Variation in the Distribution of Trace Elements in Renal Cell Carcinoma

The development of cancer is a complex, multistage process during which a normal cell undergoes genetic changes that result in phenotypic alterations and in the acquisition of the ability to invade other sites. Inductively coupled plasma optical emission spectroscopy was used to estimate the contents of Al, Ca, Cd, Cr, Cu, Fe, K, Mg, Mn, Na, P, Pb, and Zn in healthy kidney and renal cell carcinoma (RCC), and significant differences were found for all elements. Along with the progression of the malignant disease, a progressive decrease of Cd and K was observed. In fact, for Cd, the concentration in stage T4 was 263.9 times lower than in stage T1, and for K, the concentration in stage T4 was 1.73 times lower than in stage T1. Progressive accumulation was detected for P, Pb, and Zn in stage T4. For P, the concentration in stage T4 was 11.1 times higher than in stage T1; for Pb, the concentration in stage T4 was 232.7 times higher than in T1; and for Zn, the concentration in T4 was 8.452 times higher than in T1. This study highlights the marked differences in the concentrations of selected trace metals in different malignant tumor stages. These findings indicate that some trace metals may play important roles in the pathogenesis of RCC.     

Extraction and Inhibition of Enzymatic Activity of Botulinum Neurotoxins/A1, /A2, and /A3 by a Panel of Monoclonal Anti-BoNT/A Antibodies

Botulinum neurotoxins (BoNTs) are extremely potent toxins that are capable of causing death or respiratory failure leading to long-term intensive care. Treatment includes serotype-specific antitoxins, which must be administered early in the course of the intoxication. Rapidly determining human exposure to BoNT is an important public health goal. In previous work, our laboratory focused on developing Endopep-MS, a mass spectrometry-based endopeptidase method for detecting and differentiating BoNT/A–G serotypes in buffer and BoNT/A, /B, /E, and /F in clinical samples. We have previously reported the effectiveness of antibody-capture to purify and concentrate BoNTs from complex matrices, such as clinical samples. Because some antibodies inhibit or neutralize the activity of BoNT, the choice of antibody with which to extract the toxin is critical. In this work, we evaluated a panel of 16 anti-BoNT/A monoclonal antibodies (mAbs) for their ability to inhibit the in vitro activity of BoNT/A1, /A2, and /A3 complex as well as the recombinant LC of A1. We also evaluated the same antibody panel for the ability to extract BoNT/A1, /A2, and /A3. Among the mAbs, there were significant differences in extraction efficiency, ability to extract BoNT/A subtypes, and inhibitory effect on BoNT catalytic activity. The mAbs binding the C-terminal portion of the BoNT/A heavy chain had optimal properties for use in the Endopep-MS assay.     

Disseminated mycobacteriosis affecting a prosthetic aortic valve: first case of Mycobacterium peregrinum type III reported in Colombia

Rapidly growing mycobacteria are non-tuberculous mycobacteria amply present in the environment. Although they are not usually pathogenic for humans, they are opportunistic in that they can cause disease in people with disadvantageous conditions or who are immunocompromised. Mycobacterium peregrinum, an opportunistic, rapidly growing mycobacteria, belongs to the M. fortuitum group and has been reported as responsible for human cases of mycobacteriosis. A case of M. peregrinum type III is herein reported as the first in Colombia. It presented as a disseminated disease involving a prosthetic aortic valve (endocarditis) in a seventeen-year-old girl with a well-established diagnosis of prosthetic aortic valve endocarditis who was referred for a surgical replacement. Due to a congenital heart disease (subaortic stenosis with valve insufficiency), she had two previous aortic valve implantation surgeries. One year after the second implantation, the patient presented with respiratory symptoms and weight lost indicative of lung tuberculosis. A chest X-ray did not show parenchymal compromise but several Ziehl-Neelsen stains were positive. An echocardiography showed a vegetation on the prosthetic aortic valve. In blood and sputum samples, M. peregrinum type III was identified through culture, biochemical tests and hsp65 gene molecular analysis (PRA). The patient underwent a valve replacement and received a multidrug antimycobacterial treatment. Progressive recovery ensued and further samples from respiratory tract and blood were negative for mycobacteria.     

Genetic characterization of indigenous peoples from Oaxaca, Mexico, and its relation to linguistic and geographic isolation

We used 15 short tandem repeat (STR) loci (D8S1179, D21S11, D7S820, CSF1PO, D3S1358, TH01, D13S317, D16S539, D2S1338, D19S433, VWA, TPOX, D18S51, D5S818, and FGA) to genetically characterize 361 individuals from 11 indigenous populations (Amuzgo, Chinanteco, Chontal, Huave, Mazateco, Mixe, Mixteco, Triqui, Zapoteco del Istmo, Zapoteco del Valle, and Zoque) from Oaxaca, Mexico. We also used previously published data from other Mexican peoples (Maya, Chol, Tepehua, Otomí, and Mestizos from northern and central Mexico) to delineate genetic relations, for a total of 541 individuals. Average heterozygosity (H) was lower in most populations from Oaxaca (range 0.687 in Zoque to 0.756 in Chontal) than values observed in Mestizo populations from Mexico (0.758 and 0.793 in central and northern Mestizo, respectively) but higher than values observed in other Amerindian populations from South America; the same relation was true for the number of alleles (n(a) ). We tested (using the software Structure) whether major geographic or linguistic barriers to gene flow existed among the populations of Oaxaca and found that the populations appeared to constitute one or two genetic groups, suggesting that neither geographic location nor linguistics had an effect on the genetic structure of these culturally and linguistically highly diverse indigenous peoples. Moreover, we found a low but statistically significant between-population differentiation. In addition, the genetic structure of Oaxacan populations did not fit an isolation-by-distance model. Finally, using AMOVA and a Bayesian clustering approach, we did not detect significant geographic or linguistic barriers to gene flow within Oaxaca. These results suggest that the indigenous communities of Oaxaca, although culturally isolated, can be genetically defined as a large, nearly panmictic population in which migration could be a more important population mechanism than genetic drift. Finally, compared with outgroups in Mexico (both indigenous peoples and Mestizos), three groups were apparent. Among them, only the Otomí population from Hidalgo has a different culture and language.     

Plant Host Finding by Parasitic Plants: A New Perspective on Plant to Plant Communication

Plants release airborne chemicals that can convey ecologically relevant information to other organisms. These plant volatiles are known to mediate a large array of, often complex, interactions between plants and insects. It has been suggested that plant volatiles may have similar importance in mediating interactions among plant species, but there are few well-documented examples of plant-to-plant communication via volatiles, and the ecological significance of such interactions has been much debated. To date, nearly all studies of volatile-mediated interactions among plant species have focused on the reception of herbivore-induced volatiles by neighboring plants. We recently documented volatile effects in another system, demonstrating that the parasitic plant Cuscuta pentagona uses volatile cues to locate its hosts. This finding may broaden the discussion regarding plant-to-plant communication, and suggests that new classes of volatile-meditated interactions among plant species await discovery.Key Words: chemical communication, Cuscuta pentagona, host fiding, host selection, plant-plant communication, plant volatiles, parasitic plantsFor nearly 25 years, the ecological importance of plant-to-plant communication through volatiles has remained an open and much debated question. Plants exchange gases with the atmosphere and, in so doing, release plumes of volatile chemicals that can convey ecologically important information to other organisms. The potential ecological significance of these volatile cues is demonstrated by the large and growing array of interactions between plants and arthropods known to be mediated by plant volatiles. Volatiles serve as foraging cues both for insects that are beneficial to plants, such as pollinators,¹ and those that are harmful such as herbivores.^2,3 Because the volatile blends released by plants exhibit variation in response to environmental stimuli, volatiles can convey detailed information about the status of the emitting plant. Predatory and parasitic insects that feed on herbivorous insects respond preferentially to plant volatiles that are induced by insect feeding,⁴ while female herbivores use such cues to avoid laying their eggs on already-infested plants.^3,5 Moreover, the volatile blends released in response to herbivory can differ between individual herbivore species, providing highly specific cues to specialist parasitoids.⁶ Thus, plant volatiles are known to mediate complex interactions among plants and insects across multiple trophic levels.It has long been speculated that plant volatiles might have similar significance for interactions among plant species, yet there are few well-documented examples of communication between plants by way of volatile signals. Essentially all previous work on plant-to-plant communication has focused on the reception of herbivore-induced volatile signals by neighboring plants, which may use them as early warning signals to initiate their own direct and indirect defense responses. The first studies claiming to document such effects were published almost 25 years ago.^7,8 But issues with the experimental design of these early experiments and the availability of alternative explanations for their results led many ecologists to disregard the phenomenon.^9–11 Later, a number of studies demonstrated that direct and indirect plant defenses could be elicited by exposure to certain induced plant volatiles.^12–15 But many of these effects were demonstrated in airtight chambers with volatile concentrations far higher than those likely experienced in natural settings, again raising doubts about the ecological significance of plant-plant communication.^16–18 Still more recently, some researchers have provided evidence that more realistic volatile concentrations likely induce priming of the defenses of receiving plants, rather than the initiation of full scale responses,¹⁵ while others have documented volatile effects under natural conditions.^19–21 Thus, despite continuing caution about the interpretation of experiments in this area,^17,18 there is mounting evidence that plant herbivore-induced volatiles can serve as early warning signals to neighboring plants.We recently documented an entirely new class of volatile mediated interactions among plants: the role of plant volatiles in host location by parasitic plants that attach to above ground shoots of other plants. Plant parasites are important components of natural and agricultural ecosystems and play important roles in determining community structure and dynamics.^22,23 We are exploring the mechanisms of host-location and other interactions between parasitic plants in the genus Cuscuta (dodder) and their host plants. Dodder vines germinate from seeds containing limited energy reserves and, as the parasites have no roots and little photosynthetic ability, must quickly locate and attach to suitable hosts in order to survive (Fig. 1). Thus, there is presumably significant selection pressure for dodder vines to employ efficient strategies for host location, and host plant volatiles may be expected to provide relevant directional cues. Dodder seedlings exhibit a rotational growth habit (circumnutation) following germination and previous researchers have suggested that host-finding might involve random growth²⁴ or the exploitation of light cues.²⁵Open in a separate windowFigure 1Seedling of Cuscuta pentagona (A) foraging toward a 20-day-old tomato plant, (B) attaching to and beginning to grow from stems of tomato seedlings and (C) close up of C. pentagona attachment.Using a very simple experimental design, we explored the possibility that host-plant volatiles might mediate host-location by seedlings of C. pentagona. We placed a germinated seedling in a vial of water located at the center of a dry filter paper disk. A host plant (a 20-day old tomato seedling) was placed near the edge of the disk and the dodder seedling was allowed to forage for four days. By the end of the experiment the seedling would lay horizontally on the disk and we traced its position on the filter paper in order to assess the directionality of growth relative to the host plant. This experiment was replicated 30 times and our results clearly indicated directional growth toward the tomato plant (80% of the tested seedlings grew into the disk half nearest the host) demonstrating that C. pentagona seedlings were perceiving some host-derived cue.We did not observe directed growth when we tested dodder seedling response to alternative targets including pots of moist soil, artificial plants, and vials of colored water intended to mimic possible light cues. In order to confirm a role for plant volatiles in host location by C. pentagona, we tested seedling response to host plant volatiles extracted from filtered air in a volatile collection system and then released from rubber septa in the absence of any other host-derived cues. Here we observed a directed growth response similar to that exhibited toward an intact tomato seedling, confirming that host plant volatiles do provide a cue used for host location by C. pentagona. In subsequent experiments we found directed growth toward impatiens and alfalfa plants, which are attractive hosts for C. pentagona and also toward wheat plants which are poor hosts, suggesting that the host-location mechanisms operate over a wide range of host species.Since discriminating between more and less desirable host species is likely to be important in natural settings, we next explored whether dodder seedlings could distinguish volatile signals from host and nonhost plants. Cuscuta pentagona seedlings exhibited directional growth toward tomato plants in preference to wheat plants and also to extracted volatiles from tomato in preference to those from wheat, demonstrating an ability to distinguish and choose among volatiles from more and less preferred hosts.When we tested seedling responses to individual compounds from the wheat and tomato blends, we found that three compounds from tomato, α-pinene, β-myrcene, and β-phellandrene elicited directed growth. β-myrcene was also present in the wheat blend. Unexpectedly, we also found that one compound present in the wheat blend, (Z)-3-hexenyl acetate, was repellent, providing a plausible explanation for the lower attractiveness of the wheat blend. It is interesting to note that (Z)-3-hexenyl acetate is also released by tomato in response to feeding by herbivores, and we have some data suggesting that C. pentagona seedlings may find tomato seedlings infested by Heliothis virescens caterpillars less attractive than un-attacked plants (unpublished data).The discovery that some parasitic plants exploit host plant volatiles for host location provides a new perspective on volatile mediated interactions among plant species, demonstrating that plant volatiles play a role in mediating ecologically significant interactions in at least one system other than the transfer of herbivore-induced warning signals. We think it is quite likely that plant volatiles will be found to play a role in host location by other parasitic plants and perhaps even by vining plants generally. Moreover, we think it is more likely than not that more classes of volatile mediated interactions among plants remain to be discovered given the potential availability of volatile cues and the fitness benefits to be derived by plants using such cues to gather information about the identity and condition of their neighbors.     

Testing metabolic ecology theory for allometric scaling of tree size, growth and mortality in tropical forests

The theory of metabolic ecology predicts specific relationships among tree stem diameter, biomass, height, growth and mortality. As demographic rates are important to estimates of carbon fluxes in forests, this theory might offer important insights into the global carbon budget, and deserves careful assessment. We assembled data from 10 old-growth tropical forests encompassing censuses of 367 ha and > 1.7 million trees to test the theory's predictions. We also developed a set of alternative predictions that retained some assumptions of metabolic ecology while also considering how availability of a key limiting resource, light, changes with tree size. Our results show that there are no universal scaling relationships of growth or mortality with size among trees in tropical forests. Observed patterns were consistent with our alternative model in the one site where we had the data necessary to evaluate it, and were inconsistent with the predictions of metabolic ecology in all forests.     

Molecular biology in the diagnosis of deep-seated candidiasis in the critically ill non-neutropenic patient

Candida infections are an important cause of morbidity and mortality in critically ill patients. Rapid detection of the yeast in blood and other tissues by molecular biology methods has been the goal of some recent studies. An analysis of the sensitivity and specificity of these methods assayed in clinical specimens from critically ill and other patients is carried out. PCR amplification of ribosomal genes and their internal spacers showed a higher sensitivity than culture based methods. A standardization of most of the methodological steps in molecular methods is needed. Real time PCR with fluorescent probes seems to be the most interesting proposal. It has the advantage of the possible quantification of fungal presence in tissues and minimizes the samples' contamination risk.     

Structural and thermodynamic characterization of Pal, a phage natural chimeric lysin active against pneumococci

Pal amidase, encoded by pneumococcal bacteriophage Dp-1, represents one step beyond in the modular evolution of pneumococcal murein hydrolases. It exhibits the choline-binding module attaching pneumococcal lysins to the cell wall, but the catalytic module is different from those present in the amidases coded by the host or other pneumococcal phages. Pal is also an effective antimicrobial agent against Streptococcus pneumoniae that may constitute an alternative to antibiotic prophylaxis. The structural implications of Pal singular structure and their effect on the choline-amidase interactions have been examined by means of several techniques. Pal stability is maximum around pH 8.0 (Tm approximately 50.2 degrees C; DeltaHt = 183 +/- 4 kcal mol(-1)), and its constituting modules fold as two tight interacting cooperative units whose denaturation merges into a single process in the free amidase but may proceed as two well resolved events in the choline-bound state. Choline titration curves reflect low energy ligand-protein interactions and are compatible with two sets of sites. Choline binding strongly stabilizes the cell wall binding module, and the conformational stabilization is transmitted to the catalytic region. Moreover, the high proportion of aggregates formed by the unbound amidase together with choline preferential interaction with Pal dimers suggest the existence of marginally stable regions that would become stabilized through choline-protein interactions without significantly modifying Pal secondary structure. This structural rearrangement may underlie in vitro "conversion" of Pal from the low to the full activity form triggered by choline. The Pal catalytic module secondary structure could denote folding conservation within pneumococcal lytic amidases, but the number of functional choline binding sites is reduced (2-3 sites per monomer) when compared with pneumococcal LytA amidase (4-5 sites per monomer) and displays different intermodular interactions.     

Ceramide pathways modulate ethanol-induced cell death in astrocytes

We showed previously that alcohol exposure during in vivo brain development induced astroglial damage and caused cell death. Because ceramide modulates a number of biochemical and cellular responses to stress, including apoptosis, we now investigate whether ethanol-induced cell death in astrocytes is mediated by ceramide signalling pathways triggering apoptosis. Here we show that both ethanol and ceramide are able to induce apoptotic death in cultured astrocytes, in a dose-dependent manner, and that C2-ceramide addition potentiates the apoptotic effects of ethanol. Cell death induced by ethanol is associated with stimulation of neutral and acidic sphingomyelinase (SMase) and ceramide generation, as well as with activation of stress-related kinases, c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein kinase (p38) and extracellular signal-regulated kinase (ERK) pathways. We also provide evidence for the participation of JNK and p38 in ethanol-induced cell death, because pharmacological inhibitors of these kinases largely prevent the apoptosis induced by ethanol or by ethanol and C2-ceramide. Furthermore, we show that ethanol-induced ERK activation triggers the stimulation of cyclo-oxygenase-2 (COX-2) and the release of prostaglandin E2, and that blockade of the mitogen-activated protein kinase kinase (MEK)/ERK pathway by PD98059 abolishes the up-regulation of COX-2 induced by ethanol plus ceramide, and decreases the ethanol-induced apoptosis. These results strongly suggest that ethanol is able to stimulate the SMase-ceramide pathway, leading to the activation of signalling pathways implicated in cell death. These findings provide an insight into the mechanisms involved in ethanol-induced astroglial cell death during brain development.