Molecular phylogeny reveals independent origins of body scales in Entomobryidae (Hexapoda: Collembola).

Entomobryidae is the largest family in Collembola but relationships within the family have never been subjected to rigorous phylogenetic analyses. Within the family, body scales are present in many species, and are fundamental in the classification at the subfamilial and tribal levels. A molecular phylogeny was reconstructed using the nuclear 18SrRNA and partial 28SrRNA and the mitochondrial 16SrRNA to examine the evolution of scales across Entomobryidae subfamilies. These datasets were analyzed separately and combined, with parsimony, likelihood and Bayesian algorithms. Monophyly of Orchesellinae was not recovered, and it was split into a scaled clade and an unscaled clade, contradicting to all recent taxonomic conceptions. The monophyly of Entomobryinae, Seirinae and Lepidocyrtinae is well supported however within Entomobryinae, the polyphyly of Entomobryini and Willowsiini implies that classification using the presence/absence of scales is not valid. Analyses of ancestral character state reconstruction in Entomobryidae indicate that the presence of body scales have evolved independently at least five times, with a loss of scales occurring independently at least twice. A revision of the family Entomobryidae on molecular and morphological basis is clearly needed.

Stanniocalcin-1 promotes tumor angiogenesis through up-regulation of VEGF in gastric cancer cells.

BACKGROUND: Stanniocalcin-1(STC-1) is up-regulated in several cancers including gastric cancer. Evidences suggest that STC-1 is associated with carcinogenesis and angiogenic process. However, it is unclear on the exact role for STC-1 in inducing angiogenesis and tumorigeneisis. METHOD: BGC/STC cells (high-expression of STC-1) and BGC/shSTC cells (low- expression of STC-1) were constructed to investigate the effect of STC-1 on the xenograft tumor growth and angiogenesis in vitro and in vivo. ELISA assay was used to detect the expression of vascular endothelial growth factor (VEGF) in the supernatants. Neutralizing antibody was used to inhibit VEGF expression in supernatants. The expression of phosphorylated -PKCßII, phosphorylated -ERK1/2 and phosphorylated -P38 in the BGC treated with STC-1protein was detected by western blot. RESULTS: STC-1 could promote angiogenesis in vitro and in vivo, and the angiogenesis was consistent with VEGF expression in vitro. Inhibition of VEGF expression in supernatants with neutralizing antibody markedly abolished angiogenesis induced by STC-1 in vitro. The process of STC-1-regulated VEGF expression was mediated via PKCßII and ERK1/2. CONCLUSIONS: STC-1 promotes the expression of VEGF depended on the activation of PKCßII and ERK1/2 pathways. VEGF subsequently enhances tumor angiogenesis which in turn promotes the gastric tumor growth.

Determination of trace deoxyribonucleic acid by using fluorescein isothiocyanate-phenosafranine as a double-luminescent phosphorescence probe.

Using Pb(2+) as ion perturber, phenosafranine (PF) and fluorescein isothiocyanate (FITC) could emit strong and stable room temperature phosphorescence (RTP) signal on the filter paper, respectively. When they were mixed, the phenomenon that the RTP signal of PF and FITC enhanced significantly was found. And 1.12 ag DNA spot(-1) (sample volume was 0.40 µL, corresponding concentration was 2.8 × 10(-15) g mL(-1)) could cause the RTP signal of both PF and FITC to enhance sharply. The content of DNA was proportional to the ΔI(p) of PF and FITC in the system at 634 and 659 nm. Thus, a new solid substrate room temperature phosphorimetry (SSRTP) for the determination of trace DNA was established by using FITC-PF as double-luminescent phosphorescence probe. The detection limit (LD) of this method calculated by 3S(b)/k was 14 zg DNA spot(-1) for PF and 18 zg DNA spot(-1) for FITC, respectively, showing high sensitivity. It has been applied to the determination of trace DNA in practical samples and the analysis results were in accordance with those of fluorescence probe. The reaction mechanism of SSRTP for the determination of trace DNA was also discussed.

Over-expressed and truncated midkines promote proliferation of BGC823 cells in vitro and tumor growth in vivo.

AIM: To determine whether midkine (MK) and its truncated form (tMK) contribute to gastric tumorigenesis using in vitro and in vivo models. METHODS: Human MK and tMK plasmids were constructed and expressed in BGC823 (a gastric adenocarcinoma cell line) to investigate the effect of over-expressed MK or tMK on cell growth and turmorigenesis in nude mice. RESULTS: The growth of MK-transfected or tMK-transfected cells was significantly increased compared with that of the control cells, and tMK-transfected cells grew more rapidly than MK-transfected cells. The number of colony formation of the cells transfected with MK or tMK gene was larger than the control cells. In nude mice injected with MK-transfected or tMK-transfected cells, visible tumor was observed earlier and the tumor tissues were larger in size and weight than in control animals that were injected with cells without the transfection of either genes. CONCLUSION: Over-expressed MK or tMK can promote human gastric cancer cell growth in vitro and in vivo, and tMK has greater effect than MK. tMK may be a more promising gene therapeutic target compared with MK for treatment of malignant tumors.

[Cloning and expression of truncated Midkine cytokine from gastric carcinogenesis tissue in E. coli].

OBJECTIVE: To clone the truncated Midkine from gastric carcinogenesis tissue and express in Escherichia coli. METHODS: A pair of PCR specific primers were designed according to the reported human tMK cDNA sequence in Genbank. The target DNA fragment was obtained by RT-PCR from gastric carcinoma patient's carcinogenesis tissue and cloned into pMD 18 T-vector. After sequencing, the tMK nucleotide fragment was inserted into an E. coli expression vector pBV222. The recombinant plasmid was transferred into E. coli DH5alpha and an E. coli DH5alpha expressed recombinant tMK protein, DH5alpha/pBV222-tMK, was obtained. DH5alpha/pBV222-tMK was cultured and induced with 42 degrees C. RESULTS: Truncated Midkine was cloned from gastric carcinogenesis tissue and the efficiently expressed recombinant tMK protein was obtained. SDS-PAGE indicated the molecular weight of recombinant tMK protein accorded with anticipation. CONCLUSION: The tMK was expressed in gastric carcinoma of Chinese patients' carcinogenesis tissue. The efficient expression of tMK protein was actualized in E. coli by clone and recombination.

Purification of the recombinant hepatitis B virus core antigen (rHBcAg) produced in the yeast Saccharomyces cerevisiae and comparative observation of its particles by transmission electron microscopy (TEM) and atomic force microscopy (AFM).

Hepatitis B virus core antigen (HBcAg) gene (C gene) was expressed in Saccharomyces cerevisiae and the products (rHBcAg or core particles) were purified from a crude lysate of the yeast by three steps: Sephrose CL-4B chromatography, Sucrose step-gradient ultracentrifugation and CsCl-isopycnic ultracentrifugation. It has been observed that HBcAg was synthesized in yeast cells as a particle consisting of polypeptides with a molecular weight of 21.5 kDa (p21.5). Results of ELISA test and density analysis of CsCl-isopycnic ultracentrifugation indicated that the purified products (rHBcAg particles) with HBcAg antigenicity mainly located at the densities of 1.27 and 1.40 g ml(-1), respectively. Observation and analysis of the purified rHBcAg products by TEM indicated that rHBcAg peptides could mainly self-assemble into two size classes of core particles. The larger particles were approximately 30.1 nm and the smaller were approximately 21.5 nm in mean diameter. Further observation and analysis of the same rHBcAg (core) particles by AFM also indicated that rHBcAg (core) particles were similar to the native HBcAg (core) particles from infected human hepatocytes and mainly composed of two size classes of partides core. The larger particles were approximately 31.3 nm and the smaller were approximately 22.5 nm in mean diameter which was similar to the results obtained by TEM. All results from both TEM and AFM suggested that core particles (capsids) produced in S. cerevisiae possessed dimorphism.