Human Amniotic Epithelial Cells as a Tool to Investigate the Effects of Cyanidin 3-O-Glucoside on Cell Differentiation.
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ABSTRACT:
SUBMITTER: Takahashi S
PROVIDER: S-EPMC8038597 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:AimTo explore the feasibility that human amniotic epithelial cells (hAECs) have the potential to differentiate into corneal epithelial-like cells under the microenvironment replicated by spontaneously immortalized human corneal epithelial cells (S-ihCECs).MethodshAECs were isolated by enzyme digestion, and flow cytometry was used to analysis the expression of CD29/90/166/73/34 and HLA-DR. Recovered and cultured S-ihCECs, immunocytochemistry was used to detect the expression of CK3/12. The proliferation of S-ihCECs handled by different concentrations of mitomycin was detected by CCK-8. The proliferation of hAECs cultured by S-ihCECs culture media collected at different time was analyzed by CCK-8. After filtered out the optimal conditions, we collected S-ihCECs culture media for 5 days, then prepared conditioned medium to incubate hAECs, inverted phase contrast microscope and scanning electron microscope were used to observe the change of morphology in hAECs. Quantitative real-time reverse transcription-polymerase chain reaction (QRT-PCR) was carried out to evaluate the expression of Oct-4, NANOG, PAX6, and CK12 in the differentiation period. Immunocytochemistry and western bloting were used to detect the expression of CK3/12.ResultsThe culture media collected every 12h, from 20µg/mL mitomycin pretreatment S-ihCECs could significantly promote the proliferation of hAECs. In the period of differentiation, the morphology of differentiated hAECs was obviously different compared with the control group, and the distinctive CK3/12 for corneal epithelial cells was detected.ConclusionThis study showed that hAECs can differentiate into corneal epithelial-like cells by in vitro replication of the corneal epithelial microenvironment, using the culture media collected from S-ihCECs, and it is possible that S-ihCECs culture media could be used in corneal tissue engineering.
| S-EPMC3808898 | biostudies-literature
Project description:UNLABELLED:Hepatocyte transplantation to treat liver disease is largely limited by the availability of useful cells. Human amniotic epithelial cells (hAECs) from term placenta express surface markers and gene characteristics of embryonic stem cells and have the ability to differentiate into all three germ layers, including tissues of endodermal origin (i.e., liver). Thus, hAECs could provide a source of stem cell-derived hepatocytes for transplantation. We investigated the differentiation of hAECs in vitro and after transplantation into the livers of severe combined immunodeficient (SCID)/beige mice. Moreover, we tested the ability of rat amniotic epithelial cells (rAECs) to replicate and differentiate upon transplantation into a syngenic model of liver repopulation. In vitro results indicate that the presence of extracellular matrix proteins together with a mixture of growth factors, cytokines, and hormones are required for differentiation of hAECs into hepatocyte-like cells. Differentiated hAECs expressed hepatocyte markers at levels comparable to those of fetal hepatocytes. They were able to metabolize ammonia, testosterone, and 17?-hydroxyprogesterone caproate, and expressed inducible fetal cytochromes. After transplantation into the liver of retrorsine (RS)-treated SCID/beige mice, naïve hAECs differentiated into hepatocyte-like cells that expressed mature liver genes such as cytochromes, plasma proteins, transporters, and other hepatic enzymes at levels equal to adult liver tissue. When transplanted in a syngenic animal pretreated with RS, rAECs were able to engraft and generate a progeny of cells with morphology and protein expression typical of mature hepatocytes. CONCLUSION:Amniotic epithelial cells possess the ability to differentiate into cells with characteristics of functional hepatocytes both in vitro and in vivo, thus representing a useful and noncontroversial source of cells for transplantation.
| S-EPMC3103747 | biostudies-literature
Project description:Human amniotic epithelial cells (hAECs) represent an interesting clinical alternative to human embryonic (hESCs) and induced pluripotent (hiPSCs) stem cells in regenerative medicine. The potential of hAECs can be enhanced ex vivo by their partial pre-differentiation. The aim of this study was to evaluate the effectiveness of 18-day differentiation of hAECs into endodermal cells, hepatic precursor cells, and cells showing functional features of hepatocytes using culture media supplemented with high (100 ng/mL) concentrations of EGF or HGF. The cells obtained after differentiation showed changes in morphology and increased expression of AFP, ALB, CYP3A4, CYP3A7, and GSTP1 genes. HGF was more effective than EGF in increasing the expression of liver-specific genes in hAECs. However, EGF stimulated the differentiation process more efficiently and yielded more hepatocyte-like cells capable of synthesizing α-fetoprotein during differentiation. Additionally, after 18 days, GST transferases, albumin, and CYP P450s, which proved their partial functionality, were expressed. In summary, HGF and EGF at a dose of 100 ng/mL can be successfully used to obtain hepatocyte-like cells between days 7 and 18 of hAEC differentiation. However, the effectiveness of this process is lower compared with hiPSC differentiation; therefore, optimization of the composition of the medium requires further research.
| S-EPMC9317663 | biostudies-literature
Project description:Anthocyanins (ACNs) are plant secondary metabolites from the flavonoid family. Red to blue fruits are major dietary sources of ACNs (up to 1 g/100 g FW), being cyanidin-3-O-glucoside (Cy3G) one of the most widely distributed. Cy3G confers a red hue to fruits, but its content in raspberries and strawberries is low. It has a good radical scavenging capacity (RSC) against superoxide but not hydroxyl radicals, and its oxidative potential is pH-dependent (58 mV/pH unit). After intake, Cy3G can be metabolized (phases I, II) by oral epithelial cells, absorbed by the gastric epithelium (1%-10%) and it is gut-transformed (phase II & microbial metabolism), reaching the bloodstream (<1%) and urine (about 0.02%) in low amounts. In humans and Caco-2 cells, Cy3G's major metabolites are protocatechuic acid and phloroglucinaldehyde which are also subjected to entero-hepatic recycling, although caffeic acid and peonidin-3-glucoside seem to be strictly produced in the large bowel and renal tissues. Solid evidence supports Cy3G's bioactivity as DNA-RSC, gastro protective, anti-inflammatory, anti-thrombotic chemo-preventive and as an epigenetic factor, exerting protection against Helicobacter pylori infection, age-related diseases, type 2 diabetes, cardiovascular disease, metabolic syndrome and oral cancer. Most relevant mechanisms include RSC, epigenetic action, competitive protein-binding and enzyme inhibition. These and other novel aspects on Cy3G's physical-chemistry, foodomics, and health effects are discussed.
| S-EPMC6273591 | biostudies-literature
Project description:Osteoporosis, characterized by a gradual decrease in the number of osteoblasts and a gradual increase in bone resorption of osteoclasts in bone tissue, is a global chronic disease, which severely impairs the quality of life of the elderly. Therefore, it is extremely urgent to study the prevention and treatment of osteoporosis. It has been reported that anthocyanins can regulate bone metabolism and prevent osteoporosis. Cyanidin-3-O-glucoside (C3G), the most common type of anthocyanin in nature, widely exists in a variety of vegetables and fruits. Although it has been shown that C3G has multiple effects on osteoclasts, its impact(s) and underlying mechanism(s) on osteoblasts are still not clear. Here, we evaluated the effect of C3G on cell proliferation and differentiation of osteoblasts (extracted from the hip joint of patients with osteoporosis) and MC3T3-E1 (a kind of osteoblast cell line from mice). We also test the ability of osteoblasts to mineralize after C3G treatment. To find the underlying mechanism of the above effects, we further evaluated the role of the ERK signaling pathway in C3G regulation of osteoblasts. The results showed that C3G treatment enhanced osteoblast proliferation rate, osteoblast mineralization points, the mRNA levels and protein expression levels of OC (osteocalcin), and the level of ERK phosphorylation, which could be blocked by pretreatment with ERK signaling pathway inhibitor. The above results not only indicate that the ERK pathway was involved in C3G regulation of osteoblast differentiation but also provide strong suggestive evidence that osteoblasts may be promising targets in preventive and therapeutic strategies for osteoporosis.
| S-EPMC7905819 | biostudies-literature
Project description:BackgroundCD38+ NK cells are overabundant in rheumatoid arthritis (RA). Cyanidin-3-O-glucoside (C3G) is an inhibitor of CD38. This study investigated the pathogenic role of CD38+ NK cells and the effect of C3G on RA.MethodsRats with bovine type II collagen-induced arthritis (CIA) were injected with C3G. RA synovial fibroblasts (RASFs) or mononuclear cells (MNCs) were cultured with C3G. MNCs were also cocultured with CD38+ NK cells following C3G pretreatment.ResultsC3G injection significantly alleviated CIA. C3G also significantly increased the level of interleukin (IL)-10 and the regulatory T (Treg) cell proportion, and it decreased the interleukin (IL)-6 and interferon (IFN)-? levels and CD38+ NK cell proportion in rat peripheral blood and synovial fluid. Additionally, C3G significantly increased RASF apoptosis and decreased RASF proliferation and IL-6 production in the culture medium. Furthermore, C3G stimulated MNCs to increase IL-2 and IL-10 production and the Treg cell proportion, and it caused MNCs to decrease IL-6 and IFN-? production and the CD38+ NK cell proportion. Although CD38+ NK cells significantly decreased the Treg cell proportion and IL-10 level in MNCs, CD38+ NK cells that had been pretreated with C3G increased the proportion of Treg cells and IL-10 levels and decreased the IL-6 and IFN-? levels in the coculture. In CD38+ NK cells, C3G significantly increased Sirtuin 6 (Sirt6) expression and the tumor necrosis factor (TNF)-? level, and it decreased natural killer group 2D (NKG2D) expression and the IFN-? level. However, when CD38+ NK cells were treated with Sirt6 siRNA, C3G did not change the NKG2D expression, the TNF-? level sharply decreased, and the IFN-? level increased. When MNCs were cocultured with C3G-pretreated CD38+ NK cells in the presence of TNF-? and an anti-IFN-? antibody, the IL-10+ Treg cell proportion significantly increased. When MNCs were cocultured with C3G-pretreated CD38+ NK cells in the presence of IFN-? and an anti-TNF-? antibody, the IL-10+ Treg cell proportion sharply decreased. When CIA rats were injected with both C3G and the Sirt6 inhibitor OSS_128167, the rats exhibited joint inflammation and a low Treg cell proportion, but the CD38+ NK proportion was still low.ConclusionC3G has therapeutic effects on CIA and RA. C3G decreased the proportion of CD38+ cells, RASF proliferation, and proinflammatory cytokine secretion, and it increased the Treg cell proportion. C3G also elevated Sirt6 expression to suppress NKG2D expression, increase TNF-? secretion, and decrease IFN-? secretion in CD38+ NK cells, which stimulates MNCs to differentiate into Treg cells. This study also demonstrates that the inhibition of Treg cell differentiation in MNCs by CD38+ NK cells is a potential cause of the immune imbalance in RA and CIA.
| S-EPMC6819496 | biostudies-literature
Project description:The present study investigated the detailed in vitro osteogenic differentiation process and in vivo bone regenerative property of human amniotic epithelial cells (hAECs). The in vitro osteogenic differentiation process of hAECs was evaluated by biochemical staining, real-time polymerase chain reaction, and immunofluorescence. Next, β-tricalcium phosphate (β-TCP) scaffolds alone or loaded with hAECs were implanted into the alveolar defects of rats. Micro-computed tomography evaluation and histologic studies were conducted. Our results validated the in vitro osteogenic capacity of hAECs by upregulation of Runx2, osterix, alkaline phosphatase, collagen I, and osteopontin, with positive biochemical staining for osteoblasts. An epithelial-mesenchymal transformation process might be involved in the osteogenic differentiation of hAECs by increased expression of transforming growth factor-β1. Our data also demonstrated that in vivo implantation of hAECs loaded on β-TCP scaffolds, not only improved bone regeneration by direct participation, but also reduced the early host immune response to the scaffolds. The presented data indicate that hAECs possess proper osteogenic differentiation potential and a modulatory influence on the early tissue remodeling process, making these cells a potential source of progenitor cells for clinical restoration of the alveolar defect.
| S-EPMC4250213 | biostudies-literature
Project description:The objective was to assess anti-progression and stimulatory immune response effects among anthocyanins (ANC) and their metabolites on human colorectal cancer cells in vitro and in silico. Pure phenolics including delphinidin-3-O-glucoside (D3G) and its metabolites, delphinidin (DC) and gallic acid (GA), were tested alone or in combination, on HCT-116 and HT-29 human colorectal cancer cells (100-600 µg/mL). HCT-116 and HT-29 50% inhibition concentrations (µg/mL) were 396 ± 23 and 329 ± 17 for D3G; 242 ± 16 and >600 for DC; and 154 ± 5 and 81 ± 5 for GA, respectively. Using molecular docking, cyanidin-3-O-glucoside (C3G) showed the highest potential to inhibit immune checkpoints: programmed cell death protein-1 (PD-1) (-6.8 kcal/mol) and programmed death-ligand-1 (PD-L1) (-9.6 kcal/mol). C3G, D3G, DC, GA, and D3G-rich extracts decreased PD-L1 protein expression in HCT-116 cells. C3G decreased PD-L1 fluorescence intensity by 39%. ANC decreased PD-1 expression in peripheral blood mononuclear cells in monoculture by 41% and 55%, and co-culture with HCT-116 and HT-29 cells by 39% and 26% (C3G) and 50% and 51% (D3G), respectively. D3G and C3G, abundant in plant foods, showed potential for binding with and inhibiting immune checkpoints, PD-1 and PD-L1, which can activate immune response in the tumor microenvironment and induce cancer cell death.
| S-EPMC6689002 | biostudies-literature
Project description:Human amniotic epithelial cells (hAECs) are attractive candidates for regenerative medical therapy, with the potential to replace deficient cells and improve functional recovery after injury. Previous studies have demonstrated that transplantation of hAECs effectively alleviate chemotherapy-induced ovarian damage via inhibiting granulose cells apoptosis in animal models of premature ovarian failure/insufficiency (POF/POI). However, the underlying molecular mechanism accounting for hAECs-mediated ovarian function recovery is not fully understood.To investigate whether hAECs-secreting cytokines act as molecular basis to attenuate chemotherapy-induced ovarian injury, hAECs or hAEC-conditioned medium (hAEC-CM) was injected into the unilateral ovary of POF/POI mouse. Follicle development was evaluated by H&E staining at 1, 2 months after hAECs or hAEC-CM treatment. In addition, we performed a cytokine array containing 507 human cytokines on hAECs-derived serum-free conditioned medium. Finally, we further investigated whether hAECs could affect chemotherapy-induced apoptosis in primary human granulosa-lutein (hGL) cells and the tube formation of human umbilical vein endothelial cells (hUVECs) via a co-culture system in vitro.We observed the existence of healthy and mature follicles in ovaries treated with hAECs or hAEC-CM, whereas seriously fibrosis and many atretic follicles were found in the contralateral untreated ovaries of the same mouse. To distinguish cytokines involved in the process of hAECs-restored ovarian function, hAEC-CM was analyzed with a human cytokines array. Results revealed that 109 cytokines in hAEC-CM might participate in a variety of biological processes including apoptosis, angiogenesis, cell cycle and immune response. In vitro experiments, hAECs significantly inhibited chemotherapy-induced apoptosis and activated TGF-?/Smad signaling pathway within primary granulosa-lutein cells in paracrine manner. Furthermore, hAEC-CM was shown to promote angiogenesis in the injured ovaries and enhance the tube formation of human umbilical vein endothelial cells (hUVECs) in co-culture system.These findings demonstrated that paracrine might be a key pathway in the process of hAECs-mediating ovarian function recovery in animal models of premature ovarian failure/insufficiency (POF/POI).
| S-EPMC5704397 | biostudies-literature
Project description:Mitochondrial dysfunction has been identified as one of the primary factors contributing to liver diseases. Pathways that control mitochondrial biogenesis are potential therapeutic targets for the amelioration of hepatocyte dysfunction and liver disease. Research on natural pharmacological agents that ameliorate liver diseases has intensified over the last two decades. Cyanidin-3-glucoside (Cy3g), a dietary flavonoid compound extracted from a wide variety of fruits and vegetables, reportedly has several beneficial health effects. In this study, we used an adult human hepatoma cell line (HuH7) to investigate the effects of the Cy3g polyphenolic compound on mitochondrial function and biogenesis in vitro. An increase in intracellular mitochondrial reductase levels was observed after treatment with Cy3g, but cytotoxicity was not induced. In addition, mitochondrial membrane potential and ATP production were increased following Cy3g treatment. Cy3g treatment also resulted in a dose- and time-dependent upregulation of the gene expression of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1?), a transcription factor considered a master regulator of mitochondrial biogenesis and metabolism. Additionally, the expression of sirtuin 1 (SIRT1), which plays a key role in deacetylating PGC-1?, was also increased in a dose- and time-dependent manner. Cy3g treatment also increased the expression of downstream PGC-1? genes, nuclear respiratory factor 1 and mitochondrial transcription factor A (TFAM). Our results suggest that Cy3g has potential as a hepatoprotective therapeutic agent that enhances mitochondrial function and biogenesis in hepatocytes.
| S-EPMC6269359 | biostudies-literature