Role of Drosophila IKK gamma in a toll-independent antibacterial immune response

We have generated, by ethylmethane sulfonate mutagenesis, loss-of-function mutants in the Drosophila homolog of the mammalian I-kappa B kinase (IKK) complex component IKK gamma (also called NEMO). Our data show that Drosophila IKK gamma is required for the Relish-dependent immune induction of the genes encoding antibacterial peptides and for resistance to infections by Escherichia coli. However, it is not required for the Toll-DIF-dependent antifungal host defense. The results indicate distinct control mechanisms of the Rel-like transactivators DIF and Relish in the Drosophila innate immune response and show that Drosophila Toll does not signal through a IKK gamma-dependent signaling complex. Thus, in contrast to the vertebrate inflammatory response, IKK gamma is required for the activation of only one immune signaling pathway in Drosophila.     

DNA-ploidy analysis correlates with the histogenetic classification of thymic epithelial tumours

The ploidy values of the epithelial component were determined in a series of thymomas and organotypic thymic carcinomas using image cytometry and the results were compared with the histological tumour subtypes according to the histogenetic classification introduced by Marino, Müller Hermelink, and Kirchner (MMHK). Forty-six cases of thymic epithelial tumours were included in the study. After reclassification according to the MMHK classification, the distribution among the subtypes was as follows: three medullary, nine mixed type, five predominantly cortical (organoid), 16 cortical thymomas, and 13 well-differentiated thymic carcinomas. Single cell preparations were made from paraffin-embedded tumour tissue and stained according to Feulgen. Ploidy analysis was performed using an automated image analysis system. In five cases, DNA cytometry could not be performed, for technical reasons. The remaining 41 cases consisted of 11 diploid and 30 non-diploid tumours. The percentage of aneuploid tumours in the different subtypes increased from medullary (0 per cent) through mixed type (44.4 per cent), predominantly cortical (75 per cent), cortical (83.3 per cent) to well-differentiated thymic carcinomas (100 per cent). DNA-ploidy determination using image cytometry correlates with the concept of the MMHK classification of thymomas.     

A sandwich ELISA for properdin in clinical specimens

We have developed a symmetrical sandwich ELISA for measuring human properdin (P) in serum by using the globulin fraction from a commercial antiserum as the capture antibody adsorbed on the plastic. The detecting reagent was a glutaraldehyde conjugate of this Ig fraction with alkaline phosphatase. Two types of inhibition were observed in this study. First, inhibition was observed when greater than 2.5 micrograms/ml of the globulin fraction was used to coat the plates. A second type of inhibition was observed for serum dilutions less than 1/400; it was independent of the concentration of capture Ab and did not occur when purified P was assayed. The data generated with this assay could be fitted in log-log mode by a quadratic equation. The coefficient of the linear term in this equation was found to be the same for serum and purified P, within the limits of experimental error. The results for different samples run on the same plate were expressed in terms of the relative concentration of each sample required to produce an OD405 = 0.2. A sample of pooled normal human serum was run on each plate as a reference; it was assigned a titer of 100 ELISA units/ml (EU/ml). The titers of the unknown samples were expressed in terms of EU/ml by reference to this standard. For purified P, the assay could readily detect 10 ng/ml. By comparing purified P with our reference serum pool, we found that 1 EU equals 0.57 microgram. Day-to-day variation for a group of nine normal sera showed a mean difference of -0.85 EU/ml, SD 5.85 EU/ml. The mean titer for these normal sera was 78.9 EU/ml, SD 15.7 EU/ml. In three recovery experiments in which purified P was mixed with pooled normal serum, the recoveries ranged from 96 to 117%. We conclude that the sandwich ELISA constitutes an adequate immunochemical assay for human P in serum specimens.     

Association between platelet endothelial cellular adhesion molecule 1 (PECAM-1/CD31) polymorphisms and acute myocardial infarction: a study in patients from Sicily.

Adhesion of circulating cells to the arterial surface is among the first detectable events in atherogenesis. Cellular adhesion molecules, expressed by the vascular endothelium and by circulating leucocytes, mediate cell recruitment and their transendothelial migration. Platelet endothelial cellular adhesion molecule 1 (PECAM-1/CD31), involved in this migration, has been associated with the developmental course of atherosclerosis. A few studies have investigated an association between coronary heart disease and single nucleotide polymorphisms (SNPs) located in functionally important domains of the PECAM-1/CD31 gene. In particular, Ser563Asn and Gly670Arg SNPs have been described as susceptibility factors involved in acute myocardial infarction (AMI) in the Japanese male population. To confirm these observations, we studied 96 male patients (mean age 40 years; age range 20-46) affected by AMI and 118 healthy male controls (mean age 38 years, age range: 20-55), and analysed for the following PECAM-1/CD31 SNPs: Val125Leu, Asn563Ser and Gly670Arg. The frequency of the Gly670Arg polymorphism was significantly higher in patients with AMI (58.9% vs. 48.3%; P = 0.019), whereas the frequencies of the other two SNPs (Leu125Val and Ser563Asn) were not significantly different between patients and controls. By comparing the observed number of 670Arg/Arg genotypes in the patients with the expected number, calculated from the allele frequency in a healthy population, a significance of P = 0.02 (odds ratio, 2.04; 95% CI: 1.1-3.7) was obtained, supporting a recessive model of inheritance. Hence, the differences between patients and controls are significant, but relatively small. However, as AMI is a multifactorial disease, any single mutation will only provide a small or modest contribution to the risk, which also depends on environmental interaction. All in all, we believe that the results of the present study would add support to the role of pro/anti-inflammatory genotypes in determining susceptibility or resistance to immune-inflammatory diseases, including atherosclerosis.     

Pretectal jerk neuron activity during saccadic eye movements and visual stimulations in the cat

Summary The activity of jerk neurons was recorded extracellularly in the pretectum of the awake cat. The characteristic response of jerk neurons was a short, high-frequency burst that occurred after fast movements (jerks) of a large, structured visual stimulus, during saccadic eye movements in the light, and after on or off visual stimulation. Mean burst latency to pure visual jerks was 50 ms, whereas it was 30 ms to saccadic eye movements. Bursts were found to be stereotyped; the highest discharge rate was always at burst onset. Jerk neurons were not selective for stimulus parameters (such as movement amplitude or direction) except that in some neurons a weak correlation between stimulus velocity and discharge frequency was found. During saccades in the dark, clear bursts were only rarely found. In about half of the neurons, however, there was a slight but significant increase in the number of spikes above spontaneous frequency. Visual receptive fields were very large (46° horizontal and 35° vertical extent, on average). Nevertheless, the pretectal jerk neurons showed a rough retinotopic order, which was in accordance with the published retinotopy of the pretectum. Jerk neurons were found throughout the whole superficial pretectum, but preferentially in an area that corresponds to the nucleus of the optic tract (NOT) and the nucleus pretectalis posterior (NPP). Saccades were elicited by electrical stimulations at the sites where jerk neurons were recorded. The direction of the elicited saccades depended strongly on the pretectal stimulation site. A possible role of the jerk neurons as a visuomotor relay to elicit saccades or to modulate perception and attention is discussed.     

Decreased aminoacylation of mutant tRNAs in MELAS but not in MERRF patients

Mutations in human mitochondrial tRNA genes are associated witha number of multisystemic disorders. Using an assay that combinestRNA oxidation and circularization we have determined the relativeamounts and states of aminoacylation of mutant and wild-typetRNAs in tissue samples from patients with MELAS syndrome (mito-chondrial myopathy, encephalopathy, lactic acidosis, stroke-likeepisodes) and MERRF syndrome (myoclonus epilepsy with raggedred fibers), respectively. In most, but not all, biopsies fromMELAS patients carrying the A3243G substitution in the mitochondrialtRNA^Leu(UUR) gene, the mutant tRNA is under-represented amongprocessed and/or aminoacylated tRNAs. In contrast, in biopsiesfrom MERRF patients harboring the A8344G substitution in thetRNA^Lys gene neither the relative abundance nor the aminoacylationof the mutated tRNA is affected. Thus, whereas the A3243G mutationmay contribute to the pathogenesis of MELAS by reducing theamount of aminoacylated tRNA^Leu, the A8344G mutation does notaffect tRNA^Lys function in the same way. ⁺ To whom correspondence should be addressed. Tel: +1 617 4954396; Fax: +1 617 495 0758; Email: boerner@fas.harvard.edu