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Characterization of human dental pulp cells grown in chemically defined serum-free medium

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      Abstract

      Dental pulp cells (DPCs) are promising candidates for use as transplantable cells in regenerative medicine. However, ex vivo expansion of these cells typically requires culture media containing fetal bovine serum, which may cause infection and immunological reaction following transplantation. In addition, the proliferation and differentiation of DPCs markedly depend upon serum batches. Therefore, the present study examined whether DPCs could be expanded under serum-free conditions. DPCs obtained from four donors were identified to proliferate actively in the serum-free medium, STK2, when compared with those cells in control medium (Dulbecco's modified Eagle's medium containing 10% serum). The high proliferative potential with STK2 was maintained through multiple successive culture passages. DNA microarray analyses demonstrated that the gene expression profile of DPCs grown in STK2 was similar to that of cells grown in the control medium; however, a number of genes related to cell proliferation, including placental growth factor and inhibin-βE, were upregulated in the STK2 cultures. Following induction of osteogenesis, DPCs grown in STK2 induced alkaline phosphatase activity and calcification at higher levels compared with the control medium cultures, indicating maintenance of differentiation potential in STK2. This serum-free culture system with DPCs may have applications in further experimental studies and as a clinical strategy in regenerative medicine.

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      Most cited references 29

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      Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method.

       K Livak,  T Schmittgen (2001)
      The two most commonly used methods to analyze data from real-time, quantitative PCR experiments are absolute quantification and relative quantification. Absolute quantification determines the input copy number, usually by relating the PCR signal to a standard curve. Relative quantification relates the PCR signal of the target transcript in a treatment group to that of another sample such as an untreated control. The 2(-Delta Delta C(T)) method is a convenient way to analyze the relative changes in gene expression from real-time quantitative PCR experiments. The purpose of this report is to present the derivation, assumptions, and applications of the 2(-Delta Delta C(T)) method. In addition, we present the derivation and applications of two variations of the 2(-Delta Delta C(T)) method that may be useful in the analysis of real-time, quantitative PCR data. Copyright 2001 Elsevier Science (USA).
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        Gene Expression Omnibus: NCBI gene expression and hybridization array data repository.

        The Gene Expression Omnibus (GEO) project was initiated in response to the growing demand for a public repository for high-throughput gene expression data. GEO provides a flexible and open design that facilitates submission, storage and retrieval of heterogeneous data sets from high-throughput gene expression and genomic hybridization experiments. GEO is not intended to replace in house gene expression databases that benefit from coherent data sets, and which are constructed to facilitate a particular analytic method, but rather complement these by acting as a tertiary, central data distribution hub. The three central data entities of GEO are platforms, samples and series, and were designed with gene expression and genomic hybridization experiments in mind. A platform is, essentially, a list of probes that define what set of molecules may be detected. A sample describes the set of molecules that are being probed and references a single platform used to generate its molecular abundance data. A series organizes samples into the meaningful data sets which make up an experiment. The GEO repository is publicly accessible through the World Wide Web at http://www.ncbi.nlm.nih.gov/geo.
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          Postnatal human dental pulp stem cells (DPSCs) in vitro and in vivo.

          Dentinal repair in the postnatal organism occurs through the activity of specialized cells, odontoblasts, that are thought to be maintained by an as yet undefined precursor population associated with pulp tissue. In this study, we isolated a clonogenic, rapidly proliferative population of cells from adult human dental pulp. These DPSCs were then compared with human bone marrow stromal cells (BMSCs), known precursors of osteoblasts. Although they share a similar immunophenotype in vitro, functional studies showed that DPSCs produced only sporadic, but densely calcified nodules, and did not form adipocytes, whereas BMSCs routinely calcified throughout the adherent cell layer with clusters of lipid-laden adipocytes. When DPSCs were transplanted into immunocompromised mice, they generated a dentin-like structure lined with human odontoblast-like cells that surrounded a pulp-like interstitial tissue. In contrast, BMSCs formed lamellar bone containing osteocytes and surface-lining osteoblasts, surrounding a fibrous vascular tissue with active hematopoiesis and adipocytes. This study isolates postnatal human DPSCs that have the ability to form a dentin/pulp-like complex.
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            Author and article information

            Affiliations
            [1 ]Department of Dental Science for Health Promotion, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima 734-8553, Japan
            [2 ]Department of Dental and Medical Biochemistry, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima 734-8553, Japan
            [3 ]Department of Molecular Biology and Biochemistry, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima 734-8553, Japan
            [4 ]Department of Pediatric Dentistry, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima 734-8553, Japan
            [5 ]Department of Biochemistry and Molecular Biology, Nihon University School of Dentistry at Matsudo, Matsudo, Chiba 271-8587, Japan
            [6 ]Two Cells Co., Ltd., Hiroshima 734-0816, Japan
            [7 ]Natural Science Center for Basic Research and Development, Hiroshima University, Hiroshima 734-8553, Japan
            [8 ]Department of Dental Public Health, Faculty of Dental Medicine, Airlangga University, Surabaya, East Java 60132, Indonesia
            [9 ]Department of Conservative Dentistry, Faculty of Dental Medicine, Airlangga University, Surabaya, East Java 60132, Indonesia
            Author notes
            Correspondence to: Dr Katsumi Fujimoto, Department of Molecular Biology and Biochemistry, Graduate School of Biomedical and Health Sciences, Hiroshima University, 1-2-3 Kasumi, Minami-ku, Hiroshima 734-8553, Japan, E-mail: kfujimo@ 123456hiroshima-u.ac.jp
            Dr Yukio Kato, Department of Dental and Medical Biochemistry, Graduate School of Biomedical and Health Sciences, Hiroshima University, 1-2-3 Kasumi, Minami-ku, Hiroshima 734-8553, Japan, E-mail: ykato@ 123456hiroshima-u.ac.jp
            Journal
            Biomed Rep
            Biomed Rep
            BR
            Biomedical Reports
            D.A. Spandidos
            2049-9434
            2049-9442
            April 2018
            16 February 2018
            16 February 2018
            : 8
            : 4
            : 350-358
            5844140
            10.3892/br.2018.1066
            BR-0-0-1066
            Copyright: © Fujii et al.

            This is an open access article distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made.

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