Pre-synaptic nicotinic and D2 receptors functionally interact on dopaminergic nerve endings of rat and mouse nucleus accumbens

The existence of pre-synaptic auto- and hetero receptors which modulate neurotransmitter release is well documented. Emerging evidence show that in some cases these pre-synaptic receptors may also cross-talk with each other. The aim of the present work was to investigate whether acetylcholine receptors (nAChRs) and dopamine (DA) autoreceptors, which are both able to modulate DA release, functionally interact on the same nerve endings. We used rat and mouse nucleus accumbens synaptosomes pre-labeled with [³H]DA and exposed to nicotinic and dopaminergic receptor ligands. Both nicotinic agonists and 4-aminopyridine (4-AP) provoked [³H]DA release which was inhibited by quinpirole and blocked by sulpiride and raclopride. Both the inhibitory effect of quinpirole and the stimulatory effect of (−)nicotine did not change when the nAChRs or the DA receptors were desensitized. (−)Nicotine and 4-AP were able to stimulate [³H]DA overflow also in mouse synaptosomes and this overflow was partially inhibited by quinpirole. In the β₂ knockout mice quinpirole was still able to inhibit the [³H]DA overflow elicited by 4-AP. To conclude: in rat and mouse the (−)nicotine evoked-release can be modulated by D₂/D₃ autoreceptors present on the DA terminals and nAChRs function is independent from D₂/D₃ autoreceptors which themselves may function independently from the activation of nAChRs.     

Therapeutic efficacy of ipilimumab,an anti-CTLA-4 monoclonal antibody,in patients with metastatic melanoma unresponsive to prior systemic treatments: clinical and immunological evidence from three patient cases

The management of unresectable metastatic melanoma is a major clinical challenge because of the lack of reliably effective systemic therapies. Blocking cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) has recently been proposed as a strategy to enhance cell-mediated immune responses to cancer, and clinical trials have demonstrated that anti-CTLA-4 therapy can produce durable outcomes with different response patterns than cytotoxic chemotherapy. We enrolled eight out of 155 patients with advanced melanoma in a multicentre phase II trial that evaluated the activity and tolerability of ipilimumab, a fully human, anti-CTLA-4 monoclonal antibody (; NCT00289627; CA184-008). Here we report our experience with three of these patients, who experienced progressive disease after a variety of previous therapies, including prior immunotherapies, and who achieved good outcomes with ipilimumab. One patient had a partial response ongoing at 17+ months on ipilimumab despite failure with four prior therapies, and the other two patients showed durable stable disease, both still ongoing at 17+ and 20+ months, respectively. The patient achieving a partial response experienced no side effects while receiving ipilimumab. The other two patients developed immune-related adverse events (irAEs) including rash (one case; grade 2) and diarrhoea (both cases; grades 1 and 2, respectively); the histopathology of colon biopsy samples from both was suggestive of colitis, with an abundant CD8+ T-cell infiltrate. Nausea, vomiting and acute pancreatitis were also observed in one patient. In addition, immunohistochemical findings of a dense CD8+, TIA1+ and granzyme B+ lymphoid infiltrate within a biopsied lesion provide indirect evidence of functional T-cell activation induced by treatment. These case reports highlight the potential for anti-CTLA-4-based therapy in previously treated patients with advanced melanoma. Moreover, because the patterns of response to ipilimumab differ from chemotherapy, we need to understand how and when patients may respond to treatment so that appropriate clinical decisions can be made.     

Reproductive performance of swine females re-serviced after return to estrus or abortion

The objective of this study was to analyze reproductive performance in swine females re-serviced after return to estrus or abortion in comparison with females in first service (gilts or weaned females). Records used were obtained from four commercial sow herds in Brazil including 24,194 mating records from PigCHAMP research database. Three mating categories (first service in gilts or weaned sows, re-serviced after return to estrus and re-serviced after abortion) were considered for the analysis. The farrowing rate (FR) was less and return to estrus (RER), abortion rate (ABR) and total born (TB) were greater in the category re-serviced after return to estrus compared to first service category (P<0.05). The category re-serviced after abortion only differed from the first service category by a greater ABR (P<0.05). In gilts and PO2-5 females re-serviced after a return to estrus, the FR was less (72.0% and 83.2%) and RER was greater (22.3% and 12.5%) compared to first service PO2-5 sows (92.7% and 5.3%; P<0.05). A re-service after a return to estrus did not affect TB in PO > or =2 females (P>0.05) but resulted in less TB in gilts and greater TB in primiparous sows (P<0.05). In females re-serviced after a return to estrus the performance was similar (P>0.05) between the two intervals considered as regular return to estrus (18-24 days and 36-48 days). Among the intervals considered as irregular return to estrus, greater FR was observed in intermediate (25-35 days) than in early (11-17 days) or late (>48 days) intervals. The re-service after a return to estrus results in an impaired farrowing rate, with a greater impact on gilts than at older parities. Females re-serviced after abortion are more predisposed to the recurrence of this reproductive failure.     

Response of the carotenoidless mutant Rhodobacter sphaeroides growing cells to cobalt and nickel exposure

The interaction of cobalt (Co²⁺) and nickel (Ni²⁺) ions with whole cells of the photosynthetic purple bacterium Rhodobacter sphaeroides strain R26 was investigated. Active and passive uptakes were examined in cells grown in the presence of increasing amounts of Co²⁺ and Ni²⁺. Inductively coupled plasma atomic emission spectroscopy (ICP-AES), pH titration, and attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy were used to assess the role of cell envelope and metabolism in accumulating the two heavy metals. The chosen microorganism was able to uptake cobalt and nickel up to 2.2 and 0.25 mg per gram of dried cells respectively, with the largest part found bound to the cell surface. Carboxylate groups lying on the cell wall of this Gram-negative bacterium proved to be the major candidates for binding protons and metal cations. Co²⁺ was found to interfere with Mg²⁺ extracellular immobilization and transport across the membrane, indicating that these ions share binding sites on the cell envelope and ion transport systems. According to the presence of a competition mechanism, bacterial growth experiments showed that high Mg²⁺ concentrations are able to rescue R. sphaeroides from Co²⁺ toxicity.     

Isolation and expression analysis of low temperature-induced genes in white poplar (Populus alba)

Poplar is an important crop and a model system to understand molecular processes of growth, development and responses to environmental stimuli in trees. In this study, we analyzed gene expression in white poplar (Populus alba) plants subjected to chilling. Two forward suppression-subtractive-hybridization libraries were constructed from P. alba plants exposed to low non-freezing temperature for 6 or 48 h. Hundred and sixty-two cDNAs, 54 from the 6-h library and 108 from the 48-h library, were obtained. Isolated genes belonged to six categories of genes, specifically those that: (i) encode stress and defense proteins; (ii) are involved in signal transduction; (iii) are related to regulation of gene expression; (iv) encode proteins involved in cell cycle and DNA processing; (v) encode proteins involved in metabolism and energetic processes; and (vi) are involved in protein fate.Different expression patterns at 3, 6, 12, 24, 48 h at 4 °C and after a recovery of 24 h at 20 °C were observed for isolated genes, as expected according to the class in which the gene putatively belongs. Forty-four of 162 genes contained DRE/LTRE cis-elements in the 5′ proximal promoter of their orthologs in Populus trichocarpa, suggesting that they putatively belong to the CBF regulon. The results contribute new data to the list of possible candidate genes involved in cold response in poplar.     

Plasma nociceptin/orphanin FQ levels rise after spontaneous episodes of angina, but not during induced myocardial ischemia

The aim of our study was to evaluate the effects of repeated episodes of angina and induced myocardial ischemia on plasma nociceptin/orphanin FQ (N/OFQ) levels. Patients with unstable angina (23 with new onset severe angina or accelerated angina and 18 with subacute angina at rest) who had had repeated spontaneous episodes of chest pain in the last week before the study underwent myocardial perfusion single-photon emission computed tomography using adenosine infusion. Twenty subjects without clinical symptoms of angina matched for age, sex and cardiac risk factors served as a control group. N/OFQ levels were significantly (P < 0.01) higher in the patients (15.2 ± 2.1 pg/ml) than in the control group (8.5 ± 2.6 pg/ml). Blood pressure and heart rate did not significantly differ. All patients showed transient adenosine infusion myocardial ischemia that did not induce chest pain or significantly modify plasma N/OFQ levels or hemodynamic parameters. Our findings show that unstable angina is associated with a significant increase in circulating N/OFQ levels unrelated to intervening transient myocardial ischemia or hemodynamic changes. This increase is probably related to the chest pain repeatedly occurring in the course of coronary artery disease, but absent during transient adenosine-induced myocardial ischemia.     

Polyazetidine-based immobilization of redox proteins for electron-transfer-based biosensors

A highly stable functional composite film was prepared using polyazetidine prepolymer (PAP) with peroxidase from horseradish (HRP) and/or glucose oxidase (GOx). The good permeability of the PAP layer to classical electrochemical mediators, as evaluated by the determination of the diffusion coefficient of different redox molecules, is of great importance in view of the use of PAP as an immobilizing agent in second-generation biosensor development. Cyclic voltammetry of the HRP-PAP layer on a glassy carbon electrode (GCE) showed a pair of stable and quasi-reversible peaks for the HRP-Fe((III))/Fe((II)) redox couple at about -370 mV vs. Ag/AgCl electrode in pH 6.5 phosphate buffer. The electrochemical reaction of HRP entrapped in the PAP film exhibited a surface-controlled electrode process. This film and the successive modifications (HRP-PAP self-assembled monolayer (SAM) modified Au electrode) were used as a biological catalyst (hydrogen peroxide transducers) for glucose biosensors, after coupling to GOx. Both HRP/GOx-PAP and HRP/GOx-PAP SAM third generation biosensors were prepared and characterized. The use of PAP as immobilizing agent offers a biocompatible micro-environment for confining the enzyme and foreshadows the great potentiality of this immobilizing agent not only in theoretical studies on protein direct electron transfer but also from an applications point of view in the development of second- and third-generation biosensors.     

Vitamin D substrate–product relationship in idiopathic hypercalciuria

Absorptive hypercalciuria (AH) is associated with elevated levels of 1,25-dihydroxyvitamin D (1,25(OH)₂D). While no increase of 1,25(OH)₂D after oral administration of 25-hydroxyvitamin D (25OHD) at high doses has been claimed in normal subjects, a substrate–product relationship has been reported in normal children, young people after UV irradiation, older persons, postmenopausal women, primary hyperparathyroidism, renal failure, osteomalacia, and sarcoidosis. No data of this relationship in AH is available. To investigate 25OHD-1,25(OH)₂D substrate–product relationship in AH, 161 AH patients (mean age 60.9 ± 11.7 years) and 110 age- and sex-matched controls (mean age 61.5 ± 12.4 years) were studied. In 57 controls and 52 AH subjects 25OHD-1,25(OH)₂D relationship in basal conditions and after 2-week oral 25OHD (25 μg/day) administration were evaluated. In basal conditions 25OHD and 1,25(OH)₂D were correlated in both, controls and AH; 25OHD treatment was followed by an increase in serum 25OHD and 1,25(OH)₂D in both groups. However, delta responses of 25OHD and 1,25(OH)₂D to 25OHD were higher in AH suggesting an enhanced activity of 1α-hydroxylase. In conclusion, the higher response of 1,25(OH)₂D after oral 25OHD in AH patients suggests a differential capacity between both groups in handling the increases in 1,25(OH)₂D.