Cerebral malaria is a severe neurovascular complication of Plasmodium falciparum with high mortality, even after treatment with effective antimalarials. A better understanding of pathogenic mechanisms could help future development of adjunctive therapies, yet limitations in current experimental models have hindered our knowledge of the disease. We developed a 3D blood-brain barrier model with enhanced barrier properties using primary brain endothelial cells, astrocytes and pericytes. Exposure to parasite egress products increased microvascular permeability, likely due to transcriptional downregulation of junctional and vascular development genes in endothelial cells. In addition, it increased the expression of ferroptosis markers, antigen presentation and type I interferon genes across all BBB cell types. Incubation with cytoadherent schizont-stage P. falciparum -infected erythrocytes induced a similar, but highly localized transcriptional shift, along with inter-endothelial gaps at sites of parasite egress, significantly increasing permeability. These findings provide key insights into the parasite-mediated mechanisms driving brain microvascular pathogenesis in cerebral malaria.

Shiga toxin (Stx) causes significant renal microvascular injury and kidney failure in the pediatric population, and an effective targeted therapy has yet to be demonstrated. Here we established a human kidney microvascular endothelial cell line for the study of Stx mediated injuries with respect to their morphologic, phenotypic, and transcriptional changes, and modeled Stx induced thrombotic microangiopathy (TMA) in flow-mediated 3D microvessels. Distinct from other endothelial cell lines, both isolated primary and immortalized human kidney microvascular endothelial cells demonstrate robust cell-surface expression of the Stx receptor Gb3, and concomitant dose-dependent toxicity to Stx, with significant contributions from caspase-dependent cell death. Use of a glucosylceramide synthase inhibitor (GCSi) to target disruption of the synthetic pathway of Gb3 resulted in remarkable protection of kidney microvascular cells from Stx injury, shown in both cellular morphologies, caspase activation and transcriptional analysis from RNA sequencing. Importantly, these findings are recapitulated in 3D engineered kidney microvessels under flow. Moreover, whole blood perfusion through Stx-treated microvessels led to marked platelet binding on the vessel wall, which was significantly reduced with the treatment of GCSi. These results validate the feasibility and utility of a bioengineered ex vivo human microvascular model under flow to recapitulate relevant blood-endothelial interactions in STEC-HUS. The profound protection afforded by GCSi demonstrates a preclinical opportunity for investigation in human tissue approximating physiologic conditions. Moreover, this work provides a broad foundation for novel investigation into TMA injury pathogenesis and treatment. Insight Box: Shiga toxin (Stx) causes endothelial injury that results in significant morbidity and mortality in the pediatric population, with no effective targeted therapy. This paper utilizes human kidney microvascular cells to examine Stx mediated cell death in both 2D culture and flow-mediated 3D microvessels, with injured microvessels also developing marked platelet binding and thrombi formation when perfused with blood, consistent with the clinical picture of HUS. This injury is abrogated with a small molecule inhibitor targeting the synthetic pathway of the Shiga toxin receptor. Our findings shed light onto Stx-induced vascular injuries and pave a way for broad investigation into thrombotic microangiopathies.
© The Author(s) 2024. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Background Acute rheumatic fever (ARF) and its chronic sequelae, rheumatic heart disease (RHD) contributes to valvular dysfunction and significant cardiovascular disability and endocardial damage is considered the primary pathophysiological mechanism underlying ARF. This study examined peripheral blood markers of endothelial injury and function in ARF and RHD patients and compared them to healthy controls. Method In this prospective observational study, the levels of collagen intermediates, matrix metalloproteinases, tissue inhibitors of matrix metalloproteinases, brain natriuretic peptide, Anti-DNaseB, VEGF, E-selectin, VCAM, and ICAM in circulation were estimated. The study also isolated hemangioblastic and monocytic endothelial progenitor cells and their respective microvesicles from the peripheral blood of patients and control samples. Results Procollagen type I carboxy-terminal propeptide, cross-linked c-telopeptide of type I, and procollagen III c-terminal propeptide levels were higher in RHD subjects compared to patients with ARF. The ARF patients had the highest levels of matrix metalloproteinases 10 (MMP-10) followed by chronic patients and healthy controls. The ratio of tissue inhibitors of matrix metalloproteinases TIMP-1 and MMP-10 was lowest in healthy controls. At the cellular level, there were higher number of monocytic endothelial progenitor cells (EPCs) in ARF subjects as compared to healthy controls. For hemangioblastic EPCs, there was no significant difference between chronic subjects and healthy controls, though their early subtype was higher in chronic subjects. The hemangioblastic EPCs microvesicles were more abundant in ARF compared to RHD patients. Conclusion The greater number of EPCs and respective microvesicles confirm the continued disruption of the endothelium in ARF, and during the progression of the disease, the majority of EPCs undergo apoptosis. Obituary Statement This study was conceived and designed by SR, PU, and Prof. Rajnish Juneja, Professor at AIIMS. Prof. Rajnish Juneja expired in April 2018 while the study was ongoing (1). Mr. Suran Nambisan, a research fellow at NII, was part of the team who initiated the experimental work. Dr. Suran Nambisan expired in January 2023. This paper is dedicated to both of them. It was the profound love for mankind and unwavering dedication to perfection by Prof. Rajnish Juneja that brought together a remarkable team to undertake this study. Dr. Suran Nambisan embarked on his professional research journey by successfully establishing and standardizing a few intricate protocols used in this study.

A comparative analysis of flow-mediated vasodilation (FMD), vasoactive angiogenic, and fibrogenic mediators between treatment-naive and treated systemic sclerosis (SSc) patients is an unmet need. (1)To assess the FMD and different pathogenic mediators in SSc patients about endothelial dysfunction. (2) To assess the proportion of circulating endothelial cells (CECs) in treatment-naïve patients. SSc patients were grouped into treatment-naïve (Group-I, n = 24) on vasodilator (Group-II, n = 10), on vasodilator + immunosuppressive (Group-III, n = 22)]. Age-sex matched healthy controls (n = 20) were included. Endothelial dysfunction (ED) was measured radiologically using FMD. Serum levels of NO, ET1, NO/ET1, sVCAM, sICAM, TGF, IL-6, and VEGF, as well as gene expressions of eNOS, iNOS, ET-1, and TGF, were measured to assess the status of ED in various study groups. CEC was measured in Group-I and HC. CEC was used as a marker to identify a key regulator of ED in SSc. FMD was significantly decreased in all SSc patients through receiving treatment. Upregulation of serum NO and ET concentrations was noted post-treatment with an unaltered NO/ET1 ratio. NO was positively correlated with FMD (r = 0.6) and negatively with TGFβ (r =  - 0.5). ET-1 showed a negative correlation with TGFβ (r =  - 0.5) but no significant correlation with FMD. Circulating endothelial cell (CEC) was significantly higher in Group-I (3.2%) than HC (0.8%) (p = 0.002), and it showed a good correlation with NO (r =  - 0.7, p = 0.0001) and NO/ET1 (r =  - 0.6, p = 0.007). Persistent ED was observed in all SSc patients irrespective of treatment. Dysbalance in NO/ET1 ratio might be the considering factor for the underlying progression of ED. Based on our findings, it may be hypothesized that reduced NO may be a contributing factor in the pathogenesis of endothelial dysfunction in SSc.
© 2022. The Author(s), under exclusive licence to Springer Nature Switzerland AG.

Developmental engineering of living implants from different cell sources capable of stimulating bone regeneration by recapitulating endochondral ossification (ECO) is a promising strategy for large bone defect reconstruction. However, the clinical translation of these cell-based approaches is hampered by complex manufacturing procedures, poor cell differentiation potential, and limited predictive in vivo performance. We developed an adipose tissue-based developmental engineering approach to overcome these hurdles using hypertrophic cartilaginous (HyC) constructs engineered from lipoaspirate to repair large bone defects. The engineered HyC constructs were implanted into 4-mm calvarial defects in nude rats and compared with decellularized bone matrix (DBM) grafts. The DBM grafts induced neo-bone formation via the recruitment of host cells, while the HyC pellets supported bone regeneration via ECO, as evidenced by the presence of remaining cartilage analog and human NuMA-positive cells within the newly formed bone. However, the HyC pellets clearly showed superior regenerative capacity compared with that of the DBM grafts, yielding more new bone formation, higher blood vessel density, and better integration with adjacent native bone. We speculate that this effect arises from vascular endothelial growth factor and bone morphogenetic protein-2 secretion and mineral deposition in the HyC pellets before implantation, promoting increased vascularization and bone formation upon implantation. The results of this study demonstrate that adipose-derived HyC constructs can effectively heal large bone defects and present a translatable therapeutic option for bone defect repair.
© 2022 The Authors. Bioengineering & Translational Medicine published by Wiley Periodicals LLC on behalf of American Institute of Chemical Engineers.