CD44是存在於細胞膜表面的醣蛋白，主要負責細胞和細胞之間及細胞和基質之間的黏著功能，與細胞遷移，增殖，癌細胞的侵襲與轉移有關。根據報告指出，在人類牙齒發生(odontogenesis)的過程中，CD44在不同的發展階段會表現於不同的特定位置；在成人的牙齒裡，CD44表現在牙周韌帶的fibroblasts，cementocytes和牙髓中的造牙本質細胞(odontoblasts)裡；在牙胚期，表現在dental lamina和 presecretory ameloblasts；但是CD44是否在牙根未發育完全的牙齒內表現，或是否參與odontogenesis的過程目前並不清楚。因此本研究的目的是要探討CD44在未發育完全牙齒odontoblast裡的表現情形以及細胞礦物化過程中它所扮演的角色。首先，我們從門診收集拔下的第三大臼齒並經過脫鈣處理，利用免疫組織染色法(immunohistochemistry)來分析CD44在牙齒組織中分佈的情形，實驗結果顯示CD44蛋白主要表現在未發育完全的牙齒odontoblasts和根尖乳突(apical papillae)上。 我們分離這群細胞建立體外細胞培養系統，並且透過誘導細胞礦物化(in vitro mineralization)確認該群細胞硬組織生成的能力。為了進一步探討CD44參與調控礦物化的過程，我們利用人類慢病毒shRNA干擾系統(shRNA by the lentiviral system)抑制CD44的表現，結果發現經由siRNA來抑制CD44，會降低牙髓細胞之ALP activity的活性，同時降低產生礦物質小結的能力，表示牙髓細胞產生礦物質的能力受到干擾；此外在動物試驗方面，利用collagen membrane當做支架，將細胞植入SCID mice皮下，研究顯示抑制CD44也會使得牙髓細胞產生礦物質的能力降低；本實驗結果顯示CD44表現在造牙本質細胞，而且在礦物質生成能力上扮演調控的角色。

CD44 is a cell surface integral membrane glycoprotein, which plays important roles in cell–cell interactions, cell adhesion, and a variety of physiological and pathophysiological processes. The localization of CD44 during human odontogenesis has been investigated in previous studies. During odontogenesis, CD44 can be found in human dental structures at different developmental stages. In mature human teeth, it also appears that CD44 is localized in periodontal ligament fibroblasts, cementocytes and odontoblasts. But there is lacking connection in previous reports about the role of CD44 influencing the odontogenesis in teeth with immature roots. The aim of the study is to examine the expression of CD44 in the odontoblast cells and its role in the mineralization. In our study, extracted third molars were collected from dental clinics, decalcified and processed for immunolocalization of CD44. The data showed that the CD44 immunoreactivity was localized in odontoblasts in immature tooth. Whether CD44 involves in mineralization was assessed via small interfering RNA by using short hairpin RNA (by lentiviral system) and in vitro mineralization assay. Significantly reducing the alkaline phosphatase(ALP) activity and mineralized nodules formation after knockdown CD44 expression indicate a regulatory role of CD44 on mineralization of odontoblasts. Furthermore, using collagen membrane as scaffolds and implanted into SCID mice, odontoblasts with knockdown CD44 expression was repressed to produce mineralization.

Åberg, T., Wozney, J., Thesleff, I., 1997. Expression patterns of bone morphogenetic proteins (Bmps) in the developing mouse tooth suggest roles in morphogenesis and cell differentiation. Developmental Dynamics 210, 383-396.
Arany, S., Koyota, S., Sugiyama, T., 2009. Nerve growth factor promotes differentiation of odontoblast-like cells. Journal of Cellular Biochemistry 106, 539-545.
Aruffo, A., Stamenkovic, I., Melnick, M., Underhill, C.B., Seed, B., 1990. CD44 is the principal cell surface receptor for hyaluronate. Cell 61, 1303-1313.
Aubin, J.E., Heersche, J.N., Merrilees, M.J., Sodek, J., 1982. Isolation of bone cell clones with differences in growth, hormone responses, and extracellular matrix production. The Journal of Cell Biology 92, 452-461.
Banchs, F., Trope, M., 2004. Revascularization of Immature Permanent Teeth With Apical Periodontitis: New Treatment Protocol? Journal of Endodontics 30, 196-200.
Cao, J.J., Singleton, P.A., Majumdar, S., Boudignon, B., Burghardt, A., Kurimoto, P., Wronski, T.J., Bourguignon, L.Y.W., Halloran, B.P., 2005. Hyaluronan Increases RANKL Expression in Bone Marrow Stromal Cells Through CD44. Journal of Bone and Mineral Research 20, 30-40.
Chueh, L.-H., Ho, Y.-C., Kuo, T.-C., Lai, W.-H., Chen, Y.-H.M., Chiang, C.-P., 2009. Regenerative Endodontic Treatment for Necrotic Immature Permanent Teeth. Journal of Endodontics 35, 160-164.
Chueh, L.-H., Huang, G.T.J., 2006. Immature Teeth With Periradicular Periodontitis or Abscess Undergoing Apexogenesis: A Paradigm Shift. Journal of Endodontics 32, 1205-1213.
Cotti, E., Mereu, M., Lusso, D., 2008. Regenerative Treatment of an Immature, Traumatized Tooth With Apical Periodontitis: Report of a Case. Journal of Endodontics 34, 611-616.
Ding, R.Y., Cheung, G.S.-p., Chen, J., Yin, X.Z., Wang, Q.Q., Zhang, C.F., 2009. Pulp Revascularization of Immature Teeth With Apical Periodontitis: A Clinical Study. Journal of Endodontics 35, 745-749.
Fedarko, N.S., Fohr, B., Robey, P.G., Young, M.F., Fisher, L.W., 2000. Factor H Binding to Bone Sialoprotein and Osteopontin Enables Tumor Cell Evasion of Complement-mediated Attack. Journal of Biological Chemistry 275, 16666-16672.
Fedarko, N.S., Termine, J.D., Young, M.F., Robey, P.G., 1990. Temporal regulation of hyaluronan and proteoglycan metabolism by human bone cells in vitro. Journal of Biological Chemistry 265, 12200-12209.
Felszeghy, S., Hyttinen, M., Tammi, R., Tammi, M., Módis, L., 2000. Quantitative image analysis of hyaluronan expression in human tooth germs. European Journal of Oral Sciences 108, 311-317.
Felszeghy, S., Módis, L., Tammi, M., Tammi, R., 2001. The distribution pattern of the hyaluronan receptor CD44 during human tooth development. Archives of Oral Biology 46, 939-945.
Goodison, S., Urquidi, V., Tarin, D., 1999. CD44 cell adhesion molecules. Molecular Pathology 52, 189-196.
Grimme, H.U., Termeer, C.C., Bennett, K.L., Weiss, J.M., Schöpf, E., Aruffo, A., Simon, J.C., 1999. Colocalization of basic fibroblast growth factor and CD44 isoforms containing the variably spliced exon v3 (CD44v3) in normal skin and in epidermal skin cancers. British Journal of Dermatology 141, 824-832.
Gronthos, S., Brahim, J., Li, W., Fisher, L.W., Cherman, N., Boyde, A., DenBesten, P., Robey, P.G., Shi, S., 2002. Stem Cell Properties of Human Dental Pulp Stem Cells. Journal of Dental Research 81, 531-535.
Gronthos, S., Mankani, M., Brahim, J., Robey, P.G., Shi, S., 2000. Postnatal human dental pulp stem cells (DPSCs) in vitro and in vivo. Proceedings of the National Academy of Sciences 97, 13625-13630.
Horst, E., Meijer, C.J., Radaszkiewicz, T., Ossekoppele, G.J., Van Krieken, J.H., Pals, S.T., 1990. Adhesion molecules in the prognosis of diffuse large-cell lymphoma: expression of a lymphocyte homing receptor (CD44), LFA-1 (CD11a/18), and ICAM-1 (CD54). Leukemia 4, 595-599.
Hughes, D.E., Salter, D.M., Simpson, R., 1994. CD44 expression in human bone: A novel marker of osteocytic differentiation. Journal of Bone and Mineral Research 9, 39-44.
Inagaki, Y., Yoshida, K., Ohba, H., Seto, H., Kido, J.-i., Haneji, T., Nagata, T., 2010. High Glucose Levels Increase Osteopontin Production and Pathologic Calcification in Rat Dental Pulp Tissues. Journal of Endodontics 36, 1014-1020.
Inuyama, Y., Kitamura, C., Nishihara, T., Morotomi, T., Nagayoshi, M., Tabata, Y., Matsuo, K., Chen, K.-K., Terashita, M., 2010. Effects of hyaluronic acid sponge as a scaffold on odontoblastic cell line and amputated dental pulp. Journal of Biomedical Materials Research Part B: Applied Biomaterials 92B, 120-128.
Iohara, K., Nakashima, M., Ito, M., Ishikawa, M., Nakasima, A., Akamine, A., 2004. Dentin Regeneration by Dental Pulp Stem Cell Therapy with Recombinant Human Bone Morphogenetic Protein 2. Journal of Dental Research 83, 590-595.
Iwaya, S.-i., Ikawa, M., Kubota, M., 2001. Revascularization of an immature permanent tooth with apical periodontitis and sinus tract. Dental Traumatology 17, 185-187.
Jamal, H.H., Aubin, J.E., 1996. CD44 Expression in Fetal Rat Bone:In Vivoandin VitroAnalysis. Experimental Cell Research 223, 467-477.
Jernvall, J., Thesleff, I., 2000. Reiterative signaling and patterning during mammalian tooth morphogenesis. Mechanisms of Development 92, 19-29.
Jung, H.-S., Hosoya, A., Kim, J.-Y., Cho, S.-W., 2008a. The Role of BMP4 in HERS during Tooth Root Development. Journal of Oral Biosciences 50, 160-166.
Jung, I.-Y., Lee, S.-J., Hargreaves, K.M., 2008b. Biologically Based Treatment of Immature Permanent Teeth with Pulpal Necrosis: A Case Series. Journal of Endodontics 34, 876-887.
Kettunen, P., Karavanova, I., Thesleff, I., 1998. Responsiveness of developing dental tissues to fibroblast growth factors: expression of splicing alternatives of FGFR1, -2, -3, and of FGFR4; and stimulation of cell proliferation by FGF-2, -4, -8, and -9. Developmental Genetics 22, 374-385.
Knutson, J., Iida, J., Fields, G., McCarthy, J., 1996. CD44/chondroitin sulfate proteoglycan and alpha 2 beta 1 integrin mediate human melanoma cell migration on type IV collagen and invasion of basement membranes. Mol. Biol. Cell 7, 383-396.
Konig, H., Moll, J., Ponta, H., Herrlich, P., 1996. Trans-acting factors regulate the expression of CD44 splice variants. EMBO J 15, 4030-4039.
Kulseth, M.A., Kolset, S.O., Ranheim, T., 1999. Stimulation of serglycin and CD44 mRNA expression in endothelial cells exposed to TNF-[alpha] and IL-1[alpha]. Biochimica et Biophysica Acta (BBA) - General Subjects 1428, 225-232.
Kuppner, M.C., Van Meir, E., Gauthier, T., Hamou, M.F., de Tribolet, N., 1992. Differential expression of the CD44 molecule in human brain tumours. Int J Cancer 50, 572-577.
Laino, G., Graziano, A., d'Aquino, R., Pirozzi, G., Lanza, V., Valiante, S., De Rosa, A., Naro, F., Vivarelli, E., Papaccio, G., 2006. An approachable human adult stem cell source for hard-tissue engineering. Journal of Cellular Physiology 206, 693-701.
Leonardi, R., Loreto, C., Caltabiano, R., Caltabiano, C., 2006. Immunolocalization of CD44s in human teeth. Acta Histochemica 108, 425-429.
Lesley, J., Hyman, R., Kincade, P.W., 1993. CD44 and Its Interaction with Extracellular Matrix. In: Frank, J.D. (Ed.), Advances in Immunology. Academic Press, pp. 271-335.
Linde, A., Goldberg, M., 1993. Dentinogenesis. Critical Reviews in Oral Biology & Medicine 4, 679-728.
Matthiessen, M.E., Garbarsch, C., Engelbrecht Olsen, B., Hellström, S., Engström-Laurent, A., 1997. Hyaluronan in Human Deciduous Tooth Germs in the Bell Stage. Cells Tissues Organs 159, 1-7.
McKee, M.D., Zalzal, S., Nanci, A., 1996. Extracellular matrix in tooth cementum and mantle dentin: Localization of osteopontin and other noncollagenous proteins, plasma proteins, and glycoconjugates by electron microscopy. The Anatomical Record 245, 293-312.
Miyake, K., Underhill, C.B., Lesley, J., Kincade, P.W., 1990. Hyaluronate can function as a cell adhesion molecule and CD44 participates in hyaluronate recognition. The Journal of Experimental Medicine 172, 69-75.
Nakashima, M., 1994. Induction of Dentin Formation on Canine Amputated Pulp by Recombinant Human Bone Morphogenetic Proteins (BMP)-2 and -4. Journal of Dental Research 73, 1515-1522.
Nakashima, M., Reddi, A.H., 2003. The application of bone morphogenetic proteins to dental tissue engineering. Nat Biotech 21, 1025-1032.
Nakashima, M., Tachibana, K., Iohara, K., Ito, M., Ishikawa, M., Akamine, A., 2003. Induction of Reparative Dentin Formation by Ultrasound-Mediated Gene Delivery of Growth/Differentiation Factor 11. Human Gene Therapy 14, 591-597.
Nakashima, M.I., Koichiro; Zheng, Li, 2006. Gene Therapy for Dentin Regeneration with Bone Morphogenetic Proteins. Current Gene Therapy 6, 551-560.
Narayanan, K., Srinivas, R., Ramachandran, A., Hao, J., Quinn, B., George, A., 2001. Differentiation of embryonic mesenchymal cells to odontoblast-like cells by overexpression of dentin matrix protein 1. Proceedings of the National Academy of Sciences 98, 4516-4521.
Nie, X., Luukko, K., Kettunen, P., 2006. BMP signalling in craniofacial development. Int J Dev Biol 50, 511-521.
Perschl, A., Lesley, J., English, N., Trowbridge, I., Hyman, R., 1995. Role of CD44 cytoplasmic domain in hyaluronan binding. European Journal of Immunology 25, 495-501.
Peters, H., Balling, R., 1999. Teeth: where and how to make them. Trends in Genetics 15, 59-65.
Petrino, J.A., Boda, K.K., Shambarger, S., Bowles, W.R., McClanahan, S.B., 2010. Challenges in Regenerative Endodontics: A Case Series. Journal of Endodontics 36, 536-541.
Pittenger, M.F., Mackay, A.M., Beck, S.C., Jaiswal, R.K., Douglas, R., Mosca, J.D., Moorman, M.A., Simonetti, D.W., Craig, S., Marshak, D.R., 1999. Multilineage Potential of Adult Human Mesenchymal Stem Cells. Science 284, 143-147.
Schmitz, B., Park, I.A., Kaufmann, R., Thiele, J., Fischer, R., 1995. Influence of Cytokine Stimulation (Granulocyte Macrophage-Colony Stimulating Factor, lnterleukin-3 and Transforming Growth Factor-Beta-1) on Adhesion Molecule Expression in Normal Human Bone Marrow Fibroblasts. Acta Haematologica 94, 173-181.
Screaton, G.R., Bell, M.V., Bell, J.I., Jackson, D.G., 1993. The identification of a new alternative exon with highly restricted tissue expression in transcripts encoding the mouse Pgp-1 (CD44) homing receptor. Comparison of all 10 variable exons between mouse, human, and rat. Journal of Biological Chemistry 268, 12235-12238.
Screaton, G.R., Bell, M.V., Jackson, D.G., Cornelis, F.B., Gerth, U., Bell, J.I., 1992. Genomic structure of DNA encoding the lymphocyte homing receptor CD44 reveals at least 12 alternatively spliced exons. Proceedings of the National Academy of Sciences 89, 12160-12164.
Shah, N., Logani, A., Bhaskar, U., Aggarwal, V., 2008. Efficacy of Revascularization to Induce Apexification/Apexogensis in Infected, Nonvital, Immature Teeth: A Pilot Clinical Study. Journal of Endodontics 34, 919-925.
Smith, A.J., Lesot, H., 2001. Induction and Regulation of Crown Dentinogenesis: Embryonic Events as a Template for Dental Tissue Repair? Critical Reviews in Oral Biology & Medicine 12, 425-437.
Sodek, J., Ganss, B., McKee, M.D., 2000. Osteopontin. Critical Reviews in Oral Biology & Medicine 11, 279-303.
Sonoyama, W., Liu, Y., Yamaza, T., Tuan, R.S., Wang, S., Shi, S., Huang, G.T.J., 2008. Characterization of the Apical Papilla and Its Residing Stem Cells from Human Immature Permanent Teeth: A Pilot Study. Journal of Endodontics 34, 166-171.
Stamenkovic, I., Amiot, M., Pesando, J.M., Seed, B., 1989. A lymphocyte molecule implicated in lymph node homing is a member of the cartilage link protein family. Cell 56, 1057-1062.
Tammi, R., MacCallum, D., Hascall, V.C., Pienimäki, J.-P., Hyttinen, M., Tammi, M., 1998. Hyaluronan Bound to CD44 on Keratinocytes Is Displaced by Hyaluronan Decasaccharides and Not Hexasaccharides. Journal of Biological Chemistry 273, 28878-28888.
Tanikawa, R., Tanikawa, T., Hirashima, M., Yamauchi, A., Tanaka, Y., 2010. Galectin-9 induces osteoblast differentiation through the CD44/Smad signaling pathway. Biochemical and Biophysical Research Communications 394, 317-322.
Thesleff, I., 2003. Epithelial-mesenchymal signalling regulating tooth morphogenesis. Journal of Cell Science 116, 1647-1648.
Thesleff, I., Barrach, H.J., Foidart, J.M., Vaheri, A., Pratt, R.M., Martin, G.R., 1981. Changes in the distribution of type IV collagen, laminin, proteoglycan, and fibronectin during mouse tooth development. Developmental Biology 81, 182-192.
Thesleff, I., Sharpe, P., 1997. Signalling networks regulating dental development. Mechanisms of Development 67, 111-123.
Thesleff, I., Vaahtokari, A., Partanen, A.M., 1995. Regulation of organogenesis. Common molecular mechanisms regulating the development of teeth and other organs. Int J Dev Biol 39, 35-50.
Toole, B.P., 2001. Hyaluronan in morphogenesis. Seminars in Cell & Developmental Biology 12, 79-87.
Tucker, A., Sharpe, P., 2004. The cutting-edge of mammalian development; how the embryo makes teeth. Nat Rev Genet 5, 499-508.
Tucker, A.S., Al Khamis, A., Ferguson, C.A., Bach, I., Rosenfeld, M.G., Sharpe, P.T., 1999. Conserved regulation of mesenchymal gene expression by Fgf-8 in face and limb development. Development 126, 221-228.
Tziafas, D., Kodonas, K., 2010. Differentiation Potential of Dental Papilla, Dental Pulp, and Apical Papilla Progenitor Cells. Journal of Endodontics 36, 781-789.
Vainio, S., Karavanova, I., Jowett, A., Thesleff, I., 1993. Identification of BMP-4 as a signal mediating secondary induction between epithelial and mesenchymal tissues during early tooth development. Cell 75, 45-58.
Wheatley, S.C., Isacke, C.M., Crossley, P.H., 1993. Restricted expression of the hyaluronan receptor, CD44, during postimplantation mouse embryogenesis suggests key roles in tissue formation and patterning. Development 119, 295-306.
Wong, G.L., 1984. Paracrine interactions in bone-secreted products of osteoblasts permit osteoclasts to respond to parathyroid hormone. Journal of Biological Chemistry 259, 4019-4022.
Yamada, K., Yamada, T., Sasaki, T., Rahemtulla, F., Takagi, M., 1997. Light and electron microscopical immunohistochemical localization of large proteoglycans in human tooth germs at the bell stage. The Histochemical Journal 29, 167-175.
Yamashiro, T., Tummers, M., Thesleff, I., 2003. Expression of Bone Morphogenetic Proteins and Msx Genes during Root Formation. Journal of Dental Research 82, 172-176.
Yoshiba, N., Yoshiba, K., Aberdam, D., Meneguzzi, G., Perrin-Schmitt, F., Stoetzel, C., Ruch, J.V., Lesot, H., 1998. Expression and localization of laminin-5 subunits in the mouse incisor. Cell and Tissue Research 292, 143-149.
Yu, Q., Toole, B.P., 1997. Common pattern of CD44 isoforms is expressed in morphogenetically active epithelia. Developmental Dynamics 208, 1-10.
Zhang, Y., Zhang, Z., Zhao, X., Yu, X., Hu, Y., Geronimo, B., Fromm, S.H., Chen, Y.P., 2000. A new function of BMP4: dual role for BMP4 in regulation of Sonic hedgehog expression in the mouse tooth germ. Development 127, 1431-1443.
Zohar, R., Cheifetz, S., McCulloch, C.A., Sodek, J., 1998. Analysis of intracellular osteopontin as a marker of osteoblastic cell differentiation and mesenchymal cell migration. Eur J Oral Sci 106 Suppl 1, 401-407.