Collagenous substrata regulate the nature and distribution of glycosaminoglycans produced by differentiated cultures of mouse mammary epithelial cells

We have investigated the influence of culture substrata upon glycosaminoglycans produced in primary cultures of mouse mammary epithelial cells isolated from the glands of late pregnant mice. Three substrata have been used for experiments: tissue culture plastic, collagen (type I) gels attached to culture dishes, and collagen (type I) gels that have been floated in the culture medium after cell attachment. These latter gels contract significantly. Cells cultured on all three substrata produce hyaluronic acid, heparan sulfate, chondroitin sulfates and dermatan sulfate but the relative quantities accumulated and their distribution among cellular and extracellular compartments differ according to the nature of the culture substratum. Notably most of the glycosaminoglycans accumulated by cells on plastic are secreted into the culture medium, while cells on floating gels incorporate almost all their glycosaminoglycans into an extracellular matrix fraction. Cells on attached collagen gels secrete approx. 30% of their glycosaminoglycans and assemble most of the remainder into an extracellular matrix. Hyaluronic acid is produced in significant quantities by cells on plastic and attached gels but in relatively reduced quantity by cells on floating gels. In contrast, iduronyl-rich dermatan sulfate is accumulated by cells on floating gels, where it is primarily associated with the extracellular matrix fraction, but is proportionally reduced in cells on plastic and attached gels. The results are discussed in terms of polarized assembly of a morphologically distinct basal lamina, a process that occurs primarily when cells are on floating gels. In addition, as these cultures secrete certain milk proteins only when cultured on floating gels, we discuss the possibility that cell synthesized glycosaminoglycans and proteoglycans may play a role in the maintenance of a differentiated phenotype.     

Functional organization of cerebral microvasculature in a marsupial, the northern quoll (Dasyurus hallucatus)

Capillaries within the central nervous system (CNS) of eutherian mammals form meshworks with numerous anastomoses, whereas capillaries in the CNS of marsupials consist entirely of hairpin-like loops, without anastomotic interconnections. Counter-current blood flow in capillary loops may have been important in the evolutionary development of a cerebral vascular supply. However, loops are not found in eutherian mammals, perhaps because of a limited benefit to the diffusive conductance of gases.     

Interactions between SV40 T antigen and DNA polymerase alpha

Simian virus 40 large T antigen is the only viral protein required for SV40 DNA synthesis in vivo and in vitro. This complex protein recruits the cellular DNA replication apparatus to the SV40 origin and provides a good model for the initiation of cellular DNA replication. The interaction between SV40 large T antigen (TAg) and DNA polymerase alpha has been shown previously to be inhibited by murine p53, the nuclear protein product of a cellular anti-oncogene. The murine p53 protein will inhibit SV40 replication both in vivo and in vitro. Using monoclonal antibodies to TAg, p53, and polymerase alpha, we developed immunoassays to measure the complexes formed between TAg and polymerase alpha and between TAg and p53. The assays allowed us to detect the TAg-polymerase alpha and TAg-p53 complexes in lytically infected and transformed cells. The amount of TAg complexed to p53 was far lower in infected cells than in transformed cells. We used a large range of monoclonal antibodies to different sites on T antigen and found that antibodies that inhibited the formation of the TAg-polymerase alpha complex also inhibited the formation of the TAg-p53 complex. Finally, we found that the tsA58 and 5080 point mutations in TAg, previously shown to inhibit the binding of TAg to p53, also inhibit its binding to polymerase alpha. Together these results emphasize the specificity and functional importance of the TAg-polymerase alpha complex. The disruption of this interaction by the cellular anti-oncogene p53 provides an interesting model for the normal action of p53 and the effects of its removal on the regulation of cellular DNA synthesis.     

Tail-to-tail orientation of the Atlantic salmon alpha- and beta-globin genes

We report the cloning of a cDNA and two corresponding -globin genes of the Atlantic salmon (Salmo salar L.) as well as two genes for -globins. Nucleotide sequence analysis of the cDNA shows that the predicted -globin peptide comprises 148 amino acids with a calculated molecular mass of 16,127 Da and an overall amino acid similarity of 40–50% to higher vertebrates and 60–90% to fish sequences. The study of the genomic organization of - and -globin genes shows that, as is the case in Xenopus, the salmon genes are adjacent. Two sets of linked - and -globin genes were isolated and restriction-enzyme polymorphisms indicate that they belong to two distinct loci, possibly as a result of the salmon tetraploidy. In each locus the - and -globin genes are oriented 3 to 3 relative to each other with the RNA coding sequences located on opposite DNA strands. This is the first evidence for this type of arrangement found for globin genes. Moreover, while the linkage found in salmon and Xenopus supports the hypothesis of an initial tandem duplication of a globin ancestor gene, our results raise the question of the actual original orientation of the duplicated genes. Correspondence to: F. Gannon     

Getting to the root of the problem: new evidence for the use of plant root foods in Mesolithic hunter-gatherer subsistence in Europe

Vegetation History and Archaeobotany - This paper presents new evidence for the harvesting of edible plant roots and tubers at Northton, a Mesolithic hunter-gatherer site on Harris, in the Western...     

Transforming growth factor-beta 1 acts cooperatively with sodium n-butyrate to induce differentiation of normal human keratinocytes.

Growth factors with established biological activity toward cultured normal human epidermal keratinocytes (NHEKs) (e.g., transforming growth factor-beta, TGF-beta; retinoic acid, RA) initiate programmed changes in cellular maturation which differ with regard to the specific differentiation pathway (normal or abnormal) analyzed. Sodium butyrate (NaB) initiates one form of epidermal differentiation leading to enhanced cornified envelope (CE) formation which involves abrogation of the normally inhibitory effect of RA on NHEK maturation. NaB also induces TGF-beta mRNA in the maturing suprabasal compartment, suggesting that TGF-beta may play a role in NaB-initiated NHEK differentiation. Treatment with TGF-beta 1 alone, however, only marginally increased (by twofold) the number of detergent-resistant CEs compared to control NHEKs and did not alter the prevalence of fully mature enucleated CEs. TGF-beta 1 was quite effective in inducing significant levels of CE expression when used simultaneously with suboptimal concentrations of NaB. The cooperative action of suboptimal NaB and TGF-beta 1 generated numbers of CEs which, in fact, exceeded the incidence of mature CEs formed in response to optimal levels of NaB alone. Neutralizing antibodies to TGF-beta, moreover, effectively reduced the incidence of CE formation in cultures treated with optimal NaB concentrations, further implicating endogenous TGF-beta activity in the NaB-initiated NHEK differentiation model. It is suggested, therefore, that within the NaB-induced pathway of NHEK differentiation, TGF-beta can positively modulate expression of the differentiated phenotype but alone is insufficient for generation of mature CEs.     

Effect of Planting Unit Size and Sediment Stabilization on Seagrass Transplants in Western Australia

Abstract The effect of increasing planting unit size and stabilizing sediment was examined for two seagrass planting methods at Carnac Island, Western Australia in 1993. The staple method (sprigs) was used to transplant Amphibolis griffithii (J. M. Black) den Hartog and the plug method was used to transplant A. griffithii and Posidonia sinuosa Cambridge and Kuo. Transplant size was varied by increasing the number of rhizomes incorporated into a staple and increasing the diameter of plugs. Planting units were transplanted into bare sand, back into the original donor seagrass bed, or into a meadow of Heterozostera tasmanica, which is an important colonizing species. Sprigs of A. griffithii were extracted from a monospecific meadow; tied into bundles of 1, 2, 5, and 10 rhizomes; and planted into unvegetated areas. Half the units were surrounded by plastic mesh and the remainder were unmeshed. All treatments were lost within 99 days after transplanting, and although larger bundles survived better than smaller ones, no significant differences could be attributed to the effects of mesh or sprig size. Plugs of P. sinuosa and A. griffithii were extracted from monospecific meadows using polyvinyl chloride pipe of three diameters, 5, 10, and 15 cm, and planted into unvegetated areas nearby. Half the units were surrounded by plastic mesh and the remainder were unmeshed. Posidonia sinuosa plugs were also placed within a meadow of H. tasmanica (Martens ex Aschers.) den Hartog. Only 60% of A. griffithii plug sizes survived 350 days after transplanting back into the donor bed; however, survival of transplants at unvegetated areas varied considerably, and analysis of variance indicated a significant two‐way interaction between treatment and plug size. Transplants survived better when meshed (90% survived) and survival improved with increasing plug size. Posidonia sinuosa transplants survived poorly (no plugs survived beyond 220 days in bare or meshed treatments) regardless of size. Survival of 10‐ and 15‐cm plugs was markedly better than the 5‐cm plugs in vegetated areas, including the H. tasmanica meadow. The use of large seagrass plugs may be appropriate for transplantation in high‐energy wave environments.