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[CANCER RESEARCH 42, 4936-4942, December 1982] Glucose Metabolism and the Percentage of Glucose Derived from Alanine: Response to Exogenous Glucose Infusion in Tumor-bearing and NonTumor-bearing Rats Jeffrey M. Arbeit, Michael E. Burt, Lawrence V. Rubinstein, Catherine M. Gorschboth, and Murray F. Brennan1 Surgical Metabolism Section. Surgery Branch [J. M. A., M. E. B., C. M. G.. M. F. B.J, and Clinical and Diagnostic National Cancer Institute. NIH. Bethesda. Maryland 20205 ABSTRACT Glucose and alanine metabolism were investigated in nontumor-bearing (NTB) and tumor-bearing (TB) male F344 rats after a 24-hr fast and during the infusion of either 0.9% NaCI solution or glucose at 0.67 or 2.35 mg per 100 g total body weight per min. During 0.9% NaCI solution infusion, the plasma glucose level was higher (98.2 ±4.0 versus 85.8 ±8.1 mg per dl; p < 0.05), the whole-blood láclate level was lower (5.8 ± 0.8 versus 8.3 ± 1.6 mg per dl; p < 0.05), the glucose turnover rate was lower (0.72 ±0.04 versus 0.88 ±0.13 mg per 100 g total body weight per min; p < 0.05), alanine turnover rate and the percentage of glucose derived from alanine was measured by ['"CJalanine in the NTB and compared to the TB animals. In response to glucose infusions, the whole-blood lactate level rose in both groups but remained lower (7.1 ±0.9 versus 10.5 ±2.4 mg per dl at 0.67 mg per 100 g total body weight per min, p < 0.05; 9.1 ±1.1 versus 19.3 ±5.5 mg per dl at 2.35 mg per 100 g total body weight per min, p < 0.05; NTB versus TB) in the NTB than in the TB animals. The endogenous production rate of glucose as measured by [3H]glucose dis played a similar response to exogenous substrate in the NTB and TB animals but required a higher plasma glucose concen tration to effect a similar degree of suppression in the TB group. The alanine turnover rate rose to a similar level, and the percentage of glucose derived from alanine was similarly de pressed in the NTB and TB animals at each glucose infusion rate. INTRODUCTION Nutritional support tailored to the needs of the cancer patient is dependent upon the elucidation of the metabolism of TB2 hosts in the basal state and in response to exogenous substrate administration. Since the work of Cori and Cori (8) and Warburg era/. (48) in the 1920's suggesting the importance of anaerobic glucose metabolism in the substrate economy of tumors in various animal models, a number of studies in cancer patients have centered around carbohydrate metabolism. In the late 1950's, Marks and Bishop (4, 32) demonstrated abnormal glucose tolerance in cancer patients. In the 1960's, Reichard, using [14C]glucose, showed that a small proportion of humans with cancer had abnormally high glucose turnover rates (39). In the 1970's, both Holroyde e? al. (19) and Waterhouse ef al. ' To whom requests for reprints should be addressed, at Memorial Sloan Kettering Cancer Center, 1275 York Avenue, New York, N. Y. 10021. 2 The abbreviations used are: TB, tumor-bearing; NTB, non-tumor-bearing; TBW. total body weight; R0. turnover rate. Received March 4, 1981; accepted August 16, 1982. 4936 Trials Section, Biometry Branch [L. V. R.]. (51) commented, respectively, on altered glucose metabolism and increased gluconeogenesis from alanine in cancer pa tients. However, studies in humans showing significant altera tions in carbohydrate metabolism induced by the TB state are plagued by heterogeneous patient populations, significant an tecedent weight loss, and lack of appropriate controls (4, 19, 20, 28, 32,38, 39,49, 51). An animal model was chosen in this experiment which has been extensively characterized in various studies from this laboratory (2, 5, 6, 14, 29, 36, 40). The unrestrained rat with long-term arterial and venous cannulae (5) permits precise quantification of food intake, nitrogen excretion, and tumor burden in a homogeneous group with appropriate controls. The purpose of this study was not only to see whether differences existed in glucose metabolism and gluconeogenesis from ala nine during the basal state but also to see if aberrancies in glucose metabolism, if present, persisted in the face of constant exogenous substrate administration. Since gluconeogenesis is known to be suppressed in human controls by glucose admin istration (25), the percentage of glucose derived from alanine was also examined to see if it was equivalently suppressed by exogenous substrate in NTB and TB animals. Alanine was chosen because of its participation in the glucose-alanine cycle (11), because of its role as the predominant amino acid re leased from muscle during acute starvation (37), and because it is one of the most important glycogenic amino acids taken up by the liver in the postabsorptive state (41). MATERIALS AND METHODS Animals. Male F344 rats were obtained from Charles River Breed ing Laboratories (Wilmington, Mass.) and maintained on NIH-5018 rat chow (Ralston-Purina, St. Louis, Mo.) and tap water ad libitum. Preparation of Animals. Rats weighing 230 to 250 g were anesthe tized with sodium pentobarbital, 50 mg/kg ¡.p.;using aseptic tech nique, the left carotid and right external jugular veins were each cannulated, and the animals were placed in individual plastic metabolic cages (Maryland Plastics, New York, N. Y.) (5). Five to 7 days postcatheter implantation, those animals that were to be TB were inoculated s.c. in the right flank with a suspension of 1 x 106 viable (by trypan blue exclusion) cells of a methylcholanthrene-induced sarcoma (31 ). When both NTB and TB animals had returned to their preoperative weight and the TB animals had tumors 2 to 3 cm in diameter, 2-day food intake was quantified. Both groups were then fasted for 24 hr with water given ad libitum while urine was collected for total nitrogen determination by the micro-Kjeldahl method (30). An arterial blood sample was obtained, and then an i.v. continuous infusion (Harvard Apparatus, Dover, Mass.) of either 0.9% NaCI solution or glucose at 0.67 or 2.35 mg per 100 g TBW per min was begun (0.67 and 2.35 mg per 100 g TBW per min represent 0.93 and 3.2 times, respectively, the glucose turnover rate of NTB animals). Six hr later, 10 /iCi D-[33H]glucose(18.1 Ci/mmol)and 5juCiL-{U-"lC]alanine(172 mCi/mmol; CANCER RESEARCH Downloaded from cancerres.aacrjournals.org on June 17, 2017. © 1982 American Association for Cancer Research. VOL. 42 Glucose Metabolism and Gluconeogenesis [l4C]alanine New England Nuclear, Boston, Mass.) were injected, and arterial blood samples were obtained at 1, 2, 5, 10, 20, 30, 45, 60, and 90 min after tracer administration. The animals were sacrificed by air embolization, and the TBWs, carcass weights, and tumor weights were determined. Analysis of Intraarterial Substrate Levels. Samples of arterial blood, 220 to 420 n\, were placed in chilled tubes containing 22 to 24 fil sodium heparin (1000 units/ml). Whole-blood aliquots (200 /il) of the base line and 5- and 60-min blood samples were deproteinized, and the supernatant was analyzed enzymatically (17). Blood samples were centrifuged, and the plasma was separated. An aliquot of plasma was diluted with water, and glucose was measured by a glucose oxidase method (47) on an autoanalyzer (Technicon Instruments Corp., Tarrytown, N. Y.). The remaining plasma underwent membrane filter centrifugation (Amicon Centriflo Cones, CF50A), and alanine concen tration was determined on an aliquot of filtrate by a single column technique (24) on a Beckman 121 MB amino acid analyzer (Beckman Instruments, Palo Alto, Calif.). Separation of Radioactive Metabolites. The pH of the filtrate was adjusted to 2.0 to 2.1 and placed on a cation-exchange column (Kontes, Vineland, N. J.) with a 0.5-ml bed volume (18) (AG 50-X8, 200 to 400 mesh; Bio-Rad Laboratories, Richmond, Calif.). The column was washed with 2.5 ml of water, and the glucose-containing eluant. Fraction I, was collected. Amino acids, Fraction II, were removed from the column by adding 1.5 ml of 2 N NH4OH followed by 1 ml of water. Fraction I was bought to pH 7 to 8 and placed on an anion-exchange column with a 1.0-ml bed volume (AGI-X8, 200 to 400 mesh, formate form; Bio-Rad), followed by 1 ml of water, and the eluant, Fraction la, during Glucose Infusion specific activity (dpm/mmol). Calculations. The glucose R . and alanine R were calculated using noncompartmental steady state analysis (22, 23). The glucose mass, half-life, and space were calculated from the linear regression of the In specific activity-time curve of [3H]glucose (42). The glucose clearance (7), the endogenous production rate (total [3H]glucose fl0 - glucose infusion rate), and the percentage of glucose derived from alanine 100 / ([l/-'"C]glucose 2 / ([U-'"C]alanine specific activity)df specific activity)df where specific activity is in dpm/mmol] (9, 10, 45, 47), were also calculated. Statistical Analysis. Data are expressed as mean ±S.D. Statistical significance was determined by Student's 2-tailed t test for unpaired data and by covariance analysis. Since the group sample sizes are small and there is no assurance that the specific activity values are normally distributed, the significance levels were confirmed by the Fisher randomization test for unpaired data (46). The significance levels obtained by means of the 2 tests were essentially identical. RESULTS Food Intake, Nitrogen Balance, and Body Weight. The food intake did not differ between the NTB and TB animals (Table 1). The urinary nitrogen balance was significantly less negative in the TB than in the NTB animals (Table 1). The TBW was significantly increased but the carcass weight was signifi cantly decreased in the TB versus the NTB animals (Table 1). Substrate Levels. During 0.9% NaCI solution infusion, the whole-blood lactate level was significantly increased in the TB animals (Table 2). At each glucose infusion rate, the wholeblood lactate incrementally rose in both groups of animals with a statistically greater level in the TB animals (Table 2). The plasma glucose concentration remained stable in each group of animals during infusion (Chart 1). When 0.9% NaCI solution was collected. Both Fraction la and Fraction II were evaporated to dryness at room temperature (Savant Instruments, Inc., Hicksville, N. Y.). The Fraction la residue was reconstituted with water, and aliquots were taken for glucose concentration measurement and scin tillation counting (Beckman LS 355) to determine the specific activity (dpm/mmol) of [3H]glucose and [14C]glucose. The Fraction II residues was reconstituted with 0.15 N lithium citrate buffer, pH 2.0 (Beckman) (24) and placed on an amino acid analyzer modified as a fraction collector (Beckman 121M). Aliquots of the alanine fraction were placed on a Beckman 121MB amino acid analyzer and counted to determine Table 1 Body weights, food intake, and nitrogen balance of TBW8 occu GroupNTB (15)" (g)216.4 (g)216.4 (g)0 wt wt intake(g)30.2 food (mg)-212.8 N balance tumor0 pied by ±9.0e ±11.9 (15) ± 9.0(15) (15) (15) ±3.1 (15) 208.3 ±10.3 (14)Tumor3 17.3 ±8.3(14)% TB(14)TBW" 225.7 ±7.0 (14)Carcass8 7.7 ±3.7(14)48-hr628.1 ±4.5(14)24-hrc -194.1 ±21.9 (14) Determined at the end of the experiment after the 24-hr fast and the 6-hr infusion. '' Measured after animals had regained their postoperative weight loss and just prior to the 24-hr fast. c Measured during the 24-hr fast prior to the start of the 6-hr infusion. 0 Numbers in parentheses, number of animate. " Mean ±S.D. 'p<0.05. Table 2 Whole-blood lactate and plasma glucose concentrations during infusions The results are expressed as the mean of mean values of multiple time points for each rat. Lactate (mg/dl) Infusion NTB (6)"7.1 ±0.88 solutionGlucose 0.9% NaCI (5)9.1 ±0.9" mg/100gTBW/min)Glucose (0.67 mg/100gTBW/min)5.8 (2.35 ±1.1df*(4)8.3 Glucose (mg/dl) TB (5)10.5±2.4C ±1.6C (4)19.3 ±5.5c'"-e (5)98.2 NTB (6)134.3 ±4.0 (5)160.2 ±4.1a TB 8.1C124.7 ± 5.9s170.8 ± ±5.4*' (4)85.8 (5) ±12.3*(5)"(4)9 8 Mean ±S.D. Numbers in parentheses, number of animals. c p < 0.05 comparison of TB with NTB during each infusion. p < 0.05 comparison of values during each glucose infusion with values during 0.9% NaCI solution infusion within each group. * p < 0.05 comparison of values during 2.35-mg per 100 g TBW per min glucose infusion with values during 0.67mg per 100 g TBW per min glucose infusion within each group. DECEMBER 1982 4937 Downloaded from cancerres.aacrjournals.org on June 17, 2017. © 1982 American Association for Cancer Research. J. M. Arbeit et al. Plasma Glucose Concentration (mg/dl) 200 Glucose 2.35mg/ 100g TBW/ min 10.00 100 7.60 -360 0 20 40 TIME (min) 60 80 100 - 5.00 Chart 1. Plasma glucose concentration in response to 6-hr infusions of either 0.9% NaCI solution or glucose at 0.67 or 2.35 mg per 100 g TBW per min. The data are the mean for each group at each time point. Bars, S.D. The curves should be broken between —¿360 and O min. was infused, the plasma glucose concentration was signifi cantly lower in the TB than in the NTB animals (Table 2). Infusion of glucose at 0.67 mg per 100 g TBW per min failed to significantly raise the plasma glucose concentration to an equivalent level in the TB animals (Table 2). At the glucose infusion rate of 2.35 mg per 100 g TBW per min, there was no difference in the elevated plasma glucose levels in both groups of animals (Table 2). There were no statistical differences between the plasma alanine levels during 0.9% NaCI infusion in the TB and NTB animals (2). In response to glucose infusions, there was a progressive elevation in the plasma alanine levels in both TB and NTB animals with no statistical difference between the groups (2). [3H]Glucose Kinetics. The specific activity-time curves of the disappearance of [3H]glucose during 0.9% NaCI solution and incremental glucose infusion are displayed in Chart 2. Noncompartmental analysis of these curves (22, 23) shows that during 0.9% NaCI solution infusion there was a statistically significant increase in the glucose turnover in the clearance rate in the TB compared to the NTB animals (Table 3). Linear regression analysis of the In specific activity [3H]glucose-time curves demonstrates a significantly more rapid glucose half-life and a similar mass in the TB compared to the NTB animals (Table 3). In response to glucose infused at 0.67 mg per 100 g TBW per min, there was a decrease in the half-life and increases in the masses, turnover rates, and clearance rates in both groups of animals, with statistically significant differences in the half-life and clearance rates and a borderline significant difference (p = 0.06) in the turnover rate in the TB compared to the NTB animals. When glucose was infused at 2.35 mg per 100 g TBW per min, there was an additional decrease in the half-life and an increase in the mass, turnover rate, and clear ance rate in both groups, but there were no statistical differ ences between the TB and NTB animals (Table 3). Endogenous Glucose Production. The endogenous glucose production rate, as measured by [3H]glucose kinetics, was suppressed significantly in the NTB animals infused with glu cose at 0.67 mg per 100 g TBW per min (Table 3). At this infusion rate, the TB animals failed to suppress their endoge- 4938 2.50 10 20 30 40 50 60 70 90 Chart 2. Specific activity (S. A Mime curves for [3H]glucose during each in fusion, normalized for dose (dpm [3H)glucose injected) and TBW of each animal. The data are presented as the mean values at each time point. The turnover rates generated by noncompartmental analysis show a statistically significant increase with increasing glucose infusion within each group and a significant difference for the NTB (A) versus TB (O) animals infused with 0.9% NaCI solution. The top, middle, and bottom sets of curves are from animals infused with 0.9% NaCI solution and glucose at 0.67 and 2.35 mg per 100 g TBW per min, respectively. nous production and had a statistically significant elevated rate compared to the NTB animals (Table 3). During glucose infu sion at 2.35 mg per 100 g TBW per min, the endogenous glucose production fell to a similar level in both NTB and TB animals (Table 3). Linear regression and covariance analysis of the plasma glucose concentration versus the endogenous production rate for each individual animal (Chart 3) (NTB, y = -0.00555* + 1.302; TB, y = -0.00360x + 1.263) shows that, despite a high degree of correlation in the NTB animals, r = —¿0.803 (p < 0.05), and a low correlation in the TB animals, r = —¿0.364 (p > 0.05), the slopes of these regression lines were similar (p = 0.56). However, the adjusted cell means were significantly greater in the TB compared to the NTB animals (p < 0.05). [14C]Alanine Turnover Rate. The specific activity-time curves for [14C]alanine are depicted in Chart 4, according to group and infúsate. Noncompartmental analysis reveals an increase in the alanine turnover rate during incremental glucose infusion with similar results in the NTB and TB animals infused either with 0.9% NaCI solution or glucose. Percentage of Glucose Derived from Alanine. The specific activity-time curves for the appearance of ['"CJalanine are shown in Chart 5. The percentage of glucose derived from CANCER RESEARCH Downloaded from cancerres.aacrjournals.org on June 17, 2017. © 1982 American Association for Cancer Research. _ 80 TIME (min) VOL. 42 Glucose Metabolism and Gluconeogenesis during Glucose Infusion Table 3 Glucose kinetics during infusions The [3H]glucose turnover rate is calculated as: dpm 3H injected 100 x TBW J ([3H]glucose specific activity) dt The glucose half-life, mass, and space are calculated from the linear regression of the In specific activity-time curve of [3H]glucose. The glucose clearance rate is calculated as 100 x The endogenous glucose production glucose turnover rate plasma glucose concentration rate is defined as total | 'H]glucose turnover rate - glucose infusion rate. rate (mg/100 g TBW/min)0.72 (mg/ 100 (min)24.4 TBW)25.5 g ±2.4b (6)" NaCI solution ± 2.4 17.8 ±2.1° 0.88 TB(5) 22.5 ± 3.4 19.2 ±1.6d 35.5 ± 3.0d NTB (5) 0.67mg/100gTBW/min 1.29 16.7 ±1.2C 1.68 TB(4) 40.2 ± 8.8 12.9 ±1.9d'e 50.0 ± 5.3d'8 2.69 2.35 mg/100 g TBW/minGroupNTBNTB (4) 54.5 ±10.2oTurnover 2.76 TB(5)Half-life 13.6 ±0.8â„¢Mass Infusion0.9% rate (ml/ 100 g TBW/ min)0.73 glu cose production rate (mg/ 100 g TBW/min)0.72 ±0.04 ±0.13C ±0.02 ±0.04 1.03 ±0.12° 0.88 ±0.13 0.64 ±0.07d 0.96 ±0.1"(6) ±0.10 ±0.38d 1.34 ±0.27°'d 1.05 ±0.38C ±0.20tÃ--e 1.70 ±0.14"'" 0.35 ±0.170-8 ±0.43d'eClearance 1.62 ±0.24dEndogenous 0.53 ±0.42 Numbers in parentheses, number of animals. " Mean ±S.D. cp< 0.05 comparison of TB with NTB during each infusion. " p < 0.05 comparison of values during each glucose in fusion with values during 0.9% NaCI solution infusion within each group. 8 p < 0.05 comparison of values during 2.35-mg per 100 g TBW per min glucose infusion with values during 0.67-mg per 100 g TBW per min glucose infusion within each group. 2.0 o _ O e £E U) ? >e LO lo 75 100 125 150 175 200 PLASMA GLUCOSE CONCENTRATION (mg/dl) Chart 3. Graphic analysis of the response of the endogenous glucose pro duction rate to prevailing plasma glucose concentrations. The data represent the plasma glucose concentration of each animal and the resultant endogenous glucose production rate: NTB (A), y = -0.00555x + 1.302; TB (O), y -0.0036X + 1.263. Covariance analysis showed equality of slopes but a statistically significant higher group mean in the TB compared to the NTB animals. alanine decreases progressively in response to incremental glucose infusion to a similar degree in both NTB and TB animals (Table 4). DISCUSSION In this study, abnormalities of basal glucose metabolism were demonstrated in TB compared to NTB animals when the tumor comprised 7.7 ± 3.7% of the TBW, before the onset of de creased food intake, increased nitrogen excretion, and pro gressive TBW loss. These differences in glucose metabolism persisted during exogenous glucose administration. Basal al anine metabolism and the percentage of glucose derived from DECEMBER 1982 alanine were not significantly different in the TB animals com pared to the NTB animals, and this lack of significant difference between the groups also persisted during glucose infusion. Previous work in our laboratory demonstrating abnormal glucose metabolism and gluconeogenesis from alanine in TB compared to NTB rats was based on animals in their fourthday post-superior vena cava cannulation, with tumor burdens of 15 to 20% of the TBW and significantly lower carcass weights (29). With the use of a single superior vena cava catheter for simultaneous tracer injection and substrate infu sion, it is difficult to obtain blood samples at rapid intervals and to ensure that they are not contaminated. However, to accu rately define glucose and alanine kinetics, it is important to obtain early, multiple time points; thus, in our previous studies, alanine kinetics was calculated indirectly by assumptions as to glucose derived from alanine entering the glucose decay curve (29). In addition, rats at the fourth day postcannulation may still be recovering from the operative procedure (5), and pre vious reports in the literature have shown how carbohydrate metabolism can be altered by trauma (1, 26, 52). The transplantable, methylcholanthrene-induced sarcoma used in this study is first palpable at Day 14 after s.c. flank injection of 1 x 106 viable cells and kills the animal when it occupies 30 to 35% of the TBW, 35 to 40 days postinoculation (36). Thus, the animals used in this experiment were studied in the first quarter of their disease process, and their tumor burdens were comparable to those of humans with large retroperitoneal sarcomas or extensive leukemic-lymphomatous bone and extramedullary involvement (12, 27). The equivalent food intake of the TB and NTB animals in this study eliminated prior starvation as a cause for the altered glucose metabolism in the tumor bearers (35). The significantly less negative nitrogen balance in the TB compared to the NTB animals was previously documented by Mider in rats with and without Walker 256 carcinomas (34). This "nitrogen trap" (34) phenomenon has been reported by Waterhouse ef a/. (50) in 4939 Downloaded from cancerres.aacrjournals.org on June 17, 2017. © 1982 American Association for Cancer Research. J. M. Arbeit et al. 90 Chart 4. Specific activity (S.A. Mime curves for [14C]alanine during each in fusion normalized similar to curves in Chart 2. The data are the mean values at each time point. The turnover rates generated by noncompartmental analysis show a statistically significant increase within each group (Table 4) with no patients with leukemias and lymphomas. In the presence of this relative nitrogen avidity and in spite of their significantly greater TBW's, the TB animals still had depressed carcass weights compared to the NTB animals. Mider demonstrated this same result; the carcasses of his TB rats contained less lipid (33) and nitrogen (34) than did the NTB controls, and the "lost" nitrogen was found sequestered in the tumor (34). The stable plasma glucose levels during infusion of either 0.9% NaCI solution or exogenous glucose demonstrated that both groups of animals were in the steady state. The statistically significantly decreased basal plasma glucose and increased whole-blood lactate levels in the TB animals have been shown in previous work from this laboratory (6) and others (43). Studies involving in vivo tumor preparations growing in ap pendages (8, 48) or on isolated vascular pedicles (16) have documented high rates of glucose consumption and lactate production by tumors. In response to glucose infused at 0.67 mg per 100 g TBW per min, there was a statistically significantly higher wholeblood lactate level in the TB animals despite a significantly lower plasma glucose concentration. With a similar degree of hyperglycemia during glucose infusion at 2.35 mg per 100 g TBW per min, the lactate level was twice as high in the TB as in the NTB animals. One possible explanation for this greater rise in lactate concentration during glucose infusion in the TB compared to the NTB animals was demonstrated in isolated in vivo tumor preparations in which the capacity of the tumor for glucose uptake was unaffected by hyperglycemia and the effluent lactate concentration was increased (16). Clinically, 4940 difference between the NTB (A) and TB (O) animals. Top, middle, and bottom sets of curves are from animals infused with 0.9% NaCI solution and glucose at 0.67 and 2.35 mg per 100 g TBW per min, respectively. there have been reports of cancer patients receiving high glucose infusion rates in the form of total parenteral nutrition who were either in moderate lacticacidemia (20) or frank lactic acidosis (15). The lack of significant difference in the plasma alanine con centration during 0.9% NaCI solution or glucose infusion (2) implies that glucose shunting via the alanine cycle (11 ) is not increased in the TB compared to the NTB animals and stands in contrast to the above-noted differential lactate response. The close agreement of the glucose kinetic data in the NTB animals from this experiment with that reported from this lab oratory (6) and others (3, 9, 21, 22, 44) strengthens the differences demonstrated between the NTB and TB animals in this study. The significantly elevated glucose turnover and clearance rates of the TB animals infused with 0.9% NaCI solution are nearly identical to the data reported by Burt using the same noncachectic animal-tumor system but with i.p. tracer admin istration (6). This increased glucose turnover rate shown in TB animals is distinct from starvation which results in a decrease in the turnover rate (13). During glucose infusion at 0.67 mg per 100 g TBW per min, the significantly increased glucose clearance rate in the TB as compared to the NTB animals demonstrates that the aberrant glucose kinetics of the TB state persists during moderate rates of exogenous substrate administration. When glucose was infused at 2.35 mg per 100 g TBW per min, the kinetic differences between the TB and NTB groups disappeared, suggesting either that the TB animal was saturated with subCANCER RESEARCH Downloaded from cancerres.aacrjournals.org on June 17, 2017. © 1982 American Association for Cancer Research. VOL. 42 Glucose Metabolism and Gluconeogenesis during Glucose Infusion strate or, given the increased láclate level, that there was increased conversion of the infused glucose into this substrate in the TB animal. In response to incremental glucose infusion and the resultant increase in the plasma glucose, the endogenous glucose pro duction rate clearly was suppressed in the NTB animals. The TB animals behaved in a more complex manner, having a persistently elevated endogenous production rate compared to the NTB group when glucose was infused at 0.67 mg per 100 g TBW per min which then fell to a similar level when glucose was infused at 2.35 mg per 100 g TBW per 100. The data from the covariance analysis of Chart 3 reveal that the response of the endogenous glucose production rate to the prevailing plasma glucose concentration was basically similar in the NTB and TB animals but that the TB animals required a higher plasma glucose concentration to effect a similar reduction in this rate. In contrast to the glucose kinetic data, there were no signifi cant differences in the alanine turnover rates during 0.9% NaCI solution or glucose infusion between the TB and NTB animals. Coupled with the lack of significant differences in the percent age of glucose derived from alanine, these data suggest that abnormalities in glucose kinetics found in TB animals during 0.9% NaCI solution or exogenous glucose administration are not due to an increased percentage of glucose derived from alanine. However, if data from conversion of [14C]alanine to [14C]glucose are expressed as a percentage of 3-3H-derived Chart 5. Appearance of the specific activity (S.A 1 of |"C|glucoso, derived from the injected [U-"C]alanine, over time during each infusion. Normalized similar to data in Chart 2. Data are the mean values at each time point. The curves are arranged with respect to infusion similar to curves in Charts 2 and 4. A. NTB; O, TB. Table 4 Percentage of glucose derived from alanine and the alanine turnover rate The percentage of glucose from alanine is calculated as: 100 J (["CJglucose specific activity)« 2 / <['4C]alanine specific activity) dt The alanine turnover rate is: dose (dpm ['"CJalanine injected) ' TBW / (( ' 4Cplanine specific activity) dtInfusion0.9% solution0.67mg/100g NaCI (5f-b TB(5)NTB TBW/ (4)b TB(4)NTB min2.35 of glucose alanine4.00 from ±1.06° turnover rate (nmol/ 100 g min)0.93 TBW 4.14 0.942.37 ± ±0.24 0.88 0.251.41 ± ±0.29d 0.43d0.84 2.55 ± ±0.09d 0.33d2.20 1.57 ± ±0.28dl" ±0.21d'8 (4) mg/100 g TBW/ 0.96 ±0.20d'eAlanine 2.51 ±0.54d-e TB(5)% minGroupNTB "' Numbers in parentheses, number of animals. In a animals, alanine kinetics was not obtainable. c Mean ±S.D. d p < 0.05 comparison of values during each glucose infusion with values during 0.9% NaCI solution infusion within each group. " p < 0.05 comparison of values during 2.35-mg per 100 g per TBW per min glucose infusion with values during 0.67-mg per 100 g TBW per min glucose infusion within each group. DECEMBER 1982 glucose turnover, an apparent increase in the amount of glu cose derived from alanine (3.2 to 4.8%) occurs in TB animals at high glucose loads. This calculation we believe to be unjus tified, inasmuch as the sensitivity to error is high and the [14C]glucose specific activity is subject to recycling. Any com ment on increased cycling of glucose to alanine must await more definitive experiments. In conclusion, the results of this experiment define a unique state of glucose metabolism in the TB animals not explicable in terms of antecedent starvation or increased stress or injury. The increased glucose turnover of the TB animal during 0.9% NaCI solution infusion is a different response to the progressive decrease seen in starvation. The significantly greater rise in whole blood lactate during glucose infusion in this group implies continual glucose uptake and conversion to lactate by the tumor in the face of hyperglycemia. The increased clearance and endogenous glucose produc tion rates during glucose infusion at 0.67 mg per 100 g TBW per min demonstrate that glucose kinetic abnormalities persist during moderate rates of exogenous substrate administration. The lack of difference in the percentage of glucose derived from alanine and the alanine turnover rates during 0.9% NaCI solution infusion, coupled with the relative nitrogen avidity during the 24-hour fast, is strong support that gluconeogenesis from alanine is not increased in TB animals when antecedent food intake is equivalent to that of NTB animals. The compa rable suppression of the percentage of glucose derived from alanine by glucose infusion shows that TB animals are not like septic postoperative patients (25) and that glucose derived from alanine probably plays a minor role in the substrate economy of the TB host. REFERENCES 1. Allsop. J. R.. Wolfe, R. R., and Burke, J. F. Glucose kinetics and respon- 4941 Downloaded from cancerres.aacrjournals.org on June 17, 2017. © 1982 American Association for Cancer Research. J. M. Arbeit et al. siveness to insulin in the rat injured by burn. Surg. Gynecol. Obstet., 147: 565-573. 1978. 2. Arbeit, J. M., Gorschboth, C. M., and Brennan, M. F. Basal amino acid metabolism and its response to glucose infusion in rats with and without a sarcoma. Clin. Res., 29: 432A, 1981. 3. Baker, N., Shipley, R. A., Clark, R. E., and Incefy, G. E. 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