Hiroto Mitsui

Mesenchymal stem cells cultured under hypoxia escape from senescence via down-regulation of p16 and extracellular signal regulated kinase.

Hypoxia has been considered to affect the properties of tissue stem cells including mesenchymal stem cells (MSCs). Effects of long periods of exposure to hypoxia on human MSCs, however, have not been clearly demonstrated. MSCs cultured under normoxic conditions (20% pO(2)) ceased to proliferate after 15-25 population doublings, while MSCs cultured under hypoxic conditions (1% pO(2)) retained the ability to proliferate with an additional 8-20 population doublings. Most of the MSCs cultured under normoxic conditions were in a senescent state after 100days, while few senescent cells were found in the hypoxic culture, which was associated with a down-regulation of p16 gene expression. MSCs cultured for 100days under hypoxic conditions were superior to those cultured under normoxic conditions in the ability to differentiate into the chondro- and adipogenic, but not osteogenic, lineage. Among the molecules related to mitogen-activated protein kinase (MAPK) signaling pathways, extracellular signal regulated kinase (ERK) was significantly down-regulated by hypoxia, which helped to inhibit the up-regulation of p16 gene expression. Therefore, the hypoxic culture retained MSCs in an undifferentiated and senescence-free state through the down-regulation of p16 and ERK.

Prostaglandin E2 receptor type 2-selective agonist prevents the degeneration of articular cartilage in rabbit knees with traumatic instability

IntroductionOsteoarthritis (OA) is a common cause of disability in older adults. We have previously reported that an agonist for subtypes EP2 of the prostaglandin E2 receptor (an EP2 agonist) promotes the regeneration of chondral and osteochondral defects. The purpose of the current study is to analyze the effect of this agonist on articular cartilage in a model of traumatic degeneration.MethodsThe model of traumatic degeneration was established through transection of the anterior cruciate ligament and partial resection of the medial meniscus of the rabbits. Rabbits were divided into 5 groups; G-S (sham operation), G-C (no further treatment), G-0, G-80, and G-400 (single intra-articular administration of gelatin hydrogel containing 0, 80, and 400 μg of the specific EP2 agonist, ONO-8815Ly, respectively). Degeneration of the articular cartilage was evaluated at 2 or 12 weeks after the operation.ResultsONO-8815Ly prevented cartilage degeneration at 2 weeks, which was associated with the inhibition of matrix metalloproteinase-13 (MMP-13) expression. The effect of ONO-8815Ly failed to last, and no effects were observed at 12 weeks after the operation.ConclusionsStimulation of prostaglandin E2 (PGE2) via EP2 prevents degeneration of the articular cartilage during the early stages. With a system to deliver it long term, the EP2 agonist could be a new therapeutic tool for OA.

Recipient bone marrow-derived stromal cells prolong graft survival in a rat hind limb allotransplantation model

Recent studies have indicated that bone marrow‐derived stromal cells (BMSCs) have immunomodulatory properties that suppress the T cell responses that cause graft rejection. The purpose of this study is to evaluate the effect of recipient BMSCs intravenous infusion for immunomodulation in a rat vascularized composite allotransplantation model.

FRI0165 Relationship Between Serum Concentrations of Cytokines and Disease Activity in Rheumatoid Arthritis Patients Treated with Tocilizumab

Background The balance between T type 17 cells (Th17) and regulatory T cells (Treg) affects inflammation in RA pathogenesis, and both subpopulations depend on transforming growth factor (TGF)-b for their induction, but the presence of interleukin (IL)-6 disturbs this balance and induction of T17 may be increased. Objectives The aim of the present study was to investigate changes in serum concentrations of cytokines such as IL-6, IL-17, tumor necrosis factor (TNF)-a and TGF-b in patients treating with tocilizumab (TCZ), an IL-6 receptor antibody, before and after treatment. Methods Participants comprised 31 patients (23 women, 8 men) who met the 2010 ACR/EULAR classification criteria. At baseline and at weeks 24 and 52 of TCZ treatment, clinical results were obtained from clinical records and evaluations were made using the 28-joint disease activity score (DAS28-ESR). Peripheral blood was obtained at the same time and serum concentrations of cytokines were analyzed. Results Thirty patients received follow-up, and 3 patients ceased TCZ therapy because of side effects. DAS28-ESR scores decreased from 5.09 at baseline, to 2.95 in Week 24 and 2.66 in Week 52. The number of good responders was 17 in week 24, and 19 in week 52, compared to 9 moderate responders in week 24, and 7 in week 52, and 2 non-responders in week 24, and 1 in week 52, according to the clinical evaluation criteria of EULAR. Twelve patients were in remission state (DAS28-ESR) at week 56. Mean concentration of serum IL-6 was 19.8 (pg/ml) at baseline and 30.2 (pg/ml) at week 56 in good responders at week 56 (Group A), compared to 45.0 (pg/ml) at baseline and 73.6 (pg/ml) at week 56 in moderate responders at week 56 (Group B). Mean serum concentration of TNF-a was 18.6 (pg/ml) at baseline and 10.4 (pg/ml) at week 56 in Group A, and 12.6 (pg/ml) and 12.9 (pg/ml) in Group B. Mean serum concentration of TGF-β in group A was 34.8 (ng/ml) at baseline and 30.6 (ng/ml) at 56 weeks in Group A, and 37.7 (ng/ml) and 31.5 (ng/ml) in Group B. Mean serum concentration of IL-17A was 6.08 (pg/ml) at baseline and 1.76 (pg/ml) at week 56 in Group A, and 6.56 (pg/ml) and 4.25 (pg/ml) in Group B. Mean serum concentration of TNF-α was 18.6 (pg/ml) at baseline and 10.4 (pg/ml)at week 56 in Group A, and 12.6 (pg/ml) and 12.9 (pg/ml) in Group B. Conclusions As 63.3% of patients treated with TCZ were good responders and 44.4% of patients were in remission at week 52, the clinical effectiveness of TCZ therapy was confirmed. Comparing Groups A and B, serum IL-6 level was lower in good responders before treatment. Serum IL-6 level at baseline may reflect disease activity, and high IL-6 level at baseline may suggest poor prognosis after TCZ therapy. Otherwise, TNF-a showed no significant correlation with disease activity markers at baseline. Serum level of TNF-a at baseline may not indicate prognosis after TCZ therapy. Mean serum level of IL-17A was significantly lower in Group A at week 52. Mean serum level of TGF-b showed no significant differences among the Groups at baseline, but the reduction after TCZ therapy was smaller in Group A. This result indicates that a high concentration of serum IL-17A and a low concentration of serum TGF-b correlate with poor efficacy of TCZ therapy. Disclosure of Interest H. Mitsui: None declared, K. Yoshikawa Shareholder of: Chugai Pharmaceutical Co., Ltd, Y. Nagaya Shareholder of: Viomet Japan, T. Otsuka: None declared, T. Mitsui: None declared