GABAergic Interneurons at Supraspinal and Spinal Levels Differentially Modulate the Antinociceptive Effect of Nitrous Oxide in Fischer Rats

Background: The study hypothesizes that nitrous oxide (N2O) releases opioid peptide in the brain stem, which results in inhibition of [gamma]-aminobutyric acid-mediated (GABAergic) neurons that tonically inhibit the descending noradrenergic inhibitory neurons (DNIN), resulting in activation of DNIN. In the spinal cord, activation of DNIN leads to the release of norepinephrine, which inhibits nociceptive processing through direct activation of [alpha]2 adrenoceptor and indirect activation of GABAergic neurons through [alpha]1 adrenoceptor. Arising from this hypothesis, it follows that GABAergic neurons will modulate the antinociceptive effect of N2O in diametrically opposite directions at supraspinal and spinal levels. The authors have tested this tenet and further examined the effect of midazolam, a GABA-mimetic agent, on N2O-induced antinociceptive effect.

Methods: Adult male Fischer rats were administered muscimol (GABAA receptor agonist) intracerebroventricularly (icv), gabazine (GABAA receptor antagonist) intrathecally (intrathecal), or midazolam intraperitoneally (intraperitoneal). Fifteen minutes later, they were exposed to air or 75% N2O and were subjected to the plantar test after 30 min of gas exposure. In some animals administered with midazolam, gas exposure was continued for 90 min, and the brain and spinal cord were examined immunohistochemically.

Results: The N2O-induced antinociceptive effect, which was attenuated by icv muscimol, intrathecal gabazine, and intraperitoneal midazolam. Midazolam inhibited N2O-induced c-Fos expression (a marker of neuronal activation) in the pontine A7 and spinal cord.     

Interleukin-7-engineered mesenchymal cells: in vitro effects on naive T-cell population.

T-cell homeostasis is regulated by several molecules; among these, interleukin (IL)-7 plays an essential role in the survival and homeostatic proliferation of peripheral naive T cells. In a previous study, we investigated whether human mesenchymal stromal cells (MSCs) could be engineered with the IL-7 gene to produce functional level of this cytokine. In the present study, we analyzed the impact of different quantities of IL-7 produced by MSCs on the survival and proliferation of a negative immunoselected naive (CD3(+)/CD45RA(+)) T-cell population. Co-cultivation of peripheral naive T cells with MSCs producing low (16 pg/mL) or high (1000 pg/mL) IL-7 levels or in the presence of exogenous IL-7 (0.01 ng/mL and 100 ng/mL) maintained the CD3(+)/CD45RA(+) naive T-cell phenotype. Chemokine receptor CCR7(+) expression was also maintained among this T-cell population. Naive T-cell molecular characteristics were maintained as assessed by the Vbeta spectratyping complexity score, which showed the maintenance of a broad T-cell repertoire. No Th1 or Th2 differentiation was observed, as assessed by interferon-gamma or IL-4 accumulation. In contrast, only MSCs producing high amounts of IL-7 caused increased activation (CD25 31.2% +/- 12% vs 10% +/- 3.5%; P < .05), proliferation (CD71 17.8+/-7% vs 9.3%+/-3, P < .05), apoptosis (assessed by annexin V: 18.6% +/- 5% vs 14.9% +/- 2.6%; P > .05), and the phase S cell cycle (15% vs 6.9%, P > .05). Exogenous IL-7 exhibited no significant effect. In conclusion, we demonstrated that IL-7 produced by MSCs has a dose-independent effect on naive T-cell survival while exerting a dose-dependent effect on activation/proliferation. Due to the continuous production of IL-7 by engineered cells, our system is more efficacious than exogenous IL-7.     

Sodium hypochlorite-, chlorine dioxide- and peracetic acid-induced genotoxicity detected by the Comet assay and Saccharomyces cerevisiae D7 tests

Mutagenicity of drinking water is due not only to industrial,agricultural and urban pollution but also to chlorine disinfectionby-products. Furthermore, residual disinfection is used to providea partial safeguard against low level contamination and bacterialre-growth within the distribution system. The aims of this studywere to further evaluate the genotoxic potential of the worldwide used disinfectants sodium hypochlorite and chlorine dioxidein human leukocytes by the Comet assay and in Saccharomycescerevisiae strain D7 (mitotic gene conversion, point mutationand mitochondrial DNA mutability, with and without endogenousmetabolic activation) and to compare their effects with thoseof peracetic acid, proposed as an alternative disinfectant.All three disinfectants are weakly genotoxic in human leukocytes(lowest effective dose 0.2 p.p.m. for chlorine dioxide, 0.5p.p.m. for sodium hypochlorite and peracetic acid). The resultsin S.cerevisiae show a genotoxic response on the end-pointsconsidered with an effect only at doses higher (5- to 10-fold)than the concentration normally used for water disinfection;sodium hypochlorite and peracetic acid are able to induce genotoxiceffects without endogenous metabolic activation (in stationaryphase cells) whereas chlorine dioxide is effective in growingcells. The Comet assay was more sensitive than the yeast tests,with effective doses in the range normally used for water disinfectionprocesses. The biological effectiveness of the three disinfectantson S.cerevisiae proved to be strictly dependent on cell-specificphysiological/biochemical conditions. All the compounds appearto act on the DNA and peracetic acid shows effectiveness similarto sodium hypochlorite and chlorine dioxide. ¹Author to whom correspondence should be addressed. Tel: +39 0521 905608; Fax: +39 0521 905604; Email: mutgen{at}unipr.it Received on September 22, 2003; revised and accepted on November 27, 2003     

ABO antibody titres: a multisite comparative study of equivalency and reproducibility for automated solid-phase and haemagglutination titration,and manual dilution with gel column agglutination technology

BackgroundABO antibody titres are important in many clinical decisions; however, much variability is observed in titre results. For reliable and reproducible titre results, automated ABO titration methods have been developed. In this 10-site study, we evaluated the equivalency of the automated ABO titration assays on the Galileo NEO, a fully automated blood bank analyzer (Immucor, Inc.) to manual titration with gel Column Agglutination Technology (CAT), as well as the reproducibility of both methods.Materials and methodsTen different locations participated in this study. The equivalency study included 70 random samples at each site. The reproducibility study tested the same blinded 30-sample panel at each study site. Anti-A and anti-B IgM and IgG antibody titres were tested with both the automated and manual methods; additionally, dithiothreitol (DTT) treatment was used to inactivate IgM antibodies in the manual CAT method.ResultsThe equivalency between CAT manual method and Galileo NEO automated titres at each site ranged from 38 to 88%; equivalency for each isotype was 66.2% for IgM, 60.6% for IgG, and 88.5% for DTT-treated IgG. The reproducibility study evaluated the titre variation of each sample obtained from the 10 sites. The average titre ranges (in doubling dilutions) for the automated and manual methods, respectively, were 2.15±1.0 and 4.03±1.8 for IgM, and 1.53±0.7 and 4.10±1.9 for IgG; for the manual DTT-treated IgG, the average titre range was 3.45±1.8 doubling dilutions.DiscussionThe results demonstrated that the Galileo NEO automated and manual CAT ABO titres are not equivalent. However, the study also demonstrated that titre reproducibility is enhanced with the Galileo NEO automated ABO titration assays relative to the manual CAT ABO titration method. Therefore, to improve management of patients receiving care across multiple institutions, our study supports the use of automated ABO titration.     

Cytokine Pattern of Peripheral Blood Mononuclear Cells Isolated from Children Affected by Generalized Epilepsy Treated with Different Protein Fractions of Meat Sources

The objective of the present study was the evaluation of cytokine patterns in terms of TNF-α, IL-10, IL-6, and IL-1β secretion in peripheral blood mononuclear cell (PBMC) supernatants isolated from blood of children affected by generalized epilepsy and treated in vitro with myofibrillar, sarcoplasmic, and total protein fractions of meat and fish sources. Children with generalized epilepsy (EC group, n = 16) and children without any clinical signs of disease, representing a control group (CC group n = 16), were recruited at the Complex Structure of Neuropsychiatry Childhood-Adolescence of Policlinico Riuniti (Foggia, Italy). Myofibrillar (MYO), sarcoplasmic (SA), and total (TOT) protein fractions were obtained from longissimus thoracis muscle of beef (BF) and lamb (LA); from pectoralis muscle of chicken (CH); and from dorsal white muscle of sole (Solea solea, SO), European hake (Merluccius merluccius, EH), and sea bass fish (Dicentrarchus labrax, SB), respectively. PBMCs were isolated from peripheral blood of EC and CC groups, and an in vitro stimulation in the presence of 100 μg/mL for each protein fraction from different meat sources was performed. Data were classified according to three different levels of cytokines produced from the EC group relative to the CC group. TNF-α, IL-10, and IL-6 levels were not affected by different meat fractions and meat sources; on the contrary, IL-1β levels were found to be significantly affected by the tested proteins fractions, as well as different meat sources, in high-level cytokine group. On average, the protein fractions obtained from LB, BF, and CH meat sources showed a higher level of IL-1β than the protein fractions obtained from EH and SB fish samples. When all cytokine classes were analyzed, on average, a significant effect was observed for IL-10, IL-1β, and TNF-α. Data obtained in the present study evidence that the nutritional strategy based on protein from fish and meat sources may modulate the immunological cytokine pattern of infants with generalized epilepsy.     

Validation of the Italian Version of the SARC-F Questionnaire to Assess Sarcopenia in Older Adults

Background: SARC-F is a simple sarcopenia screening tool. This study aimed to examine the validity of the Italian version of SARC-F. Methods: A total of 97 elderly individuals (37/60 males/females, 65 years and older) who met the study’s selection criteria were included. SARC-F was translated into the Italian language in a culturally responsive manner. The total score was calculated by adding the scores on the five items. The participants were divided into two groups according to the total score (SARC-F < 4 vs. SARC-F ≥ 4), and their associations with various factors (handgrip test, chair stand test, and Skeletal Muscle Index assessed by DXA) have been examined by gender. In addition, the tool’s validity was analyzed by comparing it with different international working group diagnostic criteria for sarcopenia. Results: The total prevalence of sarcopenia according to the SARC-F was 14.2% and, specifically, 12.8% among men and 14.3% in women. The sensitivity of the SARC-F was (male (M): 11–50% and female (F): 22–36%) medium-low compared with the European, international, and Asian criteria of sarcopenia; however, SARC-F showed a high specificity (M: 77.3–100% and F: 79.5–100%) and a moderate Cronbach’s alpha coefficient of (0.669 (CI95%: 0.358–0.830). The participants in the SARC-F ≥ 4 group had poorer handgrip for EWGSOP2 (p < 0.001) and chair stand (p < 0.001) than the participants in the SARC-F < 4 group. Conclusions: The Italian language version of SARC-F showed high specificity, moderate reliability, and good associations with other predictive tests. The Italian version of SARC-F appears to be a useful screening tool for the diagnosis of sarcopenia in Italian elderly populations.     

Production of Dense Cu-10Sn Part by Laser Powder Bed Fusion with Low Surface Roughness and High Dimensional Accuracy

Tin-bronze alloys with a tin content of at least 10 wt% have excellent mechanical properties, wear resistance, and corrosion resistance. Among these alloys, Cu-10Sn was investigated in this study for production with the laser powder bed fusion process with a 500W Yb:YAG laser. In particular, a design of experiment (DoE) was developed in order to identify the optimal process parameters to obtain full density, low surface roughness, and high dimensional accuracy. Samples were characterized with Archimedes’ method and optical microscopy to determine their final density. It was shown that the first method is fast but not as reliable as the second one. A first mechanical characterization was performed through microhardness tests. Finally, a set of process parameters was identified to produce fully dense samples with low surface roughness and high accuracy. The results showed that the volumetric energy density could represent an approach that is too simplified, therefore limiting the direct correlation with the physical aspects of the process.     

An unusual case of postaxial polydactyly of the feet

Polydactyly is a fairly common congenital anomaly characterized by additional fingers and/or toes, generally divided into three groups: medial ray (preaxial) and central and lateral ray (postaxial). We present a case of postaxial (fibular) polydactyly of the feet, assaying its clinical and radiological features and the method of surgical correction. Clinical outcomes after initial surgery were excellent in both feet. This surgical case shows that a careful individual preoperative planning is necessary to achieve good functional and cosmetic outcome. Level of Evidence: Level V, therapeutic study.