Cell kinetic studies of chick brain cells in culture: an autoradiographic study with [3H]- and [14C]-thymidine

Labelling index, S-phase duration and cell-cycle time of proliferating brain cells from 6-day-old chick embryos in culture were investigated autoradiographically after labelling with [3H]- and/or [14C]-thymidine. The dissociated cells were cultured in the absence or in the presence of brain extract from 8-day-old chick embryos. Cultures contained essentially two cell types, which could be easily distinguished by the size of their nuclei: small nuclei identified as belonging to precursor cells of neurons and large nuclei corresponding to astroglial cells. The labelling index of astroglial cells (16.4%) was about 2 times higher than that of the neuronal cells (9.9%). Under the influence of brain extract the labelling index of neuroblasts was nearly doubled while that of the astroglial cells remained nearly unchanged. From double-labelling experiments with [3H]- and [14C]-thymidine, the same S-phase duration of about 7 hr was found for both cell types cultured with or without brain extract. A cell-cycle duration of 39 hr for neuronal and of 29 hr for astroglial cells was found. The cycle times remained constant under the influence of brain extract. From the measured data mentioned above, a growth fraction of 50% (neuroblasts) and 68% (astroglial cells) was calculated in control cultures without brain extract. After addition of brain extract, the growth fraction increased for both cell types (neuroblasts: 92%; astroglial cells: 80%). The results demonstrate that more cells proliferate in the presence of brain extract, but the durations of the S-phase and the cell cycle remain unchanged.     

Numerical study of the impact of non-Newtonian blood behavior on flow over a two-dimensional backward facing step

Endothelial cell (EC) responsiveness to shear stress is essential for vasoregulation and plays a role in atherogenesis. Although blood is a non-Newtonian fluid, EC flow studies in vitro are typically performed using Newtonian fluids. The goal of the present study was to determine the impact of non-Newtonian behavior on the flow field within a model flow chamber capable of producing flow disturbance and whose dimensions permit Reynolds and Womersley numbers comparable to those present in vivo. We performed two-dimensional computational fluid dynamic simulations of steady and pulsatile laminar flow of Newtonian and non-Newtonian fluids over a backward facing step. In the non-Newtonian simulations, the fluid was modeled as a shear-thinning Carreau fluid. Steady flow results demonstrate that for Re in the range 50-400, the flow recirculation zone downstream of the step is 22-63% larger for the Newtonian fluid than for the non-Newtonian fluid, while spatial gradients of shear stress are larger for the non-Newtonian fluid. In pulsatile flow, the temporal gradients of shear stress within the flow recirculation zone are significantly larger for the Newtonian fluid than for the non-Newtonian fluid. These findings raise the possibility that in regions of flow disturbance, EC mechanotransduction pathways stimulated by Newtonian and non-Newtonian fluids may be different.     

Calcitonin gene-related peptide and islet amyloid polypeptide stimulate insulin secretion in RINm5F cells through a common receptor coupled to a generation of cAMP

The question as to whether the homologous peptides CGRP and IAPP can regulate insulin secretion in RINm5F cells was addressed. Chicken CGRP displayed a reproducible inhibitory effect on insulin secretion within 0.1 and 1 nM concentrations and a stimulatory effect at higher concentrations. The maximal stimulatory effects on insulin secretion were obtained with 1.0 M of chicken CGRP (cCGRP), human -CGRP (h -CGRP) and human IAPP (hIAPP) which caused 246 ± 22, 302 ± 63 and 224 ± 14 percent increases of control levels, respectively (p < 0.001). Similarly, maximal accumulations of cAMP were obtained with 1.0 M of cCGRP, h -CGRP and hIAPP with the respective percent increases of control levels of 587 ± 24, 436 ± 41 and 410 ± 25 (p < 0.005). Thus the stimulatory effects on insulin secretion in RINm5F cells by cCGRP, h -CGRP and hIAPP appear to be mediated by the cAMP pathway. Chicken CGRP, the most potent peptide tested, displayed a correlated dose response stimulation of intracellular cAMP and insulin release within the concentration range of 10–1000nM. The EC₅₀ values of cCGRP for cAMP accumulation and insulin release were similar (20nM and 10 nM respectively). The stimulatory effect of IAPP on cAMP was not additive with that of cCGRP suggesting that IAPP action was mediated by CGRP receptors. This hypothesis was further sustained by a preferential inhibition of¹²⁵I[His]h -CGRP binding to RINm5F cells by cCGRP as compared to IAPP.We conclude that CGRP and IAPP, through a direct action on a chicken CGRP preferring receptor present in cells, stimulated insulin by a cAMP mediated pathway.     

Revealing the atomistic details behind the binding of B7–1 to CD28 and CTLA-4: A comprehensive protein-protein modelling study

Background

CD28 and CTLA-4 are homologous T-cell receptors that bind with B7–1 and produce two opposing immunological signals required for T-cell activation and inactivation, respectively. It has been clinically proven that specific blockade of these key protein-protein interactions at the synapse can offer immunotherapeutic benefits for cancers and autoimmune treatments. Hence, there is a growing interest towards developing anti-CD28 and anti-CTLA-4 small molecule inhibitors. To achieve this goal, it is important to understand unique molecular level fingerprint interactions that stabilize CTLA-4/B7–1 and CD28/B7–1 complexes. However, until recently, the structure of the human CD28/B7–1 complex has not been resolved experimentally, which remains a significant setback in achieving specific inhibitors against CTLA-4 or CD28.

Influence of age and exercise training on lipid metabolism in Fischer-344 rats

The influence of training on fatty acid and glyceride synthesis by liver and adipose tissue homogenates of young and old Fischer-344 rats was examined. Four groups of rats (10 animals/group) were studied: young untrained, young trained, old untrained, and old trained. Training of each group was for 10 wk at 75% maximal O2 uptake. Young rats were killed at 6 mo of age and old rats were killed at 27 mo of age. Fatty acid synthesis was assessed by measuring the activities of acetyl-CoA carboxylase, fatty acid synthase, ATP citrate-lyase, "malic" enzyme, and glucose-6-phosphate dehydrogenase. Glyceride synthesis was evaluated by determining the rate of incorporation of [14C]glycerol 3-phosphate into lipids. In addition, lipoprotein lipase activity was measured in acetone-ether powders of adipose tissue from the four groups of rats. In liver, training had no effect on fatty acid or glyceride synthesis in either group. However, aging caused a significant decrease in the activities of four of the lipogenic enzymes but had no effect on glyceride synthesis. Training caused an increase in fatty acid synthase and glyceride synthesis in adipose tissue, and aging decreased lipoprotein lipase activity. It was concluded that training enhances the synthetic capacity of lipids by adipose tissue but that aging had a more profound effect in that the activities of the enzymes involved in these processes were lower in the old rats. Furthermore, the decreased activity of lipoprotein lipase in the older rats may explain the higher plasma triglyceride levels that were observed in these animals.     

Flavonoid constituents of Ephedra alata

Two new flavonol glucosides have been identified in Ephedra alata, namely, herbacetin 8-methyl ether 3-O- glucoside-7-O-rutinoside and herbacetin 7-O-(6″-quinylglucoside). The known flavonoids vicenin II, lucenin III, kaempferol 3-rhamnoside, quercetin 3-rhamnoside and herbacetin 7-glucoside were also found. The structure of the isolated compounds was determined mostly by FABMS and ¹H NMR spectroscopy. The final structure of the new compounds and of herbacetin 7-glucoside was confirmed by ¹³C NMR spectroscopy.     

Effect of dietary chitosan on challenged <Emphasis Type="Italic">Dicentrarchus labrax</Emphasis> post larvae with <Emphasis Type="Italic">Aeromonas hydrophila</Emphasis>

Effect of different dietary squilla chitosan (Csq) concentrations: 0 (control), 0.5, 1 and 2 g 100 g^–1 diets were studied for weaned sea bass (Dicentrarchus labrax) post larvae. Post larvae were challenged with Aeromonas hydrophila after 5 feeding days, in order to monitor the prophylactic effect on the Csq fed larvae. The experiment started with an average initial weight of 50 ± 2 mg and total length of 12 ± 2 mm for post larval stage (40 days post hatch; dph), then continued feeding diets for a period of 20 days. Larvae survival percentage (%), mean total length (TL), width (W), total weight (TW), total weight gain (TWG), average daily weight (ADW) and specific growth rate (SGR) were recorded as morphometric measurements representing growth compared to the control groups. The results revealed that 1g Csq 100 g^–1 diet at P < 0.05 was the most effective concentration that achieved higher survival percentages; 94.5 ± 0.5 and 74 ± 2.0%, increasing the specific growth rate by 7.22% and 5.77% for non challenged and challenged weaned larval groups, respectively. Otherwise, the control challenged group displayed the lowest performance in all assayed parameters with the coincidental decrease in the survival % and specific growth rates. Similarly, lower growth performance was also observed at 2 g 100 g^–1 diet. Thus, the incorporation of chitosan at a level of 1g in fish diet enhanced the performance and reduced the fish mortality under stress conditions.