Background
Diabetic ulcer is one of the major complications of diabetes that accounts for about 36% of all chronic skin ulcers, and the amputation rate is up to 19.03% in individuals with diabetic ulcers [
1]. A chronic diabetic ulcer is characterized by complex etiology, prolonged treatment duration, high medical expense, predisposition to repeated recurrence, and poor clinical prognosis. Therefore, to improve the quality of patient’s life, identifying the effective therapeutic measure for diabetic chronic skin ulcer wound is essential [
2].
Mesenchymal stem cells (MSCs) are adult stem cells with the potential of multilineage differentiation; MSCs are primarily derived from the bone marrow, fat, and umbilical blood and cord [
3,
4]. Human umbilical cord mesenchymal stem cells (hUCMSCs) possess characteristics of robust proliferation and differentiation, as well as weak immunogenicity, thereby posing a great potential in the field of tissue engineering [
5]. To improve the survival ratio of hUCMSCs on the wound surface, we implanted hUCMSCs on the scaffold made of the collagen–chitosan laser drilling acellular dermal matrix (CCLDADM), which was then spread on the wound surface, to achieve the optimal therapeutic effect on the wound repair [
6]. Therefore, whether this method could be feasible for the management of refractory diabetic ulcer wound necessitates further exploration.
Wnt signaling pathway is critical for the regulation of cell proliferation and differentiation, as well as posttraumatic tissue regeneration and repair [
7]. A recent study reported that Wnt signal was attenuated in the skin wound of diabetic rat model, and its suppression slowed down the healing speed, thereby implying Wnt signaling pathway as a vital element for the regulation of diabetic skin ulcer [
8]. Therefore, the current study deployed the hUCMSCs-loaded scaffold for treatment of diabetic skin ulcer wound and explored the activation of Wnt signaling pathway in hUCMSCs seeding on the scaffold to accelerate wound healing.
Discussion
Diabetes mellitus is the leading chronic disease posing risks to human health. It not only compromises the quality of patient’s life but also gives rise to an economic burden on society and families [
2]. The primary manifestation of diabetic foot is chronic skin ulcers on lower limbs, and a subset of patients with chronic ulcers require amputation; refractory diabetic ulcers constitutes 50–70% of all amputations [
13]. Hitherto, the mechanism underlying the incurable skin wound in diabetic patients remains unclear; however, some theories propose the involvement of neuropathy, vascular remodeling, inflammatory reaction, and metabolic abnormality of the local wound [
14]. In addition to the conventional preventive measures, stem cell therapy is gradually emerging in the management of chronic diabetic ulcers [
15‐
17]. Previous studies displayed that MSCs promoted the healing of mouse cutaneous wound [
18,
19]. Nonetheless, MSCs have rarely been reported to be successful on humans. This failure might be attributed to the following: (1) intravenous perfusion of MSCs pose great safety risks; (2) when MSCs are injected subcutaneously around the wound area, differentiated MSCs cannot migrate easily to the wound; (3) only a small portion of MSCs cells smeared on the wound can attach and survive [
20,
21]. In the previous study, we adopted CCLDADM as a scaffold on skin wound to enhance stable proliferation of hUCMSCs and capillary regeneration at the wound site [
6]. However, this method on diabetic ulcers still faces considerable challenges due to incurable diabetic wounds.
Furthermore, Wnt signaling pathway is highly conserved across evolution, suggesting its essential role in embryonic development and stem cell growth and differentiation [
22]. β-Catenin is the canonical effector molecule in the Wnt signaling pathway [
23]. Wnt protein, when bound to its membrane receptors, such as FZD and LRP5/6, elicits the dissociation of β-catenin from the degradation complex, thereby attenuating its phosphorylation catalyzed by GSK3β and CK1 and diminishing the subsequent ubiquitination. Also, the activation of Wnt signaling pathway enhances the nuclear translocation of β-catenin, which then regulates the expression of target genes, such as
cyclin-
D1,
c-
Myc,
p63,
CK19, and
PCNA, which regulate the cell proliferation and differentiation [
23]. The binding of Wnt protein to its receptors is modulated by a variety of molecules: sFRPs and Wnt inhibitory factor (WIF-1). These two proteins can competitively inhibit the binding of Wnt3a to FZD via interaction with FZD, thereby restraining the stimulation of Wnt signal [
22]. The abnormal Wnt signal is closely associated with multiple diseases, such as malignancies, liver fibrosis, rheumatoid arthritis, bone development, and immune response [
24,
25]. The role of Wnt signaling pathway in above diseases is closely related to the regulation of cell proliferation and differentiation [
25]. Previous studies reported the attenuation of Wnt signal in diabetic nephropathy and diabetic ulcer wounds, implying its role in the pathogenesis of the two diseases [
8,
26]. To further explore whether Wnt pathway could modulate the therapeutic effect of hUCMSCs-loading matrix scaffold material for diabetic ulcers, the present study applied hUCMSCs-seeding CCLDADM scaffold on the skin ulcer wounds of both normal and diabetic rats. Moreover, Wnt signal agonist or antagonist was administered at the scaffold edge. The results showed that the healing rate of skin wound was significantly lower in diabetic rats than that in the normal control, along with a reduction in protein levels of cyclin-D1, c-Myc, p63, CK19, and PCNA, indicating attenuated Wnt signaling pathway [
27]. In addition, we found that the healing of skin wound was significantly accelerated in diabetic rats, following the activation of the Wnt signaling pathway, while it significantly slowed down post-suppression of the signal. The results of HE staining demonstrated that the activated pathway raised the abundance of new blood vessels and integumentary appendages in the granulation tissue, further proving that the activation of Wnt signaling pathway could enhance the therapeutic effect of the CCLDADM scaffold on diabetic ulcers.
In the present study, Wnt3a and sFRP were injected at the scaffold edge, rendering the possibility that Wnt3a and sFRP might exert the biological effect through rat innate somatic cells at the wound edge. To validate that Wnt3a and sFRP could regulate the hUCMSCs proliferation and differentiation seeding on the CCLDADM scaffold, the hUCMSCs-seeding scaffold was cultured in vitro and stimulated with Wnt3a and sFRP. The results showed that Wnt3a could promote and sFRP could suppress the proliferation and differentiation of hUCMSCs, and Wnt3a decreased the hUCMSCs death, thereby increasing the survival number of hUCMSCs on the CCLDADM scaffold.
Thus, hUCMSCs combined with CCLDADM scaffold for the treatment of refractory diabetic ulcers exhibit several advantages, such as potent differentiation of hUCMSCs and abundant source of umbilical cord without ethical issues. Furthermore, increasing development in material science proposes multiple options for engineered skin materials. The key study orientations for the next step focus on the selection of the scaffold material, improvement of the proliferation and differentiation efficiency of hUCMSCs seeded on the engineered skin, and the safety of the usage of hUCMSCs in humans.
Authors’ contributions
YH and TS carried out the literature search and drafted the manuscript. RT and GY reviewed the manuscript and corrected language and grammar. LL, YH and CL coordinated and helped to draft the manuscript. JL served as the final internal reviewer. All authors read and approved the final manuscript.