Causal relationship between diabetes mellitus, glycemic traits and Parkinson's disease: a multivariable mendelian randomization analysis.

BACKGROUND: Observational studies have indicated an association between diabetes mellitus (DM), glycemic traits, and the occurrence of Parkinson's disease (PD). However, the complex interactions between these factors and the presence of a causal relationship remain unclear. Therefore, we aim to systematically assess the causal relationship between diabetes, glycemic traits, and PD onset, risk, and progression. METHOD: We used two-sample Mendelian randomization (MR) to investigate potential associations between diabetes, glycemic traits, and PD. We used summary statistics from genome-wide association studies (GWAS). In addition, we employed multivariable Mendelian randomization to evaluate the mediating effects of anti-diabetic medications on the relationship between diabetes, glycemic traits, and PD. To ensure the robustness of our findings, we performed a series of sensitivity analyses. RESULTS: In our univariable Mendelian randomization (MR) analysis, we found evidence of a causal relationship between genetic susceptibility to type 1 diabetes (T1DM) and a reduced risk of PD (OR = 0.9708; 95% CI: 0.9466, 0.9956; P = 0.0214). In our multivariable MR analysis, after considering the conditions of anti-diabetic drug use, this correlation disappeared with adjustment for potential mediators, including anti-diabetic medications, insulin use, and metformin use. CONCLUSION: Our MR study confirms a potential protective causal relationship between genetically predicted type 1 diabetes and reduced risk of PD, which may be mediated by factors related to anti-diabetic medications.

Ferroptosis and WDFY4 as novel targets for immunotherapy of lung adenocarcinoma.

BACKGROUND: Lung cancer exhibits the world's highest mortality rate among malignant cancers worldwide, thereby presenting a significant global challenge in terms of reducing patient mortality. In the field of oncology, targeted immunotherapy has emerged as a novel therapeutic approach for lung cancer. This study aims to explore potential targets for immunotherapy in lung adenocarcinoma (LUAD) through the analysis of Ferroptosis Index (FPI) and Single Cell RNA-Sequencing (scRNA-seq) data. The findings of this research can potentially offer valuable insights for improving LUAD immunotherapy strategies and informing clinical decision-making. METHODS: Firstly, the relationship between survival and ferroptosis in LUAD patients was analyzed by FPI. Subsequently, the association between ferroptosis and infiltration and regulation of immune cells was explored by immune infiltration analysis and correlation statistics. Lastly, the relationship between major infiltrating immune cell populations and related pathways and prognosis of LUAD patients was analyzed by GSEA and GSVA. To screen out core genes regulating infiltration of immune cell populations, scRNA-seq data of cancer and para-cancerous tissues of LUAD patients were downloaded, followed by cell clustering analysis, cell identification of core subpopulations, pseudotime analysis, single-cell GSVA and pathway enrichment analysis, and identification and functional analysis of core regulatory genes. Moreover, the expression levels of core functional genes in LUAD tissue microarray were detected by immunohistochemistry, and its relationship with the prognosis of LUAD patients was verified. Finally, we used lentivirus with WDFY4 to transfect LUAD A549 cells. CCK-8, flow cytometry apoptosis detection, Scratch wound healing assay, Transwell migration assay, Xenograft nude mice model, immunohistochemical analysis and other experimental methods were used to explore the biological effects of WDFY4 on LUAD in vitro and in vivo. RESULTS: Survival analysis of FPI values in LUAD patients revealed a positive correlation between smaller FPI values and longer overall survival. Immuno-infiltration analysis and its correlation with FPI values revealed that B cells were most strongly associated with ferroptosis. Ferroptosis of cancer cells could promote infiltration and activation of B cell populations, and LUAD patients with more infiltration of B cell populations had longer long-term survival. scRNA-seq data analysis indicated that the B cell population is one of the major cell populations infiltrated by immune cells in LUAD. During the later phases of B cell differentiation in LUAD, there was a decrease in the expression levels of ACAP1, LINC00926, TLR10, MS4A1, WDFY4, and TRIM22 genes, whereas the expression levels of TMEM59, TP53INP1, and METTL7A genes were elevated. The protein-protein interaction (PPI) network analysis indicated that WDFY4 plays a crucial role in regulating B cell differentiation in LUAD. Immunohistochemical analysis of LUAD tissue microarray revealed a significant downregulation of WDFY4 expression, which was closely related to the occurrence sites of LUAD. Moreover, LUAD patients with a low WDFY4 expression exhibited a poorer prognosis. Additionally, experimental findings demonstrated that the overexpression of WDFY4 could inhibit the proliferation and metastasis of A549 cells while promoting apoptosis. It was also confirmed that WDFY4 could inhibit cancer growth in vivo. CONCLUSIONS: The results indicate that promoting infiltration and activation of B cell populations could improve the long-term survival of LUAD patients, thereby offering a potential novel immunotherapeutic approach for LUAD. Besides, the promotion of cancer cell ferroptosis and upregulation of WDFY4 expression have been shown to induce the infiltration and activation of B cell populations. Furthermore, the overexpression of WDFY4 can significantly inhibit the growth of lung adenocarcinoma in vitro and in vivo, highlighting its potential as a target for immunotherapy in LUAD.

Curcumin alleviates 1-methyl- 4-phenyl- 1,2,3,6-tetrahydropyridine- induced Parkinson's disease in mice via modulating gut microbiota and short-chain fatty acids.

Background: The microbiota-gut-brain axis has been proposed as a potential therapeutic target of PD. The effects of curcumin against Parkinson's disease have been demonstrated; however, its neuroprotective mechanisms remain unknown. Our study investigated the potential mechanisms through which curcumin ameliorates Parkinson's disease via the microbiota-gut-brain axis. Methods: Mice were randomly divided into four groups: control, Curcumin, MPTP, and MPTP + Curcumin. Motor deficits and gastrointestinal dysfunction were assessed using behavioral test, intestinal motility test, and fecal parameter measurement. The loss of dopaminergic neurons and intestinal barrier function was measured using Western blot and immunofluorescence. Shotgun metagenomic sequencing and LC-MS were parallelly performed on mice feces to investigate alterations in microbiota and metabolites. Results: Curcumin alleviated motor deficits and the loss of dopaminergic neurons in MPTP-induced mice. Curcumin ameliorated gastrointestinal and intestinal barrier dysfunctions in MPTP-induced mice. Curcumin reduced gut microbial dysbiosis and modulated carbohydrate metabolism in MPTP-induced mice. Curcumin restored short-chain fatty acid (SCFA) profiles in MPTP-induced mice. Conclusion: Concurrently, these results indicate that curcumin inhibits Parkinson's disease by regulating the gut microbiota and short-chain fatty acids.

Genotype and clinical phenotype analysis of a Family with Kennedy disease.

To investigate the clinical phenotype-genotype correlations of a family with Kennedy disease (KD) and improve our understanding of the disease. KD was confirmed after clinical phenotypic analyses, laboratory tests, polymerase chain reaction assays for cytosine-adenine-guanine (CAG) repeats, and neuro-electrophysiological tests. The disease was assessed using the KD1234 scale and the spinal and bulbar muscular atrophy functional rating scale. The average age of disease onset was 30.8 ± 2.85 years. Clinically diagnosed members had 48 CAG repeats (≥35 is abnormal) in the androgen receptor gene. The patients exhibited gynecomastia and testicular dysfunction. The lesions mainly involved the medulla oblongata and spinal cord. Progesterone and serum creatine kinase levels were significantly high. Electromyography showed chronic neurogenic damage and abnormal sensory and motor conduction in family members who did not participate in sports, exercise, or physical hobbies. Our study showed that this family had a stable inheritance of CAG repeats, and the genotype was consistent with the clinical phenotype. Gynecomastia was the first symptom, with progressive androgen resistance resulting in testicular atrophy, infertility, and sexual dysfunction. Changes in serum creatine kinase may indicate the progression or relief of symptoms, and rehabilitation may delay the progression of muscle atrophy.

δ-Opioid Receptor Activation Inhibits Ferroptosis by Activating the Nrf2 Pathway in MPTP-Induced Parkinson Disease Models.

Introduction: Recent studies suggest the involvement of ferroptosis in the pathogenesis of Parkinson disease (PD). δ-Opioid receptors (DORs) have neuroprotective effects in PD. It is not known whether the neuroprotective effects of DORs in PD are attributable to the inhibition of ferroptosis. Therefore, we aimed to investigate the role of DORs in ferroptosis in MPTP-induced PD models. Methods: To identify the influence of DORs on ferroptosis in MPTP-induced PD models, we measured the malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE) levels, analyzed the levels of ferroptosis-related proteins (GXP4 and SLC7a11) and Nrf2 expression by using western blotting, and assessed mitochondrial dysfunction by using JC-1 staining and transmission electron microscopy. Results: DOR activation reduced the 4-HNE and MDA levels, increased the GXP4 and SLC7a11 levels, and ameliorated mitochondrial dysfunction in MPTP-induced PD models. These neuroprotective effects of DORs could be blocked by Nrf2-siRNA. Thus, the effects of DORs on ferroptosis in PD models were partially controlled by Nrf2, which regulated GXP4 and SLC7a11 synthesis. Conclusion: DORs exert neuroprotective effects in PD models by inhibiting ferroptosis partially via activating the Nrf2 pathway.

Transcriptomics combined with metabolomics analysis of the mechanism of agmatine in the treatment of septic liver injury.

Background: Acute liver injury can occur at any stage of sepsis and is an important sign of multiple organ dysfunction syndrome (MODS). Studies have shown that agmatine (AGM) can effectively improve liver injury caused by sepsis. However, due to the numerous metabolites and metabolic pathways of AGM in the human body, its mechanism in treating septic liver injury is unclear. Methods: In this study, a liver injury model of septic Sprague-Dawley rats was established by cecal ligation and perforation (CLP). After AGM treatment, transcriptomics combined with metabolomics was employed to analyze the gene expression levels and metabolite changes. Results: The results showed that AGM decreased the expression levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), procalcitonin (PCT), and inflammatory factors [interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß)] in the serum of septic rats. It also reduced liver inflammatory cell infiltration and abnormal lipid metabolism, and promoted the survival rate of septic rats. In addition, 17 differentially-expressed genes were identified by transcriptomics, mainly in arginine and proline metabolism, the arachidonic acid metabolism pathway, as well as the nuclear factor kappa B (NF-κB) and AMP-activated protein kinase (AMPK)-peroxisome proliferator-activated receptor α (PPARα) signal transduction pathways. Metabolomics analysis was carried out to study the potential liver metabolism spectrum changes induced by AGM treatment. The results showed significant changes in 26 metabolites in the rat liver samples, mainly involved in arginine and proline metabolism, arachidonic acid metabolism, linoleic acid metabolism, and fatty acid metabolism. Conclusions: The integrated transcriptomics and metabolomics analysis demonstrated that AGM improved septic liver injury by regulating lipid metabolism, and reduced the inflammatory reaction by affecting fatty acid metabolism, amino acid metabolism, and the arachidonic acid metabolism pathway. The integration of transcriptomics and metabolomics provides an effective means to elucidate AGM's therapeutic pathways and biomarkers.

A T-cell-dependent antibody response (TDAR) method in BALB/c mice based on a cytometric bead array.

Most current methods to assess T-cell-dependent antibody responses (TDAR) are semi-quantitative and based on measures of antibody titer generated against a standard antigen like keyhole limpet hemocyanin (KLH). The precision, sensitivity, and convenience of TDAR assays might be improved by applying rapid, sensitive, specific cytometric bead assays (CBA). In the study here, KLH antigen was covalently coupled onto the surface of cytometric beads using immune microsphere technology, and IgM antibody capture spheres were prepared for use in pretreatment processing of samples. The working parameters associated with this novel TDAR-CBA system were optimized in orthogonal experiments. The optimal concentration of the KLH coating solution in this system was 160 µg/ml, that of the anti-KLH IgG capture spheres 6.0 × 105/ml, and the optimal dilution of fluorescein isothiocyanate (FITC)-conjugated Affini-Pure Goat Anti-Mouse IgG (H + L) was 60 µg/ml. Repeated tests indicated that this approach yielded good linearity (r2 = 0.9937) method, with a within-run precision of 3.1-4.9%, and a between-run precision of 4.4-4.9%. This new approach had a limit of detection of 113.43 ng/ml (linear range = 390.63-50 000), and an interference rate of just 0.04-3.51%. Based on these findings, it seems that a new mouse TDAR assay based on CBA can be developed that would appear to be more sensitive, accurate, and precise than the current TDAR assay approaches based on traditional ELISA.

Chinese herbal component, Praeruptorin E, enhances anti-asthma efficacy and prevents toxicity of aminophylline by targeting the NF-κB/PXR/CYP3A4 pathway.

Background: Aminophylline is widely used for the treatment of asthma, but the therapeutic dose is very close to the toxic dose, which makes this drug prone to accumulation poisoning. In the present study, we explored whether the Chinese herbal component, Praeruptorin E (PE), enhances anti-asthma efficacy and prevents the toxicity of aminophylline. Methods: First, an ovalbumin (OVA)-induced mouse model of asthma, immunohistochemistry, pathological staining, and bronchoalveolar lavage fluid (BALF) were used to detect the lung condition of asthmatic mice. The content of Th2 cytokines in serum was measured by enzyme-linked immunosorbent assay (ELISA), and the expression of related proteins was detected by Western blotting and immunofluorescence. Concentrations of theophylline and its metabolites in rat serum were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). siRNA transfection and chromatin immunoprecipitation (ChIP) were used to investigate the mechanism of PE. Results: PE was found to synergize with aminophylline to reduce the infiltration of inflammatory cells, collagen deposition, and mucus hyperplasia in the lungs of asthmatic mice. It inhibited the expression of Th2 cytokines, interleukin (IL)-4, IL-5, and IL-13; promoted lung tissue repair; and reduced the toxic effect of aminophylline on the heart. Moreover, LC-MS/MS analysis showed that PE reduced the plasma concentration of the parent theophylline and its metabolite 1,3-dimethyluric acid (1,3-DMU). PE facilitated aminophylline's suppression of nuclear factor-κB (NF-κB), and increased the expression of the xenobiotic nuclear receptor pregnane X receptor (PXR) and its primary target gene, CYP3A11 [this is the mouse homolog of cytochrome P450 3A (CYP3A)] in the asthmatic mouse liver and in the L-02 human fetal hepatocyte cell culture model. In addition, the ChIP assay revealed that PE attenuated the binding of NF-κB to the promoter region of the PXR gene and prevented the suppression of PXR gene expression by NF-κB. Conclusions: PE has a dual function in enhancing the immune regulation and anti-inflammatory effects of theophylline, as well as preventing theophylline toxicity by targeting the NF-κB/PXR/CYP3A4 axis. PE is a promising herbal medicine that will benefit asthmatics taking theophylline.

The synergistic Reduning and cefmetazole sodium treatment of severe pneumonia is mediated by the AhR-Src-STAT3 pathway.

Background: Reduning (RDN) is a common Chinese medicine preparation with antibacterial, anti-inflammatory, antiviral and immunomodulatory effects in respiratory infectious diseases. Clinically, it is used in combination with antibiotics, but its synergistic effect and mechanism in treating severe pneumonia remain unclear. Methods: A rat model of severe pneumonia and an in vitro coculture model consisting of A549 and THP-1 cells were used to observe the synergistic effect of RDN on severe pneumonia. The inflammatory cytokines were tested by enzyme-linked immunosorbent assay (ELISA). The localization of Aryl hydrocarbon receptor (AhR) in A549 cells was observed by immunofluorescence, and the interaction of AhR and signal transducer and activator of transcription 3 (STAT3) proteins was observed by co-immunoprecipitation. AhR-Src tyrosine kinase (Src)-STAT3 pathway in rats and A549 cells were examined by Western Blot. Histopathological changes were observed by Hematoxylin-eosin (HE) staining, X-ray and survival rates were used to evaluate the effects of paclitaxel on severe pneumonia rats. Results: RDN regulation of Src-STAT3-interleukin 10 (IL-10) signaling pathway activation and macrophage polarization were mediated through the nuclear receptor AhR. The expression of AhR was significantly increased after RDN treatment, and this effect was accompanied by STAT3 expression increasing. Coimmunoprecipitation confirmed an interaction between AhR and STAT3 and upregulated IL-10 expression. Silencing AhR decreased Src, STAT3, and IL-10 expression. RDN activated AhR and increased Src, STAT3, and IL-10 expression. In addition, RDN regulated the polarization of macrophages RDN combined with cefmetazole sodium significantly reduced the pulmonary bacterial load, alleviated lung injury, and reduced o inflammatory factors expression, improving their survival. Conclusions: RDN can synergistically enhance the effect of cefmetazole sodium treatment in severe pneumonia, and the mechanism may involve increasing the expression level of IL-10 mediated through the AhR-Src-STAT3 pathway, driving the polarization of macrophages, and attenuating the cytokine storm to control inflammation in severe pneumonia.

Exploring the Neuroprotective Mechanism of Curcumin Inhibition of Intestinal Inflammation against Parkinson's Disease Based on the Gut-Brain Axis.

Parkinson's disease (PD) is a chronic progressive neurodegenerative disease commonly seen in aged people, in which gastrointestinal dysfunction is the most common nonmotor symptom and the activation of the gut-brain axis by intestinal inflammation may contribute to the pathogenesis of PD. In a previous study, curcumin was considered neuroprotective in PD, and this neuroprotective mechanism may act by inhibiting intestinal inflammation. Therefore, the aim of this study was to evaluate the effect of curcumin on motor dysfunction and the loss of dopaminergic neurons in a PD mouse model, induced by N-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) using open field test and pole test behavioral assessments and the immunofluorescence and Western blot methods. Moreover, the effects of curcumin on gastrointestinal dysfunction, gastric barrier function, pro-inflammatory cytokines, and the SIRT1/NRF2 pathway in intestinal tissues in a PD mouse model were assessed using fecal parameters and intestinal dynamics, immunofluorescence, ELISA, and Western blot. A motor impairment study of an MPTP-induced mouse group prior to treatment with curcumin had a lower total movement distance and a slow average speed, while there was no statistical difference in the curcumin group. After treatment with curcumin, the total movement distance and average speed improved, the tyrosine hydroxylase (TH) rate in the substantia nigra pars compacta (SNpc) and striatum were reduced, the pyroptosis of AIM2 and caspase-1 activations were inhibited, and intestinal inflammatory factors and intestinal inflammation were reduced. Curcumin improved gastrointestinal disorders and gastrointestinal barrier function in the MPTP-induced mice and reversed MPTP-induced motor dysfunction and dopaminergic neuron loss in mice. The above effects may be partly dependent on curcumin activation of the SIRT1/NRF2 pathway in the colon. This study provides a potential opportunity to develop new preventive measures and novel therapeutic approaches that could target the gut-brain axis in the context of PD and provide a new intervention in the treatment of Parkinson's disease.

Feng-Liao-Chang-Wei-Kang Combined with 5-Fluorouracil Synergistically Suppresses Colitis-Associated Colorectal Cancer via the IL-6/STAT3 Signalling Pathway.

BACKGROUND: Colitis-associated colorectal cancer (CAC) develops from active colonic inflammation, which is characterized by the production of proinflammatory cytokines that can induce mutations. IL-6 is produced by multiple cell types located within the tumor microenvironment including tumor-infiltrating immune cells, stromal cells, and the tumor cells themselves. The aim of our study was to explore the mechanism of Feng-Liao-Chang-Wei-Kang (FLCWK) and 5-fluorouracil (5-FU) in treating CAC. METHOD: HCT116 cells were treated with 5-FU in the absence or presence of FLCWK. Cell proliferation was assayed by MTT assays. Apoptosis and the cell cycle phases were detected by flow cytometry. Western blotting and Q-PCR assays were used to detect the expression levels of proteins and genes related to the IL-6/STAT3 signalling pathway. A mouse model for CAC was established by treating animals with 12.5 mg/kg azoxymethane (AOM) followed by 3 cycles of 2.5% dextran sodium sulphate (DSS). The associated pathological changes were determined after haematoxylin and eosin (H&E) staining. The expression of related proteins and genes in various tissues was examined using immunofluorescence techniques. RESULTS: FLCWK enhanced the ability of 5-FU to promote apoptosis by inhibiting the proliferation of HCT116 cells and blocking the IL-6/STAT3 pathway. FLCWK combined with 5-FU reduced the number and size of colon tumors in mice with CAC and significantly increased their survival rate. In the CAC model, FLCWK synergized with 5-FU to inhibit the phosphorylation of STAT3, preventing IL-6/STAT3 signal transduction and thus further inducing apoptosis and inhibition of colon cancer cell proliferation. CONCLUSION: FLCWK can inhibit the activation of STAT3 by reducing the production of IL-6, thereby increasing the occurrence of colitis-related colorectal cancer with 5-FU.