Action Of Exo Research Articles

Resveratrol-primed exosomes strongly promote the recovery of motor function in SCI rats by activating autophagy and inhibiting apoptosis via the PI3K signaling pathway

Spinal cord injury (SCI) is a traumatic condition of the central nervous system (CNS) that can cause paralysis of the limbs. The molecular mechanisms of neural repair following SCI remain unclear and no effective treatment for SCI currently exists, since drugs have difficulty crossing the blood-brain barrier (BBB). The present study aimed to investigate whether exosomes could be used as specific carriers of resveratrol for induction of neuronal autophagy both in vitro and in vivo for the treatment of SCI. The results indicate that exosomes are able to enhance the solubility of resveratrol and enhance penetration of the drug through the BBB, thereby increasing its concentration in the CNS. Exosomes derived from resveratrol-treated primary microglia (Exo + Res) assisted the rehabilitation of paralyzed limbs in rats. Restoration of neural function following SCI was mediated through increased induction of autophagy and inhibition of apoptosis of neurons both in vitro and in vivo via activation of the PI3K signaling pathway. The mechanism of action of Exo + Res may be associated with the PI3K inhibitor 3-methyladenine (3-MA) in primary spinal neurons. The results suggest that Exo + Res are highly effective at crossing the BBB with good stability, suggesting they have potential for enhancing targeted drug delivery and the recovery of neuronal function in SCI therapy, likely associated with the induction of autophagy and inhibition of apoptosis via the PI3K signaling pathway.

Facile strategy to enhance the specificity and sensitivity of hairpin molecular devices for detecting pax-5a gene by an integration probe and the specific function of exonuclease Ⅲ

A label-free DNA sensor is proposed to detect the acute leukemia gene pax-5a, in which the system only relies on a functional hairpin probe (HP) containing a G-quadruplex sequence and the cleavage of exonuclease Ⅲ (Exo Ⅲ). Non-single-stranded fragments in the HP structure can also be digested, including double-stranded stems and G-quadruplex fragments, by Exo Ⅲ at a certain concentration. In the presence of pax-5a, the HP sequence can be cyclically opened through the shearing action of Exo Ⅲ, causing a large number of G-quadruplex fragments to be released. In the absence of pax-5a, the G-quadruplex fragment is digested by Exo Ⅲ. The G-quadruplex-hemin DNAzyme can catalyze the oxidation of 3, 3′, 5, 5′-tetramethylbenzidine by H2O2 to generate a detectable visible absorption signal, thereby achieving facile and visual detection of acute leukemia gene pax-5a. The detection limit of the label-free DNA sensor that we proposed is as low as 25 fM in the pax-5a concentration range of 25 fM to 25 nM. In addition, the sensor has good discrimination ability of non-target genes and excellent detection ability in complex matrices. Therefore, the DNA sensor shows great potential in biological analysis and clinical diagnosis.

Co-expression of Exo-inulinase and Endo-inulinase Genes in the Oleaginous Yeast Yarrowia lipolytica for Efficient Single Cell Oil Production from Inulin.

Yarrowia lipolytica is a promising platform for the single cell oil (SCO) production. In this study, a transformant X+N8 in which exo- and endo-inulinase genes were co-expressed could produce an inulinase activity of 124.33U/mL within 72h. However, the inulinase activity of a transformant X2 carrying a single exo-inulinase gene was only 47.33U/mL within 72h. Moreover, the transformant X+N8 could accumulate 48.13% (w/w) SCO from inulin and the cell dry weight reached 13.63g/L within 78h, which were significantly higher than those of the transformant X2 (41.87% (w/w) and 11.23g/L) under the same conditions. In addition, inulin hydrolysis and utilization of the transformant X+N8 were also more efficient than those of the transformant X2 during the fermentation process. These results demonstrated that the co-expression of the exo- and endo-inulinase genes significantly enhanced the SCO production from inulin due to the improvement of the inulinase activity and the synergistic action of exo- and endo-inulinase. Besides, over 95.01% of the fatty acids from the transformant X+N8 were C16-C18, especially C18:1 (53.10%), suggesting that the fatty acids could be used as feedstock for biodiesel production.

Highly sensitive exonuclease III-assisted fluorometric determination of silver(I) based on graphene oxide and self-hybridization of cytosine-rich ss-DNA

We report on a method for selective fluorometric determination of Ag(I) based on the use of graphene oxide (GO) and by exploiting exonuclease III-assisted signal amplification. In the presence of Ag(I), the self-hybridization of cytosine-rich ss-DNA labeled with the fluorescent tag FAM leads to the formation of a ds-DNA that is cleaved by the catalytic action of Exo III. In the absence of Ag(I) ions, however, the tagged ss-DNA is adsorbed on the surface of GO and its fluorescence is quenched. As a result, the induction of the process by Ag(I) leads to a strong amplification of fluorescence. The method allows Ag(I) ions to be determined in the 1 to 250 nM concentration range, with a 0.1 nM detection limit. The recoveries range from 93.3 to 105.1 %, with RSDs of <8.2 %.

Xylan degradation improved by a combination of monolithic columns bearing immobilized recombinant β-xylosidase from Aspergillus awamori X-100 and Grindamyl H121 β-xylanase.

Synergistic action of exo- and endohydrolazes is preferred for effective destruction of biopolymers. The main purpose of the present work was to develop an efficient tool for degradation of xylan. Macroporous lab-made monolithic columns and commercial CIM-Epoxy disk were used to immobilize the recombinant β-xylosidase from Aspergillus awamori and Grindamyl β-xylanase. The efficiency of xylan degradation using the low-loaded β-xylosidase column appeared to be four times higher than for the in-solution process and about six times higher than for the high-loaded bioreactor. Disk bioreactor with the Grindamil β-xylanase operated in a recirculation mode has shown noticeable advantages over the column design. Additionally, a system comprised of two immobilized enzyme reactors (IMERs) was tested to accelerate the biopolymer hydrolysis, yielding total xylan conversion into xylose within 20 min. Fast online monitoring HPLC procedure was developed where an analytical DEAE CIM disk was added to the two-enzyme system in a conjoint mode. A loss of activity of immobilized enzymes did not exceed 7% after 5 months of the bioreactor usage. We can therefore conclude that the bioreactors developed exhibit high efficiency and remarkable long-term stability.

The inhibitory action of exo- and endocannabinoids on [3H]GABA release are mediated by both CB1 and CB2 receptors in the mouse hippocampus

Exogenous and endogenous cannabinoids play an important role in modulating the release of neurotransmitters in hippocampal excitatory and inhibitory networks, thus having profound effect on higher cognitive and emotional functions such as learning and memory. In this study we have studied the effect of cannabinoid agonists on the potassium depolarization-evoked [3H]GABA release from hippocampal synaptosomes in the wild-type (WT) and cannabinoid 1 receptor (CB1R)-null mutant mice.All tested cannabinoid agonists (WIN55,212-2, CP55,940, HU-210, 2-arachidonoyl-glycerol, 2-AG; delta-9-tetra-hydrocannabinol, THC) inhibited [3H]GABA release in WT mice with the following rank order of agonist potency: HU-210>CP55,490>WIN55,212-2>>2-AG>THC. By contrast, 2-AG and THC displayed the greatest efficacy eliciting almost complete inhibition of evoked [3H]GABA efflux, whereas the maximal inhibition obtained by HU-210, CP55,490, and WIN55,212-2 were less, eliciting not more than 40% inhibition. The inhibitory effect of WIN55,212-2, THC and 2-AG on evoked [3H]GABA efflux was antagonized by the CB1 receptor inverse agonist AM251 (0.5μM) in the WT mice. In the CB1R knockout mice the inhibitory effects of all three agonists were attenuated. In these mice, AM251 did not antagonize, but further reduced the [3H]GABA release in the presence of the synthetic agonist WIN55,212-2. By contrast, the concentration-dependent inhibitory effects of THC and 2-AG were partially antagonized by AM251 in the absence of CB1 receptors. Finally, the inhibition of evoked [3H]GABA efflux by THC and 2-AG was also partially attenuated by AM630 (1μM), the CB2 receptor-selective antagonist, both in WT and CB1 knockout mice.Our data prove the involvement of CB1 receptors in the effect of exo- and endocannabinoids on GABA efflux from hippocampal nerve terminals. In addition, in the effect of the exocannabinoid THC and the endocannabinoid 2-AG, non-CB1, probably CB2-like receptors are also involved.

Mode of Action of Exo- and Endo-Type Cellulase from Fungi in the Hydrolysis of Various Substrates

Mode of Action of Exo- and Endo-Type Cellulase from Fungi in the Hydrolysis of Various Substrates

How is bursting mode maintained under the action of exo- and endogenous factors?

The investigation is concerned with slow oscillations of the intracellular calcium concentration, fast oscillations of the membrane potential, and their interactions during bursting generation. The question associated with functioning of the bursting mode is considered: how is the condition of simultaneous existence of the slow and fast membrane potential oscillations (necessary for the bursting mode) realized under exo- and endogenous actions that result in changes in the neuron membrane parameters? To answer the question the regions of existence of fast, slow and bursting oscillations have been constructed on different parameter planes. The study has revealed that in spite of the fact the regions of the existence of the slow and fast oscillations are sensitive to parameter variations, the condition of the bursting mode existence remains due to invariance of the spatial vicinity of these regions. Furthermore, it has been demonstrated that there exists the possibility to enhance the maximal conductances of the calcium-dependent currents simultaneously such that even strong variations of their values do not induce changes of the oscillations mode (i.e. it remains bursting).

Upper limits for endogenous oligogalacturonides and free galacturonic acid in rose cell-suspension cultures: Implications for the action of Exo- and Endo-polygalacturonases in vivo

Summary To test for the possible production of biologically-active oligogalacturonides by healthy plant cells, we fed d-[U- 14 C] glucose to rose ( Rosa sp., cv. ) cell-suspension cultures and analysed the anionic metabolites that accumulated in the medium. Acidic 14 C-polysaccharides and a novel acidic 14 C-oligosaccharide accumulated, but steady-state concentrations of [ 14 C] oligogalacturonides (degree of polymerisation 2–12) were undetectable (≤50μg/L). The product of [ 14 C]oligogalacturonide turnover would be free [ 14 C] galacturonic acid; however, this too failed to accumulate in the rose culture medium (steady-state concentration ≤200nM), even though the cultures lacked the ability to take up or metabolise exogenous d-galacturonic acid. Therefore, healthy rose cell cultures lacked sufficient endo-polygalacturonase activity (EC 3.2.1.15) to generate detectable oligogalacturonides from endogenous pectic polysaccharides. Also, the absence of detectable [ 14 C]galacturonic acid shows that exo-polygalacturonase (EC 3.2.1.67), known to be present in the apoplast of these rose cells, failed to act at an appreciable rate on any endogenous apoplastic substrate, either soluble or wall-bound. We argue that the main natural substrate of plant exo-polygalacturonase in vegetative tissues is microbially-generated pectic fragments after infection.

Mode of Action of Exo- and Endo-Type Cellulases from Irpex lacteus in the Hydrolysis of Cellulose with Different Crystallinities

The mode of action of two highly purified cellulases of exo- and endo-types from Irpex lacteus was investigated by using pure cellulosic substrates with different crystallinities derived from cellulose I. Exo-type cellulase saccharified all celluloses more effectively than endo-type enzyme, and the saccharification activities of both cellulases similarly increased with decreasing crystallinity of cellulose. The DP-lowering activity of exo-type cellulase remained similar for celluloses with higher crystallinity, while this cellulase showed a degradation mode resembling that of the endo-type enzyme for the substrates with lower crystallinity. Compared with exo-type cellulase, endo-type cellulase remarkably decreased the DP of cellulose with higher crystallinity, while this activity was abated for cellulose with lower crystallinity. Thus, the effects of both cellulases became similar in the degradation of amorphous substrates such as H3PO4-treated cellulose. Endo-type cellulase produced several kinds of cellooligosaccharide from all kinds of cellulose used, while the product of the exo-type enzyme was only cellobiose from crystalline cellulose such as cotton and cotton linter even after a 12-h incubation period. The results indicate that each cellulase shows a typical mode of action (exo or endo) for crystalline cellulose, but that their characteristic modes of attack may change with decreasing crystallinity of cellulose.

Modes of Action of Exo- and Endo-Cellulases in the Degradation of Celluloses I and II1

Cotton and Valonia-cellulose (cellulose I) were readily attacked by endo-cellulase of a highly endowise-hydrolysis type, with a sharp decrease in the degree of polymerization, while the simultaneous production of reducing sugar was low. In contrast, viscose rayon and alkali-cellulose (cellulose II) showed little lowering of the degree of polymerization by endo-cellulase, though the effect was slightly greater than with an exo-cellulase, while the simultaneous production of reducing sugar was very high. The synergistic effect of exo- and endo-cellulases on the hydrolysis of cellulose was much higher with cellulose I than with cellulose II. These results can be explained in terms of differences in the polarity of cellulose chains and in the crystallinity and/or ultrastructure of the cellulose fibers.

Models for deploymerizing enzymes: criteria for discrimination of models

Several models for the action of alpha amylase have been proposed to account for the nonrandom distribution of oligosaccharides in the amylase digests of polysaccharides. The preferred-attack model attempts to account for the nonrandom distribution by assuming that the probability for bond cleavage depends upon the position of the bond in the chain. The repetitive, or multiple-attack, model suggests that the nonrandom distribution of oligosaccharides arises because an amylase can form a cage-like complex with a substrate and attack it several times during a single encounter. The multiple-enzyme or dual-site model suggests that the nonrandom yield of oligosaccharides arises from the combined action of exo- and endo-enzymes. The effects of pH, inhibitors, and substrate chain-length on enzyme action have been studied in several laboratories to determine which of the three action-patterns best describes the action of alpha amylase. The influence of these variables on product distributions or enzyme action-patterns are mathematically modeled in the Appendix. The experimental data on porcine-pancreatic alpha amylase are discussed in the light of the derivations.