Furthermore, metabolism of PC species has been linked to NASH pathogenesis: hepatic deletion of phosphocholine cytidylyltransferase (PCYT1) and knockdown of LPCAT3, two enzymes involved in PC metabolism, results in marked reductions in VLDL secretion, a key factor in the development of that NASH in humans . PCs are synthesized in mammals by two predominant pathways (Figure 1). In the first pathway, CDP-choline, generated by the sequential actions of choline kinase (CHK) and phosphocholine cytidylyltransferase (CEPT) on dietary choline, reacts with sn-1,2 diacylglycerols (DAGs) to form PC , . A second pathway, accounting for 30% of hepatic PCs, involves sequential methylation of phosphatidylethanolamine (PE) with S-adenosylmethionine catalyzed by the enzyme phosphatidylethanolamine N-methyltransferase (PEMT) , , .
The PCs synthesized by either pathway are readily converted into TAG. Figure 1 Schematic representation of major phosphatidylcholine (PC) biosynthetic pathways in human liver. Insight into the distribution of some hepatic lipid species was recently established by Debois et al. in normal and SS human liver specimens using cluster TOF-SIMS imaging . They demonstrated periportal enrichment of ��-tocopherol and cholesterol, along with a macrovesicular enrichment of TAGs, DAGs and FAs. However, the TOF-SIMS is a hard ionization technology resulting in significant lipid fragmentation, thus hampering the detection and discrimination of intact phospholipids.
In the present study we investigated hepatic phospholipid abundance by quantitative lipidomic profiling and phospholipid localization by MALDI-IMS, a ��soft�� technique more conducive to intact lipid ionization. We determined the zonal distribution of various intact phospholipid species in situ in human liver specimens of control, SS and NASH using MALDI Imaging Mass Spectrometry (MALDI IMS). In addition, we examined the in situ hepatic localization of an important enzyme in the phosphatidylcholine biosynthetic pathway, PEMT, to investigate if altered zonation of PC biosynthetic enzymes may additionally contribute to NAFLD progression. Materials and Methods Ethics Statement Subjects gave their informed written consent before participating in this study, which was approved by the Institutional Review Board of Vanderbilt University and registered at ClinicalTrials.
gov (NCT00983463). Brefeldin_A Human Subjects Class III obese women (n=33, ages 26�C59 years old) were recruited from the Center for Surgical Weight Loss at Vanderbilt University Medical Center prior to their scheduled bariatric procedures. Exclusion criteria included a history of previous liver disease (e.g. viral or autoimmune hepatitis, or hemochromatosis), significant alcohol use, concurrent infections, a cancer diagnosis within the previous 5 years, hemoglobin A1C >7.0, and the use of anti-diabetic drugs.