Morphological variations among larval-postlarval intermediates produced by eyestalk ablation in the snapping shrimp Alpheus heterochaelis Say

The eyestalk of many crustaceans contains the X-organ, the presumptive site of production and release of many protein and peptide hormones into the hemolymph. Removal of the eyestalk deprives the animal of these hormones and is known to affect many physiological processes in the adult and developing larva. In the snapping shrimp Alpheus heterochaelis Say, eyestalk ablation performed early in larval development has profound effects on morphogenesis, causing the appearance of supernumerary larval stages, accompanied by retardation and even complete arrest of morphogenesis. In this study, we examined the effects on morphogenesis of bilateral eyestalk removal at carefully controlled intervals. We found that the crucial point for this operation-the point at which the animal attains the ability to metamorphose fully-is just before the onset of ecdysis to the third instar. Additionally, the pattern of development and morphogenesis among body segments follows a discernible double gradient pattern along the anterior-posterior axis in which the extremities of the animal attain the potential for morphogenetic advance prior to the central thorax. This pattern of morphogenesis, punctuated by ecdysis, is a continuous rather than a stepwise or compartmentalized phenomenon.     

Geographic differences in species boundaries among members of the Montastraea annularis complex based on molecular and morphological markers

The three members of the Montastraea annularis complex (M. annularis, M. franksi, and M. faveolata) are dominant reef builders in the western Atlantic whose species status has been controversial for over a decade. Although differences in colony morphology and reproductive characteristics exist, interspecific fertilizations are possible in the laboratory and genetic differentiation is slight. Here we compare the three taxa genetically and morphologically in Panama and the Bahamas, widely separated locations spanning most of their geographic ranges. In Panama, analyses of three AFLP loci, a noncoding region of the mitochondrial genome, and ITS sequences reveal that M. faveolata is strongly differentiated genetically. Discriminant function analysis also indicates no overlap with the other two species in the fine structure of the corallites that comprise the colony. Genetic analyses of larvae from interspecific crosses between M. faveolata and the other two taxa confirmed the hybrid status of the larvae, but no examples of the most probable F1 genotype were observed in the field. Although M. annularis and M. franksi were more similar, they also exhibited strong frequency differences at two AFLP loci and in the mitochondrial noncoding region, as well as distinct corallite structure. In the Bahamas, in contrast, the three taxa exhibited overlapping morphologies. Montastraeafranksi and M. annularis were indistinguishable genetically, and M. faveolata was distinct at fewer genetic loci. Once again, however, the most probable F1 genotype involving M. faveolata was not observed. Geographic differences between Panama and the Bahamas explain why past studies have come to different conclusions concerning the status of the three species. In general, the genetic and morphological data suggest a north to south hybridization gradient, with evidence for introgression strongest in the north. However, reproductive data show no such trend, with intrinsic barriers to gene flow comparable or stronger in the north.     

Hypervariable genes--experimental error or hidden dynamics

In a homogeneous group of samples, not all genes of high variability stem from experimental errors in microarray experiments. These expression variations can be attributed to many factors including natural biological oscillations or metabolic processes. The behavior of these genes can tease out important clues about naturally occurring dynamic processes in the organism or experimental system under study. We developed a statistical procedure for the selection of genes with high variability denoted hypervariable (HV) genes. After the exclusion of low expressed genes and a stabilizing log-transformation, the majority of genes have comparable residual variability. Based on an F-test, HV genes are selected as having a statistically significant difference from the majority of variability stabilized genes measured by the 'reference group'. A novel F-test clustering technique, further noted as 'F-means clustering', groups HV genes with similar variability patterns, presumably from their participation in a common dynamic biological process. F-means clustering establishes, for the first time, groups of co-expressed HV genes and is illustrated with microarray data from patients with juvenile rheumatoid arthritis and healthy controls.     

Microarray Data Analysis Toolbox (MDAT): for normalization, adjustment and analysis of gene expression data

SUMMARY: We introduce a novel Matlab toolbox for microarray data analysis. This toolbox uses normalization based upon a normally distributed background and differential gene expression based on five statistical measures. The objects in this toolbox are open source and can be implemented to suit your application. AVAILABILITY: MDAT v1.0 is a Matlab toolbox and requires Matlab to run. MDAT is freely available at http://microarray.omrf.org/publications/2004/knowlton/MDAT.zip.     

Correlation between thyroid hormone status and hepatic hyperplasia and hypertrophy caused by the peroxisome proliferator-activated receptor alpha agonist Wy-14,643

BACKGROUND: The metabolic inhibitor rotenone inhibits hepatocellular proliferation and the incidence of liver cancer resulting from exposure to the PPARalpha agonist Wy-14,643, via unknown mechanisms. Since the absence of thyroid hormones diminishes hepatomegaly, an early biomarker for the hepatocarcinogenicity induced by PPARalpha agonists, this study was undertaken to investigate whether rotenone might interference with the ability of Wy-14,643 to alter the animal thyroid status. METHODS: Male B6C3F1 mice were given Wy-14,643 (100 ppm), rotenone (600 ppm) or a mixture of both, in the feed for 7 days. Bromodeoxyuridine (BrDU), marker of cell replication, was delivered through subcutaneously implanted osmotic mini-pumps. At the end of the experiment, sera were collected and corticosterone and thyroid hormone levels were measured by solid-phase radioimmunoassay kits. In addition, liver tissue samples were stained immunohistochemically for BrDU to determine percentages of labeled cells. Further, cell surface area was determined from images generated by a Zeiss Axioplan microscope equipped with a plan Neofluar x40 0.75 na objective. Tracings of individual hepatocyte perimeters were then analyzed and cell-surface areas were calculated using MicroMeasure FL-4000. RESULTS: Wy-14,643 caused a significant increase in liver weights, hepatocyte BrDU labeling index (LI), and hepatocyte surface area. In animals which received both Wy-14,643 and rotenone simultaneously, all of these effects were significantly less pronounced compared with mice that received Wy-14,643 alone. Rotenone alone decreased liver weights, LI and surface area. The Free Thyroid Index (FTI), which provides an accurate reflection of the animal's thyroid status, was 5.0 +/- 0.3 in control mice. In animals exposed to rotenone, these values decreased to 2.0 +/- 0.9, but in animals which received Wy-14,643, levels increased significantly to 7.7 +/- 0.9. FTI values decreased to 3.4 +/- 0.8 in mice receiving both rotenone and Wy-14,643. CONCLUSION: A strong correlation was observed between the animal thyroid status and both, hepatocyte proliferation (r2 = 0.62), and hepatocyte surface area (r2 = 0.83). These results support the hypothesis that the thyroid status of the animal plays a role in PPARalpha-induced hepatocellular proliferation and liver cell enlargement. Both these events are known to contribute to the expression of liver cancer in response to the activation of PPARalpha.     

Inflammation and ischemia: macrophages activated by fibronectin fragments enhance the survival of injured cardiac myocytes

Proteolytic enzymes, released early in the course of an inflammatory response, hydrolyze fibronectin, producing fragments of the parent molecule that alter monocyte phenotype and migratory behavior. Here we test the hypothesis that macrophages, stimulated by the dominant 110-120 kd fibronectin fragments (FNf), as are found in lymphatic fluid draining sites of cardiac ischemia-reperfusion injury, produce factors that promote the survival of injured parenchymal cells. Rat splenic macrophages stimulated in vitro with purified FNf produced soluble factors that protected hypoxic rat cardiac myocytes from death by apoptosis. Addition of blocking antibodies specific for tumor necrosis factor-alpha(TNF-alpha), fibroblast growth factor-1 (FGF-1), insulin-like growth factor I (IGF-I), and leukemia inhibitory factor (LIF) partly reduced the protection against apoptosis provided to hypoxic cardiac myocytes by cell-free culture supernatants from FNf-stimulated macrophages. Complete blockade of this protection was achieved by a combination of antibodies specific for FGF-1, IGF-I, and LIF. Stimulation of human monocyte-derived macrophages in vitro with FNf significantly increased their output of TNF-alpha, FGF-1, IGF-I, and LIF. These results suggest that tissue degradation products, released in the early hours of an inflammatory response, stimulate tissue-infiltrating macrophages to protect injured but still viable parenchymal cells from death by apoptosis.     

Expression of heat shock proteins in turtle and mammal hearts: relationship to anoxia tolerance

Heat shock proteins (HSPs) may play a cardioprotective role during hypoxia or ischemia. We hypothesized that cardiac tissue from hypoxia-tolerant animals might have high levels of specific HSPs. We measured myocardial HSP60 and HSP72/73 in painted and softshell turtles during normoxia and anoxia (12 h) and after recovery (12 or 24 h). We also measured myocardial HSPs in normoxic rats and rabbits. During normoxia, hearts from the most highly anoxia-tolerant species, the painted turtle, expressed the highest levels of HSP60 (22.6+/-2.0 mg/g total protein) followed by softshells (11.5+/-0.8 mg/g), rabbits (6.8+/-0.9 mg/g), and rats (4.5+/-0.5 mg/g). HSP72/73 levels, however, were not significantly different. HSP60 levels in hearts from both painted and softshell turtles did not deviate significantly from control values after either 12 h of anoxia or 12 or 24 h of recovery. The pattern of changes observed in HSP72/73 was quite different in the two turtle species. In painted turtles anoxia induced a significant increase in myocardial HSP72/73 (from 2.8+/-0.1 mg/g normoxic to 3.9+/-0.2 mg/g anoxic, P<0.05). By 12 h of recovery, HSP72/73 had returned to control levels (2.7+/-0.1 mg/g) and remained there through 24 h (2.6+/-0.2 mg/g). In softshell turtles, HSP72/73 decreased significantly after 12 h of anoxia (from 2.4+/-0.4 mg/g normoxic to 1.3+/-0.2 mg/g anoxic, P<0.05). HSP72/73 levels were still slightly below control after 12 h of recovery (2.1+/-0.1 mg/g) and then rose to significantly above control after 24 h of recovery (4.1+/-0.7 mg/g, P<0.05). We also conclude that anoxia-tolerant and anoxia-sensitive turtles exhibit different patterns of myocardial HSP changes during anoxia and recovery. Whether these changes correlate with their relative degrees of anoxia tolerance remains to be determined.     

OCCURRENCE, COMPOSITION, AND POTENTIAL FUNCTIONS OF NORTH ATLANTIC RIGHT WHALE (EUBALAENA GLACIALIS) SURFACE ACTIVE GROUPS

Marine mammal social interactions are poorly understood primarily because of the difficulty of observing these behaviors in the wild. Observations of copulations in North Atlantic right whale surface active groups (SAGs) have led to the hypothesis that the function of these groups is for conception. The occurrence and composition of SAGs sighted from 1992 to 2001 were investigated on the basis of age, sex, and reproductive status of individuals. A total of 918 SAG events were documented. SAGs were observed in all habitat areas and in all months in which right whales were sighted. Group compositions revealed both potentially conceptive groups and nonconceptive groups. Of right whales whose age was known, 93% were first sighted in SAGs as juveniles, and more than half of all observed groups contained at least one juvenile whale. The group composition and timing of occurrence of SAGs do not support the hypothesis that all groups serve a purely conceptive function. Their functional role is likely much broader. Other potential functions include play, mating practice, or maintenance of social bonds.     

Ants Do Not Traverse the Silk of Adult Female <Emphasis Type="Italic">Nephila clavipes</Emphasis> (Linnaeus) Webs

Many organisms use chemicals to deter enemies. Some spiders can modify the composition of their silk to deter predators from climbing onto their webs. The Malaysian golden orb-weaver Nephila antipodiana (Walckenaer) produces silk containing an alkaloid (2-pyrrolidinone) that functions as a defense against ant invasion—ants avoid silk containing this chemical. In the present study, we test the generality of ants’ silk avoidance behavior in the field. We introduced three ant species to the orb webs of Nephila clavipes (Linnaeus) in the tropical rainforest of La Selva, Costa Rica. We found that predatory army ants (Eciton burchellii Westwood) as well as non-predatory leaf-cutting ants (Atta cephalotes Linnaeus and Acromyrmex volcanus Wheeler) avoided adult N. clavipes silk, suggesting that an additional species within genus Nephila may possess ant-deterring silk. Our field assay also suggests that silk avoidance behavior is found in multiple ant species.     

Short-term hypoxia/reoxygenation activates the angiogenic pathway in rat caudate putamen

In response to hypoxia, tissues have to implement numerous mechanisms to enhance oxygen delivery, including the activation of angiogenesis. This work investigates the angiogenic response of the hypoxic caudate putamen after several recovery times. Adult Wistar rats were submitted to acute hypoxia and analysed after 0 h, 24 h and 5 days of reoxygenation. Expression of hypoxia-inducible factor-1 alfa (HIF-1α) and angiogenesis-related genes including vascular endothelial growth factor (VEGF), adrenomedullin (ADM) and transforming growth factor-beta 1 (TGF-β1) was determined by both RT-PCR and ELISA. For vessel labelling, lectin location and expression were analysed using histochemical and image processing techniques (fractal dimension). Expression of Hif-1α, Vegf, Adm and Tgf-β1 mRNA rose immediately after hypoxia and this increase persisted in some cases after 5 days post-hypoxia. While VEGF and TGF-β1 protein levels increased parallel to mRNA expression, ADM remained unaltered. The quantification of the striatal vessel network showed a significant augmentation at 24 h of reoxygenation. These results reveal that not only short-term hypoxia, but also the subsequent reoxygenation period, up-regulate the angiogenic pathway in the rat caudate putamen as a neuroprotective mechanism to hypoxia that seeks to maintain a proper blood supply to the hypoxic tissue, thereby minimizing the adverse effects of oxygen deprivation.