Characterization of renal interstitial fibroblast-specific protein 1/S100A4-positive cells in healthy and inflamed rodent kidneys

Fibrosis is considered as a central factor in the loss of renal function in chronic kidney diseases. The origin of fibroblasts and myofibroblasts that accumulate in the interstitium of the diseased kidney is still a matter of debate. It has been shown that accumulation of myofibroblasts in inflamed and fibrotic kidneys is associated with upregulation of fibroblast-specific protein 1 (FSP1, S100A4), not only in the renal interstitium but also in the injured renal epithelia. The tubular expression of FSP1 has been taken as evidence of myofibroblast formation by epithelial–mesenchymal transition (EMT). The identity of FSP1/S100A4 cells has not been defined in detail. We originally intended to use FSP1/S100A4 as a marker of putative EMT in a model of distal tubular injury. However, since the immunoreactivity of FSP1 did not seem to fit with the distribution and shape of fibroblasts or myofibroblasts, we undertook the characterization of FSP1/S100A4-expressing cells in the interstitium of rodent kidneys. We performed immunolabeling for FSP1/S100A4 on thin cryostat sections of perfusion-fixed rat and mouse kidneys with peritubular inflammation, induced by thiazides and glomerulonephritis, respectively, in combination with ecto-5-nucleotidase (5NT), recognizing local cortical peritubular fibroblasts, with CD45, MHC class II, CD3, CD4 and Thy 1, recognizing mononuclear cells, with alpha smooth muscle actin (SMA), as marker for myofibroblasts, and vimentin for intracellular intermediate filaments in cells of mesenchymal origin. In the healthy interstitium of rodents the rare FSP1/S100A4+ cells consistently co-expressed CD45 or lymphocyte surface molecules. Around the injured distal tubules of rats treated for 3–4 days with thiazides, FSP1+/S100A4+, 5NT+, SMA+, CD45+ and MHC class II+ cells accumulated. FSP1+/S100A4+ cells consistently co-expressed CD45. In the inflamed regions, SMA was co-expressed by 5NT+ cells. In glomerulonephritic mice, FSP1+/S100A4+ cells co-expressed Thy 1, CD4 or CD3. Thus, in the inflamed interstitium around distal tubules of rats and of glomerulonephritic mice, the majority of FSP1+ cells express markers of mononuclear cells. Consequently, the usefulness of FSP1/S100A4 as a tool for detection of (myo)fibroblasts in inflamed kidneys and of EMT in vivo is put into question. In the given rat model the consistent co-expression of SMA and 5NT suggests that myofibroblasts originate from resident peritubular fibroblasts.Ivan Hegyi and Michel Le Hir contributed equally to the study     

Reduced compensatory growth capacity in mistimed broods of a migratory passerine

Phenotypic plasticity has recently been proposed to increase population viability when rapid anthropogenic environmental changes cannot be tracked by means of evolution. This assumes that environmental changes do not constrain phenotypic plasticity itself, which has rarely been examined in natural populations. In areas of climate warming, many long-distance migratory birds breed increasingly late relative to the period of peak food supply, and the temporal mismatch may constrain plastic life-history traits such as nestling growth. We combined 23 years of food availability and breeding data with a 3-year experimental manipulation of nestling growth trajectories in a Central European population of collared flycatchers (Ficedula albicollis) to examine the potential impact of climate-related mistimed breeding on nestling developmental plasticity. Timing of the food peak was predicted by winter climate, and the median hatching date of broods was earlier in springs with earlier food peaks. However, the adjustment of hatching date was incomplete and the population largely missed the food peak in years with very early food peaks. After imposing a temporary, experimental food shortage on nestlings, the extent of compensatory growth in body mass differed among years, and this difference was apparently related to the distance of hatching dates from the yearly food peak. Growth compensation declined with distance from the peak. These results suggest that mistimed phenology may not only create permanently adverse conditions for migratory species but it may also constrain the plastic responses of individuals to temporary disturbances. Therefore, climate change may not only favour but also restrict phenotypic plasticity.     

When to measure plumage reflectance: a lesson from Collared Flycatchers Ficedula albicollis

Sexually selected colour traits of bird plumage are widely studied. Although the plumage is replaced only at one or two yearly moults, plumage colour has long been shown to change between moults. Nevertheless, most studies measure colour weeks to months after the courtship period, typically at nestling rearing, and it is unclear whether these measurements yield relevant data concerning the primary process of sexual selection. Here we analyse repeated spectrometric data taken from male Collared Flycatchers during social courtship and nestling rearing. We show that some spectral traits are not correlated between the two measurements and that within‐individual correlation declines significantly with the likely exposure of the plumage area to damage and soiling. There is an overall decline in spectral trait exaggeration during breeding, but trait decline is not closely related to measurement latency, especially not in the damage‐exposed areas. Finally, sexual selection estimates differ depending on whether they are derived from spectra measured during courtship or during nestling rearing. These results suggest that, contrary to current practice, measurements of plumage reflectance should be made during the primary period of sexual signalling. Spectral trait decline during breeding could also be studied as a possible signal for mates and neighbours.     

Upregulation of CYP2e1 and CYP3a activities in histamine-deficient histidine decarboxylase gene targeted mice

Histamine is a biogenic amine with multiple physiological functions. Its importance in allergic inflammation is well characterized; moreover, it plays a role in the regulation of gastric acid production, various hypothalamic functions, such as food uptake, and enhancing TH2 balance during immune responses. Using histidine decarboxylase gene targeted (HDC(-/-)) BALB/c mice, we studied the effect of the absence of histamine on four cytochrome p450 enzyme activities. Their selective substrates were measured: ethoxyresorufin O-dealkylase activity of CYP1A, pentoxyresorufin O-dealkylase activity of CYP2B, chlorzoxazone 6-hydroxylase activity of CYP2E1 and ethylmorphine N-demethylase activity of CYP3A.The results indicate a significant elevation of CYP2E1 and CYP3A activities, however, no change in CYP1A and CYP2B activities was seen in HDC targeted mice compared to wild type controls with identical genetic backgrounds.     

Expression of truncated PrP targeted to Purkinje cells of PrP knockout mice causes Purkinje cell death and ataxia

PrP knockout mice with disruption of only the PrP-encoding region (Zürich I-type) remain healthy, whereas mice with deletions extending upstream of the PrP-encoding exon (Nagasaki-type) suffer Purkinje cell loss and ataxia, associated with ectopic expression of Doppel in brain, particularly in Purkinje cells. The phenotype is abrogated by co-expression of full-length PrP. Doppel is 25% similar to PrP, has the same globular fold, but lacks the flexible N-terminal tail. We now show that in Zürich I-type PrP-null mice, expression of N-terminally truncated PrP targeted to Purkinje cells also leads to Purkinje cell loss and ataxia, which are reversed by PrP. Doppel and truncated PrP probably cause Purkinje cell degeneration by the same mechanism.     

Bird predation by tawny owls (Strix aluco L.) and its effect on the reproductive performance of tits

The density of great tit Parus major L. and blue tit Parus caeruleus L. was artificially increased by placing nest-box colonies for these species in the vicinity of the nests of breeding tawny owls during 1993–1997. Bird prey composition in the owl nests, the proportion of parents disappearing from the breeding tit populations and the reproductive performance of the widowed parents were analysed. The frequency of predation on tits by tawny owls was greater in areas where tit density had been artificially increased. Owls preyed more on tits during the feeding period of owlets than during the incubation period and more in years when snow covered the ground during the incubation period than when it did not. Mortality due to predation was male biased and more females lost their mates in populations breeding near tawny owl nests. Reproductive performance of the widowed parents was lower and their body weights were lighter at the end of the nestling period than those found in birds rearing youngs with their mates. Predation by owls increased the between-year turnover in the breeding tit population: widowed parents did not return to the nesting site for the next breeding season.     

New Insights into the Methodology of L-Arginine-Induced Acute Pancreatitis

Animal models are ideal to study the pathomechanism and therapy of acute pancreatitis (AP). The use of L-arginine-induced AP model is nowadays becoming increasingly popular in mice. However, carefully looking through the literature, marked differences in disease severity could be observed. In fact, while setting up the L-arginine (2×4 g/kg i.p.)-induced AP model in BALB/c mice, we found a relatively low rate (around 15%) of pancreatic necrosis, whereas others have detected much higher rates (up to 55%). We suspected that this may be due to differences between mouse strains. We administered various concentrations (5–30%, pH = 7.4) and doses (2×4, 3×3, or 4×2.5 g/kg) of L-arginine-HCl in BALB/c, FVB/n and C57BL/6 mice. The potential gender-specific effect of L-arginine was investigated in C57BL/6 mice. The fate of mice in response to the i.p. injections of L arginine followed one of three courses. Some mice (1) developed severe AP or (2) remained AP-free by 72 h, whereas others (3) had to be euthanized (to avoid their death, which was caused by the high dose of L-arginine and not AP) within 12 h., In FVB/n and C57BL/6 mice, the pancreatic necrosis rate (about 50%) was significantly higher than that observed in BALB/c mice using 2×4 g/kg 10% L–arginine, but euthanasia was necessary in a large proportion of animals, The i.p. injection of lower L-arginine concentrations (e.g. 5–8%) in case of the 2×4 g/kg dose, or other L-arginine doses (3×3 or 4×2.5 g/kg, 10%) were better for inducing AP. We could not detect any significant differences between the AP severity of male and female mice. Taken together, when setting up the L-arginine-induced AP model, there are several important factors that are worth consideration such as the dose and concentration of the administered L arginine-HCl solution and also the strain of mice.     

Automatic target selection for structural genomics on eukaryotes

A central goal of structural genomics is to experimentally determine representative structures for all protein families. At least 14 structural genomics pilot projects are currently investigating the feasibility of high-throughput structure determination; the National Institutes of Health funded nine of these in the United States. Initiatives differ in the particular subset of "all families" on which they focus. At the NorthEast Structural Genomics consortium (NESG), we target eukaryotic protein domain families. The automatic target selection procedure has three aims: 1) identify all protein domain families from currently five entirely sequenced eukaryotic target organisms based on their sequence homology, 2) discard those families that can be modeled on the basis of structural information already present in the PDB, and 3) target representatives of the remaining families for structure determination. To guarantee that all members of one family share a common foldlike region, we had to begin by dissecting proteins into structural domain-like regions before clustering. Our hierarchical approach, CHOP, utilizing homology to PrISM, Pfam-A, and SWISS-PROT chopped the 103,796 eukaryotic proteins/ORFs into 247,222 fragments. Of these fragments, 122,999 appeared suitable targets that were grouped into >27,000 singletons and >18,000 multifragment clusters. Thus, our results suggested that it might be necessary to determine >40,000 structures to minimally cover the subset of five eukaryotic proteomes.     

Prions and the lymphoreticular system

Following intracerebral or peripheral inoculation of mice with scrapie prions, infectivity accumulates first in the spleen and only later in the brain. In the spleen of scrapie-infected mice, prions were found in association with T and B lymphocytes and to a somewhat lesser degree with the stroma, which contains the follicular dendritic cells (FDCs) but not with non-B, non-T cells; strikingly, no infectivity was found in lymphocytes from blood of the same mice. Transgenic PrP knockout mice expressing PrP restricted to either B or T lymphocytes show no prion replication in the lymphoreticular system. Therefore, splenic lymphocytes either acquire prions from another source or replicate them in dependency on other PrP-expressing cells. The essential role of FDCs in prion replication in spleen was shown by treating mice with soluble lymphotoxin-beta receptor, which led to disappearance of mature FDCs from the spleen and concomitantly abolished splenic prion accumulation and retarded neuroinvasion following intraperitoneal scrapie inoculation.