Proteomic analysis of human osteoarthritis synovial fluid

Background

Osteoarthritis is a chronic musculoskeletal disorder characterized mainly by progressive degradation of the hyaline cartilage. Patients with osteoarthritis often postpone seeking medical help, which results in the diagnosis being made at an advanced stage of cartilage destruction. Sustained efforts are needed to identify specific markers that might help in early diagnosis, monitoring disease progression and in improving therapeutic outcomes. We employed a multipronged proteomic approach, which included multiple fractionation strategies followed by high resolution mass spectrometry analysis to explore the proteome of synovial fluid obtained from osteoarthritis patients. In addition to the total proteome, we also enriched glycoproteins from synovial fluid using lectin affinity chromatography.

Results

We identified 677 proteins from synovial fluid of patients with osteoarthritis of which 545 proteins have not been previously reported. These novel proteins included ADAM-like decysin 1 (ADAMDEC1), alanyl (membrane) aminopeptidase (ANPEP), CD84, fibulin 1 (FBLN1), matrix remodelling associated 5 (MXRA5), secreted phosphoprotein 2 (SPP2) and spondin 2 (SPON2). We identified 300 proteins using lectin affinity chromatography, including the glycoproteins afamin (AFM), attractin (ATRN), fibrillin 1 (FBN1), transferrin (TF), tissue inhibitor of metalloproteinase 1 (TIMP1) and vasorin (VSN). Gene ontology analysis confirmed that a majority of the identified proteins were extracellular and are mostly involved in cell communication and signaling. We also confirmed the expression of ANPEP, dickkopf WNT signaling pathway inhibitor 3 (DKK3) and osteoglycin (OGN) by multiple reaction monitoring (MRM) analysis of osteoarthritis synovial fluid samples.

Conclusions

We present an in-depth analysis of the synovial fluid proteome from patients with osteoarthritis. We believe that the catalog of proteins generated in this study will further enhance our knowledge regarding the pathophysiology of osteoarthritis and should assist in identifying better biomarkers for early diagnosis.     

160-fold acceleration of the Smith-Waterman algorithm using a field programmable gate array (FPGA)

Background  

To infer homology and subsequently gene function, the Smith-Waterman (SW) algorithm is used to find the optimal local alignment between two sequences. When searching sequence databases that may contain hundreds of millions of sequences, this algorithm becomes computationally expensive.     

A third variant of glucose phosphate isomerase in pigs

A new variant of glucose phosphate isomerase (GPI), also known as phosphohexose isomerase (PHI), was detected in a primitive pig population.     

Microbial carbon metabolism associated with electrogenic sulphur oxidation in coastal sediments

Recently, a novel electrogenic type of sulphur oxidation was documented in marine sediments, whereby filamentous cable bacteria (Desulfobulbaceae) are mediating electron transport over cm-scale distances. These cable bacteria are capable of developing an extensive network within days, implying a highly efficient carbon acquisition strategy. Presently, the carbon metabolism of cable bacteria is unknown, and hence we adopted a multidisciplinary approach to study the carbon substrate utilization of both cable bacteria and associated microbial community in sediment incubations. Fluorescence in situ hybridization showed rapid downward growth of cable bacteria, concomitant with high rates of electrogenic sulphur oxidation, as quantified by microelectrode profiling. We studied heterotrophy and autotrophy by following ¹³C-propionate and -bicarbonate incorporation into bacterial fatty acids. This biomarker analysis showed that propionate uptake was limited to fatty acid signatures typical for the genus Desulfobulbus. The nanoscale secondary ion mass spectrometry analysis confirmed heterotrophic rather than autotrophic growth of cable bacteria. Still, high bicarbonate uptake was observed in concert with the development of cable bacteria. Clone libraries of 16S complementary DNA showed numerous sequences associated to chemoautotrophic sulphur-oxidizing Epsilon- and Gammaproteobacteria, whereas ¹³C-bicarbonate biomarker labelling suggested that these sulphur-oxidizing bacteria were active far below the oxygen penetration. A targeted manipulation experiment demonstrated that chemoautotrophic carbon fixation was tightly linked to the heterotrophic activity of the cable bacteria down to cm depth. Overall, the results suggest that electrogenic sulphur oxidation is performed by a microbial consortium, consisting of chemoorganotrophic cable bacteria and chemolithoautotrophic Epsilon- and Gammaproteobacteria. The metabolic linkage between these two groups is presently unknown and needs further study.     

Activin-A up-regulates type I activin receptor mRNA levels in human immortalized extravillous trophoblast cells.

Activin is known to play an important regulatory role in reproduction, including pregnancy. To further examine the role and signaling mechanism of activin in regulating placental function, the steady-state level of activin type I receptor (ActRI) mRNA in immortalized extravillous trophoblasts (IEVT) cells was measured using competitive PCR (cPCR). An internal standard of ActRI cDNA for cPCR was constructed for the quantification of ActRI mRNA levels in IEVT cells. ActRI mRNA levels were increased in a dose-dependent manner by activin-A with the maximal effect observed at the dose of 10 ng/ml. Time course studies revealed that activin-A had maximal effects on ActRI mRNA levels at 6 hours after treatment. The effects of activin-A on ActRI mRNA levels was blocked by follistatin, an activin binding protein, in a dose-dependent manner. In addition, inhibin-A inhibited basal, as well as activin-A-induced ActRI mRNA levels. These findings provide evidence, for the first time, that activin-A modulates ActRI mRNA levels in human trophoblast cells.     

The variability of the hepatitis B virus genome: statistical analysis and biological implications

A statistical analysis of the nucleotide sequence variability in 14 published hepatitis B virus (HBV) genomes was carried out using parametric and nonparametric methods. A parametric statistical model revealed that the different regions of the genome differed significantly in their variability. The conclusion was supported by a nonparametric kernel-density model of the HBV genome. Genes S, C, and P, region X, the precore region, and the pre-S2/pre-S1 regions were ranked in order of increasing variability. In many instances, conserved regions of the genome identified with sequences of known function in HBV biology. However, other characterized regions (such as pre-S) showed much variability despite the involvement of their encoded peptides in specific functions. Point mutations that may result in the formation of stop codons and amino acid changes may affect the clinical picture of HBV infection and may be reflected in atypical serological patterns.