Objective To investigate sequential expression of bone sialoprotein(BSP), alkaline phosphatase(ALP) and cementum attachment protein(CAP) in cementoblast(CB) during mineralized culture in vitro, and study the morphological and biologic characters of the CB in this process. Methods CB was seeded on the glass coverslips, and cultured for 6h, 12h, 1d, 2d, 3d, 4d, 5d, 6d, 8d, 10d, 12d and 16d, respectively. The expression of BSP, ALP and CAP proteins was detected using immunocytochemical method. Results 6 hours after plated, cementoblast expressed all of the three proteins. In the second and third days after plated, the cells became confluent and formed multiple layers, BSP and ALP expression decreased, and CAP did not be expressed at all. From the forth day, the cells formed cell nodules with mineralizing function. The cells in the nodules strongly expressed BSP,ALP and CAP, the cells around the nodule weakly expressed BSP and ALP, and did not express ALP. During the following 10 to 16 days, the cell nodules became mineralized nodules. Conclusion These results elucidated the changes of ALP, BSP and CAP expression as well as cell morphology during the CB proliferation, differentiation and mineralization in vitro, and provided some valuable information for studying the formation of cementum and CB proliferation, differentiation and mineralization in vivo.

Objective To study the ultrastrueture of 3 types of periodontal cells, including periodontal ligament cells (PDLC), os-teoblast cells (OB) and eementoblast cells (CB). Methods After culturing 3 kinds of periodontal cells - PDLC, OB and CB, we observed them with transmission electron microscope. Results There was rich rough endoplasmic reticulum and lots of microfilaments in the cyto-plasm of CB and PDLC cells. There was rich rough endoplasmic in the cytoplasm of OB cells. Conclusion The main characteristic ultra-structure feature of the bovine CB and PDLC was rich rough endoplasmic reticulum and microfilaments in the cytoplasm. Compared with CB and PDLC, OB contained fewer microfilaments in the cytoplasm.

Objective:To study the role of 14-3-3εand Cdc25B in germinal vesicle (GV)-stage arrest of mouse oocytes,and to pay foundation for further study on the molecular mechanism of PKA/Cdc25B/14-3-3εpathway in GV-stage arrest of mouse oocytes.Methods:The eukaryotic expression vectors of pcDNA3.1-ZEO-HA-14-3-3ε, pcDNA3.1-MYC-Cdc25B-WT, pcDNA3.1-MYC-Cdc25B-S321A, and pcDNA3.1-MYC-Cdc25B-S321D were transcribed into mRNA invitro.The mouse GV-stage oocytes were collected after superovulation and divided into no injection group,TE buffer microinjection group,14-3-3εmRNA injection group,14-3-3εmRNAs + Cdc25B-WT mRNA injection group,and 14-3-3εmRNA + Cdc25B-S321A mRNA injection group,14-3-3εmRNA+Cdc25B-S321D mRNA injection group.The protein expression levels of HA-14-3-3εand MYC-Cdc25B and the phosphorylation status of Cdc2-pTyr15 were observed by Western blotting method.The morphological changes and germinal vesicle breakdown (GVDB)rates of mouse oocytes were observed under phase-contrast microscope. Results:None of the oocytes in no injection group, TE buffer microinjection group, 14-3-3εmRNA injection group,14-3-3εmRNA + Cdc25B-WT mRNAs injection group and 14-3-3εmRNA + Cdc25B-S321D mRNA were able to undergo GVBD until at least 20 h after injection (P>0.05 );the GVBD rates of oocytes in 14-3-3εmRNA+Cdc25B-S321A mRNA group at 1 h (5.00%±0.68%),2 h (62.00%±3.56%)and 3 h (100.00%± 0.00%)after injection were significantly higher than those in no injection group and TE buffer injection group (P<0.01);the oocytes in 14-3-3εmRNA+ Cdc25B-Ser321A mRNA group at 20 h (79.00%±2.80%)after injection progressed to MII (P<0.01).Conclusion:14-3-3εcan regulate the transition from GV to GVBD of mouse oocytes by means of phosphorylation and dephosphorylation of S321-Cdc25B.

Objective: Epithelial dysplasia of the esophagus and gastric cardia is precancerous lesion, including mild, moderate and severe levels. In 2000 year, WHO recommended to replace dysplasia with intraepithelial neoplasia. Mild and moderate dysplasia were classified as low-grade intraepithelial neoplasia (LIN). Cardia adenocarcinoma was suggested to be called esophageal-gastric junction adenocarcinoma. The risk of cancer development and the rule of time evolution were detected in esophagus and esophageal-gastdc junction LIN in high incidence area of esophageal cancer in Northern China, in an effort to provide scientific data for the prevention of esophageal cancer. Methods: Between October 2001 and October 2002, two townships of Cixian were chosen to carry out endoscopic iodine staining screening cohort study. The total population aged 0-85 was 22,016, of which 6,596 aged 40-69 (3257 males and 3339 females). Except for thoese with contraindications and those who refused to join the study, 3,506 cases were finally recruited in the study, and the screening rate was 53.2%. According to WHO criteria of the pathological diagnosis, the esophageal squamous epithelium with mild and moderate dysplasia and esophageal-gastric junction with mild dysplasia were classified into LIN groups (including 616 cases). The control group contained a total of 2,478 cases without precancerous lesions and free of cancer in endoscopic screening. Results: From June to September in 2008, the cohort was followed up and 174 cases were lost, with a follow-up rate of 95.0%. Follow-up was 3,970.7 person- years in the LIN group and 16,120.0 person-years in the control group.Carcinomous conversion rates were 251.7 and 68.2/per 100,000 person- years respectively in the LIN group and the control group. The median time in the two groups was 38 and 47 months, respectively. Compared with that of the normal population, the relative risk (RR) of LIN was 3.69 (95% CI=1.57-8.69, P=0.001). Conclusion: Population with LIN are at high-risk for esophageal cancer and endoscopic examination every year is absolutely necessary.

Alveolar Epithelial Cells ; pathology ; Diagnosis, Differential ; Humans ; Hyperplasia ; diagnostic imaging ; etiology ; metabolism ; pathology ; Lung Diseases ; diagnostic imaging ; etiology ; metabolism ; pathology ; Mucin-1 ; metabolism ; Nuclear Proteins ; metabolism ; Radiography ; Thyroid Nuclear Factor 1 ; Transcription Factors ; metabolism ; Tuberous Sclerosis ; complications ; diagnostic imaging ; metabolism ; pathology

Objective To assess the continent diversion results of sigma rectum pouch after radical cystectomy. Methods The reconstruction of bladder with sigmoid was modified for treatment of 18 cases of bladder tumor.The intestine was incised over a length of 20～24 cm with the junction of sigmoid colon and rectum as the midpoint so as to create a low pressure reservoir for urine and side-to-side anastomosis was performed on the posterior borders of the rectosigmoid wall.Submucosal tunnel modified technique was em- ployed in antireflux urethral implantation,Urination has been controlled by anal sphincter.Results About 80 minutes was spent to finish a new low pressure pouch after radical cystectomy.Among 18 patients with this op- eration,the controlled emiction were good after pull out the anal duct and"J"stent in 1 week to 2 months.Af- ter 2 months,the times of urination is stable,4～5 times in daytime and 1～3 times during nighttime.Two pa- tients had nocturnal enuresis and the symptom vanished after 2 months. One patient had adhesive ileus, two patients had hyperchloremia acidosis and kaliopenia,one patient had urethral stump cancer.There is no com- plication as anastomotic block,renal function lesion and severe upper urinary tract infection. Conclusion This operative method was easy,emiction control was well,and with higher quality of life for patients.It is al- so a better alternative diversion procedure that would be easily accepted.

OBJECTIVETo investigate the optimum proportion of Mosaicplasty and genes-enhanced tissue engineering for the repair of acute osteochondral defects.

METHODSWestern blot test was conducted to detect the expression of hTGF-beta1, Col II and Aggrecan in 3 groups, including hTGF-beta1, transduction group, Adv-betagal transduction group and control group without transduction. Eighteen 6-month-old Shanghai male goats (weight: 22 to 25 kg) were used. BMSCs were isolated from the autologous bone marrow, and were subcultured to get the cells at passage 3. Thirty-six medial femoral condyles were used and divided into 6 groups named AR, AL, BR, BL, CR, and CL. Acute cylindrical defects (9 mm in diameter and 3 mm in depth)were created in the weight bearing area of the medial femoral condyle of hind limbs. In the single group, the autologous osteochondral mosaicplasty was performed to repair the defect; in the combination group, besides the mosaicplasty, the dead space between the cylindrical grafts and the host cartilage were injected with the suspension of hTGF-beta1, gene enhanced autogenous BMSCs in sodium alginate, and CaCl2 was dropped into it to form calcium alginate gels. The autologous osteochondral transplantation cover rates of group AR was 44.44% single group, AL was 44.44% combination group, BR was 33.33% single group, BL was 33.33% combination group, CR was 22.22% single group, and CL was 22.22% combination group. The goats were killed 24 weeks after operation to receive gross and histology observation, which was evaluated by the histological grading scale of O'Driscoll, Keeley and Salter. Immunohistochemistry and TEM observation were also performed.

RESULTSWestern blot test showed the expression of the hTGF-beta1, Col II and the Aggrecan in the hTGF-beta1 transduction group were significantly higher than that of the Adv-betaga1 transduction and the blank control groups. The gross and histology observation revealed that each defects of six groups had different degrees of repairing. There was no significantly difference among the BL, AR, and AL groups. But the scores of the other three groups (BR, CR, and CL) were significantly poorer than the former three groups.

CONCLUSIONMosaicplasty associated with genes enhanced tissue engineering could repair the osteochondral defects effectively. With the autologous osteochondral transplantation coverage reducing, the advantage of the combination could have a better representation.

Animals ; Blotting, Western ; Bone Diseases ; metabolism ; pathology ; therapy ; Cell Line ; Goats ; Humans ; Immunoprecipitation ; Male ; Tissue Engineering ; methods

Objective: To observe morphological changes of epithelial-mesenchymal transition in the early stage of mice renal interstitial fibrosis. Methods: Renal interstitial fibrosis was induced by unilateral ureteral obstruction(UUO) in mice. Histological and immunohistochemical methods were used to analyze pathological changes and α-SMA expression in renal tissue.Argentum hexamethylenamine staining and transmission electron microscopy were used to observe changes of the renal tubule basement membrane. Gelatin zymographic analysis was used to observe the expression of MMP2 and MMP9 in renal tissue.Results:The mice suffered from renal interstitial fibrosis were identified by histological analysis and α-SMA positive cells in renal tissue. Argentum hexamethylenamine staining and transmission electron-microscopy showed that the renal tubule basement membrane disrupted locally and renal tubule epithelial cells invaded into the renal interstitium in the early stage of renal interstitial fibrosis. Gelatin zymographic analysis showed that the expression of MMP2 and MMP9 was increased transitorily in the early stage of renal interstitial fibrosis. Conclusion: Renal tubule basement membrane disruption, renal tubule epithelial cells invasion into the renal interstitium, and the expression of MMP2 and MMP9 are involved in the development of renal interstitial fibrosis.

Objective:To explore the effect of 14-3-3εprotein on the localization of Cdc25B protein during the meiotic resumption of mouse oocytes,and to pay foundation for the further study on the molecular mechanism of 14-3-3εprotein in regulating the development of mouse oocytes.Methods:The Kunming genealology female mice aged 3 weeks were used to obtain the germinal vesicle (GV)-stage oocytes after superovulation.The GV-stage oocytes were divided into non-injection group,control siRNA injection group and 14-3-3εsiRNA injection group. The pmax-FP-Red-HA-14-3-3εexpression vector was constructed.Indirect immunofluorescence was used to observe the colocalization of 14-3-3εprotein and Cdc25B protein in the mouse oocytes;direct immunofluorescence was used to observe the subcellular localization of 14-3-3εprotein and Cdc25B protein in the mouse oocytes;14-3-3εsiRNA was microinj ected into the GV-stage oocytes;the morphology was observed under phase-contrast microscope;the germinal vesicle breakdown (GVDB)rates of the mouse oocytes were calculated;the expression level of 14-3-3εprotein and the relative expression level of Cdc2-pTyr15 protein were observed by Western blotting method;the matuation-promoting factor (MPF)activity in the oocytes was measured by autoradiography.Results:The indirect immunofluorescence and direct immunofluorescence results showed that the 14-3-3εprotein and wild Cdc25B protein were co-localized in the cytoplasm;Cdc25B was translocated from the cytoplasm to the nucleus shortly before GVBD.When the Ser321 of Cdc25B protein turned into Ala,the expression level of 14-3-3εprotein was decreased. None of the oocytes in non-injection group and control siRNA injection group were able to undergo GVBD until at least 24 h after injection,there was no significant differences in the rate of GVBD between non-injection group and control siRNA injection group (P>0.05);the GVBD rates of oocytes in 14-3-3εsiRNA injection group at 22 and 24 h after injection were significantly higher than those in non-injection group and control siRNA injection group (P<0.01);the rate of oocytes progressed to metaphaseⅡ (MII)in 14-3-3εsiRNA injection group at 24 h after injection was significantly higher than those in non-injection group and control siRNA injection group (P<0.01). Conclusion:Ser321 might be involved in the process of regulating the subcellular localization of Cdc25B by 14-3-3εprotein in the meiotic resumption of mouse oocytes.

OBJECTIVETo investigate the effects of oxidized low-density lipoprotein receptor 1 (LOX-1) on secretion of adhesive molecules mediated by ox-LDL in human umbilical endothelial cells (HUVECs).

METHODSHUVECs with different concentration of ox-LDL (0, 10, 20, 50, 100 microg/ml) were incubated for 24 h, or HUVECs were pretreated with 250 microg/ml poly (I) or 250 microg/ml carrageenan for 2 h and then incubated with 50 microg/ml ox-LDL for another 24 h. Expression of LOX-1 was determined by realtime RT-PCR and Western blot. mRNA and protein of ICAM-1, VCAM-1 and E-selectin were examined by RT-PCR and Western blot respectively.

RESULTSIncubation of HUVECs with ox-LDL (10-100 microg/ml) enhanced the expressions of LOX-1, ICAM-1 and E-selectin in a concentration-dependent manner (P < 0.01). On the contrary, ox-LDL did not affect the expression of VCAM-1 by HUVECs. The expression of LOX-1, ICAM-1 and E-selectin induced by ox-LDL were reduced in HUVECs pretreated with 250 microg/ml poly (I) or 250 microg/ml carrageenan for 2 h and then incubated with 50 microg/ml ox-LDL for 24 h. This showed that both poly (I) and carrageenan obviously decreased the expression of LOX-1, ICAM-1 and E-selectin induced by ox-LDL.

CONCLUSIONox-LDL may upregulate the expression of LOX-1, ICAM-1 and E-selectin, and LOX-1 blocker may partly inhibit this upregulation. The results suggest that the expression of inflammatory molecules induced by ox-LDL in HUVECs is mediated by LOX-1.

Cell Adhesion ; Cell Adhesion Molecules ; Cells, Cultured ; E-Selectin ; metabolism ; Endothelial Cells ; metabolism ; Endothelium, Vascular ; metabolism ; Humans ; Intercellular Adhesion Molecule-1 ; metabolism ; Lipoproteins, LDL ; biosynthesis ; RNA, Messenger ; metabolism ; Receptors, Oxidized LDL ; metabolism ; Scavenger Receptors, Class E ; metabolism ; Umbilical Veins ; cytology ; Vascular Cell Adhesion Molecule-1 ; metabolism