Conceptualizing and Developing Competence in Newborn Medicine Among Military Pediatricians.

INTRODUCTION: Competence in neonatal care is especially important for military pediatricians because military pediatricians can be asked to serve in remote duty locations with limited resources. We sought to understand how this competence is defined, developed, and assessed by military pediatric training programs. MATERIALS AND METHODS: After Institutional Review Board approval was obtained, we interviewed educators and recent graduates from every pediatric military training program to construct a shared definition of competence. We then used Kern's Six Steps for curriculum development to understand how competence is taught and assessed. RESULTS: Participants felt that competence for military pediatricians in the neonatal setting meant that learners should be able to provide a full spectrum of newborn care in any military setting. Participants confirmed that this competence was particularly important for military pediatricians because of the possibility of remote duty locations. Participants felt that specific knowledge, skills, and attitudes supported competence. Knowledge domains include distinguishing normal newborns from abnormal newborns, managing normal newborn care, managing common newborn abnormalities, and creating a safe escalation plan for complicated or uncommon newborn abnormalities. Specific skills that support competence are newborn resuscitation, delivery of effective ventilation, and neonatal circumcision. Specific attitudes that support competence are, understanding the personal limits of knowledge and understanding the resources for escalation of care. Educators use a variety of modalities to teach toward competence, including the structured curricula, bedside teaching, and simulation. According to participants, the assessment of learners occurs primarily through narrative assessment and feedback but would ideally occur through direct observation. CONCLUSIONS: Competence in the neonatal setting is particularly important for military pediatricians. Essential skills undergo differential assessment and current assessment methods differ from ideal assessment methods. Future work should focus on how these facets can support a unified curriculum in newborn medicine.

Comparing outcomes among outpatients treated for pyelonephritis with oral cephalosporins versus first-line agents.

Fluoroquinolones and trimethoprim/sulfamethoxazole (TMP-SMX) are first-line agents for acute pyelonephritis. Oral ß-lactams are second-line agents owing to reported lower efficacy rates, primarily seen with aminopenicillins rather than cephalosporins. The increase in resistance rates and adverse effects associated with first-line agents provides justification to reconsider oral cephalosporins for pyelonephritis. Therefore, the objective of this study was to determine whether there is a difference in urinary tract infection (UTI) recurrence rates between oral cephalosporins and first-line agents in the treatment of acute pyelonephritis. This was a retrospective, single-centre, observational cohort study from 1 December 2018 to 31 May 2020. The study population was adult TRICARE beneficiaries with a diagnosis of acute pyelonephritis who were treated with oral antibiotics. The two cohorts compared were first-line antibiotics (ciprofloxacin, levofloxacin and TMP-SMX) and oral cephalosporins. The primary outcome was UTI recurrence rate at 30 days, which was defined as a repeat clinic visit, emergency department visit or hospital admission for a UTI (cystitis or pyelonephritis). The secondary outcome was to determine independent risk factors for UTI recurrence. A total of 268 cephalosporin and 211 first-line cases were included. The primary composite outcome of UTI recurrence within 30 days occurred in 44 (16%) cephalosporin and 36 (17%) first-line cases (P = 0.851). Independent risk factors for UTI recurrence were chronic kidney disease and Klebsiella spp. isolation. In conclusion, there was no significant difference in UTI recurrence rates between oral cephalosporins and first-line agents in the treatment of acute pyelonephritis in the outpatient setting.

Stiffness based enrichment of leukemia cells using microfluidics.

To improve the survival rate of cancer patients, new diagnosis strategies are necessary to detect lower levels of cancer cells before and after treatment regimens. The scarcity of diseased cells, particularly in residual disease after treatment, demands highly sensitive detection approaches or the ability to enrich the diseased cells in relation to normal cells. We report a label-free microfluidic approach to enrich leukemia cells from healthy cells using inherent differences in cell biophysical properties. The microfluidic device consists of a channel with an array of diagonal ridges that recurrently compress and translate flowing cells in proportion to cell stiffness. Using devices optimized for acute T cell leukemia model Jurkat, the stiffer white blood cells were translated orthogonally to the channel length, while softer leukemia cells followed hydrodynamic flow. The device enriched Jurkat leukemia cells from white blood cells with an enrichment factor of over 760. The sensitivity, specificity, and accuracy of the device were found to be > 0.8 . The values of sensitivity and specificity could be adjusted by selecting one or multiple outlets for analysis. We demonstrate that low levels of Jurkat leukemia cells (1 in 10 4 white blood cells) could be more quickly detected using flow cytometry by using the stiffness sorting pre-enrichment. In a second mode of operation, the device was implemented to sort resistive leukemia cells from both drug-sensitive leukemia cells and normal white blood cells. Therefore, microfluidic biomechanical sorting can be a useful tool to enrich leukemia cells that may improve downstream analyses.

Periostin promotes liver fibrogenesis by activating lysyl oxidase in hepatic stellate cells.

Liver fibrosis arises from dysregulated wound healing due to persistent inflammatory hepatic injury. Periostin is a nonstructural extracellular matrix protein that promotes organ fibrosis in adults. Here, we sought to identify the molecular mechanisms in periostin-mediated hepatic fibrosis. Hepatic fibrosis in periostin-/- mice was attenuated as evidenced by significantly reduced collagen fibril density and liver stiffness compared with those in WT controls. A single dose of carbon tetrachloride caused similar acute liver injury in periostin-/- and WT littermates, and we did not detect significant differences in transaminases and major fibrosis-related hepatic gene expression between these two genotypes. Activated hepatic stellate cells (HSCs) are the major periostin-producing liver cell type. We found that in primary rat HSCs in vitro, periostin significantly increases the expression levels and activities of lysyl oxidase (LOX) and lysyl oxidase-like (LOXL) isoforms 1-3. Periostin also induced expression of intra- and extracellular collagen type 1 and fibronectin in HSCs. Interestingly, periostin stimulated phosphorylation of SMAD2/3, which was sustained despite short hairpin RNA-mediated knockdown of transforming growth factor ß (TGFß) receptor I and II, indicating that periostin-mediated SMAD2/3 phosphorylation is independent of TGFß receptors. Moreover, periostin induced the phosphorylation of focal adhesion kinase (FAK) and AKT in HSCs. Notably, siRNA-mediated FAK knockdown failed to block periostin-induced SMAD2/3 phosphorylation. These results suggest that periostin promotes enhanced matrix stiffness in chronic liver disease by activating LOX and LOXL, independently of TGFß receptors. Hence, targeting periostin may be of therapeutic benefit in combating hepatic fibrosis.

Microfluidic cell sorting by stiffness to examine heterogenic responses of cancer cells to chemotherapy.

Cancers consist of a heterogeneous populations of cells that may respond differently to treatment through drug-resistant sub-populations. The scarcity of these resistant sub-populations makes it challenging to understand how to counter their resistance. We report a label-free microfluidic approach to separate cancer cells treated with chemotherapy into sub-populations enriched in chemoresistant and chemosensitive cells based on the differences in cellular stiffness. The sorting approach enabled analysis of the molecular distinctions between resistant and sensitive cells. Consequently, the role of multiple mechanisms of drug resistance was identified, including decreased sensitivity to apoptosis, enhanced metabolism, and extrusion of drugs, and, for the first time, the role of estrogen receptor in drug resistance of leukemia cells. To validate these findings, several inhibitors for the identified resistance pathways were tested with chemotherapy to increase cytotoxicity sevenfold. Thus, microfluidic sorting can identify molecular mechanisms of drug resistance to examine heterogeneous responses of cancers to therapies.

Microfluidic Sorting of Cells by Viability Based on Differences in Cell Stiffness.

The enrichment of viable cells is an essential step to obtain effective products for cell therapy. While procedures exist to characterize the viability of cells, most methods to exclude nonviable cells require the use of density gradient centrifugation or antibody-based cell sorting with molecular labels of cell viability. We report a label-free microfluidic technique to separate live and dead cells that exploits differences in cellular stiffness. The device uses a channel with repeated ridges that are diagonal with respect to the direction of cell flow. Stiff nonviable cells directed through the channel are compressed and translated orthogonally to the channel length, while soft live cells follow hydrodynamic flow. As a proof of concept, Jurkat cells are enriched to high purity of viable cells by a factor of 185-fold. Cell stiffness was validated as a sorting parameter as nonviable cells were substantially stiffer than live cells. To highlight the utility for hematopoietic stem cell transplantation, frozen samples of cord blood were thawed and the purity of viable nucleated cells was increased from 65% to over 94% with a recovery of 73% of the viable cells. Thus, the microfluidic stiffness sorting can simply and efficiently obtain highly pure populations of viable cells.