RESUMEN
The intima, comprising the endothelium and the subendothelial matrix, plays a crucial role in atherosclerosis pathogenesis. The mechanical stress arising from disturbed blood flow (d-flow) and the stiffening of the arterial wall contributes to endothelial dysfunction. However, the specific impacts of these physical forces on the mechanical environment of the intima remain undetermined. Here, we investigated whether inhibiting collagen crosslinking could ameliorate the detrimental effects of persistent d-flow on the mechanical properties of the intima. Partial ligation of the left carotid artery (LCA) was performed in C57BL/6J mice, inducing d-flow. The right carotid artery (RCA) served as an internal control. Carotids were collected 2 days and 2 weeks after surgery to study acute and chronic effects of d-flow on the mechanical phenotype of the intima. The chronic effects of d-flow were decoupled from the ensuing arterial wall stiffening by administration of ß-aminopropionitrile (BAPN), an inhibitor of collagen crosslinking by lysyl oxidase (LOX) enzymes. Atomic force microscopy (AFM) was used to determine stiffness of the endothelium and the denuded subendothelial matrix in en face carotid preparations. The stiffness of human aortic endothelial cells (HAEC) cultured on soft and stiff hydrogels was also determined. Acute exposure to d-flow caused a slight decrease in endothelial stiffness in male mice but had no effect on the stiffness of the subendothelial matrix in either sex. Regardless of sex, the intact endothelium was softer than the subendothelial matrix. In contrast, exposure to chronic d-flow led to a substantial increase in the endothelial and subendothelial stiffness in both sexes. The effects of chronic d-flow were largely prevented by concurrent BAPN administration. In addition, HAEC displayed reduced stiffness when cultured on soft vs. stiff hydrogels. We conclude that chronic d-flow results in marked stiffening of the arterial intima, which can be effectively prevented by inhibition of collagen crosslinking.
Asunto(s)
Arterias Carótidas , Ratones Endogámicos C57BL , Rigidez Vascular , Animales , Masculino , Rigidez Vascular/efectos de los fármacos , Ratones , Arterias Carótidas/efectos de los fármacos , Arterias Carótidas/patología , Arterias Carótidas/fisiopatología , Femenino , Túnica Íntima/patología , Túnica Íntima/efectos de los fármacos , Colágeno/metabolismo , Aminopropionitrilo/farmacología , Proteína-Lisina 6-Oxidasa/metabolismo , Proteína-Lisina 6-Oxidasa/antagonistas & inhibidores , Microscopía de Fuerza Atómica , Humanos , Estrés Mecánico , Endotelio Vascular/efectos de los fármacos , Endotelio Vascular/patología , Endotelio Vascular/fisiopatología , Endotelio Vascular/metabolismoRESUMEN
The topical elastase murine model of abdominal aortic aneurysm (AAA) is enhanced when combined with ß-aminopropionitrile (BAPN)-supplemented drinking water to reliably produce true infrarenal aneurysms with behaviors that mimic human AAAs. Topically applying elastase to the adventitia of the infrarenal aorta causes structural damage to the elastic layers of the aortic wall and initiates aneurysmal dilation. Co-administering BAPN, a lysyl oxidase inhibitor, promotes sustained wall degeneration by reducing collagen and elastin crosslinking. This combination results in large AAAs that progressively expand, form intraluminal thrombus, and are capable of rupture. Refining surgical techniques, such as circumferentially isolating the entire infrarenal aortic segment, can help standardize the procedure for a consistent and thorough application of porcine pancreatic elastase despite different operators and anatomic variations between mice. Therefore, the elastase/BAPN model is a refined approach to surgically inducing AAA in mice, which may better recapitulate human aneurysms and provide additional opportunities to study aneurysm growth and rupture risk.
Asunto(s)
Aminopropionitrilo , Aneurisma de la Aorta Abdominal , Modelos Animales de Enfermedad , Elastasa Pancreática , Animales , Elastasa Pancreática/administración & dosificación , Aneurisma de la Aorta Abdominal/patología , Aminopropionitrilo/administración & dosificación , Ratones , Administración Oral , Administración Tópica , MasculinoRESUMEN
There is no treatment for acute aortic dissection (AAD) targeting inflammatory cells. We aimed to identify the new therapeutic targets associated with inflammatory cells. We characterized the specific distribution of myeloid cells of both human type A AAD samples and a murine AAD model generated using angiotensin II (ANGII) and ß-aminopropionitrile (BAPN) by single-cell RNA sequencing (scRNA-seq). We also examined the effect of an anti-interleukin-1ß (IL-1ß) antibody in the murine AAD model. IL1B+ inflammatory macrophages and classical monocytes were increased in human AAD samples. Trajectory analysis demonstrated that IL1B+ inflammatory macrophages differentiated from S100A8/9/12+ classical monocytes uniquely observed in the aorta of AAD. We found increased infiltration of neutrophils and monocytes with the expression of inflammatory cytokines in the aorta and accumulation of inflammatory macrophages before the onset of macroscopic AAD in the murine AAD model. In blocking experiments using an anti-IL-1ß antibody, it improved survival of murine AAD model by preventing elastin degradation. We observed the accumulation of inflammatory macrophages expressing IL-1ß in both human AAD samples and in a murine AAD model. Anti-IL-1ß antibody could improve the mortality rate in mice, suggesting that it may be a treatment option for AAD.
Asunto(s)
Disección Aórtica , Modelos Animales de Enfermedad , Interleucina-1beta , Macrófagos , Disección Aórtica/metabolismo , Disección Aórtica/patología , Interleucina-1beta/metabolismo , Animales , Humanos , Macrófagos/metabolismo , Macrófagos/inmunología , Ratones , Masculino , Aminopropionitrilo/farmacología , Angiotensina II/metabolismo , Inflamación/metabolismo , Inflamación/patología , Monocitos/metabolismo , Aorta/metabolismo , Aorta/patología , Ratones Endogámicos C57BL , FemeninoRESUMEN
Inflammation assumes a pivotal role in the aortic remodeling of aortic dissection (AD). Asiatic acid (AA), a triterpene compound, is recognized for its strong anti-inflammatory properties. Yet, its effects on ß-aminopropionitrile (BAPN)-triggered AD have not been clearly established. The objective is to determine whether AA attenuates adverse aortic remodeling in BAPN-induced AD and clarify potential molecular mechanisms. In vitro studies, RAW264.7 cells pretreated with AA were challenged with lipopolysaccharide (LPS), and then the vascular smooth muscle cells (VSMCs)-macrophage coculture system was established to explore intercellular interactions. To induce AD, male C57BL/6J mice at three weeks of age were administered BAPN at a dosage of 1 g/kg/d for four weeks. To decipher the mechanism underlying the effects of AA, RNA sequencing analysis was conducted, with subsequent validation of these pathways through cellular experiments. AA exhibited significant suppression of M1 macrophage polarization. In the cell coculture system, AA facilitated the transformation of VSMCs into a contractile phenotype. In the mouse model of AD, AA strikingly prevented the BAPN-induced increases in inflammation cell infiltration and extracellular matrix degradation. Mechanistically, RNA sequencing analysis revealed a substantial upregulation of CX3CL1 expression in BAPN group but downregulation in AA-treated group. Additionally, it was observed that the upregulation of CX3CL1 negated the beneficial impact of AA on the polarization of macrophages and the phenotypic transformation of VSMCs. Crucially, our findings revealed that AA is capable of downregulating CX3CL1 expression, accomplishing this by obstructing the nuclear translocation of NF-κB p65. The findings indicate that AA holds promise as a prospective treatment for adverse aortic remodeling by suppressing the activity of NF-κB p65/CX3CL1 signaling pathway.
Asunto(s)
Disección Aórtica , Quimiocina CX3CL1 , Ratones Endogámicos C57BL , Triterpenos Pentacíclicos , Transducción de Señal , Factor de Transcripción ReIA , Remodelación Vascular , Animales , Ratones , Masculino , Disección Aórtica/metabolismo , Disección Aórtica/patología , Disección Aórtica/tratamiento farmacológico , Triterpenos Pentacíclicos/farmacología , Remodelación Vascular/efectos de los fármacos , Células RAW 264.7 , Transducción de Señal/efectos de los fármacos , Factor de Transcripción ReIA/metabolismo , Quimiocina CX3CL1/metabolismo , Quimiocina CX3CL1/genética , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patología , Aminopropionitrilo/farmacología , Macrófagos/metabolismo , Macrófagos/efectos de los fármacos , Miocitos del Músculo Liso/metabolismo , Miocitos del Músculo Liso/efectos de los fármacosRESUMEN
BACKGROUND: ß-aminopropionitrile (BAPN) is a pharmacological inhibitor of LOX (lysyl oxidase) and LOXLs (LOX-like proteins). Administration of BAPN promotes aortopathies, although there is a paucity of data on experimental conditions to generate pathology. The objective of this study was to define experimental parameters and determine whether equivalent or variable aortopathies were generated throughout the aortic tree during BAPN administration in mice. METHODS: BAPN was administered in drinking water for a period ranging from 1 to 12 weeks. The impacts of BAPN were first assessed with regard to BAPN dose, and mouse strain, age, and sex. BAPN-induced aortic pathological characterization was conducted using histology and immunostaining. To investigate the mechanistic basis of regional heterogeneity, the ascending and descending thoracic aortas were harvested after 1 week of BAPN administration before the appearance of overt pathology. RESULTS: BAPN-induced aortic rupture predominantly occurred or originated in the descending thoracic aorta in young C57BL/6J or N mice. No apparent differences were found between male and female mice. For mice surviving 12 weeks of BAPN administration, profound dilatation was consistently observed in the ascending region, while there were more heterogeneous changes in the descending thoracic region. Pathological features were distinct between the ascending and descending thoracic regions. Aortic pathology in the ascending region was characterized by luminal dilatation and elastic fiber disruption throughout the media. The descending thoracic region frequently had dissections with false lumen formation, collagen deposition, and remodeling of the wall surrounding the false lumen. Cells surrounding the false lumen were predominantly positive for α-SMA (α-smooth muscle actin). One week of BAPN administration compromised contractile properties in both regions equivalently, and RNA sequencing did not show obvious differences between the 2 aortic regions in smooth muscle cell markers, cell proliferation markers, and extracellular components. CONCLUSIONS: BAPN-induced pathologies show distinct, heterogeneous features within and between ascending and descending aortic regions in mice.
Asunto(s)
Aminopropionitrilo , Aorta Torácica , Rotura de la Aorta , Modelos Animales de Enfermedad , Ratones Endogámicos C57BL , Animales , Aminopropionitrilo/toxicidad , Aminopropionitrilo/farmacología , Aorta Torácica/patología , Aorta Torácica/efectos de los fármacos , Aorta Torácica/metabolismo , Femenino , Masculino , Rotura de la Aorta/inducido químicamente , Rotura de la Aorta/patología , Rotura de la Aorta/metabolismo , Rotura de la Aorta/prevención & control , Ratones , Remodelación Vascular/efectos de los fármacos , Dilatación Patológica , Músculo Liso Vascular/patología , Músculo Liso Vascular/efectos de los fármacos , Músculo Liso Vascular/metabolismo , Factores de Edad , Factores de Tiempo , Factores Sexuales , Proliferación Celular/efectos de los fármacos , Proteína-Lisina 6-Oxidasa/metabolismoRESUMEN
Epilepsy-associated cognitive disorder (ECD), a prevalent comorbidity in epilepsy patients, has so far uncharacterized etiological origins. Our prior work revealed that lysyl oxidase (Lox) acted as a novel contributor of ferroptosis, a recently discovered cell death mode in the regulation of brain function. However, the role of Lox-mediated ferroptosis in ECD remains unknown. ECD mouse model was established 2 months later following a single injection of kainic acid (KA) for. After chronic treatment with KA, mice were treated with different doses (30â¯mg/kg, 100â¯mg/kg and 300â¯mg/kg) of Lox inhibitor BAPN. Additionally, hippocampal-specific Lox knockout mice was also constructed and employed to validate the role of Lox in ECD. Cognitive functions were assessed using novel object recognition test (NOR) and Morris water maze test (MWM). Protein expression of phosphorylated cAMP-response element binding (CREB), a well-known molecular marker for evaluation of cognitive performance, was also detected by Western blot. The protein distribution of Lox was analyzed by immunofluorescence. In KA-induced ECD mouse model, ferroptosis process was activated according to upregulation of 4-HNE protein and a previously discovered ferroptosis in our group, namely, Lox was remarkably increased. Pharmacological inhibition of Lox by BAPN at the dose of 100â¯mg/kg significantly increased the discrimination index following NOR test and decreased escape latency as well as augmented passing times within 60â¯s following MWM test in ECD mouse model. Additionally, deficiency of Lox in hippocampus also led to pronounced improvement of deficits in ECD model. These findings indicate that the ferroptosis regulatory factor, Lox, is activated in ECD. Ablation of Lox by either pharmacological intervention or genetic manipulation ameliorates the impairment in ECD mouse model, which suggest that Lox serves as a promising therapeutic target for treating ECD in clinic.
Asunto(s)
Disfunción Cognitiva , Epilepsia , Humanos , Ratones , Animales , Proteína-Lisina 6-Oxidasa/genética , Proteína-Lisina 6-Oxidasa/metabolismo , Aminopropionitrilo/farmacología , Regulación de la Expresión Génica , Modelos Animales de Enfermedad , Disfunción Cognitiva/tratamiento farmacológicoRESUMEN
BACKGROUND: Thoracic aortic dissection (TAD) is one of the most fatal cardiovascular diseases. One of its important pathological characteristics is the local inflammatory response. Many studies have found that Macrophage polarization plays an extremely critical role in the inflammatory progression and tissue remodeling of TAD. Costunolide (CTD) has an improving effect on oxidative stress and inflammation in the body. However, whether it can promote the integrity of extracellular matrix in Aortic dissection and its mechanism are still unclear. METHODS: The male C57BL/6J mice were used to construct an animal model of TAD with ß-aminopropionitrile (BAPN) (100 mg/kg/day, lasting for 28 days), and then CTD (10 mg/kg or 100 mg/kg) was injected intraperitoneally for 28 days to check the survival rate, TAD incidence, aortic morphology and other indicators of the mice. Using hematoxylin-eosin (HE), Masson, Elastin van Gieson (EVG) staining, immunofluorescence (IF), and immunohistochemical staining, the study aimed to determine the therapeutic effects of CTD on an animal model with BAPN-induced TAD. To enhance the examination of the regulatory mechanism of CTD, we conducted transcriptome sequencing on arterial tissues of mice in both the BAPN group and the BAPN + CTD100 group. Next, ANG II were used to construct TAD model in vascular smooth muscle cells (VMSCs). The effects of CTD on the proliferation, migration, invasion, and apoptosis of ANG II-induced cells are to be detected. The expression of MMP2, MMP9, P65, and p-P65 in each group will be examined using Western blot. Finally, the overexpression of IκB kinaseß (IKKß) will be established in VMSCs cells to further explore the protective function of CTD. RESULTS: The result showed that CTD significantly inhibited BAPN induced mortality and TAD incidence in the animal model, improved aortic vascular morphology, promoted the integrity of extracellular matrix in TAD, reduced tissue inflammation, reduced the accumulation of M1 macrophage, promoted M2 macrophage polarization, and reduced the expression of NF-κB pathway related proteins. Mechanistically, CTD significantly weakened the proliferation, migration, invasion, and apoptosis. p-P65 protein expression of TAD cells were induced by ANG II and IKK-ß. CONCLUSION: CTD has the potential to alleviate inflammation, VSMC apoptosis, MMP2/9 levels, and enhance extracellular matrix integrity in TAD by inhibiting the NF-κB signaling pathway.
Asunto(s)
Disección Aórtica , Disección de la Aorta Torácica , Sesquiterpenos , Masculino , Ratones , Animales , FN-kappa B/metabolismo , Metaloproteinasa 2 de la Matriz/metabolismo , Aminopropionitrilo/uso terapéutico , Aminopropionitrilo/farmacología , Ratones Endogámicos C57BL , Disección Aórtica/tratamiento farmacológico , Transducción de Señal , Inflamación/tratamiento farmacológico , Modelos Animales de EnfermedadRESUMEN
Two major constituents of exfoliation material, fibrillin-1 and lysyl oxidase-like 1 (encoded by FBN1 and LOXL1), are implicated in exfoliation glaucoma, yet their individual contributions to ocular phenotype are minor. To test the hypothesis that a combination of FBN1 mutation and LOXL1 deficiency exacerbates ocular phenotypes, the pan-lysyl oxidase inhibitor ß-aminopropionitrile (BAPN) was used to treat adult wild-type (WT) mice and mice heterozygous for a missense mutation in Fbn1 (Fbn1C1041G/+) for 8 weeks and their eyes were examined. Although intraocular pressure did not change and exfoliation material was not detected in the eyes, BAPN treatment worsened optic nerve and axon expansion in Fbn1C1041G/+ mice, an early sign of axonal damage in rodent models of glaucoma. Disruption of elastic fibers was detected only in Fbn1C1041G/+ mice, which increased with BAPN treatment, as shown by histologic and immunohistochemical staining of the optic nerve pia mater. Transmission electron microscopy showed that Fbn1C1041G/+ mice had fewer microfibrils, smaller elastin cores, and a lower density of elastic fibers compared with WT mice in control groups. BAPN treatment led to elastin core expansion in both WT and Fbn1C1041G/+ mice, but an increase in the density of elastic fiber was confined to Fbn1C1041G/+ mice. LOX inhibition had a stronger effect on optic nerve and elastic fiber parameters in the context of Fbn1 mutation, indicating the Marfan mouse model with LOX inhibition warrants further investigation for exfoliation glaucoma pathogenesis.
Asunto(s)
Aminopropionitrilo , Modelos Animales de Enfermedad , Fibrilina-1 , Síndrome de Marfan , Nervio Óptico , Proteína-Lisina 6-Oxidasa , Animales , Ratones , Adipoquinas , Aminoácido Oxidorreductasas/metabolismo , Aminoácido Oxidorreductasas/antagonistas & inhibidores , Aminoácido Oxidorreductasas/genética , Aminopropionitrilo/farmacología , Tejido Elástico/patología , Tejido Elástico/metabolismo , Tejido Elástico/ultraestructura , Fibrilina-1/genética , Fibrilinas/metabolismo , Glaucoma/patología , Presión Intraocular , Síndrome de Marfan/patología , Síndrome de Marfan/complicaciones , Ratones Endogámicos C57BL , Proteínas de Microfilamentos/metabolismo , Nervio Óptico/patología , Nervio Óptico/ultraestructura , Nervio Óptico/efectos de los fármacos , Proteína-Lisina 6-Oxidasa/metabolismo , Proteína-Lisina 6-Oxidasa/antagonistas & inhibidoresRESUMEN
Acute aortic dissection (AAD) progresses rapidly and is associated with high mortality; therefore, there remains an urgent need for pharmacological agents that can protect against AAD. Herein, we examined the therapeutic effects of cannabidiol (CBD) in AAD by establishing a suitable mouse model. In addition, we performed human AAD single-cell RNA sequencing and mouse AAD bulk RNA sequencing to elucidate the potential underlying mechanism of CBD. Pathological assays and in vitro studies were performed to verify the results of the bioinformatic analysis and explore the pharmacological function of CBD. In a ß-aminopropionitrile (BAPN)-induced AAD mouse model, CBD reduced AAD-associated morbidity and mortality, alleviated abnormal enlargement of the ascending aorta and aortic arch, and suppressed macrophage infiltration and vascular smooth muscle cell (VSMC) apoptosis. Bioinformatic analysis revealed that the pro-apoptotic gene PMAIP1 was highly expressed in human and mouse AAD samples, and CBD could inhibit Pmaip1 expression in AAD mice. Using human aortic VSMCs (HAVSMCs) co-cultured with M1 macrophages, we revealed that CBD alleviated HAVSMCs mitochondrial-dependent apoptosis by suppressing the BAPN-induced overexpression of PMAIP1 in M1 macrophages. PMAIP1 potentially mediates HAVSMCs apoptosis by regulating Bax and Bcl2 expression. Accordingly, CBD reduced AAD-associated morbidity and mortality and mitigated the progression of AAD in a mouse model. The CBD-induced effects were potentially mediated by suppressing macrophage infiltration and PMAIP1 (primarily expressed in macrophages)-induced VSMC apoptosis. Our findings offer novel insights into M1 macrophages and HAVSMCs interaction during AAD progression, highlighting the potential of CBD as a therapeutic candidate for AAD treatment.
Asunto(s)
Disección Aórtica , Cannabidiol , Animales , Humanos , Ratones , Aminopropionitrilo/farmacología , Disección Aórtica/tratamiento farmacológico , Apoptosis , Proteínas Reguladoras de la Apoptosis/metabolismo , Cannabidiol/farmacología , Cannabidiol/metabolismo , Macrófagos/metabolismo , Músculo Liso Vascular/patologíaRESUMEN
Aortic dissection (AD) is the most catastrophic vascular disease with a high mortality rate. Trimethylamine N-oxide (TMAO), a gut microbial metabolite, has been implicated in the pathogenesis of cardiovascular diseases. However, the role of TMAO in AD and the underlying mechanisms remain unclear. This study aimed to explore the effects of TMAO on AD. Plasma and fecal samples from patients with AD and healthy individuals were collected to analyze TMAO levels and gut microbial species, respectively. The plasma levels of TMAO were significantly higher in 253 AD patients compared with those in 98 healthy subjects (3.47, interquartile range (IQR): 2.33 to 5.18 µM vs. 1.85, IQR: 1.40 to 3.35 µM; p < 0.001). High plasma TMAO levels were positively associated with AD severity. An increase in the relative abundance of TMA-producing genera in patients with AD was revealed using 16S rRNA sequencing. In the angiotensin II or ß-aminopropionitrile-induced rodent model of AD, mice fed a TMAO-supplemented diet were more likely to develop AD compared to mice fed a normal diet. Conversely, TMAO depletion mitigated AD formation in the BAPN model. RNA sequencing of aortic endothelial cells isolated from mice administered TMAO revealed significant upregulation of genes involved in inflammatory pathways. The in vitro experiments verified that TMAO promotes endothelial dysfunction and activates nuclear factor (NF)-κB signaling. The in vivo BAPN-induced AD model confirmed that TMAO increased aortic inflammation. Our study demonstrates that the gut microbial metabolite TMAO aggravates the development of AD at least in part by inducing endothelial dysfunction and inflammation. This study provides new insights into the etiology of AD and ideas for its management.
Asunto(s)
Disección Aórtica , Microbioma Gastrointestinal , Metilaminas , Humanos , Ratones , Animales , Microbioma Gastrointestinal/fisiología , ARN Ribosómico 16S , Aminopropionitrilo , Células Endoteliales , Inflamación , Disección Aórtica/etiologíaRESUMEN
Thoracic aortic aneurysm and dissection (TAAD) is a life-threatening disease and currently there is no pharmacological therapy. Sympathetic nerve overactivity plays an important role in the development of TAAD. Sympathetic innervation is mainly controlled by nerve growth factor (NGF, a key neural chemoattractant) and semaphoring 3A (Sema3A, a key neural chemorepellent), while the roles of these two factors in aortic sympathetic innervation and especially TAAD are unknown. We hypothesized that genetically manipulating the NGF/Sema3A ratio by the Ngf -driven Sema3a expression approach may reduce aortic sympathetic nerve innervation and mitigate TAAD progression. A mouse strain of Ngf gene-driven Sema3a expression (namely NgfSema3a/Sema3a mouse) was established by inserting the 2A-Sema3A expression frame to the Ngf terminating codon using CRISPR/Cas9 technology. TAAD was induced by ß-aminopropionitrile monofumarate (BAPN) both in NgfSema3a/Sema3a mice and wild type (WT) littermates. Contrary to our expectation, the BAPN-induced TAAD was severer in NgfSema3a/Sema3a mice than in wild-type (WT) mice. In addition, NgfSema3a/Sema3a mice showed higher aortic sympathetic innervation, inflammation and extracellular matrix degradation than the WT mice after BAPN treatment. The aortic vascular smooth muscle cells isolated from NgfSema3a/Sema3a mice and pretreated with BAPN in vivo for two weeks showed stronger capabilities of proliferation and migration than that from the WT mice. We conclude that the strategy of Ngf -driven Sema3a expression cannot suppress but worsens the BAPN-induced TAAD. By investigating the aortic phenotype of NgfSema3a/Sema3a mouse strain, we unexpectedly find a path to exacerbate BAPN-induced TAAD which might be useful in future TAAD studies.
Asunto(s)
Aneurisma de la Aorta Torácica , Disección Aórtica , Azidas , Desoxiglucosa , Animales , Ratones , Aminopropionitrilo/efectos adversos , Aneurisma de la Aorta Torácica/genética , Aneurisma de la Aorta Torácica/inducido químicamente , Aneurisma de la Aorta Torácica/metabolismo , Desoxiglucosa/análogos & derivados , Modelos Animales de Enfermedad , Factor de Crecimiento Nervioso/genética , Factor de Crecimiento Nervioso/efectos adversos , Semaforina-3A/genéticaRESUMEN
Lysyl oxidases (LOX(L)) are enzymes that catalyze the formation of cross-links in collagen and elastin fibers during physiologic calcification of bone. However, it remains unknown whether they may promote pathologic calcification of articular cartilage, an important hallmark of debilitating arthropathies. Here, we have studied the possible roles of LOX(L) in cartilage calcification, related and not related to their cross-linking activity. We first demonstrated that inhibition of LOX(L) by ß-aminoproprionitrile (BAPN) significantly reduced calcification in murine and human chondrocytes, and in joint of meniscectomized mice. These BAPN's effects on calcification were accounted for by different LOX(L) roles. Firstly, reduced LOX(L)-mediated extracellular matrix cross-links downregulated Anx5, Pit1 and Pit2 calcification genes. Secondly, BAPN reduced collagen fibrotic markers Col1 and Col3. Additionally, LOX(L) inhibition blocked chondrocytes hypertrophic differentiation (Runx2 and COL10), pro-inflammatory IL-6 release and reactive oxygen species (ROS) production, all triggers of chondrocyte calcification. Through unbiased transcriptomic analysis we confirmed a positive correlation between LOX(L) genes and genes for calcification, hypertrophy and extracellular matrix catabolism. This association was conserved throughout species (mouse, human) and tissues that can undergo pathologic calcification (kidney, arteries, skin). Overall, LOX(L) play a critical role in the process of chondrocyte calcification and may be therapeutic targets to treat cartilage calcification in arthropathies.
Asunto(s)
Calcinosis , Cartílago Articular , Artropatías , Ratones , Humanos , Animales , Proteína-Lisina 6-Oxidasa/metabolismo , Aminopropionitrilo , Colágeno/metabolismo , Calcinosis/patología , Condrocitos/metabolismo , Hipertrofia , Cartílago Articular/metabolismoRESUMEN
BACKGROUND: Thoracic aortic aneurysm (TAA) is a silent but life-threatening cardiovascular disease. Heme oxygenase 1 (HO-1) plays an important role in the cardiovascular diseases but is poorly understood in TAA. This study aims at investigating the role of HO-1 in TAA. METHODS: Single-cell RNA sequencing, Western blot and histological assay were performed to identify specific cellular expression of HO-1 in both human and ß-aminopropionitrile (BAPN)-induced mice TAA. Zinc protoporphyrin (ZnPP), a pharmacological inhibitor of HO-1, was used to investigate whether inhibition of HO-1 could attenuate BAPN-induced TAA in rodent model. Histological assay, Western blot assay, and mRNA sequencing were further performed to explore the underlying mechanisms. RESULTS: Single-cell transcriptomic analyses of 113,800 thoracic aortic cells identified an increase of HO-1(+) macrophage in aneurysmal thoracic aorta from BAPN-induced TAA mice and TAA patients. Histological assay verified HO-1 overexpression in clinical TAA specimens, which was co-localized with CD68(+) macrophage. HO-1(+) macrophage was closely associated with pro-inflammatory response and immune activation. Inhibition of HO-1 through ZnPP significantly alleviated BAPN-induced TAA in mice and restored extracellular matrix (ECM) in vivo. Further experiments showed that ZnPP treatment suppressed the expression of matrix metalloproteinases (MMPs) in aneurysmal thoracic aortic tissues from BAPN-induced TAA mice, including MMP2 and MMP9. Macrophages from myeloid specific HO-1 knockout mice displayed weakened pro-inflammatory activity and ECM degradation capability. CONCLUSION: HO-1(+) macrophage subgroup is a typical hallmark of TAA. Inhibition of HO-1 through ZnPP alleviates BAPN-induced TAA in mice, which might work through restoration of ECM via suppressing MMP2 and MMP9 expression.
Asunto(s)
Aneurisma de la Aorta Torácica , Metaloproteinasa 2 de la Matriz , Animales , Humanos , Ratones , Aminopropionitrilo/efectos adversos , Aminopropionitrilo/metabolismo , Aorta Torácica/metabolismo , Aneurisma de la Aorta Torácica/genética , Modelos Animales de Enfermedad , Matriz Extracelular/metabolismo , Hemo-Oxigenasa 1/metabolismo , Metaloproteinasa 2 de la Matriz/metabolismo , Metaloproteinasa 9 de la Matriz/metabolismo , Ratones NoqueadosRESUMEN
Thoracic aortic dissection (TAD) is one of the most lethal aortic diseases due to its acute onset, rapid progress, and high rate of aortic rupture. The pathogenesis of TAD is not completely understood. In this mini-review, we introduce three emerging experimental mouse TAD models using β-aminopropionitrile (BAPN) alone, BAPN for a prolonged duration (four weeks) and then with added infusion of angiotensin II (AngII), or co-administration of BAPN and AngII chronically. We aim to provide insights into appropriate application of these three mouse models, thereby enhancing the understanding of the molecular mechanisms of TAD.
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Animales , Masculino , Ratones , Aminopropionitrilo/toxicidad , Disección Aórtica/patología , Angiotensina II/toxicidad , Aneurisma de la Aorta Torácica/patología , Modelos Animales de Enfermedad , Ratones Endogámicos C57BLRESUMEN
Objective: To investigate the protective role of alprostadil on aortic dissection. Methods: 26 C57BL6 male mice were divided into control group (normal drinking water, n=13) and model group (1 g·kg-1·d-1 BAPN via drinking water, n=13). On day 14, mRNA expression of inflammatory-related genes as well as EP receptor families were detected by RT-PCR (n=6 each) and EP4 protein levels were determined by Western blot (n=7 each). Another 88 mice were divided into 3 groups: control group (n=22), model group (n=33) and treatment group (n=33). The mice in model group and treatment group were applied with BAPN (1 g·kg-1·d-1) via drinking water. The mice in treatment group received additional intraperitoneal injection with alprostadil (80 μg·kg-1·d-1) for 28 days. The mice in the control and model group received equal volume intraperitoneal injection with 0.9% saline respectively. The body weight and systolic blood pressure, the mortality and morbidity were monitored from the beginning until the designed end of the study. On day 28, the mice were sacrificed and aorta were fixed, embedded and sliced, followed by staining with HE and Victoria Blue. The distribution of EP4 was determined by immunohistochemistry in control (n=6) and model group (n=6). Furthermore, the concentration of PGE1 were tested among model (n=3) and treatment group (n=4). EP4 protein expression was determined in model group (n=7) and treatment group (n=6). Results: On day 14, mRNA expression level of MCP-1 ((2.74±1.55) vs. (1.00±0.49),<0.05) and MMP2((1.38±0.42) vs. (1.00±0.27), P<0.05) was significantly upregulated in model group compared with control group. Protein expression of EP4 receptor also increased in aorta in model group compared with control group (1.48±0.51 vs. 1.00±0.19, P<0.05). In the dissection area, the EP4 expression was also enriched compared with non-dissection area, particularly in endothelial cells and inflammatory cells on day 28. BAPN applied in drinking water (model and treatment groups) successfully induced the aortic dissection in mice, some mice died of the rupture. The elastic fibers were fractured, and the infiltrated immune cells were visible in dissected tissue. False lumen was formed. There was no dissection and death in the control group. Compared with control group, the morbidity and mortality rates were significantly increased in the model group (60.6%, 20/33, 30.3%, 10/33) and the treatment group (72.7%, 24/33, 24.2%, 8/33). The mortality and morbidity rates were similar between model and treatment groups. There is no difference in terms of SBP among three groups (P>0.05). Further study showed that after alprostadil injection, the blood concentration of PGE1 was increased in treatment group ((0.540±0.041 vs. 0.436±0.012)μmol/L, P<0.05). Besides, the EP4 receptor expression was downregulated in the treatment group compared to model group (0.60±0.30 vs. 1.00±0.20, P<0.05). Conclusion: EP4 expression is upregulated in BAPN induced aortic dissection mouse model. No protective effects are observed post alprostadil treatment in this model probably due to the reduced expression of EP4.
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Animales , Masculino , Ratones , Alprostadil , Aminopropionitrilo , Disección Aórtica , Modelos Animales de Enfermedad , Células EndotelialesRESUMEN
<p><b>OBJECTIVE</b>To investigate thoracic aortic longitudinal elastic strength in β-aminopropionitrile (BAPN) treated rat model of aortic dissection (AD).</p><p><b>METHODS</b>Twenty-nine young rats (Sprague-Dawley) were divided into tow groups, control group (n = 12) and BAPN group (n = 17). Seventeen rats were treated with 0.25% BAPN mixed in feed for 6 weeks. All the rats were sacrificed in the end of experiment and aorta was harvested for biomechanical and pathological study. Longitudinal elastic strength and stress were detected and analyzed by material testing machine. Elasticity modulus as well as maximum stretching length, draw ratio, maximum load, maximum strength, and maximum extensibility was calculated according to the analysis with thickness and area of aortic media.</p><p><b>RESULTS</b>Nine BAPN-treated rats died of aortic dissecting aneurysm rupture during the experiment. The diameter of the aneurysms was (6.33 ± 1.17) mm and the length was (9 ± 5) mm. The maximum diameter significantly increased in BAPN-induced rats with AD (group B2) compared with without AD (group B1) and control group ((6.49 ± 1.20) mm vs. (1.45 ± 0.11), (1.25 ± 0.26); F = 165.257, P = 0.001 and 0.000, respectively), but there was no significance between group B1 and control group (P = 0.108). Thickness and area of aortic media in BAPN-induced rats significantly increased compared with control group (F = 27.277 and 27.153, P = 0.000 and 0.000, respectively), but there was no significance of area between group B1 and B2 (P = 0.540). Maximum stretching length, draw ratio, maximum load, maximum strength maximum extensibility and elasticity modulus were significantly decreased from group B2, group B1 to control group (P < 0.01, respectively).</p><p><b>CONCLUSIONS</b>This study built a successful model of AD. Biomechanical analysis and the decrease of maximum stretching length, draw ratio, maximum load, maximum strength maximum extensibility and elasticity modulus may explain the formation of AD partly.</p>
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Animales , Masculino , Ratas , Aminopropionitrilo , Farmacología , Disección Aórtica , Aorta , Patología , Fenómenos Biomecánicos , Modelos Animales de Enfermedad , Módulo de Elasticidad , Ratas Sprague-DawleyRESUMEN
<p><b>OBJECTIVE</b>To study the effects of lysyl oxidase (LOX) on the migration and adhesion of the human gastric cancer cell line HGC-27 cells in vitro.</p><p><b>METHODS</b>The human gastric cancer cell line HGC-27 cells were cultured in vitro, and treated with different concentration of β-aminopropionitrile (BAPN). The ability of migration was assessed by wound-healing assay. The ability of adhesion was detected by homogenous and heterogeneous adhesion experiments.</p><p><b>RESULTS</b>Compared that with 0 mmol/L BAPN, the ability of migration of the cells after treatment with 0.2 mmol/L BAPN was descended at 8, 24, 32 and 48 h; the number of cells with homogeneous adhesion was increased from (6.97 ± 0.07) × 10(3)/ml to (7.78 ± 0.11) × 10(3)/ml; and the number of cells with heterogeneous adhesion was decreased from (8.98 ± 0.15) × 10(3)/ml to (8.35 ± 0.10) × 10(3)/ml, both < 0.05. Compared with that of cells treated with 0 mmol/L and 0.2 mmol/L BAPN, the migration ability of cells after treatment with 0.3 mmol/L BAPN was descended at 8, 24, 32 and 48 h; the number of cells with homogeneous adhesion was raised to (8.02 ± 0.11) × 10(3)/ml and the number of cells with heterogeneous adhesion was down to (7.93 ± 0.07) × 10(3)/ml (P < 0.05).</p><p><b>CONCLUSION</b>LOX may promote the metastasis of cancer cells by enhancing invasion, increasing heterogeneous adhesion and decreasing homogeneous adhesion.</p>
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Humanos , Aminopropionitrilo , Farmacología , Adhesión Celular , Línea Celular Tumoral , Movimiento Celular , Relación Dosis-Respuesta a Droga , Invasividad Neoplásica , Proteína-Lisina 6-Oxidasa , Metabolismo , Fisiología , Neoplasias Gástricas , PatologíaRESUMEN
PURPOSE: To compare the CT findings of tuberculous pneumonia (TBPN) with those of bacterial pneumonia (BAPN) in diabetic patients and to evaluate the usefulness of CT in the differential diagnosis of these two diseases. MATERIALS AND METHODS: The chest CT scans of 23 diabetic patients with TBPN (M:F=21:2; mean age, 59 yrs.) and of 37 diabetic patients with BAPN (M:F=21:16; mean age, 63 yrs.) were evaluated by two radiologists with regard to low attenuation areas in regions of consolidation, cavities, air bronchogram, volume changes, ground-glass attenuation, findings of bronchogenic spread, and other associated findings. The involvement of each segment was recorded in all patients. RESULTS:The frequencies of multiple small low-attenuation areas in regions of consolidation (52%, 0%), multiple cavities (35%, 3%), loss of volume (70%, 30%) and findings of bronchogenic spread (96%, 30%) were significantly higher in TBPN than in BAPN (p<.05). Low-attenuation masses and bizarre-shaped cavities were noted only in TBPN. Large areas of ground-glass attenuation (4%, 38%) and bilateral pleural effusions (0%, 19%) were more common in BAPN, while air-bronchogram was common in both groups (96%, 86%). The involvement of the superior segment was significantly more common in TBPN (p<.05). CONCLUSION: In the diabetic patients with pulmonary consolidation, CT findings of multiple small low-attenuation areas, multiple cavities, bizarre-shaped cavities, low attenuation masses in cavities, volume loss, and findings of bronchogenic spread are more suggestive of TBPN, while large areas of ground-glass attenuation and bilateral pleural effusions are more suggestive of BAPN. CT may be useful in the differential diagnosis between TBPN and BAPN.
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Humanos , Aminopropionitrilo , Diabetes Mellitus , Diagnóstico Diferencial , Derrame Pleural , Neumonía , Neumonía Bacteriana , Tomografía Computarizada por Rayos X , Tuberculosis PulmonarRESUMEN
The purpose of this study is to evaluate the influences of the bone morphogenetic protein (BMP) on the healing of periodontal ligament and alveolar bone after replantation of tooth, and to examine the possibility of its clinical application. 45 Sprague Dawley rats weighted about 100 gram were divided into 3 experimental groups by different dose of BMP. All the upper right and left 1st molar were extracted after 5 days feeding of 0.4% beta-aminopropionitrile, and right molar were used as experimental group and left molar were used as control group. The root surface of experimental molar were treated with 25, 50 and 100ng/ml of human recombinant Bone morphogenetic protein-4 (rh-BMP-4) with micropipet, and 1M Sodium hypochloride were used on control root surface. All the experimental animals were sacrificed as 1, 2, 4, 7 and 14 days after autoreplantation of upper 1st molar into their own position. The maxilla were disected included both side of 1st molar. The collected tissue were processed from demineralization to paraffin embeding as usual procedure, and the specimens were prepared with Hematoxylin-Eosin stain for the light microscopic evaluation. The results of this study were as follows : 1. There was no significant differences between control and experimental site on 1 and 2 days after replantation of tooth. In the case of 4th days, the evidence of tissue regeneration were observed on experimental site to compare the controls. New osteoid were revealed on high concentration of BMP at 7 days after replantation, and it became more obvious at 14 days. 2. The effect of the rh-BMP-4 coated on root surface was revealed slight influences for the prolifertion of cells of periodontium and tissue regeneration as dose-dependent pattern. 3. Bony ankylosis was observed between alveolar bone and root surface due to the remarkable amount of osteoid formation on the 14 days after replantation of root. In the conclusion, it was suggested that topical application of the rhBMP-4 on the root surface has influence on the periodontal ligament and alveolar bone. The application method of BMP on the root should be designed with calculation of proper concentration.
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Animales , Humanos , Ratas , Aminopropionitrilo , Anquilosis , Proteínas Morfogenéticas Óseas , Maxilar , Diente Molar , Parafina , Ligamento Periodontal , Periodoncio , Ratas Sprague-Dawley , Regeneración , Reimplantación , Sodio , Reimplante Dental , DienteRESUMEN
Cleft palate has been studied with epidemiologic and molecular methods, and many etiologic factors have been examined closely. Among the research methods, biologic molecule research has been the most important method for cleft palate formation study. The TGF-beta played an important role in cell migration, epithelial-mesenchymal transdifferentiation, extracellular matrix synthesis and deposition. But there was not much research on the correlation cleft palate induced by beta-aminonitroproprionitrile(BAPN) and TGF-beta expression. The purpose of the present study was to examine how TGF-beta is expressed in cleft palate rats. 4 Timed-pregnant Sprague-Dawley rats were obtained on the 10th gestation day. On the 13th day of gestation, BAPN-monofumarate salts ((C3H6N2)2.C4H4O4) were individually, orally administered to 3 pregnant rats at a ratio of 1g/kg body weight. And 4 pregnant rats were sacrificed on day 20 post coitus (p.c.). The TGF-beta expression in the cleft formed rats fetuses showed the following patterns : 1. Osteoblast and mesenchymal cells of the cleft palate rats were of low expression compared with those of the control rats. 2. The cleft palate rats didn't show any difference in the TGF-beta expression of osteocyte from the control rats. 3. In western blot analysis, the thickness of band of TGF-beta in the cleft palate rats was thinner and more diluted than that of the control rats.