Advanced Therapy Medicinal Products as Potential Therapeutic Strategy against COVID-19 and Immune-Related Disorders.

Advanced Therapy Medicinal Products (ATMPs) comprise a heterogenous class of innovative medicinal products, which further require extensive preclinical and clinical assessments before their broader use in the general population [...].

Evaluation of the Regenerative Potential of Platelet-Lysate and Platelet-Poor Plasma Derived from the Cord Blood Units in Corneal Wound Healing Applications: An In Vitro Comparative Study on Corneal Epithelial Cells.

BACKGROUND: Cord blood platelet lysate (CB-PL) and cord blood platelet poor plasma (CB-PPP) have been applied with success in wound healing applications. Pathologies such as Sjogrens's Syndrome (SS) and chronic graft versus host disease (cGVHD) can lead to severe ophthalmology issues. The application of CB-PL and CB-PPP may be strongly considered for damaged cornea healing. This study aimed to the evaluation of the beneficial properties of CB-PL and CB-PPP in corneal wound healing applications. METHODS: Initially, the CB-PL and CB-PPP were produced from donated cord blood units (CBUs), followed by biochemical analysis. Corneal epithelial cells (CECs) were isolated from wistar rats and then cultured with medium containing 20% v/v either of CB-PL or CB-PPP. To define the impact of CB-PL and CB-PPP, biochemical, morphological analysis, scratch-wound assays, and immunoassays in CECs were performed. RESULTS: CB-PL and CB-PPP were characterized by good biochemical parameters, regarding their quality characteristics and biomolecule content. CECs' morphological features did not change after their cultivation with CB-PL or CB-PPP. A scratch wound assay and molecular analysis of CECs expanded with CB-PL indicated higher migratory capacity compared to those cultured with CB-PPP. CONCLUSION: CB-PL and CB-PPP exhibited good properties with respect to cell migration and proliferation, and could be considered an alternative source for eye drop production, to possibly be used in cornea wound healing applications.

Investigating the production of platelet lysate obtained from low volume Cord Blood Units: Focus on growth factor content and regenerative potential.

BACKGROUND: The regenerative potential of platelet lysate (PL) and platelet gel (PG) is mediated by the release of platelets (PLTs) growth factors. The aim of this study was the evaluation of the PL production utilizing low volume single Cord Blood Units (CBUs) and the comparison of the biomolecule content between PLs obtained from intermediate and high volume CBUs. METHODS: CBUs (n = 90) with volumes greater than 50 ml and initial platelet count > 150 × 109/L were used. CBUs were classified into the following groups: group A (50-80 ml), group B (81-110 ml) and group C (111-150 ml). The CBUs were centrifuged twice for the production of the platelet concentrate (PC), which was stored at - 80 °C for at least 48 h. Then, rapidly thawed and the biomolecule content was determined using commercial ELISA kits. The regenerative potential of PLs was evaluated using the scratch wound and in vitro angiogenesis assay. RESULTS: CBPL was produced from low volume single CBUs and contained 3.4 ± 0.3 ×109 PLTs. PL obtained from intermediate and high volume CBUs consisted of 10.2 ±â€¯0.3 and 16.1 ±â€¯0.4 × 109 PLTs. All PL groups were characterized by high biomolecule content. Gap closure was observed within 72 h after the wound assay initiation and the capillary tubes were formed in all study groups. CONCLUSION: This study provided significant evidence regarding the utilization of the low volume CBUs for the production of CB derivatives, thus can serve as healing mediators in regenerative medicine approaches.

Effect of Cord Blood Platelet Gel on wound healing capacity of human Mesenchymal Stromal Cells.

BACKGROUND: Wound healing is a dynamic process, involving the recruitment of growth factors, cytokines, chemokines and cellular populations. Recently, the Cord Blood Platelet Gel (CBPG) has been applied successfully in wound closure and tissue regeneration. Moreover, its proper combination with stem cell populations such as Mesenchymal Stromal Cells (MSCs) may positively improve the wound healing process. Based on the above data, this study aimed to the evaluation of wound healing capacity of MSCs combined with CBPG under in vitro conditions. METHODS: Initially, CBPG was developed from Cord Blood Units (CBUs). The determination of wound healing ability of MSCs was performed using the scratch wound assay. In addition, the morphological features, immunophenotypical characteristics and differentiation capacity of MSCs were evaluated. RESULTS: Scratch wound assay results showed, that CBPG could positively stimulate the MSCs migration. Moreover, MSCs cultured in presence of CBPG were characterized by elongated shape and improved stemness properties as it was indicated by flow cytometric analysis and differentiation process. CONCLUSION: These results clearly showed the beneficial effect of CBPG in combination with MSCs in wound healing. The proper combination of CBPG with stem cells strategy may enhance the healing process in patients with skin erosions.

Vitrified Wharton's jelly tissue as a biomaterial for multiple tissue engineering applications.

Wharton's Jelly (WJ) tissue is a promising biomaterial, for tissue engineering applications. However, its preservation over a long period in order to be readily available needs further optimization. A possible solution could be the vitrification and storage of WJ tissue at low temperatures. The aim of this study is to evaluate the effect of low temperature in the WJ tissue, which was stored at -196 °C, either with the vitrification or conventional cryopreservation methods. WJ tissues were isolated from human umbilical cords, cryopreserved with the above methods and remained for 1 year at -196 °C. Histological analysis of tissue's extracellular matrix (ECM), isolation, and characterization of mesenchymal stromal cells (MSCs) were performed. Histological analysis revealed the presence of ECM components such as collagen, sulfated glycosaminoglycans (sGAGs), and the presence of cell nuclei only in vitrified samples. Furthermore, MSCs were isolated and expanded successfully from vitrified WJ tissues, whereas a few viable cells were obtained from conventionally cryopreserved tissues that were not further expanded. In conclusion, this study indicated the proper preservation of vitrified WJ tissues after 1 year of storage, which eventually could be used in tissue engineering and regenerative medicine approaches.

Good mid-term outcomes after adipose-derived culture-expanded mesenchymal stem cells implantation in knee focal cartilage defects.

PURPOSE: The purpose of the present study was to investigate the mid-term outcomes of a single-stage cell-based procedure in patients with knee focal symptomatic cartilage defects using matrix-induced culture-expanded autologous AD-MSCs. It was hypothesised that the increased number of autologous AD-MSCs after culture expansion is a safe and efficient cartilage repair procedure, which improves overtime chondrogenesis in cartilage lesions METHODS: Twenty-five consecutive patients treated for a symptomatic cartilage defect were prospectively followed for 3 years. The median age of patients was 30.5 (range 16-43) with a median BMI of 23.6 kg/m2 (range 19-29) and an average size of the lesion of 3.5 cm2 (range 2-6). All patients underwent a single-stage procedure consisting in filling each defect with autologous culture-expanded mesenchymal stem cells embedded in a trimmed-to-fit commercially available biodegradable matrix. Pre-operative and post-operative evaluation included knee-related clinical and functional evaluation based on objective and subjective scores at 6, 12, 24 and 36 months and MRI evaluation of the repair tissue using the MOCART score at 12 and 24 months. RESULTS: Clinical outcomes recorded significant improvements (p < 0.05) at the final follow-up compared with baseline as following: all subcategories of KOOS Score, the IKDC subjective from 40.9 (range 20.7-65.6) to 76.9 (range 42-90.3), Tegner Activity Score from 3 (range 2-4) to 4 (range 3-4), VAS for pain from 6 (range 4-8) to 1 (range 0-3). All patients improve significantly their IKDC objective scores. The MRI findings showed complete filling of the defect and integration to the border zone for 65% of the patients. Two patients underwent post-operative biopsies and the histological analysis demonstrated the presence of hyaline-like tissue. CONCLUSIONS: Adipose-derived culture-expanded mesenchymal stem cells were shown to be an efficient and safe single-stage cell-based procedure for symptomatic, full-thickness knee chondral lesions. The findings of the present study demonstrate that all patients presented significant mid-term clinical, functional and radiological improvement. LEVEL OF EVIDENCE: IV.

Serum miRNA-based distinct clusters define three groups of breast cancer patients with different clinicopathological and immune characteristics.

Breast cancer (BCa) is a heterogeneous disease with different histological, prognostic and clinical aspects. Therefore, the need for identification of novel biomarkers for diagnosis, prognosis and monitoring of disease, as well as treatment outcome prediction remains at the forefront of research. The search for circulating elements, obtainable by simple peripheral blood withdrawal, which may serve as possible biomarkers, constitutes still a challenge. In the present study, we have evaluated the expression of 6 circulating miRNAs, (miR-16, miR-21, miR-23α, miR-146α, miR-155 and miR-181α), in operable BCa patients, with non-metastatic, invasive ductal carcinoma, not receiving neoadjuvant chemotherapy. These miRNAs, known to be involved in both tumor cell progression and immune pathways regulation, were analyzed in relation to circulating cytokines, tumor immune-cell infiltration and established prognostic clinicopathological characteristics. We have identified three different clusters, with overall low (C1), moderate (C2) or high (C3) expression levels of these six circulating miRNAs, which define three distinct groups of non-metastatic BCa patients characterized by different clinicopathological and immune-related characteristics, with possibly different clinical outcomes. Our data provide the proof-of-principle to support the notion that, up- or down-regulation of the same circulating miRNA may reflect different prognosis in BCa. Nonetheless, the prognostic and/or predictive potential of these three "signatures" needs to be further evaluated in larger cohorts of BCa patients with an, at least, 5-year clinical follow-up.

Matrix-induced adipose-derived mesenchymal stem cells implantation for knee articular cartilage repair. Two years follow-up.

The present study aims to evaluate the efficacy of matrix-induced adipose-derived mesenchymal stem cells (Ad-MSCs) for cartilage repair of focal chondral knee lesions. Twenty patients were initially treated for symptomatic full-thickness chondral defects and then prospectively followed for two years. All patients underwent a single- stage procedure consisting in filling each defect with autologous culture-expanded mesenchymal stem cells embedded in a trimmed-to-fit commercially available biodegradable matrix. Knee-related function was evaluated based on subjective scores given by two self-reported questionnaires (KOOS and IKDC). Data analysis shows significant improvements (p<0.001) in all values. The mean preoperative scores in the subscales of KOOS, as well as in the IKDC subjective score were constantly increased during the follow-up period with statistically significant differences at 6, 12 and 24 months follow-up. The findings of this study indicate that matrix- induced adipose-derived mesenchymal stem cells implantation is an effective and safe single-staged cell-based procedure to manage full-thickness focal chondral lesions of the knee.

Peripheral T cell responses to tumour antigens are associated with molecular, immunogenetic and cellular features of breast cancer patients.

PURPOSE: Breast cancer is a leading cause of cancer deaths in women, but despite steady improvements in therapies, treatment is still suboptimal. Immunotherapy holds promise as a more effective therapy for breast cancer; supporting this, our prior study showed that patients possessing HER2-reactive CD8+ T cells in blood experience survival superior to patients without these cells. Here, we define a composite set of biomarkers that identify patients with T cell responses to tumour antigens. METHODS: We assessed T cell responses following in vitro stimulation with the HER2, MUC1 and SUR tumour-associated antigens (TAA) by flow cytometry and intracellular cytokine staining in 50 breast cancer patients. We also measured HLA type, serum cytokines, tumour-infiltrating leukocytes and blood leukocyte populations. RESULTS: We found few correlations between TAA-reactive T cells and HLA type, serum cytokines and tumour-infiltrating leukocytes, whereas blood leukocyte phenotypes broadly correlated with TAA responses. This showed monocytes, natural killer cells, dendritic cells and T cells to be inversely associated with both CD4+ and CD8+ T cells reactive to tumour antigens. Moreover, combining multiple parameters improved the accuracy in predicting patients with TAA-responsive T cells. CONCLUSION: This study therefore defines composite immune profiles that identify patients responding to TAAs which may allow better personalisation of cancer therapies.

Successful short-term cryopreservation of volume-reduced cord blood units in a cryogenic mechanical freezer: effects on cell recovery, viability, and clonogenic potential.

BACKGROUND: Cord blood (CB) units are stored from weeks to years in liquid- or vapor-phase nitrogen until they are used for transplantation. We examined the effects of cryostorage in a mechanical freezer at -150°C on critical quality control variables of CB collections to investigate the possible use of mechanical freezers at -150°C as an alternative to storage in liquid- (or vapor-) phase nitrogen. STUDY DESIGN AND METHODS: A total of 105 CB units were thawed and washed at different time intervals (6, 12, 24, and 36 months). For every thawed CB unit, samples were removed and cell enumeration (total nucleated cells [TNCs], mononuclear cells [MNCs], CD34+, CD133+) was performed. In addition, viability was obtained with the use of flow cytometry, and recoveries were calculated. Also, total absolute colony-forming unit counts were performed and progenitor cell recoveries were studied by clonogenic assays. RESULTS: Significant differences (p < 0.05) were observed in certain variables (TNCs, MNC numbers, viability) when they were examined in relation with time intervals, while others (CD34+, CD133+) were relatively insensitive (p = NS) to the duration of time interval the CB units were kept in cryostorage condition. CONCLUSIONS: The data presented suggest that cryopreservation of CB units in a mechanical freezer at -150°C may represent an alternative cryostorage condition for CB cryopreservation.

Optimizing isolation culture and freezing methods to preserve Wharton's jelly's mesenchymal stem cell (MSC) properties: an MSC banking protocol validation for the Hellenic Cord Blood Bank.

BACKGROUND: Mesenchymal stem or stromal cells (MSCs) are a heterogeneous population that can be isolated from many tissues including umbilical cord Wharton's jelly (UC-WJ). Although initially limited in studies such as a hematopoietic stem cell transplantation adjuvant, an increasing number of clinical trials consider MSCs as a potential anti-inflammatory or a regenerative medicine agent. It has been proposed that creating a repository of MSCs would increase their availability for clinical applications. The aim of this study was to assess the optimal isolation and cryopreservation procedures to facilitate WJ MSC banking. STUDY DESIGN AND METHODS: Cells were isolated from UC-WJ using enzymatic digestion or plastic adhesion methods. Their isolation efficacy, growth kinetics, immunophenotype, and differentiation potential were studied, as well as the effects of freezing. Flow cytometry for common MSC markers was performed on all cases and differentiation was shown with histocytochemical staining. Finally, the isolation efficacy on cryopreserved WJ tissue fragments was tested. RESULTS: MSC isolation was successful using both isolation methods on fresh UC-WJ tissue. However, UC-WJ MSC isolation from frozen tissue fragments was impossible. Flow cytometry analysis revealed that only MSC markers were expressed on the surface of the isolated cells while differentiation assays showed that they were capable of trilinear differentiation. All the above characteristics were also preserved in isolated UC-WJ MSCs over the cryopreservation study period. CONCLUSION: These data showed that viable MSCs can only be isolated from fresh UC-WJ tissue, setting the foundation for clinical-grade banking.

Perspectives for the Use of Umbilical Cord Blood in Transplantation and Beyond: Initiatives for an Advanced and Sustainable Public Banking Program in Greece.

The umbilical cord blood (UCB) donated in public UCB banks is a source of hematopoietic stem cells (HSC) alternative to bone marrow for allogeneic HSC transplantation (HSCT). However, the high rejection rate of the donated units due to the strict acceptance criteria and the wide application of the haploidentical HSCT have resulted in significant limitation of the use of UCB and difficulties in the economic sustainability of the public UCB banks. There is an ongoing effort within the UCB community to optimize the use of UCB in the field of HSCT and a parallel interest in exploring the use of UCB for applications beyond HSCT i.e., in the fields of cell therapy, regenerative medicine and specialized transfusion medicine. In this report, we describe the mode of operation of the three public UCB banks in Greece as an example of an orchestrated effort to develop a viable UCB banking system by (a) prioritizing the enrichment of the national inventory by high-quality UCB units from populations with rare human leukocyte antigens (HLA), and (b) deploying novel sustainable applications of UCB beyond HSCT, through national and international collaborations. The Greek paradigm of the public UCB network may become an example for countries, particularly with high HLA heterogeneity, with public UCB banks facing sustainability difficulties and adds value to the international efforts aiming to sustainably expand the public UCB banking system.

Treatment with Umbilical Cord Blood Platelet Lysate Gel Improves Healing of Diabetic Foot Ulcer.

BACKGROUND: This study was conducted to examine the hypothesis that umbilical cord blood platelet lysate (UCB-PL) gel has a significant impact on the healing rate of DFU. Μethods: In this open-labeled, randomized controlled trial, 110 patients were randomized to treatment with UCB-PL gel (UCB-PL group, n = 52) every three days for one month or dressing with normal saline (control group, n = 58). All participants were followed up for 20 weeks post treatment. Ulcer surface area was assessed with the imitoMeasure application at two, four, and six weeks, and two, four and six months. This study's main outcome was the reduction in ulcer size over the six-month study period. RESULTS: The mean ulcer area at baseline was 4.1 cm2 in the UCB-PL group and 1.7 cm2 in the control group. At six months post treatment, patients on the UCB-PL treatment displayed a significant reduction in ulcer size compared to baseline 0.12 (0-8.16) in contrast to a more modest change in the control group 1.05 (0-24.7). The ulcer area was decreased at the end of the study in 40 patients (97.6%) in the UCB-PL group and 27 (73%) in the control group (Fisher's p = 0.002). CONCLUSIONS: The application of UCB-PL gel in DFU resulted in a significant reduction in ulcer size compared to regular saline dressing.

Mesenchymal stromal cell delivery as a potential therapeutic strategy against COVID-19: Promising evidence from in vitro results.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for the coronavirus disease 2019 (COVID-19) pandemic, which was initiated in December 2019. COVID-19 is characterized by a low mortality rate (< 6%); however, this percentage is higher in elderly people and patients with underlying disorders. COVID-19 is characterized by mild to severe outcomes. Currently, several therapeutic strategies are evaluated, such as the use of anti-viral drugs, prophylactic treatment, monoclonal antibodies, and vaccination. Advanced cellular therapies are also investigated, thus representing an additional therapeutic tool for clinicians. Mesenchymal stromal cells (MSCs), which are known for their immunoregulatory properties, may halt the induced cytokine release syndrome mediated by SARS-CoV-2, and can be considered as a potential stem cell therapy. AIM: To evaluate the immunoregulatory properties of MSCs, upon stimulation with COVID-19 patient serum. METHODS: MSCs derived from the human Wharton's Jelly (WJ) tissue and bone marrow (BM) were isolated, cryopreserved, expanded, and defined according to the criteria outlined by the International Society for Cellular Therapies. Then, WJ and BM-MSCs were stimulated with a culture medium containing 15% COVID-19 patient serum, 1% penicillin-streptomycin, and 1% L-glutamine for 48 h. The quantification of interleukin (IL)-1 receptor a (Ra), IL-6, IL-10, IL-13, transforming growth factor (TGF)-ß1, vascular endothelial growth factor (VEGF)-a, fibroblast growth factor (FGF), platelet-derived growth factor (PDGF), and indoleamine-2,3-dioxygenase (IDO) was performed using commercial ELISA kits. The expression of HLA-G1, G5, and G7 was evaluated in unstimulated and stimulated WJ and BM-MSCs. Finally, the interactions between MSCs and patients' macrophages were established using co-culture experiments. RESULTS: Thawed WJ and BM-MSCs exhibited a spindle-shaped morphology, successfully differentiated to "osteocytes", "adipocytes", and "chondrocytes", and in flow cytometric analysis were characterized by positivity for CD73, CD90, and CD105 (> 95%) and negativity for CD34, CD45, and HLA-DR (< 2%). Moreover, stimulated WJ and BM-MSCs were characterized by increased cytoplasmic granulation, in comparison to unstimulated cells. The HLA-G isoforms (G1, G5, and G7) were successfully expressed by the unstimulated and stimulated WJ-MSCs. On the other hand, only weak expression of HLA-G1 was identified in BM-MSCs. Stimulated MSCs secreted high levels of IL-1Ra, IL-6, IL-10, IL-13, TGF-ß1, FGF, VEGF, PDGF, and IDO in comparison to unstimulated cells (P < 0.05) after 12 and 24 h. Finally, macrophages derived from COVID-19 patients successfully adapted the M2 phenotype after co-culturing with stimulated WJ and BM-MSCs. CONCLUSION: WJ and BM-MSCs successfully produced high levels of immunoregulatory agents, which may efficiently modulate the over-activated immune responses of critically ill COVID-19 patients.

Multiple Sclerosis and MEN2 Neoplasia in a Female Patient: A Unique Co-Existence with Expanded Immunological Interest and Therapeutical Challenges, before and after Patient's COVID-19 Infection.

Multiple sclerosis (MS) and its various comorbidities that may be observed are of great interest due to the complexity of MS pathophysiology and all of the immunological changes that follow. The incidence of cancer in MS has been investigated for several years, as not only does it affect ongoing therapeutical decisions, but also, certain disease-modifying treatments (DMTs) may increase the risk of tumorigenesis. For the first time, we present a case of a female patient with pediatric-onset MS (POMS) and multiple endocrine neoplasia 2B (MEN2B) and analyze the immunological impact of these diseases on the therapeutical choice, under the umbrella of her COVID-19 infection and the SARS-CoV-2 pandemic as a whole. We also review the existing literature regarding the immunogenetic and immunological correlations between these two extremely rare diseases and discuss the most suitable treatment for our case, which seems to be an anti-CD20 agent due to a better outcome in putative MS worsening and tumor progression, when killer immunoglobulin-like receptors' (KIR) expression is reduced in natural killer (NK) cells. We also broaden our concerns on this comorbidity issue, at the same time focusing on the future research needed in this unexplored field of the comorbidity of MS and cancers.

HLA-genotyping by next-generation-sequencing reveals shared and unique HLA alleles in two patients with coexisting neuromyelitis optica spectrum disorder and thymectomized myasthenia gravis: Immunological implications for mutual aetiopathogenesis?

The exact immunopathogenesis, genetic mechanisms and triggering factors underlying myasthenia gravis (MG) and neuromyelitis optica (NMO) remain unknown and the coexistence may underline an aetiopathogenetic link be- tween these two diseases. We report the cases of two thymectomized patients with acetylcholine receptor (AChR) antibody (Ab)-positive MG who eventually developed AQP4-NMO. Next-Generation Sequencing (NGS) analysis showed that patient-1 had two HLA alleles previously associated with MG, mainly HLA-A*01:01:01 and HLA-DRB1*03:01, present in a haplotype in Caucasian MG patients (HLA-A1-B8-DR3-DQ2). Patient-2, expressed HLA-C*07:01:01, a well characterized MG risk factor and HLA-DQB1*05:02:01, previously described both in MG and NMO patients. Finally, we observed two common alleles in patient 1 and 2, HLA-DQA1*05:01:01 and HLA-DPB1*04:02:01. We believe that this study provides clinical evidence of the role of specific HLA alleles in rare forms of combined human peripheral and CNS autoimmunity, a fact that enhances the aim towards tailor-made therapeutic decision making.

Low usage rate of banked sibling cord blood units in hematopoietic stem cell transplantation for children with hematological malignancies: implications for directed cord blood banking policies.

Directed sibling cord blood banking is indicated in women delivering healthy babies who already have a sibling with a disease that is potentially treatable with an allogeneic cord blood transplant. We evaluated the effectiveness of a national directed cord blood banking program in sibling HLA-identical stem cell transplantation for hematological malignancies and the factors influencing the usage rate of the stored cord blood units. Fifty families were enrolled from which, 48 cord blood units were successfully collected and 2 collections failed due to damaged cord/placenta at delivery. Among enrolled families 4 children needed transplantation; however, only one was successfully transplanted using the collected cord blood unit containing 2×10(7) nucleated cells/kg in conjunction with a small volume of bone marrow from the same HLA-identical donor. Two children received grafts from matched unrelated donors because their sibling cord blood was HLA-haploidentical, while the fourth one received bone marrow from his HLA-identical brother, since cord blood could not be collected due to damaged cord/placenta at delivery. With a median follow-up of 6 years (range, 2-12) for the 9 remaining HLA-matched cord blood units, none from the prospective recipients needed transplantation. The low utilization rate of sibling cord blood in the setting of hematopoietic stem cell transplantation for pediatric hematological malignant diseases necessitates the development of directed cord blood banking programs that limit long-term storage for banked cord blood units with low probability of usage such as non-HLA-identical or identical to patients who are in long-term complete remission.

Modern Approaches in Cardiovascular Disease Therapeutics: From Molecular Genetics to Tissue Engineering.

Cardiovascular disease (CVD) currently represents one of the leading causes of death worldwide. It is estimated that more than 17.9 million people die each year due to CVD manifestations. Often, occlusion or stenosis of the vascular network occurs, either in large- or small-diameter blood vessels. Moreover, the obstruction of small vessels such as the coronary arteries may be related to more pronounced events, which can be life-threatening. The gold standard procedure utilizes the transplantation of secondary vessels or the use of synthetic vascular grafts. However, significant adverse reactions have accompanied the use of the above grafts. Therefore, modern therapeutic strategies must be evaluated for better disease administration. In the context of alternative therapies, advanced tissue-engineering approaches including the decellularization procedure and the 3D additive bioprinting methods, have been proposed. In this way the availability of bioengineered vascular grafts will be increased, covering the great demand that exists globally. In this Special Issue of Bioengineering, we tried to highlight the modern approaches which are focused on CVD therapeutics. This issue includes articles related to the efficient development of vascular grafts, 3D printing approaches and suitable atherosclerosis models.

Improved Repopulation Efficacy of Decellularized Small Diameter Vascular Grafts Utilizing the Cord Blood Platelet Lysate.

BACKGROUND: The development of functional bioengineered small-diameter vascular grafts (SDVGs), represents a major challenge of tissue engineering. This study aimed to evaluate the repopulation efficacy of biological vessels, utilizing the cord blood platelet lysate (CBPL). METHODS: Human umbilical arteries (hUAs, n = 10) were submitted to decellularization. Then, an evaluation of decellularized hUAs, involving histological, biochemical and biomechanical analysis, was performed. Wharton's Jelly (WJ) Mesenchymal Stromal Cells (MSCs) were isolated and characterized for their properties. Then, WJ-MSCs (1.5 × 106 cells) were seeded on decellularized hUAs (n = 5) and cultivated with (Group A) or without the presence of the CBPL, (Group B) for 30 days. Histological analysis involving immunohistochemistry (against Ki67, for determination of cell proliferation) and indirect immunofluorescence (against activated MAP kinase, additional marker for cell growth and proliferation) was performed. RESULTS: The decellularized hUAs retained their initial vessel's properties, in terms of key-specific proteins, the biochemical and biomechanical characteristics were preserved. The evaluation of the repopulation process indicated a more uniform distribution of WJ-MSCs in group A compared to group B. The repopulated vascular grafts of group B were characterized by greater Ki67 and MAP kinase expression compared to group A. CONCLUSION: The results of this study indicated that the CBPL may improve the repopulation efficacy, thus bringing the biological SDVGs one step closer to clinical application.

Optimizing Decellularization Strategies for the Efficient Production of Whole Rat Kidney Scaffolds.

BACKGROUND: Renal dysfunction remains a global issue, with chronic kidney disease being the 18th most leading cause of death, worldwide. The increased demands in kidney transplants, led the scientific society to seek alternative strategies, utilizing mostly the tissue engineering approaches. Unlike to perfusion decellularization of kidneys, we proposed alternative decellularization strategies to obtain acellular kidney scaffolds. The aim of this study was the evaluation of two different decellularization approaches for producing kidney bioscaffolds. METHODS: Rat kidneys from Wistar rats, were submitted to decellularization, followed two different strategies. The decellularization solutions used in both approaches were the same and involved the use of 3-[(3-cholamidopropyl) dimethylammonio]-1-propanesulfonate and sodium dodecyl sulfate buffers for 12 h each, followed by incubation in a serum medium. Both approaches involved 3 decellularization cycles. Histological analysis, biochemical and DNA quantification were performed. Cytotoxicity assay and repopulation of acellular kidneys were also applied. RESULTS: Histological, biochemical and DNA quantification confirmed that the 2nd approach had the best outcome regarding the kidney composition and cell elimination. Acellular kidneys from both approaches were successfully recellularized. CONCLUSION: Based on the above data, the production of kidney scaffolds with the proposed cost- effective decellularization approaches, was efficient.