T2-FLAIR Mismatch: An Imaging Biomarker for Children's MYB/MYBL1-Altered Diffuse Astrocytoma or Angiocentric Glioma.

BACKGROUND AND PURPOSE: T2-FLAIR mismatch is a highly specific imaging biomarker of IDH-mutant diffuse astrocytoma in adults. It has however also been described in MYB/MYBL1-altered low grade tumors. Our aim was to assess the diagnostic power of the T2-FLAIR mismatch in IDH-mutant astrocytoma and MYB/MYBL1-altered low-grade tumors in children and correlate this mismatch with histology. MATERIALS AND METHODS: We evaluated MR imaging examinations of all pediatric patients, performed at the Princess Máxima Center for Pediatric Oncology and the University Medical Center Utrecht between January 2012 and January 2023, with the histomolecular diagnosis of IDH-mutant astrocytoma, diffuse astrocytoma MYB/MYBL1-altered, or angiocentric glioma, and the presence of T2-FLAIR mismatch was assessed. Histologically, the presence of microcysts in the tumor (a phenomenon suggested to be correlated with T2-FLAIR mismatch in IDH-mutant astrocytomas in adults) was evaluated. RESULTS: Nineteen pediatric patients were diagnosed with either IDH-mutant astrocytoma (n = 8) or MYB/MYBL1-altered tumor (n = 11: diffuse astrocytoma, MYB- or MYBL1-altered n = 8; or angiocentric glioma n = 3). T2-FLAIR mismatch was present in 11 patients, 3 (38%) in the IDH-mutant group and 8 (73%) in the MYB/MYBL1 group. No correlation was found between T2-FLAIR mismatch and the presence of microcysts or an enlarged intercellular space in either IDH-mutant astrocytoma (P = .38 and P = .56, respectively) or MYB/MYBL1-altered tumors (P = .36 and P = .90, respectively). CONCLUSIONS: In our pediatric population, T2-FLAIR mismatch was more often found in MYB/MYBL1-altered tumors than in IDH-mutant astrocytomas. In contrast to what has been reported for IDH-mutant astrocytomas in adults, no correlation was found with microcystic changes in the tumor tissue. This finding challenges the hypothesis that such microcystic changes and/or enlarged intercellular spaces in the tissue of these tumors are an important part of explaining the occurrence of the T2-FLAIR mismatch.

IDEAL monitoring of musical skills during awake craniotomy: From step 1 to step 2.

The aim of awake brain surgery is to perform a maximum resection on the one hand, and to preserve cognitive functions, quality of life and personal autonomy on the other hand. Historically, language and sensorimotor functions were most frequently monitored. Over the years other cognitive functions, including music, have entered the operation theatre. Cases about monitoring musical abilities during awake brain surgery are emerging, and a systematic method how to monitor music would be the next step. According to the IDEAL framework for surgical innovations our study aims to present future recommendation based on a systematic literature search (PRISMA) in combination with lessons learned from three case reports from our own clinical practice with professional musicians (n = 3). We plead for structured procedures including individual tailored tasks. By embracing these recommendations, we can both improve clinical care and unravel music functions in the brain.

Awake craniotomy does not lead to increased psychological complaints.

BACKGROUND: Patients with brain tumours are increasingly treated by using the awake craniotomy technique. Some patients may experience anxiety when subjected to brain surgery while being fully conscious. However, there has been only limited research into the extent to which such surgeries actually result in anxiety or other psychological complaints. Previous research suggests that undergoing awake craniotomy surgery does not lead to psychological complaints, and that post-traumatic stress disorders (PTSD) are uncommon following this type of surgery. It must be noted, however, that many of these studies used small random samples. METHOD: In the current study, 62 adult patients completed questionnaires to identify the degree to which they experienced anxiety, depressive and post-traumatic stress complaints following awake craniotomy using an awake-awake-awake procedure. All patients were cognitively monitored and received coaching by a clinical neuropsychologist during the surgery. RESULTS: In our sample, 21% of the patients reported pre-operative anxiety. Four weeks after surgery, 19% of the patients reported such complaints, and 24% of the patients reported anxiety complaints after 3 months. Depressive complaints were present in 17% (pre-operative), 15% (4 weeks post-operative) and 24% (3 months post-operative) of the patients. Although there were some intra-individual changes (improvement or deterioration) in the psychological complaints over time, on group-level postoperative levels of psychological complaints were not increased relative to the preoperative level of complaints. The severity of post-operative PTSD-related complaints were rarely suggestive of a PTSD. Moreover, these complaints were seldom attributed to the surgery itself, but appeared to be more related to the discovery of the tumour and the postoperative neuropathological diagnosis. CONCLUSIONS: The results of the present study do not indicate that undergoing awake craniotomy is associated with increased psychological complaints. Nevertheless, psychological complaints may well exist as a result of other factors. Consequently, monitoring the patient's mental wellbeing and offering psychological support where necessary remain important.

A timeline of cognitive functioning in glioma patients who undergo awake brain tumor surgery.

BACKGROUND: The purpose of awake brain tumor surgery is to maximize the resection of the tumor and to minimize the risk of neurological and cognitive impairments. The aim of this study is to gain understanding of the development of possible postoperative cognitive deficits after awake brain tumor surgery in patients with suspected gliomas, by comparing preoperative, early postoperative, and late postoperative functioning. A more detailed timeline will be helpful in informing candidates for surgery about what to expect regarding their cognitive functioning. METHODS: Thirty-seven patients were included in this study. Cognitive functioning was measured by means of a broad cognitive screener preoperatively, days after surgery and months after surgery in patients who underwent awake brain tumor surgery with cognitive monitoring. The cognitive screener included tests for object naming, reading, attention span, working memory, inhibition, inhibition/switching, and visuoperception. We performed a Friedman ANOVA to analyze on group level. RESULTS: Overall, no significant differences were found between preoperative cognitive functioning, early postoperative cognitive functioning, and late postoperative cognitive functioning, except for performances on the inhibition task. Directly after surgery, patients were significantly slower on this task. However, in the following months after surgery, they returned to their preoperative level. CONCLUSION: The timeline of cognitive functioning after awake tumor surgery appeared overall stable in the early and late postoperative phase, except for inhibition, which is more difficult in the first days after awake brain tumor surgery. This more detailed timeline of cognitive functioning, in combination with future research, can possibly be contributing in informing patients and caregivers what to expect after awake brain tumor surgery.

The impact of etiology in lesion-symptom mapping - A direct comparison between tumor and stroke.

INTRODUCTION: Lesion-symptom mapping is a key tool in understanding the relationship between brain structures and behavior. However, the behavioral consequences of lesions from different etiologies may vary because of how they affect brain tissue and how they are distributed. The inclusion of different etiologies would increase the statistical power but has been critically debated. Meanwhile, findings from lesion studies are a valuable resource for clinicians and used across different etiologies. Therefore, the main objective of the present study was to directly compare lesion-symptom maps for memory and language functions from two populations, a tumor versus a stroke population. METHODS: Data from two different studies were combined. Both the brain tumor (N = 196) and stroke (N = 147) patient populations underwent neuropsychological testing and an MRI, pre-operatively for the tumor population and within three months after stroke. For this study, we selected two internationally widely used standardized cognitive tasks, the Rey Auditory Verbal Learning Test and the Verbal Fluency Test. We used a state-of-the-art machine learning-based, multivariate voxel-wise approach to produce lesion-symptom maps for these cognitive tasks for both populations separately and combined. RESULTS: Our lesion-symptom mapping results for the separate patient populations largely followed the expected neuroanatomical pattern based on previous literature. Substantial differences in lesion distribution hindered direct comparison. Still, in brain areas with adequate coverage in both groups, considerable LSM differences between the two populations were present for both memory and fluency tasks. Post-hoc analyses of these locations confirmed that the cognitive consequences of focal brain damage varied between etiologies. CONCLUSION: The differences in the lesion-symptom maps between the stroke and tumor population could partly be explained by differences in lesion volume and topography. Despite these methodological limitations, both the lesion-symptom mapping results and the post-hoc analyses confirmed that etiology matters when investigating the cognitive consequences of lesions with lesion-symptom mapping. Therefore, caution is advised with generalizing lesion-symptom results across etiologies.

MR Imaging and Clinical Characteristics of Diffuse Glioneuronal Tumor with Oligodendroglioma-like Features and Nuclear Clusters.

BACKGROUND AND PURPOSE: Diffuse glioneuronal tumor with oligodendroglioma-like features and nuclear clusters (DGONC) is a new, molecularly defined glioneuronal CNS tumor type. The objective of the present study was to describe MR imaging and clinical characteristics of patients with DGONC. MATERIALS AND METHODS: Preoperative MR images of 9 patients with DGONC (median age at diagnosis, 9.9 years; range, 4.2-21.8 years) were reviewed. RESULTS: All tumors were located superficially in the frontal/temporal lobes and sharply delineated, displaying little mass effect. Near the circle of Willis, the tumors encompassed the arteries. All except one demonstrated characteristics of low-to-intermediate aggressiveness with high-to-intermediate T2WI and ADC signals and bone remodeling. Most tumors (n = 7) showed a homogeneous ground-glass aspect on T2-weighted and FLAIR images. On the basis of the original histopathologic diagnosis, 6 patients received postsurgical chemo-/radiotherapy, 2 were irradiated after surgery, and 1 patient underwent tumor resection only. At a median follow-up of 61 months (range, 10-154 months), 6 patients were alive in a first complete remission and 2 with stable disease 10 and 21 months after diagnosis. The only patient with progressive disease was lost to follow-up. Five-year overall and event-free survival was 100% and 86±13%, respectively. CONCLUSIONS: This case series presents radiomorphologic characteristics highly predictive of DGONC that contrast with the typical aspects of the original histopathologic diagnoses. This presentation underlines the definition of DGONC as a separate entity, from a clinical perspective. Complete resection may be favorable for long-term disease control in patients with DGONC. The efficacy of nonsurgical treatment modalities should be evaluated in larger series.

Association between cerebral perfusion and paediatric postoperative cerebellar mutism syndrome after posterior fossa surgery-a systematic review.

BACKGROUND: Paediatric postoperative cerebellar mutism syndrome (ppCMS) is a common complication following the resection of a cerebellar tumour in children. It is hypothesized that loss of integrity of the cerebellar output tracts results in a cerebello-cerebral "diaschisis" and reduced function of supratentorial areas of the brain. METHODS: We performed a systematic review of the literature according to the PRISMA guidelines, in order to evaluate the evidence for hypoperfusion or hypofunction in the cerebral hemispheres in patients with ppCMS. Articles were selected based on the predefined eligibility criteria and quality assessment. RESULTS: Five studies were included, consisting of three prospective cohort studies, one retrospective cohort study and one retrospective case control study. Arterial spin labelling (ASL) perfusion MRI, dynamic susceptibility contrast (DSC) perfusion MRI and single photon emission computed tomography (SPECT) were used to measure the cerebral and cerebellar tissue perfusion or metabolic activity. Reduced cerebral perfusion was predominantly demonstrated in the frontal lobe. CONCLUSIONS: This systematic review shows that, after posterior fossa tumour resection, cerebral perfusion is reduced in ppCMS patients compared to patients without ppCMS. Well-powered prospective studies, including preoperative imaging, are needed to ascertain the cause and role of hypoperfusion in the pathophysiology of the syndrome.

Tinnitus treatment by vagus nerve stimulation: A systematic review.

BACKGROUND: Tinnitus is a phantom sensation of sound, which can have a negative impact on quality of life of those affected. No curative treatments are currently known. Neuromodulation by vagus nerve stimulation has emerged as a new treatment option for tinnitus, though till date the effectiveness remains unclear. Therefore, we aim to review the effect of vagus nerve stimulation on tinnitus distress and tinnitus symptom severity in patients with chronic tinnitus. METHODS: We searched Pubmed, Embase and the Cochrane Library systematically for RCTs, observational studies and case studies on the effect of VNS treatment for tinnitus on October 29, 2019. Studies including adult patients with subjective tinnitus, comparing transcutaneous or implantable VNS to placebo or no treatment or before and after application of VNS treatment on tinnitus distress and tinnitus symptom severity measured with a validated questionnaire were eligible. The risk of bias was assessed with the appropriate tool for each type of study. RESULTS: Our search identified 9 primary studies of which 2 RCTs, 5 cohort studies and 2 case series or reports. 5 studies used transcutaneous VNS treatment and 4 used implanted VNS treatment. 6 studies combined VNS treatment with sound therapy. There was a serious risk of bias in all studies, especially on confounding. Most studies reported a small decrease in tinnitus distress or tinnitus symptom severity. CONCLUSION: Due to methodological limitations and low reporting quality of the included studies, the effect of VNS on tinnitus remains unclear. To draw conclusions for which patient population and to what extent (t)VNS is beneficial in the treatment of tinnitus, a randomised controlled trial should be considered.

PI3 kinase mutations and mutational load as poor prognostic markers in diffuse glioma patients.

INTRODUCTION: Recent advances in molecular diagnostics allow diffuse gliomas to be classified based on their genetic changes into distinct prognostic subtypes. However, a systematic analysis of all molecular markers has thus far not been performed; most classification schemes use a predefined and select set of genes/molecular markers. Here, we have analysed the TCGA dataset (combined glioblastoma (GBM) and lower grade glioma (LGG) datasets) to identify all prognostic genetic markers in diffuse gliomas in order to generate a comprehensive classification scheme. RESULTS: Of the molecular markers investigated (all genes mutated at a population frequency >1.7 % and frequent chromosomal imbalances) in the entire glioma dataset, 57 were significantly associated with overall survival. Of these, IDH1 or IDH2 mutations are associated with lowest hazard ratio, which confirms IDH as the most important prognostic marker in diffuse gliomas. Subsequent subgroup analysis largely confirms many of the currently used molecular classification schemes for diffuse gliomas (ATRX or TP53 mutations, 1p19q codeletion). Our analysis also identified PI3-kinase mutations as markers of poor prognosis in IDH-mutated + ATRX/TP53 mutated diffuse gliomas, median survival 3.7 v. 6.3 years (P = 0.02, Hazard rate (HR) 2.93, 95 % confidence interval (CI) 1.16 - 7.38). PI3-kinase mutations were also prognostic in two independent datasets. In our analysis, no additional molecular markers were identified that further refine the molecular classification of diffuse gliomas. Interestingly, these molecular classifiers do not fully explain the variability in survival observed for diffuse glioma patients. We demonstrate that tumor grade remains an important prognostic factor for overall survival in diffuse gliomas, even within molecular glioma subtypes. Tumor grade was correlated with the mutational load (the number of non-silent mutations) of the tumor: grade II diffuse gliomas harbour fewer genetic changes than grade III or IV, even within defined molecular subtypes (e.g. ATRX mutated diffuse gliomas). CONCLUSION: We have identified PI3K mutations as novel prognostic markers in gliomas. We also demonstrate that the mutational load is associated with tumor grade. The increase in mutational load may partially explain the increased aggressiveness of higher grade diffuse gliomas when a subset of the affected genes actively contributes to gliomagenesis and/or progression.

The use of (18)F-FDG PET to differentiate progressive disease from treatment induced necrosis in high grade glioma.

In the follow-up of patients treated for high grade glioma, differentiation between progressive disease (PD) and treatment-induced necrosis (TIN) is challenging. The purpose of this study is to evaluate the diagnostic accuracy of FDG PET for the differentiation between TIN and PD after high grade glioma treatment. We retrospectively identified patients between January 2011 and July 2013 that met the following criteria: age >18; glioma grade 3 or 4; treatment with radiotherapy or chemoradiotherapy; new or progressive enhancement on post treatment MRI; FDG PET within 4 weeks of MRI. Absolute and relative (to contralateral white matter) values of SUVmax and SUVpeak were determined in new enhancing lesions on MRI. The outcome of PD or TIN was determined by neurosurgical biopsy/resection, follow-up MRI, or clinical deterioration. The association between FDG PET and outcome was analyzed with univariate logistic regression and ROC analysis for: all lesions, lesions >10, >15, and >20 mm. We included 30 patients (5 grade 3 and 25 grade 4), with 39 enhancing lesions on MRI. Twenty-nine lesions represented PD and 10 TIN. Absolute and relative values of SUVmax and SUVpeak showed no significant differences between PD and TIN. ROC analysis showed highest AUCs for relative SUVpeak in all lesion sizes. Relative SUVpeak for lesions >20 mm showed reasonable discriminative properties [AUC 0.69 (0.41-0.96)]. FDG PET has reasonable discriminative properties for differentiation of PD from TIN in high grade gliomas larger than 20 mm. Overall diagnostic performance is insufficient to guide clinical decision-making.

Human cytomegalovirus and primary intracranial tumours: frequency of tumour infection and lack of correlation with systemic immune anti-viral responses.

AIMS: Human cytomegalovirus (HCMV) is a ubiquitous beta human herpesvirus able to influence infected cell survival and proliferation and to modulate the host immune response. As there is accumulating evidence that HCMV is detected in primary intracranial astrocytic tumours, in this study we looked for the presence of HCMV in intracranial tumours and tried to correlate this eventual presence with the anti-HCMV systemic immunoreactivity and with the detection of HCMV in peripheral blood. METHODS: In this study, we analysed 43 glioblastomas (GBM), 14 oligodendrogliomas (OL) and 20 meningiomas (MG) by immunofluorescence (IF) targeting HCMV immediate early antigen (IE1) and by nested PCR (nPCR) amplifying HCMV glycoprotein B (gB). RESULTS: Detection of IE1 by IF showed the presence of HCMV in 70% of GBM, 57% of OL and 85% of MG, in contrast to gB nPCR, which detected HCMV in only 50% of GBM, 38% of OL and 46% of MG. Unexpectedly, HCMV DNA and antigens were detected within GBM, OL and MG of patients that exhibit negative viral serology. More surprisingly, PCR on the peripheral blood did not detect HCMV in patients with a HCMV-positive tumour. CONCLUSIONS: Our results are in agreement with previous observations demonstrating HCMV in glial tumours and highlight the presence of HCMV in meningiomas. We also showed that anti-HCMV specific systemic immunoreactivity and detection of HCMV in peripheral blood are not predictive of HCMV presence in primary intracranial tumours.

[Traumatic brain injury and subarachnoid hemorrhage as a cause of hypopituitarism: a review]. / Hypopituitarisme consécutif aux atteintes cérébrales: le traumatisme crânien et l'hémorragie sous-arachnoïdienne mis en cause.

Brain injuries namely traumatic brain injuries (TBI) and subarachnoid haemorrhage (SAH) are relevant causes of acquired adult hypopituitarism, perhaps more prevalent than ever believed. TBI represent a major health problem with an annual incidence of 300 cases per 100.000. SAH affects six new cases per 1.000.000 habitants in USA. In Belgium we estimate nearly 30.000 new TBI cases and 600 SAH cases per year. In the English literature, TBI secondary hypopituitarism has been well documented in 14 retrospective and prospective series accounting for 1.077 cases. In all these series the main pituitary deficits were: GH (14%), ACTH (14%), gonadotrope (18%), TSH (7%) and diabetes insipidus (4%). SAH was documented as a cause of hypopituitarism in three retrospective series accounting for 110 cases and in one prospective series. In all these series main pituitary deficits were GH (25%), ACTH (15%), gonadotrope (8.5%), TSH (6%) and diabetes insipidus (4%). In this review, we analyze recent data and discuss diagnostic and treatment features of secondary hypopituitarism due TBI and SAH.

[Glioblastoma, an example of translational research?]. / Les glioblastomes, un exemple de recherche translationnelle?

Among patients which develop glioblastoma multiform (GBM), recurrence is the rule despite continuous progress in surgery, radiotherapy and chemotherapy. In the adult, GBM is the most frequent and most aggressive tumour of the Central Nervous System. A better understanding of the mechanisms by which these tumours relapse could promote the use of preventive therapy and could increase patients' survival. GBM stem cells have been recently described and it was demonstrated that they are specifically implied in the experimental tumorigenesis. It is thus very attractive to speculate on a possible relationship between these GBM stem cells and the neural stem cells which are persisting in the neurogenic zones of the adult brain. In this review, we formulate and discuss the hypothesis by which, in a patient with GBM, malignant stem cells might be present in the neurogenic zones, away from the tumour mass. This hypothesis could explain the tumour relapse observed after the first treatments.

[Recent advances in the treatment of brain tumours]. / Actualités neurochirurgicales dans le traitement des tumeurs cérébrales.

Neuronavigation is a tool for image guidance surgery. Based on the principle of the GPS, it is notably used for the ablation of brain tumors. Because of their millimetre precision, neuronavigation devices bring more safety and effectiveness due to the ever increasing performances of medical imaging. However, neuronavigation presents a major pitfall as it uses a static support (the images acquired preoperatively) to perform a dynamic process (the surgical ablation). To preserve the performance of neuronavigation, it is mandatory to update the images during surgery. This is now achievable by interventional MRI, intra-operative ultrasound and the incorporation of fluorescent tracers by the tumor cells. These major tools, now available at Sart Tilman University Hospital of combined with state-of-the-art chemotherapy, radiotherapy and experimental protocols (including gene therapy) will undoubtedly improve the prognosis of brain tumors.

Rat gap junction connexin-30 inhibits proliferation of glioma cell lines.

Connexins, the structural components of gap junctions, control cell growth and differentiation and are believed to belong to a family of tumour suppressor genes. Studies on connexin localization in brain showed that several of these proteins were expressed in distinct compartments of the brain in a cell-type specific manner, indicating that different gap junctions play specific roles in the physiology of the mammalian brain. In this report, we first cloned rat connexin-30 cDNA from brain and showed that it was expressed in long-term primary culture of rat astrocytes. In order to examine the potential role of connexin-30 in tumour cell proliferation, we transfected the connexin-30 cDNA into two rat glioma cell lines (9L and C6) which have lost its expression. Transfected clones adequately expressed membrane-bound connexin-30 protein. Connexin-30-expressing clones showed slower growth, lower DNA synthesis and reduced proliferation in soft agar as compared with the parental and control cells. We concluded that connexin-30 may also probably be considered as a tumour suppressor in rat gliomas.

Nuclear factor-kappa B, cancer, and apoptosis.

The role of nuclear factor (NF)-kappa B in the regulation of apoptosis in normal and cancer cells has been extensively studied in recent years. Constitutive NF-kappa B activity in B lymphocytes as well as in Hodgkin's disease and breast cancer cells protects these cells against apoptosis. It has also been reported that NF-kappa B activation by tumor necrosis factor (TNF)-alpha, chemotherapeutic drugs, or ionizing radiations can protect several cell types against apoptosis, suggesting that NF-kappa B could participate in resistance to cancer treatment. These observations were explained by the regulation of antiapoptotic gene expression by NF-kappa B. However, in our experience, inhibition of NF-kappa B activity in several cancer cell lines has a very variable effect on cell mortality, depending on the cell type, the stimulus, and the level of NF-kappa B inhibition. Moreover, in some experimental systems, NF-kappa B activation is required for the onset of apoptosis. Therefore, it is likely that the NF-kappa B antiapoptotic role in response to chemotherapy is cell type- and signal-dependent and that the level of NF-kappa B inhibition is important. These issues will have to be carefully investigated before considering NF-kappa B as a target for genetic or pharmacological anticancer therapies.

Growth regulation of astrocytes and C6 cells by TGFbeta1: correlation with gap junctions.

Transforming growth factor (TGF) beta1 enhanced in vitro [3H]thymidine incorporation into C6 cells and reduced that of astrocytes in the presence of a high serum concentration. It concomitantly raised the gap junction intercellular communication (GJIC) in normal astrocytes but reduced the coupling of C6 cells, and respectively increased or decreased the proportion of P2-phosphorylated connexin (Cx) 43 isoform in these cells. Finally, octanol, which inhibited GJIC in both cell types, increased the thymidine incorporation in C6 cells, but neither altered the proliferation of astrocytes nor their response to TGFbeta1. These data indicate that an inhibition of gap junction intercellular communication, due to an altered phosphorylation of connexin 43, may contribute to the proliferative response of C6 glioblastoma cells to TGFbeta1.

Pharmacological modulation of the bystander effect in the herpes simplex virus thymidine kinase/ganciclovir gene therapy system: effects of dibutyryl adenosine 3',5'-cyclic monophosphate, alpha-glycyrrhetinic acid, and cytosine arabinoside.

The herpes simplex virus type 1 thymidine kinase (HSV1-tk) suicide gene/ganciclovir system was first applied to the treatment of glioblastoma tumors, but was hampered by the low gene transfection yield. Fortunately, the gap junction-dependent diffusion of phosphorylated ganciclovir metabolites from transfected cells to their neighbors proved to enhance the overall benefit of this strategy. However, as tumor cells are often gap junction-deficient, we sought to restore this property pharmacologically and hence to improve the efficacy of the treatment. We demonstrated that this approach was feasible in glioblastoma cells using dibutyryl adenosine 3',5'-cyclic monophosphate (cAMP) (100 microM) as a pharmacological inducer of gap junctions. alpha-Glycyrrhetinic acid (25 microM), on the other hand, strongly inhibited both gap junction-mediated intercellular communication and the bystander effect, thus confirming the role of gap junctions in HSV-tk-mediated bystander killing. Using cytosine arabinoside as a growth inhibitor, we underlined the role of tumor cell proliferation in the sensitivity of HSV-tk-transfected cells to ganciclovir and demonstrated its correlation with the importance of the bystander effect.