In vitro influence of the extracellular matrix in myoepithelial cells stimulated by malignant conditioned medium

In order to investigate the role of myoepithelial cell and tumor microenvironment in salivary gland neoplasma, we have performed a study towards the effect of different extracellular matrix proteins (basement membrane matrix, type I collagen and fibronectin) on morphology and differentiation of benign myoepithelial cells from pleomorphic adenoma cultured with malignant cell culture medium from squamous cell carcinoma. We have also analyzed the expression of alpha-smooth muscle actin (alpha-SMA) and FGF-2 by immunofluorescence and qPCR. Our immunofluorescence results, supported by qPCR analysis, demonstrated that alpha-SMA and FGF-2 were upregulated in the benign myoepithelial cells from pleomorphic adenoma in all studied conditions on fibronectin substratum. However, the myoepithelial cells on fibronectin substratum did not alter their morphology under malignant conditioned medium stimulation and exhibited a stellate morphology and, occasionally focal adhesions with the substratum. In summary, our data demonstrated that the extracellular matrix exerts an important role in the morphology of the benign myoepithelial cells by the presence of focal adhesions and also inducing increase FGF-2 and alpha-SMA expression by these cells, especially in the fibronectin substratum. (C) 2011 Elsevier Ltd. All rights reserved.

The polysaccharide fraction of Propionibacterium acnes modulates the development of experimental focal segmental glomerulosclerosis

The pathogenesis of focal segmental glomerulosclerosis (FSGS) appears to be associated with type-2 cytokines and podocyte dysfunction. In this study, we tested the hypothesis that immunization with the polysaccharide fraction of Propionibacterium acnes (PS), a pro-Th1 agonist, may subvert the type-2 profile and protect podocytes from adriamycin-induced glomerulosclerosis. Adriamycin injection resulted in albuminuria and increased serum creatinine in association with loss of glomerular podocin and podoplanin expression, which is consistent with podocyte dysfunction. Renal tissue analysis revealed the expression of transcripts for GATA3 and fibrogenic-related proteins, such as TGF-beta, tissue inhibitor of metalloproteinase-1 (TIMP-1) and metalloproteinase 9 (MMP9). In association with the expression of fibrogenic transcripts, we observed peri-glomerular expression of a-smooth muscle actin (alpha-SMA), indicating epithelial-to-mesenchymal transition, and increased expression of proliferating cell nuclear antigen (PCNA) in tubular cells, suggesting intense proliferative activity. Previous immunization with PS inhibited albuminuria and serum creatinine in association with the preservation of podocyte proteins and inhibition of fibrogenic transcripts and the expression of alpha-SMA and PCNA proteins. Tissue analysis also revealed that PS treatment induced expression of mRNA for GD3 synthase, which is a glycosiltransferase related to the synthesis of GD3, a ganglioside associated with podocyte physiology. In addition, PS treatment inhibited the influx of inflammatory CD8(pos) and CD11b(pos) cells to kidney tissue. Finally, PS treatment on day 4 post-ADM, a period when proteinuria was already established, was able to improve renal function. Thus, we demonstrate that the PS fraction of P. acnes can inhibit FSGS pathogenesis, suggesting that immunomodulation can represent an alternative approach for disease management. (C) 2011 Elsevier GmbH. All rights reserved.

Antifibrotic Effect of Tamoxifen in a Model of Progressive Renal Disease

Tamoxifen, a selective estrogen receptor modulator, has antifibrotic properties; however, whether it can attenuate renal fibrosis is unknown. In this study, we tested the effects of tamoxifen in a model of hypertensive nephrosclerosis (chronic inhibition of nitric oxide synthesis with L-NAME). After 30 days, treated rats had significantly lower levels of albuminuria as well as lower histologic scores for glomerulosclerosis and interstitial fibrosis than untreated controls. Tamoxifen was renoprotective despite having no effect on the sustained, severe hypertension induced by L-NAME. Tamoxifen prevented the accumulation of extracellular matrix by decreasing the expression of collagen I, collagen III, and fibronectin mRNA and protein. These renoprotective effects associated with inhibition of TGF-beta 1 and plasminogen activator inhibitor-1, and with a significant reduction in a-smooth muscle actin-positive cells in the renal interstitium. Furthermore, tamoxifen abrogated IL-1 beta- and angiotensin-II-induced proliferation of fibroblasts from both kidney explants and from the NRK-49F cell line. Tamoxifen also inhibited the expression of extracellular matrix components and the production and release of TGF-beta 1 into the supernatant of these cells. In summary, tamoxifen exhibits antifibrotic effects in the L-NAME model of hypertensive nephrosclerosis, likely through the inhibition of TGF-beta 1, suggesting that it may have therapeutic use in CKD treatment.

In vivo hydroquinone exposure causes tracheal hyperresponsiveness due to TNF secretion by epithelial cells

Hydroquinone (HQ) is the main oxidative substance in cigarette smoke and a toxic product of benzene biotransformation. Although the respiratory tract is an inlet pathway of HQ exposure, its effect on airway muscle responsiveness has not been assessed. We thus investigated the effects of low dose in vivo HQ-exposure on tracheal responsiveness to a muscarinic receptor agonist. Male Swiss mice were exposed to aerosolised 5% ethanol/saline solution (HQ vehicle; control) or 0.04 ppm HQ (1 h/day for 5 days) and tracheal rings were collected 1 h after the last exposure. HQ exposure caused tracheal hyper-responsiveness to methacholine (MCh), which was abolished by mechanical removal of the epithelium. This hyperresponsiveness was not dependent on neutrophil infiltration, but on tumour necrosis factor (TNF) secretion by epithelial cells. This conclusion was based on the following data: (1) trachea from HQ-exposed mice presented a higher amount of TNF, which was abrogated following removal of the epithelium; (2) the trachea hyperresponsiveness and TNF levels were attenuated by in vivo chlorpromazine (CPZ) treatment, an inhibitor of TNF synthesis. The involvement of HQ-induced TNF secretion in trachea mast cell degranulation was also demonstrated by the partial reversion of tracheal hyperresponsiveness in sodium cromoglicate-treated animals, and the in vivo HQ-exposure-induced degranulation of trachea connective tissue and mucosal mast cells, which was reversed by CPZ treatment. Our data show that in vivo HQ exposure indirectly exacerbates the parasympathetic-induced contraction of airway smooth muscle cells, mediated by TNF secreted by tracheal epithelial cells, clearly showing the link between environmental HQ exposure and the reactivity of airways. (C) 2012 Elsevier Ireland Ltd. All rights reserved.

Rho-kinase inhibition attenuates airway responsiveness, inflammation, matrix remodeling, and oxidative stress activation induced by chronic inflammation

Possa SS, Charafeddine HT, Righetti RF, da Silva PA, Almeida-Reis R, Saraiva-Romanholo BM, Perini A, Prado CM, Leick-Maldonado EA, Martins MA, Tiberio ID. Rho-kinase inhibition attenuates airway responsiveness, inflammation, matrix remodeling, and oxidative stress activation induced by chronic inflammation. Am J Physiol Lung Cell Mol Physiol 303: L939-L952, 2012. First published September 21, 2012; doi:10.1152/ajplung.00034.2012.-Several studies have demonstrated the importance of Rho-kinase in the modulation of smooth muscle contraction, airway hyperresponsiveness, and inflammation. However, the effects of repeated treatment with a specific inhibitor of this pathway have not been previously investigated. We evaluated the effects of repeated treatment with Y-27632, a highly selective Rho-kinase inhibitor, on airway hyperresponsiveness, oxidative stress activation, extracellular matrix remodeling, eosinophilic inflammation, and cytokine expression in an animal model of chronic airway inflammation. Guinea pigs were subjected to seven ovalbumin or saline exposures. The treatment with Y-27632 (1 mM) started at the fifth inhalation. Seventy-two hours after the seventh inhalation, the animals' pulmonary mechanics were evaluated, and exhaled nitric oxide (E-NO) was collected. The lungs were removed, and histological analysis was performed using morphometry. Treatment with Y-27632 in sensitized animals reduced E-NO concentrations, maximal responses of resistance, elastance of the respiratory system, eosinophil counts, collagen and elastic fiber contents, the numbers of cells positive for IL-2, IL-4, IL-5, IL-13, inducible nitric oxide synthase, matrix metalloproteinase-9, tissue inhibitor of metalloproteinase-1, transforming growth factor-beta, NF-kappa B, IFN-gamma, and 8-iso-prostaglandin F2 alpha contents compared with the untreated group (P < 0.05). We observed positive correlations among the functional responses and inflammation, remodeling, and oxidative stress pathway activation markers evaluated. In conclusion, Rho-kinase pathway activation contributes to the potentiation of the hyperresponsiveness, inflammation, the extracellular matrix remodeling process, and oxidative stress activation. These results suggest that Rho-kinase inhibitors represent potential pharmacological tools for the control of asthma.

Vascular dysfunction by myofibroblast activation in patients with idiopathic pulmonary fibrosis and prognostic significance

In this study, we demonstrated the importance of telomerase protein expression and determined the relationships among telomerase, endothelin-1 (ET-1) and myofibroblasts during early and late remodeling of parenchymal and vascular areas in usual interstitial pneumonia (UIP) using 27 surgical lung biopsies from patients with idiopathic pulmonary fibrosis (IPF). Telomerase+, myofibroblasts alpha-SMA+, smooth muscle cells caldesmon+, endothelium ET-1+ cellularity, and fibrosis severity were evaluated in 30 fields covering normal lung parenchyma, minimal fibrosis (fibroblastic foci), severe ( mural) fibrosis, and vascular areas of UIP by the point-counting technique and a semiquantitative score. The impact of these markers was determined in pulmonary functional tests and follow-up until death from IPF. Telomerase and ET-1 expression was significantly increased in normal and vascular areas compared to areas of fibroblast foci. Telomerase and ET-1 expression was inversely correlated with minimal fibrosis in areas of fibroblast foci and directly associated with severe fibrosis in vascular areas. Telomerase activity in minimal fibrosis areas was directly associated with diffusing capacity of the lung for oxygen/alveolar volume and ET-1 expression and indirectly associated with diffusing capacity of the lungs for carbon monoxide and severe fibrosis in vascular areas. Cox proportional hazards regression revealed a low risk of death for females with minimal fibrosis displaying high telomerase and ET-1 expression in normal areas. Vascular dysfunction by telomerase/ET-1 expression was found earlier than vascular remodeling by myofibroblast activation in UIP with impact on IPF evolution, suggesting that strategies aimed at preventing the effect of these mediators may have a greater impact on patient outcome.

Association between the C242T polymorphism in the p22phox gene with arterial stiffness in the Brazilian population

de Oliveira Alvim R, Lima Santos PCJ, Goncalves Dias R, Rodrigues MV, de Sa Cunha R, Mill JG, Junior WN, Krieger JE, Pereira AC. Association between the C242T polymorphism in the p22phox gene with arterial stiffness in the Brazilian population. Physiol Genomics 44: 587-592, 2012. First published April 10, 2012; doi:10.1152/physiolgenomics.00122.2011.-NADPH oxidase p22phox subunit is responsible for the production of reactive oxygen species in the vascular tissue. The C242T polymorphism in the p22phox gene has been associated with diverse coronary artery disease phenotypes, but the findings about the protective or harmful effects of the T allele are still controversial. Our main aim was to assess the effect of p22phox C242T genotypes on arterial stiffness, a predictor of late morbidity and mortality, in individuals from the general population. We randomly selected 1,178 individuals from the general population of Vitoria City, Brazil. Genotypes for the C242T polymorphism were detected by PCR-RFLP, and pulse wave velocity (PWV) values were measured with a noninvasive automatic device Complior. p22phox and TNF-alpha gene expression were quantified by real-time PCR in human arterial mammary smooth muscle cells. In both the entire and nonhypertensive groups: individuals carrying the TT genotype had higher PWV values and higher risk for increased arterial stiffness [odds ratio (OR) 1.93, 95% confidence interval (CI) 1.27-2.92 and OR 1.78, 95% CI 1.07-2.95, respectively] compared with individuals carrying CC + CT genotypes, even after adjustment for covariates. No difference in the p22phox gene expression according C242T genotypes was observed. However, TNF-alpha gene expression was higher in cells from individual carrying the T allele, suggesting that this genetic marker is associated with functional phenotypes at the gene expression level. In conclusion, we suggest that p22phox C242T polymorphism is associated with arterial stiffness evaluated by PWV in the general population. This genetic association shed light on the understanding of the genetic modulation on vascular dysfunction mediated by NADPH oxidase.

Using gene-network landscape to dissect genotype effects of TCF7L2 genetic variant on diabetes and cardiovascular risk

Vaquero AR, Ferreira NE, Omae SV, Rodrigues MV, Teixeira SK, Krieger JE, Pereira AC. Using gene-network landscape to dissect genotype effects of TCF7L2 genetic variant on diabetes and cardiovascular risk. Physiol Genomics 44: 903-914, 2012. First published August 7, 2012; doi:10.1152/physiolgenomics.00030.2012.-The single nucleotide polymorphism (SNP) within the TCF7L2 gene, rs7903146, is, to date, the most significant genetic marker associated with Type 2 diabetes mellitus (T2DM) risk. Nonetheless, its functional role in disease pathology is poorly understood. The aim of the present study was to investigate, in vascular smooth muscle cells from 92 patients undergoing aortocoronary bypass surgery, the contribution of this SNP in T2DM using expression levels and expression correlation comparison approaches, which were visually represented as gene interaction networks. Initially, the expression levels of 41 genes (seven TCF7L2 splice forms and 40 other T2DM relevant genes) were compared between rs7903146 wild-type (CC) and T2DM-risk (CT + TT) genotype groups. Next, we compared the expression correlation patterns of these 41 genes between groups to observe if the relationships between genes were different. Five TCF7L2 splice forms and nine genes showed significant expression differences between groups. RXR alpha gene was pinpointed as showing the most different expression correlation pattern with other genes. Therefore, T2DM risk alleles appear to be influencing TCF7L2 splice form's expression in vascular smooth muscle cells, and RXR alpha gene is pointed out as a treatment target candidate for risk reduction in individuals with high risk of developing T2DM, especially individuals harboring TCF7L2 risk genotypes.

Treatment With Methotrexate Inhibits Atherogenesis in Cholesterol-Fed Rabbits

A decrease in the number of cardiovascular events in patients with rheumatoid arthritis or psoriasis treated with methotrexate (MTX) has been observed in the literature. The aim of this study was to test whether MTX could promote anti-inflammatory effects and reduce the atherosclerotic lesions in rabbits with atherosclerosis induced by cholesterol feeding. Twenty male New Zealand rabbits were fed a 1% cholesterol diet for 60 days. Starting from day 30 of cholesterol feeding, 10 animals were treated with 4 weekly intravenous injections of MTX (4 mg/kg) and 10 with 4 weekly saline solution injections for 30 days. MTX reduced the size of the lesion areas of cholesterol-fed animals by 75% and intima-media ratio 2- fold. The drug inhibited macrophage migration into the intima by 50% and the presence of apoptotic cells by 84% but did not inhibit the intimal proliferation of smooth muscle cells. MTX treatment also diminished the positive staining area of metalloproteinase 9 in the intima, which is probably beneficial. In the tumor necrosis factor-alpha-treated human umbilical vein endothelial cell line, incubation with MTX led to downregulation of 5 pro-inflammatory genes, TNF-alpha, VAP-1, IL-1 beta, CXCL2, and TLR2, and upregulation of the antiinflammatory TGF-beta 1 gene, thus showing endothelium-protective properties. In conclusion, MTX showed direct in vivo anti-atherosclerotic action and may have potential in the treatment of this disorder.

Omega-6 polyunsaturated fatty acids prevent atherosclerosis development in LDLr-KO mice, in spite of displaying a pro-inflammatory profile similar to trans fatty acids

The development of atherosclerosis and the inflammatory response were investigated in LDLr-KO mice on three high-fat diets (40% energy as fat) for 16 weeks: trans (TRANS), saturated (SAFA) or omega-6 polyunsaturated (PUFA) fats. The following parameters were measured: plasma lipids, aortic root total cholesterol (TC), lesion area (Oil Red-O), ABCA1 content and macrophage infiltration (immunohistochemistry), collagen content (Picrosirius-red) and co-localization of ABCA1 and macrophage (confocal microscopy) besides the plasma inflammatory markers (IL-6, TNF-alpha) and the macrophage inflammatory response to lipopolysaccharide from Escherichia coli (LPS). As expected, plasma TC and TG concentrations were lower on the PUFA diet than on TRANS or SAFA diets. Aortic intima macrophage infiltration, ABCA1 content, and lesion area on PUFA group were lower compared to TRANS and SAFA groups. Macrophages and ABCA1 markers did not co-localize in the atherosclerotic plaque, suggesting that different cell types were responsible for the ABCA1 expression in plaques. Compared to PUFA, TRANS and SAFA presented higher collagen content and necrotic cores in atherosclerotic plaques. In the artery wall, TC was lower on PUFA compared to TRANS group; free cholesterol was lower on PUFA compared to TRANS and SAFA; cholesteryl ester concentration did not vary amongst the groups. Plasma TNF-alpha concentration on PUFA and TRANS-fed mice was higher compared to SAFA. No difference was observed in IL-6 concentration amongst groups. Regarding the macrophage inflammatory response to LPS, TRANS and PUFA presented higher culture medium concentrations of IL-6 and TNF-alpha as compared to SAFA. The PUFA group showed the lowest amount of the anti-inflammatory marker IL-10 compared to TRANS and SAFA groups. In conclusion, PUFA intake prevented atherogenesis, even in a pro-inflammatory condition. (c) 2012 Elsevier Ireland Ltd. All rights reserved.

Involvement of nitric oxide on the pathogenesis of irinotecan-induced intestinal mucositis: role of cytokines on inducible nitric oxide synthase activation

Purpose Intestinal mucositis and the closely associated diarrhea are common costly side effects of irinotecan. Cytokine modulators, such as thalidomide and pentoxifylline, are found capable of attenuating intestinal mucositis progression. Nitric oxide (NO) seems to be a key mediator of the antineoplastic drug toxicity. The aim of this study was to investigate the role of NO on the pathogenesis of intestinal mucositis, as well as the participation of cytokines upon inducible nitric oxide synthase (iNOS) expression in irinotecan-induced intestinal mucositis. Methods iNOS-knockout (iNOS(-/-)) and C57BL/6 (WT, wild type) animals (n = 5-6) were given either saline or irinotecan (60 mg/kg i.p for 4 days), with or without pretreatment with aminoguanidine (50 mg/kg s.c.), thalidomide (60 mg/kg s.c), infliximab (5 mg/kg i.v.), or pentoxifylline (1.7 mg/kg s.c). On day 5, diarrhea was assessed, and following euthanasia, proximal intestinal samples were obtained for myeloperoxidase (MPO) and iNOS activity, morphometric analysis, western blot and immunohistochemistry to iNOS, cytokine dosage, and for in vitro evaluation of gut contractility. Results Irinotecan induced severe diarrhea and intestinal smooth muscle over-contractility, accompanied with histopathological changes. Additionally, increased MPO and iNOS activity and iNOS immunoexpression were found in WT animals treated with irinotecan. The rise in MPO, smooth muscle over-contractility, and diarrhea were abrogated in aminoguanidine-treated and iNOS(-/-) mice. Moreover, through western blot, we verified that infliximab and pentoxifylline significantly inhibited irinotecan-induced iNOS expression. In addition, cytokine concentration was found only partially decreased in irinotecan-treated iNOS(-/-) mice when compared with wild-type animals that were given irinotecan. Conclusions This study suggests a role of nitric oxide in the pathogenesis of irinotecan-induced intestinal mucositis and also provides evidence for the participation of cytokines on iNOS induction.

Alterations in vasoconstrictor responses to the endothelium-derived contracting factor uridine adenosine tetraphosphate are region specific in DOCA-salt hypertensive rats

Uridine adenosine tetraphosphate (Up(4)A) has been recently identified as a novel and potent endothelium-derived contracting factor and contains both purine and pyrimidine moieties, which activate purinergic P2X and P2Y receptors. The present study was designed to compare contractile responses to Up(4)A and other nucleotides such as ATP (P2X/P2Y agonist), UTP (P2Y(2)/P2Y(4) agonist), UDP (P2Y(6) agonist), and alpha,beta-methylene ATP (P2X(1) agonist) in different vascular regions [thoracic aorta, basilar, small mesenteric, and femoral arteries] from deoxycorticosterone acetate-salt (DOCA-salt) and control rats. In DOCA-salt rats [vs. control uninephrectomized (Uni) rats]: (1) in thoracic aorta, Up(4)A-, ATP-, and UP-induced contractions were unchanged; (2) in basilar artery, Up(4)A-, ATP-, UTP- and UDP-induced contractions were increased, and expression for P2X(1), but not P2Y(2) or P2Y(6) was decreased; (3) in small mesenteric artery, Up(4)A-induced contraction was decreased and UDP-induced contraction was increased; expression of P2Y(2) and P2X(1) was decreased whereas P2Y(6) expression was increased; (4) in femoral artery, Up(4)A-. UTP-, and UDP-induced contractions were increased, but expression of P2Y(2), P2Y(6) and P2X(1) was unchanged. The alpha,beta-methylene ATP-induced contraction was bell-shaped and the maximal contraction was reached at a lower concentration in basilar and mesenteric arteries from Uni rats, compared to arteries from DOCA-salt rats. These results suggest that Up(4)A-induced contraction is heterogenously affected among various vascular beds in arterial hypertension. P2Y receptor activation may contribute to enhancement of Up(4)A-induced contraction in basilar and femoral arteries. These changes in vascular reactivity to Up(4)A may be adaptive to the vascular alterations produced by hypertension. (C) 2011 Elsevier Ltd. All rights reserved.

Immunohistochemical evidence for myofibroblastlike cells associated with liver injury induced by Aflatoxin B1 in rainbow trout (Oncorhynchus mykiss)

In mammalian species, profibrogenic cells are activated to become myofibroblasts in response to liver damage. Few studies have examined hepatic myofibroblasts and their role in liver damage in teleosts. The aim of the present study was to investigate the involvement of myofibroblast-like cells in rainbow trout (Oncorhynchus mykiss) with hepatic damage induced by aflatoxin B1 (AFB1). Histopathological and immunohistochemical analyses characterized alterations in the liver stroma during the carcinogenic process. Anti-human a-smoothmuscle actin (SMA) and anti-human desmin primary antibodies were used in immunohistochemistry. Only the anti-SMA reagent labelled cells in trout liver. In the livers of control fish, only smooth muscle in blood vessels and around bile ducts was labelled. In the livers from AFB1-treated fish, SMA-positive cells were present in the stroma surrounding neoplastic lesions and in areas of desmoplastic reaction. These observations indicate that in teleosts, as in mammals, the myofibroblast-like cell is involved in fibrosis associated with liver injury. Chronic liver injury induced in trout by aflatoxin may provide a useful model system for study of the evolution of such mechanisms.

Quantification of cellular action fibers alignment from confocal microscopic images.

The cellular rheology has recently undergone a rapid development with particular attention to the cytoskeleton mechanical properties and its main components - actin filaments, intermediate filaments, microtubules and crosslinked proteins. However it is not clear what are the cellular structural changes that directly affect the cell mechanical properties. Thus, in this work, we aimed to quantify the structural rearrangement of these fibers that may emerge in changes in the cell mechanics. We created an image analysis platform to study smooth muscle cells from different arteries: aorta, mammary, renal, carotid and coronary and processed respectively 31, 29, 31, 30 and 35 cell image obtained by confocal microscopy. The platform was developed in Matlab (MathWorks) and it uses the Sobel operator to determine the actin fiber image orientation of the cell, labeled with phalloidin. The Sobel operator is used as a filter capable of calculating the pixel brightness gradient, point to point, in the image. The operator uses vertical and horizontal convolution kernels to calculate the magnitude and the angle of the pixel intensity gradient. The image analysis followed the sequence: (1) opens a given cells image set to be processed; (2) sets a fix threshold to eliminate noise, based on Otsu's method; (3) detect the fiber edges in the image using the Sobel operator; and (4) quantify the actin fiber orientation. Our first result is the probability distribution II(Δθ) to find a given fiber angle deviation (Δθ) from the main cell fiber orientation θ0. The II(Δθ) follows an exponential decay II(Δθ) = Aexp(-αΔθ) regarding to its θ0. We defined and determined a misalignment index α of the fibers of each artery kind: coronary αCo = (1.72 ‘+ or =’ 0.36)rad POT -1; renal αRe = (1.43 + or - 0.64)rad POT -1; aorta αAo = (1.42 + or - 0.43)rad POT -1; mammary αMa = (1.12 + or - 0.50)rad POT -1; and carotid αCa = (1.01 + or - 0.39)rad POT -1. The α of coronary and carotid are statistically different (p < 0.05) among all analyzed cells. We discussed our results correlating the misalignment index data with the experimental cell mechanical properties obtained by using Optical Magnetic Twisting Cytometry with the same group of cells.

Hypervitaminosis A-induced hepatic fibrosis in a cat

Rationale: The excessive intake of vitamin A in the form of vitamin concentrate, supplement or vitamin-rich liver can result in hypervitaminosis A in man and animals. Although osteopathologies resulting from chronic vitamin A intoxication in cats are well characterized, no information is available concerning feline hypervitaminosis A-induced liver disease. Clinical summary: We report the first case of hepatic stellate cell lipidosis and hepatic fibrosis in a domestic cat that had been fed a diet based on raw beef liver. Radiographic examination revealed exostoses and ankylosis between vertebrae C1 and T7, compatible with deforming cervical spondylosis. Necropsy showed a slightly enlarged and light yellow to bronze liver. Microscopic and ultrastructural analyses of liver tissues revealed diffuse and severe liver fibrosis associated with hepatic stellate cell hyperplasia and hypertrophy. These cells showed immunopositive staining for α-smooth muscle actin and desmin markers. The necropsy findings of chronic liver disease coupled with osteopathology supported the diagnosis of hypervitaminosis A. Practical relevance: As in human hepatology, if there is dietary evidence to support increased intake of vitamin A, then hypervitaminosis A should be considered in the differential diagnosis of chronic liver disease in cats.