Abstract
Objective: 1) Evaluate the expression of hexosoaminidases (HEXA and HEXB) genes in cholesteatoma fibroblasts (CF) and in the inflamed synoviocytes obtained from patients who suffered from rheumatoid arthritis; and 2) Evaluate in vitro inhibition potential of pyrimethamine.
Method: Cholesteatoma fibroblasts (study group), synoviocytes (comparison group), and retroauricular skin fibroblasts (control group) were used. The following solutions of the selected inhibitor (pyrimethamine) were added to the cell culture: 20 mcg/mL, 10 mcg/mL, 3 mcg/mL, and 1.5 mcg/mL. RT-PCR was carried out before and after adding pyrimethamine.
Results: RT-PCR established elevated expression of HEXA and HEXB mRNA in cholesteatoma. HEXA mRNA in CF was 6 times higher than in controls. HEX mRNA expression is regulated by pyrimethamine. Inhibition of HEXA and HEXB mRNA expression was achieved when the highest concentration of PYR was used. Low concentrations of pyrimethamine upregulated HEX gene in CF. A difference in HEXA and HEXB expression was observed in synoviocytes. The highest concentration of pirymethamine (10 and 20 mcg/mL) caused the least elevation of HEXA expression. A slight decrease of HEXB expression was observed under the concentration of pirymethamine: 1.3 and 3 mcg/mL.
Conclusion: Pyrimethamine, depending on its concentration, contributes to regulating HEX gene expression in CF and controls. Our preliminary data do not let us establish the concentration of pyrimethamine that may significantly inhibit HEXA and HEXB expression. Further study may put new insight into the usefulness of investigating the localized inflammation in cholesteatoma.
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