SSRP1 promotes colorectal cancer progression and is negatively regulated by miR‐28‐5p

In this study, microarray data analysis, real‐time quantitative PCR and immunohistochemistry were used to detect the expression levels of SSRP1 in colorectal cancer (CRC) tissue and in corresponding normal tissue. The association between structure‐specific recognition protein 1 (SSRP1) expression and patient prognosis was examined by Kaplan‐Meier analysis. SSRP1 was knocked down and overexpressed in CRC cell lines, and its effects on proliferation, cell cycling, migration, invasion, cellular energy metabolism, apoptosis, chemotherapeutic drug sensitivity and cell phenotype‐related molecules were assessed. The growth of xenograft tumours in nude mice was also assessed. MiRNAs that potentially targeted SSRP1 were determined by bioinformatic analysis, Western blotting and luciferase reporter assays. We showed that SSRP1 mRNA levels were significantly increased in CRC tissue. We also confirmed that this upregulation was related to the terminal tumour stage in CRC patients, and high expression levels of SSRP1 predicted shorter disease‐free survival and faster relapse. We also found that SSRP1 modulated proliferation, metastasis, cellular energy metabolism and the epithelial‐mesenchymal transition in CRC. Furthermore, SSRP1 induced apoptosis and SSRP1 knockdown augmented the sensitivity of CRC cells to 5‐fluorouracil and cisplatin. Moreover, we explored the molecular mechanisms accounting for the dysregulation of SSRP1 in CRC and identified microRNA‐28‐5p (miR‐28‐5p) as a direct upstream regulator of SSRP1. We concluded that SSRP1 promotes CRC progression and is negatively regulated by miR‐28‐5p.     

Rapid discrimination of colon cancer cells with single base mutation in KRAS gene segment using laser tweezers Raman spectroscopy

Laser tweezers Raman spectroscopy (LTRS) as a label‐free and noninvasive technology has been proven to be an ideal tool for analysis of single living cells, which provides important fingerprint information without interference from surrounding environments. For the first time, LTRS system was successfully used to examine the colon cancer cells with single base mutation in KRAS gene segment, including DKS‐8 (KRAS wild‐type [WT]) and DLD‐1 (KRAS mutant‐type [MT]), HKE‐3 (KRAS WT) and HCT‐116 (KRAS MT). Spectra changes of some vital biomolecules due to the gene mutation state were sensitively recorded by our home‐made LTRS system. As a result of the comparison between DKS‐8 and DLD‐1 cells, an index of 97.5% of correct classification was obtained by combining LTRS with principle component analysis coupled with linear discriminant analysis (PCA‐LDA) statistical analysis, while an index of 97.0% of correct classification was achieved between HKE‐3 and HCT‐116 cells. Moreover, between WT cells (DKS‐8 and HKE‐3) vs MT cells (DLD‐1 and HCT‐116), the index of correct classification was 81.2%, which was quite encouraging. Our preliminary results showed that the LTRS system coupled with PCA‐LDA analysis will have a great potential for further applications in the rapid and label‐free detection of circulating tumor cells in liquid biopsy.      

不同母岩发育马尾松土壤固氮菌群落结构和丰度特征

生物固氮是马尾松林地土壤氮素的重要来源,固氮微生物群落组成和数量的变化对土壤氮素供应和地力维持起重要作用。采用池栽试验,应用荧光定量PCR（聚合酶链式反应,Polymerase Chain Reaction）技术,借助Illumina Miseq高通量测序平台,以nifH基因为标靶,研究四类母岩（变余砂岩、长石石英砂岩、石英砂岩和玄武岩）发育马尾松土壤固氮菌群落结构和丰度的差异及其与土壤化学性质之间的关系。结果表明：（1）玄武岩土壤的有机碳、全氮、碱解氮、微生物量氮、马尾松株高和地径均显著高于其他母岩（P < 0.05）。（2）四类母岩土壤nifH基因丰度差异显著,玄武岩土壤nifH基因丰度分别是变余砂岩、石英砂岩和长石石英砂岩的3.75倍、7.89倍和4.41倍。（3）四类母岩土壤固氮菌群落α多样性指数（丰富度和多样性）差异显著,且玄武岩显著高于其他母岩。四类母岩发育土壤共获得有效序列159231条,分属于6个门、14个纲、41个目、69个科和122个属。门水平上,变形菌门和蓝藻门为主要优势类群。属水平上,慢生根瘤菌属、眉藻属、根瘤菌属和固氮螺菌属为主要优势属。玄武岩土壤变形菌门、慢生根瘤菌属、根瘤菌属和固氮螺菌属相对丰度显著高于其他母岩。层次聚类和非度量多维尺度（NMDS）分析结果表明,石英砂岩和长石石英砂岩土壤固氮菌群落结构相似,玄武岩与其他母岩固氮菌群落结构差异较大。（4）土壤有机碳、全氮、碱解氮和微生物量氮是固氮菌丰度、α多样性及群落结构的主要影响因子。综上,玄武岩土壤肥力高,提高了土壤固氮菌数量和群落多样性,有利于马尾松生长。研究从微生物学角度为马尾松适地造林和氮素调控提供了科学依据。     

益生菌混合发酵条件及其对发酵胡萝卜汁色泽及风味的影响

目的 研究3株益生菌混合发酵胡萝卜汁的发酵条件对色泽、风味的影响及其贮藏特性等.方法 通过菌种配比、单因素试验及正交试验优化了胡萝卜汁的发酵条件,并利用分光测色计和气质联用仪分别研究了发酵前后胡萝卜汁的风味成分和色泽变化.结果 添加30％ (m/m)的新鲜胡萝卜汁(原液),接种3％的以1∶1∶1(v/v/v)混合的嗜热...     

Iron increases liver injury through oxidative/nitrative stress in diabetic rats: Involvement of nitrotyrosination of glucokinase

Excessive tissue iron levels are associated with the increase of oxidative/nitrative stress which contributes to tissue damage that may elevate the risk of diabetes. Therefore, we investigated the effects of iron on diabetes-associated liver injury and whether iron-related tyrosine nitration participated in this process. Rats were randomly divided into four groups: control, iron overload (300 mg/kg iron dextran, i.p.), diabetic (35 mg/kg of streptozotocin i.p. after administration of a high-fat diet) and diabetic simultaneously treated with iron. Iron supplement markedly increased diabetes-mediated liver damage and hepatic dysfunction by increasing liver/body weight ratio, serum levels of aspartate and alanine aminotransferase, and histological examination, which were correlated with elevated levels of lipid peroxidation, protein carbonyls and tyrosine nitration, oxidative metabolism of nitric oxide, and reduced antioxidant capacity. Consequently, the extent of oxidized/nitrated glucokinase was markedly increased in the iron-treated diabetic rats that contribute to a decrease in its expression and activity. Further studies revealed a significant contribution of iron-induced specific glucokinase nitration sites to its inactivation. In conclusion, iron facilitates diabetes-mediated elevation of oxidative/nitrative stress, simultaneously impairs liver GK, and can be a link between enzymatic changes and hepatic dysfunction. These findings may provide new insight on the role of iron in the pathogenesis of diabetes mellitus.     

Cigarette smoke extract inhibits angiogenesis of pulmonary artery endothelial cells: the role of calpain

Angiogenesis is an integral part of both the pulmonary inflammatory response to chronic exposure to cigarette smoke and the lung tissue remodeling associated with cigarette smoke-induced chronic obstructive pulmonary disease (COPD). To investigate the role of angiogenesis in the pathogenesis of COPD, we evaluated the effect of cigarette smoke extract (CSE) on angiogenesis of pulmonary artery endothelial cells (PAEC). Incubation of PAEC with 2.5-10% CSE resulted in a dose-dependent inhibition of endothelial monolayer wound repair. CSE also caused inhibition of tube formation on Matrigel, migration in a Boyden chamber, and proliferation of PAEC. Because calpain, a family of calcium-dependent intracellular proteases, mediates cytoskeletal signaling in endothelial motility, we explored the role of calpain in the CSE-induced inhibition of endothelial angiogenesis. Incubation of CSE resulted in a dose-dependent decrease in calpain activity. Calpain inhibitor-1, a specific inhibitor of calpain, potentiates inhibitory effect of CSE on the endothelial monolayer wound repair, tube formation, cell migration, and cell proliferation. Transfection of PAEC with antisense oligodeoxyribonucleotides of calpastatin, the major endogenous calpain inhibitor, prevented CSE-induced increase in calpastatin protein content and CSE-induced decreases in calpain activity. It also prevented CSE-induced decreases in monolayer wound repair, tube formation, and migration. These results suggest that CSE attenuates angiogenesis of PAEC and the mechanism involves inhibition of calpain. Impaired angiogenesis may impede the repair process in the lungs of cigarette smokers and contribute to the altered structural remodeling observed in the lungs of patients with cigarette smoke-related COPD.     

The effective population size of Anopheles gambiae in Kenya: implications for population structure

We estimated current and long-term effective population size (Ne) of two Anopheles gambiae (savanna cytotype) populations in Kenya. Temporal variation at nine microsatellite loci in each population sampled 7 and 9 years apart and genetic diversity in each sample were analyzed to answer the following questions. (1) Do bottlenecks occur in Kenyan populations of A. gambiae? (2) How variable are different populations with respect to their current and long-term Ne values? (3) What are the implications of these results on population structure and history? The estimates of Ne of Asembo and Jego were 6,359 and 4,258, respectively, and the lower 95% limits were 2,455 and 1,669, respectively. Thus, despite the typical observation of low density at the village level during the dry season, large populations are maintained annually. Large current Ne is consistent with previous studies showing low differentiation across the continent, especially under Wright's isolation-by-distance model. Current Ne in Asembo was 1.5-fold higher than in Jego, but this difference was not significant. Long-term Ne in Asembo (22,667) was 2.9-fold higher than that in Jego (7,855) based on the stepwise mutation model. The difference between populations was significant at both time points regardless of whether long-term Ne values were calculated based on the stepwise mutation model or the infinite-alleles model. Heterozygosity in Jego declined significantly between 1987 (59%) and 1996 (54%), whereas heterozygosity in Asembo was stable (66%-65%). Despite the relatively high and significant differentiation between Asembo and Jego (FST = 0.072-0.10, RST = 0.037- 0.038), all alleles in Jego were found in Asembo but not vice versa. All of these findings suggest that lower Ne in Jego magnifies differentiation between the two populations. The long-term Ne was biased downward, because its calculation was based on an upper bound estimate of microsatellite mutation rate. Ne values based on mtDNA and allozymes were an order of magnitude higher. Long-term Ne therefore, is probably measured in hundreds of thousands and hence does not support a recent expansion of this species from a small population.      

Peripheral hyaline blebs (podosomes) of macrophages

The plasmalemma and hyaline ectoplasm together constitute the sensory and motor organ of macrophages. The purpose of this study was to isolate this cell fraction in order to analyze it biochemically and functionally. Brief sonification of warmed rabbit lung macrophages caused release of heterodisperse hyaline blebs and filopodia, which were easily collected by differential centrifugation. Viewed in the electron microscope, these structures consisted of membrane-bounded sacs principally containing actin filaments. Some contained secondary lysosomes. They were enriched threefold over whole cell homogenates in specific adenylate cyclase activity and in trichloroacetic-acid-precipitable (125)I when derived from cells labeled with 125(I) by means of a lactoperoxidase-catalyzed reaction. These markers were found to have identical isopycnic densitites when macrophage homogenates were subjected to sedimentation in a focusing sucrose density gradient system, and these markers had densities distinct from those of other cytoplasmic organelles. These markers were therefore assumed to be associated with macrophage plasma membranes. The specific β- glucuronidase activity of the bleb fraction was similar to that of homogenates, but the blebs had considerably lower specific succinic dehydrogenase activity and RNA content, and DNA was undetectable. Electrophoresis of blebs solubilized in sodium dodecyl sulfate on polyacrylamide gels revealed polypeptides co-migrating with macrophage actin-binding protein, myosin, and actin; blebs also had EDTA-activated adenosine triphosphatase activity characteristic of myosin. The concentrations of actin-binding protein and myosin were higher in blebs than in cells or cytoplasmic extracts, whereas actin concentrations were similar (relative to extracts) or only slightly greater (than in cells). Blebs and intact cells had high lactate dehydrogenase activities in the presence but not the absence of Triton X-100. Blebs and cells oxidased 1-[(14)C]glucose, and the rate of glucose oxidation was increased substantially in the presence of latex beads. We conclude that intact sacs of plasmalemma encasing contractile proteins and cytoplasmic enzymes can be isolated from macrophages. They are enriched in myosin and actin-binding protein, indicating that the contractile apparatus is regulated in the cell periphery. These structures have the capacity to respond to environmental signals. We suggest the name "podosomes" for them because of their resemblance to macrophage pseudopodia. We propose that podosome formation results from rapid dissolution of the cortical gel when the membrane is in an actively extended configuration.