Landscape genetics informs mesohabitat preference and conservation priorities for a surrogate indicator species in a highly fragmented river system.

Poor dispersal species represent conservative benchmarks for biodiversity management because they provide insights into ecological processes influenced by habitat fragmentation that are less evident in more dispersive organisms. Here we used the poorly dispersive and threatened river blackfish (Gadopsis marmoratus) as a surrogate indicator system for assessing the effects of fragmentation in highly modified river basins and for prioritizing basin-wide management strategies. We combined individual, population and landscape-based approaches to analyze genetic variation in samples spanning the distribution of the species in Australia's Murray-Darling Basin, one of the world's most degraded freshwater systems. Our results indicate that G. marmoratus displays the hallmark of severe habitat fragmentation with notably scattered, small and demographically isolated populations with very low genetic diversity-a pattern found not only between regions and catchments but also between streams within catchments. By using hierarchically nested population sampling and assessing relationships between genetic uniqueness and genetic diversity across populations, we developed a spatial management framework that includes the selection of populations in need of genetic rescue. Landscape genetics provided an environmental criterion to identify associations between landscape features and ecological processes. Our results further our understanding of the impact that habitat quality and quantity has on habitat specialists with similarly low dispersal. They should also have practical applications for prioritizing both large- and small-scale conservation management actions for organisms inhabiting highly fragmented ecosystems.

Historic change in catchment land use and metal loading to Sydney estuary, Australia (1788-2010).

Sydney estuary has a long history of environmental degradation and is one of the most modified water ways in Australia due to a highly urbanised catchment (~77 %) and a high population (4.6 million). The objectives of the present study were to map historical land use change from European settlement (1788) to 2010 to determine catchment evolutionary pathways and to estimate catchment loading (total suspended solids, Cu, Pb and Zn) to the estuary over this period. Land use distribution in Sydney catchment, determined for seven time horizons over this period, indicated that a substantial increase in residential land use through subdivision of large estates and an increase in road area resulted in a marked increase in metal loading to Sydney estuary between 1892 and 1936. The decline in industrial activity from a maximum in 1978 (3.9 %) to 1.8 % in 2010 and the introduction of unleaded fuel during this time was accompanied by reduction in metal loading to the estuary. Land use time horizon maps enabled the creation of novel, ternary diagrams to represent temporal evolution in catchment land use. The 15 sub-catchments of Sydney estuary were combined into three major catchment categories, i.e., urban, dense urban and commercial. Present-day annual discharge of stormwater from the Sydney catchment was calculated to be 466,000 ML and annual loadings of total suspended sediment (TSS), Cu, Pb and Zn in tonnes were 49,239, 27, 37 and 57, respectively. Stormwater has superseded industry as the main source of anthropogenic metals to this estuary in recent times.

Growth and decline of shoreline industry in Sydney estuary (Australia) and influence on adjacent estuarine sediments.

Sydney estuary (Australia), like many urbanised waterways, is degraded due to an extended history of anthropogenic activity. Two major sources of contamination to this estuary are discharge by former shoreline industries and historic and contemporary catchment stormwater. The objectives of the present study were to document changes in shoreline land use from European settlement to the present day and determine the influence of this trend on the metal content of adjacent estuarine sediments. Temporal analysis of land use for seven time horizons between 1788 and 2010 showed rapid expansion of industry along much of the Sydney estuary foreshore soon after European settlement due to the benefits of easy and inexpensive access and readily available water for cooling and power. Shoreline industry attained maximum development in 1978 (32-km length) and declined rapidly to the present-day (9-km length) through redevelopment of industrial sites into medium- to high-density, high-value residential housing. Cores taken adjacent to 11 long-term industrial sites showed that past industrial practices contributed significantly to contamination of estuarine sediment. Subsurface metal concentrations were up to 35 times that of present-day surface sediment and over 100 times greater than natural background concentrations. Sedimentation rates for areas adjacent to shoreline industry were between 0.6 and 2.5 cm/year, and relaxation times were estimated at 50 to 100 years. Natural relaxation and non-disturbance of sediments may be the best management practice in most locations.

Tumor necrosis factor-alpha mediates osteopenia caused by depletion of antioxidants.

We recently found that estrogen deficiency leads to a lowering of thiol antioxidant defenses in rodent bone. Moreover, administration of agents that increase the concentration in bone of glutathione, the main intracellular antioxidant, prevented estrogen-deficiency bone loss, whereas depletion of glutathione by buthionine sulfoximine (BSO) administration provoked substantial bone loss. It has been shown that the estrogen-deficiency bone loss is dependent on TNFalpha signaling. Therefore, a model in which estrogen deficiency causes bone loss by lowering antioxidant defenses predicts that the osteopenia caused by lowering antioxidant defenses should similarly depend on TNFalpha signaling. We found that the loss of bone caused by either BSO administration or ovariectomy was inhibited by administration of soluble TNFalpha receptors and abrogated in mice deleted for TNFalpha gene expression. In both circumstances, lack of TNFalpha signaling prevented the increase in bone resorption and the deficit in bone formation that otherwise occurred. Thus, depletion of thiol antioxidants by BSO, like ovariectomy, causes bone loss through TNFalpha signaling. Furthermore, in ovariectomized mice treated with soluble TNFalpha receptors, thiol antioxidant defenses in bone remained low, despite inhibition of bone loss. This suggests that the low levels of antioxidants in bone seen after ovariectomy are the cause, rather than the effect, of the increased resorption. These experiments are consistent with a model for estrogen-deficiency bone loss in which estrogen deficiency lowers thiol antioxidant defenses in bone cells, thereby increasing reactive oxygen species levels, which in turn induce expression of TNFalpha, which causes loss of bone.

FLT3 ligand can substitute for macrophage colony-stimulating factor in support of osteoclast differentiation and function.

Although bone resorption and osteoclast numbers are reduced in osteopetrotic (op/op) mice, osteoclasts are nevertheless present and functional, despite the absence of macrophage colony-stimulating factor (M-CSF). This suggests that alternative factors can partly compensate for the crucial actions of M-CSF in osteoclast induction. It was found that when nonadherent bone marrow cells were incubated in RANKL with Flt3 ligand (FL) without exogenous M-CSF, tartrate-resistance acid phosphatase (TRAP)-positive cells were formed, and bone resorption occurred. Without FL, only macrophagelike TRAP-negative cells were present. Granulocyte-macrophage CSF, stem cell factor, interleukin-3, and vascular endothelial growth factor could not similarly replace the need for M-CSF. TRAP-positive cell induction in FL was not due to synergy with M-CSF produced by the bone marrow cells themselves because FL also enabled their formation from the hemopoietic cells of op/op mice, which lack any M-CSF. FL appeared to substitute for M-CSF by supporting the differentiation of adherent cells that express mRNA for RANK and responsiveness to RANKL. To determine whether FL can account for the compensation for M-CSF deficiency that occurs in vivo, FL signaling was blockaded in op/op mice by the injection of soluble recombinant Flt3. It was found that the soluble receptor induced a substantial decrease in osteoclast number, strongly suggesting that FL is responsible for the partial compensation for M-CSF deficiency that occurs in these mice.

Modular synthesis of pi-acceptor cyclophanes derived from 1,4,5,8-naphthalenetetracarboxylic diimide and 1,5-dinitronaphthalene.

Three neutral cyclophanes were synthesized, and their association with indole, an aromatic pi-donor, was studied. The cyclophanes were designed to contain a rigid, hydrophobic binding cavity with 1,4,5,8-naphthalenetetracarboxylic diimide or 1,5-dinitronaphthalene as the pi-acceptor. Two of the cyclophanes also contain a (S)-(valine-leucine-alanine) tripeptide unit to provide chiral hydrogen bonding interactions with guest molecules. Despite the fact that these cyclophanes contain a hydrophobic binding cavity of appropriate dimensions, their association with indole is very weak. In the case of cyclophanes derived from 1,5-dinitronaphthalene, steric interactions force the nitro groups out of the plane of the naphthalene ring, diminishing their effectiveness as pi-acceptors. A simple UV--visible titrimetric method, using N,N,N',N'-tetramethyl-1,4-phenylenediamine (TMPD) as a pi-donor, was used to rank the pi-acceptor strength of these and other aromatic units. These titrations show that 1,4,5,8-naphthalenetetracarboxylic diimide and 1,5-dinitronaphthalene derivatives are weaker pi-acceptors than viologens, which make good pi-acceptor cyclophanes. Methyl viologen is in turn a weaker pi-acceptor than anthaquinone disulfonate, suggesting that the latter may serve as a useful building block for pi-accepting cyclophane hosts.

TGF-beta 1 and IFN-gamma direct macrophage activation by TNF-alpha to osteoclastic or cytocidal phenotype.

TNF-related activation-induced cytokine (TRANCE; also called receptor activator of NF-kappaB ligand (RANKL), osteoclast differentiation factor (ODF), osteoprotegerin ligand (OPGL), and TNFSF11) induces the differentiation of progenitors of the mononuclear phagocyte lineage into osteoclasts in the presence of M-CSF. Surprisingly, in view of its potent ability to induce inflammation and activate macrophage cytocidal function, TNF-alpha has also been found to induce osteoclast-like cells in vitro under similar conditions. This raises questions concerning both the nature of osteoclasts and the mechanism of lineage choice in mononuclear phagocytes. We found that, as with TRANCE, the macrophage deactivator TGF-beta(1) strongly promoted TNF-alpha-induced osteoclast-like cell formation from immature bone marrow macrophages. This was abolished by IFN-gamma. However, TRANCE did not share the ability of TNF-alpha to activate NO production or heighten respiratory burst potential by macrophages, or induce inflammation on s.c. injection into mice. This suggests that TGF-beta(1) promotes osteoclast formation not only by inhibiting cytocidal behavior, but also by actively directing TNF-alpha activation of precursors toward osteoclasts. The osteoclast appears to be an equivalent, alternative destiny for precursors to that of cytocidal macrophage, and may represent an activated variant of scavenger macrophage.

Osteoclast lineage commitment of bone marrow precursors through expression of membrane-bound TRANCE.

Osteoclast formation from hemopoietic precursors is induced by TRANCE (also called RANKL, ODF, and OPGL), a membrane-bound ligand expressed by bone marrow stromal cells. Because soluble recombinant TRANCE is a suboptimal osteoclastogenic stimulus, and to eliminate the need for such dependence on stromal cells, membrane-bound TRANCE was expressed in hematopoietic precursors using retroviral gene transfer. Four TRANCE-expressing osteoclast cell lines were established that continuously generate large numbers of multinucleated cells and express tartrate-resistant acid phosphatase and calcitonin receptors. The multinuclear cells are long-lived and either fuse continuously with each other and with mononuclear cells to form enormous syncytia, or separate to form daughter multinuclear cells. When formed on bone, but not on plastic, the majority of multinuclear cells develop actin rings on bone, and resorb bone, suggesting that bone matrix may provide additional signals that facilitate osteoclastic functional maturation. Surprisingly, multinuclear cells originate from fusion of proliferating mononuclear cells that strongly express the mature macrophage markers F4/80 and Fc receptor, which are not expressed by osteoclasts. These results indicate that osteoclasts can be derived from F4/80-positive and Fc receptor-positive cells, and that TRANCE induces osteoclastic differentiation partly by suppressing the macrophage phenotype.

A role for TGFbeta(1) in osteoclast differentiation and survival.

Recently, tumour necrosis factor-related activation-induced cytokine (TRANCE) was shown to be necessary for osteoclast formation. We now report that TGF(beta), a cytokine enriched in bone matrix, is also required. TGF(beta) not only powerfully synergized with TRANCE for induction of osteoclast-like cells (OCL) from bone marrow precursors and monocytes, but OCL formation was abolished by recombinant soluble TGF(beta) receptor II (TGF(beta)sRII). Preincubation in TGF(beta) was as effective as simultaneous incubation with TRANCE. TGF(beta)-preincubation enhanced OCL formation at least partly by preventing the development of resistance to OCL-induction that otherwise occurs when precursors are incubated in M-CSF. OCL formed in TRANCE also showed more rapid apoptosis than OCL in TRANCE plus TGF(beta). Like TGF(beta), incubation on bone matrix prolonged and enhanced the sensitivity of precursors to OCL-induction by TRANCE, and this was reversed by TGF(beta)sRII. Taken together, this data is compelling evidence for a model in which TGF(beta) in matrix or released from bone-lining or other cells maintains and enhances the osteoclast-forming potential of precursors as they migrate towards sites of cell-bound TRANCE. Thus, the specific circumstances necessary for osteoclast formation and survival are TRANCE expression on osteoblastic cells and TGF(beta) in bone.

The role of prostaglandins and nitric oxide in the response of bone to mechanical forces.

We have developed an experimental model whereby bone is exposed to a brief episode of mechanical stimulation, which is followed by bone formation. The earliest response is in osteocytes, which express c-fos and insulin-like growth factor (IGF-1) within 30-60min. Thirty-six to 72h after loading bone matrix gene expression occurs on bone surfaces. The osteogenic response can be suppressed by a single dose of nitric oxide synthase (NOS) or prostaglandin (PG) synthase inhibitors, if these are administered just before mechanical stimulation: similar doses after stimulation have no effect. There is a later phase of indomethacin-sensitivity associated with COX-2 expression in bone at 6h. Thus, mechanically induced osteogenesis involves early expression of c-fos and IGF-1 by osteocytes, which are believed to be the strain-sensitive cells in bone. Both NOS and PG synthase, either in parallel or in sequence, are crucial to the initial transduction of the mechanical stimulus into an osteogenic response.

Solar cycle variability, ozone, and climate

Results from a global climate model including an interactive parameterization of stratospheric chemistry show how upper stratospheric ozone changes may amplify observed, 11-year solar cycle irradiance changes to affect climate. In the model, circulation changes initially induced in the stratosphere subsequently penetrate into the troposphere, demonstrating the importance of the dynamical coupling between the stratosphere and troposphere. The model reproduces many observed 11-year oscillations, including the relatively long record of geopotential height variations; hence, it implies that these oscillations are likely driven, at least in part, by solar variability.

Role of nitric oxide and prostaglandins in mechanically induced bone formation.

We have previously shown that prostaglandins (PG) and nitric oxide (NO) are required in the induction of bone formation by mechanical stimulation. We therefore tested the ability of NO donors, S-nitroso-N-acetyl-D,L-penicillamine (SNAP), and S-nitroso-glutathione (GSNO) to mimic or augment the osteogenic response of bone to a minimal mechanical stimulus. In rats administered vehicle or the vasodilator hydralazine, stimulation of the 8th caudal vertebra increased bone formation. In animals treated with SNAP or GSNO, there was significant potentiation of this osteogenic response. The bone formation rate in nonloaded vertebrae was unaffected by administration of the NO donors. We also found that while inhibition of either PG or NO production at the time of loading caused a partial suppression of c-fos mRNA expression in the loaded vertebrae, administration of indomethacin and NG-monomethyl-L-arginine together markedly suppressed c-fos expression. This suggests that although both PG and NO are required in mechanically induced osteogenesis, they appear to be generated largely independently of each other. Moreover, while exogenous NO potentiates the stimulatory effect of mechanical loading on bone formation, the lack of effect in nonloaded vertebrae suggests that NO is necessary but not sufficient for induction of bone formation.

Vitrectomy with endoscopy for management of retained lens fragments and/or posteriorly dislocated intraocular lens.

PURPOSE: To evaluate the advantages of vitrectomy combined with endoscopy for the management of retained lens fragments and/or posteriorly dislocated intraocular lens (IOL). METHODS: A consecutive series of 30 eyes with these complications treated by this technique was reviewed retrospectively. An endoscopic probe which incorporates a video channel, a fibreoptic light source, and a diode laser was used for visualization. Lens material or the IOL was extracted through the corneal wound in 18 eyes (60%). They were either aspired or grasped or lifted using perfluorocarbon liquids (PFCL), under endoscopic control. In 9 eyes (30%) pars plana phakoemulsification was performed. PFCL was used in 11 eyes (36.6%). In 16 eyes (53.3%) an IOL was sutured in the ciliary sulcus. RESULTS: Final visual acuity was > or = 20/40 in 19 eyes (63.3%), > or = 20/30 in 15 eyes (50%). Intraoperative breaks occurred early in the series in two eyes (in one case from use of the endoprobe, in the other from pars plana phakoemulsification). Poor final acuity was related to proliferative vitreoretinopathy, which developed in both cases with an intraoperative iatrogenic retinal break, senile macular degeneration, myopia and amblyopia, cystoid macular oedema, corneal oedema and high astigmatism. CONCLUSION: We found that endoscopy facilitated the management of these complications of cataract surgery once the peculiar difficulties of the technique (absence of stereoscopy, manipulation of the endoprobe, video monitor control) were mastered. Endoscopy facilitated and shortened localization of lens fragments embedded into the vitreous base for aspiration, grasping and phakoemulsification, enabled detection of small anterior retinal breaks, permitted resection of adhesions between anterior hyaloid, lens capsule and ciliary sulcus and facilitated PFCL manipulations, whatever the status of the anterior segment (corneal edema, myosis, synechiae, presence of IOL).

RoBo-1, a novel member of the urokinase plasminogen activator receptor/CD59/Ly-6/snake toxin family selectively expressed in rat bone and growth plate cartilage.

Using differential display polymerase chain reaction, we cloned a novel cDNA named RoBo-1 from rat tibia. RoBo-1 is abundantly expressed in bone, including the hypertrophic chondrocytes of the growth plate where cartilage is remodeled into bone. RoBo-1 mRNA expression increased in response to two modulators of bone metabolism, estradiol and intermittent mechanical loading, suggesting a role in bone homeostasis. The 1.6-kilobase cDNA encodes a 240-amino acid protein with a cysteine spacing pattern, suggesting that RoBo-1 is a novel member of the urokinase plasminogen activator receptor/CD59/Ly-6/snake toxin family. Furthermore, the C-terminal contains a glycosyl-phosphatidylinositol attachment site, suggesting that it is a cell surface protein similar to other mammalian members of this family. The strongest homology of RoBo-1 is to the snake serum-derived phospholipase A2 inhibitors, which uniquely contain two of the cysteine domains but are secreted proteins. Interestingly, RoBo-1 is likely the first membrane-anchored member of this family containing two cysteine domains. Thus, the tissue specificity, responsiveness to bone protective mediators, along with its relationship to the multifunctional urokinase plasminogen activator receptor/CD59/Ly-6/snake toxin family suggests that RoBo-1 may play a novel role in the growth or remodeling of bone.

Osteocytic expression of mRNA for c-fos and IGF-I: an immediate early gene response to an osteogenic stimulus.

We analyzed the expression, during the osteogenic response of bone to mechanical stimulation, of insulin-like growth factor I (IGF-I), a growth factor implicated in bone formation, and c-fos, a protooncogene in which disordered regulation specifically affects bone. Both genes were strongly expressed in osteocytes of mechanically stimulated but not control bones within 30 min of the osteogenic stimulus. IGF-I mRNA expression increased up to 6 h, was restricted to osteocytes, and was strongly suppressed by indomethacin. Although early IGF-I mRNA expression was resistant to cycloheximide, there was a degree of suppression after 6 h, raising the possibility that IGF-I expression might be prolonged by autocrine mechanisms. c-fos mRNA was increased both in osteocytes and on bone surfaces. At both sites, c-fos expression was transient, prolonged by cycloheximide, and was strongly stimulated even in the presence of indomethacin. Thus osteocytes respond to mechanical stimulation with immediate prolonged expression of IGF-I and immediate transient expression of c-fos, implicating osteocytes in the osteogenic response to mechanical stimulation. Moreover, the different spatial distribution and indomethacin sensitivity of c-fos and IGF-I gene expression suggest that at least two signaling pathways are activated in osteocytes during this process.

The prognostic utility of the Silicone Study Classification System. Silicone Study Report 9. Silicone Study Group.

OBJECTIVE: To evaluate the reproducibility and the prognostic utility of the Retina Society and Silicone Study Classification Systems in eyes after surgery for severe proliferative vitreoretinopathy (PVR). DESIGN: Subgroup analysis of the Silicone Study--a randomized, multicentered, surgical trial. SETTING: Community and university-based ophthalmology clinics. MATERIALS: Three hundred forty eyes with preoperative and intraoperative evaluations using both systems of grading PVR (reproducibility study), and 287 eyes with preoperative and intraoperative evaluations using both systems of grading PVR and with a 24-month follow-up examination (prognosis study). INTERVENTIONS: Vitrectomy for PVR with long-acting perfluoropropane gas or silicone oil as the intraocular tamponade. OUTCOME MEASURES: Retinal reattachment, visual acuity ( > or = 5/200), intraocular pressure, corneal clarity, and the need for reoperation. RESULTS: The reproducibility of the Silicone Study Classification System was 64% (type of contraction), 77% (number of clock hours), 67% (posterior PVR), 88% anterior and posterior PVR), and 94% (anterior, posterior, and subretinal PVR). The reproducibility of the Retina Society Classification System was 99%. Using the Silicone Study Classification System, location of PVR predicted visual acuity (P=.004, chi 2 test for trend) and hypotony (P=.03, chi 2 test for trend). Using the Retina Society Classification System, the grade of PVR predicted only visual acuity (P=.01, chi 2 test for trend). For eyes with anterior and posterior PVR, there was a decreasing trend in successful visual acuity outcome with increasing severity of PVR (from C-3 to D-3, P=.02, chi 2 test for trend). CONCLUSIONS: Although the classification of PVR using the Silicone Study classification System was not reproducible for the type of contraction or for posterior PVR, identification of the anteroposterior extent of the PVR was prognostic of visual acuity and hypotony at 24 months. The joint knowledge of the location of PVR (using the Silicone Study Classification System) and the tightness of the funnel for retinas with 9 to 12 clock hours involved by fixed folds (using the Retina Society Classification System) has prognostic utility for eyes with anterior and posterior PVR.

Posterior vitreoschisis. An echographic finding in proliferative diabetic retinopathy.

PURPOSE: To describe the echographic characteristics of splitting the outer posterior cortical vitreous in patients with proliferative diabetic retinopathy and vitreous hemorrhage. METHODS: The authors retrospectively reviewed the echographic findings in 270 patients who were evaluated at the Doheny Eye Institute between January 1983 to December 1989 for proliferative diabetic retinopathy and vitreous hemorrhage. None of the eyes had undergone pars plana vitrectomy before echographic examination. RESULTS: Forty-five patients (17%) had echographic evidence of splitting of the outer posterior vitreous cortex, a finding the authors have termed posterior vitreoschisis. In all patients, differentiation of the posterior vitreoschisis from a true posterior hyaloid detachment was possible, either on the initial or on serial echographic examination, by the separate detachment of the inner wall of the vitreoschisis cavity and the true posterior hyaloid from the retinal surface. The vitreoschisis cavities often were found to contain unclotted blood. In some eyes, the inner wall of the vitreoschisis cavity was adherent to the apex of the most highly elevated area of traction retinal detachment, suggesting that posterior vitreoschisis may itself result in clinically significant vitreoretinal traction, independent of the presence or extent of true posterior hyaloid separation. CONCLUSIONS: The authors' finding suggest that spontaneous splitting of the outer posterior vitreous cortex may occur in patients with proliferative diabetic retinopathy and vitreous hemorrhage, which may mimic a true posterior cortical vitreous detachment on echographic examination. Preoperative recognition of posterior vitreoschisis may be important in the surgical management of these patients.

Sequential analysis of gene expression after an osteogenic stimulus: c-fos expression is induced in osteocytes.

We have recently developed an experimental model whereby mechanical stimulation induces osteogenesis in the caudal vertebrae of rats. We used this model to assess expression of genes induced by mechanical loading. Bulk preparations of mRNA extracted after loading did not show > 2-fold increases in expression of mRNA for matrix proteins or growth factors in Northern blotting analysis. c-jun was undectable. However, c-fos showed a 4-fold increase in expression within 60 mins of loading, before returning to control levels by 4 hrs. This increase was associated with intense signals in in situ hybridization, not seen in any nonloaded vertebrae, for c-fos over cortical osteocytes: thus osteocytes respond to mechanical loading with c-fos expression so strongly as to be visible even in the bulk RNA preparations. The results represent persuasive evidence for a role for osteocytes, and for c-fos, in the osteogenic response of bone to mechanical stimulation.