Diglycine
absorption
in streptozotocin
diabetic
HAROLD P. SCHEDL, JANICE WENGER, AND SIAMAK A. ADIBI Gastruenterulugy Research Laboratories, Department of Medicine, University Iuwa College of Medicine, Iuwa City, Iuwa 52242; and Gastrointestinal and Nutritional Unit, Muntefiure Huspitul, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15213
SCHEDL, HAROLD P., JANICE WENCER, AND SIAMAK A. Anxsr. Diglycine absorption in streptozotocin diabetic rat. Am. J. Physiol. 235(5): E457-E460, 1978 or Am. J. Physiol.: Endocrinol. Metab. Gastrointest. Physiol. 4(5): E457-E460, 1978. - Because experimental diabetes in the rat is associated with changes in function of neutral and basic amino acid transport systems, we studied the effects of streptozotocininduced diabetes on dipeptide transport. Absorption was investigated by intraluminal perfusion of the entire small intestine with 20 mM diglycine in pair-fed control and diabetic rats early (3-4 days) and late (15-17 days) after induction of diabetes. In contrast to controls, which grew in body and mucosal weight, diabetics lost body weight but showed mucosal growth. Specific absorption (mmol/g dry wt mucosa per h) was the same in controls and diabetics both early (1.69 vs. 1.55) and late (1.23 vs. 1.28), but was lower in the late than in the early set. Total intestinal absorption (specific absorption x mucosal weight) was the same in early and late controls, but was greater late than early in diabetics because of increased mucosal growth. Pair feeding of controls and diabetics requires restriction of food intake of early controls because food intake of diabetics is low at the early time interval. When controls restricted to the intake of diabetics were compared with ad libitum fed controls, specific absorption was increased in the restricted group. Thus, the greater specific absorption in the early, compared to the late sets of controls and diabetics, is the result of decreased food intake in the early set, intestinal tion
adaptation;
intestinal
growth;
amino
acid absorp-
MAJORITY of amino acid residues of dietary proteins enter the intestinal mucosal cells as small oligopeptides (3, 16). This entry appears to be mediated by a carrier system that is separate from those for free amino acids (3, 16). Prior studies in the rat have shown that alteration in function of the peptide carrier system in response to changes in diet differs from that of the neutral amino acid carrier system (14, 15). In addition to its effect on nutrition, experimental diabetes also has profound effects on mucosal transport function in the rat: absorption of amino acids transported by the neutral and basic amino acid transport systems is increased when studied in vitro (20) and in vivo (12). In the absence of any information on effects of diabetes on peptide transport, we investigated this problem in the present experiments. We chose diglycine for the present studies because its intestinal transport has been THE
0363-6100/78/0000-0000$01.25
Copyright
0 1978 the American
Physiological
rat
of
extensively studied in man and animals (2, 6, 17, 19, 21). Furthermore, diglycine appears to share a common carrier mechanism with other dipeptides and tripeptides (1, 5). MATERIALS
AND
METHODS
Animals. We used male albino rats of the Simonsen strain (Simonsen Laboratories, Gilroy, CA) in the HO200 g weight range (Table 1). Animals were divided into two groups. Rats to be made diabetic were given a single intraperitoneal injection of 100 mg/kg of streptozotocin (supplied by the Upjohn Co., Kalamazoo, MI) freshly dissolved in citrate buffer (pH 4.5). Controls received the buffer only. Control and diabetic groups were pair-fed equal amounts of commercial rat chow (Teklad Mills, Madison, WI). Pair feeding was achieved as follows: a) because food intake of diabetics is decreased for several days after streptozotocin injection, food intake of controls was restricted to that of diabetics initially; b) subsequently, because diabetics developed hyperphagia about 5 days after injection, food intake of diabetics was restricted to that of controls. Food intake by pair-fed controls and diabetics increased from 7 to 22 g/day during the period prior to and during early studies and increased to a mean level of 26 g/day during the several days preceding late studies. Body weight, urine volume, and urine glucose concentrations (TesTape, Lilly) were measured daily. Diabetic animals showed urine glucose concentrations in excess of 2 g/ 100 ml. Mean (*SE) serum glucose concentrations (mg/ 100 ml) in aortic blood from these nonfasted animals were 117 -+ 17 and 195 -t- 20 for early and late control groups, respectively. For diabetics, mean (+SE) serum glucose was 425 -+ 18 in early diabetics and 530 t 23 for late diabetics. Pair feeding of controls with early diabetics requires restriction of food intake of controls to approximately half that of the ad libitum fed controls. Subsequently, for late studies, controls were fed ad libitum. To determine the effect of dietary restriction on diglycine absorption, a separate group of control rats was divided into an ad libitum fed group and a group fed at half the ad libitum intake. Perfusion procedure. For control and diabetic animals, absorption was measured “early” (3-4 days) and “late” (15-17 days) after injection. Early studies defined transport behavior of diglycine before mucosal growth Society
Downloaded from www.physiology.org/journal/ajpendo at Washington Univ (128.252.067.066) on February 13, 2019.
E457
SCHEDL,
E458 TABLE
1. Groups
studied
intestine Weight,
g Dry Wt Mucosa,
Group
Initial
g
was calculated
WENGER,
AND
ADIBI
as follows
corrected mucosal dry wt
Final
= (measured mucosal dry wt)
Controls (3-5 days)” Ad libitum fed Restricted
203 * 3 202 k 3
225 k 7” 183 IL 3cJ1
0.96 k 0.04 0.62 + 0.02”
Early (3-4 days)” Control Diabetic
191 + 2 186 -t 1
186 k 3 156 k 2’=
0.62 -c 0.02 0.65 +_ 0.02
Late (15-17 Control Diabetic
193 k 3 192 k 2
273 k 3”+’ 145 + 3C+“,f
0.93 k 0.03f 1.01 f 0.06”
days)“’
Values are means * SE. ii Time of absorption study after food l) Time of absorption study in was reduced for restricted controls. c Initial differs from final, P < relation to induction of diabetes. 0.05. d Restricted group differs from ad libitum fed group, P < 0.05. e Diabetic group differs from control group, P < 0.05. ’ Group at late period differs from both groups at early period, P < o-05-
in diabetics was increased in comparison with controls, Late studies were to define differences in absorption that appeared after greater mucosal growth was established in diabetics. Ad libitum and restricted control animals were studied after 3-5 days of the feeding regimen Rats were anesthetized by intraperitoneal Nembutal (50 mg/kg). The abdomen was opened by a midline incision, and the common bile duct was ligated. The entire small intestine was perfused as a single unit by inserting cannulas at the pylorus (inlet) and terminal ileum (outlet). The perfusion solution contained 20 mM diglycine (Grand Island Biological Co., Grand Island, NY), and 150 mM NaCl and was perfused at a constant rate (2.5 or 2.0 mllmin) using a peristaltic pump (Cassette Pump, Manostat, New York City). Preliminary studies showed that the diglycine absorption rate is linear for up to a 20 mM infusion concentration. Radioactively labeled polyethylene glycol ([14C]PEG, 10 &i/liter, New England Nuclear, Boston, MA) was used as nonabsorbable indicator. After a 30min initial wash, three lo-min samples were collected in tubes containing sulfosalicylic acid (3.5 mg/ml of sample collected) to inactivate intraluminal peptidase activity. Immediately after perfusion, blood was obtained from the bifurcation of the descending aorta for serum glucose determinations (25) of control and diabetic rats. Intestinal measurements. After blood was obtained, the intestine was stripped from the mesentery. Contents were expressed, and wet weight of the entire intestine was determined. The intestine was then slit longitudinally with a scissors and opened, mucosal side up, onto a glass plate. The mucosa was then scraped from underlying tissue with the edge of a microscope slide. Wet weights of both the mucosal scrapings and underlying tissue were determined, and the scrapings were dried in a vacuum oven at 100°C for 24 h to obtain dry weights. To correct for losses of mucosa incurred in scraping and tranferring to tared containers for weighing and drying, dry weight of mucosa for the entire small
wet wt of entire intestine mucosal wet wt + underlying tissue wet wt > This is a small correction because tissue recovery averaged 95% and ranged from 92 to 96%. AnaZyses. The test solution and samples collected were analyzed for amino acid and peptide concentration using an amino acid analyzer (Beckman model 12OC, Beckman Instruments, Fullerton, CA). [14C]PEG was measured by liquid scintillation counting (Beckman LS-250). Diglycine absorption was calculated as follows total diglycine absorption, mmollh
= [Ci - Cf (IND)] x R where Ci is initial peptide concentration in the test solution and Cf is final peptide concentration after perfusion (both in mmollml); IND is the ratio of initialto-final [l$C]PEG concentrations; R is the rate of perfusion in ml/h. specific diglycine absorption, mmol/h per g dry wt mucosa = (total absorption)/(dry
wt mucosa, g)
net water movement, % = (1 - IND) x 100 Positive values indicate net absorption. luminal appearance rate of glycine, pmol/h = [(C, x IND) - Ci] X R where Cf and Ci are final and initial glycine concentrations (pmol/ml) in the perfused solution, and the other symbols are the same as for total diglycine absorption. Data are presented as mean values t 1 SE for groups of six to nine animals. Tukey’s multiple comparison test (13) and the Student t test (9) were used for statistical analysis. RESULTS
Table 1 shows initial and final body weights and mucosal dry weight for groups studied. At the top are data from a group of normal rats divided into an ad libitum fed group and a group fed half of ad libitum intake (restricted). Shown below are data for control and diabetic groups. Ad libitum and restricted groups were studied after 3-5 days of the feeding regimen. Control and diabetic groups were studied early (3-4 days) or late (15-H days) after injection of streptozototin or buffer. Controls restricted in food intake lost weight, whereas those fed ad libitum (ad libitum fed group and late control group) gained weight. Early diabetics showed marked weight loss, which continued into the late period. Dry weight of mucosa was decreased by one-third in restricted controls as compared with ad libitum fed controls. Dry weight of mucosa was the same in control
Downloaded from www.physiology.org/journal/ajpendo at Washington Univ (128.252.067.066) on February 13, 2019.
DIGLYCINE
ABSORPTION
IN
E459
DIABETES
and diabetic groups in both early and late sets, but was increased by 50% in the late groups compared with the early groups. Absorption data are given in Table 2. Specific absorption is increased by 40% by restricting intake of controls to half their ad libitum intake. Specific absorption was the same in sets of control and diabetic groups studied early and late, but was lower in both groups of the late set compared to the early set, Total absorption was the same in ad libitum fed and restricted control groups as well as control and diabetic groups of both study periods. Absorption per 100 g body wt (Table 2) was greater in diabetics than in controls for both study periods. Early controls showed greater absorption per 100 g body wt than late controls; however, absorption of late diabetics was greater than the early diabetic group. The difference between ad libitum fed and restricted groups is not significant. Glycine is secreted into the lumen during perfusion of diglycine. Specific glycine appearance (per g dry wt mucosa) was the same for control and diabetic groups in both early and late sets. However, specific appearance (all data pmol/h) was lower in both groups in the late set (control 182 t 18; diabetic 235 t 31) compared to the early set (control 344 t 75; diabetic 394 * 33). Total appearance of glycine was the same in early and late sets of controls and diabetics because of mucosal growth in the late set. For ad libitum and restricted controls, specific appearance was significantly greater in the restricted group (299 t 26) than in the ad libitum group (169 t ll), whereas total appearance was the same. DISCUSSION
Although previous studies showed enhancement of specific absorption of neutral and basic amino acids by diabetes (12, 20), we found no effect of diabetes on specific absorption of diglycine in the rat small intestine. The lack of response of diglycine transport to the stimulus of diabetes is similar to the unresponsiveness of the imino acid-glycine transport system (12). Absorption was studied at two time intervals: early, before mucosal growth was greater in diabetics than controls, and late , when the mucosal hyperplasia response oc-
2. Diglycine
TABLE
Group
-
absorption Specific, per g Dry Wt Mucosa
Total, per Entire Small Intmtine
Per 1OO g Bdy
Wt
mmoL/h
Controls (3-5 days) Ad libitum fed Restricted Early (3-4 days) Control Diabetic Late (15-17 days)
Control Diabetic
1.60 k 0.07 2.26 + 0.W
1.52 + 0.44 1.38 + 0.26
0.68 -+ 0.04 0.76 k 0.06
1.69 k 0.05 1.55 + 0.06
1.04 k 0.04 1.00 k 0.03
0.56 + 0.03 0.65 f 0.03$
1.23 k 0.08t 1.28 * 0.11t
1.11 * 0.05 1.23 2 0.04t
0.41 k 0.02t 0.85 f o.o4=Q
Values are means * SE. libitum fed group, P < 0.05. early control and diabetic gro differs from control group, P
absorption
in streptozotocin
diabetic
HAROLD P. SCHEDL, JANICE WENGER, AND SIAMAK A. ADIBI Gastruenterulugy Research Laboratories, Department of Medicine, University Iuwa College of Medicine, Iuwa City, Iuwa 52242; and Gastrointestinal and Nutritional Unit, Muntefiure Huspitul, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15213
SCHEDL, HAROLD P., JANICE WENCER, AND SIAMAK A. Anxsr. Diglycine absorption in streptozotocin diabetic rat. Am. J. Physiol. 235(5): E457-E460, 1978 or Am. J. Physiol.: Endocrinol. Metab. Gastrointest. Physiol. 4(5): E457-E460, 1978. - Because experimental diabetes in the rat is associated with changes in function of neutral and basic amino acid transport systems, we studied the effects of streptozotocininduced diabetes on dipeptide transport. Absorption was investigated by intraluminal perfusion of the entire small intestine with 20 mM diglycine in pair-fed control and diabetic rats early (3-4 days) and late (15-17 days) after induction of diabetes. In contrast to controls, which grew in body and mucosal weight, diabetics lost body weight but showed mucosal growth. Specific absorption (mmol/g dry wt mucosa per h) was the same in controls and diabetics both early (1.69 vs. 1.55) and late (1.23 vs. 1.28), but was lower in the late than in the early set. Total intestinal absorption (specific absorption x mucosal weight) was the same in early and late controls, but was greater late than early in diabetics because of increased mucosal growth. Pair feeding of controls and diabetics requires restriction of food intake of early controls because food intake of diabetics is low at the early time interval. When controls restricted to the intake of diabetics were compared with ad libitum fed controls, specific absorption was increased in the restricted group. Thus, the greater specific absorption in the early, compared to the late sets of controls and diabetics, is the result of decreased food intake in the early set, intestinal tion
adaptation;
intestinal
growth;
amino
acid absorp-
MAJORITY of amino acid residues of dietary proteins enter the intestinal mucosal cells as small oligopeptides (3, 16). This entry appears to be mediated by a carrier system that is separate from those for free amino acids (3, 16). Prior studies in the rat have shown that alteration in function of the peptide carrier system in response to changes in diet differs from that of the neutral amino acid carrier system (14, 15). In addition to its effect on nutrition, experimental diabetes also has profound effects on mucosal transport function in the rat: absorption of amino acids transported by the neutral and basic amino acid transport systems is increased when studied in vitro (20) and in vivo (12). In the absence of any information on effects of diabetes on peptide transport, we investigated this problem in the present experiments. We chose diglycine for the present studies because its intestinal transport has been THE
0363-6100/78/0000-0000$01.25
Copyright
0 1978 the American
Physiological
rat
of
extensively studied in man and animals (2, 6, 17, 19, 21). Furthermore, diglycine appears to share a common carrier mechanism with other dipeptides and tripeptides (1, 5). MATERIALS
AND
METHODS
Animals. We used male albino rats of the Simonsen strain (Simonsen Laboratories, Gilroy, CA) in the HO200 g weight range (Table 1). Animals were divided into two groups. Rats to be made diabetic were given a single intraperitoneal injection of 100 mg/kg of streptozotocin (supplied by the Upjohn Co., Kalamazoo, MI) freshly dissolved in citrate buffer (pH 4.5). Controls received the buffer only. Control and diabetic groups were pair-fed equal amounts of commercial rat chow (Teklad Mills, Madison, WI). Pair feeding was achieved as follows: a) because food intake of diabetics is decreased for several days after streptozotocin injection, food intake of controls was restricted to that of diabetics initially; b) subsequently, because diabetics developed hyperphagia about 5 days after injection, food intake of diabetics was restricted to that of controls. Food intake by pair-fed controls and diabetics increased from 7 to 22 g/day during the period prior to and during early studies and increased to a mean level of 26 g/day during the several days preceding late studies. Body weight, urine volume, and urine glucose concentrations (TesTape, Lilly) were measured daily. Diabetic animals showed urine glucose concentrations in excess of 2 g/ 100 ml. Mean (*SE) serum glucose concentrations (mg/ 100 ml) in aortic blood from these nonfasted animals were 117 -+ 17 and 195 -t- 20 for early and late control groups, respectively. For diabetics, mean (+SE) serum glucose was 425 -+ 18 in early diabetics and 530 t 23 for late diabetics. Pair feeding of controls with early diabetics requires restriction of food intake of controls to approximately half that of the ad libitum fed controls. Subsequently, for late studies, controls were fed ad libitum. To determine the effect of dietary restriction on diglycine absorption, a separate group of control rats was divided into an ad libitum fed group and a group fed at half the ad libitum intake. Perfusion procedure. For control and diabetic animals, absorption was measured “early” (3-4 days) and “late” (15-17 days) after injection. Early studies defined transport behavior of diglycine before mucosal growth Society
Downloaded from www.physiology.org/journal/ajpendo at Washington Univ (128.252.067.066) on February 13, 2019.
E457
SCHEDL,
E458 TABLE
1. Groups
studied
intestine Weight,
g Dry Wt Mucosa,
Group
Initial
g
was calculated
WENGER,
AND
ADIBI
as follows
corrected mucosal dry wt
Final
= (measured mucosal dry wt)
Controls (3-5 days)” Ad libitum fed Restricted
203 * 3 202 k 3
225 k 7” 183 IL 3cJ1
0.96 k 0.04 0.62 + 0.02”
Early (3-4 days)” Control Diabetic
191 + 2 186 -t 1
186 k 3 156 k 2’=
0.62 -c 0.02 0.65 +_ 0.02
Late (15-17 Control Diabetic
193 k 3 192 k 2
273 k 3”+’ 145 + 3C+“,f
0.93 k 0.03f 1.01 f 0.06”
days)“’
Values are means * SE. ii Time of absorption study after food l) Time of absorption study in was reduced for restricted controls. c Initial differs from final, P < relation to induction of diabetes. 0.05. d Restricted group differs from ad libitum fed group, P < 0.05. e Diabetic group differs from control group, P < 0.05. ’ Group at late period differs from both groups at early period, P < o-05-
in diabetics was increased in comparison with controls, Late studies were to define differences in absorption that appeared after greater mucosal growth was established in diabetics. Ad libitum and restricted control animals were studied after 3-5 days of the feeding regimen Rats were anesthetized by intraperitoneal Nembutal (50 mg/kg). The abdomen was opened by a midline incision, and the common bile duct was ligated. The entire small intestine was perfused as a single unit by inserting cannulas at the pylorus (inlet) and terminal ileum (outlet). The perfusion solution contained 20 mM diglycine (Grand Island Biological Co., Grand Island, NY), and 150 mM NaCl and was perfused at a constant rate (2.5 or 2.0 mllmin) using a peristaltic pump (Cassette Pump, Manostat, New York City). Preliminary studies showed that the diglycine absorption rate is linear for up to a 20 mM infusion concentration. Radioactively labeled polyethylene glycol ([14C]PEG, 10 &i/liter, New England Nuclear, Boston, MA) was used as nonabsorbable indicator. After a 30min initial wash, three lo-min samples were collected in tubes containing sulfosalicylic acid (3.5 mg/ml of sample collected) to inactivate intraluminal peptidase activity. Immediately after perfusion, blood was obtained from the bifurcation of the descending aorta for serum glucose determinations (25) of control and diabetic rats. Intestinal measurements. After blood was obtained, the intestine was stripped from the mesentery. Contents were expressed, and wet weight of the entire intestine was determined. The intestine was then slit longitudinally with a scissors and opened, mucosal side up, onto a glass plate. The mucosa was then scraped from underlying tissue with the edge of a microscope slide. Wet weights of both the mucosal scrapings and underlying tissue were determined, and the scrapings were dried in a vacuum oven at 100°C for 24 h to obtain dry weights. To correct for losses of mucosa incurred in scraping and tranferring to tared containers for weighing and drying, dry weight of mucosa for the entire small
wet wt of entire intestine mucosal wet wt + underlying tissue wet wt > This is a small correction because tissue recovery averaged 95% and ranged from 92 to 96%. AnaZyses. The test solution and samples collected were analyzed for amino acid and peptide concentration using an amino acid analyzer (Beckman model 12OC, Beckman Instruments, Fullerton, CA). [14C]PEG was measured by liquid scintillation counting (Beckman LS-250). Diglycine absorption was calculated as follows total diglycine absorption, mmollh
= [Ci - Cf (IND)] x R where Ci is initial peptide concentration in the test solution and Cf is final peptide concentration after perfusion (both in mmollml); IND is the ratio of initialto-final [l$C]PEG concentrations; R is the rate of perfusion in ml/h. specific diglycine absorption, mmol/h per g dry wt mucosa = (total absorption)/(dry
wt mucosa, g)
net water movement, % = (1 - IND) x 100 Positive values indicate net absorption. luminal appearance rate of glycine, pmol/h = [(C, x IND) - Ci] X R where Cf and Ci are final and initial glycine concentrations (pmol/ml) in the perfused solution, and the other symbols are the same as for total diglycine absorption. Data are presented as mean values t 1 SE for groups of six to nine animals. Tukey’s multiple comparison test (13) and the Student t test (9) were used for statistical analysis. RESULTS
Table 1 shows initial and final body weights and mucosal dry weight for groups studied. At the top are data from a group of normal rats divided into an ad libitum fed group and a group fed half of ad libitum intake (restricted). Shown below are data for control and diabetic groups. Ad libitum and restricted groups were studied after 3-5 days of the feeding regimen. Control and diabetic groups were studied early (3-4 days) or late (15-H days) after injection of streptozototin or buffer. Controls restricted in food intake lost weight, whereas those fed ad libitum (ad libitum fed group and late control group) gained weight. Early diabetics showed marked weight loss, which continued into the late period. Dry weight of mucosa was decreased by one-third in restricted controls as compared with ad libitum fed controls. Dry weight of mucosa was the same in control
Downloaded from www.physiology.org/journal/ajpendo at Washington Univ (128.252.067.066) on February 13, 2019.
DIGLYCINE
ABSORPTION
IN
E459
DIABETES
and diabetic groups in both early and late sets, but was increased by 50% in the late groups compared with the early groups. Absorption data are given in Table 2. Specific absorption is increased by 40% by restricting intake of controls to half their ad libitum intake. Specific absorption was the same in sets of control and diabetic groups studied early and late, but was lower in both groups of the late set compared to the early set, Total absorption was the same in ad libitum fed and restricted control groups as well as control and diabetic groups of both study periods. Absorption per 100 g body wt (Table 2) was greater in diabetics than in controls for both study periods. Early controls showed greater absorption per 100 g body wt than late controls; however, absorption of late diabetics was greater than the early diabetic group. The difference between ad libitum fed and restricted groups is not significant. Glycine is secreted into the lumen during perfusion of diglycine. Specific glycine appearance (per g dry wt mucosa) was the same for control and diabetic groups in both early and late sets. However, specific appearance (all data pmol/h) was lower in both groups in the late set (control 182 t 18; diabetic 235 t 31) compared to the early set (control 344 t 75; diabetic 394 * 33). Total appearance of glycine was the same in early and late sets of controls and diabetics because of mucosal growth in the late set. For ad libitum and restricted controls, specific appearance was significantly greater in the restricted group (299 t 26) than in the ad libitum group (169 t ll), whereas total appearance was the same. DISCUSSION
Although previous studies showed enhancement of specific absorption of neutral and basic amino acids by diabetes (12, 20), we found no effect of diabetes on specific absorption of diglycine in the rat small intestine. The lack of response of diglycine transport to the stimulus of diabetes is similar to the unresponsiveness of the imino acid-glycine transport system (12). Absorption was studied at two time intervals: early, before mucosal growth was greater in diabetics than controls, and late , when the mucosal hyperplasia response oc-
2. Diglycine
TABLE
Group
-
absorption Specific, per g Dry Wt Mucosa
Total, per Entire Small Intmtine
Per 1OO g Bdy
Wt
mmoL/h
Controls (3-5 days) Ad libitum fed Restricted Early (3-4 days) Control Diabetic Late (15-17 days)
Control Diabetic
1.60 k 0.07 2.26 + 0.W
1.52 + 0.44 1.38 + 0.26
0.68 -+ 0.04 0.76 k 0.06
1.69 k 0.05 1.55 + 0.06
1.04 k 0.04 1.00 k 0.03
0.56 + 0.03 0.65 f 0.03$
1.23 k 0.08t 1.28 * 0.11t
1.11 * 0.05 1.23 2 0.04t
0.41 k 0.02t 0.85 f o.o4=Q
Values are means * SE. libitum fed group, P < 0.05. early control and diabetic gro differs from control group, P
