T hyroid FN A Cyt ology
Diffe re nt ia l Dia gnoses & Pit fa lls

Editor-in-Chief
Kennichi Kakudo, MD, PhD, FIAC
Associate Editor
Zhiyan Liu, MD, PhD
Associate Editor
Mitsuyoshi Hirokawa, MD, PhD, FIAC

T hyroid FN A Cyt ology
Diffe re nt ia l Dia gnoses & Pit fa lls

Editor-in-Chief
Kennichi Kakudo, MD, PhD, FIAC
Associate Editor
Zhiyan Liu, MD, PhD
Associate Editor
Mitsuyoshi Hirokawa, MD, PhD, FIACTwo illustrations in the front book cover and back are artworks
by Emiko Taniguchi, CT, CTIAC.

Thyroid FNA Cytology: Differential
Diagnoses and Pitfalls
Editor-in-Chief:
Kennichi Kakudo, MD, PhD, FIAC
Professor
Department of Pathology, Nara Hospital
Kindai University Faculty of Medicine
Nara, Japan
kakudo@thyroid.jp

Associate Editor:
Zhiyan Liu, MD, PhD
Associate Professor
Department of Pathology
Shandong University School of Medicine
Shandong, China
zhiyanliu@sdu.edu.cn

Associate Editor:
Mitsuyoshi Hirokawa, MD, PhD, FIAC
Department of Diagnostic Pathology and Cytology
Kuma Hospital
Kobe, Japanmhirokawa@kuma-h.or.jp

Printed by ONO KOUSOKU INSATSU CO.,LTD.
Published by Kakudo Medical Education
Copyright©2016 Kakudo Medical Education
Distributed by BookWay GLOBAL (https://bookway-global.com/)1

Contents
Preface to Print Edition
Thyroid FNA Cytology: Differential Diagnoses and Pitfalls
Kennichi Kakudo, Zhiyan Liu and Mitsuyoshi Hirakawa…………………… 8Book review

N. Paul Ohori…………………………………………………………………11

List of Frequently Used Abbreviations in This Book…………………………………………12
Chapter 1:

Factors Impacting Thyroid Fine Needle Aspiration Cytology and the
Algorithm for Cytological Diagnosis
Kennichi Kakudo………………………………………………………………14

Chapter 2:

Comparison of Diagnostic Systems: The American, British, Italian, andJapanese Reporting Systems of Thyroid FNA Cytology
Kennichi Kakudo, Zhiyan Liu and Kaori Kameyama…………………………23

Chapter 3:

Pitfalls in the Diagnosis of Papillary Thyroid Carcinoma
Xin Jing and Claire W. Michael………………………………………………30

Chapter 4:

Risk Stratification of Cases with Papillary Thyroid Carcinoma Type
Nuclear Features into Low-risk, High-risk, and Malignancy Categories.
Zhiyan Liu and Kennichi Kakudo……………………………………………39

Chapter 5:

Diagnostic Criteria of AUS/FLUS in the Bethesda Reporting System andthe Low-risk Indeterminate Category
Yubo Ren, Zhiyan Liu and Kennichi Kakudo…………………………………47

Chapter 6:

The New Tumor Entity “NIFTP (Non-invasive Follicular Thyroid
Neoplasm with Papillary-like Nuclear Features)” and the Conventional
Papillary Carcinoma
Zhiyan Liu, Shinya Satoh and Kennichi Kakudo ……………………………61

Chapter 7:

Cyst or Cystic Papillary Carcinoma
Kayoko Higuchi………………………………………………………………69

Chapter 8:

Tall Cell Variant of Papillary Thyroid Carcinoma vs. Conventional
Papillary Thyroid Carcinoma
Miyoko Higuchi………………………………………………………………78

Chapter 9:

Cribriform Variant of Papillary Thyroid Carcinoma
Ayana Suzuki…………………………………………………………………83

Chapter 10:

Papillary Carcinoma, Columnar Cell Variant - Diagnostic Pitfalls andDifferential Diagnoses
Chiung-Ru Lai…………………………………………………………………93

2

Chapter 11:

Subacute Thyroiditis or Papillary Carcinoma
Tiesheng Wang………………………………………………………………99

Chapter 12:

Hyalinizing Trabecular Tumor vs. Papillary Thyroid Carcinoma
Mitsuyoshi Hirokawa and Ayana Suzuki…………………………………… 107

Chapter 13:

Hashimoto’s Thyroiditis or Papillary Carcinoma
Yun Zhu and Tiesheng Wang……………………………………………… 115

Chapter 14:

MALT Lymphoma vs. Hashimoto Thyroiditis
Mitsuyoshi Hirokawa and Ayana Suzuki…………………………………… 125

Chapter 15:	Medullary (C Cell) Thyroid Carcinoma or Oxyphilic Follicular
Neoplasms
Yun Zhu and Tiesheng Wang……………………………………………… 132
Chapter 16:

Medullary (C Cell) Carcinoma or Metastatic Carcinoma to Thyroid
Junko Maruta ……………………………………………………………… 142

Chapter 17:

Intrathyroid Epithelial Thymoma/Carcinoma Showing Thymus-like
Differentiation vs. Poorly Differentiated Carcinoma
Mitsuyoshi Hirokawa and Ayana Suzuki…………………………………… 151

Chapter 18:

Parathyroid Adenoma and Its Differential Diagnoses
Kayoko Higuchi and Nami Takada………………………………………… 158

Chapter 19:

Risk-Classification of Follicular Pattern Lesions in Thyroid FNA Cytology(Part 1: Benign Follicular Pattern Lesions or Follicular Neoplasms)
Kennichi Kakudo, Emiko Taniguchi, Shinya Satoh and Kaori Kameyama…168

Chapter 20:

Risk-Classification of Follicular Pattern Lesions in Thyroid FNA Cytology(Part 2: Follicular Adenoma or Follicular Carcinoma)
Kennichi Kakudo, Kaori Kameyama, Keiko Inomata and Shinya Satoh… 175

Chapter 21:

Poorly Differentiated Carcinoma vs. Well Differentiated Carcinoma
Mitsuyoshi Hirokawa and Ayana Suzuki…………………………………… 185

Chapter 22:

Metastatic Renal Cell Carcinoma vs. Follicular Neoplasm
Ayana Suzuki ……………………………………………………………… 192

Chapter 23:

Biochemical Tests of Fine-needle Aspirate as an Adjunct to Cytologic
Diagnosis for Patients with Thyroid Cancer or PrimaryHyperparathyroidism
Shinya Satoh, Hiroyuki Yamashita and Kennichi Kakudo………………… 198

Chapter 24:

Infectious Thyroiditis
Claire W. Michael and Xin Jing…………………………………………… 207

3

Chapter 25:

Necrotic Background in Thyroid FNA Cytology: Anaplastic
Transformation or Infarction in Papillary Carcinoma
Kennichi Kakudo, Shinya Satoh, Yusuke Mori, Hiroyuki Yamashita……… 213

Chapter 26:

Complications of Fine Needle Aspiration Biopsy
Yasuhiro Ito and Mitsuyoshi Hirokawa…………………………………… 220

Chapter 27:

Cytological Features of Papillary Carcinoma on LBC Preparation,
Comparison with Conventional Preparation
Ayana Suzuki……………………………………………………………… 225

Chapter 28:

Pitfalls in Immunocytochemical Study Using Fine Needle Aspiration
Samples.
Junko Maruta……………………………………………………………… 232

Chapter 29:

Pitfalls in Molecular-based Diagnosis Using Thyroid Aspirates
Toru Takano………………………………………………………………… 237

Chapter 30:

Techniques of Thyroid Fine-Needle Aspiration
Hongxun Wu ……………………………………………………………… 243

Chapter 31:

Standard and Rapid Stains in Fine-Needle Aspiration Cytology and
Rapid On-Site Evaluation
Takashi Koshikawa ………………………………………………………… 251

Chapter 32:

Case Study and Mini-Tests (Multiple choice questions and differential
diagnosis)
Kennichi Kakudo, Masahiko Ura, Keiko Inomata, Ayana Suzuki, Yun Zhu,
Zhiyan Liu, Yubo Ren, Xin Jing, and Shinya Satoh……………………… 255

Suppemental Chapter 1
The Italian Reporting System for Thyroid Cytology
Guido Fadda…………………………………………………………………… 276
Suppemental Chapter 2
How to Follow Fine-needle Aspiration Biopsy-Proven Benign Thyroid
Nodules, Active Surveillance vs. Diagnostic Lobectomy for Indeterminate
Nodules and Individualized Treatments for Low-risk Thyroid Carcinomas
Kennichi Kakudo……………………………………………………………… 283
Suppemental Chapter 3
Cyst Fluid Only (CFO)
Nami Takada……………………………………………………………… 293
Suppemental Chapter 4
Thyroglossal Duct Cyst and Other Ectopic Thyroid Tissue in the Neck
Andrey Bychkov…………………………………………………………… 297

4

Suppemental Chapter 5
Diagnostic Clues for Thyroid Fine Needle Aspiration Cytology
Aki Ito and Ayana Suzuki………………………………………………… 309

5

Contributors(Alphabetical List)
Andrey Bychkov, MD PhD
Department of Pathology, Faculty of
Medicine, Chulalongkorn University,
Bangkok, Thailand
dr_bychkov@mail.com

Yasuhiro Ito, MD, PhD
Department of Surgery
Kuma Hospital
Kobe, Japan
ito01@kuma-h.or.jp

Guida Fadda, MD, MIAC
Division of Anatomic Pathology andHistology,
Catholic University,
Foundation “Agostino Gemelli”
University Hospital,
Rome, Italy
guido.fadda@unicatt.it

Xin Jing, MD
Department of Pathology
University of Michigan
Ann Arbor, Michigan, USA
xinjing@med.umich.edu

Kayoko Higuchi, MD, PhD, FIAC
Section of Anatomic Pathology
Aizawa Hospital
Matsumoto, Japan
byori-dr@ai-hosp.or.jp
Miyoko Higuchi, CT, IAC
Department of Laboratory Science
Kuma Hospital
Kobe, Japanhiguchi01@kuma-h.or.jp
Mitsuyoshi Hirokawa, MD, FIAC
Department of Diagnostic Pathology andCytology, Kuma Hospital
Kobe, Japanmhirokawa@kuma-h.or.jp
Keiko Inomata, PhD
Department of Laboratory Medicine
Yamashita Thyroid and Parathyroid Clinic
Fukuoka, Japankinomata@kojosen.com
Aki Ito, CT
Department of Laboratory Science,
Kuma Hospital, Kobe, Japanakiito@kuma-h.or.jp

6Kennichi Kakudo, MD, PhD, FIAC
Department of Pathology, Nara Hospital
Kindai University Faculty of Medicine
Nara, Japan
kakudo@thyroid.jp
Kaori Kameyama, MD, PhD
Division of Diagnostic Pathology
Keio University School of Medicine
Tokyo, Japan
kameyama@a5.keio.jp
Takashi Koshikawa, MD, PhD, FIAC
Department of Pathology
Shubun University Faculty of Nursing
Aichi, Japan
koshikawa.t@shubun.ac.jp
Chiung-Ru Lai, MD, FIAC
Department of Pathology
Taipei Veterans General Hospital
Taipei, Taiwan
crlai@vghtpe.gov.tw
Zhiyan Liu, MD, PhD
Department of Pathology
Shandong University School of Medicine
Shandong, China
zhiyanliu@sdu.edu.cn

Junko Maruta, CT, PhD
Noguchi Thyroid Clinic and HospitalFoundation
Oita, Japanjunko@noguchi-med.or.jp
Claire W. Michael, MD
Department of Pathology
University Hospitals Case Medical Center
Case Western Reserve University
Cleveland, OH, USAclaire.michael@uhhospitals.org
Yusuke Mori, MD, PhD
Department of Endocrine Surgery
Yamashita Thyroid and Parathyroid Clinic
Fukuoka, Japan
mori@kojosen.com
Yubo Ren, MD
Department of Pathology
Liaocheng People’s Hospital,
Shandong, China
renli676312@163.com
Shinya Satoh, MD, PhD
Department of Endocrine Surgery
Yamashita Thyroid and Parathyroid Clinic
Fukuoka, Japanshinya.satoh.48128@gmail.com
Ayana Suzuki, CT, CTIAC
Department of Laboratory Science
Kuma Hospital
Kobe, Japansuzuki01@kuma-h.or.jp
Nami Takada, CT
Department of Laboratory Science
Kuma Hospital
Kobe, Japan
takada01@kuma-h.or.jp

Osaka, Japanttakano@labo.med.osaka-u.ac.jp
Emiko Taniguchi, CT, CTIAC
Formerly Chief Cytotechnologist (retired)Department of Human Pathology
Wakayama Medical University
Wakayama, Japan
emi5106@yahoo.co.jp
Masahiko Ura, CT, CTIAC
Department of Pathology
Nara Hospital
Kindai University Faculty of Medicine
Nara, Japankensa@nara.med.kindai.ac.jp
Tiesheng Wang, MD
Department of Pathology
Jiangsu Institute of Nuclear Medicine
Wuxi, China.
1246513715@qq.com
Hongxun Wu, MD
Department of Ultrasound
Jiangsu Institute of Nuclear Medicine
Wuxi, China
wuhongxun@jsinm.org
Hiroyuki Yamashita, MD, PhD
Department of Endocrine Surgery
Yamashita Thyroid and Parathyroid Clinic
Fukuoka, Japan
yamaftc@kojosen.com
Yun Zhu, MD
Department of Pathology
Jiangsu Institute of Nuclear Medicine
Wuxi, China.
zhuyun@jsinm.org

Toru Takano, MD, PhD
Department of Metabolic Medicine
Osaka University
Graduate School of Medicine

7Preface to Print Edition
Thyroid FNA Cytology: Differential Diagnoses and Pitfalls
We are happy to provide you a print edition of, “Thyroid FNA Cytology: Differential
Diagnoses and Pitfalls”. As there is a file size limit in the eBook edition, some important
illustrations were deleted and all illustrations in the eBook edition were reduced in quality. To
circumvent these conditions, the print edition was published. In this print edition, we provide
you with all of the original illustrations in high quality and have incorporated some more
illustrations which were deleted from the eBook edition.
The editors proudly announce that this is the first and only textbook for thyroid FNA
cytology that incorporates borderline tumor categories in thyroid tumor classification, which
are hyalinizing trabecular adenoma identified by Carney et al.(1), UMP (well differentiatedtumor of uncertain malignant potential and follicular tumor of uncertain malignant potential)
proposed by Williams (2), the recent reclassification of some indolent tumors currently
classified as carcinoma into the borderline tumor category proposed by Kakudo et al. (3-7)
and NIFTP (non-invasive follicular thyroid neoplasm with papillary-like nuclear features),a nomenclature revision of non-invasive encapsulated follicular variant papillary thyroid
carcinoma proposed by Nikiforov et al. (8-15).
The E-Book edition was the first English-language textbook of thyroid FNA cytology
textbook published in Asia. In the print edition, we invited some more authors, and we would
like to highlight the new supplementary chapters only available in this print edition. Finally,
this print edition is a more comprehensive and international textbook than our eBook edition,“Thyroid FNA Cytology: Differential Diagnoses and Pitfalls”. The editors of the print edition,“Thyroid FNA Cytology: Differential Diagnoses and Pitfalls” thank all authors sincerely andappreciate their great efforts and contributions to this book.
The eBook version (reduced size version) is also available from the S&S Publications
and distributed by Smashwords (https://www.smashwords.com/books/view/655745).
Chief Editor: Kennichi Kakudo, MD, PhD, FIAC
Department of Pathology, Nara Hospital
Kindai University Faculty of Medicine
Professor Emeritus, Wakayama Medical University, Japan
Associate Editor: Zhiyan Liu, MD, PhD
Department of Pathology and Pathophysiology
Shandong University School of Medicine, China
Associate Editor: Mitsuyoshi Hirokawa, MD, PhD, FIAC
Department of Pathology
Kuma Hospital, Kobe, Japan8

References:1.

Carney JA, Hirokawa M, Lloyd RV, et al. Hyalinizing trabecular tumors of the thyroid gland are almost all
benign. Am J Surg Pathol 2008; 32:1877-89.

2.

Williams ED. Guest editorial: Two proposals regarding the terminology of thyroid tumors. Int J Surg Pathol
2000; 8:181-183.

3.

Kakudo K, Bai Y, Katayama S et al: Classification of follicular cell trumors of thyroid gland: Analysis involving
Japanese patients from one institute. Patholo Int 2009; 59:359-357.

4.

Liu Z, Zhou G, Nakamura M, et al. Encapsulated follicular thyroid tumor with equivocal nuclear changes, socalled well-differentiated tumor of uncertain malignant potential: a morphological, immunohistochemical, andmolecular appraisal. Cancer Sci 2011; 102:288-294.

5.

Kakudo K, Bai Y, Liu Z et al: Classification of thyroid follicular cell tumors: with special reference toborderline lesions. Endocrine J 2012; 59:1-12.

6.

Kakudo K, Bai Y, Liu Z, et al. Encapsulated papillary thyroid carcinoma, follicular variant: A misnomer. Pathol
Int 2012; 62:155-60.

7.

Nishigami K, Liu Z, Taniguchi E, et al. Cytological features of well-differentiated tumors of uncertain
malignant potential: Indeterminate cytology and WDT-UMP. Endocrine J 2012; 59:483-7.

8.

Liu J, Singh B, Tallini G, et al. Follicular variant of papillary carcinoma. A clinicopathologic study of aproblematic entity. Cancer 2006; 107:1255-64.

9.

Rivera M, Ricarte-Filho J, Knauf J, et al. Encapsulated papillary thyroid carcinoma: A clinic-pathologic study of
106 cases with emphasis on its morphologic subtypes (histologic growth pattern). Mod Pathol 2010; 23:1191200.

10. Ganly I, Wang L, Tuttle RM, et al. Invasion rather than nuclear features correlates with outcome in encapsulated
follicular tumors: further evidence for the reclassification of the encapsulated papillary thyroid carcinoma
follicular variant. Hum Pathol 2015; 46:657-64.
11. Nikiforov Y, Seethala RR, Tallini G, et al. Nomenclature revision for encapsulated follicular variant of papillary
thyroid carcinoma: A paradigm shift to reduce overtreatment of indolent tumors. JAMA Oncol, 2016, doi:
10.1001/jamaoncol.2016.0386 [Epub ahead of print].
12. Strickland KC, Howitt BE, Marquesee E, et al. The impact of non-invasive follicular variant of papillary thyroid
carcinoma on rates of malignancy for fine-needle aspiration diagnostic categories. Thyroid 2015; 25:987-92.
13. Liu X, Medici M, Kwong N, et al. Bethesda categorization of thyroid nodule cytology and prediction of thyroid
cancer type and prognosis. Thyroid 2015; [Epub ahead of print]
14. Faquin WC, Wong LQ, Afrogheh AH, et al. Impact of reclassifying noninvasive follicular variant of papillary
thyroid carcinoma on the risk of malignancy in the Bethesda system for reporting thyroid cytopathology. Cancer
Cytopathol 2015 Oct 12. doi: 10.1002/cncy.21631. [Epub ahead of print]
15. Maletta F, Massa F, Torregrossa L et al. Cytological features of “noninvasive follicular thyroid neoplasm withpapillary-like nuclear features” and their correlation with tumor histology. Hum Pathol 2016, doi: 10.1016/
j.humpath.2016.03.014. [Epub ahead of print]

9

Preface to eBook Edition
Thyroid FNA Cytology: Differential Diagnoses and Pitfalls
Thyroid fine needle aspiration (FNA) cytology is the most widely used clinical testfor patients with thyroid nodules. This can be attributed to its accuracy and reliability in
identifying high-risk patients who should undergo surgical treatments. It has been nearly 10
years since the National Cancer Institute of the United States of America proposed a reporting
system for thyroid FNA cytology (1). Following this recommendation, Italy, England, andJapan developed their own reporting systems comparable with the American (Bethesda)
system (2-4). These diagnostic systems have contributed to significantly better performancein thyroid cytology and improved communication among the different cytology practices.
All four diagnostic systems focus on the standardization of 1) diagnostic terminologies, 2)clinical management, and 3) risk of malignancy. However, there remain a few of pitfalls thatare important to address for cytopathologists to achieve a good performance in their practice.
This E-Book, “Thyroid FNA Cytology: Differential Diagnoses and Pitfalls”, focuses on how
to avoid such pitfalls in thyroid FNA cytology. Good performance in your practice can only
be achieved when you become familiar with these pitfalls and differential diagnoses in detail.
The thyroid experts included in this E-Book demonstrate how to bypass these pitfalls using
beautiful case presentations and detailed differential diagnoses based on their rich experiencesand evidences from the literature. We believe this approach is essential for establishing highlevel performance in thyroid FNA cytology, regardless of the diagnostic system used.
This E-Book is the first English textbook of thyroid FNA cytology published in Asia,and all authors are of Asian background. The editors thank all authors sincerely and appreciate
their great efforts on and contribution to this E-Book.
Chief Editor: Kennichi Kakudo, MD, PhD, FIAC
Associate Editor: Zhiyan Liu, MD, PhD
Associate Editor: Mitsuyoshi Hirokawa, MD, PhD, FIAC
References
1.
2.
3.

4.

10

Ali SZ and Cibas ES (ed). The Bethesda system for reporting thyroid cytopathology. Definitions, criteria andexplanatory notes. 2010, Springer, New York, USA.
Fadda G, Basolo F, Bondi A, et al. Cytological classification of thyroid nodules. Proposal of the SIAPEC-IAP
Italian consensus working group. Pathologica 2010; 102:405-6.
Lobo C, McQueen A, Beale T, et al. The UK Royal College of Pathologists thyroid fine-needle aspiration
diagnostic classification is a robust tool for the clinical management of abnormal thyroid nodules. Acta Cytol
2011; 55:499-506.
Kakudo K, Kameyama K, Miyauchi A, et al. Introducing the reporting system for thyroid fine-needle aspiration
cytology according to the new guidelines of the Japan Thyroid Association. Endocr J 2014; 61:539-52.

Book review
This book takes an unconventional but pragmatic approach of addressing many key
issues in thyroid cytopathology through the use of case studies in many chapters. I believe
this format is effective in engaging the reader to understand the subspecialty practice of
thyroid cytopathology. The text is authored by many well-known thyroid cytologists andpathologists, mostly from Asia. The first few chapters discuss some key principles regarding
thyroid cytopathology. The differences in the approaches to thyroid FNA cytology in various
countries such as the USA, UK, Italy, and Japan are detailed nicely and are very insightful;
they raise questions as to whether the differences are cultural or responses to external factors(e.g. medical-legal environment in the USA). While the expectation of having everyone
practice in exactly the same manner globally may be too idealistic, better understanding of oursimilarities and differences bring us closer together.
The chapters based on case studies are organized systematically into sections on
clinical summary, radiologic findings (with ultrasound images), cytologic findings, differential
diagnosis, gross and histopathologic diagnosis, discussion, and references. The images are
effective in addressing the main points of the authors. Sample chapter headings include
Diagnostic Criteria of AUS/FLUS, NIFTP (Non-invasive Follicular Thyroid Neoplasm
with Papillary-like Nuclear Features) and Conventional Papillary Carcinoma, Hyalinizing
Trabecular Tumor vs. Papillary Thyroid Carcinoma, Parathyroid Adenoma and Its Differential
Diagnosis, and Poorly Differentiated Carcinoma vs. Well Differentiated Carcinoma. In somechapters, the differential diagnoses are based on the cytologic findings, whereas in otherchapters, they are based on the histologic findings of the resected lesion. In either case, pitfalls
are discussed thoroughly.
The last chapters of this book cover the technical aspects of thyroid cytology. These
include procedural complications, liquid-based cytology, immunocytochemistry, moleculardiagnosis, FNA techniques, and staining procedures. As for the case study chapters, the images
illustrate the authors’ points very effectively. For some chapters, the technical procedures may
be based primarily on the author’s experience. Inquisitive readers are encouraged to broaden
their scope by surveying the up-to-date references provided by the authors.
In summary, I recommend this book for residents and fellows in pathology training
as well as those practicing thyroid cytopathology. The selection of chapter titles is excellent
and addresses many common dilemmas in daily practice. Case examples are well illustrated
and the thyroid cytology practitioners should find this book to be a valuable reference. While
the reader may have to adjust to the writing style of some authors, the text is relatively easy tofollow. Since some topics (e.g. diagnosis of AUS/FLUS) are controversial with many differing
opinions, readers may want to use the text as a foundation and use the up-to-date references tobuild on their own understanding of the topics.
N. Paul Ohori, M.D.
Department of Pathology
University of Pittsburgh Medical Center
Pittsburgh, Pennsylvania, USA

11

List of Frequently Used Abbreviations in This Book
ATA: 		The American Thyroid Association
ATC: 		Anaplastic Thyroid Carcinoma (Undifferentiated Carcinoma)
AUS: 		Atypia of Undetermined Significance
BRAF: 		B-Raf Proto-oncogene, Serine/Threonine Kinase
CEA: 		Carcinoembryonic Antigen
CNB: 		Core Needle Biopsy (Core Biopsy)
CFO:		
Cyst Flued Only
CT: 		
Computed Tomography
CV-PTC:
Cribriform Variant of Papillary Thyroid Carcinoma
FA: 		
Follicular Adenoma
FLUS: 		Follicular Lesion of Undetermined Significance
FNA: 		Fine Needle Aspiration
FNAC: 		Fine-Needle Aspiration Cytology
FN/HCN:
Follicular Neoplasm/Hurthle Cell Neoplasm
FN/SFN:
Follicular Neoplasm/Suspicious Follicular Neoplasm
FT-UMP:
Follicular Tumor of Uncertain Malignant Potential
fT3: 		free T3
fT4: 		free T4
FTC: 		Follicular Thyroid Carcinoma
FVPTC:
Follicular Variant Papillary Thyroid Carcinoma
HCN: 		Hurthle Cell Neoplasm
HE: 		
Hematoxylin and Eosin
HT: 		
Hashimoto’s Thyroiditis
IHC: 		Immunohistochemical Stain
IL: 		
Indeterminate Lesion
ITET/CASTLE:
Intrathyroid Epithelial Thymoma/Carcinoma Showing Thymus-like
Differentiation
LBC: 		Liquid-Based Cytology
LT: 		
Lymphocytic Thyroiditis
MGG: 		May Gruenwalds Giewsa
ML: 		
Malignant Lymphoma
MTC: 		Medullary Thyroid Cancer or C Cell Carcinoma
N/C: 		Nuclear/Cytoplasmic
NCI:
Nuclear Cytoplasmic Inclusions (Pseudoinclusions), Intranuclear Cytoplasmic
Inclusions (Pseudoinclusions)ND:
Non Diagnostic
NIFTP:
Non-Invasive Follicular Thyroid Neoplasm with Papillary-like Nuclear
Features
PAP: 		Papanicolaou
PAX8: 		Paired Box Gene 8
PDC: 		Poorly Differentiated Carcinoma
PHPT: 		Primary Hyperparathyroidism
PMC: 		Papillary Microcarcinoma
PTC: 		Papillary Thyroid Carcinoma
PTC-N		Papillary Thyroid Carcinoma Type Nuclear Features (Changes)
PTH: 		Parathyroid Hormone
PTMC: 		Papillary Thyroid Microcarcinoma
12

RAI: 		ROM: 		ROSE		
SM: 		TBSRTC:
TG: 		
TGD: 		TPO: 		TSH: 		TTF1: 		UC: 		
US: 		
WHAFFT:
WDT-UB:
WDT-UMP:
WHO: 		

Radioactive Iodine
Risk of Malignancy
Rapid On-site Evaluation
Suspicious for Malignancy
The Bethesda System for Reporting Thyroid Cytopathology
Thyroglobulin
Thyroglossal DuctThyroid Peroxidase
Thyroid Stimulating Hormone or Thyrotropin
Thyroid Transcription Factor 1
Undifferentiated Carcinoma (Anaplastic Thyroid Carcinoma)
Ultrasonography
Worrisome Histologic Alterations Following Fine needle aspiration of theThyroid
Well Differentiated Tumor of Uncertain Behavior
Well Differentiated Tumor of Uncertain Malignant Potential
World Health Organization

13

Chapter 1
Factors Impacting Thyroid Fine Needle Aspiration Cytology andthe Algorithm for Cytological Diagnosis
Kennichi Kakudo

1. The American Thyroid Association (ATA) Management Guidelines and Thyroid Fine
Needle Aspiration (FNA) Cytology
The ATA guidelines for thyroid nodules and differentiated thyroid carcinomas are
the most well-known and established guidelines, with a new edition published recently (1).
Recommendations (R7 to R24) regarding FNA cytology are also included in the 2015 ATA
guidelines (1). Recommendation 8 suggests performing FNA cytology if the thyroid nodule islarger than 1 cm and with intermediate or highly suspicious pattern on sonography. Diagnostic
FNA cytology is not recommended for thyroid nodules that are either purely cystic or smaller
than 1 cm (please refer to Supplemental Chapter 3). Thus, papillary microcarcinomas smaller
than 1 cm in diameter are clinically followed without cytological confirmation of malignancy.
According to the 2015 ATA guidelines, a higher risk of malignancy should be suspected in
cystic FNA samples from the USA than those from other countries. This is because according
to the ATA guidelines, cystic FNA samples are often taken from the solid portion (highrisk for cystic papillary thyroid carcinoma [PTC]) of the cystic nodule, and purely cystic
nodules should be excluded from FNA examination. The author of this textbook wishes
cytopathologists, particularly those practicing in other geographic areas, to bear in mind thatthe risk of malignancy of cystic FNA samples from the American system may be different
from the reader’s samples, in which a significant number of samples are from purely cystic
lesions (extremely low risk of malignancy).
Furthermore, important improvements in the clinical management of AUS/FLUS (lowrisk indeterminate) and FN/SFN (high-risk indeterminate) nodules have been included in the2015 ATA guidelines. Although diagnostic surgery is the traditional and established standard
for FN/SFN nodules in all clinical guidelines from Western countries, additional preoperativerisk stratification is recommended for SN/SFN nodules in the 2015 ATA guidelines. According
to Recommendation 16, the weak recommendation with moderate-quality evidence in FN/SFN
cytology suggests molecular testing for supplementation of malignancy risk assessment data inlieu of proceeding directly with surgery, after consideration of clinical and sonographic features.
Therefore, a diagnosis of FN/SFN on thyroid FNA cytology is no longer a compulsory orderof immediate diagnostic surgery from cytopathologists to clinical doctors. In Recommendation
15, molecular testing was added as a supplement in risk assessment for AUS/FLUS nodules, inaddition to the use of repeat FNA cytology and assessment of clinical and sonographic features.
This triage of patients with AUS/FLUS or FN/SFN nodules is already a standard practice in
Asian countries, and more than half of these patients with benign clinical tests are followed
without surgical intervention. Higher risks of malignancy in surgically treated patients with
AUS/FLUS or FN/SFN nodules have been reported in Korea (2, 3), China (4), and Japan (59), where risk stratification using cytological sub-classification, clinical tests, sonographic riskstratification, and molecular analyses are usually performed for patients with indeterminate
nodules. The increased role of conservative management using molecular testing and clinical/
radiologic risk stratification are emphasized in the 2015 ATA clinical guidelines (1).
14

Proliferation
Hyperplasia

NormalFollicular
Cell

Neoplasia
Benignor
Low
MalignantPotential

Hyperplastic
NoduleNIFTP
WDT‐UMP
WDT‐UB

Well
PoorlyUndifferentiatedDifferentiated DifferentiatedAggressive
variant PTC

Common PTC
Papillary
Microcarcinoma
PDC

FT‐UMP
Follicular
Adenoma

BenignMinimally
invasive
FTC

Precursor(borderline)

Anaplastic
Carcinoma

Widelyinvasive FTC

MalignantFigure 1. Progression and dedifferentiation of follicular cell neoplasia and precursor tumors in
the multistep carcinogenesis theory. FTC: follicular thyroid carcinoma; FT-UMP: follicular tumor of
uncertain malignant potential; NIFTP: non-invasive follicular thyroid neoplasm with papillary-like
nuclear features; PTC: papillary thyroid carcinoma; PDC: poorly differentiated carcinoma; WDT-UB:
well-differentiated tumor of uncertain behavior; WDT-UMP: well-differentiated tumor of uncertain
malignant potential.

2. The New Classification of Thyroid Tumors in the 4th Edition (2017) of the WHO
Classification of Endocrine Organs
The WHO classification of tumors serves as an international standard forhistopathologic diagnosis and clinical practice for neoplastic diseases in all organ systems.
The 3rd edition of endocrine tumor classification is under revision, and the 4th edition WHO
classification of endocrine organs will be published in 2017. The thyroid tumors included in
the contents of the 4th edition (Table 1) are essentially the same as those of the 3rd edition,
but with one significant change. The 4 th edition introduces a precursor tumor category
in the classification schema of follicular cell neoplasia to better adhere to the multistep
carcinogenesis theory (Figure 1). This category is the non-invasive follicular thyroid neoplasm
with papillary-like nuclear features (NIFTP), introduced by Nikiforov et al. in 2016, andwas formerly named the well-differentiated tumor of uncertain malignant potential (WDTUMP) by Williams in 2000, or the well-differentiated tumor with uncertain behavior (WDTUB) by Liu et al. in 2011 (10-12). All 109 cases of NIFTP in a patient series by Nikiforov
et al. showed a benign course, with average follow-up of more than 14 years after surgery
alone (without RAI treatments) (10). NIFTP is an encapsulated, follicular pattern tumor
characterized by the worrisome nuclear features of papillary carcinoma, and with no invasion.
This type of tumor shows significant observer disagreement among pathologists (13-15),
and is usually classified as a malignant tumor (encapsulated follicular variant PTC without
invasion) in Western practice, resulting in it being the most popular (10-30%) variant of PTC.
While the so-called encapsulated follicular variant PTC without invasion is a rare malignancy(<1%) in Asian practice, the majority of NIFTPs are classified as benign hyperplastic nodules
or follicular adenomas when evidence of invasion is not discernible (12). The use of a lower
cut-off threshold in judging PTC type nuclear features (PTC-N) by Western cytopathologists
compared with Asian cytopathologists may be the cause for such an increased number of
1. Factors Impacting Thyroid Fine Needle Aspiration Cytology

15malignant diagnoses (encapsulated follicular variant PTC), both histological and cytological,
in Western practice. This was further demonstrated in recent publications from the USA, inwhich a significant number of malignancies in the indeterminate categories were shown to
be NIFTP, and if NIFTP was no longer categorized as a malignant tumor, significant drops
in the rates of malignancy were observed in the AUS/FLUS and suspicious for malignancy
categories (16-18). Thus, there is a different diagnostic attitude regarding the diagnosis of PTC
between Western and Eastern pathologists, contributing to significant inter-observer variation(11-15). Cytopathologists in Western countries must refine their cytological diagnostic criteria
for PTC-N and adjust to the new WHO classification schema of PTC and precursor tumors(NIFTP).
Table 1. Classification of Thyroid Tumors in the 4th Edition

1
2
2A
3
4
4A
5
6
7
8
9
10
11
12
13
14
15
16A
16B
16C
16D
16E
16F
17A
17B
17C
17D
18
19

16

Follicular Adenoma
Hyalinizing Trabecular Adenoma/Tumor
Other Encapsulated Follicular Patterned Thyroid Tumors
Papillary Carcinoma
Follicular Carcinoma
Hurthle Cell Tumors
Poorly Differentiated Carcinoma
Undifferentiated Carcinoma
Squamous Cell Carcinoma
Medullary Carcinoma
Mixed Medullary and Follicular Cell Carcinomas
Mucoepidermoid Carcinoma
Sclerosing Mucoepidermoid Carcinoma with Eosinophilia
Mucinous Carcinoma
Ectopic Thymoma
Spindle Epithelial Tumor with Thymus-like Differentiation
Carcinoma showing Thymus-like Differentiation
Paraganglioma
Peripheral Nerve Sheath Tumors (including Schwannoma)
Benign Vascular Tumors
Angiosarcoma
Smooth Muscle Tumors (including leiomyoma and leiomyosarcoma)
Solitary Fibrous Tumor
Langerhans Cell Histiocytosis
Rosai-Dorfman Disease
Follicular Dendritic Cell Tumor
Primary Thyroid Lymphoma
Germ Cell Tumors
Secondary Tumors

3. Diagnostic Criteria and Observer Disagreement
Differences in the rates of thyroid tumor malignancy, proportions of histologicaltypes, and biological characteristics may exist among different geographic areas or patient
populations. However, the author of this chapter emphasizes the importance of applying
the diagnostic criteria of malignant thyroid tumors consistently throughout the world. Fromprevious observer variation studies on the diagnostic criteria of PTC-N, possibly irreconcilable
issues between the two types (Western and Eastern) of practice have been identified (1315). These differences may also be observed not only in PTC type malignancies, but also infollicular adenoma (FA)/follicular thyroid carcinoma (FTC) tumors. Recently, Cipriani et al.
from the USA reported that in a review of 66 FTC cases diagnosed over the past 50 years, 71%of the FTC diagnoses required reclassification, resulting in 36% of cases diagnosed as PTC,
8% as poorly differentiated carcinoma, and 27% as benign FA (19). The authors emphasized
the importance of strict diagnostic criteria for thyroid tumors (19). In this chapter, issues thatcause significant observer discrepancies in thyroid tumor diagnosis, resulting in difficulty
in cytologic-histologic correlation studies and comparisons of data in the literature, will behighlighted. These reasons underlying such observer variations may also be applied to thediagnostic criteria of thyroid FNA cytology. Because the majority of thyroid malignancies in
the current diagnostic schema do not recur or metastasize (20, 21), and a significant proportionof PTCs is proposed to be renamed NIFTP (precursor benign tumor) (10), the author of thischapter recommends applying stricter diagnostic criteria and retaining a more conservative
attitude in the diagnosis of thyroid carcinomas. This is because there are several thyroid tumors
currently classified as carcinomas, including papillary microcarcinoma, intrathyroidal PTC,
encapsulated follicular variant PTC, and minimally invasive FTC, whose recurrence rates areless than 5% after initial surgery and cancer specific death rates are at negligible levels (1, 22,
23).
4. Diagnostic Algorithm for Thyroid FNA Cytology (Figure 2)
Assessment of cellularity and specimen adequacy is the first step in evaluation of
FNA cytology specimens. Only adequate specimens must be processed for further cytologicalevaluation; however, special care must be paid to rare atypical cells in compromised specimens(sparse cellularity, air-drying artifact, bloody sample, poor fixation, or crush artifact) (pleaserefer to Chapter 5). Thus, in such cases, assessment of the presence of atypical cells should beindicated as inadequate, followed by a statement pointing out the presence of atypical cells,
clarifying that PTC cannot be ruled out, and that the specimen may be categorized into the
low-risk indeterminate category (Figure 2), such as indeterminate B: others in the Japanesesystem or AUS (PTC type atypia in suboptimal specimen) in the American system (please refer
to Chapters 2 and 5). If rare, but conclusive, malignant cells are found in a poor(low quality)
specimen, the specimen maybe classified in the suspicious for malignancy, but not malignancy,
category (Figure 2). In adequate smear samples, the following features are searched for:nuclear atypia (24, 25) (presence or absence of papillary carcinoma type nuclear atypia,
neuroendocrine carcinoma type nuclear features, high-grade nuclear features, and atypicallymphocytes) (Figures 3-4), and architectural abnormalities, such as nuclear crowding andoverlapping in follicular or trabecular clusters due to loss of cellular polarity, and dispersed
cells or microfollicles due to loss of cellular cohesiveness (Figures 5-7). The microfollicle
is emphasized in the Bethesda text book and the microfollicle consists of groups of 6-12
follicular cells in a small circle; sometimes with a small amount of thick luminal colloid(Figures 6 and 7) (24). These features are further graded depending on morphological features(mild, moderate, and marked) into benign, low-risk indeterminate, suspicious for malignancy,and malignancy categories for the PTC lineage (Figures 3 and 4), and benign, low-risk
1. Factors Impacting Thyroid Fine Needle Aspiration Cytology

17indeterminate, and high-risk indeterminate categories in the FA/FTC lineage (Figures 6-7)
(please refer to the scoring system by Kameyama in Chapter 20) (8, 25). These algorithms,
shown in Figures 2, 3 and 5, are summarized in Figure 8.
Sample preparation, fixation, and/or staining methods may require some modifications
for evaluation of these morphological features, an example of which is specimen adequacy formore than 180-320 follicular cells in LBC samples in comparison to more than 6 cell clustes inconventional smear (24, 26); please refer to Chapter 27 for liquid based cytology and Chapter
31 for different staining methods.
Adequacy judgement (sparse cellularity, air dry artifact,
bloody sample, poor fixation or crush artifact)
More than 6 clusters (more than 10 cells in one cluster)
Poor:

Inadequate or
Non-diagnostic

rare atypical cells
rare malignant cells(AUS) Atypia of
undetermined significance(low-risk indeterminate)
Suspicious for Malignancy

Adequate: Further evaluation on the following 2 features;
Presence or absence of PTC type nuclear features
Nuclear atypia and/or architectural atypia

Figure 2. An adequate sample must contain at least 6 clusters composed of more than 10 cells.
Special care must be paid to the possible presence of rare atypical cells in compromised specimens.
In such cases, the assessment for the presence of atypical cells should be pointed out as being
inadequate, followed by a statement indicating the presence of atypical cells or in the low-risk
indeterminate category, such as indeterminate B: others in the Japanese system or AUS (PTC typeatypia in suboptimal specimen) in the American system. If rare, but conclusive, malignant cells arefound in a poor specimen, then the specimen should be classified into the suspicious for malignancy
category.
Classification schema with nuclear atypia
A: Papillary carcinoma type nuclear features
AbsentBenign or FA/FTC lineage

Present

subtle, worrisome
incomplete, equivocal

Low-risk
Indeterminate

incomplete but diffuse
definite but focal

Suspicious
for Malignancy

conclusive
definite and diffuse

Malignancy

B: non-PTC type nuclear features related to other malignancies
1. salt and pepper chromatin in medullary (C cell) carcinoma,
2. high grade nuclear features in PDC and UC,
3. atypical lymphocytes in malignant lymphoma

Figure 3. The next step is an assessment of the presence or absence of PTC type nuclear features.
These are graded into four classes (negative, worrisome, definite but focal, and definite). Other
types of nuclear features related to thyroid malignancy may also be evaluated here. They are 1) salt
and pepper chromatin in medullary (C cell) carcinoma (please refer to Chapters 15 and 16), 2) highgrade nuclear features in PDC and UC (Please refer to Chapter 21), and 3) atypical lymphocytes inmalignant lymphoma (please refer to Chapter 14).
18

A

B

C

Figure 4. Papillary thyroid carcinoma type nuclear features. A: Subtle PTC type nuclear features,
including nuclear enlargement and nuclear irregularity with questionable grooves. B: Incomplete PTC
type nuclear features, such as pale (ground glass) nuclear chromatin with distinct small nucleoli in
addition to nuclear enlargement. C: Fully developed PTC type nuclear features are seen, such as
nuclear grooves and intranuclear cytoplasmic pseudo inclusions. (Conventional smear, Papanicolaou
stain, A and B: x200, C: x400)

Follicular pattern lesions without PTC-N

Nuclear Atypia (enlargement, hyperchromasia, distinct nucleoli)
and/or Architectural Atypia (cellularity, trabecular, nuclear
overlapping, microfollicles, dispersed cells)
AbsentPresent

Normal or Benign
mildly(FLUS) Follicular lesions of
undetermined significance(low-risk indeterminate)moderately

borderline (moderate-risk)

markedly(FN) Follicular neoplasm(high-risk indeterminate)

Figure 5. Follicular pattern lesions without PTC type nuclear features are graded into three classes,
benign, low-risk indeterminate (FLUS in the American or indeterminate A1, favorable benign in theJapanese system), and high-risk indeterminate category (indeterminate A3, favorable malignant in theJapanese system or FN in the American, Thy 3f in the British or TIR3B in the Italian systems).

1. Factors Impacting Thyroid Fine Needle Aspiration Cytology

19

A

B

Figure 6. Architectural abnormalities in follicular pattern lesions (FA/FTC lineage). A: Large trabecularclusters show nuclear crowding and overlapping with small round nuclei. Cellular cohesion is fairly
preserved and isolated cells are minimally found. Few macro-follicles containing colloid (orange red)
are also seen. B: There are small clusters forming microfollicles with distorted cellular polarity. Note
papillary thyroid carcinoma type nuclear features are not seen. (Conventional smear, Papanicolaou
stain, A: x200, B: x400).

A

B

Figure 7. Architectural abnormalities in follicular pattern lesions (FA/FTC lineage). A: Nuclear
crowding and overlapping are markedly seen in 3-dimensional, thick trabecular clusters. B:
Predominantly dispersed cells are shown as a loss of cellular cohesiveness. Few micro-follicles
containing colloid (orange red) are also seen. (Conventional smear, Papanicolaou stain, A: x200, B:x400).

20

Poor
Specimens

Adequate Specimens

Cystic sample or
Colloid only

Benign or normal
follicular cells

CellularatypiaAritechtural
atypia

PTC type
Nuclear atypia

Rareatypical cells
withsuspicious
US findings

Inadequate

No suspicious
US findings

Inadequate
Atypical cells

BenignMild
atypiaLow‐risk
Indeterminate

SevereatypiaHigh‐risk
Indeterminate

Insufficient atypia
for malignancy

Suspicious
For Malignancy

Malignancy

Figure 8. Algorithms for thyroid FNA cytology assessment, composed of three steps: 1) adequate
judgement, 2) nuclear atypia, and 3) architectural abnormality.

References
1.

Haugen BR, Alexander EK, Bible KC, et al. American Thyroid Association management guidelines for adultpatients with thyroid nodules and differentiated thyroid cancer. Thyroid 2015; 26:1-134.
2. Park JH, Yoon SO, Son EJ, et al. Incidence and malignancy rates of diagnoses in the Bethesda system forreporting thyroid aspiration cytology: an institutional experience. Korean J Pathol 2014; 48:133-9.
3. Hyeon J, Ahn S, Shin JH, et al. The prediction of malignant risk in the category ‘atypia of undeterminedsignificance/follicular lesion of undetermined significance’ of the Bethesda System for Reporting Thyroid
Cytopathology using subcategorization and BRAF mutation results. Cancer Cytopathol 2014; 122:368-76.
4. Zhu Y, Dai J, Lin X, et al. Fine needle aspiration of thyroid nodules: Experience in a Chinese population. J
Basic Clin Med 2015; 4:65-9.
5. Takezawa N, Sakamoto A, Komatsu K, et al. Cytological evaluation of the “indeterminate” category by the
Bethesda System for Reporting Thyroid Cytopathol. J Jpn Soc Clin Cytol 2014; 53:251-6 (in Japanese withEnglish abstract).
6. Maekawa M, Hirokawa M, Yanase Y, et al. Cytology of follicular thyroid tumors, quality assurance, anddifferential diagnosis. J Jpn Soc Clin Cytol 2010; 48-54 (in Japanese with English abstract).
7. Yamao N, Hirokawa M, Suzuki A, et al. Analysis of atypia of undetermined significance (AUS)/follicular lesionof undetermined significance (FLUS) in the Bethesda system for reporting thyroid cytopathology. J Jpn Soc
Clin Cytol 2014; 53:342-8 (in Japanese with English abstract).
8. Kameyama K, Sasaki E, Sugino K, et al. The Japanese Thyroid Association reporting system of thyroid
aspiration cytology and experience from a high-volume center, especially in indeterminate category. J Basic
Clin Med 2015; 4:70-4.
9. Sugino K, Kameyama K, Ito K. Characteristics and outcome of thyroid cancer patients with indeterminate
cytology. J Basic Clin Med 2015; 42:92-8.
10. Nikiforov YE, Seethala RR, Tallini G, et al. Nomenclature revision for encapsulated follicular variant ofpapillary thyroid carcinoma: A paradigm shift to reduce overtreatment of indolent tumors. JAMA Oncol, 2016
Apr 14. doi: 10.1001/jamaoncol.2016.0386 [Epub ahead of print]
11. Williams ED. Guest Editorial: Two proposals regarding the terminology of thyroid tumors. Int J Surg Pathol
2000; 8:181-3.
12. Liu Z, Zhou G, Nakamura M, et al. Encapsulated follicular thyroid tumor with equivocal nuclear changes, so-

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13.
14.
15.
16.
17.

18.

19.
20.
21.

22.
23.
24.
25.
26.

22

called well-differentiated tumor of uncertain malignant potential: a morphological, immunohistochemical, andmolecular appraisal. Cancer Sci 2011; 102:288-94.
Lloyd RV, Erickson LA, Casey MB, et al. Observer variation in the diagnosis of follicular variant of papillary
thyroid carcinoma. Am J Surg Pathol 2004; 28:1336-40.
Kakudo K, Katoh R, Sakamoto A, et al. Thyroid gland: international case conference. Endocrine Pathol 2002;
13:131-4.
Hirokawa M, Carney JA, Goellner JR, et al. Observer variation of encapsulated follicular lesions of the thyroid
gland. Am J Surg Pathol 2002; 26:1508-14.
Strickland KC, Howitt BE, Marquesee E, et al. The impact of non-invasive follicular variant of papillary thyroid
carcinoma on rates of malignancy for fine-needle aspiration diagnostic categories. Thyroid 2015; 25:987-92.
Faquin WC, Wong LQ, Afrogheh AH, et al. Impact of reclassifying noninvasive follicular variant of papillary
thyroid carcinoma on the risk of malignancy in the Bethesda system for reporting thyroid cytopathology. Cancer
Cytopathol 2015 Oct 12. doi: 10.1002/cncy.21631. [Epub ahead of print]
Maletta F, Massa F, Torregrossa L et al. Cytological features of “noninvasive follicular thyroid neoplasm withpapillary-like nuclear features” and their correlation with tumor histology. Hum Pathol 2016, doi: 10.1016/
j.humpath.2016.03.014. [Epub ahead of print]
Cipriani NA, Nagar S, Kaplan SP, et al. Follicular thyroid carcinoma: How have histologic diagnosis changedin the last half-century and what are the prognostic implications? Thyroid 2015; 25:1209-16.
Geffredo P, Cheung K, Roman SA, et al. Can minimally invasive follicular thyroid cancer be approached as abenign lesion? Ann Surg Oncol 2013; 20:767-72.
Piana S, Frasoldati A, Di Felice E, et al. Encapsulated well-differentiated follicular-patterned thyroid
carcinomas do not play a significant role in the fatality rates from thyroid carcinoma. Am J Surg Pathol 2010;
34:868-72.
Kakudo K, Bai Y, Liu Z, et al. Classification of thyroid follicular cell tumors: with special reference toborderline lesions. Endocr J 2012; 59:1-12.
Kakudo K, Bai Y, Liu Z, et al. Encapsulated papillary thyroid carcinoma, follicular variant: A misnormer. Pathol
Int 2012; 62:155-60.
Ali SZ, Cibas ES (ed). The Bethesda system for reporting thyroid cytopathology. Definitions, criteria andexplanatory notes. Springer, New York, USA, 2010, pp1-166.
Kakudo K, Kameyama K, Miyauchi A, et al. Introducing the reporting system for thyroid fine-needle aspiration
cytology according to the new guidelines of the Japan Thyroid Association. Endocr J 2014; 61: 539-52.
Michael CM, Pang Y, Pu RT et al: Cellular adequacy for thyroid aspirates prepared by ThinPrep: how manycells are needed? Diagn Cytopathol 2007; 35:792-97.

Chapter 2
Comparison of Diagnostic Systems: The American, British, Italian, andJapanese Reporting Systems of Thyroid FNA Cytology
Kennichi Kakudo, Zhiyan Liu and Kaori Kameyama

1. History of Diagnostic Systems in Different Countries
In the 2000s, the most popular reporting system of thyroid fine needle aspiration(FNA) cytology was the one recommended by the Papanicolaou society published in 1996 (1).
It had a single indeterminate category (Follicular Neoplasm: FN), with many cytopathologists
modifying it to their own institutional systems, which may have led to significant problems
for clinical doctors on how to decide the clinical management of their patients. In 2008,
the National Cancer Institute of the United States proposed a new reporting system forthyroid FNA cytology, the American (so-called Bethesda) system, in which standardizationof terminologies, clinical managements and risks of malignancy were established (2). Thisdiagnostic system sub-classified the indeterminate category into low-risk (AUS/FLUS) andhigh-risk (FN) categories (2, 3). Following this recommendation, the United Kingdom Royal
College of Pathologists (the British system) (4) and Italian Societies of Endocrinology andthe Italian Society for Anatomic Pathology and Cytology joint with the Italian Division of theInternational Academy of Pathology (the Italian system) (5) updated their diagnostic schema
to be comparable with the American system. These were again updated in 2014 (6, 7). The
Japan Thyroid Association (JTA) published its clinical guidelines for thyroid nodules in 2013(8), which included a reporting system for thyroid cytology (the Japanese system), basedon diagnostic systems used in high volume thyroid centers in Japan (9, 10). This system ischaracterized by more self-explanatory terminologies, and its principle of sub-classification(follicular pattern and papillary carcinoma lineages) of the indeterminate category, differingfrom other systems.
2. Differences among the Four National Systems (Table 1)
All four national reporting systems are comparable with one another, as shown in
Table 1 (11, 12). However, there are slight, but significant, differences among them. First,
cystic samples with fewer than 6 follicular cell clusters are classified into the non-diagnostic(Inadequate) category in the American system (2, 3). It is because the 2015 ATA guideline does
not recommend FNA cytology to purely cystic nodule therefore cystic cytology samples in theAmerican system usually have solid part suspicious for PTC. On the contrary, these samples
are classified into benign category in the Japanese system (9), because majority of them arefrom pure cystic nodules. Repeat examination of all cystic samples with fewer than 6 follicularcell clusters is not justified in the TIR 1C category in the Italian system (6, 11) and the Thy 1C
category in the British systems (7) due to its extremely low-risk nature (equal to or less thanbenign category). (Please refer to Supplemental Chapter 3) It is also stated in the 2015 ATA
guideline, although an FNA specimen found to have abundant colloid and few epithelial cells
may be considered nondiagnostic, this is also likely a benign biopsy. Second, the Americansystem recommends limiting the percentage of cases diagnosed as AUS/FLUS (low-risk
indeterminate category) to less than 7%, stating that overuse of this low-risk category as awastebasket should be avoided (2). These strict target numbers for the low-risk and high-risk
categories were not emphasized in the other reporting systems, although a proportion of less
2. Comparison of Diagnostic Systems

23

than 20% of indeterminate diagnoses was recommended by the Papanicolaou society in 1996,
which has been maintained as a common standard in all systems (1). The third characteristic isthe different diagnostic criteria of the high-risk indeterminate (TIR 3B) category in the Italiansystem compared with the American and British systems. A significant degree of nuclear
atypia serves as an inclusion criterion of the high-risk category (TIR 3B), and the risk ofmalignancy of the low-risk category (TIR 3A) in the Italian system was reported to be slightly
lower (5-10%) than those (5-15%) of the American and British systems (6, 11) (please refer toSupplemental Chapter 1). The fourth is the manner in dividing the indeterminate category. In
the Japanese system, the indeterminate category is divided into the follicular adenoma (FA)/follicular carcinoma (FTC) lineage and the PTC lineage, differing in principle from the otherthree diagnostic systems, where the indeterminate category is divided into low-risk (TIR 3A
in the Italian, Thy3a in the British, and AUS/FLUS in the American systems) and high-risk(TIR 3B in the Italian, Thy 3b in the British, and FN in the American systems) categories,
regardless of presence or absence of PTC type nuclear features (PTC-N). This difference wasfurther demonstrated when a significant number of PTC type malignancies were found inboth the low-risk and high-risk categories in the Italian, British, and American systems (2-7),
whereas the majority of PTC cases were found in the indeterminate B: others (PTC-lineage)
category in the Japanese system (9, 13-15). Sugino et al. reported that the risk of malignancy
in the indeterminate A category was 283/779 (36.3%), and only 61/235 cases (26.0%) of PTC
were classified as indeterminate A. On the other hand, they reported that while the risk ofmalignancy in the indeterminate B (Others) category was 202/270 (74.8%), and the majorityof PTC cases (174/235, 64.4%) were classified into the indeterminate B (15) category. In theJapanese system, the distinction between PTC type and FTC type malignancies can be moreaccurately achieved in cytology due to application of stricter criteria for PTC-N in both thehistological and cytological diagnoses by Asian pathologists (16-22).
Encapsulated non-invasive follicular pattern lesions with subtle (incomplete andquestionable) nuclear changes of PTC are traditionally classified into the benign category in
Asia, either as hyperplastic nodules or follicular adenomas (17-22). This diagnostic attitude
by Asian pathologists has helped accommodate a recent reclassification and renaming of the
non-invasive encapsulated follicular variant of PTC into the benign precursor tumor category
NIFTP (non-invasive follicular thyroid neoplasm with papillary-like nuclear features) by
Nikiforov et al. (23), as is the case with previously proposed borderline tumor terminologies,
such as well-differentiated tumor of uncertain malignant potential (WDT-UMP) by Williams(24) and well-differentiated tumor with uncertain behavior (WDT-UB) by Liu et al. (20, 25)(please refer to Chapters 1 and 6).
3. How to Subclassify Indeterminate Thyroid Nodules, and the Principle of theJapanese System (Figure 1)
The American, Italian, and British systems subclassify the indeterminate category
into low-risk and high-risk categories (Table 1). Their morphological criteria were establishedregardless of presence or absence of PTC type nuclear features. As a result, a significant
proportion of the malignancies in both the low-risk and high-risk categories are PTCs (2-7).
There are a number of explanations as to why PTCs are found in the follicular pattern lesionsof the indeterminate category (FN in the American system, Thy 3f in the British system, andTIR 3B in the Italian system). The most accepted explanation is that this is due to the lax
criteria for PTC-N in the histological diagnosis of encapsulated non-invasive follicular variant
PTC; the presence of subtle PTC-N in this variant is not uncommon, and may lead to diagnosis
of FN rather than PTC by cytology (2-4). The authors of this chapter believe it is importantto separate the two different lineages (FA/FTC and PTC) using stricter criteria for PTC type24

Inadequate (Non-Diagnostic)

<1%

<10%

Japanese (Japan Thyroid
Association)

Not Defined

Benign<1%

Risk of
Malignancy after
triage by Kakudo
et al reference
No.7

Not Defined

Normal or Benign

Risk of Malignancy
by Ali et alreference No. 2

Table 1. Comparison of the Italian, British, American (Bethesda), and Japanese reporting systems of thyroid cytopathology

I. Non Diagnostic

American (Bethesda)

Thy 1

I. Non Diagnostic

BritishNot DefinedThy 1c:

Itary

TIR 1

Low

Risk of
Malignancy by
Fadda et alreference No. 9

Non Diagnostic

TIR 1CThy 2/Thy 2cDiagnostic Category/
National Systems/ Risk
of Malignancy

Non-Diagnostic-Cystic
<3%

0-3%

TIR 2

II. Benign

Non-Neoplastic/Benign

5-15%

A1: 5-15%,
A2: 15-30%,
A3: 40-60%,
B: 40-60%<10%

III. (AUS/FLUS) Atypia ofUndetermined
Significance /Follicular
Lesion of Undetermined

15-30%

Indeterminate
A: Follicular Neoplasms(Follicular Pattern Lesions)
A1: Favor Benign,
A2: Borderline,
A3: Favor Malignant
B: Others (Non-Follicular
Pattern Lesions and PTC>80%

TIR 3A

15-30%

IV. (FN/SFN) Follicular
Neoplasm /Suspicious for
Follicular Neoplasm

Malignancy Suspected (NotConclusive for Malignancy)

Thy 3a: Neoplasm
Possible-Atypia/NonDiagnostic

TIR 3BThy 3f: Neoplasm
Possible-Suggesting
Follicular Neoplasm

60-75%

Low-Risk
Indeterminate Lesion
High-Risk
Indeterminate Lesion

V. Suspicious of
Malignancy
60-80%

>99%

TIR 4

Malignancy

Suspicious of
Malignancy

97-99%>95%

VI. Malignant

TIR 5

Thy 4: Suspicious of
Malignancy
Thy 5: Diagnostic of
Malignancy

Malignant25

2. Comparison of Diagnostic Systems

nuclear features, because the two types of well-differentiated follicular cell carcinomas havedifferent molecular alterations in their carcinogenesis, which should be useful for moleculardiagnosis. In addition, encapsulated follicular variant PTCs that have subtle PTC-N belong
to the FA/FTC lineage, and do not harbor BRAF mutation or RET/PTC rearrangements (20,26, 27) (please refer to Chapters 6 and 29). This distinction should also be useful for deciding
clinical management, because their treatment strategies are different (28).
In the Japanese system, follicular pattern lesions without PTC type nuclear features(Indeterminate A: Follicular Neoplasm) are further sub-classified into A1: favor benign,
A2: borderline, and A3: favor malignant (Figure 1), with a scoring system developed by
Kameyama et al. (14) using three parameters (cellularity, nuclear overlapping, and nuclear
atypia). Several studies from Japan have reported successful stratification of resection rates andrisks of malignancy using this system (13-15, 29). Cases with PTC type nuclear features arealso risk-stratified into three categories (Figure 1), Indeterminate B (40-60% risk), Malignancy
suspected (more than 80% risk), and Malignancy (more than 99% risk), using strict criteria
of PTC type nuclear features. With use of these criteria, Kameyama et al. were successful indifferentiating
and ofFTC
in their indeterminate
categories (14). A (follicular
Fig PTC
1: Risk
malignancy
in the indeterminate
neoplasms) and B (others) in the Japanese System

FA/FTC Lineage (without PTC-N)

Indeterminate A: FN
without PTC-N

A3: Favor Malignant
A2: Borderline
A1: Favor Benign

Risk
100%
90%
80%
70%
60%
50%
40%
30%
20%
10%
0%

PTC Lineage (with PTC-N)

Malignancy
Malignancy suspected

Indeterminate B: others(non-follicular pattern lesions)10-15%: Baseline risk of Malignancy
2-8% risk: Incidental papillary microcarcinoma in general population.

Figure 1. Risks of malignancy in Indeterminate A (Follicular Neoplasm: FA/FTC Lineage) and B(Others: PTC Lineage). On the left side, follicular pattern lesions without PTC type nuclear features(Indeterminate A: Follicular Neoplasm) are sub-classified into A1: favor benign, A2: borderline, andA3: favor malignant. Their risks of malignancy were reported to be 5-15%, 15-30%, and 40-60% using
three parameters (cellularity, nuclear overlapping, and nuclear atypia). On the right side, cases with
PTC type nuclear features are also risk-stratified into three categories, Indeterminate B (40-60% risk),
Malignancy suspected (more than 80% risk), and Malignancy (more than 99% risk). These numbers
are higher than those of the other three reporting systems, because strict criteria of PTC type nuclear
features were applied for the cytological diagnosis, not just the histological diagnosis.

4. Clinical Management of Patients with Indeterminate Nodules
The American and Italian systems recommend a diagnostic lobectomy for all patientswith high-risk (FN in the American system and TIR 3B in the Italian system) indeterminate
nodules (1-4, 6, 30). In this regard, FNA may be thought of as a screening test selecting
patients for surgery with higher probability of malignancy, although this clinical management
has been reported to result in overtreatment for the majority (more than 75%) of patients,
because they had benign nodules. Advising diagnostic surgery for all patients with high-risk
indeterminate nodules may result in a high proportion (60-80%) of surgery and a 15-30%
risk of malignancy in surgically-treated patients with high-risk indeterminate nodules (3126

33). It should be noted that all cases with indeterminate thyroid nodules (Thy 3a, Thy 3f, andThy 4) in the British system are referred to so-called multidisciplinary team discussions todetermine more appropriate clinical management (5, 7). In the JTA clinical guidelines, patientswith indeterminate thyroid nodules are also recommended to undergo further triage withother clinical tests to minimize unnecessary diagnostic surgeries for those who actually havebenign nodules (8, 9). This is a completely different approach from the clinical managementof the American and Italian systems. If all patients with indeterminate cytology are advised toundergo diagnostic surgery immediately and unselectively, then the efforts of sub-classificationof indeterminate A (FN), as in the Japanese system, or the use of multidisciplinary meetings,
as in the British system, should not be necessary. Alternatively, emphasis should be placedon stricter diagnostic criteria of the high-risk (FN) category to reduce the proportion of thiscategory and to avoid diagnostic surgery in patients with benign follicular pattern lesions (34,35). Abele and Levine suggested that compared with their experience of 5%, the 15% national
rate of the high-risk indeterminate category (FN) was in large part due to overdiagnosis (34),
and a stricter approach is essential if patients with high-risk indeterminate nodules are sent to
the operation room immediately and unselectively. The Japanese and British systems have set
up a different strategy to reduce the overtreatments of patients, demonstrating that integrated
clinical examination and watchful follow-up of thyroid nodules can spare many patientswith benign nodules from diagnostic surgery. The authors of this chapter believe that thisprocess may also assuage cytopathologists’ anxiety over how to reduce the rate of missing
FTC type malignancies in the benign category (9). This is because FTC cases with mild
cellular abnormality and the absence of an altered architectural pattern, cell crowding, and/or
a microfollicle formation may be placed in the indeterminate A1 (FN: favor benign), and thesepatients can be followed up without surgical intervention and be advised to undergo diagnostic
surgery only when other clinical tests reveal malignant characteristics. This ultimately helps
reduce missed diagnoses of FTC cases with mild cellular abnormality (please refer to Chapters
19 and 20).
5. Which Reporting System Has Better Performance?
The performance of these reporting systems likely depends on the manner of triaging
patients with thyroid nodules as well as the decision making process of clinical managementof patients with indeterminate nodules. Reports from Asian countries have demonstrated
that the application of strict triage criteria preoperatively using integrated clinical tests forpatients with indeterminate cytology may result in a low resection rate and a high proportionof malignancies in patients with indeterminate cytology (13-15, 22, 29). In clinical practice,
if avoidance of missing a malignancy is the highest priority, as opposed to minimization ofpatient overtreatment, diagnostic surgery should be advised for patients with indeterminate
nodules, because histological examination is ultimately necessary to determine whether the
nodule is benign or malignant. Under such circumstances, higher resection rates and lower
malignant risks have been reported in those with indeterminate nodules (31, 34, 35). Whateverreporting system is chosen to be used, the clinical management of patients with thyroid
nodules is ultimately decided between the clinical doctors (endocrinologists and endocrine
surgeons) and the patient, which significantly impacts the resection rates of thyroid nodules
and their risks of malignancy. Although these numbers are measures of performance, they are
dependent on the clinical guidelines applied to the patients. Thus, the thyroid FNA cytology
reporting system remains only one of the decisive factors for obtaining a good performancein clinical practice of thyroid tumors. The increased role of conservative management using
molecular testing and clinical/radiologic risk stratification is emphasized in the most recent
edition of 2015 American Thyroid Association clinical guidelines, and all international
2. Comparison of Diagnostic Systems

27

diagnostic systems for reporting thyroid cytology come close more and more, and, I believe,
they will have only negligible differences soon (28).
Conclusion
An alternative approach for establishing good performance in thyroid cytology is asecond opinion consultation with experts to establish more definitive diagnoses. This textbook,Thyroid FNA Cytology, Differential Diagnoses and Pitfalls, is designed to provide helpful
consultation and guarantees better performance in the differential diagnoses of difficult cases.
It also presents high quality evidence and helpful instructions on how to avoid the pitfalls ofcytology practice, regardless of the reporting system used.
References
1.

2.

3.
4.
5.

6.
7.
8.
9.
10.
11.
12.

13.
14.

15.
16.

28

The Papanicolaou Society of Cytopathology Task Force on Standards of Practice. Guidelines of the
Papanicolaou Society of Cytopathology for the examination of fine-needle aspiration specimens from thyroid
nodules. Diagn Cytopathol 1996; 15:84-9. (also simultaneously published in Mod Pathol 1996; 9:710-5)
Baloch ZW, LiVolsi VA, Asa SL, et al. Diagnostic terminology and morphologic criteria for cytologic diagnosisof thyroid lesions: a synopsis of the National Cancer Institute Thyroid Fine-Needle Aspiration State of the
Science Conference. Diagn Cytopathol 2008; 36:425-37.
Ali SZ and Cibas ES (ed). The Bethesda system for reporting thyroid cytopathology. Definitions, criteria andexplanatory notes. Springer, New York, USA, 2010, 1-166.
Fadda G, Basolo F, Bondi A, et al. Cytological classification of thyroid nodules. Proposal of the SIAPEC-IAP
Italian consensus working group. Pathologica 2010; 102:405-6.
Lobo C, McQueen A, Beale T, et al. The UK royal college of pathologists’ thyroid fine-needle aspiration
diagnostic classification: is a robust tool for the clinical management of abnormal thyroid nodules. Acta Cytol
2011; 55:499-506.
Nardi F, Basolo F, Crescenzi A, et al. Italian consensus for the classification and reporting of thyroid cytology. J
Endocrinol Invest 2014; 37:593-9.
Perros P, Colley S, Boelaert K, et al. Guidelines for the management of thyroid cancer Third edition British
Thyroid Association Chapt 5.1 pages 19-24; Clin Endocrinol 2014; 81, Suppl S1.
Japanese Thyroid Association Guidelines for Clinical Practice for the Management of Thyroid Nodules in Japan
2013; pp1-277, Nankodo Publishing Co. Tokyo, Japan (in Japanese).
Kakudo K, Kameyama K, Miyauchi A, et al. Introducing the reporting system for thyroid fine-needle aspiration
cytology according to the new guidelines of the Japan Thyroid Association. Endocr J 2014; 61:539-52.
Kakudo K, Kameyama K, Miyauchi A. History of thyroid cytology in Japan and reporting system recommended
by the Japan Thyroid Association. J Basic Clin Med 2013; 2:10-5.
Fadda G, Rossi ED. The 2014 Italian reporting system for thyroid cytology: Comparison with the national
reporting systems and future directions. J Basic Clin Med 2015; 4:46-51.
Bongiovanni M, Kakudo K, Nobile A. Performance comparison in the “follicular neoplasm” category between
the American, British, Italian, and Japanese system for reporting thyroid cytology. J Basic Clin Med 2015; 4:425.
Fujisawa T, Morimitsu E, Hirai T, et al. Fine needle aspiration cytology of follicular tumor, proposal for our
subclassification. J Jpn Soc Clin Cytol 2010; 49:42-7 (in Japanese with English abstract).
Kameyama K, Sasaki E, Sugino K, et al. The Japanese Thyroid Association reporting system of thyroid
aspiration cytology and experience from a high-volume center, especially in indeterminate category. J Basic
Clin Med 2015; 4:70-4.
Sugino K, Kameyama K, Ito K. Characteristics and outcome of thyroid cancer patients with indeterminate
cytology. J Basic Clin Med 2015; 4:92-8.
Lloyd RV, Erickson LA, Casey MB, et al. Observer variation in the diagnosis of follicular variant of papillary
thyroid carcinoma. Am J Surg Pathol 2004; 28:1336-40.

17. Kakudo K, Katoh R, Sakamoto A, et al. Thyroid gland: international case conference. Endocrine Pathol 2002;
13:131-4.
18. Hirokawa M, Carney JA, Goellner JR, et al. Observer variation of encapsulated follicular lesions of the thyroid
gland. Am J Surg Pathol 2002; 26:1508-14.
19. Liu J, Singh B, Tallini G, et al. Follicular variant of papillary carcinoma. A clinicopathologic study of aproblematic entity. Cancer 2006; 107:1255-64.
20. Liu Z, Zhou G, Nakamura M, et al. Encapsulated follicular thyroid tumor with equivocal nuclear changes, socalled well-differentiated tumor of uncertain malignant potential: a morphological, immunohistochemical, andmolecular appraisal. Cancer Sci 2011; 102:288-94.
21. Chan JK. Strict criteria should be applied in the diagnosis of encapsulated follicular variant of papillary thyroid
carcinoma. Am J Clin Pathol 2002; 117:16-8.
22. Y Zhu, Dai J, Lin X, et al. Fine needle aspiration of thyroid nodules: experience in a Chinese population. J
Basic Clin Med 2015; 4:65-9.
23. Nikiforov Y, Seethala RR, Tallini G, et al. Nomenclature revision for encapsulated follicular variant of papillary
thyroid carcinoma: A paradigm shift to reduce overtreatment of indolent tumors. JAMA Oncol, 2016, doi:
10.1001/jamaoncol.2016.0386 [Epub ahead of print].
24. Williams ED. Guest editorial: Two proposals regarding the terminology of thyroid tumors. Int I Surg Pathol
2000; 8:181-3.
25. Kakudo K, Bai Y, Liu Z, et al. Encapsulated papillary thyroid carcinoma, follicular variant: A misnomer. Pathol
Int 2012; 62:155-60.
26. Rivera M, Ricarte-Hilho J, Knauf J, et al. Molecular genotyping of papillary thyroid carcinoma follicularvariant according to its histological subtypes (encapsulated vs infiltrative) reveals distinct BRAF and RAS
mutation patterns. Mod Pathol 2010; 23:1191-200.
27. Howitt BE, Paulson VA, Barletta JA. Absence of BRAF V600E in non-infiltrative, non-invasive follicularvariant of papillary thyroid carcinoma. Histopathol 2015; 67:579-82.
28. Haugen BR, Alexander EK, Bible KC, et al. American Thyroid Association management guidelines for adultpatients with thyroid nodules and differentiated thyroid cancer. Thyroid 2015; 26:1-134.
29. Maekawa M, Hirokawa M, Yanase Y, et al. Cytology of follicular thyroid tumors, quality assurance, anddifferential diagnosis. J Jpn Soc Clin Cytol 2010; 49:48-54 (in Japanese with English abstract).
30. Takami H, Ito Y, Okamoto T, et al. Revisiting the guidelines by the Japanese Society of Thyroid Surgeons andJapan Association of Endocrine Surgeons: a gradual move towards consensus between Japanese and Western
practice in the management of thyroid carcinoma. World J Surg 2014; 38:2002-10.
31. Theoharis CG, Schofield KM, Hammers L, et al. The Bethesda thyroid fine-needle aspiration classification
system: year 1 at an academic institution. Thyroid 2009; 19:1215-23.
32. Ohori NP, Schoedel KE. Variability in the atypia of undetermined significance/follicular lesion of
undetermined significance diagnosis in the Bethesda system for reporting thyroid cytopathology: sources andrecommendation. Acta Cytol 2011; 55:492-8.
33. Bongiovanni M, Crippa S, Baloch Z, et al. Comparison of 5-tired and 6-tired diagnostic systems for thereporting of thyroid cytopathology: a multi-institutional study. Cancer Cytopathol 2012; 120:117-25.
34. Strickland KC, Howitt BE, Marqusee E, et al. The impact of noninvasive follicular variant of papillary thyroid
carcinoma on rates of malignancy for fine-needle aspiration diagnostic categories. Thyroid 2015; 25:987-92.
35. Abele JS, Levine RA. Diagnostic criteria and risk-adapted approach to indeterminate thyroid cytodiagnosis.
Cancer Cytopathol 2010; 118:415-22.
36. Renshaw AA, Gould EW. Reducing indeterminate thyroid FNAs. Cancer Cytopathol 2015; 123:237-43.

2. Comparison of Diagnostic Systems

29

Chapter 3
Pitfalls in the Diagnosis of Papillary Thyroid Carcinoma
Xin Jing and Claire W. Michael

Clinical History
A 10 year-old male presented with a two-year history of an enlarged neck, which wasconcerning for thyroid mass. He denied weight change, chills, excessively warm, diarrhea,
constipation, sweatiness, anxiety, and palpitations. His mother stated that he had difficulty
keeping up with his siblings and tired a little more easily.
Clinical Tests
TSH measured 1.6 (reference range: 0.30-5.50 mU/L), thyroid peroxidase antibody
measured 35 (reference range: 0-30 IU/mL), and free T4 measured 1.39 (0.76-1.70 ng/dL).
Ultrasound Findings
The right thyroid contained a complex cyst measuring 2.1 cm (transverse) x 3.2 cm(superior-inferior) x 1.3 cm (anterior-posterior). Color Doppler demonstrated vascularity in the
periphery. Central vascularity was not evident. Multiple non-shadowing punctate echoes were
present throughout the complex cyst (Figure 1).

Figure 1. Ultrasound examination of
the neck reveals a right thyroid nodule.

Cytological Findings
An ultrasound-guided fine needle aspiration (FNA) of the right thyroid nodule was
performed with preparations of Diff-Quik- and Papanicolaou-stained conventional smears.
Review of the Diff-Quik-stained conventional smears showed a cellular aspirate consistingof lymphocytes and follicular cells, which were arranged as single cells and/or irregularfragments/sheets (Figure 2). Some of the latter had a syncytial configuration with unevenly
distributed nuclei (Figure 3). Various shaped nuclei (round, oval, or elongated) and nuclear30

enlargement were evident. Some cells contained abundant cytoplasm, mimicking Hurthle cells(Figure 4). Detailed nuclear features were better appreciated upon examining the Papanicolaoustained conventional smears, including an irregular nuclear membrane, pale chromatin, anddistinct nucleoli (Figures 5 and 6). In addition, intranuclear grooves were occasionally seen
(Figure 7). However, no intranuclear pseudoinclusions (NCI) were identified.
Differential Diagnoses
1. Benign hyperplastic nodule
2. Lymphocytic (Hashimoto’s) thyroiditis
3. Follicular neoplasm/Hurthle cell neoplasm
4. Suspicious for papillary thyroid carcinoma (PTC)
5. PTC, including its variants

Figure 2. The aspirate is cellular withbackground lymphocytes, and single
and irregular sheets of follicular cells.
(Conventional smear, Diff-Quik stain,
x100)

Figure 3. Syncytial sheets with
unevenly distributed nuclei.
(Conventional smear, Diff-Quik stain,x400)

3. Pitfalls in the Diagnosis of Papillary Thyroid Carcinoma

31

Figure 4. Various shaped nuclei(round, oval, or elongated) and nuclear
enlargement are evident. Somecells contain abundant cytoplasm,
mimicking Hurthle cells. (Conventionalsmear, Diff-Quik stain, x400)

Figures 5 and 6. Irregular nuclear
membrane, pale chromatin anddistinct nucleoli are appreciated.
(Conventional smear, Papanicolaou
stain, x400)

32

Figure 7. Intranuclear grooves
are noted. (Conventional smear,
Papanicolaou stain, x400)

Discussions
It has been widely accepted that FNA is a useful triage method in distinguishingneoplastic/malignant nodules from non-neoplastic/benign nodules of thyroid. Greaterthan 90% of diagnostic accuracy has been reported while utilizing FNA to detect PTC (1).
Cytomorphological features associated with PTC have been well defined and The Bethesda
System for Reporting Thyroid Cytopathology (TBSRTC) provides diagnostic criteria of PTC
along with imaging illustrations and explanations. Briefly, the cytomorphological features of
PTC include hypercellularity, papillae and/or syncytial tissue fragments, nuclear enlargement,
oval or irregular shaped nuclei with crowding/overlapping/molding, intranuclear longitudinalgrooves and NCIs, powdery chromatin, marginally located micronucleoli, psammoma
bodies, and multinucleated giant cells (please refer to Chapters 4 and 5) (Tables 1 and 2) (2).
It is noteworthy to mention that none of these features itself is specific for PTC. We haveencountered benign hyperplastic nodule with monolayer sheets and/or papillary fragmentsof follicular cells, intranuclear grooves, and/or rarely NCIs; Hurthle cell-containing nodules(i.e. Hashimoto’s thyroiditis and Hurthle cell neoplasm) with NCIs; and benign hyperplastic
nodule with suboptimal preparation of smears showing foci of pale nuclei that mimic palenuclei with powdery chromatin. Similarly, Kini reported three aspirates obtained from
Hashimoto’s thyroiditis that contained NCIs (3) (please refer to Chapter 13). Faquin et al.
identified nuclear enlargement, nuclear grooves, fine chromatin, and distinct nucleoli, as wellas rare nuclear crowding and NCI in cyst-lining cells (4) (please refer to Chapters 5 and 7).
Other investigators also suggested that the presence of occasional intranuclear grooves should
be interpreted as a non-specific finding (5-7). Overall, extreme cautions should be excisedin the assessment of specimens with single or a few of the PTC-related features in order toavoid misinterpretation of these specimens as PTC. In addition, it is not uncommon that NCIs
may be easily appreciated in other malignant neoplasms involving the thyroid, i.e., medullary
thyroid carcinoma and metastatic melanoma to the thyroid. (please refer to Chapters 15 and
16)
There are limited literatures focusing on diagnostic pitfalls associated with FNA
cytology of PTC. Generally, the presence of overlapping cytological features between PTC
and non-PTC, as well as the over-interpretation of certain PTC-associated features arecontributing factors to misdiagnosis of PTC. We previously performed a retrospective review
of 22 thyroid aspirates with a histology-proven false diagnosis of PTC and identified several
3. Pitfalls in the Diagnosis of Papillary Thyroid Carcinoma

33

pitfalls attributed to the misdiagnosis. The pitfalls included misinterpretation of papillarylike tissue fragments and/or monolayer sheets with honeycomb arrangement as syncytial
fragments/sheets (please refer to Figure 6 in the Chapter 4), over-interpretation of suboptimal
intranuclear grooves or rare NCIs, while other PTC-associated features were minimal orabsent, and misinterpretation of the elongated or spindle cells that actually represented atypicalcyst lining cells (8) (please refer to Chapter 7). Others have reported cases of histology-proven
Hashimoto thyroiditis being over-interpreted as PTC (please refer to Chapter 13). The authorsof these studies demonstrated the pitfalls leading to over-diagnosis of PTC, including powdery
chromatin, occasional nuclear grooves or pseudoinclusions, and a paucity of background
lymphocytes. Furthermore, it was indicated that appreciation of lymphocytes infiltrating
follicular groups is a helpful approach to avoid over-diagnosis of Hashimoto thyroiditis as
PTC (9, 10). In addition, one study retrospectively reviewed three cases in which cytologic
evaluation raised suspicion PTC and the subsequent thyroidectomy revealed solitary papillary
hyperplastic nodule. The retrospective review of the FNA specimens showed worrisome
cytologic findings including broad flat sheets, 3-dimensional clusters, nonbranching papillae
with transgressing vessels, as well as mild to moderate nuclear pleomorphism and occasional
intranuclear grooves. The authors pointed out that the presence of short nonbranching papillae,
watery and inspissated colloid plus lack of NCIs may be useful features of distinguishingsolitary papillary hyperplastic nodule from PTC (11). Last but not the least, Pusztaszeri et al.
reported a case of histology-proven primary Langerhans cell histiocytosis of the thyroid inwhich FNA specimen was interpreted as suspicious for PTC due to the finding of cells with
nuclear enlargement, pale chromatin, and prominent nuclear grooves (12).
Besides aforementioned over-interpretation phenomenon, under-interpretationof variants of PTC poses a challenge to practicing cytopathologist as the classic featuresassociated with PTC may appear subtle in some of these variants (please refer to Chapters 6,
9 and 10). Among all variants of PTC, the follicular variant is the most common one, in which
FNA smear may display predominantly architectural atypia manifested by a microfollicularpattern, while PTC-associated nuclear features are inconspicuous. It is not uncommon thatsuch variant is interpreted as follicular neoplasm rather than PTC (please refer to Chapters
4 and 6). Nonetheless, there has been an on-going discussion of reclassifying non-invasive
follicular variant of PTC, which however, is beyond the scope of current discussion (pleaserefer to Chapters 4-6).
Histological Diagnosis
A total thyroidectomy was subsequently performed. Gross examination of the rightthyroid lobe revealed a 2.6 cm nodule, which abutted the anterior and isthmus margins (Figure8). Microscopic examination demonstrated morphological features consistent with PTC,
mixed follicular and Hurthle cell variant (Figure 9).

34

Figure 8. Gross examinationof the right thyroid lobe
revealed a 2.6 cm nodule.

Figure 9. Microscopic
examination identified
morphological features of
PTC, follicular and Hurthle cellvariants. (H&E stain, x400)

3. Pitfalls in the Diagnosis of Papillary Thyroid Carcinoma

35

Features

PTC

rare and an isolated finding

honeycomb arrangement withevenly spaced nuclei
rare, focal
minor portion
may be present
may be present
various amounts, homogenous,
hard or waterymay be present
rare
uniformed round or oval-shaped,
slightly enlarged, smooth contour,
fine chromatin
isolated, thin and/or incomplete

BFH

rare and an isolated finding

honeycomb arrangement withevenly spaced nuclei
rare, focal
minor portion
may be present
prominent, polymorphous population
various amounts, homogenous,
hard or waterymay be present
may be present, few nucleoli
uniformed round or oval-shaped,,slightly enlarged, smooth contour,
fine chromatin
isolated, thin and/or incomplete

LT/HT

Table 1. Differentiating features between PTC and benign non-neoplastic thyroid nodules

Papillary-like fragments

Intranuclear grooves

Psammomata bodies
Multinucleated giant cells
NucleiSyncytial-type sheets
Microfollicles
Hurthle cells
Lymphocytes
Colloidsyncytial arrangement with nuclear
crowding/overlapping/molding
moderate to abundant, diffuseprominent in FV variant
may be present
may be present
various amounts, may show abubble-gum appearance
may be present
may be present, numerous nucleolioval, elongate or irregular shaped,
variation in size, markedly enlarged,
Irregular contour, pale chromatinthick/longitudinal, accompanied byother architectural/nuclear atypia
Present along with other architectural/
nuclear atypia

Intranuclear pseudoinclusion

PTC: papillary thyroid carcinoma; BFH: benign follicular nodule; LT/HT: Lymphocytic/Hashimoto’s thyroiditis; FV: follicular variant

36

Features
papillae and/or syncytial sheets,
single cell microfollicles seen in FV

PTC

FN/HN

Table 2. Differentiating features between PTC and other neoplastic/malignant thyroid nodules

Architecture

MTC

various, commonly dispersed
single cell

plasmacytoid, spindled, small
blue cell, etc.,

none

Cells
Nucleicommonround, oval or spindled, salt andpepper chromatin, inconspicuous
nucleolirare and an isolated finding

amyloid may present
calretinin, synaptophysin,
chromogranin, CD56, CEA

enlarged cells with various amountof cytoplasm may have histocytoid
or squamoid appearance
oval, elongate or irregular shaped,
variation in size, markedly enlarged,
irregular contour, pale chromatin

scant amount or no colloid
TTF1 and thyroglobulin

microfollicles, trabeculae, single cells
Hurthle cell type may show transgressing
vessels
uniformed, normal to slightly enlarged
Hurthle cells may show marked
pleomorphism
uniformed round or oval-shaped, slightly
enlarged, smooth contour, fine chromatin.
Hurthle cells may show size variationand prominent nucleoli
isolated, thin and/or incompleteay

Intranuclear grooves
thick/longitudinal, accompanied byother architectural/nuclear atypia
Intranuclear pseudoinclusion present along with otherarchitectural/nuclear atypia
bubble-gum colloid may present
TTF1 and thyroglobulin
Background materials
Positive immunostaining

PTC: papillary thyroid carcinoma; FN/HN: follicular/Hurthle cell neoplam; MTC: medullary thyroid carcinoma; FV: follicular variant37

3. Pitfalls in the Diagnosis of Papillary Thyroid Carcinoma

References
1.

Renshaw AA. Accuracy of thyroid fine-needle aspiration using receiver operator characteristic curves. Am J
Clin Pathol 2001; 116(4):477-82.
2. Auger M SE, Yang GCH, Sanchez MA, et al. Papillary thyroid carcinoma and variants. In: The Bethesda system
for reporting thyroid cytopathology: definitions, criteria and explanatory notes [Internet]. New York: Springer;
2010. p91-115.
3. Kini SR. Thyroid cytopathology: an atlas and text. 1st ed. Philadelphia: Lippincott Williams & Wilkins; 2008.
p196-210.
4. Faquin WC, Cibas ES, Renshaw AA. “Atypical” cells in fine-needle aspiration biopsy specimens of benign
thyroid cysts. Cancer 2005; 105(2):71-9.
5. Gould E, Watzak L, Chamizo W, Albores-Saavedra J. Nuclear grooves in cytologic preparations. A study of theutility of this feature in the diagnosis of papillary carcinoma. Acta Cytol 1989; 33(1):16-20.
6. Rupp M, Ehya H. Nuclear grooves in the aspiration cytology of papillary carcinoma of the thyroid. Acta Cytol
1989; 33(1):21-6.
7. Francis IM, Das DK, Sheikh ZA, et al. Role of nuclear grooves in the diagnosis of papillary thyroid carcinoma.
A quantitative assessment on fine needle aspiration smears. Acta Cytol 1995; 39(3):409-15.
8. Jing X, Michael CW. Potential pitfalls for false suspicion of papillary thyroid carcinoma: a cytohistologic
review of 22 cases. Diagn Cytopathol 2012; 40(Suppl 1):E74-9.
9. Haberal AN, Toru S, Ozen O, et al. Diagnostic pitfalls in the evaluation of fine needle aspiration cytology of the
thyroid: correlation with histopathology in 260 cases. Cytopathol 2009; 20(2):103-8.
10. Harvey AM, Truong LD, Mody DR. Diagnostic pitfalls of Hashimoto’s/lymphocytic thyroiditis on fine-needle
aspirations and strategies to avoid overdiagnosis. Acta cytologica 2012; 56(4):352-60.
11. Khurana KK, Baloch ZW, LiVolsi VA. Aspiration cytology of pediatric solitary papillary hyperplastic thyroid
nodule. Arch Pathol Lab Med 2001; 125(12):1575-8.
12. Pusztaszeri MP, Sauder KJ, Cibas ES, et al. Fine-needle aspiration of primary Langerhans cell histiocytosis of
the thyroid gland, a potential mimic of papillary thyroid carcinoma. Acta Cytologica 2013; 57(4):406-12.

38

Chapter 6
The New Tumor Entity “NIFTP (Non-invasive Follicular Thyroid
Neoplasm with Papillary-like Nuclear Features)” and Conventional
Papillary Carcinoma
Zhiyan Liu, Shinya Satoh and Kennichi Kakudo

Brief Clinical Summary
The patient was a 72-year-old female who has a history of stroke. During follow up,
she was found to have a nodule in the right lobe of the thyroid. She was referred to our hospitalfor a further check-up, and under ultrasound guidance, a fine needle aspiration (FNA) was
performed for the 18 mm thyroid nodule (Figure 1).
Ultrasound Findings
Ultrasound detected a well-circumscribed nodule in the right lobe of the thyroid(Figure 1). The nodule was solid with minor cystic change. Neither irregular margin
suggestive of invasion into thyroid parenchyma nor invasion to thyroid capsule was found. No
calcification was identified in the nodule. A clinical diagnosis of a benign adenomatous nodule
was made based on the ultrasound images.

Figure 1. Ultrasound images show a well-demarcated thyroid nodule.

Cytological Findings:
A moderate number of follicular cells in follicular pattern clusters were seen in the
Papanicolaou-stained conventional smear (Figures 2-6). No macrophages suggesting cystic
change were noted in the smear background. These follicular cell clusters appeared in loosely
cohesive with moderate nuclear overlapping (Figures 2-4) or in large trabecular clusters(Figure 5). Many isolated cells with ill-defined pale cytoplasm were also noted (Figure 4). The
tumor cell nuclei were irregular in shape (Figures 3 and 5), and moderately enlarged, and they
had a few grooves with fine powdery chromatin (Figures 2-6). Nuclear crowding, overlapping,and molding were conspicuous as shown in the Figure 5. A definite nuclear cytoplasmic
6. The New Tumor Entity “NIFTP”

61inclusion (NCI) body was found in only one sample (Figure 6), but small questionable
nuclear inclusions or vacuoles (red arrows in Figures 3 and 4) and a few of nuclear grooves(yellow arrows in Figures 3 and 4) were worrisome nuclear features, suggesting papillary
thyroid carcinoma (PTC) type malignancy. Nucleoli were small and inconspicuous. Neither
psammoma bodies nor necrosis was identified in the smear background.
List of Differential Diagnoses
1. Benign, follicular adenoma
2. Benign, atypical follicular adenoma (FA)
3. Benign, Hashimoto thyroiditis
4. Indeterminate A, follicular neoplasm (FN by the Bethesda system, TIR 3B by
Italian system or Thy 3f by British system)
5. Indeterminate B, others, PTC cannot rule out (AUS by the Bethesda system, TIR
3A by the Italian system or Thy 3a by the British system)
6. Suspicious for PTC, follicular variant
7. Malignant, PTC, follicular variant
8. Malignant, PTC, conventional

Figure 2. Moderate nuclear enlargement
is seen in microfollicular or loosely
arranged clusters. (Conventional smear,
Papanicolaou stain, x400)

Figure 3. Small nuclear vacuoles (redarrows) and grooves (yellow arrows)
are shown in loosely cohesive follicular
cells. (Conventional smear, Papanicolaou
stain, x400)

62

Figure 4. Note a small nuclear
vacuole or inclusion (red arrow) andgrooves (yellow arrows) in a loosely
cohesive large cluster of follicular
cells. Numerous isolated cells arealso seen in the smear background.
(Conventional smear, Papanicolaou
stain, x400)

Figure 5. A large follicular cell cluster
forms trabecular arrangement. Nuclear
crowding, molding and irregularity
are shown. (Conventional smear,
Papanicolaou stain, x400)

Figure 6. A large nucleus with acytoplasmic inclusion is shown inthe center. This is the only nuclear
inclusion identified in the samples.
(Conventional smear, Papanicolaou
stain, x400)

6. The New Tumor Entity “NIFTP”

63Differential Diagnosis:
Differential diagnoses include atypical FA, atypical follicular cells in Hashimoto
thyroiditis, conventional PTC, follicular variant PTC, well differentiated tumor of uncertain
malignant potential (WDT-UMP), NIFTP, and follicular thyroid carcinoma (FTC).
Hashimoto thyroiditis can be ruled out by the absence of lymphocytes in the smear
background. Characteristic oxyphilic change (Hurthle) of follicular cells in Hashimoto
thyroiditis is absent (please refer to Chapter 13).
Moderate nuclear enlargement, nuclear shape irregularity, and small nuclear vacuoles in thiscase were worrisome nuclear features of PTC type malignancy. NCIs are seen in more than
80% of conventional PTC (please refer to Chapters 3 and 4), however, they are not specific for
PTC type malignancy because they are also seen in C cell (medullary) carcinomas (please refer
to Chapters 15 and 16), benign hyalinizing trabecular adenomas (please refer to Chapter 12),and in some cases of adenomatous nodules and FA (please refer to Chapters 3 and 4). The NCIof PTC are larger than one third of the nuclear diameter and have distinctive sharp contour by
a rim of condensed chromatin (please refer to Chapters 3 and 4), which was rarely found in thiscase. The minimum number of NCIs necessary for a PTC diagnosis is uncertain because thethreshold varies among observers and a significant observer variation exists. For PTC, NCIs
should be observed in many tumor cells and is found easily when examining. However thosenuclear changes as shown in the Figures 3 and 4 are incomplete nuclear features for PTC typemalignancy. Therefore it may be appropriate to classify this case in either indeterminate B,
others, PTC cannot be ruled out (AUS by the Bethesda system, TIR 3A by the Italian systemor Thy 3a by the British system) or indeterminate A, follicular neoplasm (FN by the Bethesda
system, TIR 3B by the Italian system or Thy 3f by the British system). It is also acceptable but
not advisable to classify it in suspicious for PTC (TIR 4 or Thy 4), follicular variant, or pattern
B of suspicious for malignancy by the Bethesda system, because it’s clinical management isusually different from that of indeterminate categories. A conservative approach of classifying
cases with mild PTC-N, which are intermediate between the indeterminate category andsuspicious categories, into indeterminate category is recommended. Classification of NIFTP or
WDT-UMP in suspicious for malignancy category has been reported in a high proportion fromseveral academic centers (1-4), and it should be minimized because of benign nature of thislesion. One typical NCI (Figure 6) is not sufficient for a diagnosis of malignant. A diagnosis ofmalignancy, such as conventional PTC, should not be applied to this case.
Histological Diagnosis: NIFTP (non-invasive follicular thyroid neoplasm with papillary-like
nuclear features), WDT-UMP (well differentiated tumor of uncertain malignant potential), or
non-invasive encapsulated follicular variant papillary carcinoma of the thyroid
An 18 mm well-circumscribed nodule was found in the right lobe of the thyroid gland(Figure 7). Small areas of hemorrhage and cystic change were noted on the cut surface (Figure7). It was a capsulated, follicular patterned lesion with thin a fibrous capsule (Figures 8A and
8B). Neither invasive growth beyond the tumor capsule (Figure 8) nor papillary growth was
identified (Figures 8-10). Moderately increased nuclear size and an irregular nuclear contour
similar to the cytological features were well demonstrated in the histological samples (Figures8-10). An irregular nuclear contour and ground glass (powdery chromatin) nuclei were moreclearly shown in the Figures 9 and 10. Incomplete NCIs or degenerative vacuoles were seen
as indicated with blue arrows in the Figures 9 and 10. No enlarged lymph nodes suggestingmetastasis were found during surgery, and prophylactic lymph node dissection was not carried
out.

64

Figure 7. Gross appearance of the thyroid tumor (cut surface after formalin fixation), showing an
encapsulated nodule, 18 mm in diameter, ivory-white in color, and solid with hemorrhage and cystic
change.

A

B

Figure 8. Low (A, x100) and high (B, x400) magnifications of the tumor, showing the wellencapsulation by a thin fibrous capsule. Note follicular but not papillary growth pattern. These follicles
are composed of atypical follicular cells with large irregular nuclei.

6. The New Tumor Entity “NIFTP”

65Figure 9. Incomplete
nuclear inclusions or
vacuoles are indicatedby blue arrows.
Note the irregularnuclear contour andhyperchromatic nuclear
chromatin (x400).

Figure 10. Bluearrows indicate nuclear
vacuoles in irregularnuclei. Note the purefollicular growth pattern(x400).

Explanatory Notes:
PTC is a malignant follicular cell tumor characterized by distinctive nuclear features.
Follicular variant PTC (FVPTC) is the most common sub-type of PTC, which is characterizedby PTC type nuclear features (PTC-N) and a follicular (non-papillary) growth pattern. There
are debates on diagnostic criteria of this variant and observer variation between benign FA andmalignant FVPTC has been reported to be significant (5-7). The Memorial Sloan-Kettering
Cancer Center group has clarified that there are three sub-types of FVPTC that have different
genetic profiles and different biological behaviors (8-10). None of the 57 cases of non-invasive66