Rapamycin alleviates irradiation-induced parotid injury by enhancing the whole gland homeostasis.

OBJECTIVES: Salivary gland injury is one of the most common complications of radiotherapy in head-and-neck cancers. This study investigated the mechanism by which rapamycin prevents irradiation (IR)-induced injury in the parotid glands. MATERIALS AND METHODS: Miniature pigs either received (a) no treatment (NT), (b) IR in the right parotid gland for 5 consecutive days (IR), or intraperitoneal administration of rapamycin (Rap) 1 h prior to IR (IR + Rap). Tissues were collected at three distinct time points (24 h, 4 weeks, and 16 weeks) after IR. Histological analyses, western blot, and real-time reverse transcriptase-polymerase chain reaction were performed to explore the mechanisms of IR-induced injury in the parotid gland. RESULTS: Rapamycin treatment maintained parotid salivary flow 16 weeks post-IR, preserved the number of acinar cells, and reduced parotid tissue fibrosis, as well as reduced apoptosis levels, decreased cleaved caspase-3 expression, and increased the Bcl-2/Bax ratio in the parotid glands. Autophagy marker LC3B was upregulated by rapamycin after IR, while P62 expression was downregulated. Rapamycin reduced the expression of pro-inflammatory factors and the mesenchymal tissue fibrosis following IR. CONCLUSIONS: Rapamycin maintains gland homeostasis after IR by decreasing apoptosis, reducing the expression of pro-inflammatory factors, and enhancing autophagy.

Type 2 diabetes aggravates periodontitis-induced pathological changes in the dental pulp.

OBJECTIVES: The inner mechanism of how diabetes affects dental pulp of patients with periodontitis has seldom been reported. We collected clinical samples and explored the influence of diabetes and periodontitis on the pathological change of dental pulp. METHODS: Dental pulp from healthy individuals and patients with periodontitis with or without diabetes were collected based on strict inclusion and exclusion criteria. Dental pulp was morphologically observed; advanced glycation end products (AGEs) and lectin-like oxidized low-density lipoprotein receptor-1 (LOX1) were examined. Oxidative stress (OS), inflammatory indices, and apoptotic levels were assessed. RESULTS: Morphologically, fibrous structure in the dental pulp of patients with diabetic periodontitis (DP) group was sparse and disordered, and the blood vessel wall was thickened. Diabetes related indexes as AGEs and LOX1 were upregulated. Superoxide dismutase 2 expression was decreased, and OS level was increased. Matrix metalloproteinase 3 and other relevant proinflammatory cytokines levels were increased. The elevated OS and inflammation contributed to upregulation of apoptotic levels in DP group. CONCLUSIONS: Diabetes aggravates the pathological changes in the dental pulp of periodontitis patients possibly due to upregulated AGEs and LOX1. Our results highlight the importance of early oral intervention in patients with DP.

Transient Activation of Hedgehog Signaling Inhibits Cellular Senescence and Inflammation in Radiated Swine Salivary Glands through Preserving Resident Macrophages.

Salivary gland function is commonly and irreversibly damaged by radiation therapy for head and neck cancer. This damage greatly decreases the patient's quality of life and is difficult to remedy. Previously, we found that the transient activation of Hedgehog signaling alleviated salivary hypofunction after radiation in both mouse and pig models through the inhibition of radiation-induced cellular senescence that is mediated by resident macrophages in mouse submandibular glands. Here we report that in swine parotid glands sharing many features with humans, the Hedgehog receptor PTCH1 is mainly expressed in macrophages, and levels of PTCH1 and multiple macrophage markers are significantly decreased by radiation but recovered by transient Hedgehog activation. These parotid macrophages mainly express the M2 macrophage marker ARG1, while radiation promotes expression of pro-inflammatory cytokine that is reversed by transient Hedgehog activation. Hedgehog activation likely preserves parotid macrophages after radiation through inhibition of P53 signaling and consequent cellular senescence. Consistently, VEGF, an essential anti-senescence cytokine downstream of Hedgehog signaling, is significantly decreased by radiation but recovered by transient Hedgehog activation. These findings indicate that in the clinically-relevant swine model, transient Hedgehog activation restores the function of irradiated salivary glands through the recovery of resident macrophages and the consequent inhibition of cellular senescence and inflammation.

Sphingosine-1-phosphate alleviates irradiation-induced parotid injury in a miniature pig model.

OBJECTIVES: Our aim was to verify the alleviation effect of sphingosine-1-phosphate (S1P) in a miniature pig model. MATERIAL AND METHODS: Thirty male miniature pigs were randomly separated into 10 groups in our experiment. We administered S1P through the parotid duct in a retrograde fashion 2 hr before irradiation (IR). The salivary flow rate and blood flow rate were tested 20 weeks after IR. The apoptotic level was checked at 12, 24 hr and 7 days post-IR. RESULTS: Twenty weeks after IR, the salivary flow rate of the IR-side parotid gland in IR + S1P group can be maintained at about 40% of the non-IR side, while only 20% was maintained in the IR group. The blood flow rate and microvascular density were significantly higher in the IR + S1P group than in the IR group. The apoptotic level and cleaved caspase-3 expression were downregulated in IR + S1P group, and the ratio of Bcl-2/Bax was increased. The blood flow rate and CD31 level were significantly restored at 12, 24 hr and 7 days post-IR. CONCLUSION: Sphingosine-1-phosphate may partially alleviate IR-induced parotid dysfunction by decreasing apoptosis of microvascular endothelial cells and maintaining the blood flow rate.

Development of a New Index to Assess the Difficulty Level of Surgical Removal of Impacted Mandibular Third Molars in an Asian Population.

PURPOSE: This study aimed to develop a straightforward and accurate index of impacted third molar removal difficulty through analyzing various factors to assess the difficulty level of impacted mandibular third molar (IMTM) extraction. MATERIALS AND METHODS: This prospective cohort study included 203 patients who required IMTM extraction. All patients were selected using the preset selection criteria. The present study assessed operation difficulty with operating time. A mathematical model and regression analysis were performed to explore 6 main factors (age, number of roots, degree of bone impaction, shape of roots, and impaction angle and its relation). Appropriate correction coefficients were obtained to formulate a new IMTM removal difficulty predictive index. Consistency of the κ value was checked to evaluate performance. RESULTS: Degree of bone impaction had the highest correlation coefficient (0.576), followed by shape of roots (0.359), and the lowest correlation coefficient was for number of roots. The Pederson index for these 203 patients showed that 75, 76, and 52 patients had low, moderate, and high difficulty levels, respectively, whereas the new index categorized 78, 85, and 40 patients as having low, moderate, and high difficulty. Comparison of the Pederson index and new index with operating time showed κ agreements of 65.30 and 77.9% (P < .01), suggesting that the prediction results of the new index are more objective and accurate. CONCLUSION: The newly proposed index is straightforward and efficient and exhibited promising results in κ agreement. Because of its straightforward nature, it is better suited for Chinese public hospitals with a large volume of patients who require alveolar surgery. The detection of predictor variables could be useful for graduate students, professionals, and general dental practitioners contemplating IMTM removal to assess the difficulty level of IMTM extraction.

SFRP2 enhances the osteogenic differentiation of apical papilla stem cells by antagonizing the canonical WNT pathway.

BACKGROUND: Exploring the molecular mechanisms underlying directed differentiation is helpful in the development of clinical applications of mesenchymal stem cells (MSCs). Our previous study on dental tissue-derived MSCs demonstrated that secreted frizzled-related protein 2 (SFRP2), a Wnt inhibitor, could enhance osteogenic differentiation in stem cells from the apical papilla (SCAPs). However, how SFRP2 promotes osteogenic differentiation of dental tissue-derived MSCs remains unclear. In this study, we used SCAPs to investigate the underlying mechanisms. METHODS: SCAPs were isolated from the apical papilla of immature third molars. Western blot and real-time RT-PCR were applied to detect the expression of ß-catenin and Wnt target genes. Alizarin Red staining, quantitative calcium analysis, transwell cultures and in vivo transplantation experiments were used to study the osteogenic differentiation potential of SCAPs. RESULTS: SFRP2 inhibited canonical Wnt signaling by enhancing phosphorylation and decreasing the expression of nuclear ß-catenin in vitro and in vivo. In addition, the target genes of the Wnt signaling pathway, AXIN2 (axin-related protein 2) and MMP7 (matrix metalloproteinase-7), were downregulated by SFRP2. WNT1 inhibited the osteogenic differentiation potential of SCAPs. SFRP2 could rescue this WNT1-impaired osteogenic differentiation potential. CONCLUSIONS: The results suggest that SFRP2 could bind to locally present Wnt ligands and alter the balance of intracellular Wnt signaling to antagonize the canonical Wnt pathway in SCAPs. This elucidates the molecular mechanism underlying the SFRP2-mediated directed differentiation of SCAPs and indicates potential target genes for improving dental tissue regeneration.

Negative Regulation of LINC01013 by METTL3 and YTHDF2 Enhances the Osteogenic Differentiation of Senescent Pre-Osteoblast Cells Induced by Hydrogen Peroxide.

Senescent pre-osteoblasts have a reduced ability to differentiate, which leads to a reduction in bone formation. It is critical to identify the keys that regulate the differentiation fate of senescent pre-osteoblasts. LINC01013 has an essential role in cell stemness, differentiation, and senescence regulation. This study aims to examine the role and mechanism of LINC01013 in regulating osteogenic differentiation in senescent human embryonic osteoblast cell line (hFOB1.19) cells induced by hydrogen peroxide (H2O2). The results show that LINC01013 decreased alkaline phosphatase activity, mineralization of hFOB1.19 cells in vitro, and the expression of collagen II, osteocalcin, and bone sialoprotein. LINC01013 knockdown enhances the osteogenesis of hFOB1.19 cells and rescues osteogenic differentiation impaired by H2O2. METTL3 negatively regulates LINC01013 expression, enhancing hFOB1.19 cells' osteogenesis in vitro and in vivo. METTL3 overexpression can enhance hFOB1.19 cells' osteogenic differentiation impaired by H2O2. YTHDF2 promotes LINC01013 decay, facilitating osteogenic differentiation. YTHDF2 overexpression rescues hFOB1.19 cells osteogenic differentiation impaired by H2O2. Taken together, METTL3 upregulates osteogenic differentiation by inhibiting LINC01013, and YTHDF2 accelerates LINC01013 degradation, reducing its inhibitory effect. This study highlights LINC01013 as a key regulator in the fate switching process of senescent hFOB1.19 cells, impacting osteogenic differentiation.

Shh signaling, negatively regulated by BMP signaling, inhibits the osteo/dentinogenic differentiation potentials of mesenchymal stem cells from apical papilla.

Mesenchymal stem cells (MSCs) derived from dental tissues show promise for use in tooth-related tissue regeneration, but the molecular mechanisms underlying their directed differentiation remain unclear, limiting their usefulness. Sonic Hedgehog (Shh) signaling is a major signaling pathway that regulates cell differentiation and osteogenesis. We found that when Shh signaling was activated by human recombinant SHH-N protein or by overexpression of active mutant M2-Smoothened (SMO) in stem cells from apical papilla (SCAPs), GLI1, a key downstream transcription factor and a marker of Shh signaling, was upregulated. Subsequently, in vitro osteo/dentinogenic differentiation and in vivo osteogenesis were inhibited in SCAPs. Moreover, the expression of GLI1 and SMO were downregulated by BMP signaling while osteo/dentinogenic differentiation in SCAPs was upregulated. These results provide insights into the role of Shh signaling in the directed differentiation of MSCs derived from dental tissues and suggest possible target genes for optimizing the use of stem cells of dental origin for tissue regeneration applications.

Decellularized rat submandibular gland as an alternative scaffold for dental pulp regeneration.

Introduction: Decellularized extracellular matrix has been recognized as an optimal scaffold for dental pulp regeneration. However, the limited amount of native dental pulp tissue restricts its clinical applications. The submandibular gland shares some basic extracellular matrix components and characteristics with dental pulp. However, whether decellularized submandibular gland extracellular matrix (DSMG) can be used as an alternative scaffold for dental pulp regenerative medicine is unclear. Methods: Thus, we successfully decellularized the whole rat submandibular gland and human dental pulp, and then conducted in vitro and in vivo studies to compare the properties of these two scaffolds for dental pulp regeneration. Results: Our results showed that extracellular matrix of the submandibular gland had great similarities in structure and composition with that of dental pulp. Furthermore, it was confirmed that the DSMG could support adhesion and proliferation of dental pulp stem cells in vitro. In vivo findings revealed that implanted cell-seeded DSMG formed a vascularized dental pulp-like tissue and expressed markers involved in dentinogenesis and angiogenesis. Discussion: In summary, we introduced a novel accessible biological scaffold and validated its effectiveness as an extracellular matrix-based tissue engineering scaffold for dental pulp regenerative therapy.

Spatiotemporal expression of fibroblast growth factor 4 and 10 during the morphogenesis of deciduous molars in miniature pigs.

OBJECTIVE: Fibroblast growth factors (FGFs) play pivotal roles in mediating interactions between dental epithelium and mesenchyme throughout tooth initiation and morphogenesis. This study aimed to elucidate the roles of FGF4 and FGF10 in the regulation of tooth development. DESIGN: In this study, we investigated spatiotemporal expression patterns of FGF4 and FGF10 in the third deciduous molars (DM3) of miniature pigs at the cap, early bell, and late bell stages. Pregnant miniature pigs were obtained, and the samples were processed for histological staining. Non-radioactive in situ hybridization, immunohistochemistry, and real-time PCR were used to detect mRNA and protein expression levels of FGF4 and FGF10. RESULTS: FGF4 was expressed in the dental epithelium and mesenchyme at the cap stage. At the early bell stage, epithelial expression of FGF4 was reduced while mesenchymal expression got stronger. At the late bell stage, the FGF4 expression was restricted to the inner enamel epithelium (IEE) and differentiating odontoblasts. FGF10 was expressed intensely in both epithelium and mesenchyme at the cap stage. The expression of FGF10 was concentrated in the secondary enamel knots and surrounding mesenchyme at the early bell stage. FGF10 was weakly detected in the IEE by the late bell stage. CONCLUSIONS: Our results indicated that FGF4 and FGF10 might have partially redundant functions in regulating epithelium morphogenesis. FGF4 may be involved in regulatory signaling cascades mediating interactions between the epithelium and mesenchyme. In addition, the downregulation of FGF10 expression may be associated with the cessation of mesenchymal cell proliferation and initiation of preodontoblast polarization.

Sphingosine-1-Phosphate Alleviates Irradiation Induced Salivary Gland Hypofunction through Preserving Endothelial Cells and Resident Macrophages.

Radiotherapy for head-and-neck cancers frequently causes long-term hypofunction of salivary glands that severely compromises quality of life and is difficult to treat. Here, we studied effects and mechanisms of Sphingosine-1-phosphate (S1P), a versatile signaling sphingolipid, in preventing irreversible dry mouth caused by radiotherapy. Mouse submandibular glands (SMGs) were irradiated with or without intra-SMG S1P pretreatment. The saliva flow rate was measured following pilocarpine stimulation. The expression of genes related to S1P signaling and radiation damage was examined by flow cytometry, immunohistochemistry, quantitative RT-PCR, Western blotting, and/or single-cell RNA-sequencing. S1P pretreatment ameliorated irradiation-induced salivary dysfunction in mice through a decrease in irradiation-induced oxidative stress and consequent apoptosis and cellular senescence, which is related to the enhancement of Nrf2-regulated anti-oxidative response. In mouse SMGs, endothelial cells and resident macrophages are the major cells capable of producing S1P and expressing the pro-regenerative S1P receptor S1pr1. Both mouse SMGs and human endothelial cells are protected from irradiation damage by S1P pretreatment, likely through the S1pr1/Akt/eNOS axis. Moreover, intra-SMG-injected S1P did not affect the growth and radiosensitivity of head-and-neck cancer in a mouse model. These data indicate that S1P signaling pathway is a promising target for alleviating irradiation-induced salivary gland hypofunction.

Osseointegration of a novel dental implant in canine.

This study aimed to compare and verify the osseointegration performance of a novel implant (NI) in vivo, which could provide a useful scientific basis for the further development of NIs. Thirty-two NIs treated with hydrofluoric acid and anodization and sixteen control implants (CIs) were placed in the mandibles of 8 beagles. Micro-CT showed that the trabecular number (Tb.N) significantly increased and trabecular separation (Tb.Sp) significantly decreased in the NIs at 2 weeks. Significant differences were found in the trabecular thickness, Tb.N, Tb.Sp, bone surface/bone volume ratio, and bone volume/total volume ratio between the two groups from the 2nd-4th weeks. However, there were no significant differences between the two groups in the bone volume density at 2, 4, 8, or 12 weeks or bone-implant contact at 2 or 4 weeks, but the BIC in the CIs was higher than that in the NIs at the 8th and 12th weeks. Meanwhile, the histological staining showed a similar osseointegration process between the two groups over time. Overall, the NIs could be used as new potential implants after further improvement.

Long-term transduction of miniature pig parotid glands using serotype 2 adeno-associated viral vectors.

BACKGROUND: Previously, using an adenoviral vector, we showed that miniature pigs could provide a valuable and affordable large animal model for pre-clinical gene therapy studies to correct parotid gland radiation damage. However, adenoviral vectors lead to short-term transgene expression and, ideally, a more stable correction is required. In the present study, we examined the suitability of using a serotype 2 adeno-associated viral (AAV2) vector to mediate more stable gene transfer in the parotid glands of these animals. METHODS: Heparan sulfate proteoglycan was detected by immunohistochemistry. beta-galactosidase expression was determined histochemically. An AAV2 vector encoding human erythropoietin (hEpo) was administered via Stensen's duct. Salivary and serum hEpo levels were measured using an enzyme-linked immunosorbent assay. Serum chemistry and hematological analyses were performed and serum antibodies to hEpo were measured throughout the study. Vector distribution was determined by a quantitative polymerase chain reaction. RESULTS: Transgene expression was vector dose-dependent, with high levels of hEpo being detected for up to 32 weeks (i.e. the longest time studied). hEpo reached maximal levels during weeks 4-8, but declined to approximately 25% of these values by week 32. Haematocrits were elevated from week 2. Transduced animals exhibited low serum anti-hEpo antibodies (1 : 8-1 : 16). Vector biodistribution at animal sacrifice revealed that most copies were in the targeted parotid gland, with few being detected elsewhere. No consistent adverse changes in serum chemistry or hematology parameters were seen. CONCLUSIONS: AAV2 vectors mediate extended gene transfer to miniature pig parotid glands and should be useful for testing pre-clinical gene therapy strategies aiming to correct salivary gland radiation damage.

[Increase of saliva nitrate and nitrite level in patients with oral candidiasis].

OBJECTIVE: To observe the alterations of saliva nitrate and nitrite level in patients with oral candidiasis. METHODS: Parotid saliva and whole saliva were collected from 33 patients and 34 healthy volunteers. Concentrations of nitrate and nitrite in saliva were determined by high-performance liquid chromatography. Follow-up observation was performed on 10 patients after treatment. The data were statistically analyzed with independent-samples t test or paired-samples t test at alpha = 0.05. RESULTS: There was significant increase of the concentrations and secretion rate of parotid saliva nitrate in patient group as compared with controls: (49.70 +/- 0.50) vs (21.51 +/- 0.60) mg/L (t = 2.692, P = 0.009) and (27.71 +/- 0.50) vs (12.55 +/- 0.60) microg/min (t = 2.554, P = 0.013), respectively. Significantly increased concentrations and secretion rate of nitrate and nitrite [nitrate: (6.46 +/- 0.94) vs (1.11 +/- 0.70) mg/L (t = 3.792, P = 0.000); nitrite: (8.48 +/- 0.58) vs (3.39 +/- 0.53) mg/L (t = 2.888, P = 0.005); nitrate secretion rate: (10.57 +/- 0.91) vs (2.10 +/- 0.74) microg/min (t = 3.464, P= 0.001); nitrite secretion rate: (13.91 +/- 0.55) vs (6.42 +/- 0.58) microg/min (t = 2.397, P = 0.020)] were revealed in whole saliva of patients group. Significantly decreased nitrate and nitrite levels were also observed in patients after treatment, especially the changes of parotid saliva nitrate secretion rate [(37.50 +/- 0.50) vs (14.34 +/- 0.64) microg/min (t = 3.142, P = 0.012)], whole saliva nitrate [(14.29 +/- 1.01) vs (2.59 +/- 1.03) mg/L (t = 3.475, P = 0.007)] and whole saliva nitrate secretion rate [(25.97 +/- 0.93) vs (4.12 +/- 1.00) microg/min (t = 3.922, P = 0.003)]. CONCLUSION: The present study revealed the significant increase of salivary nitrate and nitrite level in patients with oral candidiasis is considered to be associated with the host defense reaction.

Evaluation of Parotid Salivary Glucose Level for Clinical Diagnosis and Monitoring Type 2 Diabetes Mellitus Patients.

Background. To investigate the relationships among blood glucose, mixed saliva glucose, and parotid glucose in type 2 diabetes patients and to evaluate the diagnostic and monitoring value of salivary gland glucose in patients with type 2 diabetes (type 2DM). Material and Methods. Thirty patients with type 2DM and 30 healthy age- and sex-matched individuals were included in this study. Glucose levels in unstimulated mixed saliva and in unstimulated parotid saliva were measured by the glucose oxidase peroxidase method. Results. The blood glucose and parotid salivary glucose levels in type 2DM patients were significantly higher than those in the controls (P < 0.05). The blood glucose, parotid salivary glucose, and mixed salivary glucose were 7.46 ± 1.44 mmol/L, 0.18 ± 0.19 mmol/L, and 3.17 × 10-2 ± 2.84 × 10-2 mmol/L, respectively, in the type 2DM group; the corresponding glucose levels in the control group were 5.56 ± 0.71 mmol/L, 7.70 × 10-2 ± 6.02 × 10-2 mmol/L, and 3.47 × 10-2 ± 2.79 × 10-2 mmol/L. The parotid salivary and blood glucose levels in type 2DM patients were strongly correlated; the linear regression equation for blood glucose and parotid salivary glucose was Y = 6.267X + 6.360, with r = 0.810. However, mixed salivary glucose levels were not significantly different in the type 2 diabetes group compared with the control group. Conclusion. Our results suggest that parotid salivary glucose has potential as a biomarker to monitor type 2DM and as a painless, noninvasive method for the management of type 2DM.

Structural and functional characteristics of irradiation damage to parotid glands in the miniature pig.

PURPOSE: To evaluate the effects of a solitary megadose protocol of ionizing radiation (IR) on the structure and function of the miniature pig (minipig) parotid gland. METHODS AND MATERIALS: Fourteen minipigs were subjected to either 15 or 20 Gy to one parotid gland with a linear accelerator, whereas another four minipigs served as non-IR controls. Salivary flow rates and salivary chemistries were measured pre-IR and 4 and 16 weeks post-IR. A quantitative assessment of gland weight and acinar area and detailed serum chemistry and hematologic analyses were also performed. RESULTS: Parotid flow rates decreased by approximately 50% either with 20 Gy at 4 weeks, or 15 Gy at 16 weeks post-IR. In the 20 Gy group, salivary flow rates were reduced by approximately 80% at 16 weeks post-IR. A significant decrease in salivary calcium and amylase and an increase of salivary potassium levels were found in both IR groups. There were also transient alterations in serum chemistry and hematology parameters post-IR. Parotid gland weights were significantly decreased (-50%) in the 15 and 20 Gy groups at 4 and 16 weeks post-IR. Additionally, the acinar cell area in glands of both IR groups was significantly reduced from that in control glands at both the 4 and 16 weeks time points. CONCLUSION: Structural changes in salivary gland parenchyma occurred relatively early after IR, whereas the alterations in salivary output were relatively delayed. Further, reductions in salivary flow were not proportional to acinar cell area loss. Together, these findings suggest that nonparenchymal IR damage likely contributes to IR-induced salivary hypofunction.

Histone demethylase KDM2B inhibits the chondrogenic differentiation potentials of stem cells from apical papilla.

Mesenchymal stem cells (MSCs) are a reliable resource for tissue regeneration, but the molecular mechanism underlying directed differentiation remains unclear; this has restricted potential MSC applications. Histone methylation, controlled by histone methyltransferases and demethylases, may play a key role in MSCs differentiation. Previous studies determined that KDM2B can regulate the cell proliferation and osteo/dentinogenic differentiation of MSCs. It is not known whether KDM2B is involved in the other cell lineages differentiation of MSCs. Here we used the stem cells from apical papilla (SCAPs) to study the role of KDM2B on the chondrogenic differentiation potentials in MSCs. In this study, Gain- and loss-of-function assays were applied to investigate the role of KDM2B on the chondrogenic differentiation. Alcian Blue Staining and Quantitative Analysis were used to investigate the synthesis of proteoglycans by chondrocytes. Real-time RT-PCR was used to detect the expressions of chondrogenesis related genes. The Alcian Blue staining and Quantitative Analysis results revealed that overexpression of KDM2B decreased the proteoglycans production, and real-time RT-PCR results showed that the expressions of the chondrogenic differentiation markers, COL1, COL2 and SOX9 were inhibited by overexpression of KDM2B in SCAPs. On the contrary, depletion of KDM2B increased the proteoglycans production, and inhibited the expressions of COL1, COL2 and SOX9. In conclusion, our results indicated that KDM2B is a negative regulator of chondrogenic differentiation in SCAPs and suggest that inhibition of KDM2B might improve MSC mediated cartilage regeneration.

[Effect of nerve growth factor on the early phase of osseointegration around oral implants].

OBJECTIVE: To observe the early bone integration of oral implants after injection of exogenous nerve growth factor (NGF) and investigate the effects of NGF on peri-implant osseointegration. METHODS: Twelve New Zealand white rabbits were used in this study to establish bi-mandible implant model. Then local injection of 1 µg NGF was given on the right side of the mandible as experimental group and normal saline only was injected on the left side as control group once a day for seven days. The rabbits were respectively sacrificed at 2, 4 and 8 weeks after surgery. The implant-bone grinding samples were prepared and stained by toluidine blue for general observation, X-ray, histology and bone histomorphometry analysis. RESULTS: The density of the new bone around implants at 2 and 4 weeks was lower than normal bone. Compared with the control group, the quantity of new bone and bone-implant contact ratio significantly increased in the experimental group. At 8 weeks, the new bone density in both groups was similar to the normal bone. In the experimental group, the haversian system was observed. Bone contact ratio was significantly different between experimental and control group at 2 and 4 weeks, but similar at 8 weeks.[control group at 2 weeks (26.67 ± 3.88)%, 4 weeks (52.59 ± 5.07)% and 8 weeks (97.33 ± 6.75)%, experimental group at 2 weeks (42.24 ± 6.67)%, 4 weeks (72.25 ± 6.30)% and 8 weeks (99.15 ± 4.68)%]. CONCLUSIONS: Applying exogenous NGF in the early phase could accelerate the formation and maturation of trabecular bone around the implants and shorten the period of osseointegration. Nerve growth factor could promote osseointegration in the early stage of oral implantation.

Effect of same-dose single or dual field irradiation on damage to miniature pig parotid glands.

AIM: To evaluate the effect of single or dual field irradiation (IR) with the same dose on damage to miniature pig parotid glands. METHODOLOGY: Sixteen miniature pigs were divided into two IR groups (n=6) and a control group (n=4). The irradiation groups were subjected to 20 Gy X-radiation to one parotid gland using single-field or dual-field modality by linear accelerator. The dose-volume distributions between two IR groups were compared. Saliva from parotid glands and blood were collected at 0, 4, 8 and 16 weeks after irradiation. Parotid glands were removed at 16 weeks to evaluate tissue morphology. RESULTS: The irradiation dose volume distributions were significantly different between single and dual field irradiation groups (t=4.177, P=0.002), although dose volume histogramin (DVH) indicated the equal maximal dose in parotid glands. Saliva flow rates from IR side decreased dramatically at all time points in IR groups, especially in dual field irradiation group. The radiation caused changes of white blood cell count in blood, lactate dehydrogenase and amylase in serum, calcium, potassium and amylase in saliva. Morphologically, more severe radiation damage was found in irradiated parotid glands from dual field irradiation group than that from single field irradiation group. CONCLUSION: Data from this large animal model demonstrated that the radiation damage from the dual field irradiation was more severe than that of the single field irradiation at the same dose, suggesting that dose-volume distribution is an important factor in evaluation of the radiobiology of parotid glands.

[Effect of single dose irradiation to parotid gland on the structured and function changes of bilateral parotid glands in miniature pig].

OBJECTIVE: To evaluate the effects of a solitary megadose protocol of ionizing radiation (IR) to parotid gland on the structured and function changes of bilateral parotid glands in miniature pig. METHODS: Fourteen minipigs were subjected to either 15 or 20 Gy to one parotid gland with a linear accelerator, while another four minipigs served as non-IR controls. Salivary flow rates and salivary chemistries were measured pre-IR, and 4 and 16 weeks post-IR. A quantitative assessment of gland weight and acinar area, and detailed serum chemistry and hematological analyses, were also performed. RESULTS: Parotid gland weights were significantly decreased in the 15 and 20 Gy groups at 4 and 16 weeks post-IR. The acinar cell area in glands of both IR groups was significantly reduced. Parotid flow rates decreased by 60% with 15 Gy at 16 weeks post-IR. In the 20 Gy group, salivary flow rates were reduced by 80% at 16 weeks post-IR. Additionally, parotid flow rates significantly reduced in contralateral glands with 20 Gy at 16 weeks, while structure and weight did not changes in parotid glands. CONCLUSION: Structural changes in salivary gland parenchyma occurred relatively early after IR, while the alterations in salivary output were relatively delayed. Further, reductions in salivary flow were not proportional to acinar cell area loss. There isn't a significant structured change of contralateral glands, but significant reduction of parotid flow rate at this time.