• 【无细胞翻译过程中异寡聚去唾液酸糖蛋白受体复合物的组装。】 复制标题 收藏 收藏
    DOI:10.1073/pnas.87.12.4854 复制DOI
    作者列表:Sawyer JT,Doyle D
    BACKGROUND & AIMS: :We have translated RNAs for the two rat asialoglycoprotein receptor polypeptides together in a cell-free system containing dog pancreatic microsomes and immunoprecipitated the products with antibodies that distinguish the two proteins. In this system the proteins oligomerize, as judged by their coprecipitation with either of the subunit-specific antisera. Oligomerization does not occur between subunits synthesized without microsomes or between subunits synthesized on separate microsomes mixed during detergent solubilization. Thus, oligomerization occurs within the microsomal membrane. We calculate that oligomerization proceeds with an efficiency of approximately 85%. The receptor complex appears to represent a specific oligomer because it excludes a third membrane glycoprotein synthesized in the same reaction. Oligomerization of the asialoglycoprotein receptor in vitro should provide a useful system to study the assembly of a membrane-protein complex.
    背景与目标: : 我们已经在包含狗胰腺微粒体的无细胞系统中将两种大鼠去唾液酸糖蛋白受体多肽的rna翻译在一起,并用区分这两种蛋白质的抗体免疫沉淀了产物。在该系统中,蛋白质寡聚,根据它们与亚基特异性抗血清中的任何一种的共沉淀来判断。在没有微粒体的情况下合成的亚基之间或在洗涤剂增溶过程中混合的单独的微粒体上合成的亚基之间不会发生低聚。因此,低聚发生在微粒体膜内。我们计算低聚化以约85% 的效率进行。受体复合物似乎代表特定的寡聚体,因为它排除了在同一反应中合成的第三种膜糖蛋白。体外去唾液酸糖蛋白受体的寡聚化应为研究膜蛋白复合物的组装提供有用的系统。
  • 【重组人可溶性肿瘤坏死因子受体融合蛋白治疗异基因造血干细胞移植后类固醇难治性移植物抗宿主病.】 复制标题 收藏 收藏
    DOI:10.1002/ajh.20752 复制DOI
    作者列表:Busca A,Locatelli F,Marmont F,Ceretto C,Falda M
    BACKGROUND & AIMS: :Etanercept is a recombinant human soluble tumor necrosis factor (TNF-alpha) receptor fusion protein that inhibits TNF-alpha, a major mediator in the pathogenesis of graft-versus-host disease (GVHD). The purpose of our study was to evaluate the safety and efficacy of etanercept therapy in 21 patients with steroid-refractory acute GVHD (aGVHD) (n = 13) and chronic GVHD (cGVHD) (n = 8). Etanercept 25 mg was given subcutaneously twice weekly for 4 weeks followed by 25 mg weekly for 4 weeks. At the time of initiation of etanercept, 14 patients had skin, 13 had gastro-intestinal, 5 had liver, 5 had pulmonary, and 4 had oral involvement. Twelve patients (57%) completed 12 doses of therapy. Overall, 11 of 21 patients (52%) responded to the treatment with etanercept, including 6 patients (46%) with aGVHD [n = 4 complete response (CR), n = 2 partial response (PR)] and 5 patients (62%) with cGVHD (n = 1 CR, n = 4 PR). Clinical responses were most commonly seen in patients with refractory gut aGVHD with 55% of the patients having a CR and 9% having a PR. CMV reactivation occurred in 48% of patients, bacterial infections in 14% of patients, and fungal infections in 19% of patients. Fourteen patients (67%) were alive after a median follow-up of 429 days (range 71-1007 days) since initiation of etanercept. Seven patients died, 3 of infections, 2 of refractory aGVHD, and 2 of disease progression. In conclusion, our preliminary data indicate that etanercept is well tolerated and can induce a high response rate in patients with steroid-refractory aGVHD and cGVHD, particularly in the setting of GI involvement.
    背景与目标: : 依那西普是一种重组人可溶性肿瘤坏死因子 (TNF-α) 受体融合蛋白,可抑制TNF-α,TNF-α 是移植物抗宿主病 (GVHD) 发病机理中的主要介质。我们研究的目的是评估依那西普治疗21例类固醇难治性急性GVHD (aGVHD) (n = 13) 和慢性GVHD (cGVHD) (n = 8) 患者的安全性和有效性。依那西普25 mg,每周皮下注射两次,持续4周,然后每周注射25 mg,持续4周。在开始使用依那西普时,14例患者有皮肤,13例有胃肠道,5例有肝脏,5例有肺部,4例有口腔受累。12名患者 (57%) 完成12剂治疗。总体而言,21例患者中有11例 (52%) 对依那西普治疗有反应,其中6例 (46% 例) aGVHD [n = 4完全缓解 (CR),n = 2部分缓解 (PR)] 和5例 (62%) cGVHD (n = 1 CR,n = 4 PR)。临床反应最常见于难治性肠道aGVHD患者,其中55% 患者具有CR,9% 患者具有PR。48% 患者发生CMV再激活,14% 患者发生细菌感染,19% 患者发生真菌感染。自依那西普开始以来,中位随访429天 (范围71-1007天) 后,有14名患者 (67%) 还活着。7例患者死亡,3例感染,2例难治性aGVHD,2例疾病进展。总之,我们的初步数据表明,依那西普具有良好的耐受性,并且可以在类固醇难治性aGVHD和cGVHD患者中诱导高反应率,尤其是在GI受累的情况下。
  • 【第3天或第5天胚胎移植后血清雌二醇和 β-HCG测定解释妊娠结局。】 复制标题 收藏 收藏
    DOI:10.1016/s1472-6483(10)60631-1 复制DOI
    作者列表:Kumbak B,Oral E,Karlikaya G,Lacin S,Kahraman S
    BACKGROUND & AIMS: :The aim of this study was to assess the clinical value of serum oestradiol concentration 8 days after embryo transfer (D8E2) and beta-human chorionic gonadotrophin (HCG-beta) concentration 12 days after embryo transfer (D12HCG-beta) in the prediction of pregnancy and the outcome of pregnancy following assisted reproduction, taking into account the day of transfer, which was either day 3 (D3) or day 5 (D5). The objective was to improve patient counselling by giving quantitative and reliable predictive information instead of non-specific uncertainties. A total of 2035 embryo transfer cycles performed between January 2003 and June 2005 were analysed retrospectively. Biochemical pregnancy, ectopic pregnancy and first-trimester abortions were classified as non-viable pregnancies; pregnancies beyond 12 weeks gestation were classified as ongoing pregnancies (OP). Significantly higher D8E2 and D12HCG-beta were obtained in D5 transfers compared with D3 transfers with regard to pregnancy and OP (P
    背景与目标: 本研究的目的是评估胚胎移植后8天血清雌二醇浓度 (D8E2) 和胚胎移植后12天 β-人绒毛膜促性腺激素 (HCG-β) 浓度 (D12HCG-beta) 在预测妊娠和辅助生殖后妊娠结局中的临床价值,考虑到转移的日期,即第3天 (D3) 或第5天 (D5)。目的是通过提供定量和可靠的预测信息而不是非特异性不确定性来改善患者咨询。回顾性分析了2003年1月和2005年6月之间进行的2035年胚胎移植周期。生化妊娠,异位妊娠和早孕流产被归类为不可行妊娠; 妊娠12周以上的妊娠被归类为持续妊娠 (OP)。就妊娠和OP而言,与D3转移相比,D5转移获得了明显更高的D8E2和D12HCG-beta (P <或 = 0.001)。对于D3胚胎移植,D8E2预测OP的临界值为130 pg/ml (敏感性80%,特异性72%),而D12HCG-beta为98 mIU/ml (敏感性89%,特异性69%)。对于D5胚胎移植,分别为179 pg/ml (敏感性79%,特异性84%) 和257 mIU/ml (敏感性78%,特异性81%)。看来,胚胎移植后血清D8E2和D12HCG-beta浓度提供了有关IVF胚胎移植后妊娠和妊娠结局的明确信息。
  • 【细胞外钙感应受体的激活启动人胰岛的胰岛素分泌: 蛋白激酶的参与。】 复制标题 收藏 收藏
    DOI:10.1677/joe.1.06891 复制DOI
    作者列表:Gray E,Muller D,Squires PE,Asare-Anane H,Huang GC,Amiel S,Persaud SJ,Jones PM
    BACKGROUND & AIMS: :The extracellular calcium-sensing receptor (CaR) is usually associated with systemic Ca(2+) homeostasis, but the CaR is also expressed in many other tissues, including pancreatic islets of Langerhans. In the present study, we have used human islets and an insulin-secreting cell line (MIN6) to investigate the effects of CaR activation using the calcimimetic R-568, a CaR agonist that activates the CaR at physiological concentrations of extracellular Ca(2+). CaR activation initiated a marked but transient insulin secretory response from both human islets and MIN6 cells at a sub-stimulatory concentration of glucose, and further enhanced glucose-induced insulin secretion. CaR-induced insulin secretion was reduced by inhibitors of phospholipase C or calcium-calmodulin-dependent kinases, but not by a protein kinase C inhibitor. CaR activation was also associated with an activation of p42/44 mitogen-activated protein kinases (MAPK), and CaR-induced insulin secretion was reduced by an inhibitor of p42/44 MAPK activation. We suggest that the beta-cell CaR is activated by divalent cations co-released with insulin, and that this may be an important mechanism of intra-islet communication between beta-cells.
    背景与目标: : 细胞外钙感应受体 (CaR) 通常与全身Ca(2) 稳态有关,但CaR也在许多其他组织中表达,包括朗格汉斯的胰岛。在本研究中,我们使用人类胰岛和胰岛素分泌细胞系 (MIN6) 来研究使用钙拟R-568 (一种在细胞外Ca(2) 的生理浓度下激活CaR的CaR激动剂) 激活CaR的作用。CaR激活在葡萄糖的亚刺激浓度下启动了人类胰岛和MIN6细胞的显着但短暂的胰岛素分泌反应,并进一步增强了葡萄糖诱导的胰岛素分泌。磷脂酶C或钙-钙调蛋白依赖性激酶的抑制剂可减少CaR诱导的胰岛素分泌,但蛋白激酶C抑制剂不能减少。CaR激活也与p42/44丝裂原活化蛋白激酶 (MAPK) 的激活有关,并且CaR诱导的胰岛素分泌被p42/44 MAPK激活的抑制剂减少。我们建议 β 细胞CaR被与胰岛素共同释放的二价阳离子激活,这可能是 β 细胞之间胰岛内通讯的重要机制。
  • 【烟碱乙酰胆碱受体的酪氨酸磷酸化介导Grb2结合。】 复制标题 收藏 收藏
    DOI: 复制DOI
    作者列表:Colledge M,Froehner SC
    BACKGROUND & AIMS: Tyrosine phosphorylation of the nicotinic acetylcholine receptor (AChR) is associated with an altered rate of receptor desensitization and also may play a role in agrin-induced receptor clustering. We have demonstrated a previously unsuspected interaction between Torpedo AChR and the adaptor protein Grb2. The binding is mediated by the Src homology 2 (SH2) domain of Grb2 and the tyrosine-phosphorylated delta subunit of the AChR. Dephosphorylation of the delta subunit abolishes Grb2 binding. A cytoplasmic domain of the delta subunit contains a binding motif (pYXNX) for the SH2 domain of Grb2. Indeed, a phosphopeptide corresponding to this region of the delta subunit binds to Grb2 SH2 fusion proteins with relatively high affinity, whereas a peptide lacking phosphorylation on tyrosine exhibits no binding. Grb2 is colocalized with the AChR on the innervated face of Torpedo electrocytes. Furthermore, Grb2 specifically copurifies with AChR solubilized from postsynaptic membranes. These data suggest a novel role for tyrosine phosphorylation of the AChR in the initiation of a Grb2-mediated signaling cascade at the postsynaptic membrane.

    背景与目标: 烟碱乙酰胆碱受体 (AChR) 的酪氨酸磷酸化与受体脱敏率的改变有关,并且也可能在agrin诱导的受体聚集中起作用。我们已经证明了鱼雷AChR和衔接蛋白grb2之间以前未曾怀疑的相互作用。结合由Grb2的Src同源2 (SH2) 结构域和AChR的酪氨酸磷酸化 δ 亚基介导。Δ 亚基的去磷酸化消除了Grb2结合。Δ 亚基的细胞质结构域包含grb2的SH2结构域的结合基序 (pYXNX)。实际上,对应于 δ 亚基该区域的磷酸肽以相对较高的亲和力与Grb2 SH2融合蛋白结合,而在酪氨酸上缺乏磷酸化的肽则没有结合。Grb2与AChR共定位在鱼雷细胞的神经支配面上。此外,Grb2与溶解于突触后膜的AChR特异性共聚。这些数据表明,AChR的酪氨酸磷酸化在突触后膜的Grb2-mediated信号级联反应的启动中起着新的作用。
  • 【B淋巴细胞群体对IgM受体连接的敏感性差异取决于局部因素。】 复制标题 收藏 收藏
    DOI:10.1093/intimm/9.5.755 复制DOI
    作者列表:Modigliani Y,Demengeot J,Vasconcellos R,Andersson J,Coutinho A,Grandien A
    BACKGROUND & AIMS: Ligation of surface IgM on B cells responding to lipopolysaccharide (LPS) suppresses terminal differentiation and high-rate Ig secretion with no effect on proliferation. As shown here, different B cell populations show characteristic mean values of ligand concentration required for 50% inhibition, with Gaussian distributions of sensitivity to IgM receptor ligation that reflect cellular heterogeneity of 'al-or-none' inhibitions in single cells. Differential sensitivity of B cell populations to IgM ligation seems to be locally determined by the cellular environment and unrelated to the 'maturity' of the responding cells. Thus, while long-lived peritoneal B cells are 3- to 5-fold more resistant than splenic B cells, there is no difference in sensitivity between short- and long-lived B cells in the spleen. Furthermore, while B cells in bone marrow and spleen differ in sensitivity by two orders of magnitude, B cells differentiated in vitro from bone marrow pre-B cells are as resistant as splenic B cells. Moreover, bone marrow cell culture supernatants restore a high level of sensitivity in such cell populations. Differential sensitivity to IgM receptor ligation is reproduced by multivalent nominal antigen, in cell populations that show identical dose-response inhibition curves to direct activation of protein kinase C by phorbol esters. We conclude that the level of sensitivity to IgM ligation is independent of the life span or maturity of the B cell, but differentially regulated in vivo by putative tissue factors.

    背景与目标: 对脂多糖 (LPS) 响应的b细胞上的表面IgM连接抑制了终末分化和高速度Ig分泌,而对增殖没有影响。如这里所示,不同的b细胞群体显示出50% 抑制所需的配体浓度的特征平均值,其对IgM受体连接的敏感性的高斯分布反映了单细胞中 “al-or-none” 抑制的细胞异质性。B细胞群体对IgM连接的不同敏感性似乎是由细胞环境局部决定的,与响应细胞的 “成熟度” 无关。因此,尽管长寿命的腹膜b细胞的抵抗力比脾b细胞高3至5倍,但脾脏中短寿命b细胞和长寿命b细胞之间的敏感性没有差异。此外,尽管骨髓和脾脏中的b细胞的敏感性相差两个数量级,但从骨髓前b细胞体外分化的b细胞与脾b细胞一样具有抵抗力。此外,骨髓细胞培养上清液在此类细胞群体中恢复了高水平的敏感性。在显示出与佛波酯直接激活蛋白激酶C相同的剂量反应抑制曲线的细胞群体中,多价名义抗原再现了对IgM受体连接的不同敏感性。我们得出的结论是,对IgM连接的敏感性水平与b细胞的寿命或成熟度无关,但在体内受假定的组织因素的差异调节。
  • 【[非洲男性乳腺癌,与瓦加杜古大学教学医院 (布基纳法索) 的5例病例有关]。】 复制标题 收藏 收藏
    DOI: 复制DOI
    作者列表:Sano D,Dao B,Lankoandé J,Touré B,Sakandé B,Traoré SS,Wandaogo A,Dakouré R,Sanou A
    BACKGROUND & AIMS: :A retrospective study of male breast cancer was undertaken at Ouagadougou University Teaching Hospital over a 3 year period (1993-1996). Authors report 5 cases representing 4.16% of all breast cancers. The patients' mean age was 61 years. The average duration of signs and symptoms before the diagnosis was 13 months. Clinically all the 5 cases presented advanced cancers (4 T4N2M0, 1 T4N2M1 according to UICC TNM System) with size ranging from 5.5, to 11.5 cm. Histology found: 2 medullary infiltrating carcinoma, 1 canalar infiltrating carcinoma, 1 colloid mucous carcinoma and 1 lobular infiltrating carcinoma. All patients had mastectomy associated with axillary clearance in 4 cases. Radiotherapy, chemotherapy and hormonotherapy were not associated because unavailable in Burkina Faso. Three patients died: the first, 10 days after surgical treatment and the 2 others respectively after 14 and 17 months. We have lost sight 1 patients. The last one is still alive. Authors find that to get better prognosis, it is important to improve medical and technical means, to increase information and to promote early detection.
    背景与目标: : 在瓦加杜古大学教学医院进行了为期3年 (1993-1996年) 的男性乳腺癌回顾性研究。作者报告了5例代表所有乳腺癌4.16% 的病例。患者的平均年龄为61岁。诊断前症状和体征的平均持续时间为13个月。临床上所有5例均表现为晚期癌症 (根据UICC TNM系统,4个T4N2M0,1个T4N2M1),大小从5.5到11.5厘米。组织学发现: 髓质浸润性癌2例,管内浸润性癌1例,胶体粘液性癌1例,小叶浸润性癌1例。所有患者均行乳房切除术伴腋窝清除4例。放疗,化疗和激素治疗不相关,因为在布基纳法索不可用。3例患者死亡: 手术治疗后第1、10天,另外2例分别在14和17个月后死亡。我们已经看不见1个病人了。最后一个还活着。作者发现,要获得更好的预后,必须改善医疗和技术手段,增加信息并促进早期发现。
  • 【一系列赛庚胺类似物的合成,对5-HT2A,5-HT2B和5-HT2C 5-羟色胺受体的亲和力和结构-活性关系。】 复制标题 收藏 收藏
    DOI:10.1248/cpb.45.842 复制DOI
    作者列表:Honrubia MA,Rodriguez J,Dominguez R,Lozoya E,Manaut F,Seijas JA,Villaverde MC,Calleja JM,Cadavid MI,Maayani S,Sanz F,Loza MI
    BACKGROUND & AIMS: :Cyproheptadine is a drug that shows high affinity for type 2 (5-HT2) receptors. We studied a series of compounds obtained by modification of the tricyclic system of Cyp (dibenzocycloheptadiene): 2f (thioxanthene), 2g (xanthene), 2h (dihydrodibenzocycloheptadiene), 2j (diphenyl), 2i (fluorene), and 3b (phenylmethyl). Their activities at the rat cerebral cortex 5-HT2A receptor were (pKi +/- S.E.M.): 8.80 +/- 0.11 (Cyp), 8.60 +/- 0.07 (2f), 8.40 +/- 0.02 (2g), 8.05 +/- 0.03 (2h), 7.87 +/- 0.12 (2j), 6.70 +/- 0.02 (2i) and 6.45 +/- 0.02 (3b); those at the rat stomach fundus 5-HT2B receptor (pA2 +/- S.E.M.) were: 9.14 +/- 0.25 (Cyp), 8.49 +/- 0.07 (2f), 7.58 +/- 0.58 (2g), 7.02 +/- 0.14 (2h), 6.07 +/- 0.20 (2j), and undetectable (2i, 3b): and those at the pig choroidal plexus 5-HT2C receptor (pKi +/- S.E.M.) were: 8.71 +/- 0.08 (Cyp), 8.68 +/- 0.01 (2f), 8.58 +/- 0.20 (2g), 7.95 +/- 0.05 (2h), 7.57 +/- 0.04 (2j), 6.98 +/- 0.04 (2i) and 6.63 +/- 0.20 (3b). The slopes did not differ significantly from unity. The compounds exhibited the same order of activities at every type of receptor, and the most active molecules presented certain steric (butterfly conformation of the tricyclic system) and electrostatic (proton affinity on the top of the central rings) patterns. It is concluded that the activity of cyproheptadine derivatives at 5-HT2 receptors is related to these molecular features, which make feasible a common disposition to interact with all three 5-HT2 subtypes.
    背景与目标: : 赛庚啶是一种对2型 (5-HT2) 受体显示高亲和力的药物。我们研究了通过修饰Cyp (二苯并环庚二烯) 的三环体系获得的一系列化合物: 2f (噻吩),2g (xanthene),2h (二氢二苯并环庚二烯),2j (二苯基),2i (芴) 和3b (苯基甲基)。它们在大鼠大脑皮层5-HT2A受体上的活性为 (pKi +/-s.e.M.): 8.80 +/- 0.11 (Cyp),8.60 +/- 0.07 (2f),8.40 +/- 0.02 (2g),8.05 +/- 0.03 (2h),7.87 +/- 0.12 (2j),6.70 +/- 0.02 (2i) 和6.45 +/- 0.02 (3b); 大鼠胃底5-HT2B受体 (pA2 +/-s.e.M.) 为: 9.14 +/- 0.25 (Cyp),8.49 +/- 0.07 (2f),7.58 +/- 0.58 (2g),7.02 +/- 0.14 (2h),6.07 +/- 0.20 (2j) 和不可检测 (2i,3b): 猪脉络丛5-HT2C受体 (pKi +/-s.e.M.) 为: 8.71 +/- 0.08 (Cyp),8.68 +/- 0.01 (2f),8.58 +/- 0.20 (2g),7.95 +/- 0.05 (2h) 、7.57 +/- 0.04 (2j) 、6.98 +/- 0.04 (2i) 和6.63 +/- 0.20 (3b)。坡度与统一没有显着差异。这些化合物在每种类型的受体上都表现出相同的活性顺序,并且最具活性的分子呈现某些空间 (三环系统的蝴蝶构象) 和静电 (中心环顶部的质子亲和力) 模式。结论是,赛庚啶衍生物在5-HT2受体上的活性与这些分子特征有关,这使得与所有三种5-HT2亚型相互作用的共同处置是可行的。
  • 【受体激活剂NFkappaB配体和骨保护素蛋白在人根尖周囊肿和肉芽肿中的表达。】 复制标题 收藏 收藏
    DOI:10.1016/j.tripleo.2005.10.054 复制DOI
    作者列表:Menezes R,Bramante CM,da Silva Paiva KB,Letra A,Carneiro E,Fernando Zambuzzi W,Granjeiro JM
    BACKGROUND & AIMS: OBJECTIVE:The purpose of this study was to determine the expression of receptor activator of NFkappaB ligand (RANKL) and osteoprotegerin (OPG) associated with bone destruction in periapical cysts and granulomas. STUDY DESIGN:Forty human dental chronic periapical lesions were collected after periapical surgery. The lesions collected were fixed in 10% buffered formalin and histologically processed. At least 2 sections of each specimen were stained with hematoxylin and eosin for microscopic diagnosis. After that, 10 human periapical granulomas and 10 cysts were selected for immunohistochemical analysis for RANKL, OPG, and CD68+. RESULTS:Polymorphonuclear neutrophils, macrophages, endothelial cells, and lymphocytes were stained for RANKL and OPG in both lesions. Epithelial cells were also stained for RANKL and OPG in periapical cysts. Quantitative analysis was conducted and the results were expressed as a ratio of the number of immunostained cells over the total number of cells in the field (n = 100). The ratio of RANKL+/total cells was higher than OPG+/total cells in periapical granulomas (0.553 +/- 0.153 and 0.483 +/- 0.189, respectively; P < .0012; paired t test) and in cysts (0.519 +/- 0.09 and 0.339 +/- 0.117, respectively; P < .0001; paired t test). The ratios of OPG+/total cells (P < .0001; paired t test) and RANKL+/total cells (P < .0322; paired t test) were greater in granulomas than in cysts. However, the ratio RANKL+/OPG+ in granulomas (1.336 +/- 0.723) and cysts (1.404 +/- 0.385) was not significantly different. The ratio of CD68+/total cells was significantly higher in granulomas (0.381 +/- 0.040) than in cysts (0.307 +/- 0.068) (P < .0001; unpaired t test with Welch correction). CONCLUSION:Taking into account the limitations of the experimental approach employed, our findings indicate the presence of RANKL and OPG in cysts and granulomas, strongly suggesting the involvement of these gene products in the development of periapical lesions.
    背景与目标:
  • 【大鼠I型清道夫受体 (SRBI) 在卵巢中的克隆,表征和细胞分布。】 复制标题 收藏 收藏
    DOI:10.1006/bbrc.1997.6646 复制DOI
    作者列表:Mizutani T,Sonoda Y,Minegishi T,Wakabayashi K,Miyamoto K
    BACKGROUND & AIMS: :An immediately inducible gene by gonadotropin was isolated from rat ovaries primed with pregnant mare serum gonadotropin (PMSG) by using a subtraction cloning procedure. Homology analysis revealed that the gene is a rat homologue of scavenger receptor class B-I, which was recently identified as a specific receptor for high density lipoprotein (HDL). The structure of rat SRBI was determined by nucleotide sequence analysis of full-length cDNAs for SRBI. Northern blot analysis revealed that rat SRBI mRNA levels were rapidly and strongly increased within 3 h by the injection of PMSG. In situ hybridization study revealed that SRBI mRNA was strongly induced in theca interna cells of immature rat ovary stimulated with 30 IU of PMSG for 6 h. SRBI mRNA expression was also observed in corpora lutea of the adult rat ovary. These findings indicate that expression of SRBI mRNA is restricted to and induced in the ovarian steroidogenic cell types where cholesterol is used as a substrate for synthesis of steroid hormones. Our data strongly suggest that SRBI may play a significant role in the ovarian steroidogenesis by mediating selective uptake of cholesterol from HDL to ovarian theca interna cells or to corpus luteum.
    背景与目标: : 使用减法克隆程序从用妊娠母马血清促性腺激素 (PMSG) 引发的大鼠卵巢中分离出促性腺激素立即诱导的基因。同源性分析表明,该基因是清道夫受体B-I类的大鼠同源物,最近被鉴定为高密度脂蛋白 (HDL) 的特异性受体。通过SRBI全长cdna的核苷酸序列分析确定大鼠SRBI的结构。Northern印迹分析显示,通过注射PMSG,大鼠SRBI mRNA水平在3小时内迅速且强烈地增加。原位杂交研究表明,用30 IU的PMSG刺激6 h,在未成熟大鼠卵巢的卵泡膜细胞中强烈诱导SRBI mRNA。在成年大鼠卵巢的黄体中也观察到了SRBI mRNA的表达。这些发现表明,SRBI mRNA的表达仅限于并在其中胆固醇用作合成类固醇激素的底物的卵巢类固醇生成细胞类型中诱导。我们的数据强烈表明,SRBI可能通过介导胆固醇从HDL到卵巢卵泡膜细胞或黄体的选择性摄取而在卵巢类固醇生成中起重要作用。
  • 【细菌基因lfpA影响Burkholderia pseudomalei诱导的TRAP阳性多核巨细胞中降钙素受体和破骨细胞相关基因的有效诱导。】 复制标题 收藏 收藏
    DOI:10.1111/j.1462-5822.2006.00807.x 复制DOI
    作者列表:Boddey JA,Day CJ,Flegg CP,Ulrich RL,Stephens SR,Beacham IR,Morrison NA,Peak IR
    BACKGROUND & AIMS: :Burkholderia pseudomallei is a facultative intracellular pathogen and the causative agent of melioidosis, a spectrum of potentially fatal diseases endemic in Northern Australia and South-East Asia. We demonstrate that B. pseudomallei rapidly modifies infected macrophage-like cells in a manner analagous to osteoclastogenesis. These alterations include multinucleation and the expression by infected cells of mRNA for factors required for osteoclastogenesis: the chemokines monocyte chemotactic protein 1 (MCP-1), macrophage inflammatory protein 1 gamma (MIP-1gamma), 'regulated on activation normal T cell expressed and secreted' (RANTES) and the transcription factor 'nuclear factor of activated T-cells cytoplasmic 1' (NFATc1). An increase in expression of these factors was also observed after infection with Burkholderia thailandensis. Expression of genes for the osteoclast markers calcitonin receptor (CTR), cathepsin K (CTSK) and tartrate-resistant acid phosphatase (TRAP) was also increased by B. pseudomallei-infected, but not by B. thailandensis-infected cells. The expression by B. pseudomallei-infected cells of these chemokine and osteoclast marker genes was remarkably similar to cells treated with RANKL, a stimulator of osteoclastogenesis. Analysis of dentine resorption by B. pseudomallei-induced osteoclast-like cells revealed that demineralization may occur but that authentic excavation does not take place under the tested conditions. Furthermore, we identified and characterized lfpA (for lactonase family protein A) in B. pseudomallei, which shares significant sequence similarity with the eukaryotic protein 'regucalcin', also known as 'senescence marker protein-30' (SMP-30). LfpA orthologues are widespread in prokaryotes and are well conserved, but are phylogenetically distinct from eukaryotic regucalcin orthologues. We demonstrate that lfpA mRNA expression is dramatically increased in association with macrophage-like cells. Mutation of lfpA significantly reduced expression of the tested host genes, relative to the response to wild-type B. pseudomallei. We also show that lfpA is required for optimal virulence in vivo.
    背景与目标: : pseudomalei伯克霍尔德菌是一种兼性细胞内病原体,也是类鼻疽病的病原体,类鼻疽是澳大利亚北部和东南亚流行的一系列潜在致命疾病。我们证明假单胞菌以类似于破骨细胞发生的方式快速修饰感染的巨噬细胞样细胞。这些改变包括破骨细胞形成所需的因子的多核和感染细胞的mRNA表达: 趋化因子单核细胞趋化蛋白1 (MCP-1),巨噬细胞炎性蛋白1 γ (MIP-1gamma),“调节活化正常T细胞表达和分泌” (RANTES) 和转录因子 “活化T细胞胞质核因子1” (NFATc1)。感染泰国伯克霍尔德菌后,还观察到这些因子的表达增加。破骨细胞标记物降钙素受体 (CTR),组织蛋白酶K (CTSK) 和耐酒石酸酸性磷酸酶 (TRAP) 的基因表达也被假假性芽孢杆菌感染的细胞增加,但未被泰国芽孢杆菌感染的细胞增加。这些趋化因子和破骨细胞标记基因的假假单胞菌感染细胞的表达与用破骨细胞生成刺激物RANKL处理的细胞非常相似。假单胞菌诱导的破骨细胞样细胞对牙本质的吸收分析表明,可能会发生脱矿质,但在测试条件下不会发生真正的挖掘。此外,我们在假单胞菌中鉴定并鉴定了lfpA (用于内酯酶家族蛋白A),该蛋白与真核蛋白 “regucalin” (也称为 “衰老标记蛋白-30” (SMP-30)) 具有显着的序列相似性。LfpA直系同源物在原核生物中广泛存在,并且保存良好,但在系统发育上与真核生物regucalcin直系同源物不同。我们证明lfpA mRNA表达与巨噬细胞样细胞相关显着增加。相对于对野生型B.Pseudomalei的反应,lfpA的突变显着降低了所测试宿主基因的表达。我们还表明,lfpA是体内最佳毒力所必需的。
  • 【MacMARCKS与代谢型谷氨酸受体7型相互作用,并调节g蛋白介导的钙通道组成型抑制。】 复制标题 收藏 收藏
    DOI:10.1111/j.1471-4159.2006.04121.x 复制DOI
    作者列表:Bertaso F,Lill Y,Airas JM,Espeut J,Blahos J,Bockaert J,Fagni L,Betz H,El-Far O
    BACKGROUND & AIMS: :We have previously shown that the interaction of Ca2+/calmodulin with the metabotropic glutamate receptor type 7 (mGluR7) promotes the G-protein-mediated inhibition of voltage-sensitive Ca2+ channels (VSCCs) seen upon agonist activation. Here, we performed a yeast two-hybrid screen of a new-born rat brain cDNA library using the cytoplasmic C-terminal tail of mGluR7 as bait and identified macrophage myristoylated alanine-rich c-kinase substrate (MacMARCKS) as a binding protein. The interaction was confirmed in vitro and in vivo by pull-down assays, immunoprecipitation, and colocalization of mGluR7 and MacMARCKS in transfected HEK293 cells and cultured cerebellar granule cells. Binding of MacMARCKS to mGluR7 was antagonized by Ca2+/calmodulin. In neurons, cotransfection of MacMARCKS with mGluR7, but not mGluR7 mutants unable to bind MacMARCKS, reduced the G-protein-mediated tonic inhibition of VSCCs in the absence of mGluR7 agonist. These results suggest that competitive interactions of Ca2+/calmodulin and MacMARCKS with mGluR7 control the tonic inhibition of VSCCs by G-proteins.
    背景与目标: : 我们以前已经表明,Ca2/钙调蛋白与代谢型谷氨酸受体7 (mGluR7) 的相互作用促进了g蛋白介导的对激动剂激活后看到的电压敏感Ca2通道 (vscc) 的抑制。在这里,我们使用mGluR7的细胞质C末端尾巴作为诱饵进行了新生大鼠脑cDNA文库的酵母双杂交筛选,并确定了巨噬细胞肉豆蔻基化的富含丙氨酸的c激酶底物 (MacMARCKS) 作为结合蛋白。通过下拉测定,免疫沉淀以及mGluR7和MacMARCKS在转染的HEK293细胞和培养的小脑颗粒细胞中的共定位,在体外和体内证实了相互作用。Ca2 +/钙调蛋白拮抗MacMARCKS与mGluR7的结合。在神经元中,在没有mGluR7激动剂的情况下,MacMARCKS与mGluR7共转染,但不能与MacMARCKS结合的mGluR7突变体,降低了g蛋白介导的VSCCs的强直抑制。这些结果表明,Ca2/钙调蛋白和MacMARCKS与mGluR7的竞争性相互作用控制了g蛋白对VSCCs的强直抑制。
  • 【对AUG和替代引发剂密码子的识别在4位的G中增加,但通常不受5位和6位核苷酸的影响。】 复制标题 收藏 收藏
    DOI:10.1093/emboj/16.9.2482 复制DOI
    作者列表:Kozak M
    BACKGROUND & AIMS: A primer extension (toeprinting) assay was used to monitor selection by ribosomes of the first versus the second AUG codon as a function of introducing mutations on the 3' side (positions +4, +5 and +6) of the first AUG codon. Six different flanking codons starting with G (GCG, GCU, GCC, GCA, GAU and GGA) strongly augmented selection of AUG#1 when compared with matched mRNAs that had A or C instead of G in position +4. Augmentation by G in position +4 failed only when it was combined with U in position +5, as in the sequence augGUA. In contrast with the usual enhancing effect of introducing G in position +4, most mutations in position +5 had no discernible effect, as shown with the series augANA (where N = C, A, G or U) and the series augCNA. AUG codon recognition was also unaffected by mutations in position +6, as shown by testing four mRNAs that had augCCN as the start site. Thus the primary sequence context that augments the recognition of AUG start codons does not appear generally to extend beyond G in position +4. When the toeprinting assay was used with mRNAs that initiate translation at CUG instead of AUG, cugGAU was not recognized better than cugGGU, contradicting the hypothesis that initiation at non-AUG codons might be favored by A instead of G in position +5.

    背景与目标: 引物延伸 (toeprinting) 测定法用于监测第一个AUG密码子与第二个AUG密码子的核糖体选择,这是在第一个AUG密码子的3' 侧 (位置4、5和6) 引入突变的函数。与在4位具有A或C而不是G的匹配mrna相比,以G开头的六个不同的侧翼密码子 (GCG,GCU,GCC,GCA,GAU和GGA) 强烈增强了AUG #1的选择。仅当G在位置4与U在位置5结合时,G的增强才失败,如序列augGUA所示。与通常在4位引入G的增强作用相反,5位的大多数突变没有明显的作用,如augANA系列 (其中N = C,A,G或U) 和augCNA系列所示。AUG密码子识别也不受6位突变的影响,如测试四个以augCCN为起始位点的mrna所示。因此,增强对AUG起始密码子的识别的主要序列上下文通常不会扩展到4位中的G以外。当toeprinting测定法与在CUG而不是AUG启动翻译的mrna一起使用时,cugGAU的识别并不比cugGGU更好,这与以下假设相矛盾: A而不是G在5位可能有利于非AUG密码子启动。
  • 【Interseptin调节表皮生长因子受体内吞作用,泛素化和信号传导。】 复制标题 收藏 收藏
    DOI:10.1124/mol.106.028274 复制DOI
    作者列表:Martin NP,Mohney RP,Dunn S,Das M,Scappini E,O'Bryan JP
    BACKGROUND & AIMS: :Receptor tyrosine kinases (RTKs) are critical for normal cell growth, differentiation, and development, but they contribute to various pathological conditions when disrupted. Activation of RTKs stimulates a plethora of pathways, including the ubiquitylation and endocytosis of the receptor itself. Although endocytosis terminates RTK signaling, it has emerged as a requisite step in RTK activation of signaling pathways. We have discovered that the endocytic scaffolding protein intersectin (ITSN) cooperated with epidermal growth factor receptor (EGFR) in the regulation of cell growth and signaling. However, a biochemical link between ITSN and EGFR was not defined. In this study, we demonstrate that ITSN is a scaffold for the E3 ubiquitin ligase Cbl. ITSN forms a complex with Cbl in vivo mediated by the Src homology (SH) 3 domains binding to the Pro-rich COOH terminus of Cbl. This interaction stimulates the ubiquitylation and degradation of the activated EGFR. Furthermore, silencing ITSN by RNA interference attenuated EGFR internalization as well as activation of the extracellular signal-regulated kinasemitogen-activated protein kinase pathway, thereby demonstrating the importance of ITSN in EGFR function. Given the cooperativity between ITSN and additional RTKs, these results point to an important evolutionarily conserved, regulatory role for ITSN in RTK function that is necessary for both signaling from receptors as well as the ultimate termination of receptor signaling.
    背景与目标: 受体酪氨酸激酶 (rtk) 对于正常的细胞生长,分化和发育至关重要,但是当它们被破坏时,它们会导致各种病理状况。RTKs的激活刺激了多种途径,包括受体本身的泛素化和内吞作用。尽管胞吞作用终止了RTK信号传导,但它已成为RTK激活信号传导途径的必要步骤。我们发现,内吞支架蛋白intersectin (ITSN) 与表皮生长因子受体 (EGFR) 配合调节细胞生长和信号传导。然而,ITSN和EGFR之间的生化联系尚未确定。在这项研究中,我们证明了ITSN是E3泛素连接酶Cbl的支架。ITSN通过与Cbl的富含前COOH末端结合的Src同源 (SH) 3结构域在体内与Cbl形成复合物。这种相互作用刺激活化的EGFR的泛素化和降解。此外,通过RNA干扰沉默ITSN减弱了EGFR的内在化以及细胞外信号调节的激酶原激活蛋白激酶途径的激活,从而证明了ITSN在EGFR功能中的重要性。鉴于ITSN和其他RTK之间的协同作用,这些结果表明ITSN在RTK功能中具有重要的进化保守调节作用,这对于受体的信号传导以及受体信号传导的最终终止都是必需的。
  • 【正常年轻和老年妇女肠道维生素d受体,钙吸收和血清1,25二羟基维生素d之间的关系。】 复制标题 收藏 收藏
    DOI:10.1359/jbmr.1997.12.6.922 复制DOI
    作者列表:Kinyamu HK,Gallagher JC,Prahl JM,DeLuca HF,Petranick KM,Lanspa SJ
    BACKGROUND & AIMS: The exact mechanism for the decrease in intestinal calcium absorption with age is not yet understood. A decrease with age in serum 1,25-dihydroxyvitamin D (1,25(OH)2D) or a decrease in the intestinal vitamin D receptor (VDR) protein concentration are possible causes. The objective of this study was to examine the effect of age on these factors. Fifty-nine young women age 25-35 years were compared with 41 elderly women age 65-83 years who underwent measurements of VDR, calcium absorption using a 20 mg and 100 mg calcium carrier, and calciotropic hormones. Calcium absorption by both tests was lower in the elderly women compared with the young women (p < 0.05). Serum 1,25(OH)2D and duodenal VDR protein concentration were not significantly different between the two age groups. Serum 1,25(OH)2D correlated with the 20 mg calcium absorption test in both young (r = 0.35, p < 0.007) and elderly women (r = 0.58, p < 0.0001) and with the 100 mg calcium absorption in the elderly (r = 0.32; p < 0.05). VDR did not correlate with calcium absorption in young women or elderly women, nor did VDR correlate with serum 1,25(OH)2D and serum 25-hydroxyvitamin D. In summary, the decrease in calcium absorption cannot be explained by a decrease in intestinal VDR. The correlation between serum 1,25(OH)2D and both calcium absorption tests only accounts for 12-30% of the variance in the age-related change in the calcium absorption tests. Other factors, not yet understood, are responsible for the decline in calcium absorption with age.

    背景与目标: 随着年龄的增长,肠道钙吸收减少的确切机制尚不清楚。血清1,25-二羟基维生素d (1,25(OH)2D) 随着年龄的增长而降低或肠道维生素d受体 (VDR) 蛋白浓度降低是可能的原因。这项研究的目的是检查年龄对这些因素的影响。将59名年龄在25-35岁之间的年轻女性与41名年龄在65-83岁之间的老年女性进行了比较,这些女性接受了VDR,使用20 mg和100 mg钙载体的钙吸收以及钙促激素的测量。与年轻女性相比,老年女性两种测试对钙的吸收均较低 (p <0.05)。血清1,25(OH)2D和十二指肠VDR蛋白浓度在两个年龄组之间没有显着差异。血清1,25(OH)2D与青年 (r = 0.35,p <0.007) 和老年妇女 (r = 0.58,p <0.0001) 的20 mg钙吸收试验相关,与老年人的100 mg钙吸收相关 (r = 0.32; p <0.05)。VDR与年轻女性或老年女性的钙吸收无关,也与血清1,25(OH)2D和血清25-羟基维生素d无关。总而言之,钙吸收的减少不能用肠道VDR的减少来解释。血清1,25(OH)2D与两种钙吸收测试之间的相关性仅占钙吸收测试中年龄相关变化的12-30%。其他尚未了解的因素是钙吸收随年龄下降的原因。

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