Petra-Maria Schumm-Draeger

Rhythmic gene expression in pituitary depends on heterologous sensitization by the neurohormone melatonin

In mammals, many daily cycles are driven by a central circadian clock, which is based on the cell-autonomous rhythmic expression of clock genes. It is not clear, however, how peripheral cells are able to interpret the rhythmic signals disseminated from this central oscillator. Here we show that cycling expression of the clock gene Period1 in rodent pituitary cells depends on the heterologous sensitization of the adenosine A2b receptor, which occurs through the nocturnal activation of melatonin mt1 receptors. Eliminating the impact of the neurohormone melatonin simultaneously suppresses the expression of Period1 and evokes an increase in the release of pituitary prolactin. Our findings expose a mechanism by which two convergent signals interact within a temporal dimension to establish high-amplitude, precise and robust cycles of gene expression.

Molecular detection of thyroglobulin mRNA transcripts in peripheral blood of patients with thyroid disease by RT-PCR

The sensitive detection of circulating tumour cells in patients with differentiated thyroid cancer may precede the detection of relapse by other diagnostic studies – such as serum thyroglobulin – and thus may have important therapeutic and prognostic implications. We performed reverse transcription-polymerase chain reaction (RT-PCR) on blood samples from patients diagnosed with thyroid disease using two different RT-PCR sensitivities. Additionally, tissue specificity of TG mRNA-expression was determined using RNA extracts from 27 different human tissues. The lower limit of detection was 50–100 TG mRNA producing cells/ml blood using a ‘normal’ RT-PCR sensitivity and 10–20 cells/ml blood using a ‘high’ sensitivity. With the normal sensitivity TG mRNA was detected in 9/13 patients with thyroid cancer and metastasis, 63/137 patients with a history of thyroid cancer and no metastasis, 21/85 with non-malignant thyroid disease and 9/50 controls. With the high sensitivity TG mRNA was detected in 11/13 patients with thyroid cancer and metastasis, 111/137 patients with a history of thyroid cancer and no metastasis, 61/85 with non-malignant thyroid disease and 41/50 controls. Interestingly, using the normal RT-PCR sensitivity TG mRNA transcripts are specific for thyroid tissue and detectable in the peripheral blood of controls and patients with thyroid disease, which correlates with a diagnosis of metastasized thyroid cancer. However, with a high RT-PCR sensitivity, TG mRNA expression was found not to be specific for thyroid tissue and was not correlated with a diagnosis of thyroid cancer in patients. As a consequence, to date TG mRNA detected by RT-PCR in the peripheral blood cannot be recommended as a tumour marker superior to TG serum-level.

T lymphocytes, CD68-positive cells and vascularisation in thyroid carcinomas

Immunohistochemical detection and quantification of CD3-and CD45RO-positive lymphocytes and CD68-positive cells in 75 thyroid carcinomas of follicular cell origin revealed rising levels for these parameters associated with dedifferentiation. A parallel trend towards reduction of vascularisation, determined as CD31-positive blood vessels, with decreasing differentiation became evident, statistically only significant when well-differentiated follicular and anaplastic carcinomas were compared. Positive correlations could be demonstrated between the density of CD68-, CD3-, and CD45RO-positive cells as well as between the density of CD68-, and CD3-, and CD45RO-positive cells and vascularisation. These correlations were expected, as the interaction of CD68-positive cells and T lymphocytes results in the production of angiogenesis factors, ultimately leading to better vascularisation of the tumour. Nevertheless, the tumour cells themselves are variously capable of producing angiogenic substances. The obvious lack of positive correlation between the density of tumour-infiltrating cells determined in this study and vascularisation, despite reduced vascularisation in less differentiated tumours that contained increasing numbers of tumour-infiltrating cells, seems to be due to functional heterogeneity of morphologically similar, tumours.

Circulating calcitonin and carcinoembryonic antigen m-RNA detected by RT-PCR as tumour markers in medullary thyroid carcinoma.

Detection of local relapse or metastasis in medullary thyroid carcinoma (MTC) continue to pose a major diagnostic challenge. Besides established diagnostic studies such as serum calcitonin (CT) and carcinoembryonic antigen (CEA), molecular detection of circulating tumour cells may be an additional diagnostic tool for the early detection of disease recurrence. We performed reverse transcription-polymerase chain reaction (RT-PCR) on blood samples from patients diagnosed with MTC disease using primers specific for CT and CEA, respectively. CT mRNA was not detectable in peripheral blood of all patients with MTC (n = 11) and all controls (n = 32). CEA mRNA was significantly more often detected patients with MTC (72.7%) than in controls (34.4%; p = 0.038; Fisher exact test). With an example of a patient with MTC and massive tumour mass in the neck we demonstrate the failure of detection of CT mRNA over a period of 6 months, whereas CEA mRNA could be detected in peripheral blood of this patient. As a consequence, CT mRNA detected by RT-PCR in the peripheral blood can not be recommended as a tumour marker in MTC. However, the use of carcinoembryonic mRNA may provide a significant improvement in diagnosis of recurrent disease in MTC.

Polymerase chain reaction in the detection of tumor cells: new approaches in diagnosis and follow-up of patients with thyroid cancer.

Thyroid cancers are the most common endocrine malignancies and are being diagnosed with increasing frequency. In addition to other measures, diagnosis is based on fine-needle aspiration cytology examination. Recently, new assays using reverse transcription-polymerase chain reaction (PCR) are being tested to improve sensitivity and specificity of primary diagnosis and detection of recurrent thyroid cancer. In the preoperative diagnosis of thyroid cancer, several tissue- and/or tumor-specific mRNA have been described and in several cases, a higher sensitivity and specificity could be achieved using molecular techniques compared to conventional methods. In the postoperative follow-up of patients with thyroid cancer, conflicting data have been published and the use of PCR techniques revealed several problems of the molecular approach, which are based on some technical as well as biologic limitations. Despite these problems, which are discussed in detail in this review, molecular techniques may nevertheless improve the sensitivity and accuracy of fine-needle aspiration of thyroid nodules, fine-needle aspiration of metastases, and detection of recurrent disease in peripheral blood samples.

Endokrinologie: Teil II

(Insulin-Hypoglyk-ämie-Test oder Metopiron-Kurztest) angeschlossen werden [8, 9]. Neben der Glukokortikoid(primäre und sekundäre NNR-Insuffizienz) und der Mineralokortikoidsubstitution (nur primäre NNR-Insuffizienz) sollte bei jedem Patienten mit NNRInsuffizienz eine zusätzliche Substitution mit dem NNR-Androgen Dehydroepiandrosteron (DHEA) erwogen werden [10]. Bei Patientinnen mit NNR-Insuffizienz konnte in zwei Studien eindeutig ein verbessertes psychisches Wohlbefinden unter einer Substitution mit 50 mg DHEA/die belegt werden [11, 12]. Da in einer dieser Studien auch männliche Patienten mit NNR-Insuffizienz untersucht und ähnliche Effekte gemessen wurden, wird als Ursache neben der androgenen Wirkung eine zusätzliche „neurosteroidale“ Wirkung des DHEA postuliert [12]. Obwohl weder bei Patienten mit Autoimmunadrenalitis (80–90% der Patienten mit Morbus Addison) noch bei Patienten mit sekundärer NNRInsuffizienz das Nebennierenmark morphologisch verändert ist, findet man sowohl bei primärer [13] als auch bei sekundärer NNR-Insuffizienz [14] eine eingeschränkte Adrenalinsynthese des Nebennierenmarks. Dies wird auf eine funktionelle Einschränkung der Nebennierenmarkenzyme bei fehlendem Glukokortikoidangebot aus der Zona fasciculata zurückgeführt. Hatte man bisher dem daraus resultierenden „Adrenalinmangel“ keine klinische Bedeutung zugeordnet, zeigt eine neuere Studie, dass dies zu einer eingeschränkten „Stressfähigkeit“ führen kann [14]. Ob sich hieraus evtl. eine Indikation zu einer „Ersatztherapie“ des Nebennierenmarks ergibt, ist derzeit bei fehlenden weiterführenden Studien nicht abzusehen, aufgrund der pharmakologischen Voraussetzungen (parenterale Adrenalingabe) aber eher unwahrscheinlich. Nebennierenrinde und -mark