PLGA-based control release of Noggin blocks the premature fusion of cranial sutures caused by retinoic acid.

Craniosynostosis (CS), the premature and pathological fusion of cranial sutures, is a relatively common developmental disorder. Elucidation of the pathways involved and thus therapeutically targeting it would be promising for the prevention of CS. In the present study, we examined the role of BMP pathway in the all-trans retinoic acid (atRA)-induced CS model and tried to target the pathway in vivo via PLGA-based control release. As expected, the posterior frontal suture was found to fuse prematurely in the atRA subcutaneous injection mouse model. Further mechanism study revealed that atRA could repress the proliferation while promote the osteogenic differentiation of suture-derived mesenchymal cells (SMCs). Moreover, BMP signal pathway was found to be activated by atRA, as seen from increased expression of BMPR-2 and pSMAD1/5/9. Recombinant mouse Noggin blocked the atRA-induced enhancement of osteogenesis of SMCs in vitro. In vivo, PLGA microsphere encapsulated with Noggin significantly prevented the atRA-induced suture fusion. Collectively, these data support the hypothesis that BMP signaling is involved in retinoic acid-induced premature fusion of cranial sutures, while PLGA microsphere-based control release of Noggin emerges as a promising strategy for prevention of atRA-induced suture fusion.

All-Trans Retinoic Acid-Induced Craniofacial Malformation Model: A Prenatal and Postnatal Morphological Analysis.

OBJECTIVE: To characterize the prenatal and postnatal craniofacial bone development in mouse model of all-trans retinoic acid (ATRA) exposure at different ages by a quantitative and morphological analysis of skull morphology. METHODS: Pregnant mice were exposed to ATRA at embryonic day 10 (E10) and 13 (E13) by oral gavage. Skulls of mice embryos at E19.5 and adult mice at postnatal day 35 (P35) were collected for high-resolution microcomputed tomography (microCT) imaging scanning and section HE staining. Reconstruction and measurement of mouse skulls were performed for prenatal and postnatal analysis of the control and ATRA-exposed mice. RESULTS: Craniofacial malformations in mouse models caused by ATRA exposure were age dependent. ATRA exposure at E10 induced cleft palate in 81.8% of the fetuses, whereas the palatine bone of E13-exposed mice was intact. Inhibitions of maxilla and mandible development with craniofacial asymmetry induced were observed at E19.5 and P35. Compared with control and E13-exposed mice, the palatine bones of E10-exposed mice were not elevated and were smaller in dimension. Some E10-exposed mice exhibited other craniofacial abnormalities, including premature fusion of mandibular symphysis with a missing mandibular incisor and a smaller mandible. Severe deviated snouts and amorphous craniofacial suture were detected in E13-exposed mice at P35. CONCLUSION: These morphological variations in E10- and E13-exposed mice suggested that ATRA was teratogenic in craniofacial bone development in mice and the effect was age dependent.

pVEGF-loaded lipopolysaccharide-amine nanopolymersomes for therapeutic angiogenesis.

Therapeutic angiogenesis via gene delivery is promising for tissue survival and regeneration after injury or ischemia. A stable, safe and efficient gene vector is essential for successful angiogenesis. We have demonstrated that our newly developed lipopolysaccharide-amine nanopolymersomes (LNPs) have higher than 95% transfection efficiency when delivering pEGFP into mesenchymal stem cells (MSCs). To explore their clinical potential in therapeutic angiogenesis, in this study, we studied their toxicity, storage stability, protection ability to genes and efficacy to deliver therapeutic genes of pVEGF in MSCs and zebrafish. The results show that LNPs can condense pVEGF to form pVEGF-loaded nanopolymersomes (VNPs), and protect pVEGF against DNase digestion in 6 h. Both LNPs and VNPs have low toxicity to MSCs, erythrocytes and zebrafish embryos. LNPs are stable at 4 °C for at least two years with unchanged size and transfection efficiency. MSCs transfected by VNPs continuously synthesize VEGF for at least four days under control, with a peak (21.25 ng ml(-1)) ∼35-fold greater than that for the untreated group. VNPs induce significant and dose-dependent angiogenesis in zebrafish without causing death, deformity or delay in growth and development, and the induced maximal vessel area of subintestinal vessel plexus is 2.5-fold higher than that for the untreated group. Our study suggests that VNP has high potential in therapeutic angiogenesis.

Boosting Upconversion Efficiency in Optically Inert Shelled Structures with Electroactive Membrane through Electron Donation.

This study innovatively addresses challenges in enhancing upconversion efficiency in lanthanide-based nanoparticles (UCNPs) by exploiting Shewanella oneidensis MR-1, a microorganism capable of extracellular electron transfer. Electroactive membranes, rich in c-type cytochromes, are extracted from bacteria and integrated into membrane-integrated liposomes (MILs), encapsulating core-shelled UCNPs with an optically inactive shell, forming UCNP@MIL constructs. The electroactive membrane, tailored to donate electrons through the inert shell, independently boosts upconversion emission under near-infrared excitation (980 or 1550 nm), bypassing ligand-sensitized UCNPs. The optically inactive shell restricts energy migration, emphasizing electroactive membrane electron donation. Density functional theory calculations elucidate efficient electron transfer due to the electroactive membrane hemes' highest occupied molecular orbital being higher than the valence band maximum of the optically inactive shell, crucial for enhancing energy transfer to emitter ions. The introduction of a SiO2 insulator coating diminishes light enhancement, underscoring the importance of unimpeded electron transfer. Luminescence enhancement remains resilient to variations in emitter or sensitizing ions, highlighting the robustness of the electron transfer-induced phenomenon. However, altering the inert shell material diminishes enhancement, emphasizing the role of electron transfer. This methodology holds significant promise for diverse biological applications. UCNP@MIL offers an advantage in cellular uptake, which proves beneficial for cell imaging.

Single-cell transcriptomics reveals cell atlas and identifies cycling tumor cells responsible for recurrence in ameloblastoma.

Ameloblastoma is a benign tumor characterized by locally invasive phenotypes, leading to facial bone destruction and a high recurrence rate. However, the mechanisms governing tumor initiation and recurrence are poorly understood. Here, we uncovered cellular landscapes and mechanisms that underlie tumor recurrence in ameloblastoma at single-cell resolution. Our results revealed that ameloblastoma exhibits five tumor subpopulations varying with respect to immune response (IR), bone remodeling (BR), tooth development (TD), epithelial development (ED), and cell cycle (CC) signatures. Of note, we found that CC ameloblastoma cells were endowed with stemness and contributed to tumor recurrence, which was dominated by the EZH2-mediated program. Targeting EZH2 effectively eliminated CC ameloblastoma cells and inhibited tumor growth in ameloblastoma patient-derived organoids. These data described the tumor subpopulation and clarified the identity, function, and regulatory mechanism of CC ameloblastoma cells, providing a potential therapeutic target for ameloblastoma.

Dentin matrix protein 1 (DMP1) signals via cell surface integrin.

Dentin matrix phosphoprotein 1 (DMP1) is a non-collagenous, acidic extracellular matrix protein expressed chiefly in bone and dentin. We examined the DMP1 ability to engage cell-surface receptors and subsequently activate intracellular signaling pathways. Our data indeed show that the presence of extracellular DMP1 triggers focal adhesion point formation in human mesenchymal stem cells and osteoblast-like cells. We determine that DMP1 acts via interaction with αvß3 integrin and stimulates phosphorylation of focal adhesion kinase. Further biochemical characterization confirms the activation of downstream effectors of the MAPK pathways, namely ERK and JNK, after DMP1 treatment. This activation is specifically inhibitable and can also be blocked by the addition of anti-αvß3 integrin antibody. Furthermore, we show that extracellular treatment with DMP1 stimulates the translocation of phosphorylated JNK to the nucleus and a concomitant up-regulation of transcriptional activation by phosphorylated c-Jun. The evidence presented here indicates that DMP1 is specifically involved in signaling via extracellular matrix-cell surface interaction. Combined with the published DMP1-null data (Feng, J. Q., Ward, L. M., Liu, S., Lu, Y., Xie, Y., Yuan, B., Yu, X., Rauch, F., Davis, S. I., Zhang, S., Rios, H., Drezner, M. K., Quarles, L. D., Bonewald, L. F., and White, K. E. (2006) Nat. Genet. 38, 1310-1315) it can be hypothesized that DMP1 could be a key effector of ECM-osteocyte signaling.

Cephalometric analysis of craniofacial malformations in newborn mice with cleft palate induced by retinoic acid.

OBJECTIVE: To determine changes on craniofacial growth morphometrically in newborn mice with cleft palate induced by retinoic acid. DESIGN, SETTING, PARTICIPANTS, INTERVENTIONS: Gestation day 10 or 12 pregnant female C57BL/6N mice were given a single dose of all-trans retinoic acid (atRA) by gastric intubations via oral gavage. Sixty newborn mice with cleft palate (CP), 52 without CP from the experimental group, and 30 without CP from the control group were collected, and lateral cephalograms were taken of all of the mice. MAIN OUTCOME MEASURES: Cephalometric analysis of the craniofacial skeleton was performed by means of a personal computer. RESULTS: Inhibition of craniofacial growth was found in the experimental groups but not in the control groups. In the maxillary bone and mandible, the amount of growth was significantly reduced. CONCLUSIONS: These results suggest that craniofacial growth is inhibited in newborn mice with cleft palate induced by retinoic acid.

Retinoic acid inhibits osteogenic differentiation of mouse embryonic palate mesenchymal cells.

BACKGROUND: All-trans-retinoic acid (ATRA), a known teratogenic factor affecting the development of cleft palate, has been shown to adversely affect craniofacial development. In the present study, we evaluated the effects of ATRA on the osteo-/adipogenic differentiation of mouse embryonic palate mesenchymal (MEPM) cells, which served as a valid model system for investigating the mechanisms regulating osteogenesis during palatogenesis. METHODS: MEPM cells were derived from gestational day 13 C57BL/6N mouse embryos and induced to differentiate in the presence or absence of ATRA in either osteogenic medium (OM) or control medium (CM). RESULTS: Alkaline phosphatase (ALP) activity assays, von Kossa staining, and RT-PCR assays confirmed that MEPM cells underwent osteogenic differentiation when cultured in OM. Although ATRA induced ALP activity and lipid accumulation in MEPM cells, it failed to induce matrix mineralization and osteoblastic gene expression. BMPR-IB and Smad5 mRNA levels increased significantly in cells cultured in OM and declined following treatment with ATRA, whereas the expression of the BMPR-IA mRNA was up-regulated by ATRA. CONCLUSIONS: In conclusion, our results suggested that ATRA and the BMP signaling pathway cooperate to inhibit osteogenesis and promote adipogenesis of MEPM cells.

The role of RANKL and MMP-9 in the bone resorption caused by ameloblastoma.

BACKGROUND: Ameloblastoma, a common odontogenic tumor located in jaws, generally leads to severe damage to patient's complexion and masticatory function. To expand in jaws, ameloblastoma must have a mechanism of resorbing the surrounding bone. Our objective was to explore the bone-resorption mechanism of ameloblastoma by observing the role of Receptor activator of nuclear factor kappa B ligand (RANKL) and matrix metalloproteinase-9 (MMP-9) in the bone-resorption process. METHODS: In the study, the expression of RANKL and MMP-9 in ameloblastoma was detected using immunohistochemistry (IHC) and RT-PCR. Then, co-culture system of ameloblastoma cells and bone marrow cells from neonatal rabbit was erected to observe the potential of ameloblastoma cells to induce osteoclastogenesis. Finally, the induced osteoclasts were used for in vitro bone-resorption assay. In the co-culture system and the bone-resorption assay, the selective inhibitor of RANKL and MMP-9, osteoprotegerin (OPG) and tissue inhibitor of metalloproteinase-1 (TIMP-1) were, respectively, used for observing the role of RANKL and MMP-9. RESULTS: The expression of RANKL and MMP-9 in ameloblastoma was confirmed. Ameloblastoma cells were found to induce bone marrow cells from neonatal rabbit differentiate into osteoclasts with bone-resorption activity. In addition, OPG was found to, respectively, have markedly inhibitory effect on osteoclastogenesis (P<0.01), and slightly inhibitory action on bone resorption (P<0.05). CONCLUSIONS: Ameloblastoma cells had the potential to induce osteoclastogenesis. Moreover, RANKL played an essential role in the in vitro osteoclast formation and bone resorption induced by ameloblastoma cells.

Expression and role of metalloproteinase-2 and endogenous tissue regulator in ameloblastoma.

BACKGROUND: Ameloblastoma is a frequently encountered odontogenic benign tumor characterized by local invasiveness and high risk of recurrence. Matrix metalloproteinase (MMP)-2 can degrade type IV collagen, one of the major components in the basement membrane, resulting in the promotion of tumor invasion, whereas it is under the influence of an activator, membrane type 1-MMP-14 and the tissue inhibitor of metalloproteases (TIMP)-2. The aim of this study was to investigate combinatorial role played by MMP-2, TIMP-2 and MMP-14 in ameloblastoma. METHODS: Transcriptional expression of MMP-2, TIMP-2 and MMP-14 in tissue extracts of 42 ameloblastomas and 10 dental follicles was detected using reverse transcription-polymerase chain reaction, and compared difference in clinical type and local recurrence of ameloblastoma. RESULTS: MMP-2, TIMP-2 and MMP-14 mRNA were detected in all investigated ameloblastoma tissues. While MMP-2 mRNA was only expressed in eight of 10 odontotheca tissues and TIMP- 2 and MMP-14 were not found in all odontotheca tissues. The mRNA expression of MMP-2, TIMP-2 and MMP-14 were significantly higher in ameloblastoma tissues than in odontotheca tissues. The mRNA levels of TIMP-2 and MMP-14 were significantly higher in recurrent and solid/multicystic ameloblastoma tissues than in primary and unicystic ameloblastoma tissues respectively. CONCLUSIONS: Of the MMP-2/TIMP-2/MMP-14 complex, a high expression of MMP-2, TIMP-2 and MMP-14 mRNA levels may contribute to the local invasive characteristics of ameloblastoma, whereas the local invasive capacity of ameloblastoma is more likely related to a high transcriptional levels of TIMP-2 and MMP-14.

Smad2/3 is involved in growth inhibition of mouse embryonic palate mesenchymal cells induced by all-trans retinoic acid.

BACKGROUND: Cleft palate is one of the major malformations induced by retinoic acid in both rodents and humans. The purpose of the present study was to elucidate the mechanism by which all-trans retinoic acid (atRA) induces the cleft palate. METHODS: The cell cycle distribution of mouse embryonic palate mesenchymal (MEPM) cells under atRA (100 mg/kg) treatment on gestation day (GD) 10 or GD 12 were measured by immunohistochemistry and flow cytometry. The p21, phospho-Rb, Smad2/3, phospho-Smad2 and phospho-Smad3 protein expression levels were detected by western blot, respectively. Quantitative real-time PCR was performed for p21, Smad2, and Smad3 gene expression in each group under both conditions. Small interfering RNA (siRNA) was applied to inhibit Smad2/3 expression in MEPM cells and the effect was detected by western blot and flow cytometry. RESULTS: The G(0)/G(1) arrest in MEPM cells in vivo was induced by atRA on GD 10. The protein expression levels of p21, Smad2/3, phospho-Smad2, and phospho-Smad3 were increased, while phospho-Rb was decreased in MEPM after atRA treatment on GD 10. These changes were not observed on the GD 12 group. Moreover, the mRNA expression levels of p21, Smad2, and Smad3 detected by quantitative real-time PCR were almost consistent with their protein expression trends. Furthermore, p21 was partially decreased and G(0)/G(1) arrest was partially released following Smad2/3 siRNA knockdown. CONCLUSIONS: The induction of G(0)/G(1) block by atRA in MEPM cells varied with the development stage of exposure. Our study demonstrated that Smad2/3 regulation of p21 was partly required for atRA-induced cell cycle perturbations in MEPM cells.

EMMPRIN expression in tongue squamous cell carcinoma.

BACKGROUND: Extracellular matrix metalloproteinase inducer (EMMPRIN) is identified as a tumor-cell membrane protein that stimulates matrix metalloproteinases (MMPs) production. Several studies have shown that higher EMMPRIN expression is associated with shorter survival time and correlated significantly with more advanced clinico-parameters of cancer. The aim of this study was to investigate the relationship between clinico-pathologic characteristics and EMMPRIN, and prognostic significance of EMMPRIN expression in human tongue squamous cell carcinoma. METHODS: Extracellular MMP inducer expression was examined immunohistochemically on paraffin-embedded tissue specimens from 68 patients with tongue squamous cell carcinoma and who underwent radical surgeries from 1996 to 2006. The 68 patients were followed up from 1 to 119 months, with an average of 27.5 months. Nonparametric tests were performed for the comparison of EMMPRIN expression between two independent groups. Survival analysis was performed to find the prognostic significance of EMMPRIN expression. RESULTS: We found that EMMPRIN expression in tongue squamous cell carcinoma is significantly higher than that in non-cancerous epithelium adjacent to carcinoma of tongue. In addition, EMMPRIN expression is significantly associated with tumor diameter and clinical stage in the samples, but did not correlate with gender, age, tumor metastasis, and pathological grade. Finally, survival analysis indicates that EMMPRIN overexpression correlates significantly with poor overall survival in the patient cohort. CONCLUSION: These results suggest that EMMPRIN might represent an attractive target for immunotherapeutic approaches in a subgroup of patients with tongue squamous cell carcinoma.

Inhibition of ameloblastoma invasion in vitro and in vivo by inhibitor of metalloproteinase-2 activity.

BACKGROUND: Ameloblastoma is an odontogenic benign tumor characterized by local invasiveness and most of its local recurrences clinically result from local invasion. This study used matrix metalloproteinase-2 (MMP-2) inhibitor I (MMP-2I) to investigate the role played by MMP-2 activity in the local invasiveness of ameloblastoma. METHODS: The cells and xenografts of ameloblastoma were treated with MMP-2I and treatment group were compared with the control group. In vitro, the invasive activity of tumor cells was assayed in transwell cell culture chamber. Gelatinolytic activity of gelatinases and MMP-2/tissue inhibitor of matrix metalloproteinase (TIMP-2) protein expression was detected using gelatin zymography and flow cytometry. The cell viability and adhesion were evaluated using methyl thiazol tetrazolium. In vivo, bilateral subrenal capsule xenograft transplantation of ameloblastoma was performed in 10 nude mice and the invasion of ameloblastoma into the renal parenchyma was observed. RESULTS: Active-MMP-2 of conditioned media was significantly lower in treatment group than in the control group. Accordingly, potential of in vitro cell invasion, adhesion and in vivo tumor invasion were also significantly lower in the treatment group than in the control group. CONCLUSIONS: Inhibitor of MMP-2 activity suppressed the local invasive capability of ameloblastoma by decreasing MMP-2 activity. MMP-2 activity is in relation with invasive capacity of ameloblastoma.

Suppression of local invasion of ameloblastoma by inhibition of matrix metalloproteinase-2 in vitro.

BACKGROUND: Ameloblastomas are odontogenic neoplasms characterized by local invasiveness. This study was conducted to address the role of matrix metalloproteinase-2 (MMP-2) in the invasiveness of ameloblastomas. METHODS: Plasmids containing either MMP-2 siRNA or tissue inhibitor of metalloproteinase-2 (TIMP-2) cDNA were created and subsequently transfected into primary ameloblastoma cells. Zymography, RT-PCR, and Western blots were used to assess MMP-2 activity and expression of MMP-2 and TIMP-2, as well as protein levels. RESULTS: Primary cultures of ameloblastoma cells expressed cytokeratin (CK) 14 and 16, and MMP-2, but only weakly expressed CK18 and vimentin. MMP-2 mRNA and protein levels were significantly inhibited by RNA interference (P < 0.05). Both MMP-2 siRNA and TIMP-2 overexpression inhibited MMP-2 activity and the in vitro invasiveness of ameloblastoma. CONCLUSION: These results indicate that inhibition of MMP-2 activity suppresses the local invasiveness of ameloblastoma cells. This mechanism may serve as a novel therapeutic target in ameloblastomas pursuant to additional research.

[Effect analysis of modified radical surgery in tongue cancer at early stages].

OBJECTIVE: To evaluate the oncologic efficacy, surgical safety, postoperative beauty, and morbidity of an ideal procedure of modified radical surgery in tongue cancer at early stages. METHODS: Six patients with early stage squamous cell carcinoma of tongue underwent a procedure of modified radical surgery, in which an inconspicuous incision was used, the external jugular vein, sternocleidomastoid muscle, spino-accessory nerve, and greater auricular nerve were preserved, half of glosso and mouth floor were excised, and the defect was repaired instantly. Recurrence of glosso, mouth floor and neck, regional edema, shoulder dysfunction, auricle sensibility, oral cavity function and beautiful outlook on the face and neck were evaluated clinically. RESULTS: Compared with classic radical neck dissection, this procedure of modified radical surgery showed an inconspicuous incision and pretty appearance, minor edema on face and neck, better shoulder function, and sensation of auricular skin. No recurred to the glosso, mouth floor, and neck was found during follow-up. The patients showed better oncological safety, pretty appearance of tongue, and better oral function of speech, swallow and mastication. CONCLUSION: This ideal procedure of modified radical surgery in tongue cancer at early stages lessens or avoids destruction of face and neck, shoulder malfunction, numbness of auricular skin, and oral dysfunction of speech, swallow and mastication without impairment on the oncologic safety of the radical surgery, and improves the quality of life of the patients.

[Combined repair of large defect caused by radical surgery of advanced tongue cancer with rib-major pectoralis myocutaneous flap carrying costal parietal pleura].

OBJECTIVE: To explore the clinical value and safety of using rib-major pectoralis myocutaneous flap carrying costal parietal pleura in combined repair of large soft and hard tissue defect caused by radical surgery of advanced tongue cancer. METHODS: Six patients with advanced tongue carcinoma involving the floor of mouth and mandible were performed combined radical neck dissection with glossectomy and mandibulectomy, which caused large soft and hard tissue defect. Six rib-major pectoralis myocutaneous flaps carrying costal parietal pleura were transferred for immediate repair of the large defects. The rib flaps were applied for the repair of mandible, and the major pectoralis myocutaneous flaps were applied for the reconstruction of tongue and floor of mouth. RESULTS: Six patients recovered well after operation. Six rib-major pectoralis myocutaneous flaps carrying costal parietal pleura survived well; the wounds of surgical incision of the oral cavity, neck, and chest healed up. The reconstructed tongue and the lower face appearance were satisfactory, the occlusion relationships were normal; the speaking as well as swallowing functions recovered. CONCLUSIONS: It's safe and reliable to use rib-major pectoralis myocutaneous flap carrying costal parietal pleura to repair large soft and hard tissue defect in oral and maxillofacial region. Opening pleural cavity and harvest costal parietal pleura would not influence patients' thoracic movement and breath function and would not cause other complications. It's simple and safe for harvesting the composite flap. Carrying costal parietal pleura assures the sufficient blood supply of rib in the composite flap.

[Effect of stiffness of polyelectrolyte multilayer on titanium surface on bacterium adhesion].

OBJECTIVE: To provide a theoretical basis for surface modification of titanium implants, the effects of the stiffness of polyelectrolyte multilayer films on titanium surface on bacterium adhesion was explored. METHODS: Via layer-by-layer technique, catechol functionalized polyelectrolyte multilayer film (cPEM) was constructed on titanium surface by using catechol functionalized hyaluronic acid (cHA) and lipopolysaccharide-amine nanopolymersomes (NP). The stiffness of cPEM was controlled by adjusting the catechol substitution degree of cHA (5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%). Titanium samples covered with cPEM were selected as test group, and the cPEM was constructed with the lowest, medium and highest stiffness. The polished titanium was used as a control. The surface topography of titanium before and after film construction was observed by scanning electron microscopy (SEM). At 1 and 24 h after incubation, the adhesion and clonal formation of Streptococcus mutans (S. mutans) on different titanium surfaces were quantified, and their morphology and survival status were observed by SEM and laser scanning confocal microscope (LSCM). RESULTS: When the catechol grafting ratio was 5%, 30% and 70%, the lowest, medium and highest cPEM stiffness were obtained, and the cPEM stiffness were (10.69±4.54) GPa(cPEM-L), (20.99± 5.81) GPa (cPEM-M) and (32.57±6.93) GPa (cPEM-H) respectively, and the stiffness of polished titanium was (107.12±8.68) GPa (P<0.05). SEM observation showed that after cPEM coating, the titanium surface became smoother. After incubation for 1 and 24 h, the amount of adhesion and clonal formation of S. mutans on cPEM were higher than those on control titanium, and the difference was statistically significant (P<0.05). SEM images showed that for 1 h incubation, softer surfaces were beneficial for S. mutans adhering and agglomerating, while this difference nearly disappeared at 24 h. Observation under LSCM revealed that most of bacteria were alive on titanium disks at 1 h, and their amount decreased with the increase of stiffness. At 24 h, the living/dead bacterium ratios on cPEM-L and control titanium was higher than that on cPEM-M and cPEM-H, and cPEM-L surface was dominated by living bacteria, while stiffer cPEM-M and cPEM-H had more dead bacteria than living bacteria. CONCLUSIONS: Increasing the stiffness of polyelectrolyte films on titanium limits the adhesion of S. mutans. As an independent factor, stiffness influences the bacterium adhesion.

[Study on gene transfection in bone marrow mesenchymal stem cells mediated by plasmid of bone morphogenetic protein 2 loaded lipopolysaccharide-amine nanopolymersomes].

OBJECTIVE: To evaluate the combination of lipopolysaccharide-amine nanopolymersomes (LNPs), as a gene vector, with target gene and the transfection in bone marrow mesenchymal stem cells (BMSCs) so as to provide a preliminary experiment basis for combination treatment of bone defect with gene therapy mediated by LNPs and stem cells. METHODS: Plasmid of bone morphogenetic protein 2 (pBMP-2)-loaded LNPs (pLNPs) were prepared. The binding ability of pLNPs to pBMP-2 was evaluated by a gel retardation experiment with different ratios of nitrogen to phosphorus elements (N/P). The morphology ofpLNPs (N/P = 60) was observed under transmission electron microscope (TEM) and atomic force microscope (AFM). The size and Zeta potential were measured by dynamic light scattering (DLS). The resistance of pLNPs against DNase I degradation over time was explored. The viability of BMSCs, transfection efficiency, and expression of target protein were investigated after transfection by pLNPs in vitro. RESULTS: At N/P ≥ 1.5, pLNPs could completely retard pBMP-2; at N/P of 60, pLNPs was uniform vesicular shape under AFM; TEM observation demonstrated that pLNPs were spherical nano-vesicles with the diameter of (72.07 ± 11.03) nm, DLS observation showed that the size of pLNPs was (123 ± 6) nm and Zeta potential was 20 mV; pLNPs could completely resist DNase I degradation within 4 hours, and such protection capacity to pBMP-2 decreased slightly at 6 hours. The cell survival rate first increased and then decreased with the increase of N/P, and reached the maximum value at N/P of 45; the cytotoxicity was in grade I at N/P ≤ 90, which meant no toxicity for in vivo experiment. While the transfection efficiency of pLNPs increased with the increase of N/P, and reached the maximum value at N/P of 60. So it is comprehensively determined that the best N/P was 60. At 4 days, transfected BMSCs expressed BMP-2 continuously at a relatively high level at N/P of 60. CONCLUSION: LNPs can compress pBMP-2 effectively to form the nanovesicles complex, which protects the target gene against enzymolysis. LNPs has higher transfection efficiency and produces more amount of protein than polyethylenimine 25k and Lipofectamine 2000.

Application of CAD/CAM-assisted technique with surgical treatment in reconstruction of the mandible.

OBJECTIVE: The purpose of this clinical study was to explore the optimal method of reconstruct mandible defects individually and immediately. STUDY DESIGN: Three-dimensional model simulation technique and vascularized fibular osteomyocutaneous flap were used to repair 15 cases of mandible defects, which were caused by ameloblastoma. A three-dimensional computed tomography (CT) images were converted to a virtual model using CAD software and the 3-dimensional (3D) simulated resin models of skeleton and fibula were used to design the osteotomies, bone segment replacement and titanium mesh shaping preoperatively. RESULTS: Fibula flaps were alive and no complication occurred. The patients were satisfied with the results both esthetically and functionally. CONCLUSIONS: This preliminarily clinical study and case demonstrated that CAD/CAM-assisted technique with surgical treatment offers an individual anatomical reconstruction of the mandible in ameloblastoma patients. The procedures guarantee intraoperatively an exact placement of the preformed mesh even for precise reconstruction of extensive mandible defects.