Muscle type dependent increase in intramyocellular lipids during prolonged fasting of human subjects: a proton MRS study.

The amount of intramyocellular lipids in skeletal muscle was assessed by proton magnetic resonance spectroscopy during a voluntary fasting period of 120 h in four healthy lean volunteers. The aim of the study was to determine whether muscular lipid uptake in the presence of high plasma lipid levels, or lipid oxidation due to lacking glycogen as a source of energy in musculature, are the dominant effects on intramyocellular lipid levels under fasting conditions in various muscle types. Intramyocellular lipids were quantified in the tibialis anterior (mixed type I and type II fibers, predominantly type II) and the soleus muscle (predominantly type I fibers) before and after 24 h, 72 h, and 120 h of fasting. An extreme increase in intramyocellular lipids to levels of 369 % (median) was found in the tibialis anterior muscle compared to baseline value (intramyocellular lipid level prior to fasting, set to 100 %; p = 0.02). The soleus muscle with clearly higher baseline content of intramyocellular lipids (2 - 4-fold compared to tibialis anterior) revealed slightly delayed and less pronounced uptake of intramyocellular lipids during fasting to 152 % (median) after 120 h (p = 0.02). The absolute increment in intramyocellular lipids (in terms of ratios between lipid and creatine signals) was also higher in tibialis anterior than in soleus (not statistically significant). These findings indicate augmentation of the intramyocellular lipid pool during long-term elevation of plasma FFA in the presence of low plasma insulin concentrations in both muscles investigated. The rate of muscular lipid oxidation during fasting is clearly lower than the increased uptake of FFA by myocytes.     

Fatty acylation of heparan sulfate proteoglycan from human colon carcinoma cells

A number of transmembrane proteins have been recently reported to be modified by the covalent addition of saturated fatty acids which may contribute to membrane targeting and specific protein-lipid interactions. Such modifications have not been reported in cell-associated heparan sulfate proteoglycans, although these macromolecules are known to be hydrophobic. Here, we report that a cell surface heparan sulfate proteoglycan is acylated with both myristate and palmitate, two long-chain saturated fatty acids. When colon carcinoma cells were labeled with [3H]myristic acid, a significant proportion of the label was shown to be specifically incorporated into the protein core of the proteoglycan. Characterization of fatty acyl moiety in the purified proteoglycan by reverse-phase high pressure liquid chromatography revealed that approximately 60% of the covalently bound fatty acids was myristate. We further show that this relatively rare 14-carbon fatty acid was bound to the protein core via a hydroxylamine- and alkali-resistant amide bond. The remaining 40% was the more common 16-carbon palmitate, which was bound via a hydroxylamine- and alkali-sensitive thioester bond. Palmitate appeared to be added post-translationally and derived in part from intracellular elongation of myristate, a process that occurred within the first two hours and was insensitive to inhibition of protein synthesis. Acylation of heparan sulfate proteoglycan represents a novel modification of this gene product and could play a role in a number of biological functions including specific interactions with membrane receptors and ligand stabilization.     

Isolation and characterization of temperature-sensitive mutants of adenovirus type2.

Fourteen temperature-sensitive mutants of human adenovirus type2, which differed in their plaquing efficiencies at at the permissive and nonpermissive temperatures by 4 to 5 orders of magnitude, were isolated. These mutants, which could be assigned to seven complementation groups, were tested for their capacity to synthesize adenovirus DNA at the nonpermissive temperature. Three mutants in three different complementation groups proved deficient in viral DNA synthesis. The DNA-negative mutant H2ts206 complemented the DNA-negative mutants H5ts36 and H5ts125, whereas mutant H2ts201 complemented H5ts36 only. Among the DNA-negative mutants, H2ts206 synthesized the smallest amount of viral DNA at the nonpermissive temperature (39.5 C). Data obtained in temperature shift experiments indicated that a very early function was involved in temperature sensitivity. In keeping with this observation, early virus-specific mRNA was not detected in cells infected with H2ts206 and maintained at 39.5 C. Prolonged (52 h) incubation of cells infected with H2ts206 at the nonpermissive temperature led to the synthesis of a high-molecular-weight form of viral DNA.     

Modulation of motilin-induced somatostatin release in dogs by naloxone

Somatostatin release in dogs is modulated by exogenous and endogenous opioids. Since postprandial somatostatin secretion is in part due to the stimulatory effect of postprandially activated gastrointestinal hormones as well as endogenous opioids, it was of interest to determine the interaction between motilin, a known stimulus of somatostatin release, and endogenous opioids with regard to activation of D-cell function. In a group of eight conscious dogs the infusion of synthetic porcine motilin at doses of 0.05, 0.25 and 0.5 micrograms/kg X hr elicited a significant increase of peripheral vein plasma somatostatin-like immunoreactivity (SLI), confirming previously reported data. The additional infusion of the opiate receptor antagonist naloxone attenuated this SLI response, suggesting that endogenous opioids participate in motilin-induced SLI release. Since previous studies have shown that the interaction between endogenous opioids and postprandial somatostatin secretion is modified by elevated plasma glucose levels, the experiments were repeated during an IV glucose (0.2 g/min) background infusion increasing circulating glucose levels by 20-30 mg/dl. During IV glucose, the SLI response to motilin was almost abolished. In this group the addition of naloxone restored the SLI response, indicating that the inhibitory effect of elevated glucose on D-cell function is, at least in part, mediated by endogenous opioids. These data suggest that motilin has to be considered as one regulatory factor which participates in the previously observed interaction between glucose and endogenous opioids during postprandial SLI release.     

Activity of tumor-associated lymphoid cells at short intervals after administration of irradiated syngeneic and allogeneic tumor cells.

After a single intraperitoneal injection of irradiated tumor cells, host cells capable of responding against syngeneic tumors were detected in peritoneal exudates of mice. Although irradiation of the injected tumor prevented its overgrowth, it did not significantly alter the antigenicity of the tumor. Immunologic activities of tumor-associated host cells in the peritoneal cavity were continuously monitored, starting 48 hr after tumor administration. In vitro cell-mediated lysis of syngeneic tumors appeared as early as 3 days after irradiated tumor administration. In addition, peritoneal exudate cells from inoculated mice were capable of adoptively transferring immunity. Purification of these peritoneal exudate cells on nylon wool columns yielded a nonadherent Ig-negative lymphocyte fraction whose cytolysis was tumor-specific and T cell-associated. The macrophage-free lymphocyte fraction exhibited a higher in vitro activity against tumors than unpurified peritoneal exudates. This tumor-host system allowed the study of cells which directly interact with the tumor cells in vivo, starting shortly after tumor administration. The results reported in this paper show that tumor-associated lymphoid cells capable of mounting anti-tumor response in vivo and in vitro can be demonstrated as early as 3 days after tumor inoculation.     

New imaging techniques of fat, muscle and liver within the context of determining insulin sensitivity

Body fat distribution and ectopic fat deposition are important determinants of insulin sensitivity. Fat deposition in muscle and the liver, in particular, has been found to impair insulin signalling in these insulin-sensitive tissues. Thus, exact quantification of fat content may help to distinguish between different sites of insulin resistance. Increased fat deposition in the visceral compartment compared with the subcutaneous depot also represents an important factor leading to insulin resistance. Recent data clearly showed that visceral fat is a strong determinant of liver fat content. Exact quantification of fat distribution by magnetic resonance imaging and magnetic resonance spectroscopy may help to define distinct 'fat-distribution phenotypes'. This may allow a search for new candidate genes for type 2 diabetes mellitus and identify, at an early stage, individuals at risk for decline in insulin sensitivity.     

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