Qualitative identification of Aucklandiae Radix with LLE + SMA classification models based on electronic nose / 中草药

Objective: To establish an odor fingerprint with electronic nose technology to qualitatively identify Aucklandiae Radix odor from different producing areas. Methods: Eight batches of Aucklandiae Radix samples from eight different producing areas were collected. The odor information of each sample was obtained by electronic nose. The LDA algorithm based on Fisher’s identification criterion and the nonlinear dimensionality reduction LLE + SMA algorithm were used to distinguish the Aucklandiae Radix odor of different origins. Results: It was found that the LDA algorithm based on Fisher’s discriminant criterion could not distinguish the Aucklandiae Radix scent of different producing areas. Some of the samples in the place of origin had a lot of overlap, and the LLE + SMA algorithm could distinguish the odor very well. It can completely distinguish eight batches of Aucklandiae Radix samples from eight different producing areas. Conclusion: It is feasible to apply the electronic nose technology to the odor differentiation of Aucklandiae Radix from different producing areas, and provide new ideas and methods for the quality evaluation of Aucklandiae Radix.

Quality evaluation of Aucklandiae Radix based on gray correlation method and FCM algorithm / 中草药

Objective To comprehensively evaluate the Aucklandiae Radix quality by combining gray correlation method and FCM (Fuzzy C-Means) algorithm. Methods HPLC method was used to determine the content of costunolide and dehydrocostus, and the content of volatile oil was determined by steam distillation. The gray correlation method was used to sort the quality of Chinese herbal medicines, and the FCM algorithm was used to classify the quality of medicines. Results The average correlation degree of Aucklandiae Radix from different producing areas was: Yunnan > Guangdong > Guangxi > Hunan> Sichuan > Bozhou > Beijing > Hebei. FCM algorithm divided samples into three categories: Yunnan, Guangdong and Guangxi were in the high quality group; Hunan and Sichuan were in the medium quality group; Beijing, Bozhou, and Hebei were in the low quality group. Conclusion Constructing an integrated pattern recognition model system for evaluating the quality of Chinese medicinal materials, and the fuzzy clustering algorithm was adopted for the first time in Aucklandiae Radix. The purpose of this study is to form a kind of research method of Aucklandiae Radix quality evaluation and provide a way to guide and apply the new methods of modern pattern recognition and data mining in the application of traditional Chinese medicine quality evaluation.

Effect of Early Ambulation after Radiofrequency Catheter Ablation via Femoral Vein Approach / 中国康复理论与实践

Objective To investigate the effect of early ambulation on the comfort and complications in patients undergoing radiofre-quency catheter ablation via femoral vein approach. Methods From April,2016 to April,2017,120 patients were randomly assigned into control group(n=40),group A(n=40)and group B(n=40).The control group received the routine care of remaining bed rest for eight hours with the affected leg immobilized at least four hours.Group A ambulated after four hours of the affected leg immobili-zation,and group B changed their position after ablation and ambulated after four hours of bed rest.They were assessed with the General Comfort Questionnaire,Numerical Rating Scale of back pain and State Anxiety Inven-tory.Their vascular complications were recorded. Results The incidence of bleeding and hematoma was not significantly different among the groups(P>0.05).Group B was more comfortable than the control group(P<0.05).Compared with the control group,groups A and B were less in back pain intensity(F>10.376,P<0.001),and had less anxiety(t=3.278,P<0.05),less incidence of limb numbness, difficulty falling asleep, and loss of appetite and dysuria (χ2>6.409, P<0.05). Group B was the best among them. Conclusion There is little risk of early physical activity and ambulation in the vascular complications after radiofrequen-cy catheter ablation via femoral vein approach,which may reduce their discomfort.

Clinical Significance of Corrected Serum Calcium in 320 Patients with Multiple Myeloma / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To analyze the clinical significance of corrected serum calcium(CSCa) in patients with multiple myeloma.</p><p><b>METHODS</b>The serum calcium levels of 320 patients with initial multiple myeloma were measured and corrected by serum albumin and its levels measured simultaneously. The differences of serum calcium levels were analyzed before and after the correction by serum albumin.</p><p><b>RESULTS</b>There was a significant difference between serum calcium and CSCa in MM patients (2.34±0.15 vs 2.6±0.17 mmol/L). The constituent ratio of patients with hypercalcemia was from 11.3% to 23.1% after correction, the MM patients with hypocalcemia was decreased from 42.8% to 7.8% after correction, and the patients with normal calcium level were increased. There was a significant difference between serum calcium level and CSCa in I, II, III stages of MM patients respectively(P<0.05). In the 320 patients, the incidence of anemia was 80%, renal failure was 20.9%, and myeloma bone disease was 68.8%. Calcium concentration in both anemia and renal insufficiency was higher than the normal group, and the difference was more significant after correction. In 220 cases of MM receiving chemotherapy, the median progression-free survival (PFS) was 15 months, and overall survival(OS) time was 20 months. The PFS and OS time of the patients with hypercalcemia were shortened, and the difference was very significant after correction(P<0.01).</p><p><b>CONCLUSION</b>Corrected serum calcium can more sensitively to reflect the diseases serious extent, thus indicating prognosis has better effect.</p>

Growth Inhibition of Multiple Myeloma Cells Caused by MicroRNA-15a and Its Mechanisms / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To explore the effect and possible mechanisms of miR-15a on growth of multiple myeloma(MM) cells.</p><p><b>METHODS</b>MM cell lines (U266 and RPMI8226) were transfected by lentiviral particles. MM stable cell lines were selected and collected by flow cytometry (FCM). Proliferation of MM cells before and after miR-15a high expression was detected by CCK-8 method. Apoptosis of MM cells before and after miR-15a high expression was detected by AO/EB dying, Hoechst 33258 dying and FCM, respectively. Cell cycle of MM cells before and after miR-15a high expression was detected by FCM. The expressions of miR-15a, BMI-1 and BCL-2 mRNA of MM cells before and after miR-15a high expression were detected by real-time PCR. The expressions of BMI-1 protein of MM cells before and after miR-15a high expression were detected by Western blot.</p><p><b>RESULTS</b>MM stable cell lines with miR-15a high expression was acquired. CCK-8 result showed that high expression of miR-15a could inhibit growth of MM cells (U266 and RPMI8226). AO/EB dying, Hoechst 33258 dying and FCM testing results showed that high expression of miR-15a could significantly induce apoptosis of MM cells (U266 and RPMI8226). The apoptosis rates of U266 and RPMI8226 cells in high expression group and control group were 90.52% vs 37.08% and 59.40% vs 44.17%, respectively. Meanwhile, FCM testing results showed that high expression of miR-15a could induce G1 arrest of MM cells (U266 and RPMI8226), which proportion of G1 phase were 41.50%±0.64%, 45.31%±0.77%, respectively. Real-time PCR results showed that during the growth inhibition process of MM cells caused by miR-15a high expression, the expression of BCL-2 mRNA decreased, but there was no significant changes in the expression of BMI-1 mRNA, while the expression of BMI-1 protein decreased significantly.</p><p><b>CONCLUSION</b>High expression of miR-15a can induce cell cycle arrest and apoptosis of MM cells, then inhibit their growth. The mechanisms may be related with the negative regulation of BMI-1 and BCL-2 genes in post-transcription level caused by miR-15a.</p>

Anti-leukemia mechanism of miR-17 and miR-20a silencing mediated by miRNA sponge / 中国实验血液学杂志

This study was aimed to quantitatively detect the expression levels of pre-miR-17 and pre-miR-20a in acute leukemia patients and eight kinds of leukemia cell lines, and to investigate the anti-leukemia mechanism of miR-17 and miR-20a silence mediated by miRNA Sponge. Quantitative real-time PCR was used to detect the mRNA expression levels of pre-miR-17 and pre-miR-20a in patients with various types of leukemia and leukemia cell lines. The Jurkat cells over-expressing miR-17 and miR-20a were transfected with recombinant lentivirus-transfecting units targeted at miR-17 and miR-20a plus 6 µg/ml of polybrene. Then the proliferation ability and cell cycle of Jurkat cells was evaluated by CCK-8 and flow cytometry respectively. The results showed that the expression level of pre-miR-17 and pre-miR-20a in all leukemia patients was significantly higher than that in normal group(P < 0.05), the expression of pre-miR-17 and pre-miR-20a in acute lymphoid leukemia was significantly higher than that in acute myeloid leukemia(P < 0.05), and the pre-miR-17 and pre-miR-20a expression level did not correlate significantly with high white blood cell count>20.0×10(9)/L(P > 0.05). The miR-17 and miR-20a silencing mediated by miRNA Sponge led to a significant decrease of cell growth, restored G1 accumulation and increase of cell apoptosis. It is concluded that the expression of miR-17 and miR-20a is upregulated in leukemia patients, which may contribute to leukemogenesis. Over-expressed miR-17 and miR-20a promote cell growth and cell cycle progression, and inhibit apoptosis through negatively-regulating P21 and E2F1 after-transcriptionally.

Expression of MCL-1 and microRNA-29a in extranodal NK/T-cell lymphoma tissue / 中国实验血液学杂志

This study was aimed to explore the expression level and correlation of MCL-1 and miR-29a in extranodal NK/T-cell lymphoma (ENKTCL) tissue. Maxvision immunohistochemistry technique and real time fluorescent quantitative PCR were used to detect the expression level of MCL-1 and miR-29a in tissue of 20 patients with ENKTCL and 10 patients with proliferative lymphadenitis, respectively. The results showed that the expression of MCL-1 protein were higher in patients with ENKTCL than that in patients with proliferative lymphadenitis, but there were no significant correlation between MCL-1 overexpression and age, sex, Ann Arbor stage and International Prognostic Index (IPI), respectively. Correlation analysis indicated that there was significant negative correlation between miR-29a expression and MCL-1 expression (r = -0.59, P = 0.016). It is concluded that miR-29a may target MCL-1 gene, regulate its expression, then participate in tumorigenesis and development of ENKTCL.

Construction of miRNA sponge targeting miR-20a and stable expression in Jurkat leukemia cell line / 中国实验血液学杂志

This study was aimed to construct miRNA sponge targeting miR-20a and to establish a stable cell line Jurkat-S, paving the way for further research on function of miR-20a and application of RNAi in gene therapy. One pair of two-repeated oligonucleotide sequences containing bulged sites that are mispaired opposite miR-20a positions 9-12 was designed and synthesized with enzyme cutting sites. The annealed oligonucleotide fragments were subcloned into pCDNA3.0-L expressing vector. After double-enzyme cutting, the vector was ligated to the annealed oligonucleotide fragments again. Enzyme cutting and luciferase activity assay were performed for identification after four repeats. Then the ligated fragment was subcloned to lentivirus expressing vector. Virus particles were collected after the control or sponge vectors were co-transfected with the psPAX2 packaging plasmid and the envelope plasmid pMD2.G into HEK-293T cells using Lipofectamine 2000. The Jurkat cells were transfused with recombinant lentivirus-transfusing units plus 6 µg/ml of Polybrene. Real-time PCR and Western blot were used to detect the mRNA and protein expression of P21 and E2F1 after lentivirus transfusion respectively. As a result, luciferase activity assay demonstrated that the sponge targeting miR-20a was constructed successfully and the virus was packaged in 293T. The titer of virus was 5×10(7) TU/ml. Stable transfected Jurkat-S cell line was established. As was expected, the mRNA and protein level of P21 and E2F1 was upregulated significantly in Jurkat-S cells. It is concluded that the miR-20a sponge is constructed successfully, and Jurkat-S stable cell line is established, in which the expression of miR-20a is inhibited stably.

Establishment of a U266 cell line with stable Bmi-1 silencing by lentivirus-mediated RNA interference / 中国实验血液学杂志

This study was aimed to construct lentivirus-mediated shRNA expression vector targeting Bmi-1 and establish a stable cell line U266-li, so as to pave the way for further research on function of Bmi-1 and application of shRNA to gene therapy. One pair of oligonucleotide sequences targeted at human Bmi-1 mRNA were designed and synthesized. The annealed oligonucleotide fragments were subcloned into pLVTHM vector. Virus particles were collected after the control or shRNA vectors were co-transfected with the psPAX2 packaging plasmid and the plasmid pMD2.G was enveloped into HEK-293T cells by using Lipofectamine2000. The U266 cells were transduced with 5 × 10(6) recombinant lentivirus-transducing units plus 6 µg/ml of polybrene. Real-time PCR and Western blot were used respectively to detect the expression of Bmi-1 and P14 after lentivirus transduction. DNA sequencing demonstrated that the lentivirus RNAi vector of Bmi-1 was constructed successfully and the virus was packaged in 293T cells. The titer of virus was 5 × 10(7) TU/ml. Stable transfected U266 cell line was established. As was expected, the mRNA and protein levels of Bmi-1 was reduced significantly in U266 cells after lentivirus transduction, whereas the mRNA and protein levels of P14 was upregulated. It is concluded that the lentiviral RNAi vector of Bmi-1 is constructed, and U266 stable cell line is established.

Mechanism associated to enhancing the sensitivity of myeloma cells U266 to bortezomib by 2-methoxyestradiol / 中国实验血液学杂志

This study was aimed to explore the synergistic effect of 2-methoxyestradiol (2-ME2) and bortezomib (Bor) on the proliferative inhibition and apoptosis of U266 cell line and its possible mechanism. The cells were treated with 2-ME2, Bor alone and 2-ME2 combined with Bor, respectively. The cell viability and proliferative curve were detected by CCK8, the cell apoptosis was detected by caspase 3/7 activity test, cell cycle status was analyzed by flow cytometry, and real-time PCR was used to detect the mRNA expression of P21, BAX and BCL-2. The results showed that compared with cells treated with 2-ME2 or Bor alone, the proliferative potential of cells in combination group was significantly inhibited (p < 0.05), and apoptosis rate markedly increased (p < 0.05), cell cycle was arrested at G(1)-S phase, the mRNA expressive level of P21 and BAX increased, while the expression of BCL-2 decreased. It is concluded that 2-ME2 combined with Bor synergistically inhibits cell proliferation and induces apoptosis in U266 cell line. The possible mechanism may be associated with its effect of up-regulating P21 and BAX expressions.

Content determination of epigoitrin in Radix Isatidis and its preparation by RP-HPLC / 中国中药杂志

<p><b>OBJECTIVE</b>To establish an HPLC method for the content determination of epigoitrin in Radix Isatidis and its preparation, and to provide valuable data for quality control of Radix Isatidis and its preparation.</p><p><b>METHOD</b>The samples were separated on a ZORBAX SB-C18 (4. 6 mm x 150 mm, 5 microm) column with the mobile phase of acetonitrile-water-phosphoric acid-triethylamine (8.50 : 90.72 : 0.73 : 0.05) in the flow rate of 0.7 mL x min(-1). The detection wavelength was set at 245 nm. Column temperature was 30 degrees C.</p><p><b>RESULT</b>The linear range of epigoitrin was 0.0204-0.3060 microg (r = 0.9998), and the average recovery was 98.99% with the RSD was 1.31% (n = 9).</p><p><b>CONCLUSION</b>The method for quantitation of epigoitrin in Radix Isatidis and its preparation was accurate and reliable, which can be used to evaluate the quality of Radix Isatidis and its preparation.</p>