U373-MG cells express PepT2 and accumulate the fluorescently tagged dipeptide-derivative ß-Ala-Lys-N(ε)-AMCA.

Aim of this study was to examine the dipeptide transport of ß-Ala-Lys-N(ɛ)-AMCA in the human glioma cell line U373-MG and its potential regulation by diverse hormones and culture media. A mixed glial primary cell culture of the newborn rat served as reference cell system. ß-Ala-Lys-N(ɛ)-AMCA (ß-Ala-Lys-N(ɛ)-7-amino-4-methyl-coumarin-3-acetic acid) is a highly specific reporter substrate to investigate the dipeptide transport system PepT2. We were able to demonstrate that U373-MG cells express PepT2-mRNA and translocate ß-Ala-Lys-N(ɛ)-AMCA via PepT2 into the cytoplasm. Previous results demonstrated that ß-Ala-Lys-N(ɛ)-AMCA specifically accumulates in differentiated and dedifferentiated astrocytes but neither in differentiated nor dedifferentiated oligodendrocytes and in neurons. U373-MG cells were incubated with estradiol, testosterone, thyronine, dexamethasone, dibutyryl cyclic adenosine monophosphate and tetradecanoylphorbol acetate in order to detect potential substance-dependent changes in dipeptide uptake. There was no significant increase or decrease of ß-Ala-Lys-N(ɛ)-AMCA-uptake after stimulation. Northern blot analyses confirmed that PepT2-mRNA is expressed in U373-MG and glial cells but showed no regulation of PepT2-mRNA expression in both cell types. Future investigations might offer the opportunity of an anti-tumor therapy with cytotoxic agents linked to a dipeptide-derivative such as ß-Ala-Lys.

PepT2 transporter protein expression in human neoplastic glial cells and mediation of fluorescently tagged dipeptide derivative beta-Ala-Lys-Nepsilon-7-amino-4-methyl-coumarin-3-acetic acid accumulation.

OBJECT: The present study was aimed at analyzing the accumulation of the fluorescently tagged dipeptide derivative, beta-Ala-Lys-N(epsilon)-7-amino-4-methyl coumarin-3-acetic acid (AMCA), in primary cultures of human neoplastic glial cells. This molecule is a highly specific reporter used to investigate the dipeptide transport system hPepT2. METHODS: In this study the authors used immunocytochemical methods to determine the cell-specific accumulation of a small and fluorescently tagged reporter molecule named beta-Ala-Lys-N(epsilon)-AMCA to detect dipeptide transport capacity of neoplastic glial cells. Furthermore, specific mRNA levels were quantified using Northern blot analysis and the tissue distribution of hPepT2 mRNA transcripts was demonstrated with in-situ hybridization histochemical analysis. RESULTS: Recent fluorescent immunocytochemical analyses have revealed that beta-Ala-Lys-N(epsilon)-AMCA specifically accumulates within anaplastic cells of astrocytic lineage but not in anaplastic oligodendrocytes or neurons. Northern blot analysis demonstrated that human hPepT2 mRNA is specifically detected in primary cell cultures of human glioblastoma but not in oligodendroglioma. Moreover, in situ hybridization analyses revealed an astrocytic localization of hPepT2 transcripts in human glioblastoma and astrocytoma cells. The hPepT2 transcription levels were clearly dependent on the grade of glial cell differentiation: within low-grade gliomas (WHO Grade II), more hPepT2 mRNA was detected compared with tumors of a higher grade of dedifferentiation (WHO Grade IV). Analysis of expression levels of hPepT2 mRNA in human neoplastic glial cells xenografted into the brains of athymic rats (han rnu(+/+)) showed a markedly increased expression of hPepT2 after 2 weeks of growth in vivo compared with the primary counterparts grown in vitro. CONCLUSIONS: The authors concluded that expression of the hPepT2 transporter protein is a characteristic of glial cells of astrocytic lineage, and is dependent on the grade of astroglial cell differentiation and the extracellular matrix (here brain neuropil). The authors found that beta-Ala-Lys-N(epsilon)-AMCA is as an excellent reporter molecule for assessing neoplastic glial cell function and physiological characteristics.

Calcifying pseudoneoplasm of the inferior colliculus: an unusual location for a rare tumor: case report.

OBJECTIVE: Quadrigeminal plate lesions are rare and usually present with a silent clinical course. Tumors, vascular lesions, inflammatory and infectious processes have been described in this region. Calcifying pseudoneoplasms, also reported as fibro-osseous lesions, cerebral calculi, and brain stones, are unusual lesions in the central nervous system. They can be revealed by cranial radiography, computed tomography, and magnetic resonance imaging as calcified masses and should be differentiated from neoplastic, inflammatory, and vascular lesions. To the best of our knowledge, the occurrence of a calcifying pseudoneoplasm located at the quadrigeminal plate has not yet been reported. CLINICAL PRESENTATION: A 67-year-old woman with a 6-month history of several daily attacks of dizziness presented to our service. Magnetic resonance imaging studies revealed a tumor in the right inferior colliculus. This lesion was isointense on T1-weighted imaging, hypointense on T2-weighted imaging, and homogeneously enhanced with contrast. INTERVENTION: The lesion at the quadrigeminal plate was completely removed, and the patient was successfully treated without any new neurological deficit. At the time of follow-up, all preoperative symptoms had resolved. CONCLUSION: We report the first case of a calcifying pseudoneoplasm of the inferior colliculus. Complete surgical removal of this type of tumor is feasible. We propose surgical treatment in this location when this tumor becomes symptomatic.