Specific caspase interactions and amplification are involved in selective neuronal vulnerability in Huntington's disease

Huntington's disease (HD) is an autosomal dominant progressive neurodegenerative disorder resulting in selective neuronal loss and dysfunction in the striatum and cortex. The molecular pathways leading to the selectivity of neuronal cell death in HD are poorly understood. Proteolytic processing of full-length mutant huntingtin (Htt) and subsequent events may play an important role in the selective neuronal cell death found in this disease. Despite the identification of Htt as a substrate for caspases, it is not known which caspase(s) cleaves Htt in vivo or whether regional expression of caspases contribute to selective neuronal cells loss. Here, we evaluate whether specific caspases are involved in cell death induced by mutant Htt and if this correlates with our recent finding that Htt is cleaved in vivo at the caspase consensus site 552. We find that caspase-2 cleaves Htt selectively at amino acid 552. Further, Htt recruits caspase-2 into an apoptosome-like complex. Binding of caspase-2 to Htt is polyglutamine repeat-length dependent, and therefore may serve as a critical initiation step in HD cell death. This hypothesis is supported by the requirement of caspase-2 for the death of mouse primary striatal cells derived from HD transgenic mice expressing full-length Htt (YAC72). Expression of catalytically inactive (dominant-negative) forms of caspase-2, caspase-7, and to some extent caspase-6, reduced the cell death of YAC72 primary striatal cells, while the catalytically inactive forms of caspase-3, -8, and -9 did not. Histological analysis of post-mortem human brain tissue and YAC72 mice revealed activation of caspases and enhanced caspase-2 immunoreactivity in medium spiny neurons of the striatum and the cortical projection neurons when compared to controls. Further, upregulation of caspase-2 correlates directly with decreased levels of brain-derived neurotrophic factor in the cortex and striatum of 3-month YAC72 transgenic mice and therefore suggests that these changes are early events in HD pathogenesis. These data support the involvement of caspase-2 in the selective neuronal cell death associated with HD in the striatum and cortex.     

Resistances to fluid flow of model xylem vessels with simple and scalariform perforation plates

The resistances of xylem vessel walls and perforation plates has been investigated using 18 large-scale physical models made of plastic tubing into which scale models of plates were inserted. Flow of water through vessels was modelled using glycerol instead of water to keep the Reynolds number below 0.1. The technique proved easy, cheap and reliable.Results showed that perforation plate resistance is low compared with the resistance of the walls, whatever the plate morphology; plates only provided 0.6-18.6% extra resistance. Simple plates provided less resistance than scalariform plates, but because they are arranged closer together in vessels, resistance values (1.7-5.1%) overlap with those of scalariform plates. The resistance of scalariform plates varied in a systematic way with their morphology. For a given plate angle, increasing the number of bars increased resistance. For a given bar number, increasing the angle of the plate to the vessel axis also increased the resistance. However, for a given gap between bars, increasing the angle of the plate to the vessel axis decreased resistance. These results are discussed in the light of theories about the function of perforation plates.Keywords: Flow, xylem vessel, perforation plate, models.      

Proteolysis of Mutant Huntingtin Produces an Exon 1 Fragment That Accumulates as an Aggregated Protein in Neuronal Nuclei in Huntington Disease

Huntingtin proteolysis has been implicated in the molecular pathogenesis of Huntington disease (HD). Despite an intense effort, the identity of the pathogenic smallest N-terminal fragment has not been determined. Using a panel of anti-huntingtin antibodies, we employed an unbiased approach to generate proteolytic cleavage maps of mutant and wild-type huntingtin in the HdhQ150 knock-in mouse model of HD. We identified 14 prominent N-terminal fragments, which, in addition to the full-length protein, can be readily detected in cytoplasmic but not nuclear fractions. These fragments were detected at all ages and are not a consequence of the pathogenic process. We demonstrated that the smallest fragment is an exon 1 huntingtin protein, known to contain a potent nuclear export signal. Prior to the onset of behavioral phenotypes, the exon 1 protein, and possibly other small fragments, accumulate in neuronal nuclei in the form of a detergent insoluble complex, visualized as diffuse granular nuclear staining in tissue sections. This methodology can be used to validate the inhibition of specific proteases as therapeutic targets for HD by pharmacological or genetic approaches.     

Serotonin as an integrator of leech behavior and muscle mechanical performance

The obliquely striated muscle in the leech body wall has a broad functional repertoire; it provides power for both locomotion and suction feeding. It also operates over an unusually high strain range, undergoing up to threefold changes in length. Serotonin (5-HT) may support this functional flexibility, integrating behavior and biomechanics. It can act centrally, promoting motor outputs that drive body wall movements, and peripherally, modulating the mechanical properties of body wall muscle. During isometric contractions 5-HT enhances active force production and reduces resting muscle tone. We therefore hypothesized that 5-HT would increase net work output during the cyclical contractions associated with locomotion and feeding. Longitudinal strains measured during swimming, crawling and feeding were applied to body wall muscle in vitro with the timing and duration of stimulation selected to maximize net work output. The net work output during all simulated behaviors significantly increased in the presence of 100μM 5-HT relative to the 5-HT-free control condition. Without 5-HT the muscle strips could not achieve a net positive work output during simulated swimming. The decrease in passive tension associated with 5-HT may also be important in reducing muscle antagonist work during longitudinal muscle lengthening. The behavioral and mechanical effects of 5-HT during locomotion are clearly complementary, promoting particular behaviors and enhancing muscle performance during those behaviors. Although 5-HT can enhance muscle mechanical performance during simulated feeding, low in vivo activity in serotonergic neurons during feeding may mean that its mechanical role during this behavior is less important than during locomotion.     

Are we ready for therapeutic drug monitoring of biologic therapeutics?

In the previous issue of Arthritis Research & Therapy, Ducourau and colleagues report that they retrospectively detected anti-infliximab antibodies in 21% of patients with rheumatic diseases. Patients with anti-infliximab antibodies had lower serum drug concentrations. These findings contribute to the existing evidence of immunogenicity of biologicals and its clinical relevance. We argue for therapeutic drug monitoring to optimize treatment response.     

Coupling endoplasmic reticulum stress to the cell death program: role of the ER chaperone GRP78

Alterations in Ca(2+) homeostasis and accumulation of unfolded proteins in the endoplasmic reticulum (ER) lead to an ER stress response. Prolonged ER stress may lead to cell death. Glucose-regulated protein (GRP) 78 (Bip) is an ER lumen protein whose expression is induced during ER stress. GRP78 is involved in polypeptide translocation across the ER membrane, and also acts as an apoptotic regulator by protecting the host cell against ER stress-induced cell death, although the mechanism by which GRP78 exerts its cytoprotective effect is not understood. The present study was carried out to determine whether one of the mechanisms of cell death inhibition by GRP78 involves inhibition of caspase activation. Our studies indicate that treatment of cells with ER stress inducers causes GRP78 to redistribute from the ER lumen with subpopulations existing in the cytosol and as an ER transmembrane protein. GRP78 inhibits cytochrome c-mediated caspase activation in a cell-free system, and expression of GRP78 blocks both caspase activation and caspase-mediated cell death. GRP78 forms a complex with caspase-7 and -12 and prevents release of caspase-12 from the ER. Addition of (d)ATP dissociates this complex and may facilitate movement of caspase-12 into the cytoplasm to set in motion the cytosolic component of the ER stress-induced apoptotic cascade. These results define a novel protective role for GRP78 in preventing ER stress-induced cell death.     

Hybrid origin of Japanese mice "Mus musculus molossinus": evidence from restriction analysis of mitochondrial DNA

The Japanese mouse, Mus musculus molossinus, has long been considered an independent subspecies of the house mouse. A survey of restriction- site haplotypes of mitochondrial DNA (mtDNA) showed that Japanese mice have two main maternal lineages. The most common haplotype is closely related to the mtDNA of the European subspecies M. m. musculus. The other common haplotype and two minor ones are closely related to each other and to the mtDNA of an Asiatic subspecies, M. m. castaneus. Two other rare variants are probably the result of recent contamination by European M. m. domesticus. The musculus type of mtDNA is found in the southern two-thirds of Japan, whereas the common castaneus type is found in the northern third and the minor variants are found sporadically throughout Japan. The castaneus mtDNA lineage had a few minor variants, whereas the musculus lineage was completely monomorphic. By contrast, the native population of M. m. castaneus and the Chinese and Korean musculus populations were highly polymorphic. These results suggest that M. m. molossinus is a hybrid between ancestral colonies, possibly very small, of M. m. musculus and M. m. castaneus, rather than an independent subspecies.      

A new approach to quantifying morphological variation in bluegill Lepomis macrochirus

Bluegill Lepomis macrochirus showed intraspecific morphological and behavioural differences dependent on the environment. Pelagic L. macrochirus had more fusiform bodies, a higher pectoral fin aspect ratio, a larger spiny dorsal fin area and pectoral fins located farther from the centre of mass than littoral L. macrochirus (P < 0·05). The shape of the body and pectoral fins, in particular, were suggestive of adaptation for sustained high-speed and economical labriform swimming. Littoral L. macrochirus had a deeper and wider body, deeper caudal fins and wider mouths than pelagic L. macrochirus (P < 0·05). Additionally, the soft dorsal, pelvic, anal and caudal fins of littoral L. macrochirus were positioned farther from the centre of mass (P < 0·05). The size and placement of these fins suggested that they will be effective in creating turning moments to facilitate manoeuvring in the macrophyte-dense littoral habitat.