Peripheral blood neutrophil granulocytes from patients with head and neck squamous cell carcinoma functionally differ from their counterparts in healthy donors.

Solid tumors such as head and neck squamous cell carcinoma (HNSCC) display an intense interaction between tumoral factors and the immune system. Functional modulation of tumor-infiltrating and peripheral blood immune cells plays an important role during tumor progression. In this pilot study we compared biological functions of polymorphonuclear granulocytes (PMN) from the peripheral blood of HNSCC patients and healthy subjects. PMN were simultaneously isolated from the peripheral blood of HNSCC patients and healthy donors for functional analysis (apoptosis, production of reactive oxygen species (ROS), cytokine release and immunophenotyping). PMN from HNSCC patients showed a significantly lower inducible production of ROS (P = 0.02) and reduced spontaneous apoptosis (P= 0.008) compared with PMN from healthy donors. Under standard culture conditions, there was no significant difference regarding the release of inflammatory cytokines between PMN from HNSCC patients and PMN from healthy donors. Confirming previous observations, serum concentrations of PMN-related cytokines were significantly higher in the peripheral blood of HNSCC patients than in that of controls. Importantly, immunophenotyping revealed an increased number of immature PMN in PMN fractions isolated from HNSCC patients. Peripheral blood PMN from HNSCC patients and healthy donors show distinct functional differences. The presence of increased numbers of immature stages of PMN in HNSCC patients may partly contribute to the changes observed. After recruitment to and infiltration of the tumor, PMN may be further modulated in the local tumor microenvironment. This pilot study justifies functional analyses of myeloid cells in larger cohorts of patients with HNSCC.

Dendritic cell generation and CD4+ CD25high FOXP3+ regulatory t cells in human head and neck carcinoma during radio-chemotherapy.

BACKGROUND: Regulatory T cells (Treg) and dendritic cells (DC) play an important role in tumor immunity and immune escape. However, their interplay and the effects of anti-cancer therapy on the human immune system are largely unknown. METHODS: For DC generation, CD14⊃+ monocytes were enriched by immunomagnetic selection from peripheral blood of advanced head and neck squamous cell carcinoma (HNSCC) patients and differentiated into immature DC using GM-SCF and IL-4. DC maturation was induced by addition of TNFα. The frequency of CD4⊃+CD25⊃highFOXP3⊃+ Treg in HNSCC patients was analyzed before and after radio-chemotherapy (RCT) by four-color flow cytometry. RESULTS: In HNSCC patients, the frequency of Treg (0.33 ± 0.06%) was significantly (p = 0.001) increased compared to healthy controls (0.11 ± 0.02%), whereas RCT had variable effects on the Treg frequency inducing its increase in some patients and decrease in others. After six days in culture, monocytes of all patients had differentiated into immature DC. However, DC maturation indicated by CD83 up-regulation (70.7 ± 5.5%) was successful only in a subgroup of patients and correlated well with lower frequencies of peripheral blood Treg in those patients. CONCLUSION: The frequency of regulatory T cells is elevated in HNSCC patients and may be modulated by RCT. Monocyte-derived DC in HNSCC patients show a maturation deficiency ex vivo. Those preliminary data may have an impact on multimodality clinical trials integrating cellular immune modulation in patients with advanced HNSCC.

In vitro chemosensitivity of head and neck cancer cell lines.

BACKGROUND: Systemic treatment of head and neck squamous cell carcinoma (HNSCC) includes a variety of antineoplastic drugs. However, drug-resistance interferes with the effectiveness of chemotherapy. Preclinical testing models are needed in order to develop approaches to overcome chemoresistance. - METHODS: Ten human cell lines were obtained from HNSCC, including one with experimentally-induced cisplatin resistance. Inhibition of cell growth by seven chemotherapeutic agents (cisplatin, carboplatin, 5- fluorouracil, methotrexate, bleomycin, vincristin, and paclitaxel) was measured using metabolic MTT-uptake assay and correlated to clinically-achievable plasma concentrations. - RESULTS: All drugs inhibited cell growth in a concentration-dependent manner with an IC50 comparable to that achievable in vivo. However, response curves for methotrexate were unsatisfactory and for paclitaxel, the solubilizer cremophor EL was toxic. Cross-resistance was observed between cisplatin and carboplatin. - CONCLUSION: Chemosensitivity of HNSCC cell lines can be determined using the MTT-uptake assay. For DNA-interfering cytostatics and vinca alkaloids this is a simple and reproducible procedure. Determined in vitro chemosensitivity serves as a baseline for further experimental approaches aiming to modulate chemoresistance in HNSCC with potential clinical significance.

Interaction of volatile anesthetics with human Kv channels in relation to clinical concentrations.

BACKGROUND: Recent evidence shows that inhibition of human Kv3 channels by intravenous anesthetics occurs at clinical concentrations. The effects of volatile anesthetics on these human ion channels are unknown. This study was designed to establish whether minimum alveolar concentrations (MAC) of halothane, enflurane, isoflurane, and desflurane exhibit effects on Kv3 channeLs. To obtain an indication whether these findings may be specific to Kv3 channels, the effects of enflurane and isoflurane on human Kv1.1 channels were also investigated. METHODS: Kv3 channels natively expressed in SH-SY5Y cells and Kv1.1 channels expressed in HEK293 cells were measured with the whole cell patch clamp technique by standard protocols. Concentrations of volatile anesthetics were determined by gas chromatography. RESULTS: Halothane, enflurane, isoflurane, and desflurane reversibly inhibited Kv3 channels in a concentration-dependent manner. Concentrations at half-maximal effect (IC50 values) ranged between 1,800 and 4,600 microM. Hill coefficients were between 1.7 and 2.5. IC50 values for inhibition of Kv1.1 channels were 2,800 and 5,200 microM, and Hill coefficients were 3.9 and 5.6 for enflurane and isoflurane, respectively. CONCLUSION: Volatile anesthetics inhibit human Kv3 channels at clinical concentrations. At 1-3 MAC, inhibition would account on average for 2-12%. Inhibition would be highest with enflurane (between 3% and 22%) and lowest with isoflurane (between 0.2% and 3%). Kv1.1 channels would only be inhibited by enflurane at clinical concentrations (2% at 2 MAC and 8% at 3 MAC). Whether the degree of K channel inhibition by volatile anesthetics may contribute to their clinical action needs further study.

[Epileptic seizures from etomidate? The human Kv1.1 potassium channel in humans]. / Epileptische Anfälle durch Etomidat? Untersuchung an Kv1.1-Kaliumkanälen des Menschen.

OBJECTIVE: It is a matter of dispute whether etomidate exhibits an epileptogenic action. It is also disputed whether frequently observed myocloni induced by etomidate are related to seizure-like activity in the EEG. The clinical effects of anaesthetic agents reflect their molecular action. A possible epileptogenic action of etomidate should, therefore, result from action on a molecular target that has yet to be identified. Suppression of Kv1.1 channels may be associated with epilepsy in men. Inhibition of Kv1.1 channels by etomidate at clinically relevant concentrations would, thus, argue in favour of an epileptogenic action of etomidate. METHODS: Patch-clamp recordings of the pharmacological action of etomidate on cloned and functionally expressed human Kv1.1 channels. RESULTS: Etomidate inhibits human Kv1.1 channels in a concentration-dependent and reversible manner. The IC50-value is 400 microM, the Hill-coefficient is 2. The ratio of free clinical plasma concentrations and the IC50-value is 0.006. At plasma concentrations determined in a clinical setting Kv1.1 channels would be suppressed by less than 0.01%. CONCLUSION: The small effect of etomidate on human Kv1.1 channels at these concentrations questions an epileptogenic action of etomidate caused by inhibition of Kv1.1 channels.