Mesenchymal Growth Factors Research Articles

Longitudinal positron emission tomography imaging for monitoring myelin repair in the spinal cord

Novel therapeutic interventions aimed at myelin repair are now under development for neuroprotection as well as functional recovery of patients with multiple sclerosis. However, development of myelin repair therapy necessitates a noninvasive approach for measuring changes in myelin content in vivo in a quantitative fashion not yet possible using magnetic resonance imaging. For this reason, we developed a novel positron emission tomography (PET) probe, termed [11C]MeDAS, that is capable of longitudinally imaging central nervous system myelin content. The binding properties of [11C]MeDAS for myelin were systematically evaluated by in vitro and in situ fluorescent staining of the spinal cord and the brain, and by in vivo competitive blocking studies. Longitudinal PET studies were conducted in 3 rat models involving acute focal neuroinflammation in the brain, lysophosphatidylcholine (LPC)-induced focal demyelination in the spinal cord, and experimental autoimmune encephalomyelitis (EAE). Image-guided myelin repair therapy was conducted in an LPC rat model using a mesenchymal stem cell-based hepatocyte growth factor (HGF). Biodistribution and acute toxicity studies of [11C]MeDAS were also conducted. MeDAS selectively stains myelin in the spinal cord and brain. Neuroinflammation did not affect [11C]MeDAS uptake in the brain as long as the myelin sheaths remained intact. Longitudinal PET studies in LPC and EAE rat models demonstrate that [11C]MeDAS uptake changes correlate with associated myelin loss in the spinal cord. Furthermore, using [11C]MeDAS-PET, the efficacy of myelin repair therapy with HGF was longitudinally monitored in vivo. [11C]MeDAS-PET is a promising imaging marker for monitoring myelin pathology in vivo, future applications of which in humans should be achievable.

Bilaterally cleft lip and bilateral thumb polydactyly with triphalangeal component in a patient with two De novo deletions of HSA 4q32 and 4q34 involving PDGFC, GRIA2, and FBXO8 genes

We report on a newborn boy with a bilateral cleft of the primary palate, duplicated triphalangeal thumbs, and a patent foramen ovale. During childhood he had moderate developmental delay. Brain MRI at 4 years was normal. The concurrence of non-syndromic clefts of the lip/palate (CL/P) and duplicated thumbs with triphalangeal component has, to our knowledge, not been reported so far. In our case, array-CGH analysis documented two de novo deletions (∼1.2 Mb and ∼400 Kb) of the long arm of chromosome 4, containing four genes: platelet-derived growth factor C (PDGFC), glycine receptor beta subunit (GLRB), glutamate receptor ionotropic AMPA2 (GRIA2), and F-box protein 8 gene (FBXO8). PDGFC codes for a mesenchymal cell growth factor already known to be associated with clefts of the lip. Pdgfc(-/-) mice have skeletal anomalies, and facial schisis resembling human cleft/lip palate. GRIA2 codes for a ligand-activated cation channel that mediates the fast component of postsynaptic excitatory currents in neurons, and may be linked to cognitive dysfunction. FBXO8, a gene of unknown function, is a member of the F-box gene family, among which FBXW4, within the minimal duplicated region associated with human split-hand/foot malformation type 3 (SHFM type 3). The presence of overlapping deletions in patients who do not share the same phenotype of our case suggests incomplete penetrance, and a possible effect of modifier genetic factors.

Tissue-engineered blood vessel substitute by reconstruction of endothelium using mesenchymal stem cells induced by platelet growth factors

Cardiovascular diseases remain leaders as the major causes of mortality in Western society. Restoration of the circulation through construction of bypass surgical treatment is regarded as the gold standard treatment of peripheral vascular diseases, and grafts are necessary for this purpose. The great saphenous vein is often not available and synthetic grafts have their limitations. Therefore, new techniques to produce alternative grafts have been developed and, in this sense, tissue engineering is a promising alternative to provide biocompatible grafts. This study objective was to reconstruct the endothelium layer of decellularized vein scaffolds, using mesenchymal stem cells (MSCs) and growth factors obtained from platelets. Fifteen nonpregnant female adult rabbits were used for all experiments. Adipose tissue and vena cava were obtained and subjected to MSCs isolation and tissue decellularization, respectively. MSCs were subjected to differentiation using endothelial inductor growth factor (EIGF) obtained from human platelet lysates. Immunofluorescence, histological and immunohistochemical analyses were employed for the final characterization of the obtained blood vessel substitute. The scaffolds were successfully decellularized with sodium dodecyl sulfate. MSCs actively adhered at the scaffolds, and through stimulation with EIGF were differentiated into functional endothelial cells, secreting significantly higher quantities of von Willebrand factor (0.85 μg/mL; P < .05) than cells cultivated under the same conditions, without EIGF (0.085 μg/mL). Cells with evident morphologic characteristics of endothelium were seen at the lumen of the scaffolds. These cells also stained positive for fascin protein, which is highly expressed by differentiated endothelial cells. Taken together, the use of decellularized bioscaffold and subcutaneous MSCs seems to be a potential approach to obtain bioengineered blood vessels, in the presence of EIGF supplementation.

In-vitro Behavior of Human Umbilical Cord Blood Stem Cells Towards Serum Based Minimal Cytokine Growth Conditions.

We tried here to optimize the proliferation of both Hematopoietic and Mesenchymal stem cells of Umbilical Cord blood in minimal cytokine growth condition. Failing to get good results of expansion of non-adherent Hematopoietic Total Nucleated Cells and adherent Fibroblastic Mesenchymal Stem Cells derived from 10-12ml of collected Cord blood, we designed the further experimental study by increasing the volume of Cord blood sample up to 65-70ml. We harvested the non-adherent as well as adherent fraction separately derived from the primary culture of Umbilical Cord blood stem cells under the influence of growth promoting Cytokines or Growth Factors. The proliferation study was conducted by taking different combinations of two hematopoietic growth stimulatory Cytokines like stem cell factor (SCF) and Fms like tyrosine kinase-3Ligand (Flt3L) at concentrations (10ng/ml, 100ng/ml) while we preferred Mesenchymal specific growth factor i.e. basic Fibroblast growth factor (FGF-β) at its 10ng/ml concentration for adherent cells to get optimal results. The Hematopoietic and Fibroblast Colony forming abilities of the expanded stem cells were performed through Colony Forming Unit assay. Culture Medium containing cytokine combination like SCF 100ng/ml with Flt3L 10ng/ml was found to be optimal for the proliferation of hematopoietic stem cells. But the number of hematopoietic colonies like Erythroid colonies generated were less in case of media supplemented with SCF & Flt3L while more number of Myeloid colonies were observed in Growth factor supplemented media in comparison to the control one. The FGF-β supplemented media successfully enhanced the proliferation of Mesenchymal Stem Cells and exhibited its efficient Fibroblast colony forming ability. Our experimental study supports the minimal utilization of cytokines for haematopoietic and mesenchymal stem cell proliferation which may help in future safe Cord blood stem cell infusion.

Controlled delivery of mesenchymal stem cells and growth factors using a nanofiber scaffold for tendon repair

Outcomes after tendon repair are often unsatisfactory, despite improvements in surgical techniques and rehabilitation methods. Recent studies aimed at enhancing repair have targeted the paucicellular nature of tendon for enhancing repair; however, most approaches for delivering growth factors and cells have not been designed for dense connective tissues such as tendon. Therefore, we developed a scaffold capable of delivering growth factors and cells in a surgically manageable form for tendon repair. Platelet-derived growth factor BB (PDGF-BB), along with adipose-derived mesenchymal stem cells (ASCs), were incorporated into a heparin/fibrin-based delivery system (HBDS). This hydrogel was then layered with an electrospun nanofiber poly(lactic-co-glycolic acid) (PLGA) backbone. The HBDS allowed for the concurrent delivery of PDGF-BB and ASCs in a controlled manner, while the PLGA backbone provided structural integrity for surgical handling and tendon implantation. In vitro studies verified that the cells remained viable, and that sustained growth factor release was achieved. In vivo studies in a large animal tendon model verified that the approach was clinically relevant, and that the cells remained viable in the tendon repair environment. Only a mild immunoresponse was seen at dissection, histologically, and at the mRNA level; fluorescently labeled ASCs and the scaffold were found at the repair site 9days post-operatively; and increased total DNA was observed in ASC-treated tendons. The novel layered scaffold has the potential for improving tendon healing due to its ability to deliver both cells and growth factors simultaneously in a surgically convenient manner.

Crizotinib: A New Treatment Option for ALK-Positive Non-Small Cell Lung Cancer

To review the characteristics and clinical trial data of crizotinib in ALK-positive non-small cell lung cancer (NSCLC). A literature search using PubMed/MEDLINE (up to December 2012) was performed using the terms crizotinib, ALK-positive, non-small cell lung cancer, and PF-02341066. Phase 1, 2, and 3 trials evaluating the safety and efficacy of crizotinib in a cohort of patients with ALK rearrangements and advanced NSCLC were evaluated. All peer-reviewed articles with clinically relevant information were reviewed. ALK rearrangement results in an aberrant EML4-ALK fusion oncogene that constitutively activates ALK tyrosine kinase, resulting in inhibition of apoptosis and promotion of tumor cell proliferation. Approximately 3-5% of NSCLC exhibit this rearrangement. Crizotinib is an oral selective inhibitor of ALK and mesenchymal epithelial growth factor tyrosine kinases. Early phase trials with crizotinib showed improved response rates of 50-57% and extended duration of response of 6-10 months. Results of these studies led to accelerated Food and Drug Administration (FDA) approval of crizotinib. Further clinical trial results confirmed improvement in response rates, duration of response, as well as progression-free survival in ALK-positive patients with NSCLC receiving crizotinib. The drug undergoes hepatic metabolism by CYP3A4 and demonstrates autoinhibition of CY3A4, thus predisposing it to drug interactions. The most frequent toxicities with crizotinib include mild visual disturbances, nausea, vomiting, diarrhea, constipation, edema, and generally reversible, sometimes severe, elevations in aspartate aminotransferase and alanine aminotransferase. Crizotinib is a novel targeted anticancer agent that appears to be a favorable treatment option for patients with locally advanced or metastatic NSCLC that is ALK-positive as detected by an FDA-approved test.

MRNA expression and promoter DNA methylation status of CDKi p21 and p57 genes in ex vivo expanded CD34+ cells following co-culture with mesenchymal stromal cells and growth factors

BackgroundSeveral studies indicate that ex vivo cytokine-based expansion of cord blood (CB) CD34+ cells can induce cell cycle abnormality in expanded cells. Cycline-dependent kinase inhibitors, p21 and p57, are critical regulators of cell cycle. We investigated mRNA expression and promoter DNA methylation status of p21 and p57 genes in ex vivo expanded CD34+ cells in different culture conditions.Materials and methodsHuman CB CD34+ cells were cultured either in cytokine-supplemented liquid culture or in co-culture with mesenchymal stromal cells (co-culture system) in presence and absence of cytokine. After 14 days, expansion fold in total nucleated cells (TNCs), CD34+ cells, CD34+/CD38− cells and colony-forming units in culture (CFU-C) count was determined. Moreover, mRNA expression level and promoter DNA methylation status of p21 and p57 genes in expanded CD34+ cells were analyzed by real-time reverse transcriptase-polymerase chain reaction (PCR) and methylation-specific PCR technique, respectively.ResultsMaximum expansion of TNC, CD34+ cells, CD34+/CD38− cells and CFU-C was seen at day 10 in all three culture conditions. p21 and p57 mRNA expression level decreased in both cytokine liquid culture and co-culture system with cytokine, while in the co-culture system without cytokine, the mRNA expression of p21 and p57 genes relatively increased. In addition, promoter DNA methylation status of p21 and p57 genes did not change during expansion in all three culture conditions.ConclusionsCo-culture of CB CD34+ cells with mesenchymal stromal cells, especially in the absence of cytokines, maintained mRNA expression of p21 and p57 genes in ex vivo expanded CD34+ cells. Moreover, transcriptional repression of p21 and p57 genes in ex vivo expanded CD34+ cells was not mediated by promoter DNA methylation.

MiR‐16 inhibits the proliferation and angiogenesis‐regulating potential of mesenchymal stem cells in severe pre‐eclampsia

Pre-eclampsia is thought to be a systemic disease of maternal endothelial cell dysfunctions. miRNAs regulate various basic biological functions in cells, including stem cells. Mesenchymal stem cells exist in almost all tissues and are the key cellular source for tissue repair and regeneration. Our aims are to investigate whether miRNAs regulate MSCs in fetal-maternal interfaces to influence the pathogenesis of pre-eclampsia. The differential expression of miRNAs in decidua-derived mesenchymal stem cells of all patients with severe pre-eclampsia (n=20) and normal groups (n=20) was first screened by microarray analysis and validated by quantitative real-time PCR analysis. The integrated bioinformatics analysis showed that miR-16 showed the highest number of connections in the miRNA GO network and the miRNA gene network. Moreover, over-expressed miR-16 inhibited the proliferation and migration of decidua-derived mesenchymal stem cells and induced cell-cycle arrest by targeting cyclinE1. Interestingly, over-expression of miR-16 by decidua-derived mesenchymal stem cells reduced the ability of human umbilical vein endothelial cells to form blood vessels and reduced the migration of trophoblast cells. Furthermore, decidua-derived mesenchymal stem cell-expressed endothelial growth factor VEGF-A was involved in migration of trophoblast cells and human umbilical vein endothelial cells as well as tube and network formation. Importantly, the levels of cyclinE1 and VEGF-A were negatively correlated with the level of miR-16 expression in decidua-derived mesenchymal stem cells from the patients with severe pre-eclampsia. Together, these data suggest that the alteration of miR-16 expression in decidua-derived mesenchymal stem cells may be involved in the development of pre-eclampsia.

The role of adult bone marrow derived mesenchymal stem cells in the repair of tissue injuries

Mesenchymal stem cells, which reside in adult bone marrow are multipotent, have an excellent regeneration potential for tissue repair. These cells are able to differentiate in cell culture not only into mesodermal lineages but also into other lineages of ectodermal and endodermal cells. This regenerative process is assisted by application of bioactive molecules, specific growth factors and biomaterials (scaffolds). The cell therapy is successfully used in the treatment of bone defects, nonunions, osteoblasts formed from the mesenchymal stem cells. At present, there are encouraging data in the clinical practice. The mesenchymal stem cell seems to be successful in the regeneration of articular cartilage. There are further promising data for the application of mesenchymal stem cells in the treatment of myocardial infarction, neurologic diseases, liver and kidney diseases and injuries and diabetes mellitus. The aim of this review is to survey the molecular characteristics of mesenchymal stem cells and specific growth factors using the data of preclinical investigations and to call attention to their possible clinical application.

An Upstream Finite Volume Scheme for a Bone Healing Model

This paper is devoted to the introduction of a numerical scheme for a bone healing model and simulation of skull fractures. The mathematical model describes the evolution of mesenchymal stem cells, osteoblasts, bone matrix and osteogenic growth factor. WeВ propose a numerical scheme based on an implicit finite volume method constructed on an orthogonal mesh. The efficiency and robustness of the scheme are shown in simulating a skull fracture in rats.

Arthroscopic Knee Cartilage Repair With Covered Microfracture and Bone Marrow Concentrate

In recent years several single-stage cartilage repair approaches have been devised to treat focal cartilage lesions. These usually associate microfracture (MFX) and a coverage scaffold. We describe a novel arthroscopic technique that combines MFX, autologous bone marrow concentrate (BMC), and a protective scaffold. Bone marrow aspirate from the iliac crest is centrifuged to obtain BMC. The cartilage defect is debrided, MFX holes are created, and the final defect is measured by use of a bent K-wire. The scaffold is then shaped to match the defect, immersed in BMC, introduced into the joint with a grasper, and fixed in place with a mixture of fibrin glue and BMC. This technique aims to augment the original single-stage procedure with a number of mesenchymal stem cells and growth factors contained in the BMC, to increase the defect filling and the rate of hyaline-like cartilage regeneration. The procedure combining MFX, BMC, and a protective scaffold is inexpensive and reproducible and has already shown the ability to regenerate hyaline-like cartilage. Its use as an alternative to autologous chondrocyte implantation requires further investigation.

Cost-Effectiveness Analysis of Autologous Chondrocyte Implantation

Background: Autologous chondrocyte implantation (ACI) involves the use of a periosteal patch (ACI-P) as a cover for transplanted chondrocytes. Theoretically, this periosteal patch provides mesenchymal stem cells and growth factors that encourage chondrocyte development/differentiation. However, there is a significant rate of graft hypertrophy with the use of periosteum compared with using a type I/III collagen patch (ACI-C). This type I/III collagen patch, although not approved by the United States Food and Drug Administration for ACI, has been used extensively in Europe and in an “off-label” nature in the United States as a cover during ACI. Purpose: To examine the cost effectiveness of ACI and determine whether ACI-C is more cost effective than ACI-P. Study Design: Economic and decision analysis; Level of evidence, 2. Methods: Outcome data and complication rates from patients undergoing ACI (ACI-P and ACI-C) were derived from the best evidence in the literature. Costs were determined by examining the typical patient charges undergoing ACI at a local orthopaedic hospital. The costs, results, and complication rates were used to develop a decision analysis model comparing ACI-P to ACI-C. Results: The cost of ACI-P was $66,752 and for ACI-C was $66,939.50 ($187.50 difference). The cost per quality-adjusted life year (QALY) for ACI-P was $9466 compared with $9243 for ACI-C. Sensitivity analysis was performed regarding the additional cost of the type I/III collagen patch ($780) in ACI-C as well as the rate of graft hypertrophy after ACI-P (25%). This analysis revealed that the cost of the type I/III collagen patch would have to reach $1721, or the rate of graft hypertrophy after ACI-P reduced to almost 11%, before ACI-P became more cost effective than ACI-C. Conclusion: This cost-effectiveness analysis reveals that, while both ACI-P and ACI-C are cost effective, ACI-C is slightly more cost effective than ACI-P. This is likely secondary to the significant rate of patch-related complications associated with ACI-P, which is significantly reduced with ACI-C. Although the model is very sensitive to differences in outcomes between ACI-P and ACI-C, there is no high-quality evidence to suggest that there is a significant difference between the two. Thus, ACI-P becomes more cost effective if the cost of the type I/III collagen membrane is significantly increased or if the rate of graft hypertrophy after ACI-P were to be markedly reduced.

Tissue Engineering in Orthopaedics and Musculoskeletal Sciences

Orthopaedic surgery as a speciality lends itself well to tissue engineering. Many musculoskeletal tissues are frequently damaged or lost in injury and disease, and show a limited capacity for repair. Although orthopaedic surgery has been successful in replacing cartilage, the coming years will establish whether a shift from replacement towards regeneration of tissue is feasible. There has been an increasing interest in tissue engineering approaches to deal with damaged or lost tissue [1-3]. Tissue engineering is an exciting strategy that employs stem cells, scaffolds and growth factors or mediators either in isolation or in combination. It is the expanding science of generating tissue using molecular and cellular techniques, combined with material engineering principles, to replace tissue surgically. This could be in the form of cells with or without matrices. Many tissue engineering advances are being made in orthopaedics and this is partly due to the familiarity with bone marrow derived mesenchymal stem cells. In this review we will cover mesenchymal stem cells, scaffolds, bioreactors and growth factors applicable to orthopaedics and musculoskeletal sciences. This is followed by specific discussions on Dupuytren’s disease where there is early evidence of stem cell etiology, and bone and cartilage tissue engineering.

Polytherapy in Bone Regeneration: Clinical Applications and Preliminary Considerations

Polytherapy, namely the simultaneous application of three fundamental elements necessary for bone regeneration (growth factors, osteogenic cells and osteoconductive scaffolds) seems to lead to a very high success rate in the treatment of complex non-union (NU) cases and critical bone defects. NU are reported in 5-10% of long bone fractures. The use of autologous bone grafts has been long-considered the gold standard for the treatment of these cases. However the harvesting procedure from the iliac crest increases surgery time and presents some donor site complications which may be elevated. In recent years, surgeons have some alternatives to autologous grafting such as: application of organic or synthetic bone substitute, application of mesenchymal stromal cells (MSC) or growth factors (GF). In the literature there are many studies available about their application in monotherapy, but unfortunately the healing rate doesn't exceed 90%. Polytherapy seems to be a logical option to improve the healing rate, nevertheless, there are not still extensive studies that validate this strategy and moreover, some questions are not resolved.

Hepatocyte growth factor activator (HGFA): a serine protease that links tissue injury to activation of hepatocyte growth factor

Growth factors are a group of proteins that regulate a wide variety of cellular processes, including proliferation, differentiation, motility, adhesion, and apoptosis of target cells. They play crucial roles in the formation and maintenance of tissue architecture in embryonic development and adult tissue homeostasis. Because aberrations in growth factor signaling often result in pathological conditions, the activities of growth factors are tightly controlled by extracellular and intracellular regulators. Hepatocyte growth factor (HGF) is a mesenchymal cell-derived growth factor that affects various target cells, including epithelial and endothelial cells. HGF is synthesized and secreted as a latent form, and is proteolytically activated in response to tissue injury, thus participating in tissue regeneration and repair. Interestingly, HGF has a unique structural feature: it is homologous to plasminogen, a key enzyme in the fibrinolytic system. Elucidation of the regulatory mechanisms of HGF activity has revealed that a blood coagulation factor XII-like serine protease, hepatocyte growth factor activator, efficiently converts HGF from the latent form to the active form. Hepatocyte growth factor activator itself is activated downstream of the blood coagulation cascade, and links tissue injury to activation of HGF. HGF thus has structural as well as functional relevance to the blood coagulation/fibrinolytic system.

Sex Specific Expression Patterns of Some Critical Genes During Morhogenesis of Genital Tubercle and the Effect of Exogenous Estrogen

Purpose Several genes are considered to be critical for the normal morphogenesis of the external genital organs and their defects lead to congenital malformations like Hypospadias. We examined the sex-specific ontogenetic expressions of these genes during patternning of fetal mice genital tubercles (GTs) and investigated the effect of in-utero estrogen exposure. Material and Methods Expression of several genes i.e. sonic hedgehog (shh), ephrin B-2 (efn-b2), ephrin B-2 receptor (eph-b2), fibroblast growth factor-8 (fgf-8) and fgf-10 were detected in mouse GTs from fetuses collected on gestational days (GD) 12, 14, 16 and 18, and from newborns using q-PCR analysis. Pregnant dams were exposed to ethinyl estradiol (EE) or corn oil as the control. Results Efn-b2 and eph-b2 expressions increased gradually in both sex during normal development of GT. Fgf-10 mRNA showed an initial increase in both sex until GD-16, later continued to increase in females and to decrease in males. However, both male and female GTs demonstrated a significant declining pattern in terms of shh and fgf-8 expression from GD-12 until birth. In utero EE exposure led to a 50.7% and 77.7% decrease in shh and fgf-8 expressions respectively (p= 0.002 and 0.001, respectively). A similar decrease after EE exposure were observed in eph-b2, but the levels of efn-b2 and fgf-10 (mesenchymal growth factor) were not affected. Conclusions Shh and fgf-8 genes are essential for the early phase of GT outgrowth. Because their expression can be altered by exogenous estrogens, they can be involved in the defected molecular pathways leading to feminizing conditions (i.e. Hypospadias) caused by endocrine disrupters. The previously published data that they are mainly expressed on the urethral plate of the GT supports this result. Finally, efn-b2 and fgf-10 are mainly expressed in later stages of GT development, but they are more resistant to estrogenic effects.

AP-1-Controlled Hepatocyte Growth Factor Activation Promotes Keratinocyte Migration via CEACAM1 and Urokinase Plasminogen Activator/Urokinase Plasminogen Receptor

Keratinocyte migration is essential for the rapid closure of the epidermis in the process of wound healing. Mesenchymal cell-derived hepatocyte growth factor (HGF) is a central regulator of this process. However, the molecular mechanisms and relevant genes that facilitate this cellular response are still poorly defined. We used heterologous cocultures combining primary human keratinocytes and genetically modified murine fibroblasts to identify key factors mediating HGF-induced epidermal cell migration. The absence of c-Jun activity in fibroblasts completely abolished the expression of HGF in these cells and consequently altered the behavior of keratinocytes. Time-resolved expression series of keratinocytes stimulated with HGF disclosed target genes regulating HGF-dependent motility. In addition to well-established HGF-dependent wound healing-associated genes, carcinoembryogenic antigen-related cell adhesion molecule (CEACAM)-1 and the urokinase plasminogen activator (uPA)/uPA-receptor (uPAR) pathway were identified as possible mediators in HGF-induced keratinocyte migration. The functional relevance of CEACAM-1 and uPA/uPAR on epidermal cell motility was demonstrated using the HaCaT cell culture model. In conclusion, the distinct spatiotemporal regulation of genes by HGF is essential for proper epidermal cell migration in cutaneous wound healing.

High glucose concentration in cell culture medium does not acutely affect human mesenchymal stem cell growth factor production or proliferation.

Optimizing the function and proliferative capacity of stem cells is essential to maximize their therapeutic benefits. High glucose concentrations are known to have detrimental effects on many cell types. We hypothesized that human mesenchymal stem cells (hMSCs) cultured in high glucose-containing media would exhibit diminished proliferation and attenuated production of VEGF, hepatocyte growth factor (HGF), and FGF2 in response to treatment with TNF-alpha, LPS, or hypoxia. hMSCs were plated in medium containing low (5.5 mM) and high (20 mM or 30 mM) glucose concentrations and treated with TNF-alpha, LPS, or hypoxia. Supernatants were collected at 24 and 48 h and assayed via ELISA for VEGF, HGF, and FGF2. In addition, hMSCs were cultured on 96-well plates at the above glucose concentrations, and proliferation at 48 h was determined via bromo-2'-deoxy-uridine (BrdU) incorporation. At 24 and 48 h, TNF-alpha, LPS, and hypoxia-treated hMSCs produced significantly higher VEGF, HGF, and FGF2 compared with control. Hypoxia-induced VEGF production by hMSCs was the most pronounced change over baseline. At both 24 and 48 h, glucose concentration did not affect production of VEGF, HGF, or FGF2 by untreated hMSCs and those treated with TNF-alpha, LPS, or hypoxia. Proliferation of hMSCs as determined via BrdU incorporation was unaffected by glucose concentration of the media. Contrary to what has been observed with other cells, hMSCs may be resistant to the short-term effects of high glucose. Ongoing efforts to characterize and optimize ex vivo and in vivo conditions are critical if the therapeutic benefits of MSCs are to be maximized.

The role of adult bone marrow derived mesenchymal stem cells, growth factors and carriers in the treatment of cartilage and bone defects.

Human mesenchymal stem cells, which reside in adult bone marrow, have an excellent regeneration potential for tissue repair. These cells can expand in culture into lineages of mesenchymal tissues and they are able to differentiate into cartilage, bone and other tissues. This regenerative process is assisted by application of bioactive molecules, specific growth factors, signaling molecules and biomaterials (scaffolds). An articular cartilage defect has only limited capacity for healing. Presently no methods have successfully repaired cartilage. The result of preclinical in vitro and in vivo investigations using mesenchymal stem cells, inductive biomolecules or carriers is encouraging for the future clinical applications. Bone defects can be treated with traditional methods using autologous bone grafting, but these procedures are often material limited and can cause donor side morbidity. The formation of bone in small or large defects after injury requires mesenchymal stem cells capable of differentiation in osteoblasts, moreover growth factors that direct these cells into the defect. According to in vitro and in vivo animal investigations and some clinical experiments with the application of cultured mesenchymal cells, growth factors, first of all bone morphogenetic proteins (BMP-s), successful reparative results can be achieved. The application of recombinant human BMP-s is a novel therapy using osteoinductive signaling of a number of gene expression pathways involving differentiation of mesenchymal progenitor cells. The aim of this review is to study the results of preclinical investigations during the last two decades and to deal with the potential clinical application.

Cystic Adenomatoid Malformations Are Induced by Localized FGF10 Overexpression in Fetal Rat Lung

Fibroblast growth factor-10 (FGF10) is a mesenchymal growth factor, involved in epithelial and mesenchymal interactions during lung branching morphogenesis. In the present work, FGF10 overexpression was transiently induced in a temporally and spatially restricted manner, during the pseudoglandular or canalicular stages of rat lung development, by trans-uterine ultrasound-guided intraparenchymal microinjections of adenoviral vector encoding the rfgf10 transgene. The morphologic and histologic classification of the resulting malformations were dependent upon developmental stage and location. Overexpression of FGF10 restricted to the proximal tracheobronchial tree during the pseudoglandular phase resulted in large cysts lined by tall columnar epithelium composed primarily of Clara cells with a paucity of Type II pneumocytes, resembling bronchiolar type epithelium. In contrast, FGF10 overexpression in the distal lung parenchyma during the canalicular phase resulted in small cysts lined by cuboidal epithelial cells composed of primarily Type II pneumocytes resembling acinar epithelial differentiation. The cystic malformations induced by FGF10 overexpression appear to closely recapitulate the morphology and histology of the spectrum of human congenital cystic adenomatoid malformation (CCAM). These findings support a role for FGF10 in the induction of human CCAM and provide further mechanistic insight into the role of FGF10 in normal and abnormal lung development.