Differential Effect of Culture Temperature and Specific Growth Rate on CHO Cell Behavior in Chemostat Culture

Mild hypothermia condition in mammalian cell culture technology has been one of the main focuses of research for the development of breeding strategies to maximize productivity of these production systems. Despite the large number of studies that show positive effects of mild hypothermia on specific productivity of r-proteins, no experimental approach has addressed the indirect effect of lower temperatures on specific cell growth rate, nor how this condition possibly affects less specific productivity of r-proteins. To separately analyze the effects of mild hypothermia and specific growth rate on CHO cell metabolism and recombinant human tissue plasminogen activator productivity as a model system, high dilution rate (0.017 h⁻¹) and low dilution rate (0.012 h⁻¹) at two cultivation temperatures (37 and 33°C) were evaluated using chemostat culture. The results showed a positive effect on the specific productivity of r-protein with decreasing specific growth rate at 33°C. Differential effect was achieved by mild hypothermia on the specific productivity of r-protein, contrary to the evidence reported in batch culture. Interestingly, reduction of metabolism could not be associated with a decrease in culture temperature, but rather with a decrease in specific growth rate.     

The functional effect of soybean extract and isolated isoflavone on myocardial infarction and ventricular dysfunction: The soybean extract on myocardial infarction

BackgroundMyocardial infarction is a public health problem. Functional food is an alternative treatment for cardiovascular diseases.ObjectiveThe objective was to analyze the functional and anatomopathological post-myocardial-infarction effects of soybean extract (SE) and isoflavone (IF).MethodsMyocardial infarction was induced in adult Wistar rats. After 5 days, an echocardiogram was performed to determine heart rate (HR), ejection fraction (EF), systolic volume (LVESV) and diastolic volume (LVEDV). Animals with ventricular dysfunction (EF<45%) were selected for study. The animals were divided into three groups: control (n=14), SE (n=15) and IF (n=12). The IF group received 120 mg/kg/day isolated IF, and the SE group received 12.52 g/day. After 30 days, a new echocardiogram was performed. A histological exam was carried out to determine the collagen. Activity of biochemical markers [arginase, lactate dehydrogenase (LDH) and malate dehydrogenase] was measured.ResultsThe animals of the control, IF and SE groups showed a reduction in EF after the infarction (P=.432, P=.017 and P=.320, respectively). An increase of LVESV and LVEDV was observed in all groups (P=.009, P=.001 and P=.140; and P=.003, P=.008 and P=.205, respectively). A reduction of HR was found in the SE group (P=.020). There was a greater activity of LDH in the SE group. A smaller quantity of mature collagen was found in the region proximal to the myocardial infarction in the SE group.ConclusionA protective effect in the SE group was observed 30 days after the myocardial infarction.     

Are kelp holdfasts islands on the ocean floor? – indication for temporarily closed aggregations of peracarid crustaceans

During the colonisation process of islands, newly immigrating species often arrive as single individuals. Islands that have received single colonisers may subsequently harbour large populations of a species, while other islands may completely lack this species. Exchange between islands is limited, thereby strongly affecting evolutionary processes. While this concept is widely used in the context of oceanic islands or habitat patches on the mainland, it is rarely used to explain and examine the distribution patterns of marine invertebrates. Benthic marine organisms inhabiting patches with island-like features may also be restricted in their movements between patches. Once established in a patch, it may be more favourable to remain there rather than moving to another patch. Juveniles of species with direct development may recruit to the island patch of their parents. Herein, we examined the peracarid fauna in patches that have island-like features, i.e. kelp holdfasts. The number of peracarid species within an individual holdfast increased with its size. Similarly, the number of individuals per holdfast increased with holdfast size. However, several peracarid species showed a strongly aggregated distribution pattern, being highly abundant in some holdfasts and almost completely absent in others. Our results suggest that these aggregations of conspecifics may be a consequence of the peracarid reproductive biology: fully developed juveniles emerge from the female's marsupium and recruit to the immediate vicinity of their mother, showing little or no tendency to emigrate towards other patches. At present, while it is not known how long peracarid aggregations within kelp holdfasts persist, our data suggest that some juveniles may remain with the natal holdfast and possibly reproduce therein. It is concluded that, during certain time periods, reproduction rates of peracarids in a holdfast may exceed their migration rates between holdfasts.     

Branch transpiration of pine and spruce scaled to tree and canopy using needle biomass distributions

Branch water exchange and total tree water uptake were measured in a mixed Norway spruce and Scots pine stand in central Sweden during the 1995 and 1996 growing seasons. Branch transpiration was scaled to canopy level on the basis of a branch conductance model, using vertical needle-area distributions obtained by destructive sampling. Comparison with total tree water uptake scaled to canopy level showed agreement within 10%, for periods when the canopy was not affected by climatically induced stress. Comparison of scaled fluxes on individual trees showed that measurements of transpiration at branch level provide information on the direct response of transpiration to variations in weather, and furthermore that the time-lag between transpiration and tree water uptake was as much as 3 h. The vertical needle-area distribution of Scots pine was similar to that found by other authors. Needle-area distribution on Norway spruce, which has not been described before, showed that it has its largest needle area at the top of the crown. Specific needle area varied considerably both within trees and between trees. For spruce, mean specific needle area (±SD) varied from 2.4±0.5 mm² mg^–1 at the top of the crown to 7.1±1.9 mm² mg^–1 at the base. Corresponding figures for Scots pine were 3.4±2.0 and 9.1±2.1. Received: 5 March 1999 / Accepted: 17 March 2000     

Evidence of natural Wolbachia infections and molecular identification of field populations of Culex pipiens complex (Diptera: Culicidae) mosquitoes in western Turkey

Establishing reliable risk projection information about the distribution pattern of members of the Culex pipiens complex is of particular interest, as these mosquitoes are competent vectors for certain disease‐causing pathogens. Wolbachia, a maternally inherited bacterial symbiont, are distributed in various arthropod species and can induce cytoplasmic incompatibility, i.e., reduced egg hatch, in certain crosses. It is being considered as a tool for population control of mosquito disease vectors. The Aegean region is characterized by highly populated, rural, and agricultural areas and is also on the route of the migratory birds. In this study, a fragment of the 658 bp of the mitochondrial cytochrome c oxidase subunit 1 (COI) gene, which includes the barcode region, was employed to differentiate Cx. pipiens complex species found in this region. Moreover, for the first time, the prevalence of Wolbachia endobacteria in these natural populations was examined using PCR amplification of a specific wsp gene. Our results revealed a widespread (more than 90%, n=121) presence of the highly efficient West Nile virus vector Cx. quinquefasciatus in the region. We also found that Wolbachia infection is widespread; the average prevalence was 62% in populations throughout the region. This study provided valuable information about the composition of Cx. pipiens complex mosquitoes and the prevalence of Wolbachia infection in these populations in the Aegean region. This information will be helpful in tracking mosquito‐borne diseases and designing and implementing Wolbachia‐based control strategies in the region.     

Altered Thiol Chemistry in Human Amyotrophic Lateral Sclerosis-linked Mutants of Superoxide Dismutase 1

Neurodegenerative diseases share a common characteristic, the presence of intracellular or extracellular deposits of protein aggregates in nervous tissues. Amyotrophic Lateral Sclerosis (ALS) is a severe and fatal neurodegenerative disorder, which affects preferentially motoneurons. Changes in the redox state of superoxide dismutase 1 (SOD1) are associated with the onset and development of familial forms of ALS. In human SOD1 (hSOD1), a conserved disulfide bond and two free cysteine residues can engage in anomalous thiol/disulfide exchange resulting in non-native disulfides, a hallmark of ALS that is related to protein misfolding and aggregation. Because of the many competing reaction pathways, traditional bulk techniques fall short at quantifying individual thiol/disulfide exchange reactions. Here, we adapt recently developed single-bond chemistry techniques to study individual disulfide isomerization reactions in hSOD1. Mechanical unfolding of hSOD1 leads to the formation of a polypeptide loop held by the disulfide. This loop behaves as a molecular jump rope that brings reactive Cys-111 close to the disulfide. Using force-clamp spectroscopy, we monitor nucleophilic attack of Cys-111 at either sulfur of the disulfide and determine the selectivity of the reaction. Disease-causing mutations G93A and A4V show greatly altered reactivity patterns, which may contribute to the progression of familial ALS.     

Changes in public order after the opening of a medically supervised safer injecting facility for illicit injection drug users

Background

North America''s first medically supervised safer injecting facility for illicit injection drug users was opened in Vancouver on Sept. 22, 2003. Although similar facilities exist in a number of European cities and in Sydney, Australia, no standardized evaluations of their impact have been presented in the scientific literature.

Methods

Using a standardized prospective data collection protocol, we measured injection-related public order problems during the 6 weeks before and the 12 weeks after the opening of the safer injecting facility in Vancouver. We measured changes in the number of drug users injecting in public, publicly discarded syringes and injection-related litter. We used Poisson log-linear regression models to evaluate changes in these public order indicators while considering potential confounding variables such as police presence and rainfall.

Results

In stratified linear regression models, the 12-week period after the facility''s opening was independently associated with reductions in the number of drug users injecting in public (p < 0.001), publicly discarded syringes (p < 0.001) and injection-related litter (p < 0.001). The predicted mean daily number of drug users injecting in public was 4.3 (95% confidence interval [CI] 3.5–5.4) during the period before the facility''s opening and 2.4 (95% CI 1.9–3.0) after the opening; the corresponding predicted mean daily numbers of publicly discarded syringes were 11.5 (95% CI 10.0–13.2) and 5.4 (95% CI 4.7–6.2). Externally compiled statistics from the city of Vancouver on the number of syringes discarded in outdoor safe disposal boxes were consistent with our findings.

Interpretation

The opening of the safer injecting facility was independently associated with improvements in several measures of public order, including reduced public injection drug use and public syringe disposal.Many cities are experiencing epidemics of bloodborne diseases as a result of illicit injection drug use,^1,2,3 and drug overdoses have become a leading cause of death in many urban areas.^4,5,6 Public drug use also plagues many inner city neighbourhoods, and the unsafe disposal of syringes in these settings is a major community concern.^{7,8,9,10,11,12,13}In over 2 dozen European cities and, more recently, in Sydney, Australia, medically supervised safer injecting facilities, where injection drug users (IDUs) can inject previously obtained illicit drugs under the supervision of medical staff, have been established in an effort to reduce the community and public health impacts of illicit drug use.¹⁴ Inside these facilities IDUs are typically provided with sterile injecting equipment, emergency care in the event of overdose, as well as primary care services and referral to addiction treatment.^13,15 Although anecdotal reports have suggested that such sites may improve public order,¹² reduce the number of deaths from overdose¹⁶ and improve access to care,¹⁷ no standardized evaluations of their impact are available in the scientific literature.¹⁸On Sept. 22, 2003, health officials in Vancouver opened a government-sanctioned safer injecting facility as pilot project. The facility, the first in North America, is centrally located in Vancouver''s Downtown Eastside, which is the most impoverished urban neighbourhood in Canada and home to well-documented overdose and HIV epidemics among the estimated 5000 IDUs who reside there.^19,20 Federal approval for the 3-year project was granted on the condition that the health and social impacts of the facility be rigorously evaluated. Although evaluation of the facility''s impact on certain outcomes (e.g., HIV incidence) is ongoing and will take several years, it is now possible to examine the impacts of the site on public order. Therefore, we conducted this study to test the hypothesis that changes in improperly discarded syringes and public drug use would be observed after the opening of the safer injecting facility.