Chenggang Zhang

Characterization and expression of three novel differentiation-related genes belong to the human NDRG gene family.

NDRG1(N-Myc downstream regulated) is upregulated during cell differentiation, repressed by N-myc and c-myc in embryonic cells, and suppressed in several tumor cells. A nonsense mutation in the NDRG1 gene has been reported to be causative for hereditary motor and sensory neuropathy-Lom (HMSNL), indicating that NDRG1 functions in the peripheral nervous system necessary for axonal survival. Here, we cloned three human cDNAs encoding NDRG2 (371aa), NDRG3 (375aa) and NDRG4 (339aa), which are homologous to NDRG1. These three genes, together with NDRG1, constitute the NDRG gene family. The phylogenetic analysis of the family demonstrated that human NDRG1 and NDRG3 belong to a subfamily, and NDRG2 and NDRG4 to another. At amino acid (aa) level, the four members share 53–65% identity. Each of the four proteins contains an α/β hydrolase fold as in human lysosomal acid lipase. Expression of the fusion proteins NDRG2/GFP, NDRG3/GFP and NDRG4/GFP in COS-7 cells showed that all of them are cytosolic proteins. Based on UniGene cluster analysis, the genes NDRG2, NDRG3 and NDRG4 are located at chromosome 14q11.1–11.2, 20q12–11.23 and 16q21–22.1, respectively. Northern and dot blot analysis shows that all of the three genes are highly expressed in adult brain and almost not detected in the eight human cancer lines. In addition, in contrast to the relatively ubiquitous expression of NDRG1, NDRG2 is highly expressed in adult skeletal muscle and brain, NDRG3 highly expressed in brain and testis, and NDRG4 specifically expressed in brain and heart, suggesting that they might display different specific functions in distinct tissues.

Intracrine hepatopoietin potentiates AP-1 activity through JAB1 independent of MAPK pathway

Many growth factors and cytokines are involved in liver regeneration. Of them, only hepatopoietin (HPO)/ALR (augmenter of liver regeneration) is a specifically hepatotrophic factor originally identified from the cytosol of regenerating or hyperplastic hepatic cells. Previous reports indicate that extracellular HPO triggers the MAPK pathway by binding its specific receptor on the cell surface. However, its function in the cytosol of hepatocytes is unclear. Here we identified that JAB1 (Jun activation domain‐binding protein 1), a co‐activator of AP‐1, which is essential for liver regeneration, specifically interacts with intracellular HPO. JAB1 colocalizes with HPO in nuclei of hepatic cells or COS‐7 cells. As an intracrine factor, the intracellular function of HPO is to increase c‐Jun phosphorylation independent of c‐Jun amino‐terminal kinase (JNK), extracellular signal‐regulated kinase (ERK) ‐1 and ‐2, and leads to potentiation of JAB1‐mediated AP‐1 activation. Amino acids 1‐63 of HPO molecule are sufficient to bind to JAB1, but the full‐length HPO is necessary for its intracellular signaling. Taken together, these results elucidate a novel mechanism of intracrine cytokine signaling by specifically modulating the AP‐1 pathway through JAB1, in a MAPK‐independent fashion.

Identification and Characterization of Receptor for Mammalian Hepatopoietin That Is Homologous to Yeast ERV1

Hepatopoietin (HPO) is a novel polypeptide mitogen specific for hepatocytes and hepatoma cell lines, which is derived from liver and supports its regeneration. To determine whether HPO acts via a receptor-based signal transduction, recombinant human hepatopoietin was labeled by iodination and used to characterize its binding activity by specific displacement test and Scatchard analysis in primarily cultured rat hepatocytes and human hepatoma Hep-G2 cells. The binding was saturable and specific because it was replaceable by HPO but not by epidermal growth factor, transforming growth factor-α, or insulin. Scatchard analysis indicated the presence of a single class of high affinity receptor with dissociation constant (K d ) of 2 and 0.7 pm, and a receptor density of about 10,000 sites/cell and 55,000 sites/cell in the rat hepatocytes and human hepatoma cells, respectively. TheK d values were consistent with the half-maximum dose of HPO activity. Affinity cross-linking of the receptor with125I-HPO revealed a polypeptide of molecular mass approximately 90 kDa by SDS-polyacrylamide gel electrophoresis. Thus, the molecular mass of the HPO receptor was calculated to be about 75 kDa. These data demonstrated the existence of an HPO receptor in hepatocytes and hepatoma cells, which may account for biological effect.

Effects of enriched environment on morphine-induced reward in mice

Drug addiction and abuse have been extremely serious problems in our society. The effect of different environmental conditions on the susceptibility of human to drug addiction and abuse is still not fully understood. It was recently reported that enriched environment can trigger long-term modification in neural functions and might prevent the occurrence of pathogenic behaviors. Here we investigated the effects of enriched environment on morphine-induced reward in mice. We found that the enriched environment attenuated the acute morphine (10 mg/kg)-induced hyperlocomotion and repeated morphine (10 mg/kg)-induced behavioral sensitization. Moreover, the environmental condition blocked the conditioned place preference (CPP) of the mice induced by morphine (5 mg/kg). Associated with behavioral alterations, the expression of FosB-like proteins in the nucleus accumbens of the brain was down-regulated in the mice housed in the enriched environmental condition, but not in the standard environment. Taken together, these results indicate that enriched environmental condition leads to decrease in sensitivity of the mice to morphine-induced reward.

Functional characterization of novel human ARFGAP3.

ADP ribosylation factors (ARFs) are critical in the vesicular trafficking pathway. ARF activity is controlled by GTPase‐activating proteins (GAPs). We have identified recently a novel tentative ARF GAP derived from human fetal liver, ARFGAP3 (originally named as ARFGAP1). In the present study, we demonstrated that ARFGAP3 had GAP activity in vitro and remarked that the GAP activity of ARFGAP3 was regulated by phospholipids, i.e. phosphatidylinositol 4,5‐diphosphate as agonist and phosphatidylcholine as antagonist. ARFGAP3 is a predominantly cytosolic protein, and concentrated in the perinuclear region. Its transient ectopic overexpression in cultured mammalian cells reduced the constitutive secretion of secreted alkaline phosphatase, indicating that ectopic overexpression of ARFGAP3 inhibits the early secretory pathway of proteins in vivo. These results demonstrated that ARFGAP3 is a novel GAP for ARF1 and might be involved in intracellular traffic of proteins and vesicular transport as predicted.

Chronic corticosterone administration from adolescence through early adulthood attenuates depression-like behaviors in mice

There is evidence that depression may have a different neural basis at different ages. Although chronic stress and elevated glucocorticoid levels have been demonstrated to lead to the emergence of mood disorders, it remains unclear how moderate elevation of glucocorticoid levels in young animals influences depression-like behaviors and brain functions. To address this issue, the present study examines how chronic corticosterone (CORT) administration during adolescence and early adulthood influences depression-like behaviors, hypothalamic-pituitary-adrenal (HPA) axis response and hippocampal cell proliferation. Male mice were chronically administrated with CORT drinking water (20mg/L) during adolescence. After two months of treatment, serum CORT levels were measured using enzyme immunoassay. Hippocampal glucocorticoid and mineralocorticoid receptors were characterized using Western blot. Tail suspension and forced swim tests were used to assess depression-related behaviors in mice. Immunohistochemistry was performed to measure bromodeoxyuridine (BrdU) incorporation in order to assess cell proliferation in the hippocampus. Our results suggest that chronic CORT administration induced a mild but not significant elevation in basal CORT levels and attenuated the physiological responses to stress. Chronic CORT administration also reduced expression of the hippocampal mineralocorticoid receptor and decreased immobility time in both the tail suspension test and the forced swim test. Moreover, chronic CORT administration increased the BrdU immunoreactivities in the hippocampus. Taken together, these findings suggest that chronic mild elevation by CORT administration during the adolescence and early adulthood attenuates depression-like behaviors.

Antidepressive behaviors induced by enriched environment might be modulated by glucocorticoid levels

Exposure to enriched environment (EE) can influence expression of depression symptoms, however, the underlying mechanism has not been established, although neurogenesis was probably involved. It has been reported that EE stimulates glucocorticoids release. However, the role of corticosterone (CORT) in effects of EE is still unknown. To address these issues, we examined depression-like behaviors of the animals exposed to EE with low dose CORT supplement following bilateral adrenalectomy (ADX+CORT). Two months after housing, tail suspension test and forced swim test were used to assess depression-related behavior of mice. Serum CORT levels were measured by radio-immunoassay. Signals of DNA synthesis marker bromodeoxyuridine and immature neuronal marker doublecortin were measured by immunohistochemistry. Results showed that EE significantly decreased immobility time of the mice in both the tail suspension test and forced swim test, showing distinctive antidepressive behaviors. Exposure to EE also increased serum CORT level, and prevention of this increase with ADX+CORT eliminated the decrease of immobility time of the animals. Both the mice exposed to EE and those receiving ADX+CORT treatment showed enhanced newly born cells and immature granule neurons in the hippocampus. Taken together, our data suggest that glucocorticoids elevation is required for antidepressive behaviors of EE.

Characterization and tissue expression of a novel human gene npdc1.

We report the molecular characterization of a novel human homologue of mouse npdc1 (neural proliferation, differentiation and control, 1) gene, designated human npdc1 (hnpdc1). hnpdc1 was identified by large-scale sequencing of fetal liver cDNA libraries and the full-length cDNA was obtained by PCR amplification. The hnpdc1 gene, which contains nine exons, was mapped to human chromosome 15. It encodes a polypeptide of 325 amino acids, which shows high homology (77% identity) to the mouse NPDC1. Sequence analysis has shown that hNPDC1 protein contains a putative signal peptide of 34 amino acids, a transmembrane segment, and a typical bipartite nuclear localization signal. Northern blot and dot blot hybridization indicates that, just like mnpdc1, hnpdc1 mRNA is strongly expressed in adult brain (especially in hippocampus, frontal lobe and temporal lobe) and about 1.82-fold higher in adult brain than that in fetal brain. Unlike mnpdc1, however, hnpdc1 contains two transcripts instead of only one (1.5 kb), and has high expression levels in prostate, pituitary gland, and mammary glands. These results support that hNPDC1 plays a role in the control of neural cell proliferation and differentiation, and suggest that it may be involved in the development of several secretion glands.

Theophylline attenuates microwave-induced impairment of memory acquisition

Numerous studies have shown that acute microwave exposure causes cognitive deficits in animals, possibly via hyperthermia, but the biological effect of microwave exposure on memory processing is still unknown. The release of adenosine is demonstrated to be a general way for the cells to respond to metabolically stressful conditions such as hypoxia and ischemia. The present study aimed to examine whether adenosine mediates biological effects of microwave exposure on memory processing using a continuous multiple-trial inhibitory avoidance task. Results demonstrated that microwave exposure for 20 min before training impaired memory acquisition and retention performance in mice, assessed by the number of training trials and by latency to enter the dark compartment. The mice exposed to microwave radiation showed a dose-dependent hyperthermia. Moreover, the cell numbers of hippocampus were decreased in the mice receiving microwave exposure at an average power density of 50 mW/cm(2), indicating the anatomical correlation to hippocampal-amygdaloid structures corresponding with the memory disrupt of the mice. Administration of theophylline, a nonspecific adenosine receptor antagonist, 30 min before microwave exposure, completely antagonized the impairment of inhibitory avoidance acquisition but not retention. These results suggest that the adenosine regulation pathway was partially involved in microwave-induced impairment of inhibitory avoidance memory.