Decrease In Collagen Content Research Articles

Histotripsy of the Prostate in a Canine Model: Characterization of Post-Therapy Inflammation and Fibrosis.

Histotripsy is a nonthermal, noninvasive, pulsed ultrasound technology that homogenizes tissue within the targeted volume. From previous experiments, it appeared that the resultant fibrotic response from histotripsy was limited compared with the typical tissue response seen after thermoablation. The objective of this study was to characterize the inflammatory response and quantify patterns of collagen deposition 6 weeks after in vivo canine prostate histotripsy. Histotripsy was applied to the left half of eight canine prostates to produce an intraparenchymal zone of tissue homogenization. Six weeks after treatment, prostates were harvested, sectioned, and stained with hematoxylin and eosin for histologic evaluation, CD3, CD20, and Mac387 immunohistochemistry to characterize the inflammatory components, and picrosirius red staining to identify collagen. Seven of eight treated prostates exhibited only minimal residual inflammation. Visual microscopic analysis of picrosirius red slides revealed a band of dense collagen (0.5 mm wide) immediately adjacent to the cavity produced by histotripsy. This was surrounded by a second band (1 mm wide) of less dense collagen interspersed among glandular architecture. A lobar distribution of epithelial atrophy and basal cell hyperplasia reminiscent of periurethral glands and ducts was apparent surrounding the margin of the treatment cavities. Tissue loss (-31%) was apparent on the treated side of all prostates while four demonstrated a net decrease in collagen content. In vivo histotripsy of canine prostate produced a decrease in prostate volume coupled with a limited inflammatory and fibrotic response. A narrow (1.5 mm) band of fibrosis around the empty, reepithelialized treatment cavity was observed 6 weeks after treatment. In four cases, an overall reduction in collagen content was measured. Further studies are planned to correlate these histologic findings with alteration in mechanical tissue properties and to explore histotripsy strategies for treatment of benign prostatic hyperplasia that optimize tissue volume removal with minimization of fibrosis.

Diminazene protects corpus cavernosum against hypercholesterolemia-induced injury.

Angiotensin-converting enzyme 2 (ACE2) is a key enzyme of the renin angiotensin system, which breaks down angiotensin II and forms angiotensin-(1-7). In erectile tissues, it has been documented that angiotensin II contributes to the development of erectile dysfunction (ED), while treatment with angiotensin-(1-7) improves penile erection. However, the expression and function of ACE2 in erectile tissues have never been investigated. Here, we examined the expression of ACE2 in erectile tissues and its actions against hypercholesterolemia-induced corpus cavernosum (CC) injury. Hypercholesterolemic apolipoprotein E knockout (ApoE(-/-) ) mice, a well-known model of ED, were treated with diminazene aceturate (DIZE), an ACE2 activator compound, or vehicle for 3 weeks. Reactive oxygen species (ROS), collagen content, and protein expression of ACE2, neuronal nitric oxide synthase (nNOS), endothelial nitric oxide synthase (eNOS), nicotinamide adenine dinucleotide phosphate-oxidase (NADPH) subunits were evaluated in the penis of DIZE-treated and untreated ApoE(-/-) mice. Functional studies were performed in CC strips. ACE2 expression and its role in modulating nitric oxide (NO)/ROS production and fibrosis within the CC of hypercholesterolemic mice were the main outcome measures. ACE2 was expressed in smooth muscle and endothelial cells of mouse CC. Interestingly, ACE2 was downregulated in penis of hypercholesterolemic mice with ED, suggesting a protective role of ACE2 on the CC homeostasis. In accordance with that, pharmacological ACE2 activation by DIZE treatment reduced ROS production and NADPH oxidase expression, and elevated nNOS and eNOS expression and NO bioavailability in the penis of ApoE(-/-) mice. Additionally, DIZE decreased collagen content within the CC. These beneficial actions of DIZE on the CC were not accompanied by improvements in atherosclerotic plaque size or serum lipid profile. ACE2 is expressed in erectile tissue and its reduction is associated with hypercholesterolemia-induced ED. Additionally, treatment with DIZE improved hypercholesterolemia-induced CC injury, suggesting ACE2 as a potential target for treating ED. .

Revealing cytokine-induced changes in the extracellular matrix with secondary ion mass spectrometry

Cell-secreted matrices (CSMs), where extracellular matrix (ECM) deposited by monolayer cell cultures is decellularized, have been increasingly used to produce surfaces that may be reseeded with cells. Such surfaces are useful to help us understand cell–ECM interactions in a microenvironment closer to the in vivo situation than synthetic substrates with adsorbed proteins. We describe the production of CSMs from mouse primary osteoblasts (mPObs) exposed to cytokine challenge during matrix secretion, mimicking in vivo inflammatory environments. Time-of-flight secondary ion mass spectrometry data revealed that CSMs with cytokine challenge at day 7 or 12 of culture can be chemically distinguished from one another and from untreated CSM using multivariate analysis. Comparison of the differences with reference spectra from adsorbed protein mixtures points towards cytokine challenge resulting in a decrease in collagen content. This is supported by immunocytochemical and histological staining, demonstrating a 44% loss of collagen mass and a 32% loss in collagen I coverage. CSM surfaces demonstrate greater cell adhesion than adsorbed ECM proteins. When mPObs were reseeded onto cytokine-challenged CSMs they exhibited reduced adhesion and elongated morphology compared to untreated CSMs. Such changes may direct subsequent cell fate and function, and provide insights into pathological responses at sites of inflammation.

Aspirin-triggered 15-epi-lipoxin A4 signals through FPR2/ALX in vascular smooth muscle cells and protects against intimal hyperplasia after carotid ligation

Aspirin-triggered 15-epi-lipoxin A4 signals through FPR2/ALX in vascular smooth muscle cells and protects against intimal hyperplasia after carotid ligation

The role of the FPR2/ALX receptor in atherosclerosis development and plaque stability.

AimsThe formyl peptide receptor (FPR) subtype FPR2/ALX transduces pro-inflammatory responses and participates in the resolution of inflammation depending on activation. The aim of the present study was to unravel the role of FPR2/ALX signalling in atherosclerosis.Methods and resultsExpression of FPR2/ALX was analysed in 127 human carotid atherosclerotic lesions and revealed that this receptor was expressed on macrophages, smooth muscle cells (SMCs), and endothelial cells. Furthermore, FPR2/ALX mRNA levels were significantly up-regulated in atherosclerotic lesions compared with healthy vessels. In multiple regression, age, creatinine, and clinical signs of increased cerebral ischaemia were independent predictors of FPR2/ALX expression. To provide mechanistic insights into these observations, we generated Ldlr−/−xFpr2−/− mice, which exhibited delayed atherosclerosis development and less macrophage infiltration compared with Ldlr−/−xFpr2+/+ mice. These findings were reproduced by transplantation of Fpr2−/− bone marrow into Ldlr−/− mice and further extended by in vitro experiments, demonstrating a lower inflammatory state in Fpr2−/− macrophages. FPR2/ALX expression correlated with chemo- and cytokines in human atherosclerotic lesions and leucocytes. Finally, atherosclerotic lesions in Ldlr−/−xFpr2−/− mice exhibited decreased collagen content, and Fpr2−/− SMCs exhibited a profile of increased collagenase and decreased collagen production pathways.ConclusionFPR2/ALX is proatherogenic due to effects on bone marrow-derived cells, but promoted a more stable plaque phenotype through effects on SMCs. Taken together, these results suggest a dual role of FPR2/ALX signalling in atherosclerosis by way of promoting disease progression and but increasing plaque stability.

Genetic Deletion of LXRα Prevents Arsenic-Enhanced Atherosclerosis, But Not Arsenic-Altered Plaque Composition

Arsenic exposure has been linked to an increased incidence of atherosclerosis. Previously, we have shown in vitro and in vivo that arsenic inhibits transcriptional activation of the liver X receptors (LXRs), key regulators of lipid homeostasis. Therefore, we evaluated the role of LXRα in arsenic-induced atherosclerosis using the apoE(-/-) mouse model. Indeed, deletion of LXRα protected apoE(-/-) mice against the proatherogenic effects of arsenic. We have previously shown that arsenic changes the plaque composition in apoE(-/-) mice. Arsenic decreased collagen content in the apoE(-/-) model, and we have observed the same diminution in LXRα(-/-)apoE(-/-) mice. However, the collagen-producing smooth muscle cells (SMCs) were decreased in apoE(-/-), but increased in LXRα(-/-)apoE(-/-). Although transcriptional activation of collagen remained the same in SMC from both genotypes, arsenic-exposed LXRα(-/-)apoE(-/-) plaques had increased matrix metalloproteinase activity compared with both control LXRα(-/-)apoE(-/-) and apoE(-/-), which could be responsible for both the decrease in plaque collagen and the SMC invasion. In addition, arsenic increased plaque lipid accumulation in both genotypes. However, macrophages, the cells known to retain lipid within the plaque, were unchanged in arsenic-exposed apoE(-/-) mice, but decreased in LXRα(-/-)apoE(-/-). We confirmed in vitro that these cells retained more lipid following arsenic exposure and are more sensitive to apoptosis than apoE(-/-). Mice lacking LXRα are resistant to arsenic-enhanced atherosclerosis, but arsenic-exposed LXRα(-/-)apoE(-/-) mice still present a different plaque composition pattern than the arsenic-exposed apoE(-/-) mice.

CD137-inducing factors from T cells and macrophages accelerate the destabilization of atherosclerotic plaques in hyperlipidemic mice.

CD137 (4-1BB), a member of the tumor necrosis factor receptor superfamily, has been reported to be expressed in atherosclerotic plaques, and to promote lesion formation. However, the role of CD137 in mediating atherosclerotic plaque stability and the possible underlying molecular and cellular mechanisms are poorly understood. Here, apolipoprotein E-deficient (ApoE(-/-)) and CD137-deficient ApoE(-/-) (ApoE(-/-)CD137(-/-)) mice fed a chow diet for 66 wk were used. CD137 induces plaque instability, which is characterized by increased plaque necrosis, decreased collagen content, decreased vascular smooth muscle cell (VSMC) content, and increased macrophage infiltration. CD137 also increases the infiltration of effector T (Teff) cells into plaque lesion sites, resulting in increased interferon-γ (IFN-γ) expression. Interestingly, Teff-cell-derived IFN-γ inhibits collagen synthesis in atherosclerotic plaques. Furthermore, CD137 activation increases the apoptosis of VSMCs, possibly by decreasing the antiapoptotic regulator, Bcl-2, and subsequently up-regulating cleaved caspase-3. In macrophages, activation of CD137 signaling boosted the oxidized low density lipoprotein-induced expression of matrix metalloproteinase 9 via the p38 mitogen-activated protein kinase and extracellular signal-regulated kinase1/2 signaling pathways. In summary, activation of CD137 signaling decreases the stability of advanced atherosclerotic plaques via its combined effects on Teff cells, VSMCs, and macrophages.

Effect of ultraviolet light treatment on microbial contamination, some textural and organoleptic parameters of cultured sea bass fillets (Dicentrarchus labrax)

Ultraviolet (UV) light (λ = 250 nm) was assessed to increase shelf life of sea bass ( Dicentrarchus labrax ). Fillets were exposed to different doses: i) 7.9 kJ/m 2 , designed as UV 8 , ii) 15.8 kJ/m 2 , UV 16 and iii) control, untreated (C). After the treatments, the fillets were stored in a cold-room (4 ± 1 °C) up to 11 days and sampled at 0, 2, 4, 7, 9 and 11 days. The UVL treatments significantly decreased microbial counts during the first 4 days post-mortem (dpm), independently of the doses, compared to C fillets. A delay in rigor process in both UV 8 and UV 16 fillets was observed, as well as consistent improvement of textural parameters (hardness, gumminess and chewiness) at 11 dpm. These findings correlate with the lower proteolysis in myofibrillar protein fraction observed in UV 16 fillets, confirmed by electrophoretic separation. On the other hand, UV treatments decreased collagen content and increased lipid oxidation in fillets. Continuous ultraviolet light is an interesting technology for food preservation due to its ability on the destruction of microorganisms mainly used in fluid and vegetal sanitation but with a proven effect in animal origin food. The advantages of treatment with continuous ultraviolet light for the food industry could be the absence of negative effect on food quality observed with other preservation treatments. The results of the present study showed that UV treatment causes a clear inhibition of microbial growth up to 4-day cold storage, independently of the doses. A consistently higher hardness throughout cold storage and a protective action on myofibrillar protein degradation were also evidenced. Future studies must be performed to minimize the negative effect observed in the present study. • Ultraviolet light was assessed to increase shelf life of sea bass. • UVL treatments decreased microbial counts during the first 4 days post-mortem. • UVL treated fillets showed higher hardness during cold storage. • Lower proteolysis in myofibrillar protein fraction was observed in UV 16 fillets. • UVL treatments decreased collagen content and increased lipid oxidation in fillets.

Targeted delivery of vascular endothelial growth factor improves stem cell therapy in a rat myocardial infarction model

Rebuilding of infarcted myocardium by mesenchymal stem cells (MSCs) has not been successful because of poor cell survival due in part to insufficient blood supply after myocardial infarction (MI). We hypothesize that targeted delivery of vascular endothelial growth factor (VEGF) to MI can help regenerate vasculature in support of MSC therapy in a rat model of MI. VEGF-encapsulated immunoliposomes targeting overexpressed P-selectin in MI tissue were infused by tail vein immediately after MI. One week later, MSCs were injected intramyocardially. The cardiac function loss was moderated slightly by targeted delivery of VEGF or MSC treatment. Targeted VEGF+MSC combination treatment showed highest attenuation in cardiac function loss. The combination treatment also increased blood vessel density (80%) and decreased collagen content in post-MI tissue (33%). Engraftment of MSCs in the combination treatment group was significantly increased and the engrafted cells contributed to the restoration of blood vessels. From the Clinical EditorVEGF immunoliposomes targeting myocardial infarction tissue resulted in significantly higher attenuation of cardiac function loss when used in combination with mesenchymal stem cells. MSCs were previously found to have poor ability to restore cardiac tissue, likely as a result of poor blood supply in the affected areas. This new method counterbalances that weakness by the known effects of VEGF, as demonstrated in a rat model.

Diabetes alters mechanical properties and collagen fiber re-alignment in multiple mouse tendons.

Tendons function to transfer load from muscle to bone through their complex composition and hierarchical structure, consisting mainly of type I collagen. Recent evidence suggests that type II diabetes may cause alterations in collagen structure, such as irregular fibril morphology and density, which could play a role in the mechanical function of tendons. Using the db/db mouse model of type II diabetes, the diabetic skin was found to have impaired biomechanical properties when compared to the non-diabetic group. The purpose of this study was to assess the effect of diabetes on biomechanics, collagen fiber re-alignment, and biochemistry in three functionally different tendons (Achilles, supraspinatus, patellar) using the db/db mouse model. Results showed that cross-sectional area and stiffness, but not modulus, were significantly reduced in all three tendons. However, the tendon response to load (transition strain, collagen fiber re-alignment) occurred earlier in the mechanical test, contrary to expectations. In addition, the patellar tendon had an altered response to diabetes when compared to the other two tendons, with no changes in fiber re-alignment and decreased collagen content at the midsubstance of the tendon. Overall, type II diabetes alters tendon mechanical properties and the dynamic response to load.

Abstract 370: Interferon Regulatory Factors 3 and 7 Regulate Vein Graft Remodeling and Vascular Inflammation

Introduction: Type I interferons (IFN) are implicated in the development of vascular proliferative diseases. Pathway analysis by gene set enrichment analysis of hypercholesterolemic ApoE3*Leiden murine vein grafts revealed that the type I IFN pathway belonged to the top 15 of significant regulated pathways. The transcriptional regulators of type I IFN and type I IFN responsive genes are the interferon regulatory factors (IRF). Activation of TLR3 results in activation of type I IFN in a IRF3 and IRF7 dependent manner. TLR3 has been shown to be protective in vascular remodeling. The aim of this study was to investigate the role of IRF3 and IRF7 on vein graft remodeling. Methods and Results: The importance of IRFs in vein graft remodeling is illustrated by the increase in vein graft thickening in Irf3 -/- and Irf7 -/- mice in comparison to control mice (n=9/group, Irf3 -/- ; 39%, p=0.185, Irf7 -/- ; 68% p=0.003). Also an increase in outward remodeling ( Irf3 -/- ; 26%, p=0.081, Irf7 -/- ; 42%, p=0.049) was observed comparable to that of Tlr3 -/- (n=8, 52%, p<0.001) Immunohistochemical analysis revealed that both Irf3 -/- and Irf7 -/- mice showed a significant higher influx of macrophages in the vessel wall than the control mice whereas the Irf7 -/- mice also showed a significant decrease in collagen content. RNA levels of typical type I IFN responsive genes such as Mx1, Ifit1-3 and Oas2 were down regulated in the knockout vein grafts in comparison to control vein grafts. Activation of both Irf3 -/- and Irf7 -/- bone marrow derived macrophages with LPS and poly:(IC) resulted in a significant increase in TNFα production. Conclusions: IRFs regulates vein graft remodeling since Irf3 -/- and especially Irf7 -/- vein grafts show increased vessel wall thickening and outward remodeling. This increased remodeling is the result of a pro-inflammatory response as reflected by the increase in macrophages in the vein graft wall.

11β-Hydroxysteroid Dehydrogenase 1 Specific Inhibitor Increased Dermal Collagen Content and Promotes Fibroblast Proliferation

Glucocorticoids (GCs) are one of the most effective anti-inflammatory drugs for treating acute and chronic inflammatory diseases. However, several studies have shown that GCs alter collagen metabolism in the skin and induce skin atrophy. Cortisol is the endogenous GC that is released in response to various stressors. Over the last decade, extraadrenal cortisol production in various tissues has been reported. Skin also synthesizes cortisol through a de novo pathway and through an activating enzyme. 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1) is the enzyme that catalyzes the conversion of hormonally inactive cortisone to active cortisol in cells. We previously found that 11β-HSD1 negatively regulates proliferation of keratinocytes. To determine the function of 11β-HSD1 in dermal fibroblasts and collagen metabolism, the effect of a selective 11β-HSD1 inhibitor was studied in mouse tissues and dermal fibroblasts. The expression of 11β-HSD1 increased with age in mouse skin. Subcutaneous injection of a selective 11β-HSD1 inhibitor increased dermal thickness and collagen content in the mouse skin. In vitro, proliferation of dermal fibroblasts derived from 11β-HSD1 null mice (Hsd11b1−/− mice) was significantly increased compared with fibroblasts from wild-type mice. However, in vivo, dermal thickness of Hsd11b1−/− mice was not altered in 3-month-old and 1-year-old mouse skin compared with wild-type mouse skin. These in vivo findings suggest the presence of compensatory mechanisms in Hsd11b1−/− mice. Our findings suggest that 11β-HSD1 inhibition may reverse the decreased collagen content observed in intrinsically and extrinsically aged skin and in skin atrophy that is induced by GC treatment.

Manipulation of valve composition to elucidate the role of collagen in aortic valve calcification

BackgroundExtracellular matrix (ECM) disarray is found in calcific aortic valvular disease (CAVD), yet much remains to be learned about the role of individual ECM components in valvular interstitial cell (VIC) function and dysfunction. Previous clinical analyses have shown that calcification is associated with decreased collagen content, while previous in vitro work has suggested that the presence of collagen attenuates the responsiveness of VICs to pro-calcific stimuli. The current study uses whole leaflet cultures to examine the contributions of endogenous collagen in regulating the phenotype and calcification of VICs.MethodsA “top-down” approach was used to characterize changes in VIC phenotype in response to collagen alterations in the native 3D environment. Collagen-deficient leaflets were created via enzymatic treatment and cultured statically for six days in vitro. After culture, leaflets were harvested for analysis of DNA, proliferation, apoptosis, ECM composition, calcification, and gene/protein expression.ResultsIn general, disruption of collagen was associated with increased expression of disease markers by VICs in whole organ leaflet culture. Compared to intact control leaflets, collagen-deficient leaflets demonstrated increased VIC proliferation and apoptosis, increased expression of disease-related markers such as alpha-smooth muscle actin, alkaline phosphatase, and osteocalcin, and an increase in calcification as evidenced by positive von Kossa staining.ConclusionsThese results indicate that disruption of the endogenous collagen structure in aortic valves is sufficient to stimulate pathological consequences in valve leaflet cultures, thereby highlighting the importance of collagen and the valve extracellular matrix in general in maintaining homeostasis of the valve phenotype.

Effects of castration and time-on-feed on Mertolenga breed beef quality

Physicochemical characteristics were determined in the longissimus lumborum muscle, after 8 days of ageing of steers (n=12) and bulls (n=12) from Mertolenga breed slaughtered directly from pasture (day 0) or after a finishing period of 50, 100 and 150 days in a feed-lot facility. Bulls and steers presented similar live weight (averaging 388 kg), carcass weight (CW; averaging 213 kg), dressing percentage (averaging 60%), carcass fatness (11.9% CW) and carcass fat thickness (averaging 3.03 mm). Live weight, CW, carcass fatness and fat thickness increased along time-on-feed. Gender only had a negligible effect on meat characteristics, with b* and h* being the only parameters of colour affected by gender, also presenting a significant interaction gender×time-on-feed. Nevertheless, both the genders presented a high-quality grade concerning tenderness (Warner–Bratzler shear force (WBSF)). L* increased until 50 days on feed and decreased afterwards, whereas a* and C* values increased along time-on-feed. Pigment content was also affected by time-on-feed and showed a gender×time-on-feed interaction. Beef colour became darker and redder along time-on-feed, but still in a colour range highly acceptable by Portuguese consumers. Despite the increase in intramuscular fat and myofibrillar fragmentation index, as well as the decrease in collagen content of steers and bulls along time-on-feed, it did not affect the tenderness/hardness, indicating a small effect of time-on-feed in meat characteristics. Despite only small differences in carcass characteristics and meat-quality parameters that have been noticed along time-on-feed, those differences were only significant after 100 days on feed. Principal component analysis (PCA) was performed. The first PC axis (39.6% of the total variance) included colour variables a*, b* and C*, and carcass fatness, fat thickness, CW and live weight, whereas the second one (12.7% of the total variance) included h*, cooking losses and dressing-out. The principal component (PC) analysis confirmed the lack of differences between bulls and steers and indicates a differentiation of the first two periods of feeding (0 and 50 days on feed) from the two latter (100 and 150 days on feed) periods of feeding.

Loss of progesterone receptor-mediated actions induce preterm cellular and structural remodeling of the cervix and premature birth.

A decline in serum progesterone or antagonism of progesterone receptor function results in preterm labor and birth. Whether characteristics of premature remodeling of the cervix after antiprogestins or ovariectomy are similar to that at term was the focus of the present study. Groups of pregnant rats were treated with vehicle, a progesterone receptor antagonist (onapristone or mifepristone), or ovariectomized on day 17 postbreeding. As expected, controls given vehicle delivered at term while rats delivered preterm after progesterone receptor antagonist treatment or ovariectomy. Similar to the cervix before term, the preterm cervix of progesterone receptor antagonist-treated rats was characterized by reduced cell nuclei density, decreased collagen content and structure, as well as a greater presence of macrophages per unit area. Thus, loss of nuclear progesterone receptor-mediated actions promoted structural remodeling of the cervix, increased census of resident macrophages, and preterm birth much like that found in the cervix at term. In contrast to the progesterone receptor antagonist-induced advance in characteristics associated with remodeling, ovariectomy-induced loss of systemic progesterone did not affect hypertrophy, extracellular collagen, or macrophage numbers in the cervix. Thus, the structure and macrophage census in the cervix appear sufficient for premature ripening and birth to occur well before term. With progesterone receptors predominantly localized on cells other than macrophages, the findings suggest that interactions between cells may facilitate the loss of progesterone receptor-mediated actions as part of a final common mechanism that remodels the cervix in certain etiologies of preterm and with parturition at term.

Lack of Phospholipase A 2 Receptor Increases Susceptibility to Cardiac Rupture After Myocardial Infarction

Recent evidence indicates that the biological effects of secretory phospholipase A2 (sPLA2) cannot be fully explained by its catalytic activity. A cell surface receptor for sPLA2 (PLA2 receptor 1 [PLA2R]) and its high-affinity ligands (including sPLA2-IB, sPLA2-IIE, and sPLA2-X) are expressed in the infarcted myocardium. This study asked whether PLA2R might play a pathogenic role in myocardial infarction (MI) using mice lacking PLA2R (PLA2R(-/-)). MI was induced by permanent ligation of the left coronary artery. PLA2R(-/-) mice exhibited higher rates of cardiac rupture after MI compared with PLA2R wild-type (PLA2R(+/+)) mice (46% versus 21%, respectively; P=0.015). PLA2R(-/-) mice had a 31% decrease in collagen content and a 45% decrease in the number of α-smooth muscle actin-positive fibroblasts in the infarcted region compared with PLA2R(+/+) mice. PLA2R was primarily found in myofibroblasts in the infarcted region. PLA2R(-/-) myofibroblasts were impaired in collagen-dependent migration, proliferation, and activation of focal adhesion kinase in response to sPLA2-IB. Binding of sPLA2-IB to PLA2R promoted migration and proliferation of myofibroblasts through functional interaction with integrin β1, independent of the catalytic activity of sPLA2-IB. In rescue experiments, the injection of PLA2R(+/+) myofibroblasts into the infarcted myocardium prevented post-MI cardiac rupture and reversed the decrease in collagen content in the infarcted region in PLA2R(-/-) mice. PLA2R deficiency increased the susceptibility to post-MI cardiac rupture through impaired healing of the infarcted region. This might be partly explained by a reduction in integrin β1-mediated migratory and proliferative responses of PLA2R(-/-) myofibroblasts.

Antifibrotic Properties of Transarterial Oncolytic VSV Therapy for Hepatocellular Carcinoma in Rats With Thioacetamide-Induced Liver Fibrosis

Recombinant vesicular stomatitis virus (VSV) shows promise for the treatment of hepatocellular carcinoma (HCC), but its safety and efficacy when administered in a setting of hepatic fibrosis, which occurs in the majority of clinical cases, is unknown. We hypothesized that VSV could provide a novel benefit to the underlying fibrosis, due to its ability to replicate and cause cell death specifically in activated hepatic stellate cells. In addition to the ability of VSV to produce a significant oncolytic response in HCC-bearing rats in the background of thioacetamide-induced hepatic fibrosis without signs of hepatotoxicity, we observed a significant downgrading of fibrosis stage, a decrease in collagen content in the liver, and modulation of gene expression in favor of fibrotic regression. Together, this work suggests that VSV is not only safe and effective for the treatment of HCC with underlying fibrosis, but it could potentially be developed for clinical application as a novel antifibrotic agent.

Dietary lecithin improves dressing percentage and decreases chewiness in the longissimus muscle in finisher gilts

The influence of dietary lecithin at doses of 0, 4, 20 or 80g/kg fed to finisher gilts for six weeks prior to slaughter on growth performance, carcass quality and pork quality was investigated. M. longissimus lumborum (loin) was removed from 36 pig carcasses at 24h post-mortem for Warner–Bratzler shear force, compression, collagen content and colour analyses. Dietary lecithin increased dressing percentage (P=0.009). Pork chewiness and collagen content were decreased by dietary lecithin (P<0.05, respectively), suggesting that improved chewiness may be due to decreased collagen content. However, dietary lecithin had no effect on shear force, cohesiveness or hardness (P>0.05, respectively). Dietary lecithin reduced loin muscle L* values and increased a* values (P<0.05, respectively) but no changes on b* values (P=0.56). The data showed that dietary lecithin improved dressing percentage and resulted in less chewy and less pale pork.

Reproducible Porcine Model of Thoracic Aortic Aneurysm

Thoracic aortic aneurysms (TAAs) develop secondary to abnormal aortic extracellular matrix remodeling, resulting in a weakened and dilated aortic wall that progressed to rupture if left unattended. Currently, no diagnostic/prognostic tests are available for the detection of TAA disease. This is largely driven by the lack of a large animal model, which would permit longitudinal/mechanistic studies. Accordingly, the objective of the present study was to establish a reproducible porcine model of aortic dilatation, which recapitulates the structural and biochemical changes observed during human TAA development. Descending TAAs were induced in Yorkshire pigs (20-25 kg; n=7) through intra-adventitial injections of collagenase (5 mL, 0.35 mg/mL) and periadventitial application of crystalline CaCl2 (0.5 g). Three weeks after TAA induction, aortas were harvested and tissue was collected for biochemical and histological measurements. A subset of animals underwent MRI preoperatively and at terminal surgery. Results were compared with sham-operated controls (n=6). Three weeks after TAA induction, aortic luminal area increased by 38 ± 13% (P=0.018 versus control). Aortic structural changes included elastic lamellar degradation and decreased collagen content. The protein abundance of matrix metalloproteinases 3, 8, 9, and 12 increased in TAA tissue homogenates, whereas tissue inhibitors of metalloproteinases 1 and 4 decreased. These data demonstrate aortic dilatation, aortic medial degeneration, and alterations in matrix metalloproteinase/tissue inhibitors of metalloproteinase abundance, consistent with TAA formation. This study establishes for the first time a large animal model of TAA that recapitulates the hallmarks of human disease and provides a reproducible test bed for examining diagnostic, prognostic, and therapeutic strategies.

Abstract 186: ACE2 Activation Protects Corpus Cavernosum Impairment Induced by Hypercholesterolemia

Erectile dysfunction (ED) is multicausal disorder strictly associated with cardiovascular diseases. ACE2 is a key enzyme of the renin angiotensin system which plays an essential role in the cardiovascular system; however, the function of this enzyme in erectile tissues has never been investigated. Here we examined the expression of ACE2 in erectile tissues and its protective actions against hypercholesterolemic-induced corpus cavernosum injures. ACE2 expression was evaluated by immunostaining and western blot. Apolipoprotein E knockout (ApoE-/-) mice fed with Western-type diet for 11 weeks were treated with DIZE, an ACE2 activator compound, or vehicle during the last 3 weeks. Collagen and reactive oxygen species (ROS) content within the corpus cavernosum (CC) were evaluated by Sirius red and dihydroethidium staining, respectively. Protein expression of nNOS, eNOS, NADPH subunits (p67-phox and p22-phox) was assessed by western blot. Functional studies in CC strips were performed in an isometric organ bath. ACE2 was expressed in smooth muscle and endothelial cells of mouse CC. Interestingly, this enzyme was downregulated in penis from hypercholesterolemic mice. The treatment with DIZE reduced ROS production and NADPH oxidase expression in penis from ApoE-/- mice. Furthermore, DIZE treatment elevated vascular NO production (untreated-ApoE-/-: 11.3±1.1 vs. treated-ApoE-/-: 18.7±4.5, mol/L of serum NO3-) and augmented the expression of nNOS and eNOS in the penis. The NO-dependent relaxation induced by acetylcholine (ACh) was significantly increased in CC strip from DIZE-treated ApoE-/- mice (wild type 53.7%; untreated-ApoE-/- 39.3%; treated-ApoE-/- 52.1%, percentage of relaxation at 1μM of ACh). Additionally, DIZE treatment decreased collagen content within the CC. Of note, these beneficial actions of DIZE on the CC were not associated with changes in atherosclerotic plaque deposition or serum lipid profile. In summary, ACE2 activation attenuated oxidative stress, augmented NO bioavailability and decreased collagen deposition in CC of hypercholesterolemic mice. These results demonstrate that ACE2 is expressed and functional in erectile tissue, suggesting that this enzyme might have significant therapeutic benefits against ED.