Multi-lineage Lung Regeneration by Stem Cell Transplantation across Major Genetic Barriers

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Differential response to hepatic differentiation stimuli of amniotic epithelial cells isolated from four regions of the amniotic membrane

Human Amniotic Epithelial Cells (hAEC) isolated from term placenta are a promising source for regenerative medicine. However, it has long been debated whether the hAEC population consists of heterogeneous or homogeneous cells. In a previous study, we investigated the characteristics of hAEC isolated from four different regions of the amniotic membrane finding significant heterogeneity. The aim of this study was to evaluate the hepatic differentiation capability of hAEC isolated from these four regions. Human term placentae were collected after caesarean section and hAEC were isolated from four regions of the amniotic membrane (R1-R4, according to their relative distance from the umbilical cord) and treated in hepatic differentiation conditions for 14 days. hAEC-derived hepatocyte-like cells showed marked differences in the expression of hepatic markers: R4 showed higher levels of Albumin and Hepatocyte Nuclear Factor (HNF) 4α whereas R1 expressed higher Cytochrome P450 enzymes, both at the gene and protein level. These preliminary results suggest that hAEC isolated from R1 and R4 of the amniotic membrane are more prone to hepatic differentiation. Therefore, the use of hAEC from a specific region of the amniotic membrane should be taken into consideration as it could have an impact on the outcome of therapeutic applications.     

Neurofilament-like and glial fibrillary acidic protein-like immunoreactivities in rat and guinea-pig sympathetic ganglia in situ and after perturbation

Summary The presence of neurofilament (NF)-like and glial fibrillary acidic protein (GFAP)-like immunoreactivities was studied in sympathetic ganglia of adult rats and guinea pigs during normal conditions and after perturbation. In the superior cervical ganglion (SCG) of normal rats, many ganglion cells and nerve fibers show NF immunoreactivity. Some of these nerve fibers disappear after preganglionic decentralization of SCG; this indicates the presence of a mixture of preand postganglionic NF-positive nerves in the ganglion. Cuts in both preand postganglionic nerves result in a marked increase in GFAP immunoreactivity in SCG, whereas NF immunoreactivity increases in nerve cell bodies after preganglionic cuts. Only a few ganglion cells show NF immunoreactivity in the normal SCG of guinea pig. All intraganglionic NF-positive nerves are of preganglionic origin; decentralization abolishes NF immunoreactivity in these nerve fibers. The inferior mesenteric ganglion, the hypogastric nerves and colonic nerves in guinea pigs contain large numbers of strongly NF-immunoreactive nerve fibers.When the SCG of adult rat is grafted to the anterior eye chamber of adult rat recipients, both ganglionic cell bodies and nerve fibers, forming on the host iris from the grafted ganglion, are NF-positive. As only the perikarya of these neurons normally exhibit NF immunoreactivity, and the terminal iris arborizations are NF-negative, it appears that the grafting procedure causes NF immunoreactivity to become more widespread in growing SCG neurons.     

Effects of induction current and other factors on large-scale electrofusion for pronuclear transplantation of mouse eggs

In this paper, we describe the procedure of large-scale and efficient electrofusion for pronuclear transplantation in mouse eggs and the tolerance of the eggs for electric stimulus, assessed in vitro and in vivo development. The fusion chamber was arranged in parallel by dielectrodes (30-mm length, 1-mm width, and 2-mm height), and 0.3 M mannitol in distilled water was used as a fusion solution. The agglutination cleavage of enucleated eggs with karyoplast was easily orientated in parallel with electrodes by alternating current between 100 and 500 kHz at 2 and 10 V/mm. Immediately after the orientation, a direct current of 150 V/mm was given for 200 μsec twice and repeated three times to induce fusion of the enucleated eggs with karyoplast. More than five eggs, at least, can be submitted to electrofusion at the same time. The eggs that were not fused were treated again in the same manner. The proportion of eggs fused with karyoplast was increased by preincubation in M16 medium prior to submitting them to the electrofusion. When the eggs were incubated for 60 min, 80% of them were fused with karyoplast by the first electric treatment; in contrast, only 19% of the eggs were fused if they were submitted to electrofusion directly. It was found that between the CD-1 and F1 strains there was a difference in tolerance of the eggs to electric stimulus and that this was depend on the nuclei but not on cytoplasm. The proportion of development to blastocyst in the eggs fused with the pronuclear karyoplast derived from F1 (75 and 71%) was twice that of the eggs fused with the pronuclei derived from CD-1 strain (25 and 37%). After transfer to recipients, live young were obtained from both the eggs fused with karyoplast following one or two electrofusion exposures.     

A comparative study on immunosuppressive effects of cyclosporin A and FK 506 on peripheral blood lymphocytes in dogs

Immunosuppressive effects of cyclosporin A (CsA) and FK 506 (FK) on peripheral blood lymphocytes were studied in dogs in respect to mixed lymphocyte reaction, proliferative responses to recombinant interleukin-2 (rIL-2), phytohemagglutinin (PHA) and concanavalin-A (Con-A); phenotypes of OKIa1, CD3, CD8 and surface IgM; cytotoxic activity against xenogeneic tumor cells. CsA (2.0 or 5.0 mg/kg, intravenously) or FK (0.16 mg/kg, intramuscularly) was given to mongrel dogs every morning for serial 21 days. The blood concentrations of CsA, measured as trough levels by fluorescence polarization method, ranged from 37 to 350 ng/ml in dogs administered at 2.0 mg/kg and from 170 to 894 ng/ml in dogs administered at 5.0 mg/kg during treatment, respectively. In dogs treated with FK at a dose of 0.16 mg/kg, the drug concentrations in the plasma during treatment ranged from 0.16 to 1.8 ng/ml. Mixed lymphocyte reaction and proliferative responses to rIL-2, PHA and Con-A, which were declined by CsA, were not affected by FK. In contrast, the proportion of OKIa1+ cells was not affected by CsA, whereas FK decreased the proportion of OKIa1+ cells progressively during the course of treatment. Cytotoxic activity was suppressed by both CsA and FK. These results possibly indicate that CsA and FK exert their immunosuppressive effects via different mechanisms.     

Line and crossing effects in a diallel mating system with highly inbred lines of White Leghorn chickens

Summary Seven highly inbred lines of White Leghorn chickens were used in a near complete diallel mating plan during eight years. The lines originated from three different base populations selected for egg weight. Average inbreeding coefficients of parents of chicks hatching in successive years were 0.75, 0.80, 0.84, 0.86, 0.89, 0.91, 0.93 and 0.94. The composition of line, specific combining ability and reciprocal effects and their estimated values are given. These effects were estimated for age at first egg (AFE), average weight of all eggs laid to 40 weeks (EW40), body weight at 40 weeks (BW40), number of eggs to 40 weeks (EP40) and number of eggs between 41 and 60 weeks (EP60). Records of 3247 hens surviving to 40 weeks and of 3133 birds to 60 weeks could be used. Large differences between line effects could be found in all traits. They were only partly due to the preceding selection in the base populations. All specific combining effects were in the expected direction, negative for AFE and positive for EW40, BW40, EP40 and EP60. Recovery of inbreeding depression inflated these effects rather substancially. Average heterosis, defined as the relative superiority of a line combination over the mid parent value, was –11.3%, 5.8%, 7.8%, 45.1% and 35.8% for AFE, EW40, BW40, EP40 and EP60 respectively. One line showed a relative superiority in AFE of -19.3% compared to about –7.9% for all other combinations. Reciprocal or sex-linked effects were generally smaller in all traits than specific combining effects, they were considerably smaller in AFE, EP40 and EP60. General reciprocal effects could be found for several lines in one or more traits. Offspring of two lines, when used as sire lines, showed a negative correlation between reciprocal effects of egg weight and body weight.     

Analysis of the Drosophila female sterile mutation fs(1) 1304 by pole cell transplantation experiments

The Drosophila melanogaster mutant fs(1)1304 is an ovary autonomous female sterile mutant that causes abnormal morphology of the egg. Vitellogenesis proceeds at an abnormally slow rate in homozygous females. We have used pole cell transplantation to construct germ line mosaics in order to determine whether the 1304 defect depends upon the genotype of the germ line cells (oocyte or nurse cells) or the somatic line (follicle cells). We have found that the germ line is the primary target tissue where the mutant gene is expressed.