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1.
Food Chem ; 338: 128020, 2021 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-32932087

RESUMO

Plant-based protein foods are increasingly common, but data on their nutritional protein quality are scarce. This study evaluated it for seitan (wheat-based food), tofu (soya-based food), soya milk, and a pea emulsion. The true ileal digestibility (TID) of their amino acids was determined in minipigs, to calculate the digestible indispensable amino acid score (DIAAS). The TID of the proteins was high and not significantly different between the foods tested: 97% for seitan, 95% for tofu, 92% for soya milk and 94% for pea emulsion. There were only minor differences in individual amino acid TIDs. DIAAS ranking was thus essentially driven by the amino acid composition of the food: soya-based food > pea emulsion > seitan. Nevertheless, the lower TID of sulphur-containing amino acids in tofu than in soya milk induced a significant decrease in DIAAS (from 117% to 97%), highlighting the importance of the matrix effect on nutritional protein quality.


Assuntos
Aminoácidos/análise , Proteínas na Dieta/farmacocinética , Íleo/metabolismo , Proteínas de Plantas/farmacocinética , Aminoácidos/metabolismo , Aminoácidos Essenciais/análise , Aminoácidos Essenciais/metabolismo , Animais , Digestão , Íleo/efeitos dos fármacos , Valor Nutritivo , Proteínas de Plantas/metabolismo , Alimentos de Soja , Leite de Soja , Soja/química , Suínos , Porco Miniatura , Triticum/química
2.
Cell Prolif ; 53(10): e12863, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32871045

RESUMO

OBJECTIVES: Immunodeficient mice injected with human cancer cell lines have been used for human oncology studies and anti-cancer drug trials for several decades. However, rodents are not ideal species for modelling human cancer because rodents are physiologically dissimilar to humans. Therefore, anti-tumour drugs tested effective in rodents have a failure rate of 90% or higher in phase III clinical trials. Pigs are similar to humans in size, anatomy, physiology and drug metabolism rate, rendering them a desirable pre-clinical animal model for assessing anti-cancer drugs. However, xenogeneic immune rejection is a major barrier to the use of pigs as hosts for human tumours. Interleukin (IL)-2 receptor γ (IL2RG), a common signalling subunit for multiple immune cytokines including IL-2, IL-4, IL-7, IL-9, IL-15 and IL-21, is required for proper lymphoid development. MATERIALS AND METHODS: IL2RG-/Y pigs were generated by CRISPR/Cas9 technology, and examined for immunodeficiency and ability to support human oncogenesis. RESULTS: Compared to age-matched wild-type pigs, IL2RG-/Y pigs exhibited a severely impaired immune system as shown by lymphopenia, lymphoid organ atrophy, poor immunoglobulin function, and T- and NK-cell deficiency. Human melanoma Mel888 cells generated tumours in IL2RG-/Y pigs but not in wild-type littermates. The human tumours grew faster in IL2RG-/Y pigs than in nude mice. CONCLUSIONS: Our results indicate that these pigs are promising hosts for modelling human cancer in vivo, which may aid in the discovery and development of anti-cancer drugs.


Assuntos
Sistemas CRISPR-Cas/genética , Subunidade gama Comum de Receptores de Interleucina/metabolismo , Neoplasias Cutâneas/patologia , Animais , Animais Geneticamente Modificados/metabolismo , Linhagem Celular Tumoral , Modelos Animais de Doenças , Edição de Genes , Humanos , Sistema Imunitário/metabolismo , Subunidade gama Comum de Receptores de Interleucina/antagonistas & inibidores , Subunidade gama Comum de Receptores de Interleucina/genética , Linfopenia/patologia , Melanoma/metabolismo , Melanoma/patologia , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Neoplasias Cutâneas/metabolismo , Neoplasias Cutâneas/mortalidade , Taxa de Sobrevida , Suínos , Porco Miniatura , Transplante Heterólogo
3.
Plast Reconstr Surg ; 146(4): 792-798, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32970001

RESUMO

BACKGROUND: Tissue expansion relies on the ability of skin to grow in response to sustained mechanical strain. This study focuses on correlation of cellular and histologic changes with skin growth and deformation during tissue expansion. METHODS: Tissue expanders were placed underneath the skin of five Yucatan minipigs and inflated with one fill of 60 cc of saline 1 hour, 24 hours, 3 days, and 7 days before the animals were killed, or two fills of either 30 cc or 60 cc at 10 and 3 days or 14 and 7 days before the animals were killed. Skin biopsy specimens and three-dimensional photographs were used to calculate skin growth and stretch according to the authors' novel finite element analysis model. RESULTS: The mitotic index of keratinocytes in the basal layer increased 1 hour after stimulus was applied (4 percent) (p = 0.022), peaked at approximately day 3 (26 percent) (p < 0.0001), and tapered by day 7 (12.5 percent) (p = 0.012) after tissue expansion. The authors demonstrated that it is the volume per fill rather than the total volume in the expander that scales the magnitude of response. Lastly, the authors demonstrated that the ratio of deformation attributable to growth versus stretch (Fgrowth/Fstretch) after 60 cc of tissue expansion fill was 1.03 at 1 hour, 0.82 at 1 day, 0.85 at day 3, and 0.95 at 7 days. CONCLUSIONS: Peak cell proliferation occurred 3 days after tissue expansion fill and is scaled in response to stimulus magnitude. The growth component of deformation equilibrates to the stretch component at day 7, as cell proliferation has started to translate to skin growth.


Assuntos
Modelos Estatísticos , Pele/crescimento & desenvolvimento , Expansão de Tecido/métodos , Animais , Feminino , Modelos Animais , Tamanho do Órgão , Pele/anatomia & histologia , Suínos , Porco Miniatura , Fatores de Tempo
4.
Am J Physiol Heart Circ Physiol ; 319(4): H915-H926, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-32857599

RESUMO

We have previously reported enhanced Ca2+ sensitivity of coronary arteries that is dependent upon collateral circulation for their blood supply. For the current study, we hypothesized that small collateral-dependent arteries would exhibit an enhanced KCl-mediated contractile response attributable to Ca2+ sensitization and increased Ca2+ channel current. Ameroid constrictors were surgically placed around the left circumflex (LCX) artery of female Yucatan miniature swine. Eight weeks postoperatively, pigs were randomized into sedentary or exercise-trained (treadmill run; 5 days/wk; 14 wk) groups. Small coronary arteries (150-300 µm luminal diameter) were isolated from myocardial regions distal to the collateral-dependent LCX and the nonoccluded left anterior descending arteries. Contractile tension and simultaneous measures of both tension and intracellular free Ca2+ levels (fura-2) were measured in response to increasing concentrations of KCl. In addition, whole cell Ca2+ currents were also obtained. Chronic occlusion enhanced contractile responses to KCl and increased Ca2+ sensitization in collateral-dependent compared with nonoccluded arteries of both sedentary and exercise-trained pigs. In contrast, smooth muscle cell Ca2+ channel current was not altered by occlusion or exercise training. Ca2+/calmodulin-dependent protein kinase II (CaMKII; inhibited by KN-93, 0.3-1 µM) contributed to the enhanced contractile response in collateral-dependent arteries of sedentary pigs, whereas both CaMKII and Rho-kinase (inhibited by hydroxyfasudil, 30 µM or Y27632, 10 µM) contributed to increased contraction in exercise-trained animals. Taken together, these data suggest that chronic occlusion leads to enhanced contractile responses to KCl in collateral-dependent coronary arteries via increased Ca2+ sensitization, a response that is further augmented with exercise training.NEW & NOTEWORTHY Small coronary arteries distal to chronic occlusion displayed enhanced contractile responses, which were further augmented after exercise training and attributable to enhanced calcium sensitization without alterations in calcium channel current. The calcium sensitization mediators Rho-kinase and CaMKII significantly contributed to enhanced contraction in collateral-dependent arteries of exercise-trained, but not sedentary, pigs. Exercise-enhanced contractile responses may increase resting arterial tone, creating an enhanced coronary flow reserve that is accessible during periods of increased metabolic demand.


Assuntos
Sinalização do Cálcio/efeitos dos fármacos , Cálcio/metabolismo , Circulação Colateral/efeitos dos fármacos , Circulação Coronária/efeitos dos fármacos , Oclusão Coronária/fisiopatologia , Vasos Coronários/efeitos dos fármacos , Esforço Físico , Cloreto de Potássio/farmacologia , Vasoconstrição/efeitos dos fármacos , Adaptação Fisiológica , Animais , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Oclusão Coronária/metabolismo , Vasos Coronários/metabolismo , Vasos Coronários/fisiopatologia , Modelos Animais de Doenças , Feminino , Suínos , Porco Miniatura , Quinases Associadas a rho/metabolismo
5.
Clin Oral Implants Res ; 31(10): 1025-1036, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32790921

RESUMO

OBJECTIVE: To test the hypotheses of no differences in (I) percentage of bone (POB), non-mineralized tissue (NMT), and deproteinized bovine bone mineral (DBBM), and (II) ingrowth of mineralized bone after lateral guided bone regeneration (GBR) augmentation of the mandible with different ratios of DBBM and particulate autogenous bone (PAB) at different time points. MATERIAL AND METHODS: Twenty-four minipigs were randomly allocated into three groups. Lateral augmentation in 96 sites (4 in each animal) was performed unilaterally with a standardized quantity of grafting material in each animal with different ratios of DBBM and PAB (50:50, 75:25, 100:0) and autogenous bone block in combination with DBBM and covered with a collagen membrane. The percentage of different tissues in the graft and ingrowth of mineralized bone was assessed by histomorphometrical and histological analyses after 10, 20, and 30 weeks, respectively. RESULTS: The POB was 54% (50:50), 50% (75:25), and 48% (100:0) after 10 weeks, 60% (50:50), 61% (75:25), and 60% (100:0) after 20 weeks, and 63% (50:50), 62% (75:25), and 62% (100:0) after 30 weeks. There was no significant difference between the groups at any time points. There was a significant increase in POB and a significant decrease in NMT for 75:25 and 100:0 from 10 to 30 weeks. All ratios demonstrated a non-complete ingrowth of mineralized bone into the graft after 10 weeks and complete mineralization after 30 weeks. CONCLUSION: Within the limitations of the present study, it seems like addition of autogenous bone to DBBM for LRA did not affect the bone formation nor graft incorporation after 10-30 weeks of healing. However, a prolonged healing time seems to result in an increased POB for all ratios.


Assuntos
Aumento do Rebordo Alveolar , Substitutos Ósseos/farmacologia , Animais , Regeneração Óssea , Transplante Ósseo , Bovinos , Mandíbula/cirurgia , Minerais , Suínos , Porco Miniatura
6.
Cardiovasc Ther ; 2020: 3480276, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32565909

RESUMO

Wenxin Keli (WXKL) is a traditional Chinese medicine drug approved for the treatment of cardiovascular diseases. This study aimed to identify WXKL-targeting genes involved in antiarrhythmic efficacy of WXKL. The Traditional Chinese Medicine Systems Pharmacology (TCMSP) technology platform was used to screen active compounds of WXKL and WXKL-targeting arrhythmia-related genes. A pig model of myocardial ischemia (MI) was established by balloon-expanding the endothelium of the left coronary artery. Pigs were divided into the model group and WXKL group (n = 6). MI, QT interval, heart rate, and arrhythmia were recorded, and the mRNA expression of target genes in myocardial tissues was detected by PCR. Eleven active ingredients of WXKL and eight WXKL-targeting arrhythmia-related genes were screened. Five pathways were enriched, and an "ingredient-gene-path" network was constructed. WXKL markedly decreased the incidence of arrhythmia in the MI pig model (P < 0.05). The QT interval was significantly shortened, and the heart rate was slowed down in the WXKL group compared with the model group (P < 0.05). In addition, the expression of sodium channel protein type 5 subunit alpha (SCN5A) and beta-2 adrenergic receptor (ADRB2) was downregulated, while muscarinic acetylcholine receptor M2 (CHRM2) was upregulated in the WXKL group (P < 0.05). In conclusion, WXKL may shorten the QT interval and slow down the heart rate by downregulating SCN5A and ADRB2 and upregulating CHRM2 during MI. These findings provide novel insight into molecular mechanisms of WXKL in reducing the incidence of ventricular arrhythmia.


Assuntos
Potenciais de Ação/efeitos dos fármacos , Antiarrítmicos/farmacologia , Arritmias Cardíacas/prevenção & controle , Medicamentos de Ervas Chinesas/farmacologia , Frequência Cardíaca/efeitos dos fármacos , Isquemia Miocárdica/tratamento farmacológico , Potenciais de Ação/genética , Animais , Arritmias Cardíacas/genética , Arritmias Cardíacas/metabolismo , Arritmias Cardíacas/fisiopatologia , Modelos Animais de Doenças , Regulação da Expressão Gênica , Redes Reguladoras de Genes , Frequência Cardíaca/genética , Masculino , Medicina Tradicional Chinesa , Isquemia Miocárdica/genética , Isquemia Miocárdica/metabolismo , Isquemia Miocárdica/fisiopatologia , Canal de Sódio Disparado por Voltagem NAV1.5/genética , Canal de Sódio Disparado por Voltagem NAV1.5/metabolismo , Mapas de Interação de Proteínas , Receptor Muscarínico M2/genética , Receptor Muscarínico M2/metabolismo , Receptores Adrenérgicos beta 2/genética , Receptores Adrenérgicos beta 2/metabolismo , Suínos , Porco Miniatura , Fatores de Tempo
7.
PLoS One ; 15(6): e0234131, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32502216

RESUMO

BACKGROUND: Low plasma testosterone, either spontaneous or as a result of androgen deprivation therapy for prostate cancer, is associated with an increased risk of cardiovascular events. The underlying mechanism in humans is not understood. Experimental studies in mice have shown that castration facilitates atherogenesis and may increase signs of plaque vulnerability. Pigs used for translational atherosclerosis research have frequently been castrated for practical or commercial reasons, but the effect of castration on atherosclerosis has never been systematically evaluated in pigs. OBJECTIVE: To study the effect of castration on atherosclerotic plaque burden and type in genetically modified minipigs with hypercholesterolemia. METHODS: Newborn male Yucatan minipigs with transgenic overexpression of a human gain-of-function mutant of proprotein convertase subtilisin/kexin type 9 were randomized to undergo orchiectomy (n = 8) or serve as controls (n = 6). Minipigs were started on high-fat diet at 3 months of age and the amount and composition of atherosclerotic lesions were analyzed at 12 months of age. Plasma lipid profiles and behavioral parameters were also assessed. RESULTS: Plasma lipids were slightly affected to a more atherogenic profile by orchiectomy, but atherosclerotic lesion size was unaltered in the LAD, thoracic aorta, abdominal aorta, and iliac arteries. The distribution of lesion types (xanthomas, pathological intimal thickening and fibroatheromas) were also not statistically different between groups in any of the examined vascular territories. The abdominal aorta developed the most advanced stages of disease with reproducible fibroatheroma formation, and here it was found that the area of necrotic core was significantly increased in orchiectomized pigs compared with controls. Orchiectomy also reduced aggressive behavior. CONCLUSIONS: Castration does not alter the burden of atherosclerosis in hypercholesterolemic Yucatan minipigs, but may increase necrotic core area in fibroatheromas.


Assuntos
Aterosclerose/patologia , Hipercolesterolemia/patologia , Animais , Animais Geneticamente Modificados , Aorta/patologia , Aterosclerose/complicações , Dieta Hiperlipídica , Modelos Animais de Doenças , Humanos , Hipercolesterolemia/complicações , Hipercolesterolemia/genética , Artéria Ilíaca/patologia , Lipídeos/sangue , Masculino , Necrose , Orquiectomia , Pró-Proteína Convertase 9/genética , Pró-Proteína Convertase 9/metabolismo , Suínos , Porco Miniatura , Testosterona/sangue
8.
PLoS One ; 15(6): e0234772, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32555746

RESUMO

Glioblastoma is the most aggressive primary brain tumor leading to death in most of patients. It comprises almost 50-55% of all gliomas with an incidence rate of 2-3 per 100,000. Despite its rarity, overall mortality of glioblastoma is comparable to the most frequent tumors. The current standard treatment combines surgical resection, radiotherapy and chemotherapy with temozolomide. In spite of this aggressive multimodality protocol, prognosis of glioblastoma is poor and the median survival remains about 12-14.5 months. In this regard, new therapeutic approaches should be developed to improve the life quality and survival time of the patient after the initial diagnosis. Before switching to clinical trials in humans, all innovative therapeutic methods must be studied first on a relevant animal model in preclinical settings. In this regard, we validated the feasibility of intratumoral delivery of a holmium (Ho) microparticle suspension to an induced U87 glioblastoma model. Among the different radioactive beta emitters, 166Ho emits high-energy ß(-) radiation and low-energy γ radiation. ß(-) radiation is an effective means for tumor destruction and γ rays are well suited for imaging (SPECT) and consequent dosimetry. In addition, the paramagnetic Ho nucleus is a good asset to perform MRI imaging. In this study, five minipigs, implanted with our glioblastoma model were used to test the injectability of 165Ho (stable) using a bespoke injector and needle. The suspension was produced in the form of Ho microparticles and injected inside the tumor by a technique known as microbrachytherapy using a stereotactic system. At the end of this trial, it was found that the 165Ho suspension can be injected successfully inside the tumor with absence or minimal traces of Ho reflux after the injections. This injection technique and the use of the 165Ho suspension needs to be further assessed with radioactive 166Ho in future studies.


Assuntos
Neoplasias Encefálicas/radioterapia , Glioblastoma/radioterapia , Hólmio/química , Compostos Radiofarmacêuticos/administração & dosagem , Siloxanas/química , Animais , Neoplasias Encefálicas/diagnóstico por imagem , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Modelos Animais de Doenças , Estudos de Viabilidade , Feminino , Glioblastoma/diagnóstico por imagem , Glioblastoma/patologia , Humanos , Masculino , Compostos Radiofarmacêuticos/química , Compostos Radiofarmacêuticos/metabolismo , Suínos , Porco Miniatura , Tomografia Computadorizada de Emissão de Fóton Único , Transplante Heterólogo
9.
Cardiovasc Pathol ; 49: 107241, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32554057

RESUMO

In order to accelerate development of atherosclerosis(AS) in miniature swine models, varieties of strategies and methods have been explored. In addition to traditional methods such as high cholesterol feeding and balloon injury, new methods such as familial hypercholesterolemia induced by gene editing and intramural injection have been applied in recent years. Although it has been claimed that these methods have successfully aggravated lesion areas and stenosis, lesion features induced by different strategies have shown heterogeneity in morphology. In addition, time consumption, high cost, and unavailability are problems that restrict application of these AS models. Here, we summarize strategies and methods to accelerate AS models and further analyze their values, advantages, and shortcomings.


Assuntos
Artérias/patologia , Aterosclerose/etiologia , Placa Aterosclerótica , Angioplastia com Balão , Animais , Animais Geneticamente Modificados , Artérias/metabolismo , Aterosclerose/genética , Aterosclerose/metabolismo , Aterosclerose/patologia , Dieta Hiperlipídica , Modelos Animais de Doenças , Progressão da Doença , Edição de Genes , Predisposição Genética para Doença , Hipercolesterolemia/complicações , Hipercolesterolemia/genética , Suínos , Porco Miniatura/genética , Fatores de Tempo
10.
J Surg Res ; 254: 175-182, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32450418

RESUMO

BACKGROUND: Vascularized composite tissue allotransplantation (VCA) opens new possibilities for reconstruction of complex tissue defects, including upper extremity and facial transplantation. The main challenges in VCA transplantation are the side effects of long-term immunosuppression and chronic graft rejection. Translational preclinical animal models are crucial for VCA research to improve clinical outcomes and to study underlying immunologic mechanisms. Herein, we describe a novel, large animal, non-bone-bearing VCA model in inbred, swine leukocyte antigen-typed miniature swine. METHODS: Transplantation of vertical rectus abdominis myocutaneous (VRAM) flaps was performed between fully swine leukocyte antigen-mismatched miniature swine. The flaps were transferred to the posterolateral aspect of the neck of recipients and anastomosed to the common carotid artery and internal jugular vein. Different immunosuppressive drug regimens were used. Clinical graft evaluation was performed daily, and punch biopsies were taken for histology. RESULTS: Ten VRAM transplants were performed. The mean ischemia time was 89.4 min (SD ± 47), mean pedicle length 7.5 cm (SD ± 2), mean venous diameter 2.5 mm (SD ± 0.4), and mean arterial diameter 2.2 mm (SD ± 0.3). Follow-up demonstrated good correlation between clinical appearance and progression of graft rejection confirmed by histologic assessment. Complications were intraoperative cardiac arrest in one recipient and one flap loss due to venous compromise. CONCLUSIONS: VRAM transplantation in miniature swine is an appropriate preclinical VCA model, with the advantage of good clinical and histologic correlation during the course of rejection, as well as easy access to the graft. The availability of inbred, haplotyped animals allows studies across different major histocompatibility complex barriers in a non-bone-bearing VCA.


Assuntos
Rejeição de Enxerto/patologia , Reto do Abdome/transplante , Animais , Reto do Abdome/patologia , Suínos , Porco Miniatura , Transplante Heterotópico , Transplante Homólogo
11.
J Neuropathol Exp Neurol ; 79(6): 605-617, 2020 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-32386412

RESUMO

The neuropathology of mild traumatic brain injury in humans resulting from exposure to explosive blast is poorly understood as this condition is rarely fatal. A large animal model may better reflect the injury patterns in humans. We investigated the effect of explosive blasts on the constrained head minimizing the effects of whole head motion. Anesthetized Yucatan minipigs, with body and head restrained, were placed in a 3-walled test structure and exposed to 1, 2, or 3 explosive blast shock waves of the same intensity. Axonal injury was studied 3 weeks to 8 months postblast using ß-amyloid precursor protein immunohistochemistry. Injury was confined to the periventricular white matter as early as 3-5 weeks after exposure to a single blast. The pattern was also present at 8 months postblast. Animals exposed to 2 and 3 blasts had more axonal injury than those exposed to a single blast. Although such increases in axonal injury may relate to the longer postblast survival time, it may also be due to the increased number of blast exposures. It is possible that the injury observed is due to a condition akin to mild traumatic brain injury or subconcussive injury in humans, and that periventricular injury may have neuropsychiatric implications.


Assuntos
Traumatismos por Explosões/patologia , Concussão Encefálica/patologia , Encéfalo/patologia , Substância Branca/patologia , Animais , Axônios/patologia , Modelos Animais de Doenças , Masculino , Suínos , Porco Miniatura
12.
Exp Mol Pathol ; 115: 104470, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32445752

RESUMO

Sulfur mustard (SM), a dermal vesicant that has been used in chemical warfare, causes inflammation, edema and epidermal erosions depending on the dose and time following exposure. Herein, a minipig model was used to characterize wound healing following dermal exposure to SM. Saturated SM vapor caps were placed on the dorsal flanks of 3-month-old male Gottingen minipigs for 30 min. After 48 h the control and SM wounded sites were debrided daily for 7 days with wet to wet saline gauze soaks. Animals were then euthanized, and full thickness skin biopsies prepared for histology and immunohistochemistry. Control skin contained a well differentiated epidermis with a prominent stratum corneum. A well-developed eschar covered the skin of SM treated animals, however, the epidermis beneath the eschar displayed significant wound healing with a hyperplastic epidermis. Stratum corneum shedding and a multilayered basal epithelium consisting of cuboidal and columnar cells were also evident in the neoepidermis. Nuclear expression of proliferating cell nuclear antigen (PCNA) was contiguous in cells along the basal epidermal layer of control and SM exposed skin; SM caused a significant increase in PCNA expression in basal and suprabasal cells. SM exposure was also associated with marked changes in expression of markers of wound healing including increases in keratin 10, keratin 17 and loricrin and decreases in E-cadherin. Trichrome staining of control skin showed a well-developed collagen network with no delineation between the papillary and reticular dermis. Conversely, a major delineation was observed in SM-exposed skin including a web-like papillary dermis composed of filamentous extracellular matrix, and compact collagen fibrils in the lower reticular dermis. Although the dermis below the wound site was disrupted, there was substantive epidermal regeneration following SM-induced injury. Further studies analyzing the wound healing process in minipig skin will be important to provide a model to evaluate potential vesicant countermeasures.


Assuntos
Gás de Mostarda/toxicidade , Pele/patologia , Cicatrização , Animais , Caderinas/metabolismo , Diferenciação Celular/efeitos dos fármacos , Epiderme/efeitos dos fármacos , Epiderme/patologia , Proteínas de Membrana/metabolismo , Antígeno Nuclear de Célula em Proliferação/metabolismo , Pele/efeitos dos fármacos , Suínos , Porco Miniatura , Cicatrização/efeitos dos fármacos
13.
J Surg Res ; 253: 280-287, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32402853

RESUMO

BACKGROUND: The autologous vein remains the standard conduit for lower extremity and coronary artery bypass grafting despite a 30%-50% 5-y failure rate, primarily attributable to intimal hyperplasia (IH) that develops in the midterm period (3-24 mo) of graft maturation. Our group discovered that externally strengthening vein grafts by cross-linking the adventitial collagen with photochemical tissue passivation (PTP) mitigates IH in an arteriovenous model at 4 wk. We now investigate whether this effect is retained in the midterm period follow-up. METHODS: Six Hanford miniature pigs received bilateral carotid artery interposition vein grafts. In each animal, the external surface of one graft was treated with PTP before grafting, whereas the opposite side served as the untreated control. The grafts were harvested after 3 mo. Ultrasound evaluation of all vein grafts was performed at the time of grafting and harvest. The grafts were also evaluated histomorphometrically and immunohistologically for markers of IH. RESULTS: All vein grafts were patent at 3 mo except one graft in the PTP-treated group because of early technical failure. The control vein grafts had significantly greater IH than PTP-treated grafts at 3 mo, as evidenced by the intimal area (2.6 ± 1.0 mm2versus 1.4 ± 1.5 mm2, respectively, P = 0.045) and medial area (5.1 ± 1.9 mm2versus 2.7 ± 2.4 mm2, respectively, P = 0.048). The control grafts had an increased presence and proliferation of mural myofibroblasts with greater smooth muscle actin and proliferating cell nuclear antigen staining. CONCLUSIONS: PTP treatment to the external surface of the vein grafts decreases IH at 3 mo after arteriovenous grafting and may prevent future graft failure.


Assuntos
Artérias Carótidas/cirurgia , Neointima/prevenção & controle , Fotoquimioterapia/métodos , Veia Safena/transplante , Enxerto Vascular/métodos , Túnica Adventícia/efeitos dos fármacos , Túnica Adventícia/efeitos da radiação , Animais , Colágeno/química , Colágeno/efeitos dos fármacos , Colágeno/efeitos da radiação , Feminino , Corantes Fluorescentes/administração & dosagem , Luz , Neointima/diagnóstico , Neointima/etiologia , Neointima/patologia , Rosa Bengala/administração & dosagem , Veia Safena/diagnóstico por imagem , Veia Safena/patologia , Suínos , Porco Miniatura , Transplante Autólogo/efeitos adversos , Túnica Íntima/diagnóstico por imagem , Túnica Íntima/patologia , Enxerto Vascular/efeitos adversos , Grau de Desobstrução Vascular
14.
Sci Rep ; 10(1): 7278, 2020 04 29.
Artigo em Inglês | MEDLINE | ID: mdl-32350389

RESUMO

SIRT1 (silent information regulator 1) is a histone deacetylase. It can sense the energy level in cells and delay cell senescence, leading to resistance to external stress and improving metabolism. Mitral regurgitation (MR) is a common disease in cardiac surgery. However, there are no previous studies on SIRT1 and left atrial fibrosis caused by MR. In this study, we aimed to explore the regulatory effect of SIRT1 on left atrial fibrosis induced by MR. We used Guizhou miniature pigs to establish an MR model and a sham operation model after anaesthesia induction and respiratory intubation, and these model animals were followed for 30 months after the surgery. The differential distribution and expression of SIRT1 and collagen I in the left atrium was determined by immunofluorescence and Western blotting. Furthermore, we treated NIH3T3 fibroblasts (CFs) with resveratrol and Angiotensin II (Ang II) to analyse the specific mechanism involved in the development of myocardial fibrosis. The results showed that the MR model was successfully constructed. There were 8 pigs in the MR group and 6 pigs in the control group. In both the animal experiments and the cell experiments, the expression of collagen I in the MR group was increased significantly compared to that in the control group, while the expression of SIRT1 was decreased.


Assuntos
Regulação Enzimológica da Expressão Gênica , Insuficiência da Valva Mitral/enzimologia , Sirtuína 1/biossíntese , Animais , Modelos Animais de Doenças , Fibrose , Átrios do Coração/enzimologia , Átrios do Coração/patologia , Camundongos , Insuficiência da Valva Mitral/patologia , Células NIH 3T3 , Suínos , Porco Miniatura
15.
BMC Cardiovasc Disord ; 20(1): 176, 2020 04 16.
Artigo em Inglês | MEDLINE | ID: mdl-32295540

RESUMO

BACKGROUND: Renal denervation (RDN) targeting the sympathetic nerves in the renal arterial adventitia as a treatment of resistant hypertension can cause endothelial injury and vascular wall injury. This study aims to evaluate the risk of atherosclerosis induced by RDN in renal arteries. METHODS: A total of 15 minipigs were randomly assigned to 3 groups: (1) control group, (2) sham group, and (3) RDN group (n = 5 per group). All pigs were fed a high-fat diet (HFD) for 6 months after appropriate treatment. The degree of intimal thickening of renal artery and the conversion of endothelin 1 (ET-1) receptors were evaluated by histological staining. Western blot was used to assess the expression of nitric oxide (NO) synthesis signaling pathway, ET-1 and its receptors, NADPH oxidase 2 (NOX2) and 4-hydroxynonenal (4-HNE) proteins, and the activation of NF-kappa B (NF-κB). RESULTS: The histological staining results suggested that compared to the sham treatment, RDN led to significant intimal thickening and significantly promoted the production of endothelin B receptor (ETBR) in vascular smooth muscle cells (VSMCs). Western blotting analysis indicated that RDN significantly suppressed the expression of AMPK/Akt/eNOS signaling pathway proteins, and decreased the production of NO, and increased the expression of endothelin system proteins including endothelin-1 (ET-1), endothelin converting enzyme 1 (ECE1), endothelin A receptor (ETAR) and ETBR; and upregulated the expression of NOX2 and 4-HNE proteins and enhanced the activation of NF-kappa B (NF-κB) when compared with the sham treatment (all p < 0.05). There were no significant differences between the control and sham groups (all p > 0.05). CONCLUSIONS: RDN aggravated endothelial endocrine dysfunction and intimal thickening, and increased the risk of atherosclerosis in renal arteries of HFD-fed pigs.


Assuntos
Aterosclerose/etiologia , Dieta Hiperlipídica , Células Endoteliais/metabolismo , Neointima , Obesidade/metabolismo , Artéria Renal/inervação , Artéria Renal/metabolismo , Simpatectomia/efeitos adversos , Proteínas Quinases Ativadas por AMP/metabolismo , Aldeídos/metabolismo , Animais , Aterosclerose/metabolismo , Aterosclerose/patologia , Modelos Animais de Doenças , Células Endoteliais/patologia , Masculino , NADPH Oxidase 2/metabolismo , NF-kappa B/metabolismo , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Obesidade/patologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores de Endotelina/metabolismo , Artéria Renal/patologia , Transdução de Sinais , Suínos , Porco Miniatura
16.
Cancer Chemother Pharmacol ; 85(4): 805-816, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32185484

RESUMO

PURPOSE: Metarrestin is a first-in-class pyrrolo-pyrimidine-derived small molecule targeting a marker of genome organization associated with metastasis and is currently in preclinical development as an anti-cancer agent. Here, we report the in vitro ADME characteristics and in vivo pharmacokinetic behavior of metarrestin. METHODS: Solubility, permeability, and efflux ratio as well as in vitro metabolism of metarrestin in hepatocytes, liver microsomes and S9 fractions, recombinant cytochrome P450 (CYP) enzymes, and potential for CYP inhibition were evaluated. Single dose pharmacokinetic profiles after intravenous and oral administration in mice, rat, dog, monkey, and mini-pig were obtained. Simple allometric scaling was applied to predict human pharmacokinetics. RESULTS: Metarrestin had an aqueous solubility of 150 µM at pH 7.4, high permeability in PAMPA and moderate efflux ratio in Caco-2 assays. The compound was metabolically stable in liver microsomes, S9 fractions, and hepatocytes from six species, including human. Metarrestin is a CYP3A4 substrate and, in mini-pigs, is also directly glucuronidated. Metarrestin did not show cytochrome P450 inhibitory activity. Plasma concentration-time profiles showed low to moderate clearance, ranging from 0.6 mL/min/kg in monkeys to 48 mL/min/kg in mice and moderate to high volume of distribution, ranging from 1.5 L/kg in monkeys to 17 L/kg in mice. Metarrestin has greater than 80% oral bioavailability in all species tested. The excretion of unchanged parent drug in urine was < 5% in dogs and < 1% in monkeys over collection periods of ≥ 144 h; in bile-duct cannulated rats, the excretion of unchanged drug was < 1% in urine and < 2% in bile over a collection period of 48 h. CONCLUSIONS: Metarrestin is a low clearance compound which has good bioavailability and large biodistribution after oral administration. Biotransformation appears to be the major elimination process for the parent drug. In vitro data suggest a low drug-drug interaction potential on CYP-mediated metabolism. Overall favorable ADME and PK properties support metarrestin's progression to clinical investigation.


Assuntos
Sistema Enzimático do Citocromo P-450/química , Microssomos Hepáticos/metabolismo , Pirimidinas/administração & dosagem , Pirimidinas/farmacocinética , Pirróis/administração & dosagem , Pirróis/farmacocinética , Administração Oral , Animais , Biotransformação , Inibidores das Enzimas do Citocromo P-450/administração & dosagem , Inibidores das Enzimas do Citocromo P-450/farmacocinética , Sistema Enzimático do Citocromo P-450/metabolismo , Cães , Avaliação Pré-Clínica de Medicamentos , Interações Medicamentosas , Feminino , Haplorrinos , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microssomos Hepáticos/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Especificidade da Espécie , Suínos , Porco Miniatura , Distribuição Tecidual
17.
Sci Rep ; 10(1): 5291, 2020 03 24.
Artigo em Inglês | MEDLINE | ID: mdl-32210315

RESUMO

BACKGROUND: Prior studies have applied driver mutations targeting the RTK/RAS/PI3K and p53 pathways to induce the formation of high-grade gliomas in rodent models. In the present study, we report the production of a high-grade spinal cord glioma model in pigs using lentiviral gene transfer. METHODS: Six Gottingen Minipigs received thoracolumbar (T14-L1) lateral white matter injections of a combination of lentiviral vectors, expressing platelet-derived growth factor beta (PDGF-B), constitutive HRAS, and shRNA-p53 respectively. All animals received injection of control vectors into the contralateral cord. Animals underwent baseline and endpoint magnetic resonance imaging (MRI) and were evaluated daily for clinical deficits. Hematoxylin and eosin (H&E) and immunohistochemical analysis was conducted. Data are presented using descriptive statistics including relative frequencies, mean, standard deviation, and range. RESULTS: 100% of animals (n = 6/6) developed clinical motor deficits ipsilateral to the oncogenic lentiviral injections by a three-week endpoint. MRI scans at endpoint demonstrated contrast enhancing mass lesions at the site of oncogenic lentiviral injection and not at the site of control injections. Immunohistochemistry demonstrated positive staining for GFAP, Olig2, and a high Ki-67 proliferative index. Histopathologic features demonstrate consistent and reproducible growth of a high-grade glioma in all animals. CONCLUSIONS: Lentiviral gene transfer represents a feasible pathway to glioma modeling in higher order species. The present model is the first lentiviral vector induced pig model of high-grade spinal cord glioma and may potentially be used in preclinical therapeutic development programs.


Assuntos
Modelos Animais de Doenças , Vetores Genéticos/administração & dosagem , Glioma/patologia , Lentivirus/genética , Transtornos Motores/patologia , Neoplasias da Medula Espinal/patologia , Animais , Feminino , Vetores Genéticos/genética , Glioma/genética , Humanos , Masculino , Transtornos Motores/genética , Gradação de Tumores , Neoplasias da Medula Espinal/genética , Suínos , Porco Miniatura
18.
Lipids Health Dis ; 19(1): 54, 2020 Mar 25.
Artigo em Inglês | MEDLINE | ID: mdl-32213192

RESUMO

BACKGROUND: Atherosclerosis is a major contributor to cardiovascular events, however, its molecular mechanism remains poorly known. Animal models of atherosclerosis can be a valuable tool to provide insights into the etiology, pathophysiology, and complications of atherosclerosis. In particular, Tibetan minipigs are a feasible model for studying diet-related metabolic and atherosclerotic diseases. METHODS: We used vascular transcriptomics to identify differentially expressed genes (DEGs) in high fat/cholesterol (HFC) diet-fed Tibetan minipig atherosclerosis models, analyzed the DEGs gene ontology (GO) terms, pathways and protein-protein interactions (PPI) networks, and identified hub genes and key modules using molecular complex detection (MCODE), Centiscape and CytoHubba plugin. The identified genes were validated using the human carotid atherosclerosis database (GSEA 43292) and RT-PCR methods. RESULTS: Our results showed that minipigs displayed obvious dyslipidemia, oxidative stress, inflammatory response, atherosclerotic plaques, as well as increased low-density lipoprotein (LDL) and leukocyte recruitment after 24 weeks of HFC diet feeding compared to those under a regular diet. Our RNA-seq results revealed 1716 DEGs in the atherosclerotic/NC group, of which 1468 genes were up-regulated and 248 genes were down-regulated. Functional enrichment analysis of DEGs showed that the HFC diet-induced changes are related to vascular immune-inflammatory responses, lipid metabolism and muscle contraction, indicating that hypercholesterolemia caused by HFC diet can activate innate and adaptive immune responses to drive atherosclerosis development. Furthermore, we identified four modules from the major PPI network, which are implicated in cell chemotaxis, myeloid leukocyte activation, cytokine production, and lymphocyte activation. Fifteen hub genes were discovered, including TNF, PTPRC, ITGB2, ITGAM, VCAM1, CXCR4, TYROBP, TLR4, LCP2, C5AR1, CD86, MMP9, PTPN6, C3, and CXCL10, as well as two transcription factors (TF), i.e. NF-ĸB1 and SPI1. These results are consistent with the expression patterns in human carotid plaque and were validated by RT-PCR. CONCLUSIONS: The identified DEGs and their enriched pathways provide references for the development and progression mechanism of Tibetan minipig atherosclerosis model induced by the HFC diet.


Assuntos
Aterosclerose/metabolismo , Biologia Computacional/métodos , Dieta Hiperlipídica/efeitos adversos , Animais , Aterosclerose/etiologia , Aterosclerose/genética , Ontologia Genética , Contagem de Leucócitos , Masculino , Mapas de Interação de Proteínas , RNA Mensageiro/metabolismo , RNA-Seq , Suínos , Porco Miniatura , Tibet
19.
Life Sci ; 250: 117514, 2020 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-32145306

RESUMO

AIMS: Pigs are increasingly used as human metabolic disease models; however, there is insufficient research on breed-related genetic background differences. This study aimed to investigate the differential metabolic responses to high-fat and high-cholesterol (HFC) diet-induced non-alcoholic fatty liver disease (NAFLD) of two miniature pig breeds and explore the molecular mechanisms involved. MAIN METHODS: Male Wuzhishan (WZSP) and Tibetan pigs (TP) were randomly fed either a standard or an HFC diet for 24 weeks. Weight, serum lipids, bile acid, insulin resistance, liver function, liver histology, and hepatic lipid deposition were determined. RNA-Seq was used to detect the hepatic gene expression profiles. Western blot, immunohistochemistry, and qRT-PCR were used to detect the lipid and glucose metabolism-related gene expressions. KEY FINDINGS: The HFC diet caused obesity, hypertension, severe hypercholesterolemia, liver injury, increased hepatocellular steatosis and inflammation, and significantly increased serum insulin levels in both pig breeds. This diet led to higher serum and hepatic cholesterol level concentrations in WZSP and elevated fasting glucose levels in TP. Transcriptome analysis revealed that the genes controlling hepatic cholesterol metabolism and the inflammatory response were consistently regulated; lipid metabolism and insulin signaling related genes were uniquely regulated by the HFC diet in the WZSP and TP, respectively. SIGNIFICANCE: Our study demonstrated that the genetic background affects profoundly pigs' metabolic and hepatic responses to an HFC diet. These results deepened our understanding of the molecular mechanisms of HFC diet-induced NAFLD and provided a foundation for selecting the appropriate pig breeds for metabolic studies in the future.


Assuntos
Ração Animal , Colesterol na Dieta , Dieta Hiperlipídica , Hepatopatia Gordurosa não Alcoólica/metabolismo , Hepatopatia Gordurosa não Alcoólica/fisiopatologia , Transcriptoma , Animais , Glicemia/análise , Colesterol/metabolismo , Modelos Animais de Doenças , Biblioteca Gênica , Hipercolesterolemia/etiologia , Hiperlipidemias/etiologia , Hipertensão/etiologia , Inflamação/etiologia , Insulina/metabolismo , Fígado/metabolismo , Hepatopatias/etiologia , Masculino , Obesidade/etiologia , Fenótipo , Distribuição Aleatória , Especificidade da Espécie , Suínos , Porco Miniatura
20.
Circ Arrhythm Electrophysiol ; 13(4): e007614, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32189516

RESUMO

BACKGROUND: Heart rate variability (HRV) and pulse rate variability are indices of autonomic cardiac modulation. Increased pericardial fat is associated with worse cardiovascular outcomes. We hypothesized that progressive increases in pericardial fat volume and inflammation prospectively dampen HRV in hypercholesterolemic pigs. METHODS: WT (wild type) or PCSK9 (proprotein convertase subtilisin-like/kexin type-9) gain-of-function Ossabaw mini-pigs were studied in vivo before and after 3 and 6 months of a normal diet (WT-normal diet, n=4; PCSK9-normal diet, n=6) or high-fat diet (HFD; WT-HFD, n=3; PCSK9-HFD, n=6). The arterial pulse waveform was obtained from an arterial telemetry transmitter to analyze HRV indices, including SD (SD of all pulse-to-pulse intervals over a single 5-minute period), root mean square of successive differences, proportion >50 ms of normal-to-normal R-R intervals, and the calculated ratio of low-to-high frequency distributions (low-frequency power/high-frequency power). Pericardial fat volumes were evaluated using multidetector computed tomography and its inflammation by gene expression of TNF (tumor necrosis factor)-α. Plasma lipid panel and norepinephrine level were also measured. RESULTS: At diet completion, hypercholesterolemic PCSK9-HFD had significantly (P<0.05 versus baseline) depressed HRV (SD of all pulse-to-pulse intervals over a single 5-minute period, root mean square of successive differences, proportion >50 ms, high-frequency power, low-frequency power), and both HFD groups had higher sympathovagal balance (SD of all pulse-to-pulse intervals over a single 5-minute period/root mean square of successive differences, low-frequency power/high-frequency power) compared with normal diet. Pericardial fat volumes and LDL (low-density lipoprotein) cholesterol concentrations correlated inversely with HRV and directly with sympathovagal balance, while sympathovagal balance correlated directly with plasma norepinephrine. Pericardial fat TNF-α expression was upregulated in PCSK9-HFD, colocalized with nerve fibers, and correlated inversely with root mean square of successive differences and proportion >50 ms. CONCLUSIONS: Progressive pericardial fat expansion and inflammation are associated with a fall in HRV in Ossabaw mini-pigs, implying aggravated autonomic imbalance. Hence, pericardial fat accumulation is associated with alterations in HRV and the autonomic nervous system. Visual Overview: A visual overview is available for this article.


Assuntos
Tecido Adiposo/fisiopatologia , Adiposidade , Arritmias Cardíacas/etiologia , Sistema Nervoso Autônomo/fisiopatologia , Frequência Cardíaca , Hipercolesterolemia/complicações , Inflamação/etiologia , Pericárdio/fisiopatologia , Tecido Adiposo/metabolismo , Animais , Animais Geneticamente Modificados , Arritmias Cardíacas/metabolismo , Arritmias Cardíacas/fisiopatologia , Sistema Nervoso Autônomo/metabolismo , Colesterol/sangue , Modelos Animais de Doenças , Hipercolesterolemia/metabolismo , Hipercolesterolemia/fisiopatologia , Inflamação/metabolismo , Inflamação/fisiopatologia , Mediadores da Inflamação/metabolismo , Masculino , Norepinefrina/sangue , Pericárdio/metabolismo , Suínos , Porco Miniatura/genética , Fatores de Tempo , Fator de Necrose Tumoral alfa/metabolismo
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