Linc-ROR promotes mesenchymal stem cells chondrogenesis and cartilage formation via regulating SOX9 expression.

OBJECTIVE: The present study is to characterize the role of long intergenic non-coding RNA, regulator of reprogramming (linc-ROR) in bone marrow mesenchymal stem cell (BMSCs) chondrogenesis, cartilage formation and OA development. METHODS: Linc-ROR expression pattern in articular cartilage tissue sample from OA patients were studied by real-time PCR. Linc-ROR lentivirus mediated BMSCs were constructed. In vitro micromass cultured BMSCs chondrogenesis or in vivo MeHA hydrogel encapsulated BMSCs cartilage formation activity were studied. Linc-ROR associating miRNAs which repressed SOX9 expression were characterized by luciferase assay, real-time PCR and Western blot. Linc-ROR was co-transfected with miRNAs into BMSCs to study its rescue effect on SOX9 expression and chondrogenesis activity. RESULTS: Linc-ROR was down-regulated in articular cartilage tissue from OA patients and was positively correlated with the expression level of SOX9 (R2 = 0.43). Linc-ROR expression was upregulated during BMSCs chondrogenesis. Linc-ROR ectopic expression significantly promoted in vitro BMSCs chondrogenesis and in vivo cartilage formation activities as revealed by safranin O, alcian blue and COL II staining. The mRNA expression level of chondrogenesis markers including COL II, SOX9 and ACAN were increased, and the hypertrophy markers MMP13 and COL X were decreased upon linc-ROR overexpression in BMSCs. Linc-ROR functioned as a miRNA sponge for miR-138 and miR-145. Both miR-138 and miR-145 suppressed BMSCs chondrogenesis activity and SOX9 expression, while co-expression of linc-ROR displayed a rescuing effect. CONCLUSIONS: Taken together, linc-ROR modulated BMSCs chondrogenesis differentiation and cartilage formation by acting as a competing endogenous RNA for miR-138 and miR-145 and activating SOX9 expression. Linc-ROR could be considered as a new diagnostic and therapeutic target for OA treatment.

Characterization of the N-terminal repeat domain of Escherichia coli ClpA-A class I Clp/HSP100 ATPase.

The ClpA, ClpB, and ClpC subfamilies of the Clp/HSP100 ATPases contain a conserved N-terminal region of approximately 150 residues that consists of two approximate sequence repeats. This sequence from the Escherichia coli ClpA enzyme is shown to encode an independent structural domain (the R domain) that is monomeric and approximately 40% alpha-helical. A ClpA fragment lacking the R domain showed ATP-dependent oligomerization, protein-stimulated ATPase activity, and the ability to complex with the ClpP peptidase and mediate degradation of peptide and protein substrates, including casein and ssrA-tagged proteins. Compared with the activities of the wild-type ClpA, however, those of the ClpA fragment missing the R domain were reduced. These results indicate that the R domain is not required for the basic recognition, unfolding, and translocation functions that allow ClpA-ClpP to degrade some protein substrates, but they suggest that it may play a role in modulating these activities.

Determination of linear alkylbenzenesulfonates and their degradation products in water samples by gas chromatography with ion-trap mass spectrometry.

A method was developed for the analysis of linear alkylbenzenesulfonates (LAS) and their degradation products, sulfophenylcarboxylic acids (SPC), in samples of sewage effluent and river water. This method involved extraction of the samples by graphitized carbon black cartridge, esterification by a two-step thionyl chloride-trifluoroethanol derivatization procedure, and separation, identification and quantitation by ion-trap GC-MS with EI and low pressure CI modes. High selectivity with few signals was observed in the low pressure CI mass spectra of LAS and SPC. Enhanced sensitivity with protonated molecular ion chromatograms of homologous C10-C13 LAS by CI-MS permit the determination of LAS and SPC at trace concentrations in environmental samples. Recovery rates of LAS and SPC in spiked water samples ranged from 75 to 112% with R.S.D. values from 3 to 26%. The limit of quantitation for both LAS and SPC was estimated to be 0.01 microgram/l in 100 ml of water sample.

Left ventricular opacification after peripheral venous injection of a modified albumin solution.

The usefulness of a modified albumin solution was assessed in 8 dogs after peripheral venous and inferior vena cava injections. The contrast agent is a mixed solution made of glucose, albumin and glycerin, with sonicated microbubble diameter of 5.0 +/- 2.3 microns. Multiple injections (8 ml each) of this contrast agent (total 80 injections) into peripheral vein and inferior cava were performed. The blood pressure from femoral artery was measured before, during and after injections. Two-dimensional echocardiograms were recorded in a modified long axis view on videotapes for play back analysis. The pulmonary transit time and left ventricular contrast persistent time was determined for each injection. The videodensity of the region of interest (ROI) at the center of right ventricle and left ventricle was measured. The background videodensity of both ventricles was evaluated. The videodensity over the ROI of both ventricles with peak contrast enhancement was measured in all frames for 3 consecutive cardiac cycles. The peak videodensity of right and left ventricle subtracting the background videodensity of each ventricles was further calculated respectively. The injections caused no change in blood pressure or heart rate. All injections produced right ventricular contrast echo. As much as 85% of peripheral venous and 82.5% of inferior vena cava injections resulted in left ventricular contrast which was 0.68 and 0.65 as bright as that produced in the right ventricle. Pulmonary transit time and left ventricle contrast persistent time of peripheral venous injection was 4.05 +/- 0.53 and 13.67 +/- 4.28 seconds respectively. No difference of these data (3.93 +/- 0.47 and 11.65 +/- 4.66 seconds) from those produced by inferior vena cava injections were noted.(ABSTRACT TRUNCATED AT 250 WORDS)