Association of GAL-8 promoter methylation levels with coronary plaque inflammation.

BACKGROUND AND AIMS: Coronary heart disease (CHD) is a condition that carries a high risk of mortality and is associated with aging. CHD is characterized by the chronic inflammatory response of the coronary intima. Recent studies have shown that the methylation level of blood mononuclear cell DNA is closely associated with adverse events in CHD, but the roles and mechanisms of DNA methylation in CHD remain elusive. METHODS AND RESULTS: In this study, the DNA methylation status within the epigenome of human coronary tissue in the sudden coronary death (SCD) group and control (CON) group of coronary heart disease was analyzed using the Illumina® Infinium Methylation EPIC BeadChip (850 K chip), resulting in the identification of a total of 2553 differentially methylated genes (DMGs). The differentially methylated genes were then subjected to Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis, and significant differential DNA methylation was found. Among the differentially hypomethylated genes were GAL-8, LTF, and RFPL3, while the highly methylated genes were TMEM9B, ANK3, and C6orF48. These genes were mainly enriched in 10 significantly enriched pathways, such as cell adhesion junctions, among which the differentially methylated gene GAL-8 was involved in inflammatory pathway signaling. For functional analysis of GAL-8, we first examined the differences in GAL-8 promoter methylation levels among different subgroups of human coronary tissue in the CON, CHD, and SCD groups using pyrophosphate sequencing. The results revealed reduced GAL-8 promoter methylation levels in the SCD group, while the difference between the CHD and CON groups was not statistically significant (P > 0.05). The reduced GAL-8 promoter methylation level was associated with upregulated GAL-8 expression, which led to increased expression of the inflammatory markers TNF-α, IL-1ß, MCP-1, MIP-2, MMP-2, and MMP-9. This enhanced inflammatory response contributed to the accumulation of foam cells, thickening of the intima of human coronary arteries, and increased luminal stenosis, which promoted the occurrence of sudden coronary death. Next, we found that GAL-8 promoter methylation levels in PBMC were consistent with human coronary tissue. The unstable angina group (UAP) had significantly lower GAL-8 promoter methylation levels than stable angina (SAP) and healthy controls (CON) (P < 0.05), and there was a significant correlation between reduced GAL-8 promoter methylation levels and risk factors for coronary heart disease. These findings highlight the association between decreased GAL-8 promoter methylation and the presence of coronary heart disease risk factors. ROC curve analysis suggests that methylation of the GAL 8 promoter region is an independent risk factor for CHD. In conclusion, our study confirmed differential expression of GAL-8, LTF, MUC4D, TMEM9B, MYOM2, and ANK3 genes due to DNA methylation in the SCD group. We also established the consistency of GAL-8 promoter methylation alterations between human coronary tissue and patient peripheral blood monocytes. The decreased methylation level of the GAL-8 promoter may be related to the increased expression of GAL-8 and the coronary risk factors. CONCLUSIONS: Accordingly, we hypothesized that reduced levels of GAL-8 promoter methylation may be an independent risk factor for adverse events in coronary heart disease.

Blockade of CXCR4 promotes macrophage autophagy through the PI3K/AKT/mTOR pathway to alleviate coronary heart disease.

OBJECTIVE: Autophagy is important in regulating inflammation and cholesterol efflux, suggesting that targeting autophagy may slow down atherosclerosis (AS). Since the pathological basis of coronary artery disease (CAD) is atherosclerosis, it is crucial to investigate the role of autophagy in atherosclerosis. This study aimed to investigate the role of the chemokine CXC chemokine receptor 4 (CXCR4) in promoting macrophage autophagy through the phosphoinositide-3 kinase/protein kinase B/mammalian target of rapamycin (PI3K/AKT/mTOR) pathway to alleviate coronary artery disease. METHODS: The human left coronary artery and myocardium were collected to detect CXCR4, MAP1LC3(LC3) and SQSTM1(p62) expression. ApoE-/- mice were used to establish an atherosclerosis mice model, while human monocytes (THP-1) were used to establish a foam cell model and co-cultured with foam cells using siRNACXCR4. Western blotting was conducted to quantify CXCR4, PI3K/AKT/mTOR pathway protein, LC3, Beclin1 and p62 protein levels. The left coronary artery from humans and mouse aorta and myocardium were stained with Hematoxylin and Eosin (H&E), macrophages with Oil Red O staining and foam cells were assessed by Movat's staining. CXCR4 levels, PI3K/AKT/mTOR pathway protein, LC3 and p62 were detected by immunohistochemistry (IHC) and immunofluorescence assays. Detection of autophagosomes in macrophages using transmission electron microscopy. We further assessed whether the effect of CXCR4-mediated macrophage autophagy on the formation of atherosclerosis and structural changes in the myocardium was mediated via the PI3K/AKT/mTOR signaling pathway. RESULTS: CXCR4 and p62 proteins were upregulated in human coronary lesions, mouse aorta, myocardial tissue, and foam cells, while LC3II/LC3I was downregulated. p85 (P-PI3K), Ser473 (P-AKT), and Ser2448 (P-mTOR) phosphorylated proteins associated with the PI3K/AKT/mTOR pathway were detected in AS and foam cell models. Upregulated CXCR4 inhibited autophagy of macrophages and increased the severity of atherosclerotic lesions. After specific knockdown of CXCR4 by adeno-associated virus (AAV9-CXCR4-RNAi) and siRNACXCR4, the above indicators were reversed, macrophage autophagy was promoted, the severity of atherosclerotic lesions was reduced, and the disorganized arrangement of myocardial architecture was improved. CONCLUSION: Knockdown of CXCR4 reduces the extent of coronary artery disease by promoting macrophage autophagy through the PI3K/AKT/mTOR pathway to attenuate atherosclerosis.

Genetic analysis of the mitochondrial DNA control region in Tai-Kadai-speaking Dong population in southwest China.

BACKGROUND: Dong people in Southwest China are officially recognised as an ethnic group, but there has been a lack of population genetic research on this group, especially based on mitochondrial DNA data. AIM: To study the sequences and haplogroups of the mitochondrial DNA control region in a typical Dong population, and to provide help for the construction of a forensic mitochondrial DNA analysis reference database in East Asia. SUBJECTS AND METHODS: The sequences of the mitochondrial DNA control region were analysed in 200 individuals of Dong in Guizhou. The haplotype frequencies, haplogroup distribution and paired Fst values of Guizhou Dong and 51 other populations in the world were calculated and explained to explore the genetic polymorphism and population relationships. RESULTS: A total of 180 haplotypes were detected, with frequencies of 0.005-0.02. All haplotypes were assigned to 97 different haplogroups. The haplotype diversity and random matching probability were 0.998643 and 0.00635, respectively. The paired Fst values and correlation p values of 52 populations showed that the Guizhou Dong had the closest genetic relationship with the Henan Han and the Guizhou Miao in China, and were closest to the Punjab population in Pakistan and the Kashmiri population when compared with the world populations. CONCLUSIONS: Our study was based on the matrilineal genetic structure of Guizhou Dong to study mitochondrial DNA, which was helpful to promote the establishment of the forensic DNA reference database in East Asia and provide reference for anthropological research.

Luteolin Ameliorates Methamphetamine-Induced Podocyte Pathology by Inhibiting Tau Phosphorylation in Mice.

Methamphetamine (METH) can cause kidney dysfunction. Luteolin is a flavonoid compound that can alleviate kidney dysfunction. We aimed to observe the renal-protective effect of luteolin on METH-induced nephropathies and to clarify the potential mechanism of action. The mice were treated with METH (1.0-20.0 mg/kg/d bodyweight) for 14 consecutive days. Morphological studies, renal function, and podocyte specific proteins were analyzed in the chronic METH model in vivo. Cultured podocytes were used to support the protective effects of luteolin on METH-induced podocyte injury. We observed increased levels of p-Tau and p-GSK3ß and elevated glomerular pathology, renal dysfunction, renal fibrosis, foot process effacement, macrophage infiltration, and podocyte specific protein loss. Inhibition of GSK3ß activation protected METH-induced kidney injury. Furthermore, luteolin could obliterate glomerular pathologies, inhibit podocyte protein loss, and stop p-Tau level increase. Luteolin could also abolish the METH-induced podocyte injury by inactivating GSK3ß-p-Tau in cultured podocytes. These results indicate that luteolin might ameliorate methamphetamine-induced podocyte pathology through GSK3ß-p-Tau axis.

Trilobatin Alleviates Cognitive Deficits and Pathologies in an Alzheimer's Disease Mouse Model.

Alzheimer's disease (AD) is the most common neurodegenerative disease nowadays that causes memory impairments. It is characterized by extracellular aggregates of amyloid-beta (Aß), intracellular aggregates of hyperphosphorylated Tau (p-Tau), and other pathological features. Trilobatin (TLB), a natural flavonoid compound isolated from Lithocarpuspolystachyus Rehd., has emerged as a neuroprotective agent. However, the effects and mechanisms of TLB on Alzheimer's disease (AD) remain unclear. In this research, different doses of TLB were orally introduced to 3×FAD AD model mice. The pathology, memory performance, and Toll-like receptor 4- (TLR4-) dependent inflammatory pathway protein level were assessed. Here, we show that TLB oral treatment protected 3×FAD AD model mice against the Aß burden, neuroinflammation, Tau hyperphosphorylation, synaptic degeneration, hippocampal neuronal loss, and memory impairment. The TLR4, a pattern recognition immune receptor, has been implicated in neurodegenerative disease-related neuroinflammation. We found that TLB suppressed glial activation by inhibiting the TLR4-MYD88-NFκB pathway, which leads to the inflammatory factor TNF-α, IL-1ß, and IL-6 reduction. Our study shows that TLR4 might be a key target of TLB in AD treatment and suggests a multifaceted target of TLB in halting AD. Taken together, our findings suggest a potential therapeutic effect of TLB in AD treatment.

Transfer of α-synuclein from neurons to oligodendrocytes triggers myelin sheath destruction in methamphetamine administration mice.

Methamphetamine (METH), a widely abused nervous system stimulant, could induce neurotoxicity through α-synuclein (α-syn). Not much is known about the neuronal derived α-syn transmission that underlies oligodendrocyte pathology in METH mice model. In this study, we tested α-syn level, oligodendroglial pathology and autophagy lysosome pathway (ALP) function in corpus callosum in a chronic METH mice model. METH increased α-syn level in neurons and then accumulated in oligodendrocytes. METH increased phosphor-mTOR level, decreased transcription factor EB (TFEB) level and triggered autophagy lysosomal pathway (ALP) impairment, leading to myelin sheath destruction, oligodendroglial proteins loss, mature dendritic spine loss, neuron loss, and astrocyte activation. Deleting endogenous α-syn increased TFEB level, alleviated ALP deficit, and diminished neuropathology induced by METH. TFEB overexpression in oligodendrocytes exerted beneficial effects in METH mice model. These neuroprotective effects were associated with the rescued ALP machinery after oligodendroglial TFEB overexpression. Our study demonstrated, for the first time, that α-syn-TFEB axis might be involve in the METH induced myelin loss, oligodendroglial pathology, and neuropathology. In summary, targeting at the α-syn-TFEB axis might be a promising therapeutic strategy for treating METH induced oligodendroglial pathology, and to a broader view, neurodegenerative diseases.

The role of OX40L and ICAM-1 in the stability of coronary atherosclerotic plaques and their relationship with sudden coronary death.

BACKGROUND: Coronary heart disease is related to sudden death caused by multi-factors and a major threat to human health.This study explores the role of OX40L and ICAM-1 in the stability of coronary plaques and their relationship with sudden coronary death. METHODS: A total of 118 human coronary arteries with different degrees of atherosclerosis and/or sudden coronary death comprised the experimental group and 28 healthy subjects constituted the control group were isolated from patients. The experimental group was subdivided based on whether the cause of death was sudden coronary death and whether it was accompanied by thrombosis, plaque rupture, plaque outflow and other secondary changes: group I: patients with coronary atherosclerosis but not sudden coronary death, group II: sudden coronary death without any of the secondary changes mentioned above, group III: sudden coronary death with coronary artery atherosclerotic lesions accompanied by either of the above secondary changes. The histological structure of the coronary artery was observed under a light microscope after routine HE staining, and the related indexes of atherosclerotic plaque lesions were assessed by image analysis software. The expressions of OX40L and ICAM-1 were detected by real-time quantitative PCR (RT-PCR), immunohistochemistry (IHC) and Western blotting, and the correlations between the expressions and the stability of coronary atherosclerotic plaque and sudden coronary death were analyzed. RESULTS: (1) The expression of OX40L protein in the control group and the three experimental groups showed an increasing trend, and the difference between groups was statistically significant (P < 0.05). (2) The expression of the ICAM-1 protein in the control group and the three experimental groups showed a statistically significant (P < 0.05) increasing trend. (3) The expression of OX40L and ICAM-1 mRNAs increased in the control and the three experimental groups and the difference was statistically significant (P < 0.05). CONCLUSION: The expression of OX40L and ICAM-1 proteins and mRNAs is positively correlated with the stability of coronary atherosclerotic plaque and sudden coronary death.

The mitochondrial DNA control region sequences from the Chinese Miao population of southeastern China.

Background: Miao people are an officially recognised ethnic group living in southwest China, but have seldom been studied genetically, especially with respect to mtDNA data.Aim: To investigate the sequences and haplogroups of the mtDNA control region in a typical Miao population, with the aim of providing a good start for the expansion of the East Asian mtDNA reference database for forensic DNA analysis.Subjects and methods: We analysed 203 Miao individuals, looking at mtDNA control region sequences. We calculated and illustrated the haplotype frequencies, haplogroup distribution and pairwise Fst values between the Miao and six other worldwide populations to explore genetic polymorphisms and population relationships.Results: We observed 121 haplotypes with corresponding frequencies ranging from 0.0049 to 0.0690 in the Miao population. All the samples were assigned to 71 different haplogroups. The haplotype diversity and the random match probability were estimated to be 0.9844 and 0.0204, respectively. The pairwise Fst values and associated p values among seven populations suggest that the Miao population has significant differences to the other six populations, and is relatively isolated compared with them.Conclusions: Our results suggest that frequency estimates for mtDNA haplotypes in Miao ethnic groups should be determined independently rather than being pooled with other populations.

Clinicopathological Characteristics of Traumatic Head Injury in Juvenile, Middle-Aged and Elderly Individuals.

BACKGROUND Traumatic head injury is a leading cause of death and disability worldwide. How clinicopathological features differ by age remains unclear. This epidemiological study analyzed the clinicopathological features of patients with head injury belonging to 3 age groups. MATERIAL AND METHODS Data of patients with traumatic head injury were obtained from the Department of Cerebral Surgery of the Affiliated Hospital of Guizhou Medical University and the Guizhou Provincial People's Hospital in 2011-2015. Their clinicopathological parameters were assessed. The patients were divided into 3 age groups: elderly (≥65 years), middle-aged (18-64 years), and juvenile (≤17 years) individuals. RESULTS Among 3356 hospitalizations for traumatic head injury (2573 males and 783 females, 654 died (19.49%), the highest and lowest mortality rates were in the elderly and juvenile groups, respectively. Fall was the most common cause in juvenile and elderly individuals (32.79% and 43.95%, respectively), while traffic injury was most common in the elderly group (35.08%). The manners of injury differed considerably among the 3 age groups. Scalp injury, skull fracture, intracranial hematoma, and cerebral injury were the most common mechanisms in juvenile (67.32%), middle-aged (63.50%), elderly (69.56%) and middle-aged (90.44%) individuals, respectively. Scalp injury and skull fracture types differed among the groups. Epidural, subdural, and intracerebral hematomas were most common in juvenile, middle-aged, and elderly individuals, respectively. Cerebral contusion showed the highest frequency in the 3 groups, and concussion the lowest. CONCLUSIONS Patients with traumatic HI show remarkable differences in clinicopathological features among juvenile, middle-aged, and elderly individuals.

Insulin-like growth factor binding protein 5 (IGFBP5) mediates methamphetamine-induced dopaminergic neuron apoptosis.

Overexposure to methamphetamine (METH), a psychoactive drug, induces a variety of adverse effects to the nervous system, including apoptosis of dopaminergic neurons. Insulin-like growth factor binding protein 5 (IGFBP5), a member of insulin-like growth factor (IGF) system, is a pro-apoptotic factor that plays important roles in neuronal apoptosis. To test the hypothesis that IGFBP5 can mediate METH-induced neuronal apoptosis, we examined IGFBP5 mRNA and protein expression changes in PC12 cells exposed to METH (3.0mM) for 24h and in the striatum of rats following 15 mg/kg × 8 intraperitoneal injections of METH at 12h interval. We also checked the effect on neuronal apoptosis after silencing IGFBP5 expression with TUNEL staining and flow cytometry; Western blot was used for detecting the expression of apoptotic markers active-caspase3 and PARP. To elucidate the mechanisms underlying IGFBP5-mediated neuronal apoptosis, we determined the release of cytochrome c (cyto c), an apoptogenic factor, from the mitochondria after METH treatment with or without IGFBP5 knockdown. Our results showed that IGFBP5 expression was increased significantly after METH exposure in PC12 cells and in the METH-treated rats' striatum. Further, METH-exposed PC12 cells exhibited higher apoptosis-positive cell number and activity of caspase3 and PARP compared with control cells, while these changes can be blocked by silencing IGFBP5 expression. In addition, a significant increase of cyto c release from mitochondria after METH exposure was observed and it was inhibited after silencing IGFBP5 expression in PC12 cells. These results indicate that IGFBP5 plays key roles in METH-induced neuronal apoptosis and may be a potential gene target for therapeutics in METH-caused neurotoxicity.