Orthodontic tensile strain induces angiogenesis via type IV collagen degradation by matrix metalloproteinase-12.

BACKGROUND AND OBJECTIVE: During orthodontic tooth movement (OTM), periodontal ligament (PDL) is remodeled dynamically, which requires sufficient blood supply for the regeneration of PDL. However, little is known about the remodeling of blood vessels during OTM. In this study, we hypothesized that the orthodontic tensile strain upregulates matrix metalloproteinase-12 (MMP-12) expression in the tension zone and induces angiogenesis via degradation of type IV collagen (Col-IV) in vascular endothelial basement membrane during the early stage of OTM. MATERIAL AND METHODS: Temporal and spatial MMP-12 expression in the tension zone of PDL, during the early stage of OTM, were examined by immunohistochemistry in rats. Continuous tensile strain was applied to cultured human immortalized PDL cell lines (HPL cells) and MMP-12 expression was examined in vitro. Colocalization of MMP-12 and Col-IV in vivo were examined by immunohistochemistry. To investigate whether MMP-12 produced by HPL cells could degrade Col-IV, recombinant Col-IV was incubated in the culture supernatants of HPL cells. Intact Col-IV in vitro was also examined by western blot analysis. Finally, the changes in blood vessels in the PDL were examined by micro-computed tomography analysis with perfused contrast agents and by conventional histological analysis. RESULTS: Orthodontic tensile strain induced MMP-12 expression in PDL cells in vivo and in vitro. Immunohistochemistry revealed that MMP-12-positive cells were observed adjacent to the Col-IV-positive tubular area in the tension zone of PDL. MMP-12 in culture supernatant of HPL cells degraded recombinant Col-IV, and specific MMP-12 inhibitor blocked the Col-IV degradation. Micro-computed tomography analysis and conventional histological analysis demonstrated that the areas of blood vessels were increased in the tension zone of the PDL after OTM. CONCLUSION: We discovered that the orthodontic tensile strain upregulates MMP-12 expression in the tension zone of PDL and induces angiogenesis via degradation of Col-IV in the vascular endothelial basement membrane.

Structural basis of pathogen recognition by an integrated HMA domain in a plant NLR immune receptor.

Plants have evolved intracellular immune receptors to detect pathogen proteins known as effectors. How these immune receptors detect effectors remains poorly understood. Here we describe the structural basis for direct recognition of AVR-Pik, an effector from the rice blast pathogen, by the rice intracellular NLR immune receptor Pik. AVR-PikD binds a dimer of the Pikp-1 HMA integrated domain with nanomolar affinity. The crystal structure of the Pikp-HMA/AVR-PikD complex enabled design of mutations to alter protein interaction in yeast and in vitro, and perturb effector-mediated response both in a rice cultivar containing Pikp and upon expression of AVR-PikD and Pikp in the model plant Nicotiana benthamiana. These data reveal the molecular details of a recognition event, mediated by a novel integrated domain in an NLR, which initiates a plant immune response and resistance to rice blast disease. Such studies underpin novel opportunities for engineering disease resistance to plant pathogens in staple food crops.

Nrf2 activation attenuates both orthodontic tooth movement and relapse.

During orthodontic tooth movement, osteoclasts resorb the alveolar bone at the compress side of periodontium. Reactive oxygen species (ROS) works as intracellular signaling molecules of RANKL during osteoclastogenesis, although ROS has cytotoxicity against cells such as lipid oxidation. To deal with oxidative stress, cells have a defense system that is scavenging ROS by augmented antioxidative stress enzymes via transcriptional regulation with nuclear factor E2-related factor 2 (Nrf2). Previously, we reported that augmented antioxidative stress enzymes by Nrf2-gene transfer inhibited bone destruction. In the present study, we examined the effects of Nrf2 activation on osteoclastogenesis and, thereby, orthodontic tooth movement and orthodontic relapse. Mouse macrophage cell line RAW264.7 cells were used as osteoclast progenitor cells and stimulated with recombinant RANKL (100 ng/mL) with or without Nrf2 activator sulforaphane (SFN) and epigallocatechin gallate (EGCG) or ROS scavenger catechin. Osteoclastogenesis, resorption activity, and osteoclast marker gene expression were examined. Intracellular ROS was analyzed by flow cytometry. Maxillary first molars of C57BL6 male mice were moved palatally with 0.012-inch NiTi wire (100-mN force); SFN or EGCG was injected into the palatal gingiva once a week; and phosphate buffered saline was injected on the contralateral side. Tooth movement was monitored using a stone model with precise impression, and the amount of the tooth movement was compared among groups. SFN and EGCG significantly, but catechin weakly, inhibited RANKL-mediated osteoclastogenesis in vitro. Western blot analysis revealed that SFN and EGCG augmented the nuclear translocation of Nrf2 and the expression of anti-oxidative stress enzymes such as HO-1, although catechin did not. SFN and EGCG significantly, but catechin weakly, attenuated the intracellular ROS. Finally, animal experiment revealed that both SFN and EGCG successfully inhibited the orthodontic tooth movement. Additionally, SFN inhibited the relapse. These results suggest that Nrf2 activation could be therapeutic target for the anchorage enforcement in orthodontic treatment and pharmacologic retention against relapse.

Intralesional steroid injection to prevent stricture after endoscopic submucosal dissection for esophageal cancer: a controlled prospective study.

BACKGROUND AND STUDY AIMS: The frequency of stricture after endoscopic submucosal dissection (ESD) for esophageal squamous cell carcinoma with a mucosal defect involving more than three-quarters of the circumference is 70% - 90%. Stricture decreases quality of life and requires multiple endoscopic balloon dilation (EBD) sessions. We investigated the efficacy and safety of a single session of intralesional steroid injections to prevent post-ESD stricture. PATIENTS AND METHODS: We conducted a prospective study on 30 patients with esophageal squamous cell carcinoma treated by ESD, who had a more than three-quarter but less than whole circumferential defect. A single session of intralesional steroid injections was undertaken immediately after ESD. Esophagogastroduodenoscopy was performed whenever patients reported dysphagia and 2 months after ESD in patients without dysphagia. Results were compared with a historical control group of 29 patients who underwent ESD without intralesional steroid injection. The primary endpoint was the post-ESD stricture rate. Secondary endpoints were the number of EBD sessions and the complication rate. RESULTS: Compared with the historical control group, the study group had a significantly lower stricture rate (10%, 3/30 patients vs. 66%, 19/29 patients; P < 0.0001) and a lower number of EBD sessions (median 0, range 0 - 2 vs. median 2, range 0 - 15; P < 0.0001). The study group had a complication rate of 7 % (2 /30 patients), comprising a submucosal tear in one patient and bleeding in another, which were not a direct result of EBD. CONCLUSIONS: A single session of intralesional steroid injections showed promising results for the prevention of stricture after ESD for esophageal cancer.

Effects of local osteoprotegerin gene transfection on orthodontic root resorption during retention: an in vivo micro-CT analysis.

OBJECTIVES: External root resorption (ERR) is a serious complication of orthodontic treatment. Aim of this study was to evaluate the effects of local osteoprotegerin (OPG) gene transfection on ERR during retention. MATERIAL AND METHODS: Eighteen 6-week-old male Wistar rats were divided into three groups. All the rats were subjected to 2 weeks of orthodontic tooth movement followed by a 2-week retention period. During retention, the three groups of rats received local OPG gene transfection (OPG transfection group, n=6), mock vector transfection (mock group, n=6), or no injections (control group, n=6). ERR of all three groups was evaluated with in vivo micro-CT analysis at three different time points: baseline, the last day of orthodontic tooth movement, and the last day of retention. RESULTS: In the OPG transfection group, there was no significant difference between ERR at the baseline and ERR on the last day of retention. By the last day of retention, the repair ratio of ERR in the OPG transfection group was statistically higher in relation to the repair ratio of the other groups (p<0.001). CONCLUSION: The results indicated that local OPG gene transfection significantly enhanced the repair of ERR during retention. Local OPG gene transfection might therefore be a useful tool for ERR repair during retention.

Prenatal diagnosis of fetal left pulmonary artery sling.

Left pulmonary artery (LPA) sling is a very rare anomaly in which the LPA arises distally, far from the right pulmonary artery on the right side of the distal trachea, turns sharply leftwards around the trachea and courses to the left lung hilum through the space between the trachea and esophagus. LPA sling is often associated with distal tracheal narrowing, due to either intrinsic stenosis or secondary compression by the anomaly itself. To our knowledge, prenatal diagnosis of LPA sling has not been reported so far. We report a case in which LPA sling was diagnosed during fetal ultrasound examination.

Clinical features and outcomes of delayed perforation after endoscopic submucosal dissection for early gastric cancer.

Perforation is a major complication of endoscopic submucosal dissection (ESD) for early gastric cancer (EGC). However, there have been no reports on delayed perforation after ESD for EGC. We aimed to elucidate the incidence and outcomes of delayed perforation after ESD. Clinical courses in 1159 consecutive patients with 1329 EGCs who underwent ESD were investigated. Delayed perforation occurred in six patients (0.45 %). All these patients had complete en bloc resection without intraoperative perforation during ESD. Five of six perforations were located in the upper third of the stomach, while one lesion was found in the middle third. Symptoms of peritoneal irritation with rebound tenderness presented within 24 h after ESD in all cases. One patient did not require surgery because the symptoms were localized, and recovered with conservative antibiotic therapy by nasogastric tube placement. The remaining five patients required emergency surgery. There was no mortality in this case series.

Altered activity of lysophospholipase D, which produces bioactive lysophosphatidic acid and choline, in serum from women with pathological pregnancy.

Altered lipid metabolism is associated with human abnormal pregnancy, such as pre-eclampsia and preterm labor, and potentially leads to fetus loss. A causative factor for the onset and progress of the systemic multifactorial syndromes associated with the pathological pregnancy is oxidized low-density lipoprotein, an active identity of which was postulated to be lysophosphatidic acid (LPA). We previously found that LPA is produced extracellularly by plasma lysophospholipase D (lysoPLD) activity of autotaxin, a tumor cell motility-stimulating protein. In this study, a convenient assay based on the choline released from endogenous substrate or exogenous lysophosphatidylcholine (LPC) was used for comparison of serum lysoPLD activity among patients with normal and abnormal pregnancy. The serum choline-producing activity was found to be mainly due to autotaxin, and dependent on its dilution rate. There was some association between low dilution dependency of serum lysoPLD activity toward an exogenous LPC and high lysoPLD activity toward endogenous substrates in cases of patients with preterm labor and pre-eclampsia. However, there was no difference in the serum level of LPC between women with normal pregnancy and those with pathological pregnancy. These results indicate that production of bioactive LPA by lysoPLD activity is elevated by an unknown mechanism that may be related to increased availability of endogenous substrates LPC, but not its concentration in human serum. If the level of LPA in blood circulation is elevated in the pathological pregnancies in vivo, it may play a role in induction and/or progression of systemic vascular dysfunction seen patients with preterm labor or pre-eclampsia.

Heterogeneity of regional systolic function detected by tissue Doppler imaging is linked to impaired global left ventricular relaxation in hypertrophic cardiomyopathy.

OBJECTIVE: To evaluate regional and global left ventricular (LV) function and LV wall thickness (LVWT) in patients with hypertrophic cardiomyopathy (HCM). DESIGN AND SETTING: Observational study at the National Cardiovascular Centre and Nagoya University Hospital in Japan. PARTICIPANTS: Thirty-six patients with HCM and 16 patients with hypertensive LV hypertrophy (LVH). MAIN OUTCOME MEASURES: Conventional echocardiography and strain rate (SR) imaging derived from tissue Doppler imaging were performed. Systolic strain (epsilon(sys)), peak systolic SR (SR(sys)), peak early diastolic SR (SR(dia)) and LVWT were obtained from eight LV segments. LV pressure was simultaneously recorded with a high-fidelity micromanometer. RESULTS: The regional epsilon(sys) and SR(sys) were correlated with LVWT in patients with HCM (r = 0.50, p<0.001 and r = 0.63, p<0.001, respectively) but not in patients with hypertensive LVH. The standard deviations of LVWT, epsilon(sys) and SR(sys) obtained from the eight LV segments of each subject were greater for patients with HCM than for patients with hypertensive LVH. The standard deviation of LVWT was correlated with those of epsilon(sys) and SR(sys) (r = 0.55, p<0.001 and r = 0.56, p<0.001, respectively). The standard deviations of LVWT, epsilon(sys) and SR(sys) were correlated with tau (r = 0.35, p<0.05; r = 0.47, p<0.001; and r = 0.39, p<0.005, respectively). CONCLUSIONS: Heterogeneity of regional LV systolic function detected by SR imaging is in part attributable to heterogeneity of LVH and may be linked to impaired global LV relaxation in HCM.

Clodronate inhibits PGE(2) production in compressed periodontal ligament cells.

Periodontal ligament (PDL) cells play an essential role in orthodontic tooth movement. We recently reported that clodronate, a non-N-containing bisphosphonate, strongly inhibited tooth movement in rats, and thus could be a useful adjunct for orthodontic treatment. However, it is not clear how clodronate affects the responses of PDL cells to orthodontic force. In this study, we hypothesized that clodronate prevents the mechanical stress-induced production of prostaglandin E(2) (PGE(2)), interleukin-1beta (IL-1beta), and nitric oxide (NO) in human PDL cells. A compressive stimulus caused a striking increase in PGE(2) production, while the responses of IL-1beta and NO were less marked. Clodronate concentration-dependently inhibited the stress-induced production of PGE(2). Clodronate also strongly inhibited stress-induced gene expression for COX-2 and RANKL. These results suggest that the inhibitory effects of clodronate on tooth movement and osteoclasts may be due, at least in part, to the inhibition of COX-2-dependent PGE(2) production and RANKL expression in PDL cells.

Cyclical tensile force on periodontal ligament cells inhibits osteoclastogenesis through OPG induction.

The periodontal ligament (PDL) maintains homeostasis of periodontal tissue under mechanical tensile-loading caused by mastication. Occlusal load inhibits atrophic alveolar bone resorption. Previously, we discovered that continuous compressive force on PDL cells induced osteoclastogenesis-supporting activity, with up-regulation of RANKL. We hypothesized that, unlike compression, cyclical tensile force up-regulates OPG expression in PDL cells via TGF-beta up-regulation, and does not induce osteoclastogenesis-supporting activity. PDL cells were mechanically stimulated by cyclical tensile force in vitro. The conditioned media of PDL cells that had been subjected to cyclical tensile force inhibited osteoclastogenesis. Cyclical tensile force up-regulated not only RANKL mRNA expression, but also OPG mRNA expression in PDL cells. Tensile force up-regulated TGF-beta expression in PDL cells as well. Administration of neutralizing antibodies to TGF-beta inhibited OPG up-regulation under cyclical tensile-force stimulation in a dose-dependent manner. Additionally, the osteoclastogenesis-inhibitory effect of the conditioned media of PDL cells under cyclical tensile force was partially rescued by the administration of TGF-beta neutralizing antibodies. In conclusion, tensile force inhibited the osteoclastogenesis-supporting activity of PDL cells by inducing the up-regulation of OPG via TGF-beta stimulation.

Local RANKL gene transfer to the periodontal tissue accelerates orthodontic tooth movement.

It has been reported that not only selective alveolar-bone resorption, but also receptor activator of nuclear factor kappa B ligand (RANKL) expression is induced on the compressed side of an orthodontically moving tooth. Numerous reports have described the pharmacological acceleration of tooth movement (TM) through the activation of osteoclasts. However, because of rapid flush out by blood circulation, daily systemic administration or daily local injection is needed. Previously, we discovered that every-3-days OPG gene transfer to the periodontal-tissue inhibited RANKL-mediated osteoclastogenesis and diminished experimental TM. Therefore, we hypothesized that local RANKL gene transfer into the periodontal tissue would accelerate TM. The upper first molars of 6-week-old male Wistar rats were moved palatally using fixed orthodontic wires. The inactivated hemagglutinating-virus of Japan (HVJ) envelope vector containing the mouse RANKL expression plasmid was injected periodically into the palatal periodontal tissue of the upper first molars during TM. Local RANKL gene transfer significantly enhanced RANKL expression and osteoclastogenesis in periodontal tissue without any systemic effects. The TM rate was significantly increased in the RANKL gene transfer side. In conclusion, we demonstrated that transfer of the RANKL gene to the periodontal-tissue activated osteoclastogenesis and accelerated the amount of experimental TM. Local RANKL gene transfer might be a useful tool not only for shortening orthodontic treatment, but also for moving ankylosed teeth where teeth, fuse to the surrounding bone.

Progesterone induces the fibulin-1 expression in human endometrial stromal cells.

BACKGROUND: By using microarray analysis with human endometrial stromal cells (ESCs), we previously reported that the mRNA for fibulin-1, an extracellular matrix as well as a plasma glycoprotein, is up-regulated by progesterone. In the present study, we tried to clarify the spatial and temporal regulation mechanism of fibulin-1 in the human endometrium. METHODS AND RESULTS: Quantitative analysis with real-time PCR experiments on human endometrial tissues showed significantly higher fibulin-1 mRNA expressions in secretory phase endometria than in proliferative phase. Immunohistochemical studies revealed that the fibulin-1 protein is expressed in the glandular epithelium in proliferative phase endometria, and that expression switched to the stroma in secretory phase endometria. In culture experiments with ESCs, a significant increase of fibulin-1 mRNA expression was observed in cells treated with 6 alpha-methyl-17 alpha-hydroxy-progesterone acetate (MPA) or 8 bromoadenosine 3':5'-cyclic monophosphate (8-Br-cAMP). MPA stimulated the fibulin-1 mRNA expression in a dose-dependent manner, and a progesterone antagonist, RU-486, inhibited the stimulatory effect almost completely. By contrast, beta-estradiol alone did not increase the fibulin-1 mRNA expression. CONCLUSIONS: These results suggest that fiblin-1 is an important molecule that mediates progesterone action in human ESC differentiation towards implantation.

Local OPG gene transfer to periodontal tissue inhibits orthodontic tooth movement.

Previously, we discovered that RANKL expression is induced in compressed periodontal ligament cells, and that this promotes osteoclastogenesis on the compression side in orthodontic tooth movement. We hypothesized that local OPG gene transfer to the periodontium would neutralize the RANKL activity induced by mechanical compressive force, thereby inhibiting osteoclastogenesis and diminishing tooth movement. The upper first molars of six-week-old male Wistar rats were moved palatally by means of a fixed-orthodontic wire. A mouse OPG expression plasmid [pcDNA3.1(+)-mOPG] was constructed, and the production of functional OPG protein was confirmed in vitro. The inactivated HVJ envelope vector containing pcDNA3.1(+)-mOPG or PBS was injected periodically into the palatal periodontal tissue of upper first molars. When this local OPG gene transfer was performed, OPG production was induced, and osteoclastogenesis was inhibited. Local OPG gene transfer significantly diminished tooth movement. In this study, we report that OPG gene transfer to periodontal tissue inhibited RANKL-mediated osteoclastogenesis and inhibited experimental tooth movement.

Monoacylglycerols: glycolipid biosurfactants produced by a thermotolerant yeast, Candida ishiwadae.

AIMS: To isolate and characterize biosurfactants produced by a thermotolerant yeast isolated in Thailand. MATERIALS AND RESULTS: Yeast strains isolated from plant material in Thailand were first screened for the ability to produce lipase and biosurfactant. A strain Y12, identified as Candida ishiwadae by physiological tests, survived at 45 degrees C and produced relatively large amounts of biosurfactants. From the culture filtrate of this strain, two glycolipid biosurfactants, a and b, were purified by solvent fractionation, silica gel and ODS column chromatographies. Compounds a and b were determined to be monoacylglycerols; 1-linoleylglycerol and 1-oleylglycerol, respectively. Both compounds exhibited higher surfactant activities tested by the drop collapse test than several artificial surfactants such as sodium dodecyl sulphate. CONCLUSIONS: Glycolipid biosurfactants produced by a thermotolerant yeast, C. ishiwadae were characterized to be monoacylglycerols which exhibited high surfactant activities. SIGNIFICANCE AND IMPACT OF THE STUDY: A thermotolerant yeast strain, C. ishiwadae, could be a potential candidate for producing monoacylglycerols which are useful in industrial applications.

Microarray analysis of genes controlled by progesterone in human endometrial stromal cells in vitro.

The steroid hormone progesterone is a key factor in establishment and maintenance of pregnancy in the human endometrium. To obtain a global view and identify new target genes for progesterone in human endometrial stromal cells in short-term (3 days) culture, we used a screening strategy to analyze the expression of nearly 1000 human genes by DNA microarray analysis. The results showed that six genes were up-regulated (at least a two-fold increase), and 27 genes were down-regulated (at least a two-fold decrease) after progesterone treatment compared with control. Progesterone stimulated the expression of the interleukin (IL)-1 receptor type 1, fibulin-1, fibulin-2, microsomal glutathione S-transferase 1, fumarylacetoacetate hydrolase and orphan G protein-coupled receptor (RDC1). Progesterone inhibited the expression of insulin-like growth factor binding protein-5, heparin-binding epidermal growth factor-like growth factor, and IL-13 receptor alpha2. In addition, progesterone inhibited the expression of genes involved in immune modulators, DNA/chromatin-related proteins, signal transduction, transcription factors, transport proteins, enzyme, receptor and structural proteins. Our results demonstrate that microarray analysis can be used to identify progesterone-regulated genes in endometrial stromal cells, thus contributing to a more detailed understanding of the molecular mechanisms in response to progesterone in the endometrium during the preparatory period for implantation.

Analyses of dendritic cell subsets in pregnancy.

PROBLEM: Changes in the frequency of dendritic cell (DC) subsets in the peripheral blood were analyzed as pregnancy progressed, and the effects of human chorionic gonadotropin (hCG) on myeloid and lymphoid DC subsets were phenotypically and functionally examined. METHOD OF STUDY: Two major subsets of DCs were prepared from the peripheral blood by flow cytometry. Major histocompatibility complex class II molecules and adhesion/costimulatory molecules were examined before and after culture with hCG. hCG receptors on both DC subsets were analyzed by reverse transcriptase-polymerase chain reaction (RT-PCR). RESULTS: The frequency of myeloid DCs increased in the late stage of pregnancy, while that of lymphoid DCs gradually decreased. The addition of hCG (physiological concentrations in pregnancy) to cultures induced the maturation of both DC subsets in conjunction with increases in the expression of adhesion/costimulatory molecules, their stimulatory activities in allogeneic mixed lymphocyte/leukocyte reaction, and cytokine secretion (interleukin-12 and interferon-gamma). hCG receptors were found in both DC subsets by RT-PCR, suggesting that these stimulatory activities of hCG are mediated by hCG receptors on the DCs. CONCLUSIONS: hCG can modulate immune responses through the activation of myeloid and lymphoid DCs.

Cytosolic HSP90 and HSP70 are essential components of INF1-mediated hypersensitive response and non-host resistance to Pseudomonas cichorii in Nicotiana benthamiana.

SUMMARY Mitogen-activated protein kinases (MAPKs) play pivotal roles in the signal transduction pathway of plant defence responses against pathogens. A search for MAPK-interacting proteins revealed an interaction between a Nicotiana benthamiana MAPK, SIPK (NbSIPK) and cytosolic Hsp90 (NbHsp90c-1) in yeast two-hybrid assay. To study the function of Hsp90 in disease resistance, we silenced NbHsp90c-1 in N. benthamiana by virus-induced gene silencing (VIGS) with Potato virus X (PVX). NbHsp90c-1 silenced plants exhibited: (1) a stunted phenotype, (2) no hypersensitive response (HR) development after infiltration with the Phytophthora infestans protein INF1 and a non-host pathogen Pseudomonas cichorii that normally triggers HR in N. benthamiana, (3) compromised non-host resistance to P. cichorii, and (4) consistently reduced transcription levels of PR (pathogenesis related) protein genes. Similar phenotypes were observed also for plants in which a cytosolic Hsp70 (NbHsp70c-1), a gene for another class of molecular chaperon, was silenced. Hsp90 was isolated as a MAPK-interacting protein in yeast two-hybrid assay, therefore we tested the effect of NbHsp90c-1 silencing as well as NbHsp70c-1 silencing on the HR development caused by infiltration of a hyperactive potato MAPKK (StMEK1(DD)). No difference in the timing or extent of HR was found among NbHsp90c-1 silenced, NbHsp70c-1 silenced and control plants. This result indicates that observed impairment of INF1- and P. cichorii-mediated HR development in NbHsp90c-1 silenced and NbHsp70c-1 silenced plants was not caused by the abrogation in MAPK function downstream of active MAPKK that leads to HR. These findings suggest essential roles of Hsp90 and Hsp70 in plant defence signal transduction pathway upstream or independent of the MAPK cascade.

Overexpression of the wasabi defensin gene confers enhanced resistance to blast fungus ( Magnaporthe grisea) in transgenic rice.

Transgenic rice ( Oryza sativa cv. Sasanishiki) overexpressing the wasabi defensin gene, a plant defensin effective against the rice blast fungus, was generated by Agrobacterium tumefaciens-mediated transformation. Twenty-two T2 homozygous lines harboring the wasabi defensin gene were challenged by the blast fungus. Transformants exhibited resistance to rice blast at various levels. The inheritance of the resistance over generations was investigated. T3 plants derived from two highly blast-resistant T2 lines (WT14-5 and WT43-5) were challenged with the blast fungus using the press-injured spots method. The average size of disease lesions of the transgenic line WT43-5 was reduced to about half of that of non-transgenic plants. The 5-kDa peptide, corresponding to the processed form of the wasabi defensin, was detected in the total protein fraction extracted from the T3 progeny. Transgenic rice plants overproducing wasabi defensin are expected to possess a durable and wide-spectrum resistance (i.e. field resistance) against various rice blast races.

Dual regulation of osteoclast differentiation by periodontal ligament cells through RANKL stimulation and OPG inhibition.

Periodontal ligament (PDL) cells play an important role in maintaining the homeostasis of periodontal tissues. However, it is not known how PDL cells contribute to osteoclastogenesis. In this study, we examined the consequences of cell-to-cell interactions between peripheral blood mononuclear cells (PBMCs) and PDL cells during osteoclastogenesis. PBMCs were co-cultured directly or indirectly with PDL cells for two to four weeks. PBMCs that were directly co-cultured with PDL cells formed significantly more resorption pits on dentin slices than did PBMCs that were cultured alone. However, soluble factor(s) produced from PDL cells inhibited the formation of tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells. Furthermore, PDL cells expressed both receptor activator nuclear factor kappa B ligand (RANKL) and osteoprotegerin (OPG) mRNA. In conclusion, PDL cells support osteoclastogenesis through cell-to-cell contact. PDL cells might regulate osteoclastogenesis by opposing mechanisms--stimulation of resorptive activity by RANKL and inhibition by OPG--thus affecting processes such as periodontitis and orthodontic tooth movement.