pH Regulation in Human Gallbladder Bile: Study in Patients with and without Gallstones CHANTAL MARTEAU,l
BERNARD SASTRE,' NICOLA ICONOMIDIS,' HENRIPORTUGAL,' ANDRB GI~ROLAMI'
ANNE-MARIE PAULI' AND
'INSERM U. 260, Faculte' de Me'decine de la Timone, 13385 Marseille Ce'dex 5 and 'Hdpital Ste. Marguerite, 13009 Marseille, France
Samples of gallbladder bile obtained from 26 controls and 34 patients with pigment (28 cases) or cholesterol (6 cases) gallstones were studied to establish whether disturbances in regulation of the biliary pH are likely to play a role in the pathogenesis of gallstones. Samples were assayed for pH, Pco, and concentrations of sodium, bicarbonate and calcium (total and ionized). Saturation of bile in calcium carbonate was calculated. The main results of the study were as follows: (a)mean ( f S.D.) Pco, was significantly higher in gallbladder bile (6.72 f 0.36 kPa (in controls) and 7.63 2 0.29 kPa in patients) than in blood. This is consonant with previous results in animal species; it suggests that also in man a mucosal Na' H' antiport acidifies the gallbladder bile generating CO, from biliary bicarbonate. (b) Biliary pH decreased when sodium concentration increased over a range of 140 to 280 mM. The pH decreased slightly when sodium increased from 140 to 200 mM and rapidly beyond this value; the rapid pH decrease in concentrated bile was associated with bicarbonate concentrations lower than 1 to 2 mM. The results showed that bicarbonate is the main buffer of gallbladder bile. (c)The pH decrease during bile concentration was similar in patients and controls. In both groups, fully concentrated bile was unsaturated in calcium carbonate. The results suggest that gallstone formation is not due to disturbances of biliary pH regulation. However, the normal concentration process that increases Ca' concentration up to 4 mM and lowers pH values is likely to favor, in fully concentrated biles, the precipitation of calcium salts such as calcium bilirubinate. +
(HEPATOLOGY 1990;12997-1002.) Whether gallstone formation is at least due in part to disturbances of the normal concentrative properties of the gallbladder is not clearly established. In the gallbladder, biliary bicarbonate concentration and pH decrease (1-3);on the contrary, calcium concentration increases (4, 5). Recently, the consequences of these
Received June 5, 1989;accepted January 15, 1990. Address reprint requests to: Dr. Chantal Marteau, INSERM U. 260, Facult6 de MBdecine de la Timone, Bd J. Moulin, 13385 Marseille Cedex 5, France. 31/1/21184
changes on bile saturation with calcium carbonate have been studied in dogs (3). In this species, which does not spontaneously form gallstones, proton secretion from gallbladder mucosa seems to be responsible for the rapid desaturation of hepatic bile with calcium carbonate in hepatic bile. This mechanism may prevent gallstone formation because supersaturation with calcium carbonate is a prerequisite for calcium carbonate precipitation, which may serve as a nucleus for cholesterol or pigment stone formation. In man, cholelithiasis is a frequent disease and may be due to specific regulation of biliary pH and, therefore, of bile saturation with calcium carbonate as recently suggested (6). In this work, we studied pH and supersaturation with calcium carbonate of human gallbladder bile as a function of bile concentration in control subjects and patients with gallstones.
MATERIALS AND METHODS Gallbladder bile was obtained from two groups of patients who needed a perioperative cholangiography or an examination of gallbladder bile to look for cholesterol microcrystals during abdominal surgery. Bile was collected before cholecystectomy from 34 patients with gallstones and 25 patients without gallstones who were considered controls; 10 of these patients were operated for hydatid cysts, three for sequelae of acute pancreatitis (pancreatic pseudocysts), five for chronic pancreatitis and seven for various digestive disorders. Routine laboratory tests (alkaline phosphatases, ALT, conjugated bilirubin) were within the normal range for patients and controls. In patients with gallstones a preoperative cholecystogram gave a normal opacification of the gallbladder. Each bile sample was slowly withdrawn anaerobically from the gallbladder into a syringe and immediately analyzed at 37"C. Pco, and pH were measured in a Corning blood analyzer (Corning Glass Works, Corning, NY). Bicarbonate concentration was measured by titrimetry (7), which gave results similar to those calculated from the followingderivation of the Henderson-Hasselbalch equation
[HCO,-]= (Pco,. Q') x (lopH- pK2) where Q' = 0.0288 at 37" C. Pco, was expressed in Wa and p F Z = 6.1 Na' was determined by an automatic conventional procedure. Total bile salt concentration was assessed by the
997
HEPATOLOGY
MAFtTEAU ET AL.
998 280. 260. 240.
s-
220. 200.
+a
z
180. 160. 140. 120, 0
50
100
150
300
250
200
Bile Salt (mM)
A
FIG. 1. Relationship between bile salt concentration and sodium concentration. o = patients with cholesterol gallstones. y = 136 + 0 . 4 2 ~1.2 ; = 0.89.
=
controls;
=
patients with mixed gallstones;
a I12 4I 10
1
0 120
0
0
.
,
140
.
,
160
.
.
180
.
,
200
.
,
220
.
,
240
.
I
260
.
BERNARD SASTRE,' NICOLA ICONOMIDIS,' HENRIPORTUGAL,' ANDRB GI~ROLAMI'
ANNE-MARIE PAULI' AND
'INSERM U. 260, Faculte' de Me'decine de la Timone, 13385 Marseille Ce'dex 5 and 'Hdpital Ste. Marguerite, 13009 Marseille, France
Samples of gallbladder bile obtained from 26 controls and 34 patients with pigment (28 cases) or cholesterol (6 cases) gallstones were studied to establish whether disturbances in regulation of the biliary pH are likely to play a role in the pathogenesis of gallstones. Samples were assayed for pH, Pco, and concentrations of sodium, bicarbonate and calcium (total and ionized). Saturation of bile in calcium carbonate was calculated. The main results of the study were as follows: (a)mean ( f S.D.) Pco, was significantly higher in gallbladder bile (6.72 f 0.36 kPa (in controls) and 7.63 2 0.29 kPa in patients) than in blood. This is consonant with previous results in animal species; it suggests that also in man a mucosal Na' H' antiport acidifies the gallbladder bile generating CO, from biliary bicarbonate. (b) Biliary pH decreased when sodium concentration increased over a range of 140 to 280 mM. The pH decreased slightly when sodium increased from 140 to 200 mM and rapidly beyond this value; the rapid pH decrease in concentrated bile was associated with bicarbonate concentrations lower than 1 to 2 mM. The results showed that bicarbonate is the main buffer of gallbladder bile. (c)The pH decrease during bile concentration was similar in patients and controls. In both groups, fully concentrated bile was unsaturated in calcium carbonate. The results suggest that gallstone formation is not due to disturbances of biliary pH regulation. However, the normal concentration process that increases Ca' concentration up to 4 mM and lowers pH values is likely to favor, in fully concentrated biles, the precipitation of calcium salts such as calcium bilirubinate. +
(HEPATOLOGY 1990;12997-1002.) Whether gallstone formation is at least due in part to disturbances of the normal concentrative properties of the gallbladder is not clearly established. In the gallbladder, biliary bicarbonate concentration and pH decrease (1-3);on the contrary, calcium concentration increases (4, 5). Recently, the consequences of these
Received June 5, 1989;accepted January 15, 1990. Address reprint requests to: Dr. Chantal Marteau, INSERM U. 260, Facult6 de MBdecine de la Timone, Bd J. Moulin, 13385 Marseille Cedex 5, France. 31/1/21184
changes on bile saturation with calcium carbonate have been studied in dogs (3). In this species, which does not spontaneously form gallstones, proton secretion from gallbladder mucosa seems to be responsible for the rapid desaturation of hepatic bile with calcium carbonate in hepatic bile. This mechanism may prevent gallstone formation because supersaturation with calcium carbonate is a prerequisite for calcium carbonate precipitation, which may serve as a nucleus for cholesterol or pigment stone formation. In man, cholelithiasis is a frequent disease and may be due to specific regulation of biliary pH and, therefore, of bile saturation with calcium carbonate as recently suggested (6). In this work, we studied pH and supersaturation with calcium carbonate of human gallbladder bile as a function of bile concentration in control subjects and patients with gallstones.
MATERIALS AND METHODS Gallbladder bile was obtained from two groups of patients who needed a perioperative cholangiography or an examination of gallbladder bile to look for cholesterol microcrystals during abdominal surgery. Bile was collected before cholecystectomy from 34 patients with gallstones and 25 patients without gallstones who were considered controls; 10 of these patients were operated for hydatid cysts, three for sequelae of acute pancreatitis (pancreatic pseudocysts), five for chronic pancreatitis and seven for various digestive disorders. Routine laboratory tests (alkaline phosphatases, ALT, conjugated bilirubin) were within the normal range for patients and controls. In patients with gallstones a preoperative cholecystogram gave a normal opacification of the gallbladder. Each bile sample was slowly withdrawn anaerobically from the gallbladder into a syringe and immediately analyzed at 37"C. Pco, and pH were measured in a Corning blood analyzer (Corning Glass Works, Corning, NY). Bicarbonate concentration was measured by titrimetry (7), which gave results similar to those calculated from the followingderivation of the Henderson-Hasselbalch equation
[HCO,-]= (Pco,. Q') x (lopH- pK2) where Q' = 0.0288 at 37" C. Pco, was expressed in Wa and p F Z = 6.1 Na' was determined by an automatic conventional procedure. Total bile salt concentration was assessed by the
997
HEPATOLOGY
MAFtTEAU ET AL.
998 280. 260. 240.
s-
220. 200.
+a
z
180. 160. 140. 120, 0
50
100
150
300
250
200
Bile Salt (mM)
A
FIG. 1. Relationship between bile salt concentration and sodium concentration. o = patients with cholesterol gallstones. y = 136 + 0 . 4 2 ~1.2 ; = 0.89.
=
controls;
=
patients with mixed gallstones;
a I12 4I 10
1
0 120
0
0
.
,
140
.
,
160
.
.
180
.
,
200
.
,
220
.
,
240
.
I
260
.
