An Ultrasensitive Biosensor for Probing Subcellular Distribution and Mitochondrial Transport of l-2-Hydroxyglutarate.

l-2-Hydroxyglutarate (l-2-HG) is a functionally compartmentalized metabolite involved in various physiological processes. However, its subcellular distribution and mitochondrial transport remain unclear owing to technical limitations. In the present study, an ultrasensitive l-2-HG biosensor, sfLHGFRH, composed of circularly permuted yellow fluorescent protein and l-2-HG-specific transcriptional regulator, is developed. The ability of sfLHGFRH to be used for analyzing l-2-HG metabolism is first determined in human embryonic kidney cells (HEK293FT) and macrophages. Then, the subcellular distribution of l-2-HG in HEK293FT cells and the lower abundance of mitochondrial l-2-HG are identified by the sfLHGFRH-supported spatiotemporal l-2-HG monitoring. Finally, the role of the l-glutamate transporter SLC1A1 in mitochondrial l-2-HG uptake is elucidated using sfLHGFRH. Based on the design of sfLHGFRH, another highly sensitive biosensor with a low limit of detection, sfLHGFRL, is developed for the point-of-care diagnosis of l-2-HG-related diseases. The accumulation of l-2-HG in the urine of patients with kidney cancer is determined using the sfLHGFRL biosensor.

A novel variation in DEPDC5 causing familial focal epilepsy with variable foci.

Background: Disheveled, EGL-10, and pleckstrin (DEP) domain-containing protein 5 (DEPDC5) is a component of GTPase-activating protein (GAP) activity toward the RAG complex 1 (GATOR1) protein, which is an inhibitor of the amino acid-sensing branch of the mammalian target of rapamycin complex 1 (mTORC1) pathway. GATOR1 complex variations were reported to correlate with familial focal epilepsy with variable foci (FFEVF). With the wide application of whole exome sequencing (WES), more and more variations in DEPDC5 were uncovered in FFEVF families. Methods: A family with a proband diagnosed with familial focal epilepsy with variable foci (FFEVF) was involved in this study. Whole exome sequencing (WES) was performed in the proband, and Sanger sequencing was used to confirm the variation carrying status of the family members. Mini-gene splicing assay was performed to validate the effect on the alternative splicing of the variation. Results: A novel variant, c.1217 + 2T>A, in DEPDC5 was identified by WES in the proband. This splicing variant that occurred at the 5' end of intron 17 was confirmed by mini-gene splicing assays, which impacted alternative splicing and led to the inclusion of an intron fragment. The analysis of the transcribed mRNA sequence indicates that the translation of the protein is terminated prematurely, which is very likely to result in the loss of function of the protein and lead to the occurrence of FFEVF. Conclusion: The results suggest that c.1217 + 2T>A variations in DEPDC5 might be the genetic etiology for FFEVF in this pedigree. This finding expands the genotype spectrum of FFEVF and provides new etiological information for FFEVF.

Precise treatment of pelvic fracture urethral injury associated with urethrorectal fistula.

OBJECTIVE: To evaluate the effect of a new strategy of transperineal anastomotic urethroplasty (TAU) with proximal transection in treating pelvic fracture urethral injury (PFUI) associated with urethrorectal fistula (URF). PATIENTS AND METHODS: A retrospective review of all patients treated by TAU with proximal transection and fistula repair for PFUI associated with URF was performed between August 2013 and July 2022. Information on demographics, peri-operative variables, and postoperative follow-up outcomes was collected. Successful surgery was defined as restoration of a uniform urethral calibre using flexible cystoscopy (third postoperative month) without strictures or leakage, with no further interventions required. Functional outcomes, including erectile function (assessed using the five-item International Index of Erectile Function) and urinary continence, were assessed. RESULTS: Forty patients diagnosed with PFUI associated with URF and treated by TAU with proximal transection and rectal fistula repair were enrolled. Six patients (15.0%) had a history of failed urethral reconstruction. The mean stenosis length and fistula diameter were 2.9 cm and 1.2 cm, respectively. All patients underwent faecal diversion before urethroplasty. After a median (range) follow-up of 45 (3-115) months, the final success rate was 90.0% (36/40). Postoperative complications included haematoma in three patients, epididymo-orchitis in three, wound infection in one, wound bleeding in one, delayed wound healing in three, and wound numbness in three. The overall incidence of postoperative erectile dysfunction reached 75.0%, with a median (range) score of 9 (0-19). Normal continence was achieved in 31 patients (77.5%). Occasional incontinence without the need for urinal pads occurred in eight patients, whereas one patient required urinal pads. CONCLUSIONS: Transperineal anastomotic urethroplasty with proximal transection is a precise and effective surgical strategy for treating PFUI associated with URF. This strategy ensures a high success rate and improves surgical efficiency.

Orthogonality of non-coaxial Laguerre-Gaussian beams.

The conventional orthogonality of Laguerre-Gaussian (LG) beams is widely recognized and serves as the fundamental basis for mode division multiplexing utilizing LG beams as channels. However, this conventional orthogonality holds true only when two LG beams share the same axis. Whether non-coaxial LG beams, with parallel axes separated by a distance, are orthogonal to each other remains an open question. To address this issue and reveal the orthogonality of non-coaxial LG beams, we analytically derive the projection of one LG beam onto another. This projection exhibits a helical phase and a rotationally symmetric amplitude with dark rings present. These dark rings indicate that two non-coaxial LG beams are indeed orthogonal to each other when their axes are appropriately separated. Furthermore, any pair of non-coaxial LG beams within a set also exhibit orthogonality when their axes are fixed at a certain separation distance. This novel form of orthogonality predicts an innovative compound multiplexing technique that simultaneously combines mode and space, enabling more efficient packing of multiple LG beams within a single transmission aperture while maintaining parallel and closely spaced beam axes.

The value of audiovisual sexual stimulation with virtual reality in diagnosing erectile dysfunction.

Background: The traditional audiovisual sexual stimulation (AVSS) test may experience limitations including low erectile response rate and lack of unified diagnostic criteria. Aim: We aimed to explore the clinical value of AVSS with virtual reality (VR-AVSS) test in assessing erectile function and diagnosing erectile dysfunction (ED). Methods: Participants 18 to 60 years of age were screened for analysis in 3 clinical centers from June 2020 to March 2022. Demographic data, 5-item International Index of Erectile Function (IIEF-5), erectile hardness score (EHS), and self-reported symptom questions were collected. The ED patients and control patients were confirmed according to the IIEF-5 and EHS. All subjects watched a 60-minute erotic video by VR device during RigiScan recording. The parameters including tip average rigidity, tip effective erectile duration (duration of rigidity ≥60%, tip effective erectile duration), base average rigidity, and base effective erectile duration were evaluated. Outcomes: The main outcome of interest was the application of VR immersion technology to improve the traditional AVSS test. Results: A total of 301 ED cases and 100 eligible control patients were included for final analysis. Compared with control patients, ED cases had significantly lower IIEF-5 scores, EHS, positive response rate, and erectile rigidity and duration. The positive response rate of ED and control patients were 75.5% and 90.9%, respectively. The cutoff points of tip average rigidity, tip effective erectile duration, base average rigidity, and base effective erectile duration were 40.5% (sensitivity: 77.6%, specificity: 70.2%; P < .001), 4.75 minutes (sensitivity: 75.9%, specificity: 75.4%; P < .001), 48.5% (sensitivity: 77.6%, specificity: 75.1%; P < .001), and 7.75 minutes (sensitivity: 79.3%, specificity: 75.7%; P < .001). Clinical Implications: The technological superiority of VR will enable the VR-AVSS immersion test to be a more accurate detection than traditional AVSS modes. Strengths and Limitations: Our study applied VR immersion technology to establish the standard operation procedure for the AVSS test, which could effectively reduce the interference of adverse factors and minimize the detecting errors. However, the test data only included positive response subjects, so the true erectile status of men with a negative response to the AVSS test cannot be obtained. Conclusions: The VR-AVSS test can effectively improve the diagnostic accuracy of ED. The average rigidity and effective erectile duration were the optimal diagnostic parameters for excluding ED.

Discovery of novel peptide-dehydroepiandrosterone hybrids inducing endoplasmic reticulum stress with effective in vitro and in vivo anti-melanoma activities.

Steroid hybrids have emerged as a type of advantageous compound as they could offer improved pharmacological and pharmaceutical properties. Here, we report a series of novel peptide-dehydroepiandrosterone hybrids, which would effectively induce endoplasmic reticulum stress (ERS) and lead to apoptosis with outstanding in vitro and in vivo anti-melanoma effects. The lead compound IId among various steroids conjugated with peptides and pyridines showed effective in vivo activity in B16 xenograft mice: in medium- and high-dose treatment groups (60 and 80 mg/kg), compound IId would significantly inhibit the growth of tumours by 98%-99% compared to the control group, with the highest survival rate as well. Further mechanism studies showed that compound IId would damage the endoplasmic reticulum and upregulate the ERS markers C/EBP homologous protein (CHOP) and glucose-regulated protein 78 (GRP78), which could further regulate caspase and Bcl-2 family proteins and lead to cell apoptosis. The compound IId was also proven to be effective in inhibiting B16 cell migration and invasion.

Redo inferior pubectomy for failed anastomotic urethroplasty in pelvic fracture urethral injury.

Objectives: To assess the effect of redo inferior pubectomy on the management of complicated pelvic fracture urethral injury (PFUI) in patients with a history of failed anastomotic urethroplasty. Materials and methods: We retrospectively reviewed all patients receiving redo anastomotic urethroplasty with redo inferior pubectomy for failed PFUI between January 2010 and December 2021. Patients with incomplete data and those who were lost to follow-up were excluded. Successful urethroplasty was defined as the restoration of a uniform urethral caliber without stenosis or leakage and further intervention. Functional results, including erectile function and urinary continence, were evaluated. Descriptive statistical analyses were then performed. Results: Thirty-one patients were included in this study. Among them, concomitant urethrorectal fistula occurred in 2 patients, and concomitant enlarged bladder neck occurred in 1. The stenosis site was the bulbomembranous urethra in 2 patients and the prostatomembranous urethra in 29. The mean length of urethral stenosis in all patients was 3.1 cm (range, 2.0-5.0 cm). After a mean follow-up of 34.6 months, the final success rate was 96.8%. The incidence of erectile dysfunction reached 77.4% (24/31). Normal continence was achieved in 27 (87.1%) patients. One patient developed urinary incontinence of grade II requiring urinary pads because of an enlarged bladder neck. According to the Clavien-Dindo classification, postoperative complications of grade I occurred in 7 patients and grade II in 4. Conclusions: Repeat anastomotic urethroplasty with repeat inferior pubectomy provides reliable success rates for failed PFUI. In complicated cases, it should be known and mastered.

Efficient L-valine production using systematically metabolic engineered Klebsiella oxytoca.

L-Valine, a branched-chain amino acid with diversified applications, is biosynthesized with α-acetolactate as the key precursor. In this study, the metabolic flux in Klebsiella oxytoca PDL-K5, a Risk Group 1 organism producing 2,3-butanediol as the major fermentation product, was rearranged to L-valine production by introducing exogenous L-valine biosynthesis pathway and blocking endogenous 2,3-butanediol generation at the metabolic branch point α-acetolactate. After further enhancing L-valine efflux, strengthening pyruvate polymerization and selecting of key enzymes for L-valine synthesis, a plasmid-free K. oxytoca strain VKO-9 was obtained. Fed-batch fermentation with K. oxytoca VKO-9 in a 7.5 L fermenter generated 122 g/L L-valine with a yield of 0.587 g/g in 56 h. In addition, repeated fed-batch fermentation was conducted to prevent precipitation of L-valine due to oversaturation. The average concentration, yield, and productivity of produced L-valine in three cycles of repeated fed-batch fermentation were 81.3 g/L, 0.599 g/g, and 3.39 g/L/h, respectively.

Photoswitchable Radical State in Nanoporous Metal-Organic Framework Films with Embedded Hexaarylbiimidazole.

Photoresponsive materials enable dynamic remote control of their fundamental properties. The incorporation of photochromic molecules in nanoporous metal-organic frameworks (MOFs) provides a unique opportunity to tailor the material properties, including the interaction between the MOF host and guest molecules in the pores. Here, a MOF film of type HKUST-1 with embedded hexaarylbiimidazole (HABI), undergoing reversible light-induced reactions between the stable dimer state and the metastable radical state, is presented. The switching between the dimer and radical form is shown by infrared, UV-vis, and electron paramagnetic resonance (EPR) spectroscopy. By transient uptake experiments with ethanol and methanol as probe molecules, we show that the dimer-radical switching impacts the host-guest interaction and, in particular, modifies the uptake amount and diffusion coefficient of the guest molecules. For ethanol, the diffusion slows down by 75%. This research presents the first MOF material with photoswitchable (meta)stable dimer and radical molecules, and it contributes to the advancement of photoresponsive nanoporous materials.

Generation of a human haploid neural stem cell line for genome-wide genetic screening.

BACKGROUND: Haploid embryonic stem cells (haESCs) have been established in many species. Differentiated haploid cell line types in mammals are lacking due to spontaneous diploidization during differentiation that compromises lineage-specific screens. AIM: To derive human haploid neural stem cells (haNSCs) to carry out lineage-specific screens. METHODS: Human haNSCs were differentiated from human extended haESCs with the help of Y27632 (ROCK signaling pathway inhibitor) and a series of cytokines to reduce diploidization. Neuronal differentiation of haNSCs was performed to examine their neural differentiation potency. Global gene expression analysis was con-ducted to compare haNSCs with diploid NSCs and haESCs. Fluorescence activated cell sorting was performed to assess the diploidization rate of extended haESCs and haNSCs. Genetic manipulation and screening were utilized to evaluate the significance of human haNSCs as genetic screening tools. RESULTS: Human haESCs in extended pluripotent culture medium showed more compact and smaller colonies, a higher efficiency in neural differentiation, a higher cell survival ratio and higher stability in haploidy maintenance. These characteristics effectively facilitated the derivation of human haNSCs. These human haNSCs can be generated by differentiation and maintain haploidy and multipotency to neurons and glia in the long term in vitro. After PiggyBac transfection, there were multiple insertion sites in the human haNSCs' genome, and the insertion sites were evenly spread across all chromosomes. In addition, after the cells were treated with manganese, we were able to generate a list of manganese-induced toxicity genes, demonstrating their utility as genetic screening tools. CONCLUSION: This is the first report of a generated human haploid somatic cell line with a complete genome, proliferative ability and neural differentiation potential that provides cell resources for recessive inheritance and drug targeted screening.

Reducing the Permittivity of Polyimides for Better Use in Communication Devices.

Recent studies have shown that introducing fluorinated groups into polyimide (PI) molecules can effectively reduce the dielectric constant (Dk) and dielectric loss (Df) of PIs. In this paper, 2,2'-bis[4-(4-aminophenoxy) phenyl]-1,1',1',1',3,3',3'-hexafluoropropane (HFBAPP), 2,2'-bis(trifluoromethyl)-4,4'-diaminobenzene (TFMB), diaminobenzene ether (ODA), 1,2,4,5-Benzenetetracarboxylic anhydride (PMDA), 3,3',4,4'-diphenyltetracarboxylic anhydride (s-BPDA) and 3,3',4,4'-diphenylketontetracarboxylic anhydride (BTDA) were selected for mixed polymerization to find the relationship between the structure of PIs and dielectric properties. Firstly, different structures of fluorinated PIs were determined, and were put into simulation calculation to learn how structure factors such as fluorine content, the position of fluorine atom and the molecular structure of diamine monomer affect the dielectric properties. Secondly, experiments were carried out to characterize the properties of PI films. The observed change trends of performance were found to be consistent with the simulation results, and the possible basis of the interpretation of other performance was made from the molecular structure. Finally, the formulas with the best comprehensive performance were obtained respectively. Among them, the best dielectric properties were 14.3%TFMB/85.7%ODA//PMDA with dielectric constant of 2.12 and dielectric loss of 0.00698.

Regiodivergent catalytic asymmetric dearomative cycloaddition of bicyclic heteroaromatics.

The catalytic dearomative cycloaddition of bicyclic heteroaromatics including benzofurans and indoles provides rapid access to functionalized heterocyclic molecules. Because of the inherent stereoelectronic differences, the furan or pyrrole nucleus is more prone to dearomative cycloaddition than the benzene ring. Here, we realized a geometry-based differentiation approach for achieving C6-C7 and C7-C7a regioselectivity. The rotationally restricted σ bond at C7 position respectively placed the C6-C7 and C7-C7a sites of benzofurans or indoles in an optimal spatial orientation toward the axially chiral heterodiene, thus affording two enantioenriched polycyclic compounds from a single racemic heterobiaryl atropisomers. Calculation results of density functional theory interpreted the mechanism of this parallel kinetic resolution. The bioactivity of the dearomatized products was evaluated in cancer cell lines with certain compounds exhibiting interesting biological activities.

Substrate-Bound Diarylethene-Based Anisotropic Metal-Organic Framework Films as Photoactuators with a Directed Response.

Molecular machines and responsive materials open a plethora of new opportunities in nanotechnology. We present an oriented crystalline array of diarylethene (DAE)-based photoactuators, arranged in a way to yield an anisotropic response. The DAE units are assembled, together with a secondary linker, into a monolithic surface-mounted metal-organic framework (SURMOF) film. By Infrared (IR) and UV/Vis spectroscopy as well as by synchrotron X-ray diffraction, we show that the light-induced extension changes of the molecular DAE linkers multiply to yield mesoscopic and anisotropic length changes. Due to the special architecture and substrate-bonding of the SURMOF, these length changes are transferred to the macroscopic scale, leading to the bending of a cantilever and performing work. This research shows the potential of assembling light-powered molecules into SURMOFs to yield photoactuators with a directed response, presenting a path to advanced actuators.

Inhibition of Kv7/M Channel Currents by Fangchinoline.

INTRODUCTION: Voltage-gated Kv7/M potassium channels play an essential role in the control of membrane potential and neuronal excitability. Fangchinoline, a bisbenzylisoquinoline alkaloid, displays extensive biological activities including antitumor, anti-inflammatory, and antihypertension effects. In this study, we investigated the effects of fangchinoline on Kv7/M channels. METHODS: A perforated whole-cell patch technique was used to record Kv7 currents from HEK293 cells and M-type currents from mouse dorsal root ganglion (DRG) neurons. RESULTS: Fangchinoline inhibited Kv7.2/Kv7.3 currents in a concentration-dependent manner, with an IC50 of 9.5 ± 1.2 µM. Fangchinoline significantly inhibited Kv7.1, Kv7.2, Kv7.3, Kv7.4, and Kv7.3/Kv7.5 channels without selective effects. Furthermore, fangchinoline significantly slowed the activation of Kv7.1-Kv7.5 channels and inhibited native M-channel currents of DRG neurons. CONCLUSION: Taken together, our findings indicate that fangchinoline concentration-dependently inhibited Kv7/M channel currents.

Generalized Newton's rings with vortex beams.

The Newton's rings are interference patterns with concentric rings, and Newton's rings experiment is one of the most famous classic optics experiments. Here, we show that if we use a vortex beam, we can obtain generalized Newton's rings. Unlike traditional Newton's rings, the generalized ones are no longer concentric rings but spiral arms, and fork-shaped dislocations appear in spiral arms. More interesting, we reveal that both the number of spiral arms and the number of fork-shaped dislocations are equal to the value of topological charge of incident vortex beams. Our theoretical results are demonstrated experimentally. This novel interference pattern can be used for measuring the topological charge of vortex beams.

Glycated serum albumin decreases connexin 43 phosphorylation in the corpus cavernosum.

Background: Glycated serum albumin (GSA) is an early glycosylation product that participates in diabetic vascular complications. This study examined the role of GSA in early damage to the corpus cavernosum and the involved mechanisms. Methods: Nine 8-week-old male Sprague-Dawley (SD) rats (250-300 g) were divided into the control (saline vehicle, n=3) and GSA (200 µg/kg, n=6) groups. The corpus cavernosum tissues were harvested. Phosphorylated and non-phosphorylated connexin 43 (Cx43), endothelial nitric oxide synthase (eNOS), phosphatidylinositol 3-kinase (PI3K), and serine-threonine kinase (Akt) were tested by immunohistochemistry and western blotting. Human umbilical vein endothelial cells (HUVECs) overexpressing Cx43 were used to analyze the Cx43 phosphorylation sites (S368, S262, Y265, S255, and S279/282) using western blotting. Results: The expression of phosphorylated Cx43 in the penis was significantly lower in GSA-treated rats than in controls. The expression levels of p-Cx43, p-eNOS, p-PI3K, and p-Akt were significantly decreased in HUVECs exposed to GSA in dose- and time-dependent manners. The most significant impact on all four proteins was observed with 1 µg/mL of GSA for 12 h. Phosphorylation at the S368, S262, Y265, S255, and S279/282 sites of Cx43 was downregulated by GSA, and S368 was the most significantly suppressed phosphorylation site compared with the other sites. Conclusions: GSA decreases the expression of p-Cx43 in the corpus cavernosum of rats. This effect might be also related to the decreased phosphorylation of p-eNOS, p-PI3K, and p-Akt, as well as by the downregulation of phosphorylation at the S368 site.

Synthesis and Characterization of Polyimides with Naphthalene Ring Structure Introduced in the Main Chain.

In this paper, a new aromatic diamine monomer 4,4'-(2,6-naphthalenediyl)bis[benzenamine]) (NADA) was synthesized and a series of modified PI films containing naphthalene ring structure obtained by controlling the molar ratio of NADA monomer, ternary polymerization with 4,4'-oxydianiline (ODA), and pyromellitic dianhydride (PMDA). The effects of the introduction of the naphthalene ring on the free volume and various properties of PI were investigated by molecular dynamic simulations. The results show that the comprehensive properties of the modified films are all improved to some extent, with 5% thermal weight loss temperature (Td5%) of 569 °C, glass transition temperature (Tg) of 381 °C, tensile strength of 96.41 MPa, and modulus of elasticity of 2.45 GPa. Dielectric property test results show that the dielectric constant (Dk) of the film at 1 MHz is reduced from 3.21 to 2.82 and dielectric loss (Df) reduced from 0.0091 to 0.0065. It is noteworthy that the PI-1 dielectric constant is reduced from 3.26 to 3.01 at 10 GHz with only 5% NADA doping, which is expected to yield the best ratio and provide the possibility of industrial production.

Mitochondrial regulatory mechanisms in spinal cord injury: A narrative review.

Spinal cord injury is a severe central nervous system injury that results in the permanent loss of motor, sensory, and autonomic functions below the level of injury with limited recovery. The pathological process of spinal cord injury includes primary and secondary injuries, characterized by a progressive cascade. Secondary injury impairs the ability of the mitochondria to maintain homeostasis and leads to calcium overload, excitotoxicity, and oxidative stress, further exacerbating the injury. The defective mitochondrial function observed in these pathologies accelerates neuronal cell death and inhibits regeneration. Treatment of spinal cord injury by preserving mitochondrial biological function is a promising, although still underexplored, therapeutic strategy. This review aimed to explore mitochondrial-based therapeutic advances after spinal cord injury. Specifically, it briefly describes the characteristics of spinal cord injury. It then broadly discusses the drugs used to protect the mitochondria (e.g., cyclosporine A, acetyl-L-carnitine, and alpha-tocopherol), phenomena associated with mitochondrial damage processes (e.g., mitophagy, ferroptosis, and cuproptosis), mitochondrial transplantation for nerve cell regeneration, and innovative mitochondrial combined protection therapy.

Metabolic Mechanism and Physiological Role of Glycerol 3-Phosphate in Pseudomonas aeruginosa PAO1.

Pseudomonas aeruginosa is an important opportunistic pathogen that is lethal to cystic fibrosis (CF) patients. Glycerol generated during the degradation of phosphatidylcholine, the major lung surfactant in CF patients, could be utilized by P. aeruginosa. Previous studies have indicated that metabolism of glycerol by this bacterium contributes to its adaptation to and persistence in the CF lung environment. Here, we investigated the metabolic mechanisms of glycerol and its important metabolic intermediate glycerol 3-phosphate (G3P) in P. aeruginosa PAO1. We found that G3P homeostasis plays an important role in the growth and virulence factor production of P. aeruginosa PAO1. The G3P accumulation caused by the mutation of G3P dehydrogenase (GlpD) and exogenous glycerol led to impaired growth and reductions in pyocyanin synthesis, motilities, tolerance to oxidative stress, and resistance to kanamycin. Transcriptomic analysis indicates that the growth retardation caused by G3P stress is associated with reduced glycolysis and adenosine triphosphate (ATP) generation. Furthermore, two haloacid dehalogenase-like phosphatases (PA0562 and PA3172) that play roles in the dephosphorylation of G3P in strain PAO1 were identified, and their enzymatic properties were characterized. Our findings reveal the importance of G3P homeostasis and indicate that GlpD, the key enzyme for G3P catabolism, is a potential therapeutic target for the prevention and treatment of infections by this pathogen. IMPORTANCE In view of the intrinsic resistance of Pseudomonas aeruginosa to antibiotics and its potential to acquire resistance to current antibiotics, there is an urgent need to develop novel therapeutic options for the treatment of infections caused by this bacterium. Bacterial metabolic pathways have recently become a focus of interest as potential targets for the development of new antibiotics. In this study, we describe the mechanism of glycerol utilization in P. aeruginosa PAO1, which is an available carbon source in the lung environment. Our results reveal that the homeostasis of glycerol 3-phosphate (G3P), a pivotal intermediate in glycerol catabolism, is important for the growth and virulence factor production of P. aeruginosa PAO1. The mutation of G3P dehydrogenase (GlpD) and the addition of glycerol were found to reduce the tolerance of P. aeruginosa PAO1 to oxidative stress and to kanamycin. The findings highlight the importance of G3P homeostasis and suggest that GlpD is a potential drug target for the treatment of P. aeruginosa infections.

Enantioselective Synthesis of Atropisomers via Vinylidene ortho-Quinone Methides (VQMs).

Atropisomers, arising from conformational restriction, are inherently chiral due to the intersecting dissymmetric planes. Since there are numerous applications of enantiopure atropisomers in catalyst design, drug discovery, and material science, the asymmetric preparation of these highly prized molecules has become a flourishing field in synthetic chemistry. A number of catalysts, synthetic procedures, and novel concepts have been developed for the manufacture of the atropisomeric molecules. However, due to the intrinsic properties of different types of atropisomers featuring biaryl, hetero-biaryl, or non-biaryl architectures, only very few methods pass the rigorous inspection and are considered generally applicable. The development of a broadly applicable synthetic strategy for various atropisomers is a challenge. In this Account, we summarize our recent studies on the enantioselective synthesis of atropisomers using the vinylidene ortho-quinone methides (VQMs) as pluripotent intermediates.The most appealing features of VQMs are the disturbed aromaticity and axial chirality of the allene fragment. At the outset, the applications of VQMs in organic synthesis have been neglected due to their principal liabilities: ephemeral nature, extraordinary reactivity, and multireaction sites. The domestication of this transient intermediate was demonstrated by in situ catalytic asymmetric generation of VQMs, and the reactivity and selectivity were fully explored by judiciously modifying precursors and tuning catalytic systems. A variety of axially chiral heterocycles were achieved through five-, six-, seven- and nine-membered ring formation of VQM intermediates with different kinds of branched nucleophilic functional groups. The axially chiral C-N axis could be constructed from VQM intermediates via N-annulation or desymmetrization of preformed C-N scaffolds. We take advantage of the high electrophilicity of VQMs toward a series of sulfur and carbon based nucleophiles leading to atropisomeric vinyl arenes. Furthermore, chiral helical compounds were realized by cycloaddition or consecutive annulation of VQM intermediates. These achievements demonstrated that the VQMs could work as a nuclear parent for the collective synthesis of distinct and complex optically active atropisomers. Recently, we have realized the isolation and structural characterization of the elusive VQMs, which were questioned as putative intermediates for decades. The successful isolation of VQMs provided direct evidence for their existence and an unprecedented opportunity to directly investigate their reactivity. The good thermal stability and reserved reactivity of the isolated VQMs demonstrated their great potential as synthetic reagents and expanded the border of VQM chemistry.