Defining T Cell Subsets in Human Tonsils Using ChipCytometry.

ChipCytometry is a multiplex imaging method that can be used to analyze either cell suspensions or tissue sections. Images are acquired by iterative cycles of immunostaining with fluorescently labeled Abs, followed by photobleaching, which allows the accumulation of multiple markers on a single sample. In this study, we explored the feasibility of using ChipCytometry to identify and phenotype cell subsets, including rare cell types, using a combination of tissue sections and single-cell suspensions. Using ChipCytometry of tissue sections, we successfully demonstrated the architecture of human palatine tonsils, including the B and T cell zones, and characterized subcompartments such as the B cell mantle and germinal center zone, as well as intrafollicular PD1-expressing CD4+ T cells. Additionally, we were able to identify the rare tonsillar T cell subsets, mucosal-associated invariant T (MAIT) and γδ-T cells, within tonsil tissue. Using single-cell suspension ChipCytometry, we further dissected human tonsillar T cell subsets via unsupervised clustering analysis as well as supervised traditional manual gating. We were able to show that PD1+CD4+ T cells are comprised of CXCR5+BCL6high follicular Th cells and CXCR5-BCL6mid pre-follicular Th cells. Both supervised and unsupervised analysis approaches identified MAIT cells in single-cell suspensions, confirming a phenotype similar to that of blood-derived MAIT cells. In this study, we demonstrate that ChipCytometry is a viable method for single-cell suspension cytometry and analysis, with the additional benefit of allowing phenotyping in a spatial context using tissue sections.

Stimulatory MAIT cell antigens reach the circulation and are efficiently metabolised and presented by human liver cells.

OBJECTIVE: Mucosal-associated invariant T (MAIT) cells are the most abundant T cells in human liver. They respond to bacterial metabolites presented by major histocompatibility complex-like molecule MR1. MAIT cells exert regulatory and antimicrobial functions and are implicated in liver fibrogenesis. It is not well understood which liver cells function as antigen (Ag)-presenting cells for MAIT cells, and under which conditions stimulatory Ags reach the circulation. DESIGN: We used different types of primary human liver cells in Ag-presentation assays to blood-derived and liver-derived MAIT cells. We assessed MAIT cell stimulatory potential of serum from healthy subjects and patients with portal hypertension undergoing transjugular intrahepatic portosystemic shunt stent, and patients with inflammatory bowel disease (IBD). RESULTS: MAIT cells were dispersed throughout healthy human liver and all tested liver cell types stimulated MAIT cells, hepatocytes being most efficient. MAIT cell activation by liver cells occurred in response to bacterial lysate and pure Ag, and was prevented by non-activating MR1 ligands. Serum derived from peripheral and portal blood, and from patients with IBD stimulated MAIT cells in MR1-dependent manner. CONCLUSION: Our findings reveal previously unrecognised roles of liver cells in Ag metabolism and activation of MAIT cells, repression of which creates an opportunity to design antifibrotic therapies. The presence of MAIT cell stimulatory Ags in serum rationalises the observed activated MAIT cell phenotype in liver. Increased serum levels of gut-derived MAIT cell stimulatory ligands in patients with impaired intestinal barrier function indicate that intrahepatic Ag-presentation may represent an important step in the development of liver disease.

Female breast cancer treatment and survival in South Australia: Results from linked health data.

OBJECTIVE: We investigated treatment and survival by clinical and sociodemographic characteristics for service evaluation using linked data. METHOD: Data on invasive female breast cancers (n = 13,494) from the South Australian Cancer Registry (2000-2014 diagnoses) were linked to hospital inpatient, radiotherapy and universal health insurance data. Treatments ≤12 months from diagnosis and survival were analysed, using adjusted odds ratios (aORs) from logistic regression, and adjusted sub-hazard ratios (aSHRs) from competing risk regression. RESULTS AND CONCLUSION: Five-year disease-specific survival increased to 91% for 2010-2014. Most women had breast surgery (90%), systemic therapy (72%) and radiotherapy (60%). Less treatment applied for ages 80+ vs <50 years (aOR 0.10, 95% CI 0.05-0.20) and TNM stage IV vs stage I (aOR 0.13, 95% CI 0.08-0.22). Surgical treatment increased during the study period and strongly predicted higher survival. Compared with no surgery, aSHRs were 0.31 (95% CI 0.26-0.36) for women having breast-conserving surgery, 0.49 (95% CI 0.41-0.57) for mastectomy and 0.42 (95% CI 0.33-0.52) when both surgery types were received. Patients aged 80+ years had lower survival and less treatment. More trial evidence is needed to optimise trade-offs between benefits and harms in these older women. Survival differences were not found by residential remoteness and were marginal by socioeconomic status.

Power, control, communities and health inequalities. Part II: measuring shifts in power.

In the health field, there is great interest in the role empowerment might play in reducing social inequalities in health. Empowerment is understood here as the processes of developing capabilities that individuals and/or communities need to exercise control over decisions and actions impacting on their lives and health. There is a fundamental problem, however, in identifying and measuring capabilities for collective control that emerge at the level of the collective, with much of the existing literature focusing on individual measures even where community-level processes are concerned. Collective measures need to capture the dynamics of interactions within and between groups, not simply aggregate individual-level measures. This article, Part 2 in a three-part series, takes up the challenge of identifying qualitative markers of capabilities for collective control. We applied the emancipatory power framework (EPF) reported in Part 1 of the series, to qualitative data generated during a longitudinal evaluation of a major English area-based empowerment initiative, the Big Local (BL). We identified empirical 'markers' of shifts towards greater collective control pertaining to each of the 'power' dimensions in the EPF-'power within', 'power with' and 'power to'-and markers of communities exercising 'power over' other institutions/community members. These markers can usefully be applied in the evaluation planning and evaluation of empowerment initiatives. Part 3 in the series uses these markers and a second analytical framework developed during our evaluation of BL to explore how power dynamics unfold in participatory spaces in BL neighbourhoods.

SSBP1 mutations in dominant optic atrophy with variable retinal degeneration.

OBJECTIVE: Autosomal dominant optic atrophy (ADOA) starts in early childhood with loss of visual acuity and color vision deficits. OPA1 mutations are responsible for the majority of cases, but in a portion of patients with a clinical diagnosis of ADOA, the cause remains unknown. This study aimed to identify novel ADOA-associated genes and explore their causality. METHODS: Linkage analysis and sequencing were performed in multigeneration families and unrelated patients to identify disease-causing variants. Functional consequences were investigated in silico and confirmed experimentally using the zebrafish model. RESULTS: We defined a new ADOA locus on 7q33-q35 and identified 3 different missense variants in SSBP1 (NM_001256510.1; c.113G>A [p.(Arg38Gln)], c.320G>A [p.(Arg107Gln)] and c.422G>A [p.(Ser141Asn)]) in affected individuals from 2 families and 2 singletons with ADOA and variable retinal degeneration. The mutated arginine residues are part of a basic patch that is essential for single-strand DNA binding. The loss of a positive charge at these positions is very likely to lower the affinity of SSBP1 for single-strand DNA. Antisense-mediated knockdown of endogenous ssbp1 messenger RNA (mRNA) in zebrafish resulted in compromised differentiation of retinal ganglion cells. A similar effect was achieved when mutated mRNAs were administered. These findings point toward an essential role of ssbp1 in retinal development and the dominant-negative nature of the identified human variants, which is consistent with the segregation pattern observed in 2 multigeneration families studied. INTERPRETATION: SSBP1 is an essential protein for mitochondrial DNA replication and maintenance. Our data have established pathogenic variants in SSBP1 as a cause of ADOA and variable retinal degeneration. ANN NEUROL 2019;86:368-383.

Cancers of the corpus uteri treated in South Australian public hospitals: Trends in clinical management and survival across three decades.

OBJECTIVE: To investigate treatment and survival over three decades. METHODS: Clinical registry data from three major public hospitals analysed using Kaplan-Meier product-limit estimates and multivariate proportional hazard regression to determine disease-specific survival. RESULTS: Five-year survival increased from 75% to 84%. The adjusted hazard ratio (HR, 95% CI) was 0.56 (0.41, 0.77) for 2010-2016 compared with 1984-1989 and was higher for: ages 80+ years; more advanced stages; poorly differentiated tumours; and complex mixed epithelial and mesenchymal tumours and sarcomas. Treatment was by surgery (92%), radiotherapy (33%), chemotherapy (12%) and hormone therapy (10%). Adjusted analyses showed radiotherapy and hormone therapy were less common from 1990 and chemotherapy more common for 2010-2016. Treatment likelihood was lower for ages ≥80 years, mixed epithelial and mesenchymal tumours receiving surgery and chemotherapy, but higher for radiotherapy. Advanced cancers (FIGO stage IV) had less surgery but more non-surgical treatments. Marginal evidence presented of more hormone therapy for high socio-economic areas. CONCLUSIONS: Survival was equivalent to national figures for Australia and the United States, but potentially higher than for England and Wales. Cases aged 80+ years had less care and poorer survival. Findings illustrate the complementary roles of hospital and population-based registries in local service evaluation.

Pre-diagnostic colonoscopies reduce cancer mortality - results from linked population-based data in South Australia.

BACKGROUND: To investigate the association between pre-diagnostic colonoscopy and colorectal cancer mortality in South Australia. METHODS: Colonoscopy histories were obtained for colorectal cancer patients diagnosed in 2003-2013 using linked Medical Benefits Schedule (MBS) claims, hospital-inpatient and cancer-registry data. Colonoscopy histories included the year of colonoscopy, numbers of examinations, and the time from first colonoscopy to diagnosis. Histories of multiple exposures to colonoscopies, and exposures of greater than a year from initial colonoscopy to diagnosis, were regarded as indicators of screening or surveillance activity. Colonoscopies occurring within one year of diagnosis were regarded as more likely to be a response to cancer symptoms than those occurring > 1 year before diagnosis. Associations between colonoscopy history and post-diagnostic survival were analysed using sub-hazard ratios (SHRs) from competing risk regression adjusted for socio-demographic and cancer characteristics. RESULTS: Having pre-diagnostic colonoscopy was associated with an unadjusted reduction in risk of colorectal cancer death of 17% (SHR: 0.83, 95% CI 0.78-0.89). After adjusting for time period and sociodemographic characteristics, the risk of colorectal cancer death reduced by 17% for one pre-diagnostic colonoscopy examination; 27% for two pre-diagnostic colonoscopy examinations; and 45% for three or more pre-diagnostic colonoscopy examinations. Those with a time of over one year from first colonoscopy in the study window to diagnosis, when compared with less than one year, had a 17% lower risk of colorectal cancer death in this adjusted analysis. These reductions were substantially reduced or eliminated when also adjusting for less advanced stage. CONCLUSIONS: Pre-diagnostic colonoscopy, and more so, multiple colonoscopies and first colonoscopy occurring over one year from initial colonoscopy to diagnosis, were associated with longer survival post diagnosis. This was largely explained by less advanced cancer stage at the time of diagnosis.

Using hospital registries in Australia to extend data availability on vulval cancer treatment and survival.

BACKGROUND: The value of hospital registries for describing treatment and survival outcomes for vulval cancer was investigated. Hospital registry data from four major public hospitals in 1984-2016 were used because population-based data lacked required treatment and outcomes data. Unlike population registries, the hospital registries had recorded FIGO stage, grade and treatment. METHODS: Unadjusted and adjusted disease-specific survival and multiple logistic regression were used. Disease-specific survivals were explored using Kaplan-Meier product-limit estimates. Hazards ratios (HRs) were obtained from proportional hazards regression for 1984-1999 and 2000-2016. Repeat analyses were undertaken using competing risk regression. RESULTS: Five-year disease-specific survival was 70%, broadly equivalent to the five-year relative survivals reported for Australia overall (70%), the United Kingdom (70%), USA (72%), Holland (70%), and Germany (Munich) (68%). Unadjusted five-year survival tended to be lower for cancers diagnosed in 2000-2016 than 1984-1999, consistent with survival trends reported for the USA and Canada, but higher for 2000-2016 than 1984-1999 after adjusting for stage and other covariates, although differences were small and did not approach statistical significance (p ≥ 0.40). Surgery was provided as part of the primary course of treatment for 94% of patients and radiotherapy for 26%, whereas chemotherapy was provided for only 6%. Less extensive surgical procedures applied in 2000-2016 than 1984-1999 and the use of chemotherapy increased over these periods. Surgery was more common for early FIGO stages, and radiotherapy for later stages with a peak for stage III. Differences in treatment by surgery and radiotherapy were not found by geographic measures of remoteness and socioeconomic status in adjusted analyses, suggesting equity in service delivery. CONCLUSIONS: The data illustrate the complementary value of hospital-registry data to population-registry data for informing local providers and health administrations of trends in management and outcomes, in this instance for a comparatively rare cancer that is under-represented in trials and under-reported in national statistics. Hospital registries can fill an evidence gap when clinical data are lacking in population-based registries.

Control of cerebellar granule cell output by sensory-evoked Golgi cell inhibition.

Classical feed-forward inhibition involves an excitation-inhibition sequence that enhances the temporal precision of neuronal responses by narrowing the window for synaptic integration. In the input layer of the cerebellum, feed-forward inhibition is thought to preserve the temporal fidelity of granule cell spikes during mossy fiber stimulation. Although this classical feed-forward inhibitory circuit has been demonstrated in vitro, the extent to which inhibition shapes granule cell sensory responses in vivo remains unresolved. Here we combined whole-cell patch-clamp recordings in vivo and dynamic clamp recordings in vitro to directly assess the impact of Golgi cell inhibition on sensory information transmission in the granule cell layer of the cerebellum. We show that the majority of granule cells in Crus II of the cerebrocerebellum receive sensory-evoked phasic and spillover inhibition prior to mossy fiber excitation. This preceding inhibition reduces granule cell excitability and sensory-evoked spike precision, but enhances sensory response reproducibility across the granule cell population. Our findings suggest that neighboring granule cells and Golgi cells can receive segregated and functionally distinct mossy fiber inputs, enabling Golgi cells to regulate the size and reproducibility of sensory responses.

A Low-Molecular-Weight Alginate Oligosaccharide Disrupts Pseudomonal Microcolony Formation and Enhances Antibiotic Effectiveness.

In chronic respiratory disease, the formation of dense, 3-dimensional "microcolonies" by Pseudomonas aeruginosa within the airway plays an important role in contributing to resistance to treatment. An in vitro biofilm model of pseudomonal microcolony formation using artificial-sputum (AS) medium was established to study the effects of low-molecular-weight alginate oligomers (OligoG CF-5/20) on pseudomonal growth, microcolony formation, and the efficacy of colistin. The studies employed clinical cystic fibrosis (CF) isolates (n = 3) and reference nonmucoid and mucoid multidrug-resistant (MDR) CF isolates (n = 7). Bacterial growth and biofilm development and disruption were studied using cell viability assays and image analysis with scanning electron and confocal laser scanning microscopy. Pseudomonal growth in AS medium was associated with increased ATP production (P < 0.05) and the formation (at 48 h) of discrete (>10-µm) microcolonies. In conventional growth medium, colistin retained an ability to inhibit growth of planktonic bacteria, although the MIC was increased (0.1 to 0.4 µg/ml) in AS medium compared to Mueller-Hinton (MH) medium. In contrast, in an established-biofilm model in AS medium, the efficacy of colistin was decreased. OligoG CF-5/20 (≥2%) treatment, however, induced dose-dependent biofilm disruption (P < 0.05) and led to colistin retaining its antimicrobial activity (P < 0.05). While circular dichroism indicated that OligoG CF-5/20 did not change the orientation of the alginate carboxyl groups, mass spectrometry demonstrated that the oligomers induced dose-dependent (>0.2%; P < 0.05) reductions in pseudomonal quorum-sensing signaling. These findings reinforce the potential clinical significance of microcolony formation in the CF lung and highlight a novel approach to treat MDR pseudomonal infections.

Analytical approaches to support current understanding of exposure, uptake and distributions of engineered nanoparticles by aquatic and terrestrial organisms.

Initiatives to support the sustainable development of the nanotechnology sector have led to rapid growth in research on the environmental fate, hazards and risk of engineered nanoparticles (ENP). As the field has matured over the last 10 years, a detailed picture of the best methods to track potential forms of exposure, their uptake routes and best methods to identify and track internal fate and distributions following assimilation into organisms has begun to emerge. Here we summarise the current state of the field, focussing particularly on metal and metal oxide ENPs. Studies to date have shown that ENPs undergo a range of physical and chemical transformations in the environment to the extent that exposures to pristine well dispersed materials will occur only rarely in nature. Methods to track assimilation and internal distributions must, therefore, be capable of detecting these modified forms. The uptake mechanisms involved in ENP assimilation may include a range of trans-cellular trafficking and distribution pathways, which can be followed by passage to intracellular compartments. To trace toxicokinetics and distributions, analytical and imaging approaches are available to determine rates, states and forms. When used hierarchically, these tools can map ENP distributions to specific target organs, cell types and organelles, such as endosomes, caveolae and lysosomes and assess speciation states. The first decade of ENP ecotoxicology research, thus, points to an emerging paradigm where exposure is to transformed materials transported into tissues and cells via passive and active pathways within which they can be assimilated and therein identified using a tiered analytical and imaging approach.

The regulation of copper stress response genes in the polychaete Nereis diversicolor during prolonged extreme copper contamination.

Polychaetes are frequented in toxicological studies, one reason being that some members occupy shallow burrows in sediments and are maximally exposed to the contaminants that accumulate within them. We have been studying one population of the polychaete Nereis (Hediste) diversicolor exhibiting inheritable tolerance to extreme copper contamination in estuarine sediment. Using transcriptome sequencing data we have identified a suite of genes with putative roles in metal detoxification and tolerance, and measured their regulation. Copper tolerant individuals display significantly different gene expression profiles compared to animals from a nearby population living without remarkable copper levels. Gene transcripts encoding principle copper homeostasis proteins including membrane copper ion transporters, copper ion chaperones and putative metallothionein-like proteins were significantly more abundant in tolerant animals occupying contaminated sediment. In contrast, those encoding antioxidants and cellular repair pathways were unchanged. Nontolerant animals living in contaminated sediment showed no difference in copper homeostasis-related gene expression but did have significantly elevated levels of mRNAs encoding Glutathione Peroxidase enzymes. This study represents the first use of functional genomics to investigate the copper tolerance trait in this species and provides insight into the mechanism used by these individuals to survive and flourish in conditions which are lethal to their conspecifics.

A single cell atlas of frozen shoulder capsule identifies features associated with inflammatory fibrosis resolution.

Frozen shoulder is a spontaneously self-resolving chronic inflammatory fibrotic human disease, which distinguishes the condition from most fibrotic diseases that are progressive and irreversible. Using single-cell analysis, we identify pro-inflammatory MERTKlowCD48+ macrophages and MERTK + LYVE1 + MRC1+ macrophages enriched for negative regulators of inflammation which co-exist in frozen shoulder capsule tissues. Micro-cultures of patient-derived cells identify integrin-mediated cell-matrix interactions between MERTK+ macrophages and pro-resolving DKK3+ and POSTN+ fibroblasts, suggesting that matrix remodelling plays a role in frozen shoulder resolution. Cross-tissue analysis reveals a shared gene expression cassette between shoulder capsule MERTK+ macrophages and a respective population enriched in synovial tissues of rheumatoid arthritis patients in disease remission, supporting the concept that MERTK+ macrophages mediate resolution of inflammation and fibrosis. Single-cell transcriptomic profiling and spatial analysis of human foetal shoulder tissues identify MERTK + LYVE1 + MRC1+ macrophages and DKK3+ and POSTN+ fibroblast populations analogous to those in frozen shoulder, suggesting that the template to resolve fibrosis is established during shoulder development. Crosstalk between MerTK+ macrophages and pro-resolving DKK3+ and POSTN+ fibroblasts could facilitate resolution of frozen shoulder, providing a basis for potential therapeutic resolution of persistent fibrotic diseases.

Zoom behavior during visual search modulates pupil diameter and reflects adaptive control states.

Adaptive gain theory proposes that the dynamic shifts between exploration and exploitation control states are modulated by the locus coeruleus-norepinephrine system and reflected in tonic and phasic pupil diameter. This study tested predictions of this theory in the context of a societally important visual search task: the review and interpretation of digital whole slide images of breast biopsies by physicians (pathologists). As these medical images are searched, pathologists encounter difficult visual features and intermittently zoom in to examine features of interest. We propose that tonic and phasic pupil diameter changes during image review may correspond to perceived difficulty and dynamic shifts between exploration and exploitation control states. To examine this possibility, we monitored visual search behavior and tonic and phasic pupil diameter while pathologists (N = 89) interpreted 14 digital images of breast biopsy tissue (1,246 total images reviewed). After viewing the images, pathologists provided a diagnosis and rated the level of difficulty of the image. Analyses of tonic pupil diameter examined whether pupil dilation was associated with pathologists' difficulty ratings, diagnostic accuracy, and experience level. To examine phasic pupil diameter, we parsed continuous visual search data into discrete zoom-in and zoom-out events, including shifts from low to high magnification (e.g., 1× to 10×) and the reverse. Analyses examined whether zoom-in and zoom-out events were associated with phasic pupil diameter change. Results demonstrated that tonic pupil diameter was associated with image difficulty ratings and zoom level, and phasic pupil diameter showed constriction upon zoom-in events, and dilation immediately preceding a zoom-out event. Results are interpreted in the context of adaptive gain theory, information gain theory, and the monitoring and assessment of physicians' diagnostic interpretive processes.

Load Carriage and Physical Exertion Influence Soldier Emotional Responses.

INTRODUCTION: Regular aerobic exercise benefits psychological health, enhancing mood in clinical and nonclinical populations. However, single bouts of exercise exert both positive and negative effects on emotion. Exercise reliably increases emotional arousal. Its effects on emotional valence are thought to depend on an interplay between cognitive and interoceptive factors that change as a function of exercise intensity, as studied in clinical, healthy, and athlete populations. However, special populations, such as military, first responders, and endurance athletes, have unique physical exertion requirements that can coincide with additional cognitive, physical, and environmental stressors not typical of the general population. Load carriage is one such activity. The present study examined emotional valence and arousal during sustained, heavy load carriage akin to military training and operations. METHODS: Thirteen (one woman) active duty soldiers completed a V̇O2max test, a 2-h loaded (up to 50% body mass) and unloaded (empty rucksack) treadmill foot march (3 mph/4% incline) on separate days, during which they rated their exertion and emotional valence and arousal every 40 min. They also completed measures of positive and negative affect and anxiety before and every 20 min after the foot march. RESULTS: Two hours of loaded foot march led to elevated perceived exertion and less positive, more negative and anxious feelings. Higher rated exertion and more negative emotion were associated with higher percent HRmax and V̇O2peak at multiple time points. CONCLUSIONS: These results support affect exertion models such as the Dual Mode Theory, whereby physical exertion becomes less pleasant with increasing intensity, and provide insights into how affective responses applied contexts may help predict time to fatigue or failure.

CD200 ectodomain shedding into the tumor microenvironment leads to NK cell dysfunction and apoptosis.

The basis of immune evasion, a hallmark of cancer, can differ even when cancers arise from one cell type such as in the human skin keratinocyte carcinomas: basal and squamous cell carcinoma. Here we showed that the basal cell carcinoma tumor-initiating cell surface protein CD200, through ectodomain shedding, was responsible for the near absence of NK cells within the basal cell carcinoma tumor microenvironment. In situ, CD200 underwent ectodomain shedding by metalloproteinases MMP3 and MMP11, which released biologically active soluble CD200 into the basal cell carcinoma microenvironment. CD200 bound its cognate receptor on NK cells to suppress MAPK pathway signaling that in turn blocked indirect (IFN-γ release) and direct cell killing. In addition, reduced ERK phosphorylation relinquished negative regulation of PPARγ-regulated gene transcription and led to membrane accumulation of the Fas/FADD death receptor and its ligand, FasL, which resulted in activation-induced apoptosis. Blocking CD200 inhibition of MAPK or PPARγ signaling restored NK cell survival and tumor cell killing, with relevance to many cancer types. Our results thus uncover a paradigm for CD200 as a potentially novel and targetable NK cell-specific immune checkpoint, which is responsible for NK cell-associated poor outcomes in many cancers.

Human intestinal tissue-resident memory T cells comprise transcriptionally and functionally distinct subsets.

Tissue-resident memory T (TRM) cells provide key adaptive immune responses in infection, cancer, and autoimmunity. However, transcriptional heterogeneity of human intestinal TRM cells remains undefined. Here, we investigate transcriptional and functional heterogeneity of human TRM cells through study of donor-derived TRM cells from intestinal transplant recipients. Single-cell transcriptional profiling identifies two transcriptional states of CD8+ TRM cells, delineated by ITGAE and ITGB2 expression. We define a transcriptional signature discriminating these populations, including differential expression of cytotoxicity- and residency-associated genes. Flow cytometry of recipient-derived cells infiltrating the graft, and lymphocytes from healthy gut, confirm these CD8+ TRM phenotypes. CD8+ CD69+CD103+ TRM cells produce interleukin-2 (IL-2) and demonstrate greater polyfunctional cytokine production, whereas ß2-integrin+CD69+CD103- TRM cells have higher granzyme expression. Analysis of intestinal CD4+ T cells identifies several parallels, including a ß2-integrin+ population. Together, these results describe the transcriptional, phenotypic, and functional heterogeneity of human intestinal CD4+ and CD8+ TRM cells.

Carcinosarcomas of the Uterus: Prognostic Factors and Impact of Adjuvant Treatment.

BACKGROUND: Uncertainties remain about the most effective treatment for uterine carcinosarcoma (UCS), a rare but aggressive uterine cancer, due to the limited scope for randomized trials. This study investigates whether nodal excision or adjuvant therapies after hysterectomy offer a survival benefit, using multi-institutional clinical registry data from South Australia. METHODS: Data for all consecutive cases of UCS from 1980 to 2019 were extracted from the Clinical Cancer Registry. Clinical and treatment-related factors associated with disease-specific mortality (DSM) and all-cause mortality (ACM) were determined using multivariable Cox proportional hazards regression, with subgroup analyses by stage. RESULTS: Median follow-up for the 140 eligible cases was 21 months. 94% underwent hysterectomy, and 72% had an additional pelvic lymph node dissection (PLND). Furthermore, 16% received adjuvant chemotherapy; 11% adjuvant radiotherapy and 16% multimodal chemoradiotherapy, with an increase in the latter two modalities over time. DSM was reduced among those who underwent PLND (HR: 0.41; 95%CI: 0.23-0.74), adjuvant chemotherapy (HR: 0.39; 95%CI: 0.18-0.84) or multimodality treatment (HR: 0.11; 95%CI: 0.06-0.30) compared with hysterectomy alone for the whole cohort and for late stage disease (FIGO III/IV) but not for earlier stage disease, except for reduced DSM with multimodal therapy. Findings were similar for ACM. CONCLUSION: Our findings indicate better survival among those who received PLND, chemotherapy and multimodal adjuvant therapy, with the latter applying to early and late stage disease. However, cautious interpretation is warranted, due to potential "indication bias" and limited power. Further research into effective treatment modalities, ideally using prospective study designs, is needed.

Restoration of visual function in advanced disease after transplantation of purified human pluripotent stem cell-derived cone photoreceptors.

Age-related macular degeneration and other macular diseases result in the loss of light-sensing cone photoreceptors, causing irreversible sight impairment. Photoreceptor replacement may restore vision by transplanting healthy cells, which must form new synaptic connections with the recipient retina. Despite recent advances, convincing evidence of functional connectivity arising from transplanted human cone photoreceptors in advanced retinal degeneration is lacking. Here, we show restoration of visual function after transplantation of purified human pluripotent stem cell-derived cones into a mouse model of advanced degeneration. Transplanted human cones elaborate nascent outer segments and make putative synapses with recipient murine bipolar cells (BCs), which themselves undergo significant remodeling. Electrophysiological and behavioral assessments demonstrate restoration of surprisingly complex light-evoked retinal ganglion cell responses and improved light-evoked behaviors in treated animals. Stringent controls exclude alternative explanations, including material transfer and neuroprotection. These data provide crucial validation for photoreceptor replacement therapy and for the potential to rescue cone-mediated vision.