NeuroOncology

Non-NF2 mutations have a key effect on inhibitory immune checkpoints and tumor pathogenesis in skull base meningiomas Abstract Aims Skull base meningiomas represent approximately 25% of all meningiomas, nearly 20% of which are atypical or anaplastic. To date, effective medical treatments for meningiomas are still lacking. Genetic aberrations (TRAF7, KLF4, AKT1, and SMO) and the effects of genetic aberrations on the expression of inhibitory immune checkpoint molecules (PD-L1, IDO, and TDO2) in skull base meningiomas are still unclear. Methods Genetic alterations in the four genes were identified in 92 skull base meningiomas by Sanger sequencing. The expression differences in immune checkpoints between mutant and wild-type (WT) tumors were determined by immunohistochemistry (IHC) and Western blot (WB). Results The four mutations were not concurrently detected in the patients with skull base meningiomas. Among the tumors from the KLF4-mutated group, almost half were petroclival meningiomas. KLF4- and TRAF7-mutated tumors were predominantly secretory meningiomas. SMO-mutated tumors exhibited higher calcification, and half of these tumors were observed in the brain midline. Receiver operating characteristic curve analysis indicated that tumor volume can predict KLF4 and TRAF7 mutation status with high sensitivity and specificity, respectively. The IHC and WB analyses indicated that PD-L1, IDO, and TDO2 levels in tumors with TRAF7 mutations were significantly higher than those in WT tumors. Meanwhile, there was a significant difference in TDO2 between tumors with AKT1 mutations and WT tumors. Specifically, TRAF7 mutations could play a key role in skull base meningiomas by regulating the expression of inhibitory immune checkpoints and thus suppressing immune responses. Conclusions Checkpoint inhibitors may be potential strategies for targeted immunotherapies of these mutant meningiomas.

Super-early initiation of temozolomide prolongs the survival of glioblastoma patients without gross-total resection: a retrospective cohort study Abstract Objective The optimal timing of chemoradiotherapy in patients with newly diagnosed glioblastoma (GBM) remains unclear. In this study, we explored the clinical efficacy of super-early initiation of temozolomide (TMZ) in the treatment interval from surgery to radiotherapy. Methods We retrospectively reviewed the clinical data of 375 patients with GBM in our institution from 2012 to 2018. One hundred and sixty-three patients received super-early TMZ within 7 days after craniotomy based on standard Stupp protocol (super-early group, SEG), while two hundred and twelve patients underwent standard Stupp protocol alone (control group, CG). We performed propensity score matching (PSM) to reduce patient selection bias between the two groups. Results Before PSM, both median progression-free survival (PFS) and overall survival (OS) of patients in SEG were longer than those in CG (PFS 11.5 vs. 9.0 months, P = 0.0384 and OS 23.0 vs. 17.0 months, P = 0.0014). After PSM, the clinical efficacy of super-early initiation of TMZ only remained significant in term of OS, which was further validated in Cox hazard proportional model (HR = 0.583, 95% CI 0.384–0.884, P = 0.011). In the subgroup analysis, patients without gross total resection (GTR) or with O6-methylguanine DNA methyltransferase promoter methylation could benefit from super-early initiation of TMZ in both PFS and OS (P < 0.05). No significant difference of treatment emerging adverse events was observed between the two groups (P > 0.05). Conclusions This retrospective study highlights that super-early initiation of TMZ in newly diagnosed GBM may confer to survival benefit, especially for those without GTR.

LncRNA GAS5 regulates the proliferation, migration, invasion and apoptosis of brain glioma cells through targeting GSTM3 expression. The effect of LncRNA GAS5 on glioma cells Abstract Introduction To investigate the effects of lncRNA GAS5 on the proliferation, migration, invasion and apoptosis of brain glioma cells. Methods The expression levels of lncRNA GAS5 and GSTM3 in normal glial cells (HEB) and glioma cells (U251 and U87) were detected by RT-qPCR and western blot, respectively. Glioma cells were transfected with ctrl vector, pcDNA-GAS5, siRNA ctrl (siNC) or GSTM3 siRNA and the effects of lncRNA GAS5 and GSTM3 on the proliferation, migration, invasion and apoptosis of glioma cells were detected by CCK-8 assay, transwell assay and Caspase 3/7 activity assay, respectively. Results The expression of lncRNA GAS5 was significantly decreased in glioma cell lines U251 and U87 compared with normal glial cells HEB (p < 0.01). In addition, overexpression of lncRNA GAS5 inhibited the proliferation, migration and invasion of U251 and U87 cells, and promoted cell apoptosis as demonstrated by the increased activity of Caspase 3/7. Furthermore, GSTM3 was predicted as a target gene of lncRNA GAS5 by bioinformatics analysis and its expression was increased in glioma cells compared with the normal cells as indicated by western blotting and RT-qPCR experimental results. Silencing of GSTM3 with GSTM3 siRNA decreased the proliferation, migration and invasion but increased the apoptosis of glioma cell lines U251 and U87, which was similar to that the effect lncRNA GAS5 over-expression. Conclusion lncRNA GAS5 can effectively inhibit the proliferation, migration and invasion of glioma cells and promote cell apoptosis through targeting GSTM3 expression.

Correction to: Phase I-II trial of imatinib mesylate (Gleevec; STI571) in treatment of recurrent oligodendroglioma and mixed oligoastrocytoma. North central cancer treatment group study N0272 (ALLIANCE/NCCTG) The last author's first name was truncated in the initial online publication. The original article has been corrected.

Radiotherapy for recurrent intracranial epidermoid cysts without malignant transformation: a single-institution case series Abstract Introduction Recurrent intracranial epidermoid cysts may be difficult to address surgically given their proximity to critical neurovascular structures of the skull base. There are emerging reports of using radiotherapy (RT) for the treatment of recurrent epidermoid cysts. Here, we report a case series of adjuvant fractionated external beam RT for recurrent intracranial epidermoid cysts. Methods A single-institution review of all recurrent epidermoid cysts treated with adjuvant therapy between 2000 and 2017 was performed. Results Eight patients with recurrent epidermoid cysts who underwent adjuvant external beam RT were identified. Average age at initial diagnosis was 42.2 years, and median clinical follow-up after initial diagnosis and RT was 16.4 and 2.9 years, respectively. The median number of surgical resections prior to RT was 3 (range 2–5). Rationale for RT included multiple recurrent disease, rapid recurrence following prior resection, increased risk of further surgical morbidity, and patient preference. Median dose was 50.4 delivered in 1.8 Gy fractions (median 28 fractions). By the date of last follow-up, no patient has demonstrated progression, and there have been no cases of malignant degeneration. Conclusion Adjuvant RT should be considered in the context of recurrent epidermoid cysts to decrease the likelihood of further recurrences.

Identification of PEPT2 as an important candidate molecule in 5-ALA-mediated fluorescence-guided surgery in WHO grade II/III gliomas Abstract Purpose 5-aminolevulinic acid (5-ALA) fluorescence-guided surgery (FGS) appears to be a promising treatment for glioma. However, 5-ALA-mediated fluorescence cannot always be detected in grade II/III gliomas. We hypothesized that gene expression patterns in the Protoporphyrin IX (PpIX) synthesis pathway may be associated with intraoperative fluorescence status of grade II/III gliomas, and then attempted to identify the key molecule of 5-ALA-mediated fluorescence. Methods Using 50 surgically obtained specimens, which were diagnosed as grade II and III gliomas, we analyzed gene expression within the PpIX synthesis pathway to identify candidate molecules according to intraoperative 5-ALA-mediated fluorescence status. The most likely candidate gene was selected and confirmed by protein expression analysis. To evaluate the biological function of the molecule in PpIX synthesis, functional analysis was performed using specific, small interference (si)RNA in the SW-1783 human grade III glioma cell line. Results Among the genes involved in the porphyrin synthesis pathway, the mRNA expression of Peptide transporter 2 (PEPT2) in FGS fluorescence-positive gliomas was significantly higher than that in fluorescence-negative gliomas. Protein expression of PEPT2 was also significantly higher in the fluorescence-positive gliomas, which was confirmed by western blot analysis and immunofluorescence analysis. The siRNA-mediated downregulation of the mRNA and protein expression of PEPT2 led to decreased PpIX fluorescence intensity, as confirmed by fluorescence spectrum analysis. Conclusions The results suggest PEPT2 is an important candidate molecule in 5-ALA-mediated FGS in grade II/III gliomas. As the overexpression of PEPT2 was associated with higher PpIX fluorescence intensity, PEPT2 may improve fluorescence-guided resection in grade II/III gliomas.

Clinical diagnosis of attention-deficit/hyperactivity disorder in survivors of pediatric brain tumors Abstract Purpose Survivors of pediatric brain tumors often have neurodevelopmental late effects, such as inattention. Symptoms may mirror those of attention-deficit/hyperactivity disorder (ADHD), which affects ~ 5–8% of the general population. This retrospective study of survivors followed at a large tertiary care center examined the prevalence of a clinical diagnosis of ADHD, and risk factors associated with ADHD diagnosis and ADHD-related medication use. Methods A retrospective chart review of brain tumor survivors (n = 528), diagnosed between 2000 and 2015, who were at least 6 years old and 2 years from the end of tumor-directed therapy or from diagnosis, if no interventions were received. Clinical and demographic data were extracted from the medical record. Results Survivors were 55.7% male with mean age 8.15 ± 4.4 (0.0–16.0) years at brain tumor diagnosis. The most common diagnoses were low-grade glioma, medulloblastoma, and craniopharyngioma, with 52.5% of tumors supratentorial. Of the survivors, 81.3% received surgery, 40.0% radiation therapy, and 36.6% chemotherapy. Sixty-nine survivors (13.1%) had ADHD diagnoses, 105 (19.9%) had symptoms of ADHD without diagnoses, and 64 (12.1%) had ADHD medication use. ADHD diagnosis was associated with younger age at tumor diagnosis (p = 0.05) and supratentorial tumor location (p = 0.001). ADHD diagnosis was not associated with gender, tumor type, or treatment type. Conclusions Survivors of brain tumors are at increased risk of ADHD and related symptoms. The greatest increase in risk occurs for survivors with diagnoses at younger ages and supratentorial tumors. Additional research is warranted, as select survivors may benefit from behavioral or pharmacologic ADHD treatments to optimize functioning.

Combination anti-CXCR4 and anti-PD-1 immunotherapy provides survival benefit in glioblastoma through immune cell modulation of tumor microenvironment Abstract Background Emerging evidence suggests that myeloid cells play a critical role in glioblastoma (GBM) immunosuppression. Disappointing results of recent checkpoint inhibitor trials suggest that combination immunotherapy with alternative agents could be fruitful in overcoming immunosuppression. Overexpression of chemokine receptor CXCR4 is associated with poor prognosis in GBM. We investigate the treatment effects of combination immunotherapy with anti-PD-1 and anti-CXCR4 in a murine glioma model. Methods C57BL/6 mice were implanted with GL261-Luc+ glioma cells and randomized into 4 arms: (1) control (2) anti-PD-1 (3) anti-CXCR4, and (4) anti-PD-1 and anti-CXCR4 therapy. Overall survival and median survival were assessed. Cell populations were assessed by flow cytometry. Results Combination therapy conferred a significant survival benefit compared to control and monotherapy arms. Mice that received combination therapy demonstrated immune memory and decreased populations of immunosuppressive tumor-infiltrating leukocytes, such as monocytic myeloid-derived suppressor cells and microglia within the brain. Furthermore, combination therapy improved CD4+/CD8+ ratios in the brain as well as contributed to increased levels of pro-inflammatory cytokines. Conclusions Anti-CXCR4 and anti-PD-1 combination immunotherapy modulates tumor-infiltrating populations of the glioma microenvironment. Targeting myeloid cells with anti-CXCR4 facilitates anti-PD-1 to promote an antitumor immune response and improved survival rates.

Diagnostic delay and morbidity of central nervous system tumors in children and young adults: a pediatric hospital experience Abstract Introduction For children with central nervous system (CNS) tumors, survival rates remain significantly lower than other childhood malignancies with a substantial increase in disability of survivors. Given this, it is imperative that these children are identified at the earliest sign of symptom onset. Our institution aimed to identify diagnostic delays, morbidity and mortality, and specific barriers that may exist within our specific healthcare system that result in diagnostic delay. Methods A retrospective chart review was performed of newly diagnosed CNS tumors between January 1, 2008 and December 31, 2017. Results 235 patient cases were reviewed, 34 (14.5%) of which had an associated tumor predisposition syndrome. Median age at the time of diagnosis was 9 years (range 1 day to 25 years), with median number of days from symptom onset to definitive diagnosis of 42 days (interquartile range 14–120 days). Delays longer than 60 days occurred in 95 (47.5%) patients. The 10 year relative survival rate for all tumors was 86.8%. Conclusions Our institution had a shorter interval from symptom onset to diagnosis than currently reported in the literature, as well as a decrease in associated morbidity. In addition, for those with longer delays, we were able to characterize the etiology and barriers leading to these delays. With these identified, we are able to utilize this knowledge to further improve education and awareness in community members and healthcare professionals to continue to improve the time to diagnosis in the future.

A critical assessment of the quality of radiation therapy in Israel: time to initiation of treatment of spinal cord compression as an index of efficiency Abstract Introduction Radiotherapy departments function under workload pressure. We examined the process from referral to treatment initiation for spinal cord compression (SCC), one of the most daunting clinical scenarios in oncology. Methods We identified 235 patients with SCC, treated between 2013–2015. Two physicians classified cases as "emergent" or "urgent" (treatment within 24 or 72 h, respectively). Results The distribution of referrals over the week was uniform for inpatients. In contrast, there was a referral peak (62.27%) during the first two workdays for emergency ambulatory patients (p = 0.011). There were few weekend referrals in all groups (3.0%). There was a statistically shorter interval between referral and treatment for emergent versus urgent cases (0.94 days vs. 4.17 days; p < 0.0001, Bonferroni correction p < 0.0005). Conclusion Time elapsed between referral and treatment of SCC may constitute a quality index in neuro-oncology. Modern departments of radiotherapy should determine the degree to which they can successfully implement such treatment. Patients with cancer and their physicians should be taught to recognize signs of SCC to expedite intervention.

Alexandros Sfakianakis
Anapafseos 5 . Agios Nikolaos
Crete.Greece.72100
2841026182

6948891480

alsfakia@gmail.com

Biotechnology

Correction to: FabG: from a core to circumstantial catalyst In the original publication of the article, under section Polyketide synthesis: a pathway similar to fatty acid synthesis, the sentences "Phylogeny of KS domains and proteins of FAS and PKS, inferred by Bayesian estimation. Numbers above branches indicate posterior clade probability values." and "Branch length indicates number of inferred amino acid changes per position." in the first paragraph were included inadvertently.

Biocatalytic synthesis and characterization of sn -1/3 and sn -2 monoacylglycerols Abstract Objectives To investigate the lipase-catalyzed synthesis of high purity sn-1/3 and sn-2 monoacylglycerols (1/3-MAG and 2-MAG) of different fatty acids (FAs). Results The 1/3-MAGs of three FAs (16:0, 17:0, 16:1) were synthesized using lipase-catalyzed esterification of glycerol with FAs. The 2-MAGs were obtained from the ethanolysis of synthetic triacylglycerols using sn-1,3 regiospecific lipase. The effects of lipase types, substrate ratio, temperature, reaction time and lipase load on the MAG conversion were studied. Under the optimal conditions, high purities (96.74%, 95.44%, 92.96%) with acceptable isolated yields (51.00%, 54.28%, 46.00%) were obtained for 1/3-16:0-MAG, 1/3-17:0-MAG, and 1/3-16:1-MAG, respectively. For 2-16:0-MAG, 2-17:0-MAG, and 2-16:1-MAG, the purities were 92.64, 95.04, and 96.48%, with isolated yields of 50.64, 52.16, and 26.12%, respectively. The molecular structures of the synthetic compounds were confirmed by 1H NMR, and MS and the melting points were characterized by DSC. Conclusions High purity MAG isomers can be synthesized via lipase-catalyzed reactions to be building blocks for production of functional lipids, the melting points of which are largely governed by the hydrophobic interactions among the alkanyl chains.

Theophylline-inducible riboswitch accurately regulates protein expression at low level in Escherichia coli Abstract Objectives Fine-tuning of enzyme expression at low levels is an important challenge for metabolic engineers. Here, theophylline-inducible riboswitch for translational regulation was evaluated. The background expression, translation rate, and time delay for its induction was reported. Results To evaluate the effect of the amount of mRNA on its translation rate, transcription of the riboswitch RNA with red fluorescent protein (RFP) was controlled by the lac system with addition of isopropyl β-d-1-thiogalactopyranoside in Escherichia coli. Regardless of the amount of riboswitch mRNA, the translation of RFP was completely suppressed without theophylline during both growth and stationary phases. Furthermore, a strong positive correlation between theophylline concentration (0 to 1 mM) and specific RFP production rate was observed. The specific RFP production rate with the riboswitch was approximately 2.3% of that without the riboswitch. Furthermore, 60 min of time delay for RFP expression was observed after adding theophylline during the stationary phase. Conclusion Theophylline-inducible riboswitch precisely controls protein translation at low expression levels with significantly low background expression. It can emerge as a powerful tool for fine tuning of enzyme expression.

Pharmacological Notch pathway inhibition leads to cell cycle arrest and stimulates ascl1 and neurogenin2 genes expression in dental pulp stem cells-derived neurospheres Abstract Objective Human dental pulp-derived stem cells (hDPSCs) are becoming an attractive source for cell-based neurorestorative therapies. As such, it is important to understand the molecular mechanisms that regulate the differentiation of hDPSCs toward the neuronal fate. Notch signaling plays key roles in neural stem/progenitor cells (NS/PCs) maintenance and prevention of their differentiation. The aim of this study was to address the effects of Notch signaling inhibition on neurosphere formation of hDPSCs and neuronal differentiation of hDPSCs-neurospheres. Results hDPSCs were isolated from third molar teeth. The cultivated hDPSCs highly expressed CD90 and CD44 and minimally presented CD34 and CD45 surface markers. The osteo/adipogenic differentiation of hDPSCs was documented. hDPSCs were cultured in neural induction medium and N-[N-(3,5-difluorophenacetyl-l-alanyl)]-Sphenylglycine t-butyl ester (DAPT) was applied to impede Notch signaling during transformation into spheres or on the formed neurospheres. Our results showed that the size and number of neurospheres decreased and the expression profile of nestin, sox1 and pax6 genes reduced provided DAPT. Treatment of the formed neurospheres with DAPT resulted in the cleaved Notch1 reduction, G0/G1 arrest and a decline in L-lactate production. DAPT significantly reduced hes1 and hey1 genes, while ascl1 and neurogenin2 expressions augmented. The number of MAP2 positive cells improved in the DAPT-treated group. Conclusions Our findings demonstrated the Notch activity in hDPSCs-neurospheres. DAPT treatment positively regulated proneural genes expression and increased neuronal-like differentiation.

Recent advances in carbohydrate-based cancer vaccines Abstract Cancer is a complex multifactorial disease for which many promising therapeutic strategies such as immunotherapy are emerging. Malignant cells frequently express aberrant cell surface carbohydrates, which differentiate them from normal "healthy" cells. This characteristic presents a window for the development of synthetic carbohydrate antigen-based cancer vaccines which can be recognized by the immune system and can bring about T cell-dependent immune responses. Antibodies generated against the carbohydrate antigens partake in the inactivation of carbohydrate-decorated cancer cells, by slowing down tumor cell growth and inducing cancer cell apoptosis. Novel synthetic strategies for carbohydrate antigens have led to several synthetic cancer vaccine candidates. In the present review, we describe the latest progress in carbohydrate-based cancer vaccines and their clinical evaluation in various cancers.

Precursor-feeding and altered-growth conditions reveal novel blue pigment production by Rubrivivax benzoatilyticus JA2 Abstract Objective To explore the secondary metabolite biosynthetic potential of Rubrivivax benzoatilyticus JA2 using a new metabolite mining strategy. Results Combination of precursor-feeding and altered growth conditions were used to mine new biomolecules. Strain JA2 utilised l-phenylalanine as sole source of nitrogen and showed pigments production only under phenylalanine-amended aerobic cultures. Stable isotope based precursor feeding studies indicated the blue pigment consists of 4-phenyl rings derived from l-phenylalanine. The purified blue pigment displayed characteristic visible-absorption and pH-dependent color variations. Precursor-feeding under altered growth conditions activated the plausible novel aromatic pigment production in strain JA2. Conclusion Our approach unraveled the previously unknown pigment synthesis in strain JA2 and demonstrated the potential of mining strategy in discovering the hidden secondary metabolite repertoire in microorganisms.

FabG: from a core to circumstantial catalyst Abstract Core biochemical pathways such as Fatty-acid synthesis II (FAS II) is ascribed to the synthesis of fatty-acids, biotin and lipoic acid in prokaryotes. It has two dehydrogenases namely, FabG and FabI which interact with the fatty-acid chain bound to Acyl-carrier protein (ACP), a well-studied enzyme which binds to substrates of varying lengths. This protein–protein interaction 'broadens' the active site of these dehydrogenases thus, contributing to their flexible nature. This property is exploited for catalysing numerous chiral synthons, alkanes, long-chain alcohols and secondary metabolites in industries especially with FabG. FASI relegates FASII in eukaryotes making it a 'relic gene pool' and an antibacterial drug target with diverse inhibitor and substrate markush. FabG often substitutes other dehydrogenases for producing secondary metabolites in nature. This redundancy is probably due to gene duplication or addition events possibly making FabG, a progenitor to some of the complex short-chain dehydrogenases used in organisms and industries today.

MiR-499 inhibited hypoxia/reoxygenation induced cardiomyocytes injury by targeting SOX6 Abstract Objective MiR-499 has been reported to be expressed only in cardiomyocytes, and its expression would increase after acute myocardial infarction (AMI). miR-499 plays a role in the process of cardiomyocytes injury induced by hypoxia/reoxygenation (H/R), however, it still remains unclear. Results Hypoxia inhibited miR-499-5p expression and H/R induced apoptosis. SOX6 was a target gene of miR-499-5p, and high expression of miR-499-5p inhibited the expression of SOX6. MiR-499-5p reduced H9c2 cells injury by inhibiting the expression of SOX6, overexpression of which could reverse the effect of miR-499-5p on H9c2 cells. MiR-499-5p inhibited the levels of LDH and MDA, while overexpression of miR-499-5p inhibited H/R-induced cell apoptosis. MiR-499-5p could up-regulate the level of Bcl-2 and down-regulate the expression levels of Bax and caspase-3. However, SOX6 partially reversed these effects of miR-499-5p. Conclusion We proved that miR-499-5p inhibited H/R-induced cardiomyocytes injury by targeting SOX6. Our results suggested that miR-499-5p/SOX6 pathway may present a potential therapeutic target for the treatment of AMI.

Human genome-derived TOP1 matrix attachment region enhances transgene expression in the transfected CHO cells Abstract Objectives To investigate the effect of full-length fragment of DNA topoisomerase I gene (TOP1) matrix attachment regions (MARs) originating from the human genome on transgene expression in Chinese hamster ovary (CHO) cells and explore the underlying mechanisms. Results Results showed that TOP1 MAR cannot only enhance the transient and stable transgenic expression of enhanced green fluorescence protein (EGFP) but also increase long-term stability and ratio of positive colonies in transfected CHO cells with TOP1 MAR at the 5′ or 3′ ends of the EGFP expression cassette. Interestingly, the CHO cells were transfected with the 5′,3′ TOP1 MAR-containing vector featured the highest transient and stable expression, whereas those with the 3′ TOP1 MAR-containing vector exhibited the most effective stability and ratio of positive colonies. We also observed that transgene copy numbers and mRNA of egfp gene were correlated with the expression levels of EGFP protein in polyclonal CHO cells. However, the heterogeneity of expression in monoclonal CHO cells was unaffected by transgene copy number. Conclusions The findings may aid in the potential application of TOP1 MAR in expression enhancement of recombinant proteins in mammalian cells.

Pseudomonas monteilii PN1: a great potential P -nitrophenol degrader with plant growth promoting traits under drought and saline–alkali stresses Abstract Objectives To evaluate Pseudomonas monteilii strain PN1 for the removal efficiency of P-nitrophenol (PNP) in soils and its growth promotion of maize (Zea mays L.) seedlings under drought and saline–alkali stress. Results PN1 can survive in soils contaminated with PNP dosage between 90 and 155 mg/kg and considerably improved the removal PNP efficiency in soils. Drought and saline–alkali stress reduced maize seedling growth (root length, shoot height and dry or fresh weight) and improved the antioxidant enzyme activities and malondialdehyde (MDA) and proline (PRO) contents. However, maize seedlings treated with PN1 remarkably promoted their growth compared with the control. The reduction in antioxidant enzyme activities and MDA and PRO contents was significant. This result may be correlated to the increased tolerance of maize seedlings to drought and saline–alkali stress. Conclusions Application of P. monteilii PN1 can be an extremely useful approach for the development of bioinoculants in improving plant tolerance to several abiotic stresses and removing PNP in soils to ensure secure crop production.

Alexandros Sfakianakis
Anapafseos 5 . Agios Nikolaos
Crete.Greece.72100
2841026182

6948891480

alsfakia@gmail.com

Neuroradiology

European Society of Neuroradiology (ESNR)

Subtraction CT angiography in the follow-up of treated cerebral aneurysms

Application of MR morphologic, diffusion tensor, and perfusion imaging in the classification of brain tumors using machine learning scheme Abstract Purpose While MRI is the modality of choice for the assessment of patients with brain tumors, differentiation between various tumors based on their imaging characteristics might be challenging due to overlapping imaging features. The purpose of this study was to apply a machine learning scheme using basic and advanced MR sequences for distinguishing different types of brain tumors. Methods The study cohort included 141 patients (41 glioblastoma, 38 metastasis, 50 meningioma, and 12 primary central nervous system lymphoma). A computer-assisted classification scheme, combining morphologic MRI, perfusion MRI, and DTI metrics, was developed and used for tumor classification. The proposed multistep scheme consists of pre-processing, ROI definition, features extraction, feature selection, and classification. Feature subset selection was performed using support vector machines (SVMs). Classification performance was assessed by leave-one-out cross-validation. Given an ROI, the entire classification process was done automatically via computer and without any human intervention. Results A binary hierarchical classification tree was chosen. In the first step, selected features were chosen for distinguishing glioblastoma from the remaining three classes, followed by separation of meningioma from metastasis and PCNSL, and then to discriminate PCNSL from metastasis. The binary SVM classification accuracy, sensitivity and specificity for glioblastoma, metastasis, meningiomas, and primary central nervous system lymphoma were 95.7, 81.6, and 91.2%; 92.7, 95.1, and 93.6%; 97, 90.8, and 58.3%; and 91.5, 90, and 96.9%, respectively. Conclusion A machine learning scheme using data from anatomical and advanced MRI sequences resulted in high-performance automatic tumor classification algorithm. Such a scheme can be integrated into clinical decision support systems to optimize tumor classification.

Endovascular recanalization for symptomatic subacute and chronic intracranial large artery occlusion of the anterior circulation: initial experience and technical considerations Abstract Purpose This study aimed to report the clinical findings and initial clinical experience of endovascular recanalization for symptomatic subacute/chronic intracranial large artery occlusion (ILAO) of the anterior circulation. Methods From October 2015 to December 2017, 13 patients with symptomatic subacute/chronic ILAO of the anterior circulation were enrolled in this study and underwent endovascular recanalization. We collected the initial procedural results, including the rate of successful recanalization and periprocedural complications, and data pertaining to angiographic and clinical follow-up. Results Recanalization was successful in 11 of 13 patients (84.6%). Intraoperative complications occurred in four cases, including symptomatic distal embolism in three cases; one of which was simultaneously complicated with artery dissection. Intracerebral hemorrhage occurred in one case. Eleven patients underwent angiographic follow-up, and 12 patients underwent clinical follow-up. The results of the angiography follow-up (mean 6 ± 3.29 months) showed that in-stent restenosis occurred in one of the 11 successfully recanalized patients. However, the artery was occluded again in the patient who achieved thrombolysis in cerebral infarction (TICI) grade of 2a after treatment. Clinical follow-up (mean 5.8 ± 2.25 months) showed no recurrence of transient ischemic attack (TIA) or stroke in ten successfully recanalized cases. However, the patient who developed in-stent stenosis suffered TIA. Conclusions Endovascular recanalization for symptomatic subacute/chronic ILAO of anterior circulation is feasible, relatively safe, and efficacious in highly selected cases, improving patients' symptoms in the short-term. However, further larger scale pilot studies are needed to determine the efficacy and long-term outcome associated with this treatment.

Dependency of the blood oxygen level dependent-response to hyperoxic challenges on the order of gas administration in intracranial malignancies Abstract Purpose Literature reports contradicting results on the response of brain tumors to vascular stimuli measured in T2*-weighted MRI. Here, we analyzed the potential dependency of the MRI-response to (hypercapnic) hyperoxia on the order of the gas administration. Methods T2* values were quantified at 3 Tesla in eight consenting patients at rest and during inhalation of hyperoxic/hypercapnic gas mixtures. Patients were randomly divided into two groups undergoing different gas administration protocols (group A: medical air-pure oxygen–carbogen; group B: medical air–carbogen-pure oxygen). Mann-Whitney U test and Wilcoxon signed rank test have been used to proof differences in T2* regarding respiratory challenge or different groups, respectively. Results T2* values at rest for gray and white matter were 50.3 ± 2.6 ms and 46.1 ± 2.0 ms, respectively, and slightly increased during challenge. In tumor areas, T2* at rest were: necrosis = 74.1 ± 10.1 ms; edema = 60.3 ± 17.6 ms; contrast-enhancing lesions = 48.6 ± 20.7 ms; and solid T2-hyperintense lesions = 45.0 ± 3.0 ms. Contrast-enhancing lesions strongly responded to oxygen (+ 20.7%) regardless on the gas protocol (p = 0.482). However, the response to carbogen significantly depended on the order of gas administration (group A, + 18.6%; group B, − 6.4%, p = 0.042). In edemas, a different trend between group was found when breathing oxygen (group A, − 9.9%; group B, + 19.5%, p = 0.057). Conclusion Preliminary results show a dependency of the T2* response of contrast-enhancing brain tumor lesions on the order of the gas administration. The gas administration protocol is an important factor in the interpretation of the T2*-response in areas of abnormal vascular growth.

Micro-computed tomography: a new diagnostic tool in postmortem assessment of brain anatomy in small fetuses Abstract Purpose The aim of our study was to evaluate the postmortem micro-CT anatomy of early fetal human fetal brains, either in situ or isolated. Methods We studied 12 ex vivo specimens, 9 whole human fetuses (9–18 GW), and 3 isolated samples (16–26 GW). Specimens were fixed in formalin, then immersed in Lugol solution. Images were evaluated by two neuroradiologists. The depiction of CNS structures was defined based on the comparison between micro-CT images and a reference histologic anatomical Atlas of human brain development. Results Micro-CT provided informative high-resolution brain images in all cases, with the exception of one case (9 weeks) due to advanced maceration. All major CNS structures (i.e., brain hemispheres, layering, ventricles, germinal neuroepithelium, basal ganglia, corpus callosum, major cranial nerves, and structures of the head and neck) were recognizable. Conclusions Micro-CT imaging of the early fetal brain is feasible and provides high-quality images that correlate with the histological Atlas of the human brain, offering multiplanar and volumetric images that can be stored and shared for clinical, teaching, and research purposes.

Dynamic cerebellar herniation in Chiari patients during the cardiac cycle evaluated by dynamic magnetic resonance imaging Abstract Purpose Cerebellar herniation in Chiari patients can be dynamic, following the cerebrospinal fluid pulsatility during the cardiac cycle. We present a voxel intensity distribution method (VIDM) to automatically extract the pulsatility-dependent herniation in time-resolved MRI (CINE MRI) and compare it to the simple linear measurements. The degree of herniation is furthermore compared on CINE and static sequences, and the cerebellar movement is correlated to the presence of hydrocephalus and syringomyelia. Methods The cerebellar movement in 27 Chiari patients is analyzed with VIDM and the results were compared to linear measurements on an image viewer (visual inspection, VI) using a paired t test. Second, an ANOVA test is applied to compare the degree of herniation on static 3D MRI and CINE. Finally, the Pearson's correlation coefficient is calculated for the correlation between cerebellar movement and the presence of hydrocephalus and syringomyelia. Results VIDM showed significant movement in 85% of our patients. Assuming that movement < 1 mm cannot be detected reliably on an image viewer, VI identified movement in 29.6% of the patients (p = 0.002). The herniation was greater on static sequences than on CINE in most cases, but this was not statistically significant. The cerebellar movement was not correlated with hydrocephalus or syringomyelia (Pearson's coefficient < 0.3). Conclusions VIDM is a sensitive method to detect tissue movement on CINE MRI and could be used for Chiari patients, but also for the evaluation of cyst membranes, ventriculostomies, etc. The cerebellar movement appears not to correlate with hydrocephalus and syringomyelia in Chiari patients.

Altered brain diffusion tensor imaging indices in adolescents with the Fontan palliation Abstract Purpose Single ventricle heart disease (SVHD) patients show injury in brain sites that regulate autonomic, mood, and cognitive functions. However, the nature (acute or chronic changes) and extent of brain injury in SVHD are unclear. Our aim was to examine regional brain tissue damage in SVHD over controls using DTI-based mean diffusivity (MD), axial diffusivity (AD), radial diffusivity (RD), and fractional anisotropy (FA) procedures. Methods We collected two DTI series (3.0-T MRI), mood and cognitive data, from 27 SVHD and 35 control adolescents. Whole-brain MD, AD, RD, and FA maps were calculated from each series, realigned and averaged, normalized to a common space, smoothed, and compared between groups using ANCOVA (covariates, age and sex; false discovery rate, p < 0.05). Region-of-interest analyses were performed to calculate MD, AD, RD, and FA values for magnitude assessment between groups. Results SVHD patients showed impaired mood and cognitive functions over healthy adolescents. Multiple brain sites in SVHD showed increased MD values, including the insula, caudate, cingulate, hypothalamus, thalamus, medial prefrontal and frontal cortices, parahippocampal gyrus, hippocampus, precentral gyrus, amygdala, cerebellum, corpus callosum, basal forebrain, mammillary bodies, internal capsule, midbrain, fornix, and occipital, parietal, and temporal cortices, indicating chronic tissue changes. Similar areas showed either increased AD or RD values, with RD changes more enhanced over AD in SVHD compared to controls. Few brain regions emerged with increased or decreased FA values in SVHD patients over controls. Conclusion SVHD adolescents, more than a decade from their last surgical procedure, show widespread brain abnormalities in autonomic, mood, and cognitive regulatory areas. These findings indicate that brain injury is in a chronic stage in SVHD with predominantly myelin changes that may result from previous hypoxia/ischemia- or developmental-induced processes.

Meningeal enhancement depicted by magnetic resonance imaging in tumor patients: neoplastic meningitis or therapy-related enhancement? Abstract Purpose To assess the prevalence of false-positive meningeal contrast enhancement in patients with solid tumors who were undergoing chemotherapy. Methods A total of 2572 magnetic resonance imaging (MRI) examinations of the brain were retrospectively evaluated by two readers for the presence of pathological meningeal contrast enhancement conspicuous for neoplastic meningitis. These patients either had malignant melanoma, breast or lung cancer, or lymphoma. The reference standards were cerebrospinal fluid cytology results and follow-up MRI. In cases with pathological contrast enhancement that decreased upon follow-up and non-malignant cytology, the enhancement pattern was further described as pial or dural, local or diffuse, or supra- or infra-tentorial. Moreover, the underlying therapy regimes were assessed. Results The final study cohort included 78 patients (51 females, median age 57 years), of which 11 patients (14.1%) had a repeated non-malignant cytology ('pseudomeningeosis'). In one case, this finding, a granular pleocytosis, was attributed to previous radiotherapy. Of the remaining patients, seven were receiving multimodal, immunotherapy-based therapy regimens. Patients with unsuspicious cytology had a predominantly supratentorial distribution pattern in comparison to patients with neoplastic meningitis. Conclusions The overall prevalence of the presence of false-positive meningeal contrast enhancement is low (< 1%) and not associated with specific imaging patterns. We hypothesize that there is a possible relationship between immunotherapy and 'pseudomeningeosis'. Therefore, in all cases with suspected neoplastic meningitis, the cerebrospinal fluid should be analyzed to confirm the diagnosis, especially in patients undergoing immunotherapy.

Dual-energy CT angiography-derived virtual non-contrast images for follow-up of patients with surgically clipped aneurysms: a retrospective study Abstract Purpose To evaluate the diagnostic performance, image quality, artifacts, and radiation doses of virtual non-contrast (VNC) images, relative to true non-contrast (TNC) images, in patients with surgically clipped aneurysms. Methods Seventy-six patients who underwent unenhanced computed tomography (CT) and dual-energy (DE)-CT angiography after surgical clipping of 85 intracranial aneurysms were included in the study. Diagnostic performances of VNC and TNC images were compared with respect to sensitivity, specificity, and positive and negative predictive values. The results of quantitative and qualitative analyses were compared between VNC and TNC images. Radiation doses were also compared between VNC and TNC images. Results Diagnostic performance of VNC images was lower than that of TNC images; however, addition of contrast-enhanced images improved diagnostic performance. Image noise and mean attenuation of VNC images were significantly higher than those of TNC images in the centrum semiovale, cerebellum, and pons. The quality of VNC images was excellent or sufficient (85.5% for observer 1, 85.5% for observer 2), and complete acceptance of VNC images was achieved in 64.5% for observer 1 and in 71.0% for observer 2; however, the addition of contrast-enhanced images increased the level of acceptance (92.0% for observer 1, 90.9% for observer 2). Clip artifacts were significantly lower in VNC images than in TNC images. CT dose index, dose-length product, and effective dose were significantly lower without TNC images. Conclusion VNC images showed lower diagnostic performance and image quality, and higher image noise than TNC images; however, VNC images could reduce clip artifacts and radiation doses.

Alexandros Sfakianakis
Anapafseos 5 . Agios Nikolaos
Crete.Greece.72100
2841026182

6948891480

alsfakia@gmail.com

Physiological Sciences

Chronic water insufficiency induced kidney damage and energy dysregulation despite reduced food intake, which improved gut microbiota in female rats Abstract Water intake is recommended for weight loss, but the relationship between water intake and energy metabolism is not clear. We hypothesized that long-term water insufficiency would influence energy, glucose, and lipid metabolism while modulating gut microbiota. Female rats were provided with high-fat diets with different amounts of water and food intake for 6 weeks as follows: water provided for 1 h per day with food ad libitum (WRFA), water supply ad libitum plus pair feeding of with water restricted rats(WAFR), water restriction with ad libitum food for 3 weeks and water and food intake ad libitum for 3 weeks (WR-WA) and ad libitum supply of water and food (WAFA). Water intake in WRFA was about one-third of WAFR and WAFA, whereas food intake was lowered by 30% in WRFA and WAFR than WAFA. Body fat decreased in WRFA and WAFR, but WAFR decreased fat mass more than WRFA. Energy expenditure was lower in WRFA than WAFA and carbohydrate utilization was much higher in WRFA than the other groups. The peak serum glucose concentrations were lower in WAFA than the other groups and WRFA lowered serum insulin levels more than WAFA during OGTT. WRFA shrank the glomerulus with increased apoptotic cells and damaged renal tubules compared to the WAFA and WAFR. WR-WA also exhibited greater glomerular shrinkage and apoptosis that WAFA, but not as much WRFA, indicating that the kidneys were healing after water restriction damage. WRFA exacerbated dyslipidemia compared to the WAFA and WAFR groups. The gut microbiome was similarly modulated in WRFA and WAFR, compared to WAFA, but it was mainly affected by food intake, not water restriction in the host. WRFA and WAFR increased Bacteroidetes and decreased Firmicutes compared WAFA. In conclusion, chronic insufficient water intake induced renal damage, decreased energy expenditure, and exacerbated dyslipidemia in rats with reduced food intake. However, the reduction of food intake improved gut microbiome regardless of insufficient water intake and only minor effects on the microbiome were observed due to water restriction.

Natural silibinin modulates amyloid precursor protein processing and amyloid-β protein clearance in APP/PS1 mice Abstract Silibinin has been shown to attenuate cognitive dysfunction and inhibit amyloid-beta (Aβ) aggregation in Alzheimer's disease (AD) models. However, the underlying mechanism by which silibinin improves cognition remains poorly understood. In this study, we investigated the effect of silibinin on β-secretase levels, Aβ enzymatic degradation, and oxidative stress in the brains of APP/PS1 mice with cognitive impairments. Oral administration of silibinin for 2 months significantly attenuated the cognitive deficits of APP/PS1 mice in the Y-maze test, novel object recognition test, and Morris water maze test. Biochemical analyses revealed that silibinin decreased Aβ deposition and the levels of soluble Aβ1-40/1-42 in the hippocampus by downregulating APP and BACE1 and upregulating NEP in APP/PS1 mice. In addition, silibinin decreased the MDA content and increased the activities of the antioxidant enzymes CAT, SOD, and NO. Based on our findings, silibinin is a potentially promising agent for preventing AD-associated Aβ pathology.

Mild hyperbaric oxygen: mechanisms and effects Abstract Adequate oxygen supply by exposure to mild hyperbaric oxygen at appropriately high atmospheric pressure (1266–1317 hPa) and increased oxygen concentration (35–40% oxygen) has a possibility of improving the oxidative metabolism in cells and tissues without barotrauma and excessive production of reactive oxygen species. Therefore, metabolic syndrome and lifestyle-related diseases, including type 2 diabetes and hypertension, in rats were inhibited and/or improved by exposure to mild hyperbaric oxygen. It accelerated the growth-induced increase in oxidative capacity of the skeletal muscle in rats and inhibited the age-related decrease in oxidative capacity of the skeletal muscle in mice. A decrease in dopaminergic neurons in the substantia nigra of mice with Parkinson's disease was inhibited by exposure to mild hyperbaric oxygen. This review describes the beneficial effects of exposure to mild hyperbaric oxygen on some metabolic diseases and their perspectives.

Correlations between "hie-sho" interview score and progesterone, fat intake, and Kupperman index in pre- and post-menopausal women: a pilot study Abstract Japanese menopausal women who feel cold, even in a warm room, are said to be experiencing "hie-sho." We assessed the magnitude of coldness by a "hie-sho" interview score. The association between the magnitude of coldness and female hormones, fat intake, and menopausal symptoms is unknown. The aim of the present study was to elucidate the relationship between the hie-sho interview scores and female hormones, fat intake, Kupperman index in pre- (pre group) and post- (post group) menopausal women. The hie-sho interview scores, Kupperman index questionnaire results, dietary survey to analyze fat intake, and body weight were analyzed, and plasma estradiol, progesterone, and lipid levels were measured in the subjects in the pre (n = 9) and post (n = 11) groups. Plasma female hormones and fat intake were different, but the total Kupperman index was not different between pre and post groups. Plasma progesterone was positively correlated with the hie-sho score only in the post group. Plasma triglyceride was positively correlated with the hie-sho score only in the pre group. Intake of cholesterol, arachidonic acid, and docosapentaenoic acid was negatively correlated with the hie-sho score only in the pre group. The positive correlation between total Kupperman index and hie-sho score was observed only in the pre group. These results indicated that progesterone level was related to coldness in post-menopausal women. Fat intake, plasma triglyceride, and menopausal symptoms may be related to coldness in pre-menopausal women.

Muscle sympathetic nerve activity during exercise Abstract Appropriate cardiovascular adjustment is necessary to meet the metabolic demands of working skeletal muscle during exercise. The sympathetic nervous system plays a crucial role in the regulation of arterial blood pressure and blood flow during exercise, and several important neural mechanisms are responsible for changes in sympathetic vasomotor outflow. Changes in sympathetic vasomotor outflow (i.e., muscle sympathetic nerve activity: MSNA) in inactive muscles during exercise differ depending on the exercise mode (static or dynamic), intensity, duration, and various environmental conditions (e.g., hot and cold environments or hypoxic). In 1991, Seals and Victor [6] reviewed MSNA responses to static and dynamic exercise with small muscle mass. This review provides an updated comprehensive overview on the MSNA response to exercise including large-muscle, dynamic leg exercise, e.g., two-legged cycling, and its regulatory mechanisms in healthy humans.

Tabata training: one of the most energetically effective high-intensity intermittent training methods Abstract For decades, high-intensity interval/intermittent exercise training methods have been used by elite athletes to improve their performance in sports. One of the most effective training methods, i.e., 'Tabata training,' is reviewed herein from the viewpoint of the energetics of exercise. The prior research describing the metabolic profile and effects of Tabata training is also summarized, with some historical anecdotes.

Renal involvement in the pathogenesis of mineral and bone disorder in dystrophin-deficient mdx mouse Abstract Duchenne muscular dystrophy is a severe muscular disorder, often complicated with osteoporosis, and impaired renal function has recently been featured. We aimed to clarify the involvement of renal function in the pathogenesis of mineral and bone disorder in mdx mice, a murine model of the disease. We clearly revealed renal dysfunction in adult mdx mice, in which dehydration and hypercalcemia were contributed. We also examined the effects of dietary phosphorus (P) overload on phosphate metabolism. Serum phosphate and parathyroid hormone (PTH) levels were significantly increased in mdx mice by dietary P in a dose-dependent manner; however, bone alkaline phosphatase levels were significantly lower in mdx mice. Additionally, bone mineral density in mdx mice were even worsened by increased dietary P in a dose-dependent manner. These results suggested that the uncoupling of bone formation and resorption was enhanced by skeletal resistance to PTH due to renal failure in mdx mice.

Renin–angiotensin system research: from molecules to the whole body Abstract Hypertension is one of the most important risk factors and a leading cause of death from cardiovascular and cerebrovascular diseases. Based on numerous previous studies, hypertension is thought to be caused by the complex mutual interactions of genetic factors and environmental factors, such as excessive salt intake and stress. However, its detailed mechanisms are not yet clearly understood. The renin–angiotensin system (RAS) is a key hormonal system in the pathogenesis of hypertension. New knowledge is still accruing on this cascade, even after more than 120 years since the discovery of renin. To clarify the molecular mechanisms of RAS in vivo, we created transgenic mice with chronic hypertension. These mice carry the human genes encoding renin, a hypertensive enzyme, and its substrate angiotensinogen. Hypotensive mice homozygous for a targeted disruption of the angiotensinogen gene were also created. This review presents our 47-year history of RAS research.

Sodium-coupled monocarboxylate transporter 1 interacts with the RING finger- and PDZ domain-containing protein PDZRN3 Abstract Sodium-coupled monocarboxylate transporter SMCT1 (SLC5A8) mediates monocarboxylate transport in the proximal tubule of the kidney. We have identified PDZK1 and PDZ domain-containing RING finger 3 (PDZRN3) as potent binding partners of SMCT1, which has a PDZ motif (Thr–Arg–Leu), by yeast two-hybrid screening and revealed that PDZK1 enhances the transport activity of SMCT1. In this study, we aimed to characterize the interaction between SMCT1 and PDZRN3 as well as to examine how PDZRN3 regulates SMCT1 function. An interaction between SMCT1 and PDZRN3 through the PDZ motif was observed in a co-immunoprecipitation assay and yeast two-hybrid assay. A transport assay showed that PDZRN3 abolished the enhancing effect of PDZK1 on nicotinate uptake via SMCT1. Our results suggest that SMCT1 interacts with PDZRN3 and that PDZRN3 may regulate SMCT1 function by interfering with the interaction between SMCT1 and PDZK1.

Enhancement of electroencephalogram activity in the theta-band range during unmatched olfactory-taste stimulation Abstract The aim of this study was to investigate how odor stimulation affects taste perception. Electroencephalogram (EEG) signals were measured from the frontal region of the head in normal, healthy subjects, and frequency analyses were performed. Each odor stimulation was delivered while the subject was tasting chocolate, using chocolate paste as the odorant for 'matched odor stimulation,' and garlic paste for 'unmatched odor stimulation.' Differences in EEG signals appeared between the matched and unmatched arms of the study. Comparison of the frequencies of EEGs captured under the condition of unmatched odor stimulation with those captured under the condition of matched odor stimulation showed that the occupancy rate of the theta-frequency band under the condition of unmatched odor stimulation was higher than that under the condition of matched odor stimulation. Interestingly, a negative correlation existed between the occupancy rate of the theta-frequency band and the subjective feeling of chocolate sweetness. The present findings suggest that when humans receive odors that do not match with the foods being consumed, subjective feelings are disturbed and theta-band brain activity is increased while the unmatched information is cross-checked.

Alexandros Sfakianakis
Anapafseos 5 . Agios Nikolaos
Crete.Greece.72100
2841026182

6948891480

alsfakia@gmail.com

Vegetation History and Archaeobotany

Correction to: Small things can make a big difference: a comparison of pollen and macrobotanical records of some food plants from medieval and post-medieval cesspits in the Netherlands and northern Belgium In the original publication, the caption of Fig. 1 is partly missing. It has been updated in this correction.

Small things can make a big difference: a comparison of pollen and macrobotanical records of some food plants from medieval and post-medieval cesspits in the Netherlands and northern Belgium Abstract This paper presents a review of records of pollen and botanical macroremains of a selection of food plants from late and post-medieval cesspits (12th century-19th century ad) in the Netherlands and northern Belgium. The presented data demonstrate that several food plants remain largely invisible in the macrobotanical records. These are all plants from which the flowers or flower buds (Borago officinalis, Capparis, Carthamus tinctorius, Crocus sativus, Syzygium aromaticum) or leaves (Anthriscus cerefolium, Spinacia oleracea) are eaten, or that are typical components of honey (Cistus). As a result, little is known about the import or local production and consumption of these food plants in these times. This review now shows that past use of some of these plants is reflected in the pollen assemblages of (post-) medieval cesspits. For the first time, a large archaeobotanical dataset is presented, including pollen, providing information on the past use of these plants between the 12th and 19th century ad in the Netherlands and Belgium.

Palynology of Iron Age and Gallo-Roman archaeological sediments from Beaurieux Les Grèves, Aisne, France Abstract Pollen and non-pollen palynomorph analyses conducted on archaeological sediments from ditches and pits are used to investigate land-use and vegetation history around the long-lived rural settlement of Beaurieux Les Grèves in the Aisne valley, Picardy, northern France. Samples were examined from successive phases spanning the 8th century bc to the 3rd century ad. Each phase showed evidence of agriculture and human impact on the environment. The earliest occupation was during the early Iron Age, and the area around the site was shown to have been almost completely deforested and mainly pasture. The later Iron Age supported mixed agriculture, probably with pasture around the settlement site and cereal fields further away. In Gallo-Roman times there was mixed farming with considerable cereal cultivation, although the area near the site was mainly pasture or abandoned land. Late Gallo-Roman times saw a reduction in agricultural activity and some regeneration of heath and scrub vegetation. Coprophilous fungal spores are conspicuous and indicate the importance of animal husbandry on and around the site. The data from Beaurieux Les Grèves augment earlier analyses from settlements and natural deposits in the Aisne valley in suggesting almost complete deforestation for agricultural land use within the valley bottom, although some woodland undoubtedly survived on the steeper valley sides and nearby plateaux. It agrees with the pollen evidence from the wider region of north-eastern France that shows clearance of Fagus-dominated woodland for mixed agriculture in late Iron Age and Gallo-Roman times, and increasingly for arable cultivation. This was responsible for effecting a transformation from a partly wooded to a mainly open landscape.

Distribution and vegetation representation of pollen assemblages from surface sediments of Nam Co, a large alpine lake in the central Tibetan Plateau Abstract Lacustrine fossil pollen records have been widely used to reconstruct palaeovegetation and palaeoclimate changes on the Tibetan Plateau (TP). However, little is known about the vegetation representation of modern lacustrine pollen assemblages there. This paper presents the results of modern pollen investigation on 63 surface sediments from the lake basin and 37 topsoil samples from the drainage area of a large lake, Nam Co, located in the central TP. It aims to assess quantitatively the influences on lacustrine pollen assemblages of the pollen sources and sedimentary processes, and to establish vegetation representations for modern lacustrine pollen assemblages. Modern pollen assemblages from topsoils of different vegetation had diagnostic features in terms of their composition and pollen percentage. The spatial variabilities and results of principal component analysis suggested that lacustrine pollen assemblages were influenced by both the regional/local source vegetation and sedimentary processes. The lacustrine pollen assemblages were mainly homogeneous due to in-lake sedimentary processes (mixing and redistribution). An accumulation zone for lacustrine pollen assemblages was found in the deep lake basin (depth > 60 m) due to sediment focusing. The results of boosted regression tree analysis further confirmed that source vegetation was the predominant factor (85.8%) responsible for the vegetation representation of lacustrine pollen assemblages, while sedimentary processes accounted for only 14.2%. The results of discriminant analysis indicated that most lacustrine pollen assemblages (90.5%) were representative for the regional vegetation of alpine steppe in the Nam Co catchment and central TP, while only 9.5% were representative for the local meadow vegetation. Therefore, it is recommended that lacustrine pollen assemblages from deep lake basin of accumulation zone in large lakes of the TP can be used to retrieve efficiently the signals from regional vegetation and climate changes.

The native status of Pinus pinaster on serpentine soils: charcoal analysis and palaeoenvironmental history in Sierra Bermeja (southern Iberian Peninsula, Spain) Abstract Pinus pinaster (maritime pine), a conifer native to the western Mediterranean, has a broad distribution, occupying a wide variety of habitats. On certain substrata such as ultramafic (ultrabasic) rock, the indigenous nature of this conifer has traditionally been questioned by the scientific community, which has regarded it as an introduction. In Sierra Bermeja, mountains forming the largest ultramafic outcrop in western Europe, the dominant woodland formations on serpentine soils are P. pinaster and Abies pinsapo. However the variable presence, albeit isolated, of various species of arboreal Quercus and the frequent forestry plantation of P. pinaster in recent centuries have led to broad-leaved woods being generally considered as the dominant natural communities in this mountain range, so marginalizing the role of these conifers. In an attempt to settle this scientific controversy, we have carried out soil charcoal analyses from seven localities in Sierra Bermeja. The palaeoecological data we have gathered show that P. pinaster has a natural status and has been present in this mountain range during a large part of the Holocene before the changes to its natural landscape by human activities. These results are of great importance for the management and conservation of rare serpentine ecosystems.

Complex responses of vegetation diversity to Holocene climate change in the eastern Tibetan Plateau Abstract Biodiversity has attracted much attention recently due to its important relationships with ecosystem function under various global warming scenarios. However, an understanding of biodiversity mechanisms requires study over long time scales. Three high-resolution pollen records of Zoige Basin in the eastern Tibetan Plateau are used to reconstruct changes in vegetation diversity during the Holocene, allowing the mechanisms that drove the dynamic to be quantitatively explored. Rarefaction and Hill's indices are used to estimate the diversity richness and evenness based on pollen data. The results show that changes in palynological richness can be divided into five stages: an abnormal change from 10,500 to 9,000 cal bp, an obvious increase from 9,000 to 6,500 cal bp, a decreasing trend from 6,500 to 4,000 cal bp, an increasing trend after 4,000 cal bp until 1,500 cal bp, and a highly fluctuating stage from 1,500 cal bp to the present. Palynological evenness is relatively stable throughout the Holocene except for during a briefly elevated period from ca. 4,000 to 1,500 cal bp. The result of Boosted Regression Tree analysis indicates that climate is the main driving factor and the effect of temperature is stronger than that of precipitation in the study region. However, during ca. 4,000–1,500 cal bp, palynological diversity is primarily affected by vegetation structure, as shown by an increase in palynological evenness, which can in turn be explained by the climate threshold theory. This research provides a long-term, high-resolution reconstruction of palynological diversity which could be used to infer vegetation diversity change in the ecologically sensitive Tibetan Plateau. The results imply that vegetation diversity in the region may increase under global warming if human impacts are not considered.

Methods of acquisition and use of firewood among hunter-gatherer groups in Patagonia (Argentina) during the Holocene Abstract The present article examines the ways of obtaining firewood and of using it by Patagonian hunter-gatherer groups and the relationship with their high mobility. With these goals, we have selected a range of archaeological sites in varied types of vegetation: forest, forest-steppe ecotone and steppe (according to pollen reconstructions and current records) in several different latitudes of Argentinean Patagonia: Paredón Lanfré (Río Negro province); Cerro Pintado (Chubut province); Cerro Casa de Piedra 7 and Orejas de Burro 1 (Santa Cruz province). The taxa, including Nothofagus pumilio, Austrocedrus chilensis, Ribes magellanicum, Embothrium coccineum and Fabiana imbricata, found among the scattered charcoal remains in the sediments of the four Patagonian sites, show how firewood was gathered in types of vegetation similar to the ones that nowadays surround the archaeological sites. The archaeobotanical results allow us to detect differences and similarities of the supply of wood and its relationship with human mobility, site functionality and the types of occupation.

Re-analysis of archaeobotanical remains from pre- and early agricultural sites provides no evidence for a narrowing of the wild plant food spectrum during the origins of agriculture in southwest Asia Abstract Archaeobotanical evidence from southwest Asia is often interpreted as showing that the spectrum of wild plant foods narrowed during the origins of agriculture, but it has long been acknowledged that the recognition of wild plants as foods is problematic. Here, we systematically combine compositional and contextual evidence to recognise the wild plants for which there is strong evidence of their deliberate collection as food at pre-agricultural and early agricultural sites across southwest Asia. Through sample-by-sample analysis of archaeobotanical remains, a robust link is established between the archaeological evidence and its interpretation in terms of food use, which permits a re-evaluation of the evidence for the exploitation of a broad spectrum of wild plant foods at pre-agricultural sites, and the extent to which this changed during the development of early agriculture. Our results show that relatively few of the wild taxa found at pre- and early agricultural sites can be confidently recognised as contributing to the human diet, and we found no evidence for a narrowing of the plant food spectrum during the adoption of agriculture. This has implications for how we understand the processes leading to the domestication of crops, and points towards a mutualistic relationship between people and plants as a driving force during the development of agriculture.

Comparison of a recent elm decline with the mid-Holocene Elm Decline Abstract The threat of devastating pathogens on a range of tree species has increased recently, which justifies the investigation of the temporal dynamics of tree declines caused by disease. The mid-Holocene Elm Decline was a widespread and synchronous event recorded in pollen diagrams across northwest Europe. The probable drivers of this event have been debated for over 80 years but the role of disease has gathered greatest support in recent decades. Here we report pollen data at sub-decadal resolution from a closely sampled sediment core from eastern Ireland covering the last 160 years. This provides data on the structure and dynamics of the surrounding woodland before, during and after the outbreak of Dutch elm disease in the 1970s. A decline in elm was also recorded in the 1940s associated with an earlier, less virulent, outbreak of elm disease. The elm population recovered from the 1940s outbreak after 20 years but did not recover from the 1970s pandemic. The pollen data also facilitate the comparison of this disease-mediated decline with the mid-Holocene Elm Decline; the impact of human disturbance and disease resulted in different woodland compositions and the adjustment of the woodland in response to these factors was rapid. The impact of Dutch elm disease on this elm 1970s population was very similar to that recorded in the mid-Holocene Elm Decline in Britain and Ireland.

20,000 years of interactions between climate, vegetation and land use in Northern Greece Abstract Detailed knowledge about the history of vegetation, fire and land use is scarce in Northern Greece. We analysed lake sediments from Limni Zazari (Northern Greece) to reconstruct the past local vegetation and fire history with a special focus on land use and its impacts on erosion and lake eutrophication. Our data suggest a rather dense steppic vegetation after ca 20,000 cal bp (18050 cal bc). Forest expansion with Pinus sylvestris and admixed Quercus pubescens started around 14,500 cal bp (12550 cal bc). After the onset of the Holocene, mixed deciduous sub-mediterranean oak forests expanded, accompanied by rapidly decreasing soil erosion rates and increasing aquatic biological productivity. Pollen of cereals and Plantago lanceolata suggests continuous farming activities in the region after 8,200 cal bp (6250 cal bc), in agreement with archaeological evidence. Fairly closed mixed pine-oak forests dominated the landscape until ca 3,500 cal bp (1550 cal bc) that were only temporarily reduced during the Neolithic around 7,100 and 6,500 cal bp (5150 and 4550 cal bc). Land cover changes and aquatic biogeochemistry were closely linked during this period. Forest phases corresponded to lake eutrophication and hypolimnetic anoxia (meromixis), whereas during periods of deforestation (e.g. around 8,200 cal bp/6250 cal bc) soil erosion rates and lake mixing increased, while aquatic productivity decreased. After 3,500 cal bp (1550 cal bc) humans disrupted forests and open land vegetation expanded (e.g. Artemisia, Rumex-type, Cichorioideae, Chenopodiaceae). With the onset of the Iron Age (ca. 3,050 cal bp/1100 cal bc) grassland communities expanded massively and pine-oak forests gradually declined. Anthropogenic pressure on forests increased even more during the past 500 years. Finally, forest recovery during the recent decades led to decreased erosion and increased lake productivity. We conclude that over the millennia, intense pastoral and arable activities shaped both aquatic and terrestrial environments, ultimately creating a humanized vegetation mosaic in which the original natural mixed deciduous oak forests only form relict stands. Future climate warming and decreasing anthropogenic pressure may release a rapid spread of mixed deciduous oak forests around Limni Zazari.

Alexandros Sfakianakis
Anapafseos 5 . Agios Nikolaos
Crete.Greece.72100
2841026182

6948891480

alsfakia@gmail.com