Low density vascular smooth muscle (VSM) cell cultures maintained on extracelIular-matrix (ECM)-coated dishes and plated in the presence of either plasma or serum will proliferate actively when serum-containing medium is replaced by a synthetic medium supplemented with three factors: high density lipoprotein (HDL, 250,ug protein/ml) ; insulin (2 .5 pg/ ml) or somatomedin C (10 ng/ml) ; and fibroblast growth factor (FGF, 100 ng/ml) or epidermal growth factor (EGF, 50 ng/ml) . The omission of any of these three factors from the synthetic medium results in a lower growth rate of the cultures, as well as in a lower final cell density once cultures reach confluence . When cells are plated in the total absence of serum, transferrin (10ILg/ml) is also required to induce optimal cell growth .The effects of the substrate and medium supplements on the life span of VSM cultures have also been analyzed . Cultures maintained on plastic and exposed to medium supplemented with 5% bovine serum underwent 15 generations . However, when maintained on ECM-coated dishes the serum-fed cultures had a life span of at least 88 generations. Likewise, when cultures were maintained in a synthetic medium supplemented with HDL and either FGF or EGF, an effect on the tissue culture life span by the substrate was observed . Cultures maintained on plastic underwent 24 generations, whereas those maintained on ECM-coated dishes could be passaged repeatedly for 58 generations . These experiments demonstrate the influence of the ECM-substrate not only in promoting cell growth but also in increasing the longevity of the cultures .Previous studies have shown that bovine vascular smooth muscle (VSM) cells maintained on dishes coated with extracellular matrix (ECM) proliferate equally well when exposed to either plasma or serum (13). Among the plasma components that could affect the proliferation of VSM cells are the high (HDL) and low density lipoproteins (LDL), both of which have been reported to be mitogenic for that cell type when cultures are exposed to lipoprotein-deficient serum (LPDS ; 3, 5, 28) or to serum from abetalipoproteinemic subjects (24). Insulin, epidermal growth factor (EGF), and fibroblast growth factor (FGF) might also affect VSM cell proliferation, inasmuch as these agents have been shown to be mitogenic for a wide variety of cell types maintained under serum-free conditions (1, 2, 31) as well as for VSM cell cultures exposed to plasma (16,18) . Transferrin could also be required (1, 2, 31) to make iron available to the cells.To define which of these factors are important for proliferation of VSM cells, we have studied the proliferative response of low density VSM cell cultures that were plated in the presence of serum or plasma, and then exposed to a synthetic medium supplemented with LDL, HDL, insulin, and FGF, and have compared the effects of the substrate (either ECM or plastic) on the proliferative response to these agents. Our results indicate that VSM cells maintained on ECM-coated dishes proliferate actively when e...
Experimental conditions have been defined that allow bovine corneal endothelial (BCE) cells to grow in the complete absence of serum. Low density BCE cell cultures maintained on extracellular matrix (ECM)-coated dishes and plated in the total absence of serum proliferate actively when exposed to a synthetic medium supplemented with high density lipoprotein (HDL 500 micrograms protein/ml), transferrin (10 micrograms/ml), insulin (5 micrograms/ml), and fibroblast (FGF) or epidermal growth factor (EGF) added at concentrations of 100 or 50 ng/ml, respectively. Omission of any of these components results in a lower growth rate and/or final cell density of the cultures. BCE cell cultures plated on plastic dishes and exposed to the same synthetic medium grow very poorly. The longevity of BCE cultures maintained on plastic versus ECM and exposed to serum-free virus serum-containing medium has been studied. The use of ECM-coated dishes extended the life span of BCE cultures maintained in serum-supplemented medium to over 120 generations, as compared to less than 20 generations for cultures maintained on plastic. Likewise, BCE cells maintained on ECM and exposed to a synthetic medium supplemented with optimal concentrations of HDL, transferrin, insulin, and FGF underwent 85 generations, whereas control cultures maintained on plastic could not be passaged. The enhancing effect of ECM on BCE cell growth and culture longevity clearly illustrates the importance of the cell substrate in the control of proliferation of these cells.
Zooplankton are collected and sorted into two size fractions from which samples are randomly alloted to a battery of commonly used preservation techniques. We determine dry weight, ash content, and caloric content. We compute organic and inorganic losses of the samples to examine potential causes of variation in dry weight estimates. Treatments are: no preservation, preservation with one of three chemicals (75% ethanol, 5% or 10% buffered formaldehyde), preservation for 1 or 66 wk, and oven- or freeze-drying. Overall dry weight losses are independent of preservation methods. Chemical preservation reduces dry weight by 37 to 43%. Organic and inorganic losses range from 25 to 33% and 73 to 82%, respectively. Because inorganic losses are large, chemical preservation increases the caloric content of samples by 13 – 27%. Dry weight losses are somewhat size-dependent (37 versus 43% for the large and small size fraction respectively, after 66 wk of preservation). A regression of percent dry weight losses on body length (in millimetres) is obtained for our data, and published reports where formaldehyde is used as a preservative. It is: In[dry weight loss] = 4.149 − 0.576 length0.333. This relationship can be used to adjust the weight of zooplankton samples which have been preserved chemically. A survey of studies published in 1983 indicates that most authors did not adjust for dry weight losses due to preservation.
Transparency reduces the chances of detection of large planktonic animals by visual predators. An important constraint on the transparency of planktonic animals may be ingested food which could be seen through the body, thereby increasing the vulnerability of transparent zooplankton to visual predators. To test this hypothesis, we presented fed and un-fed Chaoborus larvae to juvenile coho salmon (Oncorhynchus kisutch). Overall, the presence of prey in the gut of Chaoborus increased their probability of capture by 68%. Predation risks due to the visibility of ingested food increased in proportion to meal size: larvae with nearly full gut were captured about three times faster on the average than larvae which had little food in their gut. Although Chaoborus larvae may be able to reduce this increased predation risk by migrating downward to low light levels, this behavior would reduce feeding opportunities by removing the larvae from surface waters where prey density is generally high. In this way, visual predators may limit the growth and the maximum size that can be achieved by transparent animals.
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