Fluid phase endocytosis of [125I]iodixanol in rat liver parenchymal, endothelial and Kupffer cells

Endocytosis of [125I]iodixanol was studied in vivo and in vitro in rat liver cells to determine fluid phase endocytic activity in different liver cells (hepatocytes, Kupffer cells and endothelial cells). The Kupffer cells were more active in the uptake of [l25I]iodixanol than parenchymal cells or endothelial cells. Inhibition of endocytic uptake via clathrin-coated pits (by potassium depletion and hypertonic medium) reduced uptake of [125I]iodixanol much more in Kupffer cells and endothelial cells than in hepatocytes. To gain further information about the importance of clathrin-mediated fluid phase endocytosis, the expression of proteins known to be components of the endocytic machinery was investigated. Using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting, endothelial cells and Kupffer cells were found to express approximately fourfold more rab4, rab5 and rab7 than parenchymal cells, while clathrin was expressed at a higher level in endothelial cells than in Kupffer cells and hepatocytes. Using electron microscopy it was shown that liver endothelial cells contained approximately twice as many coated pits per membrane unit than the parenchymal and Kupffer cells, thus confirming the immunoblotting results concerning clathrin expression. Electron microscopy on isolated liver cells following fluid phase uptake of horseradish peroxidase (HRP) showed that HRP-containing organelles had a different morphology in the different cell types: In the liver endothelial cells HRP was in small, tubular endosomes, while in Kupffer cells HRP was mainly found in larger structures, reminiscent of macropinosomes. Parenchymal cells contained HRP in small vacuolar endosomes with a punctuated distribution. In conclusion, we find that the Kupffer cells and the endothelial cells have a higher pinocytic activity than the hepatocytes. The hepatocytes do, however, account for most of the total hepatic uptake. The fluid phase endocytosis in liver endothelial cells depends mainly on clathrin-mediated endocytosis, while the parenchymal cells have additional clathrin-independent mechanisms that may play an important role in the uptake of plasma membrane components. In the Kupffer cells the major uptake of fluid phase markers seems to take place via a macropinocytic mechanism.     

Responses of two populations of an Iranian nickel-hyperaccumulating serpentine plant,Alyssum inflatum Nyar., to substrate Ca/Mg quotient and nickel

We selected two geographically close serpentine and non-serpentine populations of a Ni-hyperaccumulating plant (Alyssum inflatum) to investigate the influence of two common factors of serpentine soils: high Ni concentrations and low Ca/Mg quotients. Soils and plants were sampled from serpentine and non-serpentine substrates, and concentrations of Ca, Mg and Ni were measured. A hydroponic culture was used to compare growth and elemental composition responses of serpentine and non-serpentine plants to different Ca/Mg quotients and Ni concentrations in the nutrient solution. The Ca/Mg quotient for non-serpentine soils was 15 times higher than for serpentine soils, but there was no difference in the Ca/Mg quotient of plants from the two populations. In hydroponic culture, plants from both populations were able to survive at high Ca/Mg quotients. This result suggests that serpentine plants of A. inflatum do not necessarily need a substrate with a low Ca/Mg quotient for survival. Decreases in the Ca/Mg quotient in hydroponics decreased growth. The magnitude of this decrease was significantly greater in non-serpentine plants, suggesting a greater resistance of serpentine plants to low Ca/Mg quotients. Total Ni concentration in serpentine soils was 13 times higher than in non-serpentine soils, but ammonium nitrate-extractable concentrations of Ni in both soil types were similar. Ni concentrations in non-serpentine plants from their natural habitat were significantly lower than in serpentine plants, but there was no significant difference in Ni accumulation by plants of the two populations in hydroponic culture. However, increased concentrations of Ni in the hydroponic medium caused similar decreases in growth of both populations, indicating that Ni tolerance of the two populations was similar.     

Calling behaviour of the potato aphid Macrosiphum euphorbiae oviparae under laboratory and field conditions

Abstract.  1. The calling behaviour of virgin oviparae of the potato aphid Macrosiphum euphorbiae (Thomas) (Homoptera: Aphididae) was studied at three different constant temperatures under laboratory conditions. The mean age of calling for the first time decreased with a decrease in temperature from 2.9 days at 20 °C to 2.1 days at 10 °C. At all temperature regimes, the mean onset time of calling advanced from about 5 to 3 h after the onset of the photophase, and the mean time spent calling increased by > 4 h over the 8 days.
2. Cohorts of oviparae were also observed at two different periods in late summer–early autumn in the field, to examine the effects of fluctuating abiotic conditions (temperatures, wind velocity, rain) and age on calling behaviour. As under constant laboratory conditions, the mean age of calling for the first time declined with declining temperature, from 3.7 days in early September to 1.6 days at the end of September. Age-related changes in the mean onset time of calling and the mean time spent calling were much less evident under field conditions, due to the inhibitory effects of low temperatures, high winds, and rain on female calling activity.
3. The results are discussed within the context of reproductive success and address a previously proposed hypothesis suggesting that species-specific calling windows may serve as a reproductive isolating mechanism for sympatric aphid species.     

Kinetics of Cr(VI) Removal by Iron Filings in Some Soils

In this research, kinetics of Cr(VI) reduction by iron filings was investigated through a batch study in seven different soils. Chromate reduction experiments were carried out for initial Cr(VI) concentrations ranging from 20 to 100 mgkg^?1 and iron filings dosage of 0 to 5% w/w. The experimental data were analyzed using various kinetic models including zero-order, pseudo first-order, power function, Elovich, and diffusion parabolic. Results showed that the Cr(VI) reduction efficiency in the presence of all studied soils increased with increasing iron filings dosage and decreased with increasing the initial Cr(VI) concentration. The reaction rates considerably depended on pH and were higher in acidic soils. The diffusion parabolic model was the best kinetic model as evidenced by the highest determination coefficient (r²) and the lowest standard error of the estimate (SE). The rate-limiting step(s) may be transport of chromate anions across a liquid film at the interface of soil-liquid, transport in liquid-filled macropores of iron filings aggregates, or diffusion in micropores and along the particle's surface.     

Development of Acid-Resistant Alginate/Trimethyl Chitosan Nanoparticles Containing Cationic β-Cyclodextrin Polymers for Insulin Oral Delivery

In this study, the use of trimethylchitosan (TMC), by higher solubility in comparison with chitosan, in alginate/chitosan nanoparticles containing cationic β-cyclodextrin polymers (CPβCDs) has been studied, with the aim of increasing insulin uptake by nanoparticles. Firstly, TMCs were synthesized by iodomethane, and CPβCDs were synthesized within a one-step polycondensation reaction using choline chloride (CC) and epichlorohydrine (EP). Insulin–CβCDPs complex was prepared by mixing 1:1 portion of insulin and CPβCDs solutions. Then, nanoparticles prepared in a three-step procedure based on the iono-tropic pregelation method. Nanoparticles screened using experimental design and Placket Burman methodology to obtain minimum size and polydispercity index (pdI) and the highest entrapment efficiency (EE). CPβCDs and TMC solution concentration and pH and alginate and calcium chloride solution concentrations are found as the significant parameters on size, PdI, and EE. The nanoparticles with proper physicochemical properties were obtained; the size, PdI, and EE% of optimized nanoparticles were reported as 150.82 ± 21 nm, 0.362 ± 0.036, and 93.2% ± 4.1, respectively. The cumulative insulin release in intestinal condition achieved was 50.2% during 6 h. By SEM imaging, separate, spherical, and nonaggregated nanoparticles were found. In the cytotoxicity studies on Caco-2 cell culture, no significant cytotoxicity was observed in 5 h of incubation, but after 24 h of incubation, viability was decreased to 50% in 0.5 mμ of TMC concentration. Permeability studies across Caco-2 cells had been carried out, and permeability achieved in 240 min was 8.41 ± 0.39%, which shows noticeable increase in comparison with chitosan nanoparticles. Thus, according to the results, the optimized nanoparticles can be used as a new insulin oral delivery system.KEY WORDS: alginate, cationic β-cyclodextrin, insulin nanoparticle, oral delivery, trimethyl chitosan     

Improving Multi-Epitope Long Peptide Vaccine Potency by Using a Strategy that Enhances CD4+ T Help in BALB/c Mice

Peptide-based vaccines are attractive approaches for cancer immunotherapy; but the success of these vaccines in clinical trials have been limited. Our goal is to improve immune responses and anti-tumor effects against a synthetic, multi-epitope, long peptide from rat Her2/neu (rHer2/neu) using the help of CD4+ T cells and appropriate adjuvant in a mouse tumor model. Female BALB/c mice were vaccinated with P₅₊₄₃₅ multi-epitope long peptide that presents epitopes for cytotoxic T lymphocytes (CTL) in combination with a universal Pan DR epitope (PADRE) or CpG-oligodeoxynucleotides (CpG-ODNs) as a Toll-like receptor agonist adjuvant. The results show that vaccination with the multi-epitope long peptide in combination with the PADRE peptide and CpG-ODN induced expansion of subpopulations of CD4+ and CD8+ cells producing IFN-γ, the average tumor size in the vaccinated mice was less than that of the other groups, and tumor growth was inhibited in 40% of the mice in the vaccinated group. The mean survival time was 82.6 ± 1.25 days in mice vaccinated with P₅₊₄₃₅ + CpG+ PADRE. Our results demonstrate that inclusion of PADRE and CpG with the peptide vaccine enhanced significant tumor specific-immune responses in vaccinated mice.     

Characterization of Lung Fibroblasts More than Two Decades after Mustard Gas Exposure

Purpose

In patients with short-term exposure to the sulfur mustard gas, the delayed cellular effects on lungs have not been well understood yet. The lung pathology shows a dominant feature consistent with obliterative bronchiolitis, in which fibroblasts play a central role. This study aims to characterize alterations to lung fibroblasts, at the cellular level, in patients with delayed respiratory complications after short-term exposure to the sulfur mustard gas.

Methods

Fibroblasts were isolated from the transbronchial biopsies of patients with documented history of exposure to single high-dose sulfur mustard during 1985–7 and compared with the fibroblasts of control subjects.

Results

Compared with controls, patients’ fibroblasts were thinner and shorter, and showed a higher population doubling level, migration capacity and number of filopodia. Sulfur mustard decreased the in vitro viability of fibroblasts and increased their sensitivity to induction of apoptosis, but did not change the rate of spontaneous apoptosis. In addition, higher expression of alpha smooth muscle actin showed that the lung''s microenvironment in these patients is permissive for myofibroblastic differentiation.

Conclusions

These findings suggest that in patients under the study, the delayed pulmonary complications of sulfur mustard should be considered as a unique pathology, which might need a specific management by manipulation of cellular components.     

Trend of Cardio-Metabolic Risk Factors in Polycystic Ovary Syndrome: A Population-Based Prospective Cohort Study

Objective

To see the changes of cardio-metabolic risk factors overtime in polycystic ovary syndrome vs. control women.

Methods

This study was conducted on 637 participants (85 PCOS and 552 control reproductive aged, 18–45 years) of Tehran Lipid and Glucose Study (TLGS), an ongoing population-based cohort study with 12 years of follow-up. The cardiovascular risk factors of these groups were assessed in three-year intervals using standard questionnaires, history taking, anthropometric measures, and metabolic/endocrine evaluation. Generalized estimating equation was used to analyze the data.

Results

Overall mean of insulin (3.55, CI: 0.66–6.45), HOMA-IR (0.63, CI: 0.08–1.18), and HOMA-β (45.90, CI: 0.86–90.93) were significantly higher in PCOS than in healthy women after adjustment for age, BMI, and baseline levels. However, the negative interaction (follow-up years × PCOS status) of PCOS and normal women converged overtime. Comparing third follow-up with first, insulin and HOMA-IR decreased 10.6% and 5%, respectively in PCOS women; and increased 6.7% and 14.6%, respectively in controls (P<0.05). The results did not show any significant result for other cardio-metabolic variables including WC, lipid profile, FPG, 2-h PG, SBP, and DBP.

Conclusion

While the insulin level and insulin resistance rate were higher in reproductive aged PCOS than in healthy women, the difference of these risk factors decreased overtime. Thus, the metabolic consequences of PCOS women in later life may be lower than those initially anticipated.     

Four Categories of Viral Infection Describe the Health Status of Honey Bee Colonies

Honey bee virus prevalence data are an essential prerequisite for managing epidemic events in a population. A survey study was carried out for seven viruses in colonies representing a healthy Danish honey bee population. In addition, colonies from apiaries with high level Varroa infestation or high level of winter mortality were also surveyed. Results from RT-qPCR showed a considerable difference of virus levels between healthy and sick colonies. In the group of healthy colonies, no virus was detected in 36% of cases, while at least one virus was found in each of the sick colonies. Virus titers varied among the samples, and multiple virus infections were common in both groups with a high prevalence of Sacbrood virus (SBV), Black queen cell virus (BQCV) and Deformed wing virus (DWV). Based on the distribution of virus titers, we established four categories of infection: samples free of virus (C = 0), samples with low virus titer (estimated number of virus copies 0 < C < 10³), samples with medium virus titer (10³ ≤ C < 10⁷) and samples with high virus titer (C ≥ 10⁷). This allowed us to statistically compare virus levels in healthy and sick colonies. Using categories to communicate virus diagnosis results to beekeepers may help them to reach an informed decision on management strategies to prevent further spread of viruses among colonies.