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@#Vascular malformations, which mainly occur in the head and neck region, are a group of congenital disorders that cannot involute and dilate gradually as patients grow. Traditional therapeutic strategies for vascular malformations include laser therapy, sclerotherapy, interventional embolization, surgical resection, etc. However, for some cases with a relatively larger range of lesions, traditional therapeutic strategies might fall short of the goals. With the development of molecular genetics, gene mutations are currently recognized as the root cause of the occurrence of vascular malformations. The progression of vascular malformation lesions is further promoted by the activation of related pathways. Low-flow vascular malformations mainly involve activation of the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) pathway, whereas high-flow vascular malformations mainly involve activation of the rat sarcoma (RAS)/rapidly accelerated fibrosarcoma (RAF)/mitogen-activated protein kinase kinase (MAPKK)/extracellular-signal regulated protein kinase (ERK) pathway. Targeted drugs against relevant gene mutations and signaling pathways have also been applied in the treatment of vascular malformations, and previous studies have shown that the mTOR inhibitor rapamycin is effective and now widely used in the treatment of low-flow vascular malformations. The PI3K inhibitor alpelisib is also promising in the treatment of venous malformations, and the MAPKK inhibitor trametinib has shown good results in the treatment of arteriovenous malformations. Therefore, traditional therapies supplemented by targeted drugs may bring new breakthroughs to the treatment of vascular malformations.
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ObjectiveBy observing the effect of Xiaoluowan on phosphatidylinositol 3-kinase （PI3K）/protein kinase B （Akt）/mammalian target of rapamycin complex 1 （mTORC1） pathway in experimental goiter rats， this study aims to explore its therapeutic effect on experimental goiter rats. MethodSixty 5-month-old SD rats of SPF grade were purchased， half males and half females， of which 10 were used as a normal group， and the remaining rats were administrated with propylthiouracil （PTU） solution to induce nodular goiter. After successful modeling， rats were randomly divided into a model group， levothyroxine sodium tablets group， Xiaoluowan low-dose group， medium-dose group， and high-dose group， ten rats each. The levothyroxine sodium tablets group was given 15 μg·kg-1 levothyroxine sodium tablets by gavage. The Xiaoluowan low-， medium-， and high-dose groups were given （ig） Xiaoluowan low-dose （10 g·kg-1）， medium-dose （20 g·kg-1）， and high-dose （30 g·kg-1） Xiaoluowan， and the normal group and model group were administered （ig） with the same volume of 0.9% sodium chloride solution. Four weeks after the intervention， rats were sacrificed by routine intraperitoneal anesthesia using 5% phenobarbital. Subsequently， the histopathology was observed under a microscope， and serum thyroid hormone levels were measured using a Roche electrochemiluminescence immunoassay analyzer. Serum cytokines were detected by enzyme-linked immunosorbent assay （ELISA）， and neurotransmitters were measured using a high-performance liquid chromatograph. The protein level of PI3K/Akt/mTORC1 pathway was determined by Western blot. ResultAs compared with the normal group， the levels of basic fibroblast growth factor （bFGF）， vascular endothelial growth factor （VEGF）， insulin-like growth factor （IGF-1）， 5-hydroxytryptamine （5-HT）， and thyroid stimulating hormone （TSH） were increased， and PI3K， Akt， and mTORC1 protein levels were up-regulated in the model group， while the levels of norepinephrine （NE）， triiodothyronine （T3）， tetraiodothyronine （T4）， free triiodothyronine （FT3）， and free thyroid hormone （FT4） were decreased （P<0.05）. Compared with the model group， the levothyroxine sodium tablets group， and Xiaoluowan low-， medium-， and high-dose groups exhibited reduced levels of bFGF， VEGF， IGF-1， 5-HT， and TSH， and down-regulated PI3K， Akt， and mTORC1 protein levels， and increased NE， T3， T4， FT3， and FT4 levels （P<0.05）. ConclusionXiaoluowan may act on the PI3K/Akt/mTORC1 signaling pathway to play its role in the treatment of nodular goiter， and it is dose-dependent.
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Idiopathic pulmonary fibrosis （IPF） is a common， lethal interstitial lung disease characterized by airway remodeling， inflammation， alveolar destruction， and fibrosis. The mammalian target of rapamycin complex 1/4E binding protein 1 （mTORC1/4E-BP1） axis is closely related to the expression of collagen by fibroblasts， and its role in pulmonary fibrosis remains to be further elucidated. Traditional Chinese medicine （TCM） has shown promising efficacy in improving the lung function， exercise capacity， and quality of life in patients with IPF. The theory of "same treatment for different diseases" provides a TCM theoretical basis for the treatment of pulmonary fibrosis with Bupleuri Radix， while the research in western medicine has preliminarily shown that both the formulation and single herb as well as the active ingredients of Bupleuri Radix have good therapeutic effects on pulmonary fibrosis. Therefore， this review will elaborate on the role of the mTORC1/4E-BP1 axis in the pathomechanism of IPF， as well as the research results of the active components of Bupleuri Radix on the phosphoinositide 3-kinase/protein kinase B/mammalian target of rapamycin protein（PI3K/AKT/mTOR） pathway， so as to provide a reference for the treatment and drug development of IPF.
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Objective @#To investigate the activation of the mammalian target of rapamycin complex 1 (mTORC1) signaling pathway molecules during the process by which kaempferol (Kae) promotes osteogenic differentiation of mouse bone marrow mesenchymal cells (BMMCs) under cyclic and uniaxial tension.@*Methods @#BMMCs isolated and cultured in vitro were subjected to uniaxial dynamic tension with a 10% shape variable. The appropriate concentration of Kae was selected by cytotoxicity testing. The endogenous mTOR signal was inhibited by pp242. Four hours after traction, alkaline phosphatase (ALP) and osteocalcin (OCN) were detected by chemical colorimetry and ELISA, and the relative concentration of intracellular calcium was detected by flow cytometry. Phosphorylation of mTOR, 4E/BP1, and ribosomal protein S6 kinases (S6K), which are the main molecules of the endogenous mTORC1 signaling pathway, and expression of osteogenic transcription factors (Runx2 and Osterix) were detected by western blotting (WB), and mRNA expression levels of the above factors were detected by qRT-PCR.@*Results @# The cytotoxicity test showed that 10 μmol/L Kae had little inhibitory effect on cell proliferation but had the strongest osteogenic ability. Four hours after stretching, Kae effectively promoted the osteogenic differentiation of BMMCs. The expression of ALP was （153.04 ± 18.72） U/mg, the expression of OCN was （1.64 ± 0.25） U. The mRNA and protein levels of Runx2 and Osterix were upregulated, and the intracellular calcium content was decreased. The mRNA and protein phosphorylation of mTOR and S6K was upregulated, and the opposite effect was observed with 4E/BP1. After pp242 was added to inhibit mTOR signaling, mTOR and S6K mRNA and protein phosphorylation were downregulated, but 4E/BP1 mRNA and protein phosphorylation was upregulated. The osteogenic differentiation of BMMCs was also significantly inhibited, mRNA and protein expression of Runx2 and Osterix were significantly downregulated, ALP and OCN expression were downregulated, and intracellular calcium content was increased. @* Conclusion@#Kae promotes osteogenic differentiation of mouse BMMCs under uniaxial dynamic tension through the mTORC1 signaling pathway.
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Objective::To observe the effect of Shenling Baizhusan(SBS)on the mammalian target of rapamycin complex 1 (mTORC1)/signal transducers and activators of transcription 3 (STAT3) pathway in liver hepatocyte of nonalcoholic fatty liver disease(NAFLD)rats induced by high fat diet, in order to reveal the mechanism of SBS against rat NAFLD from the perspective of inflammation. Method::Totally 80 SD rats were randomly divided into 4 groups, normal control group, model group, high-dose SBP group(30 g·kg-1), and low-dose SBS group(10 g·kg-1), with 20 rats in each group. The rats of NAFLD model were established by being fed with high-fat diets for 8 weeks, and the treatment groups were fed with high or low dose of SBS respectively. After treatment for 8 weeks, blood and liver samples of rats were collected. Alanine aminotransferase (ALT), aspartate aminotransferase(AST), total cholesterol (TC), triglyceride(TG), high-density lipoprotein cholesterol(HDL-C)and low-density lipoprotein cholesterol(LDL-C)levels in blood serum were detected with automatic biochemical analyzer. The liver tissues were observed by oil red O and hematoxylin-eosin (HE) staining. Hepatocytes were isolated by type Ⅳ collagenase perfusion in vitro. Tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-5 and IL-6 in hepatocytes were detected by enzyme-linked immunosorbent assay (ELISA), and the relevant gene and proteins expressions of mTORC1 and STAT3 in hepatocytes were detected by Real-time fluorescent quantitative polymerase chain reaction (Real-time PCR) and Western blot detection respectively. Result::Compared with the normal control group, the serum levels of TG, TC, AST, ALT and LDL-C were increased significantly, the levels of TNF-α, IL-1β, IL-5 and IL-6 in hepatocytes were increased significantly, and the expression levels of mTORC1, STAT3 mRNA and proteins in hepatocytes were increased significantly(P<0.01). Compared with the model group, the hepatic lipid accumulation of the medicine intervention group was relieved significantly, the serum levels of AST, ALT, TG and LDL-C were decreased significantly, the expression levels of TNF-α, IL-1β, IL-5 and IL-6 of hepatocytes were decreased significantly, and the expressions of mTORC1, STAT3 mRNA and proteins in hepatocytes were decreased significantly(P<0.05, P<0.01). In the high-dose SBS group, the effects in improving the lipid accumulation and inhibiting the inflammatory reaction were better than those of the low-dose SBS group, and the expressions of mTORC1 and STAT3 genes and proteins in hepatocytes were significantly decreased (P<0.05, P<0.01). Conclusion::SBS can improve the fat metabolism disorder and reduce liver lipid accumulation and inflammatory reaction in NAFLD rats induced by high-fat diet. The mechanism may be correlated with the inhibition of mTORC1/STAT3 pathway relating to genes and protein expression in hepatocytes.
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Rab1A is a member of the RAB family as a small GTPase and a mammalian target of rapamycin complex 1 (mTORC1) Activator,which has been well established to mediate vesicular trafficking from the endoplasmic reticulum (ER) to the Golgi apparatus. With increasingly deep research of Rab1A,many researchers discover that Rab1A protein is also involved in mediating signal transduction,cell migration and regulation of autophagy. Meanwhile,aberrant expression of Rab1A has been linked to a range of human diseases as well,including Parkinson's disease,cardiomyopathy,and aspirin-exacerbated respiratory disease. Recently Rab1A research has gradually shifted to its role in occurrence and development of tumor,and researchers also discover that Rab1A was abnor-mally expressed in many malignant tumors such as tonguesquamous carcinoma,breast cancer,human lung cancer,hepatocellular carci-noma,colorectal cancer,gastric cancer and cervical cancer. Besides that,over expression of Rab1A plays a significant role in the pro-gression of different tumors. This article summarizes the research progress of Rab1A intumor development and signaling pathways.
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[ ABSTRACT] AIM: To investigate the influence of advanced glycosylation end products-modified bovine serum albumin (AGE-BSA) on mammalian target of rapamycin complex 1 (mTORC1), urokinase-type plasminogen activator re-ceptor ( uPAR) , and cell mobility in the podocytes, and to further explore the probable relationship.METHODS: The conditionally immortalized mouse podocyte cell line was cultured in vitro.MTT assay and immunofluorescence were used to analyze the cell viability and cytoskeleton of the podocytes treated with the stimuli and intervention agents.The activity of mTORC1 and the expression level of uPAR in normal podocytes and podocytes treated with control BSA or AGE-BSA were detected by Western blotting.The migration ability of the podocytes was determined by would-healing assay.Rapamycin was added to inhibit the activity of mTORC1 along with the addition of AGE-BSA to observe the changes of uPAR and the motility of podocytes.RESULTS:No significant difference of the cell viability or cytoskeleton in the podocytes treated with the stimuli and intervention agents was observed.AGE-BSA up-regulated the activity of mTORC1 and the expression of uPAR, and induced the high mobility of the podocytes.Rapamycin obviously reduced the high expression level of uPAR and the increase in the migration ability of podocytes caused by AGE-BSA treatment.CONCLUSION: AGE-BSA might cause the high migration of podocytes through the mTORC1/uPAR signaling pathway.
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Objective To explore the effect and its mechism of osteoprotegerin (OPG) on human umbilical vein endothelial cells(HUVECs) induced by high glucose.Methods (1) The cultured HUVECs were treated with normal glucose(5.5 mmol/L),high glucose(33 mmol/L),high glucose + OPG(0.5,1,and 2 μg/ml)as well as mannitol(5.5 mmol/L glucose+27.5 mmol/L mannitol) for 48 h,respectively.Flow cytometry assay and Hoechst 33258 staining were used to detect the cell apoptosis.The expression levels of Bcl-2 and Bax protein were measured by western blot analysis.(2) The cultured HUVECs were treated with normal glucose,high glucose,high glucose + OPG (2 μg/ml OPG),high glucose + rapamycin(10 ng/ml rapamycin) as well as high glucose + rapamycin + OPG for 24h,respectively.The expression levels of S6K,p-S6K,4EBP1 and p-4EBP1 protein were measured by western blot analysis.(3)The cultured HUVECs were treated with normal glucose,high glucose,high glucose + OPG(2 μg/ml)for 24 h,respectively.The expression levels of tuberin and p-tuberin protein were measured by western blot analysis.Results (1) Compared with normal glucose group,the apoptosis of HUVECs and the expression level of Bax was dramatically increased,and the expression of Bcl-2 was decreased significantly in high glucose group (P＜0.05).The apoptosis of HUVECs and the expression level of Bax in high glucose + OPG group was significantly lower than that in high glucose group,which was still higher than that in normal glucose group (P＜ 0.05).and the expression of Bcl-2 in high glucose + OPG group was significantly higher than that in high glucose group,which was still lower than that in normal glucose group (P＜0.05).There was no statistically difference between hyperosmolar control group and normal glucose group.(2) Compared with normal glucose group,the expression levels of p-S6K and p-4EBP1 were increased markedly in high glucose group(P＜0.05).The expression levels of p-S6K and p-4EBP1 in high glucose + OPG group were significantly lower than that in high glucose group,which were still higher than that in normal glucose group(P＜0.05).No significant difference was found between high glucose + OPG group and high glucose + rapamycin group.(3) Compared with normal glucose group,the expression level of p-tuberin was increased markedly in high glucose group (P ＜ 0.05).The expression level of p-tuberin in high glucose + OPG group was significantly lower than that in high glucose group,which was still higher than that in normal glucose group (P ＜0.05).Conclusions It suggests that the protective effect and mechanism of OPG on HUVECs cultured with high glucose may be association with tuberin/mTORC1 pathway.