Negative feedback regulation of the ileal bile acid transport system in rodents

GASTROENTEROl.OGY

1993;104:38-46

Negative Feedback Regulation of the Heal Bile Acid Transport System in Rodents JAN LILLIENAU,* LEE R. HAGEY,*

DIANE L. CROMBIE,* JORGE and ALAN F. HOFMANN*

MUNOZ,’

J. LONGMIRE-COOK,”

SARAH

*Department of Physiological Chemistry, Universit) of Lund, Lund, Sweden; *Division of Gastroenterology, Department of Medicine, University of California, San Diego, La Jolla, California; “Department of Medicine, University of Texas at San Antonio, San Antonio, Texas; and “Department of Surgery, University of Louisville, Louisville, Kentucky

Active transport of conjugated bile acids by ileal enterocytes is a key mechanism for conservation of the bile acid pool. Experiments were performed to determine whether such transport is regulated by substrate load. Methods: Using anesthetized biliary fistula guinea pigs or rats, the ileum was perfused with ursodeoxycholyltaurine at a concentration causing maximal ileal transport of this bile acid; absorption was assessed by biliary recovery. Before ileal perfusion, animals ingested one of three diets: chow, chow with added conjugated bile acid, or chow with added cholestyramine. Results: In the guinea pig, ingestion of a taurocholate-enriched diet resulted in a 75% decrease in the absorption rate of ursodeoxycholyltaurine. Similar results were obtained with cholylsarcosine (a deconjugation-dehydroxylation resistant analogue) or with chenodeoxycholylglycine, the endogenous bile acid of the guinea pig. In contrast, cholestyramine ingestion caused an increase in ursodeoxycholyltaurine absorption. In the rat, cholyltaurine or cholylsarcosine ingestion also caused decreased ileal transport. In the guinea pig, maximal down-regulation of active ileal bile acid transport occurred after 2-3 days of bile acid feeding; up-regulation required 3-4 days. Conclusions: Bile acid metabolism is regulated by feedback inhibition of active ileal transport in addition to the well-established feedback inhibition of bile acid biosynthesis in the liver. Together, these two regulatory mechanisms ensure constancy of bile acid secretion. Background:

he

of the terminal ileum actively transport conjugated bile acids from the intestinal T lumen.’ The bile acids are then carried via portal blood to the liver, secreted by the liver into bile, and transported back to the intestine. Ileal conservation of bile acids together with hepatic biosynthesis results in the accumulation of a bile acid pool,* whose circulation is necessary for efficient digestion of dietary lipids3 The ileal transport system for conjugated bile acids is now known to involve an apical bile acid/Na+ cotransporter in the apical membrane’s4 and a bile acid/anion exchange system in the basolateral membrane.’ enterocytes

Transport

processes

of

the

small

intestine

are

known to be influenced by substrate load. Two types of effects have been observed. The first is up-regulation in response to increased load. For example, the transport of glucose’ as well as essential and nonessential amino acids’ is increased by the addition of these nutrients in the diet. The second is upward regulation response to a deficiency ample,

deficiency

state in the organism.

in

For ex-

of body calcium’ or iron” greatly

increases the transport activity of the intestinal carriers for these minerals. Bile acids are the end products of cholesterol metabolism.* Regulation of bile acid biosynthesis from cholesterol by the flux of bile acids through the hepatocyte was described some years ago by Bergstrom ielsson”;

the properties

comprehensively

of this regulation

reviewed recently.”

and Danhave been

This regulation

was shown by Shefer et al.‘* to be mediated by changes in the activity of cholesterol 7&hydroxylase, which is considered to be the rate-limiting enzyme.‘3,‘4 Regulation of the activity of this enzyme has recently been shown to be at a transcriptional level.14 We hypothesized

that active ileal bile acid transport

might also be regulated by substrate load. Here, we report experiments that indicate ileal transport of conjugated bile acids is down-regulated by increased substrate load and up-regulated by decreased substrate load. Experiments were also performed to determine the time necessary for up- and down-regulation.

Materials and Methods Experimental

Design

The experimental design featured measurement of the rate of ileal absorption of a nontoxic conjugated bile acid Abbreviations used in this paper: CDCA, chenodeoxycholic acid; CDC-gly, chenodeoxycholylglycine; HP& high performance liquid chromatography; UDCA, ursodeoxycholic acid; UDC-tau, ursodeoxycholyltaurine. 0 1993 by the American Gastroenterological Association 0016~6085/93/$3.00

January 1993

NEGATIVE FEEDBACK REGULATION OF ACTIVE ILEAL BILE ACID TRANSPORT

[ursodeoxycholyltaurine concentration control),

(UDC-tau)]

after

long-term

after long-term

feeding

gated bile acids (to increase port system),

feeding

system).

conjugated

this compound

was observed

(to decrease

additional

studies

were

synthetic

conjugated

with chenodeoxycholylglycine this conjugated

Three

received

deconju-

or CDC-gly

a result,

ing 250-330

with

cholylsarcosine,

(CDC-gly)

a

in the guinea

bile acid is the predominant

biliary

was measured

was equated Because

with recovery

animals

of the perfused

each animal

rate of UDC-tau:

submaximal

of the perfu-

Details

and

proce-

MO) and used without

its

high-performance acid

were gifts from and UDCA scribed

(UDCA)

Diamalt

with

et al. l9 The

by column

mixtures

silica

chromatography.”

were synthesized

let al.” and purified

as de-

bile acids

using

(24-[14C]cholyltaurine)

[‘4C]UDC-tau)

CDCA

or taurine

and

according

wt/wt

was made by soaking 97% ethanol

pellets

of

for 5 minutes.

dried at room temperature centration in the ethanol

After

soaking,

the pellets

in were

overnight. (The bile acid consolution was 100 mmol/L for

pig chow and 130 mmol/L for rat chow, because pig chow absorbed more of the ethanol solution per

weight

chow

chow containing additional guinea

the rat chow). 0.5% wt/wt pig studies.

Cholestyramine-enriched

(Guinea

Pig Diet

006; Wayne,

Using

CDC-gly

was prepared

were performed

Animal

and

Medical

weight

the study.

ad libitum,

Subjects

in each group).

cages (one per cage) Center

of the animals

The animals

were

and the lighting

(light period

from 7

Committee,

AM

to 7

condi-

PM).

of California,

protocol

This San

numbers

053-

by intramuscular

in-

Heal Perfusion jection

with

vis, Morris

pigs were anesthetized

ketamine Plains,

Health

mg/kg;

HCl, 40 mg/kg

NJ), xylazine

Co., Kansas anesthesia

(Ketalar;

Parke-Da-

HCl, 5 mg/kg

(Fermenta

City, MO), and atropine

was maintained

once every hour.

After

laparotomy,

the

the common

The cystic duct was ligated and the

bile collected

and an incision

sulfate,

by repeating

for subsequent

analysis

low). The ileum was ligated as close as possible was made in the antimesenteric

(see be-

to the cecum, border.

The

entire small intestine was rinsed, and a tube was inserted through the incision in the distal end of the cecum. A small incision was made 30 cm proximal to the ileocecal valve, and a tube for infusion The

for

similar.

Guinea pig chow Madison, WI) or rat chow chow.

studies

by the University

was approved

pigs (n = 18)

diet for 7 days were

study

a

this procedure,

water

in

per group).

Guinea

tions were controlled

gallbladder

guinea guinea

than

daily during

to drink

bile duct was cannulated.

pig or rat chow solution

were performed

San Diego

intake

pigs (n =

to a diet con-

animals

were kept in metabolic

monitored

allowed

Diego

Guinea

studies

diet. Perfusion

Food

Animal

or cholylsarcosine

in a cholyltaurine

with

at days 1, 2, 3, or 4 (4 animals

Guinea

(24-

chromatography.

Guinea

chow.

cholyltaurine

were performed

on a cholyltaurine-enriched

quarters.

injections 0.5%

Perfusion

animal

0.05

Bile acid-enriched

mg/

of down-regulation.

of California,

Labeled

Diet containing

experiments

day) for

(270-370

was added to the diet at a concentra-

in the University

gel and

to the method

by thin-layer

mg/kg.

- day

mmol/kg

of to a

9R and 055-9R.

were

UDC-tau

with

corresponding

on a chow diet were switched

to a chow

and were at least 98% pure

and by thin-layer

cholyltaurine

glycine

conjugated

and grade

Germany).

rats, weigh-

chow enriched

for 7 days. The amount

Time course of up-regulation.

were

chromatography

pharmaceutical

chromatography

chloroform-methanol by HPLC’s

of

purification;

acid (CDCA)

AG (Raubling,

were conjugated

by Tserng

purified

Tserng

liquid

was 98%. Chenodeoxycholic

ursodeoxycholic

Sigma Chemical

further

MA)

cholylsarcosine,

day (240-270

0.5-0.7

course

All animals from

received

at days 1, 2, 3, 4, 5, or 6 (3-4

switched

Co. (St. Louis, by

mmol/kg*

cholyltaurine.

animals

maintained

previously.”

was purchased

River),

which

Time

in animals

Cholyltaurine

Wilmington,

was 175 ltmol/day,

and

24) maintained

one at

of the perfusion

were followed. Guinea

River,

or cholylsarcosine

pigs

taining

for the

Bile Acids

purity

guinea

regimens

tion of 2% by weight.

bile acid in bile.

one at maximal

concentrations.

dure have been reported

(HPLC)”

cholyltaurine

pig or rat and

gave only two values

MO) was added.

with cholyltaurine,

kg. day) for rats. Similar

and submaximal

fistula guinea

were killed at the completion

sion procedure, absorption

at maximal

biliary

Madison, cholestyr-

load on bile acid transport.

g (Charles

g (Charles

cholestyramine,

rates in the anesthetized

Premier,

and 2% wt/wt

for at least 7 days. Sprague-Dawley

dose of 0.4-0.5

pig,

feeding

350-450

bile acid ingested

bile acid in this species. Absorption

different

chow enriched

pig.

As

Teklad

Co., St. Louis,

Effect of substrate

pigs weighing

However,

8604;

Dietary Regimens

were perwas selected

BLOX

to a fine powder,

(Sigma Chemical

in

analogue resistant to deconjuStudies were also performed

gation-dehydroxylation.‘5p’6 because

amine

extensive

guinea

performed

bile-acid

Rodent

substrate

Experiments

to undergo

in the

WI) was ground

in conju-

bile acid for feeding.

gation-dehydroxylation

(Wayne

diet (as

of a diet enriched

on the guinea pig and rat. Cholyltaurine

as a prototypic

luminal

load to the ileal trans-

a bile acid sequestrant

load to the ileal transport formed

of a chow

of a diet enriched

substrate

or after long-term

cholestyramine,

at a defined

feeding

39

surgical

was inserted

procedure

Anesthesia

performed

was introduced

into the Real lumen. in the rat was quite by an intraperitoneal

injection of pentobarbital sodium (70 mg/kg, Abbott Laboratories, Chicago, IL); anesthesia tained

by a continuous

intravenous

infusion

Nembutal; was mainof 0.1 mg/

40

LILLIENAU

GASTROENTEROLOGY

ET AL.

min - kg pentobarbital ileum

solution.

The small

was made 20 cm proximal

The temperature the body 38°C

of the perfusate

temperature

by means

animals

was maintained controlled

to a rectal temperature

Jr. model

RHSY;

Fluid

NY) was used to perfuse

The morphology and

probe.

Metering

at

heating

A pump

(Lab

Oyster

Bay,

Inc.,

the ileum at a flow rate of 4 mL/

min. minutes,

without

followed

solution.

bile acids was used during by 1 hour of a 5 mmol/L

tion and during

a second

UDC-tau

this conjugated

hour

UDC-tau

of a 0.5 mmol/L

bile acid is absorbed

the ileum.

UDC-tau

from this laboratory).

appreciable

passive

is extremely

A 5 mmol/L

was used because maximal

transport

in the rat” and in the guinea this laboratory);

a 0.5 mmol/L

transport

under

nonsaturating

lected

at IO-minute

collections three)

receiving

cholestyramine-enriched

bedded

major

absorption by

at 4 mmol/L

solution.

LJDC-tau

buffer of 130 mmol/L

calcium

chloride

methyl)

aminomethane

St. Louis,

(dihydrate)

were

Bile

MO)

adjusted

before

this was always
using

in an

was

Radioactivity

ity and mass. The presence

tested

in the results

results

from

animals with

or control

receiving

results

from

the

signifi-

Because there with

individual

bile acids animals

were

receiving

animals.

Effect of Regimen on Heal Transport Down-regulation. 1. In guinea

decreased

ileal

was perfused

pigs,

Data

are summarized

addition

transport

at a luminal

of bile acids

of UDC-tau concentration

in Figto the diet

when the ileum of 5 mmol/L,

6

I t v CDCG ??CT ?? CS

was calculated;

size filter;

and the ab-

8

of dehy-

the perfusate

to

:

bile acid radioactiv-

of only UDC-tau

between

for statistical

Results

in the

using a 3-hydroxysteroid

uses was confirmed

knowledge

+- SD. Differences

were

and the

based on measurement permitted

em-

tris(hydroxyCo.,

times, thus conserving

chow

2 mmol/L

8. 24-[‘4C]UDC-tau

pore

assay. 24 This procedure

after multiple

without

After each study, the used perfusate

a 0.45 pm

the bile acid concentration be used multiple

UDC-tau

was dissolved

groups

and compared

(usually the last

and after perfusion

bile acids were replaced,

drogenase

microscopically

procedure.

was col-

buffered

of bile acids lost from the perfusate

injury animals

with H&E. Coded

obtained

ileal

base; Sigma Chemical

(100 Bq/mL).

was measured

amount

to pH

treatment

of the three

chloride,

and stained

were no differences

also

to measure

and 25 mmol/L

(Trizma

to the perfusate

perfusate

sodium

from

chow or unaltered

cance using an analysis of variance

ure

(0.5 or 5.0 mmol/L)

isotonic

mucosal

tissues

regimen.

cholestyramine

experiments

experiments

of an isotonic

sectioned,

All values are mean

trans-

were used for calculations. consisted

possible

regimen;

was ex-

Statistics

is reached

bile acid output

chow,

The tissue was fixed in formalin,

were examined

of the dietary

pooled

and the average

the dietary

in paraffin,

bile acid concentration

solution

to detect

from

were also examined.

enriched

because

observations

conditions.

intervals;

with the highest

The perfusate

sorbed

microscopically

resulting

bile acids,

in addition,

with

of the distal ileum from guinea pigs

or cholylsarcosine

solu-

well transported

pig (unpublished

performed

added

amined

UDC-tau

solely by active

the liver of the rat23 or guinea pig (unpublished

from

the first 40

was used to assess ileal transport

port; its low pKaz2 precludes from

fed cholyltaurine

sections

Perfusate

No. 1

Light Microscopy

in the

valve.

was kept at 38°C

of a thermostatically

lamp connected Pump

of the

incision

to the ileocecal

Vol. 104,

0

in the perfusate

repeatedly

by thin-layer

chromatography.

Analytical

Methods

Bile volume

was measured

radioactivity

output

gravimetrically.

in bile was determined

UDC-tau

by adding

50-

200 l.rL bile to 10 mL scintillation cocktail (ScintiVerse BD; Fisher Scientific, Tustin, CA). Radioactivity was measured using

a liquid

scintillation

counter

CA) and divided by the specific tau to give mass.

activity

(Beckman,

Fullerton,

of the infused

HPLC was used to determine the percentage conjugated bile acids in guinea pig gallbladder

UDC-

of different bile.‘s

0'

I

I

cholastymmlne

control

Dietary

I

I

con]qatad bile acids

Regimen

Figure 1. Transport rate of UDC-taurine (5.0 mmol/L) by the perfused guinea pig ileum after feedtng chow enriched with cholestyramine. unaltered chow, or chow enriched with conjugated bile acids. Each point represents data from a single perfusion. Bile acids used are indicated.

NEGATIVE FEEDBACK REGULATION OF ACTIVE ILEAL BILE ACID TRANSPORT

January 1993

Table 1. Effect of Feeding Conjugated

Bile Acids or Cholestyramine on Active Intestinal Transport of UDC-tau in the Guinea Pig

compared

Transport of UDC-tau by perfused Ileum; concentration (pno//mins kg) 5.0 mmol/L

Regimen

0.74 1.34 1.40 4.26 2.95

Control

+ + + + +

0.26 0.72 0.90 0.97 0.68

However,

animals UDC-tau

0.15 0.31 0.36 0.60 0.55

* ? t * +

0.04 0.19 0.24 0.14 0.18

(4) (3) (2) (4) (5)

(for

were no differences

a possible

(Table

showed

feeding.

(P < 0.05).

decreased tions

ileal transport

that causing maximal transof cholyltaurine to the diet under

by 75%. Enrichment

sine caused from

exceeding Addition

a similar

the perfused

endogenous

served in animals

of CDC-gly,

transport.

pig,

When

the

also caused

the lower

in UDC-tau

receiving

time a constant

level of transport

was reached

Animals

chow diet after ingesting for 1 week. Figure for active

were

sorption marized

between in Table

For the lower

was no significant

UDC-

or cholylsarco-

experimental 2.

of the effect was concentration difference

groups.

to

enriched

a

diet

3 shows that 3-4 days were required

ileal bile acid transport

to return

to control

Effect of Bile Acid Feeding on Biliary Bile Acid Composition Cholyltaurine.

In the guinea

deoxycholic

there

changed

a cholyltaurine

biotransformation

less (34%) (P < 0.05).

An

was ob-

2).

lyltaurine

UDC-tau,

feeding.

transport

for 3-7 days,

gates of bacterial

diet, but the magnitude

(P < 0.05) oc-

cholyltaurine

dietary groups. Data are summarized in Table 1. In rats, a similar decrease in ileal transport of UDCtau at a saturating concentration was observed after of a cholyltaurine-enriched

this

to cholyltaurine

2 days of cholyltaurine

cholyltaurine

ingestion

pig because

response

tau concentration (0.5 mmol/L) was infused, its rate of transport was not significantly different between

sine-enriched

of the negaon active ileal

values.

(55%) in ileal transport Feeding

acid of the guinea

UDC-tau

condi-

of the diet in cholylsarco-

decrease segment.

bile

decreased

these saturating

in the guinea

reduction

at which

Inhibition

bile acid transport

after only

additional

from

experimental

The time course

a greater

Decreased

curred

Discussion).

transport

caused by bile acid feeding

was studied

species

not ani-

2).

Up-regulation. a concentration

see

for the three

Down-regulation. tive feedback

did

the control

in UDC-tau

Time Course of Feedback

(Figure

port

than

explanation,

solution

(P < 0.05).

cholestyramine

differently

There

transport NOTE. Data are mean + SD. The number of experimental animals in each group is given in parentheses. By analysis of variance for the 5 mmol/L UDC-tau perfusate, the difference between bile acid fed animals and control animals was significant (P < 0.05). Animals receiving cholestyramine had a greater maximal transport rate than either the bile acid or the control group (P < 0.05). For the dilute perfusate (0.5 mmol/L UDC-tau), differences between the groups were not significant.

receiving

mals

the 0.5 mmol/L

0.5 mmol/L

(4) (4) (2) (4) (5)

the bile acid fed animals

transport

groups

Bile acid feeding Cholyltaurine Cholylsarcosrne CDC-gly Cholestyramine feeding

with

41

of

caused a marked

acid composition. (mostly

pig, the feeding

alteration

Bile became

enriched

in biliary

of bile

in the conju-

products

of cho-

acid and its 3 oxo- and

Table 2. Effect of Feeding Conjugated

Bile Acids or Cholestyramine on Active Intestinal Transport of UDC-tau in the Rat

in abTransport of UDC-tau by perfused ileum: concentration (umol/min . kg)

Data are sum-

Up-regulation. In guinea pigs, the feeding of a diet containing 2% cholestyramine caused a 44% increase in maximal ileal transport compared with the control animals (P < 0.05). When compared with the animals receiving bile acids in their diet, the increase in maximal ileal transport was 284% (P < 0.05). By analysis of variance, the addition of cholestyramine to the diet did not influence UDC-tau transport when the ileum was perfused with the lower concentration bile acid solution (Table 1). In rats, the addition of cholestyramine to the diet caused a 47% increase in maximal UDC-transport as

Regimen Bile acid feeding Cholyltaurine Cholylsarcosine Cholestyramine feeding Control

5.0 mmol/L

2.58 2.07 3.50 3.60

f f * f

0.28 0.16 0.59 0.53

0.5 mmol/L

(6) (4) (7) (6)

0.42 0.51 0.43 0.53

+ 0.08 + 0.28 I? 0.12 t 0.04

(6) (4) (7) (5)

NOTE. Data represent mean -c SD. The number of experimental animals in each group is grven in parentheses. By analysis of variance, for the 5 mmol/L UDC-tau solution, animals in the bile acid group had a lower maximal transport rate than animals in the control group (P < 0.05). Animals in the cholestyramine-fed group had a greater rate of maximal transport than the bile acid-fed group (P < 0.05) but not the control group (not significant). None of the differences for transport rate were significant for the 0.5 mmol/L UDC-tau perfusate.

42

LILLIENAU

ET AL.

GASTROENTEROLOGY

Data are expressed deoxycholic

as 7-deoxy

acid as well as its 3- and these are derived

cholyltaurine.

After

to normal.

occurred

much

and by 4 days, it had not yet

more rapidly,

lation

was not

bile acid composition.

influenced

taurine 2

Number

3

4

5

6

7

of Days with CT Feadlng

Figme2.

Time course of down-regulatron of active ileal bile acid transport. Transport of UDC-taurine (5.0 mmol/L) by the perfused guinea pig ileum l-7 days after animals were changed to a cholyltaurineenriched diet (Z 175 pmol/day). Animals fed regular chow served as controls. Statistical differences relative to animals fed cholyltaurine for 1 week are Indicated by *P < 0.05 and ***P < 0.001 (mean + SD).

feeding

by the induced

reported was stopped,

for cholylsarcosine,

averaging

CDC-gly

in biliary

in one animal,

occurred

ther

4 shows the time course

ary bile acid composition

of regression

to its pretreatment

are

enrich-

and rabbits.” to two

guinea

bile acids increased

con-

data are given

in Table

in3.

Histology of the Ileum By light microscopy, ileal morphology, indicating

Figure

Data

and its 7-0~0 derivative

Mean

marized in Table 3. The increased proportion of bacterial biotransformation products of cholyltaurine rapidly.

to its

bile acids in cholylsar-

was administered

in the other.

cholyl-

administra-

to cause modest

30%, of biliary

pigs, its proportion

12-0~0 derivatives). Changes in biliary bile acid composition after 1 week of feeding cholyltaurine are sum-

because

at least in rats, hamsters,

When

bile

When

(no data shown).

tion of this bile acid is known

creased

in

in the

bile slowly reverted

composition

cosine,

change

an increase

previously.‘5

not presented

siderably

extremely

caused

pretreatment

ment,

that up-regu-

of this bile acid in the administered

acid as has been 1

of ileal transport

suggesting

In the rat, cholyltaurine

Chow Only

deriva-

bile acid composi-

Up-regulation

biliary

proportion

12-0~0

of

solely from exogenous

3 days, biliary

little change

returned

No. 1

bile acids (conjugates

tives) because tion showed

Vol. 104,

cholestyramine

nor

there were no changes in that in the guinea pig neiconjugated

bile acid caused

of bilipattern.

loo I

0 Chow Only

D

1

Days with Reg.

2

3

Chow after

4 CT feodlng

1 Number

2

3

of days after corsatlon cholyltaurln~ feadlng

4 of

Figure 4. Time course of changes in proportion of 7-deoxy bile acids Figure 3. Time course of up-regulation

of active ileal bile acid transport. Transport of UDC-taurine (5.0 mmol/L) by the perfused guinea pig ileum 0, 1, 2, 3, or 4 days after animals changed to a chow diet from a diet enriched in cholyltaurine (= 175 pmol/day). Animals fed only regular chow served as control. Statistical differences relative to control animals (chow only) are indicated by *P < 0.05, **P < 0.0 1, and ***P < 0.001 (mean + SD).

(deoxycholic acid and its 3-0~0 and 12.0~0 derivatives) in biliary bile acids in relation to ileal transport of UDC-taurine (5.0 mmol/L) in animals who were changed to a chow diet after 1 week of ingesting a diet enriched in cholyltaurine. Increased bile acid transport occurred as early as day 1, whereas a decrease in the proportion of 7-deoxy bile acids was not observed until day 4. Points shown are the mean of 4 animals for each time point.

January 1993

NEGATIVE

Table 3. Bile Acid Composition

FEEDBACK

REGULATION

OF ACTIVE

ILEAL

BILE ACID

(%) of Gallbladder Bile From Guinea Pig After Ingestion of Cholyltaurine Dietary

Bile acid classa

Control

Chenodeoxycholic acid and its biotransformation products Chenodeoxycholyl conjugates” 3a-hydroxy-7-oxo-5P-cholanoyl conjugatesD Ursodeoxycholylglycine Cholic acid and its biotransformation products Cholylglycine Deoxycholylglycine 3-0x0- 12a-hydroxy-5P-cholanoyl conjugatesb 3a-hydroxy- 12.oxo-5P-cholanoyl conjugate@

53.9

TRANSPORT

43

or CDC-gly

regimen

Cholyltaurine

Chenodeoxycholylglycine

+ 9.0

6.9 + 2.6

45.5

t

16.8

38.0 + 7.6 5.9 k 1.7

a.1 * 7.3

36.6 + 20.7 3.4 * 2.0

0.4 k 0.5 1.9 f 0.6

7.0 f 2.6 31.4 + 7.9

1.2 ?I 0.0 4.9 + 2.9

36.0

+ 2.0

6.7 t 3.0

10.6

f 2.7

1.6 f 0.1

NOTE. Mean + SD. aDetermined by HPLC.‘* Ammals had ingested the bile acid for 2 days. %onjugates means glycine or taurine aminoacyl amidates.

any detectable damage to epithelial cells. Animals failed to gain weight while receiving cholyltaurine or

been shown to occur during cholic acid feeding in the

CDC-gly,

ileal absorption in bile acid conservation,

but showed normal

ceiving cholylsarcosine. induced diarrhea.

weight gain when re-

None of the dietary regimens

experimental

evidence

is regulated by lu-

minal signals. The regulation had the characteristics inhibition,

by passive mechanisms.‘,27 which is resistant to de-

conjugation-dehydroxylation15

studies provide

being decreased

of

when substrate

ity, permitted

and is devoid of toxic-

down-regulation

acids to be shown convincingly

by administered

bile

in both rodent species.

Cholyltaurine feeding greatly changed the bile acid profile in guinea pig gallbladder bile causing an in-

load was increased and increased when substrate load was decreased. The effect was much greater in the

creased proportion

guinea pig than the rat, indicating

fluenced ileal bile acid transport,

that the magnitude

because most

bile acids are rapidly absorbed

from the small intestine

that maximal active ileal transport feedback

natural, unconjugated

decreases the need for active

The use of cholylsarcosine,

Discussion These

rat.” Such deconjugation

derivatives.

of deoxycholylglycine

Although

and its 0x0

this change could also have inwe think it did not

of the effect varied between species. The experiments reported here as well as previous studies by many labo-

play a major role in the induced regulatory changes for three reasons. The first is that maximum bile acid

ratories indicate that the response of an animal to an

transport resumed its normal capacity after 4 days, at which time the bile acid profile still was very different

interruption of the enterohepatic circulation of bile acids at a level proximal to the ileum is followed by two compensatory

responses:

an increase

in hepatic

biosynthesis of bile acids (reviewed in reference 11) and an increase in active ileal transport of conjugated bile acids, as described here. Cholyltaurine

was selected as a prototypic

bile acid,

but the striking enrichment of biliary bile acids in deoxycholic acid and its 3- and 12-0~0 derivatives indicates that this compound underwent extensive dehydroxylation-deconjugation in the guinea pig. In the rat, cholyltaurine has recently been found to also undergo extensive deconjugatictl-dehydroxylationzs but this effect is less obvious because of an adaptive increase in the 7-rehydroxylation of deoxycholic acid that has

from control animals. The second is that down-regulation was induced by cholylsarcosine, which does not undergo deconjugation-dehydroxylation.15 The third is that administration of CDC-gly also decreased maximum active bile acid transport in the guinea pig, and the only change in biliary bile acid composition was a modest increase in those bile acids already present. It took about 3 days to down- or up-regulate the bile acid transport capacity in the guinea pig, which corresponds very well with the turnover time of enterocytes in rodents. 28 A slightly longer time was required for up-regulation than down-regulation, possibly because in our experiments the stimulus for down-regulation (bile acid feeding) was presented to the ileum immedi-

44

LILLIENAU

ately.

In contrast,

pletion

the stimulus

this

whether

study,

regulatory

changes

or whether

tion might change

likely

because in addition,

era1 level

might

concentrations

bile acid transport et a1.31 They

fistula,

bile acid secretion

has

been

to high

bile acids. Such intracel-

experiments showed whole

with

increased intestine

by

of

50 years ago by Bera biliary

to be homeostati-

the rhesus transport

Redinger monkey,

et these

biliary

drainage

administra-

of UDCA.42-44 Pa-

who ingest cholestyrcholesterol levels might

ileal transport minor

par-

by duodenal

39 ileal bypass, 4o cholestyramine

the relatively

ex-

ileal transport

of bile acids; this could

changes

olism induced

by cholestyramine

spite its potent

bile acid binding

in bile acid metab-

administration,45

de-

effect in the intestinal

The data reported tion for other acid

here may also provide

observations

metabolism

in

CDCA

or UDCA

creased

biliary

on induced

man.

Oral

to gallstone

an explana-

changes

in bile

administration

patients

bile acid secretion.47-52

of

causes little inWe suggest

a1.32 In

is because ileal transport down-regulates in response the increased substrate load resulting from secretion

workers

the conjugates

of total bile acids from the

into bile after 18 hours

decrease

would

from thera-

lumen.46

inhibition

that in dogs with

to be hepatotoxic

as well as the administration

explain

and

bile acids. These include

diversion,38

well up-regulate

at the basolat-

cause cell damage.30

published

cytotoxic

No. 1

as CDCA

may benefit

that should

active

level, a

tally controlled by intestinal absorption. Further indirect evidence for up-regulation of ileal bile acid transport

tion4’

patients

involves

enterocyte

appeared

tial biliary

such

are known

seems

transport

was reported

procedures

Vol. 104.

37). This possibility

transporter

in favor of feedback showed

peutic

acids

tients with hypercholesterolemia amine to lower their plasma

down-regulation the ileal

acid, which

why cholestatic

intubation,

ileal

bile

in reference

of endogenous,

al-

up-regula-

at a cellular

of the apical

might

evidence

of the ileum

of more proximal

of conjugated

lular accumulation

from altered trans-

cells of the terminal

occurred

expose

dihydroxy

deoxycholic plain

determine

For example,

apical

transport;

to

region

recruitment

If regulation

in the activity

Indirect

villous

capacity.

involve

enterocytes.

resulted

an entire

its transport

most

of bile acid

dogenous (reviewed

it is impossible

rates by individual

ileum’” tered

(de-

was more gradual.

From

man

for up-regulation

of the bile acid pool by cessation

feeding)

port

GASTROENTEROLOGY

ET AL.

but not after 6

of the administered

the data in the present

paper

tion for the well-documented

to of

bile acid. Finally,

also provide inverse

this

an explana-

relationship

be-

hours of diversion of bile acids from the intestine. Contrary to our interpretation of these data, van Til-

tween

burg et al. 33 very recently suggested that increased bile acid concentration in the distal ileum causes increased

In conclusion, these studies show for the first time that maximal active ileal bile acid transport is under

sodium

dependent

uptake

of bile

acids,

while

de-

creased bile acid concentration in the distal ileum causes decreased bile acid absorption in humans.

In the adult human, concentrations of conjugated bile acids in the terminal ileum have been reported to be 1-2 mmol/L,34p35 a concentration reto

conjugated

be

close

to

that

bile acid transport

causing

saturation

in the perfused

of cy-

of the bile acid po01.~~*~~

negative

feedback

control

in two rodent

study also shows that this regulation requires

three days to down-

species.

is rather

or up-regulate.

The

slow and Thus,

the

constancy of hepatic bile acid secretion during digestion in at least these two rodent species has at least two

Physiological and Clinical Implications

ported

cling

the bile acid pool size and the frequency

of

levels of control: formed bile acids

(a) regulation of input of newly by feedback inhibition of hepatic

biosynthesis and (b) regulation of intestinal conservation by feedback inhibition of ileal transport.

human

ileum.36 In the fetus and newborn animal with a relatively sterile small intestine, downward regulation of active transport would serve to keep the bile acid pool from expanding indefinitely once biliary bile acid secretion had begun. The opposite situation may also be envisioned. Were the bile acid pool to be depleted by, for example, an episode of viral enteritis, the ileal transport system should up-regulate and increase its efficiency of conservation of secreted bile acids until the bile acid pool had been restored. From a clinical standpoint, patients with cholestatic liver disease are likely to inappropriately conserve en-

References Wilson, FA. Intestinal absorption of bile acids. In: SG Schultz, M Field, RA Frizzell, BB Rauner, eds. The Handbook of Physiology. The Gastrointestinal System: Intestinal Absorption and Secretion. Bethesda: American Physiological Society, 1991:389-409. Hofmann AF. Enterohepatic circulation of bile acids. In: Schultz SG, ed. Handbook of Physiology. Section on the Gastrointestinal System. Bethesda: American Physiological Society 1989:567596. Borgstrom B, Barrowman JR, Lindstrom M. Role of bile acids in intestinal lipid digestion in absorption. In: Danielsson H, Sjbvall J, eds. Sterols and Bile Acids. Amsterdam: Elsevier, 1985:405425. Lueke H, Stange G, Kinne R, Murer H. Taurocholate-sodium

co-

January 1993

NEGATIVE FEEDBACK REGULATION OF ACTIVE ILEAL BILE ACID TRANSPORT

transport by brush border membrane vesicles isolated from rat ileum. Biochem J 1978; 174:95 I-958. 5. Weinberg SL, Burckhardt G, and Wilson FA. Taurocholate transport by rat intestinal basolateral membrane vesicles. Evidence for the presence of an anion exchange transport system. J Clin Invest 1986;78:44-50. 6. Solberg DH, Diamond JM. Comparison of different dietary sugars as inducers of intestinal sugar transporters. Am J Physiol 1987;252:G574-G584. 7. Karasov WH, Solberg DH, Diamond JM. Dependence of intestinal amino acid uptake on dietary protein or amino acid levels. Am J Physrol 1987;252:G614-G625. 8. Nellans HN, Kimberg DV. Cellular and paracellular calcium transport in rat ileum: effect of dietary calcium. Am J Physiol 1978;235:E726-E737. 9. Wheby MS, Jones LG, Crosby WH. Studies on iron absorption. Intestinal regulatory mechanisms. J Clin Invest 1964;43: 14331442. 10. Bergstrom S, Danielsson H. On the regulation of bile acid formation In the rat liver. Acta Physiol Stand 1958;43: 1. 1 1. Vlahcevic ZR, Heuman DM, Hylemon PB. Regulation of bile acid synthesis. Hepatology 199 1; 13:590-600. Shefer S, Hauser S, Bekersky I, Mosbach EH. Feedback regulation of bile acid biosynthesis in the rat. J Lipid Res 1969; 10:646-655. 13. Kwekkeboom J, Princen HMG, van Voorthuizen ME, Kempen HJM. Bile acids exert negative feedback control on bile acid biosynthesis in cultured pig hepatocytes by suppression of cholesterol 7ohydroxylase activity. Hepatology 1990; 12: 1209- 12 15. 14. Pandak WM, Li YC, Chiang JYL, Studer EJ, Gurley EC, Heuman DM, Vlahcevic ZR, Hylemon PB. Regulation of cholesterol 7a-hydroxylase mRNA and transcriptional activity by taurocholate and cholesterol in the chronic biliary diverted rat. J Biol Chem 1991;266:3416-3421. 12.

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values for taurine-conjugated bile salts In rats and hamsters. Am J Physiol 1986;25 1:G852-G858. 24. Turley SD, Dietschy JM. Re-evaluation of the 3a-hydroxy-steroid dehydrogenase assay for total bile acids in bile. J Lipid Res 1978; 19:924-928. 25. Zhang R, Barnes S, Diasio RB. Differentral intestinal deconjugation of taurine and glycine bile acid N-acyl amidates in rats. Am J Physiol 1992;262:G35 1-G358. 26. Oda H, Kuroki S, Yamashita H, Nakayama F. Effects of bile acid feeding on hepatic deoxycholate 7a-hydroxylase activity in the hamster. Lipids 1990;25:706-710. 27. Schiff ER, Small NC, Dietschy JM. Characterization of the kinetics of the passive and active transport mechanisms for bile acid absorption in the small intestine and colon of the rat. J Clin Invest 1972;51:1351-1362. 28. Messier B, Leblond CP. Cell proliferation and migration as revealed by radioautography after injection of thymidine-H3 into rats and mice. Am J Anat 1960; 106:247-294. 29.

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Received July 22, 1991. Accepted July 21, 1992. Address correspondence and reprint requests to: Alan F. Hofmann, M.D., Ph.D., Division of Gastroenterology, 0813, University of California, San Diego, 9500 Gilman Drive, La Jolla, California 92093-08 13. Lee R. Hagey was a predoctoral student in UCSD’s Biomedical Science Graduate Program and was supported by National Institutes of Health Training Grant HL07212. Work at UCSD was supported in part by grants-in-aid from the Falk Foundation, e.V., Germany; Burroughs-Wellcome Co., Research Triangle Park, NC; Diamalt AG, Germany; and grants DK 21506 and DK 32130 from the National Institutes of Health (PI: Dr. E. W. Moore, Medical College of Virginia, Richmond, VA). Dr. Hofmann acknowledges support from the Alexander von Humboldt Foundation. Parts of this work have been published in abstract form (Gastroenterology 1991;100:A832; Bile acids as therapeutic agents. From basic science to clinical practice. Boston: Kluwer Academic, 1991: 163-165). The authors wish to acknowledge fruitful discussions with Dr. Claudio Schteingart, statistical analyses by Dr. Joseph Steinbach, and syntheses of bile acids by Huong-Thu Ton-Nu. They also thank Vicky Huebner for preparing the manuscript.