Nanofibrous Scaffolds' Ability to Induce Mesenchymal Stem Cell Differentiation for Soft Tissue Regenerative Applications

Abstract

Mesenchymal stem cells (MSCs) have gained recognition as a highly versatile and promising cell source for repopulating bioengineered scaffolds due to their inherent capacity to differentiate into multiple cell types. However, MSC implantation techniques have often yielded inconsistent clinical results, underscoring the need for advanced approaches to enhance their therapeutic efficacy. Recent developments in three-dimensional (3D) bioengineered scaffolds have provided a significant breakthrough by closely mimicking the in vivo environment, addressing the limitations of traditional two-dimensional (2D) cell cultures. Among these, nanofibrous scaffolds have proven particularly effective, offering an optimal 3D framework, growth-permissive substrates, and the delivery of trophic factors crucial for MSC survival and regeneration. Furthermore, the selection of appropriate biomaterials can amplify the paracrine effects of MSCs, promoting both proliferation and targeted differentiation. The synergistic combination of MSCs with nanofibrous scaffolds has demonstrated remarkable potential in achieving repair, regeneration, and tissue-specific differentiation with enhanced safety and efficacy, paving the way for routine clinical applications. In this review, we examine the most recent studies (2013-2023) that explore the combined use of MSCs and nanofibrous scaffolds for differentiation into cardiogenic, epithelial, myogenic, tendon, and vascular cell lineages. Using PubMed, we identified and analyzed 275 relevant articles based on the search terms "Nanofibers", "Electrospinning", "Mesenchymal stem cells", and "Differentiation". This review highlights the critical advancements in the use of nanofibrous scaffolds as a platform for MSC differentiation and tissue regeneration. By summarizing key findings from the last decade, it provides valuable insights for researchers and clinicians aiming to optimize scaffold design, MSC integration, and translational applications. These insights could significantly influence future research directions and the development of more effective regenerative therapies.

Keywords: differentiation; electrospinning; mesenchymal stem cells; nanofibrous scaffolds.

Figure 2

(a–d) Immunofluorescent staining of differentiated hMSCs for paxillin (green), F-actin (red), and nucleus (blue) in the presence of cytoskeletal inhibitors on PCL-A nanofibers. (a) Control; (b) Cells treated with F-actin inhibitor, cytochalasin D; (c) Cells treated with nocodazole; (d) Cells treated with Y-27632, a ROCK inhibitor. In panels (c,d), the cells adopt a tetragonal morphology and show a decrease in paxillin expression. Images obtained by [19]. (e) Random orientation of cells on a nonaligned nanofiber; (f) Well-oriented stem cells morphology on an aligned nanofiber matrix at day 4. Arrow indicates the MSCs aligned along fibers. Images obtained by [20]. (g–j) SEM micrographs of MSCs after differentiation to cardiomyocytes in PVA-CS-CNT3 (g,h) and PVA-CS-CNT5 (i,j) scaffolds after 14 days of exposure to electrical stimulation and the differentiation medium. Images obtained by [23]. (k,l) PU electrospun nanofibers and cardiomyocyte-like cells on (k) NGF (–) and (l) NGF (+) PU. Images obtained by [34].

Figure 3

Morphology of (a) H-keratino/MSC on day 1; (b) day 14; and (c) day 21 on nylon and N-B.v NFM. Images obtained by [37]. (d–f) SEM photograph of MSC-derived keratinocyte attachment on fiber mats after 84 h of seeding. Images obtained by [38]. Live/dead imaging of BM-MSCs cultured in induction media at various time points: (g) 3 days; (h) 7 days; and (i) 14 days of culture. Red arrows highlight the morphological changes in BM-MSCs upon induction with epidermal differentiation media. (j–l) Immunostaining of CD105 and involucrin in epidermally differentiated BM-MSCs on PHBV nanofibers. Images obtained by [40].

Figure 5

(a) SEM images showing cell adhesion within 4 h of culture and fluorescent microscopy images of actin and nucleus-stained cells after 24 h of culture on plasma-treated and untreated aligned PCL/collagen multiscale fibers (PC). The scale bar for SEM images is 10 μm, and for fluorescent microscopy images, it is 50 μm. Images sourced from [67]. (b) (1, 2) Representative fluorescent images showing living cells (green) and dead cells (red) of HADMSC seeded on PLA MY bundles and PLGA/PLA HY bundles conditioned in TM for 21 days. (3, 4) Immunofluorescence (IF) staining for TNMD (green) and nuclei (blue) of HADMSC after 21 days of tenogenic differentiation. (5, 6) IF staining for COL1 (green) and nuclei (blue) of HADMSC after 21 days of tenogenic differentiation. Scale bar: 100 μm. Images sourced from [70].

Figure 1

A systematic search conducted using PRISMA 2020 flow diagram for updated systematic reviews, which included searches of databases and registers only. The search query (Nanofibers) and (Electrospinning) and (Mesenchymal stem cells) and (Differentiation) was used on PubMed and Web of Science over the last 10 years (2014–2024). A total number of 666 articles found in the last 10 years (2014–2024) were registered. After a systematic process of applying exclusion and inclusion criteria, 91 studies were included in this review.

Figure 4

(a–c) Immunocytochemistry assay results for bladder myosin heavy chain (MHC) protein in human adipose-derived mesenchymal stem cells (AT-MSCs) cultured on (a) tissue culture polystyrene (TCPS); (b) poly lactic-co-glycolic acid (PLGA)/polyurethane (PU); (c) poly-phosphate (poly-P)-incorporated PLGA/PU nanofibrous scaffold (PLGA/PU/poly-P). Images obtained by [51]. (d,e) SEM images of human AT-MSCs seeded on electrospun PAN-PEO nanofibers. Immunocytochemistry assay of (f) bladder myosin heavy chain (MHC) and (g) α-SMA protein in smooth muscle cell (SMC)-differentiated human AT-MSCs grown on PAN-PEO nanofibers. Images obtained by [52].

Figure 6

(a,b) Live/dead staining was performed to assess the viability of MSCs cultured on pectin nanofiber scaffolds with varying oxidation levels over a 14-day period. MSCs were cultured with TGFβ1 medium on (a) OCP25-ADH and (b) OCP50-ADH scaffolds to evaluate cell viability and proliferation, respectively. Images obtained by [82]. Scanning electron microscopy images of the PCL scaffold (c) without cells and (d) with cells. (e) An overview image showing cell coverage and alignment across the entire length of the perfused sample. Images obtained by [83].