RESUMO
OBJECTIVE@#To investigate the effects of Danmu Extract Syrup (DMS) on lipopolysaccharide (LPS)-induced acute lung injury (ALI) in mice and explore the mechanism.@*METHODS@#Seventy-two male Balb/C mice were randomly divided into 6 groups according to a random number table (n=12), including control (normal saline), LPS (5 mg/kg), LPS+DMS 2.5 mL/kg, LPS+DMS 5 mL/kg, LPS+DMS 10 mL/kg, and LPS+Dexamethasone (DXM, 5 mg/kg) groups. After pretreatment with DMS and DXM, the ALI mice model was induced by LPS, and the bronchoalveolar lavage fluid (BALF) were collected to determine protein concentration, cell counts and inflammatory cytokines. The lung tissues of mice were stained with hematoxylin-eosin, and the wet/dry weight ratio (W/D) of lung tissue was calculated. The levels of tumor necrosis factor-α (TNF-α), interleukin (IL)-6 and IL-1 β in BALF of mice were detected by enzyme linked immunosorbent assay. The expression levels of Claudin-5, vascular endothelial (VE)-cadherin, vascular endothelial growth factor (VEGF), phospho-protein kinase B (p-Akt) and Akt were detected by Western blot analysis.@*RESULTS@#DMS pre-treatment significantly ameliorated lung histopathological changes. Compared with the LPS group, the W/D ratio and protein contents in BALF were obviously reduced after DMS pretreatment (P<0.05 or P<0.01). The number of cells in BALF and myeloperoxidase (MPO) activity decreased significantly after DMS pretreatment (P<0.05 or P<0.01). DMS pre-treatment decreased the levels of TNF-α, IL-6 and IL-1 β (P<0.01). Meanwhile, DMS activated the phosphoinositide 3-kinase/protein kinase B (PI3K/Akt) pathway and reversed the expressions of Claudin-5, VE-cadherin and VEGF (P<0.01).@*CONCLUSIONS@#DMS attenuated LPS-induced ALI in mice through repairing endothelial barrier. It might be a potential therapeutic drug for LPS-induced lung injury.
Assuntos
Camundongos , Masculino , Animais , Proteínas Proto-Oncogênicas c-akt/metabolismo , Lipopolissacarídeos , Fosfatidilinositol 3-Quinases/metabolismo , Interleucina-1beta/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Claudina-5/metabolismo , Lesão Pulmonar Aguda/induzido quimicamente , Pulmão/patologia , Interleucina-6/metabolismo , Medicamentos de Ervas ChinesasRESUMO
Transcription factor IIB (TFIIB) is a general transcription factor for RNA polymerase II, exerting its influence across various biological contexts. In the majority of eukaryotes, TFIIB typically has two homologs, serving as general transcription factors for RNA polymerase I and III. In plants, however, the TFIIB-related protein family has expanded greatly, with 14 and 9 members in Arabidopsis and rice, respectively. BRP5/pollen-expressed transcription factor 2 (PTF2) proteins belong to a subfamily of TFIIB-related proteins found only in plants and algae. The prior analysis of an Arabidopsis atbrp5 mutant, characterized by a T-DNA insertion at the 5' untranslated region, demonstrated the essential role of BRP5/PTF2 during the process of pollen germination and embryogenesis in Arabidopsis. Using a rice transformation system based on CRISPR/Cas9 technology, we have generated transgenic rice plants containing loss-of-function frameshift mutations in the BRP5/PTF2 gene. Unlike in the Arabidopsis atbrp5 mutant, the brp5/ptf2 frameshift mutations were not transmitted to progeny in rice, indicating an essential role of BRP5/PTF2 in both male and female gamete development or viability. The silencing of rice BRP5/PTF2 expression through RNA interference (RNAi) had little effect on vegetative growth and panicle formation but strongly affected pollen development and grain formation. Genetic analysis revealed that strong RNAi silencing of rice BRP5/PTF2 was still transmissible to progeny almost exclusively through female gametes, as found in the Arabidopsis atbrp5 knockdown mutant. Thus, reduced rice BRP5/PTF2 expression impacted pollen preferentially by interfering with male gamete development or viability. Drawing upon these findings, we posit that BRP5/PTF2 assumes a distinct and imperative function in the realm of plant sexual reproduction.
Assuntos
Oryza , Proteínas de Plantas , Fator de Transcrição TFIIB , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Gametogênese , Regulação da Expressão Gênica de Plantas , Oryza/genética , Oryza/metabolismo , Plantas/metabolismo , Pólen/metabolismo , Fator de Transcrição TFIIB/metabolismo , Proteínas de Plantas/metabolismoRESUMO
OBJECTIVE@#To investigate the effect of adipocytes in the bone marrow microenvironment of patients with multiple myeloma (MM) on the pathogenesis of MM.@*METHODS@#Bone marrow adipocytes (BMA) in bone marrow smears of health donors (HD) and newly diagnosed MM (ND-MM) patients were evaluated with oil red O staining. The mesenchymal stem cells (MSC) from HD and ND-MM patients were isolated, and in vitro co-culture assay was used to explore the effects of MM cells on the adipogenic differentiation of MSC and the role of BMA in the survival and drug resistance of MM cells. The expression of adipogenic/osteogenic differentiation-related genes PPAR-γ, DLK1, DGAT1, FABP4, FASN and ALP both in MSC and MSC-derived adipocytes was determined with real-time quantitative PCR. The Western blot was employed to detect the expression levels of IL-6, IL-10, SDF-1α, TNF-α and IGF-1 in the supernatant with or without PPAR-γ inhibitor.@*RESULTS@#The results of oil red O staining of bone marrow smears showed that BMA increased significantly in patients of ND-MM compared with the normal control group, and the BMA content was related to the disease status. The content of BMA decreased in the patients with effective chemotherapy. MM cells up-regulated the expression of MSC adipogenic differentiation-related genes PPAR-γ, DLK1, DGAT1, FABP4 and FASN, but the expression of osteogenic differentiation-related gene ALP was significantly down-regulated. This means that the direct consequence of the interaction between MM cells and MSC in the bone marrow microenvironment is to promote the differentiation of MSC into adipocytes at the expense of osteoblasts, and the cytokines detected in supernatant changed. PPAR-γ inhibitor G3335 could partially reverse the release of cytokines by BMA. Those results confirmed that BMA regulated the release of cytokines via PPAR-γ signal, and PPAR-γ inhibitor G3335 could distort PPAR-γ mediated BMA maturation and cytokines release. The increased BMA and related cytokines effectively promoted the proliferation, migration and drug resistance of MM cells.@*CONCLUSION@#The BMA and its associated cytokines are the promoting factors in the survival, proliferation and migration of MM cells. BMA can protect MM cells from drug-induced apoptosis and plays an important role in MM treatment failure and disease progression.
Assuntos
Humanos , Osteogênese/genética , Medula Óssea/metabolismo , Mieloma Múltiplo/metabolismo , Resistencia a Medicamentos Antineoplásicos , Receptores Ativados por Proliferador de Peroxissomo/farmacologia , Diferenciação Celular , Adipogenia , Citocinas/metabolismo , Adipócitos/metabolismo , Células da Medula Óssea/metabolismo , Células Cultivadas , PPAR gama/farmacologia , Microambiente TumoralRESUMO
Here,a styrene-based polymer monolithic column poly(VBS-co-TAT-co-AHM)with reversed-phase/hydrophilic interaction liquid chromatography(RPLC/HILIC)bifunctional separation mode was success-fully prepared for capillary electrochromatography by the in situ polymerization of sodium p-styrene sulfonate(VBS)with cross-linkers 3-(acryloyloxy)-2-hydroxypropyl methacrylate(AHM)and 1,3,5-triacryloylhexahydro-1,3,5-triazine(TAT).The preparation conditions of the monolith were optimized.The morphology and formation of the poly(VBS-co-TAT-co-AHM)monolith were confirmed by scanning electron microscopy(SEM)and Fourier transform infrared spectroscopy(FT-IR).The separation perfor-mances of the monolith were evaluated systematically.It should be noted that the incorporation of VBS functional monomer can provide π-π interactions,hydrophilic interactions,and ion-exchange in-teractions.Hence,the prepared poly(VBS-co-TAT-co-AHM)monolith can achieve efficient separation of thiourea compounds,benzene series,phenol compounds,aniline compounds and sulfonamides in RPLC or HILIC separation mode.The largest theoretical plate number for N,N'-dimethylthiourea reached 1.7×105 plates/m.In addition,the poly(VBS-co-TAT-co-AHM)monolithic column showed excellent reproducibility and stability.This novel monolithic column has great application value and potential in capillary electrochromatography(CEC).
RESUMO
Objective To explore the feasibility and accuracy of sex determination by morphology of the distal humerus in North China population. Methods Observing sexually dimorphic differences of olecranon fossa shape, trochlear symmetry, trochlear constriction and angle of medial epicondyle. Each characteristic was divided into three categories: male, unknown and female. The morphological characteristics of the humerus of 70 individuals (35 males and 35 females) from the Ming and Qing Dynasties in North China were counted to compare the occurrence rate of different categories of distal humeral morphology and the accuracy rate of sex determination. Results There were statistically significant differences in the performance of the four traits of the distal humerus between males and females. The accuracy rate of angle of medial epicondyle was the highest at 76%, while the lowest accuracy rate of trochlear constriction was only 52%. The accuracy of the combined four traits was higher than the single ones, with an accuracy rate of 76% (83% for males and 68% for females). Conclusion Distal humerus morphology can be used to determine sex when skulls and pelvis are absent.
RESUMO
Objective To investigate the effect of subchronic inhalation of toluene diisocyanate (TDI) on the pathological changes, oxidative stress damage, and HO-1 expression levels in rat liver tissues. Methods Forty healthy 3-week-old SPF-grade Sprague-Dawley male rats were randomly divided into 4 groups (control group, low-dose group, medium-dose group, and high-dose group), each with 10 rats. The rats were placed in a HOPE-MED 8050A movable poison cabinet in a cage. TDI was administered to animals by inhalation at doses of 0, 3.06 mg/m3, 12.25 mg/m3, and 49.00 mg/m3, respectively, for 6 hours a day and 5 days a week, and continuously for 13 weeks. The control group was exposed to fresh air. The effect of TDI on pathological changes, oxidative stress damage and HO-1 expression in rat liver tissues was examined. Results Compared with the control group, the rats in the medium and high-dose TDI-exposed groups exhibited vacuolar changes, hepatocyte swelling, steatosis and other pathological changes. With the increase of the TDI dose, the gap between hepatocytes was widened, mitochondria were swollen and vacuolated, and mitochondrial cristae disappeared. The expression levels of HO-1 gene and protein in the liver tissues of the low, medium, and high dose groups were significantly higher than those in the control group (P<0.05). Compared with the control group, the number of HO-1 positive cells in the low, medium and high dose groups increased and the staining increased gradually, and the difference was statistically significant (P<0.05). Conclusion TDI exposure can cause oxidative damage to rat liver tissues and induce the expression levels of HO-1 gene and protein expression.
RESUMO
Objective: To evaluate the impact of interventional therapy on top of drug therapy on cardiac function and structure in heart failure with reduced ejection fraction (HFrEF) patients complicating with middle aortic syndrome caused by Takayasu arteritis (TA-MAS). Methods: It was a retrospective longitudinal study. The data of patients with TA-MAS and HFrEF, who received interventional therapy on top of drug therapy in Fuwai Hospital from January 2010 to September 2020, were collected and analyzed. Baseline clinical data (including demographic data, basic treatment, etc.) were collected through the electronic medical record system. Changes of indexes such as New York Heart Association (NYHA) classification, N-terminal pro-brain natriuretic peptide (NT-proBNP), left ventricular end diastolic diameter (LVEDD), left ventricular ejection fraction (LVEF), left ventricular mass index (LVMI) before and after therapy were analyzed. Results: A total of 10 patients were collected. There were 8 females in this patient cohort, age was (18.4±5.0) years and onset age was (15.3±5.0) years. All 10 patients received standard heart failure medication therapy in addition to hormone and/or immunosuppressive anti-inflammatory therapy, but cardiac function was not improved, so aortic balloon dilatation and/or aortic stenting were performed in these patients. The median follow-up was 3.3(1.3, 5.6) years. On the third day after interventional therapy, the clinical symptoms of the 10 patients were significantly improved, NYHA classfication was restored from preoperative Ⅲ/Ⅳ to Ⅱ at 6 months post intervention(P<0.05). Compared with preoperation, NT-proBNP (P=0.028), LVEDD (P=0.011) and LVMI (P=0.019) were significantly decreased, LVEF was significantly increased (P<0.001) at 6 months after operation. Compared with preoperation, NT-proBNP (P=0.016), LVEDD (P=0.023) and LVMI (P=0.043) remained decreased, LVEF remained increased (P<0.001) at 1 year after operation. Conclusion: Results from short and medium term follow-up show that interventional therapy on top of heart failure drug therpay can effectively improve left cardiac function and attenuate cardiac remodeling in patients with TA-MAS comorbid with HFrEF.
Assuntos
Adolescente , Criança , Feminino , Humanos , Adulto Jovem , Masculino , Insuficiência Cardíaca/cirurgia , Estudos Longitudinais , Peptídeo Natriurético Encefálico , Fragmentos de Peptídeos , Estudos Retrospectivos , Volume Sistólico , Arterite de Takayasu/cirurgia , Função Ventricular Esquerda/efeitos dos fármacos , Ventrículos do Coração/efeitos dos fármacos , Fármacos Cardiovasculares/uso terapêutico , Angioplastia com Balão , Stents , Implante de Prótese VascularRESUMO
Allergen immunotherapy (AIT), also known as allergen vaccine desensitization, is currently the only treatment that can change the natural course of allergic diseases. AIT for pregnant women is a protective factor for neonatal allergies. Existing findings suggest that AIT should not be started during pre-pregnancy, pregnancy, or lactation periods, but can be continued if has already been started before; unexpected pregnancy during AIT does not require termination; AIT shows no obvious adverse effects on fetuses, infants, and young children. The latest evidence on the effectiveness and safety of AIT during pre-pregnancy, pregnancy, and lactation periods were reviewed.
RESUMO
Objective: To compare the efficacy and safety of prolonged dual antiplatelet therapy (DAPT>1 year) in patients with stable coronary artery disease (CAD) and diabetes who were event-free at 1 year after percutaneous coronary intervention (PCI) with drug-eluting stent (DES) in a large and contemporary PCI registry. Methods: A total of 1 661 eligible patients were selected from the Fuwai PCI Registry, of which 1 193 received DAPT>1 year and 468 received DAPT ≤1 year. The primary endpoint was major adverse cardiac and cerebrovascular event (MACCE) and Bleeding Academic Research Consortium (BARC) type 2, 3 or 5 bleeding, MACCE was defined as a composite of all-cause death, myocardial infarction or stroke. Multivariate Cox regression analysis and inverse probability of treatment weighting (IPTW) Cox regression analysis were performed. Results: After a median follow-up of 2.5 years, patients who received DAPT>1 year were associated with lower risks of MACCE (1.4% vs. 3.2%; hazard ratio (HR) 0.412, 95% confidence interval (CI) 0.205-0.827) compared with DAPT ≤1 year, which was primarily caused by the lower all-cause mortality (0.1% vs. 2.6%; HR 0.031, 95%CI 0.004-0.236). Risks of cardiac death (0.1% vs. 1.5%; HR 0.051, 95%CI 0.006-0.416) and definite/probable ST (0.3% vs. 1.1%; HR 0.218, 95%CI 0.052-0.917) were also lower in patients received DAPT>1 year than those received DAPT ≤ 1 year. No difference was found between the two groups in terms of BARC type 2, 3, or 5 bleeding (5.3% vs. 4.1%; HR 1.088, 95%CI 0.650-1.821). Conclusions: In patients with stable CAD and diabetes who were event-free at 1 year after PCI with DES, prolonged DAPT (>1 year) provides a substantial reduction in ischemic cardiovascular events, including MACCE, all-cause mortality, cardiac mortality, and definite/probable ST, without increasing the clinically relevant bleeding risk compared with ≤ 1-year DAPT. Further well-designed, large-scale randomized trials are needed to verify the beneficial effect of prolonged DAPT in this population.
Assuntos
Humanos , Doença da Artéria Coronariana/terapia , Diabetes Mellitus Tipo 2 , Quimioterapia Combinada , Stents Farmacológicos , Hemorragia , Intervenção Coronária Percutânea , Inibidores da Agregação Plaquetária/uso terapêutico , Medição de Risco , Resultado do TratamentoRESUMO
Objective: To explore the influential factors of job stress suffered by workers in railway stations, the level of job stress of were measured and subjective comfort of employees targeting to working environment were reported. Methods: In March 2019, a cluster sampling study was designed to collect the personal characteristics, job characteristics and subjective comfort degree of working environment of 432 employees in Chongqing railway stations. Meanwhile, job stress was assessed using the effort-reward imbalance scale. Chi-square test was used to compare the difference of occupational stress detection rate among different stratified factors such as occupational characteristics. Logistic regression was applied to analyze the influential factors of occupational stress. Results: The detection rate of job stress of workers in the railway stations was 31.02% (134/432) . The detection rate of job stress was higher among the divorced workers in railway stations, those earning less than 5, 000 yuan per month, those with 10-20 years' length of service, those who worked as a conductor and other workers including baggageman, station master on duty and assistant engineer (χ(2)=9.61, 14.76, 23.28, 11.06, P=0.008, 0.002, 0.000, 0.011) . The detection rate of job stress was higher among those whose working environment subjective feelings were uncomfortable, the differences were statistically significant (P<0.001) . The results showed that the occupational stress of the staff in the railway stations was influenced by their subjective feeling of air quality, noise and Space Layout (P<0.05) . The risk factors of occupational stress were air quality, noise and uncomfortable space layout (OR=0.571, 0.068, 0.441, P=0.051, 0.054, 0.007) . Conductor, other (Bellboy, Duty Station Master, assistant engineer) were the risk factors of occupational stress (OR=1.884, 2.703, P=0.065, 0.019) . The employees of station A and station B were the risk factors of occupational stress (OR=4.681, 1.811, P=0.002, 0.067) . Conclusion: The higher detection rate of job stress of workers in the railway stations is correlated with the subjective comfort degree of the working environment of the workers.
Assuntos
Humanos , Estudos Transversais , Satisfação no Emprego , Estresse Ocupacional/epidemiologia , Estresse Psicológico , Inquéritos e Questionários , Local de TrabalhoRESUMO
Today, there is greater awareness on the association between oral diseases and respiration diseases after the outbreak of COVID-19. However, confusion regarding the oral health management and medical risk prevention for patients with chronic airway diseases has been remained among dental clinicians. Therefore, the dental experts of the Fifth General Dentistry Special Committee, Chinese Stomatological Association, combined with the experts of respiratory and critical care medicine, undertook the formation of consensus on the oral health management of patients with chronic airway diseases in order to help dental clinicians to evaluate medical risks and make better treatment decision in clinical practice. In the present consensus report, the relationship of oral diseases and chronic airway diseases, the oral health management and the treatment recommendations of patients with chronic airway diseases are provided.
Assuntos
Humanos , COVID-19 , Consenso , Saúde Bucal , Medicina BucalRESUMO
Objective: To investigate the sensitization characteristics of Juniperus chinensis pollen in patients with allergic rhinitis and/or allergic asthma in Beijing area, and to explore the characteristics of Juniper chinensis pollen sensitized population. Methods: Patients with suspected allergic rhinitis and/or asthma from January 2017 to December 2019 in the outpatient department of Allergy Department of Beijing Shijitan Hospital were selected in this study. Skin prick test (SPT) was performed with Juniper chinensis pollen allergen reagent to compare different age and disease allergen distribution, and to observe the sensitization characteristics of its population. All of the analyses were performed using SAS software version 9.4. Results: A total of 8 380 patients were enrolled in the end. The total positive rate of Juniper chinensis pollen SPT reached 49.92% (4 183/8 380). The positive rate of Juniper chinensis pollen SPT was highest in the 10-14 age group, reaching 60.99% (283/464). Compared with other age groups, there was a statistical difference (χ²=266.77, P<0.01). The SPT positive rate of patients aged less than 10 years increased with the increase of age, while the SPT positive rate of patients aged over 40 years decreased with the increase of age. Single Juniper chinensis pollen was less allergenic, accounting for about 25.05% (1 048/4 183), and the patients' age was (35.21±12.39) years. Regardless of single Juniper chinensis pollen or other pollen allergies, allergic rhinitis was the main disease. Among the patients with SPT positive Juniper chinensis pollen combined with other inhaled pollen allergens, willow pollen accounted for the first (74.99%). The positive rate of Juniper chinensis pollen was the highest in patients with single allergic rhinitis, accounting for 52.05% (3 797/7 295), and the rate in patients with single allergic asthma was the lowest, accounting for 17.49% (53/303), with statistically difference (χ²=138.99, P<0.01). Conclusions: Juniper chinensis pollen is highly sensitized in patients with allergic rhinitis and/or allergic asthma in Beijing . The positive rate of SPT is highest among 10-14 age group, most of which showed strong positive reaction, and allergic rhinitis is more common in Juniper chinensis pollen sensitization diseases.
Assuntos
Adolescente , Adulto , Criança , Humanos , Alérgenos , Asma , Juniperus , Pólen , Rinite Alérgica , Testes CutâneosRESUMO
Abnormal angiogenesis and regression of the diseased retinal vasculature are key processes associated with ischemic retinopathies, but the underlying mechanisms that regulate vascular remodeling remain poorly understood. Here, we confirmed the specific expression of semaphorin 3G (Sema3G) in retinal endothelial cells (ECs), which was required for vascular remodeling and the amelioration of ischemic retinopathy. We found that Sema3G was elevated in the vitreous fluid of patients with proliferative diabetic retinopathy (PDR) and in the neovascularization regression phase of oxygen-induced retinopathy (OIR). Endothelial-specific Sema3G knockout mice exhibited decreased vessel density and excessive matrix deposition in the retinal vasculature. Moreover, loss of Sema3G aggravated pathological angiogenesis in mice with OIR. Mechanistically, we demonstrated that HIF-2α directly regulated Sema3G transcription in ECs under hypoxia. Sema3G coordinated the functional interaction between ß-catenin and VE-cadherin by increasing ß-catenin stability in the endothelium through the neuropilin-2 (Nrp2)/PlexinD1 receptor. Furthermore, Sema3G supplementation enhanced healthy vascular network formation and promoted diseased vasculature regression during blood vessel remodeling. Overall, we deciphered the endothelium-derived Sema3G-dependent events involved in modulating physiological vascular remodeling and regression of pathological blood vessels for reparative vascular regeneration. Our findings shed light on the protective effect of Sema3G in ischemic retinopathies.
Assuntos
Endotélio Vascular/metabolismo , Isquemia/metabolismo , Doenças Retinianas/metabolismo , Vasos Retinianos/metabolismo , Semaforinas/metabolismo , Remodelação Vascular , beta Catenina/metabolismo , Animais , Endotélio Vascular/patologia , Feminino , Humanos , Isquemia/genética , Isquemia/patologia , Masculino , Camundongos , Camundongos Transgênicos , Doenças Retinianas/genética , Doenças Retinianas/patologia , Vasos Retinianos/patologia , Semaforinas/genética , beta Catenina/genéticaRESUMO
As sessile organisms, plants have evolved unique patterns of growth and development, elaborate metabolism and special perception and signaling mechanisms to environmental cues. Likewise, plants have complex and highly special programs for transcriptional control of gene expression. A case study for the special transcription control in plants is the expansion of general transcription factors, particularly the family of Transcription Factor IIB (TFIIB)-like factors with 15 members in Arabidopsis. For more than a decade, molecular and genetic analysis has revealed important functions of these TFIIB-like factors in specific biological processes including gametogenesis, pollen tube growth guidance, embryogenesis, endosperm development, and plant-microbe interactions. The redundant, specialized, and diversified roles of these TFIIB-like factors challenge the traditional definition of general transcription factors established in other eukaryotes. In this review, we discuss general transcription factors in plants with a focus on the expansion and functional analysis of plant TFIIB-like proteins to highlight unique aspects of plant transcription programs that can be highly valuable for understanding the molecular basis of plant growth, development and responses to stress conditions.
Assuntos
Proteínas de Arabidopsis/metabolismo , Fator de Transcrição TFIIB/fisiologia , Proteínas de Arabidopsis/genética , Proteínas Arqueais/metabolismo , Proteínas de Bactérias/metabolismo , Eucariotos/fisiologia , Regulação da Expressão Gênica de Plantas , Proteínas de Plantas/fisiologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
OBJECTIVE@#To investigate the effect of gap junction intercellular communication (GJIC) combined by connexin43 (Cx43) and its signal to the biobehavior of multiple myeloma (MM) cells, and its possible mechanism.@*METHODS@#The mesenchymal stem cell (MSC) cells were isolated and cultured from patients with MM and normal donors. The expression of connexin43 (Cx43) in MSC cells from different sources was detected by RT-PCR and Western blot. The side population (SP) cells were sorted by flow cytometry (FCM). The effect of MSC cells from different sources to the cell cycle, Cx43 expression, colony formation in vitro, stem cell related genes expression, cytokines secretion and chemoresistance in MM SP cells as well as with or without Cx43 inhibitor 18α-glycyrrhetinic acid (18α-GA) was observed.@*RESULTS@#There was no significantly difference between the MSC isolated from normal donor and MM patients. Western blot showed that Cx43 expression in SP cells was up-regulated when the cells were incubated with MSC, and medium containing 18α-GA could partially inhibit it, moreover, it was more significant in MSC cells of MM patients. The ability of colony formation of SP cells in vitro was higher than those of MM cells and MM-MSC could promote the colony formation in a co-culture manner. The effect of MM-MSC to SP cells was down-regulated after 18α-GA was added. RT-PCR showed that there was several important stem cell-related genes including c-myc, Oct-4 Klf-4, and Sox-2 were found in RPMI 8226 cells, but those cells were up-regulated in SP cells (P0.05). Cytometry bead array assays showed that MM-MSCs could secrete high level of IL-6, but the levels of IL-6, IL-10 and TGF-β increased significantly when the MM-MSCs were co-cultured with SP cells (P<0.05), especially the levels of IL-6 and IL-10 were significantly higher than cultured alone. There was no significant change in the levels of bFGF and IL-17 before and after co-cultured. The levels of IL-6, IL-10 and TGF-β in supernatant decreased significantly after GJ inhibitor 18α-GA was added. PI/Annexin V assay showed that MM cells were sensitive to bortezomib (BTZ)-induced apoptosis, but the sensitivity for SP cells was weaker. The ratio of cell apoptosis was 75.2%±0.77% and 8.12%±0.86% (P<0.001), respectively. MM-MSC could down-regulate the cell apoptosis induced by BTZ, while the sensitivity of MM cells to BTZ could be partially recovered after GJ inhibitor was added.@*CONCLUSION@#MSC derived from MM patients can enhance GJIC to maintain its "hematopoiesis" by up-regulating the expression of Cx43 in MM cells, and at the same time promote cell proliferation and drug recistance by secreting multiple cytokines, which finally contributes to the relapse of MM.
Assuntos
Humanos , Comunicação Celular , Técnicas de Cocultura , Conexina 43 , Células-Tronco Mesenquimais , Mieloma MúltiploRESUMO
Objective:To investigate the clinical effects of minimally invasive treatment of clavicular midshaft fracture with bridge-combined internal fixation system.Methods:A retrospective analysis was conducted of the 54 patients with clavicular midshaft fracture who had been admitted to Department of Trauma Orthopedics, The First Affiliated Hospital to Dali University from September 2016 to January 2018.They were assigned into 2 groups according to different treatment methods. In the observation group of 27 patients who were subjected to minimally invasive treatment with bridge-combined internal fixation system, there were 14 males and 13 females with an age of (41.8±14.8) years, and 10 cases of type 6-A1 and 17cases of type 6-C1 by the AO classification.In the control group of 27 patients who were subjected to traditional open reduction and internal plate fixation, there were 15 males and 12 females with an age of (39.6±14.1) years, and 11 cases of type 6-A1 and 16 cases of type 6-C1 by the AO classification.The 2 groups were compared in terms of operation time, incision length, intraoperative blood loss, fracture healing time, shoulder functional recovery and incidence of postoperative complications.Results:There was no significant difference between the 2 groups in their preoperative general data, showing comparability ( P>0.05). The 54 patients were followed up for an average of 13 months (from 9 to 25 months).The observation group had an incision length of (4.34±0.65)cm, significantly shorter than that [(10.64±2.14) cm] in the control group, an intraoperative blood loss of (20.9±6.2)mL, significantly less than that [(40.7±13.5) mL] in the control group, and a fracture healing time of (4.5±0.9) months, significantly shorter than that [(5.7±1.0) months] in the control group (all P<0.05). According to the Neer scoring for shoulder function at the last follow-up, the observation group had 22 excellent, 4 good and one fair cases, giving an excellent and good rate of 96.3% (26/27) while the control group had 21 excellent, 4 good, one fair and one poor cases, giving an excellent and good rate of 92.6% (25/27). There was no significant difference in operation time or functional results between the 2 groups ( P>0.05).One case of nonunion and plate breakage was observed in the control group. Conclusion:In the treatment of clavicular midshaft fracture, compared with traditional open reduction and plate fixation, the bridge-combined internal fixation system may lead to less surgical trauma, a smaller incision and faster fracture healing.
RESUMO
CRISPR/Cas-based mRNA imaging has been developed to labeling of high-abundance mRNAs. A lack of non-genetically encoded mRNA-tagged imaging tools has limited our ability to explore the functional distributions of endogenous low-abundance mRNAs in cells. Here, we developed a CRISPR-Sunspot method based on the SunTag signal amplification system that allows efficient imaging of low-abundance mRNAs with CRISPR/Cas9. Methods: We created a stable TRE3G-dCas9-EGFP cell line and generated an Inducible dCas9-EGFP imaging system for assessment of two factors, sgRNA and dCas9, which influence imaging quality. Based on SunTag system, we established a CRISPR-Sunspot imaging system for amplifying signals from single-molecule mRNA in live cells. CRISPR-Sunspot was used to track co-localization of Camk2a mRNA with regulatory protein Xlr3b in neurons. CRISPR-Sunspot combined with CRISPRa was used to determine elevated mRNA molecules. Results: Our results showed that manipulating the expression of fluorescent proteins and sgRNA increased the efficiency of RNA imaging in cells. CRISPR-Sunspot could target endogenous mRNAs in the cytoplasm and amplified signals from single-molecule mRNA. Furthermore, CRISPR-Sunspot was also applied to visualize mRNA distributions with its regulating proteins in neurons. CRISPR-Sunspot detected the co-localization of Camk2a mRNA with overexpressed Xlr3b proteins in the neuronal dendrites. Moreover, we also manipulated CRISPR-Sunspot to detect transcriptional activation of target gene such as HBG1 in live cells. Conclusion: Our findings suggest that CRISPR-Sunspot is a novel applicable imaging tool for visualizing the distributions of low-abundance mRNAs in cells. This study provides a novel strategy to unravel the molecular mechanisms of diseases caused by aberrant mRNA molecules.
Assuntos
Sistemas CRISPR-Cas/genética , Microscopia Intravital/métodos , Imagem Molecular/métodos , RNA Mensageiro/metabolismo , Imagem Individual de Molécula/métodos , Animais , Linhagem Celular Tumoral , Embrião de Mamíferos , Feminino , Hemoglobina Fetal/genética , Células HEK293 , Humanos , Microscopia Confocal/métodos , Neurônios , Cultura Primária de Células , RNA Guia de Cinetoplastídeos/genética , RNA Mensageiro/genética , Ratos , Ativação Transcricional , TransfecçãoRESUMO
Blood-brain barrier (BBB) dysfunction has been suggested to play an important role in epilepsy. However, the mechanism mediating the transition from cerebrovascular damage to epilepsy remains unknown. Here, we report that endothelial cyclin-dependent kinase 5 (CDK5) is a central regulator of neuronal excitability. Endothelial-specific Cdk5 knockout led to spontaneous seizures in mice. Knockout mice showed increased endothelial chemokine (C-X-C motif) ligand 1 (Cxcl1) expression, decreased astrocytic glutamate reuptake through the glutamate transporter 1 (GLT1), and increased glutamate synaptic function. Ceftriaxone restored astrocytic GLT1 function and inhibited seizures in endothelial Cdk5-deficient mice, and these effects were also reversed after silencing Cxcl1 in endothelial cells and its receptor chemokine (C-X-C motif) receptor 2 (Cxcr2) in astrocytes, respectively, in the CA1 by AAV transfection. These results reveal a previously unknown link between cerebrovascular factors and epileptogenesis and provide a rationale for targeting endothelial signaling as a potential treatment for epilepsy.
Assuntos
Quimiocina CXCL1/metabolismo , Quinase 5 Dependente de Ciclina/metabolismo , Células Endoteliais/metabolismo , Epilepsia/metabolismo , Gliose/metabolismo , Receptores de Interleucina-8B/metabolismo , Animais , Astrócitos/metabolismo , Astrócitos/patologia , Barreira Hematoencefálica/metabolismo , Células Cultivadas , Células Endoteliais/patologia , Epilepsia/patologia , Gliose/patologia , Ácido Glutâmico/metabolismo , Camundongos , Camundongos Knockout , Neurônios/metabolismo , Neurônios/patologia , Convulsões/metabolismo , Convulsões/patologia , Transdução de Sinais/fisiologiaRESUMO
Objective: To analyze the expression profile of plasma microRNA (miRNA) in patients with mild cognitive impairment (MCI) due to Alzheimer's disease (AD) by bioinformatics method, and explore its pathogenesis at the level of genetic regulation. Methods: Five MCI patients due to AD and five control participants were recruited. The plasma miRNA expression profiles were analyzed by miRNA microarray sequencing. Target genes of significantly up-regulated miRNAs were detected by TargetScan 7.2 database. The miRNA-gene interaction network of significantly up-regulated miRNAs was established by Cytoscape software, and the key miRNAs of the network were analyzed. The target genes of key miRNAs were analyzed by Gene Ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genome (KEGG) pathway analysis using R packages. Results: There are 13 up-regulated miRNAs in the plasma of MCI patients due to AD, and 5 of them were key miRNAs in miRNA-gene interaction network. Target genes of these miRNAs were mainly involved in biological process such as synaptic plasticity reg-ulation, Wnt signaling pathway, synaptic vesicle transport and synaptic vesicle localization, as well as Ras signaling pathway, mitogen-activated protein kinase (MAPK) signaling pathway and glycolysis/gluconeogenesis pathway. Conclusion: Five up-regulated miRNAs in plasma of MCI due to AD may be the main regulators involved in the pathological mechanism of AD, which can be used as potential biomarkers for diagnosis of MCI due to AD.
RESUMO
We compared the pharmacokinetic and pharmacodynamic profiles of desmopressin acetate after intraocular, intravenous and intragastric administration in rabbits to better understand the systemic delivery of peptide drugs through intraocular administration. Fifteen rabbits were randomly divided into three groups (intraocular administration, 7 μg·kg-1; intravenous administration, 0.7 μg·kg-1; and intragastric administration, 7 μg·kg-1). Blood samples were taken from the heart at predetermined time points after dosing and the plasma desmopressin concentration was analyzed by enzyme-linked immunosorbent assay (ELISA). Another 21 rabbits were randomly divided into three groups (intraocular administration, 7 μg·kg-1; intravenous administration, 0.7 μg·kg-1; intragastric administration, 7 μg·kg-1) for a pharmacodynamics study. Urine was collected at predetermined intervals after dosing. The pharmacokinetic parameters after intravenous administration were as follows: Cmax was 143.0 pg·mL-1; the area under the plasma concentration–time curve for desmopressin (AUC0-t) was 999.9 pg·h·mL-1. The pharmacokinetic parameters after intraocular administration were as follows: tmax was 5 min, Cmax was 125.6 pg·mL-1, AUC0-t was 873.1 pg·h·mL-1, and absolute bioavailability (F) was 8.7%. The pharmacokinetic parameters after intragastric administration were as follows: tmax was 10 min, Cmax was 104.1 pg·mL-1, AUC0-t was 451.8 pg·h·mL-1, and absolute bioavailability was 4.5%. Intraocular administration and intravenous administration of one tenth of the dosage showed a similar effect, and the urine volume remained decreased for 12 h, but urine volume increased significantly in the second collection period after intragastric administration, and there was no decrease in volume 12 h after dosing. This study demonstrates that peptide drugs such as desmopressin can be absorbed more rapidly after intraocular administration than after intragastric administration and can exert systemic therapeutic effects. In this study, the program of animal testing had been approved by the Laboratory Animal Care and Use Committee at Anhui University of Chinese Medicine.