ObjectiveTo investigate the neuroprotective effects of Tiaoseng decoction on a perimenopausal depression （PMD） rat model and to examine its regulatory influence on the estrogen receptor β （ERβ）/monoamine oxidase A （MAOA）/c-Jun N-terminal kinase （JNK） signaling pathway， thereby elucidating its potential mechanisms of action. MethodsForty-eight female Sprague-Dawley （SD） rats were divided into a sham group， a model group， a 17β-estradiol （E₂） group （2.5 × 10^-5 g·kg^-1）， and low-， medium-， and high-dose Tiaoseng decoction groups （9.69， 19.37， 38.74 g·kg^-1） by using a random number table method， with eight rats in each group. The PMD model was replicated using ovariectomy （OVX） combined with chronic unpredictable mild stress （CUMS） and was treated continuously with 17β-E₂ and different doses of Tiaoseng decoction for 28 d， once a day. Depressive-like behaviors were assessed using the sucrose preference test， open-field test， and forced swim test. Histopathological changes in the prefrontal cortex were examined using hematoxylin-eosin （HE） and Nissl staining. Enzyme-linked immunosorbent assay （ELISA） was performed to measure serum levels of E₂， luteinizing hormone （LH）， and follicle-stimulating hormone （FSH）， as well as reactive oxygen species （ROS）， malondialdehyde （MDA）， and superoxide dismutase （SOD） levels in the prefrontal cortex. Mitochondrial ultrastructure of prefrontal cortex neurons was observed via transmission electron microscopy. Terminal deoxynucleotidyl transferase dUTP nick-end labeling （TUNEL） was used to detect neuronal apoptosis in the prefrontal cortex. Immunohistochemistry （IHC） was applied to analyze the protein expression of brain-derived neurotrophic factor （BDNF）， postsynaptic density protein95 （PSD95）， and synaptophysin （SYP） in the prefrontal cortex. Immunofluorescence （IF） was conducted to evaluate the average fluorescence intensity of ERβ and MAOA in the prefrontal cortex. Western blot and real-time quantitative polymerase chain reaction （Real-time PCR） were used to determine the protein and mRNA expression levels of key molecules in the ERβ/MAOA/JNK signaling pathway in the prefrontal cortex. ResultsCompared with the sham group， the model group had significantly reduced sugar-water preference index， total distance traveled， average speed， and activity time in the central region in the open field experiment， E₂ content， and SOD content （P<0.01） and significantly reduced BDNF， PSD95， SYP， and ERβ expression and B-cell lymphoma-2 （Bcl-2）/Bcl-2-associated X protein （Bax） ratio （P<0.01）. Additionally， the model group exhibited severe histopathological damage， disrupted mitochondrial structure， cristae disappearance or fracture， swelling， and deformation in the prefrontal lobe. The immobilization time of forced swimming， TUNEL positivity， LH， FSH， MDA， ROS， MAOA， Caspase-3， p-JNK/JNK， and p-c-Jun/c-Jun were significantly increased in the model group compared with the sham group （P<0.01）. Compared with the model group， the low-， medium-， and high-dose Tiaoseng decoction groups and the 17β-E₂ group had increased sugar-water preference index， total distance traveled， average speed， and activity time in the central region in the open-field test， E₂ content， and SOD content （P<0.05， P<0.01） and elevated BDNF， PSD95， SYP， and ERβ expression and Bcl-2/Bax ratio （P<0.05， P<0.01）. In addition， histopathological damage to the prefrontal lobe was improved to different degrees， and mitochondrial structure was gradually repaired. The immobilization time of forced swimming， TUNEL positivity， LH， FSH， MDA， ROS， MAOA， Caspase-3， p-JNK/JNK and p-c-Jun/c-Jun were significantly reduced in the low-， medium-， and high-dose Tiaoseng decoction groups and the 17β-E₂ group compared with the model group （P<0.05， P<0.01）. ConclusionTiaoseng decoction has significant neuroprotective effects on PMD model rats， which can alleviate perimenopausal depressive disorder by inhibiting oxidative stress， attenuating neuronal apoptosis， and restore synaptic plasticity via ERβ/MAOA/JNK signaling pathway regulation.

Objective To investigate the expression of Cyclin F in clear cell renal cell carcinoma (ccRCC), its clinicopathological characteristics, and its effect on the biological behavior of renal cancer cell lines Methods RT-qPCR and Western blot were used to detect the mRNA and protein expression of Cyclin F in fresh ccRCC specimens. Immunohistochemistry assay was performed to detect the expression of Cyclin F protein in 80 paraffin samples. CCK-8 assay, scratch assay, and flow cytometry were conducted to determine the effects of Cyclin F overexpression on the proliferation, migration, and apoptosis of renal cancer cell lines. Results The expression of Cyclin F in cancer tissues was higher than that in adjacent tissues at the mRNA level (P<0.0285). The expression of Cyclin Fprotein in cancer tissues was lower than that in adjacent tissues (P<0.001). The analysis of clinicopathological parameters showed that the low expression of Cyclin F was correlated with WHO/ISUP grade, Ki67 index, and age (P=0.041, 0.047, 0.008,). In vitro experiments confirmed that overexpression of Cyclin F promoted the proliferation (P<0.001) and migratory ability (24 h P=0.003, 48 h P=0.0058) of the RCC 786-O cell line, while inhibiting apoptosis (P=0.0019). Conclusion The low expression of Cyclin F protein in ccRCC tissuesis associated with high ISUP grade, high Ki67 index, oldage, and prognosis of patients.

OBJECTIVE To provide a reference for optimizing and promoting the quality standards of Shechuan naolitong granules. METHODS Fifteen batches of Shechuan naolitong granules were used as samples to establish HPLC fingerprints using the Similarity Evaluation System for Chromatographic Fingerprint of Traditional Chinese Medicine （2012 edition）. Similarity evaluation and common peak identification were performed， and orthogonal partial least squares discriminant analysis （OPLS-DA） was used to assess quality differences among different batches and to screen quality differential components. Using salvianolic acid B（SAB） as the internal reference， quantitative analysis of multi-components by single-marker （QAMS） was developed to simultaneously determine geniposidic acid （GA）， chlorogenic acid （CA）， vaccarin （VA）， ferulic acid （FA） and senkyunolide I （SI）. The results were compared with those obtained by the external standard method. RESULTS A total of 13 common peaks were identified in the HPLC fingerprints of 15 batches of samples， and the similarities of the spectra were all above 0.96. Seven chromatographic peaks were identified as GA （peak 3）， CA （peak 6）， VA （peak 8）， FA （peak 9）， SI （peak 11）， SAB（peak 12） and TA（peak 13）. OPLS-DA indicated that the differential quality markers among 15 batches were peaks 5， 11 （SI）， and 12 （SAB）.Using SAB as the internal reference， the relative correction factors for GA， CA， VA， FA and SI were calculated as 1.058 4， 0.594 3， 0.643 3， 0.342 7 and 0.262 8， respectively. The mean content of GA， CA， VA， FA， SI and SAB across the 15 batches of samples were 0.155 0， 0.085 4， 0.140 3， 0.071 8， 0.072 7， 1.276 3 mg/g， respectively， showing no significant difference compared with the ESM （P＞0.05）. CONCLUSIONS The established HPLC fingerprint and QAMS are simple， efficient and economical， providing a reference for the quality control and further development of Shechuan naolitong granules.

Ischemic stroke is the leading cause of disability and mortality worldwide. The blood‒brain barrier (BBB) is the first line of defense after ischemic stroke. Disruption of the BBB induced by brain microvascular endothelial cells (BMECs) dysfunction is a key event that triggers secondary damage to the central nervous system, where blood-borne fluids and immune cells penetrate the brain parenchyma, causing cerebral edema and inflammatory response and further aggravating brain damage. Here, we develop a novel artificial mesenchymal stem cell (MSC) extracellular vesicles by integrating MSC membrane proteins into liposomal bilayers, which encapsulated miR-132-3p with protective effects on BMECs. The artificial extracellular vesicles (MSCo/miR-132-3p) had low immunogenicity to reduce non-specific clearance by the mononuclear phagocytosis system (MPS) and could target ischemia-injured BMECs. After internalization into the damaged BMECs, MSCo/miR-132-3p escaped the lysosomes via the H_II phase transition of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) and decreased cellular reactive oxygen species (ROS) and apoptosis levels by regulating the RASA1/RAS/PI3K/AKT signaling pathway. In the transient middle cerebral artery occlusion (tMCAO) models, MSCo/miR-132-3p targeted impaired brain regions (approximately 9 times the accumulation of plain liposomes at 12 h), reduced cerebral vascular disruption, protected BBB integrity, and decreased infarct volume (from 44.95% to 6.99%).

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*

Corticotomy is a clinical procedure to accelerate orthodontic tooth movement characterized by the regional acceleratory phenomenon (RAP). Despite its therapeutic effects, the surgical risk and unclear mechanism hamper the clinical application. Numerous evidences support macrophages as the key immune cells during bone remodeling. Our study discovered that the monocyte-derived macrophages primarily exhibited a pro-inflammatory phenotype that dominated bone remodeling in corticotomy by CX3CR1^CreERT2; R26^GFP lineage tracing system. Fluorescence staining, flow cytometry analysis, and western blot determined the significantly enhanced expression of binding immunoglobulin protein (BiP) and emphasized the activation of sensor activating transcription factor 6 (ATF6) in macrophages. Then, we verified that macrophage specific ATF6 deletion (ATF6^f/f; CX3CR1^CreERT2 mice) decreased the proportion of pro-inflammatory macrophages and therefore blocked the acceleration effect of corticotomy. In contrast, macrophage ATF6 overexpression exaggerated the acceleration of orthodontic tooth movement. In vitro experiments also proved that higher proportion of pro-inflammatory macrophages was positively correlated with higher expression of ATF6. At the mechanism level, RNA-seq and CUT&Tag analysis demonstrated that ATF6 modulated the macrophage-orchestrated inflammation through interacting with Tnfα promotor and augmenting its transcription. Additionally, molecular docking simulation and dual-luciferase reporter system indicated the possible binding sites outside of the traditional endoplasmic reticulum-stress response element (ERSE). Taken together, ATF6 may aggravate orthodontic bone remodeling by promoting Tnfα transcription in macrophages, suggesting that ATF6 may represent a promising therapeutic target for non-invasive accelerated orthodontics.
Animals ; Mice ; Macrophages/metabolism* ; Tumor Necrosis Factor-alpha/genetics* ; Tooth Movement Techniques/methods* ; Activating Transcription Factor 6/metabolism* ; Bone Remodeling ; Flow Cytometry ; Blotting, Western

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

Lung cancer is the malignant tumor with the highest incidence and mortality rate worldwide.For lung adenocarcinoma,identifying specific gene mutations,fusions,and giving corresponding targeted drugs can greatly improve the survival time of the patients.Among them,anaplastic lymphoma kinase(ALK)fusion occurs in 3％-7％of non-small cell lung cancer(NSCLC).In clinical practice,a variety of detection methods can be used to determine the ALK fusion status,but false negative test results are possible.This paper retrospectively analyzed the diagnosis and treatment of a patient with lung adeno-carcinoma,judged the ALK fusion status by various detection methods.Among them,immunohistochemistry(IHC)(Ventana D5F3),RNA based next-generation sequencing(RNA-based NGS)confirmed positive echinoderm microtubule associated protein like 4(EML4)-ALK fusion,while DNA-based NGS was negative.This paper analyzed the detection methods of ALK fusion,in order to clarify which detection method is the most accurate and simple to choose in different clinical cases and guide the subsequent treatment.

Objective:To investigate the effects of gambogenic acid(GNA)on the proliferation and apoptosis of CAL27 cell xenograft tumor in nude mice.Methods:18 SPF nude mice were randomly divided into 3 groups(n=6).All nude mice were inoculated with CAL27 cells at logarithmic growth stage to establish subcutaneous transplanted tumor models.The mice in low and high dose GNA groups were treated with GNA of 8.0 mg/kg iv.and 16.0 mg/kg iv.every other day,respectively,and those in the control group was given the same amount of normal saline.The tumor growth curve was plotted during drug administration.2 weeks later,the nude mice were sacrificed,the tumor tissue was removed and the tumor inhibition rate was evaluated by the tumor size measurements.TUNEL as-say was used to detect the apoptosis of transplanted tumor cells in the groups.The expression levels of AKT,Bcl-2 and PI3K proteins in tumor tissue were detected by immunohistochemistry(IHC).The toxicity and side effects of GNA on normal tissues were detected by HE staining.Results:The transplanted tumors in low and high dose GNA groups grew slowly,and the tumor weight and volume were significantly lower than those in the control group(P＜0.05),the tumor inhibition ratio of low and high dose groups was 57.58％and 83.68％respectively.TUNEL results showed that the apoptosis index of GNA low and high dose groups was higher that of control group(P＜0.05).IHC results showed that the expression of AKT,Bcl-2 and PI3K in the tumor tissues of nude mice in low and high dose GNA groups was lower than that in the control group(P＜0.05).HE results showed that GNA had not effect on normal tissues and or-gans(P＜0.05);Conclusion:GNA may induce CAL27 cell apoptosis by regulating the expression of AKT,Bcl-2 and PI3K,and in-hibites the development of human tongue squamous cell carcinoma with little effect on normal tissues and organs.

Objective To investigate the in vitro anti-inflammatory effects of Sophora davidii Hance leaves total alkaloids(SDLTAs)and possible molecular mechanisms.Methods The lipopolysaccharide(LPS)-induced inflammation model of RAW264.7 cells was used,and different concentrations of SDLTAs(50,100 and 200 μg/mL)were administered,and the effect of SDLTAs on cellular NO expression was detected by the Griess method;ELISA method was used to detect the effect of SDLTAs on the expression of IL-6,TNF-α and IL-1β;The expression of iNOS,NF-κB p65 and IκBα mRNA was detected by RT-qPCR;Western blotting was used to detecte the expres-sion of p-p38,p-p65 and p-JNK in the cells and NF-κB p65 in the nucleus.Results SDLTAs could significantly inhibit the LPS-induced inflammatory response in RAW264.7 cells.SDLTAs significantly decreased the secretion of NO,IL-6,TNF-α and IL-1β in cells(P<0.01),and significantly decreased the mRNA expressions of iNOS,NF-κB p65 and IκBα in cells(P<0.01).Significantly decreased the protein expression of p-p38,p-p65 and p-JNK in cells and NF-κB p65 in nucleus(P<0.01).Conclusion SDLTAs can exert anti-inflammatory effects by regulating the MAPK/NF-κB signalling pathway.