Neuronal injury in glaucoma persists despite effective intraocular pressure (IOP) control, necessitating neuroprotective strategies for retinal ganglion cells (RGCs). In this study, we investigated the neuroprotective role of the γ-hydroxybutyrate analog HOCPCA in a glaucoma model, focusing on its effects on CaMKII signaling, oxidative stress, and neuroinflammatory responses. Retinal tissue from high IOP animal models was analyzed via proteomics. In vitro mouse retinal explants were subjected to elevated pressure and oxidative stress, followed by HOCPCA treatment. HOCPCA significantly mitigated the RGC loss induced by oxidative stress and elevated pressure, preserving neuronal function. It restored CaMKIIα and β levels, preserving RGC integrity, while also modulating oxidative stress and neuroinflammatory responses. These findings suggest that HOCPCA, through its interaction with CaMKII, holds promise as a neuroprotective therapy for glaucoma.
Animals ; Retinal Ganglion Cells/metabolism* ; Glaucoma/pathology* ; Oxidative Stress/drug effects* ; Neuroprotective Agents/pharmacology* ; Mice ; Calcium-Calmodulin-Dependent Protein Kinase Type 2/metabolism* ; Mice, Inbred C57BL ; Disease Models, Animal ; Neuroinflammatory Diseases/drug therapy* ; Neuroprotection/drug effects* ; Male ; Intraocular Pressure/drug effects*

Glutamine provides carbon and nitrogen to support the proliferation of cancer cells. However, the precise reason why cancer cells are particularly dependent on glutamine remains unclear. In this study, we report that glutamine modulates the tumor suppressor F-box and WD repeat domain-containing 7 (FBW7) to promote cancer cell proliferation and survival. Specifically, lysine 604 (K604) in the sixth of the 7 substrate-recruiting WD repeats of FBW7 undergoes glutaminylation (Gln-K604) by glutaminyl tRNA synthetase. Gln-K604 inhibits SCFFBW7-mediated degradation of c-Myc and Mcl-1, enhances glutamine utilization, and stimulates nucleotide and DNA biosynthesis through the activation of c-Myc. Additionally, Gln-K604 promotes resistance to apoptosis by activating Mcl-1. In contrast, SIRT1 deglutaminylates Gln-K604, thereby reversing its effects. Cancer cells lacking Gln-K604 exhibit overexpression of c-Myc and Mcl-1 and display resistance to chemotherapy-induced apoptosis. Silencing both c-MYC and MCL-1 in these cells sensitizes them to chemotherapy. These findings indicate that the glutamine-mediated signal via Gln-K604 is a key driver of cancer progression and suggest potential strategies for targeted cancer therapies based on varying Gln-K604 status.
Glutamine/metabolism* ; Myeloid Cell Leukemia Sequence 1 Protein/genetics* ; Humans ; Proto-Oncogene Proteins c-myc/genetics* ; Cell Proliferation ; Signal Transduction ; Neoplasms/pathology* ; F-Box-WD Repeat-Containing Protein 7/genetics* ; Cell Survival ; Cell Line, Tumor ; Apoptosis

OBJECTIVE: To investigate chronic hepatitis C virus (HCV) infection's effect on gestational liver function, pregnancy and delivery complications, and neonatal development. METHODS: A total of 157 HCV antibody-positive (anti-HCV[+]) and HCV RNA(+) patients (Group C) and 121 anti-HCV(+) and HCV RNA(-) patients (Group B) were included as study participants, while 142 anti-HCV(-) and HCV RNA(-) patients (Group A) were the control group. Data on biochemical indices during pregnancy, pregnancy complications, delivery-related information, and neonatal complications were also collected. RESULTS: Elevated alanine aminotransferase (ALT) rates in Group C during early, middle, and late pregnancy were 59.87%, 43.95%, and 42.04%, respectively-significantly higher than Groups B (26.45%, 15.70%, 10.74%) and A (23.94%, 19.01%, 6.34%) ( P < 0.05). Median ALT levels in Group C were significantly higher than in Groups A and B at all pregnancy stages ( P < 0.05). No significant differences were found in neonatal malformation rates across groups ( P > 0.05). However, neonatal jaundice incidence was significantly greater in Group C (75.16%) compared to Groups A (42.25%) and B (57.02%) ( χ ² = 33.552, P < 0.001). HCV RNA positivity during pregnancy was an independent risk factor for neonatal jaundice ( OR = 2.111, 95% CI 1.242-3.588, P = 0.006). CONCLUSIONS Chronic HCV infection can affect the liver function of pregnant women, but does not increase the pregnancy or delivery complication risks. HCV RNA(+) is an independent risk factor for neonatal jaundice.
Humans ; Female ; Pregnancy ; Adult ; Pregnancy Complications, Infectious/epidemiology* ; Retrospective Studies ; Pregnancy Outcome ; Infant, Newborn ; Viremia/virology* ; Hepatitis C ; Hepacivirus/physiology* ; Hepatitis C, Chronic/virology* ; Young Adult ; Alanine Transaminase/blood*

A mounting body of research suggests that circRNAs significantly contribute to the develop-ment of myocardial fibrosis.The microarray results of human circular RNA expression profile indicated that circHERC4_041 expression increased in the myocardium of patients with heart failure,RT-qPCR a-nalysis confirmed that the myocardial expression level of circHERC4_041 in individuals with heart failure were considerably elevated compared to that in healthy organ donors.Fluorescence in situ hybridization(FISH)confirmed that circHERC4_041 was abundant in the cytoplasm of human cardiomyocyte AC16.Overexpression of circHERC4_041 in mouse myocardial fibroblasts(mCFs)mediated by adenovirus in-hibited the expression of fibrosis-related proteins in mCFs.Experiments involving cell proliferation,wound healing,and Transwell assays demonstrated that overexpression of circHERC4_041 suppressed the growth and mobility of mCFs(P＜0.001).Sequence analysis results suggested that circHERC4_041 con-tains potential ribosome entry sequence(IRES)and open reading frame(ORF).Western blot confirmed that circHERC4_041 could translate the 516 amino acid HERC4-516aa protein,which was mainly located in the cytoplasm of the cell.Cell functional experiments confirmed that circHERC4_041 inhibited the fi-brotic phenotype of mCFs by specifically translating HERC4-516aa(P＜0.05).The specific interaction between HERC4-516aa and transglutaminase 2(TGM2)was confirmed by IP-MS screening and Co-IP i-dentification.Further results found that the degradation of TGM2 was promoted through proteasome path-way.The overexpression of TGM2 in mCFs facilitated by adenoviral vectors could counteract the suppres-sive effects of HERC4-516aa on the fibrotic phenotype of mCFs.Therefore,this study confirmed that the HERC4-516aa protein translated by circHERC4_041 can specifically bind to TGM2 to inhibit the fibrotic phenotype of myocardial fibroblasts.

Objective:To compare the effects of standard cognitive behavioral therapy for insomnia (CBT-I) and enhanced cognitive behavioral therapy for insomnia(CBT-I Plus) in patients with chronic insomnia disorder comorbid anxiety or depressive symptoms.Methods:This prospective study included 148 patients with chronic insomnia disorder and anxiety/depression symptoms who were treated at the Sleep Disorder clinic of Shanghai Mental Health Center between July 2020 and August 2023. Participants (56 males, 92 females; aged 18-65 years, mean age 35.08±10.30 years) were randomly assigned in a 1∶2 ratio to the CBT-I group ( n=54) or CBT-I Plus group ( n=94). The CBT-I Plus group received additional treatments targeting anxiety and depressive symptoms. Treatment lasted 8 weeks, with assessment conducted at baseline, weeks 2, 4, and 8. Depression severity was measured using the 17-item Hamilton Depression Rating Scale (HAMD 17), anxiety severity with the Hamilton Anxiety Scale (HAMA), and sleep quality with the Pittsburgh Sleep Quality Index (PSQI). Paired sample t-tests were used to evaluate within-group changes, repeated-measures ANOVA compared treatment effects between groups, and ANCOVA was employed to adjust for confounding variables. Results:Significant reductions in PSQI, HAMD 17, and HAMA scores were observed in both groups after treatment: CBT-I group: PSQI ((14.15±2.54) vs. (7.50±3.35), t=13.25), HAMD 17 ((14.70±4.09) vs. (7.40±4.61), t=9.33), and HAMA ((14.94±4.11) vs. (5.56±3.67), t=12.38) (all P<0.001).CBT-I Plus group: PSQI ((14.87±3.01) vs. (7.19±3.86), t=18.75), HAMD 17 ((16.84±3.91) vs. (6.84±4.79), t=17.42), and HAMA ((15.57±3.93) vs. (6.10±4.57), t=18.39) (all P<0.001). After adjusting for HAMD 17 scores and medication use, no statistically significant between-group differences were observed in changes in PSQI, HAMD 17, and HAMA scores ( P>0.05). A significant time-by-group interaction was found for the PSQI daytime dysfunction subscale ( F=4.87, P<0.01). Conclusion:Both CBT-I and CBT-I Plus improve sleep and emotional symptoms in patients with chronic insomnia disorder and comorbid anxiety/depression symptoms. However, CBT-I Plus has no significant advantages over standard CBT-I. Further studies are needed to refine the timing and content of interventions.

This study was aimed at developing an in vitro fluorescent substrate assay for the activity of leucyl aminopeptid-ase(LAP)from Echinococcus multilocularis and comparing it with the chemical chromogenic substrate enzyme activity assay.Through the establishment of reaction conditions for the fluorescent substrate-based in vitro enzyme activity assay,we com-pared the differences between the fluorescent substrate L-Leucine-7-amido-4-methylocoumarin(Leu-AMC)and the chemical chromogenic substrate L-Leucine-4-nitroanilide(Leu-pNA)through molecular docking,inhibition rates,and precision measures.Molecular docking revealed that the fluorescent substrate Leu-AMC had higher affinity for the protein than the chemical chromogenic substrate Leu-pNA.Through analysis of the effects of varying reaction conditions on fluorescence intensi-ty,we optimized the fluorescent substrate enzyme activity assay to demonstrate favorable performance at a reaction temperature of 37℃,a pH of 9.0,a protein concentration of 800 nmol/L,and a reaction duration of 60 minutes.Leu-AMC exhibited significant and distinct responses at a 5 μmol/L substrate concentration,under varying substrate conditions.The fluo-rescent substrate assay demonstrated more significant intergroup differences than the chemical chromogenic substrate assay when various inhibitors were added.This study established a fluorescence-based enzyme activity assay for leucyl aminopeptidase from Echinococcus multilocularis by using Leu-AMC as the substrate;this method demonstrated a more significant intergroup difference and sensitivity than the chemical chromogenic substrate assay.

A mounting body of research suggests that circRNAs significantly contribute to the develop-ment of myocardial fibrosis.The microarray results of human circular RNA expression profile indicated that circHERC4_041 expression increased in the myocardium of patients with heart failure,RT-qPCR a-nalysis confirmed that the myocardial expression level of circHERC4_041 in individuals with heart failure were considerably elevated compared to that in healthy organ donors.Fluorescence in situ hybridization(FISH)confirmed that circHERC4_041 was abundant in the cytoplasm of human cardiomyocyte AC16.Overexpression of circHERC4_041 in mouse myocardial fibroblasts(mCFs)mediated by adenovirus in-hibited the expression of fibrosis-related proteins in mCFs.Experiments involving cell proliferation,wound healing,and Transwell assays demonstrated that overexpression of circHERC4_041 suppressed the growth and mobility of mCFs(P＜0.001).Sequence analysis results suggested that circHERC4_041 con-tains potential ribosome entry sequence(IRES)and open reading frame(ORF).Western blot confirmed that circHERC4_041 could translate the 516 amino acid HERC4-516aa protein,which was mainly located in the cytoplasm of the cell.Cell functional experiments confirmed that circHERC4_041 inhibited the fi-brotic phenotype of mCFs by specifically translating HERC4-516aa(P＜0.05).The specific interaction between HERC4-516aa and transglutaminase 2(TGM2)was confirmed by IP-MS screening and Co-IP i-dentification.Further results found that the degradation of TGM2 was promoted through proteasome path-way.The overexpression of TGM2 in mCFs facilitated by adenoviral vectors could counteract the suppres-sive effects of HERC4-516aa on the fibrotic phenotype of mCFs.Therefore,this study confirmed that the HERC4-516aa protein translated by circHERC4_041 can specifically bind to TGM2 to inhibit the fibrotic phenotype of myocardial fibroblasts.

OBJECTIVE To investigate the effect and potential mechanism of Ziyin Mingmu Formula in treating age-related mac-ular degeneration(AMD)by combining network pharmacology with animal model validation.METHODS Active ingredients of Ziyin Mingmu Formula were obtained from the TCMSP and BATMAN databases,and their targets were searched using the PharmMapper da-tabase.AMD disease targets were identified using the GeneCards,DrugBank,OMIM,and TTD databases.Venny analysis was per-formed to identify the intersection of active ingredient and disease targets.A protein-protein interaction(PPI)network was constructed using the String database,and core targets were screened using Cytoscape 3.9.0 software.KEGG pathway enrichment analysis was per-formed using the DAVID database.Molecular docking of key active ingredients with core targets was performed using Autodock software.A laser-induced mouse choroidal neovascularization(CNV)model was used.Optical coherence tomography angiography(OCTA)was used to assess CNV area in vivo,immunofluorescence staining was used to assess CNV area on choroidal flat mounts,and Western blot analysis was used to examine the expression of proteins involved in the vascular endothelial growth factor(VEGF)pathway.RESULTS Network pharmacology analysis identified 221 active ingredients in the Ziyin Mingmu Formula.PPI analysis i-dentified 29 core targets,including SRC,protein kinase B(AKT1),mitogen-activated protein kinase 1(MAPK1),and heat shock protein 90α family class A member 1(HSP90AA1).KEGG analysis revealed that the VEGF signaling pathway was the most highly en-riched.Molecular docking revealed that the core targets SRC,AKT1,MAPK1,and HSP90AA1 had good binding affinity with the main active ingredients,diosmetin,catechin,vestitol,and licochalcone A.Animal experiments showed that the Ziyin Mingmu Formula significantly reduced CNV area in model mice,downregulated VEGF protein expression,decreased VEGFR2,p38,and ERK1/2 pro-tein phosphorylation levels,and inhibited the VEGF signaling pathway.CONCLUSION Ziyin Mingmu Formula may inhibit CNV for-mation by regulating the VEGF signaling pathway.

Objective:To evaluate the role of hippocampal activating transcription factor 5 (ATF5) in cognitive impairment induced by neuropathic pain and the relationship with mitochondrial unfolded protein response(mtUPR) in mice.Methods:This study was conducted in 2 parts. Experiment Ⅰ Twenty-four SPF healthy male C57BL/6 mice, aged 6-8 weeks, weighing 20-25 g, were divided into 2 groups ( n=12 each) using a random number table method: sham operation group (S1 group) and neuropathic pain group (NP group). Neuropathic pain was induced by chronic constriction injury to the sciatic nerve. The mechanical paw withdrawal threshold (MWT) and thermal paw withdrawal latency (TWL) were measured before developing the model and at 7, 14, 21 and 28 days after developing the model. Mouse cognitive function was assessed using the novel object recognition test from 30-31 days after developing the model. After the end of the novel object recognition test, mice were sacrificed and the hippocampal CA1 region was harvested for determination of the expression of ATF5 (by Western blot) and the expression of ATF5 in neurons, microglia and astrocytes (by immunofluorescence double staining). Experiment Ⅱ Thirty-six SPF healthy male C57BL/6 mice, aged 6-8 weeks, weighing 20-25 g, were divided into 3 groups ( n=12 each) using a random number table method: sham operation group (S2 group), neuropathic pain + ATF5 up-regulation group (NA group), and neuropathic pain + empty virus group (NE group). On day 14 after developing the model, a virus that specifically up-regulated ATF5 expression in neurons and empty virus were injected into the hippocampal CA1 region. The MWT and TWL were measured at days 28 and 35 after developing the model. The novel object recognition test was performed on day 36 after developing the model to evaluate the cognitive function. After the end of the behavioral test, mice were sacrificed and the hippocampal CA1 region was harvested for detection of the expression of ATF5 and mtUPR marker proteins (Lon protease [LONP1] and heat shock protein 60 [HSP60]) by Western blot. Results:Experiment Ⅰ Compared with S1 group, no statistically significant change was found in the MWT and TWL before developing the model ( P>0.05), the MWT and TWL were significantly decreased on days 7, 14, 21 and 28 after developing the model, the discrimination index (DI) was decreased at day 31 after developing the model, the expression of ATF5 was down-regulated, the expression of ATF5 in neurons was down-regulated ( P<0.05), and no statistically significant change was found in the expression of ATF5 in mircrolia and astrocytes in NP group ( P>0.05). Experiment Ⅱ Compared with S2 group, the MWT and TWL were significantly decreased on days 28 and 35 after developing the model in NE group and NA group, DI was decreased, and the expression of ATF5, LONP1 and HSP60 was down-regulated in NE group ( P<0.05), and no significant change was found in NA group ( P>0.05). Compared with NE group, no significant change was found in the MWT and TWL in NA group ( P>0.05), DI was significantly increased, and the expression of ATF5, LONP1 and HSP60 was up-regulated in NA group ( P<0.05). Conclusions:Down-regulated ATF5 in the hippocampus is involved in the process of cognitive impairment caused by neuropathic pain, and the mechanism may be related to the inhibition of mtUPR.