Deciphering effect of complex organics on Anammox-sulfide autotrophic denitrification coupling system for landfill leachate treatment.

Landfill leachate, a high-strength liquid waste, is collected at landfills and then trucked to and treated at wastewater treatment facilities, where it is one of the largest contributors of nitrogen in the wastewater influent. This process costs each landfill hundreds of thousands to millions of dollars annually, mostly due to the high ammonia content. Nutrients released in effluent from wastewater treatment facilities have raised concerns over the resulting ecological and economic costs that eutrophication has on downstream water quality as well as commercial fishery and recreational uses. This issue may be addressed by combining leachate treatment, specifically nutrient removal, with commercial algae growth. The pairing of these systems has potential cost savings to biomass production, wastewater treatment, and solid waste disposal industries. The feasibility of such a remediation system was evaluated based on the biomass growth and removal of nitrogen from raw landfill leachate by a mixed algae-bacteria culture. This system was designed to treat leachate with minimal inputs and maintenance requirements, and was operated as an open semi-batch reactor in an urban greenhouse. The results, presented in Chapter 3, revealed that the average biomass growth rate was 5.6 ± 6.4 mg biomass/L/day and reached a maximum biomass density of 480 mg biomass/L. This maximum biomass density is comparable to the algae biomass industry goal of 500mg biomass/L. The maximum nitrogen removal rate measured from this system was 9.18 mgN/L/day. The ammonia removal rates of this culture increased with increasing initial ammonia concentration; maximum nitrogen removal occurred at an ammonia concentration of 80 mg N-NH3/L. At starting ammonia concentrations above 80 mg N-NH3/L a reduction in nitrogen removal was seen; this inhibition is hypothesized to be caused by ammonia toxicity. This inhibiting concentration is considerably higher than that of many other published studies. To determine if these productivity rates are feasible when the scale of the system is increased, a comparison of productivity at three significantly different scales was completed. The methodology of this comparison study, described in Chapter 4, used ammonia removal, nitrogen removal and algal growth rates as metrics of productivity. The treatment system evaluated utilized an algae-bacteria consortium to remediate raw landfill leachate and was monitored continuously for 52 weeks. The result of this comparison-of-scale study, presented in Chapter 5, determined that the rates observed in small-scale reactors (0.25L flasks) were significantly different from the rates observed in mid- (100L aquarium tanks) and large- (1000L raceway ponds) scale reactors. These results are of great significance since many predictive analyses, such as life cycle assessments (LCAs) and techno economic analyses (TEAs), use results from small scale studies as input values for large-scale predictions. This suggests that the results from small-scale experiments may not be appropriate as input data in predictive analyses of full scale algal processes. Throughout all of these studies it was observed that the amount of nitrogen removed from the liquid portion of the systems did not equal that amount of nitrogen in the biomass grown, calculated by the Redfield ratio. An evaluation of the potential nitrogen transformation pathways present in the yearlong study of the algae based leachate treatment systems, from Chapter 5, is presented in Chapter 6. The results of this evaluation concluded that while the nitrogen removal pathway of bio-assimilation was present, it was not the only nitrogen removal pathway in the treatment system. During a major part of the study (83.4% of the weeks), all nitrogen could not be accounted for in the dissolved inorganic nitrogen or biomass portions of the system. It is hypothesized that some of the unaccounted for nitrogen was lost due to volatilization of gaseous nitrogen species. Scenarios for the occurrence other nitrogen transformation pathways have been postulated based upon characteristic distribution of measured dissolved inorganic nitrogen and the nitrogen content of the biomass for each week and prior metagenomic analysis of the microbial community in the treatment system.

Effective partial denitrification of biological effluent of landfill leachate for Anammox process: Start-up, influencing factors and stable operation

The inheritance of the short SLC6A4 allele, encoding the serotonin transporter (SERT) in humans, increases susceptibility to neuropsychiatric and metabolic disorders, with aging and female sex further exacerbating these conditions. Both central and peripheral mechanisms of the compromised serotonin (5-HT) system play crucial roles in this context. Previous studies on SERT-deficient (Sert-/-) mice, which model human SERT deficiency, have demonstrated emotional and metabolic disturbances, exacerbated by exposure to a high-fat Western diet (WD). Growing evidence suggests the significance of hepatic regulatory mechanisms in the neurobiology of central nervous system disorders, supporting the 'liver-brain' concept. However, the relationship between aberrant behavior and hepatic alterations under conditions of SERT deficiency remains poorly investigated. One-year-old female Sert-/- mice and their wild-type (WT) littermates were subjected to a control diet (CD) or the WD for a duration of three weeks. The WD had a higher caloric content and was characterized by an elevated saturated fat content (21%) compared to the CD (4.5%) and contained 0.2% cholesterol. Mice were evaluated for anxiety-like behavior, exploration and locomotor activity in the open field test, as well as glucose tolerance and histological indicators of hepatic steatosis. Hepatic pro-inflammatory and metabolism-related gene expression and markers of nitrosative stress, were analyzed utilizing real-time polymerase chain reaction (RT-PCR) and correlated with behavioral and histological outcomes. In comparison to unchallenged mice, Sert-/-/WD mutants, but not the WT/WD group, had increased locomotion and anxiety-like behavior, increased hepatic steatosis, and elevated expression of insulin receptor B and pro-inflammatory cytokines interleukin-1β (Il-1β) and Tnf, as well as decreased expression of leptin receptor B. The two genotypes displayed distinct gene expression patterns of nitric oxide (NO)-related molecules inducible NO synthase (iNos) and arginase (Arg2), insulin receptor-related signaling factors: cluster of differentiation 36 (Cd36), ecto-nucleotide pyrophosphatase/phosphodiesterase (Enpp), protein tyrosine phosphatase N1 (Ptpn1), cytochrome P450 omega-hydroxylase 4A14 (Cyp4a14), acyl-CoA synthetase 1 (Acsl1) and phosphatase and tensin homolog (Pten). Furthermore, there were profound differences in correlations between molecular, histological, and behavioral measurements across the two genotypes. Our findings suggest that the genetic deficiency of SERT results in abnormal hepatic pro-inflammatory and metabolic adaptations in response to WD. The significant correlations observed between behavioral measures and pro-inflammatory and metabolic alterations in WD-fed mice suggest the importance of liver-brain interactions and their role in the aberrant behaviors exhibited by Sert-/- mutants. This study presents the first evidence that altered liver functions are associated with pathological behaviors arising from genetic SERT deficiency.

Organisms adapt to changing environments by adjusting their development, metabolism, and behavior to improve their chances of survival and reproduction. To achieve such flexibility, organisms must be able to sense and respond to changes in external environmental conditions and their internal state. Metabolic adaptation in response to altered nutrient availability is key to maintaining energy homeostasis and sustaining developmental growth. Furthermore, environmental variables exert major influences on growth and final adult body size in animals. This developmental plasticity depends on adaptive responses to internal state and external cues that are essential for developmental processes. Genetic studies have shown that the fruit fly Drosophila, similarly to mammals, regulates its metabolism, growth, and behavior in response to the environment through several key hormones including insulin, peptides with glucagon-like function, and steroid hormones. Here we review emerging evidence showing that various environmental cues and internal conditions are sensed in different organs that, via inter-organ communication, relay information to neuroendocrine centers that control insulin and steroid signaling. This review focuses on endocrine regulation of development, metabolism, and behavior in Drosophila, highlighting recent advances in the role of the neuroendocrine system as a signaling hub that integrates environmental inputs and drives adaptive responses.

Potential and limits of landfill leachate treatment in a multi-stage subsurface flow constructed wetland – Evaluation of organics and nitrogen removal

Landfill leachates are complex wastewater which has high pollution and their biological degradability is also difficult. Landfill leachate with high concentrations of contaminants must be pretreated. For this reason, the ammonium stripping process is an effective method for ammonium removal, especially for landfill leachate. With the ammonium stripping process, the ammonium and organic matter removal increases. This study aimed to investigate the applicability of the ammonium stripping process as remove ammonium (NH4+) and chemical oxygen demand (COD) from raw leachate. A new system has been designed which is called ammonium stripping tower. During this study, the ammonium stripping process was operated temperature (30-40-60oC), aeration rate (HH, HL, LL m3air/min), and hydraulic retention times (6-12-24-48 h). System performance was evaluated by ammonium, and COD parameters. The optimum conditions of the ammonium stripping process were determined at 600C temperature, HH(1m3air/min aeration rate), and hydraulic retention time (48h). The corresponding ammonium and COD removal efficiencies were about 88% and 79% respectively. The results of this study suggest that the use of an ammonium stripping process is an effective way to remove ammonium and COD concentration from raw landfill leachate.

Metabolic Control Analysis (MCA) marked a turning point in understanding the design principles of metabolic network control by establishing control coefficients as a means to quantify the degree of control that an enzyme exerts on flux or metabolite concentrations. MCA has demonstrated that control of metabolic pathways is distributed among many enzymes rather than depending on a single rate-limiting step. MCA also proved that this distribution depends not only on the stoichiometric structure of the network but also on other kinetic determinants, such as the degree of saturation of the enzyme active site, the distance to thermodynamic equilibrium, and metabolite feedback regulatory loops. Consequently, predicting the alterations that occur during metabolic adaptation in response to strong changes involving a redistribution in such control distribution can be challenging. Here, using the framework provided by MCA, we illustrate how control distribution in a metabolic pathway/network depends on enzyme kinetic determinants and to what extent the redistribution of control affects our predictions on candidate enzymes suitable as targets for small molecule inhibition in the drug discovery process. Our results uncover that kinetic determinants can lead to unexpected control distribution and outcomes that cannot be predicted solely from stoichiometric determinants. We also unveil that the inference of key enzyme-drivers of an observed metabolic adaptation can be dramatically improved using mean control coefficients and ruling out those enzyme activities that are associated with low control coefficients. As the use of constraint-based stoichiometric genome-scale metabolic models (GSMMs) becomes increasingly prevalent for identifying genes/enzymes that could be potential drug targets, we anticipate that incorporating kinetic determinants and ruling out enzymes with low control coefficients into GSMM workflows will facilitate more accurate predictions and reveal novel therapeutic targets.

Advanced nitrogen removal from mature landfill leachate based on novel step-draining partial nitrification-denitrification and Anammox process: Significance of low volume exchange ratio

Endurance training causes an increased activity of oxidative enzymes in skeletal muscle tissue (Holloszy and Booth, 1976; Bylund et al., 1977). A similar adaptation has also been described as a spontaneous phenomenon in patients with peripheral arterial occlusive disease (Holm et al., 1972; Bylund et al., 1976), who have an insufficient blood supply to the muscles even at mild exercise.

Weight loss is known to reduce resting energy expenditure (REE) and non-resting energy expenditure (NREE). Although seen in subjects with normal weight and excess weight, this decrease is variable and independent of the weight loss strategy. Adaptive thermogenesis (AT) refers to fat-free mass (FFM)-independent decreases in energy expenditure (1). AT spares energy, explains the nonlinear weight loss with time, and is related to weight loss maintenance. Weight loss-associated changes in composition of FFM, i.e., in skeletal muscle and organ masses, explained one third of AT (2). With a considerable interindividual variance, AT is considered as an individual trait that is biologically controlled. Low sympathetic nervous system (SNS) activity and decreases in leptin and T3 levels are its proposed determinants. This idea is supported by associations between AT and changes in its endocrine determinants after weight loss and during weight maintenance. However, preventing weight loss-associated decreases in serum T3 concentrations or in SNS activity did not affect REE (2). By contrast, substituting leptin in subjects who were weight reduced and weight stable normalized NREE but was without effect on REE (3). In this issue of Obesity, Rosenbaum and Leibel (4) proposed three models for AT. First, a “mechanical model” related to the “settling” of body weight where weight loss has no effect on slope and intercept of the REE-FFM regression line; second, a “threshold model” where AT is related to the decrease in fat mass below a minimum; and third, a so-called “spring loading model” with an effect of the dynamics of weight loss (4). No model fully explained AT. However, decreases in REE were consistent with the threshold model (with a maximal effect of fat mass on REE at 10% weight loss), whereas NREE declined similarly at each weight loss consistent with the third model. The calculations were performed using data obtained in patients with obesity during maintenance at different weights by controlled diet and physical activity. The major caveat of this protocol is that it is not about regulation of AT but more on control of long-term metabolic adaptations in response to reduced body weight. Since data obtained in a weight-reduced/weight-stable status reflect adaptations to reduced body weight, they do not explain regulation of AT. In fact, recent data showed that AT occurred early, within 3 days of semi-starvation, and is independent of changes in body weight and fat mass (2). AT fully established within 1 week of caloric restriction with no further changes during the following weeks (Figure 1). Thus, REE is regulated during early starvation. Although similar in magnitude, this may differ from long-term adaptations and weight maintenance. Surprisingly, neither leptin nor T3 nor SNS activity further explained AT (2). By contrast, decreases in insulin secretion and fluid retention correlated with AT, suggesting a very early threshold model unrelated to the adipocyte. Changes in resting energy expenditure (REE) and REE adjusted for fat-free mass during a 3-week period of controlled caloric restriction (at 50% of individual energy needs) and physical activity (at about 5,000 steps per day) in 32 young men who were healthy and with normal weight. Data were given separately for each week (i.e., they were calculated as differences between the last and the first day of the respective week). REE decreased during week 1. When compared with week 1, there were no further decreases in REE during weeks 2 and 3. The early decrease in REE corresponds to the nonlinear fall in body weight. The data were taken as evidence that regulation of adaptive thermogenesis occurs early, i.e., during week 1 of caloric restriction. With prolonged starvation, adaptation to reduced body weight rather than regulation of energy metabolism became more and more evident. *P ≤ 0.05; ***P ≤ 0.001. Data are from Müller et al., Am J Clin Nutr 2015;102:807-819 (see Ref. 2). Currently there is some confusion about defining AT in relation to either weight loss or weight loss maintenance (1). We need to differentiate between (i) mass-independent sparing of energy with negative energy balance and (ii) metabolic and endocrine factors related to maintenance of reduced weight. There is first evidence that the two issues are differently explained. Putting the adipocyte at the center of body weight control was a big step forward; this was also a good story. However, as far as the regulation of AT is concerned, this idea has some limitations, i.e., up to now, there is no evidence of an effect of leptin on REE in humans. By contrast, the issue of AT brings us back to a classical and “pre-adipocentric” view on endocrine and metabolic adaptation to starvation (5). From now on, taking both points of view into account will be wise.

Discovery of Heat-Tolerant Ovarian Cancer Cells and Metabolic Adaptations in Response to Heat Stress

Endotoxin triggers a reorganization of the energy metabolic pathway, including the promotion of fatty acid utilization to adapt to a high energy demand during endotoxemia. However, the factors responsible for the metabolic adaptation and characteristic pathologies resulting from defective utilization fatty acids during endotoxin response have not been fully clarified. The thioredoxin binding protein-2 (TBP-2) knockout (TBP-2) mouse is an animal model of fatty acid oxidation disorder. The aim of this study was to determine whether and how TBP-2 is involved in metabolic regulation in a lipopolysaccharide (LPS)-induced endotoxemia model in mice. Prospective animal trial. Research laboratory. TBP-2 and wild control mice. TBP-2 and wild control mice were intraperitoneally injected with LPS. Mortality, serum levels of markers of hepatorenal injuries, cytokines, insulin, glucose and lipid derivatives, and the hepatic signaling pathway regulating gluconeogenesis were investigated. Following the administration of LPS, TBP-2 mice showed a predisposition for death without any significant elevation of inflammatory cytokines, compared to the wild mice. LPS-challenged TBP-2 mice showed fat deposition in the liver and kidney, organ injuries, glycogen depletion, and elevation of serum lipid derivatives such as free fatty acids, triglyceride and cholesterol. Hyperinsulinemia and hypoglycemia were observed in TBP-2 mice after LPS injection. Death due to the LPS administration was prevented by supplementation of glucose. Phosphorylation of Akt and FoxO1, an inhibitory pathway of gluconeogenesis in the liver of LPS-challenged TBP-2 mice was demonstrated, suggesting the enhancement of insulin signaling. TBP-2 is involved in metabolic control during LPS-induced endotoxemia. After the LPS challenge, TBP-2 mice showed several characteristic aspects, such as hepatorenal injuries, and dysregulation of the lipid and glucose metabolisms. Furthermore, hypoglycemia promoted by hyperinsulinemia may be a critical risk factor for mortality in circumstances in which fatty acid utilization is impaired during endotoxemia.

Neuroblastoma is the most common childhood extracranial solid tumor. In high-risk cases, many of which are characterized by amplification of MYCN, outcome remains poor. Mutations in the p53 (TP53) tumor suppressor are rare at diagnosis, but evidence suggests that p53 function is often impaired in relapsed, treatment-resistant disease. To address the role of p53 loss of function in the development and pathogenesis of high-risk neuroblastoma, we generated a MYCN-driven genetically engineered mouse model in which the tamoxifen-inducible p53ER(TAM) fusion protein was expressed from a knock-in allele (Th-MYCN/Trp53(KI)). We observed no significant differences in tumor-free survival between Th-MYCN mice heterozygous for Trp53(KI) (n = 188) and Th-MYCN mice with wild-type p53 (n = 101). Conversely, the survival of Th-MYCN/Trp53(KI/KI) mice lacking functional p53 (n = 60) was greatly reduced. We found that Th-MYCN/Trp53(KI/KI) tumors were resistant to ionizing radiation (IR), as expected. However, restoration of functional p53ER(TAM) reinstated sensitivity to IR in only 50% of Th-MYCN/Trp53(KI/KI) tumors, indicating the acquisition of additional resistance mechanisms. Gene expression and metabolic analyses indicated that the principal acquired mechanism of resistance to IR in the absence of functional p53 was metabolic adaptation in response to chronic oxidative stress. Tumors exhibited increased antioxidant metabolites and upregulation of glutathione S-transferase pathway genes, including Gstp1 and Gstz1, which are associated with poor outcome in human neuroblastoma. Accordingly, glutathione depletion by buthionine sulfoximine together with restoration of p53 activity resensitized tumors to IR. Our findings highlight the complex pathways operating in relapsed neuroblastomas and the need for combination therapies that target the diverse resistance mechanisms at play. Cancer Res; 76(10); 3025-35. ©2016 AACR.

Lung adenocarcinoma (LUAD) is a deadly and heterogeneous subtype of non-small cell lung cancer. During LUAD progression, the emergence of alternate epithelial lineage fates in primary tumors correlates with poor outcome. The underlying mechanisms and biological consequences of these phenomena are poorly understood. Using an integrated approach, we previously examined the molecular relationship between cell differentiation states, lung cancer subtypes, and clinical outcome, to discover a novel role for several lineage-restricted transcription factors in the pathogenesis of LUAD (1). Specifically, we identified the alveolar specification factor, HOPX, as an inhibitor of metastatic progression. Herein we show that down-regulation of its function in a subset of human tumors correlates with DNA hypermethylation, aberrant tumor differentiation, and relapses. HOPX restricts the metastatic potential of LUAD cells, by modulating converging transcriptional programs of airway epithelial differentiation, malignant invasion, and metabolic adaptation in response to the extracellular microenvironment. Mechanistically, HOPX mediates its functions in part through the regulation of previously uncharacterized long-intergenic long non-coding RNAs with potential roles in lung development. Our findings demonstrate that perturbation of intrinsic cell lineage pathways is a determinant of metastasis in specific lung cancers. The implications of these findings for the origin(s) and treatment of LUAD metastasis will be further discussed. (1) Cheung, W.K.C., Zhao, M., Liu, Z., Stevens, L.E., Cao, P.D., Fang, J.E., Westbrook, T.F., and Nguyen, D.X. (2013). Cancer Cell, 23 (6): 725-738. Citation Format: Don X. Nguyen. Converging molecular effectors of airway lineage specification and metastasis in lung cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2332. doi:10.1158/1538-7445.AM2015-2332

[EN] In order to study the metabolic adaptation in response to 48 h transient doe-litter separation (DLS) in young rabbits (5 rabbits/d group) between postnatal 9 and 11 d, plasma concentrations of thyroid hormones T3 and T4, insulin, leptin, glucose, triglycerides (TG), and free fatty acids (FFA) were examined before (6-8 d), during (9-11 d), and after separation (12-16 d). T3 concentrations in newborn control rabbits gradually increased from 0.6 ng/mL at postnatal 6 d to 1.0 ng/mL at postnatal 16 d, whereas those of T4 remained fairly constant (25 ng/mL) up to postnatal 14 d, when T4 gradually declined to 8 ng/mL. T3 values of DLS newborn rabbits did not differ from those of controls at postnatal 10 and 11 d, but were lower (P<0.05) at postnatal 12 d, while T4 concentrations in DLS animals increased, although not significantly, between postnatal 10 and 12 d compared to controls. Insulin concentrations in young control rabbits ranged between 0.6 and 1.0 mg/L in the early postnatal days, whereas those for leptin averaged 2-3 ng/mL. Insulin and leptin values in DLS newborn rabbits were lower (P<0.05) at postnatal 10 and 11 d, but thereafter rebounded to levels close to those of controls. Glycaemia showed a comparable trend in both groups, ranging between 170 and 190 mg/dL up to postnatal 14 d, but thereafter decreased (P<0.05) to values of 120-130 mg/dL independently of treatment. Concentrations of TG varied greatly from day to day around a mean value of 300 mg/dL, whereas those of FFA remained at approximately the same steady-state levels from postnatal 6 to 16 d, averaging 0.8 mM without any significant differences between groups. In conclusion, these findings confirm that newborn rabbits can cope with the metabolic stress of starvation associated with DLS by lowering insulin and leptin concentrations while maintaining those of thyroid hormones, an overall endocrine response which, together with temporary increase of glucorticoids, successfully maintains an adequate energy balance.