Transmucosal triglyceride transport rates in proximal and distal rat intestine in vivo.

Transmucosal transport rates for triolein in proximal and distal intestine were compared in unanesthetized rats. Emulsified [1-14-C] triolein together with bile and pancreatic juice from donor rats was infused for 6 hr into either the duodenum or the midpoint of the small intestine at such a rate that absorption was essentially complete in both regions of the intestine. Lymph was collected from the thoracic duct during triolein infusion and for an additional 6-hr period. The decrease in the rate of lymphatic output of labeled fat was found to follow a simple exponential function in all animals. This rate of decrease (decay rate) was used to calculate the half-times of lipid turnover through the intestinal wall and the fractional output rates. Distal intestine transported lipid 40% more slowly than proximal intestine, and the difference was associated with a greater accumulation of triglyceride in the distal intestinal wall. Chylomicron synthesis and/or release is the rate-limiting step for distal lymphatic fat transport in vivo, whereas fat uptake from the lumen is rate limiting for proximal intestine.     

Starch Binding Domain-containing Protein 1/Genethonin 1 Is a Novel Participant in Glycogen Metabolism

Stbd1 is a protein of previously unknown function that is most prevalent in liver and muscle, the major sites for storage of the energy reserve glycogen. The protein is predicted to contain a hydrophobic N terminus and a C-terminal CBM20 glycan binding domain. Here, we show that Stbd1 binds to glycogen in vitro and that endogenous Stbd1 locates to perinuclear compartments in cultured mouse FL83B or Rat1 cells. When overexpressed in COSM9 cells, Stbd1 concentrated at enlarged perinuclear structures, co-localized with glycogen, the late endosomal/lysosomal marker LAMP1 and the autophagy protein GABARAPL1. Mutant Stbd1 lacking the N-terminal hydrophobic segment had a diffuse distribution throughout the cell. Point mutations in the CBM20 domain did not change the perinuclear localization of Stbd1, but glycogen was no longer concentrated in this compartment. Stable overexpression of glycogen synthase in Rat1WT4 cells resulted in accumulation of glycogen as massive perinuclear deposits, where a large fraction of the detectable Stbd1 co-localized. Starvation of Rat1WT4 cells for glucose resulted in dissipation of the massive glycogen stores into numerous and much smaller glycogen deposits that retained Stbd1. In vitro, in cells, and in animal models, Stbd1 consistently tracked with glycogen. We conclude that Stbd1 is involved in glycogen metabolism by binding to glycogen and anchoring it to membranes, thereby affecting its cellular localization and its intracellular trafficking to lysosomes.     

Evidence for multiple lytic pathways used by cytotoxic T lymphocytes

Previous data generated by ourselves and others questioned the role of degranulation as a mechanism to explain CTL-mediated cytotoxicity. In this report we examine this tissue in greater depth. CTL-mediated lysis was probed with three different inhibitors. 4,4'-diisothiocyano-2,2'-disulfonic acid stilbene inhibits degranulation in a wide range of cell types, including CTL. EGTA, through chelation of Ca2+, also inhibits degranulation processes in CTL, and would inhibit other events or processes dependent on extracellular Ca2+. We also used prolonged exposure to PMA to exhaust PKC activity in CTL. Using these inhibitors, we have defined three pathways of lysis used by CTL. One pathway requires Ca2+, is PMA sensitive, but does not depend on degranulation. The second pathway is independent of Ca2+, is not PMA sensitive, and also does not depend on degranulation. All primary CTL and cloned CTL lyse most target cells via pathway I. However, when confronted with certain target cells (which we have referred to previously as Ca2+-independent target cells), pathway II is induced. When pathway II is induced, pathway I apparently shuts down. We show here that pathway II does not depend on protein synthesis, and that it also leads to DNA solubilization in target cells. A limited number of cloned CTL use pathways I and II as just described, but use in addition, and simultaneously, a third pathway that appears to involve degranulation. This pathway is seen irregularly in most CTL clones, and may be influenced by levels of IL-2 in the culture medium.     

Covalent interaction of L-2-amino-4-oxo-5-chloropentanoic acid with rat renal phosphate-dependent glutaminase. Evidence for a specific glutamate binding site and of subunit heterogeneity

Rat renal phosphate-dependent glutaminase is rapidly inactivated by incubating with L-2-amino-4-oxo-5-chloropentanoic scid. Concentrations of phosphate, which increase the glutaminase activity, decrease the rate of inactivation by chloroketone. In addition, inactivation is not blocked by glutamine. Instead, glutamate was shown to specifically reduce the rate of chloroketone inactivation. Upon sodium lauryl sulfate-polyacrylamide gel electrophoresis, the purified glutaminase preparation exhibits at least five protein staining bands which range in molecular weight from 57,000 to 75,000. Studies with 14C-labeled chloroketone indicate that this reagent reacts with each of these peptides. The mean stoichiometry of binding was calculated to be 1.3 mol/mol of enzyme. Therefore, these results indicate that the glutaminase may contain a specific site for binding glutamate and that the purified enzyme consists of a series of related peptides which may have resulted from partial proteolysis.     

Observations on lesser-known flatworms: temnocephala

Temnocephala novae-zealandie, a flatworm epizoic on crayfish, was examined by light and electron microscopy to investigate the variation within rhabdocoel turbellarians and to provide information on the possible structural modifications in the evolution of parasitism. The sucker is clearly glandular; the tentacular glands are eosinophilic and at the surface store and often release mucus as a coiled thread; the epidermis is clearly not reduced in structure and contains septate junctions in the anterior portion. Light microscope studies documented the presence of ten pair of paranephrocytes (athrocytes). In the laboratory, egg deposition, survival on and apart from the host and another crayfish, and behavior during the host molt were observed.     

Effects of capture on biological parameters in free-ranging bighorn sheep (Ovis canadensis): evaluation of normal, stressed and mortality outcomes and documentation of postcapture survival

Blood samples and physiological data were collected from 634 bighorn sheep (Ovis canadensis) captured by four different methods between 1980 and 1986 in the western United States. These parameters were evaluated for selected physiological, biochemical and hematological values. Postcapture biological parameters were compared among bighorn sheep according to four different outcomes; normal, stressed or compromised, capture myopathy (CM) mortality, and accidental mortality. Significant differences (P less than 0.05) were noted between outcome groups relative to certain parameters: temperature, respiration, creatinine phosphokinase (CPK), lactic dehydrogenase (LDH), serum glutamic oxaloacetic transaminase (SGOT), blood urea nitrogen (BUN), glucose, white blood cell count (WBC) and plasma pH. Such differences between groups may help in evaluating the clinical status of bighorn sheep at capture, enabling one to predict those animals that might develop CM at a later date, indicate candidates for preventive medical treatment prior to release, and/or which should be followed closely to determine long-term survival. Evaluation of follow-up data (n = 77) related to outcome status and long-term survival of bighorn sheep indicated that less than 4% (3 of 77) were dead within 1 mo of capture (one of these had been classified as normal and two as stressed or compromised at capture); less than 3% (3 of 77) were dead greater than 1 mo, and less than 6 mo after capture two were classified in the stressed outcome and one as diseased. Eighty-eight percent (68 of 77) were alive from 1 mo to 5 yr after capture (53 were classified as normal, 12 as stressed or compromised and 3 as diseased), and 2% (1 of 77) had chronic CM but was still alive (this animal had been classified as normal). Of 77 sheep in the follow-up group, less than 3% (2 of 77) were not observed following capture (one was classified as normal and one as stressed and diseased). Of the fatalities, less than 3% (2 of 40) had been captured by the net-gun and less than 4% (1 of 27) by drive-net. Those two unobserved in the follow-up group also had been caught with the net-gun, 5% (2 of 40). The single surviving CM case had been captured by the net-gun. Although the net-gun appears to be one of the safest methods of capturing individual bighorn sheep, based on evaluation of capture data and biological parameters, it may not be associated with the best long-term survival in some bighorn sheep.(ABSTRACT TRUNCATED AT 400 WORDS)