A new abietane diterpene and anti-complementary constituents from Juniperus tibetica.

Anti-complementary activity-guided fractionation led to the isolation of a new abietane diterpene (1) and twenty-five known compounds (2-26) from the twigs and leaves of Juniperus tibetica. All the compounds were isolated from J. tibetica for the first time. The structure of 1 was assigned by spectroscopic data and X-ray crystallography analysis. Five lignans (2, 3, 7, 9 and 10), two flavones (19 and 22), and one coumarin (23) exhibited anti-complementary activity with CH50 values ranging from 0.3 to 3.69 mM.

Regulating the balance of Th17/Treg cells in gut-lung axis contributed to the therapeutic effect of Houttuynia cordata polysaccharides on H1N1-induced acute lung injury.

Our previous study had demonstrated that oral administration of Houttuynia cordata polysaccharides (HCP) without in vitro antiviral activity ameliorated gut and lung injuries induced by influenza A virus (IAV) in mice. However, as macromolecules, HCP was hard to be absorbed in gastrointestinal tract and had no effect on lung injury when administrated intravenously. The action mechanism of HCP was thus proposed as regulating the gut mucosal-associated lymphoid tissue (GALT). Actually, HCP treatment restored the balance of Th17/Treg cells firstly in GALT and finally in the lung. HCP reduced the expression of chemokine CCL20 in the lung and regulated the balance of Th17/Treg carrying CCR6+ (the CCL20 receptor), which was associated with specific migration of Th17/Treg cells from GALT to lung. In vitro, HCP inhibited Th17 cell differentiation through the downregulation of phospho-STAT3, whereas it promoted Treg cell differentiation by upregulating phospho-STAT5. Furthermore, its therapeutic effect was abolished in RORγt-/- or Foxp3-/- mice. These findings indicated that oral administration of macromolecular polysaccharides like HCP might ameliorate lung injury in IAV infected mice via directly regulating the balance of Th17/Treg cells in gut-lung axis. Our results provided a potential mechanism underlying the therapeutic effect of polysaccharides on pulmonary infection.

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.

[Suppression to ameloblastoma xenografts of chicken embryo chorioallantoic membrane by tissue inhibitor of metalloproteinases-2].

OBJECTIVE: To explore the invasiveness of xenografts on chicken embryo chorioallantoic membrane (CAM) after tissue inhibitor of metalloproteinase-2 (TIMP-2) gene transfection. METHODS: Fresh ameloblastoma tissues were minced into 1-2 mm3 and transplanted on the CAM. There were three groups named as control group (Empt), plasma transfection group (Lipo), and TIMP-2 gene transfection group (P). The specimens were respectively investigated by microscope indifferent spots after implanting. The volume of the xenografts and the weight of xenografts in the termination time of the experiment were recorded. The invasiveness of xenografts was divided into four grades by pathological examination. Western blot analysis was performed to investigate matrix metalloproteinase-2 (MIMP-2) and TIMP-2 protein in xenografts. RESULTS: Ameloblastoma tissues can survive on CAM and the tumor cells may invade it on 5-7 days after implanting. At 9 d after implanting, the invasiveness grades in P group were 7 in grade 0, 1 in grade 2, 0 in grade 3. The expression of TIMP-2 protein in P group was significantly higher than that in Empt group (P < 0.05). The expression of MMP-2 protein in P group was lower than that in Empt group (P < 0.05). CONCLUSION: The xenotransplanted tumor model of human ameloblastoma on CAM was successfully established. The invasiveness of ameloblastoma xenografts was suppressed might be due to TIMP-2 gene transfection.

Chronological age estimation of third molar mineralization of Han in southern China.

In order to study the chronology of age of third molar mineralization of Han in southern China, Demirjian staging method was used to determine the stage of four third molars (18, 28, 38, 48) mineralization in 3,100 Han in southern China aged 4.1-26.9 years based on radiological evidence from digital orthopantomograms. The mean age of the 3,100 patients was 15.96 +/- 4.73 years, including 1,200 male (mean age, 15.32 +/- 4.62) and 1,900 female (mean age, 16.35 +/- 4.76). Results show that there was no significant difference in mineralization between 18 and 28 and 38 and 48 of male or female. However, significant difference was observed between 28 and 38 of female at stage C; 28 was 0.25 years earlier than 38. In male, at stage G, 38 was 0.61 years earlier than 28, and 48 was 0.62 years earlier than 18. At stages D, E, F, G, and H, male 48 was 0.34, 0.66, 0.72, 1.34, and 0.76 years earlier than that of female, respectively. At stages A, D, E, F, G, and H, male 38 was 0.73, 0.26, 0.56, 0.91, 1.29, and 0.70 years earlier than that of female, respectively. At stages B, E, F, G, and H, the mineralization mean age of male 18 was 0.54, 0.50, 0.76, 0.92, and 0.58 years earlier than that of female, respectively. At stages E, F, G, and H, the mineralization mean age of male 28 was 0.51, 0.76, 0.92, and 0.49 years earlier than that of female, respectively. After reviewing the literature, the chronological mineralization age of 48, at stages D to G, of Han in southern China was 1 to 4.6 years earlier than that of Japanese and 1 to 3 years earlier than that of German. The mean age at stage H of 48 of Han in southern China was similar to Turkish, Black African, Japanese, and German, but was later than Spanish. Finally, the conclusions are: (1) in the same gender group of Han in southern China, the mineralization ages between two sides in upper or lower jaw are very similar, and (2) the chronology mean age and complete time of third molar mineralization of male were earlier than that of female.

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.

[Establishment of subcutaneous xenotransplanted tumor model of human ameloblastoma in nude mice].

OBJECTIVE: To establish an subcutaneous xenotransplanted tumor model of human ameloblastoma in nude mice. METHODS: Ameloblastoma cells were absorbed by primary culture, repeat attachment and pancreas proteolytic enzyme were both used to purify them. Then, the purified cells were implanted subcutaneously into the nude mice. The specimens were respectively investigated by microscope in different spots after implanting. RESULTS: Ameloblastoma cells can survive in all of the 8 nude mice. The xenograft can be found on 23 days after implanting. The rate of successful inocutation is 25%. The subcutaneously xenotransplanted tumor cells can be found with microscope in the inter-muscle tissues of nude mice. CONCLUSION: The subcutaneously xenotransplanted tumor model of human ameloblastoma in nude mice was successfully established and it may benefit to further studies on this tumor.

[The construction and expression of PcDNA3.1(+)/GFP-TIMP-2 in human ameloblastoma cell].

OBJECTIVE: To construct the eukaryotic expression vector of TIMP-2 gene and to explore its expression in human ameloblastoma cell in vitro. METHODS: The aimed gene fragment was obtained by RT-PCR. And then, molecmicrolar cloning technology and enzyme digestion were used to connect the gene with the plasmid PcDNA3.1(+), which can be expressed in eukaryotic cells and a report gene: green fluorescent protein gene (GFP) was already existed in the plasmid. We named the eukaryotic expression vector, which contended our aimed gene TIMP-2 as well as report gene GFP, PcDNA3.1(+)/GFP-TIMP-2. The vector was identified by PCR analysis, EcoR I and Xho I restriction analysis and Sequence analysis. After the PcDNA3.1(+)/GFP-TIMP-2 was transfected into cultured human ameloblastoma cell, RT-PCR and Flow Cytometry (FCM) and Microscope wre respectively performed to evaluate the effect of transfection and expression. RESULTS: The constructed vector PcDNA3.1(+)/GFP-TIMP-2 was proved correct by enzyme digestion and sequencing analysis. After PcDNA3.1(+)/GFP-TIMP-2 was trasnfected into cultured human ameloblastoma cell, the rate of transfection is 47.6% (Analysis report of FCM), the green fluorescence was found in plasm (observed with fluo-microwave), the expression of TIMP-2 mRNA was elevated 2.4 times compared with the control group. CONCLUSIONS: PcDNA3.1(+)/GFP-TIMP-2 was successfully constructed and it could be transfected into cultured human ameloblastoma cell. It may be benefit to further study of the relationship between the TIMP-2 gene and the behaviour of ameloblastoma.