Multi-omics-based investigation of Bifidobacterium's inhibitory effect on glioma: regulation of tumor and gut microbiota, and MEK/ERK cascade.

Glioma, the most prevalent primary tumor of the central nervous system, is characterized by a poor prognosis and a high recurrence rate. The interplay between microbes, such as gut and tumor microbiota, and the host has underscored the significant impact of microorganisms on disease progression. Bifidobacterium, a beneficial bacterial strain found in the human and animal intestines, exhibits inhibitory effects against various diseases. However, the existing body of evidence pertaining to the influence of Bifidobacterium on glioma remains insufficient. Here, we found that Bifidobacterium reduces tumor volume and prolongs survival time in an orthotopic mouse model of glioma. Experiments elucidated that Bifidobacterium suppresses the MEK/ERK cascade. Additionally, we noted an increase in the α-diversity of the tumor microbiota, along with an augmented relative abundance of Bifidobacterium in the gut microbiota. This rise in Bifidobacterium levels within the intestine may be attributed to a concurrent increase in Bifidobacterium within the glioma. Additionally, Bifidobacterium induced alterations in serum metabolites, particularly those comprised of organonitrogen compounds. Thus, our findings showed that Bifidobacterium can suppress glioma growth by inhibiting the MEK/ERK cascade and regulating tumor, and gut microbiota, and serum metabolites in mice, indicating the promising therapeutic prospects of Bifidobacterium against glioma.

The role of histone deacetylases in NLRP3 inflammasomes-mediated epilepsy.

Epilepsy is one of the most common brain disorders that not only causes death worldwide, but also affects the daily lives of patients. Previous studies have revealed that inflammation plays an important role in the pathophysiology of epilepsy. Activation of inflammasomes can promote neuroinflammation by boosting the maturation of caspase-1 and the secretion of various inflammatory effectors, including chemokines, interleukins, and tumor necrosis factors. With the in-depth research on the mechanism of inflammasomes in the development of epilepsy, it has been discovered that NLRP3 inflammasomes may induce epilepsy by mediating neuronal inflammatory injury, neuronal loss and blood-brain barrier dysfunction. Therefore, blocking the activation of the NLRP3 inflammasomes may be a new epilepsy treatment strategy. However, the drugs that specifically block NLRP3 inflammasomes assembly has not been approved for clinical use. In this review, the mechanism of how HDACs, an inflammatory regulator, regulates the activation of NLRP3 inflammasome is summarized. It helps to explore the mechanism of the HDAC inhibitors inhibiting brain inflammatory damage so as to provide a potential therapeutic strategy for controlling the development of epilepsy.

Corticotropin releasing factor neurons in the visual cortex mediate long-term changes in visual function induced by early adversity.

Early adversity can cause malfunction of the visual system in adulthood. Adult female but not male mice undergoing early chronic mild stress (ECMS) maintain ocular dominance (OD) plasticity after the critical period. How early stressful experiences have a long-term impact on it is largely unknown. Here, we observed a wide distribution of corticotropin-releasing factor (CRF)-positive neurons, which mainly colocalized with a subpopulation of GABAergic interneurons in the mouse primary visual cortex (V1). Optogenetic activation of CRF-positive neurons transfected with AAV-ChR2 evoked inhibitory currents in nearby pyramidal cells. ECMS induced a reduction in the expression of CRF mRNA in adult mouse V1. Chemogenetic activation of V1 CRF neurons impaired OD plasticity in adult ECMS females. We further showed that local administration of the corticotropin releasing factor receptor 1 (CRFR1) antagonist via an osmotic minipump into the visual cortex mimicked OD plasticity in adult ECMS females. Whole-cell recording in layer 2/3 pyramidal neurons revealed that the CRFR1 antagonist reduced the short-term depression (STD) of evoked inhibitory postsynaptic current (IPSC) in females but not in males. Likewise, CRF agonists have the opposite effect. In summary, our findings indicate that the local CRF-CRFR1 system within V1 may mediate the long-term and sex-dependent effect of early stress experiences on visual plasticity and provide a target for the prevention of visual deficits in adults with a history of early-life adversity.

MicroRNAs, Key Regulators in Glioma Progression as Potential Therapeutic Targets for Chinese Medicine.

Gliomas are tumors of the primary central nervous system associated with poor prognosis and high mortality. The 5-year survival rate of patients with gliomas received surgery combined with chemotherapy or radiotherapy does not exceed 5%. Although temozolomide is commonly used in the treatment of gliomas, the development of resistance limits its use. MicroRNAs are non-coding RNAs involved in numerous processes of glioma cells, such as proliferation, migration and apoptosis. MicroRNAs regulate cell cycle, PI3K/AKT signal pathway, and target apoptosis-related genes (e.g., BCL6), angiogenesis-related genes (e.g., VEGF) and other related genes to suppress gliomas. Evidence illustrates that microRNAs can regulate the sensitivity of gliomas to temozolomide, cisplatin, and carmustine, thereby enhancing the efficacy of these agents. Moreover, traditional Chinese medicine (e.g., tanshinone IIA, xanthohumol, and curcumin) exert antiglioma effects by regulating the expression of microRNAs, and then microRNAs inhibit gliomas through influencing the process of tumors by targeting certain genes. In this paper, the mechanisms through which microRNAs regulate the sensitivity of gliomas to therapeutic drugs are described, and traditional Chinese medicine that can suppress gliomas through microRNAs are discussed. This review aims to provide new insights into the traditional Chinese medicine treatment of gliomas.

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Biochar is beneficial to soil phosphorus (P) availability and crop growth, but the effects vary greatly across different soil types. We investigated the effects of rice straw biochar (4% of total mass) and P application (0, 30, and 90 kg P·hm-2) on soil P availability, phosphomonoesterase activity, and soybean P uptake by using lateritic red soil (pH 4.91) and cinnamon soil (pH 7.24) as test materials. The results showed that biochar application at different P levels significantly increased available P and total P in both soils. Biochar application with 30 kg P·hm-2 increased soil available P with maxima at 192.6% and 237.1% in lateritic red soil and cinnamon soil, respectively. Biochar application with 30 kg P·hm-2 in lateritic red soil significantly increased the activity of alkaline phosphomonoesterase by 78.9%, decreased the content of active organic P by 39.3%, and subsequently stimulated soybean P absorption and growth. Biochar amendment significantly reduced active organic P content in cinnamon soil, but did not affect soil phosphomonoesterase activity and plant growth. The content of active organic P was significantly negatively correlated with soil available P content. In summary, the effect of biochar on soil P availability varied across different soil types (lateritic red soil > cinnamon soil) and P levels (better at 30 kg P·hm-2). Our results could provide scientific basis for a promising application of biochar in reducing the amount of P fertilizer and increasing soybean P uptake, especially in lateritic red soil.

HDAC9 in the Injury of Vascular Endothelial Cell Mediated by P38 MAPK Pathway.

Ischemic stroke caused by atherosclerosis (AS) poses a serious threat to human life expectancy and quality. With the development of genome-wide association studies, the association of histone deacetylase 9 (HDAC9) expression of atheromatous plaques with ischemic stroke in large arteries has been revealed, but the molecular mechanisms behind this phenomenon have not been elucidated. In this study, we explored the effect of HDAC9 on the P38 mitogen activated protein kinase (P38 MAPK), a classic cellular inflammation-related pathway, by knocking down HDAC9 in vascular endothelial cells with short hairpin RNA (shRNA) and found that HDAC9 may mediate oxidized low density lipoprotein (ox-LDL)-induced inflammatory injury in vascular endothelial cells by regulating the phosphorylation level of P38 MAPK to lead to AS. It can be seen that HDAC9 may be a target to control the formation of atherosclerotic plaques. In follow-up experiments, it was verified that sodium valproate (SVA), as a HDAC9 inhibitor, can indeed antagonize the inflammatory damage of vascular endothelial cells, as well as SB203580, which is a P38 MAPK inhibitor. It proves that SVA may be a potential drug for the prevention and treatment of ischemic stroke.

Klotho overexpression improves amyloid-ß clearance and cognition in the APP/PS1 mouse model of Alzheimer's disease.

Alzheimer's disease (AD) is the most prevalent type of dementia, characterized by the presence of amyloid-ß (Aß) plaques. We previously reported that Klotho lowered Aß levels in the brain and protected against cognitive deficits in amyloid precursor protein/presenilin 1(APP/PS1) mice. However, the underlying mechanism remains unclear. In this study, we induced intracerebral Klotho overexpression in 13-month-old APP/PS1 mice by injecting lentivirus that carried full-length mouse Klotho cDNA in the lateral ventricle of the brain. We examined the effects of Klotho overexpression on cognition, Aß burden, Aß-related neuropathology, microglia transformation, and Aß transport systems in vivo. Additionally, we investigated the effects of Klotho on Aß transport at the blood-cerebrospinal fluid barrier by knocking down Klotho in primary human choroid plexus epithelial cells (HCPEpiCs). The upregulation of Klotho levels in the brain and serum significantly ameliorated Aß burden, neuronal and synaptic loss and cognitive deficits in aged APP/PS1 mice. Klotho treatment significantly inhibited NACHT, LRR, and PYD domain-containing protein 3 (NLRP3) and the subsequent transformation of microglia to the M2 type that may enhance microglia-mediated Aß clearance. Meanwhile, Klotho overexpression also regulated Aß transporter expression, which may promote Aß transporter-mediated Aß clearance. Moreover, the ability of HCPEpiCs to transport Aß in vitro was also significantly impaired by Klotho knockdown. Given the neuroprotective effect of Klotho overexpression, the present findings suggest that Klotho should be further investigated as a potential therapeutic target for AD.

miR-181a-5p inhibits the proliferation and invasion of drug-resistant glioblastoma cells by targeting F-box protein 11 expression.

Glioblastoma (GBM) is the most common malignant primary tumor in the human central nervous system. The present study aimed to explore the molecular mechanism by which microRNA (miR)-181a-5p targets the F-box protein 11 (FBXO11) in glioma cells to inhibit cell proliferation and invasion. Reverse transcription-quantitative (RT-q)PCR was performed to detect the expression levels of miR-181a-5p in U251TR cells, U251 cells, primary GBM tissues and relapsed GBM tissues in order to determine the association between miR-181a-5p and the chemoresistance of GBM cells. The expression levels of miR-181a-5p in GBM cells were modulated via transfecting miR-181a-5p mimics and inhibitors. Cell Counting Kit-8 assays were undertaken to assess the effects of miR-181a-5p on drug sensitivity and proliferation of GBM cells. Wound healing assays were performed to examine the effects of miR-181a-5p on the migratory ability of GBM cells. Furthermore, the effects of miR-181a-5p on the invasive ability of GBM cells were analyzed using an in vitro invasion assay. Flow cytometry analysis was carried out to determine whether overexpression of miR-181a-5p can promote the apoptotic rate of GBM cells. RT-qPCR and western blotting were employed to detect the effects of miR-181a-5p on mRNA and protein expression of FBX011. miR-181a-5p exhibited low expression in resistant GBM cell lines and recurrent tumor tissues. Dual-luciferase reporter assays were utilized to detect luciferase activity to verify the targeted regulatory association between miR-181a-5p and FBXO11. Upregulation of miR-181a-5p promoted the sensitivity of GBM cells to temozolomide (TMZ), increased the apoptotic rate of GBM cells and significantly inhibited the invasive and migratory capacities of GBM cells. In drug-resistant glioma cells, compared with the miR-negative control group and the blank group, the expression of miR-181a-5p was significantly upregulated (P<0.01), while the expression of FBXO11 protein was downregulated. miR-181a-5p increased the sensitivity of GBM cells to TMZ. miR-181a-5p significantly inhibited the migratory and invasive capacities of GBM cells. miR-181a-5p may become a novel effective target for the treatment of GBM. The results of dual-luciferase reporter assays indicated that miR-181a-5p could target the 3'-untranslated region of FBXO11. The underlying mechanism may be targeted inhibition of FBXO11 gene expression, or may be associated with apoptosis.

Regulation of SPARC family proteins in disorders of the central nervous system.

SPARC (secreted protein acidic and rich in cysteine) family of proteins is a class of protein involved in tissue development and repair by regulating cell adhesion, proliferation, metastasis, and growth factor signaling to affect the extracellular matrix (ECM) and interactions between cells. Although being highly valued in non-nerve tissues, studies in both cell and animal models have been revealed that SPARC family proteins may also continue to play a vital role in central nervous system (CNS) diseases and development. These SPARC family proteins are widely expressed in the CNS of normal people, and significantly increased in brain tissue following disease or injury, such as SPARC, SPARCL-1, FSTL-1 and testican. In our review, we will pay attention to the functions of SPARC family proteins in autophagy, apoptosis, angiogenesis, adipogenesis, inflammatory response, and cerebral tissue development, injury and repair mainly including but not limited to axon regeneration, formation of glial scar, neural plasticity, regulation of nerve conduction related receptors and rewiring of neural circuitry.

Iron Metabolism and Ferroptosis in Epilepsy.

Epilepsy is a disease characterized by recurrent, episodic, and transient central nervous system (CNS) dysfunction resulting from an excessive synchronous discharge of brain neurons. It is characterized by diverse etiology, complex pathogenesis, and difficult treatment. In addition, most epileptic patients exhibit social cognitive impairment and psychological impairment. Iron is an essential trace element for human growth and development and is also involved in a variety of redox reactions in organisms. However, abnormal iron metabolism is associated with several neurological disorders, including hemorrhagic post-stroke epilepsy and post-traumatic epilepsy (PTE). Moreover, ferroptosis is also considered a new form of regulation of cell death, which is attributed to severe lipid peroxidation caused by the production of reactive oxygen species (ROS) and iron overload found in various neurological diseases, including epilepsy. Therefore, this review summarizes the study on iron metabolism and ferroptosis in epilepsy, in order to elucidate the correlation between iron and epilepsy. It also provides a novel method for the treatment, prevention, and research of epilepsy, to control epileptic seizures and reduce nerve injury after the epileptic seizure.

SPARC Knockdown Reduces Glutamate-Induced HT22 Hippocampal Nerve Cell Damage by Regulating Autophagy.

Secreted protein acidic and rich in cysteine (SPARC) is a matricellular protein involved in the extracellular matrix and interactions between cells during neural development of the central nervous system (CNS). Oxidative glutamate toxicity is involved in CNS diseases, including epilepsy, Alzheimer's disease, and ischemic stroke. However, the molecular mechanism of nerve injury is not fully understood in CNS diseases. Herein, the glutamate-induced nerve damage model was used to explore the molecular mechanisms affecting nerve damage. The levels of SPARC and autophagy were increased in glutamate-induced HT22 hippocampal nerve injury. In summary, the current study confirmed that SPARC regulates autophagy in HT22 hippocampal nerve cells, and its knockdown reduces the glutamate-induced HT22 hippocampal nerve injury by inhibiting autophagy. These findings suggested that SPARC plays a crucial role in nerve injury of CNS diseases.

Lentiviral vector-mediated overexpression of Klotho in the brain improves Alzheimer's disease-like pathology and cognitive deficits in mice.

Alzheimer's disease (AD) is the most common type of senile dementia. The antiaging gene Klotho is reported to decline in the brain of patients and animals with AD. However, the role of Klotho in the progression of AD remains elusive. The present study explored the effects and underlying mechanism of Klotho in a mouse model of AD. The upregulation of cerebral Klotho expression was mediated by an intracerebroventricular injection of a lentiviral vector that encoded Klotho (LV-KL) in 7-month-old amyloid precursor protein/presenilin 1 transgenic mice. Three months later, LV-KL significantly induced Klotho overexpression in the brain and effectively ameliorated cognitive deficit and AD-like pathology in amyloid precursor protein/presenilin 1 mice. LV-KL induced autophagy activation and protein kinase B/mammalian target of rapamycin inhibition both in AD mice and BV2 murine microglia. These results suggest that the upregulation of Klotho expression in the brain may promote the autophagic clearance of amyloid beta and protect against cognitive deficits in AD mice. These findings highlight the preventive and therapeutic potential of Klotho for the treatment of AD.

The neuroprotective effects and probable mechanisms of Ligustilide and its degradative products on intracerebral hemorrhage in mice.

BACKGROUND: Intracerebral hemorrhage (ICH) is a common neurological emergency with higher mortality and disability rate than cerebral ischemia. Although diverse therapeutic interventions have been explored for potential neuroprotection from ICH, no effective drugs until now are available for improvement of survival rate or the life quality of survivors after ICH. Just like cerebral ischemia, inflammatory mechanism is highly thought to play a vital role in hemorrhagic brain injury. Ligustilide (LIG) has potent anti-inflammatory effects, which were shown to be closely related to its neuroprotective effects against ischemic brain injury. Senkyunolide H (SH) and senkyunolide I (SI) are natural degradation products of LIG, which contain the mother nucleus structure of LIG as that of phthalide. However, no reports have been retrieved about the neuroprotective effects of the three phthalide compounds on ICH, especially from the perspectives of inflammatory pathways. Accordingly, this study investigated the neuroprotective potentials and mechanisms of LIG, SH and SI on experimental ICH in mice. METHODS: ICH was induced in adult male CD-1 mice by intracerebral injection of autologous blood. LIG, SH and SI, respectively, was administrated after ICH induction. Neurological deficits, brain edema, injury volume, the number of surviving/dying neurons and inflammatory gene expression were evaluated at 3 days after ICH. RESULTS: Neurological deficits, brain edema, neuronal injury, microglia and astrocytes activation as well as peripheral immune cells infiltration were all significantly improved by LIG and SH, yet SI not. Moreover, the expression of TLR4, p-NF-kB p65, TNF-α and IL-6, was significantly downregulated by LIG and SH treatment. So was Prx1 expression and release. CONCLUSIONS: LIG and SH provide the potent neuroprotective effects against hemorrhagic stroke by inhibiting Prx1/TLR4/NF-kB signaling and the subsequent immune and neuroinflammation lesions.

Lentivirus-mediated klotho up-regulation improves aging-related memory deficits and oxidative stress in senescence-accelerated mouse prone-8 mice.

AIMS: Oxidative stress caused by aging aggravates neuropathological changes and cognitive deficits. Klotho, an anti-aging protein, shows an anti-oxidative effect. The aims of the present study were to determine the potential therapeutic effect of klotho in aging-related neuropathological changes and memory impairments in senescence-accelerated mouse prone-8 (SAMP8) mice, and identify the potential mechanism of these neuroprotective effects. MATERIALS AND METHODS: A lentivirus was used to deliver and sustain the expression of klotho. The lentiviral vectors were injected into the bilateral lateral ventricles of 7-month-old SAMP8 mice or age-matched SAMR1 mice. Three months later, the Y-maze alternation task and passive avoidance task were used to assess the memory deficits of the mice. In situ hybridization, immunohistochemistry, immunofluorescence, Nissl staining, quantitative real-time PCR and Western blot assays were applied in the following research. KEY FINDINGS: Our results showed that 3 months after injection of the lentiviral vectors encoding the full-length klotho gene, the expression of klotho in the brain was significantly increased in 10-month-old SAMP8 mice. This treatment reduced memory deficits, neuronal loss, synaptic damage and 4-HNE levels but increased mitochondrial manganese-superoxide dismutase (Mn-SOD) and catalase (CAT) expression. Moreover, the up-regulation of klotho expression decreased Akt and Forkhead box class O1 (FoxO1) phosphorylation. SIGNIFICANCE: The present study provides a novel approach for klotho gene therapy and demonstrates that direct up-regulation of klotho in the brain might improve aging-related memory impairments and decrease oxidative stress. The underlying mechanism of this effect likely involves the inhibition of the Akt/FoxO1 pathway.

Neuroprotective effect of a novel gastrodin derivative against ischemic brain injury: involvement of peroxiredoxin and TLR4 signaling inhibition.

The inhibition of extracellular inflammatory peroxiredoxin (Prx) signaling appears to be a potential therapeutic strategy for neuroinflammatory injury after acute ischemic stroke. Gastrodin (Gas) is a phenolic glycoside that is used for the treatment of cerebral ischemia, accompanied by regulation of the autoimmune inflammatory response. The present study investigated the neuroprotective effects of Gas and its derivative, Gas-D, with a focus on the potential mechanism associated with inflammatory Prx-Toll-like receptor 4 (TLR4) signaling. Gas-D significantly inhibited Prx1-, Prx2-, and Prx4-induced inflammatory responses in RAW264.7 macrophages and H2O2-mediated oxidative injury in SH-SY5Y nerve cells. In rats, intraperitoneal Gas-D administration 10 h after reperfusion following 2-h middle cerebral artery occlusion (MCAO) ameliorated neurological deficits, brain infarction, and neuropathological alterations, including neuron loss, astrocyte and microglia/macrophage activation, T-lymphocyte invasion, and lipid peroxidation. Delayed Gas-D treatment significantly inhibited postischemic Prx1/2/4 expression and spillage, TLR4 signaling activation, and inflammatory mediator production. In contrast, Gas had no significant effects in either cell model or in MCAO rats under the same conditions. These results indicate that Gas-D may be a drug candidate with an extended therapeutic time window that blocks inflammatory responses and attenuates the expression and secretome of inflammatory Prxs in acute ischemic stroke.

Neuroprotective Effect of Ligustilide through Induction of α-Secretase Processing of Both APP and Klotho in a Mouse Model of Alzheimer's Disease.

Emerging evidence suggests that alpha-processing single transmembrane proteins, amyloid precursor protein (APP) and anti-aging protein Klotho, are likely to be involved in the progression of Alzheimer's disease (AD). The natural phthalide Ligustilide (LIG) has been demonstrated to protect against aging- and amyloid-ß (Aß)-induced brain dysfunction in animal models. The present study is to investigate the effects of LIG on cognitive deficits and metabolism of both APP and Klotho and its underlying mechanism in AD double-transgenic (APP/PS1) mice and cultured human cells. Our results show that treatment with LIG significantly ameliorated memory impairment and Aß levels and plaques burden. Specifically, LIG might act as a potent enhancer of α-secretase, disintegrin, and metalloprotease 10 (ADAM10), leading to upregulation of alpha-processing of both APP and Klotho and subsequent increases in the levels of both soluble APP fragment (sAPPα) and soluble Klotho (sKL) with inhibition of IGF-1/Akt/mTOR signaling in AD mice and cultured cells. Moreover, the specific ADAM10 inhibitor (G1254023X) effectively reversed LIG-induced alpha-processing of both APP and Klotho in vitro, while Klotho gene knockdown by small interfering RNA significantly blunted LIG-mediated inhibition of IGF-1/Akt/mTOR signaling in vitro. Taken together with the reported neuroprotective effects of both sAPPα and sKL as well as autophagy induction by Akt/mTOR pathway inhibition, our findings suggest that neuroprotection of LIG against AD is associated with induction alpha-processing of APP and Klotho and potential Aß clearance. Whether LIG might induce Aß autophagic clearance and the underlying mechanisms need to be further studied.

Protective Effect of Klotho against Ischemic Brain Injury Is Associated with Inhibition of RIG-I/NF-κB Signaling.

Aging is the greatest independent risk factor for the occurrence of stroke and poor outcomes, at least partially through progressive increases in oxidative stress and inflammation with advanced age. Klotho is an antiaging gene, the expression of which declines with age. Klotho may protect against neuronal oxidative damage that is induced by glutamate. The present study investigated the effects of Klotho overexpression and knockdown by an intracerebroventricular injection of a lentiviral vector that encoded murine Klotho (LV-KL) or rat Klotho short-hairpin RNA (LV-KL shRNA) on cerebral ischemia injury and the underlying anti-neuroinflammatory mechanism. The overexpression of Klotho induced by LV-KL significantly improved neurobehavioral deficits and increased the number of live neurons in the hippocampal CA1 and caudate putamen subregions 72 h after cerebral hypoperfusion that was induced by transient bilateral common carotid artery occlusion (2VO) in mice. The overexpression of Klotho significantly decreased the immunoreactivity of glial fibrillary acidic protein and ionized calcium binding adaptor molecule-1, the expression of retinoic-acid-inducible gene-I, the nuclear translocation of nuclear factor-κB, and the production of proinflammatory cytokines (tumor necrosis factor α and interleukin-6) in 2VO mice. The knockdown of Klotho mediated by LV-KL shRNA in the brain exacerbated neurological dysfunction and cerebral infarct after 22 h of reperfusion following 2 h middle cerebral artery occlusion in rats. These findings suggest that Klotho itself or enhancers of Klotho may compensate for its aging-related decline, thus providing a promising therapeutic approach for acute ischemic stroke during advanced age.

Visfatin is involved in promotion of colorectal carcinoma malignancy through an inducing EMT mechanism.

Increasing evidences suggested visfatin, a newly discovered obesity-induced adipocytokine, is involved in promotion of cancer malignancy and correlated with worse clinical prognosis. While its effects and mechanisms on progression of colorectal cancer (CRC) remain unclear. Our clinical data show that visfatin protein is over expressed, positive associated with lymph node metastasis, high-grade tumor, and poor prognosis in 87 CRC patients. The levels of plasma visfatin are significantly upregulated in Stage IV colon cancer. Visfatin can significantly promote the in vitro migration and invasion of CRC cells via induction epithelial mesenchymal transition (EMT). It can increase the expression and nuclear translocation of Snail, a key transcription factor in regulating EMT. While silencing of Snail attenuates visfatin induced EMT. Further studies reveal visfatin can inhibit the association of Snail with GSK-3ß and subsequently suppress ubiquitylation of Snail. In addition, visfatin can increase the expression and nuclear translocation of ß-catenin, elevate its binding with Snail promoter, and then increase the transcription of Snail. While inhibitor of PI3K/Akt, LY294002, abolishes visfatin induced up regulation of Snail, Vimentin (Vim), ß-catenin, and phosphorylated GSK-3ß. In summary, our data suggest that increased expression of visfatin are associated with a more aggressive phenotype of CRC patients. It can trigger the EMT of CRC cells via Akt/GSK-3ß/ß-catenin signals.

The mechanism of apoliprotein A1 down-regulated by Hepatitis B virus.

BACKGROUND: Hepatitis B virus (HBV) infection correlated with the development of cirrhosis, liver failure and hepatocellular carcinoma (HCC), poses a huge health burden on the global community. However, the pathogenesis of chronic hepatitis B (CHB) remains unclear. Apolipoprotein A1 (ApoA1) mainly secreted by hepatocytes, represents the major protein component of high-density lipoprotein. ApoA1 secretion may be disrupted by HBV infection. In this study, we mainly investigated the molecular mechanism of ApoA1 down regulated by HBV for revealing the pathogenesis of CHB. METHODS: ApoA1 expression in livers of CHB patients as well as healthy controls were performed by Real-time PCR (RT-PCR) and Western blot. The serum ApoA1 levels were measured by Enzymed-linked immunosorbent assay (ELISA). Expression of ApoA1 mRNA and protein levels were performed by RT-PCR and Western blot in human hepatoma HepG2 cells and subline HepG2.2.15 cells. HBV expression construct, pHBV1.3 were transfected into HepG2, the changes of ApoA1 mRNA and protein expression were detected by RT-PCR and Western blot. To further study the mechanism of ApoA1 down regulation by HBV, 11 CpG islands in ApoA1 promotor were tested for DNA methylation status by MSP. HepG2.2.15 cell lines were treated with DNA methyltransferase inhibitor 5-aza-deoxycytidine (5-aza-dC), then, expression of ApoA1 mRNA and HBV particles in the supernatant, as well as ApoA1 protein levels were detected by RT-PCR and Western blot. Secretion of HBsAg and HBeAg in HepG2 cells cotransfected with pApoA1 and pHBV1.3 constructs was tested by ELISA. Meanwhile, secretion of HBsAg and HBeAg in the supernatant were quantified by ELISA in the HepG2.2.15 cells treated with 5-aza-dC plus ApoA1 siRNA. RESULTS: Expression of ApoA1 mRNA and protein levels, as well as serum ApoA1 levels in CHB patients were decreased corresponding healthy controls in vivo. In addition, the expression of ApoA1 mRNA and protein levels were down regulated in HepG2.2.15 cells correponding HepG2 cells, 11 CpG islands in ApoA1 promoter were tested for methylation status by MSP in HepG2.2.15 cells compared to HepG2 cells, while two CpG islands were found hypermethylated. Expression of ApoA1 mRNA and protein levels were increased in HepG2.2.15 cells treated with DNA methyltransferase inhibitor 5-aza-dC. Furthermore, overexpression of ApoA1 can enhance HBV expression in HepG2 cells while the inhibitory effect of 5-aza-dC on HBV expression was completely abolished by blocking 5-aza-dC-induced up-regulation of ApoA1 using RNAi. CONCLUSIONS: Epigenetic silencing of ApoA1 gene expression by CpG island DNA hypermethylation induced by HBV may contribute to the pathogenesis of CHB.