Hormonal effects on the development changes of mouse small intestinal glycolipids

Vol. 119, No. 3, 1984

BIOCHEMICAL

AND BIOPHYSICAL RESEARCH COMMUNICATIONS

March 30, 1984

Pages 1168-1173

Hormonal

Ciji

Effects Small

Sato,

Michio

on the Intestinal

Fujie, and

Department Hamamatsu

Received

February

of

13,

Development Glycolipids

Changes +

Tadayoshi Uezato Kenji Nishimura

of

Mouse

, Michiya

+

Fujita +

Biology and Department of Biochemistry University School of Medicine, Handa-cho 3600, Hamamatsu 431-31, Japan

,

1984

Summary: The composition of intestinal qlycosphingolipids during normal and hormone-perturbed development was investigated. The concentrations of glycosphingolipids of mouse small intestine were affected by the injection of thyroxine or cortisone during sucklinq and weaning periods. GDla was reduced by the hormonal treatment among major qanqliosides, GM3, GM1 and GDla, of mouse small intestine during the suckling period. In contrast, asialo GM1 was precociously produced by the treatment,which scarecely found in control suckling mouse small intestine. The results showed that these hormones were related to developmental alteration of small-intestinal qlycolipids.

There control

is

of

that

work

thyroid

of

a precocious

mucosal

cells

developmental

cells

have

not

small precocious

MATERIALS

that

far

during

hormones in

the

developmental

alteration

in

maturation (1). early

present

effects

of

the

qlycosphingolipids

hormones

on

of

these

contents previously

demonstrated pattern

the composition

known

weeks

small-intestinal

reports

under

well

of

qlycosphingolipids work

is

pattern

We have in

It

is

postnatal

glycosphingolipids

examined. changes

The

enzyme possible

of

been

(5).

intestinal

these

However,

so

the

hormones

alteration

intestine

show

glucocorticoid

change

(2-4).

the

normal

to

and

administration

causes

the

much

in

mouse

hormone-induced of

the

same

organ.

AND METHODS

ICR mice were used without regard to sex. One litter comprising at least 11 animals was divided into two groups. At 6 d of age one half of the litter received intraperitoneal injection of 50 pg/g body wt/d of cortisone 21-acetate in 0.9% NaCl for 3-4 successive days or 2-3 pg of DL-thyroxine/g body wt/d for successive days until they were killed. The other half served as controls. Animals were killed either at 14 or 28 d of age. Small intestines were throughly washed with cold 0.9% NaCl before extraction of lipids with 20 vol. of chloroform/methanol (2/l,v/v). Acidic and neutral glycolipids were isolated with the method described 0006-291X/84 $1.50 Copyright 0 1984 by Academic Press, Inc. All rights of reproduction in any form reserved.

1168

Vol. 119, No. 3, 1984

BIOCHEMICAL

AND BIOPHYSICAL RESEARCH COMMUNICATIONS

previously (5) which was partly based on that of Saito and Hakomori (6). The neutral glycolipids were eluted from a column (1.2 x 5 cm) of DEAF Sephadex A-25 (acetate form) with 150 ml of chloroform/methanol/water (30/60/8,v/v/v) and the acidic glycolipids with 50 ml of chloroform/ methanol/0.8M sodium acetate(30/60/8,v/v/v). The neutral glycolipids Wrji“? freed of phospholipids as described by Saito and Hakomori (6). with 0.1 N :JaOH at 37'C for lh, The acidic lipid fraction was treated neutralized with HCl and then dialyzed. Components of acidic and neutral glycolipids were identified with HPTLC(Merck). The loaded plate was developed with chloroform/methanol /water containing 0.5% CaC12 (60/40/9,v/v/v) for gangliosides or chloroform/methanol/water (60/40/5,v/v/v) for neutral glycolipids. The amounts of the applied samples were such that they represented the aliquots of the same amount of the wet tissue. Spots were detected as described previously (5). The standard glycolipids were GM3, GM1 and GDla extracted from l-2 week old mouse small intestine (5), and monohexosyl and dihexosyl ceramides, asialo GM1 from mouse small intestine (5). The amounts of sialic acid and hexose in each spot were determined with TLC scanner CS 920 (Shimadzu co., Ltd., Kyoto) at an analytical wavelength of 530 nm and 520 nm, respectively. RESULTS Major are

When the

that

ceramide

scarecely

(5).

the

asialo

GM1

hormonal

adult

amount

of

both

(lactosyl

(Fig.

1).

(Figs.

GM1 was

present

relative

in

2A).

both

amount

asialo

GM1

(Figs.

faster

than

at

was

14 d of

age of

2A).

Relative by

GMl,were in days

hormone-injected

the

constantly

augmented

28

found

detected

intestines

age)

thyroxine

irrespective

was

asialo

weaning, and

at

slightly

mouse

After

it

1A and

also

d of or

section,

CMH

asialo

(14

cortisone

in

was

control

mice

with

contrast,

intestine However,

increased

intestines

bands

(7,8).

method

In

small

suckling

treated

hormone-treated

1A and

of

acquired

2A).

The

CM1

the

ceramide)

and

asialo

mouse

the

detected similar

of mice

age, in

asialo similar

amounts. The

relative

in

the

to

developmental

amount

in

suckling

control

amounts

described

animals

adult

organ

were

(Fig. and

same

animals

weaned

CDH

treatment

the

and

precociously

treatment

both

for

as

was

The

1A).

in

suckling work

of

(CMH)

found

present

(Fig.

in

glycosphingolipids

monoglycosyl

G141 is

in

neutral

method

of

cortisone-treated

asialo

amounts section

and

age

(Table

GM1

was

animals,

of

the

the

above

the

results

I). 30

and

lipids

The 40%

for

respectively,

were are

summerized

results

show

the

thyroxine-treated

whereas

1169

estimated

that

as with

that

the

described

relation relative and

for

the

control

the

Vol. 119, No. 3, 1984

BIOCHEMICAL

Neutral

A

AND BIOPHYSICAL RESEARCH COMMUNICATIONS

Neutral

Acidic

B

Acidic

A

CMH

0

CMH -em----

CDH

CDH

GM3

‘T

GM3

asialoGM1

asialoGM1

GM1

GM1 -” Nor. 14

.-

._

Thyro. 14

Car. 14

Nor. 14

Thyro. 14

COr. 14

0

NM.

wro.

28

20

COr.

Nor.

28

28

Thyro.

0

1

2

Fig.

1

Fig.

was

less

showed

than 40

to

ment(Table

Table

2

qlycollpids and qangliosides at 14 d of age. The solvent were as described in the method Nor, control mote; Thyro, thyroxine-injected car , cortisone-incected mice

of

TLC of neutral qlycollpids and qangliosides small intestine at 28 d of age. The solvent and reagents were as described in the method Nor, control mice; Thyro, thyroxine-injected COI, cortisone-injected mice

of

4% at 50%

I).

TLC of neutral small intestine and reagents

for The

I.

d of

14

the

age

same

relative

(Table lipid

amount

Neutral CMH

The

adult

irrespective of

Percentage distribution and qangliosides-bound

;iqe and conditions

I).

of sialic

the

highest

hexose acid

of

asialo

mouse systems section. mice;

intestines

hormonal

treat-

the

control

glycoliplds small intestine Gangliosides (%)

glycosphingolipids (%) CDH

mice:

for

neutral mouse

in

section.

small

of

CMH was

mouse systems

GM1

GM3

GM1

GDla 16.2

14 nor.

69.1

4.1

23.0

51.0

32.8

14

50.6

6.9

28.5

14.0

38.0

55.8

6.2

34.1

a.3

38.0

18.5

35.8

57.3

6.9

Thyro.

14 car.

-3.8

Others

28 Nor.

43.0

0.5

48.9

7.6

38.9

61.1

-

28

40.2

0.4

45.7

13.7

59.6

40.4

-

43.7

0.4

39.2

16.7

57.6

42.4

-

Thyro.

28 car.

* FOT abbrevlatlon

see

the

legend

to

Figs.

1 and

2

28

‘hr.

2a

Vol. 119, No. 3, 1984

group

(70%)

the

at

BIOCHEMICAL

14

d of

cortisone-treated

ranged

around

a minor

40%

14

d of

in

contrast

4%)

but

of

to

the

asialo to

of

throxine

decreasing

at

14 d of

and

in

ganglioside

in

present by

most

prominent

the

developmental

mimicked of

the

the

GM3

intestine The

hormonal

was

the

of

GDla

(Fig.

change

in

1B and the

the

than order

order

(Table

was

that

was

whereby was

also

amounts

of

GM3

and

GM1

The

relative

in

control

in the

decrement

It

GM1

the the

the in

I).

of

in

of

decreased

the

I).

found

amount

precociouly

(Table

reversed

change

demonstrated

change

was

administration

the

as

and

control

developmental

lB),

relative

mouse

GM1

the

lipids,

Table

(about

Gb13, in

yanyliosides (Fig.

treatment

suckling

If

treatment,

the

in

I). normal

intestine

I).

(5).

neutral

treatment

greater

reversed

intestine

of

control

gangliosides Table

CDH was

component

amount

the

hormone-accelerated

whereas

same

by All

a minor

and

animals

hormonal

(Table

major

case

the

the

weaning

1B and

in

decrease

for

by

amount

as

by

considerable

the

weaned The

in

7-8% also

after

the

treatment.

treatment

accelerated

experiment.

amount

in

(Fig.

cortisone

should

was

(53%)

for

amount to

the

order, age

that

hormonal

which

by

thyroxine-treated

relative

diminish

contents

gangliosides the

GM1

they

in

were,

the

increased

present

but

the

whereas

4% in

30-409,

are

by

of

about slightly

increased

intestines

groups

and

intestine

GDla

(35%)

age

Gangliosides small

followed

regardless

component

at

age

AND BIOPHYSICAL RESEARCH COMMUNICATIONS

was

noted

that

was

suckling

control

adult

hormone-treated

intestine.

suckling

I).

DISCUSSION Previous

studies

administration

mental

weeks contents.

the

However,

(Y-11).

work

on

have

it

of

became

evident

M&jor

the

glycosphingolipid

little

alteration

accelerated

demonstrated

is

intestinal that

normal gangliosides

effects

of

compositions

known

about

a hormonal developmental markedly

of

hormonal

glycolipid

treatment

decreased

several

effects

in in

early

intestinal in

tissues

on In

contents.

change

hormonal

amount

the

the

develop-

present

postnatal ylycolipid and

gave

way

Vol. 119, No. 3, 1984 to

the

and

increasing

appearance

of

newly

synthesized

asialo

GM1

(Figs.

1

2). The

GDla

developmental

which

the

normal

that

the

weaning

accelerated previous

one

genetic

off

that

the

fetal-suckling

thyroxine

for

developmental integral

components

may

reflect

the

as

a whole.

behind

The

those

of

identified

previously

be

from

derived

accord

with

in

epithelium It

(12). the

The alone

in

and

thyroxine

based

is

by

on

the

the

hormonal

intestine in

tissue

early fractionation

light

qlycolipid

administration

was

postnatal

affected

postnatal

weeks and

the must

asialo

morphological

1172

similar and

glycolipids changes

the

membrane

asialo the

GM1

but

Forssman

postulated

to

postulation

is

GM1

present of

in

answered

tissues

observation.

the

TLC

(Fiq.

the

epithelium of

amount

was

work

by

in

localized

that

administration be

from and

mesecymal

development by

the

finding

the

that

specifically

the

latter

in

(4)

ceramide)

was

in

before

of

This GM1

the

cortisone

was

(8).

glycolipid

is

regulated

finding

of

of

Forssman

the

off

the

with

of

globotetraosyl

of

and

at

developmental

(or

the

phosphatase

those

asialo

on

phophatase

those

by

is

precisely

by

their

to

migrating

the

is

operation

before

the

or

suggests

compatible

differentiation

Forssman

whether

on

alkaline

correspond

that

glycolipids

switching

alkaline

tissue

in

switched

accelerated

both

non-epithelial

interstinq

GM1

The

membrane,

The

tha

phenomenon

is

the

ceramide

observation

question the

IA)

treatment

qlycolipids

is

migrating

(8).

and

neutral

affected

bands (Fig.

asialo

suggests

plasma

triqlycosyl

the

type

for

glycosphinqolipids

intestinal

developmental

CDH

This

of

remarkable

hormonal

assumption

rat

adult

of

TLC

and

of

Because

general

glycolipid

of

of

mechanism.

the

differentiation.

also

are

the

organ

administration

by

especially

acidic

expression

the

be

developmental

transition to

some

This

the

programme

developmental

this

to

2B).

that

hormones.

(12)

the

of

while

that

the

1B and

synthesis

and

by

disappears

(Figs.

switched

of

seems

completely

development

precociously time

transition

almost

possibility

by

BIOCHEMICAL AND BlOPHYSlCAL RESEARCH COMMUNICATIONS

not 1A).

cortisone further

work

Vol. 119, No. 3, 1984

BIOCHEMICAL

AND BIOPHYSICAL RESEARCH COMMUNICATIONS

REFERENCES 1. 2. 3. 4. 5. 6. 7. 8. 9 10. 11. 12.

Moog, F. (1981) Textbook of gastroenterology and nutrition in infancy (Lebenthal, E. ed. Raven Press, ;?ew York) 139-147 Moog, F. (1953) J. Exp. Zool. 124, 329-346 and Flooy, F. (1975) t&vBiol. 47, 173-184 Y&l, K.-y. T. and Fujita, M. (1983) J. Biochem. 94; 1483-1483 Uezato, Sato, C., Uezato, T., Fujita, PI. and ;?ishimura, K. (1982) J. Biochem. 91, 2013-2019 Saito, T and Hakomori, S. (1971) J. Lipid Res. 12, 257-259 Umesaki, Y., Suzuki, A., Kasama, T., Tohyama, K., Mutai, M. and Yamakawa, T. (1981) J Biochem. 90, 1731-1738 Hansson, G. C., Karlsson, K.-A., Lefflcr, H. and Str&berg, N. (1982) FEBS Lett. 139, 291-294 Hay, J.B. and Gray, G.M. (1970) Biochim Biophys. Acta 202, 566-568 Nishinura, K. and Shimoda, R. (1975) Jpn. J. Exp. Med. 45, 241-244 Horowitz, A.J. and Schaberg, S.M. (1979) Biochem. Pharmacol. 28, 881-895 Suzuki, A. and Yamakawa, T. (1981) J. Biochem. 90, 1541-1544

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