Wen-bin He

Nrf2 pathway activation contributes to anti-fibrosis effects of ginsenoside Rg1 in a rat model of alcohol- and CCl4-induced hepatic fibrosis.

Aim:To investigate the anti-fibrosis effects of ginsenoside Rg1 on alcohol- and CCl4-induced hepatic fibrosis in rats and to explore the mechanisms of the effects.Methods:Rats were given 6% alcohol in water and injected with CCl4 (2 mL/kg, sc) twice a week for 8 weeks. Rg1 (10, 20 and 40 mg/kg per day, po) was administered in the last 2 weeks. Hepatic fibrosis was determined by measuring serum biochemical parameters, HE staining, Massons trichromic staining, and hydroxyproline and α-SMA immunohistochemical staining of liver tissues. The activities of antioxidant enzymes, lipid peroxidation, and Nrf2 signaling pathway-related proteins (Nrf2, Ho-1 and Nqo1) in liver tissues were analyzed. Cultured hepatic stellate cells (HSCs) of rats were prepared for in vitro studies.Results:In the alcohol- and CCl4-treated rats, Rg1 administration dose-dependently suppressed the marked increases of serum ALT, AST, LDH and ALP levels, inhibited liver inflammation and HSC activation and reduced liver fibrosis scores. Rg1 significantly increased the activities of antioxidant enzymes (SOD, GSH-Px and CAT) and reduced MDA levels in liver tissues. Furthermore, Rg1 significantly increased the expression and nuclear translocation of Nrf2 that regulated the expression of many antioxidant enzymes. Treatment of the cultured HSCs with Rg1 (1 μmol/L) induced Nrf2 translocation, and suppressed CCl4-induced cell proliferation, reversed CCl4- induced changes in MDA, GPX, PCIII and HA contents in the supernatant fluid and α-SMA expression in the cells. Knockdown of Nrf2 gene diminished these actions of Rg1 in CCl4-treated HSCs in vitro.Conclusion:Rg1 exerts protective effects in a rat model of alcohol- and CCl4-induced hepatic fibrosis via promoting the nuclear translocation of Nrf2 and expression of antioxidant enzymes.

The Multifunctionality of Human Vγ9Vδ2 γδ T Cells: Clonal Plasticity or Distinct Subsets?

The dominant subset of γδ T cells in human peripheral blood expresses Vγ9 paired with Vδ2 as variable TCR elements. Vγ9Vδ2 T cells recognize pyrophosphates derived from the microbial non‐mevalonate isoprenoid biosynthesis pathway at pico‐ to nanomolar concentrations. Structurally related pyrophosphates are generated in eukaryotic cells through the mevalonate pathway involved in protein prenylation and cholesterol synthesis. However, micromolar concentrations of endogenous pyrophosphates are required to be recognized by Vγ9Vδ2 T cells. Such concentrations are not produced by normal cells but can accumulate upon cellular stress and transformation. Therefore, many tumour cells are susceptible to γδ T cell–mediated lysis owing to the overproduction of endogenous pyrophosphates. This explains why Vγ9Vδ2 T cells contribute to both anti‐infective and anti‐tumour immunity. Ex vivo analysed Vγ9Vδ2 T cells can be subdivided on the basis of additional surface markers, including chemokine receptors and markers for naïve and memory T cells. At the functional level, Vγ9Vδ2 T cells produce a broad range of cytokines, display potent cytotoxic activity, regulate αβ T cell responses, and – quite surprisingly – can act as professional antigen‐presenting cells. Thus, an exceptional range of effector functions has been assigned to a population of T cells, which all recognize invariant exogenous or endogenous pyrophosphates that are not seen by any other immune cell. Here, we discuss whether this plethora of effector functions reflects the plasticity of individual Vγ9Vδ2 T cells or can be assigned to distinct subsets.

Immobilized MICA Could Expand Human Vδ1 γδ T Cells In Vitro that Displayed Major Histocompatibility Complex Class I Chain‐Related A‐Dependent Cytotoxicity to Human Epithelial Carcinomas

Human major histocompatibility complex class I chain‐related A (MICA) is a human leucocyte antigen‐related polymorphic molecule, which is expressed on many kinds of epithelial tumours and can be recognized by the Vδ1 subset of γδ T cells. In the present study, monoclonal antibodies (MoAbs) were produced in mice immunized with recombinant MICA (rMICA)*008. It was found that MICA was expressed on ovarian and colonic tumour tissues and could be detected by these anti‐MICA MoAbs. The immobilized rMICA could induce the proliferation of human ovarian epithelial carcinoma‐ or colonic carcinoma‐derived γδ T cells of the Vδ1 phenotype in vitro. These Vδ1 T cells displayed a strong, broad‐range cytolytic activity towards tumour cell lines positive for MICA. The efficiency of this cytolytic activity depended greatly on the level of MICA expressed on the cell surface and could be inhibited by anti‐MICA MoAbs. Therefore, MICA may play an important role in immune responses against epithelial tumours and function as a stimulating factor for the growth of Vδ1 γδ T cells, whereas MICA‐reactive Vδ1 γδ T cells might serve as a new candidate for adoptive cellular therapy of tumours.

Protopanaxtriol protects against 3-nitropropionic acid-induced oxidative stress in a rat model of Huntington's disease

Aim:Protopanaxtriol (Ppt) is extracted from Panax ginseng Mayer. In the present study, we investigated whether Ppt could protect against 3-nitropropionic acid (3-NP)-induced oxidative stress in a rat model of Huntingtons disease (HD) and explored the mechanisms of action.Methods:Male SD rats were treated with 3-NP (20 mg/kg on d 1, and 15 mg/kg on d 2–5, ip). The rats received Ppt (5, 10, and 20 mg/kg, po) daily prior to 3-NP administration. Nimodipine (12 mg/kg, po) or N-acetyl cysteine (NAC, 100 mg/kg, po) was used as positive control drugs. The body weight and behavior were monitored within 5 d. Then the animals were sacrificed, neuronal damage in striatum was estimated using Nissl staining. Hsp70 expression was detected with immunohistochemistry. Reactive oxygen species (ROS) generation was measured using dihydroethidium (DHE) staining. The levels of components in the Nrf2 pathway were measured with immunohistochemistry and Western blotting.Results:3-NP resulted in a marked reduction in the body weight and locomotion activity accompanied by progressive striatal dysfunction. In striatum, 3-NP caused ROS generation mainly in neurons rather than in astrocytes and induced Hsp70 expression. Administration of Ppt significantly alleviated 3-NP-induced changes of body weight and behavior, decreased ROS production and restored antioxidant enzymes activities in striatum. Moreover, Ppt directly scavenged free radicals, increased Nrf2 entering nucleus, and the expression of its downstream products heme oxygenase-1 (HO-1) and NAD(P)H quinone oxidase 1 (NQO1) in striatum. Similar effects were obtained with the positive control drugs nimodipine or NAC.Conclusion:Ppt exerts a protective action against 3-NP-induced oxidative stress in the rat model of HD, which is associated with its anti-oxidant activity.

Downregulation of CD94/NKG2A inhibitory receptor on decreased γδ T cells in patients with systemic lupus erythematosus.

γδ T cells are characterized by recognizing conserved endogenous and stress‐induced antigens without antigen presentation. It has been show that γδ T cells play an important role in anti‐tumour/microbe responses, but their function in autoimmune diseases is yet not clear. Here, we reported the quantity and phenotype of peripheral blood γδ T cells from systemic lupus erythematosus (SLE). Both the percentages of γδ T cells in peripheral blood and among CD3+ T cells of patients with SLE were significantly decreased, regardless of disease activity. However, activating marker CD69 and HLA‐DR was upregulated, while inhibiting receptor CD94/NKG2A was downregulated in γδ T cells of patients with SLE. The expression of CD69 is negatively correlated with the quantity of γδ T cells. Moreover, the expression of CD94/NKG2A remained low even with antigen stimulation on those γδ T cells. Our results suggested that the low expression level of CD94/NKG2A upon γδ T cell activation might lead to the over‐activation of γδ T cells in patients with SLE. These findings will be useful in elucidating the roles of γδ T cells in SLE pathogenesis.

Flavin-containing monooxygenase, a new clue of pathological proteins in the rotenone model of parkinsonism.

Parkinsons disease (PD) is a major age-related neurodegenerative disorder characterized by loss of dopaminergic neurons in the substantia nigra par compacta (SNpc) and accumulation of aggregated alpha-synuclein in brain areas. Rotenone is a neurotoxin that is routinely used to model PD, thus to help us understand the mechanisms of neural death. Flavin-containing monooxygenase (FMO), usually known as an important hepatic microsomal enzyme like cytochrome P450, was found to play a role in the brain recent years. In our study we aimed to find out the role that FMO might play in PD pathology. Thus we successfully generated rotenone model in primary midbrain dopaminergic neurons and identified the apoptosis of neurons caused by rotenone. We found that in rotenone model of Parkinsonism, the expression/protein level of parkin and FMO1 were decreased accompanied by the activation of caspase 3. Blocking FMO activity by FMO inhibitor methimazole directly caused activation of caspase 3, meanwhile parkin protein level was decreased. Our data indicated that FMO, whose dysfunction could be a reason for the apoptosis of dopaminergic neurons in rotenone model, might be a new clue of pathological proteins in rotenone model of parkinsonism. Meanwhile, it was suggested that parkin function was compromised in neuro-pathological states, thereby further adding to the cellular survival stress.

Claulansine F promotes neuritogenesis in PC12 cells via the ERK signaling pathway

Aim:To study the effects of Claulansine F (Clau F), a carbazole alkaloid isolated from the stem of Clausena lansium (Lour) Skeels, on neuritogenesis of PC12 cells, and to elucidate the mechanism of action.Methods:Neuritogenesis of PC12 cells was quantified under an inverted microscope. Expression of the neurite outgrowth marker GAP-43 was detected using immunofluorescence. GAP-43 transcription was measured using RT-PCR. Cell viability was evaluated with MTT assay. The levels of phosphor-ERK1/2, phosphor-CREB, phosphor-AKT and acetylate-p53 in the cells were examined using Western blotting analyses.Results:Clau F (10–100 μmol/L) significantly increased the percentage of PC12 cells bearing neurites. Clau F markedly increased the expression of GAP-43 in the cells. The efficiency of Clau F (10 μmol/L) in increasing neuritogenesis and GAP-43 expression was comparable to that of nerve growth factor (50 ng/mL). In addition, Clau F completely blocked the proliferation of PC12 cells within 7 d of incubation, whereas it did not cause cell death in cultured rat cortical neurons. Treatment of PC12 cells with Clau F activated both ERK and AKT signaling pathways. Co-treatment of PC12 cells with the specific ERK inhibitor PD98059, but not the specific PI3K inhibitor LY294002, blocked Clau F-induced neuritogenesis and GAP-43 upregulation.Conclusion:Clau F promotes neuritogenesis in PC12 cells specifically via activation of the ERK signaling pathway.

Effect of corticosterone on developing hippocampus: short-term and long-term outcomes.

Many documents implicate that corticosterone plays a negative role in brain function, especially in learning and memory. However, less evidence confirms its direct actions on hippocampal development. In the work reported here, pro treatment, minimum corticosterone administration in infant mice, and con treatment, corticosterone deprivation by adrenalectomy, were used to examine the effects imposed by corticosterone on the structure and function of developing hippocampus. Our study shows that adrenalectomy induces decrease of plasma corticosterone levels and results in the impairment of learning performance and the degenerative changes not in CA regions of hippocampus but in dentate gyrus. Noteworthily, this damage effect is severer in 5‐week‐old mice than that in 10‐week‐old mice. In addition, the short‐term effect of minimum corticosterone administration may accelerate the development of dentate gyrus of 10‐day‐old mice. Moreover, minimum corticosterone administration during infancy contributed to the learning performance and the structural integrity of hippocampal CA regions in different developing stages, while this phenomenon was not observed in dentate gyrus. In conclusion, corticosterone is necessary for the development of dentate gyrus, especially in relatively young individuals, and administration with minimum corticosterone in infancy has a long‐term positive influence on the hippocampal structure and function in different developing stages.

Effects of cerebral glucose levels in infarct areas on stroke injury mediated by blood glucose changes

Admission hyperglycemia is considered to be related to poor outcomes of ischemic stroke. However, there is controversy regarding effects of attempts to lower blood glucose in stroke patients. This study aimed at determining the effects of blood glucose fluctuation on stroke injury by detection of cerebral glucose levels. A single intraperitoneal injection of glucose (0, 0.5, 1, 1.5 or 2 g kg−1) at 5 min before reperfusion caused blood glucose fluctuation (5–15 mmol L−1) lasting for 2 h after reperfusion. Blood glucose levels of 6–10 mmol L−1 decreased stroke injury after reperfusion for 24 h and 28 days compared with conditions of hyperglycemia (>10 mmol L−1) and hypoglycemia (<6 mmol L−1). High glucose concentration increased neuronal injury and death after oxygen–glucose deprivation. Under hyperglycemia and hypoglycemia, elevations in expression of MMP-2/-9 and decrease of tight junction proteins including occludin, claudin-5 and ZO-1 contributed to blood–brain barrier (BBB) dysfunction in infarct regions after ischemia–reperfusion injury, and reduction of hexokinase, pyruvate kinase and lactate dehydrogenase activities significantly inhibited the glucose metabolism in cortex and striatum after 24 h of reperfusion. BBB damage and reduced glucose metabolism caused accumulation of glucose in infarct areas. An obvious increase in cerebral glucose levels aggravated stroke injury.

Characterization of Complementary Determinant Region 3δ in Human MutS Homologue 2-Specific γδ T Cells

γδT cells function as sentinels in early host responses to infections and malignancies. Previously, we found ectopically expressed human MutS homologue 2 (hMSH2), recognized by γδT cells, triggered a γδT cell‐mediated cytolysis to tumor cells. However, the characteristics of hMSH2‐specific γδ Τ cells are not fully understood. In this study, we investigated the complementary determinant region (CDR) 3δ diversity of hMSH2‐specific γδ T cells. We found that the CDR3δ sequences of hMSH2‐specific γδ T cells displayed limited diversity, while the length and germline gene usage showed no differences compared with whole CDR3δ immune repertoire. There are more hydrophilic amino acids in P/N insert of hMSH2‐specific γδ T cells including the more conserved amino acid at the position 97. Our results offer clues to understanding antigen recognition pattern of γδ T cells to stress‐induced hMSH2 of tumor cells and also the mechanism of γδT cell‐mediated tumor immune surveillance.