Table Of ContentHistochemistry
of the
Salivary Glands
Author
Masahiko Mori, D.D.S., Ph.D.
Professor and Chairman
Department of Oral and Maxillofacial Surgery
Asahi University School of Dentistry
Gifu, Japan
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PREFACE
Cytoskeletal proteins and secretory materials in salivary gland tissues have been the
subject of investigation for a long time, and are also of wide interest for broad cross relations
in biology including histochemistry. Recent topics have been highlighted to evaluate the
significance of biologically active peptides such as epidermal growth factor (EGF), which
are of salivary gland origin and have been established in the mice salivary gland. It is well
known that salivary glands show a tremendous difference in their histological features among
the different glands and in different animals. Most interestingly, the histology of these glands
is obtained through sex dimorphism in rodent submandibular glands. They are strongly
responsive to male sex hormones, and sex difference probably affects their histological
appearance. The increased focus and interest in salivary gland research is due not only to
the identification of detectable cytoskeletal and secretory proteins, but also to the importance
of growth factors in biological functions.
The objective of this book is to summarize the current topics in the immunohistochemistry
of these materials of salivary gland origin. Section I principally deals with intermediate size
proteins in the different salivary gland structures in humans and animals, and with the use
of monoclonal antibodies against keratin proteins. It is very interesting to note that salivary
glands, particularly submandibular glands, are characterized by the existence of structural
differences in animal species, but keratin distributions also strongly indicate heterogeneity.
In the human salivary glands, immunohistochemistry has come to be concerned with the
normal salivary glands and their pathological lesions. Salivary glands in humans develop
many types of benign and malignant tumors, and these topics are treated in the book from
the point of view of the diagnostic pathologist who is interested in differentiating between
the specific patterns of immunohistochemical characteristics, including coexpression of in-
termediate proteins in tissues and cell structures of ductal origin. The discussion of human
salivary gland lesions is wide and comprehensive enough to benefit not only pathologists
but also clinical researchers in identifying cellular origin as well as the specific properties
of their structural and functional components.
Sections II and III emphasize the utility of the immunohistochemical approach for
elucidating the structure and related functions of the salivary glands. These include detection
of the epithelial membrane antigen (EMA), carcinoembryonic antigen (CEA), lactoferrin,
involucrin, and S-100 protein, which are the functional indicators of salivary gland secretion,
monitoring of tumors and calcium-binding mechanisms, and cellular regulations of myoe-
pithelial derived cells of salivary gland tumors. arAntitrypsin and arantichymotrypsin,
both proteinase inhibitors, are also discussed with the defense mechanism of salivary gland
tissue for bacterial or infectious lesions.
In Section IV, the discussion covers the enzymes amylase, neuron-specific enalase,
carbonic anhydrase, and lysozyme, with immunohistochemical techniques for the detection
of normal and pathologic salivary glands. It deals with the production and biosynthesis of
these enzymes, which play major roles in the secretion of saliva from the salivary gland
cells.
Section V denotes the potential functional role of granular convoluted tubule cells in
the mice submandibular gland, such as indicating biosynthesis and secretions from the tubule
cells under normal, hormone-treated, and secretagogue-treated conditions. It emphasizes the
immunohistochemical localization of the human epidermal growth factor (hEGF) in salivary
gland tumors. Growth factors signify important and complex biological involvements for
the regulation of cells and for binding to the growth receptors.
Section VI deals with the lectin binding affinities of salivary glands in humans and
animals; lectin binding patterns are conspicuously expressed in the sugar residual site in the
tissues and cells of salivary glands. Specific patterns of lectin binding in the acini of both
serous and mucous cells, and in ductal segments including intercalated, granular convoluted,
striated, and excretory ducts were conducted in the normal gland, obstructive lesions, and
tumors.
Finally, the book is a comprehensive thesis on the current interest in detailed scientific
knowledge regarding secretion, biosynthesis of growth factors in the granular convoluted
tubules, and regulation in the salivary glands, particularly in the mice submandibular glands
and in the human salivary glands.
Masahiko Mori
ACKNOWLEDGMENTS
I have much pleasure in acknowledging the help of Dr. Harold M. Fullmer, Professor
Emeritus, University of Alabama at Birmingham, for editing the manuscript of this book
for language and syntax. I first got to know him 30 years ago when he was appointed to
the chair of Histopathology at the Dental Research Institute of the National Institutes of
Health, Bethesda, Maryland. His interest in the project and his unfailing courtesy has made
it possible for me to publish this book in its present form.
The contents of this book is the product of research done at the Department of Oral and
Maxillofacial Surgery at Asahi University School of Dentistry, where many aspects of it
first appeared as doctoral dissertations, reports, or individual manuscripts. The work would
not have come to fruition without the direct and indirect support of many faculties of Asahi
University during the past 10 years. I am particularly thankful to my colleague, Dr. Yoshiaki
Takai, whose encouragement and support I enjoyed during the making of the book, as well
as to the staff of my department who extended continuous help in editing and typing the
manuscript. I also thank my friend Noriaki Umehara of the Photo Center of the university
for his help in making the photographs used in the book. Finally, I take this opportunity to
express my appreciation of Mohan Nair, Coordinating Editor, and the production staff of
CRC Press for their care and cooperation in editing and producing this text.
THE AUTHOR
Masahiko Mori, D.D.S., Ph.D., is Professor and Chairman of the Department of Oral
and Maxillofacial Surgery, Asahi University School of Dentistry, Hozumi, Gifu, Japan.
Dr. Mori graduated with a D.D.S. degree from Osaka Dental College in 1950, and did
postdoctoral work at Osaka University Medical School (1950—1951) and Osaka University
Dental School (1951—1952). He was an instructor in the Department of Oral-Maxillofacial
Surgery at Osaka University Dental School from 1952 to 1960. Dr. Mori obtained his Ph.D.
degree in 1957 from Osaka University Medical School, and was appointed Associate Pro-
fessor in the Department of Oral-Maxillofacial Surgery, Osaka University Dental School in
1960. From 1971, he has held the position of Professor and Chairman of the Department
of Oral-Maxillofacial Surgery at Gifu College of Dentistry, known since 1985 as the Asahi
University School of Dentistry.
Dr. Mori is a fellow of Royal Society of Medicine, and a member of the Royal Micro-
scopic Society, International Association for Dental Research, Federation Dentaile Inter-
national, Japanese Cancer Association, Japanese Society for Cancer Therapy, Japanese
Society of Oral and Maxillofacial Surgeons, Japanese Stomatological Society, Japanese
Association for Oral Biology, Japan Society of Histochemistry and Cytochemistry, and Japan
Society for Cell Biology. He is an honorary fellow of the Philippines College of Oral
Surgeons, and Counsellor of Harbin Medical University, People’s Republic China.
Dr. Mori has delivered over 30 invitation lectures in international meetings, and over
100 guest lectures at universities and institutes in Japan and abroad. He has published more
than 150 papers in Japanese, and more than 200 papers in English in international journals.
He is also the coauthor of five books.
Dr. Mori’s current research interests are the immunohistochemistry of cytoskeletal pro-
teins, enzymes or enzyme-related proteins, and growth factors such as EGF and NGF, and
relate to carcinogenesis of oral lesions such as oral mucosa and salivary glands.
TABLE OF CONTENTS
Section Is Cytoskeletal Proteins
Chapter 1
Keratin...........................................................................................................................................1
I. Introduction....................................................................................................................... 1
II. Proteolytic Enzyme Pretreatments and Fixations for
Immunohistochemically Detectable Keratins.................................................................1
III. Histochemical Methods....................................................................................................4
A. Indirect Method....................................................................................................4
B. PAP Method.........................................................................................................4
C. ABC Method.........................................................................................................5
IV. Keratin Distribution of Human Salivary Glands............................................................5
V. Immunohistochemical Expression of Keratin Antibodies in Animal
Salivary Glands..................................................................................................................9
VI. Immunohistochemical Patterns for Monoclonal Antibodies toK eratins...................11
VII. Keratin Distribution of Duct-Ligated Salivary Glands...............................................25
VIII. Keratin Distribution of Obstructive Lesions in Human Salivary Glands.................27
A. Degenerating Stage.............................................................................................27
B. Intermediate Stage...............................................................................................29
C. Chronic Stage.....................................................................................................29
IX. Keratin Distribution in Mucoceles...............................................................................29
X. Keratin Expression in Experimental Tumors of Mouse Salivary Glands.................31
XI. Keratin Expression in Human Salivary Gland Tumors..............................................31
A. Pleomorphic Adenomas....................................................................................35
1. Tubulo-Ductal and Cyst-Like Structures.............................................37
2. Squamous Metaplastic Cells................................................................37
3. Modified Myoepithelial Cells...............................................................37
4. Myoepithelioma or Myoepithelial Adenomas.....................................37
B. Monomorphic Adenomas...................................................................................43
C. Warthin’s Tumors..............................................................................................46
D. Adenoid Cystic Carcinoma...............................................................................47
E. Acinic Cell Carcinomas....................................................................................54
F. Mucoepidermoid Tumors...................................................................................55
G. Squamous Cell Carcinomas..............................................................................56
H. Undifferentiated Carcinomas............................................................................59
References......................................................................................................................................62
Chapter 2
Vimentin and Desmin................................................................................................................69
I. Introduction.......................................................................................................................69
IL Immunohistochemical Method.......................................................................................69
A. Polyclonal Antibodies........................................................................................69
B. Monoclonal Antibody........................................................................................69
III. Immunohistochemical Findings.....................................................................................69
A. Animal Salivary Glands....................................................................................69
B. Human Salivary Glands....................................................................................71
C. Pleomorphic Adenoma......................................................................................71
References......................................................................................................................................77
Chapter 3
Actin and Myosin......................................................................................................................79
I. Introduction.......................................................................................................................79
IL Immunohistochemical Method.......................................................................................79
III. Immunohistochemical Findings.....................................................................................79
A. Rodent Salivary Glands.....................................................................................79
B. Normal Human Salivary Glands.......................................................................79
C. Obstructive Lesions...........................................................................................79
D. Pleomorphic Adenomas.....................................................................................80
References......................................................................................................................................85
Chapter 4
Gilial Fibrillary Acidic Proteins (GFAP)..............................................................................87
I. Introduction.......................................................................................................................87
IL Immunohistochemical Method.......................................................................................87
III. Immunohistochemical Findings......................................................................................87
References......................................................................................................................................90
Section II: Tissue and Plasma Proteins
Chapter 1
Epithelial Membrane Antigen....................................................................................................93
I. Introduction......................................................................................................................93
II. Immunohistochemical Method......................................................................................93
A. Control..................................................................................................................93
III. Immunohistochemical Findings....................................................................................93
A. Normal Salivary Glands....................................................................................93
B. Obstructive Lesions...........................................................................................93
C. Salivary Gland Tumors......................................................................................94
References......................................................................................................................................98
Chapter 2
Carcinoembryonic Antigen (CEA)........................................................................................101
I. Introduction......................................................................................................................101
IL Immunohistochemical Methods....................................................................................101
A. P-CEA................................................................................................................101
B. NCAa-CEA.......................................................................................................101
C. M-CEA................................................................................................................101
D. Control................................................................................................................102
III. Immunohistochemical Findings....................................................................................102
A. Normal Salivary Glands...................................................................................102
B. Obstructive Lesions and Mucocele................................................................102
C. Salivary Gland Tumors....................................................................................102
References....................................................................................................................................108
Chapter 3
Lactoferrin................................................................................................................................109
I. Introduction......................................................................................................................109
IL Immunohistochemical Method......................................................................................109
III. Immunohistochemical Findings....................................................................................109
A. Normal Salivary Glands....................................................................................109
B. Obstructive Lesions..........................................................................................109
C. Salivary Gland Tumors....................................................................................110
References....................................................................................................................................116
Chapter 4
Involucrin..................................................................................................................................117
I. Introduction......................................................................................................................117
IL Immunohistochemical Method.....................................................................................117
III. Immunohistochemical Findings....................................................................................117
References....................................................................................................................................119
Chapter 5
S-100 Protein.............................................................................................................................121
I. Introduction.....................................................................................................................121
II. Immunohistochemical Method.....................................................................................121
A. S-100 Protein (Polyclonal Anti-S-100 Antiserum).......................................121
B. S-100 Protein a and (3 Subunits (Monoclonal Antibodies).........................121
III. Immunohistochemical Findings....................................................................................121
A. Normal Salivary Glands...................................................................................121
B. Sialoadenitis.......................................................................................................121
C. Pleomorphic Adenomas...................................................................................122
1. Reactive and Modified Myoepithelial Cells......................................122
2. Tubulo-Ductal Structures....................................................................123
a. Spindle Tumor Cells in Outer Zone....................................123
b. Round Tumor Cells...............................................................124
3. Additional Observations.....................................................................125
References....................................................................................................................................133
Chapter 6
Laminin.......................................................................................................................................137
I. Introduction.....................................................................................................................137
II. Immunohistochemical Method.....................................................................................137
III. Immunohistochemical Findings....................................................................................137
A. Normal Salivary Glands...................................................................................137
B. Pleomorphic Adenomas...................................................................................137
References....................................................................................................................................141
Section III: Proteinase Inhibitor
Chapter 1
al-Antitrypsin (al-AT)...........................................................................................................145
I. Introduction.....................................................................................................................145
II. Immunohistochemical Method.....................................................................................145
III. Immunohistochemical Findings...................................................................................145
A. Normal Salivary Glands...................................................................................145
B. Obstructive Lesions..........................................................................................145
C. Pleomorphic Adenomas...................................................................................145
References....................................................................................................................................147
Chapter 2
al-Antichymotrypsin (al-Ach).............................................................................................149
I. Introduction......................................................................................................................149
