Industrial processes with animal cells

Industrial processes involving animal cells for the production of useful products still seem to be rather uncommon. Nevertheless, during the last four decades of the last century the number of relevant processes has increased from production of virus vaccines to monoclonal antibodies and finally complex structured glycoproteins. As soon as cell lines became permanent and culture medium changed from purely biological fluids to more or less defined chemical media, large-scale cultivation could begin. The developments of the 1970s - fusion of cells to form hybridomas, and genetic engineering - triggered a second wave of products. Monoclonal antibodies and recombinant proteins for diagnosis and therapy set new challenges for the inventors. Historically, there has been no straightforward process development since the product dictates the process operation. Therefore, the scale of production covers the whole range from small multiple-unit reactors (flasks or roller bottles) up to 10,000-l single-unit batch reactors. Products with high value and small demand can be produced in multiple-unit systems whereas "bulk" products for vaccination and therapy may need large-scale bioreactors to be cost effective. All the different systems have their advantages and disadvantages and significant challenges that curb the development of effective perfusion cultures still remain.     

Identification of mutants of Arabidopsis defective in acclimation of photosynthesis to the light environment

In common with many other higher plant species, Arabidopsis undergoes photosynthetic acclimation, altering the composition of the photosynthetic apparatus in response to fluctuations in its growth environment. The changes in photosynthetic function that result from acclimation can be detected in a noninvasive manner by monitoring chlorophyll (Chl) fluorescence. This technique has been used to develop a screen that enables the rapid identification of plants defective at ACCLIMATION OF PHOTOSYNTHESIS TO THE ENVIRONMENT (APE) loci. The application of this screen to a population of T-DNA-transformed Arabidopsis has successfully led to the identification of a number of mutant lines with altered Chl fluorescence characteristics. Analysis of photosynthesis and pigment composition in leaves from three such mutants showed that they had altered acclimation responses to the growth light environment, each having a distinct acclimation-defective phenotype, demonstrating that screening for mutants using Chl fluorescence is a viable strategy for the investigation of acclimation. Sequencing of the genomic DNA flanking the T-DNA elements showed that in the ape1 mutant, a gene was disrupted that encodes a protein of unknown function but that appears to be specific to photosynthetic organisms, whereas the ape2 mutant carries an insertion in the region of the TPT gene encoding the chloroplast inner envelope triose phosphate/phosphate translocator.     

Emotional Processing,Recognition, Empathy and Evoked Facial Expression in Eating Disorders: An Experimental Study to Map Deficits in Social Cognition

Background

Difficulties in social cognition have been identified in eating disorders (EDs), but the exact profile of these abnormalities is unclear. The aim of this study is to examine distinct processes of social-cognition in this patient group, including attentional processing and recognition, empathic reaction and evoked facial expression in response to discrete vignettes of others displaying positive (i.e. happiness) or negative (i.e. sadness and anger) emotions.

Method

One hundred and thirty-eight female participants were included in the study: 73 healthy controls (HCs) and 65 individuals with an ED (49 with Anorexia Nervosa and 16 with Bulimia Nervosa). Self-report and behavioural measures were used.

Results

Participants with EDs did not display specific abnormalities in emotional processing, recognition and empathic response to others’ basic discrete emotions. However, they had poorer facial expressivity and a tendency to turn away from emotional displays.

Conclusion

Treatments focusing on the development of non-verbal emotional communication skills might be of benefit for patients with EDs.     

Captive Reptile Mortality Rates in the Home and Implications for the Wildlife Trade

The trade in wildlife and keeping of exotic pets is subject to varying levels of national and international regulation and is a topic often attracting controversy. Reptiles are popular exotic pets and comprise a substantial component of the live animal trade. High mortality of traded animals raises welfare concerns, and also has implications for conservation if collection from the wild is required to meet demand. Mortality of reptiles can occur at any stage of the trade chain from collector to consumer. However, there is limited information on mortality rates of reptiles across trade chains, particularly amongst final consumers in the home. We investigated mortality rates of reptiles amongst consumers using a specialised technique for asking sensitive questions, additive Randomised Response Technique (aRRT), as well as direct questioning (DQ). Overall, 3.6% of snakes, chelonians and lizards died within one year of acquisition. Boas and pythons had the lowest reported mortality rates of 1.9% and chameleons had the highest at 28.2%. More than 97% of snakes, 87% of lizards and 69% of chelonians acquired by respondents over five years were reported to be captive bred and results suggest that mortality rates may be lowest for captive bred individuals. Estimates of mortality from aRRT and DQ did not differ significantly which is in line with our findings that respondents did not find questions about reptile mortality to be sensitive. This research suggests that captive reptile mortality in the home is rather low, and identifies those taxa where further effort could be made to reduce mortality rates.     

How Different Are Our Perceptions of Equal-Tempered and Microtonal Intervals? A Behavioural and EEG Survey

For listeners familiar with Western twelve-tone equal-tempered (12-TET) music, a novel microtonal tuning system is expected to present additional processing challenges. We aimed to determine whether this was the case, focusing on the extent to which our perceptions can be considered bottom-up (psychoacoustic and primarily perceptual) and top-down (dependent on familiarity and cognitive processing). We elicited both overt response ratings, and covert event-related potentials (ERPs), so as to compare subjective impressions of sounds with the neurophysiological processing of the acoustic signal. We hypothesised that microtonal intervals are perceived differently from 12-TET intervals, and that the responses of musicians (n = 10) and non-musicians (n = 10) are distinct. Two-note chords were presented comprising 12-TET intervals (consonant and dissonant) or microtonal (quarter tone) intervals, and ERP, subjective roughness ratings, and liking ratings were recorded successively. Musical experience mediated the perception of differences between dissonant and microtone intervals, with non-musicians giving similar ratings for each, and musicians preferring dissonant over the less commonly used microtonal intervals, rating them as less rough. ERP response amplitude was greater for consonant intervals than other intervals. Musical experience interacted with interval type, suggesting that musical expertise facilitates the sensory and perceptual discrimination of microtonal intervals from 12-TET intervals, and an increased ability to categorize such intervals. Non-musicians appear to have perceived microtonal intervals as instances of neighbouring 12-TET intervals.     

Diminished NADPH transhydrogenase activity and mitochondrial redox regulation in human failing myocardium

Although the functional role of nicotinamide nucleotide transhydrogenase (Nnt) remains to be fully elucidated, there is strong evidence that Nnt plays a critical part in mitochondrial metabolism by maintaining a high NADPH-dependant GSH/GSSG ratio, and thus the control of cellular oxidative stress. Using real-time PCR, spectrophotometric and western blotting techniques, we sought to determine the presence, abundance and activity level of Nnt in human heart tissues and to discern whether these are altered in chronic severe heart failure. Left ventricular levels of the NNT gene and protein expression did not differ significantly between the non-failing donor (NF) and heart failure (HF) group. Notably, compared to NF, Nnt activity rates in the HF group were 18% lower, which coincided with significantly higher levels of oxidized glutathione, lower glutathione reductase activity, lower NADPH and a lower GSH/GSSG ratio. In the failing human heart a partial loss of Nnt activity adversely impacts NADPH-dependent enzymes and the capacity to maintain membrane potential, thus contributing to a decline in bioenergetic capacity, redox regulation and antioxidant defense, exacerbating oxidative damage to cellular proteins.     

Bioluminescent organs of two deep-sea arrow worms, Eukrohnia fowleri and Caecosagitta macrocephala, with further observations on Bioluminescence in chaetognaths

Bioluminescence in the deep-sea chaetognath Eukrohnia fowleri is reported for the first time, and behavioral, morphological, and chemical characteristics of bioluminescence in chaetognaths are examined. Until this study, the only known species of bioluminescent chaetognath was Caecosagitta macrocephala. The luminescent organ of that species is located on the ventral edge of each anterior lateral fin, whereas that of E. fowleri runs across the center of the tail fin on both dorsal and ventral sides. Scanning electron microscopy showed that the bioluminescent organs of both species consist of hexagonal chambers containing elongate ovoid particles-the organelles holding bioluminescent materials. No other luminous organism is known to use hexagonal packing to hold bioluminescent materials. Transmission electron microscopy of particles from C. macrocephala revealed a densely packed paracrystalline matrix punctuated by globular inclusions, which likely correspond to luciferin and luciferase, respectively. Both species use unique luciferases in conjunction with coelenterazine for light emission. Luciferase of C. macrocephala becomes inactive after 30 min, but luciferase of E. fowleri is highly stable. Although C. macrocephala has about 90 times fewer particles than E. fowleri, it has a similar bioluminescent capacity (total particle volume) due to its larger particle size. In situ observations of C. macrocephala from a remotely operated vehicle revealed that the luminous particles are released to form a cloud. The discovery of bioluminescence in a second chaetognath phylogenetically distant from the first highlights the importance of bioluminescence among deep-sea organisms.     

Siderophore-mediated cooperation and virulence in Pseudomonas aeruginosa

Why should organisms cooperate with each other? Helping close relatives that are likely to share the same genes (kin selection) is one important explanation that is likely to apply across taxa. The production of metabolically costly extracellular iron-scavenging molecules (siderophores) by microorganisms is a cooperative behaviour because it benefits nearby conspecifics. We review experiments focusing on the production of the primary siderophore (pyoverdin) of the opportunistic bacterial pathogen, Pseudomonas aeruginosa, which test kin selection theories that seek to explain the evolution of cooperation. First, cooperation is indeed favoured when individuals interact with their close relatives and when there is competition between groups of cooperators and noncooperators, such that the benefit of cooperation can be realized. Second, the relative success of cheats and cooperators is a function of their frequencies within populations. Third, elevated mutation rates can confer a selective disadvantage under conditions when cooperation is beneficial, because high mutation rates reduce how closely bacteria are related to each other. Fourth, cooperative pyoverdin production is also shown to be favoured by kin selection in vivo (caterpillars), and results in more virulent infections. Finally, we briefly outline ongoing and future work using this experimental system.     

The stability and complexity of antibody responses to the major surface antigen of Plasmodium falciparum are associated with age in a malaria endemic area

Individuals that are exposed to malaria eventually develop immunity to the disease with one possible mechanism being the gradual acquisition of antibodies to the range of parasite variant surface antigens in their local area. Major antibody targets include the large and highly polymorphic Plasmodium falciparum Erythrocyte Membrane Protein 1 (PfEMP1) family of proteins. Here, we use a protein microarray containing 123 recombinant PfEMP1-DBLα domains (VAR) from Papua New Guinea to seroprofile 38 nonimmune children (<4 years) and 29 hyperimmune adults (≥15 years) from the same local area. The overall magnitude, prevalence and breadth of antibody response to VAR was limited at <2 years and 2-2.9 years, peaked at 3-4 years and decreased for adults compared with the oldest children. An increasing proportion of individuals recognized large numbers of VAR proteins (>20) with age, consistent with the breadth of response stabilizing with age. In addition, the antibody response was limited in uninfected children compared with infected children but was similar in adults irrespective of infection status. Analysis of the variant-specific response confirmed that the antibody signature expands with age and infection. This also revealed that the antibody signatures of the youngest children overlapped substantially, suggesting that they are exposed to the same subset of PfEMP1 variants. VAR proteins were either seroprevalent from early in life, (<3 years), from later in childhood (≥3 years) or rarely recognized. Group 2 VAR proteins (Cys2/MFK-REY+) were serodominant in infants (<1-year-old) and all other sequence subgroups became more seroprevalent with age. The results confirm that the anti-PfEMP1-DBLα antibody responses increase in magnitude and prevalence with age and further demonstrate that they increase in stability and complexity. The protein microarray approach provides a unique platform to rapidly profile variant-specific antibodies to malaria and suggests novel insights into the acquisition of immunity to malaria.