Cardiovascular cell therapy represents a promising field, with several approaches currently being tested. The advanced therapy medicinal product (ATMP) for the ongoing METHOD clinical study ("Bone marrow derived cell therapy in the stable phase of chronic ischemic heart disease") consists of fresh mononuclear cells (MNC) isolated from autologous bone marrow (BM) through density gradient centrifugation on standard Ficoll-Paque. Cells are tested for safety (sterility, endotoxin), identity/potency (cell count, CD45/CD34/CD133, viability) and purity (contaminant granulocytes and platelets). The aims of the present work were (1) to optimize the cell manufacturing process in order to reduce contaminants and (2) to implement additional assays in order to improve product characterization and evaluate product stability. BM-MNC were isolated by density gradient centrifugation on Ficoll-Paque. The following process parameters were optimized throughout the study: gradient medium density; gradient centrifugation speed and duration; washing conditions. Differential cell count was performed by an automated hematology cell analyzer. Immunophenotype and cell viability were determined by flow cytometry. Functional hematopoietic and mesenchymal precursors and cells with angiogenic potential were assessed by colony-forming assays, cell invasion capacity by a fluorimetric assay. Sterility was tested using an automated microbial detection system, endotoxin by a kinetic chromogenic Limulus amebocyte lysate test. T-test was used for statistical analysis. A new manufacturing method was set up, based on gradient centrifugation on low density Ficoll-Paque, followed by 2 washing steps, of which the second one at low speed. It led to significantly higher removal of contaminant granulocytes and platelets, improving product purity; the frequencies of CD34+ cells, CD133+ cells and functional hematopoietic and mesenchymal precursors were significantly increased. The process was successfully validated according to Good Manufacturing Practices. The resulting ATMP mainly consisted of viable MNC including CD34+ and CD133+ cell subsets (2.98% ± 1.90% and 0.83% ± 1.32%, respectively), CD184/CXCR4+ cells (34% ± 15%), CD34+/CD133+/CD309+ endothelial precursors (44 ± 21 in 106 total cells), cells with invasion capacity, functional hematopoietic and mesenchymal precursors, cells with angiogenic potential; it was stable for 20 hours at 10°C. The methodological optimization described here resulted in a significant improvement of ATMP quality, a crucial issue to clinical applications in cardiovascular cell therapy.

(1) Introduction: Intraoperative preservation solutions for saphenous vein grafts may influence the endothelial structure and increase the risk of graft failure after coronary surgery. The aim of the study was to compare the efficacy of three solutions in maintaining the endothelial cell integrity of venous segments. (2) Methods: We tested the efficacy of physiological saline solution (PSS), heparinized autologous blood (HAB) and DuraGraft^® in preserving the endothelium of vein segments by evaluating the degree of endothelial cell apoptosis. Two incubation times (2 and 4 h from harvesting) were used for each solution. The quantification of apoptotic cells was computed as the intensity nuclei/intensity area ratio. (3) Results: After 2 h of ischemia, the degree of apoptosis decreased progressively across the use of DuraGraft, HAB and PSS (p = 0.004), although only the difference between DuraGraft and PSS yielded a statistical significance (p = 0.002). After 4 h, a similar decrease in apoptosis was shown across the three media; however, statistical significance was not reached. The analysis of the elapsed time (2 or 4 h of incubation) showed that this was a relevant factor in maintaining the endothelial structural integrity independently from the storage solution (test for interaction of media and time p = 0.010). (4) Conclusion: Within 2 h of incubation, endothelial structural integrity depended on the incubating medium. DuraGraft better protected the SVG against ischemic-induced apoptosis when compared to saline solution. Prolonged ischemia was associated with extended endothelium damage and none of the studied solutions protected the vein graft.

Objective: Transcatheter aortic valve replacement (TAVR) through alternative access routes is indicated in patients with severe aortic valve stenosis and diseased peripheral arteries. We analysed and compared the outcome of patients undergoing transapical (TA) and direct transaortic (TAO) TAVR procedures. Methods: Preoperative characteristics, procedural details, and thirty-day outcome of patients undergoing transapical (TA-TAVR group) and direct transaortic (TAO-TAVR group) TAVR procedures were prospectively collected and retrospectively analysed. Results: From March 2012 to March 2022, 81 TA and 82 TAO-TAVR (total: 163 cases) were performed with balloon-expanding (n = 120; 73.6%) and self-expandable (n = 43; 26.4%) valves. The mean age was 79.7 ± 6.2 and 81.9 ± 6.7 years for the TA- and TAO-TAVR groups, respectively (p = 0.032). Females were more represented in the TAO-TAVR group (56% vs. 32%; p = 0.003) while TA-TAVR patients showed a higher prevalence of previous vascular surgery (20% vs. 6%; p = 0.01), previous cardiac surgery (51% vs. 3.6%; p < 0.001), and porcelain aorta (22% vs. 5%; p = 0.001). The mean ejection fraction was 49.0 ± 14.6% (TA) and 53.5 ± 12.2% (TAO) (p = 0.035) while mean gradients were 35.6 ± 13.2 mmHg (TA) and 40.4 ± 16.1 mmHg (TAO) (p = 0.045). The median EuroSCORE-II was 5.0% (IQR: 3.0–11.0) and 3.9% (IQR: 2.5–5.4) for the TA- and TAO-TAVR groups, respectively (p = 0.005). The procedural time was shorter for TA procedures (97 min (IQR: 882–118) vs. 102 min (IQR: 88–129); p = 0.133). Mortality at day 30 was 6% in both groups (p = 1.000); the permanent pacemaker implantation rate was similar (8.6% vs. 9.7%; p = 1.000), and hospital stay was shorter for the TAO group (8 days (IQR: 6–11) vs. 10 days (IQR: 7–13); p = 0.025). Conclusions: Our results show that transapical and direct transaortic TAVR in high-risk patients with diseased peripheral arteries provide satisfactory clinical results with similar thirty-day outcomes.

Background: Surgical aortic valve replacement with rapid deployment bioprosthesis guarantees good hemodynamic results but carries the risk of paravalvular leaks. To address this issue, an annulus stabilization technique has been recently developed. Methods: Clinical and hemodynamic parameters from patients treated for aortic valve replacement with the rapid deployment bioprosthesis and a concomitant annulus stabilization technique were prospectively collected and retrospectively analyzed. Echocardiographic data at discharge and at 1-year follow-up were collected and analysed. Results: A total of 57 patients (mean age 74.3 ± 6.1 years) with symptomatic aortic valve stenosis underwent aortic valve replacement with the rapid deployment bioprosthesis and concomitant annulus stabilization technique (mean valve size: 23.8 ± 1.9 mm). Combined procedures accounted for 56.1%. Hospital mortality was 1.8% and a new pacemaker for conduction abnormalities was implanted in 10 patients. The pre-discharge echocardiographic control showed absence of paravalvular leaks of any degree in all patients with mean valve gradient of 9.6 ± 4.0 mmHg. The 1-year echocardiographic control confirmed the good valve hemodynamic (mean gradient of 8.0 ± 2.8 mmHg) and absence of leaks. Conclusion: In this preliminary clinical experience, the annulus stabilization technique prevents postoperative paravalvular leaks after rapid deployment aortic valve implantation, up to 1-year postoperatively. Studies on larger series are of paramount importance to confirm the long-term efficacy of this new surgical technique.

Introduction: Some evidence suggests that surgical minimally invasive (MIDCAB) and hybrid coronary revascularization (HCR) are safe and potentially effective at short-term follow-up. Data on long-term outcomes are more limited and inconclusive. Methods: Between February 2013 and December 2023, a total of 1997 patients underwent surgical coronary artery revascularization at our institution, of whom, 92 (4.7%) received left anterior mini-thoracotomy access (MIDCAB), either isolated (N = 78) or in combination with percutaneous coronary intervention (N = 14, HCR group). Results: After a median follow-up of 75 months (range 3.1: 149 months), cardiac mortality was 0% while overall mortality was 3%, with one in-hospital mortality and two additional late deaths. Conversion to sternotomy happened in two patients (2.1%), and surgical re-explorations occurred in five patients (4.6%), of whom three for bleeding and two for graft failure. All patients received left internal mammary (LIMA) to left anterior descending artery (LAD) grafting (100%). In the HCR group, 10 patients (72%) showed percutaneous revascularization (PCI) after MIDCAB, showing PCI on a mean of 1.6 ± 0.6 vessels and implanting 2.1 ± 0.9 drug-eluting stents. Conclusions: MIDCAB, in isolation or in association with hybrid coronary revascularization, is associated with encouraging short- and long-term results in selected patients discussed within a dedicated heart-team.