Implications of the initial mutations in membrane cofactor protein (MCP; CD46) leading to atypical hemolytic uremic syndrome

The hemolytic uremic syndrome is characterized by the triad of microangiopathic hemolytic anemia, thrombocytopenia and acute renal failure. There are two general types. One occurs in epidemic form and is diarrheal associated (D+HUS). It has a good prognosis. The second is a rare form known as atypical (aHUS), which may be familial or sporadic, and has a poor prognosis. aHUS is increasingly recognized to be a disease of defective complement regulation, particularly cofactor activity. Mutations in membrane cofactor protein (MCP; CD46) that predispose to the development of aHUS were first identified in 2003. MCP is a membrane-bound complement regulator that acts as a cofactor for the factor I-mediated cleavage of C3b and C4b deposited on host cells. More than 20 different mutations in MCP have now been identified in patients with aHUS. Many of these mutants have been functionally characterized and have helped to define the pathogenic mechanisms leading to aHUS development. Over 75% of the reported mutations cause a reduction in MCP expression, due to homozygous, compound heterozygous or heterozygous mutations. This deficiency of MCP leads to inadequate control of complement activation on endothelial cells after an initiating injury. The remaining MCP mutants are expressed, but demonstrate reduced ligand (C3b/C4b) binding capacity and cofactor activity of MCP. MCP mutations in aHUS demonstrate incomplete penetrance, indicating that additional genetic and environmental factors are required to manifest disease. MCP mutants as a cause of aHUS have a favorable clinical outcome in comparison to patients with factor H (CFH) or factor I (IF) mutations. In 90% of the renal transplants performed in patients with MCP-HUS, there has been no recurrence of the primary disease, whilst >50% of factor I or factor H deficient patients have had a prompt recurrence. This highlights the importance of defining and characterizing the underlying genetic defects in patients with aHUS.     

alpha- and epsilon-protein kinase C activity during smooth muscle cell apoptosis in response to gamma-radiation

The use of gamma-radiation in treatment of pelvic cancer is associated with injury of healthy surrounding tissues and disorders of intestinal motility; however, the cellular mechanisms involved are unclear. We tested the hypothesis that exposure of visceral smooth muscle cells (SMCs) to gamma-radiation induces apoptosis via activation of specific protein kinase C (PKC) isoforms. Cultured SMCs and slices from guinea pig ileum smooth muscle longitudinal layer (GPISMLL) were exposed to 10 to 50 Gy. Flow cytometry in gamma-radiated SMCs showed increased percentage of cells in the sub-G(0)/G(1) phase, a hallmark of apoptosis. gamma-Radiation-induced reduction in cell survival was partially but significantly alleviated with the PKC inhibitors. Sections of gamma-irradiated GPISMLL showed DNA fragmentation and apoptotic bodies analyzed by the terminal deoxynucleotidyl transferase dUTP nick-end labeling method, whereas the plasma and nuclear membranes were preserved. Confocal microscopy in gamma-radiated SMCs labeled with annexin V-fluorescein showed an increase in apoptotic cells and phosphatidylserine externalization. Contraction of GPISMLL strips in response to KCl and acetylcholine was reduced in tissues exposed to 30 and 50 Gy. gamma-Radiation of GPISMLL caused an increase in PKC activity in the particulate fraction, a decrease in the cytosolic fraction, and increased particulate/cytosolic PKC activity ratio. Western blot analysis revealed significant amounts of alpha- and epsilon-PKC in the cytosolic fraction of control GPISMLL. gamma-Radiation caused an increase in the amount of alpha- and epsilon-PKC in the particulate fraction and a decrease in the cytosolic fraction. Data suggest that gamma-radiation induces apoptosis, growth arrest, and contractile dysfunction in visceral SMCs of GPISMLL via activation and translocation of alpha- and epsilon-PKC isoforms.     

The link between dopamine function and apathy in cannabis users: an [18F]-DOPA PET imaging study

Rationale

Cannabis is the most widely used illicit drug in the world, and regular use has been associated with reduced motivation, i.e. apathy. Regular long-term cannabis use has been associated with reduced dopamine synthesis capacity. The mesolimbic dopaminergic system mediates the processing of incentive stimuli by modifying their motivational value, which in turn is modulated by endocannabinoid signalling. Thus, it has been proposed that dopaminergic dysfunction underlies the apathy associated with chronic cannabis use.

Objectives

The aim of this study was to examine the relationship between dopaminergic function and subjective apathy in cannabis users.

Methods

We measured dopamine synthesis capacity (indexed as the influx rate constant K _i ^cer ) via 3,4-dihydroxy-6-[¹⁸F]-fluoro-l-phenylalanine positron emission tomography and subjective apathy using the self-rated Apathy Evaluation Scale (AES-S) in 14 regular cannabis users.

Results

All subjects scored in excess of 34 points on the AES-S (median [interquartile range] 59.5 [7.5]), indicative of significant apathy based on normative data. K _i ^cer was inversely correlated to AES-S score in the whole striatum and its associative functional subdivision (Spearman’s rho?=??0.64, p?=?0.015 [whole striatum]; rho?=??0.69, p?=?0.006 [associative]) but not in the limbic or sensorimotor striatal subdivisions. There were no significant relationships between AES-S and current cannabis consumption (rho?=?0.28, p?=?0.34) or age of first cannabis use (rho?=?0.25, p?=?0.40).

Conclusions

These findings indicate that the reduction in striatal dopamine synthesis capacity associated with chronic cannabis use may underlie reduced reward sensitivity and amotivation associated with chronic cannabis use.     

Polydopamine nanoparticles kill cancer cells

Polydopamine (PD) is a synthetic melanin analogue of growing importance in the field of biomedicine, especially with respect to cancer research, due, in part, to its biocompatibility. But little is known about the cytotoxic effects of PD on cancer cell lines. PD is a UV-vis absorbing material whose absorbance overlaps with that of formazan salts, which are used to assess cell viability in MTT assays. In this study, a protocol has been established to eliminate the contributing absorbance of PD at 550 nm, and has been applied to characterize the cytotoxicity of PD nanoparticles in both healthy and breast cancer cell lines. Once the protocol is applied, it was found that PD is per se an antineoplastic system, meaning it selectively kills cancer cells, especially those of breast cancer, but it has no toxic effect on healthy cells. The mechanism of action could be related to the production of ROS and the alteration of iron homeostasis in lysosomes. To the best of our knowledge there are only a few examples of nanoparticle systems devoid of drugs that selectively kill cancer cells.

Polydopamine (PD) is a synthetic melanin analogue of growing importance in the field of biomedicine, especially with respect to cancer research, due, in part, to its biocompatibility.     

Absolute requirement of spermidine for growth and cell cycle progression of fission yeast (Schizosaccharomyces pombe)

Schizosaccharomyces pombe cells that cannot synthesize spermidine or spermine because of a deletion-insertion in the gene coding for S-adenosylmethionine decarboxylase (Deltaspe2) have an absolute requirement for spermidine for growth. Flow cytometry studies show that in the absence of spermidine an overall delay of the cell cycle progression occurs with some accumulation of cells in the G(1) phase; as little as 10(-6) M spermidine is sufficient to maintain normal cell cycle distribution and normal growth. Morphologically some of the spermidine-deprived cells become spherical at an early stage with little evidence of cell division. On further incubation in the spermidine-deprived medium, growth occurs in most of the cells, not by cell division but rather by cell elongation, with an abnormal distribution of the actin cytoskeleton, DNA (4', 6-diamidino-2-phenylindole staining), and calcofluor-staining moieties. More prolonged incubation in the spermidine-deficient medium leads to profound morphological changes including nuclear degeneration.     

Death,treatment decisions and the permanent vegetative state: evidence from families and experts

Some brain injured patients are left in a permanent vegetative state, i.e., they have irreversibly lost their capacity for consciousness but retained some autonomic physiological functions, such as breathing unaided. Having discussed the controversial nature of the permanent vegetative state as a diagnostic category, we turn to the question of the patients’ ontological status. Are the permanently vegetative alive, dead, or in some other state? We present empirical data from interviews with relatives of patients, and with experts, to support the view that the ontological state of permanently vegetative patients is unclear: such patients are neither straightforwardly alive nor simply dead. Having defended this view from counter-arguments we turn to the practical question as to how these patients ought to be treated. Some relatives and experts believe it is right for patients to be shifted from their currently unclear ontological state to that of being straightforwardly dead, but many are concerned or even horrified by the only legally sanctioned method guaranteed to achieve this, namely withdrawal of clinically assisted nutrition and hydration. A way of addressing this distress would be to allow active euthanasia for these patients. This is highly controversial; but we argue that standard objections to allowing active euthanasia for this particular class of permanently vegetative patients are weakened by these patients’ distinctive ontological status.