Platelet derived growth factor (PDGF) contained in Platelet Rich Plasma (PRP) stimulates migration of osteoblasts by reorganizing actin cytoskeleton

Platelet-rich plasma (PRP) is a platelet concentrate in a small volume of plasma. It is highly enriched in growth factors able to stimulate the migration and growth of bone-forming cells. PRP is often used in clinical applications, as dental surgery and fracture healing. Platelet derived growth factor (PDGF), is highly concentrated in PRP and it was shown in our previous studies to provide the chemotactic stimulus to SaOS-2 osteoblasts to move in a microchemotaxis assay. Aim of the present studies is to analyze the effects of a PRP pretreatment (short time course: 30–150 min) of SaOS-2 cells with PRP on the organization of actin cytoskeleton, the main effector of cell mobility. The results indicate that a pretreatment with PRP increases chemokinesis and chemotaxis and concomitantly induces the organization of actin microfilaments, visualized by immunocytochemistry, in a directionally elongated phenotype, which is characteristic of the cells able to move. PRP also produces a transient increase in the expression of PGDF α receptor. This reorganization is blocked by the immunoneutralization of PDGF demonstrating the responsibility of this growth factor in triggering the mechanisms responsible for cellular movements.     

Microfluidic Platform for Measuring Neutrophil Chemotaxis from Unprocessed Whole Blood

Neutrophils play an essential role in protection against infections and their numbers in the blood are frequently measured in the clinic. Higher neutrophil counts in the blood are usually an indicator of ongoing infections, while low neutrophil counts are a warning sign for higher risks for infections. To accomplish their functions, neutrophils also have to be able to move effectively from the blood where they spend most of their life, into tissues, where infections occur. Consequently, any defects in the ability of neutrophils to migrate can increase the risks for infections, even when neutrophils are present in appropriate numbers in the blood. However, measuring neutrophil migration ability in the clinic is a challenging task, which is time consuming, requires large volume of blood, and expert knowledge. To address these limitations, we designed a robust microfluidic assays for neutrophil migration, which requires a single droplet of unprocessed blood, circumvents the need for neutrophil separation, and is easy to quantify on a simple microscope. In this assay, neutrophils migrate directly from the blood droplet, through small channels, towards the source of chemoattractant. To prevent the granular flow of red blood cells through the same channels, we implemented mechanical filters with right angle turns that selectively block the advance of red blood cells. We validated the assay by comparing neutrophil migration from blood droplets collected from finger prick and venous blood. We also compared these whole blood (WB) sources with neutrophil migration from samples of purified neutrophils and found consistent speed and directionality between the three sources. This microfluidic platform will enable the study of human neutrophil migration in the clinic and the research setting to help advance our understanding of neutrophil functions in health and disease.     

Isolation and Characterisation of Chemotactic Mutants of Chlamydomonas reinhardtii obtained by Insertional Mutagenesis

The swimming behaviour of the green flagellated protist Chlamydomonas reinhardtii is influenced by several different external stimuli including light and chemical attractants. Common components are involved in both the photo- and chemo-sensory transduction pathways, although the nature and organisation of these pathways are poorly understood. To learn more about the mechanism of chemotaxis in Chlamydomonas, we have generated nonchemotactic strains by insertional mutagenesis. The arginine-requiring strain arg7-8 was transformed with DNA carrying the wild-type ARG7 gene. Of the 8630 arginine-independent transformants obtained, five are defective in their chemotaxis towards various sugars. Two of the mutants (CTX2 and CTX3) are blocked only in their response to xylose. Mutant CTX1 is blocked in its response to xylose, maltose and mannitol, but displays normal taxis to sucrose. Mutants CTX4 and CTX5 lack chemotactic responses to all sugars tested. CTX1, CTX4 and CTX5 represent novel chemotactic phenotypes not previously obtained using ultra-violet or chemical mutagenesis. Genetic analysis confirms that each mutation maps to a single nuclear locus that is unlinked to the mating-type locus. Further analysis of CTX4 indicates that the mutant allele is tagged by the transforming ARG7 DNA. CTX4 appears to be defective in a component specific for chemotactic signal transduction since it exhibits wild-type photobehavioural responses (phototaxis and photoshock) as well as the wild-type responses of EGTA-induced trans-flagellum inactivation and acid-induced deflagellation. Insertional mutagenesis has thus permitted the generation of novel chemotactic mutants that will be of value in the molecular dissection of the signalling machinery.     

盐胁迫下粪产碱菌在水稻根表的定殖和异源固氮基因的表达

异源nif LacZ融合基因在粪产碱菌A15 6 1中的表达活性随盐浓度增加而升高 ,然后逐渐降低 ,nifH LacZ融合基因可以正常表达的盐浓度在 0 .1%～0 .5 %之间 ,盐浓度为 0 .0 5 %时活性最高。A15 6 1在盐浓度为 0 .0 6 %时趋化能力最强 ,随着盐浓度的提高逐渐下降 ,当盐浓度为 3 .0 %时完全丧失趋化能力。一定的盐浓度 (0 .5 % )对固氮粪产碱菌的根表定殖有促进作用 ,该条件下根表定殖的菌体数远大于对照。 3种nif LacZ融合基因在根内的表达部位有显著差异。nifH的表达部位主要分布于根的皮层薄壁组织细胞间隙 ,在条件适宜 (无铵和微量氧 )的部位或某些特殊位置如侧根伸出部位高水平表达。盐胁迫下水稻 耐盐粪产碱菌A15 6 1的联合固氮效率明显高于A15 6 1纯培养物     

鱼类趋化因子的研究进展

趋化因子(Chemokine)是由多种细胞在致病因子刺激后分泌的一类低分子量的细胞因子,它们都具有激活和趋化白细胞的作用。趋化因子从结构上可分为四类:CC型、CXC型、CX3C型和C型;从功能上可分为两种类型:一类主要诱导白细胞到炎症部位;另一类主要是对肌体起免疫监控作用。目前,有关鱼类趋化因子的研究主要集中于CXC型和CC型两类,以及其在非特异性免疫中的作用。     

HGF/SF Induces Mesothelial Cell Migration and Proliferation by Autocrine and Paracrine Pathways

Mesothelial repair differs from that of other epithelial-like surfaces as healing does not occur solely by centripetal in-growth of cells as a sheet from the wound margins. Mesothelial cells lose their cell-cell junctions, divide, and adopt a fibroblast-like morphology while scattering across and covering the wound surface. These features are consistent with a cellular response to hepatocyte growth factor/scatter factor (HGF/SF). In this study, we examined the ability of mesothelial cells to secrete HGF/SF and investigated its possible role as an autocrine regulator of mesothelial cell motility and proliferation. We found that human primary mesothelial cells expressed HGF/SF mRNA and secreted active HGF/SF into conditioned medium as determined by ELISA and in a scattering bioassay. These cells also expressed the HGF/SF receptor, Met, as shown by RT-PCR and by Western blot analysis and immunofluorescence. Incubation of mesothelial cells with neutralizing antibodies to HGF/SF decreased cell migration to 25% of controls, whereas addition of HGF/SF disrupted cell-cell junctions and induced scattering and enhanced mesothelial cell migration. Furthermore, HGF/SF showed a small but significant mitogenic effect on all mesothelial cell lines examined. In conclusion, HGF/SF is produced by mesothelial cells and induces both motility and proliferation of these cells. These data are consistent with HGF/SF playing an autocrine role in mesothelial healing.     

Study of the oxidative metabolic function and chemotaxis of neutrophils from patients with cancer influenced by Selenium Yeast

The effects of Selenium Yeast (SY) given orally to patients with cancer on the oxidative metabolic function and chemotaxis of neutrophils were studied by Lumino-dependent Chemiluminescence (CL) assay and agarose plates assay, respectively. The results showed that the background CL and the peak value CL of neutrophils from patients with cancer 14 d after taking SY were, on the average, 4 and 1.2 times higher than those before taking SY. The chemotactic index (CI) and chemotactic differential (CD) increased in response to SY supplementation; the differences are statistically significant (p less than 0.01), and no corresponding differences could be seen in the control group (taking yeast).     

Interaction Between Neutrophils and 4-Hydroxyalkenals and Consequences on Neutrophil Motility

The lipid peroxidation product 4-hydroxynonenal (HNE) and homologous aldehydes have been found to possess chemotactic activity for rat neutrophil leukocytes in the micromolar to picomolar range, depending on the compound. Such an activity is displayed only in the presence of albumin. The mechanisms by which aldehydes could interact with neutrophils are discussed. II is proposed that albumin acts as a carrier for the aldehyde and releases them to a neutrophil receptor. At concentrations around 10^-4M, 4-hydroxyal-kenals have been found to exert toxic effects on a number of cells, including a strong depression of neutrophil motility. Finally, HNE has been found at chemotactic concentrations in the inflammatory site. The possibility that HNE is involved in the neutrophil influx into the inflammatory site is considered.     

糖皮质激素对肺泡巨噬细胞趋化活性的影响

采用测定肺泡巨噬细胞(AM)趋化性和AM产生中性粒细胞趋化因子的实验技术。观察到糖皮质激素体外直接作用于大鼠AM悬液或大鼠皮下注射7d都可抑制AM的趋化性,并且肺灌洗液中的AM数量明显减少;但注射3d组无上述效应。以糖皮质激素处理AM 16h可使AM产生的中性粒细胞趋化因子强度明显降低。结果表明糖皮质激素可抑制AM的趋化性和抑制AM产生中性粒细胞趋化因子。提示这可能是较长期应用糖皮质激素抑制免疫和炎症反应、降低肺部防御能力的部分机理。