Improvement in the intestinal processes of hydroelectrolytic absorption ...

Surg F. Fuertes TodayGuiró et al.: Octreotide and Experimental Surgical Lesions Jpn J Surg (1999) 29:419– 430

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Improvement in the Intestinal Processes of Hydroelectrolytic Absorption and Secretion in Abdominal Pathologies of Surgical Interest Treated with SMS 201-995: Experimental Protocol Fernando Fuertes Guiró1, Guido Bertolini2, and Joan Viñas Salas3 1

International University of Catalonia, Barcelona, Spain Laboratory of Clinical Epidemiology, Institute “Mario Negri”, Campus of Milan, Milan, Italy 3 Department of Surgery, University of Lleida, Lleida, Spain 2

Abstract: The hypothesis that octreotide can improve the intestinal absorption and secretion processes in a mixed group of intestinal pathologies, and that this effect varies according to the pathology in question, was tested. One hundred and twenty Wistar rats were randomly assigned to six pathology groups consisting of three intestinal occlusions including (1) complete, (2) partial, and (3) complete with strangulation, and three mesenteric vascular occlusions including (4) partial permanent, (5) total permanent, and (6) total temporary. Each group contained ten control and ten treated rats. The treated animals received octreotide (100 µg/kg body weight) while the controls were given the same quantity of saline solution every 8 h. After the observation period, the contents of the small intestine were extracted and its volume measured before and after centrifugation; the concentration and total content of Na, K, Cl, and bicarbonate was then analyzed. Samples of all the intestines at specific distances from the lesion zone were treated and stained, and then evaluated according to a specific score to quantify the lesions. The concentration and contents of electrolytes in the intestine and its volume (before and after centrifugation) were lower in the treated animals, but varied according to the pathology. There was a nonadditive influence between the type of pathology and treatment for the four electrolytes and intestinal volume. The effects of the drug make it directly or indirectly possible to decrease the intestinal lesions to improve the absorption-secretion processes. Octreotide acts on intestinal secretion and absorption in all the pathologies analyzed except for total permanent intestinal ischemia. Its action also varies according to the type of pathology involved. Key Words: somatostatin, octreotide, intestinal obstruction, mesenteric ischemia, electrolyte

Reprint requests to: F. Fuertes Guiró, Plaza de España, 3, E25002 Lleida, Spain (Received for publication on July 22, 1997; accepted on Sept. 11, 1998)

Introduction Intestinal mechanical occlusion and mesenteric ischemia provoke the intestinal dysfunction that involves all intestinal physiologic activity including absorption and secretion, electrical activity and motility, neural and hormonal control mechanisms, immunity, bacterial flora, and responses to pharmacological agents, with severe repercussions in the general organism homeostasis. There are contradictory hypotheses in the literature regarding the pathophysiologic mechanisms that alter these intestinal processes. In intestinal occlusion it has been postulated that an increase in the blood flow occurs in response to both mechanical phenomena (increase of intraluminal pressure up to 30 mmHg, excessive distension of the intestinal wall, altered peristalsis),1,2 and inflammatory phenomena (intraluminal bacteria growth and subsequent liberation of mediator substances, such as prostaglandins and endotoxins which affect the blood flow, secretion, and intestinal motility).3 Contrary to these theories, the literature of the 1960s and 1970s support the hypothesis that in the course of intestinal occlusion there is a decrease in the blood flow to the intestinal wall in response to increased intraluminal pressure.4,5 An increase in the intestinal surface due to wall distension and activation of bioactive peptides, such as vasoactive intestinal peptide (VIP), could also explain these alterations in the evolution of intestinal occlusion.6 Alterations in the processes of absorption and secretion during ischemia are due to lesions of the mucosa in the villi and the liberation of vasoactive substances. It has been postulated that the responsible agents include such local mediators as prostanoids, histamine, and VIP;7 bacteria endotoxins,8 proteolytic enzymes,9 hypoxia, and free oxygen radicals.10 When ischemia is localized in an intestinal segment, the nonischemic segment, in response to the lesion, also suffers alterations

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F. Fuertes Guiró et al.: Octreotide and Experimental Surgical Lesions

in the absorption and secretion processes.11 In cases of temporary ischemia, alterations are time-dependent and thus, during recovery the viable intestine processes of absorption and secretion are substantially altered during the first hours of revascularization.12 Somatostatin is a tetra-decapeptide hormone with paracrine functions. It inhibits some endocrine and exocrine secretions. In the small bowel, it inhibits the secretion of chloride, sodium, and water, stimulates absorption of water and electrolytes,13,14 and also diminishes intestinal peristalsis and the blood flow to the intestinal wall.15 These actions are due to a blockage of the calcium channels via either a direct action or through a modification of the cyclic nucleotides (cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP)) and by its dosedependent actions.16 Somatostatin has also been shown to have a cytoprotective effect.17 SMS 201-995, or octreotide, is a long-duration analog of somatostatin. Octreotide and its predecessor induce numerous actions in the gastrointestinal tract.18,19 Recent studies have demonstrated that octreotide is able to reduce bacterial overgrowth in hypotonic intestinal conditions.20 We hypothesize that: (1) octreotide improves intestinal absorption and secretion processes in intestinal obstruction and mesenteric ischemia, and (2) octreotide’s pharmacological actions at the intestinal level are different with respect to these intestinal pathologies.

Materials and Methods Animals One hundred and twenty Wistar rats from Charles River (Calco, taly), weighing from 400 to 450 g, were acclimated on water and Purina rat chow, ad libitum,

for 7 days prior to the experiment. The rats were randomly placed in six groups, each composed of two subgroups, as shown in Table 1. Surgery The rats were anesthetized with intraperitoneal urethane (100 mg/100 g body weight) and anesthesia was maintained during the entire experiment. By means of a medial xiphopubic laparotomy, initiated 1 cm from the xiphoid appendix, the intestinal contents were exposed, and a surgical intervention assigned to each corresponding group was performed. After the laparotomy was closed using silk 0-0 sutures, the animals were maintained at ambient temperatures between 25°C and 30°C, and hydrated with 40–80 ml/kg per 24 h of lactated Ringer’s solution, administrated subcutaneously in three doses (every 8 h), except for the treatment and control groups of total permanent mesenteric vascular occlusion, which were reoperated on at 7 h, and samples for biochemical studies were thus obtained. Thereafter, the animals were killed by a lethal dose of urethane. Surgical Interventions Complete Intestinal Occlusion (CIO). A complete intestinal occlusion was performed with a double ligature around the distal ileum, using silk 0-0, at 5 cm from the ileocecal valve. Partial Intestinal Occlusion (PIO). A partial intestinal occlusion was created by means of a double ligature of the distal ileum (at 5 cm from the ileocecal valve) interposing a 2 3 0.3 cm stainless-steel tube between the loop and the ligature, which was removed once the suture was secured.

Table 1. Design of intestinal ischemia and obstruction studies Pathology Complete intestinal occlusion (CIO) Partial intestinal occlusion (PIO) Complete intestinal occlusion 1 strangulation (CIOS) Partial permanent mesenteric vascular occlusion (PPMVO) Total permanent mesenteric vascular occlusion (TPMVO) Total temporary mesenteric vascular occlusion (TTMVO) Total

Number of animals 10 Control (CIOC) 10 Control (PIOC) 10 Control (CIOSC) 10 Control (PPMVOC) 10 Control (TPMVOC) 10 Control (TTMVOC) 60

10 Treated (CIOO) 10 Treated (PIOO) 10 Treated (CIOSO) 10 Treated (PPMVOO) 10 Treated (TPMVOO) 10 Treated (TTMVOO) 60


Complete Intestinal Occlusion with Strangulation (CIOS). The creation of a strangulated loop was performed by the ligation of the distal ileum at 5 and 10 cm from the ileocecal valve and its corresponding mesentery at the base. The animal was then observed for 15 min to ensure ischemia of the strangulated segment. Partial Permanent Mesenteric Vascular Occlusion (PPMVO). This pathology was achieved by the ligation, using silk 5-0, of the branches of the ileocecal artery and vein which supply the ileal segment between 5 and 10 cm from the ileocecal valve and the corresponding portion of the terminal arcade. The animal remained under observation for 15 min after cutoff of the blood supply to ensure that a modification in the color of the segment had taken place. Total Permanent Mesenteric Vascular Occlusion (TPMVO). Occlusion was achieved by a ligation of the superior mesenteric artery at its origin from the aorta using silk 3-0. The animal remained under observation for 15 min to verify ischemia of the loops. Total Temporary Mesenteric Vascular Occlusion (TTMVO). Occlusion was achieved by isolating and clamping the superior mesenteric artery at its origin by means of a bulldog clamp for 90 min, thus inducing the ischemia of the intestinal loops, then the laparotomic incision was temporarily closed. The clamp was then removed and the animal was observed for 15 min to ensure revascularization by means of a color change in the loops. Drug and Placebo Administration In animals of the treatment subgroups, SMS 201-995 (octreotide; Italfarmaco, Milan, Italy) was administered subcutaneously into the lateral abdomen at a dose of 100 µg/kg body weight, diluted in normal saline up to 2 ml, 8 h after a lesion developed and then every 8 h until the animal was killed, except in total permanent mesentric vascular occlusion, which received a unique dose after the laparotomy closure. The control subgroups received 2 ml of normal saline in the same periods of time as the treatment animals, except for those with total permanent mesenteric vascular occlusion, which received a unique dose after a closure of the laparotomy. Collection of Biochemical Samples at Reoperation To obtain a chemical analysis of the intestinal contents, the entire small bowel was isolated from the angle of Treitz to 5 cm above the ileocecal valve, and the con-

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tents were collected and measured in a 50-ml syringe. The volume prior to centrifugation was recorded, followed by centrifugation, after which liquids and solids were separated, and the liquid volume recorded. Samples of 5 ml were conserved at 280°C for an analysis of Na, KCl, and HCO23 . All electrolyte measurements were made prior to thawing to room temperature with the same reactive agents and the same flame photometer, and all measurements were carried out by the same technician. Histological Study After collecting the intestinal contents, a part of the intestine, corresponding to 5 cm of the tissue up to the point of obstruction in CIO, PIO, and CIOS, and 5 cm of tissue up to 5 cm of the ileocecal valve in TPMVO, PPMVO, and TTMVO was isolated, fixed in formol dilution (6%), and included in paraffin, in order to conduct the histopathologic study with hematoxylin of Harris–eosin and trichromic of Masson staining. The histological study took into consideration the following parameters: the grade of ischemia of the villi, type and extent of inflammation, the presence of edema, and the modification of the structure of the intestinal layers. Statistical Analysis The results were analyzed using an analysis of variance (ANOVA) of factorial design of 2 3 6 (two treatments, octreotide and placebo, by six pathologies, CIO, PIO, CIOS, PPVMO, PTMVO, TTMVO). The statistical difference was calculated using Fisher’s test (F-test). The study was broken down into 12 treatment groups, making nonorthogonal comparisons for an analysis of simple effects. The effects were considered significant if P , 0.01 in the F-test, and highly significant if P , 0.001 was obtained. A sensitivity analysis was performed excluding the total permanent mesenteric vascular occlusion group, since these animals were killed prior to the other subjects. A final ANOVA and comparisons were made including this group. The results are presented as the mean (SD) or mean (confidence interval of 95%). The histological score was analyzed for any significant differences with Student’s t-test. Legal and Ethical Considerations This study was carried out in accordance with the statement of the Declaration of Helsinki, the European Union specifications for animal experimentation, and the ethical principles announced by the Italian Government.

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Results Action of the Drug Independent of the Pathologies The overall average concentration of the intestinal electrolytes examined (Na, K, Cl, and HCO23 ) in the 60 animals treated with octreotide was significantly lower than in the animals in the placebo group, and the electrolytes from the total intestinal contents were also lower in the treatment group than in the control group (Table 2), and the difference was highly significant. The intestinal volume, both before and after centrifugation, was significantly lower in the treatment group than in the control group (Fig. 1). Action of Pathologies Independent of Treatments Significant differences were found among the average concentration of the intestinal levels of Na, K, Cl, and HCO23 in the six pathologies (respectively: F 5 12.10, P , 0.001; F 5 209.88, P , 0.001; F 5 66.70, P , 0.001;

Table 2. Action of octreotide independent of the pathologies Electrolyte Na Na K K Cl Cl HCO23 HCO23

Subgroup

Concentration (mEq/l)

Total contents (mEq)

Control Treated Control Treated Control Treated Control Treated

143.74 (13.71) 130.69 (7.05)*** 12.84 (5.12) 8.83 (3.44)*** 130.72 (19.75) 112.44 (9.68)*** 8.50 (2.47) 19.31 (5.98)***

1.277 (0.590) 0.632 (0.172)*** 0.104 (0.040) 0.043 (0.020)*** 1.202 (0.653) 0.544 (0.162)*** 0.070 (0.023) 0.092 (0.037)***

Intestinal concentration and total intestinal content values of four electrolytes in 60 treated and 60 control animals *** P , 0.001

F 5 89.39, P , 0.001) (Table 3). The animals with mechanical occlusion had a higher level of Na and Cl than the animals with ischemic lesions. Complete intestinal occlusion plus strangulation and total permanent mesenteric vascular occlusion produced lower intestinal concentrations of HCO23 than the other pathologies. The levels of the four electrolytes from the total intestinal contents among the six pathologies were significantly different (Na: F 5 64.13, P , 0.001; K: F 5 35.66, P , 0.001; Cl: F 5 92.28, P , 0.001; HCO23 : F 5 38.21, P , 0.001) (Table 3). The levels of Na and K from total intestinal contents were lower in the pathologies of mechanical occlusion, while less aggressive lesions (the partial intestinal occlusion and the partial permanent mesenteric vascular occlusion) produced lower levels of K. Lesions with the greatest ischemic surface (total permanent mesenteric vascular occlusion and total temporary mesenteric vascular occlusion) had the lowest levels of total intestinal HCO23 . The less aggressive lesions (partial permanent mesenteric vascular occlusion and partial intestinal occlusion) produced lower concentrations of intestinal K and total K intestinal contents than other pathologies. A complete intestinal occlusion and partial intestinal occlusion produced the highest levels of intestinal volume before and after centrifugation, with significant differences among the six pathologies (F 5 42.45, P , 0.001 and F 5 63.58, P , 0.001, respectively) (Fig. 2). Effects Between Treatment and Pathology on Intestinal Electrolyte Status The interaction test between pathology and treatment was highly significant for the intestinal concentration and total intestinal contents of all four ions (concentration: Na: F 5 13.52, P , 0.001; K: F 5 31.74, P , 0.001;

Fig. 1. Total intestinal volume both before and after centrifugation (cent) of the six pathologies independent of the treatment


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Table 3. Action of the pathologies independent of the treatments Electrolyte Na Na K K Cl Cl HCO23 HCO23

CIO

PIO

CIOS

PPMVO

TPMVO

TTMVO

[ ] (mEq/l) 140.81 (16.14) 133.78 (6.43) 147.42 (18.88) 135.33 (5.73) 130.71 (5.75) 133.26 (9.63) CT (mEq) 1.438 (0.756) 0.780 (0.273) 1.323 (0.661) 0.687 (0.119) 0.783 (0.198) 0.717 (0.327) [ ] (mEq/l) 8.22 (1.49) 6.94 (1.27) 10.38 (2.02) 18.57 (3.55) 8.53 (1.21) 12.38 (5.23) CT (mEq) 0.083 (0.047) 0.041 (0.020) 0.097 (0.057) 0.094 (0.026) 0.052 (0.014) 0.074 (0.052) [ ] (mEq/l) 133.86 (22.84) 113.49 (10.42) 140.53 (21.45) 119.29 (5.94) 112.54 (5.29) 109.79 (5.71) CT (mEq) 1.403 (0.795) 0.657 (0.265) 1.318 (0.727) 0.605 (0.104) 0.670 (0.159) 0.586 (0.254) [ ] (mEq/l) 13.74 (8.50) 16.62 (6.17) 13.31 (6.59) 6.95 (0.92) 17.23 (7.49) 15.60 (5.47) CT (mEq) 0.102 (0.033) 0.087 (0.023) 0.095 (0.015) 0.034 (0.006) 0.097 (0.038) 0.071 (0.011)

The intestinal concentration and total intestinal contents of the four electrolytes in 20 animals with each pathology are given [ ], Intestinal concentration; CT, total intestinal contents

Fig. 2. Effects of octreotide treatment by pathology on total intestinal volume before centrifugation. *** P , 0.001

Cl: F 5 30.56, P , 0.001; HCO23 : F 5 46.90, P , 0.001. Total contents: Na: F 5 42.79, P , 0.001; K: F 5 25.23, P , 0.001; Cl: F 5 57.44, P , 0.001; HCO23 : F 5 10.38, P , 0.001). When analyzing the results by specific pathology (Table 4), the Na intestinal concentration was found to be lower in the treatment subgroups than in the control ones for all pathologies except the total permanent mesenteric vascular occlusion group. These differences were also highly significant, with the exception of the partial intestinal occlusion group which showed a significant difference, and the partial permanent mesenteric vascular occlusion group where the differences were not significant (P , 0.05). The total intestinal Na levels also demonstrated a quantitative decrease in the subgroups treated with octreotide compared with the control subgroups, except for the total permanent mesenteric vascular occlusion group. The differences were highly significant for com-

plete and partial intestinal occlusion, complete intestinal occlusion plus strangulation, and total temporary mesenteric vascular occlusion. The differences were significant for partial permanent mesenteric vascular occlusion. The intestinal K concentrations and the total intestinal K contents were lower in the treatment subgroups than in the control subgroups for all pathologies, except for partial permanent mesenteric vascular occlusion. These differences were all highly significant. The concentration of intestinal Cl was lower in the treatment vs control subgroups in four of the pathologies: for complete intestinal occlusion, complete intestinal occlusion plus strangulation, and partial intestinal occlusion the differences were highly significant, and in the treatment subgroup of total temporary mesenteric vascular occlusion the differences were significant. The total intestinal Cl contents were lower in the treatment subgroups, with highly significant differences in com-

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Table 4. Effects between the treatment and pathology on the intestinal electrolyte status Pathology

[Na]

Na CT

[K]

K CT

[Cl]

Cl CT

[Bic]

Bic CT

CIOC CIOO PIOC PIOO CIOSC CIOSO PPMVOC PPMVOO TPMVOC TPMVOO TTMVOC TTMVOO

154.4 (10.6) 127.1 (4.8)*** 140.4 (4.1) 131.1 (4.7)** 159.1 (19.6) 135.6 (7.6)*** 134.3 (4.3) 127.1 (4.6)* 131.7 (5.2) 138.9 (3.6) 142.3 (1.8) 124.1 (2.9)***

2.13 (0.30) 0.73 (0.17)*** 0.98 (0.22) 0.57 (0.12)*** 1.91 (0.36) 0.72 (0.09)*** 0.89 (0.15) 0.67 (0.17)** 0.69 (0.10) 0.67 (0.13) 1.03 (0.06) 0.40 (0.03)***

9.2 (1.2) 7.2 (0.9)*** 7.9 (1.0) 5.9 (0.3)*** 11.8 (1.2) 8.8 (1.4)*** 9.2 (0.7) 7.8 (1.1)** 21.3 (2.3) 15.7 (1.9)*** 17.4 (1.2) 7.36 (0.0)***

0.12 (0.02) 0.04 (0.01)*** 0.05 (0.01) 0.02 (0.00)*** 0.14 (0.03) 0.04 (0.01)*** 0.06 (0.01) 0.04 (0.01)* 0.11 (0.02) 0.07 (0.01)*** 0.12 (0.00) 0.02 (0.00)***

154.7 (2.3) 113.0 (11.4)*** 120.4 (9.6) 106.5 (5.4)*** 159.2 (9.3) 121.8 (10.4)*** 116.4 (3.39) 108.6 (3.6)** 119.2 (4.6) 119.3 (7.2) 114.2 (2.7) 105.3 (4.0)**

2.14 (0.28) 0.66 (0.18)*** 0.85 (0.23) 0.46 (0.10)*** 1.98 (0.35) 0.65 (0.10)*** 0.77 (0.10) 0.56 (0.13)** 0.63 (0.10) 0.57 (0.10) 0.83 (0.04) 0.34 (0.03)***

5.7 (1.2) 21.7 (2.8)*** 11.0 (2.1) 22.2 (2.3)*** 7.0 (1.0) 19.6 (1.7)*** 10.1 (1.3) 24.3 (2.1)*** 6.6 (1.0) 7.2 (0.7) 10.5 (1.0) 20.7 (2.1)***

0.07 (0.02) 0.12 (0.02)*** 0.07 (0.02) 0.09 (0.01)* 0.08 (0.01) 0.10 (0.01)* 0.06 (0.01) 0.12 (0.03)*** 0.03 (0.00) 0.03 (0.00) 0.07 (0.00) 0.06 (0.01)

[ ], Intestinal concentration (mEq/l); CT, total intestinal contents (mEq) * P , 0.05; ** P , 0.01; *** P , 0.001

Fig. 3. Action of octreotide on total intestinal volume both before and after centrifugation, independent of the pathologies. *** P , 0.001

plete intestinal occlusion, complete intestinal occlusion plus strangulation, partial intestinal occlusion, and total temporary mesenteric vascular occlusion. The differences were significant for partial permanent mesenteric vascular occlusion but not significant for total permanent mesenteric vascular occlusion. There were important differences in both the concentration and total contents of intestinal HCO23 : the concentrations were higher in all the treatment subgroups except for the total permanent mesenteric vascular occlusion subgroup, with highly significant differences. The total HCO23 contents were higher in two treatment subgroups (complete intestinal occlusion and partial permanent mesenteric vascular occlusion), and the differences were highly significant, while the differences in the other subgroups were not significant.

intestinal volume, both before and after centrifugation (F 5 19.94, P , 0.001 and F 5 36.40, P , 0.001, respectively) (Figs. 3, 4). Prior to centrifugation, the total intestinal volume from complete intestinal occlusion, partial intestinal occlusion, and permanent partial mesenteric vascular occlusion was greater (P , 0.001) in the control subgroups vs the three corresponding treatment subgroups. After centrifugation, the liquid volumes were lower in the treatment subgroups of complete intestinal occlusion, partial intestinal occlusion, complete intestinal occlusion plus strangulation, partial permanent mesenteric vascular occlusion (P , 0.001), and in total temporary mesenteric vascular occlusion (P , 0.01). The differences were not significant in the total permanent mesenteric vascular occlusion subgroups.

Effects of Octreotide Treatment by Pathology on Total Intestinal Volume

Histological Results

We observed a highly significant difference between the type of pathology and type of treatment regarding the

Figures 5 to 10 show photomicrographs corresponding to an example of each condition, and the statistical results of the histological score applied to all the prepara-


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Fig. 4. Effects of octreotide treatment by pathology on total intestinal volume after centrifugation. ** P , 0.01; *** P , 0.001

b

a

tions are shown in Table 5. The control animals subjected to complete intestinal occlusion revealed lesions in the villosity compatible with a significant degree of ischemia, a situation that was not verified in the treatment animals. The vasodilation and edema in the different layers were more pronounced in the control group than in the treatment group. Although the two subgroups had acute inflammatory infiltrates in the different layers, this was more intense in the controls. The intestinal mucosal barrier disappeared on a higher surface in the controls (Fig. 5). Once these differences were quantified, they were statistically significant (Table 5).

Fig. 5a,b. Photomicrographs showing an example of small intestine treated with octreotide (a) and control (b). Pathology: CIO (hematoxilin of Harris–eosin, 310)

In almost all the treatment animals with a partial intestinal occlusion condition that were treated, the only alterations that they demonstrated was a modest acute inflammatory infiltrate in the lamina propria and in the submucosa, but the controls only presented lesions in the villosity that were compatible with the presence of a mild lesion. These quantified differences were also significant. One example of this condition is presented in Fig. 6. Most of the control animals with CIOS presented advanced intestinal ischemic lesions with a significant loss of the mucosal barrier and of the normal axial

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a

a

dimensions in comparison with the treatment group. The acute inflammatory infiltrate and edema of all layers in the latter was less than in the controls. When these differences were quantified, they were observed to be statistically significant. Two examples are shown in Fig. 7. The most important difference between the treatment and the control groups with partial permanent mesenteric vascular occlusion (Fig. 8) was the degree of ischemia of the villosities: the controls had initial

b

Fig. 6a,b. Photomicrographs showing an example of small intestine treated with octreotide (a) and control (b). Pathology: PIO (hematoxilin of Harris–eosin, 310)

b

Fig. 7a,b. Photomicrographs showing an example of small intestine treated with octreotide (a) and control (b). Pathology: CIOS (hematoxilin of Harris–eosin, 310)

ischemic lesions and there was no damage in the treatment group. The differences, once quantified, were also significant. In the total permanent mesenteric vascular occlusion animals, no differences between the treatment and control group were observed. Ischemic and massive necrotic lesions were observed in all of them, as is shown in Fig. 9. The degree of ischemic lesions of the control animals in total temporary mesenteric vascular occlusion was greater than in the treatment group, and their mucosal


a

a

427

b

Fig. 8a,b. Photomicrographs showing an example of small intestine treated with octreotide (a) and control (b). Pathology: PPMVO (hematoxilin of Harris–eosin, 310)

b Fig. 9a,b. Photomicrographs showing an example of small intestine treated with octreotide (a) and control (b). Pathology: TPMVO (Trichromic of Masson, 310) 427

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a

b Fig. 10a,b. Photomicrographs showing an example of small intestine treated with octreotide (a) and control (b). Pathology: TTMVO (hematoxilin of Harris–eosin, 310)

Table 5. Statistical results of the histological score applied to all preparations Pathology

Treated

Control

Student’s t-test

CIO PIO CIOS PPMVO TPMVO TTMVO

8.4 (2.21) 4.6 (2.14) 11.6 (2.19) 2.7 (1.46) 34.3 (3.12) 17.0 (2.46)

13.7 (2.21) 7.6 (2.14) 19.5 (2.19) 5.9 (1.46) 36.4 (3.12) 22.8 (2.46)

P , 0.0001 P , 0.005 P , 0.0001 P , 0.0001 P , 0.1 P , 0.0001

barrier was also less conserved. The inflammatory infiltrate, edema, hemorrhagic suffusion, and vasodilation in the different layers was also qualitatively and quantitatively greater and significant in the controls in comparison with most of the treatment groups (Fig. 10).

Discussion This report was designed to provide further information regarding the effects of octreotide on intestinal absorption and secretion. In physiologic conditions, the action

of octreotide is aimed at pharmacologically activating the epithelial receptors of absorption and blocking secretion at the point of onset of regulation by cAMP and Ca21,21 or in blocking the secretion of Cl (which acts as a neurotransmitter in combination with norepinephrine) at the level of neuroenterocytic junctions or in the crypts of Lieberkühn at the level of the submucosal plexus by competing with 5-hydroxytryptamine.22 In such pathologic conditions as intestinal occlusion and mesenteric ischemia, the actions of octreotide on intestinal absorption and secretion may include different mechanisms such as reduction of the intestinal blood flow by some direct action on the vessels19 (intestinal distension in the course of intestinal occlusion or mesenteric ischemia induces an increased blood flow)1,2 and a reduction in the intestinal liquids and diminished intestinal distension, and consequently, a reduction in the surface area of secretion. Octreotide may also block the vasoactive intestinal peptide (VIP), which is considered to be a mediator in the pathophysiology of intestinal occlusion and mesenteric ischemia.6,7 In mesenteric ischemia, the mechanisms of action might also be similar to an intestinal occlusion together with an inhibition of the pancreatic secretion24 (it has been postulated that the vulnerability of the intestinal mucosa to the digestive action of pancreatic proteases also increases with ischemia).9 Recently, octreotide has also shown cytoprotective effects;25 however, it also inhibits the intestinal epidermal growth factor.26 These differences in the pathophysiologic mechanisms have been demonstrated in the results obtained by grouping the animals according to pathology, independent of treatment, and observing the significant differences in the levels of the total intestinal contents of the electrolytes and total intestinal volume, as well as of the electrolyte concentrations and the quantity of water absorbed. In our report we included six diverse pathologic conditions which all alter the processes of intestinal hydroelectrolytic absorption and secretion, and we thus observed that those animals treated with octreotide presented lower levels of intestinal electrolytes and lower intestinal liquid volumes than the untreated groups except for the animals with total permanent mesenteric vascular occlusion. Octreotide affects both intestinal absorption and secretion, and this action (with different mechanisms depending on the pathology) consists of a reduction of intestinal concentrations of Na, K, Cl, and total intestinal volume and secretion of HCO23 . This significant pathology–treatment interaction demonstrates the nonadditive influence between the treatment effects and the pathological effects. Octreotide has diverse actions on intestinal absorption and secretion depending on the pathology, and vice versa. Each type of pathology influences intestinal absorption


and secretion in various ways, depending on the administration of either octreotide or a placebo. These results, which tested its action on diverse surgical models of intestinal occlusion and mesenteric ischemia, demonstrate that octreotide has a pharmacological effect which is broader than a single pathological model would demonstrate. This report suggests that the multimodel pharmacological action towards the recuperation of the intestinal processes is thus altered by various pathophysiologic mechanisms. Total permanent mesenteric vascular occlusion was the only pathology in which no response was seen to the drug, and this is understandable when we consider that ischemic occlusion is permanent and that octreotide has no action on the necrotic intestine. A statistical analysis of sensitivity to test whether or not this pathology could influence the overall results was negative. The histological results when correlated with those obtained in the biochemical study showed that the more serious the results, the greater the alteration in the electrolyte data in the intestine; consequently, the scores coincided with the severity of the condition and with the degree of alteration of the ionic values measured. Different quantitative lesions, but not qualitative ones, were observed. Essentially, the lesions observed in the control animals correspond to those observed in daily clinical practice. Therefore, according to the results obtained, octreotide reduced the consequences that the conditions have on the intestinal wall. In this sense, octreotide decreases the loss of the mucosal barrier and the appearance of signs of mucosal ischemia (Gruenhagen’s space, apice opening of the villosity, lifting and denudation, the appearance of inflammatory infiltrate, edema, and vascular dilation in all the histological layers). The reasons for the intestinal lesions observed in these conditions are considered to be multicausal. Mainly, either cellular hypoxia due to intraluminal pressure because of an increase in content, which would first affect the mucus and the flow, and worsen both hypoxia and production of oxygen free radicals (occlusive condition), or because of a temporal or permanent discontinuation of the blood irrigation of the intestine (ischemic condition), are considered to be responsible for these lesions. The second main cause of the observed lesions is the alteration in the intraluminal homeostasis (in particular the pH), with its consequent bacterial overgrowth and installation of the inflammatory cascade with all its mediators. Based on the known effects of octreotide, even though this work did not aim to directly verify them, it can thus be hypothesized that the fewer lesions observed in the treatment animals is likely to be the result of the decreased intestinal contents, due to the control of the absorption–secretion processes that decrease (a) the pressure on the intestinal wall (and at the same time, this benefit on the

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cellular integrity would make it possible to help conserve these processes), (b) its specific effects on the inflammation, and (c) its vasoactive effects, along with other reasons that are continuously being added in the literature27–29 and, as a consequence, the drug has a direct cytoprotective effect. The results of HCO23 deserve particular mention as they help to elucidate the general intestinal acid–base equilibrium. We hypothesize that the higher concentration of HCO23 observed in the treatment subgroups of complete intestinal occlusion, partial intestinal occlusion, complete intestinal occlusion plus strangulation, partial permanent mesenteric vascular occlusion, and total temporary mesenteric vascular occlusion was due, not to any direct pharmacological action, but to the indirect action of promotion of the absorption of Cl and the subsequent interchange of HCO23 /Cl, as described in physiologic conditions.30 This higher level of intestinal HCO23 assists in the regulation of pH and could explain, in part, the hypothetic reduction in intestinal bacterial flora, although we have no direct data on this. If we consider the theories regarding bacterial overgrowth and the liberation of endotoxins and prostaglandins as partial causes for alterations in intestinal absorption and secretion in the course of intestinal occlusion3 and somewhat in mesenteric ischemia,12 octreotide may be able to reduce bacterial overgrowth and thereby control both absorption and secretion.

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