31P and 13C NMR studies of isolated perfused hematopoietic cells from leukemic mice

The myeloproliferative leukemic virus (MPLV) induces within 2-3 weeks a massive infiltration of the adult mouse liver by hematopoietic leukemic cells. Since the metabolism of the infiltrated organ might be correlated with an interaction of two cell populations, it was decided to study the isolated hematopoietic cells separately. The metabolism of these cells embedded in an agarose gel and perfused with labeled substrates was investigated using 31P and 13C NMR. Using [1-13C]glucose as precursor, sequential 13C NMR spectra showed that the hematopoietic cells were able to store glucose as [1-13C]glycogen and to metabolize it through the glycolytic pathway to give [3-13C]lactate as sole end-product. The liver neoglucogenic substrates: [2-13C]pyruvate and [3-13C]alanine are not metabolized by these cells. This suggests that the tricarboxylic acid cycle was not efficient. To investigate further the glycolytic properties of the cells, 10 mM sodium azide was added to the medium containing [1-13C]glucose. When compared to the aerobic conditions, a 40% decrease of nucleotides (0.10 vs 0.17 mumole NTP/10(9) cells), a degradation of [1-13C]glycogen and an increase of ca 35% of the glycolytic rate were observed. The analysis of 13C NMR spectra of the perfusates at the end of the perfusion indicates a total conversion of [1-13C]glucose into [3-13C]lactate and [3-13C]pyruvate under anaerobic conditions. These results permit a better understanding of the metabolism of the perfused leukemic livers which are extensively infiltrated by these hematopoietic cells.     

Adiponectin 基因转染对肌细胞糖原合成和葡萄糖氧化的影响

目的：研究adiponectin对C2C12肌细胞糖原合成和葡萄糖氧化的影响。 方法： 用阳离子脂质体介导转染和随后G418筛选建立稳定转染小鼠adiponectin cDNA真核表达质粒（pcDNA3.0-mad）及空载pcDNA3.0的C2C12细胞株并鉴定。C2C12肌细胞糖代谢实验分对照组、空载体组和pcDNA3.0-mad(mad)组共3组进行，每组又分 0、0.5、5、100 nmol/L胰岛素刺激4个亚组。通过液闪测定细胞合成的糖原中［14C］的放射活性和氧化产生的［14CO2］，分别检测肌细胞的糖原合成和葡萄糖氧化情况。 结果： Western blotting和免疫组化检测证实mad组细胞表达adiponectin蛋白。Mad组葡萄糖氧化量随胰岛素浓度增加的速率较其它两组快，对照、空载体和mad组线性回归系数分别为23.34、23.23和26.06。Mad组C2C12肌细胞基础状态下和胰岛素刺激下的葡萄糖氧化和糖原合成与其它两组无显著差异（P>0.05）。 结论： 转染adiponectin基因对C2C12肌细胞葡萄糖氧化和糖原合成无显著影响。     

Herpes simplex virus protein synthesis in the presence of 2-deoxy-D-glucose.

The proteins and glycoproteins induced by herpes simplex virus type 1 (HSV-1) were labeled with [¹⁴C]amino acids or [¹⁴C]glucosamine in the presence or absence of 2-deoxy-d-glucose (deoxyglucose) and analyzed by slab gel electrophoresis. In the presence of 0.1% deoxyglucose (6.1 mM), the major envelope glycoprotein (VP123, MW 123,000) labeled with [¹⁴C]glucosamine was shifted to a component of an apparent lower molecular weight (VP123′). In the presence of increasing concentrations of deoxyglucose, there was a progressive decrease in the amount of [¹⁴C]amino acids incorporated into polypeptides which normally band in the VP123 region. Concomitant with this decrease was an increase in [¹⁴C]amino acids incorporated into a polypeptide(s) of greater electrophoretic mobility and of an apparently lower molecular weight. The polypeptide(s) was designated DG92 (MW 92,000) and was found to be predominantly associated with the nuclear fraction of HSV-1-infected cells cultured in the presence of deoxyglucose. The effects of deoxyglucose on HSV-1 polypeptide synthesis could be prevented by the addition of mannose.     

Glucose load diverts hepatic gluconeogenic product from glucose to glycogen in vivo.

Intravenous or oral administration of concentrated glucose solution into fasted rats simultaneously injected with 14C-bicarbonate resulted in an inhibition of [14C]glucose release into the blood and in an accelerated [14C]glycogen formation associated with glycogen synthetase activation and phosphorylase inactivation in the liver. The specific activity of glycogen was much higher than that of blood glucose after the glucose load, indicating that glycogen originated from gluconeogenesis rather than blood glucose. These metabolic changes induced by the glucose load were not mediated by endogenous insulin because they were observed to the same extent in rats treated with anti-insulin serum. However, they were mostly, if not totally, abolished by adrenalectomy, which suppressed gluconeogenesis and glycogenesis. Glucose tolerance was markedly impaired not only by anti-insulin serum, which inhibits peripheral glucose utilization, but also by adrenalectomy, which affects hepatic metabolism. It is concluded that a glucose load diverts the final product of hepatic gluconeogenesis from blood glucose to liver glycogen; these metabolic changes in the liver are an important determinant of glucose tolerance.     

Effect of fasting and refeeding on glucose metabolism in rat aorta

The effect of refeeding on accumulation of [14C]glucose carbon, oxidation of [14C]glucose and glycogen concentration in rat aortic intima-media was studied in rats fasted for 3 days. Accumulation of glucose carbon and glucose oxidation were determined by incubating rat aorta in vitro for 2 h with 5.6 mM 14C-labelled glucose in the medium. Refeeding with standard pellets for 2-4 h augmented [14C]glucose accumulation in rat aorta but had no significant effect on glucose oxidation. The glycogen concentration in rat aorta tended to increase. After refeeding for 16 h both [14C]glucose incorporation and [14C]glucose oxidation were increased in rat aorta. Refeeding with carbohydrate-rich pellets (92% carbohydrate) for 3 h increased blood glucose more than did protein-rich (92% protein) pellets, whereas the rise in plasma insulin was about the same. The accumulation of [14C]glucose carbon measured during the subsequent in vitro incubation for 2 h was augmented after refeeding with protein-rich pellets and slightly reduced after carbohydrate-rich pellets. Refeeding of diabetic rats for 2 h with standard pellets increased plasma insulin and markedly increased blood glucose but had no effect on aortic [14C]glucose incorporation. Intravenous infusion of glucose in normal rats for 2 h markedly raised blood glucose but did not increase the aortic glucose incorporation. Raising the ambient glucose concentration from 5 to 10 mM during incubation of normal rat aorta in vitro for 2 h slightly decreased the [14C]glucose incorporation determined during a subsequent incubation for 2 h. These results suggest that refeeding with standard pellets augments glucose incorporation and glucose oxidation in rat aorta.(ABSTRACT TRUNCATED AT 250 WORDS)     

Glyconeogenesis from lactate in frog striated muscle.

The rates of 14carbon incorporation into CO2 and glycogen from [U-14C]-lactate and [1-14C]acetate in frog sartorius muscles were compared. The rates of incorporation into CO2 were similar, while the rate of incorporation of lactate into glycogen was more than 200-fold larger than that of acetate incorporation. It is concluded that the pathway of lactate incorporation into glycogen does not involve Krebs cycle intermediates and is extramitochondrial. To test the possibility that the pathway of lactate incorporation involves net reversal of a pyruvate kinase, the changes in phosphoenolpyruvate and pyruvate concentrations during stimulation of lactate incorporation into glycogen were measured. There wer none. The mass action ratio of pyruvate kinase was calculated. This value was two orders of magnitude from the equilibrium constant and it was concluded that reversal of pyruvate kinase was a very unlikely pathway. To test the possibility that a pathway involving the oxaloacetate-to-phosphoenolpyruvate step was involved the muscles were treated with 3-mercaptopicolinate, an inhibitor of phosphoenolpyruvate carboxykinase. The treatment resulted in decreased incorporation of lactate into glycogen.     

The use of 2-deoxy-D-glucose to assess changes in tumor target cell membranes in vitro.

Following short-term suspension culture, cells from the Balb/C sarcoma Meth A were allowed to incorporate both [14C] leucine and 2-deoxy-D-glucose-1-[3H] (2DG). The 2DG is trapped as a small anionic marker of the cytosol. Deviation from the kinetics of spontaneous efflux of the markers is interpreted as reflecting perturbation of the target cell membrane. In the presence of guinea pig complement and a rabbit antiserum to Meth A, enhanced 2DG efflux was effected in a titer comparable to that detected with a 51Cr-release assay. With a number of alloantisera and syngeneic immune sera, 2DG efflux was enhanced while 51Cr-release was unaffected. Only in the presence of syngeneic immune sera from mice bearing a low tumor mass, syngeneic splenic leukocytes effect a retardation in the spontaneous 2DG efflux. Sera from animals with a large tumor mass were ineffective. Effux of proteins labeled with [14C] leucine was not altered. The phenomenon was not dependent on the presence of a heat-inactivatable syngeneic complement source. The method described provides a sensitive probe of target cell membrane permeability in the tumor model studied. The phenomenon detected is the capacity of serum, sampled relatively early in syngeneic oncogenesis, to direct syngeneic splenic leukocytes to interact with the target cell membrane differentially altering its permeability to the small cytosol marker.     

Calcium exchange in vascular smooth muscle, action of noradrenaline and lanthanum.

1. The Na, K, Ca and Mg content and the 45Ca uptake and loss were determined in rat aortae incubated in physiological solution or in solution containing LaCl3 instead of CaCl2. 2. Aortae washed in La-solution contained less Ca and Na than controls in physiological solution, the K content was not modified and the Mg content was slightly decreased. 3. In 50 mM-La solution the 45Ca diffusion space was intermediate between the values found for the [14C]sorbitol space and the [14C]inulin space, indicating that there was no Ca entry within the cell nor Ca binding at superficial sites. 45Ca loss from the tissue was directly related to the La concentration. 4. Noradrenaline increased the rate of uptake of 45Ca into the Ca fraction resistant to displacement by La. This increase was dose dependent, a response of 50% of the maximum being produced by 2 x 10(-8) noradrenaline as for the contraction. In the presence of phentolamine, the dose-effect curves for the action of noradrenaline on 45Ca uptake were displaced in a manner characteristic of competitive antagonism. The rhoA2 for phentolamine was 7-8. 5. In physiological solution, the rate of loss of 45Ca, from the Ca fraction resistant to displacement by La, was increased by noradrenaline the ED50 was 2 x 10(-8) M, and the effect was abolished by phentolamine. 6. In view of the similarity of phentolamine rhoA2 estimated by measuring noradrenaline sensitive 45Ca uptake or noradrenaline evoked contraction, it is likely that the activation of alpha-adrenergic receptors is responsible for both effects.     

Influence of altered O2 tension on substrate metabolism in perfused rat lung.

Effects of hypoxia (1.5 h) on glucose and palmitate metabolism were investigated in perfused lungs from normal rats and rats exposed for 24 h to hypobaric conditions (simulated altitude of 24,000 ft). Hypoxic lungs were ventilated with 5% O2-5% CO2 and control lungs with 21% O2-5% CO2. Blood gases and pH remained stable during the 1.5-h perfusion period. Exposure of normal rat lungs to 1.5 h of in vitro hypoxia (blood Po2=34 mmHg) significantly increased lactate production and mean arterial pulmonary pressure, but did not alter glucose uptake, pyruvate levels, and oxidation of either [U-14C]glucose or [1-14C]palmitate to CO2. Incorporation of labeled glucose and palmitate into lung lipids was also unaltered. In contrast to normal lungs, prior exposure to hypoxia for 24 h and subsequent perfusion under hypoxic conditions significantly stimulated glucose uptake (74% increase), markedly increased glucose incorporation into lung lipids, and increased oxidation of glucose to CO2. Lactate/pyruvate ratios also showed a significant 38% increase. Lung glycogen was unchanged following 24 h hypoxia. These data indicate that adaptive changes occur in metabolic processes within the lung during acute changes in O2 tension.     

Myocardial adenosine salvage rates and restoration of ATP content following ischemia.

The isolated perfused rat heart was utilized to determine the maximum rate of adenosine incorporation into adenine nucleotides and the effect of ischemia on this rate. In aerobic hearts, the rates of [8-14C]adenosine incorporation into nucleotides in nanomoles/minute per gram dry tissue were ATP 34 +/- 2, ADP 6 +/- 0.4, AMP 3 +/- 0.3, and IMP, 1 +/- 0.2. Following ischemia these values were not significantly different except for the rate of incorporation into IMP, which doubled. The extent of adenosine deamination with one pass through the coronary vasculature was the same in aerobic and postischemic hearts: 2% and 7% of the perfusate adenosine was converted to hypoxanthine and inosine, respectively. These percentages were similar at 50, 100, and 200 micron adenosine. Perfusion of aerobic hearts for 5 h with adenosine did not change ATP concentrations. Therefore, [8-14C]adenosine incorporation into ATP in these hearts appeared to represent ATP turnover. In contrast, 5 h perfusion of postischemic hearts with adenosine restored ATP concentrations to control values. The synthesis rate calculated from the increase in ATP concentration was comparable to the synthesis rate calculated from [8-14C]adenosine incorporation. Thus, incorporation of [8-14C]adenosine into ATP in postischemic hearts represented net ATP synthesis.     

Evidence for the existence of alpha- and beta-adrenoceptors on neurones and glial cells of cultured rat central nervous system--an autoradiographic study

The cellular localization of the binding of radioactive noradrenaline and alpha- and beta-adrenoceptor antagonists was studied in organotypic cultures of rat cerebellum, brain stem and spinal cord using autoradiography. In cerebellar cultures, many neurones, which appeared to be Purkinje cells, were labelled by [3H]noradrenaline and by the beta-antagonists [3H]dihydroalprenolol and [3H]carazolol, whereas no binding of the alpha-antagonists [3H]prazosin and [3H]rauwolscine was detected. In cultures of spinal cord and brain stem, [3H]noradrenaline and the beta-antagonists were bound to many large neurones. Binding of [3H] alpha-antagonists was observed to a small number of brain stem and spinal neurones, the labelling being much weaker than that produced by the [3H] beta-antagonists. The antidepressant [3H]desmethylimipramine was bound to many neurones and glial cells in cerebellar, brain stem and spinal cord cultures. Glial cells also possessed binding sites for [3H]noradrenaline and alpha- and beta-adrenoceptor antagonists, findings that are consistent with recent electrophysiological observations which indicate the existence of alpha- and beta-adrenoceptors on cultured astrocytes.     

The effects of testosterone on insulin sensitivity in male rats.

In order to examine the effects of testosterone (T) on insulin sensitivity, male rats were castrated or sham-operated, and exposed to low or high doses of T to substitute normal or to produce high serum T concentrations. Insulin sensitivity was followed by euglycaemic, hyperinsulinaemic glucose clamp measurements. An index of insulin-stimulated glucose transport was obtained in the white gastrocnemius (WG), extensor digitorum longus (EDL), red gastrocnemius (RG) and soleus (SOL) muscles after a bolus dose of [2-3H]deoxyglucose (2-DOG) when steady state was obtained in the clamp measurements. Glycogen synthesis was followed similarly with [U-14C]glucose as a labelled precursor after isolation of glycogen in the muscles mentioned, and in the liver. Castration and high T were followed by a marked insulin resistance in the clamp measurements. This was paralleled by a diminished insulin stimulation of glucose incorporation into glycogen down to about 50% of control values, apparently equally pronounced in all muscles but not found in liver glycogen synthesis. 2-DOG uptake was diminished by castration in the WG and RG muscles but was unaffected by high doses of T. Substitution of castrated rats with a low dose of T, restoring their serum T concentrations to the normal range, completely abolished these perturbations of insulin sensitivity. It is concluded that T is an important regulator of muscular insulin sensitivity, which seems to be highest in a 'window' of normal serum T concentrations.     

Arachidonic acid metabolism by rat alveolar epithelial cells

Arachidonic acid release and metabolism by stimulated cultures of rat type II alveolar epithelial cells (94 +/- 2% pure) were studied. As compared with unstimulated cultures, a marked increase in the release of [14C]arachidonic acid from prelabeled cells was observed when the cells were incubated with the calcium ionophore A23187. Radioimmunoassay of unlabeled cultures demonstrated significant increases in the production of prostaglandin E2 greater than 6-Keto-prostaglandin F1 alpha greater than prostaglandin F2 alpha greater than thromboxane B2 with A23187 stimulation. Reverse-phase high performance liquid chromatography of media from cells prelabeled with [14C]arachidonic acid confirmed the identities and relative amounts of these metabolites. As expected, the production of these cyclooxygenase products was inhibited by indomethacin. Stimulation with A23187 led to no increment in immunoreactive leukotriene C4 production, but yielded a statistically significant but quantitatively small increment in leukotriene B4 production; its production by small numbers of contaminating macrophages cannot be ruled out. Analysis by high performance liquid chromatography of media from prelabeled cells after 30 minutes stimulation revealed no peaks of radioactivity coeluting with the lipoxygenase products leukotriene B4, leukotriene C4, or 5-, 12-, or 15-hydroxy-6,8,11,14-eicosatetraenoic acid. The results indicate that rat alveolar epithelial cells have the capacity to release arachidonic acid and metabolize it to an array of cyclooxygenase products. However, after stimulation, little or no lipoxygenase products accumulated in media. Thus, the alveolar epithelium may be a source of bioactive eicosanoids with potentially important roles in pulmonary physiology and pathophysiology.     

Incorporation of choline, serine, ethanolamine and inositol into phospholipids of isolated rat mast cells.

The incorporation of labelled phospholipid precursors, [Me-14C]-choline, L-[3(-14)C]-serine, [2(-14)C]-ethanolamine and [2(-3)H]-myoinositol into the phospholipids of isolated rat mast cells was studied. The label from the different precursors were found to be essentially associated with compounds with the t.1.c.-motility of the respective phospholipids. Whereas the incorporation of [Me(-14)c]-choline and L-[3(-14)C]-serine showed evidence of saturation the incorporation of [2(-14)C]-ethanolamine was linear with time (2 h) and it was not saturated by increasing the concentration from 0.07 mM to 2.07 mM. The incorporation of [2(-3)H]-myoinositol was stimulated by Ca2+ (1 mM) or Mg2+ (1 mM), while the incorporation of the other precursors was stimulated only in the presence of both Ca2+ and Mg2+ (1 mM). Antimycin A (1muM), an inhibitor of the respiratory chain, significantly ingibited the incorporation of [Me(-14)c]-choline, L-[3(-14)C]-serine and [2(-3)H]-myoinositol but not that of [2(-14)C]-ethanolamine. The experimental system used might be a useful model for studies on the turnover of membrane phospholipids during histamine release.     

S-phase transit times as a function of age in human diploid fibroblasts

The minimum time it takes a cell to pass completely through the S phase (MIN S) was examined in human diploid fibroblasts using a sequential [14C]thymidine, [3H]thymidine, [14C]thymidine labeling protocol. MIN S appeared to be around 6-8 h for both WI-38 and MRC-5 cells. In addition, MIN S did not increase in senescent cultures. Since damage to either DNA, its polymerases, or both would result in a reduction in the rate of DNA synthesis and a corresponding increase in MIN S, this suggests that in senescent cultures at least a portion of the cells contain DNA that is relatively undamaged and DNA polymerases that exhibit normal replicative kinetics.     

Continuous synthesis of glycogen by individual worm pairs of Schistosoma mansoni inside the veins of the final host

Hamsters infected with Schistosoma mansoni were operated upon to install a permanent canula into their blood stream. After recovery of the hamster, this canula was used for the injection of radioactively labelled glucose. In this way the glycogen metabolism of S. mansoni could be studied while the parasites remained undisturbed in their natural habitat. The consecutive injection of [U-14C]glucose and [1-3H]glucose permitted an analysis of possible changes in the glycogen synthesis of individual worm pairs with time. The results showed that the synthesis of glycogen by each worm pair was fairly constant with time. Furthermore, all individual worm pairs synthesised glycogen continuously; not even 2 min passed without its formation. Only small differences in glycogen synthesis were observed between parasites isolated from different locations in the veins of the hamster. These results exclude the possibility that the worm pairs had alternating periods of glycogen synthesis and degradation, and they also disprove the idea that synthesis and degradation occur at two different sites in the bloodstream of the hamster. The experiments further showed that glycogen synthesis was proportional to the amount of glycogen already present, which in turn was shown to be proportional to the size of the parasite. From this study it can be concluded that the replenishment of the endogenous glycogen reserves of S. mansoni is not induced by a marked decrease in the glycogen levels, but occurs slowly and continuously.     

Radiolabeling of Treponema pallidum (Nichols virulent strain) in vitro with precursors for protein and RNA biosynthesis.

We observed uptake of [U-14C]serine, U-14C-labeled amino acid hydrolysates, and [2-14C]uracil by virulent Treponema pallidum in vitro for at least 96 h. No uptake of [2-14C]thymine, [1-14C]pyruvate, [U-14C]pyruvate, and [2-14C]uridine was detected. Treponemal protein and RNA biosynthetic activity was identified by erythromycin inhibition of amino acid and uracil uptake. Radioactivity due to uptake of radiolabeled amino acids by residual testicular cells in the cultures remained at background levels regardless of the presence or absence of cycloheximide. Accumulation of the radiolabeled substrates by T. pallidum proceeded at a linear rate for 48 to 96 h during incubation in vitro. The longevity of substrate uptake using the system of incubation described will facilitate future studies on the metabolism of the microorganism to help determine essential growth factors and environmental conditions for multiplication of T. pallidum in vitro.     

Release and turnover of noradrenaline in isolated median eminence: lack of negative feedback modulation

A low volume (tissue holder, 100 microliter; dead space, 300 microliter) perfusion system has been developed for measuring [3H]noradrenaline release from isolated median eminence, where supramaximal electrical field stimulation can be applied. In tissue preloaded with [3H]noradrenaline, the resting release (0.4-2% of the content) was enhanced by electrical stimulation (2-10-fold increase). That the released radioactivity in response to electrical stimulation is mainly due to release of [3H]noradrenaline was confirmed by high pressure liquid chromatography combined with radiochemical detection. Evidence has been obtained that of the stimulation-evoked release of radioactivity 70-80 percent originates from noradrenergic neurons, however, the release observed at rest was not affected by 6-hydroxydopamine pretreatment. 6-Hydroxydopamine pretreatment selectively reduced the concentration of noradrenaline of the median eminence without affecting its dopamine content. The release evoked by electrical stimulation was [Ca2+]- and tetrodotoxin-sensitive. 4-Aminopyridine enhanced both the resting and stimulation-evoked release. The ratio between the amount of [3H]noradrenaline released by two consecutive stimulation periods at 2 Hz (120 shocks) was constant, 0.94 +/- 0.08. In contrast with other noradrenergic axon terminals, the release of [3H]noradrenaline in the median eminence was not subject to negative feedback modulation, yohimbine and xylazine had no effect. This conclusion was substantiated by in vivo study showing that yohimbine, an alpha2-adrenoceptor antagonist enhanced the turnover rate of noradrenaline in the cortex but not in the median eminence. Since noradrenergic axon terminals in the median eminence do not make synaptic contact and the released noradrenaline does not modulate its own release via alpha2-adrenoceptors, it is an interesting anatomical arrangement: the modulatory alpha2-adrenoceptors are located exclusively on the terminals of the hormone-containing neurons.     

Effects of Endotoxin on Gluconeogenesis, Glycogen Synthesis, and Liver Glycogen Synthase in Mice

This study was undertaken to characterize the nature of carbohydrate loss due to endotoxin poisoning in mice and to elucidate mechanisms responsible for the changes. Female ICR mice, fasted overnight, were injected intraperitoneally with a mean lethal dose of endotoxin extracted from Salmonella typhimurium strain SR-11. Liver glycogen levels, alanine-U-(14)C and pyruvate-2-(14)C incorporation into blood glucose and liver glycogen, glucose-U-(14)C incorporation into liver glycogen, and liver glycogen synthase activities were measured at intervals after treatment. Liver glycogen in fasted mice given endotoxin was diminished significantly as early as 1 h after treatment. Liver glycogen synthase was significantly decreased in poisoned mice at 17 h. The use of actinomycin D showed that the induction of this enzyme due to fasting or hydrocortisone, or both, was inhibited by endotoxin. The incorporation of the (14)C-label from alanine-U-(14)C, pyruvate-2-(14)C, or glucose-U-(14)C into blood glucose and liver glycogen was substantially impaired in endotoxemic animals at 12 h. Decreases in incorporation occurred as early as 4 h after treatment. The progressive increase in glycogen synthase activity observed in fasted controls was not seen in endotoxin-poisoned mice. The administration of a glucose or pyruvate load to endotoxin-treated mice did not restore gluconeogenesis, glycogen synthesis, or liver glycogen synthase activity to normal levels. The in vivo activation of glycogen synthase by glucose was significantly reduced in endotoxemic animals. These changes indicate reduced carbohydrate synthesis as a probable cause for rapid sugar loss during endotoxemia in mice.     

Herpes simplex virus type 1 infection of endothelium reduces collagen and fibronectin synthesis

Protein synthesis was assessed in virus-infected and -uninfected cultures of bovine aorta endothelial cells by following the incorporation of [14C]proline into nondialyzable protein and by use of an ELISA assay. Monolayers were infected at confluency by incubating with herpes simplex virus type I at a multiplicity of infection (ratio of virus to cells) of 1.0, 1 hour prior to the introduction of the [14C]proline. Samples were taken from infected and uninfected cultures at various time points postinfection from both medium and cell layer fractions and analyzed for total 14C-labeled protein, 14C-labeled collagen, and 14C-labeled fibronectin. Medium proteins were analyzed by polyacrylamide gel electrophoresis, gel filtration, and electroimmunoblotting. No change was detected in total [14C] incorporation into nondialyzable protein in infected compared with uninfected endothelial cells; however, there was a significant decrease in the synthesis of collagen and fibronectin in infected cultures. The hydroxy[14C]proline content of fractionated medium proteins showed no significant differences between infected and uninfected cultures; therefore, the decrease in collagen synthesis cannot be explained by increased collagen degradation. These data suggest that infection of bovine endothelial cells causes a reduction in the synthesis of collagen and fibronectin.