Energetics of swelling in isolated hepatocytes: A comprehensive study

Cell swelling is now admitted as being a new principle of metabolic control but little is known about the energetics of cell swelling. We have studied the influence of hypo- or hyperosmolarity on both isolated hepatocytes and isolated rat liver mitochondria. Cytosolic hypoosmolarity on isolated hepatocytes induces an increase in matricial volume and does not affect the myxothiazol sensitive respiratory rate while the absolute value of the overall thermodynamic driving force over the electron transport chain increases. This points to an increase in kinetic control upstream the respiratory chain when cytosolic osmolarity is decreased. On isolated rat liver mitochondria incubated in hypoosmotic potassium chloride media, energetic parameters vary as in cells and oxidative phosphorylation efficiency is not affected. Cytosolic hyperosmolarity induced by sodium co-transported amino acids, per se, does not affect either matrix volume or energetic parameters. This is not the case in isolated rat liver mitochondria incubated in sucrose hyperosmotic medium. Indeed, in this medium, adenine nucleotide carrier is inhibited as the external osmolarity increases, which lowers the state 3 respiration close to state 4 level and consequently leads to a decrease in oxidative phosphorylation efficiency. When isolated rat liver mitochondria are incubated in KCl hyperosmotic medium, state 3 respiratory rate, matrix volume and membrane electrical potential vary as a function of time. Indeed, matrix volume is recovered in hyperosmotic KCl medium and this recovery is dependent on Pi-Kentry. State 3 respiratory rate increases and membrane electrical potential difference decreases during the first minutes of mitochondrial incubation until the attainment of the same value as in isoosmotic medium. This shows that matrix volume, flux and force are regulated as a function of time in KCl hyperosmotic medium. Under steady state, neither matrix volume nor energetic parameters are affected. Moreover, NaCl hyperosmotic medium allows matrix volume recovery but induces a decrease in state 3 respiratory flux. This indicates that potassium is necessary for both matrix volume and flux recovery in isolated mitochondria. We conclude that hypoosmotic medium induces an increase in kinetic control both upstream and on the respiratory chain and changes the oxidative phosphorylation response to forces. At steady state, hyperosmolarity, per se, has no effect on oxidative phosphorylation in either isolated hepatocytes or isolated mitochondria incubated in KCl medium. Therefore, potassium plays a key role in matrix volume, flux and force regulation.     

Direct anabolic metabolism of three-carbon propionate to a six-carbon metabolite occurs in vivo across tissues and species

Anabolic metabolism of carbon in mammals is mediated via the one- and two-carbon carriers S-adenosyl methionine and acetyl-coenzyme A. In contrast, anabolic metabolism of three-carbon units via propionate has not been shown to extensively occur. Mammals are primarily thought to oxidize the three-carbon short chain fatty acid propionate by shunting propionyl-CoA to succinyl-CoA for entry into the TCA cycle. Here, we found that this may not be absolute as, in mammals, one nonoxidative fate of propionyl-CoA is to condense to two three-carbon units into a six-carbon trans-2-methyl-2-pentenoyl-CoA (2M2PE-CoA). We confirmed this reaction pathway using purified protein extracts provided limited substrates and verified the product via LC-MS using a synthetic standard. In whole-body in vivo stable isotope tracing following infusion of ¹³C-labeled valine at steady state, 2M2PE-CoA was found to form via propionyl-CoA in multiple murine tissues, including heart, kidney, and to a lesser degree, in brown adipose tissue, liver, and tibialis anterior muscle. Using ex vivo isotope tracing, we found that 2M2PE-CoA also formed in human myocardial tissue incubated with propionate to a limited extent. While the complete enzymology of this pathway remains to be elucidated, these results confirm the in vivo existence of at least one anabolic three- to six-carbon reaction conserved in humans and mice that utilizes propionate.     

Membrane electrical parameters in turtle bladder measured using impedance-analysis techniques

Summary Equivalent-circuit impedance analysis experiments were performed on the urinary bladders of freshwater turtles in order to quantify membrane ionic conductances and areas, and to investigate how changes in these parameters are associated with changes in the rate of proton secretion in this tissue. In all experiments, sodium reabsorption was inhibited thereby unmasking the electrogenic proton secretion process. We report the following: (1) transepithelial impedance is represented exceptionally well by a simple equivalent-circuit model, which results in estimates of the apical and basolateral membrane ionic conductances and capacitances; (2) when sodium transport is inhibited with mucosal amiloride and serosal ouabain, the apical and basolateral membrane conductances and capacitances exhibit a continual decline with time; (3) this decline in the membrane parameters is most likely caused by subtle time-dependent changes in cell volume, resulting in changes in the areas of the apical and basolateral membranes; (4) stable membrane parameters are obtained if the tissue is not treated with ouabain, and if the oncotic pressure of the serosal solution is increased by the addition of 2% albumin; (5) inhibition of proton secretion using acetazolamide in CO₂ and HCO ₃ ^– -free bathing solutions results in a decrease in the area of the apical membrane, with no significant change in its specific conductance; (6) stimulation of proton transport with CO₂ and HCO ₃ ^– -containing serosal solution results in an increase in the apical membrane area and specific conductance. These results show that our methods can be used to measure changes in the membrane electrophysiological parameters that are related to changes in the rate of proton transport. Notably, they can be used to quantify in the live tissue, changes in membrane area resulting from changes in the net rates of endocytosis and exocytosis which are postulated to be intimately involved in the regulation of proton transport.     

Estimating the Volume Density from Thin Sections

An approximation formula for the determination of the volume density from thin sections is given. It is applicable under very mild conditions on the structure studied and has a good accuracy.     

Determination of optimal planning target volume margins in patients with gynecological cancer

PurposeTo define optimal planning target volume (PTV) margins for intensity modulated radiotherapy (IMRT) ± knee-heel support (KHS) in patients treated with adjuvant radiotherapy.MethodsComputed tomography (CT) scans ± KHS of 10 patients were taken before and at 3rd and 5th week of treatment, fused and compared with initial IMRT plans.ResultsA PTV margin of 15 mm in anteroposterior (AP) and superoinferior (SI) directions and 5 mm in lateral directions were found to be adequate without any difference between ± KHS except for the SI shifts in CTV-primary at the 3rd week. Five mm margin for iliac CTV was found to be inadequate in 10–20% of patients in SI directions however when 7 mm margin was given for iliac PTV, it was found to be adequate. For presacral CTV, it was found that the most striking shift of the target volume was in the direction of AP. KHS caused significantly less volume of rectum and bladder in the treated volume.ConclusionsPTV margin of 15 mm in SI and AP, and 5 mm in lateral directions for CTV-primary were found to be adequate. A minimum of 7 mm PTV margin should be given to iliac CTV. The remarkable shifting in presacral CTV was believed to be due to the unforeseen hip malposition of obese patients. The KHS seems not to provide additional beneficial effect in decreasing the shifts both in CTV-primary and lymphatic, however it may have a beneficial effect of decreasing the OAR volume in PTV margins.     

Malaria-induced reduction of fecundity during the first gonotrophic cycle of Anopheles Stephensi mosquitoes

Abstract. Anopheles stephensi mosquitoes which had fed upon mice infected with Plasmodium yoelii nigeriensis malaria parasites produced significantly fewer eggs than mosquitoes fed on an uninfected mouse. Fecundity reduction was more pronounced when the bloodmeal contained malaria gametocytes and the mosquitoes developed oocysts. Egg production and haematin excretion were correlated for uninfected bloodfed mosquitoes; the presence of P.y. nigeriensis in the blood affected this relationship. Reduced fecundity was associated with a significant reduction of bloodmeal size (measured by haematin excretion) in mosquitoes which ingested gametocytaemic blood. The bloodmeal size in mosquitoes fed on parasitaemic blood without gametocytes was not significantly reduced. The use of haematin assays for determination of bloodmeal size in mosquitoes is discussed.     

Evaluation of the antiparasitic activity of the chitosan-silver nanocomposites in the treatment of experimentally infested pigeons with Pseudolynchia canariensis

This study aimed to evaluate the efficacy of chitosan-silver nanocomposites in the treatment of experimentally infested pigeons with Pseudolynchia canariensis (P. canariensis) with evaluation of different immunological parameters before and after treatment. Therefore, fourteen birds were divided into 2 groups; group1(infested group including 12 birds) which subdivided into 6 sub-groups experimentally infested pigeons 2 pigeons each, and five group of them were treated with chitosan-silver nanocomposites and sub-group number 6 was treated with deltamethrin while, group 2 including two pigeons were kept as control negative ones. P. canariensis flies distributed under the wing and /or under the tail in infested group and these pigeons showed significantly lower RBCs and higher WBCs than that in non-infested pigeons. The cell mediated immune response against experimentally infested pigeons with P. canariensis was studied. P. canariensis infestation in pigeons have a negative impact on pigeon’s blood parameters, increase TNF-α and IL-1β cytokines levels. This study cleared out the role of P. canariensis in the induction of a case of oxidative stress indicated by high level of nitric oxide and malondialdehyde (MDA) with low antioxidant capacity in shape of reduced zinc concentration in the sera of experimentally infested pigeon. Chitosan-silver nanocomposite has a promising effect in the elimination of P. canariensis infestation in pigeons.