Treatment with PB125® Increases Femoral Long Bone Strength in 15-Month-Old Female Hartley Guinea Pigs.

Nuclear factor-erythroid 2-related factor-2 (Nrf2) is a transcription factor that serves as a master regulator of anti-inflammatory agents, phase I xenobiotic, and phase II antioxidant enzymes, all of which provide a cytoprotective role during disease progression. We hypothesized that oral administration of a purported phytochemical Nrf2-activator, PB125®, would increase long bone strength in aging Hartley guinea pigs, a model prone to musculoskeletal decline. Male (N = 56) and female (N = 56) guinea pigs were randomly assigned to receive daily oral treatment with either PB125® or vehicle control. Animals were treated for a consecutive 3-months (starting at 2-months of age) or 10-months (starting at 5-months of age) and sacrificed at 5-months or 15-months of age, respectively. Outcome measures included: (1) ANY-maze™ enclosure monitoring, (2) quantitative microcomputed tomography, and (3) biomechanical testing. Treatment with PB125® for 10 months resulted in increased long bone strength as determined by ultimate bending stress in female Hartley guinea pigs. In control groups, increasing age resulted in significant effects on geometric and structural properties of long bones, as well as a trending increase in ultimate bending stress. Furthermore, both age and sex had a significant effect on the geometric properties of both cortical and trabecular bone. Collectively, this work suggests that this nutraceutical may serve as a promising target and preventive measure in managing the decline in bone mass and quality documented in aging patients. Auxiliary to this main goal, this work also capitalized upon 5 and 15-month-old male and female animals in the control group to characterize age- and sex-specific differences on long bone geometric, structural, and material properties in this animal model.

Challenges of measles and rubella laboratory diagnostic in the era of elimination.

The Member States of the WHO European Region adopted the goal of measles and rubella elimination more than 10 years ago, but so far only 21 of 53 countries have reached this target. Laboratory investigation of suspected cases is essential to support disease elimination efforts. Therefore, WHO maintains a network of accredited laboratories providing high-quality testing. Laboratory investigation heavily relies on specific IgM serology and increasingly on virus detection by reverse transcription (RT)-PCR, but other methods such as IgG avidity testing and genetic characterization of virus strains have gained in importance. In elimination settings, often few samples from suspected cases are available for testing, but testing proficiency must be maintained. The predictive value of an IgM-positive result decreases and other rash-fever disease aetiologies become more important. In addition, cases with a rash after measles/rubella vaccination or with mild disease after waning of vaccine-induced antibodies are seen more often. Thus, it is necessary to perform comprehensive and potentially time-consuming and costly investigations of every suspected case using quality-controlled laboratory methods. At the same time rapid feedback to public health officers is required for timely interventions. The introduction of new laboratory methods for comprehensive case investigations requires training of staff under the supervision of WHO-accredited reference laboratories and the definition of appropriate test algorithms. Clinical, laboratory, and epidemiological data are essential for final case classification and investigation of chains of transmission in the endgame of measles and rubella elimination.

[Relapse rate within 6 months after successful ECT: a naturalistic prospective peer- and self-assessment analysis]. / Rückfallraten innerhalb von 6 Monaten nach erfolgreicher EKT--Eine naturalistische prospektive Fremd- und Selbstbeurteilungsanalyse.

BACKGROUND: Up to 100% relapse rate after successful electroconvulsive therapy (ECT) poses a challenge for patients and psychiatrists. The aim of the study was to evaluate the outcome of patients affected by major depression after the successful course of acute ECT. METHODS: 84 patients recruited in a randomized double blind multicenter study designed to investigate the optimal stimulation placement in acute ECT had a follow up under naturalistic conditions between the 5th and 7th month. Outcome, maintenance therapy and patients; attitude were evaluated with semi structured questionnaires by patients and the study raters. RESULTS: 82.14% (68/84) questionnaires of the patients and 83.3% (70/84) of the rater were returned. 98% of the patients had at least one antidepressant; only in 23% (20/68) lithium was prescribed. 35% (7/20) of the patients with lithium and 57% (16/28) without lithium had a relapse within the first 6 months (OR 0.6) in a median of 2.5 months. Only one institution offered maintenance ECT in 8.3% (7/84) patients. For 52.2% of the patients ECT was a helpful treatment an 49.3% would recommend the therapy to their relatives. The vast majority (59.4%) wishes a better information about the ECT and 21.4% feel frightening about the therapy. CONCLUSIONS: The results show a high relapse rate and highlight the meaning of maintenance medication especially for a lithium combination therapy, as stated before. In regard to the subjective sensation the patients claim a better education about the ECT and anyway one of four patients feel frightening about the therapy.

Isolation of human mesenchymal stromal cells is more efficient by red blood cell lysis.

BACKGROUND: Human mesenchymal stromal cells (MSC) have raised high hopes for tissue engineering and clinical therapy. Their isolation usually involves density fractionation of mononuclear cells (MNC) but this is difficult to standardize, especially under good manufacturing practice (GMP) conditions. MSC represent a heterogeneous mixture of cell types and the composition of subpopulations is affected by the initial steps of cell preparation. METHODS: This study describes a straightforward method for isolation of human MSC based on red blood cell (RBC) lysis with ammonium chloride. Colony formation was compared directly with Ficoll density fractionation and culture of an untreated whole bone marrow (BM) aspirate. RESULTS: After 7 days the number of fibroblastic colony-forming units (CFU-F) per milliliter of BM aspirate was slightly higher upon RBC lysis and the colonies were significantly larger compared with density fractionation, possibly because of maintenance of platelets. In contrast, colony formation was much lower in untreated BM. The heterogeneous composition of subpopulations was reflected by differences between the initial colonies with regard to growth pattern (tight or disperse) and cell morphology (round or elongated). This heterogeneous composition was not affected by the three different isolation methods. Furthermore, enrichment of CD271(+) cells resulted in the same morphologic heterogeneity. All cell preparations demonstrated the same immunophenotype using a panel of surface markers and displayed adipogenic and osteogenic differentiation potential. DISCUSSION: This study demonstrates that human MSC can be efficiently isolated by RBC lysis. This technique is faster and can be standardized more easily for clinical application of MSC.

Molecular signature of CD34(+) hematopoietic stem and progenitor cells of patients with CML in chronic phase.

In this study, we provide a molecular signature of highly enriched CD34+ cells from bone marrow of untreated patients with chronic myelogenous leukemia (CML) in chronic phase in comparison with normal CD34+ cells using microarrays covering 8746 genes. Expression data reflected several BCR-ABL-induced effects in primary CML progenitors, such as transcriptional activation of the classical mitogen-activated protein kinase pathway and the phosphoinositide-3 kinase/AKT pathway as well as downregulation of the proapoptotic gene IRF8. Moreover, novel transcriptional changes in comparison with normal CD34+ cells were identified. These include upregulation of genes involved in the transforming growth factorbeta pathway, fetal hemoglobin genes, leptin receptor, sorcin, tissue inhibitor of metalloproteinase 1, the neuroepithelial cell transforming gene 1 and downregulation of selenoprotein P. Additionally, genes associated with early hematopoietic stem cells (HSC) and leukemogenesis such as HoxA9 and MEIS1 were transcriptionally activated. Differential expression of differentiation-associated genes suggested an altered composition of the CD34+ cell population in CML. This was confirmed by subset analyses of chronic phase CML CD34+ cells showing an increase of the proportion of megakaryocyte-erythroid progenitors, whereas the proportion of HSC and granulocyte-macrophage progenitors was decreased in CML. In conclusion, our results give novel insights into the biology of CML and could provide the basis for identification of new therapeutic targets.

Calcium-ions are involved in erythrocyte invasion by equine Babesia parasites.

Ethylene glycol bis (beta-aminoethylether)-N,N,N,N-tetraacetic acid (EGTA) is a chelating agent capable of binding to positively-charged metal ions, including a calcium-ion (Ca2+). Here, we demonstrated the inhibitory effect of the chemical on the in vitro asexual growth of the equine protozoan parasites, Babesia caballi and Babesia equi. The growth of both B. caballi and B. equi was significantly inhibited in the presence of EGTA (IC50=1.27 and 2.25 mM, respectively). Under microscopical observation, increased percentages of extracellular merozoites in the total parasites were detected in both of the cultures treated with high concentrations of EGTA. In contrast, further addition of Ca2+ to the EGTA-treated cultures prevented the parasites from clearing and the percentages of extracellular merozoites from increasing. As for B. caballi, an invasion test using high-voltage pulsing proved that EGTA has an inhibitory effect to their erythrocyte invasion. These results suggest that Ca2+ is involved in erythrocyte invasion by equine Babesia parasites.

Effects of protein kinase inhibitors on the in vitro growth of Babesia bovis.

Staurosporine, Ro-31-7549, and KN-93, which are inhibitors of serine/threonine protein kinase, protein kinase C, and calcium-modulin kinase, respectively, were tested for their effects on the in vitro growth of Babesia bovis. Staurosporine was the most effective inhibitor, completely clearing the parasitaemia as early as the first day of exposure at a concentration of 100 microM. Moreover, staurosporine caused a significant increase in the percentage of extracellular merozoites, most likely due to the inhibition of erythrocyte invasion by the parasite. Although 5 mM Ro-31-7549 and KN-93 had a suppressive action, this was not enough to destroy the parasite. Interestingly, concentrations of 0.5 to 5 mM KN-93 influenced the parasitic development within the infected erythrocytes. The present study suggests that B. bovis requires, to a certain extent, the phosphorylations mediated by parasite- or host erythrocyte-protein kinases, in particular, for the processes of successful invasion of erythrocytes and intraerythrocytic development.

Host serum modifies the drug susceptibility of Babesia bovis in vitro.

Babesia parasites generally require a defined percentage of serum in the culture medium for their in vitro growth. In this study, we attempted to culture Babesia bovis in a serum-free condition. The growth pattern and morphology of B. bovis in serum-free (plain) GIT medium were unaltered as compared to those of the standard growth condition containing 40% bovine serum in M199. When exposed to the test drugs, the parasite in plain GIT medium showed clearly lower IC50 values than those in 40% serum-containing G IT medium, indicating that several serum components may interfere with the drug bio-availability. Therefore, the serum-free culture system is useful for standardizing drug test protocols and understanding the roles of serum factors in the drug test.

Successful transplantation of peripheral blood stem cells mobilized by chemotherapy and a single dose of pegylated G-CSF in patients with multiple myeloma.

Following induction therapy and 4 g/m(2) cyclophosphamide, a single dose of 12 mg polyethyleneglycol-conjugated G-CSF (pegfilgrastim; n=12) or daily doses of unconjugated G-CSF (8.5 mug/kg/day) (n=12) were administered to myeloma patients. Pegfilgrastim was associated with an earlier leukocyte recovery (12 vs 14 days) and peripheral blood CD34+ cell peak (12 vs 15 days). The peripheral blood CD34+ cell peak was lower in the pegfilgrastim group (78 vs 111/mul). Following high-dose melphalan (200 mg/m(2)) and autografting, leukocyte and platelet reconstitution was similar in both groups and stable blood counts were observed 100 days post transplant. In summary, a single dose of pegfilgrastim after chemotherapy is capable of mobilizing a sufficient number of CD34+ cells for successful autografting with early engraftment and sustained hematological reconstitution in patients with myeloma. These data provide the basis for randomized studies evaluating the optimal dose and time of pegfilgrastim as well as long-term outcome in larger cohorts of patients.

Clotrimazole, ketoconazole, and clodinafop-propargyl inhibit the in vitro growth of Babesia bigemina and Babesia bovis (Phylum Apicomplexa).

We evaluated the growth inhibitory efficacy of the imidazole derivatives, clotrimazole (CLT) and ketoconazole (KC), and the herbicide clodinafop-propargyl (CP), in in vitro cultures of Babesia bovis and B. bigemina. Clotrimazole was effective in a dose range of 15 to 60 microM (IC50: 11 and 23.5 microM), followed by KC (50 to 100 microM; IC50: 50 and 32 microM) and CP (500 microM; IC50: 265 and 390 microM). In transmission electron microscopy, extensive damage was observed in the cytoplasm of drug-treated parasites. Combinations of CLT/KC, CLT/CP and CLT/KC/CP acted synergistically in both parasites. In contrast, the combination of KC/CP was exclusively effective in B. bovis, but not in B. bigemina.