Sinensetin protects against periodontitis through binding to Bach1 enhancing its ubiquitination degradation and improving oxidative stress.

Periodontitis is a chronic inflammatory and immune reactive disease induced by the subgingival biofilm. The therapeutic effect for susceptible patients is often unsatisfactory due to excessive inflammatory response and oxidative stress. Sinensetin (Sin) is a nature polymethoxylated flavonoid with anti-inflammatory and antioxidant activities. Our study aimed to explore the beneficial effect of Sin on periodontitis and the specific molecular mechanisms. We found that Sin attenuated oxidative stress and inflammatory levels of periodontal ligament cells (PDLCs) under inflammatory conditions. Administered Sin to rats with ligation-induced periodontitis models exhibited a protective effect against periodontitis in vivo. By molecular docking, we identified Bach1 as a strong binding target of Sin, and this binding was further verified by cellular thermal displacement assay and immunofluorescence assays. Chromatin immunoprecipitation-quantitative polymerase chain reaction results also revealed that Sin obstructed the binding of Bach1 to the HMOX1 promoter, subsequently upregulating the expression of the key antioxidant factor HO-1. Further functional experiments with Bach1 knocked down and overexpressed verified Bach1 as a key target for Sin to exert its antioxidant effects. Additionally, we demonstrated that Sin prompted the reduction of Bach1 by potentiating the ubiquitination degradation of Bach1, thereby inducing HO-1 expression and inhibiting oxidative stress. Overall, Sin could be a promising drug candidate for the treatment of periodontitis by targeting binding to Bach1.

Protocol for MRI-guided virus injection in macaque deep brain regions.

Effective delivery of viruses into required brain regions is critical to the success of optogenetic or chemogenetic experiments. However, in monkeys, due to the large size and heterogeneity of their brain, precise injections in deep brain regions have been challenging. Here, we present a protocol for virus injection in monkey deep brain regions under the guidance of MRI. We describe the steps for installing the guiding grid, MRI scanning, MRI-based localization, and virus injection. For complete details on the use and execution of this protocol, please refer to Chen et al. (2023).1.

Probing the processing of facial expressions in monkeys via time perception and eye tracking.

Accurately recognizing facial expressions is essential for effective social interactions. Non-human primates (NHPs) are widely used in the study of the neural mechanisms underpinning facial expression processing, yet it remains unclear how well monkeys can recognize the facial expressions of other species such as humans. In this study, we systematically investigated how monkeys process the facial expressions of conspecifics and humans using eye-tracking technology and sophisticated behavioral tasks, namely the temporal discrimination task (TDT) and face scan task (FST). We found that monkeys showed prolonged subjective time perception in response to Negative facial expressions in monkeys while showing longer reaction time to Negative facial expressions in humans. Monkey faces also reliably induced divergent pupil contraction in response to different expressions, while human faces and scrambled monkey faces did not. Furthermore, viewing patterns in the FST indicated that monkeys only showed bias toward emotional expressions upon observing monkey faces. Finally, masking the eye region marginally decreased the viewing duration for monkey faces but not for human faces. By probing facial expression processing in monkeys, our study demonstrates that monkeys are more sensitive to the facial expressions of conspecifics than those of humans, thus shedding new light on inter-species communication through facial expressions between NHPs and humans.

Male Sex Is Not a Risk Factor for Prognosis in Postoperative Thyroid Cancer Patients: A Propensity Score Matching Study.

PURPOSE: To study whether male sex is a risk factor for prognosis in patients with differentiated thyroid cancer after 131I treatment using the propensity score matching (PSM) method. METHODS: From April 2016 to October 2021, 1948 postoperative differentiated thyroid cancer patients aged 43 (interquartile range: 34, 54) years who received 131I treatment were retrospectively enrolled and divided into male (n = 645) and female groups (n = 1303). The PSM method was adopted to process all data to reduce the influence of data bias and confounding variables. The Mann-Whitney U test and χ2 test were used for data analysis. Multivariate logistic regression was used to analyze the risk factors affecting prognosis, and the receiver operating characteristic curve was used to analyze the relationship between stimulated thyroglobulin (sTg) level, 131I dose, and poor prognosis. RESULTS: Before PSM, the proportion of male patients with poor prognosis was significantly higher than that of female patients. After PSM, there was no difference in the proportion of poor prognosis between male and female groups. Multivariate logistic regression analysis showed that male sex; high T stage, N1b stage, and M1 stage; high sTg level; and high 131I dose were risk factors for poor prognosis before PSM. After PSM, high T stage, M1 stage, high sTg level, and 131I dose were still risk factors but male sex was no longer a risk factor for poor prognosis. CONCLUSIONS: After the reduction of selection bias by PSM, male sex was no longer a risk factor for prognosis after 131I treatment of differentiated thyroid cancer. In addition, high T stage (T3 + T4 stage), M1 stage, sTg ≥10.15 ng/mL, and 131I dose ≥260 mCi were risk factors for poor prognosis.

Using eZIS of SPECT to evaluate the therapeutic effect of carotid endarterectomy.

OBJECTIVE: Stroke is an acute cerebrovascular disease with high morbidity and mortality. The main causes of ischemic stroke include carotid artery stenosis, and carotid endarterectomy (CEA) can be used to improve the blood flow of the lesion site. Regional cerebral blood flow (rCBF) can be decreased by using single photon emission computed tomography (SPECT). The Easy Z-Score imaging system (eZIS) can display the changes of rCBF as Z-Score. The purpose of this study was to determine whether eZIS of SPECT can be used to evaluate the therapeutic effect of CEA in the treatment of carotid artery stenosis. METHODS: In this study, subjects were divided into the surgery group and the control group. The surgery group included seven patients with unilateral or bilateral internal carotid artery stenosis who received CEA treatment, and the control group included 11 patients who only received conventional drug treatment but did not receive surgery. Cerebral perfusion imaging (CPI) was collected twice before and after the corresponding treatment (within 6 months). rCBF of the lesion site was measured and Z-score was calculated before and after treatment by the eZIS technique. RESULTS: The postoperative Z-score of the surgery group was 0.54 ± 2.71 compared with that of the preoperative -1.34 ± 2.68 ( P = 0.0034; t = 4.687; df = 6), while the z-score of the control group was -0.33 ± 2.58 compared with that of the pretreatment 1.84 ± 2.62 ( P = 0.0010; t = 4.618; df = 10). CONCLUSION: CEA can effectively improve the blood flow in the lesion area of patients with carotid artery stenosis. eZIS of SPECT can be used to evaluate the therapeutic effect of CEA on carotid artery stenosis visually.

Primate preoptic neurons drive hypothermia and cold defense.

Maintaining body temperature within a narrow range is vital for warm-blooded animals. In rodents, the preoptic area (POA) of the hypothalamus detects and regulates core body temperature. However, knowledge about the thermal regulation center in primates remains limited. Here, we show that activating a subpopulation of POA neurons by a chemogenetic strategy reliably induces hypothermia in anesthetized and freely moving macaques. Comprehensive monitoring of physiological parameters reveals that such hypothermia is accompanied by autonomic changes including a rise in heart rate, skeletal muscle activity, and correlated biomarkers in blood. Consistent with enhanced ambulatory movement during hypothermia, the animals show a full range of cold-defense behaviors. Resting-state fMRI confirms the chemogenetic activation of POA and charts a brain-wide network of thermoregulation. Altogether, our findings demonstrate the central regulation of body temperature in primates and pave the way for future application in clinical practice.

Mapping the emergence of visual consciousness in the human brain via brain-wide intracranial electrophysiology.

Consciousness lies at the heart of our existence and experience. To probe how perceptual consciousness emerges in the brain, we recorded brain-wide intracranial electroencephalography signals from human patients while their perceptual consciousness was effectively manipulated using the continuous flash suppression paradigm. We observed substantial differences in brain activities when visual information gradually enters consciousness. Specifically, the functional connectivity first increases and then decreases, oscillations in the low-frequency band reduce in power, and those in the high-frequency band remain unchanged. We employed random forest-based classification to characterize the transitions from no perception to subconsciousness and then to consciousness, which showed an increase in signal variance at the second transition rather than the first. Further, the frontal-parietal junction dominates the first transition, whereas the temporal-frontal lobes dominate the second transition. Finally, we identified the most relevant neuronal features associated with consciousness. Altogether, these findings shed fresh light on the emergence of visual consciousness.

A pilot behavioural and neuroimaging investigation on photothrombotic stroke models in rhesus monkeys.

BACKGROUND: Ischemic stroke leads to a long-term disability in humans and no efficient clinical therapy exists to date. The middle cerebral artery occlusion (MCAO) model in non-human primates has shown to be of value for translational stroke research. New method In the current study, a photothrombotic (PT) stroke model was established in rhesus monkeys with either a proximal or distal segment of middle cerebral artery (MCA) thrombosis. This study is the first that compares the two approaches of PT stroke in monkeys using behavioral and physiological measurements and MRI scans. RESULTS: The experiment found that infarct occurred in the MCA target regions, with all monkeys having impaired behavior reflected by deficits in neurologic function, and motor and cognition in object retrieval detour (ORD) task. The monkeys with distal MCA thrombosis developed with sequential photo-irritations of the Sylvian fissure zone, adjacent central anterior gyrus and central posterior gyrus, had similar impairments with respect to behavior and showed a tendency of a small edema volume with proximal MCA thrombosis at days 4 and 7 post PT stroke. COMPARISON WITH EXISTING METHODS: The distal MCA thrombosis developed with sequential photo-irritations might provide a consistent and well-tolerated focal ischemia in rhesus monkeys, compared with other PT stroke models which usually were singly conducted on the animal's motor cortex and had a temporal effect. CONCLUSIONS: The sequentially photo-irritated PT stroke model is a promising ischemic stroke model in rhesus monkey for studying human stroke pathology and physiology and for new therapies development.

Extracellular Vesicle-Mediated Delivery of Circular RNA SCMH1 Promotes Functional Recovery in Rodent and Nonhuman Primate Ischemic Stroke Models.

BACKGROUND: Stroke is a leading cause of adult disability that can severely compromise the quality of life of patients, yet no effective medication currently exists to accelerate rehabilitation. A variety of circular RNA (circRNA) molecules are known to function in ischemic brain injury. Lentivirus-based expression systems have been widely used in basic studies of circRNAs, but safety issues with such delivery systems have limited exploration of the potential therapeutic roles for circRNAs. METHODS: Circular RNA SCMH1 (circSCMH1) was screened from the plasma of patients with acute ischemic stroke by using circRNA microarrays. Engineered rabies virus glycoprotein-circSCMH1-extracellular vesicles were generated to selectively deliver circSCMH1 to the brain. Nissl staining was used to examine infarct size. Behavioral tasks were performed to evaluate motor functions in both rodent and nonhuman primate ischemic stroke models. Golgi staining and immunostaining were used to examine neuroplasticity and glial activation. Proteomic assays and RNA-sequencing data combined with transcriptional profiling were used to identify downstream targets of circSCMH1. RESULTS: CircSCMH1 levels were significantly decreased in the plasma of patients with acute ischemic stroke, offering significant power in predicting stroke outcomes. The decreased levels of circSCMH1 were further confirmed in the plasma and peri-infarct cortex of photothrombotic stroke mice. Beyond demonstrating proof-of-concept for an RNA drug delivery technology, we observed that circSCMH1 treatment improved functional recovery after stroke in both mice and monkeys, and we discovered that circSCMH1 enhanced the neuronal plasticity and inhibited glial activation and peripheral immune cell infiltration. CircSCMH1 binds mechanistically to the transcription factor MeCP2 (methyl-CpG binding protein 2), thereby releasing repression of MeCP2 target gene transcription. CONCLUSIONS: Rabies virus glycoprotein-circSCMH1-extracellular vesicles afford protection by promoting functional recovery in the rodent and the nonhuman primate ischemic stroke models. Our study presents a potentially widely applicable nucleotide drug delivery technology and demonstrates the basic mechanism of how circRNAs can be therapeutically exploited to improve poststroke outcomes.

Inhibition of Notch signaling pathway enhanced the radiosensitivity of breast cancer cells.

This study aimed to investigate the effect of inhibiting the Notch signaling pathway on the radiosensitivity of breast cancer cells. Human breast cancer cell lines (MCF-7 and T47D) were selected and treated with radiation of different doses. Cells were treated with Gamma secretase inhibitor (GSI) to analyze the effects of GSI on the Notch signaling, which were detected by Immunofluorescence assay, RT-qPCR, and Western blot analysis. Besides, Transwell assay, Scratch test, colony formation assay, MTT assay, and flow cytometry were conducted to show the effects of GSI on the invasion and migration, survival fraction, cell viability, and apoptosis of MCF-7 and T47D cells after radiation therapy. Moreover, cell transfection with a dominant negative mutant of RBPJ, the key transcription factor of Notch signaling pathway, were also applied to show the inhibition of Notch signaling pathway. Initially, we found that the 4 Gy radiation activated Notch signaling pathway, and enhanced the invasion and migration of MCF-7 and T47D cells. However, GSI inhibited the Notch signaling pathway, and reversed the enhancement of radiation on the migration and invasion, promoted the enhancement of apoptosis and inhibition of proliferation of MCF-7 and T47D cells induced by radiation. Except that, we also determined that GSI and dnRBPJ suppressed the upregulation of Notch signaling after radiation therapy. Our study demonstrated that inhibition of the Notch signaling pathway enhanced the radiosensitivity of breast cancer cells, which may provide evident for a beneficial adjuvant therapy in the breast cancer treatment.

RNA Binding Protein RNPC1 Inhibits Breast Cancer Cell Metastasis via Activating STARD13-Correlated ceRNA Network.

RNA binding proteins (RBPs) are pivotal post-transcriptional regulators. RNPC1, an RBP, acts as a tumor suppressor through binding and regulating the expression of target genes in cancer cells. This study disclosed that RNPC1 expression was positively correlated with breast cancer patients' relapse-free and overall survival and that RNPC1 suppressed breast cancer cell metastasis. Mechanistically, RNPC1 promotes competing endogenous RNA (ceRNA) network crosstalk among STARD13, CDH5, HOXD10, and HOXD1 (STARD13-correlated ceRNA network), which we previously confirmed in breast cancer cells through stabilizing the transcripts and thus facilitating the expression of these four genes in breast cancer cells. Furthermore, RNPC1 overexpression restrained the promotion of STARD13, CDH5, HOXD10, and HOXD1 knockdown on cell metastasis. Notably, RNPC1 expression was positively correlated with CDH5, HOXD1, and HOXD10 expression in breast cancer tissues and attenuated adriamycin resistance. Taken together, these results identified that RNPC1 could inhibit breast cancer cell metastasis via promoting a STARD13-correlated ceRNA network.

StarD13 3'-untranslated region functions as a ceRNA for TP53INP1 in prohibiting migration and invasion of breast cancer cells by regulating miR-125b activity.

Competitive endogenous messenger RNA (ceRNA) affects transcription of other RNA molecules by competitively binding common microRNAs. Previous studies have shown that TP53INP1 functions as a suppressor in tumor metastasis. Our study elucidated StarD13 messenger RNA as a ceRNA in regulating migration and invasion of breast cancer cells. MicroRNA-125b was identified to induce metastasis of MCF-7 cells and bind with both StarD13 3'UTR and TP53INP1 3'UTR. Therefore, a ceRNA interaction between StarD13 and TP53INP1 mediated by competitively binding to miR-125b was indicated. Importantly, a microRNA-125b binding site at 4546-4560 nt on StarD13 was verified more vital for this ceRNA interaction. Indirectly regulation of SPARC in inducing metastasis of breast cancer cells by StarD13 via competitively binding with TP53INP1 was further confirmed. In conclusion, our findings demonstrate a ceRNA regulatory network which could give a better understanding of metastatic mechanisms of breast cancer.

Displacement of Bax by BMF Mediates STARD13 3'UTR-Induced Breast Cancer Cells Apoptosis in an miRNA-Depedent Manner.

The balance of pro- and antiapoptotic gene expression programs dominates the apoptotic progress of cancer cells. We previously demonstrated that STARD13 3'UTR suppressed breast cancer metastasis via inhibiting epithelial-mesenchymal transition (EMT). However, the roles of STARD13 3'UTR in breast cancer apoptosis remain elusive. Here, we identified that STARD13 3'UTR promoted cell apoptosis in vitro and in vivo. Mechanistically, STARD13 3'UTR acted as a ceRNA for BMF (Bcl-2 modifying factor), thus increasing BMF expression in an miRNA-dependent manner. Meanwhile, STARD13 3'UTR enhanced the interaction of BMF/Bcl-2 to release Bax (Bcl-2 associated X protein) in breast cancer cells. Finally, we verified the ceRNA relationship between STARD13 and BMF in vivo. Collectively, these findings suggest that STARD13 3'UTR could act as a ceRNA for BMF to promote apoptosis and recognize STARD13 3'UTR as a potential therapeutic target in breast cancer cells.

The CCR2 3'UTR functions as a competing endogenous RNA to inhibit breast cancer metastasis.

Diverse RNA transcripts acting as competing endogenous RNAs (ceRNAs) can co-regulate each other's expression by competing for shared microRNAs. CCR2 protein, the receptor for CCL2, is implicated in cancer progression. However, we found that a higher CCR2 mRNA level is remarkably associated with prolonged survival of breast cancer patients. These conflicting results prompted us to study the non-coding function of CCR2 mRNA. We found that the CCR2 3' untranslated region (UTR) inhibited MDA-MB-231 and MCF-7 cell metastasis by repressing epithelial-mesenchymal transition (EMT) in vitro, and suppressed breast cancer metastasis in vivo Mechanistically, the CCR2 3'UTR modulated the expression of the RhoGAP protein STARD13 via acting as a STARD13 ceRNA in a microRNA-dependent and protein coding-independent manner. The CCR2 3'UTR blocked the activation of RhoA-ROCK1 pathway, which is the downstream effector of STARD13, and thus decreased the phosphorylation level of myosin light chain 2 (MLC2) and formation of F-actin. Additionally, the function of the CCR2 3'UTR was dependent on STARD13 expression. In conclusion, our results confirmed that the CCR2 3'UTR acts as a metastasis suppressor by acting as a ceRNA for STARD13 and thus inhibiting RhoA-ROCK1-MLC-F-actin pathway in breast cancer cells.This article has an associated First Person interview with the first author of the paper.

Akr1 attenuates methylmercury toxicity through the palmitoylation of Meh1 as a subunit of the yeast EGO complex.

BACKGROUND: We previously reported that palmitoyltransferase activity of Akr1 is required for alleviation of methylmercury toxicity in yeast. In this study, we identified a factor that alleviates methylmercury toxicity among the substrate proteins palmitoylated by Akr1, and investigated the role of this factor in methylmercury toxicity. METHODS: Gene disruption and site-directed mutagenesis were used to examine the relationship of methylmercury toxicity and vacuole function. Palmitoylation was investigated using the acyl-biotinyl exchange method. Vacuoles were stained with the fluorescent probe FM4-64. RESULTS: We found that Meh1 (alias Ego1), a substrate protein of Akr1, participates in the alleviation of methylmercury toxicity. Moreover, almost no palmitoylation of Meh1 when Akr1 was knocked out, and mutant Meh1, which is not palmitoylated, did not show alleviation of methylmercury toxicity. The palmitoylated Meh1 was involved in the alleviation of methylmercury toxicity as a constituent of EGO complex which suppresses autophagy. Methylmercury caused vacuole deformation, and this was greater in the yeasts knocking out the EGO complex subunits. 3-Methyladenine, an autophagy inhibitor, suppresses vacuole deformation and cytotoxicity caused by methylmercury. The elevated methylmercury sensitivity by Meh1 knockout almost completely disappeared in the presence of 3-methyladenine. CONCLUSIONS: Akr1 reduces methylmercury toxicity through palmitoylation of Meh1. Furthermore, the EGO complex including Meh1 reduces methylmercury toxicity by suppressing the induction of vacuole deformation caused by methylmercury. GENERAL SIGNIFICANCE: These findings propose that Meh1 palmitoylated by Akr1 may act as a constituent of the EGO complex when contributing to the decreased cytotoxicity by negatively controlling the induction of autophagy by methylmercury.

MiR-9 enhances the sensitivity of A549 cells to cisplatin by inhibiting autophagy.

OBJECTIVES: To demonstrate that miR-9 inhibits autophagy by down-regulating Beclin1 and thus enhances the sensitivity of A549 cells to cisplatin. RESULTS: MiR-9 inhibited Beclin1 expression by binding to its 3'UTR. The inhibition decreased the cisplatin-induced autophagy in A549 cells, evidenced by the decreased expression of LC3II and GFP-LC3 puncta and the increased expression of P62. Upregulation of miR-9 level enhanced the sensibility of A549 cells to cisplatin and increased the cisplatin-induced apoptosis. Overexpression of Beclin1 reversed above effects of miR-9 mimics, cisplatin-induced autophagy was increased and apoptosis was decreased. CONCLUSIONS: MiR-9 inhibits autophagy via targeting Beclin1 3'UTR and thus enhances cisplatin sensitivity in A549 cells.

Competing endogenous RNA networks of CYP4Z1 and pseudogene CYP4Z2P confer tamoxifen resistance in breast cancer.

Patients with estrogen receptor α (ERα)-positive breast cancer can be treated with endocrine therapy using anti-estrogens such as tamoxifen; nonetheless, patients often develop resistance limiting the success of breast cancer treatment. The potential mechanisms remain elusive. In detail, many miRNAs have been associated with breast cancer tamoxifen resistance, but no studies have addressed the role of miRNA-mediated competitive endogenous RNAs network (ceRNET) in tamoxifen resistance. The ceRNET between CYP4Z1 and pseudogene CYP4Z2P has been revealed to promote breast cancer angiogenesis. However, its function in tamoxifen resistance remains unclear. Here we report CYP4Z1 and CYP4Z2P were downregulated in MCF-7 cells compared with tamoxifen-resistant MCF-7-TamR cells. Enforced upregulation of CYP4Z1- or CYP4Z2P-3'UTR level renders MCF-7 Cells resistant to tamoxifen. We find that overexpression of CYP4Z1- or CYP4Z2P-3'UTR enhances the transcriptional activity of ERα through the activation of ERα phosphorylation. Furthermore, we find that CYP4Z1- and CYP4Z2P-3'UTRs increase ERα activity dependent on cyclin-dependent kinase 3 (CDK3). Reporter gene and western blot assays revealed that CYP4Z1- and CYP4Z2P-3'UTRs act as CDK3 ceRNAs. More importantly, the blocking of CYP4Z1- and CYP4Z2P-3'UTRs reversed tamoxifen resistance in MCF-7-TamR cells. Our data demonstrates that the ceRNET between CYP4Z1 and pseudogene CYP4Z2P acts as a sub-ceRNET to promote CDK3 expression in ER-positive breast cancer and is a potential therapeutic target for treatment of tamoxifen-resistant breast cancer.

[Effects of UV-B radiation on the decomposition of Cunninghamia lanceolata leaf litter].

A litterbag experiment was conducted to study the decomposition of Cunninghamia lanceolata leaf litter under ambient and reduced UV-B radiation (22.1% below ambient). Comparing with ambient treatment, the reduced treatment decreased the decomposition rate of C. lanceolata leaf litter by 69.6% (P<0.001), making the relative contents of nitrogen (N), phosphorus (P), and lignin in the litter increased by 150%, 83.3%, and 13.8%, respectively, and the release of potassium (K) and carbon (C) slowed down. In the process of litter decomposition, photo-degradation of lignin didn't play crucial role. The results suggested that UV-B radiation could accelerate the decomposition rate of C. lanceolata leaf litter, promote the release of N, P, K, and C from it, and increase the nutrients turnover rate in litter layer as well as the carbon flux on the ground, giving potential effects on the function of C. lanceolata forest as a carbon source or sink in humid subtropical China.

[Characteristics of soil seed banks in logging gaps of forests at different succession stages in Changbai Mountains].

An investigation was made on the soil seed banks in the logging gaps of Populus davidiana--Betula platyphylla secondary forest, secondary broad-leaved forest, and broad-leaved Korean pine mixed forest at their different succession stages in Changbai Mountains. Among the test forests, secondary broad-leaved forest had the highest individual density (652 ind x m(-2)) in its soil seed bank. With the succession of forest community, the diversity and uniformity of soil seed bank increased, but the dominance decreased. The seed density of climax species such as Pinus koraiensis, Abies nephrolepis, and Acer mono increased, whereas that of Maackia amurensis and Fraxinus mandshurica decreased. Moreover, the similarity in species composition between soil seed bank and the seedlings within logging gaps became higher. The individual density and similarity between soil seed bank and the seedlings in non-logging gaps were similar to those in logging gaps. All of these indicated that soil seed bank provided rich seed resources for forest recovery and succession, and the influence of soil seed bank on seedlings regeneration increased with the succession.