Dual Signaling of the Fas Receptor: Initiation of Both Apoptotic and Necrotic Cell Death Pathways

Murine L929 fibrosarcoma cells were transfected with the human Fas (APO-1/CD95) receptor, and the role of various caspases in Fas-mediated cell death was assessed. Proteolytic activation of procaspase-3 and -7 was shown by Western analysis. Acetyl-Tyr-Val-Ala-Asp-chloromethylketone and benzyloxycarbonyl-Asp(OMe)-Glu(OMe)-Val-Asp(OMe)-fluoromethylketone, tetrapeptide inhibitors of caspase-1– and caspase-3–like proteases, respectively, failed to block Fas-induced apoptosis. Unexpectedly, the broad-spectrum caspase inhibitors benzyloxycarbonyl-Val-Ala-Asp(OMe)-fluoromethylketone and benzyloxycarbonyl-Asp(OMe)-fluoromethylketone rendered the cells even more sensitive to Fas-mediated cell death, as measured after 18 h incubation. However, when the process was followed microscopically, it became clear that anti-Fas–induced apoptosis of Fas-transfected L929 cells was blocked during the first 3 h, and subsequently the cells died by necrosis. As in tumor necrosis factor (TNF)-induced necrosis, Fas treatment led to accumulation of reactive oxygen radicals, and Fas-mediated necrosis was inhibited by the oxygen radical scavenger butylated hydroxyanisole. However, in contrast to TNF, anti-Fas did not activate the nuclear factor κB under these necrotic conditions. These results demonstrate the existence of two different pathways originating from the Fas receptor, one rapidly leading to apoptosis, and, if this apoptotic pathway is blocked by caspase inhibitors, a second directing the cells to necrosis and involving oxygen radical production

A Caspase-activated Factor (CAF) Induces Mitochondrial Membrane Depolarization and Cytochrome c Release by a Nonproteolytic Mechanism

It is well established that apoptosis is accompanied by activation of procaspases and by mitochondrial changes, such as decrease in mitochondrial transmembrane potential (ΔΨm) and release of cytochrome c. We analyzed the causal relationship between activated caspases and these mitochondrial phenomena. Purified recombinant caspase-1, -11, -3, -6, -7, and -8 were incubated with mitochondria in the presence or absence of additional cellular components, after which ΔΨm was determined. At lower caspase concentrations, only caspase-8 was able to activate a cytosolic factor, termed caspase-activated factor (CAF), which resulted in decrease in ΔΨm and release of cytochrome c. Both CAF-mediated activities could not be blocked by protease inhibitors, including oligopeptide caspase inhibitors. CAF-induced cytochrome c release, but not decrease of ΔΨm, was blocked in mitochondria from cells overexpressing Bcl-2. CAF is apparently involved in decrease of ΔΨm and release of cytochrome c, whereas Bcl-2 only prevents the latter. Hence, CAF may form the link between death domain receptor–dependent activation of procaspase-8 and the mitochondrial events studied

Novel genetic loci underlying human intracranial volume identified through genome-wide association

Intracranial volume reflects the maximally attained brain size during development, and remains stable with loss of tissue in late life. It is highly heritable, but the underlying genes remain largely undetermined. In a genome-wide association study of 32,438 adults, we discovered five novel loci for intracranial volume and confirmed two known signals. Four of the loci are also associated with adult human stature, but these remained associated with intracranial volume after adjusting for height. We found a high genetic correlation with child head circumference (ρgenetic=0.748), which indicated a similar genetic background and allowed for the identification of four additional loci through meta-analysis (Ncombined = 37,345). Variants for intracranial volume were also related to childhood and adult cognitive function, Parkinson’s disease, and enriched near genes involved in growth pathways including PI3K–AKT signaling. These findings identify biological underpinnings of intracranial volume and provide genetic support for theories on brain reserve and brain overgrowth

Novel genetic loci associated with hippocampal volume

The hippocampal formation is a brain structure integrally involved in episodic memory, spatial navigation, cognition and stress responsiveness. Structural abnormalities in hippocampal volume and shape are found in several common neuropsychiatric disorders. To identify the genetic underpinnings of hippocampal structure here we perform a genome-wide association study (GWAS) of 33,536 individuals and discover six independent loci significantly associated with hippocampal volume, four of them novel. Of the novel loci, three lie within genes (ASTN2, DPP4 and MAST4) and one is found 200 kb upstream of SHH. A hippocampal subfield analysis shows that a locus within the MSRB3 gene shows evidence of a localized effect along the dentate gyrus, subiculum, CA1 and fissure. Further, we show that genetic variants associated with decreased hippocampal volume are also associated with increased risk for Alzheimer's disease (rg =-0.155). Our findings suggest novel biological pathways through which human genetic variation influences hippocampal volume and risk for neuropsychiatric illness

Skewed X-inactivation is common in the general female population

X-inactivation is a well-established dosage compensation mechanism ensuring that X-chromosomal genes are expressed at comparable levels in males and females. Skewed X-inactivation is often explained by negative selection of one of the alleles. We demonstrate that imbalanced expression of the paternal and maternal X-chromosomes is common in the general population and that the random nature of the X-inactivation mechanism can be sufficient to explain the imbalance. To this end, we analyzed blood-derived RNA and whole-genome sequencing data from 79 female children and their parents from the Genome of the Netherlands project. We calculated the median ratio of the paternal over total counts at all X-chromosomal heterozygous single-nucleotide variants with coverage ≥10. We identified two individuals where the same X-chromosome was inactivated in all cells. Imbalanced expression of the two X-chromosomes (ratios ≤0.35 or ≥0.65) was observed in nearly 50% of the population. The empirically observed skewing is explained by a theoretical model where X-inactivation takes place in an embryonic stage in which eight cells give rise to the hematopoietic compartment. Genes escaping X-inactivation are expressed from both alleles and therefore demonstrate less skewing than inactivated genes. Using this characteristic, we identified three novel escapee genes (SSR4, REPS2, and SEPT6), but did not find support for many previously reported escapee genes in blood. Our collective data suggest that skewed X-inactivation is common in the general population. This may contribute to manifestation of symptoms in carriers of recessive X-linked disorders. We recommend that X-inactivation results should not be used lightly in the interpretation of X-linked variants

Is the assumed natural history of vulvar intraepithelial neoplasia III based on enough evidence? A systematic review of 3322 published patients

To establish the true natural history of VIN III from literature data. In a systematic review, data of women with VIN III indexed in several computer databases were pooled. The effect of treatment was correlated with recurrences and progression of VIN III. Ninety-seven articles met the inclusion criteria. Data of 3322 patients were available. The mean age at diagnosis of VIN III was 46. This decreased over time, although not significantly (P = 0.08). Recurrences were seen as often after local excision as after vulvectomy. The percentage of recurrences was lower, but not absent, after free surgical margins than after involved surgical margins (P < 0.001). 6.5% of the 3322 patients progressed to an invasive vulvar carcinoma. Occult carcinomas were diagnosed in 3.2% of patients and 3.3% carcinomas were diagnosed during follow-up. The mean age at diagnosis of invasive vulvar carcinoma was 52 years. Nine percent of 88 untreated patients progressed in 12 to 96 months to invasive vulvar carcinoma. Only 1.2% of the 3322 patients showed complete regression, mostly during the first 10 months after diagnosis, 41% of which was related to pregnancy. Evidence exists that VIN III may progress to invasive vulvar carcinoma. However, the available literature suggests that the progression rate to invasive vulvar carcinoma is low. The incidence of invasion as found in this systematic review is probably even too high, because overreporting of (micro)invasive cases cannot be excluded. Only a prospective registration using a standardized pathology examination will provide information about the real natural history of VIN II