Götz Lehnerdt

Polymorphonuclear granulocytes in human head and neck cancer: enhanced inflammatory activity, modulation by cancer cells and expansion in advanced disease.

The progression of epithelial cancer is associated with an intense immunological interaction between the tumor cells and immune cells of the host. However, little is known about the interaction between tumor cells and polymorphonuclear granulocytes (PMNs) in patients with head and neck squamous cell carcinoma (HNSCC). In our study, we investigated systemic PMN‐related alterations in HNSCC, the role of tumor‐infiltrating PMNs and their modulation by the tumor microenvironment. We assessed the infiltration of HNSCC tissue by PMNs (retrospectively) and systemic PMN‐related alterations in blood values (prospectively) in HNSCC patients (n = 99 and 114, respectively) and control subjects (n = 41). PMN recruitment, apoptosis and inflammatory activity were investigated in an in vitro system of peripheral blood PMNs and a human HNSCC cell line (FaDu). HNSCC tissue exhibited considerable infiltration by PMNs, and strong infiltration was associated with poorer survival in advanced disease. PMN count, neutrophil‐to‐lymphocyte ratio and serum concentrations of CXCL8 (interleukin‐8), CCL4 (MIP‐1β) and CCL5 (RANTES) were significantly higher in the peripheral blood of HNSCC patients than in that of controls. In vitro, HNSCC‐conditioned medium inhibited apoptosis of PMNs, increased chemokinesis and chemotaxis of PMNs, induced release of lactoferrin and matrix metalloproteinase 9 by PMNs and enhanced the secretion of CCL4 by PMN. Our findings demonstrate alterations in PMN biology in HNSCC patients. In vitro, tumor‐derived factors modulate cellular functions of PMNs and increase their inflammatory activity. Thus, the interaction between HNSCC and PMNs may contribute to host‐mediated changes in the tumor microenvironment.

EBV transformation overrides gene expression patterns of B cell differentiation stages.

EBV-associated Hodgkin lymphoma (HL) and some post-transplant lymphoproliferative disease (PTLD) cases originate from pro-apoptotic germinal center (GC) B cells that have acquired destructive somatic Ig V gene mutations and were presumably rescued from apoptosis by EBV. To find out whether B cell receptor-crippled GC B cells acquire features of HL and/or PTLD cells upon EBV-infection and to reveal the impact of EBV on expression of B cell differentiation markers, we compared lymphoblastoid cell lines (LCLs) from GC B cells (including BCR-crippled GC-LCLs) to monoclonal LCLs from naïve B cells (N-LCLs). In addition, we analyzed the controversially discussed effect of EBV-infection on the GC B-cell-specific process of somatic hypermutation in vitro. Irrespective of their cellular origin, LCLs expressed CD20, CD30, CD38, AID, Pu.1, and with one exception Syk, but lacked expression of the GC B cell marker BCL-6. Interestingly, the T cell transcription factor GATA-3 that is aberrantly expressed in HL was induced in most GC-LCLs and the memory B cell marker CD27 was activated in N-LCLs. Remarkably, only 4 of 24 GC-LCLs showed significant somatic hypermutation activity, demonstrating that EBV usually silences hypermutation upon infection of GC B cells. Notably, one of three N-LCL showed a low level of intraclonal diversification. Thus, EBV-infection deregulates multiple differentiation factors and processes in B cells, leading to a largely homogenous phenotype of EBV-infected B cells in latency III.

Post-tonsillectomy haemorrhage : A retrospective comparison of abscess- and elective tonsillectomy

Conclusion. There is no increased risk of postoperative haemorrhage for abscess tonsillectomies in comparison to elective tonsillectomies. Objective. There is still controversy as regards the optimal management of peritonsillar abscess. Opponents of tonsillectomy à chaud cite an increased postoperative bleeding risk. Most authors who compared the risks of postoperative haemorrhage after tonsillectomy à chaud and tonsillectomy à froid did not take into consideration criteria such as the age and gender of the patients or the experience of the surgeon. We aimed to eliminate this bias by performing a retrospective study in which a large series of abscess tonsillectomies were compared with an age- and gender-matched group of elective tonsillectomies. Material and methods. All patients had been operated on at the Department of Otorhinolaryngology, University of Duisburg-Essen between March 1994 and August 2000. There were 350 patients in the abscess tonsillectomy group (61% male, 39% female; mean age 31.8 years; range 3–88 years) and 311 in the elective tonsillectomy comparison group (61% male, 39% female; mean age 30.0 years; range 2–83 years). Results. In the abscess tonsillectomy group, 9 patients (2.6%; confidence level 1.1–4.8%) had postoperative haemorrhages which required treatment under general anaesthesia, compared to 17 (5.5%; confidence level 3.2–8.6%) in the age- and gender-matched group of “selected” elective tonsillectomies. The difference between these two rates was not significant (p=0.056). The fairly high rate of haemorrhages in the elective tonsillectomy group was mainly due to the effect of the age-matching procedure, which excluded a considerable number of usually unproblematic tonsillectomies for tonsillar hyperplasia in young children. Moreover, our results show that there is a learning curve for surgeons performing tonsillectomies with regard to postoperative haemorrhages.

Toll-like receptor 4 single-nucleotide polymorphisms Asp299Gly and Thr399Ile in head and neck squamous cell carcinomas.

BackgroundChronic inflammation plays an important role in head and neck squamous cell carcinomas (HNSCC). This study addresses the impact of two single nucleotide polymorphisms (SNP) Asp299Gly and Thr399Ile of the toll-like receptor (TLR) 4 gene on the clinical outcome while accounting for the influence of adjuvant systemic therapy in a large cohort of HNSCC patients.MethodsGenotype analysis was done using DNA from tissue samples from 188 patients with HNSCC; TLR4 protein expression was assessed immunohistochemically in tissue microarrays. Classical survival models were used for statistical analyses.ResultsTen percent of patients with HNSCC presented with the TLR4 299Gly and 17% with the TLR4 399Ile allele. Patients with the heterozygous genotype TLR4 Asp299Gly had a significantly reduced disease-free and overall survival. Also, patients with the heterozygous genotype TLR4 Thr399Ile had a reduced disease-free survival. Notably, these associations seem to be attributable to relatively poor therapy response as e.g. reflected in a significantly shorter DFS among HNSCC patients carrying the Asp299Gly variant and receiving adjuvant systemic therapy.ConclusionAccording to this study, TLR4 299Gly und 399Ile alleles may serve as markers for prognosis of head and neck cancer in patients with adjuvant systemic therapy, particularly chemotherapy, and might indicate therapy resistance.

Detection of Circulating Tumor Cell Subpopulations in Patients with Head and Neck Squamous Cell Carcinoma (HNSCC)

Background Since image based diagnostic tools fail to detect early metastasis in head and neck squamous cell carcinoma (HNSCC) it is crucial to develop minimal invasive diagnostic methods. A promising approach is to identify and characterize circulating tumor cells (CTC) in the peripheral blood of HNSCC patients. In this pilot study, we assessed which non-hematopoietic cell types are identifiable and whether their numbers differ in pre- and postoperative blood samples. Methods 20 ml citrated peripheral blood was taken from 10 HNSCC patients before and after curative resection. CTC were enriched using density gradient centrifugation. CTC presence was verified by multi-immunofluorescence staining against cytokeratin (CK; epithelial), N-cadherin (mesenchymal); CD133 (stem-cell), CD45 (hematopoietic) and DAPI (nucleus). Individual cell type profiles were analyzed. Results We were able to detect cells with epithelial properties like CK+/N-cadherin−/CD45− and CK+/CD133−/CD45− as well as cells with mesenchymal features such as N-cadherin+/CK−/CD45− and cells with both characteristics like N-cadherin+/CK+/CD45−. We also observed cells showing stem cell-like features like CD133+/CK−/CD45− and cells with both epithelial and stem cell-like features such as CD133+/CK+/CD45−. The number of CK positive cells (p = 0.002), N-cadherin positive cells (p = 0.002) and CD133 positive cells (p = 0.01) decreased significantly after resection. Kaplan-Meier test showed that the survival was significantly shorter when N-cadherin+ cells were present after resection (p = 0.04; 474 vs. 235 days; [HR] = 3.1). Conclusions This is - to the best of our knowledge- the first pilot study identifying different CTC populations in peripheral blood of HNSCC patients and showing that these individual cell type profiles may have distinct clinical implications.

The role of tumour FoxP3 as prognostic marker in different subtypes of head and neck cancer

Expression of the forkhead transcription factor (FoxP3)--an established marker of regulatory T cells--has been found in other cell types as well, including tumour cells. Recent studies indicated that high tumour FoxP3 expression might be associated with a poor outcome of patients with several types of solid cancers. Here, we investigated the role of FoxP3 expressed by the tumour cells in the prognosis of larynx and oro-hypopharynx squamous cell carcinoma (LSCC and OHSCC)--two major subtypes of head and neck cancer. To this end, we analysed by immunohistochemistry the expression of tumour FoxP3 in tissues from 83 LSCC and 89 OHSCC patients in relation to overall survival. In multivariate analysis we found that high tumour FoxP3 expression significantly associated with poor survival in OHSCC but not in LSCC patients. In further studies, we combined the prognostic value of FoxP3 with selected markers of inflammation (cyclooxygenase-2; COX2) or with markers of enhanced tumour migration/invasion (AHNAK and CORTACTIN). Interestingly, we found that the combination of FoxP3 and AHNAK (in LSCC) or FoxP3 and CORTACTIN (in OHSCC) had significantly stronger prognostic values than either marker analysed individually. Combination of FoxP3 and COX2 enhanced the prognostic accuracy only in OHSCC. Thus, our study identifies novel individual and combination markers that might have enhanced and distinct prognostic relevance in different subtypes of head and neck cancer.

In vitro chemosensitivity of head and neck cancer cell lines

BackgroundSystemic 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.MethodsTen 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.ResultsAll 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.ConclusionChemosensitivity 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.

Acute Arterial Hemorrhage Following Radiotherapy of Oropharyngeal Squamous Cell Carcinoma

Background and Purpose:Vascular erosion is a rare but life-threatening complication after radiotherapy. The authors report on acute arterial bleeding and its therapy following radiotherapy of oropharyngeal tumors.Patients and Methods:Ten patients with oropharyngeal squamous cell carcinoma of any stage developed foudroyant acute arterial hemorrhage 3–46 months (14.4 ± 5.1 months) after primary (5/10) or adjuvant radio(chemo)therapy (R[C]T).Results:All patients had a history of recurrent minor bleeding episodes and showed deep mucosal ulcerations also outside the primary tumor region. A life-threatening arterial hemorrhage appeared in the area of these mucosal defects in the pharyngeal region. Affected vessels were the common carotid artery as well as the internal and the external portion with branches like the ascending pharyngeal and superior thyroid arteries. Treatment consisted of emergency intubation or tracheotomy followed by exposure and package of the pharynx and surgical ligature and/or embolization. 6/10 patients (all hospitalized) survived the episode, however, lethal outcome in 4/10 patients (outpatients) was related to asphyxia as a result of blood aspiration or exsanguination. None of the patients revealed evidence of persistent or recurrent tumor disease as proven by biopsy/autopsy and imaging technique.Conclusion:Vascular erosion following primary or adjuvant R(C)T represents a rare and potentially life-threatening complication requiring immediate emergency treatment involving head and neck surgeons, anesthesiologists and neuroradiologists. For patients with oropharyngeal neoplasms treated by R(C)T and showing recurrent bleeding episodes and mucosal ulceration particularly after the acute treatment phase, hospitalization with prophylactic surgical ligature or embolization of affected arteries is recommended.ZusammenfassungHintergrund und Ziel:Schädigungen von Blutgefäßen stellen eine seltene, jedoch potentiell lebensbedrohliche Komplikation nach Strahlentherapie dar. Die Autoren berichten über akute arterielle Blutungen und deren Therapie nach Strahlentherapie von oropharyngealen Tumoren.Patienten und Methodik:Zehn Patienten mit einem oropharyngealen Plattenepithelkarzinom jeglichen Tumorstadiums entwickelten 3–46 Monate (14,4 ± 5,1 Monate) nach primärer (5/10) oder adjuvanter Radio(chemo)therapie (R[C]T) eine akute arterielle Blutung (Tabelle 1).Ergebnisse:Alle Patienten zeigten in ihrer Vorgeschichte kleinere Blutungsepisoden und litten unter tiefen Schleimhautulzerationen im Pharynx auch außerhalb der Primärtumorregion (Abbildung 1). Die lebensbedrohlichen Blutungen traten im Bereich dieser Schleimhautdefekte auf. Betroffene Gefäße waren die Arteriae carotis communis, carotis interna und carotis externa mit ihren Ästen, Arteria pharyngea ascendens und Arteria thyroidea superior. Die Therapie umfasste die notfallmäßige Intubation oder Tracheotomie und Tamponade des Rachens, gefolgt entweder von einer Ligatur (Abbildung 2) und/oder Embolisation des betroffenen Gefäßes. 6/10 Patienten überlebten dieses Ereignis, 4/10 Patienten verstarben vor Erreichen der Klinik infolge des hohen Blutverlusts oder Aspiration. Keiner der Patienten zeigte zum Zeitpunkt der Blutung bildmorphologisch oder histologisch einen Residual- oder Rezidivtumor.Schlussfolgerung:Gefäßschädigungen infolge primärer oder adjuvanter R(C)T stellen eine potentiell lebensbedrohliche Komplikation dar, die einer unmittelbaren notfallmäßigen interdisziplinären Behandlung durch Kopf-Hals-Chirurgen, Anästhesisten und Neuroradiologen bedarf. Bei Patienten mit oropharyngealen Karzinomen, die nach einer R(C)T unter wiederkehrenden pharyngealen Blutungsepisoden und Schleimhautulzerationen leiden, sollte eine stationäre Überwachung mit einer eventuellen prophylaktischen Unterbindung oder einem neuroradiologisch-interventionellen Verschluss (Abbildung 3) der betroffenen Gefäße in Erwägung gezogen werden.

Bilateral peritonsillar abscesses

In the discussion about the management of the peritonsillar abscess (PTA) in regard to the pros and cons of tonsillectomy à chaud versus à froid, the risk of obscure contralateral abscesses is often neglected. To the authors’ knowledge, there are only a few series of PTA being analyzed for the abscess rate of bilateral PTA. A group of 541 abscess tonsillectomies was retrospectively analyzed for the presence of a bilateral manifestation of peritonsillar abscess. Twenty-one patients (3.88%) had bilateral abscesses. None of these had been detected prior to the operation. Of the 541 patients, 2.22% had postoperative hemorrhages that had to be arrested under general anesthesia. Within the discussion about abscess tonsillectomy versus stab incision followed by interval tonsillectomy (à froid), the rate of almost 4% bilateral abscesses should be taken into consideration as dangerous complications such as mediastinitis could develop from the remaining abscess formation of the contralateral side.

Value of DSA in the diagnostic workup of pulsatile tinnitus.

Objectives Pulsatile tinnitus (PT) is a rare complaint, but can be a symptom of life-threatening disease. It is often caused by vascular pathologies, e.g. dural arteriovenous fistula (dAVF), arteriovenous malformation (AVM) or vascularized tumors. The current diagnostic pathway includes clinical examination, cranial MRI and additional DSA. The aim of this study was to evaluate the diagnostic impact of DSA in the diagnostic workup of patients with PT in comparison to MRI alone. Methods Retrospectively, 54 consecutive patients with pulsatile tinnitus were evaluated. All patients had a diagnostic workup including cranial MRI and DSA. MRI examinations were blinded to the results of DSA and retrospectively analyzed in consensus by two experienced neuroradiologists. The MR-examinations were evaluated for each performed sequence separately: time-of-flight-angiography, ce-MRA, T2, ce-T1-sequence and ce-T1-sequence with fat saturation. Results 37 of the 54 patients revealed a pathology explaining PT on MRI, which was detected by the readers in 100% and proofed by means of DSA. 24 dAVF, four paraganglioma, two AVM and seven more pathologies were described. All patients without pathology on MRI did also not show any pathology in DSA. Conclusions MR imaging is sufficient to exclude pathology in patients with pulsatile tinnitus.