Stathmokinetic Analysis of Human Epidermal Cells in vitro

Proliferation kinetics of cultured human epidermal cells is characterized in quantitative terms. Three distinct subpopulations of keratinocytes, two of which are cycling have been discriminated by two parameter DNA/RNA flow cytometry. Based on mathematical modelling, the cell cycle parameters of the cycling subpopulations have been assessed from stathmokinetic data collected at different time points after initiation of cultures (7–15 days). the first subpopulation is composed of low-RNA cells which resemble basal keratinocytes of epidermis and which show some characteristics of stem cells; these cells have a mean generation time of approximately 100 hr. the second subpopulation consists of high-RNA cells, resembling stratum spinosum cells of epidermis, which have an average generation time of approximately 40 hr. the third subpopulation consists of non-cycling cells with G_o/G₁ DNA content, with cytochemical features similar to those of cells in granular layer of epidermis. The results based on modelling can reproduce with acceptable accuracy the actual growth curve of the cultured cell population. Analysis of kinetics and differentiation of human keratinocytes is of interest in view of the recent application of cultured epidermal cell sheets for transplantation onto burn wounds. the results of this study also reveal the existence of regulatory mechanisms associated with proliferation and differentiation in the cultured epidermal cell population.     

The centrifugal horizontal cells in the lobula plate of the blowfly, Phaenicia sericata

Intracellular recordings combined with iontophoretic injection of Procion Yellow M4RAN were used to study the anatomy and physiology of the centrifugal horizontal cells (CH-cells) in the lobula plate of the blowfly, Phaenicia sericata.Anatomy: The CH-cells comprise a set of two homolateral, giant visual interneurones (DCH, VCH) at the rostral surface of each lobula plate. Their extensive arborizations in the lobula plate possess bulbous swellings (boutons terminaux). The arborization of one cell (DCH) covers the dorsal, and the arborization of the other cell (VCH) the ventral half of the lobula plate. Their axons run jointly with those of the horizontal cells through the chiasma internum and the optic peduncle. Their protocerebral arborization possesses spines; they form a dense network together with the axonal arborization of the horizontal cells, a second type of giant homolateral cell most sensitive to horizontal motion. The protocerebral arborization of the CH-cells gives rise to a cell body fibre which traverses the protocerebrum dorsally to the oesophageal canal. The cell body lies on the contralateral side laterally and slightly dorsally to the oesophageal canal in the frontal cell body layer.Physiology: The CH-cells respond with graded potentials to rotatory movements of their surround. Cells in the right lobula plate respond with excitation (excitatory postsynaptic potentials, membrane depolarization) to clockwise motion (contralateral regressive, ipsilateral progressive), and with inhibition (inhibitory postsynaptic potentials, membrane hyperpolarization) to counterclockwise motion in either or both receptive fields; CH-cells respond to motion presented to the ipsilateral and/or contralateral eye. Cells of the left lobula plate respond correspondingly to the reverse directions of motion. Vertical pattern motion and stationary patterns are ineffective.The heterolateral H1-neurone elicits excitatory postsynaptic potentials in the DCH-cell; these postsynaptic potentials are tightly correlated 1:1 to the preceding H1-action potentíal. The delay between the peak of the action potential and the beginning of the DCH-postsynaptic potential is 1.15 msec, agreeing very well with the value reported previously for the blowfly, Calliphora (Hausen, 1976a). The synaptic input and output connections of the CH-cells are discussed.     

CAM-idling in Hoya carnosa (Asclepiadaceae)

In the leaf succulent Asclepiad Hoya carnosa (L.) R. Br., CAM photosynthesis occurred under well-watered conditions, as characterized by diurnal gas exchange and changes in titratable acidity. Following 10–12 days of severe water stress, the plants shifted from CAM to a modified CAM-idling mode of metabolism. CAM-idling was characterized by complete or almost complete stomatal closure accompanied by CAM-like diurnal changes in titratable acidity. H. carnosa plants maintained this CAM-idling mode of photosynthesis for at least 8 weeks. Upon reirrigation, the plants returned to the original CAM mode within 1 week. These results suggested that CAM-idling is a reversible, intermediate form of sustained metabolism which enables plant survival under conditions of extended drought.This work was supported in part by NSF Grant PCM 8200366 and in part by the Science and Education Administration of the United States Department of Agriculture under Competitive Grant 5901-0420-8-0018-0.     

Chimeric gene expression using maize intron in cultured cells of breadwheat

High voltage electrical pulses were used to introduce the CAT reporter gene into cultured protoplasts of breadwheat,Triticum aestivum. Four DNA constructs harboring the CAT gene and the 35S or mannipine synthase promoter were tested for levels of CAT activity 40–45 hr after electroporation of protoplasts. One construct, containing a maize intron sequence between 35S and CAT sequences, conferred 30 to 185 fold greater CAT activity over the other three constructs. Data from these experiments suggest that a maize intron or sequences with similar effects may be required in DNA constructs for efficient heterologous gene expression in cultured cells of breadwheat.Abbreviations CAT Chloramphenicol acetyl transferase - NPT II neomycin phosphotransferase - 35S the 35S promoter of Cauliflower Mosaic Virus - PEG Polyethylene glycol - MES 2-[N-morpholino] ethanesulfonic acid     

Nucleotide sequence of the Rickettsia prowazekii ATP/ADP translocase-encoding gene

The Rickettsia prowazekii ATP/ADP translocase (Tlc) gene (tlc), previously cloned in Escherichia coli was localized to a 1.6-kb chromosomal fragment. Nucleotide sequence analysis of this fragment revealed an open reading frame of 1494 bp that could encode a hydrophobic protein of 497 amino acids (aa) with an M_r of 56 668. Analysis of the deduced aa sequence revealed that it contained twelve potential membrane-spanning regions. Comparisons between the deduced aa sequence of the R. prowazekii ATP/ADP Tlc and the sequences of mitochondrial (mt) Tic revealed no detectable homologies between the rickettsial and mt sequences. The major protein synthesized in E. coli minicells containing the rickettsial gene exhibited an M_r of approx. 34000.     

Streptomycin and lincomycin resistances are selective plastid markers in cultured Nicotiana cells

Summary Resistance to streptomycin and lincomycin in plant cell culture is used as a color marker: resistant cells are green whereas sensitive cells are white on the selective medium. Streptomycin and lincomycin at appropriate concentrations do not kill sensitive Nicotiana cells. The selective value of plastid ribosomal DNA mutations, conferring resistance to streptomycin and lincomycin, was investigated by growing heteroplastidic cells on a selective medium. The heteroplastidic cells were obtained by protoplast fusion, and contained a mixed population of streptomycin resistant plastids from the N. tabacum line Nt-SR1-Kan2, and lincomycin resistant plastids from the N. plumbaginifolia line Np-LR400-Hyg1. Clones derived from protoplast fusion were selected by kanamycin and hygromycin resistance, transgenic nuclear markers. Somatic hybrids were then grown on a selective streptomycin or lincomycin medium, or in the absence of either drug to a 50 to 100 mg size callus. Southern analysis of a polymorphic region of plastid DNA (ptDNA) revealed that somatic hybrids grown on streptomycin contained almost exclusively ptDNA from the streptomycin resistant parent, somatic hybrids grown on lincomycin contained almost exclusively ptDNA from the lincomycin resistant parent whereas somatic hybrids grown in the absence of either drug contained mixed parental plastids. Sensitive ptDNA was below detection level in most clones on selective medium, but could be recovered upon subsequent culture in the presence of the appropriate drug. The drugs streptomycin and lincomycin provide a powerful selection pressure that should facilitate recovery of plastid transformants.     

Short-term influence of nitrate on acetylene reduction, net photosynthesis and nodule respiration in black alder (alnus glutinosa L. gaertn.) seedlings

The use of nitrogen-fixing trees such as black alder (Alnus glutinosa L. Gaertn.) as forest silvicultural tools has recently been recognized. The potential benefit of black alder in silvicultural practices may be reduced by nitrate fertilization. Fifteen-month-old, nodulated, black alder rooted cuttings were fertilized for 6 days with 0, 7.5 or 15 mM NO₃ to determine the influence of nitrate on acetylene reduction, nodule respiration and net photosynthesis. Acetylene reduction, net photosynthesis and nodule respiration were measured on the second, fourth and sixth days of nitrate application. Nitrate treatment significantly reduced acetylene reduction and nodule respiration by day 4. Acetylene reduction was 75% lower and nodule respiration 36% lower for the 15 mM NO₃ treatment when compared to that of the control treatment. By day 6, net photosynthesis and nodule respiration were significantly reduced by 29 and 59%, respectively, for seedlings treated with 15 mM NO₃. This study suggests that nitrate fertilization has a profound influence on nitrogenase activity and that nitrogen-fixing tree species may respond to nitrate fertilization by shifting photosynthetic rates.     

Lipid composition and food quality of some freshwater phytoplankton for cladoceran zooplankters

The nutritional value of several planktonic algae was testedby means of feeding trials with three cladoceran zooplankters.The algae were monocultures and included two blue-greens, fourgreens and four flagellates with a size range of 5–48µm. The specific growth rates of the zooplankters werechosen as the measure of the nutritional value of the algae.The three cladocerans showed large differences in growth ratein the different algae, but the two cryptomonads were withoutdoubt best suited as food for all. The fatty acid compositionfor the cryptomonads were different from the other algae. Theycontained high percentages of the polyunsaturated fatty acids20:5æ3 (EPA) and 22:6æ3 (DHA), which also are commonin fish. It is suggested that the lipid composition is a probablefactor determining the nutritional quality of the algae.     

The social development of free-ranging infantLemur catta at Berenty reserve,Madagascar

The social development of 11 free-ranging infantLemur catta was examined over the first 16 weeks of the infants' lives. By 16 weeks, infants still occasionally suckled and were carried dorsally, but on the whole, they were independent of their mothers. Sex or mother's rank was not found to affect frequency or type of play behavior. Mother's rank had no effect on frequency of maternal rejections, from the nipple or from riding, but female infants were rejected slightly more frequently than males were. Mothers tended to reject infants more severely and more frequently from dorsal riding than from the nipple. Sex and rank differences were not found with respect to behaviors determined as measures of independence; however, lower-ranking infants engaged in significantly more dependent behaviors than higher-ranking young did. It may be necessary for the infant of a low-ranking mother to maintain closer proximity to its mother for a longer period of time during infancy because such infants may be subject to abuse by higher-ranking group members and, furthermore, may not be as readily rescued in a stressful or dangerous situation as a higher-ranking infant. Sex was not found to be a factor in terms of measures of dependence. The lack of sex differences in developmental behaviors in this species may be related to female dominance, as well as to the fact that, as adults, both sexes engage in aggressive territorial behavirs.