Decreased fasting serum glucogenic amino acids with a higher compared to normal protein diet during energy restriction in women: a randomized controlled trial.

Dietary protein alters circulating amino acid (AAs) levels and higher protein intake (HP) is one means of losing weight. We examined 34 overweight and obese women (57 ± 4 years) during 6 months of energy restriction (7.3 ± 3.8% weight loss) divided into groups consuming either normal protein (NP; 18.6 energy% protein) or HP (24.3 energy% protein). There was a reduction in fasting serum glucogenic AAs (p = 0.015) that also associated with greater weight loss (p < 0.05) in the HP group, but not in the NP group. These findings have implications for nutrient prioritization during energy restriction.

New treatment options for patients with metastatic prostate cancer.

Prostate cancer is one of the most common cancers in men. When metastasised (40% of patients), classic anti-androgen therapy is the first-line treatment. Usually, this treatment becomes ineffective when castration-resistant prostate cancer (CRPC) develops. Thus far, docetaxel was the only chemotherapeutic option that has shown to be able to extend overall survival and improve quality of life in these patients. Recently, cabazitaxel and abiraterone have shown significant survival benefits for patients progressive on or after docetaxel treatment, as did enzalutamide and radium-223. In North America, immune therapy (sipuleucel-T) became available for a subgroup of CRPC patients. These new treatment options will change the treatment paradigm of patients with metastatic castration resistant prostate cancer. A multidisciplinary approach by both medical oncologists and urologists seems mandatory.

Interaction between the BDNF gene Val/66/Met polymorphism and morning cortisol levels as a predictor of depression in adult women.

BACKGROUND: Common genetic variants, such as the brain-derived neurotrophic factor (BDNF) Val/66/Met polymorphism (rs6265), are known to interact with environmental factors such as early adversity to increase the risk of subsequent major depression. Much less is known about how they interact with individual differences in cortisol, although these also represent a risk for major depression. AIMS: To determine whether this BDNF variant moderated the risk represented by higher levels of morning salivary cortisol in adult women. METHOD: We recruited 279 premenopausal women who were at high risk of major depressive disorder because of either negative self-evaluation, unsupportive core relationship or chronic subclinical symptoms of depression or anxiety. Morning salivary cortisol was measured daily for up to 10 days at entry. Participants were followed up for about 12 months by telephone calls at 3-4 monthly intervals. Major depression and severe life events were assessed through interviews at baseline and follow-up; DNA was obtained from the saliva. RESULTS: There were 53 onsets (19%) of depressive episodes during follow-up. There was a significant U-shaped relationship between adjusted morning cortisol levels at baseline and the probability of depression onset during follow-up. In total, 51% experienced at least one severe life event/difficulty, and this strongly predicted subsequent onsets of depressive episodes. The BDNF Val/66/Met genotype was not directly associated with onsets of depression or with cortisol levels, but there was significant interaction between Val/66/Met and cortisol: the association between baseline cortisol and depression was limited to those with the Val/66/Val variant. There was no interaction between life events and either this BDNF polymorphism or cortisol levels. CONCLUSIONS: Morning salivary cortisol interacts with the BDNF Val/66/Met polymorphism in predicting new depressive episodes. This paper adds to the evidence that single gene polymorphisms interact with endogenous factors to predict depression.

Methods of outcomes measurement in nail psoriasis.

BACKGROUND: Nail involvement is a common feature of psoriasis, predicting higher disease severity and greater impairment in quality of life. Valid assessment of nail psoriasis is relevant for research and routine clinical use. However, no measurement standards have currently been developed. OBJECTIVE: To identify state-of-the-art outcomes measurements in nail psoriasis by literature analysis. METHODS: Systematic Web-based literature search, followed by structured critical appraisal and consecutive descriptive report. The search focused on methodological and epidemiological publications and papers describing outcomes of clinical trials on nail psoriasis. RESULTS: Initially, 646 publications met the primary criteria. After non-relevant or replicate publications were excluded, 66 papers were analysed, including clinical trials or case reports (n = 41), reviews (n = 11) and methodological or epidemiological studies (n = 14). In total, 23 clinical outcomes measures and 15 patient-reported outcomes (PRO) tools were used. None had been validated according to recent standards. In the studies with clinical interventions (n = 41), NAPSI (Nail Psoriasis Severity Index; n = 4) or target NAPSI (n = 2) were the most often used single tools, followed by Physician's Global Assessment (n = 3). However, in 16 studies, no specifically described outcomes measures were used. CONCLUSION: Valid clinical outcomes measures in nail psoriasis are rare. Existing tools lack validation and standardisation. A need exists for accurate and scientifically sound evaluation of nail psoriasis severity in trials and clinical practice. To cover all elements of nail psoriasis, the optimal nail psoriasis assessment tool would include both PRO and physician-assessed outcomes measures.

Leukocyte infiltration and mRNA expression of IL-20, IL-8 and TNF-R P60 in psoriatic skin is driven by TNF-alpha.

Anti-TNF-alpha therapy with a chimeric monoclonal antibody (Infliximab, Remicade) has been shown to be highly effective in the treatment of skin lesions as well as arthritis in patients with psoriatic arthritis. In this study we investigated the molecular consequences of the in vivo TNF-alpha blockade with infliximab in psoriatic skin lesions of 6 patients with severe psoriatic arthritis. Biopsies from lesional and non-lesional skin were taken before and 10 weeks after the initiation of treatment. Immunohistochemistry and semiquantative RT-PCR were performed focusing on proinflammatory gene products. Immunohistochemistry, after three infusions, revealed a marked decrease in the expression of TNF-alpha, HLA-DR, CD3, CD15, ICAM-1 and LFA-1 positive cells. By semiquantitative RT-PCR, we analysed mRNA expression of IL-8, IL-20, TNF-R (TNF-R p60 and TNF-R p80), IL-1R I and IL-1R II, as well as ICAM-2. Before therapy, m-RNA for IL-8, IL-20, TNF-R p60, TNF-R p80, IL-1R II and ICAM-2 were detected in lesional skin. mRNA expression of IL-8 and IL-20 completely disappeared and mRNA expression of TNF-R p60 was reduced after therapy. This effect on IL-8 expression was paralleled by a decreased infiltration of leukocytes in psoriatic skin. These data suggest that the clinical response of anti-TNF-alpha therapy in patients with psoriasis or psoriatic arthritis may be, at least in part, caused by the inhibition of the production of proinflammatory cytokines and by the decreased expression of adhesion molecules with the consequence of an impaired migration of proinflammatory cells into the inflamed tissue. These data further support a critical role for TNF-alpha in the pathology of psoriasis.

Work-family fit: voices of parents of children with emotional and behavioral disorders.

Employed parents of children with emotional or behavioral disorders overcome significant challenges to fulfill their daily work-family responsibilities; however, their experiences in meeting these multiple demands have not been examined. The purpose of the present study was to describe the strategies these parents use and their perceptions about how caregiving and employment responsibilities can successfully fit together. Findings of five focus groups conducted with 41 employed parents whose children had been diagnosed with mental health problems are presented. Qualitative analysis was used to identify major themes across the areas of employment responsibilities and adaptations, child care arrangements, and achieving fit between job and home. Results revealed that parents experience a serious lack of community-based services and resources necessary to support work and family obligations in a meaningful way. The data suggest a need for more research and services development to support these families in their communities.

NAD degradation and regulation of CD38 expression by human monocytes/macrophages.

In recent years, evidence has accumulated that NAD+ serves as a precursor of metabolites that are involved in a number of regulatory processes. In this work we show that extracellularly added NAD+ was rapidly degraded by intact human monocytes to nicotinamide and ADP-ribose. Besides these main products, minor amounts of AMP, ADP and cADP-ribose were formed. Expression of CD38, which has been identified as NAD+-glycohydrolase (EC 3.2.2.6) degrading NAD+ into nicotinamide and ADP-ribose, was determined on freshly isolated human monocytes by flow cytometry and RT-PCR. Upon ligation with anti-CD38 mAb, CD38 underwent internalization, shedding and new expression. As monocytes possess an intracellular CD38 pool, it could serve as a source for newly expressed CD38. Differentiation of monocytes to macrophages resulted in down-regulation of surface expression of CD38. This decrease correlates with a reduction in NADase activity, indicating that the amount of functional active CD38 molecules decrease during differentiation. As CD38 mRNA was found to be diminished in macrophages, regulation of the gene product seems to occur at the level of transcription or mRNA stability.

Reversal of established delayed type hypersensitivity reactions following therapy with IL-4 or antigen-specific Th2 cells.

Delayed-type hypersensitivity reactions (DTHR) are mediated by IFN-gamma-producing CD4+ (Th1) or CD8+ T cells (Tc1) and can be prevented by steering T cells toward an IL-4-producing Th2 or Tc2 phenotype. It is currently accepted that T cells can be directed toward a Th2 or Tc2 phenotype only during the initiation of an immune response. Once established, the cytokine pattern of immune reactions is believed to be stable. Therefore, inhibition of DTHR by the induction of Th2/Tc2 responses, termed immune deviation, is considered only as a prevention but not as a therapy of harmful DTHR. Here we demonstrate that therapeutic immune deviation can reverse established contact hypersensitivity (CHS), a Th1/Tc1-mediated DTHR. One or two weeks after induction of CHS, mice received either a single cycle of IL-4 therapy or adoptive transfer of antigen-specific Th2 cells. This treatment generated a novel state of immunity that provided long-lasting protection against tissue destruction and neutrophil recruitment during subsequent antigen exposures. Therapeutic immune deviation of established CHS was dependent on CD4+ T cells and the induction of endogenous IL-4 synthesis. Thus, a population of immunoregulatory Th2 cells persists during advanced inflammatory responses that can be used for therapeutic deviation of established DTHR.

Biphasic cytotoxic mechanism of extracellular ATP on U-937 human histiocytic leukemia cells: involvement of adenosine generation.

Since extracellular ATP can exhibit cytotoxic activity in vivo and in vitro, its application has been proposed as an alternative anticancer therapy. In this study we investigated the mechanisms of ATP-induced cytotoxicity in a human leukemic cell line (U-937). ATP added as a single dose exceeding 50 microM was cytostatic or even cytotoxic for U-937 cells. Interestingly, growth inhibition by ATP (50-3500 microM) showed a biphasic dose response. Up to 800 microM, ATP was cytotoxic in a dose-dependent manner (EC(50) 90 microM). In a range between 800 and 2500 microM, cell count was markedly higher despite the higher ATP concentrations. The cytotoxic effect of ATP could be antagonized by addition of uridine as a pyrimidine source and, alternatively, by addition of the nucleoside transmembrane inhibitor dipyridamole. The apoptosis-inducing adenosine A(3) receptor was not involved in measurable quantities, since (1) adenosine did not lead to an elevation of intracellular calcium levels, and (2) an unselective A(1-3) antagonist (ULS-II-80) could not abrogate the cytotoxic effect. Experiments monitoring extracellular nucleotide metabolism confirmed the assumption that the long-term production and continuous uptake of adenosine, which is extracellularly generated by degradation of ATP, led to an intracellular nucleotide imbalance with pyrimidine starvation. The biphasic dose response to higher ATP concentrations could be explained by the rapid degradation of lower ATP concentrations (300 microM) to adenosine by serum-derived enzymes, whereas higher concentrations (900 microM) only produced small amounts of adenosine due to forward inhibition of AMP hydrolysis by prolonged high ADP levels. FACS analysis revealed that at lower adenosine concentrations (300 microM) a reversible G(1) phase arrest of the cell cycle was induced, whereas higher concentrations (1000 microM) triggered apoptosis. Considering ATP as a potential cytostatic drug, our data have important implications concerning metabolic interactions of administered nucleotides.

Treatment of psoriatic arthritis with antitumour necrosis factor-alpha antibody clears skin lesions of psoriasis resistant to treatment with methotrexate.

BACKGROUND: In inflamed skin, keratinocytes and inflammatory cells both produce large amounts of tumour necrosis factor (TNF) -alpha, a cytokine with broad effects that are relevant to inflammation. Blockade of this proinflammatory cytokine by a monoclonal anti-TNF-alpha antibody might be effectively used in the treatment of inflammatory skin diseases. OBJECTIVES: To gather information about the efficacy of an anti-TNF-alpha antibody (infliximab) in the treatment of skin lesions of psoriatic arthritis. METHODS: Six patients with progressive joint disease and psoriatic skin lesions unresponsive to methotrexate therapy were treated with anti-TNF-alpha antibody. The Psoriasis Area and Severity Index was determined before and 10 weeks after initiation of therapy. RESULTS: Improvement of psoriatic skin lesions was observed in all patients. In addition, a marked improvement of the joint disease was noted. CONCLUSIONS: Therapy with anti-TNF-alpha antibody may be an effective treatment regimen for both psoriatic arthritis and psoriatic skin lesions.

Improved short-term spatial memory but impaired reversal learning following the dopamine D(2) agonist bromocriptine in human volunteers.

RATIONALE: Studies in humans of cognitive effects of dopaminergic drugs have largely focused on tasks of working memory, with a few studies also examining executive function. OBJECTIVES: This study was designed to investigate the effects of 1.25 mg of the dopamine D(2) agonist bromocriptine on spatial working memory, planning and discrimination reversal learning in young healthy volunteers. METHODS: Twenty volunteers were tested in a double-blind, placebo-controlled, cross-over design. The cognitive assessment included tests taken from the Cambridge Neuropsychological Test Automated Battery (CANTAB) designed to test visuo-spatial recognition memory and spatial working memory. In addition, tests of spatial planning and discrimination reversal learning were used to assess the more general effects of bromocriptine. Tests of subjective feelings and motivation were also incorporated into the battery. RESULTS: Bromocriptine enhanced the spatial memory span of subjects, whilst impairing their ability to reverse a learned probabilistic discrimination. Tests of recognition memory and planning were unaffected by the drug. The findings were not explained by changes in subjective mood or motivational measures. CONCLUSIONS: The pattern of findings observed here mirror medication-dependent observations seen in Parkinson's disease. The results are discussed with reference to the different anatomical networks known to subserve performance of the differentially affected tasks.

Expression of purinergic receptors (ionotropic P2X1-7 and metabotropic P2Y1-11) during myeloid differentiation of HL60 cells.

The expression of human purinergic P2 receptors (P2X1-7 and P2Y1-11) as well as the ecto-enzymes apyrase (CD39) and 5'-nucleotidase (CD73) was investigated on the nucleic acid level during granulocytic and monocytic differentiation of HL60 cells and on peripheral human blood leukocytes. RT-PCR and dot-blot hybridization assays indicated that mRNA transcripts of all analyzed P2 receptors apart from the P2X3 receptor were expressed during myeloid development of HL60 cells, showing a distinct regulation during the course of differentiation. In blood leukocytes, transcripts of P2X5, P2X7 and all P2Y receptors, except for P2Y6, receptor were found. CD39 and CD73 showed a marked upregulation during myeloid maturation. Functional analysis of P2 receptor-mediated intracellular Ca(2+)-increase after stimulation with ATP revealed no change during granulocytic differentiation, but showed a strong attenuation in both potency and efficacy during monocytic development of HL60 cells.

Immature dendritic cells generated with low doses of GM-CSF in the absence of IL-4 are maturation resistant and prolong allograft survival in vivo.

Dendritic cells (DC) were cultured from mouse bone marrow (BM) progenitors in low concentrations of granulocyte-macrophage colony-stimulating factor (GM-CSF) (GM(lo) DC) by two different protocols. The phenotype and functional properties of these GM(lo) DC were compared to those of standard BM-DC cultures generated in high concentrations of GM-CSF (GM(hi) DC) or in low GM-CSF plus IL-4 (GM(lo)/IL-4 DC). An effect of IL-4 on maturation was observed only at low but not high doses of GM-CSF. Compared to mature DC, GM(lo) DC were phenotypically immature, weak stimulators of allogeneic and peptide-specific T cell responses, but substantially more potent in presentation of native protein. Immature GM(lo) DC were resistant to maturation by lipopolysaccharide, TNF-alpha or anti-CD40 monoclonal antibodies, as the expression of co-stimulatory molecules was not increased, and stimulatory activity in oxidative mitogenesis was not enhanced. These maturation-resistant immature GM(lo) DC induced T cell unresponsiveness in vitro and in vivo. GM(lo) DC also prolonged haplotype-specific cardiac allograft survival (from 8 days to >100 days median survival time) when they were administered 7 days (but not 3, 14 or 28 days) before transplantation. Our findings may have important implications for future studies in T cell tolerance induction in vivo.

Ecto-diadenosine 5',5'''-P1,P4-tetraphosphate (Ap4A)-hydrolase is expressed as an ectoenzyme in a variety of mammalian and human cells and adds new aspects to the turnover of Ap4A.

Ap4A and other dinucleotides participate in the regulation of hemostasis and blood pressure control. With the exception of two previously reported surface anchored ectoAp4A-hydrolases on bovine aortic endothelial and chromaffine cells, all Ap4A-hydrolases reported are intracellular or freely soluble. We demonstrated that ectoAp4A-hydrolases are present on a broad variety of cell types of different species: rat mesangial, bovine corneal epithelial, human Hep-G2 and peridontal cells. Ectoenzyme properties were evaluated on rat mesangium cells. Chromatography of purified plasma membranes on Sephacel 300 resulted in enrichment of ectoAp4A-hydrolase and in separation from ectoATPase. In contrast to ATPase, Ap4A-hydrolase was stable at room temperature. EctoAp4A-hydrolase also recognized ATP as substrate, and therefore is not highly specific. The molecular weight was 180 kD. Unlike ectoAMPase ectoAp4A-hydrolase was not attached via a glycosyl-phosphatidylinositol (GPI)-moiety. Concentrations of PI-PLC 10-100-fold higher than effective for ectoAMPase cleavage (10-100 mU/ml) plus extensively extended incubation times up to eight hours did not result in cleavage of ectoAp4A-hydrolase. The enzyme ectoAp4A-hydrolase might presage a direction for pharmaceutical manipulation in the control of blood pressure and hemostasis.

Human monocyte derived dendritic cells express functional P2X and P2Y receptors as well as ecto-nucleotidases.

We investigated the expression and function of P2 receptors and ecto-nucleotidases on human monocyte derived dendritic cells (DC). In addition we analyzed the effect of extracellular ATP on the maturation of DC. By RT-PCR, DC were found to express mRNA for several P2X (P2X1, P2X4, P2X5, P2X7) and P2Y (P2Y1, P2Y2, P2Y4, P2Y5, P2Y6, P2Y10, P2Y11) receptors. As shown by FURA-2 measurement, triggering of P2 receptors resulted in an increase in free intracellular Ca2+. In combination with Tumor necrosis factor-alpha, ATP increased the expression of the DC surface markers CD80, CD83 and CD86 indicating a maturation promoting effect. DC expressed the ecto-apyrase CD39 and the ecto-5'-nucleotidase CD73 as demonstrated by RT-PCR. Extracellular ATP was rapidly hydrolyzed by these ecto-enzymes as shown by separation of 3H-labeled ATP metabolites using a thin layer technique. These data suggest that ATP acts as a costimulatory factor on DC maturation.

Absence of severe systemic toxicity after leakage-controlled isolated limb perfusion with tumor necrosis factor-alpha and melphalan.

BACKGROUND: Severe systemic toxicity and hemodynamic changes after isolated limb perfusion (ILP) with tumor necrosis factor-alpha (TNF-alpha) and melphalan, with or without interferon-gamma, have been reported in several series. We studied whether these side effects could be precluded by preventing leakage from the isolated circuit into the systemic circulation. METHODS: Clinical and pharmacokinetic data for 20 consecutive patients with recurrent melanoma of the limbs who were treated by ILP with TNF-alpha (3-4 mg) and melphalan, with or without interferon-gamma, were studied. Leakage rates and TNF-alpha levels were determined during and after ILP and were correlated with systemic toxicity and hemodynamic changes. RESULTS: Only two patients experienced leaks (2% and 13%) during ILP. For 18 patients without leakage, the mean peak systemic TNF-alpha level was 2.8 ng/ml at 10 minutes after ILP. After leakage, the peak systemic TNF-alpha levels were 31.9 and 88.3 ng/ml at 5 minutes. Toxicity was mild and consisted mainly of fever (n = 17) and nausea/vomiting (n = 19) during the first day after ILP. Some patients developed tachycardia (n = 6), hypotension (n = 3; responding immediately to fluid challenge), a decrease in the WBC count (n = 3; grade I) or thrombocyte count (n = 11; grade I/II, no hemorrhage or therapeutic intervention), or hepatotoxicity [cytolysis (n = 15; 14 grade I/II and 1 grade IV) or hyperbilirubinemia (n = 7; grade I/II, all resolving spontaneously)]. Patients with tachycardia or hepatotoxicity exhibited significantly higher TNF-alpha levels after ILP, compared with other patients. CONCLUSIONS: Systemic toxicity after ILP with TNF-alpha is minimal and does not differ from that after ILP with melphalan alone when leakage is adequately controlled.

Presence of multiple functional polyadenylation signals and a single nucleotide polymorphism in the 3' untranslated region of the human serotonin transporter gene.

The human serotonin transporter (hSERT) gene is a candidate for involvement in the aetiology of affective disorders. In humans, multiple transcripts of the gene have been detected by northern blot analysis of brain and other tissues. We performed 3' rapid amplification of cDNA ends to identify the common sites of polyadenylation in hSERT mRNA from human JAR cells and whole blood. Two major polyadenylation sites were identified: one 567 bp downstream of the stop codon, consistent with the usage of the polyadenylation signal AATGAA, and a second site 690 bp downstream of the stop codon. The putative polyadenylation signal upstream of this site contained a single nucleotide polymorphism (AG/TTAAC). However, allelic variation at this site did not influence polyadenylation site usage, and there were no significant differences in the abundance of the two alleles of this polymorphism between 329 control subjects, 158 individuals with major depression, and 130 individuals with bipolar affective disorder. This single nucleotide polymorphism in the 3' untranslated region of the hSERT gene should provide a useful genetic marker in the evaluation of hSERT as a candidate gene influencing susceptibility to mood disorders.