A Newly Developed Chemically Defined Serum-Free Medium Suitable for Human Primary Keratinocyte Culture and Tissue Engineering Applications.

In our experience, keratinocytes cultured in feeder-free conditions and in commercially available defined and serum-free media cannot be as efficiently massively expanded as their counterparts grown in conventional bovine serum-containing medium, nor can they properly form a stratified epidermis in a skin substitute model. We thus tested a new chemically defined serum-free medium, which we developed for massive human primary keratinocyte expansion and skin substitute production. Our medium, named Surge Serum-Free Medium (Surge SFM), was developed to be used alongside a feeder layer. It supports the growth of keratinocytes freshly isolated from a skin biopsy and cryopreserved primary keratinocytes in cultured monolayers over multiple passages. We also show that keratin-19-positive epithelial stem cells are retained through serial passaging in Surge SFM cultures. Transcriptomic analyses suggest that gene expression is similar between keratinocytes cultured with either Surge SFM or the conventional serum-containing medium. Additionally, Surge SFM can be used to produce bilayered self-assembled skin substitutes histologically similar to those produced using serum-containing medium. Furthermore, these substitutes were grafted onto athymic mice and persisted for up to six months. In conclusion, our new chemically defined serum-free keratinocyte culture medium shows great promise for basic research and clinical applications.

Tie-Over Bolster Pressure Dressing Improves Outcomes of Skin Substitutes Xenografts on Athymic Mice.

The efficacy of skin substitutes is established for the treatment of burn injuries, but its use is not limited to this condition. This technology has the potential to improve the treatment of various conditions by offering highly advanced and personalized treatments. In vivo studies are challenging but essential to move to clinical use in humans. Mice are the most widely used species in preclinical studies, but the main drawback of this model is the limited surface area of the graft in long-term transplantation studies caused by the displacement and the contraction of the graft. We improved the conventional surgical procedures by stabilizing the chamber covering the graft with intramuscular sutures and by adding a tie-over bolster dressing. The current study was therefore performed to compare outcomes of skin grafts between the conventional and optimized skin graft model. Human self-assembled skin substitutes (SASSs) were prepared and grafted to athymic mice either by the conventional method or by the new grafting method. Graft healing and complications were assessed using digital photographs on postoperative days 7, 14, and 21. Similar structure and organization were observed by histological staining. The new grafting method reduced medium and large displacement events by 1.26-fold and medium and large contraction events by 1.8-fold, leading to a 1.6-fold increase in graft surface area compared to skin substitutes grafted with the usual method. This innovation ensures better reproducibility and consistency of skin substitute transplants on mice.

Frozen Elephant Trunk for Aortic Arch Reconstruction is Associated with Reduced Mortality as Compared to Conventional Techniques.

To examine the perioperative outcomes following aortic arch repair using frozen elephant trunk (FET) vs conventional elephant trunk (ET) techniques. Between 2002 and 2018, 390 patients underwent aortic repair with elephant trunk reconstruction at 9 centers: 172 patients received a FET (mean age: 65+/-13 years, 30% female, 37% aortic dissection) and 218 patients received an ET (mean age: 63+/-13 years, 37% female, 43% aortic dissection). Outcomes of interest included in-hospital mortality; stroke; and spinal cord injury (SCI). In-hospital mortality rate was 11% (n = 43) overall, 9% (n = 15) for FET and 13% (n = 28) for ET. Post-operative stroke occurred in 13% (n = 49) overall, 13% (n = 22) for FET and 12% (n = 27) for ET. The rate of post-operative SCI was 3% (n = 13) overall, 5.0% (n = 9) for FET and 2.0% (n = 4) for ET. When compared to ET, the propensity score analysis confirmed FET to be associated with lower mortality (adjusted risk difference -7.0% (95% CI -13.0 to -1.0), P = 0.02). There was no significant difference in the propensity score-adjusted risk difference for stroke between FET and ET (-0.7%, 95% CI -7.4% to 6.1%, P = 0.85), nor for SCI (3.3%, 95% CI -0.4% to 7.0%, P = 0.085) On multivariable analysis, FET was associated with lower odds of mortality (OR 0.44, 95% CI 0.21-0.95, P = 0.04), and had similar odds of stroke (OR 0.83, 95% CI 0.41-1.70, P = 0.62) and SCI (OR 2.83, 95% CI 0.83-9.60, P = 0.1). FET repair is associated with lower in-hospital mortality as compared to conventional ET, and results in similar risk of stroke and spinal cord injury. Further investigation is warranted.

Ticagrelor use and practice patterns among Canadian cardiac surgeons.

BACKGROUND AND AIM: The P2Y12 platelet receptor inhibitor ticagrelor is widely used in patients following acute coronary syndromes or in those who have received coronary stents. Bentracimab is a monoclonal antibody-based reversal agent that is being formally evaluated in a Phase 3 clinical trial. Here, we probe the knowledge, attitudes, and practice patterns of cardiac surgeons regarding their perioperative management of ticagrelor and potential application of a ticagrelor reversal agent. METHODS: A questionnaire was developed by a working group of cardiac surgeons to inquire into participants' practices and beliefs regarding ticagrelor and disseminated to practicing, Canadian-trained cardiac surgeons. RESULTS: A total of 70 Canadian-trained cardiac surgeons participated. Bleeding risk was identified as the most significant consideration when surgically revascularizing ticagrelor-treated patients (90%). There is variability in the duration of withholding ticagrelor before coronary artery bypass graft procedure in a stable patient; 44.3% wait 3 days and 32.9% wait 4 days or longer. Currently, 15.7% of cardiac surgeons prophylactically give platelet transfusions and fresh frozen plasma intraoperatively following protamine infusion in patients who have recently received ticagrelor. Interestingly, 47.1% of surveyed surgeons were aware of a reversal agent for ticagrelor, 91.4% of cardiac surgeons would consider utilizing a ticagrelor reversal agent if available, and 51.4% acknowledged that the introduction of such an agent would be a major advance in clinical practice. CONCLUSIONS: The present survey identified ticagrelor-related bleeding as a major concern for cardiac surgeons. Surgeons recognized the significant unmet need that a ticagrelor reversal agent would address.

Does adding an aortic root replacement or sinus repair during arch repair increase postoperative mortality? Evidence from the Canadian Thoracic Aortic Collaborative.

OBJECTIVES: The aim of this study was to examine the effect of the addition of an aortic root replacement or sinus repair on mortality and morbidity during aortic arch repair. METHODS: A total of 2472 patients underwent proximal or total aortic arch repair with hypothermic circulatory arrest between 2002 and 2018 at 12 centres. Multivariable logistic regressions (MV) and propensity score (PS) with inverse probability of treatment weighting (IPTW) analyses were performed. RESULTS: A total of 1099 (44.5%) patients had additional aortic root replacement (n = 934) or sinus repair (n = 165). Those with aortic root interventions were younger (61 ± 13 vs 64 ± 13 years, P < 0.001) and had less females (23% vs 35%, P < 0.001), less dissection (31% vs 36%, P = 0.004), less urgent cases (35% vs 39%, P = 0.047), more connective tissue disease (7% vs 3%, P < 0.001) and less total arch replacements (14% vs 22%, P < 0.001). On adjusted analyses, the addition of aortic root procedure was associated with increased mortality [MV: odds ratio (OR) 1.41, 95% confidence interval (CI) 1.03-1.92; PS-IPTW: risk increased by 3.7%, 95% CI 1.2-6.3%, P = 0.004]. Reoperation for bleeding was also increased with the addition of aortic root intervention (MV: OR 1.48, 95% 1.10-1.99; PS-IPTW: risk increased by 3.2%, 95% CI 0.8-5.6%, P = 0.009). The risks of stroke and dialysis-dependent renal failure were similar. When looking only at non-elective cases, the increased risk of mortality was more pronounced (MV: OR 1.60, 95% CI 1.11-2.32, P = 0.013; PS-IPTW: risk increased by 6.8%, 95 CI 1.7-11.8%, P = 0.008, and a number need to harm of 15 patients to cause 1 additional death). CONCLUSIONS: The addition of aortic root replacement or sinus repair during proximal or total aortic arch repair seems to increase postoperative mortality only in non-elective cases.

Infective Endocarditis Secondary to Injection Drug Use: A Survey of Canadian Cardiac Surgeons.

BACKGROUND: Injection drug use-associated infective endocarditis (IDU-IE) is a growing epidemic. The objective of this survey was to identify the beliefs and practice patterns of Canadian cardiac surgeons regarding surgical management of IDU-IE. METHODS: A 30-question survey was developed by a working group and distributed to all practicing adult cardiac surgeons in Canada. Data were analyzed using descriptive statistics. RESULTS: Of 146 surgeons, 94 completed the survey (64%). Half of surgeons (49%) would be less likely to operate on patients with IE if associated with IDU. In the case of prosthetic valve IE owing to continued IDU, 36% were willing to reoperate once and 14% were willing to reoperate twice or more. Most surgeons required commitments from patients before surgery (73%), and most referred patients to addiction services (81%). Some surgeons would offer a Ross procedure (10%) or homograft (8%) for aortic valve IE, and 47% would consider temporary mechanical circulatory support. Whereas only 17% of surgeons worked at an institution with an endocarditis team, 71% agreed that there was a need for one at each institution. Most surgeons supported the development of IDU-IE-specific guidelines (80%). CONCLUSIONS: Practice patterns and surgical management of IDU-IE vary considerably across Canada. Areas of clinical unmet needs include the development of a formal addiction services referral protocol for patients, the development of an interdisciplinary endocarditis team, as well as the creation of IDU-IE clinical practice guidelines.

Hybrid arch frozen elephant trunk repair: evidence from the Canadian Thoracic Aortic Collaborative.

BACKGROUND: The frozen elephant trunk (FET) technique has become an increasingly popular strategy for aortic reconstruction in the setting of extensive thoracic aortic aneurysms or dissections. The objective of this study is to report on the Canadian experience with the FET technique in both the elective and emergent settings. METHODS: A total of 167 consecutive patients (mean age 65±13 years, 30% female, 25% re-operation) underwent elective (70%) and non-elective (30%) aortic arch reconstruction with the FET technique between May 2008 and October 2019 in six centers of the Canadian Thoracic Aortic Collaborative (CTAC). In-hospital clinical endpoints and early imaging endpoints were prospectively collected and analyzed. RESULTS: All 167 patients underwent successful FET implantation. In-hospital mortality occurred in 14 patients (8%), stroke occurred in 22 patients (13%) and temporary and permanent spinal cord ischemia (SCI) occurred in 6 (3.6%) and 3 (1.8%) patients, respectively. Prolonged mechanical ventilation was required in 35 patients (21%), renal failure requiring dialysis in 14 patients (8%) and atrial fibrillation in 59 patients (36%). The median hospital and intensive care unit (ICU) lengths of stay were 3 [interquartile range (IQR): 1, 6] and 10 (IQR: 7, 17) days, respectively. The rate of type 1A endoleak was 3.6%, with the lowest rate in patients who underwent a total arch replacement with a hybrid FET graft (0%) and the highest among patients who had a hemiarch with antegrade thoracic endovascular aortic repair (TEVAR) deployment (25%). The rate of other types of endoleak and stent complications was comparatively low. CONCLUSIONS: The early CTAC experience with the FET operation demonstrates technical feasibility and good early clinical outcomes in elective and emergent patients. Further analysis is required to explore variations in technique and their potential impact on early and late outcomes.

Impact of brain protection strategies on mortality and stroke in patients undergoing aortic arch repair with hypothermic circulatory arrest: evidence from the Canadian Thoracic Aortic Collaborative.

OBJECTIVES: The aim of this study was to investigate the impact of various brain perfusion techniques and nadir temperature cooling strategies on outcomes after aortic arch repair in a contemporary, multicentre cohort. METHODS: A total of 2520 patients underwent aortic arch repair with hypothermic circulatory arrest (HCA) between 2002 and 2018 in 11 centres of the Canadian Thoracic Aortic Collaborative. Primary outcomes included mortality; stroke; a composite of mortality or stroke; and a Society of Thoracic Surgeons-defined composite (STS-COMP) end point for mortality or major morbidity including stroke, reoperation, renal failure, prolonged ventilation and deep sternal wound infection. Multivariable logistic regression and propensity score matching were performed for cerebral perfusion and nadir temperature practices. RESULTS: Antegrade cerebral perfusion was found on multivariable analysis to be protective against mortality [odds ratio (OR) 0.64, 95% confidence interval (CI) 0.48-0.86; P = 0.005], stroke (OR 0.55, 95% CI 0.37-0.81; P = 0.006), composite of mortality or stroke (OR 0.57, 95% CI 0.45-0.72; P = 0.0001) and STS-COMP (OR 0.53, 95% CI 0.41-0.67; P < 0.0001), as compared to HCA alone. Retrograde cerebral perfusion yielded similar outcomes as compared to antegrade cerebral perfusion. When compared to HCA with nadir temperature <24°C, a propensity score analysis of 647 matched pairs identified nadir temperature ≥24°C as predictor of lower mortality (OR 0.62, 95% CI 0.40-0.98; P = 0.04), stroke (OR 0.51, 95% CI 0.31-0.84; P = 0.008), composite of mortality or stroke (OR 0.62, 95% CI 0.43-0.89; P = 0.01) and STS-COMP (OR 0.64, 95% CI 0.49-0.85; P = 0.002). CONCLUSIONS: Antegrade cerebral perfusion and nadir temperature ≥24°C during HCA for aortic arch repair are predictors of improved survival and neurological outcomes.

Sex-Related Differences in Patients Undergoing Thoracic Aortic Surgery.

BACKGROUND: Contemporary outcomes after surgical management of thoracic aortic disease have improved; however, the impact of sex-related differences is poorly understood. METHODS: A total of 1653 patients (498 [30.1%] female) underwent thoracic aortic surgery with hypothermic circulatory arrest between 2002 and 2017 in 10 institutions of the Canadian Thoracic Aortic Collaborative. Outcomes of interest were in-hospital death, stroke, and a modified Society of Thoracic Surgeons-defined composite for mortality or major morbidity (stroke, renal failure, deep sternal wound infection, reoperation, prolonged ventilation). Multivariable logistic regression was used to determine independent predictors of these outcomes. RESULTS: Women were older (mean±SD, 66±13 years versus 61±13 years; P<0.001), with more hypertension and renal failure, but had less coronary disease, less previous cardiac surgery, and higher ejection fraction than men. Rates of aortic dissection were similar between women and men. Rates of hemiarch, and total arch repair were similar between the sexes; however, women underwent less aortic root reconstruction including aortic root replacement, Ross, or valve-sparing root operations (29% versus 45%; P<0.001). Men experienced longer cross-clamp and cardiopulmonary bypass times, but similar durations of circulatory arrest, methods of cerebral perfusion, and nadir temperatures. Women experienced a higher rate of mortality (11% versus 7.4%; P=0.02), stroke (8.8% versus 5.5%; P=0.01), and Society of Thoracic Surgeons-defined composite end point for mortality or major morbidity (31% versus 27%; P=0.04). On multivariable analyses, female sex was an independent predictor of mortality (odds ratio, 1.81; P<0.001), stroke (odds ratio, 1.90; P<0.001), and Society of Thoracic Surgeons-defined composite end point for mortality or major morbidity (odds ratio, 1.40; P<0.001). CONCLUSIONS: Women experience worse outcomes after thoracic aortic surgery with hypothermic circulatory arrest. Further investigation is required to better delineate which measures may reduce sex-related outcome differences after complex aortic surgery.

Differential induction of apoptosis by tumor necrosis factor-related apoptosis-inducing ligand in human ovarian carcinoma cells.

OBJECTIVES: In this study, we examine the sensitivity of a panel of ovarian carcinoma cells, which includes four primary ovarian cancer cell samples, and four normal ovarian epithelium samples to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). We also examine the intracellular regulation of TRAIL-mediated apoptosis. METHODS: The sensitivity to TRAIL was determined by short-term survival assays on seven ovarian carcinoma cell lines, four primary samples of ovarian cancer, and four normal ovarian epithelium samples. We assessed the activation of the apoptotic pathway in TRAIL-resistant and -sensitive tumor cells. The expression of TRAIL receptors was determined by flow cytometry. The protein expression of FADD, XIAP, caspase-8, caspase-3, BAX, and c-FLIP were determined by immunoblot analyses. RESULTS: We show that ovarian cancer cells display variable sensitivity to TRAIL-induced apoptosis although most cell lines have similar sensitivity to cisplatin. Normal ovarian epithelium samples were mostly sensitive to TRAIL. In sensitive cells, TRAIL induced caspase-8-dependent apoptosis, which subsequently led to activation of caspase-3. Both sensitive and resistant cells expressed caspase-8, caspase-3, FADD, XIAP, and c-FLIP at similar levels. A significant enhancement in cell death was observed in TRAIL-resistant cells when c-FLIP(L) levels were downregulated by RNA interference. CONCLUSIONS: These data suggest that sensitivity to TRAIL and chemotherapy does not necessarily correlate in human ovarian cancer cells. Cancerous cells isolated from patients with ovarian cancer show variable sensitivity to TRAIL but most normal ovarian epithelial cells are sensitive. In human ovarian cancer cells, c-FLIP(L) may participate to the regulation of the TRAIL signaling cascade.