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1.
Ecotoxicol Environ Saf ; 275: 116264, 2024 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-38564869

RESUMO

Triocresyl phosphate (TOCP) was commonly used as flame retardant, plasticizer, lubricant, and jet fuel additive. Studies have shown adverse effects of TOCP on the reproductive system. However, the potential harm brought by TOCP, especially to mammalian female reproductive cells, remains a mystery. In this study, we employed an in vitro model for the first time to investigate the effects of TOCP on the maturation process of mouse oocytes. TOCP exposure hampered the meiotic division process, as evidenced by a reduction in the extrusion of the first polar body from oocytes. Subsequent research revealed the disruption of the oocyte cell cytoskeleton induced by TOCP, resulting in abnormalities in spindle organization, chromosome alignment, and actin filament distribution. This disturbance further extended to the rearrangement of organelles within oocytes, particularly affecting the mitochondria. Importantly, after TOCP treatment, mitochondrial function in oocytes was impaired, leading to oxidative stress, DNA damage, cell apoptosis, and subsequent changes of epigenetic modifications. Supplementation with nicotinamide mononucleotide (NMN) alleviated the harmful effects of TOCP. NMN exerted its mitigating effects through two fundamental mechanisms. On one hand, NMN conferred stability to the cell cytoskeleton, thereby supporting nuclear maturation. On the other hand, NMN enhanced mitochondrial function within oocytes, reducing the excess reactive oxygen species (ROS), restoring meiotic division abnormalities caused by TOCP, preventing oocyte DNA damage, and suppressing epigenetic changes. These findings not only enhance our understanding of the molecular basis of TOCP induced oocyte damage but also offer a promising avenue for the potential application of NMN in optimizing reproductive treatment strategies.


Assuntos
Mononucleotídeo de Nicotinamida , Fosfatos , Tritolil Fosfatos , Feminino , Camundongos , Animais , Mononucleotídeo de Nicotinamida/metabolismo , Mononucleotídeo de Nicotinamida/farmacologia , Fosfatos/metabolismo , Oócitos , Citoesqueleto , Mitocôndrias , Espécies Reativas de Oxigênio/metabolismo , Mamíferos
2.
Microb Cell Fact ; 23(1): 105, 2024 Apr 09.
Artigo em Inglês | MEDLINE | ID: mdl-38594656

RESUMO

BACKGROUND: Pregnenolone and progesterone are the life-important steroid hormones regulating essential vital functions in mammals, and widely used in different fields of medicine. Microbiological production of these compounds from sterols is based on the use of recombinant strains expressing the enzyme system cholesterol hydroxylase/C20-C22 lyase (CH/L) of mammalian steroidogenesis. However, the efficiency of the known recombinant strains is still low. New recombinant strains and combination approaches are now needed to produce these steroid hormones. RESULTS: Based on Mycolicibacterium smegmatis, a recombinant strain was created that expresses the steroidogenesis system (CYP11A1, adrenodoxin reductase, adrenodoxin) of the bovine adrenal cortex. The recombinant strain transformed cholesterol and phytosterol to form progesterone among the metabolites. When 3-methoxymethyl ethers of sterols were applied as bioconversion substrates, the corresponding 3-ethers of pregnenolone and dehydroepiandrosterone (DHEA) were identified as major metabolites. Under optimized conditions, the recombinant strain produced 85.2 ± 4.7 mol % 3-methoxymethyl-pregnenolone within 48 h, while production of 3-substituted DHEA was not detected. After the 3-methoxymethyl function was deprotected by acid hydrolysis, crystalline pregnenolone was isolated in high purity (over 98%, w/w). The structures of steroids were confirmed using TLC, HPLC, MS and 1H- and 13C-NMR analyses. CONCLUSION: The use of mycolicybacteria as a microbial platform for the expression of systems at the initial stage of mammalian steroidogenesis ensures the production of valuable steroid hormones-progesterone and pregnenolone from cholesterol. Selective production of pregnenolone from cholesterol is ensured by the use of 3-substituted cholesterol as a substrate and optimization of the conditions for its bioconversion. The results open the prospects for the generation of the new microbial biocatalysts capable of effectively producing value-added steroid hormones.


Assuntos
Fitosteróis , Progesterona , Bovinos , Animais , Pregnenolona/metabolismo , Esteróis , Esteroides , Colesterol/metabolismo , Enzima de Clivagem da Cadeia Lateral do Colesterol/genética , Enzima de Clivagem da Cadeia Lateral do Colesterol/metabolismo , Mamíferos/metabolismo , Éteres
3.
Se Pu ; 42(4): 333-344, 2024 Apr.
Artigo em Chinês | MEDLINE | ID: mdl-38566422

RESUMO

17ß-Estradiol (E2), an important endocrine hormone in the mammalian body, participates in the regulation of the physiological functions of the reproductive system, mammary glands, bone, and cardiovascular system, among others. Paradoxically, despite the physiological actions of endogenous E2 (0.2-1.0 nmol/L), numerous clinical and experimental studies have demonstrated that high-dose E2 treatment can cause tumor regression and exert pro-apoptotic actions in multiple cell types; however, the underlying mechanism remains undescribed. In particular, little information of the cellular processes responding to the lethality of E2 is available. In the present study, we attempted to characterize the cellular processes responding to high-dose (µmol/L) E2 treatment using quantitative phosphoproteomics to obtain a better understanding of the regulatory mechanism of E2-induced cell death. First, the cell phenotype induced by high-dose E2 was determined by performing Cell Counting Kit-8 assay (CCK8), cell cytotoxicity analysis by trypan blue staining, and microscopic imaging on HeLa cells treated with 1-10 µmol/L E2 or dimethyl sulfoxide (DMSO) for 1-3 d. E2 inhibited cell proliferation and induced cell death in a dose- and time-dependent manner. Compared with the DMSO-treated HeLa cells, the cells treated with 5 µmol/L E2 for 2 d demonstrated >74% growth inhibition and approximately 50% cell death. Thus, these cells were used for quantitative phosphoproteomic analysis. Next, a solid-phase extraction (SPE)-based immobilized titanium ion affinity chromatography (Ti4+-IMAC) phosphopeptide-enrichment method coupled with data-independent acquisition (DIA)-based quantitative proteomics was employed for the in-depth screening of high-dose E2-regulated phosphorylation sites to investigate the intracellular processes responding to high-dose E2 treatment. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) identified over 10000 phosphorylation sites regulated by E2 and DMSO in HeLa cells. In comparison with the DMSO-treated cells, the cells treated with 5 µmol/L E2 showed 537 upregulated phosphorylation sites and 387 downregulated phosphorylation sites, with a threshold of p<0.01 and |log2(fold change)|≥1. A total of 924 phosphorylation sites on 599 proteins were significantly regulated by high-dose E2, and these sites were subjected to enrichment analysis. In addition, 453 differently regulated phosphorylation sites on 325 proteins were identified only in the E2- or DMSO-treated cell samples. These phosphorylation sites may be phosphorylated or dephosphorylated in response to high-dose E2 stimulation and were subjected to parallel enrichment analyses. Taken together, 1218 phosphorylation sites on 741 proteins were significantly regulated by high-dose E2 treatment. The functional phosphoproteins in these two groups were then analyzed using Gene Ontology (GO) and Gene Set Enrichment Analysis (GSEA) to determine the biological processes in which they participate and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway database. Consistent with the cell-phenotype data, cell cycle-related proteins were highly enriched in the two groups of E2-regulated phosphoproteins (p<0.05), indicating that high-dose E2 treatment can regulate cell proliferation. In addition, E2-regulated phosphoproteins were highly enriched in the cellular processes of ribosome biogenesis, nucleocytoplasmic transport, and messenger ribonucleic acid (mRNA) processing/splicing (p<0.05), indicating that the activation of these processes may contribute to high-dose E2-induced cell death. These results further confirm that high-dose E2 treatment inhibits protein translation and induces cell death. Furthermore, the significant upregulation of multiple phosphorylation sites associated with epidermal growth factor receptor (EGFR) and mitogen-activated protein kinases (MAPKs) MAPK1, MAPK4, and MAPK14 by high-dose E2 indicates that the EGFR and MAPK signaling pathways are likely involved in the regulation of E2-induced cell death. These phosphorylation sites likely play vital roles in E2-induced cell death in HeLa cells. Overall, our phosphoproteomic data could be a valuable resource for uncovering the regulatory mechanisms of E2 in the micromolar range.


Assuntos
Dimetil Sulfóxido , Espectrometria de Massas em Tandem , Animais , Humanos , Cromatografia Líquida , Células HeLa , Estradiol/farmacologia , Fosfoproteínas/química , Fosfoproteínas/metabolismo , Receptores ErbB/metabolismo , Fosforilação , Mamíferos/metabolismo
4.
PLoS One ; 19(4): e0298242, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38568908

RESUMO

Dinosauria debuted on Earth's stage in the aftermath of the Permo-Triassic Mass Extinction Event, and survived two other Triassic extinction intervals to eventually dominate terrestrial ecosystems. More than 231 million years ago, in the Upper Triassic Ischigualasto Formation of west-central Argentina, dinosaurs were just getting warmed up. At this time, dinosaurs represented a minor fraction of ecosystem diversity. Members of other tetrapod clades, including synapsids and pseudosuchians, shared convergently evolved features related to locomotion, feeding, respiration, and metabolism and could have risen to later dominance. However, it was Dinosauria that radiated in the later Mesozoic most significantly in terms of body size, diversity, and global distribution. Elevated growth rates are one of the adaptations that set later Mesozoic dinosaurs apart, particularly from their contemporary crocodilian and mammalian compatriots. When did the elevated growth rates of dinosaurs first evolve? How did the growth strategies of the earliest known dinosaurs compare with those of other tetrapods in their ecosystems? We studied femoral bone histology of an array of early dinosaurs alongside that of non-dinosaurian contemporaries from the Ischigualasto Formation in order to test whether the oldest known dinosaurs exhibited novel growth strategies. Our results indicate that the Ischigualasto vertebrate fauna collectively exhibits relatively high growth rates. Dinosaurs are among the fastest growing taxa in the sample, but they occupied this niche alongside crocodylomorphs, archosauriformes, and large-bodied pseudosuchians. Interestingly, these dinosaurs grew at least as quickly, but more continuously than sauropodomorph and theropod dinosaurs of the later Mesozoic. These data suggest that, while elevated growth rates were ancestral for Dinosauria and likely played a significant role in dinosaurs' ascent within Mesozoic ecosystems, they did not set them apart from their contemporaries.


Assuntos
Dinossauros , Animais , Dinossauros/anatomia & histologia , Evolução Biológica , Ecossistema , Fósseis , Osso e Ossos , Filogenia , Mamíferos
5.
Nat Commun ; 15(1): 2864, 2024 Apr 05.
Artigo em Inglês | MEDLINE | ID: mdl-38580657

RESUMO

Global climate patterns fundamentally shape the distribution of species and ecosystems. For example, Bergmann's rule predicts that homeothermic animals, including birds and mammals, inhabiting cooler climates are generally larger than close relatives from warmer climates. The modern world, however, lacks the comparative data needed to evaluate such macroecological rules rigorously. Here, we test for Bergmann's rule in Mesozoic dinosaurs and mammaliaforms that radiated within relatively temperate global climate regimes. We develop a phylogenetic model that accounts for biases in the fossil record and allows for variable evolutionary dispersal rates. Our analysis also includes new fossil data from the extreme high-latitude Late Cretaceous Arctic Prince Creek Formation. We find no evidence for Bergmann's rule in Mesozoic dinosaurs or mammaliaforms, the ancestors of extant homeothermic birds and mammals. When our model is applied to thousands of extant dinosaur (bird) and mammal species, we find that body size evolution remains independent of latitude. A modest temperature effect is found in extant, but not in Mesozoic, birds, suggesting that body size evolution in modern birds was influenced by Bergmann's rule during Cenozoic climatic change. Our study provides a general approach for studying macroecological rules, highlighting the fossil record's power to address longstanding ecological principles.


Assuntos
Dinossauros , Animais , Filogenia , Ecossistema , Modelos Biológicos , Tamanho Corporal , Mamíferos , Evolução Biológica
6.
BMC Genomics ; 25(1): 344, 2024 Apr 05.
Artigo em Inglês | MEDLINE | ID: mdl-38580899

RESUMO

BACKGROUND: Genome-wide DNA demethylation occurs in mammalian primordial germ cells (PGCs) as part of the epigenetic reprogramming important for gametogenesis and resetting the epigenetic information for totipotency. Dppa3 (also known as Stella or Pgc7) is highly expressed in mouse PGCs and oocytes and encodes a factor essential for female fertility. It prevents excessive DNA methylation in oocytes and ensures proper gene expression in preimplantation embryos: however, its role in PGCs is largely unexplored. In the present study, we investigated whether or not DPPA3 has an impact on CG methylation/demethylation in mouse PGCs. RESULTS: We show that DPPA3 plays a role in genome-wide demethylation in PGCs even before sex differentiation. Dppa3 knockout female PGCs show aberrant hypermethylation, most predominantly at H3K9me3-marked retrotransposons, which persists up to the fully-grown oocyte stage. DPPA3 works downstream of PRDM14, a master regulator of epigenetic reprogramming in embryonic stem cells and PGCs, and independently of TET1, an enzyme that hydroxylates 5-methylcytosine. CONCLUSIONS: The results suggest that DPPA3 facilitates DNA demethylation through a replication-coupled passive mechanism in PGCs. Our study identifies DPPA3 as a novel epigenetic reprogramming factor in mouse PGCs.


Assuntos
Desmetilação do DNA , Epigênese Genética , Camundongos , Feminino , Animais , Metilação de DNA , Células Germinativas/metabolismo , Genoma , Mamíferos/genética , Proteínas Cromossômicas não Histona/metabolismo
7.
BMC Genomics ; 25(1): 335, 2024 Apr 06.
Artigo em Inglês | MEDLINE | ID: mdl-38580918

RESUMO

BACKGROUND: Mammalian follicle development is characterized by extensive changes in morphology, endocrine responsiveness, and function, providing the optimum environment for oocyte growth, development, and resumption of meiosis. In cattle, the first signs of transcription activation in the oocyte are observed in the secondary follicle, later than during mouse and human oogenesis. While many studies have generated extensive datasets characterizing gene expression in bovine oocytes, they are mostly limited to the analysis of fully grown and matured oocytes. The aim of the present study was to apply single-cell RNA sequencing to interrogate the transcriptome of the growing bovine oocyte from the secondary follicle stage through to the mid-antral follicle stage. RESULTS: Single-cell RNA-seq libraries were generated from oocytes of known diameters (< 60 to > 120 µm), and datasets were binned into non-overlapping size groups for downstream analysis. Combining the results of weighted gene co-expression network and Trendy analyses, and differently expressed genes (DEGs) between size groups, we identified a decrease in oxidative phosphorylation and an increase in maternal -genes and transcription regulators across the bovine oocyte growth phase. In addition, around 5,000 genes did not change in expression, revealing a cohort of stable genes. An interesting switch in gene expression profile was noted in oocytes greater than 100 µm in diameter, when the expression of genes related to cytoplasmic activities was replaced by genes related to nuclear activities (e.g., chromosome segregation). The highest number of DEGs were detected in the comparison of oocytes 100-109 versus 110-119 µm in diameter, revealing a profound change in the molecular profile of oocytes at the end of their growth phase. CONCLUSIONS: The current study provides a unique dataset of the key genes and pathways characteristic of each stage of oocyte development, contributing an important resource for a greater understanding of bovine oogenesis.


Assuntos
Oogênese , Transcriptoma , Feminino , Bovinos , Animais , Humanos , Camundongos , Oogênese/genética , Oócitos/metabolismo , Folículo Ovariano/metabolismo , Proliferação de Células , Mamíferos/genética
8.
Sci Rep ; 14(1): 8039, 2024 04 05.
Artigo em Inglês | MEDLINE | ID: mdl-38580725

RESUMO

This study aimed to characterize the antimicrobial resistance (AMR) and virulence profiles of 67 Escherichia coli isolates obtained from faecal samples of 77 wild mammals from 19 different species, admitted in two rescue and rehabilitation centers in Costa Rica. It was possible to classify 48% (n = 32) of the isolates as multidrug-resistant, and while the highest resistance levels were found towards commonly prescribed antimicrobials, resistance to fluoroquinolones and third generation cephalosporins were also observed. Isolates obtained from samples of rehabilitated animals or animals treated with antibiotics were found to have significantly higher AMR levels, with the former also having a significant association with a multidrug-resistance profile. Additionally, the isolates displayed the capacity to produce α-haemolysins (n = 64, 96%), biofilms (n = 51, 76%) and protease (n = 21, 31%). Our results showed that AMR might be a widespread phenomenon within Costa Rican wildlife and that both free-ranging and rehabilitated wild mammals are potential carriers of bacteria with important resistance and virulence profiles. These results highlight the need to study potential sources of resistance determinants to wildlife, and to determine if wild animals can disseminate resistant bacteria in the environment, potentially posing a significant threat to public health and hindering the implementation of a "One Health" approach.


Assuntos
Infecções por Escherichia coli , Escherichia coli , Animais , Costa Rica , Saúde Pública , Farmacorresistência Bacteriana , Mamíferos , Animais Selvagens/microbiologia , Infecções por Escherichia coli/veterinária , Infecções por Escherichia coli/microbiologia , Antibacterianos/farmacologia , Bactérias , Centros de Reabilitação
9.
Biol Direct ; 19(1): 26, 2024 Apr 06.
Artigo em Inglês | MEDLINE | ID: mdl-38582839

RESUMO

Ischemic stroke is a sudden and acute disease characterized by neuronal death, increment of reactive gliosis (reactive microglia and astrocytes), and a severe inflammatory process. Neuroinflammation is an early event after cerebral ischemia, with microglia playing a leading role. Reactive microglia involve functional and morphological changes that drive a wide variety of phenotypes. In this context, deciphering the molecular mechanisms underlying such reactive microglial is essential to devise strategies to protect neurons and maintain certain brain functions affected by early neuroinflammation after ischemia. Here, we studied the role of mammalian target of rapamycin (mTOR) activity in the microglial response using a murine model of cerebral ischemia in the acute phase. We also determined the therapeutic relevance of the pharmacological administration of rapamycin, a mTOR inhibitor, before and after ischemic injury. Our data show that rapamycin, administered before or after brain ischemia induction, reduced the volume of brain damage and neuronal loss by attenuating the microglial response. Therefore, our findings indicate that the pharmacological inhibition of mTORC1 in the acute phase of ischemia may provide an alternative strategy to reduce neuronal damage through attenuation of the associated neuroinflammation.


Assuntos
Isquemia Encefálica , Microglia , Camundongos , Animais , Alvo Mecanístico do Complexo 1 de Rapamicina , Doenças Neuroinflamatórias , Isquemia Encefálica/tratamento farmacológico , Isquemia Encefálica/genética , Serina-Treonina Quinases TOR/uso terapêutico , Isquemia , Sirolimo/farmacologia , Sirolimo/uso terapêutico , Mamíferos
10.
Sci Rep ; 14(1): 8103, 2024 04 06.
Artigo em Inglês | MEDLINE | ID: mdl-38582880

RESUMO

Antimicrobial resistance genes (ARG), such as extended-spectrum ß-lactamase (ESBL) and carbapenemase genes, are commonly carried on plasmids. Plasmids can transmit between bacteria, disseminate globally, and cause clinically important resistance. Therefore, targeting plasmids could reduce ARG prevalence, and restore the efficacy of existing antibiotics. Cobalt complexes possess diverse biological activities, including antimicrobial and anticancer properties. However, their effect on plasmid conjugation has not been explored yet. Here, we assessed the effect of four previously characterised bis(N-picolinamido)cobalt(II) complexes lacking antibacterial activity on plasmid conjugation in Escherichia coli and Klebsiella pneumoniae. Antimicrobial susceptibility testing of these cobalt complexes confirmed the lack of antibacterial activity in E. coli and K. pneumoniae. Liquid broth and solid agar conjugation assays were used to screen the activity of the complexes on four archetypical plasmids in E. coli J53. The cobalt complexes significantly reduced the conjugation of RP4, R6K, and R388 plasmids, but not pKM101, on solid agar in E. coli J53. Owing to their promising activity, the impact of cobalt complexes was tested on the conjugation of fluorescently tagged extended-spectrum ß-lactamase encoding pCTgfp plasmid in E. coli and carbapenemase encoding pKpQILgfp plasmid in K. pneumoniae, using flow cytometry. The complexes significantly reduced the conjugation of pKpQILgfp in K. pneumoniae but had no impact on pCTgfp conjugation in E. coli. The cobalt complexes did not have plasmid-curing activity, suggesting that they target conjugation rather than plasmid stability. To our knowledge, this is the first study to report reduced conjugation of clinically relevant plasmids with cobalt complexes. These cobalt complexes are not cytotoxic towards mammalian cells and are not antibacterial, therefore they could be optimised and employed as inhibitors of plasmid conjugation.


Assuntos
Anti-Infecciosos , Infecções por Klebsiella , Animais , Escherichia coli/genética , Klebsiella pneumoniae/genética , Ágar/farmacologia , beta-Lactamases/genética , beta-Lactamases/farmacologia , Antibacterianos/farmacologia , Plasmídeos/genética , Anti-Infecciosos/farmacologia , Infecções por Klebsiella/microbiologia , Testes de Sensibilidade Microbiana , Mamíferos/genética
11.
Proc Natl Acad Sci U S A ; 121(15): e2322563121, 2024 Apr 09.
Artigo em Inglês | MEDLINE | ID: mdl-38557192

RESUMO

Mammalian switch/sucrose nonfermentable (mSWI/SNF) ATPase degraders have been shown to be effective in enhancer-driven cancers by functioning to impede oncogenic transcription factor chromatin accessibility. Here, we developed AU-24118, an orally bioavailable proteolysis-targeting chimera (PROTAC) degrader of mSWI/SNF ATPases (SMARCA2 and SMARCA4) and PBRM1. AU-24118 demonstrated tumor regression in a model of castration-resistant prostate cancer (CRPC) which was further enhanced with combination enzalutamide treatment, a standard of care androgen receptor (AR) antagonist used in CRPC patients. Importantly, AU-24118 exhibited favorable pharmacokinetic profiles in preclinical analyses in mice and rats, and further toxicity testing in mice showed a favorable safety profile. As acquired resistance is common with targeted cancer therapeutics, experiments were designed to explore potential mechanisms of resistance that may arise with long-term mSWI/SNF ATPase PROTAC treatment. Prostate cancer cell lines exposed to long-term treatment with high doses of a mSWI/SNF ATPase degrader developed SMARCA4 bromodomain mutations and ABCB1 (ATP binding cassette subfamily B member 1) overexpression as acquired mechanisms of resistance. Intriguingly, while SMARCA4 mutations provided specific resistance to mSWI/SNF degraders, ABCB1 overexpression provided broader resistance to other potent PROTAC degraders targeting bromodomain-containing protein 4 and AR. The ABCB1 inhibitor, zosuquidar, reversed resistance to all three PROTAC degraders tested. Combined, these findings position mSWI/SNF degraders for clinical translation for patients with enhancer-driven cancers and define strategies to overcome resistance mechanisms that may arise.


Assuntos
Adenosina Trifosfatases , Neoplasias de Próstata Resistentes à Castração , Masculino , Humanos , Ratos , Camundongos , Animais , Adenosina Trifosfatases/genética , Adenosina Trifosfatases/metabolismo , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/genética , Linhagem Celular , Cromatina , Mamíferos/genética , Antagonistas de Receptores de Andrógenos , DNA Helicases/genética , Proteínas Nucleares/genética , Fatores de Transcrição/genética
12.
Molecules ; 29(7)2024 Mar 22.
Artigo em Inglês | MEDLINE | ID: mdl-38611696

RESUMO

Collagens are fundamental constituents of the extracellular matrix and are the most abundant proteins in mammals. Collagens belong to the family of fibrous or fiber-forming proteins that self-assemble into fibrils that define their mechanical properties and biological functions. Up to now, 28 members of the collagen superfamily have been recognized. Collagen biosynthesis occurs in the endoplasmic reticulum, where specific post-translational modification-glycosylation-is also carried out. The glycosylation of collagens is very specific and adds ß-d-galactopyranose and ß-d-Glcp-(1→2)-d-Galp disaccharide through ß-O-linkage to hydroxylysine. Several glycosyltransferases, namely COLGALT1, COLGALT2, LH3, and PGGHG glucosidase, were associated the with glycosylation of collagens, and recently, the crystal structure of LH3 has been solved. Although not fully understood, it is clear that the glycosylation of collagens influences collagen secretion and the alignment of collagen fibrils. A growing body of evidence also associates the glycosylation of collagen with its functions and various human diseases. Recent progress in understanding collagen glycosylation allows for the exploitation of its therapeutic potential and the discovery of new agents. This review will discuss the relevant contributions to understanding the glycosylation of collagens. Then, glycosyltransferases involved in collagen glycosylation, their structure, and catalytic mechanism will be surveyed. Furthermore, the involvement of glycosylation in collagen functions and collagen glycosylation-related diseases will be discussed.


Assuntos
Colágeno , Glicosiltransferases , Humanos , Animais , Glicosilação , Matriz Extracelular , Processamento de Proteína Pós-Traducional , Mamíferos
13.
Molecules ; 29(7)2024 Mar 24.
Artigo em Inglês | MEDLINE | ID: mdl-38611729

RESUMO

Royal jelly (RJ) is recognized as beneficial to mammalian health. Multilineage differentiation potential is an important property of mesenchymal stem cells (MSCs). C2C12 cells have an innate ability to differentiate into myogenic cells. Like MSCs, C2C12 cells can also differentiate into osteoblast- and adipocyte-lineage cells. We recently reported that RJ enhances the myogenic differentiation of C2C12 cells. However, the effect of RJ on osteoblast or adipocyte differentiation is still unknown. Here in this study, we have examined the effect of RJ on the osteoblast and adipocyte differentiation of C2C12 cells. Protease-treated RJ was used to reduce the adverse effects caused by RJ supplementation. To induce osteoblast or adipocyte differentiation, cells were treated with bone morphogenetic proteins (BMP) or peroxisome proliferator-activated receptor γ (PPARγ) agonist, respectively. RNA-seq was used to analyze the effect of RJ on gene expression. We found that RJ stimulates osteoblast and adipocyte differentiation. RJ regulated 279 genes. RJ treatment upregulated glutathione-related genes. Glutathione, the most abundant antioxidative factor in cells, has been shown to promote osteoblast differentiation in MSC and MSC-like cells. Therefore, RJ may promote osteogenesis, at least in part, through the antioxidant effects of glutathione. RJ enhances the differentiation ability of C2C12 cells into multiple lineages, including myoblasts, osteoblasts, and adipocytes.


Assuntos
Antioxidantes , Ácidos Graxos , Animais , Diferenciação Celular , Glutationa , Mioblastos , Mamíferos
14.
Molecules ; 29(7)2024 Apr 03.
Artigo em Inglês | MEDLINE | ID: mdl-38611878

RESUMO

Exosomes are multifunctional, cell-derived nanoscale membrane vesicles. Exosomes derived from certain mammalian cells have been developed as angiogenesis promoters for the treatment of myocardial ischemia-reperfusion injury, as they possess the capability to enhance endothelial cell proliferation, migration, and angiogenesis. However, the low yield of exosomes derived from mammalian cells limits their clinical applications. Therefore, we chose to extract exosome-like nanoparticles from the traditional Chinese medicine Salvia miltiorrhiza, which has been shown to promote angiogenesis. Salvia miltiorrhiza-derived exosome-like nanoparticles offer advantages, such as being economical, easily obtainable, and high-yielding, and have an ideal particle size, Zeta potential, exosome-like morphology, and stability. Salvia miltiorrhiza-derived exosome-like nanoparticles can enhance the cell viability of Human Umbilical Vein Endothelial Cells and can promote cell migration and improve the neovascularization of the cardiac tissues of myocardial ischemia-reperfusion injury, indicating their potential as angiogenesis promoters for the treatment of myocardial ischemia-reperfusion injury.


Assuntos
Exossomos , Traumatismo por Reperfusão Miocárdica , Nanopartículas , Salvia miltiorrhiza , Humanos , Animais , 60489 , Traumatismo por Reperfusão Miocárdica/tratamento farmacológico , Células Endoteliais da Veia Umbilical Humana , Fatores de Transcrição , Mamíferos
15.
Molecules ; 29(7)2024 Apr 04.
Artigo em Inglês | MEDLINE | ID: mdl-38611899

RESUMO

2,6-Diaryl-4H-tetrahydro-thiopyran-4-ones and corresponding sulfoxide and sulfone derivatives were designed to lower the major toxicity of their parent anti-kinetoplatidal diarylideneacetones through a prodrug effect. Novel diastereoselective methodologies were developed and generalized from diarylideneacetones and 2,6-diaryl-4H-tetrahydro-thiopyran-4-ones to allow the introduction of a wide substitution profile and to prepare the related S-oxides. The in vitro biological activity and selectivity of diarylideneacetones, 2,6-diaryl-4H-tetrahydro-thiopyran-4-ones, and their S-sulfoxide and sulfone metabolites were evaluated against Trypanosoma brucei brucei, Trypanosoma cruzi, and various Leishmania species in comparison with their cytotoxicity against human fibroblasts hMRC-5. The data revealed that the sulfides, sulfoxides, and sulfones, in which the Michael acceptor sites are temporarily masked, are less toxic against mammal cells while the anti-trypanosomal potency was maintained against T. b. brucei, T. cruzi, L. infantum, and L. donovani, thus confirming the validity of the prodrug strategy. The mechanism of action is proposed to be due to the involvement of diarylideneacetones in cascades of redox reactions involving the trypanothione system. After Michael addition of the dithiol to the double bonds, resulting in an elongated polymer, the latter-upon S-oxidation, followed by syn-eliminations-fragments, under continuous release of reactive oxygen species and sulfenic/sulfonic species, causing the death of the trypanosomal parasites in the micromolar or submicromolar range with high selectivity indexes.


Assuntos
Doença de Chagas , Pró-Fármacos , Piranos , Safrol/análogos & derivados , Compostos de Sulfidrila , Humanos , Animais , Óxidos , Oxirredução , Mamíferos
16.
Int J Mol Sci ; 25(7)2024 Mar 22.
Artigo em Inglês | MEDLINE | ID: mdl-38612409

RESUMO

Limonoids are extremely diversified in plants, with many categories of products bearing an intact, rearranged or fragmented oxygenated scaffold. A specific subgroup of fragmented or degraded limonoids derives from the tetranortriterpenoid prieurianin, initially isolated from the tree Trichilia prieuriana but also found in other plants of the Meliaceae family, including the more abundant species Aphanamixis polystachya. Prieurianin-type limonoids include about seventy compounds, among which are dregeanin and rohitukin. Prieurianin and analogs exhibit insecticidal, antimicrobial, antiadipogenic and/or antiparasitic properties but their mechanism of action remains ill-defined at present. Previous studies have shown that prieurianin, initially known as endosidin 1, stabilizes the actin cytoskeleton in plant and mammalian cells via the modulation of the architecture and dynamic of the actin network, most likely via interference with actin-binding proteins. A new mechanistic hypothesis is advanced here based on the recent discovery of the targeting of the chaperone protein Hsp47 by the fragmented limonoid fraxinellone. Molecular modeling suggested that prieurianin and, to a lesser extent dregeanin, can form very stable complexes with Hsp47 at the protein-collagen interface. Hsp-binding may account for the insecticidal action of the product. The present review draws up a new mechanistic portrait of prieurianin and provides an overview of the pharmacological properties of this atypical limonoid and its chemical family.


Assuntos
Inseticidas , Limoninas , Meliaceae , Animais , Limoninas/farmacologia , Citoesqueleto de Actina , Actinas , Antiparasitários , Inseticidas/farmacologia , Mamíferos
17.
Int J Mol Sci ; 25(7)2024 Mar 23.
Artigo em Inglês | MEDLINE | ID: mdl-38612422

RESUMO

As compounds of natural origin enter human body, it is necessary to investigate their possible interactions with the metabolism of drugs and xenobiotics in general, namely with the cytochrome P450 (CYP) system. Phytic acid (myo-inositol hexaphosphoric acid, IP6) is mainly present in plants but is also an endogenous compound present in mammalian cells and tissues. It has been shown to exhibit protective effect in many pathological conditions. For this paper, its interaction with CYPs was studied using human liver microsomes, primary human hepatocytes, the HepG2 cell line, and molecular docking. Docking experiments and absorption spectra demonstrated the weak ability of IP6 to interact in the heme active site of CYP1A. Molecular docking suggested that IP6 preferentially binds to the protein surface, whereas binding to the active site of CYP1A2 was found to be less probable. Subsequently, we investigated the ability of IP6 to modulate the metabolism of xenobiotics for both the mRNA expression and enzymatic activity of CYP1A enzymes. Our findings revealed that IP6 can slightly modulate the mRNA levels and enzyme activity of CYP1A. However, thanks to the relatively weak interactions of IP6 with CYPs, the chances of the mechanisms of clinically important drug-drug interactions involving IP6 are low.


Assuntos
Ácido Fítico , Xenobióticos , Humanos , Animais , Simulação de Acoplamento Molecular , Sistema Enzimático do Citocromo P-450 , RNA Mensageiro , Mamíferos
18.
Int J Mol Sci ; 25(7)2024 Mar 25.
Artigo em Inglês | MEDLINE | ID: mdl-38612449

RESUMO

Stress granules (SGs) are membraneless ribonucleoprotein (RNP)-based cellular foci formed in response to stress, facilitating cell survival by protecting against damage. Mammalian spermatogenesis should be maintained below body temperature for proper development, indicating its vulnerability to heat stress (HS). In this study, biotin tracer permeability assays showed that the inhibition of heat-induced SG assembly in the testis by 4-8 mg/kg cycloheximide significantly increased the percentage of seminiferous tubules with a damaged blood-testis barrier (BTB). Western blot results additionally revealed that the suppression of heat-induced SG assembly in Sertoli cell line, TM4 cells, by RNA inference of G3bp1/2 aggravated the decline in the BTB-related proteins ZO-1, ß-Catenin and Claudin-11, indicating that SGs could protect the BTB against damage caused by HS. The protein components that associate with SGs in Sertoli cells were isolated by sequential centrifugation and immunoprecipitation, and were identified by liquid chromatography with tandem mass spectrometry. Gene Ontology and KEGG pathway enrichment analysis revealed that their corresponding genes were mainly involved in pathways related to proteasomes, nucleotide excision repair, mismatch repair, and DNA replication. Furthermore, a new SG component, the ubiquitin associated protein 2 (UBAP2), was found to translocate to SGs upon HS in TM4 cells by immunofluorescence. Moreover, SG assembly was significantly diminished after UBAP2 knockdown by RNA inference during HS, suggesting the important role of UBAP2 in SG assembly. In addition, UBAP2 knockdown reduced the expression of ZO-1, ß-Catenin and Claudin-11, which implied its potential role in the function of the BTB. Overall, our study demonstrated the role of SGs in maintaining BTB functions during HS and identified a new component implicated in SG formation in Sertoli cells. These findings not only offer novel insights into the biological functions of SGs and the molecular mechanism of low fertility in males in summer, but also potentially provide an experimental basis for male fertility therapies.


Assuntos
Barreira Hematotesticular , DNA Helicases , Masculino , Animais , Camundongos , Proteínas de Ligação a Poli-ADP-Ribose , RNA Helicases , Proteínas com Motivo de Reconhecimento de RNA , Grânulos de Estresse , beta Catenina , RNA , Claudinas , Mamíferos
19.
Proc Natl Acad Sci U S A ; 121(16): e2313820121, 2024 Apr 16.
Artigo em Inglês | MEDLINE | ID: mdl-38598343

RESUMO

In primates, high-acuity vision is mediated by the fovea, a small specialized central region of the retina. The fovea, unique to the anthropoid lineage among mammals, undergoes notable neuronal morphological changes during postnatal maturation. However, the extent of cellular similarity across anthropoid foveas and the molecular underpinnings of foveal maturation remain unclear. Here, we used high-throughput single-cell RNA sequencing to profile retinal cells of the common marmoset (Callithrix jacchus), an early divergent in anthropoid evolution from humans, apes, and macaques. We generated atlases of the marmoset fovea and peripheral retina for both neonates and adults. Our comparative analysis revealed that marmosets share almost all their foveal types with both humans and macaques, highlighting a conserved cellular structure among primate foveas. Furthermore, by tracing the developmental trajectory of cell types in the foveal and peripheral retina, we found distinct maturation paths for each. In-depth analysis of gene expression differences demonstrated that cone photoreceptors and Müller glia (MG), among others, show the greatest molecular divergence between these two regions. Utilizing single-cell ATAC-seq and gene-regulatory network inference, we uncovered distinct transcriptional regulations differentiating foveal cones from their peripheral counterparts. Further analysis of predicted ligand-receptor interactions suggested a potential role for MG in supporting the maturation of foveal cones. Together, these results provide valuable insights into foveal development, structure, and evolution.


Assuntos
Callithrix , Retina , Humanos , Animais , Recém-Nascido , Callithrix/anatomia & histologia , Retina/metabolismo , Fóvea Central/fisiologia , Células Fotorreceptoras Retinianas Cones , Macaca , Mamíferos
20.
Sci Adv ; 10(15): eadl5952, 2024 Apr 12.
Artigo em Inglês | MEDLINE | ID: mdl-38598639

RESUMO

N-methyl-d-aspartate receptors (NMDARs) and other ionotropic glutamate receptors (iGluRs) mediate most of the excitatory signaling in the mammalian brains in response to the neurotransmitter glutamate. Uniquely, NMDARs composed of GluN1 and GluN3 are activated exclusively by glycine, the neurotransmitter conventionally mediating inhibitory signaling when it binds to pentameric glycine receptors. The GluN1-3 NMDARs are vital for regulating neuronal excitability, circuit function, and specific behaviors, yet our understanding of their functional mechanism at the molecular level has remained limited. Here, we present cryo-electron microscopy structures of GluN1-3A NMDARs bound to an antagonist, CNQX, and an agonist, glycine. The structures show a 1-3-1-3 subunit heterotetrameric arrangement and an unprecedented pattern of GluN3A subunit orientation shift between the glycine-bound and CNQX-bound structures. Site-directed disruption of the unique subunit interface in the glycine-bound structure mitigated desensitization. Our study provides a foundation for understanding the distinct structural dynamics of GluN3 that are linked to the unique function of GluN1-3 NMDARs.


Assuntos
Receptores de Glicina , Receptores de N-Metil-D-Aspartato , Animais , Receptores de N-Metil-D-Aspartato/metabolismo , 6-Ciano-7-nitroquinoxalina-2,3-diona , Microscopia Crioeletrônica , Glicina/metabolismo , Neurotransmissores , Mamíferos/metabolismo
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