Polymyxin-induced neuromuscular weakness: a case report.

Polymyxin-induced neuromuscular blockade is a rare but potentially fatal condition, with majority of cases that were reported between 1962 and 1973. We describe a patient who developed hypercapnic respiratory failure after initiation of polymyxin for multi-drug resistant Escherichia Coli bacteremia, due to polymyxin-induced neuromuscular dysfunction. After cessation of polymyxin, he regained full strength, had complete resolution of ptosis, and was successfully extubated. In light of the renewed use of polymyxin in this era of antimicrobial-resistance, this case aims to raise awareness about this rare but life-threatening condition, which is easily reversible with early recognition and prompt discontinuation of the drug.

Factors associated with ablation-related thrombus extension following microfoam versus radiofrequency saphenous vein closure.

OBJECTIVE: Polidocanol endovenous microfoam ablation (MFA) is approved by the US Food and Drug Administration for great saphenous vein (GSV) closure, yet there are few published data on the subsequent risk of ablation-related thrombus extension (ARTE). Recent societal practice guidelines recommend against routine postprocedure duplex ultrasound (DU) examination after thermal ablation of the GSV in asymptomatic patients. At present, limited data do not allow this recommendation to extend to MFA. Our aim is to identify characteristics and outcomes associated with ARTE following MFA vs radiofrequency ablation (RFA). METHODS: A retrospective review of a prospectively maintained database was conducted of patients who underwent MFA and RFA closure of incompetent above-knee GSVs. Patients treated for isolated tributary vein treatment or did not have a postprocedure DU examination within 48 to 72 hours were not included. Patients were classified into two groups: ARTE and no ARTE. Demographic data, Clinical, Etiologic, Anatomic and Pathophysiologic class, Venous Clinical Severity Score, operative details, postprocedure (48-72 hours) DU findings, and adverse events were analyzed. Variables that were significant on univariate analysis were evaluated using multivariate logistic regression with the primary outcome being development of ARTE. RESULTS: Between June 2018 and February 2023, 800 limbs were treated with either MFA (n = 224) or RFA (n = 576). Ninety-six GSVs treated with MFA met the study criteria. One hundred fifty successive GSVs treated with RFA during the same period were included as a comparison group. There was no statistically significant difference in baseline demographics between the two groups. Six patients (2.4%) demonstrated ARTE on postoperative DU examination at 48 to 72 hours (MFA, n = 5 [5.2%]; RFA, n = 1 [0.7%]; P = .02). Saphenous vein ablation with MFA (P = .045) and a vein diameter of >10 mm (P = .017) were associated with ARTE on both univariable and multivariable analysis. All patients who developed ARTE were treated with oral anticoagulants (mean, 15.6 days). Body mass index, Clinical, Etiologic, Anatomic and Pathophysiologic class, Venous Clinical Severity Score, microfoam volume, operative time, and prior deep venous thrombosis were not predictive of ARTE. CONCLUSIONS: ARTE after above-knee GSV closure occurred more frequently after MFA. Our results suggest that a saphenous vein diameter of >10 mm may be associated with ARTE. Despite this finding, all patients with ARTE were treated with short-term anticoagulation with no related complications. Until larger studies with high-risk subgroups have been studied after MFA, DU examination should be performed routinely after this procedure and patients with ARTE anticoagulated until the thrombus retracts caudal to the saphenofemoral junction or is no longer present on DU examination. Current societal guidelines recommending against routine post-thermal ablation DU examination should not be applied to similar patients after saphenous nonthermal MFA ablation.

Increased body mass index and vein diameter are associated with incomplete target vein closure following microfoam ablation of incompetent saphenous veins.

OBJECTIVE: Patient characteristics and risk factors for incomplete or non-closure following thermal saphenous vein ablation have been reported. However, similar findings have not been clearly described following commercially manufactured polidocanol microfoam ablation (MFA). The objective of our study is to identify predictive factors and outcomes associated with non-closure following MFA of symptomatic, refluxing saphenous veins. METHODS: A retrospective review of a prospectively maintained patient database was performed from procedures in our Ambulatory Procedure Unit. All consecutive patients who underwent MFA with commercially manufactured 1% polidocanol microfoam for symptomatic superficial vein reflux between June 2018 and September 2022 were identified. Patients treated for tributary veins only, without truncal vein ablation, were excluded. Patients were then stratified into groups: complete closure (Group I) and non-closure (Group II). Preoperative demographics, procedural details, and postoperative outcomes were analyzed. Preoperative variables that were significant on univariate analysis (prior deep venous thrombosis [DVT], body mass index [BMI] ≥30 kg/m2, and vein diameter) were entered into a multivariate logistic regression model with the primary outcome being vein non-closure. RESULTS: Between June 2018 and September 2022, a total of 224 limbs underwent MFA in our ambulatory venous center. Of these, 127 limbs in 103 patients met study inclusion criteria. Truncal veins treated included the above-knee great saphenous vein (Group I: n = 89, 77% vs Group II: n = 7, 58%; P = .14), below-knee great saphenous vein (Group I: n = 7, 6% vs Group II: n = 0; P = .38), anterior accessory saphenous vein (Group I: n = 17, 15% vs Group II: n = 4, 33%; P = .12, and small saphenous vein (Group I: n = 4, 4% vs Group II: n = 1, 8%; P = .41). Complete closure (Group I) occurred in 115 limbs, and 12 limbs did not close (Group II) based on postoperative duplex ultrasound screening. The mean BMI in Group II (36.1 ± 6.4 kg/m2) was significantly greater than Group I (28.6 ± 6.1 kg/m2) (P < .001). Vein diameter of ≥10.2 mm was independently associated with truncal vein non-closure with an odds ratio of 4.8. The overall mean foam volume was 6.2 ± 2.7 ml and not different between the two cohorts (Group I: 6.2 ± 2.6 ml vs Group II: 6.3 + 3.5 ml; P = .89). Post MFA improvement in symptoms was higher in Group I (96.9%) compared with Group II (66.7%) (P = .001). The mean postoperative Venous Clinical Severity Score was also lower in Group I (8.0 ± 3.0) compared with Group II (9.9 ± 4.2) (P = .048). The overall incidences of ablation-related thrombus extension and DVT were 4.7% (n = 6) and 1.6% (n = 2), and all occurred in Group I. All were asymptomatic and resolved with anticoagulation. CONCLUSIONS: Microfoam ablation of symptomatic, refluxing truncal veins results in excellent overall closure rates and symptomatic relief. BMI ≥30 kg/m2 and increased vein diameter are associated with an increased risk of saphenous vein non-closure following MFA. Non-closure is associated with less symptomatic improvement and a lower post-procedure reduction in Venous Clinical Severity Score. Despite the incidence of ablation-related thrombus extension and DVT in this study being higher than reported rates following thermal ablation, MFA is safe for patients with early postoperative duplex ultrasound surveillance and selective short-term anticoagulation.

Enhanced external counter pulsation therapy in patients with symptomatic and severe intracranial steno-occlusive disease: a randomized clinical trial protocol.

Intracranial stenosis is prevalent among Asians and constitutes a common cause of cerebral ischemia. While the best medical therapy carries stroke recurrence rates in excess of 10% per year, trials with intracranial stenting have been associated with unacceptable peri-procedural ischemic events. Cerebral ischemic events are strongly related to the severity of intracranial stenosis, which is high in patients with severe intracranial stenosis with poor vasodilatory reserve. Enhanced External Counter Pulsation (EECP) therapy is known to improve myocardial perfusion by facilitating the development of collateral blood vessels in the heart. In this randomized clinical trial, we evaluate whether EECP therapy may be useful in patients with severe stenosis of intracranial internal carotid (ICA) or middle cerebral artery (MCA). The review of literature, methods of evaluation, status of currently used therapeutic approaches, and trial protocol have been presented. Clinical trial registration: ClinicalTrials.gov, Identifier: NCT03921827.

Early results following comparison of radiofrequency and microfoam ablation of large diameter truncal veins demonstrate high closure rates and symptomatic relief.

OBJECTIVE: Endovenous closure of truncal veins with a large diameter (LD) (≥8 mm) has been associated with higher risk of post ablation thrombus propagation into the deep venous system. Similar findings after Varithena microfoam ablation (MFA) have not been characterized. The study objective was to analyze outcomes after both radiofrequency ablation (RFA) and MFA and of LD truncal veins. METHODS: A retrospective review of a prospectively maintained database was performed. All patients who underwent MFA and RFA for LD symptomatic truncal vein reflux (≥8 mm) were identified. All patients had postoperative duplex (48-72 hours) scanning. Patients underwent subsequent clinical follow-up at 3 to 6 weeks. Demographic data, CEAP Classification, Venous Clinical Severity Score, procedure details, adverse thrombotic events, and follow-up data were abstracted. RESULTS: Between June 2018 and September 2022, 784 consecutive limbs (RFA, n = 560; MFA, n = 224) underwent truncal vein (great, accessory, and small saphenous) closure for symptomatic reflux. Sixty-six limbs in the MFA group met inclusion criteria. Sixty-six consecutive limbs treated with RFA during the same time period were included as a comparison group. The mean truncal vein diameter treated was 10.5 mm (RFA, 10.0 mm; MFA, 10.9 mm). Twenty-nine limbs (44%) in the RFA group underwent concomitant phlebectomy. Tributary veins were concomitantly sclerosed in 34 MFA limbs (52%). Total procedural times were shorter in the MFA group (MFA, 31.6 minutes vs RFA, 55.7 minutes) (P < .001). Immediate closure rates were 100% and 95% in the RFA and MFA groups, respectively. Venous Clinical Severity Scores improved after treatment in both groups (RFA, from 9.5 to 7.8; P ≤ .001) (MFA, from 11.3 to 9.0; P ≤ .001). In the RFA and MFA groups, 83% and 79% of venous ulcers healed during the study period, respectively. Symptomatic superficial phlebitis occurred after RFA in 11% and 17% MFA. The incidence of postablation proximal deep venous thrombus extension was 3.0% in the RFA group and 6.1% in the MFA group, which was not statistically significant. All resolved with short-term oral anticoagulant therapy. No remote deep venous thromboses or pulmonary emboli occurred in either group. CONCLUSIONS: High early closure rates, symptom relief and ulcer healing rates can be achieved after RFA and MFA of LD saphenous veins. Both techniques can be used safely across a wide array of CEAP classes. Longer term studies are required to characterize the durability of MFA closure and sustained symptom relief in LD truncal veins.

Comparison of outcomes following polidocanol microfoam and radiofrequency ablation of incompetent thigh great and accessory saphenous veins.

BACKGROUND: Microfoam ablation (MFA) and radiofrequency ablation (RFA) are both approved by the Food and Drug Administration for treatment of proximal saphenous truncal veins. The objective of our study was to compare early postoperative outcomes between MFA and RFA following treatment of incompetent thigh saphenous veins. METHODS: A retrospective review of a prospectively maintained database was conducted of patients who underwent treatment of incompetent great saphenous veins (GSVs) or anterior accessory saphenous veins (AASVs) in the thigh. All the patients underwent duplex ultrasound of the treated leg at 48 to 72 hours postoperatively. Patients were excluded from analysis if concomitant stab phlebectomy was performed. Demographic data, CEAP (clinical, etiologic, anatomic, pathophysiologic) class, venous clinical severity score (VCSS), and adverse events were recorded. RESULTS: Between June 2018 and September 2022, 784 consecutive limbs (RFA, n = 560; MFA, n = 224) underwent venous closure for symptomatic reflux. A total of 200 consecutive thigh GSVs and ASVs treated within the study period using either MFA (n = 100) or RFA (n = 100) were identified. The patients were predominantly women (69%) with a mean age of 64 years. The preoperative CEAP classification was similar between the MFA and RFA groups. The mean preoperative VCSS was 9.4 ± 2.6 for the RFA patients and 9.9 ± 3.3 for the MFA patients. Among the RFA patients, the GSV was treated in 98% and the AASV in 2% compared with the GSV in 83% and the AASV in 17% in the MFA group (P < .001). The mean operative time was 42.4 ± 15.4 minutes in the RFA group and 33.8 ±16.9 minutes in the MFA group (P < .001). The median follow-up was 64 days for the study cohort. The mean postoperative VCSS declined to 7.3 ± 2.1 in the RFA group and 7.8 ± 2.9 in the MFA group. Complete closure occurred in 100% of the limbs after RFA and 90% after MFA (P = .005). Eight veins were partially closed and two remained patent following MFA. The incidence of superficial phlebitis was 6% and 15% (P = .06) after RFA and MFA, respectively. Overall, symptomatic relief was 90% following RFA and 89.5% following MFA. The complete ulcer healing rate for the entire cohort was 77.8%. Deep venous proximal thrombus extension (RFA, 1%; vs MFA, 4%; P = .37) and remote deep vein thrombosis (RFA, 0%; vs MFA, 2%; P = .5) showed a trend toward being higher following MFA but the difference did not reach statistical significance. All were asymptomatic and resolved with short-term anticoagulation therapy. CONCLUSIONS: MFA and RFA are both safe and effective for treating incompetent thigh saphenous veins, with excellent symptomatic relief and a low incidence of postprocedure adverse thrombotic events. RFA resulted in improved complete closure rates following initial treatment compared with MFA. The operative times were shorter with MFA. Both modalities can be used for patients with active venous ulcers with good healing rates. Longer term studies are required to characterize the durability of MFA closure for above knee truncal veins.

Using gene panels in the diagnosis of neuromuscular disorders: A mini-review.

The diagnosis of inherited neuromuscular disorders is challenging due to their genetic and phenotypic variability. Traditionally, neurophysiology and histopathology were primarily used in the initial diagnostic approach to these conditions. Sanger sequencing for molecular diagnosis was less frequently utilized as its application was a time-consuming and cost-intensive process. The advent and accessibility of next-generation sequencing (NGS) has revolutionized the evaluation process of genetically heterogenous neuromuscular disorders. Current NGS diagnostic testing approaches include gene panels, whole exome sequencing (WES), and whole genome sequencing (WGS). Gene panels are often the most widely used, being more accessible due to availability and affordability. In this mini-review, we describe the benefits and risks of clinical genetic testing. We also discuss the utility, benefits, challenges, and limitations of using gene panels in the evaluation of neuromuscular disorders.

Autoimmune glial fibrillary acidic protein astrocytopathy masquerading as tuberculosis of the central nervous system: a case series.

We describe the case history of three patients with meningoencephalitis who were initially treated for presumed tuberculous meningoencephalitis before being diagnosed with autoimmune glial fibrillary acidic protein (GFAP) astrocytopathy. We highlight the overlapping clinical features between autoimmune GFAP astrocytopathy and tuberculous meningoencephalitis and the challenges in early diagnosis, as both entities respond to an initial course of steroids accompanying antituberculous medications. Early evaluation of GFAP-immunoglobulin G in the cerebrospinal fluid of patients who present with aseptic meningoencephalitis could reveal autoimmune GFAP astrocytopathy, which responds favorably to immunotherapy.

Factors influencing protective behaviours during haze episodes in Singapore: A population-based study.

INTRODUCTION: Haze is a recurrent problem in Southeast Asia. Exposure to haze is linked to ophthalmic, respiratory and cardiovascular diseases, and mortality. In this study, we investigated the role of demographic factors, knowledge and perceived risk in influencing protective behaviours during the 2013 haze in Singapore. METHODS: We evaluated 696 adults in a cross-sectional study. Participants were sampled via a 2-stage simple random sampling without replacement from a large residential district in Singapore in 2015. The questionnaire measured the participant's knowledge, perceived risk and behaviours during the Southeast Asian haze crisis in 2013. Reliability and validity of the questionnaire were assessed using comparative fit index (≥0.96) and root mean square error of approximation (≤0.05). We performed structural equation modelling to examine the relationship between the hypothesised factors and protective behaviours. RESULTS: More than 95% of the individuals engaged in at least 1 form of protective behaviour. Knowledge was strongly associated with protective behaviours via direct effect (ß=0.45, 95% CI 0.19-0.69, P<0.001) and indirect effect through perceived risk (ß=0.18, 95% CI 0.07-0.31, P=0.002). Perceived risk was associated with protective behaviours (ß=0.28, 95% CI:0.11-0.44, P=0.002). A lower household income and ethnic minority were associated with protective behaviours. A lower education level and smokers were associated with lower knowledge of haze. A higher education and ethnic minority were associated with a lower perceived risk. Wearing of N95 masks was associated with other haze-related protective behaviours (ß=0.24, 95% CI 0.08-0.37, P=0.001). CONCLUSION: Knowledge was associated with protective behaviours, suggesting the importance of public education. Efforts should target those of lower education level and smokers. The wearing of N95 masks correlates with uptake of other protective behaviours.

Powered Air Purifying Respirator (PAPR) restores the N95 face mask induced cerebral hemodynamic alterations among Healthcare Workers during COVID-19 Outbreak.

BACKGROUND AND AIM: COVID-19 pandemic has resulted in an unprecedented increased usage of Personal protective equipment (PPE) by healthcare-workers. PPE usage causes headache in majority of users. We evaluated changes in cerebral hemodynamics among healthcare-workers using PPE. METHODS: Frontline healthcare-workers donning PPE at our tertiary center were included. Demographics, co-morbidities and blood-pressure were recorded. Transcranial Doppler (TCD) monitoring of middle cerebral artery was performed with 2-MHz probe. Mean flow velocity (MFV) and pulsatility index (PI) were recorded at baseline, after donning N95 respirator-mask, and after donning powered air-purifying respirator (PAPR), when indicated. End-tidal carbon-dioxide (ET-CO2) pressure was recorded for participants donning PAPR in addition to the N95 respirator-mask. RESULTS: A total of 154 healthcare-workers (mean age 29 ± 12 years, 67% women) were included. Migraine was the commonest co-morbidity in 38 (25%) individuals while 123 (80%) developed de-novo headache due to N95 mask. Donning of N95 respirator-mask resulted in significant increase in MFV (4.4 ± 10.4 cm/s, p < 0.001) and decrease in PI (0.13 ± 0.12; p < 0.001) while ET-CO2 increased by 3.1 ± 1.2 mmHg (p < 0.001). TCD monitoring in 24 (16%) participants donning PAPR and N95 respirator mask together showed normalization of PI, accompanied by normalization of ET-CO2 values within 5-min. Combined use of N95 respirator-mask and PAPR was more comfortable as compared to N95 respirator-mask alone. CONCLUSION: Use of N95 respirator-mask results in significant alterations in cerebral hemodynamics. However, these effects are mitigated by the use of additional PAPR. We recommend the use of PAPR together with the N95 mask for healthcare-workers doing longer duties in the hospital wards.

Impact of the COVID-19 pandemic on hyperacute stroke treatment: experience from a comprehensive stroke centre in Singapore.

The Coronavirus disease 2019 (COVID-19) pandemic is rapidly evolving and affecting healthcare systems across the world. Singapore has escalated its alert level to Disease Outbreak Response System Condition (DORSCON) Orange, signifying severe disease with community spread. We aimed to study the overall volume of AIS cases and the delivery of hyperacute stroke services during DORSCON Orange. This was a single-centre, observational cohort study performed at a comprehensive stroke centre responsible for AIS cases in the western region of Singapore, as well as providing care for COVID-19 patients. All AIS patients reviewed as an acute stroke activation in the Emergency Department (ED) from November 2019 to April 2020 were included. System processes timings, treatment and clinical outcome variables were collected. We studied 350 AIS activation patients admitted through the ED, 206 (58.9%) pre- and 144 during DORSCON Orange. Across the study period, number of stroke activations showed significant decline (p = 0.004, 95% CI 6.513 to - 2.287), as the number of COVID-19 cases increased exponentially, whilst proportion of activations receiving acute recanalization therapy remained stable (p = 0.519, 95% CI - 1.605 to 2.702). Amongst AIS patients that received acute recanalization therapy, early neurological outcomes in terms of change in median NIHSS at 24 h (-4 versus -4, p = 0.685) were largely similar between the pre- and during DORSCON orange periods. The number of stroke activations decreased while the proportion receiving acute recanalization therapy remained stable in the current COVID-19 pandemic in Singapore.

The evolution of open abdominal aortic aneurysm repair at a tertiary care center.

BACKGROUND: The characteristics of and indications for open abdominal aortic aneurysm (AAA) repair have evolved over time. We evaluated these trends through the experience at a tertiary care academic center. METHODS: A retrospective review was conducted for patients undergoing open AAA repair (inclusive of type IV thoracoabdominal aortic aneurysms) from 2005 to 2018 at an academic institution. Trends over time were evaluated using the Spearman test; Cox regression was used to determine predictors of mortality and to generate adjusted survival curves. RESULTS: There were 628 patients (71.5% male; 88.2% white) with a mean age of 70.5 ± 9.4 years who underwent open AAA repair with a mean aneurysm diameter of 6.2 ± 1.5 cm. The median length of stay was 10 days, and the median intensive care unit length of stay was 3 days. Urgent repair was undertaken in 21.1%; 22.3% were type IV thoracoabdominal aortic aneurysm repairs, and 9.9% were performed for explantation. Our series favored a retroperitoneal approach in the majority of cases (82.5%). The proximal clamp sites were supraceliac (46.1%), suprarenal (29.1%), and infrarenal (24.8%), with approximately a third requiring renal artery reimplantation. The average cross-clamp time was 25.5 ± 14.9 minutes; the mean renal ischemia time for supraceliac and suprarenal clamp sites was 28.4 ± 12.3 minutes and 23.5 ± 12.7 minutes, respectively. Postoperative renal dysfunction occurred in 19.6% of the overall cohort, with 6.2% requiring hemodialysis. Of those requiring postoperative hemodialysis, the majority (75%) received an urgent repair. The in-hospital mortality was 2.3% for elective cases vs 20.9% for urgent repair, and 29.8% of patients were discharged to rehabilitation, with an overall 30-day readmission rate of 7.9%. Over time, there were trends of increased aneurysm repair complexity, with decreasing infrarenal clamp sites, increasing supraceliac clamp sites, increasing proportion of explantations, and increasing need for bifurcated grafts. The acuity of aneurysm repair likewise changed, with the proportion of urgent repairs increasing over time, largely attributable to the rise in explantations. Clamp site influenced the frequency of perioperative complications. Urgent repairs and age at operation were associated with mortality, whereas mortality was not associated with need for explantation and clamp location. CONCLUSIONS: Aneurysm repair reflected increasing complexity over time, with the need for explantation among urgent repairs significantly on the rise. Urgency and clamp location independently predicted long-term mortality, even after adjustment for age. These findings underscore the changing landscape of open AAA repair in the current era.

Heart-Kidney and Heart-Liver Transplantation Provide Immunoprotection to the Cardiac Allograft.

BACKGROUND: Prior single-center studies suggest that kidney and liver allografts are immunoprotective toward transplanted hearts. The broader effects of the simultaneous transplantation of kidney or liver on protection from rejection are unclear. METHODS: The United Network for Organ Sharing database for heart transplantation was queried from 1987 to 2015 and stratified into patients undergoing heart-liver transplantation (HLT) (n = 192), heart-kidney transplantation (HKT) (n = 1,174), and heart-only transplantation (HT) (n = 61,471). Perioperative and follow-up data were compared between HT versus HLT and HT versus HKT groups using analysis of variance (continuous), chi-square test (categorical), and Kaplan-Meier curves (survival). RESULTS: HKT patients were older (51.2 ± 13.4 years of age) compared with HT patients (45.6 ± 19.2 years of age; p < 0.0001), with higher rate of diabetes (33.8% versus 14.8%; p < 0.0001) and dialysis (49.7% versus 2.1%; p < 0.0001). HKT (46.2%) and HLT (49.5%) patients had more urgent need for transplantation (status 1A) compared with HT patients (32%; p < 0.0001). Acute rejection episodes before discharge were lower in the HLT group (7.1% versus 3.1%; p = 0.03). Ten-year patient survivals were similar for HT (53.6%) versus HKT (56.7%) (p = 0.13) versus HLT (60.4%) (p = 0.09). Treatment for rejection during the first posttransplant year was lower in HLT (2.1%) and HKT (8.4%) compared with HT (17.4%) (p < 0.0001 for both). Cox multivariate analysis showed that cardiac allograft survival was improved in HKT (odds ratio, 0.58; 95% confidence interval [CI], 0.49 to 0.70; p < 0.0001). Additionally, HKT (hazard ratio, 0.52; 95% CI, 0.45 to 0.60; p < 0.0001) and HLT (hazard ratio, 0.24; 95% CI, 0.15 to 0.39; p < 0.0001) were associated with improved freedom from rejection. CONCLUSIONS: Nationally, HKT and HLT have equivalent postoperative outcomes as HT. Simultaneous kidney or liver transplantation confers an improved clinical and immunologic outcome.

Intrinsic cellular chirality regulates left-right symmetry breaking during cardiac looping.

The vertebrate body plan is overall symmetrical but left-right (LR) asymmetric in the shape and positioning of internal organs. Although several theories have been proposed, the biophysical mechanisms underlying LR asymmetry are still unclear, especially the role of cell chirality, the LR asymmetry at the cellular level, on organ asymmetry. Here with developing chicken embryos, we examine whether intrinsic cell chirality or handedness regulates cardiac C looping. Using a recently established biomaterial-based 3D culture platform, we demonstrate that chick cardiac cells before and during C looping are intrinsically chiral and exhibit dominant clockwise rotation in vitro. We further show that cells in the developing myocardium are chiral as evident by a rightward bias of cell alignment and a rightward polarization of the Golgi complex, correlating with the direction of cardiac tube rotation. In addition, there is an LR polarized distribution of N-cadherin and myosin II in the myocardium before the onset of cardiac looping. More interestingly, the reversal of cell chirality via activation of the protein kinase C signaling pathway reverses the directionality of cardiac looping, accompanied by a reversal in cellular biases on the cardiac tube. Our results suggest that myocardial cell chirality regulates cellular LR symmetry breaking in the heart tube and the resultant directionality of cardiac looping. Our study provides evidence of an intrinsic cellular chiral bias leading to LR symmetry breaking during directional tissue rotation in vertebrate development.

Epithelial Cell Chirality Revealed by Three-Dimensional Spontaneous Rotation.

Our understanding of the left-right (LR) asymmetry of embryonic development, in particular the contribution of intrinsic handedness of the cell or cell chirality, is limited due to the confounding systematic and environmental factors during morphogenesis and a ack of physiologically relevant in vitro 3D platforms. Here we report an efficient two-layered biomaterial platform for determining the chirality of individual cells, cell aggregates, and self-organized hollow epithelial spheroids. This bioengineered niche provides a uniform defined axis allowing for cells to rotate spontaneously with a directional bias toward either clockwise or counterclockwise directions. Mechanistic studies reveal an actin-dependent, cell-intrinsic property of 3D chirality that can be mediated by actin cross-linking via α-actinin-1. Our findings suggest that the gradient of extracellular matrix is an important biophysicochemical cue influencing cell polarity and chirality. Engineered biomaterial systems can serve as an effective platform for studying developmental asymmetry and screening for environmental factors causing birth defects.

Lineage-Specific Chiral Biases of Human Embryonic Stem Cells during Differentiation.

Left-right symmetry breaking is a complex developmental process and an important part of embryonic axis development. As of yet, the biophysical mechanism behind LR asymmetry establishment remains elusive for the overall asymmetry of embryos as well as for the organ-specific asymmetry. Here, we demonstrate that inherent cellular chirality is observable in the cells of early embryonic stages using a 3D Matrigel bilayer system. Differentiation of human embryonic stem cells to three lineages corresponding to heart, intestine, and neural tissues demonstrates phenotype-specific inherent chiral biases, complementing the current knowledge regarding organ development. The existence of inherent cellular chirality early in development and its correlation with organ asymmetry implicate cell chirality as a possible regulator in LR symmetry breaking.

Cell chirality: emergence of asymmetry from cell culture.

Increasing evidence suggests that intrinsic cell chirality significantly contributes to the left-right (LR) asymmetry in embryonic development, which is a well-conserved characteristic of living organisms. With animal embryos, several theories have been established, but there are still controversies regarding mechanisms associated with embryonic LR symmetry breaking and the formation of asymmetric internal organs. Recently, in vitro systems have been developed to determine cell chirality and to recapitulate multicellular chiral morphogenesis on a chip. These studies demonstrate that chirality is indeed a universal property of the cell that can be observed with well-controlled experiments such as micropatterning. In this paper, we discuss the possible benefits of these in vitro systems to research in LR asymmetry, categorize available platforms for single-cell chirality and multicellular chiral morphogenesis, and review mathematical models used for in vitro cell chirality and its applications in in vivo embryonic development. These recent developments enable the interrogation of the intracellular machinery in LR axis establishment and accelerate research in birth defects in laterality.This article is part of the themed issue 'Provocative questions in left-right asymmetry'.