A standard tissue as a control for histochemical and immunohistochemical staining

The variable quality of histochemical and immunohistochemical staining of tissues may be attributed to pre-analytical and analytical variables. Both categories of variables frequently are undefined or inadequately controlled during specimen collection and preparation. Pre-analytical variables may alter the molecular composition of tissues, which results in variable staining; such variations may cause problems when different tissues are used as staining controls. We developed a standard tissue for use as a staining control. Our standard tissue contains five components: 1) nine combined human cell lines mixed with stroma from human spleen; 2) a squamous cancer cell line, A431; 3) fungus; 4) transverse sections of the mosquitofish and 5) normal human spleen. The first three components were embedded in HistoGel^? and all components were processed to paraffin and used to construct a single standard paraffin block. The muscles of mosquitofish and arteries of the spleen are positive controls for eosin staining, while other tissues are useful for assessing hematoxylin staining. The mosquitofish tissues also are excellent controls for the Masson trichrome stain and all mucin-related histochemical stains that we tested. The goblet cells of the intestine and skin stained strongly with Alcian blue, pH 2.5 (AB-2.5), mucicarmine, colloidal iron, periodic acid Schiff (PAS) or PAS-hematoxylin (PASH) and combination stains such as colloidal iron-PASH. Cell lines were not useful for evaluating histochemical stains except for PASH. The splenic stroma was a useful control for AB-2.5; however, eosin and mucin stains stained cell lines poorly, probably due to their rapid growth and associated loss of some differentiated characteristics such as production of mucins. Nevertheless, the cell lines were a critical control for immunohistochemical stains. Immunostaining of specific cell lines was consistent with the presence of markers, e.g., EGFr in DU145 cells. The cell lines expressed a wide range of markers, so they were useful controls for immunohistochemical staining including EGFr, HER2, E-cadherin, cytokeratins, Ki67, PCNA, estrogen receptor, progesterone receptor, CD3, CD20 and CD45, activated (cleaved) caspase 3 and Bcl-2. The cell lines also were a control for the TUNEL stain.     

Rich dynamics of a hepatitis B viral infection model with logistic hepatocyte growth

Chronic hepatitis B virus (HBV) infection is a major cause of human suffering, and a number of mathematical models have examined within-host dynamics of the disease. Most previous HBV infection models have assumed that: (a) hepatocytes regenerate at a constant rate from a source outside the liver; and/or (b) the infection takes place via a mass action process. Assumption (a) contradicts experimental data showing that healthy hepatocytes proliferate at a rate that depends on current liver size relative to some equilibrium mass, while assumption (b) produces a problematic basic reproduction number. Here we replace the constant infusion of healthy hepatocytes with a logistic growth term and the mass action infection term by a standard incidence function; these modifications enrich the dynamics of a well-studied model of HBV pathogenesis. In particular, in addition to disease free and endemic steady states, the system also allows a stable periodic orbit and a steady state at the origin. Since the system is not differentiable at the origin, we use a ratio-dependent transformation to show that there is a region in parameter space where the origin is globally stable. When the basic reproduction number, R ₀, is less than 1, the disease free steady state is stable. When R ₀ > 1 the system can either converge to the chronic steady state, experience sustained oscillations, or approach the origin. We characterize parameter regions for all three situations, identify a Hopf and a homoclinic bifurcation point, and show how they depend on the basic reproduction number and the intrinsic growth rate of hepatocytes.     

EXAMINING HYPOTHESES GENERATED BY FIELD MEASURES OF SEXUAL SELECTION ON MALE LIZARDS,UTA PALMERI

I examined sexual selection in the iguanid lizard Uta palmeri by measuring phenotypic selection in a cohort of males. Relative fitness was estimated by copulation rate from one breeding season, and I analyzed selection on five morphological traits (snout–vent length, mass, jaw length, head width, and head depth) and on male territory quality. Only territory quality and head depth were identified as direct targets of selection in a linear selection gradient analysis. Head depth was suggested to also be subject to quadratic selection. All traits exhibited significant directional selection differentials, suggesting indirect selection also was present because of the correlation of these traits with direct targets of selection. I used these results to generate hypotheses about the mechanisms of selection. For traits not identified as direct targets of selection (snout–vent length, mass, head width, jaw length), I could accept the null hypothesis of no female preference for the analyzed male traits; if these morphological traits were preferred by females in mate choice, they would have been identified as direct targets of selection. Exploring possible functional relationships within the cohort, I found that all five morphological traits contributed to explaining variation in territorial status. And in staged aggressive interactions between males that were similar in snout–vent length and mass, winning was associated only with greater head depth and not with head width or jaw length. Several possible interpretations of these results are presented. This study suggests that differential mating success arising from variation in territory quality gives rise to indirect selection on morphology. The possible mechanisms giving rise to the proposed direct selection on head depth require further study.     

Signals and behavioural responses are not coupled in males: aggression affected by replacement of an evolutionarily lost colour signal

Male Sceloporus virgatus lack the blue abdominal patches which are used during aggressive encounters in other Sceloporus lizards. Herein we report that, despite having lost this signal, males have retained a behavioural response to experimentally restored blue abdominal patches. We tested two adaptive hypotheses: selection acted primarily upon signallers or selection acted upon both signallers and receivers. The first predicts that only the signal is lost and that male interactions should be affected by the restoration of blue patches. The latter predicts that both the signal and behavioural response are lost and the display of the restored blue patches should have no effect on male-male interactions. We compared the behaviour of receivers in paired encounters where one male (signaller) had blue-painted abdominal patches to a set of trials where both males had white-painted abdomens, unmanipulated abdomens or a novel-painted pattern. The receivers of the blue-painted signal were more likely to display submissive behaviour. The receivers in either the unmanipulated, white-painted or novel-painted signal trials were more likely to display neutral behaviour. These results support the hypothesis that receivers have retained a behavioural response and selection has acted primarily on the signaller. We believe this is the first documentation of males responding to an evolutionarily lost signal in conspecific males.     

Interspecific variation of body shape and sexual dimorphism in three coexisting species of the genus <Emphasis Type="Italic">Petrotilapia</Emphasis> (Teleostei: Cichlidae) from Lake Malawi

Differences in color patterns have been the most used feature in describing cichlid species belonging to genus Petrotilapia from Lake Malawi. In this study, we quantified morphological variation in body shape within and among three coexisting Petrotilapia species using landmark-based geometric morphometric methods. Statistic analyses revealed significant body shape differences among species but not between sexes. Post hoc multiple comparisons based on Mahalanobis distances revealed that P. nigra was significantly different from P. genalutea and Petrotilapia sp., whereas the latter two were not significantly different. The splines generated showed that the most pronounced variation was in the head region, in which P. nigra had a relatively longer and deeper head than the other two. The most clear-cut distinction was in gape length; P. genalutea had the longest gape, followed by Petrotilapia sp., whereas P. nigra had the shortest gape. Body depth was shallower in P. nigra than the others. When comparing sexes by their centroid size, ANOVA revealed that males were bigger than females. Therefore, we conclude that color is not the only feature that can distinguish these congeners. We discuss the observed sexual dimorphism in terms of sexual selection and relate morphological variation among species to feeding behavior, which may help explain their coexistence in nature.     

Trehalose Increases Freeze-Thaw Damage in Liposomes Containing Chloroplast Glycolipids

Chloroplast thylakoids contain three classes of glycolipids, monogalactosyldiacylglycerol (MGDG), digalactosyldiacylglycerol (DGDG), and sulfoquinovosyldiacylglycerol (SQDG). We have investigated the stability of large unilamellar vesicles made from egg phosphatidylcholine (EPC) and different chloroplast glycolipids during freezing to -18 degreesC, as a function of the presence of three sugars: glucose, sucrose, or trehalose. Contrary to the situation in thylakoids, where cryoprotection increases from glucose < sucrose < trehalose, liposomes containing 50% DGDG showed the opposite behavior. In fact, carboxyfluorescein leakage increased over the control values (freezing in the absence of sugar) in the presence of trehalose. This effect was not seen in vesicles made from pure EPC, or a mixture of EPC and MGDG, or EPC and SQDG. Liposomes made from mixtures of all three glycolipids, however, showed even more leakage in the presence of trehalose than liposomes containing only DGDG and EPC. Copyright 1998 Academic Press.     

Negative Association Between Conspicuous Visual Display and Chemosensory Behavior in Two Phrynosomatid Lizards

Communication in one sensory modality can influence communication in others. Lizards in many phrynosomatid species use primarily visual but also chemical signals. The striped plateau lizard, Sceloporus virgatus , exhibits evolutionary loss of a male color signal that in many species is used during aggressive postural displays towards conspecific males. These patches are used similarly in Urosaurus , the sister genus to Sceloporus . We compared a species in which a color signal has been lost, S. virgatus , to a species retaining the ancestral character state of blue abdominal display patches, Urosaurus ornatus , the common tree lizard, to test two hypotheses: (i) conspicuous postural displays that reveal the abdominal patch location are used less in the species that has lost the color patches; and (ii) potential chemical signals are used more in the species with the color loss. We analyzed both visual display behavior (push-up, full-show) and chemosensory behavior (tongue flick and nose tap) of male lizards following their introduction to a resident conspecific male in his home terrarium. Resident males performed very low rates of all behaviors, but intruders exhibited sufficient behavior for analysis.
Supporting the first hypothesis, S. virgatus were less likely than U. ornatus to perform full-show, a display that reveals abdominal skin. Male S. virgatus were more likely to perform push-up than U. ornatus , although S. virgatus performed push-up infrequently. Push-up is a postural display that does not specifically reveal the abdominal patch location. Supporting the second hypothesis, S . virgatus were more likely to perform chemosensory behaviors and performed them at a greater rate than did U. ornatus . Work comparing more closely related species is warranted to determine whether a negative association between conspicuous visual displays and chemosensory behavior is a general pattern.     

Losing the trait without losing the signal: Evolutionary shifts in communicative colour signalling

Colour signalling traits are often lost over evolutionary time, perhaps because they increase vulnerability to visual predators or lose relevance in terms of sexual selection. Here, we used spectrometric and phylogenetic comparative analyses to ask whether four independent losses of a sexually selected blue patch are spectrally similar, and whether these losses equate to a decrease in conspicuousness or to loss of a signal. We found that patches were lost in two distinct ways: either increasing reflectance primarily at very long or at very short wavelengths, and that species with additional colour elements (UV, green and pink) may be evolutionary intermediates. In addition, we found that patch spectral profiles of all species were closely aligned with visual receptors in the receiver's retina. We found that loss of the blue patch makes males less conspicuous in terms of chromatic conspicuousness, but more conspicuous in terms of achromatic contrast, and that sexual dimorphism often persists regardless of patch loss. Dorsal surfaces were considerably more cryptic than were ventral surfaces, and species in which male bellies were the most similar in conspicuousness to their dorsal surfaces were also the most sexually dimorphic. These results emphasize the consistent importance of sexual selection and its flexible impact on different signal components through evolutionary time.