• Title/Summary/Keyword: 생체외 실험

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Effects of Compositions of Saponin Fraction from Korean Red Ginseng in the Relaxation of Rabbit and Rat Corpus Cavernosum (토끼와 흰쥐 음경해면체 이완작용에 미치는 홍삼사포닌 분획별 효과)

  • Choi Young Deuk;Park Jin Ah;Choi Hyung Ki;Nam Ki Yeul
    • Journal of Ginseng Research
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    • v.23 no.1 s.53
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    • pp.13-20
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    • 1999
  • We previously reported that Korean red ginseng (KRG) has a relaxation effect on the smooth muscles of corpus cavernosum via nitric oxide (NO) pathway and calcium and potassium channels. However, it is suggested that the active ingredients of KRG might be different depending on the sources of preparation, and there might be differences in actions for different compositions. We first investigated the composition of KRG saponins according to the extractions of the various sources of KRG, then with these extractions the relaxation effects were evaluated in vitro and hemodynamical in vivo using New Zealand white rabbit and rat corpus cavernosum. The total compositions of ginsenoside $(G-Rb_1,\;-Rb_2,\;-Rc,\;-Rd,\;G-Re,\;-Rf,\;-Rg_1)$ in fractionated KRG saponin designated as TS-1, TS-2, TS-3 were $41\%,\;40\%,\;and\;62\%,$ respectively, and the ratios of PD saponin and PT saponin (PD/PT) were 1,55, 1.72, 2.25, and 2.61, the values of which were statistically significant. In vitro studies using the rabbit corpus cavernosal muscle strips, the KRG saponin relaxed cavernosal strips in a dose-dependent manner, and same results were observed in in vivo studies, that KRG saponin increased the intracavernosal pressure in the rat. There was difference in the efficacy according to fractionation techniques. The differences in the total contents of ginsenosides did not affect relaxation, rather PT saponin content was statistically related to the degree of cavernosal relaxation, and this action presumed to be mediated by NO pathway and calcium and potassium channels. In conclusion, KRG exerts relaxation which is a key step in erection via combination of effects on NO system or calcium and potassium channels. The efficacy of this action is different to the sources of ginseng, which is affected by the different composition of ginsenosides $(G-Rb_1,\;-Rb_2,\;-Rc,\;-Rd,\;G-Re,\;-Rf,\;-Rg_1).$ Thus the further studies on the active ingredients such as minor ginsenosides and non-saponin components of red ginseng with maximum potency should be sought.

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Preparation and Characterization of PEG-PLA(PLGA) Micelles for Solubilization of Rosiglitazone (Rosiglitazone 가용화를 위한 PEG-PLA(PLGA) 고분자 미셀의 제조 및 특성분석)

  • Kim, Yon-Hwan;Im, Jeong-Hyuk;Min, Hyun-Su;Kim, Jun-Ki;Lee, Yong-Kyu;Park, Go-Eun;Cho, Kwang-Jae;Huh, Kang-Moo
    • Polymer(Korea)
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    • v.34 no.3
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    • pp.274-281
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    • 2010
  • In this study, PEG-PLA(or PLGA) amphiphilic di-block copolymers were synthesized by ring opening polymerization of D,L-lactide(or glycolide) and applied to polymeric micelle system for solubilization of a rosiglitazone as diabetes drug. The drug could be efficiently loaded into the polymer micelle by solid dispersion technique, and the drug-loaded micelles were characterized and evaluated as a drug delivery carrier by fluorescence spectrometer, DSC, and DLS measurements. The colloidal stability of drug loaded micelles in aqueous media could be enhanced by addition of 2-hydroxy-N-picolylnitinamide as a hydrotropic agent. The polymer micelles also showed biocompatible and nontoxic properties in vitro cell viability using MTT assay, and the drug loaded micelles were observed to be more effective than free drug for decreasing glucose in blood of rats.

Preparation and Release Behavior of Ipriflavone-Loaded PLGA Microsphere for Tissue Engineered Bone (이프리플라본을 함유한 생분해성 PLGA 미립구의 제조 및 조직공학적 골재생을 위한 영향평가)

  • So, Jung-Won;Jang, Ji-Wook;Kim, Soon-Hee;Kim, Geun-Ah;Choi, Jin-Hee;Rhee, John-M.;Son, Young-Suk;Min, Byoung-Hyun;Khang, Gil-Son
    • Polymer(Korea)
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    • v.33 no.1
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    • pp.26-32
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    • 2009
  • The aim of this research was to prepare microparticulate systems based on poly (lactide-co-glycolide)(PLGA) for the local release of ipriflavone in order to reduce bone loss. We developed the IP loaded PLGA microspheres using relatively simple oil-in-water(O/W) solvent evaporation method. HPLC was used to perform the in vitro release test of IP and morphology of cell attached on the micro-spheres was investigated using SEM. Cytotoxicity was assayed by cell counting kit-8 (CCK-8) test. Osteogenic differential cells were analyzed by ALP activity. Through RT-PCR analysis, we observed osteocalcin, ALP, and Type I collagen mRNA expression. The release of IP in vitro was more prolonged over 42 days and IP/PLGA microspheres showed the improvement on the cell proliferation, ALP activity and RT-PCR comparing with control (only PLGA). This initial research will be used to direct future work involved in developing this composite injectable bone tissue engineering system.

Identification and Characterization of Secreted Phosphoprotein 2 as a Novel Bioactive Protein for Myocardial Differentiation (심근세포로의 분화에 관여하는 새로운 생리활성 단백질 SPP2의 발굴)

  • Sejin Jeon
    • Journal of Life Science
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    • v.33 no.1
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    • pp.64-72
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    • 2023
  • Despite several advances in identification of cardiac transcription factors, there are still needs to find new bioactive molecules that promote cardiomyogenesis from stem cells to highly efficient myocardial differentiation. We analyzed Illumina expression microarray data of mouse embryonic stem cells (mESCs)-derived cardiomyocytes. 276 genes were upregulated (≥ 4fold) in mESCs-derived cardiomyocytes compared undifferentiated ESCs. Secreted phosphoprotein 2 (Spp2) is one of candidates and is known to inhibit bone morphogenetic protein 2 (BMP2) signal transduction as a pseudoreceptor for BMP2. However, its function in cardiomyogenesis is unknown. We confirmed that Spp2 expression increased during the differentiation into functional cardiomyocytes using mESCs, TC-1/Kh2 and E14. Interestingly, Spp2 secretion transiently increased 3 days after formation of embryoid bodies (EBs), indicating that the extracellular secretion of Spp2 is involved in the differentiation of ESCs into cardiomyocytes. To characterize Spp2, we performed experiments using the C2C12 mouse myoblast cell line, which has the property of shifting the differentiation pathway from myoblastic to osteoblastic by treatment with BMP2. Similar to the differentiation of ESCs, transcription of Spp2 increased as C2C12 myoblasts differentiated into myotubes. In particular, Spp2 secretion increased dramatically in the early stage of differentiation. Furthermore, treatment with Spp2-Flag recombinant protein promoted the differentiation of C2C12 myoblasts into myotubes. Taken together, we suggest a novel bioactive protein Spp2 that differentiates ESCs into cardiomyocytes. This may be useful for understanding the molecular pathways of cardiomyogenesis and for experimental or clinical promotion of stem cell therapy for ischemic heart diseases.

The Role of c-Jun N-terminal Kinase in the Radiation-Induced Lung Fibrosis (방사선에 의한 폐 섬유화증에서 c-Jun N-terminal Kinase(JNK)의 역할)

  • Uh, Soo-Taek;Hong, Ki-Young;Lee, Young-Mok;Kim, Ki-Up;Kim, Do-Jin;Moon, Seung-Hyuk;Kim, Yong-Hoon;Park, Choon-Sik;Yeom, Uk;Kim, Eun-Suk;Choi, Doo-Ho
    • Tuberculosis and Respiratory Diseases
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    • v.50 no.4
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    • pp.450-461
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    • 2001
  • Background : The underlying pathogenesis of radiation-induced lung fibrosis (RTLF) has not been very well defined. However, the role of TGF-$\beta$ in the generation of RTLF has been a major focus because there is an increase in the expression of both the TGF-${\beta}m$-RNA and its protein preceding RTLF lesions. The down stream signal after a TGF-$\beta$ stimulated lung fibrosis includes the activation of many mediators such as Smad and c-Jun N-terminal kinase (JNK) through TAK1. It is we hypothesized that JNK activation may play a pivotal role in RTLF pathogenesis through increased transcription of the fibrogenic cytokines. The present study evaluates JNK activity in alveolar macrophages after irradiation and the relationship between JNK activity and the amount of collagen in the lung tissues. Methods : C57BL/6 mice(20-25 gr, males) received chlorotetracycline(2g/L) in their drinking water 1 week prior to irradiation and continuously there after. The mice were irradiated once with 1400 cGy of $60CO{\gamma}$-ray over the whole chest. The cellular composition of the whole lung bronchoalveoalr lavage fluids(BALF), elastin expression in the lung tissues, the level of hydroxyproline in lung tissues, and an in vitro JNK assay was measured before irradiation and one, four, and eight weeks after irradiation (RT). Results : The volumes of BALF retrieved from instilled 4 mL of saline with 2% heparin were 3.7-3.8 mL for each group. The cell numbers were similar before($4.1{\times}10^4{\pm}0.5{\times}10^4/mL$) and 1 week($3.1{\times}10^4{\pm}0.5{\times}10^4/mL$) after RT. At four and eight weeks after RT, the cell number reached to $14.0{\times}10^4{\pm}1.5{\times}10^4mL$ and $10.0{\times}10^4{\pm}1.3{\times}10^4/mL$, respectively. There we no changes in the lymphocytes and neutrophils population observed in the BALF after RT. The H-E stain of the lung tissues did not show any structural and fibrotic change in the lung tissues at 4 and 8 weeks after RT. In addition, the amount of elastin and collagen were not different on Verhoeff staining of the lung tissues before RT to eight weeks after RT. The hydroxyproine content was measured with the left lung dissected from the left main bronchus. The lung were homogenized and hydrolyzed with 6 N Hel for 12 hours at $110^{\circ}C$ then measured as previously described. The content of hydroxyproline, standardized with a lung protein concentration, reached a peak 4 weeks after RT, and thereafter showed a plateau. AnIn vitro JNK assay using c-$Jun_{1-79}$-GST sepharose beads were performed with the alveolar macrophages obtained from the BAL. JNK activity was not detected prior to RT, However, the JNK activity increased from one week after RT and reached a peak four weeks after RT. Conclusion : JNK may be involved in the pathogenesis because the JNK activity showed similar pattern observed with the hydroxyproine content. However, it is necessary to clarify that the JNK increases the transcription of fibrogenic cyiokines through the transcription factor.

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The Change of Podocyte ${\beta}$-Catenin by Puromycin Aminonucleoside (Puromycin aminonucleoside 투여에 따른 사구체 족세포 ${\beta}$-catenin의 변화)

  • Choi, Ji-Young;Ahn, Eun-Mi;Park, Hye-Young;Shin, Jae-Il;Ha, Tae-Sun
    • Childhood Kidney Diseases
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    • v.15 no.2
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    • pp.138-145
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    • 2011
  • Purpose : To test whether the expression of ${\beta}$-catenin, a component of podocyte as a filtration molecule, would be altered by puromycin aminonucleoside (PAN) in the cultured podocyte in vitro. Methods : We cultured rat glomerular epithelial cells (GEpC) with various concentrations of PAN and examined the distribution of ${\beta}$-catenin by confocal microscope and measured the change of ${\beta}$-catenin expression by Western blotting and reverse transcriptase-polymerase chain reaction (RT-PCR). Results :We found that ${\beta}$-catenin relocalized from peripheral cytoplasm to inner cytoplasm, therefore, intercellular separations were seen in confluently cultured cells by high concentrations of PAN in immunofluorescence views. In Western blotting of GEpC, PAN ($50{\mu}g/mL$) decreased ${\beta}$-catenin expression by 34.9% at 24 hrs and 34.3% at 48 hrs, compared to those in without PAN condition (P<0.05). In RT-PCR, high concentrations ($50{\mu}g/mL$) of PAN also decreased ${\beta}$-catenin mRNA expression similar to protein suppression by 25.4% at 24 hrs and 51.8% at 48 hrs (P<0.05). Conclusion : Exposure of podocytes to PAN in vitro relocates ${\beta}$-catenin internally and reduces ${\beta}$-catenin mRNA and protein expression, which could explain the development of proteinuria in experimental PAN-induced nephropathy.

Cultural Conditions Affecting Gametophyte Propagation and Sporophyte Formation of Asplenium trichomanes L. (차꼬리고사리의 전엽체 증식 및 포자체 형성에 영향을 미치는 배양조건)

  • Jang, Bo Kook;Lee, Ki Cheol;Lee, Cheol Hee
    • Proceedings of the Plant Resources Society of Korea Conference
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    • 2018.10a
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    • pp.41-41
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    • 2018
  • 차꼬리고사리(Asplenium trichomanes L.)는 남방계식물로 제주도에 자생하는 것으로 알려져 있으며, 잎에 광택이 있고 총생하는 식물이다. 주로 실내 외 조경 및 분화소재로 이용되며, 한방에서는 철각봉미초라하여 뿌리를 포함한 전초를 이질, 임병, 만성질염, 월경불순 및 요통의 약재로 사용한다. 또한 식물구계학적 특정식물종 IV급, 적색자료목록 준위협 (NT)에 분류된 식물로 보호가 필요하다. 본 연구는 차꼬리고사리의 대량생산을 위한 기내전엽체 증식 및 기외포자체 형성조건을 구명하고자 수행되었다. 실험재료는 포자를 기내 발아시켜 전엽체를 획득한 다음 8주 간격으로 계대하면서 확보하였다. 전엽체의 증식과 생육에 적합한 배지를 비교하고자, 1/4, 1/2, 1, 2MS와 Knop배지를 조성하여 배양하였다. 배양은 전엽체 300mg을 메스로 균일하게 다지는 방법을 이용하였으며, 배양환경은 온도 $25{\pm}1.0^{\circ}C$, 광도 $30{\pm}1.0{\mu}mol{\cdot}m^{-2}{\cdot}s^{-1}$, 광주기 16/8h(light/dark)로 조절되었다. 실험결과, 1MS배지에 배양된 전엽체의 생체중이 4.3g으로 가장 많이 증가하였다. 형태형성발달도 하트형의 전엽체로 정상적으로 유도되었으며, 생식기관도 관찰되었다. 전엽체로부터 포자체의 형성을 유도하고자, 원예상토, 피트모스, 펄라이트 및 마사토의 비율을 5종류로 달리하여 혼합된 토양을 사각분($7.5{\times}7.5{\times}7.5cm$)에 충진하였다. 전엽체 1g과 증류수 25mL를 핸드블랜더로 10초간 분쇄하여 토양표면에 분주하는 실험방법을 사용하였다. 재배환경은 온도 $25{\pm}1.0^{\circ}C$, 광도 $43{\pm}2.0{\mu}mol{\cdot}m^{-2}{\cdot}s^{-1}$, 광주기 16/8h(light/dark)로 유지하면서 12주간 재배되었다. 실험결과, 원예상토 단용, 원예상토와 펄라이트가 2:1(v:v)로 혼합된 토양, 원예상토와 마사토가 2:1(v:v)로 혼합된 토양에서 각 31.7, 24.3, 19.3개의 포자체가 생산되었다. 한편 포자체의 생육은 원예상토 단용 토양에서 엽수, 엽장, 엽폭 등의 수치가 비교적 우수하였다. 따라서 차꼬리고사리의 전엽체는 MS배지에 배양하고 증식된 전엽체를 원예상토에 분주하여 포자체의 형성을 유도하는 것이 가장 효과적이었다.

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Detection of Carbonic Anhydrase in the Gills of Rainbow Trout (Oncorhynchus mykiss) (무지개 송어 rainbow trout, Oncorhynchus mykiss의 아가미에서의 carbonic anhydrase의 존재)

  • Kim, Soo Cheol;Choi, Kap Seong;Kim, Jung Woo;Choi, Myeong Rak;Han, Kyeong Ho;Lee, Won Kyo;Kho, Kang Hee
    • Journal of Life Science
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    • v.23 no.12
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    • pp.1557-1561
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    • 2013
  • Carbonic anhydrase isozymes are a widespread, zinc-containing metalloenzyme family. The enzyme catalyzes the reversible inter-conversion of $CO_2$ and $HCO_3$. This reaction is the main role played by CA enzymes in physiological conditions. This enzyme has been found in virtually all organisms, and at least 16 isozymes have been isolated in mammals. Unlike mammals, there is little information available regarding CA isozymes in the tissues of non-mammalian groups, such as fish. Carbonic anhydrase is very important in the osmotic and acid-base regulation in fish. It is well-known that the gills of fish play the most important role in acid-base relevant ion transfer, the transfer of $H^+$ and/or $HCO_3^-$, for the maintenance of systemic pH. Rainbow trout, Oncorhynchus mykiss, is the most important freshwater fish species in the aquaculture industry of Korea, with annual production increasing each year. In addition, environmental toxicology research has shown that rainbow trout is known to be the species that is most susceptible to environmental toxins. Consequently, carbonic anhydrase was detected in rainbow trout, Oncorhynchus mykiss. The isolated protein showed the specific band with a molecular weight of 30 kDa and pI of 7.0, and it was identified as being carbonic anhydrase. The immunohistochemical result demonstrated that the carbonic anhydrase was located in the epithelial cells of the gills.

Safety of Nano-sized Bee Pollen in both In-vitro and In-vivo Models (생체 외 및 생체 내 실험조건에서 나노화 벌 화분의 안전성 규명)

  • Pyeon, Hae-In;So, Soojeong;Bak, Jia;Lee, Seunghyun;Lee, Seungmin;Suh, Hwa-Jin;Lim, Je-Oh;Kim, Jung-Woo;Kim, Sun Youn;Lee, Se Ra;Lee, Yong Hyun;Chung, Il Kyung;Choi, Yun-Sik
    • Journal of Life Science
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    • v.28 no.5
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    • pp.605-614
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    • 2018
  • Bee pollen has an outer wall which is resistant to both acidic and basic solutions and even the digestive enzymes in the gastrointestinal tract. Therefore, the oral bioavailability of bee pollen is only 10-15%. A previous study reported on wet-grinding technology which increased the extraction of active ingredients from bee pollen by 11 times. This study was designed to investigate the safety of wet-ground bee pollen. First, a single dose of wet-ground bee pollen was tested in both rats and beagle dogs at dosages of 5, 10, and 20 g/kg and 1.5, 3, and 6 g/kg, respectively. In rats, compound-colored stools were found in those administered 10 g/kg or more of wet-ground bee pollen. In beagle dogs, 6 g/kg of wet-ground bee pollen induced diarrhea in one male for four hours. However, no obvious clinical signs were found through the end of the experiment in rats and beagle dogs. In addition, no histological abnormality was found in all animals. The data indicates that a single dose of up to 20 g/kg of wet-ground bee pollen is safe. Next, the genetic toxicity of nano-sized bee pollen was tested. This study employed a bacterial reverse mutation test, a micronucleus assay, and a chromosomal aberration assay. In the micronucleus assay, there was no genetic toxicity up to the dosage of 2 g/kg. There was also no genetic toxicity in the bacterial reverse mutation test and chromosomal aberration assay. This data provides important information in developing nano-sized bee pollen into more advanced functional foods and herbal medicines.

Effect of resin cement color on the color of commercially available zirconia crown (레진시멘트 색상이 상용 지르코니아 크라운의 색상에 미치는 영향)

  • Ha, Hyeon-Seung;Lim, Bum-Soon;Rhee, Sang-Hoon
    • Korean Journal of Dental Materials
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    • v.45 no.4
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    • pp.233-242
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    • 2018
  • The purpose of this study is to evaluate the effect of resin cement color on the color of commercially available zirconia crown. The zirconia and resin cements used for the experiment were $NuSmile^{(R)}$ ZR Zirconia LT Shade (LT), $RelyX^{TM}$ U200 TR, A2, and A3O (TR, A2, A3O). The disks of zirconia and resin cements with diameters of 5 mm and thicknesses of 1 mm were prepared. Five disks were made for each specimen. The CIE $L^*a^*b^*$ values of zirconia, resin cements and the combinations thereof were measured on black and white backgrounds, respectively, using a spectrophotometer. The color effect of resin cement on the color of the zirconia crown was evaluated by calculating translucency parameter (TP), contrast ratio (CR), and color differences (${\Delta}E{^*}_{ab}$) based on the measured CIE $L^*a^*b^*$ values. The statistical significances were verified by one-way ANOVA and the Tukey-multiple comparisons tests. As a result, the TP and CR values were decreased (p<0.05) and increased, respectively, in the combination of zirconia and resin cement disks compared to zirconia disk per se. When using the black background, the ${\Delta}E{^*}_{ab}$ values between zirconia and the combination of the zirconia and three resin cement disks were imperceptible level. The A3O showed the lowest ${\Delta}E{^*}_{ab}$ value among three resin cements. When using the white background, the ${\Delta}E{^*}_{ab}$ values between zirconia and the combination of zirconia and TR resin cement (LT/TR) disks showed acceptable level. However, the ${\Delta}E{^*}_{ab}$ values between zirconia and the combination of zirconia and A2 resin cement (LT/A2) disks showed unacceptable level. Meanwhile, the ${\Delta}E{^*}_{ab}$ values between zirconia and the combination of zirconia and A3O resin cement (LT/A3O) disks showed perceptible but acceptable level. Within the limits of this study, the colors of resin cements did not cause unacceptable color changes of zirconia except the combination of LT/A2 on the white background. The resin cement that gave the least color changes to zirconia was A3O. This means that the resin cement A3O is recommended to use for minimizing color changes when cementing commercially available zirconia crown to tooth.