Adjustable Removable Traction Appliance With Surgically Assisted Eruption for Impacted Maxillary Central Incisors.

Context: Impacted maxillary central incisors (MCIs) can seriously affect children's appearance, verbal abilities, and maxillofacial development. Clinically, a combination of surgically assisted eruption and orthodontic traction is the treatment modality most acceptable to dentists and children's families. However, previously used traction methods have been complex and required a long treatment time. Objective: The study intended to evaluate the clinical effects of the use of the research team's adjustable removable traction appliance combined with a surgically assisted eruption of impacted MCIs. Design: The research team performed a controlled prospective study. Setting: The study took place at Department of Orthodontics, Hefei Stomatological, Hospital. Participants: 10 patients with impacted MCIs, aged 7-10 years, who had visited the hospital between September 2017 and December 2018. Intervention: The research team assigned the impacted MCIs to the intervent ion group and contral ateral normal MCIs to thecontrol group. For the intervention group, the research team performed a surgical eruption and inserted the adjustable removable traction appliance. The control group received no treatments. Outcome Measures: Postintervention, the research team determined the mobility of both groups' teeth. At baseline and immediately postintervention for both groups, the team performed cone-beam computed tomography (CBCT) and measured root length, apical-foramen width, volume, surface area, and root-canal wall thickness for the labial and palatal sides. For both groups, after the intervention group's treatments, the team: (1) performed electric pulp testing and periodontal probing on the participants' teeth; (2) measured and documented pulp vitality, gingival index, periodontal probing depth, and gingival height (GH)for the labial and palatal sides; and (3) measured labial-and-palatal, alveolar bone level and alveolar bone thickness. Results: At baseline, the intervention group showed delayed root development, and that group's root length was significantly shorter (P < .05) and apical-foramen width (P < .05) was significantly greater than those of the control group. The intervention group's treatment success rate was 100%. And the intervention group did not have any adverse reactions, such as tooth loosening, gingival redness and swelling, or bleeding. Postintervention, the intervention group's labial GH was significantly higher than that of the control group, at 10.58 ± 0.45 mm and 9.47 ± 0.31 mm, respectively (P = .000). The increase in the intervention group's root length postintervention was significantly greater than that of the control group, at 2.80 ± 1.09 mm and 1.84 ± 0.97 mm, respectively (P < .05). The intervention group also had significantly greater decrease in the apical-foramen width than the control group did, at 1.79 ± 0.59 mm and 0.96 ± 0.40 mm, respectively (P < .05). At the end of traction, the intervention group had significantly higher labial-and-palatal alveolar-bone levels, at 1.77 ± 0.37 mm and 1.23 ± 0.21 mm, respectively, than the control group did, at 1.25 ± 0.26 mm (P = .002) and 1.05 ± 0.15 mm (P = .036), respectively. The labial alveolar-bone thickness in the intervention group was thinner than that of the control group, at 1.49 ± 0.31 mm and 1.80 ± 0.11 mm, respectively (P = .008). The volume and surface area (P < .01) of the intervention group's impacted teeth had increased significantly postintervention (both P < .01), but both were significantly smaller than those of the control group, both at baseline and postintervention. Conclusions: An adjustable removable traction appliance combined with a surgically assisted eruption can be a reliable treatment for impacted MCIs and can provide root development and a good periodontal-pulp condition postintervention.

Compensation Method for Sensor Network Clock Error Based on Cyclic Symmetry Algorithm.

Since the existing methods cannot evaluate the time delay of different layers of sensor networks, there are some problems such as the low precision of clock error compensation, high time delay, and low efficiency of communication in sensor networks. To solve this problem, a method of clock error compensation in sensor networks based on a cyclic symmetry algorithm is proposed. Based on the basic theory of cyclic symmetry, the cyclic symmetry matrix of the sensor network is constructed; in the communication process, all nodes are extended to get the cumulative delay rate of the sensor network in the specified time domain. Using the cumulative delay rate of the cyclic network and the sensor network, the autoregressive integral sliding mode control model is established to compensate for the cumulative error of clock synchronization. The simulation results show that the compensation accuracy of this method is more than 98%, which can effectively reduce the delay of sensor network. It improves the communication efficiency of the sensor network, meets the communication requirements of the sensor network, and has a very broad application prospect.

ALDH1A3 affects colon cancer in vitro proliferation and invasion depending on CXCR4 status.

BACKGROUND: Aldehyde dehydrogenase (ALDH) has been widely used as a marker of cancer stem cells (CSCs). However, the ALDH family includes 19 members, and the most relevant isoforms and their biological functions in cancer biology are still controversial. METHODS: We examined ALDH enzyme activity and the mRNA expression of 19 ALDH members in 58 human cell lines. The biological effect and mechanism of knocking down ALDH1A3 with siRNA and shRNA in cell lines were explored. Finally, the relationship between ALDH1A3 and CXCR4 was analysed in a large panel of cell lines. RESULTS: ALDH1A3 is the key isoform that contributed to Aldefluor positivity in cell lines. Knocking down ALDH1A3 in different cancer cells conferred opposite phenotypes due to differential effects on CXCR4 expression. There was a significant negative correlation between ALDH1A3 and CXCR4 in 58 human cell lines. CONCLUSIONS: ALDH1A3 was the main contributor to Aldefluor positivity in human cell lines, and its contrasting effects might arise from differences in CXCR4 expression.

Intrinsic gene changes determine the successful establishment of stable renal cancer cell lines from tumor tissue.

Human tumor cell lines, especially those with complete data and follow-up, are important tools in tumor biological studies. Clear cell renal cell cancer (ccRCC) is not sensitive to radiotherapy or chemotherapy, and treatment of patients with distant metastasis relies on targeted therapy. Here, we report the establishment of seven new ccRCC stable cell lines that were continuously cultured for more than 20 generations among 81 cases of renal cell cancer. Moreover, gene expression and methylation in the established cell lines, in those that had a finite in vitro life span of less than 10 generations, and in cells that originated from the same culture at different generations were profiled using microarrays. Genes including SLC34A2, SEPP1, SULT1C4 and others were differentially expressed in established cell lines and finite cell lines, and changes in their expression might be caused by methylation or demethylation. The expression level of SLC34A2 was related not only to the life span in vitro culture but also to tumor size. Additionally, six of the seven new ccRCC cell lines had VHL deletions or termination mutations. So in addition to the establishment of seven new ccRCC cell lines with complete clinical data, we conclude that genes such as SLC34A2 and VHL play key roles in the continuous in vitro growth and development of ccRCC.

Cloning and expression of a novel target fusion protein and its application in anti-tumor therapy.

BACKGROUNDS: Epidermal growth factor (EGF) is a 53 amino acid polypeptide and its receptor EGFR is an established therapeutic target for anti-tumor therapy. Two major categories of EGFR-targeted drugs include monoclonal antibodies (mAbs) and small molecular tyrosine kinase inhibitors (TKIs). However, drug resistance occurs in a significant proportion of patients due to EGFR mutations. Since EGFR can maintain activation while abrogating the activity of mAbs or TKIs, or bypass signaling functions while successfully circumventing the EGF-EGFR switch, developing new mechanism-based inhibitors is necessary. METHODS: In this study, based on the principle of tumor immunotherapy, a recombinant protein pLLO-hEGF was constructed. The N-terminal portion contains three immunodominant epitopes from listeriolysin O (LLO) and the C-terminal includes EGF. To use EGF as a target vector to recognize EGFR-expressing cancer cells, immunodominant epitopes could enhance immunogenicity of tumor cells for immune cell activation and attack. RESULTS: Recombinant protein pLLO-hEGF was successfully expressed and showed strong affinity to cancer cells. Also, pLLO-hEGF could significantly stimulate human lymphocyte proliferation and the lymphocytes demonstrated enhanced killing potency in EGFR-expressing cancer cells in vitro and in vivo. CONCLUSION: This study can provide novel strategies and directions in tumor biotherapy.

[Correlation of Contrast-enhanced Ultrasound with Expression of Hypoxia Inducible Factor-1α in Transplanted Mice Mammary Cancer].

OBJECTIVE: To investigate the correlation of contrast-enhanced pattern with expression of hypoxia inducible factor-1α (HIF-1α) and microvessel density (MVD) in mice breast cancer. METHODS: A total of 22 mice were implanted with breast cancer cells (Ca761) subcutanously in the thigh. The tumors were examined with conventional ultrasound and contrast-enhanced ultrasound (CEUS) on days 4,6,7,8,9,10,and 11 after implantation and then sacrificed. Three or four mice were included each time. Expressions of HIF-1α and MVD in cancer tissues were detected immunohistochemically. Correlation of contrast-enhanced patterns with expression of HIF-1α and MVD in breast cancer was analyzed. RESULTS: Mice were divided into 3 groups according to the tumor volume:group 1 (volume<0.05 cm(3),n=5),group 2 (volume 0.05-0.75 cm(3),n=9),and group 3 (volume>0.75 cm(3),n=8). The CEUS pattern was different in different groups:four mice in group 1 presented as type 1 (peripheral ring enhancement with no enhancement within the tumor) and 1 case presented as type 2 (peripheral ring enhancement with deep penetration). Most mice in group 2 presented as type 3 (homogeneous or heterogeneous enhancement in the whole tumor,n=5). In group 3,most mice presented as type 4 (peripheral ring enhancement with focal nodular enhancement within the tumor,n=7). Contrast-enhanced pattern was significantly different in different volume groups (P<0.01). Enhanced pattern (type 1-4) was closely correlated with tumor volume (r=0.841,P<0.05). The expression of HIF-1α was negatively correlated with enhanced patterns (type 1-4) (r=-0.596,P＝0.003),but not with tumor volume (P>0.05). There was no significant difference in MVD values between different enhanced patterns (type 1-4),and there was no correlation between the MVD and tumor volumes (P>0.05). CONCLUSION: CEUS can be used as a noninvasive tool to monitor tumor angiogenesis in tumor and the enhanced patterns may reflect the expression of HIF-1α inside the tumor.

Ubiquitin-specific peptidase USP22 negatively regulates the STAT signaling pathway by deubiquitinating SIRT1.

BACKGROUND/AIMS: The ubiquitin-specific peptidase USP22 mediates various cellular and organismal processes, such as cell growth, apoptosis, and tumor malignancy. However, the molecular mechanisms that regulate USP22 activity remain poorly understood. Here we identify STAT3 as a new USP22 interactor. METHODS: · We used western blotting and RT-PCR to measure key protein, acetylated STAT3, and mRNA levels in HEK293 and colorectal cancer cell lines transfected with expression plasmids or specific siRNAs. Co-immunoprecipitation was used to demonstrate protein-protein interaction and protein complex composition. RESULTS: USP22 overexpression down-regulated STAT3 acetylation by deubiquitinating SIRT1. The three proteins were found to be present in a single protein complex. SiRNA-mediated depletion of endogenous USP22 resulted in SIRT1 destabilization and elevated STAT3 acetylation. Consistent with this finding, USP22 also down-regulated the expression of two known STAT3 target genes, MMP9 and TWIST. CONCLUSION: We show that USP22 is a new regulator of the SIRT1-STAT3 signaling pathway and report a new mechanistic explanation for cross talk between USP22 and the SIRT1-STAT pathways.

CD133 affects the invasive ability of HCT116 cells by regulating TIMP-2.

CD133 is widely expressed in colorectal cancer (CRC) tissues and cell lines. This protein has been used as a marker of CRC cancer stem cells, although the function and mechanism of CD133 in CRC invasion and metastasis remain unclear. In our study, we examined the role of CD133 in CRC invasion in vitro and investigated the mechanism involved in CD133-related invasion. CD133(high) and CD133(low) HCT116 cells were isolated, and the proliferation and invasive ability of these two subpopulations were tested. CD133(high) HCT116 cells exhibited greater proliferation and invasion compared with CD133(low) HCT116 cells. CD133 knockdown (using CD133 small-interfering [si]RNA) inhibited the invasive activity of CD133si-HCT116 cells. For the first time, we found that the expression of tissue inhibitor of matrix metalloproteinases-2 (TIMP-2) was down-regulated in CD133si-HCT116 cells. In addition, for the TIMP-2si-HCT116 cells (transfected with TIMP-2 siRNA), in vitro invasion was significantly decreased, whereas the expression of CD133 remained unchanged. Finally, the metalloproteinase 2 and MEK/ERK signaling pathways were examined, and no significant change was observed after the knockdown of CD133 or TIMP-2 in HCT116 cells. In conclusion, we demonstrated that CD133 plays an important role in HCT116 cell invasion, and for the first time, we found that CD133 knockdown significantly down-regulated TIMP-2 expression, which suggests that CD133 affects the invasive ability of HCT116 cells by regulating TIMP-2.

[Enhanced growth inhibition by combined two pathway inhibitors on K-ras mutated non-small cell lung cancer cells].

OBJECTIVE: To evaluate the effect of combined targeting of MEK and PI3K signaling pathways on K-ras mutated non-small cell lung cancer cell line A549 cells and the relevant mechanisms. METHODS: A549 cells were treated with different concentrations of two inhibitors. Growth inhibition was determined by MTT assay. According to the results of MTT test, the cells were divided into four groups: the control group, PI3K inhibitor group (GDC-0941,0.5 and 5.0 µmol/L), combination group I (0.5 µmol/L AZD6244+0.5 µmol/L GDC-0941) and combination group II (5.0 µmol/L AZD6244+5.0 µmol/L GDC-0941). The cell cycle and apoptosis were analyzed by flow cytometry. The expression of proteins related to apoptosis was tested with Western blot. RESULTS: Both GDC-0941 and AZD6244 inhibited the cell proliferation. The combination group II led to a stronger growth inhibition. The combination group I showed an antagonistic effect and combination group II showed an additive or synergistic effect. Compared with the control group, the combination group I led to reduced apoptotic rate [(20.70 ± 0.99)% vs. (18.65 ± 0.92 )%, P > 0.05]; Combination group II exhibited enhanced apoptotic rate [(37.85 ± 3.18)% vs. (52.27 ± 4.36)%, P < 0.01]. In addition, in the combination group II, more A549 cells were arrested in G0/G1 phase and decreased S phase (P < 0.01), due to the reduced expressions of CyclinD1 and Cyclin B1, the increased cleaved PARP and the diminished ratio of Bcl-2/Bax. CONCLUSIONS: For single K-ras mutated NSCLC cell line A549 cells, combination of RAS/MEK/ERK and PI3K/AKT/mTOR inhibition showed synergistic effects depending on the drug doses. Double pathways targeted therapy may be beneficial for these patients.

Assessment of the osteogenic effect after maxillary sinus floor elevation and simultaneous implantation with or without bone grafts by analyzing trabecular bone parameters: a retrospective study.

OBJECTIVE: The aim of this population-based retrospective study was to compare the osteogenic effect of newly formed bone after maxillary sinus floor elevation (MSFE) and simultaneous implantation with or without bone grafts by quantitatively analyzing trabecular bone parameters. METHODOLOGY: A total of 100 patients with missing posterior maxillary teeth who required MSFE and implantation were included in this study. Patients were divided into two groups: the non-graft group (n=50) and the graft group (n=50). Radiographic parameters were measured using cone beam computed tomography (CBCT), and the quality of newly formed bone was analyzed by assessing trabecular bone parameters using CTAn (CTAnalyzer, SkyScan, Antwerp, Belgium) software. RESULTS: In the selected regions of interest, the non-graft group showed greater bone volume/total volume (BV/TV), bone surface/total volume (BS/TV), trabecular number (Tb. N), and trabecular thickness (Tb. Th) than the graft group (p<0.001). The non-graft group showed lower trabecular separation (Tb. Sp) than the graft group (p<0.001). The incidence of perforation and bleeding was higher in the graft group than in the non-graft group (p<0.001), but infection did not significantly differ between groups (p>0.05). Compared to the graft group, the non-graft group showed lower postoperative bone height, gained bone height and apical bone height (p<0.001). CONCLUSION: MSFE with and without bone grafts can significantly improve bone formation. In MSFE, the use of bone grafts hinders the formation of good quality bone, whereas the absence of bone grafts can generate good bone quality and limited bone mass.

[Observation on the growth and metastasis of cross-strain transplanted tumors in different mouse strains].

OBJECTIVE: Mouse tumors were subcutaneously transplanted into different mouse strains and their growth and metastatic properties were checked, to explore the possibility of establishing animal tumor models in different mouse strains other than their normal host strains. METHODS: Seven mouse tumor cell lines: H22, S180, U14, FC, Ca761, SMG-A and DCS were transplanted into C57BL/6J, ICR or KM mice, and their tumorigenicity, growth and metastasis were recorded and analyzed. RESULTS: The tumor formation rate of H22 cells in both the C57BL/6J and ICR mice was 100%, but the growth of H22 tumors was significantly faster in the C57BL/6J (2.8 ± 0.4)g than in the ICR mice (1.5 ± 0.5)g at the 17th day after transplantation (P<0.001). The S180 tumors grew stably in C57BL/6J mice and the tumor formation rate was 100%. The U14 inoculated into C57BL/6J and KM mice showed both lymphatic and lung metastasis and formed significantly larger tumors in KM mice [(12.6 ± 3.4)g] than that in the C57BL/6J mice [(10.2 ± 2.2)g] on the 32rd day after transplantation (P = 0.002). Transplantation of FC, Ca761, and SMG-A did not form tumors or the tumors were completely regressed later in C57BL/6J mice. DCS cells formed tumors in C57BL/6J mice, but some of the tumors regressed. The retained tumors were passaged in C57BL/6J mice, and the substrain DCS-C57 cells was established which showed stable growth and had a 100% tumor formation rate and 100% lung metastasis rate in C57BL/6J mice. CONCLUSIONS: Cross-strain transplanted tumors can be successfully established by inoculation of poorly differentiated and highly malignant tumor cells into different mouse strains. Some highly immunogenic tumor cells may form tumor, however, the tumors are regressed later, and can not establish cross-strain transplanted tumors in other mouse strains. Stable transplanted tumor models can be obtained from the partially regressed tumors after continuous passages in vivo.

[Growth inhibition of combined pathway inhibitors on KRAS mutated non-small cell lung cancer cell line].

OBJECTIVE: To investigate the effect of the selective PI3K inhibitor and MEK inhibitor on KRAS and PTEN co-mutated non-small cell lung cancer cell line NCI-H157 and the relevant mechanisms. METHODS: NCI-H157 was cultured routinely and treated with different concentrations of the two inhibitors. Cell proliferation was detected by MTT cell cycle assay. Based on the MTT results the cells were divided into four groups: the control group, PI3K inhibitor group (GDC-0941, 0.5 and 5.0 µmol/L), combination group I (0.5 µmol/L AZD6244 + 0.5 µmol/L GDC-0941) and combination group II (5.0 µmol/L AZD6244 + 5.0 µmol/L GDC-0941). Colony formation assay was performed to detect colony formation efficiency. The cell cycle and apoptosis were analyzed by flow cytometry. The expression of protein related to apoptosis was tested with Western blot. RESULTS: Cell growth was inhibited by the two inhibitors. Combination groups led to stronger cell proliferation inhibition: combination group Ishowed synergistic effect of their actions and combination group II showed an additive effect; in both groups, there were decreased colony number [(77.2 ± 1.54)/well vs (61.50 ± 2.12)/well, P < 0.01] and [(51.00 ± 4.00)/ well vs (22.50 ± 3.53)/well, P < 0.01]; and enhanced apoptotic ratios [(18.30 ± 0.82)% vs (21.32 ± 0.56)%, P < 0.01] and [(27.14 ± 1.58)% vs (42.45 ± 4.42)%, P < 0.01]. In addition, compared to the PI3K inhibitor alone group, the NCI-H157 cells in the combination groups showed increased G0/G1 phase and decreased S phase (P < 0.01). Western blotting showed that the combination groups demonstrated significantly decreased expression of cyclin D1 and cyclin B1, increased p21 and cleaved PARP and decreased bcl-2/bax ratio, compared to the PI3K inhibitor only group. CONCLUSION: The combined inhibition of PI3K (AZD6244) and MEK (GDC-0941) has synergistic effects on the proliferation of NCI-H157 cells, but such effects appear to be in a dose-dependent manner.

Identification of lactylation gene CALML5 and its correlated lncRNAs in cutaneous melanoma by machine learning.

As a product of glycolysis, lactate contributes to cancer proliferation, immunosuppression, and metastasis via histone lactylation. However, the relationship between cutaneous melanoma (CM) and lactylation-associated genes and lncRNAs has remained unclear. In this study, 4 mechanism learning algorithms and integrated bioinformatic analyses were employed to identify the core lactylation-associated genes and lncRNAs. Subsequently, 2 risk signatures based on the hub lactylation-associated genes and lncRNAs were constructed for CM patients. As a result, CALML5 was identified as a core lactylation-associated gene in CM, and its expression was found to be associated with patients survival and immune infiltration, suggesting its relevance as a potential therapeutic target. Additionally, this study provided clarification on hub CALML5-associated lncRNAs in CM, offering insights into their roles in the disease. Meanwhile, 2 identified risk signatures were both strongly linked to the prognosis and cancer growth of CM, underscoring their potential as valuable prognostic indicators. Furthermore, mechanistic analyses suggested a significant association between the risk signature and the immune microenvironment in CM, highlighting potential immune-related implications in disease progression. In conclusion, we propose that lactylation-associated genes and lncRNAs hold promise as potential targets in CM. Moreover, our findings revealed a significant correlation between lactylation and the immune microenvironment, providing crucial insights for guiding individualized treatment strategies in CM.

[Effect of forced E-cadherin expression on adhesion and proliferation of human breast carcinoma cells].

OBJECTIVE: To investigate the role that E-cadherin (E-cad) plays on cell adhesion and proliferation of human breast carcinoma. METHODS: E-cad expression vector was transfected into an E-cad-negative human breast carcinoma MDA-MB-231 cells. G418 was used to screen positive clones. E-cad, ß-catenin (ß-cat) and cyclin D1 expressions of these clones were confirmed by Western blot. Their cell-cell and cell-matrix adhesion abilities were detected. E-cad/ß-catenin interaction was confirmed by immunoprecipitation. Cell proliferation was evaluated by MTT. Cell apoptosis was analyzed by flow cytometry. Direct two-step immunocytochemistry was used to detect the localization of ß-cat. RESULT: E-cad(+) cell strains Ecad-231-7 and Ecad-231-9 were established. When cultured in ultra-low-binding dishes Ecad-231 cells grow in suspension while Ecad-231-7 and Ecad-231-9 cells grow in large clamps. When co-cultured with HCT116 cells, the average adhesion rates at 30 min are 39.0%, 60.0% and 59.5% for MDA-MB-231, Ecad-231-7 and Ecad-231-9 respectively. The average detachment rates by EDTA for 5 min are 37.4%, 4.2% and 7.4% respectively. So E-cad expression enhanced hemotypic and heterotypic cell-cell adhesion and cell-matrix adhesion. Forced exogenously expressed E-cad could combine with endogenous ß-cat, whereas down stream cyclin D1 expression was significantly decreased, as evidenced by Western blot. The rates of cell apoptosis of MDA-MB-231, Ecad-231-7 and Ecad-231-9 were 1.8%, 2.0% and 2.1%. Expression of E-cad had no obvious effect on the apoptosis of tumor cells with regular culture. ß-cat increased in the cytoplasma. CONCLUSIONS: Two monoclonal tumor cell strains (Ecad-231-7 and Ecad-231-9) stably expressing E-cad were successfully established. E-cad could enhance adhesion and inhibit proliferation of human breast carcinoma cells through a pathway involving ß-cat and cyclin D1.

In vivo longitudinal and multimodal imaging of hypoxia-inducible factor 1α and angiogenesis in breast cancer.

BACKGROUND: Angiogenesis and hypoxia-inducible factor 1α (HIF-1α) play major roles in solid tumors. This study aimed to establish a longitudinal and multimodal imaging model for in vivo evaluation of HIF1α and angiogenesis in breast cancer. METHODS: By transfection of a 5 hypoxia-responsive element (HRE)/green fluorescent protein (GFP) plasmid, the cell line Ca761-hre-gfp was established, which emitted green fluorescence triggered by HIF-1α under hypoxia. The cells were subjected to CoCl2-simulated hypoxia to confirm the imaging strategy. We grew Ca761-hre-gfp cells in the left rear flanks of twelve 615 mice. Experiments were conducted on days 4, 9, 15, and 19. For in vivo analysis, Ca761-hre-gfp subcutaneous allografted tumors were imaged in vivo using contrast-enhanced ultrasound (CEUS) and fluorescence imaging (FLI) during tumor development. The tumor size, CEUS peak intensity, and FLI photons were measured to evaluate tumor growth, angiogenesis, and HIF-1α activity, respectively. After each experiment, three mice were randomly sacrificed and tumor specimens were collected to examine HIF-1α activity and the microvessel density (MVD). RESULTS: In vitro, both green fluorescence and HIF-1α expression were detected in Ca761-hre-gfp cells treated with CoCl2, indicating the suitability of the cells to detect HIF-1α activity. In vivo, HIF-1α activity first increased and then decreased, which was significantly correlated with angiogenic changes (r = 0.803, P = 0.005). These changes were confirmed by immunohistochemical staining of HIF-1α and MVD. CONCLUSIONS: The findings validated the Ca761-hre-gfp murine allograft model for reliable evaluation of HIF-1α activity and angiogenesis longitudinally using both molecular and pre-clinical non-invasive imaging modalities. The cell line may be useful for studies of anti-HIF pathway therapies.

[Expression and function of VAP-33 in murine dendritic cell sarcoma].

OBJECTIVE: To elucidate the expression and function of VAP-33 gene in dendritic cell sarcoma (DCS) cell line. METHODS: The expression of VAP-33 in DCS cells was investigated by mass spectrum with immunoprecipitation membrane protein. DCS cells were treated with antigens in different dosages (150, 850, and 1500 microl) for 24, 48 and 72 h respectively. Cell morphology and phagocytosis activity of DCS cells were measured. Indirect immunofluorescence, confocal microscopy and Western blotting were used to study the distribution and expression changes of VAP-33. Moreover, DCS cells were treated with 0.5 mol/L insulin for 20 min first and followed by Western blotting to detect changes of VAP-33 and glucose transfer protein 4 (GLUT-4) in the total cellular protein, cytoplasmic protein and membrane protein. Confocal microscopy was used to document the expression and distribution changes of VAP-33 and GLUT-4 in DCS cells. RESULTS: VAP-33 expression was obtained at the cell membrane and in the cytoplasm of DCS cells. Upon antigen stimulation, DCS cells showed more active phagocytosis and morphologically became more elongated with branched protrusions. The expression of VAP-33 was decreased by the antigen stimulation. Upon the insulin stimulation, the expression of VAP-33 and GLUT-4 were increased and co-localized. CONCLUSIONS: VAP-33 expression in DCS originated from the dendritic cells (DC) seemed relating to the vesicle transportation during antigen processing in DC. Additionally, VAP-33 and GLUT-4 also take part in the glucose transportation in the cells.

[Establishment of green-fluorescent protein expressing tumor metastasis models].

OBJECTIVE: To establish a green-fluorescent protein (GFP) labeled tumor metastasis model and to evaluate its biological characteristics. METHODS: Human gastric carcinoma cell MGC-803 and murine cervical carcinoma cell U14 were transfected with the plasmid pEGFP-N1 and the efficiency of transfection was assessed 24 h later. Limited dilution was employed to screen and establish monoclonal cell strains, MGC-803-GFP and U14-GFP. The two fluorescent tumor cell stains were transplanted into BALB/c-nu mice and C57BL/6J mice respectively. The latency period of tumor mass appearance and the growth curve in vivo were documented. The tumor growth and metastasis were evaluated in vivo by the Viviperception Fluorescence Imagining System (VFIS). Expressions of CD44 and E-cadherin in tumor tissue were monitored by immunohistochemistry. RESULTS: The efficiency of pEGFP-N1 transfection of MGC-803 cells and U14 cells were 30% and 60%, respectively. Monoclonal GFP(+) cell strains-MGC-803-GFP and U14-GFP were established. The latency periods of tumor formation of MGC-803-GFP and U14-GFP were 3-5 days and 2-4 days, respectively. Their tumorigenicity rates were 100% in both. The tumor growth of MGC-803-GFP was well defined by the VFIS. Only one mouse was shown to harbor lymphatic metastasis by VFIS, 60 days after transplantation. The metastasis process of U14-GFP was depicted through VFIS on 27, 37 and 52 days post-transplantation. The incidence of pulmonary metastasis and lymphatic metastasis of U14-GFP was 67% and 100% respectively when the tumor volume was >or=5 cm3. CD44 was positive and E-cadherin was negative in both tumors by immunohistochemistry. CONCLUSIONS: Successfully established two monoclonal tumor cell strains stably expressing GFP: MGC-803-GFP and U14-GFP. Transplantation of these cells into mice can establish tumor metastasis models which could be used for future visualized tumor research in vivo.

Assessment of the osteogenic effect after maxillary sinus floor elevation and simultaneous implantation with or without bone grafts by analyzing trabecular bone parameters: a retrospective study

Abstract Objective: The aim of this population-based retrospective study was to compare the osteogenic effect of newly formed bone after maxillary sinus floor elevation (MSFE) and simultaneous implantation with or without bone grafts by quantitatively analyzing trabecular bone parameters. Methodology: A total of 100 patients with missing posterior maxillary teeth who required MSFE and implantation were included in this study. Patients were divided into two groups: the non-graft group (n=50) and the graft group (n=50). Radiographic parameters were measured using cone beam computed tomography (CBCT), and the quality of newly formed bone was analyzed by assessing trabecular bone parameters using CTAn (CTAnalyzer, SkyScan, Antwerp, Belgium) software. Results: In the selected regions of interest, the non-graft group showed greater bone volume/total volume (BV/TV), bone surface/total volume (BS/TV), trabecular number (Tb. N), and trabecular thickness (Tb. Th) than the graft group (p<0.001). The non-graft group showed lower trabecular separation (Tb. Sp) than the graft group (p<0.001). The incidence of perforation and bleeding was higher in the graft group than in the non-graft group (p<0.001), but infection did not significantly differ between groups (p>0.05). Compared to the graft group, the non-graft group showed lower postoperative bone height, gained bone height and apical bone height (p<0.001). Conclusion: MSFE with and without bone grafts can significantly improve bone formation. In MSFE, the use of bone grafts hinders the formation of good quality bone, whereas the absence of bone grafts can generate good bone quality and limited bone mass.

[Effect of inhibitor of differentiation-1 on murine dendritic cell sarcoma cells].

OBJECTIVE: To investigate the effect of down-expression of inhibitor of differentiation-1 (Id-1) on the differentiation of dendritic cell sarcoma (DCS) cells in vitro. METHODS: Down-regulation of the expression of Id-1 in DCS cells was performed by RNAi, and confirmed by protein and mRNA quantitative analyses. Cellular differentiation and biological behavior including malignant phenotypes of the cells were evaluated. All experiments included negative (no treatment group and no-target siRNA) and positive (induction-differentiation drug sodium butyrate) controls. RESULTS: When the expression of Id-1 was down regulated, the DCS cells showed more mature morphology including cell enlargement, longer cellular extensions, more branches, and decreased nuclear/plasma ratio. Differentiation marker expression (Id-2 and CD86) was also increased. RNAi treated cells at 24 and 48 hours, showed increase percentage of cells at G0/G1 phase and less cells at S phase (P < 0.01). Importantly, the abilities of cell proliferation, colony formation and invasiveness were significantly decreased (P < 0.01), as evidenced by MTT, colony formation and transwell assays respectively. CONCLUSION: RNAi inhibition of Id-1 protein can induce differentiation of malignant solid tumor cells along with reversion of their malignant phenotype.

Angiogenesis Research in Mouse Mammary Cancer Based on Contrast-enhanced Ultrasonography: Exploratory Study.

RATIONALE AND OBJECTIVES: The objective of this study was to investigate the contrast-enhanced ultrasound (CEUS) characteristics of tumor angiogenesis in mouse mammary cancer. MATERIALS AND METHODS: Twenty-four mice were examined with ultrasound and CEUS at 2-12 days after implantation. Four to five mice were assessed daily, and one to three mice were then sacrificed for histology. All of the histologic slides were reviewed and correlated with CEUS findings. RESULTS: A total of 46 cases of ultrasound examination had been performed in 24 mice. The mice were classified into three groups according to the tumor growth: group 1 (2~6 days after implantation, n = 20 cases), group 2 (7~9 days after implantation, n = 15 cases), and group 3 (10~12 days after implantation, n = 11 cases). In group 1, all tumors presented as a homogeneous hypoechoic mass with no color Doppler signals. However, three CEUS patterns were observed: 14 tumors presented as type I (peripheral ring enhancement with no enhancement within the tumor), 4 tumors presented as type II (peripheral ring enhancement with deep penetration), and 2 tumors presented as type III (homogeneous or heterogeneous enhancement in the entire tumor). In group 2, there was only difference in the echo (heterogeneous or not) and color Doppler signals (with or without) among the tumors in conventional ultrasound, but four CEUS patterns were observed and most presented as type III (53.3%, 8/15). In group 3, most tumors presented as a heterogeneous solid mass (81.8%, 9/11) with color signals (100%, 11/11), and almost all tumors presented as enhancement of type IV (peripheral ring enhancement with focal nodular enhancement) (90.9%, 10/11).The histologic results showed that the enhanced areas mainly corresponded to tumor cells, large tortuous vessels, and an inflammatory cell infiltrate. Nonenhanced areas corresponded to large areas of necrotic tissue or tumor cells, which arranged loosely with the small zone of necrosis. CONCLUSIONS: CEUS could image the progression of vessel formation. Moreover, most importantly, CEUS is able to identify angiogenesis before the change of tumor color Doppler, and presents different enhanced patterns at different tumor growth times, which corresponded to tumor histologic features.