Long-term in vitro cell culture of sinclair swine melanoma

Summary The melanoma of Sinclair swine exhibits several characteristics similar to human melanoma but demonstrates an unusually high incidence of spontaneous regression. A total of 66 finite cell lines derived from 21 swine melanotic lesions, both cutaneous and visceral, were studied in vitro over their life spans of up to 14 months. The growth characteristics of the cultures varied with the age of the swine from which the tumors were obtained. Cell cultures of tumors obtained from swine aged less than 2 months grew steadily in culture with a population-doubling time of 120 to 180 hr until growth and division ceased after a maximum of 25 to 35 population doublings (6 to 8 passages). Cell culture of tumors obtained from swine aged 3 months or older showed a biphasic growth pattern with an early slow growth rate (population-doubling time 120 to 160 hr), which shifted after 3 to 6 passages to a faster rate (80 to 110 hr population-doubling time) until termination of growth and division after a maximum of 75 to 85 population doublings (18 to 20 passages). The cultures were morphologically heterogeneous including cuboidal, spindle and dendritic cell types. Electron microscopy showed classic melanosomes only in the primary and passage 1 cultures although vesicular inclusions were numerous in later-passage cells. However, continued melanin synthesis was indicated by the spectroscopic characteristics of material obtained from medium of passage 8 cultures and by DOPA staining of cultures as advanced as passage 18. This work was supported by a grant from the NIH, NCI (2 P01 CA 08023-11A1).     

Protein metabolism. 6. Trichostrongylus colubriformis: skeletal protein catabolism in primary and secondary infections of the guinea pig

The urinary excretion of 3-methylhistidine (3-MH) was used as an index of muscle protein catabolism in primary and secondary infections of the guinea pig with Trichostrongylus colubriformis and in uninfected animals fed quantitatively reduced rations. Catabolism, which was depressed in all three groups, was directly related to a fall in food consumption. Possible explanations for the greater depression of catabolism in the primary infection than in the uninfected guinea pigs and its fall in the secondary infection in spite of little change in consumption are briefly discussed. It was concluded that the faster rate of whole-body protein turnover reported earlier in this series on protein metabolism in intestinal nematode infection was not partly due to a faster rate of muscle protein catabolism. It was shown that the urinary excretion of 3-MH could be validly expressed in terms of unit creatinine.     

Cycloheximide resistance in Chinese hamster cells. II. Induction of Chm resistance in Chinese hamster cells by N-nitrosomethylurea.

Mutant Chinese hamster ovarian (CHO) cells with a resistance to 7-10(-7) and 8-10(-7) M cycloheximide (CHM) were induced at mutation rates of 1.9-5.2-10(-3) and 1.6-1.8-10(-3) respectively after treatment with N-nitrosomethylurea (NMU) at 100 mug/ml. The induced mutation rates differed by two orders of magnitude from the spontaneous rate of mutation to CHM resistance.     

Heritability of the backtest response in piglets and its genetic correlations with production traits

The backtest response of a pig gives an indication of its coping style, that is, its preferred strategy to cope with stressful situations, which may in turn be related to production traits. The objective of this study was therefore to estimate the heritability of the backtest response and estimate genetic correlations with production traits (birth weight, growth, fat depth and loin depth). The backtest was performed by placing the piglet on its back for 60 s and the number of struggles (NrS) and vocalizations (NrV), and the latency to struggle and vocalize (LV) was recorded. In total, 992 piglets were subjected to the backtest. Heritability estimates for backtest traits were statistically moderate (although high for behavioral traits), with LV having the highest heritability estimate (0.56±0.10, P<0.001) and NrS having the lowest estimate (0.37±0.09, P<0.001). Backtest traits also had high genetic correlations with each other, with vocalization traits (NrV and LV) having the highest (−0.94±0.03, P<0.001), and NrS with NrV the lowest correlation (0.70±0.09, P<0.001). No significant correlations were found between backtest traits and production traits, but correlations between NrS and birth weight (−0.38±0.25), and NrV and loin depth (−0.28±0.19) approached significance (P=0.07). More research into genotype-by-environment interactions may be needed to assess possible connections between backtest traits and production traits, as this may depend on the circumstances (environment, experiences, etc.). In conclusion, heritability estimates of backtest traits are high and it would therefore be possible to select for them. The high genetic correlations between backtest traits indicate that it may be possible to only consider one or two traits for characterization and selection purposes. There were no significant genetic correlations found between backtest traits and production traits, although some of the correlations approached significance and hence warrant further research.     

Bioavailability of two organic forms of zinc in comparison to zinc sulphate for weaning pigs fed a diet composed mainly of wheat,barley and soybean meal

This study was performed to compare the bioavailability of two organic zinc compounds, a zinc glycinate complex and a zinc amino acid chelate with that of zinc sulphate in growing pigs fed a basal diet composed mainly of wheat, barley and soybean meal. The experiment included 96 pigs with an average body weight of 8 kg, allotted to ten groups of nine to ten pigs each. The first group received the basal diet, containing 42 mg of native zinc per kg, without zinc supplementation over a period of five weeks. The other nine groups received the basal diet supplemented with 15, 30 or 50 mg of zinc/kg as zinc sulphate, zinc glycinate or the zinc amino acid chelate. Pigs fed the unsupplemented diet had a lower growth performance (body weight gain, feed conversion ratio) than the other nine groups. Supplementation of 15 mg zinc/kg diet (irrespective of zinc form) was sufficient to yield optimum growth performance. Plasma zinc concentration and activity of alkaline phosphatase were rising with increasing zinc supplementation levels up toa maximum reached at a supplementary level of 30 or 50 mg/kg diet for activity of alkaline phosphatase and plasma zinc concentration, respectively. The response of those parameters to zinc supplementation did, however, not differ between thethree zinc compounds considered. The apparent digestibility of zinc from the diet was also not different for the three zinc compounds. In conclusion, these findings show that the bioavailability of the two organic zinc compounds did not differ from that of zinc sulphate in growing pigs fed a diet with wheat, barley and soybean meal as major components.     

Effect of early antibiotic intervention on specific bacterial communities and immune parameters in the small intestine of growing pigs fed different protein level diets

Antibiotics are designed to affect gut microbiota and subsequently gut homeostasis. However, limited information exists about short- and long-term effects of early antibiotic intervention (EAI) on gut homeostasis (especially for the small intestine) of pigs following antibiotic withdrawal. We investigated the impact of EAI on specific bacterial communities, microbial metabolites and mucosal immune parameters in the small intestine of later-growth-stage pigs fed with diets differing in CP levels. Eighteen litters of piglets were fed creep feed with or without antibiotics from day 7 to day 42. At day 42, pigs within each group were offered a normal- or low-CP diet. Five pigs per group were slaughtered at days 77 and 120. At day 77, EAI increased Enterobacteriaceae counts in the jejunum and ileum and decreased Bifidobacterium counts in the jejunum and ileum (P < 0.05). Moreover, tryptamine, putrescine, secretory immunoglobulin (Ig) A and IgG concentrations in the ileum and interleukin-10 (IL-10) mRNA and protein levels in the jejunum and ileum were decreased in pigs with EAI (P < 0.05). At day 120, EAI only suppressed Clostridium cluster XIVa counts in the jejunum and ileum (P < 0.05). These results suggest that EAI has a short-term effect on specific bacterial communities, amino acid decarboxylation and mucosal immune parameters in the small intestine (particularly in the ileum). At days 77 and 120, feeding a low-CP diet affected Bifidobacterium, Clostridium cluster IV, Clostridium cluster XIVa and Enterobacteriaceae counts in the jejunum or ileum (P < 0.05). Moreover, feeding a low-CP diet increased the concentrations of Igs in the jejunum and decreased pro-inflammatory cytokines levels in the jejunum and ileum (P < 0.05). At day 120, feeding a low-CP diet increased short-chain fatty acid concentrations, reduced ammonia and spermidine concentrations and up-regulated genes related to barrier function in the jejunum and ileum (P < 0.05). These results suggest that feeding a low-CP diet changes specific bacterial communities and intestinal metabolite concentrations and modifies mucosal immune parameters. These findings contribute to our understanding on the duration of the impact of EAI on gut homeostasis and may provide basis data for nutritional modification in young pigs after antibiotic treatment.     

Changes in soil properties and resistance to concentrated flow across a 25‐year passive restoration chronosequence of grasslands on the Chinese Loess Plateau

Revegetation represents an effective measure for preventing soil erosion on the Loess Plateau. However, the effects of revegetation‐induced changes in soil and root properties on soil resistance to concentrated flow erosion (SRC) remain unclear. This study sampled soils and roots across a 25‐year chronosequence from farmland to grasslands of different ages (3, 7, 10, 18, and 25 years) to quantify variations in soil and root properties (soil bulk density, SBD; soil disintegration rate, SDR; saturated hydraulic conductivity, SHC; organic matter content, OMC; water‐stable aggregate, WSA; mean weight diameter, MWD; root mass density, RMD; root length density, RLD; and root surface area density, RSAD) and their effects on SRC. Farmland and grassland SRCs were obtained using a hydraulic flume. Soil properties and root density gradually improved with restoration time. In terms of the comprehensive soil property index calculated via principal component analysis, grassland values were 0.66 to 1.94 times greater than farmland values. Grassland SRCs increased and gradually stabilized (>18 years) over time and were 1.60 to 8.26 times greater than farmland SRC. SRC improvement was significantly related to increases in OMC, SHC, WSA, and MWD and decreases in SBD and SDR over time. SRC was effectively simulated by the Hill curve of RMD, RLD, and RSAD. SDR, SHC, and RMD (0.5–1.0 mm) affected SRC the most. This study scientifically describes how revegetation improves soil quality and soil resistance to flow erosion, and suggests that vegetations rich in 0.5–1.0 mm roots should be preferred during revegetation.     

Improvement of Intracellular Traffic System by Overexpression of KDEL Receptor 1 in Antibody‐Producing CHO Cells

The localization of soluble endoplasmic reticulum (ER) chaperones in the cell organelle is mediated by the C‐terminal KDEL (lysine, aspartic acid, glutamic acid and leucine) motif. This motif is recognized by the KDEL receptor, a seven‐transmembrane protein that cycles between the ER and cis‐Golgi to capture missorted KDEL chaperones from post‐ER compartments in a pH‐dependent manner. The KDEL receptor's target chaperones have a substantial role in protein folding and assembly. In this study, the gene expression level of KDEL receptor 1 shows a moderate upregulation during either ER stress or growth of Chinese hamster ovary (CHO) cells in batch culture, while the ER chaperones show higher upregulation. This might indicate the possibility of saturation of the ER retention machinery or at least hindered retention during late stage batch culture in recombinant CHO cells. KDELR1 is overexpressed in a monoclonal antibody‐producing CHO cell line to improve the intracellular chaperone retention rate in the ER. An increase in the specific productivity of IgG1 by 13.2% during the exponential phase, and 23.8% in the deceleration phase of batch culture is observed. This is the first study to focus on the ER retention system as a cell engineering target for enhancing recombinant protein production.     

杉木采伐迹地营造阔叶树种对土壤微生物生态化学计量特征的影响

研究土壤微生物生物量及其生态化学计量特征对造林树种转换的响应,对深入了解森林生态系统土壤养分循环和有效性具有重要意义。本研究以1993年春在二代杉木采伐迹地上营造的26年生米老排和杉木人工林为对象,采用氯仿熏蒸法测定了0～40 cm土层土壤微生物生物量碳(MBC)、氮(MBN)、磷(MBP)的变化。结果表明: 与杉木人工林相比,米老排人工林0～10 cm土层MBC和0～20 cm土层MBN和MBP均显著提高, 0～20 cm土层MBC/MBP和10～20 cm土层MBN/MBP显著降低。两种人工林所有土层MBC/MBN均无差异。相关分析显示,土壤含水率、总有机碳、总氮、全磷、有效磷与MBC、MBN和MBP呈显著正相关,而与MBC/MBP和MBN/MBP呈显著负相关。逐步线性回归分析表明,MBN和MBP主要受土壤总氮和有效磷的影响,而MBC/MBP和MBN/MBP主要受有机碳和有效磷的驱动。研究表明,造林树种从杉木转换成米老排能够增加表层土壤微生物生物量,加速氮磷养分周转,增加土壤氮磷养分供应能力。米老排人工林土壤MBP的增加可能在一定程度上缓解了树木生长的磷限制。