Menstrual Blood-derived Stem Cells Research Articles

Menstrual Blood-Derived Endometrial Stem Cells Ameliorate Ovarian Senescence by Relieving Oxidative Stress-Induced Inflammation.

Senescence is a degenerative process that occurs with ageing, and in the female reproductive system, senescence occurs earlier in the ovaries than in other tissues and organs, which implies a decrease in oocyte quality and exhaustion of the primordial follicular pool, leading to impaired ovarian function and an inability to maintain normal fertility. Unfortunately, the development of curative and effective treatments for ovarian senescence is still a considerable challenge. Currently, mesenchymal stem cells (MSCs)-based therapies for treating various refractory diseases, especially ovarian dysfunction, have been extensively studied and confirmed. However, the mechanisms by which MSCs improve ovarian senescence are not yet clear. Therefore, in this study, a mouse ageing model was generated via the intraperitoneal injection of a D-galactose (D-gal) solution, and the effects of menstrual blood-derived endometrial stem cells (MenSCs) transplantation on the ovarian follicle count, fibrosis level, and degree of apoptosis were evaluated. Subsequently, an ovarian granulosa cell ageing model was induced with H2O2, and CCK-8 assays, flow cytometry, mitochondrial membrane potential analysis and Western blotting were subsequently performed to further investigate the potential mechanism by which MenSCs ameliorate cellular oxidative damage. Overall, our findings demonstrated that MenSCs treatment can increase the cellular antioxidant capacity by activating the NRF2/HO-1 signalling pathway and further ameliorate the inflammatory ovarian microenvironment, apoptosis and dysfunction in ageing mice. These results provide reliable evidence and support for MenSCs-based therapy for ovarian senescence.

Menstrual blood-derived mesenchymal stem cells combining with platelet-rich plasma infusion in endometrium repair.

Thin endometrium caused by various factors affects the conception rate of females worldwide; however, current medications are still insufficient. Therefore, a novel approach is needed. We previously reported the effect of menstrual blood-derived mesenchymal stem cells (MenSCs) in ameliorating ethanol-induced endometrial injuries. In the present study, we aimed to investigate whether the effect of MenSCs is enhanced by a combination of platelet-rich plasma (PRP) infusion. An endometrial injury mouse model was established by infiltrating 95% ethanol for 15 s into the uterus, followed by MenSCs, PRP, or MenSCs + PRP treatment. Pathological changes were observed by HE staining. The expression of CK18, vimentin, ItgαVβ3, and VEGF was determined using IHC staining and WB blotting. Compared with the model, MenSCs, PRP, and MenSCs + PRP treatments significantly improved endometrial damage and thickness, with the combined therapy displaying the most pronounced efficacy. The density of CK18-, vimentin-, and ItgαVβ3 positive cells increased most significantly in the MenSCs + PRP group of mice. In addition, the protein expression of CK18, vimentin, and VEGF was significantly upregulated after MenSCs + PRP, MenSCs, and PRP treatment, with MenSCs + PRP therapy showing the best efficacy. MenSCs + PRP therapy is more beneficial for ameliorating ethanol-induced endometrial damage than MenSCs or PRP alone, providing a basis for the investigation of novel approaches for treating thin endometria.

Ameliorative Potential of Menstrual Blood-derived Endometrial Stem Cells in Alcohol-Induced Fatty Liver Disease

Background: Alcohol-induced fatty liver disease (AFLD) begins with steatosis and may progress to a range of pathological liver changes, including fibrosis, cirrhosis, and complications. Menstrual blood-derived endometrial stem cells (MenSCs) have shown potential therapeutic effects against various types of liver damage. However, the liver-protective effects of MenSCs in AFLD are not well understood. This study aimed to evaluate the therapeutic effects of MenSCs on AFLD progression. Methods: MenSCs were sourced from women in good health (N=5, 25-34 years old). Male C57BL/6 mice were separated into three distinct groups to establish the mouse models. The AH/- MenSCs group received MenSCs (5×105 cells/mouse) transplantation through tail injection on the 7th and 13th days following the initiation of the alcohol-induced fatty liver model. The therapeutic effects of MenSCs transplantation in AFLD mouse models were subsequently explored using qPCR, Western blotting, histopathological examination, and mRNA sequencing analysis. Results: MenSCs significantly improved liver function and reduced lipid accumulation in AFLD. Treatment with MenSCs was also found to reduce the expression levels of inflammatory cytokines and profibrotic markers in the liver tissues of the mouse model. Additionally, the MenSCs- treated group demonstrated a significant reduction in endoplasmic reticulum (ER) stress and oxidative stress, along with an increase in autophagic activity. Conclusion: The findings provided preliminary evidence of the multifaced protective effects of MenSCs in AFLD.

Intrauterine adhesions repair with menstrual blood-derived mesenchymal stem cells via CXCL13-CXCR5 signal axis and its mechanism

BackgroudIntrauterine Adhesions (IUA) is a common gynecological disease which is seriously endangers the reproductive function of women without any ideal treatment. Some researchers found Menstrual Blood-derived Mesenchymal Stem Cells (MenSCs) can repair of damaged endometrium, however, has not been fully clarified. This study aims to evaluate the therapeutic effects of MenSCs in IUA and the repair mechanism in vivo.MethodsThis study is Laboratory-based study. To evaluate the therapeutic effects of MenSCs in IUA, We cultivated MenSCs, established mouse endometrial injury model, observed the uterine morphology and degree of endometrial fibrosis and compared the expression of CXC chemokine ligand-13 (CXCL13)、CXC chemokine receptor-5 (CXCR5)、Plasmin Activating Inhibitor-1(Pai-1), Transforming Growth Faction-β1(TGF- β1) and Matrix Metalloproteinase-9 (Mmp-9) among each groups. GraphPad Prism 8.0 was used for statistical processing. Data were expressed as mean ± SD. Statistical comparisons among groups were performed with one-way ANOVA. P < 0.05 were considered statistically significant.ResultsWe successfully cultured and identified MenSCs and established mice model of uterine adhesion. After treatment with MenSCs, endometrial morphology of mice was partially restored, endometrial thickness was increased, and glands were multipiled. The concentrations of CXCL13 and CXCR5 were significantly increased by immunofluorescence detection compared with the control group. The results of RT-qPCR showed that the expressions of Pai-1 and Mmp-9 were significantly lower than those of the control group.ConclusionsMenSCs may reduce endometrial fibrosis and the down-regulating expression of Pai-1、Mmp-9 and CXCL13-CXCR5 axis were involved in the process of MenSCs repaired IUA.

Exosomes with Engineered Brain Derived Neurotrophic Factor on Their Surfaces Can Proliferate Menstrual Blood Derived Mesenchymal Stem Cells: Targeted Delivery for a Protein Drug.

Despite the efficacy of brain derived neurotrophic factor (BDNF) in neuro-regenerative medicine, it can't pass the blood-brain barrier. Recently, exosomes have been harnessed for targeted delivery of therapeutics into brain. Given these facts, an engineered exosome capable of BDNF expression on the surface would be an amenable tool for drug delivery. The BDNF gene was cloned into a plex-lamp lentiviral vector and virus particles were packaged using the Torano method. HEK293T cells were transduced by the purified viruses to produce and purify recombinant exosomes displaying the fusion protein on their surfaces. Western blot, Zeta sizer, TEM, and ELISA methods were used for exosome characterization. The effect of engineered exosomes on menstrual blood-derived mesenchymal stem cells (Mens-MSCs) proliferation was evaluated by cell counting assay, MTT assay, and qPCR on the bcl2 and nestin genes. Approximately, 1.8 × 108 TdU/ml of the viral particles was purified from the transfected cells and transduced into the HEK293T. Western blot and ELISA methods confirmed the surface display of the LAMP-BDNF fusion. TEM graphs and Zeta sizer results confirmed the morphology and the size of purified exosomes. Treatment of Mens-MSCs with the targeted exosomes augmented the expression level of bcl2 and nestin genes, increased the cell proliferation, and elevated the cell number. Chimeric BDNF on the exosome surface could retain its biological activity and elevate the expression of bcl2 and nestin genes. Moreover, these exosomes are capable of elevating the Mens-MSCs proliferation.

Menstrual blood-derived mesenchymal stem cells pretreated by low-intensity pulsed ultrasound repair endometrial injury by inhibiting the PI3K/AKT pathway

Research questionWhat is the effect and underlying mechanism of low-intensity pulsed ultrasound (LIPUS)-treated menstrual blood-derived mesenchymal stem cells (MenSCs) on intrauterine adhesions (IUA)? DesignFirst, MenSCs were identified and exposed to LIPUS. Then, the proliferation, migration, invasion, cytokine secretion, and ability to differentiate into human endometrial epithelial cells (HEECs) of LIPUS-treated MenSCs were characterized. In vitro, human endometrial stromal cells (HESCs) were treated with 10 ng/mL TGF-β1 to simulate IUA and then indirectly co-cultured with LIPUS-treated MenSCs. In vivo, IUA rat models were constructed and intrauterinely transplanted with LIPUS-treated MenSCs. Further, the morphology, structure, and levels of fibrosis and repair-related factors of the uterus were detected. In the mechanism study, IGF-1 was applied to verify whether the PI3K/AKT pathway participated in the repair of endometrial injury by LIPUS-treated MenSCs. ResultsIn vitro, LIPUS treatment showed beneficial effects on MenSCs by promoting cell proliferation and migration, inhibiting apoptosis, and enhancing the expression of EGF, HGF, and VEGF. It also facilitated the differentiation of MenSCs into HEECs while reducing fibrosis levels in TGF-β1-treated HESCs by decreasing p-PI3K and p-AKT levels. However, these effects were reversed with the use of insulin-like growth factor-1 (IGF-1). In vivo, transplantation of LIPUS-treated MenSCs resulted in increased uterine length, width, and weight. The transplanted cells also improved endometrial structure completeness, reduced inflammatory infiltration, increased endometrial thickness and gland abundance, and decreased endometrial fibrosis. Additionally, increased levels of endometrial repair-related proteins and receptivity-related markers were observed after transplantation of LIPUS-treated MenSCs. ConclusionLIPUS-treated MenSCs repaired endometrial injury by inhibiting the PI3K/AKT pathway.

A thermosensitive hydrogel for the sustained delivery of exosomes extracted from menstrual blood mesenchymal stem cells and frizzled antibody on triple-negative breast cancer cells in vitro

Due to the heterogeneity of triple-negative breast cancer (TNBC), conventional treatments usually respond poorly and effective targeted therapies for this type of cancer are rare. Wnt signaling plays an important role in breast cancer metastasis and proliferation, as shown by analyzes of gene expression profiling and genome-wide sequencing. In addition, the role of endometrial cells in regulating angiogenesis during the menstrual cycle as an angiostatic state associated with the end of the cycle has been well established. Therefore, exosomes of menstrual blood-derived stem cells (Exo-Mens) could play a role as effective inhibitors of tumor-induced angiogenesis. However, repeated administration and systemic toxicity are major problems with conventional drug delivery methods. To minimize these drawbacks and maximize the therapeutic benefits of the drugs, hydrogels provide convenient methods for drug delivery. The aim of this study was therefore to investigate the effects of hydrogels containing Exo-Mens and Frizzled antibodies (anti-FZ) on breast cancer cells. First, the optimization and characterization of the hydrogel were performed, then the cell lines (MDA-MB-231 and HUVEC) were studied in four groups: Hydrogel (control), Hydrogel with anti-FZ, Exo-Mens and FZ + Exo. First, the IC50 of anti-FZ and Exo-Mens was determined and the degradability and release pattern of the investigated groups were measured. Western blot (VEGF, HIF-1α) and qRT-PCR (VEGF, HIF-α and KDR) were performed for the HUVEC cell line and scratch tests, colony formation and apoptosis for the MDA-MB-231 cell line. In addition, invasion and migration were performed for both cell lines. Our results showed that the optimal hydrogel is a suitable option for sustained release of the studied groups. The loaded compounds could be released within 7 days. The angiogenic activity of dual anti-FZ and Exo-Mens within the hydrogel, showed a significant shift in the expression of pro-angiogenic genes and related proteins, including HIF-1α, VEGF-A and KDR, in the treated HUVECs. The hydrogel treatment groups were also able to reduce migration and invasion in both cell lines compared to the control group. In addition, a reduction in colony formation and an increase in apoptosis were observed, especially in the cancer cells exposed to anti-FZ. Angiogenesis as well as proliferation and invasion of tumor cells were inhibited by the developed hydrogel system containing anti-FZ, Exo-Mens and a combination of both with slow release.

Granulosa cell insight: unraveling the potential of menstrual blood-derived stem cells and their exosomes on mitochondrial mechanisms in polycystic ovary syndrome (PCOS)

BackgroundPolycystic ovary syndrome (PCOS) presents a significant challenge in women’s reproductive health, characterized by disrupted folliculogenesis and ovulatory dysfunction. Central to PCOS pathogenesis are granulosa cells, whose dysfunction contributes to aberrant steroid hormone production and oxidative stress. Mitochondrial dysfunction emerges as a key player, influencing cellular energetics, oxidative stress, and steroidogenesis. This study investigates the therapeutic potential of menstrual blood-derived stem cells (MenSCs) and their exosomes in mitigating mitochondrial dysfunction and oxidative stress in PCOS granulosa cells.MethodsUsing a rat model of PCOS induced by letrozole, granulosa cells were harvested and cultured. MenSCs and their exosomes were employed to assess their effects on mitochondrial biogenesis, oxidative stress, and estrogen production in PCOS granulosa cells.ResultsResults showed diminished mitochondrial biogenesis and increased oxidative stress in PCOS granulosa cells, alongside reduced estrogen production. Treatment with MenSCs and their exosomes demonstrated significant improvements in mitochondrial biogenesis, oxidative stress levels, and estrogen production in PCOS granulosa cells. Further analysis showed MenSCs' superior efficacy over exosomes, attributed to their sustained secretion of bioactive factors. Mechanistically, MenSCs and exosomes activated pathways related to mitochondrial biogenesis and antioxidative defense, highlighting their therapeutic potential for PCOS.ConclusionsThis study offers insights into granulosa cells mitochondria’s role in PCOS pathogenesis and proposes MenSCs and exosomes as a potential strategy for mitigating mitochondrial dysfunction and oxidative stress in PCOS. Further research is needed to understand underlying mechanisms and validate clinical efficacy, presenting promising avenues for addressing PCOS complexity.

Menstrual Blood Stem Cells-Derived Exosomes as Promising Therapeutic Tools in Premature Ovarian Insufficiency Induced by Gonadotoxic Systemic Anticancer Treatment.

Gonadotoxicity resulting from systemic and locoregional cancer treatments significantly threatens women's reproductive health, often culminating in premature ovarian insufficiency. These therapies, particularly alkylating agents and ionizing radiation, induce DNA damage and apoptosis in ovarian follicles, leading to infertility, amenorrhea, and estrogen deficiency, which exacerbate risks of osteoporosis and cardiovascular diseases. Existing fertility preservation methods do not prevent immediate ovarian damage, underscoring the need for innovative protective strategies. Menstrual blood-derived stem cells (MenSC) and their extracellular vesicles (EV) present promising regenerative potential due to their therapeutic cargo delivery and pathway modulation capabilities. Preclinical studies demonstrate that MenSC-derived EV ameliorate premature ovarian insufficiency by inhibiting granulosa cell apoptosis, promoting angiogenesis, and activating pivotal pathways such as SMAD3/AKT/MDM2/P53. However, comprehensive research is imperative to ensure the safety, efficacy, and long-term effects of MenSC-derived EV in clinical practice. In this review, we update the current knowledge and research regarding the use of MenSC-derived EV as a novel therapeutic weapon for ovarian regeneration in the context of gonadotoxicity induced by systemic anticancer treatment.

Selection of reference miRNAs for RT-qPCR assays in endometriosis menstrual blood-derived mesenchymal stem cells.

Choosing appropriate reference genes or internal controls to normalize RT-qPCR data is mandatory for the interexperimental reproducibility of gene expression data obtained by RT-qPCR in most studies, including those on endometriosis. Particularly for miRNAs, the choice for reference genes is challenging because of their physicochemical and biological characteristics. Moreover, the retrograde menstruation theory, mesenchymal stem cells in menstrual blood (MenSCs), and changes in post-transcriptional regulatory processes through miRNAs have gained prominence in the scientific community as important players in endometriosis. Therefore, we originally explored the stability of 10 miRNAs expressions as internal control candidates in conditions involving the two-dimensional culture of MenSCs from healthy women and patients with endometriosis. Here, we applied multiple algorithms (geNorm, NormFinder, Bestkeeper, and delta Ct) to screen reference genes and assessed the comprehensive stability classification of miRNAs using RefFinder. Pairwise variation calculated using geNorm identified three miRNAs as a sufficient number of reference genes for accurate normalization. MiR-191-5p, miR-24-3p, and miR-103a-3p were the best combination for suitable gene expression normalization. This study will benefit similar research, but is also attractive for regenerative medicine and clinics that use MenSCs, miRNA expression, and RT-qPCR.

Single-cell RNA sequencing reveals reduced intercellular adhesion molecule crosstalk between activated hepatic stellate cells and neutrophils alleviating liver fibrosis in hepatitis B virus transgenic mice post menstrual blood-derived mesenchymal stem cell transplantation.

Liver fibrosis can cause hepatitis B virus (HBV)-associated hepatocellular carcinoma. Menstrual blood-derived mesenchymal stem cells (MenSCs) can ameliorate liver fibrosis through paracrine. Single-cell RNA sequencing (scRNA-seq) may be used to explore the roadmap of activated hepatic stellate cell (aHSC) inactivation to target liver fibrosis. This study established HBV transgenic (HBV-Tg) mouse model of carbon tetrachloride (CCl4)-induced liver fibrosis and demonstrated that MenSCs migrated to the injured liver to improve serological indices and reduce fibrotic accumulation. RNA-bulk analysis revealed that MenSCs mediated extracellular matrix accumulation and cell adhesion. Liver parenchymal cells and nonparenchymal cells were identified by scRNA-seq in the control, CCl4, and MenSC groups, revealing the heterogeneity of fibroblasts/HSCs. A CellChat analysis revealed that diminished intercellular adhesion molecule (ICAM) signaling is vital for MenSC therapy. Specifically, Icam1 in aHSCs acted on Itgal/Itgb2 and Itgam/Itgb2 in neutrophils, causing decreased adhesion. The expression of Itgal, Itgam, and Itgb2 was higher in CCl4 group than in the control group and decreased after MenSC therapy in neutrophil clusters. The Lcn2, Pglyrp1, Wfdc21, and Mmp8 had high expression and may be potential targets in neutrophils. This study highlights interacting cells, corresponding molecules, and underlying targets for MenSCs in treating HBV-associated liver fibrosis.

Transplantation of hyaluronic acid and menstrual blood-derived stem cells accelerated wound healing in a diabetic rat model

Diabetic wounds require a multifactorial approach because several factors are involved in its occurrence. Herein we investigated whether transplantation of hyaluronic acid (HA) in combination with menstrual blood derived stem cells (MenSCs) could promote healing in diabetic rats. Thirty days after induction of diabetes, sixty animals were randomly planned into four equal groups: the untreated group, HA group, MenSC group, and HA+MenSC group. Sampling was done for histological, molecular, and tensiometrical assessments. Our results indicated that the wound contraction rate, volumes of new epidermis and dermis, collagen density, as well as tensiometrical parameter were considerably increased in the treatment groups compared to the untreated group and these changes were more obvious in the HA+MenSC ones. In addition, the expression levels of TGF-β and VEGF genes were significantly upregulated in treatment groups in comparison with the untreated group and were greater in the HA+MenSC group. This is while expression levels of TNF-α and IL-1β genes were more considerably downregulated in the HA+MenSC group than the other groups. We concluded that the combined use of HA and MenSCs has more effects on diabetic wound healing.

Human Menstrual Blood-Derived Stem Cells Protect against Tacrolimus-Induced Islet Dysfunction via Cystathionine β-Synthase Mediated IL-6/STAT3 Inactivation.

Diabetes imposes a huge burden worldwide. Islet transplantation is an alternative therapy for diabetes. However, tacrolimus, a kind of immunosuppressant after organ transplantation, is closely related to post-transplant diabetes mellitus. Mesenchymal stem cells (MSCs) have attracted interest for their potential to alleviate diabetes. In vivo experiments revealed that human menstrual blood-derived stem cells (MenSCs) treatment improved tacrolimus-induced blood glucose, body weight, and glucose tolerance disorders in mice. RNA sequencing was used to analyze the potential therapeutic targets of MenSCs. In this study, we illustrated that cystathionine β-synthase (CBS) contributed to tacrolimus -induced islet dysfunction. Using β-cell lines (MIN6, β-TC-6), we demonstrated that MenSCs ameliorated tacrolimus-induced islet dysfunction in vitro. Moreover, MenSC reduced the tacrolimus-induced elevation of CBS levels and significantly enhanced the viability, anti-apoptotic ability, glucose-stimulated insulin secretion (GSIS), and glycolytic flux of β-cells. We further revealed that MenSCs exerted their therapeutic effects by inhibiting CBS expression to activate the IL6/JAK2/STAT3 pathway. In conclusion, we showed that MenSCs may be a potential strategy to improve tacrolimus-induced islet dysfunction.

Enhanced therapeutic potential of Flotillins-modified MenSCs by improve the survival, proliferation and migration.

Menstrual blood-derived endometrial stem cells (MenSCs) have attracted increasing interest due to their excellent safety, and lack of ethical dilemma as well as their ability to be periodically obtained in a noninvasive manner. However, although preclinical research as shown the therapeutic potential of MenSCs in several diseases, their poor cell survival and low engraftment at disease sites reduce their clinical efficacy. Flotillins (including Flot1 and Flot2) are implicated in various cellular processes, such as vesicular trafficking, signal transduction, cell proliferation, migration and apoptosis. In this study, we aimed to determine the effects of Flotillins on MenSCs survival, proliferation and migration. Our experimental results show that MenSCs were modified to overexpress Flot1 and/or Flot2 without altering their intrinsic characteristics. Flot1 and Flot2 co-overexpression promoted MenSC viability and proliferation capacity. Moreover, Flot1 or Flot2 overexpression significantly promoted the migration and inhibited the apoptosis of MenSCs compared with the negative control group, and these effects were stronger in the Flot1 and Flot2 gene co-overexpression group. However, these effects were significantly reversed after Flot1 and/or Flot2 knockdown. In conclusion, our results indicate that Flot1 and Flot2 overexpression in MenSCs improved their proliferation and migration and inhibited their apoptosis, and this might be an effective approach to improve the efficiency of cell-based therapies.

Experimental study on the effects of human amniotic membrane in combination with menstrual blood-derived stem cells on wound healing in a diabetic rat model

One of the serious challenges in diabetic patients is the occurrence of complications caused by the disease. One of the most important side effects is wounding in limbs. Due to the multifactorial nature of these wounds, treatments require a multifaceted approach. Therefore, the aim of the present study was whether the human amniotic membrane (HAM) in combination with menstrual blood-derived stem cells (MenSCs) could promote wound healing in diabetic rats. Thirty days after induction of diabetes, the animals were randomly allocated into four equal groups (n=15): the control group, HAM group, MenSC group, and HAM+MenSC group. Sampling was done on days 7, 14, and 21 for histological, molecular, and tensiometrical evaluations. The results showed that the wound healing rate, collagen deposition, volumes of new epidermis and dermis, as well as tensiometrical characteristics were significantly increased in the treatment groups compared to the control group, and these changes were more obvious in the HAM+MenSC ones (P<0.05). Moreover, the expression levels of TGF-β, bFGF, and VEGF genes were considerably increased in treatment groups compared to the control group and were greater in the HAM+MenSC group (P<0.05). This is while expression levels of TNF-α and IL-1β decreased more significantly in the HAM+MenSC group than the other groups (P<0.05). We concluded that the combined use of HAM and MenSCs has a more significant effect on diabetic wound healing.

Menstrual blood-derived mesenchymal stem cells combined with collagen I gel as a regenerative therapeutic strategy for degenerated disc after discectomy in rats

BackgroundAnnulus fibrosis (AF) defects have been identified as the primary cause of disc herniation relapse and subsequent disc degeneration following discectomy. Stem cell-based tissue engineering offers a promising approach for structural repair. Menstrual blood-derived mesenchymal stem cells (MenSCs), a type of adult stem cell, have gained attention as an appealing source for clinical applications due to their potential for structure regeneration, with ease of acquisition and regardless of ethical issues.MethodsThe differential potential of MenSCs cocultured with AF cells was examined by the expression of collagen I, SCX, and CD146 using immunofluorescence. Western blot and ELISA were used to examine the expression of TGF-β and IGF-I in coculture system. An AF defect animal model was established in tail disc of Sprague-Dawley rats (males, 8 weeks old). An injectable gel containing MenSCs (about 1*106/ml) was fabricated and transplanted into the AF defects immediately after the animal model establishment, to evaluate its repairment properties. Disc degeneration was assessed via magnetic resonance (MR) imaging and histological staining. Immunohistochemical analysis was performed to assess the expression of aggrecan, MMP13, TGF-β and IGF-I in discs with different treatments. Apoptosis in the discs was evaluated using TUNEL, caspase3, and caspase 8 immunofluorescence staining.ResultsCoculturing MenSCs with AF cells demonstrated ability to express collagen I and biomarkers of AF cells. Moreover, the coculture system presented upregulation of the growth factors TGF-β and IGF-I. After 12 weeks, discs treated with MenSCs gel exhibited significantly lower Pffirrmann scores (2.29 ± 0.18), compared to discs treated with MenSCs (3.43 ± 0.37, p < 0.05) or gel (3.71 ± 0.29, p < 0.01) alone. There is significant higher MR index in disc treated with MenSCs gel than that treated with MenSCs (0.51 ± 0.05 vs. 0.24 ± 0.04, p < 0.01) or gel (0.51 ± 0.05 vs. 0.26 ± 0.06, p < 0.01) alone. Additionally, MenSCs gel demonstrated preservation of the structure of degenerated discs, as indicated by histological scoring (5.43 ± 0.43 vs. 9.71 ± 1.04 in MenSCs group and 10.86 ± 0.63 in gel group, both p < 0.01), increased aggrecan expression, and decreased MMP13 expression in vivo. Furthermore, the percentage of TUNEL and caspase 3-positive cells in the disc treated with MenSCs Gel was significantly lower than those treated with gel alone and MenSCs alone. The expression of TGF-β and IGF-I was higher in discs treated with MenSCs gel or MenSCs alone than in those treated with gel alone.ConclusionMenSCs embedded in collagen I gel has the potential to preserve the disc structure and prevent disc degeneration after discectomy, which was probably attributed to the paracrine of growth factors of MenSCs.

Biological characteristics of Muse cells derived from MenSCs and their application in acute liver injury and intracerebral hemorrhage diseases

The increasing interest in multilineage differentiating stress-enduring (Muse) cells within the field of regenerative medicine is attributed to their exceptional homing capabilities, prolonged viability in adverse conditions, and enhanced three-germ-layer differentiate ability, surpassing their parent mesenchymal stem cells. Given their abundant sources, non-invasive collection procedure, and periodic availability, human menstrual blood-derived endometrium stem cells (MenSCs) have been extensively investigated as a potential resource for stem cell-based therapies. However, there is no established modality to isolate Muse cells from MenSCs and disparity in gene expression profiles between Muse cells and MenSCs remain unknown. In this study, Muse cells were isolated from MenSCs by long-time trypsin incubation method. Muse cells expressed pluripotency markers and could realize multilineage differentiation in vitro. Compared with MenSCs, Muse cells showed enhanced homing ability and superior therapeutic efficacy in animal models of acute liver injury (ALI) and intracerebral hemorrhage (ICH). Furthermore, the RNA-seq analysis offers insights into the mechanism underlying the disparity in trypsin resistance and migration ability between Muse and MenSCs cells. This research offers a significant foundation for further exploration of cell-based therapies using MenSCs-derived Muse cells in the context of various human diseases, highlighting their promising application in the field of regenerative medicine.

Menstrual Blood-Derived Endometrial Stem Cell Transplantation Improves Male Reproductive Dysfunction in T1D Mice by Enhancing Antioxidative Capacity.

Diabetes is known to negatively affect male reproduction. Recent clinical results have confirmed that mesenchymal stem cell (MSC)-based therapies are safe and effective for the treatment of diabetes. However, the effect and potential mechanism through which MSC transplantation improves diabetes-derived male reproductive dysfunction are still unknown. In the present study, we first established a male T1D mouse model through intraperitoneal injection of streptozotocin for five consecutive days. Subsequently, we evaluated the blood glucose levels, fertility, and histology and immunology of the pancreas, testes, and penis of T1D mice with or without transplantation of menstrual blood-derived endometrial stem cells (MenSCs) or umbilical cord mesenchymal stem cells (UCMSCs). Glucose was added to the medium in which the Leydig cells were cultured to imitate high glucose-injured cell viability. Subsequently, we evaluated the cellular viability, ROS levels, and mitochondrial membrane potential of Leydig cells treated with or without MenSC-conditioned medium (MenSC-CM) using a CCK8 assay, immunofluorescence, and flow cytometry. The targeted proteins are involved in the potential mechanism underlying MenSC-derived improvements, which was further validated via Western blotting. Collectively, our results indicated that MenSC transplantation significantly ameliorated reproductive dysfunction in male T1D mice by enhancing cellular antioxidative capacity and promoting angiogenesis. This study provides solid evidence and support for the application of MSCs to improve diabetes-induced male reproductive dysfunction.

MALAT1/miR-7-5p/TCF4 Axis Regulating Menstrual Blood Mesenchymal Stem Cells Improve Thin Endometrium Fertility by the Wnt Signaling Pathway.

Thin endometrium (TE) is a significant factor contributing to fertility challenges, and addressing this condition remains a central challenge in reproductive medicine. Menstrual blood-derived mesenchymal stem cells (MenSCs) play a crucial role in tissue repair and regeneration, including that of TE. The Wnt signaling pathway, which is highly conserved and prevalent in eukaryotes, is essential for cell proliferation, tissue development, and reproductive functions. MALAT1 is implicated in various transcriptional and molecular functions, including cell proliferation and metastasis. However, the combined effects of the Wnt signaling pathway and the long non-coding RNA (lncRNA) MALAT1 on the regulation of MenSCs' regenerative capabilities in tissue engineering have not yet been explored. To elucidate the regulatory mechanism of MALAT1 in TE, we analyzed its expression levels in normal endometrium and TE tissues, finding that low expression of MALAT1 was associated with poor clinical prognosis. In addition, we conducted both in vitro and in vivo functional assays to examine the role of the MALAT1/miR-7-5p/TCF4 axis in cell proliferation and migration. Techniques such as dual-luciferase reporter assay, fluorescent in situ hybridization, and immunoblot experiments were utilized to clarify the molecular mechanism. To corroborate these findings, we established a TE model and conducted pregnancy experiments, demonstrating a strong association between MALAT1 expression and endometrial fertility. In conclusion, our comprehensive study provides strong evidence supporting that lncRNA MALAT1 modulates TCF4 expression in the Wnt signaling pathway through interaction with miR-7-5p, thus enhancing MenSCs-mediated improvement of TE and improving fertility.

Transcriptome profile analysis in spinal cord injury rats with transplantation of menstrual blood-derived stem cells.

Menstrual blood-derived stem cells (MenSCs) are vital in treating many degenerative and traumatic disorders. However, the underlying molecular mechanisms remain obscure in MenSCs-treating spinal cord injury (SCI) rats. MenSCs were adopted into the injured sites of rat spinal cords at day 7 post surgery and the tissues were harvested for total RNA sequencing analysis at day 21 after surgery to investigate the expression patterns of RNAs. The differentially expressed genes (DEGs) were analyzed with volcano and heatmap plot. DEGs were sequentially analyzed by weighted gene co-expression network, functional enrichment, and competitive endogenous RNAs (ceRNA) network analysis. Next, expression of selected miRNAs, lncRNAs, circRNAs and mRNAs were validated by quantitative real-time polymerase chain reaction (qRT-PCR). Bioinformatics packages and extra databases were enrolled to scoop the genes functions and their interaction relationships. A total of 89 lncRNAs, 65 circRNAs, 120 miRNAs and 422 mRNAs were significantly upregulated and 65 lncRNAs, 72 circRNAs, 74 miRNAs, and 190 mRNAs were significantly downregulated in the MenSCs treated rats compared to SCI ones. Current investigation revealed that MenSCs treatment improve the recovery of the injured rats and the most significantly involved pathways in SCI regeneration were cell adhesion molecules, nature killer cell mediated cytotoxicity, primary immunodeficiency, chemokine signaling pathway, T cell receptor signaling pathway and B cell receptor signaling pathway. Moreover, the lncRNA-miRNA-mRNA and circRNA-miRNA-mRNA ceRNA network of SCI was constructed. Finally, the protein-protein interaction (PPI) network was constructed using the top 100 DE mRNAs. The constructed PPI network included 47 nodes and 70 edges. In summary, the above results revealed the expression profile and potential functions of differentially expressed (DE) RNAs in the injured spinal cords of rats in the MenSCs-treated and SCI groups, and this study may provide new clues to understand the mechanisms of MenSCs in treating SCI.