Interactions between epidermal growth factor-mediated autocrine regulation and linoleic acid-stimulated growth of a human prostate cancer cell line.

Human prostate cancer (PC) cell lines possess epidermal growth factor (EGF) receptors and secrete EGF-related polypeptides. We used an EGF receptor-blocking antibody (anti-EGF.R) to demonstrate a functional autocrine loop, as well as the interaction between this and the effects of linoleic acid (LA), an omega-6 fatty acid, on PC cell growth. The anti-EGF.R competed effectively with [125I]EGF for receptors on DU145 PC cells, and on a high-passage DU145 variant (DU145M); when added to the culture medium, it suppressed both DU145 and DU145M cell growth in a dose-dependent manner. LA, a precursor for eicosanoid synthesis, had little effect on DU145 cell growth rate but stimulated DU145M growth in a concentration-related manner over a range of 0.25-2.0 micrograms/ml. anti-EGF.R (10(-9) M) caused suppression of LA-stimulated growth of DU145M cells in serum-free medium, which was prevented by the addition of 2 nM EGF. We conclude that an EGF.R-mediated autocrine loop is involved in PC cell growth regulation and that at least one site of action may be the synthesis of eicosanoids from their LA precursor.     

Biosynthesis of two stage-specific membrane proteins during transformation of Plasmodium gallinaceum zygotes into ookinetes

We have studied the synthesis and expression of surface proteins in zygotes of Plasmodium gallinaceum during their transformation to mature ookinetes. The cells were biosynthetically labelled in vitro using [35S]methionine and proteins were immunoprecipitated with rabbit anti-ookinete serum or monoclonal antibodies. Early zygotes (approx. 2 h post-gametogenesis and fertilization) synthesized and expressed on their surface a protein of Mr 26 000 as observed under reducing conditions on polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate (SDS-PAGE) (31 000 under non-reducing conditions) and continued to do so for 8-10 h; thereafter synthesis of the Mr 26 000 protein declined and little or none was synthesized in the mature ookinetes (greater than 20 h post-gametogenesis). Between 3-5 h post-gametogenesis, zygotes also began to synthesize a protein of Mr 28 000 (34 000 under non-reducing conditions). Synthesis and expression of this surface protein continued throughout development; and the Mr 28 000 protein was the predominant surface protein synthesized by the mature ookinete. Mr 26 000 and Mr 28 000 proteins have been designated earlier as PgO-1 and PgO-2 respectively (Carter and Kaushal, Mol. Biochem. Parasitol. (1984) 13, 235-241). Neither protein was synthesized in the gametocytes prior to gametogenesis. Both proteins could be labelled with [3H]glucosamine or [3H]mannose. When zygotes were incubated with [3H]palmitic acid both PgO-1 and PgO-2 bound fatty acids in covalent linkage. The two proteins do not otherwise appear to be structurally related. They were differentially immunoprecipitated by different monoclonal antibodies and gave rise to distinct patterns of peptides following digestion with proteases such as Staphylococcus aureus V-8, trypsin and chymotrypsin.     

Phase I pharmacokinetic trial of the selective oral epidermal growth factor receptor tyrosine kinase inhibitor gefitinib ('Iressa', ZD1839) in Japanese patients with solid malignant tumors.

BACKGROUND: This phase I dose-escalating study investigated the tolerability and toxicity of the selective epidermal growth factor receptor tyrosine kinase inhibitor gefitinib ('Iressa', ZD1839) in Japanese patients with solid tumors. Thirty-one patients were included. PATIENTS AND METHODS: Patients initially received a single oral dose of gefitinib followed by 10-14 days of observation. Oral gefitinib was subsequently administered on 14 consecutive days, every 28 days. Dose escalation was from 50 mg/day to a maximum of 925 mg/day or dose-limiting toxicity (DLT). RESULTS: Most adverse events were mild (grade 1/2); the most frequent were an acne-like rash and gastrointestinal effects. Two of six patients at 700 mg/day had DLT; no further dose escalation occurred. C(max) was reached within 3-7 h and exposure to gefitinib increased with dose. Mean terminal half-life following multiple dosing was 50.1 h (range 27.8-79.7 h). A partial response (duration 35-361 days) was observed in five of the 23 patients with non-small-cell lung cancer over a range of doses (225-700 mg/day), and seven patients with a range of tumors had disease stabilization (duration 40-127 days). CONCLUSIONS: In conclusion, gefitinib showed a favorable tolerability profile in Japanese patients. The safety profile, pharmacokinetic parameters and antitumor activity observed in our study are comparable to those observed in patients from the USA and Europe.     

炭疽毒素受体2表达受miR-145-5p调控在胰腺癌的作用机制研究

目的：探讨炭疽毒素受体2(anthrax toxin receptor 2,ANTXR2)在胰腺癌组织和细胞中的表达、临床意义和细胞功能,验证miR-145-5p在胰腺癌细胞中调控ANTXR2的作用机制。方法：分析GEO和TCGA数据库中胰腺癌的转录数据和临床数据。通过CCK-8实验、流式细胞术、划痕实验和Transwell实验进行细胞功能验证。采用miRNA靶点预测数据库反向预测可能通过ceRNA机制抑制ANTXR2表达的miRNA,并使用双荧光素酶报告实验进行验证。结果：ANTXR2在胰腺癌组织和细胞中的表达均上调（均P<0.05）,ANTXR2上调预示着原发肿瘤等级（P<0.05）、肿瘤细胞分化等级（P<0.05）和总体生存期（P<0.05,HR=1.72）均较差。ANTXR2可以促进胰腺癌细胞增殖、迁移和侵袭（均P<0.05）。ANTXR2是miR-145-5p下游的靶标（P<0.01）,miR-145-5p可以通过抑制ANTXR2的表达抑制胰腺癌细胞增殖、迁移和侵袭。结论：ANTXR2在胰腺癌组织和细胞中过表达,促进胰腺癌细胞增殖、迁移和侵...     

上调miR-145表达抑制卵巢癌HO8910细胞侵袭、迁移及促进TβR-Ⅱ蛋白表达

目的:探究微小RNA-145(microRNA-145,miR-145)对卵巢癌(ovarian cancer,OC)HO8910细胞侵袭、迁移及TβR-Ⅱ蛋白水平的影响.方法:将某大学研究实验室细胞库提供的OC细胞HO8910分为ZW组(无转染HO8910细胞)、ZN组(转染miR-145-NC)、YJ组(转染miR...     

miR-145调控肝癌腹水模型Th9细胞升高的机制研究

目的:探讨miR-145调控肝癌腹水模型Th9细胞升高的作用机制。方法:构建小鼠H22肝癌腹水模型。造模两周后处死小鼠并分离出脾脏组织,流式细胞术分析肝癌腹水组（MA组）与正常对照组（Control 组）小鼠脾脏Th9细胞表达水平。ELISA法检测脾脏IL-9表达水平。RT-PCR法检测脾脏miR-145表达水平。Western Blot法检测脾脏中PI3K/Akt/mTOR/P70S6K/HIF-1α相关蛋白表达水平。分选小鼠脾脏CD4+T细胞,随机分为miR-145 mimics组和NC组,分别应用miR-145-5P mimics、阴性对照寡核苷酸（negative control,NC）进行转染,RT-PCR检测各组miR-145、HIF-1α mRNA及IL-9 mRNA表达水平。结果:与Control 组相比,MA组Th9细胞及其细胞因子IL-9表达均升高（P<0.05）,miR-145表达降低（P<0.05）,p-PI3K、p-Akt、mTOR、p-mTOR、p-P70S6K、HIF-1α蛋白均升高（P<0.05）。与NC组相比,miR-145 mimics组miR-145显著升高,HIF-1α mRNA及IL-9 mRNA表达下降。结论:肝癌腹水中Th9细胞升高可能与miR-145下降导致的PI3K/Akt/mTOR/P70S6K/HIF-1α通路激活有关。     

miR-155在结肠癌组织中表达的临床病理意义

目的:探讨结肠癌组织及细胞株中microRNA-155(miR-155)表达的临床病理意义.方法:采用RT-qPCR检测40例结肠癌组织及癌旁组织、结肠癌细胞株中miR-155的mRNA的表达,并对其临床病理指标进行分析,分析抑制miR-155表达对结肠癌细胞增殖、克隆形成及凋亡影响.结果:RT-qPCR检测结果表明40例结肠癌组织中miR-155相对表达量明显高于癌旁组织(P<0.01).不同分化结肠癌细胞miR-155相对表达量差异有统计学意义(P<0.05),其中低分化结肠癌细胞中表达均高于中、高分化结肠癌细胞(P<0.05~P<0.01).miR-155的异常高表达与病人肿瘤较大、分化程度差、淋巴结发生转移情况及远处有转移均有关(P<0.05~P<0.01),而与年龄和性别无关(P>0.05).抑制miR-155可明显抑制结肠癌细胞增殖、克隆形成.结论:结肠癌组织中高表达miR-155可以作为预测恶性程度、淋巴转移的有效指标,miR-155在结肠癌病人的临床靶向治疗中具有一定的价值.     

金纳米颗粒表面催化发夹组装无酶信号放大快速荧光法检测miRNA-721

目的 构建一种自催化发夹组装（CHA）无酶信号放大的靶标快速荧光法,用于检测靶标miRNA。方法 首先在金纳米颗粒（AuNPs）表面修饰5-羧基荧光素（FAM）标记的DNA发夹探针H1,H1的荧光被金纳米粒子猝灭。加入靶标miRNA会导致AuNPs上的H1标记荧光素远离AuNPs而发射荧光,随后H1和H2之间发生循环自组装,靶标循环被利用,导致荧光信号放大。将20 μL探针AuNPs-H1,50 μL 3 μmol/L探针 H2（退火缓冲液为20 mmol/L Tris-HCl,100 mmol/L NaCl,5 mmol/L MgCl2,pH 7.4）,与50 μL不同浓度的miRNA-721（0.1 ~ 5 μmol/L）在30℃下共反应20 min,最后在480 nm激发波长下测定体系的荧光强度。结果 以急性心肌炎生物标志物miRNA-721为模型靶标,在优化的实验条件下得到520 nm处的相对荧光强度变化值（?F = F – F0）与miRNA-721浓度呈良好的线性关系,拟合线性方程?F = 29232lgC – 52435（R2 = 0.9910）。该荧光法检测限为1.23 nmol/ L。在正常人血清中的加标回收率为92.71% ~ 104.02%。一次完整的miRNA分析可以在20 min内完成。结论 该方法可用于生物样品中miRNA-721的检测,为急性心肌炎的早期诊断提供了一种快速检测手段。     

乳腺肿瘤患者血清miRNA-222、miRNA-21检测的临床意义

目的 探讨血清miRNA-222、miRNA-21水平对于乳腺肿瘤患者的临床意义.方法 选择经病理确诊并分类的35例乳腺癌患者和32例乳腺良性肿瘤患者;同时选35例女性健康体检者作为对照.采用RT-PCR的方法测定所有受试者的血清miRNA-222、miRNA-21的相对表达水平,分析血清miRNA-222、miRNA...     

长链非编码 RNALOC100288637在原发性肝癌中差异性表达的生物学意义及相关机制研究

目的：研究长链非编码 RNA （LncRNA ） LOC100288637在肝癌中的表达差异,对肝癌细胞增殖的影响及相关机制。方法分析GEO数据集GSE58043和GSE10694,得出LOC100288637及hsa‐miR‐101‐3p在肝癌组织中差异表达,RNA‐hy‐brid分析LOC100288637与hsa‐miR‐101‐3p的结合具有可能性。逆转录‐聚合酶链式反应在组织和细胞水平检测LOC100288637表达量。荧光原位杂交观察LOC100288637在细胞中的定位。敲低LOC100288637后CCK‐8检测细胞增殖情况。过表达hsa‐miR‐101‐3p后,检测 LOC100288637的变化。结果 LOC100288637在肝癌组织中的表达量高于癌旁组织（ P＜0．05）;LOC100288637在肝癌细胞系中的表达量高于肝细胞系（P＜0．05）。LOC100288637在 HepG2细胞核质均有表达,以细胞质居多。敲低LOC100288637后 HepG2细胞增殖活性降低（P＜0．01）。过表达hsa‐miR‐101‐3p后,LOC100288637表达量下降（P＜0．05）。结论 LncRNA LOC100288637可能在肝癌发生,发展过程中起重要作用并接受hsa‐miR‐101‐3p靶向调控影响肝癌细胞的增殖。