Hematopoietic cells are recruited to and co-opted by the growing tumor making expansive tumor growth possible. Although several cell types become associated with the growing tumor, macrophages play a fundamental role. The movement of macrophages across the basement membrane and through the extracellular matrix to the tumor site requires the activation of proteases, such as plasmin, at their cell surface. The proteolytic aspect of macrophage recruitment may represent an exploitable aspect of tumor growth in terms of therapeutic strategies. Here I show that the S100A10 protein facilitates the infiltration of macrophages into the site of tumor growth by stimulating the generation of the protease plasmin at their surface. Using a mouse model in which wild-type (WT) and S100A10-null mice are inoculated with tumor cells, a decrease in tumor-associated macrophages (TAMs) and greatly diminished tumor growth in tumors grown in S100A10-null mice was observed. Although tumor growth in S100A10-null mice could be restored by intraperitoneal injection of WT macrophages, S100A10-null macrophages only restored tumor growth when directly injected into the tumor. Lastly, selective depletion of macrophages from a WT mouse by liposome encapsulated clodronate treatment resulted in similar tumor growth deficits as in the S100A10-null mouse. These results highlight a new role for the S100A10 protein in the recruitment of TAMs to the tumor site and demonstrate a potential therapeutic strategy in which the tumor associated cells may be targeted.

Background Ovarian follicular fluid serves as the microenvironment for a maturing oocyte prior to ovulation. Previous studies have shown that gilts fed a high fibre (HF) diet before ovulation have improved fertility compared to gilts fed a control (C) diet, including a higher proportion of metaphase II oocytes following in vitro maturation (IVM). Hypothesis The molecular composition of porcine follicular fluid (pFF) was altered by the diet and that these alterations conferred the fertility benefits. Aims The aim of this study was to compare the protein composition of pFF from pigs fed a control diet with pFF of pigs fed a high fibre diet, to identify whether a high fibre diet fed to pigs during their oestrous cycle altered the composition of pFF. Additionally, the pFF of fertile animals was compared with the pFF of non-fertile animals to identify whether pFF composition was associated with fertility; fertile animals produced an embryo following in vitro fertilisation (IVF). Differences in the molecular composition were to be used to ascertain the potential underlying mechanism(s) involved in dietary induced improvements to oocyte maturation. Results The protein composition of pooled pFF from 12 HF-pigs and 12 C-pigs was compared by liquid chromatography tandem mass spectrometry (LC-MS/MS). Additionally, within each dietary group, the composition of pooled pFF from pigs whose oocytes produced blastocysts following in vitro fertilisation (C-Bl and HF-Bl) was compared with pFF from pigs whose oocytes did not produce blastocysts (C-No and HF-No respectively; n=6 per group). These proteomic analyses identified differentially expressed proteins, associated with several canonical pathways including acute phase response signalling, complement system and LXR/RXR activation, as determined by Ingenuity Pathway Analysis. Quantitative western blots revealed the differential expression of candidates associated with these canonical pathways. Plasminogen expression was lower (P≤0.05) in pFF of HF-pigs compared to pFF of C-pigs. In pFF from C-Bl gilts, apolipoprotein A4 (P≤0.01) and apolipoprotein M (P≤0.05) expression were higher compared to pFF from C-No gilts. Plasmin expression was lower (P≤0.05) in pFF from HF-Bl gilts compared to pFF from C-Bl gilts. Due to the interest in the differentially expressed apolipoproteins (involved in cholesterol and lipid efflux), a targeted metabolomic analysis was carried out to measure the concentration of nine fatty acids (FAs) in pFF of individual pigs in C-No, C-Bl, HF-No, HF-Bl groups (n=6 per group); adrenic, arachadonic, arachidic, dihomo- γ-linolenic, docosapentaenoic, erucic, linoleic, palmitoleic and oleic acids were measured by LC-MS/MS. The analysis revealed the lower concentration of linoleic acid (LA, p≤0.05) and higher concentration of erucic acid (P≤0.05) in HF-pFF compared to C-pFF. Following the results of the targeted metabolomic analysis, cumulus-oocytecomplexes (COCs) were matured in TCM 199 medium supplemented with 0 (No-LA), 50, 100 or 200 μM LA for 44 hours (n = 320 per treatment). COC diameters were measured and the COCs were categorised into "full", "partial" or "no" expansion. COCs were denuded, fixed and stained to determine their stage of maturation. IVM with 200 μM LA resulted in the reduced diameter of COCs (p≤0.01), fewer COCs with full cumulus expansion (p≤0.05) and fewer metaphase II oocytes (p≤0.05). Discussion Plasminogen is the precursor to plasmin, a proteolytic enzyme involved in weakening the follicular wall prior to ovulation. The lower expression of plasminogen and plasmin in pFF of high fibre pigs implies a delay in the accumulation of the inflammatory proteins required for ovulation. The delay in ovulation can result in the lengthening of the oocyte maturation process, leading to more mature oocytes, as observed in the previous studies. A disruption in the expression of apolipoproteins may also occur in high fibre-fed pigs. The increase in apolipoproteins associated with blastocyst development was only observed with pFF of control pigs but not high fibre pigs. An alteration in lipid homeostasis in the high fibre pigs could potentially affect oocyte energy consumption. LA concentration was also lower in pFF of high fibre pigs. LA is an essential fatty acid, indicating that the difference in concentration is directly from the diet. The lower levels of LA can potentially be beneficial to oocyte maturation, which is substantiated by the negative effects of a high LA concentration on IVM of abattoir derived oocytes.

La cicatrisation de défauts osseux permet tout au plus une réparation de l'os et dans peu de cas, une régénération ad integrum. Le développement de biomatériaux issus de l'ingénierie tissulaire en vue d'une régénération osseuse est donc un enjeu majeur. Le but de cette étude a été d'évaluer si des peptides GXXPG issus de l'élastine sont capables de favoriser la différenciation ostéoblastique de cellules mésenchymateuses dérivées de la moelle osseuse humaine (CMMO) ainsi que la formation de la matrice osseuse et sa minéralisation. Pour y répondre, nous avons utilisés les lattis de collagène de type I (COL1). La contraction de lattis « flottants » (LF) stimule l'expression de marqueurs de l'ostéoblaste (Runx-2, BSP…) par les CMMO ainsi que la minéralisation de la matrice osseuse. Cette différenciation ostéoblastique est aussi associée à l'activation de la cascade MT1-MMP/MMP-2/MMP-13. Nous montrons ensuite que les peptides GXXPG stimulent de façon dose-dépendante l'expression de marqueurs ostéoblastiques comme Runx-2 via S-Gal. Sur « coating » de COL1, ils stimulent la différenciation ostéoblastique des CMMO, la formation de la matrice osseuse et sa minéralisation. Enfin, dans des conditions « inflammatoires » créées par l'ajout de plasminogène (Plg) exogène, ces peptides conservent une activité ostéogénique sous contraintes mécaniques ou non. Plg seul induit également la différenciation ostéoblastique. Bien que les peptides GXXPG stimulent la production d'enzymes à activité collagénolytique (MT1-MMP, MMP-1), la lyse des LF n'est pas significative. En conclusion, les peptides GXXPG apparaissent comme des biomolécules pharmacologiques prometteuses pour la régénération osseuse. / Bone healing leads in only a few cases to an ad integrum regeneration, but most often to an incomplete tissue repair. Thus, the development of new biomaterials from tissue engineering in order to promote bone regeneration is a major goal. The purpose of our study was to evaluate if GXXPG peptides, derived from elastin, are able to favor human bone marrow mesenchymal cells (HBMC) to mature osteoblasts and bone matrix formation and mineralization.To this end, we used type I collagen (COL1) lattices. Floating lattice (LF) contraction stimulates osteoblasts markers expression (Runx-2, BSP…) by HMBC and bone matrix mineralization. Osteoblast differentiation is also associated to MT1-MMP/MMP-2/MMP-13 proteolytic cascade activation. We then showed that GXXPG peptides stimulate osteoblast markers like Runx-2 in a dose-dependent manner, an effect which involves S-Gal receptor. On a type I collagen coating model, these peptides also promote CMMO differentiation into osteoblast, bone matrix formation and mineralization. Finally, under “inflammatory” conditions, which can be catalyzed by plasminogen (Plg) supplementation, these peptides keep their ability to induce osteogenic responses in HBMC, even under mechanical stress. Plg alone is also able to promote osteoblast differentiation. Although GXXPG peptides stimulate collagenolytic enzymes (MT1-MMP, MMP-1) production, collagen degradation in LF is not significant. To conclude, GXXPG peptides appear as promising pharmacological biomolecules in bone regeneration.