Extracellular matrix formation by epithelial cells from human polycystic kidney cysts in culture

Cells from the cysts of patients with autosomal dominant polycystic kidney disease (PKD) were grown in vitro under standard conditions without the aid of collagen-pretreated surfaces, and both the synthesis and composition of the extracellular matrix were investigated. At confluence, PKD cells presented the typical features of epithelial cells, but showed a different collagen composition from fibroblasts. Compared with normal tubular epithelia (NTE), PKD monolayers produced an excess of extracellular matrix, which accounted for 30% of the total incorporation of [³H] proline, although this value was considerably lower (by a factor of 10) in the case of NTE. Immunohistochemical and electrophoretic techniques revealed a complex collagen composition in the extracellular matrix which included [α(III)]₃ and collagen IV. However, part of the collagen components remained unidentified in spite of the fact that they exhibited a typical M_r of α₁(I) and α₂(I) in the presence of urea. Immunoprecipitation with monospecific antibodies and Northern blotting with specific probes failed to recognize α₁(I) and α₂(I), but demonstrated their presence in fibroblasts. Purification and cyanogen bromide digestion demonstrated a strong interhomology in fingerprint peptide composition among the uncharacterized collagens synthesized by PKD cells, thus suggesting a common identity. These observations document a markedly augmented production of extracellular matrix by PKD cultured cells in vitro, and show the presence of collagens which do not share homologies with the major collagen molecules. A better characterization of extracellular matrix composition is central to any comprehension of the cystogenetic mechanisms in vivo.     

Multiple facets of biodiversity are threatened by mining-induced land-use change in the Brazilian Amazon

Aim

Mining is increasingly pressuring areas of critical importance for biodiversity conservation, such as the Brazilian Amazon. Biodiversity data are limited in the tropics, restricting the scope for risks to be appropriately estimated before mineral licensing decisions are made. As the distributions and range sizes of other taxa differ markedly from those of vertebrates—the common proxy for analysis of risk to biodiversity from mining—whether mining threatens lesser-studied taxonomic groups differentially at a regional scale is unclear.

Location

Brazilian Amazon.

Methods

We assess risks to several facets of biodiversity from industrial mining by comparing mining areas (within 70 km of an active mining lease) and areas unaffected by mining, employing species richness, species endemism, phylogenetic diversity and phylogenetic endemism metrics calculated for angiosperms, arthropods and vertebrates.

Results

Mining areas contained higher densities of species occurrence records than the unaffected landscape, and we accounted for this sampling bias in our analyses. None of the four biodiversity metrics differed between mining and nonmining areas for vertebrates. For arthropods, species endemism was greater in mined areas. Mined areas also had greater angiosperm species richness, phylogenetic diversity and phylogenetic endemism, although less species endemism than unmined areas.

Main Conclusions

Unlike for vertebrates, facets of angiosperm and arthropod diversity are relatively higher in areas of mining activity, underscoring the need to consider multiple taxonomic groups and biodiversity facets when assessing risk and evaluating management options for mining threats. Particularly concerning is the proximity of mining to areas supporting deep evolutionary history, which may be impossible to recover or replace. As pressures to expand mining in the Amazon grow, impact assessments with broader taxonomic reach and metric focus will be vital to conserving biodiversity in mining regions.     

Innervation of human omental arteries and veins and vasomotor response to noradrenaline, neuropeptide Y, substance P and vasoactive intestinal peptide

Human omental arteries and veins are supplied with nerve fibers containing noradrenaline (NA) and neuropeptide Y (NPY); these two agents probably co-exist in perivascular sympathetic nerve fibers. Substance P (SP)- or vasoactive intestinal peptide (VIP)-containing fibers could not be detected. In studies on isolated omental vessels NA produced constriction. The results of blockade experiments suggest that human omental arteries are equipped predominantly with alpha 1-adrenoceptors and omental veins with a mixture of alpha 1- and alpha 2-adrenoceptors. NPY at a concentration of 10(-7) M or higher had a weak contractile effect on veins and virtually no effect on arteries. NPY at a concentration of 3 X 10(-8) M shifted the NA concentration response curve to the left in arteries (pD2 = 5.8 for NA versus 6.6. for NA in the presence of NPY; P less than 0.001) but not in veins. Both SP and VIP relaxed arteries precontracted with NA or prostaglandin F2 alpha (PGF2 alpha). The potency of SP as a relaxant agent was similar in arteries and veins; the effect of VIP was elicited at lower concentrations in veins than in arteries.     

Ca2+/calmodulin-dependent protein kinase II is an essential mediator in the coordinated regulation of electrocyte Ca2+-ATPase by calmodulin and protein kinase A

The aim of this study was to investigate (a) whether Ca2+/calmodulin-dependent protein kinase II (CaM kinase II) participates in the regulation of plasma membrane Ca2+-ATPase and (b) its possible cross-talk with other kinase-mediated modulatory pathways of the pump. Using isolated innervated membranes of the electrocytes from Electrophorus electricus L., we found that stimulation of endogenous protein kinase A (PKA) strongly phosphorylated membrane-bound CaM kinase II with simultaneous substantial activation of the Ca2+ pump (approximately 2-fold). The addition of cAMP (5-50 pM), forskolin (10 nM), or cholera toxin (10 or 100 nM) stimulated both CaM kinase II phosphorylation and Ca2+-ATPase activity, whereas these activation processes were cancelled by an inhibitor of the PKA alpha-catalytic subunit. When CaM kinase II was blocked by its specific inhibitor KN-93, the Ca2+-ATPase activity decreased to the levels measured in the absence of calmodulin; the unusually high Ca2+ affinity dropped 2-fold; and the PKA-mediated stimulation of Ca2+-ATPase was no longer seen. Hydroxylamine-resistant phosphorylation of the Ca2+-ATPase strongly increased when the PKA pathway was activated, and this phosphorylation was suppressed by inhibition of CaM kinase II. We conclude that CaM kinase II is an intermediate in a complex regulatory network of the electrocyte Ca2+ pump, which also involves calmodulin and PKA.     

One-step purification of metallothionein extracted from two different sources

We describe a one-step purification of hepatic metallothionein from the Amazon fish Colossoma macropomum injected with cadmium and from the copper-loaded metallothionein from the yeast Saccharomyces cerevisiae, performed by affinity chromatography through metal-chelating columns. Yeast metallothionein was purified from Cu2+-loaded resin and eluted by a continuous EDTA gradient whereas hepatic metallothionein extracted from fishes was purified by Ni2+-loaded resin and eluted by a continuous imidazol gradient. Purified metallothioneins were evaluated by SDS-PAGE and characterized by UV spectra of the apo- and Cd2+-loaded protein. This method allowed high purity and yield as well as rapid one-step extraction of both metal-loaded and apoprotein.     

Bioclimatic conditions in Mexico City – an assessment

 Bioclimatic conditions have been assessed for a large urban area located in the tropical highlands of central Mexico using the indices (in °C) of resultant temperature (RT) and effective temperature (ET). The well-developed heat island effect the city generates, reduces the number of nights categorized as cold (ET between 5 and 15° C) to cool (ET from 15 to 18.5° C). Most days fall in the cold to cool range and during the warm season (April to June) the bioclimate of Mexico City is mostly within the neutral (comfort) range. The effect of the nocturnal (to the west) and daytime (to the east of the town) heat island is noticeable in the central and northern sectors. The daytime heat island located in these regions, albeit small (urban air temperature 2–3° C greater than rural), compared with the nocturnal heat island intensity (9–10° C) still adds energy to the already heated afternoon urban air. ET values in the north and central sectors approach the threshold for comfort (ET of 25° C) during the warm months around noon. It is not surprising to find that as the nocturnal heat island has increased over the years (1921–1985) as the city grew, so has the ET for the central district and indicating the dominating role of temperature in the ET index. Assessment of the diurnal cycle of bioclimatic conditions in downtown Mexico City by means of two empirical indices (effective temperature and thermopreferendum) throughout the years gave similar results to those obtained from the application of Fanger’s predicted mean vote (PMV) model. An attempt has been made to characterize four bioclimatic zones in the capital city. Received: 8 March 1994 / Revised: 15 July 1996 / Accepted: 9 September 1996     

Length–weight relationships of three estuarine species in the Comacchio Lagoon,Po River delta,Italy

The length–weight relationships (LWRs) for three native fish species of the Comacchio Lagoon (northwestern Adriatic Sea, Italy) were determined. The species Zosterisessor ophiocephalus (Pallas, 1814), Pomatoschistus canestrini (Ninni, 1883) and Knipowitschia panizzae (Verga, 1841), were sampled monthly from 2010 to 2014. Both P. canestrini and K. panizzae are listed as protected species in the Habitat Directive and the Berne Convention and there is no information on their LWRs in FishBase.     

Effect of aeration and agitation on extractive fermentation of clavulanic acid by using aqueous two‐phase system

In this work, the effects of agitation and aeration rates on aqueous two‐phase system (ATPS)‐based extractive fermentation of clavulanic acid (CA) by Streptomyces variabilis DAUFPE 3060 were investigated through a 2² full factorial design, where oxygen transfer rate (OTR) and oxygen uptake rate (OUR) were selected as the responses. Aeration rates significantly influenced cell growth, OUR, and CA yield, while OTR was practically the same in all the runs. Under the intermediate agitation (950 rpm) and aeration conditions (3.5 vvm) of the central point runs, it was achieved OTR of 1.617 ± 0.049 mmol L^?1 h^?1, OUR of 0.132 ± 0.030 mmol L^?1 h^?1, maximum CA production of 434 ± 4 mg L^?1, oxygen mass transfer coefficient of 33.40 ± 2.01 s^?1, partition coefficient of 66.5 ± 1.5, CA yield in the top and bottom phases of 75% ± 2% and 19% ± 1%, respectively, mass balance of 95% ± 4% and purification factor of 3.8 ± 0.1. These results not only confirmed the paramount role of O₂ supply, broth composition and operational conditions in CA ATPS‐extractive fermentation, but also demonstrated the possibility of effectively using this technology as a cheap tool to simultaneously produce and recover CA. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:1444–1452, 2016     

Strong spatial structure,Pliocene diversification and cryptic diversity in the Neotropical dry forest spider Sicarius cariri

The Brazilian Caatinga is part of the seasonally dry tropical forests, a vegetation type disjunctly distributed throughout the Neotropics. It has been suggested that during Pleistocene glacial periods, these dry forests had a continuous distribution, so that these climatic shifts may have acted as important driving forces of the Caatinga biota diversification. To address how these events affected the distribution of a dry forest species, we chose Sicarius cariri, a spider endemic to the Caatinga, as a model. We studied the phylogeography of one mitochondrial and one nuclear gene and reconstructed the paleodistribution of the species using modelling algorithms. We found two allopatric and deeply divergent clades within S. cariri, suggesting that this species as currently recognized might consist of more than one independently evolving lineage. Sicarius cariri populations are highly structured, with low haplotype sharing among localities, high fixation index and isolation by distance. Models of paleodistribution, Bayesian reconstructions and coalescent simulations suggest that this species experienced a reduction in its population size during glacial periods, rather than the expansion expected by previous hypotheses on the paleodistribution of dry forest taxa. In addition to that, major splits of intraspecific lineages of S. cariri took place in the Pliocene. Taken together, these results indicate S. cariri has a complex diversification history dating back to the Tertiary, suggesting the history of dry forest taxa may be significantly older than previously thought.