Efficacy Evaluation and Tracking of Bone Marrow Stromal Stem Cells in a Rat Model of Renal Ischemia-Reperfusion Injury.

OBJECTIVES: The aim of this study was to evaluate the effects of bone marrow stromal stem cells (BMSCs) on renal ischemia-reperfusion injury (RIRI) and dynamically monitor engrafted BMSCs in vivo for the early prediction of their therapeutic effects in a rat model. METHODS: A rat model of RIRI was prepared by clamping the left renal artery for 45 min. One week after renal artery clamping, 2 × 106 superparamagnetic iron oxide- (SPIO-) labeled BMSCs were injected into the renal artery. Next, MR imaging of the kidneys was performed on days 1, 7, 14, and 21 after cell transplantation. On day 21, after transplantation, serum creatinine (Scr) and urea nitrogen (BUN) levels were assessed, and HE staining and TUNEL assay were also performed. RESULTS: The body weight growth rates in the SPIO-BMSC group were significantly higher than those in the PBS group (P < 0.05), and the Scr and BUN levels were also significantly lower than those in the PBS group (P < 0.05). HE staining showed that the degree of degeneration and vacuole-like changes in the renal tubular epithelial cells in the SPIO-BMSC group was significantly better than that observed in the PBS group. The TUNEL assay showed that the number of apoptotic renal tubular epithelial cells in the SPIO-BMSC group was significantly lower than that in the PBS group. The T2 value of the renal lesion was the highest on day 1 after cell transplantation, and it gradually decreased with time in both the PBS and SPIO-BMSC groups but was always the lowest in the SPIO-BMSC group. CONCLUSION: SPIO-labeled BMSC transplantation can significantly promote the recovery of RIRI and noninvasive dynamic monitoring of engrafted cells and can also be performed simultaneously with MRI in vivo for the early prediction of therapeutic effects.

A dual-mode nanoparticle based on natural biomaterials for photoacoustic and magnetic resonance imaging of bone mesenchymal stem cells in vivo.

Stem cell imaging in vivo is critical to elucidate the homing, distribution, survival, and repair mechanisms and to evaluate the therapeutic effects of engrafted stem cells. Unfortunately, unimodal imaging of stem cells does not simultaneously satisfy all current requirements owing to their intrinsic limitations. Obviously, bimodal or multimodal imaging of stem cells is a promising strategy for circumventing this issue. This study aimed to design and synthesize a novel dual-modal polyethylene glycol-modified magnetic nanoparticle (Fe3+-PEG-MNP) based on natural biomaterials including melanin and Fe ions for photoacoustic (PA) and magnetic resonance (MR) imaging of stem cells in vivo. The Fe3+-PEG-MNPs were characterized and their PA/MR imaging capability and cytotoxicity were evaluated. Bone marrow mesenchymal stem cells (BM-MSCs) labeled with Fe3+-PEG-MNPs were subjected to PA and MR imaging in vitro and in vivo. Consequently, Fe3+-PEG-MNPs displayed many superior properties, including ultra-small particle size, higher stability, water solubility, easy labeling of cells, lower cytotoxicity, high biosafety, excellent capability of PA/MR imaging, high sensitivity and long-term monitoring in vitro and in vivo. In particular, PA and MR signals of labeled BM-MSCs were maintained for at least 35 and 28 d, respectively, in vivo. Therefore, Fe3+-PEG-MNPs are ideal dual-modal PA/MR nanoparticles for non-invasive and effective monitoring of engrafted stem cells in vivo.

A promising magnetic resonance stem cell tracer based on natural biomaterials in a biological system: manganese(II) chelated to melanin nanoparticles.

BACKGROUND: Melanin and manganese are both indispensable natural substances that play crucial roles in the human body. Melanin has been used as a multimodality imaging nanoplatform for biology science research because of its natural binding ability with metal ions (eg, 64Cu2+, Fe3+, and Gd3+). Because of its effects on T1 signal enhancement, Mn-based nanoparticles have been used in magnetic resonance (MR) quantitative cell tracking in vivo. Stem cell tracking in vivo is an essential technology used to characterize engrafted stem cells, including cellular viability, biodistribution, differentiation capacity, and long-term fate. METHODS: In the present study, manganese(II) ions chelated to melanin nanoparticles [MNP-Mn(II)] were synthesized. The characteristics, stem cell labeling efficiency, and cytotoxicity of the nanoparticles were evaluated. MR imaging of the labeled stem cells in vivo and in vitro were also further performed. In T1 relaxivity (r1), MNP-Mn(II) were significantly more abundant than Omniscan. Bone marrow-derived stem cells (BMSCs) can be labeled easily by coincubating with MNP-Mn(II), suggesting that MNP-Mn(II) had high biocompatibility. RESULTS: Cell Counting Kit-8 assays revealed that MNP-Mn(II) had almost no cytotoxicity when used to label BMSCs, even with a very high concentration (1,600 µg/mL). BMSCs labeled with MNP-Mn(II) could generate a hyperintense T1 signal both in vitro and in vivo, and the hyperintense T1 signal in vivo persisted for at least 28 days. CONCLUSION: Taken together, our results showed that MNP-Mn(II) possessed many excellent properties for potential quantitative stem cell tracking in vivo.

Effective tracking of bone mesenchymal stem cells in vivo by magnetic resonance imaging using melanin-based gadolinium3+ nanoparticles.

Tracking transplanted stem cells is necessary to clarify cellular properties and improve transplantation success. In this study, we designed and synthesized melanin-based gadolinium3+ (Gd3+ )-chelate nanoparticles (MNP-Gd3+ ) of ∼7 nm for stem cell tracking in vivo. MNP-Gd3+ possesses many beneficial properties, such as its high stability and sensitivity, shorter T1 relaxation time, higher cell labeling efficiency, and lower cytotoxicity compared with commercial imaging agents. We found that the T1 relaxivity (r1 ) of MNP-Gd3+ was significantly higher than that of Gd-DTPA; the nanoparticles were taken up by bone mesenchymal stem cells (BMSCs) via endocytosis and were broadly distributed in the cytoplasm. Based on an in vitro MTT assay, no cytotoxicity of labeled stem cells was observed for MNP-Gd3+ concentrations of less than 800 µg/mL. Furthermore, we tracked MNP-Gd3+ -labeled BMSCs in vivo using 3.0T MRI equipment. After intramuscular injection, MNP-Gd3+ -labeled BMSCs were detected, even after four weeks, by 3T MRI. We concluded that MNP-Gd3+ nanoparticles at appropriate concentrations can be used to effectively monitor and track BMSCs in vivo. MNP-Gd3+ nanoparticles have potential as a new positive MRI contrast agent in clinical applications. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 131-137, 2017.

Effects of Magnetically Guided, SPIO-Labeled, and Neurotrophin-3 Gene-Modified Bone Mesenchymal Stem Cells in a Rat Model of Spinal Cord Injury.

Despite advances in our understanding of spinal cord injury (SCI) mechanisms, there are still no effective treatment approaches to restore functionality. Although many studies have demonstrated that transplanting NT3 gene-transfected bone marrow-derived mesenchymal stem cells (BMSCs) is an effective approach to treat SCI, the approach is often low efficient in the delivery of engrafted BMSCs to the site of injury. In this study, we investigated the therapeutic effects of magnetic targeting of NT3 gene-transfected BMSCs via lumbar puncture in a rat model of SCI. With the aid of a magnetic targeting cells delivery system, we can not only deliver the engrafted BMSCs to the site of injury more efficiently, but also perform cells imaging in vivo using MR. In addition, we also found that this composite strategy could significantly improve functional recovery and nerve regeneration compared to transplanting NT3 gene-transfected BMSCs without magnetic targeting system. Our results suggest that this composite strategy could be promising for clinical applications.

Visual bone marrow mesenchymal stem cell transplantation in the repair of spinal cord injury.

An important factor in improving functional recovery from spinal cord injury using stem cells is maximizing the number of transplanted cells at the lesion site. Here, we established a contusion model of spinal cord injury by dropping a weight onto the spinal cord at T7-8. Superparamagnetic iron oxide-labeled bone marrow mesenchymal stem cells were transplanted into the injured spinal cord via the subarachnoid space. An outer magnetic field was used to successfully guide the labeled cells to the lesion site. Prussian blue staining showed that more bone marrow mesenchymal stem cells reached the lesion site in these rats than in those without magnetic guidance or superparamagnetic iron oxide labeling, and immunofluorescence revealed a greater number of complete axons at the lesion site. Moreover, the Basso, Beattie and Bresnahan (BBB) locomotor rating scale scores were the highest in rats with superparamagnetic labeling and magnetic guidance. Our data confirm that superparamagnetic iron oxide nanoparticles effectively label bone marrow mesenchymal stem cells and impart sufficient magnetism to respond to the external magnetic field guides. More importantly, superparamagnetic iron oxide-labeled bone marrow mesenchymal stem cells can be dynamically and non-invasively tracked in vivo using magnetic resonance imaging. Superparamagnetic iron oxide labeling of bone marrow mesenchymal stem cells coupled with magnetic guidance offers a promising avenue for the clinical treatment of spinal cord injury.

High expression of transform growth factor beta 1 in gastric cancer confers worse outcome: results of a cohort study on 184 patients.

BACKGROUND/AIMS: Transform growth factors beta (TGFbeta) plays different roles at different stages of tumor development. TGFbeta1 is one isoform of TGFbeta, with complex secretion mechanism and bidirectional functions. This study was to investigate TGFbeta1 expression and its clinical significance in different clinicopathological subgroups of gastric cancer (GC) patients. METHODOLOGY: Tumor and peritumoral tissues from 184 GC patients were constructed into three tumor tissue microarrays. The expression of TGFbeta1 was analyzed by immunohistochemistry methods. RESULTS: TGFbeta1 was mainly expressed in the cytoplasm and membrane of GC cells. Low TGFbeta1 expression was observed in 82 (44.6%) tumor and 28 (68.3%) peritumoral tissues, and high expression was observed in 102 (55.4%) tumor and 13 (31.7%) peritumoral tissues. TGFbeta1 expression was significantly higher in tumor than peritumoral tissues (chi2 = 7.554, P = 0.006). The high expression of TGFbeta1 was related to worse overall survival (OS) (P = 0.040). TGFbeta1 expression was higher in the old and intestinal type GC than in the young (P = 0.017) and in diffuse type GC (P = 0.015), respectively. Patients with high TGFbeta1 expression had a worse survival in young people, female, diffuse type GC, poor differentiation, and lymph nodes metastasis. Multivariate Cox proportional hazards analysis showed that age, pathological grading, serosal invasion and TGFbeta1 expression were independent risk factors. CONCLUSIONS: High TGFbeta1 expression may indicate poor prognosis of GC patients and warrant more active treatment against TGFbeta1.

Long term follow up and retrospective study on 533 gastric cancer cases.

BACKGROUND: Gastric cancer (GC) is the third leading cause of cancer death in China and the outcome of GC patients is poor. The aim of the research is to study the prognostic factors of gastric cancer patients who had curative intent or palliative resection, completed clinical database and follow-up. METHODS: This retrospective study analyzed 533 GC patients from three tertiary referral teaching hospitals from January 2004 to December 2010 who had curative intent or palliative resection, complete clinical database and follow-up information. The GC-specific overall survival (OS) status was determined by the Kaplan-Meier method, and univariate analysis was conducted to identify possible factors for survival. Multivariate analysis using the Cox proportional hazard model and a forward regression procedure was conducted to define independent prognostic factors. RESULTS: By the last follow-up, the median follow-up time of 533 GC patients was 38.6 mo (range 6.9-100.9 mo), and the median GC-specific OS was 25.3 mo (95% CI: 23.1-27.4 mo). The estimated 1-, 2-, 3- and 5-year GC-specific OS rates were 78.4%, 61.4%, 53.3% and 48.4%, respectively. Univariate analysis identified the following prognostic factors: hospital, age, gender, cancer site, surgery type, resection type, other organ resection, HIPEC, LN status, tumor invasion, distant metastases, TNM stage, postoperative SAE, systemic chemotherapy and IP chemotherapy. In multivariate analysis, seven factors were identified as independent prognostic factors for long term survival, including resection type, HIPEC, LN status, tumor invasion, distant metastases, postoperative SAE and systemic chemotherapy. CONCLUSIONS: Resection type, HIPEC, postoperative SAE and systemic chemotherapy are four independent prognostic factors that could be intervened for GC patients for improving survival.

Transplantation of neurotrophin-3-expressing bone mesenchymal stem cells improves recovery in a rat model of spinal cord injury.

BACKGROUND: This study aimed to investigate the therapeutic effects of transplanting neutrophin-3 (NT-3)-expressing bone marrow-derived mesenchymal stem cells (BMSCs) in a rat model of spinal cord injury (SCI). METHODS: Forty-eight adult female Sprague-Dawley rats were randomly assigned to three groups: the control, BMSC, and NT-3-BMSC groups. BMSCs were infected with NT-3-DsRed or DsRed lentivirus and injected into the cerebrospinal fluid (CSF) via lumbar puncture (LP) 7 days after SCI in the NT-3-BMSC and BMSC groups, respectively. The hind-limb motor function of all rats was recorded using the Basso, Beattie, and Bresnahan (BBB) locomotor rating scale on days 1, 3, 7, 14, 21, 28, and 35 after transplantation. Haematoxylin-eosin (HE) staining, immunofluorescence labelling, and western blotting were performed at the final time point. RESULTS: Expressions of NT-3, brain-derived neurotrophic factor (BDNF), and vascular endothelial growth factor (VEGF) proteins increased significantly in the NT-3-BMSC group, and hind-limb locomotor functions improved significantly in the NT-3-BMSC group compared with the other two groups. The cystic cavity area was smallest in the NT-3-BMSC group. In the NT-3-BMSC group, neurofilament 200 (NF200) and glial fibrillary acidic protein (GFAP) expression levels around the lesions were significantly increased and decreased, respectively. CONCLUSIONS: Our findings demonstrate that transplantation of NT-3 gene-modified BMSCs via LP can strengthen the therapeutic benefits of BMSC transplantation. We observed that these modified cells increased locomotor function recovery, promoted nerve regeneration, and improved the injured spinal cord microenvironment, suggesting that it could be a promising treatment for SCI.

Tumor invasion unit in gastric cancer revealed by QDs-based in situ molecular imaging and multispectral analysis.

In tumor tissues, cancer cells, tumor infiltrating macrophages and tumor neo-vessels in close spatial vicinity with one another form tumor invasion unit, which is a biologically important tumor microenvironment of metastasis to facilitate cancer invasion and metastasis. Establishing an in situ molecular imaging technology to simultaneously reveal these three components is essential for the in-depth investigation of tumor invasion unit. In this report, we have developed a computer-aided algorithm by quantum dots (QDs)-based multiplexed molecular imaging technique for such purpose. A series of studies on gastric cancer tumor tissues demonstrated that the tumor invasion unit was correlated with major unfavorable pathological features and worse clinical outcomes, which illustrated the significantly negative impacts and predictive power of tumor invasion unit on patient overall survival. This study confirmed the technical advantages of QDs-based in situ and simultaneous molecular imaging of key cancer molecules to gain deeper insights into the biology of cancer invasion.

Lymph node ratio is a better prognosticator than lymph node status for gastric cancer: A retrospective study of 138 cases.

To study the clinical significance of lymph node ratio (LNR) in gastric cancer (GC), this study analyzed 613 patients with GC who underwent surgical resection. Of 613 patients with GC, 138 patients who had >15 lymph nodes (LNs) resected and radical resection were enrolled into the final study. All major clinicopathological data were entered into a central database. LNR was defined as the ratio of the number of metastatic LNs to the number of removed LNs. In order to determine the best cut-off points for LNR, the log-rank test and X-tile were used. LNR was then substituted for lymph node status (pN) in the 7th American Joint Committee on Cancer tumor-node-metastases (TNM) staging system and this was defined as the tumor-node ratio-metastases (TRM) staging system. Pearson's correlation coefficient (r) was used to study the correlations among the number of removed LNs, pN and LNR. The Kaplan-Meier survival curve was used to study the survival status, and the log-rank test and Cox proportional hazards model were used to identify the independent factors for survival. Receiver operating characteristic curve analysis was used to determine the predictive value of the parameters. By the time of last follow-up (median follow-up period, 38.3 months; range, 9.9-97.7 months), the median overall survival (OS) was 23.9 months [95% confidence interval (CI), 18.8-29.0 months]. The 1-, 2-, 3- and 5-year survival rates were 76.8, 57.2, 50.0 and 46.4%, respectively. The cut-off points were 0, 0.5 and 0.8 (R0, LNR=0; R1, LNR ≤0.5; R2, 0.5> LNR ≤0.8; and R3, LNR >0.8). Univariate and multivariate analyses revealed that both LNR and pN were independent prognostic factors for GC. LNR could better differentiate OS in patients than LN. In addition, the TRM staging system was better at predicting the clinical outcomes than the TNM staging system, and LNR was better than pN. In conclusion, LNR was a better prognosticator than pN for GC.