A platform to reproducibly evaluate human colon permeability and damage.

The intestinal epithelium comprises diverse cell types and executes many specialized functions as the primary interface between luminal contents and internal organs. A key function provided by the epithelium is maintenance of a barrier that protects the individual from pathogens, irritating luminal contents, and the microbiota. Disruption of this barrier can lead to inflammatory disease within the intestinal mucosa, and, in more severe cases, to sepsis. Animal models to study intestinal permeability are costly and not entirely predictive of human biology. Here we present a model of human colon barrier function that integrates primary human colon stem cells into Draper's PREDICT96 microfluidic organ-on-chip platform to yield a high-throughput system appropriate to predict damage and healing of the human colon epithelial barrier. We have demonstrated pharmacologically induced barrier damage measured by both a high throughput molecular permeability assay and transepithelial resistance. Using these assays, we developed an Inflammatory Bowel Disease-relevant model through cytokine induced damage that can support studies of disease mechanisms and putative therapeutics.

Breast cancer polygenic risk scores are associated with short-term risk of poor prognosis breast cancer.

PURPOSE: Polygenic risk scores (PRS) for breast cancer may help guide screening decisions. However, few studies have examined whether PRS are associated with risk of short-term or poor prognosis breast cancers. The study purpose was to evaluate the association of the 313 SNP breast cancer PRS with 2-year risk of poor prognosis breast cancer. METHODS: We evaluated the association of breast cancer PRS with breast cancer overall, ER + and ER- breast cancer, and poor prognosis breast cancer diagnosed within 2 years of a negative mammogram among a cohort of 3657 women using logistic regression adjusted for age, breast density, race/ethnicity, year of screening, and genetic ancestry principal components. Breast cancers were considered poor prognosis if they were metastatic, positive lymph nodes, ER/PR + HER2- and > 2 cm, ER/PR/HER2-, or HER2 + and > 1 cm. RESULTS: Of the 308 breast cancers, 137 (44%) were poor prognosis. The overall breast cancer PRS was significantly associated with breast cancer diagnosis within 2 years (OR 1.39, 95% CI 1.23-1.57, p < 0.001). The breast cancer PRS was also associated specifically with diagnosis of poor prognosis disease (OR 1.24, 95% CI 1.03-1.49, p = 0.018), but was more strongly associated with good prognosis cancer (OR 1.52 95% CI 1.29-1.80 p = 3.60 × 10-7) The ER + PRS was significantly associated with ER/PR + breast cancer (OR 1.41, 95% CI 1.24-1.61, p < 0.001) and the ER- PRS was significantly associated with ER- breast cancer (OR 1.48, 95% CI 1.08-2.02, p = 0.015). CONCLUSION: Breast cancer PRS was independently and significantly associated with diagnosis of both breast cancer overall and poor prognosis breast cancer within 2 years of a negative mammogram, suggesting PRS may help guide decisions about screening intervals and supplemental screening.

Mesenchymal stromal cell-derived extracellular vesicles reduce lung inflammation and damage in nonclinical acute lung injury: Implications for COVID-19.

Mesenchymal stem cell derived extracellular vesicles (MSC-EVs) are bioactive particles that evoke beneficial responses in recipient cells. We identified a role for MSC-EV in immune modulation and cellular salvage in a model of SARS-CoV-2 induced acute lung injury (ALI) using pulmonary epithelial cells and exposure to cytokines or the SARS-CoV-2 receptor binding domain (RBD). Whereas RBD or cytokine exposure caused a pro-inflammatory cellular environment and injurious signaling, impairing alveolar-capillary barrier function, and inducing cell death, MSC-EVs reduced inflammation and reestablished target cell health. Importantly, MSC-EV treatment increased active ACE2 surface protein compared to RBD injury, identifying a previously unknown role for MSC-EV treatment in COVID-19 signaling and pathogenesis. The beneficial effect of MSC-EV treatment was confirmed in an LPS-induced rat model of ALI wherein MSC-EVs reduced pro-inflammatory cytokine secretion and respiratory dysfunction associated with disease. MSC-EV administration was dose-responsive, demonstrating a large effective dose range for clinical translation. These data provide direct evidence of an MSC-EV-mediated improvement in ALI and contribute new insights into the therapeutic potential of MSC-EVs in COVID-19 or similar pathologies of respiratory distress.

Relationship of established risk factors with breast cancer subtypes.

BACKGROUND: Breast cancer is a heterogeneous disease, divided into subtypes based on the expression of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2). Subtypes have different biology and prognosis, with accumulating evidence of different risk factors. The purpose of this study was to compare breast cancer risk factors across tumor subtypes in a large, diverse mammography population. METHODS: Women aged 40-84 without a history of breast cancer with a screening mammogram at three United States health systems from 2006 to 2015 were included. Risk factor questionnaires were completed at mammogram visit, supplemented by electronic health records. Invasive tumor subtype was defined by immunohistochemistry as ER/PR+HER2-, ER/PR+HER2+, ER, and PR-HER2+, or triple-negative breast cancer (TNBC). Cox proportional hazards models were run for each subtype. Associations of race, reproductive history, prior breast problems, family history, breast density, and body mass index (BMI) were assessed. The association of tumor subtypes with screen detection and interval cancer was assessed using logistic regression among invasive cases. RESULTS: The study population included 198,278 women with a median of 6.5 years of follow-up (IQR 4.2-9.0 years). There were 4002 invasive cancers, including 3077 (77%) ER/PR+HER2-, 300 (8%) TNBC, 342 (9%) ER/PR+HER2+, and 126 (3%) ER/PR-HER2+ subtype. In multivariate models, Black women had 2.7 times higher risk of TNBC than white women (HR = 2.67, 95% CI 1.99-3.58). Breast density was associated with increased risk of all subtypes. BMI was more strongly associated with ER/PR+HER2- and HER2+ subtypes among postmenopausal women than premenopausal women. Breast density was more strongly associated with ER/PR+HER2- and TNBC among premenopausal than postmenopausal women. TNBC was more likely to be interval cancer than other subtypes. CONCLUSIONS: These results have implications for risk assessment and understanding of the etiology of breast cancer subtypes. More research is needed to determine what factors explain the higher risk of TNBC for Black women.

Risk factors for an advanced breast cancer diagnosis within 2 years of a negative mammogram.

BACKGROUND: Identifying women at risk for advanced interval cancers would allow better targeting of mammography and supplemental screening. The authors assessed risk factors for advanced breast cancer within 2 years of a negative mammogram. METHODS: The authors included 293,520 negative mammograms performed from 2006 to 2015 among 74,736 women. Breast cancers were defined as advanced if they were >2 cm, were >1 cm and triple-negative or human epidermal growth factor receptor 2-positive, had positive lymph nodes, or were metastatic. Cox proportional hazards modeling was used to evaluate associations of age, breast density, menopause, mammogram type, prior breast biopsy, body mass index (BMI), and a family history of breast cancer with a cancer diagnosis within 2 years of a negative mammogram. Models were stratified by year since a negative mammogram. RESULTS: Among 1345 breast cancers, 357 were advanced (26.5%), and 988 (73.5%) were at an early stage. Breast density, prior biopsy, and family history were associated with an increased risk of both advanced and early-stage cancers. Overweight and obese women had a 40% higher risk of early-stage cancer only in year 2 (overweight hazard ratio [HR], 1.41; 95% confidence interval [CI], 1.19-1.67; P < .001; obese HR, 1.41; 95% CI, 1.17-1.70; P < .001). Obese women had a 90% increased risk of advanced cancer in year 1 (HR, 1.90; 95% CI, 1.14-3.18; P = .014), and both overweight and obese women had a 40% or greater increased risk in year 2 (overweight HR, 1.55; 95% CI, 1.14-2.07; P = .005; obese HR, 1.42; 95% CI, 1.00-2.01; P = .051). CONCLUSIONS: A higher BMI was associated with an advanced breast cancer diagnosis within 2 years of a negative mammogram. These results have important implications for risk assessment, screening intervals, and use of supplemental screening.

High-throughput continuous-flow microfluidic electroporation of mRNA into primary human T cells for applications in cellular therapy manufacturing.

Implementation of gene editing technologies such as CRISPR/Cas9 in the manufacture of novel cell-based therapeutics has the potential to enable highly-targeted, stable, and persistent genome modifications without the use of viral vectors. Electroporation has emerged as a preferred method for delivering gene-editing machinery to target cells, but a major challenge remaining is that most commercial electroporation machines are built for research and process development rather than for large-scale, automated cellular therapy manufacturing. Here we present a microfluidic continuous-flow electrotransfection device designed for precise, consistent, and high-throughput genetic modification of target cells in cellular therapy manufacturing applications. We optimized our device for delivery of mRNA into primary human T cells and demonstrated up to 95% transfection efficiency with minimum impact on cell viability and expansion potential. We additionally demonstrated processing of samples comprising up to 500 million T cells at a rate of 20 million cells/min. We anticipate that our device will help to streamline the production of autologous therapies requiring on the order of 10[Formula: see text]-10[Formula: see text] cells, and that it is well-suited to scale for production of trillions of cells to support emerging allogeneic therapies.

Uncovering mutation-specific morphogenic phenotypes and paracrine-mediated vessel dysfunction in a biomimetic vascularized mammary duct platform.

The mammary gland is a highly vascularized tissue capable of expansion and regression during development and disease. To enable mechanistic insight into the coordinated morphogenic crosstalk between the epithelium and vasculature, we introduce a 3D microfluidic platform that juxtaposes a human mammary duct in proximity to a perfused endothelial vessel. Both compartments recapitulate stable architectural features of native tissue and the ability to undergo distinct forms of branching morphogenesis. Modeling HER2/ERBB2 amplification or activating PIK3CA(H1047R) mutation each produces ductal changes observed in invasive progression, yet with striking morphogenic and behavioral differences. Interestingly, PI3KαH1047R ducts also elicit increased permeability and structural disorganization of the endothelium, and we identify the distinct secretion of IL-6 as the paracrine cause of PI3KαH1047R-associated vascular dysfunction. These results demonstrate the functionality of a model system that facilitates the dissection of 3D morphogenic behaviors and bidirectional signaling between mammary epithelium and endothelium during homeostasis and pathogenesis.

Different associations of tumor PIK3CA mutations and clinical outcomes according to aspirin use among women with metastatic hormone receptor positive breast cancer.

INTRODUCTION: The relationships among PIK3CA mutations, medication use and tumor progression remains poorly understood. Aspirin use post-diagnosis may modify components of the PI3K pathway, including AKT and mTOR, and has been associated with lower risk of breast cancer recurrence and mortality. We assessed time to metastasis (TTM) and survival with respect to aspirin use and tumor PIK3CA mutations among women with metastatic breast cancer. METHODS: Patients with hormone receptor positive, HER2 negative (HR+/HER2-) metastatic breast cancer treated in 2009-2016 who received tumor genotyping were included. Aspirin use between primary and metastatic diagnosis was extracted from electronic medical records. TTM and survival were estimated using Cox proportional hazards regression. RESULTS: Among 267 women with metastatic breast cancer, women with PIK3CA mutated tumors had longer TTM than women with PIK3CA wildtype tumors (7.1 vs. 4.7 years, p = 0.008). There was a significant interaction between PIK3CA mutations and aspirin use on TTM (p = 0.006) and survival (p = 0.026). PIK3CA mutations were associated with longer TTM among aspirin non-users (HR = 0.60 95% CI:0.44-0.82 p = 0.001) but not among aspirin users (HR = 1.57 0.86-2.84 p = 0.139). Similarly, PIK3CA mutations were associated with reduced mortality among aspirin non-users (HR = 0.70 95% CI:0.48-1.02 p = 0.066) but not among aspirin users (HR = 1.75 95% CI:0.88-3.49 p = 0.110). CONCLUSIONS: Among women who develop metastatic breast cancer, tumor PIK3CA mutations are associated with slower time to progression and mortality only among aspirin non-users. Larger studies are needed to confirm this finding and examine the relationship among aspirin use, tumor mutation profile, and the overall risk of breast cancer progression.

Performance of Breast Cancer Risk-Assessment Models in a Large Mammography Cohort.

BACKGROUND: Several breast cancer risk-assessment models exist. Few studies have evaluated predictive accuracy of multiple models in large screening populations. METHODS: We evaluated the performance of the BRCAPRO, Gail, Claus, Breast Cancer Surveillance Consortium (BCSC), and Tyrer-Cuzick models in predicting risk of breast cancer over 6 years among 35 921 women aged 40-84 years who underwent mammography screening at Newton-Wellesley Hospital from 2007 to 2009. We assessed model discrimination using the area under the receiver operating characteristic curve (AUC) and assessed calibration by comparing the ratio of observed-to-expected (O/E) cases. We calculated the square root of the Brier score and positive and negative predictive values of each model. RESULTS: Our results confirmed the good calibration and comparable moderate discrimination of the BRCAPRO, Gail, Tyrer-Cuzick, and BCSC models. The Gail model had slightly better O/E ratio and AUC (O/E = 0.98, 95% confidence interval [CI] = 0.91 to 1.06, AUC = 0.64, 95% CI = 0.61 to 0.65) compared with BRCAPRO (O/E = 0.94, 95% CI = 0.88 to 1.02, AUC = 0.61, 95% CI = 0.59 to 0.63) and Tyrer-Cuzick (version 8, O/E = 0.84, 95% CI = 0.79 to 0.91, AUC = 0.62, 95% 0.60 to 0.64) in the full study population, and the BCSC model had the highest AUC among women with available breast density information (O/E = 0.97, 95% CI = 0.89 to 1.05, AUC = 0.64, 95% CI = 0.62 to 0.66). All models had poorer predictive accuracy for human epidermal growth factor receptor 2 positive and triple-negative breast cancers than hormone receptor positive human epidermal growth factor receptor 2 negative breast cancers. CONCLUSIONS: In a large cohort of patients undergoing mammography screening, existing risk prediction models had similar, moderate predictive accuracy and good calibration overall. Models that incorporate additional genetic and nongenetic risk factors and estimate risk of tumor subtypes may further improve breast cancer risk prediction.

Breastfeeding indicators among a nationally representative multi-ethnic sample of New Zealand children.

AIMS: To describe breastfeeding initiation and duration, and demographic associations with breastfeeding duration within a representative sample of New Zealand infants. METHODS: In 6,685 singletons enrolled in the Growing Up in New Zealand cohort we described breastfeeding initiation (96%), any (94%) and exclusive (93%) breastfeeding (EBF) duration. We used adjusted relative risk (RR) and 95% confidence intervals (CI) to describe associations with breastfeeding duration. RESULTS: Breastfeeding initiation occurred for 97%. Sixteen percent were EBF to age six months and 13% were breastfed to age 24 months. Exclusive breastfeeding for ≥4 months was less likely for children of mothers of Maori (RR=0.80, 95% CI 0.73-0.87), Pacific (0.90, 95% CI 0.83-0.98) or Asian (0.80, 95% CI 0.74-0.86) ethnicity. Children of mothers aged 20-29 years (1.24, 95% CI 1.04-1.49); ≥30 years (1.36, 95% CI 1.14-1); with a tertiary education (1.14, 95% CI 1.08-1.21); or planned pregnancy (1.14, 95% CI 1.08-1.21); and children with older siblings (RR=1.31, 95% CI 1.17-1.47) were more likely to be exclusively breastfed for ≥4 months. Children were more likely to be breastfed ≥6 months if their mother was aged 20-29 (1.26, 95% CI 1.10-1.45) or ≥30 years (1.40, 95% CI 1.22-1.61), had a tertiary education (1.11, 95% CI 1.06-1.59) or planned pregnancy (1.11, 95% CI 1.06-1.15), or if they had older siblings (1.04, 95% CI 1.00-1.08). CONCLUSION: In New Zealand, most children are initially breastfed, however a large proportion did not receive the recommended duration of any or exclusive breastfeeding. Maternal age, education, parity and pregnancy planning identify children at risk of shorter duration of breastfeeding and EBF, and maternal ethnicity identifies children at risk of shorter EBF duration.