Electronic cigarettes (e‐cigarettes) are nicotine delivery devices advertised as a healthier alternative to conventional tobacco products, but their rapid rise in popularity outpaces research on potential health consequences. As conventional tobacco use is a risk factor for osteoporosis, this study examines whether exposure to electronic liquid (e‐liquid) used in e‐cigarettes affects bone‐forming osteoblasts. Human MG‐63 and Saos‐2 osteoblast‐like cells were treated for 48 hours with 0.004%‐4.0% dilutions of commercially available e‐liquids of various flavors with or without nicotine. Changes in cell viability and key osteoblast markers, runt‐related transcription factor 2 and Col1a1, were assessed. With all e‐liquids tested, cell viability decreased in a dose‐dependent manner, which was least pronounced in flavorless e‐liquids, most pronounced in cinnamon‐flavored e‐liquids and occurred independently of nicotine. Col1a1, but not runt‐related transcription factor 2, mRNA expression was upregulated in response to coffee‐flavored and fruit‐flavored e‐liquids. Cells treated with a non‐cytotoxic concentration of fruit‐flavored Mango Blast e‐liquid with or without nicotine showed significantly increased collagen type I protein expression compared to culture medium only. We conclude that the degree of osteotoxicity is flavor‐dependent and occurs independently of nicotine and that flavored e‐liquids reveal collagen type I as a potential target in osteoblasts. This study elucidates potential consequences of e‐cigarette use in bone.

The heavy metal cadmium is a widespread environmental contaminant that has gained public attention due to the global increase in cadmium-containing electronic waste. Human exposure to cadmium is linked to the pathogenesis of osteoporosis. We previously reported cadmium induces apoptosis and decreases alkaline phosphatase mRNA expression via extracellular signal-regulated protein kinase (ERK) activation in Saos-2 bone-forming osteoblasts. This study examines the mechanisms of cadmium-induced osteotoxicity by investigating roles of Ca+2/calmodulin-dependent protein kinase (CAMK) pathways. Saos-2 or MG-63 cells were treated for 24 or 48 h with 5 μM CdCl2 alone or in combination with calmodulin-dependent phosphodiesterase (PDE) inhibitor CGS-9343β; calmodulin-dependent kinase kinase (CAMKK) inhibitor STO-609; or calmodulin-dependent kinase II (CAMKII) inhibitor KN-93. CGS-9343β protected against cadmium-induced toxicity and attenuated ERK activation; STO-609 enhanced toxicity and exacerbated ERK activation, whereas KN-93 had no detectable effect on cadmium-induced toxicity. Furthermore, CGS-9343β co-treatment attenuated cadmium-induced apoptosis; but CGS-9343β did not recover cadmium-induced decrease in ALP activity. The major findings suggest the calmodulin-dependent PDE pathway facilitates cadmium-induced ERK activation leading to apoptosis, whereas the CAMKK pathway plays a protective role against cadmium-induced osteotoxicity via ERK signaling. This research distinguishes itself by identifying pleiotropic roles for CAMK pathways in mediating cadmium’s toxicity in osteoblasts.

Cohesin rings interact with DNA and modulate the expression of thousands of genes. NIPBL loads cohesin onto chromosomes, and WAPL takes it off. Haploinsufficiency for NIPBL causes a developmental disorder, Cornelia de Lange syndrome (CdLS), that is modeled by Nipbl +/− mice. Mutations in WAPL have not been shown to cause disease or gene expression changes in mammals. Here, we show dysregulation of >1000 genes in Wapl Δ /+ embryonic mouse brain. The patterns of dysregulation are highly similar in Wapl and Nipbl heterozygotes, suggesting that Wapl mutations may also cause human disease. Since WAPL and NIPBL have opposite effects on cohesin’s association with DNA, we asked whether decreasing Wapl dosage could correct phenotypes seen in Nipbl +/− mice. Gene expression and embryonic growth are partially corrected, but perinatal lethality is not. Our data are consistent with the view that cohesin dynamics play a key role in regulating gene expression.

Maternal loss of imprinting (LOI) at the H19/IGF2 locus results in biallelic IGF2 and reduced H19 expression and is associated with Beckwith-Wiedemann syndrome (BWS). We use mouse models for LOI to understand the relative importance of Igf2 and H19 mis-expression in BWS phenotypes. Here we focus on cardiovascular phenotypes and show that neonatal cardiomegaly is exclusively dependent on increased Igf2. Circulating IGF2 binds cardiomyocyte receptors to hyperactivate mTOR signaling, resulting in cellular hyperplasia and hypertrophy. These Igf2-dependent phenotypes are transient: cardiac size returns to normal once Igf2 expression is suppressed postnatally. However, reduced H19 expression is sufficient to cause progressive heart pathologies including fibrosis and reduced ventricular function. In the heart, H19 expression is primarily in endothelial cells (ECs) and regulates EC differentiation both, in vivo and in vitro. Finally, we establish novel mouse models to show that cardiac phenotypes depend on H19 lncRNA interactions with Mirlet7 microRNAs.