[Show abstract][Hide abstract] ABSTRACT: According to a Prognoscan database, upregulation of Bruton's tyrosine kinase (Btk) is associated with low overall survival in ovarian cancer patients. We found that spheroids-forming ovarian cancer cell, which highly expressed cancer stem-like cell (CSC) markers and Btk, were cisplatin resistant. We next treated CSCs and non-CSCs by a combination of ibrutinib and cisplatin. We found that chemoresistance was dependent on Btk and JAK2/STAT3, which maintained CSC by inducing Sox-2 and prosurvival genes. We suggest that addition of ibrutinib to cisplatin may improve treatment outcome in ovarian cancer.
[Show abstract][Hide abstract] ABSTRACT: The aberrant activation of Wnt/β-catenin signaling plays an important role in the carcinogenesis and progression of hepatocellular carcinoma (HCC). Therefore, the Wnt/β-catenin signaling molecules are attractive candidates for the development of targeted therapies for this disease. The present study showed that destruxin B (DB) inhibits the proliferation and induces the apoptosis of HCC cells by decreasing the protein expression of anti-apoptotic Bcl-2 and Bcl-xL and increasing the expression of the proapoptotic protein Bax. More importantly, DB also attenuates Wnt-signaling in HCC cells by downregulating β-catenin, Tcf4, and β-catenin/Tcf4 transcriptional activity, which results in the decreased expression of β-catenin target genes, such as cyclin D1, c-myc, and survivin. Furthermore, DB affects the migratory and invasive abilities of Sk-Hep1 cells through the suppression of markers of the epithelial-mesenchymal transition (EMT). A synergistic anti-proliferative and migratory effect was achieved using the combination of DB and sorafenib in Sk-Hep1 cells. In conclusion, DB acts as a novel Wnt/β-catenin inhibitor and reduces the aggressiveness and invasive potential of HCC by altering the cells' EMT status and mobility. DB in combination with sorafenib may be considered for future clinical use for the management of metastatic HCC.
Toxicology in Vitro 01/2014; 28(4). DOI:10.1016/j.tiv.2014.01.002 · 3.21 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Lung cancer is the leading cause of cancer deaths worldwide and current therapies fail to treat this disease in majority of cases. Antrodia camphorata (AC), is a medicinal mushroom being widely used as food dietary supplement for cancer prevention. The sesquiterpene lactone antrocin is the most potent among more than one hundred secondary metabolites isolated from AC. However, the molecular mechanisms of antrocin-mediated anti-cancer effects remain unclear. In this study, we found that antrocin inhibited cell proliferation in two non-small cell lung cancer cells, namely H441 (wild-type EGFR, IC50= 0.75μM) and H1975 (gefitnib-resistant mutant T790M, IC50=0.83μM). Antrocin dose-dependently suppressed colony formation and induced apoptosis as evidenced by activated caspase-3 and increased Bax/Bcl2 ratio. Gene profiling studies indicated that antrocin downregulated JAK/STAT signaling pathway. We further demonstrated that antrocin suppressed both constitutively activated and IL-6-induced STAT3 phosphorylation and its subsequent nuclear translocation. Such inhibition is found to be achieved through the suppression of JAK2 and interaction between STAT3 and ERK. Additionally, antrocin increased microRNA let-7c expression and suppressed STAT signaling. The combination of antrocin and JAK2/STAT3 gene silencing significantly increased apoptosis in H441 cells. Such dual interruption of JAK2 and STAT3 pathways also induced downregulation of antiapoptotic protein mcl-1 and increased caspase-3 expression. In vivo, intraperitoneal administration of antrocin significantly suppressed the growth of lung cancer tumor xenografts. Our results indicate that antrocin may be a potential therapeutic agent for human lung cancer cells through constitutive inhibition of JAK2/STAT3 pathway.
[Show abstract][Hide abstract] ABSTRACT: SCOPE: Tumor-associated macrophages (TAMs) have been shown to promote metastasis and malignancy. Pterostilbene, a natural stilbene isolated from blueberries, has been suggested for anti-cancer effects. Here, we explored the potential cancer stem cells (CSCs)/TAM modulating effects of pterostilbene in breast cancer. METHODS AND RESULTS: Using flowcytometric and Boyden chamber assay, we showed MCF7 and MDA-MB-231 cells cocultured with M2 TAMs exhibited increased percentage of CD44(+) /CD24(-) CSC population and migratory/invasive abilities. RT-PCR results showed that CD44(+) /CD24(-) cells expressed an increased level of HIF-1α, β-catenin, Twist1, and NF-κB and enhanced tumor sphere forming ability. Additionally, pterostilbene treatment dose dependently overcame M2 TAM-induced enrichment of CSCs and metastatic potential of breast cancer cells. Mechanistically, pterostilbene suppressed NFκB, Twist1, vimentin, and increased E-cadherin expression. Using siRNA technique, we demonstrated that pterostilbene-mediated NFκB downregulation was correlated to an increased amount of microRNA 448. Finally, pterostilbene-mediated suppression in tumorigenesis and metastasis was validated by noninvasive bioluminescence in mice bearing M2 TAM cocultured MDA-MB-231 tumor. CONCLUSION: Pterostilbene effectively suppresses the generation of CSCs and metastatic potential under the influence of M2 TAMs via modulating EMT associated signaling pathways, specifically NF-κB/miR488 circuit. Thus, pterostilbene could be an ideal anti-CSC agent in clinical settings.
[Show abstract][Hide abstract] ABSTRACT: For many malignancies, radiation therapy remains the second option only to surgery in terms of its curative potential. However, radiation-induced tumor cell death is limited by a number of factors, including the adverse response of the tumor microenvironment to the treatment and either intrinsic or acquired mechanisms of evasive resistance, and the existence of cancer stem cells (CSCs). In this study, we demonstrated that using different doses of irradiation led to the enrichment of CD133(+) Mahlavu cells using flow cytometric method. Subsequently, CD133(+) Mahlavu cells enriched by irradiation were characterized for their stemness gene expression, self-renewal, migration/invasion abilities, and radiation resistance. Having established irradiation-enriched CD133(+) Mahlavu cells with CSC properties, we evaluated a phytochemical, pterostilbene (PT), found abundantly in blueberries, against irradiation-enriched CSCs. It was shown that PT treatment dose-dependently reduced the enrichment of CD133(+) Mahlavu cells upon irradiation; PT treatment also prevented tumor sphere formation, reduced stemness gene expression, and suppressed invasion and migration abilities as well as increasing apoptosis of CD133(+) Mahlavu CSCs. Based on our experimental data, pterostilbene could be used to prevent the enrichment of CD133(+) hepatoma CSCs and should be considered for future clinical testing as a combined agent for HCC patients.
Evidence-based Complementary and Alternative Medicine 06/2013; 2013:258425. DOI:10.1155/2013/258425 · 1.88 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Drug resistance and tumor recurrence are major obstacles in treating lung cancer patients. Accumulating evidence considers lung cancer stem cells (CSCs) as the major contributor to these clinical challenges. Agents that can target lung CSCs could potentially provide a more effective treatment than traditional chemotherapy. Here, we utilized the side-population (SP) method to isolate lung CSCs from A549 and PC-9 cell lines. Subsequently, a high throughput platform, connectivity maps (CMAPs), was used to identify potential anti-CSC agents. An antibiotic, antimycin A (AMA), was identified as a top candidate. SP A549 cells exhibited an elevated stemness profile, including Nanog, β -catenin, Sox2, and CD133, and increased self-renewal ability. AMA treatment was found to suppress β -catenin signaling components and tumor sphere formation. Furthermore, AMA treatment decreased the proliferation of gefitinib-resistant PC-9/GR cells and percentage of SP population. AMA demonstrated synergistic suppression of PC-9/GR cell viability when combined with gefitinib. Finally, AMA treatment suppressed tumorigenesis in mice inoculated with A549 SP cells. Collectively, we have identified AMA using CMAP as a novel antilung CSC agent, which acts to downregulate β -catenin signaling. The combination of AMA and targeted therapeutic agents could be considered for overcoming drug resistance and relapse in lung cancer patients.
Evidence-based Complementary and Alternative Medicine 06/2013; 2013:910451. DOI:10.1155/2013/910451 · 1.88 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Platinum-based chemotherapy is the first-line treatment for non-small cell lung cancer, but recurrence occurs in most patients. Recent evidence suggests that CD133+ cells are the cause of drug resistance and tumor recurrence. However, the correlation between chemotherapy and regulation of CD133+ cells has not been investigated methodically. In this study, we revealed that CD133+ lung cancer cells labeled by a human CD133 promoter-driven GFP reporter exhibited drug resistance and stem cell characteristics. Treatment of H460 and H661 cell lines with low-dose cisplatin (IC20) was sufficient to enrich CD133+ cells, to induce DNA damage responses, and to up-regulate ABCG2 and ABCB1 expression, which therefore increased the cross-resistance to doxorubicin and paclitaxel. This cisplatin-induced enrichment of CD133+ cells was mediated through Notch signaling as judged by increased levels of cleaved Notch1 (NICD1). Pretreatment with the γ-secretase inhibitor, DAPT, or Notch1 shRNAs remarkably reduced the cisplatin-induced enrichment of CD133+ cells and increased the sensitivity to doxorubicin and paclitaxel. Ectopic expression of NICD1 reversed the action of DAPT on drug sensitivity. Immunohistochemistry showed that CD133+ cells were significantly increased in the relapsed tumors in 3 of 6 lung cancer patients, who have received cisplatin treatment. A similar effect was observed in animal experiments as cisplatin treatment increased Notch1 cleavage and the ratio of CD133+ cells in engrafted tumors. Intratumoral injection of DAPT with cisplatin treatment significantly reduced CD133+ cell number. Together, our results demonstrated that cisplatin induces the enrichment of CD133+ cells, leading to multi-drug resistance by the activation of Notch signaling.
Cancer Research 11/2012; 73(1). DOI:10.1158/0008-5472.CAN-12-1733 · 9.28 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: RATIONALE: Cancer stem cell (CSC) theory has drawn much attention, with evidence supporting the contribution of stem cells to tumor initiation, relapse and therapy resistance. OBJECTIVES: To screen drugs that target CSCs to improve the current treatment outcome and overcome drug resistance in lung cancer patients. METHODS: We used publicly available embryonic stem cell and CSC-associated gene signatures to query the Connectivity Map for potential drugs that can, at least in part, reverse the gene expression profile of CSCs. High scores were noted for several phenothiazine-like anti-psychotic drugs, including trifluoperazine. We then treated lung CSCs with different EGFR mutation status with trifluoperazine to examine its anti-CSC properties. Lung CSCs resistant to epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI) or cisplatin were treated with trifluoperazine plus gefitinib or trifluoperazine plus cisplatin. Animal models were used for in vivo validation of the anti-CSC effect and synergistic effect of trifluoperazine with gefitinib. MEASUREMENTS AND MAIN RESULTS: We demonstrated that trifluoperazine inhibited CSC tumor spheroid formation and down-regulated the expression of CSC markers (CD44/CD133). Trifluoperazine inhibited Wnt/β-catenin signaling in gefitinib-resistant lung cancer spheroids. The combination of trifluoperazine with either gefitinib or cisplatin overcame drug resistance in lung CSCs. Trifluoperazine inhibited the tumor growth and enhanced the inhibitory activity of gefitinib in lung cancer metastatic and orthotopic CSC animal models. CONCLUSIONS: Using in silico drug screening via Connectivity Map followed by empirical validations, we repurposed an existing phenothiazine-like anti-psychotic drug, trifluoperazine, as a potential anti-CSC agent that could overcome EGFR-TKI and chemotherapy resistance.
American Journal of Respiratory and Critical Care Medicine 09/2012; 186(11). DOI:10.1164/rccm.201207-1180OC · 11.99 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Sulforaphane (SFN) has been indicated for the prevention and suppression of tumorigenesis in solid tumors. Herein, we evaluated SFN's effects on imatinib (IM)-resistant leukemia stem cells (LSCs). CD34(+)/CD38(-) and CD34(+)/CD38(+) LSCs were isolated from KU812 cell line flowcytometrically. Isolated LSCs showed high expression of Oct4, CD133, β-catenin, and Sox2 and IM resistance. Differentially, CD34(+)/CD38(-) LSCs demonstrated higher BCR-ABL and β-catenin expression and imatinib (IM) resistance than CD34(+)/CD38(+) counterparts. IM and SFN combined treatment sensitized CD34(+)/CD38(-) LSCs and induced apoptosis, shown by increased caspase 3, PARP, and Bax while decreased Bcl-2 expression. Additionally, the combined treatment reduced BCR-ABL and β-catenin and MDR-1 protein expression. Mechanistically, IM and SFN combined treatment resensitized LSCs by inducing intracellular reactive oxygen species (ROS). Importantly, β-catenin-silenced LSCs exhibited reduced glutathione S-transferase pi 1 (GSTP1) expression and intracellular GSH level, which led to increased sensitivity toward IM and SFN. We demonstrated that IM and SFN combined treatment effectively eliminated CD34(+)/CD38(-) LSCs. Since SFN has been shown well tolerated in both animals and human, this regimen could be considered for clinical trials.
Journal of Agricultural and Food Chemistry 06/2012; 60(28):7031-9. DOI:10.1021/jf301981n · 3.11 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Adipose-derived stem cells (ADSCs) have been shown to be pluoripotent and explored for their usage in tissue engineering. Previously, we have established a cell-based approach comprised of platelet-enriched plasma and osteo-progenitor cells for treating osteoporosis in an ovariectomized-senescence-accelerated mice (OVX-SAMP8) model. In the present study, we intend to explore the feasibility of using ADSCs as a cell-based therapeutic approach for treating osteoporosis, and to examine the effects of aging on the pluoripotency of ADSCs and the efficiency of bone formation both in vitro and in vivo. Flow cytometry was used to characterize ADSCs isolated from young and aged female SAMP8 mice and showed that the highly positive expression of surface markers such as CD44 and CD105 and negative for CD34 and CD45. Therefore, to compare the aging effects on the growth kinetics and differentiation potential of young and aged ADSCs, we found that there was a significant decline in both the proliferation rate (approximately 13.3%) and osteo-differentiation potential in aged ADSC. Subsequently, young and aged ADSCs were transplanted into the bone marrow of osteoporotic mice (OVX-SAMP8) to evaluate their bone formation ability. ADSC transplants were shown effective in restoring bone mineral density in the right/left knees, femurs and spine, 4 months post-transplantation; mice which received young ADSC transplants showed significantly higher bone regeneration (an average of 24.3% of improved BMD) over those received aged ADSCs. In conclusion, these findings showed that aging impedes osteoporosis-ameliorating potential of ADSC by diminishing osteogenic signal, and that ADSC could be used as a potential cell-based therapy for osteoporosis.
[Show abstract][Hide abstract] ABSTRACT: In continuation to our studies toward the identification of direct anti-cancer targets, here we showed that destruxin B (DB) from Metarhizium anisopliae suppressed the proliferation and induced cell cycle arrest in human colorectal cancer (CRC) HT29, SW480 and HCT116 cells. Additionally, DB induced apoptosis in HT29 cells by decreased expression level of anti-apoptotic proteins Bcl-2 and Bcl-xL while increased pro-apoptotic Bax. On the other hand, DB attenuated Wnt-signaling by downregulation of β-catenin, Tcf4 and β-catenin/Tcf4 transcriptional activity, concomitantly with decreased expression of β-catenin target genes cyclin D1, c-myc and survivin. Furthermore, DB affected the migratory and invasive ability of HT29 cells through suppressed MMPs-2 and -9 enzymatic activities. We also found that DB targeted the MAPK and/or PI3K/Akt pathway by reduced expression of Akt, IKK-α, JNK, NF-κB, c-Jun and c-Fos while increased that of IκBα. Finally, we demonstrated that DB inhibited tumorigenesis in HT29 xenograft mice using non-invasive bioluminescence technique. Consistently, tumor samples from DB-treated mice demonstrated suppressed expression of β-catenin, cyclin D1, survivin, and endothelial marker CD31 while increased caspase-3 expression. Collectively, our data supports DB as an inhibitor of Wnt/β-catenin/Tcf signaling pathway that may be beneficial in the CRC management.
[Show abstract][Hide abstract] ABSTRACT: An ideal anticancer strategy should target only the malignant cells but spare the normal ones. In this regard, we established a platform, consisting of an antigen-delivering vehicle and a protein vaccine, for developing an immunotherapeutic approach with the potential for eliminating various cancer types. Mesenchymal stem cells (MSCs) have been demonstrated capable of targeting tumors and integrating into the stroma. Moreover, we have developed a protein vaccine PE(ΔIII)-E7-KDEL3 which specifically recognized E7 antigen and elicited immunity against cervical cancer. Taking advantage of tumor-homing property of MSCs and PE(ΔIII)-E7-KDEL3, we used E6/E7-immortalized human MSCs (KP-hMSCs) as an E7 antigen-delivering vehicle to test if this protein vaccine could effectively eliminate non-E7-expressing tumor cells. Animals which received combined treatment of KP-hMSCs and PE(ΔIII)-E7-KDEL3 demonstrated a significant inhibition of tumor growth and lung-metastasis when compared to PE(ΔIII)-E7-KDEL3 only and KP-hMSCs only groups. The efficiency of tumor suppression correlated positively to the specific immune response induced by PE(ΔIII)-E7-KDEL3. In addition, this combined treatment inhibited tumor growth via inducing apoptosis. Our findings indicated that KP-hMSCs could be used as a tumor-targeting device and mediate antitumor effect of PE(ΔIII)-E7-KDEL3. We believe this strategy could serve as a platform for developing a universal vaccine for different cancer types.
[Show abstract][Hide abstract] ABSTRACT: The aim of this study was to develop a new diagnostic and therapeutic approach for the treatment of osteoporosis. Previously, we demonstrated that intraosseous transplantation of platelet-rich plasma (PRP) treated-osteoblast-like cells into ovariectomized senescence-accelerated mice (OVX-SAMP8) prevented the development of osteoporosis. In continuation, we aimed to explore the complex etiology of osteoporosis using this platform. An inverse relationship between bone marrow adipogenesis and osteogenesis has been suggested in the development of osteoporosis but the underlying mechanisms remain poorly described. To address these issues, we used PRP to inhibit adipocyte differentiation by promoting osteoblastic differentiation in adipocytes. In addition, a positive correlation between an increase in bone marrow adipocytes and bone loss was established. We assessed this relationship using an osteoporotic animal disease model which consisted of young (for prevention) and old (for treatment) OVX-SAMP8 mice. This animal model demonstrated that PRP treatment mainly exerted its action via promoting bone regeneration but also appeared to suppress adipogenesis within the marrow. The findings and methodology of this study could potentially be applied in the prevention and treatment of osteoporosis.
[Show abstract][Hide abstract] ABSTRACT: This study was carried out to provide a platform for the pre-clinical evaluation of anti-cancer properties of a unique CAM (complementary and alternative medicine) agent,
alcohol extract (ACAE), in a mouse model with the advantageous non-invasive
bioluminescence molecular imaging technology.
analyses on the proliferation, migration/invasion, cell cycle and apoptosis were performed on ACAE-treated non-small cell lung cancer cells, H441GL and control CGL1 cells.
, immune-deficient mice were inoculated subcutaneously with H441GL followed by oral gavages of ACAE. The effect of ACAE on tumor progression was monitored by non-invasive bioluminescence imaging. The proliferation and migration/invasion of H441GL cells were inhibited by ACAE in a dose-dependent manner. In addition, ACAE induced cell cycle arrest at G0/G1 phase and apoptosis in H441GL cells as shown by flow cytometric analysis, Annexin-V immunoflourescence and DNA fragmentation.
bioluminescence imaging revealed that tumorigenesis was significantly retarded by oral treatment of ACAE in a dose-dependent fashion. Based on our experimental data, ACAE contains anti-cancer properties and could be considered as a potential CAM agent in future clinical evaluation.
Evidence-based Complementary and Alternative Medicine 03/2011; 2011(1):914561. DOI:10.1093/ecam/nep228 · 1.88 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The PI3K/Akt/mTOR pathway is considered to be an attractive target for the development of novel anticancer molecules. This paper reports for the first time that a small molecule, antrocin (MW = 234), from Antrodia camphorata was a potent antagonist in various cancer types, being highest in metastatic breast cancer MDA-MB-231 cells (MMCs) with an IC(50) value of 0.6 μM. Antrocin was a superior antiproliferator in MMCs as compared with doxorubicin and cisplatin, prevents colony formation, and was nontoxic to nontumorgenic MCF10A and HS-68 cells. Antrocin induced dose-dependent apoptosis in MMCs and caused cleavage of caspase-3 and poly(ADP-ribose) polymerase. Antrocin also caused a time-dependent decrease in protein expression of anti-apoptotic Bcl-2, Bcl-xL, survivin, and their mRNA, with concomitant increase in pro-apoptotic Bax and cytosolic cytochrome c. In a mechanistic study, antrocin suppressed the phosphorylation of Akt and its downstream effectors mTOR, GSK-3β, and NF-κB. Furthermore, down-regulation of Akt by small interfering RNA prior to antrocin treatment resulted in enhanced cell growth inhibition and apoptosis. Thus, antrocin as an Akt/mTOR dual inhibitor has broad applicability in the development of a clinical trial candidate for the treatment of metastatic breast cancer.
Chemical Research in Toxicology 02/2011; 24(2):238-45. DOI:10.1021/tx100318m · 4.19 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The aim of this study was to examine the underlying signaling mechanisms of arsenic trioxide (ATO)-mediated anticancer effects and the responsible biomarker(s) for the acquired resistance in human heptatocellular carcinoma (HCC).
The therapeutic effects of ATO were examined using 2 characteristically distinct HCC cell lines, Hep-J5 (overexpressing HIF-1α/GRP78) and SK-Hep-1 (the matched control). ATO-mediated proliferation inhibition, oxidative stress, and apoptosis were analyzed using flowcytometric analysis and western blotting. The role of HIF-1α and GRP78 in HCC resistance to ATO treatment was determined using RNA silencing and inhibitor approaches.
SK-Hep-1 cells, lacking both HIF-1α and GRP78 expressions were responsive to ATO-induced apoptosis via an oxidative-nitrosative mechanism. Intracellular glutathione depletion and lipid peroxidation have been identified as the early cascade of events preceding apoptosis via cytochrome c release and the severe drop of mitochondrial membrane potential (MMP). Conversely, Hep-J5 cells, with normoxic coexpression of HIF-1α and GRP78, were resistant to ATO-induced apoptosis. GRP78-silenced Hep-J5 cells remained resistant to ATO treatment. In contrast, ATO resistance in Hep-J5 cells was overcome by the addition of YC-1, a HIF-1α inhibitor.
HIF-1α was identified as the major positive modifier for ATO resistance acquisition in HCC, and it represents a prime molecular target for overcoming ATO resistance.
[Show abstract][Hide abstract] ABSTRACT: Osteoarthritis is characterized by an imbalance in cartilage homeostasis, which could potentially be corrected by mesenchymal stem cell (MSC)-based therapies. However, in vivo implantation of undifferentiated MSCs has led to unexpected results. This study was undertaken to establish a model for preconditioning of MSCs toward chondrogenesis as a more effective clinical tool for cartilage regeneration.
A coculture preconditioning system was used to improve the chondrogenic potential of human MSCs and to study the detailed stages of chondrogenesis of MSCs, using a human MSC line, Kp-hMSC, in commitment cocultures with a human chondrocyte line, hPi (labeled with green fluorescent protein [GFP]). In addition, committed MSCs were seeded into a collagen scaffold and analyzed for their neocartilage-forming ability.
Coculture of hPi-GFP chondrocytes with Kp-hMSCs induced chondrogenesis, as indicated by the increased expression of chondrogenic genes and accumulation of chondrogenic matrix, but with no effect on osteogenic markers. The chondrogenic process of committed MSCs was initiated with highly activated chondrogenic adhesion molecules and stimulated cartilage developmental growth factors, including members of the transforming growth factor beta superfamily and their downstream regulators, the Smads, as well as endothelial growth factor, fibroblast growth factor, insulin-like growth factor, and vascular endothelial growth factor. Furthermore, committed Kp-hMSCs acquired neocartilage-forming potential within the collagen scaffold.
These findings help define the molecular markers of chondrogenesis and more accurately delineate the stages of chondrogenesis during chondrocytic differentiation of human MSCs. The results indicate that human MSCs committed to the chondroprogenitor stage of chondrocytic differentiation undergo detailed chondrogenic changes. This model of in vitro chondrogenesis of human MSCs represents an advance in cell-based transplantation for future clinical use.