSThe incidence of Ph chromosome was 17.98% (16/89) and 31.25% (35/112) by CC and DD-FISH, respectively. The mean positive rate of Ph+cells by FISH was 66.23% (ranging 18.5%-99%). Of the 35 Ph+ALL patients by FISH, 25 were successfully karyotyped by CC which included 5 normal karyotypes, 20 abnormal karyotypes including 16 Ph chromosome and 13 complex abnormalities.

CONCLUSIONThe incidence of Ph chromosome was 31.25% in adult with B-ALL. DD-FISH with BCR-ABL probe provides a powerful technique for the diagnosis of Ph+B-ALL. It is an important supplement to the CC analysis. DD-FISH technique should be used as a routine method for the diagnosis for adult acute B-ALL.

Adolescent ; Adult ; Aged ; B-Lymphocytes ; metabolism ; pathology ; Chromosome Aberrations ; Color ; Female ; Humans ; In Situ Hybridization, Fluorescence ; methods ; Interphase ; Karyotyping ; Male ; Middle Aged ; Philadelphia Chromosome ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; genetics ; pathology

In order to evaluate the cytogenetic features and clinical significance of chronic myeloid leukemia (CML), chromosome preparation of bone marrow cells was made by using 24-hour culture, and R-banding technique was employed for karyotyping in 362 patients with CML. The patients were divided into two groups of chronic phase (CP) and blast crisis (BC). The results showed that the incidence of additional chromosome, variant translocation and Philadelphia (Ph) negative, bcr/abl positive CML with abnormal chromosomes in CP group were 70 cases (26.1%), 19 cases (7.1%), 4 cases (1.5%), and were 50 cases (53.2%), 8 cases (8.5%), 4 cases (4.3%) in BC group. Among the 362 cases, 324 cases (89.5%) were Ph positive. Classic translocation was found in 297 cases (91.7%) and variant translocation in 27 cases (8.3%), including 13 cases of simple variant, 13 cases of complex variant and 1 case of marked Ph. Special karyotypes were found in 120 out of 362 cases. Analysis of these karyotypes demonstrated that the most common numerical abnormalities were +Ph (21.7%), +8 (10.0%), +21 (10.0%), +19 (7.5%) and structure abnormalities were i(17q) (13.3%). In conclusion, compared to chronic phase, the incidence of additional chromosome, variant translocation and so on are much higher at in blast crisis. It is feasible to evaluate the progress of the disease by karyotype analysis.
Adolescent ; Adult ; Aged ; Aged, 80 and over ; Child ; Female ; Humans ; Karyotyping ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; genetics ; Male ; Middle Aged ; Philadelphia Chromosome ; Young Adult

This study was aimed to investigate the characteristics of chromosome karyotype abnormality in patients with acute myeloid leukemia. 379 cases of de novo acute myeloid leukemia were enrolled in this study. Chromosome preparations were made on bone marrow cells by using direct method or short-term culture. Chromosome karyotypes were analyzed by R-banding technique. The results indicated that 216 out of 379 patients had clonal chromosome aberrations with the percentage of 56.99%, including 19 kinds of balanced translocations and 70 kinds of chromosome gain or loss. The most common structure and numerical abnormalities were t(15;17) and -Y with the percentage of 25.86% and 5.80%, respectively. -Y was accompanied by t(8;21) in 90.9% of the -Y abnormality cases, which accounted for 40.81% of t(8;21) positive cases. The abnormality of M(3) was significantly higher than the other FAB subtypes (p < 0.05). No significance was found between the male and female groups for the chromosome aberrations (p > 0.05). In conclusion, some specific chromosome aberrations are correlated with specific FAB subtype, which may contribute to the clinical diagnosis and subtyping of the disease.
Adolescent ; Adult ; Aged ; Aged, 80 and over ; Child ; Child, Preschool ; Chromosome Aberrations ; Chromosome Disorders ; genetics ; Female ; Humans ; Karyotyping ; Leukemia, Myeloid, Acute ; genetics ; Male ; Middle Aged ; Young Adult

The aim of this study was to explore cytogenetic characteristics of acute promyelocytic leukemia (APL) and compare the interphase fluorescence in situ hybridization (I-FISH) with conventional cytogenetic (CC) analysis. A total number of 157 APL patients were recruited in this study, and the I-FISH and CC were applied to analyze cytogenetic features. Chromosome samples of bone marrow cells were prepared by short-term culture. Out of all 157 cases, 136 were observed with CC assay, 66 with I-FISH, of which 45 samples were analyzed with both methods. The results showed that among all 136 CC samples, t(15;17)(q22;q21) was found in 120 cases, of which 107 cases was isolated t(15;17)(q22;q21) abnormality, 13 cases was complex abnormalities and 12 case without mitotic figure. Among all 66 cases of I-FISH group, PMI/RARα fusion gene was found in 64 cases (97.0%), suggesting that I-FISH group was more sensitive than CC group (p = 0.041). It is concluded that combination of I-FISH and CC techniques plays a pivotal role for diagnosis and detection of minimal residual disease in APL.
Adolescent ; Adult ; Aged ; Aged, 80 and over ; Child ; Child, Preschool ; Cytogenetic Analysis ; methods ; Female ; Humans ; In Situ Hybridization, Fluorescence ; methods ; Infant ; Karyotyping ; Leukemia, Promyelocytic, Acute ; diagnosis ; genetics ; Male ; Middle Aged ; Young Adult

Although the etiology of inflammatory bowel disease is still uncertain, increasing evidence indicates that the excessive activation of NLRP3 inflammasome plays a major role. Norisoboldine (NOR), an alkaloid isolated from Radix Linderae, has previously been demonstrated to inhibit inflammation and IL-1β production. The present study was to examine the effect of NOR on colitis and the underlying mechanism related to NLRP3 inflammasome activation. Our results showed that NOR alleviated colitis symptom in mice induced by 2, 4, 6-trinitrobenzene sulfonic acid (TNBS). Moreover, it significantly reduced expressions of cleaved IL-1β, NLRP3 and cleaved Caspase-1 but not ASC in colons of mice. In THP-1 cells, NOR suppressed the expressions of NLRP3, cleaved Caspase-1 and cleaved IL-1β but not ASC induced by lipopolysaccharide (LPS) and adenosine triphosphate (ATP). Furthermore, NOR could activate aryl hydrocarbon receptor (AhR) in THP-1 cells, inducing CYP1A1 mRNA expression, and promoting dissociation of AhR/HSP90 complexes, association of AhR and ARNT, AhR nuclear translocation, XRE reporter activity and binding activity of AhR/ARNT/XRE. Both siAhR and α-naphthoflavone (α-NF) markedly diminished the inhibition of NOR on NLRP3 inflammasome activation. In addition, NOR elevated Nrf2 level and reduced ROS level in LPS- and ATP-stimulated THP-1 cells, which was reversed by either siAhR or α-NF treatment. Finally, correlations between activation of AhR and attenuation of colitis, inhibition of NLRP3 inflammasome activation and up-regulation of Nrf2 level in colons were validated in mice with TNBS-induced colitis. Taken together, NOR ameliorated TNBS-induced colitis in mice through inhibiting NLRP3 inflammasome activation via regulating AhR/Nrf2/ROS signaling pathway.
Alkaloids ; administration & dosage ; Animals ; Colitis ; chemically induced ; drug therapy ; genetics ; immunology ; Drugs, Chinese Herbal ; administration & dosage ; Humans ; Inflammasomes ; drug effects ; immunology ; Interleukin-1beta ; genetics ; immunology ; Lindera ; chemistry ; Male ; Mice ; Mice, Inbred BALB C ; NF-kappa B ; genetics ; immunology ; Receptors, Aryl Hydrocarbon ; agonists ; genetics ; metabolism ; Trinitrobenzenesulfonic Acid ; adverse effects

BACKGROUND: Single subcortical infarction (SSI) is caused by two main etiological subtypes, which are branch atheromatous disease (BAD) and cerebral small vessel disease (CSVD)-related SSI. We applied the Beijing version of the Montreal Cognitive Assessment (MoCA-BJ), the Shape Trail Test (STT), and the Stroop Color and Word Test (SCWT) to investigate the differences in cognitive performance between these two subtypes of SSI. METHODS: Patients with acute SSIs were prospectively enrolled. The differences of MoCA-BJ, STT, and SCWT between the BAD group and CSVD-related SSI group were analyzed. A generalized linear model was used to analyze the associations between SSI patients with different etiological mechanisms and cognitive function. We investigated the correlations between MoCA-BJ, STT, and SCWT using Spearman's correlation analysis and established cut-off scores for Shape Trail Test A (STT-A) and STT-B to identify cognitive impairment in patients with SSI. RESULTS: This study enrolled a total of 106 patients, including 49 and 57 patients with BAD and CSVD-related SSI, respectively. The BAD group performances were worse than those of the CSVD-related SSI group for STT-A (83 [60.5-120.0] vs. 68 [49.0-86.5], P = 0.01), STT-B (204 [151.5-294.5] vs. 153 [126.5-212.5], P = 0.015), and the number of correct answers on Stroop-C (46 [41-49] vs. 49 [45-50], P = 0.035). After adjusting for age, years of education, National Institutes of Health Stroke Scale and lesion location, the performance of SSI patients with different etiological mechanisms still differed significantly for STT-A and STT-B. CONCLUSIONS BAD patients were more likely to perform worse than CSVD-related SSI patients in the domains of language, attention, executive function, and memory. The mechanism of cognitive impairment after BAD remains unclear.
Cerebral Infarction ; Cerebral Small Vessel Diseases ; Cognitive Dysfunction/etiology* ; Executive Function ; Humans ; Mental Status and Dementia Tests

This study investigated the mechanism of Danggui Shaoyao Powder(DSP) against mitophagy in rat model of Alzheimer's disease(AD) induced by streptozotocin(STZ) based on PTEN induced putative kinase 1(PINK1)-Parkin signaling pathway. The AD rat model was established by injecting STZ into the lateral ventricle, and the rats were divided into normal group, model group, DSP low-dose group(12 g·kg~(-1)·d~(-1)), DSP medium-dose group(24 g·kg~(-1)·d~(-1)), and DSP high-dose group(36 g·kg~(-1)·d~(-1)). Morris water maze test was used to detect the learning and memory function of the rats, and transmission electron microscopy and immunofluorescence were employed to detect mitophagy. The protein expression levels of PINK1, Parkin, LC3BⅠ/LC3BⅡ, and p62 were assayed by Western blot. Compared with the normal group, the model group showed a significant decrease in the learning and memory function(P<0.01), reduced protein expression of PINK1 and Parkin(P<0.05), increased protein expression of LC3BⅠ/LC3BⅡ and p62(P<0.05), and decreased occurrence of mitophagy(P<0.01). Compared with the model group, the DSP medium-and high-dose groups notably improved the learning and memory ability of AD rats, which mainly manifested as shortened escape latency, leng-thened time in target quadrants and elevated number of crossing the platform(P<0.05 or P<0.01), remarkably activated mitophagy(P<0.05), up-regulated the protein expression of PINK1 and Parkin, and down-regulated the protein expression of LC3BⅠ/LC3BⅡ and p62(P<0.05 or P<0.01). These results demonstrated that DSP might promote mitophagy mediated by PINK1-Parkin pathway to remove damaged mitochondria and improve mitochondrial function, thereby exerting a neuroprotective effect.
Rats ; Animals ; Mitophagy ; Alzheimer Disease/genetics* ; Powders ; Protein Kinases/metabolism* ; Ubiquitin-Protein Ligases/metabolism*