VHL suppresses UBE3B-mediated breast tumor growth and metastasis.

Protein homeostasis is predominantly governed through post-translational modification (PTM). UBE3B, identified as an oncoprotein, exhibits elevated protein levels in breast cancer. However, the impact of PTM on UBE3B remains unexplored. In this study, we show that VHL is a bona fide E3 ligase for UBE3B. Mechanistically, VHL directly binds to UBE3B, facilitating its lysine 48 (K48)-linked polyubiquitination at K286 and K427 in a prolyl hydroxylase (PHD)-independent manner. Consequently, this promotes the proteasomal degradation of UBE3B. The K286/427R mutation of UBE3B dramatically abolishes the inhibitory effect of VHL on breast tumor growth and lung metastasis. Additionally, the protein levels of UBE3B and VHL exhibit a negative correlation in breast cancer tissues. These findings delineate an important layer of UBE3B regulation by VHL.

Periodontitis exacerbates pulmonary hypertension by promoting IFNγ+ T cell infiltration in mice.

Uncovering the risk factors of pulmonary hypertension and its mechanisms is crucial for the prevention and treatment of the disease. In the current study, we showed that experimental periodontitis, which was established by ligation of molars followed by orally smearing subgingival plaques from patients with periodontitis, exacerbated hypoxia-induced pulmonary hypertension in mice. Mechanistically, periodontitis dysregulated the pulmonary microbiota by promoting ectopic colonization and enrichment of oral bacteria in the lungs, contributing to pulmonary infiltration of interferon gamma positive (IFNγ+) T cells and aggravating the progression of pulmonary hypertension. In addition, we identified Prevotella zoogleoformans as the critical periodontitis-associated bacterium driving the exacerbation of pulmonary hypertension by periodontitis, and the exacerbation was potently ameliorated by both cervical lymph node excision and IFNγ neutralizing antibodies. Our study suggests a proof of concept that the combined prevention and treatment of periodontitis and pulmonary hypertension are necessary.

Locational and functional characterization of PI4KB in the mouse embryo.

Phosphatidylinositol 4-kinase beta (PI4KB) is a member of the PI4K family, which is mainly enriched and functions in the Golgi apparatus. The kinase domain of PI4KB catalyzes the phosphorylation of phosphatidylinositol to form phosphatidylinositol 4-phosphate, a process that regulates various sub-cellular events, such as non-vesicular cholesterol and ceramide transport, protein glycosylation, and vesicle transport, as well as cytoplasmic division. In this study, a strain of PI4KB knockout mouse, immunofluorescence, reverse transcription polymerase chain reaction and microinjection were used to characterize the cytological location and biological function of PI4KB in the mouse embryos. we found that knocking down Pi4kb in mouse embryos resulted in embryonic lethality at around embryonic day (E) 7.5. Additionally, we observed dramatic fluctuations in PI4KB expression during the development of preimplantation embryos, with high expression in the 4-cell and morula stages. PI4KB colocalized with the Golgi marker protein TGN46 in the perinuclear and cytoplasmic regions in early blastomeres. Postimplantation, PI4KB was highly expressed in the epiblast of E7.5 embryos. Treatment of embryos with PI4KB inhibitors was found to inhibit the development of the morula into a blastocyst and the normal progression of cytoplasmic division during the formation of a 4-cell embryo. These findings suggest that PI4KB plays an important role in mouse embryogenesis by regulating various intracellular vital functions of embryonic cells.

The dynamic recruitment of LAB proteins senses meiotic chromosome axis differentiation in C. elegans.

During meiosis, cohesin and meiosis-specific proteins organize chromatin into an axis-loop architecture, coordinating homologous synapsis, recombination, and ordered chromosome segregation. However, how the meiotic chromosome axis is assembled and differentiated with meiotic progression remains elusive. Here, we explore the dynamic recruitment of two long arms of the bivalent proteins, LAB-1 and LAB-2, in Caenorhabditis elegans. LAB proteins directly interact with the axis core HORMA complexes and weak interactions contribute to their recruitment. LAB proteins phase separate in vitro, and this capacity is promoted by HORMA complexes. During early prophase, synapsis oppositely regulates the axis enrichment of LAB proteins. After the pachytene exit, LAB proteins switch from a reciprocal localization pattern to a colocalization pattern, and the normal dynamic pattern of LAB proteins is altered in meiotic mutants. We propose that LAB recruitment senses axis differentiation, and phase separation of meiotic structures helps subdomain establishment and accurate segregation of the chromosomes.

Oral pathogens exacerbate Parkinson's disease by promoting Th1 cell infiltration in mice.

BACKGROUND: Parkinson's disease (PD) is a common chronic neurological disorder with a high risk of disability and no cure. Periodontitis is an infectious bacterial disease occurring in periodontal supporting tissues. Studies have shown that periodontitis is closely related to PD. However, direct evidence of the effect of periodontitis on PD is lacking. Here, we demonstrated that ligature-induced periodontitis with application of subgingival plaque (LIP-SP) exacerbated motor dysfunction, microglial activation, and dopaminergic neuron loss in 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD mice. RESULTS: The 16S rRNA gene sequencing revealed that LIP-SP induced oral and gut dysbiosis. Particularly, Veillonella parvula (V. parvula) and Streptococcus mutans (S. mutans) from oral ligatures were increased in the fecal samples of MPTP + LIP-SP treated mice. We further demonstrated that V. parvula and S. mutans played crucial roles in LIP-SP mediated exacerbation of motor dysfunction and neurodegeneration in PD mice. V. parvula and S. mutans caused microglial activation in the brain, as well as T helper 1 (Th1) cells infiltration in the brain, cervical lymph nodes, ileum and colon in PD mice. Moreover, we observed a protective effect of IFNγ neutralization on dopaminergic neurons in V. parvula- and S. mutans-treated PD mice. CONCLUSIONS: Our study demonstrates that oral pathogens V. parvula and S. mutans necessitate the existence of periodontitis to exacerbate motor dysfunction and neurodegeneration in MPTP-induced PD mice. The underlying mechanisms include alterations of oral and gut microbiota, along with immune activation in both brain and peripheral regions. Video Abstract.

Brown adipocyte mineralocorticoid receptor deficiency impairs metabolic regulation in diet-induced obese mice.

Activation of brown adipose tissue (BAT) contributes to energy dissipation and metabolic health. Although mineralocorticoid receptor (MR) antagonists have been demonstrated to improve metabolism under obesity, the underlying mechanisms remain incompletely understood. We aimed to evaluate the role of BAT MR in metabolic regulation. After 8 weeks of high-fat diet (HFD) feeding, BAT MR KO (BMRKO) mice manifested significantly increased bodyweight, fat mass, serum fasting glucose, and impaired glucose homeostasis compared with littermate control (LC) mice, although insulin resistance and fasting serum insulin were not significantly changed. Metabolic cage experiments showed no change in O2 consumption, CO2 production, or energy expenditure in obese BMRKO mice. RNA sequencing analysis revealed downregulation of genes related to fatty acid metabolism in BAT of BMRKO-HFD mice compared with LC-HFD mice. Moreover, H&E and immunohistochemical staining demonstrated that BMRKO exacerbated HFD-induced macrophage infiltration and proinflammatory genes in epididymal white adipose tissue (eWAT). BMRKO-HFD mice also manifested significantly increased liver weights and hepatic lipid accumulation, an increasing trend of genes related to lipogenesis and lipid uptake, and significantly decreased genes related to lipolytic and fatty acid oxidation in the liver. Finally, the level of insulin-induced AKT phosphorylation was substantially blunted in eWAT but not liver or skeletal muscle of BMRKO-HFD mice compared with LC-HFD mice. These data suggest that BAT MR is required to maintain metabolic homeostasis, likely through its regulation of fatty acid metabolism in BAT and impacts on eWAT and liver.

Periodontitis exacerbates atherosclerosis through Fusobacterium nucleatum-promoted hepatic glycolysis and lipogenesis.

AIMS: Positive associations between periodontitis (PD) and atherosclerosis have been established, but the causality and mechanisms are not clear. We aimed to explore the causal roles of PD in atherosclerosis and dissect the underlying mechanisms. METHODS AND RESULTS: A mouse model of PD was established by ligation of molars in combination with application of subgingival plaques collected from PD patients and then combined with atherosclerosis model induced by treating atheroprone mice with a high-cholesterol diet (HCD). PD significantly aggravated atherosclerosis in HCD-fed atheroprone mice, including increased en face plaque areas in whole aortas and lesion size at aortic roots. PD also increased circulating levels of triglycerides and cholesterol, hepatic levels of cholesterol, and hepatic expression of rate-limiting enzymes for lipogenesis. Using 16S ribosomal RNA (rRNA) gene sequencing, Fusobacterium nucleatum was identified as the most enriched PD-associated pathobiont that is present in both the oral cavity and livers. Co-culture experiments demonstrated that F. nucleatum directly stimulated lipid biosynthesis in primary mouse hepatocytes. Moreover, oral inoculation of F. nucleatum markedly elevated plasma levels of triglycerides and cholesterol and promoted atherogenesis in HCD-fed ApoE-/- mice. Results of RNA-seq and Seahorse assay indicated that F. nucleatum activated glycolysis, inhibition of which by 2-deoxyglucose in turn suppressed F. nucleatum-induced lipogenesis in hepatocytes. Finally, interrogation of the molecular mechanisms revealed that F. nucleatum-induced glycolysis and lipogenesis by activating PI3K/Akt/mTOR signalling pathway in hepatocytes. CONCLUSIONS: PD exacerbates atherosclerosis and impairs lipid metabolism in mice, which may be mediated by F. nucleatum-promoted glycolysis and lipogenesis through PI3K/Akt/mTOR signalling in hepatocytes. Treatment of PD and specific targeting of F. nucleatum are promising strategies to improve therapeutic effectiveness of hyperlipidaemia and atherosclerosis.

CYLD deubiquitinates plakoglobin to promote Cx43 membrane targeting and gap junction assembly in the heart.

During heart maturation, gap junctions assemble into hemichannels and polarize to the intercalated disc at cell borders to mediate electrical impulse conduction. However, the molecular mechanism underpinning cardiac gap junction assembly remains elusive. Herein, we demonstrate an important role for the deubiquitinating enzyme cylindromatosis (CYLD) in this process. Depletion of CYLD in mice impairs the formation of cardiac gap junctions, accelerates cardiac fibrosis, and increases heart failure. Mechanistically, CYLD interacts with plakoglobin and removes lysine 63-linked polyubiquitin chains from plakoglobin. The deubiquitination of plakoglobin enhances its interaction with the desmoplakin/end-binding protein 1 complex localized at the microtubule plus end, thereby promoting microtubule-dependent transport of connexin 43 (Cx43), a key component of gap junctions, to the cell membrane. These findings establish CYLD as a critical player in regulating gap junction assembly and have important implications in heart development and diseases.

Anti-diabetic effects of linarin from Chrysanthemi Indici Flos via AMPK activation.

Objective: The purpose of this study is to investigate the anti-diabetic effects of linarin, a flavonoid extracted from Chrysanthemi Indici Flos (CIF), and its potential mechanisms. Methods: The effects of linarin on cell viability and glucose consumption in HepG2 cells were measured. Meanwhile, monosodium glutamate (MSG) mouse model was constructed to monitor the changes of insulin tolerance, glucose tolerance, triglyceride and cholesterol. The protein expression levels of p-AMPK, p-ACC, PEPCK and p-GS were detected by Western blot. Results: Linarin could increase the relative glucose consumption of HepG2 cells, improve insulin tolerance and glucose tolerance, and decrease the levels of triglyceride and cholesterol of MSG mice. Simultaneously, the expression levels of p-AMPK and p-ACC in HepG2 cells and the liver tissue of MSG mice were increased, while the expression levels of PEPCK and p-GS were decreased after treatment with linarin. Conclusion: Insulin resistance could be ameliorated by linarin in type 2 diabetes, and its mechanism may be related to AMPK signaling pathway.

Periodontitis Salivary Microbiota Aggravates Ischemic Stroke Through IL-17A.

Although epidemiological studies suggest that periodontitis is tightly associated with ischemic stroke, its impact on ischemic stroke and the underlysing mechanisms are poorly understood. Recent studies have shown that alteration in gut microbiota composition influences the outcomes of ischemic stroke. In the state of periodontitis, many oral pathogenic bacteria in the saliva are swallowed and transmitted to the gut. However, the role of periodontitis microbiota in the pathogenesis and progression of ischemic stroke is unclear. Therefore, we hypothesized that the periodontitis salivary microbiota influences the gut immune system and aggravates ischemic stroke. Mice receiving gavage of periodontitis salivary microbiota showed significantly worse stroke outcomes. And these mice also manifested more severe neuroinflammation, with higher infiltration of inflammatory cells and expression of inflammatory cytokines in the ischemic brain. More accumulation of Th17 cells and IL-17+ γδ T cells were observed in the ileum. And in Kaede transgenic mice after photoconversion. Migration of CD4+ T cells and γδ T cells from the ileum to the brain was observed after ischemic stroke in photoconverted Kaede transgenic mice. Furthermore, the worse stroke outcome was abolished in the IL-17A knockout mice. These findings suggest that periodontitis salivary microbiota increased IL-17A-producing immune cells in the gut, likely promoted the migration of these cells from the gut to the brain, and subsequently provoked neuroinflammation after ischemic stroke. These findings have revealed the role of periodontitis in ischemic stroke through the gut and provided new insights into the worse outcome of ischemic stroke coexisting with periodontitis in clinical trials.

Osteoblast MR deficiency protects against adverse ventricular remodeling after myocardial infarction.

RATIONALE: Mineralocorticoid receptor (MR) antagonists have been clinically used to treat heart failure. However, the underlying cellular and molecular mechanisms remain incompletely understood. METHODS AND RESULTS: Using osteoblast MR knockout (MRobko) mouse in combination with myocardial infarction (MI) model, we demonstrated that MR deficiency in osteoblasts significantly improved cardiac function, promoted myocardial healing, as well as attenuated cardiac hypertrophy, fibrosis and inflammatory response after MI. Gene expression profiling using RNA sequencing revealed suppressed expression of osteocalcin (OCN) in calvaria from MRobko mice compared to littermate control (MRfl/fl) mice with or without MI. Plasma levels of undercarboxylated OCN (ucOCN) were also markedly decreased in MRobko mice compared to MRfl/fl mice. Administration of ucOCN abolished the protective effects of osteoblast MR deficiency on infarcted hearts. Mechanistically, ucOCN treatment promoted proliferation and inflammatory cytokine secretion in macrophages. Spironolactone, an MR antagonist, significantly inhibited the expression and secretion of OCN in post-MI mice. More importantly, spironolactone decreased plasma levels of ucOCN and inflammatory cytokines in heart failure patients. CONCLUSIONS: MR deficiency in osteoblasts alleviates pathological ventricular remodeling after MI, likely through its regulation on OCN. Spironolactone may work through osteoblast MR/OCN axis to exert its therapeutic effects on pathological ventricular remodeling and heart failure in mice and human patients.

Macrophage Nuclear Receptor Corepressor 1 Deficiency Protects Against Ischemic Stroke in Mice.

Microglia/macrophage activation plays an essential role in Ischemic stroke (IS). Nuclear receptor corepressor 1 (NCoR1) has been identified as a vital regulator in macrophages. The present study aims to explore the functions of macrophage NCoR1 in IS. Macrophage NCoR1 knockout (MNKO) mice and littermate control mice were subjected to middle cerebral artery occlusion (MCAO). Our data showed that macrophage NCoR1 deficiency significantly reduced the infarct size and infarct volume as well as brain edema after MCAO. Additionally, MNKO induced less microglia/macrophage infiltration and activation, neuroinflammation, apoptosis of neuronal cells, and BBB disruption in brains after IS. Mechanistic studies revealed that NCoR1 interacted with LXRß in microglia and MNKO impaired the activation of the Nuclear factor-κB signaling pathway in brains after IS. Our data demonstrated that macrophage NCoR1 deficiency inhibited microglia/macrophage activation and protected against IS. Targeting NCoR1 in microglia/macrophage may be a potential approach for IS treatment.

Primary cilia in retinal pigment epithelium development and diseases.

Retinal pigment epithelium (RPE) is a highly polarized epithelial monolayer lying between the photoreceptor layer and the Bruch membrane. It is essential for vision through participating in many critical activities, including phagocytosis of photoreceptor outer segments, recycling the visual cycle-related compounds, forming a barrier to control the transport of nutrients, ions, and water, and the removal of waste. Primary cilia are conservatively present in almost all the vertebrate cells and acts as a sensory organelle to control tissue development and homeostasis maintenance. Numerous studies reveal that abnormalities in RPE lead to various retinal diseases, such as age-related macular degeneration and diabetic macular oedema, but the mechanism of primary cilia in these physiological and pathological activities remains to be elucidated. Herein, we summarize the functions of primary cilia in the RPE development and the mutations of ciliary genes identified in RPE-related diseases. By highlighting the significance of primary cilia in regulating the physiological and pathological processes of RPE, we aim to provide novel insights for the treatment of RPE-related retinal diseases.

Effects of 2,3',4,4',5-pentachlorobiphenyl exposure during pregnancy on DNA methylation in the testis of offspring in the mouse.

Polychlorinated biphenyls (PCBs) are persistent organic pollutants, and the widespread use of PCBs has had adverse effects on human and animal health. This study experiment explored the effects of 2,3',4,4',5-pentachlorobiphenyl (PCB118) on the mammalian reproductive system. PCB118 was administered to pregnant mice from 7.5 to 12.5 days of gestation; F1 mice were obtained and the reproductive system of F1 male mice was examined. PCB118 damaged the reproductive system in male F1 mice, as evidenced by negative effects on the testicular organ coefficient (testes weight/bodyweight), a decrease in the diameter of seminiferous tubules and a significant reduction in the anogenital distance in 35-day-old F1 mice. In addition, methylation levels of genomic DNA were reduced, with reductions in the expression of the DNA methyltransferases DNMT1, DNMT3A and DNMT3B, as well as that of the epigenetic regulatory factor ubiquitin like with PHD and ring finger domains 1 (Uhrf1). Together, the results of this study provide compelling evidence that exposure of pregnant mice to PCB118 during primordial germ cell migration in the fetus affects the reproductive system of the offspring and decreases global methylation levels in the testis.

Maternal exposure to Di-(2-ethylhexyl) phthalate (DEHP) activates the PI3K/Akt/mTOR signaling pathway in F1 and F2 generation adult mouse testis.

Di-(2-ethylhexyl) phthalate (diethylhexyl phthalate, DEHP) can cause male reproductive damage in rodents and human. Moreover, DEHP is known to promote transgenerational inheritance of adult-onset disease in subsequent generations after maternal exposure during fetal gonadal development. The PI3K/Akt/mTOR signaling pathway has been implicated in germ cell survival following testicular damage. In this study, a F0 gestation DEHP exposure and transgenerational inheritance testis injury model was established to study the testis injury phenotype and the expression and activation of members of PI3K/Akt/mTOR signaling pathway in the testis of F1-F3 generation mice. We found that the bodyweight and the anogenital distance (AGD) are reduced only in F1 mice, the sperm motility and deformity decreased in F1-F3 mice, and the testicular histomorphology damagedin F1-F3 mice; however the sperm motility and deformity rates are increased and the histomorphological injury is repaired during the transgenerational process. We also found the activation of PI3K/Akt/mTOR signaling pathway is enhanced in F1 and F2, and the number of apoptotic cells is decreased in F3 generation mice compared to the control group. These results suggest that the PI3K/Akt/mTOR signaling pathway may be activated to promote the proliferation and differentiation and protect testicular cells from apoptosis in the F1 and F2 generation mice after direct exposure to DEHP.

Multivalent weak interactions between assembly units drive synaptonemal complex formation.

The synaptonemal complex (SC) is an ordered but highly dynamic structure assembled between homologous chromosomes to control interhomologous crossover formation, ensuring accurate meiotic chromosome segregation. However, the mechanisms regulating SC assembly and dynamics remain unclear. Here, we identified two new SC components, SYP-5 and SYP-6, in Caenorhabditis elegans that have distinct expression patterns and form distinct SC assembly units with other SYPs through stable interactions. SYP-5 and SYP-6 exhibit diverse in vivo SC regulatory functions and distinct phase separation properties in cells. Charge-interacting elements (CIEs) are enriched in SC intrinsically disordered regions (IDRs), and IDR deletion or CIE removal confirmed a requirement for these elements in SC regulation. Our data support the theory that multivalent weak interactions between the SC units drive SC formation and that CIEs confer multivalency to the assembly units.

Electrophysiological and Neurochemical Considerations of Distinct Neuronal Populations in the Rat Pedunculopontine Nucleus and Their Responsiveness Following 6-Hydroxydopamine Lesions.

The pedunculopontine nucleus (PPN) is composed of a morphologically and neurochemically heterogeneous population of neurons, which is severely affected by Parkinson's disease (PD). However, the role of each subtype of neurons within the PPN in the pathophysiology of PD has not been completely elucidated. In this study, we present the discharge profiles of three classified subtypes of PPN neurons and their alterations after 6-hydroxydopamine (6-OHDA) lesion. Following 6-OHDA lesion, the spike timing of the Type II (GABAergic) and Type III (glutamatergic) neurons had phase-lock with the oscillations in the delta and beta band frequency range in the PPN, respectively. Morphological evidence has shown distinct alteration in three kinds of neurons after 6-OHDA lesion. These findings revealed that the changes in the firing characteristics of neurons in PPN in hemi-parkinsonism rats are closely associated with damaged neuronal morphology, which would make contributions to the divergence of dysfunctions in Parkinsonism.

Effects of 2,3',4,4'5-pentachlorobiphenyl exposure during pregnancy on epigenetic imprinting and maturation of offspring's oocytes in mice.

Polychlorinated biphenyls (PCBs) are a class of organic pollutants that have been widely found in the environment. The chemical 2,3',4,4'5-pentachlorobiphenyl (PCB118) is an important dioxin-like PCB compound with strong toxicity. PCB118 can accumulate in adipose tissue, serum and milk in mammals, and it is highly enriched in the follicular fluid. In this study, pregnant mice were exposed to 0, 20 and 100 µg/kg/day of PCB118 during pregnancy at the fetal primordial germ cell migration stage. The methylation patterns of the imprinted genes H19, Snrpn, Peg3 and Igf2r as well as the expression levels of Dnmt1, 3a, 3b and 3l, Uhrf1, Tet2 and Tet3 in fully grown germinal vesicle oocytes were measured in offspring. The rates of in vitro maturation, in vitro fertilization, oocyte spindle and chromosomal abnormalities were also calculated. The results showed that prenatal exposure to PCB118 altered the DNA methylation status of differentially methylated regions in some imprinted genes, and the expression levels of Dnmt1, 3a, and 3l, Uhrf1 and Tet3 were also changed. In addition, PCB118 disturbed the maturation process of progeny mouse oocytes in a dose-dependent manner. Therefore, attention should be paid to the potential impacts of PCB118-contaminated dietary intake during pregnancy on the offspring's reproductive health.

Salt-Assisted Synthesis of 3D Porous g-C3N4 as a Bifunctional Photo- and Electrocatalyst.

Graphitic carbon nitride (g-C3N4), characterized with a suitable bandgap, has aroused great interest as a robust and efficient catalyst for solar energy utilization. Herein, we introduce a new strategy to fabricate a three-dimensional (3D) porous g-C3N4 by a facile NaCl-assisted ball-milling strategy. The porous structure-induced advantages, such as a higher specific surface area, more efficient charge separation, and faster electron-transfer efficiency, enable the 3D porous g-C3N4 to achieve impressive properties as a bifunctional catalyst for both photocatalytic hydrogen evolution and electrocatalytic oxygen evolution reaction (OER). As a result, the 3D porous g-C3N4 exhibits a hydrogen evolution rate of 598 µmol h-1 g-1 with an apparent quantum yield of 3.31% at 420 nm for photocatalytic H2 generation, which is much higher than that of the bulk g-C3N4. Simultaneously, the porous g-C3N4 also presents an attractive OER performance with a low onset potential of 1.47 V (vs reversible hydrogen electrode) in an alkaline electrolyte after rational cobalt-doping. Accordingly, the NaCl-assisted ball-milling strategy paves the way to the rational design of a controllable porous structure.