Bone marrow-derived fibroblast migration via periostin causes irreversible arthrogenic contracture after joint immobilization.

Joint contracture causes distressing permanent mobility disorder due to trauma, arthritis, and aging, with no effective treatment available. A principal and irreversible cause of joint contracture has been regarded as the development of joint capsule fibrosis. However, the molecular mechanisms underlying contracture remain unclear. We established a mouse model of knee joint contracture, revealing that fibrosis in joint capsules causes irreversible contracture. RNA-sequencing of contracture capsules demonstrated a marked enrichment of the genes involved in the extracellular region, particularly periostin (Postn). Three-dimensional magnetic resonance imaging and immunohistological analysis of contracture patients revealed posterior joint capsule thickening with abundant type I collagen (Col1a2) and POSTN in humans. Col1a2-GFPTG ; Postn-/- mice and chimeric mice with Col1a2-GFPTG ; tdTomatoTG bone marrow showed fibrosis in joint capsules caused by bone marrow-derived fibroblasts, and POSTN promoted the migration of bone marrow-derived fibroblasts, contributing to fibrosis and contracture. Conversely, POSTN-neutralizing antibody attenuated contracture exacerbation. Our findings identified POSTN as a key inducer of fibroblast migration that exacerbates capsule fibrosis, providing a potential therapeutic strategy for joint contracture.

Wheat germ cell-free system-based production of malaria proteins for discovery of novel vaccine candidates.

One of the major bottlenecks in malaria research has been the difficulty in recombinant protein expression. Here, we report the application of the wheat germ cell-free system for the successful production of malaria proteins. For proof of principle, the Pfs25, PfCSP, and PfAMA1 proteins were chosen. These genes contain very high A/T sequences and are also difficult to express as recombinant proteins. In our wheat germ cell-free system, native and codon-optimized versions of the Pfs25 genes produced equal amounts of proteins. PfCSP and PfAMA1 genes without any codon optimization were also expressed. The products were soluble, with yields between 50 and 200 mug/ml of the translation mixture, indicating that the cell-free system can be used to produce malaria proteins without any prior optimization of their biased codon usage. Biochemical and immunocytochemical analyses of antibodies raised in mice against each protein revealed that every antibody retained its high specificity to the parasite protein in question. The development of parasites in mosquitoes fed patient blood carrying Plasmodium falciparum gametocytes and supplemented with our mouse anti-Pfs25 sera was strongly inhibited, indicating that both Pfs25-3D7/WG and Pfs25-TBV/WG retained their immunogenicity. Lastly, we carried out a parallel expression assay of proteins of blood-stage P. falciparum. The PCR products of 124 P. falciparum genes chosen from the available database were used directly in a small-scale format of transcription and translation reactions. Autoradiogram testing revealed the production of 93 proteins. The application of this new cell-free system-based protocol for the discovery of malaria vaccine candidates will be discussed.

Peritoneal penetration and pharmacodynamic exposure of intravenous cefozopran in abdominal surgery patients.

This study aimed to examine the peritoneal penetration and pharmacodynamic exposure of intravenous cefozopran. Cefozopran 1 g was administered to 10 patients before abdominal surgery. Venous blood and peritoneal fluid (PF) samples were obtained at the end of the 0.5-h infusion and at 1, 2, 3, 4, 5 and 6 h thereafter. Drug concentrations in plasma and PF were determined, analysed pharmacokinetically and used for a Monte Carlo simulation with minimum inhibitory concentration (MIC) data. Cefozopran penetrated well into PF, with a mean (+/-standard deviation) maximum drug concentration in PF/plasma ratio of 0.65+/-0.17 (n=10) and area under the concentration-time curve ratio of 0.92+/-0.13. The probabilities of attaining the pharmacodynamic exposure target (PTA), defined as 70% of the time above the MIC, in PF were almost identical to those in plasma. The PTAs were 95-100% against Escherichia coli, Enterobacter spp. and Staphylococcus aureus with 0.5 g every 12 h; however, 1 g every 8 h or 1 g every 6 h was required for 93-98% PTA against Pseudomonas aeruginosa. These results should help us to understand the peritoneal pharmacokinetics of cefozopran whilst also helping to choose the appropriate dosage for surgical intra-abdominal infections.

Enzymatic characterization of the Plasmodium vivax chitinase, a potential malaria transmission-blocking target.

The chitinase (EC 3.2.1.14) of the human malaria parasite Plasmodium falciparum, PfCHT1, has been validated as a malaria transmission-blocking vaccine (TBV). The present study aimed to delineate functional characteristics of the P. vivax chitinase PvCHT1, whose primary structure differs from that of PfCHT1 by having proenzyme and chitin-binding domains. The recombinant protein rPvCHT1 expressed with a wheat germ cell-free system hydrolyzed 4-methylumbelliferone (4MU) derivatives of chitin oligosaccharides (beta-1,4-poly-N-acetyl glucosamine (GlcNAc)). An anti-rPvCHT1 polyclonal antiserum reacted with in vitro-obtained P. vivax ookinetes in anterior cytoplasm, showing uneven patchy distribution. Enzymatic activity of rPvCHT1 shared the exclusive endochitinase property with parallelly expressed rPfCHT1 as demonstrated by a marked substrate preference for 4MU-GlcNAc(3) compared to shorter GlcNAc substrates. While rPvCHT1 was found to be sensitive to the general family-18 chitinase inhibitor, allosamidin, its pH (maximal in neutral environment) and temperature (max. at approximately 25 degrees C) activity profiles and sensitivity to allosamidin (IC50=6 microM) were different from rPfCHT1. The results in this first report of functional rPvCHT1 synthesis indicate that the P. vivax chitinase is enzymatically close to long form Plasmodium chitinases represented by P. gallinaceum PgCHT1.

Spontaneous osteonecrosis of the lateral femoral condyle of the knee: a report of 11 cases.

BACKGROUND: Spontaneous osteonecrosis (SON) of the lateral femoral condyle of the knee joint is very rare and there have been only a few articles about this condition. MATERIALS: We reviewed data for 11 patients (7 men, 4 women) with unusual SON of the lateral femoral condyle of the knee. The average age of patients at onset was 61.9 years (range 47-76 years). No patient had underlying disease or had undergone steroid therapy, although one underwent lateral meniscectomy. RESULTS: According to Aglietti's radiographic classification, three patients had stage 1 disease, two had stage 2 disease, three had stage 3 disease, one had stage 4 disease, and two had stage 5 at first examination. The average alignment of affected limbs on standing was 5.9 degrees valgus (range 0 degrees -11 degrees ). Although the osteonecrotic lesion was seen on the lateral side, the mechanical axis passed the medial compartment in three patients. Six patients were treated conservatively and the remaining five required surgery, which consisted of lateral unicompartmental knee arthroplasties. CONCLUSION: The pathology of the necrosis of the lateral femoral condyle was considered to be different from that of the medial femoral condyle regarding clinical features, limb alignment, and radiographic findings.

Recurrent hemarthrosis after knee joint arthroplasty: etiology and treatment.

This study reports the results for 10 patients with recurrent hemarthrosis after knee joint arthroplasty. The average interval between arthroplasty and the first instance of hemarthrosis was at 26 months, and the average number of hemarthroses per patient was 3.8. In 3 patients, the bleeding responded to simple conservative measures. The remaining 7 needed surgery; there were 6 arthroscopic synovectomies and 1 polyethylene revision. Impingement of the proliferative synovium was observed in only 2 patients during surgical intervention. In the 2 patients in whom arthroscopic management was successful, another procedure with an electric coagulator, in addition to a formal synovectomy, was performed. The use of a coagulator may be helpful for direct coagulation when arthroscopic management is selected, although open synovectomy is curative in most cases.