Since cardiovascular diseases are the leading cause of death worldwide, autologous endothelial colony-forming cells (ECFCs) are often considered for tissue engineering applications and expanded in vitro , e.g., to seed vascular scaffolds as a vascular substitute or within EC regenerative cell therapies. Despite the simple isolation procedure, success rates vary markedly between individuals. The primary goal of this study was to define pre-collection parameters in participant blood donors that reveal the potential of generating high-quality ECFCs for vascular regeneration therapy. ECFCs were isolated from peripheral blood mononuclear cells from 43 healthy blood donors (23 females, 20 males) and characterized regarding their expandability for therapeutic applications. In addition to data on blood pressure, heart rate, and vascular function – using pulse-wave velocity (PWV) and flow-mediated dilation (FMD) examinations –, questionnaires regarding physical activity, family history, lifestyle factors, and medication were collected. A key finding is the association between the number of giant cells in ECFC cultures and arterial stiffness in blood donors, as reflected by PWV > 8.5 m/s and an FMD < 5%, as well as the absence or at least a significantly reduced proliferative capacity of ECFCs in vitro . It was also found that donors with a higher number of cardiovascular risk factors – such as higher age or body mass index, lack of exercise, and high blood pressure – not only exhibited increased vascular stiffness, but also ECFC isolates that showed reduced ex vivo expandability. Future studies must investigate whether lifestyle changes could also have a positive effect on vascular quality or ECFC quality.
Introduction Chemotherapy, especially with alkylating agents, can impair spermatogonial stem cells in young males, leading to infertility. This study evaluates the therapeutic efficacy of bone-marrow-derived Mesenchymal Stem Cells (BM-MSCs) in restoring testicular function of the dysfunctional testes in a chemo-ablated infertile rat model. Method Wistar male rats were randomly divided into three groups: the first control group without any treatment; the second group was infertile chemo-ablated, generated through the intraperitoneal injections of 15 mg/kg of Busulfan, twice with a two-week interval; and the third group was chemo-ablated and subsequently treated with intra-testicular injections of enriched MSCs. The BM-MSCs were cultured and sorted using magnetic-activated cell sorting (MACS), and purity was confirmed by Fluorescence-Activated Cell Sorting (FACS) analysis of positive and negative cell-surface markers. Purified MSCs were injected into the testes of the infertile rats. Results were evaluated after 2-10 weeks of treatment. After 2 weeks of MSC treatment, mating trials for these males were conducted every 2 weeks, and their testes, as well as serum samples; were collected for therapeutic assessment of MSCs. Results Histological assessments of the excised testes revealed a gradual, significant improvement within 10 weeks of treatment. Additionally, the expression levels of stem cell and sperm maturation markers were elevated in the testes of treated animals, indicating recovery of spermatogenesis. Hormonal analyses demonstrated normalisation of FSH and Testosterone levels after treatment. Mating studies also demonstrated that the MSC-treated group delivered 7-8 pups per litter, whereas chemo-ablated animals remained totally infertile. Conclusions Overall findings support MSC transplantation as a promising approach to restore testicular function and fertility, inspiring continued exploration in regenerative medicine.
Objective: This study evaluated the feasibility of constructing tubular grafts using adipose-derived stem cells and smooth muscle cells seeded onto bladder acellular matrix scaffolds for repairing long-segment urethral defects. Methods: A tissue-engineered urethra was constructed in dogs using autologous adipose-derived stem cells (ADSCs) and smooth muscle cells (SMCs) seeded onto bladder acellular matrix grafts (BAMGs). First, autologous ADSCs and SMCs were isolated and cultured separately from canine tissues via enzymatic digestion. ADSCs were characterized by flow cytometry and assessed for multi-lineage differentiation potential. Fresh canine bladder tissues were decellularized to prepare BAMGs. Expanded ADSCs were seeded onto one side of the BAMGs, and SMCs onto the other side. After one week of in vitro co-culture, the cell-scaffold constructs were rolled into tubular tissue-engineered urethras. Twenty-four male Beagle dogs were subjected to creation of a 4-cm anterior urethral defect and were randomly divided into experimental and control groups, with 12 dogs in each group. In the experimental group, the defects were repaired with the tissue-engineered urethra constructed from autologous ADSCs/SMCs and BAMGs, whereas the control group received BAMG alone. At 1, 2, 4, and 8 weeks postoperatively, tissue samples were harvested for evaluation of regenerative outcomes; at 8 weeks postoperatively, retrograde urethrography and cystourethroscopy were performed at the terminal assessment of each animal to assess the repair efficacy. Results: ADSCs cultured in vitro were predominantly spindle-shaped and immunophenotypically characterized as CD90 + CD29 + CD34 − CD45 − , with confirmed adipogenic and osteogenic differentiation by Oil Red O and Alizarin Red S staining, respectively. ADSCs and SMCs seeded onto BAMGs showed favorable biocompatibility and growth.All experimental animals (n = 12) maintained patent urethras without stricture. In the control group, 2 animals died within the first 2 weeks, and of the 10 animals available for 8-week assessment, 6 developed overt urethral strictures. Experimental grafts displayed satisfactory epithelialization and smooth surfaces, while controls showed contracture, scarring, and calculi. Histologically, experimental animals exhibited well-organized multilayered urothelium and muscle with progressive regeneration over time, in contrast to the extensive fibrosis and scant smooth muscle seen in controls. At 8 weeks, cystoscopy and urethrography confirmed smooth mucosa and unobstructed lumens without extravasation in the experimental group, versus overt urethral collapse and obstruction in the control group. Conclusions: These findings provide preclinical evidence that a tubularized scaffold based on bladder acellular matrix and co-seeded with ADSCs and SMCs is a promising strategy for repairing 4-cm circumferential urethral defects in a healthy canine model. Because this was an acute-defect study in an otherwise healthy urethra with follow-up limited to 8 weeks, the findings should be interpreted as a promising preclinical strategy rather than an established alternative to current autologous tissue-based procedures.
Introduction Cell processing facilities commonly employ manual wiping to decontaminate biosafety cabinet (BSC) surfaces during changeovers between manufacturing lots. Although wiping is intended to remove contaminants, it may also inadvertently redistribute them across surfaces or transfer them to operators’ gloves. Quantitative data on the impact of wiping techniques, repetition, and surface materials on the dispersion and transfer of biological contaminants within BSCs remain limited. This study systematically evaluated the spread of contamination during wiping operations in BSCs, focusing on three representative indicators: DNA, protein, and viable bacteria. Methods The redistribution of contaminants on stainless steel (SUS304) and polyethylene terephthalate surfaces was evaluated using DNA, protein, and Bacillus subtilis endospores as model indicators. Two wiping protocols were evaluated: multi-pass wiping (reusing the same sheet) and replacement wiping (using a new sheet after the source area), each performed either once or four times with ethanol-moistened nonwoven sheets under standardized mechanical conditions based on EN 16615:2015. Residual DNA and protein were quantified fluorometrically (n = 6 per condition), whereas bacterial surface contamination was measured by contact culture (n = 12 per condition). Bacterial transfer to gloves was evaluated across increasing numbers of wiping sheets (n = 16 per sheet-number condition). Results DNA and protein residues predominantly persisted in the source area under all conditions, with minimal transfer to adjacent areas. However, measurable residues persisted in the source area even after four wiping cycles. By contrast, viable bacteria were readily redistributed to the affected area during multi-pass wiping, whereas bacterial detection in the affected area was less frequent during replacement wiping. Bacterial transfer to gloves decreased monotonically with an increasing number of wiping sheets (Cramér's V = 0.82; Cochran–Armitage trend test, p = 1.7 × 10−14). Conclusions Wiping operations within BSCs serve both as decontamination steps and potential vehicles for contamination redistribution. These findings provide quantitative evidence for implementing replacement wiping strategies, increasing the frequency of sheet changes, and incorporating nonmicrobial residue indicators into changeover validation for cell processing facilities.
Introduction As a new therapeutic approach, regenerative medicine is expected to provide novel tools for the treatment of diseases that cannot be adequately addressed via the use of conventional therapies, and it has now been introduced into treatment strategies for ophthalmic diseases where its efficacy (i.e., improvement of not only corneal endothelial cell [CEC] density, but also best-corrected visual acuity [BCVA]) and safety for the treatment of corneal endothelial failure has recently been reported. While the value of new keratoplasty methods has traditionally been assessed primarily in terms of clinical outcomes and healthcare costs, successful treatment in the clinical setting using corneal regenerative medicine products may generate broader benefits for society. For example, restoration of BCVA may reduce the need for disability pension expenditure. With that said, such societal benefits have received relatively limited attention in the overall evaluation of regenerative medicine products. Hence, using a neltependocel (i.e., a human allogeneic CEC-processed product) as a case study, the purpose of this present study was to evaluate the potential societal value of cultured human CEC transplantation through cost offsets from avoided disability pension and break-even life expectancy. Methods In this retrospective study, the transplantation therapy-related BCVA improvement, disability pension grade change, benefits amount, medical cost, break-even life expectancy, abridged life tables, and potential cost savings were evaluated through a scenario-based analysis using publicly available data sources. Results As part of the scenario-based analysis, in 2 of 12 cases in a phase 3 clinical trial, BCVA improvement corresponding to a disability pension grade change from Grade 1 to non-eligibility criteria was observed. The annual Grade 1 disability basic pension benefit amount of the Japan Pension Service was ¥1,059,125 Japanese yen (i.e., $6746 U S. dollars). Initial therapy costs were equivalent to ¥10,153,730 ($64,673). The follow-up costs at 2-weeks and 1-, 3-, and 6-months postoperative were ¥10,967 ($70), ¥11,534 ($73), ¥12,101 ($77), and ¥13,274 ($85), respectively. The annual follow-up costs after 6-months postoperative were estimated to be ¥26,548 ($169). The break-even life expectancy was estimated to be 9.88 years. In cases where the disability pension grade improved, even at 65 years of age, which corresponds to the old-age (elderly) population, the potential cost savings were estimated to be ¥9,902,668 ($63,074) for male and ¥14,972,621 ($95,367) for female. Conclusions The findings in this study through a scenario-based analysis reveal that the surgical transplantation of neltependocel, a novel regenerative-medicine treatment for corneal endothelial failure, is estimated to potentially result in reductions in disability pension expenditures that exceed the initial and subsequent follow-up costs.
