Evolving paradigms for repair of tissues by adult stem/progenitor cells (MSCs)

• ? Paradigm I: the haematopoietic niche
• ? Paradigm II: engraftment/differentiation
  • ‐ Early observations on engraftment and differentiation
  • ‐ Technical challenges in testing paradigm II
  • ‐ The impetus to test the paradigm II in clinical trials
  • ‐ Tests of the paradigm II with local administrations
  • ‐ Tests of paradigm II with systemic infusion
• ? Paradigm III: transient ‘quasi‐niches’
  • ‐ Unusual features of MSCs in culture
  • ‐ Cross‐talk with injured tissues
  • ‐ Modulation of inflammation in paradigm III
  • ‐ Modulation of apoptosis in paradigm III
  • ‐ Modulation of immune reactions
  • ‐ Paradigm III and the similarities to paradigm I
• ? Conclusions/perspectives
  • ‐ Why is administration of MSCs beneficial?
  • ‐ Better assays for the potency of MSCs?
  • ‐ Are MSCs pericytes?
  • ‐ Therapies with recombinant proteins?
  • ‐ Additional questions in developing therapies with MSCs

In this review, we focus on the adult stem/progenitor cells that were initially isolated from bone marrow and first referred to as colony forming units‐fibroblastic, then as marrow stromal cells and subsequently as either mesenchymal stem cells or multipotent mesenchymal stromal cells (MSCs). The current interest in MSCs and similar cells from other tissues is reflected in over 10,000 citations in PubMed at the time of this writing with 5 to 10 new publications per day. It is also reflected in over 100 registered clinical trials with MSCs or related cells ( http//www.clinicaltrials.gov ). As a guide to the vast literature, this review will attempt to summarize many of the publications in terms of three paradigms that have directed much of the work: an initial paradigm that the primary role of the cells was to form niches for haematopoietic stem cells (paradigm I); a second paradigm that the cells repaired tissues by engraftment and differentiation to replace injured cells (paradigm II); and the more recent paradigm that MSCs engage in cross‐talk with injured tissues and thereby generate microenvironments or ‘quasi‐niches’ that enhance the repair tissues (paradigm III).     

Gene therapy in Alzheimer’s disease – potential for disease modification

• ? Introduction
• ? Targets and ongoing research
  • ‐ NGF
    • ‐ Neurotrophic function of NGF
    • ‐ Levels of NGF in AD
    • ‐ Role of NGF in AD
    • ‐ NGF as a therapeutic agent
    • ‐ Development of NGF gene therapy
    • ‐ In vivo gene delivery of NGF
  • ‐ BDNF
    • ‐ Neurotrophic function of BDNF
    • ‐ BDNF levels in AD
    • ‐ BDNF function in AD
    • ? Towards BDNF gene therapy
  • ‐ Neprilysin
    • ‐ Role of neprilysin in AD
    • ‐ Neprilysin levels in AD
    • ‐ Gene delivery of neprilysin in AD animal models
• ? Potential gene therapy target candidates
  • ‐ APOE
  • ‐ ECE
  • ‐ Cathepsin B
  • ‐ Other Aβ degrading enzymes
• ? Down‐regulation of AD‐associated proteins by siRNA
  • ‐ BACE1
  • ‐ APP
• ? Concluding remarks

Alzheimer’s disease (AD) is the major cause of dementia in the elderly, leading to memory loss and cognitive decline. The mechanism underlying onset of the disease has not been fully elucidated. However, characteristic pathological manifestations include extracellular accumulation and aggregation of the amyloid β‐peptide (Aβ) into plaques and intracellular accumulation and aggregation of hyperphosphorylated tau, forming neurofibrillary tangles. Despite extensive research worldwide, no disease modifying treatment is yet available. In this review, we focus on gene therapy as a potential treatment for AD, and summarize recent work in the field, ranging from proof‐of‐concept studies in animal models to clinical trials. The multifactorial causes of AD offer a variety of possible targets for gene therapy, including two neurotrophic growth factors, nerve growth factor and brain‐derived neurotrophic factor, Aβ‐degrading enzymes, such as neprilysin, endothelin‐converting enzyme and cathepsin B, and AD associated apolipoprotein E. This review also discusses advantages and drawbacks of various rapidly developing virus‐mediated gene delivery techniques for gene therapy. Finally, approaches aiming at down‐regulating amyloid precursor protein (APP) and β‐site APP cleaving enzyme 1 levels by means of siRNA‐mediated knockdown are briefly summarized. Overall, the prospects appear hopeful that gene therapy has the potential to be a disease modifying treatment for AD.     

Virus diseases of lupins

• I. INTRODUCTION
• II. APHID-TRANSMITTED VIRUSES

• (a) Potyvirus group

• (i) Bean yellow mosaic virus
• (ii) Clover yellow vein virus
• (iii) Bean common mosaic virus
• (iv) Peanut mottle virus
• (v) Bidens mottle virus

• (b) Cucumovirus group

• (i) Cucumber mosaic virus
• (ii) Peanut stunt virus

• (c) Alfalfa mosaic virus
• (d) Fabavirus group: broad bean wilt virus
• (e) Pea enation mosaic virus
• (f) Luteovirus group: soybean dwarf virus
• (g) Rhabdovirus group: lettuce necrotic yellows virus

• III. THRIPS-TRANSMITTED VIRUSES
• Tomato spotted wilt virus

• IV. NEMATODE-TRANSMITTED VIRUSES

• (a) Tobravirus group: pea early browning virus
• (b) Nepovirus group: tomato black ring virus

• V. VIRUS-LIKE DISEASES

• (a) Lupin leaf curl ‘virus’
• (b) ‘Lupin witches’ broom disease

• VI. CONTROL VII. CONCLUSIONS
• VII. CONCLUSIONS

     

Island biogeography and landscape ecology of mammals inhabiting fragmented, temperate rain forests

• 1 We expanded the island biogeography paradigm to test whether mammalian communities of the heavily fragmented temperate rain forests of the Olympic Peninsula were influenced by local environmental conditions, biogeographic factors (fragment area and isolation) and characteristics of the surrounding landscape.
  • 2 We used live‐trapping, sign surveys and infra‐red triggered cameras to compare distributions of non‐volant mammals among fragments and between fragments and other principal landscape components (continuous old‐growth, riparian corridors, second‐growth forest and clearcuts).
    • 3 Of the 24 species of non‐volant mammals detected during our studies, 18 occurred in at least one fragment.
      • 4 Species richness of old‐growth mammals was not significantly correlated with fragment area or isolation, per se, but was significantly and positively correlated with the amount of old‐growth fragments and old second‐growth (41–159 years) in the surrounding landscape (r² = 0.95, P < 0.005).
        • 5 Distributions of three old‐growth dependent species [shrew‐mole (Neurotrichus gibbsii), Douglas squirrel (Tamiasciurus douglasii) and Trowbridge shew (Sorex trowbridgii)] were significantly associated with local environmental conditions within the fragment, with geographical isolation from continuous old‐growth and riparian corridors, and with the amount of old‐growth and old second growth in the adjacent matrix.

     

Somatostatin receptor biology in neuroendocrine and pituitary tumours: part 2 – clinical implications

• ? Introduction
• ? SSTR subtype tissue distribution and its relevance to tumour imaging and treatment
• ? Conclusions

     

On the role of dynamic atmosphere–vegetation interactions under increasing radiative forcing

• 1 A zero‐dimensional model of local atmosphere–vegetation interaction is presented. The model includes essentials of water related two‐way feedbacks, such as the influence of vegetation on evapotranspiration, and the impact of temperature and drought on biomass growth and mortality. The simple model serves as a framework for the preliminary investigation of vegetation related feedbacks under climate change scenarios.
  • 2 Model simulations for a mid‐latitude forest area for an increasing external forcing indicate a transient growth of biomass up to a critical forcing, where drought stress begins to dominate the response. Beyond, biomass decreases, reinforced by the reduced evapotranspiration of a diminished vegetation, leading to an additional temperature increase (biomass–evapotranspiration feedback).
    • 3 The implementation of an additional feedback loop based on the hypothesis that drought stress implies not only a reduction in above‐ground biomass, but also a net reduction in roots and therefore a reduction of the amount of water accessible to the plants for transpiration, leads to the occurrence of a second stable state in the atmosphere–vegetation system. In the bistable regime, a moderate perturbation can trigger an abrupt change of state.
      • 4 The present conceptual investigations underline the importance of a dynamic vegetation subsystem in transient climate change, and stress in particular the possible role of feedbacks related to root dynamics.

     

TOXIC EFFECTS OF OXYGEN ON CEREBRAL METABOLISM

Abstract— The effect of 100% O₂ at one atmosphere on carbohydrate metabolism in cell-free homogenates of rat brain was studied under different experimental conditions. The principal findings were the following:

• 1 Compared to 10% O₂-90% N₂, oxygen at one atmosphere inhibited metabolism of α-oxoglutarate and depressed the net synthesis of ATP. With glucose as substrate, accumulation of ATP was also markedly inhibited but substrate utilization was only slightly affected.

• 2 Glycolysis in brain was relatively resistant to oxygen toxicity, except in the presence of added Cu²⁺.

• 3 With α-oxoglutarate as the substrate, inhibition of the formation of ATP occurred earlier than inhibition of substrate utilization, indicating the particular sensitivity of oxidative phosphorylation to inactivation by oxygen in vitro.

• 4 Cu²⁺ and Fe²⁺ accentuated oxygen toxicity but appeared to act by different mechanisms. Co²⁺ exerted a protective effect.

• 5 The sulphydryl compounds, dithiothreitol and reduced glutathione, strongly diminished the toxic effect of oxygen.

     

Physical properties of the bovine encephalitogenic protein; molecular weight and conformation

Abstract— The highly basic encephalitogenic protein isolated from bovine spinal cord was studied by various physicochemical methods:

• 1 The molecular weight was determined by sedimentation equilibrium, by calculation from the data on sedimentation coefficients and intrinsic viscosities, by measurement of intrinsic viscosity in the presence of concentrated guanidine hydrochloride (according to the method of Tanford , Kawahara and Lapanie , 1967), and by exclusion chromatography on Sephadex G-100 columns. All values obtained were in good agreement and indicated a molecular weight of approximately 18,000–20,000.

• 2 Studies of sedimentation velocities in the presence and absence of 6 m -guanidine-HCl indicated that there was a significant difference in the values of sedimentation coefficients.

• 3 The same conditions were applied to the measurements of viscosity; the difference was small but significant. These findings and the magnitude of the intrinsic viscosity suggested that this protein was in a disordered configuration. From these data, it is concluded that the protein was apparently monodispersed, in the presence or absence of the denaturing agent. This protein behaved like a polyelectrolyte in neutral aqueous solution.

• 4 The measurements of optical rotatory dispersion also confirmed that this protein existed in a disordered configuration.

     

METABOLISM OF AMINO ACIDS IN INCUBATED SLICES OF MOUSE BRAIN1

• 1 Slices of mouse brain were incubated with [U-¹⁴C]alanine, valine, leucine, phenylalanine, proline, histidine, lysine, arginine or aspartic acid, and the extent of metabolism was estimated by analyses utilizing paper chromatography of the tissue extracts and with an amino acid analyser.

• 2 The metabolism of Ala and Asp was high; of Leu and Pro, moderate; and of Lys, Arg and Phe, low; the metabolism of Val and His was not significant. The time-course of metabolism in most cases showed varying rates, indicating heterogeneous metabolic compartments for the amino acids.

• 3 Production of CO₂ was high from Asp, moderate from Ala, and low from Leu; the other amino acids were not oxidized to CO₂ to any significant extent. A large portion of the metabolized label was trapped in the form of Glu or Asp.

• 4 Metabolism increased with increasing concentration of amino acid to some extent and was largely inhibited by omission of glucose, by anaerobic conditions, or by cyanide. Although these conditions also inhibit uptake, the time-course and extent of inhibition uptake and metabolism were different.

• 5 With Asp, Ala and Phe, metabolism was lowest in slices from pons-medulla; the brain area exhibiting the highest metabolism differed for each amino acid. The metabolism of Asp was lower in brain samples from newborn than in those from adults; the metabolism of Leu was higher in slices from newborn brain.

• 6 The results indicate that the majority of the amino acids can be metabolized in brain tissue and that the metabolic rates are influenced by a number of factors, among them the level of amino acids and the level of available energy.

     

THE COMPARTMENTATION OF GLUTAMATE AND ITS METABOLITES IN FRACTIONS OF NEURON CELL BODIES AND NEUROPIL; STUDIED BY INTRAVENTRICULAR INJECTION OF [U-14C]GLUTAMATE

Abstract—

• 1 The in vivo metabolism of glutamate in rat neuron cell bodies and neuropil was studied after intraventricular injection of (U-¹⁴C)glutamic acid followed by separation of the tissue into neuronal and neuropil fractions.

• 2 The losses of amino acid and of radioactivity during the fractionation were equivalent. Recoveries were: glutamate, 32; glutamine, 15; aspartate, 25; GABA, 41; alanine, 30 per cent. In the washed cell fractions glutamine was 45 per cent and alanine 132 per cent higher in the neuronal fraction, glutamate was 62, GABA 77 and aspartate 95 per cent of neuropil levels. This contrasted with results obtained previously for in vitro incorporation. Calculation from these results indicated that 28 per cent of the original cell suspension was neuronal, 72 per cent neuropil. In the final cell preparations, 29 per cent of the neuron cell bodies and 26 per cent of the neuropil were recovered.

• 3 Specific activity of glutamate in the neuronal fraction 15 min after injection was higher than in the original suspension, but had declined to 30 per cent of its initial value by 2 h. In the neuropil, specific activity of glutamate was below that of the cell suspension at 15 min, but at later times rose above it by up to 40 per cent.

• 4 Radioactivity was detected in aspartate and glutamine 15 min after injection and GABA by 60 min after injection. In the original cell suspension the specific activity of glutamine was higher than that of glutamate at all times (the Waelsch effect) but aspartate and GABA were lower than glutamate.

• 5 In the neuronal fraction the specific activity of glutamine was below that of glutamate at all times, indicating a precursor-product relationship. In the neuropil fraction, glutamine specific activity remained above glutamate for the first hour.

• 6 These results are discussed in relation to the interpretation of the Waelsch effect in terms of metabolic compartmentation.

     

Subcellular distribution of aromatic amino acid transaminases in rat brain

Abstract— The subcellular distributions of tyrosine transaminase, DOPA transaminase, tryptophan transaminase and 5-hydroxytryptophan (5-HTP) transaminase were studied in rat brain.

• 1 For all of these transaminases 60-81 per cent of the total activities were found in the crude mitochondrial fraction. Tyrosine transaminase was the most active enzyme.

• 2 Tyrosine transaminase and DOPA transaminase had very similar distributions in all fractions, but the distribution of tryptophan transaminase and 5-HTP transaminase differed in the microsomal (Mic) and synaptic vesicle (M₂) fractions. Only 5-HTP transaminase was highly concentrated in the M₂ fraction.

• 3 DOPA transaminase was inhibited by dopamine and 5-HT, but these compounds had no effect on 5-HTP transaminase. Both enzymes were completely inhibited by m-hydroxybenzoyloxyamine.

     

ILSI/HESI maternal toxicity workshop summary: maternal toxicity and its impact on study design and data interpretation

Workshops on maternal toxicity were held at the annual Society of Toxicology, Teratology Society, and European Teratology Society meetings in 2009. Speakers presented background information prior to a general discussion on this topic. The following recommendations/options are based on the outcome of the discussions at the workshops:

• 1. A comprehensive evaluation of all available data from general toxicity studies, range‐finding Developmental and Reproductive Toxicology (DART) studies, class effects, structure–activity relationships, exposure studies, etc. is essential for appropriate dose selection for definitive DART studies. The intent is to avoid marked maternal toxicity leading to mortality or decreased body weight gains of greater than 20% for prolonged periods.

• (a) Evaluate alternative endpoints for dose selection and data interpretation (e.g., target tissue effects and pharmacology) for biotherapeutics.
• (B) Evaluate additional maternal parameters based on effects and/or target organs observed in short‐term (e.g., 2‐ or 4‐week) general toxicity studies.

• 2. Evaluate all available data to determine a cause–effect relationship for developmental toxicity.

• (a) Conduct a pair‐feeding/pair‐watering study as a follow‐up.
• (b) Evaluate individual data demonstrating maternal toxicity in the mother with adverse embryo–fetal outcomes in the litter associated with the affected mother.
• (c) Conduct single‐dose studies at increasing doses as a complement to conventional embryo–fetal toxicity studies for certain classes of compounds that affect the hERG channel.

• 3. Support statements that embryo–fetal effects are caused by maternal toxicity and/or exaggerated pharmacology, especially for malformations.

• (a) Provide mechanistic or other supporting data.
• (b) Establish the relevance of the DART findings in animals for human exposures. Birth Defects Res (Part B) 92:36–51, 2010. © 2011 Wiley‐Liss, Inc.

     

Vegetation gradients in Atlantic Europe: the use of existing phytosociological data in preliminary investigations on the potential effects of climate change on British vegetation

• 1 This paper aims to demonstrate the use of available vegetation data from the phytosociological literature in preliminary analyses to generate hypotheses regarding vegetation and climate change.
  • 2 Data for over 3000 samples of calcareous grassland, mesotrophic grassland, heath and woodland vegetation were taken from the literature for a region in the west of Atlantic Europe and subjected to ordination by detrended correspondence analysis in order to identify the main gradients present.
    • 3 Climate data were obtained at a resolution of 0.5° from an existing database. The relationship between vegetation composition and climate was investigated by the correlation of the mean scores for the first two ordination axes for each 0.5° cell with the climate and location variables.
      • 4 The ordinations resulted in clear geographical gradients for calcareous grasslands, heaths and woodlands but not for mesotrophic grasslands. Significant correlations were shown between some of the vegetation gradients and the climate variables, with the strongest relationships occurring between the calcareous grassland gradients and July temperature, latitude and oceanicity. Some of the vegetation gradients were also inferred to reflect edaphic factors, management and vegetation history.
        • 5 Those gradients that were related to temperature were hypothesized to reflect the influence of a progressively warmer climate on species composition, providing a baseline for further studies on the influence of climate change on species composition.
        • 6 The validity of the literature data was assessed by the collection of an original set of field data for calcareous grasslands and the subsequent ordination of a dataset containing samples from both the literature and the field. The considerable overlap between the samples from the literature and the field suggest that literature data can be used, despite certain limitations. Such preliminary analyses, using readily available data, can thus achieve useful results, thereby saving lengthy and costly field visits.

     

EFFECT OF THYROID HORMONE ON THE BIOCHEMICAL MATURATION OF RAT BRAIN: CONVERSION OF GLUCOSE-CARBON INTO AMINO ACIDS

• 1 The rapid and extensive conversion of glucose-carbon into amino acids is an index of the final coordination of the mechanisms underlying energy metabolism in the adult brain. This phenomenon develops in the rat during a short period extending from 10 to about 19 days after birth. The underlying factors have been analysed.

• 2 The development of the pattern of distribution of glucose-carbon characteristic of the adult brain was markedly influenced by the thyroid state of the animals. The age-curve for the conversion of glucose-carbon into brain amino acids was displaced to the left after treatment with thyroid hormone (T₃) in infancy thus indicating an accelerated maturation. Conversely, neonatal thyroidectomy resulted in a significant retardation in the conversion of glucose-carbon into amino acids.

• 3 The specific radioactivity of glutamate increased five-fold in the brain of normal rats from the 10th to the 19th day of age. The values (as a percentage of those for littermate controls) were 220 in the case of the 10 day-old thyroid treated rats and about 30 for the 19 day-old thyroid deficient animals. At the age of 10 days neither treatment affected the concentration of glutamate which was also only slightly less than the control values in the brain of 19 day-old thyroid deficient animals (–17 per cent).

• 4 Specific pool(s) of glutamate associated with the formation of GABA can be demonstrated in the brain of 19 day-old rats after administration of [U-¹⁴C]glucose as a result of anoxia post mortem. These pools did not develop in the brain of 10 day-old animals. Neonatal thyroidectomy retarded the development of these glutamate pools.

• 5 Evidence is summarized which indicates that the development of the rapid conversion of glucose-carbon into amino acids reflects the enlargement, during maturation, of the metabolic compartments which are associated with neuronal processes.

     

CHANGES IN POLYSOMES OF THE DEVELOPING RAT BRAIN

Abstract— Rat brain polysomes were prepared from a deoxycholate-treated postmito-chondrial supernatant in the presence of 2% bentonite and 1 mg/ml of yeast RNA to prevent partial degradation during preparation.

• 1 The polysomal preparations had an absorption maximum at 260 mμ and an absorption minimum at 235 mμ. The ratio of absorption maximum to minimum and the RNA to protein ratio were 1·58 and 1·06 respectively in 6-day-old rat brain polysomes. The sedimentation patterns showed six distinct peaks with sedimentation coefficients of 235S, 185S, 173S, 135S, 100S and 80S, indicating that these preparations have the characteristics of pure heavy polysomes.

• 2 The rate of [¹⁴C]phenylalanine incorporation into brain polysomal protein was maximal at approximately 10 days of age and decreased thereafter. A similar progressive reduction with increasing age was found in the stimulation of phenylalanine incorporation by the addition of 60 μg/tube of polyuridylic acid. However, the incorporation of phenylalanine into young rat brain polysomes was usually greater even with the addition of polyuridylic acid than in the older animals.

• 3 The comparative studies on sucrose density gradient centrifugation of polysomes between young and adult rat brains showed a considerable decrease of heavy polysomes in the older animals.

• 4 The effect of various factors on the stability of brain polysomes from both ages has been studied. The rates of RNA, protein and acid-soluble phosphorus release from polysomes of the adult rat brains were usually greater in the presence of high salt concentration, ethylenediaminetetra-acetic acid and urea than those from the corresponding preparations of younger animals. On the basis of evidence obtained from the above results it suggested that the adult brain polysomes were more unstable than those of younger animals.

• 5 The amount of polysomal RNA linearly increased up to the first 20 days after birth and then levelled off. The ratio of G + C/A + U of polysomal RNA was less in the young rat brains, falling to 1·30 as compared to 1·50 in older animals. The differences were statistically significant at less than a 1% level of confidence.

• 6 Polysomal preparations also contained RNase, phosphomonoesterase, phospho-diesterase and 5′-nucleotidase activities which cannot be washed off. The specific activities of these enzymes were generally higher in young rat brains than those in the adult.