Baculovirus transduction of rat articular chondrocytes: roles of cell cycle

BACKGROUND: We have previously demonstrated highly efficient baculovirus transduction of primary rat articular chondrocytes, thus implicating the possible applications of baculovirus in gene-based cartilage tissue engineering. However, baculovirus-mediated gene expression in the chondrocytes is transient. METHODS: In this study, we attempted to prolong the expression by supertransduction, but uncovered that after long-term culture the chondrocytes became more refractory to baculovirus transduction. Therefore, the correlation between baculovirus-mediated enhanced green fluorescent protein (EGFP) expression and cell cycle was investigated by comparing the cycling chondrocytes and chondrocytes rich in quiescent cells, in terms of EGFP expression, virus uptake, cell cycle distribution, nuclear import and methylation of viral DNA. RESULTS: We demonstrated, for the first time, that baculovirus-mediated transduction of chondrocytes is correlated with the cell cycle. The chondrocytes predominantly in G2/M phase were approximately twice as efficient in EGFP expression as the cycling cells, while the cells in S and G1 phases expressed EGFP as efficiently as the cycling cells. Notably, the chondrocyte populations rich in quiescent cells resulted in efficient virus uptake, but less effective nuclear transport of baculoviral DNA and higher degree of methylation, and hence poorer transgene expression. CONCLUSIONS: These findings unravel the practical limitations when employing baculovirus in cartilage tissue engineering. The implications and possible solutions are discussed.     

Expansion on specific substrates regulates the phenotype and differentiation capacity of human articular chondrocytes

In this study, we investigated if monolayer expansion of adult human articular chondrocytes (AHAC) on specific substrates regulates cell phenotype and post-expansion multilineage differentiation ability. AHAC isolated from cartilage biopsies of five donors were expanded on plastic dishes (PL), on dishes coated with collagen type II (COL), or on slides coated with a ceramic material (Osteologic, OS). The phenotype of expanded chondrocytes was assessed by flow cytometry and real-time RT-PCR. Cells were then cultured in previously established conditions promoting differentiation toward the chondrogenic or osteogenic lineage. AHAC differentiation was assessed histologically, biochemically, and by real-time RT-PCR. As compared to PL-expanded AHAC, those expanded on COL did not exhibit major phenotypic changes, whereas OS-expanded cells expressed (i) higher bone sialoprotein (BSP) (22.6-fold) and lower collagen type II (9.3-fold) mRNA levels, and (ii) lower CD26, CD90 and CD140 surface protein levels (1.4-11.1-fold). Following chondrogenic differentiation, COL-expanded AHAC expressed higher mRNA levels of collagen type II (2.3-fold) and formed tissues with higher glycosaminoglycan (GAG) contents (1.7-fold), whereas OS-expanded cells expressed 16.5-fold lower collagen type II and generated pellets with 2.0-fold lower GAG contents. Following osteogenic differentiation, OS-expanded cells expressed higher levels of BSP (3.9-fold) and collagen type I (2.8-fold) mRNA. In summary, AHAC expansion on COL or OS modulated the de-differentiated cell phenotype and improved the cell differentiation capacity respectively toward the chondrogenic or osteogenic lineage. Phenotypic changes induced by AHAC expansion on specific substrates may mimic pathophysiological events occurring at different stages of osteoarthritis and may be relevant for the engineering of osteochondral tissues.     

Tissue engineering strategies for cartilage generation--micromass and three dimensional cultures using human chondrocytes and a continuous cell line

Utilizing ATDC5 murine chondrogenic cells and human articular chondrocytes, this study sought to develop facile, reproducible three-dimensional models of cartilage generation with the application of tissue engineering strategies, involving biodegradable poly(glycolic acid) scaffolds and rotating wall bioreactors, and micromass pellet cultures. Chondrogenic differentiation, assessed by histology, immunohistochemistry, and gene expression analysis, in ATDC5 and articular chondrocyte pellets was evident by the presence of distinct chondrocytes, expressing Sox-9, aggrecan, and type II collagen, in lacunae embedded in a cartilaginous matrix of type II collagen and proteoglycans. Tissue engineered explants of ATDC5 cells were reminiscent of cartilaginous structures composed of numerous chondrocytes, staining for typical chondrocytic proteins, in lacunae embedded in a matrix of type II collagen and proteoglycans. In comparison, articular chondrocyte explants exhibited areas of Sox-9, aggrecan, and type II collagen-expressing cells growing on fleece, and discrete islands of chondrocytic cells embedded in a cartilaginous matrix.     

Medical ozone therapy as a potential treatment modality for regeneration of damaged articular cartilage in osteoarthritis

Osteoarthritis (OA) is the most common degenerative joint disease and a growing health problem affecting more than half of the population over the age of 65. It is characterized by inflammation in the cartilage and synovium, resulting in the loss of joint structure and progressive damage to the cartilage. Many pro-inflammatory mediators are elevated in OA, including reactive oxygen species (ROS) such as nitric oxide (NO) and hydrogen peroxide (H₂O₂). Damaged articular cartilage remains a challenge to treat due to the limited self-healing capacity of the tissue and unsuccessful biological interventions. This highlights the need for better therapeutic strategies to heal damaged articular cartilage. Ozone (O₃) therapy has been shown to have positive results in the treatment of OA; however the use of O₃ therapy as a therapeutic agent is controversial. There is a perception that O₃ is always toxic, whereas evidence indicates that when it is applied following a specified method, O₃ can be effective in the treatment of degenerative diseases. The mechanism of action of O₃ therapy in OA is not fully understood and this review summarizes the use of O₃ therapy in the treatment of damaged articular cartilage in OA.     

The synthesis of cartilage collagen by rabbit and human chondrocytes in primary cell culture

Summary This report describes a method for preparing primary cell cultures of differentiated rabbit sternal and human vertebral cartilage cells. These cell cultures were shown to synthesize primarily α1 chains, which is taken to mean that at least 82% of the collagen produced is cartilage specific collagen (type II). This work was supported in part by grant HD-05505 from NIH.     

A distinct cohort of progenitor cells participates in synovial joint and articular cartilage formation during mouse limb skeletogenesis

The origin, roles and fate of progenitor cells forming synovial joints during limb skeletogenesis remain largely unclear. Here we produced prenatal and postnatal genetic cell fate-maps by mating ROSA-LacZ-reporter mice with mice expressing Cre-recombinase at prospective joint sites under the control of Gdf5 regulatory sequences (Gdf5-Cre). Reporter-expressing cells initially constituted the interzone, a compact mesenchymal structure representing the first overt sign of joint formation, and displayed a gradient-like distribution along the ventral-to-dorsal axis. The cells expressed genes such as Wnt9a, Erg and collagen IIA, remained predominant in the joint-forming sites over time, gave rise to articular cartilage, synovial lining and other joint tissues, but contributed little if any to underlying growth plate cartilage and shaft. To study their developmental properties more directly, we isolated the joint-forming cells from prospective autopod joint sites using a novel microsurgical procedure and tested them in vitro. The cells displayed a propensity to undergo chondrogenesis that was enhanced by treatment with exogenous rGdf5 but blocked by Wnt9a over-expression. To test roles for such Wnt-mediated anti-chondrogenic capacity in vivo, we created conditional mutants deficient in Wnt/β-catenin signaling using Col2-Cre or Gdf5-Cre. Synovial joints did form in both mutants; however, the joints displayed a defective flat cell layer normally abutting the synovial cavity and expressed markedly reduced levels of lubricin. In sum, our data indicate that cells present at prospective joint sites and expressing Gdf5 constitute a distinct cohort of progenitor cells responsible for limb joint formation. The cells appear to be patterned along specific limb symmetry axes and rely on local signaling tools to make distinct contributions to joint formation.     

Extent of cell differentiation and capacity for cartilage synthesis in human adult adipose‐derived stem cells: Comparison with fetal chondrocytes

This study evaluated the extent of differentiation and cartilage biosynthetic capacity of human adult adipose‐derived stem cells relative to human fetal chondrocytes. Both types of cell were seeded into nonwoven‐mesh polyglycolic acid (PGA) scaffolds and cultured under dynamic conditions with and without addition of TGF‐β1 and insulin. Gene expression for aggrecan and collagen type II was upregulated in the stem cells in the presence of growth factors, and key components of articular cartilage such as glycosaminoglycan (GAG) and collagen type II were synthesized in cultured tissue constructs. However, on a per cell basis and in the presence of growth factors, accumulation of GAG and collagen type II were, respectively, 3.4‐ and 6.1‐fold lower in the stem cell cultures than in the chondrocyte cultures. Although the stem cells synthesized significantly higher levels of total collagen than the chondrocytes, only about 2.4% of this collagen was collagen type II. Relative to cultures without added growth factors, treatment of the stem cells with TGF‐β1 and insulin resulted in a 59% increase in GAG synthesis, but there was no significant change in collagen production even though collagen type II gene expression was upregulated 530‐fold. In contrast, in the chondrocyte cultures, synthesis of collagen type II and levels of collagen type II as a percentage of total collagen more than doubled after growth factors were applied. Although considerable progress has been achieved to develop differentiation strategies and scaffold‐based culture techniques for adult mesenchymal stem cells, the extent of differentiation of human adipose‐derived stem cells in this study and their capacity for cartilage synthesis fell considerably short of those of fetal chondrocytes. Biotechnol. Bioeng. 2010;107: 393–401. © 2010 Wiley Periodicals, Inc.     

Mechanisms of synovial joint and articular cartilage formation: recent advances, but many lingering mysteries

Synovial joints are elegant, critically important, and deceptively simple biomechanical structures. They are comprised of articular cartilage that covers each end of the opposing skeletal elements, synovial fluid that lubricates and nourishes the tissues, ligaments that hold the skeletal elements in check, and a fibrous capsule that insulates the joints from surrounding tissues. Joints also exhibit an exquisite arrays of shapes and sizes, best exemplified by the nearly spherical convex femoral head articulating into a nearly spherical concave hip acetabulum, or a phalangeal joint with two condyles on the distal side articulating in reciprocally-shaped sockets on the opposing proximal side. Though few in number, joint tissues are highly specialized in structure and function. This is illustrated by articular cartilage with its unique extracellular matrix, unique biomechanical resilience, its largely avascular nature, and its ability to persist through life with minimal turnover of its cells and components. The fact that interest in synovial joints has remained unabated for decades is a reflection of their fundamental importance for organism function and quality of life, and for their susceptibility to a variety of acquired and congenital conditions, most importantly arthritis. This has led to many advances in this field that encompass molecular genetics to biomechanics to medicine. Regrettably, what continues to be poorly understood are the mechanisms by which synovial joints actually form in the developing embryo. If available, this information would be not only of indisputable biological interest, but would also have significant biomedical ramifications, particularly in terms of designing novel tissue regeneration or reconstruction therapies. This review focuses on recent advances in understanding the mechanisms of synovial joint formation in the limbs, and places and discusses the information within the context of classic studies and the many mysteries and questions that remain unanswered.     

CCN2: a master regulator of the genesis of bone and cartilage

CCN family member 2 (CCN2), also known as connective tissue growth factor (CTGF), has been suggested to be an endochondral ossification genetic factor that has been termed “ecogenin”, because in vitro studies revealed that CCN2 promotes the proliferation and differentiation of growth-plate chondrocytes, osteoblasts, and vascular endothelial cells, all of which play important roles in endochondral ossification. In addition to its action toward these three types of cells, CCN2 was recently found to promote the formation of osteoclasts in vitro, which cells play an important role in the replacement of cartilage by bone during endochondral ossification, thus strengthening the “ecogenin” hypothesis. For confirmation of this hypothesis, transgenic mice over-expressing CCN2 in cartilage were generated. The results proved the hypothesis; i.e., the over-expression of CCN2 in cartilage stimulated the proliferation and differentiation of growth-plate chondrocytes, resulting in the promotion of endochondral ossification. In addition to its “ecogenin” action, CCN2 had earlier been shown to promote the differentiation of various cartilage cells including articular cartilage cells. In accordance with these findings, cartilage-specific overexpression of CCN2 in the transgenic mice was shown to protect against the development of osteoarthritic changes in aging articular cartilage. Thus, CCN2 may also play a role as an anti-aging (chondroprotective) factor, stabilizing articular cartilage. CCN2 also had been shown to promote intramembranous ossification, regenerate cartilage and bone, and induce angiogenesis in vivo. For understanding of the molecular mechanism underlying such multifunctional actions, yeast two-hybrid analysis, protein array analysis, solid-phase binding assay, and surface plasmon resonance (SPR) analysis have been used to search for binding partners of CCN2. ECMs such as fibronectin and aggrecan, growth factors including BMPs and FGF2 and their receptors such as FGFR1 and 2 and RANK, as well as CCN family members themselves, were shown to bind to CCN2. Regarding the interaction of CCN2 with some of them, various binding modules in the CCN2 molecule have been identified. Therefore, the numerous biological actions of CCN2 would depend on what kinds of binding partners and what levels of them are present in the microenvironment of different types of cells, as well as on the state of differentiation of these cells. Through this mechanism, CCN2 would orchestrate various signaling pathways, acting as a signal conductor to promote harmonized skeletal growth and regeneration.     

Growth and shaping of metacarpal and carpal cartilage anlagen: application of morphometry to the development of short and long bone. A study of human hand anlagen in the fetal period

A histological and morphometric analysis of human metacarpal and carpal anlagen between the 16th and 22nd embryonic weeks was carried out with the aim of studying the establishment of the respective anlage architecture. No differences in the pattern of growth were documented between the peripheral and central zones of the metacarpal epiphyses and those of the carpals. The regulation of longitudinal growth in long bone anlagen occurred in the transition zone between the epiphysis and the diaphysis (homologous to the metaphyseal growth plate cartilage in more advanced developmental stage of the bone). Comparative zonal analysis was conducted to assess the chondrocyte density, the mean chondrocyte lacunar area, the paired chondrocyte polarity in the orthogonal longitudinal and transverse planes, and the lacunar shape transformation in the metacarpal. In transition from epiphysis to diaphysis chondrocyte density decreased and mean lacunar area increased. No significant differences in the chondrocyte maturation cycle were observed between proximal/distal metacarpal epiphyses and the carpal anlagen. The number of paired chondrocyte oriented along the growth vector was significantly higher in both proximal/distal transition zones between epiphysis and diaphysis. Human metacarpals shared with experimental models (like mice and nonmammal tetrapods) an early common chondrocyte maturation cycle but with a different timing due to the slower embryonic and fetal developmental rate of human anlagen.     

A validated model of GAG deposition,cell distribution,and growth of tissue engineered cartilage cultured in a rotating bioreactor

In this work a new phenomenological model of growth of cartilage tissue cultured in a rotating bioreactor is developed. It represents an advancement of a previously derived model of deposition of glycosaminoglycan (GAG) in engineered cartilage by (i) introduction of physiological mechanisms of proteoglycan accumulation in the extracellular matrix (ECM) as well as by correlating (ii) local cell densities and (iii) tissue growth to the ECM composition. In particular, previously established predictions and correlations of local oxygen concentrations and GAG synthesis rates are extended to distinguish cell secreted proteoglycan monomers free to diffuse in cell surroundings and outside from the engineered construct, from large aggrecan molecules, which are constrained within the ECM and practically immovable. The model includes kinetics of aggregation, that is, transformation of mobile GAG species into immobile aggregates as well as maintenance of the normal ECM composition after the physiological GAG concentration is reached by incorporation of a product inhibition term. The model also includes mechanisms of the temporal evolution of cell density distributions and tissue growth under in vitro conditions. After a short initial proliferation phase the total cell number in the construct remains constant, but the local cell distribution is leveled out by GAG accumulation and repulsion due to negative molecular charges. Furthermore, strong repulsive forces result in expansion of the local tissue elements observed macroscopically as tissue growth (i.e., construct enlargement). The model is validated by comparison with experimental data of (i) GAG distribution and leakage, (ii) spatial‐temporal distributions of cells, and (iii) tissue growth reported in previous works. Validation of the model predictive capability—against a selection of measured data that were not used to construct the model—suggests that the model successfully describes the interplay of several simultaneous processes carried out during in vitro cartilage tissue regeneration and indicates that this approach could also be attractive for application in other tissue engineering systems. Biotechnol. Bioeng. 2010. 105: 842–853. © 2009 Wiley Periodicals, Inc.     

Directed differentiation of human mesenchymal stem cells toward a cardiomyogenic fate commitment through formation of cell aggregates

Cell morphology is known to modulate the multipotential lineage commitment of stem cells. We provide a new strategy to induce the early lineage commitment of human mesenchymal stem cells (hMSCs) toward a cardiomyogenic fate through the formation of cell aggregates. A surface-immobilized polyamidoamine dendrimer with fifth generation of dendron structure was used during the culturing of hMSCs. These hMSCs cultured on the G5 surface formed aggregates through active migration and division. More than 22% of cardiac troponin-T (cTnT)-positive (cTnT⁺) cells in aggregates formed on the dendrimer surface; the population formed on the dendrimer surface was higher than that in conventional culture vessel. When cell aggregate was reseeded onto a fresh G5 surface, single cells migrated out of the aggregates, proliferated, and formed new aggregates. This passage method, accompanied with repetitive aggregate dispersion and formation, was applied to cultures over 40 days. The proportion of cTnT⁺ cells increased to 62% by the end of third passage. Our results suggest that culturing hMSCs on G5 surface results in directed commitment of the hMSCs toward a cardiomyocyte-like fate.     

Differentiation and mineralization in chick chondrocytes maintained in a high cell density culture: A model for endochondral ossification

Summary Chondrocytes isolated from the proliferative and differentiating zones of 3-wk-old chick growth plates were cultured in the presence of 10% fetal bovine serum (FBS) and ascorbic acid for up to 21 d in a high cell density culture within Eppendorf tubes. The proliferative, differentiating, and calcification properties of the chondrocytes were examined by immunolocalization and by enzyme histochemical and biochemical methods. The cells maintained a chondrocyte phenotype throughout culture: they were round in shape and synthesized both collagen type II and proteoglycans. The expression of a hypertrophic phenotype was evident by Day 3 of culture and from this time onwards characteristics of terminal differentiation were observed. The cells were positive for both alkaline phosphatase (ALP) activity and c-myc protein and the surrounding matrix stained strongly for collagen type X. Small foci of mineralization associated with individual chondrocytes were first evident by Day 6 and more widespread areas of mineralization occupying large areas of matrix were present by Day 15. Mineralization occurred without the addition of exogenous phosphate to the medium. This culture system displays characteristics that are similar in both morphological and developmental terms to that of chick chondrocyte differentiation and calcification in vivo and therefore offers an excellent in vitro model for endochondral ossification.     

Human blood-derived fibrin releasates: Composition and use for the culture of cell lines and human primary cells

We have evaluated the capacity of two human blood fractions to substitute for FBS as growth medium supplement for human and animal cell cultures. Non-anticoagulated blood from volunteer donors (N = 13) was centrifuged to isolate a supernatant serum (SS) and a platelet-rich fibrin (PRF) clot which was squeezed to extract the releasate (PRFR). Both materials were characterized for the content in PDGF-AB, TGF-β1, VEGF, bFGF, EGF, IGF, total protein, albumin, IgG, IgM IgA, fibrinogen, cholesterol, triglycerides, various chemistry analytes and hemoglobin. Cell growth promoting activity of pooled SS and PRFR at 1, 5, and 10% in growth medium was evaluated over 7 days using human (HEK293, MG-63) and animal (SIRC, 3T3) cell lines and two human primary cultures (gingival fibroblasts and periodontal ligaments). Viable cell count was compared to that in cultures in FBS free-medium and 10% FBS supplement. SS and PRFR at 1-10% stimulated cell growth significantly more than FBS-free medium and in a way similar to 10% FBS in all cultures apart from 3T3. These two human blood-derived fibrin releasates are equally efficient to substitute for FBS as supplement for cell cultures and could be useful for specialized applications in regenerative medicine, dentistry and oral implantology, or cell therapy.     

Identification of the antigenic epitopes in staphylococcal enterotoxins A and E and design of a superantigen for human cancer therapy

Monoclonal antibodies have a potential for cancer therapy that may be further improved by linking them to effector molecules such as superantigens. Tumor targeting of a superantigen leads to a powerful T cell attack against the tumour tissue. Encouraging results have been observed preclinically and in patients using the superantigen staphylococcal enterotoxin A, SEA. To further improve the concept, we have reduced the reactivity to antibodies against superantigens, which is found in all individuals. Using epitope mapping, antibody binding sites in SEA and SEE were found around their MHC class II binding sites. These epitopes were removed genetically and a large number of synthetic superantigens were produced in an iterative engineering procedure. Properties such as decreased binding to anti-SEA as well as higher selectivity to induce killing of tumour cells compared to MHC class II expressing cells, were sequentially improved. The lysine residues 79, 81, 83 and 84 are all part of major antigenic epitopes, Gln204, Lys74, Asp75 and Asn78 are important for optimal killing of tumour cells while Asp45 affects binding to MHC class II. The production properties were optimised by further engineering and a novel synthetic superantigen, SEA/E-120, was designed. It is recognised by approximately 15% of human anti-SEA antibodies and have more potent tumour cell killing properties than SEA. SEA/E-120 is likely to have a low toxicity due to its reduced capacity to mediate killing of MHC class II expressing cells. It is produced as a Fab fusion protein at approximately 35 mg/l in Escherichia coli.     

Isolation,expansion and neural differentiation of stem cells from human plucked hair: a further step towards autologous nerve recovery

Stem cells from the adult hair follicle bulge can differentiate into neurons and glia, which is advantageous for the development of an autologous cell-based therapy for neurological diseases. Consequently, bulge stem cells from plucked hair may increase opportunities for personalized neuroregenerative therapy. Hairs were plucked from the scalps of healthy donors, and the bulges were cultured without prior tissue treatment. Shortly after outgrowth from the bulge, cellular protein expression was established immunohistochemically. The doubling time was calculated upon expansion, and the viability of expanded, cryopreserved cells was assessed after shear stress. The neuroglial differentiation potential was assessed from cryopreserved cells. Shortly after outgrowth, the cells were immunopositive for nestin, SLUG, AP-2α and SOX9, and negative for SOX10. Each bulge yielded approximately 1 × 10⁴ cells after three passages. Doubling time was 3.3 (±1.5) days. Cellular viability did not differ significantly from control cells after shear stress. The cells expressed class III β-tubulin (TUBB3) and synapsin-1 after 3 weeks of neuronal differentiation. Glial differentiation yielded KROX20- and MPZ-immunopositive cells after 2 weeks. We demonstrated that human hair follicle bulge-derived stem cells can be cultivated easily, expanded efficiently and kept frozen until needed. After cryopreservation, the cells were viable and displayed both neuronal and glial differentiation potential.     

Basic studies on gene therapy of human malignant melanoma by use of the human interferon beta gene entrapped in cationic multilamellar liposomes. 1. Morphology and growth rate of six melanoma cell lines used in transfection experiments with the human interferon beta gene

Six cell lines of human malignant melanoma: A375, A375.2, G361, HMV-1, MM8.1 and WM115 were seeded at densities of 1 × 10⁴ cells/ml, 2 × 10⁴ cells/ml or 3 × 10⁴ cells/ml of RPMI medium supplemented with 10% fetal calf serum and antibiotics in a humidified atmosfere of 5% CO₂ at 37°C. A375 cells were also grown in Dulbecco's minimum Eagle's medium (DMEM medium). The morphology was studied by phase contrast light microscopy. At 4 days after seeding the colonies of A375 cells and HMV-1 cells were oval-shaped, the cells were polyhedrical and were making contact with each other regularly. The remaining cells were scattered, more elongated, and made contact randomly. G361 cells and MM8.1 cells tended to form superposed layers before 100% confluency was achieved. There were great differences in the growth rate and doubling time of melanoma cells. The doubling time in day 1 was short (around 6-12 h) in the case of A375, G361 and HMV-1 cells, longer (around 18h) in the case of MM8.1 cells and very long (ranging between 26 and 89 h) for A375.2 and WM115 cells. There were also differences in the doubling time of cells as a function of the cell density at seeding. On the other hand, except for MM8.1 cells, there were differences between the doubling time in day 2 compared to day 1.     

Microenvironment at tissue injury, a key focus for efficient stem cell therapy: A discussion of mesenchymal stem cells

Stem cell therapy is not a new field, as indicated by the success of hematopoietic stem cell reconstitution for various hematological malignancies and immune-mediated disorders. In the case of tissue repair, the major issue is whether stem cells should be implanted, regardless of the type and degree of injury. Mesenchymal stem cells have thus far shown evidence of safety, based on numerous clinical trials, particularly for immune-mediated disorders. The premise behind these trials is to regulate the stimulatory immune responses negatively. To apply stem cells for other disorders, such as acute injuries caused by insults from surgical trauma and myocardial infarction, would require other scientific considerations. This does not imply that such injuries are not accompanied by immune responses. Indeed, acute injuries could accompany infiltration of immune cells to the sites of injuries. The implantation of stem cells within a milieu of inflammation will establish an immediate crosstalk among the stem cells, microenvironmental molecules, and resident and infiltrating immune cells. The responses at the microenvironment of tissue injury could affect distant and nearby organs. This editorial argues that the microenvironment of any tissue injury is a key consideration for effective stem cell therapy.