Fatty acid binding protein 4 expression in cerebral vascular malformations: implications for vascular remodelling.

AIM: Arteriovenous malformations (AVM) and cavernous malformations (CM) are the most commonly encountered cerebral vascular malformations, which are dynamic lesions with de novo growth potentials. Postnatal angiogenesis and vasculogenesis have been postulated to play a role in the pathogenesis of these malformations. Fatty acid binding protein 4 (FABP4) is an intracellular lipid chaperone, which is expressed in a subset of endothelial cells. FABP4 enhances the angiogenic responses of endothelial cells and is not expressed in normal cerebral vasculature. Herein, we investigated the hypothesis that FABP4 expression may be up-regulated in AVM and CM. METHODS: The abundance of FABP4 expression was analysed by immunohistochemistry on 35 paraffin-embedded AVM and CM sections. FABP4-expressing cells were further characterized by double immunofluorescence using antibodies against various cell-specific markers. RESULTS: Heterogenous FABP4 expression was detected in 100% AVM and 78% of CM samples. Endothelial cell FABP4 expression was present in 65% and 43% of AVM and CM, respectively. Interestingly, a population of FABP4-positive perivascular cells was detected in 100% of AVM and 86% of CM sections examined. These cells were negative for markers of macrophages and smooth muscle cells, but expressed vimentin, a marker of mesenchymal cells, including fibroblasts. CONCLUSION: FABP4 expression is detected in AVM and CM in a subset of endothelial cells and some perivascular fibroblast-like vimentin-positive cells.

Hypercholesterolemia impairs immunity to tuberculosis.

We demonstrate that apolipoprotein E -deficient (ApoE(-/-)) mice are highly susceptible to tuberculosis and that their susceptibility depends on the severity of hypercholesterolemia. Wild-type (WT) mice and ApoE(-/-) mice fed a low-cholesterol (LC) or high-cholesterol (HC) diet were infected with approximately 50 CFU Mycobacterium tuberculosis Erdman by aerosol. ApoE(-/-) LC mice were modestly more susceptible to tuberculosis than WT LC mice. In contrast, ApoE(-/-) HC mice were extremely susceptible, as evidenced by 100% mortality after 4 weeks with tuberculosis. The lung pathology of ApoE(-/-) HC mice was remarkable for giant abscess-like lesions, massive infiltration by granulocytes, elevated inflammatory cytokine production, and a mean bacterial load approximately 2 log units higher than that of WT HC mice. Compared to WT HC mice, the gamma interferon response of splenocytes restimulated ex vivo with M. tuberculosis culture filtrate protein was delayed in ApoE(-/-) HC mice, and they failed to control M. tuberculosis growth in the lung. OT-II cells adoptively transferred into uninfected ApoE(-/-) HC mice had a weak proliferative response to their antigen, indicating impaired priming of the adaptive immune response. Our studies show that ApoE(-/-) deficiency is associated with delayed expression of adaptive immunity to tuberculosis caused by defective priming of the adaptive immune response and that elevated serum cholesterol is responsible for this effect.

Tuberculosis susceptibility of diabetic mice.

Increased susceptibility to infections, including tuberculosis (TB), is a major cause of morbidity and mortality in patients with diabetes. Despite the clinical importance of this problem, little is known about how diabetes impairs protective immunity. We modeled this phenomenon by infecting acute (< or = 1 mo) or chronic (> or = 3 mo) diabetic mice with a low aerosol dose of Mycobacterium tuberculosis (Mtb) Erdman. Diabetes was induced by streptozotocin (STZ) treatment of C57BL/6 mice, while another mouse strain and diabetes model were used to confirm key observations. Lungs from acute diabetic and euglycemic mice had similar bacterial burdens, cytokine expression profiles, and histopathology. In contrast, chronic diabetic mice had > 1 log higher bacterial burden and more inflammation in the lung compared with euglycemic mice. The expression of adaptive immunity was delayed in chronic diabetic mice, shown by reduced early production of IFN-gamma in the lung and by the presence of fewer Mtb antigen (ESAT-6)-responsive T cells compared with euglycemic mice within the first month of infection. However, after 2 months of TB disease proinflammatory cytokines levels were higher in chronic diabetic than euglycemic mice. Here we show that Mtb infection of STZ-treated mice provides a useful model to study the effects of hyperglycemia on immunity. Our data indicate that the initiation of adaptive immunity is impaired by chronic hyperglycemia, resulting in a higher steady-state burden of Mtb in the lung.

Positive transcriptional regulatory element located within exon 1 of elastin gene.

Elastin gene transcription is cell type specific and developmentally regulated, but the promoter often exhibits relatively weak activity in transient transfections of cells that express elastin at high levels. To search for positive-acting regulatory sequences, we isolated genomic clones spanning the mouse elastin gene and extensive 5'- and 3'-flanking regions. Restriction fragments of potential regulatory regions were ligated 5' or 3' relative to the active promoter to test for enhancer activity in transient transfections of fetal rat lung fibroblasts, which express elastin at high levels, and distal lung epithelial cells, which do not express detectable elastin. Fragments of intron 1 did not exhibit significant enhancer activity. Inclusion of the 84-bp exon 1 and adjacent 5'-untranslated region increased activity of the elastin promoter approximately sixfold compared with parental constructs. Transfections with constructs of varying promoter length showed that as little as 40 bp of the 5' end of exon 1 confers enhanced activity in elastin-expressing rat lung fibroblasts, but these constructs had variable activity in lung epithelial cell lines. This region, localized between the transcription start site and extending into exon 1, binds Sp1 in nuclear extracts from elastin-expressing cells. These studies indicate a role for the 5' end of the first exon of the elastin gene in regulating strong transcriptional activity in elastogenic cells.

Modulation of the membrane-binding domain of tau protein: splicing regulation of exon 2.

Tau is a microtubule-associated protein whose transcript undergoes complex regulated splicing in the mammalian nervous system. The N-terminal domain of the protein interacts with the axonal membrane, and is modulated by regulated inclusion of exons 2 and 3. These two tau exons are alternatively spliced cassettes, in which exon 3 never appears independently of exon 2. Previous work with tau minigene constructs indicated that exon 2 resembles a constitutive exon. In this study, we show that exon 2 is regulated by a combination of exonic and intronic enhancers and silencers. Furthermore, we demonstrate that known splicing regulators affect the ratio of exon 2 isoforms. Lastly, we tentatively pinpoint the site of action of several splicing factors which regulate tau exon 2.

Modulation of the membrane-binding projection domain of tau protein: splicing regulation of exon 3.

Tau is a microtubule-associated protein whose transcript undergoes complex regulated splicing in the mammalian nervous system. The N-terminal domain of the protein interacts with the axonal membrane, and is modulated by differential inclusion of exons 2 and 3. These two tau exons are alternatively spliced cassettes, in which exon 3 never appears independently of exon 2. Previous work with tau minigene constructs indicated that exon 3 is intrinsically suboptimal and its primary regulator is a weak branch point. In this study, we confirm the role of the weak branch point in the regulation of exon 3 but also show that the exon is additionally regulated by a combination of exonic enhancers and silencers. Furthermore, we demonstrate that known splicing regulators affect the ratio of exon 3 isoforms, Lastly, we tentatively pinpoint the site of action of several splicing factors which regulate tau exon 3.