Gene expression of growth differentiation factors in the developing periodontium of rat molars.

Growth and differentiation factors (GDF) 5, 6, and 7 are known to play roles in tendon and ligament formation, and are therefore probably involved in the formation of periodontal ligament. In this study, we sought to determine temporal and spatial expression of GDF-5, -6, and -7 mRNA in developing periodontal tissue of rat molars using in situ hybridization. GDF gene expression in the periodontal ligament was first detected in cells associated with the initial process of periodontal ligament fiber bundle formation. Gene signals were also detected in cells located along the alveolar bone and cementum surfaces, the insertion sites of periodontal ligaments, during the course of root formation. GDF expression in these cells were down-regulated after completion of root formation. Our results appeared to suggest the involvement of GDF-5, -6, and -7 in the formation of the dental attachment apparatus.

LPS-stimulated human gingival fibroblasts inhibit the differentiation of monocytes into osteoclasts through the production of osteoprotegerin.

Periodontitis is an inflammatory bone disease caused by Gram-negative anaerobic bacteria, but the precise mechanism of bone destruction remains unknown. Activated T lymphocytes secrete receptor activator of NF-kappaB ligand (RANKL) and support the differentiation of monocytes into mature osteoclasts. The purpose of this study was to examine the expression of RANKL and its inhibitor, osteoprotegerin (OPG), in inflamed gingival tissue and to clarify the role of human gingival fibroblasts (HGFs) in osteoclastogenesis regulated by RANKL. HGFs and gingival mononuclear cells (GMCs) were obtained from chronic periodontitis patients during routine periodontal surgery. Expression of OPG and RANKL mRNA in gingival tissue and HGFs was examined with RT-PCR. OPG production was measured using ELISA. Expression of RANKL, CD4, CD8 and CD69 on GMCs was determined by flow-cytometry using RANK-Fc fusion protein and the respective monoclonal antibodies. Osteoclastogenesis by RANKL was assayed by counting the number of tartarate-resistant acid phosphatase (TRAP)-positive cells after culturing human peripheral blood monocytes with recombinant human RANKL and macrophage-colony stimulating factor (M-CSF) for 10 days. OPG and RANKL mRNA were expressed in 80% (16/20) and 25% (5/20) of periodontitis lesions, respectively. OPG, but not RANKL, mRNA was expressed within HGFs. OPG mRNA expression and production by HGFs was augmented by LPS stimulation. All GMC samples expressed CD69, and two of five GMC samples expressed RANKL. The culture supernatant of LPS-stimulated gingival fibroblasts significantly reduced the number of TRAP positive cells generated by culturing monocytes with RANKL and M-CSF. The present study suggests that LPS-stimulated HGFs inhibit monocyte differentiation into osteoclasts through the production of OPG.

Noggin blocks osteoinductive activity of porcine enamel extracts.

Enamel extracts induce biomineralization such as osteogenesis and cementogenesis, but the molecular component responsible for this activity remains uncertain. We fractionated enamel extracts from developing pig teeth and isolated the osteoinductive fraction. Proteins from pig enamel scrapings were extracted under alkaline conditions (pH 10.8) and fractionated with the use of a Sephadex G-100 (size exclusion) column. The ability of each fraction to enhance alkaline phosphatase (ALP) activity was assayed in ST2 cells, a mouse bone marrow stromal cell line. The osteoinductive fraction of enamel extracts (OFE) was found in fractions 44 and 45, which induced ST2 cells to express the phenotype of bone-forming osteoblasts, and to form mineralized nodules. Furthermore, the ALP activity of ST2 cells exposed to OFE was reduced by noggin, an antagonist of BMPs, and OFE reacted with BMP-2/4 antibody in dot-blot analysis. These results indicate that OFE contains BMPs that contribute to the induction of biomineralization.

Intermittent hypoxia increases ventilation and Sa(O2) during hypoxic exercise and hypoxic chemosensitivity.

The purpose of this study was 1) to test the hypothesis that ventilation and arterial oxygen saturation (Sa(O2)) during acute hypoxia may increase during intermittent hypoxia and remain elevated for a week without hypoxic exposure and 2) to clarify whether the changes in ventilation and Sa(O2) during hypoxic exercise are correlated with the change in hypoxic chemosensitivity. Six subjects were exposed to a simulated altitude of 4,500 m altitude for 7 days (1 h/day). Oxygen uptake (VO2), expired minute ventilation (VE), and Sa(O2) were measured during maximal and submaximal exercise at 432 Torr before (Pre), after intermittent hypoxia (Post), and again after a week at sea level (De). Hypoxic ventilatory response (HVR) was also determined. At both Post and De, significant increases from Pre were found in HVR at rest and in ventilatory equivalent for O2 (VE/VO2) and Sa(O2) during submaximal exercise. There were significant correlations among the changes in HVR at rest and in VE/VO2 and Sa(O2) during hypoxic exercise during intermittent hypoxia. We conclude that 1 wk of daily exposure to 1 h of hypoxia significantly improved oxygenation in exercise during subsequent acute hypoxic exposures up to 1 wk after the conditioning, presumably caused by the enhanced hypoxic ventilatory chemosensitivity.

Cardiovascular response to hypoxia after endurance training at altitude and sea level and after detraining.

The purpose of this study was to elucidate 1) the effects of endurance exercise training during hypoxia or normoxia and of detraining on ventilatory and cardiovascular responses to progressive isocapnic hypoxia and 2) whether the change in the cardiovascular response to hypoxia is correlated to changes in the hypoxic ventilatory response (HVR) after training and detraining. Seven men (altitude group) performed endurance training using a cycle ergometer in a hypobaric chamber of simulated 4,500 m, whereas the other seven men (sea-level group) trained at sea level (K. Katayama, Y. Sato, Y. Morotome, N. Shima, K. Ishida, S. Mori, and M. Miyamura. J. Appl. Physiol. 86: 1805-1811, 1999). The HVR, systolic and diastolic blood pressure responses (DeltaSBP/DeltaSa(O(2)), DeltaDBP/DeltaSa(O(2))), and heart rate response (DeltaHR/DeltaSa(O(2)); Sa(O(2)) is arterial oxygen saturation) to progressive isocapnic hypoxia were measured before and after training and during detraining. DeltaSBP/DeltaSa(O(2)) increased significantly in the altitude group and decreased significantly in the sea-level group after training. The changed DeltaSBP/DeltaSa(O(2)) in both groups was restored during 2 wk of detraining, as were the changes in HVR, whereas there were no changes in the DeltaDBP/DeltaSa(O(2)) and DeltaHR/DeltaSa(O(2)) throughout the experimental period. The changes in DeltaSBP/DeltaSa(O(2)) after training and detraining were significantly correlated with those in HVR. These results suggest that DeltaSBP/DeltaSa(O(2)) to progressive isocapnic hypoxia is variable after endurance training during hypoxia and normoxia and after detraining, as is HVR, but DeltaDBP/DeltaSa(O(2)) and DeltaHR/DeltaSa(O(2)) are not. It also suggests that there is an interaction between the changes in DeltaSBP/DeltaSa(O(2)) and HVR after endurance training or detraining.

Reproducibility of the oxygen uptake efficiency slope in normal healthy subjects.

BACKGROUND: To elucidate the intertest agreement of the oxygen uptake efficiency slope (OUES) in comparison with those of the maximal oxygen uptake (VO2max) and the ventilatory anaerobic threshold (VAT). EXPERIMENTAL DESIGN: A comparative study. SETTING: Institutional practice. A total of 19 healthy volunteers underwent two sessions of maximal exercise testing with an interval of no more than 7 days. The testing was conducted on a cycle ergometer with the work rate increased by either 20, 30, or 40 Watts (W)/min so that the subject would reach exhaustion within 9 to 12 min of exercise. VAT was defined as the level of oxygen uptake (VO2) at which either an increase in the ventilatory equivalent for oxygen without a concomitant increase in the ventilatory equivalent for carbon dioxide or a change in the slope of the linear relationship between carbon-dioxide production (VCO2) and VO2 occurred. OUES was determined by the following equation: VO2 = a log VE + b, where VE was minute ventilation and "a" was the OUES. Intertest reproducibility was assessed by coefficient of repeatability (COR). RESULTS: The intertest reproducibility of VO2max and OUES were excellent (COR = 570 ml/min [16%] and 740 [20%], respectively). VAT showed poor agreement between the two tests (COR = 650 ml/min [31%]). CONCLUSIONS: Results show that OUES is reproducible and reliable, supporting the clinical usefulness of this index.

Ventilatory chemosensitive adaptations to intermittent hypoxic exposure with endurance training and detraining.

The present study was performed to clarify the effects of intermittent exposure to an altitude of 4,500 m with endurance training and detraining on ventilatory chemosensitivity. Seven subjects (sea-level group) trained at sea level at 70% maximal oxygen uptake (VO2 max) for 30 min/day, 5 days/wk for 2 wk, whereas the other seven subjects (altitude group) trained at the same relative intensity (70% altitude VO2 max) in a hypobaric chamber. VO2 max, hypoxic ventilatory response (HVR), and hypercapnic ventilatory response, as an index of central hypercapnic chemosensitivity (HCVR) and as an index of peripheral chemosensitivity (HCVRSB), were measured. In both groups VO2 max increased significantly after training, and a significant loss of VO2 max occurred during 2 wk of detraining. HVR tended to increase in the altitude group but not significantly, whereas it decreased significantly in the sea-level group after training. HCVR and HCVRSB did not change in each group. After detraining, HVR returned to the pretraining level in both groups. These results suggest that ventilatory chemosensitivity to hypoxia is more variable by endurance training and detraining than that to hypercapnia.

Gene expression of growth and differentiation factors-5, -6, and -7 in developing bovine tooth at the root forming stage.

Growth and differentiation factors (GDF)-5, -6, and -7 are members of the bone morphogenetic protein (BMP) family. Previous studies suggest their importance in bone development and in tendon/ligament morphogenesis. The cells of the dental attachment apparatus, cementum, periodontal ligament, and alveolar bone proper are derived from the dental follicle proper. In this study, we investigated the expression of GDF-5, -6, and -7 genes in tissues of the bovine incisor tooth germ at the root forming stage. The results demonstrate distinct expression of GDFs in both the dental follicle and the odontoblast layer. While GDF-5 and -6 mRNAs were expressed in both the dental follicle and the odontoblast layer, GDF-7 mRNA expression was detected only in the dental follicle. These results indicate that GDFs, expressed in the bovine tooth germ including the dental follicle, may be potent regulatory molecules in the development of the dental attachment apparatus.