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Reduced 5-methylcytosine level as a potential progression predictor in patients with T1 or non-invasive urothelial carcinoma.

Chung CJ, Chang CH, Chuu CP, Yang CR, Chang YH, Huang CP, Chen WC, Chung MC, Chang H - Int J Mol Sci (2014)

Bottom Line: Low DNMT1 levels were only associated with UC with squamous differentiation (p = 0.0365).Neither 5-MeC nor DNMT1 levels were associated with UC recurrence.In conclusion, a low 5-MeC level could predict the progression of UC invasion into muscle.

View Article: PubMed Central - PubMed

Affiliation: Department of Health Risk Management, College of Public Health, China Medical University, Taichung 40402, Taiwan. cjchung1010@gmail.com.

ABSTRACT
This study aims to elucidate the level of DNA methylation in urothelial carcinomas (UCs) using 5-methylcytosine (5-MeC) immunohistochemistry (IHC). We examined the relationship among 5-MeC levels, DNA (cytosine-5)-methyltransferase 1 (DNMT1) immunostaining levels, and clinicopathologic features. Tissue samples included 23 normal urothelia and 150 urothelial neoplasia, which comprised 40 non-invasive and 110 invasive UCs. The levels of 5-MeC and DNMT1 were assessed based on their immunoreactivities and then divided into low and high levels. In addition, we collected information on clinical variables, pathologic features, and recurrent status from patient questionnaires and medical records. Chi-square test and multivariate logistic regression model were used for analyses. Results showed that 5-MeC levels were positively associated with DNMT1 levels in UC (p = 0.0288). Both 5-MeC and DNMT1 were low in approximately 50% (76/150) of UC. The percentage of low 5-MeC levels was higher in invasive UC (65/110; 59%) than in normal urothelia (2/23; 13%) and non-invasive UC (18/40; 45%). Clinical factors were independently associated with low 5-MeC levels after adjusting for age and sex, including cancer stages II-IV, presence of UC in situ, and marked inflammation. Low 5-MeC levels in stage I invasive UC were not significantly different from those of non-invasive tumors (p = 0.8478). Low DNMT1 levels were only associated with UC with squamous differentiation (p = 0.0365). Neither 5-MeC nor DNMT1 levels were associated with UC recurrence. In conclusion, a low 5-MeC level could predict the progression of UC invasion into muscle.

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Related in: MedlinePlus

Immunohistochemical intensity representated as H-score by imaging software counts. The A–C photography are the sample 1. The D–F photography are the sample 2. The positive color in our experiment is brown. The photography (A,D) are initially changed to brown-color channels (B,E) before intensity counting. The positive pixel counts (C,F) shows red color as strong positivity (staining intensity grade 3), orange color as medium positivity (grade 2), yellow color as weak positivity (grade 1) and blue color as negativity (grade 0), and finally quantifies the areas and intensities to get H-scores ranged from 0 to 300. The H-score of sample 1 by 175 is higher than sample 2 by 133. Bar = 200 µm.
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ijms-16-00677-f003: Immunohistochemical intensity representated as H-score by imaging software counts. The A–C photography are the sample 1. The D–F photography are the sample 2. The positive color in our experiment is brown. The photography (A,D) are initially changed to brown-color channels (B,E) before intensity counting. The positive pixel counts (C,F) shows red color as strong positivity (staining intensity grade 3), orange color as medium positivity (grade 2), yellow color as weak positivity (grade 1) and blue color as negativity (grade 0), and finally quantifies the areas and intensities to get H-scores ranged from 0 to 300. The H-score of sample 1 by 175 is higher than sample 2 by 133. Bar = 200 µm.

Mentions: The 5-MeC- or DNMT1-positive cells presented a brown color and were localized at the nuclei and/or cytoplasm of the normal urothelia and urothelial cancer cells. The interstitial stroma exhibited 5-MeC and DNMT1 immunonegativity. H-scoring system was used to assess staining intensity by employing imaging software (Aperio Technologies Inc., Vista, CA, USA) to obtain an objective evaluation and avoid subjective interpretation. The grades of staining intensity were four-tiered as follows: 0, negative staining; 1, weak staining; 2, medium staining; and 3, strong staining. The H-score represented the sum of the mean value in each grade multiplied by the proportion of positive cells for each tissue core (Figure 3).


Reduced 5-methylcytosine level as a potential progression predictor in patients with T1 or non-invasive urothelial carcinoma.

Chung CJ, Chang CH, Chuu CP, Yang CR, Chang YH, Huang CP, Chen WC, Chung MC, Chang H - Int J Mol Sci (2014)

Immunohistochemical intensity representated as H-score by imaging software counts. The A–C photography are the sample 1. The D–F photography are the sample 2. The positive color in our experiment is brown. The photography (A,D) are initially changed to brown-color channels (B,E) before intensity counting. The positive pixel counts (C,F) shows red color as strong positivity (staining intensity grade 3), orange color as medium positivity (grade 2), yellow color as weak positivity (grade 1) and blue color as negativity (grade 0), and finally quantifies the areas and intensities to get H-scores ranged from 0 to 300. The H-score of sample 1 by 175 is higher than sample 2 by 133. Bar = 200 µm.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4307268&req=5

ijms-16-00677-f003: Immunohistochemical intensity representated as H-score by imaging software counts. The A–C photography are the sample 1. The D–F photography are the sample 2. The positive color in our experiment is brown. The photography (A,D) are initially changed to brown-color channels (B,E) before intensity counting. The positive pixel counts (C,F) shows red color as strong positivity (staining intensity grade 3), orange color as medium positivity (grade 2), yellow color as weak positivity (grade 1) and blue color as negativity (grade 0), and finally quantifies the areas and intensities to get H-scores ranged from 0 to 300. The H-score of sample 1 by 175 is higher than sample 2 by 133. Bar = 200 µm.
Mentions: The 5-MeC- or DNMT1-positive cells presented a brown color and were localized at the nuclei and/or cytoplasm of the normal urothelia and urothelial cancer cells. The interstitial stroma exhibited 5-MeC and DNMT1 immunonegativity. H-scoring system was used to assess staining intensity by employing imaging software (Aperio Technologies Inc., Vista, CA, USA) to obtain an objective evaluation and avoid subjective interpretation. The grades of staining intensity were four-tiered as follows: 0, negative staining; 1, weak staining; 2, medium staining; and 3, strong staining. The H-score represented the sum of the mean value in each grade multiplied by the proportion of positive cells for each tissue core (Figure 3).

Bottom Line: Low DNMT1 levels were only associated with UC with squamous differentiation (p = 0.0365).Neither 5-MeC nor DNMT1 levels were associated with UC recurrence.In conclusion, a low 5-MeC level could predict the progression of UC invasion into muscle.

View Article: PubMed Central - PubMed

Affiliation: Department of Health Risk Management, College of Public Health, China Medical University, Taichung 40402, Taiwan. cjchung1010@gmail.com.

ABSTRACT
This study aims to elucidate the level of DNA methylation in urothelial carcinomas (UCs) using 5-methylcytosine (5-MeC) immunohistochemistry (IHC). We examined the relationship among 5-MeC levels, DNA (cytosine-5)-methyltransferase 1 (DNMT1) immunostaining levels, and clinicopathologic features. Tissue samples included 23 normal urothelia and 150 urothelial neoplasia, which comprised 40 non-invasive and 110 invasive UCs. The levels of 5-MeC and DNMT1 were assessed based on their immunoreactivities and then divided into low and high levels. In addition, we collected information on clinical variables, pathologic features, and recurrent status from patient questionnaires and medical records. Chi-square test and multivariate logistic regression model were used for analyses. Results showed that 5-MeC levels were positively associated with DNMT1 levels in UC (p = 0.0288). Both 5-MeC and DNMT1 were low in approximately 50% (76/150) of UC. The percentage of low 5-MeC levels was higher in invasive UC (65/110; 59%) than in normal urothelia (2/23; 13%) and non-invasive UC (18/40; 45%). Clinical factors were independently associated with low 5-MeC levels after adjusting for age and sex, including cancer stages II-IV, presence of UC in situ, and marked inflammation. Low 5-MeC levels in stage I invasive UC were not significantly different from those of non-invasive tumors (p = 0.8478). Low DNMT1 levels were only associated with UC with squamous differentiation (p = 0.0365). Neither 5-MeC nor DNMT1 levels were associated with UC recurrence. In conclusion, a low 5-MeC level could predict the progression of UC invasion into muscle.

Show MeSH
Related in: MedlinePlus