Twenty-first century brain banking: practical prerequisites and lessons from the past: the experience of New York Brain Bank,Taub Institute,Columbia University

Generally accepted methods for processing postmortem brains are lacking, despite the efforts of pioneers in the field, and the growing awareness of the importance of brain banking for investigating the pathogenesis of illnesses unique to humans. Standardizing methods requires compromises, institutional or departmental mindset promoting collaboration, and the willingness to share ideas, information, and samples. A sound balance between competition and institutional interests is needed to best fulfill the tasks entrusted to health care institutions. Thus, a potentially widely accepted protocol design involves tradeoffs. We successfully integrated brain banking within the operation of the department of pathology. We reached a consensus whereby a brain can be utilized for diagnosis, research, and teaching. Thus, routing brains away from residency programs is avoided. The best diagnostic categorization possible is being secured and the yield of samples for research maximized. Thorough technical details pertaining to the actual processing of brains donated for research were recently published. Briefly, one-half of each brain is immersed in formalin for performing the neuropathologic evaluation, which is combined with the teaching task. The contralateral half is extensively dissected at the fresh state to obtain samples ready for immediate disbursement once categorized diagnostically. The samples are tracked electronically, which is crucial. This important tracking system is described separately in this issue. This report focuses on key lessons learned over the past 25 years of brain banking including successful solutions to originally unforeseen problems.     

Acute lung inflammation and ventilator-induced lung injury caused by ATP via the P2Y receptors: an experimental study

Background

Many studies associated the main polyphenolic constituent of green tea, (-)-Epigallocatechin-3-gallate (EGCG), with inhibition of cancers, invasion and metastasis. To date, most of the studies have focused on the effect of EGCG on cell proliferation or death. Since cell migration is an important mechanism involved in tumor invasion, the aim of the present work was to target another approach of the therapeutic effect of EGCG, by investigating its effect on the cell migratory behavior.

Methods

The effect of EGCG (at concentrations lower than 10 μg/ml) on the migration speed of invasive cells was assessed by using 2D and 3D models of cell culture. We also studied the effects of EGCG on proteinases expression by RT-PCR analysis. By immunocytochemistry, we analyzed alterations of vimentin organization in presence of different concentrations of EGCG.

Results

We observed that EGCG had an inhibitory effect of cell migration in 2D and 3D cell culture models. EGCG also inhibited MMP-2 mRNA and protein expression and altered the intermediate filaments of vimentin.

Conclusion

Taken together, our results demonstrate that EGCG is able to inhibit the migration of bronchial tumor cells and could therefore be an attractive candidate to treat tumor invasion and cell migration.     

mrpA (all1838), a gene involved in alkali and Na sensitivity, may also have a role in energy metabolism in the cyanobacterium Anabaena sp. strain PCC 7120

Anabaena sp. PCC7120 contains a gene, mrpA (all1838), which forms part of a seven gene-cluster (all1843–all1837) with significant sequence similarity to bacterial operons that putatively code for a multicomponent cation/proton antiporter involved in alkaline pH adaptation and salt resistance. We previously showed that growth and photosynthesis were inhibited in a strain mutated in mrpA, denoted as PHB11, particularly at alkaline pH. Here, we show that respiration was also impaired in the mutant independently of the external pH. In addition, at high pH, less ATP and vegetative cell ferredoxin were present in PHB11, which also showed lower levels of ferredoxin-NADP⁺ oxidoreductase (FNR). Ferredoxin and FNR are involved in the generation of reductant NADPH in cyanobacteria. These results suggest an energetic role of mrpA (and perhaps of the whole mrp-gene cluster) in Anabaena sp. PCC 7120 that is further supported by the significant similarity of putative Anabaena Mrp proteins to membrane subunits of complex I.     

Metal bridges between the PhoQ sensor domain and the membrane regulate transmembrane signaling

Bacterial histidine kinases respond to environmental stimuli by transducing a signal from an extracytosolic sensor domain to a cytosolic catalytic domain. Among them, PhoQ promotes bacterial virulence and is tightly repressed by the divalent cations such as calcium and magnesium. We have determined the crystal structure of the PhoQ sensor domain from Salmonella typhimurium in the Ca2+-bound state, which reveals a highly negatively charged surface that is in close proximity to the inner membrane. This acidic surface binds at least three Ca2+, which mediate the PhoQ-membrane interaction. Mutagenesis analysis indicates that structural integrity at the membrane proximal region of the PhoQ sensor domain promotes metal-mediated repression. We propose that depletion or displacement of divalent cations leads to charge repulsion between PhoQ and the membrane, which initiates transmembrane signaling through a change in orientation between the PhoQ sensor domain and membrane. Therefore, both PhoQ and the membrane are required for extracytosolic sensing and transmembrane signaling.     

Long-term intake of egg white hydrolysate attenuates the development of hypertension in spontaneously hypertensive rats

This paper evaluates the effect of the long-term intake of a hydrolysate of egg white with pepsin (HEW), with a potent angiotensin converting enzyme inhibitory activity, on the development of hypertension of spontaneously hypertensive rats (SHR). After being weaned, male 3-week-old SHR were randomly divided into five groups that were given until the 20th week of life the following drinking fluids: (1) tap water, (2) non-treated egg white 1 g/kg/day, (3) captopril 100 mg/kg/day, (4) HEW 0.5 g/kg/day, and (5) HEW 1 g/kg/day. From the 20th to 25th week of life, animals from all groups were given tap water. Systolic blood pressure (SBP) and diastolic blood pressure (DBP) were measured weekly in the rats, from the 6th to 25th week of life, by the tail cuff method. Development of hypertension was attenuated in the groups treated with captopril and HEW (P<0.001 vs. the group that drunk tap water). At the 20th week of life, the arterial blood pressure values of the different groups of rats were: tap water (SBP = 219.5 +/- 5.7, DBP = 167 +/- 3.7), non-treated egg white (SBP = 206.4 +/- 1.43, DBP = 166.4 +/- 4.9), captopril (SBP = 131.7 +/- 2.74, DBP = 91.5 +/- 1.62), HEW 0.5 g/kg/day (SBP = 182.9 +/- 4.64, DBP = 127.5 +/- 2.1) and HEW 1 g/kg/day (SBP = 177.7 +/- 4.72, DBP = 120.1 +/- 2.4). SBP and DBP increased in the treated SHR when the corresponding antihypertensive treatment was removed. In spite of this, SBP remained lower in the SHR that had received captopril and HEW than in the SHR of the control groups (P<0.05). The present results suggest that HEW could be used as a functional food with antihypertensive activity.     

Ultrastructural analysis and identification of envelope proteins of "Candidatus Chloracidobacterium thermophilum" chlorosomes

Chlorosomes are sac-like, light-harvesting organelles that characteristically contain very large numbers of bacteriochlorophyll (BChl) c, d, or e molecules. These antenna structures occur in chlorophototrophs belonging to some members of the Chlorobi and Chloroflexi phyla and are also found in a recently discovered member of the phylum Acidobacteria, "Candidatus Chloracidobacterium thermophilum." "Ca. Chloracidobacterium thermophilum" is the first aerobic organism discovered to possess chlorosomes as light-harvesting antennae for phototrophic growth. Chlorosomes were isolated from "Ca. Chloracidobacterium thermophilum" and subjected to electron microscopic, spectroscopic, and biochemical analyses. The chlorosomes of "Ca. Chloracidobacterium thermophilum" had an average size of ～100 by 30 nm. Cryo-electron microscopy showed that the BChl c molecules formed folded or twisted, sheet-like structures with a lamellar spacing of ～2.3 nm. Unlike the BChls in the chlorosomes of the green sulfur bacterium Chlorobaculum tepidum, concentric cylindrical nanotubes were not observed. Chlorosomes of "Ca. Chloracidobacterium thermophilum" contained a homolog of CsmA, the BChl a-binding, baseplate protein; CsmV, a protein distantly related to CsmI, CsmJ, and CsmX of C. tepidum, which probably binds a single [2Fe-2S] cluster; and five unique polypeptides (CsmR, CsmS, CsmT, CsmU, and a type II NADH dehydrogenase homolog). Although "Ca. Chloracidobacterium thermophilum" is an aerobe, energy transfer among the BChls in these chlorosomes was very strongly quenched in the presence of oxygen (as measured by quenching of fluorescence emission). The combined analyses showed that the chlorosomes of "Ca. Chloracidobacterium thermophilum" possess a number of unique features but also share some properties with the chlorosomes found in anaerobic members of other phyla.     

Quantitative trait loci and underlying candidate genes controlling agronomical and fruit quality traits in octoploid strawberry <Emphasis Type="Italic">(Fragaria</Emphasis> × <Emphasis Type="Italic">ananassa</Emphasis>)

Breeding for fruit quality traits in strawberry (Fragaria × ananassa, 2n = 8x = 56) is complex due to the polygenic nature of these traits and the octoploid constitution of this species. In order to improve the efficiency of genotype selection, the identification of quantitative trait loci (QTL) and associated molecular markers will constitute a valuable tool for breeding programs. However, the implementation of these markers in breeding programs depends upon the complexity and stability of QTLs across different environments. In this work, the genetic control of 17 agronomical and fruit quality traits was investigated in strawberry using a F₁ population derived from an intraspecific cross between two contrasting selection lines, ‘232’ and ‘1392’. QTL analyses were performed over three successive years based on the separate parental linkage maps and a pseudo-testcross strategy. The integrated strawberry genetic map consists of 338 molecular markers covering 37 linkage groups, thus exceeding the 28 chromosomes. 33 QTLs were identified for 14 of the 17 studied traits and approximately 37% of them were stable over time. For each trait, 1–5 QTLs were identified with individual effects ranging between 9.2 and 30.5% of the phenotypic variation, indicating that all analysed traits are complex and quantitatively inherited. Many QTLs controlling correlated traits were co-located in homoeology group V, indicating linkage or pleiotropic effects of loci. Candidate genes for several QTLs controlling yield, anthocyanins, firmness and l-ascorbic acid are proposed based on both their co-localization and predicted function. We also report conserved QTLs among strawberry and other Rosaceae based on their syntenic location.     

Recognition and specificity determinants of the human cbx chromodomains

The eight mammalian Cbx proteins are chromodomain-containing proteins involved in regulation of heterochromatin, gene expression, and developmental programs. They are evolutionarily related to the Drosophila HP1 (dHP1) and Pc (dPc) proteins that are key components of chromatin-associated complexes capable of recognizing repressive marks such as trimethylated Lys-9 and Lys-27, respectively, on histone H3. However, the binding specificity and function of the human homologs, Cbx1-8, remain unclear. To this end we employed structural, biophysical, and mutagenic approaches to characterize the molecular determinants of sequence contextual methyllysine binding to human Cbx1-8 proteins. Although all three human HP1 homologs (Cbx1, -3, -5) replicate the structural and binding features of their dHP counterparts, the five Pc homologs (Cbx2, -4, -6, -7, -8) bind with lower affinity to H3K9me3 or H3K27me3 peptides and are unable to distinguish between these two marks. Additionally, peptide permutation arrays revealed a greater sequence tolerance within the Pc family and suggest alternative nonhistone sequences as potential binding targets for this class of chromodomains. Our structures explain the divergence of peptide binding selectivity in the Pc subfamily and highlight previously unrecognized features of the chromodomain that influence binding and specificity.