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Nanopore sequencing has been widely used for the real-time detection and surveillance of pathogens with portable MinION. Nanopore adaptive sequencing can enrich on-target sequences without additional pretreatment. In this study, the performance of adaptive sequencing was evaluated for viral genome enrichment of clinical respiratory samples. Ligation-based nanopore adaptive sequencing (LNAS) and rapid PCR-based nanopore adaptive sequencing (RPNAS) workflows were performed to assess the effects of enrichment on nasopharyngeal swab samples from human adenovirus (HAdV) outbreaks. RPNAS was further applied for the enrichment of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from nasopharyngeal swab samples to evaluate sensitivity and timeliness. The RPNAS increased both the relative abundance (7.87-12.86-fold) and data yield (1.27-2.15-fold) of HAdV samples, whereas the LNAS increased only the relative abundance but had no obvious enrichment on the data yield. Compared with standard nanopore sequencing, RPNAS detected the SARS-CoV-2 reads from two low-abundance samples, increased the coverage of SARS-CoV-2 by 36.68-98.92%, and reduced the time to achieve the same coverage. Our study highlights the utility of RPNAS for virus enrichment directly from clinical samples, with more on-target data and a shorter sequencing time to recover viral genomes. These findings promise to improve the sensitivity and timeliness of rapid identification and genomic surveillance of infectious diseases.
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Background: Comparing the outcomes of debridement and total hip arthroplasty (THA) with antibiotic-loaded spacer implantation and subsequent THA for the treatment of patients affected by primary advanced septic arthritis (SA) of the hip in adults. Methods: All of the 20 patients (20 hips) underwent two-stage surgery. Nine patients were submitted to surgical debridement first and then THA (group 1), while 11 patients were treated with antibiotic-loaded spacer and subsequent THA (group 2). Patients were evaluated based on the recurrence of infection, Harris hip score, visual analogue scale (VAS) pain score, and leg length discrepancy. Results: No cases of infection, deep vein thrombosis, death, and loosening of the hip prosthesis were observed during follow-up. The mean follow-up time was 29.09 ± 10.80 months in group 1 and 28.22 ± 14.80 months in group 2. Before the THA surgery, the mean leg length discrepancy was 2.80 ± 2.03 cm in group 1 and 0.50 ± 0.23 cm in group 2 (P < 0.05). In the latest follow-up, the Harris hip scores of patients were 90.33 ± 4.85 in group 1 and 94.36 ± 2.34 in group 2 (P < 0.05), respectively. There was no statistically significant difference in the VAS pain score of the hip between the two groups (P > 0.05). Conclusions: Debridement and antibiotic-loaded spacer and subsequent THA were effective in eradicating the infection for advanced SA. However, antibiotic-loaded spacer and subsequent THA was superior for effectively maintaining the length of the lower limb and function of the hip.
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Artrite Infecciosa , Artroplastia de Quadril , Prótese de Quadril , Adulto , Antibacterianos/uso terapêutico , Artrite Infecciosa/tratamento farmacológico , Artrite Infecciosa/etiologia , Artrite Infecciosa/cirurgia , Artroplastia de Quadril/efeitos adversos , Articulação do Quadril/cirurgia , Prótese de Quadril/efeitos adversos , Humanos , Dor , Estudos Retrospectivos , Resultado do TratamentoRESUMO
OBJECTIVE: To explore whether there exist undiscovered transphyseal vasculature-canal compound structures in immature femurs and tibias, and reveal their potential oncological impact. METHODS: This investigation was divided into a morphological study and a clinical study. In the morphological part, a new-identified anatomic structure was investigated by using radiographical, anatomical, and histological methodologies. Twenty-eight 1-mm-slice thickness magnetic resonance images of pediatric knees were generated and 10 pediatric knees were dissected to verify the existence and universality, observe the radiographic and anatomic characteristics, and determined the located region of this structure. Hematoxylin-eosin staining, immunofluorescence, and angiography procedures were performed to illustrate its histological feature, molecular identification, and vascular origination, respectively. In the clinical part, 38 pediatric osteosarcoma patients were enrolled from January 2014 to December 2020. A descriptive clinical study including 13 typical participants was conducted to investigate the oncological significance of this new-identified structure. Meanwhile, the discrepancy in transphyseal osteosarcoma extension between different physeal regions was evaluated in a cross-sectional study. RESULTS: In the morphological study, we discovered a new-found vasculature-canal compound structure, intercondylar transphyseal complex (ITC), which originated from the middle genicular vessels, traversed the whole epiphysis, and breached the intact open physis in the immature proximal tibia or distal femur. The components of ITC included the juxta-articular, epiphyseal, and transphyseal segments of vessels, the canals that traverse the entire epiphysis and physis and enclosed the vessels, vascular foramina on articular facet and foramina-covered synovium. Depending on the location, ITCs can be divided into three types: femoral ITC, anterior tibial ITC, and posterior tibial ITC. Clinically, the ITC may facilitate intercondylar transphyseal sarcomatous dissemination without damaging the adjacent physeal cartilage. Compared to bilateral condylar physes, more osteosarcomas transgressed the open growth plates through intercondylar regions in which ITC was located (P = 0.022). CONCLUSION: As the "gap" on intact open physis, ITC, which is a new-identified compound structure in intercondylar regions of immature femur or tibia, may promote intercondylar transphyseal tumor extension. Moreover, the identification and characterization of ITC subvert some traditional comprehensions about physis and may provide novel perspectives for pediatric osteosarcomas.
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Neoplasias Ósseas , Osteossarcoma , Ligamento Cruzado Anterior , Neoplasias Ósseas/diagnóstico por imagem , Criança , Estudos Transversais , Epífises/diagnóstico por imagem , Fêmur/anatomia & histologia , Fêmur/diagnóstico por imagem , Lâmina de Crescimento , Humanos , Osteossarcoma/diagnóstico por imagem , Tíbia/diagnóstico por imagemRESUMO
Objective: This research aims to refresh the limited understanding about the canal and vascular structures within the epiphysis and metaphysis of the tibia and femur and their oncological significance. Methods: This study was started with characterization of a novel structure using radiographs and anatomic dissections, followed by a descriptive clinical study with 55 participants to investigate the effects of tumors on this novel discovery and a retrospective cohort study with 82 participants to investigate whether the structure would be a risk factor for tumor recurrence after the curettage of giant cell tumor of bone. Results: A new anatomical knee structure, the Lijianmin-Chengkun (LC) complex, was discovered in healthy adults, and its clinical implications were examined in this study. This new-found anatomical structure is composed of an epiphyseal and metaphyseal canal which surrounds a blood vessel, foramen, and foramen-covered synovium. All LC complexes showed similar radiographical, anatomical, and histological characteristics and were located within specific tibial and femoral intercondylar regions. These LC complexes seem to facilitate tumor residue and extension and may be a risk factor for tumor recurrence after curettage of femoral and tibial giant cell tumors (P = 0.031). Conclusion: The LC complexes are related to local tumor recurrence and bidirectional tumor dissemination between intraosseous and intraarticular regions. These findings have opened up a new perspective and may provide new targets for intervention in malignant and aggressive tumors around the knee joint.
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The aim of this study was to investigate the expression of interferon regulatory factor 4 (IRF4) and the inflammatory response in secondary injury of intracerebral haemorrhage (ICH). Twelve SD rats were randomly divided into a sham group and an ICH group, with 6 rats in each group. A rat model of ICH was established by injecting collagenase type IV into the right striatum of rats. The expression of IRF4 was measured by western blot and immunohistochemistry 48 h after ICH. In addition, 15 mm of hemin-induced PC12 cell injury was used to simulate an in vitro ICH model. IRF4 expression was detected by immunofluorescence (IF). Moreover, the inflammatory cytokines (IL-1b and IL-6) were measured by ELISA. The behavioural score of ICH rats was the lowest at 48 h after operation. The expression of IRF4 was significantly higher in the striatal tissue of ICH rats compared with the sham group (p < 0.05). Meanwhile, IF results showed that hemin induced the upregulation of IRF4 expression in rat pheochromocytoma cells PC12. In addition, IL-1b and IL-6 levels were significantly increased in the serum of ICH rats and in the supernatant of hemin-induced PC12 cells (p < 0.01). The inflammation in ICH is related to the increase of IRF4. It provides a theoretical basis for the clinical treatment of ICH.
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Hemorragia Cerebral , Inflamação , Animais , Citocinas/metabolismo , Fatores Reguladores de Interferon , Ratos , Ratos Sprague-DawleyRESUMO
BACKGROUND: Lung biopsy tissue samples can be used for infection detection and cancer diagnosis. Metagenomic next-generation sequencing (mNGS) has the potential to further improve diagnosis. METHODS: From July 2018 to May 2020, lung biopsy samples of 133 patients with suspected pulmonary infection or abnormal imaging findings were collected and subjected to clinical microbiological testing, Illumina and Nanopore sequencing to identify pathogens. The neural networks were pretrained by extracting features of human reads from 2,095 metagenomic next-generation sequencing results, and the human reads of lung biopsy samples were entered into the validated pipeline to predict the risk of cancer. FINDINGS: Based on the pathogen-cancer detection pipeline, the Illumina platform showed 77·6% sensitivity and 97·6% specificity compared to the composite reference standard for infection diagnosis. However, the Nanopore platform showed 34·7% sensitivity and 98·7% specificity. mNGS identified more fungi, which was confirmed by subsequent pathological examination. M. tuberculosis complex was weakly detected. For cancer detection, compared with histology, the Illumina platform showed 83·7% sensitivity and 97·6% specificity, diagnosing an additional 36 cancer patients, of whom half had abnormal imaging findings (pulmonary shadow, space-occupying lesions, or nodules). INTERPRETATION: For the first time, we have established a pipeline to simultaneously detect pathogens and cancer based on Illumina sequencing of lung biopsy tissue. This pipeline efficiently diagnosed cancer in patients with abnormal imaging findings. FUNDING: This work was supported by the National Key Research and Development Program of China and National Natural Science Foundation of China.
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Biópsia , Pneumopatias/diagnóstico , Pneumopatias/etiologia , Pulmão/patologia , Metagenômica , Neoplasias/complicações , Adulto , Idoso , Biópsia/métodos , Gerenciamento Clínico , Suscetibilidade a Doenças , Feminino , Instabilidade Genômica , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Metagenoma , Metagenômica/métodos , Pessoa de Meia-Idade , Neoplasias/diagnóstico , Adulto JovemRESUMO
Glioma is one of the most commonly observed tumours, representing approximately 75% of brain tumours in the adult population. Generally, glioma therapy includes surgical resection followed by radiotherapy and chemotherapy. The transcription factor STAT3 (signal transducer and activator of transcription 3) is a promising target for the treatment of cancer and several other diseases. At nanomolar concentrations, SD-36 induces rapid cellular degradation of STAT3 but cannot degrade other STAT proteins. The current study demonstrates the therapeutic efficacies of the STAT3 degraders SD-36 against glioma, as well as understanding the elucidating mechanisms and identifying molecular markers that determine cell sensitivity to STAT3 degraders. Glioma cell lines possessed similar response patterns to SD-36 but different responses to the STAT3 inhibitor Stattic. SD-36 potently induced apoptosis in glioma cells along with a reduction in Mcl-1 levels, which are critical for mediating the induction of apoptosis and enhancing TMZ-induced apoptosis. Accordingly, SD-36 sensitizes the antitumour effect of TMZ in patient-derived xenograft. In addition, the downregulation of Mcl-1 expression-mediated antitumour effect of SD-36 was analysed in cell-derived xenograft. These observations need to be validated clinically to confirm the efficacy of STAT3 degraders in glioma.
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Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Neoplasias do Sistema Nervoso Central , Glioma , Fator de Transcrição STAT3/antagonistas & inibidores , Animais , Linhagem Celular Tumoral , Neoplasias do Sistema Nervoso Central/tratamento farmacológico , Neoplasias do Sistema Nervoso Central/metabolismo , Glioma/tratamento farmacológico , Glioma/metabolismo , Humanos , CamundongosRESUMO
Traumatic spinal cord injury (SCI) often causes micturition dysfunction. We recently discovered a low level of spinally-derived dopamine (DA) that regulates recovered bladder and sphincter reflexes in SCI female rats. Considering substantial sexual dimorphic features in the lower urinary tract, it is unknown if the DA-ergic mechanisms act in the male. Histological analysis showed a similar distribution of tyrosine hydroxylase (TH)+ neurons in the lower cord of male rats and the number increased following thoracic SCI. Subsequently, focal electrical stimulation in slices obtained from L6/S1 spinal segments of SCI rats elicited detectable DA release with fast scan cyclic voltammetry. Using bladder cystometrogram and external urethral sphincter (EUS) electromyography in SCI male rats, intravenous (i.v.) administration of SCH 23390, a D1-like receptor (DR1) antagonist, induced significantly increased tonic EUS activity and a trend of increased residual volume, whereas activation of these receptors with SKF 38393 did not influence the reflex. Meanwhile, blocking spinal D2-like receptors (DR2) with remoxipride had no effect but stimulating these receptors with quinpirole elicited EUS bursting to increase voiding volume. Further, intrathecal delivery of SCH 23390 and quinpirole resulted in similar responses to those with i.v. delivery, respectively, which indicates the central action regardless of delivery route. In addition, metabolic cage assays showed that quinpirole increased the voiding frequency and total voiding volume in spontaneous micturition. Collectively, spinal DA-ergic machinery regulates recovered micturition reflex following SCI in male rats; spinal DR1 tonically suppress tonic EUS activity to enable voiding and activation of DR2 facilitates voiding.
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Dopamina/metabolismo , Traumatismos da Medula Espinal/metabolismo , Traumatismos da Medula Espinal/fisiopatologia , Bexiga Urinária/metabolismo , Bexiga Urinária/fisiopatologia , Micção/fisiologia , Animais , Antagonistas de Dopamina/administração & dosagem , Eletromiografia/métodos , Feminino , Masculino , Ratos , Ratos Wistar , Receptores de Dopamina D1/antagonistas & inibidores , Receptores de Dopamina D1/metabolismo , Receptores de Dopamina D2/agonistas , Receptores de Dopamina D2/metabolismo , Traumatismos da Medula Espinal/tratamento farmacológico , Vértebras Torácicas/lesões , Bexiga Urinária/efeitos dos fármacos , Micção/efeitos dos fármacosRESUMO
The proteasome inhibitor bortezomib (BTZ) is a potent first-line anticancer drug for multiple myeloma; nonetheless, it induced peripheral neuropathy. It has been suggested that many cytokines may play a role in mediating neuropathic pain, but the underlying molecular mechanism is not fully understood. Recent studies have shown that neuropathic pain is closely related to the purinergic ligand-gated ion channel 7 receptor (P2X7R), one of the P2X receptors, which is richly expressed in glial cells. P2X7-p38 pathway is correlated with microglia- and satellite glial cell- (SGC-) mediated neuropathic pain. However, the association of P2X7R and p38MAPK in mediating BTZ-induced neuropathic pain remains unclear. In this study, the relationship between P2X7R activation and p38 phosphorylation in the dorsal root ganglion (DRG) and spinal dorsal horn (SDH) in the development and maintenance of BTZ-induced neuropathic pain was elucidated. The results showed that BTZ increased mechanical thresholds in rats, accompanied with upregulation of P2X7R expression and p38MAPK phosphorylation, indicating that P2X7R and p38MAPK are key molecules in the development and maintenance of BTZ-induced neuropathic pain. Inhibiting p38MAPK phosphorylation with SB203580 resulted in downregulation of P2X7R expression levels. Inhibition of P2X7R with Brilliant Blue G (BBG) reversed neuropathic pain might decrease through the expression of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and IL-6 via inhibiting p38MAPK phosphorylation. The P2X7R/p38MAPK signaling pathway in SGCs of DRG and microglia of SDH might be a potential pharmacological target behind this mechanism as an opportunity to relieve BTZ-induced neuropathic pain.
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Bortezomib/efeitos adversos , Neuralgia/induzido quimicamente , Neuralgia/metabolismo , Receptores Purinérgicos P2X7/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Citocinas/genética , Citocinas/metabolismo , Ativação Enzimática , Gânglios Espinais/efeitos dos fármacos , Gânglios Espinais/metabolismo , Hiperalgesia/complicações , Masculino , Neuralgia/complicações , Fosforilação , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos Wistar , Corno Dorsal da Medula Espinal/efeitos dos fármacos , Corno Dorsal da Medula Espinal/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/genéticaRESUMO
Glioma is a brain tumour that is often diagnosed, and temozolomide (TMZ) is a common chemotherapeutic drug used in glioma. Yet, resistance to TMZ is a chief hurdle towards curing the malignancy. The current work explores the pathways and involvement of miR-3116 in the TMZ resistance. miR-3116 and FGFR1 mRNA were quantified by real-time PCR in malignant samples and cell lines. Appropriate assays were designed for apoptosis, viability, the ability to form colonies and reporter assays to study the effects of the miR-3116 or FGFR1. The involvement of PI3K/AKT signalling was assessed using Western blotting. Tumorigenesis was evaluated in an appropriate xenograft mouse model in vivo. This work revealed that the levels of miR-3116 dipped in samples resistant to TMZ, while increased miR-3116 caused an inhibition of the tumour features mentioned above to hence augment TMZ sensitivity. miR-3116 was found to target FGFR1. When FGFR1 was overexpressed, resistance to TMZ was augmented and reversed the sensitivity caused by miR-3116. Our findings further confirmed PI3K/AKT signalling pathway is involved in this action. In conclusion, miR-3116 sensitizes glioma cells to TMZ through FGFR1 downregulation and the PI3K/AKT pathway inactivation. Our results provide a strategy to overcome TMZ resistance in glioma treatment.
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Glioma/tratamento farmacológico , MicroRNAs/genética , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Temozolomida/farmacologia , Animais , Apoptose/genética , Carcinogênese/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Dacarbazina/farmacologia , Resistencia a Medicamentos Antineoplásicos/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glioma/genética , Glioma/patologia , Humanos , Camundongos , Proteína Oncogênica v-akt/genética , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas c-akt/genética , Transdução de Sinais/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Mesenchymal stromal injection is a promising therapy for traumatic brain injury (TBI). The aim of this study was to explore the effects of the HIF-1α/SDF-1/CXCR4 axis on neuron repair in TBI rats through improving the bone marrow-derived mesenchymalstromal cells (BMSCs) migration. TBI rat models were established. The rats were treated with exogenous SDF-1, and then the neuronal apoptosis in TBI rats was measured. BMSCs from rats were collected, and the roles of NF-κB p65 expression in nuclei, overexpression of SDF-1 and HIF-1α, as well as downregulation of CXCR4 in BMSC migration were identified. HIF-1α- and SDF-1- treated BMSCs were transplanted into TBI rats, after which the neuronal apoptosis and activity of the HIF-1α/SDF-1/CXCR4 axis were detected. Consequently, we found SDF-1 elevated the HIF-1α/SDF-1/CXCR4 activity and presented protective roles in TBI rat hippocampal neurons with reduced neuronal apoptosis. SDF-1 promoted BMSC migration in vitro, and co-effects of SDF-1 and HIF-1α showed strong promotion, while CXCR4 inhibition suppressed BMSC migration. BMSC transplantation activated the HIF-1α/SDF-1/CXCR4 axis and reduced neuronal apoptosis in TBI rats. To conclude, our study demonstrated that the HIF-1α/SDF-1/CXCR4 axis could enhance BMSC migration and alleviate neuronal damage and apoptosis in TBI rats. This study provided novel options for TBI therapy.
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Lesões Encefálicas Traumáticas/genética , Quimiocina CXCL12/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Receptores CXCR4/genética , Animais , Apoptose/genética , Lesões Encefálicas Traumáticas/metabolismo , Lesões Encefálicas Traumáticas/patologia , Movimento Celular/genética , Modelos Animais de Doenças , Hipocampo/metabolismo , Hipocampo/patologia , Humanos , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/patologia , Neurônios/metabolismo , Neurônios/patologia , Ratos , Transdução de Sinais/genética , Células Estromais/metabolismo , Células Estromais/patologiaRESUMO
BACKGROUND: Hypertensive cerebral hemorrhage (HCH) is a potentially life-threatening neurological condition with an extremely high morbidity and mortality. In recent years, neuroendoscopy has been used to treat intracerebral hemorrhage (ICH). However, the choice of neuroendoscopic surgery versus craniotomy for patients with intracerebral hemorrhages is controversial. AIM: We conducted this meta-analysis to assess the efficacy of neuroendoscopic surgery compared with craniotomy in patients with supratentorial hypertensive ICH. METHODS: A systematic electronic search was conducted of online electronic databases: PubMed, Embase, and the Cochrane Library updated on December 2017. The meta-analysis only included randomized controlled studies. RESULTS: Three randomized controlled trials met our inclusion criteria. The pooled analysis of death showed that neuroendoscopic surgery decreased the rate of death when compared with craniotomy (RR = 0.58, 95% CI 0.26-1.29; p = .18). The pooled result of complications indicated that neuroendoscopic surgery has a tendency toward lower complications (RR = 0.37, 95% CI 0.28-0.49; p < .001). CONCLUSIONS: Our results suggested that neuroendoscopic surgery has lower complications, but no superior advantages in morbidity rates. Since the advantage of neuroendoscopic surgery has been performed in some area, the continuation of multi-center comparative investigation with craniotomy may be necessary. Moreover, some efforts need to be taken in selecting appropriate patients with different treatments.
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Craniotomia , Hemorragia Intracraniana Hipertensiva/cirurgia , Neuroendoscopia , Complicações Pós-Operatórias , Craniotomia/efeitos adversos , Craniotomia/métodos , Humanos , Neuroendoscopia/efeitos adversos , Neuroendoscopia/métodos , Avaliação de Processos e Resultados em Cuidados de Saúde , Complicações Pós-Operatórias/epidemiologia , Complicações Pós-Operatórias/etiologia , Ensaios Clínicos Controlados Aleatórios como AssuntoRESUMO
Adult skeletal muscle regeneration after injury depends on normal myoblast function. However, the intrinsic mechanisms for the control of myoblast behaviors are not well defined. Herein, we identified Pim1 kinase as a novel positive regulator of myoblast behaviors in vitro and muscle regeneration in vivo. Specifically, knockdown of Pim1 significantly restrains the proliferation and accelerates the apoptosis of myoblasts in vitro, indicating that Pim1 is critical for myoblast survival and amplification. Meanwhile, we found that Pim1 kinase is increased and translocated from cytoplasm into nucleus during myogenic differentiation. By using Pim1 kinase inhibitor, we proved that inhibition of Pim1 activity prevents myoblast differentiation and fusion, suggesting the necessity of Pim1 kinase activity for proper myogenesis. Mechanistic studies demonstrated that Pim1 kinase interacts with myogenic regulator MyoD and controls its transcriptional activity, inducing the expression of muscle-specific genes, which consequently promotes myogenic differentiation. Additionally, in skeletal muscle injury mouse model, deletion of Pim1 hinders the regeneration of muscle fibers and the recovery of muscle strength. Taken together, our study provides a potential target for the manipulation of myoblast behaviors in vitro and the myoblast-based therapeutics of skeletal muscle injury.
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Desenvolvimento Muscular/genética , Músculo Esquelético/fisiologia , Mioblastos/metabolismo , Proteínas Proto-Oncogênicas c-pim-1/metabolismo , Regeneração/genética , Animais , Apoptose/genética , Linhagem Celular , Núcleo Celular/enzimologia , Núcleo Celular/metabolismo , Proliferação de Células/genética , Sobrevivência Celular/ética , Bases de Dados Genéticas , Regulação da Expressão Gênica/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Desenvolvimento Muscular/fisiologia , Músculo Esquelético/enzimologia , Músculo Esquelético/lesões , Músculo Esquelético/metabolismo , Proteína MyoD/metabolismo , Mioblastos/enzimologia , Fosforilação , Proteínas Proto-Oncogênicas c-pim-1/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-pim-1/genética , Regulação para CimaRESUMO
STUDY DESIGN: Animal experiment: a rat model of lumbar disc herniation (LDH) induced painful radiculopathies. OBJECTIVE: To investigate the role and mechanism of AMP-activated protein kinase (AMPK) in dorsal root ganglia (DRG) neurons in LDH-induced painful radiculopathies. SUMMARY OF BACKGROUND DATA: Overactivation of multiple pain signals in DRG neurons triggered by LDH is crucial to the development of radicular pain. AMPK is recognized as a cellular energy sensor, as well as a pain sensation modulator, but its function in LDH-induced pain hypersensitivity remains largely unknown. METHODS: The LDH rat model was established by autologous nucleus pulposus transplantation into the right lumbar 5 (L5) nerve root. At different time points after AMPK agonist metformin (250âmg/kg/d) or mammalian target of rapamycin (mTOR) inhibitor rapamycin (5âmg/kg) intraperitoneal administration, thermal and mechanical sensitivity were evaluated by measuring paw withdrawal latency (PWL) and 50% paw withdrawal thresholds (PWT). The levels of AMPK, mTOR, and p70S6K phosphorylation were determined by Western blot. We also investigated the proportion of p-AMPK positive neurons in the right L5 DRG neurons using immunofluorescence. RESULTS: LDH evoked persistent thermal hyperalgesia and mechanical allodynia on the ipsilateral paw, as indicated by the decreased PWL and 50% PWT. These pain hypersensitive behaviors were accompanied with significant inhibition of AMPK and activation of mTOR in the associated DRG neurons. Pharmacological activation of AMPK in the DRG neurons not only suppressed mTOR/p70S6K signaling, but also alleviated LDH-induced pain hypersensitive behaviors. CONCLUSION: We provide a molecular mechanism for the activation of pain signals based on AMPK-mTOR axis, as well as an intervention strategy by targeting AMPK-mTOR axis in LDH-induced painful radiculopathies. LEVEL OF EVIDENCE: N/A.
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Proteínas Quinases Ativadas por AMP/metabolismo , Gânglios Espinais/metabolismo , Hiperalgesia/metabolismo , Degeneração do Disco Intervertebral/metabolismo , Deslocamento do Disco Intervertebral/metabolismo , Radiculopatia/metabolismo , Proteínas Quinases S6 Ribossômicas 70-kDa/antagonistas & inibidores , Serina-Treonina Quinases TOR/antagonistas & inibidores , Animais , Modelos Animais de Doenças , Gânglios Espinais/enzimologia , Hiperalgesia/enzimologia , Degeneração do Disco Intervertebral/enzimologia , Deslocamento do Disco Intervertebral/enzimologia , Masculino , Metformina/farmacologia , Neurônios/enzimologia , Neurônios/metabolismo , Núcleo Pulposo/enzimologia , Núcleo Pulposo/metabolismo , Dor/enzimologia , Dor/metabolismo , Fosforilação , Radiculopatia/enzimologia , Ratos , Ratos Wistar , Proteínas Quinases S6 Ribossômicas 70-kDa/metabolismo , Transdução de Sinais , Sirolimo/farmacologia , Raízes Nervosas Espinhais/enzimologia , Raízes Nervosas Espinhais/metabolismo , Serina-Treonina Quinases TOR/metabolismoRESUMO
Coactivator-associated arginine methyltransferase 1 (CARM1) is involved in a variety of biological processes in different cell types and disease conditions, including myogenesis. However, the specific function of CARM1 in skeletal muscle wasting under pathologic conditions remains unclear. Here, we identify CARM1 as a novel participant in muscular atrophy. Increases in CARM1 protein levels correlated positively with the loss of muscle mass upon denervation in mice. Notably, the knockdown of CARM1 represses the progression of muscle wasting and the expression of the atrophy-related genes Atrogin-1 and MuRF1 in vivo and in vitro. With respect to the underlying mechanism, we show that CARM1 interacts with and asymmetrically dimethylates FoxO3 (a specific transcription factor that controls atrophy-related gene expression). This methylation modification by CARM1 is required for FoxO3-dependent transcription. Accordingly, a CARM1 methyltransferase inhibitor also restrains the expression of Atrogin-1 and MuRF1 and myotube atrophy. Furthermore, CARM1 knockdown induces a remarkable myofiber autophagic deficit during the atrophy process. Altogether, our study identifies a crucial regulator of skeletal muscle atrophy and suggests that CARM1 is a potential target for the prevention of muscle atrophy.
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Autofagia , Proteína Forkhead Box O3/metabolismo , Fibras Musculares Esqueléticas/metabolismo , Proteína-Arginina N-Metiltransferases/metabolismo , Animais , Linhagem Celular , Dexametasona , Masculino , Metilação , Camundongos Endogâmicos C57BL , Modelos Biológicos , Denervação Muscular , Atrofia Muscular/patologia , Tamanho do Órgão , Ligação ProteicaRESUMO
BACKGROUND: DL-3-n-butylphthalide (NBP) is a drug for treating acute ischemic stroke, and may play a neuroprotective role by acting on multiple active targets. The aim of this study was to predict the target proteins of NBP in mammalian cells. METHODS: The similarity ensemble approach search tool (SEArch), one of the commonly used public bioinformatics tools for target prediction, was employed in the experiment. The molecular docking of NBP to target proteins was performed by using the three-dimensional (3-D) crystal structure, substrate free. The software AutoDock Vina was used for all dockings. The binding targets of NBP were illustrated as 3-D and 2-D diagrams. RESULTS: Firstly, the results showed that NBP bounded to the same binding site on NAD(P)H quinone oxidoreductases (NQO1) as the substrate FAD, leading to competitive inhibition for the catalytic site with -7.2 kcal/mol. This might break the 3-D structure of NQO1 and bring about P53 degradation, resulting in a decrease of p53-mediated apoptosis in ischemic brain cells. Secondly, NBP might exert its therapeutic effect on acute ischemic stroke via modulating indoleamine 2,3-dioxygenase (IDO) bioactivity after associating with it. NBP could alleviate the depression following ischemic stroke by inhibiting IDO. Thirdly, NBP might modulate the function of NADH-ubiquinone oxidoreductase by competitively embedding itself into this complex, further affecting mitochondrial respiration in cerebrovascular diseases as an anti-oxidant agent. CONCLUSIONS: Three potential target proteins of NBP were identified, which may provide a novel aspect for better understanding the protective effects of NBP on the nervous system at the molecular level.
RESUMO
Long non-coding RNAs play a significant role in cancer metastasis. Studies have demonstrated that LncRNA NEAT1 promotes cancer progression. We aimed to explore whether NEAT1 regulated growth and invasion in breast cancer cells. We provided evidence that lncRNA NEAT1 was up-regulated in breast cancer cell lines and tissues. NEAT1 promoted invasion through inducing Epithelial-mesenchymal transition (EMT) and NEAT1 played a role in 5-fluorouracil (5-FU) resistance. In addition, we revealed a reciprocal repression between NEAT1 and miR-211. Furthermore, the EMT-inducer HMGA2 was identified as a down-stream target of miR-211. LncRNA NEAT1 induced EMT and 5-FU resistance through the miR-211/HMGA2 axis. Our findings suggest that lncRNA NEAT1 could be a new diagnostic biomarker and therapy target for breast cancer.
Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Proteína HMGA2/metabolismo , MicroRNAs/genética , RNA Longo não Codificante/genética , Animais , Neoplasias da Mama/diagnóstico , Linhagem Celular Tumoral , Proliferação de Células/genética , Transição Epitelial-Mesenquimal/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos , Pessoa de Meia-Idade , Invasividade Neoplásica , Metástase Neoplásica , Fenótipo , Prognóstico , Regulação para CimaRESUMO
A novel ultra-sensitive fluorescent sensor for monitoring microRNA (miRNA) in living cells was constructed by utilizing a hybridization chain reaction (HCR) as the signal amplification with a carbon nitride nanosheet (CNNS) as a carrier. The Cy5-labeled hairpin DNA could be adsorbed onto the surface of CNNS, resulting in fluorescence quenching of Cy5. When treated with complementary miRNA, the fluorescence was recovered because miRNA could efficiently trigger an HCR, which led to the release of the HCR products from the CNNS. This intracellular HCR strategy can be used for ultra-sensitive monitoring of intracellular miRNA. The main advantages of the proposed method are its simplicity, high sensitivity, high specificity and low toxicity for monitoring low-level biomarkers.
Assuntos
Corantes Fluorescentes/química , MicroRNAs/análise , Nanoestruturas/química , Nitrilas/química , Animais , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , Corantes Fluorescentes/farmacologia , Nitrilas/farmacologia , Hibridização de Ácido Nucleico , Células PC12 , Ratos , Espectrometria de Fluorescência , Relação Estrutura-AtividadeRESUMO
Restoring the contractile function of long-term denervated skeletal muscle (SKM) cells is difficult due to the long period of denervation, which causes a loss of contractility. Although sensory innervation is considered a promising protective approach, its effect is still restricted. In this study, we introduced insulin-like growth factor-1 (IGF-1) as an efficient protective agent and observed that IGF-1 potentiated the effects of sensory protection by preventing denervated muscle atrophy and improving the condition of denervated muscle cells in vivo and in vitro. IGF-1-induced Akt phosphorylation suppressed the mitochondrial outer-membrane protein Mul1 expression, which is a key step on preserving contractile property of sensory innervated SKM cells. Mul1 overexpression interfered with the balance between mitochondrial fusion and fission and was a key node for blocking the effects of IGF-1 that preserved the contractility of sensory-innervated SKM cells. Activation of AMP-activated protein kinase α (AMPKα), a mitochondrial downstream target, could block the effects of IGF-1. These data provide novel evidence that might be applied when searching for new approaches to improve the functional condition of long-term denervated SKM cells by increasing sensory protection using the IGF-1 signalling system to modulate the balance between mitochondrial fusion and fission.