General anesthetic action profile on the human prefrontal cortex cells through comprehensive single-cell RNA-seq analysis.

The cellular and molecular actions of general anesthetics to induce anesthesia state and also cellular signaling changes for subsequent potential "long term" effects remain largely elusive. General anesthetics were reported to act on voltage-gated ion channels and ligand-gated ion channels. Here we used single-cell RNA-sequencing complemented with whole-cell patch clamp and calcium transient techniques to examine the gene transcriptome and ion channels profiling of sevoflurane and propofol, both commonly used clinically, on the human fetal prefrontal cortex (PFC) mixed cell cultures. Both propofol and sevoflurane at clinically relevant dose/concentration promoted "microgliosis" but only sevoflurane decreased microglia transcriptional similarity. Propofol and sevoflurane each extensively but transiently (<2 h) altered transcriptome profiling across microglia, excitatory neurons, interneurons, astrocytes and oligodendrocyte progenitor cells. Utilizing scRNA-seq as a robust and high-through put tool, our work may provide a comprehensive blueprint for future mechanistic studies of general anesthetics in clinically relevant settings.

A Network Meta-Analysis of Retreatment Rates following Bevacizumab, Ranibizumab, Aflibercept, and Laser for Retinopathy of Prematurity.

TOPIC: To compare bevacizumab, ranibizumab, aflibercept, and laser treatment as primary therapies for retinopathy of prematurity (ROP) in terms of retreatment rate. CLINICAL RELEVANCE: Anti-VEGF agents are increasingly used as primary treatment for ROP and may provide superior outcomes compared with laser in posterior disease. Head-to-head comparisons between different anti-VEGFs are lacking. METHODS: We searched CENTRAL, Embase, MEDLINE, and CINAHL databases for randomized controlled trials and nonrandomized comparative studies that had been reported as of March 2022. We included studies that used bevacizumab, ranibizumab, aflibercept or laser for ROP with comparable cohorts and treatment criteria. Studies were evaluated by the Grading of Recommendations, Assessment, Development and Evaluation framework, and those with biased case selection, nonrandomized case-control, or lack of control group were excluded. Frequentist meta-analyses of proportions determined the absolute primary retreatment rate of each modality and Bayesian network meta-analyses compared pairs of treatments in type 1 and Zone I ROP. RESULTS: In all, 30 studies (4686 eyes) were included in the network meta-analyses. For type 1 ROP, single-treatment success rates (i.e., likelihood of needing no further treatment) were 89.3% (95% confidence interval [CI]: 83.8%-93.8%; n = 1552) for laser, 87.0% (95% CI: 78.6%-93.8%; n = 2081) for bevacizumab, 80.7% (95% CI: 62.0%-94.4%; n = 326) for aflibercept, and 74.0% (95% CI: 62.7%-84.1%; n = 727) for ranibizumab. Bayesian network meta-analysis indicates that laser treatment is associated with a significant 62% (95% credible interval [CrI]: 16%-83%) reduction in retreatment risk compared with ranibizumab, while no significant difference was found among other pairwise comparisons. The mean ± standard error of the mean times to secondary treatment following primary aflibercept (12.96 ± 0.47 weeks) and bevacizumab (11.36 ± 0.54 weeks) therapy were significantly longer than that for primary ranibizumab (9.29 ± 0.43weeks) therapy (P = 7 × 10-7 and P = 9 × 10-3, respectively). For Zone I ROP, single-treatment success rates were 91.2% (95% CI: 83.6-96.9; n = 231) for bevacizumab, 78.3% (95% CI: 61.4-91.9; n = 100) for ranibizumab, and 65.9% (95% CI: 41.4-87.2; n = 158) for laser treatment. In this case, Bayesian network meta-analysis suggests that primary bevacizumab is associated with a significant 67% (95% CrI:10%-90%) reduction in retreatment risk compared with laser treatment. CONCLUSIONS: Laser was associated with a lower rate of retreatment than ranibizumab in type 1 ROP (Zones I and II combined), while bevacizumab was associated with a lower rate of retreatment than laser in Zone I ROP. Aflibercept and bevacizumab demonstrate longer duration of action than ranibizumab for ROP.

Ciliary IFT88 Protects Coordinated Adolescent Growth Plate Ossification From Disruptive Physiological Mechanical Forces.

Compared with our understanding of endochondral ossification, much less is known about the coordinated arrest of growth defined by the narrowing and fusion of the cartilaginous growth plate. Throughout the musculoskeletal system, appropriate cell and tissue responses to mechanical force delineate morphogenesis and ensure lifelong health. It remains unclear how mechanical cues are integrated into many biological programs, including those coordinating the ossification of the adolescent growth plate at the cessation of growth. Primary cilia are microtubule-based organelles tuning a range of cell activities, including signaling cascades activated or modulated by extracellular biophysical cues. Cilia have been proposed to directly facilitate cell mechanotransduction. To explore the influence of primary cilia in the mouse adolescent limb, we conditionally targeted the ciliary gene Intraflagellar transport protein 88 (Ift88fl/fl ) in the juvenile and adolescent skeleton using a cartilage-specific, inducible Cre (AggrecanCreERT2 Ift88fl/fl ). Deletion of IFT88 in cartilage, which reduced ciliation in the growth plate, disrupted chondrocyte differentiation, cartilage resorption, and mineralization. These effects were largely restricted to peripheral tibial regions beneath the load-bearing compartments of the knee. These regions were typified by an enlarged population of hypertrophic chondrocytes. Although normal patterns of hedgehog signaling were maintained, targeting IFT88 inhibited hypertrophic chondrocyte VEGF expression and downstream vascular recruitment, osteoclastic activity, and the replacement of cartilage with bone. In control mice, increases to physiological loading also impair ossification in the peripheral growth plate, mimicking the effects of IFT88 deletion. Limb immobilization inhibited changes to VEGF expression and epiphyseal morphology in Ift88cKO mice, indicating the effects of depletion of IFT88 in the adolescent growth plate are mechano-dependent. We propose that during this pivotal phase in adolescent skeletal maturation, ciliary IFT88 protects uniform, coordinated ossification of the growth plate from an otherwise disruptive heterogeneity of physiological mechanical forces. © 2022 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

Role of Ciliary Protein Intraflagellar Transport Protein 88 in the Regulation of Cartilage Thickness and Osteoarthritis Development in Mice.

OBJECTIVE: Mechanical and biologic cues drive cellular signaling in cartilage development, health, and disease. Primary cilia proteins, which are implicated in the transduction of biologic and physiochemical signals, control cartilage formation during skeletal development. This study was undertaken to assess the influence of the ciliary protein intraflagellar transport protein 88 (IFT88) on postnatal cartilage from mice with conditional knockout of the Ift88 gene (Ift88-KO). METHODS: Ift88fl/fl and aggrecanCreERT2 mice were crossed to create a strain of cartilage-specific Ift88-KO mice (aggrecanCreERT2 ;Ift88fl/fl ). In these Ift88-KO mice and Ift88fl/fl control mice, tibial articular cartilage thickness was assessed by histomorphometry, and the integrity of the cartilage was assessed using Osteoarthritis Research Society International (OARSI) damage scores, from adolescence through adulthood. In situ mechanisms of cartilage damage were investigated in the microdissected cartilage sections using immunohistochemistry, RNAScope analysis, and quantitative polymerase chain reaction. Osteoarthritis (OA) was induced in aggrecanCreERT2 ;Ift88fl/fl mice and Ift88fl/fl control mice using surgical destabilization of the medial meniscus (DMM). Following tamoxifen injection and DMM surgery, the mice were given free access to exercise on a wheel. RESULTS: Deletion of Ift88 resulted in progressive reduction in the thickness of the medial tibial cartilage in adolescent mice, as well as marked atrophy of the cartilage in mice during adulthood. In aggrecanCreERT2 ;Ift88fl/fl mice at age 34 weeks, the median thickness of the medial tibial cartilage was 89.42 µm (95% confidence interval [95% CI] 84.00-93.49), whereas in Ift88fl/fl controls at the same age, the median cartilage thickness was 104.00 µm (95% CI 100.30-110.50; P < 0.0001). At all time points, the median thickness of the calcified cartilage was reduced. In some mice, atrophy of the medial tibial cartilage was associated with complete, spontaneous degradation of the cartilage. Following DMM, aggrecanCreERT2 ;Ift88fl/fl mice were found to have increased OARSI scores of cartilage damage. In articular cartilage from maturing mice, atrophy was not associated with obvious increases in aggrecanase-mediated destruction or chondrocyte hypertrophy. Of the 44 candidate genes analyzed, only Tcf7l2 expression levels correlated with Ift88 expression levels in the microdissected cartilage. However, RNAScope analysis revealed that increased hedgehog (Hh) signaling (as indicated by increased expression of Gli1) was associated with the reductions in Ift88 expression in the tibial cartilage from Ift88-deficient mice. Wheel exercise restored both the articular cartilage thickness and levels of Hh signaling in these mice. CONCLUSION: Our results in a mouse model of OA demonstrate that IFT88 performs a chondroprotective role in articular cartilage by controlling the calcification of cartilage via maintenance of a threshold of Hh signaling during physiologic loading.