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Developing robust methodology for the sustainable production of red blood cells in vitro is essential for providing an alternative source of clinical-quality blood, particularly for individuals with rare blood group phenotypes. Immortalized erythroid progenitor cell lines are the most promising emergent technology for achieving this goal. We previously created the erythroid cell line BEL-A from bone marrow CD34+ cells that had improved differentiation and enucleation potential compared to other lines reported. In this study we show that our immortalization approach is reproducible for erythroid cells differentiated from bone marrow and also from far more accessible peripheral and cord blood CD34+ cells, consistently generating lines with similar improved erythroid performance. Extensive characterization of the lines shows them to accurately recapitulate their primary cell equivalents and provides a molecular signature for immortalization. In addition, we show that only cells at a specific stage of erythropoiesis, predominantly proerythroblasts, are amenable to immortalization. Our methodology provides a step forward in the drive for a sustainable supply of red cells for clinical use and for the generation of model cellular systems for the study of erythropoiesis in health and disease, with the added benefit of an indefinite expansion window for manipulation of molecular targets., Graphical abstract, Here Daniels et al. demonstrate reproducible immortalization of erythroid cells derived from bone marrow and peripheral and cord blood CD34+ cells and reveal a molecular signature of immortalization. This methodology provides a step toward the sustainable production of red cells for clinical use and for the generation of cellular model systems.

Graphical abstract, In vitro generation of red blood cells has become a goal for scientists globally. Directly, in vitro-generated red blood cells (RBCs) may close the gap between blood supply obtained through blood donation and high demand for therapeutic uses. In addition, the cells obtained can be used as a model for haematologic disorders to allow the study of their pathophysiology and novel treatment discovery. For those reasons, a number of RBC culture systems have been established and shown to be successful; however, the cost of each millilitre of packed RBC is still extremely high. In order to reduce the cost, we aim to see if we can reduce the number of factors used in the existing culture system. In this study, we examined how well haematopoietic stem cells proliferate and differentiate into mature red blood cells with modified culture system. • Absence of extra heparin or insulin or both from the erythroid differentiation media did not affect haematopoietic stem cell proliferation and differentiation. Therefore, we show that the cost and complexity of erythroid culture can be reduced, which may improve the feasibility of in vitro generation of red blood cells.

Highlights • Pneumatosis intestinalis is characterized by the presence of air localizing in the submucosa and subserosa layers of the bowel wall. • Because of its risk of impending perforation, emergency surgery is generally required to be a definite treatment. • Colonic stenting can be used as a safe alternative procedure in the selected patient., Introduction Pneumatosis intestinalis is one of serious conditions following mechanical bowel obstruction. Emergency surgery is generally required to be a definite treatment in these patients of pneumatosis intestinalis, because of its risk of bowel ischemia and perforation. Since the operation in unprepared colon usually resulted in unfavorable outcome, the use of colonic stent is considered one of potential options as a bridge to definitive surgery. Presently, there is no widely published report of using colonic stent in these patients, particularly for stepping to curative surgery. Therefore, we herein report a case of obstructing sigmoid cancer with pneumatosis intestinalis who underwent successfully emergency metallic stent placement to convert from emergency to elective surgery. Presentation of case A 50-year-old woman presented with 3-day history of abdominal pain and obstipation. Abdominal computed tomography demonstrated a short segment of circumferential luminal narrowing at sigmoid colon, the presence of pneumatosis intestinalis at cecum, including ascending colon, and no extraluminal air. We performed colonoscopy and placed the metallic stent. The patient was then improved. After 1 week, the patient underwent elective hand-assisted laparoscopic sigmoidectomy and was discharged 5 days later. Pathological report showed stage IIa sigmoid cancer. The patient had no local recurrence or distant metastasis in 1 year follow up. Conclusion In obstructing colonic patient with pneumatosis intestinalis, nonsurgical treatment by colonic stenting can be used in selected patient as a bridge to definitive surgery. This will result in decreased morbidity and mortality and lower rate of stoma formation.

Background Patient-derived organoids (PDOs) are versatile experimental models for cancer research. At the Faculty of medicine Siriraj Hospital, we have recently established live biobank of over seventy PDO models from stage I-IV Thai colorectal cancer patients. Our biobank included at least 10 patients who underwent neoadjuvant chemotherapy. We aimed to apply this resource for N-of-1 co-clinical trials, for the discovery of novel therapeutic agents and for biomarker development. We report here our application of the developed PDO models for predicting clinical outcome of colorectal cancer patients. Methods Surgically resected colorectal specimens from both normal and cancerous tissue were digested and embedded in appropriate matrices to form PDO models. Subsequently, all successful PDO models were collected in Siriraj live biobank together with their clinical data. Herein, we characterized our derived PDO models by pathological and molecular techniques. Some models were profiled with panel sequencing to assess for the causal driver mutations. Drug sensitivity of each PDO model to standard-of-care drugs were generated and compared with clinical outcome in each patient. Results We successfully established PDO models from different colorectal sites, of different pathological grades and staging. We identified heterogeneous morphologies of PDO models which appear to be corresponding well with the pathological and genomic profiles of each specimen origin. Although the PDO models from different colorectal cancer patients exhibited variable response to 5-fluorouracil (5-Fu), Leucovorin and Oxaliplatin, the response patterns are consistent with the clinical outcome of the individual patients as determined by tumor regression grade or radiologic imaging for neoadjuvant cases. We are examining the use of drug sensitivity profiling of PDO models for prioritizing anti-cancer treatment choices in a prospective clinical study. Conclusions The Siriraj live biobank of PDO from colorectal cancer patients can serve as a useful resource for N-of-1 co-clinical trials, and for comparison of drug response among patients from different ethnic backgrounds. Legal entity responsible for the study Faculty of Medicine Siriraj Hospital, Mahidol University, Thailand. Funding The National Research Council of Thailand and Siriraj foundation, Siriraj Hospital. Disclosure All authors have declared no conflicts of interest.