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Table of Contents   
ORIGINAL RESEARCH  
Year : 2011  |  Volume : 22  |  Issue : 2  |  Page : 256-259
Expression of heat shock protein70 in oral submucous fibrosis and oral squamous cell carcinoma: An immunohistochemical study


1 Department of Oral and Maxillofacial Pathology, Meenakshi Ammal Dental College, Porur, Chennai, India
2 Department of Oral and Maxillofacial Pathology, Sri Ramachandra University, Porur, Chennai, India
3 Department of Community Medicine, Sri Ramachandra University, Porur, Chennai, India

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Date of Submission24-Aug-2010
Date of Decision29-Nov-2010
Date of Acceptance18-Apr-2011
Date of Web Publication27-Aug-2011
 

   Abstract 

Background: Heat shock proteins are a highly conserved group of protective cellular proteins whose synthesis is increased in response to a variety of environmental or pathophysiological stresses. Heat shock proteins are useful biomarkers for carcinogenesis in tissues and signal the degree of differentiation and the aggressiveness of cancers. Regulation of heat shock protein 70 (HSP70) expression in oral submucous fibrosis is not known much, and the aim of this study was to evaluate HSP70 expression in oral submucous fibrosis and oral squamous cell carcinoma by immunohistochemical method and to understand the role of HSP70 in tumorigenesis.
Materials and Methods: Immunohistochemical method was used to detect HSP70 expression in normal oral mucosa, oral submucous fibrosis (n=30) and oral squamous cell carcinoma (n=20). HSP70 immunoreactivity was correlated with histological and clinicopathological features.
Results: A significant increase in expression of HSP70 was observed (P<0.000) as the tissue progressed from oral submucous fibrosis towards oral squamous cell carcinoma.
Conclusion: HSP70 is synthesized upon stress situations arising in cells of all living organisms. Expression of HSP70 indicates that stress plays an important role as a predisposing factor in oral submucous fibrosis and its subsequent progression to oral squamous cell carcinoma.

Keywords: Heat shock protein70, immunohistochemistry, oral squamous cell carcinoma, oral submucous fibrosis

How to cite this article:
Thubashini M, Malathi N, Kannan L. Expression of heat shock protein70 in oral submucous fibrosis and oral squamous cell carcinoma: An immunohistochemical study. Indian J Dent Res 2011;22:256-9

How to cite this URL:
Thubashini M, Malathi N, Kannan L. Expression of heat shock protein70 in oral submucous fibrosis and oral squamous cell carcinoma: An immunohistochemical study. Indian J Dent Res [serial online] 2011 [cited 2019 Oct 14];22:256-9. Available from: http://www.ijdr.in/text.asp?2011/22/2/256/84299
The primary goal of all living cells is to survive. The ability of organisms to confront and survive adverse changes in their environmental circumstance represents an integral aspect of evolution. [1] Cells from all organisms appear to employ a rather common and ubiquitous defensive mechanism when confronted with abrupt changes in their local environmental circumstance. This response is referred to as the heat shock response or stress response and entails the rapid and coordinated increased expression of a group of proteins referred to as the heat shock proteins or stress proteins.

Heat shock proteins are a highly conserved group of protective cellular proteins whose synthesis is increased in response to a variety of environmental or pathophysiological stresses, including anoxia, ischemia, heavy metal ions, ethanol, nicotine, surgical stress, viral agents, starvation, inflammation, water deprivation and nitrogen deficiency. These proteins play an important role in the maintenance of cellular homeostasis, both under normal conditions and during stress. [1],[2] Heat shock proteins are overexpressed in a wide range of human cancers and are implicated in tumor cell proliferation, differentiation, invasion, metastasis, death and recognition by the immune system. [3] They are useful biomarkers for carcinogenesis in tissues and signal the degree of differentiation and the aggressiveness of cancers. [3]

Heat shock proteins originally were discovered and reported by Ferrucio Ritossa in 1962. [1],[4] He observed that heat shock produced chromosomal puffs in the salivary glands of fruit flies (Drosophilia melanogaster). [4]

Heat shock proteins are broadly classified into 5 major families according to the apparent molecular weight, amino acid sequence homologies and their functional aspects:

  • HSP100 family (100-104 KDa),
  • HSP90 family (82-90 KDa),
  • HSP70 family (68-75 KDa),
  • HSP60 family (58-65 KDa) and
  • the small HSP family (15-30 KDa). [1]


Many of the stress proteins are present continuously - constitutive expression. Expression of other proteins is increased by stress - stress inducible. [2]

Currently there is limited information on the regulation of heat shock protein70 expression in oral submucous fibrosis, and this study was aimed at understanding the up-regulation of heat shock protein70 in oral submucous fibrosis and comparing its expression with that in oral squamous cell carcinoma by immunohistochemical method.


   Materials and Methods Top


Collection of samples

The study group consisted of 50 formalin-fixed, paraffin-embedded tissues, of which 30 were oral submucous fibrosis and 20 were oral squamous cell carcinoma retrieved from the Department of Oral and Maxillofacial Pathology, Faculty of Dental Sciences, Sri Ramachandra University, Chennai. Data on relevant features like age; sex; site; and habits, including the duration and types of habits, were obtained from the records of the patients. Breast carcinoma tissue was used as positive control [Figure 1]. Normal gingival tissue was taken as negative control [Figure 2].
Figure 1: Weak expression of HSP70 in oral submucous fibrosis (OSMF)

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Figure 2: Intense expression of HSP70 in OSMF (×10)

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Immunohistochemical kit (super-sensitive polymer horse radish peroxidase (HRP) detection system) was obtained from Biogenex Laboratories along with primary antibody (anti-human heat shock protein70).

Analysis of heat shock protein70 immunoreactivity

The highly stress-inducible heat shock protein70 is present within the cytosol/ nucleus/ nucleolus and expressed at high levels in the cells experiencing stress. The immunostained slides were evaluated semi-quantitatively and graded as weak expression or intense expression.


   Results Top


Age of patients ranged from 5 to 72 years with a mean age of 38.28 years and standard deviation of 11.6073. Increased expression of heat shock protein70 was seen in patients aged 25 to 50 years in cases of oral submucous fibrosis; and more than 50 years, in cases of oral squamous cell carcinoma. Chi-square values were found to be 9.97 and 8.25, respectively. P values were 0.007 and 0.016, respectively which were statistically significant [Table 1].
Table 1: Heat shock protein 70 expression with study group


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Weak expression of heat shock protein70 was seen in 66.6% of oral submucous fibrosis patients and 5% of oral squamous cell carcinoma patients. Intense expression of heat shock protein70 was seen in 33.3% of oral submucous fibrosis patients and 95% of oral squamous cell carcinoma patients. Chi square test (test of significance) provided a value of 18.733; and P value was <0.000, which was statistically significant [Figure 3]. There was no statistical correlation between heat shock protein 70 expression and gender, habits and duration of habits was insignificant.
Figure 3: HSP70 expression with study groups

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   Discussion Top


The present times are of importance for the subject of the role of heat shock protein70 in cancer, the assessment of which is greatly contributory to both basic and clinical cancer research. Protein misfolding is a major cause of a number of human diseases. By maintaining cellular proteins in a folding-competent state, heat shock proteins70 (HSPs70) in coordination with other chaperones play an important role in cellular viability. [5] They also assist in the repair of denatured proteins or promote their degradation after stress or injury. [5]

Our results showed increased heat shock protein70 expression between 25 and 50 years of age in patients with oral submucous fibrosis; and with increasing age, in patients with oral squamous cell carcinoma. This difference in intensity of expression was found to be statistically significant (P<0.007) [Table 1]. The difference in heat shock protein70 expression between genders in the study groups was statistically insignificant. Our results were generally in agreement with previous reports. According to Shinn lee et al., there was no significant difference in heat shock protein70 expression with respect to gender in oral squamous cell carcinoma cases. [6] Kaur et al. observed that there was no association between heat shock protein70 expression and gender of the patients with premalignant lesions. [7] They showed significant correlation of heat shock protein70 expression with premalignant condition and oral squamous cell carcinoma. [6],[7]

Expression of heat shock protein70 wuth the study group revealed weak staining [Figure 4] in 66.6% of the oral submucous fibrosis cases and intense staining [Figure 5] in 95% of the oral squamous cell carcinoma cases. The Pearson chi-square test provided a value of 18.733; and P value was <0.000, which was statistically significant [Figure 3]. As the tissue progressed from oral submucous fibrosis towards oral squamous cell carcinoma, a significant increase in expression of heat shock protein70 was observed (P<0.000). Out of 30 samples of oral submucous fibrosis, weak heat shock protein70 expression was observed in 20 cases [Figure 4]; and intense heat shock protein70 expression [Figure 6] and [Figure 7], in 10 cases. This intense heat shock protein70 expression was observed in advanced cases of oral submucous fibrosis (Pindborg histological criteria). Apart from the intensity of staining, a characteristic pattern of staining was observed. Mild oral submucous fibrosis showed weak heat shock protein70 expression extending from basal to lower half of the epithelium [Figure 4]. In the remaining 10 cases of advanced oral submucous fibrosis, heat shock protein70 expression was seen in the entire thickness of the epithelium [Figure 6] and [Figure 7]. According to the study by Shinn lee et al., [6] it can be substantiated that heat shock protein70 is up-regulated by oxidative stress and a variety of heavy metals, such as copper. Earlier studies have demonstrated that areca nut has a high copper content and generates reactive oxygen species during chewing; [8] it is therefore feasible to suggest that high levels of copper released and reactive oxygen species generated may at least in part be responsible for the up-regulation of heat shock protein70 in areca quid-associated oral submucous fibrosis and oral squamous cell carcinoma.
Figure 4: Intense expression of HSP70 in OSMF (×40)

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Figure 5: HSP70 expression in oral squamous cell carcinoma

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Figure 6: Positive control - breast carcinoma

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Figure 7: Negative control - normal gingival tissue

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Kaur et al. [9] showed strong nuclear and cytoplasmic heat shock protein70 expression in oral squamous cell carcinoma, which was similar to our study suggesting that heat shock protein70 is expressed differentially during carcinogenesis and may be implicated in tumor development.


   Conclusion Top


All these results suggest that increased heat shock protein70 expression is critical for tumor growth and might be important as a prognostic factor for patients with oral cancer. Heat shock protein70 expression was seen mostly in cells under stress rather than normal cells and it took place in the cytoplasm and nuclei of tumor cells but not in the non-neoplastic cells.

Heat shock protein70 is synthesized upon stress situations arising in cells of all living organisms. The fact that tumor, but not normal, cells express heat shock protein70 suggests tumor cells are under stress. Thus stress plays an important role as a predisposing factor in oral submucous fibrosis and also in its subsequent progression to oral squamous cell carcinoma.


   Acknowledgment Top


My heartfelt thanks to Yuvarani, Dr. Deepika, Dr. Priyadarshini for their help and support.

 
   References Top

1.Mukhopadhyay I, Nazir A, Saxena DK, Chowdhuri DK. Heat shock response: HSP70 in environmental monitoring. J Biochem Mol Toxicol 2003;17:249-54.  Back to cited text no. 1
[PUBMED]  [FULLTEXT]  
2.Whitley D, Goldberg SP, Jordan WD. Heat shock proteins: A review of the molecular chaperones. J Vasc Surg 1993;29:748-51.  Back to cited text no. 2
    
3.Ciocca DR, Calderwood SK. Heat shock proteins in cancer: diagnostic, prognostic, predictive, and treatment implications. Cell Stress Chaperones 2005;10:86-103.  Back to cited text no. 3
[PUBMED]  [FULLTEXT]  
4.Ritossa F. A new puffing pattern induced by heat shock and DNP in Drosophilia. Experientia 1962;18:571-3.  Back to cited text no. 4
    
5.Sharma D, Maison DC. HSP 70 structure, function, regulation and influence on yeast prions. Protein Pept Lett 2009;16:571-81.  Back to cited text no. 5
    
6.Lee SS, Tsai CH, Ho YC, Chang YC. The upregulation of Heat Shock Protein70 expression in areca quid chewing - associated oral squamous cell carcinomas. Oral oncol 2008;44:884-90.   Back to cited text no. 6
[PUBMED]  [FULLTEXT]  
7.Kaur J, Srivastava A, Ralhan R. Expression of 70 kDa heat shock protein in oral lesions: marker of biological stress or pathogenicity. Oral oncol 1998;34:496-501.  Back to cited text no. 7
[PUBMED]  [FULLTEXT]  
8.Tilakaratne WM, Klinikowski MF, Saku T, PetersTJ,Warnakulasuriya S. Oral submucous fibrosis:review on aetiology and pathogenesis. Oral Oncol 2006;42:561-8.   Back to cited text no. 8
    
9.Kaur J, Ralhan R. Differential expression of 70kda heat shock protein in human oral tumorigenesis. Int J Cancer 1995;63:774-9.  Back to cited text no. 9
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Correspondence Address:
M Thubashini
Department of Oral and Maxillofacial Pathology, Meenakshi Ammal Dental College, Porur, Chennai
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0970-9290.84299

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    Figures

  [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7]
 
 
    Tables

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