Eckhardt E R, van de Heijning B J, van Erpecum K J, Renooij W, VanBerge-Henegouwen G P
Department of Gastroenterology, University Hospital Utrecht, The Netherlands.
J Lipid Res. 1998 Mar;39(3):594-603.
The inter-mixed micellar/vesicular (non-phospholipid-associated) bile salt concentration (IMC) can be rapidly measured in model biles by centrifugal ultrafiltration, thus allowing reliable separation of vesicular and micellar cholesterol carriers by gel filtration with an elution buffer containing bile salts at the correct IMC (Donovan, J. M., and A. A. Jackson. 1993. J. Lipid Res. 34: 1121-1129). We adapted this method to the more complex human gallbladder bile and examined the relationship between cholesterol solubilization and crystallization in gallbladder biles from 10 cholesterol gallstone patients. The IMC (mean +/- SEM) was 9.67 +/- 1.97 (range 3.56-35.02) mM with significant enrichment with hydrophilic bile salt species. Upon gel filtration of these biles with an eluant buffer containing 10 major bile salts at concentrations according to their IMC, cholesterol was found to be solubilized mainly in mixed micelles. Vesicles were detected in all 10 biles after separation by KBr density gradient ultracentrifugation but in only 5 of these biles with the IMC method. Biles without vesicles had a lower CSI (1.15 +/- 0.12 vs. 1.90 +/- 0.28, P < 0.05), a higher total lipid concentration (11.9 +/- 2.3 vs. 5.9 +/- 1.1, P < 0.05), and a higher bile salt/ (bile salt + phospholipid) ratio (0.83 +/- 0.01 vs. 0.74 +/- 0.04, P = 0.07). For both IMC and ultracentrifugation methods, vesicular cholesterol concentration showed a negative correlation with crystal observation time and a positive correlation with cumulative crystal score during 21 days. Our data indicate that methods such as density gradient ultracentrifugation overestimate vesicular cholesterol solubilization in human biles.
通过离心超滤可快速测定模型胆汁中混合的胶束/囊泡（非磷脂相关）胆盐浓度（IMC），这样就能用含有正确IMC的胆盐洗脱缓冲液通过凝胶过滤可靠地分离囊泡和胶束胆固醇载体（多诺万，J.M.，和A.A.杰克逊。1993年。《脂质研究杂志》34：1121 - 1129）。我们将此方法应用于更复杂的人体胆囊胆汁，并研究了10名胆固醇结石患者胆囊胆汁中胆固醇溶解与结晶之间的关系。IMC（平均值±标准误）为9.67±1.97（范围3.56 - 35.02）mM，亲水性胆盐种类显著富集。用含有10种主要胆盐且浓度根据其IMC的洗脱缓冲液对这些胆汁进行凝胶过滤时，发现胆固醇主要溶解在混合胶束中。通过KBr密度梯度超速离心分离后，在所有10份胆汁中均检测到囊泡，但用IMC方法仅在其中5份胆汁中检测到。无囊泡的胆汁具有较低的胆固醇饱和度指数（1.15±0.12对1.90±0.28，P < 0.05）、较高的总脂质浓度（11.9±2.3对5.9±1.1，P < 0.05）以及较高的胆盐/（胆盐 + 磷脂）比值（0.83±0.01对0.74±0.04，P = 0.07）。对于IMC和超速离心方法，囊泡胆固醇浓度在21天内与晶体观察时间呈负相关，与累积晶体评分呈正相关。我们的数据表明，诸如密度梯度超速离心等方法高估了人体胆汁中囊泡胆固醇的溶解度。
