BACKGROUND & AIMS: :Under normoxic conditions the alpha-subunit of hypoxia-inducible factor (HIF-1alpha) protein is targeted for degradation by the von Hippel-Lindau (VHL) tumor suppressor protein acting as an E3 ubiquitin ligase. Recently, we developed a hypoxia-targeting protein, TOP3, which consisted of procaspase-3 with the VHL-mediated protein destruction motif of HIF-1alpha. This design enables procaspase-3 to be regulated similarly with HIF-1alpha, being degraded under normoxia while stabilized under hypoxia. Furthermore, stabilized TOP3 was cleaved by the hypoxic stress-induced endogenous caspases and thus the procaspase-3 was converted to active caspase-3 specifically under hypoxic conditions. These data demonstrated that the VHL-mediated protein destruction motif of HIF-1alpha endowed procaspase-3 with hypoxia-specific cytotoxicity.

背景与目标： 在常氧条件下，缺氧诱导因子（HIF-1alpha）蛋白的α亚基被von Hippel-Lindau（VHL）肿瘤抑制蛋白（作为E3泛素连接酶）降解。最近，我们开发了一种低氧靶向蛋白TOP3，该蛋白由具有HIF-1alpha的VHL介导的蛋白破坏基序的procaspase-3组成。这种设计使procaspase-3与HIF-1alpha相似，在常氧下降解，而在低氧下稳定。此外，稳定化的TOP3被低氧应激诱导的内源性半胱天冬酶裂解，因此procaspase-3特别是在低氧条件下转化为活性caspase-3。这些数据表明，HIF-1α的VHL介导的蛋白质破坏基序赋予了procaspase-3具有缺氧特异性细胞毒性。

BACKGROUND & AIMS: Recent evidence suggests that Chlamydia trachomatis can persist in the female upper genital tract in an unculturable state. Since unsuspected C. trachomatis infection has been associated with adverse in-vitro fertilization (IVF) outcome we sought to detect further evidence of C. trachomatis in the genital tracts of women undergoing IVF. The prevalence and distribution of antibodies to the major structural proteins of C. trachomatis in paired follicular fluid and sera of women undergoing IVF were examined. Sera and follicular fluid samples from 149 women were assayed for immunoglobulin (Ig)G and IgA antibodies to two C. trachomatis antigens, the major outer membrane protein (MOMP) and a recombinant lipopolysaccharide (rLPS) fragment. Additionally, the expression of human 60 kDa heat shock protein (hsp 60) in follicular fluid was determined. All cervical and follicular fluid samples were negative for C. trachomatis by polymerase chain reaction, ligase chain reaction and DNA probe. Sera from 60% of the subjects were positive for antichlamydial rLPS IgG; 36% were positive for anti-MOMP IgG. Similarly, rLPS-directed and MOMP-directed IgA were detected in sera of 34 and 14% of the subjects respectively. IgG antibodies to MOMP and rLPS were detected in 42 and 41% of the follicular fluid examined respectively. Anti-MOMP IgA was identified in 8.7% of the follicular fluid while 27.5% were positive for anti-rLPS IgA. Human hsp 60 expression was documented in 11.6% of the follicular fluid tested. IgA antibodies to both MOMP (P = 0.03) and rLPS (P = 0.02) in follicular fluid were associated with a failure to become pregnant after embryo transfer. IgG antibodies in sera and follicular fluid and IgA antibodies in sera were unrelated to IVF outcome. Similarly only anti-MOMP IgA (P = 0.02) and anti-rLPS IgA (P = 0.04) in follicular fluid were correlated with human hsp 60 expression in follicular fluid. The unique association between IgA antibodies to two chlamydial antigens in follicular fluid and both hsp 60 expression and IVF failure provides further support for the possibility that a persistent upper genital tract chlamydial infection contributes to IVF failure in some women.

背景与目标： 最近的证据表明，沙眼衣原体可以在女性上生殖道中以无法培养的状态持续存在。由于意外的沙眼衣原体感染与不良的体外受精（IVF）结果相关联，我们试图寻找接受IVF的女性生殖道中沙眼衣原体的进一步证据。检查了在进行IVF的妇女的成对卵泡液和血清中沙眼衣原体主要结构蛋白的抗体的分布情况。分析了来自149名妇女的血清和卵泡液样品中针对两种沙眼衣原体抗原，主要外膜蛋白（MOMP）和重组脂多糖（rLPS）片段的免疫球蛋白（Ig）G和IgA抗体。另外，测定了人60kDa热休克蛋白（hsp 60）在卵泡液中的表达。通过聚合酶链反应，连接酶链反应和DNA探针，所有子宫颈和卵泡液样品均为沙眼衣原体阴性。 60％的受试者血清抗衣原体rLPS IgG阳性； 36％的抗MOMP IgG阳性。同样，分别在34％和14％的受试者血清中检测到了rLPS导向和MOMP导向的IgA。在分别检查的卵泡液中有42％和41％检出了针对MOMP和rLPS的IgG抗体。在8.7％的卵泡液中发现了抗MOMP IgA，而抗rLPS IgA的阳性率为27.5％。在测试的卵泡液中有11.6％记录了人类hsp 60的表达。卵泡液中针对MOMP（P = 0.03）和rLPS（P = 0.02）的IgA抗体与胚胎移植后无法怀孕有关。血清和卵泡液中的IgG抗体以及血清中的IgA抗体与IVF结果无关。同样，卵泡液中只有抗MOMP IgA（P = 0.02）和抗rLPS IgA（P = 0.04）与人hsp 60在卵泡液中的表达相关。卵泡液中两种衣原体抗原的IgA抗体与hsp 60表达和IVF衰竭之间的独特联系为某些女性持续性上生殖道衣原体感染导致IVF衰竭的可能性提供了进一步的支持。

BACKGROUND & AIMS: The attachment protein or G protein of the A2 strain of human respiratory syncytial virus (RSV) was digested with trypsin and the resultant peptides separated by reverse-phase high-performance liquid chromatography (HPLC). One tryptic peptide produced a mass by matrix-assisted laser desorption/ionization (MALDI) time-of-flight (TOF) mass spectrometry (MS) corresponding to residues 152-187 with the four Cys residues of the ectodomain (residues 173, 176, 182, and 186) in disulfide linkage and absence of glycosylation. Sub-digestion of this tryptic peptide with pepsin and thermolysin produced peptides consistent with disulfide bonds between Cys173 and Cys186 and between Cys176 and Cys182. Analysis of ions produced by post-source decay of a peptic peptide during MALDI-TOF-MS revealed fragmentation of peptide bonds with minimal fission of an inter-chain disulfide bond. Ions produced by this unprecedented MALDI-induced post-source fragmentation corroborated the existence of the disulfide arrangement deduced from mass analysis of proteolysis products. These findings indicate that the ectodomain of the G protein has a non-glycosylated subdomain containing a "cystine noose."

背景与目标： 用胰蛋白酶消化人呼吸道合胞病毒（RSV）A2株的附着蛋白或G蛋白，并通过反相高效液相色谱（HPLC）分离得到的肽。一种胰蛋白酶肽通过基质辅助激光解吸/电离（MALDI）飞行时间（TOF）质谱（MS）产生质量，对应于带有胞外域的四个Cys残基的残基152-187（残基173、176， 182和186）中的二硫键和不存在糖基化作用。用胃蛋白酶和嗜热菌蛋白酶亚消化该胰蛋白酶消化的肽产生的肽与Cys173和Cys186之间以及Cys176和Cys182之间的二硫键一致。对在MALDI-TOF-MS过程中由消化性肽的源后降解产生的离子进行的分析显示，肽键断裂且链间二硫键的裂变最小。由这种前所未有的MALDI诱导的源后裂解产生的离子证实了从蛋白水解产物的质量分析推导出的二硫键结构的存在。这些发现表明，G蛋白的胞外域具有一个非糖基化的亚域，其中包含一个“胱氨酸套索”。

BACKGROUND & AIMS: The mammalian transcription factor LSF (CP2/LBP-1c) binds cellular promoters modulated by cell growth signals. We demonstrate here that LSF-DNA-binding activity is strikingly regulated by induction of cell growth in human peripheral T lymphocytes. Within 15 min of mitogenic stimulation of these cells, the level of LSF-DNA-binding activity increased by a factor of five. The level of LSF protein in the nucleus remained constant throughout this interval. However, a rapid decrease in the electrophoretic mobility of LSF, attributable to phosphorylation, correlated with the increase in DNA-binding activity. pp44 (ERK1) phosphorylated LSF in vitro on the same residue that was phosphorylated in vivo, specifically at amino acid position 291, as indicated by mutant analysis. As direct verification of the causal relationship between phosphorylation and DNA-binding activity, treatment in vitro of LSF with phosphatase both increased the electrophoretic mobility of the protein and decreased LSF-DNA-binding activity. This modulation of LSF-DNA-binding activity as T cells progress from a resting to a replicating state reveals that LSF activity is regulated during cell growth and suggests that LSF regulates growth-responsive promoters.

背景与目标： 哺乳动物转录因子LSF（CP2 / LBP-1c）结合受细胞生长信号调节的细胞启动子。我们在这里证明，LSF-DNA结合活性是由人类外周血T淋巴细胞的细胞生长诱导来显着调节的。在这些细胞的促有丝分裂刺激的15分钟内，LSF-DNA结合活性的水平提高了五倍。在整个间隔期间，核中LSF蛋白的水平保持恒定。但是，可归因于磷酸化的LSF电泳迁移率的快速下降与DNA结合活性的增加有关。突变分析表明，pp44（ERK1）在体外磷酸化的同一残基上，特别是在氨基酸位置291处磷酸化LSF。作为磷酸化与DNA结合活性之间因果关系的直接验证，体外用磷酸酶处理LSF既增加了蛋白质的电泳迁移率，又降低了LSF-DNA结合活性。当T细胞从静止状态发展到复制状态时，对LSF-DNA结合活性的这种调节揭示了LSF活性在细胞生长过程中受到调节，并暗示LSF调节生长响应性启动子。

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： ：Etanercept是一种重组人类可溶性肿瘤坏死因子（TNF-alpha）受体融合蛋白，可抑制TNF-alpha（一种在移植物抗宿主病（GVHD）发病机理中的主要介体）。本研究的目的是评估依那西普治疗21例激素抵抗性急性GVHD（aGVHD）（n = 13）和慢性GVHD（cGVHD）（n = 8）患者的安全性和有效性。每周两次皮下给予Etanercept 25 mg，持续4周，然后每周25 mg，持续4周。依那西普开始治疗时，有14例皮肤，13例胃肠，5例肝，5例肺，4例经口受累。 12名患者（57％）完成了12剂治疗。总体上，在21名患者中有11名患者（52％）对依那西普治疗有反应，包括6名患者（46％）患有aGVHD [n = 4完全缓解（CR），n = 2部分缓解（PR）]和5例（62 ％）和cGVHD（n = 1 CR，n = 4 PR）。临床反应最常见于顽固性肠aGVHD患者，其中55％的患者为CR，9％的患者为PR。 CMV重新激活发生在48％的患者中，细菌感染发生在14％的患者中，而真菌感染发生在19％的患者中。自依那西普开始接受中位随访429天（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死亡，2例疾病进展。总之，我们的初步数据表明，依那西普耐受性好，在类固醇难治性aGVHD和cGVHD患者中可引起较高的应答率，尤其是在胃肠道受累的情况下。

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

BACKGROUND & AIMS: PURPOSE:Peptide growth factors are known accelerators of corneal wound healing, probably mediated through increased proliferation of the cells; however, information about their effect on keratocyte motility is lacking. The influence of peptide growth factors on keratocyte migratory activity was investigated, using the following growth factors: platelet derived growth factor (PDGF-BB), epidermal growth factor (EGF), acidic fibroblast growth factor (aFGF), basic fibroblast growth factor (bFGF), insulin-like growth factor-I (IGF-I) and transforming growth factor-beta-1 (TGF-beta 1).

METHODS:Keratocytes were seeded on gels of type 1 collagen, growth factor added, and the cells left to migrate for 72 hours. Subsequently, the number of keratocytes at the different levels in the collagen gel was evaluated by optically sectioning the gel at 20 microns, intervals, with an inverted phase contrast microscope.

RESULTS:PDGF, EGF and bFGF at 10 ng/ml, all increased the number of keratocytes at the different levels of the gel as compared to a non-stimulated control (p < 0.05 or p < 0.01, students t-test). TGF-beta proved to be a strong inhibitor of keratocyte migration, decreasing the number of keratocytes observed at every level in the gel (p < 0.05 and p < 0.01, students t-test), whereas no effect of IGF-I and aFGF was found. During the 72 hours of migration, no contraction of the collagen gels was observed. Autoradiography of histological sections of the gels showed that during the 72-hour period only TGF-beta and 10% fetal bovine serum induced an increase in keratocyte proliferation.

CONCLUSION:PDGF, EGF and bFGF increase keratocyte migration, independent of proliferation in a collagen gel invasion assay and might promote corneal wound healing, not only by increasing cell proliferation, but also through increased motility.

背景与目标： 目的：肽生长因子是角膜伤口愈合的已知促进剂，可能是通过细胞增殖的增加来介导的。然而，缺乏有关它们对角膜细胞运动性影响的信息。使用以下生长因子研究了肽生长因子对角膜细胞迁移活性的影响：血小板衍生生长因子（PDGF-BB），表皮生长因子（EGF），酸性成纤维细胞生长因子（aFGF），碱性成纤维细胞生长因子（bFGF） ），胰岛素样生长因子I（IGF-I）和转化生长因子β-1（TGF-beta 1）。

方法：将角质形成细胞接种在1型胶原蛋白，添加了生长因子，细胞迁移了72小时。随后，通过倒置相差显微镜以20微米的间隔对凝胶进行光学切片，评估胶原蛋白凝胶中不同水平的角质形成细胞的数量。

结果：PDGF与未刺激的对照组相比，10 ng / ml的EGF和bFGF均增加了凝胶水平不同时的角膜细胞数量（p <0.05或p <0.01，学生t检验）。 TGF-β被证明是一种强烈的角膜细胞迁移抑制剂，可减少凝胶中各个水平上观察到的角膜细胞数量（p <0.05和p <0.01，学生t检验），而IGF-I和aFGF则没有作用成立。在迁移的72小时内，未观察到胶原凝胶的收缩。凝胶组织学切片的放射自显影显示，在72小时内，只有TGF-β和10％的胎牛血清诱导了角膜细胞增殖的增加。

结论：PDGF，EGF bFGF和bFGF可增加角膜细胞迁移，而不受胶原凝胶入侵试验中的增殖的影响，并且不仅可以通过增加细胞增殖，而且可以通过增加运动性来促进角膜伤口愈合。