Verhoeff-Van Gieson Staining Research Articles

Ex vivo expansion of regulatory T cells from abdominal aortic aneurysm patients inhibits aneurysm in humanized murine model

ObjectiveAbdominal aortic aneurysm (AAA) is a chronic inflammatory disease. Studies of human aneurysm tissue demonstrate dense inflammatory cell infiltrates with CD4+ T cells predominating. Regulatory T cells (Tregs) play an important role in inhibiting pro-inflammatory T cell proliferation, therefore, limiting collateral tissue destruction. The aim of this study was to investigate whether ex vivo augmentation of human Tregs attenuates aneurysm formation in humanized murine model of AAA. MethodsCirculating Treg population in AAA patients and age- and gender-matched controls were determined by real-time polymerase chain reaction and flow cytometry. To create humanized murine model of AAA, irradiated Rag1-deficient (Rag1-/-) mice, without mature T lymphocytes, at 7 weeks of age were given 5 × 106 of human CD4+ T cells intraperitoneally. Then the mice underwent CaCl2 aneurysm induction. Aortic diameters were measured before and at 6 weeks after aneurysm induction. Aortic tissue was collected for histology and protein extraction. Verhoeff-Van Gieson stain was used for staining elastic fiber. CD4+ T cells in the aortic tissue were detected by immunohistochemical staining. ResultsIn human peripheral blood mononuclear cells, the proportion of Tregs are decreased in AAA patients compared with matched control patients with significant vascular disease. We first validated the role of Tregs in the CaCl2 model of AAA. To determine the role of human T cells in AAA formation, Rag1-/- mice, resistant to CaCl2-aneurysm induction, were transplanted with human CD4+ T cells. Human CD4+ T cells were able to drive aneurysm formation in Rag1-/- mice. We show that ex vivo augmentation of human Tregs by interleukin-2 resulted in decreased aneurysm progression. ConclusionsThese data suggest that the ex vivo expansion of human Tregs may be a potential therapeutic strategy for inhibiting progression of AAA.

P3113Functional deficiency in activating fcgamma receptor protects against aortic abdominal aneurysm in mice

Abstract Background and aim Abdominal aortic aneurysm (AAA) is a degenerative disorder characterized by a localized and permanent dilation of the aorta. Pathological features of AAA include proteolysis, vascular smooth muscle cells (VSMC) apoptosis, oxidative stress and inflammation. Previous studies have demonstrated the role of innate and adaptive immunity in the initiation and progression of AAA. However, the specific mechanisms of humoral, antibody-mediated, immune response elicited by self and non-self-antigens are not completely known. IgG Fc receptors (FcgammaR) play an important role in the initiation and regulation of many immunological and inflammatory processes, thus providing a link between humoral and cellular immune responses. In this work, we focus on IgG immune response against antigens exposed in the damaged vessel, analysing the specific role played by FcgammaR in the pathogenesis of AAA. Methods and results AAA was induced by aortic elastase perfusion in wild-type (WT) C57BL/6 mice (n=24 males, 12 weeks old). Compared with healthy aortas, AAA tissue showed IgG and IgM deposition and increased expression levels of activating (IA, IIIA and IVA) and inhibitory (IIB) FcgammaR (5-, 3-, 10, and 2-fold increases; p<0.005) at day 14 post-elastase perfusion. To explore the functional contribution of activating FcgammaR to AAA formation, parallel experiments were performed in mice deficient in gamma-chain (gamma-KO), the common signalling subunit of FcgammaRI, III and IV. Compared with WT mice, gamma-KO mice (n=21) exhibited lower IgG and IgM deposits (21±4% and 28±12% vs WT, respectively; p<0.02) and decreased AAA lesions with reduced aortic expansion (% vs WT: aortic wall thickness, 67±3%; aortic diameter, 55±3%; p<0.0001). AAA lesions from gamma-KO mice showed less disruption of elastin layers (Verhoeff-van Gieson staining) and reduced loss of medial VSMC (α-actin immunofluorescence). Inflammatory markers such as leukocyte content (CD68+ macrophages, Ly6G+ neutrophils, CD45R+ B cells, and CD3+ T lymphocytes) and the gene expression of chemokines (MCP-1, RANTES), adhesion molecules (ICAM-1), cytokines (TNFα, IFNg, IL-10) and metalloproteinases (MMP9) were all significantly lower in AAA lesions from gamma-KO mice than those from WT mice. In vitro, cross-linking of FcgammaR with fibrinogen-containing immune complexes triggers the gene expression of proinflammatory cytokines and chemokines in both VSMC and bone marrow-derived macrophages. Conclusion Activating FcgammaR participate in AAA formation by promoting inflammatory cell recruitment, and cytokine and matrix-degrading protease expression. Therefore, modulation of FcgammaR-dependent responses could be a promising therapeutic option for the treatment of AAA. Acknowledgement/Funding MINECO/FEDER (SAF2015-63696-R), ISCIII (FIS/FEDER PI17/01495), Spanish Society of Arteriosclerosis, La Caixa (HR17-00247), Conchita Rabago Foundation

The Lumbar Ligamentum Flavum Does Not Have Two Layers and Is Confluent with the Interspinous Ligament: Anatomical Study with Application to Surgical and Interventional Pain Procedures

Numerous authors over the years have reported that the lumbar ligamentum flavum has two layers. Our routine cadaveric dissections raised the question whether this understanding is correct, as we always have observed only one layer. Thus, the goal of this cadaveric study was to reevaluate the layers of the ligamentum flavum. Twenty lumbar levels from five fresh-frozen cadaveric specimens were used in this study. After dissection of the lumbar spine, the ligamentum flavum and interspinous ligament were exposed. Each lumbar level was transected through the zygapophyseal joint, and hematoxylin and eosin staining, Masson's trichrome staining and Verhoeff-van Gieson staining were performed. Continuation of the interspinous ligament and ligamentum flavum were observed invariably. There was no evidence of the existence of a two-layered ligamentum flavum. The lumbar ligamentum flavum does not consist of two layers, but is confluent instead with the interspinous ligament that attaches to the zygapophyseal joints. To convey this anatomy better, we suggest describing the lumbar ligamentum flavum as a structure that consists of interlaminar and interspinous parts. Precise knowledge of the ligamentum flavum's anatomy can be of clinical value, particularly when epidural anesthesia or lumbar puncture are performed. Clin. Anat. 32:34-40, 2019. © 2019 Wiley Periodicals, Inc.

Morphological Analysis of Elastic Fibers in Various Grades of Oral Squamous Cell Carcinoma and Epithelial Dysplasia Using Verhoeff-Van Gieson Stain.

BackgroundOral squamous cell carcinoma (OSCC) is the sixth most common malignancy in India. The aggressiveness of OSCC is analyzed not only based on the dysplastic features and tumor infiltration pattern, but also by means of the stromal changes that pave the way for an invasion into the connective tissue. The role of elastic fibers in the progression of OSCC is still unknown because of sparse literature and the masking effect of overlying inflammatory cells and the lower number of elastic fibers in the lamina propria. The present study provides further insight into the qualitative assessment of elastic fibers in various grades of dysplasia and OSCC.ObjectivesTo analyze the morphological changes exhibited by the elastic fibers in epithelial dysplasia and OSCC.Materials and methodsTwo sections were cut from each of 60 samples of varying grades of OSCC and 60 samples of varying grades of epithelial dysplasia followed by staining with hematoxylin and eosin and Verhoeff–Van Gieson stain.ResultsStatistically significant results were obtained for qualitative analysis of elastic fibers. A change in density and orientation to overlying epithelium and tumor islands was seen on progressing from well-differentiated to poorly differentiated OSCC and in progressing grades of dysplasia.ConclusionThe uniqueness of this study lies in the exploration of elastic fibers in dysplasia and well-differentiated OSCC, a less explored field. The study of the connective tissue stromal changes can be used as an adjunct to histological grading.

Atorvastatin Protects Against Cerebral Aneurysmal Degenerative Pathology by Promoting Endothelial Progenitor Cells (EPC) Mobilization and Attenuating Vascular Deterioration in a Rat Model.

BackgroundEndothelial injury is the early pathological change of cerebral aneurysm (CA) formation. In addition to its lipid-lowering activity, atorvastatin (ATR) also reportedly promotes vascular repair via mobilizing endothelial progenitor cells (EPC). Here, we investigated the influence of ATR on vascular worsening after CA induction in rats.Material/MethodsAdult male Sprague-Dawley rats were randomly assigned to 3 groups: a control (CTR) group, a CA group, and a CA+ATR treatment group. Circulating EPC level and hematological and lipid profiles were measured 3 months after CA induction. Verhoeff-Van Gieson staining and transmission electron microscopy were performed to assess pathological changes in the artery wall. RT-PCR was also performed to evaluate the expression of inflammation-related genes in the aneurysmal wall.ResultsATR significantly restored the impaired level of circulating EPC without changing hematological and lipid profiles 3 months after CA induction. ATR markedly inhibited endothelial injury, media thinning, and CA enlargement, accompanied by reduced vascular inflammation.ConclusionsOur preliminary results demonstrate that the mobilization of EPC and improvement of endothelial function by ATR contribute to the prevention of cerebral aneurysm. Further studies are warranted to investigate the detailed mechanism.

Third Harmonic Generation Imaging using Common Histological Dyes

Histology has long relied on optical microscopy of tissue stained with colorful absorptive dyes, which enables easy visual differentiation between cell nuclei and various extracellular structures. Histological stains are presently the gold standard in diagnoses by medical pathologists. While these stains provide an effective way to visualize tissue components, they still require thin sectioning which results in only two dimensional data that cannot be easily stitched in three dimensions. As tissue inherently exists as a three dimensional structure, it is important to develop histological methods to investigate it in such form while maintaining the high spatial resolution of light microscopy. Third harmonic generation (THG) imaging is a nonlinear microscopy technique that is gaining attention in the biomedical community. In this technique, three incident photons are combined into one by light-matter interactions with the sample, resulting in high contrast and inherent three dimensional optical sectioning. While in practice THG only typically occurs at optical heterogeneities and interfaces within the biological sample, some materials present enhanced THG emission due to resonance effects. These materials have also been called harmoniphores. Hematoxylin (H), a dye that is often used in combination with eosin (E) as part of the H&E stain, has previously been reported as an effective harmoniphore for distinguishing cell nuclei by 3D THG imaging. In this work, we apply trans THG imaging to skin and liver sections to show that in addition to H&E, other commonly used histological dyes in elastic and collagen staining can serve as effective THG harmoniphores. We find that elastic fibers can be easily visualized by THG microscopy using Verhoeff Van Gieson (VVG) staining while collagen is directly highlighted by THG imaging of picrosirius red staining. By combining THG imaging with common histological dyes, 3D histological imaging could be achieved.

Abstract WP341: Experimental Internal Carotid Artery Stenosis Models Pathogenic Features of Moyamoya Syndrome

Introduction: Moyamoya is an arteriopathy defined by the progressive stenosis of the intracranial internal carotid arteries accompanied by the formation of abnormal vascular networks. To address the paucity of research on the adult-onset Moyamoya Syndrome, which completely lacks any animal model for evaluation, our lab created a novel surgical technique, termed internal carotid artery stenosis (ICAS), that attempts to model the cerebrovascular changes seen in the human Moyamoya Syndrome. We hypothesized that the ICAS model would mimic the key proposed pathogenic features of Moyamoya Syndrome; namely, intimal hyperplasia of the major vessels of the cerebrovasculature leading to vessel stenosis and the formation of small, compensatory collateral arteries. Methods: ICAS (n=9) and sham (n=6) surgeries were performed in a randomized fashion on male C57Bl/6 mice (age=16 weeks). Brain tissue sections and RNA were collected 28-days post-surgery. Intimal hyperplasia was assessed with a modified Verhoeff-van Gieson stain. Expression of genes associated with human Moyamoya Syndrome (VEGF, SDF-1α, MMP-9, bFGF, VCAM-1, and MCP-1) was quantified by qPCR. MatLab and PRISM software were used for data analysis. Results: ICAS induces intimal hyperplasia of the cerebrovasculature ( p =0.033). ICAS increases genetic expression of VEGF in the whole brain (fold change=0.60, p =0.050) and SDF-1α in the striatum (fold change=0.59, p =0.016), and decreases genetic expression of MMP-9 in the ipsilateral cortex (fold change=0.39, p =0.044). Furthermore, a trend indicates that ICAS may increase genetic expression of VCAM-1 in the ipsilateral striatum (fold change=0.33, p =0.109). Conclusions: The present finding that ICAS induces intimal hyperplasia and alters genetic expression supports ICAS as a relevant model of the proposed pathology of Moyamoya Syndrome. The upregulation of VEGF, SDF-1α, and VCAM-1 is indicative of angiogenesis, hypoxia-induced vasculogenesis, and ischemia-induced inflammation, while the downregulation of MMP-9 may indicate decreased extracellular remodeling in the cortex. While investigation of the long-term effects of coil-placement is needed, ICAS may provide a crucial model for development of therapies for Moyamoya Syndrome.

Ginkgo biloba Extract Protects Mesenteric Arterioles of Old Rats via Improving Vessel Elasticity through Akt/FoxO3a Signaling Pathway

Previous studies have shown that Ginkgo biloba extract (GBE) dietary diminished salt-related elevation of blood pressure and ameliorated ischemic diseases. However, whether GBE could improve vascular elasticity to protect mesenteric arterioles of old rats is still elusive. In this study, we aimed to investigate the effects of GBE on vascular elasticity of old rats and its possible underlying mechanism. Morphological changes of mesenteric arterioles were observed using hematoxylin and eosin and Verhoeff-Van Gieson staining, and diameters of mesenteric arterioles under various pressure were detected after GBE administration. In addition, phosphorylation level of Akt and FoxO3a proteins from mesenteric arterioles were detected. The results implicated that GBE treatment narrowed endothelial cell gap and increased the curvature of inner elastic membrane with reduced middle layer collagen fiber. Meanwhile, compared with young rats, old rats appeared to have lower vascular elasticity while GBE treatment at 50, 100, and 200mg/kg dosage through intragastric administration per day for 3weeks could effectively improve the vascular elasticity under different pressures in a dose-dependent manner. Furthermore, phosphorylation level of Akt and FoxO3a was also reduced in GBE-treated rats. This is the first report to indicate that GBE might exert protective effect on mesenteric arterioles of old rats via improving vascular elasticity and Akt/FoxO3a signaling pathway might be involved in this action.

A case of papular elastorrhexis.

ENWEndNote BIBJabRef, Mendeley RISPapers, Reference Manager, RefWorks, Zotero AMA Jiang M, Bu W, Chen X, Gu H. A case of papular elastorrhexis. Advances in Dermatology and Allergology/Postępy Dermatologii i Alergologii. 2019;36(1):117-118. doi:10.5114/ada.2019.82832. APA Jiang, M., Bu, W., Chen, X., & Gu, H. (2019). A case of papular elastorrhexis. Advances in Dermatology and Allergology/Postępy Dermatologii i Alergologii, 36(1), 117-118. https://doi.org/10.5114/ada.2019.82832 Chicago Jiang, Meng, Wenbo Bu, Xu Chen, and Heng Gu. 2019. "A case of papular elastorrhexis". Advances in Dermatology and Allergology/Postępy Dermatologii i Alergologii 36 (1): 117-118. doi:10.5114/ada.2019.82832. Harvard Jiang, M., Bu, W., Chen, X., and Gu, H. (2019). A case of papular elastorrhexis. Advances in Dermatology and Allergology/Postępy Dermatologii i Alergologii, 36(1), pp.117-118. https://doi.org/10.5114/ada.2019.82832 MLA Jiang, Meng et al. "A case of papular elastorrhexis." Advances in Dermatology and Allergology/Postępy Dermatologii i Alergologii, vol. 36, no. 1, 2019, pp. 117-118. doi:10.5114/ada.2019.82832. Vancouver Jiang M, Bu W, Chen X, Gu H. A case of papular elastorrhexis. Advances in Dermatology and Allergology/Postępy Dermatologii i Alergologii. 2019;36(1):117-118. doi:10.5114/ada.2019.82832.

Pebbled skin over axilla and umbilicus

Pseudoxanthoma elasticum or Gronblad–Strandberg syndrome is characterized by progressive calcification and degeneration of the elastic fibers throughout the body. They are noted in the second or third decade and comprise flat-topped, discrete-to-confluent yellowish papules in a linear or reticular pattern over flexures, and periumbilical skin giving a “cobblestone,” “plucked chicken skin,” or “Moroccan leather” appearance. We report a case of 13-year-old-child with yellowish pebbled skin over axilla and umbilicus. Angioid streaks were present on ophthalmological examination. On histopathology, fragmented and curled fibers were present in the dermis giving ravelled wool appearance. Von Kossa and Verhoeff-van gieson stain fibers stains showed calcified and fragmented elastic fibers in the mid-dermis.

Comparative mechanical, morphological, and microstructural characterization of porcine mitral and tricuspid leaflets and chordae tendineae

BackgroundHealthy function of tricuspid valve (TV) structures is essential to avoid tricuspid regurgitation (TR) and may significantly improve disease prognosis. Mitral valve (MV) structures have been extensively studied, but little is known about the TV and right-sided heart diseases. Therefore, clinical decisions and finite element (FE) simulations often rely heavily on MV data for TV applications, despite fundamentally different mechanical and physiological environments. Method/ResultsTo bridge this gap, we performed a rigorous mechanical, morphological, and microstructural characterization of the MV and TV leaflets and chordae in a porcine model. Planar biaxial testing, uniaxial testing, second harmonic generation imaging and Verhoeff Van Gieson staining were performed. Morphological parameters, tissue moduli, extensibility, and anisotropy were quantified and compared. No major differences in leaflet mechanics or structure were found between TV and MV; chordal mechanics, morphology, and structure were found to compensate for anatomical and physiological loading differences between the valves. No differences in chordal mechanics were observed by insertion point within a leaflet; the septal tricuspid leaflet (STL) and posterior mitral leaflet (PML) did not have distinguishable strut chords, and the STL had the shortest chords. Within a valve, chords from septally-located leaflets were more extensible. MV chords were stiffer. ConclusionsThis study presents the first rigorous comparative mechanical and structural dataset of MV and TV structures. Valve type and anatomical location may be stronger predictors of chordal mechanics. Chords from septally-located leaflets differ from each other and from their intravalvular counterparts; they merit special consideration in surgical and computational applications. Statement of SignificanceA better understanding of the tricuspid valve (TV) and its associated structures is important for making advancements towards the repair of tricuspid regurgitation. Mitral valve structures have been extensively studied, but little is known about the TV and right-sided heart diseases. Clinical decisions and computational simulations often rely heavily on MV data for TV applications, despite fundamentally different environments. We therefore performed a rigorous mechanical, morphological, and microstructural characterization of atrioventricular leaflets and chordae tendineae in a porcine model.Finding that valve type and anatomical location may be strong predictors of chordal mechanics, chords from septally-located leaflets differ from each other and from their intravalvular counterparts; they merit special consideration in surgical and computational applications.

Alpha-pinene attenuates UVA-induced photoaging through inhibition of matrix metalloproteinases expression in mouse skin

AimThe present study was designed to examine the role of alpha-pinene (AP) against skin photoaging in UVA-irradiated mice. Materials and methodsSwiss albino mice were subjected to UVA-irradiation at the rate of 10 J/cm2 per day for ten days, totally mouse received 100 J/cm2. One hour prior to each UVA-exposure, the mouse skin was topically treated with AP (100 mg kg/b·wt). Biochemical methods were employed to study the status of antioxidant enzymes and lipid peroxidation. Histopathological observations were performed using hematoxylin and eosin (H & E) and Verhoeff van Gieson (VVG) staining in the mouse skin. The inflammatory and apoptotic protein expression was studied by immunohistochemical and Western blot methods. The mRNA expression of matrix metalloproteinases was determined by qRT-PCR and Western blot analysis. Key findingsWe found that AP pretreatment substantially ameliorated UVA-induced depletion of antioxidant enzymes and prevented UVA-induced lipid peroxidation in the mouse skin. Further, AP effectively inhibited UVA-induced activation of pro-angiogenic (iNOS and VEGF), inflammatory proteins (TNF-α, IL-6, and COX-2) expression and prevented the activation of NF-κB p65 in the mouse skin. Additionally, AP inhibited UVA-mediated apoptotic mediators (Bax, Bcl-2, caspase-3 and caspase 9) expression in the mouse skin. Moreover, AP inhibited mRNA expression of matrix metalloproteinases (MMP-13 and MMP-9) and tissue type IV collagenase (MMP-2) expression in the mouse skin. Histological studies showed that AP remarkably prevented the dermal tissue damage in UVA-irradiated mice. ConclusionThus, AP treatment effectively prevented UVA-induced photoaging probably through its antioxidant property.

Three-dimensional structure of the orbicularis retaining ligament: an anatomical study using micro-computed tomography

The orbicularis retaining ligament (ORL) is an important structure for maintaining the eyelid and cheek skin and contouring the characteristic facial appearance. However, the ORL is a delicate structure that is easily damaged in manual dissection. This study aimed to comprehensively investigate the ORL using a micro-computed tomography (mCT) with phosphotungstic acid (PTA) preparation for the acquisition of its three-dimensional information non-destructively. Twenty-two specimens were obtained from non-embalmed human cadaver (mean age 73.7 years). Multidirectional images of the mCT showed that the ORL consisted of continuous tiny plates with a multilayered plexiform shape. The modified Verhoeff Van Gieson staining and immunofluorescence revealed a ligamentous tissue consisting of multiple fibroelastic bundles. The preorbicularis fibres of the ORL had more layers and a more intricate arrangement than its retro-orbicularis fibres. The number, complexity and ambiguity of the ORL fibres increased in the lateral area and their density and extent increased near the dermis. Its dermal anchorage was shown as a confluence of its fibroelastic tissue into the dermis. The ORL comprises a multilayered meshwork of very thin continuous fibroelastic plates and its related cutaneous deformities might be a complicated outcome of subcutaneous tissue shrinkage, lipid accumulation and ORL retention.

Fibre Interplay in the Pregnant Uterus of the Frugivorous Bat Eidolon Helvum

Introduction There is currently a lack of adequate information on the mating, reproduction and the reproductive anatomy of the frugivorous bat Eidolon helvum. We, therefore, investigated their uterus during the non-pregnant and pregnant states for adequate information and comparison between the two states to ascertain what histological adaptations occur during pregnancy in relation to collagen and elastic fibers. Materials and Methods A total of 47 female bats (E. helvum) were used, out of which 24 were pregnant and 23 were not. They were harvested on the Obafemi Awolowo University Campus. The animals were carefully screened, assumed to be presumably healthy and then sacrificed by cervical dislocation. The left and right uterine limbs with the uterine body and part of the placenta were fixed in 10% formal saline, processed for paraffin embedding and sectioned at 5 μm with a rotary microtome. The sections were stained using Verhoeff-van Gieson stain to demonstrate collagen and elastic fibers. Results The results showed that both uterine limbs were histologically active during pregnancy. The prevalent fiber was the elastic type during the non-pregnant state, and collagen type during pregnancy. Conclusion We conclude that elastic fibers tend to obey Hooke's law during pregnancy in the uterus of the frugivorous bat E. helvum, which signifies a form of histomorphological response or adaptation to pregnancy in the uterus of this bat species.

Application of Combined Special Staining in the Observation of Vascular Pathology

Objective To evaluate the value of combined special staining technique in observing pathological changes in blood vessels. Methods Totally 999 vascular specimen were harvested from patients with complete medical records,clear diagnosis,and age≥18 years in the Pathology Department of Beijing Anzhen Hospital from January 2014 to September 2017.All specimen were stained with HE,Verhoef-Van Gieson,AB/PAS,and Masson. Then,the result of HE staining was compared with that of the combined special staining (Verhoef-Van Gieson,AB/PAS,and Masson).Results HE staining only showed a small amount of elastic fiber fracture. In contrast,the combined special staining clearly showed the pathological changes including fractures,decreased elastic fibers,and aggregation of extracellular mucous matrix. Diseases in these 999 patients included aneurysm of sinus of valsalva (SVA) (n=3),aortic root aneurysm (ARA) (n=177),thoracic aortic aneurysm (TAA) (n=78),abdominal aortic aneurysm (AAA) (n=17),total thoracoabdominal aortic aneurysm (tTAAA) (n=32),and aortic dissection (AD) (n=692). The median scores of combined special staining of SVA (Z=3.857,P=0.040),ARA (Z=14.307,P=0.000),TAA (Z=26.939,P=0.000),AAA (Z=22.412,P=0.000),tTAAA (Z=15.926,P=0.000),and AD (Z=39.213,P=0.000) were significantly higher than that of HE staining. Conclusion The combined Verhoeff-Van Gieson,AB/PAS,and Masson special staining is an effective technique for observing pathological changes of elastic fibers and mucus in blood vessels.

Abstract 316: Chronic Intermittent Hypoxia Attenuates BAPN Induced Thoracic Aortic Aneurysms in Mice

Objective: Aortic aneurysm (AA) is a life-threatening cardiovascular disorder due to the predisposition for dissection and rupture. Chronic intermittent hypoxia (CIH) has protective effects on heart and brain against ischemia injury. However, whether CIH prevents against AA formation was not elucidated. The present study aimed to investigate the effect of CIH treatment on thoracic aortic aneurysms (TAA) in mice. Approach and Results: Seventy-five C57BL/6 mice 3 weeks of age were divided into three groups, i.e. Control group ( 21% O 2 , 24 h per day, 4 weeks, n = 15); β-aminopropionitrile (BAPN) group (oral administration BAPN 1g / kg / day, 21% O 2 , 24 h per day, 4 weeks, n = 30); CIH+BAPN group ( 5% -21% O 2 90s / cycle, 8h per day, 4 weeks; BAPN 1g / kg / day, n = 30). We found that CIH significantly reduced the incidence of TAA (18 of 30 versus 9 of 30; p<0.05), and decreased maximal aortic diameter in BAPN group by hematoxylin and eosin staining (1.71 ± 0.35 versus 1.23± 0.15 mm; p <0.05). CIH significantly reduced elastin breaks in BAPN group by Verhoeff-van Gieson staining. The apoptosis of vascular smooth muscle cells was detected by TUNEL staining increased in BAPN group and markedly reversed by CIH. Moreover, RNA-Seq analysis and bioinformatics analysis were performed to screen the differential expression genes and possible pathway. CIH significantly down-regulated the expression of inflammation signaling pathways, up-regulated the expression of PI3K/AKT survival pathway and extracellular matrix homeostasis related pathway in BAPN induced TAA mice. Confirmed by real-time quantitative reverse-transcription-polymerase chain reaction (qRT-PCR) assays, CIH significantly inhibited the expression of IL-1β, TNF-α, MMP-9 and Cathepsin S, and increased the expression of hypoxia inducible factor-1α (HIF-1α), Lysyl oxidase (LOX), ITGB1 and COL1A2. Our findings also showed that the expression of MMP-9, MMP-2, cathepsin S, IL-1β, MCP-1 and Mac-2 increased in BAPN group and markedly decreased in CIH+BAPN group by immunohistochemistry staining. Besides, CIH also significantly increased the protein expression of HIF-1α, LOX and p-AKT by western blot analysis. Conclusions: The findings from our study suggest that CIH prevented the TAA development of BAPN-induced mice.

Abstract 312: Upregulation of Notch3 Contributes to Aneurysm Formation in a Murine Model of Marfan Syndrome

Notch3 functions as a receptor for membrane-bound ligands (Jag1, DLL1) in signaling pathways for cell-cell communication, gene regulation and control of cell differentiation during embryonic and adult vascular development. However, the role of Notch3 in Marfan Syndrome (MFS), which the main cardiovascular complication is thoracic aneurysms that results in high mortality, has yet to be fully elucidated. Previously we have shown that during aneurysm development, aortic smooth muscle cells (SMCs) in mgR mice, a murine model of MFS, display premature switch from a synthetic to a more contractile phenotype. Therefore we hypothesized that Notch3 could be playing a key role in this aberrant differentiation and aneurysm formation. To test this hypothesis we examined Notch3 mRNA and protein levels in the aortic tissue and SMCs from 1, 4, and 8 weeks-old mgR mice and wild type controls by real time-PCR, Western blot, and immunohistochemistry. Aortic histological changes were examined by Verhoeff-Van Gieson (VVG) elastin staining. In addition, mice were treated with DAPT (N-[N-(3,5-Difluorophenacetyl)-L-alanyl]-S-phenylglycine t-butyl ester), a known γ-secretase inhibitor. The effects of notch signaling inhibition on aneurysm rupture were evaluated. We found that aortic mRNA and protein levels of Notch3 in mgR mice was upregulated ~2 fold by 1 week, and 4-fold at 4 and 8 weeks compared to WT littermate controls. The expression levels of upstream and downstream targets of Notch3, Jagged1 and p21, respectively, were significantly increased in the aorta of mgR mice compared to wild type controls. DAPT treatment prolonged the median lifespan of mgR mice from 16 weeks (untreated) to 32 weeks (DAPT treated). The results from VVG staining of aortic tissue showed that DAPT treatment inhibited aortic elastic degradation in mgR mice. Taken together, our findings indicate that Notch3 is upregulated in the early aortic development and continues to increase during the course of the aneurysm formation in mgR mice. The increase in Notch3 expression may contribute to aneurysm development in MFS. This study lays the groundwork to further examine Notch3 as a novel potential target for treatment.

Elastic Staining in Differentiating Between Follicular Streamers and Follicular Scars in Horizontal Scalp Biopsy Sections.

Accurate identification of follicular streamers versus follicular scars on horizontal scalp biopsy sections is important to differentiate between cicatricial and noncicatricial alopecias. However, distinguishing between them can be difficult especially in nonoptimal specimens. The utility of elastic Verhoeff-Van Gieson (EVG) stain in cicatricial alopecias has been described on vertical sections, but its evaluation on horizontal sections has not been performed. Sixty-four cicatricial alopecias (25 lichen planopilaris, 29 central centrifugal cicatricial alopecias, and 10 discoid lupus erythematosus) and 53 noncicatricial alopecias (34 androgenic alopecia, 8 telogen effluvium, and 11 alopecia areata) were identified, and EVG staining was performed on horizontal sections. In follicular streamers, EVG highlighted an intact elastic network composed of delicate and thin elastic fibers circumferentially surrounding the angiofibrotic streamer without elastic network attenuation, loss, clumping, thickening, or recoil. In follicular scars, EVG demonstrated central attenuation and loss of the elastic network with peripheral clumping and recoil of elastic fibers. In cases where distinguishing between follicular streamers and scars is difficult, EVG may be a helpful adjunctive tool in this regard and aid in allowing better discrimination between cicatricial and noncicatricial alopecias.

Functional anatomy of the nasal bones and adjacent structures. Consequences for nasal surgery

The periosteum of the nasal bones, the periosteal-perichondrial nasal envelope, and the cartilaginous support of the bony vault were studied in serial coronal sections of four human cadaver noses. To differentiate between the various tissue components, the sections were stained according to Mallory-Cason and Verhoeff-Van Gieson stain. The results demonstrated: 1. the presence of clearly distinguishable layers of the periosteum covering the nasal bones; 2. the presence of a continuous periosteal-perichondrial covering of the bony and cartilaginous nasal vaults; 3. the way the cartilaginous support of the bony vault is constructed. The findings described in the present study may have clinical relevance in nasal surgery.

Effects of Aqueous Extract of Persea americana (Mill.) Leaf on the Myocardium of Left Ventricle of High Salt Fed Adult Wistar Rat

Introduction This study was designed to show that Persea americana extract possess the ability to protect the myocardium of left ventricle against injury caused by high salt diet in adult Wistar rats. Method Forty healthy Wistar rats of both sexes weighing 120–150 g were randomly assigned into 8 groups of 5 rats each (Groups A, B, C, D, E, F, G and H). Rats in groups A, F, G and H were fed with standard laboratory pellets, while groups B, C, D and E were fed on the high-salt diet for four weeks. Concomitantly, daily administration of 50 mg kg-1, 100 mg kg-1 and 150 mg kg-1 of the Persea americana extract were given orally to groups C&F, D&G and E&H respectively while rats in groups A and B were administered distilled water. The rats were sacrificed under ketamine anesthesia (30mg/kg i.m). The left ventricle of the heart was excised, processed in paraffin wax and stained with haematoxylin and eosin and Verhoeff-Van Gieson stains. One-way ANOVA was used to analyze data, followed by Student Newman-keuls (SNK) test for multiple comparison. Result Results revealed that there was statistically significant (p < 0.05) difference in body weight change across all experimental groups; which was significantly lower in high salt fed groups. It was revealed that there were morphological alterations in the myocardium of left ventricle in group B while Persea americana protected myocardium in other experimental groups. Conclusion In conclusion, high salt diet induced myocardium alterations which were significantly protected by oral administration of Persea americana extract.