Development of a MET-targeted single-chain antibody fragment as an anti-oncogene targeted therapy for breast cancer.

The usage of monoclonal antibodies (mAbs) and antibody fragments, as a matter associated with the biopharmaceutical industry, is increasingly growing. Harmonious with this concept, we designed an exclusive modeled single-chain variable fragment (scFv) against mesenchymal-epithelial transition (MET) oncoprotein. This scFv was newly developed from Onartuzumab sequence by gene cloning, and expression using bacterial host. Herein, we examined its preclinical efficacy for the reduction of tumor growth, invasiveness and angiogenesis in vitro and in vivo. Expressed anti-MET scFv demonstrated high binding capacity (48.8%) toward MET-overexpressing cancer cells. The IC50 value of anti-MET scFv against MET-positive human breast cancer cell line (MDA-MB-435) was 8.4 µg/ml whereas this value was measured as 47.8 µg/ml in MET-negative cell line BT-483. Similar concentrations could also effectively induce apoptosis in MDA-MB-435 cancer cells. Moreover, this antibody fragment could reduce migration and invasion in MDA-MB-435 cells. Grafted breast tumors in Balb/c mice showed significant tumor growth suppression as well as reduction of blood-supply in response to recombinant anti-MET treatment. Histopathology and immunohistochemical assessments revealed higher rate of response to therapy. In our study, we designed and synthetized a novel anti-MET scFv which could effectively suppress MET-overexpressing breast cancer tumors.

Development of an RNA sequencing-based prognostic gene signature in multiple myeloma.

Several prognostic gene signatures have been developed to predict the clinical outcome in patients with multiple myeloma (MM). The most salient disadvantage of the previous signatures is their non-reproducibility in external datasets. Given the disadvantages and the superiority of RNA sequencing over microarrays in transcriptome profiling to produce more reliable outputs, we sought to develop a reproducible RNA sequencing-based prognostic gene signature for MM. Genes significantly associated with survival were detected in The Cancer Genome Atlas (TCGA) MM RNA sequencing dataset (MMRF-CoMMpass) (n = 412) through a strict pipeline containing four rigid filters. The reproducibility of the selected genes was checked in an independent dataset (GSE24080), containing 559 newly diagnosed patients with MM. The RNA sequencing-based prognostic signature was reconstructed based on the final genes in the training dataset (MMRF-CoMMpass) and externally validated in five independent datasets (i.e. GSE2658, GSE13624, GSE9782, GSE6477 and GSE57317), containing 1461 MM cases. The RNA sequencing-based signature was reconstructed using finally five reproducible genes: CCT2, CKS1B, PRKDC, NONO and UBE2A. This signature was able to robustly discriminate between low- and high-risk patients in both training and validation datasets (Ps ≤ 0·001). Our signature was also independent of and more powerful than the routine MM prognostic factors (i.e. ß2-microglobulin, albumin, age and sex) (Ps ≤ 0·01). Treatment regimens had no effect on RNA sequencing-based signature insofar as this signature succeeded in predicting the clinical outcome in various treatment groups (Ps ≤ 0·001).

Macrophage polarization by MSC-derived CXCL12 determines tumor growth.

BACKGROUND: Phenotypic and functional heterogeneity of macrophages is known to be the main reason for their ability to regulate inflammation and promote tumorigenesis. Mesenchymal stem cells (MSCs) are one of the principal cells commonly found in the tumor stromal niche, with capability of macrophage phenotypic switching. The objective of this study was to evaluate the role of C-X-C motif chemokine ligand 12 (CXCL12) produced by marrow-derived MSCs in the phenotypic and functional pattern of bone marrow-derived macrophages (BMDMs). METHODS: First, the CRISPR/Cas9 system was used for the CXCL12 gene knock-out in MSCs. Then, coculture systems were used to investigate the role of MSCsCXCL12-/- and MSCsCXCL12+/+ in determination of macrophage phenotype. To further analyze the role of the MSC-derived CXCL12 niche, cocultures of 4T1 mammary tumor cells and macrophages primed with MSCsCXCL12-/- or MSCsCXCL12+/+ as well as in-vivo limiting dilution assays were performed. RESULTS: Our results revealed that the expression of IL-4, IL-10, TGF-ß and CD206 as M2 markers was significantly increased in macrophages co-cultured with MSCsCXCL12+/+ , whereas the expression of IL-6, TNF-α and iNOS was conversely decreased. The number and size of multicellular tumor spheroids were remarkably higher when 4T1 cells were cocultured with MSCCXCL12+/+-induced M2 macrophages. We also found that the occurrence of tumors was significantly higher in coinjection of 4T1 cells with MSCCXCL12+/+-primed macrophages. Tumor initiating cells were significantly decreased after coinjection of 4T1 cells with macrophages pretreated with MSCsCXCL12-/-. CONCLUSIONS: In conclusion, our findings shed new light on the role of MSC-derived CXCL12 in macrophage phenotypic switching to M2, affecting their function in tumorigenesis.

Impairment of endothelial progenitor cells function in patient with mustard gas intoxication.

Background: Sulfur mustard (SM), also known as mustard gas, was first widely used in the Iraq-Iran. After SM exposure, the most prominent clinical signs are the development of extensive non-healing skin wounds and pulmonary signs, persisting over long time. Since the most frequent complications in SM-intoxicated patients are respiratory and dermatologic lesions, and with respect to the important role of endothelial progenitor cells (EPCs) in the pathophysiology of these lesion, we conducted this study to recognize the potential effects of SM on biological features of EPCs in patients exposed with this gas.Methods: In this study, 30 patients with the history of SM exposure during the Iran-Iraq war (1984-1988), 27 patients with pulmonary signs with no history of SM exposure and 20 healthy participants were included. Cell population and function of EPCs were assessed 4 years post-exposure. For this purpose, circulating EPCs (cEPCs) were harvested and cultivated, then the biological features of these cells, including migratory, proliferative, and tubulogenic activities were analyzed. We also measured serum antioxidants levels and mRNA levels of some proangiogenic factors in EPCs from SM-intoxicated patients.Results: Our results showed lesser number of cEPCs in patients exposed with SM, which was associated with decreased proliferative, migratory, and tubulogenic activity of these cells. Also, we found the lesser serum activity of SOD, GPX and MDA in the SM group than in the healthy control group.Conclusions: SM exposure resulted in decreased proliferation and migration of EPCs, which was associated with decreased tubule formation and angiogenic factors.

Correction to: Low-level laser irradiation at a high power intensity increased human endothelial cell exosome secretion via Wnt signaling.

After publication of this paper, the authors determined an error in Fig. 4. Below is the correct Fig. 4.

Evaluating the effect of remote ischemic preconditioning on kidney ischemia-reperfusion injury.

BACKGROUND: Acute kidney injury is a high-risk complication in a variety of clinical situations mostly due to ischemia-reperfusion (IR) injuries. The novel idea of remote ischemic preconditioning (rIPC) was proposed to prevent serious ischemia sequels. To address the controversy of previous reports, the current study was performed to assess the effect of rIPC on kidney IR injury. MATERIALS AND METHODS: Male BALB/c mice were exposed to either rIPC or sham intervention, 24 h before kidney IR. In two independent sets of experiments, rIPC was accomplished by inducing three cycles of 5 min ischemia with 5 min reperfusion intervals through the ligation of the left external iliac artery or infrarenal abdominal aorta. Kidney IR injury was performed by left renal pedicle occlusion for 35 min and simultaneous right nephrectomy. After 48 h, mice were sacrificed for the assessment of kidney function and structure. RESULTS: According to the serum urea and creatinine, as well as histopathological measures, none of the exploited rIPC procedures could significantly protect against kidney IR injury. CONCLUSION: Based on our findings and the divergent results of previous animal and human studies, it can be concluded that the renoprotective effects of rIPC are minimal, if any, and are not robustly detectable.

Distinct Tie2 tyrosine phosphorylation sites dictate phenotypic switching in endothelial progenitor cells.

Angiogenesis is a regulated process involving the proliferation, migration, and remodeling of different cell types particularly mature endothelial and their progenitor cells, nominated as endothelial progenitor cells (EPCs). Tie2/Tek is a tyrosine kinase receptor expressed by endothelial cells that induces signal transduction pathways involved in endothelial biology. To address the potential importance of the various tyrosine residues of Tie2 in EPC development, we generated a series of Tie2 tyrosine mutated (Y1106F, Y1100F, and Y1111F) EPCs and then assess the biological features of these cells. Clonogenic, tubulogenic, proliferative, migratory, and functional properties of these cells were analyzed. Next, GFP-positive EPCs containing Tie2 tyrosine mutations were systemically transplanted into sublethaly irradiated mice to analyze the potency of these cells for marrow reconstitution. We found that mutation in the Tie2 tyrosine 1106 residue directed EPCs toward a mature endothelial phenotype, which was associated with augmented tubulogenic and migratory properties, and increased phosphorylation of the active site (tyrosine 992) as well as increased vascular perfusion in the in vivo Matrigel plug assay. Moreover, transplantation of 1106 Tie2 mutant EPCs failed to reconstitute the bone marrow after myeloablation, whereas transplantation of EPCs with the 1100 or 1111 Tie2 tyrosine mutation resulted in bone marrow engraftment, leading to improved survival of recipient mice. Our findings demonstrate that the tyrosine 1106 residue in Tie2 plays a key role to maintain the stemness features of EPCs.

Improved angiogenic activity of endothelial progenitor cell in diabetic patients treated with insulin plus metformin.

Type 2 diabetes (T2DM) is associated with an increased vascular disease. Moreover, endothelial progenitor cell (EPC) function is impaired in diabetic patients. Decreased EPC number plays a critical role in reduced endothelial repair and development of the vascular disorder. To determine the effect of metformin and insulin plus metformin on functional activity of EPCs, 130 participants were divided into three groups (group 1: healthy control; group 2: metformin; group 3: insulin plus metformin). The concentration of EPCs in the circulation was first quantified. Thereafter, circulating EPCs (cEPCs) were harvested and the biological features of these cells including proliferative, clonogenicity, tubulogenic, and migratory properties were analyzed after expansion. The serum protein levels of some proangiogenic factors were also measured. Our results showed greater numbers of cEPCs in control and in diabetic patients treated with insulin plus metformin than in metformin-treated patients. Insulin plus metformin therapy was associated with augmented proliferative, clonogenicity, migratory, and tubulogenic activity of cEPCs in patients with T2DM. Increased serum concentrations of angiogenic factors were also observed in patients treated with insulin plus metformin. Western blot analysis showed increased protein levels of pTie-2/Tie2 and Pakt/AKT in cEPCs harvested from T2DM, treated with insulin metformin plus. This study showed that treatment with insulin plus metformin in diabetic patients is associated with increased mobilization of EPCs into the circulation, with potential beneficial effect in vascular protection in diabetic patients.

Comparative measurement of FeLV load in hemolymphatic tissues of cats with hematologic cytopenias.

BACKGROUND: Feline leukemia virus (FeLV) is a serious viral infection in cats. FeLV is found in some tissues, such as spleen, lymph nodes and epithelial tissues. However, there is controversy about the organ in which the virus can be reliably detected in infected cats. The purpose of this study was to determine the level of viral infection in hemolymphatic tissues, including blood, bone marrow and spleen by reverse-transcriptase quantitative polymerase chain reaction (RT-qPCR). RESULTS: A total of 31 cats with clinical signs of FeLV infection associated with at least a single lineage hematologic cytopenia were included in this study. Peripheral blood, bone marrow and spleen samples were obtained from each cat. Complete blood counts, biochemical tests, and a rapid test to detect FeLV p27 antigen in blood samples of cats were performed. Of 31 cats, 9 had anemia alone, 4 had thrombocytopenia alone, 2 had neutropenia alone, 9 had bicytopenia of anemia and thrombocytopenia, 3 had bicytopenia of anemia and neutropenia, and 4 had pancytopenia. FeLV RNA was then detected by RT-qPCR in the whole blood, bone marrow and spleen. Viral RNA copy numbers were detected in all cats by RT-qPCR whereas 24 out of 31 cats were positive for the serum FeLV antigen. We detected a significantly greater number of viral RNA in the spleen compared with the whole blood and bone marrow. CONCLUSION: Spleen is a site where FeLV is most frequently detected in cats with hematologic cytopenias.

Increased circulation mobilization of endothelial progenitor cells in preterm infants with retinopathy of prematurity.

Retinopathy of prematurity (ROP) is a result of increased pathological neoangiogenesis of the retina in preterm infants. Cells responsible for the pathogenesis of ROP are unclear, but some evidence indicates that bone marrow derived cells are involved in this disorder. Endothelial progenitor cells (EPCs), play a role in angiogenesis in response to tissue ischemia or endothelial damage. In this study, the number of cEPCs in preterm infants with ROP was determined to identify whether the circulation mobilization of EPCs is associated with ROP. We evaluated 99 participants in this study: 22 preterm infants with ROP, 35 preterm infants without ROP, and 42 full-term infants. The release of EPCs in the circulation was first quantified. Thereafter, cEPCs were harvested and cultivated, then the biological features of these cells including migratory, proliferative, and tubulogenic activities were analyzed. The mRNA levels of some proangiogenic factors were also measured in preterm infants. Our results showed greater numbers of cEPCs in infants with ROP, which was associated with increased serum concentrations of angiogenic factors and with augmented proliferative, migratory, and tubulogenic activity of these cells. Western blotting showed increased protein levels of VEGF and HIF-α in cEPCs harvested from ROP infants. This study showed that ROP in preterm infants is associated with increased mobilization of EPCs into the circulation. Therefore, increased cEPCs along with elevated levels of angiogenic factors and tubulogenesis suggest that these cells may play a role in the development and progression of ROP.

Hyperbilirubinemia-induced pro-angiogenic activity of infantile endothelial progenitor cells.

OBJECTIVES: Bilirubin, a by-product of heme degradation, is suggested to have a role for vascular protection. There is increasing evidence that bilirubin may directly affect the function and secretory activity of endothelial cells. In this study, potential effect of hyperbilirubinemia on biological features of circulation endothelial progenitor cells (cEPCs) isolated from infants was investigated. METHODS: Circulation concentration, differentiation and migratory activity of cEPCs isolated from infants with (n = 111) or without (n = 73) hyperbilirubinemia were analyzed. Then, the potential beneficial effect of conditioned medium of cEPCs from infants with or without hyperbilirubinemia was examined on experimental mouse wounds. RESULTS: Our results revealed significantly higher percentages of cEPCs in infants with hyperbilirubinemia. Cell proliferation, and migratory properties of cEPCs isolated and expanded from infants with hyperbilirubinemia were significantly improved. Also, the conditioned medium of cEPCs from hyperbilirubinemic infants possessed a superior beneficial effect on wound healing, which was associated with increased protein levels of VEGF, IL-10, and Pho-ERK/ERK, and decreased TNF-α in the wound tissues. CONCLUSIONS: Our results showed that hyperbilirubinemia can activate migration, proliferating and angiogenic properties of cEPCs. Hyperbilirubinemia can promote the proangiogenic secretory activity of cEPCs, thereby resulting in enhancement of their regenerative wound healing properties.

Circulating endothelial progenitor cells in pregnant women with premature rupture of membranes: potential association with placental disorders.

The frequency of preterm labour has risen over the last few years. Plasma oestrogen concentrations differ between patients who deliver before term and those who deliver at term. Oestrogen can influence the kinetics of circulating endothelial progenitor cells (cEPCs). Here, we attempted to identify the potential association of cEPCs with the incidence of complications typical of prematurity. The study groups consisted of 60 pregnant women with premature rupture of membranes (PROM; less than 37 weeks) and 50 term pregnant women (more than 38 weeks). cEPCs were isolated from term pregnant women and pregnant women with PROM and then migratory, proliferative, tubulogenic and functional properties of these cells along with serum secretion of important EPC chemotactic cytokines were analysed. In addition, the effect of 17ß-oestradiol on biological features of cEPCs harvested from pregnant women was investigated. Our results showed that an increased concentration of oestrogen in women with PROM was associated with increased numbers of cEPCs, with these cells having increased oestrogen receptor α expression together with augmented proliferative, migratory and colony-formation properties. 17ß-oestradiol induced proliferation, migration and angiogenic secretory activity of cEPCs from pregnant women. Overall, circulation mobilisation of EPCs in pregnant women may be associated with placental disorders.

Low-level laser irradiation at a high power intensity increased human endothelial cell exosome secretion via Wnt signaling.

The distinct role of low-level laser irradiation (LLLI) on endothelial exosome biogenesis remains unclear. We hypothesize that laser irradiation of high dose in human endothelial cells (ECs) contributes to the modulation of exosome biogenesis via Wnt signaling pathway. When human ECs were treated with LLLI at a power density of 80 J/cm2, the survival rate reduced. The potential of irradiated cells to release exosomes was increased significantly by expressing genes CD63, Alix, Rab27a, and b. This occurrence coincided with an enhanced acetylcholine esterase activity, pseudopodia formation, and reduced zeta potential value 24 h post-irradiation. Western blotting showed the induction of LC3 and reduced level of P62, confirming autophagy response. Flow cytometry and electron microscopy analyses revealed the health status of the mitochondrial function indicated by normal ΔΨ activity without any changes in the transcription level of PINK1 and Optineurin. When cells exposed to high power laser irradiation, p-Akt/Akt ratio and in vitro tubulogenesis capacity were blunted. PCR array and bioinformatics analyses showed the induction of transcription factors promoting Wnt signaling pathways and GTPase activity. Thus, LLLI at high power intensity increased exosome biogenesis by the induction of autophagy and Wnt signaling. LLLI at high power intensity increases exosome biogenesis by engaging the transcription factors related to Wnt signaling and autophagy stimulate.

ß3-Adrenergic Regulation of EPC Features Through Manipulation of the Bone Marrow MSC Niche.

Mesenchymal stem cells (MSCs) reside in a specific niche in the bone marrow, however, biological features of this niche are still not fully understood. Given the interactions of MSCs with endothelial cells in different tissues, bone marrow MSC niche may influence the biological features of endothelial progenitor cells (EPCs). To understand the role of the sympathetic nervous system in regulation of the MSC niche, we examined whether the manipulation of the MSC niche via ß3-adrenergic signals will affect EPC features. A selective ß3 agonist (BRL37344) or a ß3 antagonist (SR59230A) was administered in mice for 2 weeks to determine the potential effects of these regimens on the population of CD133+ stem cells in the bone marrow. Then, bone marrow-derived MSCs and EPCs were harvested and expanded from the mice to examine the effect of changes in the MSC niche on EPC features. Improved MSC colony forming potency with increased bone marrow stromal cell-derived factor 1 (SDF-1) (also known as C-X-C motif chemokine 12 [CXCL12]) expression was shown as a result of intensification of the bone marrow adrenergic signals through BRL37344 injection. On the other hand, the blockage of these signals limited the expression level of SDF-1 and resulted in bone marrow enrichment of CD133+ cells. Manipulation of the MSC niche and decreased SDF-1 expression via SR59230A injection also prompted EPCs to form more colonies with augmented proliferation and differentiation capacity. Overall, our results indicate that the ß3-adrenergic signals regulate the MSC niche, thereby resulting in modulation of EPC biological features. J. Cell. Biochem. 118: 4753-4761, 2017. © 2017 Wiley Periodicals, Inc.

Circulation Enrichment of Functional Endothelial Progenitor Cells by Infantile Phototherapy.

Phototherapy is the most common therapy used for severe jaundice. There is increasing evidence that phototherapy can directly affect the expression and function of cell surface receptors including adhesion molecules, cytokines, and growth factor receptors. In this study, the effect of two infantile phototherapy regimens, including single and intensive phototherapy was investigated on biological features of circulation endothelial progenitor cells (cEPCs), as well as on serum secretion of two important chemotactic cytokines, SDF-1 and VEGF. Sixty infants diagnosed with severe hyperbilirubinemia and exposed to phototherapy were enrolled in this study. cEPCs were isolated before and after phototherapy and then migratory, proliferative, tubulogenic, and functional properties of these cells were analyzed. Our results revealed that intensive phototherapy markedly increased the release of EPCs into the circulation, and augmented the serum concentrations of both SDF-1 and VEGF cytokines. Cell proliferation, tubulogenic, and migratory properties of cEPCs isolated and expanded from infants with intensive phototherapy were significantly improved. cEPCs from infants with intensive phototherapy also showed greater levels of acetylated low-density lipoprotein and lectin binding. Overall, our results showed that the intensive phototherapy regimen can mobilize functional EPCs into the circulation through up-regulation of serum levels of VEGF and SDF-1, indicating phototherapy as an effective modality for improvement of stem cell mobilization in the therapeutic regenerative medicine. J. Cell. Biochem. 118: 330-340, 2017. © 2016 Wiley Periodicals, Inc.

The Improvement of Respiratory Performance After Phototherapy-Induced EPC Mobilization in Preterm Infants With RDS.

Many infants who develop bronchopulmonary dysplasia (BPD) are born with serious respiratory distress syndrome (RDS), which is associated with impaired vascular and alveolar growth. RDS is a breathing disorder that mostly affects preterm infants and occurs in infants whose lungs have not yet been fully developed. The use of surfactant in RDS treatment does not necessarily prevent BPD. Endothelial progenitor cells (EPCs) may contribute to lung angiogenesis for the prevention and treatment of BPD. The aim of this study was to evaluate the therapeutic efficacy of phototherapy for EPC release in preterm infants born with RDS. Seventy-five RDS preterm infants were divided into two groups: RDS with phototherapy and RDS without phototherapy. Respiratory indices were recorded for each patient. Circulating EPCs were isolated before and after phototherapy and colony forming efficiency, chemotactic, tubulogenic, proliferative, and functional properties of these cells were analyzed. Our results showed that phototherapy increased the release of EPCs into the circulation in RDS preterm infants, with augmentation of cell proliferation, tubulogenic, chemotactic, and proliferative properties of EPCs. Phototherapy-induced EPC release was associated with improved lung function as was recorded by significantly decrease in continuous positive airway pressure (CPAP) days, CPAP plus ventilator days, and PCO2 along with a significant increase in PO2 and PaO2 /FiO2 , resulting in markedly decreased rate of BPD occurrence in preterm infants with RDS. Overall, phototherapy is touted as a promising modality for the amelioration of respiratory performance and prohibition of BPD occurrence in RDS preterm infants. J. Cell. Biochem. 118: 594-604, 2017. © 2016 Wiley Periodicals, Inc.

Endothelial Progenitor Cell Mobilization in Preterm Infants With Sepsis Is Associated With Improved Survival.

Microvascular dysfunction plays a key role in the pathology of sepsis, leading to multi-organ failure, and death. Circulating endothelial progenitor cells (cEPCs) are critically involved in the maintenance of the vascular homeostasis in both physiological and pathological contexts. In this study, concentration of cEPCs in preterm infants with sepsis was determined to recognize whether the EPC mobilization would affect the clinical outcome of infantile sepsis. One hundred and thirty-three preterm infants (81 with sepsis and 52 without sepsis) were enrolled in this study. The release of EPCs in circulation was first quantified. Thereafter, these cells were cultivated and biological features of these cells such as, proliferation and colony forming efficiency were analyzed. The levels of chemoattractant cytokines were also measured in infants. In mouse models of sepsis, effects of VEGF and SDF-1 as well as anti-VEGF and anti-SDF-1 were evaluated in order to shed light upon the role which the EPC mobilization plays in the overall survival of septic animals. Circulating EPCs were significantly higher in preterm infants with sepsis than in the non-sepsis group. Serum levels of VEGF, SDF-1, and Angiopoietin-2 were also higher in preterm infants with sepsis than in control non-sepsis. In the animal experiments, injection of VEGF and SDF-1 prompted the mobilization of EPCs, leading to an improvement in survival whereas injection of anti-VEGF and anti-SDF-1 was associated with significant deterioration of survival. Overall, our results demonstrated the beneficial effects of EPC release in preterm infants with sepsis, with increased mobilization of these cells was associated with improved survival. J. Cell. Biochem. 118: 3299-3307, 2017. © 2017 Wiley Periodicals, Inc.

ECM-Dependence of Endothelial Progenitor Cell Features.

Preserving self-renewal, multipotent capacity, and large-scale expansion of highly functional progenitor cells, including endothelial progenitor cells (EPCs), is a controversial issue. These current limitations, therefore, raise the need of developing promising in vitro conditions for prolonged expansion of EPCs without loss of their stemness feature. In the current study, the possible role of three different natural extracellular substrates, including collagen, gelatin, and fibronectin, on multiple parameters of EPCs such as cell morphology, phenotype, clonogenic, and vasculogenic properties was scrutinized. Next, EPCs from GFP-positive mice were pre-expanded on each of these ECM substrates and then systemically transplanted into sublethaly irradiated mice to analyze the potency of these cells for marrow reconstitution. Our results revealed considerable promise for fibronectin for EPC expansion with maintenance of stemness characteristics, whereas gelatin and collagen matrices directed the cells toward a mature endothelial phenotype. Transplantation of EPCs pre-expanded on fibronectin resulted in widespread distribution and appropriate engraftment to various tissues with habitation in close association with the microvasculature. In addition, fibronectin pre-expanded cells were gradually enriched in the bone marrow after transplantation, resulting in marrow repopulation and hematologic recovery, leading to improved survival of recipient mice whereas gelatin- and collagen-expanded cells failed to reconstitute the bone marrow. This study demonstrated that, cell characteristics of in vitro expanded EPCs are determined by the subjacent matrix. Fibronectin-expanded EPCs are heralded as a source of great promise for bone marrow reconstitution and neo-angiogenesis in therapeutic bone marrow transplantation. J. Cell. Biochem. 117: 1934-1946, 2016. © 2016 Wiley Periodicals, Inc.

Detection of Critical Genes Associated with Overall Survival (OS) and Progression-Free Survival (PFS) in Reconstructed Canine B-Cell Lymphoma Gene Regulatory Network (GRN).

Canine B-cell lymphoma GRN was reconstructed from gene expression data in the STRING and MiMI databases. Critical genes of networks were identified and correlations of critical genes with overall survival (OS) and progression-free survival (PFS) were evaluated. Significant changes were detected in the expressions of GLUL, CD44, CD79A, ARF3, FOS, BLOC1S1, FYN, GZMB, GALNT3, IFI44, CD3G, GNG2, ESRP1, and CCND1 in the STRING network and of PECAM1, GLUL, CD44, GDI1, E2F4, TLE1, CD79A, UCP2, CCND1, FYN, RHOQ, BIN1, and A2M in the MiMI network. Final survival analysis highlighted CCND1 and FOS as genes with significant correlations with OS and PFS.

Endothelial juxtaposition of distinct adult stem cells activates angiogenesis signaling molecules in endothelial cells.

Efficacy of therapeutic angiogenesis needs a comprehensive understanding of endothelial cell (EC) function and biological factors and cells that interplay with ECs. Stem cells are considered the key components of pro- and anti-angiogenic milieu in a wide variety of physiopathological states, and interactions of EC-stem cells have been the subject of controversy in recent years. In this study, the potential effects of three tissue-specific adult stem cells, namely rat marrow-derived mesenchymal stem cells (rBMSCs), rat adipose-derived stem cells (rADSCs) and rat muscle-derived satellite cells (rSCs), on the endothelial activation of key angiogenic signaling molecules, including VEGF, Ang-2, VEGFR-2, Tie-2, and Tie2-pho, were investigated. Human umbilical vein endothelial cells (HUVECs) and rat lung microvascular endothelial cells (RLMECs) were cocultured with the stem cells or incubated with the stem cell-derived conditioned media on Matrigel. Following HUVEC-stem cell coculture, CD31-positive ECs were flow sorted and subjected to western blotting to analyze potential changes in the expression of the pro-angiogenic signaling molecules. Elongation and co-alignment of the stem cells were seen along the EC tubes in the EC-stem cell cocultures on Matrigel, with cell-to-cell dye communication in the EC-rBMSC cocultures. Moreover, rBMSCs and rADSCs significantly improved endothelial tubulogenesis in both juxtacrine and paracrine manners. These two latter stem cells dynamically up-regulated VEGF, Ang-2, VREGR-2, and Tie-2 but down-regulated Tie2-pho and the Tie2-pho/Tie-2 ratio in HUVECs. Induction of pro-angiogenic signaling in ECs by marrow- and adipose-derived MSCs further indicates the significance of stem cell milieu in angiogenesis dynamics.