p NMR Studies of Rat Liver Cold Preservation with Histidine-Buffered Lactobionate Solution

The efficiency of a preservation medium, histidine-buffered lactobionate solution (HBLS), was determined by measuring post-ischemic recoveries of ATP and intracellular pH under Krebs-Henseleit buffer (KHB) perfusion. We used NMR spectroscopy to study the effect of 24-h cold ischemia, followed by 4°C then 37°C reperfusion on the isolated rat liver. Three media were compared: University of Wisconsin solution (UW-lactobionate); Bretschneider's solution (HTK); HBLS and HBLS supplemented with 2 mM Gly and 2 mM Cys (HBLSg2) or with 10 mM Gly and 2 mM Cys (HBLSg10). All values were compared to control values measured during pre-ischemic cold perfusion with KHB (ATP = 8.60 ± 0.6 μmol/g of dry weigh and pH_in = 7.41 ± 0.05). The main result from ³¹p NMR data concerned ATP recovery during cold reperfusion, which was significantly higher in the HBLS group (112 ± 10%) as compared to the UW and HTK groups (around 66%). The presence of glycine decreased ATP recovery (88 ± 8% in HBLSg2, 79 ± 15% in HBLSg10). Higher values of recovered pH_in were observed in livers stored in histidine buffered solutions (around 7.30) as compared to UW (around 7.20); histidine was by ¹³C NMR proved to accumulate in the liver cells, thus ensuring a good buffering capacity. The thermal transition induced a decrease in both ATP level and pH_in in all groups. This might be the result of a stimulation of the carbohydrate metabolism (as demonstrated by ¹³C NMR) especially when glycine was present in the storage solution.     

Simulation of bone tissue formation within a porous scaffold under dynamic compression

A computational model of mechanoregulation is proposed to predict bone tissue formation stimulated mechanically by overall dynamical compression within a porous polymeric scaffold rendered by micro-CT. Dynamic compressions of 0.5–5% at 0.0025–0.025 s⁻¹ were simulated. A force-controlled dynamic compression was also performed by imposing a ramp of force from 1 to 70 N. The model predicts homogeneous mature bone tissue formation under strain levels of 0.5–1% at strain rates of 0.0025–0.005 s⁻¹. Under higher levels of strain and strain rates, the scaffold shows heterogeneous mechanical behaviour which leads to the formation of a heterogeneous tissue with a mixture of mature bone and fibrous tissue. A fibrous tissue layer was also predicted under the force-controlled dynamic compression, although the same force magnitude was found promoting only mature bone during a strain-controlled compression. The model shows that the mechanical stimulation of bone tissue formation within a porous scaffold closely depends on the loading history and on the mechanical behaviour of the scaffold at local and global scales.     

OSIP1 is a self-assembling DUF3129 protein required to protect fungal cells from toxins and stressors

Secreted proteins are key players in fungal physiology and cell protection against external stressing agents and antifungals. Oak stress-induced protein 1 (OSIP1) is a fungal-specific protein with unknown function. By using Podospora anserina and Phanerochaete chrysosporium as models, we combined both in vivo functional approaches and biophysical characterization of OSIP1 recombinant protein. The P. anserina OSIP1^Δ mutant showed an increased sensitivity to the antifungal caspofungin compared to the wild type. This correlated with the production of a weakened extracellular exopolysaccharide/protein matrix (ECM). Since the recombinant OSIP1 from P. chrysosporium self-assembled as fibers and was capable of gelation, it is likely that OSIP1 is linked to ECM formation that acts as a physical barrier preventing drug toxicity. Moreover, compared to the wild type, the OSIP1^Δ mutant was more sensitive to oak extractives including chaotropic phenols and benzenes. It exhibited a strongly modified secretome pattern and an increased production of proteins associated to the cell-wall integrity signalling pathway, when grown on oak sawdust. This demonstrates that OSIP1 has also an important role in fungal resistance to extractive-induced stress.     

OdhI dephosphorylation kinetics during different glutamate production processes involving Corynebacterium glutamicum

In Corynebacterium glutamicum, the activity of the 2-oxoglutarate dehydrogenase complex was shown to be controlled by the phosphorylation of a 15-kDa protein OdhI by different serine/threonine protein kinases. In this paper, the phosphorylation status and kinetics of OdhI dephosphorylation were assessed during glutamate producing processes triggered by either a biotin limitation or a temperature upshock from 33°C to 39°C. A dephosphorylation of OdhI in C. glutamicum 2262 was observed during the biotin-limited as well as the temperature-induced glutamate-producing process. Deletion of pknG in C. glutamicum 2262 did not affect the phosphorylation status of OdhI during growth and glutamate production phases triggered by a temperature upshock, though a 40% increase in the specific glutamate production rate was measured. These results suggest that, under the conditions analyzed, PknG is not the kinase responsible for the phosphorylation of OdhI in C. glutamicum 2262. The phosphorylation status of OdhI alone is, as expected, not the only parameter that determines the performance of a specific strain, as no clear relation between the specific glutamate production rate and OdhI phosphorylation level was demonstrated.     

Decrease in oxidative phosphorylation yield in presence of butyrate in perfused liver isolated from fed rats

Background  

Butyrate is the main nutrient for the colonocytes but the effect of the fraction reaching the liver is not totally known. A decrease in tissue ATP content and increase in respiration was previously demonstrated when livers were perfused with short-chain fatty acids (SCFA) such as butyrate, or octanoate.     

Nucleopeptidic Bioconjugates Containing a Sulfamide Bridge: Linkage via the Mitsunobu Reaction

Abstract

The synthesis of compounds connecting unprotected 2′-deoxyribonucleosides (T,dC,dG,dA) with N-Boc sulfamoyl derivatives of natural aminoacid esters (Phe, Asp) was carried out by Mitsunobu reaction, using regiospecific coupling. The created link was a priori non-hydrolyzable in biological conditions.     

Triple Helix Forming α-Oligonucleotides Containing 5-Methylcytosine and/or 5-Bromouracil

Abstract

Stability of αββ triplexes constituted with an α-strand containing 5-methylcytosine and/or 5-bromouracil was studied at a wide pH range. Introduction of 5-methylcytosine increases hybridization stability of the triplexes while introduction 5-broumouracil does not change it.