This study examined the ability of (i) pure nisin, (ii) nisin-producing Lactococcus lactis strain CHCC5826, and (iii) the non-nisin-producing L. lactis strain CHCH2862 to affect the composition of the intestinal microbiota of human flora-associated rats. The presence of both the nisin-producing and the non-nisin-producing L. lactis strains significantly increased the number of Bifidobacterium cells in fecal samples during the first 8 days but decreased the number of enterococci/streptococci in duodenum, ileum, cecum, and colon samples as detected by selective cultivation. No significant changes in the rat fecal microbiota were observed after dosage with nisin. Pearson cluster analysis of denaturing gradient gel electrophoresis profiles of the 16S rRNA genes present in the fecal microbial population revealed that the microbiota of animals dosed with either of the two L. lactis strains were different from that of control animals dosed with saline. However, profiles of the microbiota from animals dosed with nisin did not differ from the controls. The concentrations of nisin estimated by competitive enzyme-linked immunosorbent assay (ELISA) were approximately 10-fold higher in the small intestine and 200-fold higher in feces than the corresponding concentrations estimated by a biological assay. This indicates that nisin was degraded or inactivated in the gastrointestinal tract, since fragments of this bacteriocin are detected by ELISA while an intact molecule is needed to retain biological activity.

Background: The transdifferentiation of hepatic stellate cells (HSCs) into myofibroblasts is a major mechanism for stroma development in hepatic metastasis, but their regulatory pathways remain unclear. Transdifferentiated HSCs from fibrotic liver express high levels of the fibrillar collagen receptor discoidin domain receptor 2 (DDR2), but it is unclear if DDR2 plays a direct profibrogenic role in the tumour microenvironment. Aim: To assess the impact of DDR2 on the prometastatic role of HSC-derived myofibroblasts. Methods: Hepatic metastases were induced in DDR2 ^−/− and DDR2 ^+/+ mice by intrasplenic injection of MCA38 colon carcinoma cells, and their growth and features were characterised. Stromagenic, angiogenic and cancer cell proliferation responses were quantified in metastases by immunohistochemistry. The adhesion-, migration- and proliferation-stimulating activities of supernatants from primary cultured DDR2 ^−/− and DDR2 ^+/+ HSCs, incubated in MCA38 cell-conditioned medium, were evaluated in primary cultured liver sinusoidal endothelium cells (LSECs) and MCA38 cells. Gene expression signatures from freshly isolated DDR2 ^−/− and DDR2 ^+/+ HSCs were compared and DDR2-regulated genes were studied by RT-PCR under basal conditions and after stimulation with MCA38 tumour-conditioned media. Results: Metastases were increased three fold in DDR2 ^−/− livers, and contained a higher density of α−smooth muscle actin-expressing myofibroblasts, CD31-expressing microvessels and Ki67-expressing MCA38 cells than metastases in DDR2 ^+/+ livers. Media conditioned by MCA38-activated DDR2 ^−/− HSCs significantly increased adhesion, migration and proliferation of LSECs and MCA38 cells, compared with DDR2 ^+/+ HSCs. DDR2 deficiency in HSCs led to decreased gene expression of interferon γ-inducing factor interleukin (IL)-18 and insulin-like growth factor-I; and increased gene expression of prometastatic factors IL-10, transforming growth factor (TGF)β and vascular endothelial growth factor (VEGF), bone morphogenetic protein-7 and syndecan-1. MC38 tumour-conditioned media further exacerbated expression changes in DDR2-dependent IL-10, TGFβ and VEGF genes. Conclusion: DDR2 deficiency fosters the myofibroblast transdifferentiation of tumour-activated HSCs, generating a prometastatic microenvironment in the liver via HSC-derived factors. These findings underscore the role of stromal cells in conditioning the hepatic microenvironment for metastases through altered receptor–stroma interactions.

Discoidin domain receptors (DDR1 and DDR2) are tyrosine kinase receptors for fibrillar collagen implicated in postnatal development, tissue repair, and primary and metastatic cancer progression. While DDR1 has been described in tumor cells, DDR2 has been localized in the tumor stroma, but its presence in the tumor cells remains unknown. The aim of this study was to elucidate the role of DDR2 signaling in tumor cells during hepatic metastasis progression. DDR2 expression and phosphorylation in cultured human A375 melanoma cells was documented by Western blot analysis. A375 cells were stably transfected with a small interfering RNA (siRNA) against DDR2 and two clones were selected: A375R2-70 and A375R2-40, with 70 and 40% of the DDR2 protein expression respectively, compared to mock-transfected cells (A375R2-100). Development of experimental liver metastasis by intrasplenic inoculation of A375R2-70 and A37R2-40 clones was reduced by 60 and 75%, respectively, measured as tumor volume, compared to livers injected with A375R2-100 cells. Accordingly, A375R2-70 and A37R2-40 clones showed reduced in vitro gelatinase activity and JNK phosphorylation, compared to mock transfected cells, with maximal inhibition in A375R2-40. Additionally, A375 melanoma, SK-HEP hepatoma and HT-29 colon carcinoma human cell lines transiently transfected with siRNA against DDR2 also showed reduced proliferation and migration rates compared to mock-transfected ones. In conclusion, DDR2 promotes A375 melanoma metastasis to the liver and the underlying mechanism implicates regulation of metalloproteinase release, cell growth and chemotactic invasion of the host tissue.