RESUMO
Biomolecular condensates have emerged as an important subcellular organizing principle1. Replication of many viruses, including human respiratory syncytial virus (RSV), occurs in virus-induced compartments called inclusion bodies (IBs) or viroplasm2,3. IBs of negative-strand RNA viruses were recently shown to be biomolecular condensates that form through phase separation4,5. Here we report that the steroidal alkaloid cyclopamine and its chemical analogue A3E inhibit RSV replication by disorganizing and hardening IB condensates. The actions of cyclopamine and A3E were blocked by a point mutation in the RSV transcription factor M2-1. IB disorganization occurred within minutes, which suggests that these molecules directly act on the liquid properties of the IBs. A3E and cyclopamine inhibit RSV in the lungs of infected mice and are condensate-targeting drug-like small molecules that have in vivo activity. Our data show that condensate-hardening drugs may enable the pharmacological modulation of not only many previously undruggable targets in viral replication but also transcription factors at cancer-driving super-enhancers6.
Assuntos
Condensados Biomoleculares/virologia , Vírus Sincicial Respiratório Humano/efeitos dos fármacos , Alcaloides de Veratrum/farmacologia , Replicação Viral/efeitos dos fármacos , Animais , Antivirais/farmacologia , Linhagem Celular , Feminino , Humanos , Corpos de Inclusão , Pulmão/virologia , Camundongos , Camundongos Endogâmicos BALB C , Vírus Sincicial Respiratório Humano/fisiologia , Fatores de Transcrição , Proteínas ViraisRESUMO
The ciliary muscle constitutes a crucial element in refractive regulation. Investigating the pathophysiological mechanisms within the ciliary muscle during excessive contraction holds significance in treating ciliary muscle dysfunction. A guinea pig model of excessive contraction of the ciliary muscle induced by drops pilocarpine was employed, alongside the primary ciliary muscle cells was employed in in vitro experiments. The results of the ophthalmic examination showed that pilocarpine did not significantly change refraction and axial length during the experiment, but had adverse effects on the regulatory power of the ciliary muscle. The current data reveal notable alterations in the expression profiles of hypoxia inducible factor 1 (HIF-1α), ATP2A2, P53, α-SMA, Caspase-3, and BAX within the ciliary muscle of animals subjected to pilocarpine exposure, alongside corresponding changes observed in cultured cells treated with pilocarpine. Augmented levels of ROS were detected in both tissue specimens and cells, culminating in a significant increase in cell apoptosis in in vivo and in vitro experiments. Further examination revealed that pilocarpine induced an increase in intracellular Ca2+ levels and disrupted MMP, as evidenced by mitochondrial swelling and diminished cristae density compared to control conditions, concomitant with a noteworthy decline in antioxidant enzyme activity. However, subsequent blockade of Ca2+ channels in cells resulted in downregulation of HIF-1α, ATP2A2, P53, α-SMA, Caspase-3, and BAX expression, alongside ameliorated mitochondrial function and morphology. The inhibition of Ca2+ channels presents a viable approach to mitigate ciliary cells damage and sustain proper ciliary muscle function by curtailing the mitochondrial damage induced by excessive contractions.
Assuntos
Apoptose , Cálcio , Senescência Celular , Pilocarpina , Animais , Pilocarpina/farmacologia , Cobaias , Apoptose/efeitos dos fármacos , Cálcio/metabolismo , Senescência Celular/efeitos dos fármacos , Corpo Ciliar/metabolismo , Masculino , Células Cultivadas , Espécies Reativas de Oxigênio/metabolismoRESUMO
AIM: During hypertension-induced endothelial dysfunction, periodic mechanical stretching (MS) activates related inflammatory pathways and leads to endothelial damage, but the underlying mechanisms remain unknown. The present study aimed to determine the injury of HUVECs caused by overstretching and the role of HMGB1-RAGE pathway in HUVECs after injury. MAIN METHODS AND KEY FINDINGS: Human umbilical vein endothelial cells (HUVECs) were cultured and seeded in BioFlex™ plates (six wells). Cells were exposed to 5% (physiological state) and 20% (pathological state) mechanical stretch at 1 Hz for 12 or 24 h in a Flexcell-5000™, with unstretched cells serving as controls. It was found that excessive MS can inhibit cell viability, proliferation, and tube-forming ability resulting in disordered cell arrangement and orientation, slowing cell migration. All these changes cause endothelial damage compared to physiological MS. Endothelial cells (ECs) promote cell migration and self-repair after injury by increasing the High-mobility group box 1 (HMGB1) expression. Experiments and protein prediction networks have shown that HMGB1 can also promote the expression of downstream protein bFGF by binding to receptor for advanced glycation end products (RAGE). Interestingly, VEGF protein expression did not change significantly during this repair process, implying that bFGF replaces the role of VEGF in vascular endothelial repair. SIGNIFICANCE: The present study demonstrates that in the context of endothelial injury caused by excessive MS, the HMGB1/RAGE/bFGF pathway is activated and promotes endothelial repair after injury. Therefore, understanding these mechanisms will help find new therapies for diseases such as hypertension, atherosclerosis, and aneurysm formation.
Assuntos
Proteína HMGB1 , Hipertensão , Humanos , Receptor para Produtos Finais de Glicação Avançada/genética , Receptor para Produtos Finais de Glicação Avançada/metabolismo , Proteína HMGB1/metabolismo , Células Endoteliais da Veia Umbilical Humana/metabolismo , Fator A de Crescimento do Endotélio Vascular , Transdução de Sinais , Fator 2 de Crescimento de FibroblastosRESUMO
The effects of prenatal opioid exposure in adult animals has been widely studied, but little is known about the effects of prenatal opioid on adolescents. Most of the risk behaviors associated with drug abuse are initiated during adolescence. The developmental state of the adolescent brain makes it vulnerable to initiate drug use and susceptible to drug-induced brain changes. In this study, pregnant rats were subcutaneously injected with an increasing dose of morphine (5 mg/kg, 7 mg/kg, 10 mg/kg) for 9 days since the gestation day 11. The effects of prenatal morphine (PNM) on learning and memory, anxiety- and depressive- like behavior, morphine induced conditioned place preference (CPP) as well as locomotor sensitization were tested in both adolescent and adult rats. The results showed that: (1) PNM decreased anxiety-like behavior in both adolescent and adult female rats, but not males; (2) PNM decreased depressive-like behavior in adolescent but increased depressive -like behavior in adult females; (3) PNM increased low dose morphine induced locomotor sensitization in females; (4) PNM decreased tyrosine hydroxylase (TH) expression in the prefrontal cortex but decreased dopamine D1 receptor expression in the nucleus-accumbens (NAc) in female rats. These results suggested that PNM altered the emotional and addictive behavior mainly in female rats, with female rats being less anxiety and depressive during adolescence, but more depressive in adult, and more sensitive to low dose morphine induced locomotor activity sensitization, which might be mediated in part by the differential expression of the TH, dopamine D1 receptors in the female brain.
Assuntos
Comportamento Aditivo , Morfina , Efeitos Tardios da Exposição Pré-Natal , Analgésicos Opioides/efeitos adversos , Animais , Comportamento Animal , Condicionamento Clássico , Feminino , Masculino , Morfina/efeitos adversos , Núcleo Accumbens/metabolismo , Gravidez , Efeitos Tardios da Exposição Pré-Natal/fisiopatologia , Efeitos Tardios da Exposição Pré-Natal/psicologia , Ratos , Fatores Sexuais , Tirosina 3-Mono-Oxigenase/metabolismoRESUMO
Domoic acid (DA) is a potent neurotoxin produced by toxigenic Pseudo-nitzschia blooms and quickly transfers to the benthic anaerobic environment by marine snow particles. DA anaerobic biotransformation is driven by microbial interactions, in which trace amounts of DA can cause physiological stress in marine microorganisms. However, the underlying response mechanisms of microbial community to DA stress remain unclear. In this study, we utilized an anaerobic marine DA-degrading consortium GLY (using glycine as co-substrate) to systematically investigate the global response mechanisms of microbial community during DA anaerobic biotransformation.16S rRNA gene sequencing and metatranscriptomic analyses were applied to measure microbial community structure, function and metabolic responses. Results showed that DA stress markedly changed the composition of main species, with increased levels of Firmicutes and decreased levels of Proteobacteria, Cyanobacteria, Bacteroidetes and Actinobacteria. Several genera of tolerated bacteria (Bacillus and Solibacillus) were increased, while, Stenotrophomonas, Sphingomonas and Acinetobacter were decreased. Metatranscriptomic analyses indicated that DA stimulated the expression of quorum sensing, extracellular polymeric substance (EPS) production, sporulation, membrane transporters, bacterial chemotaxis, flagellar assembly and ribosome protection in community, promoting bacterial adaptation ability under DA stress. Moreover, amino acid metabolism, carbohydrate metabolism and lipid metabolism were modulated during DA anaerobic biotransformation to reduce metabolic burden, increase metabolic demands for EPS production and DA degradation. This study provides the new insights into response of microbial community to DA stress and its potential impact on benthic microorganisms in marine environments.
Assuntos
Diatomáceas , Microbiota , Aminoácidos/metabolismo , Anaerobiose , Bactérias/metabolismo , Biotransformação , Diatomáceas/química , Diatomáceas/genética , Diatomáceas/metabolismo , Matriz Extracelular de Substâncias Poliméricas/química , Matriz Extracelular de Substâncias Poliméricas/metabolismo , Glicina , Ácido Caínico/análogos & derivados , Toxinas Marinhas/análise , Toxinas Marinhas/toxicidade , Proteínas de Membrana Transportadoras/metabolismo , Neurotoxinas , RNA Ribossômico 16SRESUMO
The good treatment of skin defects has always been a challenge in the medical field, and the emergence of tissue engineering skin provides a new idea for the treatment of injured skin. However, due to the single seed cells, the tissue engineering skin has the problem of slow vascularization at the premonitory site after implantation into the human body. Cell co-culture technology can better simulate the survival and communication environment of cells in the human body. The study of multicellular co-culture hopes to bring a solution to the problem of tissue engineering. In this paper, human skin fibroblasts (HSFs) and human vascular endothelial cells (HVECs) were co-cultured in Transwell. The Cell Counting Kit 8 (CCK8), Transwell migration chamber, immunofluorescence, Western blot (WB), and real time quantitative PCR (RT-qPCR) were used to study the effects of HVECs on cell activity, migration factor (high mobility group protein 1, HMGB1) and vascularization factor (vascular endothelial growth factor A, VEGFA and fibroblast growth factor 2, FGF2) secretion of HSFs after co-cultured with HVECs in the Transwell. The biological behavior of HSFs co-cultured with HVECs was studied. The experimental results are as follows: (1) The results of cck8 showed that HVECS could promote the activity of HSFs. (2) HVECs could significantly promote the migration of HSFs and promote the secretion of HMGB1. (3) HVECs could promote the secretion of VEGFA and FGF2 of HSFs. (4) The HVECs and HSFs were inoculated on tissue engineering scaffolds at the ratio of 1:4 and were co-cultured and detected for 7 days. The results showed that from the third day, the number of HSFs was significantly higher than that of the control group without HVECs.
Assuntos
Células Endoteliais , Proteína HMGB1 , Humanos , Técnicas de Cocultura , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo , Proteína HMGB1/metabolismo , Fator 2 de Crescimento de Fibroblastos/metabolismo , Fibroblastos/metabolismoRESUMO
Payment for Ecosystem Services (PES) has been widely accepted as a policy tool for promoting ecological and social progress. However, PES development and implementation in traditional agricultural and pastoral areas are often more challenging than in other areas. The contradiction between ecological protection and people's livelihood development in traditional agricultural and pastoral areas is related to developing country's sustainable development strategy. Based on this, we evaluate the PES (ERCCP: the Ecological Relocation and Capital Compensation Program) program in Tianzhu County, as a case study to investigated the impact of ERCCP on the local natural and social ecosystems on a 20-year scale. The results of indicated that ERCCP has achieved "win-win" gains of restoring environment and promoting socioeconomic development: in the 10 years since ERCCP was implemented, the area of forest land and grassland increased significantly, increasing by 1135.6 ha and 919.62 ha, respectively. 57.5% of farmland was converted to grassland and 30.8% to forest, respectively. In addition, 92.2% and 7.5% of bare land were replaced by grassland and forest, respectively, indicating a gradual recovery of green land during this period. We also analyzed the effects of ERCCP on social systems, and found that the change of agro-pastoralists' attitude towards ERCCP promoted the transfer of labor force from the primary industry to the tertiary industry, accelerated the development of urbanization, and made the poor population completely out of poverty by 2020. In addition, we predict that the income level of households, the PES return on investment of local governments, and the value of regional ecosystem services will increase significantly after 2025. In this context, We establish a theoretical model to explain the win-win plan for the coordinated development of ecosystem services and regional well-being to explore the sustainability of PES and provided a typical case for the similar research area in the world, especially in the areas with the ecological fragility and poverty problems.
Assuntos
Conservação dos Recursos Naturais , Ecossistema , Agricultura , China , Fazendas , Florestas , HumanosRESUMO
Mutations in FASTKD2, a mitochondrial RNA binding protein, have been associated with mitochondrial encephalomyopathy with isolated complex IV deficiency. However, deficiencies related to other oxidative phosphorylation system (OXPHOS) complexes have not been reported. Here, we identified three novel FASTKD2 mutations, c.808_809insTTTCAGTTTTG, homoplasmic mutation c.868C>T, and heteroplasmic mutation c.1859delT/c.868C>T, in patients with mitochondrial encephalomyopathy. Cell-based complementation assay revealed that these three FASTKD2 mutations were pathogenic. Mitochondrial functional analysis revealed that mutations in FASTKD2 impaired the mitochondrial function in patient-derived lymphocytes due to the deficiency in multi-OXPHOS complexes, whereas mitochondrial complex II remained unaffected. Consistent results were also found in human primary muscle cell and zebrafish with knockdown of FASTKD2. Furthermore, we discovered that FASTKD2 mutation is not inherently associated with epileptic seizures, optic atrophy, and loss of visual function. Alternatively, a patient with FASTKD2 mutation can show sinus tachycardia and hypertrophic cardiomyopathy, which was partially confirmed in zebrafish with knockdown of FASTKD2. In conclusion, both in vivo and in vitro studies suggest that loss of function mutation in FASTKD2 is responsible for multi-OXPHOS complexes deficiency, and FASTKD2-associated mitochondrial disease has a high degree of clinical heterogenicity.
Assuntos
Estudos de Associação Genética , Predisposição Genética para Doença , Mitocôndrias/genética , Doenças Mitocondriais/diagnóstico , Doenças Mitocondriais/genética , Mutação , Proteínas Serina-Treonina Quinases/genética , Trifosfato de Adenosina/metabolismo , Alelos , Sequência de Aminoácidos , Animais , Linhagem Celular , Respiração Celular/genética , Criança , Pré-Escolar , Análise Mutacional de DNA , Feminino , Técnicas de Silenciamento de Genes , Estudos de Associação Genética/métodos , Humanos , Linfócitos/imunologia , Linfócitos/metabolismo , Mitocôndrias/metabolismo , Doenças Mitocondriais/metabolismo , Modelos Moleculares , Fosforilação Oxidativa , Linhagem , Fenótipo , Conformação Proteica , Proteínas Serina-Treonina Quinases/química , Sequenciamento do Exoma , Peixe-ZebraRESUMO
TOM70 is a member of the TOM complex that transports cytosolic proteins into mitochondria. Here, we identified two compound heterozygous variants in TOMM70 [c.794C>T (p.T265M) and c.1745C>T (p.A582V)] from a patient with severe anemia, lactic acidosis, and developmental delay. Patient-derived immortalized lymphocytes showed decreased TOM70 expression, oligomerized TOM70 complex, and TOM 20/22/40 complex compared with expression in control lymphocytes. Functional analysis revealed that patient-derived cells exhibited multi-oxidative phosphorylation system (OXPHOS) complex defects, with complex IV being primarily affected. As a result, patient-derived cells grew slower in galactose medium and generated less ATP and more extracellular lactic acid than did control cells. In vitro cell model compensatory experiments confirmed the pathogenicity of TOMM70 variants since only wild-type TOM70, but not mutant TOM70, could restore the complex IV defect and TOM70 expression in TOM70 knockdown U2OS cells. Altogether, we report the first case of mitochondrial disease-causing mutations in TOMM70 and demonstrate that TOM70 is essential for multi-OXPHOS assembly. Mutational screening of TOMM70 should be employed to identify mitochondrial disease-causing gene mutations in the future.
Assuntos
Acidose Láctica/genética , Anemia/genética , Deficiências do Desenvolvimento/genética , Proteínas de Transporte da Membrana Mitocondrial/genética , Acidose Láctica/patologia , Anemia/patologia , Criança , Deficiências do Desenvolvimento/patologia , Humanos , Masculino , Mitocôndrias/genética , Doenças Mitocondriais/genética , Doenças Mitocondriais/patologia , Proteínas do Complexo de Importação de Proteína Precursora Mitocondrial , Mutação/genética , Fosforilação Oxidativa , Sequenciamento do ExomaRESUMO
STUDY QUESTION: What is the expression level of T-cadherin in endometriosis, and does T-cadherin play a role in regulating invasion and migration of endometrial stromal cells? SUMMARY ANSWER: T-cadherin expression was reduced in ectopic endometriotic lesions compared to eutopic endometrium, and T-cadherin overexpression inhibited the invasion and migration of endometrial stromal cells. WHAT IS KNOWN ALREADY: Endometriosis is a disease that involves active cell invasion and migration. T-cadherin can inhibit cell invasion, migration and proliferation in various cancer cells, but its role in endometriosis has not been investigated. STUDY DESIGN, SIZE, DURATION: We explored the expression status of T-cadherin in 40 patients with and 24 without endometriosis. We also isolated endometrial stromal cells to study the invasion, migration and signaling pathway regulation of T-cadherin overexpression. PARTICIPANTS/MATERIALS, SETTING, METHODS: Patients were recruited at the Guangzhou Women and Children's Medical Center to study the expression levels of T-cadherin. The expression of T-cadherin was detected by immunohistochemistry staining and western blot. H-score was used to evaluate the staining intensity of T-cadherin. The correlation between T-cadherin expression levels (H-score) and endometriosis patients' age, stage, lesion size and adhesion was analyzed. Endometrial stromal cells from patients with and without endometriosis were isolated, and cell invasion and migration were detected by transwell assays after T-cadherin overexpression. The expression of vimentin in T-cadherin-overexpressed cells was detected by western blot. After T-cadherin overexpression, the phosphorylation profile of signaling pathway proteins was detected with the Proteome Profiler Human Phospho-Kinase Array Kit. MAIN RESULTS AND THE ROLE OF CHANCE: There was no difference in the expression of T-cadherin in the normal endometrium of control patients and the eutopic endometrium of endometriotic patients, but it was significantly decreased in the ectopic endometrium of endometriotic patients, compared with control endometrium and eutopic endometrium of endometriosis patients (P < 0.0001, for both). Western blot analysis also showed that the expression of T-cadherin was decreased in ectopic endometriotic lesions, but not the normal control endometrium or the endometriotic eutopic endometrium. The results of transwell assays indicated that T-cadherin overexpression inhibited the invasion and migration of endometrial stromal cells. In addition, T-cadherin overexpression promoted the phosphorylation of HSP27 (S78/S82) and JNK 1/2/3 (T183/Y185, T221/Y223) and decreased the expression of vimentin, MMP2 and MMP9 in eutopic endometriosis stromal cells. LARGE-SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: The control group were patients with benign gynecological conditions (e.g. uterus myoma, endometrial or cervical polyp), which may have genetic or epigenetic variations associated with T-cadherin expression and signaling pathways. The case numbers of involved endometriosis and control patients were limited. This study only used endometrial stromal cells from patients with or without endometriosis. Ideally, ectopic endometrial stromal cells of the ovarian endometriotic lesions should also be utilized to explore the function of T-cadherin. WIDER IMPLICATIONS OF THE FINDINGS: Further investigation of the role of T-cadherin in endometriosis may generate new potential therapeutic targets for this complex disorder. STUDY FUNDING AND COMPETING INTEREST(S): This study was supported by the Natural Science Foundation of Guangdong Province (2016A030313495), National Natural Science Foundation of China (81702567, 81671406, 31871412), the Science and Technology Programs of Guangdong (2017A050501021), Medical Science Technology Research Fund of Guangdong Province (A2018075), the Science and Technology Programs of Guangzhou City (201704030103), Internal Project of Family Planning Research Institute of Guangdong Province (S2018004), Post-doc initiation fund of Guangzhou (3302) and Post-doc science research initiation fund of Guangzhou Women and Children's Medical Center (20160322). There are no conflicts of interest.
Assuntos
Endometriose , Caderinas , China , Endométrio , Feminino , Humanos , Células EstromaisRESUMO
Objectives Leigh syndrome (LS) is one of the most common mitochondrial diseases and has variable clinical symptoms. However, the genetic variant spectrum of this disease is incomplete. Methods Next-generation sequencing (NGS) was used to identify the m.14430A > G (p.W82R) variant in a patient with LS. The pathogenesis of this novel complex I (CI) variant was verified by determining the mitochondrial respiration, assembly of CI, ATP, MMP and lactate production, and cell growth rate in cybrids with and without this variant. Results A novel m.14430A > G (p.W82R) variant in the NADH dehydrogenase 6 (ND6) gene was identified in the patient; the mutant loads of m.14430A > G (p.W82R) in the patient were much higher than those in his mother. Although the transmitochondrial cybrid-based study showed that mitochondrial CI assembly remains unaffected in cells with the m.14430G variant, control cells had significantly higher endogenous and CI-dependent mitochondrial respiration than mutant cells. Accordingly, mutant cells had a lower ATP, MMP and higher extracellular lactate production than control cells. Notably, mutant cells had impaired growth in a galactose-containing medium when compared to wild-type cells. Conclusions A novel m.14430A > G (p.W82R) variant in the ND6 gene was identified from a patient suspected to have LS, and this variant impaired mitochondrial respiration by decreasing the activity of mitochondrial CI.
Assuntos
Doença de Leigh/genética , NADH Desidrogenase/deficiência , NADH Desidrogenase/genética , Linhagem Celular Tumoral , Pré-Escolar , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Doença de Leigh/enzimologia , Masculino , Mutação de Sentido IncorretoRESUMO
Endometrial polyps (EPs) are outgrowths in the endometrium with unknown etiology. The fact that EPs can often recur after surgical removal suggests that EPs are not induced by random events but by continuous or recurrent processes in patients. We previously demonstrated that the risk of EP development was positively associated with overactive Th17 responses. However, the requirements of Th17 upregulation are yet unclear. Here, we recruited 26 women with symptomatic EP and 24 without EP, and peripheral mononuclear cells were harvested for the examination of circulating immunity. Compared to controls without EP, the patients with symptomatic EP presented significantly elevated levels of monocyte activation. The circulating monocytes from patients secreted higher levels of tumor necrosis factor (TNF), interleukin (IL)-1ß, IL-6 and IL-23 directly ex vivo and with LPS stimulation. In memory CD4+ T cells, monocytes were not required for IL-17 expression, but the presence of activated monocytes significantly increased the secretion of IL-17. In naive CD4+ T cells, activated monocytes were required for significant IL-17 secretion and RORC transcription. Interestingly, the monocytes from EP individuals were significantly more potent in promoting Th17 differentiation from naive CD4+ T cells than the monocytes from controls. Furthermore, we showed that monocyte-mediated Th17 differentiation required the secretion of TNF, IL-1ß and IL-6. Together, this study demonstrated activated monocytes supported Th17 inflammation in patients with EP.
Assuntos
Endométrio/patologia , Macrófagos/citologia , Pólipos/imunologia , Células Th17/citologia , Adulto , Linfócitos T CD4-Positivos/citologia , Estudos de Casos e Controles , Diferenciação Celular , Citocinas/biossíntese , Feminino , Humanos , Interleucina-17/biossíntese , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Monócitos/metabolismo , Pólipos/metabolismo , Recidiva , Células Th17/metabolismoRESUMO
Endometrial polyps (EPs) are localized benign overgrowths at the endometrium, with currently unknown aetiology and pathogenesis. Although symptoms of EP can be alleviated or resolved by hysteroscopic polypectomy, a significant fraction of individuals develop recurrent EPs after initial EP removal. In rare cases, EPs may also undergo malignant transformation. In-depth understanding of the mechanisms that are involved in EP development is urgently needed. Recent works indicate that dysregulations in the immune system participate in the development of a variety of symptoms, such as aging, obesity and hypertension, many of which are EP risk factors. Based on these discoveries, we investigated the cellular immune system in premenopausal women with and without EP. Compared to EP-free controls, the women with EP presented significantly higher RORC expression but unchanged TBX21 and FOXP3 expression in the circulating CD4+ T cells. When stimulated with PMA/ionomycin, CD4+ T cells from women with EP presented significantly higher interferon (IFN)-γ and interleukin (IL)-17 secretion, and lower transforming growth factor (TGF)-ß secretion. Hysteroscopic polypectomy did not significantly alter the composition of CD4+ T cells, as the women with EP presented a similar upregulation of Th17 inflammation and a downregulation of regulatory T cell (Treg) response postoperatively. Notably, in women that developed recurrent EP, the CD4+ T cells presented higher preoperative and postoperative RORC, IFN-γ, and IL-17 expression, as well as lower postoperative FOXP3 and TGF-ß expression, than hysteroscopic polypectomy-treated women without EP recurrence. These data demonstrated an association between CD4+ T cell imbalance and recurrent EP development.
Assuntos
Linfócitos T CD4-Positivos/citologia , Endométrio/patologia , Pólipos/imunologia , Adulto , Linfócitos T CD4-Positivos/metabolismo , Contagem de Células , Feminino , Regulação da Expressão Gênica , Humanos , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/genética , Pólipos/genética , Pólipos/cirurgia , Pré-Menopausa , Período Pré-Operatório , Recidiva , RiscoRESUMO
The increasing need for carbon-neutral, low-emission transportation sector has led to the development of advanced biofuels with tailor-made production and combustion processes. Even though the large-scale deployment of these advanced biofuels also increases the risk for their release into the environment, their toxic potency remains largely unknown. To identify hazardous biofuel candidates as early as possible, the fuel development process can be expanded by "Green Toxicology". To demonstrate such early Green Toxicology testing, this study investigates the aquatic toxicity for the two biofuel candidates 2-methyltetrahydrofuran (2-MTHF) and 2-methylfuran (2-MF) on Daphnia magna. We performed the prolonged acute immobilisation assay (96â¯h) and the D. magna reproduction test. 2-MF induced acute effects on D. magna that were two orders of magnitude stronger than those of 2-MTHF. Furthermore, both substances affected the growth and reproductive output of D. magna in a 21 d reproduction test, with 2-MF already inducing effects with concentrations one to two orders of magnitude lower than those of 2-MTHF. Thus, our assessment of the aquatic toxicity suggests that further biofuel development should focus on 2-MTHF. Furthermore, the acute immobilisation test with D. magna was identified as a promising tool for a rapid and sensitive "Green Toxicology" screening of further biofuel candidates.
Assuntos
Biocombustíveis/toxicidade , Daphnia/efeitos dos fármacos , Animais , Bioensaio , Furanos/toxicidade , Reprodução/efeitos dos fármacos , Testes de Toxicidade Aguda , Poluentes Químicos da Água/toxicidadeRESUMO
The arsenate (As(V)) biotransformation by Microcystis aeruginosa in a medium with different concentrations of nitrogen (N) and phosphorus (P) has been studied under laboratory conditions. When 15µg/L As(V) was added, N and P in the medium showed effective regulation on arsenic (As) metabolism in M. aeruginosa, resulting in significant differences in the algal growth among different N and P treatments. Under 0.2mg/L P treatment, increases in N concentration (4-20mg/L) significantly stimulated the cell growth and therefore indirectly enhanced the production of dimethylarsinic acid (DMA), the main As metabolite, accounting for 71%-79% of the total As in the medium. Meanwhile, 10-20mg/L N treatments accelerated the ability of As metabolization by M. aeruginosa, leading to higher contents of DMA per cell. However, As(V) uptake by M. aeruginosa was significantly impeded by 0.5-1.0mg/L P treatment, resulting in smaller rates of As transformation in M. aeruginosa as well as lower contents of As metabolites in the medium. Our data demonstrated that As(V) transformation by M. aeruginosa was significantly accelerated by increasing N levels, while it was inhibited by increasing P levels. Overall, both P and N play key roles in As(V) biotransformation processes.
Assuntos
Arseniatos/metabolismo , Biotransformação , Microcystis/metabolismo , Nitrogênio/química , Nitrogênio/metabolismo , Fósforo/química , Fósforo/metabolismo , Poluentes Químicos da ÁguaRESUMO
Haplogroup G predisposes one to an increased risk of osteoarthritis (OA) occurrence, while haplogroup B4 is a protective factor against OA onset. However, the underlying mechanism is not known. Here, by using trans-mitochondrial technology, we demonstrate that the activity levels of mitochondrial respiratory chain complex I and III are higher in G cybrids than in haplogroup B4. Increased mitochondrial oxidative phosphorylation (OXPHOS) promotes mitochondrial-related ATP generation in G cybrids, thereby shifting the ATP generation from glycolysis to OXPHOS. Furthermore, we found that lower glycolysis in G cybrids decreased cell viability under hypoxia (1% O2) compared with B4 cybrids. In contrast, G cybrids have a lower NAD(+)/NADH ratio and less generation of reactive oxygen species (ROS) under both hypoxic (1% O2) and normoxic (20% O2) conditions than B4 cybrids, indicating that mitochondrial-mediated signaling pathways (retrograde signaling) differ between these cybrids. Gene expression profiling of G and B4 cybrids using next-generation sequencing technology showed that 404 of 575 differentially expressed genes (DEGs) between G and B4 cybrids are enriched in 17 pathways, of which 11 pathways participate in OA. Quantitative reverse transcription PCR (qRT-PCR) analyses confirmed that G cybrids had lower glycolysis activity than B4 cybrids. In addition, we confirmed that the rheumatoid arthritis pathway was over-activated in G cybrids, although the remaining 9 pathways were not further tested by qRT-PCR. In conclusion, our findings indicate that mtDNA haplogroup G may increase the risk of OA by shifting the metabolic profile from glycolysis to OXPHOS and by over-activating OA-related signaling pathways.
Assuntos
DNA Mitocondrial/genética , Haplótipos , Mitocôndrias/genética , Proteínas Mitocondriais/genética , Osteoartrite/genética , Transdução de Sinais/genética , Linhagem Celular Tumoral , DNA Mitocondrial/metabolismo , Feminino , Humanos , Masculino , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Proteínas Mitocondriais/metabolismo , Osteoartrite/metabolismo , Fosforilação Oxidativa , Fatores de RiscoRESUMO
By using next-generation sequencing targeted to MitoExome including the entire mtDNA and exons of 1033 genes encoding the mitochondrial proteome, we described here a novel m.11240C>T mutation in the mitochondrial ND4 gene from a patient with Leigh syndrome. High mutant loads of m.11240C>T were detected in blood, urinary epithelium, oral mucosal epithelium cells, and skin fibroblasts of the patient. Decreased mitochondrial complex I activity was found in transmitochondrial cybrids containing the m.11240C>T mutation with biochemical analysis. Furthermore, functional investigations confirmed that mitochondria with the m.11240C>T variant exhibited lower adenosine triphosphate-related mitochondrial respiration. However, complex I assembly in mutant cybrids was not affected. While this mutation was located in the fourth hydrophobic trans-membrane region of ND4 gene, we suggested that mutation of m.11240C>T might impair the proton pumping channel of complex I but had little effect on the complex I assembly. In conclusion, we identified m.11240C>T as a novel mitochondrial disease-related mtDNA mutation.
Assuntos
DNA Mitocondrial/genética , Complexo I de Transporte de Elétrons/genética , Doença de Leigh/genética , Proteínas Mitocondriais/genética , NADH Desidrogenase/genética , Trifosfato de Adenosina/metabolismo , Substituição de Aminoácidos/genética , Sequência de Bases , Linhagem Celular Tumoral , Pré-Escolar , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Doença de Leigh/diagnóstico , Masculino , Mitocôndrias/genética , Mitocôndrias/metabolismo , Consumo de Oxigênio/fisiologia , Polimorfismo de Nucleotídeo Único , Análise de Sequência de DNARESUMO
A novel red-emitting Lu3Al5O12:Mn4+ (LuAG:Mn4+) phosphor was synthesized by a solid-state reaction. The emission-band position is shifted to red region by gradually increasing the amount of substitution of Lu3+ for Y3+, eventually yielding fully Y3Al5O12:Mn4+ (YAG:Mn4+). The compared structural and optical properties of the phosphors were investigated. From the experimentally measured spectroscopic data, crystal field parameter Dq and Racah parameters B and C are calculated to be 2127, 1464, and 3620 cm-1 in LuAG:Mn4+, respectively. In YAG:Mn4+, Dq, B, and C are calculated to be 2082, 1524, and 3740 cm-1, respectively. Impressively, Ca2+/Li+/Mg2+/Sr2+/Sc3+/Na+ dopants were found to be beneficial for enhancing Mn4+ luminescence, and the related mechanisms were systematically discussed.
RESUMO
Two series of phosphors of Ca8MgSc(PO4)7:Dy3+ and Sr8MgSc(PO4)7:Dy3+ single-phase white-emitting phosphors with high thermal emission stability are synthesized by the high-temperature solid-state reaction. The crystal structure, photoluminescence (PL), PL excitation (PLE), and thermal PL quenching spectra of Ca8MgSc(PO4)7:xDy3+ and Sr8MgSc(PO4)7:xDy3+ were investigated and compared in detail. Upon excitation at 387 nm, M8MgSc(PO4)7:xDy3+ (M = Ca/Sr) showed white emission centered at 480, 571, 660, and 754 nm. The white-emitting Dy-phosphor Ca8MgSc(PO4)7:Dy3+ (CMSP:Dy) had good terminal stability. The emission intensity of Ca8MgSc(PO4)7:Dy3+ still remained 95.2% of that at room temperature at 160 °C, and remained 77.3% at 300 °C under 387 nm excitation.
RESUMO
STUDY QUESTION: Is chloride channel-3 (ClC-3) involved in regulating the biological behavior of endometrial stromal cells (ESCs)? SUMMARY ANSWER: ClC-3 promotes endometriotic cell migration and invasion. WHAT IS KNOWN ALREADY: ClC-3 plays a significant role in the migration and invasion of various kinds of cells. STUDY DESIGN, SIZE, DURATION: An ITALIC! in vitro investigation of the effect of ClC-3 on the migration and invasion of ectopic ESCs from patients with endometriosis. PARTICIPANTS/MATERIALS, SETTING, METHODS: The ectopic and eutopic endometrial samples from 43 female patients with endometriosis and the endometrial samples from 39 non-endometriotic female patients were collected. Primary cells from these samples were isolated and cultured. Real-time RT-PCR, immunohistochemistry and western blot were used to detect the expression of ClC-3 and matrix metalloproteinase 9 (MMP-9). Small interfering RNA (siRNA) technology was employed to knock down ClC-3 expression. The migration and invasion ability of ESCs was measured by the transwell assay with uncoated or Matrigel-coated membranes. MAIN RESULTS AND THE ROLE OF CHANCE: The expression of ClC-3 mRNA and proteins was significantly up-regulated in the ectopic tissues from endometriotic patients, while that in the eutopic endometrial tissues of the same patients did not significantly differ from that in non-endometriotic patients. The migration and invasion ability and MMP-9 expression was increased in the ESCs from ectopic endometrial tissues. The knockdown of ClC-3 expression by ClC-3 siRNA inhibited ESC migration and invasion and attenuated the expression of MMP-9. ClC-3 expression level was well-correlated to the clinical characteristics and symptoms of endometriosis patients, including infertility, dysmenorrhea, chronic pelvic pain, dyspareunia and diameter of endometriosis lesion. LIMITATIONS, REASONS FOR CAUTION: Further studies are needed to examine the regulatory mechanism of estrogen on ClC-3 expression of ESCs. WIDER IMPLICATIONS OF THE FINDINGS: ClC-3 is involved in the migration and invasion processes of ESCs and can regulate MMP-9 expression. Up-regulation of ClC-3 expression may contribute to endometriosis development by regulating MMP-9 expression. STUDY FUNDING/COMPETING INTERESTS: This work was supported by the National Natural Science Foundation of China (81173064, 81272223, 81273539), the Ministry of Education of China (20124401110009), the Natural Science Foundation of Guangdong Province (S2011010001589) and the Science and Technology Programs of Guangdong (2013B051000059), Guangzhou (2013J500015) and Dongguan (2011108102006). The authors have no conflict of interest.