Correlation between vertebral bone marrow fat and abdomen fat: a prospective study / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the correlation between the lumbar bone marrow fat and abdominal fat.</p><p><b>METHODS</b>A total of 68 individuals (32 men and 36 women, aged 21-74 years with a median of 49.5 years) were included in this study. All the subjects underwent spectroscopic examination of the third lumber vertebra with the single voxel method on a 1.5T MR scanner to measure the fat fraction (FF%). Quantitative CT was also performed for measurement of the abdomen subcutaneous adipose tissue (SAT) and visceral adipose tissue (VAT). The measurements were compared between subjects aged ≥50 years and those below 50 years, respectively,in male or female subjects.</p><p><b>RESULTS</b>In male subjects, BMI, FF%, VAT or SAT showed no significant differences between the two age groups (P>0.05), and FF% was not correlated with BMI, VAT or SAT (r=0.109, 0.034, 0.066, respectively; P>0.05). In the female subjects, BMI, FF%, VAT and SAT differed significantly between the two age groups (P<0.05), and in ≥50 years group, FF% showed a positive correlation with VAT (r=0.499, P<0.05) but was not correlated with SAT (r=0.221, P>0.05); in<50 years group, FF% was not correlated with VAT or SAT (r=0.076, -0.067, respectively; P>0.05).</p><p><b>CONCLUSION</b>FF% is positively correlated with VAT in female subjects aged beyond 50 years, but is not correlated with VAT or SAT in male subjects or in younger female subjects.</p>

Chromosomal Aberrations in Myelodysplastic Syndromes / 中国实验血液学杂志

Myelodysplastic syndromes (MDS) are a group of clonal hematopioetic disorders characterized by myelodysplasia, decreased peripheral blood cells and high-risk of transformation into acute leukemia. MDS are often accompanied by a variety of chromosomal aberrations which play a role in disease pathogenesis, and are crucial in diagnosis and prognostic evaluation of this disease. About half of the patients with MDS have chromosomal abnormalities, mainly unbalanced chromosomal aberration. Different chromosomal aberration types are associated with different clinical outcome of this disease. Though balanced chromosomal translocations are not common in MDS, it seems that the patients with them have a higher leukemia transformation rate than those with other type of chromosomal aberrations. In this review, the chromosomal aberrations in MDS and their clinical significance for diagnosis and prognosis are briefly summarized.

Simultaneous determination of seven index components in Refined Coronary Tablet by HPLC / 中草药

Objective: To develop an HPLC method for the simultaneous determination of tanshinone IIA, paeoniflorin, salvianolic acid B, ferulic acid, safflor yellow A, ligustilide, and danshensu in Refined Coronary Tablets. Methods: The chromatographic separation was achieved on a Zorbax Eclipse XDB-C18 column (250 mm × 4.6 mm, 5.0 μm) with methanol-acetonitrile (25∶75, A)-0.1% formic acid (B) as mobile phases at the flow rate of 1.0 mL/min for gradient elution: 0-10.0 min, 95% B; 10.0-16.0 min, 95%-85% B; 16.0-18.0 min, 85% B; 18.0-22.0 min, 85%-75% B; 22.0-26.0 min, 75%-65% B; 26.0-40.0 min, 65%-15% B; Detection with variable wavelength: 0-19.0 min was 270 nm, 19.0-22.0 min was 230 nm, 22.0-27.0 min was 320 nm, 27.0-40.0 min was 402 nm, and the column temperature was 30 ℃. Its linear relationship, precision, repeatability, stability, and recoveries were investigated. Results: The results showed that the seven active components were well separated and showed good linearity, tanshinone IIA 0.4-8.0 mg/L (r = 0.999 5), paeoniflorin 1.2-24.0 mg/L (r = 0.999 1), salvianolic acid B 3.2-64.0 mg/L (r = 0.999 3), ferulic acid 0.08-1.60 mg/L (r = 0.999 5), safflor yellow A 1.2-24.0 mg/L (r = 0.999 3), ligustilide 0.24-4.80 mg/L (r = 0.999 7), and danshensu 0.32-6.40 mg/L (r = 0.999 7). The precision was good, and RSD was less than 2.0%. The repeatability was good, and RSD was less than 2.0%. The stability was good in 12 h. The average recoveries were between 98.05%-101.27%, and RSD was less than 2.0%. The contents of target components in Refined Coronary Tablets, tanshinone was 0.704-0.797 mg/g, paeoniflorin was 3.124-3.411 mg/g, salvianolic acid B was 7.129-7.611 mg/g, ferulic acid was 0.180-0.198 mg/g, safflor yellow A was 2.718-2.966 mg/g, ligustilide was 0.590-0.683 mg/g, and danshensu was 0.811-0.899 mg/g. Conclusion: The method is accurate, sensitive, credible, and repeatable. It can be applied to the quality control of Refined Coronary Tablets.

JAK2V617F mutation and TNF-α expression in myeloproliferative neoplasms and their correlation / 中国实验血液学杂志

This study was aimed to explore the JAK2V617F mutation and TNF-α expression in patients with myeloproliferative neoplasm (MPN), and the relation between them so as to provide theoretical basis for clinical practice and target therapy. Sixty-two confirmed BCR-ABL-negative MPN patients and 15 healthy adults were enrolled in this study. The peripheral blood mononuclear cells of the patients and healthy controls were divided into two parts, one part was used to extract DNA, the other one was used to extract mRNA and reverse-transcribe into cDNA. Real-time fluorescent quantitative PCR was used to detect JAK2V617F mutation proportion and the expression level of TNF-α. The results showed that the positive rate of JAK2V617F mutation in MPN patients was 64.52% (40/62) , including 54.28% in essential thrombocythemia (ET) patients (19/35), 94.74% in polycythemia vera (PV) patients (18/19) and 37.50% in myelofibrosis (MF) (3/8) patients. Mutation proportions of JAK2V617F in ET, PV and MF patients were 0.838 ± 0.419, 4.417 ± 0.658, 2.746 ± 2.009 respectively. The expression of TNF-α in ET, PV and MF patients were higher than that in healthy controls: 1.7, 7.0, 8.2-fold (P < 0.05) respectively. In addition, TNF-α expression was correlated with JAK2V617F allele burden (Pearson r = 0.610,R(2) = 0.372,P = 0.005). It is concluded that TNF-α plays an important role in the pathogenesis of MPN, the TNF-α expression increases and is different in ET,PV and MF patients,which correlates with JAK2V617F allele burden.

Detection of chromosome abnormalities in myelodysplastic syndromes with interval fluorescence in situ hybridization / 中国实验血液学杂志

This study was aimed to investigate the value of interval fluorescence in situ hybridization (FISH) in detection of abnormal karyotypes of patients with myelodysplastic syndromes (MDS). Conventional cytogenetics (CC) and interval FISH methods were carried out to analyze the bone marrow cells in 80 cases of MDS and 20 normal people. The results showed that using FISH, 53.8% cases of MDS (43/80) were found with abnormal karyotypes which was higher than 21.3% detected by CC method. There was significant difference between the 2 methods in detecting abnormal karyotypes in MDS (P < 0.05). Among all World Health Organization (WHO) subtypes, more chromosome abnormal were detected by FISH than by CC, especially for refractory anemia (RA) and refractory cytopenia with multilineage dysplasia (RCMD) groups. The detecting rate in patients with intermediate risk of International Prognostic Scoring System (IPSS) also had a statistical difference between FISH and CC methods. It is concluded that the FISH is more sensitive than CC in detection of abnormal karyotypes in MDS and is informative for the cases with karyotype failure or normal karyotype tested by CC. It is mainly embodied in the intermediate risk cases of IPSS. In addition, patients with RA and RCMD may benefit more from FISH for diagnosis compared with other WHO subtypes.

Effect of erlotinib on proliferation and differentiation of JAK2V617F-positive cells in vitro / 中国实验血液学杂志

The aim of this study was to investigate the effect of erlotinib on proliferation and differentiation of JAK2V617F-positive cells in vitro, and to provide experimental evidence of erlotinib for potential target therapy in polycythemia vera. Colony forming assays were used to detect the effect of erlotinib on differentiation of hematopoietic progenitor cells from bone marrow of polycythemia vera patients, and MTT method was used to measure the proliferation of HEL cell line containing the JAK2V617F mutation. The results showed that erlotinib 5 µmol/L inhibited the differentiation of JAK2V617F-positive hematopoietic progenitor cells into hematopoietic colonies in vitro, while it had almost no effect on normal hematopoietic progenitor cells from the patients. Erlotinib had inhibitory effect on the proliferation of HEL cell line in a dose dependent manner. The IC(50) was 4.1 µmol/L. It is concluded that erlotinib can inhibit proliferation and differentiation of JAK2V617F-positive cells to a certain extent in vitro.

Inhibitory effect of gefitinib and lapatinib on proliferation of HEL cells / 中国实验血液学杂志

This study was aimed to investigate the therapeutic effect of two molecular targeted therapeutic drugs, tyrosine kinase inhibitors gefitinib and lapatinib, on JAK2 V617F positive myeloproliferative disorders (MPD). The human leukemia cell line (HEL cell line) carrying JAK2 V617F mutation was treated with gefitinib (0.5, 1, 5, 10, 25 µmol/L) and lapatinib (0.5, 1, 2, 4, 8, 16 µmol/L) respectively. MTT method was used to detect HEL cell proliferation. The apoptotic rate and cell cycle were measured by flow cytometry. The results showed that gefitinib could significantly inhibit the proliferation of HEL cells in a dose-dependent manner, it's correlation coefficients for 24 and 48 h were 0.991 and 0.895 respectively. IC(50) at 48 h was 5.4 µmol/L. Gefitinib could effectively induce apoptosis of HEL cells in a dose-dependent manner (r = 0.896). Otherwise, gefitinib could arrest HEL cells at G(0)/G(1) phase. The inhibitory effect of lapatinib was less than gefitinib, it's IC(50) of inhibiting proliferation of HEL cells was 19.6 µmol/L. It is concluded that both gefitinib and lapatinib can inhibit the proliferation of HEL cells. These two tyrosine kinase inhibitors can be used for researching of targeted therapy of JAK2 V617 positive MPD.

JAK2V617F mutation and p-STAT5 protein expression in peripheral blood cells of patients with myeloproliferative neoplasm and their relations with clinical features / 中国实验血液学杂志

This study was aimed to explore the JAK2V617F mutation and p-STAT5 expression in patients with myeloproliferative neoplasm (MPN), and investigate their relations with clinical characteristics so as to provide theoretical basis for clinical practice and target therapy. Forty-five confirmed BCR-ABL-negative MPN patients and 15 healthy adults were enrolled in this study. Real-time fluorescent quantitative PCR and Western blot were respectively used to detect JAK2V617F mutation proportion and p-STAT5 expression level. In addition, their relations with clinical characteristics of MPN were analyzed. The results showed that the positive rate of JAK2V617F mutation in MPN patients was 73.3% (33/45), including 83.3% in polycythemia vera (PV) patients (20/24), 68.8% in essential thrombocythemia (ET) patients (11/16) and 40.0% in idiopathic myelofibrosis (IMF) patients (2/5). Mutation proportions of JAK2V617F in PV, ET and IMF patients were 0.472 ± 0.245, 0.216 ± 0.162, 0.435 ± 0.239 respectively; gray values of p-STAT5 protein in PV, ET and IMF patients were 1.396 ± 0.758, 0.760 ± 0.623, 0.792 ± 0.612 respectively. JAK2V617F mutation proportion and p-STAT5 protein expression level showed a linear correlation (P < 0.05). PV patients with higher JAK2V617F mutation proportion had higher white blood cell count, hemoglobin level and hematocrit, but lower platelet count; ET patients with higher mutation proportion showed older and higher white blood cell count, hemoglobin level and hematocrit, there was no significant difference between platelet count; IMF patients with higher JAK2V617F mutation proportion showed lower white blood cell count, platelet count, hemoglobin level and hematocrit. Patients with JAK2V617F positive mutation were more likely complicated by splenomegaly, bleeding and thrombotic events. It is concluded that the incidence rate of JAK2V617F mutation is high in patients with MPN. Higher mutation proportion always connected with higher expression of p-STAT5, and easily complicates by splenomegaly and thrombotic events.

Progress of study on JAK2V617F mutation in myeloproliferative neoplasm / 中国实验血液学杂志

Myeloproliferative neoplasms (MPN) are clonal hematopoietic stem cell diseases characterized by proliferation of one or more myeloid cell lineages in the bone marrow and increased mature and immature cells in peripheral blood. As the most important discovery in recent studies of MPN, JAk2V617F mutation is considered to closely relate with the pathogenesis of MPN. The mutated JAK2 lost self-inhibition, and then, the sustained activation leads to a series of disorders in downstream signal transduction pathways, eventually resulting in malignant cell proliferation. A variety of methods have been used in quantitative/qualitative detection of JAK2V617F mutation, and researches about JAK2V617F mutation and its clinical features have also made some progress. However, it must be noted that there are still some unsolved problems, such as the role of JAk2V617F mutation in pathogenesis of MPN needs further exploration, effective targeted therapy for JAK2 is a attractive topic, and the application of JAK2V617F mutation in disease diagnosis also requires a deep research. In this review, the latest progress from different aspects is summarized briefly, including JAK2 and JAK2V617F mutation, effects of JAK2V617F mutation on the pathogenesis, clinical correlation of JAK2V617F with MPN, and targeting therapy.

Diagnostic significance of Th1/Th2 cytokine pattern in childhood hemophagocytic lymphohistiocytosis / 中华儿科杂志

<p><b>OBJECTIVE</b>To illustrate the diagnostic value of Th1/Th2 cytokine pattern in childhood hemophagocytic lymphohistiocytosis (HLH) and its diagnostic accuracy.</p><p><b>METHOD</b>The BD(TM) CBA Human Th1/Th2 Cytokine Kit II was used to measure the serum Th1 and Th2 cytokines, including Interferon-gamma (IFN-γ), tumor necrosis factor (TNF), interleukin (IL)-10, IL-6, IL-4 and IL-2 in 50 patients with de novo HLH admitted to our hospital from Oct. 2005 to Aug. 2009. The above cytokine levels were also determined in 250 healthy volunteers and 235 patients with sepsis as controls.</p><p><b>RESULT</b>The primary features of these patients were prolonged high-grade fever (50/50), hepatomegaly (44/50), splenomegaly (38/50), hemocytopenia (47/50), hyperferritinemia (49/50), coagulopathy (44/50), hemophagocytosis in bone marrow (42/50), liver dysfunction (42/50) and hypertriglyceridemia (42/50). The IFN-γ, TNF, IL-10, IL-6, IL-4 and IL-2 levels for healthy children were (4.6 ± 1.8) ng/L, (4.0 ± 1.2) ng/L, (6.5 ± 1.3) ng/L, (6.0 ± 1.5) ng/L, (2.9 ± 0.8) ng/L and (2.6 ± 0.7) ng/L, while the median levels of them in acute phase of HLH children were 1138.5 (49.2 - 5000.0) ng/L, 3.4 (1.0 - 25.1) ng/L, 740.5 (26.5 - 5000.0) ng/L, 66.1 (3.9 - 4472.6) ng/L, 3.9 (1.0-32.8) ng/L and 4.0 (1.0 - 51.1) ng/L, respectively. The cytokine levels decreased to 9.1 (1.9 - 180.1) ng/L, 2.9 (1.0 - 11.0) ng/L, 11.4 (2.9 - 184.2) ng/L, 6.5 (1.0 - 44.8) ng/L, 2.7 (1.0 - 6.5) ng/L and 4.1 (1.0 - 12.0) ng/L respectively after remission. The IFN-γ, IL-10 and IL-6 levels in acute phase were significantly higher than those after remission and those of the healthy control (P all < 0.001). IL-4, IL-2 and TNF slightly elevated or at normal range in acute phase of HLH. The patients with sepsis showed a different cytokine pattern, with an extremely high level of IL-6 (median: 251.3 ng/L, range: 8.4- > 5000.0 ng/L) and moderately elevated level of IL-10 (median: 46.5 ng/L, range: 3.1 - 5000.0 ng/L), whereas IFN-γ was only slightly elevated (median: 9.2 ng/L, range: 1.3 - 498.8 ng/L). When the criteria for HLH set as the following: IFN-γ > 100 ng/L, IL-10 > 60 ng/L and the concentration of IFN-γ higher than that of IL-6, the specificity reached as high as 98.7% and the sensitivity was 88.0% for the diagnosis of HLH among patients with HLH and sepsis. Meanwhile, the positive predictive value (PPV) and negative predictive value (NPV) could reach 93.6% and 97.5%, respectively.</p><p><b>CONCLUSION</b>The significant increase of IFN-γ and IL-10 with slightly increased level of IL-6 is a sensitive and specific cytokine pattern for childhood HLH, which is helpful for its diagnosis and differential diagnosis.</p>

Involvement of CD4+CD25+ regulatory T cells in the pathogenesis of polycythaemia vera / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Regulatory T cells (T(reg)) have been shown to play an important role in the regulation of hematopoietic activity. However, there is no information about the effect of T(reg) cells in the pathogenesis of polycythaemia vera (PV).</p><p><b>METHODS</b>In this study, we investigated the percentage and function of T(reg) cells in the peripheral blood of 21 PV patients and 25 healthy donors. T(reg) cells were identified and characterized as CD4+CD25+ FOXP3+ by flow cytometry. The suppressive activity of CD4+CD25+ T(reg) cells was assessed by the proliferation and cytokine secretion of the co-cultured CD4+CD25- fractions.</p><p><b>RESULTS</b>The results showed that the percentage of T(reg) cells in the peripheral blood of PV patients significantly increased compared to healthy controls ((10.93 +/- 4.02)% vs (5.86 +/- 1.99)%, P < 0.05). Moreover, the mRNA and protein expression of FOXP3 was higher in CD4+CD25+ T(reg) cells. Coordinately, when co-cultured with the activated CD4+CD25- cells, the CD4+CD25+ T(reg) cells showed enhanced suppressive function in PV. Yet, the underlying mechanism for the increased frequency and function of CD4+CD25+ T(reg) cells is still to be clarified.</p><p><b>CONCLUSION</b>T(reg) cells expansion might account for the abnormal T cell immunity in PV patients and thus contribute to the pathogenesis of PV.</p>

Long-term follow-up of treatment outcome and prognosis on 46 children with acute promyelocytic leukemia / 中国当代儿科杂志

<p><b>OBJECTIVE</b>Acute promyelocytic leukemia (APL) is a specific type of hematopoietic malignancy, accounting for 10% of the de novo acute myeloid leukemia (AML). The data on long-term outcome of APL in children are limited. The aim of this study was to investigate the clinical biological features, diagnosis, prognosis and long-term survival of childhood APL.</p><p><b>METHODS</b>A total of 46 children with newly diagnosed APL from April 1998 to October 2005 were enrolled into this study. Induction treatment containing all-trans retinoic acid (ATRA) plus daunorubicin (DNR) or pirarubicin (THP) was performed on these patients, followed by 6 courses of chemotherapy consolidation: DNR, homoharringtonine or etoposide plus Ara-C. A maintenance therapy was then administered once 3-6 months. The total period of treatment was 2.5 years.</p><p><b>RESULTS</b>Of the 39 patients who had completed the regular treatment, 36 (92.3%) achieved a complete remission. The 5-year cumulative incidence of relapse (CIR) was 28.6%. The estimated overall survival (OS) rates at 1, 3 and 5 years were (86.1 +/- 5.8)%, (76.1 +/- 7.5)% and (70.2 +/- 8.9)% respectively, while the event free survival (EFS) rates were (78.4 +/- 6.8)%, (63.6 +/- 8.7)% and (53.1 +/- 10.0)% respectively. The 5-year OS rate of patients with WBC less than or equal to 10.0 X 10(9)/L was (81.4 +/- 10.3)%, which was significantly higher than that with WBC greater than 10.0 X 10(9)/L[(51.6 +/- 14.7)%, P < 0.05]. Five patients with RT-PCR positive for PML/RARalpha S (short) subtype died eventually although all of them achieved CR, but none of the 13 patients with PML/RARalpha L (long) subtype died.</p><p><b>CONCLUSIONS</b>Remission induction therapy with ATRA + DNR or THP is effective and safe for newly diagnosed childhood APL. The remission induction therapy combined with chemotherapy containing high/intermediate dose Ara-C can improve the long-term survival rates of APL patients. High WBC count and S subtype of PML-RARa are two poor prognostic factors for children with APL.</p>

Application of denaturing high performance liquid chromatography to mutation detection of the c-kit gene in mastocytosis / 中国实验血液学杂志

The aim of study was to detect mutation of tyrosine domain of c-kit gene in mastocytosis using denaturing high performance liquid chromatography technique, and to investigate the significance of this gene mutation in diagnosis and therapy of mastocytosis. Genomic DNA was obtained from bone marrow or peripheral blood leukocytes using the phenol/chloroform method from 7 mastocytosis patients. PCR was performed with AmpliTaq Gold DNA polymerase and 100 ng genomic DNA to amplify the entire coding sequence and exon-intron boundaries of c-kit exon 17 approximately exon 19. Denaturing high-performance liquid chromatography (DHPLC) analysis was performed on a WAVE DNA Fragment Analysis System. Each PCR product was mixed with an equal quantity of amplified human placental DNA (served as normal control) and was denatured at 95 degrees C for 5 minutes, followed by slowly cooling down to room temperature by 1.5 degrees C per minute to allow heteroduplexes formation. All the conditions for the DHPLC analysis, including melting temperature and buffer gradients were determined using the Transgenomic software Navigator. Samples with extra peaks or with different peak form on DHPLC were directly sequenced using the BigDye Terminator Cycle Sequencing Reaction kit and ABI 3100 Genetics Analyser. The results showed that DHPLC revealed an aberrant peak in one patient in exon 17 and the D816V mutation was identified by direct sequencing. The other two patients had an extra peak for exon 18/19 and direct sequencing revealed a conservative sequence change (L862L) within exon 18. It is concluded that denaturing high performance liquid chromatography is a high efficiency and reliable technique for mutation detection of c-kit gene and the detection results would be helpful for the selection of therapy in mastocytosis.