Photon Factory Activity Report 2018 #36 (2019)

BL-4A/2017G692, 2018G644

Evaluation of Copper Accumulation in The Liver of Chronic Bile Stasis bySynchrotron Radiation X-ray
Akihiro MATSUURA , Miyuki KINEBUCHI
Fujita Health University, Reserach Integrity Office, Quantum Beam Diagnostics, Graduate Schoolof Medicine, Molecular Pathology
Kutsukake, Toyoake 470-1192, Japan
1*11 Introduction
Copper and iron are essential nutrients for all organisms.
Both metals in excess are toxic with strong redox-activity.
Humans as well as higher vertebrates developed theingenious strategy to manage these double–edged swords;
the effective absorption and excretion system for copper,
and the strictly regulated absorption and no specificexcretion system for iron. However, medical interests ofcopper in the beginning had been focused on the toxic
effect, acute poisoning or chronic exposure by mine
pollution. Since the discovery of mammalian coppertransporters, ATP7A and ATP7B in 1993, molecularmechanisms of copper homeostasis and trafficking
pathways in physiological and pathological conditions
have been widely explored. We would like to introduce
about what copper toxicosis is, namely Wilson disease(WD), and several problems encountered in clinics andlaboratories. In this report, we show results of copper
detection in copper storage conditions by synchrotronbased technique.

22(parenchyma) and surrounding portion (stroma, including
fibrous tissues, bile ducts and vessels). To understandprecise localization of copper signals, further study ofother cases with high-resolution microbeam X-ray analysis
is undergoing.
3.2 Neonatal livers
It has been noticed that human neonates presented
physiologic newborn jaundice 2 to 4 days after birth anddisappeared at 1 to 2 weeks. The incidence is about 98% inJapanese and about 60% in Caucasians. It could beoccurred in the breast-feeding babies (breastfeedingjaundice). If the blood type of mother and baby was not
appropriate (mother-infant blood type mismatch) such as
Rh and ABO types, infants developed pathologicaljaundice due to over degradation of red blood cells
followed by hyperbilirubinemia [1, 2]. We examinednewborn livers by X-ray fluorescence (XRF) analysis. The
copper signals were stronger than control adult liver andthe copper signals were diffusely distributed throughout
the newborn liver.

2 Experiment
Tissue sections were prepared from tissue blocks asstandard pathological laboratory technique. The section tobe measured was mounted on the folder. The synchrotronradiation X-ray beam was irradiated and the fluorescence
X-ray was measured based on energy dispersive
spectrometry. Each sample was measured at least twotimes to standardize variation. Two dimentional image wascreated with Analysis software.
3 Results and Discussion
There are several examples of cooper accumulation I theliver including physiological and pathological conditions.
3.1 Chronic bile stasis
The bile duct obstruction including bile stone,cholecystitis, bile duct obstruction due to tumor can cause
chronic bile stasis. Because the bile contains a large
amount of copper, the bile static condition can induce
copper retention in the liver. An extreme example is biliaryatresia, congenital bile obstruction with unknown reason.
Therefore, we examined the liver explant at the living livertransplantation due to advanced biliary atresia, in whichchronic bile stasis caused liver cirrhosis. As shown in
Figure 1, macroscopic H&E staining view did not show
clear cirrhotic nodules. In contrast, copper signals clearly
demonstrated cirrhotic nodules. Of interest, the copper
signals were much stronger in the border of nodule

Figure 1. Copper localization in chronic bile stasis (BA).
Hematoxylin and eosin stain (left) and Copper imaging(right)
4. Conclusion
Synchrotron X-ray fluorescence analysis (SXRF) is usefulto evaluate the copper accumulation and distribution invarious copper storage conditions [3, 4]. A unique
distribution pattern found in patients with biliary atresia
implied that copper storage mechanism is different from
Wilson disease. To conduct more high-resolution analysis,
the technical improvement is important [5]. We hope thatscientists in the field of X-ray optics would continue hard
works to bring biomedical researchers to the next step,beyond “precision medicine”.

Photon Factory Activity Report 2018 #36 (2019)

Acknowledgement
We thank BL-4A user’s group members (especially, Drs.
Takahashi and Iida) for continuous help of the maintenanceof BL-4A.
References[1] MacSween’s Pathology of the Liver, 7th edition (2017).
[2] Kitazawa J et al., Pediatr Intern 46, 726 (2004).
[3] Kinebuchi M and Matsuura A. Acute Med Surg 2, 169(2015).
[4] Kinebuchi M et al., Clin Exp Immunol 139, 144 (2005).
[5] Matsuura A and Kinebuchi M. PFACR 2013b 31, 315(2014)* amatsuu@fujita-hu.ac.jp