Acute multiple organ failure in adult mice deleted for the developmental regulator Wt1

There is much interest in the mechanisms that regulate adult tissue homeostasis and their relationship to processes governing foetal development. Mice deleted for the Wilms' tumour gene, Wt1, lack kidneys, gonads, and spleen and die at mid-gestation due to defective coronary vasculature. Wt1 is vital for maintaining the mesenchymal-epithelial balance in these tissues and is required for the epithelial-to-mesenchyme transition (EMT) that generates coronary vascular progenitors. Although Wt1 is only expressed in rare cell populations in adults including glomerular podocytes, 1% of bone marrow cells, and mesothelium, we hypothesised that this might be important for homeostasis of adult tissues; hence, we deleted the gene ubiquitously in young and adult mice. Within just a few days, the mice suffered glomerulosclerosis, atrophy of the exocrine pancreas and spleen, severe reduction in bone and fat, and failure of erythropoiesis. FACS and culture experiments showed that Wt1 has an intrinsic role in both haematopoietic and mesenchymal stem cell lineages and suggest that defects within these contribute to the phenotypes we observe. We propose that glomerulosclerosis arises in part through down regulation of nephrin, a known Wt1 target gene. Protein profiling in mutant serum showed that there was no systemic inflammatory or nutritional response in the mutant mice. However, there was a dramatic reduction in circulating IGF-1 levels, which is likely to contribute to the bone and fat phenotypes. The reduction of IGF-1 did not result from a decrease in circulating GH, and there is no apparent pathology of the pituitary and adrenal glands. These findings 1) suggest that Wt1 is a major regulator of the homeostasis of some adult tissues, through both local and systemic actions; 2) highlight the differences between foetal and adult tissue regulation; 3) point to the importance of adult mesenchyme in tissue turnover.     

Platelet-activating factor and kinin-dependent vascular leakage as a novel functional activity of the soluble terminal complement complex

The infrequent occurrence of septic shock in patients with inherited deficiencies of the terminal complement components experiencing meningococcal disease led us to suspect that the terminal complement complex is involved in vascular leakage. To this end, the permeabilizing effect of the cytolytically inactive soluble terminal complement complex (SC5b-9) was tested in a Transwell system measuring the amount of fluorescein-labeled BSA (FITC-BSA) leaked through a monolayer of endothelial cells. The complex caused increased permeability to FITC-BSA after 15 min as opposed to the prompt response to bradykinin (BK). The effect of SC5b-9 was partially reduced by HOE-140 or CV-3988, two selective antagonists of BK B2 and platelet-activating factor receptors, respectively, and was completely neutralized by the mixture of the two antagonists. Also, DX-88, a specific inhibitor of kallikrein, partially inhibited the activity of SC5b-9. The permeabilizing factor(s) released after 30 min of incubation of endothelial cells with SC5b-9 caused a prompt leakage of albumin like BK. Intravital microscopy confirmed both the extravasation of circulating FITC-BSA across mesenteric microvessels 15 min after topical application of SC5b-9 and the complete neutralization by the mixture of HOE-140 and CV-3988. SC5b-9 induced opening of interendothelial junctions in mesenteric endothelium documented by transmission electron microscopy.     

Prolonged cell cycle transit is a defining and developmentally conserved hemopoietic stem cell property

Adult mouse hemopoietic stem cells (HSCs) are typically quiescent and enter and progress through the cell cycle rarely in steady-state bone marrow, but their rate of proliferation can be dramatically enhanced on demand. We have studied the cell cycle kinetics of HSCs in the developing fetal liver at a stage when they expand extensively. Despite that 100% of fetal liver HSCs divide within a 48-h period, their average cell cycle transit time (10.6 h) is twice that of their downstream progenitors, translating into a prolonged G(1) transit and a period of relative quiescence (G(0)). In agreement with their prolonged G(1) transit when compared with hemopoietic progenitors, competitive transplantation experiments demonstrate that fetal HSCs are highly enriched in G(1) but also functional in S-G(2)-M. This observation combined with experimental data demonstrating that adult HSCs forced to expand ex vivo also sustain a uniquely prolonged cell cycle and G(1) transit, demonstrate at least in part why purified HSCs at any state of development or condition are highly enriched in the G(0)-G(1) phases of the cell cycle. We propose that a uniquely prolonged cell cycle transit is a defining stem cell property, likely to be critical for their maintenance and self-renewal throughout development.     

Myeloid and lymphoid contribution to non-haematopoietic lineages through irradiation-induced heterotypic cell fusion

Recent studies have suggested that regeneration of non-haematopoietic cell lineages can occur through heterotypic cell fusion with haematopoietic cells of the myeloid lineage. Here we show that lymphocytes also form heterotypic-fusion hybrids with cardiomyocytes, skeletal muscle, hepatocytes and Purkinje neurons. However, through lineage fate-mapping we demonstrate that such in vivo fusion of lymphoid and myeloid blood cells does not occur to an appreciable extent in steady-state adult tissues or during normal development. Rather, fusion of blood cells with different non-haematopoietic cell types is induced by organ-specific injuries or whole-body irradiation, which has been used in previous studies to condition recipients of bone marrow transplants. Our findings demonstrate that blood cells of the lymphoid and myeloid lineages contribute to various non-haematopoietic tissues by forming rare fusion hybrids, but almost exclusively in response to injuries or inflammation.     

Experimental evidence for paternal effects on offspring growth rate in Arctic charr (Salvelinus alpinus)

Sexual selection theory predicts that females should choose males that signal viability and quality. However, few studies have found fitness benefits among females mating with highly ornamented males. Here, we use Arctic charr (Salvelinus alpinus), a teleost fish with no parental care, to investigate whether females could gain fitness benefits by mating with highly ornamented and large-sized males. Carotenoid-based coloration signalled by males during spawning is believed to be an indicator of good genes for this species. Paternal effects on offspring size (body length and dry body mass) were examined experimentally by crossing eggs and sperm in vitro from 12 females and 24 males in a split-brood design and raising larvae to 30 days past hatching. We clearly demonstrated that there was a relationship between offspring size and paternal coloration. However, a negative interaction between paternal length and coloration was evident for offspring length, indicating that positive effects of paternal coloration were only present for smaller males. Thus, the red spawning coloration of the male Arctic charr seems to be an indicator of good genes, but the effect of paternal coloration on offspring length, an indicator of 'offspring quality', is size dependent.     

Molecular and phenotypic divergence in the bluethroat (Luscinia svecica) subspecies complex

Subspecies complexes may provide valuable insights into the early stages of the speciation process. The bluethroat (Luscinia svecica) consists of many morphologically distinct subspecies that differ most strikingly in the ornamental colour pattern of the male throat. We investigated the genetic and phenotypic differentiation in this subspecies complex, using (i) microsatellite genotyping (11 loci) of a sample of 364 individuals from bluethroat populations in Europe and Asia, and (ii) spectrometric and morphological measurements of a sample of 131 museum skin specimens. Population genetic analyses, based on microsatellite allele frequency variation, revealed a slight but significant overall population differentiation (F(ST) = 0.042). There was a well-differentiated southern group of subspecies with white or no throat spots and a less-differentiated northern group of chestnut-spotted populations. Phylogenetic analyses indicated that the southern all-blue and white-spotted forms are ancestral to the chestnut-spotted subspecies. In addition to the qualitative variation in throat plumage pattern already described in the literature, we found significant quantitative variation among subspecies in hue, chroma and brightness of the ultraviolet (UV)/blue throat coloration, and this variation seemed to be unrelated to the phylogenetic distance between subspecies.     

An exploratory trial of cyclooxygenase type 2 inhibitor in HIV-1 infection: downregulated immune activation and improved T cell-dependent vaccine responses

Chronic HIV infection is characterized by chronic immune activation and dysfunctional T cells with elevated intracellular cyclic AMP (cAMP), which inhibits the T cell activation capability. cAMP may be induced by prostaglandin E(2) following lipopolysaccharide (LPS)-induced upregulation of cyclooxygenase type 2 (COX-2) in monocytes due to the elevated LPS levels in patients with chronic HIV infection. This hypothesis was tested using celecoxib, a COX-2 inhibitor, for 12 weeks in HIV-infected patients without antiretroviral treatment in a prospective, open, randomized exploratory trial. Thirty-one patients were randomized in the trial; 27 completed the study, including 13 patients on celecoxib. Celecoxib reduced chronic immune activation in terms of CD38 density on CD8(+) T cells (-24%; P = 0.04), IgA levels (P = 0.04), and a combined score for inflammatory markers (P < 0.05). Celecoxib further reduced the inhibitory surface receptor programmed death 1 (PD-1) on CD8(+) T cells (P = 0.01), including PD-1 on the HIV Gag-specific subset (P = 0.02), enhanced the number of CD3(+) CD4(+) CD25(+) CD127(lo/-) Treg or activated cells (P = 0.02), and improved humoral memory recall responses to a T cell-dependent vaccine (P = 0.04). HIV RNA (P = 0.06) and D dimers (P = 0.07) tended to increase in the controls, whereas interleukin-6 (IL-6) possibly decreased in the treatment arm (P = 0.10). In conclusion, celecoxib downmodulated the immune activation related to clinical progression of chronic HIV infection and improved T cell-dependent functions in vivo.     

Protection against Atlantic halibut nodavirus in turbot is induced by recombinant capsid protein vaccination but not following DNA vaccination

Fish nodaviruses (betanodaviruses) are small, non-enveloped icosahedral single-stranded positive-sense RNA viruses that can cause viral encephalopathy and retinopathy (VER) in a number of cultured marine teleost species, including Atlantic halibut (Hippoglossus hippoglossus). A recombinant protein vaccine and a DNA vaccine were produced, based on the same capsid-encoding region of the Atlantic halibut nodavirus (AHNV) genome, and tested for protection in juvenile turbot (Scophthalmus maximus). Vaccine efficacy was demonstrated in the fish vaccinated with recombinant capsid protein but not in the DNA-vaccinated fish, despite the fact that in vivo expression of the DNA vaccine-encoded antigen was confirmed by RNA in situ hybridisation and immunohistochemistry. Combined DNA and recombinant vaccine administration did not improve the effect of the latter. Surprisingly, fish vaccinated with 50 microg recombinant protein demonstrated a threefold lower survival rate than the two groups that received 10 microg recombinant protein. Neither the recombinant protein vaccine nor the DNA vaccine induced anti-viral antibodies 9 weeks after immunisation, while antibodies reactive with the recombinant protein were detectable mainly in fish vaccinated with 50 microg recombinant protein. The study also demonstrates evidence of viral replication inside the myocytes of intramuscularly challenged fish.