Volatile emissions of plant tissue cultures

Summary The low-molecular-weight volatiles released by a variety of plant tissue cultures were examined by gas chromatography. Callus cultures invariably produced carbon dioxide, ethylene, acetaldehyde and ethanol. In cultures with developed shoots, ethanol was absent and acetaldehyde was detected only rarely.     

Tissue microenvironment and the genetic control of hair pigment patterns in mice

This study was conducted to assess microenvironmental variability within integumental tissue of genetically identical mice with respect to a specific cellular response: cyclic synthesis of yellow and black pigment by hair bulb melanocytes. Crosses were performed within and between inbred strains of mice that were isogenic with the exception of a single gene substitution at the agouti locus. Agouti locus genes included the A^vy, A^w, A, a^td, a^t, a^x, a^m, and a alleles. The pigment patterns of dorsal, flank, and ventral hairs of the first and third hair generations and of hairs growing in special integumentary areas such as the pinna, tail, and hind foot were studied. It was found that the amount of yellow pigment synthesized by hair bulb melanocytes within genetically identical mice is both agedependent and conditioned by the integumentary environment. Furthermore, the special integumentary regions produce hairs with a variety of pigment patterns in which the distribution and relative amounts of black and yellow pigments do not necessarily conform to dominance relationships expected among agouti locus alleles as judged by their effects on the pigmentation of the dorsal pelage. We conclude that within genetically uniform integumental tissues, microenvironmental differences occur and are reflected as alterations in the metabolic pattern of differentiated cells.     

Preservation of proteins in mummified tissues

Protein material was extracted from the dessicated tissues of several Egyptian mummies and a frozen Eskimo. The distribution and degree of preservation of high molecular weight protein was analyzed by gel filtration, protein assays, amino acid analysis, and polyacrylamide gel electrophoresis. The protein has undergone considerable degradation although some high molecular weight protein (C. 130,000 daltons) remains intact. Amino acid analysis of the extracted protein indicates the basic amino acids have undergone a chemical modification and may represent a point of preferential breakdown in the polypeptide chain. Atomic absorption spectrophotometry of tissue cations suggests a correlation between degree of preservation of mummified tissue and levels of sodium salts (natron) in the tissue.     

Hormonal effects on diosgenin biosynthesis and growth in Dioscorea deltoidea tissue cultures

Analysis of Dioscorea deltoidea tissue cultures grown in the presence of 2,4-D, indole-3-butyric acid, isopentenyladenine, benzyladenine and GA singly and in combination showed that the medium with 2,4-D most consistently favored diosgenin production. GA and high benzyladenine concentrations were toxic.     

Conditions affecting primary cell cultures of functional adult rat hepatocytes

Summary The conditions for obtaining representative, primary adult rat hepatocyte cultures were explored. The methods applied included enzymatic liver perfusion which was nondestructive to hepatocytes, the prevention of aggregation of dissociated cells and the selective attachment of viable cells. These procedures yielded a recovery of 50% of the liver cells which gave rise to cultures representing 14% of the total liver cells. The cultures were composed of homogeneous epithelial-like cells cytologically similar to hepatocytes and possessed a number of liver-specific enzymes. There was virtually no cell division initially and most cells died between 24 and 48 hr. Insulin enhanced the attachment of the liver cells, altered their morphology, but did not prolong cell survival. This study was supported by grant no. BC 133 from the American Cancer Society.     

Growth of human muscle in tissue culture

Summary A method is described for the culture of normal and diseased human muscle cells. Cell outgrowth was obtained from 63/63 biopsies, and cells differentiated to form myotubes in 57/63 biopsies. The culture technique used readily permitted the growth of both normal and diseased human muscle cells. This work was supported by grants from the Muscular Dystrophy Group of Great Britain and the Medical Research Council.     

Hormonal mechanisms for differentiation in plant tissue culture

Summary Cells possess extraordinary powers to organize their molecular processes not only to maintain a cell in a given steady state but also to recognize that state during differentiation. Regulation of these organizational forces appears to be under the control of chemical factors, and a hormonal concept of regulation has evolved. Hormones have been considered to act by reacting with a specific target site. This may be part of their mode of action, but I would like to suggest that a hormone enters and becomes part of a total molecular resonance system. In so doing, the entire molecular system of the cell is modified. Of the known plant hormones, the cytokinins, because of their role in experimentally induced cell division and differentiation, serve as a probe of hormonal involvement in differentiation. Cultured somatic cells of tobacco plants can be induced to undergo differentiation by addition of cytokinin and auxin to the medium. Studies of the cytokinin hormones show a series of diverse molecular involvements. The archetype cytokinin, N⁶-(Δ²-isopentenyl) adenosine (i⁶Ado), occurs in some molecular species of tRNA where it plays a vital role in the codon-anticodon interaction of tRNA and m-RNA. i⁶Ado under-goes extensive metabolism in the tobacco tissue. It is either degraded to adenosine or converted to derivatives that possess biological activity. It is perhaps, therefore, more correct to consider the hormone function as being derived from this total metabolic web. The normal somatic cells of tobacco cultures spontaneously change occasionally into an autonomous form that requires no external growth factors. This line of cells synthesizes i⁶Ado. The metabolic web of the hormone-dependent strain can be perturbed by added auxin but such is not the case in the autonomous strain. These data provide some insight into the altered state of cytokinin activity in which a cell line changes into an autonomous form. Curiously, in become independent of the requirement for exogenous cytokinin, the autonomous tissue becomes sensitive to added cytokinin. i⁶Ado also inhibits the growth of lines of mammalian cancer cells grown in culture. Presented in the formal symposium on Information Transfer in Eukaryotic Cells, at the 26th Annual Meeting of the Tissue Culture Association, Montreal, Quebec, June 2–5, 1975.     

Fate of 3H-thymidine labelled myogenic cells in regeneration of muscle isografts

Summary Intact and denervated extensor digitorum longus (EDL) muscles of 20-day-old inbred Lewis-Wistar rats were labelled with ³H-thymidine. Ninety minutes after the injection of the isotope 4.0% of the nuclei were labelled in the intact (i.e. innervated) and 9.6% in the muscles, denervated 3 days before administration of the isotope. The labelled EDL muscles were grafted into the bed of the previously removed EDL muscles of inbred animals and these isografts were studied 30 days later.In the EDL muscles, regenerated from innervated isografts only occasionally labelled endothelial cells were found whereas in the muscles regenerated from denervated isografts also parenchymal muscle nuclei were regularly labelled. The incidence of labelled nuclei in the regenerated EDL muscles was, however, about 20 times lower than in the donor EDL muscles. The present experiments provide a direct proof of utilization of donor satellite cell nuclei for regeneration in grafted muscle tissue. With respect to the low incidence of labelled nuclei in regenerated EDL muscles, other sources of cells apparently also contribute to the regeneration process.     

Early development and pathogenesis of Lobatostoma manteri Rohde (Trematoda: Aspidogastrea)

The prosobranch Planaxis sulcatus is reported as a new natural host of Lobatostoma manteri at Heron Island, Great Barrier Reef. Planaxis sulcatus and Cerithium moniliferum were experimentally infected with large numbers of eggs. The larvae hatch in the stomach and migrate immediately along the ducts of the digestive gland into the digestive follicles. The larvae feed on the secretion and probably epithelial cells of the follicles. The acetabulum is used for adhesion to the epithelium and contributes to its erosion. In heavily infected snails, the digestive follicles disappear gradually and the larvae live in cavities lined by a flattened epithelium, parts of which show secretory activity. In snails dissected 47–49 and 65–66 days after infection, the cavities are fused, forming several large spaces which communicate with each other; only small parts of the epithelium are still secretory. Concentrations of amoebocytes occur in the walls of the digestive gland and in the wall between digestive gland and stomach of infected Planaxis. Some young worms were found in the stomach of Planaxis. No tissue reactions were seen around the stomach except in the wall between digestive gland and stomach. In Cerithium with 65–67 days old infection, the cavities contain much detritus and disintegrating cells, the epithelium is practically non-secretory and surrounded by loose connective tissue. In larvae with a body length of approximately 0·5–0·6 mm, the acetabulum begins to divide into alveoli and its anterior end grows forward; the anterior alveoli gradually increase in size and new alveoli are formed in the posterior undivided zone. In two specimens of approximately 1·3 mm body length, the whole adhesive disk was divided into half the number of alveoli usually found in adults. Allometric shifts during growth of the worms are analysed.     

Isoperoxidases from tobacco tissue cultures

Two anodic isoperoxidases (A₁ and A₂) from tobacco tissue culture W-38 and two cathodic isoperoxidases (C₃ and C₄) from tobacco suspension culture WR-132 have been separated and characterized. Molecular weights for each of the isoperoxidases have been determined by two different methods. Only C₄ contained a carbohydrate component. The substrate specificity and the pH optima for the four enzymes with each of five substrates were determined.