A higher aspect ratio enhanced bioaccumulation and altered immune responses due to intravenously-injected aluminum oxide nanoparticles

Aluminum oxide nanoparticles (AlO NP) have been widely utilized in a variety of areas, including in the optical, biomedical and electronic fields and in the overall development of nanotechnologies. However, their toxicological profiles are still not fully developed. This study compared the distribution and immunotoxicity of two rod-types of AlO NP. As reported previously, the two types of AlO NP had different aspect ratios (long-type: 6.2?±?0.6, short-type: 2.1?±?0.4), but the size and surface charge were very similar. On Day 14 after a single intravenous (IV) injection (1.25 or 5?mg/kg), both AlO NP accumulated primarily in the liver and spleen and altered the levels of redox response-related elements. The accumulated level was higher in mice exposed to the long-type AlO NP compared to the short-type. Additionally, it was noted that the levels of IL-1β, IL-8 and MCP-1 were enhanced in the blood of mice exposed to both types of AlO NP and the percentages of neutrophils and monocytes among all white blood cells were increased only in mice injected with the long-type AlO NP (5?mg/kg). In addition, as compared to the control, co-expression of CD80 and CD86 (necessary for antigen presentation) on splenocytes together with a decreased expression of chemotaxis-related marker (CD195) was attenuated by exposure to the AlO NP, especially the long-type. Taken together, the data suggest that accumulation following a single IV injection with rod-types of AlO NP is strengthened by a high aspect ratio and, subsequently, this accumulation has the potential to influence immune functions in an exposed host.     

Low noise bipolar photodiode array protein chip based on on-chip bioassay for the detection of <Emphasis Type="Italic">E. coli</Emphasis> O157:H7

A bipolar photodiode array (PDA) protein chip is presented for the detection of E. coli O157:H7. Through unique design of the bipolar PDA microchip, the device was able to detect E. coli O157:H7 directly on the surface of the bipolar PDA. The bipolar PDA microchip maintained low noise level in the entire process of on-chip protein assay and demonstrated high performance of analog signal processing. At every reaction step of the on-chip bioassay, stability of wet photodiode detection elements was confirmed by monitoring the variance of their photosignals with respect to the irradiated red beam. The background signal represented less than 1.8% variance with respect to maximum signal of photodiode detection elements. As a result of using the on-chip bioassay, any complicated optical alignment and components could be removed in the constructed protein chip. This protein chip enables direct optical detection of E. coli O157:H7 eliminating the need of conventional expensive microplate reader that is incompatible with size of sampling platform of protein chip. The independence of the constructed protein chip on conventional microplate reader can contribute greatly to further miniaturization of protein chip and field usable lab-on-a-chip.     

Structure of the protein core of the glypican Dally-like and localization of a region important for hedgehog signaling

Glypicans are heparan sulfate proteoglycans that modulate the signaling of multiple growth factors active during animal development, and loss of glypican function is associated with widespread developmental abnormalities. Glypicans consist of a conserved, approximately 45-kDa N-terminal protein core region followed by a stalk region that is tethered to the cell membrane by a glycosyl-phosphatidylinositol anchor. The stalk regions are predicted to be random coil but contain a variable number of attachment sites for heparan sulfate chains. Both the N-terminal protein core and the heparan sulfate attachments are important for glypican function. We report here the 2.4-Å crystal structure of the N-terminal protein core region of the Drosophila glypican Dally-like (Dlp). This structure reveals an elongated, α-helical fold for glypican core regions that does not appear homologous to any known structure. The Dlp core protein is required for normal responsiveness to Hedgehog (Hh) signals, and we identify a localized region on the Dlp surface important for mediating its function in Hh signaling. Purified Dlp protein core does not, however, interact appreciably with either Hh or an Hh:Ihog complex.Glypicans are heparan sulfate proteoglycans (HSPGs) that consist of an approximately 450 amino acid N-terminal protein domain followed by an approximately 100 amino acid stalk region that is attached to the outer cell membrane via a glycosyl-phosphatidylinositol anchor (1). The N-terminal domain of most glypicans is proteolytically processed by a furin-like convertase to produce two chains that remain connected by disulfide bonds (2). This processing appears required for some but not all glypican activity (2, 3). The stalk regions of glypicans are predicted to be largely random coil and typically contain 1–5 heparan sulfate attachment sites (1, 4). Six glypicans are present in humans and mice (glypican-1, -2, -3, -4, -5, and -6); two are present in Drosophila [Dally and Dally-like (Dlp)] (1). Based on sequence similarity, glypicans assort into two subfamilies with glypican-1, -2, -4, -6, and Dlp in one family and glypican-3, -5, and Dally in another (1).Glypicans are active in development in both vertebrates and invertebrates. Loss of Dally in fruit flies results in defects in brain, eye, wings, antennae, and genitalia (5). Loss of glypican-3 in humans is responsible for Simpson–Golabi–Behmel overgrowth syndrome, in which widespread visceral and skeletal abnormalities are present along with a predisposition to tumor formation (6). Loss of glypican-6 has recently been shown to cause omodysplasia, a genetic disorder characterized by variable heart defects, cognitive delay, skeletal and facial abnormalities, and shortness of stature (7). Much of the function of glypicans is attributable to modulation of signaling by several heparin-binding growth factors active during development including members of the fibroblast growth factor, Hedgehog (Hh), Wnt, and transforming growth factor-β families (8–15). Each of these factors functions as a morphogen to elicit distinct concentration-dependent responses within target cells, and glypicans have been shown to be required both for normal response to these factors as well as to establish their proper distribution (9, 10, 12, 16–21). The heparan sulfate attachments of glypicans are clearly important for mediating interactions with these growth factors and downstream signaling components (22, 23), but recent work has demonstrated a role for the N-terminal protein domain, which lacks heparan sulfate modifications, in mediating responsiveness to at least Wnt and Hh signals (23–26).Curiously, glypicans appear able to play both positive and negative roles in mediating Hh signaling. The protein region of Dally-like contributes positively to Drosophila Hh responsiveness, and the developmental defects in omodysplasia, particularly the bone growth defects, are suggestive of a positive role for glypican-6 function in response to Indian hedgehog (7). Notably, glypican-4 and glypican-6 are most similar to Dlp (vs. Dally) and complement Dlp function in a Drosophila cultured cell-based Hh signaling assay (25). In contrast, the protein region of glypican-3, which is more similar to Dally than Dally-like, is a negative regulator of Hh responsiveness in the mouse (24, 25, 27, 28). Based on sequence homology and functional phenotypes, it has thus been speculated that the two major subfamilies of glypicans have evolved opposing activities in Hh signal responsiveness (25).To investigate the molecular basis for glypican function, we have undertaken structural and functional characterization of the N-terminal protein domain of Dlp and report here its 2.4-Å crystal structure. We show that the N-terminal protein domains of glypicans adopt an elongated α-helical structure with no evident homology to any known structure. We have used structure-guided mutagenesis to identify a localized region on the Dlp surface important for the ability of Dlp to mediate Hh signal response. These results are most consistent with Dlp functioning as a binding protein in Hh signaling, but we are unable to detect high-affinity interactions between Dlp and either Hh or an Hh:Ihog complex. These results establish a molecular basis for mapping and comparing functional regions of different glypicans.