OBJECTIVE: To study the regulatory effect of Cheng's Juanbi Decoction (JBT) on autophagy in rheumatoid arthritis fibroblast-like synoviocytes (RA-FLS) and role of PI3K/Akt/mTOR signaling axis in the mechanism mediating this effect. METHODS: CCK8 assay was used to determine the optimal concentration and treatment time of JBT for inhibiting the viability of RA- FLS. The effect of freeze-dried powder of JBT, RAPA, or both on morphology of the autophagosomes in RA-FLS was observed under transmission electron microscope, and the changes in the number of autophagosomes and autolysosomes were observed with autophagy double-labeled adenovirus experiment. RT-qPCR and Western blotting were used to detect the expression levels of the related indicators. RESULTS: The results of CCK8 assay showed that treatment with 0.5 mg/mL JBT for 12 h produced the optimal effect for inhibiting RA-FLS viability. Observation with transmission electron microscope and the results of the autophagy double-labeled adenovirus experiment both showed the presence of a small number of autophagosomes in control RA-FLS group, and treatment with JBT significantly increased the number of autophagosomes and lowered the number of autophagolysosomes in the cells. Compared with the control cells and the cells treated with JBT or RAPA alone, the cells treated with both JBT and RAPA showed significantly decreased mRNA levels of PI3K, Akt and mTOR (P < 0.01) but without significant changes in their protein expressions (P > 0.05); the combined treatment significantly inhibited the protein expressions of p-PI3K, p-Akt, p-mTOR, and P62 (P < 0.05) and upregulated the protein expressions of Beclin-1 and LC3B (P < 0.05) in the cells. CONCLUSION JBT can inhibit the survival rate of RA-FLS and increase the level of autophagy possibly through a mechanism that down-regulates PI3K/Akt/mTOR signaling pathway.
Humans ; Phosphatidylinositol 3-Kinases ; Autophagy ; Fibroblasts ; Synoviocytes ; Arthritis, Rheumatoid ; Adenoviridae ; TOR Serine-Threonine Kinases

Amino acids are fundamental nutrients for protein synthesis and cell growth (increase in cell size). Recently, many compelling evidences have shown that the level of amino acids is sensed by extra- or intra-cellular amino acids sensor(s) and regulates protein synthesis/degradation. Mammalian target of rapamycin complex 1 (mTORC1) is placed in a central position in cell growth regulation and dysregulation of mTOR signaling pathway has been implicated in many serious human diseases including cancer, diabetes, and tissue hypertrophy. Although amino acids are the most potent activator of mTORC1, how amino acids activate mTOR signaling pathway is still largely unknown. This is partly because of the diversity of amino acids themselves including structure and metabolism. In this review, current proposed amino acid sensing mechanisms to regulate mTORC1 and the evidences pro/against the proposed models are discussed.
Amino Acids ; Humans ; Hypertrophy ; Multiprotein Complexes ; Sirolimus ; TOR Serine-Threonine Kinases

To detemine the expression pattern of mTOR complex subunits Raptor and Rictor in the hair follicles of mice at different hair follicle stages, and to explore its significance. 
 Methods: Immunostaining of Ki-67, a proliferative marker, was used to determine the precise hair follicle stages of mouse dorsal skin at different postnatal time points. Real-time PCR was used to detect the mRNA expression of Raptor and Rictor in mouse dorsal skin at 43 days after birth (P43, early telogen), 56 days after birth (P56, mid-telogen), 69 days after birth (P69, late telogen) and 74 days after birth (P74, early anagen). The expression intensity and localization of Raptor and Rictor at different stages of hair cycle were tested by co-immumostaining.
 Results: Ki-67 immunostaining showed that the time points (P43, P56, P69, P74) and hair follicle stages (early telogen, mid-telogen, late telogen, early anagen) of the dorsal skin were consistent with each other. The results of real-time PCR and immunostaining were consistent, showing that the expression of Raptor and Rictor did not changed in the early-, mid-, late telogen, and early anagen. However, Raptor was specifically expressed in the bulge where hair follicle stem cells (HFSCs) are residing in, and Rictor was mainly detected in inner root sheath (IRS) cells. 
 Conclusion: The expression of Raptor and Rictor does not altered in the hair follicles at different hair follicle stages, but Raptor and Rictor are specifically expressed in the HFSCs and IRS cells, respectively, indicating that Raptor might be a molecular marker for HFSCs, and Rictor might be involved in the maintenance of IRS and formation of hair shaft.
Animals ; Hair ; Hair Follicle ; Mice ; Rapamycin-Insensitive Companion of mTOR Protein ; Raptors ; Skin ; TOR Serine-Threonine Kinases

Immune-mediated hemocytopenia is a common cytopenic diseases without bone marrow hematopoietic abnormalities, the patient's quality of life is significantly reduced when first-line treatments are ineffective. Rapamycin, possesses a clear mechanism of targeting mTOR protein, can upregulate regulatory T cells and induces apoptosis of specific cells, by regulating the lymphocyte subsets, so as to treat various types of immune-mediated hemocytopenia with a certain therapeutic effect. In this reviews, the action mechanism and clinical application of rapamycin in immune thrombocytopenia（ITP）, autoimmune hemolytic anemia（AIHA）, acquired aplastic anemia and autoimmune lymphoproliferative syndrome（ALPS） etc. are summarized.
Anemia, Aplastic ; Humans ; Quality of Life ; Sirolimus ; therapeutic use ; T-Lymphocytes, Regulatory ; TOR Serine-Threonine Kinases

Mammalian target of rapamycin (mTOR) is an intracellular signaling pathway molecule which regulates various fundamental physiological processes. The mTOR signaling pathway plays an important role in synaptic plasticity, information transmission and processing, and neuroregulation. Dysregulation of the mTOR signaling pathway is generally considered to be related to the pathogenesis of autism spectrum disorder (ASD); meanwhile, the mTOR inhibitor can ameliorate the symptoms of ASD. The role of mTOR in the pathogenesis of ASD is summarized in this article to provide a theoretical basis for targeted therapy of ASD.
Animals ; Autism Spectrum Disorder ; Humans ; Signal Transduction ; Sirolimus ; TOR Serine-Threonine Kinases

The mammalian target of rapamycin(mTOR)is a serine/threonine protein kinase that regulates protein synthesis and degradation,cytoskeletal formation,and cell longevity.Autophagy,a catabolic process necessary for the maintenance of intracellular homeostasis,is essential for cell survival,whereas mTOR is the crucial regulator of autophagy.Alzheimer's disease(AD)is the most common cause of progressive dementia in the elderly.It has been shown that disorders of mTOR and autophagy signaling pathways are closely related to AD.In the present review,we describe the regulatory roles of mTOR signaling and autophagy pathway in AD brain and introduce drugs for AD acting via modulation of autophagy and mTOR.
Alzheimer Disease ; pathology ; Autophagy ; Humans ; Signal Transduction ; TOR Serine-Threonine Kinases ; metabolism

OBJECTIVE: This study aimed to investigate the expression patterns and relationship of microtubule-associated protein 1 light chain 3 (LC3B) and mammalian target of rapamycin (mTOR) in oral leukoplakia (OLK) in smokers and never-smokers. This work also analyzed the relationship between smoking and the carcinogenic potential of OLK. METHODS: Immunohistochemistry was used to detect the expression of LC3B and mTOR in 120 patients with OLK. Clinical data from 120 smokers and never-smokers with OLK were analyzed. Subsequently, the relationships among LC3B and mTOR expression, clinical factors, and smoking were analyzed. RESULTS: Smoking and nonsmoking patients with OLK differed in terms of gender, age, lesion location, pathological typing, and carcinogenic situation. The positive rate of LC3B in never-smokers was higher than that in smokers. Whereas the positive rate of mTOR in smokers was higher than that in the corresponding never-smokers, and the differences were statistically significant (P<0.05). Smoking was positively correlated with the positive rate of mTOR (P<0.05), and had no significant correlation with LC3B expression. The positive rates of LC3B and mTOR were negatively correlated with the intensity of smoking (P<0.05). CONCLUSIONS The effect of smoking habits on OLK may be linked to the expression of proteins that are directly associated with autophagy.
Animals ; Autophagy ; Humans ; Leukoplakia, Oral ; Microtubule-Associated Proteins ; Smokers ; TOR Serine-Threonine Kinases

Animals ; Autophagy ; genetics ; Genes ; Humans ; Neoplasms ; genetics ; pathology ; Signal Transduction ; TOR Serine-Threonine Kinases ; metabolism

In the tumor microenvironment, metabolic reprogramming can impact metabolic characteristics of T cells, thus inducing immunosuppression to promote tumor immune escape. The mammalian target of rapamycin (mTOR) signaling pathway plays an important role in regulating diverse functions of various immune cells. This review mainly focuses on the molecular mechanism of mTOR signaling in regulating cellular energy metabolism process, and the activation status of mTOR signaling under different nutritional environments. In addition, it also summarizes the role of the mTOR signaling in regulatory T cell (Tregs) metabolism and function in current studies, and evaluates the potential of mTOR as a clinical immunotherapeutic target and its current application challenges.
Immunosuppression Therapy ; Metabolic Reprogramming ; Signal Transduction ; Sirolimus ; T-Lymphocytes, Regulatory ; TOR Serine-Threonine Kinases ; Humans

This study aimed to investigate the effect of lipopolysaccharide (LPS) on lipophagy in hepatocytes and the underlying mechanism. Human hepatoma cell line HepG2 was cultured in vitro, treated with 0.1 mmol/L palmitic acid (PA), and then divided into control group (0 μg/mL LPS), LPS group (10 μg/mL LPS), LPS+DMSO group and LPS+RAPA (rapamycin, 10 μmol/L) group. Lipid accumulation in hepatocytes was observed by oil red O staining. The autophagic flux of the cells was assessed using confocal laser scanning microscope after being transfected with autophagy double-labeled adenovirus (mRFP-GFP-LC3). The level of intracellular lipophagy was visualized by the colocalization of lipid droplets (BODIPY 493/503 staining) and lysosomes (lysosome marker, lysosomal associated membrane protein 1, LAMP1). The expression levels of mammalian target of rapamycin (mTOR), phosphorylated mTOR (p-mTOR), ribosome protein subunit 6 kinase 1 (S6K1), p-S6K1, LC3II/I and P62 protein were examined by Western blot. The results showed that the number of red lipid droplets stained with oil red O was significantly increased in LPS group compared with that in control group (P < 0.001). Moreover, in LPS group, the number of autophagosomes was increased, while the number of autophagolysosomes and the colocalization rate of LAMP1 and BODIPY were significantly decreased (P < 0.05). Meanwhile, the ratios of p-mTOR/mTOR and p-S6K1/S6K1, the ratio of LC3II/LC3I and the protein expression of P62 were significantly increased (P < 0.05) in LPS group. Furthermore, compared with LPS+DMSO group, RAPA treatment obviously reduced the number of lipid droplets and autophagosomes, and raised the number of autophagolysosomes and the colocalization rate of LAMP1 and BODIPY (P < 0.05). In conclusion, the results demonstrate that LPS inhibits lipophagy in HepG2 cells via activating mTOR signaling pathway, thereby aggravating intracellular lipid accumulation.
Autophagy ; Hep G2 Cells ; Humans ; Lipopolysaccharides ; Palmitic Acid ; Signal Transduction ; TOR Serine-Threonine Kinases