## 2 Cell organelles and metabolic pathways that contribute to HCV infection
### Assembly of VLDL  
VLDLs are released by hepatocytes in the liver and have a similar architecture to lipid droplets in that they are composed of a TAG- and CE-rich hydrophobic core that is bounded by a layer of phospholipid and protein [39]. The proteins that coat VLDL consist of non-exchangeable (apolipoprotein B [apoB]) and exchangeable apoproteins (principally apoCI-III and apoE) [40]. ApoB is a long, amphipathic protein of 4536 amino acids [41,42] that is essential for assembly of VLDL. Assembly is considered to proceed through two stages of lipidation of apoB. Firstly, apoB is lipidated by microsomal transfer protein (MTP), which is an ER lumenal protein. MTP is a heterodimer of two proteins with molecular weights of 88 and 58 kDa, the smaller of which is protein disulfide isomerase [43]. MTP both binds to apoB and facilitates transfer of lipid to the protein during its translation, ultimately leading to the production of a precursor VLDL (pre-VLDL) particle that is poorly lipidated [44]. Bulk addition of lipid constitutes the second stage in VLDL assembly, generating a mature, secretion-competent particle. However, the intracellular location and processes involved in lipidation of pre-VLDL are not well understood. It appears that the further addition of lipid to pre-VLDL does not rely on either MTP or the need for ongoing TAG synthesis [45,46]. Indeed, all the available evidence indicates that lipid droplets supply the lipid that is needed to convert pre-VLDL to a mature particle [47]. By way of example, increasing the expression levels of adipocyte differentiation-relatedprotein (ADRP), the major protein on the surface of lipid droplets in hepatocyte-derived cells [48], stimulates accumulation of droplets and reduces secretion of VLDL from rat hepatocytes [49].  
Mobilization of lipid from droplets relies on several enzymes involved in TAG and cholesterol metabolism. Transfer of TAG and CE from a lipid droplet to pre-VLDL seems to require de-esterification followed by re-esterification. A number of reports have investigated involvement of several enzymes including triacylglycerglycerol hydrolase (TGH), DGAT1, and ACAT2. TGH converts TAG to acylglycerols and fatty acids [50]. Expression of TGH in mice and rat hepatoma cells gives a modest rise in triglyceride secretion [50,51]. As stated above, DGAT1 catalyzes the last stage in the synthesis of TAG and there is evidence to suggest that the enzyme participates in hepatic VLDL assembly [52]. By contrast, DGAT2 appears to direct TAG for lipid storage [52]. ACAT2 plays a major contribution to CE formation in the liver and intestine [53,54]. Studies in rat cells have shown that overexpression of human DGAT1 and ACAT2 individually leads to increased secretion of VLDL [55]. Interestingly, compared to ACAT1, ACAT2 induced higher levels of VLDL secretion, thereby providing evidence for its predominant role in CE formation in the liver. However, it should be noted that there are conflicting data from overexpression studies in rodents, [for example [56]], which illustrates the difficulty with formal demonstration that any individual enzyme can stimulate VLDL secretion.