Safety, tolerability, and pharmacokinetics of l-ornithine phenylacetate in patients with acute liver injury/failure and hyperammonemia.

Cerebral edema remains a significant cause of morbidity and mortality in patients with acute liver failure (ALF) and has been linked to elevated blood ammonia levels. l-ornithine phenylacetate (OPA) may decrease ammonia by promoting its renal excretion as phenylacetylglutamine (PAGN), decreasing the risk of cerebral edema. We evaluated the safety, tolerability, and pharmacokinetics of OPA in patients with ALF and acute liver injury (ALI), including those with renal failure. Forty-seven patients with ALI/ALF and ammonia ≥60 µM were enrolled. Patients received OPA in a dose escalation scheme from 3.3 g every 24 hours to 10 g every 24 hours; 15 patients received 20 g every 24 hours throughout the infusion for up to 120 hours. Plasma phenylacetate (PA) concentrations were uniformly below target (<75 µg/mL) in those receiving 3.3 g every 24 hours (median [interquartile range] 5.0 [5.0] µg/mL), and increased to target levels in all but one who received 20 g every 24 hours (150 [100] µg/mL). Plasma [PAGN] increased, and conversion of PA to PAGN became saturated, with increasing OPA dose. Urinary PAGN clearance and creatinine clearance were linearly related (r = 0.831, P < 0.0001). Mean ammonia concentrations based on the area under the curve decreased to a greater extent in patients who received 20 g of OPA every 24 hours compared with those who received the maximal dose of 3.3 or 6.7 g every 24 hours (P = 0.046 and 0.022, respectively). Of the reported serious adverse events (AEs), which included 11 deaths, none was attributable to study medication. The only nonserious AEs possibly related to study drug were headache and nausea/vomiting. CONCLUSION: OPA was well-tolerated in patients with ALI/ALF, and no safety signals were identified. Target [PA] was achieved at infusion rates of 20 g every 24 hours, leading to ammonia excretion in urine as PAGN in proportion to renal function. Randomized, controlled studies of high-dose OPA are needed to determine its use as an ammonia-scavenging agent in patients with ALF. (Hepatology 2018;67:1003-1013).

Pharmacotherapies that specifically target ammonia for the prevention and treatment of hepatic encephalopathy in adults with cirrhosis.

BACKGROUND: Hepatic encephalopathy is a common complication of cirrhosis, with high related morbidity and mortality. Its presence is associated with a wide spectrum of change ranging from clinically obvious neuropsychiatric features, known as 'overt' hepatic encephalopathy, to abnormalities manifest only on psychometric or electrophysiological testing, 'minimal' hepatic encephalopathy. The exact pathogenesis of the syndrome is unknown but ammonia plays a key role. Drugs that specifically target ammonia include sodium benzoate, glycerol phenylbutyrate, ornithine phenylacetate, AST-120 (spherical carbon adsorbent), and polyethylene glycol. OBJECTIVES: To evaluate the beneficial and harmful effects of pharmacotherapies that specifically target ammonia versus placebo, no intervention, or other active interventions, for the prevention and treatment of hepatic encephalopathy in people with cirrhosis. SEARCH METHODS: We searched the Cochrane Hepato-Biliary Controlled Trials Register, CENTRAL, MEDLINE, Embase, and three other databases to March 2019. We also searched online trials registries such as ClinicalTrials.gov, European Medicines Agency, WHO International Clinical Trial Registry Platform, and the Food and Drug Administration for ongoing or unpublished trials. In addition, we searched conference proceedings, checked bibliographies, and corresponded with investigators. SELECTION CRITERIA: We included randomised clinical trials comparing sodium benzoate, glycerol phenylbutyrate, ornithine phenylacetate, AST-120, and polyethylene glycol versus placebo or non-absorbable disaccharides, irrespective of blinding, language, or publication status. We included participants with minimal or overt hepatic encephalopathy or participants who were at risk of developing hepatic encephalopathy. DATA COLLECTION AND ANALYSIS: Two review authors independently extracted data from the included reports. The primary outcomes were mortality, hepatic encephalopathy, and serious adverse events. We undertook meta-analyses and presented results using risk ratios (RR) or mean differences (MD), both with 95% confidence intervals (CIs), and I2 statistic values as a marker of heterogeneity. We assessed bias control using the Cochrane Hepato-Biliary domains and the certainty of the evidence using GRADE. MAIN RESULTS: We identified 11 randomised clinical trials that fulfilled our inclusion criteria. Two trials evaluated the prevention of hepatic encephalopathy while nine evaluated the treatment of hepatic encephalopathy. The trials assessed sodium benzoate (three trials), glycerol phenylbutyrate (one trial), ornithine phenylacetate (two trials), AST-120 (two trials), and polyethylene glycol (three trials). Overall, 499 participants received these pharmacotherapies while 444 participants received a placebo preparation or a non-absorbable disaccharide. We classified eight of the 11 trials as at 'high risk of bias' and downgraded the certainty of the evidence to very low for all outcomes.Eleven trials, involving 943 participants, reported mortality data, although there were no events in five trials. Our analyses found no beneficial or harmful effects of sodium benzoate versus non-absorbable disaccharides (RR 1.26, 95% CI 0.49 to 3.28; 101 participants; 2 trials; I2 = 0%), glycerol phenylbutyrate versus placebo (RR 0.65, 95% CI 0.11 to 3.81; 178 participants; 1 trial), ornithine phenylacetate versus placebo (RR 0.73, 95% CI 0.35 to 1.51; 269 participants; 2 trials; I2 = 0%), AST-120 versus lactulose (RR 1.05, 95% CI 0.59 to 1.85; 41 participants; 1 trial), or polyethylene glycol versus lactulose (RR 0.50, 95% CI 0.09 to 2.64; 190 participants; 3 trials; I2 = 0%).Seven trials involving 521 participants reported data on hepatic encephalopathy. Our analyses showed a beneficial effect of glycerol phenylbutyrate versus placebo (RR 0.57, 95% CI 0.36 to 0.90; 178 participants; 1 trial; number needed to treat for an additional beneficial outcome (NNTB) 6), and of polyethylene glycol versus lactulose (RR 0.19, 95% CI 0.08 to 0.44; 190 participants; 3 trials; NNTB 4). We did not observe beneficial effects in the remaining three trials with extractable data: sodium benzoate versus non-absorbable disaccharides (RR 1.22, 95% CI 0.51 to 2.93; 74 participants; 1 trial); ornithine phenylacetate versus placebo (RR 2.71, 95% CI 0.12 to 62.70; 38 participants; 1 trial); or AST-120 versus lactulose (RR 1.05, 95% CI 0.59 to 1.85; 41 participants; 1 trial).Ten trials, involving 790 participants, reported a total of 130 serious adverse events. Our analyses found no evidence of beneficial or harmful effects of sodium benzoate versus non-absorbable disaccharides (RR 1.08, 95% CI 0.44 to 2.68; 101 participants; 2 trials), glycerol phenylbutyrate versus placebo (RR 1.63, 95% CI 0.85 to 3.13; 178 participants; 1 trial), ornithine phenylacetate versus placebo (RR 0.92, 95% CI 0.62 to 1.36; 264 participants; 2 trials; I2 = 0%), or polyethylene glycol versus lactulose (RR 0.57, 95% CI 0.18 to 1.82; 190 participants; 3 trials; I2 = 0%). Likewise, eight trials, involving 782 participants, reported a total of 374 non-serious adverse events and again our analyses found no beneficial or harmful effects of the pharmacotherapies under review when compared to placebo or to lactulose/lactitol.Nine trials, involving 733 participants, reported data on blood ammonia. We observed significant reductions in blood ammonia in placebo-controlled trials evaluating sodium benzoate (MD -32.00, 95% CI -46.85 to -17.15; 16 participants; 1 trial), glycerol phenylbutyrate (MD -12.00, 95% CI -23.37 to -0.63; 178 participants; 1 trial), ornithine phenylacetate (MD -27.10, 95% CI -48.55 to -5.65; 231 participants; 1 trial), and AST-120 (MD -22.00, 95% CI -26.75 to -17.25; 98 participants; 1 trial). However, there were no significant differences in blood ammonia concentrations in comparison with lactulose/lactitol with sodium benzoate (MD 9.00, 95% CI -1.10 to 19.11; 85 participants; 2 trials; I2 = 0%), AST-120 (MD 5.20, 95% CI -2.75 to 13.15; 35 participants; 1 trial), and polyethylene glycol (MD -29.28, 95% CI -95.96 to 37.39; 90 participants; 2 trials; I2 = 88%). FUNDING: Five trials received support from pharmaceutical companies while four did not; two did not provide this information. AUTHORS' CONCLUSIONS: There is insufficient evidence to determine the effects of these pharmacotherapies on the prevention and treatment of hepatic encephalopathy in adults with cirrhosis. They have the potential to reduce blood ammonia concentrations when compared to placebo, but their overall effects on clinical outcomes of interest and the potential harms associated with their use remain uncertain. Further evidence is needed to evaluate the potential beneficial and harmful effects of these pharmacotherapies in this clinical setting.

Safety of ornithine phenylacetate in cirrhotic decompensated patients: an open-label, dose-escalating, single-cohort study.

AIMS: Confirm in patients with cirrhosis and gastrointestinal bleeding the safety of ornithine phenylacetate (OP) and assess the pharmacokinetic profile of OP and its effects on plasma ammonia. BACKGROUND: OP is a drug that has shown experimentally to decrease hyperammonemia and improve hepatic encephalopathy. OP is safe in healthy subjects and in stable patients with cirrhosis, but there are no data in decompensated cirrhosis. METHODS: We performed a study to assess safety and tolerance of OP in cirrhotic patients after an episode of upper gastrointestinal bleeding.Ten patients were included within 24 hours of an upper gastrointestinal bleeding. OP was administered as a continuous infusion up to a maximum of 10 g/24 h (0.42 g/h) for 5 days. The infusion was started at 33% of the target dose and increased at 12-hour intervals achieving target dose at 24 hours. Ammonia was also assessed in control group of 10 patients. RESULTS: No severe adverse events were observed. Mild adverse events were reported in 4 patients. Plasma ammonia (baseline: 80±43 µmol/L) showed a progressive drop between baseline and 36 hours (42±15 µmol/L), 72 hours (44±15 µmol/L), 96 hours (40±24 µmol/L), and 120 hours (33±14 µmol/L). Plasma ammonia at 24 hours was significantly higher in the control group. Plasma glutamine showed a significant decrease (-37% at day 5) and its excretion in urine as phenylacetylglutamine, a progressive rise (52±35 mmol at day 5). CONCLUSIONS: OP is a safe and well-tolerated drug in decompensated cirrhotics that may decrease plasma ammonia by inducing its appearance as phenylacetylglutamine in urine.

Phase I trial and pharmacokinetic studies of alpha-difluoromethylornithine--an inhibitor of polyamine biosynthesis.

alpha-Difluoromethylornithine (DFMO), an enzyme-activated, irreversible inhibitor of ornithine decarboxylase, blocks polyamine biosynthesis and has antitumor effects in animal tumor models as well as in athymic mice implanted with human small cell carcinoma. This study was designed to determine the maximally tolerated dose of oral DFMO administered every six hours for 28 days to patients with advanced solid tumors or lymphomas. DFMO levels were measured using an ion exchange chromatographic assay and pharmacokinetic studies were performed in patients treated at each dose level. Twenty-two patients received 24 courses of DFMO. The drug was generally well tolerated. Thrombocytopenia was the dose-limiting toxicity and gastrointestinal side effects were also seen. Thrombocytopenia developed in 11 of 16 patients who had received prior chemotherapy but the four patients who had no prior chemotherapy had no decrease in the platelet count. The steady state level of DFMO achieved at the highest dose (3 g/m2) were found to be within the range needed for inhibition of ornithine decarboxylase in cell-culture systems as well as for the inhibitory activity against various human tumors in vitro. A DFMO dose of 2.25 g/m2 every six hours is recommended for phase II studies in patients previously treated with cytotoxic drugs.

Sequential inhibition of polyamine synthesis. A phase I trial of DFMO (alpha-difluoromethylornithine) and methyl-GAG [methylglyoxal-bis(guanylhydrazone)].

Both DFMO and methyl-GAG inhibit sequential enzymatic reactions in the pathway of polyamine biosynthesis. Since polyamines may be important factors in proliferation of cancer cells, we initiated a phase-I study of these agents in patients with advanced cancer. DFMO was given by mouth at a constant daily dose of 4 g/m2 starting on day 1 of the treatment protocol. The dose of methyl-GAG ranged from 200 to 700 mg/m2 administered IV every 2 weeks beginning on day 4. Twenty-two patients were entered into the protocol. Toxic reactions to this therapy were dose-related and included nausea, fatigue, diarrhea, and myelosuppression. One patient with colon cancer experienced a greater than 50% decrease in measurable disease but developed severe myelotoxicity. While DFMO was well tolerated, the combination of drugs appeared to cause substantially more hematologic and gastrointestinal toxicity than encountered during our recent experience with methyl-GAG used alone. We suggest that future studies of this drug combination carefully evaluate levels of polyamines and inhibition of enzymatic activity to minimize toxicity.

Ornithine alpha-ketoglutarate in nutritional support.

Ornithine alpha-ketoglutarate (OKG) is a salt formed of two molecules of ornithine and one molecule of alpha-ketoglutarate. OKG has been successfully used by the enteral and parenteral route in burn, traumatized, and surgical patients and in chronically malnourished subjects. According to the metabolic situation, OKG treatment decreases muscle protein catabolism and/or increases synthesis. In addition, OKG promotes wound healing. The mechanism of action of OKG is not fully understood, but the secretion of anabolic hormones (insulin, human growth hormone) and the synthesis of metabolites (glutamine, polyamines, arginine, ketoacids) may be involved.

L-ornithine alpha-ketoglutarate in HIV infection: effects on muscle, gastrointestinal, and immune functions.

OBJECTIVES: There have been claims that l-ornithine alpha-ketoglutarate (OKG) exerts anticatabolic, anabolic, and immunomodulating properties. This study aimed at quantifying the effects of OKG on muscle force, body composition, and immune function in outpatients infected with the human immunodeficiency virus (HIV) and presenting weight loss. METHODS: Forty-six HIV(+) patients were included in a double-blind, prospective, randomized, controlled trial for 12 wk (10 g/d of OKG or isonitrogenous placebo and nutritional counseling). Podometry, handgrip strength, step test, triceps skinfold thickness, 50-kHz bioelectrical impedance, 3-d diet record, CD4 cell count, HIV-1 RNA concentration (viral load), and gastrointestinal symptoms were assessed at 0, 4, 8, and 12 wk. RESULTS: At baseline, patients (OKG, n = 22; placebo, n = 24) has similar CD4 counts (338 +/- 172 and 310 +/- 136 cells/mL), viral load (3.6 +/- 1.3 and 3.5 +/- 1.3 log(10) copies/mL), body mass index (20.0 +/- 2.4 and 20.6 +/- 3.0 kg/m(2)), weight loss (9.0 +/- 3.12 and 9.4 +/- 3.0 kg), and food intake (2509 +/- 962 and 2610 +/- 808 kcal/d). Twenty-nine patients completed the protocol. Both groups increased their body mass index (P = 0.02 versus baseline) and triceps skinfold thickness (P < 0.01 versus baseline). They showed a similar positive correlation between handgrip strength and fat-free mass. Frequency of gastrointestinal symptoms increased in the OKG group (86% versus 54% in the placebo group, P = 0.025). No other differences were observed between groups. CONCLUSIONS: All patients increased their body mass index and triceps skinfold thickness due to food supplementation and diet counseling. Oral OKG failed to improve nutritional, functional, or immunologic status in these weight-losing HIV(+) patients and had important gastrointestinal side effects.

Ornithine-vasopressin gangrene and reconstruction of the upper lip. Case report.

A bilateral cleft lip and palate case received ornithine-vasopressin intra-operatively in preparation for a vasoconstricted field of the various lip segments prior to the repair of the cleft lip. A cyanotic tinge appeared immediately. This eventually led to total necrosis of the upper lip. Various relevant blood tests were done and a mild thrombocytosis was found. Surgical reconstruction of the upper lip was performed by means of a forked cross-lip flap--the main blood supply coming from the columella--as well as by means of an inferiorly pedicled cheek flap from the para-nasal area.

[Retinal degeneration after intravitreal injection of ornithine. 1. Early change after administration].

Retinal damage 1 to 7 days after intravitreal injection of 1-ornithine hydrochloride solution into monkey eyes was investigated. Clinically the retinal pigment epithelium (RPE) showed edema, and hyperfluorescence on fluoroangiograms, predominantly in the equatorial region at 3 to 7 days. No retinal changes were seen at the posterior pole. Histopathologically, RPE in the equatorial region showed severe damage 24 hours after injection, becoming liquefactively necrotic and degenerated at 3 to 7 days. Early changes in RPE consisted of an increase in the number of secondary lysosomes, disappearance of basal infolding, destruction of most of the smooth endoplasmic reticulum, shortening of microvilli, and marked swelling of mitochondria. These RPE changes were seen slightly in the posterior pole. There were slight changes in the inner retina including photoreceptors. In eyes injected intravitreally by hi-osmosis of 3.5% NaCl solution, with equivalent osmolarity to ornithine solution, mild changes of microvilli and mitochondria were seen, but not necrotic changes. Our experiment showed that intravitreal injection of a small amount of 1-ornithine hydrochloride induced severe RPE damage. Selective toxicity of ornithine to RPE appeared mainly in the equatorial region.

Efficacy and safety of ornithine alpha-ketoglutarate in heel pressure ulcers in elderly patients: results of a randomized controlled trial.

OBJECTIVE: Pressure ulcers affect predominantly the elderly and nutritional status is a known risk factor. Guidelines on pressure ulcers provide recommendation on nutritional management. Ornithine alpha-ketoglutarate (OKG) is an adjuvant treatment in undernourished elderly patients or in patients with hypercatabolism states. It is a precursor of different amino-acids which play a role in the process of healing. The objective of the study is to determine the efficacy of OKG on pressure ulcer area reduction after six weeks of treatment. DESIGN: Multi-centre, international, randomized, comparative, double blind, parallel groups, placebo-controlled study. PARTICIPANTS: 160 patients (ITT population) aged over 60 years with a heel pressure ulcer at stage II or III. INTERVENTION: Patients received OKG (n=85) or placebo (n=75) once a day for 6 weeks. MEASUREMENTS: Ulcer area was measured each week, using a tracer. The primary endpoint was the percentage reduction of the surface at the final visit: [(Wound areatn - Wound areat0)/ (Wound areat0)]. RESULTS: At inclusion, ulcer area distribution deviated from normal distribution (median ulcer area OKG 6.6 cm(2), placebo 3.9 cm2, p=0.044, Mann-Whitney test). As healing is strongly related to baseline ulcer area, the abnormal distribution was a major bias. Therefore it was decided to perform the analysis on 2 sub-groups of patients according to the mean ulcer area, i.e. above or below 8 cm2. The mean wound area reduction for baseline area 8 cm2 no between group differences on either parameter was detected. When closure rate is considered, a significant difference in favor of OKG group is observed (- 0.07 cm2/day in the OKG group and - 0.04 cm2/day in the placebo groups respectively p=0.007, Mann-Whitney test). Thirty serious adverse events were reported in 28 patients (15 allocated to OKG and 13 to placebo). None of them was considered treatment related. CONCLUSION: This clinical trial supports a potential benefit of OKG 10g daily in the subgroup of patients with pressure ulcers

Ornithine supplementation and insulin release in bodybuilders.

Ornithine supplementation has gained popularity with athletes because of its alleged potential to release anabolic hormones, factors governing skeletal muscle hypertrophy. Three female and nine male bodybuilders served as subjects in a study to test the effectiveness of oral ornithine in bringing about the release of insulin, an anabolic hormone. After an overnight fast, subjects were administered 40, 100, or 170 mg.kg-1 L-ornithine.HCl by mouth in a random fashion on three consecutive Saturday mornings. Blood samples were drawn at baseline (T = 0), 45, and 90 min afterward. Serum ornithine levels were elevated (p < 0.01) at T = 45 and 90 min for all three dosage levels. However, serum insulin did not change from baseline levels at any dose of ornithine. The present findings show that ornithine is not an insulin secretagogue.

Difluoromethylornithine and leukocyte interferon: a phase I study in cancer patients.

Difluoromethylornithine (DFMO), an irreversible inhibitor of ornithine decarboxylase, and human leukocyte interferon (IFN-alpha) have synergistic anti-tumor activities in vivo in B 16 melanoma and in vitro against several human cancer cell lines. We have, therefore, carried out a phase I combination study with DFMO plus alpha interferon in the following manner: DFMO was maintained at a steady dose for the first four levels, 1.5 g/m2 every 6 hr. IFN-alpha was given in 100% increments ranging from 0.4 X 10(6)U/m2 to 3.2 X 10(6)U/m2 i.m. daily. At the fifth dose level both IFN-alpha and DFMO were raised by 100 and 50% respectively. From levels one through four the combination was well tolerated with no dose interruptions required because of G.I. toxicity or myelosuppression. However, at dose level 5, one-third of the patients required dose cessation and decrease due to nausea, vomiting and diarrhea. We conclude that for phase II studies the maximal tolerated dose is 3.2 million units of IFN-alpha/m2 and 1.5 g/m2 of DFMO every 6 hr. Of 12 patients with metastatic melanoma, 2 had partial remissions lasting 58+ and 36+ weeks. Two additional patients had minor responses lasting 29 and 32+ weeks. Minor responses were observed in a patient with colon carcinoma and a patient with renal carcinoma. The clinical activity of the combination is currently being pursued in a phase II study among patients with metastatic malignant melanoma.

Phase I study of alpha-difluoromethylornithine and methyl-GAG.

alpha-Difluoromethylornithine (DFMO) is a potent inhibitor of the synthesis of putrescine (pu) and spermidine (sd) in some benign and malignant tissues. Intracellular deprivation of pu and sd has been shown to induce an enhanced uptake of polyamine-analogs such as methyl-GAG (MGBG). The purpose of this study was to investigate the tolerance and the toxicity of the combination of DFMO and MGBG. Thirty-six patients received 4 X 2 g of DFMO/day orally and every 2 weeks 250-500 mg/m2 of MGBG as a 2-hr infusion, starting on day 14. Besides the well known acute and late side-effects of methyl-GAG, dose-limiting toxicity consisted also of thrombocytopenia, leucopenia, dyspnea, hemolysis and jaundice. The maximal tolerated dose of MGBG for one course was 350 mg/m2 and for repeated courses 250 mg/m2, due to cumulative toxicity. Furthermore, after 8 weeks of continuous administration of DFMO 70% of the patients had a severe hearing loss, which was reversible after a treatment delay of 4-6 weeks. Since the hearing loss prohibited the continuous use of DFMO, two different schedules of intermittent DFMO-administration together with two different infusion periods of MGBG have been investigated in 15 patients. In none of these patients did hearing loss occur. The schedule of continuous administration of 4 X 2 g of DFMO/day orally for 21 days and 250 mg/m2 of MGBG as a 24-hr infusion on days 7, 14 and 21, repeated on day 42, was tolerated best. In 28 evaluable patients two partial remissions were seen. Pretreatment with DFMO significantly enhanced the toxicity of MGBG and the combination of both drugs produced side-effects not seen with either drug alone.

Phase I evaluation of intravenous difluoromethylornithine--a polyamine inhibitor.

Difluoromethylornithine (DFMO), a non-competitive inhibitor of ornithine decarboxylase (ODC), the rate limiting enzyme of the polyamine synthetic pathway was evaluated in a Phase I trial. Intravenous DFMO was given to twenty patients with refractory leukemia by continuous infusion in doses from 5.5 to 64 g/m2. Toxicity clearly attributable to the drug was not severe and other than nausea and vomiting did not increase with dose. The previously reported ototoxicity which occurred with the oral form appeared to be less frequent. Loss of hearing which improved when the drug was stopped was seen in four patients, three of whom were simultaneously receiving aminoglycosides. Anorexia occurred in some patients at all doses. Vomiting, necessitating dosage reduction, was a significant problem at the highest dose administered. No patient achieved a remission but there was stabilization or decrease in circulating blast cells in several patients. This growth inhibition did not appear to be dosage related.

Heterogeneity in ornithine cytotoxicity of bovine retinal pigment epithelial cells in primary culture.

Gyrate atrophy of the choroid and retina is a chorioretinal degeneration caused by hyperornithinemia and a deficiency of ornithine-delta-aminotransferase (OAT). We recently showed that ornithine exhibits cytotoxicity to human retinal pigment epithelial (RPE) cell lines treated with the OAT inhibitor, 5-fluoromethylornithine (5-FMOrn), and suggested that this system may be an in vitro model of gyrate atrophy. In the present study, in order to apply this system to primary cultured RPE cells, we freshly prepared RPE cells from bovine eyes and studied the effect of ornithine on cell damage. Two phenotypes, epithelioid and fusiform, which coexisted in the primary culture and epithelioid phenotype cells, but not fusiform ones, were severely damaged and partially detached from the substrate by 10 m m ornithine and 0.5 m m 5-FMOrn. Neither ornithine nor 5-FMOrn alone exhibited such cytotoxicity to both phenotypes of RPE cells. Proline significantly prevented the ornithine-induced cytotoxicity. Epithelioid and fusiform phenotypes isolated from the primary culture showed different distribution of actin filaments. A combination of ornithine and 5-FMOrn time-dependently inhibited [(3)H]thymidine incorporation in the epithelioid, but not fusiform, cells. Proline prevented the inhibition of [(3)H]thymidine incorporation by ornithine in 5-FMOrn-treated epithelioid cells. Furthermore, l -azetidine-2-carboxylic acid, a collagen synthesis inhibitor, reduced [(3)H]thymidine incorporation in epithelioid, but not fusiform, cells, which was reversed by proline. These results demonstrate that the epithelioid phenotype of bovine RPE cells becomes susceptible to ornithine following inactivation of OAT. The phenotypic cells and its prevention by proline may provide insight into biochemical triggers that induce gyrate atrophy.