Is a high glycogen content beneficial or detrimental to the ischemic rat heart? A controversy resolved

A high glycogen level may be beneficial to the ischemic heart by providing glycolytic ATP or detrimental by increasing intracellular lactate and protons. To determine the effect of high glycogen on the ischemic myocardium, the glycogen content of Langendorff-perfused rat hearts was either depleted or elevated before 32 minutes of low-flow (0.5 mL/min) ischemia with Krebs-Henseleit buffer with or without 11 mmol/L glucose, followed by 32 minutes of reperfusion with buffer containing 11 mmol/L glucose. 31P nuclear magnetic resonance spectra were acquired sequentially throughout. Further experiments involved early reperfusion or the addition of HOE 694, a Na+-H+ exchange inhibitor, during reperfusion. When glucose was supplied throughout ischemia, no ischemic contracture occurred, and postischemic recovery of contractile function was highest, at 88% of preischemic function. In the absence of glucose, normal-glycogen hearts underwent ischemic contracture at 5 minutes, had an end-ischemic pH of 6.87, and recovered to 54%, whereas in high-glycogen hearts, contracture was delayed to 13 minutes, the end-ischemic pH was 6.61, and functional recovery decreased to 13%. Contracture onset coincided with the decrease in glycolysis, which occurred as glycogen became fully depleted. Functional recovery in the high-glycogen hearts increased to 89% when reperfused before contracture and to 56% when reperfused in the presence of HOE 694. Thus, during brief ischemia in the high-glycogen hearts, ischemic glycogen depletion and contracture were avoided, and the hearts were protected from injury. In contrast, during prolonged ischemia in the high-glycogen hearts, glycogen became fully depleted, and myocardial injury occurred; the injury was exacerbated by the lower ischemia pH in these hearts, leading to increased Na+-H+ exchange during reperfusion. The contradictory findings of past studies concerning the effect of high glycogen on the ischemic myocardium may thus be due to differences in the extent of glycogen depletion during ischemia.     

Effect of in vivo exposure to iodine-131 on the frequency and persistence of micronuclei in human lymphocytes

The validity of the micronucleus test as a biomarker of chromosome damage in dividing mammalian cells is well established. This assay was used to study the response of peripheral lymphocytes of a 34-yr-old male patient following treatment with 131I ablative radiation therapy following a total thyroidectomy. Coincidentally, 8 mo before diagnosis, the patient had provided a blood sample for an in vitro study of micronucleus induction following exposure to graded doses of x-rays. The background frequency in the unexposed culture showed a mean count of 6.0 micronuclei per 1000 binucleated (first division) lymphocytes, while mean values of 18.5, 29.0, 41.0, 61.0 and 75.5 micronuclei/1000 cells were observed following x-ray doses of 5, 10, 15, 20, and 25 cGy, respectively. These data fit a nonthreshold, linear dose-response function (y = 2.78x + 3.71; r = .99). Eight months after the in vitro x-ray study, the subject was diagnosed with thyroid cancer. Surgery was performed, and 5 wk later the patient received 1.78 GBq (48 mCi) of 131I as adjuvant radiation therapy. Blood was drawn 11 d after the radiation treatment and at monthly intervals thereafter to analyze the frequency and persistence of micronuclei. The first posttreatment sample showed 35.5 micronuclei per 1000 binucleate cells. Based on the linear dose-response equation from the earlier study, the sixfold increase in micronucleus frequency suggests a dose to the peripheral blood of approximately 11 cGy. The cytogenetic dose estimate compares to approximately 30 cGy using a new model based on external whole-body counting data. Nine consecutive monthly samples have been analyzed to date. Although the micronucleus count has fluctuated (four- to sixfold above background), the frequency after 8 mo is equivalent to the first posttreatment sample. Data show that radiation-induced cellular lesions persist for months following relatively brief radiation exposure to a medical isotope. Results of this study support the conclusion that the lymphocyte micronucleus test is a rapid, sensitive, and perhaps quantitative biomarker of low-dose (< 25 cGy) radiation exposure.     

Effects of designs of Class 2 preparations on resistance of teeth to fracture

This study was designed to evaluate the effect of isthmus width on the strength of the remaining tooth structure in maxillary first premolars restored with class 2 silver amalgam restorations. One hundred ten sound maxillary first premolars freshly extracted as a part of orthodontic treatment were collected. The teeth were then divided into 10 groups of 11 teeth each. Ideal class 2 mesio-occlusal, disto-occlusal and mesio-occlusodistal cavities with different isthmus widths i.e. one half the intercuspal distance, one third the intercuspal distance and one fourth the intercuspal distance and one fourth the intercuspal distance were prepared. Compressive forces were applied using a Universal Testing Machine and load to the point of fracture was determined. Intact teeth produced the best tooth fracture resistance i.e. 105.4 MPa, followed by preparations with isthmus width of one fourth, one third and one half intercuspal distance in the order.     

Multi-institutional survey of graduates of pediatric anesthesia fellowship: assessment of training and current professional activities

We surveyed all the graduates of four fellowship programs in pediatric anesthesia between 1985 and 1993 to assess their current professional activities, their evaluation of fellowship training, and their opinions on future directions of such training. One-hundred ninety-one (62%) of the graduates responded. Nearly all of the respondents had sought fellowship training for pediatric anesthesia and thought that the training was worthwhile. At the time of the survey, 40% worked in a children's hospital, 72% had university or affiliate positions, and 54% had a practice that was > 50% pediatric. Those with > or = 12 mo fellowship and/or board certification in pediatrics were the most likely to have a pediatric-dedicated practice. Seventy percent of the respondents thought that fellowship training should be for 12 mo, and the proportion of respondents who recommended inclusion of training in pain management and clinical research was greater than the number who had actually received such training. Fifty-eight percent of respondents supported restriction of fellowship positions in the future, but 83% did not support a mandatory 2-yr fellowship with research training. We conclude that fellowships in pediatric anesthesia seem to be successful in providing training that is not only satisfying to the trainees, but that is also followed by active involvement in the care of children and in the training of residents and fellows in anesthesia. Additional information should be gathered to assess the impact of this training on pediatric care, to formulate a standardized curriculum, and to justify support for such training in the future. Implications: We surveyed graduates of four fellowship programs in pediatric anesthesia (1985-1993) to assess current professional activities, fellowship training, and future directions of such training. Fellowships in pediatric anesthesia seem to provide training that is satisfying to trainees and that is followed by active involvement in the care of children.     

Effect of deletion from the carboxyl terminus of the 12 S subunit on activity of transcarboxylase

Transcarboxylase from Propionibacterium shermanii is a biotin-containing enzyme which catalyzes the reversible transfer of a carboxyl group from methylmalonyl-CoA to pyruvate. The central hexameric 12 S subunit of the enzyme associates with six 6 S subunits in the complete enzyme complex. We have constructed a series of cloned genes which encode COOH-terminal truncations of the 12 S subunit. Five of these subunits, which remained soluble following expression in Escherichia coli and were missing from 39 to 97 COOH-terminal amino acids, were purified and compared to the full-length subunit after enzyme complexes were assembled in vitro. All of the truncated subunits were 90% as active in the transcarboxylase reaction as wild type except the reaction containing the shortest complex, TC-12 S (1-507), which had 54% of the wild type activity (TC-12 S-WT). The reduced activity was not due to a lack of CoA ester binding sites or the Km for substrate. However, TC-12 S (1-507) was slower to form than TC-12 S-WT and had more incomplete complexes as judged by high performance liquid chromatography gel filtration profiles and electron microscopy. Isolated TC-12 S (1-507) was 70-80% as active as TC-12 S-WT. We also noted that the truncated form was heat-labile compared to wild type. We conclude that the COOH-terminal region of the 12 S subunit plays a role in assembly and stability of the hexamer and also affects the binding of 6 S subunits to form enzyme complexes. Once complexes do form, the catalytic capacity of TC-12 S (1-507) is almost the same as TC-12 S-WT.     

Sucrose consumption in mice: major influence of two genetic loci affecting peripheral sensory responses

Individual variability in sucrose consumption is prominent in humans and other species. To investigate the genetic contribution to this complex behavior, we conducted behavioral, electrophysiological, and genetic studies, using male progeny of two inbred mouse strains (C57BL/6ByJ [B6] and 129/J [129]) and their F2 hybrids. Two loci on Chromosome (Chr) 4 were responsible for over 50% of the genetic variability in sucrose intake. These loci apparently modulated intake by altering peripheral neural responses to sucrose. One locus affected the response threshold, whereas the other affected the response magnitude. These findings suggest that the majority of difference in sucrose intake between male B6 and 129 mice is due to polymorphisms of two genes that influence receptor or peripheral nervous system activity.     

Potentiation of apoptosis by flavopiridol in mitomycin-C-treated gastric and breast cancer cells

Flavopiridol (L86-8275) is a synthetic flavone currently undergoing Phase I clinical trials. It is active against a series of human cancer cell lines and has been shown to inhibit a broad range of protein kinases, including cyclin-dependent kinases and protein kinase C (PKC). Previous studies have shown that the PKC-specific inhibitor safingol significantly enhances the induction of apoptosis by mitomycin-C (MMC) in gastric cancer cells. Because flavopiridol can potentially inhibit PKC, we elected to determine the extent to which flavopiridol would promote MMC-induced apoptosis in both gastric and breast cancer cells. For these studies, MKN-74 gastric cancer cells and MDA-MB-468 breast cancer cells were exposed to either no drug, 1 microgram/ml MMC alone, 300 nM flavopiridol alone, or a combination of chemotherapy with flavopiridol for 24 h. Sequence specificity was also examined by first exposing cells to MMC for 24 h followed by flavopiridol for 24 h or to the same drugs in the reverse order. Apoptosis was measured by quantitative fluorescence microscopy of nuclear chromatin condensation in cells stained with the dye, bisbenzimide trihydrochloride. Exposure of MKN-74 cells to flavopiridol alone induced apoptosis in 12 +/- 1% of the cells, and exposure to MMC alone induced apoptosis in 10 +/- 1%. However, the combination of flavopiridol and MMC increased the induction of apoptosis to 55 +/- 3% of the cells (P < 0.005 for the drug combination versus flavopiridol alone). Pretreatment with the PKC activator 3-phorbol 12-myristate 13-acetate only partially reversed this effect (43 +/- 1%; P < 0.025). In MDA-MB-468 cells, flavopiridol alone induced apoptosis in 17 +/- 1% of the cells, and MMC alone induced apoptosis in 10 +/- 1% of the cells. The combination of flavopiridol and MMC increased the percentage of MDA-MB-468 cells undergoing apoptosis to 58 +/- 4% (P < 0.005 for the drug combination versus flavopiridol alone). Sequential treatment with MMC followed by flavopiridol induced apoptosis in 63 +/- 2% of the MKN-74 cells (P < 0.05 versus the concomitant drug combination) and in 76 +/- 2% of the MDA-MB-468 cells (P < 0.025 versus the concomitant drug combination), whereas flavopiridol followed by MMC did not increase the induction of apoptosis in either cell line. As determined by the terminal deoxynucleotidyl transferase labeling of the 3' ends of DNA fragments produced in apoptotic cells, the induction of apoptosis with the combination of flavopiridol and MMC occurred to MKN-74 cells in all phases of the cell cycle (i.e., G0-G1, S, and G2-M). These results indicate that flavopiridol potentiates the cytotoxic effect of the chemotherapeutic agent MMC by promoting drug-induced apoptosis in tumor cells. Sequencing studies suggest that MMC followed by flavopiridol or simultaneous treatment is superior to flavopiridol followed by MMC. The enhancement of MMC-induced apoptosis by flavopiridol may be partially PKC dependent and is not associated with one specific region of the cell cycle.