RESUMO
BACKGROUND: Exposure to ionizing radiation has been linked to cardiovascular diseases. However, the impact of moderate doses of radiation on abdominal aortic aneurysm (AAA) remains unknown. METHODS: Angiotensin II-infused Apoe-/- mice were irradiated (acute, 1 Gray) either 3 days before (Day-3) or 1 day after (Day+1) pomp implantation. Isolated primary aortic vascular smooth muscle cells (VSMCs) were irradiated (acute 1 Gray) for mechanistic studies and functional testing in vitro. RESULTS: Day-3 and Day+1 irradiation resulted in a significant reduction in aorta dilation (Control: 1.39+/-0.12; Day-3: 1.12+/-0.11; Day+1: 1.15+/-0.08 mm, P<0.001) and AAA incidence (Control: 81.0%; Day-3: 33.3%, Day+1: 53.3%) compared to the non-irradiated group. Day-3 and Day+1 irradiation led to an increase in collagen content in the adventitia (Thickness control: 23.64+/-2.9; Day-3: 54.39+/-15.5; Day+1 37.55+/-10.8 mm, P = 0.006). However, the underlying protective mechanisms were different between Day-3 and Day+1 groups. Irradiation before Angiotensin II (AngII) infusion mainly modulated vascular smooth muscle cell (VSMC) phenotype with a decrease in contractile profile and enhanced proliferative and migratory activity. Irradiation after AngII infusion led to an increase in macrophage content with a local anti-inflammatory phenotype characterized by the upregulation of M2-like gene and IL-10 expression. CONCLUSION: Moderate doses of ionizing radiation mitigate AAA either through VSCM phenotype or inflammation modulation, depending on the time of irradiation.
Assuntos
Angiotensina II , Aneurisma da Aorta Abdominal , Músculo Liso Vascular , Miócitos de Músculo Liso , Radiação Ionizante , Animais , Aneurisma da Aorta Abdominal/patologia , Aneurisma da Aorta Abdominal/metabolismo , Aneurisma da Aorta Abdominal/etiologia , Camundongos , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/efeitos da radiação , Músculo Liso Vascular/patologia , Angiotensina II/farmacologia , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/efeitos da radiação , Miócitos de Músculo Liso/patologia , Masculino , Modelos Animais de Doenças , Interleucina-10/metabolismo , Interleucina-10/genética , Colágeno/metabolismo , Proliferação de Células/efeitos da radiaçãoRESUMO
Pulsed dye lasers are used effectively in the treatment of psoriasis with long remission time and limited side effects. It is, however, not completely understood which biological processes underlie its favorable outcome. Pulsed dye laser treatment at 585-595 nm targets hemoglobin in the blood, inducing local hyperthermia in surrounding blood vessels and adjacent tissues. While the impact of destructive temperatures on blood vessels has been well studied, the effects of lower temperatures on the function of several cell types within the blood vessel wall and its periphery are not known. The aim of our study is to assess the functionality of isolated blood vessels after exposure to moderate hyperthermia (45 to 60°C) by evaluating the function of endothelial cells, smooth muscle cells, and vascular nerves. We measured blood vessel functionality of rat mesenteric arteries (n=19) by measuring vascular contraction and relaxation before and after heating vessels in a wire myograph. To this end, we elicited vascular contraction by addition of either high potassium solution or the thromboxane analogue U46619 to stimulate smooth muscle cells, and electrical field stimulation (EFS) to stimulate nerves. For measurement of endothelium-dependent relaxation, we used methacholine. Each vessel was exposed to one temperature in the range of 45-60°C for 30 seconds and a relative change in functional response after hyperthermia was determined by comparison with the response per stimulus before heating. Non-linear regression was used to fit our dataset to obtain the temperature needed to reduce blood vessel function by 50% (Half maximal effective temperature, ET50). Our findings demonstrate a substantial decrease in relative functional response for all three cell types following exposure to 55°C-60°C. There was no significant difference between the ET50 values of the different cell types, which was between 55.9°C and 56.9°C (P>0.05). Our data show that blood vessel functionality decreases significantly when exposed to temperatures between 55°C-60°C for 30 seconds. The results show functionality of endothelial cells, smooth muscle cells, and vascular nerves is similarly impaired. These results help to understand the biological effects of hyperthermia and may aid in tailoring laser and light strategies for selective photothermolysis that contribute to disease modification of psoriasis after pulsed dye laser treatment.
Assuntos
Lasers de Corante , Animais , Ratos , Masculino , Lasers de Corante/uso terapêutico , Miócitos de Músculo Liso/fisiologia , Miócitos de Músculo Liso/efeitos da radiação , Vasodilatação/efeitos da radiação , Vasodilatação/fisiologia , Temperatura , Músculo Liso Vascular/efeitos da radiação , Músculo Liso Vascular/fisiologia , Células Endoteliais/efeitos da radiação , Células Endoteliais/fisiologia , Vasoconstrição/efeitos da radiação , Vasoconstrição/fisiologia , Endotélio Vascular/efeitos da radiação , Ratos WistarRESUMO
INTRODUCTION: Peripheral artery disease (PAD) is a highly morbid condition in which impaired blood flow to the limbs leads to pain and tissue loss. Previously we identified 670 nm electromagnetic energy (R/NIR) to increase nitric oxide levels in cells and tissue. NO elicits relaxation of smooth muscle (SMC) by stimulating potassium efflux and membrane hyperpolarization. The actions of energy on ion channel activity have yet to be explored. Here we hypothesized R/NIR stimulates vasodilation through activation of potassium channels in SMC. METHODS: Femoral arteries or facial arteries from C57Bl/6 and Slo1-/- mice were isolated, pressurized to 60 mmHg, pre-constricted with U46619, and irradiated twice with energy R/NIR (10 mW/cm2 for 5 min) with a 10 min dark period between irradiations. Single-channel K+ currents were recorded at room temperature from cell-attached and excised inside-out membrane patches of freshly isolated mouse femoral arterial muscle cells using the patch-clamp technique. RESULTS: R/NIR stimulated vasodilation requires functional activation of the large conductance potassium channels. There is a voltage dependent outward current in SMC with light stimulation, which is due to increases in the open state probability of channel opening. R/NIR modulation of channel opening is eliminated pharmacologically (paxilline) and genetically (BKca α subunit knockout). There is no direct action of light to modulate channel activity as excised patches did not increase the open state probability of channel opening. CONCLUSION: R/NIR vasodilation requires indirect activation of the BKca channel.
Assuntos
Radiação Eletromagnética , Subunidades alfa do Canal de Potássio Ativado por Cálcio de Condutância Alta/metabolismo , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/efeitos da radiação , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/efeitos da radiação , Transdução de Sinais/efeitos da radiação , Vasodilatação/efeitos da radiação , Animais , Estimulação Elétrica/métodos , Terapia por Estimulação Elétrica/métodos , Artéria Femoral/metabolismo , Técnicas de Inativação de Genes , Subunidades alfa do Canal de Potássio Ativado por Cálcio de Condutância Alta/genética , Potenciais da Membrana/efeitos da radiação , Camundongos , Camundongos Knockout , Óxido Nítrico/metabolismo , Técnicas de Patch-Clamp , Doença Arterial Periférica/metabolismo , Doença Arterial Periférica/terapiaRESUMO
Background Chronic inflammation through cellular senescence, known as the senescence-associated secretory phenotype, is a mechanism of various organ diseases, including atherosclerosis. Particularly, ionizing radiation (IR) contributes to cellular senescence by causing DNA damage. Although previous clinical studies have demonstrated that radiotherapy causes atherosclerosis as a long-term side effect, the detailed mechanism is unclear. This study was conducted to investigate the relationship between radiation-induced atherosclerosis and senescence-associated secretory phenotype in murine carotid arteries. Methods and Results Partial ligation of the left carotid artery branches in 9-week-old male apolipoprotein E-deficient mice was performed to induce atherosclerosis. The mice received total body irradiation at a dose of 6 Gy using gamma rays at 2 weeks post operation. We compared the samples collected 4 weeks after IR with unirradiated control samples. The IR and control groups presented pathologically progressive lesions in 90.9% and 72.3% of mice, respectively. Plaque volume, macrophage accumulation, and phenotype switching of vascular smooth muscle cells were advanced in the IR group. Irradiated samples showed increased persistent DNA damage response (53BP1 [p53 binding protein 1]), upregulated cyclin-dependent kinase inhibitors (p16INK4a and p21), and elevated inflammatory chemokines expression (monocyte chemotactic protein-1, keratinocyte-derived chemokine, and macrophage inflammatory protein 2). Conclusions IR promoted plaque growth in murine carotid arteries. Our findings support the possibility that senescence-associated secretory phenotype aggravates atherogenesis in irradiated artery. This mice model might contribute to mechanism elucidation of radiation-induced atherosclerosis.
Assuntos
Aterosclerose/etiologia , Artéria Carótida Primitiva/efeitos da radiação , Macrófagos/patologia , Miócitos de Músculo Liso/efeitos da radiação , Placa Aterosclerótica/etiologia , Lesões Experimentais por Radiação/complicações , Fluxo Sanguíneo Regional/fisiologia , Animais , Apolipoproteínas E/deficiência , Aterosclerose/metabolismo , Aterosclerose/patologia , Artéria Carótida Primitiva/patologia , Senescência Celular/efeitos da radiação , Quimiocinas/biossíntese , Progressão da Doença , Relação Dose-Resposta à Radiação , Macrófagos/metabolismo , Macrófagos/efeitos da radiação , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Placa Aterosclerótica/metabolismo , Placa Aterosclerótica/patologia , Lesões Experimentais por Radiação/metabolismo , Lesões Experimentais por Radiação/patologiaRESUMO
Bronchial thermoplasty (BT) delivers targeted radiofrequency energy to bronchial airway walls and results in the partial ablation of the airway smooth muscle that is responsible for bronchoconstriction. It is approved for the treatment of severe persistent asthma. Multiple, large clinical trials including a recent "real-world" study demonstrate significant improvements in asthma-related quality of life, reduction in asthma exacerbations, emergency department visits, and hospitalizations after BT that is sustained out to 5 years. In this article, we review the state of the art of BT treatment in severe persistent asthma and share a decade of BT research and clinical experience. We share our personal experience and introduce the three "I"s (identification, implementation, and intense follow-up) that we believe promote successful patient outcomes and help build a successful BT program.
Assuntos
Asma/terapia , Brônquios/patologia , Termoplastia Brônquica/métodos , Miócitos de Músculo Liso/efeitos da radiação , Terapia por Radiofrequência/métodos , Broncoconstrição , Ensaios Clínicos como Assunto , Progressão da Doença , Humanos , Miócitos de Músculo Liso/fisiologia , Qualidade de Vida , Ablação por Radiofrequência , Resultado do TratamentoRESUMO
The long-term adverse effects of radiotherapy on cardiovascular disease are well documented. However, the underlying mechanisms responsible for this increased risk are poorly understood. Previous studies using rigorous smooth muscle cell (SMC) lineage tracing have shown abundant SMC investment into atherosclerotic lesions, where SMCs contribute to the formation of a protective fibrous cap. Studies herein tested whether radiation impairs protective adaptive SMC responses during vascular disease. To do this, we exposed SMC lineage tracing (Myh11-ERT2Cre YFP+) mice to lethal radiation (1,200 cGy) followed by bone marrow transplantation prior to atherosclerosis development or vessel injury. Surprisingly, following irradiation, we observed a complete loss of SMC investment in 100% of brachiocephalic artery (BCA), carotid artery, and aortic arch lesions. Importantly, this was associated with a decrease in multiple indices of atherosclerotic lesion stability within the BCA. Interestingly, we observed anatomic heterogeneity, as SMCs accumulated normally into lesions of the aortic root and abdominal aorta, suggesting that SMC sensitivity to lethal irradiation occurs in blood vessels of neural crest origin. Taken together, these results reveal an undefined and unintended variable in previous studies using lethal irradiation and may help explain why patients exposed to radiation have increased risk for cardiovascular disease.
Assuntos
Aterosclerose/patologia , Tronco Braquiocefálico/efeitos da radiação , Músculo Liso Vascular/efeitos da radiação , Miócitos de Músculo Liso/efeitos da radiação , Animais , Aorta Abdominal/patologia , Aorta Abdominal/efeitos da radiação , Aterosclerose/etiologia , Medula Óssea/efeitos da radiação , Transplante de Medula Óssea , Tronco Braquiocefálico/patologia , Diferenciação Celular/efeitos da radiação , Modelos Animais de Doenças , Humanos , Masculino , Camundongos , Camundongos Knockout para ApoE , Músculo Liso Vascular/citologia , Irradiação Corporal TotalRESUMO
PURPOSE: To study the effect of x-ray radiotherapy on vascular smooth muscle cells (VSMCs) and elucidate the mechanisms in preventing neointimal hyperplasia of prosthetic vascular grafts. MATERIALS AND METHODS: In model I, twelve mongrel dogs underwent revascularization with prosthetic grafts and half the dogs underwent irradiation of the grafts at 28 Gy. In model II, human VSMCs (hVSMCs) were maintained and divided into six groups to which external radiation was applied at six different doses: 0 Gy, 2 Gy, 8 Gy, 16 Gy, 24 Gy and 30 Gy. In both models, specimens were harvested and examined by using morphological, immunological, cellular and molecular methods. RESULTS: After irradiation, the neointima thickness was significantly lower in irradiated groups (p≤0.01). The radiotherapy could up-regulate p27kip1, and down-regulate proliferating cell nuclear antigen (PCNA) and S phase kinase associated protein 2 (Skp2). X-ray irradiation inhibits the proliferation of hVSMCs via acting on G1/S phase of cell cycle. The apoptosis of hVSMCs increased significantly with dose and time. The expression of PCNA and Skp2 were decreased after a first increasing trend with dose, but had a significant negative correlation with time. The expression of p27kip1 had a significant positive correlation with dose and time. CONCLUSIONS: Postoperative external fractionated irradiation after prosthetic vessel replacement of the abdominal aorta suppressed the development of hyperplasia in the graft neointima in the short term. There was a prominent time- and dose-dependent inhibition of VSMC proliferation by radiation when it was administered.
Assuntos
Aorta Abdominal/efeitos da radiação , Aorta Abdominal/cirurgia , Apoptose/efeitos dos fármacos , Implante de Prótese Vascular/instrumentação , Prótese Vascular , Proliferação de Células/efeitos da radiação , Músculo Liso Vascular/efeitos da radiação , Músculo Liso Vascular/cirurgia , Miócitos de Músculo Liso/efeitos da radiação , Neointima , Animais , Aorta Abdominal/metabolismo , Aorta Abdominal/patologia , Implante de Prótese Vascular/efeitos adversos , Células Cultivadas , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Cães , Relação Dose-Resposta à Radiação , Humanos , Hiperplasia , Modelos Animais , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Antígeno Nuclear de Célula em Proliferação/metabolismo , Proteínas Quinases Associadas a Fase S/metabolismo , Fatores de TempoRESUMO
Vascular restenosis after injury of blood vessel has been implicated in various responses including apoptosis, migration, and proliferation in vascular smooth muscle cells (VSMCs) stimulated by diverse growth factors underlying platelet-derived growth factor (PDGF). Previous studies evaluated the effects of low-power laser (LPL) irradiation over various wavelength ranges on VSMC events in normal and pathologic states. However, whether VSMC responses are affected by LPL irradiation remains unclear. The purpose of this study is to explore the effects of LPL (green diode laser 532-nm pulsed wave of 300 mW at a spot diameter of 1 mm) irradiation on the responses, apoptosis, migration, and proliferation of VSMCs. The effect of LPL irradiation was tested on VSMCs through cytotoxicity, proliferation, migration, and apoptotic assays. Aortic ring assay was used to assess the effect of LPL irradiation on aortic sprout outgrowth. Protein expression levels were determined by western blotting. LPL irradiation did not affect VSMC viability but slightly attenuated PDGF-BB-induced proliferation in VSMCs. In addition, LPL irradiation inhibited PDGF-BB-evoked migration of VSMCs. Aortic sprout outgrowth in response to PDGF-BB was diminished in cells treated with LPL. In contrast, LPL irradiation evoked apoptosis in VSMCs in the presence of PDGF-BB. Similarly, activation of caspase-3 and Bax, as well as p38 mitogen-activated protein kinase (MAPK), in VSMCs treated with PDGF-BB was enhanced by exposure to LPL. These findings indicate that LPL irradiation induces vascular apoptosis via p38 MAPK activation and simultaneously inhibits VSMC proliferation and migration in response to PDGF-BB.
Assuntos
Apoptose/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Terapia com Luz de Baixa Intensidade , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/patologia , Miócitos de Músculo Liso/efeitos da radiação , Proteínas Proto-Oncogênicas c-sis/farmacologia , Animais , Aorta/citologia , Becaplermina , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Masculino , Miócitos de Músculo Liso/efeitos dos fármacos , Ratos Sprague-DawleyRESUMO
Skin flap grafting is a form of transplantation widely used in plastic surgery. However, ischemia/reperfusion injury is the main factor which reduces the survival rate of flaps following grafting. We investigated whether photobiomodulation (PBM) precondition prior to human adipose-derived stromal cell (hASC) spheroid (PBM-spheroid) transplantation improved skin tissue functional recovery by the stimulation of angiogenesis and tissue regeneration in skin flap of mice. The LED had an emission wavelength peaked at 660 ± 20 nm (6 J/cm2, 10 mW/cm2). The expression of angiogenic growth factors in PBM-spheroid hASCs was much greater than that of not-PBM-treated spheroid or monolayer-cultured hASCs. From immunochemical staining analysis, the hASCs of PBM-spheroid were CD31+, KDR+, and CD34+, whereas monolayer-cultured hASCs were negative for these markers. To evaluate the therapeutic effect of hASC PBM-spheroid in vivo, PBS, monolayer-cultured hASCs, and not-PBM-spheroid were transplanted into a skin flap model. The animals were observed for 14 days. The PBM-spheroid hASCs transplanted into the skin flap ischemia differentiated into endothelial cells and remained differentiated. Transplantation of PBM-spheroid hASCs into the skin flap ischemia significantly elevated the density of vascular formations through angiogenic factors released by the skin flap ischemia and enhanced tissue regeneration at the lesion site. Consistent with these results, the transplantation of PBM-spheroid hASCs significantly improved functional recovery compared with PBS, monolayer-cultured hASCs, and not-PBM-spheroid treatment. These findings suggest that transplantation of PBM-spheroid hASCs may be an effective stem cell therapy for the treatment of skin flap ischemia.
Assuntos
Tecido Adiposo/citologia , Isquemia/terapia , Terapia com Luz de Baixa Intensidade , Regeneração/efeitos da radiação , Pele/irrigação sanguínea , Esferoides Celulares/citologia , Células-Tronco/citologia , Retalhos Cirúrgicos/irrigação sanguínea , Animais , Diferenciação Celular/efeitos da radiação , Sobrevivência Celular/efeitos da radiação , Modelos Animais de Doenças , Células Endoteliais/citologia , Células Endoteliais/efeitos da radiação , Células Epiteliais/citologia , Células Epiteliais/efeitos da radiação , Humanos , Isquemia/patologia , Camundongos , Miócitos de Músculo Liso/citologia , Miócitos de Músculo Liso/efeitos da radiação , Neovascularização Fisiológica/efeitos da radiação , Pele/patologia , Esferoides Celulares/efeitos da radiação , Transplante de Células-Tronco , Células-Tronco/efeitos da radiação , CicatrizaçãoRESUMO
Arresting or regressing growth of abdominal aortic aneurysms (AAAs), localized expansions of the abdominal aorta are contingent on inhibiting chronically overexpressed matrix metalloproteases (MMPs)-2 and -9 that disrupt elastic matrix within the aortic wall, concurrent with providing a stimulus to augmenting inherently poor auto-regeneration of these matrix structures. In a recent study we demonstrated that localized, controlled and sustained delivery of doxycycline (DOX; a tetracycline-based antibiotic) from poly(lactic-co-glycolic acid) nanoparticles (PLGA NPs), enhances elastic matrix deposition and MMP-inhibition at a fraction of the therapeutically effective oral dose. The surface functionalization of these NPs with cationic amphiphiles, which enhances their arterial uptake, was also shown to have pro-matrix regenerative and anti-MMP effects independent of the DOX. Based on the hypothesis that the incorporation of superparamagnetic iron oxide NPs (SPIONs) within these PLGA NPs would enhance their targetability to the AAA site under an applied external magnetic field, we sought to evaluate the functional effects of NPs co-encapsulating DOX and SPIONs (DOX-SPION NPs) on elastic matrix regeneration and MMP synthesis/activity in vitro within aneurysmal smooth muscle cell (EaRASMC) cultures. The DOX-SPION NPs were mobile under an applied external magnetic field, while enhancing elastic matrix deposition 1.5-2-fold and significantly inhibiting MMP-2 synthesis and MMP-2 and -9 activities, compared to NP-untreated control cultures. These results illustrate that the multifunctional benefits of NPs are maintained following SPION co-incorporation. Additionally, preliminary studies carried out demonstrated enhanced targetability of SPION-loaded NPs within proteolytically-disrupted porcine carotid arteries ex vivo, under the influence of an applied external magnetic field. Thus, this dual-agent loaded NP system proffers a potential non-surgical option for treating small growing AAAs, via controlled and sustained drug release from multifunctional, targetable nanocarriers. STATEMENT OF SIGNIFICANCE: Proactive screening of high risk elderly patients now enables early detection of abdominal aortic aneurysms (AAAs). There are no established drug-based therapeutic alternatives to surgery for AAAs, which is unsuitable for many elderly patients, and none which can achieve restore disrupted and lost elastic matrix in the AAA wall, which is essential to achieve growth arrest or regression. We have developed a first generation design of polymer nanoparticles (NPs) for AAA tissue localized delivery of doxycycline, a modified tetracycline drug at low micromolar doses at which it provides both pro-elastogenic and anti-proteolytic benefits that can augment elastic matrix regenerative repair. The nanocarriers themselves are also uniquely chemically functionalized on their surface to also provide them pro-elastin-regenerative & anti-matrix degradative properties. To provide an active driving force for efficient uptake of intra-lumenally infused NPs to the AAA wall, in this work, we have rendered our polymer NPs mobile in an applied magnetic field via co-incorporation of super-paramagnetic iron oxide NPs. We demonstrate that such modifications significantly improve wall uptake of the NPs with no significant changes to their physical properties and regenerative benefits. Such NPs can potentially stimulate structural repair in the AAA wall following one time infusion to delay or prevent AAA growth to rupture. The therapy can provide a non-surgical treatment option for high risk AAA patients.
Assuntos
Preparações de Ação Retardada/administração & dosagem , Dextranos/administração & dosagem , Doxiciclina/administração & dosagem , Nanopartículas de Magnetita/administração & dosagem , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/efeitos da radiação , Regeneração/efeitos dos fármacos , Animais , Células Cultivadas , Preparações de Ação Retardada/química , Preparações de Ação Retardada/efeitos da radiação , Dextranos/efeitos da radiação , Doxiciclina/química , Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/efeitos da radiação , Campos Magnéticos , Nanopartículas de Magnetita/efeitos da radiação , Masculino , Nanocápsulas/administração & dosagem , Nanocápsulas/química , Nanocápsulas/efeitos da radiação , Doses de Radiação , Ratos , Ratos Sprague-Dawley , Regeneração/efeitos da radiaçãoRESUMO
Whole brain radiation therapy for the treatment of tumors can sometimes cause cognitive impairment. Memory deficits were noted in up to 50% of treated patients over a short period of several months. In addition, an increased rate of dementia in young patients has been noted over the longer term, i.e. years. A deficit in neurogenesis after irradiation has been postulated to be the main cause of cognitive decline in patients, but recent data on irradiation therapy for limited parts of the brain appear to indicate other possibilities. Irradiation can directly damage various types of cells other than neuronal stem cells. However, this paper will focus on injury to brain vasculature leading to cognitive decline since vessels represent a better therapeutic target for drug development than other cells in the brain because of the blood-brain barrier.
Assuntos
Vasos Sanguíneos/lesões , Vasos Sanguíneos/efeitos da radiação , Encéfalo/efeitos da radiação , Transtornos Cognitivos/etiologia , Neoplasias/radioterapia , Radioterapia/efeitos adversos , Vasos Sanguíneos/patologia , Humanos , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/efeitos da radiaçãoRESUMO
Paclitaxel (PTX) has been recognized as a promising drug for intervention of vascular reconstructions. However, it is still difficult to achieve local drug delivery in a spatio-temporally controllable manner under real-time image guidance. Here, we introduce an ultrasound (US) triggered image-guided drug delivery approach to inhibit vascular reconstruction via paclitaxel (PTX)-loaded microbubbles (PLM) in a rabbit iliac balloon injury model. PLM was prepared through encapsulating PTX in the shell of lipid microbubbles via film hydration and mechanical vibration technique. Our results showed PLM could effectively deliver PTX when exposed to US irradiation and result in significantly lower viability of vascular smooth muscle cells. Ultrasonographic examinations revealed the US signals from PLM in the iliac artery were greatly increased after intravenous administration of PLM, making it possible to identify the restenosis regions of iliac artery. The in vivo anti-restenosis experiments with PLM and US greatly inhibited neointimal hyperplasia at the injured site, showing an increased lumen area and reduced the ratio of intima area and the media area (I/M ratio). No obvious functional damages to liver and kidney were observed for those animals. Our study provided a promising approach to realize US triggered image-guided PTX delivery for therapeutic applications against iliac restenosis.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Constrição Patológica/terapia , Sistemas de Liberação de Medicamentos/métodos , Artéria Ilíaca/efeitos dos fármacos , Microbolhas , Paclitaxel/farmacologia , Animais , Constrição Patológica/patologia , Composição de Medicamentos , Sistemas de Liberação de Medicamentos/instrumentação , Artéria Ilíaca/patologia , Músculo Liso Vascular/efeitos da radiação , Miócitos de Músculo Liso/efeitos da radiação , Coelhos , Ondas UltrassônicasRESUMO
OBJECTIVE: The aim of the study was to investigate the differentiation potential of adipose-derived stem cells (ADSCs) when cocultured with smooth muscle cells (SMCs), and to determine the role of low-intensity laser irradiation (LILI). BACKGROUND DATA: ADSCs isolated from adipose tissue are isolated with ease and in large amounts. SMCs constitute most parts of the intestinal, urinary, reproductive, and cardiovascular systems. LILI has been found to have positive effects on different cell types, including ADSCs. METHODS: The study used ADSCs (Stempro Adipose Derived Stem Cells-R7788-115) and SMCs (SKU-T-1 American Type Culture Collection HTB-114) cell lines. These cell lines were cocultured in a 1:1 ratio with and without growth factors and then exposed to LILI using 636 nm at 5 J/cm2. RESULTS: Cell viability and proliferation increased significantly in the cocultured groups that were exposed to LILI alone, as well as in combination with growth factors. Further, there was a significant decrease in the expression of stem cell markers with a concomitant increase in SMC markers. CONCLUSIONS: These results suggest that ADSCs have the ability to differentiate into SMCs when cocultured with SMCs, whereas LILI potentially augments the differentiation potential and need. This further highlights the significant role that LILI has to offer ADSC therapy in regenerative medicine.
Assuntos
Adipócitos/efeitos da radiação , Diferenciação Celular/efeitos da radiação , Terapia com Luz de Baixa Intensidade/métodos , Células-Tronco Mesenquimais/efeitos da radiação , Miócitos de Músculo Liso/efeitos da radiação , Adipócitos/fisiologia , Tecido Adiposo/citologia , Diferenciação Celular/fisiologia , Proliferação de Células/fisiologia , Proliferação de Células/efeitos da radiação , Sobrevivência Celular/efeitos da radiação , Células Cultivadas , Técnicas de Cocultura/métodos , Humanos , Células-Tronco Mesenquimais/fisiologia , Miócitos de Músculo Liso/fisiologia , Papel (figurativo) , Sensibilidade e EspecificidadeRESUMO
Age-associated cardiovascular diseases are at least partially ascribable to vascular cell senescence. Replicative senescence (RS) and stress-induced premature senescence (SIPS) are provoked respectively by endogenous (telomere erosion) and exogenous (H2O2, UV) stimuli resulting in cell cycle arrest in G1 and G2 phases. In both scenarios, mitochondria-derived ROS are important players in senescence initiation. We aimed to define whether a mtDNA-transcribed long-non-coding-RNA (lncRNA), ASncmtRNA-2, has a role in vascular aging and senescence. Aortas of old mice, characterized by increased senescence, showed an increment in ASncmtRNA-2 expression. In vitro analysis of Endothelial Cells (EC) and Vascular Smooth Muscle Cells (VSMC) established that ASncmtRNA-2 is induced in EC, but not in VSMC, during RS. Surprisingly, ASncmtRNA-2 is not upregulated in two different EC SIPS scenarios, treated with H2O2 and UV. The p16 gene displayed similar ASncmtRNA-2 expression patterns, suggesting a possible co-regulation of the two genes. Interestingly, the expression of two miRNAs, hsa-miR-4485 and hsa-miR-1973, with perfect homology to the double strand region of ASncmtRNA-2 and originating at least in part from a mitochondrial transcript, was induced in RS, opening to the possibility that this lncRNA functions as a non-canonical precursor of these miRNAs. Cell cycle analysis of EC transiently over-expressing ASncmtRNA-2 revealed an accumulation of EC in the G2/M phase, but not in the G1 phase. We propose that ASncmtRNA-2 in EC might be involved in the RS establishment by participating in the cell cycle arrest in G2/M phase, possibly through the production of hsa-miR-4485 and hsa-miR-1973. This article is part of a Special Issue entitled: Mitochondria.
Assuntos
Envelhecimento/metabolismo , Células Endoteliais da Veia Umbilical Humana/metabolismo , Mitocôndrias/metabolismo , Miócitos de Músculo Liso/metabolismo , RNA Longo não Codificante/genética , RNA/genética , Envelhecimento/genética , Animais , Aorta/citologia , Aorta/metabolismo , Sequência de Bases , Senescência Celular , Inibidor p16 de Quinase Dependente de Ciclina/genética , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos da radiação , Regulação da Expressão Gênica , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/efeitos da radiação , Humanos , Peróxido de Hidrogênio/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/genética , MicroRNAs/metabolismo , Mitocôndrias/genética , Dados de Sequência Molecular , Miócitos de Músculo Liso/citologia , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/efeitos da radiação , RNA/metabolismo , RNA Longo não Codificante/metabolismo , RNA Mitocondrial , Transdução de Sinais , Raios UltravioletaRESUMO
Because of high exposure to systemic noxae, vascular endothelial cells (EC) have to ensure distinct damage defense and regenerative mechanisms to guarantee vascular health. For meaningful toxicological drug assessments employing embryonic stem cell (ESC)-based in vitro models, functional competence of differentiated progeny and detailed knowledge regarding damage defense mechanisms are essential. Here, mouse ESCs (mESC) were differentiated into functionally competent vascular cells (EC and smooth muscle cells [SMC]). mESC, EC, and SMC were comparatively analyzed regarding DNA repair and DNA damage response (DDR). Differentiation was accompanied by both congruent and unique alterations in repair and DDR characteristics. EC and SMC shared the downregulation of genes involved cell cycle regulation and repair of DNA double-strand breaks (DSBs) and mismatches, whereas genes associated with nucleotide excision repair (NER), apoptosis, and autophagy were upregulated when compared with mESC. Expression of genes involved in base excision repair (BER) was particularly low in SMC. IR-induced formation of DSBs, as detected by nuclear γH2AX foci formation, was most efficient in SMC, the repair of DSBs was fastest in EC. Together with substantial differences in IR-induced phosphorylation of p53, Chk1, and Kap1, the data demonstrate complex alterations in DDR capacity going along with the loss of pluripotency and gain of EC- and SMC-specific functions. Notably, IR exposure of early vascular progenitors did not impair differentiation into functionally competent EC and SMC. Summarizing, mESC-based vascular differentiation models are informative to study the impact of environmental stressors on differentiation and function of vascular cells.
Assuntos
Diferenciação Celular/efeitos da radiação , Células-Tronco Embrionárias/efeitos da radiação , Células Progenitoras Endoteliais/efeitos da radiação , Músculo Liso Vascular/efeitos da radiação , Miócitos de Músculo Liso/efeitos da radiação , Células-Tronco Pluripotentes/efeitos da radiação , Animais , Apoptose/genética , Apoptose/efeitos da radiação , Proteínas Reguladoras de Apoptose/genética , Proteínas Reguladoras de Apoptose/metabolismo , Autofagia/genética , Autofagia/efeitos da radiação , Biomarcadores/metabolismo , Ciclo Celular/genética , Ciclo Celular/efeitos da radiação , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular , Quebras de DNA de Cadeia Dupla , Reparo do DNA , Células-Tronco Embrionárias/metabolismo , Células-Tronco Embrionárias/patologia , Células Progenitoras Endoteliais/metabolismo , Células Progenitoras Endoteliais/patologia , Regulação da Expressão Gênica , Histonas/metabolismo , Camundongos , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Neovascularização Fisiológica/genética , Neovascularização Fisiológica/efeitos da radiação , Células-Tronco Pluripotentes/metabolismo , Células-Tronco Pluripotentes/patologia , RNA Mensageiro/metabolismo , Fatores de TempoRESUMO
Ultrasound, a non-invasive therapy method, is a potential tool for medical applications, but its biological effects on vascular smooth muscle cells (VSMCs) have not been characterized. The aim of this study was to explore the effect and possible apoptotic mechanism of VSMCs that were induced by low-frequency ultrasound (LFU). Cell viability and apoptosis of A7r5 cells were evaluated after treating A7r5 cells with a continuous 45-kHz 1.0-W/cm(2) ultrasound (exposure time of 0, 10, 20, 30, and 35 s) by MTT assay and flow cytometry. At the optimum ultrasound exposure condition (30 s), gene chip analysis was performed, and the apoptotic signaling pathway was confirmed by reverse transcription-polymerase chain reaction and Western blot. As measured by flow cytometry, LFU significantly induced A7r5 cell apoptosis. Comparing the ultrasound group with the control group, the protein expression of caspase-9 and caspase-3 was increased by 50 and 57%, respectively; the caspase-3 mRNA level was increased by 37.5%. These findings indicate that an intrinsic pathway plays a major role in apoptosis that is induced by LFU and that LFU can induce A7r5 cell apoptosis via caspase-9- and caspase-3-dependent pathways.
Assuntos
Aorta/metabolismo , Apoptose/efeitos da radiação , Miócitos de Músculo Liso/metabolismo , Animais , Aorta/citologia , Aorta/efeitos da radiação , Caspase 3/biossíntese , Caspase 9/biossíntese , Linhagem Celular , Proliferação de Células/efeitos da radiação , Sobrevivência Celular/efeitos da radiação , Regulação Enzimológica da Expressão Gênica/efeitos da radiação , Miócitos de Músculo Liso/efeitos da radiação , RNA Mensageiro/biossíntese , Ratos , SomRESUMO
PURPOSE: γ-rays (IR) cause an increase in intracellular calcium [Ca(2+)], alters contractility and triggers apoptosis via the activation of protein kinase C in intestinal guinea pig smooth muscle cells. The present study investigated the role of the mitochondria in these processes and characterized proteins involved in IR-induced apoptosis. MATERIALS AND METHODS: Intestinal smooth muscle cells were exposed to 10-50 Gy from a (60)Co γ-source. Reactive oxygen species (ROS) levels were measured by colourimetry with a fluorescente probe. Protein expression was analyzed by immunoblotting and immunofluorescence. RESULTS: Apoptosis was inhibited by glutathione, possible by inhibiting the generation or scavenging ROS. Apoptosis was mediated by the mitochondria releasing cytochrome c leading to caspase 3 activation. IR increased the expression of the cyclins A, B2 and E and led to unbalanced cellular growth in an absorption dose-dependent manner. However, radiation did not induce alterations in the mitochondrial ultrastructure or in transmembrane electric potential. In contrast, IR increased the nuclear expression of cytoplasmic proteins and cyclins A and E. CONCLUSION: Smooth muscle cells subjected to IR undergo mitochondrial-mediated apoptosis that involves oncoproteins activation and preserves mitochondrial structure. IR also cause alterations in the expression and localization of both pro- and anti-apoptotic proteins.
Assuntos
Apoptose/fisiologia , Sinalização do Cálcio/fisiologia , Mitocôndrias Musculares/fisiologia , Contração Muscular/fisiologia , Miócitos de Músculo Liso/citologia , Miócitos de Músculo Liso/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Animais , Apoptose/efeitos dos fármacos , Cálcio/metabolismo , Sinalização do Cálcio/efeitos da radiação , Ciclo Celular/fisiologia , Ciclo Celular/efeitos da radiação , Proteínas de Ciclo Celular/metabolismo , Células Cultivadas , Relação Dose-Resposta à Radiação , Raios gama , Cobaias , Mitocôndrias Musculares/efeitos da radiação , Contração Muscular/efeitos da radiação , Proteínas Musculares/metabolismo , Miócitos de Músculo Liso/efeitos da radiação , Doses de RadiaçãoRESUMO
Focal drug delivery to a vessel wall facilitated by intravascular ultrasound and microbubbles holds promise as a potential therapy for atherosclerosis. Conventional methods of microbubble administration result in rapid clearance from the bloodstream and significant drug loss. To address these limitations, we evaluated whether drug delivery could be achieved with transiently stable microbubbles produced in real time and in close proximity to the therapeutic site. Rat aortic smooth muscle cells were placed in a flow chamber designed to simulate physiological flow conditions. A flow-focusing microfluidic device produced 8 µm diameter monodisperse microbubbles within the flow chamber, and ultrasound was applied to enhance uptake of a surrogate drug (calcein). Acoustic pressures up to 300 kPa and flow rates up to 18 mL/s were investigated. Microbubbles generated by the flow-focusing microfluidic device were stabilized with a polyethylene glycol-40 stearate shell and had either a perfluorobutane (PFB) or nitrogen gas core. The gas core composition affected stability, with PFB and nitrogen microbubbles exhibiting half-lives of 40.7 and 18.2 s, respectively. Calcein uptake was observed at lower acoustic pressures with nitrogen microbubbles (100 kPa) than with PFB microbubbles (200 kPa) (p < 0.05, n > 3). In addition, delivery was observed at all flow rates, with maximal delivery (>70% of cells) occurring at a flow rate of 9 mL/s. These results demonstrate the potential of transiently stable microbubbles produced in real time and in close proximity to the intended therapeutic site for enhancing localized drug delivery.
Assuntos
Sistemas de Liberação de Medicamentos/instrumentação , Fluoresceínas/farmacocinética , Microbolhas/uso terapêutico , Técnicas Analíticas Microfluídicas/instrumentação , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/efeitos da radiação , Sonicação/instrumentação , Animais , Células Cultivadas , Desenho de Equipamento , Análise de Falha de Equipamento , RatosRESUMO
Methods of controllable, noncontact rotation of optically trapped microscopic objects have garnered significant attention for tomographic imaging and microfluidic actuation. Here, we report development of a fiber-optic spanner and demonstrate controlled rotation of smooth muscle cells. The rotation is realized by introducing a transverse offset between two counterpropagating beams emanating from single-mode optical fibers. The rotation speed and surrounding microfluidic flow could be controlled by varying balanced laser beam powers. Further, we demonstrate simultaneous translation and rotation of the fiber-optically trapped cell by varying the laser power of one fiber-optic arm.
Assuntos
Separação Celular/instrumentação , Rastreamento de Células/instrumentação , Tecnologia de Fibra Óptica/instrumentação , Técnicas Analíticas Microfluídicas/instrumentação , Miócitos de Músculo Liso/citologia , Miócitos de Músculo Liso/fisiologia , Pinças Ópticas , Movimento Celular/fisiologia , Polaridade Celular , Células Cultivadas , Desenho de Equipamento , Análise de Falha de Equipamento , Humanos , Luz , Miócitos de Músculo Liso/efeitos da radiação , RotaçãoRESUMO
Hemodialysis vascular access dysfunction is a major cause of morbidity and hospitalization in the hemodialysis population at a cost of well over $1 billion per annum. Venous stenosis (due to venous neointimal hyperplasia [VNH]) is the most common cause of polytetrafluroethylene PTFE) dialysis access graft and arteriovenous fistula (AVF) failure. Despite the magnitude of the clinical problem, however, there are currently no effective therapies for this condition. We and others have previously demonstrated that VNH in PTFE dialysis grafts and AVF is composed of smooth muscle cells/myofibroblasts, endothelial cells within neointimal microvessels, and peri-graft macrophages. Radiation therapy blocks the proliferation and activation of all these cell types. The current review will dissect out the available in vitro, experimental, and clinical data on the use of radiation therapy for vascular stenosis in general, and for dialysis access dysfunction in particular. It is important to try and identify whether there is still a role for radiation therapy in this specific clinical setting. We believe that this is a critically important question to answer in view of the huge unmet clinical need that is currently associated with hemodialysis vascular access dysfunction.