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1 SUPPLEMENTARY INFORMATION doi: 1.138/nnno Sili nd titnium dioxide nnoprtiles use pregnny omplitions in mie Kohei Ymshit, Ysuo Yoshiok, Kzum Higshisk, Kzuy Mimur, Yuki Morishit, Mstoshi Nozki, Tokuyuki Yoshid, Toshinou Ogur, Hiromi Neshi, Kzuy Ngno, Ysuhiro Ae, Hruhiko Kmd, Youko Monoe, Tkyoshi Imzw, Hise Aoshim, Yuihi Kwi, Tdnori Myumi, Shin-ihi Tsunod, Norio Itoh, Tomoki Yoshikw, Itru Yngihr, Shigeru Sito, Ysuo Tsutsumi nture nnotehnology Mmilln Pulishers Limited. All rights reserved.
2 supplementry informtion doi: 1.138/nnno Methods Prtile properties. Prtile suspensions were diluted to.1 mg ml -1 (7, 3, msp1, nno-tio 2, fullerene, 7-C nd 7-N) with wter. The size of prtiles nd the zet potentil were mesured using Zetsizer Nno-ZS (Mlvern Instruments, UK). The men size nd the size distriution of prtiles were mesured y dynmi light sttering. The zet potentil ws mesured y lser doppler eletrophoresis. The shpe of sili prtiles nd nno-tio 2 were oserved using TEM. Prior to TEM nlysis, 7, 7-C nd 7-N were stined with 2% urnium ette. Smples were pled on 2-mesh ron-oted opper TEM grids (Nissin EM. Tokyo, Jpn) nd oserved y TEM. Fluoresent dissoition. One hundred mg ml -1 DY-676-leled sili prtiles in PBS were inuted for 24 hours t 37 C. After entrifugtion for 2 min t 12 rpm, the superntnt ontining dissoited DY-676 ws removed nd the resulting preipittion ws re-dissolved in 1 ml of PBS. Fluoresene intensity of re-dissolved solution ws mesured using TriStr LB 941 spetrophotometer (Berthold Tehnologies, Germny). Stility of the fluoresent lel ws evluted s reltive rtes (% of ontrol). Plsm iohemil nlysis. Liver funtion ws evluted y the plsm level of AST nd ALT. Nephrotoxiity ws evluted y the plsm level of lood BUN. These mrkers were ssyed y iohemil utonlyzer, FUJI DRI-CHEM 7 (Fujifilm, Tokyo, Jpn). The level of sflt-1 in plsm ws mesured with n enzyme-linked immunosorent ssy (ELISA) kit (R&D Systems, Minnepolis, MN) ording to the mnufturer s instrutions. Blood pressure nlysis. Mie were restrined for 1 min in plsti holder. Systoli lood pressure ws determined using progrmmed sphygmomnometer (MMK-11, Muromhi Kiki, Tokyo, Jpn). Eh reported vlue represents the verge of three reordings tken t 1-min intervls. Hemtology nlysis. The numer of white lood ells, lymphoytes, monoytes, grnuloytes, red lood ells, nd pltelets nd the mount of hemogloin in whole lood were mesured y n utonlyzer (VetSn HMII Hemtology System; Axis, Union City, CA). 2 nture nnotehnology Mmilln Pulishers Limited. All rights reserved.
3 doi: 1.138/nnno supplementry informtion 7 3 msp1 d 7-C e 7-N f TiO 2 g 3 nm h 3 nm i nm j 3 nm k 3 nm l 1 nm 1 nm nm 1 nm 1 nm 1 nm 3 nm m n Intensity (%) Stility of the fluoresent lel (%) Size (nm) msp1 TiO2 Fullerene C 6 7-C 7-N Supplementry Figure S1. Properties of sili nd titnium dioxide prtiles. TEM imges of sili nd titnium dioxide prtiles used in this study: 7 (,g), 3 (,h), msp1 (,i), 7-C (d,j), 7-N (e,k), nd TiO 2 (f,l). Size distriution of 7, 3, msp1, 7-C, 7-N nd TiO 2 in diluted wter ws mesured y dynmi light sttering method (m). Dissoition of fluoresent DY-676 from the sili prtiles in PBS ws evluted fter 24 hours of in vitro inution t 37 C (n). All dt represent mens ± SD (n = 3). nture nnotehnology Mmilln Pulishers Limited. All rights reserved.
4 supplementry informtion doi: 1.138/nnno Supplementry Figure S2. Summry of the physiohemil properties of nnoprtiles. Prtile Primry prtile size (nm) Hydrodynmi dimeter (nm) Men zet potentil (mv) msp TiO Fullerene C C N Informtion from tehnil dtsheet of produts. 4 nture nnotehnology Mmilln Pulishers Limited. All rights reserved.
5 doi: 1.138/nnno supplementry informtion AST (U/L) ALT (U/L) 4 2 BUN (mg/dl) Supplementry Figure S3. Chnges in tissue dmge mrkers in mouse plsm. Pregnnt BALB/ mie were treted with 1 l of.8 mg mouse -1 sili prtiles (7, 3, or msp1) or PBS (ontrol) intrvenously vi til vein on two onseutive dys, t GD16 nd GD17. At GD18, AST (), ALT (), nd BUN () levels in plsm were evluted (n = 7). All dt represent mens ± SEM. 3 7 nture nnotehnology Mmilln Pulishers Limited. All rights reserved.
6 supplementry informtion doi: 1.138/nnno Blood pressure (mmhg) msp Dys post oitus Hert rte (et/minute) msp Dys post oitus Supplementry Figure S4. Blood pressure nd hert rte. Pregnnt BALB/ mie were treted with 1 l of.8 mg mouse -1 sili prtiles (7, 3, or msp1) or PBS (ontrol) intrvenously vi til vein on two onseutive dys, t GD16 nd GD17. Blood pressure () nd hert rte () were reorded t GD14 18 (n = ). Eh of estimtions shown in (,) ws the verge of five reordings tken t 1-min intervls. All dt re presented s mens ± SEM. 6 nture nnotehnology Mmilln Pulishers Limited. All rights reserved.
7 doi: 1.138/nnno supplementry informtion 8 2. White lood ell ( 1 3 ells/mm 3 ) Lymphoyte ( 1 3 ells/mm 3 ) Grnuloyte ( 1 3 ells/mm 3 ) d Monoyte ( 1 3 ells/mm 3 ) e Red lood ell ( 1 6 ells/mm 3 ) g f Hemogloin (g/dl) Pltelet ( 1 3 ells/mm 3 ) Supplementry Figure S. Chnges in hemtologil prmeters in mouse lood. Pregnnt BALB/ mie were treted with 1 l of.8 mg mouse -1 sili prtiles (7, 3, or msp1) or PBS (ontrol) intrvenously vi til vein on two onseutive dys, t GD16 nd GD17. At GD18, the numers of white lood ells (), lymphoytes (), grnuloytes (), monoytes (d), red lood ells (e), nd pltelets (g), s well s the mount of hemogloin (f), were evluted (n = 11 2). All dt represent mens ± SEM. ( P <.1 y ANOVA) nture nnotehnology Mmilln Pulishers Limited. All rights reserved.
8 supplementry informtion doi: 1.138/nnno Mternl ody weight (g) (.2 mg mouse -1 ) 7 (.4 mg mouse -1 ) 7 (.8 mg mouse -1 ) Dys post oitus Uterine weight (g) mg mouse -1.4 mg mouse -1.8 mg mouse -1 d Fetl resorption rte (%) mg mouse -1.4 mg mouse -1.8 mg mouse -1 Fetl weight (g) mg mouse -1.4 mg mouse -1.8 mg mouse -1 Supplementry Figure S6. Dose-dependent toxiity of 7. Pregnnt BALB/ mie were treted with 1 l of.2,.4, or.8 mg mouse -1 7 or PBS (ontrol) intrvenously vi til vein on two onseutive dys, t GD16 nd GD17. () Chnges in mternl ody weights. Mternl ody weights were evluted dily (n = 24). Sttistilly signifint differene from ontrol mie (P <.1 y ANOVA). ( d) Anlysis of pregnny omplitions s funtion of injeted dose of 7. At GD18, uterine weights (), fetl resorption rtes (), nd fetl weights (d) were evluted (,; n = 24, d; n = 4 212). All dt represent mens ± SEM. ( P <., P <.1 y ANOVA) 8 nture nnotehnology Mmilln Pulishers Limited. All rights reserved.
9 doi: 1.138/nnno supplementry informtion mg mouse -1.4 mg mouse -1.8 mg mouse d e 2 TiO2 C6 Heprin (-) Heprin (+) 7 7-C 7-N Supplementry Figure S7. sflt-1 levels in plsm. Pregnnt BALB/ mie were treted intrvenously vi til vein on two onseutive dys, t GD16 nd GD17 with the following: 7, 3, or msp1 (.8 mg mouse -1 ) (); 7 (.2,.4, or.8 mg mouse -1 ) (); fullerene C 6 or nno-tio 2 (.8 mg mouse -1 ) (); 7, 7-C, or 7-N (.8 mg mouse -1 ) (d); 7 (.8 mg mouse -1 ) with or without tretment with heprin (e). At GD18, the levels of sflt-1 in eh set of treted mie were mesured y ELISA (n = 8). All dt represent mens ± SEM. ( P <., P <.1 y ANOVA ( d) nd Student s t-tests (e)) nture nnotehnology Mmilln Pulishers Limited. All rights reserved.
RESEARCH ARTICLE. Supplemental Figure 5
11.5 2 2 11. RESEARCH ARTICLE RBC ( 1 12 /L) 1.5 1. 9.5 PLT ( 1 9 /L) 1 16 14 HGB (g/l) 19 1 17 16 9. 12 4 4 46 Cellulr & Moleulr Immunology dvne online pulition, PCV (%) 44 MCV (fl) 46 44 ; doi:1.13/mi.214.16
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