Published online Aug 7, 2016. doi: 10.3748/wjg.v22.i29.6690
Peer-review started: May 12, 2015
First decision: September 9, 2015
Revised: April 1, 2016
Accepted: May 4, 2016
Article in press: May 4, 2016
Published online: August 7, 2016
Processing time: 445 Days and 12.6 Hours
AIM: To explore the feasibility of using hypericin as an optical imaging probe with affinity for cholesterol for differential fluorescent detection of human gallstones.
METHODS: Cholesterol, mixed and pigment stones from cholecystectomy patients were incubated with hypericin or solvent. After 72 h, the stones were analysed for fluorescence (365 nm) and treated with 2-propanol/dimethyl sulfoxide for high performance liquid chromatography (HPLC) analysis. Rats with virtual gallbladder containing human cholesterol, mixed or pigment gallstones (VGHG) received 5 mg/kg hypericin or solvent and VGHG rats with cholesterol stones were given different hypericin doses (5-15 mg/kg). Twelve hours later, the stones were analysed at 365 nm. Biliary excretion and metabolites of hypericin were assessed in common bile duct (CBD) cannulated rats for 9 h using fluorospectrometry, HPLC and matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS).
RESULTS: Homogeneous high fluorescence was seen on cholesterol stones either pre-incubated with hypericin or extracted from VGHG rats receiving hypericin. Mixed stones showed a dotted fluorescent pattern, whereas pigment and solvent-treated ones lacked fluorescence. HPLC showed 7.68, 6.65 and 0.08 × 10-3 M of cholesterol in extracts from cholesterol, mixed, and pigment gallstones, respectively. Hypericin accounted for 2.0, 0.5 and 0.2 × 10-6 M in that order. On cholesterol stones from VGHG rats receiving different hypericin doses, a positive correlation was observed between dose and fluorescence. In the bile from CBD-cannulated rats, fluorescence represented 20% of the injected dose with two peaks in 9 h. HPLC analysis revealed that hypericin conjugates reached 60% of the peak area. By MALDI-TOF MS, hypericin-glucuronide was detected.
CONCLUSION: This study proves the potential use of hypericin for differential fluorescent detection of human gallstones regarding their chemical composition.
Core tip: Cholelithiasis refers to cholesterol, mixed and pigment gallstones in the gallbladder. Although the current diagnostic methods may detect their presence in the biliary system, none of them may offer an in situ differential diagnosis of gallstones regarding their chemical composition. Hypericin is a fluorophore plant pigment tending to bind membrane lipids and is excreted via bile. Because cholesterol is likely the target that hypericin binds, hypericin could become a potential optical imaging agent for differential fluorescent detection of gallstones. Such imaging procedures would allow identifying the origin, composition and formation of gallstones in patients and permit adopting strategies to minimize cholelithiasis recurrence.