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Toxicon : Official Journal of the... Jun 2024Presynaptic- or β-neurotoxicity of secreted phospholipases A (sPLA) is a complex process. For full expression of β-neurotoxicity, the enzymatic activity of the toxin...
Presynaptic- or β-neurotoxicity of secreted phospholipases A (sPLA) is a complex process. For full expression of β-neurotoxicity, the enzymatic activity of the toxin is essential. However, it has been shown that not all toxic effects of a β-neurotoxin depend on its enzymatic activity, for example, the inhibition of mitochondrial cytochrome c oxidase. The main objective of this study was to verify whether it is possible to observe and study the phospholipase-independent actions of β-neurotoxins by a standard ex vivo twitch-tension experimental approach. To this end, we compared the effects of a potent snake venom β-neurotoxin, ammodytoxin A (AtxA), and its enzymatically inactive mutant AtxA(D49S) on muscle contraction of the mouse phrenic nerve-hemidiaphragm preparation. While AtxA significantly affected the amplitude of the indirectly evoked isometric muscle contraction, the resting tension of the neuromuscular (NM) preparation, the amplitude of the end-plate potential (EPP), the EPP half decay time and the resting membrane potential, AtxA(D49S) without enzymatic activity did not. From this, we can conclude that the effects of AtxA independent of enzymatic activity cannot be studied with classical electrophysiological measurements on the isolated NM preparation. Our results also suggest that the inhibition of cytochrome c oxidase activity by AtxA is not involved in the rapid NM blockade by this β-neurotoxin, but that its pathological consequences are rather long-term. Interestingly, in our experimental setup, AtxA upon direct stimulation reduced the amplitude of muscle contraction and induced contracture of the hemidiaphragm, effects that could be interpreted as myotoxic.
PubMed: 38942241
DOI: 10.1016/j.toxicon.2024.107833 -
Journal of Ethnopharmacology Jun 2024The traditional medicinal formulation, Qifu-yin (QFY), has been widely prescribed for Alzheimer's disease (AD) treatment in China, yet the comprehensive mechanisms...
ETHNOPHARMACOLOGICAL RELEVANCE
The traditional medicinal formulation, Qifu-yin (QFY), has been widely prescribed for Alzheimer's disease (AD) treatment in China, yet the comprehensive mechanisms through which QFY mitigates AD pathology remain to be fully delineated.
AIM OF THE STUDY
This study aimed to explore the therapeutic implications of QFY on the synaptic injury and oxidative stress in the hippocampus of APPswe/PS1dE9 (APP/PS1) mice, with a concerted effort to elucidate the molecular mechanisms related to synaptic preservation and memory improvement.
MATERIALS AND METHODS
The components of QFY were identified by ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). The neuroprotective effects of QFY was evaluated using six-month-old male APP/PS1 mice. Subsequent to a 15 days of QFY regimen, spatial memory was assessed utilizing the Morris water maze (MWM) test. Amyloid-beta (Aβ) aggregation was detected via immunostaining, while the quantification of Aβ and Aβ was achieved through enzyme-linked immunosorbent assay (ELISA). Transmission electron microscopy (TEM) was used to investigate the synaptic structure and mitochondrial morphology. Golgi staining was applied to examine dendritic spine density. Reactive oxygen species (ROS), 3-nitrotyrosine (3-NT) and 4-hydroxy-nonenal (4-HNE) assays were employed to assess oxidative stress. The expression profiles of Aβ metabolism-associated enzymes and the Keap1/Nrf2/ARE signaling pathway were determined by Western blot.
RESULTS
A total of 20 principal compounds in QFY were identified. QFY mitigated memory deficits of APP/PS1 mice, including reducing escape latency and search distance and increasing the time and distance spent in the target quadrant. In addition, QFY increased platform crossings of APP/PS1 mice in the probe trial of MWM tests. TEM analysis showed that QFY increased synapse number in the CA1 region of APP/PS1 mice. Further studies indicated that QFY elevated the expression levels of Post synaptic density protein 95 (PSD95) and synaptophysin, and mitigated the loss of dendritic spine density in the hippocampus of APP/PS1 mice. QFY has been shown to ameliorated the structural abnormalities of mitochondria, including mitochondrial dissolution and degradation, up-regulate ATP synthesis and membrane potential in the hippocampus of APP/PS1 mice. Moreover, QFY activated the Keap1/Nrf2/ARE signaling pathway in the hippocampus of APP/PS1 mice, which might contribute to the neuroprotective effects of QFY.
CONCLUSION
QFY activates the Keap1/Nrf2/ARE signaling, and protects against synaptic and mitochondrial dysfunction in APP/PS1 mice, proposing a potential alternative therapeutic strategy for AD management.
PubMed: 38942156
DOI: 10.1016/j.jep.2024.118497 -
ACS Nano Jun 2024Super-resolution imaging, especially a single-molecule localization approach, has raised a fluorophore engineering revolution chasing sparse single-molecule dark-bright...
Super-resolution imaging, especially a single-molecule localization approach, has raised a fluorophore engineering revolution chasing sparse single-molecule dark-bright blinking transforms. Yet, it is a challenge to structurally devise fluorophores manipulating the single-molecule blinking kinetics. In this pursuit, we have developed a triggering strategy by innovatively integrating the photoactivatable nitroso-caging strategy into self-blinking sulfonamide to form a nitroso-caged sulfonamide rhodamine (NOSR). Our fluorophore demonstrated controllable self-blinking events upon phototriggered caging unit release. This exceptional blink kinetics improved the super-resolution imaging integrity on microtubules compared to self-blinking analogues. With the aid of paramount single-molecule fluorescence kinetics, we successfully reconstructed the ring structure of nuclear pores and the axial morphology of mitochondrial outer membranes. We foresee that our synthetic approach of photoactivation and self-blinking would facilitate rhodamine devising for super-resolution imaging.
PubMed: 38941491
DOI: 10.1021/acsnano.4c03809 -
Biomeditsinskaia Khimiia Jun 2024Type 1 diabetes mellitus (T1DM) is the most severe form of diabetes, which is characterized by absolute insulin deficiency induced by the destruction of pancreatic beta...
Type 1 diabetes mellitus (T1DM) is the most severe form of diabetes, which is characterized by absolute insulin deficiency induced by the destruction of pancreatic beta cells. The aim of this study was to evaluate the effect of a structural analogue of apelin-12 ((NαMe)Arg-Pro-Arg-Leu-Ser-His-Lys-Gly-Pro-Nle-Pro-Phe-OH, metilin) on hyperglycemia, mitochondrial (MCh) respiration in permeabilized cardiac left ventricular (LV) fibers, the myocardial energy state, and cardiomyocyte membranes damage in a model of streptozotocin (STZ) diabetes in rats. Metilin was prepared by solid-phase synthesis using the Fmoc strategy and purified using HPLC. Four groups of animals were used: initial state (IS); control (C), diabetic control (D) and diabetic animals additionally treated with metilin (DM). The following parameters have been studied: blood glucose, MCh respiration in LV fibers, the content of cardiac ATP, ADP, AMP, phosphocreatine (PCr) and creatine (Cr), the activity of creatine kinase-MB (CK-MB) and lactate dehydrogenase (LDH) in blood plasma. Administration of metilin to STZ-treated rats decreased blood glucose, increased state 3 oxygen consumption, the respiratory control ratio in MCh of permeabilized LV fibers, and increased the functional coupling of mitochondrial CK (mt-CK) to oxidative phosphorylation compared with these parameters in group D. In STZ-treated animals metilin administration caused an increase in the PCr content and prevention of the loss of total creatine (ΣCr=PCr+Cr) in the diabetic hearts, as well as restoration of the PCr/ATP ratio in the myocardium and a decrease in the activity of CK-MB and LDH in plasma to initial values. Thus, metilin prevented energy disorders disturbances in cardiomyocytes of animals with experimental T1DM.
Topics: Animals; Diabetes Mellitus, Experimental; Rats; Male; Diabetes Mellitus, Type 1; Energy Metabolism; Intercellular Signaling Peptides and Proteins; Rats, Wistar; Myocytes, Cardiac; Mitochondria, Heart; Blood Glucose; Myocardium; Streptozocin
PubMed: 38940202
DOI: 10.18097/PBMC20247003135 -
Frontiers in Bioscience (Landmark... Jun 2024This study investigated the mechanism by which tazarotene-induced gene 1 (TIG1) inhibits melanoma cell growth. The main focus was to analyze downstream genes regulated...
BACKGROUND
This study investigated the mechanism by which tazarotene-induced gene 1 (TIG1) inhibits melanoma cell growth. The main focus was to analyze downstream genes regulated by TIG1 in melanoma cells and its impact on cell growth.
METHODS
The effects of TIG1 expression on cell viability and death were assessed using water-soluble tetrazolium 1 (WST-1) mitochondrial staining and lactate dehydrogenase release assays. RNA sequencing and Western blot analysis were employed to investigate the genes regulated by TIG1 in melanoma cells. Additionally, the correlation between expression and its downstream genes was analyzed in a melanoma tissue array.
RESULTS
TIG1 expression in melanoma cells was associated with decreased cell viability and increased cell death. RNA-sequencing (RNA-seq), quantitative reverse transcription PCR (reverse RT-QPCR), and immunoblots revealed that TIG1 expression induced the expression of Endoplasmic Reticulum (ER) stress response-related genes such as Homocysteine-responsive endoplasmic reticulum-resident ubiquitin-like domain member 1 (HERPUD1), Binding immunoglobulin protein (BIP), and DNA damage-inducible transcript 3 (DDIT3). Furthermore, analysis of the melanoma tissue array revealed a positive correlation between expression and the expression of , , and . Additionally, attenuation of the ER stress response in melanoma cells weakened the impact of TIG1 on cell growth.
CONCLUSIONS
TIG1 expression effectively hinders the growth of melanoma cells. TIG1 induces the upregulation of ER stress response-related genes, leading to an increase in caspase-3 activity and subsequent cell death. These findings suggest that the ability of retinoic acid to prevent melanoma formation may be associated with the anticancer effect of TIG1.
Topics: Humans; Endoplasmic Reticulum Stress; Melanoma; Cell Line, Tumor; Cell Survival; Gene Expression Regulation, Neoplastic; Cell Death; Apoptosis; Cell Proliferation; Membrane Proteins
PubMed: 38940043
DOI: 10.31083/j.fbl2906233 -
Biomaterials Science Jun 2024The thioredoxin system is involved in cancer development and therefore is a promising target for cancer chemotherapy. Thioredoxin reductase (TrxR) is a key component of...
The thioredoxin system is involved in cancer development and therefore is a promising target for cancer chemotherapy. Thioredoxin reductase (TrxR) is a key component of the thioredoxin (Trx) system, and is overexpressed in many cancers to inhibit apoptosis-related proteins. Alternatively, inhibition of thioredoxin reductase and upregulation of apoptosis factors provide a therapeutic strategy for anti-tumor treatment. In this study, an ultrasound-activatable -organosilica nanomedicine was prepared by integrating chloroquine (CQ) into hollow mesoporous organosilica (CQ@MOS). The -organosilica nanomedicine can inhibit the activity of thioredoxin reductase, elevate cellular reactive oxygen species (ROS) levels, upregulate the pro-apoptotic factors in the c-Jun N-terminal kinase (JNK) apoptosis pathway and induce autophagy inhibition, further resulting in mitochondrial membrane potential (MMP) depolarization and cellular ATP content decrease, ultimately causing significant damage to tumor cells. Moreover, CQ@MOS can efficiently deliver chloroquine into cancer cells and promote an enhanced sonodynamic effect for effective anti-tumor chemotherapy and sonodynamic therapy. This study may enlighten us on a new anti-tumor strategy and suggest its promising applications in cancer treatments.
PubMed: 38939985
DOI: 10.1039/d4bm00583j -
Frontiers in Pharmacology 2024Mitochondria-associated endoplasmic reticulum membranes (MAMs) act as physical membrane contact sites facilitating material exchange and signal transmission between... (Review)
Review
Mitochondria-associated endoplasmic reticulum membranes (MAMs) act as physical membrane contact sites facilitating material exchange and signal transmission between mitochondria and endoplasmic reticulum (ER), thereby regulating processes such as Calipid transport, mitochondrial dynamics, autophagy, ER stress, inflammation, and apoptosis, among other pathological mechanisms. Emerging evidence underscores the pivotal role of MAMs in cardiovascular diseases (CVDs), particularly in aging-related pathologies. Aging significantly influences the structure and function of the heart and the arterial system, possibly due to the accumulation of reactive oxygen species (ROS) resulting from reduced antioxidant capacity and the age-related decline in organelle function, including mitochondria. Therefore, this paper begins by describing the composition, structure, and function of MAMs, followed by an exploration of the degenerative changes in MAMs and the cardiovascular system during aging. Subsequently, it discusses the regulatory pathways and approaches targeting MAMs in aging-related CVDs, to provide novel treatment strategies for managing CVDs in aging populations.
PubMed: 38939842
DOI: 10.3389/fphar.2024.1389202 -
JACS Au Jun 2024The characterization of intrinsically disordered regions (IDRs) in membrane-associated proteins is of crucial importance to elucidate key biochemical processes,...
The characterization of intrinsically disordered regions (IDRs) in membrane-associated proteins is of crucial importance to elucidate key biochemical processes, including cellular signaling, drug targeting, or the role of post-translational modifications. These protein regions pose significant challenges to powerful analytical techniques of molecular structural investigations. We here applied magic angle spinning solid-state nuclear magnetic resonance to quantitatively probe the structural dynamics of IDRs of membrane-bound α-synuclein (αS), a disordered protein whose aggregation is associated with Parkinson's disease (PD). We focused on the mitochondrial binding of αS, an interaction that has functional and pathological relevance in neuronal cells and that is considered crucial for the underlying mechanisms of PD. Transverse and longitudinal N relaxation revealed that the dynamical properties of IDRs of αS bound to the outer mitochondrial membrane (OMM) are different from those of the cytosolic state, thus indicating that regions generally considered not to interact with the membrane are in fact affected by the spatial proximity with the lipid bilayer. Moreover, changes in the composition of OMM that are associated with lipid dyshomeostasis in PD were found to significantly perturb the topology and dynamics of IDRs in the membrane-bound state of αS. Taken together, our data underline the importance of characterizing IDRs in membrane proteins to achieve an accurate understanding of the role that these elusive protein regions play in numerous biochemical processes occurring on cellular surfaces.
PubMed: 38938811
DOI: 10.1021/jacsau.4c00323 -
The EMBO Journal Jun 2024Members of the SLC25 mitochondrial carrier family link cytosolic and mitochondrial metabolism and support cellular maintenance and growth by transporting compounds...
Members of the SLC25 mitochondrial carrier family link cytosolic and mitochondrial metabolism and support cellular maintenance and growth by transporting compounds across the mitochondrial inner membrane. Their monomeric or dimeric state and kinetic mechanism have been a matter of long-standing debate. It is believed by some that they exist as homodimers and transport substrates with a sequential kinetic mechanism, forming a ternary complex where both exchanged substrates are bound simultaneously. Some studies, in contrast, have provided evidence indicating that the mitochondrial ADP/ATP carrier (SLC25A4) functions as a monomer, has a single substrate binding site, and operates with a ping-pong kinetic mechanism, whereby ADP is imported before ATP is exported. Here we reanalyze the oligomeric state and kinetic properties of the human mitochondrial citrate carrier (SLC25A1), dicarboxylate carrier (SLC25A10), oxoglutarate carrier (SLC25A11), and aspartate/glutamate carrier (SLC25A13), all previously reported to be dimers with a sequential kinetic mechanism. We demonstrate that they are monomers, except for dimeric SLC25A13, and operate with a ping-pong kinetic mechanism in which the substrate import and export steps occur consecutively. These observations are consistent with a common transport mechanism, based on a functional monomer, in which a single central substrate-binding site is alternately accessible.
PubMed: 38937634
DOI: 10.1038/s44318-024-00150-0 -
Signal Transduction and Targeted Therapy Jun 2024The ORF9b protein, derived from the nucleocapsid's open-reading frame in both SARS-CoV and SARS-CoV-2, serves as an accessory protein crucial for viral immune evasion by...
The ORF9b protein, derived from the nucleocapsid's open-reading frame in both SARS-CoV and SARS-CoV-2, serves as an accessory protein crucial for viral immune evasion by inhibiting the innate immune response. Despite its significance, the precise regulatory mechanisms underlying its function remain elusive. In the present study, we unveil that the ORF9b protein of SARS-CoV-2, including emerging mutant strains like Delta and Omicron, can undergo ubiquitination at the K67 site and subsequent degradation via the proteasome pathway, despite certain mutations present among these strains. Moreover, our investigation further uncovers the pivotal role of the translocase of the outer mitochondrial membrane 70 (TOM70) as a substrate receptor, bridging ORF9b with heat shock protein 90 alpha (HSP90α) and Cullin 5 (CUL5) to form a complex. Within this complex, CUL5 triggers the ubiquitination and degradation of ORF9b, acting as a host antiviral factor, while HSP90α functions to stabilize it. Notably, treatment with HSP90 inhibitors such as GA or 17-AAG accelerates the degradation of ORF9b, leading to a pronounced inhibition of SARS-CoV-2 replication. Single-cell sequencing data revealed an up-regulation of HSP90α in lung epithelial cells from COVID-19 patients, suggesting a potential mechanism by which SARS-CoV-2 may exploit HSP90α to evade the host immunity. Our study identifies the CUL5-TOM70-HSP90α complex as a critical regulator of ORF9b protein stability, shedding light on the intricate host-virus immune response dynamics and offering promising avenues for drug development against SARS-CoV-2 in clinical settings.
Topics: Humans; Cullin Proteins; SARS-CoV-2; Virus Replication; HSP90 Heat-Shock Proteins; COVID-19; Ubiquitination; HEK293 Cells; Benzoquinones; Protein Stability; Vero Cells; Viral Proteins; Lactams, Macrocyclic
PubMed: 38937432
DOI: 10.1038/s41392-024-01874-5