Influences of the priming procedure and saline circulation conditions on polyvinylpyrrolidone in vitro elution from polysulfone membrane dialyzers

In hemodialysis (HD), the patient's blood is purified via circulation in an extracorporeal circuit containing a dialyzer. In the manufacturing process of polysulfone (PSu) membrane dialyzers, the membranes are hydrophilized via the addition of the hydrophilic agent polyvinylpyrrolidone (PVP) to increase their hydraulic permeability. The elution of PVP from the membrane reduces the membrane's hydraulic permeability, and the eluted PVP could cause adverse effects in the human body. Therefore, it is important to identify the factors that induce PVP elution from PSu dialyzer membranes to improve the efficiency and safety of HD. In the present study, experimental circuits connecting each of the three types of PSu membrane dialyzers that had been sterilized, using gamma irradiation, autoclaving, or in-line steam methods, were prepared. After the dialyzers were primed, saline was circulated in the circuits at a flow rate of 100 mL/min or 200 mL/min. At 0, 2, 4, 6, and 8 h after circulation was initiated, the amount of PVP eluted from the PSu membranes in vitro was determined. In this experimental setting, longer the circulation duration, greater the amount of PVP eluted from the PSu membranes of the tested dialyzers; however, the flow rate did not influence the in vitro elution of PVP. Furthermore, the immersion of the dialyzer membranes in saline for 24 h strongly facilitated the in vitro elution of PVP. In sum, these results suggest that the duration of PSu membrane incubation in saline is a determinant of the level of PVP elution from the PSu membrane dialyzers.     

A Highlights from MBoC Selection: Serine 129 phosphorylation of membrane-associated α-synuclein modulates dopamine transporter function in a G protein–coupled receptor kinase–dependent manner

Most α-synuclein (α-syn) deposited in Lewy bodies, the pathological hallmark of Parkinson disease (PD), is phosphorylated at Ser-129. However, the physiological and pathological roles of this modification are unclear. Here we investigate the effects of Ser-129 phosphorylation on dopamine (DA) uptake in dopaminergic SH-SY5Y cells expressing α-syn. Subcellular fractionation of small interfering RNA (siRNA)–treated cells shows that G protein–coupled receptor kinase 3 (GRK3), GRK5, GRK6, and casein kinase 2 (CK2) contribute to Ser-129 phosphorylation of membrane-associated α-syn, whereas cytosolic α-syn is phosphorylated exclusively by CK2. Expression of wild-type α-syn increases DA uptake, and this effect is diminished by introducing the S129A mutation into α-syn. However, wild-type and S129A α-syn equally increase the cell surface expression of dopamine transporter (DAT) in SH-SY5Y cells and nonneuronal HEK293 cells. In addition, siRNA-mediated knockdown of GRK5 or GRK6 significantly attenuates DA uptake without altering DAT cell surface expression, whereas knockdown of CK2 has no effect on uptake. Taken together, our results demonstrate that membrane-associated α-syn enhances DA uptake capacity of DAT by GRKs-mediated Ser-129 phosphorylation, suggesting that α-syn modulates intracellular DA levels with no functional redundancy in Ser-129 phosphorylation between GRKs and CK2.     

Plant communities and environmental factors in the Guayana Highlands: monitoring for conservation under future climate change

The Guayana Highlands (GH) constitute a highly diverse, but relatively poorly studied Neotropical biome, comprised of ～50 flat-topped mountain summits (called tepuis). Previous studies based on warming forecasts for the region suggested that an upward displacement of environmental conditions of 500–700 m could occur by 2100, potentially resulting in the extinction of c. 50% of its endemic flora due to total habitat loss. To assess the ecological responses of the species to climate change, and select the appropriate conservation measures, long-term monitoring of the GH plant communities will be necessary. In this study, the baseline state for future comparisons was established for the best explored tepui in terms of its flora, Roraima-tepui (2810 m), through a floristic characterization of its different vegetation types. We also identified the environmental gradients underlying the major plant communities, and assessed the effects of human activities on the chemistry of soils and water at three field camps. Our results yielded five main community types: three meadows, one shrubland, and one forest, with their corresponding diagnostic species. The herbaceous communities were mainly influenced by the presence of flat sandy soils, with varying flooding capacity. Shrublands and forests were characterized by irregular organic soils with very low pH. Finally, pH values below 3 were measured on an organic soil of a field camp, although further studies will be necessary to attribute this deviation to human activities.     

Identification of Biological Properties of Intralymphatic Tumor Related to the Development of Lymph Node Metastasis in Lung Adenocarcinoma

Background

Intralymphatic tumors in the extratumoral area are considered to represent the preceding phase of lymph node metastasis. The aim of this study was to clarify the biological properties of intralymphatic tumors susceptible to the development of lymph node metastasis, with special reference to the expression of cancer initiating/stem cell (CIC/CSC) related markers in cancer cells and the number of infiltrating stromal cells.

Material and Methods

Primary lung adenocarcinomas with lymphatic permeation in the extratumoral area were retrospectively examined (n = 107). We examined the expression levels of CIC/CSC related markers including ALDH1, OCT4, NANOG, SOX2 and Caveolin-1 in the intralymphatic cancer cells to evaluate their relationship to lymph node metastasis. Moreover, the number of infiltrating stromal cells expressing CD34, α-smooth muscle actin, and CD204 were also evaluated.

Results

Among the intralymphatic tissues, low ALDH1 expression in cancer cells, high SOX2 expression in cancer cells, and a high number of CD204(+) macrophages were independent predictive factors for lymph node metastasis (P = 0.004, P = 0.008, and P = 0.028, respectively). Among these factors, only low ALDH1 expression in cancer cells was significantly correlated with the farther spreading of lymph node metastasis (mediastinal lymph node, pathological N2) (P = 0.046) and the metastatic lymph node ratio (metastatic/resected) (P = 0.028). On the other hand, in the primary tumors, ALDH1 expression in the cancer cells was not associated with lymph node metastasis. Intralymphatic cancer cells expressing low ALDH1 levels exhibited lower E-cadherin expression levels than cancer cells with high levels of ALDH1 expression (P = 0.015).

Conclusions

Intralymphatic cancer cells expressing low levels of ALDH1 and infiltrating macrophages expressing CD204 have a critical impact on lymph node metastasis. Our study also highlighted the significance of evaluating the biological properties of intralymphatic tumors for tumor metastasis.     

Translesion DNA Synthesis and Mutagenesis in Prokaryotes

The presence of unrepaired lesions in DNA represents a challenge for replication. Most, but not all, DNA lesions block the replicative DNA polymerases. The conceptually simplest procedure to bypass lesions during DNA replication is translesion synthesis (TLS), whereby the replicative polymerase is transiently replaced by a specialized DNA polymerase that synthesizes a short patch of DNA across the site of damage. This process is inherently error prone and is the main source of point mutations. The diversity of existing DNA lesions and the biochemical properties of Escherichia coli DNA polymerases will be presented. Our main goal is to deliver an integrated view of TLS pathways involving the multiple switches between replicative and specialized DNA polymerases and their interaction with key accessory factors. Finally, a brief glance at how other bacteria deal with TLS and mutagenesis is presented.Within the context of this review, we will limit the notion of DNA lesions to chemically altered bases, although the sugar-phosphodiester backbone is also subject to various types of chemical attack leading, for example, to single-strand breaks. Lesions may be spontaneous (e.g., depurinations), induced endogenously (e.g., by reactive oxygen species), induced by radiations (UV light, X rays) or by chemicals. Treatments that induce DNA lesions cause mutations and cancer and are therefore referred to as mutagens or carcinogens. Carcinogens fall into large chemical families of compounds such as aromatic amides, polycyclic hydrocarbons, and nitrosamines. Carcinogens are not necessarily synthetic; for example, some are natural plant metabolites (e.g., Aflatoxin B1, aristolochic acid, etc.). In addition, some drugs used in cancer chemotherapy such as platinum derivatives form covalent DNA adducts and as such are also carcinogens. Drugs from the thiopurine family, such as azathioprine widely used as immunosuppressants in organ transplant patients, form DNA adducts upon interaction with sunlight and promote skin cancer (Zhang et al. 2007).     

Diet of Japanese eels Anguilla japonica in the Kojima Bay-Asahi River system, Japan

The diet of Japanese eels, Anguilla japonica, was investigated using stomach content and stable isotope analyses. Stable isotope enrichment of carbon and nitrogen (Δδ¹³C and Δδ¹⁵N) was first estimated for A. japonica by comparing the isotopic signatures (δ¹³C and δ¹⁵N) of reared eels to that of their food. The estimated isotope enrichment was then applied to the diet estimation of A. japonica in the Kojima Bay-Asahi River system, Japan, combined with conventional stomach content analysis. Stable isotope enrichment varied among tissues, from 0.2‰ to 0.8‰ for carbon and from 1.3‰ to 2.1‰ for nitrogen. Nitrogen isotope enrichment of A. japonica muscle estimated in this study was 2.1‰, which was different from the previously reported mean δ¹⁵N enrichment of several animals of 3.4‰. These results indicate that isotope-based diet estimations for A. japonica need to use species- and tissue-specific values of isotope enrichment. In the diet analysis, stomach contents and stable isotopes revealed that (1) A. japonica appear to usually feed on a single type of prey species in each feeding session, (2) principal prey species were mud shrimp, Upogebia major, in brackish Kojima Bay and crayfish, Procambarus clarkia, in the Asahi River, (3) A. japonica in Kojima Bay primarily depend on the pelagic food web as a carbon source due to mud shrimp being filter feeders and eels in the Asahi River primarily depend on the littoral food web. Based on these results and the recently reported eel movements between Kojima Bay and the Asahi River, it appears that A. japonica can adapt to various feeding environments as opportunists, but also utilize the food resources by targeting a single type of prey species during a single feeding session.     

Ab initio fragment molecular orbital calculations on the specific interactions between human,mouse and rat PPARα and GW409544

To elucidate the specific interactions between the peroxisome proliferator-activated receptor (PPARα) and ligand GW409544 (GW), we obtained the solvated structures of the PPARα+GW complexes for human, mouse and rat by classical molecular mechanics calculations, and investigated their electronic properties by ab initio fragment molecular orbital calculations. The results indicate that the positively charged amino acids (Lys and Arg) of PPARα make a major contribution to the binding between PPARα and GW. In addition, it was clarified that Ser280 and Tyr314 of human and rat PPARα have a large attractive interaction with GW, while Ser280, Tyr314 and His440 of mouse PPARα have large interaction. These results on the difference in specific interactions between human and mouse/rat PPARα will be useful for predicting the effects of new chemicals on the human body based on the biomedical studies for the experimental animals such as mouse and rat.     

Bioproduction of prostaglandins in a transgenic liverwort, Marchantia polymorpha

Prostaglandins are biologically active substances used in a wide range of medical treatments. Prostaglandins have been supplied mainly by chemical synthesis; nevertheless, the high cost of prostaglandin production remains a factor. To lower the cost of prostaglandin production, we attempted to produce prostaglandins using a liverwort, Marchantia polymorpha L., which accumulates arachidonic acid, which is known as a substrate of prostaglandins. Here we report the first bioproduction of prostaglandins in plant species by introducing a cyclooxygenase gene from a red alga, Gracilaria vermiculophylla into the liverwort. The transgenic liverworts accumulated prostaglandin F_2α, prostaglandin E₂ and prostaglandin D₂ which were not detected in the wild-type liverwort. Moreover, we succeeded in drastically increasing the bioproduction of prostaglandins using an in vitro reaction system with the extracts of transgenic liverworts.