日常臨床において義歯を装着した患者が,粘膜部の疼痛を訴えることがよくある.しかし,不適合義歯がどの程度,患者にストレスとなっているかの具体的指標はない.そこで健常者に実験的口蓋床を用いて口蓋粘膜の局所に圧刺激を与え,その時の唾液中のコルチゾール濃度とα-アミラーゼの活性変動を測定することにより,口蓋粘膜への不快刺激によるストレス反応を客観的に評価することを目的とした.被験者は,健常有歯顎者(男性6名,女性7名;平均年齢26.4±1.8歳)とした.実験は,口蓋床装着(以下D),局所加圧プレートを付与した口蓋床装着(以下DP)および口蓋床非装着(以下N)をランダムな順で各20分間装着させ唾液を採取した.局所加圧プレートは正中口蓋縫合線と左右第一大臼歯の近心頬側咬頭間線が交叉する点に直径2.5mm,厚さ0.5mmのアクリル板を口蓋床の内面に接着した.実験の前後に不安状態を測定できるSTAIを記入させた.各条件の終了後に疼痛および違和感についてのVASをDPおよびDの条件で記入させた.得られた唾液サンプルは,コルチゾールおよびα-アミラーゼ専用の分析キットを用いて測定した.測定結果を比較検討した結果,コルチゾール濃度およびα-アミラーゼ活性値は,DとNとの間には有意差はみられなかったがDPはNおよびDに比べ有意に増加していた(p<0.01).コルチゾール濃度,α-アミラーゼ活性値および,それぞれの増加の程度を表すDP/D比をそれぞれ求め,STAIおよびVASとの関係を調べた.α-アミラーゼとSTAI特性不安との間(r=0.658,p<0.05)およびα-アミラーゼのDP/D比と疼痛のVASとの間(r=0.603,p<0.05)に正の相関がみられた.以上の結果から,口蓋粘膜への局所圧刺激はコルチゾール濃度およびα-アミラーゼ活性値を上昇させることが明らかとなった.また,疼痛などの不快感を伴う不適合義歯は患者にとってストレッサとなることが示唆された.さらに不安傾向にある者は,不安傾向にない者と比較して,同程度の口蓋部への刺激でも,よりストレスを生じやすい可能性が示唆された.

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下顎枝矢状分割術は顎変形症に対して広く用いられているが,術後経過中に後戻りを経験することがある.後戻りの原因の一つとして,術後咬合力などの生体力学的影響が考えられる.そこで,われわれは術後の咬合力や閉口筋筋力が下顎骨や骨接合プレートに及ぼす応力分布の推移について三次元有限要素法を用い経時的に応力解析を行い検討した.すなわち,CT画像データから下顎枝矢状分割術後1か月,3か月,6か月および12か月の下顎骨モデルを作製し,各モデルに咬合力および閉口筋筋力を付与し近位骨片,遠位骨片,骨接合部および骨接合プレートにおける相当応力を調べた.近位および遠位骨片における相当応力は術後1か月から術後12か月にかけて経時的に増大傾向を認めた.特に,各期間において下顎枝前縁部に高い応力の集中を認めた.骨接合部における相当応力は術後1か月から術後6か月にかけて応力の増大を認めたが,術後12か月では減少していた.術後の咬合力および閉口筋筋力により,近位骨片の下顎枝前縁部に応力の集中を認め,術後3か月から6か月の間に骨接合部の骨切り線の上端と下端に引っ張り応力の集中を認めたことから,術後6か月までは遠位骨片に時計回りの回転方向の応力が加わることが生体力学的解析で判明した.以上のことから,下顎枝矢状分割術を施行した際,術後3か月から6か月の間に後戻りの危険性のあることが示唆された.

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本研究の目的は,インプラント体周囲の歯槽骨に及ぼすPRP(Platelet-rich plasma)の影響を明らかにすることである.現在までPRPに関して様々な報告がなされているが,インプラント埋入後の骨形成初期過程においてPRP中のどのような成長因子が,インプラント体周囲の新生骨形成を促進しているのか,さらに,骨質,新生骨量および術前と術後の新生骨の骨石灰化速度の違いについては明確にされていない.そこで本研究は免疫組織化学的染色ならびに組織学的手法で上記の点を観察した.雌性ビーグル成犬6頭を用い,実験的にインプラント埋入時の骨裂開を想定した歯槽骨欠損を作製し,欠損部にインプラント体を埋入した.それぞれのインプラント体周囲の骨欠損部に血餅群,自家骨のみを移植した群,PRPのみを移植した群およびPRP+自家骨を移植した群に分けて比較検討した.インプラント体周囲の骨欠損部へPRPを併用することにより,以下の結果を得た.1.組織学的所見において,PRP+自家骨移植群に幼若な新生血管および新生骨を認めた.2.免疫組織化学的所見において,初期過程に放出される成長因子はいずれの過程においてもTGF-βおよびbFGFが高値を示した.また,PRP+自家骨移植群において早期のVEGF陽性反応を認めた.3.骨組織形態計測法による観察において,骨質,新生骨量および術前と術後の新生骨の骨石灰化速度が明らかになり,PRP+自家骨移植群では他の群と比較して骨質,新生骨量および骨石灰化速度のすべてにおいて高値を示していた.また,PRP移植群に比べ自家骨移植群に新生骨形成能があることを認めた.さらに,Villanueva bone stainは新生骨の観察には有効な手法であると考えられた.以上の結果より,インプラント体周囲の骨欠損部に対する自家骨移植にPRPを併用すると,組織の創傷治癒に必要な血管の増生を促進し,長期的な骨再生に効果があることが示唆された.

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There are many methods for regeneration of periodontal ligament damaged by periodontal disease. Although enamel matrix derivative (EMD) is one of the most effective methods, its mechanism is not well understood. We explored how the cell proliferation, differentiation and gene expression of periodontal ligament fibroblasts (HPdLF) are influenced by peptides derived from EMD. In addition, we studied hybridization using microarray analysis. Microarray analysis showed that the expression of genes related to osteogenesis increased with the use of EMD. Our study showed that peptides derived from EMD are beneficial to periodontal ligament cells.

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We created periodontal defects on the palatal roots of normal and type 2 diabetes model rat maxillary first molars in order to examine the relation between advanced glycation end products (AGEs) and type III collagen. We observed the pathology and histology of the early wound healing. The experimental group showed poor formation of type III collagen and increased amorphous blood capillary localization of AGEs in vascular endothelium. It is suggested that AGEs inhibit formation collagen and interfere with the early wound healing process.

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To prevent alveolar bone resorption in periodontitis, it is important to understand the changes in expression of various molecules during differentiation of osteoclasts from their precursor cells. Since osteoclasts originate from monocytes and macrophages, we looked at changes in mRNA expression of osteoprotegerin (OPG), receptor activator of NF κ-B ligand (RANKL) and receptor activator of NF κ-B (RANK) during differentiation of the THP-1 cell, a human monocytic leukemia cell line, into macrophages. We demonstrated that OPG, a secretory protein and a competitive inhibitor of RANK binding to RANKL, was induced when THP-1 monocytes differentiated into macrophages. In contrast, expression of RANK or RANKL did not change during the differentiation. The induction of OPG mRNA in macrophages possibly influences the regulation of the differentiation of osteoclasts. This work was supported in part by Grant 07-05 from Osaka Dental University Research Funds.

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Mesenchymal stem cells (MSCs) are highly proliferative and can differentiate into multiple lineages. It has been demonstrated that S100 A4, a S100 calcium-binding protein, can act as an inhibitor of mineralization. We investigated the effect of S100 A4 inhibition by short interfering RNA (siRNA), which is a modified siRNA with improved silencing longevity. We also studied the specific effect of siRNA on the expression of many genes related to osteogenic differentiation. Inhibition of S100 A4 by siRNA resulted in increased expression of many genes related to osteogenic differentiation. This study provided a tool for better understanding of osteogenesis differentiation in MSCs.

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We investigated the relationship between discomfort and stress from an experimental acrylic plate covering the palatal mucosa (exp. plate) using localized pressure plates of varying thickness. The fully dentate subjects were examined under the following five experimental conditions: without an exp. plate (N), with an exp. plate (D), and with a combination of an exp. plate and localized pressure plates of thickness of 0.1mm, 0.2mm and 0.5mm (DP 1, DP 2, and DP 3, respectively). These apparatuses were placed in the oral cavity under pressure for 5min and then left in place at rest for 15min. Cortisol concentration and α-amylase activity were measured using Enzyme-Linked Immunosorbent Assay (ELISA) and a kinetic measurement kit. We found a significant increase in the salivary cortisol concentration and α-amylase activity caused by the increase in discomfort resulting from localized pressure.

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We measured the retentive force and negative pressure generated between the inner and outer crowns of electroformed telescope crowns having 0°, 2°, 4° and 6° taper with glycerin solutions of different viscosities. The peak retentive force was significantly greater at a glycerin concentration of 20% or more, when compared with no solution. The increase in maximum negative pressure was greater at 0° or 2° than at 4° or 6°, and greater increases were observed with glycerin concentrations of 60% and 80% compared with lower concentrations. At 2°, the retentive force did not increase significantly as the glycerin concentration increased when holes were present in the outer crowns. The above findings suggest that the viscosity of saliva and the negative pressure generated between the inner and outer crowns effectively increases the retentive force in a 2° tapered electroformed telescope crown.

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We examined the detection of the human papillomaviruses (HPV) p53, mutant type p53, Rb and p16 in oral dysplasias and oral squamous cell carcinomas. We used 10 samples each of irritation fibromas (IF), low grade dysplasias (LGD), high grade dysplasias (HGD) and squamous cell carcinomas (SCC) obtained from Osaka Dental University Hospital. We detected the DNA of HPV16 and HPV18 by polymerase chain reaction (PCR) and the mRNA of HPV by in situ hybridization (ISH). We also studied the immunohistochemical expression of p53, mutant type p53, Rb and p16. HPV16 was detected in almost all cases of LGD and SCC, but in no cases of ISH. Marked expression of mutant type p53 and Rb was seen in HPV16 positive LGD and SCC. HPV infections were detected by PCR in almost all cases of HGD and SCC. Specimens that showed over expression of mutant type p53 and Rb were almost all positive for HPV16. It is suggested that HPV16 may play an important role in the transformation of p53 and Rb.

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Natural killer (NK) cells exhibit their cytotoxicity through adhesion to the target cells via adhesion molecules. We investigated the effect of the adhesion molecules on NK cell cytotoxicity. Cytotoxicity assay revealed that cells of the NK cell line NK 92 MI kill the cells of the melanoma cell line WM 35. Fluorescence-activated cell scanning (FACS) analysis confirmed the expression of CD 11 a/CD 18 (LFA-1) by NK 92 MI cells and CD 54 by WM 35 cells. Next, we investigated whether the cytotoxicity was dependent on the adhesion molecule CD 54. The anti-CD 54 neutralizing antibody significantly suppressed the killing of WM 35 cells by NK 92 MI cells. These results suggest that the killing of WM 35 cells by NK 92 MI cells might be CD 54-dependent.

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It is known that the N-methyl-D-aspartate (NMDA) receptor is important for learning/memory capability and neuropathic pain. Although it has been reported that NMDA receptor antagonists cause learning/memory disorder, they are effective for neuropathic pain. We investigated the relation between the NMDA receptor and learning/memory capability using neuropathic pain chronic constriction injury (CCI) model rats on the 8-arm radial maze and the step-through type passive avoidance apparatus. The CCI model rats performed better on the tests than either the normal animals or those that had received NMDA receptor antagonists. These findings suggest that the neuropathic pain of the CCI rats raised their learning/memory ability through the NMDA receptor.

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During anesthesia, muscarinic parasympathetic antagonists can be used as a preoperative medication on order to reduce salivary flow, tracheobronchial secretions, and pharyngeal secretions, as well as to decrease the acidity of gastric secretions. We did an in vivo study to examine whether muscarinic receptor subtypes play a role in the exocrine function of the rat submandibular gland. The effects of atropine, pirenzepine, methoctramine, 4-diphenylacetoxy-N-dimethylpiperidinium (4-DAMP) and glycopyrrolate were examined on secretion from the submandibular gland evoked by acetylcholine in pentobarbital-anesthetized rats. Glycopyrrolate caused less elevation in heart rate compared with other anticholinergics. Atropine, 4-DAMP and glycopyrrolate markedly inhibited acetylcholine-evoked fluid responses. Pirenzepine showed significantly lower inhibitory potency than atropine, 4-DAMP or glycopyrrolate, while methoctramine had even less of an inhibitory effect. Pirenzepine, 4-DAMP and glycopyrrolate significantly inhibited both protein concentration and amylase activity in the acetylcholine-evoked submandibular saliva, while methoctramine did not affect the responses. The reduction of the protein concentration and amylase activity in submandibular saliva caused by pirenzepine, 4-DAMP and glycopyrrolate were inhibited by N^G-Nitro-L-arginine methyl ester (L-NAME). Thus, it might be effective to administer anticholinergic drugs together with L-NAME as premedication. These results suggest that glycopyrrolate significantly decreases the salivary secretion, and has significantly fewer side effects than other muscarinic parasympathetic antagonists. Further controlled studies are required to determine the safety, efficacy and patient tolerance of glycopyrrolate.

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Brief exposure to volatile anesthetics before prolonged ischemia exerts a cardioprotective effect. We investigated whether extracellular signal-regulated kinase (ERK) is involved in sevoflurane-induced preconditioning. Isolated perfused guinea pig hearts were subjected to 30min global ischemia and 120min reperfusion (CTL). Sevofluraneinduced preconditioning was elicited by administration of sevoflurane at one minimum alveolar concentration with 10min washout before ischemia (SEVO). Infarct size was significantly reduced in SEVO compared to CTL. Phosphorylation of ERK at 10min after reperfusion was significantly increased in SEVO compared with CTL. Activation of ERK plays an important role in sevoflurane-induced preconditioning. This work was supported in part by Grant 07-06 from Osaka Dental University Research Funds.

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