Regulation of autoinducer 2 production and luxS expression in a pathogenic Edwardsiella tarda strain

Edwardsiella tarda is a bacterial pathogen that can infect both humans and animals. TX1, an Ed. tarda strain isolated from diseased fish, was found to produce autoinducer 2 (Al-2)-like activity that was growth phase dependent and modulated by growth conditions. The gene coding for the Al-2 synthase was cloned from TX1 and designated luxS(Et). LuxS(Et) was able to complement the Al-2 mutant phenotype of Escherichia coli strain DH5 alpha. Expression Of luxS(Et) correlated with Al-2 activity and was increased by glucose and decreased by elevated temperature. The effect of glucose was shown to be mediated through the cAMP-CRP complex, which repressed luxS(Et) expression. Overexpression of luxS(Et) enhanced Al-2 activity in TX1, whereas disruption of luxS(Et) expression by antisense RNA interference (i) reduced the level of Al-2 activity, (ii) impaired bacterial growth under various conditions, (iii) weakened the expression of genes associated with the type III secretion system and biofilm formation, and (iv) attenuated bacterial virulence. Addition of exogenous Al-2 was able to complement the deficiencies in the expression of TTSS genes and biofilm production but failed to rescue the growth defects. Our results (i) demonstrated that the Al-2 activity in TX1 is controlled at least in part at the level of luxS(Et) expression, which in turn is regulated by growth conditions, and that the temporal expression of luxS(Et) is essential for optimal bacterial infection and survival; and (ii) suggested the existence in Ed. tarda of a LuxS/Al-2-mediated signal transduction pathway that regulates the production of virulence-associated elements.

Construction and characterization of a live, attenuated esrB mutant of Edwardsiella tarda and its potential as a vaccine against the haemorrhagic septicaemia in turbot, Scophthamus maximus (L.)

The esrB gene of Edwardsiella tarda, which encodes a regulator protein of the type III secretion system, was mutated by the unmarked deletion method and reintroduced by allelic exchange into the chromosome of E. tarda LSE40 by means of the suicide vector pRE 112. The LSE40 esrB mutant was highly attenuated when inoculated intraperitoneally into turbot Scophthamus maximus L., showing a 50% lethal dose of 10(8.1) cfu/fish. The esrB mutants were not recoverable from the internal organs at 14 days post-inoculation. Vaccination with a single dose of 10(5)-10(7) cfu/fish of the esrB mutant elicited significant protection against the wildtype strain of E. tarda LSE40 (relative percentage survival > 50%). The protection correlated well with the antibody titres in the serum of vaccinated fish. (c) 2006 Elsevier Ltd. All rights reserved.

REE and epsilon(Nd) of clay fractions in sediments from the eastern Pacific Ocean: Evidence for clay sources

Clay fractions in the non-calcareous surface sediments from the eastern Pacific were analyzed for clay minerals, REE and Nd-143/Nd-144. Montmorillonite/illite ratio (M/I ratio), total REE contents (Sigma REE), LREE/HREE ratio and cerium anomaly (delta Ce) may effectively indicate the genesis of clay minerals. Clay fractions with M/I ratio > 1, delta Ce < 0.85, Sigma REE > 400 mu g/g, LREE/HREE ratio approximate to 4, and REE patterns similar to those of pelagic sediments are terrigenous and autogenetic mixed clay fractions and contain more autogenetic montmorillonite. Clay fractions with M/I ratio < 1, delta Ce=0.86 to 1.5, Sigma REE=200 to 350 mu g/g, LREE/HREE ratio approximate to 6 and REE distribution patterns similar to that of China loess are identified as terrigenous clay fraction. The Nd-143/Nd-144 ratios or epsilon(Nd) values of clay fractions inherit the features of terrigenous sources of clay minerals. Clay fractions are divided into 4 types according to epsilon(Nd) values. Terrigenous clay minerals of type I with the eNd values of -8 to -6 originate mainly from North American fluvial deposits. Those of type 11 with the epsilon(Nd) Values of -9 to -7 are mainly from the East Asia and North American fluvial deposits. Those of type III with epsilon(Nd) values of -6 to -3 could come from the central and eastern Pacific volcanic islands. Those of type IV with epsilon(Nd) values of -13 to -12 may be from East Asia eolian. The terrigenous and autogenetic mixed clay fractions show patchy distributions, indicating that there are volcanic or hot-spot activities in the eastern Pacific plate, while the terrigenous clay fractions cover a large part of the study area, proving that the terrigenous clay minerals are dominant in the eastern Pacific.

Mineralogy of smectites in the surface sediments from the East Pacific and its significance

XRD, TEM, SEM and EDS are employed to analyze smectites in the clay fraction of the surface sediments from the East Pacific. It is shown from the XRD results that the clay fraction consists of about 20% smectites. Three types of smectites are identified, Fe-rich (Type I), Fe, Mg-rich (Type II) and Na,Ca smectite (Type III), and most of them are not well-crystallized. Type I is widely distributed in sediments, showing honeycomblike in the SEM, and aggregated or dispersive hairlike, or cloudy assemblage with a bit curl near its edge in the TEM. This type is considered to be typomorphic type of authigenic smectite in the East Pacific. Type II is similar to Type I in micromorphology in the TEM, showing a transition micronite, while Type III is tabletlike in the TEM with an unclear edge. Type I may be altered from volcanics and some of them even precipitated from the low subthermal water. Type II could also be formed in the ocean floor, while Type III comes from dry and distant continental area. This study suggests that the characteristic of chemical composition and morphology of smectite may give a clue to understand sediment source, origin of minerals and sedimentation in the deep sea.

Dynamics of the buoyant plume off the Pearl River Estuary in summer

Field measurements of salinity, wind and river discharge and numerical simulations of hydrodynamics from 1978 to 1984 are used to investigate the dynamics of the buoyant plume off the Pearl River Estuary (PRE), China during summer. The studies have shown that there are four major horizontal buoyant plume types in summer: Offshore Bulge Spreading (Type I), West Alongshore Spreading (Type II), East Offshore Spreading (Type III), and Symmetrical Alongshore Spreading (Type IV). River mouth conditions, winds and ambient coastal currents have inter-influences to the transport processes of the buoyant plume. It is found that all of the four types are surface-advected plumes by analysing the vertical characteristic of the plumes, and the monthly variations of the river discharge affect the plume size dominantly. The correlation coefficient between the PRE plume size and the river discharge reaches 0.85 during the high river discharge season. A wind strength index has been introduced to examine the wind effect. It is confirmed that winds play a significant role in forming the plume morphology. The alongshore wind stress and the coastal currents determine the alongshore plume spreading. The impact of the ambient currents such as Dongsha Current and South China Sea (SCS) Warm Current on the plume off the shelf has also assessed. The present study has demonstrated that both the river discharge and wind conditions affect the plume evolution.

陕北地区石家湾-李家岔探区三叠系延长组石油成藏条件分析

Shijiawan –Lijiacha area, lying on the northeastern part of the Shanbei Slope of Ordos Basin, was selected as studying area. The previous explorations proved that the 2nd segment and 6th segment of the Yanchang Formation are the most important oil-bearing formations. It is indicated that the sedimentary facies and reservoir characteristics restricted the hydrocarbon accumulation regularity. Therefore, with petrology methodologies, such as outcrop observation, core description, geophysical logging interpretation, thin section determination, scanning electron microscope, as well as rock property analysis, the reservoirs was were systematically studied and characterized. The sedimentary micro-facies, seals, reservoir-seal combines, migration pathways and entrapping modes were taken into account. The author tempted to establish a base for further studies on reservoirs and on petroleum geology, and to provide some reliably geological evidences for later prospect activities. It was found that the sediments in the 2nd and 3rd segments of the Yanchang Formation in Shijiawan –Lijiacha area were deposited in braided rivers, and most sandy-bodies were identified as channel sandbars. The 4+5th and 6th segments were principally deposited in deltaic-plain environment, consisting of corresponding sub-facies such as distributary channels, natural levee, crevasse-splay and marsh. The skeleton sandy-bodies were identified as sandy sediments of distributary channels. The sand grains in reservoir in studied area possess generally low mineralogical maturity and moderate structural maturity, and the form of pores may be classified into intergranular types and dissolved types. Most reservoirs of Yanchang Formation in Shijiawan –Lijiacha area belong to extreme low-porosity low-permeability ones (type III), and the 2nd sediments belongs to low permeability one (type II) and the 6th segment belong to super low-permeability one(type Ⅳ). The reservoirs in the 2nd segment behave more heterogeneous than those in the 6th segment. The statistic analysis results show that, for 6th and 4+5th segments, the high quality reservoir-seal combines may be found everywhere in the studied area except in the northwest and the southwest parts; and for 1st and 2nd segments, in the northeast, central and southwest parts Petroleum migration happened in the duration of the Early Cretaceous period in both lateral and vertical directions. The migration paths were mainly constructed by permeable sandy-bodies. The superimposed channel sandy-bodies consist of the principal part of the system of carriers. the vertical fractures, that may travel through the seals between reservoirs, offered the vertical paths for migrating oil. It may be synthesized that oil coming from south kitchens migrated first laterally in carriers in the 6th segment. When arrived at the studied area, oil will migration laterally or/and vertical within both the sandy-bodies and fractures, in a climbing-stair way. The results demonstrate that the oil was entrapped in traps structure-lithology and/or lithology traps. In some cases, the hydrodynamic force may help to trap oil. Accumulation of oil in the area was mainly controlled by sedimentary facies, seals, structure, and heterogeneity of reservoir in the 2nd, 4+5th and 6th segments. Especially, the oil distributions in both the 2nd and 6th segments were obviously influenced by seals in the 4+5th segment. The existence of seals in 1st segment seems important for accumulation in the 2nd segment.

港西地区河流相砂岩油藏剩余油分布研究

The Gangxi oil field has reached a stage of high water production. The reservoir parameters, such as reservoir physical characteristics, pore structure, fluid, have obviously changed. This thesis therefore carries out a study of these parameters that control reservoir characteristics, physical and chemical actions that have taken place within the reservoirs due to fluid injection, subsequent variations of reservoir macroscopic physical features, microscopic pore structures, seepages, and formation fluid properties. This study rebuilds a geologic model for this oil field, establishes a log-interpreting model, proposes a methodology for dealing with large pore channels and remnant oil distribution, and offers a basis for effective excavation of potential oil, recovery planning, and improvement of water-injection techniques. To resolve some concurrent key problems in the process of exploration of the Gangxi area, this thesis carries out a multidisciplinary research into reservoir geology, physical geography, reservoir engineering, and oil-water well testing. Taking sandstone and flow unit as objects, this study establishes a fine geologic model by a quantificational or semi-quantificational approach in order to understand the remnant oil distribution and the reservoir potential, and accordingly proposes a plan for further exploration. By rebuilding a geological model and applying reservoir-engineering methods, such as numerical simulation, this thesis studies the oil-water movement patterns and remnant-oil distribution, and further advances a deployment plan for the necessary adjustments and increase of recoverable reserves. Main achievements of this study are as follows: 1. The Minghazhen Formation in the Gangxi area is featured by medium-sinuosity river deposits, manifesting themselves as a transitional type between typical meandering and braided rivers. The main microfacies are products of main and branch channels, levee, inter-channel overflows and crevasse-splay floodplains. The Guantao Group is dominantly braided river deposit, and microfacies are mainly formed in channel bar, braided channel and overbank. Main lithofacies include conglomerate, sandstone, siltstone and shale, with sandstone facies being the principal type of the reservoir. 2. The reservoir flow unit of the Gangxi area can be divided into three types: Type I is a high-quality heterogeneous seepage unit, mainly distributed in main channel; Type II is a moderate-quality semi-heterogeneous seepage unit, mainly distributed in both main and branch channels, and partly seen within inter-channel overflow microfacies; Type III is a low-quality, relatively strong heterogeneous seepage unit, mainly distributed in inter-channel overflow microfacies and channel flanks. 3. Flow units and sedimentary microfacies have exerted relatively strong controls on the flowing of underground oil-water: (1) injection-production is often effective in the float units of Type I and II, whilst in the same group of injection-production wells, impellent velocity depends on flow unit types and injection-production spacing; (2) The injection-production of Type III flow unit between the injection-production wells of Type I and II flow units, however, are little effective; (3) there can form a seepage shield in composite channels between channels, leading to inefficient injection and production. 4. Mainly types of large-scale remnant-oil distribution are as follows: (1) remnant oil reservoir of Type III flow unit; (2) injection-production well group of remnant oil area of Type III flow unit; (3) remnant oil reservoirs that cannot be controlled by well network, including reservoir featured by injection without production, reservoir characterized by production without injection, and oil reservoir at which no well can arrive; (4) remnant oil area where injection-production system is not complete. 5. Utilizing different methods to deal with different sedimentary types, sub-dividing the columns of up to 900 wells into 76 chronostratigraphic units. Four transitional sandstone types are recognized, and contrast modes of different sandstone facies are summarized Analyzing in details the reservoirs of different quality by deciphering densely spaced well patterns, dividing microscopic facies and flow units, analyzing remnant oil distribution and its effect on injection-production pattern, and the heterogeneity. Theory foundation is therefore provided for further excavation of remnant oil. Re-evaluating well-log data. The understanding of water-flood layers and conductive formations in the Gangxi area have been considerably improved, and the original interpretations of 233 wells have changed by means of double checking. Variations of the reservoirs and the fluid and formation pressures after water injection are analyzed and summarized Studies are carried out of close elements of the reservoirs, fine reservoir types, oil-water distribution patterns, as well as factors controlling oil-gas enrichment. A static geological model and a prediction model of important tracts are established. Remaining recoverable reserves are calculated of all the oil wells and oil-sandstones. It is proposed that injection-production patterns of 348 oil-sandstones should be adjusted according to the analysis of adaptability of all kinds of sandstones in the injection-production wells.

东营凹陷沙三中亚段湖相浊积岩储层特征及油气成藏规律研究

Turbidity sandstone reservoirs have been an important field of hydrocarbon exploration and development in the basins all over the world, as well as in China. Lithologic pools are composed of turbidity sandstones and other sandstones are frequently found in the Jiyang Depression that is a Mesozoic-Cenozoic non-marine oil-bearing basin. The Dongying Sag lies in the sedimentary center of the basin. The subtle traps with turbidity reservoirs are generally difficult to be predicted and described by using current techniques. The studies on turbidity reservoirs plays thus an important theoretical and theoretical practical role in exploration and development in the Jiyang Depression. The attention is, in this thesis, focused on the petrologic properties and oil accumulating behaviors in lake turbidity sedimentary systems in the middle part of the third section of Shahejie Formation in the Dongying Sag, especially in Dongxin area, which lies on the central uplift of the Sag. The paper has disclosed the origin types of turbidity sandstones, distribution pattern and controlling factors of turbidity sandstones, and set up hydrocarbon accumulation patterns of the middle part of the third section of Shahejie Formation in Dongxin, based on nonmarine high resolution sequence stratigraphy, event sedimentology and new theories of hydrocarbon forming. By studying prediction method and technology of turbidity sandstone reservoirs, using precise geological model developing, new techniques of high resolution seismic inversion constrained by logging, the paper has forecast low permeability turbidity sandstone reservoirs and pointed out advantage exploration aims to progressive exploration and development. The paper has obtained mainly many productions and acknowledges as follows: 1.Turbidity sandstone reservoirs of the third section of Shahejie Formationin Dongying Sag are formed in such specifical geological background as rift and extension of basin. The inherited Dongying delta and transgression make up many turbidity distribution areas by overlaying and joining together. The hydrocarbon migrates from depression area to adjacent turbidity sandstone continuously. Accumulation area which is sufficient in oil is formed. 2.The paper has confirmed distinguishable sign of sequence boundary , established stratigraphic framework of Dongying Sag and realized isotime stratigraphic correlation. Es3 of Dongying delta is divided into eleven stages. Among them, the second period of the lower section in Es3, the sixth period of the middle section in Es3, the third period of the upper section in Es3 correspond to eleven sedimentary isotime surface in seismic profile, namely Es3 is classified into eleven Formations. 3.According to such the features of turbidity sandstone as deep in burial, small in area, strong in subtle property, overlaying and joining together and occurring in groups, management through fault and space variations of restriction quantum are realized and the forecast precision of turbidity sandstone by using precise geological model developing, new techniques of high resolution seismic inversion constrained by logging, based on the analysis of all kinds of interwell seismic inversion techniques. 4.According to the features of low permeable turbidity sandstone reservoirs, new method of log interpretation model is put forward. At the same time, distinguish technology of familiar low resistivity oil layer in the turbidity sandstone reservoirs is studied based on petrophysical laboratory work and "four properties" interrelationship between lithological physical Jogging and bearing hydrocarbon properties. Log interpretation model and reservoir index interpretation model of low resistivity oil layer are set up. So the log interpretation precision is improved. 5.The evolution law and its difference of the turbidity sandstone are embodies as follows: the source of sediments come from the south and east of the study area in the middle period of Es3. East source of sediments is pushed from west to east. However, the south source supply of sediments in the early and middle period of Es3 is in full, especially in Es3. subsequently, the supply is decreased gradually. Turbidity fan moves back toward the south and the size of fan is minished accordingly. The characteristic of turbidity sandstone in Dongying Sag is different in different structural positions. Dongxin in the middle-east of the central lift and Niuzhuang Sag He in Dongying delta front and prodelta deep lake subfacies. Although the turbidity sandstone of the two areas root in the Dongying delta sedimentary system, the sand body has different remarkably characteristic. 6.The sedimentary model of the turbiditys in study area have three types as follows: (1) collapse turbidity fan in respect of delta; (2) fault trench turbidity fan; (3) other types of microturbidity sandstone. Middle fan and outer fan, can be found mainly in sublacustrine fan. Middle fan includes braided channel microfacies, central microfacies and braided interchannel microfacies, which is main prospecting oil-bearing subfacies. The middle section of the third section of Shahejie Formation in study area (for example the central lift) can be divided into middle-lower and upper part. The middle-lower part is characteristic of turbidity fan. The upper part is sedimented mainly by delta-collapse fan. 7.The turbidity reservoirs of the middle part of the third section of Shahejie Formation in study area characterize by low maturity both in component and texture, strong in diagenesis and low in permeability. The reservoir can be classified into four types. Type III is the body of reservoir and comprises two types of H a and HI b. M a belongs to middle porosity - low permeability reservoir and distributes in the central lift. Hlb belongs to low porosity - low permeability and distributes in Haojia region. 8.A11 single sand body of lens turbidity reservoir of the middle part of the third section of Shahejie Formation in study area are surrounded by thick dark source rocks. The oil-water system is complex and behaves that every sandstone is single seal unit. The water body is 1/3-1-5 of the sand body. The edge water is not active. The gas exists in the top of reservoir in the form of mixed gas. For far-range turbidity fan with big scale channel, the area and volume of sand body is large and the gap is big in oil packing degree. There are lots of edge water and bottom water, and the latter increases rapidly during the course of development. 9.By exerting the modern hydrocarbon forming theories, the third section of Shahejie Formation in study area belongs to abnormally pressured fluid compartment. The lithological reservoir of the third section of Shahejie Formation is formed in the compartment. The reservoir-formed dynamic system belongs to lower self-source enclosed type. The result and the practice indicate that the form and accumulation of lithological oil reservoirs are controlled by the temperature and pressure of stratum, microfacies, thickness of sand body, fault and reservoir heterogeneity. 10. Based on studies above, the emphases focus on in south and north part of Dongying structure, west Dongxin region and south part Xinzhen structure in the application of production. The practice proves that the turbidity sandstone reservoirs in Ying 11 block and the fault-lithological reservoirs in Xin 133 block have been obtained significant breakthrough. The next target is still sandstone groups of the third section of Shahejie Formation in the bordering areas of Dongxin region for instance Xin 149 area, He 89 area, Ying 8 area etc.

Elevação do seio maxilar versus colocação de implantes curtos

Projeto de Pós-Graduação/Dissertação apresentado à Universidade Fernando Pessoa como parte dos requisitos para obtenção do grau de Mestre em Medicina Dentária

Hydrocarbon Potential of the Infra-salt Strata

This schematic geological cross-section of Angola offshore is representative of the majority of the Atlantic-type divergent margins. It illustrates the main geological features allowing to understand the different petroleum systems occurring, particularly, in South Atlantic divergent margins : (i) Pre-Pangea rocks (Precambrian granite-gneiss basement, volcanic rocks an/ or Paleozoic sediments, more or less, metamorphosed), which lie underneath the pre-rifting unconformity (PRU), in blue in the cross-section ; (ii) The rift-type basins developed during the lengthening of the Pangea supercontinent ; (iii) The breakup unconformity (BUU), which highlight the upper limit of the rift-type basins, in which organic rich lacustrine shales with a parallel internal configuration are potential source-rocks (organic matter type I) ; (iv) The SDRs (seaward dipping reflectors), which, generally, do not have any generating hydrocarbon potential (just 5 m of lacustrine shales are known in Austral basin) ; (v) The BUU is fossilized by SDRs (subaerial volcanism) or by margin infra-salt sediments (forming the mistakenly called by some American geoscientists "sag basin") ; (vi) The Loeme salt basin, which is a twin of the Brazilian salt basin, that is to say, that both basins have always been individualized ; (vii) The transgressive (backstepping) and regressive (forestepping) phases of the post-Pangea continental encroachment cycle ; (v) The interface between these sedimentary phases, correspond to the emplacement of potential marine source-rocks (organic matter type-II) ; (vi) Potential dispersive source rocks (organic matter type III) are possible in the regressive sedimentary interval.

Uniformly convergent finite element and finite difference methods for singularly perturbed ordinary differential equations

This thesis is concerned with uniformly convergent finite element and finite difference methods for numerically solving singularly perturbed two-point boundary value problems. We examine the following four problems: (i) high order problem of reaction-diffusion type; (ii) high order problem of convection-diffusion type; (iii) second order interior turning point problem; (iv) semilinear reaction-diffusion problem. Firstly, we consider high order problems of reaction-diffusion type and convection-diffusion type. Under suitable hypotheses, the coercivity of the associated bilinear forms is proved and representation results for the solutions of such problems are given. It is shown that, on an equidistant mesh, polynomial schemes cannot achieve a high order of convergence which is uniform in the perturbation parameter. Piecewise polynomial Galerkin finite element methods are then constructed on a Shishkin mesh. High order convergence results, which are uniform in the perturbation parameter, are obtained in various norms. Secondly, we investigate linear second order problems with interior turning points. Piecewise linear Galerkin finite element methods are generated on various piecewise equidistant meshes designed for such problems. These methods are shown to be convergent, uniformly in the singular perturbation parameter, in a weighted energy norm and the usual L2 norm. Finally, we deal with a semilinear reaction-diffusion problem. Asymptotic properties of solutions to this problem are discussed and analysed. Two simple finite difference schemes on Shishkin meshes are applied to the problem. They are proved to be uniformly convergent of second order and fourth order respectively. Existence and uniqueness of a solution to both schemes are investigated. Numerical results for the above methods are presented.

Chondrogenesis and mineralization during in vitro culture of human mesenchymal stem cells on three-dimensional woven scaffolds.

Human mesenchymal stem cells (hMSCs) and three-dimensional (3D) woven poly(ɛ-caprolactone) (PCL) scaffolds are promising tools for skeletal tissue engineering. We hypothesized that in vitro culture duration and medium additives can individually and interactively influence the structure, composition, mechanical, and molecular properties of engineered tissues based on hMSCs and 3D poly(ɛ-caprolactone). Bone marrow hMSCs were suspended in collagen gel, seeded on scaffolds, and cultured for 1, 21, or 45 days under chondrogenic and/or osteogenic conditions. Structure, composition, biomechanics, and gene expression were analyzed. In chondrogenic medium, cartilaginous tissue formed by day 21, and hypertrophic mineralization was observed in the newly formed extracellular matrix at the interface with underlying scaffold by day 45. Glycosaminoglycan, hydroxyproline, and calcium contents, and alkaline phosphatase activity depended on culture duration and medium additives, with significant interactive effects (all p < 0.0001). The 45-day constructs exhibited mechanical properties on the order of magnitude of native articular cartilage (aggregate, Young's, and shear moduli of 0.15, 0.12, and 0.033 MPa, respectively). Gene expression was characteristic of chondrogenesis and endochondral bone formation, with sequential regulation of Sox-9, collagen type II, aggrecan, core binding factor alpha 1 (Cbfα1)/Runx2, bone sialoprotein, bone morphogenetic protein-2, and osteocalcin. In contrast, osteogenic medium produced limited osteogenesis. Long-term culture of hMSC on 3D scaffolds resulted in chondrogenesis and regional mineralization at the interface between soft, newly formed engineered cartilage, and stiffer underlying scaffold. These findings merit consideration when developing grafts for osteochondral defect repair.

Irisin and FNDC5 in retrospect: An exercise hormone or a transmembrane receptor?

FNDC5 (fibronectin domain-containing [protein] 5) was initially discovered and characterized by two groups in 2002. In 2011 FNDC5 burst into prominence as the parent of irisin, a small protein containing the fibronectin type III domain. Irisin was proposed to be secreted by skeletal muscle cells in response to exercise, and to circulate to fat tissue where it induced a transition to brown fat. Since brown fat results in dissipation of energy, this pathway is of considerable interest for metabolism and obesity. Here I review the original discoveries of FNDC5 and the more recent discovery of irisin. I note in particular three problems in the characterization of irisin: the antibodies used to detect irisin in plasma lack validity; the recombinant protein used to demonstrate activity in cell culture was severely truncated; and the degree of shedding of soluble irisin from the cell surface has not been quantitated. The original discovery proposing that FNDC5 may be a transmembrane receptor may deserve a new look.

Irisin - a myth rather than an exercise-inducible myokine.

The myokine irisin is supposed to be cleaved from a transmembrane precursor, FNDC5 (fibronectin type III domain containing 5), and to mediate beneficial effects of exercise on human metabolism. However, evidence for irisin circulating in blood is largely based on commercial ELISA kits which are based on polyclonal antibodies (pAbs) not previously tested for cross-reacting serum proteins. We have analyzed four commercial pAbs by Western blotting, which revealed prominent cross-reactivity with non-specific proteins in human and animal sera. Using recombinant glycosylated and non-glycosylated irisin as positive controls, we found no immune-reactive bands of the expected size in any biological samples. A FNDC5 signature was identified at ~20 kDa by mass spectrometry in human serum but was not detected by the commercial pAbs tested. Our results call into question all previous data obtained with commercial ELISA kits for irisin, and provide evidence against a physiological role for irisin in humans and other species.

Rare hereditary COL4A3/COL4A4 variants may be mistaken for familial focal segmental glomerulosclerosis.

Focal segmental glomerulosclerosis (FSGS) is a histological lesion with many causes, including inherited genetic defects, with significant proteinuria being the predominant clinical finding at presentation. Mutations in COL4A3 and COL4A4 are known to cause Alport syndrome (AS), thin basement membrane nephropathy, and to result in pathognomonic glomerular basement membrane (GBM) findings. Secondary FSGS is known to develop in classic AS at later stages of the disease. Here, we present seven families with rare or novel variants in COL4A3 or COL4A4 (six with single and one with two heterozygous variants) from a cohort of 70 families with a diagnosis of hereditary FSGS. The predominant clinical finding at diagnosis was proteinuria associated with hematuria. In all seven families, there were individuals with nephrotic-range proteinuria with histologic features of FSGS by light microscopy. In one family, electron microscopy showed thin GBM, but four other families had variable findings inconsistent with classical Alport nephritis. There was no recurrence of disease after kidney transplantation. Families with COL4A3 and COL4A4 variants that segregated with disease represent 10% of our cohort. Thus, COL4A3 and COL4A4 variants should be considered in the interpretation of next-generation sequencing data from such patients. Furthermore, this study illustrates the power of molecular genetic diagnostics in the clarification of renal phenotypes.