Long‐term temporal and spatial trends in eutrophication status of the Baltic Sea

Much of the Baltic Sea is currently classified as ‘affected by eutrophication’. The causes for this are twofold. First, current levels of nutrient inputs (nitrogen and phosphorus) from human activities exceed the natural processing capacity with an accumulation of nutrients in the Baltic Sea over the last 50–100 years. Secondly, the Baltic Sea is naturally susceptible to nutrient enrichment due to a combination of long retention times and stratification restricting ventilation of deep waters. Here, based on a unique data set collated from research activities and long‐term monitoring programs, we report on the temporal and spatial trends of eutrophication status for the open Baltic Sea over a 112‐year period using the HELCOM Eutrophication Assessment Tool (HEAT 3.0). Further, we analyse variation in the confidence of the eutrophication status assessment based on a systematic quantitative approach using coefficients of variation in the observations. The classifications in our assessment indicate that the first signs of eutrophication emerged in the mid‐1950s and the central parts of the Baltic Sea changed from being unaffected by eutrophication to being affected. We document improvements in eutrophication status that are direct consequences of long‐term efforts to reduce the inputs of nutrients. The reductions in both nitrogen and phosphorus loads have led to large‐scale alleviation of eutrophication and to a healthier Baltic Sea. Reduced confidence in our assessment is seen more recently due to reductions in the scope of monitoring programs. Our study sets a baseline for implementation of the ecosystem‐based management strategies and policies currently in place including the EU Marine Strategy Framework Directives and the HELCOM Baltic Sea Action Plan.     

Nod-like Receptor Protein 3 (NLRP3) Inflammasome Activation and Podocyte Injury via Thioredoxin-Interacting Protein (TXNIP) during Hyperhomocysteinemia

NADPH oxidase-derived reactive oxygen species (ROS) have been reported to activate NLRP3 inflammasomes resulting in podocyte and glomerular injury during hyperhomocysteinemia (hHcys). However, the mechanism by which the inflammasome senses ROS is still unknown in podocytes upon hHcys stimulation. The current study explored whether thioredoxin-interacting protein (TXNIP), an endogenous inhibitor of the antioxidant thioredoxin and ROS sensor, mediates hHcys-induced NLRP3 inflammasome activation and consequent glomerular injury. In cultured podocytes, size exclusion chromatography and confocal microscopy showed that inhibition of TXNIP by siRNA or verapamil prevented Hcys-induced TXNIP protein recruitment to form NLRP3 inflammasomes and abolished Hcys-induced increases in caspase-1 activity and IL-1β production. TXNIP inhibition protected podocytes from injury as shown by normal expression levels of podocyte markers, podocin and desmin. In vivo, adult C57BL/6J male mice were fed a folate-free diet for 4 weeks to induce hHcys, and TXNIP was inhibited by verapamil (1 mg/ml in drinking water) or by local microbubble-ultrasound TXNIP shRNA transfection. Evidenced by immunofluorescence and co-immunoprecipitation studies, glomerular inflammasome formation and TXNIP binding to NLRP3 were markedly increased in mice with hHcys but not in TXNIP shRNA-transfected mice or those receiving verapamil. Furthermore, TXNIP inhibition significantly reduced caspase-1 activity and IL-1β production in glomeruli of mice with hHcys. Correspondingly, TXNIP shRNA transfection and verapamil attenuated hHcys-induced proteinuria, albuminuria, glomerular damage, and podocyte injury. In conclusion, our results demonstrate that TXNIP binding to NLRP3 is a key signaling mechanism necessary for hHcys-induced NLRP3 inflammasome formation and activation and subsequent glomerular injury.     

Evaluation of a novel methacrylate‐based protein a resin for the purification of immunoglobulins and Fc‐fusion proteins

Protein A affinity chromatography is a central part of most commercial monoclonal antibody and Fc‐fusion protein purification processes. In the last couple years an increasing number of new Protein A technologies have emerged. One of these new Protein A technologies consists of a novel, alkaline‐tolerant, Protein A ligand coupled to a macroporous polymethacrylate base matrix that has been optimized for immunoglobulin (Ig) G capture. The resin is interesting from a technology perspective because the particle size and pore distribution of the base beads are reported to have been optimized for high IgG binding and fast mass transfer, while the Protein A ligand has been engineered for enhanced alkaline tolerance. This resin was subjected to a number of technical studies including evaluating dynamic and static binding capacities, alkaline stability, Protein A leachate propensity, impurity clearance, and pressure–flow behavior. The results demonstrated similar static binding capacities as those achieved with industry standard agarose Protein A resins, but marginally lower dynamic binding capacities. Removal of impurities from the process stream, particularly host cell proteins, was molecule dependent, but in most instances matched the performance of the agarose resins. This resin was stable in 0.1 M NaOH for at least 100 h with little loss in binding capacity, with Protein A ligand leakage levels comparable to values for the agarose resins. Pressure–flow experiments in lab‐scale chromatography columns demonstrated minimal resin compression at typical manufacturing flow rates. Prediction of resin compression in manufacturing scale columns did not suggest any pressure limitations upon scale up. © 2014 American Institute of Chemical Engineers Biotechnol. Prog., 30:1125–1136, 2014     

Mechanism of intramolecular recyclization and deletion formation following transformation of Escherichia coli with linearized plasmid DNA.

The deletion end-points of a number of type I (less than monomeric) plasmid deletants obtained by transforming recA+ or recA- E. coli with linear pBR322 DNA were determined by DNA sequencing. In both monodirectional and bidirectional deletions the recyclization point was normally characterized by recombination between directly repeated sequences of between 4 and 10 bp present on each arm of the linearized pBR322 molecule. Frequently, short tracts of uninterrupted homology involved in recombinational recircularization were embedded in regions of relative non-homology. A model predicting the probability of matching sequences in either end of a linear plasmid molecule is presented. It is proposed that exonucleolytic processing of the exposed termini of linear plasmid molecules generates substrates for subsequent recombinational recyclization and deletion. The activity of host recombination and repair functions in recircularizing linear DNA molecules explains the generation of many of the aberrant recombinant DNA constructs obtained during gene cloning procedures.     

Antibody-mediated in vitro neutralization of human immunodeficiency virus type 1 abolishes infectivity for chimpanzees.

This study was undertaken to establish whether antibody directed against the human immunodeficiency virus type 1 (HIV-1) principal gp120 type-specific neutralization determinant can abolish the infectivity of HIV-1 in chimpanzees. Challenge inocula of the IIIb virus isolate were mixed in vitro with either immunoglobulin G (IgG) from an uninfected chimpanzee, nonneutralizing IgG from an HIV-seropositive human, a virus-neutralizing murine monoclonal antibody directed against the HIV-1 IIIb isolate, or virus-neutralizing IgG from a chimpanzee infected with the IIIb isolate. Both neutralizing antibodies were directed against the principal neutralization determinant of the challenge isolate. Establishment of infection following inoculation of each virus-antibody mixture into chimpanzees was assessed by virus-specific antibody development and by virus isolation. No protective effect was noted either with the control IgG or with the nonneutralizing anti-HIV IgG. By contrast, the polyclonal chimpanzee virus-neutralizing IgG prevented HIV-1 in vivo infection, while the neutralizing monoclonal antibody notably decreased the infectivity of the challenge virus. Hence, antibody to the gp120 principal neutralization determinant is able both to prevent HIV-1 infection in vitro and to inhibit infection in vivo.     

A Pilot Study of Demographic and Dopaminergic Genetic Contributions to Weight Change in Kidney Transplant Recipients

Kidney transplant recipients often experience a significant amount of weight gain in the first year post-transplantation. While demographic factors such as age, race, and sex have been associated with weight gain in this population, these factors do not explain all of the variability seen. A number of studies have suggested that genetics also plays a critical role in weight changes. Recently, alterations in the activity of the neurotransmitter dopamine have been associated with weight change, and gene expression studies in kidney transplant recipients have supported this association. The purpose of this pilot study is to first examine the feasibility and methodology, and then to examine the associations of age, race, sex, and genotype for 13 SNPs and 3 VNTRs in 9 dopaminergic pathway genes (ANKK1, DRD2, DRD3, DRD4, SLC6A3/DAT1, MAOA, MAOB, COMT, CPE) for associations with percent weight change at 12 months post-transplantation. Seventy kidney transplant recipients had demographic and clinical data collected as a part of a larger observational study. DNA was extracted from repository buffy coat samples taken at the time of transplant, and genotyped using Taqman and PCR based methods. Three SNPs were independently associated with percent weight change: ANKK1 rs1800497 (r = -0.28, p = 0.05), SLC6A3/DAT1 rs6347 (p = 0.046), and CPE rs1946816 (p = 0.028). Stepwise regression modelling confirmed the combined associations of age (p = 0.0021), DRD4 VNTR 4/5 genotype (p = 0.0074), and SLC6A3/DAT1 rs6347 CC genotype (p = 0.0009) and TT genotype (p = 0.0004) with percent weight change in a smaller sample (n = 35) of these kidney transplant recipients that had complete genotyping. These associations indicate that there may be a genetic, and an age component to weight changes post transplantation.     

Effect of acute head-down tilt on skeletal muscle cross-sectional area and proton transverse relaxation time

Conley, Michael S., Jeanne M. Foley, Lori L. Ploutz-Snyder,Ronald A. Meyer, and Gary A. Dudley. Effect of acute head-down tilt on skeletal muscle cross-sectional area and proton transverse relaxation time. J. Appl. Physiol.81(4): 1572-1577, 1996.This study investigated changes inskeletal muscle cross-sectional area (CSA) evoked by fluid shifts thataccompany short-term 6° head-down tilt (HDT) or horizontal bedrest, the time course of the resolution of these changes afterresumption of upright posture, and the effect of altered muscle CSA, inthe absence of increased contractile activity, on proton transverserelaxation time (T₂). Averagemuscle CSA and T₂ were determinedby standard spin-echo magnetic resonance imaging. Analyses wereperformed on contiguous transaxial images of the neck and calf. After aday of normal activity, 24 h of HDT increased neck muscle CSA 19 ± 4 (SE)% (P < 0.05) whilecalf muscle CSA decreased 14 ± 3%(P < 0.05). The horizontal posture(12 h) induced about one-half of these responses: an 11 ± 2%(P < 0.05) increase in neck muscleCSA and an 8 ± 2% decrease (P < 0.05) in the calf. Within 2 h after resumption of upright posture, neckand calf muscle CSA returned to within 0.5% (P > 0.05) of the values assessedafter a day of normal activity, with most of the change occurringwithin the first 30 min. No further change in muscle CSA was observedthrough 6 h of upright posture. Despite these large alterations inmuscle CSA, T₂ was not altered bymore than 1.1 ± 0.6% (P > 0.05)and did not relate to muscle size. These results suggest that posturalmanipulations and subsequent fluid shifts modeling microgravity elicitmarked changes in muscle size. Because these responses were notassociated with alterations in muscleT₂, it does not appear that simple movement of water into muscle can explain the contrast shift observed after exercise.

     

Specificity of resistance training responses in neck muscle size and strength

This study examined hypertrophy after head extension resistance training to assess which muscles of the complicated cervical neuromuscular system were used in this activity. We also determined if conventional resistance exercises, which are likely to evoke isometric action of the neck, induce generalized hypertrophy of the cervical muscle. Twenty-two active college students were studied. [mean (SE) age, weight and height: 21 (1) years, 71 (4) kg and 173 (3) cm, respectively]. Subjects were assigned to one of three groups: RESX (head extension exercise and other resistance exercises), RES (resistance exercises without specific neck exercise), or CON (no training). Groups RESX (n = 8) and RES (n = 6) trained 3 days/week for 12 weeks with large-muscle mass exercises (squat, deadlift, push press, bent row and mid-thigh pull). Group RESX also performed three sets of ten repetitions of a head extension exercise 3 days/week with a load equal to the 3 × 10 repetition maximum (RM). Group CON (n = 8) was a control group. The cross-sectional area (CSA) of nine individual muscles or muscle groups was determined by magnetic resonance imaging (MRI) of the cervical region. The CSA data were averaged over four contiguous transaxial slices in which all muscles of interest were visible. The 3 × 10 RM for the head extension exercise increased for RESX after training [from 17.9 (1.0) to 23.9 (1.4) kg, P < 0.05] but not for RES [from 17.6 (1.4) to 17.7 (1.9)␣kg] or CON [from 10.1 (2.2) to 10.3 (2.1) kg]. RESX showed an increase in total neck muscle CSA after training [from 19.5 (3.0) to 22.0 (3.6) cm², P < 0.05], but RES and CON did not [from 19.6 (2.9) to 19.7 (2.9)␣cm² and 17.0 (2.5) to 17.0 (2.4) cm², respectively]. This hypertrophy for RESX was due mainly to increases in CSA of 23.9 (3.2), 24.0 (5.8), and 24.9 (5.3)% for the splenius capitis, and semispinalis capitis and cervicis muscles, respectively. The lack of generalized neck muscle hypertrophy in RES was not due to insufficient training. For example, the CSA of their quadriceps femoris muscle group, as assessed by MRI, increased by 7 (1)% after this short-term training (P < 0.05). The results suggest that: (1) the splenius capitis, and semispinalis capitis and cervicis muscles are mainly responsible for head extension; (2) short-term resistance training does not provide a sufficient stimulus to evoke neck muscle hypertrophy unless specific neck exercises are performed; and (3) the postural role of head extensors provides modest loading in bipeds. Accepted: 15 October 1996