Agapetesheana, a new species of A.ser.Longifiles (Ericaceae) from Yunnan, China.

Agapetesheana Y. H. Tong & J. D. Ya (Ericaceae), a new species from Lüchun Xian, Yunnan Province, China is described and illustrated. This new species is assigned to Agapetessect.Agapetesser.Longifiles Airy Shaw. It is closest to A.inopinata Airy Shaw and A.oblonga Craib, but differs in having bead-like tubers, leaf blade with a wholly serrulate margin, subulate and much longer calyx lobes, much larger corollas that are carmine, green at the apex and maroon on angles, and longer stamens without spurs on the back.

Chinese guidelines for treatment of adult primary immune thrombocytopenia.

Primary immune thrombocytopenia (ITP) is a bleeding disorder commonly encountered in clinical practice. The International Working Group (IWG) on ITP has published several landmark papers on terminology, definitions, outcome criteria, bleeding assessment, diagnosis, and management of ITP. The Chinese consensus reports for diagnosis and management of adult ITP have been updated to the 4th edition. Based on current consensus positions and new emerging clinical evidence, the thrombosis and hemostasis group of the Chinese Society of Hematology issued Chinese guidelines for management of adult ITP, which aim to provide evidence-based recommendations for clinical decision making.

[Determination of six active ingredients in different parts of Belamcanda chinensis and Iris tectorum and their anti-inflammatory activity].

In order to explore the anti-inflammatory activity and active ingredient basis from the leaves of the Belamcanda chinensis and Iris tectorum, we established an HPLC method for simultaneous determination of six anti-inflammatory active ingredient contents in the root of the B. chinensis and I. tectorum as well as their leaves with different dry methods, and the anti-inflammatory effects of the extract were studied by the mouse ear swelling experiment. The HPLC analysis was performed on an Agilent WondaSil© C18-WR(4.6 mm×250 mm，5 µm)，with isocratic elution of acetonitrile-0.1% ortho-phosphoric acid solution at a flow rate of 1. 0 mL·min⁻¹ and the detection was carried out at 265 nm. The chemical compositions of the B. chinensis and I. tectorum are similar but the contents of them are obviously different. Both rhizome and leaf extract of B. chinensis and I. tectorum had inhibitory effects on inflamed mice induced by dimethylbenzene and had anti-inflammatory effects by animal experiment, which could lay the material and active foundation for the development of the non-medicinal parts of the B. chinensis and I. tectorum.

[Changes of Neutrophil CD64 in Patients with Hematological Mali-gnancies Combined with Bacterial Infections].

OBJECTIVE: To explore the changes of CD64 on surface of neutrophils in patients with hematological malignancies combined with bacterial infections. METHODS: Ninety-seven patients with hematological tumor admitted in our hospital from August 2014 to February 2016 were selected and divided into 2 groups: infection group(50 cases) and noninfection group(47 cases) according to their symptoms, physical sings and blood culture results for microbiologic detection. The CD46 index on surface of neutrophils, serum C- reactive protein (CRP), neutrophil count (NC) and procalcitonin (PCT) level were detected by flow cytometry. After treatment of infection patients, the CD46 index, CRP, PCT and NC were detected again. RESULTS: Before antibacterial treatment of patients, the CD64 index in infection group was significantly higher than that in noninfection group, the CRP, PCT and NC levels in infection group also were higher than those in noninfection group; after antibacterial treatment, the CD64 index, CRP, PCT and NC levels in infection group decresed. CONCLUSION: The CD64 index in hematologic malignancy patients complicated with bacterial infections significantly increased, after antibacterial treatment these indexes decreases, confirming the value of CD64 in the diagnosis of bacterial infections for patients with hematologic malignancies.

[Significance of Plasma HMGB1, IFN-γ, IL-4 and TLR4 Expression on CD4+T Cell Surface in Patients with Immune Thrombocytopenia].

OBJECTIVE: To investigate the effects of plasma HMGB1, IFN-γ, IL-4 and CD4+T cell surface TLR4 expression on the immune thrombocytopenia(ITP). METHODS: Twenty-five patients diagnosed as ITP in our hospital from October 2014 to October 2015, and 20 healthy persons as controls were selected. The ELISA was used to detect the plasma levels of HMGB1, IFN-γ and IL-4 in 2 groups; the flow cytometry was used to detect the expression level of TLR4 on the surface of CD4+ cells. The relatienship between different parameters was analyzed. RESULTS: Before treatment, the plasma HMGB1 level in ITP group was significantly higher than that in control group (t=4.259, P<0.01), while after treatment it significantly decreased and close to level in control group (t=1.267, P>0.05). The plasma IFN-γ level detected in ITP group before and after treatment was not significantly different from level in control group (P>0.05), while the IL-4 level in ITP group before treatment was significantly lower than that in control grup (t=2.107, P<0.05), but the IL-4 level in ITP group after treatment was significantly higher than that in control group (t=2.107, P<0.05). The plasma IFN-γ/IL-4 ratio in ITP group before treatment was significantly higher than that in control group (t=5.436, P<0.01), but it obviously decreased and was slightly lower than that in control group after treatment. The expression level of TLR4 in ITP group before and after treatment was all significantly lower than that in control group (P<0.01). The HMGB1 level in ITP group was directly proportional to the CD3+ content (r=0.824, P<0.01), however it not significantly related with CD4+ content (r=0.074, P>0.05), but than the HMGB1 level in ITP group was directly proportional to CD8+ content (r=0.844, P<0.01) and to IL-4 content (r=0.784, P<0.01), and was inversely proportional to IFN-γ level(r=-0.814,P<0.01),and also was inversely proportional to IFN-γ /IL-4 ratio(r=-0.887,P<0.01),and directly proportional to TLR4 expression level(r=0.772,P<0.01). CONCLUSION: The HMGB1 and TLR4 expression levels in ITP patients are higher, and clinical therapy can relieve the disease by targetly control in HMGB1 and TLR4 expression.

[Role of CD8(+)T cells and their secreted cytokines in the pathogenesis of aplastic anemia].

Aplastic anemia(AA) is mostly considered as an immune-mediated bone marrow failure syndrome, characterized by pancytopenia and bone marrow hypoplasia. The pathogenesis of AA is complicated, until now it is not fully understood. Further study on the pathological mechanism will be helpful for the diagnosis and treatment of AA. CD8(+) T cells and their secreted cytokines play important roles in the abnormal immunity during the process of AA. Thus, this review focuses on the role of CD8(+) T cells and their secreted cytokines in the pathogenesis of AA.

[Expression of IL-27 in multiple myeloma and its cell lines].

OBJECTIVE: To study the expression and significance of IL-27 in patients with multiple myeloma (MM) and in the supernatant of MM cell lines U266 and RPMI8226 cells culture medium. METHODS: A total of 20 MM patients and 20 controls were enrolled in this study. The expressions of IL-27 and IL-6 in MM patient's blood plasma, and the expression of IL-27 in U266 and RPMI8226 culture supernatant were measured by enzyme-linked immunosorbent assay (ELISA). The mRNA expression of IL-27 in mononuclear cells of MM patients' peripheral blood was measured by real-time quantitative polymerase chain reaction (RT-PCR). RESULTS: The levels of IL-27 in plasma of MM patients and normal controls were (61.82 ± 8.01) ng/L and (8.29 ± 4.41) ng/L (P<0.05), and those of IL-6 were (45.62 ± 1.24) ng/L and (2.27 ± 0.18) ng/L (P<0.05), respectively. The levels of IL-27 in U266 and RPMI8226 culture supernatant were (50.06 ± 5.72) ng/L and (335.47 ± 41.88) ng/L. RT-PCR revealed that the levels of IL-27 mRNA were up-regulated in MM patients compared to controls. CONCLUSION: High expression of IL-27 is observed in MM patients and MM cell lines U266 and RPMI8226 cells. IL-27 may play a important role in pathogenesis of MM.

Design and synthesis of pyrido[3,2-α]carbazole derivatives and their analogues as potent antitumour agents.

A series of pyrido[3,2-α]carbazole derivatives and their analogues have been prepared and evaluated for their antitumour activity against human lung cancer A549 cells and colon cancer HT29 cells. The intermediates 4a-4k are successfully synthesized from 1a-1k and ethyl 2-(3-bromopyridin-2-yl)acetate by Knoevenagel condensation and intramolecular Heck-type reaction, and this is a novel and efficient synthetic approach to the core scaffold of the target compounds. These target compounds have shown an interesting antitumour profile towards the tested cell lines with IC50 values ranging from 0.07 µM to 4.45 µM. Among all the compounds synthesized, 8 compounds show higher potency than R16, 12 compounds are as potent as R16, and 6 compounds are less potent than R16. The best compound 24 is 7 times and approximately 10 times as potent as R16 against A549 and HT29 cells, respectively.

[The role of CXCR3 and its ligand I-TAC in the pathogenesis of immune thrombocytopenic purpura].

OBJECTIVE: To investigate the roles of the chemokine receptor CXCR3 and its ligand I-TAC in the pathogenesis of immune thrombocytopenic purpura (ITP). METHODS: A total of 48 ITP patients were enrolled in this study: 30 with newly diagnosed or relapse ITP and 18 in remission after treatment, and 24 healthy volunteers were as controls. IFNγ and I-TAC in plasma were detected by ELISA. The mRNA expression of CXCR3 in the peripheral blood mononuclear cells (PBMNCs) was determined by quantitative RT-PCR. RESULTS: The IFNγ level in the plasma of ITP patients before the treatment was obviously increased than those in the remission group and controls [(71.45 ± 17.62) ng/L vs (36.94 ± 14.86) ng/L and (25.28 ± 12.85) ng/L, all P < 0.05] and those in the remission group was higher than in the controls (P < 0.05). In contrast, there were no statistic differences of the levels of I-TAC among the three groups [(455.56 ± 144.70) ng/L, (488.24 ± 164.70) ng/L and (382.97 ± 167.43) ng/L, P > 0.05]. Both ITP patients before the treatment and remission groups expressed more CXCR3 mRNA [6.76 (3.03, 37.00), 1.76 (0.45, 14.18) vs 0.12 (0.04, 0.28), P < 0.05]. After effective therapy, CXCR3 mRNA expression decreased, while it was still higher than that in the controls. CONCLUSIONS: Our data demonstrate that Th1 cytokine (IFNγ) dominance is reflected in ITP. Simultaneously, the CXCR3(+) cell may play a role in cell-mediated immunity through chemotaxis in ITP.

Characterization of the conversion between CD133+ and CD133- cells in colon cancer SW620 cell line.

The state of cancer stem cells (CSC) under reversible fluctuations, which has been revealed in breast cancer cells most recently, suggests that subpopulations with distinct phenotypes and functions within cancer cells can undergo inter-conversion. To investigate the possibility in colon cancer cells, we employed CD133 as the CSC marker, and characterized CD133 expression pattern and the biological features of the CD133 (+) and CD133 (-) subsets. Flow cytometry revealed that CD133 was bimodally expressed in SW620 cells among eight colon cancer cell lines. The CD133 (+) clonal SW620 cells displayed a differential gene expression profile, higher cellular reactive oxygen species (ROS), enhanced tumorigenesis and resistance to 5-fluorouracil. The conversion in term of the CD133 phenotype of the sorted cells was observed in vitro and in vivo. The fraction of the CD133 (+) cells decreased from 99% to 80% in the sorted CD133 (+) population while rising from 5 to 10% in the sorted CD133 (-) population during the first 20-day cultivation and then stayed almost unchanged. A fraction (about 20%) of the CD133 (+) clonal cells lost their CD133 marker while about 10% of the CD133 (-) clonal cells acquired the CD133 marker. 5-Azacytidine enhanced the fraction of the CD133 (+) cells in both of the CD133 (+) and CD133 (-) clonal cells. Our data demonstrate that CD133 expression is dynamic and reversible, and reveal the inter-conversion between the CD133 (+) and the CD133 (-) SW620 cells, suggesting that the CD133 phenotype of SW620 cell population is retained by the conversion between the two cell subsets.

Newly discovered angiogenesis inhibitors and their mechanisms of action.

In the past decade, the success of angiogenesis inhibitors in clinical contexts has established the antiangiogenic strategy as an important part of cancer therapy. During that time period, we have discovered and reported 17 compounds that exert potent inhibition on angiogenesis. These compounds exhibit tremendous diversity in their sources, structures, targets and mechanisms. These studies have generated new models for further modification and optimization of inhibitory compounds, new information for mechanistic studies and a new drug candidate for clinical development. In particular, through studies on the antiangiogenic mechanism of pseudolaric acid B, we discovered a novel mechanism by which the stability of hypoxia-inducible factor 1α is regulated by the transcription factor c-Jun. We also completed a preclinical study of AL3810, a compound with the potential to circumvent tumor drug resistance to a certain extent. All of these findings will be briefly reviewed in this article.

[The immunocyte subsets and their clinical significance in the peripheral blood of 35 patients with immune thrombocytopenic purpura].

OBJECTIVE: To explore the clinical significance of immunocyte subsets before and after immunosuppressive therapy in the peripheral blood of patients with immune thrombocytopenic purpura (ITP). METHODS: The percentages of immunocyte subsets in the peripheral blood of 35 patients with ITP and 20 healthy controls were detected by flow cytometry, including CD(3)(+), CD(4)(+), CD(8)(+), CD(+)(56), CD(19)(+) lymphocytes and CD(4)(+)/CD(8)(+). RESULTS: The percentages of CD(3)(+) T lymphocyte (61.58 ± 6.45)%, CD(4)(+) T lymphocyte (28.38 ± 4.89)% and the ratio of CD(4)(+)/CD(8)(+) 0.99 ± 0.22 in patients with ITP were lower than those in healthy controls [(67.85 ± 4.68)%, (38.00 ± 3.37)%, 1.54 ± 0.13, all P < 0.05]. After immunosuppressive therapy, the percentages of CD(3)(+)T lymphocyte (69.41 ± 5.03)%, CD(4)(+)T lymphocyte (38.17 ± 3.18)% and the ratio of CD(4)(+)/CD(8)(+) 1.60 ± 0.15 recovered to control levels. The percentages of CD(8)(+)T lymphocyte (29.20 ± 4.50)% and CD(19)(+)B lymphocyte (17.74 ± 4.14)% were higher than those in healthy controls [(24.82 ± 2.93)% and (12.09 ± 3.51)%, all P < 0.05]. After the immunosuppressive therapy, the percentages of CD(8)(+)T lymphocyte (24.06 ± 3.02)% and CD(19)(+)B lymphocyte (10.90 ± 3.55)% recovered to control levels. There were no significant difference of the percentage of CD(56)(+) lymphocyte among ITP patients (15.80 ± 2.85)%, ITP patients after immunosuppressive therapy (15.16 ± 2.77)% and healthy controls (16.36 ± 2.75)%. CONCLUSION: The aberrant immunocyte subsets are involved in the pathogenesis of ITP, and detection of immunocyte subsets might be helpful for the diagnosis and determination of therapeutic outcome of ITP.

D11, a novel glycosylated diphyllin derivative, exhibits potent anticancer activity by targeting topoisomerase IIα.

Glycosylated natural products are reliable platforms for the development of anticancer drugs, simply due to the important features added by sugar appendages to the shape and the stereoelectronic properties of natural scaffolds. Herein, we indentified D11, a novel diphyllin glycoside with acetylated D-quinovose sugar moiety, as a potent topoisomerase IIα (Topo IIα) inhibitior. This peculiar sugar moiety endows D11 an optimal conformation with a high binding affinity for Topo IIα via hydrogen bonding to the entrance of ATPase pocket, thereby helping achieve more potent Topo II inhibition activity compared to the aglycon diphyllin. Further biochemical insights manifested that D11 significantly inhibited Topo IIα ATPase-catalyzed ATP hydrolysis in an ATP-dependent, but a DNA-independent manner. All these underlie the consequent superiority of D11 in the in vitro proliferation inhibition, apoptosis induction and the in vivo remarkable antitumor potency in xenograft mouse model. Moreover, D11 treatment also displayed no obvious body weight loss and other apparent toxicities, indicative of the restricted side effects by the virtue of sugar attachment. Taken together, a defined glycosylated manipulation of diphyllin may be a promising alternative approach in the development of novel Topo II inhibitors.

Increased accumulation of hypoxia-inducible factor-1α with reduced transcriptional activity mediates the antitumor effect of triptolide.

BACKGROUND: Hypoxia-inducible factor-1α (HIF-1α), a critical transcription factor to reduced O2 availability, has been demonstrated to be extensively involved in tumor survival, aggressive progression, drug resistance and angiogenesis. Thus it has been considered as a potential anticancer target. Triptolide is the main principle responsible for the biological activities of the Traditional Chinese Medicine tripterygium wilfordii Hook F. Triptolide possesses great chemotherapy potential for cancer with its broad-spectrum anticancer, antiangiogenesis, and drug-resistance circumvention activities. Numerous biological molecules inhibited by triptolide have been viewed as its possible targets. However, the anticancer action mechanisms of triptolide remains to be further investigated. Here we used human ovarian SKOV-3 cancer cells as a model to probe the effect of triptolide on HIF-1α. RESULTS: Triptolide was observed to inhibit the proliferation of SKOV-3 cells, and meanwhile, to enhance the accumulation of HIF-1α protein in SKOV-3, A549 and DU145 cells under different conditions. Triptolide did not change the kinetics or nuclear localization of HIF-1α protein or the 26 S proteasome activity in SKOV-3 cells. However, triptolide was found to increase the levels of HIF-1α mRNA. Unexpectedly, the HIF-1α protein induced by triptolide appeared to lose its transcriptional activity, as evidenced by the decreased mRNA levels of its target genes including VEGF, BNIP3 and CAIX. The results were further strengthened by the lowered secretion of VEGF protein, the reduced sprout outgrowth from the rat aorta rings and the inhibitory expression of the hypoxia responsive element-driven luciferase reporter gene. Moreover, the silencing of HIF-1α partially prevented the cytotoxicity and apoptosis triggered by triptolide. CONCLUSIONS: The potent induction of HIF-1α protein involved in its cytotoxicity, together with the suppression of HIF-1 transcriptional activity, indicates the great therapeutic potential of triptolide as an anticancer drug. Meanwhile, our data further stress the possibility that HIF-1α functions in an unresolved nature or condition.

Naphthalimides induce G(2) arrest through the ATM-activated Chk2-executed pathway in HCT116 cells.

Naphthalimides, particularly amonafide and 2-(2-dimethylamino)-6-thia-2-aza-benzo[def]chrysene-1,3-diones (R16), have been identified to possess anticancer activities and to induce G(2)-M arrest through inhibiting topoisomerase II accompanied by Chk1 degradation. The current study was designed to precisely dissect the signaling pathway(s) responsible for the naphthalimide-induced cell cycle arrest in human colon carcinoma HCT116 cells. Using phosphorylated histone H3 and mitotic protein monoclonal 2 as mitosis markers, we first specified the G(2) arrest elicited by the R16 and amonafide. Then, R16 and amonafide were revealed to induce phosphorylation of the DNA damage sensor ataxia telangiectasia-mutated (ATM) responding to DNA double-strand breaks (DSBs). Inhibition of ATM by both the pharmacological inhibitor caffeine and the specific small interference RNA (siRNA) rescued the G(2) arrest elicited by R16, indicating its ATM-dependent characteristic. Furthermore, depletion of Chk2, but not Chk1 with their corresponding siRNA, statistically significantly reversed the R16- and amonafide-triggered G(2) arrest. Moreover, the naphthalimides phosphorylated Chk2 in an ATM-dependent manner but induced Chk1 degradation. These data indicate that R16 and amonafide preferentially used Chk2 as evidenced by the differential ATM-executed phosphorylation of Chk1 and Chk2. Thus, a clear signaling pathway can be established, in which ATM relays the DNA DSBs signaling triggered by the naphthalimides to the checkpoint kinases, predominantly to Chk2,which finally elicits G(2) arrest. The mechanistic elucidation not only favors the development of the naphthalimides as anticancer agents but also provides an alternative strategy of Chk2 inhibition to potentiate the anticancer activities of these agents.

Proteasome-dependent degradation of Chk1 kinase induced by the topoisomerase II inhibitor R16 contributes to its anticancer activity.

The novel naphthalimide derivative R16 has been demonstrated to exhibit potent in vitro and in vivo anticancer activity by inhibiting topoisomerase II (Top2). R16 induces G(2) arrest via an ATM-activated Chk2-executed pathway, accompanied by reducing Chk1. In this study, R16 was demonstrated to trigger time and concentration-dependent Chk1 reduction which was unrelated to the mRNA level and HSP90-involved degradation. Pretreatment of HCT116 cells with the proteasome inhibitors MG132 or lactacystin prevented Chk1 decline induced by R16, accompanied by significant accumulation of ubiquitinated Chk1 protein, indicating the involvement of ubiquitin-proteasome pathway. Meanwhile, R16 also resulted in loss of Chk1 function. By site-specifically mutating the phosphorylation sites of Chk1 protein at Ser317 or at Ser345, we further demonstrated that R16-triggered Chk1 reduction was associated with its apoptotic induction and cell killing. In conclusion, the data reveal that the novel Top2 inhibitor R16 induces degradation of Chk1 via the ubiquitin-proteasome pathway, impairing the function of Chk1 and thus contributing to the anticancer activity of R16.