2-Mercaptobenzothiazole (MBT) is employed for the first time as a matrix for the analysis of lipids from tissue extracts using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. We demonstrate that the performance of MBT is superior to that of the matrixes commonly employed for lipids, due to its low vapor pressure, its low acidity, and the formation of small crystals, although because of the strong background at low m/z, it precludes detection of species below approximately 500 Da. This inconvenience can be partly overcome with the formation of Cs adducts. Using a polymer-based dual calibration, a mass accuracy of approximately 10 ppm in lipid extracts and of approximately 80 ppm in tissues is achieved. We present spectra from liver and brain lipid extracts where a large amount of lipid species is identified, in both positive and negative ion modes, with high reproducibility. In addition, the above-mentioned special properties of MBT allow its employment for imaging mass spectrometry. In the present work, images of brain and liver tissues showing different lipid species are presented, demonstrating the advantages of the employment of MBT.
The relevant structural, energetics, and regulatory roles of lipids are universally acknowledged. However, the high variability of lipid species and the large differences in concentrations make unraveling the role played by the different species in metabolism a titanic task. A recently developed technique, known as imaging mass spectrometry, may shed some light on the field, as it enables precise information to be obtained on the location of lipids in tissues. A review of the state of the art of the technique is presented in this manuscript, including detailed analysis of sample-preparation steps, data handling, and the identification of the species mapped so far.
Zonation affects liver parenchymal cell function and metabolism as well as nonparenchymal cell activation, but whether VLDL production is zonated has yet to be elucidated. Infection induces enhanced VLDL secretion by the liver. Ex vivo studies were undertaken to examine the liver heterogeneity for VLDL formation and secretion and their in vivo response to endotoxin. Highly pure periportal (PP) and perivenous (PV) hepatocytes were isolated from fasted lipopolysaccharide-treated, fasted, and fed rats. They were used to assess their capacity to release VLDL-apolipoprotein B (apoB) and lipid classes in relation to de novo lipid synthesis and the expression of genes crucial to VLDL production. Despite the common superior ability of PP hepatocytes for lipid release and zonal differences in lipid synthesis, zonated secretion of VLDL particles was observed in septic but not in normal fed or fasted livers. The endotoxininduced apoB secretion was more accentuated in PP hepatocytes; this was accompanied by a preferential PP increase in apoB and microsomal triglyceride transfer protein mRNA levels, whereas lipogenesis indicators were, if anything, similarly modified in hepatocytes of either acinar origin. We conclude that PP and PV hepatocytes exhibited similar capabilities for VLDL formation/secretion in normal conditions; however, the endotoxic pressure did zonate periportally. Septic hyperlipidemia and hyperlipoproteinemia have been postulated to be components of the innate defense against infection in humans and experimental models (1). The insertion of the lipid A component of Gram-negative bacterial lipopolysaccharide (LPS; or endotoxin) in the phospholipid layer of lipoproteins diminishes toxicity (2), as it enables endotoxin clearance and excretion by hepatic parenchymal cells (3). Septic hypertriglyceridemia is mainly caused by the accumulation of VLDL particles in the plasma (4) attributable to many changes in their metabolism (5, 6; see recent review in 7), including increased secretion by the liver (8).VLDL particles are assembled at the endoplasmic reticulum (ER) of hepatocytes in a process that depends absolutely upon the cellular availability of lipids, such as triglycerides (TGs), phospholipids, cholesterol, and cholesteryl esters, to correctly translate and translocate apolipoprotein B (apoB) to the lumen. ApoB-100 and apoB-48 are the essential apolipoproteins for VLDL assembly in the rat (9). They are expressed in excess, and the protein that is not lipidated to a sufficient extent is targeted to and eventually degraded by proteasomes (10). Correct apoB lipidation and translocation are controlled by the microsomal triglyceride transfer protein (MTP), whose binding and lipid transfer activities are probably the major determinant in VLDL secretion (11-13). The assembly of VLDL is a complex process that includes two major lipidation steps. A relatively small, dense, TG-poor lipoprotein particle is formed in the ER by cotranslational loading of apoB chains with lipid. Then, bulk lipidation and final m...
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