The dopamine transporter is the primary means of inactivating synaptic dopamine as well as a major site of action for psychostimulants (such as cocaine and amphetamine) and for neurotoxins that induce parkinsonism. In the present study, a human dopamine transporter partial cDNA clone obtained by polymerase chain reaction exhibited 87% and 89% identity at the nucleic acid and amino acid levels, respectively, with transmembrane domains 3-5 of the rat homolog. This clone was used to quantitate human dopamine transporter mRNA by nuclease protection assay. The postmortem content of dopamine transporter mRNA in the substantia nigrae of 18-to 57-yr-old subjects was relatively constant, while in subjects >57 yr old, a precipitous (>95%) decline in substantia nigra dopamine transporter mRNA was evident. In contrast, tyrosine hydroxylase mRNA in the same samples declined in a linear manner with increasing age. In situ hybridization experiments confirmed the profound loss of dopamine transporter gene expression in melanin-positive (presumptive dopamine) nigral neurons. These data may begin to shed light on compensatory changes occurring in human dopamine neurons during normal aging.The actions of many neurotransmitters are terminated by rapid reuptake into presynaptic nerve endings via neurotransmitter-specific, high-affinity, Na+-dependent membrane transporter proteins. The transporter for the neurotransmitter dopamine (DA) is apparently the receptor through which psychostimulants such as cocaine and amphetamine exert their potent reinforcing properties, leading to psychostimulant abuse and dependence (1). In addition, the DA transporter mediates the active accumulation of neurotoxins such as 1-methyl-4-phenylpyridine into DA neurons (2, 3), resulting in DA cell death and a parkinsonian syndrome. Thus, understanding the regulation of human DA transporter gene expression is an important step toward elucidation of the molecular bases of psychostimulant drug abuse and of neurotoxin-induced (and quite possibly idiopathic) parkinsonism.Ligand binding to the human DA transporter decreases with age (4, 5), in keeping with other evidence for an age-related loss of DA neurons (6). Nevertheless, surprisingly little is known about the nature of normal age-related changes in human DA neurons, a process apparently distinct from the neuronal changes seen in Parkinson disease (6, 7). In the present study, a human DA transporter cDNA clone was obtained by polymerase chain reaction, exploiting predicted sequence homology with the recently cloned, functionally related human norepinephrine transporter (8). With the use of this clone, a precipitous, age-related loss of DA transporter mRNA was observed in human substantia nigra, while tyrosine hydroxylase mRNA, another phenotypic marker of DA neurons, decreased more linearly with age. These data may shed light on some compensatory changes occurring in human DA neurons during the normal aging process. MATERIALS AND METHODSHuman substantia nigrae used for biochemical analyses (n = 36) w...

BackgroundThe aim of this study was to evaluate the outcomes of patients with inflammatory breast cancer (IBC), with emphasis on the role of molecular subtypes and radiotherapy.MethodsA retrospective cohort study to investigate overall survival (OS) and breast cancer-specific mortality (BCSM) in patients with IBC was conducted using data obtained by the Surveillance, Epidemiology, and End Results (SEER) program from 2010–2013. Cox multivariate regression was used to calculate the adjusted Hazard Ratios (aHR).Results403 patients were eligible for this study. Patients in the group with hormone receptors (HR)+/HER2- subtype had an OS of 79.6% compared with 89.0 % in the group with (HR)+/HER2+ subtype and 76.8% in the HR-/HER2+ group and 62.9% in the triple-negative (TN) group. BCSM was 16.3% for the HR+/HER2- group, 9.8% for the HR+/HER2+ group, 21.7% for the HR-/HER2+ group, and 30.5% for the TN group. For distant metastases, the results showed that there was a high probability of bone metastasis in HR-positive groups, brain and liver metastasis in HER2-positive groups, and lung metastasis in the TN group. Multivariate analysis demonstrated that estrogen receptor and HER2 positivity were associated with better survival and that the TN subtype had a poorer OS and BCSM compared with other subtypes (P<0.05). Furthermore, patients who received radiotherapy were more likely to have improved survival (P< 0.05).ConclusionInflammatory breast cancer appears to alter the prognosis in association with the receptor status and molecular subtypes. Radiotherapy was still considered to be a crucial treatment for patients with IBC.

Parkinson's disease (PD) patients vary widely in their response to levodopa treatment, and this variation may be partially genetic in origin. We determined whether particular dopamine and opioid receptor polymorphisms were associated with risk of earlier onset of dyskinesia side effects during levodopa therapy. Smoking status was also examined. The 92 subjects were recruited from the movement disorders clinic of a neurology practice associated with a medical school. All were adult-onset PD patients who had been taking levodopa at least 5 years and/or had developed levodopa-induced dyskinesia. Carrying the G-allele of the A118G single nucleotide coding region polymorphism of the mu opioid receptor, as well as a history of never smoking, were independently associated with increased risk of earlier onset of dyskinesia (P=0.05 and 0.02, respectively). One genotype of the D2 dopamine receptor intronic dinucleotide repeat polymorphism (14 repeats/15 repeats, with frequency of 6%) was also associated with earlier dyskinesia (P=0.003). History of smoking has previously been associated with reduced risk of developing PD. Our results suggest that smoking history may also influence the response to levodopa, with contribution comparable to those of individual genes including the mu opioid receptor and D2 dopamine receptor.

Although our previous studies found Pre-B-cell colony-enhancing factor (PBEF) as a highly up-regulated gene in acute lung injury that could stimulate expressions of other inflammatory cytokines, the underlying molecular mechanisms remain to be fully elucidated. Growing evidence indicates that PBEF is a nicotinamide phosphoribosyltransferase involved in the mammalian salvage pathway of NAD synthesis. This study was designed to determine whether the effect of PBEF to stimulate expressions of inflammatory cytokines depends on its enzymatic activity. We prepared two human PBEF mutant (H247E and H247A) recombinant proteins and overexpressing constructs for their overexpressions in A549 cells and confirmed that enzymatic activities of both mutants were nearly or completely abolished. Two mutants stimulated interleukin-8 (IL-8) expression at both the mRNA level and protein level just as equally effective as the wild-type PBEF did. These effects were due to the increased transcription, not the mRNA stability, of the IL-8 gene. Reporter gene assays and gel shift experiments indicated that AP-1 transcription factor is required to mediate these effects. SB203580, a p38 MAPK pathway inhibitor, and JNK inhibitor 1 can attenuate these effects. Both PBEF mutants similarly stimulated the expression of two other inflammatory cytokines: IL-16 and CCR3. These results indicate that PBEF stimulated expression of IL-8, IL-16, and CCR3 via its non-enzymatic activity. This effect is AP-1-dependent, in part via the p38 MAPK pathway and the JNK pathway. This finding reveals a new insight, which may manifest a novel role of PBEF in the pathogenesis of acute lung injury and other inflammatory disorders. Acute lung injury (ALI)2 and its more severe form, acute respiratory distress syndrome (ARDS), are characterized by inflammation of the lung parenchyma leading to impaired gas exchange with concomitant systemic release of inflammatory mediators causing inflammation, hypoxemia, and frequently resulting in multiple organ failure (1, 2). Although ALI/ARDS was first described in 1967 by Ashbaugh et al. (3), its mortality and morbidity remain high (4). More studies are warranted to elucidate its molecular pathogenesis and to identify new diagnostic and therapeutic targets to ALI/ARDS.In our previous study on animal models of ALI and human patients with ARDS, we identified pre-B-cell colony-enhancing factor (PBEF) as a biochemical and genetic marker in ALI (5). Our findings were confirmed and extended in a separate and larger population (Ͼ1000 patients) by Bajwa et al. (6). In further studies, we demonstrated that overexpression of PBEF can augment the expression of inflammatory cytokines and dysregulate pulmonary cell barrier function, whereas inhibition of PBEF expression by its cognate small interference RNA has the opposite effects (7-9). PBEF has been confirmed as a nicotinamide phosphoribosyltransferase (Nampt) involved in the mammalian salvage pathway of NAD synthesis (10,11). Whether the enzymatic activity of PBEF is involved in its effe...

No abstract

Dopamine transporter mRNA levels in the rat substantia nigra were quantified using a sensitive nuclease protection assay with a highly homologous human dopamine transporter cDNA clone. The same probe was also used to visualize dopamine transporter mRNA in the substantia nigra by in situ hybridization. Repeated cocaine administration (15 mg/kg, twice a day for 6.5 days) resulted in a greater than 40% decrease in nigral dopamine transporter mRNA levels. In contrast, dopamine transporter mRNA levels were unchanged after either acute treatment (4 h before death) or repeated cocaine treatment followed by a 72-h withdrawal period. Thus, blockade of the dopamine transporter by repeated cocaine administration may result in the down-regulation of dopamine transporter gene expression in dopamine neurons.

Background/Aims: Chronic renal allograft dysfunction (CRAD) is a leading cause of long-term renal allograft loss. Oxidative stress may account for the nonspecific interstitial fibrosis and tubular atrophy that occur in CRAD. An antioxidant intervention via Nrf2 signaling pathway activation might be a promising therapy for some kidney diseases. The present paper investigates whether there is an association between oxidative stress alleviation via sulforaphane-induced Nrf2-HO-1/NQO-1 signaling pathway activation and CRAD improvement. Methods: F344 rat kidneys were orthotopically transplanted into Lewis rat recipients to establish CRAD models. Sulforaphane was administered at 1.5 mg/kg intraperitoneally once daily. Renal function and 24-hour urinary protein were monitored for variations for 24 weeks after transplantation. After 24 weeks, renal histopathology was evaluated according to the Banff criteria after hematoxylin and eosin, Masson’s trichrome and periodic acid-Schiff stainings. Additionally, intrarenal oxidative stress was assessed by the indicators malondialdehyde, 8-isoprostane, oxidized-low density lipoprotein and 8-hydroxy-2’-deoxyguanosine, as well as the activity levels of the antioxidant enzymes total superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase and γ-glutamylcysteine synthetase. Nrf2, HO-1 and NQO-1 expression levels were determined via immunohistochemical and Western blot analyses. Results: The sulforaphane-induced Nrf2-HO-1/NQO-1 signaling pathway activation, as demonstrated by immunohistochemical and Western blot analyses, delayed the progression of serum creatinine and blood urea nitrogen, particularly lowering the 24-hour urinary protein levels of CRAD. The semi-quantified histopathological changes were also alleviated. Evidence of oxidative stress alleviation, as indicated by a concurrent decrease in the indicators and sustained levels of antioxidant enzymes activity, was found in the renal allografts after sulforaphane intervention. Conclusion: Oxidative stress alleviation caused by continuous sulforaphane-induced Nrf2-HO-1/NQO-1 signaling pathway activation is associated with functional and morphological improvements of CRAD.

BackgroundThe genetic polymorphisms of glutathione S-transferase (GSTs) have been suspected to be related to the development of lung cancer while the current results are conflicting, especially in the Chinese population.MethodsData on genetic polymorphisms of glutathione S-transferase Mu 1 (GSTM1) from 68 studies, glutathione S-transferase theta 1 (GSTT1) from 17 studies and GSTM1-GSTT1 from 8 studies in the Chinese population were reanalyzed on their association with lung cancer risk. Odds ratios (OR) were pooled using forest plots. 9 subgroups were all or partly performed in the subgroup analyses. The Galbraith plot was used to identify the heterogeneous records. Potential publication biases were detected by Begg's and Egger's tests.Results71 eligible studies were identified after screening of 1608 articles. The increased association between two vital GSTs genetic polymorphisms and lung cancer risk was detected by random-effects model based on a comparable heterogeneity. Subgroup analysis showed a significant relationship between squamous carcinoma (SC), adenocarcinoma (AC) or small cell lung carcinoma (SCLC) and GSTM1 null genotype, as well as SC or AC and GSTT1 null genotype. Additionally, smokers with GSTM1 null genotype had a higher lung cancer risk than non-smokers. Our cumulative meta-analysis demonstrated a stable and reliable result of the relationship between GSTM1 null genotype and lung cancer risk. After the possible heterogeneous articles were omitted, the adjusted risk of GSTs and lung cancer susceptibility increased (fixed-effects model: ORGSTM1 = 1.23, 95% CI: 1.19 to 1.27, P<0.001; ORGSTT1 = 1.18, 95% CI: 1.10 to 1.26, P<0.001; ORGSTM1-GSTT1 = 1.33, 95% CI: 1.10 to 1.61, P = 0.004).ConclusionsAn increased risk of lung cancer with GSTM1 and GSTT1 null genotype, especially with dual null genotype, was found in the Chinese population. In addition, special histopathological classification of lung cancers and a wide range of gene-environment and gene-gene interaction analysis should be taken into consideration in future studies.