Seasonal variation of neutral lipids in Pinus sylvestris L. sapwood and heartwood

Summary The lipid levels and fatty acid composition of three fractions (free fatty acids, triacylglycerols and sterol/triterpenoid esters) extracted from the sapwood and heartwood of three stems of Pinus sylvestris were determined to investigate both seasonal changes in sapwood and possible metabolic changes related to heartwood formation. Seasonal changes were observed only in the amount of the free fatty acids in the sapwood: the level of free fatty acids was greatest at the beginning and end of the growing season. In the January and March samples the amount of the free fatty acid fraction in the sapwood was very small. The amount of the other fractions remained at the same level throughout the study. Marked seasonal changes in the fatty acid composition occurred only in the free fatty acid fraction of the sapwood: the saturation grade increased during the winter.     

Quantitation of autoradiographic grains in different zones of articular cartilage with image analyzer

Summary A novel method is introduced for the estimation of grain numbers in autoradiographic sections of articular cartilage with an image analyzer. It is based on separation of grains from the underlying structures by gray level thresholding and determination of the percentage of total area occupied by grains in a relatively large measuring field. The mean grain size is used as a reference to calculate grain numbers per cell profile and per unit area of tissue in various zones of bovine articular cartilage labelled with ³⁵S-sulphate in tissue culture. The results demonstrate considerable zonal differences as well as site related topographic variation in the rate of ³⁵S-sulphate incorporation. The largest site-related variation in the grain counts was observed in the superficial zone, suggesting a delicate control of proteoglycan synthesis in this zone.The IBAS program used in this work is available from Dr. J.J. Parkkinen or through Bitnet or EARN mail: MLAMMI at FINKUO     

Primer protein of bacteriophage M2 exposes the RGD receptor site upon linking the first deoxynucleotide

Summary Using electroporation with the phage PRD1 genome, we set up a high-frequency DNA transfer system for a linear dsDNA molecule with 5-covalently linked terminal proteins. The transfer was saturated when more than 100 ng of PRD1 genome was used. Electroporation efficiency was about four orders of magnitude higher than that obtained with transfection. Removal of the terminal protein abolished plaque formation, which could not be rescued by supplying the terminal protein or phage DNA polymerase or both in trans.     

Demonstration of chondroitin sulphate and glycoproteins in articular cartilage matrix using periodic acid-Schiff (PAS) method

Summary Staining of articular cartilage by the periodic acid-Schiff (PAS) method was measured using microspectrophotometry. Standard PAS technique with 2 h oxidation produced a distinct Schiff reaction in the cartilage sections. The staining increased with depth of the articular cartilage demonstrating distribution of the glycoproteins. The modified PAS method included a second, longer periodic acid treatment, which made the uronic acid of glycosaminoglycans PAS-positive. The modified PAS method proved to be highly specific for chondroitin sulphate, which was determined from the samples with gas chromatography. A statistically significant correlation between the Schiff reactivity and galactosamine content of the sections was observed. It is concluded that for articular cartilage standard and modified PAS methods are useful procedures for demonstrating local changes of glycoproteins and chondroitin sulphate, respectively.     

Indentation stiffness of young canine knee articular cartilage—Influence of strenuous joint loading

The indentation stiffness of knee articular cartilage subjected to strenuous physical training (SPT: treadmill running 20 km day⁻¹ for 15 weeks, n = 6) of young Beagles was tested and compared to that obtained from age-matched (55 weeks, n = 9) controls. The mathematical solution for the shear modulus, as determined from indentation of an elastic layer bonded to a rigid half space, was extended to small Poisson's ratios and applied to the analysis of cartilage response after a step stress (0.39 MPa) application. In these measurements with an impervious, plane-ended indenter, the equilibrium deformation was systematically greater than values predicted from the instant response by the linear biphasic theory. Therefore, the accurate determination of Poisson's ratio from the creep curves was not possible. The mean shear modulus (calculated by using the deformation at 900 s after load application and assuming a constant Poisson's ratio of 0.40 for the matrix) of canine knee articular cartilage was 0.37 MPa. While the cartilage thickness was not affected by SPT, the cartilage of the lateral tibial plateau was stiffer (13.3%, p<0.05) than that in controls. However, in the femoral condyles, the stiffness was at the control level or even below. Our results on cartilage structure and properties suggest that SPT, in contrast to our previous findings with moderate training, does not necessarily improve the biological properties of articular cartilage in young animals.     

Application of selected cationic dyes for the semiquantitative estimation of glycosaminoglycans in histological sections of articular cartilage by microspectrophotometry

Summary Selected commonly used cationic dyes, viz. Thionin, Safranin O, Toluidine Blue O, Dimethylmethylene Blue, Cuprolinic Blue, Cupromeronic Blue,N, N-Diethylpseudoisocyanine, and a modified PAS-method, and staining method, with a variety of alternative procedures, e.g., variation of pH, use of the critical electrolyte concentration method, and blocking reactions (methylation-saponification, carboxymethylation), were tested to select optimal staining procedures for the semiquantitative histochemical estimation of glycosaminoglycans by microspectrophotometry in sections of articular cartilage. The methods were carried out on 3 m-thick paraffin and 1 m-thick glycolmethacrylate sections of bovine articular cartilage. The staining intensity of the sections was measured from spots 25 m apart using a leitz MPV 3 microspectrophotometer, starting at the surface of the cartilage and ending up at the tidemark. The result was compared with the fixed-charge density graph determined from the adjacent articular cartilage.Of the dyes tested, Thionin and Safranin O proved to be excellent cationic dyes for the histochemical quantification of cartilage matrix proteoglycans, since the staining intensity curves showed a linear correlation (r=0.900–0.995) with the fixed charge density curves from the adjacent cartilage. Also, the stain distribution was consistently uniform across the sections. In 1 m-thick glycolmethacrylate sections, the Safranin O staining gradient showed almost perfect identity with the fixed-charge density curve. Cuprolinic Blue and Cupromeronic Blue combined with the critical electrolyte concentration technique were also useful for the microspectrophotometric assays of glycosaminoglycans, but the presence of metachromasia should be checked prior to the measurements. The reliability of blocking procedures for quantitative histochemical work was not convincing.     

Growth patterns in young adult monozygotic twin pairs discordant and concordant for obesity.

Weight discordance is very rare in monozygotic (MZ) twin pairs; when found, however, such pairs are advantageous in the search for either environmental or epigenetic causes and consequences of obesity. We analyzed the growth patterns of young adult MZ pairs discordant and concordant for obesity. Screening 5 consecutive birth cohorts (1975-1979) of 22- to 27-year-old Finnish twins (the FinnTwin16 study), we found 14 obesity discordant (Body Mass Index [BMI] difference > or = 4 kg/m2) MZ pairs out of 658. Ten pairs participated in clinical studies. Nine concordant pairs (BMI difference < or = 2 kg/m2) were examined as controls. Lifetime measured heights and weights recorded in hospitals and health centers were traced manually. Height development was similar in all the co-twins of both groups. The weight differences between the co-twins of the discordant pairs began to emerge at 18 years leading to an average discordance of 16.4 kg, 5.6 kg/m2 (p for both = .005) at 25.7 years. The heavier co-twin weighed 221 g (p = .066), 1.0 kg/m2 (p = .01) more already at birth than the leaner, but the differences waned by 6 months of age and reappeared only after adolescence. Both the leaner and the heavier co-twins of the discordant pairs weighed more than expected by the singleton reference values (Cole et al., 1998) after 8 years. The concordant co-twins, on the other hand, grew similarly and after 6 months, their mean growth was not distinguishable from the singleton patterns. Young adulthood represents a critical period of gaining weight irrespective of genetic background in this twin sample.     

Variability of nutrient limitation in the Archipelago Sea,SW Finland

Over a two year study period, zooplankton was sampledin Gazi Bay, Kenya, using a 335 μm mesh size Bongonet. Two Way Indicator Species Analysis (TWINSPAN)classification technique demonstrated that rainfalland tidal regime had substantial influence on thezooplankton community structure. Samples collectedduring the rainy season months clustered together whentreated with TWINSPAN. Furthermore, theclustering was more pronounced for neap tidesamples than for spring tide ones. Samples obtainedduring spring tide did not give a clear cut pattern. Canonical Correspondence Analysis (C.C.A.) confirmedthese findings, a clustering together of rainy/neaptide samples; and little separation (based onenvironmental variables) between samplingstations. This revised version was published online in August 2006 with corrections to the Cover Date.     

Complexing of glycofuranosides with calcium ion,and its effect on their acid-catalysed solvolysis

Stability constants for the calcium-ion complexes of several methyl aldo-furanosides have been determined in aqueous solution by using a potentiostatic technique with an electrode that is selective for calcium ions. The results obtained have been verified by examining the chromatographic behaviour of the compounds on a strong cation-exchange resin in the Ca²⁺ form. The rate constants for the acid-catalyzed hydrolysis and methanolysis of a few 4-chlorophenyl aldofuranosides having different complexing abilities have been determined at various concentrations of calcium chloride. The dependences of the observed salt effects on the complexing ability of the substrate are discussed.     

The rate of calcium extraction during EDTA decalcification from thin bone slices as assessed with atomic absorption spectrophotometry

Summary The rate of calcium extraction with EDTA (ethylenediamine tetraacetic acid) from thin bone slices (300 m-2mm thick) was determined by aid of an atomic absorption spectrophotometer. A 0.5 mm thick bone slice was completely decalcified with 15% (0.40 M), 8% (0.22 M), and 4% (0.11 M) EDTA in 24 h, 3 days, and 5 days, respectively (vol. 15 ml, temp. 4° C, pH 7.4). At 37 and 60° C the speed of demineralization was slightly increased as compared with that at 20° C, while no difference was observed between 4 and 20° C. Bone slices with a thickness of 0.3, 0.5, 1 and 2 mm were decalcified-in the same order-in 24 h, 2, 3, and 5 days (8% EDTA, 4° C, pH 7.4). At pH 7.4, the decalcification rate was a little slower than at pH 5.0 and 8.5. Agitation did not affect the decalcifying velocity, nor did the volume of the agent, except when the volume was very small. The demineralization of ordinary bone, containing both compact and spongy bone, was found to be more rapid than that of homogeneous bone reported earlier. The acidic buffers and New Decalc^®, which served as reference substances, exerted a more vigorous decalcifying effect than EDTA. K formate/formic acid buffer, pH 3.15, demineralized a 1 mm thick bone slice in 24 h, and 2 days was needed with Na lactate/lactic acid buffer, pH 3.70. With New Decalc^®, pH 0.9, the corresponding demineralization was accomplished in 1.5 h. Atomic absorption spectrophotometer proved to be a useful tool in the evaluation of calcium extraction velocity from bone slices.