In this in vitro study, we have investigated the ability of Taraxacum officinale (dandelion) to inhibit adipocyte differentiation and lipogenesis in 3T3-L1 preadipocytes. HPLC analysis of the three plant extracts used in this study-leaf and root extracts and a commercial root powder-identified caffeic and chlorogenic acids as the main phenolic constituents. Oil Red O staining and triglyceride levels analysis showed decreased lipid and triglyceride accumulation, respectively. Cytotoxicity was assessed with the MTT assay showing non-toxic effect among the concentrations tested. DNA microarray analysis showed that the extracts regulated the expression of a number of genes and long non-coding RNAs that play a major role in the control of adipogenesis. Taken together, our results indicate that the dandelion extracts used in this study may play a significant role during adipogenesis and lipid metabolism, and thus, supporting their therapeutic interest as potential candidates for the treatment of obesity.
Adipose tissue dysfunction constitutes a primary defect in obesity and might link this disease to severe chronic health problems. We aimed to evaluate the antioxidant activity of three extracts from Taraxacum officinale (dandelion) as well as their effects on mature 3T3-L1 adipocytes concerning intracellular lipid accumulation and cytotoxicity, this would give indications regarding therapeutic interest of dandelion as potential anti-obesity candidate. Antioxidant activities of extracts from dandelion roots and leaves were evaluated in vitro using 1,1-diphenyl-2-picrylhyorazyl (DPPH) and Ferric Reducing Antioxidant Power (FRAP) methods at the concentration range used in cellular assays (300–600 µg/mL). The influence of the extracts on mature 3T3-L1 adipocyte viability was determined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Lipid content was determined by Oil-red-O staining. The extracts showed effective antioxidant activity correlating with total flavonoid and polyphenol contents. However, the functionality level was weakly associated with the antioxidant activity. Further, our data demonstrated that mature 3T3-L1 adipocytes reduced in size and number when incubated with the extracts, which suggests a significant increase in lipolysis activity. Particularly, leaf extract and crude powdered root of dandelion reduced triglyceride accumulation in mature 3T3-L1 adipocytes to a greater extent that the extract from the root. Our study shows anti-lipogenic effects of dandelion extracts on adipocytes as well as radical scavenging and reducing activity. Importantly, along with previous results indicating that cell populations cultivated in the presence of the dandelion extracts decrease in 3T3-L1 adipogenesis capacity, these results suggests that these extracts might represent a treatment option for obesity-related diseases by affecting different processes during the adipocyte life cycle.
Neonatal livers are a potential source of good-quality hepatocytes for clinical transplantation. We compared viability and function of neonatal hepatocytes (NHs) and adult hepatocytes (AHs) and report their clinical use both intraportally and in alginate microbeads. Following isolation from donor livers, hepatocyte function was assessed using albumin, alpha-1-antitrypsin, and factor VII. Metabolic function was investigated by measuring resorufin conjugation, ammonia metabolism, uridine diphosphate glucuronosyltransferase enzyme activity, and cytochrome P450 (CYP) function following induction. Activation of the instant blood-mediated inflammatory reaction by NHs and AHs was investigated using an in vitro blood perfusion model, and tissue factor expression was analyzed using real-time polymerase chain reaction (RT-PCR). Clinical hepatocyte transplantation (HT) was undertaken using standard protocols. Hepatocytes were isolated from 14 neonatal livers, with an average viability of 89.4% ± 1.8% (mean ± standard error of the mean) and average yield of 9.3 × 10 ± 2.0 × 10 cells/g. Hepatocytes were isolated from 14 adult livers with an average viability of 78.6% ± 2.4% and yield 2.2 × 10 ± 0.5 × 10 cells/g. NHs had significantly higher viability after cryopreservation than AHs, with better attachment efficiency and less plasma membrane leakage. There were no differences in albumin, alpha-1-antitrypsin, and factor VII synthesis between NHs and AHs (P > 0.05). Neonatal cells had inducible phase 1 enzymes as assessed by CYP function and functional phase 2 enzymes, in which activity was comparable to AHs. In an in vitro blood perfusion model, AHs elicited increased thrombus formation with a greater consumption of platelets and white cells compared with NHs (28.3 × 10 versus 118.7 × 10 and 3.3 × 10 versus 6.6 × 10 ; P < 0.01). Intraportal transplantation and intraperitoneal transplantation of alginate encapsulated hepatocytes was safe, and preliminary data suggest the cells may activate the immune response to a lesser degree than adult cells. In conclusion, we have shown NHs have excellent cell viability, function, and drug metabolism making them a suitable alternative source for clinical HT. Liver Transplantation 24 394-406 2018 AASLD.
Hepatocyte transplantation has emerged as an alternative to liver transplant for liver disease. Hepatocytes encapsulated in alginate microbeads have been proposed for the treatment of acute liver failure, as they are able to provide hepatic functions while the liver regenerates. Furthermore, they do not require immunosuppression, as the alginate protects the hepatocytes from the recipient's immune cells. Mesenchymal stromal cells are very attractive candidates for regenerative medicine, being able to differentiate into cells of the mesenchymal lineages and having extensive proliferative ability. When co-cultured with hepatocytes in two-dimensional cultures, they exert a trophic role, drastically improving hepatocytes survival and functions. In this study we aimed to (i) devise a high throughput system (HTS) to allow testing of a variety of different parameters for cell encapsulation and (ii) using this HTS, investigate whether mesenchymal stromal cells could have beneficial effects on the hepatocytes when co-encapsulated in alginate microbeads. Using our HTS platform, we observed some improvement of hepatocyte behavior with MSCs, subsequently confirmed in the low throughput analysis of cell function in alginate microbeads. Therefore, our study shows that mesenchymal stromal cells may be a good option to improve the function of hepatocytes microbeads. Furthermore, the platform developed may be used for HTS studies on cell encapsulation, in which several conditions (e.g., number of cells, combinations of cells, alginate modifications) could be easily compared at the same time.
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