Structure-based virtual screening and molecular docking approaches to identify potential inhibitors against KIF2C to combat glioma.

Glioma, a kind of malignant brain tumor, is extremely lethal. Kinesin family member 2C (KIF2C) was found to have an aberrant expression in several cancer types, including lung cancer and glioma. KIF2C may therefore be a useful therapeutic target for the treatment of glioma. In the current study, new drug candidates that may function as KIF2C enzyme inhibitors were discovered. MTi OpenScreen was used to carry out the structure-based virtual screening of an inbuilt drug library containing 150,000 compounds. These compounds belong to different classes, such as natural product-based compounds (NP-lib), purchasable approved drugs (Drugs-lib), and food constituents compound collection (FOOD-lib). Based on their binding affinities, a total of 84 compounds were further pushed to calculate ADMET properties. The compounds (16) meeting the ADMET cutoff ranges were then further docked to the receptor to find their plausible binding modes using the Glide tool's standard precision (SP) technique. The docking results were examined using the Glide gscore, and the best binding compounds (Rimacalib and Sarizotan) were chosen to test their stability with KIF2C protein through molecular dynamics (MD) simulation. Similarly, Principal Component Analysis and cross-correlation matrix were also examined. The MM/GBSA binding free energies showed a considerable energy contribution in the binding of hits with the KIF2C. Collectively, these findings strongly suggest the potential of the lead compounds to inhibit the biological function of KIF2C, emphasizing the need for further investigation in this area.Communicated by Ramaswamy H. Sarma.

The anterior gradient homologue 2 (AGR2) co-localises with the glucose-regulated protein 78 (GRP78) in cancer stem cells, and is critical for the survival and drug resistance of recurrent glioblastoma: in situ and in vitro analyses.

BACKGROUND: Glioblastomas (GBs) are characterised as one of the most aggressive primary central nervous system tumours (CNSTs). Single-cell sequencing analysis identified the presence of a highly heterogeneous population of cancer stem cells (CSCs). The proteins anterior gradient homologue 2 (AGR2) and glucose-regulated protein 78 (GRP78) are known to play critical roles in regulating unfolded protein response (UPR) machinery. The UPR machinery influences cell survival, migration, invasion and drug resistance. Hence, we investigated the role of AGR2 in drug-resistant recurrent glioblastoma cells. METHODS: Immunofluorescence, biological assessments and whole exome sequencing analyses were completed under in situ and in vitro conditions. Cells were treated with CNSTs clinical/preclinical drugs taxol, cisplatin, irinotecan, MCK8866, etoposide, and temozolomide, then resistant cells were analysed for the expression of AGR2. AGR2 was repressed using single and double siRNA transfections and combined with either temozolomide or irinotecan. RESULTS: Genomic and biological characterisations of the AGR2-expressed Jed66_GB and Jed41_GB recurrent glioblastoma tissues and cell lines showed features consistent with glioblastoma. Immunofluorescence data indicated that AGR2 co-localised with the UPR marker GRP78 in both the tissue and their corresponding primary cell lines. AGR2 and GRP78 were highly expressed in glioblastoma CSCs. Following treatment with the aforementioned drugs, all drug-surviving cells showed high expression of AGR2. Prolonged siRNA repression of a particular region in AGR2 exon 2 reduced AGR2 protein expression and led to lower cell densities in both cell lines. Co-treatments using AGR2 exon 2B siRNA in conjunction with temozolomide or irinotecan had partially synergistic effects. The slight reduction of AGR2 expression increased nuclear Caspase-3 activation in both cell lines and caused multinucleation in the Jed66_GB cell line. CONCLUSIONS: AGR2 is highly expressed in UPR-active CSCs and drug-resistant GB cells, and its repression leads to apoptosis, via multiple pathways.

A paediatric dysembryoplastic neuroepithelial tumour (DNET) with deregulated stem cell markers: a case report.

Background: Dysembryoplastic neuroepithelial tumours (DNETs) are rare, with only a few reported lethal cases. Currently, there are focused efforts by neuro-oncology professionals to reveal the molecular characterisations of individual central nervous system tumours (CNSTs). Here, we report the status of cancer stem cell (CSC) genes associated with resilience and drug resistance in a paediatric DNET, since the deregulations and variations of CSC genes may prove critical to these tumours' molecular characterisations. Case Description: Immunofluorescence, clonogenic assay and whole exome sequencing (WES) were applied to the patient's tissue and its corresponding cell line. The case is for of a 6-year-old boy with intractable epilepsy and unremarkable physical and neurological examinations. Following magnetic resonance imaging (MRI) and histopathological tests, the patient was diagnosed with DNET. The child underwent a right posterior temporoparietooccipital neuronavigation-assisted craniotomy. Several CSC markers were upregulated in situ, including the metastasis-related protein, anterior gradient 2 (AGR2; 67%), and the Wnt-signalling-related protein, frizzled class receptor 9 (FZD9; 79%). The cell line possessed a similar DNA profile as the original tissue, stained positive for the tumorigenic marker [BMI1 proto-oncogene (BMI)] and CSC markers, and displayed drug resistance. Variants identified in the tissue DNA, which are listed in the catalogue of somatic mutations in cancer (COSMIC) database for genes previously known to be necessary for the development of the embryonic brain, included variants in the cell division cycle 27 (CDC27) gene. Conclusions: we report the in situ and in vitro presence of CSCs in a paediatric DNET.

The efficacy of interactive group psychoeducation for children with leukaemia: A randomised controlled trial.

OBJECTIVE: To evaluate an interactive group psychoeducation programme for children treated for leukaemia. METHODS: A longitudinal randomised controlled study across four UK hospitals with an immediate (N = 26) and delay control group (N = 32). The intervention covered the pathophysiology of leukaemia, its treatment, side effects and the importance of positive health behaviours. Primary outcomes were parent-reported child health related quality of life (HRQoL) and behavioural difficulties. Secondary outcomes were child-reported HRQoL, cancer-specific HRQoL, child confidence, caregiver burden, and treatment anxiety. Measures were completed pre- and immediately post-intervention, and at 13 and 26-weeks follow-up. Change over time was analysed using multilevel modelling. Acceptability questionnaires rated the intervention on benefits, recommendations, and barriers to participation. RESULTS: The intervention significantly improved parent-reported child HRQoL but did not have a significant effect on other outcomes. Acceptability of the intervention was high. CONCLUSIONS: This study provides initial evidence that interactive group psychoeducation is acceptable to families and improves HRQoL in children with leukaemia. Difficulties with recruitment removed power to detect effect sizes that are plausible for psychoeducational interventions. PRACTISE IMPLICATIONS: Further studies to explore the potential of psychoeducation to improve outcomes for children with leukaemia and an examination of barriers to participation within this population are warranted.

Overlapping variants in the blood, tissues and cell lines for patients with intracranial meningiomas are predominant in stem cell-related genes.

OBJECTIVE: Bulk tissue genomic analysis of meningiomas identified common somatic mutations, however, it often excluded blood-related variants. In contrast, genomic characterisation of primary cell lines that can provide critical information regarding growth and proliferation, have been rare. In our work, we identified the variants that are present in the blood, tissues and corresponding cell lines that are likely to be predictive, tumorigenic and progressive. METHOD: Whole-exome sequencing was used to identify variants and distinguish related pathways that exist in 42 blood, tissues and corresponding cell lines (BTCs) samples for patients with intracranial meningiomas. Conventional sequencing was used for the confirmation of variants. Integrative analysis of the gene expression for the corresponding samples was utilised for further interpretations. RESULTS: In total, 926 BTC variants were detected, implicating 845 genes. A pathway analysis of all BTC genes with damaging variants indicated the 'cell morphogenesis involved in differentiation' stem cell-related pathway to be the most frequently affected pathway. Concordantly, five stem cell-related genes, GPRIN2, ALDH3B2, ASPN, THSD7A and SIGLEC6, showed BTC variants in at least five of the patients. Variants that were heterozygous in the blood and homozygous in the tissues or the corresponding cell lines were rare (average: 1.3 ± 0.3%), and included variants in the RUNX2 and CCDC114 genes. An analysis comparing the variants detected only in tumours with aggressive features indicated a total of 240 BTC genes, implicating the 'homophilic cell adhesion via plasma membrane adhesion molecules' pathway, and identifying the stem cell-related transcription coactivator NCOA3/AIB1/SRC3 as the most frequent BTC gene. Further analysis of the possible impact of the poly-Q mutation present in the NCOA3 gene indicated associated deregulation of 15 genes, including the up-regulation of the stem cell related SEMA3D gene and the angiogenesis related VEGFA gene. CONCLUSION: Stem cell-related pathways and genes showed high prevalence in the BTC variants, and novel variants in stem cell-related genes were identified for meningioma. These variants can potentially be used as predictive, tumorigenic and progressive biomarkers for meningioma.

Resting-state electroencephalogram in learning-disabled children: power and connectivity analyses.

The neurophysiological underpinnings of learning disabilities remain unknown. In this clinical study, we recorded electroencephalograms for a large sample of children with learning disabilities (LD) and healthy control children (n=216) during resting states in which the eyes were either open or closed. We calculated the power and lagged phase coherence in six main frequency bands (delta, theta, lower and upper alpha, and lower and upper beta) to re-evaluate the question of whether children with LD show frontal theta power increases and posterior alpha band decreases on the basis of patterns of electroencephalogram oscillation, which could then be considered as evidence for the so-called 'maturational delay hypothesis.' We identified a general (not restricted to frontal electrodes) power increase in the theta band and no accompanying concomitant alpha band decrease at the posterior electrode position. In addition, we observed increased beta band power at frontal electrodes for LD children. With respect to lagged phase coherence, which is a coherence measure not influenced by volume conduction, we identified decreased coherence for children with LD in the upper alpha band during the eyes closed condition. We interpret this LD-specific resting-state activation pattern as indicating a suboptimally functioning neural resting-state network that provides a detrimental 'starting point' for task-specific brain activations.

Public awareness of central nervous system tumors in the Kingdom of Saudi Arabia.

OBJECTIVE: To investigate individuals` knowledge about central nervous system tumors (CNST) signs and symptoms and risk factors, as well as their readiness to seek medical advice. The signs and symptoms associated with CNSTs are often vague, and failure to recognize them could lead to delays in seeking help and possibly fatal results. METHODS: This was a cross-sectional survey that utilized 2 delivery methods. A total of 1,500 personally delivered and 1,500 online self-administered questionnaires were completed in parallel between June 2015 and June 2016 for the occupants of the Kingdom of Saudi Arabia. RESULTS: Significant differences were observed for the sociodemographic characteristics of participants recruited via the 2 methods. The most recognized symptom was "Headaches" (45.2%), and the most recognized risk factor was "Radioactive location/occupation" (84.1%). Overall knowledge scores were low, significantly predicted by employment and cancer contact (p<0.05), while the scores significantly higher for participants who were willing to see their doctors within a week (p<0.005). The most recognized barrier to seeking help was "Worry about what the doctor might find" (74.0%). CONCLUSION: The level of awareness of CNSTs was low. Using a questionnaire delivered in 2 different ways enabled the recruitment of sample pools with different sociodemographic characteristics.

In situ characterization of stem cells-like biomarkers in meningiomas.

BACKGROUND: Meningioma cancer stem cells (MCSCs) contribute to tumor aggressiveness and drug resistance. Successful therapies developed for inoperable, recurrent, or metastatic tumors must target these cells and restrict their contribution to tumor progression. Unfortunately, the identity of MCSCs remains elusive, and MSCSs' in situ spatial distribution, heterogeneity, and relationship with tumor grade, remain unclear. METHODS: Seven tumors classified as grade II or grade III, including one case of metastatic grade III, and eight grade I meningioma tumors, were analyzed for combinations of ten stem cell (SC)-related markers using immunofluorescence of consecutive sections. The correlation of expression for all markers were investigated. Three dimensional spatial distribution of markers were qualitatively analyzed using a grid, designed as a repository of information for positive staining. All statistical analyses were completed using Statistical Analysis Software Package. RESULTS: The patterns of expression for SC-related markers were determined in the context of two dimensional distribution and cellular features. All markers could be detected in all tumors, however, Frizzled 9 and GFAP had differential expression in grade II/III compared with grade I meningioma tissues. Correlation analysis showed significant relationships between the expression of GFAP and CD133 as well as SSEA4 and Vimentin. Data from three dimensional analysis showed a complex distribution of SC markers, with increased gene hetero-expression being associated with grade II/III tumors. Sub regions that showed multiple co-staining of markers including CD133, Frizzled 9, GFAP, Vimentin, and SSEA4, but not necessarily the proliferation marker Ki67, were highly associated with grade II/III meningiomas. CONCLUSION: The distribution and level of expression of CSCs markers in meningiomas are variable and show hetero-expression patterns that have a complex spatial nature, particularly in grade II/III meningiomas. Thus, results strongly support the notion of heterogeneous populations of CSCs, even in grade I meningiomas, and call for the use of multiple markers for the accurate identification of individual CSC subgroups. Such identification will lead to practical clinical diagnostic protocols that can quantitate CSCs, predict tumor recurrence, assist in guiding treatment selection for inoperable tumors, and improve follow up of therapy.

Pleomorphism and drug resistant cancer stem cells are characteristic of aggressive primary meningioma cell lines.

BACKGROUND: Meningioma tumors arise in arachnoid membranes, and are the most reported central nervous system (CNS) tumors worldwide. Up to 20% of grade I meningioma tumors reoccur and currently predictive cancer stem cells (CSCs) markers for aggressive and drug resistant meningiomas are scarce. METHODS: Meningioma tissues and primary cell lines were investigated using whole transcriptome microarray analysis, immunofluorescence staining of CSCs markers (including CD133, Sox2, Nestin, and Frizzled 9), and drug treatment with cisplatin or etoposide. RESULTS: Unsupervised hierarchical clustering of six meningioma samples separated tissues into two groups. Analysis identified stem cells related pathways to be differential between the two groups and indicated the de-regulation of the stem cell associated genes Reelin (RELN), Calbindin 1 (CALB1) and Anterior Gradient 2 Homolog (AGR2). Immunofluorescence staining for four tissues confirmed stemness variation in situ. Biological characterization of fifteen meningioma primary cell lines concordantly separated cells into two functionally distinct sub-groups. Pleomorphic cell lines (NG type) grew significantly faster than monomorphic cell lines (G type), had a higher number of cells that express Ki67, and were able to migrate aggressively in vitro. In addition, NG type cell lines had a lower expression of nuclear Caspase-3, and had a significantly higher number of CSCs co-positive for CD133+ Sox2+ or AGR2+ BMI1+. Importantly, these cells were more tolerant to cisplatin and etoposide treatment, showed a lower level of nuclear Caspase-3 in treated cells and harbored drug resistant CSCs. CONCLUSION: Collectively, analyses of tissues and primary cell lines revealed stem cell associated genes as potential targets for aggressive and drug resistant meningiomas.

Detection of ß-Thalassemia Mutations Using TaqMan Single Nucleotide Polymorphism Genotyping Assays.

AIMS: Sickle-cell anemia and ß-thalassemia are two of the most common autosomal recessive disorders in the developing world. The severity of the problem and the pressure it exerts on the health services in the Kingdom of Saudi Arabia forced the introduction of a national premarital screening program to lessen its impact on the society. Furthermore, a significant effort has been exerted in the elucidation of the genetic causes of such diseases to facilitate diagnosis and detection of carriers. METHODS: We have designed and validated the use of custom TaqMan(®) genotyping assays for the rapid detection of IVS-I-1 (G>A), IVS-I-5 (G>C), codon 39 (C>T), and IVS-I-110 (G>A) mutations in transfusion-dependent ß-thalassemia patients' cohort. RESULTS: We demonstrated that IVS-I-5 (rs33915217) is the most common single-nucleotide variant in our cohort, with the variant allele constituting 26% of the total alleles investigated. However, this variant was not found in 352 alleles screened from buccal swab DNA obtained from healthy volunteers. CONCLUSION: The TaqMan single nucleotide polymorphism (SNP) genotyping assays are a rapid, accurate, and cost-effective method for the initial screening of ß-thalassemia cases, which will minimize the need for direct sequencing of the HBB gene, thus reducing detection costs and increasing throughput.

Breast cancer awareness and breast self-examination in Northern Saudi Arabia. A preliminary survey.

OBJECTIVE: To elicit knowledge of breast cancer, perception of occurrence, and behavior in relation to breast self-examination (BSE). METHODS: A cross-sectional survey was carried out at the Department of Pathology, Medical College, University of Hail, Hail, Kingdom of Saudi Arabia for local occupants from Hail city and its rural neighborhood between September 2010 and February 2012. A personal interview-administered descriptive questionnaire and both descriptive and inferential statistics were used. RESULTS: A total of 1000 participants agreed to be involved, out of which 87.7% were females, 7.2% were males and 5.1% had undisclosed gender. The age range for participants was 12-66 years. Out of all participants, 44% did not know that breast cancer is an abnormal growth and 78% failed to recognise its multi-factorial nature, with Increased age being the least recognised single risk factor 4.8%. Scores showed that 61.5% had a low level of breast cancer related knowledge. Out of the participants who knew of someone who had breast cancer 73%, 50.1% said the disease was discovered at a Late stage mainly by Chance. Data for BSE indicated that 50.1% of female participants >16 years old did not practice BSE, and Fear was the main declared perceived reason. CONCLUSION: This study demonstrates a low level of fundamental knowledge of breast cancer and fear to practice BSE.

Inter-trial variability in sensory-evoked cortical hemodynamic responses: the role of the magnitude of pre-stimulus fluctuations.

Brain imaging techniques utilize hemodynamic changes that accompany brain activation. However, stimulus-evoked hemodynamic responses display considerable inter-trial variability and the sources of this variability are poorly understood. One of the sources of this response variation could be ongoing spontaneous hemodynamic fluctuations. We recently investigated this issue by measuring cortical hemodynamics in response to sensory stimuli in anesthetized rodents using 2-dimensional optical imaging spectroscopy. We suggested that sensory-evoked cortical hemodynamics displayed distinctive response characteristics and magnitudes depending on the phase of ongoing fluctuations at stimulus onset due to a linear superposition of evoked and ongoing hemodynamics (Saka et al., 2010). However, the previous analysis neglected to examine the possible influence of variability of the size of ongoing fluctuations. Consequently, data were further analyzed to examine whether the size of pre-stimulus hemodynamic fluctuations also influenced the magnitude of subsequent stimulus-evoked responses. Indeed, in the case of all individual trials, a moderate correlation between the size of the pre-stimulus fluctuations and the magnitudes of the subsequent sensory-evoked responses were observed. However, different correlations between the size of the pre-stimulus fluctuations and magnitudes of the subsequent sensory-evoked cortical hemodynamic responses could be observed depending on their phase at stimulus onset. These analyses suggest that both the size and phase of pre-stimulus fluctuations in cortical hemodynamics contribute to inter-trial variability in sensory-evoked responses.

Linear superposition of sensory-evoked and ongoing cortical hemodynamics.

Modern non-invasive brain imaging techniques utilize changes in cerebral blood flow, volume and oxygenation that accompany brain activation. However, stimulus-evoked hemodynamic responses display considerable inter-trial variability even when identical stimuli are presented and the sources of this variability are poorly understood. One of the sources of this response variation could be ongoing spontaneous hemodynamic fluctuations. To investigate this issue, 2-dimensional optical imaging spectroscopy was used to measure cortical hemodynamics in response to sensory stimuli in anesthetized rodents. Pre-stimulus cortical hemodynamics displayed spontaneous periodic fluctuations and as such, data from individual stimulus presentation trials were assigned to one of four groups depending on the phase angle of pre-stimulus hemodynamic fluctuations and averaged. This analysis revealed that sensory evoked cortical hemodynamics displayed distinctive response characteristics and magnitudes depending on the phase angle of ongoing fluctuations at stimulus onset. To investigate the origin of this phenomenon, "null-trials" were collected without stimulus presentation. Subtraction of phase averaged "null trials" from their phase averaged stimulus-evoked counterparts resulted in four similar time series that resembled the mean stimulus-evoked response. These analyses suggest that linear superposition of evoked and ongoing cortical hemodynamic changes may be a property of the structure of inter-trial variability.

Polarographic Electrode Measures of Cerebral Tissue Oxygenation: Implications for Functional Brain Imaging.

The changes in blood flow, blood volume and oxygenation that accompany focal increases in neural activity are collectively referred to as the hemodynamic response and form the basis of non-invasive neuroimaging techniques such as blood oxygen level dependent (BOLD) functional magnetic resonance imaging. A principle factor influencing blood oxygenation, the cerebral metabolic rate of oxygen consumption is poorly understood and as such, data from imaging techniques are difficult to interpret in terms of the underlying neural activity. In particular how neurometabolic changes vary temporally, spatially and in magnitude remains uncertain. Furthermore knowledge of which aspects of neural activity are closely reflected by metabolic changes is essential for the correct interpretation of cognitive neuroscience studies in terms of information processing. Polarographic electrode measurements of cerebral tissue oxygenation in animal models following presentation of sensory stimuli have started to address these issues. Early studies demonstrated both increases and decreases in tissue oxygenation following neural activation. However a recent series of elegant studies in the cat visual system demonstrated a tight spatial and temporal coupling between evoked peri-synaptic activity and oxygen consumption following presentation of visual stimuli.

A microarray study of gene and protein regulation in human and rat brain following middle cerebral artery occlusion.

BACKGROUND: Altered gene expression is an important feature of ischemic cerebral injury and affects proteins of many functional classes. We have used microarrays to investigate the changes in gene expression at various times after middle cerebral artery occlusion in human and rat brain. RESULTS: Our results demonstrated a significant difference in the number of genes affected and the time-course of expression between the two cases. The total number of deregulated genes in the rat was 335 versus 126 in the human, while, of 393 overlapping genes between the two array sets, 184 were changed only in the rat and 36 in the human with a total of 41 genes deregulated in both cases. Interestingly, the mean fold changes were much higher in the human. The expression of novel genes, including p21-activated kinase 1 (PAK1), matrix metalloproteinase 11 (MMP11) and integrase interactor 1, was further analyzed by RT-PCR, Western blotting and immunohistochemistry. Strong neuronal staining was seen for PAK1 and MMP11. CONCLUSION: Our findings confirmed previous studies reporting that gene expression screening can detect known and unknown transcriptional features of stroke and highlight the importance of research using human brain tissue in the search for novel therapeutic agents.

Cellular prion protein is increased in the plasma and peri-infarcted brain tissue after acute stroke.

The physiologic properties of the normal cellular prion protein (PrP(C)) have not been established fully, although recent evidence showed its upregulation in cerebral ischaemia. Using patients, animal models, and in vitro studies we aimed to identify in detail the expression and localization of PrP(C) in ischemic stroke. Patients in acute phase of ischaemic stroke had increased plasma levels of circulating PrP(C) as compared to healthy age- and gender-matched controls (3.1 +/- 1.4 vs. 1.9 +/- 0.7 ng/ml, P = 0.002). Immunohistochemistry showed increased expression of PrP(C) in the soma of peri-infarcted neurones as well as in the endothelial cells (EC) of micro-vessels and inflammatory cells in peri-infarcted brain tissue from patients who survived for 2-34 days after an initial stroke. The same pattern was repeated 1-48 hr after MCAO. RT-PCR showed increased gene expression of PrP(C) by human foetal neurons (HFN) after 12 hr of oxygen glucose deprivation (OGD), which remained increased after 24 hr reperfusion. Western blotting confirmed that protein expression was similarly upregulated, and fluorescent labeling showed a notable increase in peri-nuclear and axonal PrP(C) staining intensity. Increased plasma PrP(C) seems to reflect endogenous expression in acute stroke-affected brain tissue. Increased cellular expression in peri-infarcted regions may influence hypoxia-induced cell damage, although the effects on EC survival and angiogenesis remain to be elucidated.

Expression of basic fibroblast growth factor mRNA and protein in the human brain following ischaemic stroke.

Our previous work has demonstrated that angiogenesis occurs in the damaged brain tissue of patients surviving acute ischaemic stroke and increased microvessel density in the penumbra is associated with longer patient survival. The brain is one of the richest sources of FGF-2 and several studies have noted its angiogenic and neuroprotective effects in the nervous system. These findings led us to investigate the expression and localisation of both FGF-2 mRNA and protein in brain tissue collected within 12 h of death from 10 patients who survived for between 24 h and 43 days after acute stroke caused by thrombosis or embolus. Western blot analysis demonstrated increased FGF-2 protein expression in both grey and white matter in the infarcted core and the penumbra region compared to the normal contralateral hemisphere of all 10 patients studied. Using indirect immunoperoxidase staining of paraffin embedded sections, we observed the presence of FGF-2 in neurones, astrocytes, macrophages and endothelial cells. In situ hybridisation was used to localise and quantify mRNA expression in ischaemic brain tissue of the same 10 patients. The expression of FGF-2 in the penumbra of all patients was significantly raised compared with infarcted tissue and normal-looking contralateral hemisphere. In addition, serum FGF-2 was significantly increased between 1 and 14 days (P<0.001) in many patients with both ischaemic stroke (n=28) and intra-cerebral haemorrhage (n=16) compared with age-matched control subjects undergoing routine medical examinations (n=20). We suggest that up-regulation of FGF-2 is one of the mechanisms that leads to angiogenesis and neuro-protection in the penumbra region after acute stroke in man.