OBJECTIVETo determine the effect of type I transmembrane protein (IRE1alpha) deletions on the cell cycle of human periodontal ligament fibroblasts (hPDLFs) cells.

METHODSBased on the IRE1alpha deletions, a full-length model was successfully constructed. Moreover, overlapping polymerase chain reaction mutagenesis facilitated the establishment of two deletion mutants of IREla (pD-Kinase, pD-Rnase). The full-length model and two mutant eukaryotic expression vectors were transfected into hPDLFs cells. Western blot analysis was performed to identify the expression in the cells. The changes in the cell cycle of hPDLFS cells were detected by flow cytometry (FCM).

RESULTSThe two deletion mutants of IRE1alpha with eukaryotic expression vectors were successfully constructed and correctly expressed in hPDLFs cells based on Western blot analysis. Under stress conditions, the FCM assay showed that cell percentage of S phases increased, whereas that of G1 phases decreased in the IRE1alpha group (P < 0.05) compared with the control group of tunicamycin (TM) treatment. Moreover, the cell percentage of the S phases decreased, whereas that of the G1 phases increased in the D-Rnase group (P < 0.05) compared with the control. The deletion mutant D-Kinase had no influence on hPDLFS cell proliferation and cycle (P>0.05).

CONCLUSIONUnder stress conditions, IRE1alpha can improve the cell cycle of hPDLFs cells from the G1 to the S phase. The deletion mutant D-Rnase cause hPDLFs cell growth arrest at the G1 phase, whereas deletion mutant D-Kinase has no significant effect.

Cell Cycle ; Cell Proliferation ; Endoribonucleases ; Fibroblasts ; Humans ; Periodontal Ligament ; Protein-Serine-Threonine Kinases ; Transfection

OBJECTIVETo investigate the effect of insulin-like growth factor- I (IGF- I) on the proliferation and alkaline phosphatase (ALP) activity of human periodontal ligament cells (hPDLCs) under three-dimensional (3D) culture system.

METHODSThe hPDLCs were cultured from periodontium of human teeth by the outgrowth method. Rotary cell culture system (RCCS) was enrolled to set 3D culture system. Samples were set to four groups: Negative control group, positive control group (3D group, IGF-I group), and experimental group (3D with IGF- I group). Proliferation was tested with methylthiazolyl tetrazolium (MTT), and ALP activity was assayed by spectrophotometer at 1, 3, 5, 7 d respectively.

RESULTSCompared with that of negative control group, cell proliferation increased significantly in 3D with IGF-I group since 3 d (P < 0.05). Besides, the cell proliferation of 3D with IGF-I group was significantly higher than that of 3D group (P < 0.05). ALP activity of 3D with IGF- I group was significantly higher than that of negative control group, and 3D group at 3, 5, 7 d (P < 0.05).

CONCLUSIONIGF-I significantly promotes the proliferation and ALP activity of hPDLCs under 3D culture system.

Alkaline Phosphatase ; Cell Differentiation ; Cell Proliferation ; Cells, Cultured ; Humans ; Insulin-Like Growth Factor I ; Periodontal Ligament ; Somatomedins

To explore a new lyophilized preservation methods for human platelets, platelets were pre-treated with aldehyde, human albumin or trehalose was added to the system of condensed cooling as protectant to stabilize the structure of platelets. The optimal resuspending buffer was also selected in the study. The morphological changes of platelets were observed by using electron microscopy after lyophilization, and the expression of membrane proteins on platelets was detected also after lyophilization. The results indicated that the recovery rate of platelets treated with aldehyde was generally more than 60%. Aggregative ability was reduced a little than the platelet untreated. 5% of human albumin had an advantage over 40 mmol/L of trehalose in respect of the preservation effect. In the way of keeping aggregative ability, PPP was obviously better than PBS. The results of electron microscopy displayed that organelles including mitochondria and excreted granules could be observed distinctly. Whereas, expression of membrane proteins of platelet treated with aldehyde was evidently dropped as compared with those of the fresh platelet. In conclusion, aldehyde as a novel protective agent, has excellent effects on lyophilization of platelets and is worthy to be further studied.
Albumins ; pharmacology ; Aldehydes ; pharmacology ; Blood Platelets ; cytology ; drug effects ; Blood Preservation ; methods ; Freeze Drying ; Humans ; Reproducibility of Results ; Trehalose ; pharmacology

Objective To investigate the therapeutic effect of modified dorsal metacarpal artery island flap of index finger without skin graft in repairing different types of soft tissue defect of thumb tip.Methods From August,2015 to October,2017,25 patients with soft tissue defect of the thumb tip were included in the study.Sixteen cases were males and 9 cases were females.Seventeen defects were in thumb dorsal and 8 cases were in thumb pulp.A modified dorsal island flap of index finger was used and the dorsal metacarpal superficial vein fascial flap could be harvest and combined to repair the thumb pulp defect if necessary.A relaying perforator flap pedicled on the second dorsal metacarpal artery was raised through the same incision to cover the donor site without skin graft.Followed-up was made by clinic,telephone and WeChat.Results The patients were followed-up for 6 to 18 months.All flaps survived completely without complications.The color,texture and contour of the flaps was good.Only 1 linear scar was left in the dorsum of the hand and no skin grafts.The second dorsal metacarpal artery flap was used to cover the donor site.Twenty-one cases (84％) were satisfied with the postoperative appearance of the thumb.The function was assessed as excellent in 16 fingers,good in 6 fingers and fair in 3 fingers.No complication occurred in the donor site.Conclusion It is possible to use the modified dorsal island flap of index finger to repair different types of thumb tip defects.A second dorsal metacarpal artery flap can be used as a relaying flap to cover the donor site without skin grafts.

OBJECTIVETo explore the CT angiography features of the involved arterial branches of the spontaneous isolated superior mesentery artery dissection(SISMAD).

METHODSClinical and CT angiogram data of 6 patients with SISMAD confirmed by dual source energy CT angiography were analyzed retrospectively. The CT angiography features and prognosis of patients were investigated.

RESULTSAverage age of 6 patients was(45.83±5.60) years old. Ten branches of jejunoileal artery were involved in 4 patients. All the dissections of the superior mesenteric arteries(SMA) were extended from its proximal to its end. Intramural hematoma was found in all the involved branches, which showed eccentric target sign in the transverse section and sandwich sign in the longitudinal axial section. In 7 branches of 3 patients, the involved branches originated from the false lumina of the SMA. In one patient, the intramural hematoma of the jejunoileal arteries was found to be completely absorbed in a repeated CT angiography one year late.

CONCLUSIONSFor the SISMAD, the involvement of the branches of the SMA may be associated with the extension of the dissection of the SMA. The inflation of the false lumina of the branches is worse than that of the SMA. With the improvement of the dissection of the SMA, the intramural hematoma of the branches can be absorbed. The conservative treatment may be the first choice for SISMAD without emergency.

Angiography ; Humans ; Mesenteric Artery, Superior ; diagnostic imaging ; Middle Aged ; Prognosis ; Retrospective Studies ; Tomography, X-Ray Computed

OBJECTIVETo investigate the effects of hepatitis C virus (HCV) on transcription regulation genes of host cells by gene chip assays in cultured cells with intact HCV genome.

METHODSHuh-7 hepatoma cells were cultured and infected with in vitro constructed HCV. The total RNAs, proteins and cell culture supernatants of HCV infected cells and control cells were isolated. Proteins and cell culture supernatants were used to detect the HCV replication and protein expression in cell culture system. The HCV protein expression was detected with Western blotting. Released HCV from infected cells was analyzed by real-time fluorescence quantitative PCR. Total RNA was qualified using 10 g/L agarose gel electrophoresis. cRNA was synthesized, fluorescence labeled and purified, then hybridized with Agilent oligo microarray (20173 probes). Differential expression of genes related to transcription in cell culture system was analyzed.

RESULTHCV was positive in cell culture supernatants and HCV protein expression was also positive according to Western blotting results. Eleven up-regulated and 11 down-regulated genes related to transcription were found after Agilent gene chip screening.

CONCLUSIONIntact hepatitis C virus cell culture system provides an useful tool for study on the affects of HCV infection on transcription regulation genes in host cells.

Cell Line, Tumor ; Gene Expression Profiling ; Gene Expression Regulation, Neoplastic ; Genome, Viral ; Hepacivirus ; genetics ; growth & development ; Hepatocytes ; virology ; Humans ; Liver Neoplasms ; genetics ; pathology ; virology ; Transcription, Genetic

Objective To determine the effect of type Ⅰ transmembrane protein (IRE1α) deletions on the cell cycle of human periodontal ligament fibroblasts (hPDLFs) cells. Methods Based on the IRE1α deletions, a full-length model was successfully constructed. Moreover, overlapping polymerase chain reaction mutagenesis facilitated the establishment of two deletion mutants of IRE1α (pD-Kinase, pD-Rnase). The full-length model and two mutant eukaryotic expression vectors were transfected into hPDLFs cells. Western blot analysis was performed to identify the expression in the cells. The changes in the cell cycle of hPDLFS cells were detected by flow cytometry (FCM). Results The two deletion mutants of IRE1α with eukaryotic expression vectors were successfully constructed and correctly expressed in hPDLFs cells based on Western blot analysis. Under stress conditions, the FCM assay showed that cell percentage of S phases increased, whereas that of G1 phases decreased in the IRE1α group (P<0.05) compared with the control group of tunicamycin (TM) treatment. Moreover, the cell percentage of the S phases decreased, whereas that of the G1 phases increased in the D-Rnase group (P<0.05) compared with the control. The deletion mutant D-Kinase had no influence on hPDLFS cell proliferation and cycle (P>0.05). Conclusion Under stress conditions, IRE1α can improve the cell cycle of hPDLFs cells from the G1 to the S phase. The deletion mutant D-Rnase cause hPDLFs cell growth arrest at the G1 phase, whereas deletion mutant D-Kinase has no significant effect.

OBJECTIVETo explore the potential markers of colorectal cancer metastasis and the influence of 5-FU on differentially expressed proteins by using proteomic technology, and to elucidate the mechanism of colorectal cancer metastasis.

METHODSHuman colorectal carcinoma cell lines of different metastatic potential, Lovo and SW480 were conventionally cultured, and the protein was extracted. 50% inhibitory concentration (IC(50)) of 5-FU to these two cell lines was measured by MTT assay. Proteins of these two cell lines after intervention by 5-FU at IC(50) were extracted, then 2-dimensional gel electrophoresis was conducted for the proteins. The differential protein spots were examined by mass spectrometry and analyzed by bioinformatics. Difference of expressed proteins in two cell lines before and after the intervention of 5-FU was validated by Western blot and immunofluorescence.

RESULTSEleven differentially expressed proteins were identified by 2-dimensional gel electrophoresis and mass spectrometry. The hnRNP K protein and PDI were selected to be examined by Western blot and immunofluorescence. Results revealed that the expression of hnRNP K in Lovo was higher than that in SW480, while the expression of PDI was lower in Lovo. After intervention by 5-FU at IC(50), the expression of hnRNP K in Lovo decreased more as compared to SW480, while the expression of PDI in SW480 increased more as compared to Lovo.

CONCLUSIONThere are significant differences in expression of hnRNP K and PDI proteins between Lovo and SW480 cell lines, and the proteins alter regularly after 5-FU intervention.

Biomarkers, Tumor ; blood ; Cell Line, Tumor ; Colorectal Neoplasms ; metabolism ; pathology ; Fluorouracil ; pharmacology ; Humans ; Neoplasm Metastasis ; Proteomics

Objective To investigate the characteristic of subtypes and genetic diversity of HIV-1 circulating in Hubei province and its molecular epidemiological linkages with regard to risk factors of viral transmission.Methods plasma samples of 80 diagnosed individuals was characterized.The gene fragments of gag were amplified by reverse transcriptase polymerase chain reaction (RT-PCR) and HIV-1 genotypes were determined based on the nucleotide sequences of gag region.Results Seven HIV-1 group M subtypes or CRF including B,B',G,CRF01-AE,CRF07-BC,CRF08-BC and CRF15-01B were identified.CRF01-AE was found to be the most dominant subtype (48.4％) followed by CRF7-BC (22.6％) and B' (12.9％).Conclusion The data from this study indicate the existence of multiple HIV-1 subtypes or CRFs in Hubei province and the surveillance of HIV-1 gene variation should be paid more attention to.

Objectives To screen the high-risk population of stroke in China using color Doppler flow imaging (CDFI)and to establish a stroke risk prediction model in Chinese population in order to prevent and treat stroke early. Methods Forty-one base hospitals and 715 286 people in the project areas of the first 6 provinces of China conducted routine physical examinations and investigated the related risk factors for cardiocerebrovascular diseases from July 2011 to April 2012 using a cross-sectional study,among them 61 860 patients underwent carotid CDFI screening,and 49 386 of them were high-risk population (exposed to≥3 risk factors). The bilateral common carotid interma-media thickness (IMT),the number of plaques and the degree of carotid stenosis were screened and documented. And whether carotid IMT thickening or not,with or without carotid plaques,and degree of carotid artery stenotic rate 0-49% and≥50% were performed by multivariate logistic regression analysis with the risk factors for stroke,respectively. Results (1)Logistic regression analysis showed that hypertension,atrial fibrillation,smoking,and lack of physical exercise were the independent risk factors for carotid IMT thickening (hypertension:OR,1. 17;95%CI 1. 12-1. 22;atrial fibrillation:OR,1. 15;95%CI 1. 09-1. 21;smoking:OR,1. 13;95%CI 1. 08-1. 17;and lack of physical exercise:OR,1. 12;95%CI 1.08-1. 16). (2)Hypertension,atrial fibrillation, smoking,and diabetes were the independent risk factors for carotid plaque and carotid artery stenosis rate≥50%(carotid plaque,hypertension:OR,1. 55;95%CI 1. 47-1. 62;atrial fibrillation:OR,1. 13;95%CI 1.06-1. 21;smoking:OR,1. 16;95%CI 1. 11-1. 22;and diabetes:OR,1. 30;95%CI 1. 24-1. 37). Carotid stenosis rate≥50%,hypertension:OR,1. 78;95%CI 1.55-2. 03;atrial fibrillation:OR,1. 59;95%CI 1. 39-1. 81;smoking:OR,1. 33;95%CI 1. 20-1. 48;and diabetes:OR,1. 30;95%CI 1. 17-1. 45. Simple obesity did not increase the incidences of carotid atherosclerotic plaque and carotid artery stenosis ≥50%(OR,0. 78, 0.83;95%CI 0. 75-0. 82 ,0. 75-0. 92,respectively). Conclusions Neck vascular ultrasound can be used as a valuable means for screening high-risk population and detecting risk factors of stroke. It has an important clinical significance for the early diagnosis and treatment of carotid atherosclerosis disease.