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The tubular gland of the chicken oviduct is an attractive system for protein expression as large quantities of proteins are deposited in the egg, the production of eggs is easily scalable and good manufacturing practices for therapeutics from eggs have been established. Here we examined the ability of upstream and downstream DNA sequences of ovalbumin, a protein produced exclusively in very high quantities in chicken egg white, to drive tissue-specific expression of human mAb in chicken eggs. To accommodate these large regulatory regions, we established and transfected lines of chicken embryonic stem (cES) cells and formed chimeras that express mAb from cES cell-derived tubular gland cells. Eggs from high-grade chimeras contained up to 3 mg of mAb that possesses enhanced antibody-dependent cellular cytotoxicity (ADCC), nonantigenic glycosylation, acceptable half-life, excellent antigen recognition and good rates of internalization.

SummaryThe gap in knowledge of the molecular mechanisms underlying differentiation of human pluripotent stem cells (hPSCs) into the mesenchymal cell lineages hinders the application of hPSCs for cell-based therapy. In this study, we identified a critical role of muscle segment homeobox 2 (MSX2) in initiating and accelerating the molecular program that leads to mesenchymal stem/stromal cell (MSC) differentiation from hPSCs. Genetic deletion of MSX2 impairs hPSC differentiation into MSCs. When aided with a cocktail of soluble molecules, MSX2 ectopic expression induces hPSCs to form nearly homogeneous and fully functional MSCs. Mechanistically, MSX2 induces hPSCs to form neural crest cells, an intermediate cell stage preceding MSCs, and further differentiation by regulating TWIST1 and PRAME. Furthermore, we found that MSX2 is also required for hPSC differentiation into MSCs through mesendoderm and trophoblast. Our findings provide novel mechanistic insights into lineage specification of hPSCs to MSCs and effective strategies for applications of stem cells for regenerative medicine.

U lcerative colitis is a chronic, relapsing inflammatory disease of the colon, leading to a significant burden and disability for patients. [1][2][3][4] Current therapeutic options include mesalamine, glucocorticoids, immunosuppressive agents, and biologics. However, these available treatments are not effective in more than one third of patients and can be associated with adverse effects that limit their use. [5][6][7][8] New treatments are needed to provide sustained improvements in symptomatic and endoscopic outcomes in a higher proportion of patients with ulcerative colitis. 9 Janus kinases (JAK1, JAK2, JAK3, and tyrosine kinase [TYK] 2) are intracellular TYKs. They are activated by binding of a cytokine ligand, leading to recruitment, phosphorylation, and activation of signal transducers and activators of transcription, 10,11 which control many functions of innate and adaptive immunity, hematopoiesis, and cellular processes, including cell growth, survival, differentiation, and migration. 11 Increasingly, JAK inhibition has been evaluated as a target for the management of many immunemediated diseases, including ulcerative colitis. Tofacitinib, a pan-JAK inhibitor, has shown efficacy in 3 phase 3 placebocontrolled studies in patients with moderately to severely active ulcerative colitis and has been approved for the treatment of ulcerative colitis. 12 Upadacitinib is a once daily, oral, small-molecule therapy that was engineered to have increased selectivity for JAK1 over JAK2, JAK3, and TYK2. 13 We report the results of a phase 2b trial, investigating the dose response, efficacy, and safety of upadacitinib in patients with moderately to severely active ulcerative colitis. Methods Trial Design and OversightThe overarching U-ACHIEVE program comprises 3 studies: a phase 2b dose-ranging induction study (study 1), a phase 3 doseconfirming induction study (study 2), and a phase 3 maintenance study (study 3). Here, we report the results of the primary and secondary efficacy endpoints and safety from study 1. This was a multicenter, randomized, double-blind, placebo-controlled trial, conducted from October 2016 through April 2018, at 142 sites in 28 countries. A total of 250 patients were randomized in study 1 part 1; after the enrollment in this study part was completed, an additional 132 patients were enrolled in study 1 part 2 and were randomly assigned into groups receiving upadacitinib 30 mg and 45 mg once daily to avoid interrupting the study activities and to provide a sufficient number of clinical responders for the maintenance portion of the study. An exploratory analysis for the combined results of study 1 parts 1 and 2 is provided in the Supplementary Materials (Supplementary Tables 1-4). The complete study design of study 1 is shown in Supplementary Figure 1.This study was conducted per the International Conference on Harmonisation guidelines, applicable regulations, and the Declaration of Helsinki. The trial was registered at ClinicalTrials.gov (NCT02819635). The protocol was approved by institutio...

SummaryHuman pluripotent stem cells (hPSCs) provide an unlimited source for generating various kinds of functional blood cells. However, efficient strategies for generating large-scale functional blood cells from hPSCs are still lacking, and the mechanism underlying human hematopoiesis remains largely unknown. In this study, we identified myeloid ectopic viral integration site 1 homolog (MEIS1) as a crucial regulator of hPSC early hematopoietic differentiation. MEIS1 is vital for specification of APLNR+ mesoderm progenitors to functional hemogenic endothelial progenitors (HEPs), thereby controlling formation of hematopoietic progenitor cells (HPCs). TAL1 mediates the function of MEIS1 in HEP specification. In addition, MEIS1 is vital for megakaryopoiesis and thrombopoiesis from hPSCs. Mechanistically, FLI1 acts as a downstream gene necessary for the function of MEIS1 during megakaryopoiesis. Thus, MEIS1 controls human hematopoiesis in a stage-specific manner and can be potentially manipulated for large-scale generation of HPCs or platelets from hPSCs for therapeutic applications in regenerative medicine.

Background Gut microbiome alterations are closely related to human health and linked to a variety of diseases. Although great efforts have been made to understand the risk factors for multiple myeloma (MM), little is known about the role of the gut microbiome and alterations of its metabolic functions in the development of MM. Results Here, in a cohort of newly diagnosed patients with MM and healthy controls (HCs), significant differences in metagenomic composition were discovered, for the first time, with higher bacterial diversity in MM. Specifically, nitrogen-recycling bacteria such as Klebsiella and Streptococcus were significantly enriched in MM. Also, the bacteria enriched in MM were significantly correlated with the host metabolome, suggesting strong metabolic interactions between microbes and the host. In addition, the MM-enriched bacteria likely result from the regulation of urea nitrogen accumulated during MM progression. Furthermore, by performing fecal microbiota transplantation (FMT) into 5TGM1 mice, we proposed a mechanistic explanation for the interaction between MM-enriched bacteria and MM progression via recycling urea nitrogen. Further experiments validated that Klebsiella pneumoniae promoted MM progression via de novo synthesis of glutamine in mice and that the mice fed with glutamine-deficient diet exhibited slower MM progression. Conclusions Overall, our findings unveil a novel function of the altered gut microbiome in accelerating the malignant progression of MM and open new avenues for novel treatment strategies via manipulation of the intestinal microbiota of MM patients.

High throughput single-cell RNA-seq has been successfully implemented to dissect the cellular and molecular features underlying hematopoiesis. However, an elaborate and comprehensive transcriptome reference of the whole blood system is lacking. Here, we profiled the transcriptomes of 7,551 human blood cells representing 32 immunophenotypic cell types, including hematopoietic stem cells, progenitors and mature blood cells derived from 21 healthy donors. With high sequencing depth and coverage, we constructed a single-cell transcriptional atlas of blood cells (ABC) on the basis of both protein-coding genes and long noncoding RNAs (lncRNAs), and showed a high consistence between them. Notably, putative lncRNAs and transcription factors regulating hematopoietic cell differentiation were identified. While common transcription factor regulatory networks were activated in neutrophils and monocytes, lymphoid cells dramatically changed their regulatory networks during differentiation. Furthermore, we showed a subset of nucleated erythrocytes actively expressing immune signals, suggesting the existence of erythroid precursors with immune functions. Finally, a web portal offering transcriptome browsing and blood cell type prediction has been established. Thus, our work provides a transcriptional map of human blood cells at single-cell resolution, thereby offering a comprehensive reference for the exploration of physiological and pathological hematopoiesis.

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