一种新型免疫层析技术在临床乳腺癌Her-2蛋白定量检测中的应用

建立一种靶点蛋白质快速定量检测方法。在原有侧向流动免疫层析技术的基础上,通过优化层析材料和纳米微球的均一性、改进检测区的检测方法,经逐点扫描技术,建立标准浓度曲线,以达到对临床靶点蛋白质的定量检测。以乳腺癌组织中的Her2表达为例,通过对已知浓度样品的检测,验证本技术方法的准确度大于96%。另外,以蛋白质免疫印迹作为组织中特定蛋白质检测金标准,分析临床肿瘤组织中Her2蛋白的含量,其准确率也达到95.5%,而免疫组织化学方法检测准确率仅为69.58%。新型免疫层析法检测结果与靶向治疗患者的愈后密切相关(P<0.01)。改进后的新型免疫层析方法能够准确地对临床靶点蛋白质进行定量检测,而且结合侧向流动技术的简单、快速和易用性,这种新型检测方法可以广泛应用于临床组织标本、血液标本和体液标本中靶点蛋白质的临场定量检测,在一定程度上可以替代免疫组化技术。     

De novo organogenesis and plant regeneration in <Emphasis Type="Italic">Pongamia pinnata</Emphasis>, oil producing tree legume

Role of Thidiazuron (TDZ) in inducing adventitious organogenesis in Pongamia was studied. TDZ at different concentrations (0, 0.45, 2.27, 4.54, 6.71, 9.08, 11.35, 13.12 and 22.71 μM) were used for induction of caulogenic bud formation in deembryonated cotyledon explants. Each cotyledon was cut into three segments and identified as proximal, middle and distal. Duration of TDZ exposure, influence of the segment and orientation of the explant were studied. TDZ at 11.35 μM concentration was optimum for the induction of shoots and rapid elongation. Shoots induced at higher concentration elongated after several passages in growth regulator free medium, thereby extending the period of differentiation. Exposure of the explant for 20 days yielded more number of buds than 10 days. Proximal segment of the cotyledon was more responsive. Contact of abaxial surface in the medium was more effective and generated more buds than the adaxial side. Buds differentiated and elongated on transfer to MS basal medium for 8–12 passages of 15 days each. Rooting and elongation of shoots was achieved in charcoal supplemented half-strength MS medium. Rooted plantlets survived on transfer to sand soil mixture. The plants were hardened and transferred to green house. This is the first report on in vitro regeneration of Pongamia pinnata via adventitious organogenesis using TDZ. This protocol may find application in studies in genetic transformation, isolation of somaclonal variants and in induction of mutants. It also provides a system to study the inhibitory role of TDZ on shoot differentiation.     

多西紫杉醇修饰的人工晶体与眼组织相容性分析

目的：探讨经多西紫杉醇修饰的人工晶体对眼组织相容性的影响。方法：按照随机数字表法将32 只日本大耳兔分为两组： 实验组通过手术植入表面经多西紫杉醇修饰处理后的疏水性人工晶体，对照组植入疏水性人工晶体。比较两组人工晶体亲水角、 术后24 小时光耀斑块计数以及人工晶体周围组织炎症浸润数。结果：实验组的亲水角小于对照组，差异有统计学意义（P<0.05）。 实验组光耀斑块计数低于对照组，差异有统计学意义（P<0.05）。实验组家兔人工晶体周围组织炎症浸润计数低于对照组，差异有 统计学意义（P<0.05）。结论：人工晶体表面经多西紫杉醇修饰后，其亲水性、与眼组织的组织相容性增加，且可缓解光耀斑炎症感 染和降低并发症的发生，有重要的临床参考价值。     

Probing binding mechanism of interleukin-6 and olokizumab: in silico design of potential lead antibodies for autoimmune and inflammatory diseases

Computer-aided antibody engineering has been successful in the design of new biologics for disease diagnosis and therapeutic interventions. Interleukin-6 (IL-6), a well-recognized drug target for various autoimmune and inflammatory diseases such as rheumatoid arthritis, multiple sclerosis, and psoriasis, was investigated in silico to design potential lead antibodies. Here, crystal structure of IL-6 along with monoclonal antibody olokizumab was explored to predict antigen–antibody (Ag???Ab)-interacting residues using DiscoTope, Paratome, and PyMOL. Tyr56, Tyr103 in heavy chain and Gly30, Ile31 in light chain of olokizumab were mutated with residues Ser, Thr, Tyr, Trp, and Phe. A set of 899 mutant macromolecules were designed, and binding affinity of these macromolecules to IL-6 was evaluated through Ag???Ab docking (ZDOCK, ClusPro, and Rosetta server), binding free-energy calculations using Molecular Mechanics/Poisson Boltzman Surface Area (MM/PBSA) method, and interaction energy estimation. In comparison to olokizumab, eight newly designed theoretical antibodies demonstrated better result in all assessments. Therefore, these newly designed macromolecules were proposed as potential lead antibodies to serve as a therapeutics option for IL-6-mediated diseases.     

Stress-shielding induced bone remodeling in cementless shoulder resurfacing arthroplasty: a finite element analysis and in vivo results

Cementless surface replacement arthroplasty (CSRA) of the shoulder was designed to preserve the individual anatomy and humeral bone stock. A matter of concern in resurfacing implants remains the stress shielding and bone remodeling processes. The bone remodeling processes of two different CSRA fixation designs, conical-crown (Epoca RH) and central-stem (Copeland), were studied by three-dimensional (3-D) finite element analysis (FEA) as well as evaluation of contact radiographs from human CSRA retrievals. FEA included one native humerus model with a normal and one with a reduced bone stock quality. Compressive strains were evaluated before and after virtual CSRA implantation and the results were then compared to the bone remodeling and stress-shielding pattern of eight human CSRA retrievals (Epoca RH n=4 and Copeland n=4). FEA revealed for both bone stock models increased compressive strains at the stem and outer implant rim for both CSRA designs indicating an increased bone formation at those locations. Unloading of the bone was seen for both designs under the central implant shell (conical-crown 50–85%, central-stem 31–93%) indicating high bone resorption. Those effects appeared more pronounced for the reduced than for the normal bone stock model. The assumptions of the FEA were confirmed in the CSRA retrieval analysis which showed bone apposition at the outer implant rim and stems with highly reduced bone stock below the central implant shell. Overall, clear signs of stress shielding were observed for both CSRAs designs in the in vitro FEA and human retrieval analysis. Especially in the central part of both implant designs the bone stock was highly resorbed. The impact of these bone remodeling processes on the clinical outcome as well as long-term stability requires further evaluation.     

BMP signaling in vascular biology and dysfunction

The vascular system is critical for developmental growth, tissue homeostasis and repair but also for tumor development. Bone morphogenetic protein (BMP) signaling has recently emerged as a fundamental pathway of the endothelium by regulating cardiovascular and lymphatic development and by being causative for several vascular dysfunctions. Two vascular disorders have been directly linked to impaired BMP signaling: pulmonary arterial hypertension and hereditary hemorrhagic telangiectasia. Endothelial BMP signaling critically depends on the cellular context, which includes among others vascular heterogeneity, exposure to flow, and the intertwining with other signaling cascades (Notch, WNT, Hippo and hypoxia). The purpose of this review is to highlight the most recent findings illustrating the clear need for reconsidering the role of BMPs in vascular biology.     

The effect of calcium and vitamin D supplementation on osteoporotic rabbit bones studied by vibrational spectroscopy

Fourier transform infrared spectroscopy is utilized to examine the effects of increased calcium, vitamin D, and combined calcium-vitamin D supplementation on osteoporotic rabbit bones with induced inflammation. The study includes different bone sites (femur, tibia, humerus, vertebral rib) in an effort to explore possible differences among the sites. We evaluate the following parameters: mineral-to-matrix ratio, carbonate content, and non-apatitic species (labile acid phosphate and labile carbonate) contribution to bone mineral. Results show that a relatively high dose of calcium or calcium with vitamin D supplementation increases the bone mineralization index significantly. On the other hand, vitamin D alone is not as effective in promoting mineralization even with high intake. Mature B-type apatite was detected for the group with calcium supplementation similar to that of aged bone. High vitamin D intake led to increased labile species concentration revealing bone formation. This is directly associated with the suppression of pro-inflammatory cytokines linked to induced inflammation. The latter is known to adversely alter bone metabolism, contributing to the aetiopathogenesis of osteoporosis. Thus, a high intake of vitamin D under inflammation-induced osteoporosis does not promote mineralization but suppresses bone resorption and restores metabolic balance.     

Bone marrow-derived mesenchymal stem cells repair severe acute pancreatitis by secreting miR-181a-5p to target PTEN/Akt/TGF-β1 signaling

BackgroundSevere acute pancreatitis (SAP) is associated with high morbidity and mortality. Bone marrow mesenchymal stem cells (BMSCs) have shown obvious protective effect on SAP. However, little is known about the underlying mechanism. The objective of this study is to unravel the role and regulatory mechanism of miR-181a-5p in BMSCs-mediated pancreatic repair.MethodsBMSCs were isolated from Sprague-Dawley rats and characterized by flow cytometry and Oil Red O staining. Sodium taurocholate- and caerulein-induced models were used as SAP models in vivo and in vitro, respectively. Pancreatic injury were evaluated by H&E and histopathological analysis, as well as by measuring levels of amylase, lipase and cytokines. qRT-PCR and western blotting were performed to detect the level of miR-181a-5p and the protein levels of PTEN/Akt, respectively. ELISA was conducted to detect the levels of TNF-α, IL-1β, IL-6, angiopoietin, IL-4, IL-10 and TGF-β1. The apoptotic rate of AR42 J cells was quantitated by concurrent staining with Annexin-V-FITC and PI.ResultsBMSCs significantly attenuated pancreatic injury in SAP rats by reducing inflammatory infiltration and necrosis, and this effect was abolished by CXCR4 agonist AMD3100. ADM3100 exhibited more severe pancreatic injury and decreased miR-181a-5p levels in the pancreas and serum compared to SAP group. Overexpression of miR-181a-5p in BMSCs (BMSCs-miR-181a-5p) markedly potentiated the protective effect of BMSCs by reducing histological damage and levels of amylase and lipase. Moreover, BMSCs-miR-181a-5p dramatically reduced levels of angiopoietin, TNF-α, IL-1β and IL-6, but induced the levels of IL-4 and IL-10. In caerulein-treated AR42 J cells, co-culturing of BMSCs-miR-181a-5p alleviated caerulein-induced increase of amylase and lipase, and apoptosis via PTEN/Akt/TGF-β1 signaling.ConclusionBMSCs alleviate SAP and reduce inflammatory responses and apoptosis by secreting miR-181a-5p to target PTEN/Akt/TGF-β1 signaling. Hence, BMSCs-miR-181a-5p could serve as potential therapeutic target for SAP.     

Nitrate induced iron deficiency chlorosis in Juncus acutiflorus

Chlorosis caused by iron deficiency is commonly associated with high bicarbonate levels in the soil. However, in rare cases such chlorosis has been observed in soils with high nitrate levels. In a dutch rich-fen, chlorosis has been noted in stands of Juncus acutiflorus at locations where groundwater containing high levels of nitrate reached the surface. Experiments revealed that the chlorosis could be attributed to iron deficiency although iron levels in the shoots were well above the known physiological threshold values for iron deficiency. It is postulated that increased nitrate assimilation leads to an increased apoplastic pH and to a concomitant immobilisation of iron and/or lower iron (III) reduction. Moreover free amino acid levels were markedly higher in the iron deficient plants in the field. It was found, however, that the percentage of nitrogen present as free amino acids was not influenced directly by low iron levels but mainly by the C/N ratios in the shoots. Nowadays, nitrate concentrations in ground water as high 1000 µM are no longer an exception in the Netherlands. We propose that strongly increased nitrate inputs may cause iron stress in natural vegetations, especially in wet habitats.     

Direct somatic embryogenesis from axes of mature peanut embryos

Summary Plant regeneration via somatic embryogenesis was obtained in peanut (Arachis hypogaea L.) from axes of mature zygotic embryos. The area of greatest embryogenic activity was a 2-mm region adjacent to and encircling the epicotyl. Somatic embryogenesis was evaluated on Murashige and Skoog media supplemented with a variety of auxin treatments. Maximum production occurred on medium supplemented with 3 mg · liter⁻¹ 4-amino-3,5,6-trichloropicolinic acid. Explant cultures were transferred to half-strength medium supplemented with 1 mg · liter⁻¹ gibberellic acid for somatic embryo germination and early plantlet growth. Plantlets, transferred to soil, were placed in a greenhouse and grown to maturity.