Novel biallelic splice-site BBS1 variants in Bardet-Biedle syndrome: a case report of the first Japanese patient.

PURPOSE: To report the clinical and genetic features of a 9-year-old female Japanese patient with Bardet-Biedl syndrome (BBS). METHODS: Genetic analysis using whole-exome sequencing (WES) was performed for the patient and her parents to identify disease-causing variants. Reverse transcriptase-polymerase chain reaction (RT-PCR) was performed to investigate the impact of splice-site variants. Comprehensive ophthalmic and systemic examinations, including electroretinography (ERG), were performed. RESULTS: In the patient, WES identified novel compound heterozygous splice-site variants (c.124+2T>G and c.723+2T>G) in the BBS1 gene, and RT-PCR revealed skipping of exons 2 and 8 (p.N17AfsX56 and p.T198_K241del). Each parent had one of the variants. Ophthalmologically, the patient's decimal best-corrected visual acuity was 0.6 in the right eye and 0.4 in the left eye. Funduscopy revealed no apparent retinal degeneration or narrowed blood vessels in the periphery, but macular abnormalities were found on fundus autofluorescence imaging and optical coherence tomography images. Unexpectedly, non-recordable responses in rod ERG were found, with a non-recordable response of the right eye and an extremely reduced and delayed a-wave of the left eye in standard ERG, non-recordable responses in cone ERG, and extremely decreased responses in 30 Hz flicker ERG. Finally, the patient fulfilled four primary features of BBS diagnostic criteria: rod-cone dystrophy, polydactyly, central obesity, and learning disabilities, being diagnosed with BBS. CONCLUSIONS: This is the first report of a BBS patient with biallelic splice-site BBS1 variants in the Japanese population. Disparity between funduscopic and ERG findings may be a feature of BBS1-associated rod-cone dystrophy.

Polyamine regulating protein antizyme binds to ATP citrate lyase to accelerate acetyl-CoA production in cancer cells.

Antizyme (AZ) regulates cellular polyamines (i.e., putrescine, spermidine, and spermine) through binding to ornithine decarboxylase and subsequent ubiquitin-independent degradation of the enzyme protein by the 26S proteasome. Screening for AZ-binding proteins using a yeast two-hybrid system identified ATP citrate lyase (ACLY), a cytosolic enzyme which catalyzes the production of acetyl-CoA that is used for lipid anabolism or acetylation of cellular components. We confirmed that both AZ1 and AZ2 bind to ACLY and AZ colocalizes with ACLY to the cytoplasm. Unexpectedly, neither AZ1 nor AZ2 accelerated ACLY degradation. Additionally, purified AZ, particularly AZ1, increased the activity of purified ACLY in a dose-dependent manner in vitro, suggesting that AZ activates ACLY through protein-protein interaction. Polyamines themselves had no effect on the ACLY activity in vitro. Knockdown of AZ1 and/or AZ2 in human cancer cells significantly decreased the ACLY activity as well as cellular levels of acetyl-CoA and cholesterol. Our results are the first to show the crosstalk between polyamine and acetyl-CoA metabolism. We hypothesize that AZ may promote acetyl-CoA synthesis to downregulate spermidine and spermine through acetylation.

Combined conservative treatment and lymphatic venous anastomosis for severe lower limb lymphedema with recurrent cellulitis.

BACKGROUND: Lymphedema may be treated either conservatively or surgically. Although conservative therapy is the first-line treatment, some patients are refractory to it and repeat severe cellulitis. We usually perform lymphaticovenous anastomosis (LVA) for lymphedema patients, and LVA can reduce the frequency of cellulitis. CASE REPORT: A 67-year-old woman who had undergone a radical hysterectomy, pelvic lymphadenectomy, and postoperative radiotherapy for cervical cancer at the age 50 years. She developed lymphedema in both legs, and high-pressure compression stockings caused lymphorrhea in the groin and thigh, resulting in recurrent episodes of cellulitis. Lymphoscintigraphy revealed dilation of the lymphatic vessels in both legs. Results of an indocyanine green test revealed dermal backflow throughout the lower body. After wearing low-pressure stocking, we performed LVA to reduce cellulitis. After confirming the result of LVA, the patients started wearing high-pressure stocking. The patient underwent a subsequent LVA, 3 months after the first, to further improve edema. The lymphorrhea resolved, and cellulitis did not recur. CONCLUSIONS: The combination of surgical treatment and conservative treatment is important for severe lymphedema treatment. Although conservative treatment is usually said to be the first-line treatment, LVA can antecede in cases refractory to conservative treatment.

Rho-kinase regulation of TNF-α-induced nuclear translocation of NF-κB RelA/p65 and M-CSF expression via p38 MAPK in mesangial cells.

The small GTPase Rho and its downstream effector, Rho-associated coiled-coil containing protein kinase (Rho-kinase), regulate a number of cellular processes, including organization of the actin cytoskeleton, cell adhesion, and migration. While pharmacological inhibitors of Rho-kinase signaling are known to block renal inflammation, the molecular basis for this effect is unclear. Here, we provide evidence that proinflammatory TNF-α promotes mesangial expression of macrophage colony-stimulating factor (M-CSF), a key regulator for the growth and differentiation of mononuclear phagocytes, in a Rho-kinase-dependent manner. Consistent with this observation, TNF-α-mediated renal expression of M-CSF in insulin-resistant db/db mice was downregulated by Rho-kinase inhibition. Small interfering RNA-facilitated knockdown of Rho-kinase isoforms ROCK1 and ROCK2 indicated that both isoforms make comparable contributions to regulation of M-CSF expression in mesangial cells. From a mechanistic standpoint, Western blotting and EMSA showed that Rho-kinase and its downstream target p38 MAPK regulate nuclear translocation of NF-κB RelA/p65 and subsequent DNA binding activity, with no significant effects on IκBα degradation and RelA/p65 phosphorylation. Moreover, we showed that Rho-kinase-mediated cytoskeletal organization is required for the nuclear uptake of RelA/p65. Collectively, these findings identify Rho-kinase as a critical regulator of chemokine expression and macrophage proliferation.

MicroRNA-21 is a candidate driver gene for 17q23-25 amplification in ovarian clear cell carcinoma.

BACKGROUND: Epithelial ovarian cancer (EOC) is the most common cause of gynecological malignancy-related mortality. Ovarian clear cell carcinoma (CCC) has unique clinical characteristics and behaviors that differ from other histological types of EOC, including a frequent association with endometriosis and a highly chemoresistant nature, resulting in poor prognosis. However, factors underlying its malignant behavior are still poorly understood. Aberrant expression of microRNAs has been shown to be involved in oncogenesis, and microRNA-21 (miR-21) is frequently overexpressed in many types of cancers. The aim of this study was to investigate the role of miR-21 in 17q23-25 amplification associated with CCC oncogenesis. METHODS: We identified 17q23-25 copy number aberrations among 28 primary CCC tumors by using a comparative genomic hybridization method. Next, we measured expression levels of the candidate target genes, miR-21 and PPM1D, for 17q23-25 amplification by real-time RT-PCR analysis and compared those data with copy number status and clinicopathological features. In addition, immunohistochemical analysis of PTEN (a potential target of miR-21) was performed using the same primary CCC cases. We investigated the biological significance of miR-21 overexpression in CCC using a loss-of-function antisense approach. RESULTS: 17q23-25 amplification with both miR-21 overexpression and PTEN protein loss was detected in 4/28 CCC cases (14.2%). The patients with 17q23-25 amplification had significantly shorter progression-free and overall survival than those without 17q23-25 amplification (log-rank test: p = 0.0496; p = 0.0469, respectively). A significant correlation was observed between miR-21 overexpression and endometriosis. Both PTEN mRNA and PTEN protein expression were increased by miR-21 knockdown in CCC cells. We also confirmed that miR-21 directly bound to the 3'-untranslated region of PTEN mRNA using a dual-luciferase reporter assay. CONCLUSIONS: MiR-21 is a possible driver gene other than PPM1D for 17q23-25 amplification in CCC. Aberrant expression of miR-21 by chromosomal amplification might play an important role in CCC carcinogenesis through the regulation of the PTEN tumor suppressor gene.

Multiple forms of mouse antizyme inhibitor 1 mRNA differentially regulated by polyamines.

Antizyme inhibitor 1 (Azin1), a positive regulator of cellular polyamines, is induced by various proliferative stimuli and repressed by polyamines. It has been reported that the translational repression of Azin1 by polyamines involves an upstream open reading frame on the mRNA, but little has been known about polyamine effect on its transcription or splicing. We found multiple forms of Azin1 transcripts formed by alternative splicing and initiation of transcription from putative alternative start sites. One of the novel splice variants, Azin1-X, has a premature termination codon on 5' extension of exon 7, encodes a C-terminal truncated form of protein (Azin1ΔC), and is subject to nonsense-mediated mRNA decay. 2-Difluoromethylornithine (DFMO), an inhibitor of polyamine synthesis, increased both transcription from the canonical transcription start site and the ratio of the full-length mRNA to Azin1-X mRNA, whereas polyamines show the opposite effect. Thus, polyamines regulate two novel steps of Azin1 expression, namely the transcription and a particular splicing pattern, both of which may affect the level of mRNA encoding the full-length active Azin1 protein.

High-accuracy diagnosis and regional classification of lymphedema using indocyanine green fluorescent lymphography after gynecologic cancer treatment.

BACKGROUND: Secondary lymphedema is defined as swelling of the limbs caused by retention of lymph after cancer therapy. We diagnosed lymphedema using indocyanine green (ICG) fluorescent lymphography and developed a classification based on 12 regional types of edema in the lower bodies, with the goal of improved understanding of the pathology. METHODS: The subjects were 72 consecutive female patients aged 25 to 88 years (mean, 54.5 years) with secondary lymphedema of the lower extremities and abdominal area. The traditional diagnosis of lymphedema was stages 0, 1, 2, 3 and 4 in 5, 11, 19, 24, and 13 patients, respectively. All patients were examined by ICG lymphography. RESULTS: Features of dermal backflow were noted in most patients after cancer therapy, and the incidence was particularly high after radiotherapy. Regional analysis of lymphedema was classified into 12 types (A to L, definitions are given for major categories). The number of patients (number receiving radiation therapy in parentheses) in each type were A, 1 (0); B, 3 (1); C, 13 (1); D, 1 (0); E, 2 (0); F, 0 (0); G, 1 (0); H, 7 (3); I, 13 (3); J, 6 (2); K, 20 (3); and L, 5 (2). CONCLUSIONS: The ICG test permits definite diagnosis of lymphedema at a very early stage and in mild cases. The regional analysis enables establishment of policies for conservative or surgical treatment (for example, lymphaticovenous anastomosis) for individual regions, thereby facilitating more effective lymphedema treatment.

Predictive lymphatic mapping: a method for mapping lymphatic channels in patients with advanced unilateral lymphedema using indocyanine green lymphography.

In severe lymphedema, indocyanine green lymphography cannot be used to map lymphatic channels before lymphaticovenular anastomosis (LVA) because linear lymphatics cannot be detected in a severely affected leg. Here, we describe a new method, which we refer to as predictive lymphatic mapping, to predict the location of lymphatics for anastomosis in unilateral lymphedema, thereby improving surgical accuracy and efficiency. The approach consists of marking anatomical landmarks and joining selected landmarks with fixed lines. The distance from these fixed lines to lymphatic channels mapped by indocyanine green lymphography in the unaffected leg is then measured, scaled up based on the difference in circumference between the legs, and transposed to the affected leg. To date, we have used this method in 5 cases of unilateral or asymmetric lymphedema of the lower extremities. In no cases have we failed to find a lymphatic channel suitable for LVA within a 2-cm incision. These results suggest that predictive lymphatic mapping is a useful additional tool for surgeons performing LVA under local anesthesia, which will help to improve the accuracy of incisions and the efficiency of surgery.

The effect of lymphatico-venous anastomosis for an intractable ulcer at the lower leg in a marked obese patient.

Secondary lymphedema occurs after trauma, cancer surgery, or obesity, and wounds in lymphedema can easily become intractable. We report positive results using lymphatico-venous anastomosis (LVA) to treat a post-traumatic lymph fistula and an intractable ulcer in a severely obese patient. A 41-year-old male (BMI 51.8), one year prior, had a traffic injury, and had an 18-cm contusion in his right leg. Six months later, lymph leakage in a 14 cm × 8 cm region and a 5 cm × 3 cm skin ulcer occurred in the center of the wound. We made a diagnosis of lymphedema resulting from obesity, accompanied with lymphorrhea and intractable ulcer. He was unable to reach his legs owing to obesity, making complex physical therapy impossible. We performed LVA under local anesthesia. The lymphorrhea healed 2 weeks after the operation and had not recurred 3 months after the operation. The leg lymphedema improved after the surgery without the compression therapy. In cases of intractable ulcers, suspected of being caused by lymphostasis, treatments indicated for lymphedema, for example LVA, may possibly allow satisfactory wound healing. © 2013 Wiley Periodicals, Inc. Microsurgery 34:64-67, 2014.

Rho-kinase inhibition prevents the progression of diabetic nephropathy by downregulating hypoxia-inducible factor 1α.

The small GTPase Rho and its effector Rho-kinase are involved in the pathogenesis of diabetic nephropathy. Accumulating evidence shows that hypoxia-inducible factor-1α (HIF-1α) is a key regulator of renal sclerosis under diabetic conditions. However, the interactions of Rho-kinase and HIF-1α in the development of renal dysfunction have not been defined. Here, we assessed whether Rho-kinase blockade attenuates HIF-1α induction and the subsequent fibrotic response using type 2 diabetic mice and cultured mesangial cells. Fasudil, a Rho-kinase inhibitor, reduced urinary albumin excretion, mesangial matrix expansion, and the expression of fibrotic mediators in db/db mice. Mechanistically, HIF-1α accumulation and the expression of its target genes that contribute to diabetic glomerulosclerosis were also prevented by fasudil in the renal cortex. In mesangial cells, Rho/Rho-kinase signaling was activated under hypoxic conditions. Further in vitro studies showed that pharmacological and genetic inhibition of Rho-kinase promoted proteasomal HIF-1α degradation, which subsequently suppressed HIF-1-dependent profibrotic gene expression by upregulation of prolyl hydroxylase 2. Thus, we found a previously unrecognized renoprotective mechanism for the effects of Rho-kinase inhibition and this could be a potential therapeutic target for the treatment of diabetic nephropathy.

[A case of early gastric cancer responding to adjuvant chemotherapy for treatment of colon cancer].

A 63-year-old man underwent colonoscopy owing to a positive fecal occult blood test, and he was diagnosed as having advanced sigmoid colon cancer. Esophagogastroduodenoscopy( EGD), performed as a preoperative examination, revealed an 8 mm early gastric cancer in the lower body of his stomach. We performed laparoscopic sigmoid colon resection and D3 lymphadenectomy first because the patient had advanced sigmoid colon cancer. The histopathological diagnosis was Stage II; however, vascular invasion was apparent. Some authors have reported that chemotherapy with 5-fluorouracil (FU) is effective against early gastric cancer; therefore, we administered postoperative adjuvant chemotherapy comprising uracil and tegafur( UFT)+Leucovorin( LV) tablets before ESD for early gastric cancer. Two months later, follow-up EGD showed that the gastric cancer had become flat and small. Five months later, it resembled a scar, and examination of a biopsy showed no malignant finding. We continued to administer chemotherapy to the patient for 6 months. Nine months after the discontinuation of chemotherapy, EGD showed only a scar and biopsy revealed no malignant finding.

Using indocyanine green fluorescent lymphography and lymphatic-venous anastomosis for cancer-related lymphedema.

Advances in cancer therapy have increased the importance of improvement of quality of life after cancer survival. Cancer-related lymphedema or secondary lymphedema that occurs after lymph node dissection in resection of tumors of abdominal visceral organs can impair quality of life. However, standard curative treatment for secondary lymphedema has not been established. This may be due to the lack of a method for early diagnosis of lymphedema, and because of selection of conservative treatment such as compression therapy to delay edema progression in many cases. To develop a curative approach, we have performed definite diagnosis of early-stage lymphedema using magnetic resonance imaging and an indocyanine green fluorescent lymphography, followed by surgical treatment with lymphatic-venous anastomosis using supermicrosurgery. Herein, we report the first case of secondary lymphedema in which we performed early diagnosis and surgery using these techniques and achieved an almost complete cure of lymphedema. We suggest that early diagnostic imaging and early microsurgery is the key of lymphedema treatment.

Low-invasive lymphatic surgery and lymphatic imaging for completely healed intractable pudendal lymphorrhea after gynecologic cancer treatment.

Lower limb lymphedema and an accompanying lymphatic fistula (lymphorrhea) occur as complications after gynecologic surgery to treat cancer. Herein, we report the case of a 68-year-old woman who underwent resection and radiotherapy because of uterine cervical cancer (stage 2a) 20 years previously. Left lower limb and pudendal lymphedema and continuous lymphorrhea developed soon after surgery. Conservative treatment was administered; however, the edema increased, and a pudendal lymphatic fistula and cellulitis developed repeatedly. Lymphovascular anastomosis (LVA) and lymph vessel ligation were performed after preoperative evaluation via lymphoscintigraphy and indocyanine green (ICG) lymphography. A radioisotope injected into the first interdigit pedal region flowed into the pudendal region via the inguinal lymph nodes at preoperative lymphoscintigraphy. Linear patterns were observed up to the half level of the crus, and stardust patterns occurred over the lower abdominal and pudendal regions at ICG lymphography. During surgery, ICG lymphography was also used to identify the site of the fistula. With the patient under local anesthesia, LVA was applied in the half crus and left inguinal regions, followed by ligation and division of lymph vessels flowing into the fistula. The region around the fistula was excised as a 1 × 3-cm tissue block. As of 5 months after surgery, no recurrence of lymphatic fistula or exacerbation of lymphedema has occurred. This case shows the effectiveness of preoperative ICG lymphography and lymphoscintigraphy followed by treatment via lymph vessel ligation and LVA for curative resolution of a lymphatic fistula.

Lymphatic-venous anastomosis for the radical cure of a large pelvic lymphocyst.

Therapeutic efficacy of lymphatic-venous anastomosis (LVA) has been shown, but expansion of the indication is desirable because LVA is a procedure with low invasiveness and is applicable over a wide area. This is the first reported case of intractable pelvic lymphocyst for which LVA was effective. LVA may be useful for pelvic lymphocyst at an early stage after cancer resection and lymph node dissection.

Lower limb lymphedema treated with lymphatico-venous anastomosis based on pre- and intraoperative icg lymphography and non-contact vein visualization: A case report.

Lymphatico-venous anastomosis (LVA) is used to resolve lymph retention in lymphedema. However, the postoperative outcome of lower limb lymphedema is poorer than that for upper limb lymphedema, because of the location lower than the heart level. Improvement of the therapeutic outcome requires application of as many anastomoses as possible in a limited operation time, particularly since there is a positive correlation between the number of anastomoses and the therapeutic effect of LVA. In this case, we described a method to increase the efficiency of lymphatico-venous anastomosis for bilateral severe lower limb lymphedema through efficient identification of lymph vessels and veins suitable for anastomosis using indocyanine green (ICG) contrast imaging and AccuVein, a noncontact vein visualization system, respectively. Ten LVAs were succeeded at seven incisions, and the operation time was 3 hours and 5 minutes. Accuvein can be used for identification of subcutaneous venules with a diameter of about 0.5-1.0 mm. We used this approach in surgery for a case of bilateral lower limb lymphedema, with a resultant improvement in the surgical outcome.

Antegrade and retrograde lymphatico-venous anastomosis for cancer-related lymphedema with lymphatic valve dysfuction and lymphatic varix.

In healthy people, no retrograde lymph flow occurs because of valves in collecting lymph vessels. However, in secondary lymphedema after lymph node dissection, lymph retention and lymphatic hypertension occurs and valvular dysfunction induces retrograde lymph flow. In this case reported, we focused on retrograde lymph flow and performed retrograde lymphatico-venous anastomosis (LVA) simultaneously with antegrade LVA. A 67-year-old Japanese woman had worsening edema in her right thigh and hip area for 3 years. She had previously undergone extended hysterectomy with lymph node dissection for endometrial cancer 8 years before. Indocyanine green test showed antegrade and retrograde lymph flow. Four LVAs were made in the right medial thigh and right lower abdominal area under local anesthesia. Lymphedema showed rapid improvement within 12 months and compression therapy was not required at 24 months after LVA. Retrograde LVA has a possibility of a more efficacy for secondary lymphedema.

Upper-limb lymphedema treated aesthetically with lymphaticovenous anastomosis using indocyanine green lymphography and noncontact vein visualization.

We have described a procedure to minimize surgical wounds, in which lymph vessels and skin venules are identified by indocyanine green (ICG) lymphography and the AV300 noncontact visualization system (AccuVein, Cold Spring Harbor, NY), respectively. This approach allows accurate decisions regarding sites of incision for lymphatic venous anastomosis (LVA). This method was applied in a patient with right upper-limb lymphedema after breast cancer therapy. The low-invasive procedure can be used before and during surgery. The incision size is minimal, and the incision site is at the joint area. Thus, we aim to establish this approach as a standard method for identifying lymph vessels and veins that are suitable for LVA. This innovative vascular-imaging machine makes LVA less invasive and more effective without side effects.

The Selective Serotonin 2A Receptor Antagonist Sarpogrelate Prevents Cardiac Hypertrophy and Systolic Dysfunction via Inhibition of the ERK1/2-GATA4 Signaling Pathway.

Drug repositioning has recently emerged as a strategy for developing new treatments at low cost. In this study, we used a library of approved drugs to screen for compounds that suppress cardiomyocyte hypertrophy. We identified the antiplatelet drug sarpogrelate, a selective serotonin-2A (5-HT2A) receptor antagonist, and investigated the drug's anti-hypertrophic effect in cultured cardiomyocytes and its effect on heart failure in vivo. Primary cultured cardiomyocytes pretreated with sarpogrelate were stimulated with angiotensin II, endothelin-1, or phenylephrine. Immunofluorescence staining showed that sarpogrelate suppressed the cardiomyocyte hypertrophy induced by each of the stimuli. Western blotting analysis revealed that 5-HT2A receptor level was not changed by phenylephrine, and that sarpogrelate suppressed phenylephrine-induced phosphorylation of ERK1/2 and GATA4. C57BL/6J male mice were subjected to transverse aortic constriction (TAC) surgery followed by daily oral administration of sarpogrelate for 8 weeks. Echocardiography showed that 5 mg/kg of sarpogrelate suppressed TAC-induced cardiac hypertrophy and systolic dysfunction. Western blotting revealed that sarpogrelate suppressed TAC-induced phosphorylation of ERK1/2 and GATA4. These results indicate that sarpogrelate suppresses the development of heart failure and that it does so at least in part by inhibiting the ERK1/2-GATA4 signaling pathway.

The change of antizyme inhibitor expression and its possible role during mammalian cell cycle.

Antizyme inhibitor (AIn), a homolog of ODC, binds to antizyme and inactivates it. We report here that AIn increased at the G1 phase of the cell cycle, preceding the peak of ODC activity in HTC cells in culture. During interphase AIn was present mainly in the cytoplasm and turned over rapidly with the half-life of 10 to 20 min, while antizyme was localized in the nucleus. The level of AIn increased again at the G2/M phase along with ODC, and the rate of turn-over of AIn in mitotic cells decreased with the half-life of approximately 40 min. AIn was colocalized with antizyme at centrosomes during the period from prophase through late anaphase and at the midzone/midbody during telophase. Thereafter, AIn and antizyme were separated and present at different regions on the midbody at late telophase. AIn disappeared at late cytokinesis, whereas antizyme remained at the cytokinesis remnant. Reduction of AIn by RNA interference caused the increase in the number of binucleated cells in HTC cells in culture. These findings suggested that AIn contributed to a rapid increase in ODC at the G1 phase and also played a role in facilitating cells to complete mitosis during the cell cycle.

Investigation of the effects of urea cycle amino acids on the expression of ALB and CEBPB in the human hepatocellular carcinoma cell line FLC-4.

Cell lines are powerful tools for research into liver function at the molecular level. However, they are generally unsuitable for rigorously assessing the effects of amino acid composition, because many lines require serum-containing medium for their maintenance. Here, we aimed to investigate the effects of ornithine and arginine, which are included in the characteristic metabolic process in hepatocyte, on a human hepatoma-derived cell line (FLC-4) that can be cultured in serum-free medium. FLC-4 cells were cultured under the following three conditions: + ornithine/ - arginine, - ornithine/ - arginine, and -ornithine/ + arginine. Albumin expression evaluated by quantitative polymerase chain reaction and enzyme-linked immunosorbent assay and showed no obvious differences based on the presence of ornithine or arginine. However, the mRNA levels of two liver-enriched transcription factors (CEBPB and HNF1A), which are involved in regulating albumin expression, were significantly higher in cells grown in medium-containing arginine than that in cells grown in ornithine-containing medium. Western blotting showed that the levels both activating and inhibitory C/EBPß isoforms were significantly increased in cells grown in arginine medium. Furthermore, we have found that depletion of both ornithine and arginine, the polyamine sources, in the medium did not cause polyamine deficiency. When ornithine and arginine were depleted, albumin production was significantly reduced at the mRNA level, CEBPB mRNA levels were increased, and the level of activating form of C/EBPß was increased. The results of this study suggest that in hepatocyte, these two amino acids might have different functions, and because of which they elicit disparate cellular responses.