Validation of Tissue Microarrays for the Study of Immunosuppressive Agent-induced Nephrotoxicity

Beom Jin Lim; Ji Hong Kim; Hyeon Joo Jeong

doi:10.4285/jkstn.2013.27.3.114

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Lim, Kim, and Jeong: Validation of Tissue Microarrays for the Study of Immunosuppressive Agent-induced Nephrotoxicity

Original Article

J Korean Soc Transplant 2013;27(3):114-120.

Published online: 11 September 2013

DOI: https://doi.org/10.4285/jkstn.2013.27.3.114

Validation of Tissue Microarrays for the Study of Immunosuppressive Agent-induced Nephrotoxicity

Beom Jin Lim, M.D.¹, Ji Hong Kim, M.D.², Hyeon Joo Jeong, M.D.^1,

¹Department of Pathology, Yonsei University College of Medicine, Seoul, Korea

²Department of Pediatrics, Yonsei University College of Medicine, Seoul, Korea

본 연구는 2012년도 대한이식학회 연구비 지원에 의하여 이루어졌음. 120-749 Tel: 02-2228-1776, Fax: 02-362-0860 E-mail: jeong10@yuhs.ac

본 연구는 대한이식학회 젊은연구자 연구비(2011)에 의해 이루어졌음

Received 8 May 201327 June 2013 Accepted 30 June 2013

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Background:

Tissue microarray analysis (TMA) is a high-throughput method for histologic evaluation, immunohistochemistry, and in situ hybridization using paraffin embedded tissue. Despite its high efficiency as an experimental tool, TMA is limited because it only contains a very small tissue fragment from each case. Therefore, the purpose of this study was to evaluate the validity of TMA in a study of nephrotoxicity caused by immunosuppressants.

Methods:

Male Sprague-Dawley rats were treated with vehicle (n=16), cyclosporine (n=23), and cyclosporine plus losartan (n=13) for a maximum of 7 weeks. After animal sacrifice, renal tissues were embedded in paraffin and processed into slides for microscopic examination using conventional methods and the TMA technique. Acute tubular injury, vascular hyaline change, and interstitial fibrosis were scored in both conventional and TMA slides. The number of interstitial macrophages was counted after ED-1 immunohistochemistry and the results also compared between conventional and TMA slides.

Results:

The degree of acute tubular injury and interstitial fibrosis showed a significant agreement between conventional and TMA methods (κ value, 0.79 and 1.00, respectively). The number of interstitial macrophages counted in conventional and TMA slides showed a significant correlation as well (r=0.934, P＜0.001). However, the degree of vascular hyaline changes showed less agreement between conventional and TMA methods (κ value, 0.40).

Conclusions:

TMA is a useful and reliable method for the study of nephrotoxicity induced by immunosuppressive agents. TMA also reflects the findings of conventional methods, especially for acute and chronic tubular and interstitial changes.

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Keywords: Tissue microarray analysis, Acute kidney injury, Tubulointerstitial fibrosis, Cyclosporine

REFERENCES

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3). Solez K, Colvin RB, Racusen LC, Haas M, Sis B, Mengel M, et al. Banff 07 classification of renal allograft pathology: updates and future directions. Am J Transplant. 2008; 8:753–60.

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Fig. 1.

A sample of tissue microarray (TMA). (A) 1:1 photography of a paraffin block and (B) a PAS-stained slide. Each TMA contained upto 30 tissue cores. Tissue cores were 3 mm in diameter and obtained from corticomedullary junction. Representative photography of cores (C, ×40; D, ×200).

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Fig. 2.

ED-1 positive cell infiltration observed in conventional and tissue microarray analysis (TMA) slides. (A) In conventional slides, the number of ED-1 positive cells in interstitium was markedly increased after cyclosporine treatment, and remained in low level after cyclosporine and losartan combination treatment. (B) The result obtained from TMA slides showed same tendency in each group. Abbreviations: HPF, high power field; CsA, cyclosporine; LSRT, losartan.

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Fig. 3.

Correlation of ED-1 positive cell counts obtained from conventional and tissue microarray analysis (TMA) slides. The number of ED-1 positive cells counted in conventional and TMA slides showed a significant correlation in each case (γ=0.934, P＜0.001). Abbreviation: HPF, high power field.

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Table 1.

Comparison of the severity of tubular injury between conventional and tissue microarray analysis slides

Tubular injury	Conventional slides		TMA slides		κ value
Tubular injury	Mild	Severe	Mild	Severe	κ value
Vehicle	16 (100)	0 (0)	16 (100)	0 (0)	0.79
CsA	5 (21.7)	18 (78.3)	7 (30.4)	16 (69.6)
CsA+LSRT	12 (92.3)	1 (7.7)	13 (100)	0 (0)

Data are presented as number (%).

Abbreviations: TMA, tissue microarray analysis; CsA, cyclosporine; LSRT, losartan.

Table 2.

Comparison of the severity of vascular hyaline change between conventional and tissue microarray analysis slides

Vascular hyaline change	Conventional slides		TMA slides		κ value
Vascular hyaline change	Mild	Severe	Mild	Severe	κ value
mVehicle	16 (100)	0 (0)	16 (100)	0 (0)	0.40
CsA	12 (52.2)	11 (47.8)	19 (82.6)	4 (17.4)
CsA+LSRT	10 (76.9)	3 (23.1)	11 (84.6)	2 (15.4)

Data are presented as number (%).

Abbreviations: TMA, tissue microarray analysis; CsA, cyclosporine; LSRT, losartan.

Table 3.

Comparison of the degree of interstitial fibrosis between conventional and tissue microarray analysis slides

Interstitial fibrosis	Conventional slides		TMA slides		κ value
Interstitial fibrosis	Mild	Severe	Mild	Severe	κ value
Vehicle	16 (100)	0 (0)	16 (100)	0 (0)	1.00
CsA	17 (73.9)	6 (26.1)	17 (73.9)	6 (26.1)
CsA+LSRT	13 (100)	0 (0)	13 (100)	0 (0)

Data are presented as number (%).

Abbreviations: TMA, tissue microarray analysis; CsA, cyclosporine; LSRT, losartan.

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