RESUMO
Pathologic evaluation is crucial to the study of medical devices and integral to the Food and Drug Administration and other regulatory entities' assessment of device safety and efficacy. While pathologic analysis is tailored to the type of device, it generally involves at a minimum gross and microscopic evaluation of the medical device and associated tissues. Due to the complex nature of some implanted devices and specific questions posed by sponsors, pathologic evaluation inherently presents many challenges in accurately assessing medical device safety and efficacy. This laboratory's experience in numerous collaborative projects involving veterinary pathologists, biomedical engineers, physicians, and other scientists has led to a set of interrelated assessments to determine pathologic end points as a means to address these challenges and achieve study outcomes. Thorough device evaluation is often accomplished by utilizing traditional paraffin histology, plastic embedding and microground sections, and advanced imaging modalities. Combining these advanced techniques provides an integrative, comprehensive approach to medical device pathology and enhances medical device safety and efficacy assessment.
Assuntos
Aprovação de Equipamentos/normas , Segurança de Equipamentos/normas , Equipamentos e Provisões/normas , Patologia/métodos , Animais , Aprovação de Equipamentos/legislação & jurisprudência , Equipamentos e Provisões/efeitos adversos , Técnicas Histológicas/métodos , Técnicas Histológicas/normas , Humanos , Modelos Animais , Estados Unidos , United States Food and Drug AdministrationRESUMO
BACKGROUND: Inflammatory bowel disease (IBD) increases the risk of developing colon cancer. This risk is higher in men compared to women, implicating a role for female hormones in the protection against this disease. Studies from our laboratory demonstrated that estradiol (E2) protects against inflammation-associated colon tumor formation when administered following chemical carcinogen and induction of chronic colitis. AIM: This study seeks to better understand the effect of E2 on acute colitis in the presence and absence of estrogen receptor ß (ERß). METHODS: Inflammation was induced by 2,4,6-trinitrobenzenesulfonic acid in wild-type (WT) and ERß knockout (ERßKO) mice implanted with a control or E2-containing pellet and killed 5 days later. Inflammation and injury were scored by a pathologist. Apoptosis and proliferation were assessed by immunohistochemistry. Cytokines were measured by multiplex analysis. RESULTS: E2 treatment reduced inflammation in the middle colon in WT mice and the distal colon in ERßKO mice compared to control mice. WT mice had reduced IL-6, IL-12, IL-17, GM-CSF, IFN-γ, MCP-1, MIP-1α, and TNF-α, and ERßKO had reduced IL-6 and IFN-γ expression in response to E2. Injury scores were lower in E2-treated ERßKO mice compared to control ERßKO mice. ERßKO mice had increased proliferation in the basal third of crypts in the distal colon and decreased apoptosis in the proximal colon. CONCLUSIONS: These data suggest that E2 has differential protective effects against acute colitis in the presence or absence of ERß and provide insight into how E2 may protect against IBD.
Assuntos
Colite/tratamento farmacológico , Colite/metabolismo , Estradiol/farmacologia , Receptor beta de Estrogênio/metabolismo , Estrogênios/farmacologia , Animais , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Colite/induzido quimicamente , Neoplasias do Colo/induzido quimicamente , Neoplasias do Colo/prevenção & controle , Citocinas/análise , Citocinas/efeitos dos fármacos , Receptor beta de Estrogênio/análise , Receptor beta de Estrogênio/genética , Feminino , Imuno-Histoquímica , Mediadores da Inflamação/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ácido TrinitrobenzenossulfônicoRESUMO
Perturbations in DNA damage, DNA repair, apoptosis and cell proliferation in the base of the crypt where stem cells reside are associated with colorectal cancer (CRC) initiation and progression. Although the transformation of leucine-rich repeat-containing G protein-coupled receptor 5 (Lgr5)(+) cells is an extremely efficient route towards initiating small intestinal adenomas, the role of Lgr5(+) cells in CRC pathogenesis has not been well investigated. Therefore, we further characterized the properties of colonic Lgr5(+) cells compared to differentiated cells in Lgr5-EGFP-IRES-creER(T2) knock-in mice at the initiation stage of carcinogen azoxymethane (AOM)-induced tumorigenesis using a quantitative immunofluorescence microscopy approach. At 12 and 24h post-AOM treatment, colonic Lgr5(+) stem cells (GFP(high)) were preferentially damaged by carcinogen, exhibiting a 4.7-fold induction of apoptosis compared to differentiated (GFP(neg)) cells. Furthermore, with respect to DNA repair, O(6)-methylguanine DNA methyltransferase (MGMT) expression was preferentially induced (by 18.5-fold) in GFP(high) cells at 24h post-AOM treatment compared to GFP(neg) differentiated cells. This corresponded with a 4.3-fold increase in cell proliferation in GFP(high) cells. These data suggest that Lgr5(+) stem cells uniquely respond to alkylation-induced DNA damage by upregulating DNA damage repair, apoptosis and cell proliferation compared to differentiated cells in order to maintain genomic integrity. These findings highlight the mechanisms by which colonic Lgr5(+) stem cells respond to cancer-causing environmental factors.
Assuntos
Transformação Celular Neoplásica/efeitos dos fármacos , Homeostase/efeitos dos fármacos , Mucosa Intestinal/citologia , Células-Tronco/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Carcinógenos/toxicidade , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/fisiologia , Neoplasias Colorretais/patologia , Neoplasias Colorretais/fisiopatologia , Dano ao DNA/efeitos dos fármacos , Dano ao DNA/fisiologia , Reparo do DNA/efeitos dos fármacos , Reparo do DNA/fisiologia , Modelos Animais de Doenças , Técnicas de Introdução de Genes , Homeostase/fisiologia , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/patologia , Camundongos , Mutagênicos/toxicidade , Receptores Acoplados a Proteínas G/metabolismo , Células-Tronco/metabolismo , Células-Tronco/patologiaRESUMO
BACKGROUND: Endogenous ochronosis is caused by a defect in the enzyme homogentisate 1,2-dioxygenase (HGD), which results in abnormal pigment deposition in the skin and urine abnormalities. Ochronosis previously has not been described histologically or ultrastructurally in a domestic animal species. HYPOTHESIS/OBJECTIVES: To describe the clinical, histopathological and ultrastructural findings in a case of aberrant pigmentation in a cat with features that resemble ochronosis. ANIMAL: A 5-year-old, spayed female Domestic short hair cat presented with multiple black cutaneous plaques on the face and progressive lethargy. The cat's urine turned brown when exposed to air. The familial history of the cat was unknown. METHODS: Clinical examination; histopathology, electron microscopy and mass/energy dispersive X-ray spectroscopy of tissues. RESULTS: Septic peritonitis and additional pigment in the spleen, intestine and lymph node were found at postmortem examination. The pigment was determined to be an organic compound and had a similar histological appearance, staining properties, ultrastructure and composition to ochronotic pigment. No mutations were found in exons 3, 6, 8 and 13 of the HGD gene in the cat. CONCLUSIONS AND CLINICAL IMPORTANCE: To the best of the authors' knowledge, this is the first report of a condition resembling ochronosis in a domestic animal species that has been evaluated with histopathology and advanced imaging techniques. It provides an additional differential in cases of aberrant pigmentation in cats.
RESUMO
During colitis, activation of two inflammatory T cell subsets, Th17 and Th1 cells, promotes ongoing intestinal inflammatory responses. n-6 polyunsaturated fatty acid- (PUFA-) derived eicosanoids, such as prostaglandin E2 (PGE2), promote Th17 cell-mediated inflammation, while n-3 PUFA antagonize both Th17 and Th1 cells and suppress PGE2 levels. We utilized two genetic mouse models, which differentially antagonize PGE2 levels, to examine the effect on Th17 cells and disease outcomes in trinitrobenzene sulfonic acid- (TNBS-) induced colitis. Fat-1 mice contain the ω3 desaturase gene from C. elegans and synthesize n-3 PUFA de novo, thereby reducing the biosynthesis of n-6 PUFA-derived eicosanoids. In contrast, Fads1 Null mice contain a disrupted Δ5 desaturase gene and produce lower levels of n-6 PUFA-derived eicosanoids. Compared to Wt littermates, Fat-1 and Fads1 Null mice exhibited a similar colitic phenotype characterized by reduced colonic mucosal inflammatory eicosanoid levels and mRNA expression of Th17 cell markers (IL-17A, RORγτ, and IL-23), decreased percentages of Th17 cells and, improved colon injury scores (P ≤ 0.05). Thus, during colitis, similar outcomes were obtained in two genetically distinct models, both of which antagonize PGE2 levels via different mechanisms. Our data highlight the critical impact of n-6 PUFA-derived eicosanoids in the promotion of Th17 cell-mediated colonic inflammation.
Assuntos
Ácido Araquidônico/química , Colite/tratamento farmacológico , Colite/imunologia , Eicosanoides/química , Eicosanoides/uso terapêutico , Inflamação/imunologia , Inflamação/metabolismo , Células Th1/metabolismo , Células Th17/metabolismo , Animais , Colite/metabolismo , Feminino , Masculino , CamundongosRESUMO
Epidermal growth factor receptor (EGFR)-mediated signaling is required for optimal intestinal wound healing. Since n-3 polyunsaturated fatty acids (PUFA), specifically docosahexaenoic acid (DHA), alter EGFR signaling and suppress downstream activation of key signaling pathways, we hypothesized that DHA would be detrimental to the process of intestinal wound healing. Using a mouse immortalized colonocyte model, DHA uniquely reduced EGFR ligand-induced receptor activation, whereas DHA and its metabolic precursor eicosapentaenoic acid (EPA) reduced wound-induced EGFR transactivation compared with control (no fatty acid or linoleic acid). Under wounding conditions, the suppression of EGFR activation was associated with a reduction in downstream activation of cytoskeletal remodeling proteins (PLCγ1, Rac1, and Cdc42). Subsequently, DHA and EPA reduced cell migration in response to wounding. Mice were fed a corn oil-, DHA-, or EPA-enriched diet prior to intestinal wounding (2.5% dextran sodium sulfate for 5 days followed by termination after 0, 3, or 6 days of recovery). Mortality was increased in EPA-fed mice and colonic histological injury scores were increased in EPA- and DHA-fed mice compared with corn oil-fed (control) mice. Although kinetics of colonic EGFR activation and downstream signaling (PLCγ1, Rac1, and Cdc42) were delayed by both n-3 PUFA, colonic repair was increased in EPA- relative to DHA-fed mice. These results indicate that, during the early response to intestinal wounding, DHA and EPA uniquely delay the activation of key wound-healing processes in the colon. This effect is mediated, at least in part, via suppression of EGFR-mediated signaling and downstream cytoskeletal remodeling.
Assuntos
Ácidos Docosa-Hexaenoicos/administração & dosagem , Ácido Eicosapentaenoico/administração & dosagem , Receptores ErbB/metabolismo , Processamento de Proteína Pós-Traducional , Cicatrização , Animais , Ácido Araquidônico/metabolismo , Movimento Celular , Células Cultivadas , Colite/induzido quimicamente , Colite/tratamento farmacológico , Colite/patologia , Colo/efeitos dos fármacos , Colo/patologia , Óleo de Milho/administração & dosagem , Sulfato de Dextrana , Suplementos Nutricionais , Ácidos Docosa-Hexaenoicos/fisiologia , Ácido Eicosapentaenoico/fisiologia , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias/metabolismo , Neuropeptídeos/metabolismo , Consumo de Oxigênio , Fosforilação , Transdução de Sinais , Ácido alfa-Linolênico/farmacologia , Ácido alfa-Linolênico/fisiologia , Proteína cdc42 de Ligação ao GTP/metabolismo , Proteínas rac de Ligação ao GTP/metabolismo , Proteínas rac1 de Ligação ao GTPRESUMO
Arachidonic acid (20:4(Δ5,8,11,14), AA)-derived eicosanoids regulate inflammation and promote cancer development. Previous studies have targeted prostaglandin enzymes in an attempt to modulate AA metabolism. However, due to safety concerns surrounding the use of pharmaceutical agents designed to target Ptgs2 (cyclooxygenase 2) and its downstream targets, it is important to identify new targets upstream of Ptgs2. Therefore, we determined the utility of antagonizing tissue AA levels as a novel approach to suppressing AA-derived eicosanoids. Systemic disruption of the Fads1 (Δ5 desaturase) gene reciprocally altered the levels of dihomo-γ-linolenic acid (20:3(Δ8,11,14), DGLA) and AA in mouse tissues, resulting in a profound increase in 1-series-derived and a concurrent decrease in 2-series-derived prostaglandins. The lack of AA-derived eicosanoids, e.g., PGE2 was associated with perturbed intestinal crypt proliferation, immune cell homeostasis, and a heightened sensitivity to acute inflammatory challenge. In addition, null mice failed to thrive, dying off by 12 weeks of age. Dietary supplementation with AA extended the longevity of null mice to levels comparable to wild-type mice. We propose that this new mouse model will expand our understanding of how AA and its metabolites mediate inflammation and promote malignant transformation, with the eventual goal of identifying new drug targets upstream of Ptgs2.
Assuntos
Modelos Animais de Doenças , Eicosanoides/metabolismo , Ácidos Graxos Dessaturases/metabolismo , Animais , Ácido Araquidônico/administração & dosagem , Dessaturase de Ácido Graxo Delta-5 , Suplementos Nutricionais , Eicosanoides/deficiência , Ácidos Graxos Dessaturases/deficiência , Ácidos Graxos Dessaturases/genética , Inflamação , Camundongos , Camundongos Endogâmicos C57BL , Camundongos KnockoutRESUMO
The ligand-activated transcription factor peroxisome proliferator-activated receptor (PPAR)-δ is highly expressed in colonic epithelial cells; however, the role of PPARδ ligands, such as fatty acids, in mucosal inflammation and malignant transformation has not been clarified. Recent evidence suggests that the anti-inflammatory/chemoprotective properties of fish oil (FO)-derived n-3 polyunsaturated fatty acids (PUFAs) may be partly mediated by PPARδ. Therefore, we assessed the role of PPARδ in modulating the effects of dietary n-3 PUFAs by targeted deletion of intestinal epithelial cell PPARδ (PPARδ(ΔIEpC)). Subsequently, we documented changes in colon tumorigenesis and the inflammatory microenvironment, i.e., local [mesenteric lymph node (MLN)] and systemic (spleen) T cell activation. Animals were fed chemopromotive [corn oil (CO)] or chemoprotective (FO) diets during the induction of chronic inflammation/carcinogenesis. Tumor incidence was similar in control and PPARδ(ΔIEpC) mice. FO reduced mucosal injury, tumor incidence, colonic STAT3 activation, and inflammatory cytokine gene expression, independent of PPARδ genotype. CD8(+) T cell recruitment into MLNs was suppressed in PPARδ(ΔIEpC) mice. Similarly, FO reduced CD8(+) T cell numbers in the MLN. Dietary FO independently modulated MLN CD4(+) T cell activation status by decreasing CD44 expression. CD11a expression by MLN CD4(+) T cells was downregulated in PPARδ(ΔIEpC) mice. Lastly, splenic CD62L expression was downregulated in PPARδ(ΔIEpC) CD4(+) and CD8(+) T cells. These data demonstrate that expression of intestinal epithelial cell PPARδ does not influence azoxymethane/dextran sodium sulfate-induced colon tumor incidence. Moreover, we provide new evidence that dietary n-3 PUFAs attenuate intestinal inflammation in an intestinal epithelial cell PPARδ-independent manner.
Assuntos
Adenocarcinoma/tratamento farmacológico , Transformação Celular Neoplásica/efeitos dos fármacos , Colite/tratamento farmacológico , Neoplasias do Colo/tratamento farmacológico , Gorduras Insaturadas na Dieta/farmacologia , Óleos de Peixe/farmacologia , Deleção de Genes , PPAR delta/metabolismo , Adenocarcinoma/genética , Adenocarcinoma/imunologia , Animais , Antígeno CD11a/biossíntese , Antígeno CD11a/imunologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/imunologia , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/imunologia , Doença Crônica , Colite/genética , Colite/imunologia , Neoplasias do Colo/genética , Neoplasias do Colo/imunologia , Citocinas/biossíntese , Gorduras Insaturadas na Dieta/imunologia , Gorduras Insaturadas na Dieta/metabolismo , Feminino , Óleos de Peixe/imunologia , Receptores de Hialuronatos/biossíntese , Receptores de Hialuronatos/imunologia , Mucosa Intestinal/imunologia , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , PPAR delta/genética , PPAR delta/imunologia , Fator de Transcrição STAT3/biossíntese , Baço/efeitos dos fármacos , Baço/imunologiaRESUMO
During colon inflammation, Th17 cells and immunosuppressive regulatory T cells (Treg) are thought to play promotive and preventative roles, respectively. Dietary (n-3) PUFA favorably modulate intestinal inflammation in part by downregulating T-cell activation and functionality. We used the Fat-1 mouse, a genetic model that synthesizes long-chain (n-3) PUFA de novo, to test the hypothesis that (n-3) PUFA protect against colonic inflammation by modulating the polarization of Treg and Th17 cells during colitis. Male and female wild-type (WT) and Fat-1 mice were administered dextran sodium sulfate (DSS) in the drinking water (2.5%) to induce acute (5 d DSS) or chronic (3 cycles DSS) colitis and the percentage of Treg and Th17 cells residing locally [colonic lamina propria (cLP)] and systemically (spleen) was determined by flow cytometry. The percentage of Treg in either tissue site was unaffected by genotype (P > 0.05); however, during chronic colitis, the percentage of Th17 cells residing in both the spleen and cLP was lower in Fat-1 mice compared to WT mice (P < 0.05). Colonic mucosal mRNA expression of critical Th17 cell cytokines and chemokine receptors (IL-17F, IL-21, and CCR6) were lower, whereas expression of the Th17 cell suppressive cytokine, IL-27, was greater in Fat-1 mice compared to WT mice during chronic colitis (P < 0.05). Moreover, colon histological scores were improved in Fat-1 mice (P < 0.05). Collectively, these results demonstrate for the first time, to our knowledge, that (n-3) PUFA can modulate the colonic mucosal microenvironment to suppress Th17 cell accumulation and inflammatory damage following the induction of chronic colitis.
Assuntos
Colite/imunologia , Ácidos Graxos Ômega-3/administração & dosagem , Células Th17/imunologia , Animais , Doença Crônica , Colite/metabolismo , Colite/patologia , Citocinas/metabolismo , Feminino , Citometria de Fluxo , Masculino , Camundongos , Camundongos MutantesRESUMO
Both fish oil (FO) and curcumin have potential as anti-tumour and anti-inflammatory agents. To further explore their combined effects on dextran sodium sulphate (DSS)-induced colitis, C57BL/6 mice were randomised to four diets (2 × 2 design) differing in fatty acid content with or without curcumin supplementation (FO, FO+2 % curcumin, maize oil (control, MO) or MO+2 % curcumin). Mice were exposed to one or two cycles of DSS in the drinking-water to induce either acute or chronic intestinal inflammation, respectively. FO-fed mice exposed to the single-cycle DSS treatment exhibited the highest mortality (40 %, seventeen of forty-three) compared with MO with the lowest mortality (3 %, one of twenty-nine) (P = 0·0008). Addition of curcumin to MO increased (P = 0·003) mortality to 37 % compared with the control. Consistent with animal survival data, following the one- or two-cycle DSS treatment, both dietary FO and curcumin promoted mucosal injury/ulceration compared with MO. In contrast, compared with other diets, combined FO and curcumin feeding enhanced the resolution of chronic inflammation and suppressed (P < 0·05) a key inflammatory mediator, NF-κB, in the colon mucosa. Mucosal microarray analysis revealed that dietary FO, curcumin and FO plus curcumin combination differentially modulated the expression of genes induced by DSS treatment. These results suggest that dietary lipids and curcumin interact to regulate mucosal homeostasis and the resolution of chronic inflammation in the colon.
Assuntos
Colite/dietoterapia , Colo/metabolismo , Curcumina/uso terapêutico , Citocinas/metabolismo , Suplementos Nutricionais , Óleos de Peixe/uso terapêutico , Regulação da Expressão Gênica , Doença Aguda , Animais , Anti-Inflamatórios não Esteroides/efeitos adversos , Anti-Inflamatórios não Esteroides/uso terapêutico , Doença Crônica , Colite/imunologia , Colite/metabolismo , Colite/patologia , Colo/efeitos dos fármacos , Colo/imunologia , Colo/patologia , Curcumina/efeitos adversos , Citocinas/genética , Sulfato de Dextrana/administração & dosagem , Sulfato de Dextrana/toxicidade , Óleos de Peixe/efeitos adversos , Perfilação da Expressão Gênica , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/imunologia , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Irritantes/administração & dosagem , Irritantes/toxicidade , Masculino , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/genética , NF-kappa B/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Distribuição Aleatória , Análise de SobrevidaRESUMO
We sought to develop a reversible staining protocol using micro-computed tomography (micro-CT) paired with a radiopaque contrast agent that allows for three-dimensional in situ visualization and characterization of atherosclerotic plaques. Atherosclerotic porcine coronary arteries were dissected from surrounding myocardium and incubated in iohexol at various concentrations and incubation times and then imaged using direct radiography. Line profiles were generated across the artery x-ray to determine effectiveness of the radiopaque contrast agent to penetrate the tissue. Our studies revealed that, to sufficiently delineate tissue constructs, the minimum effective iohexol concentration and incubation time were 240 mgI/mL for 1 hour. Among all groups, 24 hours of de-staining brought radiopacity back to control levels. After iohexol incubation, micro-CT was performed. Our findings demonstrate that extended staining times and a minimum iohexol concentration of 240 mgI/mL are required for effective tissue perfusion, which eliminates the diffusion distribution profile inherent to the ability of the contrast agent to traverse tissue layers.â¢Iohexol enhances ex vivo micro-CT imaging of atherosclerotic coronary arteriesâ¢Iohexol allows for improved tissue segmentation during micro-CT image analysisâ¢Effectiveness of iohexol penetration of the tissue was dependent on concentration and duration of incubation.
RESUMO
BACKGROUND: Duchenne muscular dystrophy (DMD) is an X-linked inherited myopathy that causes progressive skeletal and cardiac muscle disease. Heart lesions were described in the earliest DMD reports, and cardiomyopathy is now the leading cause of death. However, diagnostics and treatment for cardiomyopathy have lagged behind those for appendicular and respiratory skeletal muscle disease. Most animal model studies have been done in the mdx mouse, which has a relatively mild form of cardiomyopathy. Dogs with the genetically homologous condition, Golden Retriever muscular dystrophy (GRMD), develop progressive cardiomyopathy analogous to that seen in DMD. Previous descriptive studies of GRMD cardiomyopathy have mostly been limited to selective sampling of the hearts from young dogs. METHODS AND RESULTS: We systematically assessed cardiac lesions in 31 GRMD and carrier dogs aged 3 to 76 months and a separate cohort of 2-10-year-old normal hounds. Both semi-quantitative lesion scoring and quantitation of the cross-sectional area of fibrosis distinguished dogs with GRMD disease from normal dogs. The carriers generally had intermediate involvement but had even greater fibrosis than GRMD dogs. Fatty infiltration was the most prominent feature in some older GRMD dogs. Vascular hypertrophy was increased in GRMD dogs and correlated positively with lesion severity. Purkinje fiber vacuolation was also increased but did not correlate with lesion severity. Histopathologic changes correlated with late gadolinium enhancement on cardiac MRI. CONCLUSION: These features are generally compatible with those of DMD and further validate GRMD as a useful model to study cardiomyopathy pathogenesis and treatment. Additionally, the nature of some degenerative lesions suggests that functional hypoxia or non-thrombotic ischemia may contribute to disease progression.
RESUMO
BACKGROUND: Histology of thrombosis events in left ventricular assist devices (LVADs) may point to differences between the etiology of either ingested or de novo thrombus formation within LVADs. Materials ingested by the pump would have features suggestive of lifting and folding, whereas thrombi formed de novo would have uniform, parallel layers. This study tested this hypothesis in a cohort of explanted HeartWare Ventricular assist devices (HVADs) (Medtronic, Miami Lakes, Florida). METHODS: Histology of thrombi from 59 explanted HVAD pumps were classified as presumed ingested, presumed de novo, or undeterminable on the basis of pre-defined criteria. The apparent size and location of the thrombotic materials were noted. RESULTS: Histologically, all thrombotic materials were either presumed to be ingested (73%; 95 of 130 total histology cassettes examined) or of undeterminable origin (27%; 35 of 130 histology cassettes). Undetermined origin commonly was due to a lack of sufficient material for analysis. The larger materials (>800 mm3) tended to be in the inflow region. The most common finding was smaller thrombotic materials (<150 mm3) within the pump (64%; 38 of 59 HVADs); when these smaller materials were ingested by the pump, they were most often found within the smaller flow pathways within the pump. CONCLUSIONS: Our study suggests that the thrombi within HVAD pumps are commonly ingested materials rather than de novo thrombus formation within the pump. Further research to understand the source of this ingested material and the consideration to mitigate this complication should be considered.
Assuntos
Insuficiência Cardíaca/terapia , Coração Auxiliar/efeitos adversos , Trombose/etiologia , Falha de Equipamento , Humanos , Estudos RetrospectivosRESUMO
BACKGROUND AND AIMS: Visualization of arterial lesions in situ can enhance understanding of atherosclerosis progression and potentially improve experimental therapies. Conventional histology methods for assessing atherosclerotic lesions are robust but are destructive and may prevent further tissue analysis. The objective of the current study was to evaluate a novel, nondestructive method for visualization and characterization of atherosclerotic lesions as an alternative or complementary to routine histology. Thus, we tested the hypothesis that micro-computed tomography (micro-CT) paired with an iodine-based radiopaque stain would effectively characterize atherosclerotic plaques in a manner comparable to routine histology while maintaining sample integrity and providing whole-volume data. METHODS: We examined porcine coronary arteries with varying degrees of atherosclerosis, using micro-CT in the absence and presence of iohexol (240 mgI/ml). Following iohexol washout, routine histological assessment of the samples was performed with hematoxylin and eosin and Masson's trichrome. RESULTS: Iohexol staining generated soft tissue delineation and subsequent atherosclerotic plaque assessment via augmented radiopacity, permitting three-dimensional (3D) reconstruction of these lesions, maintaining in situ architecture. Although plaque distribution and arterial wall tissue layers were discernible, micro-CT was incapable of discriminating cell types comprising the plaque. Calcium phosphate deposition was readily located and visualized in 3D space, independent of iohexol. CONCLUSIONS: The results of this study establish micro-CT, combined with a diffusible radiopaque contrast agent, as a powerful imaging modality for visualizing in situ architecture of atherosclerotic plaques. Our findings demonstrate that micro-CT can be used to identify plaque distribution and calcium deposition complementary to routine histological analysis.
Assuntos
Iodo , Placa Aterosclerótica , Animais , Meios de Contraste , Vasos Coronários/diagnóstico por imagem , Placa Aterosclerótica/diagnóstico por imagem , Suínos , Microtomografia por Raio-XRESUMO
Distal limb wounds are common injuries sustained by horses and their healing is fraught with complications due to equine anatomy, prevalence of infection, and challenges associated with wound management. Gallium is a semi-metallic element that has been shown to possess antimicrobial properties and aid in wound healing in various preclinical models. The effects of Gallium have not been studied in equine wound healing. Therefore, the objective of this study was to compare healing rates between gallium-treated and untreated wounds of equine distal limbs and to demonstrate the antimicrobial effects of gallium on wounds inoculated with S. aureus. Using an established model of equine wound healing we demonstrated beneficial effects of 0.5% topical gallium maltolate on equine wound healing. Specifically we documented reduced healing times, reduced bioburden, and reduced formation of exuberant granulation tissue in wounds treated with gallium maltolate as compared with untreated wounds. Gallium appeared to exert its beneficial effects via its well-described antimicrobial actions as well as by altering the expression of specific genes known to be involved in wound healing of horses and other animals. Specifically, gallium maltolate appeared to increase expression of transforming growth factor-ß in both infected and un-infected wounds. Further work is needed to document the effects of gallium on naturally occurring equine wounds and to compare the effects of gallium with other wound treatment options. These data, however, suggest that gallium may be an attractive and novel means of improving equine distal limb wound healing.
Assuntos
Antibacterianos/uso terapêutico , Doenças dos Cavalos/tratamento farmacológico , Traumatismos da Perna/tratamento farmacológico , Compostos Organometálicos/uso terapêutico , Pironas/uso terapêutico , Infecções Estafilocócicas/tratamento farmacológico , Administração Tópica , Animais , Antibacterianos/administração & dosagem , Carga Bacteriana , Citocinas/genética , Citocinas/metabolismo , Doenças dos Cavalos/metabolismo , Cavalos , Traumatismos da Perna/metabolismo , Traumatismos da Perna/veterinária , Compostos Organometálicos/administração & dosagem , Pironas/administração & dosagem , Infecções Estafilocócicas/metabolismo , Infecções Estafilocócicas/veterinária , CicatrizaçãoRESUMO
We recently demonstrated that (n-3) PUFA trigger the induction of apoptosis in the colon by enhancing phospholipid oxidation and mitochondrial Ca2+ accumulation. To further elucidate the mechanisms regulating oxidative stress-induced apoptosis in vivo, a 2 x 2 experiment was designed using both wild type (control) and manganese-dependent superoxide dismutase (SOD2) heterozygous knockout mice (SOD2(+/-)), which exhibit increased mitochondrial oxidative stress. Mice were fed diets differing only in the type of fat [corn oil or fish oil containing (n-3) PUFA] at 15% by weight for 4 wk. Dietary (n-3) PUFA treatment enhanced (22%) apoptosis in colonic crypts. In addition, SOD2 haploinsufficiency enhanced (20%) apoptosis, which was further increased (36%) by (n-3) PUFA feeding. Dietary lipid source and genotype interactively modulated nitrotyrosine levels (P = 0.027) and inflammation (P = 0.032). These findings demonstrate that the proapoptotic effects of (n-3) PUFA are enhanced in oxidatively stressed SOD2(+/-) mice. Thus, (n-3) PUFA appear to promote an oxidation-reduction imbalance in the intestine, which may directly or indirectly trigger apoptosis and thereby reduce colon cancer risk.
Assuntos
Apoptose/efeitos dos fármacos , Colo/enzimologia , Ácidos Graxos Ômega-3/farmacologia , Superóxido Dismutase/deficiência , Animais , Colo/citologia , Colo/efeitos dos fármacos , Primers do DNA , Triagem de Portadores Genéticos , Imuno-Histoquímica , Mucosa Intestinal/citologia , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/enzimologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Superóxido Dismutase/genéticaRESUMO
Inflammatory bowel disease is a complex collection of disorders. Microbial dysbiosis as well as exposure to toxins including xenoestrogens are thought to be risk factors for inflammatory bowel disease development and relapse. Bisphenol-A has been shown to exert estrogenic activity in the colon and alter intestinal function, but the role that xenoestrogens, such as bisphenol-A , play in colonic inflammation has been previously described but with conflicting results. We investigated the ability of bisphenol-A to exacerbate colonic inflammation and alter microbiota metabolites derived from aromatic amino acids in an acute dextran sulfate sodium-induced colitis model. Female C57BL/6 mice were ovariectomized and exposed to bisphenol-A daily for 15 days. Disease activity measures include body weight, fecal consistency, and rectal bleeding. Colons were scored for inflammation, injury, and nodularity. Alterations in the levels of microbiota metabolites derived from aromatic amino acids known to reflect phenotypic changes in the gut microbiome were analyzed. Bisphenol-A exposure increased mortality and worsened disease activity as well as inflammation and nodularity scores in the middle colon region following dextran sulfate sodium exposure. Unique patterns of metabolites were associated with bisphenol-A consumption. Regardless of dextran sulfate sodium treatment, bisphenol-A reduced levels of tryptophan and several metabolites associated with decreased inflammation in the colon. This is the first study to show that bisphenol-A treatment alone can reduce microbiota metabolites derived from aromatic amino acids in the colon which may be associated with increased colonic inflammation and inflammatory bowel disease. Impact statement As rates of inflammatory bowel disease rise, discovery of the mechanisms related to the development of these conditions is important. Environmental exposure is hypothesized to play a role in etiology of the disease, as are alterations in the gut microbiome and the metabolites they produce. This study is the first to show that bisphenol-A alone alters tryptophan and microbiota metabolites derived from aromatic amino acids in a manner consistent with autoimmune diseases, specifically inflammatory bowel diseases, regardless of dextran sulfate sodium treatment. These findings indicate a potential mechanism by which bisphenol-A negatively affects gut physiology to exacerbate inflammation.
Assuntos
Aminoácidos Aromáticos/metabolismo , Compostos Benzidrílicos/metabolismo , Colite/patologia , Estrogênios não Esteroides/metabolismo , Microbioma Gastrointestinal/efeitos dos fármacos , Fenóis/metabolismo , Animais , Compostos Benzidrílicos/administração & dosagem , Colite/induzido quimicamente , Colo/patologia , Modelos Animais de Doenças , Estrogênios não Esteroides/administração & dosagem , Feminino , Camundongos Endogâmicos C57BL , Fenóis/administração & dosagem , Análise de SobrevidaRESUMO
Cardiovascular implantable electronic devices (CIEDs) typically incorporate leads that directly contact the endocardium. Post-explant pathology evaluation of formalin-fixed CIED lead implant sites and downstream organs (i.e., lungs) can provide useful safety data to the US Food and Drug Administration; however, current regulatory guidelines do not mandate how the safety data are collected. In this paper, we outline a protocol for preclinical pathology evaluation of leads associated with CIEDs, which includes formalin fixation of the heart and lungs, gross evaluation, and qualitative and quantitative histologic evaluation. We recommend fixation of the whole heart with leads in situ alongside intratracheal formalin infusion; this enables rapid and effective preservation of target tissues and increases histologic quality to allow for accurate qualitative and quantitative pathology evaluation. Overall, we believe that our approach to pathology evaluation of leads may maximize information acquired from preclinical studies, leading to more accurate safety assessments. SUMMARY: This article introduces an established method for pathology evaluation and analysis of cardiac leads recommended for companies and researchers that seek approval from a regulatory body.
Assuntos
Desfibriladores Implantáveis/efeitos adversos , Reação a Corpo Estranho/patologia , Pulmão/patologia , Miocárdio/patologia , Marca-Passo Artificial/efeitos adversos , Fixação de Tecidos/métodos , Animais , Remoção de Dispositivo , Segurança de Equipamentos , Fixadores/farmacologia , Reação a Corpo Estranho/diagnóstico por imagem , Formaldeído/farmacologia , Pulmão/diagnóstico por imagem , Microtomia , Modelos Animais , Inclusão em Parafina , Perfusão , Desenho de Prótese , Medição de Risco , Microtomografia por Raio-XRESUMO
Low dietary folate intake is associated with an increased risk for colon cancer; however, relevant genetic animal models are lacking. We therefore investigated the effect of targeted ablation of two folate transport genes, folate binding protein 1 (Folbp1) and reduced folate carrier 1 (RFC1), on folate homeostasis to elucidate the molecular mechanisms of folate action on colonocyte cell proliferation, gene expression, and colon carcinogenesis. Targeted deletion of Folbp1 (Folbp1(+/-) and Folbp1(-/-)) significantly reduced (P < 0.05) colonic Folbp1 mRNA, colonic mucosa, and plasma folate concentration. In contrast, subtle changes in folate homeostasis resulted from targeted deletion of RFC1 (RFC1(+/-)). These animals had reduced (P < 0.05) colonic RFC1 mRNA and exhibited a 2-fold reduction in the plasma S-adenosylmethionine/S-adenosylhomocysteine. Folbp1(+/-) and Folbp1(-/-) mice had larger crypts expressed as greater (P < 0.05) numbers of cells per crypt column relative to Folbp1(+/+) mice. Colonic cell proliferation was increased in RFC1(+/-) mice relative to RFC1(+/+) mice. Microarray analysis of colonic mucosa showed distinct changes in gene expression specific to Folbp1 or RFC1 ablation. The effect of folate transporter gene ablation on colon carcinogenesis was evaluated 8 and 38 weeks post-azoxymethane injection in wild-type and heterozygous mice. Relative to RFC1(+/+) mice, RFC1(+/-) mice developed increased (P < 0.05) numbers of aberrant crypt foci at 8 weeks. At 38 weeks, RFC1(+/-) mice developed local inflammatory lesions with or without epithelial dysplasia as well as adenocarcinomas, which were larger relative to RFC1(+/+) mice. In contrast, Folbp1(+/-) mice developed 4-fold (P < 0.05) more lesions relative to Folbp1(+/+) mice. In conclusion, Folbp1 and RFC1 genetically modified mice exhibit distinct changes in colonocyte phenotype and therefore have utility as models to examine the role of folate homeostasis in colon cancer development.
Assuntos
Proteínas de Transporte/genética , Transformação Celular Neoplásica/genética , Neoplasias do Colo/genética , Proteínas de Membrana Transportadoras/genética , Receptores de Superfície Celular/genética , Animais , Azoximetano , Carcinógenos , Proteínas de Transporte/antagonistas & inibidores , Proteínas de Transporte/biossíntese , Ciclo Celular/genética , Transformação Celular Neoplásica/induzido quimicamente , Transformação Celular Neoplásica/metabolismo , Colo/metabolismo , Colo/fisiologia , Neoplasias do Colo/induzido quimicamente , Neoplasias do Colo/metabolismo , Receptores de Folato com Âncoras de GPI , Perfilação da Expressão Gênica , Inativação Gênica , Predisposição Genética para Doença , Rim/metabolismo , Rim/fisiologia , Masculino , Moduladores de Transporte de Membrana , Proteínas de Membrana Transportadoras/antagonistas & inibidores , Proteínas de Membrana Transportadoras/biossíntese , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Análise de Sequência com Séries de Oligonucleotídeos , Lesões Pré-Cancerosas/induzido quimicamente , Lesões Pré-Cancerosas/genética , Lesões Pré-Cancerosas/metabolismo , Receptores de Superfície Celular/antagonistas & inibidores , Receptores de Superfície Celular/biossíntese , Proteína Carregadora de Folato Reduzido , Reação em Cadeia da Polimerase Via Transcriptase Reversa , S-Adenosil-Homocisteína/sangue , S-Adenosil-Homocisteína/metabolismo , S-Adenosilmetionina/sangue , S-Adenosilmetionina/metabolismoRESUMO
Non-steroidal anti-inflammatory drugs (NSAIDs) are among the most frequently used classes of medications in the world, yet they induce an enteropathy that is associated with high morbidity and mortality. A major limitation to better understanding the pathophysiology and diagnosis of this enteropathy is the difficulty of obtaining information about the primary site of injury, namely the distal small intestine. We investigated the utility of using mRNA from exfoliated cells in stool as a means to surveil the distal small intestine in a murine model of NSAID enteropathy. Specifically, we performed RNA-Seq on exfoliated cells found in feces and compared these data to RNA-Seq from both the small intestinal mucosa and colonic mucosa of healthy control mice or those exhibiting NSAID-induced enteropathy. Global gene expression analysis, data intersection, pathway analysis, and computational approaches including linear discriminant analysis (LDA) and sparse canonical correlation analysis (CCA) were used to assess the inter-relatedness of tissue (invasive) and stool (noninvasive) datasets. These analyses revealed that the exfoliated cell transcriptome closely mirrored the transcriptome of the small intestinal mucosa. Thus, the exfoliome may serve as a non-invasive means of detecting and monitoring NSAID enteropathy (and possibly other gastrointestinal mucosal inflammatory diseases).