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Bone & Joint Research Jul 2024The antidiabetic agent metformin inhibits fibrosis in various organs. This study aims to elucidate the effects of hyperglycaemia and metformin on knee joint capsule...
AIMS
The antidiabetic agent metformin inhibits fibrosis in various organs. This study aims to elucidate the effects of hyperglycaemia and metformin on knee joint capsule fibrosis in mice.
METHODS
Eight-week-old wild-type (WT) and type 2 diabetic (db/db) mice were divided into four groups without or with metformin treatment (WT met(-/+), Db met(-/+)). Mice received daily intraperitoneal administration of metformin and were killed at 12 and 14 weeks of age. Fibrosis morphology and its related genes and proteins were evaluated. Fibroblasts were extracted from the capsules of 14-week-old mice, and the expression of fibrosis-related genes in response to glucose and metformin was evaluated in vitro.
RESULTS
The expression of all fibrosis-related genes was higher in Db met(-) than in WT met(-) and was suppressed by metformin. Increased levels of fibrosis-related genes, posterior capsule thickness, and collagen density were observed in the capsules of db/db mice compared with those in WT mice; these effects were suppressed by metformin. Glucose addition increased fibrosis-related gene expression in both groups of mice in vitro. When glucose was added, metformin inhibited the expression of fibrosis-related genes other than cellular communication network factor 2 () in WT mouse cells.
CONCLUSION
Hyperglycaemia promotes fibrosis in the mouse knee joint capsule, which is inhibited by metformin. These findings can help inform the development of novel strategies for treating knee joint capsule fibrosis.
PubMed: 38955349
DOI: 10.1302/2046-3758.137.BJR-2023-0384.R1 -
The Journal of Physical Chemistry. B Jul 2024Protein labeling through transient and repetitive hybridization of short, fluorophore-labeled DNA oligonucleotides has become widely applied in various optical...
Protein labeling through transient and repetitive hybridization of short, fluorophore-labeled DNA oligonucleotides has become widely applied in various optical super-resolution microscopy methods. The main advantages are multitarget imaging and molecular quantification. A challenge is the high background signal originating from the presence of unbound fluorophore-DNA labels in solution. Here, we report the self-quenching of fluorophore dimers conjugated to DNA oligonucleotides as a general concept to reduce the fluorescence background. Upon hybridization, the fluorescence signals of both fluorophores are restored. We expand the toolbox of fluorophores suitable for self-quenching and report their spectra and hybridization equilibria. We apply self-quenched fluorophore-DNA labels to stimulated emission depletion microscopy and single-molecule localization microscopy and report improved imaging performances.
PubMed: 38955346
DOI: 10.1021/acs.jpcb.4c02065 -
Acta Tropica Jun 2024Toxoplasmosis, a zoonotic parasitic disease caused by Toxoplasma gondii (T. gondii), is prevalent worldwide. The fact should be emphasized that a considerable proportion... (Review)
Review
Toxoplasmosis, a zoonotic parasitic disease caused by Toxoplasma gondii (T. gondii), is prevalent worldwide. The fact should be emphasized that a considerable proportion of individuals infected with T. gondii may remain asymptomatic; nevertheless, the condition can have severe implications for pregnant women or immunocompromised individuals. The current treatment of toxoplasmosis primarily relies on medication; however, traditional anti-toxoplasmosis drugs exhibit significant limitations in terms of efficacy, side effects, and drug resistance. The life cycles of T. gondii are characterized by distinct stages and its body morphology goes through dynamic alterations during the growth cycle that are intricately governed by a wide array of post-translational modifications (PTMs). Ubiquitin (Ub) signaling and ubiquitin-like (Ubl) signaling are two crucial post-translational modification pathways within cells, regulating protein function, localization, stability, or interactions by attaching Ub or ubiquitin-like proteins (Ubls) to target proteins. While these signaling mechanisms share some functional similarities, they have distinct regulatory mechanisms and effects. T. gondii possesses both Ub and Ubls and plays a significant role in regulating the parasite's life cycle and maintaining its morphology through PTMs of substrate proteins. Investigating the role and mechanism of protein ubiquitination in T. gondii will provide valuable insights for preventing and treating toxoplasmosis. This review explores the distinctive characteristics of Ub and Ubl signaling in T. gondii, with the aim of inspiring research ideas for the identification of safer and more effective drug targets against toxoplasmosis.
PubMed: 38955322
DOI: 10.1016/j.actatropica.2024.107283 -
Biochimica Et Biophysica Acta. Reviews... Jun 2024The cell division cycle-associated protein (CDCA) family is important in regulating cell division. High CDCA expression is significantly linked to tumor development.... (Review)
Review
The cell division cycle-associated protein (CDCA) family is important in regulating cell division. High CDCA expression is significantly linked to tumor development. This review summarizes clinical and basic studies on CDCAs conducted in recent decades. Furthermore, it systematically introduces the molecular expression and function, key mechanisms, cell cycle regulation, and roles of CDCAs in tumor development, cell proliferation, drug resistance, invasion, and metastasis. Additionally, it presents the latest research on tumor diagnosis, prognosis, and treatment targeting CDCAs. These findings are pivotal for further in-depth studies on the role of CDCAs in promoting tumor development and provide theoretical support for their application as new anti-tumor targets.
PubMed: 38955314
DOI: 10.1016/j.bbcan.2024.189147 -
Biochimica Et Biophysica Acta. General... Jun 2024Diabetic stress acts on the cardiac tissue to induce cardiac hypertrophy and fibrosis. Diabetes induced activated renin angiotensin system (RAS) has been reported to...
BACKGROUND
Diabetic stress acts on the cardiac tissue to induce cardiac hypertrophy and fibrosis. Diabetes induced activated renin angiotensin system (RAS) has been reported to play a critical role in mediating cardiac hypertrophy and fibrosis. Angiotensin converting enzyme (ACE) in producing Angiotensin-II, promotes cardiomyocyte hypertrophy and fibrotic damage. ACE2, a recently discovered molecule structurally homologous to ACE, has been reported to be beneficial in reducing the effect of RAS driven pathologies.
METHODS
In vivo diabetic mouse model was used and co-labelling immunostaining assay have been performed to analyse the fibrotic remodeling and involvement of associated target signaling molecules in mouse heart tissue. For in vitro analyses, qPCR and western blot experiments were performed in different groups for RNA and protein expression analyses.
RESULTS
Fibrosis markers were observed to be upregulated in the diabetic mouse heart tissue as well as in high glucose treated fibroblast and cardiomyocyte cells. Hyperglycemia induced overexpression of YAP1 leads to increased expression of β-catenin (CTNNB1) and ACE with downregulated ACE2 expression. The differential expression of ACE/ACE2 promotes TGFB1-SMAD2/3 pathway in the hyperglycemic cardiomyocyte and fibroblast resulting in increased cardiac fibrotic remodeling.
CONCLUSION
In the following study, we have reported YAP1 modulates the RAS signaling pathway by inducing ACE and inhibiting ACE2 activity to augment cardiomyocyte hypertrophy and fibrosis in hyperglycemic condition. Furthermore, we have shown that hyperglycemia induced dysregulation of ACE-ACE2 activity by YAP1 promotes cardiac fibrosis through β-catenin/TGFB1 dependent pathway.
PubMed: 38955313
DOI: 10.1016/j.bbagen.2024.130666 -
Gene Jun 2024Osteoarthritis (OA) is a progressive condition affecting the joints that lacking effective therapy. However, the underlying molecular mechanism has not been fully...
BACKGROUND
Osteoarthritis (OA) is a progressive condition affecting the joints that lacking effective therapy. However, the underlying molecular mechanism has not been fully clarified.
METHODS
A model of OA was established in Sprague-Dawley (SD) rats through intra-articularly injected with monoiodoacetate (MIA). Western blot analysis was used to identify the levels of UBE2I and hnRNPA2B1 in articular cartilage. Overexpression and siRNA vectors for UBE2I were constructed and transfected into rat chondrocytes. CCK-8, TUNEL and transwell assay were utilized to assess the cell viability, apoptosis and migration ability. Western blot analysis was used to determine the levels of chondrogenic-specific genes including SOX9, COL2A1, Aggrecan, and PRG4. Then, molecular interactions were confirmed by immunoprecipitation.
RESULTS
We observed significant upregulation of UBE2I and hnRNPA2B1 expression in articular cartilage samples of OA. The Pearson correlation analysis revealed positive correlation between UBE2I and hnRNPA2B1 levels. Functional experiments showed that increased UBE2I expression significantly suppressed cell growth, migration, and reduced the expression of chondrogenic-specific genes, while decreasing UBE2I levels had the opposite effects. Molecular interactions between UBE2I and hnRNPA2B1were determined via co-localization and immunoprecipitation. SUMO1 and SUMO3 proteins were enriched by immunoprecipitation using hnRNPA2B1 antibodies. Rescue experiments were performed using SUMOylation inhibitor (2-D08) and SUMOylation activator (N106). Overexpression of UBE2I increased the expression of hnRNPA2B1 in the cytoplasm and decreased the level in the nucleus, which was reversed by the treatment of 2-D08. Conversely, UBE2I knockdown and N106 treatment had the opposite effect.
CONCLUSIONS
UBE2I modulated the nuclear translocation of hnRNPA2B1 by promoting SUMOylation in OA.
PubMed: 38955308
DOI: 10.1016/j.gene.2024.148740 -
Journal of Microbiological Methods Jun 2024Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) is the first-line method for the rapid identification of most cultured...
Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) is the first-line method for the rapid identification of most cultured microorganisms. As for Streptomyces strains, MALDI-TOF MS identification is complicated by the characteristic incrustation of colonies in agar and the strong cell wall of Actinomycetes cells requiring the use of alternative protein extraction protocols. In this study, we developed a specific protocol to overcome these difficulties for the MALDI-TOF MS identification of Actinomycetes made on solid medium. This protocol includes incubation of colony removed from agar plate with the beta-agarase enzyme, followed by a mechanical lysis and two washes by phosphate buffer and ethanol. Twenty-four Streptomyces and two Lentzea strains isolated from Algerian desertic soils were first identified by 16S rRNA sequencing as gold standard method, rpoB gene was used as a secondary gene target when 16S rRNA did not allow species identification. In parallel the isolates were identified by using the MALDI-TOF MS protocol as reported. After the expansion of the database with the inclusion of this MSP, the strains were analyzed again in MALDI Biotyper, and all were identified. This work demonstrates that the rapid identification of Actinomycetes can be obtained without protein extraction step frequently used in MALDI-TOF mass spectrometry with this type of microorganisms.
PubMed: 38955305
DOI: 10.1016/j.mimet.2024.106984 -
Biochimica Et Biophysica Acta.... Jun 2024Many bacterial processes are powered by the sodium motive force (smf) and in case of pathogens, the smf contributes to virulence. Vibrio cholerae, the causative agent of...
Many bacterial processes are powered by the sodium motive force (smf) and in case of pathogens, the smf contributes to virulence. Vibrio cholerae, the causative agent of Cholera disease, possesses a Na-translocating NADH:quinone oxidoreductase (NQR), a six-subunit membrane protein assembly. The 3D structure of NQR revealed the arrangement of the six subunits NqrABCDEF, the position of all redox cofactors (four flavins, two [2Fe-2S] centers) and the binding sites for the substrates NADH (in NqrF) and ubiquinone (in NqrB). Upon oxidation of NADH, electrons are shuttled twice across the membrane, starting with cytoplasmic FAD and electron transfer to the [2Fe2S] cluster and from there to an intra-membranous [2Fe-2S] cluster, periplasmic FMN, FMN and from there to riboflavin. This riboflavin is located at the cytoplasmic entry site of the sodium channel in NqrB, and it donates an electron to ubiquinone-8 positioned at the cytoplasmic aspect of NqrB. Targeting the substrate binding sites of NQR is a promising strategy to identify new inhibitors against many bacterial pathogens. Detailed structural information on the binding mode of natural inhibitors and small molecules in the active sites of NQR is now available, paving the way for the development of new antibiotics. The NQR shows different conformations as revealed in recent cryo-EM and crystallographic studies combined with spectroscopic analyses. These conformations represent distinct steps in the catalytic cycle. Considering the structural and functional data available, we propose a mechanism of Na-NQR based on conformational coupling of electron transfer and Na translocation reaction steps.
PubMed: 38955304
DOI: 10.1016/j.bbabio.2024.149485 -
International Journal of Biological... Jun 2024To improve the techno-functional properties of rapeseed protein (RP), this work tried to regulate the molecular structure of RP via inducing the co-assembly of RP with...
To improve the techno-functional properties of rapeseed protein (RP), this work tried to regulate the molecular structure of RP via inducing the co-assembly of RP with zein and whey protein (WP). The results showed that WP and zein mainly regulate the folding process of RP through hydrophobic and disulfide bonds, thereby altering the structural conformation and forming stable complex RP (CRP). WP addition not only increased the number of surface charges and hydrophilicity of proteins, but also decreased their sizes, improved the water solubility, as well as the availability of active groups. These changes significantly increased the foaming capacity (from 60 % to 147 %) and in vitro gastric digestion rate (from 10 % to 60 %) of CRP. Besides, WP also contributed to the formation of gels and the regulation of their textural profiles. Comparatively, zein improved the hydrophobicity of CRP and balanced degree of intermolecular forces, which effectively increased the emulsifying activity index of CRP from 22 m/g to 90 m/g. Zein decreased the hardness, springiness and water-holding capacity of gel, but increased its gumminess and chewiness. Overall, both WP and zein effectively changed the structural conformation of RP, and improved its techno-functional properties, which provides an effective strategy to modify protein.
PubMed: 38955302
DOI: 10.1016/j.ijbiomac.2024.133441 -
International Journal of Biological... Jun 2024Influenza viruses contribute significantly to the global health burden, necessitating the development of strategies against transmission as well as effective antiviral...
Influenza viruses contribute significantly to the global health burden, necessitating the development of strategies against transmission as well as effective antiviral treatments. The present study reports a biomimetic strategy inspired by the natural antiviral properties of mucins. A bovine serum albumin (BSA) conjugate decorated with the multivalent neuraminidase inhibitor Zanamivir (ZA-BSA) was synthesized using copper-free click chemistry. This synthetic pseudo-mucin exhibited potent neuraminidase inhibitory activity against several influenza strains. Virus capture and growth inhibition assays demonstrated its effective absorption of virion particles and ability to prevent viral infection in nanomolar concentrations. Investigation of the underlying antiviral mechanism of ZA-BSA revealed a dual mode of action, involving disruption of the initial stages of host-cell binding and fusion by inducing viral aggregation, followed by blocking the release of newly assembled virions by targeting neuraminidase activity. Notably, the conjugate also exhibited potent inhibitory activity against oseltamivir-resistant neuraminidase variant comparable to the monomeric Zanamivir. These findings highlight the application of multivalent drug presentation on protein scaffold to mimic mucin adsorption of viruses, together with counteracting drug resistance. This innovative approach has potential for the creation of antiviral agents against influenza and other viral infections.
PubMed: 38955298
DOI: 10.1016/j.ijbiomac.2024.133564