NTP finds common indoor pollutant absorbed and distributed in rodents

The common indoor pollutant alpha-pinene is systemically absorbed and distributed to mammary glands of rodents following inhalation exposure, according to scientists in the NIEHS Division of the National Toxicology Program (DNTP). The research may be relevant to human health following exposure to alpha-pinene.

Alpha-pinene comes from the oil of coniferous trees and is a major component in turpentine. Human exposure to alpha-pinene can occur via the lumber industry, use of turpentine, and use of consumer products that contain alpha-pinene as a fragrance or flavoring ingredient. However, there is little data regarding the potential hazard from exposure to alpha-pinene.

Male and female rats and mice were exposed to alpha-pinene by whole body inhalation, at concentrations relevant to human exposures. After exposure, levels of alpha-pinene and its potential reactive metabolite, alpha-pinene oxide, were assessed in blood and mammary glands to estimate toxicokinetic parameters.

The researchers found alpha-pinene and alpha-pinene oxide in the blood and mammary glands and that levels increased with the exposure concentration. Generally, alpha-pinene and metabolite levels were higher in rats than in mice and in mammary glands than in blood. The findings may be important in providing context and relating animal toxicity findings to human exposures. (VP)

Citation: Waidyanatha S, Hackett M, Black SR, Stout MD, Fennell TR, Silinski MR, Watson SL, Licause J, Robinson VG, Sparrow B, Fernando RA, Cooper S, Rider CV. 2021. Toxicokinetic evaluation of the common indoor air pollutant, alpha-pinene, and its potential reactive metabolite, alpha-pinene oxide, following inhalation exposure in rodents. Toxicol Appl Pharmacol 418:115496.

Snapshots reveal how damaged nucleotides elude DNA repair safeguards

Using high-resolution time-lapse X-ray crystallography, NIEHS researchers and their collaborator demonstrated that DNA polymerase mu, involved in double strand break repair, uses a third metal site to insert damaged and undamaged nucleotides into DNA. Their findings shed light on how the incorporation of damaged nucleotides promotes the development of cancer and other diseases.

Mechanical stress or chemical agents arising from normal cellular processes or environmental exposures can cause breaks in one or both strands of the DNA double helix. Double strand breaks are the most deleterious of DNA lesions; their repair involves filling in gaps in DNA, a process mediated by DNA polymerases, and fusing of broken ends. This process is error-prone, and defective repair can lead to dangerous chromosome rearrangements underlying cancer and other diseases.

Oxidative stress generated by normal metabolism can also damage nucleotides, the building blocks of DNA. To examine whether insertion of damaged nucleotides, in this case oxidized dGTP, can exacerbate inefficiency of DNA repair, the researchers generated molecular snapshots of DNA polymerase mu in action using time-lapse X-ray crystallography. Oxidation elicited alterations in the nucleotide molecular attributes that allowed it to elude cellular defenses against insertion into DNA, promoting pro-mutagenic DNA repair. (FP)

Citation: Jamsen JA, Sassa A, Shock DD, Beard WA, Wilson SH. 2021. Watching a double strand break repair polymerase insert a pro-mutagenic oxidized nucleotide. Nat Commun 12(1):2059.

The association between body fat and puberty in girls

Girls who have higher total body fat (TBF) exhibit aberrations in hormonal and clinical indicators of puberty, according to NIEHS scientists and collaborators at the U.S. Centers for Disease Control and Prevention and the National Cancer Institute. It had been previously observed that girls who are either overweight or have obesity achieve both thelarche, the onset of breast development, and menarche, the first occurrence of menstruation, earlier than girls of normal weight. Additionally, girls with greater TBF spend shorter times in puberty.

The scientists conducted a four-year study in a cohort of 90 pubertal girls who were either overweight/obese or normal weight. Using a variety of imaging techniques and hormonal tests, the researchers discovered that levels of several reproductive hormones were initially similar between girls with lower and higher TBF, but after one year, hormones increased in those with higher TBF. Furthermore, girls with higher TBF exhibited slower breast development but experienced menarche sooner. These results indicate an association between higher TBF and irregularities in breast development and hormone levels, warranting further study into how body fat mediates these effects. (NA)

Citation: Ortega MT, McGrath JA, Carlson L, Flores Poccia V, Larson G, Douglas C, Sun BZ, Zhao S, Beery B, Vesper HW, Duke L, Botelho JC, Filie AC, Shaw ND. 2021. Longitudinal investigation of pubertal milestones and hormones as a function of body fat in girls. J Clin Endocrinol Metab 106(6):1668–1683. (Story)

A novel approach generates high-resolution images of proteins

NIEHS researchers and collaborators at Duke University and the National Cancer Institute developed a new approach to cryo-electron tomography (Cryo-ET). Such tomography provides atomic resolution data from cell sections and enables determination of macromolecular structures in cellulo, meaning through isolated live cells. Visualizing proteins in cellulo allows scientist to observe interactions between macromolecules that may be lost if they were extracted.

However, widespread use of atomic resolution Cryo-ET has been limited because datasets can take weeks to collect. To overcome that problem, the researchers created what is called beam image-shift electron cryo-tomography (BISECT), which shortens data acquisition time to a few days. The technique combines slow mechanical targeting with movement of the electron beam to multiply the number of areas imaged. Targeting errors are corrected by integrating geometrical constraints related to the tilting of the specimen. With this technique and others, the scientists were able to obtain an atomic resolution map in a couple of days.

BISECT combines methods that speed up data collection, increase precision in locating and focusing on the target, and improve data processing. It also allows collection of the necessary data for high-resolution, three-dimensional reconstructions of molecular structures by Cryo-ET to occur up to 10 times faster. (MH)

Citation: Bouvette J, Liu HF, Du X, Zhou Y, Sikkema AP, da Fonseca Rezende E Mello J, Klemm BP, Huang R, Schaaper RM, Borgnia MJ, Bartesaghi A. 2021. Beam image-shift accelerated data acquisition for near-atomic resolution single-particle cryo-electron tomography. Nat Commun 12(1):1957.

Breaking the feedback loop of allergic asthma

NIEHS researchers and their collaborators at the National Institute of Diabetes and Digestive and Kidney Diseases identified target molecules that reduce symptoms of allergic asthma in animal models. Allergic asthma is characterized by lung inflammation and airway constriction. The scientists found that specialized white blood cells called eosinophils, a sugar molecule known as uridine diphosphate-glucose (UDP-G), and the P2Y14 receptor worked together to produce asthma symptoms. The findings could lead to improved strategies for treating asthma.

After performing procedures that gave mice asthma, team members determined that UDP-G increases eosinophil recruitment to the airway and may also affect responses of other molecules to promote the allergen-induced immune response. The scientists also used genetically modified mice that lacked the P2Y14 receptor and observed that eosinophil infiltration and airway hyperresponsiveness were suppressed in this mouse model of allergic asthma.

Results from this study showed that P2Y14 antagonists — molecules that bind to P2Y14 but don’t activate it — may help to control allergy-induced asthma, especially in forms of the disease that are resistant to common treatments. (SR)

Citation: Karcz TP, Whitehead GS, Nakano K, Nakano H, Grimm SA, Williams JG, Deterding LJ, Jacobson KA, Cook DN. 2021. UDP-glucose and P2Y14 receptor amplify allergen-induced airway eosinophilia. J Clin Invest 131(7):e140709. (Story)