Atherogenesis is associated with elevated levels of low-density lipoprotein (LDL) and its oxidized form (oxLDL) in the blood. The liver is an important scavenger organ for circulating oxLDLs. The present study aimed to examine endocytosis of mildly oxLDL (the major circulating form of oxLDLs) in liver sinusoidal endothelial cells (LSECs) and the involvement of the scavenger receptors stabilin-1 and stabilin-2 in this process. Freshly isolated LSECs, Kupffer cells (KCs), and stabilin-1- and stabilin-2-transfected human embryonic kidney cells were incubated with fluorescently labeled or radiolabeled oxLDLs [oxidized for 3 h (oxLDL(3)), 6 h, or 24 h (oxLDL(24))] to measure endocytosis. The intracellular localization of oxLDLs and stabilins in LSECs was examined by immunofluorescence and immunogold electron microscopy. Whereas oxLDL(24) was endocytosed both by LSECs and KCs, oxLDL(3) (mildly oxLDL) was taken up by LSECs only. The LSEC uptake of oxLDLs was significantly inhibited by the scavenger receptor ligand formaldehyde-treated serum albumin. Uptake of all modified LDLs was high in stabilin-1-transfected cells, whereas stabilin-2-transfected cells preferentially took up oxLDL(24), suggesting that stabilin-1 is a more important receptor for mildly oxLDLs than stabilin-2. Double immunogold labeling experiments in LSECs indicated interactions of stabilin-1 and stabilin-2 with oxLDL(3) on the cell surface, in coated pits, and endocytic vesicles. LSECs but not KCs endocytosed mildly oxLDL. Both stabilin-1 and stabilin-2 were involved in the LSEC endocytosis of oxLDLs, but experiments with stabilin-transfected cells pointed to stabilin-1 as the most important receptor for mildly oxLDL.

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Liver sinusoidal endothelial cells (LSECs) play an essential role in systemic waste clearance by effective endocytosis of blood-borne waste macromolecules. We aimed to study LSECs' scavenger function during aging, and whether age-related morphological changes (eg, defenestration) affect this function, in F344/BN F1 rats. Endocytosis of the scavenger receptor ligand formaldehyde-treated serum albumin was significantly reduced in LSECs from old rats. Ligand degradation, LSEC protein expression of the major scavenger receptors for formaldehyde-treated serum albumin endocytosis, stabilin-1 and stabilin-2, and their staining patterns along liver sinusoids, was similar at young and old age, suggesting that other parts of the endocytic machinery are affected by aging. Formaldehyde-treated serum albumin uptake per cell, and cell porosity evaluated by electron microscopy, was not correlated, indicating that LSEC defenestration is not linked to impaired endocytosis. We report a significantly reduced LSEC endocytic capacity at old age, which may be especially important in situations with increased circulatory waste loads.

The existence of a bone marrow (BM) niche--the location in which hematopoietic stem cells (HSCs) reside--was proposed more than 30 years ago. Recent data suggest that the interaction of HSCs with cellular and extracellular components within the BM is critical for HSC regulation. The tracking of immunofluorescently labeled, prospectively isolated HSCs to and within the BM cavity allows the assessment of the regulatory processes involved in (1) homing, which involves transendothelial migration into the BM; (2) lodgment, including transmarrow migration through the extravascular space; and (3) BM reconstitution. Together, such analyses provide a better understanding of the cellular and extracellular components involved in the regulation of HSC quiescence and differentiation. Homing and lodgment of transplanted HSCs, the first critical steps in engraftment, involve multiple interactions between HSCs and the BM microenvironment. Herein, we describe a refined method of analyzing homing efficiency and spatial distribution of HSCs harvested from endosteal and/or central BM regions; we also review alternate methods. Using these techniques, microenvironment modifications within the recipient or surface protein-expression modifications on donor HSCs in animal models provide insights into components influencing the homing, lodgment, and engraftment processes.

Background:Matrix metalloproteinase-8 (MMP-8) has oncosuppressive properties in various cancers. We attempted to assess MMP-8 function in oral tongue squamous cell carcinoma (OTSCC).Methods:MMP-8 overexpressing OTSCC cells were used to study the effect of MMP-8 on proliferation, apoptosis, migration, invasion and gene and protein expression. Moreover, MMP-8 functions were assessed in the orthotopic mouse tongue cancer model and by immunohistochemistry in patient samples.Results:MMP-8 reduced the invasion and migration of OTSCC cells and decreased the expression of MMP-1, cathepsin-K and vascular endothelial growth factor-C (VEGF-C). VEGF-C was induced by transforming growth factor-β1 (TGF-β1) in control cells, but not in MMP-8 overexpressing cells. In human OTSCC samples, low MMP-8 in combination with high VEGF-C was an independent predictor of poor cancer-specific survival. TGF-β1 treatment also restored the migration of MMP-8 overexpressing cells to the level of control cells. In mouse tongue cancer, MMP-8 did not inhibit metastasis, possibly because it was eliminated in the peripheral carcinoma cells.Conclusions:The suppressive effects of MMP-8 in OTSCC may be mediated through interference of TGF-β1 and VEGF-C function and altered proteinase expression. Together, low MMP-8 and high VEGF-C expression have strong independent prognostic value in OTSCC.

Factor V (FV) and factor X (FX) activate and complex to form prothrombinase which subsequently cleaves prothrombin (PT), converting it to active thrombin. Thrombin cleaved osteopontin (tcOPN) contains a cryptic binding site for a 4 b 1 and a 9 b 1 integrins. We have previously shown that hematopoietic stem cells (HSC) bind to tcOPN via this site resulting in a decrease in their proliferation and differentiation. Therefore, tcOPN and the factors required for its generation are important components of the HSC niche. Herein we show mature megakaryocytes (MM, 8N) contain FV, FX, and PT mRNA and protein. Furthermore, we show 8N, 16N, 32N, and 64N MM all release the required factors to enable thrombin cleavage of OPN. Importantly, mice devoid of the myeloproliferative leukemia protein (Mpl), c-Mpl 2/2 mice, contain only approximately 10% of normal megakaryocyte numbers, showed significantly reduced FX and tcOPN protein levels in endosteal bone marrow (BM). In addition, WT hematopoietic progenitors and HSC showed reduced homing to the BM of c-Mpl 2/2 mice. This is the first report identifying MM as a key cellular component in the production of tcOPN in situ, allowing the BM microenvironment to self regulate HSC biology via tcOPN.

Red blood cells squeeze through micro-capillaries as part of blood circulation in the body.

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