Methods: Human umbilical cord-derived MSCs were cultured in media supplemented with 10% FBS (control), UCS, or HPL. These groups were further treated with nano-curcumin (0.3 μM) or crocin (2.5 μM), either individually or in combination. Cell proliferation was measured using the MTT assay, apoptosis was assessed by Annexin V/PI flow cytometry, and pluripotency gene expression (Sox2, Nanog, Oct4) was analyzed by RT-qPCR.

Results: UCS and HPL supplements significantly increased MSC proliferation compared to the FBS control (p < 0.001). Specifically, UCS reduced the population doubling time by approximately 50%. Supplementation with crocin reduced apoptosis by up to 30% (p = 0.04) and significantly enhanced the expression of the pluripotency genes Sox2 and Nanog, particularly in cultures supplemented with HPL. In contrast, nano-curcumin inhibited MSC proliferation and increased apoptosis across all tested conditions.

Discussion: The results demonstrate that UCS and HPL are effective, viable alternatives to FBS, promoting superior MSC expansion. The anti-apoptotic and stemness-enhancing properties of crocin highlight its potential as a valuable additive for improving culture quality and cell survival. The cytotoxic effects observed with nano-curcumin underscore a critical need for dose-optimization studies. The primary limitation of this study is the use of fixed concentrations for the supplements, which warrants further investigation across a range of doses.

Conclusion: UCS and HPL are robust, ethically sound replacements for FBS in MSC biomanufacturing. Crocin can further enhance culture outcomes by improving cell survival and maintaining stemness. These findings support the development of optimized, xeno-free culture systems for scalable MSC production, which is crucial for advancing regenerative medicine therapies.]]> https://www.benthamscience.com/article/1494402026-03-18Introduction: Therapeutic angiogenesis is a new potential strategy for treating Peripheral Arterial disease (PAD). Human Umbilical Cord Blood Mesenchymal Stem Cells (hUCB-MSCs) and their secreted exosomes can effectively promote the formation of new blood vessels, making them important targets for research on therapeutic angiogenesis.

Aim: This study investigated the impact of hUCB-MSCs and their derived exosomes on the proliferation and migration of vascular endothelial tip cells.

Methods: The cultivation and identification of endothelial tip cells, hUCB-MSCs, and exosomes were conducted, followed by co-culturing hUCB-MSCs with tip cells and incubating exosomes with tip cells. qPCR was utilized to assess the expression levels of microRNAs in exosomes, as well as the expression levels of cell proliferation-related markers, miR-21-5p, and TGF-β1 in tip cells. Western blotting was used to analyze the levels of key factors associated with cell proliferation and apoptosis. Furthermore, CCK-8 assay, EdU staining, Transwell assay, and flow cytometry were utilized to evaluate cell viability, proliferation, migration, and apoptosis, respectively.

Results: hUCB-MSCs/exosomes significantly enhanced tip cell proliferation and migration, while inhibiting apoptosis, with exosomes demonstrating superior efficacy. miR-21-5p, found within exosomes, was identified as a key factor downregulating TGF-β1 within tip cells. Furthermore, heightened levels of miR-21-5p were observed to enhance the proliferation and migration of tip cells while simultaneously inhibiting apoptosis. Notably, the impact of miR-21-5p was counteracted upon exposure to TGF-β1.

Conclusion: hUCB-MSC-derived exosomes, enriched with miR-21-5p, enhance endothelial tip cell function through targeted TGF-β1 suppression, offering a viable avenue for clinical interventions in PAD treatment.

Methods: Immunofluorescence staining was used to analyze the expression and localization of laminin and ICC in paraffin-embedded colon sections from HSCR patients. Whole-mount preparations and confocal microscopy were employed to visualize the ICC network. Laminin and c-Kit expression levels were evaluated by Western blot and qPCR. Isolated ICCs were treated with laminin-targeting siRNA or exogenous laminin protein. The effects on c-Kit expression, cell viability, and apoptosis were assessed via Western blot, qRT-PCR, MTT assay, and TUNEL staining.

Results: Laminin and ICCs were localized in the muscle layers and intermuscular regions, with laminin partially colocalizing with ICCs. In HSCR colon segments, laminin and ICC expression were significantly reduced, and ICC networks were disrupted (p < 0.05). Silencing laminin decreased c- Kit expression, ICC viability, and increased apoptosis, whereas exogenous laminin restored c-Kit expression, enhanced viability, and reduced apoptosis (p < 0.05).

Discussion: Our findings suggest laminin deficiency contributes to ICC loss in HSCR, impairing intestinal motility. This aligns with prior ECM-neural crest cell studies but contrasts with reports of elevated laminin in whole-tissue analyses, possibly due to regional or temporal differences. Limitations include reliance on rodent ICC models.

Conclusion: Laminin supports ICC viability and prevents apoptosis. Reduced laminin expression in HSCR contributes to the loss of ICC, disrupting pacemaker activity and impairing colonic motility.]]> https://www.benthamscience.com/article/1496622026-03-18Introduction: Fibroblast growth factor 2 (FGF2) plays a crucial role in regulating the osteogenic differentiation of progenitor cells. However, the process by which this occurs is not yet fully understood. In this study, we aimed to investigate whether FGF2 stimulates the osteogenesis of precursor cells through the yes-associated protein (YAP) and large tumor suppressor kinases 1/2 (LATS1/2).

Methods: Human bone marrow stromal cells (hBMSCs) were cultured in osteogenic medium supplemented with FGF2 at concentrations of 2 ng/mL, 10 ng/mL, and 50 ng/mL for 2, 7, or 21 days. Alizarin red staining was performed to identify mineralization after 21 days of culture. RT-qPCR was conducted to detect the mRNA expression of Yap, Lats1, Lats2, Runx2, Bglap, and β-Actin. Immunofluorescence staining was carried out to detect the protein expression of YAP and LATS1/2. Data was analyzed with a p-value set at 0.05.

Results: Mineralization was most significant at 10 ng/ml of FGF2 for 7 days and increased with concentrations of FGF2 from 0 ng/ml to 10 ng/ml for 7 days (p < 0.05) but decreased at the high concentration of 50 ng/ml for 2 days (p < 0.05). mRNA expression of Yap, Runx2, and Bglap increased in concordance with the increasing mineralization levels, but Lats1/2 mRNA decreased. mRNA expression levels were dose-dependent when FGF2 was added for 7 days (p < 0.05) and time-dependent when FGF2 concentration was at 10 ng/ml (p < 0.05). At the protein level, YAP increased while LATS1/2 decreased, indicating that LATS1/2 decreased, and YAP increased at higher mineralization levels when hBMSCs were cultured with 10 ng/ml of FGF2 for 7 days.

Discussion: Consistent with our results, prior research has also indicated that lower concentrations of FGF2 enhance cell proliferation, thereby increasing the cell population for later osteogenic differentiation. However, excessive expansion can negatively affect differentiation. The mechanism of FGF2 regulation in stem cell osteogenic differentiation needs more exploration.

Conclusion: Optimal concentrations and durations of FGF2 are critical for the osteogenic differentiation of hBMSCs. Moreover, it has been observed that mineralization correlates well with increasing YAP and decreasing LATS1/2 during osteogenic differentiation.