Maintaining soil organic carbon (SOC) in frequently tilled, intensive organic vegetable production systems is a challenge that is not well understood. Compost and cover crops are often used to add ...organic matter to the soil in these systems. Compost contributes relatively stabilized carbon (C) while cover crops provide readily degradable (labile) organic matter. Our objectives were to quantify C inputs, and to assess the effects of urban yard-waste compost, winter cover crop frequency and cover crop type on SOC and labile C stocks during eight years of intensive, organic production that usually included two vegetable crops per year in a long-term systems study in Salinas, California. Total C inputs from pelleted fertilizer, compost, vegetable transplant potting mix, vegetable residue and cover crops, including estimates of below ground inputs, ranged from 40 to 108 Mg ha-1 in the five systems evaluated. Following a rapid decline in SOC stocks in year 1, compost had the largest effect on SOC stocks increasing mean SOC over years 2 to 8 by an average of 9.4 Mg ha-1, while increased cover crop frequency (annual vs. quadrennial) led to an additional 3.4 Mg ha-1 increase. In contrast, cover cropping frequency had the largest effect on permanganate oxidizable labile C (POX-C), increasing POX-C by 26% after 8 years. Labile POX-C was well correlated with microbial biomass C and nitrogen. Compost had the greatest effect on total SOC stocks, while increasing cover crop frequency altered the composition of SOC by increasing the proportion of labile C. These results suggest that frequent winter cover cropping has a greater potential than compost to increase nutrient availability and vegetable yields in high-input, tillage intensive vegetable systems.
In this chapter we describe conventional methods used for preparing renal tissue for transmission electron microscopy. We also describe a relatively new technique, serial block face scanning electron ...microscopy. Protocols are given for processing, sectioning, and imaging of tissue along with methods for obtaining quantitative data from the results.
Highly potent and broadly neutralizing anti-HIV-1 antibodies (bNAbs) have been used to prevent and treat lentivirus infections in humanized mice, macaques, and humans. In immunotherapy experiments, ...administration of bNAbs to chronically infected animals transiently suppresses virus replication, which invariably returns to pre-treatment levels and results in progression to clinical disease. Here we show that early administration of bNAbs in a macaque simian/human immunodeficiency virus (SHIV) model is associated with very low levels of persistent viraemia, which leads to the establishment of T-cell immunity and resultant long-term infection control. Animals challenged with SHIV
by mucosal or intravenous routes received a single 2-week course of two potent passively transferred bNAbs (3BNC117 and 10-1074 (refs 13, 14)). Viraemia remained undetectable for 56-177 days, depending on bNAb half-life in vivo. Moreover, in the 13 treated monkeys, plasma virus loads subsequently declined to undetectable levels in 6 controller macaques. Four additional animals maintained their counts of T cells carrying the CD4 antigen (CD4
) and very low levels of viraemia persisted for over 2 years. The frequency of cells carrying replication-competent virus was less than 1 per 10
circulating CD4
T cells in the six controller macaques. Infusion of a T-cell-depleting anti-CD8β monoclonal antibody to the controller animals led to a specific decline in levels of CD8
T cells and the rapid reappearance of plasma viraemia. In contrast, macaques treated for 15 weeks with combination anti-retroviral therapy, beginning on day 3 after infection, experienced sustained rebound plasma viraemia when treatment was interrupted. Our results show that passive immunotherapy during acute SHIV infection differs from combination anti-retroviral therapy in that it facilitates the emergence of potent CD8
T-cell immunity able to durably suppress virus replication.
Systems differed by annual compost additions (0 vs. 7.6 Mg ha-1 before each vegetable crop), cover crop type (legume-rye, mustard or cereal rye alone) and cover cropping frequency (quadrennially vs. ...annually planted). https://doi.org/10.1371/journal.pone.0307250.g005 thumbnail Download: * PPT PowerPoint slide * PNG larger image * TIFF original image Fig 10. Carbon inputs, soil organic carbon and permanganate oxidizable carbon ANOVA F-statistics and significance. https://doi.org/10.1371/journal.pone.0307250.t001 Supporting information S3 Fig. Soil organic carbon stocks over 8 years in three organic vegetable systems in Salinas, CA that all received annual compost additions (7.6 Mg ha-1 before each vegetable crop) and differed by annually planted cover crop type. https://doi.org/10.1371/journal.pone.0307250.s001 (TIF) S1 Table.
Histone protein modifications control fate determination during normal development and dedifferentiation during disease. Here, we set out to determine the extent to which dynamic changes to histones ...affect the differentiated phenotype of ordinarily quiescent adult glomerular podocytes. To do this, we examined the consequences of shifting the balance of the repressive histone H3 lysine 27 trimethylation (H3K27me3) mark in podocytes. Adriamycin nephrotoxicity and subtotal nephrectomy (SNx) studies indicated that deletion of the histone methylating enzyme EZH2 from podocytes decreased H3K27me3 levels and sensitized mice to glomerular disease. H3K27me3 was enriched at the promoter region of the Notch ligand Jag1 in podocytes, and derepression of Jag1 by EZH2 inhibition or knockdown facilitated podocyte dedifferentiation. Conversely, inhibition of the Jumonji C domain-containing demethylases Jmjd3 and UTX increased the H3K27me3 content of podocytes and attenuated glomerular disease in adriamycin nephrotoxicity, SNx, and diabetes. Podocytes in glomeruli from humans with focal segmental glomerulosclerosis or diabetic nephropathy exhibited diminished H3K27me3 and heightened UTX content. Analogous to human disease, inhibition of Jmjd3 and UTX abated nephropathy progression in mice with established glomerular injury and reduced H3K27me3 levels. Together, these findings indicate that ostensibly stable chromatin modifications can be dynamically regulated in quiescent cells and that epigenetic reprogramming can improve outcomes in glomerular disease by repressing the reactivation of developmental pathways.
Efficient use of nitrogen (N) is essential to protect water quality in high-input organic vegetable production systems, but little is known about the long-term effects of organic management on N mass ...balances. We measured soil N and tabulated N inputs (organic fertilizers, compost, irrigation water, atmospheric deposition, cover crop seed, vegetable transplant plugs and fixation by legume cover crops) and exports in harvested crops (lettuce, broccoli) over eight years to calculate soil surface and soil system N mass balances for the Salinas Organic Cropping Systems study in Salinas, CA. Our objectives were to 1) quantify the long-term effects of compost, cover crop frequency and cover crop type on soil N, cover crop and vegetable crop N uptake, and yield, and 2) tabulate N balances to assess the effects of these factors on N export in harvested crops, soil N storage and potential N loss. Results show that across all systems only 13 to 23% of N inputs were exported in harvest. Annual compost applications increased soil N stocks but had little effect on vegetable N uptake or yield, increasing the cumulative soil system N balance surplus over eight years by 999 kg ha-1, relative to the system receiving organic fertilizers alone. Annually planted winter cover crops increased N availability, crop uptake and export; however, biological N fixation by legumes negated the positive effect of increased harvest exports on the balance surplus in the legume-rye cover cropped system. Over eight years, rye cover crops improved system performance and reduced the cumulative N surplus by 384 kg ha-1 relative to the legume-rye mixture by increasing N retention and availability without increasing N inputs. Reduced reliance on external compost inputs and increased use of annually planted non-legume cover crops can improve efficient N use and cropping system yield, consequently improving environmental performance.
Exposure to ionizing radiation (IR) can result in the development of cutaneous fibrosis, for which few therapeutic options exist. We tested the hypothesis that bone marrow-derived mesenchymal stem ...cells (BMSC) would favorably alter the progression of IR-induced fibrosis. We found that a systemic infusion of BMSC from syngeneic or allogeneic donors reduced skin contracture, thickening, and collagen deposition in a murine model. Transcriptional profiling with a fibrosis-targeted assay demonstrated increased expression of interleukin-10 (IL-10) and decreased expression of IL-1β in the irradiated skin of mice 14 days after receiving BMSC. Similarly, immunoassay studies demonstrated durable alteration of these and several additional inflammatory mediators. Immunohistochemical studies revealed a reduction in infiltration of proinflammatory classically activated CD80(+) macrophages and increased numbers of anti-inflammatory regulatory CD163(+) macrophages in irradiated skin of BMSC-treated mice. In vitro coculture experiments confirmed that BMSC induce expression of IL-10 by activated macrophages, suggesting polarization toward a regulatory phenotype. Furthermore, we demonstrated that tumor necrosis factor-receptor 2 (TNF-R2) mediates IL-10 production and transition toward a regulatory phenotype during coculture with BMSC. Taken together, these data demonstrate that systemic infusion of BMSC can durably alter the progression of radiation-induced fibrosis by altering macrophage phenotype and suppressing local inflammation in a TNF-R2-dependent fashion.
Epigenetic regulation of oxidative stress is emerging as a critical mediator of diabetic nephropathy. In diabetes, oxidative damage occurs when there is an imbalance between reactive oxygen species ...generation and enzymatic antioxidant repair. Here, we investigated the function of the histone methyltransferase enzyme enhancer of zeste homolog 2 (EZH2) in attenuating oxidative injury in podocytes, focusing on its regulation of the endogenous antioxidant inhibitor thioredoxin interacting protein (TxnIP). Pharmacologic or genetic depletion of EZH2 augmented TxnIP expression and oxidative stress in podocytes cultured under high-glucose conditions. Conversely, EZH2 upregulation through inhibition of its regulatory microRNA, microRNA-101, downregulated TxnIP and attenuated oxidative stress. In diabetic rats, depletion of EZH2 decreased histone 3 lysine 27 trimethylation (H3K27me3), increased glomerular TxnIP expression, induced podocyte injury, and augmented oxidative stress and proteinuria. Chromatin immunoprecipitation sequencing revealed H3K27me3 enrichment at the promoter of the transcription factor Pax6, which was upregulated on EZH2 depletion and bound to the TxnIP promoter, controlling expression of its gene product. In high glucose-exposed podocytes and the kidneys of diabetic rats, the lower EZH2 expression detected coincided with upregulation of Pax6 and TxnIP. Finally, in a gene expression array, TxnIP was among seven of 30,854 genes upregulated by high glucose, EZH2 depletion, and the combination thereof. Thus, EZH2 represses the transcription factor Pax6, which controls expression of the antioxidant inhibitor TxnIP, and in diabetes, downregulation of EZH2 promotes oxidative stress. These findings expand the extent to which epigenetic processes affect the diabetic kidney to include antioxidant repair.
Positive allosteric modulators (PAMs) of the mu-opioid receptor (MOR) have been hypothesized as potentially safer analgesics than traditional opioid drugs. This is based on the idea that PAMs will ...promote the action of endogenous opioid peptides while preserving their temporal and spatial release patterns and so have an improved therapeutic index. However, this hypothesis has never been tested. Here, we show that a mu-PAM, BMS-986122, enhances the ability of the endogenous opioid Methionine-enkephalin (Met-Enk) to stimulate G protein activity in mouse brain homogenates without activity on its own and to enhance G protein activation to a greater extent than β-arrestin recruitment in Chinese hamster ovary (CHO) cells expressing human mu-opioid receptors. Moreover, BMS-986122 increases the potency of Met-Enk to inhibit GABA release in the periaqueductal gray, an important site for antinociception. We describe in vivo experiments demonstrating that the mu-PAM produces antinociception in mouse models of acute noxious heat pain as well as inflammatory pain. These effects are blocked by MOR antagonists and are consistent with the hypothesis that in vivo mu-PAMs enhance the activity of endogenous opioid peptides. Because BMS-986122 does not bind to the orthosteric site and has no inherent agonist action at endogenously expressed levels of MOR, it produces a reduced level of morphine-like side effects of constipation, reward as measured by conditioned place preference, and respiratory depression. These data provide a rationale for the further exploration of the action and safety of mu-PAMs as an innovative approach to pain management.
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