Squalene-epoxidase-catalyzed 24(S),25-epoxycholesterol synthesis promotes trained-immunity-mediated antitumor activity.

The importance of trained immunity in antitumor immunity has been increasingly recognized, but the underlying metabolic regulation mechanisms remain incompletely understood. In this study, we find that squalene epoxidase (SQLE), a key enzyme in cholesterol synthesis, is required for ß-glucan-induced trained immunity in macrophages and ensuing antitumor activity. Unexpectedly, the shunt pathway, but not the classical cholesterol synthesis pathway, catalyzed by SQLE, is required for trained immunity induction. Specifically, 24(S),25-epoxycholesterol (24(S),25-EC), the shunt pathway metabolite, activates liver X receptor and increases chromatin accessibility to evoke innate immune memory. Meanwhile, SQLE-induced reactive oxygen species accumulation stabilizes hypoxia-inducible factor 1α protein for metabolic switching into glycolysis. Hence, our findings identify 24(S),25-EC as a key metabolite for trained immunity and provide important insights into how SQLE regulates trained-immunity-mediated antitumor activity.

Successful treatment of a B/T MPAL patient by chemo-free treatment with venetoclax, azacitidine, and blinatumomab.

B/T mixed phenotype acute leukemia (MPAL), which represents only 2-3% of all MPAL cases, is classified as a high-risk leukemia subtype. Adults diagnosed with B/T MPAL have a notably low 3-year survival rate, estimated at 20-40%. The rarity and undercharacterization of B/T MPAL present substantial challenges in identifying an optimal treatment protocol. This report aims to shed light on this issue by presenting a case in which a patient with a complex karyotype was treated using a combination of venetoclax, azacitidine, and blinatumomab. This novel, chemo-free regimen resulted in the patient achieving both hematologic and molecular complete remission, with no severe organ or hematological toxicity observed. Notably, the patient continued to maintain molecular remission for 1 year following the transplantation. Based on these findings, the combination of venetoclax, azacitidine, and blinatumomab could be considered a potential therapeutic approach for B/T MPAL patients, meriting further investigation.

Disparate macrophage responses are linked to infection outcome of Hantan virus in humans or rodents.

Hantaan virus (HTNV) is asymptomatically carried by rodents, yet causes lethal hemorrhagic fever with renal syndrome in humans, the underlying mechanisms of which remain to be elucidated. Here, we show that differential macrophage responses may determine disparate infection outcomes. In mice, late-phase inactivation of inflammatory macrophage prevents cytokine storm syndrome that usually occurs in HTNV-infected patients. This is attained by elaborate crosstalk between Notch and NF-κB pathways. Mechanistically, Notch receptors activated by HTNV enhance NF-κB signaling by recruiting IKKß and p65, promoting inflammatory macrophage polarization in both species. However, in mice rather than humans, Notch-mediated inflammation is timely restrained by a series of murine-specific long noncoding RNAs transcribed by the Notch pathway in a negative feedback manner. Among them, the lnc-ip65 detaches p65 from the Notch receptor and inhibits p65 phosphorylation, rewiring macrophages from the pro-inflammation to the pro-resolution phenotype. Genetic ablation of lnc-ip65 leads to destructive HTNV infection in mice. Thus, our findings reveal an immune-braking function of murine noncoding RNAs, offering a special therapeutic strategy for HTNV infection.

Akkermansia muciniphila-derived small extracellular vesicles attenuate intestinal ischemia-reperfusion-induced postoperative cognitive dysfunction by suppressing microglia activation via the TLR2/4 signaling.

Akkermansia muciniphila (AKK) bacteria improve the functions of theere intestinal and blood-brain barriers (BBB) via their extracellular vesicles (AmEvs). However, their role in postoperative cognitive dysfunction (POCD) and its underlying mechanisms remain unclear. To investigate, we used C57BL/6 J mice divided into five groups: Sham, POCD, POCD+Akk, POCD+Evs, and POCD+Evs + PLX5622. POCD was induced through intestinal ischemia-reperfusion (I/R). The mice's cognitive function was assessed using behavioral tests, and possible mechanisms were explored by examining gut and BBB permeability, inflammation, and microglial function. Toll-like receptor (TLR) 2/4 pathway-related proteins were also investigated both in vitro and in vivo. PLX5622 chow was employed to eliminate microglial cells. Our findings revealed a negative correlation between AKK abundance and POCD symptoms. Supplementation with either AKK or AmEvs improved cognitive function, improved the performance of the intestinal barrier and BBB, and decreased inflammation and microglial activation in POCD mice compared to controls. Moreover, AmEvs treatment inhibited TLR2/4 signaling in the brains of POCD mice and LPS-treated microglial cells. In microglial-ablated POCD mice, however, AmEvs failed to protect BBB integrity. Overall, AmEvs is a potential therapeutic strategy for managing POCD by enhancing gut and BBB integrity and inhibiting microglial-mediated TLR2/4 signaling.

Insomnia in epilepsy is associated with nocturnal seizures and anxiety.

PURPOSE: The purpose of this study was to identify the factors associated with insomnia in patients with epilepsy (PWE) and provide evidence for clinical prevention and treatment. METHODS: PWE who visited our epilepsy clinic from December 2021 to December 2022 were enrolled in our study. All participants completed the Insomnia Severity Index (ISI), Epworth Sleepiness Scale (ESS), Self-rating Anxiety Scale (SAS), and Self-rating Depression Scale (SDS). Based on their ISI scores, they were categorized into two groups: PWE with insomnia (ISI score ≥ 10) and PWE without insomnia (ISI score < 10). Univariate analysis and stepwise logistic regression analysis were conducted to identify the factors associated with insomnia in PWE. RESULTS: A total of 196 Chinese PWE were recruited in this study(men, 39.8 %). Of these, 39 PWE(19.9 %) had insomnia.The incidence of nocturnal seizures (43.6 %vs19.7 %), depression (46.2 %vs9.6 %), anxiety (59.0 %vs11.5 %), and excessive daytime sleepiness(EDS,28.2 %vs5.7 %) in PWE with insomnia were significantly higher than in those without insomnia(all p＜0.01). Univariate regression analysis showed that seizures greater than or equal to once per month, nocturnal seizures, anxiety, depression, and EDS may associate with insomnia in PWE(all p＜0.05). Stepwise logistic regression analysis demonstrated that nocturnal seizures (OR = 2.611,95 % CI 1.040-6.478, P = 0.038) and anxiety (mild OR = 4.830,95 %CI 1.741-13.186, P = 0.002;moderate OR = 24.239,95 %CI 4.719-183.935, P＜0.001; severe OR = 37.653,95 %CI 4.931-782.741, P = 0.002) were independently associated with insomnia in PWE. CONCLUSION: PWE with insomnia are more likely to experience depression and EDS. Nocturnal seizures and anxiety are identified as independent factors associated with insomnia in PWE. Furthermore, Anxiety has a greater impact on insomnia in PWE and the likelihood of insomnia has increased significantly with the aggravation of anxiety levels.

The Associations of Maternal Blood Hemoglobin and Serum Triglyceride Levels and the Risk of Preterm Delivery.

The abnormal hemoglobin (HGB) and serum lipid concentrations during pregnancy will increase the risk of preterm delivery. Our study aimed to explore the correlation between prenatal HGB and serum lipid levels and preterm delivery. We enrolled 215 mother-infant pairs in a pilot cohort study. The logistic regression model and Restricted Cubic Spline model (RCS) were used to investigate the levels of prenatal blood HGB and serum lipid such as triglyceride (TG), total cholesterol, high-density lipoprotein, low density lipoprotein and preterm delivery. The results showed that moderate levels of prenatal blood HGB (OR=0.28; 95%CI: 0.10, 0.75, p-trend=0.018) and high level of serum TG (OR=0.29; 95%CI: 0.10, 0.84, p-trend=0.022) level were negatively associated with the risk of preterm delivery. The joint effect results showed that compared with lower level of prenatal blood HGB (≤123.13 g/l) and TG (≤3.7 mmol/l), we found that high levels prenatal blood HGB and serum TG (OR=0.32, 95%CI: 0.12, 0.89) had a negative association with the risk of preterm delivery. Moreover, prenatal blood HGB and serum TG levels had negative linear dose-effect relationships with the risk of preterm delivery in overall and girl group (p<0.05). Moderate levels of prenatal blood HGB and high level of serum TG were negatively associated with the risk of preterm delivery. The joint effect of high levels prenatal HGB and prenatal serum TG in the normal range were negatively correlated with preterm delivery. Moreover, the underlying mechanisms should be clarified in future studies.

Functional network connectivity patterns predicting the efficacy of repetitive transcranial magnetic stimulation in the spectrum of Alzheimer's disease.

BACKGROUND: Neuro-navigated repetitive transcranial magnetic stimulation (rTMS) is potentially effective in enhancing cognitive performance in the spectrum of Alzheimer's disease (AD). We explored the effect of rTMS-induced network reorganization and its predictive value for individual treatment response. METHODS: Sixty-two amnestic mild cognitive impairment (aMCI) and AD patients were recruited. These subjects were assigned to multimodal magnetic resonance imaging scanning before and after a 4-week stimulation. Then, we investigated the neural mechanism underlying rTMS treatment based on static functional network connectivity (sFNC) and dynamic functional network connectivity (dFNC) analyses. Finally, the support vector regression was used to predict the individual rTMS treatment response through these functional features at baseline. RESULTS: We found that rTMS at the left angular gyrus significantly induced cognitive improvement in multiple cognitive domains. Participants after rTMS treatment exhibited significantly the increased sFNC between the right frontoparietal network (rFPN) and left frontoparietal network (lFPN) and decreased sFNC between posterior visual network and medial visual network. We revealed remarkable dFNC characteristics of brain connectivity, which was increased mainly in higher-order cognitive networks and decreased in primary networks or between primary networks and higher-order cognitive networks. dFNC characteristics in state 1 and state 4 could further predict individual higher memory improvement after rTMS treatment (state 1, R = 0.58; state 4, R = 0.54). CONCLUSION: Our findings highlight that neuro-navigated rTMS could suppress primary network connections to compensate for higher-order cognitive networks. Crucially, dynamic regulation of brain networks at baseline may serve as an individualized predictor of rTMS treatment response. RELEVANCE STATEMENT: Dynamic reorganization of brain networks could predict the efficacy of repetitive transcranial magnetic stimulation in the spectrum of Alzheimer's disease. KEY POINTS: • rTMS at the left angular gyrus could induce cognitive improvement. • rTMS could suppress primary network connections to compensate for higher-order networks. • Dynamic reorganization of brain networks could predict individual treatment response to rTMS.

Premature mortality risk in individuals with convulsive epilepsy: Results from a longitudinal, prospective, population-based study.

OBJECTIVE: To assess premature mortality and identify associated risk factors among individuals with convulsive epilepsy in resource-poor settings using a longitudinal, prospective, population-based approach. METHOD: The study recruited people with convulsive epilepsy who underwent assessment and management of epilepsy at primary healthcare centers in rural Northwest China, including newly diagnosed individuals and previously identified prevalent cases. All participants were confirmed to have epilepsy by neurologists according to strict criteria and were followed up monthly by primary care physicians. Demographic data and cause of death (COD) were obtained from death certificates or verbal autopsies conducted by neurologists, following the International Classification of Diseases, 10th Edition. The standardized mortality ratio (SMR) and proportionate mortality ratio (PMR) for each cause of death were estimated using the Cause-Of-Death Surveillance Dataset of China (2020). Survival analysis was used to identify risk factors associated with all-cause mortality and death directly due to epilepsy. RESULTS: During 5.9 years of follow-up with 40,947 person-years, there were 781 (11.2%) deaths among 6967 participants. The risk of premature death in people with convulsive epilepsy was 2.7-fold higher than that in the general population. Young participants had a significantly higher risk (standardized mortality ratio 26.5-52.5) of premature death. The proportionate mortality ratio was higher for cerebrovascular disease (15%), sudden unexpected death in epilepsy (SUDEP) (13.4%), cardiovascular disease (11.7%), status epilepsy (SE) (11.3%), and epilepsy-related accidents (14.0%) than other premature mortality cause of deaths. Additionally, the highest standardized proportional mortality ratio (SPMR) was observed from drowning in all cause of death (10.4, 95% confidence interval [CI]: 7.6-13.8), followed by burning (9.0, 95% CI: 3.7-18.9). Factors that increased the risk of all-cause mortality included male sex, late age of onset, short disease duration, high body mass index, monotherapy, and the frequency of generalized tonic-clonic seizures (GTCS). High frequency of generalized tonic-clonic seizures (> 3 attacks in the last year) was an independent risk factor for premature death directly due to epilepsy (including sudden unexpected death in epilepsy, status epilepsy, and epilepsy-related accidents), while early age of onset (≤ 14 years) and long duration of epilepsy (> 20 years) were independent risk factors for sudden unexpected death in epilepsy. In addition, short duration of epilepsy (≤ 20 years) was an independent risk factor for status epilepsy. CONCLUSIONS: This study demonstrated that individuals with poorly controlled seizures are more likely to experience premature death, with most deaths being epilepsy-related and preventable. These findings underline the importance of effective seizure treatment and the potential impact on reducing premature mortality among people with convulsive epilepsy.

Derivation and validation of a new prediction model for sudden unexpected death in epilepsy based on a longitudinal prospective population-based cohort.

OBJECTIVE: We conducted a population-based, prospective cohort study with a large sample size in Ningxia Province of the Northwest, a rural area in China, by developing a model to specifically assess risk factors of sudden unexpected death in epilepsy (SUDEP) in people with convulsive epilepsy by clinical variables. METHODS: Participants with convulsive epilepsy were recruited from January 1, 2008, to April 28, 2022, in rural Northwest China. They received regular assessments and management of epilepsy at the primary healthcare level and were followed up monthly. Information on the cause of death and relevant clinical details was obtained from death certificates or neurologist-conducted verbal autopsies. Survival analysis was employed to identify potential risk factors associated with SUDEP. RESULTS: Five variables were independently associated with SUDEP: generalized tonic-clonic seizures (GTCS) with ≥1 attack during the preceding month, GTCS with >3 attacks during the preceding year, body mass index (BMI) ≥24, age of onset ≤14 years, and duration >20 years. The area under receiver operator characteristic (ROC) curve (AUC) value (95% CI) of the model was 0.789 (0.735-0.843) in the derivation dataset and 0.830 (0.758-0.902) in the validation dataset. There was agreement between the observed and predicted probabilities of SUDEP. CONCLUSIONS: This study establishes that high GTCS frequency, early age of onset, long duration of epilepsy, and being overweight are associated with an increased risk of SUDEP in individuals with convulsive epilepsy. The study also developed and validated a personalized prediction model to accurately assess the risk of SUDEP.

Establishment and characterization of an immortalized bovine intestinal epithelial cell line.

Primary bovine intestinal epithelial cells (PBIECs) are an important model for studying the molecular and pathogenic mechanisms of diseases affecting the bovine intestine. It is difficult to obtain and grow PBIECs stably, and their short lifespan greatly limits their application. Therefore, the purpose of this study was to create a cell line for exploring the mechanisms of pathogen infection in bovine intestinal epithelial cells in vitro. We isolated and cultured PBIECs and established an immortalized BIEC line by transfecting PBIECs with the pCI-neo-hTERT (human telomerase reverse transcriptase) recombinant plasmid. The immortalized cell line (BIECs-21) retained structure and function similar to that of the PBIECs. The marker proteins characteristic of epithelial cells, cytokeratin 18, occludin, zonula occludens protein 1 (ZO-1), E-cadherin and enterokinase, were all positive in the immortalized cell line, and the cell structure, growth rate, karyotype, serum dependence and contact inhibition were normal. The hTERT gene was successfully transferred into BIECs-21 where it remained stable and was highly expressed. The transport of short-chain fatty acids and glucose uptake by the BIECs-21 was consistent with PBIECs, and we showed that they could be infected with the intestinal parasite, Neospora caninum. The immortalized BIECs-21, which have exceeded 80 passages, were structurally and functionally similar to the primary BIECs and thus provide a valuable research tool for investigating the mechanism of pathogen infection of the bovine intestinal epithelium in vitro.

In dairy cattle, the intestine is essential for productivity as it contributes nearly 10% of the total metabolizable energy. The intestinal epithelium is at risk of infection from constant exposure to pathogenic microorganisms, which seriously endangers an animal's health, but no bovine intestinal epithelial cell line has been developed so far for research on intestine -related diseases. Thus, the goal of this study was to create an immortalized cell line from isolated primary bovine intestinal epithelial cells. The expression of an exogenous human telomerase reverse transcriptase (hTERT) gene can circumvent the Hayflick limit by maintaining telomere integrity and we used transfection with a plasmid expressing the hTERT gene to convert primary intestinal epithelial cells into an immortalized cell line, which we then characterized. The results showed that the immortalized cell line (BIECs-21) was structurally and functionally similar to the primary bovine intestinal epithelial cells (BIECs) and thus provided a valuable research tool for investigating the mechanism of pathogen infection of the bovine intestinal epithelium in vitro.

Structural and biochemical characterization of Schlafen11 N-terminal domain.

Schlafen11 (SLFN11) is one of the most studied Schlafen proteins that plays vital roles in cancer therapy and virus-host interactions. Herein, we determined the crystal structure of the Sus scrofa SLFN11 N-terminal domain (NTD) to 2.69 Å resolution. sSLFN11-NTD is a pincer-shaped molecule that shares an overall fold with other SLFN-NTDs but exhibits distinct biochemical characteristics. sSLFN11-NTD is a potent RNase cleaving type I and II tRNAs and rRNAs, and with preference to type II tRNAs. Consistent with the codon usage-based translation suppression activity of SLFN11, sSLFN11-NTD cleaves synonymous serine and leucine tRNAs with different efficiencies in vitro. Mutational analysis revealed key determinates of sSLFN11-NTD nucleolytic activity, including the Connection-loop, active site, and key residues essential for substrate recognition, among which E42 constrains sSLFN11-NTD RNase activity, and all nonconservative mutations of E42 stimulated RNase activities. sSLFN11 inhibited the translation of proteins with a low codon adaptation index in cells, which mainly dependent on the RNase activity of the NTD because E42A enhanced the inhibitory effect, but E209A abolished inhibition. Our findings provide structural characterization of an important SLFN11 protein and expand our understanding of the Schlafen family.

Ramelteon improves blood-brain barrier of focal cerebral ischemia rats to prevent post-stroke depression via upregulating occludin.

Post-stroke depression (PSD) negatively affects the prognosis of post-stroke animals. Ramelteon has neuroprotection for chronic ischemia animals, but the effect and the biological mechanism of it on PSD is still unclear. This study explored the effects of ramelteon with prophylactic administration on blood-brain barrier in rats with middle cerebral artery occlusion (MCAO) and the oxygen-glucose deprivation/reperfusion (OGD/R) bEnd.3 cells and found that ramelteon pretreatment improved the depressive-like behaviors and decreased infarct area in MCAO rats. Also, this study found ramelteon pretreatment improved viability and inhibited permeability in OGD/R cells. In addition, this study found that MCP-1, TNF-α, and IL-1 levels were raised in the MCAO rats and that occludin protein and mRNA levels were decreased in the MCAO and the OGD/R models, while the Egr-1 level was up-regulated. All of these were antagonized by ramelteon pretreatment. In addition, overexpression of Egr-1 could reverse the effect of 100 nM ramelteon pretreatment on FITC and occludin levels in OGD/R cells. In short, this study has demonstrated that the protective effect on PSD of ramelteon pretreatment on MCAO rats is related to the development of BBB permeability and that ramelteon regulates occludin to protect the BBB by inhibiting Egr-1.

A Nanovaccine Based on Adjuvant Peptide FK-13 and l-Phenylalanine Poly(ester amide) Enhances CD8+ T Cell-Mediated Antitumor Immunity.

Cancer vaccines have shown promise as effective means of antitumor immunotherapy by inducing tumor antigen-specific T cell immunity. In this study, a novel peptide-based tumor nanovaccine that boosts antigen presentation and elicits effective antitumor immunity is developed. The adjuvant characteristics of an antimicrobial peptide-derived core peptide, FK-13, are investigated and used it to generate a fusion peptide named FK-33 with tumor antigen epitopes. l-phenylalanine-based poly(ester amide) (Phe-PEA), 8p4, is also identified as a competent delivery vehicle for the fusion peptide FK-33. Notably, the vaccination of 8p4 + FK-33 nanoparticles (8FNs) in vivo induces dendritic cell activation in the lymph nodes and elicits robust tumor antigen-specific CD8+ T cell response. The nanovaccine 8FNs demonstrate significant therapeutic and prophylactic efficacy against in situ tumor growth, effectively inhibit tumor metastasis, and significantly prolong the survival of tumor-bearing mice. Moreover, 8FNs can incorporate different tumor antigens and exhibit a synergistic therapeutic effect with antiprogrammed cell death protein 1 (PD-1) therapy. In summary, 8FNs represent a promising platform for personalized cancer vaccines and may serve as a potential combinational modality to improve current immunotherapy.

Goats with low levels of AAV antibody may serve as candidates for large animal gene therapy.

AAV vector-mediated gene therapy has been proposed as a feasible strategy for several eye diseases. However, AAV antibodies in the serum prior to treatment hinder the transduction efficiency and thus the therapeutic effect. Therefore, it is necessary to evaluate AAV antibodies in the serum before gene therapy. As large animals, goats are more closely related to humans than rodents and more economically available than nonhuman primates. Here, we first evaluated the AAV2 antibody serum level in rhesus monkeys before AAV injection. Then, we optimized a cell-based neutralizing antibody assay for detecting AAV antibodies in the serum of Saanen goats and evaluated the consistency of the cell-based neutralizing antibody assay and ELISA for goat serum antibody evaluation. The cell-based neutralizing antibody assay showed that the percentage of macaques with low antibody levels was 42.86%; however, there were no macaques with low antibody levels when the serum was evaluated by ELISA. The proportion of goats with low antibody levels was 56.67% according to the neutralizing antibody assay and 33. 33% according to the ELISA, and McNemar's test showed that the results of the two assays were not significantly different (P = 0.754), but that their consistency is poor (Kappa = 0.286, P = 0.114). Moreover, longitudinal evaluation of serum antibodies before and after intravitreal injection of AAV2 in goats revealed that the level of AAV antibodies increased and transduction inhibition subsequently increased, as reported in humans, indicating that transduction inhibition should be taken into account at different stages of gene therapy. In summary, starting with an evaluation of monkey serum antibodies, we optimized a detection method of goat serum antibodies, providing an alternative large animal model for gene therapy, and our serum antibody measurement method may be applied to other large animals.

Taurine alleviates oxidative stress in porcine mammary epithelial cells by stimulating the Nrf2-MAPK signaling pathway.

The high incidence of oxidative stress in sows during late gestation and lactation affects mammary gland health, milk yield, and milk quality. Recently, we found that supplementing maternal diets with 1% taurine improved antioxidant capability and enhanced growth performance in offspring; however, the mechanisms underlying these are unknown. This study aimed to investigate the cytoprotective effects and the mechanism of taurine in mitigating oxidative stress in porcine mammary epithelial cells (PMECs). PMECs were pretreated with 0-2.0 mM taurine for 12 h and then subjected to oxidative injury with 500 µM hydrogen peroxide (H2O2). Pretreatment with taurine attenuated decreased cell viability, enhanced superoxide dismutase, and reduced the intracellular reactive oxygen species accumulation after H2O2 exposure. Taurine also prevented H2O2-induced endoplasmic reticulum stress. Nuclear factor erythroid 2-related factor 2 (Nrf2) was essential to the cytoprotective effects of taurine on PMECs, as Nrf2 knockdown significantly inhibited taurine-induced cytoprotection against oxidative stress. Moreover, we confirmed that Nrf2 induction by taurine was mediated through the inactivation of the p38/MAPK pathway. Overall, taurine supplementation has beneficial effects on redox balance regulation and may protect against oxidative stress in lactating animals.

Effects of Combined Intervention of rTMS and Neurotransmitter Drugs on the Brain Functional Networks in Patients with Cognitive Impairment.

Alzheimer's disease (AD) causes extensive neural network dysfunction. Memantine and donepezil are commonly used as monotherapy or in combination with non-drug interventions, such as repetitive transcranial magnetic stimulation (rTMS), for its treatment. However, no studies have reported any differences between the effects of combined neurotransmitter and rTMS interventions versus rTMS alone on the brain networks of patients with cognitive impairment. Therefore, it is crucial to explore the advantages of different intervention methods to guide clinical practice. We used resting-state functional magnetic resonance imaging (rs-fMRI) to investigate the impact of neurotransmitter superimposed rTMS and rTMS alone on the brain functional network of patients with cognitive impairment. We divided patients with cognitive impairment who had received rTMS into two groups based on whether they received neurotransmitters: the combined intervention group and the rTMS-alone intervention group. We conducted rs-fMRI scans and comprehensively assessed cognitive function in these patients. To examine the effects of the superimposed interventions, we utilized independent component analysis to evaluate the functional connectivity of brain networks in these patients. Compared to the rTMS-alone intervention group, co-intervention of neurotransmitter drugs and rTMS exhibited potential for cognitive enhancement via the reconstructed inter-network connectivity of the cerebellum and the enhanced intra-network connectivity of the frontal-parietal regions in these patients with cognitive impairment. We hypothesized that the combination of neurotransmitter drugs and rTMS intervention could have greater clinical benefits than rTMS intervention alone, leading to improved cognitive function in patients with cognitive impairment.

Molecular mechanism of valine and its metabolite in improving triglyceride synthesis of porcine intestinal epithelial cells.

An insufficient energy supply to intestinal epithelial cells decreases production performance in weaned piglets. Triglycerides are the main energy source for intestinal epithelial cells in piglets. The present study aimed to investigate the effects and mechanisms of valine supplementation on triglyceride synthesis in porcine intestinal epithelial (IPEC-J2) cells. Valine supplementation in the medium significantly increased the content of triglycerides, fat droplets, and long-chain fatty acids (C17:0, C18:0, C20:0, C18:1, C20:1, and C22:1) (P < 0.05). Valine metabolite (3-hydroxyisobutyrate [3-HIB]) concentration increased significantly in the valine-supplemented group (P < 0.05). Silencing of the 3-HIB synthase enzyme 3-hydroxyisobutyryl-CoA hydrolase (HIBCH) in IPEC-J2 cells significantly reduced the triglyceride concentration and lipid droplet synthesis. Further studies found that 3-HIB supplementation in the medium significantly increased the concentration of triglycerides, lipid droplets, and unsaturated fatty acids (C16:1, C18:1, C18:2, C18:3, C20:3, C20:4, and C20:5) (P < 0.05) by upregulating the expression of proteins involved in fatty acid transport (CD36) and fatty acid binding protein 3 (FABP3) or triglyceride synthesis (DGAT1) (P < 0.05), indicating that 3-HIB mediates valine-enhanced triglyceride synthesis in IPEC-J2 cells. In conclusion, our results demonstrated that valine enhanced triglyceride synthesis in IPEC-J2 cells via increasing the 3-HIB concentration, which may promote fatty acid transport via upregulation of proteins related to fatty acid transporter. These findings provide new insights into the mechanisms through which valine participates in lipid metabolism.

Selective activation of cholinergic neurotransmission from the medial septal nucleus to hippocampal pyramidal neurones improves sepsis-induced cognitive deficits in mice.

BACKGROUND: Sepsis-associated encephalopathy is characterised by cognitive dysfunction, and might be mediated by deficits in neurotransmission. Reduced cholinergic neurotransmission in the hippocampus impairs memory function. We assessed real-time alterations of acetylcholine neurotransmission from the medial septal nucleus to the hippocampus, and explored whether sepsis-induced cognitive deficits can be relieved by activating upstream cholinergic projections. METHOD: Lipopolysaccharide (LPS) injection or caecal ligation and puncture (CLP) was used to induce sepsis and associated neuroinflammation in wild-type and mutant mice. Adeno-associated viruses for calcium and acetylcholine imaging, and for optogenetic and chemogenetic modulation of cholinergic neurones were injected into the hippocampus or medial septum, and a 200-µm-diameter optical fibre was implanted to collect acetylcholine and calcium signals. Cholinergic activity of the medial septum was manipulated and combined with cognitive assessment after LPS injection or CLP. RESULTS: Intracerebroventricular LPS injection reduced postsynaptic acetylcholine (from 0.146 [0.001] to 0.0047 [0.0005]; p=0.004) and calcium (from 0.0236 [0.0075] to 0.0054 [0.0026]; p=0.0388) signals in hippocampal Vglut2-positive glutamatergic neurones, whereas optogenetic activation of cholinergic neurones in the medial septum reversed LPS-induced reductions in these two signals. Intraperitoneal LPS injection decreased acetylcholine concentration in the hippocampus (476 [20] pg ml-1 to 382 [14] pg ml-1; p=0.0001). Reduction in long-term potentiation (238 [23] % to 150 [12] %; p=0.0082) and enhancement of hippocampal pyramidal neurone action potential frequency (5.8 [1.5] Hz to 8.2 [1.8] Hz; p=0.0343) were relieved, and neurocognitive performance was improved by chemogenetic activation of cholinergic innervation of the hippocampus 3 days after LPS injection in septic mice. CONCLUSIONS: Systemic or local LPS reduced cholinergic neurotransmission from the medial septum to hippocampal pyramidal neurones, and their selective activation alleviated defects in hippocampal neuronal function and synaptic plasticity and ameliorated memory deficits in sepsis model mice through enhanced cholinergic neurotransmission. This provides a basis for targeting cholinergic signalling to the hippocampus in sepsis-induced encephalopathy.

Vitrectomy and ILM peeling in rhesus macaque: pitfalls and tips for success.

BACKGROUND: The non-human primate (NHP) model is ideal for pre-clinical testing of novel therapies for human retinal diseases due to its similarity to the human visual system. However, intra-ocular delivery of gene therapy or cell transplantation to the retina gets hampered by the sticky vitreous body and poorly permeable inner limiting membrane (ILM) in primates. Although vitrectomy and ILM peeling are commonly performed in patients, many pitfalls exist in carrying out these procedures in the rhesus macaque, which have not been reported previously. METHODS: We summarised common surgical pitfalls after performing vitrectomy and ILM peeling in four eyes of two rhesus macaques (one male and one female). We provided corresponding hands-on technical tips based on our surgical experience and literature search. Orbital CT scans were compared between adult rhesus macaques and humans. High-resolution surgical videos were recorded to demonstrate each critical surgical step. RESULTS: Due to size difference, poor post-operative compliance, and high-standard requirements of a controlled experiment, there were eleven common surgical pitfalls during vitrectomy and ILM peeling in rhesus macaque. Falling into these pitfalls may produce discomfort, add fatigue, cause surgical complications, or even lead to the exclusion of the NHP from an experimental group. CONCLUSION: Recognition and circumvention of these pitfalls during vitrectomy and ILM peeling in NHP are essential. By focusing on these surgical pitfalls, we can better carry out preclinical tests of novel therapies for retinal diseases in the NHP model.