Exploring the critical therapeutic window: Dose-frequency optimization of human umbilical cord mesenchymal stem cells for preclinical asthma treatment

Human umbilical cord blood-derived mesenchymal stem cells (HUCB-MSCs) have been studied in several models of immune-mediated disease because of their unique immunomodulatory properties. We hypothesized that HUCB-MSCs could suppress the inflammatory response in postischemic kidneys and attenuate early renal injury. In 8- to 10-wk-old male C57BL/6 mice, bilateral ischemia-reperfusion injury (IRI) surgery was performed, and 1 × 10(6) HUCB-MSCs were injected intraperitoneally 24 h before surgery and during reperfusion. Renal functional and histological changes, HUCB-MSC trafficking, leukocyte infiltration, and cytokine expression were analyzed. Renal functional decline and tubular injury after IRI were attenuated by HUCB-MSC treatment. PKH-26-labeled HUCB-MSCs trafficked into the postischemic kidney. Although numbers of CD45-positive leukocytes in the postischemic kidney were comparable between groups, the expression of interferon-γ in the postischemic kidney was suppressed by HUCB-MSC treatment. The rapid decrease in intrarenal VEGF after IRI was markedly mitigated by HUCB-MSC treatment. In inflammatory conditions simulated in a cell culture experiment, VEGF secretion from HUCB-MSCs was substantially enhanced. VEGF inhibitor abolished the renoprotective effect of HUCB-MSCs after IRI. Flow cytometry analysis revealed the decreased infiltration of natural killer T cells and increased number of regulatory T cells in postischemic kidneys. In addition, these effects of HUCB-MSCs on kidney infiltrating mononuclear cells after IRI were attenuated by VEGF inhibitor. HUCB-MSCs attenuated renal injury in mice in the early injury phase after IRI, mainly by humoral effects and secretion of VEGF. Our results suggest a promising role for HUCB-MSCs in the treatment of renal IRI.

Mitigating effects of hUCB-MSCs on the hematopoietic syndrome resulting from total body irradiation

Combinatorial effects of conception and governor vessel electroacupuncture and human umbilical cord blood-derived mesenchymal stem cells on pathomorphologic lesion and cellular apoptosis in rats with cerebral ischemia/reperfusion

To investigate the effects of human umbilical cord blood-derived mesenchymal stem cells(HUCMSC) on the leukemic cell line HL-60 and acute lymphoblastic leukemia cell line Jurkat as well as the role of CXCL12/CXCR4. HL-60 cells and Jurkat cells were co-cultured with human umbilical cord blood mesenchymal stem cell (HUCMSC), and the model was treated with G-CSF, AMD3100 and their combination. The cell viability and cell cycle were measured by Cell Counting Kit-8 (CCK-8), the apoptosis and the cell-cycle analysis were assessed by flow cytometry with the Annexin V/PI double staining. The expression of surface CXCR4 protein and total CXCR4 protein of leukemic cells were detected by flow cytometry and Western blot respectively. HUCMSC could decrease the viability of HL-60 cells and Jurkat cells, as well as the percentage of apoptotic cells, they could also increase the number of G0/G1 cells, while G-CSF and AMD3100 could reduce the proliferation of HL-60 cells and Jurkat cells in HUCMSC co-culture model, destructed the anti-apoptotic effect of HUCMSC on HL-60 cells and Jurkat cells, and the combination of 2 drugs resulted in a synergistic effect. The G-CSF could reduce the expression of surface CXCR4 protein and total CXCR4 protein in leukemic cells, while AMD3100 could only decrease the expression of surface CXCR4 protein of leukemia cell membrane, having no effect on the expression of CXCR4 protein in cytoplasm. Human umbilical cord blood mesenchymal stem cells can inhibit the proliferation and apoptosis of acute leukemia cells and increase the number of G0/G1 phase cells in leukemic cells. The AMD3100 can decrease the expression of surface CXCR4 protein in leukemia cells, G-CSF can decrease expression of total CXCR4 protein as well as membrane CXCR4 protein. Both of them can block the CXCL12/CXCR4 signal axis, weakening the relationship between leukemia cells and microenvironment. And on the basic of HUCMSC influenced leukemia cells' growth and proliferation, the cell viability will be weakened, its apoptosis will be promoted, and the percentage of G0/G1 phase cells in leukemia cells will be decreased.

Induction of Human Umbilical Cord Blood Mesenchymal Stem Cells into Nerve-Like Cells by Salvia Miltiorrhiza.

Mesenchymal stem cells (MSCs) have multiple immunomodulatory properties and hold therapeutic potential for inflammatory diseases. However, the therapeutic and immunologic effects of human umbilical cord blood-derived MSCs (huMSCs) remain largely unexamined for asthma. This study was to investigate the immunomodulatory properties of huMSCs in an ovalbumin (OVA)-induced murine asthma model. Mice were injected intraperitoneally with OVA and an aluminium hydroxide adjuvant. huMSCs were administered via the tail vein (5×105 cells/100uL) to female BALB/c mice prior to the initial OVA challenge. The effects of huMSCs were assessed by investigating airway hyperresponsiveness, histological changes, inflammatory cell numbers, serum allergen-specific antibodies, cytokine production in spleen, lung tissue, and bronchoalveolar lavage (BAL) fluid as well as expansion of regulatory T cells. Administration of huMSCs significantly reduced methacholine bronchial hyperresponsiveness and eosinophil counts in BAL cells. Similarly, there was a significant decrease in serum OVA-specific IgE and IgG1 levels along with Th2 cytokine production (IL-4, IL-5, and IL-13) in the lung and spleen tissues, whereas increased percentage of regulatory T cells was observed after treatment with huMSCs. Our results suggest that huMSC treatment reduces OVA-induced allergic inflammation, which could be mediated by regulatory T cells.

Human mesenchymal stem cells (MSCs) are promising therapeutics for autoimmune diseases due to their immunomodulatory effects. In particular, human umbilical cord blood-derived MSCs (hUCB-MSCs) have a prominent therapeutic effect on atopic dermatitis (AD). However, the underlying mechanism is unclear. This study investigated the role of transforming growth factor-beta (TGF-β) in the therapeutic effect of hUCB-MSCs on AD. Small interfering RNA (siRNA)-mediated depletion of TGF-β disrupted the therapeutic effect of hUCB-MSCs in a mouse model of AD by attenuating the beneficial changes in histopathology, mast cell infiltration, tumor necrosis factor-alpha (TNF-α) expression, and the serum IgE level. To confirm that hUCB-MSCs regulate secretion of TNF-α, we investigated whether they inhibit TNF-α secretion by activated LAD2 cells. Coculture with hUCB-MSCs significantly inhibited secretion of TNF-α by LAD2 cells. However, this effect was abolished by siRNA-mediated depletion of TGF-β in hUCB-MSCs. TNF-α expression in activated LAD2 cells was regulated by the extracellular signal-related kinase signaling pathway and was suppressed by TGF-β secreted from hUCB-MSCs. In addition, TGF-β secreted by hUCB-MSCs inhibited maturation of B cells. Taken together, our findings suggest that TGF-β plays a key role in the therapeutic effect of hUCB-MSCs on AD by regulating TNF-α in mast cells and maturation of B cells.

Several studies have demonstrated that human umbilical cord blood-derived mesenchymal stem cells can promote neural regeneration following brain injury. However, the therapeutic effects of human umbilical cord blood-derived mesenchymal stem cells in guiding peripheral nerve regeneration remain poorly understood. This study was designed to investigate the effects of human umbilical cord blood-derived mesenchymal stem cells on neural regeneration using a rat sciatic nerve crush injury model. Human umbilical cord blood-derived mesenchymal stem cells (1 × 106) or a PBS control were injected into the crush-injured segment of the sciatic nerve. Four weeks after cell injection, brain-derived neurotrophic factor and tyrosine kinase receptor B mRNA expression at the lesion site was increased in comparison to control. Furthermore, sciatic function index, Fluoro Gold-labeled neuron counts and axon density were also significantly increased when compared with control. Our results indicate that human umbilical cord blood-derived mesenchymal stem cells promote the functional recovery of crush-injured sciatic nerves.

RETRACTED: Exosomal miR-22-3p from human umbilical cord blood-derived mesenchymal stem cells protects against lipopolysaccharid-induced acute lung injury

The objective of the current study was to investigate the ability of human umbilical cord blood‑derived mesenchymal stem cells (HUCB‑MSCs) to undergo chondrogenic differentiation, by co‑culture with rabbit chondrocytes. The aim was to obtain more seed cells for tissue engineering research and lay the foundation for the clinical repair of cartilage defects. The studies were performed using isolated rabbit cartilage cells and HUCB‑MSCs in vitro, which were co‑cultured in a 2:1 or 3:1 ratio with or without insulin‑like growth factor‑1 (IGF‑1). Following 7 and 14 days in culture, cell morphology was observed in each group. RNA and protein were extracted to assess the expression levels of aggrecan (ACAN) and collagen type II (COL2A) using quantitative polymerase chain reaction (qPCR) and western blotting, respectively. Groups of cells that were co‑cultured exhibited significantly higher expression levels of ACAN and COL2A mRNA and protein, compared with the reduced effect of IGF‑1 at days 7 and 14 in culture. The addition of IGF‑1 was found to potentiate these effects. Specifically, at day 7, cells co‑cultured at a ratio of 2:1 had a greater induction of ACAN and COL2A compared with cells co‑cultured at a 3:1 ratio. However, following 14 days culture, cells co‑cultured at a 3:1 ratio with additional IGF‑1 exhibited a greater induction of ACAN and COL2A compared with cells co‑cultured at a ratio of 2:1. Human chondrocytes may be successfully induced by co‑culture of HUCB‑MSCs with rabbit chondrocytes, thus providing a theoretical basis to obtain seed cells with the capacity to differentiate into multiple cell types, with low immunogenicity. Notably, these cells may provide a valuable resource for tissue engineering.

Introduction: Androgenic alopecia (AGA), a hair loss condition caused by dihydrotestosterone binding to hair follicle receptors, negatively impacts quality of life for both men and women. Current treatments like minoxidil and finasteride have limitations, highlighting the need for alternative therapies, such as human umbilical cord blood-derived mesenchymal stem cells (HUCB-MSCs). Methods: In this study, forty-eight adult male Wistar albino rats (3 months old) were used. The control group (Group I) received no treatment, while the other rats underwent AGA induction via daily subcutaneous testosterone injections (100 mg/kg). These rats developed alopecia and were divided into three groups: AGA (Group II), AGA plus daily minoxidil spray (Group III), and AGA plus a single intradermal injection of HUCB-MSCs (1 mL containing 1 × 105 cells, Group IV). After 4 weeks, the rats were sacrificed, and skin specimens were prepared for histological analysis using H&E, Masson’s trichrome, and immunohistochemical staining for CK 19, vascular endothelial growth factor (VEGF), and TUNEL antibodies. Results: It was shown that HUCB-MSC treatment reversed structural damage to hair and follicles, normalizing conditions within 1-week post-injection. The treatment enhanced the anagen phase, suppressed telogen and catagen phases, reduced apoptosis, and increased VEGF and CK 19 immune reactions. Observational follow-up for Groups III and IV revealed that while the minoxidil group experienced significant hair loss after 37 days, the stem cell group exhibited dense and long hair covering the treated area. Conclusion: HUCB-MSC therapy demonstrated superior efficacy over minoxidil with no observed side effects, indicating its potential as a promising alternative for AGA treatment.

Background: Loss of calcium/calmodulin-dependent kinase II (CaMKII) after ischemic stroke exacerbates cell death by sensitizing vulnerable neurons to excitotoxic glutamate signaling and inducing neurotoxicity. Our recent study demonstrated the potential of human umbilical cord blood-derived mesenchymal stem cells (HUCB-MSCs) treatment in inhibiting apoptosis after ischemic stroke. In this study, we aimed to investigate the effect of focal cerebral ischemia and/or reperfusion on CaMKII expression and the role of HUCB-MSCs treatment on CaMKII regulation. To our knowledge this is the first study that demonstrates the possible involvement of CaMKII in HUCB-MSCs-mediated neuroprotection after ischemic stroke. Methods: Male Sprague-Dawley rats were obtained and randomly assigned to various groups. Rats were subjected to a two-hour middle cerebral artery occlusion (MCAO) procedure followed by seven days reperfusion. HUCB-MSCs (0.25x106cells/animal) were intravenously injected via tail vein 24 hours post-MCAO to designated animals. Rats brain tissues obtained seven days after reperfusion from various groups were subjected to real-time PCR, immunoblot and immunofluorescence analysis. Results: CaMKII protein expression did not change in animals subjected to 1h, 2h, and 4h of ischemia without reperfusion. However, CaMKII expression is significantly reduced when the animals were subjected to ischemia followed by one day reperfusion. The loss of CaMKII was persistent until 14 days after reperfusion. All CaMKII isoforms (CaMKIIα, CaMKIIγ and CaMKIIδ), except CaMKIIβ, were downregulated. CaMKII expression in neurons was also reduced in the ischemic hemisphere. HUCB-MSCs treatment 24h after reperfusion revert CaMKII mRNA and protein expression. Significant co-localization of CaMKII with neurons was also noticed in rats subjected to stem cell treatment. Conclusions: HUCB-MSCs-mediated neuroprotection after ischemic stroke could be mediated by upregulation of neuronal CaMKII expression.

Accumulating evidence suggests that oxidative DNA damage plays a critical role in cell death associated with ischemic stroke. Endogenous oxidative DNA damage can be detected in the ischemic brain during the stages that precedes the manifestation of cell death and is believed to trigger cell death via various intracellular signaling pathways. Inhibiting the signaling associated with DNA damage induction or facilitating the signaling associated with the DNA repair process can be neuroprotective against brain injury after ischemic stroke. Recent reports demonstrated that human umbilical cord blood-derived mesenchymal stem cells (HUCB-MSCs) prevented the upregulation of apoptotic signaling pathway molecules and thereby attenuated the extent of apoptosis after focal cerebral ischemia as well as improved the neurological recovery. Therefore, we hypothesized that HUCB-MSCs treatment after focal cerebral ischemia prevents the overexpression of molecules associated with DNA damage induction as well as augments the expression of molecules associated with DNA repair process. In order to test our hypothesis, we administered HUCB-MSCs (0.25x106cells/animal) intravenously via tail vein to male Sprague-Dawley rats that were subjected to a two-hour middle cerebral artery occlusion followed by one-day reperfusion. Ischemic brain tissues obtained from various groups seven days’ post reperfusion were subjected to DNA damage signaling pathway PCR microarray. Our results demonstrated the induction of both DNA damage inducing and repair genes after focal cerebral ischemia and reperfusion. HUCB-MSCs treatment downregulated the DNA damage inducing genes and upregulated the DNA repair genes without disturbing the endogenous defense mechanisms.

Background/Aims: Stem cell treatment is one of the potential treatment options for ischemic stroke. We recently demonstrated a protective effect of human umbilical cord blood-derived mesenchymal stem cells (HUCB-MSCs) in a rat model of ischemic stroke. The treatment attenuated apoptosis and prevented DNA damage. A collection of published studies, including several from our laboratory, indicated the induction and detrimental role for several matrix metalloproteinases (MMPs) in post-stroke brain injury. We hypothesized that the HUCB-MSCs treatment after focal cerebral ischemia prevents the dysregulation of MMPs and induces the expression of endogenous tissue inhibitors of metalloproteinases (TIMPs) to neutralize the elevated activity of MMPs. Methods: To test our hypothesis, we administered HUCB-MSCs (0.25 million cells/animal and 1 million cells/animal) intravenously via tail vein to male Sprague-Dawley rats that were subjected to a transient (two-hour) right middle cerebral artery occlusion (MCAO) and one-day reperfusion. Ischemic brain tissues obtained from various groups of rats seven days after reperfusion were subjected to real-time PCR, immunoblot, and immunofluorescence analysis. Results: HUCB-MSCs treatment prevented the induction of MMPs, which were upregulated in ischemia-induced rats that received no treatment. HUCB-MSCs treatment also prevented the induction of TIMPs expression. The extent of prevention of MMPs and TIMPs induction by HUCB-MSCs treatment is similar at both the doses tested. Conclusion: Prevention of stroke-induced MMPs upregulation after HUCB-MSCs treatment is not mediated through TIMPs upregulation.

The ability of mesenchymal stem cells (MSCs) to differentiate into neural cells makes them potential replacement therapeutic candidates in neurological diseases. Presently, overexpression of brain-derived neurotrophic factor (BDNF), which is crucial in the regulation of neural progenitor cell differentiation and maturation during development, was sufficient to convert the mesodermal cell fate of human umbilical cord blood-derived MSCs (hUCB-MSCs) into a neuronal fate in culture, in the absence of specialized induction chemicals. BDNF overexpressing hUCB-MSCs (MSCs-BDNF) yielded an increased number of neuron-like cells and, surprisingly, increased the expression of neuronal phenotype markers in a time-dependent manner compared with control hUCB-MSCs. In addition, MSCs-BDNF exhibited a decreased labeling for MSCs-related antigens such as CD44, CD73, and CD90, and decreased potential to differentiate into mesodermal lineages. Phosphorylation of the receptor tyrosine kinase B (TrkB), which is a receptor of BDNF, was increased significantly in MSC-BDNF. BDNF overexpression also increased the phosphorylation of β-catenin and extracellular signal-regulated kinases (ERKs). Inhibition of TrkB availability by treatment with the TrkB-specific inhibitor K252a blocked the BDNF-stimulated phosphorylation of β-catenin and ERKs, indicating the involvement of both the β-catenin and ERKs signals in the BDNF-stimulated and TrkB-mediated neural differentiation of hUCB-MSCs. Reduction of β-catenin availability using small interfering RNA-mediated gene silencing inhibited ERKs phosphorylation. However, β-catenin activation was maintained. In addition, inhibition of β-catenin and ERKs expression levels abrogated the BDNF-stimulated upregulation of neuronal phenotype markers. Furthermore, MSC-BDNF survived and migrated more extensively when grafted into the lateral ventricles of neonatal mouse brain, and differentiated significantly into neurons in the olfactory bulb and periventricular astrocytes. These results indicate that BDNF induces the neural differentiation of hUCB-MSCs in culture via the TrkB-mediated phosphorylation of ERKs and β-catenin and following transplantation into the developing brain.