• Journal of Genetic Medicine
• /
• v.1 no.1
• /
• pp.5-10
• /
• 1997

The amino acids formed by degradation of proteins ingested produce ammonia. The ammonia which is broken down and excreted as urea through a process known as the Klebs-Hensleit cycle or the urea cycle (Rezvani, 1995). The urea cycle consists of five enzymes necessary for the synthesis of carbamyl phosphate, citrulline, argininosuccinate, arginine, and urea: carbamyl phosphate synthetase (CPS), ornithine transcarbamylase (OTC), argininosuccinate synthetase (AS), argininosuccinate lyase (AL), and arginase (ARG) (Lloyd, 1992). Congenital deficiencies of the enzymes involved in the urea cycle are diseases that are almost fatal without treatment, showing symptoms like vomiting, lethargy, dyspnea, and coma due to hyperammonemia coming from the accumulation of ammonia and metabolic precursors resulting from the deficiency of one of these enzymes (Batshaw and Brusilow, 1983). Among these, the disease manifested by the congenital deficiency of argininosuccinate synthetase (AS) which is associated with the formation of argininosuccinate in citrulline is called argininosuccinate synthetase deficiency or citrullinemia. There have been two reports on this so far in Korea; one in July 1987 by Kim et al. and the other by Park et al. in 1995. We are to report a case of successful treatment of a child with citrullinemia who was transferred to our hospital due to dyspnea, lethargy, feeding difficulties, convulsions and cyanosis together with some document studies related to this case.

• Proceedings of the PSK Conference
• /
• 2003.04a
• /
• pp.231.2-231.2
• /
• 2003

The efficient EPO (Erythropoietin) expression system in Chinese Hamster Ovary (CHO) cells was devised through the removal of ammonium ion accumulated in the media by introducing urea cycle enzymes. Previously, we developed C05 cell by transfecting the carbamoly phosphate synthase (CPS) and ornithine transcarbamoylase (OTC) into the EPO expressing CHO cell, IBE. (omitted)

• Journal of Microbiology and Biotechnology
• /
• v.13 no.2
• /
• pp.217-224
• /
• 2003

Previously, we developed a CHO cell line (CHO-OTC1-Al9) that expresses the first two enzymes in the urea cycle and exhibits a higher ammonia-removing ability and faster growth rate than a vector-controlled CHO cell line (CHO-neo-5). The current study was undertaken to develop a cell line with an ammonia-removing ability higher than the cell line developed previously. To accomplish this, CHO cell lines expressing the first three, first four, or all five enzymes of the urea cycle were constructed using a stable transfection method. Finally, the CHO-AS-16, CHO-AL-19, and CHO-Arg-11 cell lines expressing the first three, first four, and all five enzymes of the urea cycle, respectively, were selected and found to exhibit higher ammonia-removing ability than the CHO-OTC1-Al9 cell line. Among the three selected cell lines, CHO-AL-19 showed the highest ammonia-removing ability and highest cell viability at a higher cell density, with 40% and 15% lower ammonia concentration in the, culture media than that of CHO-neo-5 and CHO-OTC1-A19 cell lines, respectively. CHO-AL-19 also showed 44% and 10% higher cell viability than the CHO-neo-5 and CHO-OTC-Al9 cell lines, at a higher cell density, respectively. The ammonia concentrations in the culture media were expressed as the ammonia concentration/cell, and the CHO-AL-19 cells revealed 45-60% and 20% lower ammonia concentration/cell than the CHO-neo-5 and CHO-OTC1-Al9 cells, respectively.

• Journal of The Korean Society of Inherited Metabolic disease
• /
• v.5 no.1
• /
• pp.76-87
• /
• 2005

Various disorders cause hyperammonemia during childhood. Amongthem are those caused by inherited defects in urea synthesis and related metabolic pathways. These disorders can be grouped into two types: disorders of the enzymes that comprise the urea cycle, and disorders of the transporters or metabolites of theamino acids related to the urea cycle. Principal clinical features of these disorders are caused by elevated levels of blood ammonium. Additional disease-specific symptoms are related to the particular metabolic defect. These specific clinical manifestations are often due to an excess or lack of specific amino acids. Treatment of urea cycle disorders and related metabolic diseases consists of nutritional restriction of proteins, administration of specific amino acids, and use of alternative pathways for discarding excess nitrogen. Although combinations of these treatments are extensively employed, the prognosis of severe cases remains unsatisfactory. Liver transplantation is one alternative for which a better prognosis is reported.

• Proceedings of the PSK Conference
• /
• 2002.04a
• /
• pp.78-78
• /
• 2002

• Journal of Microbiology and Biotechnology
• /
• v.14 no.4
• /
• pp.844-851
• /
• 2004

Efficient mammalian erythropoietin (EPO)-expression systems are required for therapeutic applications. The accumulation of ammonia is a major problem in the production of recombinant proteins in cultured animal cells. To counter this problem we introduced the first two genes of the urea cycle, carbamoyl phosphate synthetase (CPSI) and ornithine transcarbamylase (OTC), into IBE Chinese Hamster Ovary (CHO) cells by stable transfection. The resulting cell line, CO5, had a higher growth rate and accumulated less ammonia per cell than the parental cell line, IBE. In addition, it produced 2 times more EPO than the parent, and the purified EPO contained a higher proportion of acidic isoforms with approximately 15% more sialic acid.

• 한국생물공학회:학술대회논문집
• /
• 2001.11a
• /
• pp.66-69
• /
• 2001

Previously we developed a CHO cell line (CHO-OTC1-A19) expressing the first two enzymes of urea cycle. This cell line showed higher ammonia removal activity and faster growth rate than the vector controlled CHO cells (CHO-neo-5). The purpose of this study was to develop a cell line with higher ammonia removal activity than the cell line developed previously. To accomplish this, we constructed stable CHO cell lines expressing the first three, the first four, or all five enzymes of urea cycle by the stable transfection method. We finally selected CHO-AL-19 cell line expressing the first three, the first four enzymes of the cycle with higher ammonia activity than CHO-OTC1-A19 and CHO-n대-5 cell lines: 40% and 15% higher than those of CHO-neo-5 and CHO-OTC1-A19 cell lines 72 hour after culture started, respectively. It also showed 44% and 10% higher cell viability than CHO-neo-5 and CHO- OTC1-A19 cell lines at higher cell density. In addition, CHO-AL-19 cells showed 45%-60% and about 20% lower ammonia concentration per cell than those of CHO-neo-% and CHO-OTC1-A19 cell lines, respectively. These results indicate that CHO-AL-19 could be used in the production of human therapeutic proteins with higher efficiency.

• Proceedings of the PSK Conference
• /
• 2003.10b
• /
• pp.170.3-171
• /
• 2003

An efficient Erythropoietin (EPO)-expression system in mammalian cells is required for massive production for therapeutic use. Ammonium ion is a major problem in the production of valuable recombinant proteins in cultured animal cells. Therefore, it is of importance to devise a system by which a high productivity of human therapeutic recombinant protein can be maintained or enhanced under low ammonium concentration. To reduce the ammonium ion accumulated in EPO producing Chinese Hamster Ovary (CHO) ceels, IBE, we introduced the first two genes of the urea cycle, carbamoyl phosphate synthetase (CPSI) and arnithine transcarbamoylase (OTC), into IBE using a stable transfection method. (omitted)

• Proceedings of the PSK Conference
• /
• 2002.10a
• /
• pp.338.1-338.1
• /
• 2002

The efficient Erythropoietin(EPO)-expression system in mammalian cells is required for massive production for therapeutic use. Ammonium ion is a major problem in the production of useful proteins by cultured animal cells and therefore it is of importance to devise a system by which a high productivity of human therapeutic recombinant protein can be maintained or enhanced under low ammonium concentration. (omitted)

• YAKHAK HOEJI
• /
• v.51 no.3
• /
• pp.214-218
• /
• 2007

In the previous reports, we developed the CO5 by introducing genes for the first and second urea cycle enzymes, carbamoyl phosphate synthetase I (CPS I) and ornithine transcarbamoylase (OTC) into the IBE cell lines producing erythropoietin (EPO). The CO5 have been found out to have 15-20% higher cell growth rate and produce 2-times more EPO than the parental cell line, IBE. To investigate the role of CPS I in CO5 cell line for the cell growth and amount of EPO, we knock-downed CPS I gene expression via siRNA treatment. Expression level of EPO in cell lysate of CO5 was 3-5 fold higher than that of IBE. After siRNA treatment, the cell growth of CO5 was decreased 8-21% and the EPO productivity in the cell Iysate was significantly decreased. However, these changes of the cell growth and EPO productivity were not observed in IBE. These results indicate that CPS I gene expression is important for the increased cell growth and EPO productivity of CO5 cell line.