• Journal of Veterinary Science
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• v.23 no.2
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• pp.40.1-40.13
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• 2022

Background: Somatic cell nuclear transfer (SCNT) is used widely in cloning, stem cell research, and regenerative medicine. The type of donor cells is a key factor affecting the SCNT efficiency. Objectives: This study examined whether urine-derived somatic cells could be used as donors for SCNT in pigs. Methods: The viability of cells isolated from urine was assessed using trypan blue and propidium iodide staining. The H3K9me3/H3K27me3 level of the cells was analyzed by immunofluorescence. The in vitro developmental ability of SCNT embryos was evaluated by the blastocyst rate and the expression levels of the core pluripotency factor. Blastocyst cell apoptosis was examined using a terminal deoxynucleotidyl transferase dUTP nick end-labeling assay. The in vivo developmental ability of SCNT embryos was evaluated after embryo transfer. Results: Most sow urine-derived cells were viable and could be cultured and propagated easily. On the other hand, most of the somatic cells isolated from the boar urine exhibited poor cellular activity. The in vitro development efficiency between the embryos produced by SCNT using porcine embryonic fibroblasts (PEFs) and urine-derived cells were similar. Moreover, The H3K9me3 in SCNT embryos produced from sow urine-derived cells and PEFs at the four-cell stage showed similar intensity. The levels of Oct4, Nanog, and Sox2 expression in blastocysts were similar in the two groups. Furthermore, there is a similar apoptotic level of cloned embryos produced by the two types of cells. Finally, the full-term development ability of the cloned embryos was evaluated, and the cloned fetuses from the urine-derived cells showed absorption. Conclusions: Sow urine-derived cells could be used to produce SCNT embryos.

• Nuclear Medicine and Molecular Imaging
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• v.41 no.3
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• pp.241-246
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• 2007

Purpose: $[^{11}C]6-OH-BTA-1$ ([N-methyl-$^{11}C$]2-(4'-methylaminophenyl)-6-hydroxybenzothiazole, 1), a -amyloid imaging agent for the diagnosis of Alzheimer's disease in PET, can be labeled with higher yield by a simple loop method. During the synthesis of $[^{11}C]1$, we found the formation of by-products in various solvents, e.g., methylethylketone (MEK), cyclohexanone (CHO), diethylketone (DEK), and dimethylformamide (DMF). Materials and Methods: In Automated radiosynthesis module, 1 mg of 4-aminophenyl-6-hydroxybenzothiazole (4) in 100 l of each solvent was reacted with $[^{11}C]methyl$ triflate in HPLC loop at room temperature (RT). The reaction mixture was separated by semi-preparative HPLC. Aliquots eluted at 14.4, 16.3 and 17.6 min were collected and analyzed by analytical HPLC and LC/MS spectrometer. Results: The labeling efficiencies of $[^{11}C]1$ were $86.0{\pm}5.5%$, $59.7{\pm}2.4%$, $29.9{\pm}1.8%$, and $7.6{\pm}0.5%$ in MEK, CHO, DEK and DMF, respectively. The LC/MS spectra of three products eluted at 14.4, 16.3 and 17.6 mins showed m/z peaks at 257.3 (M+1), 257.3 (M+1) and 271.3 (M+1), respectively, indicating their structures as 1, 2-(4'-aminophenyl)-6-methoxybenzothiazole (2) and by-product (3), respectively. Ratios of labeling efficiencies for the three products $([^{11}C]1:[^{11}C]2:[^{11}C]3)$ were $86.0{\pm}5.5%:5.0{\pm}3.4%:1.5{\pm}1.3%$ in MEK, $59.7{\pm}2.4%:4.7{\pm}3.2%:1.3{\pm}0.5%$ in CHO, $9.9{\pm}1.8%:2.0{\pm}0.7%:0.3{\pm}0.1%$ in DEK and $7.6{\pm}0.5%:0.0%:0.0%$ in DMF, respectively. Conclusion: The labeling efficiency of $[^{11}C]1$ was the highest when MEK was used as a reaction solvent. As results of mass spectrometry, 1 and 2 were conformed. 3 was presumed.

• The Korean Journal of Nuclear Medicine
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• v.33 no.6
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• pp.527-536
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• 1999

Purpose: Radiotracers that bind to the central benzodiazepine receptor are useful for the investigation of various neurological and psychiatric diseases. [C-11]Flumazenil, a benzodiazepine antagonist, is the most widely used radioligand for central benzodiazepine receptor imaging by PET. We synthesized 3-(2-[F-18]fluoro)flumazenil, a new fluorine-18 ($t_{1/2}$= 110 min) labeled analogue of benzodiazepine receptor imaging agent, and evaluated in vivo for biodistribution in mice. Materials and Methods: Flumazenil (Ro 15-1788) was synthesized by a modification of the reported method. Precursor of 3-(2-[F-18]fluoro)flumazenil, the tosylated flumazenil derivative was prepared by the tosylation of the ethyl ester by ditosylethane. [F-18] labeling of tosyl substitued flumazenil precursor was performed by adding F-18 ion at $85^{\circ}C$ in the hot ceil for 20 min. The reaction mixture was trapped by C18 cartridge, washed with 10% ethanol, and eluted by 40% ethanol. Bidistribution in mice was determined after intravenous injection. Results: The total chemical yield of tosylated flumazenil derivative was ${\sim}40%$. The efficiency of labeling 3-(2-[F-18]fluoro)flumazenil was 66% with a total synthesis time of 50 min. Brain uptakes of 3-(2-[F-18]fluoro)flumazenil at 10, 30, 60 min after injection, were $2.5{\pm}0.37,\;2.2{\pm}0.26,\;2.1{\pm}0.11$ and blood activities were $3.7{\pm}0.43,\;3.3{\pm}0.07,\;3.3{\pm}0.09%ID/g$, respectively. Conclusion: We synthesized a tosylated flumazenil derivative which was successfully labeled with no-carrier-added F-18 by nucleophilic substitution.

• The Korean Journal of Nuclear Medicine
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• v.32 no.1
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• pp.81-88
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• 1998

The purpose of this study was to examine the radiolabeling and biodistribution of $^{188}Re(V)$-DMSA as a therapeutic cancer radiopharmaceutical. We made a DMSA kit($NaHCO_3$ 1.5 mg, meso-2,3-dimercaptosuccinic acid 1.0 mg, L(+)-ascorbic acid 0.7 mg, $SnCl_2{\cdot}2H_2O$ 0.34 mg, pH 2.9) for labeling with $^{188}Re$. In this kit, $^{188}ReO_4{^-}$ 5 mCi/2 ml added and boiled at $100^{\circ}C$ for 3 hr in water bath. The final pH adjusted to 7.5 with 7% $NaHCO_3$ solution. We checked the labelling efficacy with TLC-SG(n-butanol : acetic acid : $H_2O$ = 3 : 2 : 3) and examined the stability both in room temperature and in serum at $37^{\circ}C$. Biodistribution(1, 3, 13, 24, 48 hr) of $^{188}Re(V)$-DMSA compound was evaluated in Sarcoma 180 tumor-bearing mice. Each labeling efficiency and stability at room temperature for 48 hours was over 98% and 95%, respectively. The stability in serum were 82%(6 hr) and 85%(48 hr). Tumor uptake of $^{188}Re(V)$-DMSA in Sarcoma 180-bearing mice were $0.66{\pm}0.15%$(1 hr), $0.51{\pm}0.10%$(3 hr), $0.19{\pm}0.05%$(24 hr) and $0.13{\pm}0.02%$(48 hr). These result are consistent with those of $^{99m}Tc(V)$-DMSA which were reported previously. In conclusion, $^{188}Re(V)$-DMSA may be a useful therapeutic radiopharmaceutical for treating some cancers and metastatic bone lesion.

• Journal of Chest Surgery
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• v.35 no.3
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• pp.171-176
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• 2002

Background: Blood-foreign interaction cause activation of coagulation and inflammatory process that may lead to multiorgan dysfunction and determine the surgical outcomes. Of the methods for assessing the biocompatibility, the platelet adhesion study is considered as the most valuable evaluation step in blood-foreign interaction. As the most studies have used in-vitro or ex-vivo conditions, we have developed a technique of quantification for platelet adhesion on the blood contact surface by using in-vivo injection of radioactive platelets. Material and Method: A coupled bypass circuit was designed to connect the proximal and descending thoracic aorta in 6 piglets(20∼25 Kg). One side of the circuit tube was consisted of a heparin coated PVC tube(10mm in ID, n=6, Experimental group), and the other, a non-heparin coated PVC tube(10mm in ID, n=6, Control group). After cannulation, the blood was circulated through the circuit for 2 hours. Platelet concentrate was prepared from homologous pig blood 24 hours before the experiment. The platelet concentrate was incubated with Tc-99m-HMPAO for 30 min and then centrifuged for 10 min. The supernatant was discarded and the radio-labeling efficacy was measured. The radio-labeled platelet concentrate was mixed with the autologous plasma to make the volume 5 ml, and the mixture was injected intravenously into the experimental animal. After 2 hour circulation, 5 pieces of the specimen(10mm in length each) were obtained from each PVC tube. The radioisotopes were counted with a gamma counter(Cobra ll, Packard, USA), and the ratio of radioisotope count was compared between the control and experimental group. Result: The radioisotope count number was 537.3221.1 Ci/min in the control group and 311.1 184.5 Ci/min in the experimental group(p=0.0104). The ratio between the groups was 1 to 0.58 (p=0.004). Conclusion: In vivo quantification using technetium-99m-HMPAO labeled platelets is simple and reproducible in evaluating platelet adhesion on a foreign surface. We suggest this technique to be a useful tool for blood compatibility test.

• The Korean Journal of Nuclear Medicine
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• v.39 no.1
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• pp.26-33
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• 2005

Purpose: The purpose of this study is to evaluate migration of technetium-99m hexamethylpropylene amine oxime ($^{99m}Tc$-HMPAO) labeled immature and mature dendritic cells (DC) in the mouse. Methods: DC were collected from bone marrow (BM) of tibiae and femurs of mice. Immature and mature DC from BM cells were radiolabeled with $^{99m}Tc$-HMPAO. To evaluate the functional and phenotypic changes of DC from radiolabeling, the allogeneic mixed lymphocyte reaction (MLR) and fluorescence-activated cell sorting (FACS) analysis were performed before and after labeling with $^{99m}Tc$-HMPAO. Migration of intravenously injected DC (iv-DC) was assessed by serial gamma camera images of mice with or without subcutaneous tumor. Percent injected dose per gram (%ID/g) was calculated in lungs, liver, spleen, kidneys, and tumor through dissection of each mice after 24 hours of injection. Results: Labeling efficiency of immature and mature DC were $60.4{\pm}5.4%\;and\;61.8{\pm}6.7%$, respectively. Iv-DC initially appeared in the lungs, then redistributed mainly to liver and spleen. Migration of mature DC to spleen was significantly higher than that of immature DC ($38.3{\pm}4.0%\;vs.\;32.2{\pm}4.1%$ in control group, $40.4{\pm}4.1%\;vs.\;35.9{\pm}3.8%$ in tumor group; p<0.05). Migration to tumor was also significantly higher in mature DC than in immature DC ($2.4{\pm}0.3%\;vs\;1.7{\pm}0.2%$; p=0.034). Conclusion: Assessment of migration pattern of DC in mice was possible using $^{99m}Tc$-HMPAO labeled immature and mature DC. Migration of mature DC to spleen and tumor was higher than that of immature DC when they were i.v. injected.

• The Korean Journal of Nuclear Medicine
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• v.39 no.3
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• pp.200-208
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• 2005

Purpose: Tc-99m labeled diethylenetriaminepentaacctic acid (DTPA)-coupled galactosylated human serum albumin (GSA) is a currently used imaging agent for asialoglycoprotein receptor (ASGPR) of the liver, but, it has several shortcomings. Recently a new ASGPR imaging agent, $^{99m}Tc$-lactosylated human serum albumin (LSA), with simple labeling procedure, high labeling efficiency, high stability was developed. In order to assess the feasibility of the $^{99m}Tc$-LSA as a ASGPR imaging radiopharmaceuticals, we performed biodistribution study of the tracer in liver injured mice model and the results were compared with histolgic data. Materals and Methods: To induce hepatic damage in ICR mice, diethylnitrosamine (DEN) ($60mg/kg/week{\times}5time$, low dose or $180mg/kg/week{\times}2times$, high dose) and thioacetamide (TAA) ($50mg/kg{\times}1time$) were administrated intraperitoneally. Degree of liver damage was evaluated by tissue hematoxilin-eosin stain, and expression of asialoglycoprotein receptor (ASGPR) was assessed by immunohistochemistry using ASGPR antibody. $^{99m}Tc$-LSA was intravenously administrated via tail vein in DEN or TAA treated mice, and biodistribution study of the tracer was also performed. Results: DEN treated mice showed ballooning of hepatocyte and inflammatory cell infiltration in low dose group and severe hapatocyte necrosis in high dose group, and low dose group showed higher ASGPR staining than control mice in immunohistochemical staining. TAA treated mice showed severe hepatic necrosis. $^{99m}Tc$-LSA Biodistribution study showed that mice with hepatic necrosis induced by high dose DEN or TAA revealed higher blood activity and lower liver activity than control mice, due to slow clearance of the tracer by the liver. The degree of liver uptake was inversely correlated with the degree of histologic liver damage. But low dose DEN treated mice with mild hepatic injury showed normal blood clearance and hepatic activity, partly due to overexpression of ASGPR in mice with mild degree hepatic injury. Conclusion: Liver uptake of $^{99m}Tc$-LSA was inversely correlated with degree of histologic hepatic injury in DEN and TAA treated mice. These results support that $^{99m}Tc$-LSA can be used to evaluate the liver status in liver disease patients.

• Journal of Korean Society of Forest Science
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• v.112 no.2
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• pp.157-172
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• 2023

Controlled pollination (CP) is an important method in tree breeding programs because CP quickly generates desirable genotypes and rapidly maximizes genetic gains. However, few studies have evaluated the efficiency and success rate of CP in the breeding program of Pinus densiflora. To evaluate CP and the management of control pollinated progenies, we used 159 individuals in CB2 × KW40 or KW40 × CB2 populations that were established in 2015. After genotyping microsatellite loci, we estimated whether the number of primers was sufficient or not. Then, we performed pedigree analysis. The result showed that the number of primers was sufficient. By pedigree analysis, we found out that 60 of 159 individuals had been generated by the mating between CB2 and KW40. In the maternity analysis, there was evidence to indicate the possibility of management problems. Therefore, we excluded 54 individuals and repeated the pedigree analysis. In the second analysis, 47 of 105 individuals were generated by the mating between CB2 and KW40. To increase the efficiency of CP in tree breeding programs, several precautions are required. It is necessary to identify the exact clone names of the mother and father trees. In addition, CP processes should be performed properly, including deciding on the schedule of CP and the isolation of female strobili or flowers. Finally, the monitoring of hybrid progenies management after mating is important. Molecular markers should be used to identify the clone names of the mother and father trees and for monitoring post hoc management. This study provides a reference for tree breeding programs for the future control pollination of pine species.

• The Korean Journal of Nuclear Medicine
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• v.27 no.2
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• pp.223-232
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• 1993

Technetium labeled isonitrile analogues are widely used as myocardial perfusion imaging agents. We synthesized and characterized a new isonitrile compound, ethyl 3-isocyanobutyrate(EIB). Proton and $^{13}C$ NMR spectroscopy and thin layer chromatography with a $C_{18}$ coat was performed. EIB was easily labeled with $^{99m}TcO_4^-$- with sodium dithionite. The labeling efficiency measured by RP-HPLC was over 95%. The labeled product was stable with dilution in normal saline and with prolonged incubation at room temperature. There was no formation of secondary products or free $^{99m}TcO_4^-$. In vivo kinetics study of $^{99m}Tc$ (I) labeled EIB in rabbits showed adequate myocardial uptake, good contrast against lung background, and relatively rapid liver clearance. The heart to lung ratio was over 2.5 and the heart to liver ratio was approximately from 0.4 to 5 at 60 minutes post injection. Hepatic clearance of $^{99m}Tc-MIBI$ was faster ($t_{1/2}$=6 minutes) than that of $^{99m}Tc-MIBI$. In vivo kinetics observed in dog was similar to that in rabbit but there was faster gallbladder filling, and thus lower liver background. SPECT imaging of the canine myocardium showed favorable imaging characteristics. However, biodistribution in mice demonstrated a myocardial % injected dose/organ of less than 0.1%. This was thought to be due to interspecies difference in plasma esterase activity. In human plasma, $^{99m}Tc$ ( I ) labeled EIB was stable for at least 2 hours, without production of secondary products by HPLC. We conclude that ethyl 3-isocyanobutyrate may be a potential new myocardial perfusion imaging agent and deserves further investigation as to its usefulness for clinical use.

• Journal of Radiopharmaceuticals and Molecular Probes
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• v.1 no.1
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• pp.53-61
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• 2015

Lutetium-177 ($T_{1/2}=6.71day$) is an adequate radionuclide for therapy, which has both beta emission ($E_{max}=497keV$) for therapeutic effect and gamma emission (113 and 208 keV) for imaging. $^{177}Lu$ labeled ethylenediamine-N,N,N',N'-tetrakis (methylene phosphonic acid) (EDTMP) and 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraaminomethylenephosphonate (DOTMP) have been proposed as radiopharmaceuticals for bone pain palliation. In this study, we compared radiochemistry and biodistribution of $^{177}Lu$-EDTMP and $^{177}Lu$-DOTMP. EDTMP and DOTMP were synthesized, and 1 mg of each was labeled with $^{177}Lu$ at pH 7~8 with high efficiency (>98%). For comparative biodistribution studies, $^{177}Lu$-EDTMP or $^{177}Lu$-DOTMP were injected into ICR-mice through tail vein, and then biodistribution data were obtained as percentages of injected dose per gram of tissue (% ID/g). Urine excretions of both agents in mice were checked for 7 days. Rat images were also obtained after injection of $^{177}Lu$-EDTMP or $^{177}Lu$-DOTMP. $^{177}Lu$-DOTMP (100% at 1 min) showed faster labeling than $^{177}Lu$-EDTMP (100% at 30 min). Both of them were stable at least for 21 days at room temperature. High bone uptakes were found for both $^{177}Lu$-EDTMP and $^{177}Lu$-DOTMP: 38.0 and 34.1% ID/g at 3 hr, respectively; and 33.2 and 18.8% ID/g at 7 day, respectively. Rapid excretions to urine were found for both agents ($^{177}Lu$-EDTMP: 56%, $^{177}Lu$-DOTMP: 63% at 1 day). Other organs showed very low uptakes. Rat images of both $^{177}Lu$-EDTMP and $^{177}Lu$-DOTMP showed high bone uptakes and low soft tissue uptakes. In conclusion, both $^{177}Lu$-EDTMP and $^{177}Lu$-DOTMP showed high potential as bone pain palliation agents. $^{177}Lu$-EDTMP showed higher bone uptake and slower bone clearance in mice than those of $^{177}Lu$-DOTMP.