Bone Marrow Mononuclear Cell Transplantation Research Articles

Autologous bone marrow mononuclear cell therapy improves symptoms in patients with end-stage peripheral arterial disease and reduces inflammation-associated parameters

Background aimsThe purpose of this study was to evaluate the effect of autologous bone marrow mononuclear cells (BM-MNCs) on symptoms and perfusion indices in severely symptomatic patients with peripheral arterial disease (PAD) without further option for endovascular or surgical revascularization. MethodsOnly patients with severe symptomatic PAD (Fontaine class IIb-IV, Rutherford category 3–6) not amenable for revascularization were treated. Bone marrow from both cristae iliacae was harvested; MNCs were isolated by the Ficoll density-gradient method and transplanted by means of intra-arterial and intramuscular injection in the index limb. Functional (pain score, ulcer healing, maximum walking distance) and perfusion indices such as ankle-brachial-index and transcutaneous oxygen pressure were documented before and after BM-MNC therapy. Additionally, serum concentration of C-reactive protein and interleukin-6 were measured as markers of inflammation before and after BM-MNC treatment. ResultsSixteen consecutive patients (four women; mean age, 63.0 ± 13 years) were treated with a mean dose of 4.2 ± 2.2 × 108 BM-MNCs. At 6 months' follow-up, ankle-brachial-index, transcutaneous oxygen pressure and maximum walking distance significantly increased, whereas C-reactive protein and interleukin-6 conversely decreased (P < 0.01 versus baseline values), resulting in 88% limb salvage, 75% pain reduction and 71% complete wound healing and/or reduction of ulcer size. One major and one minor amputation were performed, both in patients with Rutherford category 6. ConclusionsAutologous BM-MNC therapy in patients with end-stage PAD improves tissue perfusion indices and decreases markers of inflammation. If our observations could be confirmed by large-scale, randomized controlled trials, BM-MNC transplantation could become an alternative therapeutic option for patients with end-stage PAD.

Bone marrow mononuclear cell transplantation promotes therapeutic angiogenesis via upregulation of the VEGF–VEGFR2 signaling pathway in a rat model of vascular dementia

Bone marrow mononuclear cells (BMMNCs) are important for angiogenesis after stroke. We investigated the effects of BMMNCs on cognitive function, angiogenesis, and the vascular endothelial growth factor (VEGF)-VEGF receptor 2 (VEGFR2) signaling pathway in a rat model of vascular dementia. We transplanted BMMNCs into rats that had undergone permanent bilateral occlusion of the common carotid arteries (2VO) and observed their migration in vivo. On day 28, we assessed cognitive function with the Morris Water Maze test and examined vascular density and white matter damage within the corpus striatum by staining with fluorescein lycopersicon esculentum (tomato) lectin or Luxol fast blue. We evaluated expression of VEGF, rapidly accelerated fibrosarcoma 1 (Raf1), and extracellular-signal-regulated kinases 1 and 2 (ERK1/2) in the ischemic hemisphere by Western blot analysis on day 7 after cell transplantation. Contribution of the VEGF-VEGFR2 signaling pathway was confirmed by using VEGFR2 inhibitor SU5416. BMMNCs penetrated the blood-brain barrier and reached the ischemic cortex and white matter or incorporated into vascular walls of 2VO rats. BMMNC-treated 2VO rats had better learning and memory, higher vascular density, and less white matter damage than did vehicle-treated rats. The beneficial effects of BMMNCs were abolished by pretreatment of rats with SU5416. Protein expression of VEGF and phosphorylated Raf1 and ERK1/2 was also significantly increased by BMMNC treatment, but this upregulation was reversed by SU5416. BMMNCs can enhance angiogenesis, reduce white matter damage, and promote cognitive recovery in 2VO rats. The angiogenic effect may result from upregulation of the VEGF-VEGFR2 signaling pathway.

Autologous bone marrow mononuclear cell transplantation in ischemic heart failure: A prospective, controlled, randomized, double-blind study of cell transplantation combined with coronary bypass

Bone marrow mononuclear cell (BMMC) transplantation for heart failure has shown inconsistent therapeutic efficacy. We enrolled 104 ischemic heart failure patients scheduled for coronary artery bypass surgery (CABG). After 4- to 12-week pharmacotherapy optimization, 39 patients with left ventricular ejection fraction (LVEF) of ≤45% received injections of BMMC or vehicle intra-operatively into the myocardial infarction border area in a randomized, double-blind manner. The median number of cells injected was 8.4 × 10(8) (interquartile range [IQR]: 5.2 × 10(8) to 13.5 × 10(8)). We measured LV function and myocardial scar size by magnetic resonance imaging (MRI), and viability by positron emission tomography (PET) and single-photon emission computed tomography (SPECT), pre-operatively and after 1-year follow-up. LVEF, the pre-defined primary end-point measure, improved by a median of 5.6% in the control group (IQR 0.2 to 10.1) and by 4.8% in the BMMC group (IQR -0.5 to 8.2) (p = 0.59). Wall thickening in injected segments rose by a median of 4.5% among controls (IQR -18.1 to 23.9) and by 5.5% in the BMMC group (IQR -6.6 to 26.5) (p = 0.68). Changes in viability by PET and SPECT did not differ between groups. Myocardial scar size by MRI in injected segments rose by a median of 5.1% among controls (IQR -3.3 to 10.8), but fell by 13.1% in the BMMC group (IQR -21.4 to -6.5) (p = 0.0002). BMMC therapy combined with CABG failed to improve LV systolic function, or viability, despite reducing myocardial scar size.

The effect of bone marrow mononuclear cells transplantation on brain injury after intracerebral hemorrhage in rats

Objective To investigate the effect of intravenously transplanted bone marrow mononuclear cells(BMMNCs) on brain injury and inflammation after intracerebral hemorrhage(ICH) in rats. Methods Experimental ICH models were performed by stereotaxic injection collagenase Ⅳ into caudate putamen, rats that underwent ICH were randomly divided into four groups: sham operation group, ICH group, PBS group, BMMNC-treated group.The BMMNCs were injected intravenously into rats after ICH. The neurobehavioral function was evaluated on days 1, 3, 7, 14 by the modified neurological severity score, and the brain edema was examined by wet-dry weighting method on day 3 after cell transplantation. Immumofluorecence staining was used to identify the number of activation of microglia and infiltration of neutrophils in the brain after ICH. Results The neurological score in BMMNC-treated group on days 7, 14 was significantly improved compared with those in ICH group and PBS group(P<0.05). Compared to the ICH group((81.09±0.83)%) and PBS group((80.99±0.79)%), there was a significant decrease in the brain water content in BMMNC-treated group((78.62±0.97)%)(P<0.05). The number of activation of microglia and infiltration of neutrophils were both significantly lower in BMMNC-treated group ((55.8±22.1)/mm2, (49.6±12.9)/mm2) compared to ICH group and PBS group(respectively(125.0±20.7) /mm2, (86.8±13.6/mm2))(P<0.01). Conclusion Administration of BMMNCs can significantly reduce edema and improve neurologic function by inhibiting the activation of microglia and infiltration of neutrophils. Key words: Bone marrow mononuclear cells; Intracerebral hemorrhage; Edema; Microglia activation; Neutrophil infiltration

Opposite effects of bone marrow-derived cells transplantation in MPTP-rat model of Parkinson's disease: a comparison study of mononuclear and mesenchymal stem cells.

The 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) animal model is a useful tool to study Parkinson's disease (PD) and was used in the present study to investigate the potential beneficial as well as deleterious effects of systemic bone-marrow mononuclear cell (BMMC) or mesenchymal stem cell (BM-MSC) transplantation. MPTP administration resulted in a breakdown of the blood-brain barrier and motor impairment in the open field test 24 h after surgery. Three and 7 days after receiving the lesion, the injured animals showed remaining motor impairment compared to the sham groups along with a significant loss of tyrosine hydroxylase-immunoreactive (TH-ir) cells in the substantia nigra pars compacta (SNpc). The MPTP-lesioned rats treated with BMMCs immediately after lesioning exhibited motor impairment similar to the MPTP-saline group, though they presented a significantly higher loss of TH-ir cells in the SNpc compared to the MPTP-saline group. This increased loss of TH-ir cells in the SNpc was not observed when BMMC transplantation was performed 24 h after MPTP administration. In contrast, in the MPTP animals treated early with systemic BM-MSCs, no loss of TH-ir cells was observed. BMMCs and BM-MSCs previously labeled with CM-DiI cell tracker were found in brain sections of all transplanted animals. In addition, cells expressing CD45, an inflammatory white blood cell marker, were found in all brain sections analyzed and were more abundant in the MPTP-BMMC animals. In these animals, Iba1+ microglial cells showed also marked morphological changes indicating increased microglial activation. These results show that systemic BMMC transplantation did not ameliorate or prevent the lesion induced by MPTP. Instead, BMMC transplantation in MPTP-lesioned rats accelerated dopaminergic neuronal damage and induced motor impairment and immobility behavior. These findings suggest that caution should be taken when considering cell therapy using BMMCs to treat PD. However, systemic BM-MSC transplantation that reaches the injury site and prevents neuronal damage after an MPTP infusion could be considered as a potential treatment for PD during the early stage of disease development.

Intra-Spinal Bone Marrow Mononuclear Cells Transplantation Inhibits the Expression of Nuclear Factor-κB in Acute Transection Spinal Cord Injury in Rats.

ObjectiveTo assess the effect of bone marrow mononuclear cells (BMMNCs) transplantation in the expression of nuclear factor-κB (NF-κB) in spinal cord injury (SCI) in rats.MethodsBMMNCs were isolated from tibia and femur by a density gradient centrifugation. After establishment of acute transection SCI, rats were divided into experiment (BMMNCs), experiment control (0.1 M PBS infused) and sham surgery groups (laminectomy without any SCI). Locomotor function was assessed weekly for 5 weeks post-injury using BBB locomotor score and urinary bladder function daily for 4 weeks post-injury. Activity of NF-κB in spinal cord was assessed by immunohistochemistry and reverse transcriptase polymerase chain reaction.ResultsAt each time point post-injury, sham surgery group had significantly higher Basso, Beattie, Bresnahan locomotor and urinary bladder function scores than experiment and experiment control group (p<0.05). At subsequent time interval there were gradual improvement in both experiment and experiment control group, but experiment group had higher score in comparison to experiment control group (p<0.05). Comparisons were also made for expression of activated NF-κB positive cells and level of NF-κB messenger RNA in spinal cord at various time points between the groups. Activated NF-κB immunoreactivity and level of NF-κB mRNA expression were significantly higher in control group in comparison to experiment and sham surgery group (p<0.05).ConclusionBMMNCs transplantation attenuates the expression of NF-κB in injured spinal cord tissue and thus helps in recovery of neurological function in rat models with SCI.

Brain Protection Therapy for Ischemic Brain Injury

Therapeutic strategies for protecting the brain against ischemic injury aim to salvage the ischemic penumbra or to expand the therapeutic time window, or both. We have investigated the protective effects of several agents using a rat model of transient focal cerebral ischemia. Our experimental studies have identified the following agents with attractive protective mechanisms against brain ischemia: the free-radical scavenger edaravone (MCI-186), the immunosuppressant tacrolimus (FK506), bone marrow stromal cells (BMSCs) /bone marrow mononuclear cells (BMMCs), the ω3 polyunsaturated fatty acid eicosapentaenoic acid (EPA), and the hydroxymethyl glutaryl coenzyme A reductase inhibitor atorvastatin. The free-radical scavenger MCI-186 has a Bax/Bcl-2-dependent antiapoptotic mechanism in a rat model of transient focal cerebral ischemia. The therapeutic time window for FK506 in rat transient focal ischemia is from 60 to 120 minutes after ischemia induction, and mild hypothermia (35°C) expands the therapeutic time window up to 120 minutes after ischemia induction. Intravenous transplantation of BMSCs 120 minutes after ischemia induction, which is out of the therapeutic time window for FK506 alone, is neuroprotective when performed in conjunction with FK506 injection. Intra-arterial transplantation of BMMCs is more protective against transient focal cerebral ischemia than is intravenous transplantation of BMMCs. Repeated intravenous transplantation of BMMCs achieves further neuroprotection against ischemic brain injury. Pretreatment with ethyl-EPA inhibits endothelial Rho-kinase activation and reduces tissue oxidative stress following transient ischemia, resulting in strong neuroprotection. The long-term protective effects of atorvastatin against brain ischemia require continuous oral administration after ischemia/reperfusion. All of the agents used in our experimental studies are established therapies for ischemic stroke or other disorders. Further investigations are needed before such experimental brain protection strategies can be applied clinically.

Flap Prefabrication and Stem Cell–Assisted Tissue Expansion

Total face skin and soft-tissue defects remain one of the biggest challenges in reconstructive surgery. Reconstruction of the entire face with uniform coverage and delicate features is difficult to achieve. To avoid the patchwork result seen in multiple flaps and skin grafts, 1 monoblock flap that has similar color, texture, and thickness might be an ideal option to minimize the incisional scars and several surgical procedures but is unavailable with current approaches because of the lack of sufficient matched tissue and the unreliable blood supply for such a large flap. To acquire a monoblock flap for full face reconstruction, we combine the prefabricated flaps, skin overexpansion, and bone marrow mononuclear stem cell transplantation for total facial resurfacing. In this article, we present our experience from our case series that provides universally matched skin and near-normal facial contour. It is a reliable and an excellent reconstructive option for massive facial skin defect.

Effect of repeated allogeneic bone marrow mononuclear cell transplantation on brain injury following transient focal cerebral ischemia in rats

AimsTransplantation of bone marrow mononuclear cells (BMMCs) exerts neuroprotection against cerebral ischemia. We examined the therapeutic timepoint of allogeneic BMMC transplantation in a rat model of focal cerebral ischemia, and determined the effects of repeated transplantation outside the therapeutic window. Main methodsMale Sprague–Dawley rats were subjected to 90minute focal cerebral ischemia, followed by intravenous administration of 1×107 allogeneic BMMCs or vehicle at 0, 3 or 6h after reperfusion or 2×107 BMMCs 6h after reperfusion. Other rats administered 1×107 BMMCs at 6h after reperfusion received additional BMMC transplantation or vehicle 9h after reperfusion. Infarct volumes, neurological deficit scores and immunohistochemistry were evaluated 24 or 72h after reperfusion. Key findingsInfarct volumes at 24h were significantly decreased in transplantation rats at 0 and 3h, but not at 6h, after reperfusion, compared to vehicle-treatment. Even high dose BMMC transplantation at 6h after reperfusion was ineffective. Repeated BMMC transplantation at 6 and 9h after reperfusion reduced infarct volumes and significantly improved neurological deficit scores at 24 and 72h. Immunohistochemistry showed repeated BMMC transplantation reduced ionized calcium-binding adapter molecule 1, 4-hydroxy-2-nonenal and 8-hydroxydeoxyguanosine expression at 24 and 72h after reperfusion. SignificanceIntravenous allogeneic BMMCs were neuroprotective following transient focal cerebral ischemia, and the therapeutic time window of BMMC transplantation was >3h and <6h after reperfusion in this model. Repeated transplantation at 6 and 9h after reperfusion suppressed inflammation and oxidative stress in ischemic brains, resulting in improved neuroprotection.

Early Transplantation of Bone Marrow Mononuclear Cells Promotes Neuroprotection and Modulation of Inflammation After Status Epilepticus in Mice by Paracrine Mechanisms

Status epilepticus (SE) is a severe clinical manifestation of epilepsy associated with intense neuronal loss and inflammation, two key factors involved in the pathophysiology of temporal lobe epilepsy. Bone marrow mononuclear cells (BMMC) attenuated the consequences of pilocarpine-induced SE, including neuronal loss, in addition to frequency and duration of seizures. Here we investigated the effects of BMMC transplanted early after the onset of SE in mice, as well as the involvement of soluble factors produced by BMMC in the effects of the cell therapy. Mice were injected with pilocarpine for SE induction and randomized into three groups: transplanted intravenously with 1×10(7) BMMC isolated from GFP transgenic mice, injected with BMMC lysate, and saline-treated controls. Cell tracking, neuronal counting in hippocampal subfields and cytokine analysis in the serum and brain were performed. BMMC were found in the brain 4h following transplantation and their numbers progressively decreased until 24h following transplantation. A reduction in hippocampal neuronal loss after SE was found in mice treated with live BMMC and BMMC lysate when compared to saline-treated, SE-induced mice. Moreover, the expression of inflammatory cytokines IL-1β, TNF-α, IL-6 was decreased after injection of live BMMC and to a lesser extent, of BMMC lysate, when compared to SE-induced controls. In contrast, IL-10 expression was increased. Analysis of markers for microglia activation demonstrated a reduction of the expression of genes related to type 1-activation. BMMC transplantation promotes neuroprotection and mediates anti-inflammatory effects following SE in mice, possibly through the secretion of soluble factors.

Preconditioning via Angiotensin Type 2 Receptor Activation Improves Therapeutic Efficacy of Bone Marrow Mononuclear Cells for Cardiac Repair

BackgroundThe therapeutic efficiency of bone marrow mononuclear cells (BMMNCs) autologous transplantation for myocardial infarction (MI) remains low. Here we developed a novel strategy to improve cardiac repair by preconditioning BMMNCs via angiotensin II type 2 receptor (AT2R) stimulation.Methods and ResultsAcute MI in rats led to a significant increase of AT2R expression in BMMNCs. Preconditioning of BMMNCs via AT2R stimulation directly with an AT2R agonist CGP42112A or indirectly with angiotensin II plus AT1R antagonist valsartan led to ERK activation and increased eNOS expression as well as subsequent nitric oxide generation, ultimately improved cardiomyocyte protection in vitro as measured by co-culture approach. Intramyocardial transplantation of BMMNCs preconditioned via AT2R stimulation improved survival of transplanted cells in ischemic region of heart tissue and reduced cardiomyocyte apoptosis and inflammation at 3 days after MI. At 4 weeks after transplantation, compared to DMEM and non-preconditioned BMMNCs group, AT2R stimulated BMMNCs group showed enhanced vessel density in peri-infarct region and attenuated infarct size, leading to global heart function improvement. ConclusionsPreconditioning of BMMNCs via AT2R stimulation exerts protective effect against MI. Stimulation of AT2R in BMMNCs may provide a new strategy to improving therapeutic efficiency of stem cells for post MI cardiac repair.

The effects of bone marrow mononuclear cells on neurodegeneration after ischemia in mice

Objective To investigate the effects of bone marrow mononuclear cells (BMMNCs) on neurodegeneration after cerebral ischemia in mice. Methods Adult male mice were randomly divided into PBS treated group and BMMNCs treated group. The model was established with suture emboli method; BMMNCs were isolated with density gradient centrifugation method.The production of cytokines from BMMNCs including interleukin-10 (IL-10), vascular endothelial growth factor (VEGF) and insulinlike growth factor-I (IGF-1) were tested by enzyme-linked immunosorbent assay (ELISA) under normoxic or hypoxic conditions.Mice were injected with PBS or BMMNCs via tail vein 6 h after ischemia reperfusion.Neurological functional deficits, the activity of microglial cell and neurodegenration were assessed on day 1, 4, 7 after MCAO, and infarct volume was assessed on day 7. Results (1) Compared with BMMNCs under normoxic conditions, BMMNCs under hypoxic conditions secreted more IL-10, VEGF, IGF-1 after 24h of cultivation(P<0.05). (2) BMMNCs transplantation significantly reduced neurologic functional deficits and the number of activated microglia and degenerated neurons on days 4, 7 compared with PBS treated groups (P<0.05). (3) Compared with PBS treated group((37.1±6.9)%), BMMNCs transplantation significantly reduced infarct volume((27.8±7.5)%)(P<0.05). Conclusion Bone marrow mononuclear cells can significantly inhibit neurodegeneration, promote neurological recover and reduce infarct volume through inhibition of microglial cell activation by secretion of cytokines including IL-10, VEGF and IGF-1. Key words: Ischemic stroke; Bone marrow mononuclear cells; Cell transplantation; Cytokines; Microglia; Neurodegeneration

Extracellular High Mobility Group Box 1 Plays a Role in the Effect of Bone Marrow Mononuclear Cell Transplantation for Heart Failure

Transplantation of unfractionated bone marrow mononuclear cells (BMCs) repairs and/or regenerates the damaged myocardium allegedly due to secretion from surviving BMCs (paracrine effect). However, donor cell survival after transplantation is known to be markedly poor. This discrepancy led us to hypothesize that dead donor BMCs might also contribute to the therapeutic benefits from BMC transplantation. High mobility group box 1 (HMGB1) is a nuclear protein that stabilizes nucleosomes, and also acts as a multi-functional cytokine when released from damaged cells. We thus studied the role of extracellular HMGB1 in the effect of BMC transplantation for heart failure. Four weeks after coronary artery ligation in female rats, syngeneic male BMCs (or PBS only as control) were intramyocardially injected with/without anti-HMGB1 antibody or control IgG. One hour after injection, ELISA showed that circulating extracellular HMGB1 levels were elevated after BMC transplantation compared to the PBS injection. Quantitative donor cell survival assessed by PCR for male-specific sry gene at days 3 and 28 was similarly poor. Echocardiography and catheterization showed enhanced cardiac function after BMC transplantation compared to PBS injection at day 28, while this effect was abolished by antibody-neutralization of HMGB1. BMC transplantation reduced post-infarction fibrosis, improved neovascularization, and increased proliferation, while all these effects in repairing the failing myocardium were eliminated by HMGB1-inhibition. Furthermore, BMC transplantation drove the macrophage polarization towards alternatively-activated, anti-inflammatory M2 macrophages in the heart at day 3, while this was abolished by HMGB1-inhibition. Quantitative RT-PCR showed that BMC transplantation upregulated expression of an anti-inflammatory cytokine IL-10 in the heart at day 3 compared to PBS injection. In contrast, neutralizing HMGB1 by antibody-treatment suppressed this anti-inflammatory expression. These data suggest that extracellular HMGB1 contributes to the effect of BMC transplantation to recover the damaged myocardium by favorably modulating innate immunity in heart failure.

Autologous Bone Marrow Mononuclear Cells Transplant in Patients With Critical Leg Ischemia: Preliminary Clinical Results

Stem cell transplant can induce vasculogenesis and improve the blood supply to an ischemic region, offering hope for chronic lower extremity ischemic diseases. Bone marrow mononuclear cells are one of the sources for stem cell transplants. We sought to observe the safety and efficacy of autologous bone marrow mononuclear cells transplant for treating critical limb ischemia. Eligible patients were randomized 1:1 to receive placebo (0.9% NaCl) or 1 × 107 piece/mL bone marrow mononuclear cell transplant. For 6 months, patients' skin ulcers, ankle-brachial index, and rest pain were examined and recorded before and after treatment. Six months after the bone marrow mononuclear cells transplant, clinical symptoms like rest pain and skin ulcers gradually abated (P < .05). Ankle-brachial index also increased after the transplant (P < .01). Autologous bone marrow mononuclear cells transplant for treatment of patients with chronic limb ischemia is safe, effective, and feasible.

GW24-e1170 The effects and safety of autologous intracoronary Bone Marrow Mononuclear Cells transplantation in cardiomyopathy

ObjectivesTo invest the effects and safety of autologous bone marrow–derived mononuclear cells (BMMNCs) transplantation in dilated cardiomyopathy. We report the results of the randomised trial of BMMNCs therapy in 20...

Bone Marrow Mononuclear Cell Transplantation Increases Metalloproteinase-9 and 13 and Decreases Tissue Inhibitors of Metalloproteinase-1 and 2 Expression in the Liver of Cholestatic Rats

Liver fibrosis results from chronic injury followed by activation of macrophages and fibrogenic cells like myofibroblasts and activated hepatic stellate cells. These fibrogenic cells express α-smooth muscle actin (α-SMA) and produce excessive extracellular matrix (ECM), with disorganization and loss of function of hepatic parenchyma. It is known that increased levels of metalloproteinases (MMPs) in liver fibrosis are associated with reduction of the pathologic ECM and fibrosis resolution. Recently, it has been shown that bone marrow mononuclear cells (BMMNCs) may reduce collagen and α-SMA expression, and ameliorate liver function in cholestatic rats. Therefore, this study aimed to analyze MMP-2, MMP-9 and MMP-13, and tissue inhibitors of MMPs (TIMPs)-1 and TIMP-2 in the liver of cholestatic rats transplanted with BMMNC. Animals were divided into normal rats, cholestatic rats obtained after 14 and 21 days of bile duct ligation (BDL), and rats obtained after 14 days of BDL that received BMMNCs and were killed after 7 days. MMP and TIMP expression was assessed by Western blotting, along with α-SMA, CD68 and CD11b expression by confocal microscopy. Western blotting analysis showed that 14-day BDL animals had significantly reduced amounts of MMP-2 and MMP-13, but increased amounts of MMP-9 compared to normal rats. After 21 days of BDL, overall MMP amounts were decreased and TIMPs were increased. BMMNC transplantation significantly increased MMP-9 and MMP-13, and decreased TIMP expression. Increased MMP activity was confirmed by zymography. MMP-9 and MMP-13 were expressed by macrophages near fibrotic septa, suggesting BMMNC may stimulate MMP production in fibrotic livers, contributing to ECM degradation and hepatic regeneration.

A pilot study of autologous CD34-depleted bone marrow mononuclear cell transplantation via the hepatic artery in five patients with liver failure

Background aimsMany rodent experiments and human studies on stem cell therapy have shown promising therapeutic approaches to liver diseases. We investigated the clinical outcomes of five patients with liver failure of various causes who received autologous CD34-depleted bone marrow-derived mononuclear cell (BM-MNC) transplantation, including mesenchymal stromal cells, through the hepatic artery. MethodsCD34-depleted BM-MNCs were obtained from five patients waiting for liver transplantation by bone marrow aspiration and using the CliniMACS CD34 Reagent System (Miltenyi Biotech, Bergisch Gladbach, Germany), and autologous hepatic artery infusion was performed. The causes of hepatic decompensation were hepatitis B virus (HBV), hepatitis C virus (HCV), propylthiouracil-induced toxic hepatitis and Wilson disease. ResultsSerum albumin levels improved 1 week after transplantation from 2.8 g/dL, 2.4 g/dL, 2.7 g/dL and 1.9 g/dL to 3.3 g/dL, 3.1 g/dL, 2.8 g/dL and 2.6 g/dL. Transient liver elastography data showed some change from 65 kPa, 33 kPa, 34.8 kPa and undetectable to 46.4 kPa, 19.8 kPa, 29.1 kPa and 67.8 kPa at 4 weeks after transplantation in a patient with Wilson disease, a patient with HCV, and two patients with HBV. Ascites decreased in two patients. One of the patients with HBV underwent liver transplantation 4 months after the infusion, and the hepatic progenitor markers (cytokeratin [CD]-7, CD-8, CD-9, CD-18, CD-19, c-Kit and epithelial cell adhesion molecule [EpCAM]) were highly expressed in the explanted liver. ConclusionsSerum albumin levels, liver stiffness, liver volume, subjective healthiness and quality of life improved in the study patients. Although these findings were observed in a small population, the results may suggest a promising future for autologous CD34-depleted BM-MNC transplantation as a bridge to liver transplantation in patients with liver failure.

In vivo monitoring of arterially transplanted bone marrow mononuclear cells in a rat transient focal brain ischemia model using magnetic resonance imaging

Objectives: Intra-arterial transplantation of bone marrow mononuclear cells (BMMCs) effectively improves neuronal function and limits the infarct size. We monitored the fate of BMMCs labeled with super paramagnetic iron oxide (SPIO) until 7 days after the transplantation using high-field magnetic resonance imaging (MRI).Methods: Male Sprague–Dawley rats were subjected to 90-minute focal ischemia using the intraluminal suture technique followed by transplantation of 1×107 BMMCs or vehicle only via the ipsilateral carotid artery immediately after reperfusion. Autologous BMMCs were labeled with SPIO by electroporation prior to ischemia. MRI studies were performed at 1 hour, 24 hours, 3 days, and 7 days after reperfusion on each rat. The total infarct volume and the volume of negative dots were measured on T2-weighted images and T2*-weighted images, respectively.Results: One hour after BMMC transplantation, we confirmed wide spread distribution of BMMCs in the ischemic hemisphere as a negative dot. The volume of negative dot normalized by hemispheric volume decreased rapidly and was seldom seen at the seventh day after transplantation. The infarct volume was significantly smaller in the transplanted group than the vehicle group at 24 hours and 7 days after reperfusion.Discussion: The present study established in vivo monitoring of intra-arterial transplanted SPIO-labeled BMMCs immediately after reperfusion using MRI of a rat transient focal ischemia model. The accumulation of BMMCs in ischemic lesion at the acute stage of ischemia can be part of the conditions to limit the infarct size.

Bone-marrow mononuclear cells reduce neurodegeneration in hippocampal CA1 layer after transient global ischemia in rats

Global cerebral ischemia (GCI) results in death of the pyramidal neurons in the CA1 layer of the hippocampus. In this study we used the four-vessel occlusion (4VO) model of GCI to investigate a potential neuroprotective role of bone-marrow mononuclear cells (BMMCs) transplantation. BMMCs (3×107) were injected through the carotid artery, 1 or 3 days after ischemia (DAI), and the number of cells undergoing degeneration was investigated in brains at 7 DAI. A significant decrease in the number of dying cells was observed in the treated group, compared to animals treated with saline. Biodistribution of the injected cells (1 or 3 DAI) was investigated by 99mTechnetium labeling of the BMMCs and subsequent image analysis 2h after transplantation. In addition, the presence of CellTrace™-labeled BMMCs was investigated in tissue sections of the hippocampal area of these transplanted animals. BMMCs treatment significantly reduced the number of FJ-C positive cells in the hippocampal CA1 layer at 7 DAI. We also observed a decrease in the number of activated microglia/macrophage (ED1-positive cells) in the BMMCs-treated group compared with the untreated group. Our data show that BMMCs are able to modulate the microglial response and reduce neurodegeneration in the CA1 layer.

Comparison of the therapeutic effects of bone marrow mononuclear cells and microglia for permanent cerebral ischemia

In this study we transplanted bone marrow mononuclear cells (BM-MNCs) or microglia into rats that had undergone permanent cerebral ischemia and observed the distribution or morphology of transplanted cells in vivo. In addition, we compared the effects of BM-MNCs and microglia on infarct volume, brain water content, and functional outcome after permanent cerebral ischemia. BM-MNCs and microglia were obtained from femur and brain, respectively, of newborn rats. Adult rats were injected with vehicle or 3 million BM-MNCs or microglia via the tail vein 24h after permanent middle cerebral artery occlusion (pMCAO). The distribution or morphologic characteristics of transplanted BM-MNCs (double stained with BrdU/Cd34 or BrdU/CD45) and microglia (double stained with BrdU/Iba-1) were detected with immunofluorescent staining at 3 or 7 and 14 days after pMCAO. Functional deficits were assessed by the modified neurologic severity score at 1, 3, 7 and 14 days after pMCAO. Brain water content was assessed at 3 days, and infarct volume was determined at 14 days. We observed more BrdU/CD45 and BrdU/Iba-1 double-stained cells than BrdU/CD34 double-stained cells around the infarcted area. Some infused microglia showed the morphology of innate microglia at 7 days after pMCAO, and the number increased at 14 days. BM-MNC-treated rats showed significantly reduced infarct volume and brain water content compared to vehicle- and microglia-treated rats. In addition, BM-MNC treatment reduced neurologic deficit scores compared to those in the other groups. The results provide evidence that infusion of BM-MNCs, but not microglia, is neuroprotective after permanent cerebral ischemia.