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.

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.

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.

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.

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.

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.

Monitoring TNM stage of female breast cancer and survival across the South Australian population, with national and international TNM benchmarking: A population-based cohort study.

OBJECTIVE: Using linked cancer registry and administrative data to monitor, tumour, node and metastases (TNM) stage and survival from female breast cancer in Australia. METHOD: Analysis of 2000-2014 diagnoses with linked population-based data to investigate: (1) sociodemographic predictors of advanced stage (stages III and IV), using unadjusted and adjusted logistic regression; and (2) sociodemographic factors and stage as predictors of breast cancer survival using competing risk regression. DESIGN: Population-based registry cohort. SETTING AND PARTICIPANTS: 14 759 South Australian women diagnosed in 2000-2014. PRIMARY AND SECONDARY OUTCOME MEASURES: Stage and survival. RESULTS: At diagnosis, 46% of women were classified as stage I, 39% as stage II, 12% as stage III and 4% as stage IV. After adjusting for sociodemographic factors, advanced stage was more common: (1) for ages <50 years; and although not statistically significant, for ages 80+ years; and (2) in women from socioeconomically disadvantaged areas. Compared with 2000-2004 diagnoses, stage and sociodemographic adjusted risks (sub-HRs (SHRs)) of breast cancer death were lower in 2005-2009 (SHR 0.75, 95% CI 0.67 to 0.83) and 2010-2015 (SHR 0.57, 95% CI 0.48 to 0.67). Compared with stage I, the SHR was 3.87 (95% CI 3.32 to 4.53) for stage II, 10.87 (95% CI 9.22 to 12.81) for stage III, and 41.97 (95% CI 34.78 to 50.65) for stage IV. Women aged 70+ years at diagnosis and those living in the most socioeconomically disadvantaged areas were at elevated risk of breast cancer death, independent of stage and sociodemographic factors. CONCLUSIONS: Stage varied by age, diagnostic period and socioeconomic status, and was a stronger predictor of survival than other statistically significant sociodemographic predictors. Achieving earlier diagnosis outside the original BreastScreen target of 50-69 years (as applying <2014) and in residents of socioeconomically disadvantaged areas likely would increase cancer survival at a population level.

Time from diagnosis to treatment of colorectal cancer in a South Australian clinical registry cohort: how it varies and relates to survival.

OBJECTIVES: Some early studies indicated lower survival with longer time from diagnosis to cancer treatment, but others showed the reverse. We investigated time to treatment of colorectal cancer and associations with survival. SETTING AND PARTICIPANTS: Clinical registry data for colorectal cancer cases diagnosed in 2000-2010 at four major public hospitals in South Australia and treated by surgery (n=1675), radiotherapy (n=616) and/or systemic therapy (n=1556). DESIGN: A historic cohort design, with rank-order tests for ordinal clinical and sociodemographic predictors and multiple logistic regression for comparing time from diagnosis to treatment. Unadjusted Kaplan-Meier estimates and adjusted Cox proportional hazards regression were used to investigate disease-specific survival by time to treatment. OUTCOME MEASURES: Time to treatment and survival from diagnosis to death from colorectal cancer. RESULTS: Treatment (any type) commenced for 87% of surgical cases <60 days of diagnosis, with 80% having surgery within this period. Of those receiving radiotherapy, 59% began this treatment <60 days, and of those receiving systemic therapy, the corresponding proportion was 56%. Adjusted analyses showed treatment delay >60 days was more likely for rectal cancers, 2006-2010 diagnoses, residents of northern than other metropolitan regions and for surgery, younger ages <50 years and unexpectedly, those residing closer to metropolitan services. Adjusting for clinical and sociodemographic factors, and diagnostic year, better survival occurred in <2 years from diagnosis for time to treatment >30 days. Survival in the 3-10 years postdiagnosis generally did not differ by time to treatment, except for lower survival for any treatment >90 days for surgical cases. CONCLUSIONS: The lower survival <2 years from diagnosis for treatment <30 days of diagnosis is consistent with other studies attributed to preferencing more complicated cases for earlier care. Lower 3-10 years survival for surgical cases first treated >90 days from diagnosis is consistent with previously reported U-shaped relationships.

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.

Exploring the added value of hospital-registry data for showing local service outcomes: cancers of the ovary, fallopian tube and peritoneum.

OBJECTIVES: To explore the added value of hospital-registry data on invasive epithelial ovarian, tubal and peritoneal cancers. DESIGN: Historic cohort analyses. METHODS: Unadjusted and adjusted regression. SETTING: Major South Australian hospitals. PARTICIPANTS: 1596 women (1984-2015 diagnoses). RESULTS: 5-Year and 10-year survival was 48% and 41%, respectively, equivalent to relative survival for Australia and the USA. After adjusting for age, clinical and geographic factors, risk of ovarian cancer death was 25% lower in 2010-2015 than 1984-1989. Women generally had surgical treatment (87%) in their first round of care. This was more common for younger patients (adjusted OR (95% CIs) 0.17 (0.04 to 0.65) for 80+ vs <40 years) and earlier International Federation of Gynecology and Obstetrics stages (adjusted OR 0.48 (0.13 to 1.78) for stage IIIB/C and 0.13 (0.04 to 0.45) for stage IV vs stage IA). Most (74%) had systemic therapy, which was more common for advanced stages (adjusted ORs >15.0 for stages III and IV vs stage IA). Few (9%) had radiotherapy. Women generally had systemic therapy (74%), without difference by service accessibility and socioeconomic disadvantage, suggesting equity. However, surgery was less common for residents of the most compared with least remote areas (adjusted OR 0.49 (0.24 to 0.99)); and more common prior to adjustment in the highest versus lowest socioeconomic category (unadjusted OR 1.55 (1.01 to 2.39)), but this elevation did not apply after adjustment (adjusted OR 0.19 (0.63 to 2.25)), with the difference largely explained by stage. CONCLUSIONS: Hospital-registry data add value for assessing local service delivery. Equivalent survival to Australia-wide and USA survival, and temporal gains after adjusting for stage and other patient characteristics are reassuring. Survival gains may reflect therapeutic benefits of more extensive surgery and improved chemotherapy regimens.

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.

Targeted disruption of the extracellular polymeric network of Pseudomonas aeruginosa biofilms by alginate oligosaccharides.

Acquisition of a mucoid phenotype by Pseudomonas sp. in the lungs of cystic fibrosis (CF) patients, with subsequent over-production of extracellular polymeric substance (EPS), plays an important role in mediating the persistence of multi-drug resistant (MDR) infections. The ability of a low molecular weight (Mn = 3200 g mol-1) alginate oligomer (OligoG CF-5/20) to modify biofilm structure of mucoid Pseudomonas aeruginosa (NH57388A) was studied in vitro using scanning electron microscopy (SEM), confocal laser scanning microscopy (CLSM) with Texas Red (TxRd®)-labelled OligoG and EPS histochemical staining. Structural changes in treated biofilms were quantified using COMSTAT image-analysis software of CLSM z-stack images, and nanoparticle diffusion. Interactions between the oligomers, Ca2+ and DNA were studied using molecular dynamics (MD) simulations, Fourier transform infrared spectroscopy (FTIR) and isothermal titration calorimetry (ITC). Imaging demonstrated that OligoG treatment (≥0.5%) inhibited biofilm formation, revealing a significant reduction in both biomass and biofilm height (P < 0.05). TxRd®-labelled oligomers readily diffused into established (24 h) biofilms. OligoG treatment (≥2%) induced alterations in the EPS of established biofilms; significantly reducing the structural quantities of EPS polysaccharides, and extracellular (e)DNA (P < 0.05) with a corresponding increase in nanoparticle diffusion (P < 0.05) and antibiotic efficacy against established biofilms. ITC demonstrated an absence of rapid complex formation between DNA and OligoG and confirmed the interactions of OligoG with Ca2+ evident in FTIR and MD modelling. The ability of OligoG to diffuse into biofilms, potentiate antibiotic activity, disrupt DNA-Ca2+-DNA bridges and biofilm EPS matrix highlights its potential for the treatment of biofilm-related infections.

Female breast cancer management and survival: The experience of major public hospitals in South Australia over 3 decades-trends by age and in the elderly.

BACKGROUND: Clinical registry data from major South Australian public hospitals were used to investigate trends in invasive breast-cancer treatment and survival by age. METHODS: Disease-specific survival was calculated for the 1980 to 2013 diagnostic period using Kaplan-Meier product-limit estimates, with a censoring of live cases on December 31, 2014. Cox proportional hazards regression was used to examine differences in survival by age and tumour characteristic. First-round treatments following diagnosis were analysed, using multiple logistic regression to adjust for confounding. RESULTS: Five-year survival increased from 75% in the 1980s to 87% in 2000 to 2013, consistent with national trends, and with increases occurring irrespective of age. There was an increased use of breast conserving surgery, radiotherapy, chemotherapy, and hormone treatments. Five-year survival was lower for women aged 80+ years, increasing from 65% in the 1980s to 74% in 2000 to 2013. Lower survival in these older women persisted after adjusting for TNM stage, other clinical variables, and diagnostic year, without evidence of a reduced disparity over time. Older women were less likely to have surgery, radiotherapy, and chemotherapy throughout 1980 to 2013. By comparison, their use of hormone therapy was elevated. The adjusted relative odds of mastectomy (as opposed to breast conserving surgery) were lower for the 80+ year age range. CONCLUSIONS: Breast-cancer survival increases applied to all ages, including 80+ years, but poorer outcomes persisted in this older group and the gap did not reduce. A key question is whether the best trade-off now exists between optimally therapeutic cancer treatment and accommodations for frailty and co-morbidity in the aged, or whether opportunities exist for better trade-offs and better survival. Local registry data are important for describing local service activity and outcomes by age for local service providers, health administrations and consumer groups; monitoring disparities; and indicating effects of local initiatives.

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.

Breast cancer screening-opportunistic use of registry and linked screening data for local evaluation.

RATIONALE: Screening has been found to reduce breast cancer mortality at a population level in Australia, but these studies did not address local settings where numbers of deaths would generally have been too low for evaluation. Clinicians, administrators, and consumer groups are also interested in local service outcomes. We therefore use more common prognostic and treatment measures and survivals to gain evidence of screening effects among patients attending 4 local hospitals for treatment. AIMS AND OBJECTIVES: To compare prognostic, treatment, and survival measures by screening history to determine whether expected screening effects are occurring. METHODS: Employing routine clinical registry and linked screening data to investigate associations of screening history with these measures, using unadjusted and adjusted analyses. RESULTS: Screened women had a 10-year survival from breast cancer of 92%, compared with 78% for unscreened women; and 79% of screened surgical cases had breast conserving surgery compared with 64% in unscreened women. Unadjusted analyses indicated that recently screened cases had earlier tumor node metastasis stages, smaller diameters, less nodal involvement, better tumor differentiation, more oestrogen and progesterone receptor positive lesions, more hormone therapy, and less chemotherapy. Radiotherapy tended to be more common in screening participants. More frequent use of adjunctive radiotherapy applied when breast conserving surgery was used. CONCLUSIONS: Results confirm the screening effects expected from the scientific literature and demonstrate the value of opportunistic use of available registry and linked screening data for indicating to local health administrations, practitioners, and consumers whether local screening services are having the effects expected.

Colorectal cancer treatment and survival: the experience of major public hospitals in south Australia over three decades.

BACKGROUND: Registry data from four major public hospitals indicate trends in clinical care and survival from colorectal cancer over three decades, from 1980 to 2010. MATERIALS AND METHODS: Kaplan-Meier product- limit estimates and Cox proportional hazards models were used to investigate disease-specific survival and multiple logistic regression analyses to explore first-round treatment trends. RESULTS: Five-year survivals increased from 48% for 1980-1986 to 63% for 2005-2010 diagnoses. Survival increases applied to each ACPS stage (Australian Clinico-Pathological Stage), and particularly stage C (an increase from 38% to 68%). Risk of death from colorectal cancer halved (hazards ratio: 0.50 (0.45, 0.56)) over the study period after adjusting for age, sex, stage, differentiation, primary sub-site, health administrative region, and measures of socioeconomic status and geographic remoteness. Decreases in stage were not observed. Survivals did not vary by sex or place of residence, suggesting reasonable equity in service access and outcomes. Of staged cases, 91% were treated surgically with lower surgical rates for older ages and more advanced stage. Proportions of surgical cases having adjuvant therapy during primary courses of treatment increased for all stages and were highest for stage C (an increase from 5% in 1980-1986 to 63% for 2005-2010). Radiotherapy was more common for rectal than colonic cases. Proportions of rectal cases receiving radiotherapy increased, particularly for stage C where the increase was from 8% in 1980-1986 to 60% in 2005-2010. The percentage of stage C colorectal cases less than 70 years of age having systemic therapy as part of their first treatment round increased from 3% in 1980-1986 to 81% by 1995-2010. Based on survey data on uptake of adjuvant therapy among those offered this care, it is likely that all these younger patients were offered systemic treatment. CONCLUSIONS: We conclude that pronounced increases in survivals from colorectal cancer have occurred at major public hospitals in South Australia due to increases in stage-specific survivals. Use of adjuvant therapies has increased and the patterns of change accord with clinical guideline recommendations. Reasons for sub-optimal use of radiotherapy for rectal cases warrant further investigation, including the potential for limited rural access to impede uptake of treatments at metropolitan-based radiotherapy centres.

Gene therapy restores vision in rd1 mice after removal of a confounding mutation in Gpr179.

The rd1 mouse with a mutation in the Pde6b gene was the first strain of mice identified with a retinal degeneration. However, AAV-mediated gene supplementation of rd1 mice only results in structural preservation of photoreceptors, and restoration of the photoreceptor-mediated a-wave, but not in restoration of the bipolar cell-mediated b-wave. Here we show that a mutation in Gpr179 prevents the full restoration of vision in rd1 mice. Backcrossing rd1 with C57BL6 mice reveals the complete lack of b-wave in a subset of mice, consistent with an autosomal recessive Mendelian inheritance pattern. We identify a mutation in the Gpr179 gene, which encodes for a G-protein coupled receptor localized to the dendrites of ON-bipolar cells. Gene replacement in rd1 mice that are devoid of the mutation in Gpr179 successfully restores the function of both photoreceptors and bipolar cells, which is maintained for up to 13 months. Our discovery may explain the failure of previous gene therapy attempts in rd1 mice, and we propose that Grp179 mutation status should be taken into account in future studies involving rd1 mice.

Developmentally regulated expression of hemoglobin subunits in avascular tissues.

We investigated the spatio-temporal profile of hemoglobin subunit expression in developing avascular tissues. Significant up-regulation of hemoglobin subunits was identified in microarray experiments comparing blastocyst inner cell masses with undifferentiated embryonic stem (ES) cells. Hemoglobin expression changes were confirmed using embryoid bodies (derived from in vitro differentiation of ES cells) to model very early development at pre-vascular stages of embryogenesis; i.e. prior to hematopoiesis. We also demonstrate, using RT-PCR, Western blotting and immunocytochemistry, expression of adult and fetal mouse hemoglobin subunits in the avascular ocular lens at various stages of development and maturation. Hemoglobin proteins were expressed in lens epithelial cells (cytoplasmic) and cortical lens fiber cells (nuclear and cell-surface-associated); however, a sensitive heme assay demonstrated negligible levels of heme in the developing lens postnatally. Hemoglobin expression was also observed in the developing eye in corneal endothelium and retinal ganglion cells. Gut sections showed, in addition to erythrocytes, hemoglobin protein staining in rare, individual villus epithelial cells. These results suggest a paradigm shift: hemoglobin subunits are expressed in the avascular lens and cornea and in pre-hematopoietic embryos. It is likely, therefore, that hemoglobin subunits have novel developmental roles; the absence of the heme group from the lens would indicate that at least some of these functions may be independent of oxygen metabolism. The pattern of expression of hemoglobin subunits in the perinuclear region during lens fiber cell differentiation, when denucleation is taking place, may indicate involvement in the apoptosis-like signaling processes occurring in differentiating lens fiber cells.