Abstract
Cesarean section has been increasingly used, stem cells have a potential to improve the repairing of scarred uteri, but its mechanism is still unclear. Cell labeling and tracking are becoming increasingly important in stem cell research. In this study, we evaluated the safety and efficacy of labeling human Umbilical Cord Mesenchymal Stem Cells (hUCMSCs) with Superparamagnetic Iron Oxide Nanoparticles (SPIONs), both in vitro and in vivo, with a particular focus on their application as Magnetic Resonance Imaging (MRI) contrast agents injected into the uterus. Monkeys were used as animal models because their structure is similar to that of humans. These results demonstrate that hUCMSCs could be effectively labeled with SPIONs without compromising cellular proliferation or differentiation potential. Furthermore, a minimal impact on cell surface marker expression and immunocytochemical staining for vimentin was observed. MR scanning was conducted both in vitro and in vivo, along with pathological staining, to confirm the effective labeling of SPIONs without significant cytotoxicity. This labeling persisted for more than 600 days, enabling non-invasive, dynamic monitoring of the injection site via MRI. The safety and effective labeling of human Umbilical Cord Mesenchymal Stem Cells (hUCMSCs) with Superparamagnetic Iron Oxide Nanoparticles (SPIONs) for MRI contrast enhancement in uterine repair were conducted in this study. Our findings show no adverse effects on cell proliferation or differentiation, with sustained SPION labeling over 600 days. These findings could help improve scarred uterus regeneration, offering a non-invasive tracking method for stem cell therapies in regenerative medicine.
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Introduction
Cesarean section is the most frequently performed surgery in obstetrics and gynecology. Currently, cesarean section rates in the United States are approximately 20–30%, and in China, it is around 41%1. This rate is expected to increase further owing to China’s implementation of the two-child policy2. On one hand, cesarean section plays a crucial role in addressing dystocia, managing severe pregnancy-related conditions, and safeguarding the lives of mothers and infants; On the other hand, cesarean section inflicts full-thickness damage to the uterus, with 60%−70% of uterine incision scars exhibiting healing defects postoperatively3. The risk of postpartum complications, including hemorrhage, infection, and pelvic adhesions, is significantly elevated compared with vaginal delivery. Additionally, cesarean sections result in a significant number of women with scarred uteri. The presence of a scarred uterus significantly increases the risk of high-risk pregnancies and their associated complications such as placenta previa and uterine rupture. Therefore, it is crucial to repair the scarred uteri.
Cesarean section-induced full-layer uterine injuries encompass injuries to the serous, myometrial, and endometrial layers. Although cells in the serous and endometrial layers can regenerate and undergo functional repair, regenerating the uterine muscle layer is challenging. Typically, this layer transforms into fibrous connective tissue and undergoes fibrous repair rather than regenerative repair, limiting the restoration of full function. The recent emergence of stem cell technologies has offered promising avenues to address this challenge.
Stem cells possess self-renewal and tissue-differentiation abilities. Research on stem cells is booming, and they are indispensable in tissue engineering studies aimed at repairing tissue and organ damage, making them one of the most popular research topics in regenerative medicine4. Among them, human umbilical cord mesenchymal stem cells (hUCMSCs) are a type of pluripotent stem cell that is easy to obtain, harmless to the human body, has strong proliferation and differentiation abilities, and has lower immunogenicity compared to mesenchymal stem cells from other sources5,6. Currently, they are the mainstream seed cells for stem cell applications. Using cellular molecular imaging methods, transplanted stem cells can be dynamically monitored, and their biological activity and histological effects can be assessed in real time, which is of great significance in stem cell mechanism and application research7,8,9.
Magnetic Resonance Imaging (MRI) is widely used in medical imaging owing to its safety and noninvasiveness10. However, MRI imaging often lacks sufficient contrast and may require contrast agents for enhancement. Magnetic Nanoparticles (MNPs), particularly Superparamagnetic Iron Oxide Nanoparticles (SPIONs), offer promising advantages owing to their unique surface and quantum size effects. They possess excellent dispersibility, tunable particle size, thermal stability, and biocompatibility11,12. Apart from serving as contrast agents in MRI, SPIONs have potential applications in magnetic hyperthermia, magnetic drug targeting, and biological separation13,14,15. Despite the significant potential of SPIONs in MRI imaging, their application in labeling stem cells for MRI imaging is still in its early stages16,17,18. Further research is required to assess its safety and efficacy19.
Currently, the application of SPIONs in cellular imaging faces a major challenge20: inadequate Enhanced Permeability and Retention (EPR) effect, that is, the need to prolong tissue retention time21,22,23,24,25. The main influencing factors of nanoparticles include particle size, spatial hindrance, hydrophilicity, and surface charge26,27,28. Proper modification of the SPIONs surface can prevent it from being attacked and cleared by soluble molecules, such as plasma proteins and opsonins recognized by the immune system29,30. The preparation of SPIONs using polymer coatings is simple and highly controllable31,32,33. For example, when PEG-modified SPIONs were functionalized with PEG, significant improvements were achieved in the specific surface area, structural uniformity, hydrophilicity, and adsorption capacity of SPIONs34,35,36,37.
This study aimed to identify the safety and efficacy of SPIONs labeling of hUCMSCs in vitro and trace the injection of SPIONs-labeled hUCMSCs in vivo. Monkeys were used as animal models for cesarean sections, and SPIONs-labeled hUCMSCs were injected into the monkey uterus. Furthermore, we traced and monitored the transplanted SPIONs-labeled hUCMSCs using MRI.
Method
Cell culture
Human umbilical cord mesenchymal stem cells (hUCMSCs) were purchased from Beijing Health&Biotech group (Beijing, China). The hUCMSCs were suspended in DMEM/F12 + GlutaMAX™ -I (1X) medium (Gibco) + 10% fetal bovine serum (Gibco) and were maintained at 37 ℃ in 5% CO2. The culture medium was changed every 2–3 days. The cells were passaged using 0.25% trypsin-EDTA (Gibco) at 80% confluence. The cells were obtained and used for experiments at passage 5 (P5) to reduce senescence-related effects.
Cell labeling
Previously, hUCMSCs in passage 5 were incubated in DMEM/F12 + GlutaMAX™ -I (1X) medium (Gibco), with SPIONs (PEG coating Fe3O4 nanoparticles, 30 nm, Ruixi Biotechnology Co., Xi’an, China) (50, 40, 30, 25, 20, 10, 0 µg/mL), at 80% confluence, for 24 h. After washing with phosphate buffer (PBS, pH 6.8) (Biosharp) three times, the supernatant was removed, and the cells were resuspended in PBS at a concentration adjusted to 1 × 106/mL. The cells were characterized by microscopy (DMi1, Leica) after Prussian blue staining (Solarbio). In addition, the cells and labeling were identified using transmission electron microscopy (TEM) (H−7500, Hitachi).
Cell viability assessment
After labeling for 1, 2, 3, 4, 5, 6, and 7 days, the labeled hUCMSCs were seeded in 96-well plates at a density of 10,000 cells/well in culture medium at 37℃ in 5% CO2. Proliferation was monitored daily for seven days. For monitoring, the supernatant was discarded, and 100 µL of culture medium and 10 µL of the Cell Counting Kit-8 (CCK-8) detection reagent (Dojindo) were added in a dark room. The cells were then incubated at 37℃ in 5% CO2 for 1–4 h. Absorbance was measured at 450 nm using an ELISA reader (EL10B, BIOBASE). The Cell Viability (calculated using rate) was:
Where ASample is the absorbance of the wells containing the SPIONs-labeled cell sample and CCK−8 detection reagent. ABlank refers to the absorbance of the wells containing the culture medium and CCK-8 detection reagent without cells. A0 is the absorbance of the wells containing unlabeled cells and the CCK-8 detection reagent. Cell viability refers to the proliferative or cytotoxic activity of cells.
Both the negative control (sterile polyethylene) and positive control (polyurethane with an organic tin stabilizer) were included for comparison, with three replicates per group.
Differentiation potential test
The possible effects of SPIONs-labeling on adipogenesis, osteogenesis, and chondrogenic differentiation were assessed. Briefly, labeled cells were seeded on 6-well plates at 1 × 105/well, differentiated in adipogenic induction medium (Pythonbio), osteogenesis induction medium (Cyagen), and chondrogenesis induction medium (Pythonbio). The cells were then incubated at 37℃ in 5% CO2. The medium was changed every second day. After 30 days, the induced cells were fixed with 4% paraformaldehyde (Leagene) and stained with Oil Red O, alizarin red, and Alcian Blue. The interference was observed using an optical microscope (DMi1, Leica). Six replicates were performed for each treatment group.
Cell surface marker expression
After labeling for 24 h, cells were digested and resuspended in PBS at a concentration of 5 × 106/mL. The prepared cells (100 µL) were incubated in the dark for 30 min with the following antibodies: 5 µL FITC Mouse Anti-Human CD90, 5 µL phycoerythrin (PE) Mouse Anti-Human CD44, 5 µL PerCP-CyTM5.5 Mouse Anti-Human CD105, APC Mouse Anti-Human CD73, 20 µL PE hMSC Negative Cocktail, and 20 µL hMSC Positive Cocktail (Human MSC Analysis Kit, BD). A flow cytometer (DxFLEX, Beckman Coulter) was used to analyze the results.
Immunocytochemistry staining of vimentin
To characterize the labeled hUCMSCs and to determine the effects of labeling, cells were seeded on coverslips in 24-well plates, incubated at 37℃ in 5% CO2. After 24 h, the coverslips were fixed in paraformaldehyde (4%) (Leagene), permeabilized with 0.2% Triton X−100, blocked with goat serum, incubated with Goat anti-Rabbit IgG-Cy3, incubated with Vimentin Mouse Monoclonal antibody (Proteintech), and Goat anti-Mouse IgG-FITC (Absin). A laser scanning confocal microscope (LSM 880, Zeiss) was used for staining.
In vitro MR imaging
The labeled hUCMSCs were digested and suspended in PBS in 1 mL plastic tubes. PBS and unlabeled cells were used as negative controls. MRI images were obtained using a 3.0 T MRI scanner (Discovery MR750 3.0T, GE HealthCare).
Laboratory animals
The cynomolgus monkeys (Macaca fascicularis) used in this study were female reproductive-age individuals, aged 4–7 years and weighing 4–8 kg. Healthy monkeys with a gestational age of 140–160 days (late pregnancy) were selected, totaling 2 animals. All monkeys were housed individually in stainless steel cages with free access to drinking water. The breeding environment was controlled at a temperature of 16–26℃, relative humidity of 40%–80%, and a 12 h light/12 h dark photoperiod. The animals were provided and raised by Guangdong Landao Biotechnology Co., Ltd. The animal experimental protocol involved in this study was approved by the Institutional Animal Care and Use Committee (IACUC) of Guangdong Landao Biotechnology Co., Ltd., and humane care was provided in accordance with the 3R principles (Replacement, Reduction, Refinement) for laboratory animal use.
Anesthesia and euthanasia
For surgical and MRI procedures with monkeys, anesthesia was induced with intramuscular ketamine (8 mg/kg) and maintained with 1–2% isoflurane inhalation. After surgery, continuous 3 days of intramuscular injection of oxytocin (10 IU) and continuous 7 days of intramuscular injection of ceftriaxone sodium (0.5 g) were administered. Euthanasia was performed via intravenous overdose of pentobarbital sodium (150 mg/kg) following deep anesthesia. Every effort was made to minimize animal number and suffering.
Surgical procedure
Under anesthesia, a 4 cm transverse suprapubic skin incision was made, the rectus sheath and muscles were bluntly separated, and the peritoneum was opened to expose the gravid uterus. A 1 cm transverse incision was created in the lower uterine segment, the membranes were ruptured manually, and the fetus was delivered. Immediately, 10 IU oxytocin was injected into the myometrium, the uterine incision was closed with interrupted 3−0 Vicryl sutures in a single layer, the peritoneum and sheath were re-approximated continuously, and the skin was closed intradermally with 4−0 Vicryl.
In vivo MR imaging
Two monkeys (Macaca fascicularis) were purchased from Guangdong Landau Biotechnology Co. Ltd. (Guangzhou, China). They were in their first pregnancy at term and used for in vivo MR Imaging. Six months after cesarean section, the abdomen was opened, the uterus was exposed, and 1 mL hUCMSCs (1 × 106/mL) with different concentrations of labeling SPIONs (100, 50, and 40 µg/mL for Monkey No.C102124, and 25, 20, and 10 µg/mL for Monkey No.C102122) in PBS was injected into the uterus (anterior wall, fundus, and posterior wall, respectively) before the abdomen was closed.
MRI was performed at 1, 7, 28, and 604 days after injection. An MRI scanner was used to observe signal changes in the abdominal and pelvic cavities.
Ex vivo histology
On the 667th day after the labeled cells injected, the uteruses of the two monkeys were removed, three pieces of anterior wall, fundus, posterior wall, and a normal piece (as negative control) of the uterus were cut out, fixed with 4% polyformaldehyde, kept at−80 ℃. The samples were sliced and stained with hematoxylin-eosin (HE), Masson’s trichrome, and Prussian Blue. Staining results were observed under a microscope.
Statistical analysis
All data are presented as mean ± standard deviation (SD). Statistically significant differences between groups were analyzed using the independent-sample t test and multivariate analysis of variance (ANOVA) test conducted using SPSS 20.0. P < 0.05 was considered statistically significant.
Ethics statement
All experimental protocols involving cynomolgus monkey (Macaca fascicularis) were approved by the Institutional Animal Care and Use Committee (IACUC) of Guangdong Landau Biotechnology Co., Ltd. (Approval No. LDACU 20190410−01). Throughout the entire experimental process, the 3R principles (Replacement, Reduction, Refinement) for laboratory animal use were strictly followed.
All methods were carried out in accordance with relevant guidelines and regulations, including the ARRIVE guidelines, and the American Veterinary Medical Association (AVMA) Guidelines for the Euthanasia of Animals (2020). Every effort was made to minimize the number of animals used and reduce their suffering, ensuring the highest standards of animal welfare and ethical compliance.
Experiment
Labeling characterization
The hUCMSCs adhered within 24 h, grew, and became spindle-like (Fig. 1A-B). After SPIONs labeling, there was no significant change in cell morphology. Blue iron particles were observed using an optical microscope after Prussian Blue staining, in contrast to the unlabeled cells (Fig. 1C-D). Under TEM, vesicular inclusions containing small iron particles were observed in the cytoplasm, with higher labeled concentrations indicating the presence of more particles in the cells (Fig. 1E-F).
Characterization of SPIONs-labeled HUCMSCs. (A). Unlabeled hUCMSCs. (B). SPIONs-labeled hUCMSCs. (C). Prussian blue stained unlabeled hUCMSCs. (D). Prussian blue stained SPIONs-labeled hUCMSCs. (E). Unlabeled hUCMSCs were observed using TEM. (F). SPIONs-labeled hUCMSCs observed by TEM.
Labeling viability
As shown in Fig. 2, the grow curves within 7 days of the hUCMSCs labeled with different concentrations of SPIONs were similar. The growth rate of 50 µg/mL labeling was slightly lower than that at other concentrations. The growth rates of 10, 20, and 25 µg/mL hUCMSCs increased by more than 100% compared to those of unlabeled hUCMSCs. These and previous results indicate that SPIONs do not cause serious injury to the hUCMSCs.
Cell Viability (cell survival rate). The cell viability of hUCMSCs labeled with different concentrations of SPIONs was analyzed using the CCK−8 assay. The growth rate of unlabeled hUCMSCs was defined as 1.0(n ≥ 3).
Cell surface marker expression
As shown in Fig. 3, unlabeled and SPIONs-labeled hUCMSCs were positive for the human MSC surface markers CD73, CD90, CD105 and CD44, comparing to the negative control (incubated antibodies without cells). According to these results, labeling had no influence on cell surface markers.
Flow cytometry analysis. Identification of cell surface markers of hUCMSCs labeled with different concentrations of SPIONs. (A) FITC CD90. (B) PC5.5 CD105. (C) APC CD73. (D) PE CD44. There was no obvious signal difference between the unlabeled hUCMSCs and SPIONs-labeled hUCMSCs. Both cell groups showed positive signals for these surface markers compared with the negative control group.
Differentiation potential test
Adipogenic, osteogenic, and chondrogenic differentiation of unlabeled and SPIONs-labeled SPIONs were verified by Oil Red O-positive intracellular lipid-rich vacuoles, calcium nodule formation, and Alcian Blue-positive proteoglycans, respectively. After inducing adipogenic differentiation for 30 days, red oil droplets were found in the cytoplasm and stained with Oil Red O (Fig. 4A-B). After inducing osteogenic differentiation for 30 days, dense red nodule-like precipitates were observed in the cells stained with Alizarin Red (Fig. 4C-D). After inducing chondrogenic differentiation for 30 days, cartilage balls were observed, which turned blue after Alcian Blue staining (Fig. 4E-F). No significant differences were found between the unlabeled and labeled groups. These results indicate that the differentiation potential of hUCMSCs was not affected by SPIONs-labeling.
Differentiation test. (A). Oil Red O staining of unlabeled hUCMSCs. (B). Oil Red O staining of induced SPIONs-labeled hUCMSCs. (C). Alizarin Red staining of induced unlabeled hUCMSCs. (D). Alizarin Red staining of induced SPIONs-labeled hUCMSCs. (E). Alcian Blue staining of the induced unlabeled hUCMSCs. (F). Alcian Blue staining of induced SPIONs-labeled hUCMSCs.
Immunocytochemistry staining of vimentin
Vimentin was detected by immunocytochemical staining (Figure S1), and there was no significant difference between labeled and unlabeled hUCMSCs.
In vitro MR imaging
The scanning sequence used for SPIONs-labeled hUCMSCs was OAx T2 FLAIR. The T2 signal of labeled cells was lower than that of the negative control (unlabeled cells). Cells labeled with 50 µg/mL SPIONs showed the lowest signal compared to cells labeled with 40, 30, 25, 20, 10, and 0 µg/mL SPIONs (Fig. 5A).
MRI of the monkey uterus after SPIONs-labeled HUCMSCs injection. (A). In vitro MRI of SPIONs-labeled hUCMSCs. (B and E): Seven days after the injection. (C and F): 28 days after injection. (D and G): 604 days after injection. Signal changes in uterus were marked with yellow arrowheads. For different labeling concentrations, a greater signal loss was observed at higher concentrations. (B, C, and D) show no obvious signal location change. (E, F, and G) also show no obvious changes in the signal location. (D and G) show a reduction in signal loss compared with that of (B, C, E, and F).
In vivo MR imaging
Following uterine injection of SPIONs-labeled hUCMSCs, enhanced areas of T2 loss signals (scanning sequence: FIESTA-C ARC Pel, OAx T2 FRFSE 3.0, SAG T2 PROPELLER, and SAG T1 FSE) were observed in monkeys, not only at 7 and 28 days, but also at 604 days, without location change. The hypointensity signal of 100 µg/mL SPIONs labeled hUCMSCs was higher than that of other concentrations (Fig. 5B-G). The signal location remains stable at all times. As the injection sites were not co-planar and cell concentrations varied, the MRI images provide qualitative evidence of localization rather than a quantitative comparison of signal intensity. These results demonstrated that the SPIONs label persisted at the injection site for over 600 days, allowing for long-term in vivo tracking.
Furthermore, the brains and kidneys of the two monkeys were MR scanned 604 days after injection. No suspicious enhanced areas of T2 signal loss was detected. These results indicated that there was no enrichment of SPIONs-labeled hUCMSCs in the brain, kidney, and liver (Figure S2).
Pathological results
The anterior wall of the uterus healed well, with sutures absorbed, and the scars were difficult to identify.
HE staining
There was no significant difference in the nuclei between the SPIONs-labeled hUCMSCs injected group and the negative control group, with a normal structure, clear nucleus, and abundant cytoplasm (Figure S3A-C).
Masson staining
Collagen fibers were blue, cytoplasm, muscle, and cellulose were red, and nuclei were dark blue. There were no significant differences found between the SPIONs-labeled hUCMSCs injected and negative control groups (Figure S3D-F).
Prussian blue staining
Blue staining particles were found in the SPIONs-labeled hUCMSCs injected group, but not in the negative control group. This result indicates that the SPIONs inside the hUCMSCs were well located and metabolized without deposition (Fig. 6).
Histological study by prussian blue staining. Prussian blue stained particles were found in the anterior wall of the uterus, after 604 days after injection of SPIONs-labeled hUCMSCs.
Discussion
In this study, the labeling safety of SPIONs in hUCMSCs was assessed in vitro and in vivo, application as MRI contrast agent injected into the uterus was confirmed. According to our study, hUCMSCs can be effectively labeled with SPIONs without a significant influence on cellular proliferation and differentiation potential. Cell surface marker expression and immunocytochemical staining of vimentin further demonstrated its minimal impact. MR scanning, both in vitro and in vivo, as well as pathological staining, was conducted. Two monkeys were used for in vivo scanning and pathological tests. These results confirmed that SPIONs labeling is effective without hUCMSCs injury, and these SPIONs-labeled hUCMSCs have the potential to serve as MRI contrast agents. This labeling was localized for a considerable period of time, allowing for dynamic colonization.
Monkeys were used as models of scarred uteri in our study. Rodents, such as mice, rats, rabbits, dogs, and other small animals, are normally used to establish disease models for preclinical research. These animals are cost-effective and easy to obtain and maintain, but their physiological structures and genetic makeup differ significantly from humans, leading to some bias when the research data are applied to clinical and human references. As non-human primates, monkeys, are highly similar to humans in terms of genetic, physiological, and immunometabolic aspects, with a genetic homology of 92–94% with humans38. Animal models established using monkeys can generate experimental data that are conducive to clinical translation. Furthermore, compared to rodents, crab-eating macaques have a longer lifespan and metabolism that is more similar to that of humans, allowing for more thorough observation and study of drug metabolism and histopathological changes in organs. Compared to other non-human primates, monkeys also have the advantages of being docile in temperament, having a short reproductive cycle, and being small in size, making them more suitable for use in toxicology, pharmacology, physiology, reproductive biology, and chronic disease model research39, and they are increasingly becoming an important experimental animal40. Our group, originating from obstetrics, has been conducting research on scarred uteri for over a decade. Utilizing primate models for clinical translation research was a highlight of this study. In models of scarred uterus, considering the significant differences in shape, size, litter size, etc., between the uterus of rodents such as mice and rabbits and that of humans, as well as the substantial mechanistic differences in post-cesarean section wound closure, drug administration, and recovery abilities, using crab-eating macaques, which are similar to humans in physiological, genetic, and biological characteristics, establishing scarred uterus models is an important research tool for studying the repair of uterine full-layer injury, especially the repair of the muscle layer.
The model monkeys, after undergoing cesarean section surgery and injection of SPIONs-labeled hUCMSCs, lived healthily for over two years without any evidence of health impairment. The uteri healed as scarred uteri. MRI resonance imaging of the brain, liver, spleen, and kidneys revealed no changes in organ morphology or particle deposition. HE staining and Masson staining revealed no alterations in cell morphology. Results from in vitro experiments showed that SPIONs labeling had no effect on biological activity, differentiation potential, surface markers, or vimentin expression in stem cells. An important issue regarding stem cell transplantation and efficacy is the duration of local implantation. In this study, even after 604 days, particles implanted in the uterine fundus, anterior wall, and posterior wall could still be detected on MR. Prussian blue staining of uterine samples collected 604 days later revealed SPIONs particles. These results indicate that labeling stem cells with SPIONs for implantation into the body allows a sufficiently long duration of local implantation.
Previous studies41 have shown that iron nanoparticles may accumulate in vivo, cause adverse health effects owing to the ion environment, disrupt immune regulation homeostasis, or induce autophagy before apoptosis through mitochondrial dysfunction and endoplasmic reticulum stress. It has been reported that42,43 nanoparticles predominantly circulate in the body through the lymphatic system and are removed from the circulation by macrophages via the spleen. Current literature consensus that exogenous stem cells seldom survive long-term in immunocompetent hosts and argue that the most plausible explanation for the signal persistence is the phagocytosis and retention of SPIONs by local macrophages after the clearance of the initially transplanted hUCMSCs44,45. Our study demonstrates long-term SPIONs signal retention, but also likely not long-term stem cell survival. The investigation of metabolic and immunological changes, cell apoptosis, and cell fate marked by SPIONs is not yet comprehensive, and there were limitations due to the insufficient sample size of monkey models. The next translational milestone is to refine tracking accuracy and rigorously assess the safety profile46, further extensive research with larger sample sizes and more detailed improvements may be needed for preclinical application of SPIONs-labeling.
Data availability
The datasets generated for this study are available on request to the corresponding author.
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Acknowledgements
We thank Professor Xiaoling Guo, Obstetrician Consultant of Foshan Women and Children Hospital, for creating an excellent scientific research environment for us.
Funding
This work was funded by the Science and Technology Project in Key Areas of Foshan (Grant No.2020001005861), Guangdong Basic and Applied Basic Research Foundation (Grant No. 2021A1515110802), Project of Foshan Science and Technology (2220001003887, 2220001004102).
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Huiting Ma and Yingchun Wan contributed equally to this work. They performed the idea together, while Huiting Ma performed the investigation and manuscript, Yingchun Wan performed the data analysis and manuscript editing. Xiuyin Shen, Xin Luo, Shuzhen Wu, and Xiafen Lu contributed to the research part. Fengying Chen and Weibin Liao contributed to the MRI testing part. Qingjian Deng, Ting Chen and Xiaotie Chu contributed to the animal monitoring during the operation. Zhengping Liu supervised this work, managed the project, and edited the manuscript. All authors have approved the submission of this manuscript.
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This study was approved by the Institutional Animal Care and Use Committee (IACUC) of Guangdong Landau Biotechnology Co. Ltd. (Guangzhou, China).
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Ma, H., Wan, Y., Shen, X. et al. SPIONs-labeled hUCMSCs for in vitro safety analysis and in vivo tracking in scarred monkey uteri. Sci Rep 16, 14199 (2026). https://doi.org/10.1038/s41598-026-45156-2
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DOI: https://doi.org/10.1038/s41598-026-45156-2
