The Generation of Monoclonal Antibodies that Bind Preferentially to Adrenal Chromaffin Cells and the Cells of Embryonic Sympathetic Ganglia

Adrenal chromaffin cells, sympathetic neurons, and small intensely fluorescent (SIF) cells are each derived from the neural crest, produce catecholamines, and share certain morphological features. These cell types are also partially interconvertible in cell culture (Doupe et al., 1985a,b; Anderson and Axel, 1986). Thus, these cells are said to be members of the sympathoadrenal (SA) lineage and could share a common progenitor. To investigate the origins of this lineage further, we used the cyclophosphamide immuno-suppression method (Matthew and Patterson, 1983) to generate five monoclonal antibodies (SA1-5) that bind strongly to chromaffin cells, with little or no labeling of sympathetic neurons or SIF cells in frozen sections from adult rats. Competition experiments indicate that these antibodies bind to at least three distinct epitopes in tissue sections. The SA antibodies also label most of the cells of embryonic sympathetic ganglia and adrenal primordia. Labeling of sympathetic ganglia appears as the cells initially coalesce and express high levels of tyrosine hydroxylase (TH). Not all TH+ cells in the embryo are SA 1-5+, however; carotid body SIF cells, nodose ganglion TH+ cells, and the transiently TH+ cells in the dorsal root ganglia do not display detectable SA 1-5 labeling. Thus, the expression of these markers for the SA 1-5 lineage is selective. SA antigen expression is hormonally controlled; removal of glucocorticoid and addition of NGF to cultured adrenal chromaffin cells result in the loss of SA 1-5 labeling. These results suggest that the presumed precursors for sympathetic neurons and SIF cells initially express chromaffin cell markers

Isolation of the Progenitor Cells of the Sympathoadrenal Lineage from Embryonic Sympathetic Ganglia with the SA Monoclonal Antibodies

Our previous articles in this series described the production of five monoclonal antibodies (SA1-5) that bind to adrenal chromaffin cells and to cells in embryonic sympathetic ganglia and adrenal primordia (Carnahan and Patterson, 1991), and the downregulation of the sympathoadrenal (SA) antigens in vivo as neuronal markers begin to be expressed (Anderson et al., 1991). These results support the hypothesis that sympathetic neurons and adrenal chromaffin cells are derived from a common embryonic progenitor that displays both neuron- and chromaffin cell-specific markers. We have taken advantage of the fact that at least some of the SA antigens are expressed on the cell surface to isolate SA+ cells from embryonic day 14.5 rat superior cervical, sympathetic ganglia by fluorescence-activated cell sorting. This population of cells is significantly enriched in the expression of markers (tyrosine hydroxylase and neurofilament) found in the putative progenitors in situ. Growth in glucocorticoid maintains the expression of the SA antigens in the sorted cells and induces the chromaffin cell marker enzyme phenylethanolamine N-methyl transferase. In contrast, growth of the sorted cells in basic fibroblast growth factor, NGF, and insulin results in the rapid loss of SA 1 expression and the outgrowth of neurites. The ability to manipulate the fate of the SA+ cells in vitro confirms the suggestion from the in vivo observations that the SA+ cells in the ganglia are at least bipotential progenitors, capable of differentiating along the chromaffin or neuronal pathways

Antibody Markers Identify a Common Progenitor to Sympathetic Neurons and Chromaffin Cells in vivo and Reveal the Timing of Commitment to Neuronal Differentiation in the Sympathoadrenal Lineage

Using specific antibody markers and double-label immunofluorescence microscopy, we have followed the fate of progenitor cells in the sympathoadrenal (SA) sublineage of the neural crest in developing rat embryos. Such progenitors are first recognizable in the primordial sympathetic ganglia at embryonic day 11.5 (E11.5), when they express tyrosine hydroxylase. At this stage, the progenitors also coexpress neuronal markers such as SCG 10 and neurofilament, together with a series of chromaffin cell markers called SA 1–5 (Carnhan and Patterson, 1991 a). The observation of such doubly labeled cells is consistent with the hypothesis that these cells represent a common progenitor to sympathetic neurons and adrenal chromaffin cells. Subsequent to E 11.5, expression of the chromaffin markers is extinguished in the sympathetic ganglia but retained by cells within the adrenal gland. Concomitant with the loss of the SA 1-5 immunoreactivity in sympathetic ganglia, a later sympathetic neuron-specific marker, B2, appears. In dissociated cell suspensions, some B2+ cells that coexpress SA 1 are seen. This implies a switch in the antigenic phenotype of developing sympathetic neurons, rather than a replacement of one cell population by another. The SA 1--B2 transition does not occur for the majority of cells within the adrenal primordium. In vitro, most B2+ cells fail to differentiate into chromaffin cells in response to glucocorticoid. Instead, they continue to extend neurites and then die. Taken together, these data imply that the SA 1--B2 transition correlates with a loss of competence to respond to an inducer of chromaffin differentiation. Thus, the development of SA derivatives is controlled both by environmental signals and by changes in the ability of differentiating cells to respond to such signals

Geoeconomic variations in epidemiology, ventilation management, and outcomes in invasively ventilated intensive care unit patients without acute respiratory distress syndrome: a pooled analysis of four observational studies

Background: Geoeconomic variations in epidemiology, the practice of ventilation, and outcome in invasively ventilated intensive care unit (ICU) patients without acute respiratory distress syndrome (ARDS) remain unexplored. In this analysis we aim to address these gaps using individual patient data of four large observational studies. Methods: In this pooled analysis we harmonised individual patient data from the ERICC, LUNG SAFE, PRoVENT, and PRoVENT-iMiC prospective observational studies, which were conducted from June, 2011, to December, 2018, in 534 ICUs in 54 countries. We used the 2016 World Bank classification to define two geoeconomic regions: middle-income countries (MICs) and high-income countries (HICs). ARDS was defined according to the Berlin criteria. Descriptive statistics were used to compare patients in MICs versus HICs. The primary outcome was the use of low tidal volume ventilation (LTVV) for the first 3 days of mechanical ventilation. Secondary outcomes were key ventilation parameters (tidal volume size, positive end-expiratory pressure, fraction of inspired oxygen, peak pressure, plateau pressure, driving pressure, and respiratory rate), patient characteristics, the risk for and actual development of acute respiratory distress syndrome after the first day of ventilation, duration of ventilation, ICU length of stay, and ICU mortality. Findings: Of the 7608 patients included in the original studies, this analysis included 3852 patients without ARDS, of whom 2345 were from MICs and 1507 were from HICs. Patients in MICs were younger, shorter and with a slightly lower body-mass index, more often had diabetes and active cancer, but less often chronic obstructive pulmonary disease and heart failure than patients from HICs. Sequential organ failure assessment scores were similar in MICs and HICs. Use of LTVV in MICs and HICs was comparable (42·4% vs 44·2%; absolute difference -1·69 [-9·58 to 6·11] p=0·67; data available in 3174 [82%] of 3852 patients). The median applied positive end expiratory pressure was lower in MICs than in HICs (5 [IQR 5-8] vs 6 [5-8] cm H2O; p=0·0011). ICU mortality was higher in MICs than in HICs (30·5% vs 19·9%; p=0·0004; adjusted effect 16·41% [95% CI 9·52-23·52]; p<0·0001) and was inversely associated with gross domestic product (adjusted odds ratio for a US$10 000 increase per capita 0·80 [95% CI 0·75-0·86]; p<0·0001). Interpretation: Despite similar disease severity and ventilation management, ICU mortality in patients without ARDS is higher in MICs than in HICs, with a strong association with country-level economic status