Regulation of Alternative Pathway Activation and C3a Production by Adipose Cells

Adipose tissue is a major source of adpisin/factor D of the alternative pathway of complement. Adipose tissue also expresses the two other complement components which are involved in the first activation step of the alternative pathway, factor B and C3, and this step is activated in adipose tissue, producing C3a/Acylation Stimulating Protein (C3a/ASP), a stimulator of triglyceride synthesis. Complement activation is a highly regulated process, however, nothing is known about regulation of complement activation in adipose tissue. To gain insight into the nature of adipose complement activation and its regulation, we have now examined the expression of several complement activation regulatory genes, and analyzed the production of C3a/ASP in lean vs. obese, adipsin-deficient mice. We found that undifferentiated preadipocytes expressed the mRNAs encoding the negative regulatory proteins Crry and factor H, but expression of both genes was decreased upon differentiation. The positive regulator properdin, as well as Crry and factor H, were found in adipose tissue. None of these genes was regulated in murine genetic obesity. To investigate the relative levels of complement activation in lean vs. adipsin-deficient obese mice, we developed a radioimmunoassay for measurement of murine C3a/ASP in plasma. We report that there was no significant difference in the level of C3a in lean vs. obese plasma; however, we found a positive correlation between C3a and plasma triglyceride levels in normal lean mice.     

Mapping of a candidate region for susceptibility to inclusion body myositis in the human major histocompatibility complex

 Inclusion body myositis (IBM) is a form of idiopathic inflammatory myopathy of unknown aetiology. A strong association with HLA class II (HLA-DR3) suggested a role for genes in the human major histocompatibility complex (MHC) in the predisposition to this disease. In this study, we have taken advantage of the ancestral haplotype (AH) concept and historical recombinations to map for a possible susceptibility gene(s) in the MHC. We performed detailed typing of three MHC-related HSP70 genes and defined allelic combinations in the context of MHC AH. We also modified existing methods to give a simple and accurate method for typing two TNF microsatellites. Using the HSP70 and TNF markers and HLA-DR, –B, and C4 typing of our patients with IBM, we defined a potential site for the MHC-associated susceptibility gene(s) in the region between HLA-DR and C4. Received: 16 July 1998 / Revised: 14 January 1999     

Lauric acid alleviates insulin resistance by improving mitochondrial biogenesis in THP-1 macrophages

Molecular Biology Reports - Mitochondrial dysfunction plays a crucial role in the central pathogenesis of insulin resistance and type 2 diabetes mellitus. Macrophages play important roles in the...     

Phosphorylation of centromeric histone H3 variant regulates chromosome segregation in Saccharomyces cerevisiae

The centromeric histone H3 variant (CenH3) is essential for chromosome segregation in eukaryotes. We identify posttranslational modifications of Saccharomyces cerevisiae CenH3, Cse4. Functional characterization of cse4 phosphorylation mutants shows growth and chromosome segregation defects when combined with kinetochore mutants okp1 and ame1. Using a phosphoserine-specific antibody, we show that the association of phosphorylated Cse4 with centromeres increases in response to defective microtubule attachment or reduced cohesion. We determine that evolutionarily conserved Ipl1/Aurora B contributes to phosphorylation of Cse4, as levels of phosphorylated Cse4 are reduced at centromeres in ipl1 strains in vivo, and in vitro assays show phosphorylation of Cse4 by Ipl1. Consistent with these results, we observe that a phosphomimetic cse4-4SD mutant suppresses the temperature-sensitive growth of ipl1-2 and Ipl1 substrate mutants dam1 spc34 and ndc80, which are defective for chromosome biorientation. Furthermore, cell biology approaches using a green fluorescent protein–labeled chromosome show that cse4-4SD suppresses chromosome segregation defects in dam1 spc34 strains. On the basis of these results, we propose that phosphorylation of Cse4 destabilizes defective kinetochores to promote biorientation and ensure faithful chromosome segregation. Taken together, our results provide a detailed analysis, in vivo and in vitro, of Cse4 phosphorylation and its role in promoting faithful chromosome segregation.     

Toxicological effects of inorganic nanoparticles on human lung cancer A549 cells

Many researches have shown that anionic clays can be used as delivery carriers for drug or gene molecules due to their efficient cellular uptake in vitro, and enhanced permeability and retention effect in vivo. It is, therefore, highly required to establish a guideline on their potential toxicity for practical applications. The toxicity of anionic clay, layered metal hydroxide nanoparticle, was evaluated in two human lung epithelial cells, carcinoma A549 cells and normal L-132 cells, and compared with that in other human cancer cell lines such as cervical adenocarcinoma cells (HeLa) and osteosarcoma cells (HOS). The present nanoparticles showed little cytotoxic effects on the proliferation and viability of four cell lines tested at the concentrations used (<250 μg/ml) within 48 h. However, exposing cancer cells to high concentrations (250-500 μg/ml) for 72 h resulted in an inflammatory response with oxidative stress and membrane damage, which varied with the cell type (A549 > HOS > HeLa). On the other hand, the toxicity mechanism seems to be different from that of other inorganic nanoparticles frequently studied for biological and medicinal applications such as iron oxide, silica, and single walled carbon nanotubes. Iron oxide caused cell death associated with membrane damage, while single walled carbon nanotube induced oxidative stress followed by apoptosis. Silica triggered an inflammation response without causing considerable cell death for both cancer cells and normal cells, whereas layered metal hydroxide nanoparticle did not show any cytotoxic effects on normal L-132 cells in terms of inflammation response, oxidative stress, and membrane damage at the concentration of less than 250 μg/ml. It is , therefore, highly expected that the present nanoparticle can be used as a efficient vehicle for drug delivery and cancer cell targeting as well.     

In vitro cytotoxicity screening of water-dispersible metal oxide nanoparticles in human cell lines

In this study, we present in vitro cytotoxicity of iron oxide (Fe₃O₄) and manganese oxide (MnO) using live/dead cell assay, lactate dehydrogenase assay, and reactive oxygen species detection with variation of the concentration of nanoparticles (5–500 μg/ml), incubation time (18–96 h), and different human cell lines (lung adenocarcinoma, breast cancer cells, and glioblastoma cells). The surface of nanoparticles is modified with polyethyleneglycol-derivatized phospholipid to enhance the biocompatibility, water-solubility, and stability under an aqueous media. While the cytotoxic effect was negligible for 18 h incubation even at highest concentration of 500 μg/ml, MnO nanoparticle represented higher level of toxicity than those of Fe₃O₄ and the commercial medical contrast reagent, Feridex after 2 and 4 day incubation time. However, the cytotoxicity of Fe₃O₄ is equivalent or better than Feridex based on the live/dead cell viability assay. The engineered MnO and Fe₃O₄ exhibited excellent stability compared with Feridex for a prolonged incubation time.