Expanded Multiplexing on Sensor-Constrained Microfluidic Partitioning Systems.

Microfluidics can split samples into thousands or millions of partitions, such as droplets or nanowells. Partitions capture analytes according to a Poisson distribution, and in diagnostics, the analyte concentration is commonly inferred with a closed-form solution via maximum likelihood estimation (MLE). Here, we present a new scalable approach to multiplexing analytes. We generalize MLE with microfluidic partitioning and extend our previously developed Sparse Poisson Recovery (SPoRe) inference algorithm. We also present the first in vitro demonstration of SPoRe with droplet digital PCR (ddPCR) toward infection diagnostics. Digital PCR is intrinsically highly sensitive, and SPoRe helps expand its multiplexing capacity by circumventing its channel limitations. We broadly amplify bacteria with 16S ddPCR and assign barcodes to nine pathogen genera by using five nonspecific probes. Given our two-channel ddPCR system, we measured two probes at a time in multiple groups of droplets. Although individual droplets are ambiguous in their bacterial contents, we recover the concentrations of bacteria in the sample from the pooled data. We achieve stable quantification down to approximately 200 total copies of the 16S gene per sample, enabling a suite of clinical applications given a robust upstream microbial DNA extraction procedure. We develop a new theory that generalizes the application of this framework to many realistic sensing modalities, and we prove scaling rules for system design to achieve further expanded multiplexing. The core principles demonstrated here could impact many biosensing applications with microfluidic partitioning.

Malignant ascites in pancreatic cancer: Pathophysiology, diagnosis, molecular characterization, and therapeutic strategies.

Malignant ascites is the accumulation of fluid in the peritoneum as a result of advanced cancer and often signifies the terminal phase of the disease. Management of malignant ascites remains a clinical challenge as symptom palliation is the current standard of cure. Previously, studies examining malignant ascites largely focused on ovarian and gastric cancer. In recent years, there has been a significant increase in research on malignant ascites in pancreatic cancer. Malignant ascites is usually diagnosed based on positive cytology, but cytology is not always diagnostic, indicating the need for novel diagnostic tools and biomarkers. This review aims to summarize the current understanding of malignant ascites in pancreatic cancer and the recent advances in the molecular characterization of malignant ascites fluid from patients with pancreatic cancer including analysis of soluble molecules and extracellular vesicles. Current standard of care treatment options such as paracenteses and diuretics are outlined along with new emerging treatment strategies such as immunotherapy and small-molecule based therapies. New potential investigative directions resulting from these studies are also highlighted.

A rare case of sporadic mismatch repair deficient pancreatic ductal adenocarcinoma that responded to ipilimumab and nivolumab combination treatment: case report.

Background: Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignant disease with a poor prognosis. Despite high efficacy in multiple cancers, immunotherapy has had very little success in treating PDAC due to unfavorable characteristics such as low tumor mutational burden (TMB), low microsatellite instability (MSI), and non-immunogenic tumor microenvironment. Recently, however, there have been reports of rare PDAC cases with high TMB and DNA mismatch repair deficiency (dMMR) that have demonstrated positive response to immunotherapy. All these cases have also presented with Lynch Syndrome, or germline mutations in MMR genes. Case Description: Here, we report a 57-year-old male with stage IV PDAC whose tumor profile revealed high TMB, high MSI, and dMMR, but no germline mutations in genes associated with hereditary cancers including those associated with Lynch Syndrome. After a series of ineffective treatments, the patient showed positive response to combined ipilimumab and nivolumab immunotherapy. To our knowledge, this is the first report of an advanced PDAC case with sporadic dMMR, high TMB, and no Lynch Syndrome having a good response to immunotherapy. Conclusions: This case further supports TMB and high MSI/dMMR being possible biomarkers for immunotherapy of PDAC as well as highlights the importance of both germline and somatic testing of patients with PDAC.

Chylous ascites in colorectal surgery: A systematic review.

BACKGROUND: Chylous ascites is a rare complication in colorectal surgery with limited evidence. AIM: To systematically review all available evidence to describe the incidence, clinical presentation, risk factors and management strategies. METHODS: The systematic review was performed through PubMed, MEDLINE, EMBASE and Cochrane and cross-checked up to November 2020. The data collated included: Demographics, indications (benign vs malignant), site of disease, surgical approach, extent of lymphadenectomy, day to and method of diagnosis of chylous ascites and management strategies. RESULTS: A total of 28 studies were included in the final analysis (426 cases). Patient age ranged from 31 to 89 years. All except one case were performed for malignancy. Of the 426 cases, 195 were right-colonic, 121 left-colonic, 103 pelvic surgeries and 7 others. The majority were diagnosed during the same inpatient stay by recognition of typical drain appearance and increased volume. Three cases were diagnosed during outpatient visits with increased abdominal distention and subsequently underwent paracentesis. Most cases were managed successfully non-operatively (fasting with prolonged drainage, total parenteral nutrition, somatostatin analogues or a combination of these). Only three cases required surgical intervention after failing conservative management and subsequently resolved completely. Risk factors identified include: Right-colonic surgery/ tumour location, extent of lymphadenectomy and number of lymph nodes harvested. CONCLUSION: Chylous ascites after colorectal surgery is a relatively rare complication. Whilst the majority of cases resolved without surgical intervention, preventative measures should be undertaken such as meticulous dissection and clipping of lymphatics during lymphadenectomy to prevent morbidity.

Supercritical carbon dioxide decellularization of plant material to generate 3D biocompatible scaffolds.

The use of plant-based biomaterials for tissue engineering has recently generated interest as plant decellularization produces biocompatible scaffolds which can be repopulated with human cells. The predominant approach for vegetal decellularization remains serial chemical processing. However, this technique is time-consuming and requires harsh compounds which damage the resulting scaffolds. The current study presents an alternative solution using supercritical carbon dioxide (scCO2). Protocols testing various solvents were assessed and results found that scCO2 in combination with 2% peracetic acid decellularized plant material in less than 4 h, while preserving plant microarchitecture and branching vascular network. The biophysical and biochemical cues of the scCO2 decellularized spinach leaf scaffolds were then compared to chemically generated scaffolds. Data showed that the scaffolds had a similar Young's modulus, suggesting identical stiffness, and revealed that they contained the same elements, yet displayed disparate biochemical signatures as assessed by Fourier-transform infrared spectroscopy (FTIR). Finally, human fibroblast cells seeded on the spinach leaf surface were attached and alive after 14 days, demonstrating the biocompatibility of the scCO2 decellularized scaffolds. Thus, scCO2 was found to be an efficient method for plant material decellularization, scaffold structure preservation and recellularization with human cells, while performed in less time (36 h) than the standard chemical approach (170 h).

Laying the foundation: teaching policy and advocacy to medical trainees.

BACKGROUND: A novel and comprehensive two-year health policy curriculum was developed and implemented for family medicine residents at two University of Toronto-affiliated teaching sites. AIM: To evaluate the impact of the curriculum on residents' knowledge of health policy issues, and its usefulness to their learning. METHOD: The evaluation included a pre-post delivery assessment of residents' content-based knowledge of issues in the Canadian healthcare system. Residents were also asked to evaluate the content, process and usefulness of the health policy curriculum. RESULTS: At the end, more than two-thirds of residents had a better understanding of the Canadian healthcare system. The overall pre-post scores showed that residents retained content-based facts in some detail. However, more importantly, residents' positive evaluations of the curriculum indicated they were engaged, enthusiastic and recognized its importance for their learning. CONCLUSION: Despite residents' positive evaluations, questions remain as to how best to assess the success of health policy curricula. Moving beyond the popular pre-post test, less traditional approaches might complement standard program evaluation methods in future. As educators increasingly develop curricula aimed at non-biomedical expertise, we must consider how we can most meaningfully evaluate long-term impact on graduates' approach to clinical practice and their engagement in health system advocacy.