Original Article

Qualitative expression of hypoxia‑inducible factor‑1α in

malignant transformation of oral submucous fibrosis: An
immunohistochemical study
Treville Pereira, Ridima Surve, Subraj Shetty, Swati Gotmare
Department of Oral and Maxillofacial Pathology and Microbiology, D. Y. Patil Deemed to be University School of Dentistry,
Navi Mumbai, Maharashtra, India

Abstract Background: Oral submucous fibrosis (OSF) is a precancerous condition predominantly seen in people of Asian
descent. About 7%–12% of OSF patients develop oral squamous cell carcinoma. Morphological features of OSF,
especially fibrosis, suggest a possibility of hypoxic environment in diseased tissues. Neovascularization and
increased glycolysis, represent adaptations to a hypoxic microenvironment that are correlated with tumor
invasion and metastasis. The adaptation of cells to hypoxia appears to be mediated through hypoxia‑inducible
factor‑1α (HIF‑1α). HIF‑1α is said to be associated with malignant transformation of epithelium in other sites.
Aim: The aim of this study was to investigate the relationship between the expression of HIF‑1α in OSMF
and its role in malignant transformation.
Materials and Methods: A retrospective study which included 20 histopathologically diagnosed cases of
OSF was conducted. A qualitative evaluation of HIF‑1α was performed. Statistical analysis was carried out
using the IBM Statistical Package for Social Sciences 20.0 version (IBM Corporation, Armonk, NY, USA).
Results: Results showed an increased expression of HIF‑1α in OSF.
Conclusion: HIF‑1α appears to play a role in malignant transformation of OSF.

Keywords: Epithelial dysplasia, hypoxia, hypoxia‑inducible factor‑1α, malignant dysplasia, oral submucous
fibrosis

Address for correspondence: Dr. Treville Pereira, Department of Oral and Maxillofacial Pathology and Microbiology, D. Y. Patil Deemed to be University
School of Dentistry, Sector 7, Nerul, Navi Mumbai ‑ 400 706, Maharashtra, India.
E‑mail: trevillepereira@gmail.com
Submitted: 05-Aug-2019, Accepted: 18-Dec-2019, Published: 08-May-2020

INTRODUCTION habits of chewing tobacco, betel quid and areca nut in

the Indian subcontinent. Survival rates for OPC have not
Oral and pharyngeal cancers (OPC) are considered significantly improved over the past three decades.[1]
an important part of the global cancer burden. Oral
cancer is the sixth most common malignancy. More than The concept of stepwise development of cancer in
90% of all oral cancers are squamous cell carcinoma the oral mucosa, i.e., the initial presence of a precursor
(oral squamous cell carcinoma [OSCC]). The risk of lesion subsequently developing into cancer is well
developing OSCC is increased by the highly prevalent
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How to cite this article: Pereira T, Surve R, Shetty S, Gotmare S. Qualitative

DOI: expression of hypoxia‑inducible factor-1α in malignant transformation of
10.4103/jomfp.JOMFP_234_19 oral submucous fibrosis: An immunohistochemical study. J Oral Maxillofac
Pathol 2020;24:106-12.

106 © 2020 Journal of Oral and Maxillofacial Pathology | Published by Wolters Kluwer - Medknow
 Pereira, et al.: Role of HIF‑1α in malignant transformation of OSF

established.[2] Many OSCC has been documented to be MATERIALS AND METHODS

associated with or preceded by a precancerous lesion,
especially leukoplakia.[3] It is observed that the malignant This retrospective study was carried out in the Department
transformation potential increases with the severity of of Oral and Maxillofacial Pathology, D. Y. Patil Dental
epithelial dysplasia with a malignant transformation rate College, Navi Mumbai, India. Relevant history of each
ranging from 3% to 33%.[4] individual was obtained from the archives. Twenty blocks
of formalin‑fixed and paraffin‑embedded tissues were
Hypoxia is a common feature of many cancers and randomly selected from the archives. The five subjects
contributes to local and systematic cancer progression.[5] It for the control group were selected from age‑ and
reflects the imbalance between oxygen consumption by the sex‑matched individuals. These individuals were ruled out
rapidly proliferating cancer cells and the insufficient oxygen for the presence of any systemic illness. Further, these
delivery due to poor vascularization and blood supply and individuals did not have any oral tissue abuse habits or
is a common feature of solid tumors, including head and any obvious oral lesions. Individuals who had a history
neck squamous cell carcinoma (HNSCC).[6] of oral tissue abuse habits such as tobacco chewing,
smoking, areca nut chewing and alcohol consumption were
Oral submucous fibrosis (OSF) is a chronic, complex, included in the study. Relevant history of each individual
premalignant condition of the oral cavity, characterized was checked thoroughly to rule out any systemic illness or
by juxta‑epithelial inflammatory reaction and progressive any prior therapy. Individuals who had taken antibiotics
fibrosis of the submucosal tissues. The molecules involved in the previous month for any tooth infection, those on
in the biological pathways of the fibrotic process appear anticoagulant therapy, chemotherapy, radiation therapy
to be either down or upregulated at different stages of were excluded from the study. The study was carried out
the disease. over a period of 6 months. Written informed consent and
institutional ethical committee clearance were obtained
Hypoxia causes cell death if it is severe or prolonged. for the study.
However, cancer cells acclimatize to this hostile
environment. [7] Protection against hypoxia in solid Clinical staging of severity of fibrosis was carried out based
tumors is an important step in tumor development on the study conducted by Lai et al.[10]
and progression. One system in hypoxia protection
of tumor cells is represented by the hypoxia‑inducible The paraffin blocks of the study individuals were sectioned
factor‑1α (HIF‑1α) under hypoxic conditions, especially with a semi‑automatic microtome (Leica, Germany) into
in angiogenesis and carcinogenesis. HIF‑1α is a two tissue sections of 3 μm thickness. One section was
heterodimeric protein consisting of an alpha and beta stained with hematoxylin and eosin (H & E).
subunit, in which the HIF‑1α subunit mediates HIF‑1α
function as a transcription factor. [8] In human solid Further, the H & E stained sections were histopathological
tumors, where hypoxia is found, it serves as a selective graded using the criteria given by Utsunomiya et al.,[11]
environment for survival of aggressive cancer cells which is based on the concept of the Pindborg and Sirsat
and as protection from anticancer therapies. [9] The grading system.
expression of HIF‑1α has been demonstrated in a
variety of cancers, including renal, bladder, colorectal, Immunohistochemistry
breast, ovarian, endometrial, cervical and head and neck The other 3 µm section of the tissue was cut and
carcinomas.[5] Morphological features of OSF, especially transferred to APES (Sigma‑Aldrich Chemical Co., USA)
fibrosis, suggest a possibility of the hypoxic environment coated slides and incubated overnight at room temperature.
in diseased tissues. The adaptation of cells to hypoxia After warming in a slide warmer for 15 min, the sections
appears to be mediated through HIF‑1α. were deparaffinized in three changes of fresh xylene each
for 5 min followed by dehydration in a series of absolute
The purpose of this study was to investigate the expression alcohol each for 5 min. Endogenous peroxidases were
on HIF1‑α in OSF through its effect on the staining blocked with peroxide block (BioGenex Life sciences Pvt.,
intensity by analyzing the mean blood vessel and mean Ltd, CA, USA) for 15 min at room temperature and washed
fibroblast density. The study also aimed at finding out a in distilled water followed by citrate buffer (pH 6.0) wash
correlation between the staging of the disease, the grading for 10 min. Antigen retrieval was undertaken with a help
of epithelial dysplasia and the staining intensity based on of BioGenex antigen retrieval system. The sections were
epithelial dysplasia, if any. immersed in citrate buffer solution and placed into the
Journal of Oral and Maxillofacial Pathology | Volume 24 | Issue 1 | January-April 2020 107
 Pereira, et al.: Role of HIF‑1α in malignant transformation of OSF

BioGenex antigen retrieval system and heated for 15 min. mild (+), and cases in between the two extremes were
The system was allowed to cool to room temperature by graded into the moderate (++) category.
placing it under running tap water, and later, the slides
were washed with distilled water for 5 min. With an Statistical analysis
intention to block endogenous biotin, the sections were Descriptive and inferential statistical analyses were
incubated with a blocking agent (BioGenex Life sciences carried out in the present study. Results on continuous
Pvt., Ltd., CA, USA) for 15 min. Excess power block measurements were presented on mean ± standard
solution was drained, and the sections were incubated with deviation, and results on categorical measurement were
primary monoclonal antibody of HIF‑1α (BioGenex Life presented in number (%). The level of significance was
Sciences Pvt., Ltd, CA, USA) for 1 h and later thoroughly fixed at P = 0.05 and any value ≤0.05 was considered to
washed with citrate buffer. For further enhancement be statistically significant.
of the staining, the sections were then incubated with
the anti‑mouse secondary antibody (super‑enhancer) Analysis of variance (ANOVA) was used to find the
significance of study parameters between the groups
for 30 min followed by two consecutive buffer washes;
(intergroup analysis). Further post hoc analysis was carried
each for 5 min. Horseradish peroxidase (BioGenex Life
out if the values of ANOVA test were significant.
Sciences Pvt., Ltd., CA, USA) was added to the sections and
incubated for 30 min. The chromogen diaminobenzidine
The Statistical software IBM Statistical Package for Social
was prepared just prior to use by mixing one drop of Sciences statistics 20.0 (IBM Corporation, Armonk,
chromogen to 1 ml of the buffer in a mixing vial and later NY, USA) was used for the analyses of data.
added over the sections. After 5 min, the sections were
washed in buffer followed by water and counterstained OBSERVATIONS AND RESULTS
with Harris hematoxylin, air dried, cleared and mounted
with dibutyl phthalate xylene. The study comprised of 20 OSF (study group) and
five healthy controls (control). The study population
Assessment of the hypoxia‑inducible factor‑1α staining comprised of 16 males and four females with a mean
intensity age of 29 years, and the control group comprised of
Cells positive for HIF‑1α were regarded as fibroblasts and three males and two females. The study group as well
blood vessels. Fibroblasts and blood vessels were counted as control group showed a male predominance. Clinical
using a Leica research microscope with provision for staging of the patients was made depending upon the
photomicrograph (Model no. DM1000 LED, Germany). mouth opening as suggested by Lai et al. There were
The stained sections were first screened at low power seven cases in Stage 1, five cases in Stage 2, seven cases
(×10) to determine the areas of most intense staining in Stage 3 and one case in Stage 4. We observed that the
(Hot spot method). Fibroblasts and blood vessel counting majority of the study group falls in Stage 1 and Stage 3.
was then performed under high power (×40) magnification. We further did a comparison of staining intensity with
Five high‑power fields (HPFs) with the highest number grades of epithelial dysplasia (P = 0.199), comparison of
of these cells were chosen. The representative areas epithelial dysplasia with clinical staging (P = 0.283) and a
were carefully scanned from left to right of every slide comparison of epithelial dysplasia with histopathological
to avoid recounting of same areas. The cells for each grading (P = 0.283). All the above values were statistically
case was the average of the number in these five chosen insignificant.
HPFs and expressed as the number of fibroblasts or
blood vessels per HPF. The mean of 10 values was The collected data were then statically analyzed to find
calculated and expressed as mean (standard deviation). All a significance of the blood vessels in correlation to the
immunohistochemistry (IHC)‑stained slides along with the different grades of staining intensity. Among the 20 cases
corresponding H & E slides were evaluated by two qualified with respect to staining intensity, nine cases showed
observers to minimize the subjective bias. mild staining intensity with a mean of 7.13 (4.267), six
cases showed moderate staining intensity with a mean
The section was considered either as negative or positive of 6.50 (3.779) and five cases showed a severe staining
according to the absence or presence of brown staining in intensity with a mean of 7.49 (6.316). The P = 0.594
epithelial or stromal cells. The positive cases were graded [Table 1 and Figure 1]. When the data were analyzed to
into three categories depending on the intensity of staining. find the significance of the fibroblasts in co‑relation to
Homogeneous dark brown staining was considered as the different grades of staining intensity it was observed
strong (+++) and light faint staining was considered as that among the 20 cases with respect to staining intensity,
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 Pereira, et al.: Role of HIF‑1α in malignant transformation of OSF

Figure 1: Comparison of blood vessels in terms of (mean [standard Figure 2: Comparison of fibroblast in terms of (mean [standard
deviation]) among different grades of dysplasia using analysis of deviation]) among different grades of dysplasia using analysis of
variance test variance test

nine cases showed mild staining intensity with a mean Table 1: Comparison of blood vessels in terms of (mean±standard
of 13.26 (8.792), six cases shows moderate staining deviation) among different grades of staining intensity using
analysis of variance test
intensity with a means of 45.20 (25.652) and five cases
Staining intensity n Mean±SD F P
showed a severe staining intensity with a mean of Mild 9 7.13±4.267 0.536 0.594
70.16 (13.723). The P < 0.001 which was highly significant Moderate 6 6.50±3.779
[Table 2 and Figure 2]. Severe 5 9.32±6.316
Total 20 7.49±4.590
DISCUSSION SD: Standard deviation

HNSCC is the sixth most common malignancy worldwide. Table 2: Comparison of fibroblast in terms of (mean±standard
deviation) among different grades of staining intensity using
OSCC remains a significant health burden across the analysis of variance test
globe.[12] It usually arises from a preexisting malignant Staining intensity n Mean±SD F P
disorder, and occasionally de novo; but in either case from Mild 9 13.26±8.792 20.002 <0.001**
within a field of precancerized epithelium. Important risk Moderate 6 45.20±25.652
factors related to the carcinoma itself that are associated Severe 5 70.16±13.723
Total 20 37.07±28.684
with a poor prognosis include large size of the tumor at the **P<0.001: Highly significant. SD: Standard deviation
time of diagnosis, the presence of metastases in regional
lymph nodes, and a deep invasive front of the tumor.[13]
disease is a genetic process that later becomes evident at
Despite advances in therapeutic approaches, percentages
the cellular level and ultimately at the clinical level. The
of morbidity of OSCC and mortality have not improved
use of molecular markers allows diagnosis and staging of
significantly during the past 30 years.[14] Oral cancers arise
tissue change before changes in cell morphology occurs
from sustained, stepwise accumulations of mutations
and certainly before tissue changes become clinically visible.
resulting in the transition of normal mucosa to dysplasia
to invasive carcinoma over time.[15] Ultimately, the use of molecular markers in diagnosis may
lead to survival and less treatment‑associated morbidity
Furthermore, many OSCCs are believed to develop through early recognition of and intervention for at‑risk
from oral premalignant disorders (OPMD) and early oral lesions.[16]
detection and diagnosis of these premalignant lesions
should be possible. Identification of high‑risk OPMD Hypoxia is a common feature and contributes to local and
and intervention at premalignant stages could constitute systemic cancer progression, resistance to therapy and poor
one of the keys to reducing mortality, morbidity and outcome.[17] Oxygen concentrations are markedly reduced
cost of treatment associated with SCC. There are several in many human cancers compared with normal tissue,
OPMD that precede the development of OSCC. The most and major mechanism mediating adaptive responses to
commonly encountered are erythroplakia, leukoplakia and reduced oxygen availability (hypoxia) is the regulation of
OSF.[12] Several potential markers of molecular changes in transcription by HIF‑1α.[18] Hypoxia is a state of reduced
oral premalignant lesions and conditions have been studied, oxygen availability or decreased oxygen partial pressures
and interest in the genetic changes associated with these below critical thresholds, and it restricts or even abolishes
lesions is increasing. Progression from benign to malignant the function of organs, tissues or cells. In solid tumors,
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 Pereira, et al.: Role of HIF‑1α in malignant transformation of OSF

oxygen delivery to the respiring neoplastic and stromal cells and ulceration of the epithelium by inducing apoptosis. In
is frequently reduced or even abolished by deteriorating addition, the overexpression of HIF‑1α is seen in OSF,
diffusion geometry, severe structural abnormalities of which indicates changes in cell proliferation, maturation
tumor microvessels and disturbed microcirculation. and metabolic adaptation, increasing the possibility of
Neovascularization and increased glycolysis represent malignant transformation. [6] On degradation, HIF‑1
adaptations to a hypoxic microenvironment that are becomes inactive and the cells lacking oxygen supply are
correlated with tumor invasion and metastasis.[19] unable to survive and undergo apoptosis. However, under
hypoxic conditions as in OSF, HIF‑1α is stabilized and on
The development of hypoxic microenvironment is caused stabilization, it translocates to the nucleus where it forms
by the imbalance between oxygen consumption and oxygen the functional transcription factor HIF‑1. The functional
delivery. The rapidly proliferating HNSCC has insufficient HIF‑1α helps in cell survival under hypoxia and thus helps
vascularization with poor blood supply. The hypoxic stress cell proliferation promoting tumorigenesis.[25] HIF‑1α is
stimulates solid tumors to upregulate the expression of a rarely expressed in the normal oral mucosa. However, a
variety of oncogenes such as HIF and endothelial growth significant HIF‑1α expression was found in OSF in the
factor which enhances irregular vascular endothelial cell basal and suprabasal layers of epithelium. This indicates
proliferation and differentiation. The adaptation of cells the role of hypoxia in malignant transformation of OSF.
to hypoxia appears to be mediated through HIF‑1α which Thus, the upregulation of HIF‑1α is an early event in
is said to be associated with malignant transformation of carcinogenesis.[26]
epithelium in other sides. It appears that HIF‑1α plays a
significant role in both prostate and cervical carcinogenesis Dunkel et al.[27] found that the CD44 low HIF‑1α high
at early stages.[20] signature was associated with poorer disease‑free survival.
Uehara et al.[7] found expression of HIF‑1α in OSCC is
Little is known about the role of HIF‑1α in early events likely to be of great value in treatment planning and to
of carcinogenesis in the oral cavity. Therefore, the aim of predict the prognosis of OSCC. Lee et al.[28] found that
our study was to evaluate the expression of HIF‑1α in HIF‑1a expression is significantly upregulated in areca
OSF by IHC. quid chewing‑associated OSCC. Zheng et al.[29] in their
reverse transcription‑polymerase chain reaction study
Hypoxia is a common environmental stress factor measured mRNA levels of HIF‑1α and found HIF‑1α
and is associated with physiological and pathological mRNA levels were significantly increased in carcinoma of
conditions related to cancer invasion and metastasis.[21] the tongue, and a positive correlation was observed with
The process of tumor cell invasion involves degradation pathological differentiation grade. On the contrary, dos
of extracellular matrix and matrix metalloproteinases Santos et al.[9] showed a significant relationship between
which play an important role in invasion in OSCC.[22] The strong HIF‑1α protein expression and lower local disease
higher incidence of metastasis could also be explained relapse (P = 0.002) and increased local disease‑free survival
by the fact that the glycolytic products lactase and acid (P = 0.013). In a recent study on prostate carcinogenesis, it
induce secretion of matrix‑degrading hyaluronidase and was reported that the upregulation of HIF‑1α is an early
metalloproteinase by tumor‑associated fibroblasts create event.[30] They showed a gradual increase in the expression
a tumor microenvironment favorable for tumor cell of HIF‑1α from benign prostatic hyperplasia, prostatic
migration.[23] intraepithelial hyperplasia to prostatic cancer compared
with normal prostatic epithelium.
OSF is histopathologically characterized by fibrosis of
subepithelial connective tissue. Collagens are the major In the present study conducted, we hypothesized a
structural component of extracellular matrix; hence, precise possible role of HIF‑1α in progression and malignant
regulation of collagen metabolism is essential to maintain transformation of OSF and to investigate the role
the normal integrity of connective tissue.[24] of hypoxia in OSF and its relationship to epithelial
dysplasia. We have demonstrated the evidence of marked
With the progression of the disease process of OSF, upregulation of HIF‑1α in a reasonably large number
the production of collagen type 1 is increased, and the of OSF samples by conducting IHC and then by using
degradation of collagen is reduced by up to 75%. Extensive the hot‑spot method, the counting of the stained blood
fibrosis of the connective tissue causes a reduction vessels and fibroblast was done. Although this finding was
of vascularity, resulting in subsequent hypoxia in both not consistent as in few cases, staining was restricted to
fibroblasts and surface epithelium. Hypoxia causes atrophy the dysplastic zone only. In the past literature, increased
110 Journal of Oral and Maxillofacial Pathology | Volume 24 | Issue 1 | January-April 2020
 Pereira, et al.: Role of HIF‑1α in malignant transformation of OSF

expression of HIF‑1α was accompanied by an increase in 4. Scully C, Bagan JV, Hopper C, Epstein JB. Oral cancer: Current and
future diagnostic techniques. Am J Dent 2008;21:199‑209.
the number of blood vessels in OSF cases. However, in the 5. Lin PY, Yu CH, Wang JT, Chen HH, Cheng SJ, Kuo MY, et al. Expression
present study, it was seen that the number of fibroblasts was of hypoxia‑inducible factor‑1 alpha is significantly associated with the
more as compared to the number of blood vessels which progression and prognosis of oral squamous cell carcinomas in Taiwan.
J Oral Pathol Med 2008;37:18‑25.
supported our hypothesis that the process of OSF is due
6. Li JZ, Gao W, Chan JY, Ho WK, Wong TS. Hypoxia in head and neck
to increased fibrosis which is initiated by the fibroblast and squamous cell carcinoma. ISRN Otolaryngol 2012;2012:8.
thus there is increased amount of hypoxia leading to an 7. Uehara M, Sano K, Ikeda H, Nonaka M, Asahina I. Hypoxia‑inducible
increased expression of HIF‑1α. It has also been reported factor 1 alpha in oral squamous cell carcinoma and its relation to
prognosis. Oral Oncol 2009;45:241‑6.
that some of the molecules which have a direct relationship 8. Fillies T, Werkmeister R, van Diest PJ, Brandt B, Joos U, Buerger H.
to carcinogenesis exhibit a correlation with HIF‑1α levels. HIF1‑alpha overexpression indicates a good prognosis in early stage
One of the most important molecules in this regard is squamous cell carcinomas of the oral floor. BMC Cancer 2005;5:84.
9. dos Santos M, Mercante AM, Louro ID, Gonçalves AJ, de Carvalho MB,
vascular endothelial growth factor which is responsible for
da Silva EH, et al. HIF1‑alpha expression predicts survival of
angiogenesis, a key event in carcinogenesis.[31‑33] patients with squamous cell carcinoma of the oral cavity. PLoS One
2012;7:e45228.
CONCLUSION 10. Lai DR, Chen HR, Lin LM, Huang YL, Tsai CC. Clinical evaluation
of different treatment methods for oral submucous fibrosis. A 10‑year
Hypoxia is a common characteristic of locally advanced experience with 150 cases. J Oral Pathol Med 1995;24:402‑6.
11. Utsunomiya H, Tilakaratne WM, Oshiro K, Maruyama S, Suzuki M,
solid tumors that has been associated with diminished Ida‑Yonemochi H, et al. Extracellular matrix remodeling in oral submucous
therapeutic response and more recently with malignant fibrosis: Its stage‑specific modes revealed by immunohistochemistry and
progression, that is, an increasing probability of recurrence, in situ hybridization. J Oral Pathol Med 2005;34:498‑507.
12. Dionne KR, Warnakulasuriya S, Zain RB, Cheong SC. Potentially
locoregional spread and distant metastasis. Emerging malignant disorders of the oral cavity: Current practice and future
evidence indicates that the effect of hypoxia on malignant directions in the clinic and laboratory. Int J Cancer 2015;136:503‑15.
progression is mediated by a series of hypoxia‑induced 13. Feller L, Lemmer J. Oral squamous cell carcinoma: Epidemiology, clinical
proteomic and genomic changes activating angiogenesis, presentation and Treatment. J Cancer Therapy 2012;3:263‑8.
14. Califano J, van der Riet P, Westra W, Nawroz H, Clayman G, Piantadosi S,
anaerobic metabolism and other processes that enable et al. Genetic progression model for head and neck cancer: Implications
tumor cells to survive or escape their oxygen‑deficient for field cancerization. Cancer Res 1996;56:2488‑92.
environment. Thus, hypoxia should be considered an 15. Gillenwater A, Papadimitrakopoulou V, Richards‑Kortum R. Oral
premalignancy: New methods of detection and treatment. Curr Oncol
independent adverse prognostic factor in patients with Rep 2006;8:146‑54.
head and neck cancers.[34] Since this is a preliminary study, 16. Epstein JB, Zhang L, Rosin M. Advances in the diagnosis of oral
a larger sample size is a must. In addition, there should premalignant and malignant lesions. J Can Dent Assoc 2002;68:617‑21.
be approximately equal number of samples in each 17. de Lima PO, Jorge CC, Oliveira DT, Pereira MC. Hypoxic condition
and prognosis in oral squamous cell carcinoma. Anticancer Res
histopathological group. In conclusion, our data indicate 2014;34:605‑12.
that HIF‑1α appears to play a role in the malignant 18. Semenza GL. Defining the role of hypoxia‑inducible factor 1 in cancer
transformation of OSF. Further, over‑expression of biology and therapeutics. Oncogene 2010;29:625‑34.
19. Höckel M, Vaupel P. Tumor hypoxia: Definitions and current clinical,
HIF‑1α may at least be partly responsible for the biologic, and molecular aspects. J Natl Cancer Inst 2001;93:266‑76.
progression of fibrosis. Finally, our data suggest that it 20. Chaudhary M, Bajaj S, Bohra S, Swastika N, Hande A. The domino
may be possible to use HIF‑1α as a marker for malignant effect: Role of hypoxia in malignant transformation of oral submucous
transformation of OSF. fibrosis. J Oral Maxillofac Pathol 2015;19:122‑7.
21. Miyazaki Y, Hara A, Kato K, Oyama T, Yamada Y, Mori H, et al. The
effect of hypoxic microenvironment on matrix metalloproteinase
Financial support and sponsorship expression in xenografts of human oral squamous cell carcinoma. Int
Nil. J Oncol 2008;32:145‑51.
22. Warnakulasuriya S, Johnson NW, van der Waal I. Nomenclature and
Conflicts of interest classification of potentially malignant disorders of the oral mucosa.
J Oral Pathol Med 2007;36:575‑80.
There are no conflicts of interest. 23. Meijer TW, Kaanders JH, Span PN, Bussink J. Targeting hypoxia, HIF‑1,
and tumor glucose metabolism to improve radiotherapy efficacy. Clin
REFERENCES Cancer Res 2012;18:5585‑94.
24. Ekanayaka RP, Tilakaratne WM. Oral submucous fibrosis: Review on
1. Saman DM. A review of the epidemiology of oral and pharyngeal mechanisms of malignant transformation. Oral Surg Oral Med Oral
carcinoma: Update. Head Neck Oncol 2012;4:1. Pathol Oral Radiol 2016;122:192‑9.
2. Sadiq H, Gupta P, Singh N, Thakar SS, Prabhakar I, Thakral J. Various 25. Warburg O. On the origin of cancer cells. Science 1956;123:309‑14.
grading systems of the oral epithelial dysplasia: A review. Int J Adv 26. Priyadharshni B. Classification system for oral submucous grading‑a
Health Sci 2015;1:20‑6. review. Int J Sci Res 2014;3:740‑44.
3. Speight PM, Farthing PM, Bouquot JE. The pathology of oral cancer 27. Dunkel J, Vaittinen S, Koivunen P, Laranne J, Mäkinen MJ, Tommola S,
and precancer. Curr Diagn Pathol 1996;3:165‑76. et al. Tumoral expression of CD44 and HIF1α predict stage I oral cavity

Journal of Oral and Maxillofacial Pathology | Volume 24 | Issue 1 | January-April 2020 111
 Pereira, et al.: Role of HIF‑1α in malignant transformation of OSF

squamous cell carcinoma outcome. Laryngoscope Investig Otolaryngol carcinomas. J Surg Oncol 2006;94:242‑7.
2016;1:6‑12. 32. Lekas A, Lazaris AC, Deliveliotis C, Chrisofos M, Zoubouli C, Lapas D,
28. Lee SS, Tsai CH, Yang SF, Ho YC, Chang YC. Hypoxia inducible et al. The expression of hypoxia‑inducible factor‑1alpha (HIF‑1alpha)
factor‑1α expression in areca quid chewing‑associated oral squamous and angiogenesis markers in hyperplastic and malignant prostate tissue.
cell carcinomas. Oral Dis 2010;16:696‑701. Anticancer Res 2006;26:2989‑93.
29. Zheng Y, Ni Y, Huang X, Wang Z, Han W. Overexpression of HIF‑1α 33. Jiang CQ, Liu ZS, Qian Q, He YM, Yuan YF, Ai ZL. Relationship
indicates a poor prognosis in tongue carcinoma and may be associated of hypoxia‑inducible factor 1 alpha (HIF‑1alpha) gene expression
with tumour metastasis. Oncol Lett 2013;5:1285‑9. with vascular endothelial growth factor (VEGF) and microvessel
30. Zhong H, Semenza GL, Simons JW, De Marzo AM. Up‑regulation density (MVD) in human colorectal adenoma and adenocarcinoma. Ai
of hypoxia‑inducible factor 1alpha is an early event in prostate Zheng 2003;22:1170‑4.
carcinogenesis. Cancer Detect Prev 2004;28:88‑93. 34. Zhu GQ, Tang YL, Li L, Zheng M, Jiang J, Li XY, et al. Hypoxia inducible
31. Giatromanolaki A, Sivridis E, Simopoulos C, Polychronidis A, factor 1α and hypoxia inducible factor 2α play distinct and functionally
Gatter KC, Harris AL, et al. Hypoxia inducible factors 1alpha and 2alpha overlapping roles in oral squamous cell carcinoma. Clin Cancer Res
are associated with VEGF expression and angiogenesis in gallbladder 2010;16:4732‑41.

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