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Abnormal proliferation of pulmonary artery smooth muscle cells (PASMCs) is one of the critical pathological mechanisms of pulmonary hypertension (PH), and therefore is gradually being adopted as an important direction for the treatment of PH. Metallothioneins (MTs) have been reported to be associated with PH, but the underlying mechanisms are not fully understood. Here, we demonstrated that the expression level of metallothionein 3 (MT3) was significantly increased in pulmonary arterioles from PH patients and chronic hypoxia-induced rat and mouse PH models, as well as in hypoxia-treated human PASMCs. Knockdown of MT3 significantly inhibited the proliferation of human PASMCs by arresting the cell cycle in the G1 phase, while overexpression of MT3 had the opposite effect. Mechanistically, we found that MT3 increased the intracellular zinc (Zn2+) concentration to enhance the transcriptional activity of metal-regulated transcription factor 1 (MTF1), which promoted the expression of autophagy-related gene 5 (ATG5), facilitating autophagosome formation. More importantly, MT3-induced autophagy and proliferation of human PASMCs were largely prevented by knockdown of MTF1 and ATG5. Therefore, in this study, we identified MT3-Zinc-MTF1-ATG5 as a novel pathway that affects PASMC proliferation by regulating autophagosome formation, suggesting that MT3 may be a novel target for the treatment of PH.
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Proliferação de Células , Metalotioneína 3 , Miócitos de Músculo Liso , Artéria Pulmonar , Zinco , Artéria Pulmonar/citologia , Artéria Pulmonar/metabolismo , Animais , Humanos , Zinco/metabolismo , Camundongos , Ratos , Miócitos de Músculo Liso/metabolismo , Masculino , Autofagossomos/metabolismo , Proteína 5 Relacionada à Autofagia/metabolismo , Proteína 5 Relacionada à Autofagia/genética , Ratos Sprague-Dawley , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Autofagia , Hipertensão Pulmonar/metabolismo , Camundongos Endogâmicos C57BL , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Fator MTF-1 de Transcrição , Metalotioneína/metabolismo , Metalotioneína/genéticaRESUMO
Some previous researches have demonstrated that appropriate mechanical stimulation can enhance bone formation. However, most studies have employed the strain energy density (SED) method for predicting bone remodeling, with only a few considering the potential impact of wall fluid shear stress (FSS) on this process. To bridge this gap, the current study compared the prediction of bone formation and resorption via SED and wall FSS by using fluid-solid coupling numerical simulation. Specifically, 8-week-old female Sprague-Dawley rats were subjected to stretching of the eighth caudal vertebra using a custom-made device. Based on micro-computed tomography images, a three-dimensional model integrating fluid-solid coupling was created to represent compact bone, cancellous bone, and bone marrow. The animals were grouped into control, 1 Hz, and 10 Hz categories, wherein a tensile displacement load of 1000 µÎµ was applied to the loading end. The results revealed that SED values tended to increase with elevated porosity, whereas wall FSS values decreased it. Notably, wall FSS demonstrated the higher predictive accuracy for cancellous bone resorption than SED. These findings support the notion that fluid flow within cancellous bone spaces can significantly impact bone resorption. Therefore, the findings of this study contribute to a more comprehensive understanding of the role of wall FSS in bone remodeling, providing a theoretical support for the dynamic evolution of bone structures under mechanical stimulation.
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BACKGROUND: Hyperproliferation of pulmonary arterial smooth muscle cells (PASMCs) and consequent pulmonary vascular remodeling are the crucial pathological features of pulmonary hypertension (PH). Protein methylation has been shown to be critically involved in PASMC proliferation and PH, but the underlying mechanism remains largely unknown. METHODS: PH animal models were generated by treating mice/rats with chronic hypoxia for 4 weeks. SMYD2-vTg mice (vascular smooth muscle cell-specific suppressor of variegation, enhancer of zeste, trithorax and myeloid Nervy DEAF-1 (deformed epidural auto-regulatory factor-1) domain-containing protein 2 transgenic) or wild-type rats and mice treated with LLY-507 (3-cyano-5-{2-[4-[2-(3-methylindol-1-yl)ethyl]piperazin-1-yl]-phenyl}-N-[(3-pyrrolidin-1-yl)propyl]benzamide) were used to investigate the function of SMYD2 (suppressor of variegation, enhancer of zeste, trithorax and myeloid Nervy DEAF-1 domain-containing protein 2) on PH development in vivo. Primary cultured rat PASMCs with SMYD2 knockdown or overexpression were used to explore the effects of SMYD2 on proliferation and to decipher the underlying mechanism. RESULTS: We demonstrated that the expression of the lysine methyltransferase SMYD2 was upregulated in the smooth muscle cells of pulmonary arteries from patients with PH and hypoxia-exposed rats/mice and in the cytoplasm of hypoxia-induced rat PASMCs. More importantly, targeted inhibition of SMYD2 by LLY-507 significantly attenuated hypoxia-induced pulmonary vascular remodeling and PH development in both male and female rats in vivo and reduced rat PASMC hyperproliferation in vitro. In contrast, SMYD2-vTg mice exhibited more severe PH phenotypes and related pathological changes than nontransgenic mice after 4 weeks of chronic hypoxia treatment. Furthermore, SMYD2 overexpression promoted, while SMYD2 knockdown suppressed, the proliferation of rat PASMCs by affecting the cell cycle checkpoint between S and G2 phases. Mechanistically, we revealed that SMYD2 directly interacted with and monomethylated PPARγ (peroxisome proliferator-activated receptor gamma) to inhibit the nuclear translocation and transcriptional activity of PPARγ, which further promoted mitophagy to facilitate PASMC proliferation and PH development. Furthermore, rosiglitazone, a PPARγ agonist, largely abolished the detrimental effects of SMYD2 overexpression on PASMC proliferation and PH. CONCLUSIONS: Our results demonstrated that SMYD2 monomethylates nonhistone PPARγ and inhibits its nuclear translocation and activation to accelerate PASMC proliferation and PH by triggering mitophagy, indicating that targeting SMYD2 or activating PPARγ are potential strategies for the prevention of PH.
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Histona-Lisina N-Metiltransferase , Hipertensão Pulmonar , Hipóxia , Mitofagia , Músculo Liso Vascular , Miócitos de Músculo Liso , PPAR gama , Artéria Pulmonar , Ratos Sprague-Dawley , Animais , Humanos , Masculino , Camundongos , Ratos , Proliferação de Células , Células Cultivadas , Histona-Lisina N-Metiltransferase/metabolismo , Histona-Lisina N-Metiltransferase/genética , Hipertensão Pulmonar/metabolismo , Hipertensão Pulmonar/etiologia , Hipertensão Pulmonar/patologia , Hipertensão Pulmonar/genética , Hipóxia/complicações , Hipóxia/metabolismo , Metilação , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , PPAR gama/metabolismo , Artéria Pulmonar/patologia , Artéria Pulmonar/metabolismo , Remodelação VascularRESUMO
Uphill walking is a common task encountered in daily life, with steeper inclines potentially imposing greater biomechanical and neuromuscular demands on the human body. The heel-to-toe drop (HTD) in footwear may influence the biomechanical and neuromuscular pattern of uphill walking; but the impact remains unclear. Adjustments in HTD can modulate biomechanical and neuromuscular patterns, mitigating the demands and optimizing the body's response to different inclinations. We hypothesize that adjustments in HTD can modulate biomechanical and neuromuscular patterns, mitigating the demands and optimizing the body's response to different inclinations. Nineteen healthy men walked on an adjustable slope walkway, with varied inclinations (6°, 12°, 20°) and HTD shoes (10mm, 25mm, 40 mm), while the marker positions, ground reaction forces and electromyography data were collected. Our study reveals that gait temporo-spatial parameters are predominantly affected by inclination over HTD. Inclination has a more pronounced effect on kinematic variables, while both inclination and HTD significantly modulate kinetic and muscle synergy parameters. This study demonstrates that an increase in the inclination leads to changes in biomechanical and neuromuscular responses during uphill walking and the adjustment of HTD can modulate these responses during uphill walking. However, the present study suggests that an increased HTD may lead to elevated loads on the knee joint and these adverse effects need more attention.
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Previous studies have shown that ferroptosis of vascular smooth muscle cells (VSMCs) is involved in the development of aortic dissection (AD) and that histone methylation regulates this process. SP2509 acts as a specific inhibitor of lysine-specific demethylase 1 (LSD1), which governs a variety of biological processes. However, the effect of SP2509 on VSMC ferroptosis and AD remains to be elucidated. This aim of this study was to investigate the role and underlying mechanism of SP2509-mediated histone methylation on VSMC ferroptosis. Here, a mouse model of AD was established, and significantly reduced levels of H3K4me1 and H3K4me2 (target of SP2509) were found in the aortas of AD mice. In VSMCs, SP2509 treatment led to a dose-dependent increase in H3K4me2 levels. Furthermore, we found that SP2509 provided equivalent protection to ferrostatin-1 against VSMC ferroptosis, as evidenced by increased cell viability, decreased cell death and lipid peroxidation. RNA-sequencing analysis and subsequent experiments revealed that SP2509 counteracted cystine deficiency-induced response to inflammation and oxidative stress. More importantly, we demonstrated that SP2509 inhibited the expression of TFR and ferritin to reduce intracellular iron levels, thereby effectively blocking the process of ferroptosis. Therefore, our findings indicate that SP2509 protects VSMCs from multiple stimulus-induced ferroptosis by reducing intracellular iron levels, thereby preventing lipid peroxidation and cell death. These findings suggest that SP2509 may be a promising drug to alleviate AD by reducing iron deposition and VSMC ferroptosis.
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Ferroptose , Ferro , Músculo Liso Vascular , Miócitos de Músculo Liso , Ferroptose/efeitos dos fármacos , Animais , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/patologia , Camundongos , Ferro/metabolismo , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/patologia , Estresse Oxidativo/efeitos dos fármacos , Humanos , Modelos Animais de Doenças , Peroxidação de Lipídeos/efeitos dos fármacos , Fenilenodiaminas/farmacologia , Masculino , Sobrevivência Celular/efeitos dos fármacos , Histonas/metabolismo , Histonas/genética , Histona Desmetilases/metabolismo , Histona Desmetilases/genética , Camundongos Endogâmicos C57BL , CicloexilaminasRESUMO
Coronary restenosis is an important cause of poor long-term prognosis in patients with coronary heart disease. Here, we show that lysine methyltransferase SMYD2 expression in the nucleus is significantly elevated in serum- and PDGF-BB-induced vascular smooth muscle cells (VSMCs), and in tissues of carotid artery injury-induced neointimal hyperplasia. Smyd2 overexpression in VSMCs (Smyd2-vTg) facilitates, but treatment with its specific inhibitor LLY-507 or SMYD2 knockdown significantly inhibits VSMC phenotypic switching and carotid artery injury-induced neointima formation in mice. Transcriptome sequencing revealed that SMYD2 knockdown represses the expression of serum response factor (SRF) target genes and that SRF overexpression largely reverses the inhibitory effect of SMYD2 knockdown on VSMC proliferation. HDAC3 directly interacts with and deacetylates SRF, which enhances SRF transcriptional activity in VSMCs. Moreover, SMYD2 promotes HDAC3 expression via tri-methylation of H3K36 at its promoter. RGFP966, a specific inhibitor of HDAC3, not only counteracts the pro-proliferation effect of SMYD2 overexpression on VSMCs, but also inhibits carotid artery injury-induced neointima formation in mice. HDAC3 partially abolishes the inhibitory effect of SMYD2 knockdown on VSMC proliferation in a deacetylase activity-dependent manner. Our results reveal that the SMYD2-HDAC3-SRF axis constitutes a novel and critical epigenetic mechanism that regulates VSMC phenotypic switching and neointimal hyperplasia.
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Trabeculae bone undergoes directional growth along the applied force under physiological loading. The growth of bone structure relies on the coordinated interplay among osteocytes, osteoblasts, and osteoclasts. Under normal circumstances, bone remodeling maintains a state of equilibrium. Excessive bone formation can lead to osteosclerosis, while excessive bone resorption can result in osteoporosis and osteonecrosis. The investigation of the structural characteristics of trabeculae and the mechanotransduction between bone cells plays a vital role in the treatment of bone-related diseases. In this study, a fluid-solid coupling model of the entire vertebral bone was established based on micro-CT images obtained from rat tail vertebrae subjected to tensile loading experiments. The flow characteristics of bone marrow and the mechanical response of osteocytes in different regions under physiological loading were investigated. The results revealed a U-shaped distribution of wall fluid shear stress (FSS) along the longitudinal axis in trabecular bone, with higher FSS regions exhibiting greater mechanical stimulation on osteocytes. These findings elucidate a positive correlation between the mechanical microenvironment among osteocytes, osteoblasts, and osteoclasts, providing potential strategies for the prevention and treatment of bone diseases.
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Cross-country sit-skiers use double poling (DP) technique to drive the slide. The aim of this study is to analyze how poling camber angle affect the capacity of power output and biomechanical parameters of the DP process. Twenty-four non-disabled college students (24.67 ± 1.46 years old) were recruited to perform three successive 30-s maximal effort tests with different poling camber angles of 0°, 15°, 24° and 30° using a sit-skiing ergometer. The biomechanical parameters, output power and muscle activation of the subjects were analyzed. The results showed that DP output power increased with the increase of poling camber angle at 15° (597.78 ± 150.31 J), 24° (610.94 ± 158.96 J, P = 0.011) and 30° (629.10 ± 168.78 J, P < 0.001) compared with 0° (590.65 ± 148.95 J). However, effective output power decreased with the increase of camber angle. Poling with camber angle of 24° had the shortest cycle time 1.53 ± 0.17 s, compared with other abduction angle (0°, 1.57 ± 0.19 s, 15°, 1.55 ± 0.16 s, and 30°, 1.56 ± 0.19 s). Compared with 0° (1.02 ± 0.14 m), the cycle distance significantly increased at poling camber angles of 24° (1.07 ± 0.12 m, P = 0.029) and 30° (1.11 ± 0.13 m, P < 0.001). With the increase of poling camber angle, the shoulder and elbow joint range of motions and joint moments were significantly increased. This study found that poling with shoulder abducted increased the output power but decreased the efficiency. By analyzing the poling angle and poling force, we find that the optimal poling camber angle may depend on the terrain or the skiing speed. These results may guide the competition techniques and tactics in the matches, and may further influence the strength-training programs of cross-country sit-skiing athletes.
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Desempenho Atlético , Esqui , Humanos , Adulto Jovem , Adulto , Teste de Esforço , Fenômenos Biomecânicos/fisiologia , Esqui/fisiologia , Ergometria , Músculo Esquelético/fisiologia , Consumo de Oxigênio/fisiologia , Desempenho Atlético/fisiologiaRESUMO
Introduction: Musculoskeletal simulation has been widely used to analyze athletes' movements in various competitive sports, but never in ski jumping. Aerodynamic forces during ski jumping take-off have been difficult to account for in dynamic simulation. The purpose of this study was to establish an efficient approach of musculoskeletal simulation of ski jumping take-off considering aerodynamic forces and to analyze the muscle function and activity. Methods: Camera-based marker-less motion capture was implemented to measure the take-off kinematics of eight professional jumpers. A suitable full-body musculoskeletal model was constructed for the simulation. A method based on inverse dynamics iteration was developed and validated to estimate the take-off ground reaction force. The aerodynamic forces, which were calculated based on body kinematics and computational fluid dynamics simulations, were exerted on the musculoskeletal model as external forces. The activation and joint torque contributions of lower extremity muscles were calculated through static optimization. Results: The estimated take-off ground reaction forces show similar trend with the results from past studies. Although overall inconsistencies between simulated muscle activation and EMG from previous studies were observed, it is worth noting that the activation of the tibialis anterior, gluteus maximus, and long head of the biceps femoris was similar to specific EMG results. Among lower extremity extensors, soleus, vastus lateralis, biceps femoris long head, gluteus maximus, and semimembranosus showed high levels of activation and joint extension torque contribution. Discussion: Results of this study advanced the understanding of muscle action during ski jumping take-off. The simulation approach we developed may help guide the physical training of jumpers for improved take-off performance and can also be extended to other phases of ski jumping.
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BACKGROUND: Aberrant proliferation of vascular smooth muscle cells (VSMCs) is the cause of neointima formation followed by vascular injury. Autophagy is involved in this pathological process, but its function is controversial. Recently, we found that methyltransferase like 3 (METTL3) inhibited VSMC proliferation by activating autophagosome formation. Moreover, we also demonstrated that METTL3 reduced the levels of phosphorylated mammalian target of rapamycin (p-mTOR) and cyclin dependent kinase 1 (p-CDK1/CDC2), which were critical for autophagy and proliferation regulation. However, whether mTOR and CDK1 mediated the function of METTL3 on autophagy and proliferation in VSMCs remains unknown. RESULTS: We showed that the activator of mTOR, MHY1485 largely reversed the effects of METTL3 overexpression on VSMC autophagy and proliferation. Rapamycin, the inhibitor of mTOR, obviously nullified the pro-proliferation effects of METTL3 knockdown by activating autophagy in VSMCs. Unexpectedly, mTOR did not contribute to the impacts of METTL3 on migration and phenotypic switching of VSMCs. On the other hand, by knockdown of CDK1 in VSMC with METTL3 deficiency, we demonstrated that CDK1 was involved in METTL3-regulated proliferation of VSMCs, but this effect was not mediated by autophagy. CONCLUSIONS: We concluded that mTOR but not CDK1 mediated the role of METTL3 on VSMC proliferation and autophagy.
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Our previous studies have demonstrated that macrophages (RAW264.7) have a special ability for sensing the gradient of fluid shear stress (FSS) and migrate toward the low-FSS region. However, the molecular mechanism regulating this phenomenon is still unclear. In this study, we examined the transcriptome genes in RAW264.7 cells, MC3T3-E1 osteoblasts, mesenchymal stem cells, canine renal epithelial cells, and periodontal ligament cells. The expression levels of genes related to cell migration, force transfer, and force sensitivity in the Ca2+ signaling pathway were analyzed. We observed that the transient receptor potential cation channel type 2 (TRPV2) was highly expressed in RAW264.7 cells. Furthermore, we used lentiviral transfection to knockdown TRPV2 expression in RAW264.7 cells and studied the effect of TRPV2 on the migration of RAW264.7 cells under a gradient FSS field. The results showed that compared with normal cells, TRPV2-knockdown cells had impaired ability for sensing FSS gradient to migrate toward the low-FSS region and lower intracellular calcium response to FSS stimulation. This study may reveal the molecular mechanism of regulating the directional migration of macrophages under a gradient FSS field.
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Osteoblastos , Transdução de Sinais , Animais , Cães , Camundongos , Linhagem Celular , Macrófagos , Osteoblastos/metabolismo , Células RAW 264.7 , Estresse MecânicoRESUMO
BACKGROUND: E1A-associated 300-kDa protein (P300), an endogenous histone acetyltransferase, contributes to modifications of the chromatin landscape of genes involved in multiple cardiovascular diseases. Ferroptosis of vascular smooth muscle cells (VSMCs) is a novel pathological mechanism of aortic dissection. However, whether P300 regulates VSMC ferroptosis remains unknown. METHODS: Cystine deprivation (CD) and imidazole ketone erastin (IKE) were used to induce VSMC ferroptosis. Two different knockdown plasmids targeting P300 and A-485 (a specific inhibitor of P300) were used to investigate the function of P300 in the ferroptosis of human aortic smooth muscle cells (HASMCs). Cell counting kit-8, lactate dehydrogenase and flow cytometry with propidium iodide staining were performed to assess the cell viability and death under the treatment of CD and IKE. BODIPY-C11 assay, immunofluorescence staining of 4-hydroxynonenal and malondialdehyde assay were conducted to detect the level of lipid peroxidation. Furthermore, co-immunoprecipitation was utilized to explore the interaction between P300 and HIF-1α, HIF-1α and P53. RESULTS: Compared with normal control, the protein level of P300 was significantly decreased in HASMCs treated with CD and IKE, which was largely nullified by the ferroptosis inhibitor ferrostatin-1 but not by the autophagy inhibitor or apoptosis inhibitor. Knockdown of P300 by short-hairpin RNA or inhibition of P300 activity by A-485 promoted CD- and IKE-induced HASMC ferroptosis, as evidenced by a reduction in cell viability and aggravation of lipid peroxidation of HASMCs. Furthermore, we found that hypoxia-inducible factor-1α (HIF-1α)/heme oxygenase 1 (HMOX1) pathway was responsible for the impacts of P300 on ferroptosis of HASMCs. The results of co-immunoprecipitation demonstrated that P300 and P53 competitively bound HIF-1α to regulate the expression of HMOX1. Under normal conditions, P300 interacted with HIF-1α to inhibit HMOX1 expression, while reduced expression of P300 induced by ferroptosis inducers would favor HIF-1α binding to P53 to trigger HMOX1 overexpression. Furthermore, the aggravated effects of P300 knockdown on HASMC ferroptosis were largely nullified by HIF-1α knockdown or the HIF-1α inhibitor BAY87-2243. CONCLUSION: Thus, our results revealed that P300 deficiency or inactivation facilitated CD- and IKE-induced VSMC ferroptosis by activating the HIF-1α/HMOX1 axis, which may contribute to the development of diseases related to VSMC ferroptosis.
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Ferroptose , Músculo Liso Vascular , Humanos , Heme Oxigenase-1/genética , Heme Oxigenase-1/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismoRESUMO
The behavior of vascular smooth muscle cells (VSMCs) contributes to the formation of neointima. We previously found that EHMT2 suppressed autophagy activation in VSMCs. BRD4770, an inhibitor of EHMT2/G9a, plays a critical role in several kinds of cancers. However, whether and how BRD4770 regulates the behavior of VSMCs remain unknown. In this study, we evaluate the cellular effect of BRD4770 on VSMCs by series of experiments in vivo and ex vivo. We demonstrated that BRD4770 inhibited VSMCs' growth by blockage in G2/M phase in VSMCs. Moreover, our results demonstrated that the inhibition of proliferation was independent on autophagy or EHMT2 suppression which we previous reported. Mechanistically, BRD4770 exhibited an off-target effect from EHMT2 and our further study reveal that the proliferation inhibitory effect by BRD4770 was associated with suppressing on SUV39H2/KTM1B. In vivo, BRD4770 was also verified to rescue VIH. Thus, BRD4770 function as a crucial negative regulator of VSMC proliferation via SUV39H2 and G2/M cell cycle arrest and BRD4770 could be a molecule for the therapy of vascular restenosis.
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Músculo Liso Vascular , Neointima , Humanos , Neointima/metabolismo , Proliferação de Células , Movimento Celular , Células Cultivadas , Histona-Lisina N-MetiltransferaseRESUMO
BACKGROUND: Under external loading, the fluid shear stress (FSS) in the porous structures of bones, such as trabecular or lacunar-canalicular cavity, can influence the biological response of bone cells. However, few studies have considered both cavities. The present study investigated the characteristics of fluid flow at different scales in cancellous bone in rat femurs, as well as the effects of osteoporosis and loading frequency. METHODS: Sprague Dawley rats (3 months old) were divided into normal and osteoporotic groups. A multiscale 3D fluid-solid coupling finite element model considering trabecular system and lacunar-canalicular system was established. Cyclic displacement loadings with frequencies of 1, 2, and 4 Hz were applied. FINDINGS: Results showed that the wall FSS around the adhesion complexes of osteocyte on the canaliculi was higher than that on the osteocyte body. Under the same loading conditions, the wall FSS of the osteoporotic group was smaller than that of the normal group. The fluid velocity and FSS in trabecular pores exhibited a linear relationship with loading frequency. Similarly, the FSS around osteocytes also showed the loading frequency-dependent phenomenon. INTERPRETATION: The high cadence in movement can effectively increase the FSS level on osteocytes for osteoporotic bone, i.e., expand the space within the bone with physiological load. This study might help in understanding the process of bone remodeling under cyclic loading and provide the fundamental data for the development of strategies for osteoporosis treatment.
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Remodelação Óssea , Osso Esponjoso , Líquido Extracelular , Análise Numérica Assistida por Computador , Osteócitos , Osteoporose , Ratos Sprague-Dawley , Resistência ao Cisalhamento , Estresse Mecânico , Osteoporose/fisiopatologia , Osso Esponjoso/fisiologia , Osteócitos/fisiologia , Feminino , Animais , Líquido Extracelular/fisiologia , Imageamento Tridimensional , RatosRESUMO
OBJECTIVE: This study aimed to investigate the changes in oocytes at the transcriptome level after applying continuous microvibrational mechanical stimulation to human immature oocytes during in vitro maturation. METHODS: The discarded germinal-vesicle stage (GV) oocytes with no fertilization value after oocytes retrieval in assisted reproduction cycles were collected. Part of them was stimulated with vibration (n = 6) at 10 Hz for 24 h after obtaining informed consent; the other was cultured in static condition (n = 6). Single-cell transcriptome sequencing was used to detect the differences in oocyte transcriptome compared with the static culture group. RESULTS: The applied 10-Hz continuous microvibrational stimulation altered the expression of 352 genes compared with the static culture. Gene Ontology (GO) analysis suggested that the altered genes were mainly enriched with 31 biological processes. The mechanical stimulation upregulated 155 of these genes and downregulated 197 genes. Among them, the genes related to mechanical signaling, such as protein localization to intercellular adhesion (DSP and DLG-5) and cytoskeleton (DSP, FGD6, DNAJC7, KRT16, KLHL1, HSPB1, MAP2K6), were detected. DLG-5, which was related to protein localization to intercellular adhesion, was selected for immunofluorescence experiments based on the transcriptome sequencing results. The protein expression of DLG-5 in the microvibration-stimulated oocytes was higher than that in the static culture oocytes. CONCLUSIONS: Mechanical stimulation affects the transcriptome during oocyte maturation, causing the express changes in intercellular adhesion and cytoskeleton-related genes. We speculate that the mechanical signal may be transmitted to the cell through DLG-5 protein and cytoskeleton-related protein to regulate cellular activities.
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Técnicas de Maturação in Vitro de Oócitos , Transcriptoma , Humanos , Técnicas de Maturação in Vitro de Oócitos/métodos , Transcriptoma/genética , Oócitos/metabolismo , Oogênese/genética , Núcleo Celular , Proteínas de Choque Térmico/genética , Chaperonas Moleculares/genética , Proteínas dos Microfilamentos/genética , Proteínas dos Microfilamentos/metabolismoRESUMO
Wearable robots have been growing exponentially during the past years and it is crucial to quantify the performance effectiveness and to convert them into practical benchmarks. Although there exist some common metrics such as metabolic cost, many other characteristics still needs to be presented and demonstrated. In this study, we developed an integrated evaluation (IE) approach of wearable exoskeletons of lower limb focusing on human performance augmentation. We proposed a novel classification of trial tasks closely related to exoskeleton functions, which were divided into three categories, namely, basic trial at the preliminary phase, semi-reality trial at the intermediate phase, and reality trial at the advanced phase. In the present study, the IE approach has been exercised with a subject who wore an active power-assisted knee (APAK) exoskeleton with three types of trial tasks, including walking on a treadmill at a certain angle, walking up and down on three-step stairs, and ascending in 11-storey stairs. Three wearable conditions were carried out in each trial task, i.e. with unpowered exoskeleton, with powered exoskeleton, and without the exoskeleton. Nine performance indicators (PIs) for evaluating performance effectiveness were adopted basing on three aspects of goal-level, task-based kinematics, and human-robot interactions. Results indicated that compared with other conditions, the powered APAK exoskeleton make generally lesser heart rate (HR), Metabolic equivalent (METs), biceps femoris (BF) and rectus femoris (RF) muscles activation of the subject at the preliminary phase and intermediate phase, however, with minimal performance augmentation at advanced phase, suggesting that the APAK exoskeleton is not suitable for marketing and should be further improved. In the future, continuous iterative optimization for the IE approach may help the robot community to attain a comprehensive benchmarking methodology for robot-assisted locomotion more efficiently.
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Exoesqueleto Energizado , Dispositivos Eletrônicos Vestíveis , Humanos , Extremidade Inferior/fisiologia , Caminhada/fisiologia , Joelho , Fenômenos Biomecânicos/fisiologia , Marcha/fisiologiaRESUMO
The subacromial impingement syndrome is a high-incidence injury for cross-country sit-skiing skier, which is often accompanied by muscle imbalance. However, at present, no musculoskeletal model has been identified for this sport. Thus, this research aimed to establish a bilateral upper extremity trunk (BUET) musculoskeletal model suitable for cross-country sit-skiing based on OpenSim software and verify the function of the model. By splicing three existing OpenSim models, an upper limb model with 17 segments, 35 degrees of freedom, and 472 musculotendon actuators was established. The clavicle and scapula were modeled as individual bodies and then connected to the torso through a three-degrees-of-freedom rotational joint and to the clavicle through a weld joint, respectively. The five lumbar vertebrae were established separately and coupled into a three-degree-of-freedom joint. Kinematics, kinetic, and EMG signal data of five 15-s maximal effort interval tests were obtained by using seven cameras, ergometers, and surface EMG synchronous collection. Based on the resulting rotator cuff muscle geometry of the model, simulated muscle activation patterns were comparable to experimental data, and muscle-driven ability was proven. The model will be available online ( https://simtk.org/projects/bit ) for researchers to study the muscle activation of shoulder joint movement.
Assuntos
Esqui , Esqui/lesões , Esqui/fisiologia , Extremidade Superior/fisiologia , Ergometria , Eletromiografia , Tronco/fisiologia , Fenômenos Biomecânicos/fisiologia , Músculo Esquelético/fisiologiaRESUMO
Prevention of neointima formation is the key to improving long-term outcomes after stenting or coronary artery bypass grafting. RNA N6 -methyladenosine (m6 A) methylation has been reported to be involved in the development of various cardiovascular diseases, but whether it has a regulatory effect on neointima formation is unknown. Herein, we revealed that methyltransferase-like 3 (METTL3), the major methyltransferase of m6 A methylation, was downregulated during vascular smooth muscle cell (VSMC) proliferation and neointima formation. Knockdown of METTL3 facilitated, while overexpression of METTL3 suppressed the proliferation of human aortic smooth muscle cells (HASMCs) by arresting HASMCs at G2/M checkpoint and the phosphorylation of CDC2 (p-CDC2) was inactivated by METTL3. On the other hand, the migration and synthetic phenotype of HASMCs were enhanced by METTL3 knockdown, but inhibited by METTL3 overexpression. The protein levels of matrix metalloproteinase 2 (MMP2), MMP7 and MMP9 were reduced, while the expression level of tissue inhibitor of metalloproteinase 3 was increased in HASMCs with METTL3 overexpression. Moreover, METTL3 promoted the autophagosome formation by upregulating the expression of ATG5 (autophagy-related 5) and ATG7. Knockdown of either ATG5 or ATG7 largely reversed the regulatory effects of METTL3 overexpression on phenotypic switching of HASMCs, as evidenced by increased proliferation and migration, and predisposed to synthetic phenotype. These results indicate that METTL3 inhibits the phenotypic switching of VSMCs by positively regulating ATG5-mediated and ATG7-mediated autophagosome formation. Thus, enhancing the level of RNA m6 A or the formation of autophagosomes is the promising strategy to delay neointima formation.
Assuntos
Autofagossomos , Metiltransferases , Músculo Liso Vascular , Humanos , Movimento Celular , Proliferação de Células , Células Cultivadas , Metaloproteinase 2 da Matriz/metabolismo , Metiltransferases/genética , Metiltransferases/metabolismo , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Neointima/metabolismo , Fenótipo , RNA/metabolismoRESUMO
Objective: This research aims to study the effects of continuous microvibration stimulation on the parthenogenetic development of human germinal vesicle oocytes. Methods: Ninety-five discarded germinal vesicle oocytes from intracytoplasmic sperm injection treatment (ICSI) cycles performed at Amcare Women's & Children's Hospital between January and December 2021 were used for conventional static culture as well as 10 Hz microvibration culture. We investigated the differences between the two groups in terms of oocyte maturation rate, parthenogenetic activation rate, and parthenogenetic blastocyst formation rate. Results: The static culture and 10 Hz microvibration culture of 95 oocytes showed that the parthenogenetic blastocyst formation rate in the microvibration culture group was significantly higher than those in the traditional static culture group. Conclusion: A continuous microvibration stimulation can significantly improve the parthenogenetic developmental potential of human immature oocytes.
Assuntos
Oócitos , Sêmen , Criança , Humanos , Masculino , Feminino , Injeções de Esperma Intracitoplásmicas , Desenvolvimento EmbrionárioRESUMO
A variety of programmed cell death types have been shown to participate in the loss of smooth muscle cells (SMCs) during the development of aortic dissection (AD), but it is still largely unclear whether ferroptosis is involved in the development of AD. In the present study, we found that the expression of key ferroptosis regulatory proteins, solute carrier family 7 member 11 (SLC7A11), ferroptosis suppressor protein 1 (FSP1) and glutathione peroxidase 4 (GPX4) were downregulated in aortas of Stanford type A AD (TAAD) patients, and liproxstatin-1, a specific inhibitor of ferroptosis, obviously abolished the ß-aminopropionitrile (BAPN)-induced development and rupture of AD in mice. Furthermore, the expression of methyltransferase-like 3 (METTL3), a major methyltransferase of RNA m6A, was remarkably upregulated in the aortas of TAAD patients, and the protein levels of METTL3 were negatively correlated with SLC7A11 and FSP1 levels in human aortas. Overexpression of METTL3 in human aortic SMCs (HASMCs) inhibited, while METTL3 knockdown promoted SLC7A11 and FSP1 expression. More importantly, overexpression of METTL3 facilitated imidazole ketone erastin- and cystine deprivation-induced ferroptosis, while knockdown of METTL3 repressed ferroptosis of HASMCs. Overexpression of either SLC7A11 or FSP1 largely abrogated the effect of METTL3 on HASMC ferroptosis. Therefore, we have revealed that ferroptosis is a critical cause of AD in both humans and mice and that METTL3 promotes ferroptosis of HASMCs by inhibiting the expression of SLC7A11 and FSP1. Thus, targeting ferroptosis or m6A RNA methylation is a potential novel strategy for the treatment of AD.