EXPRESSION OF HEXOKINASE II IN ORAL KERATOTIC LESIONS WITH OR WITHOUT INFLAMMATION
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1
Vita-Salute San Raffaele University, Department of Dentistry, Italy
Aim. Keratotic lesions, histologically classified as areas of hyperorthokeratosis and hyperparakeratosis, are the most recurrent clinical finding in the oral cavity. Their etiology is various, but frequently the cause is due to the friction against sharp dental edges and incongruous prostheses and also parafunctional habits might play a role as well. However, other inflammatory diseases such as oral lichen planus and lichenoid reactions have the clinical appearance of keratoses. Hexokinase II is an enzyme that is capable to phosphorylate glucose to produce glucose-6-phosphate, representing the first step in most of the glucose metabolic pathways and it is expressed mainly in insulin-dependent tissues such as skeletal muscle, cardiac muscle, and fatty tissues. This enzyme can bind to the outer mitochondrial membrane, modulating the inflammatory response and anti-apoptotic effects, in favor of the cell survival, especially in cancer cells.
Materials and Methods. This study has been performed in a cohort of 35 patients, afferent the Oral Medicine and Oral Pathology Unit, Dept. of Dentistry, San Raffaele Hospital Milano, showing a clinical lesion compatible with keratosis. In order to confirm the objective and clinical diagnosis of the oral medicine expert, after obtaining an informed consent, incisional biopsies were performed, following the 14 days-rule. The specimens were routinely processed by the pathologist in order to highlight the presence of any sign of chronic lichenoid inflammation that presented the typical lymphocytic band aspect in the submucosa. Serial sections from the samples were dyed with the immunohistochemical technique using the streptavidin-peroxidase method and monoclonal antibodies anti-HK II (GeneTex, CA USA). The staining intensity was classified as 0,1,2,3 for respectively no staining, weak, moderate and strong intensity by two observers, that analyzed independently the cellular pattern of expression to exclude any possible bias before conveying to the final evaluation. Statistical analysis was performed using a JMP 9.01 software, using the Pearson Chi-square, and whereas it was possible, the Fischer's exact test.
Results. The pathologic examination allowed the stratification of the samples by the presence of the chronic inflammation as follow: 22 keratotic lesions associated with chronic lichenoid infiltrate (62.86%) and 15 keratoses without (37.14%). Analyzing the staining intensity of HK II in those groups it was possible to underline some interesting findings; for the lesions characterized by inflammatory elements the staining was: absent in 1 out of 22 (4.55%), weak in 6 out of 22 (27.27%), moderate in 13 out of 22 (59.09%) and strong in 2 out of 22 (9.09%). On the counterpart, in the lesions without inflammation, the staining was absent in 6 out of 13 (46,16%), weak in 4 out of 13 (30.77%), moderate in 2 out of 13 (15.38%) and intense in 1 out of 13 (7.69%). These data entail a statistically significant difference in the staining intensity of HK II between the keratotic lesions with and without inflammation (p= 0.0130). Moreover, we proceeded to evaluate the patterns of expression of HK II that was localized more frequently in the cytoplasm of the epithelium cells. This expression was confirmed by the granular aspect of the dye that further suggested the well-known association with specific mitochondrial structures. However, it was also evidenced the peri-nuclear pattern of HK II in some tissutal elements. These characteristics have been evaluated in the two groups with the following results: in the lesions without inflammation, 1 sample out of 22 was not assessable (4.55%) because the dye wasn't present. 11 lesions out of 22 (50%) presented an expression just at the cytoplasmic level while 10 out of 22 (45.45%) showed some elements with peri-nuclear expression whereas, in the inflammatory lesions, 6 lesions out 13 were not assessable (46.15%) because the dye wasn't present. 4 lesions out of 13 (30.77%) showed a cytoplasmic pattern and 3 lesions out 13 (23.08%) had a peri-nuclear pattern too. These data revealed a statistical significance between the two groups (p=0.0118).
Discussion. The major marker's expression of glucose metabolism as HK II in the keratoses with an inflammatory substrate is an important finding: the evidence-based literature did not report any proof about the role of this enzyme in the inflammatory diseases of the oral mucosa. One of the supposed causes could be the increased energetic supply typical of the tissues stressed by an inflammatory stimulus. Furthermore, this enzyme is overexpressed in many of the human neoplasms, including the oral squamous cell carcinoma (OSCC). This could suggest a predominant role of inflammation in the oral carcinogenesis as already evidenced in other districts: in 4 out of 22 hyperkeratotic lesions associated with inflammation, (18.18%) there were dysplastic alterations, while dysplasia was observed only in 1 out of 13 keratoses without inflammation (7.69%). The inflammatory response involves the vascular, connective and epithelial structures leading to a local hypoxia. The enzyme HIF-1 (Hypoxia-Inducible Factor 1) induce the hyperexpression of proteins such as HK II involved in the glycolysis and cell survival. Moreover, it is involved as well in the regulation of inflammatory pathways through its action on the glucose metabolism. In fact, elevated levels of glucose could activate specific pro-inflammatory responses. To confirm the role of hypoxia it is remarkable to report the results of the expression of cellular patterns of HK II: the peri-nuclear pattern, that is prevalent in inflammatory lesions could be the epiphenomenon of a mitochondrial peri-nuclear clustering, due to the association between hexokinase and mitochondrial structures as previously investigated into an animal study. The mitochondrial clustering is essential for the transcription of genes connected with hypoxia. Consequently, the triad composed of inflammation, hypoxia and glucose metabolism should be further investigated in order to study the expression of other proteins involved in these processes and to confirm our results.
References
1. Tameka A. Clemons, Luis H. Toledo-Pereyra. Hexokinase: a glycolytic enzyme with an inflammatory ischemia and reperfusion connection. J Invest Surg 2015; 28(6): 301-302.
2. Roberts DJ, Miyamoto S. Hexokinase II integrates energy metabolism and cellular protection: Akting on mitochondria and TORCing to autophagy. Cell Death Differ 2015 Feb; 22(2): 248-257.
3. Al-Mehdi AB, Pastukh VM, Swiger BM et al. Perinuclear mitochondrial clustering creates an oxidant-rich nuclear domain required for hypoxia-induced transcription. Sci Signal
Keywords:
keratoses,
Inflammation,
glucose metabolism,
hypoxia,
Hexokinase II
Conference:
5th National and 1st International Symposium of Italian Society of Oral Pathology and Medicine., Ancona, Italy, 19 Oct - 20 Oct, 2018.
Presentation Type:
Poster Presentation
Topic:
Oral Diseases
Citation:
Moretti
M,
Lissoni
A,
Gastaldi
G,
Arrigoni
G,
Doglioni
C and
Abati
S
(2019). EXPRESSION OF HEXOKINASE II IN ORAL KERATOTIC LESIONS WITH OR WITHOUT INFLAMMATION.
Front. Physiol.
Conference Abstract:
5th National and 1st International Symposium of Italian Society of Oral Pathology and Medicine..
doi: 10.3389/conf.fphys.2019.27.00057
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Received:
05 Nov 2018;
Published Online:
09 Dec 2019.
*
Correspondence:
Dr. Mattia Moretti, Vita-Salute San Raffaele University, Department of Dentistry, Milan, Lombardy, 20132, Italy, fedemurkez@gmail.com