The effect of chemo- and chemoimmunotherapy on the presence of circulating melanoma cells in peripheral blood. Preliminary results.

Reverse transcription and polymerase chain reaction (RT/PCR) with primers specific for tyrosinase allow for a new method of early detection of individual melanoma cells in peripheral blood. Using this test the effect of chemo- and chemoimmunotherapy on the spread of early micrometastatic cancer cells has been evaluated. No significant correlations have been found between RT/PCR results on the one hand and stage of disease, a kind of the therapy protocol used and usage of the therapy as an adjuvant or palliative on the other hand. Thus, although the RT/PCR test for detection of circulating individual melanoma cells might help in identification of minimal residual disease in some patients, it has no application for routine staging of more advanced disease and in monitoring the response to therapy.

[Preliminary evaluation of a method for detection of individual malignant melanoma cells in peripheral blood while monitoring the course of the malignancy and its treatment]. / Wstepna ocena metody wykrywania pojedynczych komórek czerniaka zlossliwego we krwi obwodowej w monitorowaniu przebiegu choroby nowotworowej i jej leczenia.

Using reverse transcription and polymerase chain reaction (PCR) with primers specific for tyrosinase the individual melanoma cells were detected in peripheral blood of patients in different stages of disease, after excision of primary lesion and prior and after chemotherapy. No relation between stage of disease (including situations with overt generalized spread of melanoma) and probability of positive PCR reaction detecting transcript for tyrosinase gene was found. Many patients in III and IV stages were negative for prolonged periods. Therefore, this method cannot be used for monitoring of all patients, because many of them are negative prior as well as after chemotherapy. With regard to the effects of therapy, the patients differed one to another and although some persons positive prior treatment became negative thereafter, a similar number of initially negative patients became positive after treatment.

Serum cytokine levels (IL-4, IL-6, IL-8, G-CSF, GM-CSF) in burned patients.

The presence and concentration of selected cytokines (interleukin 4 (IL-4), interleukin 6 (IL-6), interleukin 8 (IL-8), granulocyte colony-stimulating factor (G-CSF) and granulocyte-macrophage colony-stimulating factor (GM-CSF) were evaluated in the sera of 12 burned patients (6-90 per cent body surface area). The presence of cytokines in the sera of 20 healthy volunteers (control group) was always undetectable (< 2 pg/ml). In sera of the burned patients the concentrations of IL-4 or GM-CSF were also below the test sensitivity levels, while G-CSF and IL-6 were present throughout all the observation period and IL-8 was detectable at the onset of massive infections. The serum concentrations of G-CSF and IL-6 increased during the episodes of clinically and bacteriologically detectable infections. Their increases were, however, observable 12-24 h later than the other infection symptoms. Similar increases in G-CSF and IL-6 levels have been detected during corrective surgery (covering of granulation tissue with skin grafts). It may be concluded that serum G-CSF and IL-6 levels in burned patients may be considered as diagnostic factors, but the delays in the reaction to the massive infection do not allow us to use them for predicting the time of onset of the infection.

Mouse model of infected wound.

Local Pseudomonas aeruginosa infection of experimental wound in mice was performed. Suspension of living bacteria was injected beneath skin muscle after skin window had been made on the back of animals. On day 1, 2, 3 and 8 mice were sacrificed, biopsies were taken, number of colony forming units in muscle specimens was estimated and histological examination of skin muscle flaps was performed. The number of bacteria dropped from the very beginning of experiment to the eighth day after infection, still remaining higher than critical value of clinical infection (10(5) CFU per 1 g). Histologically, healing processes beneath skin started from the third day and on the eight day the wounds were practically healed. Developed model of infected wound may be useful to study the antibacterial agents' effectiveness.