RESUMO
Lactoferrin (LF), known to be present in mammalian milk, has been reported to promote the proliferation of osteoblasts and suppress bone resorption by affecting osteoclasts. However, the mechanisms underlying the effects of human sources LF on osteoblast differentiation have not yet been elucidated, and almost studies have used LF from bovine sources. The presented study aimed to characterize the molecular mechanisms of bovine lactoferrin (IF-I) and human recombinant lactoferrin (LF-II) on MC3T3-E1 pre-osteoblast cells. MC3T3-E1 cells were treated with LF, ascorbic acid, and ß-glycerophosphate (ß-GP). Cell proliferation was analyzed using the MTT assay. Alkaline phosphatase activation and osteopontin expression levels were evaluated via cell staining and immunocytochemistry. The differentiation markers were examined using quantitative real-time PCR. The cell viability assay showed the treatment of 100 µg/mL LF significantly increased; however, it was suppressed by the simultaneous treatment of ascorbic acid and ß-GP. Alizarin red staining showed that the 100 µg/mL treatment of LF enhanced calcification. Quantitative real-time PCR showed a significant increase in osterix expression. The results suggest that treatment with both LFs enhanced MC3T3-E1 cell differentiation and promoted calcification. The mechanisms of calcification suggest that LFs are affected by an increase in osterix and osteocalcin mRNA levels.
RESUMO
PURPOSE: Mohs' paste, which is composed of zinc chloride and zinc oxide starch, is used for hemostasis of superficial malignancy in the clinical setting. We investigated the concentration of intramuscular zinc in mice after Mohs' paste application and evaluated its relationship with angiogenesis from the perspective of blood flow levels within 24 h. METHODS: Male C57BL/6JJmsSlc mice were administered single dose of Mohs' paste at 25%, 50%, and 75% after unilateral hind limb surgery, and glycerin, a viscosity modifier, was administered to the control group (0%). Hind limb blood flow levels were measured with a laser Doppler perfusion imaging system (n = 6). The amounts of intramuscular zinc and vascular endothelial growth factor-A (VEGF-A) expression were analyzed using inductively coupled plasma mass spectrometry (ICP-MS) and western blotting, respectively (n = 5 or 3). RESULTS: Blood flow levels were significantly decreased in the 50% group after 8 h, and significantly decreased in the 25% and 50% groups after 24 h. Intramuscular zinc was significantly increased in the 50% and 75% groups after 8 h. Western blotting showed that VEGF-A levels were significantly increased in the 25% and 50% groups after 8 h. Based on analytical experiments and biological investigation, we predicated the pharmacological effect of Mohs' paste and found over 50% of it is critical in the blood flow and angiogenesis suppression after more than 8 h of its application. CONCLUSIONS: The results suggest that the mechanism of blood flow suppression is independent of VEGF-A levels and might suppress future angiogenesis. Our findings support that of previous studies, in which Mohs' paste was expected to induce hemostasis and suppress angiogenesis. It is an excellent ointment that facilitates hemostasis by suppressing blood flow regardless of angiogenesis, and may be apt for situations where hemostasis is required in the clinical setting.
