The genetic legacy of the expansion of Bantu-speaking peoples in Africa.

The expansion of people speaking Bantu languages is the most dramatic demographic event in Late Holocene Africa and fundamentally reshaped the linguistic, cultural and biological landscape of the continent1-7. With a comprehensive genomic dataset, including newly generated data of modern-day and ancient DNA from previously unsampled regions in Africa, we contribute insights into this expansion that started 6,000-4,000 years ago in western Africa. We genotyped 1,763 participants, including 1,526 Bantu speakers from 147 populations across 14 African countries, and generated whole-genome sequences from 12 Late Iron Age individuals8. We show that genetic diversity amongst Bantu-speaking populations declines with distance from western Africa, with current-day Zambia and the Democratic Republic of Congo as possible crossroads of interaction. Using spatially explicit methods9 and correlating genetic, linguistic and geographical data, we provide cross-disciplinary support for a serial-founder migration model. We further show that Bantu speakers received significant gene flow from local groups in regions they expanded into. Our genetic dataset provides an exhaustive modern-day African comparative dataset for ancient DNA studies10 and will be important to a wide range of disciplines from science and humanities, as well as to the medical sector studying human genetic variation and health in African and African-descendant populations.

A Jovian analogue orbiting a white dwarf star.

Studies1,2 have shown that the remnants of destroyed planets and debris-disk planetesimals can survive the volatile evolution of their host stars into white dwarfs3,4, but few intact planetary bodies around white dwarfs have been detected5-8. Simulations predict9-11 that planets in Jupiter-like orbits around stars of ≲8 M☉ (solar mass) avoid being destroyed by the strong tidal forces of their stellar host, but as yet, there has been no observational confirmation of such a survivor. Here we report the non-detection of a main-sequence lens star in the microlensing event MOA-2010-BLG-477Lb12 using near-infrared observations from the Keck Observatory. We determine that this system contains a 0.53 ± 0.11 M☉ white-dwarf host orbited by a 1.4 ± 0.3 Jupiter-mass planet with a separation on the plane of the sky of 2.8 ± 0.5 astronomical units, which implies a semi-major axis larger than this. This system is evidence that planets around white dwarfs can survive the giant and asymptotic giant phases of their host's evolution, and supports the prediction that more than half of white dwarfs have Jovian planetary companions13. Located at approximately 2.0 kiloparsecs towards the centre of our Galaxy, it is likely to represent an analogue to the end stages of the Sun and Jupiter in our own Solar System.

High-resolution structural information of membrane-bound α-synuclein provides insight into the MoA of the anti-Parkinson drug UCB0599.

α-synuclein (αS) is an intrinsically disordered protein whose functional ambivalence and protein structural plasticity are iconic. Coordinated protein recruitment ensures proper vesicle dynamics at the synaptic cleft, while deregulated oligomerization on cellular membranes contributes to cell damage and Parkinson's disease (PD). Despite the protein's pathophysiological relevance, structural knowledge is limited. Here, we employ NMR spectroscopy and chemical cross-link mass spectrometry on 14N/15N-labeled αS mixtures to provide for the first time high-resolution structural information of the membrane-bound oligomeric state of αS and demonstrate that in this state, αS samples a surprisingly small conformational space. Interestingly, the study locates familial Parkinson's disease mutants at the interface between individual αS monomers and reveals different oligomerization processes depending on whether oligomerization occurs on the same membrane surface (cis) or between αS initially attached to different membrane particles (trans). The explanatory power of the obtained high-resolution structural model is used to help determine the mode-of-actionof UCB0599. Here, it is shown that the ligand changes the ensemble of membrane-bound structures, which helps to explain the success this compound, currently being tested in Parkinson's disease patients in a phase 2 trial, has had in animal models of PD.

Circumvention of Topoisomerase IIα Intron 19 Intronic Polyadenylation in Acquired Etoposide-Resistant Human Leukemia K562 Cells.

DNA topoisomerase IIα (TOP2α; 170 kDa, TOP2α/170) is an essential enzyme for proper chromosome dysjunction by producing transient DNA double-stranded breaks and is an important target for DNA damage-stabilizing anticancer agents, such as etoposide. Therapeutic effects of TOP2α poisons can be limited due to acquired drug resistance. We previously demonstrated decreased TOP2α/170 levels in an etoposide-resistant human leukemia K562 subline, designated K/VP.5, accompanied by increased expression of a C-terminal truncated TOP2α isoform (90 kDa; TOP2α/90), which heterodimerized with TOP2α/170 and was a determinant of resistance by exhibiting dominant-negative effects against etoposide activity. Based on 3'-rapid amplification of cDNA ends, we confirmed TOP2α/90 as the translation product of a TOP2α mRNA in which a cryptic polyadenylation site (PAS) harbored in intron 19 (I19) was used. In this report, we investigated whether the resultant intronic polyadenylation (IPA) would be attenuated by blocking or mutating the I19 PAS, thereby circumventing acquired drug resistance. An antisense morpholino oligonucleotide was used to hybridize/block the PAS in TOP2α pre-mRNA in K/VP.5 cells, resulting in decreased TOP2α/90 mRNA/protein levels in K/VP.5 cells and partially circumventing drug resistance. Subsequently, CRISPR/CRISPR-associated protein 9 with homology-directed repair was used to mutate the cryptic I19 PAS (AATAAA→ACCCAA) to prevent IPA. Gene-edited clones exhibited increased TOP2α/170 and decreased TOP2α/90 mRNA/protein and demonstrated restored sensitivity to etoposide and other TOP2α-targeted drugs. Together, results indicated that blocking/mutating a cryptic I19 PAS in K/VP.5 cells reduced IPA and restored sensitivity to TOP2α-targeting drugs. SIGNIFICANCE STATEMENT: The results presented in this study indicate that CRISPR/CRISPR-associated protein 9 gene editing of a cryptic polyadenylation site (PAS) within I19 of the TOP2α gene results in the reversal of acquired resistance to etoposide and other TOP2-targeted drugs. An antisense morpholino oligonucleotide targeting the PAS also partially circumvented resistance.

Use of CRISPR/Cas9 with Homology-Directed Repair to Gene-Edit Topoisomerase IIß in Human Leukemia K562 Cells: Generation of a Resistance Phenotype.

DNA topoisomerase IIß (TOP2ß/180; 180 kDa) is a nuclear enzyme that regulates DNA topology by generation of short-lived DNA double-strand breaks, primarily during transcription. TOP2ß/180 can be a target for DNA damage-stabilizing anticancer drugs, whose efficacy is often limited by chemoresistance. Our laboratory previously demonstrated reduced levels of TOP2ß/180 (and the paralog TOP2α/170) in an acquired etoposide-resistant human leukemia (K562) clonal cell line, K/VP.5, in part due to overexpression of microRNA-9-3p/5p impacting post-transcriptional events. To evaluate the effect on drug sensitivity upon reduction/elimination of TOP2ß/180, a premature stop codon was generated at the TOP2ß/180 gene exon 19/intron 19 boundary (AGAA//GTAA→ATAG//GTAA) in parental K562 cells (which contain four TOP2ß/180 alleles) by CRISPR/Cas9 editing with homology-directed repair to disrupt production of full-length TOP2ß/180. Gene-edited clones were identified and verified by quantitative polymerase chain reaction and Sanger sequencing, respectively. Characterization of TOP2ß/180 gene-edited clones, with one or all four TOP2ß/180 alleles mutated, revealed partial or complete loss of TOP2ß mRNA/protein, respectively. The loss of TOP2ß/180 protein correlated with decreased (2-{4-[(7-chloro-2-quinoxalinyl)oxy]phenoxy}propionic acid)-induced DNA damage and partial resistance in growth inhibition assays. Partial resistance to mitoxantrone was also noted in the gene-edited clone with all four TOP2ß/180 alleles modified. No cross-resistance to etoposide or mAMSA was noted in the gene-edited clones. Results demonstrated the role of TOP2ß/180 in drug sensitivity/resistance in K562 cells and revealed differential paralog activity of TOP2-targeted agents. SIGNIFICANCE STATEMENT: Data indicated that CRISPR/Cas9 editing of the exon 19/intron 19 boundary in the TOP2ß/180 gene to introduce a premature stop codon resulted in partial to complete disruption of TOP2ß/180 expression in human leukemia (K562) cells depending on the number of edited alleles. Edited clones were partially resistant to mitoxantrone and XK469, while lacking resistance to etoposide and mAMSA. Results demonstrated the import of TOP2ß/180 in drug sensitivity/resistance in K562 cells and revealed differential paralog activity of TOP2-targeted agents.

Effects of hsa-miR-9-3p and hsa-miR-9-5p on Topoisomerase IIß Expression in Human Leukemia K562 Cells with Acquired Resistance to Etoposide.

DNA topoisomerase IIα (TOP2α/170; 170 kDa) and topoisomerase IIß (TOP2ß/180; 180 kDa) are targets for a number of anticancer drugs, whose clinical efficacy is attenuated by chemoresistance. Our laboratory selected for an etoposide-resistant K562 clonal subline designated K/VP.5. These cells exhibited decreased TOP2α/170 and TOP2ß/180 expression. We previously demonstrated that a microRNA-9 (miR-9)-mediated posttranscriptional mechanism plays a role in drug resistance via reduced TOP2α/170 protein in K/VP.5 cells. Here, it is hypothesized that a similar miR-9 mechanism is responsible for decreased TOP2ß/180 levels in K/VP.5 cells. Both miR-9-3p and miR-9-5p are overexpressed in K/VP.5 compared with K562 cells, demonstrated by microRNA (miRNA) sequencing and quantitative polymerase chain reaction. The 3'-untranslated region (3'-UTR) of TOP2ß/180 contains miRNA recognition elements (MRE) for both miRNAs. Cotransfection of K562 cells with a luciferase reporter plasmid harboring TOP2ß/180 3'-UTR plus miR-9-3p or miR-9-5p mimics resulted in statistically significant decreased luciferase expression. miR-9-3p and miR-9-5p MRE mutations prevented this decrease, validating direct interaction between these miRNAs and TOP2ß/180 mRNA. Transfection of K562 cells with miR-9-3p/5p mimics led to decreased TOP2ß protein levels without a change in TOP2ß/180 mRNA and resulted in reduced TOP2ß-specific XK469-induced DNA damage. Conversely, K/VP.5 cells transfected with miR-9-3p/5p inhibitors led to increased TOP2ß/180 protein without a change in TOP2ß/180 mRNA and resulted in enhancement of XK469-induced DNA damage. Taken together, these results strongly suggest that TOP2ß/180 mRNA is translationally repressed by miR-9-3p/5p, that these miRNAs play a role in acquired resistance to etoposide, and that they are potential targets for circumvention of resistance to TOP2-targeted agents. SIGNIFICANCE STATEMENT: Results presented here indicate that miR-9-3p and miR-9-5p play a role in acquired resistance to etoposide via decreased DNA topoisomerase IIß 180 kDa protein levels. These findings contribute further information about and potential strategies for circumvention of drug resistance by modulation of microRNA levels. In addition, miR-9-3p and miR-9-5p overexpression in cancer chemoresistance may lead to future validation as biomarkers of responsiveness to DNA topoisomerase II-targeted therapy.

A cross-linguistic approach to treating speech sound disorders in bilingual children.

The present study explores generalisation of production skills across languages when treating speech sound disorders in bilingual children. Early work suggests that treating shared sounds across languages may facilitate cross-linguistic generalisation. Thus, selecting shared sounds across languages as targets may have clinical advantages. In this study, we asked if cross-linguistic generalisation can be facilitated for targets using shared sounds in bilingual children with phonological delays from Spanish (L1) into English (L2) when treating only the L1. Two Spanish-English bilingual children between the ages of 5;0-5;3 with speech sound disorders participated in an intervention with shared sounds as targets. Each child received two sessions per week of therapy that included both linguistically-based and motor-based approaches. Accuracy of targets was assessed within and across languages using a single-subject case design. Results show increased accuracy of targets and generalisation of sounds across languages when treatment was administered only in the L1. Specific growth varied per target and child. The implications affect how we select treatment targets in bilingual children. Future studies should explore additional ways to select targets to increase generalisation of skills and replicate with additional participants.

Trends in breast and prostate cancer screening and diagnostic procedures during the COVID-19 pandemic in central Massachusetts.

PURPOSE: We calculated rates of breast and prostate cancer screening and diagnostic procedures performed during the COVID-19 pandemic through December 2021 compared to the same months in 2019 in a large healthcare provider group in central Massachusetts. METHODS: We included active patients of the provider group between January 2019 and December 2021 aged 30-85 years. Monthly rates of screening mammography and digital breast tomosynthesis, breast MRI, total prostate specific antigen (PSA), and breast or prostate biopsy per 1,000 people were compared by year overall, by age, and race/ethnicity. Completed procedures were identified by relevant codes in electronic health record data. RESULTS: Rates of screening mammography, tomosynthesis, and PSA testing reached the lowest levels in April-May 2020. Breast cancer screening rates decreased 43% in March and 99% in April and May 2020, compared to 2019. Breast cancer screening rates increased gradually beginning in June 2020 through 2021, although more slowly in Black and Hispanic women and in women aged 75-85. PSA testing rates decreased 34% in March, 78% in April, and 53% in May 2020, but rebounded to pre-pandemic levels by June 2020; trends were similar across groups defined by age and race/ethnicity. CONCLUSION: The observed decline in two common screening procedures during the COVID-19 pandemic reflects the impact of the pandemic on cancer early detection and signals potential downstream effects on the prognosis of delayed cancer diagnoses. The slower rate of return for breast cancer screening procedures in certain subgroups should be investigated to ensure all women return for routine screenings.

Imaging Assessment of Pancreatic Cancer Resectability After Neoadjuvant Therapy: AJR Expert Panel Narrative Review.

Despite important innovations in the treatment of pancreatic ductal adenocarcinoma (PDAC), PDAC remains a disease with poor prognosis and high mortality. A key area for potential improvement in the management of PDAC, aside from earlier detection in patients with treatable disease, is the improved ability of imaging techniques to differentiate treatment response after neoadjuvant therapy (NAT) from worsening disease. It is well established that current imaging techniques cannot reliably make this distinction. This narrative review provides an update on the imaging assessment of pancreatic cancer resectability after NAT. Current definitions of borderline resectable PDAC, as well as implications for determining likely patient benefit from NAT, are described. Challenges associated with PDAC pathologic evaluation and surgical decision making that are of relevance to radiologists are discussed. Also explored are the specific limitations of imaging in differentiating the response after NAT from stable or worsening disease, including issues relating to protocol optimization, tumor size assessment, vascular assessment, and liver metastasis detection. The roles of MRI as well as PET and/or hybrid imaging are considered. Finally, a short PDAC reporting template is provided for use after NAT. The highlighted methods seek to improve radiologists' assessment of PDAC treatment response after NAT.

CRISPR/Cas9 Genome Editing of the Human Topoisomerase IIα Intron 19 5' Splice Site Circumvents Etoposide Resistance in Human Leukemia K562 Cells.

An essential function of DNA topoisomerase IIα (TOP2α; 170 kDa, TOP2α/170) is to resolve DNA topologic entanglements during chromosome disjunction by introducing transient DNA double-stranded breaks. TOP2α/170 is an important target for DNA damage-stabilizing anticancer drugs, whose clinical efficacy is compromised by drug resistance often associated with decreased TOP2α/170 expression. We recently demonstrated that an etoposide-resistant K562 clonal subline, K/VP.5, with reduced levels of TOP2α/170, expresses high levels of a novel C-terminal truncated TOP2α isoform (90 kDa, TOP2α/90). TOP2α/90, the translation product of a TOP2α mRNA that retains a processed intron 19 (I19), heterodimerizes with TOP2α/170 and is a resistance determinant through a dominant-negative effect on drug activity. We hypothesized that genome editing to enhance I19 removal would provide a tractable strategy to circumvent acquired TOP2α-mediated drug resistance. To enhance I19 removal in K/VP.5 cells, CRISPR/Cas9 was used to make changes (GAG//GTAA AC →GAG//GTAA GT ) in the TOP2α gene's suboptimal exon 19/intron 19 5' splice site (E19/I19 5' SS). Gene-edited clones were identified by quantitative polymerase chain reaction and verified by sequencing. Characterization of a clone with all TOP2α alleles edited revealed improved I19 removal, decreased TOP2α/90 mRNA/protein, and increased TOP2α/170 mRNA/protein. Sensitivity to etoposide-induced DNA damage (γH2AX, Comet assays) and growth inhibition was restored to levels comparable to those in parental K562 cells. Together, the results indicate that our gene-editing strategy for optimizing the TOP2α E19/I19 5' SS in K/VP.5 cells circumvents resistance to etoposide and other TOP2α-targeted drugs. SIGNIFICANCE STATEMENT: Results presented here indicate that CRISPR/Cas9 gene editing of a suboptimal exon 19/intron 19 5' splice site in the DNA topoisomerase IIα (TOP2α) gene results in circumvention of acquired drug resistance to etoposide and other TOP2α-targeted drugs in a clonal K562 cell line by enhancing removal of intron 19 and thereby decreasing formation of a truncated TOP2α 90 kDa isoform and increasing expression of full-length TOP2α 170 kDa in these resistant cells. Results demonstrate the importance of RNA processing in acquired drug resistance to TOP2α-targeted drugs.

Improved transient silencing of gene expression in the mosquito female Aedes aegypti.

Gene silencing using RNA interference (RNAi) has become a widely used genetic technique to study gene function in many organisms. In insects, this technique is often applied through the delivery of dsRNA. In the adult female Aedes aegypti, a main vector of human-infecting arboviruses, efficiency of gene silencing following dsRNA injection varies greatly according to targeted genes. Difficult knockdowns using dsRNA can thus hamper gene function analysis. Here, by analysing silencing of three different genes in female Ae. aegypti (p400, ago2 and E75), we show that gene silencing can indeed be dsRNA sequence dependent but different efficiencies do not correlate with dsRNA length. Our findings suggest that silencing is likely also gene dependent, probably due to gene-specific tissue expression and/or feedback mechanisms. We demonstrate that use of high doses of dsRNA can improve knockdown efficiency, and injection of a transfection reagent along with dsRNA reduces the variability in efficiency between replicates. Finally, we show that gene silencing cannot be achieved using siRNA injection in Ae. aegypti adult females. Overall, this work should help future gene function analyses using RNAi in adult females Ae. aegypti, leading toward a better understanding of physiological and infectious processes.

GGCX mutations in a patient with overlapping pseudoxanthoma elasticum/cutis laxa-like phenotype.

Pseudoxanthoma elasticum (PXE) is a multisystem disorder characterized by ectopic mineralization of connective tissues with primary manifestations in the skin, eyes and the cardiovascular system. The classic forms of PXE are caused by mutations in the ABCC6 gene encoding the ABCC6 protein, expressed primarily in the liver. Cutis laxa (CL) manifests with loose and sagging skin with loss of recoil. In 2009 we investigated a 19-year-old patient with overlapping cutaneous features of PXE and CL, together with alpha thalassaemia. Genetic analysis failed to identify pathogenic mutations in ABCC6. More recently we developed a gene-targeted panel of next-generation sequencing technology. This panel has 29 genes, 22 of which, including ABCC6 and GGCX, are associated with ectopic mineralization phenotypes. Mutation analysis in the patient identified two heterozygous GGCX mutations: c.200_201delTT in exon 2 and c.763G>A, p.V255M in exon 7. The GGCX gene encodes a γ-glutamyl carboxylase necessary for activation of blood coagulation factors in the liver. The p.V255M mutation was previously reported to result in reduced γ-glutamyl carboxylase activity in vitro, while the c.200_201delTT mutation is novel. Previous studies reported that mutations in GGCX cause overlapping PXE/CL skin phenotypes in association with or without multiple vitamin K-dependent coagulation factor deficiency. Our patient had loose redundant skin, moderate-to-severe angioid streaks and characteristic calcification of elastic structures in the mid dermis, consistent with PXE/CL overlap, but no coagulation abnormalities. Our studies expand the GGCX mutation landscape in patients with PXE-like phenotypes.

Effect of Decreasing the ACR TI-RADS Point Assignment for Punctate Echogenic Foci When They Occur in Mixed Solid and Cystic Thyroid Nodules.

OBJECTIVE. Using the American College of Radiology (ACR) Thyroid Imaging Reporting and Data System (TI-RADS), this study sought to determine whether decreasing the point assignment for punctate echogenic foci in mixed solid and cystic nodules would reduce the number of benign mixed solid and cystic nodules that were biopsied and would not substantially increase the number of missed mixed carcinomas MATERIALS AND METHODS. A multiinstitutional database of 3422 pathologically proven thyroid nodules was evaluated to identify all mixed solid and cystic nodules with punctate echogenic foci. We determined the numbers of mixed benign and malignant nodules that would receive ACR TI-RADS recommendations of fine-needle aspiration, follow-up, and no further evaluation if the points assigned to punctate echogenic foci were changed from 3 points to 1 or 2 points. RESULTS. A total of 287 mixed nodules were adequately characterized for evaluation. When the number of points assigned to punctate echogenic foci was changed from 3 points to 1 point, the point categories changed for 198 mixed nodules. Seven carcinomas would not undergo biopsy, but six of those seven would receive follow-up, and 44 benign nodules would not undergo biopsy. When 2 points were assigned to punctate echogenic foci, the point categories changed for 66 mixed nodules. Three carcinomas would not undergo biopsy, but all three of these would receive follow-up, and eight benign nodules would not undergo biopsy. CONCLUSION. Consideration should be given to decreasing the number of points assigned to punctate echogenic foci in mixed solid and cystic thyroid nodules, given the substantial decrease in the number of benign nodules requiring biopsy and the recommendation of follow-up for any carcinoma 1 cm or larger that did not undergo biopsy.

Inclusion of Thyroid Nodule Location in American College of Radiology TI-RADS Scoring: Impact on System Performance.

Emerging data suggest that the location of thyroid nodules influences malignancy risk. The purpose of this study was to explore the impact of including location in American College of Radiology Thyroid Imaging Reporting and Data System (ACR TI-RADS) scoring. Four of five revised scoring algorithms that added 1 or 2 points to higher-risk locations were associated with lowered accuracy due to lower specificity. However, an algorithm that added 1 point to isthmic nodules did not differ significantly from ACR TI-RADS in accuracy; one additional isthmic cancer was diagnosed for each 10.3 additional benign nodules recommended for biopsy.

Analysis of Malignant Thyroid Nodules That Do Not Meet ACR TI-RADS Criteria for Fine-Needle Aspiration.

OBJECTIVE. Compared with other guidelines, the American College of Radiology (ACR) Thyroid Imaging Reporting and Data System (TI-RADS) has decreased the number of nodules for which fine-needle aspiration is recommended. The purpose of this study was to evaluate the characteristics of malignant nodules that would not be biopsied when the ACR TI-RADS recommendations are followed. MATERIALS AND METHODS. We retrospectively reviewed a total of 3422 thyroid nodules for which a definitive cytologic diagnosis, a definitive histologic diagnosis, or both diagnoses as well as diagnostic ultrasound (US) examinations were available. All nodules were categorized using the ACR TI-RADS, and they were divided into three groups according to the recommendation received: fine-needle aspiration (group 1), follow-up US examination (group 2), or no further evaluation (group 3). RESULTS. Of the 3422 nodules, 352 were malignant. Of these, 240 nodules were assigned to group 1, whereas 72 were assigned to group 2 and 40 were included in group 3. Sixteen of the 40 malignant nodules in group 3 were 1 cm or larger, and, on the basis of analysis of the sonographic features described in the ACR TI-RADS, these nodules were classified as having one of five ACR TI-RADS risk levels (TR1-TR5), with one nodule classified as a TR1 nodule, eight as TR2 nodules, and seven as TR3 nodules. If the current recommendation of no follow-up for TR2 nodules was changed to follow-up for nodules 2.5 cm or larger, seven additional malignant nodules and 316 additional benign nodules would receive a recommendation for follow-up. If the current size threshold (1.5 cm) used to recommend US follow-up for TR3 nodules was decreased to 1.0 cm, seven additional malignant nodules and 118 additional benign nodules would receive a recommendation for follow-up. CONCLUSION. With use of the ACR TI-RADS, most malignant nodules that would not be biopsied would undergo US follow-up, would be smaller than 1 cm, or would both undergo US follow-up and be smaller than 1 cm. Adjusting size thresholds to decrease the number of missed malignant nodules that are 1 cm or larger would result in a substantial increase in the number of benign nodules undergoing follow-up.

Radiation safety education and diagnostic imaging in pediatric patients with surgically treated hydrocephalus: the patient and family perspective.

PURPOSE: Surgically treated hydrocephalus patients are frequently imaged with head computed tomography (CT), and risk/benefit communication with families is inconsistent and unknown. We aimed to educate patients and caregivers about radiation safety in CT and explore their communication preferences. METHODS: We conducted a pediatric CT radiation safety and diagnostic imaging educational workshop for patients and caregivers at a national conference on hydrocephalus to characterize current practice and desired communication about CT imaging. Our workshop consisted of an interactive educational intervention with pre-/post-session surveys followed by feedback from participants. RESULTS: Our session included 34 participants (100% response rate for surveys) with 28 being parents of individuals with hydrocephalus. A total of 76% (n = 26) participants showed an increase in knowledge after the session (p < 0.01). All participants (N = 34) uniformly desired risk/benefit discussions before CT scans. However, 71% stated that they were not informed of risks/benefits of CT scans by a medical professional. Following the session, the number of participants indicating that informed consent should be obtained before CT scans increased from 30 to 33. Respondents also revealed that 14% of children and young adults had received > 100 CT scans for shunt evaluation with the median being 25 scans (IQR 20). CONCLUSIONS: Caregivers desire and deserve to be empowered through education and social support, and continuously engaged through sharing decisions and co-designing care plans. The neurosurgical community is in an ideal position to collaborate with radiologists, primary care providers, and parents in the development and testing of credible, high-quality online and social media resources.

hsa-miR-9-3p and hsa-miR-9-5p as Post-Transcriptional Modulators of DNA Topoisomerase IIα in Human Leukemia K562 Cells with Acquired Resistance to Etoposide.

DNA topoisomerase IIα protein (TOP2α) 170 kDa (TOP2α/170) is an important target for anticancer agents whose efficacy is often attenuated by chemoresistance. Our laboratory has characterized acquired resistance to etoposide in human leukemia K562 cells. The clonal resistant subline K/VP.5 contains reduced TOP2α/170 mRNA and protein levels compared with parental K562 cells. The aim of this study was to determine whether microRNA (miRNA)-mediated mechanisms play a role in drug resistance via decreased expression of TOP2α/170. miRNA-sequencing revealed that human miR-9-3p and miR-9-5p were among the top six of those overexpressed in K/VP.5 compared with K562 cells; validation by quantitative polymerase chain reaction demonstrated overexpression of both miRNAs. miRNA recognition elements (MREs) for both miRNAs are present in the 3'-untranslated region (UTR) of TOP2α/170. Transfecting K562 cells with a reporter plasmid harboring the TOP2α/170 3'-UTR together with either miR-9-3p or miR-9-5p mimics resulted in a statistically significant decrease in luciferase expression. Mutating the miR-9-3p or miR-9-5p MREs prevented this decrease, demonstrating direct interaction between these miRNAs and TOP2α/170 mRNA. Transfection of K562 cells with miR-9-3p or miR-9-5p mimics led to decreased TOP2α/170 protein levels without a change in TOP2α/170 mRNA and resulted in attenuated etoposide-induced DNA damage (gain-of-miRNA-inhibitory function). Conversely, transfection of miR-9-3p or miR-9-5p inhibitors in K/VP.5 cells (overexpressed miR-9 and low TOP2α/170) led to increased TOP2α/170 protein expression without a change in TOP2α/170 mRNA levels and resulted in enhancement of etoposide-induced DNA damage (loss-of-miRNA-inhibitory function). Taken together, these results strongly suggest that these miRNAs play a role in and are potential targets for circumvention of acquired resistance to etoposide. SIGNIFICANCE STATEMENT: Results presented here indicate that miR-9-3p and miR-9-5p decrease DNA topoisomerase IIα protein 170 kDa expression levels in acquired resistance to etoposide. These findings contribute new information about and potential strategies for circumvention of drug resistance by modulation of microRNA levels. Furthermore, increased expression of miR-9-3p and miR-9-5p in chemoresistant cancer cells may support their validation as biomarkers of responsiveness to DNA topoisomerase II-targeted therapy.

Imaging angiogenesis in patients with head and neck squamous cell carcinomas by [68Ga]Ga-DOTA-E-[c(RGDfK)]2 PET/CT.

PURPOSE: Angiogenesis plays an important role in the growth and metastatic spread of solid tumours and is characterised by the expression of integrins on the cell surface of endothelial cells. Radiolabelled RGD peptides specifically target angiogenesis-related αvß3 integrins, expressed on the activated endothelial cells of sprouting blood vessels. Here, we validated the feasibility of 68Ga[Ga]-DOTA-E-[c(RGDfK)]2 (68Ga-RGD) PET/CT to visualise angiogenesis in patients with oral squamous cell carcinoma (OSCC). METHODS: Ten patients with OSCC and scheduled for surgical resection including elective neck dissection received an intravenously administration of 68Ga-RGD (42 ± 8 µg; 214 ± 9 MBq). All patients subsequently underwent dynamic (n = 5) or static PET/CT imaging (n = 5) for 60 min or for 4 min/bed position at 30, 60 and 90 min after injection, respectively. Quantitative tracer uptake in tumour lesions was expressed as standardised uptake values (SUV). Additionally, tumour tissue was immunohistochemically stained for αvß3 integrin to assess the expression pattern. RESULTS: 68Ga-RGD tumour accumulation was observed in all patients. At 60 min post injection, tumour SUVmax ranged between 4.0 and 12.7. Tracer accumulation in tumour tissue plateaued at 10 min after injection. Uptake in background tissue did not change over time, resulting in tumour-to-muscle tissue of 6.4 ± 0.7 at 60 min post injection. CONCLUSIONS: 68Ga-RGD PET/CT of αvß3 integrin expression in OSCC patients is feasible with adequate tumour-to-background ratios. It will provide more insight in angiogenesis as a hallmark of the head and neck squamous cell carcinomas' tumour microenvironment. TRIAL REGISTRATION: https://eudract.ema.europa.eu no. 2015-000917-31.

Quantitative Framework for Risk Stratification of Thyroid Nodules With Ultrasound: A Step Toward Automated Triage of Thyroid Cancer.

OBJECTIVE. The purpose of this study was to explore whether a quantitative framework can be used to sonographically differentiate benign and malignant thyroid nodules at a level comparable to that of experts. MATERIALS AND METHODS. A dataset of ultrasound images of 92 biopsy-confirmed nodules was collected retrospectively. The nodules were delineated and annotated by two expert radiologists using the standardized Thyroid Imaging Reporting and Data System lexicon of the American College of Radiology. In the framework studied, quantitative features of echogenicity, texture, edge sharpness, and margin curvature properties of thyroid nodules were analyzed in a regularized logistic regression model to predict malignancy of a nodule. The framework was validated by leave-one-out cross-validation technique, and ROC AUC, sensitivity, and specificity were used to compare with those obtained with six expert annotation-based classifiers. RESULTS. The AUC of the proposed method was 0.828 (95% CI, 0.715-0.942), which was greater than or comparable to that of the expert classifiers, for which the AUC values ranged from 0.299 to 0.829 (p = 0.99). Use of the proposed framework could have avoided biopsy of 20 of 46 benign nodules in a curative strategy (at sensitivity of 1, statistically significantly higher than three expert classifiers) or helped identify 10 of 46 malignancies in a conservative strategy (at specificity of 1, statistically significantly higher than five expert classifiers). CONCLUSION. When the proposed quantitative framework was used, thyroid nodule malignancy was predicted at the level of expert classifiers. Such a framework may ultimately prove useful as the basis for a fully automated system of thyroid nodule triage.

Crosslinked network microstructure of carbon nanomaterials promotes flaw-tolerant mechanical response.

Carbon nanomaterials, such as carbon nanotubes (CNTs) and carbon nanofibers (CNFs), are chemically inert in their highly graphitic forms. Various post processing methods can activate their surfaces to enhance their interactions with a host matrix in a nanocomposite. Chemical surface functionalization is used often. This method however can lead to major strength loss in nanomaterials stemming from induced surface defects (changing sp2 bonds to sp3 bonds). In this manuscript, we have experimentally studied the mechanical properties of the individual, pyrolysis-fabricated CNFs. These CNFs have a highly crosslinked 3D network of C-C bonds. The strength of CNFs has been studied as a function of O/C ratio. The loss in strength due to functionalization has been compared to that of other carbon nanomaterials with layered strcutures (CNT and graphene). Comparisons were also made with carbon microfibers. Fracture strength estimations of the critical flaw size in CNFs, CNTs and graphene were also made. The results revealed that despite having high surface area, carbon nanomaterials with crosslinked microstructure are resilient to flaws as big (deep) as 10-30 nm, while nanomaterials with layered structure (such as CNTs) experience a dramatic loss in strength with much lower flaw sizes. Hence, it seems that graphitic nanomaterials such as graphene and CNT have high strenght that, although higher than CNFs, comes at a cost to flaw tolerance and robustness. Since failure is often progressive, this work demonstrates a benefit that crosslinked nanomaterials have over highly graphitic ones, such as CNTs, in load bearing applications.