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The synthesis of 12α-hydroxylated bile acids (12HBAs) and non-12α-hydroxylated bile acids (non-12HBAs) occurs via classical and alternative pathways, respectively. The composition of these BAs is a crucial index for pathophysiologic assessment. However, accurately differentiating 12HBAs and non-12HBAs is highly challenging due to the limited standard substances. Here, we innovatively introduce 12α-hydroxysteroid dehydrogenase (12α-HSDH) as an enzymatic probe synthesized by heterologous expression in Escherichia coli, which can specifically and efficiently convert 12HBAs in vitro under mild conditions. Coupled to the conversion rate determined by liquid chromatography-high resolution mass spectrometry (LC-HRMS), this enzymatic probe allows for the straightforward distinguishing of 210 12HBAs and 312 non-12HBAs from complex biological matrices, resulting in a BAs profile with a well-defined hydroxyl feature at the C12 site. Notably, this enzyme-driven LC-HRMS approach can be extended to any molecule with explicit knowledge of enzymatic transformation. We demonstrate the practicality of this BAs profile in terms of both revealing cross-species BAs heterogeneity and monitoring the alterations of 12HBAs and non-12HBAs under asthma disease. We envisage that this work will provide a novel pattern to recognize the shift of BA metabolism from classical to alternative synthesis pathways in different pathophysiological states, thereby offering valuable insights into the management of related diseases.
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Ácidos e Sais Biliares , Espectrometria de Massas , Ácidos e Sais Biliares/metabolismo , Ácidos e Sais Biliares/química , Ácidos e Sais Biliares/análise , Cromatografia Líquida , Animais , Escherichia coli/enzimologia , Escherichia coli/metabolismo , Humanos , CamundongosRESUMO
Glutathione (GSH) redox control and arginine metabolism are critical in regulating the physiological response to injury and oxidative stress. Quantification assessment of the GSH/arginine redox metabolism supports monitoring metabolic pathway shifts during pathological processes and their linkages to redox regulation. However, assessing the redox status of organisms with complex matrices is challenging, and single redox molecule analysis may not be accurate for interrogating the redox status in cells and in vivo. Herein, guided by a paired derivatization strategy, we present a new ultraperformance liquid chromatography tandem mass spectrometry (UPLC-MS/MS)-based approach for the functional assessment of biological redox status. Two structurally analogous probes, 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) and newly synthesized 2-methyl-6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (MeAQC), were set for paired derivatization. The developed approach was successfully applied to LPS-stimulated RAW 264.7 cells and HDM-induced asthma mice to obtain quantitative information on GSH/arginine redox metabolism. The results suggest that the redox status was remarkably altered upon LPS and HDM stimulation. We expect that this approach will be of good use in a clinical biomarker assay and potential drug screening associated with redox metabolism, oxidative damage, and redox signaling.
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Arginina , Glutationa , Oxirredução , Espectrometria de Massas em Tandem , Animais , Arginina/metabolismo , Arginina/análise , Arginina/química , Glutationa/metabolismo , Glutationa/análise , Camundongos , Espectrometria de Massas em Tandem/métodos , Células RAW 264.7 , Carbamatos/metabolismo , Carbamatos/química , Cromatografia Líquida de Alta Pressão , Lipopolissacarídeos/farmacologia , Aminoquinolinas/químicaRESUMO
BACKGROUND: The causal relationship between daytime napping and the risk of Parkinson's disease (PD) remains unclear, with prospective studies providing limited evidence. This study investigated the association between daytime napping frequency and duration and PD incidence and explored the causality relationship between this association by conducting Mendelian randomization (MR) analysis. METHODS: This prospective cohort study included 393,302 participants, and accelerometer-measured daytime napping data were available only for 78,141 individuals. Cox proportional hazards regression was used to estimate the association between the daytime napping frequency and duration and the PD risk. The role of the systemic immune-inflammation index (SII) in the association between daytime napping frequency and PD risk was assessed through mediation analyses. Moreover, the causal association between the daytime napping frequency and the PD risk was preliminarily explored by conducting two-sample MR analyses. RESULTS: The median follow-up duration was 12.18 years. The participants who reported napping sometimes or usually exhibited a significantly higher PD risk than those who never/rarely napped during the day [sometimes: hazard ratio (HR), 1.13; 95% confidence interval (CI), 1.03-1.23; usually: HR, 1.33; 95% CI, 1.14-1.55], and SII played a mediating role in this association. However, the MR analyses did not indicate that the daytime napping frequency and PD risk were significantly associated. The participants napping for over 1 h exhibited a significantly elevated PD risk (HR, 1.54; 95% CI, 1.11-2.16). Moreover, no significant interaction was identified between napping frequency or duration and genetic susceptibility to PD (P for interaction > 0.05). CONCLUSIONS: In this study, increased daytime napping frequency and duration were associated with an increased PD risk, but no causal relationship was observed between napping frequency and PD risk in the MR analysis. Larger GWAS-based cohort studies and MR studies are warranted to explore potential causal relationships.
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Análise da Randomização Mendeliana , Doença de Parkinson , Sono , Humanos , Doença de Parkinson/genética , Doença de Parkinson/epidemiologia , Estudos Prospectivos , Masculino , Feminino , Pessoa de Meia-Idade , Incidência , Sono/fisiologia , Idoso , Fatores de Risco , Modelos de Riscos Proporcionais , AdultoRESUMO
Lipopolysaccharide (LPS)-activated pro-inflammatory responses play a critical role in sepsis, a life-threatening condition. This study investigates the role of origin recognition complex subunit 6 (ORC6) in LPS responses in macrophages and monocytes. Silencing ORC6 using targeted shRNA significantly reduced LPS-induced expression and production of IL-1ß (interleukin-1 beta), TNF-α (tumor necrosis factor alpha), and IL-6 (interleukin-6) in THP-1 human macrophages, peripheral blood mononuclear cells (PBMCs), and bone marrow-derived macrophages (BMDMs). Additionally, ORC6 knockout (KO) via the CRISPR/Cas9 method in THP-1 macrophages inhibited LPS-induced pro-inflammatory responses, while ectopic overexpression of ORC6 enhanced LPS-induced expression and production of pro-inflammatory cytokines. ORC6 is crucial for the activation of the nuclear factor kappa B (NFκB) signaling cascade in macrophages and monocytes. LPS-induced NFκB activation was largely inhibited by ORC6 silencing or KO, but potentiated following ORC6 overexpression. Mechanistically, ORC6 associated with nuclear p65 after LPS stimulation, an interaction necessary for NFκB activation. Overexpression of ORC6 did not recover the reduced pro-inflammatory response to LPS in THP-1 macrophages with silenced p65. Furthermore, the NFκB inhibitor BMS-345,541 nearly eliminated the pro-inflammatory response enhanced by ORC6 overexpression in response to LPS. Further studies revealed that ORC6 depletion inhibited NFκB activation induced by double-stranded RNA (dsRNA) and high mobility group box 1 (HMGB1) in THP-1 macrophages. In vivo experiments demonstrated that macrophage-specific knockdown of ORC6 protected mice from LPS-induced septic shock and inhibited LPS-stimulated production of IL-1ß, TNF-α, and IL-6 in mouse serum. ORC6 silencing also inhibited LPS-induced NFκB activation in ex vivo cultured PBMCs following macrophage-specific knockdown of ORC6. These findings highlight ORC6 as a pivotal mediator in LPS-induced NFκB activation and the pro-inflammatory response in sepsis, suggesting that targeting ORC6 could be a novel therapeutic strategy for managing sepsis and related inflammatory conditions.
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Inflamação , Lipopolissacarídeos , Macrófagos , NF-kappa B , Complexo de Reconhecimento de Origem , Lipopolissacarídeos/farmacologia , Humanos , Animais , NF-kappa B/metabolismo , Inflamação/metabolismo , Inflamação/genética , Macrófagos/metabolismo , Camundongos , Complexo de Reconhecimento de Origem/metabolismo , Complexo de Reconhecimento de Origem/genética , Células THP-1 , Camundongos Endogâmicos C57BL , Transdução de Sinais , Masculino , Monócitos/metabolismoRESUMO
AIM: To explore the levels of neutrophil extracellular traps (NETs) in patients with periodontitis and examine their effects on keratinization, barrier function of human gingival keratinocytes (HGKs) and the associated mechanisms. MATERIALS AND METHODS: Saliva, gingival crevicular fluid (GCF), clinical periodontal parameters and gingival specimens were collected from 10 healthy control subjects and 10 patients with stage II-IV periodontitis to measure the NET levels. Subsequently, mRNA and protein levels of keratinization and barrier indicators, as well as intracellular calcium and epithelial barrier permeability, were analysed in HGKs after NET stimulation. RESULTS: The study showed that NET levels significantly elevated in patients with periodontitis, across multiple specimens including saliva, GCF and gingival tissues. Stimulation of HGKs with NETs resulted in a decrease in the expressions of involucrin, cytokeratin 10, zonula occludens 1 and E-cadherin, along with decreased intracellular calcium levels and increased epithelial barrier permeability. Furthermore, the inhibition of keratinization by NETs is ERK-KLF4-dependent. CONCLUSIONS: This study indicates that NETs impair the barrier function of HGKs and suppress keratinization through ERK/KLF4 axis. These findings provide potential targets for therapeutic approaches in periodontitis to address impaired gingival keratinization.
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Armadilhas Extracelulares , Gengiva , Líquido do Sulco Gengival , Queratinócitos , Periodontite , Humanos , Armadilhas Extracelulares/metabolismo , Gengiva/metabolismo , Líquido do Sulco Gengival/química , Queratinócitos/metabolismo , Periodontite/metabolismo , Periodontite/imunologia , Feminino , Masculino , Adulto , Pessoa de Meia-Idade , Fator 4 Semelhante a Kruppel , Saliva/metabolismo , Saliva/química , Cálcio/metabolismo , Cálcio/análise , Estudos de Casos e Controles , Epitélio , Queratinas/metabolismo , Caderinas/metabolismo , Caderinas/análiseRESUMO
Liquid-liquid phase separation (LLPS) partitions cellular contents, underlies the formation of membraneless organelles and plays essential biological roles. To date, most of the research on LLPS has focused on proteins, especially RNA-binding proteins. However, accumulating evidence has demonstrated that RNAs can also function as 'scaffolds' and play essential roles in seeding or nucleating the formation of granules. To better utilize the knowledge dispersed in published literature, we here introduce RNAPhaSep (http://www.rnaphasep.cn), a manually curated database of RNAs undergoing LLPS. It contains 1113 entries with experimentally validated RNA self-assembly or RNA and protein co-involved phase separation events. RNAPhaSep contains various types of information, including RNA information, protein information, phase separation experiment information and integrated annotation from multiple databases. RNAPhaSep provides a valuable resource for exploring the relationship between RNA properties and phase behaviour, and may further enhance our comprehensive understanding of LLPS in cellular functions and human diseases.
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Bases de Dados de Ácidos Nucleicos , Organelas/química , Transição de Fase , Proteínas de Ligação a RNA/química , RNA/química , Software , Animais , Células Eucarióticas/citologia , Células Eucarióticas/metabolismo , Humanos , Internet , Anotação de Sequência Molecular , Organelas/metabolismo , Plantas/química , Plantas/genética , Plantas/metabolismo , RNA/classificação , RNA/genética , RNA/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismoRESUMO
Recruitment and accumulation of reactive astrocytes around senile plaques are common pathological features of Alzheimer's disease (AD), with unclear mechanisms. Chemerin, an adipokine implicated in neuroinflammation, acts through its receptor, chemokine-like receptor 1 (CMKLR1), which also functions as a receptor for amyloid ß (Aß). The impact of the chemerin/CMKLR1 axis on astrocyte migration towards Aß plaques is unknown. Here we investigated the effect of CMKLR1 on astrocyte migration around Aß deposition in APP/PS1 mice with Cmklr1 knockout (APP/PS1-Cmklr1-/-). CMKLR1-expressed astrocytes were upregulated in the cortices and hippocampi of 9-month-old APP/PS1 mice. Chemerin mainly co-localized with neurons, and its expression was reduced in the brains of APP/PS1 mice, compared to WT mice. CMKLR1 deficiency decreased astrocyte colocalization with Aß plaques in APP/PS1-Cmklr1-/- mice, compared to APP/PS1 mice. Activation of the chemerin/CMKLR1 axis promoted the migration of primary cultured astrocytes and U251 cells, and reduced astrocyte clustering induced by Aß42. Mechanistic studies revealed that chemerin/CMKLR1 activation induced STING phosphorylation. Deletion of STING attenuated the promotion of the chemerin/CMKLR1 axis relative to astrocyte migration and abolished the inhibitory effect of chemerin on Aß42-induced astrocyte clustering. These findings suggest the involvement of the chemerin/CMKLR1/STING pathway in the regulation of astrocyte migration and recruitment to Aß plaques/Aß42.
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Doença de Alzheimer , Astrócitos , Quimiocinas , Peptídeos e Proteínas de Sinalização Intercelular , Placa Amiloide , Receptores de Quimiocinas , Animais , Astrócitos/metabolismo , Quimiocinas/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/genética , Camundongos , Receptores de Quimiocinas/metabolismo , Receptores de Quimiocinas/genética , Placa Amiloide/metabolismo , Placa Amiloide/patologia , Doença de Alzheimer/metabolismo , Doença de Alzheimer/genética , Doença de Alzheimer/patologia , Humanos , Peptídeos beta-Amiloides/metabolismo , Camundongos Knockout , Movimento Celular , Transdução de Sinais , Camundongos Transgênicos , Camundongos Endogâmicos C57BLRESUMO
Epithelial keratinocyte proliferation is an essential element of wound repair, and chronic wound conditions, such as diabetic foot, are characterized by aberrant re-epithelialization. In this study, we examined the functional role of retinoic acid inducible-gene I (RIG-I), a key regulator of epidermal keratinocyte proliferation, in promoting TIMP-1 expression. We found that RIG-I is overexpressed in keratinocytes of skin injury and underexpressed in skin wound sites of diabetic foot and streptozotocin-induced diabetic mice. Moreover, mice lacking RIG-I developed an aggravated phenotype when subjected to skin injury. Mechanistically, RIG-I promoted keratinocyte proliferation and wound repair by inducing TIMP-1 via the NF-κB signaling pathway. Indeed, recombinant TIMP-1 directly accelerated HaCaT cell proliferation in vitro and promoted wound healing in Ddx58-/- and diabetic mice in vivo. In summary, we demonstrated that RIG-I is a crucial factor that mediates epidermal keratinocyte proliferation and may be a potential biomarker for skin injury severity, thus making it an attractive locally therapeutic target for the treatment of chronic wounds such as diabetic foot.
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Diabetes Mellitus Experimental , Pé Diabético , Animais , Camundongos , Movimento Celular , Proliferação de Células , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/metabolismo , Pé Diabético/genética , Pé Diabético/metabolismo , Queratinócitos/metabolismo , NF-kappa B/metabolismo , Transdução de Sinais , Pele/metabolismo , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Cicatrização/genéticaRESUMO
The specification of endoderm cells to prospective hepatoblasts is the starting point for hepatogenesis. However, how a prospective hepatoblast gains the hepatic fate remains elusive. Previous studies have shown that loss-of-function of either hhex or prox1a alone causes a small liver phenotype but without abolishing the hepatocyte differentiation, suggesting that absence of either Hhex or Prox1a alone is not sufficient to block the hepatoblast differentiation. Here, via genetic studies of the zebrafish two single (hhex-/- and prox1a-/-) and one double (hhex-/-prox1a-/-) mutants, we show that simultaneous loss-of-function of the hhex and prox1a two genes does not block the endoderm cells to gain the hepatoblast potency but abolishes the hepatic differentiation from the prospective hepatoblast. Consequently, the hhex-/-prox1a-/- double mutant displays a liverless phenotype that cannot be rescued by the injection of bmp2a mRNA. Taken together, we provide strong evidences showing that Hhex teams with Prox1a to act as a master control of the differentiation of the prospective hepatoblasts towards hepatocytes.
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Fígado , Peixe-Zebra , Animais , Diferenciação Celular/genética , Hepatócitos , Estudos Prospectivos , Proteínas Repressoras , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/genéticaRESUMO
OBJECTS: Joint morphology is a risk factor for hip osteoarthritis (HOA) and could explain ethnic differences in HOA prevalence. Therefore, we aimed to compare the prevalence of radiographic HOA (rHOA) and hip morphology between the predominantly White UK Biobank (UKB) and exclusively Chinese Shanghai Changfeng (SC) cohorts. METHODS: Left hip iDXA scans were used to quantify rHOA, from a combination of osteophytes (grade ≥1) and joint space narrowing (grade ≥1), and hip morphology. Using an 85-point Statistical Shape Model (SSM) we evaluated cam (alpha angle ≥60°) and pincer (lateral centre-edge angle (LCEA) ≥45°) morphology and acetabular dysplasia (LCEA <25°). Diameter of femoral head (DFH), femoral neck width (FNW), and hip axis length (HAL) were also obtained from these points. Results were adjusted for differences in age, height, and weight and stratified by sex. RESULTS: Complete data were available for 5924 SC and 39,020 White UKB participants with mean ages of 63.4 and 63.7 years old. rHOA prevalence was considerably lower in female (2.2% versus 13.1%) and male (12.0% and 25.1%) SC compared to UKB participants. Cam morphology, rarely seen in females, was less common in SC compared with UKB males (6.3% versus 16.5%). Composite SSM modes, scaled to the same overall size, revealed SC participants to have a wider femoral head compared to UKB participants. FNW and HAL were smaller in SC compared to UKB, whereas DFH/FNW ratio was higher in SC. CONCLUSIONS: rHOA prevalence is lower in Chinese compared with White individuals. Several differences in hip shape were observed, including frequency of cam morphology, FNW, and DFH/FNW ratio. These characteristics have previously been identified as risk factors for HOA and may contribute to observed ethnic differences in HOA prevalence.
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This study aims to examine the effect of superfine powder and aqueous extract of Polygonati Rhizomaon on natural perimenopausal syndrome in rats and explore the underlying mechanism. To be specific, a total of 60 female SD rats(14-15 months old) with estrous cycle disorder were screened by the vaginal smear and randomized into model control group, ß-estradiol 3-benzoate group(0.1 mg·kg~(-1)), superfine powder of Polygonati Rhizoma group(0.25, 0.5 g·kg~(-1)) and aqueous extract of Polygonati Rhizoma group(0.25, 0.5 g·kg~(-1)), and another 10 female SD rats(14-15 months old) were selected as the youth control group. The administration lasted 6 weeks. Then the perimenopausal syndrome-related indexes such as body temperature, microcirculatory blood flow of face and ear, vertigo period, salivary secretion, grip force, and bone strength were determined and open field test was conducted. The immune system-related indexes such as the wet weight and index of thymus and spleen, percentage of T lymphocytes and subgroups in peripheral blood, and hematological indexes were measured. In addition, the ovary-related indexes such as estrous cycle, the wet weight and index of uterus and ovary, ovarian tissue morphology, and cell apoptosis were determined. Moreover, hypothalamus-pituitary-ovary axis(HPO)-related indexes such as serum sex hormone levels, cytochrome P450 family 11 subfamily A member 1(CYP11A1), cytochrome P450 family 19 subfamily A member 1(CYP19A1), and cytochrome P450 family 17 subfamily A member 1(P450 17A1) in ovarian tissue were measured. The results showed that the superfine powder and aqueous extract of Polygonati Rhizoma significantly decreased body temperature(anal, facial and dorsal temperature), microcirculatory blood flow in the ear, and vertigo period, increased salivary secretion, grip force, bone strength, total distance and total speed in the open field test, wet weight and index of thymus and spleen, lymphocyte ratio, CD3~+ level, and CD4~+/CD8~+ ratio, reduced neutrophil number and ratio, estrous cycle disorder ratio, and number of ovarian apoptotic cells, raised wet weight and index of uterus, wet weight of ovary, levels of inhibin B(INHB), estradiol(E_2), anti-müllerian hormone(AMH), and ovarian CYP11A1 and CYP19A1, decreased follicle-stimulating hormone(FSH) and luteinizing hormone(LH) content, and improved ovarian tissue morphology. It is suggested that the superfine powder and aqueous extract of Polygonati Rhizoma can improve the symptoms associated with natural perimenopausal syndrome in rats and enhance ovarian function and immune function. The mechanism is that they regulate HPO axis function by increasing estrogen synthesis.
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Enzima de Clivagem da Cadeia Lateral do Colesterol , Perimenopausa , Feminino , Animais , Ratos , Ratos Sprague-Dawley , Microcirculação , Pós , Citocromo P-450 CYP1A1RESUMO
Acetaldehyde (ACD), the first metabolite of ethanol, is implicated in several of ethanol's actions, including the reinforcing and aversive effects. The neuronal mechanisms underlying ACD's aversive effect, however, are poorly understood. The lateral habenula (LHb), a regulator of midbrain monoaminergic centers, is activated by negative valence events. Although the LHb has been linked to the aversive responses of several abused drugs, including ethanol, little is known about ACD. We, therefore, assessed ACD's action on LHb neurons in rats. The results showed that intraperitoneal injection of ACD increased cFos protein expression within the LHb and that intra-LHb infusion of ACD induced conditioned place aversion in male rats. Furthermore, electrophysiological recording in brain slices of male and female rats showed that bath application of ACD facilitated spontaneous firing and glutamatergic transmission. This effect of ACD was potentiated by an aldehyde dehydrogenase (ALDH) inhibitor, disulfiram (DS), but attenuated by the antagonists of dopamine (DA) receptor (DAR) subtype 1 (SCH23390) and subtype 2 (raclopride), and partly abolished by the pretreatment of DA or DA reuptake blocker (GBR12935; GBR). Moreover, application of ACD initiated a depolarizing inward current (IACD) and enhanced the hyperpolarizing-activated currents in LHb neurons. Bath application of Rp-cAMPs, a selective cAMP-PKA inhibitor, attenuated ACD-induced potentiation of EPSCs and IACD Finally, bath application of ZD7288, a selective blocker of hyperpolarization-activated cyclic nucleotide-gated channels, attenuated ACD-induced potentiation of firing, EPSCs, and IACD These results show that ACD exerts its aversive property by exciting LHb neurons via multiple cellular mechanisms, and new treatments targeting the LHb may be beneficial for alcoholism.SIGNIFICANCE STATEMENT Acetaldehyde (ACD) has been considered aversive peripherally and rewarding centrally. However, whether ACD has a central aversive property is unclear. Here, we report that ACD excites the lateral habenula (LHb), a brain region associated with aversion and negative valence, through multiple cellular and molecular mechanisms. Intra-LHb ACD produces significant conditioned place aversion. These results suggest that ACD's actions on the LHb neurons might contribute to its central aversive property and new treatments targeting the LHb may be beneficial for alcoholism.
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Acetaldeído/farmacologia , Aprendizagem da Esquiva/efeitos dos fármacos , Habenula/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Animais , Dissulfiram/farmacologia , Antagonistas de Dopamina/farmacologia , Inibidores da Captação de Dopamina/farmacologia , Ácido Glutâmico/metabolismo , Habenula/fisiologia , Masculino , Neurônios/fisiologia , Proteínas Proto-Oncogênicas c-fos/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores Dopaminérgicos/metabolismo , Transmissão Sináptica/efeitos dos fármacosRESUMO
Collision cross section (CCS) values generated from ion mobility mass spectrometry (IM-MS) have commonly been employed to facilitate lipid identification. However, this is hindered by the limited available lipid standards. Recently, CCS values were predicted by means of computational calculations, though the prediction precision was generally not good and the predicted CCS values of the lipid isomers were almost identical. To address this challenge, a least absolute shrinkage and selection operator (LASSO)-based prediction method was developed for the prediction of lipids' CCS values in this study. In this method, an array of molecular descriptors were screened and optimized to reflect the subtle differences in structures among the different lipid isomers. The use of molecular descriptors together with a wealth of standard CCS values for the lipids (365 in total) significantly improved the accuracy and precision of the LASSO model. Its accuracy was externally validated with median relative errors (MREs) of <1.1% using an independent data set. This approach was demonstrated to allow differentiation of cis/trans and sn-positional isomers. The results also indicated that the LASSO-based prediction method could practically reduce false-positive identifications in IM-MS-based lipidomics.
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Espectrometria de Mobilidade Iônica , Lipidômica , Espectrometria de Mobilidade Iônica/métodos , Isomerismo , Lipídeos/análiseRESUMO
INTRODUCTION: Traditional methods to derive experimentally-generated relative correction factors (RCFs) for the quantitative analysis of herbal multi-components by single marker (QAMS) method require reference standards and multiple validations with different instruments and columns, which hampers high throughput implementation. OBJECTIVES: To effectively reduce the application amounts of raw material and provide higher and more stable accuracy, this study aimed to develop a method to computationally generate RCFs of herbal components. MATERIALS AND METHODS: This strategy included the published data collection, calibration curves screening, computer algorithm-based RCFs generation and accuracy validation. RESULTS: Using the in silico approach, we have successfully produced 133 RCFs for the multi-component quantitative analysis of 63 widely used herbs. CONCLUSION: Compared with conventional RCFs, this in silico method would be a low cost and highly efficient way to produce practical RCFs for the QAMS method.
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Medicamentos de Ervas Chinesas , Cromatografia Líquida de Alta Pressão , Simulação por ComputadorRESUMO
Hexavalent chromium contamination is a global environmental issue and usually reoccurs in alkaline reduced chromite ore processing residues (rCOPR). The oxidation of Cr(III) solids in rCOPR is one possible cause but as yet little studied. Herein, we investigated the oxidation of Cr(OH)3, a typical species of Cr(III) in rCOPR, at alkaline pH (9-11) with δ-MnO2 under oxic/anoxic conditions. Results revealed three pathways for Cr(III) oxidation under oxic conditions: (1) oxidation by oxygen, (2) oxidation by δ-MnO2, and (3) catalytic oxidation by Mn(II). Oxidations in the latter two were efficient, and oxidation via Pathway 3 was continuous and increased dramatically with increasing pH. XANES data indicated feitknechtite (ß-MnOOH) and hausmannite (Mn3O4) were the reduction products and catalytic substances. Additionally, a kinetic model was established to describe the relative contributions of each pathway at a specific time. The simulation outcomes showed that Cr(VI) was mainly formed via Pathway 2 (>51%) over a short time frame (10 days), whereas in a longer-term (365 days), Pathway 3 predominated the oxidation (>78%) with an increasing proportion over time. These results suggest Cr(III) solids can be oxidized under alkaline oxic conditions even with a small amount of manganese oxides, providing new perspectives on Cr(VI) reoccurrence in rCOPR and emphasizing the environmental risks of Cr(III) solids in alkaline environments.
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Compostos de Manganês , Óxidos , Cromo , OxirreduçãoRESUMO
Late blight is a devastating potato disease worldwide, caused by Phytophthora infestans. The P. infestans strain 2013-18-306 from Yunnan is a "supervirulent race" that overcomes all 11 known late blight resistance genes (R1 to R11) from Solanum demissum. In a previous study, we identified a diploid wild-type potato JAM1-4 (S. jamesii) with high resistance to 2013-18-306. dRenSeq analysis indicated the presence of novel R genes in JAM1-4. RNA-Seq was used to analyze the late blight resistance response genes and defense regulatory mechanisms of JAM1-4 against 2013-18-306. Gene ontology enrichment and KEGG pathway analysis showed that many disease-resistant pathways were significantly enriched. Analysis of differentially expressed genes (DEGs) revealed an active disease resistance mechanism of JAM1-4, and the essential role of multiple signal transduction pathways and secondary metabolic pathways comprised of SA-JA-ET in plant immunity. We also found that photosynthesis in JAM1-4 was inhibited to promote the immune response. Our study reveals the pattern of resistance-related gene expression in response to a super race strain of potato late blight and provides a theoretical basis for further exploration of potato disease resistance mechanisms, discovery of new late blight resistance genes, and disease resistance breeding.
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Phytophthora infestans , Solanum tuberosum , China , Diploide , Doenças das PlantasRESUMO
BACKGROUND: CXCL12(chemokine ligand 12, CXCL12) and its receptors CXCR4 are widely expressed in maternal-fetal interface and plays an adjust role in materno-fetal dialogue and immune tolerance during early pregnancy. This study aimed to evaluate the role and mechanism of self-derived CXCL12 in modulating the functions of human first-trimester endometrial epithelial cells (EECs) and to identify the potential protein kinase signaling pathways involved in the CXCL12/CXCR4's effect on EECs. METHODS: The expression of CXCL12 and CXCR4 in EECs was measured by using immunohistochemistry, immunofluorescence, real-time polymerase chain reaction and enzyme-linked immunosorbent assay. The effects of EEC-conditioned medium (EEC-CM) and recombinant human CXCL12 (rhCXCL12) on EEC migration and invasion in vitro were evaluated with migration and invasion assays. In-cell western blot analysis was used to examine the phosphorylation of protein kinase B (AKT), extracellular regulated protein kinases (ERKs) and phosphatidylinositol 3-kinase (PI3K) after CXCL12 treatment. RESULTS: CXCL12 and CXCR4 were both expressed in human first-trimester EECs at the mRNA and protein level. Both EEC-CM and rhCXCL12 significantly increased the migration and invasion of EECs (P < 0.05), which could be blocked by neutralizing antibodies against CXCR4 (P < 0.05) or CXCL12 (P < 0.05), respectively. CXCL12 activated both PI3K/AKT and ERK1/2 signaling and CXCR4 neutralizing antibody effectively reduced CXCL12-induced phosphorylation of AKT and ERK1/2. LY294002, a PI3K-AKT inhibitor, was able to reverse the promotive effect of EEC-CM or rhCXCL12 on EEC migration and invasion. CONCLUSIONS: Human first-trimester EECs promoted their own migration and invasion through the autocrine mechanism with CXCL12/CXCR4 axis involvement by activating PI3K/AKT signaling. This study contributes to a better understanding of the epithelium function mediated by chemokine and chemokine receptor during normal pregnancy.
Assuntos
Comunicação Autócrina/genética , Movimento Celular/genética , Quimiocina CXCL12/fisiologia , Endométrio/citologia , Receptores CXCR4/fisiologia , Técnicas de Cultura de Células , Células Epiteliais/fisiologia , Feminino , Humanos , Fosfatidilinositol 3-Quinase/fisiologia , Gravidez , Primeiro Trimestre da Gravidez/fisiologia , Proteínas Proto-Oncogênicas c-akt/fisiologia , Transdução de Sinais/genéticaRESUMO
AbaR-type genomic islands (AbaRs) are important elements responsible for antimicrobial resistance in Acinetobacter baumannii This study performed a large-scale identification of AbaRs to understand their distribution and compositions of antimicrobial resistance genes. We identified 2.89-kb left-end and 1.87-kb right-end conserved sequences (CSs) and developed a bioinformatics approach to identify AbaRs, using the CSs as signatures, in 3,148 publicly available genomes. AbaRs were prevalent in A. baumannii, being found in 2,091 genomes. They were sparse in other Acinetobacter species and confined only to this genus. Results from 111 complete genomes showed that over 85% of AbaRs resided on chromosomes. The external flanks adjacent to the inverted repeats available in all identified CSs were mapped to an AbaR-free chromosome or searched in the NCBI database for empty loci to define insertion sites. Surprisingly, 84 insertion sites with diverse origins were revealed, including 51 scattered on the chromosome, 20 plasmid borne, 12 located on prophages, transposons, ISAba1, complex AbaRs, and genomic islands of other types, and one uncharacterized, and some were strongly associated with clonal lineages. Finally, we found 994 antimicrobial resistance genes covering 28 unique genes from 70.9% (299/422) of intact AbaRs currently available. The resistance gene profiles displayed an apparent clonal lineage-specific pattern, highlighting the distinct features of AbaRs in global clone 1 (GC1) and GC2. The tet(B) gene was highly specific to the AbaRs in GC2. In conclusion, AbaRs have diverse insertion sites on the chromosome and mobile genetic elements (MGEs) and display distinct antimicrobial resistance gene profiles in different clonal lineages.
Assuntos
Acinetobacter baumannii/genética , Proteínas de Bactérias/genética , DNA Bacteriano/genética , Farmacorresistência Bacteriana Múltipla/genética , Ilhas Genômicas/genética , Infecções por Acinetobacter/tratamento farmacológico , Infecções por Acinetobacter/microbiologia , Acinetobacter baumannii/efeitos dos fármacos , Antibacterianos/farmacologia , Elementos de DNA Transponíveis/genética , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Testes de Sensibilidade Microbiana/métodos , Plasmídeos/genética , Análise de Sequência de DNA/métodosRESUMO
Gut microbiota and their metabolites (short-chain fatty acids, SCFAs) are associated with the pathogenesis of rheumatoid arthritis (RA). Total Clematis triterpenoid saponins (CTSs) prepared from Clematis mandshurica Rupr. possess therapeutic benefits for arthritic diseases. However, the poor pharmacokinetic properties of CTSs have obstructed the translation of these natural agents to drugs. Here, we examined the effects of CTSs on arthritis symptoms, gut microbiota and SCFAs in rats with collagen-induced arthritis (CIA). Our results showed that the arthritis index scores of CIA rats treated with CTSs were significantly lower than those of the model group. Most importantly, CTSs moderated gut microbial dysbiosis and significantly downregulated the total SCFA concentration in CIA rats. Compared to the control group, CTSs treatment have no significant side effects on the gut microbiota and SCFA metabolism in normal rats. Two differential analyses (LEfSe and DESeq2) were combined to study the details of the changes in gut microbiome, and twenty-four marker taxa at the genus level were identified via a comparison among control, model and CIA rats treated with high doses of CTSs. In particular, the mostly significantly increased gram-negative (G-) and decreased gram-positive (G+) genera in CIA rats were well restored by CTSs. The observed SCFA concentrations demonstrated that CTSs tend to maintain the balance of the gut microbiota. The data presented herein suggest that CTSs could ameliorate arthritis-associated gut microbial dysbiosis and may be potential adjuvant drugs that could provide relief from the gastrointestinal damage caused as a side effect of commonly used drugs.
Assuntos
Artrite Experimental/tratamento farmacológico , Clematis/química , Disbiose/prevenção & controle , Ácidos Graxos Voláteis/metabolismo , Microbioma Gastrointestinal/efeitos dos fármacos , Saponinas/uso terapêutico , Triterpenos/uso terapêutico , Animais , Artrite Experimental/microbiologia , Disbiose/microbiologia , Feminino , Ratos , Ratos Wistar , Saponinas/isolamento & purificação , Triterpenos/isolamento & purificaçãoRESUMO
BACKGROUND: Despite the fact that miRNAs play pivotal roles in various human malignancies, their molecular mechanisms influencing RCC are poorly understood. METHODS: The expression of miRNAs from RCC and paired normal renal specimens was analysed by a combined computational and experimental approach using two published datasets and qRT-PCR assays. The functional role of these miRNAs was further identified by overexpression and inhibition assays in vivo and in vitro. Western blots, luciferase assays, and chromatin immunoprecipitation were performed to investigate the potential mechanisms of these miRNAs. RESULTS: Bioinformatics analysis and qRT-PCR revealed that miR-532-5p was one of the most heavily downregulated miRNAs. Overexpression of miR-532-5p inhibited RCC cell proliferation, while knockdown of miR-532-5p promoted cell proliferation. Mechanistic analyses indicated that miR-532-5p directly targets KRAS and NAP1L1. Interestingly, ETS1 suppressed the transcription of miR-532-5p by directly binding a special region of its promoter. Moreover, high levels of ETS1, as an oncogene in RCC, were significantly associated with poor survival in a large cohort of RCC specimens. CONCLUSIONS: Our work presents a road map for the prediction and validation of a miR-532-5p/KRAS-NAP1L1/P-ERK/ETS1 axis feedback loop regulating cell proliferation, which could potentially provide better therapeutic avenues for treating RCC.