Longitudinal Monitoring of Intra-Tumoural Heterogeneity Using Optical Barcoding of Patient-Derived Colorectal Tumour Models.

Geno- and phenotypic heterogeneity amongst cancer cell subpopulations are established drivers of treatment resistance and tumour recurrence. However, due to the technical difficulty associated with studying such intra-tumoural heterogeneity, this phenomenon is seldom interrogated in conventional cell culture models. Here, we employ a fluorescent lineage technique termed "optical barcoding" (OBC) to perform simultaneous longitudinal tracking of spatio-temporal fate in 64 patient-derived colorectal cancer subclones. To do so, patient-derived cancer cell lines and organoids were labelled with discrete combinations of reporter constructs, stably integrated into the genome and thus passed on from the founder cell to all its clonal descendants. This strategy enables the longitudinal monitoring of individual cell lineages based upon their unique optical barcodes. By designing a novel panel of six fluorescent proteins, the maximum theoretical subpopulation resolution of 64 discriminable subpopulations was achieved, greatly improving throughput compared with previous studies. We demonstrate that all subpopulations can be purified from complex clonal mixtures via flow cytometry, permitting the downstream isolation and analysis of any lineages of interest. Moreover, we outline an optimized imaging protocol that can be used to image optical barcodes in real-time, allowing for clonal dynamics to be resolved in live cells. In contrast with the limited intra-tumour heterogeneity observed in conventional 2D cell lines, the OBC technique was successfully used to quantify dynamic clonal expansions and contractions in 3D patient-derived organoids, which were previously demonstrated to better recapitulate the heterogeneity of their parental tumour material. In summary, we present OBC as a user-friendly, inexpensive, and high-throughput technique for monitoring intra-tumoural heterogeneity in in vitro cell culture models.

Epidemiology of pregnant patients with major trauma in Victoria.

OBJECTIVE: Trauma is one of the most common contributors to maternal and foetal morbidity and mortality. The aim of the present study was to describe the characteristics and outcomes of major trauma in pregnant patients using a population-based registry. METHODS: Registry-based study using data from the Victorian State Trauma Registry (VSTR), a population-based database of all hospitalised major trauma (death due to injury, Injury Severity Score [ISS] ≥12, admission to an intensive care unit [ICU] for more than 24 h and requiring mechanical ventilation for at least part of their ICU stay or urgent surgery) in Victoria, Australia, from 1 July 2007 to 30 June 2019. Pregnant patients with major trauma were identified on the VSTR. We summarised patient data using descriptive statistics. RESULTS: Over the 12-year study period, there were 63 pregnant major trauma patients. Fifty-two (82.5%) patients sustained injuries resulting from road transport collisions. The maternal survival rate was 98.4% and the foetal survival rate was 88.9%. Thoracic injury was the most common injury (25/63), followed by abdominal injury (23/63). Eighty-six percent of the third trimester patients (19/22) were transported directly to a major trauma service with capacity for definitive care of the pregnancy. CONCLUSION: The present study demonstrated road transport injury was the most common mechanism of injury and both maternal survival rates and foetal survival rates were high. This information is essential for trauma care system planning and public health initiatives to improve the clinical management and outcomes of pregnant women with major trauma.

Threat of COVID-19 impacting on a quaternary healthcare service: a retrospective cohort study of administrative data.

OBJECTIVES: The threat of a pandemic, over and above the disease itself, may have significant and broad effects on a healthcare system. We aimed to describe the impact of the SARS-CoV-2 pandemic (during a relatively low transmission period) and associated societal restrictions on presentations, admissions and outpatient visits. DESIGN: We compared hospital activity in 2020 with the preceding 5 years, 2015-2019, using a retrospective cohort study design. SETTING: Quaternary hospital in Melbourne, Australia. PARTICIPANTS: Emergency department presentations, hospital admissions and outpatient visits from 1 January 2015 to 30 June 2020, n=896 934 episodes of care. INTERVENTION: In Australia, the initial peak COVID-19 phase was March-April. PRIMARY AND SECONDARY OUTCOME MEASURES: Separate linear regression models were fitted to estimate the impact of the pandemic on the number, type and severity of emergency presentations, hospital admissions and outpatient visits. RESULTS: During the peak COVID-19 phase (March and April 2020), there were marked reductions in emergency presentations (10 389 observed vs 14 678 expected; 29% reduction; p<0.05) and hospital admissions (5972 observed vs 8368 expected; 28% reduction; p<0.05). Stroke (114 observed vs 177 expected; 35% reduction; p<0.05) and trauma (1336 observed vs 1764 expected; 24% reduction; p<0.05) presentations decreased; acute myocardial infarctions were unchanged. There was an increase in the proportion of hospital admissions requiring intensive care (7.0% observed vs 6.0% expected; p<0.05) or resulting in death (2.2% observed vs 1.5% expected; p<0.05). Outpatient attendances remained similar (30 267 observed vs 31 980 expected; 5% reduction; not significant) but telephone/telehealth consultations increased from 2.5% to 45% (p<0.05) of total consultations. CONCLUSIONS: Although case numbers of COVID-19 were relatively low in Australia during the first 6 months of 2020, the impact on hospital activity was profound.

Pancreatic cancer: An update on diagnosis and management.

BACKGROUND: Pancreatic cancer has the highest mortality rate among all main cancer types and is the fourth leading cause of cancer death in Australia. OBJECTIVE: This review focuses on the 95% of pancreatic cancers that arise as pancreatic ductal adenocarcinoma, with the aim to summarise current recommendations for diagnosis and treatment. DISCUSSION: No cardinal symptoms for pancreatic cancer exist. Weight loss combined with abdominal symptoms or back pain in individuals aged ≥60 years prompts urgent computed tomography of the abdomen, while individuals aged ≥40 years with jaundice require direct specialist referral. Pancreatic cancer is categorised as resectable, borderline resectable, locally advanced or metastatic. Resectable disease is treated with surgical resection and adjuvant chemotherapy. Borderline resectable and locally advanced disease are treated with neoadjuvant therapy, followed by surgical exploration if the disease is non-progressive. Metastatic and unresectable disease is treated with chemotherapy or best supportive care. Nutritional support is required for most patients.

Management of biliary tract complications after orthotopic liver transplantation.

INTRODUCTION: Despite improved survival, biliary complications remain a significant cause of morbidity following orthotopic liver transplantation. The aim of this study was to review the incidence, treatment and optimum management pathway of biliary complications at the Scottish Liver Transplant Unit. MATERIALS AND METHODS: All patient data were collected prospectively onto a database at the Scottish Liver Transplant Unit with review of hospital records for validation. RESULTS: A total of 379 consecutive orthotopic liver transplants were performed in 333 adult patients between November 1992 and September 2001. Biliary complications occurred in 55 grafts (51 patients) (14.6%) and their incidence decreased with time. Biliary complications occurred in 29 (10.9%) of the 265 choledocho-choledochostomies compared with 14 (25%) of the 56 with T-tubes. Twenty-eight biliary leaks occurred, 22 of which were anastomotic. Seventeen anastomotic leaks were successfully treated non-operatively. Eight patients with biliary leaks subsequently developed an anastomotic stricture. Of the 30 anastomotic strictures, stent insertion was successful in resolving six of 14 (42%) early anastomotic strictures compared with one of 12 (8%) late anastomotic strictures (p = 0.0479). Six (38%) of the 16 early anastomotic strictures required surgery for complete resolution, compared with 12 (86%) of the 14 late anastomotic strictures (p = 0.0106). CONCLUSION: The incidence of biliary complications has decreased with time. The abandonment of choledocho-choledochostomy over a T-tube has been justified. A combination of conservative, endoscopic, and radiological management has been effective in treating biliary leaks and early anastomotic stricture. However endoscopic or radiological stenting was ineffective in the management of late anastomotic strictures, which were best treated by surgical intervention.