Predicting anaerobic biogasification potential of ingestates and digestates of a full-scale biogas plant using chemical and biological parameters

The aim of this work was to develop simple and fast tests to predict anaerobic biogasification potential (ABP) of ingestates and digestates from a biogas plant. Forty-six samples of both ingestates and digestates were collected within an eight-month observation period and were analyzed in terms of biological and chemical parameters, namely, ABP test, oxygen demand in a 20-h respirometric test (OD20), total solids (TS), volatile solids (VS), total organic carbon (TOC), total Kjeldahl nitrogen (TKN), ammonia, cell solubles (CS), acid detergent fibers (ADF), lignin (ADL), cellulose, and hemicellulose. Considering both quantitative (VS and TOC) and qualitative aspects (OD20 and CS) of organic matter (OM), four models (linear regressions; 0.80相似文献   

Effect of salt and pH on the reductive half-reaction of Mycobacterium tuberculosis FprA with NADPH

Despite a number of studies, the formation of the Michaelis complexes between ferredoxin-NADP (+) reductases and NADP(H) eluded detailed investigations by rapid kinetic techniques because of their high formation rates. Moreover, the reversible nature of the reaction of hydride ion transfer between these enzymes and NADPH prevented the obtainment of reliable estimates of the rate constant of the hydride transfer step. Here we show that by working at a high salt concentration, the mechanism of the reaction with NADPH of FprA, a Mycobacterium tuberculosis homologue of adrenodoxin reductase, is greatly simplified, making it amenable to investigation by rapid reaction techniques. The approach presented herein allowed for the first time the observation of the formation of the Michaelis complex between an adrenodoxin reductase-like enzyme and NADPH, and the determination of the related rate constants for association and dissociation. Furthermore, the rate constant for the reaction of hydride ion transfer between NADPH and FAD could be unambiguously assessed. It is proposed that the approach described should be applicable to other ferredoxin reductase enzymes, providing a valuable experimental tool for the study of their kinetic properties.     

Role of α2‐macroglobulin in regulating amyloid β‐protein neurotoxicity: protective or detrimental factor?

alpha2-Macroglobulin (alpha2M) has been identified as a carrier protein for beta-amyloid (Abeta) decreasing fibril formation and affecting the neurotoxicity of this peptide. The alpha2-macroglobulin receptor/low density lipoprotein receptor related protein (LRP) is involved in the internalization and degradation of the alpha2M/Abeta complexes and its impairment has been reported to occur in Alzheimer's disease. Previous studies have shown alpha2M to determine an enhancement or a reduction of Abeta toxicity in different culture systems. In order to clarify the role of alpha2M in Abeta neurotoxicity, we challenged human neuroblastoma cell lines with activated alpha2M in combination with Abeta. Our results show that in neuroblastoma cells expressing high levels of LRP, the administration of activated alpha2M protects the cells from Abeta neurotoxicity. Conversely, when this receptor is not present alpha2M determines an increase in Abeta toxicity as evaluated by MTT and TUNEL assays. In LRP-negative cells transfected with the full-length human LRP, the addition of activated alpha2M resulted to be protective against Abeta-induced neurotoxicity. By means of recombinant proteins we ascribed the neurotoxic activity of alpha2M to its FP3 fragment which has been previously shown to bind and neutralize transforming growth factor-beta. These studies provide evidence for both a neuroprotective and neurotoxic role of alpha2M regulated by the expression of its receptor LRP.     

The bone and the kidney

Renal tubular diseases may present with osteopenia, osteoporosis or osteomalacia, as a result of significant derangements in body electrolytes. In case of insufficient synthesis of calcitriol, as in renal failure, the more complex picture of renal osteodystrophy may develop. Hypothetically, also disturbed renal production of BMP-7 and Klotho could cause bone disease. However, the acknowledgment that osteocytes are capable of producing FGF23, a phosphaturic hormone at the same time modulating renal synthesis of calcitriol, indicates that it is also bone that can influence renal function. Importantly, a feed-back mechanism exists between FGF23 and calcitriol synthesis, while Klotho, produced by the kidney, determines activity and selectivity of FGF23. Identification of human diseases linked to disturbed production of FGF23 and Klotho underlines the importance of this new bone-kidney axis. Kidney and bone communicate reciprocally to regulate the sophisticated machinery responsible for divalent ions homeostasis and for osseous or extraosseous mineralisation processes.     

Analysis of alpha-fetoprotein heterogeneity by an improved crossed immunoaffinoelectrophoretic technique

The crossed immunoaffinoelectrophoretic procedure described here for the analysis of alpha-fetoprotein heterogeneity includes three major modifications of a procedure previously used by others: impregnation of the second dimension gel with methylated sugar derivatives; rigorous gel washing at 37 degrees C with the same derivatives; and an immunostaining technique. These simple changes have resulted in clear, sharp electrophoretic patterns at levels of total alpha-fetoprotein as low as 50 ng/ml, a sensitivity not previously approached with the method. Individual peaks representing as little as 7.5 ng/ml can be identified.     

Microsporogenesis of the feathers (fertile branches derived from annual buds) in Vitis vinifera,cv Picolit giallo

Abstract

Using light and electron microscopy, we have studied the microsporogenesis and tapetal development of the feathers in two different low producing clones of Picolit giallo (sp. Vitis vinifera). In these clones while the productivity of the main branches (fertile branches originated from buds, formed in the previous year, that remained silent during the winter) is very low, that of the feathers (fertile branches derived from annual buds) is always normal.

The microsporogenesis and tapetal development proceed normally in almost all the examined anthers; it is remarkable that at the tetrad stage the tapetal cells appear well structured without any degeneration symptom, unlike what observed for the main branches. Moreover in most of the mature anthers the pollen grains are numerous, pleinty of organelles and show sometimes thickenings in the callose layer under their wall. The tapetal cells of these anthers have disappeared. Only in few anthers we observed the presence of collapsed pollen grains and tapetal cells with anomalous development, that are still present when the pollen grains are mature. This rare situation for the feathers is on the contrary frequent for the main branches.     

Proteomics of human primary osteoarthritic chondrocytes exposed to extremely low-frequency electromagnetic fields (ELF EMFs) and to therapeutic application of musically modulated electromagnetic fields (TAMMEF)

Osteoarthritis (OA) is the most frequent joint disease, characterized by degradation of extracellular matrix and alterations in chondrocyte metabolism. Some authors reported that electromagnetic fields (EMFs) can positively interfere with patients affected by OA, even though the nature of the interaction is still debated. Human primary osteoarthritic chondrocytes isolated from the femoral heads of OA-patients undergoing to total hip replacement, were cultured in vitro and exposed 30?min/day for two weeks to extremely-low-frequency electromagnetic field (ELF) with fixed frequency (100?Hz) and to therapeutic application of musically modulated electromagnetic fields (TAMMEF) with variable frequencies, intensities and waveforms. Sham-exposed (S.E.) cells served as control group. Cell viability was measured at days 2, 7 and 14. After two weeks, cell lysates were processed using a proteomic approach. Chondrocyte exposed to ELF and TAMMEF system demonstrated different viability compared to untreated chondrocytes (S.E.). Proteome analysis of 2D-Electrophoresis and protein identification by mass spectrometry showed different expression of proteins derived from nucleus, cytoplasm and organelles. Function analysis of the identified proteins showed changes in related-proteins metabolism (glyceraldeyde-3-phosphate-dehydrogenase), stress response (Mn-superoxide-dismutase, heat-shock proteins), cytoskeletal regulation (actin), proteinase inhibition (cystatin-B) and inflammation regulatory functions (S100-A10, S100-A11) among the experimental groups (ELF, TAMMEF and S.E.). In conclusion, EMFs do not cause damage to chondrocytes, besides stimulate safely OA-chondrocytes and are responsible of different protein expression among the three groups. Furthermore, protein analysis of OA-chondrocytes treated with ELF and the new TAMMEF systems could be useful to clarify the pathogenetic mechanisms of OA by identifying biomarkers of the disease.     

The Shc protein Rai enhances T-cell survival under hypoxia

Hypoxia occurs in physiological and pathological conditions. T cells experience hypoxia in pathological and physiological conditions as well as in lymphoid organs. Indeed, hypoxia-inducible factor 1α (HIF-1α) affects T cell survival and functions. Rai, an Shc family protein member, exerts pro-survival effects in hypoxic neuroblastoma cells. Since Rai is also expressed in T cells, we here investigated its role in hypoxic T cells. In this work, hypoxia differently affected cell survival, proapoptotic, and metabolic programs in T cells, depending upon Rai expression. By using Jurkat cells stably expressing Rai and splenocytes from Rai^−/−mice, we demonstrated that Rai promotes T cell survival and affects cell metabolism under hypoxia. Upon exposure to hypoxia, Jurkat T cells expressing Rai show (a) higher HIF-1α protein levels; (b) a decreased cell death and increased Akt/extracellular-signal-regulated kinase phosphorylation; (c) a decreased expression of proapoptotic markers, including caspase activities and poly(ADP-ribose) polymerase cleavage; (d) an increased glucose and lactate metabolism; (e) an increased activation of nuclear factor-kB pathway. The opposite effects were observed in hypoxic splenocytes from Rai^−/−mice. Thus, Rai plays an important role in hypoxic signaling and may be relevant in the protection of T cells against hypoxia.     

Chromatin organization in Paracentrotus lividus eggs

After purification by buoyant density centrifugation in ethidium bromide - CsCl gradient and electrophoretic fractionation, the DNA fragments isolated from P. lividus egg nuclei incubated with micrococcal nuclease exhibit a typical oligomeric pattern. Analysis of chromatin samples digested to an increasing extent by micrococcal nuclease reveals that the structural organization of egg chromatin is heterogeneous, both in terms of repeat size and degree of sensitivity to nuclease attack. The nucleosomal repeats of P. lividus sperms and embryos up to the mesenchyme blastula stage have also been determined, for comparison.     

Highly efficient CRISPR-Cas9-mediated gene knockout in primary human B cells for functional genetic studies of Epstein-Barr virus infection

Gene editing is now routine in all prokaryotic and metazoan cells but has not received much attention in immune cells when the CRISPR-Cas9 technology was introduced in the field of mammalian cell biology less than ten years ago. This versatile technology has been successfully adapted for gene modifications in human myeloid cells and T cells, among others, but applications to human primary B cells have been scarce and limited to activated B cells. This limitation has precluded conclusive studies into cell activation, differentiation or cell cycle control in this cell type. We report on highly efficient, simple and rapid genome engineering in primary resting human B cells using nucleofection of Cas9 ribonucleoprotein complexes, followed by EBV infection or culture on CD40 ligand feeder cells to drive in vitro B cell survival. We provide proof-of-principle of gene editing in quiescent human B cells using two model genes: CD46 and CDKN2A. The latter encodes the cell cycle regulator p16^INK4a which is an important target of Epstein-Barr virus (EBV). Infection of B cells carrying a knockout of CDKN2A with wildtype and EBNA3 oncoprotein mutant strains of EBV allowed us to conclude that EBNA3C controls CDKN2A, the only barrier to B cell proliferation in EBV infected cells. Together, this approach enables efficient targeting of specific gene loci in quiescent human B cells supporting basic research as well as immunotherapeutic strategies.