Identification and characterization of flavonoid 3-O-glycosyltransferase gene CtUF3GT from safflower (Carthamus tinctorius L.) / 药学学报

UDP-glucose: flavonoid 3-O-glucosyltransferase (UF3GT) uses flavones, dihydroflavonol or anthocyanin as the acceptor and uridine 5′-diphosphate-sugar as the donor to catalyze the production of flavonoid 3-O-glycoside compounds. Based on sequence homology and transcriptome data, we screened and cloned a UF3GT gene named CtUF3GT (GenBank No. OM948976) from safflower. Biological information analysis demonstrate that CtUF3GT has highly conserved PSPG motif. The open reading frame of CtUF3GT is 1 446 bp, encoding 481 amino acids, with a presumed molecular weight of 52.36 kD and a theoretical isoelectric point of 5.33. Multiple sequence alignment indicate that CtUF3GT has a high homology with UF3GT from Asteraceae, and phylogenetic analysis showed that CtUF3GT clusters with functional identified UF3GTs from other species. The purified recombinant protein glucosylated kaempferol and quercetin to biosynthesis of kaempferol 3-O-glucoside and quercetin 3-O-glucoside, respectively. And CtUF3GT prefered to use kaempferol as substrate. qRT-PCR analysis showed that the UF3GT gene was most highly expressed in flowers, followed by leaves, with very low expression in bracts and stems, and no expression in roots. The expression of UF3GT gene showed a trend of increasing and then decreasing at different stages of flower development. The expression of CtUF3GT gene in safflower with different flower color was highly significant (P < 0.01) at S1, S2, S5, S6 and S7 stages of flower development, in which the expression of CtUF3GT in white safflower was 5.3 and 3.1 times higher than that in red safflower at S6 and S7 stages. This study lays the foundation for further exploring the role of CtUF3GT in the mechanism of safflower flavonoid secondary metabolite biosynthesis and accumulation.

Effects of Prostaglandin E Receptor 4 Agonist on the Expression of Stem Factors in Human CD34 Cells / 中国实验血液学杂志

OBJECTIVE@#To investigate the effect of prostaglandin E recoptor 4 antagonist (EPA) on the self-renewal ability of human CD34 cells and its mechamism.@*METHODS@#The peripheral blood hematopoietic stem cell of 20 healthy donors received the G-CSF-mobilization were collected, then the human CD34 cells were sorted out by MACS microbead kit. The human CD34 cells were treated with DMSO (control group), EPA (EPA group) and EPA+EPA antagonist (EPA+EPA group) for 72 hours. The differential genes and pathways related with CD34 cell stemness were detected by Thermogram and Pathway enrichment analysis. and then the expression levels of protein and gene (β-catenin, Nanog, Oct4, Sox2, Stat3, AKT, P38) were detected by qRT-PCR and Western blot respectively.@*RESULTS@#EPA could elevate the mRNA and protein expression of β-catenin, Nanog, Oct4, Sox2, in comparison with control group, however, mRNA and protein expression of STAT3, AKT, P38 were not changed. When human CD34 cell were cultured with EPA+XAV939 it was found that the mRNA and protein expression of β-catenin was downregulated, moreover the mRNA and protein expression of Nanog, Oct4, Sox2 were reduced.@*CONCLUSION@#EPA can upregulate stemness factors-β-catenin, Nanog, Oct4 and Sox2 in human CD34 cell in vitro, but not STAT3, AKT and P38.

Segmented cloning and expression of the pigment epithelium derived factor and its effect on the proliferation of human SCL-1 cells / 医学研究生学报

Objective Studies are rarely reported on the effect of short peptides of the pigment epithelium derived factor (PEDF) on the proliferation of human cutaneous squamous (SCL-1) cells. The purpose of this study is to investigate segmented cloning and expression of the PEDF protein and observe its effect on the proliferation of human SCL-1 cells. Methods The target genes of PEDF1, PEDF2 and PEDF3 were amplified by PCR and the recovered fragments subjected to double digestion of NheⅠ and Hind Ⅲ and inserted into the pET28a(+) plasmid. The product was transformed into human E coli BL21 and induced to express, followed by isolation and purification of the fusion protein. CCK-8 assay was used to detect the proliferation of the SCL-1 cells with PEDF1, PEDF2 and PEDF3 at 100, 400, 800 and 1000 nmol/L at 24, 48 and 72 hours. Results The prokaryotic expression vectors of PEDF1, PEDF2 and PEDF3 were successfully constructed, and their fusion proteins prepared, with the molecular weight of 18 000, 17 000 and 13 000, respectively. The proliferation of the SCL-1 cells was significantly decreased in the 800 and 1000 nmol/L PEDF3 groups compared with that in the 0 nmol/L PEDF3 group at 24 hours (0.16 ± 0.03 and 0.78 ± 0.07 vs 1.00 ± 0.00, P < 0.05), inhibited in a concentration-dependent manner in the 400, 800 and 1000 nmol/L PEDF3 groups at 48 hours (P < 0.05), markedly lower in the 800 and 1000 nmol/L PEDF3 groups at 72 hours (0.53 ± 0.05 and 0.51 ± 0.05) than in the in the 400 and 0 nmol/L PEDF3 groups (0.60 ± 0.05 and 1.00 ± 0.00) (P < 0.05). Conclusion The PEDF fusion proteins were successfully segmentally cloned and expressed and PEDF3 inhibited the proliferation of SCL-1 cells, which has paved the ground for further screening of active functional short peptides of PEDF.

Simultaneous determination of five constituents in Maxing Kanggan Granules by HPLC / 中成药

AIM To establish an HPLC method for the simultaneous content determination of five constituents in Maxing Kanggan Granules (Isatidis Radix,Houttuyniae Herba,Scutellariae Radix,etc.).METHODS The analysis of 70％ ethanol extract of this drug was performed on a 30 ℃ thermostatic Wondasil C18 column (250 mm × 4.6 mm,5 μm),with the mobile phase comprising of acetonitrile-0.1％ phosphoric acid flowing at 1.0 mL/min in a gradient elution manner,and the detection wavelengths were set at 245,327,280 and 250 nm.RESULTS (R,S)-goitrin,chlorogenic acid,baicalin,baicalein and glycyrrhizic acid showed good linear relationships within the ranges of 0.24-6.05 μg/mL (r =0.999 9),0.70-17.40 μg/mL (r =0.999 9),28.59-714.70 μg/mL (r =0.999 6),1.67-41.65 μg/mL (r =0.999 5) and 2.59-64.80 μg/mL (r =0.999 7),whose average recoveries were 99.53％,99.41％,98.66％,98.31％ and 99.25％ with the RSDs of 1.02％,1.05％,1.26％,0.85％ and 1.43％,respectively.CONCLUSION This simple,accurate and reproducible method can be used for the quality control of Maxing Kanggan Granules.

Investigation of 1q21 amplification in patients with multiple myeloma using I-FISH and cIg-FISH / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To investigate the prevlance of 1q21 amplification in patients with multiple myeloma (MM) and its correlation with the progression and prognosis of the disease.</p><p><b>METHODS</b>1q21 amplification was detected in 48 patients with MM using cytoplasmic light chain immunofluorescence with fluorescence in situ hybridization analysis (cIg-FISH) and interphase fluorescence in situ hybridization (I-FISH) analysis combined with CD138 immunomagnetic cell sorting (MACS).</p><p><b>RESULTS</b>1q21 amplification (≥ 3 red signals) was detected in 26/48(54.2%) cases by cIg-FISH and 31/48 (64.6%) cases by I-FISH combined with CD138 MACS. There was a good consistency between the two methods (P>0.05). The mortality of patients with 1q21 amplification was significantly higher than those without (P< 0.05). No significant difference was detected in terms of sex, age, Durie-Salmon stage, subgroup and international staging system (ISS) stage between patients with 1q21 amplification and those without (P>0.05).</p><p><b>CONCLUSION</b>The frequency of 1q21 amplification in MM is high. There was also an association between the amplification and poor prognosis. cIg-FISH is consistent with CD138 MACS combined with I-FISH.</p>

MicroRNA and multiple myeloma / 中国实验血液学杂志

MicroRNA (miRNA) represents a new class of endogenous, non-coding small RNA molecules that functions as gene regulator. They can repress the expression of target genes at the post-transcriptional level by base pairing to the 3'-untranslated region of the respective target mRNAs. miRNA plays important roles in the regulation of cell proliferation, metabolism, apoptosis and differentiation. The dysregulation of miRNA expression contributes to tumorigenesis. This review summarizes recent progress of research on the characteristics and function of miRNA, as well as the role of miRNA in multiple myeloma development, abnormal karyotype in chromosomes and drug-resistant.

Investigation of chromosome 1 aberrations in patients with multiple myeloma using cIg-FISH method and its significance / 中华血液学杂志

<p><b>OBJECTIVE</b>To investigate the incidence of 1q21 amplification and 1p12 deletion, and analyze the correlation between these aberrations with disease progression, prognosis and outcome in patients with multiple myeloma (MM).</p><p><b>METHODS</b>Cytoplasm light chain immunofluorescence with simultaneous interphase fluorescence in situ hybridization (cIg-FISH) was used to detecte the 1q21 amplification and 1p12 deletion in 48 patients with MM.</p><p><b>RESULTS</b>1q21 amplification (≥ 3 red signals) was determined in 26 of 48(54.2%) cases. The mortality of patients with 1q21 amplification was significantly higher than that of those lacking 1q21 amplification (P < 0.05). The sex, age, D-S stage, subgroup and ISS stage between patients with and without 1q21 amplification had no significant difference (P > 0.05). There was a significant difference in D-S stage and mortality between patients with 3 and with 4 copies of 1q21 (P < 0.05). No significant difference in sex, age, subgroup, ISS stage, and isotype was found between them (P > 0.05). 1p12 deletion (< 2 green signals) was found in 14 of 48 (29.2%) cases. There was no significant difference in sex, age, D-S stage, ISS stage, isotype, subgroup, and mortality between patients with and without 1p12 deletion.</p><p><b>CONCLUSION</b>The frequency of chromosome 1 aberrations in multiple myeloma is high and 1q21 amplification is a poor prognosis factor.</p>

microRNA-21 and microRNA-30b expression in multiple myeloma / 中华血液学杂志

<p><b>OBJECTIVE</b>To investigate the expression of miR-21 and miR-30b in multiple myeloma (MM).</p><p><b>METHODS</b>Peripheral blood mononuclear cells from patients with MM were cultured at 2.5 x 10(6) cells/ml in alpha-MEM supplemented with 10% of fetal bovine serum, antibiotics, RANKL (50 ng/ml), and macrophage colony-stimulating factor (25 ng/ml) for 10 to 14 days to obtain osteoclasts with bone-resorbing activity. Primary myeloma cells were purified from 12 MM patients. Of them, 8 samples were cocultured with osteoclasts and 4 as noncocultured control. The expression of miR-21 and miR-30b was detected by real-time PCR.</p><p><b>RESULTS</b>The viability of MM cells recovered from cocultures was higher than those of noncocultured control. After cocultured with osteoclasts, primary myeloma cells from eight patients exhibited a 1.3- 5.9-fold increase in miR-21 expression and 1.38- 4.32-fold decrease in miR-30b expression compared with controls. In highly purified plasma cells from 3 healthy subjects, 12 MM patients and 11 MM cell lines, the expression of miR-21 was 1.9 +/- 0.8, 6.5 +/- 4.9 and 35.1 +/- 36.2, respectively; the expression of miR-30b was 13.6 +/- 1.8, 7.2 +/- 6.3 and 4.5 +/- 1.9, respectively.</p><p><b>CONCLUSIONS</b>miR-21 acts as an oncogene and miR-30b a tumor suppressor gene in MM.</p>

Progressive transformation of lymph node germinal centers: a case report and literature review / 中华血液学杂志

<p><b>OBJECTIVE</b>To improve the understanding of progressive transformation of lymph node germinal centers (PTGC) and to explore its clinical, histopathologic and immunohistochemical features and the differential diagnosis between the related disease of germinal center hyperplasia.</p><p><b>METHODS</b>The clinical manifestation, laboratory bindings, treatment and outcome of a patient with PTGC were presented.</p><p><b>RESULTS</b>The main manifestation of the patient was painless peripheral lymphadenopathy. Histopathologic examination of an axillary lymph node showed reactive follicular hyperplasia and the progressive transformation changes germinal centers. The borderline between the germinal center and the mantle layer was obscured. The cells in the progressive transforming germinal centers were positive for CD20(+), CD5(+), CDw75(+).</p><p><b>CONCLUSION</b>PTGC is a rare lymphoid disorder. Histopathology and immunohistochemistry are important basis of the diagnosis.</p>

Therapeutic efficacy of bortezomib-based chemotherapy on 40 patients with multiple myeloma / 中国实验血液学杂志

This study was aimed to investigate the therapeutic efficacy and adverse events of bortezomib-based chemotherapy for 40 patients with multiple myeloma. 16 newly diagnosed patients and 11 patients with refractory/relapse myeloma were treated with bortezomib, dexamethasone and thalidomide; 7 newly diagnosed patients and 4 patients with refractory/relapse myeloma were treated with bortezomib and dexamethasone; 2 newly diagnosed patients were treated with bortezomib, melphalan and thalidomide. Cycles were repeated every 28 or 35 days, all the patients were treated for 2 to 8 cycles. The therapeutic efficacy and adverse events were evaluated according to International Myeloma Working Group Uniform Response Criteria. The results indicated that the median follow-up duration was 13 months, the total response rate was 72.5%, among which 16 patients achieved complete response (CR), 13 achieved partial response (PR). The main side effects included gastrointestinal symptoms, peripheral neuropathy, thrombocytopenia, respiratory infection, herpes zoster and urinary retention and so on. The adverse events were ameliorated by treatment and decrease of the bortezomib dose. It is concluded that bortezomib-based chemotherapy is effective in the treatment of either newly diagnosed or refractory/relapse MM patients and the adverse events are tolerable and manageable for patients.

Investigation of the molecular changes in patients with multiple myeloma by fluorescence in situ hybridization / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To investigate the incidence and prognosis of 1q21 amplification, 13q14 deletion, TP53 gene deletion and IgH translocation in patients with multiple myeloma (MM).</p><p><b>METHODS</b>Interphase fluorescence in situ hybridization (I-FISH) with four different specific probes for the regions containing 1q21, 13q14.3 (D13S319), 14q32 and TP53 gene were performed in 43 MM patients.</p><p><b>RESULTS</b>Among the 43 MM patients, 1q21 amplification was observed in 28 (65.1%) cases, 13q14 deletion in 30 (69.7%) cases, TP53 gene deletion in 8 (18.6%) cases, and IgH translocation in 29 (67.4%) cases. The mortality of MM patients with 1q21 amplification, 13q14 deletion or TP53 gene deletion was higher than those without them.</p><p><b>CONCLUSION</b>There is high frequency of 1q21 amplification, 13q14 deletion, TP53 gene deletion and IgH translocation in multiple myeloma, and 1q21 amplification, 13q14 deletion and TP53 gene deletion are poor prognosis factors for MM patients.</p>

Autocrine effect of vascular endothelial growth factor on the proliferation of HaCaT cells / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To determine the autocrine effect of vascular endothelial growth factor (VEGF) on epidermal keratinocytes HaCaT cells.</p><p><b>METHODS</b>Cultured HaCaT cells were treated with various concentrations of VEGF(165) (0,1,5,10,25,50,100 ng/ml) or Avastin (0,0.063,0.125,0.25,0.50,1.0,2.0 mg/ml) in vitro. HaCaT cell proliferation was determined by MTT assay and the cell migration was measured by migration assay. The effect of VEGF(165) (10 ng/ml) on phosphorylation of ERK1/2 was detected in HaCaT cells pretreated or not pretreated with Avastin (0.5 mg/ml).</p><p><b>RESULTS</b>VEGF enhanced the proliferation and migration of HaCaT cells in a dose-dependent manner, while Avastin inhibited the effects of VEGF also in a dose-dependent manner. VEGF(165) (10 ng/ml) induced the phosphorylation of ERK1/2 in HaCaT cells,but which was blocked by Avastin (0.5 mg/ml).</p><p><b>CONCLUSION</b>VEGF enhanced the proliferation and migration of HaCaT cells in a dose-dependent manner, while Avastin inhibited the effects of VEGF also in a dose-dependent manner. VEGF(165) (10 ng/ml) induced the phosphorylation of ERK1/2 in HaCaT cells,but which was blocked by Avastin (0.5 mg/ml).</p>

Analysis of clinical factors related to genotype B and C chronic hepatitis B infection in Zhenjiang area / 中华传染病杂志

Objective To compare the differences of clinical characteristics between genotype B and C chronic hepatitis B(CHB)patients and to summarize clinical factors related to genotype C hepa- titis B virus(HBV)infection.Methods Seventy eight CHB patients who were diagnosed with genotype B or C infection by liver puncture biopsy and genotyping were enrolled.Their serum HBV DNA levels were detected.Severe hepatitis,liver cirrhosis,hepatocellular carcinoma and HBeAg positive rate were analyzed to determine the pathologic inflammation and fibrosis degree of liver tissue.Chi square test and Logistic multiple regression analysis were employed for the statistical analysis.Results The serum albumin and pre-protein were lower in genotype C CHB patients than that in genotype B.The alanine aminotrans- ferase,total bilirubin and prothrombin time were higher in genotype C CHB patients than that in genotype B.The rates of genotype C patients increased significantly with the grade of liver necroin- flammation progressing from GO to G4(1.8%,11.1%,20.4%,33.3%,33.3%) and the stage of liver fibrosis progressing from SO to S4(5.6%,5.6%,14.8%,33.3%,40.7%),but the rates of genotype B patients did not change significantly with the grade of liver necroinflammation(16.7%, 25.0%,25.0%,20.8%,12.5%)and stage of liver fibrosis progressing(16.7%,29.2%%,20.8%, 16.7%,16.7%).There was statistical significance in grades of liver necroinflammation(X~2= 11.49,P=0.022)and stages of liver fibrosis(X~2=13.56,P=0.006)between genotype B and gen- otype C patients.The rates of genotype C CHB patients were higher than,similar with and lower than the rates of genotype B patients of HBV DNA level above 1.0?10~6 copy/mL,between 5.0?10~2-1.0?10~6 copy/mL and under 5.0?10~2 copy/mL,respectively(51.8% vs 12.5%,35.2% vs 45.8% and 13.0% vs 41.7%).There was statistical significance of HBV loads between genotype B and genotype C patients(X~2=13.25,P=0.001).HBeAg positive rate in genotype C patients was significantly higher than that in genotype B patients(61.1% vs 25.0%,X~2=8.67,P=0.003).The rates of decompensated cirrhosis,compensated cirrhosis and no-cirrhosis in genotype C patients were higher than,similiar with and lower than the rates in genotype B patients,respectively(40.7% vs 4.2%,22.2% vs 20.8% and 37.0% vs 75.0%).There was statistical significance of the rate of cirrhosis between genotype B and genotype C patients (X~2=12.47,P=0.002).Conclusions The degree of liver necroinflammation and fibrosis,the HBeAg positive rate and the incidence of cirrhosis are all related with genotype C HBV infection.

The expression of CDld in systemic lupus erythematosus patients with antiphospholipid syndrome / 中华风湿病学杂志

Objective To investigate the association of CDId with APS and laboratory data,by examin- ing the CDld expression of PBMC from systemic lupus erythematosus(SLE)patients with antiphospholipid syndrome.Methods The CDld expression of 20 SLE patients with APS,30 SLE controls and 23 normal con- trois were detected by flow cytometry.The relationship between CDld expression and clinical,laboratory data of patients with SLE was analysed.Results The expression of CDld in PBMC of SLE patients with APS was higher than normal controls,but had no difference with SLE controls.The expression of CDld in monocyte group of SLE patients with APS was higher than normal controls,which had no difference with SLE controls. Highly significant correlations was found between CDld expression and SLEDAI while,negative correlation between CDld expression and complements was discovered.Conclusion The abnormal expression of CDld may play a role in the pathogenesis of SLE,it may be a parameter for assessing disease activity.