OBJECTIVETo study the biomechanical change of the craniovertebral junction in conditions of atlas assimilation.

METHODSMimics software was used to process CT data of the craniovertebral junction in a health adult to obtain the three-dimensional reconstruction and the cloudy points of C1, C2 and part of the occipital bone. Then the cloudy points were imported into the Abaqus 6. 8 software to establish the occipito-atlantoaxial finite element model in normal structure. According to the established model in normal structure, the model in conditions of atlas assimilation was set by changing the model parameters. Both models of normal structure and atlas assimilation were loaded with 1. 5 N . m static moment to simulate four motions of flexion, extension, lateral bending and axial rotation respectively. The movement characteristics,joint stress force and ligament deformation was analyzed.

RESULTSUnder 1. 5 N . m moment, in model of atlas assimilation the C1-C2 range of movement decreased from 13. 55° to 11.88° in flexion,increased from 13. 22° to 15. 24° in extension and from 4. 05° to 4. 23° in lateral bending and remained unchanged in axial rotation when compared with the normal model. In flexion movement, the contact force of the atlanto-dental joint increased from 1. 59 MPa to 3. 28 MPa and the deflection of apical ligament, tectorial membrane and alar ligament increased 129. 1%, 157. 6% and 75. 1% respectively when compared with the normal model.

CONCLUSIONSThe normal C1-C2 motion mode is destructed in conditions of atlas assimilation, leading to the changes of the range of movement,joint stress force and the ligament deformation at C1 C2 junction. The atlantoaxial instability will likely occur in flexion motion.

Atlanto-Axial Joint ; physiology ; Biomechanical Phenomena ; Cervical Atlas ; physiology ; Cervical Vertebrae ; Finite Element Analysis ; Humans ; Imaging, Three-Dimensional ; Joint Instability ; Ligaments, Articular ; Occipital Bone ; Range of Motion, Articular ; Rotation

Objective:To explore caspase-6's effect on the apoptosis of osteosarcoma cell line SOSP 607. Methods: The expression level of caspase-6 in the osteosarcoma cell line SOSP-9067 was examined by RT-PCR method. The adeno-virus adv5 vector with caspase-6 gene was constructed and transferred into the osteosarcoma cell line SOSP-9607 by lipo-fection. The cell survival rate after transfection was assayed by MTT method. The cell morphological changes were observed by microscope and electron microscope, the apoptosis of transferred cells were examined by gel electrophoresis. Results:No expression of caspase-6 was examined in the osteosarcoma cell line SOSP-9607. A high expression of caspase-6 was identified by RT-PCR after the transfection. The cell growth curve declined after transferring caspase-6. Electrophoresis of DNA displayed the apoptosis ladder. Conclusion: Transferring caspase-6 into the osteosarcoma line SOSP-9607 may inhibit the growth of the osteosarcoma cell line SOSP-9607 and this effect might be achieved by inducing apoptosis.

MicroRNAs (miRNAs) , a species of small noncoding RNA, could regulate gene expression bv binding to the 3'-untranslated region of target mR NA at post-transcription level.MicroRNAs play important roles in various human biological processes such as differentiation, cell proliferation, and apoptoMs.Abnormal expression of miRNAs is implicated in carcinogenesis and progression of various cancers, indicating that miRNAs could be served as molecular biomarkers for diagnosis and treatment of cancer.In this resiew, the author summarize the most common altered miRNAs expression profiles and their possible roles in promoting cell proliferation, tumor metastasis,and chemotherapeutic resistance in gastric cancer.

OBJECTIVETo identify genes associated with metastasis suppression and to investigate the molecular mechanism of osteosarcoma metastasis.

METHODSThe subtracted cDNA library of low metastatic human osteosarcoma cell line SOSP-9607 was constructed using suppression subtractive hybridization. Partial clones were sequenced. The acquired sequence data were aligned against the GenBank nucleotide database using Blastn to search for sequence matches. The interested clone was used to perform Northern blot and reverse transcriptase-PCR (RT-PCR) analysis on mRNA isolated from low metastatic cell line SOSP-9607 and OS-9901, high metastatic cell line SOSP-M and three pulmonic metastatic nodules of nude mice.

RESULTSA cDNA clone from low metastatic cell line SOSP-9607 subtracted cDNA library was identified as telomeric repeat binding factor 2 (TERF2) by sequence analysis and Blastn search. Northern blot and RT-PCR analysis demonstrated that TERF2 expressed highly in low metastatic cell line SOSP-9607 and OS-9901, but not in high metastatic cell line SOSP-M and three pulmonic metastatic nodules.

CONCLUSIONTERF2 may be important for suppressing metastasis of osteosarcoma.

Animals ; Base Sequence ; Blotting, Northern ; DNA-Binding Proteins ; genetics ; Gene Expression Regulation, Neoplastic ; Humans ; Mice ; Mice, Nude ; Molecular Sequence Data ; Neoplasm Metastasis ; genetics ; Neoplasm Transplantation ; Nucleic Acid Hybridization ; methods ; Osteosarcoma ; genetics ; pathology ; RNA, Messenger ; genetics ; metabolism ; Telomeric Repeat Binding Protein 2 ; Transplantation, Heterologous ; Tumor Cells, Cultured

In the face of the worldwide threat of severe acute respiratory syndrome (SARS) to human life, some of the most urgent challenges are to develop fast and accurate analytical methods for early diagnosis of this disease as well as to create a safe anti-viral vaccine for prevention. To these ends, we investigated the antigenicity of the spike protein (S protein), a major structural protein in the SARS-coronavirus (SARS-CoV). Based upon the theoretical analysis for hydrophobicity of the S protein, 18 peptides were synthesized. Using Enzyme-Linked Immunosorbent Assay (ELISA), these peptides were screened in the sera from SARS patients. According to these results, two fragments of the S gene were amplified by PCR and cloned into pET-32a. Both S fragments were expressed in the BL-21 strain and further purified with an affinity chromatography. These recombinant S fragments were confirmed to have positive cross-reactions with SARS sera, either by Western blot or by ELISA. Our results demonstrated that the potential epitope regions were located at Codons 469-882 in the S protein, and one epitope site was located at Codons 599-620. Identification of antigenic regions in the SARS-CoV S protein may be important for the functional studies of this virus or the development of clinical diagnosis.
Antigens, Viral ; immunology ; Chromatography, High Pressure Liquid ; Cloning, Molecular ; Electrophoresis, Polyacrylamide Gel ; Enzyme-Linked Immunosorbent Assay ; Genetic Vectors ; Humans ; Mass Spectrometry ; Membrane Glycoproteins ; genetics ; immunology ; metabolism ; Molecular Weight ; Peptide Fragments ; chemistry ; Recombinant Proteins ; genetics ; immunology ; SARS Virus ; genetics ; immunology ; metabolism ; Spike Glycoprotein, Coronavirus ; Viral Envelope Proteins ; genetics ; immunology ; metabolism

In order to develop clinical diagnostic tools for rapid detection of the SARS-CoV (severe acute respiratory syndrome-associated coronavirus) and to identify candidate proteins for vaccine development, the C-terminal portion of the nucleocapsid (NC) gene was amplified using RT-PCR from the SARS-CoV genome, cloned into a yeast expression vector (pEGH), and expressed as a glutathione S-transferase (GST) and Hisx6 double-tagged fusion protein under the control of an inducible promoter. Western analysis on the purified protein confirmed the expression and purification of the NC fusion proteins from yeast. To determine its antigenicity, the fusion protein was challenged with serum samples from SARS patients and normal controls. The NC fusion protein demonstrated high antigenicity with high specificity, and therefore, it should have great potential in designing clinical diagnostic tools and provide useful information for vaccine development.
Antigens, Viral ; immunology ; Cloning, Molecular ; Enzyme-Linked Immunosorbent Assay ; Genetic Vectors ; Genome, Viral ; Humans ; Nucleocapsid Proteins ; genetics ; immunology ; Recombinant Fusion Proteins ; genetics ; isolation & purification ; metabolism ; SARS Virus ; genetics ; immunology ; Yeasts ; genetics

The nucleocapsid protein (N protein) has been found to be an antigenic protein in a number of coronaviruses. Whether the N protein in severe acute respiratory syndrome-associated coronavirus (SARS-CoV) is antigenic remains to be elucidated. Using Western blot and Enzyme-linked Immunosorbent Assay (ELISA), the recombinant N proteins and the synthesized peptides derived from the N protein were screened in sera from SARS patients. All patient sera in this study displayed strong positive immunoreactivities against the recombinant N proteins, whereas normal sera gave negative immunoresponses to these proteins, indicating that the N protein of SARS-CoV is an antigenic protein. Furthermore, the epitope sites in the N protein were determined by competition experiments, in which the recombinant proteins or the synthesized peptides competed against the SARS-CoV proteins to bind to the antibodies raised in SARS sera. One epitope site located at the C-terminus was confirmed as the most antigenic region in this protein. A detailed screening of peptide with ELISA demonstrated that the amino sequence from Codons 371 to 407 was the epitope site at the C-terminus of the N protein. Understanding of the epitope sites could be very significant for developing an effective diagnostic approach to SARS.
Blotting, Western ; Enzyme-Linked Immunosorbent Assay ; Epitopes ; chemistry ; immunology ; Humans ; Nucleocapsid Proteins ; chemistry ; immunology ; Peptide Fragments ; chemical synthesis ; Plasmids ; Recombinant Proteins ; immunology ; isolation & purification ; metabolism ; SARS Virus ; genetics ; immunology ; metabolism

Heme is a key cofactor in aerobic life, both in eukaryotes and prokaryotes. Because of the high reactivity of ferrous protoporphyrin IX, the reactions of heme in cells are often carried out through heme-protein complexes. Traditionally studies of heme-binding proteins have been approached on a case by case basis, thus there is a limited global view of the distribution of heme-binding proteins in different cells or tissues. The procedure described here is aimed at profiling heme-binding proteins in mouse tissues sequentially by 1) purification of heme-binding proteins by heme-agarose, an affinity chromatographic resin; 2) isolation of heme-binding proteins by SDS-PAGE or two-dimensional electrophoresis; 3) identification of heme-binding proteins by mass spectrometry. In five mouse tissues, over 600 protein spots were visualized on 2-DE gel stained by Commassie blue and 154 proteins were identified by MALDI-TOF, in which most proteins belong to heme related. This methodology makes it possible to globally characterize the heme-binding proteins in a biological system.
Animals ; Carrier Proteins ; biosynthesis ; genetics ; Electrophoresis, Gel, Two-Dimensional ; Electrophoresis, Polyacrylamide Gel ; Heme ; chemistry ; Hemeproteins ; biosynthesis ; genetics ; Mass Spectrometry ; Mice ; Mice, Inbred ICR ; Protein Binding ; Proteins ; chemistry ; Proteome ; Proteomics ; methods ; Sepharose ; chemistry ; Tissue Distribution

ObjectiveTo investigate the clinical diagnostic value of extra-spindle pole-like protein 1 （ESPL1） in the progression of hepatitis B virus （HBV）-related liver fibrosis. MethodsA total of 228 patients with HBV infection who were admitted to The First Affiliated Hospital of Guangxi Medical University from June 2017 to August 2023 were enrolled. The transient elastography system FibroScan was used to determine liver stiffness measurement （LSM） for all patients， and according to the LSM value， they were divided into non-liver fibrosis group with 80 patients， mild liver fibrosis group with 83 patients， advanced liver fibrosis group with 30 patients， and liver cirrhosis group with 35 patients. ELISA was used to measure the serum level of ESPL1. The Kruskal-Wallis H test was used for comparison of the serum level of ESPL1 between the four groups； the Spearman correlation analysis was used to investigate the correlation between ESPL1 and LSM； the receiver operating characteristic （ROC） curve was used to analyze the value of serum ESPL1 in predicting the progression of liver fibrosis. ResultsThe liver cirrhosis group had a significantly higher serum level of ESPL1 than the non-liver fibrosis group and the mild liver fibrosis group （both P<0.05）， and the advanced liver fibrosis group and the mild liver fibrosis group had a significantly higher serum level of ESPL1 than the non-liver fibrosis group （both P<0.05）. The correlation analysis showed that there was a positive correlation between serum ESPL1 and LSM in the patients with HBV infection and varying degrees of liver fibrosis （r=0.515， P<0.001）. Serum ESPL1 had an area under the ROC curve （AUC） of 0.809 in predicting liver cirrhosis and an AUC of 0.638 in predicting advanced liver fibrosis， with a sensitivity of 87.5% and 100%， respectively， and a specificity of 59.7% and 31.3%， respectively. ConclusionThere is a certain correlation between serum ESPL1 and HBV-related liver fibrosis， and higher serum ESPL1 may indicate a higher degree of liver fibrosis. Serum ESPL1 is expected to become one of the serum markers for assisting in the diagnosis of liver cirrhosis and an important clinical method for dynamically monitoring the progression of liver fibrosis in patients with HBV infection.