Microporosity of the substratum regulates differentiation of MDCK cells in vitro

Summary We have analyzed the ability of the physical substratum to modulate both the ultrastructural and protein synthetic characteristics of the Madin-Darby canine kidney (MDCK) renal cell line. When MDCK cells were seeded on Millipore Millicell CM microporous membrane cell culture inserts they demonstrated a more columnar organization with an increase in cell density sixfold greater than the same cells seeded on conventional plastic substrata. After 1 wk postseeding on the microporous membrane a partial basal lamina was noted, with a contiguous basement membrane being apparent after 2 wk. One-dimensional sodium dodecyl sulfate gel electrophoresis was used to analyze detergent-solubilized proteins from MDCK cells maintained on plastic substrata vs. microporous membranes. When proteins were pulse-labeled with [³⁵S]methionine, a 55 kDa protein was evident in the cytosolic extract of cells grown on collagen, laminin, and nontreated plastic substrata; but this labeled protein was not evident in similar extracts from cells grown on collagen and laminin-coated microporous membranes. To test if the polarized, basement-membrane secreting phenotype of the MDCK cells could be generated on a microporous membrane without pretreatment with any extracellular matrix (ECM) components, cells were seeded on the Millipore Millicell HA (cellulosic) microporous membrane. This type of substrata does not need a coating of ECM components for cell attachment. A partial basement membrane was formed below cells where the basal surface of the cell was planar, but not in areas where the cell formed large cytoplasmic extensions into the filter. This led us to the conclusion that the microporous nature of the substrata can dictate both ultrastructural and protein synthetic activities of MDCK cells. Furthermore, we suggest that both the planar nature of the basal surface and the microporosity of the substrate are corequisites for the deposition of the basement membrane.     

Twenty years of numerical syntaxonomy

The development of numerical syntaxonomy during its first 20 yr is reviewed. The use of methods of numerical classification and ordination is the dominating feature of the development. National and local phytosociological data banks were established, large data sets handled and many important vegetation monographs were methodically based on multivariate data analysis. Particularly the development in Italy, the Netherlands, Czechoslovakia, and Sweden contributed to new theoretical elements of numerical syntaxonomy. Ordination became a common tool of searching for reticulate synsystematic relations between community types. The most popular ordination techniques have been Principal Components Analysis and Detrended Correspondence Analysis. Hierarchical agglomerative techniques of clustering still prevail in classification, although the divisive strategy of TWINSPAN has also become an effective tool for phytosociological clustering and table sorting. Extensive program packages, also for personal computers have now become standard equipment for many vegetation scientists.     

A new dissimilarity measure and a new optimality criterion in phytosociological classification

A new dissimilarity measure, Uppsala dissimilarity, is proposed. It is a Manhattan-type measure in between the Canberra and Gower measures, based on the differences between scores in relevés compared, but it also takes both the sums of scores and the difference between maximum and minimum score into account. The measure is considered realistic for phytosociological material.A new optimality criterion has been developed after unsatisfactory results had been obtained with the DOL criterion (Popma et al. 1983) which was developed previously by our group. Problems with DOL were especially met when the criterion was applied to the distribution of only one species over the cluster array obtained. The new criterion takes both internal cluster homogeneity and between-cluster dissimilarity into account. Between-cluster dissimilarity is calculated for all other clusters and not only for the nearest neighbour, as in DOL. The new criterion has both an unweighted form: SOM, and a form with weighting for cluster size: SWOM.This new criterion was successfully applied to the evaluation of the sharpness of distribution of individual species over cluster arrays, under the name of SIM: species indication measure and SWIM, species weighted indication measure.The measures were applied to some test data. Differences between the unweighted and weighted forms were found which could not be easily interpreted.Some remarks are made on the coherence of d-SAHN and h-SAHN approaches in agglomerative clustering within the new strategy proposed.Abbreviations DOL = Detection of Optimal Level - S(W)IM = Species (Weighted) Indication Measure - S(W)OM = Standardized (Weighted) Optimality Measure - UD = Uppsala Dissimilarity measure - WPGMA = Weighted Pair-Group Method Average linking clustering - SAHN = Sequential Agglomerative Hierarchical Non-overlapping clustering     

Monitoring juvenile stocks of flatfish in the Wadden Sea and the coastal areas of the southeastern North Sea

Results are presented of an on-going monitoring program, started in 1970, of the demersal fish population in the Wadden Sea, Scheldt estuary, and along the continental coast between the Belgian-French border and Esbjerg. Particular attention is given to long-term trends in overall abundance and annual variations in spatial distribution of 0- and 1-group plaice and sole in relation to year-class strength and to variations in growth rate. Recruitment level in both plaice and sole appeared to be higher in the 1980s than in the 1970s. The most important nursery areas for plaice are the German Bight and the Wadden Sea, and the relative contributions of these areas to the total plaice stock in different years are relatively stable. In contrast, the most important nurseries for sole are along the continental coast, and the contribution of different parts of these nurseries vary considerably from year to year. Abundant year-classes of sole can originate from northern as well as from southern parts of the nurseries. Relations between survey results and year-class estimates from Virtual Population Analysis (VPA) for plaice are better compared to sole. No trends have been observed in mean length at age 0 and 1 during the years the surveys have been carried out. It is concluded that the quality of the continental coastal area and the Wadden Sea as nursery area for plaice and sole has not declined during the period of investigation. Presented at the VI International Wadden Sea Symposium (Biologische Anstalt Helgoland, Wattenmeerstation Sylt, D-2282 List, FRG, 1–4 November 1988)     

Effects of Solar Radiation on Photoorientation,Motility and Pigmentation in a Freshwater Cryptomonas

The effects of solar radiation on motility, photoorientation and pigmentation have been studied in a freshwater Cryptomonas species. The diaphototactic orientation performed by the cells is impaired within about 90 min of solar radiation. Likewise, the percentage of motile cells within the population and the average velocity of the swimming cells decreases within about the same exposure time. This effect is not due to a thermal stress but rather seems to be caused by the solar UV-B component, since decreasing short wavelength UV radiation by means of an artificial ozone filter or UV cut-off filters increased the tolerated exposure time. Solar radiation also bleached the photosynthetic pigments of the cells as shown by absorption difference spectra.     

Semisynthesis of Arg15, Glu15, Met15, and Nle15-aprotinin involving enzymatic peptide bond resynthesis

The semisynthesis of homologues of aprotinin, the bovine pancreatic trypsin inhibitor, is described. The P₁ lysine¹⁵ residue was replaced by two methods. The first procedure, which consisted of two enzymatic steps for the incorporation of other amino acids has previously been described. The second approach consisted of six steps of both enzymatic and chemical nature. The modified inhibitor, in which the lysine¹⁵-alanine¹⁶ peptide bond is hydrolyzed, was used as the starting material. All carboxyl groups of the modified inhibitor were esterified with methanol; the lysine¹⁵ methylester group was then selectively hydrolyzed. Afterward, lysine¹⁵ itself was split off. Arginine, glutamic acid, methionine, andl-2-aminohexanoic acid (norleucine, Nle) were incorporated using water-soluble carbodiimide combined with an acylation catalyst. The methylester group was used to prevent polymerization. The reactive-site peptide bonds were resynthesized using either chymotrypsin or trypsin.     

Effects of tracheal airway occlusion on hyoid muscle length and upper airway volume

Complex relationships exist among electromyograms (EMGs) of the upper airway muscles, respective changes in muscle length, and upper airway volume. To test the effects of preventing lung inflation on these relationships, recordings were made of EMGs and length changes of the geniohyoid (GH) and sternohyoid (SH) muscles as well as of tidal changes in upper airway volume in eight anesthetized cats. During resting breathing, tracheal airway occlusion tended to increase the inspiratory lengthening of GH and SH. In response to progressive hypercapnia, the GH eventually shortened during inspiration in all animals; the extent of muscle shortening was minimally augmented by airway occlusion despite substantial increases in EMGs. SH lengthened during inspiration in six of eight animals under hypercapnic conditions, and in these cats lengthening was greater during airway occlusion even though EMGs increased. Despite the above effects on SH and GH length, upper airway tidal volume was increased significantly by tracheal occlusion under hypercapnic conditions. These data suggest that the thoracic and upper airway muscle reflex effects of preventing lung inflation during inspiration act antagonistically on hyoid muscle length, but, because of the mechanical arrangement of the hyoid muscles relative to the airway and thorax, they act agonistically to augment tidal changes in upper airway volume. The augmentation of upper airway tidal volume may occur in part as a result of the effects of thoracic movements being passively transmitted through the hyoid muscles.     

Relationship between exercise-induced muscle damage and enzyme release in rats.

The relationship between the amount of exercise-induced muscle damage and the release of creatine kinase (CK), aspartate aminotransferase (AST), and lactate dehydrogenase (LD) was studied. Gender differences in enzyme release and histological damage were also studied. Serial pre- and postexercise blood samples were drawn from untrained male and female catheterized Wistar rats that ran 1.5 or 2.5 h on a treadmill (incline 10 degrees). Three days postexercise, muscle damage was quantified morphometrically in five different hindlimb and forearm muscles. The 1.5 and 2.5 h of exercise elicited histological damage only in the soleus muscle. Significant plasma CK, AST, and LD elevations were found immediately postexercise both in male and female rats. However, the enzyme release was significantly greater in males than in females. Part of this could be explained by differences in clearance rates between males and females. No gender difference in amount of histological damage was found. The actual volume of histological muscle damage was significantly less than the calculated muscle damage based on enzyme release. An increase in the exercise duration from 1.5 to 2.5 h resulted in a disproportional increase in both histological muscle damage and muscle enzyme release. From the present study it is concluded that muscle enzyme release is not clearly reflected in histological muscle damage.