Atacama Clear for Complex 3D Imaging of Organs.

3D reconstructive imaging is a powerful strategy to interrogate the global architecture of tissues. We developed Atacama Clear (ATC), a novel method that increases 3D imaging signal-to-noise ratios (SNRs) while simultaneously increasing the capacity of tissue to be cleared. ATC potentiated the clearing capacity of all tested chemical reagents currently used for optical clearing by an average of 68%, and more than doubled SNRs. This increased imaging efficacy enabled multiplex interrogation of tough fibrous tissue and specimens that naturally exhibit high levels of background noise, including the heart, kidney, and human biopsies. Indeed, ATC facilitated visualization of previously undocumented adjacent nephron segments that exhibit notoriously high autofluorescence, elements of the cardiac conduction system, and the distinct human glomerular tissue layers, at single cell resolution. Moreover, ATC was validated to be compatible with fluorescent reporter proteins in murine, zebrafish, and 3D stem cell model systems. These data establish ATC for 3D imaging studies of challenging tissue types.

A pancreatic cancer organoid platform identifies an inhibitor specific to mutant KRAS.

KRAS mutations, mainly G12D and G12V, are found in more than 90% of pancreatic ductal adenocarcinoma (PDAC) cases. The success of drugs targeting KRASG12C suggests the potential for drugs specifically targeting these alternative PDAC-associated KRAS mutations. Here, we report a high-throughput drug-screening platform using a series of isogenic murine pancreatic organoids that are wild type (WT) or contain common PDAC driver mutations, representing both classical and basal PDAC phenotypes. We screened over 6,000 compounds and identified perhexiline maleate, which can inhibit the growth and induce cell death of pancreatic organoids carrying the KrasG12D mutation both in vitro and in vivo and primary human PDAC organoids. scRNA-seq analysis suggests that the cholesterol synthesis pathway is upregulated specifically in the KRAS mutant organoids, including the key cholesterol synthesis regulator SREBP2. Perhexiline maleate decreases SREBP2 expression levels and reverses the KRAS mutant-induced upregulation of the cholesterol synthesis pathway.

Checkpoint kinase 2 controls insulin secretion and glucose homeostasis.

After the discovery of insulin, a century ago, extensive work has been done to unravel the molecular network regulating insulin secretion. Here we performed a chemical screen and identified AZD7762, a compound that potentiates glucose-stimulated insulin secretion (GSIS) of a human ß cell line, healthy and type 2 diabetic (T2D) human islets and primary cynomolgus macaque islets. In vivo studies in diabetic mouse models and cynomolgus macaques demonstrated that AZD7762 enhances GSIS and improves glucose tolerance. Furthermore, genetic manipulation confirmed that ablation of CHEK2 in human ß cells results in increased insulin secretion. Consistently, high-fat-diet-fed Chk2-/- mice show elevated insulin secretion and improved glucose clearance. Finally, untargeted metabolic profiling demonstrated the key role of the CHEK2-PP2A-PLK1-G6PD-PPP pathway in insulin secretion. This study successfully identifies a previously unknown insulin secretion regulating pathway that is conserved across rodents, cynomolgus macaques and human ß cells in both healthy and T2D conditions.

Comparative Analysis of 2D and 3D Metrics for Evaluation of Postoperative Outcomes Following Endoscopic Suturectomy for Sagittal Craniosynostosis.

To assess whether 3-dimensional (3D) volumetrics can be used to track and evaluate postoperative course of patients treated with endoscopic suturectomy for nonsyndromic sagittal synostosis, we compared changes in 2-dimensional (2D) measurements along with 3D volumetric correlates throughout the period of helmet therapy. Forty-six patients treated at our institution with endoscopic suturectomy for sagittal synostosis were retrospectively reviewed. Head circumference (HC), cephalic index (CI), and total cranial volumes (TCVs) were measured at 3 timepoints following surgery using optical surface scans obtained for helmet orthotics. All measurements showed significant differences between timepoints on the analysis of variance ( P <0.001). There was a significant correlation between CI and TCV (r=0.35, P =0.004) and between HC and TCV (r=0.81, P <0.001). The normalized rate of change over the course of treatment was significantly higher for TCV (36.7%) than for CI (8.8%) and HC (8.4%, P <0.001), with no difference between HC and CI. The authors conclude that 3D metrics were able to reliably follow the course of postoperative 2D metrics. There was a direct and linear correlation between HC and CI with TCV. Total cranial volumes showed the highest rate of sustained change at every timepoint. Although CI and HC plateau after the first measurement, TCV continues to adapt over the course of treatment. These results demonstrate the feasibility and value of volumetrics from 3D imaging to provide a more comprehensive evaluation of postoperative surgical outcomes than traditional 2D metrics without the ionizing radiation traditionally utilized for CT to obtain 3D metrics.

Polysomnography-guided mandibular distraction osteogenesis in Pierre Robin sequence patients.

STUDY OBJECTIVES: Craniofacial malformations with micrognathia cause high grades of obstructive sleep apnea (OSA) measured by polysomnography (PSG). Mandibular distraction osteogenesis is a novel procedure for upper airway obstruction relief. Our primary objective was to describe the utilization of PSGs to improve obstruction in patients undergoing mandibular distraction. METHODS: This is a retrospective study. Patients with micrognathia and severe upper airway obstruction, presenting with severe OSA diagnosed by PSG, were included from a single tertiary care center between 2015 and 2019. PSGs were done (1) prior to surgery, (2) once the cosmetic goal was achieved (Post-Op 1), and (3) if residual moderate-to-severe OSA was seen, every 2 nights until mild or no OSA was achieved (Post-Op 2). RESULTS: Thirteen patients were included. The median age at surgery was 1.1 months (10 days-3 months). All 13 patients had baseline severe OSA, with a median obstructive apnea-hypopnea index of 33 events/h and a median O2 nadir of 73%. Post-Op 1 PSG was done at a median of 6 days after surgery. Median first postoperative obstructive apnea-hypopnea index in all 13 patients was 6.8 events/h, with a median O2 nadir of 87%. A median additional distraction of 3 mm was needed beyond the traditionally recommended advancement. Long-term follow-up studies at or after 1 year were done in 5 patients, all showing persistent nonsevere OSA. CONCLUSIONS: This is the first case series utilizing PSGs as a guide for mandibular distraction osteogenesis in patients with micrognathia showing the need for jaw overcorrection to achieve resolution of OSA. CITATION: Kochhar R, Modi V, de Silva N, et al. Polysomnography-guided mandibular distraction osteogenesis in Pierre Robin sequence patients. J Clin Sleep Med. 2022;18(7):1749-1755.

Gram-Negative Ventriculostomy-Associated Infections Predict Shunt Dependency in Stroke Diagnoses and Other Brain Injuries.

BACKGROUND: Several predictors have been studied for shunt dependency after stroke and other brain injuries. However, little is known about the association between ventriculostomy-associated infections (VAIs) and impaired cerebrospinal fluid (CSF) outflow. Moreover, gram-negative (GN) VAIs induce a potent neuroinflammatory process and are clinically challenging to treat. OBJECTIVE: To assess if GN-VAIs predict ventriculoperitoneal shunt (VPS) dependency. METHODS: Retrospective analysis of postprocedure infection rates was performed in 586 patients with external ventricle drainage (EVD) placed on site between 2012 and 2018. We collected sex, age, stroke and nonstroke related, location of EVD placement, type of hospital, EVD duration, and EVD exchange. RESULTS: Among 586 patients requiring an EVD, 55 developed a VAI. Most were caused by gram-positive (GP) pathogens (61.8%). A total of 120 patients required a conversion from EVD to VPS. Patients with VAIs had higher rates of VPS placement (49.09% vs 17.65%, P < .001), whereas patients with GN-VAIs had significantly higher rates of EVD conversion to VPS (77.78% vs 35.29%, P = .012) compared with GP-VAIs. The multivariate analysis showed that GN-VAIs were an independent predictor for shunt dependency (odds ratio = 12.896; 95% CI 3.407-48.82, P < .001). In receiver operating characteristics analysis, those less than 44.5 yr of age and more than 12 d of EVD duration were identified as the best cutoff values to discriminate the development of GN-VAI. CONCLUSION: Patients who experience a GN VAI are in greater risk of impaired CSF outflow, thus requiring VPS placement.

Impact of hospital-acquired complications in long-term clinical outcomes after subarachnoid hemorrhage.

OBJECTIVE: Patients with subarachnoid hemorrhage (SAH) usually have prolonged hospitalizations due to the need to closely monitor their neurological status. Therefore, these patients have higher risk of experiencing hospital-acquired complications (HACs), which can complicate their clinical course and recovery. However, there is no evidence on the impact of HACs of long-term clinical outcomes. We aimed to identify if HACs are independent risk factors for poor clinical outcomes at 12-18 months of follow-up. PATIENTS AND METHODS: Retrospective analysis of 323 patients with SAH diagnosis from 2013 until June 2018. We collected patient-related factors (age, sex, body mass index (BMI), ethnicity), comorbidities (hypertension, smoke status, diabetes, coronary heart diseases, prothrombotic diseases and hypercholesterolemia), clinical variables (Hunt-Hess grade, modified Fisher grade, treatment, delayed cerebral ischemia), aneurysm characteristics (location, size) and HACs (pneumonia, deep vein thrombosis (DVT), urinary tract infection (UTI), external ventricular drainage (EVD) infections, sepsis, hyponatremia and acute respiratory distress syndrome). Poor outcomes were defined as mRS ≥ 3. RESULTS: 204 patients were included in the primary analysis. 82 (40.2%) experienced one or more HACs during their hospital course. Patients that developed HACs have significantly increased ICU (12.1 ±â€¯6.6 vs 24.3 ±â€¯23.6, p < .001) and hospital (18.7 ±â€¯14.2 vs 35.3 ±â€¯26.3, p < .001) length of stays. Moreover, patients with HACs had significant higher rates of delayed cerebral ischemia, non-routine discharge and poor outcomes at 90 days. 177 patients had complete follow-ups at 12-18 months, HACs were independent risk factors for poor functional outcomes at 12-18 months after adjusting for demographic, comorbidities and clinical variables [OR = 3.205, 95% CI 1.231-8.347, p < 0.001]. CONCLUSIONS: HACs are an independent risk factor of sustaining poor clinical outcomes 12-18 months after a SAH. Furthermore, HACs are significantly related with the occurrence of DCI, with non-routine discharge and 90-day poor functional outcomes.