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Journal of Lipid Research, Vol. 46, 942-948, May 2005
Copyright © 2005 by American Society for Biochemistry and Molecular Biology

Biliary cholesterol crystallization characterized by single-crystal cryogenic electron diffraction

Daphne Weihs^1,*, Judith Schmidt^*, Ilana Goldiner, Dganit Danino^2,*, Moshe Rubin, Yeshayahu Talmon^3,* and Fred M. Konikoff

^* Department of Chemical Engineering, Technion–Israel Institute of Technology, Haifa 32000, Israel
Department of Gastroenterology, Tel Aviv Sourasky Medical Center and Minerva Center for Cholesterol Gallstones and Lipid Metabolism in the Liver, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv 69978, Israel
Department of Surgery, Felsenstein Research Center, Rabin Medical Center, Petah-Tikva 49100, Israel

³ To whom correspondence should be addressed. e-mail: ishi{at}tx.technion.ac.il

Cholesterol crystals are the building blocks of cholesterol gallstones. The exact structure of early-forming crystals is still controversial. We combined cryogenic-temperature transmission electron microscopy with cryogenic-temperature electron diffraction to sequentially study crystal development and structure in nucleating model and native gallbladder biles. The growth and long-term stability of classic cholesterol monohydrate (ChM) crystals in native and model biles was determined. In solutions of model bile with low phospholipid-to-cholesterol ratio, electron diffraction provided direct proof of a novel transient polymorph that had an elongated habit and unit cell parameters differing from those of classic triclinic ChM. This crystal is exactly the monoclinic ChM phase described by Solomonov and coworkers (Biophysical J., In press) in cholesterol monolayers compressed on the air–water interface. We observed no evidence of anhydrous cholesterol crystallization in any of the biles studied.

In conclusion, classic ChM is the predominant and stable form in native and model biles. However, under certain (low phospholipid) conditions, transient intermediate polymorphs may form. These findings, documenting single-crystal analysis in bulk solution, provide an experimental approach to investigating factors influencing biliary cholesterol crystal nucleation and growth as well as other processes of nucleation and crystallization in liquid systems.

Abbreviations: ChIn, cholesterol intermediate; ChM, cholesterol monohydrate; cryo-ED, cryogenic temperature electron diffraction; cryo-TEM, cryogenic temperature transmission electron microscopy; CSI, cholesterol saturation index; TLC, total lipid concentration

Supplementary key words bile • gallstones • cholesterol monohydrate • anhydrous cholesterol

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