Protective Geometry and Reproductive Anatomy as Candidate Determinants of Clutch Size Variation in Pentatomid Bugs.

AbstractMany animals lay their eggs in clusters. Eggs on the periphery of clusters can be at higher risk of mortality. We asked whether the most commonly occurring clutch sizes in pentatomid bugs could result from geometrical arrangements that maximize the proportion of eggs in the cluster's interior. Although the most common clutch sizes do not correspond with geometric optimality, stink bugs do tend to lay clusters of eggs in shapes that protect increasing proportions of their offspring as clutch sizes increase. We also considered whether ovariole number, an aspect of reproductive anatomy that may be a fixed trait across many pentatomids, could explain observed distributions of clutch sizes. The most common clutch sizes across many species correspond with multiples of ovariole number. However, there are species with the same number of ovarioles that lay clutches of widely varying size, among which multiples of ovariole number are not overrepresented. In pentatomid bugs, reproductive anatomy appears to be more important than egg mass geometry in determining clutch size uniformity. In addition, our analysis demonstrates that groups of animals with little variation in ovariole number may nonetheless lay a broad range of clutch shapes and sizes.

Ambulatory function in motor incomplete spinal cord injury: a magnetic resonance imaging study of spinal cord edema and lower extremity muscle morphometry.

STUDY DESIGN: This research utilized a cross-sectional design. OBJECTIVES: Spinal cord edema length has been measured with T2-weighted sagittal MRI to predict motor recovery following spinal cord injury. The purpose of our study was to establish the correlational value of axial spinal cord edema using T2-weighted MRI. We hypothesized a direct relationship between the size of damage on axial MRI and walking ability, motor function and distal muscle changes seen in motor incomplete spinal cord injury (iSCI). SETTING: University-based laboratory in Chicago, IL, USA. METHODS: Fourteen participants with iSCI took part in the study. Spinal cord axial damage ratios were assessed using axial T2-weighted MRI. Walking ability was investigated using the 6-min walk test and daily stride counts. Maximum plantarflexion torque was quantified using isometric dynomometry. Muscle fat infiltration (MFI) and relative muscle cross-sectional area (rmCSA) were quantified using fat/water separation magnetic resonance imaging. RESULTS: Damage ratios were negatively correlated with distance walked in 6 min, average daily strides and maximum plantarflexion torque, and a negative linear trend was found between damage ratios and lower leg rmCSA. While damage ratios were not significantly correlated with MFI, we found significantly higher MFI in the wheelchair user participant group compared to community walkers. CONCLUSIONS: Damage ratios may be useful in prognosis of motor recovery in spinal cord injury. The results warrant a large multi-site research study to investigate the value of high-resolution axial T2-weighted imaging to predict walking recovery following motor incomplete spinal cord injury.

Fanconi anemia signaling and Mus81 cooperate to safeguard development and crosslink repair.

Individuals with Fanconi anemia (FA) are susceptible to bone marrow failure, congenital abnormalities, cancer predisposition and exhibit defective DNA crosslink repair. The relationship of this repair defect to disease traits remains unclear, given that crosslink sensitivity is recapitulated in FA mouse models without most of the other disease-related features. Mice deficient in Mus81 are also defective in crosslink repair, yet MUS81 mutations have not been linked to FA. Using mice deficient in both Mus81 and the FA pathway protein FancC, we show both proteins cooperate in parallel pathways, as concomitant loss of FancC and Mus81 triggered cell-type-specific proliferation arrest, apoptosis and DNA damage accumulation in utero. Mice deficient in both FancC and Mus81 that survived to birth exhibited growth defects and an increased incidence of congenital abnormalities. This cooperativity of FancC and Mus81 in developmental outcome was also mirrored in response to crosslink damage and chromosomal integrity. Thus, our findings reveal that both pathways safeguard against DNA damage from exceeding a critical threshold that triggers proliferation arrest and apoptosis, leading to compromised in utero development.

Involvement of Bcl-2-associated transcription factor 1 in the differentiation of early-born retinal cells.

Retinal progenitor proliferation and differentiation are tightly controlled by extrinsic cues and distinctive combinations of transcription factors leading to the generation of retinal cell type diversity. In this context, we have characterized Bcl-2-associated transcription factor (Bclaf1) during rodent retinogenesis. Bclaf1 expression is restricted to early-born cell types, such as ganglion, amacrine, and horizontal cells. Analysis of developing retinas in Bclaf1-deficient mice revealed a reduction in the numbers of retinal ganglion cells, amacrine cells and horizontal cells and an increase in the numbers of cone photoreceptor precursors. Silencing of Bclaf1expression by in vitro electroporation of shRNA in embryonic retina confirmed that Bclaf1 serves to promote amacrine and horizontal cell differentiation. Misexpression of Bclaf1 in late retinal progenitors was not sufficient to directly induce the generation of amacrine and horizontal cells. Domain deletion analysis indicated that the N-terminal domain of Bclaf1 containing an arginine-serine-rich and a bZip domain is required for its effects on retinal cell differentiation. In addition, analysis revealed that Bclaf1 function occurs independently of its interaction with endogenous Bcl-2-related proteins. Altogether, our data demonstrates that Bclaf1expression in postmitotic early-born cells facilitates the differentiation of early retinal precursors into retinal ganglion cells, amacrine cells, and horizontal cells rather than into cone photoreceptors.

2,3,7,8-Tetrachlorodibenzo-p-dioxin poly(ADP-ribose) polymerase (TiPARP, ARTD14) is a mono-ADP-ribosyltransferase and repressor of aryl hydrocarbon receptor transactivation.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD)-inducible poly(ADP-ribose) polymerase (TiPARP/ARTD14) is a member of the PARP family and is regulated by the aryl hydrocarbon receptor (AHR); however, little is known about TiPARP function. In this study, we examined the catalytic function of TiPARP and determined its role in AHR transactivation. We observed that TiPARP exhibited auto-mono-ADP-ribosyltransferase activity and ribosylated core histones. RNAi-mediated knockdown of TiPARP in T-47D breast cancer and HuH-7 hepatoma cells increased TCDD-dependent cytochrome P450 1A1 (CYP1A1) and CYP1B1 messenger RNA (mRNA) expression levels and recruitment of AHR to both genes. Overexpression of TiPARP reduced AHR-dependent increases in CYP1A1-reporter gene activity, which was restored by overexpression of AHR, but not aryl hydrocarbon receptor nuclear translocator. Deletion and mutagenesis studies showed that TiPARP-mediated inhibition of AHR required the zinc-finger and catalytic domains. TiPARP and AHR co-localized in the nucleus, directly interacted and both were recruited to CYP1A1 in response to TCDD. Overexpression of Tiparp enhanced, whereas RNAi-mediated knockdown of TiPARP reduced TCDD-dependent AHR proteolytic degradation. TCDD-dependent induction of AHR target genes was enhanced in Tiparp(-/-) mouse embryonic fibroblasts compared with wildtype controls. Our findings show that TiPARP is a mono-ADP-ribosyltransferase and a transcriptional repressor of AHR, revealing a novel negative feedback loop in AHR signalling.

Dual energy X-ray absorptiometry of the knee in spinal cord injury: methodology and correlation with quantitative computed tomography.

STUDY DESIGN: Comparison of diagnostic tests; methodological validation. OBJECTIVES: Primary: to investigate the precision and reliability of a knee bone mineral density (BMD) assessment protocol that uses an existing dual energy X-ray absorptiometry (DXA) forearm acquisition algorithm in individuals with spinal cord injury (SCI). Secondary: to correlate DXA-based knee areal BMD with volumetric BMD assessments derived from quantitative computed tomography (QCT). SETTING: Academic medical center, Chicago, IL, USA. PARTICIPANTS: a convenience sample of 12 individuals with acute SCI recruited for an observational study of bone loss and 34 individuals with chronic SCI who were screened for a longitudinal study evaluating interventions to increase BMD. MAIN OUTCOME MEASURES: Root-mean-square standard deviation (RMS-SD) and intra/inter-rater reliability of areal BMD acquired at three knee regions using an existing DXA forearm acquisition algorithm; correlation of DXA-based areal BMD with QCT-derived volumetric BMD. RESULTS: The RMS-SD of areal BMD at the distal femoral epiphysis, distal femoral metaphysis and proximal tibial epiphysis averaged 0.021, 0.012 and 0.016 g cm(-2), respectively, in acute SCI and 0.018, 0.02 and 0.016 g cm(-2) in chronic SCI. All estimates of intra/inter-rater reliability exceeded 97% and DXA-based areal BMD was significantly correlated with QCT-derived volumetric BMD at all knee regions analyzed. CONCLUSIONS: Existing DXA forearm acquisition algorithms are sufficiently precise and reliable for short-term assessments of knee BMD in individuals with SCI. Future work is necessary to quantify the reliability of this approach in longitudinal investigations and to determine its ability to predict fractures and recovery potential. SPONSORSHIP: This work was funded by the Department of Defense, grant number DOD W81XWH-10-1-0951, with partial support from Merck & Co, Inc.

Effect of bone morphogenetic protein-2, demineralized bone matrix and systemic parathyroid hormone (1-34) on local bone formation in a rat calvaria critical-size defect model.

AIM: To determine the potential of recombinant human bone morphogenetic protein-2 (rhBMP-2) soak-loaded on to an absorbable collagen sponge (ACS) to induce local bone formation compared with the clinical reference demineralized bone matrix (DBM) and to investigate potential additive/synergistic effects of exogenous parathyroid hormone (PTH). METHODS: Critical-size (8 mm), through-through calvaria osteotomy defects in 160 adult male Sprague-Dawley rats were randomized to receive one of eight interventions: rhBMP-2/ACS, DBM, ACS, or serve as controls (empty defects) combined or not with systemic PTH. Ten animals from each group were followed for 4 and 8 wks for radiographic and histometric analysis. Multivariable analysis was used to assess the effect of experimental intervention and healing time on local bone formation. RESULTS: In the multivariable analysis, rhBMP-2/ACS exhibited significantly greater histologic bone formation than control (ß ± SE: 54.76 ± 5.85, p < 0.001) and ACS (ß ± SE: 9.14 ± 3.31, p = 0.007) whereas DBM showed significantly less bone formation than control (ß ± SE: -32.32 ± 8.23, p < 0.001). Overall, PTH did not show a significant effect on bone formation (ß ± SE: 2.72 ± 6.91, p = 0.70). No significant differences in histological defect closure were observed between 4 and 8 wks for all but the control group without PTH. CONCLUSION: rhBMP-2/ACS significantly stimulates local bone formation whereas bone formation appears significantly limited by DBM. Systemic application of PTH provided no discernible additive/synergistic effects on local bone formation.

zA map for sequence analysis of the Arabidopsis thaliana genome.

Arabidopsis thaliana has emerged as a model system for studies of plant genetics and development, and its genome has been targeted for sequencing by an international consortium (the Arabidopsis Genome Initiative; http://genome-www. stanford.edu/Arabidopsis/agi.html). To support the genome-sequencing effort, we fingerprinted more than 20,000 BACs (ref. 2) from two high-quality publicly available libraries, generating an estimated 17-fold redundant coverage of the genome, and used the fingerprints to nucleate assembly of the data by computer. Subsequent manual revision of the assemblies resulted in the incorporation of 19,661 fingerprinted BACs into 169 ordered sets of overlapping clones ('contigs'), each containing at least 3 clones. These contigs are ideal for parallel selection of BACs for large-scale sequencing and have supported the generation of more than 5.8 Mb of finished genome sequence submitted to GenBank; analysis of the sequence has confirmed the integrity of contigs constructed using this fingerprint data. Placement of contigs onto chromosomes can now be performed, and is being pursued by groups involved in both sequencing and positional cloning studies. To our knowledge, these data provide the first example of whole-genome random BAC fingerprint analysis of a eucaryote, and have provided a model essential to efforts aimed at generating similar databases of fingerprint contigs to support sequencing of other complex genomes, including that of human.

Identification of Nymphal Instars of emMecidea/em emmajor/em Sailer and Mecidea minor Ruckes (Hemiptera: Heteroptera: Pentatomidae: Mecideini) from the Southwestern and Central United States.

The mecideine stink bug genus Mecidea is represented in America north of Mexico by three species: Mecidea major Sailor, Mecidea minor Ruckes, and Mecidea longula Stål. M. major and M. minor are widely distributed, occurring collectively from the Midwest to California. M. longula is known only from south Florida. The life histories of M. major and M. minor have been published including laboratory rearing from egg to adult and descriptions of the immature stages. However, no key has been developed for identification of the nymphs of these two species. Here, we present a key to the nymphs of these taxa to the species and instar levels.

In search of a function for BCLAF1.

BCLAF1 was originally identified as a protein that interacts with antiapoptotic members of the Bcl2 family. Initial studies indicated a role for this protein as an inducer of apoptosis and repressor of transcription. Subsequent studies have shown that BCLAF1 plays criticals roles in a wide range of processes that are not normally associated with actions of Bcl2 family members, including lung development, T-cell activation, and control of the lytic infection program of Kaposi's sarcoma-associated herpesvirus. Here, we provide an overview of findings from past studies that both support and challenge the role of BCLAF1 in cell death and transcriptional control. We also present recent findings from our laboratory and others indicating a role for BCLAF1 in post-transcriptional processes that impact mRNA metabolism, instead of a direct role for this protein in apoptosis or transcription.

Biochemical and ultrastructural studies of proteoheparan sulfates synthesized by PYS-2, a basement membrane-producing cell line.

The mouse teratocarcinoma-derived cell line, PYS-2, has been shown to produce laminin, a basement membrane-specific glycoprotein. In these studies we demonstrate that PYS-2 cells synthesize and secrete into the culture medium a proteoglycan which contains only heparan sulfate as its sulfated polysaccharide side chains, as well as type IV procollagen and laminin. The apparent molecular weights of the proteoglycan and its heparan sulfate side chain were estimated to be 400,000 and 25,000, respectively, by gel chromatography. A proteoheparan sulfate with properties closely similar, if not identical, to those of the proteoglycan in the medium, together with two heparan sulfate single chains of different molecular size, were extracted from the cell layer with 2% SDS in the presence of protease inhibitors. Ultrastructurally, a fine fibrillar intercellular matrix was recognized which contained discrete 100-200 A diameter ruthenium red-positive granules interspersed throughout the filamentous meshwork. The PYS-2 cultures were shown by immunofluorescence to react with antibodies against the heparan sulfate-containing proteoglycan isolated from the mouse EHS sarcoma (Hassell, J. R., P. G. Robey, H. J. Barrach, J. Wilczek, S. I. Rennard, and G. R. Martin. 1980. Proc. Natl. Acad. Sci. U. S. A. 77:4494-4498). Immunoelectron microscopic examination, using the same antibodies, revealed that the proteoheparan sulfate was located not only at the edges but also within the interstices of the matrix. These findings indicate that PYS-2 cells synthesize and secrete a proteoglycan with properties similar to those of basement membrane proteoglycan. These cells may therefore serve as a useful model system for the study of the biosynthesis and structure of basement membranes.

Duplication of CaMV 35S Promoter Sequences Creates a Strong Enhancer for Plant Genes.

A variant of the cauliflower mosaic virus 35S promoter with transcriptional activity approximately tenfold higher than that of the natural promoter was constructed by tandem duplication of 250 base pairs of upstream sequences. The duplicated region also acted as a strong enhancer of heterologous promoters, increasing the activity of an adjacent and divergently transcribed transferred DNA gene several hundredfold, and to a lesser extent, that of another transferred DNA gene from a remote downstream position. This optimized enhancer element should be very useful for obtaining high levels of expression of foreign genes in transgenic plants.

A role for Mus81 in the repair of chromium-induced DNA damage.

Hexavalent chromium (Cr[VI]) is a toxic environmental contaminant that is capable of producing a broad spectrum of DNA damage. The ability of Cr[VI] to induce mutagenesis and neoplastic transformation has been attributed to its genotoxic action, however our understanding of molecular mechanisms involved in the repair of Cr[VI]-induced DNA damage remains incomplete. Here, we report that Mus81, an enzyme that participates with Eme1 in the resolution of replication fork damage caused by certain lesions, is involved in the repair of Cr[VI]-induced DNA damage. Mus81-deficient cells were found to be more susceptible to Cr[VI]-induced proliferation arrest and more sensitive to the long-term cytotoxic effects of Cr[VI] than isogenic wild-type cells. Following Cr[VI] exposure, Mus81-deficient cells displayed a lag in the disappearance of Rad51 foci, exhibited elevated replication-associated gamma-H2AX and showed an increased incidence of chromosomal instability compared to wild-type cells. Our findings support a role for Mus81 in the resolution of replication-associated DNA damage associated with this genotoxic agent, by converting Cr[VI]-DNA lesions into a form more amenable for homologous recombination.

Assessing the metabolic and toxic effects of anticonvulsant doses of polyunsaturated fatty acids on the liver in rats.

Polyunsaturated fatty acids (PUFA), at high doses, have been demonstrated to possess anticonvulsant properties in animal seizure models. Little is known, however, about the possible metabolic or adverse effects of PUFA at these high, anticonvulsant doses. The goal of the present study was to assess the metabolic and potential adverse effects of high-dose PUFA administration to rats. Adult male rats received a fatty acid mixture containing alpha-linolenic and linoleic acid in a 1 to 4 ratio, intraperitoneally, for 3 wk. After sacrifice, livers were isolated and analyzed for fatty acid composition and for mRNA expression of HMG-CoA lyase, catalase, and glutathione S-transferases A1 and A4, markers for ketosis, antioxidant defense, and phase II xenobiotic metabolism, respectively. Chronic administration of the PUFA mixture decreased hepatic levels of total lipids--and several fatty acids within total lipids--without altering mRNA expression of HMG-CoA lyase, a metabolic marker of ketosis. The PUFA mixture did not affect mRNA expression of catalase or glutathione S-transferases A1 and A4, which are involved in antioxidant defense and phase II xenobiotic metabolism. These findings suggest that PUFA, given for 3 wk at anticonvulsant doses, result in significant changes in liver lipid metabolism, but do not alter measured genetic markers of liver toxicity.

Changes in the relative expression pattern of multiple vitellogenin genes in adult male and larval zebrafish exposed to exogenous estrogens.

Production of the lipoprotein vitellogenin (Vg) is induced in fish upon exposure to estrogens and is a biomarker of endocrine disruption in fish. In some fish, three types of Vg (VgA, VgB, and VgC) are recognized and transcribed from at least three distinct Vg genes (vtg). We investigated expression of vtg coding for Vg1A/B, Vg2A/B, and VgC in adult male and larval zebrafish exposed to various estrogenic substances. Quantitative PCR was conducted for transcripts of each vtg and a control gene (beta-actin). Male fish were exposed to 17beta-estradiol (E2) and 17alpha-ethinylestradiol, total RNA was extracted from excised liver, and histopathology of liver, trunk kidney, and gonads was conducted. Larval fish were exposed to 10 different estrogenic substances and total RNA was extracted from groups of whole larvae. In adult male fish, the relative fold change varied, but pattern of expression change (i.e., Vg1A/B > Vg2A/B > VgC) was consistent. Larger males exposed to E2 had significantly higher induction of each vtg. In larval zebrafish, the relative fold change in vtg expression varied according to specific estrogenic substance tested, but the pattern of change (i.e., Vg2A/B > Vg1A/B > VgC) was consistent for each substance that induced vtg.

The experience of group weight loss efforts among lesbians.

OBJECTIVE: To describe experiences of group weight loss efforts among lesbians participating in a predominantly lesbian weight loss group. METHODS: A qualitative study (N = 14 self-identified overweight lesbians) was conducted, incorporating phenomenology and grounded theory in methodology. Focus groups were analyzed using a Template Analysis style. RESULTS: Several themes were identified that contributed to the weight loss experience of the participants. These themes contributed to the development of a model depicting history, components, and relationships among concepts leading to positive weight loss experiences for lesbians. CONCLUSIONS: Participants had long histories of shame and fear surrounding weight loss attempts. Weight loss group participants needed cultural connectivity and a sense of safety and acceptance to address issues contributing to weight gain and in order to lose weight and maintain weight loss. Although more research is needed before implementation of a sexual minority women-specific weight loss program, these data are the basis for further exploration into the development of such a program.

Nymphal development of Rhopalus maculatus and Chorosoma schillingii (Hemiptera: Heteroptera: Rhopalidae: Rhopalinae) and development of trichobothrial pattern in Rhopalidae.

The nymphal morphology of the family Rhopalidae is well known for several species, most of which are pests. Ontogenetic changes in morphology are described in detail for two members from two tribes of the subfamily Rhopalinae, Rhopalini: Rhopalus (Aeschyntelus) maculatus (Fieber) and Chorosomini: Chorosoma schillingii (Schilling). Keys to the nymphal instars of both species are provided. The special types of nymphal setae and lateral abdominal processes are described in R. maculatus. The ontogenetic development of abdominal trichobothria is described, and the known trichobothrial patterns within the Rhopalidae are discussed.

Systematic relationship between Piezodorus guildinii and P. hybneri (Hemiptera: Heteroptera: Pentatomidae) and diagnostic characters to separate species.

The pentatomid genus Piezodorus presently contains about 12 species, all of which, with one exception [i.e., P. guildinii (Westwood)], have been reported from the Old World; P. guildinii apparently is limited to the New World. P. guildinii and P. hybneri (Gmelin) are similar in appearance and have been considered by some as sister species. The following characteristics we consider valid as diagnostic to separate the two species: for males, the shape and length of the vesica, differences in the anatomy of the conjunctival appendages and penial plate, and shape of the pygophore and parameres; and for females, differences in the anatomy of the first and second gonocoxae, ninth paratergites, and spermathecae.

Detection of acute infections during HIV testing in North Carolina.

BACKGROUND: North Carolina has added nucleic acid amplification testing for the human immunodeficiency virus (HIV) to standard HIV antibody tests to detect persons with acute HIV infection who are viremic but antibody-negative. METHODS: To determine the effect of nucleic acid amplification testing on the yield and accuracy of HIV detection in public health practice, we conducted a 12-month observational study of methods for state-funded HIV testing. We compared the diagnostic performance of standard HIV antibody tests (i.e., enzyme immunoassay and Western blot analysis) with an algorithm whereby serum samples that yielded negative results on standard antibody tests were tested again with the use of nucleic acid amplification. A surveillance algorithm with repeated sensitive-less-sensitive enzyme immunoassay tests was also evaluated. HIV infection was defined as a confirmed positive result on a nucleic acid amplification test or as HIV antibody seroconversion. RESULTS: Between November 1, 2002, and October 31, 2003, 109,250 persons at risk for HIV infection who had consented to HIV testing presented at state-funded sites. There were 606 HIV-positive results. Established infection, as identified by standard enzyme immunoassay or Western blot analysis, appeared in 583 participants; of these, 107 were identified, with the use of sensitive-less-sensitive enzyme immunoassay tests, as recent infections. A total of 23 acutely infected persons were identified only with the use of the nucleic acid amplification algorithm. With all detectable infections taken into account, the sensitivity of standard antibody testing was 0.962 (95 percent confidence interval, 0.944 to 0.976). There were two false positive results on nucleic acid amplification tests. The specificity and positive predictive value of the algorithm that included nucleic acid amplification testing were greater than 0.999 (95 percent confidence interval, 0.999 to >0.999) and 0.997 (95 percent confidence interval, 0.988 to >0.999), respectively. Of the 23 acute HIV infections, 16 were detected at sexually transmitted disease clinics. Emergency measures for HIV prevention protected 48 sex partners and one fetus from high-risk exposure to HIV. CONCLUSIONS: The addition of nucleic acid amplification testing to an HIV testing algorithm significantly increases the identification of cases of infection without impairing the performance of diagnostic testing. The detection of highly contagious, acutely infected persons creates new opportunities for HIV surveillance and prevention.