Use of phlorizin binding to demonstrate induction of intestinal glucose transporters

Summary We used specific binding of phlorizin to the intact intestinal mucosa in order to measure glucose transport site density in intestines of mice fed a high-carbohydrate or no-carbohydrate diet. Nonspecific binding varied with intestinal position but showed only modest dependence on diet. Specific binding to glucose transporters was 1.9 times greater in jejunum of high-carbohydrate mice than of no-carbohydrate mice; this ratio was the same as the ratio for V_max values of actived-glucose uptake between the two diet groups. The gradient in specific binding of phlorizin along the intestine paralleled the gradient in V_max of glucose transport. These results directly demonstrate that the increase in intestinal glucose transport caused by a high-carbohydrate diet is due to induction of glucose transporter. They also indicate that the normal positional graident in glucose transport along the intestine arises from a gradient in transporters, induced by the normal gradient in luminal glucose concentration.     

Calcium-ryanodine receptor complex. Solubilization and partial characterization from skeletal muscle junctional sarcoplasmic reticulum vesicles

The Ca2+-ryanodine receptor complex is solubilized in functional form on treating sarcoplasmic reticulum (SR) vesicles from rabbit fast skeletal muscle with 3-[(3-cholamidopropyl)dimethylammonio]-1-propane-sulfonate (CHAPS) (1 mg/mg protein) and 1 M NaCl at pH 7.1 by shaking for 30 min at 5 degrees C. The heavy membrane preparations obtained from pyrophosphate homogenates frequently exhibit junctional feet and appear to be derived primarily from the terminal cisternae of the SR. The characteristics of [3H]ryanodine binding are similar for the soluble receptor and the heavy SR vesicles with respect to dependence on Ca2+, pharmacological specificity for inhibition by six ryanoids and ruthenium red, and lack of sensitivity to voltage-dependent Ca2+-channel blockers, inositol 1,4,5-trisphosphate, or doxorubicin. In contrast, the cation sensitivity is decreased on receptor solubilization. The soluble receptor is modulated by cyclic nucleotides and rapidly denatured at 50 degrees C. Saturation experiments reveal a single class of receptors (Kd = 9.6 nM), whereas kinetic measurements yield a calculated association constant of 5.5 X 10(6) min-1 M-1 and a dissociation constant of 5.7 X 10(-4) min-1, suggesting that the [3H]ryanodine receptor complex ages with time to a state which is recalcitrant to dissociation. Sepharose chromatography shows that the receptor complex consists primarily of two protein fractions, one of apparent Mr 150,000-300,000 and a second, the [3H]ryanodine binding component, of approximately Mr 1.2 X 10(6). Preliminary analysis of the soluble receptor preparation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis reveals subunits of Mr greater than 200,000 and major bands of calsequestrin and Ca2+-transport ATPase. These findings indicate that [3H]ryanodine binds to the Ca2+-induced open state of the channel involved in the release of contractile Ca2+.     

Distinct groups of plasmids correlated with bacteriocin production in Staphylococcus aureus

The genetic basis of bacteriocin (Bac) production by six strains of Staphylococcus aureus was examined. Gene transfer experiments (in which the plasmids were tagged with the erythromycin resistance transposon Tn551) and plasmid-elimination experiments by growth at 43 degrees C associated bacteriocin production with a particular plasmid in each strain. The Bac plasmids could be separated into two distinct groups: the first comprised plasmids larger than 40 kb, which did not specify immunity to bacteriocins; the second comprised small plasmids (8.0-10.4 kb) which also specified immunity to bacteriocins. The sequence relations among the small plasmids (pRJ6, pRJ9, pRJ10 and pRJ11) were investigated by comparing restriction enzyme digest patterns and by hybridization. Plasmids pRJ10 and pRJ11 were indistinguishable and very closely related to plasmid pRJ9. Plasmid pRJ6, although different from the others, shared regions of sequence homology with them. No homology was found between plasmids pRJ6 or pRJ9 and the large Bac plasmids.     

PERFUSION FOR MYOCARDIAL REVASCULARIZATION WITHOUT AN ARTIFICIAL OXYGENATOR (New Method to Reduce Surgical Morbidity)

Thirteen patients were submitted to direct myocardial revascularization (saphenous vein graft) without the use of an artificial oxygenator. The perfusion was done by a left ventricle-to-aorta bypass and autogenous oxygenation. Most patients had three grafts implanted plus endarterectomy of the distal right coronary artery. There was one hospital death that was apparently not related to the method used. Perfusion time ranged from 45 minutes to 4 hours. Body temperature during perfusion was kept between 25 and 30 degrees C. Perfusion flow was maintained between 25 to 50 ml per kg of body weight per minute. Ischemic, hypothermic cardiac arrest was employed. We demonstrated for the first time that perfusion for this kind of heart surgery could be done with no artificial oxygenators and, apparently, is safer for the patients. There were no bleeding problems even in perfusions as long as 4 hours. There was no respiratory dysfunction, and artificial respiration was used for only 6 to 12 hours. The patients awoke at the end of surgery with no signs or symptoms of central nervous system damage, and vasopressor drugs were rarely used after surgery. Although the experience is very small, it suggests that many postoperative problems, especially those related to bleeding and respiratory dysfunction may be reduced or eliminated by this new method.     

Evaluation of several enrichment procedures for the isolation of recombinant plasmid DNA

A number of methods for the selective enrichment of recombinant plasmids were examined; these include alkaline phosphatase treatment of the restricted pBR322 vector, as well as a combination of this and S1 nuclease treatment of the ligated mixture of pBR322 and pCR1 plasmids orS. griseus DNA followed by D-cycloserine treatment to enrich for cells carrying recombinant molecules. The relative efficiencies of these methods were compared.     

Comparison of forest above‐ground biomass from dynamic global vegetation models with spatially explicit remotely sensed observation‐based estimates

Gaps in our current understanding and quantification of biomass carbon stocks, particularly in tropics, lead to large uncertainty in future projections of the terrestrial carbon balance. We use the recently published GlobBiomass data set of forest above‐ground biomass (AGB) density for the year 2010, obtained from multiple remote sensing and in situ observations at 100 m spatial resolution to evaluate AGB estimated by nine dynamic global vegetation models (DGVMs). The global total forest AGB of the nine DGVMs is 365 ± 66 Pg C, the spread corresponding to the standard deviation between models, compared to 275 Pg C with an uncertainty of ~13.5% from GlobBiomass. Model‐data discrepancy in total forest AGB can be attributed to their discrepancies in the AGB density and/or forest area. While DGVMs represent the global spatial gradients of AGB density reasonably well, they only have modest ability to reproduce the regional spatial gradients of AGB density at scales below 1000 km. The 95th percentile of AGB density (AGB₉₅) in tropics can be considered as the potential maximum of AGB density which can be reached for a given annual precipitation. GlobBiomass data show local deficits of AGB density compared to the AGB₉₅, particularly in transitional and/or wet regions in tropics. We hypothesize that local human disturbances cause more AGB density deficits from GlobBiomass than from DGVMs, which rarely represent human disturbances. We then analyse empirical relationships between AGB density deficits and forest cover changes, population density, burned areas and livestock density. Regression analysis indicated that more than 40% of the spatial variance of AGB density deficits in South America and Africa can be explained; in Southeast Asia, these factors explain only ~25%. This result suggests TRENDY v6 DGVMs tend to underestimate biomass loss from diverse and widespread anthropogenic disturbances, and as a result overestimate turnover time in AGB.