Secreted Phospholipase A2 Involvement in Neurodegeneration: Differential Testing of Prosurvival and Anti-Inflammatory Effects of Enzyme Inhibition

There is increased interest in the contribution of secreted phospholipase A2 (sPLA2) enzymes to neurodegenerative diseases. Systemic treatment with the nonapeptide CHEC-9, a broad spectrum uncompetitive inhibitor of sPLA2, has been shown previously to inhibit neuron death and aspects of the inflammatory response in several models of neurodegeneration. A persistent question in studies of sPLA2 inhibitors, as for several other anti-inflammatory and neuroprotective compounds, is whether the cell protection is direct or due to slowing of the toxic aspects of the inflammatory response. To further explore this issue, we developed assays using SY5Y (neuronal cells) and HL-60 (monocytes) cell lines and examined the effects of sPLA2 inhibition on these homogeneous cell types in vitro. We found that the peptide inhibited sPLA2 enzyme activity in both SY5Y and HL-60 cultures. This inhibition provided direct protection to SY5Y neuronal cells and their processes in response to several forms of stress including exposure to conditioned medium from HL-60 cells. In cultures of HL-60 cells, sPLA2 inhibition had no effect on survival of the cells but attenuated their differentiation into macrophages, with regard to process development, phagocytic ability, and the expression of differentiation marker CD36, as well as the secretion of proinflammatory cytokines TNF-α and IL-6. These results suggest that sPLA2 enzyme activity organizes a cascade of changes comprising both cell degeneration and inflammation, processes that could theoretically operate independently during neurodegenerative conditions. The effectiveness of sPLA2 inhibitor CHEC-9 may be due to its ability to affect both processes in isolation. Testing potential anti-inflammatory/neuroprotective compounds with these human cell lines and their conditioned media may provide a useful screening tool prior to in vivo therapeutic applications

Putative Nanobacteria Represent Physiological Remnants and Culture By-Products of Normal Calcium Homeostasis

Putative living entities called nanobacteria (NB) are unusual for their small sizes (50–500 nm), pleomorphic nature, and accumulation of hydroxyapatite (HAP), and have been implicated in numerous diseases involving extraskeletal calcification. By adding precipitating ions to cell culture medium containing serum, mineral nanoparticles are generated that are morphologically and chemically identical to the so-called NB. These nanoparticles are shown here to be formed of amorphous mineral complexes containing calcium as well as other ions like carbonate, which then rapidly acquire phosphate, forming HAP. The main constituent proteins of serum-derived NB are albumin, fetuin-A, and apolipoprotein A1, but their involvement appears circumstantial since so-called NB from different body fluids harbor other proteins. Accordingly, by passage through various culture media, the protein composition of these particles can be modulated. Immunoblotting experiments reveal that antibodies deemed specific for NB react in fact with either albumin, fetuin-A, or both, indicating that previous studies using these reagents may have detected these serum proteins from the same as well as different species, with human tissue nanoparticles presumably absorbing bovine serum antigens from the culture medium. Both fetal bovine serum and human serum, used earlier by other investigators as sources of NB, paradoxically inhibit the formation of these entities, and this inhibition is trypsin-sensitive, indicating a role for proteins in this inhibitory process. Fetuin-A, and to a lesser degree albumin, inhibit nanoparticle formation, an inhibition that is overcome with time, ending with formation of the so-called NB. Together, these data demonstrate that NB are most likely formed by calcium or apatite crystallization inhibitors that are somehow overwhelmed by excess calcium or calcium phosphate found in culture medium or in body fluids, thereby becoming seeds for calcification. The structures described earlier as NB may thus represent remnants and by-products of physiological mechanisms used for calcium homeostasis, a concept which explains the vast body of NB literature as well as explains the true origin of NB as lifeless protein-mineralo entities with questionable role in pathogenesis

Species Interactions during Diversification and Community Assembly in an Island Radiation of Shrews

Closely related, ecologically similar species often have adjacent distributions, suggesting competitive exclusion may contribute to the structure of some natural communities. In systems such as island archipelagos, where speciation is often tightly associated with dispersal over oceanic barriers, competitive exclusion may prevent population establishment following inter-island dispersal and subsequent cladogenesis.) species in the Philippines are the result of competitive exclusion preventing secondary invasion of occupied islands. We first compare ecological niche models between two widespread, allopatric species and find statistical support for their ecological similarity, implying that competition for habitat between these species is possible. We then examine dispersion patterns among sympatric species and find some signal for overdispersion of body size, but not for phylogenetic branch length. Finally, we simulate the process of inter-island colonization under a stochastic model of dispersal lacking ecological forces. Results are dependent on the geographic scope and colonization probability employed. However, some combinations suggest that the number of inter-island dispersal events necessary to populate the archipelago may be much higher than the minimum number of colonization events necessary to explain current estimates of species richness and phylogenetic relationships. If our model is appropriate, these results imply that alternative factors, such as competitive exclusion, may have influenced the process of inter-island colonization and subsequent cladogenesis.We interpret the combined results as providing tenuous evidence that similarity in body size may prevent co-occurrence in Philippine shrews and that competitive exclusion among ecologically similar species, rather than an inability to disperse among islands, may have limited diversification in this group, and, possibly other clades endemic to island archipelagos

Indicators of "Healthy Aging" in older women (65-69 years of age). A data-mining approach based on prediction of long-term survival

<p>Abstract</p> <p>Background</p> <p>Prediction of long-term survival in healthy adults requires recognition of features that serve as early indicators of successful aging. The aims of this study were to identify predictors of long-term survival in older women and to develop a multivariable model based upon longitudinal data from the Study of Osteoporotic Fractures (SOF).</p> <p>Methods</p> <p>We considered only the youngest subjects (<it>n </it>= 4,097) enrolled in the SOF cohort (65 to 69 years of age) and excluded older SOF subjects more likely to exhibit a "frail" phenotype. A total of 377 phenotypic measures were screened to determine which were of most value for prediction of long-term (19-year) survival. Prognostic capacity of individual predictors, and combinations of predictors, was evaluated using a cross-validation criterion with prediction accuracy assessed according to time-specific AUC statistics.</p> <p>Results</p> <p>Visual contrast sensitivity score was among the top 5 individual predictors relative to all 377 variables evaluated (mean AUC = 0.570). A 13-variable model with strong predictive performance was generated using a forward search strategy (mean AUC = 0.673). Variables within this model included a measure of physical function, smoking and diabetes status, self-reported health, contrast sensitivity, and functional status indices reflecting cumulative number of daily living impairments (HR ≥ 0.879 or RH ≤ 1.131; P < 0.001). We evaluated this model and show that it predicts long-term survival among subjects assigned differing causes of death (e.g., cancer, cardiovascular disease; P < 0.01). For an average follow-up time of 20 years, output from the model was associated with multiple outcomes among survivors, such as tests of cognitive function, geriatric depression, number of daily living impairments and grip strength (P < 0.03).</p> <p>Conclusions</p> <p>The multivariate model we developed characterizes a "healthy aging" phenotype based upon an integration of measures that together reflect multiple dimensions of an aging adult (65-69 years of age). Age-sensitive components of this model may be of value as biomarkers in human studies that evaluate anti-aging interventions. Our methodology could be applied to data from other longitudinal cohorts to generalize these findings, identify additional predictors of long-term survival, and to further develop the "healthy aging" concept.</p

Ageing, adipose tissue, fatty acids and inflammation

A common feature of ageing is the alteration in tissue distribution and composition, with a shift in fat away from lower body and subcutaneous depots to visceral and ectopic sites. Redistribution of adipose tissue towards an ectopic site can have dramatic effects on metabolic function. In skeletal muscle, increased ectopic adiposity is linked to insulin resistance through lipid mediators such as ceramide or DAG, inhibiting the insulin receptor signalling pathway. Additionally, the risk of developing cardiovascular disease is increased with elevated visceral adipose distribution. In ageing, adipose tissue becomes dysfunctional, with the pathway of differentiation of preadipocytes to mature adipocytes becoming impaired; this results in dysfunctional adipocytes less able to store fat and subsequent fat redistribution to ectopic sites. Low grade systemic inflammation is commonly observed in ageing, and may drive the adipose tissue dysfunction, as proinflammatory cytokines are capable of inhibiting adipocyte differentiation. Beyond increased ectopic adiposity, the effect of impaired adipose tissue function is an elevation in systemic free fatty acids (FFA), a common feature of many metabolic disorders. Saturated fatty acids can be regarded as the most detrimental of FFA, being capable of inducing insulin resistance and inflammation through lipid mediators such as ceramide, which can increase risk of developing atherosclerosis. Elevated FFA, in particular saturated fatty acids, maybe a driving factor for both the increased insulin resistance, cardiovascular disease risk and inflammation in older adults

Characterization of Granulations of Calcium and Apatite in Serum as Pleomorphic Mineralo-Protein Complexes and as Precursors of Putative Nanobacteria

Calcium and apatite granulations are demonstrated here to form in both human and fetal bovine serum in response to the simple addition of either calcium or phosphate, or a combination of both. These granulations are shown to represent precipitating complexes of protein and hydroxyapatite (HAP) that display marked pleomorphism, appearing as round, laminated particles, spindles, and films. These same complexes can be found in normal untreated serum, albeit at much lower amounts, and appear to result from the progressive binding of serum proteins with apatite until reaching saturation, upon which the mineralo-protein complexes precipitate. Chemically and morphologically, these complexes are virtually identical to the so-called nanobacteria (NB) implicated in numerous diseases and considered unusual for their small size, pleomorphism, and the presence of HAP. Like NB, serum granulations can seed particles upon transfer to serum-free medium, and their main protein constituents include albumin, complement components 3 and 4A, fetuin-A, and apolipoproteins A1 and B100, as well as other calcium and apatite binding proteins found in the serum. However, these serum mineralo-protein complexes are formed from the direct chemical binding of inorganic and organic phases, bypassing the need for any biological processes, including the long cultivation in cell culture conditions deemed necessary for the demonstration of NB. Thus, these serum granulations may result from physiologically inherent processes that become amplified with calcium phosphate loading or when subjected to culturing in medium. They may be viewed as simple mineralo-protein complexes formed from the deployment of calcification-inhibitory pathways used by the body to cope with excess calcium phosphate so as to prevent unwarranted calcification. Rather than representing novel pathophysiological mechanisms or exotic lifeforms, these results indicate that the entities described earlier as NB most likely originate from calcium and apatite binding factors in the serum, presumably calcification inhibitors, that upon saturation, form seeds for HAP deposition and growth. These calcium granulations are similar to those found in organisms throughout nature and may represent the products of more general calcium regulation pathways involved in the control of calcium storage, retrieval, tissue deposition, and disposal