Identification of phospholipase D from cabbage as N-terminally acetylated PLD2

Recently, the genes of two isoenzymes of phospholipase D from white cabbage (PLD1 and PLD2) with molecular masses of 91.7 and 91.9 kDa, respectively, have been sequenced and expressed in Escherichia coli [Schäffner, I., Rücknagel, K.-P., Mansfeld, J., and Ulbrich-Hofmann, R. (2002). Eur. J. Lipid Sci. Technol. 104: 79–87]. Both enzymes are highly homologous (91% identity) and behave very similarly. Phospholipase D purified from white cabbage leaves (PLD_cab) is compared with the two recombinant enzymes in sodium dodecylsulfate and native polyacrylamide gel electrophoresis, isoelectric focusing, N-terminal sequencing, and mass spectrometry after tryptic digestion. As a result, PLD_cab clearly can be assigned to PLD2. In contrast to recombinant PLD2, however, PLD_cab is N-terminally acetylated.     

Hydrostatic pressurization and depletion of trapped lubricant pool during creep contact of a rippled indenter against a biphasic articular cartilage layer

This study presents an analysis of the contact of a rippled rigid impermeable indenter against a cartilage layer, which represents a first simulation of the contact of rough cartilage surfaces with lubricant entrapment. Cartilage was modeled with the biphasic theory for hydrated soft tissues, to account for fluid flow into or out of the lubricant pool. The findings of this study demonstrate that under contact creep, the trapped lubricant pool gets depleted within a time period on the order of seconds or minutes as a result of lubricant flow into the articular cartilage. Prior to depletion, hydrostatic fluid load support across the contact interface may be enhanced by the presence of the trapped lubricant pool, depending on the initial geometry of the lubricant pool. According to friction models based on the biphasic nature of the tissue, this enhancement in fluid load support produces a smaller minimum friction coefficient than would otherwise be predicted without a lubricant pool. The results of this study support the hypothesis that trapped lubricant decreases the initial friction coefficient following load application, independently of squeeze-film lubrication effects.     

Toward the identification of liver toxicity markers: a proteome study in human cell culture and rats

The effects of toxic and nontoxic compound treatments were investigated by high resolution custom developed 2-11 pH gradient NEPHGE (non equilibrium pH gradient electrophoresis) two-dimensional electrophoresis. Two models were compared: (i) in vivo rat and (ii) the human cell line HepG2, to test their suitability in a proteomics based approach to identify a toxicity marker. 163 and 321 proteins were identified from the rat liver and the HepG2 proteome. These represent various isoforms of 113 and 194 different NCBI annotated gene sequences, respectively. Nine compounds were selected to induce proteome variations associated with liver toxicity and metabolism. The rat liver proteome database consists of 78 gels, the HepG2 database of 52 gels. Variant proteins were assessed regarding their usefulness as a toxicity marker by evaluating their treatment specificity against multiple control treatments. Thirteen potential toxicity marker proteins were found in rat liver and eight in HepG2. Catalase and carbamoylphosphate synthetase-1 isoforms were found to be significantly changed after treatment by 4/4 and 3/4 toxic compounds in rat liver, respectively. Aldo-keto-reductase family 1, member C1 was implicated for 3/4 liver cell toxic compounds in HepG2. Our approach was able to differentiate the quality of potential toxicity markers and provided useful information for an ongoing characterization of more compounds in a wider number of toxicity classes.     

Peroxynitrite flux-mediated LDL oxidation is inhibited by manganese porphyrins in the presence of uric acid

We have studied the role of three Mn(III)porphyrins differing in charge, alkyl substituent length and reactivity, on LDL exposed to low fluxes of peroxynitrite (PN) in the presence of uric acid. Mn(III)porphyrins (5 microM, MnTE-2-PyP(5+), MnTnOct-2-PyP(5+), and MnTCPP(3-)) plus uric acid (300 microM) inhibited cholesteryl ester hydroperoxide formation, changes in REM as well as spared alpha- and gamma-tocopherol. MnTnOct-2-PyP(5+), the more lipophilic compound, was the most effective in protecting LDL lipids, while MnTCPP(3-) exerted the lesser protection. Mn(III)porphyrins react fast with PN ( approximately 10(5)-10(7) M(-1) s(-1)) to yield a O=Mn(IV) complex. The stoichiometry of uric acid consumption was approximately 1.7 moles per mol of PN, in agreement with reactions with both the O=Mn(IV) complex and nitrogen dioxide. A shift from an anti- to a pro-oxidant action of the Mn(III)porphyrin was observed after uric acid was significantly consumed, supporting competition reactions between LDL targets and uric acid for the O=Mn(IV) complex. Overall, the data is consistent with the catalytic reduction of PN in a cycle that involves a one electron oxidation of Mn(III) to Mn(IV) by PN followed by the reduction back to Mn(III) by uric acid. These antioxidant effects should predominate under in vivo conditions having plasma uric acid concentration range between 150 and 500 microM.     

Juvenile hormone effect on DNA synthesis and apoptosis in caste-specific differentiation of the larval honey bee (Apis mellifera L.) ovary

Caste-specific differentiation of the honey bee ovary commences in the last larval instar. In this process, formation of germ cell clusters by synchronous and incomplete mitoses occurs in the queen ovary, whereas in the worker ovary programmed cell death is the dominant feature. BrdU and TUNEL labeling were used to study dynamics of cell proliferation and apoptosis-dependent DNA degradation in ovaries of naturally developing queens and workers, as well as in juvenile hormone-treated worker larvae. Cell proliferation in ovaries of last-instar queen larvae generally exceeded that in workers, except for the late feeding phase. This inversion in cell proliferation patterns coincided with the onset of apoptosis in worker ovaries, as evidenced by TUNEL labeling. Juvenile hormone application to early-fifth-instar worker larvae had two noticeable effects. First, it diminished the number of S-phase nuclei in ovaries of late feeding-phase workers, bringing them to queen-like levels. Second, it prevented the induction of apoptotic DNA degradation. Caste-specific regulation of cell division in connection with programmed cell death can thus be attributed to the previously described differences in juvenile hormone titer in queen and worker larvae, adding a new facet to this hormone's multiple functions.     

Plant homologue of human excision repair gene ERCC1 points to conservation of DNA repair mechanisms

Nucleotide excision repair (NER), a highly versatile DNA repair mechanism, is capable of removing various types of DNA damage including those induced by UV radiation and chemical mutagens. NER has been well characterized in yeast and mammalian systems but its presence in plants has not been reported. Here it is reported that a plant gene isolated from male germline cells of lily (Lilium longiflorum) shows a striking amino acid sequence similarity to the DNA excision repair proteins human ERCC1 and yeast RAD10. Homologous genes are also shown to be present in a number of taxonomically diverse plant genera tested, suggesting that this gene may have a conserved function in plants. The protein encoded by this gene is able to correct significantly the sensitivity to the cross-linking agent mitomycin C in ERCC1-deficient Chinese hamster ovary (CHO) cells. These findings suggest that the NER mechanism is conserved in yeast, animals and higher plants.     

14C-菲在营养液-火山石-小麦有控系统中的迁移和转化

利用放射性同位素示踪技术研究了¹⁴C标记菲在有控系统中的迁移转化。结果表明,¹⁴C-菲在有控系统中降解较快,施入药品24d后仅有0.32%的¹⁴C-菲存留在植物、营养液和火山石中。植物吸收的¹⁴C放射性大部分被结合到植物组织中,其它¹⁴C主要以菲的极性代谢物形态存在。     

Chemoradiotherapy or chemotherapy as adjuvant treatment for resected gastric cancer: should we use selection criteria?

BackgroundThe management of gastric adenocarcinoma is essentially based on surgery followed by adjuvant treatment. Adjuvant chemotherapy (CT) as well as chemoradiotherapy (CTRT) have proven their effectiveness in survival outcomes compared to surgery alone. However, there is little data comparing the two adjuvant approaches. This study aimed to compare the prognosis and survival outcomes of patients with gastric adenocarcinoma operated and treated by adjuvant radio-chemotherapy or chemotherapyMaterials and methodsWe retrospectively evaluated 80 patients with locally advanced gastric cancer (LGC) who received adjuvant treatment. We compared survival outcomes and patterns of recurrence of 53 patients treated by CTRT and those of 27 patients treated by CT.ResultsAfter a median follow-up of 38.48 months, CTRT resulted in a significant improvement of the 5-year PFS (60.9% vs. 36%, p = 0.03) and the 5-year OS (55.9% vs. 33%, p = 0.015) compared to adjuvant CT. The 5-year OS was significantly increased by adjuvant CTRT (p = 0.046) in patients with lymph node metastasis, and particularly those with advanced pN stage (p = 0.0078) and high lymph node ratio (LNR) exceeding 25% (p = 0.012). Also, there was a significant improvement of the PFS of patients classified pN2–N3 (p = 0.022) with a high LNR (p = 0.018). CTRT was also associated with improved OS and PFS in patients with lymphovascular and perineural invasion (LVI and PNI) compared to chemotherapy.ConclusionThere is a particular survival benefit of adding radiotherapy to chemotherapy in patients with selected criteria such as lymph node involvement, high LNR LVI, and PNI.     

Differential protein expression profile in the liver of pikeperch (Sander lucioperca) larvae fed with increasing levels of phospholipids

A comparative proteomic approach was used to assess the protein expression profile in the liver of 34 days old pikeperch larvae fed from day 10 post hatching, with three isoproteic and isolipidic formulated diets varying by their phospholipid (PL) contents (% dry diet weight): 1.4% (PL1), 4.7% (PL5) and 9.5% (PL9). Using 2D-DIGE minimal labelling of liver extracts, we were able to show 56 protein spots with a differential intensity (p < 0.05) depending on the dietary PL content. Among these spots, 11 proteins were unambiguously identified using nanoLC-MS/MS tandem mass spectrometry. In the PL9 larvae, our results indicate that the glycolytic pathway could be down-regulated due to the under-expression of the fructose biphosphate aldolase B and the phosphoglucomutase 1. Meanwhile, propionyl coenzyme A carboxylase (a gluconeogenic enzyme) was under-expressed. In addition, another gluconeogenic and lipogenic enzyme, pyruvate carboxylase, was identified in 3 different spots as being under-expressed in fish fed with the intermediate PL level (PL5). A high PL content increased the expression of sarcosine dehydrogenase, an enzyme involved in methionine metabolism, along with vinculin, a structural protein. Moreover, several stress proteins (glutathione S-transferase M, glucose regulated protein 75 and peroxiredoxin-1) were modulated in response to the dietary PL level and fatty acid composition. In the larvae fed with the lowest dietary PL content (PL1), over-expression of both GSTM and GRP75 might indicate a cellular stress in this experimental treatment, while the under-expression of Prx1 might indicate a lower defence against oxidative stress. In conclusion, this nutriproteomic approach showed significant modifications of protein expression in the liver of pikeperch larvae fed different PL contents, highlighting the importance of these nutrients and their influence on metabolism processes and on stress response.