A.M. García-Martínez

Age‐related increase in the immunoproteasome content in rat hippocampus: molecular and functional aspects

Alterations in the proteasome activity in the CNS have been described during aging. However, a detailed study of all proteasome subunits is actually lacking. We have analyzed, in vivo, the age‐related modifications in the molecular composition of hippocampal proteasomes. We found that the immunoproteasome/proteasome ratio was increased in aged hippocampus. The processing of the low‐molecular‐mass protein (LMP)7/β5i subunit, practically absent in young hippocampus, was increased in aged animals. Among the potential factors underlying these modifications we evaluated the neuroinflammation and the transcription factor Zif268. Lipopolysaccharide (LPS)‐induced neuroinflammation in young rats, up‐regulated the expression of immunoproteasome subunits and increased the processing of the LMP7/β5i protein. Moreover, the hydrophobicity of cellular peptides, analyzed by liquid chromatography, increased in both, young LPS‐injected animals and aged rats, suggesting that immunoproteasomes including the LMP7/β5i subunit could, at least in part, account for this modification. Also, the mRNA expression of the transcription factor Zif268, which down‐regulates the immunoproteasome subunit LMP7/β5i by binding to sequences within the promoter regions, was decreased in both, aged hippocampus and young LPS‐injected animals. Finally, we found that spatial memory training in young animals, a situation in which the expression of Zif268 is increased, modified the mRNA expression of the constitutive and catalytic subunits in an opposite manner. Based on present data, we propose that the age‐related increases in the content of hippocampal immunoproteasome is mostly because of neuroinflammatory processes associated to aging.

Application of MCPA herbicide on soils amended with biostimulants: short-time effects on soil biological properties.

In this paper we studied in the laboratory the effect of MCPA herbicide at a rate of 1.5lha(-1) (manufactures rate recommended) on biological properties of a Plagic Antrosol amended with four biostimulants (WCDS, wheat condensed distillers soluble; PA-HE, hydrolyzed poultry feathers; CGHE, carob germ enzymatic extract; and RB, rice bran extract). Seven hundred grams of soil were mixed with WCDS at a rate of 10%, CGHE at a rate of 4.7%, PA-HE at a rate of 4.3%, and RB at a rate of 4.4%, respectively, in order to applying the same amount of organic matter to the soil (16.38 g organic matter). An unamended polluted and amended non-polluted soil were used as control. For all treatments, the soil ergosterol, dehydrogenase, urease, and phosphatase activities were measured at two incubation times (0 and 60 d). The 16S rDNA-DGGE profiles in all treatments were determined at the beginning and end of the incubation period. The results indicated that at the end of the incubation period and compared with the control soil, the dehydrogenase, urease and phosphatase activities and ergosterol decreased 39.3%, 20%, 15.7% and 56.5%, respectively in the non-organic amended polluted soil. The application of organic matter to unpolluted soil increased the enzymatic activities and ergosterol. However, this stimulation was higher in the soil amended with RB, followed by PA-HE, WCDS and CGHE. The application of herbicide in organic-amended soils decreased the enzymatic activities and ergosterol content. However, this decrease was lower than for the non-amended herbicide polluted soil. Possibly the low molecular weight protein content easily assimilated by soil microorganisms and the adsorption capacity of humic substances are responsible for less inhibition of these enzyme activities and soil ergosterol. The 16S rDNA-DGGE profiles indicated that herbicide did not negatively affect soil bacterial biodiversity.

Relationships between biological and chemical parameters on the composting of a municipal solid waste.

The aim of this work was to study the combined relationships between biological and chemical parameters with the humification index (HI) and degree of polymerisation (DP) during the composting of a municipal solid waste. During the composting, biological parameters increased during the first 15 days. After the second week, the dehydrogenase, urease, beta-glucosidase, phosphatase and arylsulphatase activities decreased 64%, 50.1%, 49.6%, 24.1% and 58.3%, respectively, compared with their initial values, possibly due to the decreasing of the water-soluble carbon content (WSC). The WSC contents decreased possibly due to the degradation of the microorganisms as carbon and energy sources. The HI increased (66% compared to the initial values) indicating an increase in the structural complexity of the humic substances. The factorial analysis indicates a relationship between the biological properties, WSC and HI and DP. The linear regression analysis indicates the high correlation coefficients found between the HI and DP with chemical and biological properties, and therefore, it can be concluded that combination of chemical and biological parameters can be used to determine the compost maturity.

Protection against free radicals (UVB irradiation) of a water-soluble enzymatic extract from rice bran. Study using human keratinocyte monolayer and reconstructed human epidermis

The antioxidant capacity of a water-soluble enzymatic extract from rice bran (EERB) has been tested in two cell models: keratinocyte monolayers and human reconstructed epidermis. Cells were incubated in culture medium in presence of different amounts of EERB and were UVB irradiated. Cell population assessment (MTT assay) and MDA (malonaldehyde) production were evaluated. The EERB did not induce cytotoxic effect for concentrations inferior or equal to 100 microg/mL. Human keratinocyte monolayers were protected of irradiation preventing 33% the lipid peroxidation process at concentration of 10 microg/ml of EEBR. In reconstructed human epidermis, 100 microg/mL decreased lipid peroxidation process by 44% (p<0.01) with regards to irradiated negative control. This effect was comparable to that of vitamin E at 600 microg/mL. Our data indicate that EERB is potentially able to efficiently counteract UVB-induced response. The EERB, diluted at 10% with water has very good skin compatibility. This product showed a sun protection factor of 4.8+/-0.3. Thus we can propose EERB as a useful natural standardized extract in skin photoprotection with promising applications in the field of dermatology.

Enzymatic Vegetable Organic Extracts as Soil Biochemical Biostimulants and Atrazine Extenders

The purpose of this study was to gather information on the potential effects of organic biostimulants on soil activity and atrazine biodegradation. Carob germ enzymatic extract (CGEE) and wheat condensed distiller solubles enzymatic extract (WCDS-EE) have been obtained using an enzymatic process; their main organic components are soluble carbohydrates and proteins in the form of peptides and free amino acids. Their application to soil results in high biostimulation, rapidly increased dehydrogenase, phosphatase and glucosidase activities, and an observed atrazine extender capacity due to inhibition of its mineralization. The extender capacity of both extracts is proportional to the protein/carbohydrate ratio content. As a result, these enzymatic extracts are highly microbially available, leading to two independent phenomena, fertility and an atrazine persistence that is linked to increased soil activity.

Effects of Prochloraz fungicide on soil enzymatic activities and bacterial communities.

We studied in the laboratory the effect of Prochloraz fungicide on the biological properties (soil enzymatic activities and soil bacterial communities) of a Plaggic Anthrosol. Five hundred grams of soil (<2mm) was mixed with three dosages of Prochloraz (1, 2, and 4 l ha(-1)) for 83 days. A non-Prochloraz polluted soil was used as control. Following commercial recommendations, fungicide was applied four times during the incubation experiment. For all treatments, the soil ergosterol and levels of dehydrogenase, urease, β-glucosidase, and phosphatase activity were measured at nine different times (0, 1, 21, 22, 41, 42, 62, 63, and 83 days). The 16S rDNA-DGGE profiles in all treatments were determined at the beginning and end of the incubation period. At the end of the experiment, a significant decrease in ergosterol by 72.3%, 80.8%, and 83.1%, compared with control soil, was observed when 1, 2, and 4 l ha(-1), respectively, was added. Soil enzymatic activities increased when the Prochloraz applied to the soil increased, possibly because the fungicide is used by bacterial communities as a source of energy and nutrients. The 16S rDNA-DGGE profiles indicated that the fungicide did not negatively affect soil bacterial biodiversity. These results suggested that the fungicide Prochloraz has a very interesting agronomic effect, possibly due to the negative effect on soil fungal population stimulating the growth of soil bacterial activity.

Obtaining edaphic biostimulants/biofertilizers from different sewage sludges. Effects on soil biological properties

We studied the influence of six edaphic biostimulants/biofertilizers (BSs) manufactured by the pH-stat method from different sewage sludge (SS): SS1 (an anaerobic mature sludge, one year old), SS2 (an aerobic young sludge, without maturation) and SS3 (an aerobic mature sludge, four months old), not previously autoclaved (A) and autoclaved (B), by analysing their effects on soil biological properties. Soil enzymatic activities were measured at 1, 3, 5, 7, 15, 30 and 60 days of the incubation period, whereas the 16S rDNA-DGGE profiles were determined at 0, 5 and 60 days. The enzymatic activities were significantly stimulated. The highest stimulation was found in the B2 treatment followed by B3, A2, A3, B1 and A1 treatments. Increasing the number of lower molecular weight proteins in the BS enhances the stimulation of soil enzymatic activities. The application of BS caused at 5 days of the incubation period temporal variations in the soil bacterial community structure.

Central Administration of Aminoprocalcitonin Inhibits Food Intake and Stimulates the Hypothalamic-Pituitary-Adrenal Axis in Rats via the Corticotrophin-Releasing Factor System

Aminoprocalcitonin (N‐PCT), a neuroendocrine peptide derived from procalcitonin, reduces food intake and body weight when administered centrally in rats. We have recently shown that N‐PCT is expressed in brain areas known to be involved in energy homeostasis, including the paraventricular nucleus (PVN) of the hypothalamus, which contains a prominent population of corticotrophin‐releasing factor (CRF)‐synthesising neurones. CRF plays a pivotal role in the regulation of the hypothalamic‐pituitary adrenal (HPA) axis and food intake. However, little is known about functional interactions of N‐PCT and CRF. In the present study, we found endogenous N‐PCT protein in the rat PVN. We also showed N‐PCT immunoreactivity in PVN co‐localised with NeuN, a neuronal marker, or glial fibrillary acidic protein, an astrocyte marker. Double staining immunohistochemistry revealed that N‐PCT co‐localised with CRF in parvocellular neurones of the PVN. Intracerebroventricular N‐PCT administration increased CRF mRNA and content in the hypothalamus, suggesting that N‐PCT stimulates the HPA axis and suppresses food intake and body weight via CRF‐dependent pathways. In keeping with this, i.c.v. co‐injection of D‐Phe‐CRF12‐41, a CRF receptor antagonist, significantly attenuated N‐PCT‐induced reduction in food intake and body weight in a dose‐dependent manner. Furthermore, i.c.v. administration of N‐PCT increased plasma adrenocorticotrophic hormone and corticosterone concentrations and induced the expression of Fos protein, a marker of neuronal activity, in parvocellular CRF neurones. These data collectively support the hypothesis that N‐PCT inhibits food intake and body weight and stimulates the HPA axis via CRF‐mediated pathways.

Response of Biological Properties to the Application of Banvel ® (2,4-D + MCPA + Dicamba) Herbicide in Soils Amended with Biostimulants

In this paper the effect of Banvel® (2,4-D + MCPA + Dicamba) herbicide at a rate of 1.5 L ha−1 (manufactures rate recommended) in the biological properties of a Plagic Antrosol amended with four biostimulants (WCDS, wheat condensed distillers soluble; PA-HE, hydrolyzed poultry feathers; CGHE, carob germ enzymatic extract; and RB, rice bran extract) was studied. Seven hundred grams of soil were mixed with WCDS at a rate of 10%, CGHE at a rate of 4.7%, PA-HE at a rate of 4.3%, and RB at a rate of 4.4%, respectively, in order to apply the same amount of organic matter to the soil (16.38 g organic matter). An unamended polluted and amended non-polluted soil was used as control. For all treatments, the soil ergosterol, dehydrogenase, urease, and phosphatase activities were measured at two incubation times (1 and 60 days). The 16S rDNA-DGGE profiles in all treatments were determined at the beginning and end of the incubation period. The results indicated that at the end of the incubation period and compared with the control soil, the dehydrogenase, urease and phosphatase activities and ergosterol decreased 46.4, 17, 19.1 and 47.8%, respectively in the non-organically amended polluted soil. The application of organic matter to unpolluted soil increased phosphatase activity. However, this stimulation was higher in the soil amended with PA-HE, followed by RB, WCDS and CGHE. The application of herbicide to the amended soils decreased enzymatic activities and ergosterol content. However, these decreases were lower than those in the non-amended herbicide polluted soil. Possibly the low molecular weight proteins easily assimilated by soil microorganisms provided with the biostimulants are responsible for the lower inhibition of these enzyme activities and soil ergosterol in the amended soils. The 16S rDNA-DGGE profiles indicated that herbicide did not negatively affect soil bacterial biodiversity.