Confluent transmural endothelialisation without hyperplasia progression in a loop graft isolation model

Abstracts / Cardiovascular Pathology 22 (2013) e29–e52

cause the failure of ECM components or trigger specific cellular responses in disease progression.

http://dx.doi.org/10.1016/j.carpath.2013.01.020

Heart rate variability with postural changes might be useful for detection of symptomatic mitral valve prolapse syndrome in Taiwanese patients Ya-chu Chena, Lung-Wen Tsaib, Ing-Fang Yangc, Chung-Kai Tsengd, Ten-Fang Yanga,b a National Chiao Tung University, Hsin Chu City, Taiwan b Taipei Medical University Hospital, Taipei City, Taiwan c Jen Chi General Hospital, Taipei City, Taiwan d China Medical University Hospital, Taichung City, Taiwan Purpose: To evaluate if HRV parameters with postural changes can be used to differentiate between symptomatic MVPS patients and normal controls. Methods: A total of 72 symptomatic patients (4 males and 68 females) had been echocardiographically diagnosed as having MVPS from the cardiology clinic and 101 healthy university students (51 males and 50 females) were recruited as normal control for the present study. A locally developed HRV system with one modified lead II ECG was used to record the tracing. All the records were taken during the daytime to avoid the influence of diurnal changes. The subjects were asked to rest at least 5 minutes before taking the records and postural alterations (lying, sitting and standing). Results: Time domain parameters between MVPS and normal group was statistically significantly different in lying and sitting position (Pb.05), whereas frequency domain parameters with postural changes were shown to have no significant differences in all postures. Conclusion: For time domain, HRV symptomatic MVP group is statistically significant different to normal control in lying and sitting positions. Lying and sitting female MVP Time domain parameters were statistically significant different to normal; whereas male only lying RMSSD and NN50 were significantly different. For the frequency domain, HRV cannot significantly differentiate symptomatic MVP group from normal control. Table 1 Time domain HRV parameters HRV parameters

Symptomatic MVP (n=72)

Normal group (n=101)

P value

SDNN (ms) lying RMSSD (ms) NN50 SDNN (ms) sitting RMSSD (ms) NN50 SDNN (ms) standing RMSSD (ms) NN50

37.42±24.45 29.91±24.58 28.65±36.99 36.87±17.43 28.78±16.05 36.06±48.64 32.21±16.95 20.22±16.11 11.93±22.73

51.50±21.38 41.41±18.38 64.68±49.99 50.50±22.36 35.74±16.02 53±45.22 36.23±14.41 18.47±9.51 11.23±18.79

.00 .00 .00 .00 .00 .01 .05 NS NS

http://dx.doi.org/10.1016/j.carpath.2013.01.021

A research of heart rate variability between symptomatic mitral valve prolapse syndrome and normal patients Ya-chu Chena, Lung-Wen Tsaib, Ing-Fang Yangc, Chung-Kai Tsengd, Ten-Fang Yanga,b a National Chiao Tung University, Hsin Chu City, Taiwan b Taipei Medical University Hospital, Taipei City, Taiwan c Jen Chi General Hospital, Taipei City, Taiwan d China Medical University Hospital, Taichung City, Taiwan

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Purpose: To evaluate if HRV parameters is different between MVPS and normal patients. Methods: A total of 72 symptomatic echocardiographically documented MVPS patients (4 males and 68 females) and 101 healthy students (51 males and 50 females) were recruited as normals for this study. A local HRV system with one modified lead II was used. All recordings were taken during daytime to avoid diurnal ANS influences. The subjects were asked to rest 5 minutes before recording and postural alterations (lying, sitting, and standing). Results: For MVPs, no difference was found in time domain between male and female. For normal male and female, frequency domain was statistically significantly different only in LF/HF ratio, whereas postural changes were shown to have significant effects in LF/HF ratio in all postures in frequency domain. Table 1 Frequency domain HRV: FFT and AR LF/HF ratio HRV parameters

MVP male (n=4)

MVP female (n=68)

P value

Normal male (n=51)

Normal female (n=50)

P value

FFT LF/HF (lying) AR LF/HF (lying) FFT LF/HF (sitting) AR LF/HF (sitting) FFT LF/HF (standing) AR LF/HF (standing)

2.84 ± 2.84

1.18 ± 0.97

0.04

1.65 ± 0.97

0.96 ± 1.16

0.0009

2.89 ± 2.89

1.19 ± 0.97

0.04

1.67 ± 0.95

0.96 ± 1.15

0.0006

2.29 ± 2.29

1.31±1.10

0.03

2.45 ±1.91

1.60 ±1.77

0.01

2.06 ±2.06

1.34 ±1.17

NS

2.52±2.09

1.64 ±1.86

0.01

3.60 ±1.80

2.95±4.38

NS

5.09 ±3.34

3.64 ±3.53

0.02

2.95±3.88

2.89 ±3.89

NS

5.38±4.08

3.91 ±4.09

0.04

⁎Pb.05 statistically significant.

Conclusion: For time domain, HRV cannot be used to differentiate male from female in all positions, but the sample size is not large enough. For frequency domain, HRV LF/HF ratio might be a useful tool in all three postures for the detection of gender differences in symptomatic MVPS and normal group.

http://dx.doi.org/10.1016/j.carpath.2013.01.022

Confluent transmural endothelialisation without hyperplasia progression in a loop graft isolation model Tim Pennel University of Cape Town Purpose: We have recently described an isolation looped vascular graft model that clearly separates transmural from transanastomotic endothelialisation as distinctly discrete events. The present study used the same isolation model to demonstrate that neointimal tissue formation accompanying transmural endothelialisation reaches an early, stable equilibrium. Methods: High-porosity polyurethane (PU) grafts (ID1.7 mm; 150 μm pore size) were interposed (‘welded’) between low-porosity ePTFE (ID1.7 mm; IND 15–25 μm) segments and implanted in the abdominal aorta of Wistar rats for 6, 8, 12 and 24 weeks (n=8/time point). Looping the graft increased their length to 8cm. Light, immune-fluorescence (CD31) and scanning electron microscopy were used for image analysis. Results: The transanastomotic outgrowth edge never traversed the ePTFE endothelial-free zone on the proximal or distal segment (proximal: 25.93±5.90 mm at 6 weeks and 8.71±4.94 mm at 24 weeks, P=.0005), resulting in 100% isolation from the mid-graft endothelium. Transmural midgraft endothelial coverage reached preconfluence at 6 weeks (85±15%) and confluence between week 12

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Abstracts / Cardiovascular Pathology 22 (2013) e29–e52

(94±11%) and 24 (85±28%). One of the 24-week loop grafts did not anchor in the retroperitoneum resulting in the absence midgraft endothelial coverage. There was no progression of mid-graft neointimal tissue hyperplasia at 6, 8, 12, and 24 weeks (54.99±52.02 μm; 45.00±58.39; 45.84±46.60; 47.36±37.08 μm, P=.77). Conclusions: A looped interposition-graft model provides sufficient isolation length to clearly distinguish transanastomotic from transmural endothelialisation for up to half a year. Ingrowth from surrounding tissue appears essential for transmural mid-graft endothelialisation. The lack of ongoing mid-graft luminal narrowing confirms that sub- endothelial tissue proliferation due to transmural growth is of no concern.

http://dx.doi.org/10.1016/j.carpath.2013.01.023

The effects of waveform on the generation of flow dependent phospho-Smad2 and Akt species Alex Obrejanu, Robert D. Shepherd, Kristina D. Rinker University of Calgary, Calgary, AB, Canada Purpose: Endothelial cells are affected by fluid flow through mechanotransduction-induced signaling events. We previously identified a novel signaling pathway, involving Smad2 and Akt, in which linker phosphorylated Smad2 is generated and translocates to the nucleus in a flow-dependent manner. The Smad co-repressor transforming growth interacting factor (TGIF) also localizes in a similar fashion. Levels of Smad2 were shown to follow the pattern of Akt phosphorylation through inhibitor studies. Due to the nature of our system, however, we were not able to determine if these events were controlled by the magnitude of fluid shear stress, or if they were dependent on flow waveform. The work presented here was undertaken to address this question. Methods: HAEC were exposed to 20 hours of 2 and 10 dyne/cm2 shear stress from both pulsatile and steady flow in a parallelplate flow loop. Expression levels of molecular targets were determined by real-time RT-PCR and western blotting. Tissue samples from a partial carotid ligation mouse model, in which oscillatory flow was induced in the affected artery, were analyzed using confocal microscopy. Results: Akt phosphorylation profiles showed no significant difference between flow patterns, whereas linker Smad2 phosphorylation was less robust under steady flow relative to pulsatile. Each phosphorylation increased at higher shear stress. Murine tissue studies were largely consistent with in vitro studies. Conclusions: The modulation of Smad2 and Akt signaling from fluid flow is complex, involving both shear stress magnitude and flow patterning. Further work is necessary to completely define this pathway's controlling parameters.

http://dx.doi.org/10.1016/j.carpath.2013.01.024

Effects of blood volume expansion fluids on human aortic endothelial cells Kogan Leea, Andrew Walkera, Robert D. Shepherda, Clifton Johnstonb, Gary Dobsonc, Kristina D. Rinkera a University of Calgary, Calgary, AB, Canada b Dalhousie University, Halifax, Nova Scotia, Canada c Alberta Health Services, Calgary, AB, Canada

Purpose: Blood volume replacement is a critical issue for individuals with severe blood loss or hypovolemic shock. However, it is unclear

whether crystalloid or colloid solutions are most appropriate for fluid resuscitation. Crystalloid solutions replace volume on a less than equivalent basis, and infusion of large volumes is associated with edema and pulmonary complications. Hydroxyethyl starch (HES) colloids resist hydrolysis, allowing them to be effective osmotic agents, and induce plasma volume expansion of up to 1.45 times the infused volume. HES have also been reported to positively influence endothelial physiology, while simultaneously exhibiting some negative clinical effects. This study investigated endothelial exposure to both crystalloid and HES under static and flow conditions to evaluate impacts on inflammatory mediators. Methods: HAEC were treated with 25% (v/v) pentaspan, voluven, and normal saline, supplemented with dexamethasone and/or TNF-alpha as required. Monocyte adhesion molecules expression was determined by immunoblotting, and monocyte adhesion was measured by microscopy. Viscosity dependent shear stresses were determined for flow studies. Results: Each of the solutions tested impacted adhesion molecule expression in static culture, with the HES reducing ICAM-1 and VCAM-1. This trend was maintained upon pre- treatment with TNFalpha, and supported by monocyte adhesion data. Under flow, saline increased adhesion molecule expression, while Voluven provided mixed results. Viscosity dependent shear stress also participated in determining experimental outcomes. Conclusions: HES appear to be more beneficial than saline to HAEC physiology based upon two factors; their chemical effect and their ability to maintain blood viscosity at near normal values.

http://dx.doi.org/10.1016/j.carpath.2013.01.025

Collagen fiber architecture in the longitudinal-radial plane of ascending aorta is distinctly different among bicuspid aortic valve and tricuspid aortic valve patients with ascending aortic aneurysm Alkiviadis Tsamisa, Julie A. Phillippia, Salvatore Pastab, Antonio D'Amorea,b, Simon C. Watkinsa, William Wagnera, David A. Vorpa, Thomas G. Gleasona a University of Pittsburgh, Pittsburgh, PA b Fondazione RiMED Palermo, Italy Purpose: To characterize the fiber architecture in the longitudinal (LONG)-radial (RAD) plane of human ascending thoracic aortic aneurysm (ATAA) and control ascending thoracic aorta (ATA). We hypothesize that previously reported lower delamination strength of ATAA with bicuspid aortic valve (BAV) compared with tricuspid aortic valve (TAV) is related to fiber architectural anomalies in the LONGRAD plane of the tissue. Methods: Artificially dissected human tissue samples from BAVATAA, TAV-ATAA and control ATA were examined under multi-photon microscopy. The medial-adventitial and medial-intimal halves were viewed in the LONG-RAD plane. Images were processed using an automated image-based tool to obtain the fiber amplitude of angular undulation (AAU). Results: For collagen fibers of the medial-intimal half, AAU was lower in BAV compared with TAV-ATAA and unchanged from control ATA. AAU was higher in TAV-ATAA compared with control ATA. These results suggest that collagen fibers in this region of the aorta are less undulated about LONG axis in BAV than in TAV-ATAA, and more undulated about LONG axis in TAV-ATAA than in control ATA. In the medial-adventitial half, the AAU of collagen was higher in BAV and unchanged in TAV-ATAA compared with control ATA. Conclusions: The weakened delamination strength previously observed for BAV-ATAA may be related to the present findings of reduced collagen fiber undulation about the LONG axis in the medial-