This study aimed to investigate the effects of caffeoylquinic acids from Erigeron breviscapus(EBCQA) on cognitive impairment and mitochondrial dysfunction in Alzheimer's disease(AD), and to explore its underlying mechanisms. The impacts of EBCQA on paralysis, β-amyloid(Aβ) oligomerization, and mRNA expression of mitophagy-related genes [PTEN-induced putative kinase 1(PINK1) homolog-encoding gene pink-1, Parkin homolog-encoding gene pdr-1, Bcl-2 interacting coiled-coil protein 1(Beclin 1) homolog-encoding gene bec-1, microtubule-associated protein 1 light chain 3(LC3) homolog-encoding gene lgg-1, autophagic adapter protein 62(p62) homolog-encoding gene sqst-1] were examined in the AD Caenorhabditis elegans CL4176 model, along with mitochondrial functions including adenosine triphosphate(ATP) content, enzyme activities of mitochondrial respiratory chain complexes Ⅰ,Ⅲ, and Ⅳ, and mitochondrial membrane potential. Additionally, the effects of EBCQA on the green fluorescent protein(GFP)/red fluorescent protein from Discosoma sp.(DsRed) ratio, the expression of phosphatidylethanolamine-modified and GFP-labeled LGG-1(PE-GFP::LGG-1)/GFP-labeled LGG-1(GFP::LGG-1), and GFP-labeled SQST-1(GFP::SQST-1) proteins were investigated in transgenic C. elegans strains. The effect of EBCQA on paralysis was further evaluated after RNA interference(RNAi)-mediated suppression of the pink-1 and pdr-1 genes in CL4176 strain. An AD rat model was established through intraperitoneal injection of D-galactose and intragastric administration of aluminum trichloride. The effects of β-nicotinamide mononucleotide(NMN) and EBCQA on learning and memory ability, neuronal morphology, mitophagy occurrence, mitophagy-related protein expression(PINK1, Parkin, Beclin 1, LC3-Ⅱ/LC3-Ⅰ, p62), and mitochondrial functions(ATP content; enzyme activities of mitochondrial respiratory chain complexes Ⅰ, Ⅲ, and Ⅳ; mitochondrial membrane potential) were investigated in this AD rat model. The results showed that EBCQA delayed paralysis onset in the CL4176 strain, reduced Aβ oligomer formation, and upregulated the mRNA expression levels of lgg-1, bec-1, pink-1, and pdr-1, while downregulating sqst-1 mRNA expression. EBCQA also enhanced ATP content, mitochondrial membrane potential, and the activities of mitochondrial respiratory chain complexes Ⅰ, Ⅲ, and Ⅳ. Furthermore, EBCQA improved the PE-GFP::LGG-1/GFP::LGG-1 ratio, reduced GFP::SQST-1 expression, and decreased the GFP/DsRed ratio. Notably, the ability of EBCQA to delay paralysis was significantly reduced following RNAi-mediated suppression of pink-1 and pdr-1 in CL4176 strain. In AD rats, the administration of NMN or EBCQA significantly improved learning and memory, restored neuronal morphology in the hippocampus, increased autophagosome numbers, and upregulated the expression of PINK1, Parkin, Beclin 1, and the LC3-Ⅱ/LC3-Ⅰ ratio, while reducing p62 expression. Additionally, the treatment with NMN or EBCQA both elevated ATP content, mitochondrial respiratory chain complex Ⅰ, Ⅲ, and Ⅳ activities, and mitochondrial membrane potential in the hippocampus. The above findings indicate that EBCQA improves cognitive impairment and mitochondrial dysfunction in AD, possibly through activation of PINK1/Parkin-mediated mitophagy.
Animals ; Alzheimer Disease/psychology* ; Mitophagy/drug effects* ; Mitochondria/genetics* ; Caenorhabditis elegans/metabolism* ; Ubiquitin-Protein Ligases/genetics* ; Cognitive Dysfunction/physiopathology* ; Rats ; Protein Kinases/genetics* ; Humans ; Male ; Disease Models, Animal ; Caenorhabditis elegans Proteins/genetics* ; Drugs, Chinese Herbal/administration & dosage*

Endosomes are characterized by the presence of various phosphoinositides that are essential for defining the membrane properties. However, the interplay between endosomal phosphoinositides metabolism and innate immunity is yet to be fully understood. Here, our findings highlight the evolutionary continuity of RAB-10/Rab10's involvement in regulating innate immunity. Upon infection of Caenorhabditis elegans with Pseudomonas aeruginosa, an increase in RAB-10 activity was observed in the intestine. Conversely, when RAB-10 was absent, the intestinal diacylglycerols (DAGs) decreased, and the animal's response to the pathogen was impaired. Further research revealed that UNC-16/JIP3 acts as an RAB-10 effector, facilitating the recruitment of phospholipase EGL-8 to endosomes. This leads to a decrease in endosomal phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) and an elevation of DAGs, as well as the activation of the PMK-1/p38 MAPK innate immune pathway. It is noteworthy that the dimerization of UNC-16 is a prerequisite for its interaction with RAB-10(GTP) and the recruitment of EGL-8. Moreover, we ascertained that the rise in RAB-10 activity, due to infection, was attributed to the augmented expression of LET-413/Erbin, and the nuclear receptor NHR-25/NR5A1/2 was determined to be indispensable for this increase. Hence, this study illuminates the significance of endosomal PI(4,5)P2 catabolism in boosting innate immunity and outlines an NHR-25-mediated mechanism for pathogen detection in intestinal epithelia.
Animals ; Caenorhabditis elegans/genetics* ; Endosomes/immunology* ; Caenorhabditis elegans Proteins/immunology* ; Phosphatidylinositol 4,5-Diphosphate/immunology* ; Immunity, Innate ; Pseudomonas aeruginosa/immunology* ; rab GTP-Binding Proteins/genetics* ; Diglycerides/metabolism*

OBJECTIVE: To conduct eukaryotic expression of the leucine-rich repeat containing 15 (LRRC15), a differentially expressed protein in excretory secretory antigens of Taenia solium cysticercus, and predict its antigen epitope. METHODS: The molecular weight, stability, amino acid sequence composition, isoelectric point and T lymphocyte epitope of the LRRC15 protein were predicted using the bioinformatics online softwares ExPASy-PortParam and Protean. The full-length splicing primers were designed using PCR-based accurate synthesis, and the LRRC15 gene was synthesized. The recombinant pcDNA3.4-LRRC15 plasmid was constructed and transfected into HEK293 cells to express the LRRC15 protein. In addition, the LRRC15 protein was characterized by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting. RESULTS: The recombinant pcDNA3.4-LRRC15 plasmid was successfully constructed, which expressed the target LRRC15 protein with an approximately molecular weight of 70 kDa. Bioinformatics prediction with the ExPASy-PortParam software showed that LRRC15 was a hydrophilic protein, which was consisted of 644 amino acids and had a molecular weight of 69.89 kDa and an isoelectric point of 5.6. The molecular formula of the LRRC15 protein was C₃₀₇₃H₄₉₄₂N₈₄₆O₉₅₃S₂₈ and had an instability coefficient is 50.3, indicating that LRRC15 was an instable protein. Bioinformatics prediction with the Protean software showed that the dominant T-cell antigen epitopes were located in 292 to 295, 353 to 361, 521 to 526 and 555 to 564 amino acids of the LRRC15 protein, and the T-cell antigen epitopes with a high hydrophilicity, good flexibility, high surface accessibility and high antigenicity index were found in 122 to 131, 216 to 233, 249 to 254, 333 to 343, 358 to 361, 368 to 372, 384 to 386, 407 to 412, 445 to 450, 469 to 481, 553 to 564, 588 to 594, 607 to 617 and 624 to 639 amino acids. Following transfection of the recombinant pcDNA3.4-LRRC15 plasmid into HEK293 cells, SDS-PAGE and Western blotting identified LRRC15 proteins in cell secretory culture media, cell lysis supernatants and sediments. The LRRC15-His fusion protein was purified from the cell culture medium, and SDS-PAGE identified a remarkable band at approximately 70 kDa, while Western blotting successfully recognized the band of the recombinant LRRC15 protein. CONCLUSIONS The eukaryotic expression and antigen epitope prediction of the LRRC15 protein in the excretory secretory antigens of T. solium cysticercus have been successfully performed, which provides insights into further understandings of its biological functions.
Amino Acids ; Animals ; Antigens, Helminth/genetics* ; Cysticercus/genetics* ; Epitopes/genetics* ; Eukaryota ; HEK293 Cells ; Humans ; Leucine-Rich Repeat Proteins ; Membrane Proteins ; Taenia solium/genetics*

Zn²⁺ is required for the activity of many mitochondrial proteins, which regulate mitochondrial dynamics, apoptosis and mitophagy. However, it is not understood how the proper mitochondrial Zn²⁺ level is achieved to maintain mitochondrial homeostasis. Using Caenorhabditis elegans, we reveal here that a pair of mitochondrion-localized transporters controls the mitochondrial level of Zn²⁺. We demonstrate that SLC-30A9/ZnT9 is a mitochondrial Zn²⁺ exporter. Loss of SLC-30A9 leads to mitochondrial Zn²⁺ accumulation, which damages mitochondria, impairs animal development and shortens the life span. We further identify SLC-25A25/SCaMC-2 as an important regulator of mitochondrial Zn²⁺ import. Loss of SLC-25A25 suppresses the abnormal mitochondrial Zn²⁺ accumulation and defective mitochondrial structure and functions caused by loss of SLC-30A9. Moreover, we reveal that the endoplasmic reticulum contains the Zn²⁺ pool from which mitochondrial Zn²⁺ is imported. These findings establish the molecular basis for controlling the correct mitochondrial Zn²⁺ levels for normal mitochondrial structure and functions.
Animals ; Caenorhabditis elegans/metabolism* ; Cation Transport Proteins/genetics* ; Homeostasis ; Mitochondria/metabolism* ; Zinc/metabolism*

OBJECTIVE: To investigate the changes of lipid metabolism and stress response of adult C.elegans exposed to non-freezing low temperature and explore the possible mechanism. METHODS: The survival rate and activity of adult C.elegans cultured at 20℃ or 4℃ were observed.Lipid metabolism of the cultured adult C.elegans was evaluated using oil red O staining and by detecting the expressions of the genes related with lipid metabolism.The effects of low temperature exposure on stress level of adult C.elegans were evaluated using mitochondrial fluorescence staining and by detecting the expression levels of stress-related genes and antioxidant genes at both the mRNA and protein levels. RESULTS: The lifespan and activity of adult C.elegans exposed to low temperature were significantly reduced with decreased lipid accumulation (P < 0.05) and decreased expressions of genes related with fatty acid synthesis and metabolism (fat-5, fat-6, fat-7, fasn-1, nhr-49, acs-2 and aco-1;P < 0.01).Cold stress significantly increased the expressions of heat shock proteins hsp-70 and hsp16.2(P < 0.05) but lowered the number of mitochondria (P < 0.0001) and the expressions of atfs-1, sod-2, sod-3 and gpx-1(P < 0.05).Knockout of fat-5, nhr-49 or both fat-5 and fat-6 obviously enhanced the sensitivity of C.elegans to cold stress as shown by further reduced activity (P < 0.05) and reduced survival rate at 24 h (P < 0.0001) under cold stress. CONCLUSION Exposure to a low temperature at 4℃ results in lowered lipid metabolism of adult C.elegans accompanied by a decreased mitochondrial number and quality control ability, which triggers high expressions of stress-related genes and causes reduction of antioxidant capacity, thus callsing lowered activity and reduced lifespan of C.elegans.
Animals ; Antioxidants/metabolism* ; Caenorhabditis elegans ; Caenorhabditis elegans Proteins/genetics* ; Cold-Shock Response ; Lipid Metabolism ; Lipid Metabolism Disorders ; Longevity/genetics*

OBJECTIVE: This study explored the rejuvenation mechanisms of Thai polyherbal medicines using different approaches, including in vitro methods, as well as a well-defined nematode model, Caenorhabditis elegans. METHODS: THP-R-SR012 decoction was selected from 23 polyherbal medicines, based on metal-chelating and chain-breaking antioxidant capacities. The influences of this extract on the survival and some stress biomarkers of C. elegans under paraquat-induced oxidative stress were evaluated. Furthermore, lifespan analysis and levels of lipofuscin accumulation were examined in senescent nematodes. The phytochemical profile of THP-R-SR012 was analyzed. RESULTS: Supplementation with THP-R-SR012 decoction significantly increased the mean lifespan and reduced the oxidative damage to C. elegans under oxidative stress conditions. Further, THP-R-SR012 supplementation slightly influenced the lifespan and the level of lipofuscin accumulation during adulthood. Antioxidant-related phytochemical constituents of THP-R-SR012 decoction were rutin, naringenin, 3,4-dihydroxybenzoic acid, gallic acid, glycyrrhizic acid, demethoxycurcumin and 18α-glycyrrhetinic acid. CONCLUSION The antioxidant potential of THP-R-SR012 was due to its scavenging properties, its enhancement of antioxidant-related enzyme activities, and the presence of the antioxidant-related compound. These results support the traditional use of THP-R-SR012 decoction as a tonic for nourishing and strengthening the whole body.
Animals ; Antioxidants/pharmacology* ; Caenorhabditis elegans/metabolism* ; Caenorhabditis elegans Proteins/metabolism* ; Oxidative Stress ; Plant Extracts/pharmacology* ; Reactive Oxygen Species ; Rejuvenation ; Thailand

Oxygen levels are unequal in different living geographical locations of human and related to normal physiology of health. The reduction of oxygen level in the body can lead to a variety of diseases, such as stroke caused by cerebral ischemia and hypoxia. In the recent years, many studies have elucidated the molecular and cellular mechanisms of organism response to different oxygen concentrations by using the nematode Caenorhabditis elegans (C. elegans) as model organism. C. elegans can escape hypoxia or hyperoxia and adapt to the ambient oxygen environments, and there are different response and regulation mechanisms in different degrees of hypoxia environment. In this paper, recent advances in the reaction of nematodes to different oxygen concentrations and the underlying mechanism were reviewed.
Animals ; Caenorhabditis elegans ; Caenorhabditis elegans Proteins ; Humans ; Hypoxia ; Oxygen

In this study, we aimed to evaluate the toxic effects, changes in life span, and expression of various metabolism-related genes in Caenorhabditis elegans, using RNA interference (RNAi) and mutant strains, after 3-bromopyruvate (3-BrPA) treatment. C. elegans was treated with various concentrations of 3-BrPA on nematode growth medium (NGM) plates, and their survival was monitored every 24 h. The expression of genes related to metabolism was measured by the real-time fluorescent quantitative polymerase chain reaction (qPCR). Nematode survival in the presence of 3-BrPA was also studied after silencing three hexokinase (HK) genes. The average life span of C. elegans cultured on NGM with 3-BrPA was shortened to 5.7 d compared with 7.7 d in the control group. hxk-1, hxk-2, and hxk-3 were overexpressed after the treatment with 3-BrPA. After successfully interfering hxk-1, hxk-2, and hxk-3, the 50% lethal concentration (LC₅₀) of all mutant nematodes decreased with 3-BrPA treatment for 24 h compared with that of the control. All the cyp35 genes tested were overexpressed, except cyp-35B3. The induction of cyp-35A1 expression was most obvious. The LC₅₀ values of the mutant strains cyp-35A1, cyp-35A2, cyp-35A4, cyp-35B3, and cyp-35C1 were lower than that of the control. Thus, the toxicity of 3-BrPA is closely related to its effect on hexokinase metabolism in nematodes, and the cyp-35 family plays a key role in the metabolism of 3-BrPA.
Animals ; Caenorhabditis elegans/metabolism* ; Caenorhabditis elegans Proteins/genetics* ; Cytochrome P-450 Enzyme System/genetics* ; Hexokinase/physiology* ; Pyruvates/toxicity* ; RNA, Messenger/analysis*

In order to discover lifespan-extending compounds made from natural resources, activity-guided fractionation of Zingiber officinale Roscoe (Zingiberaceae) ethanol extract was performed using the Caenorhabditis elegans (C. elegans) model system. The compound 6-gingerol was isolated from the most active ethyl acetate soluble fraction, and showed potent longevity-promoting activity. It also elevated the survival rate of worms against stressful environment including thermal, osmotic, and oxidative conditions. Additionally, 6-gingerol elevated the antioxidant enzyme activities of C. elegans, and showed a dose-depend reduction of intracellular reactive oxygen species (ROS) accumulation in worms. Further studies demonstrated that the increased stress tolerance of 6-gingerol-mediated worms could result from the promotion of stress resistance proteins such as heat shock protein (HSP-16.2) and superoxide dismutase (SOD-3). The lipofuscin levels in 6-gingerol treated intestinal worms were decreased in comparison to the control group. No significant 6-gingerol-related changes, including growth, food intake, reproduction, and movement were noted. These results suggest that 6-gingerol exerted longevity-promoting activities independently of these factors and could extend the human lifespan.
Caenorhabditis elegans* ; Caenorhabditis* ; Eating ; Ethanol ; Ginger* ; Heat-Shock Proteins ; Humans ; Lipofuscin ; Longevity* ; Natural Resources ; Reactive Oxygen Species ; Reproduction ; Superoxide Dismutase ; Survival Rate

Sec61β, a subunit of the Sec61 translocon complex, is not essential in yeast and commonly used as a marker of endoplasmic reticulum (ER). In higher eukaryotes, such as Drosophila, deletion of Sec61β causes lethality, but its physiological role is unclear. Here, we show that Sec61β interacts directly with microtubules. Overexpression of Sec61β containing small epitope tags, but not a RFP tag, induces dramatic bundling of the ER and microtubule. A basic region in the cytosolic domain of Sec61β is critical for microtubule association. Depletion of Sec61β induces ER stress in both mammalian cells and Caenorhabditis elegans, and subsequent restoration of ER homeostasis correlates with the microtubule binding ability of Sec61β. Loss of Sec61β causes increased mobility of translocon complexes and reduced level of membrane-bound ribosomes. These results suggest that Sec61β may stabilize protein translocation by linking translocon complex to microtubule and provide insight into the physiological function of ER-microtubule interaction.
Animals ; COS Cells ; Caenorhabditis elegans Proteins ; genetics ; metabolism ; Cell Line, Tumor ; Cercopithecus aethiops ; Endoplasmic Reticulum ; metabolism ; Homeostasis ; Humans ; Microtubules ; metabolism ; SEC Translocation Channels ; deficiency ; genetics ; metabolism