Ependymal polarity defects coupled with disorganized ciliary beating drive abnormal cerebrospinal fluid flow and spine curvature in zebrafish.

Idiopathic scoliosis (IS) is the most common spinal deformity diagnosed in childhood or early adolescence, while the underlying pathogenesis of this serious condition remains largely unknown. Here, we report zebrafish ccdc57 mutants exhibiting scoliosis during late development, similar to that observed in human adolescent idiopathic scoliosis (AIS). Zebrafish ccdc57 mutants developed hydrocephalus due to cerebrospinal fluid (CSF) flow defects caused by uncoordinated cilia beating in ependymal cells. Mechanistically, Ccdc57 localizes to ciliary basal bodies and controls the planar polarity of ependymal cells through regulating the organization of microtubule networks and proper positioning of basal bodies. Interestingly, ependymal cell polarity defects were first observed in ccdc57 mutants at approximately 17 days postfertilization, the same time when scoliosis became apparent and prior to multiciliated ependymal cell maturation. We further showed that mutant spinal cord exhibited altered expression pattern of the Urotensin neuropeptides, in consistent with the curvature of the spine. Strikingly, human IS patients also displayed abnormal Urotensin signaling in paraspinal muscles. Altogether, our data suggest that ependymal polarity defects are one of the earliest sign of scoliosis in zebrafish and disclose the essential and conserved roles of Urotensin signaling during scoliosis progression.

Development of Phenyl-substituted Isoindolinone- and Benzimidazole-type Cereblon Ligands for Targeted Protein Degradation.

Thalidomide, pomalidomide and lenalidomide, collectively referred to as immunomodulatory imide drugs (IMiDs), are frequently employed in proteolysis-targeting chimeras (PROTACs) as cereblon (CRBN) E3 ligase-recruiting ligands. However, their molecular glue properties that co-opt the CRL4CRBN to degrade its non-natural substrates may lead to undesired off-target effects for the IMiD-based PROTAC degraders. Herein, we reported a small library of potent and cell-permeable CRBN ligands, which exert high selectivity over the well-known CRBN neo-substrates of IMiDs by structure-based design. They were further utilized to construct bromodomain-containing protein 4 (BRD4) degraders, which successfully depleted BRD4 in the tested cells. Overall, we reported a series of functionalized CRBN recruiters that circumvent the promiscuity from traditional IMiDs, and this study is informative to the development of selective CRBN-recruiting PROTACs for many other therapeutic targets.

Heme-binding protein CYB5D1 is a radial spoke component required for coordinated ciliary beating.

Coordinated beating is crucial for the function of multiple cilia. However, the molecular mechanism is poorly understood. Here, we characterize a conserved ciliary protein CYB5D1 with a heme-binding domain and a cordon-bleu ubiquitin-like domain. Mutation or knockdown of Cyb5d1 in zebrafish impaired coordinated ciliary beating in the otic vesicle and olfactory epithelium. Similarly, the two flagella of an insertional mutant of the CYB5D1 ortholog in Chlamydomonas (Crcyb5d1) showed an uncoordinated pattern due to a defect in the cis-flagellum. Biochemical analyses revealed that CrCYB5D1 is a radial spoke stalk protein that binds heme only under oxidizing conditions. Lack of CrCYB5D1 resulted in a reductive shift in flagellar redox state and slowing down of the phototactic response. Treatment of Crcyb5d1 with oxidants restored coordinated flagellar beating. Taken together, these data suggest that CrCYB5D1 may integrate environmental and intraciliary signals and regulate the redox state of cilia, which is crucial for the coordinated beating of multiple cilia.

The role of PANoptosis in renal vascular endothelial cells: Implications for trichloroethylene-induced kidney injury.

Trichloroethylene (TCE), a widely distributed environmental chemical contaminant, is extensively dispersed throughout the environment. Individuals who are exposed to TCE may manifest occupational medicamentose-like dermatitis due to trichloroethylene (OMDT). Renal impairment typically manifests in the initial phase of OMDT and is intricately linked to the disease progression and patient outcomes. Although recombinant human tumor necrosis factor-α receptor II fusion protein (rh TNFR:Fc) has been employed in the clinical management of OMDT, there was no substantial improvement in renal function observed in patients following one week of treatment. This study primarily examined the mechanism of TNFα- and IFNγ-induced endothelial cells (ECs) PANoptosis in TCE-induced kidney injury and hypothesized that the synergistic effect of TNFα and IFNγ could be the key factor affecting the efficacy of rh TNFR:Fc therapy in OMDT patients. A TCE-sensitized mouse model was utilized in this study to investigate the effects of TNFα and IFNγ neutralizing antibodies on renal vascular endothelial cell PANoptosis. The gene of interferon regulatory factor 1 (IRF1) in human umbilical vein endothelial cells (HUVEC) was silenced by using small interfering RNA (siRNA), and the cells were then treated with TNFα and IFNγ recombinant protein to investigate the mechanism of TNFα combined with IFNγ-induced PANoptosis in HUVEC. The findings indicated that mice sensitized to TCE exhibited increased levels of PANoptosis-related markers in renal endothelial cells, and treatment with TNFα and IFNγ neutralizing antibodies resulted in a significant reduction in PANoptosis and improvement in renal function. In vitro experiments demonstrated that silencing IRF1 could reverse TNFα and IFNγ-induced PANoptosis in endothelial cells. These results suggest that the efficacy of rh TNFR:Fc may be influenced by TNFα and IFNγ-mediated PANoptosis in kidney vascular endothelial cells. The joint application of TNFα and IFNγ neutralizing antibody represented a solid alternative to existing therapeutics.

Multifunctional Cellulose Nanocrystals Electrolyte Additive Enable Ultrahigh-Rate and Dendrite-Free Zn Anodes for Rechargeable Aqueous Zinc Batteries.

The design of aqueous zinc (Zn) chemistry energy storage with high rate-capability and long serving life is a great challenge due to its inhospitable coordination environment and dismal interfacial chemistry. To bridge this big gap, herein, we build a highly reversible aqueous Zn battery by taking advantages of the biomass-derived cellulose nanocrystals (CNCs) electrolyte additive with unique physical and chemical characteristics simultaneously. The CNCs additive not only serves as fast ion carriers for enhancing Zn2+ transport kinetics but regulates the coordination environment and interface chemistry to form dynamic and self-repairing protective interphase, resulting in building ultra-stable Zn anodes under extreme conditions. As a result, the engineered electrolyte system achieves a superior average coulombic efficiency of 97.27 % under 140 mA cm-2, and steady charge-discharge for 982 h under 50 mA cm-2, 50 mAh cm-2, which proposes a universal pathway to challenge aqueous Zn chemistry in green, sustainable, and large-scale applications.

Diptoindonesin G is a middle domain HSP90 modulator for cancer treatment.

HSP90 inhibitors can target many oncoproteins simultaneously, but none have made it through clinical trials due to dose-limiting toxicity and induction of heat shock response, leading to clinical resistance. We identified diptoindonesin G (dip G) as an HSP90 modulator that can promote degradation of HSP90 clients by binding to the middle domain of HSP90 (Kd = 0.13 ± 0.02 µM) without inducing heat shock response. This is likely because dip G does not interfere with the HSP90-HSF1 interaction like N-terminal inhibitors, maintaining HSF1 in a transcriptionally silent state. We found that binding of dip G to HSP90 promotes degradation of HSP90 client protein estrogen receptor α (ER), a major oncogenic driver protein in most breast cancers. Mutations in the ER ligand-binding domain (LBD) are an established mechanism of endocrine resistance and decrease the binding affinity of mainstay endocrine therapies targeting ER, reducing their ability to promote ER degradation or transcriptionally silence ER. Because dip G binds to HSP90 and does not bind to the LBD of ER, unlike endocrine therapies, it is insensitive to ER LBD mutations that drive endocrine resistance. Additionally, we determined that dip G promoted degradation of WT and mutant ER with similar efficacy, downregulated ER- and mutant ER-regulated gene expression, and inhibited WT and mutant cell proliferation. Our data suggest that dip G is not only a molecular probe to study HSP90 biology and the HSP90 conformation cycle, but also a new therapeutic avenue for various cancers, particularly endocrine-resistant breast cancer harboring ER LBD mutations.

A Modular Chemistry Platform for the Development of a Cereblon E3 Ligase-Based Partial PROTAC Library.

Proteolysis targeting chimeras (PROTACs) are a promising therapeutic strategy to selectively promote the degradation of protein targets by exploiting the ubiquitin-proteasome system. Among the limited number of E3 ligase ligands discovered for the PROTAC technology, ligands of cereblon (CRBN) E3 ligase, such as pomalidomide, thalidomide, or lenalidomide, are the most frequently used for the development of PROTACs. Our group previously reported that a phenyl group could be tolerated on the C4-position of lenalidomide as the ligand of CRBN to develop PROTACs. Herein, we report a modular chemistry platform for the efficient attachment of various ortho-, meta-, and para-substituted phenyls to the C4-position of the lenalidomide via Suzuki cross-coupling reaction, which allows the systematic investigation of the linker effect for the development of PROTACs against any target. We examined the substrate scope by preparing twelve lenalidomide-derived CRBN E3 ligase ligands with different linkers.

Zebrafish: an important model for understanding scoliosis.

Scoliosis is a common spinal deformity that considerably affects the physical and psychological health of patients. Studies have shown that genetic factors play an important role in scoliosis. However, its etiopathogenesis remain unclear, partially because of the genetic heterogeneity of scoliosis and the lack of appropriate model systems. Recently, the development of efficient gene editing methods and high-throughput sequencing technology has made it possible to explore the underlying pathological mechanisms of scoliosis. Owing to their susceptibility for developing scoliosis and high genetic homology with human, zebrafish are increasingly being used as a model for scoliosis in developmental biology, genetics, and clinical medicine. Here, we summarize the recent advances in scoliosis research on zebrafish and discuss the prospects of using zebrafish as a scoliosis model.

E2f5 is a versatile transcriptional activator required for spermatogenesis and multiciliated cell differentiation in zebrafish.

E2f5 is a member of the E2f family of transcription factors that play essential roles during many cellular processes. E2f5 was initially characterized as a transcriptional repressor in cell proliferation studies through its interaction with the Retinoblastoma (Rb) protein for inhibition of target gene transcription. However, the precise roles of E2f5 during embryonic and post-embryonic development remain incompletely investigated. Here, we report that zebrafish E2f5 plays critical roles during spermatogenesis and multiciliated cell (MCC) differentiation. Zebrafish e2f5 mutants develop exclusively as infertile males. In the mutants, spermatogenesis is arrested at the zygotene stage due to homologous recombination (HR) defects, which finally leads to germ cell apoptosis. Inhibition of cell apoptosis in e2f5;tp53 double mutants rescued ovarian development, although oocytes generated from the double mutants were still abnormal, characterized by aberrant distribution of nucleoli. Using transcriptome analysis, we identified dmc1, which encodes an essential meiotic recombination protein, as the major target gene of E2f5 during spermatogenesis. E2f5 can bind to the promoter of dmc1 to promote HR, and overexpression of dmc1 significantly increased the fertilization rate of e2f5 mutant males. Besides gametogenesis defects, e2f5 mutants failed to develop MCCs in the nose and pronephric ducts during early embryonic stages, but these cells recovered later due to redundancy with E2f4. Moreover, we demonstrate that ion transporting principal cells in the pronephric ducts, which remain intercalated with the MCCs, do not contain motile cilia in wild-type embryos, while they generate single motile cilia in the absence of E2f5 activity. In line with this, we further show that E2f5 activates the Notch pathway gene jagged2b (jag2b) to inhibit the acquisition of MCC fate as well as motile cilia differentiation by the neighboring principal cells. Taken together, our data suggest that E2f5 can function as a versatile transcriptional activator and identify novel roles of the protein in spermatogenesis as well as MCC differentiation during zebrafish development.

HMGB 1 acetylation mediates trichloroethylene-induced immune kidney injury by facilitating endothelial cell-podocyte communication.

More and more clinical evidence shows that occupational medicamentose-like dermatitis due to trichloroethylene (OMDT) patients often present immune kidney damage. However, the exact mechanisms of cell-to-cell transmission in TCE-induced immune kidney damage remain poorly understood. The present study aimed to explore the role of high mobility group box-1 (HMGB 1) in glomerular endothelial cell-podocyte transmission. 17 OMDT patients and 34 controls were enrolled in this study. We observed that OMDT patients had renal function injury, endothelial cell activation and podocyte injury, and these indicators were associated with serum HMGB 1. To gain mechanistic insight, a TCE-sensitized BALB/c mouse model was established under the interventions of sirtuin 1 (SIRT 1) activator SRT 1720 (0.1 ml, 5 mg/kg) and receptor for advanced glycation end products (RAGE) inhibitor FPS-ZM 1 (0.1 ml, 1.5 mg/kg). We identified HMGB 1 acetylation and its endothelial cytoplasmic translocation following TCE sensitization, but SRT 1720 abolished the process. RAGE was located on podocytes and co-precipitated with extracellular acetylated HMGB 1, promoting podocyte injury, while SRT 1720 and FPS-ZM 1 both alleviated podocyte injury. The results demonstrate that interventions to upstream and downstream pathways of HMGB 1 may weaken glomerular endothelial cell-podocyte transmission, thereby alleviating TCE-induced immune renal injury.

Multistimuli-responsive fluorescence behaviours of aggregation-induced emission small-molecule and electrospun nanofibre films for acid-base vapour sensing.

Multistimuli-responsive fluorescent materials have garnered great research interest benefited from their practical applications. Two twisted-structure compounds containing tetraphenylethylene (TPE) as the aggregation-induced emission (AIE) group and a pyridine unit as the acid reaction site to obtain new multistimuli-responsive fluorescent compounds (namely, TPECNPy: TPECNPy-2 and TPECNPy-3) were successfully synthesized through a one-step Knoevenagel condensation reaction. The multiple-stimuli response process of TPECNPy was investigated by means of photoluminescence (PL) spectra and emission colour. The results showed that both TPECNPy compounds with excellent AIE abilities displayed reversible emission wavelength and colour changes in response to multiple external stimuli, including grinding-fuming by CH2 Cl2 or annealing and HCl-NH3 vapour fuming. More importantly, fluorescent nanofibre films were prepared by electrospinning a solution of TPECNPy mixed with cellulose acetate (CA), and these exhibited reversible acid-induced discolouration, even with only 1 wt% TPECNPy. The results of this study may inspire strategies for designing multistimuli-responsive materials and preparing fluorescent sensing nanofibre films.

The role of Kupffer cell activation in immune liver damage induced by trichloroethylene associated with the IFN-γ/STAT1 signaling pathway.

Trichloroethylene (TCE) is a metal detergent commonly used in industry that can enter the human body through the respiratory tract and skin, causing occupational medicamentosa-like dermatitis due to TCE (OMDT) and multiple organ damage, including liver failure. However, the pathogenesis of liver injury remains unclear. Kupffer cells (KCs) are important tissue macrophages in the body because the polarization of KCs plays a crucial role in immune-mediated liver injury. However, the mechanism of KCs polarization in TCE-induced immune liver injury has not been thoroughly elucidated. In this study, we investigated the effect of TCE-induced KCs polarization on liver function and signal transduction pathways using the TCE sensitization model developed by our group. BALB/c mouse skin was exposed to TCE for sensitization, and an increase in the expression of M1 macrophage-specific markers (CD16/CD32, iNOS), M1 macrophage-specific cytokines IL-1ß, and IFN-γ, P-JAK-1 and P-STAT1 levels were also found to be dramatically increased. When using low doses of gadolinium trichloride (GdCl3), the expression of these proteins and mRNA was significantly reduced. This phenomenon indicates that GdCl3 blocks TCE-induced polarization of KCs and suggests that the IFN-γ/STAT1 signaling pathway may be involved in the polarization process of KCs. These findings clarify the relationship between the polarization of KCs and immune liver injury and highlight the importance of further study of immune-mediated liver injury in TCE-sensitized mice.

E2f4 is required for intestinal and otolith development in zebrafish.

E2f4 is a multifunctional transcription factor that is essential for many cellular processes. Although the role of E2f4 during cell cycle progression has been investigated in great detail, less is known about E2f4 during embryonic development. Here, we investigated the role of E2f4 during zebrafish development. Zebrafish e2f4 mutants displayed ectopic otolith formation due to abnormal ciliary beating in the otic vesicle. The beating defects of motile cilia were caused by abnormal expression of ciliary motility genes. The expression of two genes, lrrc23 and ccdc151, were significantly decreased in the absence of E2f4. In addition to that, e2f4 mutants also displayed growth retardation both in the body length and body weight and mostly died at around 6 months old. Although food intake was normal in the mutants, we found that the microvilli of the intestinal epithelia were significantly affected in the mutants. Finally, the intestinal epithelia of e2f4 mutants also displayed reduced cell proliferation, together with an increased level of cell apoptosis. Our data suggested a tissue-specific role of E2f4 during zebrafish development, which is distinct from the traditional views of this protein as a transcription repressor.

TNF-α/TNFR1 regulates the polarization of Kupffer cells to mediate trichloroethylene-induced liver injury.

We have previously shown trichloroethylene (TCE) induced immune liver injury, and TNF-α/TNFR1 pathway as a probably mechanism underlying the immune damage, but the pathogenic mechanism is still unclear. The study aims to investigate whether TNF-α and its receptors regulate Kupffer cell polarization and downstream inflammation signaling pathways during TCE sensitization, to clarify the mechanism of TCE-mediated immune liver injury. 6-8 weeks old SPF BALB/c female mice were used to establish a TCE sensitization model. We found that in the TCE sensitization positive group, liver injury was aggravated, Kupffer cells activated and polarized to M1 type. The expression of M1 Kupffer cell marker proteins CD11c and CD16/32 increased in the TCE positive group, so did TNF-α and TNFR1 in liver. The expression of P-IKK protein, PP65 protein and P-STAT3 protein increased in the TCE sensitization positive group, and the downstream inflammatory factors IL-1ß and IL-6 also increased in the TCE sensitization positive group. After using the TNFR1 inhibitor R7050, we found that M1 Kupffer cell polarization, TNF-α expression, signal pathway expression and inflammatory factors IL-1ß and IL-6 expression declined, and the liver damage relieved. Briefly, the use of R7050 to inhibit TNF-α/TNFR1 changing the polarization of liver M1 Kupffer cell, thereby inhibiting the activation of related downstream signaling pathways and reducing the secretion of inflammatory factors. TNF-α/TNFR1 regulates the polarization of M1 Kupffer cells inflammatory play an important role in liver immune damage.

Effects of mitochondrial reactive oxygen species-induced NLRP3 inflammasome activation on trichloroethylene-mediated kidney immune injury.

This study aimed to investigate the activating mechanism of the NLRP3 inflammasome in trichloroethylene-sensitized mice. In total, 88 BALB/c female mice were used to establish the trichloroethylene (TCE)-sensitized mouse model. Some of the mice received MitoTEMPO, MCC 950 or soluble recombinant CD59-Cys to inhibit mitochondrial reactive oxygen species (mtROS) production, NLRP3 assembly, or C5b-9 formation. Mouse tubular epithelial cell expression levels of NLRP3, ASC, Caspase 1, IL-1ß, IL-18 and mitochondrial antiviral signaling protein (MAVS) were detected by western blot. Mitochondrial numbers, membrane potential (ΔΨm) and mtROS were detected by using MitoScene Green II, JC-1 dye and MitoSOX Red indicator, respectively. Tubular epithelial cell calcium levels were detected by a Fluo-8 no wash calcium assay kit. Human kidney-2 (HK-2) cells were cultured and stimulated by C5b6 and normal human serum (NHS) to verify the role of C5b-9-induced mitochondrial ROS in activating NLRP3 inflammasome. Urine α1-MG, ß2-MG, and mtROS production and calcium levels were increased, while mitochondrial numbers were decreased in TCE-sensitized positive mice. After treatment with MitoTEMPO, renal tubular injury was alleviated, JC-1 fluorescence and mitochondrial numbers were significantly increased, and mitochondrial ROS were inhibited. The NLRP3 inflammasome was activated in TCE-sensitized positive mice, while Mito TEMPO inhibited MAVS expression and NLRP3 inflammasome activation. The in vitro studies proved that C5b-9 can induce mtROS release and activate the assembly of NLRP3 inflammasome in HK-2 cells. In conclusion, in TCE-sensitized positive mouse renal tubular epithelial cells, C5b-9 caused calcium influx and thus induced mitochondrial injury and mtROS overexpression, finally inducing MAVS expression and NLRP3 inflammasome activation and kidney injury.

Endothelin-1 down-regulated vascular endothelial growth factor A is involved in trichloroethene-induced kidney injury.

The mechanism of kidney injury in occupational medicamentosa-like dermatitis due to trichloroethylene exposure is not well understood. This study aimed to investigate the role of endothelin-1 (ET-1)/vascular endothelial-derived growth factor-A (VEGF-A) in trichloroethylene (TCE)-induced renal injury. Forty BALB/c female mice were used in this study to build the TCE-sensitization mouse model. Transmission electron microscopic observation, histological examination, periodic acid-Schiff staining, serum urea nitrogen, creatinine, and urinary total protein levels were used to reflect renal injury. Glypican1, syndecan1, ET-1 and VEGF-A protein levels were measured by western blot. Serum ET-1 level was also measured. Tumor necrosis factor alpha (TNF-α) and vascular cell adhesion molecule-1 (VCAM-1) were detected by immunohistochemistry. The results showed that TCE-sensitized mouse kidneys were damaged and accompanied by increased serum ET-1. After treatment with CGS 35066, the inhibitor of endothelin converting enzyme-1 (ECE-1), kidney ET-1, TNF-α and VCAM-1 levels decreased, and renal function improved in TCE+CGS 35066-sensitized positive mice. In addition, kidney VEGF-A, glomerular endothelial cell glypican1 and syndecan1 levels increased, and endothelial cell damage was alleviated after treatment with CGS 35066. The results suggest that inhibiting ECE-1 could alleviate glomerular endothelial cell injury by inhibiting ET-1 expression, thus promoting endothelial cell repair by upregulating VEGF-A.

Distribution pattern of floor plate neurons in zebrafish.

The floor plate (FP) is a critical signaling center for the development of neural tube and body axis, and is localized at the ventral midline of the neural tube. Multiple types of neurons are present in the floor plate, while the distribution pattern of these neuronal cells remains unclear. By using transgenic zebrafish lines that specifically label different neuronal cells, we investigated the distribution pattern of these neurons in the floor plate region. Our results showed that foxj1a, sox2, clusterin and gfap genes were expressed in the medial floor plate (MFP), consisting of a single row of cells. The cerebrospinal fluid-contacting neurons (CSF-cNs), also named as Kolmer-Agduhr interneurons (KA' and KA" neurons), were located on the lateral sides of MFP. The foxj1a and pkd2l1 genes were expressed in the KA" neurons, which were located to the ventral terminal gap of wedge-shaped MFP cells. The neighboring KA" neurons were separated by neurons expressing Gfap, Olig2 or Sox2. In contrast, the KA' neurons were positive for Foxj1a +/Pkd2l1+/Olig2+, and were localized to the dorsal side of KA" neurons. Similarly, the Sox2 or Olig2 expressing neurons were intermingled with KA' neurons along the anterior-posterior axis in these regions. Further pharmaceutical treatment demonstrated that interference of Notch signaling resulted in the abnormal distribution of floor plate neurons together with strong dorsal body curvature at 3 days post fertilization in the zebrafish larvae. Our data showed the gene expression patterns and relative positions of the floor plate neurons; and suggested a potential role of Notch signaling during floor plate development.

C5b-9 membrane attack complex activated NLRP3 inflammasome mediates renal tubular immune injury in trichloroethylene sensitized mice.

Trichloroethylene (TCE) induced occupational medicamentosa-like dermatitis (OMLDT) in patients is accompanied, typically, by renal damage. But the role of C5b-9 and IL-1ß in TCE-sensitized mouse renal tubular damage is unclear. This study aimed to investigate whether TCE-sensitized mouse renal tubular epithelial cell damage was induced by NLRP3 inflammasome and whether NLRP3 inflammasome was activated by sublytic C5b-9. In total, 52 specific pathogen-free BALB/c female mice, 6- to 8-week-old, were used for establishing the TCE-sensitized mouse model. Renal tubular epithelial cells were isolated and used for determining the sublytic level of C5b-9. Kidney histological examination, serum neutrophil gelatinase associated lipocalin (NGAL) level were used for kidney damage evaluation. Renal protein levels of C5b-9, NLRP3, ASC, Caspase-1, IL-1ß, and IL-18 were measured. The renal lesions, serum NGAL level, renal NLRP3, ASC, Caspase-1 and IL-1ß protein levels all increased significantly in TCE sensitized positive group. However, pretreatment with recombinant protein sCD59-Cys inhibited the expression of C5b-9, NLRP3 inflammasome, IL-1ß, IL-18, and attenuated renal tubular epithelial cell damage. The sublytic C5b-9 activated NLRP3 inflammasome and aggravated renal tubular epithelial cell damage. Pretreatment with recombinant protein sCD59-Cys blocked the expression of the NLRP3 inflammasome by inhibiting the expression of C5b-9, and alleviating renal tubular epithelial cell damage.

Radial spoke proteins regulate otolith formation during early zebrafish development.

Radial spokes are structurally conserved, macromolecular complexes that are essential for the motility of 9 + 2 motile cilia. In Chlamydomonas species, mutations in radial spoke proteins result in ciliary motility defects. However, little is known about the function of radial spoke proteins during embryonic development. Here, we investigated the role of a novel radial spoke protein, leucine-rich repeat containing protein 23 (Lrrc23), during zebrafish embryonic development. Mutations in lrrc23 resulted in a selective otolith formation defect during early ear development. Similar otolith defects were also present in the radial spoke head 3 homolog ( rsph3) and radial spoke head 4 homolog A ( rsph4a) radial spoke mutants. Notably, the radial spoke protein mutations specifically affected ciliary motility in the otic vesicle (OV), whereas motile cilia in other organs functioned normally. Via high-speed video microscopy, we found that motile cilia formation was stochastic and transient in the OV. Importantly, all the motile cilia in the OV beat circularly, in contrast to the planar beating pattern of typical 9 + 2 motile cilia. We identified the key time frame for motile cilia formation during OV development. Finally, we showed that the functions of radial spoke proteins were conserved between zebrafish and Tetrahymena. Together, our results suggest that radial spoke proteins are essential for ciliary motility in the OV and that radial spoke-regulated OV motile cilia represent a unique type of cilia during early zebrafish embryonic development.-Han, X., Xie, H., Wang, Y., Zhao, C. Radial spoke proteins regulate otolith formation during early zebrafish development.

Discovery of 2,3'-diindolylmethanes as a novel class of PCSK9 modulators.

Proprotein convertase subtilisin/kexin type 9 (PCSK9) promotes the degradation of low density lipoprotein receptor (LDLR). Anti-PCSK9 agents have been approved for the treatment of hypercholesterolemia. We recently discovered a series of small-molecule PCSK9 modulators that contains a relatively small pharmacophore of 2,3'-diindolylmethane with molecular weights around only 250. These molecules can significantly lower the amount of PCSK9 protein in a cell-based phenotypic assay. Our SAR studies yielded compound 16 with a IC50-value of 200 nM. No obvious cytotoxicity was observed at concentrations below 50 µM.