Glucose Transport by Rat Small Intestine after Extensive Small-Bowel Resection ERNEST URBAN, MB, BS, FRACP, FACP, and DIANE P. HALEY, BS

In rats 50 cm of proximal or distal small intestine were resected, preserving duodenum and terminal ileum. Glucose transport was studied 5-6 weeks later, using everted gut sacs from duodenum, ileum, and also from a midgut segment consisting of intestine located preresection at mid-small intestine. Sham-operated animals served as controls: The inner (serosal) fluid medium in sacs from duodenum and midgut gained glucose; ileal sac serosal medium lost glucose. Proximal resection resulted in significant growth of duodenal and midgut mucosa. Duodenal transport specific activity (transport per gram dry mucosa) decreased from control values, but mucosal growth compensated so sac transport capacity (transport per centimeter sac length) remained unchanged. Midgut transport specific activity remained unchanged, thus sac transport capacity directly mirrored increased mucosal mass. Ileal sac serosal medium now accumulated glucose; there was no mucosal growth. Transport specific activity and sac transport capacity of ileum increased in parallel. After distal resection there was no alteration of either duodenal and midgut mucosal masses or transport specific activities, hence sac transport capacities remained unchanged. Ileal sac serosal medium also accumulated glucose, but now both transport specific activity and mucosal mass increased. The resultant increased sac transport capacity was identical to that of ileum after proximal resection. In all sacs from control and resected animals uphill [14C]glucose concentration differences developed between medium and mucosa. Activity of the mucosal uptake process, assessed in terms of a ratio of mucosal intracellular f u i d radioactivity to mucosal medium radioactivity, usually mirrored altered transport specific activity. This indicates that the increased undercoats tissue mass that accompanied increased mucosal mass did not critically affect transport. The most striking findings were: (1) decreased duodenal transport specific activity after proximal resection with mucosal growth compensating; and (2) identical adaptations of ileal segment transport capacities after either proximal or distal small-bowel resections, although mechanisms differed. The present study provides a base for further examinations of carrier-mediated hexose transport after extensive loss o f small intestine.

Functional and morphological adaptations of remaining intestine after extensive small-bowel resecFrom the Division of Gastroenterology, Department of Medicine, The University of Texas Health Science Center at San Antonio and the Audie L. Murphy Veterans Administration Hospital, San Antonio, Texas 78284. This study was supported by the National Institute of Arthritis, Metabolism and Digestive Disease Research Grant 1 RO 1 AM16840. A part was presented at the 29th Annual Meeting of the Southern Society for Clinical Investigation, New Orleans, Louisiana, February I, 1975, and has appeared previously in abstract form (i). Address for reprint requests: Dr. Ernest Urban, Gastroenterology ( l l l B ) , Audie L. Murphy Memorial Veterans Hospital, 7400 Merton Minter Boulevard San Antonio, Texas 78284.

tion have been previously described (2-13). Subsequent long-term survival depends largely on the transport capacity of remaining intestine. Glucose, the sole hydrolytic product of starch, also forms 50% of hexoses resulting from the hydrolysis of sucrose and lactose. In man, dietary carbohydrate accounts for about half the ingested calories and 80% is eventually hydrolyzed to glucose before absorption (14). Hence characterization of intestinal adaptation for glucose transport assumes major significance. The few prior studies in man (4), dog (12), and the rat (2, 5, 11), have shown enhanced glucose transport, but the studies have either been limited

Digestive Diseases, Vol. 23, No. 6 (June 1978)

0002-9211/78/0600-0531505.60/1 9 1978DigestiveDiseaseSystems, Inc.

531

URBAN AND HALEY or distal small intestine was resected (Figure I) and continuity of the intestine restored by end-to-end anastomosis. Sham-operated animals underwent simple intestinal transection at mid-small intestine and reanastomosis without removal of intestinal tissue (Figure 1). Details of operative procedures have been described (9). Preparation of Gut Sacs. Five to six weeks after intestinal resection or sham operation the animals were fasted for 24 hr, anesthetized with intraperitoneal sodium pentobarbital, and the abdomen was opened through a midline incision. Duodenum and small intestine were rapidly removed and rinsed with ice-cold saline and placed in sodium-Ringer solution (15), pH 7.2, 4~ C, and gassed with 95% 02-5% CO2. Everted gut sacs each approximately 5 cm long were prepared by the technique of Wilson and Wiseman (16) from duodenum, midgut, and ileum. The location of the duodenal and ileal tissues used to prepare the sacs was identical in the three groups of animals, but the intestine used to prepare sacs from midgut was dependent on the type of prior intestinal operation (Figure 1). In sham-operated rats, two midgut sacs were made--one just proximal to and the other just distal to the site of transection and reanastomosis (M1 and M2,

or their major emphasis has been on other facets. We utilized everted gut sacs to examine glucose transport in duodenum, midgut, and ileum of rats 56 weeks after 50 cm resection of either proximal or distal small bowel. The experiments demonstrated adaptations of glucose transport that varied with the intestinal segment studied and with the type of resection.

MATERIALS AND METHODS Sprague-Dawley strain male albino rats weighing about 120 g, allowed free access to standard commercial rat laboratory chow (Wayne Lab Blox, Allied Mills, Libertyville, Illinois) and to tap water, were divided into three groups: proximal small-bowel resection, distal smallbowel resection, and sham-operated animals. Food but not water was withheld for 24 hr before and 24 hr after intestinal resection or sham operation. Intestinal Resection. Under intraperitoneal sodium pentobarbital anesthesia (40 mg/kg), 50 cm of either proximal

TRANSECT 50cm PROXIMAL TO ILEOCECAL JUNCTION

,1,

MI

ileum

I M2 midgut

duodenum

5cm BEYOND LIGAMENT OF TREITZ

PROXIMAL RESECTIO~N//~ ~t {f V

I

f ~777777J77JJJ77J777JA /I 50cm EXCISED '--'-

ileum " ~

J M2 - ' midgut

duodenum

DISTAL ~ RESECTION~

IO cm PROXIMAL TO ILEOCECAL JUNCTION

"l

I

L.~

~///////////////////A

MI L midgut ' -

50 cm EXCISED

_l -'

ileum

duodenum Fig 1. Diagram showing locations of intestinal transection for sham-operated rats, excised segments for proximal and distal resections, and segments of small interstine utilized for gut sacs 5-6 weeks later. Note that in the sham-operated animals, gut sacs were made from intestine both proximal (M1) and distal (M2) to the site of midgut intestinal transection, allowing comparisons with corresponding midgut segments from each type of resection.

532

Digestive Diseases, Vol. 23, No. 6 (June 1978)

GLUCOSE TRANSPORT AFTER INTESTINAL RESECTION Figure 1); in resected rats only one midgut sac was prep a r e d - i n proximally resected rats the site was immediately distal to the site of reanastomosis (M2, Figure 1); in distally resected rats the site was immediately proximal to the site of reanastomosis (M1, Figure 1). Scar tissue at the site of the prior intestinal reanastomosis was readily identifiable at sacrifice. The sacs were filled with approximately 0.3 ml sodiumRinger solution (15) containing 5 mM glucose, about 0.08 b~Ci/ml [14C]D-glucose (uniformally labeled, specific activity 15 mCi/mM; New England Nuclear Corp., Boston, Massachusetts) to give a specific activity of 16 ~Ci/mmole and 0.5 /xCi/ml [methoxy-aH] inulin (molecular weight 5000, specific activity 50-150 mCi/g; New England Nuclear). The sacs were incubated for 60 min at 37~ in 10 ml of the same medium using flasks aerated with 95% 02-5% CO2, and agitated in a Dubnoff metabolic shaking incubator. Sacs whose inner fluid volume decreased during incubation were rejected. (Calculations showed either markedly reduced or absent glucose transport indicating a sac with a leak or otherwise compromised function.) After incubation the sacs were measured for length, opened lengthwise, spread on a glass plate, and a mucosal fraction separated from undercoats by scraping with the edge of a microscope slide. The same person (D.P.H.) performed all tissue scrapings. The two tissue fractions were weighed, dried in a vacuum oven for 24 hr, reweighed, and solubilized with Protosol (New England Nuclear). '4C and aH radioactivities of the tissue fractions and of the initial and final inner (serosal) and outer (mucosal) sac solutions were measured by two-channel liquid scintillation counting. The counts were corrected for quench and radioactivity expressed as disintegrations per minute (dpm) for subsequent calculations. Calculations. In the following formula the subscripts i and f refer to initial (preincubation) and final (postincubation) values. Glucose transport was defined as the net increase of 14C radioactivity, in the inner sac medium and calculated as follows: (If14CVf) -

Glucose transports =

(]il4CVi)

S&'4C(Lsor D,)T

where 114C is the inner sac solution [14C]glucoseradioactivity in dpm/ml; V is the volume of inner sac solution in ml; SA14Cis the specific activity of glucose in the medium in dpm//zmol glucose; L is the length of the sac in cm; D is the dry weight of sac mucosa in g; T is the time of incubation in hours. Transport data were thus calculated in two different ways: micromoles per hour per cm intestinal sac length and micromoles per hour per gram dry mucosa. Water content of the sac mucosa and underlying tissue after incubation was calculated from the difference of wet and dry tissue weights and expressed as per cent of wet tissue weight: Percent tissue water =

100(Wr- De)

Ws

where Wis the wet weight of tissue and D its dry weight. Digestive Diseases, Vol. 23, No. 6 (June 1978)

Inulin space extracellular fluid (ECF) of the sac mucosa and undercoats was calculated as percent of tissue water using the following formula:

ECF= (T}HWr)IO0 SiaH(Wr Ds) where Tall is the tissue [all]inulin radioactivity in dpm/ml of tissue water and Sail is the sodium-Ringer solution [all]inulin radioactivity in dpm/ml. The other symbols have the same meaning as above. We assumed no entry of [all]inulin into the intracellular fluid compartment (ICF) and by the end of the 1-hr incubation, tissue ECF inulin space had reached steady state. All results are given as mean values _+ 1 SEN. The Student's t test was used for statistical comparison of differences between the means from sham-operated and proximally resected animals and between the sham-operated and distally resected group. P values of less than 0.05 were considered to indicate statistically significant differences.

RESULTS Table 1 shows data of body weights of the shamo p e r a t e d a n d p r o x i m a l l y and d i s t a l l y r e s e c t e d groups of experimental animals at sacrifice. There were no significant differences between the groups, consistent with our prior experience (9, 10). Also shown are the dry weights of m u c o s a and underlying tissues (mg/cm) for the three experimental groups. Proximal intestinal resection resulted in significant increases in m u c o s a and undercoats of duodenum and midgut (P < 0.01 in each instance) while ileal tissues did not increase when compared to the sham-operated group. Distal small-bowel resection did not result in significant increases in either m u c o s a or undercoats of d u o d e n u m or midgut but both ileal m u c o s a and undercoats increased markedly from shams (P < 0.01). Glucose Transport. Table 2 summarizes net glucose transport in the gut sacs after 1 hr incubation in micromoles glucose per gram dry m u c o s a and per centimeter sac length in the sham-operated and proximal and distal resection groups for duodenum, midgut, and ileum. In the sham-operated group of animals, the serosal medium of everted gut sacs from d u o d e n u m and midgut gained glucose; serosal medium in sacs from ileum showed a net loss of glucose. Changes of transport after resection will be described by two terms: (1) mucosal transport specific activity defined as transport per gram dry mucosa; and (2) sac transport capacity defined as transport per centimeter sac length. Proximal Resection. There was significant mucosal growth in d u o d e n u m and midgut (Table 1). In du-

533

URBAN

AND

HALEY

TABLE 1. GROUPS STUDIED: CONTROL AND RESECTED RATS*

Body wt (g, final) Dry wt of intestinal tissues (mg/cm) Mucosa Duodenum Midgut (MI) Midgut (M2) Ileum Undercoats Duodenum Midgut(M0 Midgut (Ms)

Ileum

Sham operated

Proximal resection

resection

( N = 11)

(N = 10)

(N

324 • 13

334 • 18

5.8 5.6 5.7 5.1

• 0.3 + 0.3 -+ 0.6 -+ 0.3

9.3 • 0.4t

7,9 6.3 5.9 6,8

-+ 0.4 + 0.4 • 0.4 • 0,4

8,0 • 0.4t 5.9 + 0.6 9.9 -+ 0,5t 9.0 +- 0.6t 7.1 • 0,8

Distal = 12)

-343 + 20

6.5 -+ 0:3 6,2 +- 0.4 7.4 -+ 0.4? 8,5 • 0.5 6.7 +- 0,4 10.6 • 0.4t

*Values are means -+ SEM; N = number of animals studied. Body weights (final) were at sacrifice 5-6 weeks after resection or sham operation; other data were obtained from gut sacs after t hr incubation. tSignificant difference from corresponding data of sham-operated animals.

odenum, mucosal transport specific activity decreased 53% (P < 0.01), but the 60% increase in duodenal mucosal mass compensated so that sac transport capacity remained unchanged. In contrast to duodenum, midgut mucosal transport specific activity did not change significantly but the 41% increase in mucosal mass resulted in 47% increase of sac transport capacity (P < 0.02). In ileum where serosal medium of the gut sacs now accumulated glucose, mucosal transport specific activity increased (P < 0.02) but mucosal mass did not change so the increased sac transport capacity (P < 0.02) directly mirrored the increased mucosal transport specific activity.

Distal Resection. Mucosal mass increased significantly only in ileum (45%; Table 1). Mucosal transport specific activity did not change in duodenum or midgut. In ileum, where serosal medium of the sacs now also accumulated glucose, mucosal transport specific activity increased (P < 0.01). Hence, compared to shams, the sac transport capacities of duodenum and midgut remained unchanged but ileal sac transport capacity, reflecting the combination of increased mucosal transport specific activity and mucosal mass, increased (P < 0,01). Glucose Gradients. Figure 2 summarizes [~4C]glucose concentration of outer and inner sac media, mucosa, and undercoats after 1 hr in-

TABLE 2. GLUCOSE TRANSPORT*

Mucosal transport specific activity (/~mol/g dry mucosa in 1 hr) Duodenum Midgut (M1) Midgut(M2)

Ileum Sac transport capacity (/zmol/cm sac length in 1 hr) Duodenum Midgut (MI) Midgut (M2)

Ileum

Sham operated

Proximal resection

resection

(N = tt)

( N = 10)

( N = 12)

36.2 +- 4.7t

69.4 • 4.5 69.2 - 12.7

77.5 70.4 66.8 -10.4

+ -+ + +

9.8 13.3 17.6 1.8

0.43 0,42 0.36 -0.05

+_ 0.05 +- 0.05 - 0,06 -+ 0,01

72.2 -+ I 1 . 1 4.0 + 6.1t

0.38 -- 0.03 0.53 -+ 0,05t 0.02 -+ 0.03?

Distal

1 . 3 -+ 3.2t

0.39 - 0.02 0.45 -+ 0.06 0:02 -+ 0.02?

*Values are means -+ SEM; N = number of sacs incubated. ?Significant difference from corresponding data of sham-operated animals.

534

Digestive Diseases, VoL 23, No, 6 (June 1978)

GLUCOSE TRANSPORT AFFER INTESTINAL RESECTION

A iLEUM

MIDGUT

DUODENUM

M2

o

4-

/,

~,

z

o o

l'

3L9

,, i

1

I

i

i

MUC MUC- U. SER. MED. OSA CTS MEO.

I

I

i

!i~ " t

I

i

MUC MUC" O. SEN. MED OSA CTS MED.

MUC MUC" U. SER. MED OSA CTS, MED.

I

I

i

MUr MUC- U SER MED. OSA CTS, MED

B iLEUM

MIDGUT

DUODENUM

M2

E

~6-

i,

E

%

40

Z

8

ttl

2-

B

t,9

I

!

~uc,*uc-

d,

"1

G. s~.

'

I

MUC MUC- U SER. MED. OSA CTS, MED

MED, OSA CTS MED

MUC MUC. U. SER. MED OSA CTS MED

C 9=

M 6-

ILEUM

MIDGUT

DUODENUM

]

- -

,,

{----3

i' i

~.~

MUC. MUC- O. SER. WE{}. OSA CTS MED.

i

I

i

MUC MUC- U. SER MED. OSA CTS IdED

i

I

i

I

MUC, MIX;- U. SER MED. OSA CTS.IVIED

Fig 2. Concentrations of radioactivity across gut sacs prepared from the various intestinal segments of (A) sham-operated (10 observations), (B) distally resected (10 observations), and (C) proximally resected (12 observations) animals after incubation for 1 hr as described in Materials and Methods. Radioactivities of mucosa and undercoats (U. CTS.) are expressed per ml tissue water. M1 and M2 of midgut represents data from midgut tissues located as shown in Figure 1. All values are mean -+ SEM. Prior to incubation, mucosal and serosal media had identical [14C]glucose concentrations of about 170,000 dpm/ml (specific activity approximately 34,000 dpnv'/zmot glucose) and tissues were free of radioactivity. *P < 0.05 compared to sham.

cubation of all sacs, The postincubation radioactivities of mucosa and undercoats are expressed in dprrdml total tissue water and are directly comparable to posfincubation inner (serosal) and outer Digestive Diseases, Vol. 23, No. 6 (June 1978)

(mucosal) media radioactivities. In each instance a 14C uphill concentration difference developed between the mucosal medium and mucosal tissue. The mucosat 14C accumulation was greater from proxi-

535

URBAN AND HALEY real than from distal small intestine, indicating an accumulation mechanism most active proximally. After proximal resection duodenal mucosal 14C concentration decreased (P < 0.05) and ileal mucosal 14C concentration increased (P < 0.02) in parallel with the changes of mucosal transport specific activities in these two tissues (Table 2). After distal resection, the rise in mean ileal mucosal 14C concentration failed to reach statistical significance, even though ileal mucosal transport specific activity increased (Table 2). In all instances the concentration differences from mucosa to the inner sac medium were downhill, consistent with a diffusion process after extrusion of [14C]glucose from the mucosal cells. There were no apparent effects from the increased mass of undercoat tissues (Table 1). Tissue Water. Postincubation water content of mucosa and undercoats of all sacs are depicted in Figure 3. Also shown is the inulin space ECF for each tissue. The intracellular fluid (ICF) space is the difference between total tissue water and ECF inulin space. The total water content of mucosa was about 87% in all sacs, the differences between groups were not significant. Similarly the total water content of undercoats was about 83%, and differences were also not significant. However, the division of total tissue water into ECF and ICF was altered in the resected groups. Proximal Resection. ECF of mucosa (Figure 3a) d e c r e a s e d 41% in d u o d e n u m (P < 0.001), decreased 44% in midgut (P < 0.001), and decreased 35% in ileum (P < 0.01) compared to the sham-operated group. ECF of undercoats (Figure 3b) decreased 37% in duodenum (P < 0.001) and 52% in midgut (P < 0.001), but the decrease in ECF of undercoats in ileum (23%) did not quite reach statistical significance (P = 0.07). Distal Resection. ECF of mucosa (Figure 3a) did not change in duodenum but decreased 22% in midgut (P < 0.02) and 24% in ileum (P < 0.02). There were no significant changes of ECF in undercoats of duodenum, midgut, or ileum (Figure 3b). Mucosal Tissue Glucose Uptake

Although this study was not designed to measure postincubation mucosal ICF [14C]glucose concentrations directly, approximations were calculated from p o s t i n c u b a t i o n mucosal tissue water ['4C]glucose concentrations (Figure 3). We assumed that, at the end of the 1-hr incubation period, the concentration of 14C radioactivity in ECF of mucosa

536

was identical to that of the serosal medium (Figure 2). Mucosal ICF 14C concentrations were then calculated as the difference between total and ECF 14C radioactivities of mucosal tissue. Activity of the glucose uptake process by mucosa of the various segments was compared by means of a mucosal uptake ratio defined as the ratio of '4C radioactivity in ICF of mucosa to mucosal medium. These calculated mucosal uptake ratios are shown in Table 3. The ratios should be a measure of activity of the first step in transport from mucosal to serosal medium. Comparisons of the mucosat uptake ratios between sham-operated and the two types of resections (Table 3) with glucose transport (Table 2), and mucosal mass (Table 1) show the following: Proximal Resection. In duodenum the mucosal uptake ratio (Table 3) decreased (P < 0.01). Mucosal transport specific activity decreased, but sac transport capacity did not change (Table 2). Mucosal mass increased (Table 1). The data are consistent with the previously described functional immaturity related to increased mucosal cell turnover after intestinal resection (6, 7, 1t). In midgut there was no change in mucosal uptake ratio (Table 3) or in mucosal transport specific activity (Table 2), implying no alteration in mucosal cell transport function. The increased midgut sac transport capacity (Table 2) thus was solely the result of increased mucosal mass (Table 1). In ileum both the mucosal uptake ratio increased (P < 0.05, Table 3) and mucosal transport specific activity increased (Table 2) without alteration of mucosal mass (Table 1), here implying increased mucosal celt transport activity as the adaptive mechanism: Distal Resection. In both duodenum and midgut neither the mucosal uptake ratios (Table 3), mucosal transport specific activities (Table 2), nor mucosal masses (Table 1) were altered by resection. In ileum the m u c o s a l uptake ratio was unaltered (Table 3), b u t transport specific activity increased (however, less than in ileum after proximal resection, Table 2). There was also significant mucosal growth (Table 1) and increased sac transport capacity (Table 2). This suggests that the more important factor of the adaptive response was increased mucosal mass, but altered mucosal cell transport function and/or effects of undercoat tissues cannot be excluded. In general, after proximal or distal resection the correlations between altered mucosal transport specific activities, sac transport capacities (Table 2), and mucosat mass (Table 1) with altered mucosal Digestive Diseases, VoL 23, No. 6 (June 1978)

GLUCOSE TRANSPORT AFTER INTESTINAL RESECTION I00 -

MI 80-

ILEUM

MIDGUT

DUODENUM

M

I

rY Lul .~

60-

FZ Ld o 40rY uJ

I

20%,%,% o-

A

SHAM PROX DIST (10) RES RES (lO) 02)

I00 -

SHAM DIST (10) RES

(~2)

SHAM PROX RES (10) 00)

MIDGUT

DUODENUM

SHAM PROX (10) RES 0o)

DIST RES (12)

ILEUM M2

Mi

i-arm i-,,w,~

80-

82 Ld

} 60 t..Z I.d o 4O LLI

20-

x\\

~

.x.\"~

"\"

,N\'s

,~xx xxx

8

SHAM PROX DIST (10) RES RES (~0) (12)

SHAM DIST (10) RES 02)

SHAM PRO\ (10) RES (10)

SHAM PROX (10) RES (10)

DIST RES (12)

Fig 3. Percent water content of (A) mucosa and (B) undercoats of gut sacs prepared from the various intestinal segments of s h a m - o p e r a t e d and the two types of resected animals after 1 hr incubation. The combined height of the shaded and clear portion of each bar represents total tissue water. The shaded portion r e p r e s e n t s extracellular fluid determined from pH]inulin content, the remaining clear part r e p r e s e n t s intracellular fluid content taken as being the difference between total tissue water and extracellular fluid. M1 and M2 of midgut r e p r e s e n t data from midgut tissues whose location is shown in Figure 1. All values are m e a n -+ SEM. N u m b e r s in parentheses are the n u m b e r of observations. *P < 0.05 c o m p a r e d to sham.

uptake ratios (Table 3) imply that the movement of glucose from mucosal to serosal media was a mucosal function that was not critically affected by changes in undercoat tissues. DISCUSSION Everted gut sacs have been widely used to study intestinal transport. We incubated the sacs for 1 hr even though uptake of substrate is by then no longer Digestive Diseases, Vol. 23, No. 6 (June 1978)

linear with time. Incubation resulted in substrate differences (Figure 2), indicating viability. Equilibration of inulin within tissues as a marker of ICF/ ECF distribution is a slow process (17, 18), and there are varying tissue thicknesses (Table 1). Thus to ensure inulin equilibration we incubated the sacs 1 hr and did not explore kinetics of mucosal accumulation and release. Also considered was the use of a nonmetabolizable transport substrate such as 3-O-methyl-D-glucose. Its transport is much slower

537

URBAN AND HALEY TABLE 3. MUCOSALUPTAKERATIO* Sham operated ( N = 10)

Duodenum Midgut (M1) Midgut (M2) ileum

4.66 3.32 3.16 1.73

+ 0.10 + 0.07 -+ 0.09 -+ 0.07

Proximal resection ( N = 10)

Distal resection ( N = 12)

4.21 +- 0.127

4.59 -+- 0.09 3.35 - 0.08

3.19 +- 0.10 1.99 -+ 0.09t

1.76 -+ 0.06

*The uptake ratio is defined as ICF mucosal radioactivity/mucosal medium radioactivity after 1 hr incubation and was calculated using data depicted in Figures 2 and 3 (see text). Values are means - SEM; N = number of observations. tSignificant difference from corresponding data of the sham-operated group.

than glucose (19), an important limiting factor in transport measurements in distal small bowel. Glucose, however, also is metabolized, supplies energy to the intestinal mucosa, and there is greater glucose utilization proximally than distally. Intestinal resections of similar magnitude to those of this study have been shown not to alter the magnitude of small-bowel mucosal glucose metabolism in the remnant (20). In that study approximately 5% of the glucose present was metabolized by mucosal homogenates from proximal small intestine and less than 1% by mucosal homogenates of distal small bowel when incubated for 1 hr at 37~ C. These data also imply that after incubation most of the measured radioactivity can be expected to be glucose. As a primary function, our experiments explored only differences between similar sites in sham-operated and resected rats. Hence we decided to use glucose as transport substrate in this study. Thus, within inherent limitations of the technique, this in vitro study with gut sacs examines in detail the adaptation of glucose transport in the rat in remaining intestine after either extensive proximal or distal small-bowel resection. Sham-operated animals exhibited proximal-to-distal gradients of sac transport capacity (transport/cm), mucosal transport specific activity (transport/g), and intestinal mass as well as the failure of in vitro ileum to move glucose against concentration differences, all in agreement with prior descriptions of glucose t r a n s p o r t in normal rats (19, 21, 22). Resection altered sac transport capacity by two mechanisms: (1) changes of mucosal mass, and (2) changes of mucosal transport specific activity. The changes of mucosal transport specific activity generally paralleled altered mucosal uptake ratios. The ratios were considered to reflect muco-

538

sal activity in the translocation of glucose from mucosal to serosal media. Proximal small-bowel resection was associated with significant increases of mucosal mass in duodenum and midgut. Transport specific activity of mucosa decreased in duodenum and was unchanged in midgut. In ileum where gut sacs accumulated glucose, transport specific activity increased. In duodenum, the increase in mass compensated for the less active mucosal transport and sac transport capacity remained the same as in the sham-operated animals. The mechanism for the decreased duodenal mucosal transport specific activity is unclear. It may be the result of increased rate of mucosal epithelial cell turnover and consequent functional immaturity (6, 7, 11). A similar finding in duodenum has been described for calcium transport after proximal resection (9). It is significant that in normal animals, transport rates of glucose and calcium are maximal in duodenum and are likely to be most affected by functional immaturity. In midgut the situation was simpler--mucosal transport specific activity did not change and sac transport capacity inc r e a s e d in parallel with the increased m u c o s a l mass. In contrast to duodenum and midgut, ileal mucosal transport specific activity increased and ileal sac transport capacity increased without any increased mucosal mass. Distal resection did not alter mucosal mass in duodenum or midgut; mucosal mass increased only in ileum. Mucosal transport specific activities of duodenum and midgut, and thus sac transport capacities, were the same as in shams. However, in ileum the sacs gained glucose. Transport specific activity increased. This, in association with increased mucosal mass, resulted in increased sac transport capacity. However, the mucosal uptake ratio was unchanged from the sham-operated group, implying that mucosal cell uptake activity was not altered by distal resection. Undercoats mass increased and may have critically affected net glucose transport. We think this is unlikely because increased undercoats mass in other sites did not alter the relationship between net glucose transport and mucosal uptake ratio. We speculate that there must have been some increased mucosal cell transport function in ileum after distal resection (but less than in ileum after proximal resection). This, coupled with increased mucosal mass, resulted in the increased ileal sac transport capacity. It is noteworthy that the ileal sac transport ,capacities after either proximal or distal resections were identical, ie, the same Digestive Diseases, Vol. 23, No. 6 (June 1978)

GLUCOSE TRANSPORT AFTER INTESTINAL RESECTION functional adaptations for glucose were achieved after either type of resection, albeit by different combinations of mechanisms. Transport capacities of remaining bowel after intestinal resections are the most important parameter determining subsequent long-term nutritional status and survival. In the rat as much as 80% small intestine (2) has been resected without long-term loss of body weight. In this study, 50 cm of small intestine, approximating 50% of small bowel (23) were resected without loss of body weight, in agreement with prior experience (9-11, 13) even though nutritional intake after small-bowel resection has been shown to differ in the two types of resections. Proximally resected rats maintain body weight and growth patterns by compensatory hyperplasia while distally resected rats increase food intake (13). We did not measure food intake but allowed ad libitum access. Ileal sacs from the sham-operated group of this study behaved tike normal rat ileum which does not move glucose against a concentration gradient in vitro (21). The present study demonstrates the induction of glucose transport in terminal ileum after both proximal and distal small-bowel resection with and without increased mucosal mass (Table 1). Induction of glucose transport in the distal small intestine has also been described in semistarved rats in association with reduced intestinal mass (21, 24). Thus in ileal mucosa, there must reside an induceable reserve transport mechanism for glucose, responsive to increased absorptive demands of the animal, that is independent of mucosal mass. An ileal reserve for calcium transport after small-bowel resection has also been described (9) lending further emphasis to the ileum being an organ with major functional reserves. Accumulation of sugar within intestinal tissues is a reproducible means of showing active substrate movement against concentration differences (19). This study demonstrates (Figure 2) that after incubation glucose concentration differences were present between mucosal medium and mucosal tissues in all sacs. The greatest concentration differences were found in proximal intestine in all three experimental groups. The proximal-to-distal mucosal accumulation gradients of the shams persisted after resection. This indicates that the greatest active transport function continues to reside in proximal intestine after either type of resection. The close parallelism b e t w e e n altered m u c o s a t transport specific activities and mucosal uptake ratios confirm the validity of transport data calculated Digestive Diseases, Vol. 23, No. 6 (June 1978)

from accumulations of radioactivities in inner sac media. The data also show that transport function of mucosal epithelial cells is adaptive. The accumulation of radioactivity in mucosa and the subsequent downhill concentration differences to serosal medium are consistent with an active transport step at the mucosal cell brush border and subsequent diffusion of glucose across the basal and lateral faces of the epithelial cell (14, 19). This relationship was not critically altered by intestinal resection. Accumulation of osmotically active substrate within the cell during the process of translocation will result in increased intracellular space. Water can be trapped by intestine during solute transport, both sugars and amino acids being osmotically active (17, 18, 25-27). Tritiated inulin has been widely used to measure ECF, although apparent differences of inulin space using 3H- and 14C-labeled inulin moieties have been reported (26). Another comparison between the two labeled inulins, however, gave values that agreed within 4% (18). In this study we used [3H]inulin, all gut sacs were treated in an identical manner, and only comparisons of [3H]inulin spaces between the sham-operated and resected groups were of interest. The comparisons should be valid. As shown in Figure 3, proximal resection resulted in decreased ECF inulin space in mucosa of duodenum, midgut, and ileum; distal resection resulted in decreased inulin space of midgut and ileal mucosal tissues. There were no changes in total water content of mucosa. This implies increased ICF in these mucosal tissues, presumably the consequence of osmotically active intracellular substrate accumulations. Intracellular sugar, assumed to remain free, osmotically active, and accumulating as a result of transport, will result in water movement into mucosal cells during transport. Consequently there will be cellular swelling and decreased ECF: in other words, an inverse relationship should result between altered glucose transport and ECF changes after resection. Comparisons of mucosal transport specific activities in Table 2 with altered water partitioning in Figure 3 do not support this simple concept. Active transport of glucose in vitro is strictly sodium dependent (19, 28), and sodium influx is coupled to sugar influx in a 1:1 ratio (29). Therefore, glucose-facilitated sodium influx after resection cannot also account for the altered water distribution. Intestinal sodium transport is, however, only partially dependent on glucose absorption, and

539

URBAN A N D H A L E Y

there are regional differences: jejunal sodium absorption has been shown to be much more strongly glucose dependent than ileal sodium absorption not only in the rat (30-32), but also in rabbit (33) and in man (34, 35). Furthermore glucose-dependent sodium transport has been shown to increase in ileum after jejunectomy (8). An earlier preliminary" report has also demonstrated that resection may effect sodium transport independent of sodium-coupled glucose transport, but regional differences were not studied (3). Although intracellular sodium concentrations are low in normal intestinal mucosal cells (28), we speculate that specific adaptation of sodium transport independent of sodium-coupled glucose transport may play a leading role in the altered ICF/ECF partitioning described here. ACKNOWLEDGMENTS

The authors are grateful to Dr. Harold P. Schedl for valuable suggestions during the preparation of the manuscript. We thank Betty Medina for her secretarial skill.

REFERENCES 1, Urban E, Haley D: Tissue water partition during glucose transport after extensive intestinal resection in the rat (abstract). Clin Res 23:46A, 1975 2. Bury KD: Carbohydrate digestion and absorption after massive resection of the small intestine. Surg Gynecol Obstet 135:177-187, t972 3~ Bury K, Titson MD, Wright HK: Increased unidirectional active sodium transport in ileal remnants following jejunectomy (abstract). Gastroenterology 62:728, 1972 4. DoMing RH, Booth CC: Functional compensation after small bowel resection in man. Lancet 2:146-147, t966 5, Dowling RH, Booth CC: Structural and functional changes following small intestinal resection in the rat. Clin Sci 32:I39-149, t967 6. Dowling RH, Gleeson MH: Cell turnover following small bowel resection and by-pass. Digestion 8:176-190, 1973 7. Hanson WR, Osborne JW: Epithelial cell kinetics in the small intestine of the rat 60 days after resection of 70 per cent of the ileum and jejunum. Gastroenterology 60:10871097, 1971 8. Tillson MD, Clifford B: A glucose-stimulated non-electrogenic sodium pump in ileal remnants after jejunectomy. J Surg Res 18:405-408, 1975 9, Urban E, Pena M: In vivo calcium transport by rat small intestine after massive small bowel resection. Am J Physiot 226:1304-1308, 1974 10. Urban E: Serum proteins in the rat after extensive proximal or distal small bowel resection, Digestion 13:316-319, 1975 1l. Weser E, Hernandez MH: Studies of small bowel adaptation after intestinal resection in the rat. Gastroenterology 60:6% 75, 1971 12. Wilmore DW, Holtzapple PG, Dudrick SJ, Cerda JJ: Transport studies, morphological changes and histochemical findings in intestinal epithelial cells following massive bowel resection. Surg Forum 22:361-363, 1971 540

13. Young EA, Weser E: Nutritional adaptation after small bowel resection in rats. J Nutr 104:994-1001, 1974 14. Gray GM: Carbohydrate digestion and absorption: Role of the small intestine. N Engl J Med 292:1225-1230, 1975 15. Peterson SC, Goldner AM, Curran PF: Glycine transport in rabbit ileum. Am J Physiol 219:1027-1032, 1970 16. Wiseman G: Sacs of everted intestine: Technic for study of intestinal absorption in vitro. Methods in Medical Research, Vol. 9. JH Quastel (ed). Chicago, Year Book Medical Publishers, t96t, pp 287-292 I7. Czaky TZ, Esp0sito G: Osmotic swelling of intestinal epithelial cells during active sugar transport. Am J Physiol 217:753-755, 1969 I8. Schultz SG, Fuisz RE, Curran PE: Amino acid and sugar transport in rabbit ileum. J Gen Physiol 49:849-866, 1966 19. Crane RK: Absorption of sugars. Handbook of Physiology Section 6, Alimentary Canal, Intestinal Absorption Vol. 3, CF Code (ed). Washington, American Physiological Society, 1968, pp 1323-t351 20. Nakayama H, Weser E: Adaptation of small bowel after intestinal resection: Increase in the pentose phosphate pathway. Biochim Biophys Acta 279:4t6-432, t972 2 t. I-Iindmarsh JT, Kilby D, Ross B, Wiseman G: Further studies on intestinal active transport during semi-starvation. J Physiol 118:207-218, 1967 22. Rider AK, Schedl HP, Nokes G, Shining S: Small intestinal glucose transport Proximal-distal kinetic gradient. J Gen Physiol 50:1171-1182, 1967 23. Miller DL: Rat small intestine: Development, composition and effects of perfusion. Am J Dig Dis 16:247-254, 1971 24, Esposito G: Intestinal absorption of sugars in semi-starved rats. Proc Soc Exp Biol Med. 125:452-455, 1967 25. Armstrong WMcD, Musselman DL, Reitzug HC: Sodium, potassium and water content of isolated bullfrog small intestinal epithelia, Am J PhysioI 219:t023-1026, 1970 26. Esposito G, Czaky TZ: Extracellular space in the epithelium of rats' small intestine. Am J Physiol 226:50-55, 1974 27. Jackson MJ; Cassidy MM: Epithelial cell swelling during the incubation of a rat small intestine in vitro. Experientia 25:492-493, 1969 28. Schultz SG, Frizzell RA, Nellans HN: Ion transport by mammalian small intestine. Annu Rev Physiol 36:51-91, 1974 29. Goldner AM: Sodium-dependent sugar transport in the intestine. Metabolism 22:649-656, 1973 30. BalTyRJC, Eggenton J, Smyth DH: Sodium pumps in the rat small intestine in relation to hexose transfer and metabolism. J Physiol 204:299-310, 1969 31. Dennhardt R, Haberich FJ: The action of actively transported sugars on the sodium, potassium and fluid transport in the jejunum and ileum of the rat. Pfleugers Arch 345:221236, 1973 32. Humprheys MH, Earley LE: The mechanism of decreased intestinal sodium and water absorption after acute volume expansion in the rat. J Clin Invest 50:2355-2367, 1971 33. Fromm D: Na and C1 transport across isolated proximal small intestine of the rabbit. Am J Physiot 224:1t0-116, 1973 34. Binder HJ: Sodium transport across isolated human jejunum. Gastroenterology 67:231-236, 1974 35. Fordtran JS: Stimulation of active and passive sodium absorption by sugars in the human jejunum. J Clin Invest 55:728-737, 1975 Digestive Diseases, VoL 23, No. 6 (June 1978)