Lin28a has diverse functions including regulation of cancer, reprogramming and regeneration, but whether it promotes injury or is a protective reaction to renal injury is unknown. We studied how Lin28a acts in unilateral ureteral obstruction (UUO)-induced renal fibrosis following unilateral ureteral obstruction, in a mouse model. We further defined the role of Lin28a in transforming growth factor (TGF)-signaling pathways in renal fibrosis through in vitro study using human tubular epithelium-like HK-2 cells. In the mouse unilateral ureteral obstruction model, obstruction markedly decreased the expression of Lin28a, increased the expression of renal fibrotic markers such as type I collagen, -SMA, vimentin and fibronectin. In TGF--stimulated HK-2 cells, the expression of Lin28a was reduced and the expression of renal fibrotic markers such as type I collagen, -SMA, vimentin and fibronectin was increased. Adenovirus-mediated overexpression of Lin28a inhibited the expression of TGF--stimulated type I collagen, -SMA, vimentin and fibronectin. Lin28a inhibited TGF--stimulated SMAD3 activity, via inhibition of SMAD3 phosphorylation, but not the MAPK pathway ERK, JNK or p38. Lin28a attenuates renal fibrosis in obstructive nephropathy, making its mechanism a possible therapeutic target for chronic kidney disease.
An excessive and prolonged increase in glucose levels causes β-cell dysregulation, which is accompanied by impaired insulin synthesis and secretion, a condition known as glucotoxicity. Although it is known that both Lin28a and Lin28b regulate glucose metabolism, other molecular mechanisms that may protect against glucotoxicity are poorly understood. We investigated whether Lin28a overexpression can improve glucotoxicity-induced β-cell dysregulation in INS-1 and primary rat islet cells. INS-1, a rat insulinoma cell line was cultured and primary rat islet cells were isolated from SD-rats. To define the effect of Lin28a in chronic high glucose-induced β-cell dysregulation, we performed several in vitro and ex-vivo experiments. Chronic exposure to high glucose led to a downregulation of Lin28a mRNA and protein expression, followed by a decrease in insulin mRNA expression and secretion in β-cells. The mRNA and protein expression levels of PDX-1 and BETA2, were reduced; The levels of apoptotic factors, including c-caspase3 and the Bax/Bcl-2 ratio, were increased due to glucotoxicity. Adenovirus-mediated Lin28a overexpression in β-cells reversed the glucotoxicity-induced reduction of insulin secretion and insulin mRNA expression via regulation of β-cell-enriched transcription factors such as PDX-1 and BETA2. Adenovirus-mediated overexpression of Lin28a downregulated the glucotoxicity-induced upregulation of c-caspase3 levels and the Bax/Bcl-2 ratio, while inhibition of endogenous Lin28a by small interfering RNA resulted in their up-regulation. Lin28a counteracted glucotoxicity-induced downregulation of p-Akt and p-mTOR. Our results suggest that Lin28a protects pancreatic β-cells from glucotoxicity through inhibition of apoptotic factors via the PI3 kinase/Akt/mTOR pathway.
Rhus verniciflua is widely known for its antioxidant, antibacterial, anticancer, and antiaging efficacy and α-glucosidase inhibition. This study was designed whether Rhus verniciflua extracts inhibit the IgE-antigen-mediated allergic reaction in RBL-2H3 mast cells, and it further investigated the FcεRI- and arachidonate-signaling by which Rhus verniciflua extracts exert its antiallergic effects. IgE-antigen-sensitized RBL-2H3 mast cells were investigated for the cytotoxicity of Rhus verniciflua extracts and β-hexosaminidase release, and inflammatory mediators (e.g., TNF-α, IL-4, IL-6, histamine, and PGD2) were then assessed. Additionally, we examined expressions of genes involved in arachidonate- and FcεRI-signaling pathway in RBL-2H3. Rhus verniciflua extracts inhibited β-hexosaminidase release and production of the inflammatory mediators in RBL-2H3. Rhus verniciflua extracts reduced amounts of histamine and expressions of FcεRI signaling-related genes such as Lyn and Syk and phosphorylation of extracellular signal-regulated kinase in mast cells. Finally, in late allergic responses, Rhus verniciflua extracts reduced PGD2 release and COX-2 and cPLA2 phosphorylation expressions from IgE-antigen-mediated mast cells. Lastly, 250–500 mg/kg RVE significantly attenuated the Ag/IgE-induced passive cutaneous anaphylaxis (PCA) reaction in mice. These findings provide novel information on the molecular mechanisms underlying the antiallergy properties of Rhus verniciflua extracts in FcɛRI-mediated allergic reaction.
Enhancement of β-cell proliferation plays an important role in maintaining β-cell mass and function, and in improving pancreatic β-cell survival before transplantation. Extracellular matrix (ECM) components increase the adhesion and proliferation of β-cells, and the RGD-modified elastin-like polypeptide (RGD-ELP, REP) has been described as a bioactive matrix. In this study, we investigated whether REP could enhance β-cell adhesion and proliferation and elucidated the signaling pathways involved. Methods: We investigated the effect of REP on cell adhesion, proliferation and insulin secretion via assays using Rin-m and rat islets. Crystal violet, CCK-8, and BrdU assay, FACS, western blot, real time q-PCR analyses and insulin ELISA were examined. To explain the associated mechanisms, phosphorylation of Akt and extracellular signal-regulated kinase (Erk) were measured. Results: REP more increased the adhesion, proliferation and survival of Rin-m cells compared to elastin-like poly peptide (ELP) without RGD-motif. The enhancement of β-cell proliferation by REP was associated with increased cyclin D1, cyclin D2 and cdk6, and decreased p27 levels. When β-cells were cultured on REP, Erk and the phosphatidylinositol 3-kinase (PI3-kinase) downstream effector, Akt was stimulated. Treatment with the Erk pathway inhibitor and PI3-kinase inhibitor decreased REP-induced β-cell adhesion and proliferation, and regulated REP-induced cell cycle proteins. Additionally, REP increased the mRNA and protein levels of insulin and its transcription factor, PDX-1, and insulin secretion. Conclusions: Our results demonstrate that the up-regulation of the PI3K/Akt and Erk signaling pathways and the regulation of cell cycle proteins by REP could serve as effective strategies for improving pancreatic β-cell adhesion and proliferation.
This study was conducted to produce water-soluble polysaccharide extracts (WSP) from perilla seed meal (PSM), a by-product of perilla seed oil extraction, using enzymatic hydrolysis. The aim was to confirm the potential of this method to obtain functional materials from PSM. The cellulose and hemicellulose fractions from PSM were hydrolyzed using Celluclast 1.5 L and Viscozyme L, respectively. The yield of WSP from both fractions increased with an increase in hydrolysis time. Physical properties of WSP, including solubility, oil-holding capacity, and emulsification properties, were increased by enzymatic hydrolysis. WSP produced by hydrolysis had greater antioxidant activity than WSP obtained without hydrolysis; the highest activity was observed after a hydrolysis reaction of 24 h. WSP retarded glucose and bile acid absorption. The data indicate that WSP from PSM have potentially valuable functional and biological activities. Características de los extractos de polisacáridos solubles en agua producidos a partir de harina de semillas de perilla mediante hidrólisis enzimática RESUMEN Este estudio se llevó a cabo para producir, mediante hidrólisis enzimática, extractos de polisacáridos solubles en agua (WSP) a partir de harina de semilla de perilla (PSM), un subproducto de la extracción de aceite de semilla de perilla. El objetivo de este ejercicio era confirmar el potencial de este método para obtener materiales funcionales a partir de la PSM. Las fracciones de celulosa y hemicelulosa de PSM se hidrolizaron utilizando Celluclast 1,5 L y Viscozyme L, respectivamente. El rendimiento de la PSM de ambas fracciones aumentó con el incremento del tiempo de hidrólisis. Las propiedades físicas de la PSM, entre ellas la solubilidad, la capacidad de retención de aceite y las propiedades de emulsificación, también se elevaron con la hidrólisis enzimática. Los WSP producidos por hidrólisis mostraron mayor actividad antioxidante que aquellos obtenidos sin aplicar hidrólisis; la mayor actividad se observó después de una reacción de hidrólisis de 24 horas. Los WSP retardaron la absorción de glucosa y de ácidos biliares. Los datos permiten concluir que los WSP producidos a partir de PSM tienen actividades funcionales y biológicas potencialmente valiosas.
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