Background: Heterogeneity of endothelial cells (ECs) is a hallmark of the vascular system which may impact the development and management of vascular disorders. Despite the tremendous progress in differentiation of human embryonic stem cells (hESCs) towards endothelial lineage, differentiation into arterial and venous endothelial phenotypes remains elusive. Additionally, current differentiation strategies are hampered by inefficiency, lack of reproducibility, and use of animal-derived products.Methods: To direct the differentiation of hESCs to endothelial subtypes, H1- and H9-hESCs were seeded on human plasma fibronectin and differentiated under chemically defined conditions by sequential modulation of glycogen synthase kinase-3 (GSK-3), basic fibroblast growth factor (bFGF), bone morphogenetic protein 4 (BMP4) and vascular endothelial growth factor (VEGF) signaling pathways for 5 days. Following the initial differentiation, the endothelial progenitor cells (CD34⁺CD31⁺cells) were sorted and terminally differentiated under serum-free conditions to arterial and venous ECs. The transcriptome and secretome profiles of the two distinct populations of hESC-derived arterial and venous ECs were characterized. Furthermore, the safety and functionality of these cells upon in vivo transplantation were characterized.Results: Sequential modulation of hESCs with GSK-3 inhibitor, bFGF, BMP4 and VEGF resulted in stages reminiscent of primitive streak, early mesoderm/lateral plate mesoderm, and endothelial progenitors under feeder- and serum-free conditions. Furthermore, these endothelial progenitors demonstrated differentiation potential to almost pure populations of arterial and venous endothelial phenotypes under serum-free conditions. Specifically, the endothelial progenitors differentiated to venous ECs in the absence of VEGF, and to arterial phenotype under low concentrations of VEGF. Additionally, these hESC-derived arterial and venous ECs showed distinct molecular and functional profiles in vitro. Furthermore, these hESC-derived arterial and venous ECs were nontumorigenic and were functional in terms of forming perfused microvascular channels upon subcutaneous implantation in the mouse.Conclusions: We report a simple, rapid, and efficient protocol for directed differentiation of hESCs into endothelial progenitor cells capable of differentiation to arterial and venous ECs under feeder-free and serum-free conditions. This could offer a human platform to study arterial–venous specification for various applications related to drug discovery, disease modeling and regenerative medicine in the future.

This study aimed to answer the research question: How reliable is ChatGPT in automated essay scoring (AES) for oral and maxillofacial surgery (OMS) examinations for dental undergraduate students compared to human assessors? Sixty-nine undergraduate dental students participated in a closed-book examination comprising two essays at the National University of Singapore. Using pre-created assessment rubrics, three assessors independently performed manual essay scoring, while one separate assessor performed AES using ChatGPT (GPT-4). Data analyses were performed using the intraclass correlation coefficient and Cronbach's α to evaluate the reliability and inter-rater agreement of the test scores among all assessors. The mean scores of manual versus automated scoring were evaluated for similarity and correlations. A strong correlation was observed for Question 1 (r = 0.752–0.848, p < 0.001) and a moderate correlation was observed between AES and all manual scorers for Question 2 (r = 0.527–0.571, p < 0.001). Intraclass correlation coefficients of 0.794–0.858 indicated excellent inter-rater agreement, and Cronbach’s α of 0.881–0.932 indicated high reliability. For Question 1, the mean AES scores were similar to those for manual scoring (p > 0.05), and there was a strong correlation between AES and manual scores (r = 0.829, p < 0.001). For Question 2, AES scores were significantly lower than manual scores (p < 0.001), and there was a moderate correlation between AES and manual scores (r = 0.599, p < 0.001). This study shows the potential of ChatGPT for essay marking. However, an appropriate rubric design is essential for optimal reliability. With further validation, the ChatGPT has the potential to aid students in self-assessment or large-scale marking automated processes.

Three-dimensional (3D) biofabrication techniques that enable the production of multicellular tissue equivalents for applications in basic biology, drug screening and regenerative medicne. Fabrication of these tissue constructs with in-built microvasculature enables recapitulation of the biological environment of the native tissues. Here, we present the fabrication of 3D vascularized tissue constructs containing microvascular networks using human embryonic stem cell (hESC)-derived endothelial cells (ECs) and pericytes encapsulated within a fibrin-based matrix and cultured under chemically defined conditions. Firstly, by manipulating the developmental signaling pathways under chemically defined culture conditions, hESCs were efficiently differentiated to hESC-ECs and hESC-pericytes through intermediate stages of lateral plate and paraxial mesoderm respectively. Next, encapsulation of these hESC-derived vascular cells within fibrin-based matrix and culture under chemically defined conditions, result in self-assembly of hESC-ECs into a network of microvessels within a period of 6–9 d. With the supporting influence of hESC-pericytes, the microvascular network with lumen was stable for at least 3 weeks. Quantification of the fractal dimensions of the microvascular networks demonstrate the increasing complexity of the vascular network with increasing endothelial cell densities. Dextran permeation studies in the presence or absence of vasodilating agent (histamine) showed the presence of hollow lumen, modulation of barrier properties of the microvasculature and its functional response to histamine. Hence, this versatile in vitro 3D model of vascularized constructs generated under chemically defined conditions is well suited to study early angiogenesis for in vitro drug testing applications and provide a clinically amenable, fundamental step towards fabrication of complex and functional tissues for regenerative applications in the future.

Periodontitis involves complex interplay of bacteria and host immune response resulting in destruction of supporting tissues of the tooth. Toll‐like receptors (TLRs) play a role in recognizing microbial pathogens and eliciting an innate immune response. Recently, the potential application of multipotent stem cells and pluripotent stem cells including human embryonic stem cells (hESCs) in periodontal regenerative therapy has been proposed. However, little is known about the impact of periodontopathogens on hESC‐derived progenies. This study investigates the effects of heat‐killed periodontopathogens, namely, Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans, on TLR and cytokine expression profile of hESC‐derived progenies, namely, fibroblasts (hESC‐Fib) and mesenchymal stem cells (hESC‐MSCs). Additionally, the serotype‐dependent effect of A. actinomycetemcomitans on hESC‐derived progenies was explored. Both hESC‐Fib and hESC‐MSCs constitutively expressed TLR-2 and TLR-4. hESC‐Fib upon exposure to periodontopathogens displayed upregulation of TLRs and release of cytokines (IL‐1β, IL‐6, and IL‐8). In contrast, hESC‐MSCs were largely nonresponsive to bacterial challenge, especially in terms of cytokine production. Further, exposure of hESC‐Fib to A. actinomycetemcomitans serotype c was associated with higher IL‐8 production than serotype b. In contrast, the hESC‐MSCs displayed no serotype‐dependent response. Differential response of the two hESC progenies implies a phenotype‐dependent response to periodontopathogens and supports the concept of immunomodulatory properties of MSCs.

Background: Jaw osteoradionecrosis (ORN) is a complication in patients with previous head and neck radiotherapy. Its incidence increases with dental extractions. Hence, this review aimed to evaluate the efficacy of adjunctive treatment modalities undertaken at the time of extraction in previous head and neck radiotherapy patients in preventing ORN. Methods: A systematic review was conducted, where studies with data on ORN incidence after extraction with or without adjunctive interventions were included. Meta‐analyses were conducted to estimate the pooled prevalence of ORN per intervention and the pooled odds ratio for incidence of ORN between interventions. Results: In total, 1520 patients in 29 studies were included. Interventions identified were hyperbaric oxygen (HBO), pentoxifylline‐tocopherol (PENTO), antibiotics (ABX), platelet‐rich fibrin and photobiomodulation. The pooled prevalence of ORN for HBO (4.6%), PENTO (3.4%) and ABX (3.8%) was significantly lower than the Control (17.6%). For studies with direct comparisons between groups, HBO had lower but not significant odds of developing ORN than the Control (OR 0.27) and ABX (OR 0.57). Conclusions: HBO, PENTO and ABX may reduce the incidence of ORN compared to no intervention. Given that all three have similar incidences of ORN, ABX may be the most cost‐effective and accessible adjunctive modality.