Curcumin is a chemical component extracted from the rhizome of Zingiberaceae and Araceae. Studies have described various effects of curcumin including reducing blood fat, choleretic actions, and anti-tumor, anti-inflammatory, and anti-oxidation effects [13–16]. Several studies have indicated that the effect of curcumin on SREBPs is related to the mechanism of curcumin regulating blood lipid metabolism [4, 5, 8–12]. Kumar et al  found that curcumin can regulate the expression of NPC1L1 through SREBP-2 to inhibit cholesterol absorption in Caco-2 cells. We used a mouse model of cholesterol gallstone formation induced by a high-fat diet to show these results. Our preliminary study further confirmed that curcumin can reduce cholesterol gallstone formation and inhibit NPC1L1. As we investigated how curcumin affects NPC1L1 expression through SREBP-2, we found different results from previous studies. Although curcumin lowered SREBP-2 mRNA levels, as in other studies, we did not see simultaneously down-regulated SREBP-2 protein levels. In the Caco-2 cell line, SREBP-2 mRNA expression decreased with higher doses (Fig. 1a) and times (Fig. 1b) of curcumin treatment. However, we did not find pSREBP-2 precursor levels to be inhibited (Fig. 2). Western blotting showed that while pSREBP-2 (almost 130 kDa) wasn’t significantly down-regulated, the amount of mSREBP-2 protein (55–70 kDa) decreased (Fig. 2). The SREBP-2 mRNA levels were inconsistent with pSREBP-2 levels, which was particularly evident through the curcumin time course. Regardless of whether curcumin was administered long-term (0 ~ 72 h) or short-term (within 24 h), the pSREBP-2 did not decrease following with the mRNA inhibition. Kang et al  found that the curcumin-mediated reduction of SREBP-2 promoter activity was dependent on the inhibition of SP-1. Consistent with Kang’s study, we also found downregulated transcriptional and translational levels of SP-1 at 24 h treatment, but this down-regulation was relieved after 48 h treatment (Fig. 1b, Fig. 2b). This suggests that curcumin’s effect on SREBP-2 may not involve SP-1, or that there may be another pathway. Considering the amount that mSREBP-2 protein decreased after curcumin treatment and that SREBP-2 is activated as a transcription factor through the proteolytic process [6, 7], we hypothesized that the effect of curcumin on SREBP-2 may depend on its proteolytic process. Endoplasmic reticulum co-localization showed that curcumin modulated SREBP-2 distribution within the cell, decreased mSREBP-2 in the nucleus, and inhibited the proteolysis process (Fig. 3c). In Brown and Goldstein's research [6, 7], they suggest that the SREBP-2/SCAP complex is the real substrate of S1P and key for the proteolytic process of SREBP-2. Therefore, we also detected SCAP in our study. SCAP and SREBP-2 showed consistent changes in transcription and translation. These results suggest that SCAP may not be a target of curcumin. As S1P protein levels decreased after curcumin treatment, curcumin may inhibit the expression of SREBP-2 by inhibiting the expression of S1P.
Several other studies have demonstrated inconsistency between SREBP-2 transcription and translation levels. Field et al  showed in rat liver cells that increased cholesterol in Caco-2 cells inhibited proteolysis of SREBP-2. The mSREBP-2 was decreased but pSREBP-2 levels were not significantly changed, and this process was accompanied by inhibition of SREBP-2 gene expression. Liu et al  reported that the change of SREBP-2 protein was not statistically significant after curcumin treatment compared with controls. Sato et al  found that the SREBP-2 gene includes a Sterol Regulatory Element (SRE) identical to the one on the promoter sequence of the human LDL receptor, thus SREBP-2 may regulate its own expression through modulating sterol levels. When the proteolytic process of SREBP-2 was inhibited, mSREBP-2 transcription factor levels decreased, which affected the expression of cholesterol metabolism related proteins and also the expression of SREBP-2. These results may explain why the expression of SREBP-2 mRNA does not follow the protein level.
Some studies have reported downregulated SREBP-2 upon curcumin treatment[9, 11, 12]. However, these studies do not show the molecular weight of the protein markers in their figures, making it hard to determine the molecular weight of the SREBP-2 detected in their western blots. Thus, the detected protein may be mSREBP-2 rather than pSREBP-2. Liu et al  studied the function of curcumin using antibodies against SREBP-2 purchased from Santa Cruz which have only been used to detect pSREBP-2. Our results indicate that curcumin first affects the SREBP-2 proteolysis process rather than inhibiting its protein expression. Protein expression inhibition may be a long-term effect. The inhibitory effect of curcumin on the transcription of SREBP-2 mRNA persisted long-term, with decreased mRNA of the pSREBP-2 detected after 96 hours treatment (Fig. 2b). When SREBP-2 proteolysis is inhibited, pSREBP-2 may be degraded by other pathways, and its gene expression may remain continuously inhibited, eventually leading to down-regulation of the pSREBP-2 protein expression. This mechanism still requires further experimentation to fully understand.
The exact mechanism for how curcumin reduces the hydrolysis of SREBP-2 and deceases mSREBP-2 protein levels through inhibiting S1P expression remains unknown. However, we conclude here that the mechanism of curcumin treatment on SREBP-2 is not through short-term inhibition of protein expression, although curcumin can indeed inhibit the transcription of SREBP-2 mRNA long-term. Thus, it appears that curcumin plays an important role via inhibiting the activity of SREBP-2 rather than directly affecting its expression of gene and protein.