ABSTRACT
Background Acute cadmium (Cd) exposure can cause damage to multiple tissues, with the kidney being the primary target organ. The development of Cd-induced acute kidney injury involves complex mechanisms, in which autophagy and oxidative stress play crucial roles. Objective To investigate the effect of 10-hydroxy-2-decenoic acid (10-HDA) on kidney injury in mice exposed to cadmium, and provide experimental basis for studying the pathogenesis and prevention of Cd poisoning. Methods Thirty-five male C57BL/6 mice were divided into 7 groups (each of 5 mice): control group (normal saline, intraperitoneal injection), CdCl2 group (4 mg·kg−1, intraperitoneal injection), intervention groups ( 4 mg·kg−1 CdCl2, intraperitoneal Injection + 50, 100, 150, or 200 mg·kg−1 10-HDA, oral gavage), and 10-HDA group (150 mg·kg−1, oral gavage). All treatments were given for 14 d. Twenty-four hours after the last infection, physiological indicators [blood urea nitrogen (BUN), creatinine (CRE), malondialdehyde (MDA), and superoxide dismutase (SOD)], histopathological indicators, autophagy-related proteins (Atg7, Atg5, Beclin-1, and LC3), and mitochondrial autophagy-related proteins (PINK1 and Parkin) were detected to examine the effect of 10-HDA on kidney injury caused by CdCl2. Results Compared with the control group, the body weight of mice in the CdCl2 group was significantly reduced (P<0.01); compared with the CdCl2 group, the body weight of mice after intervention with different concentrations of 10-HDA was significantly increased (P<0.01). CdCl2 significantly increased BUN and CRE in the serum samples compared with the control group (P<0.01), which was significantly reduced to varying degrees after 100, 150, and 200 mg·kg−1 10-HDA intervention (P<0.01). MDA significantly increased and SOD significantly decreased in the renal cortex following CdCl2 administration compared with the control group (P<0.01), which was resolved following 10-HDA administration at different concentrations (P<0.01). In histopathological studies, 10-HDA restored injured kidney tissues induced by CdCl2. The expression levels of autophagy proteins Atg7 and LC3-II/I were significantly increased (P<0.05), and the expression level of Beclin-1 was significantly decreased (P<0.05) in the CdCl2 group compared with the control group. The expression levels of Atg7 were reduced to varying degrees after treatment with designed concentrations of 10-HDA, the expression levels of LC3-II/I were also reduced in the 50, 150, and 200 mg·kg−1 10-HDA intervention groups, and the expression levels of Beclin-1 were increased in the 50, 100, and 150 mg·kg−1 10-HDA intervention groups (P<0.05). The expression levels of PINK1 and Parkin in the CdCl2 group and the 50 mg·kg−1 10-HDA intervention group were lower than those in the control group (P<0.01). Compared with the CdCl2 group, the expression levels of PINK1 increased to varying degrees after 100, 150, and 200 mg·kg−1 10-HDA intervention, and the expression levels of Parkin increased in all 10-HDA intervention groups (P<0.01). Conclusion The intervention using 10-HDA can lessen acute kidney injury caused by CdCl2, reduce the expression of autophagy-related proteins, and increase the expression of mitochondrial autophagy-related proteins.
ABSTRACT
Objective:To study the protective effects and mechanisms of melatonin (MTn) on lipopolysaccharide (LPS) and hypoxic-ischemic(HI) induced white matter damage (WMD) in neonatal rats.Methods:Seventy-two 3-day-old newborn Sprague-Dawley (SD) rats were randomly assigned into sham operation group (the sham group), model group (the HI group) and MTn intervention group (the HI+MTn group) ( n=24 for each group). For the sham group, only dissection of the right common carotid artery was performed without ligation. Animal models of WMD were established using LPS pretreatment and HI method in both the HI group and HI+MTn group. The HI+MTn group received MTn intraperitoneal injection (15 mg/kg, 1 h before LPS injection and then once daily). The HI group and the sham group received equal volume of normal saline containing 1% ethanol intraperitoneal injection. The rats were sacrificed on d7 of experiment and periventricular white matter (PVWM) was collected for hematoxylin-eosin (HE) and TUNEL staining to determine WMD and apoptosis. The distribution and morphology of microglial cells in the PVWM were studied using IBA1 immunofluorescence staining. Reactive oxygen species (ROS) kit was used to detect ROS. The expression of nucleotide-binding domain-like receptor protein 3 (NLRP3) inflammasomes, interleukin (IL)-1β, IL-18 and mitochondrial autophagy markers (pink1 and parkin) were determined using real-time quantitative PCR. Results:Compared with the sham group, the HI group showed WMD, cell degeneration and necrosis,increased cell apoptosis and increased expressions of NLRP3 inflammasomes and downstream inflammatory factors (IL-1β and IL-18) in PVWM. Compared with the HI group,the HI+MTn group showed reduced WMD, cell apoptosis, microglia infiltration and inflammatory factors expression. MTn increased pink1 and parkin expression and reduced ROS production in PVWM.Conclusions:MTn reduces ROS production by enhancing mitochondrial autophagy and inhibits NLRP3 inflammasomes hyperactivation to alleviate endotoxin- and HI-induced WMD in neonatal rats.
ABSTRACT
OBJECTIVE@#To explore the protective effect of bloodletting acupuncture at twelve Jing-well points on hand (BAJP) on acute hypobaric hypoxia (AHH)-induced brain injury in rats and its possible mechanisms.@*METHODS@#Seventy-five Sprague Dawley rats were divided into 5 groups by a random number table (n=15), including control, model, BAJP, BAJP+3-methyladenine (3-MA), and bloodletting acupuncture at non-acupoint (BANA, tail tip blooding) groups. After 7-day pre-treatment, AHH models were established using hypobaric oxygen chambers. The levels of S100B, glial fibrillary acidic protein (GFAP), superoxide dismutase (SOD), and malondialdehyde (MDA) in serum were measured by enzyme-linked immunosorbent assay. Hematoxylin-eosin staining and the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling method were used to assess hippocampal histopathology and apoptosis. Transmission electron microscopy assay was used to observe mitochondrial damage and autophagosomes in hippocampal tissues. Flow cytometry was used to detect mitochondrial membrane potential (MMP). The mitochondrial respiratory chain complexes I, III and IV activities and ATPase in hippocampal tissue were evaluated, respectively. Western blot analysis was used to detect the protein expressions of Beclin1, autophagy protein 5 (ATG5), microtubule-associated protein 1 light chain 3 beta (LC3B), phosphatase and tensin homolog induced kinase 1 (PINK1), and Parkin in hippocampal tissues. The mRNA expressions of Beclin1, ATG5 and LC3-II were analyzed by quantitative real-time polymerase chain reaction.@*RESULTS@#BAJP treatment reduced hippocampal tissue injury and inhibited hippocampal cell apoptosis in AHH rats. BAJP reduced oxidative stress by decreasing S100B, GFAP and MDA levels and increasing SOD level in the serum of AHH rats (P<0.05 or P<0.01). Then, BAJP increased MMP, the mitochondrial respiratory chain complexes I, III and IV activities, and the mitochondrial ATPase activity in AHH rats (all P<0.01). BAJP improved mitochondrial swelling and increased the autophagosome number in hippocampal tissue of AHH rats. Moreover, BAJP treatment increased the protein and mRNA expressions of Beclin1 and ATG5 and LC3-II/LC3-I ratio in AHH rats (all P<0.01) and activated the PINK1/Parkin pathway (P<0.01). Finally, 3-MA attenuated the therapeutic effect of BAJP on AHH rats (P<0.05 or P<0.01).@*CONCLUSION@#BAJP was an effective treatment for AHH-induced brain injury, and the mechanism might be through reducing hippocampal tissue injury via increasing the PINK1/Parkin pathway and enhancement of mitochondrial autophagy.
ABSTRACT
Aim To investigate the effects of total saponins from Trillium tschonoskii maxim(TST)on cognitive impairment and mitochondrial autophagy in aging rats induced by D-galactose(D-gal). Methods Male SD rats were randomly divided into normal control group,model group(D-gal,subcutaneous injection),intervention group(TST,low,medium and high dose groups by intragastric administration),with 10 rats in each group,and administered for 6 weeks. Morris water maze was used to evaluate the cognitive function. HE and Nissl staining were used to test the hippocampal and brain cortex morphology. Immunohistochemistry staining was applied to detect the localization expression of Pink1 and Parkin. Western blot was employed to detect the expressions of Pink1,Parkin,LC3-Ⅱ,p62 and Beclin1. Results Compared with the normal control group,the escape latency time was prolonged and the number of crossing platform decreased in D-gal model group(P<0.05). The number of neurons in hippocampus significantly decreased. The positive cells labeled by Pink1 and Parkin staining in hippocampus significantly decreased. The expressions of Pink1,Parkin,LC3-Ⅱ and Beclin1 were markedly reduced,while the expression of p62 was significantly raised(P<0.05). Compared with D-gal model group,the escape latency time of TST dose groups was shortened,the Times of crossing the platform was more,and the time of staying in the platform quadrant increased(P<0.05). The number of neurons in hippocampus significantly increased. The positive cells labeled by Pink1 and Parkin staining in hippocampus significantly increased. The expressions of Pink1,Parkin,LC3-Ⅱ and Beclin1 in hippocampus were apparently up-regulated,while the protein expression of p62 was evidently down-regulated(P<0.05). Conclusions TST has neuroprotective effects on the learning and memory capacities in aging rats induced by D-gal,which may be related to the increasing levels of Pink1,Parkin,LC3-Ⅱ and Beclin1 proteins and the activation of mitochondrial autophagy.
ABSTRACT
Aim To investigate the potential mechanism of osthole promoting autophagy in cervical cancer HeLa cells. Methods HeLa cells were treated with various concentrations of Osthole(0,10,20,40,80,160,240,320 mg·L-1). MTT was used to detect cell vitality. Transmission electron microscopy(TEM)was used to observe the morphology of HeLa cells after osthole intervention. Mondane sulfonyl cadaverine(MDC)staining was used to dectect the level of autophagy. Western blot was employed to analyze the expression levels of mitochondrial protein MFN1 and DPR1. JC-1 flourescence probe was applied to detect mitochondrial membrane potential. Flow cytometry was used to deteminet the release of reactive oxygen species(ROS). A transplanted tumor model of cervical cancer was established in vivo in nude mice. Western blot was used to detect the protein expression levels of PINK1,Parkin and LC3Ⅱ/. Results Osthole could inhibit the proliferation of HeLa cells significantly. Transmission electron microscopy showed that typical autophagosomes were formed in HeLa cells after osthole intervention. The fluorescence intensity of MDC was enhanced. The expression of mitochondrial fusion protein MFN1 was down-regulated after HeLa cells pretreated with osthole,and mitochondrial fission protein DRP1 was up-regulated. Mitochondrial membrane potential decreased. ROS production of HeLa cells was increased by flow cytometry,which could be reversed by autophagy inhibitor 3-MA. Tumor weight in nude mice was inhibited by osthole obviously,which might restrain cervical cancer. Western blot result indicated that the key factors of mitochondrial autophagy PINK1,Parkin and LC3Ⅱ/ratio were up-regulated in HeLa cells. Conclusions Osthole could induce autophagy in HeLa cells and its mechanism may be related to ROS production and PINK1/Parkin pathway.
ABSTRACT
OBJECTIVE@#To summarize the role of chondrocyte mitochondrial homeostasis imbalance in the pathogenesis of osteoarthritis (OA) and analyze its application prospects.@*METHODS@#The recent literature at home and abroad was reviewed to summarize the mechanism of mitochondrial homeostasis imbalance, the relationship between mitochondrial homeostasis imbalance and the pathogenesis of OA, and the application prospect in the treatment of OA.@*RESULTS@#Recent studies have shown that mitochondrial homeostasis imbalance, which is caused by abnormal mitochondrial biogenesis, the imbalance of mitochondrial redox, the imbalance of mitochondrial dynamics, and damaged mitochondrial autophagy of chondrocytes, plays an important role in the pathogenesis of OA. Abnormal mitochondrial biogenesis can accelerate the catabolic reaction of OA chondrocytes and aggravate cartilage damage. The imbalance of mitochondrial redox can lead to the accumulation of reactive oxygen species (ROS), inhibit the synthesis of extracellular matrix, induce ferroptosis and eventually leads to cartilage degradation. The imbalance of mitochondrial dynamics can lead to mitochondrial DNA mutation, decreased adenosine triphosphate production, ROS accumulation, and accelerated apoptosis of chondrocytes. When mitochondrial autophagy is damaged, dysfunctional mitochondria cannot be cleared in time, leading to ROS accumulation, which leads to chondrocyte apoptosis. It has been found that substances such as puerarin, safflower yellow, and astaxanthin can inhibit the development of OA by regulating mitochondrial homeostasis, which proves the potential to be used in the treatment of OA.@*CONCLUSION@#The mitochondrial homeostasis imbalance in chondrocytes is one of the most important pathogeneses of OA, and further exploration of the mechanisms of mitochondrial homeostasis imbalance is of great significance for the prevention and treatment of OA.
Subject(s)
Humans , Reactive Oxygen Species/metabolism , Chondrocytes/metabolism , Osteoarthritis/metabolism , Homeostasis , Mitochondria/metabolism , Cartilage, Articular/metabolismABSTRACT
ObjectiveTo establish a rat model of diabetic wound by feeding on a high-fat and high-sugar diet combined with intraperitoneal injection of streptozotocin （STZ） and surgical preparation of full-thickness skin defects， observe the effect of cinnamaldehyde on the wound healing of diabetes rats， and explore the therapeutic mechanism of cinnamaldehyde in improving wound healing of diabetes rats based on the PTEN-induced putative kinase （PINK1）/Parkin pathway-mediated mitochondrial autophagy. MethodForty-eight male SD rats were randomly divided into blank group （n=12） and diabetes group （n=36）. The diabetes group was further randomly divided into model group， cinnamaldehyde group， and Beifuxin group， with 12 rats in each group. The blank group and the model group received routine disinfection with physiological saline after creating the wounds， while the cinnamaldehyde group received topical application of polyethylene glycol 400 （PEG 400） gel containing 4 μmol·L-1 cinnamaldehyde， and the Beifuxin group received topical application of Beifuxin gel. Dressings were changed once daily. The wound healing rate of each group was observed. On the 7th and 14th days after intervention， the wound tissues of the rats were collected. Hematoxylin and eosin （HE） staining was performed to observe the pathological changes in the local tissues. Immunohistochemistry （IHC） was used to detect the expression of interleukin-6 （IL-6）， tumor necrosis factor-α （TNF-α）， vascular endothelial growth factor （VEGF）， and collagen fibers. Immunofluorescence （IF） and Real-time polymerase chain reaction （Real-time PCR） were used to detect the protein， and mRNA expression of PINK1， Parkin， microtubule-associated protein 1 light chain 3 Ⅱ （LC3 Ⅱ）. ResultAfter intraperitoneal injection of STZ， compared with the blank group， the random blood glucose values of rats in the diabetic group increased significantly （P<0.01）， all higher than 16.7 mmol·L-1， and persistently hyperglycemic for some time after modeling. Compared with the blank group， the model group showed poor growth and healing of granulation tissue in the wounds， and the wound healing rate decreased （P<0.01）. On the 7th day after intervention， the blank group had squamous epithelial coverage on the wounds. Compared with the blank group， the model group only had a small amount of scab at the wound edges， with a large number of infiltrating inflammatory cells in the wounds. The protein expression levels of IL-6 and TNF-α in the tissues increased （P<0.01）， and the protein and mRNA levels of PINK1， Parkin， and LC3Ⅱ decreased （P<0.01）. On the 14th day after the intervention， the granulation tissue in the wounds of the blank group was mature and well-healed. Compared with the blank group， the model group still had infiltrating inflammatory cells and red blood cell exudation. The protein expression levels of VEGF and collagen fibers in the tissues decreased （P<0.01）， and the protein and mRNA expression levels of PINK1， Parkin， and LC3Ⅱ increased （P<0.01）. Compared with the model group， the cinnamaldehyde group and the Beifuxin group showed better wound healing， with increased wound healing rates （P<0.01）. On the 7th day after intervention， the protein expression levels of IL-6 and TNF-α in the tissues decreased （P<0.01）， and the protein and mRNA expression levels of PINK1， Parkin， and LC3Ⅱ increased （P<0.01）. On the 14th day after intervention， the protein expression levels of VEGF and collagen fibers in the tissues increased （P<0.01）， and the protein and mRNA expression levels of PINK1， Parkin， and LC3Ⅱ decreased （P<0.01）. ConclusionCinnamaldehyde can promote the wound healing of diabetes rats by increasing the wound healing rate， reducing the levels of inflammatory factors IL-6 and TNF-α， and increasing the levels of VEGF and collagen fibers. Its mechanism may be related to the regulation of the PINK1/Parkin signaling pathway， activation of mitochondrial autophagy， inhibition of inflammatory responses， and promotion of angiogenesis and collagen synthesis， thereby promoting the wound healing of diabetes rats.
ABSTRACT
Objective To evaluate the effect and mechanism of nicotinamide mononucleotide (NMN) on ischemia-reperfusion injury (IRI) induced by donor liver after cardiac death in rat models. Methods Rat models of orthotopic liver transplantation were established by "magnetic ring + double cuff" method. SD rats were randomly divided into the sham operation group (Sham group), orthotopic liver transplantation group (OLT group), NMN treatment + orthotopic liver transplantation group (NMN group), NMN+sirtuin-3 (Sirt3) inhibitor (3-TYP) + orthotopic liver transplantation group (NMN+3-TYP group), respectively. Pathological changes and hepatocyte apoptosis of the rats were observed in each group. Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels were determined. Superoxide dismutase (SOD) and malondialdehyde (MDA) contents in liver tissues were detected. The expression levels of Sirt3, microtubule-associated protein 1 light chain 3 (LC3)Ⅱ, PTEN-induced putative kinase 1 (PINK1), Parkin and translocase of the outer mitochondrial membrane 20 (TOMM20) in liver tissues were measured. Postoperative survival of the rats in each group was analyzed. Results Compared with the Sham group, serum ALT and AST levels were higher in the OLT group. Compared with the OLT group, the levels of ALT and AST were decreased in the NMN group. Compared with the NMN group, the levels of ALT and AST were increased in the NMN +3-TYP group (all P < 0.05). The liver tissue structure of rats in the Sham group was basically normal. In the OLT group, pathological changes, such as evident congestion, vacuolar degeneration and hepatocyte necrosis, were observed in the liver tissues. Compared with the Sham group, Suzuki score and apoptosis rate were higher in the OLT group. Suzuki score and apoptosis rate in the NMN group were lower than those in the OLT group. Suzuki score and apoptosis rate in the NMN+3-TYP group were higher compared with those in the NMN group (all P < 0.05). Compared with the Sham group, the SOD content was decreased, whereas the MDA content was increased in the OLT group. Compared with the OLT group, the SOD content was increased, whereas the MDA content was decreased in the NMN group. Compared with the NMN group, the SOD content was decreased, whereas the MDA content was increased in the NMN+3-TYP group (all P < 0.05). Compared with the Sham group, the relative expression levels of Sirt3 and TOMM20 proteins were down-regulated, whereas those of PINK1, Parkin and LC3Ⅱproteins were up-regulated in the OLT group. Compared with the OLT group, the relative expression levels of Sirt3, PINK1, Parkin and LC3Ⅱproteins were up-regulated, whereas that of TOMM20 protein was down-regulated in the NMN group. Compared with the NMN group, the relative expression levels of PINK1, Parkin and LC3Ⅱproteins were down-regulated, whereas that of TOMM20 protein was up-regulated in the NMN+3-TYP group (all P < 0.05). In the Sham group, the 7 d survival rate of rats was 100%, 50% in the OLT group, 75% in the NMN group and 58% in the NMN+3-TYP group, respectively. Conclusions NMN may enhance the antioxidative capacity of the liver, induce PINK1/Parkin-mediated mitochondrial autophagy, and alleviate IRI of the liver by up-regulating Sirt3, thereby playing a protective role in the donor liver after cardiac death.
ABSTRACT
Mitochondrial autophagy is a process to clear dysfunctional mitochondria in the cytoplasm to maintain the integrity of mitochondrial function and cell homeostasis. Mitochondrial autophagy is a complex physiological process， which can maintain the balance of mitochondrial quality and quantity， cell survival under starvation and harsh conditions， and the stability of the intracellular environment. Its molecular mechanism involves a variety of proteins. Many factors can induce mitochondrial autophagy， such as starvation， oxidative stress， hypoxia， depolarization， and other stresses. The accumulation of unfolded proteins can also induce mitochondrial autophagy. In recent years， as a research hotspot， the abnormality of mitochondrial morphology and function is closely related to the occurrence of a variety of diseases. The research on mitochondrial autophagy and the pathogenesis of clinical diseases has attracted more attention， such as tumors， cardiovascular diseases， liver diseases， nervous system diseases， and glucose metabolism disorders. It has been found that regulating mitochondrial autophagy may inspire the treatment of some diseases. Meanwhile， clinical researchers have paid more attention to traditional Chinese medicine （TCM）. As revealed by in-depth research， Chinese medicine has a certain value in regulating mitochondrial autophagy. The research on the pathogenesis of mitochondrial autophagy in related diseases and the intervention of Chinese medicine has found that there are many reports on the regulation of mitochondrial autophagy by Chinese medicine in tumors， cardiovascular diseases， and nervous system diseases. However， the mechanism of mitochondrial autophagy， the balance of mitochondrial autophagy， and the difference in the activation or inhibition of mitochondrial autophagy by Chinese medicine remain unclear. The regulation of mitochondrial autophagy has become a new research target strategy of Chinese medicine in the prevention and treatment of diseases. This paper reviewed the available literature in recent years to provide reference materials for the regulation of mitochondrial autophagy by Chinese medicine and ideas for the follow-up research of Chinese medicine in mitochondrial autophagy.
ABSTRACT
Mitochondrial autophagy is a process to clear dysfunctional mitochondria in the cytoplasm to maintain the integrity of mitochondrial function and cell homeostasis. Mitochondrial autophagy is a complex physiological process， which can maintain the balance of mitochondrial quality and quantity， cell survival under starvation and harsh conditions， and the stability of the intracellular environment. Its molecular mechanism involves a variety of proteins. Many factors can induce mitochondrial autophagy， such as starvation， oxidative stress， hypoxia， depolarization， and other stresses. The accumulation of unfolded proteins can also induce mitochondrial autophagy. In recent years， as a research hotspot， the abnormality of mitochondrial morphology and function is closely related to the occurrence of a variety of diseases. The research on mitochondrial autophagy and the pathogenesis of clinical diseases has attracted more attention， such as tumors， cardiovascular diseases， liver diseases， nervous system diseases， and glucose metabolism disorders. It has been found that regulating mitochondrial autophagy may inspire the treatment of some diseases. Meanwhile， clinical researchers have paid more attention to traditional Chinese medicine （TCM）. As revealed by in-depth research， Chinese medicine has a certain value in regulating mitochondrial autophagy. The research on the pathogenesis of mitochondrial autophagy in related diseases and the intervention of Chinese medicine has found that there are many reports on the regulation of mitochondrial autophagy by Chinese medicine in tumors， cardiovascular diseases， and nervous system diseases. However， the mechanism of mitochondrial autophagy， the balance of mitochondrial autophagy， and the difference in the activation or inhibition of mitochondrial autophagy by Chinese medicine remain unclear. The regulation of mitochondrial autophagy has become a new research target strategy of Chinese medicine in the prevention and treatment of diseases. This paper reviewed the available literature in recent years to provide reference materials for the regulation of mitochondrial autophagy by Chinese medicine and ideas for the follow-up research of Chinese medicine in mitochondrial autophagy.
ABSTRACT
OBJECTIVE@#To investigate the inhibitory effect of 27-P-coumayl-ursolic acid (27-P-CAUA), the active ingredient in triterpenoids from the leaves of Ilex latifolia Thunb, against breast cancer cells and explore the underlying mechanism.@*METHODS@#CCK-8 assay was used to assess the changes in viability of breast cancer HCC-1806 cells after 27-P-CAUA treatment for 24, 48, or 72 h. The inhibitory effect of 27-P-CAUA on proliferation of the cells was determined by clonogenic assay. JC-1 was used to detect the changes in mitochondrial membrane potential and flow cytometry was performed for analyzing cell apoptosis following 27-P-CAUA treatment. Immunofluorescence assay was used to observe the expression of cl-caspase-3 and P62 in the treated cells. Western blotting was performed to observe the effect of 27-P-CAUA and chloroquine pretreatment on the expressions of LC3I/II, P62 and HER2 signaling pathway proteins in the cells.@*RESULTS@#The results of CCK-8 and clonogenic assays showed that 27-P-CAUA treatment significantly inhibited the proliferation of HCC-1806 cells (P < 0.01) with IC50 values of 81.473, 48.392 and 18.467 μmol/L at 24, 48, and 72 h, respectively. 27-P-CAUA treatment also caused obvious changes in mitochondrial membrane potential (P < 0.01) and induced cell apoptosis in HCC-1806 cells with a 3.34% increase of the early apoptosis rate. Immunofluorescence assay revealed a significant increase of cl-caspase3 expression in 27-P-CAUA-treated HCC-1806 cells, and treatment with 40 μmol/L 27-P-CAUA resulted in significant cell apoptosis (P < 0.01). 27-P-CAUA obviously reduced the expression of LC3II, caused P62 degradation and induced autophagy in HCC-1806 cells. Chloroquine pretreatment obviously blocked the autophagy-inducing effect of 27-P-CAUA. 27-P-CAUA treatment also inhibited the phosphorylation of HER2 and AKT proteins and progressively lowered the expressions of HER2 and phosphorylated AKT protein in HCC-1806 cells (P < 0.01).@*CONCLUSION@#27-P-CAUA can inhibit the proliferation and induce mitochondrial autophagy and apoptosis of HCC-1806 cells by inhibiting the HER2/PI3K/AKT signaling pathway.
Subject(s)
Female , Humans , Apoptosis , Autophagy , Breast Neoplasms , Carcinoma, Hepatocellular/metabolism , Cell Line, Tumor , Cell Proliferation , Liver Neoplasms/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Signal TransductionABSTRACT
Objective To investigate the expression of mitochondrial autophagy-associated protein PINK1 and Parkin in parotid pleomorphic adenoma (PA) and carcinoma ex pleomorphic adenoma(CA-EX-PA). Methods The expression of PINK1 and Parkin were detected by immunohistochemistry in 24 cases of normal parotid gland tissues, 32 cases of PA tissues and 42 cases of CA-EX-PA tissues. The correlations of PINK1 and Parkin expression with the clinicopathologic characteristics of CA-EX-PA patients were analyzed. Results The positive rates of PINK1 in normal parotid gland, PA and CA-EX-PA tissues were 100%, 91% and 67% respectively; and those of Parkin were 100%, 88% and 52% respectively. The expression rates of PINK1 and Parkin in PA and CA-EX-PA tissues were significantly lower than those in normal tissues (P < 0.05). The expression of PINK1 and Parkin protein were positively correlated in CA-EX-PA tissues (r=0.877, P < 0.01). In CA-EX-PA tissues, the expression of PINK1 and Parkin were associated with tumor invasion, lymph node metastasis and TNM stage (P < 0.05). Conclusion The expression of PINK1 and Parkin are decreased in PA and CA-EX-PA tissues. The decrease of mitochondrial autophagy activity might play important roles in the development, invasion and metastasis of parotid gland tumors.
ABSTRACT
OBJECTIVE：To investig ate the effects of tenuifolin （TEN）on brain mitochondrial autophagy in Aizheimer ’s disease（AD）model mice. METHODS ：Totally 50 male APP/PS1 double transgenic mice were randomly divided into model group，TEN medium-dose+ 3-MA group [TEN 40 mg/（kg·d）+autophagy inhibitor 3-MA 30 mg/（kg·d）] and TEN low-dose ， medium-dose and high-dose groups [ 20，40，80 mg/（kg·d）]，with 10 mice in each group. In addition ，10 wild-type homologous mice were included in normal control group. Administration groups were intragastrically given corresponding drug solution ；normal control group and model group were intragastrically given 0.3％ sodium carboxymethyl cellulose solution ，once a day ，0.01 mL/g， for consecutive 3 months. After last administration ，positive expression [measured by integrated optical density （IOD）] of microtubule associated protein 1 light chain 3（LC3）in neuron was detected ；mRNA expressions of LC3，ubiquitin-binding protein p62，Cathepsin D ，Rab7，phosphatase and tensin homolog deleted on chromosome ten gene-induced putative kinase 1（PINK1） and E 3 ligase（Parkin）as well as protein expressions of LC 3，p62，PINK1 and Parkin were detected in brain mitochondria. RESULTS：Compared with normal control group ，IOD value of LC 3 in neuron as well as mRNA and protein expressions of LC 3， p62，PINK1 and Parkin in brain mitochondria were all increased significantly in model group （P＜0.05 or P＜0.01），while mRNA expressions of Cathepsin D and Rab 7 were decreased significantly （P＜0.05 or P＜0.01）. Compared wit h model group ，IOD values of LC 3（except for TEN low-dose and medium-dose groups ） in neuron ，mRNA expressions of LC 3，Cathepsin D ，Rab7， PINK1（except for TEN low-dose group ）and Parkin （except for TEN low-dose group ） in brain mitochondria as well as protein expressions of LC 3 （except for TEN medium-dose group），PINK1（except fo r TEN high-dose group decreased significantly）and Parkin （except for TEN low-dose group decreased significantly ）were increased significantly in TEN low-dose ， medium-dose and high-dose groups （P＜0.05 or P＜0.01）；mRNA（except for TEN low-dose group ）and protein expressions of p62 were decreased significantly （P＜0.05 or P＜0.01）. Compared with TEN medium-dose group ，the changes of above indexes were inhibited significantly in TEN medium-dose + 3-MA group （P＜0.05 or P＜0.01）. CONCLUSIONS ：TEN can induce mitophagy in brain tissue of AD model mice by activating PINK 1/Parkin signaling pathway and improve lysosome function.
ABSTRACT
Ischemic heart disease (IHD), which has been considered to be exclusively caused by stenosis or occlusion of coronary artery, is a significant cause of morbidity and mortality worldwide. Mitochondrial dysfunction is the main pathological basis of ischemic heart disease and reperfusion injury, and moderate mitochondrial autophagy can selectively remove damage proteins and organelles to maintain intracellular homeostasis, so mitochondrial autophagy is important for maintaining the homeostasis of cardiomyocytes. Natural drugs from plants are widely used in ischemic heart disease. In recent years, more and more natural drugs have been proven to alleviate myocardial cell damage after ischemia/reperfusion through mitochondrial autophagy. This paper reviews the research progress of natural drugs from plants medicines regulating mitochondrial autophagy in the treatment of ischemia heart disease.
ABSTRACT
Liver is the main place of drug metabolism. Mitochondria of hepatocytes are important targets of drug-induced liver injury. Mitochondrial autophagy could maintain the healthy operation of mitochondria in cells and the stable proliferation of cells. Therefore, the use of mitochondrial autophagy to remove damaged mitochondria is an important strategy of anti-drug-induced liver injury. Active ingredients that could enhance mitochondrial autophagy are contained in many traditional Chinese medicines, which could regulate the mitochondrial autophagy to alleviate relevant diseases. However, there are only a few reports on how to accurately and efficiently identify and evaluate such components targeting mitochondria from traditional Chinese medicine. Liquid chromatography-mass spectro-metry(LC-MS) combined with serum pharmacology in vivo can be used to accurately and efficiently find active ingredients of traditional Chinese medicine acting on mitochondrial targets. This paper reviewed the research ideas and methods of traditional Chinese medicine ingredients for increasing the hepatotoxicity of mitochondrial autophagy, in order to provide new ideas and methods for the study of active ingredients of traditional Chinese medicine targeting mitochondria.
Subject(s)
Humans , Chemical and Drug Induced Liver Injury , Drug-Related Side Effects and Adverse Reactions , Drugs, Chinese Herbal/toxicity , Medicine, Chinese Traditional , MitochondriaABSTRACT
We aimed to detect whether the effect of apigenin (Apig) on the myocardial infarction-induced cardiomyocyte injury ofmouse myocardial cells and acute myocardial infarction (AMI) mice was through regulating Parkin expression viamiR-103-1-5p. The myocardial infarction cardiomyocyte model (Hypoxia/reoxygenation) was first constructed, thenthe mouse myocardial cells were treated with Apig, and the expression of miR-103-1-5p was decreased and theexpression of Parkin was increased by qRT-PCR and Western blot. It was confirmed by miRNA pulldown andluciferase reporter system that miR-103-1-5p in mouse myocardial cells can bind to Parkin mRNA and inhibit Parkinexpression. Next, a lentiviral vector silenced Parkin and overexpressing miR-103-1-5p was constructed and transfectedinto Apig-treated cells. Autophagy was detected by mitochondrial autophagy marker proteins [atypical protein kinaseC (aPKC)-interacting protein (p62) and bcl-2/Adenovirus E1B 19-kd interacting protein 3 (BNIP3)] via Western blot,mitochondrial function was detected by JC-1 probe, and apoptosis was detected by flow cytometry. It was confirmedthat Apig regulated mitochondria autophagy through miR-103-1-5p and Parkin, which ultimately affected cardiomyocyte death. Finally, an AMI mouse model was constructed, and then the mice were treated with Apig. Theinfarct size was detected by triphenyl tetrazolium chloride (TTC) staining, and the Apig relieved the myocardialinfarction. The expression of miR-103-1-5p was decreased and the expression of Parkin was increased by qRT-PCRand Western blot. The above results simplified that the cardio protection of Apig and miR-103-1-5p against injury ofmyocardial infarction cardiomyocyte by targeting Parkin. These results provided a novel treatment against myocardialinfarction cardiomyocyte.
ABSTRACT
BACKGROUND: In addition to FUNDC1, Bnip3 and Nix mitochondrial autophagy pathways in mammals that are closely related to the hypoxic environment, Pinkl/Parkin is the main mitochondrial autophagy pathway after brain injury. PINK1 acts upstream of Parkin, and the Pinkl/Parkin pathway is used to mediate loss of mitochondrial surface functional proteins or structural polyubiquitination, and considered as a marker for autophagosome selection, which has an important influence on autophagy-dependent degradation of depolarized mitochondria. OBJECTIVE: To analyze the role of Pinkl/Parkin-mediated mitochondrial autophagy in rats with brain injury after hypertensive intracerebral hemorrhage. METHODS: The study was approved by the Laboratory Animal Ethical Committee of North Sichuan Medical College. Forty-five male Wistar rats of SPF grade were randomly divided into normal, model and autophagy inhibitor groups. The rats in the model and autophagy inhibitor groups were used to prepare hypertensive cerebral hemorrhage model. Before modeling 2 µL of PBS solution containing trimethyl adenine (100 nmol/L) was intraperitoneally injected in the inhibitor group, and the model and the normal groups were intraperitoneally injected with 2 µL of PBS. The water content, mitochondrial autophagy, mitochondrial membrane potential, cerebral infarction, mitochondrial LC3, Parkin protein and PINK1 protein expression in brain tissue were detected. RESULTS AND CONCLUSION: (1) Compared with the normal group, the autophagy staining and mitochondrial staining expression in the model group was increased, and the number of co-localized positive cells was increased (both P < 0.05). Compared with the model group, the autophagy staining and mitochondrial staining expression in the autophagy inhibitor group was decreased, and the number of co-localized positive cells was decreased (both P < 0.05). (2) Compared with the normal group, the infarction proportion of the brain in the model group was increased (P < 0.05). Compared with the model group, the infarction proportion of the brain in the autophagy inhibitor group was increased (P < 0.05). (3) Compared with the normal group, the mitochondrial staining intensity in the model group was decreased, and the autophagy staining intensity was increased (both P < 0.05). Compared with the model group, the mitochondrial staining intensity in the autophagy inhibitor group was decreased, and the autophagy staining intensity was increased (both P < 0.05). (4) Compared with the normal group, the expression levels of PINK1, LC3 and Parkin in the model group were increased (P < 0.05). Compared with the model group, the expression levels of PINK1, LC3 and Parkin in the autophagy inhibitor group were decreased (P < 0.05). In summary, mitochondrial autophagy can alleviate the degree of brain injury after hypertensive intracerebral hemorrhage. The Pinkl/Parkin pathway plays an important role in the mitochondrial autophagy.
ABSTRACT
Objective: To investigate the inhibitory effect of mitochondrial dynamics changes on the tumor growth of transplanted melanoma in the APP/PS1 the mice and its mechanism. Methods: The male C57BL/6J (C57) and APP/PS1 mice were divided into C57 transplantation tumor group (n = 7) and APP/PS1 transplantation tumor group (n = 7). The tumor appearance time was observed and the tumor volume of the mice in two groups were calculated and the growth curves of the mice in two groups were drawn. The morphology of tumor tissue of the mice in two groups was observed under light microscope, the expression levels of mitofusion 2 (Mfn2), dynamin related protein (Drpl), fission 1 (Fis1), ubiquitin and polyubiquitin, LC3- II, PTEN induced putative kinase 1 (PINK1) and Parkin proteins in tumor tissue of the mice in two groups were detected by Western blotting method. Results: The tumor tissue under the skin of the mice in C57 transplantation tumor group and APP/PS1 transplantation tumor group was found with the melanoma granules in the tumor cells. Compared with C57 transplantation tumor group, the tumor appearance time of the mice in APP/PS1 transplantation tumor group was late, and the tumor volume was decreased (P < 0. 0 5); the expression level of Mfn2 protein in tumor tissue of the mice was decreased (P < 0. 0 5); the expression levels of Drpl and Fisl proteins were increased (P < 0. 0 5), and the expression levels of ubiquitin and polyubiqitin, LC3- II, PINK1 and Parkin proteins in tumor tissue of the mice in APP/PS1 transplantation tumor group were all increased (P < 0. 0 5). Conclusion: The growth of tumor in the APP/PS1 mice with transplanted melanoma is slow, and its mechanism may be related to PINK1/Parkin pathway involved in the mitochondrial autophagy.
ABSTRACT
The aim of this paper was to investigate whether the mechanism of salvianolic acid B in protecting H9 c2 cardiomyocytes from hypoxia/reoxygenation injury is related to the regulation of mitochondrial autophagy mediated by NIX. H9 c2 cardiomyocytes were cultured in vitro and divided into normal group, model group and salvianolic acid B group(50 μmol·L~(-1)). Hypoxia/reoxygenation injury model was established by hypoxia for 4 h and reoxygenation for 2 h. In normal group, high glucose DMEM medium was used for culture. Those in model group were cultured with DMEM medium without glucose and oxygen, and no drugs for hypoxia and reoxyge-nation. In salvianolic acid B group, salvianolic acid B prepared by glucose-free DMEM medium was added during hypoxia, and the other process was as same as the model group. The cell viability was evaluated by CCK-8 assay. The leakage of lactate dehydrogenase(LDH) was detected by microplate method. The levels of intracellular reactive oxygen species(ROS) and mitochondrial membrane potential(ΔΨm) were measured by chemical fluorescence method. The level of intracellular adenosine triphosphate(ATP) was mea-sured by fluorescein enzyme method. The autophagy related proteins LC3-Ⅰ, LC3-Ⅱ, apoptosis related protein cleaved caspase-3 and mitochondrial autophagy receptor protein NIX were detected by Western blot. As compared with the normal group, the activity of H9 c2 cardiomyocytes and ATP level were decreased(P<0.05); LDH leakage and ROS production were increased(P<0.01); ΔΨm was decreased(P<0.01); LC3-Ⅱ/LC3-Ⅰ ratio, cleaved caspase-3 and NIX protein expression levels were increased(all P<0.05) in the model group. As compared with the model group, the activity of cells and ΔΨm were significantly increased(P<0.01); ATP level was increased(P<0.05); LDH leakage and ROS generation were decreased(P<0.01); LC3-Ⅱ/LC3-Ⅰ ratio was decreased(P<0.01); cleaved caspase-3 and NIX expression levels were decreased(P<0.05) in the salvianolic acid B group. The protective effect of salvianolic acid B on hypoxia/reoxygenation injury of H9 c2 cardiomyocytes may be associated with inhibiting mitochondrial auto-phagy. The specific mechanism may be related to inhibiting the activation of mitochondrial autophagy mediated by NIX, increasing ΔΨm, reducing ROS production, reducing the expression of cleaved caspase-3, LC3-Ⅱ, and increasing cell viability.
Subject(s)
Humans , Apoptosis , Autophagy , Benzofurans , Cell Hypoxia , Cell Survival , Hypoxia , Myocytes, CardiacABSTRACT
Background@#Acute lung injury (ALI) is characterized by an acute inflammatory process, and oxidative stress in the lung tissue leads to a lack of effective therapeutics. This study aimed to identify whether the overexpression of transcription factor EB (TFEB) regulates mitophagy to protect against lipopolysaccharide (LPS)-induced ALI.@*Methods@#We detected the expression of inflammatory factors, cytochrome c (Cyt.c) and nicotinamide adenine dinucleotide phosphate (NADPH), and autophagy-related proteins and observed the changes in lung histopathology induced by ALI in rats and the changes in the cell ultrastructure of primary alveolar type II epithelial cells induced by changing the expression of TFEB in the context of ALI.@*Results@#The overexpression of TFEB could reduce the expression of proinflammatory factors, such as IL-1 and IL-6, and increase the expression of anti-inflammatory factors, such as IL-10, both in vitro and in vivo. In addition, the overexpression of TFEB could reduce the Cyt.c and NADPH levels both in vivo and in vitro. The overexpression of TFEB could upregulate the expression of autophagy-related proteins, such as lysosomal-associated membrane protein 1 (LAMP1), microtubule-associated protein light chain 3B (LC3B), and Beclin both in vivo and in vitro, and promote mitochondrial autophagy. The overexpression of TFEB significantly improved the histopathologic changes induced by LPS-induced ALI in rats. However, low TFEB expression produced the opposite results.@*Conclusion@#TFEB overexpression can decrease inflammation and mitochondrial damage in the lung tissue and alveolar epithelial cells through regulating mitochondrial autophagy to protect against LPS-induced ALI. Therefore, TFEB is likely a potential therapeutic target in LPS-induced ALI.