Pentagastrin protects against stress ulceration in rats

Pentagastrin protects against stress ulceration in rats

Pentagastrin Protects Against Stress Ulceration in Rats K. TAKEUCHI, Ph.D., and LEONARD R. JOHNSON, Ph.D. Department of Physiology, University of T...

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Pentagastrin Protects Against Stress Ulceration in Rats K. TAKEUCHI,

Ph.D., and LEONARD R. JOHNSON, Ph.D.

Department of Physiology,

University of Texas Medical School, Houston, Texas

Stress ulcer formation was examined in the following four groups of 18 rats each: (a) chow fed, (b) fed an isocaloric amount of a liquid diet, (c) fed a liquid diet plus pentagastrin injections (250 pg/kg, 3 times/ day), and (d) fed a liquid diet plus histamine (20 mg/ kg, 3 times/day). The diet regimen lasted 10 days, and injections of saline, pentagastrin, or histamine were given over the last 7 days before water immersion induced stress. Six rats from each group were killed before stress (zero time) and after 4 and 20 hr of stress. Serum gastrin and oxyntic gland mucosal DNA synthesis were significantly lower before stress in ail liquid-fed rats compared with chow-fed controls. At this time, the ulcer index was zero in all groups. DNA synthesis decreased, and the ulcer index increased progressively in all groups with exposure to stress. After 4 hr of stress, the ulcer index was 3.8 in the group receiving pentagastrin, compared with 14.1 and 10.7 in the animals receiving liquid diet alone or liquid diet with histamine injections, respectively. During stress, DNA synthesis in the group receiving pentagastrin decreased significantly less than in all other groups. There was significant correlation (r = 0.782; P < 0.01) between uJcer index and the decrease in DNA synthesis. The authors conclude that rats fed liquid diets are more susceptible to restraint-induced stress ulcers because of their lower endogenous gastrin levels and that exogenous pentagastrin increases the resistance of these animals to ulceration. The susceptibility of all groups of animals to stress ulceration was directly correlated with decreases in DNA synthesis. Received July 21, 1978. Accepted September 28, 1978. Address requests for reprints to: Dr. L. R. Johnson, Department of Physiology, University of Texas Medical School at Houston, P.O. Box 20708, Houston, Texas 77030. This study was supported by NIH grants AM-18184 and AM16505. The authors are grateful to Mr. Paul D. Guthrie and Ms. Robin Bailey for technical assistance. 0 1979 by the American Gastroenterological Association 0018-5085/79/020327-08$02.00

A normal gastric mucosa is the manifestation of balance between the forces acting to destroy the lining of the stomach and those acting to maintain or rebuild it. Theoretically, an ulcer can develop if either regeneration decreases or destruction increases sufficiently to result in an imbalance. The destructive factors have been thoroughly studied: acid, pepsin, aspirin, ethanol, bile, and a host of events labeled as “stress”; but little is known about alterations in regenerative forces that could lead to ulcer formation. Previous studies from other laboratorie? have demonstrated that during stress, the gastric mucosa has a decreased rate of cellular proliferation, decreased DNA synthesis,‘-” and an overall loss of RNA.“,’ These investigators have suggested that the inhibition of DNA and RNA synthesis and mitosis, with resulting failure to replace extruded epithelial cells, contributes to the development of gastric erosions. Hypophysectomy causes gastric mucosal atrophy, decreased RNA content, and loss of parietal and chief cells.5.6 The effects of hypophysectomy on the gastric mucosa are largely prevented by growth hormone but not by other pituitary hormones.5 Prompted by these observations, Vanamee et al.’ hypothesized that growth hormone treatment might protect animals from stress ulceration. Rats were injected with varying doses of growth hormone, 1 and 10 hr into a 24-hr period of restraint stress. Growth hormone provided partial but significant protection against ulcer formation that was directly related to the dose of hormone administered.’ During the past several years, it has become evident that gastrin is an important trophic hormone in the mucosa of the oxyntic gland area.’ The administration of exogenous gastrin restores normal structure to oxyntic mucosa from rats made hypogastrinemic by antrectomy’ or total parenteral feeding.‘” Growth hormone appears to be necessary to maintain both antral and serum gastrin levels, because

328

TAKEUCHI AND JOHNSON

hypophysectomy reduces gastrin levels to a small fraction of normal.” Treatment of hypophysectomized rats with growth hormone restores serum gastrin levels to normal and significantly increases antral gastrin content.” Pentagastrin is able to restore normal growth of the duodenum and pancreas in hypophysectomized rats.” The current study tested the hypothesis that the trophic action of gastrin is important in maintaining the resistance of the gastric mucosa to stress ulcers.

Methods Animals Male Sprague Dawley rats with a mean body weight of 180 g were divided into four groups and treated as follows: Group A consumed equal measured amounts of normal Purina rat chow, and the animals were injected with saline 3 times/day; group B drank a liquid diet isocaloric to the chow consumed by group A, and the rats were injected with saline; group C received the liquid diet plus 250 pg/kg diet 3 times/day; group D received liquid diet plus 20 mg histamine/kg diet pentagastrin 2 HCl three times/day. All diets were maintained for 10 days before an experiment, and injections of NaCl, pentagastrin, and histamine were given during the last 7 of those 10 days. The liquid diet consisted of 50 ml/day of a solution containing 25% dextrose and 4.25% amino acids (FreAmine II, McGaw Laboratories, Glendale, Calif.) with the following additives per liter: 10 meq NcCl, 15 meq Na acetate, 25 meq KCl, 35 meq K acetate, 10 mEq NaHPO,, 3.5 meq Mg SO.,, 2 g calcium gluconate, and 0.5 cc of a parenteral multivitamin concentrate (M.V.I., U.S. Vitamins Corp., Tuckahoe, N.Y.). This diet contained 1.2 kcal/cc. Daily examination of the urine showed that it was free of glucose and did not contain abnormal amounts of acetone.

Gastric Secretion Basal gastric secretion was measured in 24 rats, six from each of the four groups, to determine the effect of the IO-day dietary and injection regimen on acid and pepsin secretion. After an 18-hr fast, the pylorus was ligated under ether anesthesia and the abdominal wound closed by suturing. Ligation occurred 8 hr after the last injection of saline, histamine, or pantagastrin. At this time, the secretory response was over for several hours. The animals were kept for 5 hr in separate cages with wide mesh bottoms and then killed with ether. The stomachs were removed, and their contents were collected and centrifuged. The volume of each sample was determined to the nearest 0.1 ml and the acid concentration measured by automatic titration to pH 7.0 using 0.2 N NaOH. The pepsin concentration was determined with a modification’3 of the Anson14 hemoglobin method and expressed as milligrams of pepsin per milliliter by reading the trichloroacetic acid supernate at 280 pm and comparing it with different pepsin standards (hog pepsin, three times crystallized, Pentex Biochemicals, Kankakee, Ill.).

GASTROENTEROLOGY,

Vol. 76, No. 2

Stress Rats were stressed by placing them in a stress cage and immersing it in 23°C water to the rat’s xyphoid process. Eighteen animals from each group were used in one experiment. Six of these were sacrificed without stress (0 hr stress), and six were killed after 4 hr and six after 20 hr of stress. In another experiment, from an additional 12 chow-fed rats (group A type), six animals were stressed for 7 hr and 6 for 14 hr. In a third study, after an overnight fast, 6 group A and 6 group B rats were pylorus ligated and stressed for 20 hr to measure gastric secretion. All rats were killed (0 hr stress) or subjected to stress 8 hr after the last injection of saline, pentagastrin, or histamine. Stress began at 8 AM in the 4- and 7-hr experiments and at 4 PM in the 14. and 20-hr studies.

Biochemical

Measurements

and Ulcer Jndex

After the stress period, rats were anesthetized with ether. A midline incision was made, and blood was withdrawn directly from the heart for radioimmunoassay of gastrin. The stomach was removed, opened along the greater curvature, and rinsed with cold saline. The antrum was carefully dissected from the oxyntic gland area and weighed. Lesions occurred only in the oxyntic gland area. The lengths of these were measured, and the total length of the lesions was recorded as the ulcer index for each rat. The ulcer indices from each member of the four groups were summed and divided by 6 to give the group index. The rate of DNA synthesis in mucosa scraped from the oxyntic gland area was measured by incubating the tissue at 37°C for 30 min in Eagle’s minimal essential culture medium containing 2 &i/ml of [“Hlthymidine (5 Ci/mmol, Amersham, England). Flasks were gassed continuously with 95% oxygen and 5% carbon dioxide during the incubation. The reaction was stopped with 0.4 N perchloric acid containing carrier thymidine at 5 mM concentration. Samples were washed in 0.2 N perchloric acid, and RNA was removed by hydrolyzing in 0.3 N KOH for 90 min at 37’C. DNA and protein were reprecipitated with 10% perchloric acid. The RNA content of the supernatant was determined by using the orcinol reaction.15 After standing on ice for 10 min, the DNA-containing tubes were centrifuged and the supernatant discarded. DNA in the residual pellet was solubilized in 10% perchloric acid heated to 70°C for 20 min. Using calf thymus DNA as a standard, the DNA content of the samples was determined by the Burton” procedure as modified by Giles and Myers.” The incorporation of [3H]thymidine into DNA was determined by counting 0.5 ml of the DNA-containing filtrate in a Beckman liquid scintillation system. (Beckman Instruments, Inc., Spinco Div., Palo Alto, Calif.) DNA and RNA content were expressed as micrograms per 100 milligrams of 100 mg mucosa and DNA synthesis as disintegration per minute per milligram tissue.

Gastrin nized

Determination

After weighing, the antral samples were homogeand extracted in boiling water according to the

February

Table

PENTAGASTRIN

1979

1.

Basal

Gastric

Secretion

During

Group B, liquid

chow Volume (ml/Zft hr) Acid output (peq/hr) Pepsin output (mg/hr) ” Mean + ” P < 0.05 ’ P < 0.05 I’P i 0.01 ” P < 0.05

9.9 * 0.5 149.5 +- 13.6 2.11 + 0.18

Observations

0.5 8.6’ 0.13”

Group D, + histamino

7.8 f 0.7” 113.6 f 15.4” 1.62 + 0.07”

Results Gastric

Secretion

As shown in Table 1, both volume and acid output were significantly decreased in groups B and D compared with chow-fed controls (group A). Injection with pentagastrin (group C) maintained basal secretion at control levels. The same essential findings applied to pepsin secretion. Output of pepsin from rats in group C was, however, higher than that in either group B or group D.

E#ect

of IO-Day Treatments

on Four Groups

Effects of Fasting Because stressed animals would necessarily be fasted, the effects of 20-hr fasting were compared with various periods of stress (Figures 1 and 2). All

of Rats”

Group

A,

chow Body weight (g) Oxyntic gland weight (mg/lOOg body wt) Antral weight (mg/lOO g body wt) Serum gastrin (fmol/ml) Antral gastrin (pmol/antrum) Ulcer index (mm) I’Animals arc identical to I’P < 0.05 compared with ’P < 0.01 compared with I’P < 0.001 compared with ” P -C 0.01 compared with

those killed A. A and C. A. A.

at 0 hr of stress:

213.0 312.0 58.0 156.9 857.2 0 mean

at Zero Time

Animals receiving liquid diets gained significantly less weight than those in the chow-fed group (Table 2). There were no differences in the weights of any of the liquid-fed animals. The weight of the oxyntic gland portion of the stomach in animals from groups B and D was significantly lower than the weight of the same tissue from chow-fed and pentagastrin-injected rats. Pentagastrin completely prevented the loss in oxyntic gland weight observed in the animals in the other two groups fed liquid diets (Table 2). There were no significant differences in the weights of the gastric antrums from the 4 groups of rats. Both serum and antral gastrin levels were significantly lower in all groups fed liquid diets compared with group A. Serum gastrin was reduced to only 20% of normal, and there were no significant differences between gastrin levels in groups B, C, and D. Except in one animal in group D, there were no lesions present in the stomach at zero time.

Means were compared by the Student’s t-test for values and were considered to be significantly if F’< 0.05.

2.

9.2 f 138.9 * 1.85 f

liquid

SEM: n = 6. comparc!d with A. compared with B. compared with A. compared with B and D.

Statistics

Table

Group C, + pentagastrin

liquid

7.3 f 0.9’1 103.1 + 12.1” 1.54 f 0.09~’

method described by Lichtenberger et al.“’ Serum samples and antral extracts were frozen for subsequent radioimmunoassay of gastrin. The gastrin assay was performed according to the method outlined by Yalow and Berson,“’ with the use of an antibody (1296) kindly donated by Dr. John H. Walsh and characterized by Dockray and Walsh.‘” Serum gastrin copcentration was expressed as femtomoles per milliliter and antral gastrin as picomoles per antrum.

unpaired different

Groups of Rats”

5 hr in Each of Four

Group A,

32H

PROTECTS AGAINST STRESS ULCERS

f * f f +

5.2 7.0 4.0 16.5 53.2

+ SEM: n = 6.

Group B, liquid 189.4 290.5 64.0 25.8 336.8 0

f 2.3” f 4.5’ f 5.0 + 4.5” -+.4X.4”

Group C. liquid + pentagastrin 195.6 324.2 56.4 39.3 473.6 0

f f * f +

2.6” 14.5 4.0 14.6” 66.0“

Group D, liquid + histamine 190.5 283.2 59.1 34.5 344.2 0.6

I & f -c f f

3.7” 10.3’ 2.3 16.0” 70.3” 0.4

330

TAKEUCHI AND JOHNSON

GASTROENTEROLOGY,

Effects of Pentagastrin

r30

LO

Hours of Stress

Figure

Z.Oxyntic gland mucosal DNA synthesis in DPM [3H]thymidine incorporated into DNA per milligram tissue in animals from group A (chow fed). Bars compare nonfasted control animals with ZO-hr fasting by itself and with various periods of stress, which include fasting. The dark line shows the ulcer indices in the groups of animals. Points and bars are the means of six observations. Vertical lines are standard errors of the P < 0.05 when compared with control. At 14 means. ?? and 20 hr, *P < 0.001 compared with control or 20 hr fasting.

these rats were from the chow-fed group A. Fasting caused a significant 25% decrease in DNA synthesis but no ulcer formation. DNA synthesis decreased dramatically during stress, so that after 14 or 20 hr of stress, the rate of thymidine incorporation was approximately one-third of normal and one-half of the level found in 20-hr fasted rats (P < 0.001). The ulcer index increased with the duration of stress in a pattern similar to the decrease in DNA synthesis (Figure 1). Neither the RNA nor the DNA content of the oxyntic gland mucosa of chow-fed rats decreased significantly during the 20-hr fasting period. After 14 and 20 hr of stress, however, there were significant (P < 0.01) decreases in total mucosal RNA (Figure 2), whereas the DNA content did not change. 1200 2 $000

DNA

0

RNA

t

E” 8 800 y \

ng

L

20

I

Hours

Acid Secretion

ml 14

T

Mucosal DNA synthesis of rats fed liquid diets was significantly lower than it was in those fed chow (Table 3, 0 hr). The level of DNA synthesis in pentagastrin-treated rats (group C) was, however, significantly greater than the corresponding measurement for groups B and D. Liquid-fed animals not receiving pentagastrin (groups B and D) had significantly less mucosal DNA and RNA than group A. Pentagastrin injection maintained RNA and DNA content at control levels. After 4 hr of stress, the ulcer index in animals fed the liquid diet alone (group B) increased dramatically (Figure 3). Pentagastrin treatment prevented this increase in ulcer formation, and animals in group C did not have significantly more ulceration than those in group A. At all times, ulcer formation in the histamine-injected rats (group D) was. almost identical to that in group B. (The data from group D is not depicted on Figure 3 in order to make the figure more readable). After 20 hr of stress, the ulcer index for group A increased dramatically to 23.5 f 1.7 to approximately the amount of ulceration found after 20 hr of stress in groups B and D. The ulcer indices for groups B and D after 20 hr of stress were 22.5 f 3.2 and 23.9 f 0.7, respectively. Ulcer formation was also increased significantly at 20 hr in group C. The ulcer index of 14.0 f 2.5 in the gastrintreated group, however, was significantly lower than those for all other groups. Changes in DNA synthesis during stress are shown in Figure 4 and Table 3. After 4 hr, DNA synthesis was significantly reduced from O-hr levels in groups B and D. After 20 hr of stress, DNA synthesis in group A rats had dropped to one-third of the O-hr level (P < 0.001). The other groups also had further decreases after 20 hr of stress, but these were small in comparison with those of the chow-fed rats. Oxyntic gland mucosal DNA content did not significantly decrease with stress in any of the four groups. RNA content, on the other hand, decreased significantly after 20 hr of stress in all groups except the one injected with pentagastrin (Table 3).

t 31’1

m

Vol. 76, No. 2

of Stress

Figure 2. DNA and RNA content of oxyntic gland mucosa from rats in Figure 1. +P < 0.01 compared with control.

During Stress

Figure 5 depicts the ulcer indices and acid secretion in pylorus-ligated rats from groups A and B during 20 hr of stress. Pylorus ligation greatly increased ulceration in the liquid-fed animals to approximately twice that in animals fed normal solid chow. This greater incidence, however, could not be attributed to changes in acid secretion, because volume and concentration of acid secreted during the 20 hr were essentially equal in the two groups.

February

1979

Table

3.

PENTAGASTRIN

Clxyntic

Gland

Mucosal

DNA

Synthesis

and

DNA

and

PROTECTS

4

A, chow

342.5 k 33.0

B, liquid C, liquid

+ pentagastrin

D, liquid

+ histamine

133.7 zk 17.4c 187.7 + 14.8=f 144.1 f 9.6”

1,199.3

f

B, liquid C, liquid D, liquid

891.6 1,025.6 946.1

rt 15.6” + 20.6 + 28.1”

+ pentagastrin + histamine

” Mean +SE:M; n = bP < 0.001 compared ’ P < 0.01 compared d P < 0.05 compared ’ P < 0.01 compared f P < 0.05 compared

6. with with with with with

20 _

[3H]Thymidine incorporation (dpm/mg tissue) 296.7 +- 31.4

118.5 f

84.2 f 12.7d 148.4 f 14.3 98.7 + 12.8”

387.9 & 10.4 285.8 +- 5.7” 366.6 + 7.0 310.3 * 14.5c

A, chow

331

ULCERS

of stress

0

A, Chow B, liquid C, liquid + pentagastrin D, liquid + histamine

STRESS

RNA Content During Stress” Hours

Group _____

AGAINST

54.2

5.4b

76.2 + 9.8e 123.9 + 7.gd 84.0 k 4.4b

DNA content wg/lOO mg tissue 381.7 -t 7.3 280.7 + 6.1 329.1 + 25.3 299.1 + 6.7

34.16 + 294.1 + 342.5 + 290.2 +

18.1 7.8 13.8 9.1

RNA content pg/lOO mg tissue 1,134.4 + 62.2

982.5

36.4”

800.6 986.5 933.4

f f +

66.0 35.7 63.3

f

757.1 rt 13.gb 967.1 +- 29.8 826.5 -t 16.7”

0 hr stress. group A. 0 hr stress. 0 hr stress. groups B and D.

Discussion Many different laboratories have shown that gastrin is an important trophic hormone specific to the pancreas and mucosa of the gastrointestinal tract.z’-2” The purpose of this study was to test the

hypothesis that the trophic action of gastrin might be an important factor in the resistance to gastric stress ulceration. The use of restraint to produce gastric lesions in small animals is a useful and reproducible experimental model to study the pathogenesis and treatment of stress ulcer in humans.24.25 The method of restraint used in these studies has been

0 Chow

A Liquid ??Liquid

400

I-

O Chow

+ PG

A Liquid

% .p 300

??Liquid + PG

E” & p 200 : s E f

I---

0 Figure

3. Ulcer index rats. PG = for 7 days chow and *hP < 0.01 with chow Points are observations.

4 Hours of Stress

20

at 0,4, and 20 hr of stress in three groups of pentagastrin injected 3 times/day 250 pg/kg preceding stress. *P < 0.05 compared with liquid + PG, P < 0.01 compared with 0 hr; compared with 4 hr, and P < 0.05 compared and liquid: +P < 0.001 compared with 4 hr. means and standard errors of means of six

100

%

0 I

0 Figure

u

I

4 Hours of Stress

,

20

4. DNA synthesis in oxyntic gland mucosa of rats described in Figure 3 *P < 0.05 compared with 0 hr; *+P < 0.001 compared with 0 hr. PG = pentagastrin.

332

TAKEUCHI

GASTROENTEROLOGY,

AND JOHNSON

ml 10

50

a

20-

3

loO-

Figure

T

N.S. T

60

7

P 40!. x 308 f

mEq/L

Chow

Liquid

6

- 60

4

- 40

2

- 20

0 Chow

Liquid

_ n ”

5. Ulcer

indices and acid secretion in pylorus-ligated rats from groups A (chow) and B (liquid) after 20 hr of stress. Means and standard errors of means of six rats in each group.

well characterized and is a reliable way to produce oxyntic gland stress u1cers.28.*7Before the animals were subjected to stress, three of the four groups of rats were placed on liquid diets to lower endogenous gastrin levels. This served two purposes: first, it allowed the authors to examine whether rats with low endogenous levels of gastrin were more susceptible to stress ulceration, and secondly, it produced a gastrin-depleted animal, which would be more sensitive to the effects of the exogenous hormone. The liquiddiet regimen resulted in hypoplasia of the gastric mucosa, as indicated by decreased weight and DNA and RNA content similar to that produced by other models causing gastrin depletion.“~‘“~‘”As was also the case in antrectomized’ and fasted rats” and rats maintained by total parenteral nutrition,‘0 exogenous gastrin largely prevented the mucosal hypoplasia in the liquid-fed animals. The most dramatic finding in the current study was the ability of pentagastrin to protect gastrin-depleted rats from stress ulcer. After 4 hr of stress, liquid-fed rats treated with pentagastrin had an ulcer index of 3.8 f 1.3 compared with 14.1 f 2.9 for liquid-fed rats injected with saline (group B) and 10.7 f 2.5 for liquid-fed rats treated with histamine. After 20 hr of stress, the gastrin-injected rats had significantly fewer and less severe ulcers than even the chow-fed animals. This was a surprising finding, because serum gastrin levels were normal in this group. The large increase in the ulcer index between 4 and 20 hr of stress in group A was correlated with a 60% decrease in DNA synthesis from 296.7 f 31.4 at 4 hr to 118.5 f 5.4 at 20 hr. It was equally interesting that DNA synthesis decreased least in the pentagastrin-treated animals, even though it was proceeding at a higher rate at zero time than it was in groups B and D. Figure 6 is a plot of the relationship between ulcer index and the corresponding decrease in DNA synthesis from one time point to the

Vol. 76, No. 2

next (O-4 hr and 4-20 hr) in all four groups of rats. The correlation coefficient was 0.782, which was significant at P < 0.01. Although ulcer formation was highly correlated with decreases in DNA synthesis, the interpretation of this relationship must be a cautious one. First, it is obvious that decreases in DNA synthesis alone do not produce ulcers. None of the animals had ulcers at zero time (before stress), despite more than 50% decreases in DNA synthesis in the liquid-fed animals. Secondly, it is important to emphasize that ulcer formation was not correlated with the absolute rate of DNA synthesis but with the percent change from one time period to the next. Thirdly, a correlation does not prove causality. However, of the parameters that were measured, namely, acid secretion, DNA content, RNA content, and DNA synthesis, a decrease in DNA synthesis was the best indication of the events occurring in the mucosa that ultimately led to ulceration. Numerous theories have been proffered to explain the mechanism of stress ulceration. Menguy et al.” subjected rats to hypovolemic shock and found significant decreases in gastric mucosal ATP and glycogen levels compared with levels in liver and muscle. They proposed that the severely compromised energy metabolism in the gastric mucosa wasthe cause of this tissue’s susceptibility to hypovolemic shock. This theory was supported to some extent by the finding that starved rats were more susceptible to stress ulcer than those maintained on i.v. nutrition.‘” However, it would appear that nutrition played no role in the current study. All rats received equal amounts of calories, and the three groups of liquidfed animals were maintained identically. Imondi et

30

1

0 0 0

/

/

/ 0

+

/

Chow

A Liquid k u 3

lo/

/

m Liquid + PG *Liquid

10

20

30

40

50

Decrease in DNA Synthesis Figure

60

+ H

70

(O/O)

6. Ulcer index plotted against the change in DNA synthesis for 48 rats. Each point shows mean and standard errors of mean for 6 observations. Two time periods are shown for each of the 4 groups. r = CWX?..which is significant at P < 0.01. PG = pentagastrin, H = histamine.

February1979

al.” have also studied nucleic acid metabolism during stress ulcer formation. They found decreases in DNA synthesis in gastric, duodenal, jejunal, ileal, and colonic: mucosa during stress, but RNA content and synthesis were significantly decreased only in the gastric mucosa. They hypothesized that because ulcers did not occur in any tissue but the stomach, a decrease in RNA synthesis was an important etiologic factor. RNA synthesis and protein synthesis are prerequisites for DNA synthesis,“” so that a decrease in RNA eventually would lead to cell necrosis. The authors’ results support these findings in general, except that they observed significant ulceration at 4 hr in groups B and D without decreases in RNA content (Table 3). In their hypotheses about the etiology of gastric stress ulcers based on differences when compared with other tissues, it is interesting that the authors of the above-mentioned studies have neglected the facts that neither liver and skeletal muscle” nor the rest of the gastrointestinal tract3 secrete acid. Therefore, the failure of the coionic mucosa, for example, to develop ulcerations in the study by Imondi et al.” can be explained by its lack of contact with acid as easily as its ability to maintain RNA levels. The current data certainly do not elucidate the mechanism of stress ulcer formation, but they do prove that the proliferative or regenerative capacity of the mucosa is of paramount importance, in resisting stress. This idea is not a new one, but, to the authors’ knowledge, the only other attempt to test it involved the use of growth hormone. In that study, Vanamee et al.’ administered growth hormone to rats at the beginning of and during a 24hr period of stress. Growth hormone provided significant protection against ulceration, but no other measurements were made to elucidate the mechanisms of protection. In any case, it is doubtful whether the trophic effects of growth hormone could play a major role in such a short time period. The intervals between injection of a trophic substance and the stimulation of DNA synthesi9’ and cell division’” are of the order of 16 and 20 hr, respectively. Its beneficial effects could just as easily have been caused by early alterations of metabolism. The importance of the proliferative capacity of the mucosa as a factor in the resistance to stress ulcer is further emphasized by the seemingly paradoxical ability of gastrin, a stimulator of acid secretion, to protect against ulcers. Note that at the start of the stress period, the two groups (A and C) resistant to ulcer formation at 4 hr had almost 50% higher basal rates of acid secretion than the other two groups. The following conclusions can be drawn from the authors’ data. First, rats with low endogenous gastrin levels are more susceptible to stress ulcers than

PENTAGASTRIN

PROTECTS

AGAINST

STRESS

ULCERS

333

animals with normal gastrin levels. Secondly, chronic pretreatment of gastrin-depleted rats with pentagastrin significantly protects against stress ulcer formation. Thirdly, the protective effect of gastrin is due to its trophic action on the gastric mucosa. Furthermore, these findings suggest that the regulation of the regenerative factors influencing the gastric mucosa are at least as important as the regulation of the much-studied destructive factors.

References 1. Kim Y, Kerr RJ, Lipkin M: Cell proliferation during the development of stress erosions in the mouse stomach. Nature (Lond) 215:1180-1181, 1967 2. Lahtiharju A, Rytomaa T: DNA synthesis in fore and grandular stomach and in skin after nonspecific stress in mice. Exp Cell Res 46:593-596, 1967

3. Imondi

4.

5. 6.

7.

8. 9.

10.

11.

12.

13.

14. 15. 16.

AR, Balis ME, Lipkin M: Nucleic acid metabolism in the gastrointestinal tract of the mouse during fasting and restraint-stress. Exp Mol Path01 9:339-348, 1968 Ludwig WM, Lipkin M: Biochemical and cytological alterations in gastric mucosa of guinea pigs under restraint stress. Gastroenterology 56:895-902, 1969 Crean GP: The endocrine system and the stomach. Vitam Horm 21:215-280, 1963 Jacobson ED, Magnani TJ: Some effects of hypophysectomy on gastrointestinal function and structure. Gut 5:473-479, 1964 Vanamee P, Winawer SJ, Sherlock P, et al: Decreased incidence of restraint-stress induced gastric erosions in rats treated with bovine growth hormone. Proc Sot Exp Biol Med 135:259-262, 1970 Johnson, LR: The trophic action of gastrointestinal hormones. Gastroenterology 70:278-288, 1976 Johnson, LR, Chandler AM: RNA and DNA of gastric and duodenal mucosa in antrectomized and gastrin treated rats. Am J Physiol 224:937-940, 1973 Johnson LR, Lichtenberger LM, Copeland, et al: Action of gastrin on gastrointestinal structure and function. Gastroenterology 68:1184-1192, 1975 Enochs MR, Johnson LR: Effect of hypophysectomy and growth hormone on serum and antral gastrin levels in the rat. Gastroenterology 70:727-732, 1978 Mayston PD, Barrowman JA: Influence of chronic administration of pentagastrin on the pancreas in hypophysectomized rats. Gastroenterology 64:391-399, 1973 Northrup JR, Kunitz M, Herriot RM: Crystalline Enzymes. Second edition. New York, Columbia University Press, 1948, p 303-307 Anson ML: The estimation of pepsin, trypsin, papain and cathepepsin with hemoglobin. J Gen Physiol 22:79-89, 1938 Ceriotti G: Determination of nucleic acids in animal tissues. J Biol Chem 214:59-70, 1955 Burton K: A study of the conditions and mechanism of the di-

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