A simple protocol to establish a conditionally immortalized mouse podocyte cell line.

Podocytes are specialized terminally differentiated cells in the glomerulus that are the primary target cells in many glomerular diseases. However, the current podocyte cell lines suffer from prolonged in vitro differentiation and limited survival time, which impede research progress. Therefore, it is necessary to establish a cell line that exhibits superior performance and characteristics. We propose a simple protocol to obtain an immortalized mouse podocyte cell (MPC) line from suckling mouse kidneys. Primary podocytes were cultured in vitro and infected with the SV40 tsA58 gene to obtain immortalized MPCs. The podocytes were characterized using Western blotting and quantitative real-time PCR. Podocyte injury was examined using the Cell Counting Kit-8 assay and flow cytometry. First, we successfully isolated an MPC line and identified 39 °C as the optimal differentiation temperature. Compared to undifferentiated MPCs, the expression of WT1 and synaptopodin was upregulated in differentiated MPCs. Second, the MPCs ceased proliferating at a nonpermissive temperature after day 4, and podocyte-specific proteins were expressed normally after at least 15 passages. Finally, podocyte injury models were induced to simulate podocyte injury in vitro. In summary, we provide a simple and popularized protocol to establish a conditionally immortalized MPC, which is a powerful tool for the study of podocytes.

Menin Reduces Parvalbumin Expression and is Required for the Anti-Depressant Function of Ketamine.

Dysfunction of parvalbumin (PV) neurons is closely involved in depression, however, the detailed mechanism remains unclear. Based on the previous finding that multiple endocrine neoplasia type 1 (Protein: Menin; Gene: Men1) mutation (G503D) is associated with a higher risk of depression, a Menin-G503D mouse model is generated that exhibits heritable depressive-like phenotypes and increases PV expression in brain. This study generates and screens a serial of neuronal specific Men1 deletion mice, and found that PV interneuron Men1 deletion mice (PcKO) exhibit increased cortical PV levels and depressive-like behaviors. Restoration of Menin, knockdown PV expression or inhibition of PV neuronal activity in PV neurons all can ameliorate the depressive-like behaviors of PcKO mice. This study next found that ketamine stabilizes Menin by inhibiting protein kinase A (PKA) activity, which mediates the anti-depressant function of ketamine. These results demonstrate a critical role for Menin in depression, and prove that Menin is key to the antidepressant function of ketamine.

A Low-Profile Dielectric Resonator Filter with Wide Stopband for High Integration on PCB.

A low-profile dielectric resonator (DR) filter is proposed to achieve the feature of high integration and wide stopband. The high integration is due to the structure of printed circuit board (PCB) substrate instead of metal cavity, which can be easily integrated with other planar circuits. Thus, the proposed design can improve the integration level and reduce installation errors. Moreover, the out-of-band harmonics can be well suppressed by the structure combined with introducing rectangular hollowing in the center of the dielectric block, coupling the feed and loading 1/4λ wavelength branch. For demonstration, it is fabricated and measured. The simulated and experimental results with good agreement are presented, the insertion loss is as low as 1.1 dB, the profile height is only 0.77λg, and the stopband reaches 2.61f0.

Salvianolic acid B ameliorates retinal deficits in an early-stage Alzheimer's disease mouse model through downregulating BACE1 and Aß generation.

Alzheimer's disease (AD) is a neurodegenerative disease with subtle onset, early diagnosis remains challenging. Accumulating evidence suggests that the emergence of retinal damage in AD precedes cognitive impairment, and may serve as a critical indicator for early diagnosis and disease progression. Salvianolic acid B (Sal B), a bioactive compound isolated from the traditional Chinese medicinal herb Salvia miltiorrhiza, has been shown promise in treating neurodegenerative diseases, such as AD and Parkinson's disease. In this study we investigated the therapeutic effects of Sal B on retinopathy in early-stage AD. One-month-old transgenic mice carrying five familial AD mutations (5×FAD) were treated with Sal B (20 mg·kg-1·d-1, i.g.) for 3 months. At the end of treatment, retinal function and structure were assessed, cognitive function was evaluated in Morris water maze test. We showed that 4-month-old 5×FAD mice displayed distinct structural and functional deficits in the retinas, which were significantly ameliorated by Sal B treatment. In contrast, untreated, 4-month-old 5×FAD mice did not exhibit cognitive impairment compared to wild-type mice. In SH-SY5Y-APP751 cells, we demonstrated that Sal B (10 µM) significantly decreased BACE1 expression and sorting into the Golgi apparatus, thereby reducing Aß generation by inhibiting the ß-cleavage of APP. Moreover, we found that Sal B effectively attenuated microglial activation and the associated inflammatory cytokine release induced by Aß plaque deposition in the retinas of 5×FAD mice. Taken together, our results demonstrate that functional impairments in the retina occur before cognitive decline, suggesting that the retina is a valuable reference for early diagnosis of AD. Sal B ameliorates retinal deficits by regulating APP processing and Aß generation in early AD, which is a potential therapeutic intervention for early AD treatment.

Amelioration of olfactory dysfunction in a mouse model of Parkinson's disease via enhancing GABAergic signaling.

BACKGROUND: Olfactory dysfunction is among the earliest non-motor symptoms of Parkinson's disease (PD). As the foremost pathological hallmark, α-synuclein initiates the pathology in the olfactory pathway at the early stage of PD, particularly in the olfactory epithelium (OE) and olfactory bulb (OB). However, the local neural microcircuit mechanisms underlying olfactory dysfunction between OE and OB in early PD remain unknown. RESULTS: We observed that odor detection and discrimination were impaired in 6-month-old SNCA-A53T mice, while their motor ability remained unaffected. It was confirmed that α-synuclein increased and accumulated in OB but not in OE. Notably, the hyperactivity of mitral/tufted cells and the excitation/inhibition imbalance in OB were found in 6-month-old SNCA-A53T mice, which was attributed to the impaired GABAergic transmission and aberrant expression of GABA transporter 1 and vesicular GABA transporter in OB. We further showed that tiagabine, a potent and selective GABA reuptake inhibitor, could reverse the impaired olfactory function and GABAergic signaling in OB of SNCA-A53T mice. CONCLUSIONS: Taken together, our findings demonstrate potential synaptic mechanisms of local neural microcircuit underlying olfactory dysfunction at the early stage of PD. These results highlight the critical role of aberrant GABAergic signaling of OB in early diagnosis and provide a potential therapeutic strategy for early-stage PD.

Selective C-O Bond Forming Reactions at Indole-C2-Position toward Polycyclic Indolone or Indolinone Derivatives Tethered with Medium-Sized Rings.

A methodology involving the chemoselective synthesis of tetracyclic [1,3]oxazino[3,2-a]indol-4-one or tetracyclic [1,3]oxazino[3,2-a]indoline-4-one tethered with a medium-sized ring by cross dehydrogenative coupling (CDC) or nucleophilic addition (NA) reaction has been developed. [1,3]Oxazino[3,2-a]indol-4-one compounds fused with a medium-sized ring were constructed through a CDC reaction in the presence of I2 and K2CO3. Whereas, [1,3]oxazino[3,2-a]indoline-4-ones tethered with a medium-sized ring were obtained with a TfOH system by NA reaction.

The Efficacy and Safety of Brolucizumab for the Treatment of nAMD: A Systematic Review and Meta-Analysis.

Introduction: As demonstrated in pivotal clinical trials, brolucizumab can be used to treat neovascular age-related macular degeneration (nAMD) because it antagonizes vascular endothelial growth factor (VEGF) in the vitreous. However, brolucizumab may cause retinal vasculitis obliterans in the presence of inflammation in the eyes. In the present study, a meta-analysis of randomized controlled trials (RCTs) was conducted to evaluate the efficacy and safety of brolucizumab. Methods: ClinicTrail.gov., Embase, Cochrane Library, and PubMed were retrieved from inception until 31 December 2021 for RCTs assessing the efficacy and safety of brolucizumab. Changes in best corrected visual acuity (BCVA) and central sub-field thickness (CSFT) and incidence of adverse events, serious adverse events, and serious ocular adverse events were extracted from eligible RCTs. A meta-analysis was performed using RevMan 5.4.1. Results: A total of six RCTs with 3,574 participants were finally involved in this meta-analysis. The changes of best corrected visual acuity (BCVA) showed no statistically significant difference between the brolucizumab-treated group and aflibercept-treated group. Brolucizumab induced higher central sub-field thickness (CSFT) reduction than the control agent (aflibercept). The incidence of adverse events was similar between the brolucizumab group and control group (OR 0.63, 95% CI 0.37 to 1.08, p = 0.09), and brolucizumab caused fewer serious adverse events (OR 0.78, 95% CI 0.63 to 0.95, p = 0.01). However, brolucizumab could lead to more serious ocular adverse events than Lucentis and aflibercept (OR 2.15, 95% CI 1.11 to 4.16, p = 0.02). Conclusion: Brolucizumab was non-inferior to other anti-VEGF agents in improving BCVA and decreasing CSFT. But it caused more serious ocular adverse events which is worthy of special attention by ophthalmologists.

Straightforward Stereoselective Synthesis of Seven-Membered Oxa-Bridged Rings through In Situ Generated Cycloheptenol Derivatives.

An iodine-mediated stereoselective synthesis of seven-membered oxa-bridged rings via in situ generated cycloheptenols was reported. This process was realized through the combination of C-C σ-bond cleavage and C-O bond-forming reactions in a one-pot fashion from simple and easily accessible raw materials. The formation of carbon radicals initiated by I2 was the key to the reaction.

Difunctionalization of Alkynones by Base-Mediated Reaction with α,α-Dithioketones.

A novel 1,2-difunctionalization of alkynones via an umpolung strategy for the synthesis of tetrasubstituted olefins has been developed. This procedure is realized by a formal C-C σ-bond cleavage reaction of cyclic α,α-dithioketones and subsequent deprotection. Notable features of this approach include excellent yields, mild reaction conditions, a broad substrate scope, and operational simplicity.

Atom-Economic Synthesis of Highly Functionalized Bridged Ring Systems Initiated by Ring Expansion of Indene-1,3-dione.

An atom economic procedure for the regioselective synthesis of bridged seven-membered-ring compounds from simple reactants such as ynones and indene-1,3-dione has been developed. This process was realized through the one-pot reactions of ring-expansion of indene-1,3-dione with alkynyl ketones and successive formal [4+2] cycloaddition. The Michael addition reaction is the key for the regioselectivity of the formal [4+2] cycloaddition.

Selective Synthesis of Pyrano[3,2-b]indoles or Cyclopenta[b]indoles Tethered with Medium-Sized Rings via Cascade C-C σ-Bond Cleavage and C-H Functionalization.

Highly atom-economical tandem reactions have been developed for the synthesis of pyrano[3,2-b]indoles or cyclopenta[b]indoles tethered with 7-, 8-, or 9-membered rings. These reactions first undergo a carbon-carbon σ-bond cleavage reaction of cyclic ß-ketoesters. Next, in the presence of CuCl2 and Ag2CO3, intramolecular O-H/C-H coupling occurs to give pyrano[3,2-b]indoles. This is the first example for capture of the enoloxyl radical of the intramolecular C-O bond formation reaction, whereas C3 nucleophilic addition afforded cyclopenta[b]indoles using TsOH·H2O.

Differential orthogonal frequency division multiplexing communication in water pipeline channels.

The problem of information transmission through water pipelines is addressed and a class of methods based on differentially encoded orthogonal frequency division multiplexing (OFDM) is proposed. Specifically, two methods are investigated; one based on conventional differential encoding with an estimation of the average time of arrival and another based on double encoding. Results show that the first approach improves performance in the low signal to noise ratio (SNR) region, while the second is better suited at high SNR. Adopting these techniques closes the performance gap between differential OFDM systems and coherent OFDM systems while retaining the benefits of computational simplicity.

Cyclin-Dependent Kinase 5-Dependent BAG3 Degradation Modulates Synaptic Protein Turnover.

BACKGROUND: Synaptic protein dyshomeostasis and functional loss is an early invariant feature of Alzheimer's disease (AD), yet the unifying etiological pathway remains largely unknown. Knowing that cyclin-dependent kinase 5 (CDK5) plays critical roles in synaptic formation and degeneration, its phosphorylation targets were reexamined in search of candidates with direct global impacts on synaptic protein dynamics, and the associated regulatory network was also analyzed. METHODS: Quantitative phosphoproteomics and bioinformatics analyses were performed to identify top-ranked candidates. A series of biochemical assays was used to investigate the associated regulatory signaling networks. Histological, electrochemical, and behavioral assays were performed in conditional knockout, small hairpin RNA-mediated knockdown, and AD-related mice models to evaluate the relevance of CDK5 to synaptic homeostasis and functions. RESULTS: Among candidates with known implications in synaptic modulations, BAG3 ranked the highest. CDK5-mediated phosphorylation on S297/S291 (mouse/human) destabilized BAG3. Loss of BAG3 unleashed the selective protein degradative function of the HSP70 machinery. In neurons, this resulted in enhanced degradation of a number of glutamatergic synaptic proteins. Conditional neuronal knockout of Bag3 in vivo led to impairment of learning and memory functions. In human AD and related mouse models, aberrant CDK5-mediated loss of BAG3 yielded similar effects on synaptic homeostasis. Detrimental effects of BAG3 loss on learning and memory functions were confirmed in these mice, and such effects were reversed by ectopic BAG3 reexpression. CONCLUSIONS: Our results highlight that the neuronal CDK5-BAG3-HSP70 signaling axis plays a critical role in modulating synaptic homeostasis. Dysregulation of the signaling pathway directly contributes to synaptic dysfunction and AD pathogenesis.

Selective Insertion of Alkynes into C-C σ Bonds of Indolin-2-ones: Transition-Metal-Free Ring Expansion Reactions to Seven-Membered-Ring Benzolactams or Chromone Derivatives.

An unprecedented ring expansion reaction of indolin-2-ones with alkynyl ketones under transition-metal-free conditions has been developed. Base-promoted selective cleavage of a C-C σ bond of the amide is the key in this process, which provides a straightforward and efficient way to synthesize seven-membered-ring benzolactams or chromone derivatives. The significant advantages of this method include readily accessible starting materials, wide scope and functional group tolerance, and high atom economy.

Controllable synthesis of pyrido[2,3-b]indol-4-ones or indolo[3,2-b]quinolines via formal intramolecular C(sp2)-H functionalization.

A novel Fe-catalyzed protocol for the controllable synthesis of pyrido[2,3-b]indol-4-ones or indolo[3,2-b]quinolines has been developed by using indole-2-carboxylic derivatives as starting materials. Indole-2-carboxenamines were transformed into pyrido[2,3-b]indol-4-ones through intramolecular N-H/C-H coupling, in which a carbonyl 1,2-migration was involved. Whereas, when indole-2-carboxarylamines were employed, indolo[3,2-b]quinolones were produced through direct N-H/C-H coupling. The desired products were obtained under mild reaction conditions in moderate to good yields with wide substrate scope. The natural product quindolinone was conveniently prepared by this reaction.

Understanding Mechanism of Adsorption in the Decolorization of Aqueous Methyl Violet (6B) Solution by Okra Polysaccharides: Experiment and Theory.

Optimal conditions for ultrasonic-assisted extraction of polysaccharide from Chinese okra were found using response surface methodology. The okra polysaccharide (OPS) was used for the adsorption of methyl violet 6B (MV). Conditions for maximal adsorption efficiency of MV were established. The mechanism of MV adsorption was investigated by the characterization and physicochemical analysis of OPS before and after the adsorption of MV. Both infrared (IR) analysis and molecular dynamics (MD) simulation suggest that MV adsorption by OPS was an electrostatic interaction between MV and oxygen-containing groups of OPS. Further, the results of first-principles calculation were in agreement with IR spectroscopy measurements and MD simulation, which were all consistent with the suggested adsorption mechanism. Optimization of okra extraction conditions, maximized efficiency of MV adsorption by OPS, and the understanding of the adsorption mechanism are the highlights of this work, providing a reference for promising applications of OPS in the treatment of wastewater in textile, paper, and other industries.

Synthesis of cyano-substituted carbazoles via successive C-C/C-H cleavage.

A highly efficient protocol for the synthesis of polysubstituted cyano carbazoles has been developed. This process is realized through the tandem reactions of Cs2CO3 promoted C-C σ-bond activation of α-cyano ketones followed by Fe-catalyzed selective C-H and/or C-C bond activations. Two of the sp2 C-H bonds are functionalized, and two of the C-C σ-bonds are cleaved involving a 1,2-acyl migration during the reaction process.

Cobalt-catalyzed regioselective syntheses of indeno[2,1-c]pyridines from nitriles and diynes bearing propargyl fragments.

A highly efficient CoI2/o-phenanthroline catalyzed cycloaddition reaction of diynes bearing TBS protected propargylic alcohol fragments with nitriles has been developed. This methodology offers regioselective access, with good functional group tolerance, to various indeno[2,1-c]pyridine derivatives in moderate to excellent yields. It was found that o-phenanthroline as a bidentate nitrogen ligand showed high efficacy in this cycloaddition reaction.

Sequential C-C σ-Bond Cleavage/(sp2) C-O Bond Formation via C-H Functionalization toward Pyranoindolones Fused with Medium-Sized Rings.

An atom-economical procedure for the synthesis of pyranoindolones fused with 7- or 8-membered rings has been developed. This process is realized through the sequential Cs2CO3-promoted C-C σ-bond cleavage of cyclic ketoesters and a ZnI2-mediated selective C-H/O-H coupling reaction. Two of the C-C σ-bonds are cleaved and one of the sp2 C-H bonds is functionalized during the reaction process. Easily accessible starting materials and broad substrate scope make this protocol attractive in synthetic organic chemistry.

Palladium-Catalyzed Dual C(sp2)-H Functionalization of Indole-2-carboxamides Involving a 1,2-Acyl Migration: A Synthesis of Indolo[3,2- c]quinolinones.

A novel Pd/Cu catalytic system to construct indolo[3,2- c]quinolinones has been developed starting from indole-2-carboxamides. Substrates were transformed into indolo[3,2- c]quinolinones through dual C(sp2)-H functionalization of the indole moiety and arene, in which a carbonyl 1,2-migration was involved. The corresponding products were obtained in moderate to excellent yields with a wide substrate scope.