Hereditary cystatin C amyloid angiopathy: identification of the disease-causing mutation and specific diagnosis by polymerase chain reaction based analysis

Summary Hereditary cystatin C amyloid angiopathy (HCCAA) is a dominantly inherited disease characterized by amyloidosis, dementia and fatal cerebral hemorrhage of young adults. A method for rapid and simple diagnosis of HCCAA is described. It is based upon oligonucleotide-directed enzymatic amplification of a 275-bp genomic DNA segment containing exon 2 of the cystatin C gene from a blood sample, followed by digestion of the amplification product with AluI. Loss of an AluI recognition site in the amplified DNA segment from HCCAA patients results in a deviating band-pattern at agarose gel electrophoresis, compared with that obtained from normal subjects or unaffected HCCAA family members. In a population of 9 patients with manifest HCCAA, 14 patients with other causes of brain hemorrhage and 16 healthy individuals, the diagnostic procedure displayed a sensitivity and specificity for HCCAA of 100%. Amplified DNA segments from 4 HCCAA patients of four different families were analyzed by nucleotide sequencing; the HCCAA-causing mutation in all families was found to be a single TA substitution in the codon for amino acid residue 68 of cystatin C.     

Protein arginine methyltransferase 5 (PRMT5) activates WNT/β-catenin signalling in breast cancer cells via epigenetic silencing of DKK1 and DKK3

Protein arginine methyltransferase 5 (PRMT5) activity is dysregulated in many aggressive cancers and its enhanced levels are associated with increased tumour growth and survival. However, the role of PRMT5 in breast cancer remains underexplored. In this study, we show that PRMT5 is overexpressed in breast cancer cell lines, and that it promotes WNT/β-CATENIN proliferative signalling through epigenetic silencing of pathway antagonists, DKK1 and DKK3, leading to enhanced expression of c-MYC, CYCLIN D1 and SURVIVIN. Through chromatin immunoprecipitation (ChIP) studies, we found that PRMT5 binds to the promoter region of WNT antagonists, DKK1 and DKK3, and induces symmetric methylation of H3R8 and H4R3 histones. Our findings also show that PRMT5 inhibition using a specific small molecule inhibitor, compound 5 (CMP5), reduces PRMT5 recruitment as well as methylation of H3R8 and H4R3 histones in the promoter regions of DKK1 and DKK3, which consequently results in reduced expression CYCLIN D1 and SURVIVIN. Furthermore, CMP5 treatment either alone or in combination with 5-Azacytidine and Trichostatin A restored expression of DKK1 and DKK3 in TNBCs. PRMT5 inhibition also altered the growth characteristics of breast cancer cells and induced their death. Collectively, these results show that PRMT5 controls breast cancer cell growth through epigenetic silencing of WNT/β-CATENIN pathway antagonists, DKK1 and DKK3, resulting in up-regulation of WNT/β-CATENIN proliferative signalling.     

Perceived Stress at Work Is Associated with Lower Levels of DHEA-S

Background

It is known that long-term psychosocial stress may cause or contribute to different diseases and symptoms and accelerate aging. One of the consequences of prolonged psychosocial stress may be a negative effect on the levels of dehydroepiandrosterone (DHEA) and its sulphated metabolite dehydroepiandrosterone sulphate (DHEA-S). The aim of this study is to investigate whether levels of DHEA and DHEA-S differ in individuals who report perceived stress at work compared to individuals who report no perceived stress at work.

Methods

Morning fasting DHEA-S and DHEA levels were measured in serum in a non-stressed group (n = 40) and a stressed group (n = 41). DHEA and DHEA-S levels were compared between the groups using ANCOVA, controlling for age.

Results

The mean DHEA-S levels were 23% lower in the subjects who reported stress at work compared to the non-stressed group. Statistical analysis (ANCOVA) showed a significant difference in DHEA-S levels between the groups (p = 0.010). There was no difference in DHEA level between the groups.

Conclusions

This study indicates that stressed individual have markedly lower levels of DHEA-S. Given the important and beneficial functions of DHEA and DHEA-S, lower levels of DHEA-S may constitute one link between psychosocial stress, ill health and accelerated ageing.     

Modelled population growth based on reproduction differs from life tables based on age determination in Danish raccoon dogs ( Nyctereutes procyonoides )

The population of raccoon dogs (Nyctereutes procyonoides) in Denmark has increased rapidly from 1995 when the first was recorded until today where 3291 raccoon dogs are trapped, shot by hunters or road killed. The aims of this study are to present the first data on reproduction and life tables of raccoon dogs in Denmark and to compare mortality from modelled life tables with game bag records and sampled raccoon dogs in different age groups. In this study, the uteri of 89 adult females (> 10 months) were examined for placental scars (PSC), and 561 individuals (289 males, 272 females) were aged using pulp cavity width and dental lines in canine teeth. The litter size of raccoon dogs in Denmark is to date the largest litter size recorded in the wild (mean ± SE) 10.8 ± 0.4, range 1–16 pubs and fecundity 8.4 ± 0.6 pubs. The percent-reproducing females are 78–83%, based on dark and all PSC, respectively. A significant difference was found between the proportion of individuals composing the different age groups based on age determination of individuals collected (Ntage) and the modelled number of individuals in age groups based on fecundity and different mortality rate (Ntmodel), X2 = 8, p < 0.05. The discrepancy between the relatively high reproduction and lifetables may be due to older and more experienced animals that avoid culling. A low population density in a newly founded Danish population of raccoon dogs, together with a milder climate where raccoon dogs can forage during the winter, may cause an exceptionally high reproduction in Danish raccoon dogs.     

A two-tiered mechanism by which Cdc42 controls the localization and activation of an Arp2/3-activating motor complex in yeast

The establishment of cell polarity in budding yeast involves assembly of actin filaments at specified cortical domains. Elucidation of the underlying mechanism requires an understanding of the machinery that controls actin polymerization and how this machinery is in turn controlled by signaling proteins that respond to polarity cues. We showed previously that the yeast orthologue of the Wiskott-Aldrich Syndrome protein, Bee1/Las17p, and the type I myosins are key regulators of cortical actin polymerization. Here, we demonstrate further that these proteins together with Vrp1p form a multivalent Arp2/3-activating complex. During cell polarization, a bifurcated signaling pathway downstream of the Rho-type GTPase Cdc42p recruits and activates this complex, leading to local assembly of actin filaments. One branch, which requires formin homologues, mediates the recruitment of the Bee1p complex to the cortical site where the activated Cdc42p resides. The other is mediated by the p21-activated kinases, which activate the motor activity of myosin-I through phosphorylation. Together, these findings provide insights into the essential processes leading to polarization of the actin cytoskeleton.     

Dynamics of yeast Myosin I: evidence for a possible role in scission of endocytic vesicles

Cortical actin patches are dynamic structures required for endocytosis in yeast. Recent studies have shown that components of cortical patches localize to the plasma membrane in a precisely orchestrated manner, and their movements at and away from the plasma membrane may define the endocytic membrane invagination and vesicle scission events, respectively. Here, through live-cell imaging, we analyze the dynamics of the highly conserved class I unconventional myosin, Myo5, which also localizes to cortical patches and is known to be involved in endocytosis and actin nucleation. Myo5 exhibits a pattern of dynamic localization different from all cortical patch components analyzed to date. Myo5 associates with cortical patches only transiently and remains stationary during its brief cortical lifespan. The peak of Myo5 association with cortical patches immediately precedes the fast movement of Arp2/3 complex-associated structures away from the plasma membrane, thus correlating precisely with the proposed vesicle scission event. To further test the role of Myo5, we generated a temperature-sensitive mutant myo5 allele. In the myo5 mutant cells, Myo5 exhibits a significantly extended cortical lifespan as a result of a general impairment of Myo5 function, and Arp2 patches exhibit an extended slow-movement phase prior to the fast movement toward the cell interior. The myo5 mutant cells are defective in fluid-phase endocytosis and exhibit an increased number of invaginations on the membrane. Based on these results, we hypothesize that the myosin I motor protein facilitates the membrane fusion/vesicle scission event of endocytosis.     

Influence of voluntary exercise on hypothalamic norepinephrine

We combined hypothalamic tissue and plasma determinations ofnorepinephrine, dihydroxyphenylalanine, and dihydroxyphenylglycol withmeasurements of abdominal fat in voluntary running rats to examine therelationship among exercise training, hypothalamic and sympatheticnervous function, and body fat stores. The hypothalamic concentrationsof norepinephrine, dihydroxyphenylalanine, and dihydroxyphenylglycolwere reduced after exercise training(P < 0.01), with the amount ofnorepinephrine being strongly associated with the plasma norepinephrine(r = 0.58, P < 0.05) and dihydroxyphenylglycol (r = 0.65, P = 0.01) concentrations. Exercisetraining resulted in a diminution in abdominal fat mass(P < 0.01). A strongrelationship existed between fat mass and hypothalamic norepinephrinecontent (r = 0.83, P < 0.001). The presence of apositive relationship between the arterial and hypothalamicnorepinephrine levels provides presumptive evidence of an associationbetween noradrenergic neuronal activity of the hypothalamus andsympathetic nervous function. The observation that abdominal fat massis linked with norepinephrine in the hypothalamus raises thepossibility that alterations in body fat stores provide an afferentsignal linking hypothalamic function and the activity of thesympathetic nervous system.