Chronic cerebral hypoperfusion leads to white matter injury (WMI), which plays a significant role in contributing to vascular cognitive impairment. While 13-docosenamide is a type of fatty acid amide, it remains unclear whether it has therapeutic effects on chronic cerebral hypoperfusion. In this study, we conducted bilateral common carotid artery stenosis (BCAS) surgery to simulate chronic cerebral hypoperfusion-induced WMI and cognitive impairment. Our findings showed that 13-docosenamide alleviates WMI and cognitive impairment in BCAS mice. Mechanistically, 13-docosenamide specifically binds to cannabinoid receptor 1 (CNR1) in oligodendrocyte precursor cells (OPCs). This interaction results in an upregulation of ubiquitin-specific peptidase 33 (USP33)-mediated CNR1 deubiquitination, subsequently increasing CNR1 protein expression, activating the phosphorylation of the AKT/mTOR pathway, and promoting the differentiation of OPCs. In conclusion, our study suggests that 13-docosenamide can ameliorate chronic cerebral hypoperfusion-induced WMI and cognitive impairment by enhancing OPC differentiation and could serve as a potential therapeutic drug.
Animals ; Oligodendrocyte Precursor Cells/metabolism* ; Mice ; Cell Differentiation/drug effects* ; Male ; Receptor, Cannabinoid, CB1/metabolism* ; Mice, Inbred C57BL ; Ubiquitin Thiolesterase/metabolism* ; Ubiquitination/drug effects* ; Carotid Stenosis/complications* ; Cognitive Dysfunction/drug therapy*

Microglia, the resident immune cells in the central nervous system (CNS), rapidly transition from a resting to an active state in the acute phase of ischemic brain injury. This active state mediates a pro-inflammatory response that can exacerbate the injury. Targeting the pro-inflammatory response of microglia in the semi-dark band during this acute phase may effectively reduce brain injury. Shionone (SH), an active ingredient extracted from the dried roots and rhizomes of the genus Aster (Asteraceae), has been reported to regulate the inflammatory response of macrophages in sepsis-induced acute lung injury. However, its function in post-stroke neuroinflammation, particularly microglia-mediated neuroinflammation, remains uninvestigated. This study found that SH significantly inhibited lipopolysaccharide (LPS)-induced elevation of inflammatory cytokines, including interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), and inducible nitric oxide synthase (iNOS), in microglia in vitro. Furthermore, the results demonstrated that SH alleviated infarct volume and improved behavioral performance in middle cerebral artery occlusion (MCAO) mice, which may be attributed to the inhibition of the microglial inflammatory response induced by SH treatment. Mechanistically, SH potently inhibited the phosphorylation of serine-threonine protein kinase B (AKT), mammalian target of rapamycin (mTOR), and signal transducer and activator of transcription 3 (STAT3). These findings suggest that SH may be a potential therapeutic agent for relieving ischemic stroke (IS) by alleviating microglia-associated neuroinflammation.
Animals ; Microglia/immunology* ; Mice ; Male ; Mice, Inbred C57BL ; Brain Ischemia/immunology* ; Neuroinflammatory Diseases/drug therapy* ; Neuroprotective Agents/administration & dosage* ; Interleukin-1beta/genetics* ; STAT3 Transcription Factor/genetics* ; TOR Serine-Threonine Kinases/genetics* ; Tumor Necrosis Factor-alpha/genetics* ; Proto-Oncogene Proteins c-akt/immunology* ; Nitric Oxide Synthase Type II/genetics* ; Lipopolysaccharides

Chronic cerebral hypoperfusion leads to white matter injury (WMI), which subsequently causes neurodegeneration and even cognitive impairment. However, due to the lack of treatment specifically for WMI, novel recognized and effective therapeutic strategies are urgently needed. In this study, we found that honokiol and magnolol, two compounds derived from Magnolia officinalis, significantly facilitated the differentiation of primary oligodendrocyte precursor cells (OPCs) into mature oligodendrocytes, with a more prominent effect of the former compound. Moreover, our results demonstrated that honokiol treatment improved myelin injury, induced mature oligodendrocyte protein expression, attenuated cognitive decline, promoted oligodendrocyte regeneration, and inhibited astrocytic activation in the bilateral carotid artery stenosis model. Mechanistically, honokiol increased the phosphorylation of serine/threonine kinase (Akt) and mammalian target of rapamycin (mTOR) by activating cannabinoid receptor 1 during OPC differentiation. Collectively, our study indicates that honokiol might serve as a potential treatment for WMI in chronic cerebral ischemia.
Magnolia ; White Matter ; Brain Ischemia/metabolism* ; Oligodendroglia/metabolism*

Abnormal synchronous neuronal activity has been widely detected by brain imaging of autistic patients, but its underlying neural mechanism remains unclear. Compared with wild-type mice, our in vivo two-photon imaging showed that transgenic (Tg1) mice over-expressing human autism risk gene MeCP2 exhibited higher neuronal synchrony in the young but lower synchrony in the adult stage. Whole-cell recording of neuronal pairs in brain slices revealed that higher neuronal synchrony in young postnatal Tg1 mice was attributed mainly to more prevalent giant slow inward currents (SICs). Both in vivo and slice imaging further demonstrated more dynamic activity and higher synchrony in astrocytes from young Tg1 mice. Blocking astrocytic gap junctions markedly decreased the generation of SICs and overall cell synchrony in the Tg1 brain. Furthermore, the expression level of Cx43 protein and the coupling efficiency of astrocyte gap junctions remained unchanged in Tg1 mice. Thus, astrocytic gap junctions facilitate but do not act as a direct trigger for the abnormal neuronal synchrony in young Tg1 mice, revealing the potential role of the astrocyte network in the pathogenesis of MeCP2 duplication syndrome.

The nucleocapsid (N) protein of SARS-CoV-2 has been reported to have a high ability of liquid-liquid phase separation, which enables its incorporation into stress granules (SGs) of host cells. However, whether SG invasion by N protein occurs in the scenario of SARS-CoV-2 infection is unknow, neither do we know its consequence. Here, we used SARS-CoV-2 to infect mammalian cells and observed the incorporation of N protein into SGs, which resulted in markedly impaired self-disassembly but stimulated cell cellular clearance of SGs. NMR experiments further showed that N protein binds to the SG-related amyloid proteins via non-specific transient interactions, which not only expedites the phase transition of these proteins to aberrant amyloid aggregation in vitro, but also promotes the aggregation of FUS with ALS-associated P525L mutation in cells. In addition, we found that ACE2 is not necessary for the infection of SARS-CoV-2 to mammalian cells. Our work indicates that SARS-CoV-2 infection can impair the disassembly of host SGs and promote the aggregation of SG-related amyloid proteins, which may lead to an increased risk of neurodegeneration.
Amyloidogenic Proteins/metabolism* ; Amyotrophic Lateral Sclerosis/genetics* ; Animals ; COVID-19 ; Cytoplasmic Granules/metabolism* ; Mammals ; SARS-CoV-2 ; Stress Granules

Objective:To explore the differential diagnosis methods between nonclassical 21-hydroxylase deficiency(NC21-OHD) and polycystic ovary syndrome(PCOS).Methods:The clinical data of 31 women with NC21-OHD were compared with those of 29 women with PCOS.Results:Women with NC21-OHD showed a higher prevalence of adrenal hyperplasia and lower likelihood of polycystic ovary(PCO) than those with PCOS( P<0.05), with lower height( P<0.05). The levels of adrenocorticotropic hormone (ACTH), 17-hydroxyprogesterone(17-OHP), androstenedione(AD), total testosterone(TT), and progesterone were higher in women with NC21-OHD compared with those with PCOS( P<0.05). Women with PCOS had higher levels of luteinizing hormone(LH) and higher ratio of LH to follicle stimulating hormone(FSH) than those with NC21-OHD( P<0.05). The best two identification indexes for the two diseases were 17-OHP and progesterone, with the optimal cut-off points 3.34 ng/ml(sensitivity 89.7%, specificity 93.1%) and 0.64 ng/ml(sensitivity 90.0%, specificity 75.9%), respectively. During the 1-day mid-dose dexamethasone suppression test(DST), women with NC21-OHD had higher inhibition rates of 17-OHP, progesterone, AD, and TT( P<0.01) than those with PCOS. Their optimal cut-off values of suppression rates were 73.5%(sensitivity 95.2%, specificity 100.0%), 55.5%(sensitivity 100%, specificity 88.9%), 61.4%(sensitivity 84.2%, specificity 100.0%), 68.3%(sensitivity 65.0%, specificity 100.0%), respectively. Conclusion:The clinical manifestations of women with NC21-OHD are similar to those with PCOS. 17-OHP is the best differential indicator and the 1-day mid-dose DST plays an important role in the identification of the two diseases. Genetic analysis is the gold standard for distinguishing the two diseases.

The fcl gene encodes GDP-fucose synthase, which catalyzes two-step differential isomerase and reductase reactions in the synthesis of GDP-L-fucose from GDP-D-mannose. It also participates in the biosynthesis of amino sugar and ribose sugar, and is one of the key enzymes to regulate the metabolism of sugar and nucleotides in organisms. The presence of fcl gene in Saccharopolyspora pogona was found through sequencing result of genome. The mutant S. pogona-fcl and S. pogona-Δfcl were constructed by gene engineering technology. The results showed that the gene had an effects on growth and development, protein expression and transcriptional level, insecticidal activity, and biosynthesis of butenyl-spinosyn of Saccharopolyspora pogona. The results of HPLC analysis showed that the yield of butenyl-spinosyn in S. pogona-Δfcl was 130% compared with that in S. pogona, which reduced by 25% in S. pogona-fcl. The results of determination of insecticidal activity showed that S. pogona-Δfcl had a stronger insecticidal activity against Helicoverpa armigera than that of S. pogona, while the S. pogona-fcl had a lower insecticidal activity against Helicoverpa armigera compared with S. pogona. Scanning electron microscopy (SEM) was used to observe the morphology of the mycelia. It was found that the surface of the S. pogona-Δfcl was wrinkled, and the mycelium showed a short rod shape. There was no significant difference in mycelial morphology between S. pogona-fcl and S. pogona. Aboved all showed that deletion of fcl gene in S. pogona hindered the growth and development of mycelia, but was beneficial to increase the biosynthesis of butenyl-spinosyn and improve insecticidal activity. Whereas the fcl gene over-expression was not conducive to the biosynthesis of butenyl-spinosyn and reduced their insecticidal activity. SDS-PAGE results showed that the difference of protein expression among the three strains was most obvious at 96 hours, which was identified by real-time fluorescence quantitative polymerase chain reaction, the results showed that there were significant differences of related genes in transcriptional levels among the three strains. Based on the results of the study, a network metabolic control map was constructed to analyze the effect of fcl gene on growth and the regulation pathway of butenyl-spinosyn biosynthesis, which provided an experimental basis for revealing the regulation mechanism of butenyl-spinosyn biosynthesis and related follow-up studies.
Bacterial Proteins ; Genetic Engineering ; Insecticides ; Macrolides ; Saccharopolyspora

To study the chemical constituents from the bark of Myrica rubra, fourteen compounds were isolated from the methanolic extract using various chromatographic techniques, including silica gel, Sephadex LH-20 and preparative HPLC. Their structures were identified on the basis of chemical properties and spectroscopic data, as 3, 5-dimethoxy-4-hydroxymyricanol (1), myricanol (2), myricanone (3), myricanol 11-sulfate (4), myricitrin (5), quercetin (6), quercetin-3-rhamnoside (7), tamarixol (8), uvaol (9), ursolic acid (10), taraxerol (11), myricadiol (12), β-sitosterol (13) and β-daucosterol (14). Among them, compound 1 is a new compound, named as 3, 5-dimethoxy-4-hydroxymyricanol, compounds 8, 9 were isolated from the genus Myrica for the first time.

Proteolytic processing of viral polyproteins is indispensible for the lifecycle of coronaviruses. The main protease (M(pro)) of SARS-CoV is an attractive target for anti-SARS drug development as it is essential for the polyprotein processing. M(pro) is initially produced as part of viral polyproteins and it is matured by autocleavage. Here, we report that, with the addition of an N-terminal extension peptide, M(pro) can form a domain-swapped dimer. After complete removal of the extension peptide from the dimer, the mature M(pro) self-assembles into a novel super-active octamer (AO-M(pro)). The crystal structure of AO-M(pro) adopts a novel fold with four domain-swapped dimers packing into four active units with nearly identical conformation to that of the previously reported M(pro) active dimer, and 3D domain swapping serves as a mechanism to lock the active conformation due to entanglement of polypeptide chains. Compared with the previously well characterized form of M(pro), in equilibrium between inactive monomer and active dimer, the stable AO-M(pro) exhibits much higher proteolytic activity at low concentration. As all eight active sites are bound with inhibitors, the polyvalent nature of the interaction between AO-M(pro) and its polyprotein substrates with multiple cleavage sites, would make AO-M(pro) functionally much more superior than the M(pro) active dimer for polyprotein processing. Thus, during the initial period of SARS-CoV infection, this novel active form AOM(pro) should play a major role in cleaving polyproteins as the protein level is extremely low. The discovery of AOM(pro) provides new insights about the functional mechanism of M(pro) and its maturation process.
Coronavirus ; metabolism ; Cysteine Endopeptidases ; Endopeptidases ; metabolism ; Humans ; Peptides ; chemistry ; metabolism ; Polyproteins ; chemistry ; metabolism ; Protein Binding ; SARS Virus ; chemistry ; metabolism ; Viral Proteins