Drosophila basement membrane collagen col4a1 mutations cause severe myopathy

Recent data from clinical and mammalian genetic studies indicate that COL4A1 mutations manifest with basement membrane defects that result in muscle weakness, cramps, contractures, dystrophy and atrophy. In-depth studies of mutant COL4A1-associated muscle phenotype, however, are lacking and significant details of the muscle-specific pathomechanisms remain unknown. In this study, we have used a comprehensive set of Drosophila col4a1 and col4a2 mutants and a series of genetic and mutational analyses, gene, protein expression, and immunohistochemistry experiments in order to establish a Drosophila model and address some of these questions. The Drosophila genome contains two type IV collagen genes, col4a1 and col4a2. Mutant heterozygotes of either gene are viable and fertile, whereas homozygotes are lethal. In complementation analysis of all known mutants of the locus and a complementation matrix derived from these data we have identified the dominant lesions within the col4a1, but not within the col4a2 gene. Expression of a col4a1 transgene partially rescued the dominant and recessive mutant col4a1 alleles but not the col4a2 mutations that were all recessive. Partial complementation suggested that col4a1 gene mutations have strong antimorph effect likely due to the incorporation of the mutant protein into the triple helix. In col4a1 mutants, morphological changes of the oviduct muscle included severe myopathy with centronuclear myofibers leading to gradual development of female sterility. In larval body wall muscles ultrastructural changes included disturbance of A and I bands between persisting Z bands. In the most severely affected DTS-L3 mutant, we have identified four missense mutations within the coding region of the col4a1 gene two of which affected the Y within the Gly-X-Y unit and a 3' UTR point mutation. In conclusion, our Drosophila mutant series may serve as an effective model to uncover the mechanisms by which COL4A1 mutations result in compromised myofiber-basement membrane interactions and aberrant muscle function.     

Effects of anaphylaxis mediators on partitioned pulmonary vascular resistance during ragweed shock in dogs

We examined theeffect of anaphylactic shock on the longitudinal distribution ofpulmonary vascular resistance (PVR) in ragweed-sensitized dogs in whichPVR was partitioned into an upstream arterial component (Rus) and adownstream venous and capillary component (Rds). We also assessedwhether Rus and Rds would be reduced by pretreatment with histamineH₁- andH₂-receptor blocking agents andwith cyclooxygenase and lipoxygenase pathway inhibitors.Anesthetized animals were examined on separate occasions 3 wk apart inwhich one of the treatments was randomly given. The pulmonary arterialocclusion technique was used to determine segmental pressure drops.During ragweed challenge, PVR increased 4 times compared with thepreshock value (3.04 vs. 12.07 mmHg · l¹ · min;P < 0.05). Although both Rus and Rds increased postshock, the greatestrelative increase occurred in Rds. None of the treatments reducedpartitioned resistances compared with no treatment. Our results showthat, under conditions of anaphylactic shock, increases in Rus and Rdscould not be ascribed to release of histamine or products of thecyclooxygenase and lipoxygenase pathways.

     

Interendothelial junctions in kidney vessels

Summary The interendothelial junctions of all segments of the renal vasculature have been studied in eight species using the freeze-fracture technique. Three types of junctions have been found. Combinations of tight and gap junction elements are characteristic for interlobular arteries and proximal afferent arterioles. Continuous tight junction strands not subdivided into individual particles are typical for the glomerular arterioles close to the glomerulus and the vasa recta. The interendothelial junctions of glomerular and peritubular capillaries and cortical veins are characterized by slight elevations decorated with sparse arrays of particles on the P-face of the endothelial cell plasma membrane.These studies were supported by the German Research Foundation within the SFB 90 Cardiovascular System.     

Expression of Transglutaminase K in Normal Cervix Tissue and Cervix Carcinomas

The localization and expression of transglutaminase K has been investigated immunohistochemically in normal cervix tissue (n=15) and in cervix carcinomas (n=23). The distribution of the transglutaminase K was compared with the staining patterns of cytokeratin 10, Ki-67, p53, and oestrogen and progesterone receptors in these tumours. Weak to strong membrane-bound immunoreactivity for transglutaminase K was detected in almost all cervix carcinomas analyzed. The immunostaining was heterogeneous, with visual differences between individual tumour cells. 66.7% of normal cervix tissues revealed no immunoreactivity for the transglutaminase K. In normal cervix tissue, the immunoreactivity was confined to upper cervix layers, predominantly to the superficial and intermediate cell layers. The intensity of both the immunostaining and the number of transglutaminase K-positive cells were upregulated in cervix carcinomas as compared to normal cervix tissue. When the coexpressions of transglutaminase K with markers of proliferation and differentiation were analyzed, no statistically significant correlation was found. Our findings indicate that (1) transglutaminase K is upregulated at the protein level in cervix carcinomas as compared to normal cervix tissue; (2) upregulation of the transglutaminase K in cervix carcinoma is not exclusively induced by alterations of epithelial differentiation or proliferation, but by different, unknown mechanisms; and (3) upregulation of transglutaminase K in cervix carcinomas may play an important role for the regulation of tumour invasive properties by modulating cell–cell interactions.     

APOE ε4 moderates abnormal CSF-abeta-42 levels,while neurocognitive impairment is associated with abnormal CSF tau levels in HIV+ individuals – a cross-sectional observational study

Background

Cerebrospinal fluid (CSF) biomarkers Aβ1-42, t-tau and p-tau have a characteristic pattern in Alzheimer’s Disease (AD). Their roles in HIV-associated neurocognitive disorder (HAND) remains unclear.

Methods

Adults with chronic treated HIV disease were recruited (n = 43, aged 56.7 ± 7.9; 32% aged 60+; median HIV duration 20 years, >95% plasma and CSF HIV RNA <50 cp/mL, on cART for a median 24 months). All underwent standard neuropsychological testing (61% had HAND), APOE genotyping (30.9% carried APOE ε4 and 7.1% were ε4 homozygotes) and a lumbar puncture. Concentrations of Aβ1-42, t-tau and p-tau were assessed in the CSF using commercial ELISAs. Current neurocognitive status was defined using the continuous Global Deficit Score, which grades impairment in clinically relevant categories. History of HAND was recorded. Univariate correlations informed multivariate models, which were corrected for nadir CD4-T cell counts and HIV duration.

Results

Carriage of APOE ε4 predicted markedly lower levels of CSF Aβ1-42 in univariate (r = -.50; p = .001) and multivariate analyses (R² = .25; p < .0003). Greater levels of neurocognitive impairment were associated with higher CSF levels of p-tau in univariate analyses (r = .32; p = .03) and multivariate analyses (R² = .10; p = .03). AD risk prediction cut-offs incorporating all three CSF biomarkers suggested that 12.5% of participants had a high risk for AD. Having a CSF-AD like profile was more frequent in those with current (p = .05) and past HIV-associated dementia (p = .03).

Conclusions

Similarly to larger studies, APOE ε4 genotype was not directly associated with HAND, but moderated CSF levels of Aβ1-42 in a minority of participants. In the majority of participants, increased CSF p-tau levels were associated with current neurocognitive impairment. Combined CSF biomarker risk for AD in the current HIV+ sample is more than 10 times greater than in the Australian population of the same age. Larger prospective studies are warranted.

     

Lysozyme, a mediator of sepsis that produces vasodilation by hydrogen peroxide signaling in an arterial preparation

In septic shock, systemic vasodilation and myocardial depression contribute to the systemic hypotension observed. Both components can be attributed to the effects of mediators that are released as part of the inflammatory response. We previously found that lysozyme (Lzm-S), released from leukocytes, contributed to the myocardial depression that develops in a canine model of septic shock. Lzm-S binds to the endocardial endothelium, resulting in the production of nitric oxide (NO), which, in turn, activates the myocardial soluble guanylate cyclase (sGC) pathway. In the present study, we determined whether Lzm-S might also play a role in the systemic vasodilation that occurs in septic shock. In a phenylephrine-contracted canine carotid artery ring preparation, we found that both canine and human Lzm-S, at concentrations similar to those found in sepsis, produced vasorelaxation. This decrease in force could not be prevented by inhibitors of NO synthase, prostaglandin synthesis, or potassium channel inhibitors and was not dependent on the presence of the vascular endothelium. However, inhibitors of the sGC pathway prevented the vasodilatory activity of Lzm-S. In addition, Aspergillus niger catalase, which breaks down H(2)O(2), as well as hydroxyl radical scavengers, which included hydroquinone and mannitol, prevented the effect of Lzm-S. Electrochemical sensors corroborated that Lzm-S caused H(2)O(2) release from the carotid artery preparation. In conclusion, these results support the notion that when Lzm-S interacts with the arterial vasculature, this interaction results in the formation of H(2)O(2), which, in turn, activates the sGC pathway to cause relaxation. Lzm-S may contribute to the vasodilation that occurs in septic shock.