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Phase-contrast micrographs and gene expression profiles of human cryopreserved hepatocytes 7 days after the start of culture. Hepatocytes were cultured with 1-30 µmol/L (10 µmol/L in quantitative PCR experiment) of myricetin as indicated in the lower margins. (a-e) The morphologies of human hepatocytes, lot. FLO. Scale bar: 500 µm (f-j) those of lot. EJW. Scale bar: 100 µm. (k-s) Quantitative PCR profiles of parenchymal hepatocyte specific genes. Samples were taken from EJW at culture day 7. ALB, albumin; CYP3A4, cytochrome P450 3A4; AAT, alpha1-antitrypsin; TAT, tyrosine aminotransferase; TDO2, tryptophan 2,3-dioxygenase; HNF4, hepatocyte nuclear factor-4 alpha; CK18, cytokeratin 18; CPS1, carbamoylphosphate synthetase I; OTC, ornithine transcarbamylase. The primers used are described in Supplementary Materials, Table S1. Asterisks indicate statistically significant results (* p < 0.05) compared to controls using one-way ANOVA and Fisher's protected least significant difference (PLSD) test.
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To improve the therapeutic potential of hepatocyte transplantation, the effects of the mitogen-activated protein kinase kinase 4 (MKK4) inhibitor, myricetin (3,3′,4′,5,5′,7-hexahydroxylflavone) were examined using porcine and human hepatocytes in vitro and in vivo. Hepatocytes were cultured, showing the typical morphology of hepatic parenchymal cel...
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Context 1
... human hepatocytes (lot. EJW and FLO) were cultured with 1-30 µmol/L myricetin (Figure 2a-j). The phase-contrast micrographs on culture day 7 showed the cells treated with 1 and 3 µmol/L of myricetin in both lots. ...
Context 2
... to porcine hepatocytes, human cells cultured at higher concentrations (10 and 30 µmol/L of myricetin) appeared to be attenuated, and the cell layer looked sparse. In terms of the functions of hepatic differentiation, nine typical genes were assayed (Figure 2k-s). Hepatocytes treated with 1 µmol/L and 3 µmol/L of myricetin showed a significant increase in the expression of all genes, including carbamoylphosphate synthetase I (r) and ornithine transcarbamylase (s), the key enzymes in the ammonia metabolizing urea cycle. ...
Citations
... [19] PHH are typically frozen in University of Wisconsin solution with 10%-15% DMSO and 5% glucose. [19] Recently, modifications have been reported to improve the quality of cryopreserved PHH, such as encapsulation of hepatocytes, [39] alternative cryopreservation media to University of Wisconsin solution, [40] the addition of membrane stabilizer [41] and antioxidative chemicals (myricetin), [42] and bulk droplet vitrification. [43] Despite these improvements, the variability between different batches of PHH remains a major challenge. ...
Numerous studies have shown that hepatocyte transplantation is a promising approach for liver diseases, such as liver-based metabolic diseases and acute liver failure. However, it lacks strong evidence to support the long-term therapeutic effects of hepatocyte transplantation in clinical practice. Currently, major hurdles include availability of quality-assured hepatocytes, efficient engraftment and repopulation, and effective immunosuppressive regimens. Notably, cell sources have been advanced recently by expanding primary human hepatocytes by means of dedifferentiation in vitro. Moreover, the transplantation efficiency was remarkably improved by the established preparative hepatic irradiation in combination with hepatic mitogenic stimuli regimens. Finally, immunosuppression drugs, including glucocorticoid and inhibitors for co-stimulating signals of T cell activation, were proposed to prevent innate and adaptive immune rejection of allografted hepatocytes. Despite remarkable progress, further studies are required to improve in vitro cell expansion technology, develop clinically feasible preconditioning regimens, and further optimize immunosuppression regimens or establish ex vivo gene correction-based autologous hepatocyte transplantation.
... [19] PHH are typically frozen in University of Wisconsin solution with 10%-15% DMSO and 5% glucose. [19] Recently, modifications have been reported to improve the quality of cryopreserved PHH, such as encapsulation of hepatocytes, [39] alternative cryopreservation media to University of Wisconsin solution, [40] the addition of membrane stabilizer [41] and antioxidative chemicals (myricetin), [42] and bulk droplet vitrification. [43] Despite these improvements, the variability between different batches of PHH remains a major challenge. ...
Numerous studies have shown that hepatocyte transplantation (HTx) is a promising approach for liver diseases, such as liver-based metabolic diseases and acute liver failure. However, it lacks strong evidence to support the long-term therapeutic effects of HTx in clinical practice. Currently, major hurdles include availability of quality-assured hepatocytes, efficient engraftment and repopulation, and effective immunosuppressive regimens. Notably, cell sources have been advanced by recent technologies to expand primary human hepatocytes via dedifferentiation in vitro. Moreover, the transplantation efficiency was remarkably improved by the established preparative hepatic irradiation in combination with hepatic mitogenic stimuli regimens. Furthermore, immunosuppression drugs, including glucocorticoid and inhibitors for co-stimulating signals of T cell activation, were proposed to prevent innate and adaptive immune rejection of allografted hepatocytes. Despite remarkable progresses, further studies are required to improve in vitro cell expansion technology, develop clinically feasible preconditioning regimens, and further optimize immunosuppression regimens or establish ex vivo gene correction-based autologous HTx.
... 20) It is also a powerful radiosensitizer that enhances the radiotherapeutic-suppression of NSCLC cell lines. 21) Moreover, studies have reported that myricetin has an inhibitory impact on the liver cancer cells and conversely harbors a protective effect on the normal liver cells, 15,22) suggesting the selective activity of myricetin against tumor cells. ...
Lung cancer is the leading cause of cancer-related deaths worldwide, synthesizing and screening of novel anti-cancer drugs provides an alternative therapeutic strategy for renewal of the chemotherapy regimens against lung cancer. To this end, several compounds were synthesized based on the modification of the original myricetin, and their anti-tumor activity against the human non-small cell lung cancer (NSCLC) A549 cells were measured. Among the myricetin derivatives, S4-10 has displayed the highest antitumor efficacy in dose-dependent manner. The proliferation of A549 cells were significantly attenuated by given 6 µM of S4-10 both in vitro and in vivo. Further, the treatment of S4-10 also results in the inhibition of cell migration and invasiveness and the induction of cell apoptosis and G2 cycle arrest of A549 cells. Moreover, we found that S4-10 inhibits the progression of A549 cells through the sterol biosynthetic-cell apoptosis axis. These findings shed the light of developing S4-10 as a promising treatment agent for NSCLC.
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... The Lycium chinense combined to myricetin has favourable efficacy on antioxidant activity and the regenerative ratio of residue liver tissue after 70% PH in rats [46]. A report showed that human hepatocytes grow gradually its size for 3, 6 and 10 weeks after transplanting of myricetin-treated hepatocytes [47]. Collectively, myricetin might be an effective agent to relieve inflammation and to accelerate cell proliferation during LR. ...
We intended to identify favourable metabolite(s) and pharmacological mechanism(s) of gut microbiota (GM) for liver regeneration (LR) through network pharmacology. We utilized the gutMGene database to obtain metabolites of GM, and targets associated with metabolites as well as LR-related targets were identified using public databases. Furthermore, we performed a molecular docking assay on the active metabolite(s) and target(s) to verify the network pharmacological concept. We mined a total of 208 metabolites in the gutMGene database and selected 668 targets from the SEA (1,256 targets) and STP (947 targets) databases. Finally, 13 targets were identified between 61 targets and the gutMGene database (243 targets). Protein–protein interaction network analysis showed that AKT1 is a hub target correlated with 12 additional targets. In this study, we describe the potential microbe from the microbiota (E. coli), chemokine signalling pathway, AKT1 and myricetin that accelerate LR, providing scientific evidence for further clinical trials.
... Dietary flavonols are bioavailable molecules with human health benefits (e.g., antioxidant, cardioprotection, antibacterial, antiviral and anticancer activities). Using porcine and human hepatocytes in vitro and in vivo, Cui et al. found that myricetin could maintain liver functions involved in the mitogenactivated protein kinase pathway (Cui et al., 2019). Quercetin inhibits ROS-mediated hepatocarcinogenesis by upregulating enzymatic (e.g., SOD, GPx, CAT) and non-enzymatic (e.g., GSH, total glutathione) antioxidant defense systems (Rather & Bhagat, 2020). ...
Flavonoids are a group of natural polyphenol substances abundant in vegetables, fruits, grains, and tea. As plant secondary metabolites, flavonoids play essential roles in many biological processes and responses to environmental factors in plants. Flavonoids are common in human diets and have antioxidant effects as well as other bioactivities (e.g., antimicrobial and anti-inflammatory properties), which reduce the risk of disease. Flavonoid bioactivity depends on structural substitution patterns in their C6-C3-C6 rings. However, reviews of plant flavonoid distribution and biosynthesis, as well as the health benefits of its bioactivity, remain scarce. Therefore, in the present review, we systematically summarize recent progress in the research of plant flavonoids, focusing on their biosynthesis (pathway and transcription factors) and bioactive mechanisms based on epidemic evidence, in vitro and in vivo research, and bioavailability in the human body. We also discuss future opportunities in flavonoid research, including biotechnology, therapeutic phytoproducts, and dietary flavonoids.
... On the other hand, the flavonoid called myricetin (Myr) is a polyphenolic compound. The Myr is known for promoting antioxidative and cytoprotective effects in several cell lines [13,14]. Recent reports demonstrate that myricetin could be applied as an antibacterial agent [15], an anti-inflammatory agent [16], and it could be employed in anti-tumoral treatments [17,18]. ...
In this work, manganese iron oxide nanoparticles (MnFe2O4NPs) were synthesized by hydrothermal method using sodium dodecyl sulfate, sodium hydroxide, iron chloride, and manganese chloride. Four morphologies (observed by electron microscopy) were selected to be studied: flakes, rough-octahedrons, regular-octahedrons, and icosahedrons. The four samples were functionalized with the myricetin flavonoid (Myr). Hence, tetraethyl orthosilicate was employed to coat a SiO2 layer on the nanoparticles, and then, Myr was added for obtaining the so-call MnFe2O4NPs/Myr. The fast Fourier transform infrared spectroscopy (FTIR) confirmed the SiO2 coating formation and the Myr-functionalization. In addition, both Raman spectroscopy, and X-ray photoelectron spectroscopy (XPS), demonstrated the high density of oxygen vacancies in the flakes and rough-octahedrons MnFe2O4NPs. This amount was higher than that observed in the regular-octahedrons and icosahedrons MnFe2O4NPs. Among the synthesized samples, the icosahedrons MnFe2O4NPs exhibited the maximum efficiency for Myr-functionalization due to 0.53 MnFe2O4NPs/SiO2 mass rate for coupling 1 mg of Myr. This efficiency was attributed to the low oxygen vacancies concentration on the surface of the nanoparticles. The Myr-functionalization promoted a size stability enhancement (about twice) in all the samples, with no significant changes in size across 112 days (stored at RT).
... However, although liver transplantation is an effective treatment, patients have to take immunosuppressive drugs on a lifelong basis, and risk of rejection and need for dietary restrictions exist long after transplantation. While cell transplantation and gene therapy are being examined as novel treatments [5,[8][9][10], their safety and therapeutic efficacies have not yet been determined. From this point of view, human disease models in which clinical equivalent treatments are executable are preferable for the development of novel medical technologies. ...
... Thus, the severe fulminant OTCD model was established. The well-known OTCD model animals, sparse-fur (spf/1) mice (ID156, Balb/c background spontaneous mutation mice) show 1/6 the level of OTC activity (wild-type mice: 317.8 ± 51.5 nmol/min/mg, OTCD mice: 53.7 ± 33.5 nmol/min/mg, [mean ± SD, n = 3], unpublished data) and survive for approximately two months (median, 47 days; range: 20-52) [5,10]. In our previous study, hepatocyte transplantation two days after birth significantly prolonged the survival time in OTCD mice [10]. ...
... The well-known OTCD model animals, sparse-fur (spf/1) mice (ID156, Balb/c background spontaneous mutation mice) show 1/6 the level of OTC activity (wild-type mice: 317.8 ± 51.5 nmol/min/mg, OTCD mice: 53.7 ± 33.5 nmol/min/mg, [mean ± SD, n = 3], unpublished data) and survive for approximately two months (median, 47 days; range: 20-52) [5,10]. In our previous study, hepatocyte transplantation two days after birth significantly prolonged the survival time in OTCD mice [10]. A mild OTCD model will be more sensitive to detecting the effectiveness of treatments in pigs. ...
To develop novel medical technologies, pig disease models are invaluable especially in the final stages of translational research. Recently, we established a genetically engineered ornithine transcarbamylase-deficient (OTCD) pig strain. Here, we report its characterization and treatment responsiveness. OTCD pigs were obtained by mating an OTCD carrier female (OTC-Xc.186_190delXWT) with a wild-type male. Due to the X-linked recessive mode of inheritance, the disease phenotype emerged only in males. Medication with nitrogen-scavenging agents was based on a clinical protocol. OTCD pigs were born smaller than their wild-type and carrier littermates, showing anemia and faltering. Biochemically, high levels of urinary orotic acid and loss of OTC activity were observed. The natural life course of OTCD pigs was characterized by a decrease in arterial percentage saturation of oxygen and body temperature, as well as an increase in blood ammonia levels; the pigs died in 24.0 ± 5.0 h (mean ± SD, n = 6). The established standard medication composed with nitrogen-scavenging agents and transfusion nearly doubled the survival time to 42.4 ± 13.7 h (n = 6). Our OTCD pig model appropriately mimicked the human pathology. Along with established protocols in handling and medication, this is a first step in developing a large animal disease model that is useful for translational research into novel medical technologies, such as cell transplantation and gene therapy, as well as in relation to urea cycle disorder.
... Bulk-droplet-vitrified hepatocytes had significantly higher viability, better morphology, and higher metabolic activity than non-droplet frozen hepatocytes 93 . Aside from changing the cryopreservation techniques, addition of cell-survival signal, such as myricetin (inhibitor of mitogen-activated protein kinase kinase 4 (MKK4) in culture after thawing cryopreserved hepatocytes aids both the cell survival in vitro and after transplantation in immunodeficient mice 94 . ...
The origins of low-temperature tissue storage research date back to the late 1800s. Over half a century later, osmotic stress was revealed to be a main contributor to cell death during cryopreservation. Consequently, the addition of cryoprotective agents (CPAs) such as dimethyl sulfoxide (DMSO), glycerol (GLY), ethylene glycol (EG), or propylene glycol (PG), although toxic to cells at high concentrations, was identified as a necessary step to protect against rampant cell death during cryopreservation. In addition to osmotic stress, cooling and thawing rates were also shown to have significant influence on cell survival during low temperature storage. In general, successful low-temperature cell preservation consists of the addition of a CPA (commonly 10% DMSO), alone or in combination with additional permeating or non-permeating agents, cooling rates of approximately 1ºC/min, and storage in either liquid or vapor phase nitrogen. In addition to general considerations, cell-specific recommendations for hepatocytes, pancreatic islets, sperm, oocytes, and stem cells should be observed to maximize yields. For example, rapid cooling is associated with better cryopreservation outcomes for oocytes, pancreatic islets, and embryonic stem cells while slow cooling is recommended for cryopreservation of hepatocytes, hematopoietic stem cells, and mesenchymal stem cells. Yields can be further maximized by implementing additional pre-cryo steps such as: pre-incubation with glucose and anti-oxidants, alginate encapsulation, and selecting cells within an optimal age range and functional ability. Finally, viability and functional assays are critical steps in determining the quality of the cells post-thaw and improving the efficiency of the current cryopreservation methods.
... [8] Natural flavanols in herbs regulate the function and survival of hepatocytes both in-vitro and invivo strengthening the complement immunity indirectly. [9] Siddha literatures encourage "Microbial antagonism" through regular use of millets, prebiotics (fiber), probiotics (buttermilk, fermented porridge) for maintenance of healthy gut. The beneficial gut microflora improves the development and function of mucosal T cells subsets, specifically intraepithelial [Downloaded free from http://www.jrsm.in on Tuesday, November 1, 2022, IP: 24.163.96.211] lymphocytes and lamina propria CD 4 T cells. ...
This study evaluates the antibacterial effectiveness of Origanum vulgare hydroethanolic extract, both independently and in combination with antibiotics, against Escherichia coli strains associ-ated with avian colibacillosis—a significant concern for the poultry industry due to the rise of antibiotic-resistant E. coli. The urgent demand for new treatments is addressed by analyzing the extract's phytochemical makeup via High-Performance Liquid Chromatography (HPLC), which identified sixteen phenolic compounds. Antibacterial activity was determined through agar diffu-sion and the measurement of minimum inhibitory and bactericidal concentrations (MIC and MBC), showing moderate efficacy (MIC: 3.9 to 7.8 mg/ml, MBC: 31.2 to 62.4 mg/ml). Combining the extract with antibiotics like ampicillin and tetracycline significantly amplified antibacterial activity, indicating a synergistic effect and highlighting the importance of combinatory treat-ments against resistant strains. Further analysis revealed the extract's mechanisms of action in-clude disrupting bacterial cell membrane integrity and inhibiting ATPase/H+ proton pumps, es-sential for bacterial survival. Moreover, the extract effectively inhibited and eradicated biofilms, which is crucial for preventing bacterial colonization. Regarding cytotoxicity, the extract showed no hemolytic effect at concentrations ranging from 1 to 9 mg/ml. These results suggest Origa-num vulgare extract, particularly when used with antibiotics, offers a promising strategy for managing avian colibacillosis, providing both direct antibacterial benefits and moderating antibi-otic resistance, thus potentially reducing the economic impact of the disease on the poultry in-dustry.
