Transfusion of donor-derived red blood cells (RBCs) is the most common form of cell therapy. Production of transfusion-ready cultured RBCs (cRBCs) is a promising replacement for the current, fully donor-dependent therapy. A single transfusion unit, however, contains 2 × 10 12 RBC, which requires large scale production. Here, we report on the scale-up of cRBC production from static cultures of erythroblasts to 3 L stirred tank bioreactors, and identify the effect of operating conditions on the efficiency of the process. Oxygen requirement of proliferating erythroblasts (0.55-2.01 pg/cell/h) required sparging of air to maintain the dissolved oxygen concentration at the tested setpoint (2.88 mg O 2 /L). Erythroblasts could be cultured at dissolved oxygen concentrations as low as 0.7 O 2 mg/ml without negative impact on proliferation, viability or differentiation dynamics. Stirring speeds of up to 600 rpm supported erythroblast proliferation, while 1800 rpm led to a transient halt in growth and accelerated differentiation followed by a recovery after 5 days of culture. Erythroblasts differentiated in bioreactors, with final enucleation levels and hemoglobin content similar to parallel cultures under static conditions.
Dysregulation of the gut microbiome has been shown to perpetuate neuroinflammation, alter intestinal permeability, and modify repetitive mild traumatic brain injury (RmTBI)-induced deficits. However, there have been no investigations regarding the comparative effects that the microbiome may have on RmTBI in adolescents and adults. Therefore, we examined the influence of microbiome depletion prior to RmTBI on microbial composition and metabolome, in adolescent and adult Sprague Dawley rats. Rats were randomly assigned to standard or antibiotic drinking water for 14 days, and to subsequent sham or RmTBIs. The gut microbiome composition and metabolome were analysed at baseline, 1 day after the first mTBI, and at euthanasia (11 days following the third mTBI). At euthanasia, intestinal samples were also collected to quantify tight junction protein (TJP1 and occludin) expression. Adolescents were significantly more susceptible to microbiome depletion via antibiotic administration which increased pro-inflammatory composition and metabolites. Furthermore, RmTBI induced a transient increase in ‘beneficial bacteria’ (Lachnospiraceae and Faecalibaculum) in only adolescents that may indicate compensatory action in response to the injury. Finally, microbiome depletion prior to RmTBI generated a microbiome composition and metabolome that exemplified a potentially chronic pathogenic and inflammatory state as demonstrated by increased Clostridium innocuum and Erysipelatoclostridium and reductions in Bacteroides and Clostridium Sensu Stricto. Results highlight that adolescents are more vulnerable to RmTBI compared to adults and dysbiosis prior to injury may exacerbate secondary inflammatory cascades.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.