NF-kappaB factor c-Rel mediates neuroprotection elicited by mGlu5 receptor agonists against amyloid beta-peptide toxicity

Opposite effects of nuclear factor-kappaB (NF-kappaB) on neuron survival rely on activation of diverse NF-kappaB factors. While p65 is necessary for glutamate-induced cell death, c-Rel mediates prosurvival effects of interleukin-1beta. However, it is unknown whether activation of c-Rel-dependent pathways reduces neuron vulnerability to amyloid-beta (Abeta), a peptide implicated in Alzheimer's disease pathogenesis. We show that neuroprotection elicited by activation of metabotropic glutamate receptors type 5 (mGlu5) against Abeta toxicity depends on c-Rel activation. Abeta peptide induced NF-kappaB factors p50 and p65. The mGlu5 agonists activated c-Rel, besides p50 and p65, and the expression of manganese superoxide dismutase (MnSOD) and Bcl-X(L). Targeting c-Rel expression by RNA interference suppressed the induction of both antiapoptotic genes. Targeting c-Rel or Bcl-X(L) prevented the prosurvival effect of mGlu5 agonists. Conversely, c-Rel overexpression or TAT-Bcl-X(L) addition rescued neurons from Abeta toxicity. These data demonstrate that mGlu5 receptor activation promotes a c-Rel-dependent antiapoptotic pathway responsible for neuroprotection against Abeta peptide.     

Calcium bioavailability and kinetics of calcium ascorbate and calcium acetate in rats

The objective was to investigate the bioavailability and mechanism of calcium absorption of calcium ascorbate (ASC) and calcium acetate (AC). A series of studies was performed in adult Sprague-Dawley male rats. In the first study, each group of rats (n = 10/group) was assigned to one of the five test meals labeled with (45)Ca: (i) 25 mg calcium as heated ASC or (ii) unheated ASC, (iii) 25 mg calcium as unheated AC, (iv) 3.6 mg Ca as unheated ASC, or (v) unheated AC. Femur uptake indicated better calcium bioavailability from ASC than AC at both calcium loads. A 5-min heat treatment partly reduced bioavailability of ASC. Kinetic studies were performed to further investigate the mechanism of superior calcium bioavailability from ASC. Two groups of rats (n = 10/group) received oral doses of 25 mg Ca as ASC or AC. Each dose contained 20 micro Ci (45)Ca. Two additional groups of rats (n = 10/group) received an intravenous injection (iv) of 10 micro Ci (45)Ca after receiving an unlabeled oral dose of 25 mg calcium as ASC or AC. Sequential blood samples were collected over 48 hrs. Urine and fecal samples were collected every 12 hrs for 48 hrs and were analyzed for total calcium and (45)Ca content. Total calcium and (45)Ca from serum, urine, and feces were fitted by a compartment kinetics model with saturable and nonsaturable absorption pathways by WinSAAM (Windows-based Simulation Analysis and Modeling). The difference in calcium bioavailability between the two salts was due to differences in saturable rather than passive intestinal absorption and not to endogenous secretion or calcium deposition rate. The higher bioavailability of calcium ascorbate was due to a longer transit time in the small intestine compared with ASC.     

Glomerulopathy and mutations in NPHS1 and KIRREL2 in soft-coated Wheaten Terrier dogs

Dogs of the soft-coated wheaten terrier breed (SCWT) are predisposed to adult-onset, genetically complex, protein-losing nephropathy (average onset age = 6.3 ± 2.0 years). A genome-wide association study using 62 dogs revealed a chromosomal region containing three statistically significant SNPs (p _raw ≤ 4.13 × 10^?8; p _genome ≤ 0.005) when comparing DNA samples from affected and geriatric (≥14 years) unaffected SCWTs. Sequencing of candidate genes in the region revealed single nucleotide changes in each of two closely linked genes, NPHS1 and KIRREL2, which encode the slit diaphragm proteins nephrin and Neph3/filtrin, respectively. In humans, mutations in nephrin and decreased expression of Neph3 are associated with podocytopathy and protein-losing nephropathy. The base substitutions change a glycine to arginine in the fibronectin type 3 domain of nephrin and a proline to arginine in a conserved proline-rich region in Neph3. These novel mutations are not described in other species, nor were they found in 550 dogs of 105 other breeds, except in 3 dogs, including an affected Airedale terrier, homozygous for both substitutions. Risk for nephropathy is highest in dogs homozygous for the mutations (OR = 9.06; 95 % CI = 4.24–19.35). This is the first molecular characterization of an inherited podocytopathy in dogs and may serve as a model for continued studies of complex genetic and environmental interactions in glomerular disease.     

Vegetation Pattern Formation Due to Interactions Between Water Availability and Toxicity in Plant–Soil Feedback

Development of a comprehensive theory of the formation of vegetation patterns is still in progress. A prevailing view is to treat water availability as the main causal factor for the emergence of vegetation patterns. While successful in capturing the occurrence of multiple vegetation patterns in arid and semiarid regions, this hypothesis fails to explain the presence of vegetation patterns in humid environments. We explore the rich structure of a toxicity-mediated model of the vegetation pattern formation. This model consists of three PDEs accounting for a dynamic balance between biomass, water, and toxic compounds. Different (ecologically feasible) regions of the model’s parameter space give rise to stable spatial vegetation patterns in Turing and non-Turing regimes. Strong negative feedback gives rise to dynamic spatial patterns that continuously move in space while retaining their stable topology.     

High-throughput luciferase reporter assay for small-molecule inhibitors of microRNA function

MicroRNAs (miRNAs) are endogenous, single-stranded, noncoding RNAs of 21 to 23 nucleotides that regulate gene expression, typically by binding the 3' untranslated regions of target messenger RNAs. It is estimated that miRNAs are involved in the regulation of 30% of all genes and almost every genetic pathway. Recently, the misregulation of miRNAs has been linked to various human diseases including cancer and viral infections, identifying miRNAs as potential targets for drug discovery. Thus, small-molecule modifiers of miRNAs could serve as lead structures for the development of new therapeutic agents and be useful tools in the elucidation of detailed mechanisms of miRNA function. As a result, we have developed a high-throughput screen for potential small-molecule regulators of the liver-specific microRNA miR-122, which is involved in hepatocellular carcinoma development and hepatitis C virus infection. Our small-molecule screen employs a Huh7 human hepatoma cell line stably transfected with a Renilla luciferase sensor for endogenous miR-122. The assay was optimized and validated using an miR-122 antisense agent and a previously identified small-molecule miR-122 inhibitor. The described reporter assay will enable the high-throughput screening of small-molecule miR-122 inhibitors and can be readily extended to other miRNAs.