Human Bone Marrow-derived Mononuclear Cells Research Articles

Exploring the Effects of Human Bone Marrow-Derived Mononuclear Cells on Angiogenesis In Vitro.

Cell therapies involving the administration of bone marrow-derived mononuclear cells (BM-MNCs) for patients with chronic limb-threatening ischemia (CLTI) have shown promise; however, their overall effectiveness lacks evidence, and the exact mechanism of action remains unclear. In this study, we examined the angiogenic effects of well-controlled human bone marrow cell isolates on endothelial cells. The responses of endothelial cell proliferation, migration, tube formation, and aortic ring sprouting were analyzed in vitro, considering both the direct and paracrine effects of BM cell isolates. Furthermore, we conducted these investigations under both normoxic and hypoxic conditions to simulate the ischemic environment. Interestingly, no significant effect on the angiogenic response of human umbilical vein endothelial cells (HUVECs) following treatment with BM-MNCs was observed. This study fails to provide significant evidence for angiogenic effects from human bone marrow cell isolates on human endothelial cells. These in vitro experiments suggest that the potential benefits of BM-MNC therapy for CLTI patients may not involve endothelial cell angiogenesis.

Safety and effects of intravitreal or subretinal injections of human bone marrow‐derived mononuclear cells to P23H‐1 and RCS rats

AbstractPurposeTo determine the safety and effects of intravitreal or subretinal injections of human bone marrow‐derived mononuclear cells (HBMNCs) in two different rat models of hereditary photoreceptor (FR) degeneration.Methods21‐days‐old Homozygous P23H‐1 albino rats, Royal College of Surgeons (RCS) pigmented rats or control SD and PVG rats received one intravitreal injection or one subretinal injection of HBMNCs (5µl; 50,000 cells/µl). As controls, intact animals of P23H‐1, RCS, SD and PVG strains were used. All animals were immunosuppressed with oral cyclosporine and intraperitoneal dexamethasone and were processed 7, 15, 30 or 60 days after transplantation. The eyes were cross‐sectioned in a cryostat and the sections immunodetected to visualize HBMNCs (□‐human CD45), photoreceptors (□‐Recoverin), astrocytes and Müller cells (□‐GFAP), or microglial cells (□‐Iba‐1). Sections were examined under fluorescence and confocal microscopy to investigate retinal structure and photoreceptor cell survivalResultsIn HBMNCs treated eyes no macro or microscopic anomalies, tumours, necrosis, inflammation or infective phenomena were observed. In the intravitreally injected eyes, HBMNCs‐CD45+ were found in the vitreous and attached to the inner limiting membrane. In the eyes with subretinal injections, HBMNCs‐CD45+ were found forming a layer in the subretinal space. When compared to control animals, the transplanted eyes of P23H‐1 and RCS rats showed decreased GFAP expression, indicating a reduction of gliosis. However, we did not observe increased photoreceptor survival in dystrophic animals after transplantation.ConclusionsHBMNCs injection to the vitreous or subretinal space in immunosupresed rats lack adverse effects and in P23H‐1 and RCS rats decrease the retinal gliosis but do not elicit photoreceptor neuroprotective effects.

Metabolism Regulates Cellular Functions of Bone Marrow-Derived Cells used for Cardiac Therapy.

Administration of bone marrow-derived mononuclear cells (BMC) may increase cardiac function after myocardial ischemia. However, the functional capacity of BMC derived from chronic heart failure (CHF) patients is significantly impaired. As modulation of the energy metabolism allows cells to match the divergent demands of the environment, we examined the regulation of energy metabolism in BMC from patients and healthy controls (HC). The glycolytic capacity of CHF-derived BMC is reduced compared to HC, whereas BMC of metabolically activated bone marrow after acute myocardial infarction reveal increased metabolism. The correlation of metabolic pathways with the functional activity of cells indicates an influence of metabolism on cell function. Reducing glycolysis without profoundly affecting ATP-production reversibly reduces invasion as well as colony forming capacity and abolishes proliferation of CD34(+) CD38(-) lin(-) hematopoietic stem and progenitor cells (HSPC). Ex vivo inhibition of glycolysis further reduced the pro-angiogenic activity of transplanted cells in a hind limb ischemia model in vivo. In contrast, inhibition of respiration, without affecting total ATP production, leads to a compensatory increase in glycolytic capacity correlating with increased colony forming capacity. Isolated CD34(+) , CXCR4(+) , and CD14(+) cells showed higher glycolytic activity compared to their negative counterparts. Metabolic activity was profoundly modulated by the composition of media used to store or culture BMC. This study provides first evidence that metabolic alterations influence the functional activity of human HSPC and BMC independent of ATP production. Changing the balance between respiration and glycolysis might be useful to improve patient-derived cells for clinical cardiac cell therapy. Stem Cells 2016;34:2236-2248.

Cell-based therapy by implanted human bone marrow-derived mononuclear cells improved bone healing of large bone defects in rats.

QUESTION/AIM: Cell-based therapy by cultivated stem cells (mesenchymal stem cells [MSC] and endothelial progenitor cells [EPC]) in a large-sized bone defect has already shown improved vascularization and new bone formation. However, these methods are clinically afflicted with disadvantages. Another heterogeneous bone marrow cell population, the so-called human bone marrow-derived mononuclear cells (BMC), has nevertheless been used clinically and showed improved vascularization in ischemic limbs or in the myocardium. For clinical use, a certified process has been established; thus, BMC were isolated from bone marrow aspirate by density gradient centrifugation, washed, cleaned, and given back to patients within several hours. This investigation tested the ability of human BMC seeded on beta-tricalcium phosphate (β-TCP) and placed into a large bone defect in rats to improve the bone healing process in vivo. Human EPC were isolated from buffy coat, and MSC or BMC, respectively, were isolated from bone marrow aspirate by density gradient centrifugation. 1.0×10(6) cells were loaded onto 750 μL β-TCP (0.7-1.4 mm). Large femoral defects (6 mm) in athymic rats were created surgically and stabilized with an internal fixateur. The remaining defects were filled with β-TCP granules alone (group 1), β-TCP+EPC/MSC (group 2), or β-TCP+BMC (group 3). After 8 weeks, histomorphometric analysis (new bone formation), radiological microcomputer tomography analysis (bony bridging), and biomechanical testing (three-point bending) were achieved. Moreover, a tumorigenicity study was performed to evaluate the safety of BMC implantation after 26 weeks. For statistical analysis, the Kruskal-Wallis test was used. Eight weeks after implantation of EPC/MSC or BMC, respectively, we detected a more significant new bone formation compared to control. In group 2 and 3, bony bridging of the defect was seen. In the control group, more chondrocytes and osteoid were detected. In the BMC and EPC/MSC group, respectively, less chondrocytes and a significantly more advanced bone formation were observed. The biomechanical stability of the bone regenerate was significantly enhanced if BMC and EPC/MSC, respectively, were implanted compared to control. Moreover, no tumor formation was detected either macroscopically or histologically after 26 weeks of BMC implantation. Implanted BMC suggest that a heterogeneous cell population may provide a powerful cellular therapeutic strategy for bone healing in a large bone defect in humans.

Improvement of liver fibrosis by infusion of cultured cells derived from human bone marrow

We develop "autologous bone marrow cell infusion (ABMi) therapy" for the treatment of human decompensated liver cirrhosis and confirm the efficacy and safety of this treatment in multicenter clinical studies. With the goal of further expanding the applications of ABMi, we first cultured human bone marrow cells and then determined whether a cell fraction found to be effective in improving liver fibrosis can be amplified. Cells harvested after two passages (P2 cells) consistently contained approximately 94% mesenchymal stem cells (MSCs); conversely, the cells harvested after only medium change (P0 cells) contained many macrophages. MSCs (2.8 × 10(8)) in P2 cells were harvested from 3.8 × 10(8) bone marrow-derived mononuclear cells after 22 days. DNA-chip analysis also showed during the culturing step that bone marrow-derived cells decreased with macrophage phenotype. The infused 5 × 10(5) P2 cells significantly improved liver fibrosis in the nonobese diabetic/severe combined immunodeficient (NOD-SCID) mouse carbon tetrachloride (CCl4) liver cirrhosis model and induced the expression of matrix metalloproteinase (MMP)-9 and suppressed expressions of alpha smooth muscle actin (αSMA), tumor necrosis factor alpha (TNFα) and transforming growth factor beta (TGFβ) in the liver. Cultured human bone marrow-derived cells (P2 cells) significantly inhibited liver fibrosis. The increase of MMP-9 and suppressed activation of hepatic stellate cells (HSCs) through the regulation of humoral factors (TNFα and TGFβ) contribute to the improvement of liver fibrosis by MSCs comprising about 94% of P2 cells. MSCs in cultured human bone marrow-derived mono-nuclear cells (BM-MNCs) proliferate sufficiently in cell therapy, so we believe our cultured bone marrow-derived cell therapy can lead to expanded clinical applications and enable outpatient therapy.

PTK787/ZK 222584 inhibits tumor growth promoting mesenchymal stem cells: Kinase activity profiling as powerful tool in functional studies

Bone marrow (BM)-derived mesenchymal stem cells (MSCs) have been shown to favour tumor growth, suggesting the relevance of pharmaceutical inhibition of MSCs for the treatment of malignancies. We tested the effect of PTK787/ZK 222584 (PTK) on the outgrowth of MSCs from human bone marrow-derived mononuclear cells (MNCs) and the migration and tube formation capacity of MSCs in vitro. PTK dose- dependently inhibited the outgrowth of BM-MSCs from BM-MNCs (LC50 1.12 μM PTK), while hematopoietic colony formation (HCF) was only slightly hampered (13 ± 19% at 1 μM PTK, and stable at ~50% at higher concentrations of PTK). Addition of 10 μM PTK inhibited proliferation of MSCs by 74 ± 6.6% compared to control (p

Angiotensin II Receptor Blocker Inhibits Neointimal Hyperplasia Through Regulation of Smooth Muscle–Like Progenitor Cells

Angiotensin II (ATII) type 1 receptor (AT1R) blocker (ARB) has been shown to inhibit neointimal formation. Bone marrow-derived mononuclear cells (BM-MNCs) give rise to smooth muscle (SM)-like cells at injured arterial wall and contribute to neointimal formation. However, role of the renin-angiotensin system in the homing process of SM-like cells during neointimal formation is unknown. When human BM-MNCs and peripheral blood MNCs (PB-MNCs) were cultured under treatment with PDGF-BB and bFGF, these cells gave rise to SM-like cells with expression of alphaSMA, SMemb, and SM1 proteins. RT-PCR showed the expression of AT1R, ATII type 2 receptor (AT2R), alphaSMA, and SMemb mRNAs. ATII accelerated the differentiation of SM-like cells, which was inhibited by an ARB CV11974 (P<0.05). We then examined the effects of ATII, CV11974, and AT2R antagonist PD123319 on neointimal formation and BM-derived SM-like cell incorporation at injured arteries in vivo. BM from green fluorescence protein (GFP)-transgenic mice was transplanted to irradiated WT mice. GFP-BM chimera mice were subjected to wire injury on the left femoral artery. ATII (100 ng/kg/min) stimulated whereas CV11974 (1 mg/kg/d) inhibited neointimal formation. Number of GFP+ alphaSMA+ cells at neointima correlated with the intima/media ratio (r=0.69, P<0.05). BM-derived SM-like progenitor cells contributed to the neointimal formation after arterial injury. ATII accelerated whereas ARB suppressed this process. These are new aspects of the ARB-mediated inhibition of atherosclerotic disease progression.