|Year : 2018 | Volume
| Issue : 3 | Page : 207-209
Down regulation of serum protein expression in oral submucous fibrosis: A proteomic study
Nimi Susan Mathew, Prashanth Shenoy, Laxmikanth Chatra, KM Veena, Rachana V Prabhu, Tashika Kuttappa
Department of Oral Medicine and Radiology, Yenepoya Dental College, Mangalore, Karnataka, India
|Date of Submission||22-Apr-2018|
|Date of Acceptance||10-Jul-2018|
|Date of Web Publication||18-Oct-2018|
Dr. Prashanth Shenoy
Department of Oral Medicine and Radiology, Yenepoya Dental College, Derlakatte, Mangalore - 575 018, Karnataka
Source of Support: None, Conflict of Interest: None
| Abstract|| |
Background: Oral submucous fibrosis (OSMF) is a potentially malignant condition, which affects the oral cavity and the oropharynx. Despite the high prevalence rates of the disease and the morbidity and mortality related to the malignant transformation of 3%–19%, that is, OSMF into oral squamous cell carcinoma, there are fewer methods to identify the malignant transformation associated with OSMF. Dissecting the molecular aspects of these disorders will significantly contribute to improved prevention, early diagnosis, and clinical management. The complementary process involved in decreasing of the cellular components is called downregulation and the vice versa is known as upregulation. Materials and Methods: To predict the malignant transformation of OSMF, a study was conducted for quantitative proteomic profiling on the serum of histopathologically proven patients with OSMF to identify the candidate protein biomarkers. Among the differentially expressed proteins, only the downregulated proteins are included in the scope of this article. Results: During the study, 133 proteins were identified from the plasma of the case samples who histopathologically confirmed OSMF. These proteins were identified using mass spectrometric analysis. During the analysis, 14 proteins were differentially expressed. A fold change of more than 1.5 indicates upregulated proteins and less than 0.6 indicates downregulated proteins when compared with the control group. Conclusion: This study depicts the scope of the candidate protein biomarkers that can aid in definitive diagnosis of OSMF through a minimally invasive technique and help in the early detection and better treatment planning to reduce the morbidity of the disease in the future.
Keywords: Body fluid proteomics, OFFGEL electrophoresis, oral submucous fibrosis, protein biomarkers, proteome discoverer, secreted proteins, serum, strong cation exchange chromatography
|How to cite this article:|
Mathew NS, Shenoy P, Chatra L, Veena K M, Prabhu RV, Kuttappa T. Down regulation of serum protein expression in oral submucous fibrosis: A proteomic study. J Indian Acad Oral Med Radiol 2018;30:207-9
|How to cite this URL:|
Mathew NS, Shenoy P, Chatra L, Veena K M, Prabhu RV, Kuttappa T. Down regulation of serum protein expression in oral submucous fibrosis: A proteomic study. J Indian Acad Oral Med Radiol [serial online] 2018 [cited 2019 Mar 24];30:207-9. Available from: http://www.jiaomr.in/text.asp?2018/30/3/207/243662
| Introduction|| |
Proteins are a highly conserved group of protective cellular substances whose synthesis is increased in response to a variety of environmental or pathophysiological stress. Proteins are useful biomarkers for carcinogenesis in tissue and to alert the degree of differentiation and aggressiveness of cancer. Oral submucous fibrosis (OSMF) is a potentially malignant oral condition, which is caused by the habit of chewing areca nut. Since its first description in the 1950s, numerous epidemiological, biochemical, histopathological, and genetic studies have been reported. Although most studies point out to the cause and effect of areca nut, coadditive factors are also implicated in the progression and malignant transformation of this condition. Biochemical investigations have concentrated on outlining such changes in the blood, serum, or tissues of these patients and have given significant insights on the possible pathogenesis of OSMF. The aim of our study was to evaluate the proteins that are downregulated in the serum of biopsy-proven OSMF cases to find potential biomarkers.
| Materials and Methods|| |
The case and the control samples were obtained from the Department of Oral Medicine and Radiology, Yenepoya Dental College, Mangalore, after the approval of the Institutional Ethical Review Board. Blood samples were collected from five patients who were clinically and histopathologically proven to have OSMF. The case samples were compared with control samples of similar age and gender. These healthy volunteers were identified with chewing habit without lesion. A written consent was obtained from the individuals before undergoing biopsy procedure for histopathological confirmation. After the confirmation, a written consent was obtained for including the case samples in the study. The individuals who had a history of consuming areca nut in any form were selected for the study.
The blood samples were collected in a vacutainer with anticoagulant substance from the case and control groups. These samples were stored in an icebox and transported to the laboratory (YU-IOB).
Sample preparation and analysis
Depletion of high abundant proteins from plasma
The depletion of high abundant proteins was carried out with MARS 14 column (multiple affinity removal systems; Agilent Technologies, Santa Clara, CA, USA; Cat# 5188-6557; 4.6 × 50 mm) using high-performance liquid chromatography-based method with buffers A and B. About 40 μL of plasma volume was used for depletion, after which tryptic digestion of depleted plasma samples – insolution digestion of proteins – was carried out. After that, Tandem mass tag (TMT) labeling, that is, labeling of the samples, was carried out with TMT mass tags 10-plex kit (Thermo Fisher Scientific, Germany). This step was followed by strong cation exchange (SCX) fractionation which was carried out to resolve the peptides based on their ionic property. After that Liquid chromatography–mass spectrometry analysis was done where the peptides from SCX fractionation were analyzed on Thermo Scientific Orbitrap Fusion Tribrid mass spectrometer (Thermo Fisher Scientific) connected to Easy-nLC-1200 nanoflow liquid chromatography system (Thermo Fisher Scientific). MS/MS data analysis was done: mass spectrometry-derived data were analyzed with Proteome Discoverer software, version 2.1 (Thermo Fisher Scientific) with SEQUEST and Mascot (version 2.5; Matrix Science, London, UK) search algorithms. It was searched against Human Ref Seq 81 Protein Database supplemented with common contaminants.
| Results and Observation|| |
During the study, 133 proteins were identified from the plasma of the case samples who histopathologically confirmed OSMF. These proteins were identified using mass spectrometric analysis. During the analysis, 14 proteins were differentially expressed. A fold change of more than 1.5 indicates upregulated proteins and less than 0.6 indicates downregulated proteins when compared with the control group [Table 1].
|Table 1: Gene characteristics of the down-regulated proteins expressed in the serum of oral submucous fibrosis|
Click here to view
| Discussion|| |
APOA2 protein: Apolipoprotein A2 (gene ID: 336)
This gene encodes apolipoprotein (apo-) A-II, which is the second most abundant protein of the high-density lipoprotein (HDL) particles. The protein is found in plasma as a monomer, homodimer, or heterodimer with apolipoprotein D. Defects in this gene may result in apolipoprotein A-II deficiency or hypercholesterolemia.
This protein is involved in various biological processes such as acute inflammatory response, cellular protein metabolism, cholesterol efflux, cholesterol homeostatsis, chylomicron assembly, chylomicron remodeling, diacylglycerol catabolic process, HDL particle assembly basically host–virus interaction, and lipid transport.
GSTZ1 protein (gene ID: 2954)
This gene is a member of the glutathione S-transferase super-family, which encodes multifunctional enzymes important in detoxification of electrophilic molecules, including carcinogens, mutagens, and several therapeutic drugs, by conjugation with glutathione. This enzyme catalyzes the conversion of maleylacetoacetate to fumarylacetoacatate, which is one of the steps in the phenylalanine/tyrosine degradation pathway. Deficiency of a similar gene in mouse causes oxidative stress. Several transcript variants of this gene encode multiple protein isoforms.
This protein is involved in glutathione derivative biosynthetic process, glutathione metabolic process, and L-phenylalanine catabolic process. Basically, these proteins are identified in phenylalanine and tyrosine catabolism.
APOC1 protein (gene ID: 341)
This gene encodes a member of the apolipoprotein C1 family. This gene is expressed primarily in the liver, and it is activated when monocytes differentiate into macrophages. The encoded protein plays a central role in HDL and very low-density lipoprotein metabolism. This protein has also been shown to inhibit cholesteryl ester transfer protein in plasma. A pseudogene of this gene is located 4 kb downstream in the same orientation, on the same chromosome. This gene is mapped to chromosome 19, where it resides within an apolipoprotein gene cluster. Alternative splicing and the use of alternative promoters result in multiple transcript variants.
This protein is involved in biological processes such as cholesterol efflux, cholesterol metabolic process, HDL remodeling, and lipid and lipoprotein metabolic process, basically in lipid transport.
Across the data collected, we compared the correlation between age, duration of habit, and degree of epithelial dysplasia with the differentially expressed proteins among the case and control groups. However, no correlation was found in the study population. A larger cohort should be studied to check for further comparisons.
| Conclusion|| |
This is the first de novo proteomic biomarker study of human serum in OSMF. During the study, we identified 133 proteins from OSMF serum proteome using high-resolution mass spectrometry. This study revealed 11 proteins to be upregulated and 3 proteins to be downregulated when compared with the control group. During the mass spectrometric analysis, the differentially expressed downregulated proteins showed no correlation when compared with the protein database available for oral squamous cell carcinoma. This could be because the samples selected for the study were in their early stages, but these observations could differ in the advanced stages of OSMF.
Hence, validation of proteins in a larger cohort of clinical samples in clinically appropriate conditions and stages could prove useful in the identification of potential biomarkers for the early detection and transformation of OSMF into squamous cell carcinoma. This study depicts the scope of the candidate protein biomarkers, which can aid in definitive diagnosis of OSMF through a minimally invasive technique and help in early detection and better treatment planning to reduce the morbidity of the disease in the future.
The knowledge and education of dentists in detecting oral cancer at its precancerous phase is the key to prevent its progression to later stages. To improve early detection, it is imperative to increase the healthcare providers. The depth of knowledge about oral cancer, their risk factors, and the most common oral precancerous conditions will help the oral physicians to diagnose and treat the patients in an advanced level.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
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.
] [Full text]
Kamath VV, Satelur K, Komali Y. Biochemical markers in oral submucous fibrosis: A review and update. Dent Res J (Isfahan) 2013;10:576-84.