CELL PROLIFERATION IN THE ERYTHROID COMPARTMENT OF GUINEA-PIG BONE MARROW: STUDIES WITH 3H-THYMIDINE

Single and multiple injections of ³H-TdR have been used for measuring the rate of proliferation in morphologically defined cell populations of guinea-pig bone marrow that are committed to erythroid differentiation. The conclusions are based on the analysis of absolute cell numbers in the maturational compartments, the labeling and mitotic indices, labeled mitotic curves, pulse and chase grain counts over dividing and interphase cells, and on the rate of labeling during multiple, repeated injections of ³H-TdR. The average duration of S and the rate of cycling is similar in all maturational compartments of the erythron. The majority of cells progress to the next maturational compartment by the time they divide for the second time. All proerythroblasts and basophilic erythroblasts are in cycle. Polychromatic erythroblasts incapable of incorporating ³H-TdR reach the orthochromatic population in the span of 5–6 hr. The orthochromatic population is renewed every 20–24 hr. The number of divisions between the proerythroblast and orthochromatic erythroblast does not exceed four and some cells may undergo only two divisions during the maturation pathway. Cell input from a progenitor cell population contributes to the maintenance of the erythron. The kinetic behavior of progenitor cells is similar to that of proerythroblasts. By the time of their second division, progenitor cells may reach either the proerythroblast or basophilic erythroblast compartments. The kinetic behavior of basophilic transitional cells corresponds to the predicted behavior of the erythroblast progenitor cell pool. Several of the conclusions are based on the assumption that grain count halving is the result of cell division. In view of the evidence discussed, this assumption in the present studies seems justified.     

Discovery and evaluation of spirocyclic derivatives as antagonists of the neuropeptide Y5 receptor

A novel series of spirocyclic derivatives was synthesized and evaluated as NPY Y5R antagonists for the treatment of obesity. Cis and trans analogs 7a and 8a were equipotent in a Y5R binding assay (K(i)'s ≤ 1 nM) and displayed good stability in human and rat liver microsome preparations. Compound 7a failed to demonstrate weight loss activity in a diet-induced obese (DIO) rat model at unbound drug levels in the brain that exceeded the Y5R K(i) value by 25-fold over a 24-h time-period.     

Extensive copy-number variation of the human olfactory receptor gene family

As much as a quarter of the human genome has been reported to vary in copy number between individuals, including regions containing about half of the members of the olfactory receptor (OR) gene family. We have undertaken a detailed study of copy-number variation of ORs to elucidate the selective and mechanistic forces acting on this gene family and the true impact of copy-number variation on human OR repertoires. We argue that the properties of copy-number variants (CNVs) and other sets of large genomic regions violate the assumptions of statistical methods that are commonly used in the assessment of gene enrichment. Using more appropriate methods, we provide evidence that OR enrichment in CNVs is not due to positive selection but is because of OR preponderance in segmentally duplicated regions, which are known to be frequently copy-number variable, and because purifying selection against CNVs is lower in OR-containing regions than in regions containing essential genes. We also combine multiplex ligation-dependent probe amplification (MLPA) and PCR to assay the copy numbers of 37 candidate CNV ORs in a panel of ~50 human individuals. We confirm copy-number variation of 18 ORs but find no variation in this human-diversity panel for 16 other ORs, highlighting the caveat that reported intervals often overrepresent true CNVs. The copy-number variation we describe is likely to underpin significant variation in olfactory abilities among human individuals. Finally, we show that both homology-based and homology-independent processes have played a recent role in remodeling the OR family.     

Respondent Driven Sampling: Determinants of Recruitment and a Method to Improve Point Estimation

Introduction

Respondent-driven sampling (RDS) is a variant of a link-tracing design intended for generating unbiased estimates of the composition of hidden populations that typically involves giving participants several coupons to recruit their peers into the study. RDS may generate biased estimates if coupons are distributed non-randomly or if potential recruits present for interview non-randomly. We explore if biases detected in an RDS study were due to either of these mechanisms, and propose and apply weights to reduce bias due to non-random presentation for interview.

Methods

Using data from the total population, and the population to whom recruiters offered their coupons, we explored how age and socioeconomic status were associated with being offered a coupon, and, if offered a coupon, with presenting for interview. Population proportions were estimated by weighting by the assumed inverse probabilities of being offered a coupon (as in existing RDS methods), and also of presentation for interview if offered a coupon by age and socioeconomic status group.

Results

Younger men were under-recruited primarily because they were less likely to be offered coupons. The under-recruitment of higher socioeconomic status men was due in part to them being less likely to present for interview. Consistent with these findings, weighting for non-random presentation for interview by age and socioeconomic status group greatly improved the estimate of the proportion of men in the lowest socioeconomic group, reducing the root-mean-squared error of RDS estimates of socioeconomic status by 38%, but had little effect on estimates for age. The weighting also improved estimates for tribe and religion (reducing root-mean-squared-errors by 19–29%), but had little effect for sexual activity or HIV status.

Conclusions

Data collected from recruiters on the characteristics of men to whom they offered coupons may be used to reduce bias in RDS studies. Further evaluation of this new method is required.     

Predation risk is unlikely to account for the failure of subordinate speckled warblers Chthonicola sagittata to help at the nest

The predator avoidance hypothesis suggests that the failure of subordinate birds to provision nestlings in communally breeding species is a consequence of increased predation risk. Parents exclude subordinates from the nest area and thus reduce the frequency of predator-attracting visits when the nest is most vulnerable, leading to increased reproductive success. I evaluated this hypothesis for the speckled warbler Chthonicola sagittata , a group-living member of the Pardalotidae in which subordinate males never feed nestlings or fledglings even though they are unrelated to the primary pair, compete for copulations and sometimes sire young in the brood. Parents did not modify provisioning behaviour relative to the risk of nest predation; provisioning rates to 10 d-old nestlings were similar on high and low risk territories. Furthermore, there was no evidence that parents modified the timing of deliveries or adjusted the relative size of deliveries in relation to predation risk. The condition (residual mass) of nestlings differed between high and low risk territories because nestlings on high risk territories had smaller tarsi but similar body mass to those at low risk. Tarsus length was the result of parental phenotype, not modified provisioning behaviour. Given that parents were unresponsive to predation risk, it seems unlikely that predation can account for the failure of subordinates to provision at the nest.     

Human proximity and habitat fragmentation are key drivers of the rangewide bonobo distribution

Habitat loss and hunting threaten bonobos (Pan paniscus), Endangered (IUCN) great apes endemic to lowland rainforests of the Democratic Republic of Congo. Conservation planning requires a current, data-driven, rangewide map of probable bonobo distribution and an understanding of key attributes of areas used by bonobos. We present a rangewide suitability model for bonobos based on a maximum entropy algorithm in which data associated with locations of bonobo nests helped predict suitable conditions across the species’ entire range. We systematically evaluated available biotic and abiotic factors, including a bonobo-specific forest fragmentation layer (forest edge density), and produced a final model revealing the importance of simple threat-based factors in a data poor environment. We confronted the issue of survey bias in presence-only models and devised a novel evaluation approach applicable to other taxa by comparing models built with data from geographically distinct sub-regions that had higher survey effort. The model’s classification accuracy was high (AUC = 0.82). Distance from agriculture and forest edge density best predicted bonobo occurrence with bonobo nests more likely to occur farther from agriculture and in areas of lower edge density. These results suggest that bonobos either avoid areas of higher human activity, fragmented forests, or both, and that humans reduce the effective habitat of bonobos. The model results contribute to an increased understanding of threats to bonobo populations, as well as help identify priority areas for future surveys and determine core bonobo protection areas.     

Optimizing the expression of chimeric genes in plant cells

Summary The Serratia marcescens chiA gene encodes a secreted chitinase activity which contributes to the fungal growth inhibition exhibited by this bacterium. The coding region from the chiA gene was fused to the promoter and 3 polyadenylation region of the Agrobacterium nopaline synthase gene. Site-directed mutagenesis of specific nucleotides surrounding the initiating AUG of the coding sequence of this chimeric gene resulted in up to an eight-fold increase in the amount of chitinase protein detected in transformed plant tissue. Analysis of the chiA mRNA indicated that these nucleotides also affected mRNA levels. At least 50% of the chitinase protein produced in transformed tobacco cells was the same molecular weight as the S. marcescen secreted protein.     

Modeling perimenopause in Sprague-Dawley rats by chemical manipulation of the transition to ovarian failure

Various age-related diseases increase in incidence during perimenopause. However, our understanding of the effects of aging compared with hormonal changes of perimenopause in mediating these disease risks is incomplete, in part due to the lack of an experimental perimenopause model. We therefore aimed to determine whether manipulation of the transition to ovarian failure in rats via the use of 4-vinylcyclohexene diepoxide (VCD) could be used to model and accelerate hormonal changes characteristic of perimenopause. We examined long-term (11 to 20 mo), dose-dependent effects of VCD on reproductive function in 1- and 3-mo-old female Sprague-Dawley rats. Twenty-five daily doses of VCD (80 or 160 mg/kg daily compared with vehicle alone) depleted ovarian follicles in a dose-dependent fashion in rats of both ages, accelerated the onset of acyclicity, and caused dose-dependent increases in follicle-stimulating hormone that exceeded those naturally occurring with age in control rats but left serum levels of 17β-estradiol unchanged, with continued ovarian production of androstenedione. High-dose VCD caused considerable nonovarian toxicities in 3-mo-old Sprague-Dawley rats, making this an unsuitable model. In contrast, 1-mo-old rats had more robust dose-dependent increases in follicle-stimulating hormone without evidence of systemic toxicity in response to either VCD dose. Because perimenopause is characterized by an increase in follicle-stimulating hormone with continued secretion of ovarian steroids, VCD acceleration of an analogous hormonal milieu in 1-mo-old Sprague-Dawley rats may be useful for probing the hormonal effects of perimenopause on age-related disease risk.     

FGF signaling through FGFR1 is required for olfactory bulb morphogenesis

During development, the embryonic telencephalon is patterned into different areas that give rise to distinct adult brain structures. Several secreted signaling molecules are expressed at putative signaling centers in the early telencephalon. In particular, Fgf8 is expressed at the anterior end of the telencephalon and is hypothesized to pattern it along the anteroposterior (AP) axis. Using a CRE/loxP genetic approach to disrupt genes in the telencephalon, we address the role of FGF signaling directly in vivo by abolishing expression of the FGF receptor Fgfr1. In the Fgfr1-deficient telencephalon, AP patterning is largely normal. However, morphological defects are observed at the anterior end of the telencephalon. Most notably, the olfactory bulbs do not form normally. Examination of the proliferation state of anterior telencephalic cells supports a model for olfactory bulb formation in which an FGF-dependent decrease in proliferation is required for initial bulb evagination. Together the results demonstrate an essential role for Fgfr1 in patterning and morphogenesis of the telencephalon.