Promotion of adipose stem cell transplantation using GelMA hydrogel reinforced by PLCL/ADM short nanofibers

Adipose-derived mesenchymal stem cells (ADSCs) show poor survival after transplantation, limiting their clinical application. In this study, a series of poly(l-lactide-co-ϵ-caprolactone) (PLCL)/acellular dermal matrix (ADM) nanofiber scaffolds with different proportions were prepared by electrospinning. By studying their morphology, hydrophilicity, tensile mechanics, and biocompatibility, PLCL/ADM nanofiber scaffolds with the best composition ratio (PLCL:ADM = 7:3) were selected to prepare short nanofibers. And based on this, injectable gelatin methacryloyl (GelMA) hydrogel loaded with PLCL/ADM short nanofibers (GelMA-Fibers) was constructed as a transplantation vector of ADSCs. ADSCs and GelMA-Fibers were co-cultured, and the optimal loading concentration of PLCL/ADM nanofibers was investigated by cell proliferation assay, live/dead cell staining, and cytoskeleton staining in vitro. In vivo investigations were also performed by H&E staining, Oil red O staining, and TUNEL staining, and the survival and apoptosis rates of ADSCs transplanted in vivo were analyzed. It was demonstrated that GelMA-Fibers could effectively promote the proliferation of ADSCs in vitro. Most importantly, GelMA-Fibers increased the survival rate of ADSCs transplantation and decreased their apoptosis rate within 14 d. In conclusion, the constructed GelMA-Fibers would provide new ideas and options for stem cell tissue engineering and stem cell-based clinical therapies.


Introduction
Stem cell-related tissue engineering is considered one of the most influential and promising fields in life sciences [1]. Adipose mesenchymal stem cells (ADSCs) have abundant sources and can selfrenewal multidirectional differentiation and paracrine secretion. The transplantation of ADSCs has become the focus of many preclinical and clinical cell therapy studies [2]. It has been demonstrated to promote wound healing [3], neurogenesis [4], liver regeneration [5], and treat arteriovenous fistula [6,7], retinal degenerative diseases [8], autoimmune encephalomyelitis [9], articular cartilage defects [10] and other diseases [11][12][13][14][15][16][17]. However, with stem cell transplantation alone, cells float due to the lack of adhesion sites, resulting in relatively low retention and survival rates of transplanted ADSCs [1,18,19]. In addition, the inadequate blood supply, premature differentiation, and apoptosis after transplantation limit the application of ADSCs transplantation. Therefore, creating an environment that allows the growth and adhesion of ADSCs has become an urgent problem, promotes rapid revascularization at the transplantation site, and inhibits the differentiation and apoptosis of transplanted ADSCs to improve the survival rate.
The microenvironment surrounding stem cells is critical in determining their survival, retention, and therapeutic potential. Gelatin methacryloyl (GelMA) hydrogel is a popular biodegradable hydrogel [20,21], which has a three-dimensional structure suitable for cell growth and adherence that mimics the natural extracellular matrix [22,23]. Therefore, the injectable GelMA has been used as a vehicle for stem cell delivery in tissue engineering [24][25][26][27] to provide an adhesive growth scaffold for the initial stage of stem cell transplantation. Compared with open surgery as an administration route, minimally invasive surgery reduces the complications and surgical recovery time effectively [28]. However, hydrogels generally exhibit poor mechanical properties and lack fibrous structures providing suitable support for cellular activity. Studies have shown that adding electrospun short fibers into hydrogels as reinforcements can improve hydrogels' mechanical properties and biological function, reinforce fixation and promote cell proliferation [29,30]. Acellular dermal matrix (ADM) preserves the extracellular stroma and threedimensional structure of the dermal tissue, shows low immunogenicity and high biocompatibility, promotes cell proliferation and revascularization, which has been widely used to promote wound healing [31][32][33]. However, ADM has poor mechanical properties and fast degradation [34], which is a disadvantage for homogenizing into short fibers and being applied in vivo. To overcome these problems we propose to use poly(l-lactide-co-ε-caprolactone) (PLCL) to prepare the short fibers. PLCL is obtained from the polymerization reaction of L-lactic (LA) and εcaprolactone (CL), which has high toughness and fast degradation rate [35]. Single needle type electrostatic spinning PLCL/ADM composite nanofibers were prepared into short nanofibers by high-speed homogenization. Compared with core/shell fibers, PLCL/ADM composite short fibers can possess faster degradation efficiency [36], rapid release of active ingredients, and improved microenvironment of transplanted stem cells. Moreover, the preparation process of singleneedle spinning is more straightforward, and the preparation cost is low.
Short nanofibers have been combined with GelMA for tissue engineering. Dubey incorporated antibiotic-eluting fiber-based microparticles in GelMA hydrogel to gather antimicrobial and angiogenic properties [37]. Gelatin methacrylate/thiolated pectin hydrogels carrying electrospun core/shell fibers of melatoninpolymethylmethacrylate (PMMA)/Tideglusib-silk fibroin were designed as an injectable hydrogel for vital pulp regeneration [38]. An injectable and photo cross-linkable gelatin methacryloyl hydrogel was chosen with ciprofloxacin-eluting short nanofibers for oral infection ablation [39]. However, none of the above studies have explored stem cell transplantation. In addition, Kim developed a multi-spheroid-loaded thin-sectioned ADM carrier [40]. Nevertheless, this stem cell transplantation system requires surgical implantation into the animal, a non-injectable stem cell transplantation system that can cause unnecessary harm to the recipient.
In this study, a series of PLCL/ADM nanofiber scaffolds with different composition ratios were prepared by electrospinning. After comprehensive characterization, including morphology, hydrophilicity, tensile mechanics, and biocompatibility, nanofiber scaffolds with optimal composition ratio were chosen and homogenized into short nanofibers. And the injectable GelMA hydrogel loaded with PLCL/ADM short nanofibers (GelMA-Fibers) was constructed as a transplantation vector for ADSCs. This strategy improves the survival rate of ADSCs transplantation, facilitates the continued biological role of ADSCs in the transplantation recipient site, and provides a new idea and option for clinical treatment.

Preparation of the PLCL/ADM short nanofibers
The obtained PLCL/ADM nanofiber scaffolds were cut into 3 × 3 cm 2 pieces and dispersed in deionized water. The homogenized short nanofiber dispersions were fabricated using high-speed homogenization (FSH-2 A) at 10 000 rpm for 30 min. The dispersions were placed in a refrigerator at −80 • C overnight. After 48 h of freeze-drying, dried PLCL/ADM short nanofibers were obtained. The short nanofibers were then sterilized by fumigation with 75% ethanol for 6 h and UV irradiation for 30 min.

Characterization
The morphology of PLCL/ADM nanofiber scaffolds was observed by scanning electron microscope (SEM, TM-1000, Hitachi). One hundred fibers on SEM photographs were randomly selected, and the average diameter of the fibers was measured using ImageJ software. The hydrophilic and hydrophobic properties of PLCL/ADM nanofiber scaffolds were examined using a contact angle analyzer (WCA, SA100). The nanofiber scaffolds were characterized using Fourier transform infrared spectroscopy (FTIR, Nicolet Is50) in the 4000-400 cm −1 with a resolution of 4 cm −1 . Tensile tests were performed on samples of 50 mm in length and 10 mm in width (sample thickness was measured by a spiral micrometer) using a microcomputer-controlled electronic universal testing machine (WDW-5 G, Jinan Hengsi Shengda Co., Ltd China). The scaffolds were tested at a tensile speed of 20 mm min −1 until fracture. For each scaffold, three samples were tested. The viscosity-shear rate of GelMA hydrogel and GelMA + Fibers was tested by a TA DHR-2 rheometer (TA Instruments, United States).

In vitro cytocompatibility of the PLCL/ADM nanofiber scaffolds
Human waste adipose tissue samples were derived from the liposuction operations of one healthy human specimen after excluding other diseases. The present research was allowed by the Medical Ethics Committee of the Affiliated Hospital of Qingdao University (QYFYWZLL27421). The isolation and culture of human ADSCs were performed as previously described [23]. Human adipose tissue was digested in collagenase type I solution (C8140, Solarbio) at 37 • C for 45 min with agitation. The samples were filtered with 200 µm mesh filters and centrifugated at 1200 rpm for 12 min to separate the stromal cell pellet from adipocytes. Cell pellets were resuspended in Minimum Essential Medium (MEM, Gibco) containing 10% FBS and 1% penicillin/streptomycin (Solarbio) and cultured at 37 • C with 5% CO 2 . After the ADSCs were attached, the medium was replaced every 2 d. After 7 d of incubation, the ADSCs were passaged at 90% confluence. ADSCs were seeded on the PLCL/ADM nanofiber scaffolds (10:0, 7:3, 5:5, 3:7, and 0:10) (w/w) at 8 × 10 3 cells/well density. As the control group, ADSCs were seeded on the tissue culture plates (TCP). Cell proliferation was investigated after 1, 3, 5, and 7 d of culture using Cell Counting Kit-8 (CCK-8, NCM). Briefly, the medium was replaced by 10% CCK-8 solution and incubated for 2 h at 37 • C protected from light, after which the absorbance at 450 nm was measured using a microplate reader (Spectramax ABS, Molecular Devices). After 3 d of cultivation, the F-actin cytoskeleton fluorescence staining was performed. Cells were first fixed with 4% paraformaldehyde (PFA, Biosharp) for 10 min, then permeabilized with Phosphate buffer saline (PBS) containing 0.1% Triton X-100 (9002-93-1, Solarbio) for 20 min and blocked with 4% bovine serum albumin (BSA, 9048-46-8, Solarbio) in PBS for 1 h at room temperature. After blocking, the F-actin cytoskeleton was stained with FITCconjugated phalloidin (1:1000, Abcam) diluted in PBS with 4% BSA for 40 min in the dark. Finally, each sample was washed with PBS and sealed with an antifading mounting medium containing DAPI (S2110, Solarbio). The cytoskeleton and morphology of the ADSCs were observed under a fluorescence microscope (Nikon A1MP, Japan). Three parallel samples were set up for each group in the above experiments.

In vitro cytocompatibility of the GelMA-fibers
ADSCs were blended with the GelMA + (0%, 2%, 4%, 6%) Fiber prepolymers at 1 × 10 6 cells/mL density. 200 µl GelMA + (0%, 2%, 4%, 6%) Fibers prepolymers were added per well in 24-well plates and were irradiated with the UV light (405 nm) for 14 s to obtain the photo-curable hydrogels. In this way, ADSCs were cultured in three dimensions within GelMA-Fibers. The ADSCs were cultured in the GelMA-Fibers in MEM with 10% FBS and 1% penicillin/streptomycin. After 1, 3, and 5 d of culture, cell proliferation was investigated using the CCK-8 kit as described above. In addition, after 3 d of culture, live/dead assays and cell F-actin cytoskeleton fluorescence staining were performed. Cell F-actin cytoskeleton fluorescence staining was performed as described previously. Live/dead staining was operated using the live/dead cytotoxicity kit (EFL-CLD-001) according to the manufacturer's instructions. Briefly, cells were incubated with PI working solution for 10 min and then incubated with AM working solution for 30 min. Live and dead cells were stained with AM (green) and PI (red), respectively. Cells were imaged under an inverted fluorescence microscope (IX53, OLYMPUS, Japan). Three parallel samples were set up for each group in the above experiments.

In vivo investigation
To reduce the rejection reaction, rat ADSCs (ADRS-C106, HyCyte) were selected to replace the human ADSCs used in the cell experiments. Rat ADSCs were fluorescently labeled with CM-DiI (C7000, Invitrogen) before injection. CM-DiI was diluted to 1 mg ml −1 in dimethyl sulfoxide (DMSO, Solarbio) and further to 1 µg ml −1 in D-PBS (Solarbio). ADSCs were incubated with CM-DiI for 5 min at 37 • C and then protected from light for 20 min at 4 • C. Excess dye was removed by PBS washing, and ADSCs were digested and centrifuged to obtain ADSCs precipitate. Healthy male Sprague-Dawley (SD) rats (6 weeks, 180-220 g) were purchased from Jinan Pengyue Experimental Animal Breeding Co., Ltd (Shandong, China). All animal procedures were performed in strict accordance with the regulations of the Institutional Animal Care and Use Committee of China and approved by the ethical review committees of Affiliated Hospital of Qingdao University. Three injection sites were identified on the back of each SD rat, and the injection sites were randomly divided into ADCSs + PBS group (Control group), ADCSs + GelMA group, and ADSCs + GelMA + 6%Fibers group (n = 5). First, the SD rats were anesthetized, and the back of each rat was shaved and sterilized. Before injection, ADSCs precipitates were resuspended in 200 µl PBS, 1 ml GelMA prepolymers, and 1 ml GelMA + 6%Fibers prepolymers at a concentration of 1 × 10 6 cells, respectively. Prepolymers were cross-linked with the UV light at 405 nm for 14 s. Finally, the ADCSs mixture was injected into the dorsal intradermal mesoderm of rats. A distinct dermatome elevation was visible, marking the extent of the dermatome. Rats were sacrificed on days 7 and 14 after injection, and the whole skin and intradermal graft tissue within the marked area were excised for H&E staining, oil red O staining, and TUNEL staining. Nine rats were operated on at each time point. Oil red staining was performed using fresh tissues (n = 3). H&E staining (n = 3) and TUNEL staining (protected from light, n = 3) were performed using tissues fixed with 4% paraformaldehyde. The cells were quantified using ImageJ software. The transplantation survival rate of the ADSCs was calculated according to the formula (n = 5):

Statistical analysis
Origin 6.0 and Graphpad Prism software were used for statistical analysis of all data. One-way ANOVA was used to observe the significance of differences between groups. All pairwise comparisons were performed using the Student's t-test. Results were considered statistically significant at * P < 0.05, * * P < 0.01, and * * * P < 0.001.

Characterization of PLCL/ADM nanofiber scaffolds
Surface morphology is one of the most important characteristics of electrospun materials [48]. The quality of short nanofibers is directly affected by the uniformity of the diameter of electrospun nanofibers. SEM images of PLCL/ADM nanofibers were presented in figure 1(A). The nanofibers of each group were almost uniform in morphology, without beads, and in a randomly stacked state. The fibers adhere to each other and form a porous mesh-like structure similar to a 'spider web' . Especially the PLCL/ADM composited groups had ideal and uniform fiber diameters ( figure S2). The chemical characterization of the PLCL/ADM nanofiber scaffolds was confirmed by FTIR spectroscopy analysis ( figure 1(B)). FTIR spectrum showed a strong C=O stretching peak at approximately 1639 cm −1 and an N-H stretching peak at 3295 cm −1 , indicating that ADM contains a significant amount of collagen [49]. Moreover, the above two absorption peaks vibrated more obviously with increased ADM content, indicating that PLCL was successfully doped with ADM. In addition, the ADM contents showed a gradient variation, which was consistent with the experimental design. The contact angle presented information about the hydrophilicity of fiber surfaces. As shown in figure 1(C And the hydrophobicity is beneficial to maintain the stability and intact morphology of the short nanofiber scaffolds.  The toughness and elasticity of the scaffolds are essential to maintain the relative structural integrity during the homogenization [50]. Injectable GelMA requires structural support from short nanofibers to improve the overall structural stability of GelMA. Therefore, to explore the mechanical properties of PLCL/ADM nanofibers, we investigated the effects of different composition ratios on 8.12 ± 0.16 MPa, 6.17 ± 0.69 MPa, 6.49 ± 0.71 MPa, and 4.87 ± 0.65 MPa, respectively, indicating that the PLCL and PLCL:ADM = 7:3 group had better strength and could resist relatively high stresses, which was statistically significant compared with the ADM group. Besides, the elongation at the break (figure 2(C)) of the five groups were 424.59 ± 10.02%, 289.30 ± 7.65%, 172.78 ± 8.64%, 67.635 ± 15.11% and 27.49 ± 0.36%, respectively. The elongation at the break of the fibers was higher with increasing PLCL content, and the PLCL, PLCL:ADM = 7:3, and PLCL:ADM = 5:5 groups showed significant differences compared to the ADM group. The Young's modulus (figure 2(D)) of the five groups were 6.16 ± 1.75 MPa, 5.42 ± 1.60 MPa, 45.43 ± 3.28 MPa, 142.57 ± 7.66 MPa, and 88.08 ± 10.20 MPa, respectively. The Young's modulus of PLCL and PLCL:ADM = 7:3 group was lower than the other groups, and the results were statistically different. The PLCL:ADM = 7:3 group had higher mechanical strength, better elongation at break, and the lowest Young's modulus among the composite nanofibers. The better the mechanical properties of the nanofibers, the more they can give stronger mechanical support to the hydrogel after homogenizing into short nanofibers. Therefore, it can prevent the graft system from being squeezed by the tissue and spread to the non-graft area, providing a stable attachment structure to the stem cells and facilitating their growth and proliferation.

In vitro cytocompatibility of the PLCL/ADM nanofiber scaffolds
Biocompatibility is one of the most important properties and basic essential characteristics of biomedical materials [51]. ADSCs were cultured on PLCL/ADM nanofibers with different composition ratios, with TCP as a control group. As shown in figure 3, the cell proliferation rate in the three PLCL/ADM nanofiber scaffolds groups was significantly higher than that of the PLCL and ADM groups on days 5. This is because the collagen and laminin contained in PLCL/ADM nanofiber scaffolds could effectively promote the rapid proliferation of ADSCs compared with PLCL nanofibers. However, due to the strong hydrophilicity and easy dissolution of ADM nanofibers in water, they could not maintain a good nanofiber structure, resulting in ADSCs being unable to attach well to ADM nanofibers. Therefore, the cell proliferation rate in the ADM nanofibers group was not higher than in the PLCL/ADM nanofiber scaffold groups. It could be seen that the PLCL/ADM nanofiber scaffolds had excellent biocompatibility and facilitated cell survival and proliferation. The morphology and fluorescence images of ADSCs on day 3 were shown in figure 4. The morphology of ADSCs was observed by fluorescence microscopy on day 3 and day 5, respectively. The average area of each cell in different groups was measured ( figure S3). The results showed that the cytoskeleton of ADSCs was extended to varied degrees in different groups. Cells in all experimental groups except the ADM group showed good adhesion and cytoskeleton extensibility. The nanofibrous scaffold with the composition ratio of PLCL:ADM = 7:3 was the most conducive to cell attachment and extension, significantly different from the other groups. Almost all cells exhibited a long shuttle cell shape similar to normal ADSCs. No significant differences were observed in the growth pattern of cells in the PLCL/ADM group. Since both mechanical properties Figure 5. CCK-8 results of ADSCs co-cultured with GelMA-Fibers at different short nanofiber loading concentrations (0%, 2%, 4%, and 6%). * P < 0.05, * * P < 0.01, * * * P < 0.001. and biocompatibility need to be considered in the design and preparation of biomaterials, nanofibers with a composition ratio of PLCL:ADM = 7:3 were chosen to prepare short nanofibers in subsequent experiments.

GelMA-Fibers promoted the proliferation of ADSCs in vitro
ADSCs were co-cultured with GelMA-Fibers for 3 d. As shown in figure 5, the best proliferation activity of ADSCs was observed in the GelMA + 6%Fibers group. As the number of loaded nanofibers in GelMA hydrogel decreased, the number of ADSCs decreased accordingly, and there was a significant difference between the groups. The results of live/dead staining were shown in figure 6. The red fluorescence (dead cells) was rarely found, indicating that GelMA-Fibers was not toxic to the cells. The green fluorescence (living cells) could cover the entire field of view, indicating the cells' good survival and proliferation status. It noted that GelMA-Fibers had good biocompatibility with cells. In addition, the GelMA + 6%Fibers group showed the best effect on cell proliferation. Therefore, GelMA loaded with 6% short nanofibers was selected for subsequent experiments. As shown in figure 7, ADSCs showed regular morphology, good extension, and long spindle shape by the third day of co-culture with GelMA-Fibers, and there was no significant difference between them and the control group ( figure S4). Therefore, it can be concluded that injectable GelMA hydrogel's biocompatibility will improve with the increase in the number of loaded short nanofibers, provided that injectability is guaranteed. The shear-thinning test has been tested, and the result was in figure S5. The development of the shear-thinning test showed that GelMA hydrogel and GelMA + 6%Fibers exhibited a shearthinning characteristic, which meant that a syringe could inject them. Figure S5 also reflects another perspective: GelMA + 6%Fibers have stronger mechanical properties than GelMA hydrogel. Therefore, injectable GelMA hydrogel loaded with 6% PLCL/ADM short nanofibers was selected for animal experiments.

GelMA-Fibers enhanced the survival rate of ADSCs in vivo
ADSCs transplanted display meager survival rates in severely hostile microenvironments, such as ischemic conditions [52], which severely undermine the therapeutic effect [53]. Providing a more prosperous blood supply to transplanted tissues early in stem cell transplantation will undoubtedly improve the efficiency of stem cell transplantation significantly. H&E staining was used to observe the histological changes ( figure 8(A)). After 7 d, there were no considerable differences in the vascularization of the surrounding tissues between the GelMA group and the GelMA + 6%Fibers group. While after 14 d, the number of blood vessels increased in the surrounding tissues of the GelMA + 6%Fibers group. The vascularization of the GelMA + 6%Fibers group was preferable to the GelMA group. In addition, ADSCs were located inside GelMA hydrogel, relatively stable, and heavily survived, while the transplanted tissues in the control group were free under the skin of rats and could not be precisely localized.
After 14 d, there were a large number of adipocyte cavities in and around the transplantation site of the control group, indicating that ADSCs differentiated into adipocytes quickly. The oil red O staining results on the 14th day of the transplanted tissue were shown in figure 8(B). ADSCs in GelMA and GelMA + 6%Fibers groups began to differentiate into adipocytes. Lipid droplets in the cytoplasm of adipocytes were tiny, so the adipocytes in the GelMA and GelMA + 6%Fibers groups could be considered newly differentiated adipocytes. Diffuse adipocytes could be found in the subcutaneous tissue of the graft area in the control group, and the lipid droplets in the cytoplasm were generally fuller. Besides, the adipocytes were more significant than those in the GelMA and GelMA + 6%Fibers groups. It is reasonable to speculate that without the protection of GelMA as   a cell carrier, the ADSCs in the control group differentiated into adipocytes earlier and lost stem cell function.
CM-DiI is a long-chain carbocyanine membrane probe that does not transfer from labeled to unlabeled cells [54,55]. The photostable fluorescence, excellent cellular retention, and minimal cytotoxicity of CM-DiI make it particularly suitable for long-term labeling and tracking of cells [56][57][58]. Therefore, in vivo experiments were conducted using CM-DiI live cell tracer labeled ADSCs to determine the survival rate after transplanting into rats. The examination was performed using fluorescence microscopy on postoperative days 7 and 14 (figure 9), and the survival rate of ADSCs was calculated ( figure 10(A)). Meanwhile, TUNEL staining was performed on the tissue sections (figure 9) to obtain the apoptosis rate of the transplanted cells ( figure 10(B)). A large number of CM-DiI labeled cells with red fluorescence could be found in both GelMA + 6%Fibers and GelMA groups, and these cells were evenly distributed in the field of view. The red fluorescence in the control group was lower than in the other groups, showing a significant difference. Therefore, it could be concluded that GelMA hydrogel can provide the transplanted stem cells with stable structural protection and prevent them from wandering and spreading to the non-transplanted area. In addition, the survival rate of ADSCs in the GelMA + 6%Fibers group was significantly higher than that in the GelMA group, indicating that PLCL/ADM short nanofibers loading could improve the survival rate of ADSCs transplantation in vivo. In a previous study, Li et al applied advanced glycation end products to ADSCs and introduced the siRNA of the advanced glycation end products receptor into ADSCs. The treated ADSCs were injected into the wounds of diabetic mice to promote wound healing, and less than 20% of the ADSCs survived after 7 d [59]. In this study, the ADSCs in the GelMA + 6%Fibers group achieved a survival rate of 69.44 ± 2.98% on 14 d, while GelMA provided a relatively favorable microenvironment for the transplantation of ADSCs and reduced the apoptosis rate.

Conclusion
In this study, PLCL/ADM nanofiber scaffolds with different proportions were prepared by electrospinning to investigate their tensile mechanics and effects on the proliferation of ADSCs. It was confirmed that PLCL:ADM = 7:3 nanofiber scaffolds had excellent mechanical properties and biocompatibility and were used to prepare short nanofibers. Injectable GelMA hydrogel loaded with PLCL/ADM short nanofibers was constructed as the transplantation vector of ADSCs. It was demonstrated that GelMA-Fibers promoted the proliferation of ADSCs in vitro. In vivo investigation was also carried out by H&E staining, Oil red O staining, and TUNEL staining. The survival rate and apoptosis rate of ADSCs in vivo transplantation were analyzed. It was proved that, as a transplantation vector, GelMA-Fibers provided a living environment conducive to adhesion and growth for stem cells at the early stage of transplantation. In addition, GelMA-Fibers also promoted the vascularization of tissues around transplantation and prevented stem cells from premature differentiation and dissociation to the non-transplant area. Most importantly, GelMA-Fibers can improve the survival rate and reduce the apoptosis rate of ADSCs grafts within 14 d. In conclusion, GelMA-Fibers would provide new ideas and options for stem cell tissue engineering and clinical treatment based on stem cells.

Data availability statement
All data that support the findings of this study are included within the article (and any supplementary files). The application of stem cell therapy and brown adipose tissue transplantation in metabolic disorders