The triterpene saponin ginsenoside Rh2 has been shown to have antiproliferative effects on various cancer cells. However, the effect of Rh2 on the cell cycle and its underlying molecular mechanism in human leukemia cells are not fully understood. In this study, we found that Rh2 inhibited the proliferation of human leukemia cells concentration- and time-dependently with an IC 50 of ~38 µM. DNA flow cytometric analysis indicated that Rh2 blocked cell cycle progression at the G 1 phase in HL-60 and U937 cells, and this was found to be accompanied by the downregulations of cyclin-dependent kinase (CDK) 4, CDK6, cyclin D1, cyclin D2, cyclin D3 and cyclin E at the protein level. However, CDK inhibitors (CDKIs), such as p21 CIP1/WAF1 and p27 KIP1 , were gradually upregulated after Rh2 treatment at the protein and messenger RNA (mRNA) levels. In addition, Rh2 markedly enhanced the bindings of p21 CIP1/WAF1 and p27 KIP1 to CDK2, CDK4 and CDK6, and these bindings reduced CDK2, CDK4 and CDK6 activities. Furthermore, Rh2 induced the differentiation of HL-60 cells as demonstrated by biochemical assays and the expression levels of cell surface antigens. In addition, treatment of HL-60 cells with Rh2 significantly increased transforming growth factor-β (TGF-β) production, and cotreatment with TGF-β neutralizing antibody prevented the Rh2-induced downregulations of CDK4 and CDK6, upregulations of p21 CIP1/WAF1 and p27 KIP1 levels and the induction of differentiation. These results demonstrate that the Rh2-mediated G 1 arrest and the differentiation are closely linked to the regulation of TGF-β production in human leukemia cells.
Patho-physiological conditions with high oxidative stress, such as conditions associated with increased denatured heme-proteins, are associated with enhanced adipogenic response. This effect predominantly manifests as adipocyte hypertrophy characterized by dysfunctional, pro-inflammatory adipocytes exhibiting reduced expression of anti-inflammatory hormone, adiponectin. To understand how increased levels of cellular heme, a pro-oxidant molecule, modulates adipogenesis; the following study was designed to evaluate effects of heme on adipogenesis in human mesenchymal stem cells (hMSCs) and mouse pre-adipocytes (3T3L1). Experiments were conducted in the absence and in the presence of a superoxide dismutase (SOD) mimetic (tempol, 100 µM). Heme (10 µM) increased (P<0.05) adipogenesis in hMSCs and mouse pre-adipocytes, where tempol alone (100 µmol/L) attenuated adipogenesis in these cells (P<0.05). Tempol also reversed heme-induced increase in adipogenesis in both hMSCs and mouse pre-adipocytes (P<0.05). In addition, heme exposed 3T3L1 exhibited reduced (P<0.05) expression of transcriptional regulator-sirtuin 1 (Sirt1), along with, increased (P<0.05) expression of adipogenic markers peroxisome proliferators-activated receptor-gamma (PPARγ), C/EBPα, and aP2. These effects of heme were rescued (P<0.05) in cells concurrently treated with heme and tempol (P<0.05) and prevented in cells over-expressing Sirt1. Taken together, our results indicate that heme-induced oxidative stress inhibits Sirt1, thus un-inhibiting adipogenic regulators such as PPARγ and C/EBPα; which in turn induce increased adipogenesis along with adipocyte hypertrophy in pre-adipocytes. Anti-oxidant induced offsetting of these effects of heme supports the role of heme-dependent oxidative stress in mediating such events.
A superabsorbent polymer (SAP) is a special polymer material that can absorb up to 500 times its own weight of pure water, but has a problem that it does not biodegrade itself and cause environmental pollution. Therefore, we aim to prepare a biodegradable SAP by using biomass‐based IA. The SAP must be able to retain absorbed water and absorb water under a given pressure. We have carried out studies to improve the surface hardness of the SAP to enhance absorption of water under a given pressure by surface‐crosslinking. Four types of surface‐crosslinkers, ethylene glycol diglycidyl ether (EGDGE), ethylene carbonate (EC), 1,4‐butanediol (BD), or glycerol, were used. We confirmed the water absorption capacity of the SAP by measuring its centrifuge retention capacity (CRC) and absorbency under load (AUL). The structural characteristics of the SAP were confirmed by attenuated total reflection (ATR) and X‐ray photoelectron spectroscopy (XPS), and the surface characteristics were confirmed by scanning electron microscopy (SEM).
TGF-β/Smad signaling is a major pathway in progressive fibrotic processes, and further studies on the molecular mechanisms of TGF-β/Smad signaling are still needed for their therapeutic targeting. Recently, peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) was shown to improve renal fibrosis, making it an attractive target for chronic kidney diseases (CKDs). Here, we show the mechanism by which PGC-1α regulates the TGF-β/Smad signaling pathway using HK-2 cell lines stably overexpressing empty vector (mock cells) or human PGC1α (PGC1α cells). Stable PGC-1α overexpression negatively regulated the expression of TGF-β-induced epithelial-mesenchymal transition (EMT) markers (fibronectin, E-cadherin, vimentin, and α-SMA) and EMT-related transcription factors (Snail and Slug) compared to mock cells, inhibiting fibrotic progression. Interestingly, among molecules upstream of Smad2/3 activation, the gene expression of only TGFβRI, but not TGFβRII, was downregulated in PGC-1α cells. In addition, the downregulation of TGFβRI by PGC-1α was associated with the upregulation of let-7b/c, miRNA for which the 3′ untranslated region (UTR) of TGFβRI contains a binding site. In conclusion, PGC-1α suppresses TGF-β/Smad signaling activation via targeting TGFβRI downregulation by let-7b/c upregulation.
Abstract IL-1β is a key mediator of sterile inflammation in response to endogenous particulates, a type of damage-associated molecular pattern (DAMPs) molecule derived from damaged cells. Despite the well-known role of sterile particulates such as monosodium urate (MSU) crystals as inflammasome inducers in monocytes/macrophages, little is known regarding how pro-IL-1β synthesis is induced under sterile inflammatory conditions. We provide evidence that MSU crystals post-transcriptionally induce the rapid production of pro-IL-1β in human primary monocytes. Metabolic labeling and pull-down assays for newly-synthesized proteins clearly showed that MSU crystals rapidly, within 30 min, induce the synthesis of pro-IL-1β as well as global proteins. Notably, MSU crystal-induced pro-IL-1β synthesis is selectively dependent on the p38 MAPK pathway, whereas global protein synthesis is mediated via the mTOR, ERK1/2, and p38 pathways. Furthermore, inhibition of Mnk1, a substrate of p38, blocked MSU crystal-induced pro-IL-1β synthesis downstream of eIF4E phosphorylation. In addition, the p38 MAPK pathway leading to phosphorylation of MK2 was also critical for stabilization of pro-IL-1β mRNA following MSU stimulation. Our findings demonstrate that post-transcriptional regulation via p38 MAPK plays a central role in the rapid synthesis of pro-IL-1β in response to MSU crystals, which is an essential step for IL-1β production in human monocytes.
The side effects of cisplatin, a widely used chemotherapeutic agent, include nephrotoxicity. Previous studies have reported that cisplatin induces ferroptosis and lipid peroxide accumulation. Ferroptosis, a type of regulated cell death, is characterized by iron-dependent lipid peroxidation. Although previous studies have examined the regulation of ferroptosis in acute kidney injury (AKI), the regulatory mechanism of ferroptosis has not been elucidated. Here, the ability of activated farnesoid X receptor (FXR) to attenuate cisplatin-induced AKI through the regulation of ferroptosis was examined. FXR deficiency exhibited more ferroptosis responses, such as increase in lipid peroxidation, iron content and heme oxygenase 1 protein, and a decrease in glutathione/glutathione disulfide ratio and glutathione peroxidase 4 levels in HK2 cells and mice. Increased blood urea nitrogen, serum creatinine, and ferroptotic responses in the cisplatin-induced AKI mouse model were mitigated upon treatment with the FXR agonist GW4064 but were exacerbated in FXR knockout mice. RNA sequencing analysis revealed that ferroptosis-associated genes were novel targets of FXR. FXR agonist upregulated the expression of lipid and glutathione metabolism-related genes and downregulated cell death-related genes. Additionally, chromatin immunoprecipitation assays, using mice renal tissues, revealed that agonist-activated FXR could bind to its known target genes (Slc51a, Slc51b, Osgin1, and Mafg) and ferroptosis-related genes (Aifm2, Ggt6, and Gsta4). Furthermore, activated FXR-dependent MAFG, a transcriptional repressor, could bind to Hmox1, Nqo1, and Tf in the renal tissues of FXR agonist-treated mice. These findings indicate that activated FXR regulates the transcription of ferroptosis-related genes and protects against cisplatin-induced AKI.
The neuroendocrine response to episodes of acute stress is crucial for survival whereas the prolonged response to chronic stress can be detrimental. Learning and memory are particularly susceptible to stress with cognitive deficits being well characterized consequences of chronic stress. Although there is good evidence that acute stress can enhance cognitive performance, the mechanism(s) for this are unclear. We find that hippocampal slices, either prepared from rats following 30 min restraint stress or directly exposed to glucocorticoids, exhibit an N-methyl-d-aspartic acid receptor-independent form of long-term potentiation. We demonstrate that the mechanism involves an NMDA receptor and PKA-dependent insertion of Ca2+ -permeable AMPA receptors into synapses. These then trigger the additional NMDA receptor-independent form of LTP during high frequency stimulation.
SIRT1, a highly conserved NAD+-dependent protein deacetylase, is a key metabolic sensor that directly links nutrient signals to animal metabolic homeostasis. Although SIRT1 has been implicated in a number of hepatic metabolic processes, the mechanisms by which hepatic SIRT1 modulates bile acid metabolism are still not well understood. Here we report that deletion of hepatic SIRT1 reduces the expression of farnesoid X receptor (FXR), a nuclear receptor that regulates bile acid homeostasis. We provide evidence that SIRT1 regulates the expression of FXR through hepatocyte nuclear factor 1α (HNF1α). SIRT1 deficiency in hepatocytes leads to decreased binding of HNF1α to the FXR promoter. Furthermore, we show that hepatocyte-specific deletion of SIRT1 leads to derangements in bile acid metabolism, predisposing the mice to development of cholesterol gallstones on a lithogenic diet. Taken together, our findings indicate that SIRT1 plays a vital role in the regulation of hepatic bile acid homeostasis through the HNF1α/FXR signaling pathway.
Renal fibrosis is a chronic pathological process that seriously endangers human health. However, the current therapeutic options for this disease are extremely limited. Previous studies have shown that signaling factors such as JAK2/STAT3, Smad3, and Myd88 play a regulatory role in renal fibrosis, and β-elemene is a plant-derived sesquiterpenoid organic compound that has been shown to have anti-inflammatory, anti-cancer, and immunomodulatory effects. In the present study, the anti-fibrotic effect of β-elemene was demonstrated by in vivo and in vitro experiments. It was shown that β-elemene inhibited the synthesis of extracellular matrix-related proteins in unilateral ureteral obstruction mice, and TGF-β stimulated rat interstitial fibroblast cells, including α-smooth muscle actin, vimentin, and connective tissue growth factor, etc. Further experiments showed that β-elemene reduced the expression levels of the above-mentioned fibrosis-related proteins by blocking the phosphorylation of JAK2/STAT3, Smad3, and the expression or up-regulation of MyD88. Notably, knockdown of MyD88 attenuated the phosphorylation levels of STAT3 and Smad3 in TGF-β stimulated NRK49F cell, which may be a novel molecular mechanism by which β-elemene affects renal interstitial fibrosis. In conclusion, this study elucidated the anti-interstitial fibrosis effect of β-elemene, which provides a new direction for future research and development of drugs related to chronic kidney disease.
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