A three-dimensional in vitro culture environment of a novel polymer scaffold, yielding chondroprogenitors and mesenchymal stem cells in human chondrocytes derived from osteoarthritis-affected cartilage tissue.

OBJECTIVE: We evaluated the expression of stem/progenitor biomarkers in osteoarthritic tissue derived chondrocytes cultured using a three-dimensional (3D) thermo-reversible gelation polymer (TGP). METHODS: The chondrocytes from discarded biopsy tissues obtained from human elderly patients with osteoarthritis were cultured using the 3D-TGP up to six weeks. RESULTS: The chondrocytes grew in a tissue-like manner, without de-differentiation into fibroblasts, and the cells thus tissue-engineered were proven positive for CD49e, OCT4, CD-105 and STRO-1 by immunohistochemistry. CONCLUSION: This study establishes the efficacy of this 3D-TGP platform for clinically useable in-vitro tissue-engineered cartilage for improvising the clinical outcome of cell therapy for cartilage repair.

The clinical significance of CD49e and CD56 for multiple myeloma in the novel agents era.

Multiple myeloma (MM) is a hematological malignancy characterized by the proliferation of abnormal plasma cells in bone marrow. Flow cytometry distinguishes between normal and abnormal plasma cells by evaluating cluster of differentiation (CD) 56 and CD19 expression patterns. Moreover, immunophenotyping of mature plasma cell 1 (MPC-1) and very late antigen-5 (CD49e) identifies the maturity of MM as mature (MPC-1+, CD49e+), intermediate (MPC-1+, CD49e-), or immature (MPC-1-, CD49e-). We retrospectively examined the effects of surface marker expression and maturity subtype on overall survival (OS) and time to next treatment (TNT) among 55 patients (25 males, 30 females) with symptomatic MM. All patients were treated with regimens containing bortezomib (BOR) (n = 39) or lenalidomide (LEN) (n = 16) as the initial treatment. Median age at diagnosis was 72 years (range: 36-88). The lack of CD56, an aberrant marker, was associated with significantly worse prognosis compared with CD56+ MM (median OS: 24 vs. 60 months, respectively; p = 0.0050). In CD49e+ MM, defined as mature type, no significant difference was seen in TNT of the initial treatment, regardless of whether it was a BOR-based regimen or LEN + dexamethasone (Ld) therapy. On the other hand, in CD49e- MM, defined as immature/intermediate type, TNT of Ld therapy was significantly longer than that of BOR-based regimens (median TNT: undefined vs. 12 months, respectively; p = 0.0043). These results suggest that Ld therapy is more effective than BOR-based therapy for CD49e- MM and thus may aid regimen-related decisions in the novel agents era.

Evaluation of CD49e as a distinguishing marker for human articular cartilage derived chondroprogenitors.

BACKGROUND: Cell-based therapy in cartilage repair can benefit from the use of chondroprogenitors; a cell type classified as mesenchymal stem cells, demonstrating reduced hypertrophy. Fibronectin, routinely used to isolate chondroprogenitors, classically binds to α5ß1 integrins (CD49e + CD29), of which CD49e is said to be highly expressed in progenitors. The aim of our study was to assess the specificity of CD49e as a distinguishing marker for chondroprogenitors; because studies report low expression in fresh chondrocytes (FCs), but recent conflicting data has exhibited incremental expression of CD49e in cultured chondrocytes. METHODS: FCs were isolated from three human osteoarthritic knee joints and CD49e- cells (sorted by flow cytometry) were cultured in adherent and non-adherent conditions and reassessed for CD49e and CD29 at multiple time points. Colony-forming efficiency (CFE) following fibronectin adhesion assay was calculated for FC, CD49e+ and CD49e- cells. RESULTS: A statistically significant increase in CD49e and CD29 expression was seen in both adherent and non-adherent cultures of CD49e- cells (P < 0.01), as early as 24 h. All groups grew clonally and CFE was similar without any significant difference. CD49e- chondrocytes turned positive when cultured, possibly due to an inherent phenotypic drift, seen after release from cartilage and not because of plastic adherence or chondroprogenitor overgrowth, as non-adherent cultures also showed high expression. CONCLUSIONS: As the specificity of CD49e is questionable, there is a pressing need for a specific differentiating marker, to isolate a pure population of chondroprogenitors, as this cell type shows inherent chondrogenesis and reduced hypertrophy, both requisites for cartilage repair.

Comparative analysis of fresh chondrocytes, cultured chondrocytes and chondroprogenitors derived from human articular cartilage.

INTRODUCTION: Interest in chondroprogenitors arose due to their inherent stem cell like properties, and their initial characterization was based on identification of a small percentage of CD49e positive cells in cultured chondrocytes (CC). It was further noted that when fresh chondrocytes (FC; reported to express low CD49e) were subjected to fibronectin adhesion assay, an isolate of chondroprogenitors was obtained, which was highly positive for CD49e, thus making it a distinguishing marker for this cell population. However, this notion was challenged when reports demonstrated high CD49e expression in CC as well. Therefore, our aim was to compare CD49e expression in FC, CC and chondroprogenitors. METHODS: Chondrocytes and chondroprogenitors were isolated from articular cartilage of osteoarthritic joints from three patients. Assessment of classic fibronectin receptor (CD49e, CD29), positive (CD105, CD73, CD90) and negative (CD45, CD34) mesenchymal stem cell marker expression in all groups was performed, as chondroprogenitors fulfill the minimal criteria laid down by International Society for Cellular Therapy. Following this, adipogenic, osteogenic and chondrogenic differentiation was assessed by Oil red O, Alizarin Red and Alcian Blue staining respectively. RESULTS AND CONCLUSION: Our observations indicate that FC show significantly low surface marker expression as compared to CC and chondroprogenitors, whereas no significant difference was seen in values when CC and chondroprogenitors were compared. Moreover, comparable results were exhibited when trilineage differentiation potential was compared across groups. Since CC and chondroprogenitors show similar characteristics, there is a pressing need for a specific differentiating marker to isolate a pure population of chondroprogenitors.

Plasma cell maturity as a predictor of prognosis in multiple myeloma.

In this study, the impact of plasma cell maturity on the prognoses of multiple myeloma (MM) patients in the era of novel agents was investigated. Myeloma cell maturity was classified via immunophenotyping: myeloma cells showing mature plasma cell 1 (MPC-1)-positive and CD49e-positive cells were considered mature type; MPC-1-positive and CD49e-negative cells were considered intermediate type; and MPC-1-negative cells were considered immature type. This study included 87 newly diagnosed MM patients who were initially treated with bortezomib and/or chemotherapy. Myeloma cell maturity was a critical factor affecting overall survival (OS) in the cohort, with median OS not reached in mature-type, 50 months in intermediate-type, and 20 months in immature-type cells. Multivariate analysis showed that immature type and stage III according to the International Staging System were both independent prognostic factors affecting OS. The findings of this study demonstrate the clinical importance of myeloma cell classification according to immunophenotyping using MPC-1 and CD49e antibodies to determine patient prognosis in this era of novel therapeutic agents.

Clinical significance of granule-containing myeloma cells in patients with newly diagnosed multiple myeloma.

The clinical features and prognostic significance of myeloma cells containing granules remain unclear. The purpose of this retrospective study was to investigate the clinical significance of granule-containing myeloma cells in patients with newly diagnosed multiple myeloma (NDMM). We retrospectively analyzed the records of 122 patients diagnosed with NDMM between January 2007 and December 2013. Granule-containing myeloma cells were defined as myeloma cells that exhibited three or more granules in their cytoplasm by May-Giemsa staining. The patients were classified into two groups, the granule-containing myeloma (GM) and nongranule-containing myeloma (non-GM) groups, depending on the proportion of myeloma cells that contained granules (cut-off value: 10%). There were 25 (20.5%) patients in the GM group. Patients in the GM group displayed significantly higher CD56 and CD49e expression than those in the non-GM group (t-test, P = 0.027 and 0.042). None of the patient characteristics differed significantly between the two groups. There was no significant difference in the chemotherapy profiles of the two groups, and the overall response rates of the two groups were similar. During the median follow-up period of 33.9 months, the overall survival (OS) in the GM group was similar to that in the non-GM group; 4-year OS of the GM and non-GM groups were 78.5% and 51.9%, respectively (P = 0.126). We concluded that cases of NDMM involving granule-containing myeloma cells are not infrequent. Moreover, CD56 and CD49e expression was significantly higher in the presence of myeloma cell populations, and the presence of granules did not affect survival.