Gene expression of matrix metalloproteinases 1, 3, and 9 by chondrocytes in osteoarthritic human knee articular cartilage is zone and grade specific

OBJECTIVES: Matrix metalloproteinases (MMPs) are thought to be major mediators of cartilage destruction. Osteoarthritis (OA) is characterised by cartilage degradation. This study explores gene expression of three MMPs in articular chondrocytes during the histological development of the cartilage lesion of OA. METHODS: Biopsy specimens of human normal and OA cartilage, classified into four grades on the basis of histology, were probed for MMPs 1, 3, and 9 using 35S-labelled cDNA probes. The signal was measured at four different depths (zones) using an automated image analyser and compared with signal from sections probed with lambda DNA. Rheumatoid synovium was used as a positive control for MMP gene expression. RESULTS: Rheumatoid tissue contained mRNA for all three MMPs. Expression in chondrocytes varied with the depth of the chondrocyte in the cartilage and the histomorphological extent of the OA changes. There was no detectable mRNA signal for these three MMPs in normal cartilage. In general, in OA, MMP-1 gene expression was greatest in the superficial cartilage in established disease. By contrast mRNAs for MMP-3 and 9 were expressed deeper in the cartilage, MMP-9 early in disease and MMP-3 with a biphasic pattern in early and late stage disease, most pronounced in the latter. This was a consequence of differential expression in single cells and chondrocyte clusters in late disease. CONCLUSION: The data indicate that expression of genes for MMPs 1, 3, and 9 is differentially regulated in human articular chondrocytes and, in individual cells, is related to the depth of the chondrocyte below the cartilage surface and the nature and extent of the cartilage lesion.     

Timolol gel versus acetazolamide in the prophylaxis of ocular hypertension after phacoemulsification

PURPOSE: To compare postoperative intraocular pressure (IOP) after administration of acetazolamide and timolol following phacoemulsification and intraocular lens implantation. SETTING: Ophthalmic Consultants of Long Island, Rockville Centre, New York, USA. METHODS: Sixty patients were included in a prospective, randomized, masked trial. The patients received either two doses of oral, sustained-release acetazolamide (Diamox Sequels) or a single dose of topical timolol 0.5% gel (Timoptic XE). Intraocular pressure was measured by Goldmann applanation tonometry preoperatively and 1 day postoperatively. RESULTS: Mean preoperative IOP was 16.4 mm Hg. One day postoperatively, it was 19.5 mm Hg in the oral acetazolamide group and 15.9 mm Hg in the timolol gel group. One patient in the acetazolamide group developed significant adverse reactions. CONCLUSION: Prophylactic use of topical timolol 0.5% gel for viscoelastic-induced ocular hypertension after cataract extraction appears to offer better IOP control than oral acetazolamide and has potentially fewer adverse systemic effects.     

Prediction of outcome and early vs. late improvement in OCD patients treated with cognitive behaviour therapy and pharmacotherapy

Several factors must be taken into account when deciding which specimen preparation technique(s) to use. These factors include the amount of material available, ease of preparing this material due to its properties and familiarity, location and size of the region of interest, amount of information sought, facilities accessible, and time available by the researcher to devote to the preparation of the specimen. The more popular specimen preparations for thin films, namely, electropolishing, cleaving, crushing, mechanical thinning followed by ion milling, and ultramicrotomy are discussed and the more unusual techniques such as extraction/replication, photochemical etching, lithography and reactive ion etching (RIE), chemically assisted ion beam etching (CAIBE), and precision polishing-based techniques are described. Their advantages and disadvantages in the context of the above factors are discussed. Suggestions for increasing one's success rate in preparing specimens are given. The role of transmission electron microscopy (TEM) analysis is considered since it rarely stands in isolation from other physical analytical techniques, nor is it often used as a quick diagnostic tool. Conservation of material by the minimization of the amount of material used (or destroyed) by TEM specimen preparation, and conservation of one's time by performing TEM analysis only on "worthy" samples should be given maximum consideration.     

Estimation of variance components for somatic cell counts to determine thresholds for uninfected quarters

The objective of this study was to determine the factors affecting somatic cell count (SCC), to estimate variance components of these factors, and to calculate and evaluate the thresholds for intramammary infection based on SCC. The infection status from 22,467 quarter milk samples from 544 cows in seven herds was determined. Infections status was the most important factor affecting SCC. The increase in SCC was more pronounced for major pathogens than for minor pathogens. Even after adjustment for infection status, the interaction between stage of lactation and parity was significant. For culture-negative samples within a lactation, the shape of the SCC curve was inversely related to the shape of the milk production curve. The shape of the SCC curve was flat for first lactation cows compared with the shape of the SCC curve for cows in subsequent lactations. The effect of clinical mastitis on SCC was significant. The use of SCC thresholds for specific parities and stages of lactation to detect intramammary infection improved quality parameters only slightly over a fixed threshold of 200,000 cells/ml.     

Requirement for Rho in integrin signalling

Overnight culture of Swiss 3T3 cells in serum-free medium leads to loss of focal adhesions and associated actin stress fibres, although the cells remain well spread. The small GTP-binding protein Rho is required for the formation of stress fibres and focal adhesions induced by growth factors such as lysophosphatidic acid (LPA) in serum-starved Swiss 3T3 cells, and for the LPA-induced tyrosine phosphorylation of several focal adhesion proteins. Plating of cells on extracellular matrix proteins also stimulates protein tyrosine phosphorylation and the formation of stress fibres and focal adhesions in the absence of added growth factors. These responses were inhibited in cells scrape-loaded with the Rho inhibitor C3 transferase. Focal adhesion and stress fibre formation was also triggered by addition of a peptide GRGDS, which is recognised by a number of integrins and is contained within the cell binding domain of a variety of extracellular matrix proteins. The activity of the GRGDS peptide was blocked by microinjecting cells with C3 transferase, suggesting that peptide binding to integrins stimulates a Rho-dependent assembly of focal adhesions. These experiments indicate that Rho is involved in signalling downstream of integrins.     

The impact of pre-clerking clinics on surgical operation cancellations: a prospective audit

Pre-clerking of all patients undergoing elective general surgical operations was introduced at our hospital in an attempt to reduce an unacceptably high operation cancellation rate. A prospective audit has been performed on the effect of this policy on the cancellation rate. Before the introduction of pre-clerking there was a marked seasonal variation in the number of patients who failed to attend for surgery, which could be explained by absence on holiday. This seasonal variation disappeared after the start of pre-clerking clinics, but there has been no reduction in the number of cancellations for medical reasons.     

Immunohistochemical localization of p21(WAF1/CIP1) in normal, hyperplastic, and neoplastic uterine tissues

Assuming that the chemical reactions used to synthesize a combinatorial library member are successful, then knowledge of the specific reaction sequence is equivalent to knowing the member's chemical identity. Because the determination of chemical identity is typically not automatable and requires a substantial amount of material, schemes that encode a member's reaction history onto the reaction platform are of value. The primary benefits of encoding are relational nomenclature (all methods) and automated handling (some methods). Encoding methods evaluated to date are spatial, graphical, chemical, spectrometric, electronic, and physical.     

Cloning and disruption of the antigenic catalase gene of Aspergillus fumigatus

Aspergillus fumigatus possesses two catalases (described as fast and slow on the basis of their electrophoretic mobility). The slow catalase has been recognized as a diagnostic antigen for aspergillosis in immunocompetent patients. The antigenic catalase has been purified. The enzyme is a tetrameric protein composed of 90-kDa subunits. The corresponding cat1 gene was cloned, and sequencing data show that the cat1 gene codes for a 728-amino-acid polypeptide. A recombinant protein expressed in Pichia pastoris is enzymatically active and has biochemical and antigenic properties that are similar to those of the wild-type catalase. Molecular experiments reveal that CAT1 contains a signal peptide and a propeptide of 15 and 12 amino acid residues, respectively. cat1-disrupted mutants that were unable to produce the slow catalase were as sensitive to H2O2 and polymorphonuclear cells as the wild-type strain. In addition, there was no difference in pathogenicity between the cat1 mutant and its parental cat1+ strain in a murine model of aspergillosis.