Exogenous Slit2 does not affect ureteric branching or nephron formation during kidney development

In an attempt to elucidate the role of Slit2 invertebrate kidney development, the effect of adding exogenous human Slit2 protein (hSlit2) to developing murine metanephric kidney explants was examined. To confirm the activity of the recombinant Slit2 protein, neurons from 8 day old chick sympathetic nerve chain dorsal root ganglia were cultured with hSlit2 protein, which induced significant neurite branching and outgrowth. Using kidney explants as a model system, metanephric development in the presence of hSlit2 protein was examined. Addition of hSlit2 up to a final concentration of 1 mug/ml had no detectable effect on the formation of nephrons or on branching morphogenesis of the ureteric tree after 2 or 4 days in culture, as assessed via immunofluorescence for the markers WT1 and calbindin 28K respectively. Similarly, maturation of the nephrogenic mesenchyme occurred in a phenotypically normal fashion. In situ analysis of the Slit receptors, Robot and Robot, the vasculogenic markers VEGFA and Flk-1, and the stromal cell marker BF2 displayed no difference in comparison to controls.

Movement through slits: cellular migration via the Slit family

First isolated in the fly and now characterised in vertebrates, the Slit proteins have emerged as pivotal components controlling the guidance of axonal growth cones and the directional migration of neuronal precursors. As well as extensive expression during development of the central nervous system (CNS), the Slit proteins exhibit a striking array of expression sites in non-neuronal tissues, including the urogenital system, limb primordia and developing eye. Zebrafish Slit has been shown to mediate mesodermal migration during gastrulation, while Drosophila slit guides the migration of mesodermal cells during myogenesis. This suggests that the actions of these secreted molecules are not simply confined to the sphere of CNS development, but rather act in a more general fashion during development and throughout the lifetime of an organism. This review focuses on the non-neuronal activities of Slit proteins, highlighting a common role for the Slit family in cellular migration.

Facilitation of renal autoregulation by angiotensin II is mediated through modulation of nitric oxide

Aims: This study was designed to investigate the influence of angiotensin II (Ang II) and nitric oxide (NO) on autoregulation of renal perfusion. Methods: Autoregulation was investigated in isolated perfused kidneys (IPRK) from Sprague-Dawley rats during stepped increases in perfusion pressure. Results: Ang II (75-200 pM) produced dose-dependent enhancement of autoregulation whereas phenylephrine produced no enhancement and impaired autoregulation of GFR. Enhancement by Ang II was inhibited by the AT(1) antagonist, Losartan, and the superoxide scavenger, Tempol. Under control conditions nitric oxide synthase (NOS) inhibition by 10 muM N-omega-nitro-L-arginine methyl ester (L-NAME) facilitated autoregulation in the presence of non-specific cyclooxygenase (COX) inhibition by 10 muM indomethacin. Both COX and combined NOS/COX inhibition reduced the autoregulatory threshold concentration of Ang II. Facilitation by 100 pM Ang II was inhibited by 100 muM frusemide. Methacholine (50 nM) antagonised Ang II-facilitated autoregulation in the presence and absence of NOS/COX inhibition. Infusion of the NO donor, 1 muM sodium nitroprusside, inhibited L-NAME enhancement of autoregulation under control conditions and during Ang II infusion. Conclusions: The results suggest than an excess of NO impairs autoregulation under control conditions in the IPRK and that endogenous and exogenous NO, vasodilatory prostaglandins and endothelium-derived hyperpolarizing factor (EDHF) activity antagonise Ang II-facilitated autoregulation. Ang II also produced a counterregulatory vasodilatory response that included prostaglandin and NO release. We suggest that Ang II facilitates autoregulation by a tubuloglomerular feedback-dependent mechanism through AT(1) receptor-mediated depletion of nitric oxide, probably by stimulating generation of superoxide.

In vivo and in vitro models demonstrate a role for caveolin-1 in the pathogenesis of ischaemic acute renal failure

Caveolae and their proteins, the caveolins, transport macromolecules; compartmentalize signalling molecules; and are involved in various repair processes. There is little information regarding their role in the pathogenesis of significant renal syndromes such as acute renal failure (ARF). In this study, an in vivo rat model of 30 min bilateral renal ischaemia followed by reperfusion times from 4 h to 1 week was used to map the temporal and spatial association between caveolin-1 and tubular epithelial damage (desquamation, apoptosis, necrosis). An in vitro model of ischaemic ARF was also studied, where cultured renal tubular epithelial cells or arterial endothelial cells were subjected to injury initiators modelled on ischaemia-reperfusion (hypoxia, serum deprivation, free radical damage or hypoxia-hyperoxia). Expression of caveolin proteins was investigated using immunohistochemistry, immunoelectron microscopy, and immunoblots of whole cell, membrane or cytosol protein extracts. In vivo, healthy kidney had abundant caveolin-1 in vascular endothelial cells and also some expression in membrane surfaces of distal tubular epithelium. In the kidneys of ARF animals, punctate cytoplasmic localization of caveolin-1 was identified, with high intensity expression in injured proximal tubules that were losing basement membrane adhesion or were apoptotic, 24 h to 4 days after ischaemia-reperfusion. Western immunoblots indicated a marked increase in caveolin-1 expression in the cortex where some proximal tubular injury was located. In vitro, the main treatment-induced change in both cell types was translocation of caveolin-1 from the original plasma membrane site into membrane-associated sites in the cytoplasm. Overall, expression levels did not alter for whole cell extracts and the protein remained membrane-bound, as indicated by cell fractionation analyses. Caveolin-1 was also found to localize intensely within apoptotic cells. The results are indicative of a role for caveolin-1 in ARF-induced renal injury. Whether it functions for cell repair or death remains to be elucidated. Copyright (C) 2003 John Wiley Sons, Ltd.

ATP-dependent K+ channels in renal ischemia reperfusion injury

ATP-dependent K+ channels (K-ATP) account for most of the recycling of K+ which enters the proximal tubules cell via Na, K-ATPase. In the mitochondrial membrane, opening of these channels preserves mitochondrial viability and matrix volume during ischemia. We examined KATP channel modulation in renal ischemia-reperfusion injury (IRI), using an isolated perfused rat kidney (IPRK) model, in control, IRI, IRI + 200 muM diazoxide (a K-ATP opener), IRI + 10 muM glibenclamide (a K-ATP blocker) and IRI + 200 muM diazoxide + 10 muM glibenclamide groups. IRI was induced by 2 periods of warm ischemia, followed by 45 min of reperfusion. IRI significantly decreased glomerular filtration rate (GFR) and increased fractional excretion of sodium (FENa) (p < 0.01). Neither diazoxide nor glibenclamide had an effect on control kidney function other than an increase in renal vascular resistance produced by glibenclamide. Pretreatment with 200 muM diazoxide reduced the postischemic increase in FENa (p < 0.05). Adding 10 muM glibenclamide inhibited the diazoxide effect on postischemic FENa (p < 0.01). Histology showed that kidneys pretreated with glibenclamide demonstrated an increase in injure in the thick ascending limb of outer medulla (p < 0.05). Glibenclamide significantly decreased post ischemic renal vascular resistance (p < 0.05). but had no significant effect on other renal function parameters. Our results suggest that sodium reabsorption is improved by K-ATP activation and blockade of K-ATP channels during IRI has an injury enhancing effect on renal epithelial function and histology. This may be mediated through K-ATP modulation in cell and or mitochondrial inner membrane.

Repetitive brief ischemia: Intermittent reperfusion during ischemia ameliorates the extent of injury in the perfused kidney

Acute renal failure commonly follows reduced renal perfusion or ischemia. Reperfusion is essential for recovery but can itself cause functional and structural injury to the kidney. The separate contributions of ischemia and of reperfusion were examined in the isolated perfused rat kidney. Three groups were studied: brief (5 min) ischemia, 20 min ischemia, and repetitive brief ischemia (4 periods of 5 min) with repetitive intervening reperfusion of 5 min. A control group had no intervention, the three ischemia groups were given a baseline perfusion of 30 min before intervention and all groups were perfused for a total of 80 min. In addition, the effects of exogenous (NO)-N-. from sodium nitroprusside and xanthine oxidase inhibition by allopurinol were assessed in the repetitive brief ischemia-reperfusion model. Brief ischemia produced minimal morphological injury with near normal functional recovery. Repetitive brief ischemia reperfusion caused less functional and morphological injury than an equivalent single period of ischemia (20 min) suggesting that intermittent reperfusion is less injurious than ischemia alone over the time course of study. Pretreatment with allopurinol improved renal function after repetitive brief ischemia-reperfusion compared with the allopurinol-untreated repetitive brief ischemia-reperfusion group. Similarly, sodium nitroprusside reduced renal vascular resistance but did not improve the glomerular filtration rate or sodium reabsorption in the repetitive brief ischemia-reperfusion model. Thus, these studies show that the duration of uninterrupted ischemia is more critical than reperfusion in determining the extent of renal ischemia-reperfusion injury and that allopurinol, in particular, counteracts the oxidative stress of reperfusion.

The Wilms' Tumour Suppressor Protein, WT1, Undergoes CRM1-independent Nucleocytoplasmic Shuttling

The Wilms' tumour suppressor gene (WT1) encodes a zinc finger-containing nuclear protein essential for kidney and urogenital development. Initially considered a transcription factor, there is mounting evidence that WT1 has a role in post-transcriptional processing. Using the interspecies heterokaryon assay, we have demonstrated that WT1 can undergo nucleocytoplasmic shuttling. We have also mapped the region responsible for nuclear export to residues 182-324. Our data add further complexity to the role of WT1 in trancriptional and post-transcriptional regulation. (C) 2003 Published by Elsevier B.V. on behalf of the Federation of European Biochemical Societies.

Birth in the northern quoll, Dasyurus hallucatus (Marsupialia : Dasyuridae)

Birth has been observed in a number of marsupial species and, in the studies to date, the newborn have crawled up to or across to the pouch. The method of birth in the quoll, a dasyurid, differs greatly from that observed in other marsupials. Births were recorded at normal speed using hand-held digital video cameras. Birth was heralded by a release of about 1 mL of watery fluid from the urogenital sinus followed by gelatinous material contained in either one or two tubes emanating from the sinus. The newborn, still encased in their placental membranes, were in the gelatinous material within a column. To exit this column, they had to grasp a hair and wriggle about 1 cm across to the pouch. In the pouch the newborn young had to compete for a teat. Although the quolls possessed 8 teats, the number of young in the pouch immediately after birth was 17, 16, 6, 16, 13 and 11 for each of the 6 quolls filmed. While birth has been described previously in another two dasyurids, the observers did not describe birth as reported here for the quoll. Nevertheless the movement of the newborn from the sinus to the pouch is so quick that this could have previously been missed. Filming birth from beneath and from the side allowed for a greater understanding of the birth process. Further studies are required to determine whether this use of a gelatinous material is part of the birth process in all dasyurids.

Birth of Koalas Phascolarctos cinereus at Lone Pine Koala Sanctuary following artificial insemination

This paper documents the successful development of an artificial insemination (AI) programme for the Koala Phascolurctos cinereus. The protocols for trials involving two methods to induce ovulation and two insemination techniques are described. In Trial 1, interrupted coitus using a 'teaser'♂ successfully induced ovulation in nine Koalas. Five ♀♀ were inseminated while conscious using a modified 'foley catheter' (Cook insemination catheter) resulting in the births of two offspring. The other four ♀♀ were anaesthetized and inseminated using a technique which allowed visualization of the most cranial portion of the urogenital sinus, where semen was deposited using a 3.5 Fr. 'Tom-cat catheter' (urogen-itoscopic insemination). Three of the four ♀♀ inseminated by this technique produced pouch young. Microsatellite analysis of DNA from the pouch young excluded the teaser ♀♀ as possible sires, confirming that all offspring were sired by donor sperm. In Trial 2, eight ♀♀ were induced to ovulate by injecting them with 250 International Units of human chorionic gonadotrophin (hCG). A luteal phase was confirmed in all eight ♀♀ but only one gave birth following urogenitoscopic insemination. The Koala pouch young in this study are the first of any marsupial to be conceived and born following A1 procedures. Details of the A1 procedures used are presented and the significance of A1 to the conservation biology of P. cinereus discussed.

Sexual dysfunction in the Australian population

AIM: To describe the prevalence and forms of sexual dysfunction experienced by Australians, and compare these with people in the United States. METHODS: A cross sectional, telephone interview survey of a randomly selected sample of men (n = 876) and women (n = 908) aged between 18 and 59 years on the electoral roll in all states and territories of Australia (response rate = 61%) was conducted between November 1999 and April 2000. RESULTS: Large proportions of Australian men (55.0%) and women (60.5%) reported at least one sexual problem within the preceding year. More serious sexual dysfunction (> 3 symptoms) was observed in 13.2% of men and 19.7% of women. DISCUSSION: Australia and the USA are similar with regard to the high rate of symptoms of sexual dysfunction observed in the population. The low rate of treatment points to both patient and doctor reluctance to discuss sexual performance and practices.

Determinação do sexo em catfish de canal (Ictalurus punctatus) utilizando endoscopia

A identificação precoce do sexo em peixes é um procedimento importante para o desenvolvimento de estratégias que facilitem o manejo no plantel de reprodutores. A endoscopia pode ser utilizada para essa identificação, embora em peixes seu uso ainda seja restrito. Diante disso, o objetivo deste trabalho foi validar o uso do endoscópio rígido via poro urogenital para determinação do sexo do catfish de canal (Ictalurus punctatus). O equipamento utilizado foi um endoscópio Multipurpose Rigid™ 30º com 2,7 mm de diâmetro e 18 cm de comprimento. Foram examinados 60 peixes com peso variando entre 86 e 1.032 gramas nos quais foi introduzido o endoscópio via poro genital e, logo em seguida, eutanaziados para confirmação do sexo. Amostras de tecido gonadal foram coletadas para avaliação histológica do estádio de maturação e confirmação do sexo. Os resultados mostraram que dos 60 peixes examinados, 57 tiveram confirmação positiva do sexo via endoscopia, o que corresponde a 95% da amostra. A utilização da endoscopia para determinação do sexo em I. punctatus via poro genital é viável.

Role of malnutrition and parasite infections in the spatial variation in children’s anaemia risk in Northern Angola

Anaemia has a significant impact on child development and mortality and is a severe public health problem in most countries in sub-Saharan Africa. Nutritional and infectious causes of anaemia are geographically variable and anaemia maps based on information on the major aetiologies of anaemia are important for identifying communities most in need and the relative contribution of major causes. We investigated the consistency between ecological and individual-level approaches to anaemia mapping, by building spatial anaemia models for children aged ≤15 years using different modeling approaches. We aimed to a) quantify the role of malnutrition, malaria, Schistosoma haematobium and soil-transmitted helminths (STH) for anaemia endemicity in children aged ≤15 years and b) develop a high resolution predictive risk map of anaemia for the municipality of Dande in Northern Angola. We used parasitological survey data on children aged ≤15 years to build Bayesian geostatistical models of malaria (PfPR≤15), S. haematobium, Ascaris lumbricoides and Trichuris trichiura and predict small-scale spatial variation in these infections. The predictions and their associated uncertainty were used as inputs for a model of anemia prevalence to predict small-scale spatial variation of anaemia. Stunting, PfPR≤15, and S. haematobium infections were significantly associated with anaemia risk. An estimated 12.5%, 15.6%, and 9.8%, of anaemia cases could be averted by treating malnutrition, malaria, S. haematobium, respectively. Spatial clusters of high risk of anaemia (>86%) were identified. Using an individual-level approach to anaemia mapping at a small spatial scale, we found that anaemia in children aged ≤15 years is highly heterogeneous and that malnutrition and parasitic infections are important contributors to the spatial variation in anemia risk. The results presented in this study can help inform the integration of the current provincial malaria control program with ancillary micronutrient supplementation and control of neglected tropical diseases, such as urogenital schistosomiasis and STH infection.

Role of malnutrition and parasite infections in the spatial variation in children’s anaemia risk in northern Angola

Anaemia is known to have an impact on child development and mortality and is a severe public health problem in most countries in sub-Saharan Africa. We investigated the consistency between ecological and individual-level approaches to anaemia mapping by building spatial anaemia models for children aged ≤15 years using different modelling approaches. We aimed to (i) quantify the role of malnutrition, malaria, Schistosoma haematobium and soil-transmitted helminths (STHs) in anaemia endemicity; and (ii) develop a high resolution predictive risk map of anaemia for the municipality of Dande in northern Angola. We used parasitological survey data for children aged ≤15 years to build Bayesian geostatistical models of malaria (PfPR≤15), S. haematobium, Ascaris lumbricoides and Trichuris trichiura and predict small-scale spatial variations in these infections. Malnutrition, PfPR≤15, and S. haematobium infections were significantly associated with anaemia risk. An estimated 12.5%, 15.6% and 9.8% of anaemia cases could be averted by treating malnutrition, malaria and S. haematobium, respectively. Spatial clusters of high risk of anaemia (>86%) were identified. Using an individual-level approach to anaemia mapping at a small spatial scale, we found that anaemia in children aged ≤15 years is highly heterogeneous and that malnutrition and parasitic infections are important contributors to the spatial variation in anaemia risk. The results presented in this study can help inform the integration of the current provincial malaria control programme with ancillary micronutrient supplementation and control of neglected tropical diseases such as urogenital schistosomiasis and STH infections.