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1.
PLoS One ; 8(9): e73532, 2013.
Article in English | MEDLINE | ID: mdl-24086284

ABSTRACT

Using quantitative PCR-based miRNA arrays, we comprehensively analyzed the expression profiles of miRNAs in human and mouse embryonic stem (ES), induced pluripotent stem (iPS), and somatic cells. Immature pluripotent cells were purified using SSEA-1 or SSEA-4 and were used for miRNA profiling. Hierarchical clustering and consensus clustering by nonnegative matrix factorization showed two major clusters, human ES/iPS cells and other cell groups, as previously reported. Principal components analysis (PCA) to identify miRNAs that segregate in these two groups identified miR-187, 299-3p, 499-5p, 628-5p, and 888 as new miRNAs that specifically characterize human ES/iPS cells. Detailed direct comparisons of miRNA expression levels in human ES and iPS cells showed that several miRNAs included in the chromosome 19 miRNA cluster were more strongly expressed in iPS cells than in ES cells. Similar analysis was conducted with mouse ES/iPS cells and somatic cells, and several miRNAs that had not been reported to be expressed in mouse ES/iPS cells were suggested to be ES/iPS cell-specific miRNAs by PCA. Comparison of the average expression levels of miRNAs in ES/iPS cells in humans and mice showed quite similar expression patterns of human/mouse miRNAs. However, several mouse- or human-specific miRNAs are ranked as high expressers. Time course tracing of miRNA levels during embryoid body formation revealed drastic and different patterns of changes in their levels. In summary, our miRNA expression profiling encompassing human and mouse ES and iPS cells gave various perspectives in understanding the miRNA core regulatory networks regulating pluripotent cells characteristics.


Subject(s)
Embryonic Stem Cells/metabolism , Gene Expression Profiling , Induced Pluripotent Stem Cells/metabolism , MicroRNAs/genetics , Animals , Cell Line , Cluster Analysis , Humans , Mice , Polymerase Chain Reaction , Principal Component Analysis
2.
PLoS One ; 8(7): e67739, 2013.
Article in English | MEDLINE | ID: mdl-23861797

ABSTRACT

BACKGROUND: The purpose of this study was to determine whether autologous mesenchymal stem cells (MSCs) implantation improves endothelial dysfunction in a rabbit ischemic limb model. METHODS: We evaluated the effect of MSC implantation on limb blood flow (LBF) responses to acetylcholine (ACh), an endothelium-dependent vasodilator, and sodium nitroprusside (SNP), an endothelium-independent vasodilator, in rabbits with limb ischemia in which cultured MSCs were implanted (n = 20) or saline was injected as a control group (n = 20). LBF was measured using an electromagnetic flowmeter. A total of 10(6) MSCs were implanted into each ischemic limb. RESULTS: Histological sections of ischemic muscle showed that capillary index (capillary/muscle fiber) was greater in the MSC implantation group than in the control group. Laser Doppler blood perfusion index was significantly increased in the MSC implantation group compared with that in the control group. LBF response to ACh was greater in the MSC group than in the control group. After administration of N(G)-nitro-L-arginine, a nitric oxide synthase inhibitor, LBF response to ACh was similar in the MSC implantation group and control group. Vasodilatory effects of SNP in the two groups were similar. CONCLUSIONS: These findings suggest that MSC implantation induces angiogenesis and augments endothelium-dependent vasodilation in a rabbit ischemic model through an increase in nitric oxide production.


Subject(s)
Hindlimb/blood supply , Ischemia/therapy , Mesenchymal Stem Cell Transplantation , Recovery of Function/physiology , Acetylcholine/pharmacology , Animals , Bone Marrow Cells/cytology , Bone Marrow Cells/physiology , Hemodynamics/drug effects , Hindlimb/physiopathology , Ischemia/physiopathology , Male , Mesenchymal Stem Cells/cytology , Mesenchymal Stem Cells/physiology , Neovascularization, Physiologic/drug effects , Neovascularization, Physiologic/physiology , Nitroarginine/pharmacology , Nitroprusside/pharmacology , Rabbits , Transplantation, Autologous , Vasodilation/drug effects , Vasodilation/physiology , Vasodilator Agents/pharmacology
3.
J Mater Sci Mater Med ; 22(12): 2765-72, 2011 Dec.
Article in English | MEDLINE | ID: mdl-21901371

ABSTRACT

JH8194 induces osteoblast differentiation, although it was originally designed to improve antifungal activity. This suggests that JH8194 is useful for implant treatment. Therefore, the aim of this study was to evaluate the osseointegration capacity of JH8194-modified titanium dental implant fixtures (JH8194-Fi). The implants were randomly implanted into the edentulous ridge of dog mandibles. Healing abutments were inserted immediately after implant placement. Three weeks later, peri-implant bone levels, the first bone-to-implant contact points, and trabecular bone formation surrounding the implants were assessed by histological and digital image analyses based on microcomputed tomography (microCT). The histological analysis revealed an enhancement of mature trabecular bone around the JH8194-Fi compared with untreated fixtures (control-Fi). Similarly, microCT combined with analysis by Zed View™ also showed increased trabecular bone formation surrounding the JH8194-Fi compared with the control-Fi (Student's t-test, P < 0.05). JH8194 may offer an alternative biological modification of titanium surfaces to enhance trabecular bone formation around dental implants, which may contribute to the transient acquirement of osseointegration and the long-term success of implant therapy.


Subject(s)
Antimicrobial Cationic Peptides/administration & dosage , Bone and Bones/physiology , Dental Implants , Histatins/administration & dosage , Titanium/chemistry , Animals , Bone and Bones/pathology , Coated Materials, Biocompatible , Dogs , Histatins/chemistry , Mandible/pathology , Osseointegration , Osteoblasts/cytology , Prostheses and Implants , Surface Properties , Tomography, X-Ray Computed , X-Ray Microtomography
4.
Clin Vaccine Immunol ; 18(5): 860-6, 2011 May.
Article in English | MEDLINE | ID: mdl-21346056

ABSTRACT

Since its emergence, the 2009 pandemic H1N1 virus has spread rapidly throughout the world. Previously, we reported that most individuals born after 1920 do not have cross-reactive virus-neutralizing antibodies against pandemic (H1N1) 2009 virus, indicating that they were immunologically naïve to the pandemic virus prior to its emergence. This finding provided us with an excellent opportunity for a seroepidemiological investigation of the transmission mode of the pandemic virus in the community. To gain insight into its transmission within communities, we performed a serosurvey for pandemic virus infection with schoolchildren at an elementary school in Tokyo, Japan, and their parents. We observed a high prevalence of neutralizing antibodies to the pandemic virus in the children at this school, although the percentage of children positive for the neutralizing antibodies varied among classrooms. While a much lower prevalence was observed among parents, seropositivity of the parents correlated with that of their schoolchildren. Moreover, many adults appeared to have experienced asymptomatic infection with the pandemic virus. These data suggest that the pandemic virus was readily transmitted among schoolchildren in elementary schools and that it was also transmitted from schoolchildren to their parents.


Subject(s)
Antibodies, Viral/blood , Influenza A Virus, H1N1 Subtype/immunology , Influenza, Human/epidemiology , Influenza, Human/virology , Adult , Antibodies, Neutralizing/blood , Asymptomatic Diseases , Child , Disease Transmission, Infectious , Family Health , Female , Humans , Influenza, Human/transmission , Male , Middle Aged , Parents , Seroepidemiologic Studies , Students , Tokyo/epidemiology
5.
Int J Mol Sci ; 11(4): 1458-1470, 2010 Apr 02.
Article in English | MEDLINE | ID: mdl-20480030

ABSTRACT

The objective of this study was to evaluate the effect of titanium immobilized with a cationic antimicrobial peptide (JH8194) derived from histatin on the biofilm formation of Porphyromonas gingivalis and differentiation of osteoblastic cells (MC3T3-E1). The titanium specimens (Ti) were immobilized with JH8194, according to the method previously described. The colonization of P. gingivalis on JH8194-Ti was significantly lower than that on control- and blocking-Ti. JH8194-Ti enhanced the mRNA expressions of Runx2 and OPN, and ALPase activity in the MC3T3-E1, as compared with those of control- and blocking-Ti. These results, taken together, suggested the possibility that JH8194-Ti may be a potential aid to shorten the period of acquiring osseointegration.


Subject(s)
Antimicrobial Cationic Peptides/pharmacology , Cell Differentiation/drug effects , Histatins/chemistry , Titanium/chemistry , Alkaline Phosphatase/genetics , Alkaline Phosphatase/metabolism , Animals , Antimicrobial Cationic Peptides/chemistry , Biofilms/drug effects , Cell Line , Cell Proliferation/drug effects , Core Binding Factor Alpha 1 Subunit/genetics , Core Binding Factor Alpha 1 Subunit/metabolism , Histatins/metabolism , Immobilized Proteins/chemistry , Immobilized Proteins/pharmacology , Mice , Osteoblasts/drug effects , Osteoblasts/metabolism , Osteopontin/genetics , Osteopontin/metabolism , Porphyromonas gingivalis/physiology , RNA, Messenger/metabolism
6.
Biochem Biophys Res Commun ; 396(2): 193-8, 2010 May 28.
Article in English | MEDLINE | ID: mdl-20362549

ABSTRACT

The Philadelphia chromosome-positive blastoma, maintained by serial subcutaneous transplantation in nude mice, is a highly proliferating biological mass consisting of homogenous CD34(+)CD38(-) myeloblastoid cells. These cells newly evolved from pluripotent leukemia stem cells of chronic myeloid leukemia in the chronic phase. Therefore, this mass may provide a unique tool for better understanding cellular and molecular mechanisms of self-renewal of leukemia stem cells. In this paper, we demonstrated that intravenously injected blastoma cells can cause Ph+ blastic leukemia with multiple invasive foci in NOD/SCID mice but not in nude mice. In addition, using an in vitro culture system, we clearly showed that blastoma cell adhesion to OP9 stromal cells accelerates blastoma cell proliferation that is associated with up-regulation of BMI1 gene expression; increased levels of beta-catenin and the Notch1 intra-cellular domain; and changed the expression pattern of variant CD44 forms, which are constitutively expressed in these blastoma cells. These findings strongly suggest that adhesion of leukemic stem cells to stromal cells via CD44 might be indispensable for their cellular defense against attack by immune cells and for maintenance of their self-renewal ability.


Subject(s)
Neoplastic Stem Cells/pathology , Pluripotent Stem Cells/pathology , Precursor Cell Lymphoblastic Leukemia-Lymphoma/pathology , Animals , Cell Adhesion , Cell Culture Techniques , Gene Expression Regulation, Leukemic , Humans , Hyaluronan Receptors/genetics , Hyaluronan Receptors/metabolism , Mice , Mice, Inbred NOD , Mice, SCID , Neoplasm Transplantation , Nuclear Proteins/genetics , Polycomb Repressive Complex 1 , Precursor Cell Lymphoblastic Leukemia-Lymphoma/genetics , Proto-Oncogene Proteins/genetics , Receptor, Notch1/metabolism , Repressor Proteins/genetics , beta Catenin/metabolism
7.
Yakugaku Zasshi ; 129(3): 381-4, 2009 Mar.
Article in Japanese | MEDLINE | ID: mdl-19252397

ABSTRACT

To apply human mesenchymal stem cells (hMSC) to regenerative medicines, it is necessary to multiply hMSC in vitro in a short period. In addition, it is desirable that the medium which is used for the hMSC multiplication is not supplemented with the serum, because the addition of the serum has risks of infection. In this study, we cultured hMSC with three kinds of medium used for multiplying hMSC (DMEM, MSCGM, STK2) and compared hMSC proliferation in each medium. As a result, it was confirmed that hMSC proliferation was significantly higher in STK2 medium which is a novel serum-free medium developed for hMSC multiplication. Moreover, we compared the hMSC proliferation in these media under the environment that assumed bone reproduction. When we cultured hMSC in each medium with hydroxyapatite (HAp), the proliferative inhibition by HAp depended on the additive amount, and the degree of the proliferative inhibition was different among the media but the lowest inhibitory effect was observed in STK2 medium.


Subject(s)
Cell Proliferation/drug effects , Culture Media, Serum-Free/pharmacology , Durapatite/pharmacology , Mesenchymal Stem Cells/cytology , Cells, Cultured , Depression, Chemical , Dose-Response Relationship, Drug , Humans , Stimulation, Chemical
8.
Genes Cells ; 14(3): 407-24, 2009 Mar.
Article in English | MEDLINE | ID: mdl-19228201

ABSTRACT

Although ex vivo expanded mesenchymal stem cells (MSC) have been used in numerous studies, the molecular signature and in vivo distribution status of MSC remain unknown. To address this matter, we identified numerous human MSC-characteristic genes--including nine transcription factor genes--using DNA microarray and real-time RT-PCR analyses: Most of the MSC-characteristic genes were down-regulated 24 h after incubation with osteogenesis-, chondrogenesis- or adipogenesis-induction medium, or 48-72 h after knockdown of the nine transcription factors. Furthermore, knockdowns of ETV1, ETV5, FOXP1, GATA6, HMGA2, SIM2 or SOX11 suppressed the self-renewal capacity of MSC, whereas those of FOXP1, SOX11, ETV1, SIM2 or PRDM16 reduced the osteogenic- and/or adipogenic potential. In addition, immunohistochemistry using antibodies for the MSC characteristic molecules--including GATA6, TRPC4, FLG and TGM2--revealed that MSC-like cells were present near the endosteum and in the interior of bone marrow of adult mice. These findings indicate that MSC synthesize a set of MSC markers in vitro and in vivo, and that MSC-characteristic transcription factors are involved in MSC stemness regulation.


Subject(s)
Gene Expression , Mesenchymal Stem Cells/metabolism , Transcription Factors/genetics , Fibroblasts/cytology , Filaggrin Proteins , Gene Knockdown Techniques , Humans , Oligonucleotide Array Sequence Analysis , Skin/cytology
9.
J Biol Chem ; 283(23): 16259-67, 2008 Jun 06.
Article in English | MEDLINE | ID: mdl-18390540

ABSTRACT

Brain-derived neurotrophic factor (BDNF), recognized as essential in the developing nervous system, is involved in differentiation and proliferation in non-neuronal cells, such as endothelial cells, osteoblasts, and periodontal ligament cells. We have focused on the application of BDNF to the regeneration of periodontal tissue and indicated that BDNF promotes the regeneration of experimentally created periodontal defects. Cementoblasts form cementum, mineralized tissue, which is key to establishing a functional periodontium. The application of BDNF to the regeneration of periodontal tissue requires elucidation of the mechanism by which BDNF regulates the functions of cementoblasts. In this study, we examined how BDNF regulates the mRNA expression of bone/cementum-related proteins (alkaline phosphatase (ALP), osteopontin (OPN), and bone morphogenetic protein-2 (BMP-2)) in cultures of immortalized human cementoblast-like (HCEM) cells. BDNF elevated the mRNA levels of ALP, OPN, and BMP-2 in HCEM cells. Small interfering RNA (siRNA) for TRKB, a high affinity receptor of BDNF, siRNA for ELK-1, which is a downstream target of ERK1/2, and PD98059, an ERK inhibitor, obviated the increase in the mRNA levels. BDNF increased the levels of phosphorylated ERK1/2 and Elk-1, and the blocking of BDNF signaling by treatment with siRNA for TRKB and PD98059 suppressed the phosphorylation of ERK1/2 and Elk-1. Furthermore, BDNF increased the levels of phosphorylated c-Raf, which activates the ERK signaling pathway. These findings provide the first evidence that the TrkB-c-Raf-ERK1/2-Elk-1 signaling pathway is required for the BDNF-induced mRNA expression of ALP, OPN, and BMP-2 in HCEM cells.


Subject(s)
Brain-Derived Neurotrophic Factor/pharmacology , Dental Cementum/metabolism , Gene Expression Regulation/drug effects , Periodontal Ligament/physiology , Regeneration/drug effects , Bone and Bones/cytology , Bone and Bones/metabolism , Brain-Derived Neurotrophic Factor/metabolism , Cell Differentiation/drug effects , Cell Differentiation/physiology , Cell Line, Transformed , Cell Proliferation/drug effects , Dental Cementum/cytology , Gene Expression Regulation/physiology , Humans , Nervous System/embryology , Organ Specificity/drug effects , Organ Specificity/physiology , Periodontal Ligament/cytology , RNA, Messenger/biosynthesis , Regeneration/physiology
10.
Tissue Eng ; 13(10): 2405-17, 2007 Oct.
Article in English | MEDLINE | ID: mdl-17596118

ABSTRACT

Bone marrow stromal cells (BMSCs) are valuable in tissue engineering and cell therapy, but the quality of the cells is critical for the efficacy of therapy. To test the quality and identity of transplantable cells, we identified the molecular markers that were expressed at higher levels in BMSCs than in fibroblasts. Using numerous BMSC lines from tibia, femur, ilium, and jaw, together with skin and gum fibroblasts, we compared the gene expression profiles of these cells using DNA microarrays and low-density array cards. The differentiation potential of tibia and femur BMSCs was similar to that of iliac BMSCs, and different from jaw BMSCs, but all BMSC lines had many common markers that were expressed at much higher levels in BMSCs than in fibroblasts; several BMSC markers showed discrete expression patterns between jaw and other BMSCs. The common markers are probably useful in routine tests, but their efficacy may depend upon the passage number or donor age. In our study the passage number markedly altered the expression levels of several markers, while donor age had little effect on them. Considering the effects of in vivo location of BMSCs and passage, magnitude of increase in expression levels, and interindividual differences, we identified several reliable markers -- LIF, IGF1, PRG1, MGP, BMP4, CTGF, KCTD12, IGFBP7, TRIB2, and DYNC1I1 -- among many candidates. This marker set may be useful in a routine test for BMSCs in tissue engineering and cell therapy.


Subject(s)
Aging/metabolism , Bone Marrow Cells/cytology , Bone Marrow Cells/metabolism , Mesenchymal Stem Cells/cytology , Mesenchymal Stem Cells/metabolism , Stromal Cells/cytology , Stromal Cells/metabolism , Adult , Biomarkers/metabolism , Cell Differentiation , Cells, Cultured , Female , Gene Expression Profiling/methods , Humans , Male , Middle Aged , Reverse Transcriptase Polymerase Chain Reaction/methods , Tissue Donors
11.
Br J Haematol ; 134(2): 208-12, 2006 Jul.
Article in English | MEDLINE | ID: mdl-16846479

ABSTRACT

A pilot study was undertaken using a fludarabine-based conditioning regimen to improve haematopoietic cell transplantation (HCT) from alternative donors in 27 Fanconi anaemia (FA) patients. Patients were conditioned with 150-180 mg/m2 of fludarabine, 40 mg/kg of cyclophosphamide, 5-10 mg/kg of antithymocyte globulin, and 300-450 cGy of thoracoabdominal/total body irradiation. One patient who received unrelated cord blood transplantation failed to engraft, another patient died of sepsis. The 1-year overall survival was 96.3% (95% CI, 89-100). This conditioning regimen exerted an immunosuppressive effect that enabled durable engraftment in alternative donor HCT without severe toxicity.


Subject(s)
Fanconi Anemia/therapy , Hematopoietic Stem Cell Transplantation/methods , Transplantation Conditioning/methods , Child , Child, Preschool , Combined Modality Therapy , Cyclophosphamide/therapeutic use , Drug Therapy, Combination , Fanconi Anemia/radiotherapy , Female , Graft vs Host Disease/etiology , Humans , Infant , Male , Pilot Projects , Treatment Outcome , Vidarabine/analogs & derivatives , Vidarabine/therapeutic use , Whole-Body Irradiation
12.
J Periodontol ; 77(6): 1003-7, 2006 Jun.
Article in English | MEDLINE | ID: mdl-16734575

ABSTRACT

BACKGROUND: Recently, there have been an increased number of basic and clinical reports indicating the superior potential of bone marrow-derived mesenchymal stem cells (MSCs) for tissue regeneration. In periodontal treatment, previous animal studies indicated that autotransplantation of bone marrow MSCs into experimental periodontal defects enhanced periodontal tissue regeneration. However, mechanisms for periodontal tissue regeneration with MSCs are still unclear. The purpose of this study was to elucidate the behavior of transplanted MSCs in periodontal defects. METHODS: Bone marrow MSCs were isolated from beagle dogs, labeled with green fluorescent protein (GFP), and expanded in vitro. The expanded MSCs were mixed with atelocollagen (2% type I collagen) at final concentrations of 2 x 10(7) cells/ml and transplanted into experimental Class III periodontal defects. Localizations of GFP and proliferating cell nuclear antigen (PCNA)-positive cells were evaluated by immunohistochemical analysis. RESULTS: Four weeks after transplantation, the periodontal defects were almost regenerated with periodontal tissue. Cementoblasts, osteoblasts, osteocytes, and fibroblasts of the regenerated periodontal tissue were positive with GFP. PCNA-positive cells were present in regenerating connective tissue. CONCLUSION: These findings suggest that transplanted mesenchymal stem cells could survive and differentiate into periodontal tissue cells, resulting in enhancement of periodontal tissue regeneration.


Subject(s)
Alveolar Process/surgery , Mesenchymal Stem Cell Transplantation , Mesenchymal Stem Cells/physiology , Animals , Bone Marrow Cells/cytology , Bone Regeneration/physiology , Cattle , Cell Differentiation/physiology , Dogs , Female , Green Fluorescent Proteins , Proliferating Cell Nuclear Antigen
13.
J Periodontol ; 77(5): 800-7, 2006 May.
Article in English | MEDLINE | ID: mdl-16671871

ABSTRACT

BACKGROUND: We have previously shown that cultured human periodontal ligament (HPL) cells produce nerve growth factor (NGF) and express mRNA of tyrosine kinase receptor (trkA), a high-affinity receptor of NGF. These findings suggest that NGF modulates the differentiation and proliferation of the periodontal ligament cells by paracrine and autocrine functions in vivo. Endothelial cells also express NGF and trkA. Therefore, NGF may regulate functions of periodontal ligament cells and endothelial cells during periodontal tissue regeneration. METHODS: Effects of NGF on expressions of bone/cementum-related proteins (osteocalcin [OC], bone sialoprotein [BSP], bone morphogenetic protein [BMP-7], core binding factor alpha [Cbfa-1], and type I collagen), calcification in HPL cells, and proliferation and mRNA expression of vascular endothelial growth factor (VEGF), an endothelial cell mitogen, in human microvascular endothelial cells (HMVECs) were examined. RESULTS: NGF elevated mRNA levels of OC, BSP, BMP-7, Cbfa-1, and type I collagen and enhanced mineral deposition in cultures of HPL cells. Furthermore, NGF stimulated mRNA expressions of VEGF-A and VEGF-B and cell proliferation in HMVEC. CONCLUSION: These findings suggest that the functional regulation of periodontal ligament cells and endothelial cells by NGF might result in the acceleration of periodontal tissue regeneration in vivo.


Subject(s)
Endothelial Cells/drug effects , Nerve Growth Factor/pharmacology , Periodontal Ligament/cytology , Bone Morphogenetic Proteins/metabolism , Cell Culture Techniques , Cell Proliferation/drug effects , Collagen Type I/metabolism , Core Binding Factor Alpha 1 Subunit/metabolism , Dose-Response Relationship, Drug , Endothelial Cells/metabolism , Humans , Integrin-Binding Sialoprotein , Osteocalcin/metabolism , RNA, Messenger/drug effects , RNA, Messenger/metabolism , Sialoglycoproteins/metabolism , Time Factors
14.
Oncology ; 69(3): 246-55, 2005.
Article in English | MEDLINE | ID: mdl-16127291

ABSTRACT

OBJECTIVES: The receptor for advanced glycation end products (RAGE) is a newly recognized factor regulating cancer cell invasion and metastasis. Nevertheless, the involvement of RAGE in the development and progression of oral squamous cell carcinomas has not been elucidated. This study investigated the expression of RAGE in ten oral squamous cell carcinoma cell lines including primary and metastatic cell lines and its association with invasion and metastasis. METHODS: Reverse transcriptase-polymerase chain reaction, antisense phosphorothioate (S)-oligodeoxynucleotide assay, preparation of antibody, immunohistochemical staining, immunoblot analysis, migration assay, in vitro invasion assay, and wound-healing assay were used. RESULTS: RAGE protein expression of metastatic cancer cells treated with RAGE antisense S-oligodeoxynucleotide was significantly reduced compared to that of sense S-oligodeoxynucleotide-treated cells. The migration assay showed that invasive activity was significantly reduced in metastatic cancer cells treated with RAGE antisense S-oligodeoxynucleotide. Similarly, during invasion assays, numbers of invading cells were also reduced with the addition of RAGE antisense S-oligodeoxynucleotide-treated cells. A wound-healing assay showed that only a few RAGE antisense S-oligodeoxynucleotide-treated cancer cells migrated into the scraped area, whereas sense S-oligodeoxynucleotide-treated cells showed many budding nests in the scraped area of the metastatic cell lines. Immunohistochemically, oral squamous cell carcinoma cells in the tumour mesenchymal border were often immunopositive, whereas basal cells in the normal mucosa were scarcely positive. CONCLUSIONS: These results suggest that RAGE expression appears to be closely associated with the invasiveness of oral squamous cell carcinoma and represents a promising candidate for assessing the future therapeutic potential in treating patients with oral carcinoma.


Subject(s)
Carcinoma, Squamous Cell/metabolism , Carcinoma, Squamous Cell/pathology , Gene Expression Regulation, Neoplastic , Glycation End Products, Advanced/metabolism , Mouth Neoplasms/metabolism , Mouth Neoplasms/pathology , Blotting, Western , Cadherins/metabolism , Carcinoma, Squamous Cell/genetics , Cell Line, Tumor , Cell Movement , Humans , Matrix Metalloproteinase 9/metabolism , Mouth Neoplasms/genetics , Neoplasm Invasiveness , Oligodeoxyribonucleotides, Antisense/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Wound Healing
15.
Clin Calcium ; 15(7): 99-104, 2005 Jul.
Article in Japanese | MEDLINE | ID: mdl-15995304

ABSTRACT

A major goal of periodontal therapy is to reconstruct healthy periodontium destroyed by periodontal diseases. Basic studies have revealed that transplantation of mesenchymal stem cells (MSC) into periodontal defects promotes regeneration of periodontal tissue. We have developed a novel method for periodontal therapy using MSC. Human bone marrow cells are obtained from the iliac crest and expanded in vitro at Cell and Tissue Engineering Center in Hiroshima University Hospital. MSC are, then, isolated and mixed with Atelocollagen at final concentrations of 2 x 10(7) cells/mL. These MSC in Atelocollagen are transplanted into periodontal osseous defects at the periodontal surgery. The results in all seven patients who received the own MSC transplantation have shown good clinical course. Further basic studies and the continuous clinical trial are needed to prove the effectiveness of the clinical application.


Subject(s)
Mesenchymal Stem Cell Transplantation/methods , Periodontal Diseases/therapy , Humans , Periodontium/physiology , Regeneration/physiology , Tissue Engineering
16.
Biochem Biophys Res Commun ; 332(1): 297-303, 2005 Jun 24.
Article in English | MEDLINE | ID: mdl-15896330

ABSTRACT

To characterize mesenchymal stem cells (MSC), we compared gene expression profiles in human bone marrow MSC (11 lines) and human fibroblasts (4 lines) by RT-PCR and real time PCR. Messenger RNA levels of MHC-DR-alpha, MHC-DR-beta, MHC-DR-associated protein CD74, tissue factor pathway inhibitor-2, and neuroserpin were much higher in MSC than in fibroblasts, even in the presence of large interindividual variations. Those of adrenomedullin, apolipoprotein D, C-type lectin superfamily member-2, collagen type XV alpha1, CUG triplet repeat RNA-binding protein, matrix metalloproteinase-1, protein tyrosine kinase-7, and Sam68-like phosphotyrosine protein/T-STAR were lower in MSC than in fibroblasts. FACS analysis showed that cell surface expression of MHC-DR was also higher in MSC than in fibroblasts. MHC-DR expression decreased after osteogenic differentiation, whereas the expression of adrenomedullin-a potent stimulator of osteoblast activity-along with collagen XV alpha1 and apolipoprotein D increased after osteogenic differentiation. The marker genes identified in this study should be useful for characterization of MSC both in basic and clinical studies.


Subject(s)
Biomarkers/metabolism , Cell Separation/methods , Fibroblasts/cytology , Fibroblasts/metabolism , Gene Expression Profiling/methods , Mesenchymal Stem Cells/cytology , Mesenchymal Stem Cells/metabolism , Cells, Cultured , Humans
17.
J Bone Miner Res ; 20(3): 399-409, 2005 Mar.
Article in English | MEDLINE | ID: mdl-15746984

ABSTRACT

UNLABELLED: We isolated and expanded BMSCs from human alveolar/jaw bone at a high success rate (70%). These cells had potent osteogenic potential in vitro and in vivo, although their chondrogenic and adipogenic potential was less than that of iliac cells. INTRODUCTION: Human bone marrow stromal cells (BMSCs) have osteogenic, chondrogenic, and adipogenic potential, but marrow aspiration from iliac crest is an invasive procedure. Alveolar BMSCs may be more useful for regenerative medicine, because the marrow can be aspirated from alveolar bone with minimal pain. MATERIALS AND METHODS: In this study, alveolar bone marrow samples were obtained from 41 patients, 6-66 years of age, during the course of oral surgery. BMSCs were seeded and maintained in culture with 10% FBS and basic fibroblast growth factor. In addition, BMSCs were induced to differentiate into osteoblasts, chondrocytes, or adipocytes in appropriate medium. RESULTS AND CONCLUSION: From a small volume (0.1-3 ml) of aspirates, alveolar BMSCs expanded at a success ratio of 29/41 (70%). The success rate decreased with increasing donor age, perhaps because of age-dependent decreases in the number and proliferative capacity of BMSCs. The expanded BMSCs differentiated into osteoblasts under osteogenic conditions in 21-28 days: the mRNA levels of osteocalcin, osteopontin, and bone sialoprotein, along with the calcium level, in alveolar BMSC cultures were similar to those in iliac cultures. However, unlike iliac BMSC, alveolar BMSC showed poor chondrogenic or adipogenic potential, and similar differences were observed between canine alveolar and iliac BMSCs. Subsequently, human alveolar BMSCs attached to beta-tricalcium phosphate were transplanted into immunodeficient mice. In transplants, new bone formed with osteoblasts and osteocytes that expressed human vimentin, human osteocalcin, and human GAPDH. These findings suggest that BMSCs have distinctive features depending on their in vivo location and that alveolar BMSCs will be useful in cell therapy for bone diseases.


Subject(s)
Bone Marrow Cells/cytology , Bone Marrow Cells/physiology , Ilium/cytology , Ilium/physiology , Jaw/cytology , Jaw/physiology , Regenerative Medicine , Adipocytes/physiology , Cell Differentiation/physiology , Cell Separation , Cells, Cultured , Chondrocytes/physiology , Chondrogenesis/physiology , Humans , Stromal Cells/physiology
18.
J Sep Sci ; 27(12): 1033-6, 2004 Aug.
Article in English | MEDLINE | ID: mdl-15352724

ABSTRACT

The ability of reversed-phase high-performance liquid chromatography (RP-HPLC) to separate some positionally isomeric disaturated and monounsaturated triacylglycerols (TAGs) as intact species is demonstrated for the first time. Mobile phases of acetonitrile modified with methanol, ethanol, 2-propanol, 1-propanol, 1-butanol, acetone, or dichloromethane were tested for the separation of POP-PPO, PLP-PPL, PEP-PPE, and PDP-PPD (P-palmitic, O-oleic, L-linoleic, E-eicosapentaenoic, D-docosahexaenoic acid residue) on a single RP-HPLC column. The resolution improved with increasing number of double bonds in the acyl residues. While POP and PPO were only partially resolved, PDP and PPD were fully separated with all tested mobile phases, except those containing methanol. Also separated were the four TAGs having the same equivalent carbon number (ECN = 42), PEP, PPE, PDP, and PPD, on a single RP-HPLC column with mobile phase acetonitrile-2-propanol (70:30, v/v) at 0.8 mL/min. In all cases the isomer with the unsaturated acyl residue in either 1- or 3-position was retained more strongly than the respective 2-isomer.


Subject(s)
Chromatography, High Pressure Liquid/methods , Triglycerides/analysis , Triglycerides/chemistry , Chromatography, Liquid , Docosahexaenoic Acids/analysis , Eicosapentaenoic Acid/analysis , Linoleic Acid/analysis , Oleic Acid/analysis , Protein Isoforms , Time Factors
19.
J Biomech ; 36(3): 431-42, 2003 Mar.
Article in English | MEDLINE | ID: mdl-12594991

ABSTRACT

A technique was developed to create a reproducible femoral neck fracture in vitro using 5-month-old JW/CSK series male rabbits. Force attenuation of a newly developed damping material was also evaluated using this model. Ten pairs of the femora with smaller deviations in length and weight were harvested and cleaned of soft tissue. Either a right or left of each pair of the specimens was randomly selected and put into either the control or the experimental group, both of which contained equal numbers of the right and left femora. The specimens were attached to an L-shaped plate and embedded in a resin from the proximal diaphysis to the distal end so as to maintain a consistent position of the femora. They were mounted and fixed on a pedestal slanted in the coronal plane at 20 degrees. The impact load testing was conducted using an impact mallet dropped from a height of 3 cm. The impact load was applied onto the femoral head. To the specimens in the experimental group, attenuated impact forces were loaded through the damping material, but those in the control group were subjected to forces directly transmitted without the material. All the impact testing was performed in a temperature and humidity controlled chamber. All of the femoral specimens exposed to the direct impact forces (controlled group) sustained fracture at the neck. The fracture line passed from the base of the femoral head laterally and to the calcar area just proximal to the minor trochanter medially. The location of each fracture line was almost identical among the specimens. None of the specimens that were exposed to the impact force through the damping material (experimental group) sustained fracture macroscopically and roentgenographically.


Subject(s)
Femoral Neck Fractures/physiopathology , Fracture Fixation/methods , Weight-Bearing , Animals , Compressive Strength , Epoxy Resins/therapeutic use , In Vitro Techniques , Male , Models, Biological , Rabbits , Reference Values , Stress, Mechanical
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