Case-control study of fetal microchimerism and breast cancer

Background

Prior pregnancy is known to protect against development of breast cancer. Recent studies have demonstrated that pregnancy has the capacity to establish small numbers of immunologically active fetal-derived cells in the mother, a phenomenon known as fetal microchimerism (FMc). We asked whether presence of FMc, routinely acquired during pregnancy, is a protective factor for breast cancer.

Methodology/Principal Findings

DNA extracts from peripheral blood specimens were obtained from a population-based case-control study of risk factors for breast cancer in women 21 to 45 years old. Specimens were tested with quantitative PCR for presence and concentrations of male DNA presumed to derive from prior pregnancies with a male fetus. Odds ratios (OR) and 95% confidence intervals (CI) were estimated with consideration of multiple established reproductive and environmental risk factors for breast cancer. FMc results were generated on 99 parous women, 54 with primary invasive breast cancer and 45 general population controls. FMc prevalence was 56% (25/45) and 26% (14/54) in controls and cases, respectively. Women harboring FMc were less likely to have had breast cancer (OR = 0.29, 95% CI 0.11–0.83; p = 0.02, adjusting for age, number of children, birth of a son, history of miscarriage, and total DNA tested). In addition, FMc concentrations were higher in controls versus cases (p = 0.01). Median concentrations were 2 (0–78) and 0 (0–374) fetal genomes/10⁶ maternal genomes in controls and cases, respectively.

Conclusions

Results suggest that the enigma of why some parous women are not afforded protection from breast cancer by pregnancy might in part be explained by differences in FMc. Mechanistic studies of FMc-derived protection against breast cancer are warranted.     

Genome Scan of M. tuberculosis Infection and Disease in Ugandans

Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), is an enduring public health problem globally, particularly in sub-Saharan Africa. Several studies have suggested a role for host genetic susceptibility in increased risk for TB but results across studies have been equivocal. As part of a household contact study of Mtb infection and disease in Kampala, Uganda, we have taken a unique approach to the study of genetic susceptibility to TB, by studying three phenotypes. First, we analyzed culture confirmed TB disease compared to latent Mtb infection (LTBI) or lack of Mtb infection. Second, we analyzed resistance to Mtb infection in the face of continuous exposure, defined by a persistently negative tuberculin skin test (PTST-); this outcome was contrasted to LTBI. Third, we analyzed an intermediate phenotype, tumor necrosis factor-alpha (TNFα) expression in response to soluble Mtb ligands enriched with molecules secreted from Mtb (culture filtrate). We conducted a full microsatellite genome scan, using genotypes generated by the Center for Medical Genetics at Marshfield. Multipoint model-free linkage analysis was conducted using an extension of the Haseman-Elston regression model that includes half sibling pairs, and HIV status was included as a covariate in the model. The analysis included 803 individuals from 193 pedigrees, comprising 258 full sibling pairs and 175 half sibling pairs. Suggestive linkage (p<10⁻³) was observed on chromosomes 2q21-2q24 and 5p13-5q22 for PTST-, and on chromosome 7p22-7p21 for TB; these findings for PTST- are novel and the chromosome 7 region contains the IL6 gene. In addition, we replicated recent linkage findings on chromosome 20q13 for TB (p = 0.002). We also observed linkage at the nominal α = 0.05 threshold to a number of promising candidate genes, SLC11A1 (PTST- p = 0.02), IL-1 complex (TB p = 0.01), IL12BR2 (TNFα p = 0.006), IL12A (TB p = 0.02) and IFNGR2 (TNFα p = 0.002). These results confirm not only that genetic factors influence the interaction between humans and Mtb but more importantly that they differ according to the outcome of that interaction: exposure but no infection, infection without progression to disease, or progression of infection to disease. Many of the genetic factors for each of these stages are part of the innate immune system.     

Three-Dimensional High-Resolution Second-Harmonic Generation Imaging of Endogenous Structural Proteins in Biological Tissues

We find that several key endogenous protein structures give rise to intense second-harmonic generation (SHG)—nonabsorptive frequency doubling of an excitation laser line. Second-harmonic imaging microscopy (SHIM) on a laser-scanning system proves, therefore, to be a powerful and unique tool for high-resolution, high-contrast, three-dimensional studies of live cell and tissue architecture. Unlike fluorescence, SHG suffers no inherent photobleaching or toxicity and does not require exogenous labels. Unlike polarization microscopy, SHIM provides intrinsic confocality and deep sectioning in complex tissues. In this study, we demonstrate the clarity of SHIM optical sectioning within unfixed, unstained thick specimens. SHIM and two-photon excited fluorescence (TPEF) were combined in a dual-mode nonlinear microscopy to elucidate the molecular sources of SHG in live cells and tissues. SHG arose not only from coiled-coil complexes within connective tissues and muscle thick filaments, but also from microtubule arrays within interphase and mitotic cells. Both polarization dependence and a local symmetry cancellation effect of SHG allowed the signal from species generating the second harmonic to be decoded, by ratiometric correlation with TPEF, to yield information on local structure below optical resolution. The physical origin of SHG within these tissues is addressed and is attributed to the laser interaction with dipolar protein structures that is enhanced by the intrinsic chirality of the protein helices.     

Spindle Dynamics and the Role of γ-Tubulin in Early Caenorhabditis elegans Embryos

gamma-Tubulin is a ubiquitous and highly conserved component of centrosomes in eukaryotic cells. Genetic and biochemical studies have demonstrated that gamma-tubulin functions as part of a complex to nucleate microtubule polymerization from centrosomes. We show that, as in other organisms, Caenorhabditis elegans gamma-tubulin is concentrated in centrosomes. To study centrosome dynamics in embryos, we generated transgenic worms that express GFP::gamma-tubulin or GFP::beta-tubulin in the maternal germ line and early embryos. Multiphoton microscopy of embryos produced by these worms revealed the time course of daughter centrosome appearance and growth and the differential behavior of centrosomes destined for germ line and somatic blastomeres. To study the role of gamma-tubulin in nucleation and organization of spindle microtubules, we used RNA interference (RNAi) to deplete C. elegans embryos of gamma-tubulin. gamma-Tubulin (RNAi) embryos failed in chromosome segregation, but surprisingly, they contained extensive microtubule arrays. Moderately affected embryos contained bipolar spindles with dense and long astral microtubule arrays but with poorly organized kinetochore and interpolar microtubules. Severely affected embryos contained collapsed spindles with numerous long astral microtubules. Our results suggest that gamma-tubulin is not absolutely required for microtubule nucleation in C. elegans but is required for the normal organization and function of kinetochore and interpolar microtubules.     

Extending Ethnoprimatology: Human–Alloprimate Relationships in Managed Settings

The majority of studies in ethnoprimatology focus on areas of sympatry where humans and nonhuman primates (hereafter, primates) naturally coexist. We argue that much can be gained by extending the field’s scope to incorporate settings where humans manage most aspects of primates’ lives, such as zoos, laboratories, sanctuaries, and rehabilitation centers (hereafter, managed settings). We suggest that the mixed-methods approach of ethnoprimatology, which facilitates examination of both humans’ and primates’ responses to one another, can reveal not only how humans’ ideas about primates shape management strategies, but also how those management strategies affect primates’ lives. Furthermore, we note that a greater focus on managed settings will strengthen links between ethnoprimatology and primate rights/welfare approaches, and will introduce new questions into discussions of ethics in primatology. For example, managed settings raise questions about when it might be justifiable to restrict primates’ freedom for a “greater good,” and the desirability of making primates’ lives more “natural” even if this would decrease their well-being. Finally, we propose that because ethnoprimatology is premised on challenging false dichotomies between categories of field site—specifically, between “natural” and “unnatural” free-ranging populations—it makes sense for ethnoprimatologists to examine settings in which humans exert considerable control over primates’ lives, given that the distinction between “wild” and “captive” is similarly unclear.     

Transfer of methyl groups from N-dimethylnitrosamine to glycerolipids in rat liver.

We have examined the in vivo labeling of lipids after a single intraperitoneal injection of the carcinogen, (C14) dimethylnitrosamine, into rats. Liver was most active in incorporating (C14) methyl groups into lipids (0.91% of the injected dose) and 80% of the activity appeared in sn-3-phosphatidyl-choline. Chromatographic analysis of the products (and derivatives) formed after treatment of the (C14) phosphatidylcholine with phospholipase A2 (EC 3.1.1.4) and phospholipase C (EC 3.1.4.3) demonstrated that 89% of the radioactivity was in the choline moiety. These results indicate the transfer of methyl groups to lipids occurred via the lipid methylation pathway that converts phosphatidylethanolamine to phosphatidylcholine.     

Regional differentiation of fat bodies in larvae of the indianmeal moth,Plodia interpunctella

Larvae of the Indianmeal moth, Plodia interpunctella, contain two morphologically distinct fat bodies. Tan-colored, highly tracheated fat body located posteriorly in the abdomen was the predominant fat body tissue during the early larval instars. White, sheet fat body located more anteriorly became the predominant type during the fifth (last) larval instar and eventually occupied most of the space of the hemocoel. Ultrastructural morphology of tan fat body showed the tissue to be composed of cells containing numerous, large, spherical mitochondria, with only few lipid, glycogen, or protein storage structures. In contrast, white fat body was composed of cells that in later larval stages had organelles typical of storage functions. Both fat bodies produced storage proteins during the late fifth instar, whereas only white fat body accumulated the storage proteins. Tan fat body dispersed and apparently autolyzed in pharate pupae, whereas the white fat body metamorphosed and persisted into the adult stage. These observations indicate that fat body of the Indianmeal moth is functionally and morphologically differentiated along the anterior-posterior axis into two regional subgroups of cells.