Additional Supporting Information may be found in the online version of this article.

HEP_23441_sm_SupFig1.tif24136KSupporting Fig. 1: Effect of 5HT on proliferation of hepatocellular cancer cells. (A) 3H-thymidin-incorporation was determined in Huh7 and HepG2. After 48h serum deprivation cells were labelled and stimulated for 24h with FCS 10%, SFM or 5HT (in the absence of FCS). Dose response curve of 5HT compared to SFM revealed a maximum at 100 μM 5HT (n=6, *p<0.01). (B) Same experimental conditions as in (A) were used for BrdU- and Ki67-staining. Overlay with Hoechst illustrated less cells in SFM and a similiar number of cells in media containing FCS or 5HT. (C) Quantification of BrdU-staining showed significantly less cells in SFM (n=3, *p<0.05). A similar observation was made with Ki67 staining. The ratio of BrdU or Ki67 positive cells to the total number of cells (Hoechst) was equal under the three different conditions.
HEP_23441_sm_SupFig2.tif3517KSupporting Fig. 2: (A) Experimental setup for concentration dependent experiments. (B) Agonists to 5HT receptors HTR1, HTR2A, HTR7 do not increase the survival of Huh7, but phorbol-myristate-acetate (PMA) and α-Me-HTP (selective for HTR2B) increase survival two- to three-fold (n=4, *p<0.01). (C) Accordingly, antagonists to 5HT receptor HTR1, HTR2A and HTR7 have no effect on survival of Huh7 (n=3). Ritanserin, a general 5HT antagonist of HTR2 abolished the survival effect of 5HT (n=8, *p<0.001). (D) Identification of the HT2B receptor in Huh7 and HepG2 by western blot analysis.
HEP_23441_sm_SupFig3.tif8201KSupporting Fig. 3: Illustration of the suggested mechanism. Black lines indicate activation of p70S6K and 4E-BP1, red lines illustrate inhibition.
HEP_23441_sm_SupFig4.tif5215KSupporting Fig. 4: HeLa cells (human cervical cancer) (A) and MCF7 cells (human breast cancer) (B) were treated with four concentrations of 5HT for 24h after an initial serum deprivation of 48h. MTT assay was used to quantify survival. 5HT did not increase survival in these cells compared to serum free media (SFM) and media containing 10% fetal calf serum (FCS) (n=6 per group). HEF5 cells (human fibroblasts) (C) and PC3 (human prostate cancer) (D) were subsequently treated with 100 μM 5HT and survival was compared with standard culture conditions (FCS) and SFM using MTT assay. 5HT did not increase survival (n=6 per group).
HEP_23441_sm_SupTab.doc27KSupporting Table. 1: Cross tabulation for HTR2B and p-p70S6K of 168 HCC biopsies

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