High frequency of vitamin B12 deficiency in asymptomatic individuals homozygous to MTHFR C677T mutation is associated with endothelial dysfunction and homocysteinemia.

The aim of this study was to examine the association of homozygosity for the methylenetetrahydrofolate reductase (MTHFR) C677T mutation and vitamin B12 deficiency in 360 asymptomatic individuals and to investigate forearm endothelial function in C677T homozygotes. MTHFR C677T mutation and levels of vitamin B12, folic acid, and homocysteine were measured in study participants. Frequency of homozygosity for the C677T mutation was 67/360 (18.6%). Homocysteine levels were elevated in homozygous compared with heterozygous subjects or those without the mutation (20.6 +/- 18.8 vs. 9.4 +/- 3.2 mumol/l; P < 0.0001). The number of subjects with vitamin B12 deficiency (<150 pmol/l) was significantly higher among the homozygote than the heterozygote subjects or subjects without mutation [20/67 (29.8%) vs. 27/293 (9.2%); P < 0.0001]. Homozygote subjects had 4.2 times higher probability of having B12 deficiency (95% confidence interval = 2.1-8.3). Forearm endothelial function was assessed in 33 homozygote and 12 control subjects. Abnormal endothelial function was observed in homozygous subjects and was worse in homozygote subjects with vitamin B12 deficiency. Endothelial function was normalized after B12 and folic acid treatment. We found that homozygosity for the C677T mutation is strongly associated with B12 deficiency. Coexistence of homozygosity for the C677T mutation and B12 deficiency is associated with endothelial dysfunction and can be corrected with vitamin B12 and folic acid treatment.

Telomerase activity and cytokeratin 20 as markers for the detection and followup of transitional cell carcinoma: an unfulfilled promise.

PURPOSE: Telomerase activity compensates for the erosion of chromosomes and it has been detected in a wide variety of human tumors. Cytokeratin 20, an intermediate filament of epithelial cells, is expressed particularly in the urinary tract. These 2 molecules are candidates to become markers for the detection and followup of bladder carcinoma. We evaluate whether each molecule may serve as a potential marker and whether the 2 combined would improve the detection or followup of bladder carcinoma in a noninvasive manner. MATERIALS AND METHODS: We obtained 44 morning urine samples from patients with transitional cell carcinoma patients and 26 from age matched patients with a wide variety of clinical disorders but no malignancy of any kind. A telomerase polymerase chain reaction-enzyme-linked immunosorbent assay kit was used to determine telomerase activity and cytokeratin 20 expression was determined by nested reverse transcriptase-polymerase chain reaction. RESULTS: All samples tested positive for cytokeratin 8 expression, which verified epithelial cells in the urine samples. Of the 44 transitional cell carcinoma cases of all stages and grades 37 (84.1%) were positive for telomerase activity, 36 (81.8%) were positive for cytokeratin 20 expression and 65.9% were double positive. Of the 29 controls with various clinical conditions other that malignancy 22 (75.9%) were positive for telomerase activity, 13 (44.83%) were positive for cytokeratin 20 expression and 34.6% were double positive. CONCLUSIONS: Telomerase activity and cytokeratin 20 expression are not specific for malignancy and may be detected in many nonmalignant pathological conditions. Therefore, their use as potential markers of bladder carcinoma should be carefully reevaluated.

Urinary cytokeratin 20 as a marker for transitional cell carcinoma.

OBJECTIVES: To determine if detection of cytokeratin 20 (CK20) gene expression, by reverse transcriptase-polymerase chain reaction (RT-PCR) in urine from transitional cell carcinoma (TCC) patients, can provide a new noninvasive tool for the follow-up of patients with urothelial carcinoma of the bladder. METHODS: Urine was collected from 95 patients previously diagnosed as TCC during their follow-up, and from 27 healthy volunteers. All patients had a transurethal resection of tumor or biopsies obtained from 'suspicious' areas in the bladder. RNA was extracted from cells collected from the urine and RT-PCR was performed with specific primers for the amplification of cytokeratin 8, a general marker for epithelial cells, and of CK 20, a marker for TCC urothelium. RESULTS: CK20 expression was detected in 86.7% of TCC patients, and only in 3.3% of healthy volunteers (specificity 96.7%). Strong correlation was found between tumor grade and expression of CK20 in urine. All grade III and IV tumors demonstrated positive CK20 expression (100% sensitivity), whereas the sensitivity for lower grades was between 71 and 80%. Among 11 patients with a previous biopsy-proven diagnosis of TCC and a current negative biopsy, in 9 patients CK20 expression was detected. Further follow-up of these patients for a period of 6 months revealed recurrence of TCC in 4 patients. CONCLUSION: CK20 detection in urine cells is a simple, noninvasive method with a high potential to become the marker of choice for monitoring and follow-up of TCC patients. More information is needed regarding CK20 expression in nonmalignant urological disease, to evaluate its use for routine screening purposes.