Identification and establishment of sorting method for isolating CD11b+ myeloid cells in human hepatic metastases from colorectal cancer

AIM: To establish a method for obtaining specific cells in solid tumor tissue by sorting of CD11b⁺ myeloid cells in hepatic metastases from colorectal cancer.METHODS: Tumor tissues were prepared into single cell suspension by mechanical method combined with enzyme digestion, and then the CD11b⁺ myeloid cells were isolated by flow cytometry. The sorted cells were identified by immunocytochemistry. The viability and morphologiy of the sorted cells were evaluated by Giemsa and Typan blue staining. The cell purity was evaluated by flow cytometry.RESULTS: Sufficient numbers of CD11b⁺ cells with high purity were isolated by sorting with flow cytometry from the single cell suspension prepared by mechanical and enzyme digestion. The purity of the cells was confirmed by statistical analysis (P<0.05). The positive rates of the cells before and after sorting were significantly different (P<0.01). The positive cells were verified by immunocytochemical method. Meanwhile, the sorted cells had complete morphology and good activity.CONCLUSION: The CD11b⁺ myeloid cells in solid tumor tissue can be isolated by flow cytometry from the machine-enzyme digestion suspension with high purity, good activity and complete morphology.     

Effect of miR-29b-mediated TGF-β/Smad signaling pathway on progression of hepatic fibrosis in rats

AIM: To investigate the role of microRNA-29b (miR-29b)-mediated TGF-β/Smad signaling pathway in the activation of hepatic stellate cells (HSC) and its effect on the progression of hepatic fibrosis in rats.METHODS: Hepatic liver fibrosis rat model was established, and its HSC were isolated. Normal rat HSC were also obtained and identified in vitro. RT-qPCR and Western blot were used to detect the alterations of miR-29b, TGF-β/Smad signaling pathway-related proteins and liver fibrosis marker proteins in the acquired cells. Finally, the direct targeting binding of miR-29b to TGF-β1 was identified by dual-luciferase reporter assay system.RESULTS: With the activation of HSC, the expression of miR-29b gradually decreased (P<0.01), while the expression of collagen type I and α-smooth muscle actin gradually increased (P<0.01). At the same time, the expression of Smad2/3/4 was significantly increased, and the expression of Smad7 was significantly decreased (P<0.01). Dual-luciferase reporter assay showed that miR-29b bound directly to "UCUCUCCGU" in the 3'UTR of TGF-β1, indicating that TGF-β1 was a downstream target gene of miR-29b.CONCLUSION: miR-29b may be involved in the inhibition of HSC activation and migration, thereby inhibiting the process of liver fibrosis. The biological function of miR-29b may be through the direct targeting of TGF-β1, thus regulating and inhibiting the TGF-β/Smad signaling pathway.     

Microbial use of organic amendments in saline soils monitored by changes in the C/C ratio

An incubation experiment was carried out to investigate whether salinity at high pH has negative effects on microbial substrate use, i.e. the mineralization of the amendment to CO₂ and inorganic N and the incorporation of amendment C into microbial biomass C. In order to exploit natural differences in the ¹³C/¹²C ratio, substrate from two C₄ plants, i.e. highly decomposed and N-rich sugarcane filter cake and less decomposed N-poor maize leaf straw, were added to two alkaline Pakistani soils differing in salinity, which had previously been cultivated with C₃ plants. In soil 1, the additional CO₂ evolution was equivalent to 65% of the added amount in the maize straw treatment and to 35% in the filter cake treatment. In the more saline soil 2, the respective figures were 56% and 32%. The maize straw amendment led to an identical immobilization of approximately 48 μg N g⁻¹ soil over the 56-day incubation in both soils compared with the control soils. In the filter cake treatment, the amount of inorganic N immobilized was 8.5 μg N g⁻¹ higher in soil 1 than in soil 2 compared with the control soils. In the control treatment, the content of microbial biomass C₃-C in soil 1 was twice that in soil 2 throughout the incubation. This fraction declined by about 30% during the incubation in both soils. The two amendments replaced initially similar absolute amounts of the autochthonous microbial biomass C, i.e. 50% of the original microbial biomass C in soil 1 and almost 90% in soil 2. The highest contents of microbial biomass C₄-C were equivalent to 7% (filter cake) and 11% (maize straw) of the added C. In soil 2, the corresponding values were 14% lower. Increasing salinity had no direct negative effects on microbial substrate use in the present two soils. Consequently, the differences in soil microbial biomass contents are most likely caused indirectly by salinity-induced reduction in plant growth rather than directly by negative effects of salinity on soil microorganisms.     

叶甲科部分种类 Cyt b基因序列及系统发育关系初探

测定分析了叶甲科(Chrysomelidae)15种昆虫mtDNA_Cyt b基因部分序列,结果显示,在获得的390 bp序列中,198个核苷酸位点为多态性位点(约占50.8%),序列间碱基差异平均值为18.9%,A T含量为72.7%,碱基转换呈现明显的TC偏向性,颠换主要以TA为主,转换/颠换(R)值为0.8。依据分子数据建立了该15种昆虫的系统发育关系,结果表明:跳甲亚科(Alticinae)和叶甲亚科(Chrysomelinae)的亲缘关系较近,萤叶甲亚科(Galerucinae)与前两者的关系较远。萤叶甲亚科中瓢萤叶甲为最早分化的类群,长跗萤叶甲属(Monolepta)、凹翅萤叶甲属(Paleosepharia)、长刺萤叶甲属(Atrachya)能形成姊妹群。克萤叶甲属(Cneorane)和守瓜属(Aulacophora)的亲缘关系较近。基于蛋白质序列构建的系统树的置信值高于核苷酸序列建立的系统发育树。     

GM1 gangliosidosis in a Japanese domestic cat: a new variant identified in Hokkaido,Japan

A male Japanese domestic cat with retarded growth in Hokkaido, Japan, showed progressive motor dysfunction, such as ataxia starting at 3 months of age and tremors, visual disorder and seizure after 4 months of age. Finally, the cat died of neurological deterioration at 9 months of age. Approximately half of the peripheral blood lymphocytes had multiple abnormal vacuoles. Magnetic resonance imaging showed bisymmetrical hyperintensity in the white matter of the parietal and occipital lobes in the forebrain on T2-weighted and fluid-attenuated inversion recovery images, and mild encephalatrophy of the olfactory bulbs and temporal lobes. The activity of lysosomal acid β-galactosidase in leukocytes was negligible, resulting in the biochemical diagnosis of GM1 gangliosidosis. Histologically, swollen neurons characterized by accumulation of pale, slightly granular cytoplasmic materials were observed throughout the central nervous system. Dysmyelination or demyelination and gemistocytic astrocytosis were observed in the white matter. Ultrastructually, membranous cytoplasmic bodies were detected in the lysosomes of neurons. However, genetic analysis did not identify the c.1448G>C mutation, which is the single known mutation of feline GM1 gangliosidosis, suggesting that the cat was affected with a new variant of the feline disease.     

植物多肽激素PA1b的纯化鉴定及功能研究

通过稀醋酸低温浸提、海藻酸吸附、盐析、SephadexG25凝胶过滤、Sephacel离子交换层析和高效液相色谱等方法,从豌豆种子中分离纯化出PA1b及其多种异构体,并通过质谱和氨基酸测序进行了鉴定,结果表明:PA1b能抗胃蛋白酶(pepsin)水解,其高度的酶稳定性是由于分子内具有胱氨酸结构模体。此外,PA1b能明显延长体外培养的胰岛β细胞的存活时间,并影响胰岛β细胞的功能。     

Bioprospecting of Less-Polar Fractions of Ericaria crinita and Ericaria amentacea: Developmental Toxicity and Antioxidant Activity

Ericaria crinita and Ericaria amentacea from the Adriatic Sea (Croatia) were investigated with respect to the presence of less-polar compounds for the first time after fractionation by solid-phase extraction (SPE). The composition of less-polar fractions of freeze-dried E. crinita (FdEc) and E. amentacea (FdEa) were analyzed by high-performance liquid chromatography–high-resolution mass spectrometry with electrospray ionization (UHPLC-ESI-HRMS). The major identified compounds were: amides of higher aliphatic acids (palmitoleamide, linoleamide, palmitamide, oleamide and erucamide) and related compounds, carotenoid (fucoxanthin), chlorophyll derivatives (pheophytin a and b and their derivatives) and higher terpenes (loliolide, isoamijiol with its oxidation product), β-stigmasterol and (3β,6α)-14-methylergosta-8,24(28)-diene-3,6-diol). The toxic effects observed on the less-polar fractions obtained from Ericaria species on zebrafish Danio rerio embryos could be associated with the high abundance of all five detected amides. The antioxidant activity of the fractions was evaluated by means of five independent assays, including the reduction of the radical cation (ABTS), the oxygen radical absorbance capacity (ORAC), ferric-reducing antioxidant power (FRAP), the 2,2-diphenyl-1-picryl-hydrazyl (DPPH) assay and the Folin–Ciocalteu method. A higher antioxidant activity of E. amentacea in comparison to that of the E. crinita fractions was found with IC₅₀ concentrations of 0.072 and 1.177 mg/mL, respectively. The correlation between the activity and the chemical composition revealed that the synergistic effect of different compounds impacted their antioxidant response.     

四种鱚属鱼类线粒体Cyt b基因的序列变异及系统发育研究

测定和分析了4种鱚属(Sillago)鱼类:斑鱚(Sillago aeolus)、亚洲鱚(S. asiatica)、多鳞鱚(S. sihama)和少鳞鱚(S. japonica)线粒体细胞色素b基因序列片段,比较了不同鱚属鱼类种间的序列差异,探讨了彼此间系统发育关系和分类地位。结果显示,4种鱼类平均核苷酸组成为T 29.9%、C 29.2%、A 21.5%、G 19.3%,种间平均净遗传距离在0.157到0.280之间。最大似然法构建的系统树显示,少鳞鱚和亚洲鱚首先聚类、再与多鳞鱚和斑鱚相聚,斑鱚是最晚分化出的鱼类。基于Cyt b基因序列核苷酸分歧速率计算得出4种鱼类发生遗传分化时间在距今785—1400年前的第三纪中新世(Miocene)。