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Pericardial patches are commonly used in vascular surgery to close arteriotomies. The mechanism of early healing after patch implantation is still not well defined. We used a rat aortic patch model to assess pericardial patch healing and examined Ephrin-B2, a marker of arterial identity, expression within the post-implantation patch. We also determined whether endothelial progenitor cells (EPC) are associated with early patch healing in the arterial environment.
Wistar rats (200–250 grams) underwent infrarenal aortic arteriotomy and then closure via bovine or porcine pericardial patch angioplasty. Control groups included subcutaneously implanted patches. Patches were harvested at 0–30 days and analyzed by histology, immunohistochemistry, immunofluorescence and Western blot as well as quantitative PCR.
Prior to implantation, pericardial patches are largely composed of collagen and are acellular. Following arterial implantation, increasing numbers of CD68-positive cells as well as Ephrin-B2 and CD34 dual-positive cells are found within both bovine and porcine pericardial patches, whereas the infiltrating cells are negative for vWF and α-actin. Porcine patches have a luminal monolayer of cells at day 7, compared to bovine patches that have fewer luminal cells. Subcutaneously implanted patches do not attract Ephrin-B2/CD34-positive cells. By day 30, both bovine and porcine pericardial patches develop a neointima that contains Ephrin-B2, CD34, and VEGFR2-positive cells.
Both CD68-positive and Ephrin-B2 and CD34 dual-positive cells infiltrate the pericardial patch early after implantation. Arteriotomy closure via pericardial patch angioplasty shows patch adaptation to the arterial environment that may involve a foreign body response as well as localization of EPC. Arterial remodeling of pericardial patches support endothelialization and may represent a paradigm of healing of scaffolds used for tissue engineering.
Citation: Li X, Jadlowiec C, Guo Y, Protack CD, Ziegler KR, Lv W, et al. (2012) Pericardial Patch Angioplasty Heals via an Ephrin-B2 and CD34 Positive Cell Mediated Mechanism. PLoS ONE 7(6): e38844. https://doi.org/10.1371/journal.pone.0038844
Editor: Gian Paolo Fadini, University of Padova, Italy
Received: January 23, 2012; Accepted: May 11, 2012; Published: June 13, 2012
This is an open-access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.
Funding: This work was supported by the National Institute of Health (R01-HL095498-01 to AD); the American Vascular Association William J. von Liebig Award; as well as with the resources and the use of facilities at the Veterans Affairs Connecticut Healthcare System, West Haven, Connecticut. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Competing interests: Bovine pericardial patches (Vascuguard) were kindly provided by Synovis Life Technologies (St. Paul, Minnesota, United States of America). Porcine pericardial patches (dCELL) were kindly provided by Tissue Regenix Ltd (York, United Kingdom). This does not alter the authors' adherence to all the PLoS ONE policies on sharing data and materials, as detailed online in the guide for authors.
The use of patch angioplasty during carotid endarterectomy and other arterial closure has become standard in vascular surgery as patch use is associated with improved patient outcomes including reduced risk of restenosis [1], [2], [3]. Debates regarding the optimal patch material, however, continue, and have prompted investigation into which material offers optimal performance. The bovine pericardial patch was first introduced in the 1990s and has become a commonly used material for closure of the surgical angioplasty site [4], [5]. Pericardium possesses many merits including biocompatibility, easy handling, minimal suture line bleeding, and ability to perform Doppler investigations through the patch. Interestingly, despite use of bovine pericardial patches in human patients for almost two decades, long-term results for this biomaterial remain poorly documented, and the mechanism of healing after patch implantation likewise remains unknown. In particular, it is not even known whether cells infiltrate the patch or whether these patches support endothelialization when placed into the arterial circulation.
Historically, bioprosthetic heart valves were one of the first successful applications of engineered tissue, and the introduction of gluteraldehyde preservation in 1969 only served to further enhance their clinical utility. Preliminary results using early unfixed bioprosthetic tissues found them to be immunogenic and susceptible to early degradation, weakening, and failure. The application of gluteraldehyde cross-linking to these engineered tissues decreased the observed immunogenicity and likewise increased product durability in vivo. The success of this application quickly became evident in the increasingly common usage of bioprosthetic cardiac valves. Similarly, the gluteraldehyde-fixed umbilical cord was also an important successful arterial conduit for human patients with peripheral vascular disease [6], [7].
Along with increased clinical usage came the early observation that high concentrations of gluteraldehyde used for preservation allowed for less in vivo remodeling and had an increased tendency to calcify [8]. Newer generation bioprosthetics were able to successfully use lower concentrations of gluteraldehyde while preserving reduced immunogenicity [9]. Modern bioprosthetics, such as the latest generation of the bovine pericardial patch, continue to use low concentrations of gluteraldehyde in combination with decellularization, the removal of cellular and nuclear material; in addition the gluteraldehyde is removed prior to surgical implantation into patients. These methods have led to the most recent generation of bioprosthetic patches that may allow for in vivo remodeling and adaptation. Studies using the rat subcutaneous implantation model have found that these properties allow for fibroblast ingrowth and cellular incorporation coupled with preservation of durability [9]. However, it is not known to what extent remodeling and arterial adaptation, such as endothelialization, occur with patch closure, as well as any differences that might occur in pericardial patches that are not fixed with gluteraldehyde.
Prior work in vein grafts has shown the complexity of venous adaptation to the arterial environment [10]. We have previously shown that arterialized vein grafts show increased cellularity and matrix in all layers of the vessel wall, as well as diminished expression of the venous marker Eph-B4, yet they fail to express Ephrin-B2, a marker of arterial identity [11]. Since pericardium is not derived from a vessel, and therefore does not express either Ephrin-B2 nor Eph-B4, it is not clear whether this biomaterial is capable of adopting an arterial identity and expressing Ephrin-B2 [12].
Accordingly, we examined commercially-available pericardial patches in an arterial patch closure model to determine the mechanisms by which these patches support arterial healing. We hypothesize that once placed into an arterial environment, the pericardial patches will support cellular influx and these entering cells express the arterial marker Ephrin-B2 in addition to CD34, a marker of neovascularization; in addition, we hypothesize that pericardial patches support endothelialization of the patch on its luminal surface. We also tested the hypothesis that pericardial patches that are not crosslinked with gluteraldehyde have similar patterns of healing.
Animal Model
All experiments were approved by the Institutional Animal Care and Use Committee at the Yale University School of Medicine (IACUC# 2010–10896). Young (6–8 week-old) male Wistar rats (mean weight, 209±34 g) were use for patch implantation (n = 24). Anesthesia was performed via isoflurane inhalation with surgery performed using a dissecting microscope (Leica MZ 95, Germany). An abdominal midline incision was made and the infrarenal abdominal aorta was exposed and dissected free of surrounding structures. Following subcutaneous heparin injection (100 U/100 g) the infrarenal aorta was clamped. A longitudinal 3 mm arteriotomy was then made on the anterior aortic wall and a pericardial patch (3 mm×1 mm) was sutured in place using interrupted 10-0 nylon (Figure 1; bovine: Synovis Life Technologies, St. Paul, MN, USA; porcine: Tissue Regenix, York, United Kingdom). After completion of the angioplasty, the micro-clamps were removed and aortic flow was restored. The abdomen was then closed using 5-0 Dacron sutures. Additional age-matched Wistar rats were used for subcutaneous patch implantation (n = 12). Animals were sacrificed on postoperative days 1, 3, 7, or 30. No immunosuppressive agents were given at any time.
A) The rat aorta after arteriotomy and prior to patch placement; the white arrow shows the glistening intima on the posterior wall; ruler markings are 1mm. Scale bar, 1000 microns. B) Completed bovine patch closure. Scale bar, 1000 microns. C) Completed porcine patch closure. Scale bar, 500 microns. D) Explanted aortic segment containing patch after harvesting and flushing. Scale bar, 1000 microns. In each graph, A = aorta; P = pericardial patch.
Rats were anesthetized with isoflurane, and tissues were fixed by transcardial perfusion of phosphate buffered saline (PBS) followed by 10 percent formalin. Tissue was removed and fixed overnight in 10 percent formalin followed by a 24-hour immersion in 70 percent alcohol. Tissue was then embedded in paraffin and sectioned. H&E stained sections were used for cell counts; 4 random representative high power fields were counted from each patch and the mean number of cells in each high power field recorded.
Tissue sections were de-paraffinized using xylene and a graded series of alcohols. Sections were then incubated in peroxidase blocking solution and stained using antibodies against von Willebrand Factor (Dako, A0082), α-actin (Thermo Scientific, MS-113), or CD68 (Thermo Scientific, MS-397) overnight at 4°C. Immunohistochemical detection was performed using DAB as well as NovaRED substrate (Vector). Sections were counterstained with Mayer's Hematoxylin.
Tissue sections were de-paraffinized and stained using primary antibodies against CD31 (BD Pharmingen, 550300), CD34 (R&D, AF4117) Ephrin-B2 (Novus, NBP1-48610), or ED-1 (Abcam). Appropriate fluorophore-conjugated secondary antibodies were selected. An antifade reagent with DAPI nuclear stain (Invitrogen, P-36931) was applied prior to cover slip placement.
PCR Analysis
Total RNA was isolated from tissue and patches using TRIzol Reagent (Invitrogen) and RNA was cleaned using the RNeasy Mini kit with digested DNase I (QIAGEN). For quantification of total RNA, each sample was measured using the RiboGreen RNA Assay kit (Invitrogen). RT was performed using the SuperScript III First-Strand Synthesis Supermix (Invitrogen) according to the manufacturer's instructions. Real-time quantitative PCR was performed using SYBR Green Supermix (Bio-Rad Laboratories) and amplified for 40 cycles using the iQ5 Real-Time PCR Detection system (Bio-Rad Laboratories). All samples were normalized by GAPDH amplification. The sequences for the primers to amplify Ephrin-B2 and GAPDH were synthesized (Keck Oligonucleotide Synthesis Facility, Yale University, New Haven, CT) and are shown in Table 1. Primers for CD34 were purchased (SABiosciences, PPR55773A-200). Patches were analyzed individually, without combination of samples.
Western Blot Analysis
For protein extraction, the inferior vena cava, the pericardial patch, and the thoracic aorta were carefully harvested separately and snap-frozen in liquid nitrogen. Samples were crushed and mixed with buffer including protease inhibitors (Roche, Complete Mini 12108700) prior to sonication (5 sec) and centrifugation (800 g, 20 min). Equal amounts of protein from each experimental group were loaded for SDS-PAGE, followed by Western blot analysis. The membranes were probed with antibodies against Ephrin-B2 (Novus, NBP1-48610), CD34 (R&D, AF4117), or GAPDH (Cell Signaling, 14C10). Membrane signals were detected using the ECL detection reagent (Amersham, Buckinghamshire, UK). Patches were analyzed individually, without combination of samples.
Bovine patches
To define the pattern of arterial healing with a common clinically-used pericardial patch, bovine pericardial patches were examined in a rat angioplasty model (Vascuguard; Synovis Life Technologies, St. Paul, MN, USA). Prior to implantation bovine pericardial patches are acellular (Figure 2). After implantation into the rat aorta, cells increasingly infiltrated into the patch, between the collagen fibers, with the number of infiltrating cells directly correlating with length of time implanted (Figure 2). Cells were present throughout the angioplasty patch; however, with increasing time, cells migrated more centrally into the interior of the patch compared to the fewer number of cells present on the patch surface (Figure 2). Although subcutaneously implanted patches also had an increasing cellular infiltrate, this infiltrate remained predominantly on the periphery of the patch (Figure 2).
A) Photomicrographs of bovine pericardial patches, implanted into either an arterial or subcutaneous position, and shown at either high or low magnification. Scale bar, 40 microns. B) Graph of cell count in the patch over time; mean of 4 high power fields per patch; n = 2. *, p<0.0001.
Arterial patches were also examined during the first week after implantation for potential markers of cellular identity on the infiltrating cells (Figure 3). At these early time points, an endothelial cell layer was not readily apparent; infiltrating cells were negative for vWF and α-actin, markers of vascular endothelial cells and smooth muscle cells respectively, at all time points (Figure 3). Immunofluorescence for CD31, an alternative vascular endothelial cell marker, confirmed lack of endothelial cells in arterial patches on days 1, 3, and 7 (data not shown). Some of the infiltrating cells stained positively for CD68, consistent with their being macrophages (Figure 3). In comparison, the cells infiltrating into subcutaneously implanted patches developed positive staining for vWF, α-actin and CD68 by postoperative day 7 (Figure 3).
Scale bar, 10 microns, except 40 microns for subcutaneous histology. n = 3; representative pictures are shown. Immunofluorescence shows CD34 and Ephrin-B2-positive cells colocalize in the middle of the bovine arterial patch, day 7. Photomicrographs of immunofluorescence: blue color, DAPI; green color, CD34; red color, Ephrin-B2; merge, green + red. n = 3; representative pictures are shown. Scale bar, 10 microns. Similarly, merge of CD34 and VEGFR2 is shown. Bar graphs show increased expression of Ephrin-B2 and CD34 mRNA and protein in arterial, but not subcutaneous, patches. n = 4−5. Panels A, B, D, and E, mRNA; Panels C and F, protein.
Since some of the cells infiltrating into arterial patches were not positively stained with CD68, we examined these cells for markers that would suggest endothelial or arterial progenitor cell identity. Arterial patches (day 7) were examined using immunofluorescence for both CD34 and Ephrin-B2 (Figure 3). Approximately half of the infiltrating cells were positive for CD34 and the number of cells appeared to increase, in a centrally concentrated position, over time; similarly, Ephrin-B2 was detected on these cells in a mostly similar pattern and that typically colocalized with CD34 (Figure 3). However, cells infiltrating into the subcutaneously implanted patches stained negative for both CD34 and Ephrin-B2 (Figure 3). Similarly, some of the cells infiltrating into arterial patches were dually-positive for CD34 and VEGFR2 by day 7, and none of these cells were detected in subcutaneously implanted patches (Figure 3).
To confirm the presence of Ephrin-B2/CD34 dual-positive cells infiltrating into the arterial patches but not into the subcutaneously placed patches, patches were explanted postoperatively and mRNA extracted from them. qPCR analysis showed that arterial patches had significant expression of both Ephrin-B2 (Figure 3A) and CD34 (Figure 3B) postoperatively, whereas there was minimal Ephrin-B2 (Figure 3D) and CD34 (Figure 3E) expression present in subcutaneously placed patches,. The increase in Ephrin-B2 expression in arterial patches also increased over time when compared to the amount present in the inferior vena cava, although it did not reach the amount present in aorta (data not shown).
Patches were similarly examined for protein expression (Figure 3). Western blot analysis confirmed the presence of Ephrin-B2 and CD34 protein expression within the patches when placed in the arterial (Figure 3C) but not the subcutaneous position (Figure 3F). All levels of protein expression were lower in the angioplasty patch groups when compared to native aorta; no protein expression was found in the pre-implantation patch (Figures 3C, 3F).
Porcine patches
To determine the effect of gluteraldehyde cross-linkage of the patch, we examined porcine pericardial patches that are decellularized by a different manufacturing process that does not use gluteraldehyde (dCELL; Tissue Regenix, York, United Kingdom). Examination of these porcine patches with immunofluorescence shows that these patches are acellular prior to implantation (Figure 4), similar to bovine patches (Figure 3). Also similar to bovine patches, cells infiltrating into the porcine patches stained positively for CD34, Ephrin-B2, VEGFR2, or CD68 (Figure 4). However, unlike bovine patches that showed predominantly cellular infiltration into the middle of the patch by day 7 (Figures 2, 3), the luminal surface of the porcine patch is covered by cells as early as day 7 after implantation; in addition, these luminal cells are CD34 positive, with some of the cells also being CD34 and Ephrin-B2 dual positive (Figure 4) as well as CD34 and VEGFR2 dual positive (Figure 4). Western blot confirmed that the patches contain Ephrin-B2 and CD34 protein at day 7 (Figure 4B).
7. A) Porcine pericardial patches show infiltrating cells positive for CD34, Ephrin-B2, VEGFR2, or CD68; CD34-positive cells are present as a surface monolayer, with some cells dually positive for CD34 and Ephrin-B2 or CD34 and VEGFR2; first panel is the day 0 (preimplantation) control. Photomicrographs of immunofluorescence: blue color, DAPI; green color, CD34; red color, Ephrin-B2 or VEGFR2; merge, green + red. n = 3; representative pictures are shown. Scale bar, 10 microns. B) Western blot of Ephrin-B2 and CD34, in porcine pericardial patch, day 0 (preimplantation) or day 7.
Patches were followed to 30 days; both bovine and porcine patches demonstrated a thick layer of luminal neointima (Figure 5A) that was of similar thickness between bovine and porcine patches (Figure 5B). Examination of the explanted 30-day patches with immunofluorescence showed that the neointima of both patches was composed of cells that were positive for CD34, Ephrin-B2, and VEGFR2, with minimal differences between bovine and porcine neointima (Figure 5C). Western blot confirmed the presence of Ephrin-B2 and CD34 protein in both bovine and porcine pericardial patches (Figure 5D).
30. A) Representative photomicrographs of bovine and porcine patches, low (10x) and high (40x) magnification, scale bar 80 and 30 (porcine, 40) microns. Arrow shows the neointima; arrowhead shows the patch; L, lumen. B) Bar graph of mean neointimal thickness, bovine and porcine patches, day 30. C) Photomicrographs of immunofluorescence: blue color, DAPI; green color, CD34; red color, Ephrin-B2 or VEGFR2; merge, green + red. n = 2; representative pictures are shown. Scale bar, 10 microns. D) Western blot of Ephrin-B2 or CD34, in bovine or porcine pericardial patch, day 0 (preimplantation) or day 30; n = 2.
We show that pericardial patches support cell infiltration after implantation into the arterial circulation. Within the first week of implantation of a bovine pericardial patch into an arterial position but not in a subcutaneous position, infiltrating cells express CD68, Ephrin-B2 as well as CD34, but not vWF or alpha-actin, in the middle of the patch and with fewer cells on the luminal surface. Porcine pericardial patches show a similar pattern of cells infiltrating into the middle of the patch, with additional detection of luminal cells that are positive for CD34, Ephrin-B2, and VEGFR2. These results suggest that surgical closure of an artery with a pericardial patch supports arterial remodeling by endothelialization of the luminal surface, at least in the short term, not by ingrowth of endothelial cells or smooth muscle cells, but rather by influx of arterial progenitor cells. However, by 30 days, both bovine and porcine patches develop similar amounts of luminal neointima, suggesting that long term patency is maintained by endothelialization, regardless of patch origin.
Acquisition of an arterial identity in the patch
A critical finding of this study is the detection of infiltrating cells that express the arterial identity molecule, Ephrin-B2, as well as a time-dependent increase in Ephrin-B2 mRNA expression in the patches. This finding suggests that the mean number of Ephrin-B2 positive cells increases within the patch in time-dependent manner. Ephrin-B2 and its receptor Eph-B4 are differentially expressed in arteries and veins, with Ephrin-B2 an arterial marker and Eph-B4 a venous marker [13], [14]. The Ephrins and the Ephs are the ligands and receptors, respectively, of the largest of the 14 subfamilies of the receptor tyrosine kinase families [15]. In the developing cardiovascular system, ephrin and Eph molecules control the angiogenic remodeling of blood vessels and lymphatic vessels and play essential roles in endothelial cells as well as in supporting pericytes and vascular smooth muscle cells [16]. However, not only are Ephrin-B2 and Eph-B4 expressed during embryonic development and determination of the vasculature, but Ephrin-B2 and Eph-B4 remain persistently expressed in adult vessels. We have previously shown that Eph-B4 is active in adult veins and regulates venous remodeling [17].
During vein graft adaptation to the arterial circulation, veins lose their expression of Eph-B4, but they also fail to express an arterial identity and do not increase expression of Ephrin-B2 or members of the delta-notch pathway [11], [18]. However, we show that pericardial patches, unlike vein grafts, are capable of gaining arterial identity, suggesting that pericardial patches can adapt to the arterial circulation. Interestingly, arterial identity appears to be expressed by cells infiltrating into the patch, whereas vein graft adaptation is characterized by intimal hyperplasia intrinsic to the vein graft, with smooth muscle cell proliferation [11].
EPC play a role in patch remodeling
EPC play a critical role in vascular remodeling and adaptation. EPC elicit neovascularization in response to ischemia, and repair injured endothelium [19]._ENREF_15 Accordingly, autologous EPC therapy has emerged as a promising treatment option for critical limb ischemia secondary to its ability to promote neovascularization and remodeling of pre-existent vessels [20], [21], [22]. EPC are characterized by their unique surface marker expression, and although many markers have been studied and described, CD34 and VEGFR2 expression continue to be a recognized identification method [19], [23]. We used the marker CD34 to identify progenitor cells within the implanted patch, and expression of CD34 increased in a time-dependent manner from day-1 through day-7, and continued to be present in the neointima at day 30. Since the definition of EPC is not completely agreed upon, with lack of consensus as to the definitive definition and markers [24], [25], it is clear that our identification of CD34-positive cells may not definitively identify them as classic EPC. However, colocalization of CD34-positive cells with Ephrin-B2 as well as VEGFR2 (Figure 3) suggests that these cells infiltrating the patch are arterial progenitor cells, although CD34 is present on surface endothelial cells, preventing our ability to distinguish EPC from endothelial cells (Figures 4, 5). Nevertheless, finding arterial progenitor cells infiltrating into the patch is consistent with the data that have found that coronary stent implantation may induce mobilization and differentiation of bone marrow cells into endothelial or smooth muscle cells [26]. EPC promote vascular graft patency, and circulating EPC play an important role in endothelial protection and integrity [27], [28]. Current clinical trials are attempting to exploit these characteristics by designing coated stents that attract EPC in situ, with the belief that stents that are capable of achieving this will have improved adaptation and be better equipped to heal endothelium [29], [30]. Application of these findings to the pericardial patch may help direct more EPC to the patch and aid in transfer of endothelial cells.
Subcutaneous patches fail to attract EPC
Examination of the subcutaneously implanted patches did not reveal the presence of either CD34-positive or Ephrin-B2-positive cells, thus failing to show arterial adaptation. As compared to the arteriotomy patches, subcutaneously patches lacked exposure to arterial shear flow as well as EPC exposure from within the circulating blood pool. A confounding factor is the reduced surgical trauma and associated lesser inflammatory response of a subcutaneous implantation compared to an abdominal laparotomy, since EPC are mobilized from the bone marrow into the peripheral circulation in response to surgical trauma [26], [31].
Macrophages are known mediators of inflammation and their presence within surgical sites is well established. As expected, a large number of inflammatory cells were identified in the neighboring tissues surrounding the patch, with many cells infiltrating into the subcutaneously placed patch. We observed some macrophages infiltrating into the arterial patches, although it appeared that fewer macrophages infiltrated into the arterial patches compared to the large numbers that infiltrated into the subcutaneously placed patches.
One difference that might account for these results is the effect of the patch environment on macrophage recruitment. The extracellular matrix adjacent to the subcutaneous patch may facilitate macrophage migration as compared to the environment of the arterial patches. Arterial flow and/or pressure may exert a protective effect on the angioplasty patches that alters the inflammatory response, although the outside surface of the arterial patch is still exposed to the retroperitoneal tissues. This suggests that macrophages may need a specific signal to infiltrate the patch, or that the blood flow and/or pressure inside the vessel prevents a chemoattractive signal gradient from being established. An alternative consideration is that the early time course in this study may prevent observation of a later macrophage influx. Since our model only describes the course of healing until 30 days after implantation, longer time data points are needed for a full description of patch healing.
Bovine and porcine patches develop neointima
Although porcine patches show earlier luminal cells compared to bovine patches (Figure 4), both patches develop a similar amount of neointima on their luminal surface by 30 days (Figure 5). As such, the significance of the earlier endothelialization of the porcine patches is not clear, as it may simply reflect differences in timing between bovine and porcine patch healing. Examination of bovine patches processed without gluteraldehyde could potentially examine this issue, but these patches are not currently available for surgical implantation in the vascular system. However, since both patches develop luminal neointima by 30 days, we believe that the similarities between the bovine and porcine pericardial patches, despite potential differences in their manufacturing processes, will lead to similar clinical outcomes when used in human patients. Since both patches support endothelialization and development of luminal neointima, this mode of healing predicts favorable long-term outcome, with good patency rates and minimal rates of thrombosis, infection, or pseudoaneurysm formation [3], [32].
In summary, the results from our surgical angioplasty model show that the pericardial patch is capable of supporting luminal endothelialization, promoting vessel remodeling and arterial adaptation. Ephrin-B2, CD34 and VEGFR2 positive cells were successfully identified within the implanted patch and may represent arterial or EPC. Increased Ephrin-B2 expression in the healing artery is distinctly different that the loss of Ephrin-B2/Eph-B4 expression characteristic of vein graft adaptation, suggesting that arterial and venous remodeling are fundamentally different. Arterial remodeling of pericardial patches may represent a paradigm of healing of scaffolds used for tissue engineering.
Bovine pericardial patches (Vascuguard) were kindly provided by Synovis Life Technologies (St. Paul, MN, USA). Porcine pericardial patches (dCELL) were kindly provided by Tissue Regenix Ltd (York, United Kingdom).
Author Contributions
Conceived and designed the experiments: XL CS AD. Performed the experiments: XL CJ YG CP KZ WL CY. Analyzed the data: XL CJ YG CP KZ WL CY CS AD. Contributed reagents/materials/analysis tools: CJ YG CP KZ WL CY. Wrote the paper: XL CS AD. Approved the final manuscript: XL CJ YG CP KZ WL CY CS AD.
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Celebrity Quotes of the Week: Keke Palmer Checks Wendy Williams on Her Purple Couch | Keke Palmer | Celebrities | BET
Plus, words from Mahershala Ali, Taraji P. Henson and more.
Plus, words from Mahershala Ali, Taraji P. Henson and more.
Everything that immediately required a screenshot.
Suddenly, we're craving chocolate.
The Saved by the Bell star is prepared to fight.
Starring Queen Latifah as the blues icon, the film breaks down barriers.
The actress was blasted for her remarks related to First Lady Michelle Obama.
The Just Keke host offers real relationship advice and spills her celebrity crushes.
Audience member Ashley Joy gets a glam-cam makeover for her very first Instagram photo, which will feature Keke Palmer.
The Playmakers — Kevin, Jason and Anthony — discuss their popular YouTube videos that they say poke fun at the Black church all in love.

subset: commoncrawl


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Prot Pally 906e 54 trait 3/10M Prot Paladin LF Mythic Raid Team that raids during 10:30pm-2:30am, M-F. Those are my hours of availability. Lost my raid team do to a schedule change, we were 3/10 M. Btag: KillerCars#1225.1
Arms Warrior (900 ilvl) LF Guild Hey. 900 ilvl arms warrior (51T) LF raiding guild that raids on weekdays. 7/7 H EN, 3/3 H ToV, 10/10H NH exp. Good hours would be around 7ish to 11/12ishh server time. Been playing arms warrior since forever and I follow up on all current content news, rotation, and abilities. Logs are posted below (https://www.warcraftlogs.com/rankings/character/67142/latest/). Numbers are very good, however, I also don't have DoS, which will boost it exponentially. Add me on bnet SubieDriftin#1487 if you are interested in having me as a raider :)1
Rogue / Ret / Lock LF Guild Hey, two friends of mine and I are looking for a new raiding guild. Our last one fell apart due to lack of uh...everything. We are looking to move to A52 cause we want to play on Horde once again. We are all 10/10 heroic experience and have raided mythic content a lot in the past and would like to find a guild that is in a similar spot. In turns of raid times, we're looking for something around 8-11 PM(no later than 11 EST). No Fridays or Saturdays - we're open for most other days. our equipped ilvls and traits are... Rogue = 902 54 traits Ret = 896 43 traits Lock = 891 44 traits Both the ret and lock are re-rolls from tanks so that's why their traits are not 54 but i mean...in 7.2 that's not going to matter that much Here is our armory Rogue: http://us.battle.net/wow/en/character/stormrage/Finch%C3%AEe/simple Ret: http://us.battle.net/wow/en/character/stormrage/B%C3%B6rnsinner/simple Lock: http://us.battle.net/wow/en/character/stormrage/Shadokouu/simple Let me know if you are interested in us! :D Thanks6
7/10M, [A] <Gods of the Arena> LF A FEW DPS ! <Gods of the Arena> is an Alliance Mythic raiding guild on the east coast PvE server Stormrage. We are an established raiding guild with the top performing players on our server. Our only requirement is that you excel at your class and have a good attitude. ************************************************************* Current Progression 7/10M Nighthold 9/10M EN/ToV ************************************************************* Raid days: Fri-Sat (10:30pm-2:00am EST). Optional activities on Monday and Sunday. Wednesdays are when we clear our farm content. ------------- Currently Recruiting: Melee DPS -Windwalker Monk (high) -Retribution Paladin (med) Ranged DPS -Elemental Shaman (high) -Frost Mage (med) Looking for all high quality players regardless if the class is listed or not. If you want a home for the rest of your WoW career we can meet those needs. If you think you can compete on a hardcore level, then we encourage you to contact us. -------------- About us: We are one of the many guilds looking to prove ourselves and compete for a top spot on the competitive server Stormrage. A stable guild with a solid infrastructure that has taken shape since Legion launch. Our raid team spots are based on merit, not favoritism. Players who work hard and constantly strive to improve will be given opportunity. We also have an active high level Mythic+ scene and are in construction of a pvp scene. What we can provide: ● A guild that will invest in you as much as you have invested in it. Your hard work and dedication will not be ignored. Go that extra mile and we will meet you there. ● A place where you will be regarded with respect and your voice will be heard. We appreciate your thoughts, ideas, and insight to strategies. ● PvP, Family, and Mythic+ rank. If you have friends/family that aren't active in the PvE raiding scene but are interested in other content that WoW provides What we desire: ● Players with a strong raiding background, a willingness to improve, succeed, and in-depth knowledge about their class. ● People looking to be part of a team for the long term. We are looking for players that want to spend the rest of their raiding career here. ● The drive to be the best there ever was at your class and spec ● We want each and every one of our members to enjoy raiding here with each other and we recruit like-minded individuals. ● Most importantly: New Memes Our recruitment information can be visited on our discord chat: (Work in progress) Contact info: <strong/> If you have any questions about anything, feel free to message any Gods of the Arena officer: Demotank, Divinelion (Casey#1547), Roycerx, Dirgedancer, and Alphøns (Motiv3#1599). Good luck on the search for your new home !!0
<Apocalypse Army> 3/10 M Late Night We are 3/10M Working towards Mythic Krosus and further Progression Raid Times 12AM to 3AM Server(EST) Tuesday,Wednesday,Sunday Most of the Core has been raiding together for years and have many Heroic(now Mythic) kills from back in the day. We have a pretty solid group right now that is progressing nicely. We'd like to add a few DPS and one healer into the fold. We will be willing to look at other classes too. This is just what we'd like to get. If you feel confident in your dps contact us. Xiazaki Wispurwind Randyrustrod Battletag: Cerebral#1634 (If none of us are on feel free to contact anyone else in the guild for more info,Most people should be able to help!!)2
[H]Area-52 <The Stardard Method> 5/10 H LFM "The Stardard Method" - newly established Horde guild on the Area 52 server. We are looking to recruit a handful of raiders who would be interested in pushing for the Cutting Edge achievement for every new tier of raiding. We recently transferred over from alliance and are new to the server. We hope to build a strong community of raiders where everyone can excel and push content. So far we have completed one of our raid cycles and the following raids, EN on normal and Heroic. TOV on normal. Normal NH. 5/10H NH. We are currently in need of both ranged and melee DPS, as well as a DPS with a maintained tank off spec. We are especially interested in the following classes. Death knight - Frost/UH Warlock - Any spec Priest - Shadow Pally - Ret Druid - Balance/Feral Shaman- Enhance/Ele We raid 3 days a week ( Wednesday, Thursday, Saturday ) for 3 hours at a time ( 9-12 EST ). We use a loot council system in which we prioritize everyone getting their 2-set then 4-set and the rest is rolling for BiS pieces and the council will distribute the gear based on the highest upgrades for the better of the team. The guild offers Cauldrons, and food at every raid. Pre pots and flasks will also be available as we are able to stock up on them. We also run a fair amount of M+ keys every week and do what we can to push as many keys as we can. All in all we like to have fun while chasing what we want and not letting others tell us how to play our game. We utilize discord for raiding communication as well as just to hang out with each other. Please feel free to reply to this thread or add the following B-net tags for more information. Shrui#1663 Dekappatater#1312 You can also visit our Guild website at the following- http://stardardmethod.shivtr.com/ Thank you for your time and consideration. Happy Gaming!1
H-US A-52 Purify 10/10H 1/10M LFM M Raid Team Purify is a raid focused guild that is currently progressing through Heroic and focused on moving quickly into Mythic. We are currently looking for more skilled players to augment our current raid roster to achieve these goals as efficiently as possible. We raid the normal 9 hour raid week, and as such are looking for serious minded players who understand that this is a time commitment. Loot System: We Use RC Loot Council And Gear Is Distributed Based on Attendance, BIS, Performance And Amount of work put in outside of regular raid times. =====Who We Want===== Fun people who excel at their class/spec and who bring knowledge of their class and raid mechanics. We expect you to come prepared to raid with Flasks, Potions and Food. We also expect you to put in time outside of raids. Raid Times: Tuesday 10:00PM-1:00AM (EST) Wednesday 10:00PM-1:00AM (EST) Thursday 10:00PM-1:00AM (EST) =====Recruitment Needs===== Tanks : Guardian Druid, Prot Paladin, Brewmaster Monk Healers : Resto Shaman OR Holy Priest DPS : All DPS Should have at least 40 traits in their MS weapon. Mage Shadow Priest Elemental Shaman Rogue Frost DK Monk Fury/Arms Warrior Havoc Demon Hunter All Exceptional DPS will be considered Contact Us: Officer-Bic#11211 Officer-Craps#11870 kbomb#19008
870 Resto Druid LF Guild Looking for raiding guild. Very coach-able been playing since BC off and on. If you would like to know more add Kl0wN1#16662
[H] <BlackHat Mafia> 10/10H LF DPS for M NH Hey everyone! <BlackHat Mafia> is a 2 night a week semi-casual raiding guild looking to fill our core 20 man raid team for Nighthold. We are currently looking for more DPS, preferably ranged. We are in need of a mage and a warlock but we're interested in talking to any class/spec. Players with a usable offspec is a plus! We currently raid Tuesday/Thursday from 8 PM EST to 12 AM EST and are 10/10 H as of now. We're a guild that started from a bunch of college friends who came back to WoW at the beginning of Legion. Our main goals are killing bosses and enjoying ourselves while doing it. We want to get into Mythic ASAP and our goal is to down M Gul'dan before ToS. We are a 'semi-casual' guild in that we only raid two nights a week but we expect everyone to take raid seriously and put their best foot forward every single night--we know how valuable everyone's time is and would rather not waste it. Our raid environment is one of the most, if not the most, important thing to us as a guild. We pride ourselves on the fact we don't really "rage" or yell at others and overall have chill raid nights. We are a fairly close-knit group and in general very laid back. It is, after all, a game and everyone should be prioritizing having fun while playing it. We want raiders who can reliably commit to our schedule and put in the work outside of raid to be ready with pots/flasks/food. We know real life comes first and other obligations or emergencies come up; all we ask is that you let us know as far ahead in advance as possible so we can plan around it, especially come mythic progression. While we are semi-casual, we want to progress and clear content so we want raiders who know their class and perform decently. We are more than happy to sit down and help improve people's individual performances as long as the effort is being put in from both sides. Our loot is distributed via loot council. As a whole we don't care too much about loot as individuals. As I said before, our goal is progress and loot is just a tool to get to that progress and that is a philosophy most of us agree with. We have done a loot council the whole expansion up to this point and have yet to have any complaints of fairness so I think we do a pretty good job. We as a council strive to be very transparent about loot distribution and will have no problem explaining how we made the decision. We are looking for more DPS to round out our core roster. We prefer ranged DPS but any good, well rounded player will be considered. If interested, please reply to this post or feel free to add me on BNET: JohnnyCage#1493 TL;DR: 10/10 H Tuesday/Thursday (8PM-12AM EST) Guild LF More DPS for Nighthold and beyond. Add JohnnyCage#1493 for more info/questions.3
LF Mythic Ready Disc Priest [H] <I N F E R N O> is looking for a high end heroic/mythic ready Disc priest to join our main raid. If interested please reach out my myself or another officer or apply on our website for fastest turnaround. We are 10/10 Heroic NH. Link to our main recruitment thread is below for reference. https://us.battle.net/forums/en/wow/topic/20752236325?page=1 http://inferno-area52.enjin.com/home Guild Master - Rudonkulous [ReDoNk56#1461] Guild Master - Sikowanalize [Flyingduck#1665] Officer- Deathsdefense [deathdefense#1563] Officer - Frankfutter [frankfutter#1767] Officer - Zuhrim [Suhrim#11667]12
Our Sweet Guild - 4/10M Open DPS recruitment! Raiding Tues/Thurs/Sun 9pm-12am EST, and are currently looking for any and all dps. Preferably ranged, but open to some melee Any questions or information are to Dare, or Dare#1202 Cheers20
Chemical Imbalance 5/10 H LFM 2x/week Chemical Imbalance is looking for more dps(ranged preferred) to start off ToS strong. We could also make room for a tank if you were so inclined. We are 5/10 Heroic NH, and kind of bummed about it. We lost quite a few players to personal/life situations over the past few months, and being a fairly tight guild, it hit us hard. Most of us have been raiding together since BC, so we all get along well. We don't tolerate drama(esp about loot) because...it is a game and ain't nobody got time for that. We raid Fri and Sat 9-12 server(Eastern). We expect raiders to know fights, have food and flask(although we usually have cauldrons), potions, and enchants. 870ish min ilvl. Bellatrix#1309 or Alchemy#17071
890 Holy Priest LF late night raiding guild. Hey everyone im a 890 Holy Priest who is trying to find a new home. Im currently on a low pop realm looking to move to Area 52 to raid. Ive done the whole hardcore raiding in the past and its just not for me anymore. Id like to find a guild with a good sense of humor and knows how to have fun raiding without making it a job. I have 10/10 N NH exp with 4/10 H NH experience. If anyone is interested you can add my battle tag and we can talk further. BakaBob#114217
[H] <Blood Eagle> Midday Heroic Raiding <Blood Eagle> is a midday raiding guild that is currently 5/10 H NH. We are reworking our raid team, and are looking to add some Healers and DPS in order to reignite our Heroic progression. We raid Wednesday and Friday from 11AM - 2PM EST. PVPers and casual players welcome. Respond here or contact LokiDalton#1867 or Trikan#1704 for more details.1
Sparkle Squad 5/10 H Sparkle Squad is in need of a Resto Shaman, Rogue, Balance druid, and a Warrior dps. We raid Fridays/Saturdays at 8 server and clear the older tiers anytime throughout the week, we also run keystones quite often. If interested in joining reply here or add Nic#1453 on battlenet and send a message, thanks.2
Returning Player Looking for a new home I started playing WoW again about a month ago and I figure that since it has been so long since I played (MoP), that it would be a good idea to start over and experience the Horde side of the game. To that end, I am currently leveling a character (just started and still under level 10) and I would like to find a guild that I could run dungeons and raids with once I get to max level and get geared. I have a few hours each night during the week that I could play (with being able to play more on the weekends). I also am open to leveling several different classes dependent on what is needed. I normally play healer but could play a different class if needed as well.3
[H] <Dishonored> 5/10M LFM <Dishonored> is a semi-hardcore mythic raiding guild on Area 52. We have 2 mythic raid teams, both earned Cutting Edge in Emerald Nightmare. Team Noodle is 4/10M, and is looking for a few skilled players to round out our roster. We use RCLootCouncil as our loot system, and Discord for voice. All raid consumables such as flasks, potions, food, and tomes are provided. We raid Tues/Thurs/Friday 9:30-1 EST. Currently recruiting: RDPS Warlock Hunter Boomkin Priest MDPS Rogue Death Knight Monk Warrior Paladin Demon Hunter We are willing to consider exceptional players regardless of class. Whisper Syreth, Caylyyn, Nakedstone, or Rippasta for more information. Syreth#115667
[H] <Negatory> 7/10M recruiting <Negatory> is a Horde raiding guild of 7+ years on Area 52 which raids current content. We are currently looking for a few members to fill in our raid roster. That said, please bare in mind that we are seeking a GUILDIE, not a raid robot. We are not interested in a player who will only appear online at raid time, we want active players who are social and mesh well with our existing group. HEALER: (NOTE: We are only looking for one healer) - Restoration Druid - Mistweaver Monk - Holy Priest - Discipline Priest DAMAGE: - Hunter - Warlock These are our higher priority classes/specs, however ALL exceptional DPS and healers should apply if interested. Raid-viable off-specs are highly desirable. Progression: Current: 7/10M 7/7M 2/3M T18: 13/13M with M Archimonde mounts farmed for the entire raid team Raid Schedule: Tuesday through Thursday: 8:30 pm - 11:30 pm EST What you can expect from us: - an active guild with plenty of social opportunities - a progression-minded raid team - guild repairs - some assistance with gear enchantments and flasks/potions - lady voices in the Discord What we expect from you: - knowledgeable in your class (all specs) - research fights in advance of new raids/encounters - willingness to make adjustments to playstyle upon request - gear properly gemmed/enchanted at all times - prepared with food, flasks, and potions - not a douche Please seek out Everly (Yummies#1193) or Snobby (Snobby#1357) in game for more information or apply at: http://negatory.shivtr.com3
Previous US 54th Feral Druid coming back Hello everyone, I'm just coming back to the game after over 2 years off. I was raiding with Freakshow as a Core Raider, while also being the primary Recruitment Officer when I was available over the past several years now, but they unfortunately don't have room for me anymore. Back in Siege of Org, I was the 54th ranked feral druid in the US (3rd on Area 52), with very high percentiles, and I was part of a guild that downed Garrosh before the patch. Siege was a time when I was very serious about raiding, so please use that to base your opinions. I would love to get back to that level of raiding again, and I am fully capable. (Also remember that Mythic didn't come around until the latter part of Siege) I am a bit under geared right now (Currently 886 Equipped), as I have only been back for about 1 month now. I really want to get back into Mythic raiding as fast as possible, although I know I still have some work to do in regards to my artifact. Here are some logs from my raiding history, with most recent posted first (My most recent logs are a bit sloppy due to not fully understanding the mechanics yet, so please don't read into them too much)- https://www.warcraftlogs.com/rankings/character/49200/latest#bracket=15 https://www.warcraftlogs.com/rankings/server/18/5#difficulty=4&class=Druid&spec=Feral I know some guilds have age requirements, so I'm about to turn 25. Please feel free to contact me if you need more details Green#136419
Gearing up - 872 BM Hunter LF Weekend Guild Hello! I am currently in the process of gearing up my BM hunter and will be looking to join a laid back weekend raiding guild soon. Times I am available are: Fridays - 8:00p - 11:00p EST Saturdays - 12:00p - 8:00p EST Sundays - 9:00a - 6:00p EST I have 2/7M, 3/3H, 10/10N and 10/10H experience on my main character. If you think I would be a fit for your guild please leave your spamarino below. =D Thank you!6
[H]<Winds of Winter>1/10M LF dps/heal /2 day Winds of Winter is currently looking for a few more players to fill out our roster for M raiding. We're a fun group, most of which have been playing together for quite a while. We meme around a lot, but get content cleared. All of our players currently are above the 75 % tile and we expect the same from you! We only raid 2 nights a week; Tuesday and Wednesday 8-11 server. We run several 15+ keys each week and have a solid number of players regularly on. We have been 1/10 M for a while now, as our roster isn't complete. Add me on battle net, send a whisper for more info. nwtwozero#1572 Currently we are looking for dps and a healer. Preference to.... - Ret pal - DK - DH - Shaman1
[H] Prot Pally LF Raiding Guild 891 Current progress: 10/10N and NH 5/10H - Mythic 16+ Have been raiding/tanking/healing/dps since Vanilla. Have several realm firsts. i'm available to raid at any time after 6pm eastern Mon-Friday. 40+ Artifact Talent Points if you have any questions, my b.tag is pokerbehind#13675
<Artifice> LFM for 7.2 Raiding Artifice is looking to rebuild after a hiatus back in HFC. Formerly a mythic raiding guild, we aim to build back up a roster capable of tackling mythic content. We are currently looking for mainly DPS and Healers of all specs and classes with an emphasis on: DPS: Mage - High Boomkin/Feral - Medium Spriest - High Demon Hunter - Medium Arms/Fury - Medium Warlock - Medium Healers: Holy Paladin Resto Druid MW Monk Resto Shaman Tanks: N/A If your class/role is not listed do not hesitate to get in touch. Our current raid days and times are: Tues/Thurs 9pm - 12am EST(st) The possibility of a third day being added on at the start of each tier is a possibility, but the availability of the group will be taken into account when and if that discussion comes up. If interested, feel free to get in touch with GM: Stripercop(JONATHAN#1760) or Realtylee(Tyzula#11580)0
Top 70 US Healer reroll to A-52 Hey thank for stopping by, as the title says I am looking for a reroll on A-52 for tomb progression. Due to certain real life issues I had to take off most of Nighthold so far and still wont be 100% available again until late April / Early May. Progression before I had to quit was 7/7 M - US 61st / 3/3M - US 67th (done with my alliance Mistweaver -http://us.battle.net/wow/en/character/kelthuzad/Celerie/simple 3/10M (before i had to quit) - US 66 (Done with my alliance priest -http://us.battle.net/wow/en/character/kelthuzad/Celeriy/simple The reason I am rerolling is due to after myself taking a break many key players from my old guild quit as well due to the burnout of Legion. With that being the case my old guild is still going strong but isn't the guild I was apart of and I feel very little connection to it now. Logs from before I took my break are available upon request. Looking for similar or slightly lesser progression minded people for Tomb release. This characters gear / AP lvl will be up to par by tomb or my alliance Mistweaver will be transfered and faction changed upon Tomb release. For any further questions leave a remark below or add me on Bnet: Celerie#117262
<Lowest Expectations> 10/10H Nighthold <Lowest Expectations> Is currently recruiting for our mythic raid team! Come check us out on our website! See us stream and check out our boss fights to see if we are a team you want to play with! https://lowestexpectations.gamerlaunch.com For our raid team food, flasks, enchants, and repairs are provided. What we expect in return is to be on time to raid, participate in discord with raiders and casuals, and run mythic plus groups in house to help casual members. Not into raiding? That's okay too! We are always looking for active quality players to join us for mythic plus dungeons and world questing. We also have a casual raids including H-EN, H-ToV, and N-NH scheduled throughout the week that ALL guild members are invited to attend. Core Group Information: Raid Times: Tuesday 9PM-12PM EST Thursday 9PM-12PM EST Current Needs: RANGE DPS MDPS Requirements: Looking for 880 item level or higher. Have 50 (+) traits in your artifact. Minimum DPS 500k (+) single target. Have a viable OS. Current Progression: 10/10 Heroic Nighthold 10/10 Normal Nighthold Previous Progression: 2/3 Heroic ToV 4/7M Emerald Nightmare Tyråç (Zyn#1113): Guild Master Inxy (Reahli#1781): Recruitment21
♠<TFC>♠ 2 day 10pm EST 4/10M NH LF DPS (Recruitment updated 3/24/17) Raiding Schedule: Tue/Thu 10PM-1AM EST (Server time), Wednesday 10PM-1AM EST for Heroic night! (Not mandatory) About Us: Our recruitment video(watch it for the lulz): https://youtu.be/DV1yYivjy64 <Titanforged Chromosomes> 4/10M NH with Spellblade @ 9%, 7/7M EN, 2/3M ToV on Area 52 is recruiting! We offer you a stable raiding atmosphere led by experienced raiders that have been around since BC, with the guild itself established over 6 years ago. Since we only raid 2 days a week, we aim to progress as much as possible while still having a great time, taking care of business, and working towards our goals. We aim to do as much in 2 nights as other guilds do in 3. We looking for players that reflect that type of skill level and goal. Many of our players come from top 100 guilds that don't want to/have the time to raid 4-5 days a week. Just because we only raid 2 days a week doesn't mean that we aren't active the rest of the week, though. We push +20 keys and do a lot of alt runs. We encourage you to hop in Discord and hang out with us because we are a very social and active guild. Perks: We provide flasks for everyone that raids as well as 1000g/day guild bank repairs. Voice: Discord. You are required to have a working Mic/Headset. Loot System: RC Loot Council. New members are not excluded from receiving loot. Recruitment Recent logs are required to apply. Must be minimum 10/10H with a minimum equipped ilvl of 895. If you are an exceptional player and your class/spec isn't listed below, please still contact us! R DPS: Marksman Hunter: High Beast Mastery Hunter: Medium Elemental Shaman: High Shadow Priest: Medium Boomkin: Low Warlock: Low Mage: Low Melee DPS: Frost DK: Low Feral Druid: Low Assassination Rogue: Medium Arms/Fury Warrior: Closed Ret Paladin: High Windwalker Monk: Low Survival Hunter: Impress us Heals: (All healers that apply for this specific spot must have a viable dps offspec). Holy Priest: Medium Disc Priest: Medium Resto Druid: Medium Sign up @ http://tfc-guild.enjin.com/ and fill out the app. Contact me @Hajari#1287 for any further questions! If I'm not online, contact Kicker03#1475.370
897 Fire mage looking for raiding guild My guild recently split up after killing heroic guldan. I am looking for a fun and progressive guild looking to get into mythic nighthold. My preferred nights of raiding would be tues/thurs from 9pm-1amest but i can probably be flexible. I have a 897 firemage with 51 traits and a 870 affliction lock with about 40 traits. Let me know if your guild sounds like a good fit. Update...Still looking for a guild and it seems like most mythic guilds raid times I cannot...I am thinking heroic only may be better for my schedule...let me know if this works. Thanks!12
900 Havoc DH LF Mythic Raiding Guild I'm currently looking for a mythic raiding guild due to a change in my work schedule. I was 7/7M Emerald Nightmare(cutting edge) and currently 3/10M Nighthold. I'm looking for a guild that raids between these hours: 4:00PM-9:30PM EST 3:00PM-8:30PM CST 1:00PM-6:30PM PST I have had 54 traits since the 7.1.5 patch and currently have BIS legendary AoTG. I'd prefer you to contact me in game Frostys#1524, I'm willing to transfer as needed.1
LF late evening raiding guild Played since vanilla on multiple characters, healed since WotLK. I'm looking for a guild that raids around 10pm CST to 1-2am CST. Active community is a plus, I take an active role within my guilds, running dungeons, gearing alts etc. If a trial is necessary by all means I will come based upon that. Bnet tag: Mercy#146321
897 equip Bear lf guild (H) looking for a casual setting but progression minded group of adults for raiding. Looking for Tuesday and Thursday 9pm-12 server time. currently 9/10H exp with guldan at sub 10%.... would like to push into mythic as well i also have 3 other tank alts that i play randomly and usually do a +15 weekly if interested or have more questions hit me up here or bnet bridge#1686 or discord bridge#7045 (best way)6
Just Us 3/10M 10/10H Recruiting <Just Us> on Area 52 Horde is looking for more core members for Mythic Nighthold. We are a laid back guild, however during progression we expect our members to be on point, on time, and have the required addons/consumables. We us Discord for voice chat and RC Loot Council for our loot system. Raid times: Tuesday/Wednesday/Thursday/ 7 to 10 est. Current Progression: 10/10H 3/10M The main goal of <Just Us> is to push mythic content and to do that we need to expand our core group with some of the below specs. Healer: Holy Priest(Shadow Off-spec) Resto Druid(Boomkin Off-spec) Holy Paladin DPS: Warrior Rogue Warlock Ele Shaman Shadow Priest Boomkin Hunter These specs along with any exeptional players are welcomed to contact our officers listed below If you are interested in joining, here's how you can contact us! Rothandria - Roth#1580 Btag/Roth#9553 on Discord Zalindae - Zalindae#1170 Btag/Zali#3654 on Discord Saltso - Waronne#1951 Btag9
Is it possible? progression raiding AND fun? Is it possible to be progression oriented AND have fun? We believe it is! We pride ourselves on being good while having fun. If you are looking for completely silent raids with minimal callouts and interaction, then this might not be the raid team for you. If you can parse high while doing shots and singing Disney songs into discord while doing Heroic Gul'dan, then I think this just might be the guild for you. We are a progression raiding guild that is 9/10H with sights on being a mythic raiding team. We recently lost the bulk of our raiding team and are rebuilding with an experienced team of officers and raid leaders that are passionate about progression. While we only raid 6 hours a week, we still managed to be 9/10H despite having continually fluctuating raid members. We would like to get a consistent team for a mythic raid team within the next 4-6 months. I'm aware of the length timeline, but we are looking for people that will be willing to commit to a genuine progression team. It would be nice to jump into mythics sooner, but I have been doing this for some time and I understand the reality of building a solid team. We are currently raiding H NH at those times with whatever members we have available to us, so those who are interested in joining should be prepared to jump right into the routine with us. I am the GM so please feel free to contact me through this forum or through my BNET. Battle Tag: Eranith#17621
(H) RK LF All 9/10H Mon-Weds 8:30-11:30EST We pride ourselves on being good while having fun. If you are looking for completely silent raids with minimal callouts and interaction, then this might not be the raid team for you. If you can parse high while doing shots and singing Disney songs into discord while doing Heroic Gul'dan, then I think this just might be the guild for you. We are a progression raiding guild that is 9/10H with sights on being a mythic raiding team. We recently lost the bulk of our raiding team and are rebuilding with an experienced team of officers and raid leaders that are passionate about progression. While we only raid 6 hours a week, we still managed to be 9/10H despite having continually fluctuating raid members. We would like to get a consistent team for a mythic raid team within the next 4-6 months. I'm aware of the length timeline, but we are looking for people that will be willing to commit to a genuine progression team. It would be nice to jump into mythics sooner, but I have been doing this for some time and I understand the reality of building a solid team. We are currently raiding H NH at those times with whatever members we have available to us, so those who are interested in joining should be prepared to jump right into the routine with us. Our current roster consists of the following. Tanks 1/2 Healers 2/5 DPS 4/13 Once again, our raid times are: Mon-Weds 8:30-11:30EST I am the GM so please feel free to contact me through this forum or through our guild discord or my BNET. Battle Tag: Eranith#1762 Red Knights Discord: https://discord.gg/nyb5XuY2
[H] <Titan Group> 9/10H LFM <Titan Group> is a very laid back, but driven guild looking for more members to fill our raid roster. While we fit in the "casual" raiding style, we get our work done and looking forward to progress into mythic NH. We also do mythic 15+ runs each week. We are currently looking for ANY and ALL specs. However, ranged DPS would be especially nice. We only ask that you are 880+ with 45 traits in your main spec and are looking to have FUN in the Azeroth! We usually raid on Tuesdays, but are flexible depending on IRL things, but expect at least one day per week (usually 9PM EST- 12AM EST) We provide all food, flasks, and tomes. Tl;dr Looking for a drama-free, laid back, casual raiding guild that knows mechanics while also having fun into the late hours of the night! Message Lulixa Bnet - Lulithiel#17601
LF Active Raiding/Mythic+ Guild Hello, i'm currently rerolling to my warrior so im still in the gearing process at ilvl 873 right now, however i have experience with 10/10 N and 6/10 H NH, prefer to have a late raid time around 11-midnight, any day of the week, or early morning 8am-11am, however i can usually work around times, in addition looking for a guild that has an active player base and who run mythic+s a lot, and just has fun, in addition to transmog/achieve runs. Active discord is a big plus too :D! Message me in game or on here, Alt code is alt-129 for the u. Also can add me on Btag if thats easier Lokarash#1769.0
LF Late Night Guild Hello all. I'm looking for a late night raiding guild. Something between 10pm PST and 4am PST ( which would be 1am and 7 am EST). I know its an EST based server, but I also know that people from every where play here. I started playing during vanilla on Proudmoore Horde. That faction was always low population, but really died when Aussies got a free realm transfer. After all these years, server population is one of the most important things to me. Area 52 has one of the highest for a PvE realm. I have a 900 resto/balance main, also a 890 arcane mage and an 885 unholy dk, who are all decently set up. With my druid, I'm 10/10 H , 2 /10 M. I'd like to find a guild that at least is 9/10 H working on Guldan, with solid plans of doing Mythic raids soon. But one that's 1/10M would be ideal, so we can progress together. I'm considering transferring, just wondering if the times for a raid are even a thing here. I'm currently on a PST server, and even here, its hard to find a late night guild. But Area 52 has like twice the population, so I figured I'd ask. Thanks for reading, and thanks for your time.2
<Sacred Lotus> 10/10H Looking for Raiders! <Sacred Lotus - Area 52> We are a guild that previously transferred from a really low populated realm mannoroth on alliance side. Long story short, our guild master and his wife ended up leaving the game due to real world issues. Our main core raiders are still here looking to continue and we are looking for some dedicated raiders to continue onward into mythic content. Our guild consists of 9 core members at the moment and we are looking to rebuild our mythic team. We are currently 10/10 in heroic nighthold and we are looking to go straight into mythic once we recruit a few more players. Looking for 2 healers, preferably a Mistweaver monk or a Holy priest. We are also recruiting dps both melee and ranged. (exceptional players are always welcome to try). You can send me an in game whisper or mail and I will get back to you as soon as I can or one of our officers for more details. Our raid times are as follows. Monday 8 to 11 PM Eastern time. Tuesday 8 to 11 PM Eastern time. Thursday 8 to 11 PM Eastern time. You are expected to be on 15 minutes prior. (Joebusyguns, Sassius, Faulks, or Wolfladylove)3
[H] Mistweaver LH Mythic Raiding Guild Solid healer LF a mythic progression guild on A52. 54/54/50 traits, bis legendaries , and healing gear sets for raiding and for mythic+ dungeons. Currently the #2 monk on A52 in mythic+ score on wowprogress. LF a guild that either raids weekday evenings (except Wednesday unless it's after 10 pm), or weekend mornings. I enjoy optimizing my character and learning how to play my class better during different encounters. High raid attendance, and punctual. I research fights before raids and bring all required consumables. Hope to hear from you! Btag: Shiv#16591
<Michigan Mafia> Looking for more Looking to join a hardcore raiding guild? Looking to be gkicked for being a scrub dps or a fail healer/tank? Want to have bragging rights of being the top guild on the server? Well thats not us. We are a pretty laid back bunch looking to expand our roster of active players. All types of players are welcome. Not 110 yet? No problem. Not at 54 traits yet? No problem. Still havent unlocked Arcway/CoS? No problem. Raid Times: SAT-SUN 6pm - ? Raid Status: Cleared normal EN and first boss in NH. Trying to find our place on where we should be as its a work in progress. Sometime during the week we try to fit in normal EN for gearing as well as doing mythic and mythic+ groups. Alt runs can be pretty fun as well. If your looking for a new home in a relaxed and friendly guild, check us out. Head on over to our site and apply today! http://mm.wowhordes.com0
849 Fury Warrior LF Guild I just leveled this toon and love this class so much I'm going to make it my main; I'm looking for a guild filled with cool people to play with. I am willing to push progression once I get geared.1
<Insømnia> AFTERNOON Heroic/Mythic Raiding [H] <Insømnia> (Area 52) is currently recruiting for our Heroic/Mythic NH progression teams! Do you find your days filled with time that you don't want to spend pugging in group finder? Then <Insømnia> is the group for you. The afternoon EST slot is what we're here to make great. Our team is currently 10/10H NH, with our sights set on becoming the premier high-end content guild for the daytime slot. We raid from 3pm - 6pm EST, Tues/Wed/Thurs. We are splitting into 2 teams, so recruitment needs are a bit broad. Current needs: Tanks: Pref. druid, but all classes considered Healers: Need heals (bonus points for DPS offspec), all classes considered Melee DPS: Not high priority but open to exceptional players Ranged DPS: High priority, all classes considered While we expect you to transfer your character if we bring you on permanently, cross-realm trials are absolutely fine. Area 52 is an active, highly populated server. For more information, please contact myself, Claie, at my Battle.net tag: Raere#1131.6
[H] Untold (7/7M+3/10M) Reforming Recruitment Information: Our team was previously 7/7M EN (Cutting Edge) + 3/10M (10/10H) NH. We are looking to rebuild our roster. Immediate positions are available. We are looking for players that: -Have exceptional attendance -Are competitive (logs) -Team players -Can accept constructive criticism Raid Times: Tues, Wed, Thurs :: 10:30PM - 1:30AM Server Time(EST) We start invites at 10:15pm Server Time(EST) and expect everyone to be on at that time. *If you cannot make these days/times regularly, you can still apply for a flex position or as a casual member. Currently Recruiting: *You can message any of our guild officers in-game to apply *Strong off-spec is a plus iLvL Requirement: 890iLvL Recruitment Priority: DPS (Ranged>Melee) *We are looking for players that want to be competitive for a spot on our core team. -- RANGED DPS -- -Elemental Shaman Very High -Shadow Priest Very High -Balance Druid Very High -Warlock Very High -Mage High -- MELEE DPS -- -Retribution Paladin Very High -Warrior High -Demon Hunter High -Enhancement Shaman Medium -- Heals -- -Restoration Druid w/ DPS offspec Very High -Restoration Shaman w/ DPS offspec Very High *Currently in need of a strong healer with a strong DPS offspec. -- Tanks -- -Brewmaster Monk Very High -Guardian Druid Very High -Protection Warrior Medium -Protection Paladin Medium *Any exceptional players are welcome to apply. If you can perform, we are interested. Addons/Requirements: We require the following addons: -DBM or Bigwigs -EPGP Lootmaster -Angry Assignments -Exorsus Raid Tools Voice: -Discord Guild Information: We started out as a tight knit 10m guild in MoP. We cleared Heroic SoO followed by Mythic SoO when the patch hit. WoD was a difficult expansion for us as a guild. We transitioned from 10m to 20m in order to raid the most challenging content. Through hardships, we strengthened our core and maintained progression. At the end of the expansion, we were able to solidify our spot in the top 15 of the server (13/13M). We've all played WoW for a long time and know our classes well and we do expect the same from any applicant. :) We take our raiding seriously, but we like to have a good time while doing it. A lot of us enjoy playing other games together as well; including but not limited to Diablo III, Hearthstone, Overwatch, etc. etc. Please message me or any of the other officers in game with these BattleTags to apply: Kaethur09#1857 :: (Guild Master) Thanks for your time reading the post and we look forward to hearing from you. :)44
Looking for names Below is a list of names that I'm currently searching for. If you have ownership of said name please contact me and we can talk GOLD. I'm willing to pay a hefty price for certain ones. I must resort to this because Blizzard no longer releases inactive names. Names: acid, asteroid, attack, assault, aura, ban, bass, black, blind, bounty, BUNNY, classic, dab, death, decent, decoy, defeat, duck, easy, eclipse, electric, enormous, epic, ether, faith, furious, gorgeous, green, happy, hate, heal, hidden, hit, honor, lag, lame, laser, light, liquid, lose, loser, magic, medic, monk, moon, nice, nightmare, power, rampage, rare, rat, rave, raw, respect, roll, sick, sin, smoke, spirit, star, sun, taunt, theory, thunder, toxic, trance, trap, trust, vehicle, weapon, weird, white, win, winter, wise, zen. Feel free to contact me through this forum post or in game Rat#115085
[H] <Relic>7/10H LF Rogue/Lock 4 H/M NH <Relic> is a semi-hardcore guild whose core is comprised of old-school raiders who may not have the time to raid as much as they used to due to real life commitments! We love to have fun and not take ourselves seriously, but still work hard and progress as a team. We are a very active guild with members all over the world, so there's always someone online! We'll join you for old content runs to get transmog gear, work on fun achievements, kick butt in Mythic+, or do whatever your heart desires. <Relic> raids once a week on Sundays, 8pm to midnight EST (Area 52 Server Time) with an optional day on Wednesday to help gear newer players or gear alts (also at 8 pm EST). Our current progression is 7/10H NH, 10/10 N NH, 7/7 H EN and 3/3 H ToV with very minimal pulls. We are ready to head into Mythics, we just need you.. We are currently looking for assorted DPS including but not limited to: Rogue and Warlock. However, we'll trial anyone who's interested! If this sounds like fun to you, please feel free to contact one of the following: - Kaerie on Area 52 or add Skyfire#1480 to Battle.net - Jinto on Area 52 or Jinto#1580 to Battle.net - Svenofnine on Area 52 or Svenofnine#1383 to Battle.net Thank you, and have an amazing day!79
Tues/Thur 9-12 Fresh guild mix of friends geared and not. Looking for folks at least 870. Guild went 9/10 normal on its first night tuesday. Will be finishing normal and starting heroic Thursday. Feel free to add my battle tag jonathan#1760 or in game whisper stripercop. Guild plans on gathering for the roaster and farming heroic. 7.2 will be a harder push with a plan for mythic content once undergeared players are caught up.0
887 RET LF Raiding Guild I'm ideally looking for something Fri/Sat night. I'm pretty solid, almost never miss raid and generally look into fights before there have been pulls made. Also, I unfortunately can't be part of guilds called things like "Fruit Punch Society" no offense to fruit punch!6
<RotLA> 10/10H Tues/Thurs 7-1030 <Raiders of the Lost Art> Looking for more core raiders to fill out mythic team for immediate progression. Need approximately three to five reliable raiders who are commonly minded and dedicated. Bunch of friends who run a relaxed but dedicated raid environment. No drama, adults only; looking for new friends who fit in and can run with us. 10/10H farm. Tues/Thurs 7-1030 EST. Ideally ranged DPS, other exceptional players welcome. We are heavy on plate / pally. Exceptional hunter, monk, mage, shaman DPS would be great. Mythic + and casual players also welcome.0
WTS M+15/ Nightbane/ Heroic Guldan Straight Cash Homie is finally back!!! once again offering sales runs, this time for your Legion content of choice! Hello and welcome to SCH Carry group. We are a group of friends who have been running and selling Challenge mode (Mythic +) carries since MoP. We are a very experienced group of friends who are ready to help you get your achievements, mounts, or loot. Nighthold progressions has finally begun, we are spread out among 3 different top tier guilds. scheduling will most likely be before raids, or sometimes late night after raids if that works better for you. At least until Nighthold progressions is over. 10/10 Full Heroic Nighthold Personal loot - 600k Heroic Gul'dan Personal loot - 300k M+ dungeon prices are listed dungeon by dungeon. If you purchase more than one at a time, discounts will be given. M+ Prices: M+ 7-9 - 75k M+ 10 - 150k M+ 12 - 200k M+ 15 - 1 mil Come get your in time +15 dungeon achievement completed today! We will provide the key for any dungeon you decided to buy. No need to try and pug to boost your key, we got you covered. Karazhan Prices: Karazhan - 150k Nightbane - 300k Karazhan w/ Nightbane - 350k If you'd like to book a run, or have any questions, feel free to add me: TMac#1531 If you are on a different server and would still like to book a run. You're in luck SCH is on multiple different server clusters, and we accept gold on just about any server. most of us have alts spread everywhere. As a final disclaimer WE DO NOT SELL FOR REAL MONEY, we will NEVER ask you to "get on skype to talk about it" or pressure you into pursuing "alternative payment methods". We do these sales to fuel our progression, so every run you book with us goes towards killing new bosses.1
< E Pluribus Unum > recruiting WE are currently rebuilding an old guild on server and looking for raiders to fill out ranks, mainly dps at this time. WE will start in normal nh so people can get used to each other before we push and put any strain on the group. We also accept non raiders and casual players as well. We only require people to be 18+ yrs old and able to function in a guild / group enviroment. For more Info reply to post or hit me up in game.7
2 Friends LF a Raiding Guild EDIT: Thank you everyone for your interest, but my friend and I have found a raiding home. I apologize for getting back to this post so late. I talked to a few of you and I appreciate everyone's interest in taking both of us in but we have found a home so I am now going to try and close this post.22


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text: Season 3 Category page 564pages on this wiki Add New Page Edit History Talk0 Sorting type: Most visited Most visited Alphabetical | This category lists every single episode from Season 3. Pages in category "Season 3" The League of Villains (episode) Lady Sings the News The N-Men The Tomorrow Boys Attack of the Twonkies Stranded Lights! Camera! Danger! My Big Fat Spy Wedding Crouching Jimmy, Hidden Sheen The Trouble with Clones Jimmy Goes to College King of Mars Who's Your Mommy One of Us Who Framed Jimmy Neutron? Fundemonium Back 1 2 Next Retrieved from "http://jimmyneutron.wikia.com/wiki/Category:Season_3?oldid=36541" Ad blocker interference detected! Wikia is a free-to-use site that makes money from advertising. We have a modified experience for viewers using ad blockers Wikia is not accessible if you've made further modifications. Remove the custom ad blocker rule(s) and the page will load as expected. Categories: Episodes


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UPDATE: Pamela Hupp Granted Change of VenueLawyers for accused killer Pamela Hupp go to court this morning to argue for a change of venue in her highly-publicized case.
Prosecutor: Hupp Too Well Known Nationwide to Move TrialAccused killer Pamela Hupp wants her murder trial moved out of St. Charles county, because of heavy pre-trial publicity.
St. Louis Jury to Hear Another Baby Powder CaseA Tennessee woman, 56-year-old Nora Daniels, will testify she used Johnson and Johnson talcum powder for 35 years before she was diagnosed with ovarian cancer.
Nephew Allegedly Kills His Uncle, Burns His Body After ArgumentThe argument between family members began when the nephew was driving his 4-wheeler in his uncles driveway and making ruts.
Man Accused in Second Murder Arrested After Woman's Body Found in LandfillThe suspect, 51-year-old Paulren Stepter, was charged Wednesday with first-degree murder in the death of 23-year-old Brandy Morrison.
UPDATED: Trial Date Set for Maplewood's 'Weed Lady'Alice Hezel's attorney hopes the trial settles the issue once and for all.
Extradition Hearing for Alleged Granite City Kidnapper TodayChristopher Derleth is scheduled to appear before a judge in Charleston, West Virginia for his extradition hearing.
Jurors To Be Selected in Baby Powder TrialA pool of 50 jurors picked last week will be whittled today down to 12 plus alternates.
New Trial for Reginald Clemons Pushed Back 6 MonthsClemons was one of four men convicted in the 1991 deaths of 20-year-old Julie Kerry and her 19-year-old sister, Robin.
Missouri Appeals Court Hears Penny Hubbard CaseThe Appeals Court has to decide before Friday.
Final Testimony Wednesday in Trial of Missouri Clergy ProtestersThose protesters, sometimes called the Medicaid 23, each face up to $500 in fines and up to six months behind bars.
Trial for Missouri Clergy Who Protested in Capitol BeginsMore than 20 clergy members dressed in religious garb and bearing crosses packed in a crowded Missouri courtroom Tuesday as their trial began.