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Summary Current human pluripotent stem cells lack the transcription factor circuitry that governs the ground state of mouse embryonic stem cells (ESC). Here, we report that short-term expression of two components, NANOG and KLF2, is sufficient to ignite other elements of the network and reset the human pluripotent state. Inhibition of ERK and protein kinase C sustains a transgene-independent rewired state. Reset cells self-renew continuously without ERK signaling, are phenotypically stable, and are karyotypically intact. They differentiate in vitro and form teratomas in vivo. Metabolism is reprogrammed with activation of mitochondrial respiration as in ESC. DNA methylation is dramatically reduced and transcriptome state is globally realigned across multiple cell lines. Depletion of ground-state transcription factors, TFCP2L1 or KLF4 , has marginal impact on conventional human pluripotent stem cells but collapses the reset state. These findings demonstrate feasibility of installing and propagating functional control circuitry for ground-state pluripotency in human cells. [ABSTRACT FROM AUTHOR]

Canine hepatozoonosis is a worldwide protozoal disease caused by Hepatozoon canis and Hepatozoon americanum and is transmitted by ixodid ticks, Rhipicephalus and Amblyomma spp., respectively. H. canis infection is widespread in Africa, Europe, South America, and Asia, including Japan. The objective of this study was to study the distribution pattern and diversity of H. canis in naturally infected dogs in nine Japanese islands and peninsulas. Therefore, 196 hunting dogs were randomly sampled during the period from March to September 2011 and the ages and sexes were identified. Direct microscopy using Giemsa-stained blood smears revealed H. canis gametocytes in the peripheral blood of 45 (23.6 %) dogs. Polymerase chain reaction (PCR) was performed on EDTA-anticoagulated blood, initially with the common primer set (B18S-F and B18S-R) amplifying the 1,665-bp portion of the 18S rRNA gene, and then with the specific primer set (HepF and HepR) amplifying about 660 bp fragments of the same gene. Based on PCR, 84 (42.9 %) dogs were positive using the common primer and 81 (41.3 %) were positive using the specific primer. The current investigation indicated that all screened areas, except for Sado Island and Atsumi Peninsula, were infected. Yaku Island had the highest infection rate (84.6 % in males and 100.0 % in females), while Ishigaki Island showed the lowest infection rates (8.3 % in males and 17.7 % in females). Both sexes were infected with no significant difference. However, diversity of infection among the surveyed islands and peninsulas was significantly different ( P < 0.05). Although H. canis has previously been reported in dogs in Japan, the higher infection rate described in the current study and the diversity of infection in a wide range of islands strongly encourage prospective studies dealing with the prevention and treatment of the infection in dogs, as well as control of ticks. [ABSTRACT FROM AUTHOR]

OBJECTIVE--Growth of white adipose tissue takes place in normal development and in obesity. A pool of adipose progenitom is responsible for the formation of new adipocytes and for the potential of this tissue to expand in response to chronic energy overload. However, factors controlling self-renewal of human adipose progenitors are largely unknown. We investigated the expression profile and the role of activin A in this process. RESEARCH DESIGN AND METHODS---Expression of INHBA/ activin A was investigated in three types of human adipose progenitors. We then analyzed at the molecular level the function of activin A during human adipogenesis. We finally investigated the status of activin A in adipose tissues of lean and obese subjects and analyzed macrophage-induced regulation of its expression. RESULTS--INHBA/activin A is expressed by adipose progenitors from various fat depots, and its expression dramatically decreases as progenitors differentiate into adipocytes. Activin A regulates the number of undifferentiated progenitors. Sustained activation or inhibition of the activin A pathway impairs or promotes, respectively, adipocyte differentiation via the C/EBPβLAP and Smad2 pathway in an autocrine/paracrine manner. Activin A is expressed at higher levels in adipose tissue of obese patients compared with the expression levels in lean subjects. Indeed, activin A levels in adipose progenitors are dramatically increased by factors secreted by macrophages derived from obese adipose tissue. CONCLUSIONS--Altogether, our data show that activin A plays a significant role in human adipogenesis. We propose a model in which macrophages that are located in adipose tissue regulate adipose progenitor self-renewal through activin A. Diabetes 59: 2513-2521, 2010 [ABSTRACT FROM AUTHOR]

Abstract: The mucosal vaccination is a non-invasive alternative approach for not only mucosal pathogens but also parenteral pathogens, since it induces both mucosal and systemic immunoreactions. The purpose of this study was to evaluate the application of intranasal (i.n.) immunization with a recombinant leishmanial protein against Leishmania infection. BALB/c mice were i.n. administered 1–3 times with Leish-111f plus cholera toxin (CT) adjuvant (Leish-111f/CT). Splenocytes from i.n. immunized mice produced high level of IFN-γ but not IL-4 in response to Leish-111f. When infected with 1×106 of Leishmania major promastigotes 2 weeks after the final administration, lesion development was completely controlled in all mice i.n. administered with Leish-111f/CT. Mice i.n. administered with Leish-111f alone showed neither cytokine productions nor lesion control even after 6 administrations, suggesting the importance of CT adjuvant. This report demonstrated for the first time that i.n. administration of a recombinant leishmanial protein induces Th1 type immunity and protects mice from Leishmania infection. [Copyright &y& Elsevier]

Ectopic expression of defined sets of genetic factors can reprogram somatic cells to create induced pluripotent stem (iPS) cells. The capacity to direct human iPS cells to specific differentiated lineages and to their progenitor populations can be used for disease modeling, drug discovery, and eventually autologous cell replacement therapies. During mouse cardiogenesis, the major lineages of the mature heart, cardiomyocytes, smooth muscle cells, and endothelial cells arise from a common, multipotent cardiovascular progenitor expressing the transcription factors Isl1 and Nkx2.5. Here we show, using genetic fate-mapping, that Isl1+ multipotent cardiovascular progenitors can be generated from mouse iPS cells and spontaneously differentiate in all 3 cardiovascular lineages in vivo without teratoma. Moreover, we report the identification of human iPS-derived ISL1+ progenitors with similar developmental potential. These results support the possibility to use patient-specific iPS-generated cardiovascular progenitors as amodel to elucidate the pathogenesis of congenital and acquired forms of heart diseases. [ABSTRACT FROM AUTHOR]

Abstract: Detection of the initial site of Toxoplasma gondii reactivation in brain tissue is difficult because the number of latent cysts is small and reactivation is a transient event. To detect the early stage of reactivation in mouse brain tissue, we constructed a cyst-forming strain of T. gondii in the tachyzoite stage, specifically expressing red fluorescence. The PLK strain of T. gondii was stably transfected with a red fluorescent protein gene, DsRed Express, under the control of a tachyzoite-specific SAG-1 promoter and the resulting parasite was designated as PLK/RED. Tachyzoites of PLK/RED growing in Vero cells showed red fluorescence. When C57BL/6J mice were i.p. infected with tachyzoites of PLK/RED, red fluorescent tachyzoites were detected in their brains at the fourth day p.i. However, red fluorescent tachyzoites were not detected in BALB/c mice latently infected with PLK/RED, although non-fluorescent cysts were detected in their brains. After treatment of latently infected mice with dexamethasone for 1 month, the mice showed neurological symptoms. In mice with symptoms, red fluorescent tachyzoites were again detected in their brains and in other organs. To detect the initial site of reactivation, BALB/c mice latently infected with the strain were treated with dexamethasone for 3 weeks, and brains were excised before any symptoms appeared. Excised brains were examined for red fluorescence-positive sites. By a histological study of red fluorescent-positive sites, we detected a cyst containing red fluorescent zoites, which still had a PAS stain-positive cyst wall. A few red fluorescent zoites breaking away from the cyst were also observed. The stage-specific expression of fluorescent protein facilitates detection of a rare transient event and makes it possible to detect the initial site of reactivation. [Copyright &y& Elsevier]

Neurotransmitter receptors play a key role in most research on antipsychotic drugs, but little is known about the effects of these drugs on the plasma membrane in the central nervous system. Therefore, we investigated whether chlorpromazine (CPZ), a typical, phenothiazine antipsychotic drug, affects the plasma membrane integrity in the rat brain, and if so, whether these membrane alterations can be prevented by dietary supplementation with vitamin E, which has been shown to be an antioxidant and also a membrane-stabilizer. Leakage of [18F]2-fluoro-2-deoxy-D-glucose ([18F]FDG)-6-phosphate from rat striatal slices and decrease in 1,6-diphenyl-1,3,5-hexatriene fluorescence anisotropy were used as indexes for plasma membrane permeabilization and fluidization, respectively. CPZ induced leakage of [18F] FDG-6-phosphate from striatal slices, and the leakage was delayed in the vitamin E-supplemented group compared to that in the normal diet group. The decrease in plasma membrane anisotropy induced by CPZ was significantly attenuated by vitamin E supplementation. Chronic treatment with α-phenyl-N-tert-butyl nitrone, a free radical scavenger, had no effect on CPZ-induced plasma membrane permeabilization, and the treatment with CPZ did not induce lipid peroxidation. CPZ can reduce plasma membrane integrity in the brain, and this reduction can be prevented by vitamin E via its membrane-stabilizing properties, not via its antioxidant activity. [ABSTRACT FROM AUTHOR]

We compared the potency of the interaction of three antipsychotic drugs, i.e. chlorpromazine (CPZ), haloperidol (Hal) and sulpiride (Sul), with the plasma membrane in the rat brain. CPZ loading (⩾ 100 μM) dose-dependently increased both membrane permeability (assessed as [18F]2-fluoro-2-deoxy-D-glucose-6-phosphate release from brain slices) and membrane fluidity (assessed as the reduction in the plasma membrane anisotropy of 1,6-diphenyl-1,3,5-hexatriene). On the other hand, a higher concentration of Hal (1 mM) was required to observe these effects. However, Sul failed to change membrane permeability and fluidity even at a high concentration (1 mM). These results indicated the following ranking of the potency to interact with the membrane: CPZ>Hal>Sul. The difference among antipsychotic drugs in the potency to interact with the plasma membrane as revealed in the present study may be partly responsible for the difference among the drugs in the probability of inducing extrapyramidal side-effects such as parkinsonism and tardive dyskinesia. [ABSTRACT FROM AUTHOR]

Summary: Mesenchymal stem cells (MSCs) are defined as cells that undergo sustained in vitro growth and are able to give rise to multiple mesenchymal lineages. Although MSCs are already used in regenerative medicine little is known about their in vivo behavior and developmental derivation. Here, we show that the earliest wave of MSC in the embryonic trunk is generated from Sox1+ neuroepithelium but not from mesoderm. Using lineage marking by direct gfp knock-in and Cre-recombinase mediated lineage tracing, we provide evidence that Sox1+ neuroepithelium gives rise to MSCs in part through a neural crest intermediate stage. This pathway can be distinguished from the pathway through which Sox1+ cells give rise to oligodendrocytes by expression of PDGFRβ and A2B5. MSC recruitment from this pathway, however, is transient and is replaced by MSCs from unknown sources. We conclude that MSC can be defined as a definite in vivo entity recruited from multiple developmental origins. [Copyright &y& Elsevier]

The article examines whether differentiation potential of parthenogenetic embryonic stem cells (pES) is improved by nuclear transfer (NT). Several second- and third-generation NT-pES cell lines were established from original pES cell by NT. These were examined for their potential to differential in vivo and in vitro.

Abstract: To enhance the efficacy of an inactivated vaccine against pseudorabies virus (PRV), we evaluated the adjuvant properties of Fc domain of IgG. A cell line expressing mouse IgG Fc chimera on its surface was established. We found that when PRV was propagated in the cells expressing the Fc chimera, PRV virion incorporated the Fc. Immunization of BALB/c mice with inactivated PRV harboring Fc, which had been propagated in the cells expressing Fc on its surface, induced higher antibody production against PRV and protected mice more effectively from lethal challenge of virulent strain, comparing to the immunization with normal inactivated virus. Virus harboring Fc has a great potential as a new inactivated vaccine. [Copyright &y& Elsevier]

Describes the expression of P23 of the coccidian parasite Cryptosporidium parvum in Toxoplasma gondii. Expression of sporozoites in Toxoplasma gondii; Evaluation of effects in a mouse model; Possible use of the vector as a tool for the production of a recombinant vaccine against cryptosporidiosis in animals.

An improved method for constructing canine herpesvirus (CHV) recombinants expressing foreign genes by using the lacZ-TK gene cassette as a double selectional marker was developed. A recombinant CHV carrying the lacZ-TK gene at a targeted gene locus was constructed and used as a parental virus for generating new recombinants. The parental virus formed blue plaques and was sensitive to TK-specific drugs, while newly generated recombinants, in which the lacZ-TK gene was replaced with the desired foreign gene, become both resistant to the TK-specific drugs and formed white plaques. Recombinants were isolated by using the combination of drug selection and color selection. This improved method allows construction of recombinant CHV with great ease, because the drug selection can enrich the frequency of recombinant CHV from 0.01–0.1% to 10–80%. This method was employed to construct a recombinant CHV that expressed rabies virus (RV) glycoprotein (G protein). [ABSTRACT FROM AUTHOR]

• Seventeen novel marmoset embryonic stem cell lines were established. • Stem cells were established from naturally and in vitro fertilized embryos. • First feeder-free marmoset embryonic stem cell lines from in vitro fertilized embryos. • The feeder-free method was efficient method for establishing embryonic stem cell lines. Pluripotent stem cells (PSCs), embryonic stem cells (ESCs), and induced PSCs (iPSCs) are excellent tools for studying embryonic development in organisms and classified into naïve and primed states. ESC-derived germline chimera individuals can be produced by injecting naïve ESCs/iPSCs into preimplantation embryos, and conversion of primed human ESCs/iPSCs into a naïve state provides insights into epiblast cell features. Non-human ESCs/iPSCs are alternatives to human naïve ESCs/iPSCs, which elicit ethical issues. In this study, we used the common marmoset (Callithrix jacchus) as an animal model. Since 1996, 16 marmoset ESC lines have been established. Because most of these ESC lines are female and were derived >10 years ago, new ESCs, particularly male marmoset ESC lines, are needed. Here, we successfully established 17 novel marmoset ESC lines, including six male ESC lines from in vitro -fertilized (IVF) embryos and 12 ESC lines under feeder-free conditions. This report is the first to establish ESC lines using feeder-free conditions and IVF preimplantation blastocysts in marmosets, and these novel ESC lines could potentially facilitate future non-human primate ESC studies. [ABSTRACT FROM AUTHOR]

Presents a study that attempted to detect antibodies to Neospora caninum in cattle by enzyme-linked immunosorbent assay with truncated NcSRS2 expressed in Escherichia coli. Background on the Neospora caninum parasite; Description of the invasion of the parasite into domestic animals such as cattle, dogs, goats, sheep and horses; Methods for detecting antibodies to N. canimum; Need for the implementation of immunization trials with animals to determine the potency of recombinant NcSRS2t expressed in E. coli as a potential subunit vaccine to control N. caninum infection.