Cytotoxicity of Exogenous Acetoacetate in Lithium Salt Form Is Mediated by Lithium and Not Acetoacetate.

BACKGROUND/AIM: The ketogenic diet has recently gained interest as potential adjuvant therapy for cancer. Many researchers have endeavored to support this claim in vitro. One common model utilizes treatment with exogenous acetoacetate in lithium salt form (LiAcAc). We aimed to determine whether the effects of treatment with LiAcAc on cell viability, as reported in the literature, accurately reflect the influence of acetoacetate. MATERIALS AND METHODS: Breast cancer and normal cell lines were treated with acetoacetate, in lithium and sodium salt forms, and cell viability was assessed. RESULTS: The effect of LiAcAc on cells was mediated by Li ions. Our results showed that the cytotoxic effects of LiAcAc treatment were significantly similar to those caused by LiCl, and also treatment with NaAcAc did not cause any significant cytotoxic effect. CONCLUSION: Treatment of cells with LiAcAc is not a convincing in vitro model for studying ketogenic diet. These findings are highly important for interpreting previously published results, and for designing new experiments to study the ketogenic diet in vitro.

Indigoid dyes by group E monooxygenases: mechanism and biocatalysis.

Since ancient times, people have been attracted by dyes and they were a symbol of power. Some of the oldest dyes are indigo and its derivative Tyrian purple, which were extracted from plants and snails, respectively. These 'indigoid dyes' were and still are used for coloration of textiles and as a food additive. Traditional Chinese medicine also knows indigoid dyes as pharmacologically active compounds and several studies support their effects. Further, they are interesting for future technologies like organic electronics. In these cases, especially the indigo derivatives are of interest but unfortunately hardly accessible by chemical synthesis. In recent decades, more and more enzymes have been discovered that are able to produce these indigoid dyes and therefore have gained attention from the scientific community. In this study, group E monooxygenases (styrene monooxygenase and indole monooxygenase) were used for the selective oxygenation of indole (derivatives). It was possible for the first time to show that the product of the enzymatic reaction is an epoxide. Further, we synthesized and extracted indigoid dyes and could show that there is only minor by-product formation (e.g. indirubin or isoindigo). Thus, group E monooxygenase can be an alternative biocatalyst for the biosynthesis of indigoid dyes.

Complex lipid metabolic remodeling is required for efficient hepatitis C virus replication.

The hepatitis C virus (HCV) life cycle is tightly linked to the host cell lipid metabolism with the endoplasmic reticulum-derived membranous web harboring viral RNA replication complexes and lipid droplets as virion assembly sites. To investigate HCV-induced changes in the lipid composition, we performed quantitative shotgun lipidomic studies of whole cell extracts and subcellular compartments. Our results indicate that HCV infection reduces the ratio of neutral to membrane lipids. While the amount of neutral lipids and lipid droplet morphology were unchanged, membrane lipids, especially cholesterol and phospholipids, accumulated in the microsomal fraction in HCV-infected cells. In addition, HCV-infected cells had a higher relative abundance of phosphatidylcholines and triglycerides with longer fatty acyl chains and a strikingly increased utilization of C18 fatty acids, most prominently oleic acid (FA [18:1]). Accordingly, depletion of fatty acid elongases and desaturases impaired HCV replication. Moreover, the analysis of free fatty acids revealed increased levels of polyunsaturated fatty acids (PUFAs) caused by HCV infection. Interestingly, inhibition of the PUFA synthesis pathway via knockdown of the rate-limiting Δ6-desaturase enzyme or by treatment with a high dose of a small-molecule inhibitor impaired viral progeny production, indicating that elevated PUFAs are needed for virion morphogenesis. In contrast, pretreatment with low inhibitor concentrations promoted HCV translation and/or early RNA replication. Taken together our results demonstrate the complex remodeling of the host cell lipid metabolism induced by HCV to enhance both virus replication and progeny production.

Neurovascular Specifications in the Alzheimer-Like Brain of Mice Affected by Focal Cerebral Ischemia: Implications for Future Therapies.

Alzheimer's disease (AD), the most frequent type of dementia, is a prototypical neurodegenerative disease, but shares with stroke certain common risk factors. Consequently, how vascular pathology may modulate AD pathogenesis has gained scientific attention. Therefore, aside from typical features of AD (e.g., amyloid-ß, tau hyperphosphorylation, and cholinergic dysfunction), changes within the 'neurovascular unit' (NVU) are of particular interest. This study focused on cholinergic, choline acetyltransferase (ChAT)-immunopositive, and tyrosine hydroxylase (TH)-containing neurons in association with the vasculature to explore the neurovascular complex of the AD brain affected by stroke. Wild-type and triple-transgenic (3xTg) mice of different ages underwent unilateral permanent focal cerebral ischemia. Histochemical analyses comprised diverse neuronal and vascular NVU components, and markers of AD. Immunofluorescence labeling confirmed the existence of Aß deposits and phospho-tau together with glial reactions and morphologically altered endothelia, visualized by Solanum tuberosum lectin. Twenty-four hours after ischemia induction, immunoreactivities for ChAT and TH declined in the ischemia-affected striatum and, at least in part, in the ischemic border zone and ipsilateral neocortex. Correlation analyses indicated simultaneous degeneration of neuronal and vascular components. A trend for more severe affection of ChAT was observed in younger as compared with older mice. The present findings suggest complex interactions within the NVU of the AD-like brain affected by ischemia, comprising alterations of the cholinergic system in conjunction with vascular pathology. Hence, it may be worthwhile to explore the impact of a cellular stabilization approach on vascular and glial elements in AD in terms of a potential disease-alleviating strategy.