Zdeněk Čada

Head and neck squamous cancer stromal fibroblasts produce growth factors influencing phenotype of normal human keratinocytes

Epithelial–mesenchymal interaction between stromal fibroblasts and cancer cells influences the functional properties of tumor epithelium, including the tumor progression and spread. We compared fibroblasts prepared from stroma of squamous cell carcinoma and normal dermal fibroblasts concerning their biological activity toward normal keratinocytes assessed by immunocytochemistry and profiling of gene activation for growth factors/cytokines by microarray chip technology. IGF-2 and BMP-4 were determined as candidate factors responsible for tumor-associated fibroblast activity that influences normal epithelia. This effect was confirmed by addition of recombinant IGF-2 and BMP4, respectively, to the culture medium. This hypothesis was also verified by inhibition experiments where blocking antibodies were employed in the medium conditioned by cancer-associated fibroblast. Presence of these growth factors was also detected in tumor samples.

Upregulation of IL‐6, IL‐8 and CXCL‐1 production in dermal fibroblasts by normal/malignant epithelial cells in vitro: Immunohistochemical and transcriptomic analyses

Considering an analogy between wound healing and tumour progression, we studied chemokine and cytokine transcription and expression in normal fibroblasts by co‐culture and in situ.

Smooth muscle actin-expressing stromal fibroblasts in head and neck squamous cell carcinoma: Increased expression of galectin-1 and induction of poor prognosis factors

Tumor stroma is an active part influencing the biological properties of malignancies via molecular cross‐talk. Cancer‐associated fibroblasts play a significant role in this interaction. These cells frequently express smooth muscle actin and can be classified as myofibroblasts. The adhesion/growth‐regulatory lectin galectin‐1 is an effector for their generation. In our study, we set the presence of smooth muscle actin‐positive cancer‐associated fibroblasts in relation to this endogenous lectin and an in vivo competitor (galectin‐3). In squamous cell carcinomas of head and neck, upregulation of galectin‐1 presence was highly significantly correlated to presence of smooth muscle actin‐positive cancer‐associated fibroblasts in the tumor (p = 4 × 10−8). To pinpoint further correlations on the molecular level, we applied microarray analyses to the transcription profiles of the corresponding tumors. Significant correlations of several transcripts were detected with the protein level of galectin‐1 in the cancer‐associated fibroblasts. These activated genes (MAP3K2, TRIM23, PTPLAD1, FUSIP1, SLC25A40 and SPIN1) are related to known squamous‐cell‐carcinoma poor‐prognosis factors, NF‐κB upregulation and splicing downregulation. These results provide new insights into the significance of presence of myofibroblasts in squamous cell carcinoma.

Phenotypic characterization of human keratinocytes in coculture reveals differential effects of fibroblasts from benign fibrous histiocytoma (dermatofibroma) as compared to cells from its malignant form and to normal fibroblasts.

BACKGROUND Benign and malignant fibrous histiocytoma present with a considerable difference concerning cellular organization in their vicinity. OBJECTIVE Normally appearing epithelium covers the malignant form in contrast to hyperplastic epidermis for benign tumors. It is an open question as to whether the tumor-associated fibroblasts are capable to affect phenotypic features of normal keratinocytes, prompting this comparative analysis. METHODS Fibroblasts were isolated from benign and malignant fibrous histiocytomas, respectively, and also from normal dermis. The resulting cell populations were thoroughly characterized immunocytochemically using a large panel of antibodies. The three fibroblast preparations were cocultured with normal interfollicular keratinocytes. Their phenotype was characterized for distinct properties including differentiation and proliferation. RESULTS Fibroblasts prepared from both tumor types were phenotypically practically identical with normal dermal fibroblasts. Their activities on keratinocytes were different. Cells prepared from benign fibrous histiocytoma were capable to effect strong expression of keratin 19 and production of a galectin-1-rich extracellular matrix. Fibroblasts isolated from malignant fibrous histiocytoma led to a phenotype very similar to that when keratinocytes were cocultured with normal dermal fibroblasts. CONCLUSION Fibroblasts prepared from benign fibrous histiocytoma were biologically active on keratinocytes in a particular manner. Our results on fibroblast activity are suggested to be relevant for morphologic differences observed in vivo between normal epidermis and epidermis adjacent to the studied tumor types.

Vertigo Perception and Quality of Life in Patients after Surgical Treatment of Vestibular Schwannoma with Pretreatment Prehabituation by Chemical Vestibular Ablation.

Surgical removal of vestibular schwannoma causes acute vestibular symptoms, including postoperative vertigo and oscillopsia due to nystagmus. In general, the dominant symptom postoperatively is vertigo. Preoperative chemical vestibular ablation can reduce vestibular symptoms postoperatively. We used 1.0 mL of 40 mg/mL nonbuffered gentamicin in three intratympanic installations over 2 days, 2 months preoperatively in 10 patients. Reduction of vestibular function was measured by the head impulse test and the caloric test. Reduction of vestibular function was found in all gentamicin patient groups. After gentamicin vestibular ablation, patients underwent home vestibular exercising for two months. The control group consisted of 10 patients who underwent only home vestibular training two months preoperatively. Postoperative rates of recovery and vertigo in both groups were evaluated with the Glasgow Benefit Inventory (GBI), the Glasgow Health Status Inventory (GHSI), and the Dizziness Handicap Inventory questionnaires, as well as survey of visual symptoms by specific questionnaire developed by us. There were no statistically significant differences between both groups with regard to the results of questionnaires. Patients who received preoperative gentamicin were more resilient to optokinetic and optic flow stimulation (p < 0.05). This trial is registered with clinical study registration number NCT02963896.

Microarray Analysis of Serum mRNA in Patients with Head and Neck Squamous Cell Carcinoma at Whole-Genome Scale

With the increasing demand for noninvasive approaches in monitoring head and neck cancer, circulating nucleic acids have been shown to be a promising tool. We focused on the global transcriptome of serum samples of head and neck squamous cell carcinoma (HNSCC) patients in comparison with healthy individuals. We compared gene expression patterns of 36 samples. Twenty-four participants including 16 HNSCC patients (from 12 patients we obtained blood samples 1 year posttreatment) and 8 control subjects were recruited. The Illumina HumanWG-6 v3 Expression BeadChip was used to profile and identify the differences in serum mRNA transcriptomes. We found 159 genes to be significantly changed (Storeys P value <0.05) between normal and cancer serum specimens regardless of factors including p53 and B-cell lymphoma family members (Bcl-2, Bcl-XL). In contrast, there was no difference in gene expression between samples obtained before and after surgery in cancer patients. We suggest that microarray analysis of serum cRNA in patients with HNSCC should be suitable for refinement of early stage diagnosis of disease that can be important for development of new personalized strategies in diagnosis and treatment of tumours but is not suitable for monitoring further development of disease.

Inter-individual Variability in Processing of the Sémont Liberatory Manoeuvre

Aim: Sémont liberatory manoeuvre is used in the treatment of posterior semicircular canal in patients with benign paroxysmal positional vertigo. The aim of the study was to determine the interindividual and intra-individual variability in the processing of the manoeuvre and to defi ne the parameters that aff ect it the most. Methods: Three experienced therapists applied the manoeuvre bilaterally in 10 healthy probands. Inertial measurement unit consisting of the accelerometer and gyroscope recorded the trajectory and speed. Angle changes were placed in the coordinate reference system. Accuracy of the manoeuvre was evaluated based on the deviations from the projected sensory plane. Results: Parameters aff ecting the accuracy of the manoeuvre included height of a proband (p = 0.0252), phase of the movement (p < 0.0001), therapist and the side of the movement. The eff ect of these factors is the most pronounced when combined. We assessed interactions of the phase of the movement and the height of the probands (p = 0.0130), the therapist and the phase of movement (p = 0.0001), the therapist and the height of the probands (p < 0.0252). The largest magnitude of deviation from the sensory plane was in the range of –37.17° to 31° with a standard deviation of 16.6°. Conclusions: The data highlight high variability in the implementation of the Sémont liberatory manoeuvre, whether measurements by a single therapist or interindividually between the therapists are compared. Analysis of the impact of these deviations on therapeutic effi cacy in patients with benign paroxysmal positional vertigo and detailed analysis of infl uencing parameters should be the subject of further research. Autoři deklarují, že v souvislosti s předmětem studie nemají žádné komerční zájmy. The authors declare they have no potential confl icts of interest concerning drugs, products, or services used in the study. Redakční rada potvrzuje, že rukopis práce splnil ICMJE kritéria pro publikace zasílané do biomedicínských časopisů. The Editorial Board declares that the manuscript met the ICMJE “uniform requirements” for biomedical papers. M. Stehlíková1, O. Čakrt1, I. Bodlák2, Z. Čada3, R. Černý4, J. Jeřábek4 1 Klinika rehabilitace a tělovýchovného lékařství 2. LF UK a FN v Motole, Praha 2 Princip a.s., Praha 3 Klinika otorinolaryngologie a chirurgie hlavy a krku 1. LF UK a FN v Motole, Praha 4 Neurologická klinika 2. LF UK a FN v Motole, Praha Mgr. Mariana Stehlíková Klinika rehabilitace a tělovýchovného lékařství 2. LF UK a FN Motol V Úvalu 84 150 06 Praha 5 e-mail: mariana.stehlikova@fnmotol.cz Přijato k recenzi: 19. 8. 2014 Přijato do tisku: 13. 10. 2014