[The technique characteristic and clinical application of HX straight-wire appliance].

HX straight-wire appliance (HX-SWA) is designed and adjusted by doctors of Orthodontic Department, West China College of Stomatology, Sichuan University. It is a set of appliance that is constructed according to normal occlusion features of the Chinese people, including the prescription of tip, torque, in/out, counter-tip, counter-rotation and overcorrection in brackets and buccal tubes. Some ingredients of the appliance are different from that of the most popular Roth straight-wire appliance in the world wide. Over a period of the last 10 years, doctors in our hospital kept on summarizing treatment experience with HX-SWA, which will help more doctors promote practice and results in orthodontic clinic.

Centrifugal forces within usually-used magnitude elicited a transitory and reversible change in proliferation and gene expression of osteoblastic cells UMR-106.

Centrifugation is an important step in biochemical and molecular biological researches. But the effects of centrifugal stress on cells are still unclear. In this study, osteoblastic cells UMR-106 were subjected to a moderate centrifugal stress at 209 x g for 10 min. Then the cell proliferation and gene transcription after centrifugation were analyzed with flow cytometry and Real-time RT-PCR techniques, respectively. The result showed that the cell proliferation and mRNA expression of Runx2/Cbfa1, Collagen I and osteocalcin changed shortly after centrifugal loading, but recovered to pre-load levels within 24 h. A dose-response study of exposure cells to centrifugal force at 209, 253 and 301 x g showed that the centrifugal forces within usually-used range can rapidly influenced the mRNA expression of the osteoblast-specific genes, but no statistical differences were found among the three centrifugal magnitudes. And the fast regulation in the investigated genes was proved to be related to increased c-fos mRNA levels and subsequent activation of RTK and integrity of cytoskeleton construction. The result showed that the osteoblastic cells displayed a fast auto-regulation to usually-used centrifugal stress through multiple signal pathways.

[Changes of the acetylcholine receptor on rat's superficial masseter muscles after functional mandibular advancement].

OBJECTIVE: The purpose of this study was to evaluate the maximum binding (Bmax) and affinity Kd value changes of acetylcholine receptor (n-AchR) on rat's superficial masseter muscles after functional mandibular advancement. METHODS: Forty 5-week-old male SD rats were randomly divided into experimental and control groups. The mimic functional appliances were only used in experimental groups, and the rats of two groups were killed after 1, 3, 7, 14 days. Radio-ligand binding assay (RBA) was applied to determine the Bmax and Kd value of n-AchR on the superficial masseter muscles. RESULTS: The Bmax of n-AchR in experimental groups was higher than that in the control groups at all time points, and the differences had statistical significance. The Kd value of n-AchR was higher in experimental group than that in control groups in 1, 3, 7 days, while Kd value in experimental group of n-AchR was lower than that in control groups in 14 days. The differences had statistical significance in 7 and 14 days, but the differences had no statistical significance in 1 and 3 days. CONCLUSION: The functional orthopedics can increase the Bmax and affinity of n-AchR on rapid growing rat's superficial masseter muscles.

Effects of tensile stress on the alpha1 nicotinic acetylcholine receptor expression in maxillofacial skeletal myocytes.

This study was to investigate the alterations of the alpha1 nicotinic acetylcholine receptor (nAChR) levels under tensile stress stimulation in maxillofacial skeletal myocytes. The skeletal muscle satellite cells from two to three days post-natal BALB/C mice's maxillofacial muscle were collected for primary cell culture. The second passage cells in the loaded groups were subjected to cyclic tensile stress (0.5 Hz, 2000 micro strain) produced by a four point bending system for 2, 4, 6, 8, or 12 h. In the control groups, cells were cultured on similar plates and kept in the same incubator without mechanical stress loading. The examination of nAChR alpha1 receptor expression was performed by receptor binding of [125I] a-bungarotoxin. The nAChR alpha1 mRNA transcript level was analyzed by semi-quantitative RT-PCR. The result showed that the nAChR alpha1 receptor expression was elevated significantly in stress-stimulated group (P < 0.05). An increase of nAChR alpha1 in mRNA transcript level was also observed in stress groups as compared with controls (P < 0.05). It is concluded that nAChR was a possible molecular mechanism which might play an important role in mechanotransduction of tensile stress loading on maxillofacial skeletal myocytes.

The role of extracellular matrix, integrins, and cytoskeleton in mechanotransduction of centrifugal loading.

The study was aimed to investigate the role of the "extracellular matrix (ECM)-integrins-cytoskeleton" signal pathway in mechanotransduction of centrifugal loading. MG-63 osteoblasts were exposed to centrifugal loading at 209xg for 10 min. Uncentrifuged cells and centrifuged cells that have been trypsinized and suspended in liquors were designed as control. The changes in F-actin and alpha-actin cytoskeleton, gene transcription of ECM components, and integrins expression were analyzed by LSCM, Real-Time RT-PCR and FCM, respectively. A temporary and fast reversible change was observed in F-actin and alpha-actin cytoskeleton. And the change was paralleled with the fast autoregulation in gene transcription of ECM components of fibronection, osteopontin and Collagen I, and integrins expression of both alpha2 and beta1 subunits. The result suggested that cytoskeleton was a possible mechanical sensor to centrifugal stimuli, and the cytoskeleton regulation to centrifugal loading was in an ECM-dependent and integrin-mediated manner.

A novel missense mutation in the paired domain of human PAX9 causes oligodontia.

PAX9 and MSX1 are transcription factors that play essential roles in craniofacial and limb development. In humans, mutations in both genes are associated with nonsyndromic and syndromic oligodontia, respectively. We screened one family with nonsyndromic oligodontia for mutations in PAX9 and MSX1. Single stranded conformational polymorphism (SSCP) analysis and sequencing revealed a novel heterozygous C139T transition in PAX9 in the affected members of the family. There were no mutations detected in the entire coding sequence of MSX1. The C139T mutation, predicted to result in the substitution of an arginine by a tryptophan (R47W) in the N-terminal subdomain, affected conserved residues in the PAX9 paired domain. To elucidate the pathogenic mechanism producing oligodontia phenotype caused by this mutation, we analyzed the binding of wild-type and mutant PAX9 paired domain protein to double-stranded DNA targets. The R47W mutation dramatically reduced DNA binding suggesting that the mutant protein with consequent haploinsufficiency results in a clinical phenotype.

NF-kappaB responds to mechanical strains in osteoblast-like cells, and lighter strains create an NF-kappaB response more readily.

This study was to examine the early responses of nuclear factor kappa B (NF-kappaB) to mechanical strains in MG-63. MG-63 cells were subjected to cyclic uniaxial compressive or tensile strain, produced by a four-point bending system, at 1000 microstrain or 4000 microstrain for 5 min, 15 min, 30 min and 1h, respectively. Control cells received the same treatment with no mechanical stress loading. Expression of NF-kappaB (p60) was measured by Western blotting. NF-kappaB responded rapidly to mechanical stimuli in MG-63 cells. NF-kappaB was activated by cyclic uniaxial stretch at 1000 microstrain while it was restrained under a compressive strain environment at 1000 microstrain (P<0.001). The effects reversed for tension and compression at 4000 microstrain (P<0.001). Furthermore, strains at 1000 microstrain affected NF-kappaB expression much easier than those at 4000 microstrain. This indicates that there may be different responding mechanisms or mechanotransduction pathways for different mechanical stimuli.

Osteoblast cytoskeletal modulation in response to compressive stress at physiological levels.

Biomechanical force is one of the major epigenetic factors that determine the form and differentiation of skeletal tissues. In this study, osteoblastic cells UMR-106 were exposed to compressive forces at 1000 mustrain and 4000 mustrain via a four-point bending system, and analyzed by MTT and LSCM techniques. Cell proliferation activity decreased shortly after loading but recovered to normal levels within 24 h. And the cytoskeleton depolymerized at first, but then gradually repolymerized. To find out the role of cytoskeleton in mechanotransduction, we examined the relationship between cytoskeleton construction and c-fos expression. A transient stress-induced upregulation in c-fos mRNA and c-Fos protein was discovered when cells were exposed to physiological forces. And the upregulation in c-fos expression was blocked by cytochalasin D (Depolymerizing agent of microfilament). It gave clues that the organization of cytoskeleton was an important link in transcriptional control in response to low-mechanical stimulation.

Effects of different magnitudes of cyclic stretch on Na+-K+-ATPase in skeletal muscle cells in vitro.

The Na(+)-K(+)-ATPase, which plays a major role in modulation of skeletal muscle excitability and contractility, is one of the marker enzymes that senses the mechanical strain and adapts to the stimuli. Although many papers had been published on the effects of mechanical stress on Na(+)-K(+)-ATPase in aortic smooth muscle cells, little was known about the effects of different magnitudes of mechanical stretch on Na(+)-K(+)-ATPase in skeletal muscle cells. In the present study, we determined the effect of different magnitudes(6%, 12%, or 25% elongation) of cyclic stretch on the activity of the Na(+)-K(+)-ATPase and investigated possible mechanisms that might be involved in the action of stretch. The results showed the application of different magnitudes of cyclic stretch induced a magnitude-dependent increase of Na(+)-K(+)-ATPase activity in cultured skeletal muscle cells. Furthermore, inhibition of ionic fluxes through SACs prevented the action of stretch on Na(+)-K(+)-ATPase activity. The stretch-induced increase in Na(+)-K(+)-ATPase activity was not blocked by Actinomycin D. No significant changes in mRNA and total cell protein levels of Na(+)-K(+)-ATPase were detected after stretched continuous for 24 h. However, cyclic stretch increased cell surface expression of Na(+)-K(+)-ATPase alpha(1)- and alpha(2)-subunit proteins by 1.3- and 1.75-fold, respectively, and the increases in Na(+)-K(+)-ATPase activity and cell surface expression were abolished by LY-294002. These data indicated that cyclic stretch induced a "magnitude-dependent" increase of Na(+)-K(+)-ATPase activity in cultured skeletal muscle cells in vitro. The upregulation involved translocation of Na(+)-K(+)-ATPase alpha(1)- and alpha(2)-subunits to plasma membrane, not increased gene transcription. These results suggested a novel nontranscriptional mechanism for regulation of Na(+)-K(+)-ATPase in skeletal muscle cells by cyclic stretch.

Cyclic stretch translocates the alpha2-subunit of the Na pump to plasma membrane in skeletal muscle cells in vitro.

The Na+-K+-ATPase and its regulation is important for maintaining membrane potential and transmembrane Na(+) gradient in all skeletal muscle cells and thus is essential for cell survival and function. In our previous study, cyclic stretch activated the Na pump in cultured skeletal muscle cells. Presently, we investigated whether this stimulation was the result of translocation of Na+-K+-ATPase from endosomes to the plasma membrane, and also evaluated the role of phosphatidylinositol 3-kinase (PI 3-kinase), the activation of which initiated vesicular trafficking and targeting of proteins to specific cell compartments. Skeletal muscle cells were stretched at 25% elongation continuous for 24h using the Flexercell Strain Unit. The plasma membrane and endosome fractions were isolated and Western blotted to localize the Na+-K+-ATPase alpha1- and alpha2-subunit protein. The results showed stretch increased Na+-K+-ATPase alpha1- and alpha2-subunit protein expression in plasma membrane fractions and decreased it in endosomes. The alpha2-subunit had a more dynamic response to mechanical stretch. PI 3-kinase inhibitors (LY294002) blocked the stretch-induced translocation of the Na+-K+-ATPase alpha2-subunit, while LY294002 had no effect on the transfer of alpha1-subunit. We concluded that cyclic stretch mainly stimulated the translocation of the alpha2-subunit of Na+-K+-ATPase from endosomes to the plasma membrane via a PI 3-kinase-dependent mechanism in cultured skeletal muscle cells in vitro, which in turn increased the activity of the Na pump.

Early responses of osteoblast-like cells to different mechanical signals through various signaling pathways.

This study was to examine the effects of mechanical stimuli alone and coupled with some inhibitors of related signaling pathways on early cellular responses. MG-63 cells were subjected to cyclic uniaxial compressive or tensile strain at 4000 microstrain, produced by four-point bending system. The effects of mechanical strains alone and coupled with inhibitors of microfilament and receptor tyrosine kinase (RTK) on activation of extracellular signal-regulated kinase (ERK), c-fos mRNA, and c-Fos protein were examined. ERK could be activated by mechanical stimuli in 5 min and so could be c-fos mRNA and c-Fos protein in 30 min. Tensile stress had a more obvious effect than compressive one. Early cellular responses were connected with cytoskeleton and RTK pathways during the transduction of mechanical signals. The property of strains was an influential factor for the activation effects.

WITHDRAWN: NF-kappaB is involved in early cellular response to mechanical stimuli in osteoblast-like cells in vitro.

Ahead of Print article withdrawn by publisher.

[Substance P regulates function of osteoclasts via neurokinin-1 receptor].

OBJECTIVE: To investigate the effects of substance P on cultured rat osteoclasts. METHODS: Neurokinin-1 (NK1) receptor expression in osteoclasts was examined by immunohitochemical method, and changes of bone resorption activity caused by substance P and NK1 receptor antagonists were detected by pit formation assay. RESULTS: Immunoreactivity for NK1 receptor was distributed in the cytoplasm of osteoclasts. The average of pit formation areas significantly increased with addition of substance P (10(-7)-10(-4) mol/L) (P < 0.05), but the number of pitformations did not change (P > 0.05). NK1 receptor antagonists inhibited the enhancement of the bone resorption by substance P addition. CONCLUSION: The findings suggested that substance P may stimulate osteoclasts and result in bone resorption by the mediation of NK1 receptor.

[Effects of programmed cell death on human dental follicle cells and changes of programmed cell death under different hydrostatic pressures].

OBJECTIVE: Tooth eruption requires the presence of the dental follicle (DF) around the unerupted tooth. This study is to investigate programmed cell death on human dental follicle cells and changes of programmed cell death under different hydrostatic pressures: 0, 50 and 100 kPa. METHODS: Human dental follicles from third mandibular molars were surgically removed from adolescents who need for orthodontics treatment after informed content, then trypsinized and cultured. Human dental follicle cells were divided into three groups according to different hydrostatic pressures: 0, 50 and 100 kPa and their programmed cell death were labeled by using TdT-medi-ated-dUTP nick and labeling (TUNEL). RESULTS: Dental follicle cells cultured were elongate shape and exhibited fibroblastic characteristics. Compared with 0 kPa, programmed cell death cells on human dental follicle cells were increased 0.23% and 31.65% under 50 kPa and 100 kPa hydrostatic pressures respectively. 100 kPa group increased significantly (P < 0.01). CONCLUSION: It suggested that programmed cell death occured in human dental follicle cells cultured in vitro and was influenced by different hydrostatic pressures. Hydrostatic pressure may improve tooth erup-tion through dental follicle.

[The clinic skill in fixed appliance based on characteristics of Chinese normal occlusion].

OBJECTIVE: To study the bracket placement and arch wire bending based on ethnic differences and individual differences of normal occlusion. METHODS: The prominence, tip, torque, upper first molar offset of crown and arch form between Chinese and Caucasian normal occlusion were compared. RESULTS: The results showed the ethnic differences of prominence, tip, torque, upper first molar offset of crown and arch form between Chinese and Caucasian normal occlusion. The placement of bracket was influenced by the crown morphology. CONCLUSION: The adjustments of the bracket placement and arch wire bending with Edgewise and pre-adjusted appliance are necessary to adapt to ethnic difference and individual difference.

[Cephalometric evaluation of the effects of pendulum appliance on various vertical growth patterns].

OBJECTIVE: The aim of this study was to investigate the effects of the pendulum appliance in treating dental Class II patients with various vertical growth patterns. METHODS: The samples (n = 30) were divided into three groups equally based on their FMA. Pretreatment and post-treatment cephalometric radiographs were taken to measure the changes. RESULTS: The amount of upper molar distalization in the low-angle group was the fewest, and that in the high-angle group was the most. Upper molars had been intruded insignificantly. The amount of anchorage loss at the first premolars and overjet increased at incisors was different in the three groups. The biggest change happened in the low-angle group, and the smallest in the high-angle group. CONCLUSION: The results of this study showed that pendulum appliance could move the upper molars distally in a short period of time. The upper molars in different groups were intruded insignificantly. Pendulum appliance could be used to move the upper molars distally in high-angle cases.

[The effects of growth hormone on rabbit's mandibular condylar chondrocytes proliferation and secretion in vitro].

OBJECTIVE: To study the effect of growth hormone (GH) on the proliferation and type II collagen secretion of chondrocytes of mandibular condyle in rabbit in vitro. METHODS: Flow cytometry (FCM) and immunohistochemical technique were employed to observe the possible changes. RESULTS: (1) The exogenic GH can enhance the proliferation and synthesis of DNA of the chondrocytes of mandibular condyle in rabbit in vitro. The suitable concentration of GH is 10 microg/ml. The synthesis of DNA reaches the highest level after 12 hours, while the proliferation index (PI) hits the highest after 24 hours. (2) GH (10 microg/ml) can stimulate the secretion of type II collagen of the chondrocytes. CONCLUSION: The exogenic GH can enhance the proliferation, the synthesis of DNA and the secretion of type II collagen of the chondrocytes of mandibular condyle in rabbit in vitro.

[Changes of sarco(endo)plasmic reticulum Ca2+-Mg2+-ATPase activity of cultured myoblast induced by cyclic stretch].

OBJECTIVE: To evaluate the effect of stretch on sarco(endo)plasmic reticulum Ca2+-Mg2+ ATPases activity and mRNA level and study the remodeling reaction of muscle in a variety of mechanical environments. METHODS: Myoblast from maxillofacial skeletal muscle of one-week-old male Sprague-Dawley rat was cultured and stretched cyclicly using a four-point bend device. Inorganic Phosphorus test was used to compare the activity of Ca2+-Mg2+ ATPases of myoblast before and after stretch. RT-PCR was also used to observe the Ca2+-Mg2+ ATPases mRNA level. RESULTS: The activity of Ca2+-Mg2+ ATPases of myoblast down regulated significantly in 4 hours. During the period of 8 hours to 24 hours, up-regulation followed then returned to control level at the 48 hour point. RT-PCR showed that Ca2+-Mg2+ ATPases mRNA level were elevated by stretch, particularly at 2 hour and 48 hour point. CONCLUSION: The results suggested a transcriptional control of Ca2+-Mg2+ ATPases activity was involved in the muscle remodeling process induced by stretch.

[The effect of fluid shear stress on the NO synthesis of rat osteoblast-like cells].

This study sought to elucidate the function of NO during the signal transduction wherein fluid shear stress regulates the proliferation and differentiation of osteoblast cells. The isolated rat osteoblast-like cells were exposed to fluid shear stress 12 dyn/cm2 for 5, 10, 15, 30, 60 and 120 min respectively with the use of a flow chamber. The NO release was examined. After the exposure to fluid shear stress, the NO synthesis of rat primary osteoblast-like cells increased significantly (P<0.05) when compared with the control. After 60 minutes of exposure, the release of NO began to increase significantly (P<0.05), but no significant increase as such was seen in the control (P>0.05). NO synthesis may be one of the signal transduction pathways which transduce the fluid shear stress into osteoblast cells. In early stage, it may be induced by cNOS and in late stage by iNOS.