Migration and turnover of entero-endocrine and caveolated cells in the epithelium of the descending colon, as shown by radioautography after continuous infusion of 3H-thymidine into mice

Adult male mice were given a continuous infusion of about 0.5 muCi of 3H-thymidine per gram body weight per day for periods varying from 1 to 60 days. Semithin sections of descending colon were cut from/plastic-embedded blocks and stained by a method combining silver impregnation and iron hematoxylin, by which argentaffin entero-endocrine cells and caveolated cells could be identified. From radioautographs, the labeling index of these cells was determined. One to three days after the beginning of 3H-thymidine infusion, label is observed in some of the stained entero-endocrine cells in the bottom of the crypts; the apices of these cells reach the crypt lumen and are joined to neighboring cells by terminal bars (junctional complexes). After five to seven days, labeled entero-endocrine cells are seen on the sides of the crypts, where their base stretches along the basement membrane and their apex has lost its terminal bar connections to neighboring cells. Finally, by 13 and 24 days, labeled cells are observed within the epithelium at the mucosal surface. The turnover time, which is taken to be equal to the mean time required for migration from site of origin to site of loss on the mucosal surface, has been estimated at 23.3 days. This is much longer than the 4.6 days required by the two main cell types of the epithelium -- vacuolated-columnar and mucous cells -- to travel the same route. It is likely that, after entero-endocrine cells lose their terminal bar attachment to other epithelial cells, they migrate independently and very slowly. Labeled caveolated cells are first seen in the crypt bottom one day after the beginning of 3H-thymidine infusion. By three to five days, they are on the sides of the crypts; their base is stretched along the basement membrane, but their apex retains its attachment to neighboring cells by terminal bars. By seven days, labeled caveolated cells are on the mucosal surface. Their turnover time has been assessed at 8.2 days. This is, again, longer than for the two main types to which they are bound by terminal bars throughout migration. The discrepancy is explained by the caveolated cells arising deeper in the crypts than most vacuolated-columnar and mucous cells.     

Pulsed-field gel electrophoresis analysis of the genome of Streptomyces ambofaciens strains

The genome of four Streptomyces ambofaciens strains from different geographical origins (ATCC15154, DSM40697, ETH9247 and ETH 11317) was analysed by pulsed-field gel electrophoresis (PFGE). The PFGE technique has allowed the study of the extrachromosomal content of these strains and the characterization of their genomic DNA by restriction analyses. Electrophoretic migration of undigested DNA allowed us to detect a 80 kb-length linear molecule with concatemeric forms in S. ambofaciens ATCC15154. These extrachromosomal molecules were shown to be homologous to the circular plasmid pSAM1 (80 kb) suggesting that pSAM1 could exist not only in circular form but also in linear form. In the same way a 45 kb-length linear molecule was detected in S. ambofaciens ETH9427 and ETH11317. In contrast, no extrachromosomal DNA could be detected in S. ambofaciens DSM40697. The analysis of the macrorestriction patterns using the rate-cutting enzymes AseI and DraI indicated a close relationship between the DSM- and ETH- strains. Indeed, three types of restriction patterns were distinguished: while S. ambofaciens ETH9427 and ETH11317 were characterized by the same pattern and share more than 75% of comigrating fragments with the strain DSM40697, S. ambofaciens ATCC15154 exhibited a restriction pattern different from the other three. The total genome sizes of S. ambofaciens ATCC15154, DSM40697, ETH9427 and ETH11317 were estimated to be about 6500, 8000, 8200 and 8200 kb, respectively.     

Soluble and particulate inositol 1,4,5-trisphosphate 5-phosphatases show common antigenic determinants

Inositol 1,4,5-trisphosphate 5-phosphatase catalyses the dephosphorylation of the phosphate in the 5-position from inositol 1,4,5-trisphosphate and inositol 1,3,4,5-tetrakisphosphate. One particulate and two soluble enzymes were previously described in bovine brain. In this study, we have obtained a precipitating antiserum against soluble type I inositol 1,4,5-trisphosphate 5-phosphatase. The particulate, but not the soluble type II enzyme, was immunoprecipitated by the serum. Inositol 1,4,5-triphosphate 5-phosphatase activity from crude extracts of rat brain, human platelets and rat liver were immmunoprecipitated by the same antibodies, suggesting the existence of common antigenic determinant among inositol 1,4,5-trisphosphate 5-phosphatases of diverse sources.     

UK-73,093: A non-peptide neurotensin receptor antagonist

UK-73,093 was identified in a screening program as a compound able to displace [³H]-neurotensin from its bovine brain receptor. We describe the discovery of this compound, species differences in receptor affinity and its characterization as a functional neurotensin antogonist in vitro and in vivo.     

Visualization of chromosome assembly during the S and G2 stages of the cycle and chromosome disassembly during the G1 stage in semithin sections of mouse duodenal crypt cells and other cells

Previous examination of dividing cells in the isthmus of the mouse pyloric antrum by using semithin (0.5-micron-thick) Epon sections revealed that the prophasic condensation of chromosomes began early in the DNA-synthesizing (S) stage. In order to examine whether the same observation could be made in other proliferating cell types, the crypt base columnar cells in mouse duodenum and the hepatocytes of the rat 48 hr after partial hepatectomy were investigated by morphologic and radioautographic techniques. When crypt base columnar cells were studied in semithin Epon sections, the four phases of mitosis showed the characteristic features described by classical cytologists. Moreover, the proportion of cells in prophase and telophase was high. To relate the mitotic phases to the stages of the cell cycle, the "frequency of labeled mitoses method" provided the duration of the cell cycle, 12.3 hr, and of the S stage, 7.3 hr. From the frequency of the occurrence of mitotic phases, it was estimated that metaphase lasted 0.3 hr and anaphase 0.11 hr, in line with previous estimates. However, the durations of prophase and telophase were long, 5.9 and 1.9 hr, respectively. The whole mitotic process took over 8 hr. From the duration of prophase and cycle stages, it was calculated that 67% of the S stage was occupied by prophasic cells. In fair agreement with this estimate, 68% of the labeled cells 10 min after a 3H-thymidine injection were found to be in prophase. In regenerating hepatocytes, the morphological features and frequency of prophase and telophase cells were similar to those observed in duodenal crypt cells. While the cycle time was not measured and, therefore, the duration of cycle stages and mitotic phases could not be estimated, it is likely that their duration would be of the same order of magnitude. In conclusion, the mitotic process in duodenal crypt cells takes over 8 hr. Moreover, the crypt cells, like antral isthmal cells, show features of early prophase soon after they enter the S stage of the cycle.     

Cellular stages in cartilage formation as revealed by morphometry, radioautography and type II collagen immunostaining of the mandibular condyle from weanling rats

The role played by cell addition, cell enlargement, and matrix deposition in the endochondral growth of the condyle was assessed in weanling rats by four approaches making use of the light microscope: morphometry, 3H-thymidine radioautography, 3H-proline radioautography, and immunostaining for the cartilage-specific type II collagen. From the articular surface down, the condyle may be divided into five layers made up of cells embedded in a matrix: 1) the articular layer composed of static cells in a matrix rich in fibers presumed to be of type I collagen, 2) the polymorphic cell layer including the progenitor cells from which arise the cells undergoing endochondral changes, 3) the flattened cell layer in which cells produce a precartilagenous matrix devoid of type II collagen while undergoing differentiation in two stages: a "chondroblast" stage and a short "flattened chondrocyte" stage when intracellular type II collagen elaboration begins, 4) the upper hypertrophic cell layer, in which cells are "typical chondrocytes" that enlarge at a rapid rate, actively produce type II collagen, and deposit it into a cartilagenous matrix, and 5) the lower hypertrophic cell layer, composed of chondrocytes at a stage of terminal enlargement while the cartilagenous matrix is adapting for mineralization. 3H-thymidine radioautographic results indicate that the turnover time of progenitor cells in the polymorphic cell layer is about 2.9 days. The time spent by cells at each stage of development is estimated to be 1.4 days as chondroblasts, 0.5 days as flattened chondrocytes, 2.3 days as the chondrocytes of the upper hypertrophic cell layer, and 1.1 days as those of the lower hypertrophic cell layer. Calculations referring to a 1 x 1-mm square-sided column extending from the articular surface to the zone of vascular invasion provide the daily rate of cell addition (0.0077 mm3), extracellular matrix deposition (0.0127 mm3), and cell enlargement (0.0302 mm3). Hence the respective contribution of the three factors to condyle growth is in a ratio of about 1:1.6:4. This result emphasizes the role played by cell enlargement in the overall growth of the condyle.     

Cellobiose metabolism in Erwinia: genetic study

Summary The study of mutants of Erwinia specifically unable to ferment cellobiose indicates that the mutations are clustered between arg and ile on the chromosome of this organism. In vivo cloning of the genes responsible for cellobiose utilization lead to a plasmid, pBEC2, which complements all Erwinia Clb^- specific mutants. When introduced into wild-type E. coli it allows this organism to use cellobiose, arbutin and salicin; it also complements bglB and bglC mutants of Escherichia coli indicating that arbutin and salicin utilization is due to the products of the pBEC2 cloned genes. From the characterization of mutants pleiotropically affected in the utilization of various carbon sources, including cellobiose, arbutin and salicin, it is proposed that the three--glucosides are substrates of the phosphoenolpyruvate-dependent phosphotransferase system (PTS).