Search

ObjectiveStromal barriers, such as the abundant desmoplastic stroma that is characteristic of pancreatic ductal adenocarcinoma (PDAC), can block the delivery and decrease the tumour-penetrating ability of therapeutics such as tumour necrosis factor-related apoptosis-inducing ligand (TRAIL), which can selectively induce cancer cell apoptosis. This study aimed to develop a TRAIL-based nanotherapy that not only eliminated the extracellular matrix barrier to increase TRAIL delivery into tumours but also blocked antiapoptotic mechanisms to overcome TRAIL resistance in PDAC.DesignNitric oxide (NO) plays a role in preventing tissue desmoplasia and could thus be delivered to disrupt the stromal barrier and improve TRAIL delivery in PDAC. We applied an in vitro–in vivo combinatorial phage display technique to identify novel peptide ligands to target the desmoplastic stroma in both murine and human orthotopic PDAC. We then constructed a stroma-targeted nanogel modified with phage display-identified tumour stroma-targeting peptides to co-deliver NO and TRAIL to PDAC and examined the anticancer effect in three-dimensional spheroid cultures in vitro and in orthotopic PDAC models in vivo.ResultsThe delivery of NO to the PDAC tumour stroma resulted in reprogramming of activated pancreatic stellate cells, alleviation of tumour desmoplasia and downregulation of antiapoptotic BCL-2 protein expression, thereby facilitating tumour penetration by TRAIL and substantially enhancing the antitumour efficacy of TRAIL therapy.ConclusionThe co-delivery of TRAIL and NO by a stroma-targeted nanogel that remodels the fibrotic tumour microenvironment and suppresses tumour growth has the potential to be translated into a safe and promising treatment for PDAC.

The short supply of donor corneas is exacerbated by the unsuitability of donors with insufficient endothelial cell density. Few studies have investigated promoting corneal endothelial cell proliferation to increase the endothelial cell density. We hypothesize that pre‐transplantation treatment of proliferative tissue‐cultivated corneas may increase corneal endothelial cell density. We observed that the airlift cultures were superior to immersion cultures with respect to both transparency and thickness. In this tissue culture system, we observed that lysophosphatidic acid increased the rabbit corneal endothelial cell density, number of BrdU‐positive cells and improve wound healing. We also observed an indirect effect of lysophosphatidic acid on corneal endothelial cell proliferation mediated by the stimulation of interleukin‐1β secretion from stromal cells. Human corneal tissues treated with lysophosphatidic acid or interleukin‐1β contained significantly more Ki‐67‐positive cells than untreated group. The lysophosphatidic acid‐ or interleukin‐1β‐treated cultured tissue remained hexagon‐shaped, with ZO‐1 expression and no evidence of the endothelial‐mesenchymal transition. Our novel protocol of tissue culture may be applicable for eye banks to optimize corneal grafting.

After the discovery of endogenous dinitrosyl iron complexes (DNICs) as a potential biological equivalent of nitric oxide (NO), bioinorganic engineering of [Fe(NO)2] unit has emerged to develop biomimetic DNICs [(NO)2Fe(L)2] as a chemical biology tool for controlled delivery of NO. For example, water-soluble DNIC [Fe2(μ-SCH2CH2OH)2(NO)4] (DNIC-1) was explored for oral delivery of NO to the brain and for the activation of hippocampal neurogenesis. However, the kinetics and mechanism for cellular uptake and intracellular release of NO, as well as the biocompatibility of synthetic DNICs, remain elusive. Prompted by the potential application of NO to dermato-physiological regulations, in this study, cellular uptake and intracellular delivery of DNIC [Fe2(μ-SCH2CH2COOH)2(NO)4] (DNIC-2) and its regulatory effect/biocompatibility toward epidermal cells were investigated. Upon the treatment of DNIC-2 to human fibroblast cells, cellular uptake of DNIC-2 followed by transformation into protein-bound DNICs occur to trigger the intracellular release of NO with a half-life of 1.8 ± 0.2 h. As opposed to the burst release of extracellular NO from diethylamine NONOate (DEANO), the cell-penetrating nature of DNIC-2 rationalizes its overwhelming efficacy for intracellular delivery of NO. Moreover, NO-delivery DNIC-2 can regulate cell proliferation, accelerate wound healing, and enhance the deposition of collagen in human fibroblast cells. Based on the in vitro and in vivo biocompatibility evaluation, biocompatible DNIC-2 holds the potential to be a novel active ingredient for skincare products.

Objective: To assess the effects of exoskeleton robot-assisted passive range of motion for induction training in combination with conventional hand rehabilitation in patients with chronic stroke.Design: Single-cohort feasibility study.Subjects: Chronic stroke with severe upper extremity hemiparesis.Methods: Thirty sessions of therapy over a period of 10 weeks. Each session started with 30 min robot-assisted passive range of motion for the hand, followed by 30 min conventional hand rehabilitation. The Fugl-Meyer Assessment for upper extremity, arm subscore of Motricity Index, Functional Independence Measure and Fugl-Meyer assessment for sensation (Fugl-Meyer assessment-sensory) were conducted at pre-intervention (pre) and after the 16th (16-post) and 30th (30-post) sessions of interventions.Results: Twelve patients with chronic stroke were recruited. The Fugl-Meyer assessment for upper extremity (16-post vs 30-post, p = 0.011), arm subscore of Motricity Index (pre vs 30-post, p = 0.012) and Functional Independence Measure (pre vs 30-post, p = 0.007; 16- post vs 30-post, p = 0.016) improved significantly after the therapy. However, FMA-sensory did not change significantly.Conclusion: Exoskeleton robot-assisted passive range of motion of the hand using an exoskeleton can be considered as an induction therapy before starting conventional therapy for hand rehabilitation in patients with chronic stroke. Further randomized control trials are needed to verify the therapeutic benefits. LAY ABSTRACTMotor recovery of hand dexterity is challenging during the chronic phase of stroke. Patients achieve different levels of hand function during the acute or subacute phase of stroke. Those receiving conventional physical therapy during the chronic phase of stroke usually experience difficulty in hand dexterity improvement after achieving motor recovery plateau. This pilot study investigated the effects of robot-assisted passive range of motion training in combination with conventional rehabilitation on hand function in a cohort of patients with chronic stroke who underwent follow-up at an outpatient rehabilitation clinic. The affected upper extremity function, strength and general function improved significantly after the therapy for the 12 patients recruited to this study. Using robot-assisted passive range of motion training as an induction therapy in combination with conventional rehabilitation may be beneficial for patients with chronic stoke who have impairment of hand function.

Background The pumping function of corneal endothelial cells (CECs) plays a pivotal role in the maintenance of corneal water homeostasis. Corneal endothelial dysfunction (CED) leads to corneal edema and opacity, but with the exception of keratoplasty, no optimal therapeutic strategies have been established for CED. In this study, we aimed to investigate the ameliorative effect of ascorbic acid (AA) on CED and the underlying mechanism of action in the corneal endothelium. Methods Rabbit corneal endothelial damage was induced by anterior chamber injection of benzalkonium chloride (BAK). AA was topically administered to the corneal surface, and the transparency and thickness of the cornea were assessed by external eye photography, slit-lamp photography, and ultrasonic pachymetry. To further analyze the mechanism, rabbit CECs and immortalized human CECs (B4G12 cells) were cultured. A ferric reducing/antioxidant and AA (FRASC) assay was performed to measure the AA concentration. Cell proliferation was evaluated by cell counting and bromodeoxyuridine (BrdU) labeling assays, and protein expression was examined by liquid chromatography–mass spectrometry (LC/MS) and immunoblotting. The involvement of glucose transporter 1 (GLUT1) and phospho-ERK was evaluated via GLUT1-siRNA and phospho-ERK inhibitor (PD98059) treatment. Interpretation We observed that topical AA ameliorates BAK-induced rabbit corneal endothelial damage. Furthermore, we demonstrated that AA is transported into B4G12 cells via GLUT1, and afterward, AA increases ERK phosphorylation and promotes cell proliferation. Our findings indicate that CEC proliferation stimulated via the noncanonical AA-GLUT1-ERK axis contributes to AA-enhanced healing of CED.

The major biological methyl donor, S-adenosylmethionine (adoMet) synthesis occurs mainly in the liver. Methionine adenosyltransferase 1A (MAT1A) and glycine N-methyltransferase (GNMT) are two key enzymes involved in the functional implications of that variation. We collected 42 RNA-seq data from paired hepatocellular carcinoma (HCC) and its adjacent normal liver tissue from the Cancer Genome Atlas (TCGA). There was no mutation found in MAT1A or GNMT RNA in the 42 HCC patients. The 11,799 genes were annotated in the RNA-Seq data, and their expression levels were used to investigate the phenotypes of low MAT1A and low GNMT by Gene Set Enrichment Analysis (GSEA). The REACTOME_TRANSLATION gene set was enriched and visualized in a heatmap along with corresponding differences in gene expression between low MAT1A versus high MAT1A and low GNMT versus high GNMT. We identified 43 genes of the REACTOME_TRANSLATION gene set that are powerful prognosis factors in HCC. The significantly predicted genes were referred into eukaryotic translation initiation (EIF3B, EIF3K), eukaryotic translation elongation (EEF1D), and ribosomal proteins (RPs). Cell models expressing various MAT1A and GNMT proved that simultaneous restoring the expression of MAT1A and GNMT decreased cell proliferation, invasion, as well as the REACTOME_TRANSLATION gene EEF1D, consistent with a better prognosis in human HCC. We demonstrated new findings that downregulation or defect in MAT1A and GNMT genes can enrich the protein-associated translation process that may account for poor HCC prognosis. This is the first study demonstrated that MAT1A and GNMT, the 2 key enzymes involved in methionine cycle, could attenuate the function of ribosome translation. We propose a potential novel mechanism by which the diminished GNMT and MAT1A expression may confer poor prognosis for HCC.

Background and aims Renal dysfunction is a potent risk factor for cardiovascular diseases, including stroke. This study aimed to evaluate the impact of admission estimated glomerular filtration rate (eGFR) levels on short-term (1-month) and long-term (1-year) mortality in patients with acute ischemic stroke. Methods From the Taiwan Stroke Registry data, we classified ischemic stroke patients, identified from April 2006 to December 2015, into 5 groups by eGFR at admission: ≥ 90, 60–89, 30–59, 15–29, and Results Among 52,732 ischemic stroke patients, 1480 died within one month. The 1-month mortality rate was over 5-fold greater in patients with eGFR Conclusions After acute ischemic stroke, patients with reduced eGFR are at elevated risks of short-term and long-term deaths in a graded relationship.

Multicellular spheroids show three-dimensional (3D) organization with extensive cell–cell and cell–extracellular matrix interactions. Owing to their native tissue-mimicking characteristics, mesenchymal stem cell (MSC) spheroids are considered promising as implantable therapeutics for stem cell therapy. Herein, we aim to further enhance their therapeutic potential by tuning the cultivation parameters and thus the inherent niche of 3D MSC spheroids. Significantly increased expression of multiple pro-regenerative paracrine signaling molecules and immunomodulatory factors by MSCs was observed after optimizing the conditions for spheroid culture. Moreover, these alterations in cellular behaviors may be associated with not only the hypoxic niche developed in the spheroid core but also with the metabolic reconfiguration of MSCs. The present study provides efficient methods for manipulating the therapeutic capacity of 3D MSC spheroids, thus laying solid foundations for future development and clinical application of spheroid-based MSC therapy for regenerative medicine.

We investigated the impact of serum cholesterol levels on 30-day mortality after ischemic stroke in dialysis patients.From the Taiwan Stroke Registry data, we identified 46,770 ischemic stroke cases, including 1101 dialysis patients and 45,669 nondialysis patients from 2006 to 2013.Overall, the 30-day mortality was 1.46-fold greater in the dialysis group than in the nondialysis group (1.75 versus 1.20 per 1000 person-days). The mortality rates were 1.64, .62, 2.82, and 2.23 per 1000 person-days in dialysis patients with serum total cholesterol levels of 120 mg/dL, 120-159 mg/dL, 160-199 mg/dL, and ≥200 mg/dL, respectively. Compared to dialysis patients with serum total cholesterol levels of 120-159 mg/dL, the corresponding adjusted hazard ratios of mortality were 4.20 (95% confidence interval [CI] = 1.01-17.4), 8.06 (95% CI = 2.02-32.2), and 6.89 (95% CI = 1.59-29.8) for those with cholesterol levels of 120 mg/dL, 160-199 mg/dL, and ≥200 mg/dL, respectively.Dialysis patients with serum total cholesterol levels of ≥160 mg/dL or 120 mg/dL on admission are at an elevated hazard of 30-day mortality after ischemic stroke.

Compromised pumping function of the corneal endothelium, due to loss of endothelial cells, results in corneal edema and subsequent visual problems. Clinically and experimentally, oxidative stress may cause corneal endothelial decompensation after phacoemulsification. Additionally, in vitro and animal studies have demonstrated the protective effects of intraoperative infusion of ascorbic acid (AA). Here, we established a paraquat-induced cell damage model, in which paraquat induced reactive oxygen species (ROS) production and apoptosis in the B4G12 and ARPE-19 cell lines. We demonstrate that oxidative stress triggered autophagic flux blockage in corneal endothelial cells and that addition of AA ameliorated such oxidative damage. We also demonstrate the downregulation of Akt phosphorylation in response to oxidative stress. Pretreatment with ascorbic acid reduced the downregulation of Akt phosphorylation, while inhibition of the PI3K/Akt pathway attenuated the protective effects of AA. Further, we establish an in vivo rabbit model of corneal endothelial damage, in which an intracameral infusion of paraquat caused corneal opacity. Administration of AA via topical application increased its concentration in the corneal stroma and reduced oxidative stress in the corneal endothelium, thereby promoting corneal clarity. Our findings indicate a perioperative strategy of topical AA administration to prevent oxidative stress-induced damage, particularly for those with vulnerable corneal endothelia.

Islet transplantation has been demonstrated to be a promising therapy for type 1 diabetes mellitus. Although it is a minimally invasive operating procedure and provides easy access for graft monitoring, subcutaneous transplantation of the islet only has limited therapeutic outcomes, owing to the poor capacity of skin tissue to foster revascularization in a short period. Herein, 3D cell spheroids of clinically accessible umbilical cord blood mesenchymal stem cells and human umbilical vein endothelial cells are formed and employed for codelivery with β cells subcutaneously. The 3D stem cell spheroids, which can secrete multiple proangiogenic and prosurvival growth factors, induce robust angiogenesis and prevent β cell graft death, as indicated by the results of in vivo bioluminescent tracking and histological analysis. These experimental data highlight the efficacy of the 3D stem cell spheroids that are fabricated using translationally applicable cell types in promoting the survival and function of subcutaneously transplanted β cells.

In the absence of adequate oxygen, cancer cells that are grown in hypoxic solid tumors resist treatment using antitumor drugs (such as doxorubicin, DOX), owing to their attenuated intracellular production of reactive oxygen species (ROS). Hyperbaric oxygen (HBO) therapy favorably improves oxygen transport to the hypoxic tumor tissues, thereby increasing the sensitivity of tumor cells to DOX. However, the use of HBO with DOX potentiates the ROS-mediated cytotoxicity of the drug toward normal tissues. In this work, we hypothesize that regional oxygen treatment by an implanted oxygen-generating depot may enhance the cytotoxicity of DOX against malignant tissues in a highly site-specific manner, without raising systemic oxygen levels. Upon implantation close to the tumor, the oxygen-generating depot reacts with the interstitial medium to produce oxygen in situ, effectively shrinking the hypoxic regions in the tumor tissues. Increasing the local availability of oxygen causes the cytotoxicity of DOX that is accumulated in the tumors to be significantly enhanced by the elevated production of ROS, ultimately allaying the hypoxia-induced DOX resistance in solid malignancies. Importantly, this enhancement of cytotoxicity is limited to the site of the tumors, and this feature of the system that is proposed herein is unique.

Patients with diabetes mellitus are prone to develop refractory wounds. They exhibit reduced synthesis and levels of circulating hydrogen sulfide (H

Cell transplantation via direct intramuscular injection is a promising therapy for patients with ischemic diseases. However, following injections, retention of transplanted cells in engrafted areas remains problematic, and can be deleterious to cell-transplantation therapy. In this Progress Report, a thermoresponsive hydrogel system composed of aqueous methylcellulose (MC) blended with phosphate-buffered saline is constructed to grow cell sheet fragments and cell bodies for the treatment of ischemic diseases. The as-prepared MC hydrogel system undergoes a sol-gel reversible transition upon heating or cooling at ≈32 °C. Via this unique property, the grown cell sheet fragments (cell bodies) can be harvested without using proteolytic enzymes; consequently, their inherent extracellular matrices (ECMs) and integrative adhesive agents remain well preserved. In animal studies using rats and pigs with experimentally created myocardial infarction, the injected cell sheet fragments (cell bodies) become entrapped in the interstices of muscular tissues and adhere to engraftment sites, while a minimal number of cells exist in the group receiving dissociated cells. Moreover, transplantation of cell sheet fragments (cell bodies) significantly increases vascular density, thereby improving the function of an infarcted heart. These experimental results demonstrate that cell sheet fragments (cell bodies) function as a cell-delivery construct by providing a favorable ECM environment to retain transplanted cells locally and consequently, improving the efficacy of therapeutic cell transplantation.

This article reviews recent progress in the development of carbon dioxide (CO

Cellular cardiomyoplasty has been limited by poor graft retention after cell transplantation. To ensure good retention of the engrafted cells, a microfluidic device was used to fabricate spherical porous beads of poly(D,L-lactic-co-glycolic acid) as a platform for cell delivery. The beads thus obtained had a relatively uniform size, a highly porous structure, and a favorably interconnected interior architecture, to facilitate the transportation of oxygen and nutrients. These porous beads were loaded with human amniotic fluid stem cells (hAFSCs) to generate cellularized microscaffolds. Live/dead assay demonstrated that most of the cells in the porous constructs were viable. The hAFSCs that were grown in beads formed a complex three-dimensional organization with well-preserved extracellular matrices (ECM) according to their porous structure. Retention of the administered beads was clearly identified at the site of engraftment following an experimentally induced myocardial infarction in a rat model. The results of echocardiography, magnetic resonance imaging, and histological analyses suggest that the transplantation of hAFSC beads into an infarcted heart could effectively maintain its gross morphology, prevent successive ventricular expansion, and thereby improve the post-infarcted cardiac function. Immunofluorescent staining revealed that the microenvironment that was provided by the infarcted myocardium might offer cues for the induction of the engrafted hAFSCs into angiogenic and cardiomyogenic lineages. Our results demonstrate that the cellularized beads with endogenously secreted ECM were of sufficient physical size to be entrapped in the interstitial tissues following transplantation, thereby benefiting the infarcted heart.

Cell transplantation is a promising strategy for therapeutic treatment of ischemic heart diseases. In this study, cord blood mesenchymal stem cells (cbMSCs) and human umbilical vein endothelial cells (HUVECs) in the form of core-shell bodies (cbMSC/HUVEC bodies) were prepared to promote vascularization and restore heart functions in an experimentally-created myocardial infarction (MI) rat model. Saline, cbMSC bodies and HUVEC bodies were used as controls. In vitro results indicated that cbMSC/HUVEC bodies possessed the capability of heterotypic assembly of cbMSCs and HUVECs into robust and durable tubular networks on Matrigel. The up-regulated gene expressions of VEGF and IGF-1 reflected the robust expansion of tubular networks; in addition, the augmented levels of SMA and SM22 suggested smooth muscle differentiation of cbMSCs, possibly helping to improve the durability of networks. Moreover, according to the in vivo echocardiographic, magnetic resonance and computed-tomographic results, transplantation of cbMSC/HUVEC bodies benefited post-MI dysfunction. Furthermore, the vascularization analyses demonstrated the robust vasculogenic potential of cbMSC/HUVEC bodies in vivo, thus contributing to the greater viable myocardium and the less scar region, and ultimately restoring the cardiac function. The concept of core-shell bodies composed of perivascular cells and endothelial cells may serve as an attractive cell delivery vehicle for vasculogenesis, thus improving the cardiac function significantly.

A recurring obstacle in cell-base strategies for treating ischemic diseases is the significant loss of viable cells that is caused by the elevated levels of regional reactive oxygen species (ROS), which ultimately limits therapeutic capacity. In this study, aggregates of human umbilical vein endothelial cells (HUVECs) and cord-blood mesenchymal stem cells (cbMSCs), which are capable of inducing therapeutic angiogenesis, are prepared. We hypothesize that the concurrent delivery of an antioxidant N-acetylcysteine (NAC) may significantly increase cell retention following the transplantation of HUVEC/cbMSC aggregates in a mouse model with hindlimb ischemia. Our in vitro results demonstrate that the antioxidant NAC can restore ROS-impaired cell adhesion and recover the reduced angiogenic potential of HUVEC/cbMSC aggregates under oxidative stress. In the animal study, we found that by scavenging the ROS generated in ischemic tissues, NAC is likely to be able to establish a receptive cell environment in the early stage of cell transplantation, promoting the adhesion, retention, and survival of cells of engrafted aggregates. Therapeutic angiogenesis is therefore enhanced and blood flow recovery and limb salvage are ultimately achieved. The combinatory strategy that uses an antioxidant and HUVEC/cbMSC aggregates may provide a new means of boosting the therapeutic efficacy of cell aggregates for the treatment of ischemic diseases.

Although the induction of neovascularization by cell-based approaches has demonstrated substantial potential in treating myocardial infarction (MI), the process of cell-mediated angiogenesis and its correlation with therapeutic mechanisms of cardiac repair remain elusive. In this work, three-dimensional (3D) aggregates of human umbilical vein endothelial cells (HUVECs) and cord-blood mesenchymal stem cells (cbMSCs) are constructed using a methylcellulose hydrogel system. By maximizing cell-cell and cell-ECM communications and establishing a hypoxic microenvironment in their inner cores, these cell aggregates are capable of forming widespread tubular networks together with the angiogenic marker αvβ3 integrin; they secret multiple pro-angiogenic, pro-survival, and mobilizing factors when grown on Matrigel. The aggregates of HUVECs/cbMSCs are exogenously engrafted into the peri-infarct zones of rats with MI via direct local injection. Multimodality noninvasive imaging techniques, including positron emission tomography, single photon emission computed tomography, and echocardiography, are employed to monitor serially the beneficial effects of cell therapy on angiogenesis, blood perfusion, and global/regional ventricular function, respectively. The myocardial perfusion is correlated with ventricular contractility, demonstrating that the recovery of blood perfusion helps to restore regional cardiac function, leading to the improvement in global ventricular performance. These experimental data reveal the efficacy of the exogenous transplantation of 3D cell aggregates after MI and elucidate the mechanism of cell-mediated therapeutic angiogenesis for cardiac repair.

Background Discharge disposition planning is vital for poststroke patients. We investigated clinical factors associated with discharging patients to nursing homes, using the Taiwan Stroke Registry data collected from 39 major hospitals. Methods We randomly assigned 21,575 stroke inpatients registered from 2006 to 2008 into derivation and validation groups at a 3-to-1 ratio. We used the derivation group to develop a prediction model by measuring cumulative risk scores associated with potential predictors: age, sex, hypertension, diabetes mellitus, heart diseases, stroke history, snoring, main caregivers, stroke types, and National Institutes of Health Stroke Scale (NIHSS). Probability of nursing home care and odds ratio (OR) of nursing home care relative to home care by cumulative risk scores were measured for the prediction. The area under the receiver operating characteristic curve (AUROC) was used to assess the model discrimination against the validation group. Results Except for hypertension, all remaining potential predictors were significant independent predictors associated with stroke patient disposition to nursing home care after discharge from hospitals. The risk sharply increased with age and NIHSS. Patients with a cumulative risk score of 15 or more had an OR of 86.4 for the nursing home disposition. The AUROC plots showed similar areas under curves for the derivation group (.86, 95% confidence interval [CI], .85-.87) and for the validation group (.84, 95% CI, .83-.86). Conclusions The cumulative risk score is an easy-to-estimate tool for preparing stroke patients and their family for disposition on discharge.

Cell-based therapeutic neovascularization is a promising method for treating ischemic disorders. In this work, human umbilical vein endothelial cells (HUVECs) were thoroughly premixed with cord-blood mesenchymal stem cells (cbMSCs) and cultivated to form three-dimensional (3D) cell aggregates for cellular cardiomyoplasty. In the in vitro study, tubular networks were formed at day 1 after the co-culturing of dissociated HUVECs and cbMSCs on Matrigel; however, as time progressed, the grown tubular networks regressed severely. Conversely, when 3D cell aggregates were grown on Matrigel, mature and stable tubular networks were observed over time, under the influence of their intensive cell-extracellular matrix (ECM) interactions and cell-cell contacts. 3D cell aggregates were transplanted into the peri-infarct zones of rats with myocardial infarction (MI) via direct intramyocardial injection. Based on our pinhole single photon emission computed tomography (SPECT) myocardial-perfusion observations, echocardiographic heart-function examinations and histological analyses, the engrafted 3D cell aggregates considerably enhanced the vascular densities and the blood flow recovery in the ischemic myocardium over those of their dissociated counterparts, thereby reducing the size of perfusion defects and restoring cardiac function. These results demonstrate that the intramuscular delivery of 3D cell aggregates of HUVECs/cbMSCs can be a valuable cell-based regenerative therapeutic strategy against MI.

The proximity of cells in three-dimensional (3D) organization maximizes the cell-cell communication and signaling that are critical for cell function. In this study, 3D cell aggregates composed of human umbilical vein endothelial cells (HUVECs) and cord-blood mesenchymal stem cells (cbMSCs) were used for therapeutic neovascularization to rescue tissues from critical limb ischemia. Within the cell aggregates, homogeneously mixed HUVECs and cbMSCs had direct cell-cell contact with expressions of endogenous extracellular matrices and adhesion molecules. Although dissociated HUVECs/cbMSCs initially formed tubular structures on Matrigel, the grown tubular network substantially regressed over time. Conversely, 3D HUVEC/cbMSC aggregates seeded on Matrigel exhibited an extensive tubular network that continued to expand without regression. Immunostaining experiments show that, by differentiating into smooth muscle cell (SMC) lineages, the cbMSCs stabilize the HUVEC-derived tubular network. The real-time PCR analysis results suggest that, through myocardin, TGF-β signaling regulates the differentiation of cbMSCs into SMCs. Transplantation of 3D HUVEC/cbMSC aggregates recovered blood perfusion in a mouse model of hindlimb ischemia more effectively compared to their dissociated counterparts. The experimental results confirm that the transplanted 3D HUVEC/cbMSC aggregates enhanced functional vessel formation within the ischemic limb and protected it from degeneration. The 3D HUVEC/cbMSC aggregates can therefore facilitate the cell-based therapeutic strategies for modulating postnatal neovascularization.