PEGylation of protein-based MRI contrast agents improves relaxivities and biocompatibilities

Magnetic resonance imaging (MRI) has emerged as a leading diagnostic technique in clinical and preclinical settings. However, the application of MRI to assess specific disease markers for diagnosis and monitoring drug effect has been severely hampered by the lack of desired contrast agents with high relaxivities, and optimized in vivo retention time. We have reported the development of protein-based MRI contrast agents (ProCA1) by rational design of Gd^3 + binding sites into a stable protein resulting in significantly increased longitudinal (r₁) and transverse (r₂) relaxivities compared to Gd-DTPA. Here, we report a further improvement of protein contrast agents ProCA1 for in vivo imaging by protein modification with various sizes of polyethylene glycol (PEG) chain. PEGylation results in significant increases of both r₁ and r₂ relaxivities (up to 200%), and these high relaxivities persist even at field strengths up to 9.4 T. In addition, our experimental results demonstrate that modified contrast agents have significant improvement of in vivo MR imaging and biocompatibilities including dose efficiency, protein solubility, blood retention time and decreased immunogenicity. Such improvement can be important to the animal imaging and pre-clinical research at high or ultra-high field where there is an urgent need for molecular imaging probes and optimized contrast agent.     

The acyl homoserine lactone receptor, SdiA, of Escherichia coli and Salmonella enterica serovar Typhimurium does not respond to indole

In this study, we tested the hypothesis that the SdiA proteins of Escherichia coli and Salmonella enterica serovar Typhimurium respond to indole. While indole was found to have effects on gene expression and biofilm formation, these effects were not sdiA dependent. However, high concentrations of indole did inhibit N-acyl-l-homoserine lactone (AHL) sensing by SdiA. We conclude that SdiA does not respond to indole but indole can inhibit SdiA activity in E. coli and Salmonella.     

The importance of craniofacial sutures in biomechanical finite element models of the domestic pig

Craniofacial sutures are a ubiquitous feature of the vertebrate skull. Previous experimental work has shown that bone strain magnitudes and orientations often vary when moving from one bone to another, across a craniofacial suture. This has led to the hypothesis that craniofacial sutures act to modify the strain environment of the skull, possibly as a mode of dissipating high stresses generated during feeding or impact. This study tests the hypothesis that the introduction of craniofacial sutures into finite element (FE) models of a modern domestic pig skull would improve model accuracy compared to a model without sutures. This allowed the mechanical effects of sutures to be assessed in isolation from other confounding variables. These models were also validated against strain gauge data collected from the same specimen ex vivo. The experimental strain data showed notable strain differences between adjacent bones, but this effect was generally not observed in either model. It was found that the inclusion of sutures in finite element models affected strain magnitudes, ratios, orientations and contour patterns, yet contrary to expectations, this did not improve the fit of the model to the experimental data, but resulted in a model that was less accurate. It is demonstrated that the presence or absence of sutures alone is not responsible for the inaccuracies in model strain, and is suggested that variations in local bone material properties, which were not accounted for by the FE models, could instead be responsible for the pattern of results.     

Ovarian cancer gene therapy using HPV-16 pseudovirion carrying the HSV-tk gene

Ovarian cancer is the leading cause of death from all gynecological cancers and conventional therapies such as surgery, chemotherapy, and radiotherapy usually fail to control advanced stages of the disease. Thus, there is an urgent need for alternative and innovative therapeutic options. We reason that cancer gene therapy using a vector capable of specifically delivering an enzyme-encoding gene to ovarian cancer cells will allow the cancer cell to metabolize a harmless prodrug into a potent cytotoxin, which will lead to therapeutic effects. In the current study, we explore the use of a human papillomavirus (HPV) pseudovirion to deliver a herpes simplex virus thymidine kinase (HSV-tk) gene to ovarian tumor cells. We found that the HPV-16 pseudovirion was able to preferentially infect murine and human ovarian tumor cells when administered intraperitoneally. Furthermore, intraperitoneal injection of HPV-16 pseudovirions carrying the HSV-tk gene followed by treatment with ganciclovir led to significant therapeutic anti-tumor effects in murine ovarian cancer-bearing mice. Our data suggest that HPV pseudovirion may serve as a potential delivery vehicle for ovarian cancer gene therapy.     

CONDITIONAL PRODUCTION OF A FUNCTIONAL FISH GROWTH HORMONE IN THE TRANSGENIC LINE OF NANNOCHLOROPSIS OCULATA (EUSTIGMATOPHYCEAE)1

Plasmid phr‐YPGHc, containing the fish growth hormone (GH) cDNA driven by a heat shock protein 70A promoter and a RUBISCO SSU 2 promoter, was transferred into the protoplast of marine microalga Nannochloropsis oculata (Droop) D. J. Hibberd by electroporation. Four transgenic clones were obtained in which the transferred phr‐YPGHc was integrated into the genome and existed stably at least until the 50th generation. When we treated these transgenic microalgae by heat shock, the heterologous fish GH was produced in the amount of 0.42 to 0.27 μg · mL^?1 from the 50 mL of medium. We incubated artemia with the wildtype and transgenic N. oculata for 6 h and then fed these microalgae‐treated artemia to red‐tilapia larvae. After feeding, the growth of larvae that were fed artemia incubated with transgenic microalgae was greater (i.e., statistically significant: P < 0.05) than that of larvae that were fed artemia incubated with nontransgenic microalgae: 316% versus 104% in weight gain, and 217% versus 146% in body length increase, respectively. Therefore, the N. oculata enables production of functional GH, and we propose that it might be an excellent bioreactor material.     

New costimulatory families: signaling lymphocytic activation molecule in adaptive allergic responses

The generation of an allergic immune response requires at least two signals for complete activation of T cells. Costimulatory molecules are integral to the second signal. In this review, we analyze the costimulatory molecule signaling lymphocytic activation molecule (SLAM) and other recently described SLAM family members. We highlight recent findings that position SLAM as critical for allergic inflammation and its role in modulation of cytokine secretion. Furthermore, a possible role of SLAM as a link between the adaptive and innate immune response is also discussed. Understanding the role of costimulatory molecules, including SLAM and SLAM family members, may elucidate mechanisms involved in the allergic immune response, and suggest potential therapeutic opportunities.     

C57BL/6 and BALB/c bronchoalveolar macrophages respond differently to exercise.

Macrophages from prototypical Th1 strains (e.g., C57BL/6) and Th2 strains (e.g., BALB/c) are classified as M-1 and M-2 phenotypes. We investigated the different phagocytic responses between M-1 and M-2 bronchoalveolar macrophages (BAMs) under resting and two various exercise conditions. At rest, M-1 BAMs showed higher phagocytic capacity of unopsonized particles, higher expression of MARCO (macrophage receptor with collagenous structure), and higher generation of NO than M-2 BAMs. Severe exercise, but not moderate exercise, significantly enhanced both phagocytosis of unopsonized particles and expression of MARCO in M-2 BAMs. In contrast, M-1 BAMs were unaffected by either exercise protocol. The phagocytosis of unopsonized particles was largely mediated by MARCO, especially in M-1 BAMs. Secreted products from cultured M-2 BAMs isolated after severe exercise, but not those from M-1 BAMs, enhanced BAM phagocytosis. The cultured M-1 BAMs secreted phagocytosis inhibitors, and this effect could be blocked by NO antagonists. Moreover, the extent of phagocytosis suppression induced by M-1 BAM-secreted products correlated with their production of nitrite/nitrate. Exogenous NO donors as well as NO derivatives, nitrite and nitrate, suppressed the BAM phagocytosis. We propose that while the severe exercise-enhanced phagocytosis in M-2 BAMs was largely mediated by MARCO up-regulation and secretion of stimulators, the lack of exercise effect in M-1 BAMs could be partially due to the constitutive secretion of NO-related suppressors. In conclusion, genetically different mice use different strategies in regulating BAM activity under resting conditions and in response to various exercise paradigms.     

锐化蝙蝠听皮层神经元频率调谐的柱特征

用双声刺激和多管电极方法在 6只大棕蝠 (bigbrownbat,Eptesicusfuscus)的 98个神经元上研究了锐化 (sharpening)蝙蝠听皮层 (primaryauditorycortex ,AC)神经元频率调谐的柱特征。结果发现 ,电极直插在 1个电极通道内连续记录到多个神经元时 ,它们锐化频率调谐的抑制性调谐曲线或抑制区基本相似。电极与AC表面呈 45°斜向推入使其跨越多个功能柱时 ,可观察到锐化频率调谐的抑制区构成也随电极进入不同的功能柱而发生相应的改变。两种不同的电极插入方式均证明锐化AC神经元频率调谐的神经抑制呈柱状组构。这些神经元组合起来排列在同一听觉功能柱内 ,构成AC频率分析的基本功能组构单位“微频率处理器”。实验中还观察到多峰频率调谐曲线神经元 ,它们在声通讯和声定位中不同波谱区域的时间匹配中起作用。此外 ,也有理由认为多峰调谐神经元亦被用于作为复杂波谱信息的“高级调谐预处理器” ,从而极大地提高了神经元对频率分析的能力。为研究锐化频率调谐的神经抑制机制 ,用多管电极电泳γ -氨基丁酸 (γ aminobutyricacid ,GA BA)能a受体拮抗剂荷包牡丹碱 (bicuculline ,Bic)至所记录的神经元 ,发现能大部分或几乎全部取消抑制区 ,从而表明在正常情况下GABA能抑制参与构成锐化AC神经元频率调谐的抑制区 ,