An Optimized Extract, Named Self-Growth Colony, from Platelet-Rich Plasma Shows Robust Skin Rejuvenation and Anti-Ageing Properties: A Novel Technology in Development of Cosmetics.

INTRODUCTION: Inspired by application of platelet-rich plasma (PRP) in skin treatment during injuries, an extracting method was developed here to recover high amounts of cytokines and growth factors from PRP; this prepared extract was named as self-growth colony (SGC). METHODS: In optimization of SGC preparation, various parameters were tested, for example, centrifugation force, freeze-thaw, sonication, and inclusion of calcium chelator. The amounts of cytokines and growth factors, including platelet factor 4, ß-thromboglobulin, epidermal growth factor, vascular endothelial growth factor, platelet-derived growth factor, were measured by ELISA assay. RESULTS: By comparing to PRP, the prepared SGC contained a significant higher amount of measured growth factors. In addition, the degradation of growth factors within SGC during the storage was calibrated, which showed better stability as compared to that of PRP preparation. Having possible application in skin care, the optimized SGC was chemically standardized by using the enrichment of growth factors. Application of SGC in cultured keratinocytes stimulated the wound healing of injured cultures. In line to this notion, SGC was applied onto human skin, and thereafter the robust improvement of skin properties was revealed. CONCLUSIONS: The potential application of SGC in treating skin rejuvenation and ageing, as well as its elaborated application for medical purpose, that is, wound healing, was illustrated.

The binding of kaempferol-3-O-rutinoside to vascular endothelial growth factor potentiates anti-inflammatory efficiencies in lipopolysaccharide-treated mouse macrophage RAW264.7 cells.

BACKGROUND: Vascular Endothelial Growth Factors (VEGFs) are a group of growth factor in regulating development and maintenance of blood capillary. The VEGF family members include VEGF-A, placenta growth factor (PGF), VEGF-B, VEGF-C and VEGF-D. VEGF receptor activation leads to multiple complex signaling pathways, particularly in inducing angiogenesis. Besides, VEGF is produced by macrophages and T cells, which is playing roles in inflammation. In macrophages, VEGF receptor-3 (VEGFR-3) and its ligand VEGF-C are known to attenuate the release of pro-inflammatory cytokines. METHODS: Immunoprecipitation and molecular docking assays showed the binding interaction of kaempferol-3-O-rutinoside and VEGF-C. Western blotting and qRT-PCR methods were applied to explore the potentiating effect of kaempferol-3-O-rutinoside in VEGF-C-mediated expressions of proteins and genes in endothelial cells and LPS-induced macrophages. Enzyme-linked immunosorbent assay (ELISA) was employed to reveal the release of proinflammatory cytokines in LPS-induced macrophages. Immunofluorescence assay was performed to determine the effect of kaempferol-3-O-rutinoside in regulating nuclear translocation of NF-κB p65 subunit in the VEGF-C-treated cultures. In addition, Transwell® motility assay was applied to detect the ability of cell migration after drug treatment in LPS-induced macrophages. RESULTS: We identified kaempferol-3-O-rutinoside, a flavonoid commonly found in vegetable and fruit, was able to act on cultured macrophages in inhibiting inflammatory response, and the inhibition was mediated by its specific binding to VEGF-C. The kaempferol-3-O-rutinoside-bound VEGF-C showed high potency to trigger the receptor activation. In LPS-treated cultured macrophages, applied kaempferol-3-O-rutinoside potentiated inhibitory effects of exogenous applied VEGF-C on the secretions of pro-inflammatory cytokines, i.e. IL-6 and TNF-α, as well as expressions of nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). This inhibition was in parallel to transcription and translocation of NF-κB. Moreover, the binding of kaempferol-3-O-rutinoside with VEGF-C suppressed the LPS-induced migration of macrophage. CONCLUSION: Taken together, our results suggested the pharmacological roles of kaempferol-3-O-rutinoside in VEGF-C-mediated anti-inflammation.

Piceatannol, a Natural Analog of Resveratrol, Exerts Anti-angiogenic Efficiencies by Blockage of Vascular Endothelial Growth Factor Binding to Its Receptor.

Piceatannol is also named as trans-3,4,3',5'-tetrahydroxy-stilbene, which is a natural analog of resveratrol and a polyphenol existing in red wine, grape and sugar cane. Piceatannol has been proved to possess activities of immunomodulatory, anti-inflammatory, antiproliferative and anticancer. However, the effect of piceatannol on VEGF-mediated angiogenesis is not known. Here, the inhibitory effects of piceatannol on VEGF-induced angiogenesis were tested both in vitro and in vivo models of angiogenesis. In human umbilical vein endothelial cells (HUVECs), piceatannol markedly reduced the VEGF-induced cell proliferation, migration, invasion, as well as tube formation without affecting cell viability. Furthermore, piceatannol significantly inhibited the formation of subintestinal vessel in zebrafish embryos in vivo. In addition, we identified the underlying mechanism of piceatannol in triggering the anti-angiogenic functions. Piceatannol was proposed to bind with VEGF, thus attenuating VEGF in activating VEGF receptor and blocking VEGF-mediated downstream signaling, including expressions of phosphorylated eNOS, Erk and Akt. Furthermore, piceatannol visibly suppressed ROS formation, as triggered by VEGF. Moreover, we further determined the outcome of piceatannol binding to VEGF in cancer cells: piceatannol significantly suppressed VEGF-induced colon cancer proliferation and migration. Thus, these lines of evidence supported the conclusion that piceatannol could down regulate the VEGF-mediated angiogenic functions with no cytotoxicity via decreasing the amount of VEGF binding to its receptors, thus affecting the related downstream signaling. Piceatannol may be developed into therapeutic agents or health products to reduce the high incidence of angiogenesis-related diseases.

Kaempferol, a Major Flavonoid in Ginkgo Folium, Potentiates Angiogenic Functions in Cultured Endothelial Cells by Binding to Vascular Endothelial Growth Factor.

Kaempferol is a major flavonoid in Ginkgo Folium and other edible plants, which is being proposed here to have roles in angiogenesis. Angiogenesis is important in both physiological and pathological development. Here, kaempferol was shown to bind with vascular endothelial growth factor (VEGF), probably in the heparin binding domain of VEGF: this binding potentiated the angiogenic functions of VEGF in various culture models. Kaempferol potentiated the VEGF-induced cell motility in human umbilical vein endothelial cells (HUVECs), as well as the sub-intestinal vessel sprouting in zebrafish embryos and formation of microvascular in rat aortic ring. In cultured HUVECs, application of kaempferol strongly potentiated the VEGF-induced phosphorylations of VEGFR2, endothelial nitric oxide synthase (eNOS) and extracellular signal-regulated kinase (Erk) in time-dependent and concentration-dependent manners, and in parallel the VEGF-mediated expressions of matrix metalloproteinases (MMPs), MMP-2 and MMP-9, were significantly enhanced. In addition, the potentiation effect of kaempferol was revealed in VEGF-induced migration of skin cell and monocyte. Taken together, our results suggested the pharmacological roles of kaempferol in potentiating VEGF-mediated functions should be considered.

Synthesis and preliminary evaluation of n.c.a. iodoquine: a novel radiotracer with high uptake in cells with high ALDH1 expression.

PURPOSE: Chloroquine has demonstrated high affinity for aldehyde dehydrogenase 1A1 (ALDH1), an enzyme expressed in the highly tumorigenic CD133+ brain tumor initiating subpopulation. The purpose of this study is to report the novel synthesis of a chloroquine analogue, n.c.a. iodoquine, and the in vitro and in vivo uptake in cells with high ALDH1 content. METHODS AND MATERIALS: Iodoquine was synthesized in novel no-carrier-added forms (n.c.a.) for both 125I and 123I. I25I IQ and 18F FDG cell uptake assays were performed in the L1210 and L1210cpa (cyclophosphamide resistant), A549, and MG456 glioblastoma cell lines. Uptake was expressed as a percent of the administered activity. 125I IQ biodistribution studies assessed organ uptake at 1, 4, and 24 hours after IV administration (n= 15 total; 5 mice/timepoint). Radiation dosimetry estimates were calculated using standard OLINDA/EXM software. In vivo imaging of 123I IQ uptake in MG456 glioblastoma mouse model (n=10) was performed with small animal high resolution micro-SPECT. Autoradiography and histology co-localized radiotracer and tumor biodistribution. Uptake in MG456 glioblastoma tumors was quantified with gamma counting. RESULTS: L1210 cpa (high ALDH1) showed significantly higher 125I IQ uptake compared to the parental L1210 (low ALDH1) for all time points through 4 hours (20.7% ± 1.4% versus 11.0% ± 0.5%; 21.3% ± 0.9% versus 11.0% ± 0.4%; 20.6% ± 0.7% versus 9.4% ± 0.3%; and 15.7% ± 0.7% versus 7.5% + 0.4% at 30 minutes, and 1, 2 and 4 hours, respectively; p < 0.001 for all time points). In the CD133+ fraction of MG456 glioblastoma cell line, IQ uptake was significantly higher compared to FDG at all time points through 4 hours (81.5% ± 0.9% versus 1.3% ± 0.1%; 88.8% ± 0.4% versus 1.3% ± 0.1%; 87.8% ± 2.1% versus 1.7% ± 0.2%; and 87.0% ± 2.4% versus 1.8% ± 0.1 at 30 minutes, and 1, 2 and 4 hours, respectively; p > 0.001 for all time points). The A549 lung cancer cell line also showed high IQ uptake through 4 hours. IQ normal biodistribution studies showed rapid renal excretion and very low normal background brain activity after IV administration. In vivo micro-SPECT images showed mild uptake in larger MG456 glioblastomas (n=6) as verified with autoradiography and histology. Gamma well counter uptake in large tumors was 2.3% ± 0.48% ID/g (n=5). CONCLUSION: Iodoquine localizes to cells with high ALDH1 content. Cell assays show high 125I IQ uptake in the MG456 cell line, and in vivo micro-SPECT imaging showed mild 123I IQ uptake in MG456 glioblastomas. Further studies are necessary to investigate 131I IQ as a potential therapeutic agent targeting the highly tumorigenic CD133+ brain tumor stem cell subpopulation.

Detection of human CYP2C8, CYP2C9, and CYP2J2 in cardiovascular tissues.

The cytochrome P450 (P450) enzymes CYP2C8, CYP2C9, and CYP2J2 metabolize arachidonic acid to epoxyeicosatrienoic acids, which are known to be vital in regulation of vascular tone and cardiovascular homeostasis. Because there is limited information regarding the relative expression of these P450 enzymes in cardiovascular tissues, this study examined the expression of CYP2C8, CYP2C9, and CYP2J2 mRNA and protein in human heart, aorta, and coronary artery samples by real-time polymerase chain reaction, immunoblotting, and immunohistochemistry. CYP2J2 and CYP2C9 mRNA levels were highly variable in human hearts, whereas CYP2C8 mRNA was present in lower abundance. CYP2J2 mRNA was approximately 10(3) times higher than CYP2C9 or CYP2C8 in human heart. However, CYP2C9 mRNA was more abundant than CYP2J2 or CYP2C8 in one ischemic heart. In human aorta, mean CYP2C9 mRNA levels were approximately 50 times higher than that of CYP2J2 and 5-fold higher than that of CYP2C8. In human coronary artery, mean values for CYP2C9 mRNA were approximately 2-fold higher than that of CYP2J2 mRNA and 6-fold higher than that of CYP2C8 mRNA. Immunoblotting results show relatively high levels of CYP2J2 and CYP2C8 protein in human hearts, which was confirmed by immunohistochemistry. CYP2C9 protein was also detected at high levels in one ischemic heart by immunoblotting. CYP2C9 was present at higher levels than CYPJ2 in aorta and coronary artery, whereas CYP2C8 protein was below the limits of detection. The expression of CYP2J2 and CYP2C8 in human heart, and CYPC9 and CYP2J2 in aorta and coronary artery is consistent with a physiological role for these enzymes in these tissues.