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Hematopoietic stem cells (HSCs) from different sources show varied repopulating capacity, and HSCs lose their stemness after long-time ex vivo culture. However, the underlying mechanisms of the stemness differences because of the cell sources and the culture stimulation are not fully understood. Here, we applied single-cell RNA-seq (scRNA-seq) to analyze the naïve and stimulated human CD34+ cells from cord blood (CB) and (mPB). We collected over 16,000 single-cell data to construct a comprehensive trajectory inference map and characterized the HSC population on the hierarchy top, which is under quiescent state. Then we compared HSCs in CB to those in mPB and HSCs of naïve samples to those of cultured samples, and identified stemness-related genes (SRGs) associated with culture time (CT-SRGs) and cell source (CS-SRGs), respectively. Interestingly, CT-SRGs and CS-SRGs share genes enriched in the signaling pathways such as mRNA catabolic process, Translational initiation, Ribonucleoprotein complex biogenesis and Cotranslational protein targeting to membrane, suggesting dynamic protein translation and processing may be a common requirement for stemness maintenance. Meanwhile, CT-SRGs are enriched in pathways involved in glucocorticoid and corticosteroid response that affect HSCs homing and engraftment. In contrast, CS-SRGs specifically contain genes related purine and ATP metabolic process which is important to initiate hematopoiesis. Finally, we presented an application through a small-scale drug screening using Connectivity Map (CMap) against CT-SRGs and found a small molecule cucurbitacin I, targeting STAT3/JAK2, can efficiently expand HSCs ex vivo while maintaining its stemness. These results indicate SRGs revealed by scRNA-seq can provide helpful insights to understand the stemness differences under diverse circumstances, and CT-SRGs can be a valuable database to identify candidates enhancing functional HSC expansion during ex vivo culture.

Sexual reproduction plays an important role in the population and community structure maintenance in scleractinian corals, particularly in growth and development after larvae settlement. Despite this, only few studies have been conducted to monitor the post-settlement growth and development of scleractinian coral larvae. This study aimed to obtain this data by collecting adult Galaxea fascicularis colonies from Weizhou Island coast (21°00′-21°10′ N, 109°00′-109°15′ E) and recording their complete life cycle, from fertilization to recruitment, for one year ex situ. The results demonstrated that G. fascicularis could reproduce sexually when reared in a tank for 2 y. Their embryo produced a "prawn chip" that lacked a blastocoel, indicating that G. fascicularis is a complex coral. Larvae evidently showed association with zooxanthellae four days after settlement and completed metamorphosis after 1 month. The mean diameter of G. fascicularis recruits was 4.74 ± 1.12 mm, and the survival rate was only 5.60% in the rst year, which may be attributed to their competition with algae. To increase their survival rate without the interference of algae, we suggest that the recruits of G. fascicularis be reared ex situ to complete metamorphosis (1 month) and then transferred to the eld. This study improved our understanding of the early life cycle of scleractinian corals.

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