How paediatric HIV services weathered the COVID-19 storm in Tshwane District, South Africa.

Background: The coronavirus disease 2019 (COVID-19) pandemic disrupted paediatric HIV services across South Africa. Shortly before COVID-19, updated national HIV guidelines were released. Objectives: This study describes COVID-19's impact on paediatric HIV services in Tshwane District, South Africa. Method: A retrospective review of National Institute for Communicable Diseases and District Health Information System data for Tshwane District from April 2019 to March 2022. Data included: Early Infant Diagnosis (EID), HIV viral load (VL) and CD4 monitoring and HIV management among children (< 15 years) living with HIV (CLHIV). Pre-pandemic (2019/2020) and pandemic periods (2020/2021, 2021/2022) were compared. Results: Year-on-year, HIV testing improved at 10 weeks, 6 months, and 18 months, whereas birth testing decreased. HIV EID case rates were 485 (2019/2020), 410 (2020/2021) and 454 (2021/2022). HIV EID test positivity was 0.77% - 1.2%. Antiretroviral treatment initiation declined from 2019/2020 to 2020/2021, but improved in 2021/2022.Initial HIV VL and CD4 testing declined, with HIV VL testing increasing in 2021/2022, and CD4 testing further declining. HIV VL suppression rate among CLHIV ranged from 69% to 73%. Conclusion: Initially, COVID-19 resulted in reduced paediatric HIV services as children disengaged from care. Indicators eventually recovered to proximate pre-pandemic levels; however, compensatory increases did not occur. Thus, some children may not have returned to care.

Skeletal fluorosis secondary to methoxyflurane use for chronic pain.

Skeletal fluorosis is rare and occurs secondary to chronic high amounts of fluoride consumption, manifesting as diffuse osteosclerosis, skeletal pain, connective tissue calcification, and increased fracture risk. Methoxyflurane is a volatile, fluorinated hydrocarbon-inhaled analgesic, and the maximum recommended dose is 15 mL (99.9 % w/w) per wk. A rodent study found increased skeletal fluoride after methoxyflurane exposure. However, skeletal fluorosis secondary to methoxyflurane use in humans has rarely been reported. We present the case of a 47-yr-old female with diffuse osteosclerosis secondary to fluorosis from methoxyflurane use for chronic pain, presenting with 3 yr of generalized bony pain and multiple fragility fractures. Lumbar spine BMD was elevated. CT and radiographs demonstrated new-onset marked diffuse osteosclerosis, with calcification of interosseous membranes and ligaments, and a bone scan demonstrated a grossly increased uptake throughout the skeleton. Biochemistry revealed an elevated alkaline phosphatase and bone turnover markers, mild secondary hyperparathyroidism with vitamin D deficiency, and mild renal impairment. Zoledronic acid, prescribed for presumed Paget's disease, severely exacerbated bony pain. Urinary fluoride was elevated (7.3 mg/L; reference range < 3.0 mg/L) and the patient revealed using methoxyflurane 9 mL per wk for 8 yr for chronic pain. A decalcified bone biopsy revealed haphazardly arranged cement lines and osteocytes lacunae and canaliculi, which was consistent with an osteosclerotic process. Focal subtle basophilic stippling around osteocyte lacunae was suggestive of fluorosis. Although fluorosis is not a histological diagnosis, the presence of compatible histology features was supportive of the diagnosis in this case with clinical-radiological-pathological correlation. Skeletal fluorosis should be considered as a cause of acquired diffuse osteosclerosis. Methoxyflurane should not be recommended for chronic pain. The risk of repeated low-dose exposure to fluoride from methoxyflurane use as analgesia may be greater than expected, and the maximum recommended dose for methoxyflurane may require re-evaluation to minimize skeletal complications. Abbreviated abstract: Skeletal fluorosis is rare and occurs secondary to chronic high amounts of fluoride consumption, manifesting as diffuse osteosclerosis, skeletal pain, connective tissue calcification, and increased fracture risk. We present the case of a 47-yr-old female with skeletal fluorosis secondary to long-term methoxyflurane for chronic pain. The risk of repeated low-dose exposure to fluoride from methoxyflurane use for analgesia may be greater than expected, and the maximum recommended dose for methoxyflurane may require re-evaluation to minimize skeletal complications.

Retention in care of infants diagnosed with HIV at birth: Beyond the diagnostic strategy.

Background: Birth HIV point-of-care (POC) tests curtail analytical testing issues and expedite diagnosis, potentially allowing for earlier mother-infant pair engagement and improved outcomes. Many children are lost post antiretroviral therapy (ART) initiation within the first 6 months of follow-up. Objectives: We compared 6-month retention in care, HIV viral load (VL) suppression and mortality among infants diagnosed with HIV at birth, using laboratory-based versus POC HIV PCR testing. Method: From 2018 to 2019, infants exposed to HIV underwent birth HIV PCR POC testing at Kalafong Provincial Tertiary Hospital in Tshwane District. Their outcomes were compared to a historical control born between 2014 and 2016, who exclusively underwent laboratory-based HIV PCR testing. Both groups received comparable HIV care following national guidelines. Results: Fifty-seven infants were studied (POC: 27; Control: 30). The POC turnaround time was significantly shorter (POC: 15.5 h [IQR: 4.3-24.7], Control: 68.3 h [IQR 46.0-93.9]; p = < 0.0001). Both populations had the same elapsed time from HIV diagnosis to ART initiation (median: 13 days, POC: IQR 8-21 days; Control: IQR 9-36 days). Six infants were never initiated (POC: 2 [7%]; Control: 4 [13%]). At 6 months, overall care retention was 72% (41/57), higher among the Control group (Control 23/30, 77%; POC: 18/27, 67%). HIV viral suppression at 6 months was higher among the POC group (POC: 14/18, 78%; Control: 9/19, 47%, p = 0.09). No deaths were reported. Conclusion: Poor care retention at 6 months post ART initiation is concerning. Initial mother-infant visits should be effectively utilised to assess and manage potential risk factors for loss of follow-up. Contribution: This study highlights the ongoing need to find workable solutions to improve retention in care, thereby ensuring the benefits of expedited HIV diagnosis and ART initiation.

POU2F3-expressing intraepithelial small-cell carcinoma with mixed small-cell carcinoma and conventional-type urothelial carcinoma of the urinary bladder.

Based on lineage-specific transcription factors, small-cell neuroendocrine carcinoma (SmCC) of the urinary bladder has recently been subtyped into three molecular subtypes: ASCL1, NEUROD1 and POU2F3. The latter is a master transcriptional regulator of tuft cells (TCs) which are rare solitary cells found in various mucosal epithelia such as the gastrointestinal tract, but which have not been reported in the bladder. The POU2F3 subtype shows low or absent neuroendocrine marker expression. A case of mixed SmCC and conventional-type urothelial carcinoma (CUC) of the urinary bladder with POU2F3-expressing intraepithelial small-cell carcinoma in keeping with a tuft cell phenotype, arising in association with intestinal metaplasia (IM) is described. The presence of POU2F3-expressing cells in normal urothelium, cystitis cystica glandularis and IM of the urinary bladder is demonstrated in separate cases of cystitis cystica glandularis with IM. Also, POU2F3 expression is identified in a subset of bladder SmCC.

Identification of aberrant luminal progenitors and mTORC1 as a potential breast cancer prevention target in BRCA2 mutation carriers.

Inheritance of a BRCA2 pathogenic variant conveys a substantial life-time risk of breast cancer. Identification of the cell(s)-of-origin of BRCA2-mutant breast cancer and targetable perturbations that contribute to transformation remains an unmet need for these individuals who frequently undergo prophylactic mastectomy. Using preneoplastic specimens from age-matched, premenopausal females, here we show broad dysregulation across the luminal compartment in BRCA2mut/+ tissue, including expansion of aberrant ERBB3lo luminal progenitor and mature cells, and the presence of atypical oestrogen receptor (ER)-positive lesions. Transcriptional profiling and functional assays revealed perturbed proteostasis and translation in ERBB3lo progenitors in BRCA2mut/+ breast tissue, independent of ageing. Similar molecular perturbations marked tumours bearing BRCA2-truncating mutations. ERBB3lo progenitors could generate both ER+ and ER- cells, potentially serving as cells-of-origin for ER-positive or triple-negative cancers. Short-term treatment with an mTORC1 inhibitor substantially curtailed tumorigenesis in a preclinical model of BRCA2-deficient breast cancer, thus uncovering a potential prevention strategy for BRCA2 mutation carriers.

Molecular and Pathologic Characterization of YAP1-Expressing Small Cell Lung Cancer Cell Lines Leads to Reclassification as SMARCA4-Deficient Malignancies.

PURPOSE: The classification of small cell lung cancer (SCLC) into distinct molecular subtypes defined by ASCL1, NEUROD1, POU2F3, or YAP1 (SCLC-A, -N, -P, or -Y) expression, paves the way for a personalized treatment approach. However, the existence of a distinct YAP1-expressing SCLC subtype remains controversial. EXPERIMENTAL DESIGN: To better understand YAP1-expressing SCLC, the mutational landscape of human SCLC cell lines was interrogated to identify pathogenic alterations unique to SCLC-Y. Xenograft tumors, generated from cell lines representing the four SCLC molecular subtypes, were evaluated by a panel of pathologists who routinely diagnose thoracic malignancies. Diagnoses were complemented by transcriptomic analysis of primary tumors and human cell line datasets. Protein expression profiles were validated in patient tumor tissue. RESULTS: Unexpectedly, pathogenic mutations in SMARCA4 were identified in six of eight SCLC-Y cell lines and correlated with reduced SMARCA4 mRNA and protein expression. Pathologist evaluations revealed that SMARCA4-deficient SCLC-Y tumors exhibited features consistent with thoracic SMARCA4-deficient undifferentiated tumors (SMARCA4-UT). Similarly, the transcriptional profile SMARCA4-mutant SCLC-Y lines more closely resembled primary SMARCA4-UT, or SMARCA4-deficient non-small cell carcinoma, than SCLC. Furthermore, SMARCA4-UT patient samples were associated with a YAP1 transcriptional signature and exhibited strong YAP1 protein expression. Together, we found little evidence to support a diagnosis of SCLC for any of the YAP1-expressing cell lines originally used to define the SCLC-Y subtype. CONCLUSIONS: SMARCA4-mutant SCLC-Y cell lines exhibit characteristics consistent with SMARCA4-deficient malignancies rather than SCLC. Our findings suggest that, unlike ASCL1, NEUROD1, and POU2F3, YAP1 is not a subtype defining transcription factor in SCLC. See related commentary by Rekhtman, p. 1708.

DNA Mismatch Repair Gene Variant Classification: Evaluating the Utility of Somatic Mutations and Mismatch Repair Deficient Colonic Crypts and Endometrial Glands.

Germline pathogenic variants in the DNA mismatch repair (MMR) genes (Lynch syndrome) predispose to colorectal (CRC) and endometrial (EC) cancer. Lynch syndrome specific tumor features were evaluated for their ability to support the ACMG/InSiGHT framework in classifying variants of uncertain clinical significance (VUS) in the MMR genes. Twenty-eight CRC or EC tumors from 25 VUS carriers (6xMLH1, 9xMSH2, 6xMSH6, 4xPMS2), underwent targeted tumor sequencing for the presence of microsatellite instability/MMR-deficiency (MSI-H/dMMR) status and identification of a somatic MMR mutation (second hit). Immunohistochemical testing for the presence of dMMR crypts/glands in normal tissue was also performed. The ACMG/InSiGHT framework reclassified 7/25 (28%) VUS to likely pathogenic (LP), three (12%) to benign/likely benign, and 15 (60%) VUS remained unchanged. For the seven re-classified LP variants comprising nine tumors, tumor sequencing confirmed MSI-H/dMMR (8/9, 88.9%) and a second hit (7/9, 77.8%). Of these LP reclassified variants where normal tissue was available, the presence of a dMMR crypt/gland was found in 2/4 (50%). Furthermore, a dMMR endometrial gland in a carrier of an MSH2 exon 1-6 duplication provides further support for an upgrade of this VUS to LP. Our study confirmed that identifying these Lynch syndrome features can improve MMR variant classification, enabling optimal clinical care.

A Matched Molecular and Clinical Analysis of the Epithelioid Haemangioendothelioma Cohort in the Stafford Fox Rare Cancer Program and Contextual Literature Review.

BACKGROUND: Epithelioid haemangioendothelioma (EHE) is an ultra-rare malignant vascular tumour with a prevalence of 1 per 1,000,000. It is typically molecularly characterised by a WWTR1::CAMTA1 gene fusion in approximately 90% of cases, or a YAP1::TFE3 gene fusion in approximately 10% of cases. EHE cases are typically refractory to therapies, and no anticancer agents are reimbursed for EHE in Australia. METHODS: We report a cohort of nine EHE cases with comprehensive histologic and molecular profiling from the Walter and Eliza Hall Institute of Medical Research Stafford Fox Rare Cancer Program (WEHI-SFRCP) collated via nation-wide referral to the Australian Rare Cancer (ARC) Portal. The diagnoses of EHE were confirmed by histopathological and immunohistochemical (IHC) examination. Molecular profiling was performed using the TruSight Oncology 500 assay, the TruSight RNA fusion panel, whole genome sequencing (WGS), or whole exome sequencing (WES). RESULTS: Molecular analysis of RNA, DNA or both was possible in seven of nine cases. The WWTR1::CAMTA1 fusion was identified in five cases. The YAP1::TFE3 fusion was identified in one case, demonstrating unique morphology compared to cases with the more common WWTR1::CAMTA1 fusion. All tumours expressed typical endothelial markers CD31, ERG, and CD34 and were negative for pan-cytokeratin. Cases with a WWTR1::CAMTA1 fusion displayed high expression of CAMTA1 and the single case with a YAP1::TFE3 fusion displayed high expression of TFE3. Survival was highly variable and unrelated to molecular profile. CONCLUSIONS: This cohort of EHE cases provides molecular and histopathological characterisation and matching clinical information that emphasises the molecular patterns and variable clinical outcomes and adds to our knowledge of this ultra-rare cancer. Such information from multiple studies will advance our understanding, potentially improving treatment options.

Prevalence of subclinical lung cancer detected at autopsy: a systematic review.

BACKGROUND: Lung cancer screening in high-risk populations with low-dose computed tomography is supported by international associations and recommendations. Overdiagnosis is considered a risk of screening with associated harms. The aim of this paper is to determine the prevalence of subclinical lung cancer diagnosed post-mortem to better understand the reservoir of subclinical lung cancer. METHODS: We searched EMBASE, PubMed, and MEDLINE databases from inception until March 2022 with no language restrictions. We considered all studies with ≥100 autopsies in adults. Two reviewers independently assessed eligibility of studies, extracted data, and assessed risk of bias of included studies. We performed a meta-analysis using a random-effects model for prevalence of subclinical lung cancer diagnosed post-mortem with sensitivity and subgroup analyses. RESULTS: A total of 13 studies with 16 730 autopsies were included. Pooled prevalence was 0.4% (95% CI 0.20 to 0.82%, I2 = 84%, tau2 = 1.19, low certainty evidence,16 730 autopsies). We performed a sensitivity analysis excluding studies which did not specify exclusion of children in their cohort, with a pooled prevalence of subclinical lung cancer of 0.87% (95% CI 0.48 to 1.57%, I2 = 71%, tau2 = 0.38, 6998 autopsies, 8 studies). CONCLUSIONS: This is the first published systematic review to evaluate the prevalence of post-mortem subclinical lung cancer. Compared to autopsy systematic reviews in breast, prostate and thyroid cancers, the pooled prevalence is lower in lung cancer for subclinical cancer. This result should be interpreted with caution due to the included studies risk of bias and heterogeneity, with further high-quality studies required in target screening populations.

Diverse values of nature for sustainability.

Twenty-five years since foundational publications on valuing ecosystem services for human well-being1,2, addressing the global biodiversity crisis3 still implies confronting barriers to incorporating nature's diverse values into decision-making. These barriers include powerful interests supported by current norms and legal rules such as property rights, which determine whose values and which values of nature are acted on. A better understanding of how and why nature is (under)valued is more urgent than ever4. Notwithstanding agreements to incorporate nature's values into actions, including the Kunming-Montreal Global Biodiversity Framework (GBF)5 and the UN Sustainable Development Goals6, predominant environmental and development policies still prioritize a subset of values, particularly those linked to markets, and ignore other ways people relate to and benefit from nature7. Arguably, a 'values crisis' underpins the intertwined crises of biodiversity loss and climate change8, pandemic emergence9 and socio-environmental injustices10. On the basis of more than 50,000 scientific publications, policy documents and Indigenous and local knowledge sources, the Intergovernmental Platform on Biodiversity and Ecosystem Services (IPBES) assessed knowledge on nature's diverse values and valuation methods to gain insights into their role in policymaking and fuller integration into decisions7,11. Applying this evidence, combinations of values-centred approaches are proposed to improve valuation and address barriers to uptake, ultimately leveraging transformative changes towards more just (that is, fair treatment of people and nature, including inter- and intragenerational equity) and sustainable futures.

Bridging Massive Acetabular Defects With the Triflange Cup: 10- to 28-Year Results.

BACKGROUND: Massive acetabular bone loss, encountered complex primary or revision total hip arthroplasty, remains a reconstructive challenge. The custom triflange cup reliably achieves both early fixation and longer-term stability. This study presents the 10-year minimum three-surgeon follow-up of acetabular defects managed with a custom triflange component. METHODS: All patients who underwent a custom triflange acetabular component implantation from January 1992 to December 2009 were identified. Demographics, implant data, outcomes, and reoperations were collected and analyzed. Bone defects in all cases were Paprosky type IIIA, IIIB, or IV. A total of 233 patients (241 hips) underwent implantation of a custom triflange during the study period. There were 81 patients (83 hips) who died prior to minimum follow-up and 84 patients (88 hips) had minimum follow-up of 10 years (mean 15.2; range, 10 to 28), or failure prior to 10 years. RESULTS: Complications requiring additional surgery occurred in 43 hips (49%). There were 10 revisions for failure (11.4%); four were due to recurrent infection, three for aseptic loosening, and one for recurrent infection with all revised to a new triflange. There was one patient who was resected to a Girdlestone for infection and one patient was revised for infection to a bipolar hemiprosthesis due to a healed discontinuity. CONCLUSION: To our knowledge, this study represents the largest cohort and longest follow-up in the current literature and demonstrates excellent survivorship and clinical results at an average of 15 years follow-up. The component was retained in 89% of cases.

Differential expression of genes controlling lymphocyte differentiation and migration in two distinct endotypes of type 1 diabetes.

AIMS: Morphological studies of pancreas samples obtained from young people with recent-onset type 1 diabetes have revealed distinct patterns of immune cell infiltration of the pancreatic islets suggestive of two age-associated type 1 diabetes endotypes that differ by inflammatory responses and rates of disease progression. The objective of this study was to investigate whether these proposed disease endotypes are associated with pathological differences in immune cell activation and cytokine secretion by applying multiplexed gene expression analysis to pancreatic tissue from recent-onset type 1 diabetes cases. METHODS: RNA was extracted from samples of fixed, paraffin-embedded pancreas tissue from type 1 diabetes cases characterised by endotype and from controls without diabetes. Expression levels of 750 genes associated with autoimmune inflammation were determined by hybridisation to a panel of capture and reporter probes and these were counted as a measure of gene expression. Normalised counts were analysed for differences in expression between 29 type 1 diabetes cases and 7 controls without diabetes, and between the two type 1 diabetes endotypes. RESULTS: Ten inflammation-associated genes, including INS, were significantly under-expressed in both endotypes and 48 genes were more highly expressed. A different set of 13 genes associated with the development, activation and migration of lymphocytes was uniquely overexpressed in the pancreas of people developing diabetes at younger age. CONCLUSIONS: The results provide evidence that histologically defined type 1 diabetes endotypes differ in their immunopathology and identify inflammatory pathways specifically involved in disease developing at a young age, essential for a better understanding of disease heterogeneity.

Methods for assessment of the tumour microenvironment and immune interactions in non-small cell lung cancer. A narrative review.

Non-small cell lung cancer (NSCLC) is one of the leading causes of cancer death worldwide. Immunotherapy with immune checkpoint inhibitors (ICI) has significantly improved outcomes in some patients, however 80-85% of patients receiving immunotherapy develop primary resistance, manifesting as a lack of response to therapy. Of those that do have an initial response, disease progression may occur due to acquired resistance. The make-up of the tumour microenvironment (TME) and the interaction between tumour infiltrating immune cells and cancer cells can have a large impact on the response to immunotherapy. Robust assessment of the TME with accurate and reproducible methods is vital to understanding mechanisms of immunotherapy resistance. In this paper we will review the evidence of several methodologies to assess the TME, including multiplex immunohistochemistry, imaging mass cytometry, flow cytometry, mass cytometry and RNA sequencing.

PI3Kα Translocation Mediates Nuclear PtdIns(3,4,5)P3 Effector Signaling in Colorectal Cancer.

The canonical view of PI3Kα signaling describes phosphatidylinositol-3,4,5-trisphosphate (PtdIns(3,4,5)P3) generation and activation of downstream effectors at the plasma membrane or at microtubule-bound endosomes. Here, we show that colorectal cancer (CRC) cell lines exhibit a diverse plasma membrane-nuclear distribution of PI3Kα, controlling corresponding levels of subcellular PtdIns(3,4,5)P3 pools. PI3Kα nuclear translocation was mediated by the importin ß-dependent nuclear import pathway. By PtdIns(3,4,5)P3 affinity capture mass spectrometry done in the presence of SDS on CRC cell lines with PI3Kα nuclear localization, we identified 867 potential nuclear PtdIns(3,4,5)P3 effector proteins. Nuclear PtdIns(3,4,5)P3 interactome proteins were characterized by noncanonical PtdIns(3,4,5)P3-binding domains and showed overrepresentation for nuclear membrane, nucleolus, and nuclear speckles. The nuclear PtdIns(3,4,5)P3 interactome was enriched for proteins related to RNA metabolism, with splicing reporter assays and SC-35 foci staining suggesting a role of epidermal growth factor-stimulated nuclear PI3Kα signaling in modulating pre-mRNA splicing. In patient tumors, nuclear p110α staining was associated with lower T stage and mucinous histology. These results indicate that PI3Kα translocation mediates nuclear PtdIns(3,4,5)P3 effector signaling in human CRC, modulating signaling responses.

Safety and Feasibility of a Novel Externally Cooled Bronchoscopic Radiofrequency Ablation Catheter for Ablation of Peripheral Lung Tumours: A First-In-Human Dose Escalation Study.

BACKGROUND: Radiofrequency ablation (RFA) is an established modality for percutaneous ablation of non-small cell lung cancer (NSCLC) in medically inoperable patients but is underutilized clinically due to side effects. We have developed a novel, completely endobronchial RFA catheter with an externally cooled electrode. OBJECTIVES: The objective of this study was to establish the safety and feasibility of bronchoscopic RFA using a novel, externally cooled catheter for ablation of peripheral NSCLC. METHODS: Patients with stage I biopsy-confirmed NSCLC underwent bronchoscopic RFA of tumour 7 days prior to lobectomy. The RFA catheter was delivered bronchoscopically to peripheral NSCLC lesions, guided by radial endobronchial ultrasound, with positioning confirmed using intra-procedural cone beam CT. Pre-operative CT chest and histologic examination of resected specimens were used to establish distribution/uniformity of ablation and efficacy of tumour ablation. RESULTS: RFA in the first patient was complicated by dispersal of heated saline due to cough, resulting in ICU admission. The patient recovered fully and underwent uncomplicated lobectomy. Subsequently, the protocol was altered to mandate neuromuscular blockade with a pre-determined dose escalation, with algorithm-restricted energy (kJ) and irrigated saline volume (mL) constraints. A further 10 patients consented and seven underwent successful bronchoscopic RFA of peripheral NSCLC. No significant adverse events were noted. Ablation zone included tumour in all cases (proportion of tumour ablated ranged 8-72%), with uniform necrosis of tissue within ablation zones observed at higher energy levels. Ablation zone diameter correlated with RFA energy delivered (R2 = 0.553), with maximum long axis diameter of ablation zone 3.1 cm (22.9 kJ). CONCLUSION: Bronchoscopic RFA using an externally cooled catheter is feasible, appears safe, and achieves uniform ablation within the treatment zone. Uncontrolled escape of heated saline poses a major safety risk but can be prevented procedurally through neuromuscular blockade and by limiting irrigation.

Cell signaling activation and extracellular matrix remodeling underpin glioma tumor microenvironment heterogeneity and organization.

PURPOSE: Tumor cells thrive by adapting to the signals in their microenvironment. To adapt, cancer cells activate signaling and transcriptional programs and migrate to establish micro-niches, in response to signals from neighboring cells and non-cellular stromal factors. Understanding how the tumor microenvironment evolves during disease progression is crucial to deciphering the mechanisms underlying the functional behavior of cancer cells. METHODS: Multiplex immunohistochemistry, spatial analysis and histological dyes were used to identify and measure immune cell infiltration, cell signal activation and extracellular matrix deposition in low-grade, high-grade astrocytoma and glioblastoma. RESULTS: We show that lower grade astrocytoma tissue is largely devoid of infiltrating immune cells and extracellular matrix proteins, while high-grade astrocytoma exhibits abundant immune cell infiltration, activation, and extensive tissue remodeling. Spatial analysis shows that most T-cells are restricted to perivascular regions, but bone marrow-derived macrophages penetrate deep into neoplastic cell-rich regions. The tumor microenvironment is characterized by heterogeneous PI3K, MAPK and CREB signaling, with specific signaling profiles correlating with distinct pathological hallmarks, including angiogenesis, tumor cell density and regions where neoplastic cells border the extracellular matrix. Our results also show that tissue remodeling is important in regulating the architecture of the tumor microenvironment during tumor progression. CONCLUSION: The tumor microenvironment in malignant astrocytoma, exhibits changes in cell composition, cell signaling activation and extracellular matrix deposition during disease development and that targeting the extracellular matrix, as well as cell signaling activation will be critical to designing personalized therapy.

Shariant platform: Enabling evidence sharing across Australian clinical genetic-testing laboratories to support variant interpretation.

Sharing genomic variant interpretations across laboratories promotes consistency in variant assertions. A landscape analysis of Australian clinical genetic-testing laboratories in 2017 identified that, despite the national-accreditation-body recommendations encouraging laboratories to submit genotypic data to clinical databases, fewer than 300 variants had been shared to the ClinVar public database. Consultations with Australian laboratories identified resource constraints limiting routine application of manual processes, consent issues, and differences in interpretation systems as barriers to sharing. This information was used to define key needs and solutions required to enable national sharing of variant interpretations. The Shariant platform, using both the GRCh37 and GRCh38 genome builds, was developed to enable ongoing sharing of variant interpretations and associated evidence between Australian clinical genetic-testing laboratories. Where possible, two-way automated sharing was implemented so that disruption to laboratory workflows would be minimized. Terms of use were developed through consultation and currently restrict access to Australian clinical genetic-testing laboratories. Shariant was designed to store and compare structured evidence, to promote and record resolution of inter-laboratory classification discrepancies, and to streamline the submission of variant assertions to ClinVar. As of December 2021, more than 14,000 largely prospectively curated variant records from 11 participating laboratories have been shared. Discrepant classifications have been identified for 11% (28/260) of variants submitted by more than one laboratory. We have demonstrated that co-design with clinical laboratories is vital to developing and implementing a national variant-interpretation sharing effort. This approach has improved inter-laboratory concordance and enabled opportunities to standardize interpretation practices.

Using weighted expert judgement and nonlinear data analysis to improve Bayesian belief network models for riverine ecosystem services.

Rivers are a key part of the hydrological cycle and a vital conduit of water resources, but are under increasing threat from anthropogenic pressures. Linking pressures with ecosystem services is challenging because the processes interconnecting the physico-chemical, biological and socio-economic elements are usually captured using heterogenous methods. Our objectives were, firstly, to advance an existing proof-of-principle Bayesian belief network (BBN) model for integration of ecosystem services considerations into river management. We causally linked catchment stressors with ecosystem services using weighted evidence from an expert workshop (capturing confidence among expert groups), legislation and published literature. The BBN was calibrated with analyses of national monitoring data (including non-linear relationships and ecologically meaningful breakpoints) and expert judgement. We used a novel expected index of desirability to quantify the model outputs. Secondly, we applied the BBN to three case study catchments in Ireland to demonstrate the implications of changes in stressor levels for ecosystem services in different settings. Four out of the seven significant relationships in data analyses were non-linear, highlighting that non-linearity is common in ecosystems, but rarely considered in environmental modelling. Deficiency of riparian shading was identified as a prevalent and strong influence, which should be addressed to improve a broad range of societal benefits, particularly in the catchments where riparian shading is scarce. Sediment load had a lower influence on river biology in flashy rivers where it has less potential to settle out. Sediment interacted synergistically with organic matter and phosphate where these stressors were active; tackling these stressor pairs simultaneously can yield additional societal benefits compared to the sum of their individual influences, which highlights the value of integrated management. Our BBN model can be parametrised for other Irish catchments whereas elements of our approach, including the expected index of desirability, can be adapted globally.

99m Tc-Sestamibi Thyroid Scintigraphy in Amiodarone-Induced Thyrotoxicosis : Functional Imaging-Histopathologic Correlation.

ABSTRACT: 99m Tc-sestamibi thyroid scintigraphy (STS) can aid in differentiating between types 1 and 2 amiodarone-induced thyrotoxicosis (AIT). We present a consecutive case series of 4 men (aged 56-75 years) in whom both 99m Tc-STS and thyroid histology were consistent with a diagnosis of type 2 AIT, representing the first reported histopathologic correlation for this diagnostic test. Median amiodarone treatment duration was 26 months (range, 10-39 months), and amiodarone was discontinued a median of 3 months preoperatively (range, 2-4 months) in all 4 cases. 99m Tc-STS is a promising functional imaging modality, which has the potential to aid clinicians in the diagnostic workup and treatment of AIT.