Intravenous ketamine infusion for a patient with treatment-resistant major depression: a 10-month follow-up.

WHAT IS KNOWN AND OBJECTIVE: Ketamine in a subanaesthetic dose has been shown to produce rapid antidepressant effects. Here, we describe a long-term follow-up case of a Korean patient with severe major depression who received repeated ketamine intravenous therapy (KIT). CASE DESCRIPTION: A 49-year-old woman with a 6-year history of treatment-resistant major depression was given KIT once every 1 or 2 weeks over 10 months, for a total of 36 treatments. Her mood stabilized, and she showed a nearly 50% reduction in the severity of her depressive symptom. WHAT IS NEW AND CONCLUSION: Long-term repeated KIT may be an option for alleviating treatment-resistant and relapsing major depression. Further research and large clinical trials are needed on the optimum KIT protocol, including dose, dosing interval, total number of treatments and when to stop.

Immunologic characteristics of human gingival fibroblasts in response to oral bacteria.

BACKGROUND AND OBJECTIVE: There is ample evidence that gingival fibroblasts (GFs) participate in the immune response to oral bacteria and serve as immune-regulatory cells. The objective of this study was to investigate the innate immune response of GFs to oral bacteria. MATERIAL AND METHODS: Human GFs were cocultured with relatively less-pathogenic (Leptotrichia wadei, Fusobacterium nucleatum and Campylobacter gracilis) and pathogenic red-complex bacteria. The expression of mRNA for antimicrobial peptides [AMPs; namely human beta defensins (HBDs)], chemokines with antimicrobial activity [chemokine C-X-C motif (CXCL)10, CXCL11 and chemokine C-C motif ligand 20 (CCL20)] and proinflammatory mediators [interleukin (IL)6 and IL8] and the levels of CXCL11, CCL20, IL-6 and IL-8 accumulated in supernatants were analyzed using real-time PCR and ELISA, respectively. The proteolytic activities of CXCL11, CCL20, IL-6 and IL-8 produced by six species of bacteria were also determined. RESULTS: The relatively less-pathogenic bacteria strongly up-regulated the expression of antimicrobial chemokines and proinflammatory mediators, whereas the red-complex bacteria stimulated low levels, or often suppressed, expression of these factors. Regarding the regulation of AMPs, the inhibition of HBD3, HBD106 and HBD107 mRNAs by Porphyromonas gingivalis was noticeable; however, differences between the two bacterial groups were not conspicuous. Differential degradation of proteins by the six bacterial species was observed: P. gingivalis and Treponema denticola degraded proteins well, whereas the other species degraded proteins to a relatively lower degree. CONCLUSION: The invasion of red-complex bacteria into gingival connective tissue can suppress the immune response of GFs and can be a source of persistent infection in connective tissue.

Obesity is inversely related to the risks of dental caries in Korean adults.

OBJECTIVE: This study investigated whether obesity is linked with dental caries. This study hypothesized that obesity may influence the number or prevalence of dental caries in Korean adults. SUBJECTS AND METHODS: Data were derived from Korea National Health and Nutrition Examination Survey performed at 2008-2010. Lifestyle, sociodemographic, and biochemical variables were analyzed. Indices related to obesity, sarcopenia, and metabolic syndrome were investigated. Finally, caries index and oral health behaviors were included for the analysis. RESULTS: The caries index was inversely associated with increasing body mass index (BMI) and body fat quartile (all p-values <.01). Subjects with high waist circumference who met the inclusion criteria of metabolic syndrome were less likely to have decayed tooth (p-value = .0009). Subjects with a BMI of 25 or more showed about 20% less prevalence of dental caries experience than normal individuals with a BMI of 18.5 to 23 glucose (odds ratio [95% confidence intervals] = 0.808 [0.684-0.956]). Similarly, subjects with total body fat in the highest quartile revealed about 20% less prevalence of caries experiences that those with body fat proportion in the lowest quartile (0.84 [0.672,1.049]). CONCLUSIONS: Obesity was inversely associated with occurrence or severity of dental caries in Korean adults.

Type 2 diabetes as a risk indicator for dental caries in Korean adults: the 2011-2012 Korea national health and nutrition examination survey.

OBJECTIVES: The associations between type 2 diabetes (T2D) and untreated dental caries was examined. This study hypothesized that there would be a positive association between T2D and the prevalence of decayed permanent teeth (DT) in representative Korean adults. METHODS: The information was derived from the Korea National Health and Nutrition Examination Survey conducted in 2011-2012. Sociodemographic and lifestyle variables, anthropometric and biochemical status, metabolic health and glucose tolerance status, oral health behaviors, and dental caries index were evaluated. RESULTS: The number of DT had a positive association with degree of fasting plasma glucose (FPG) level, and glycated hemoglobin (HbA1c) (p-value = 0.045 and 0.007, respectively). The levels of FPG and HbA1c increased with the number of DT (p for trend = 0.009 and 0.004, respectively). The prevalence of untreated caries uncontrolled T2D participants was about 26% higher than those with normal glucose tolerance levels after adjusting for potential confounders including diets and socioeconomic status (OR [95% CI] = 1.26 [1.02, 1.56]). CONCLUSIONS: T2D is an independent risk indicator for untreated caries in Korean adults.

Mitochondrial modulation decreases the bortezomib-resistance in multiple myeloma cells.

Multiple myeloma (MM) is an incurable hematological malignancy that causes most patients to eventually relapse and die from their disease. The 20S proteasome inhibitor bortezomib has emerged as an effective drug for MM treatment; however, intrinsic and acquired resistance to bortezomib has already been observed in MM patients. We evaluated the involvement of mitochondria in resistance to bortezomib-induced cell death in two different MM cell lines (bortezomib-resistant KMS20 cells and bortezomib-sensitive KMS28BM cells). Indices of mitochondrial function, including membrane potential, oxygen consumption rate and adenosine-5'-triphosphate and mitochondrial Ca(2+) concentrations, were positively correlated with drug resistance of KMS cell lines. Mitochondrial genes including CYPD, SOD2 and MCU were differentially expressed in KMS cells. Thus, changes in the expression of these genes lead to changes in mitochondrial activity and in bortezomib susceptibility or resistance, and their combined effect contributes to differential sensitivity or resistance of MM cells to bortezomib. In support of this finding, coadministration of bortezomib and 2-methoxyestradiol, a SOD inhibitor, rendered KMS20 cells sensitive to apoptosis. Our results provide new insight into therapeutic modalities for MM patients. Studying mitochondrial activity and specific mitochondrial gene expression in fresh MM specimens might help predict resistance to proapoptotic chemotherapies and inform clinical decision-making.

Three-dimensional soft tissue changes after reduction malarplasty in female patients.

The aim of this study was to determine the three-dimensional soft tissue changes after reduction malarplasty. Soft tissue changes relative to the amount of movement of the zygomatic bone were studied. Pre- and postoperative cone beam computed tomography images of 21 female patients were superimposed. The anterior-most point of the body osteotomy (point A), arch osteotomy site (point D), and points dividing line A-D into thirds (points B and C) were marked on lateral view images. The vertical distances from the midsagittal line to the centre of the zygomatic bone and the outer prominence of the soft tissue were measured on the coronal view of each image. The proportion of the change in soft tissue to that of the bone before and after surgery was calculated for each point. The relationship between body mass index and the soft tissue change ratio, and the differences in soft tissue changes at each point were analysed. Mean soft tissue changes for points A, B, C, and D were 53.43%, 66.66%, 63.67%, and 57.23%, respectively. The amount of soft tissue change at point B was greater than that at points A and D, which were osteotomy sites. There was no statistical correlation between body mass index and the soft tissue change ratio at each point.

Mutant DLX 3 disrupts odontoblast polarization and dentin formation.

Tricho-dento-osseous (TDO) syndrome is an autosomal dominant disorder characterized by abnormalities in the thickness and density of bones and teeth. A 4-bp deletion mutation in the Distal-Less 3 (DLX3) gene is etiologic for most cases of TDO. To investigate the in vivo role of mutant DLX3 (MT-DLX3) on dentin development, we generated transgenic (TG) mice expressing MT-DLX3 driven by a mouse 2.3 Col1A1 promoter. Dentin defects were radiographically evident in all teeth and the size of the nonmineralized pulp was enlarged in TG mice, consistent with clinical characteristics in patients with TDO. High-resolution radiography, microcomputed tomography, and SEM revealed a reduced zone of mineralized dentin with anomalies in the number and organization of dentinal tubules in MT-DLX3 TG mice. Histological and immunohistochemical studies demonstrated that the decreased dentin was accompanied by altered odontoblast cytology that included disruption of odontoblast polarization and reduced numbers of odontoblasts. TUNEL assays indicated enhanced odontoblast apoptosis. Expression levels of the apoptotic marker caspase-3 were increased in odontoblasts in TG mice as well as in odontoblastic-like MDPC-23 cells transfected with MT-DLX3 cDNA. Expression of Runx2, Wnt 10A, and TBC1D19 colocalized with DLX3 expression in odontoblasts, and MT-DLX3 significantly reduced expression of all three genes. TBC1D19 functions in cell polarity and decreased TBC1D19 expression may contribute to the observed disruption of odontoblast polarity and apoptosis. These data indicate that MT-DLX3 acts to disrupt odontoblast cytodifferentiation leading to odontoblast apoptosis, and aberrations of dentin tubule formation and dentin matrix production, resulting in decreased dentin and taurodontism. In summary, this TG model demonstrates that MT-DLX3 has differential effects on matrix production and mineralization in dentin and bone and provides a novel tool for the investigation of odontoblast biology.

In vivo impact of a 4 bp deletion mutation in the DLX3 gene on bone development.

Distal-less 3 (DLX3) gene mutations are etiologic for Tricho-Dento-Osseous syndrome. To investigate the in vivo impact of mutant DLX3 on bone development, we established transgenic (TG) mice expressing the c.571_574delGGGG DLX-3 gene mutation (MT-DLX3) driven by a mouse 2.3 Col1A1 promoter. Microcomputed tomographic analyses demonstrated markedly increased trabecular bone volume and bone mineral density in femora from TG mice. In ex vivo experiments, TG mice showed enhanced differentiation of bone marrow stromal cells to osteoblasts and increased expression levels of bone formation markers. However, TG mice did not show enhanced dynamic bone formation rates in in vivo fluorochrome double labeling experiments. Osteoclastic differentiation capacities of bone marrow monocytes were reduced in TG mice in the presence of osteoclastogenic factors and the numbers of TRAP(+) osteoclasts on distal metaphyseal trabecular bone surfaces were significantly decreased. TRACP 5b and CTX serum levels were significantly decreased in TG mice, while IFN-gamma levels were significantly increased. These data demonstrate that increased levels of IFN-gamma decrease osteoclast bone resorption activities, contributing to the enhanced trabecular bone volume and mineral density in these TG mice. These data suggest a novel role for this DLX-3 mutation in osteoclast differentiation and bone resorption.

Comparison of endoscopic ultrasonography and conventional endoscopy for prediction of depth of tumor invasion in early gastric cancer.

BACKGROUND AND STUDY AIMS: This study aimed to compare the diagnostic accuracy of endoscopic ultrasonography (EUS) with that of conventional endoscopy for staging depth of invasion (T staging) in early gastric cancer. PATIENTS AND METHODS: A total of 955 patients with suspected early gastric cancer were prospectively registered. EUS staging was carried out prospectively by a single endoscopist using either miniprobe or radial EUS depending on the endoscopic appearance of the tumor. Conventional endoscopy staging was performed retrospectively by consensus between two endoscopists who were blinded to the EUS staging. Conventional endoscopy staging was conducted on the basis of endoscopic features such as surface nodularity and fold convergence. Patients underwent either surgical (n = 586) or endoscopic resection (n = 369) with curative intent. The staging accuracy of each test was compared with the pathological staging of the resected specimen. RESULTS: The presence of a T1m tumor was histologically confirmed in 644 cases (67.4 %) and that of a T1sm tumor in 311 cases (32.6 %). The overall accuracy of EUS staging was 67.4 % (644 / 955) and that of conventional endoscopy staging was 73.7 % (704 / 955) ( P < 0.001). The accuracy of miniprobe EUS was significantly higher than that of radial EUS (79.5 % vs. 59.6 %, P < 0.001), but did not differ significantly from that of conventional endoscopy (79.0 %). CONCLUSIONS: EUS does not substantially impact on pretreatment T staging of patients with early gastric cancer compared with conventional endoscopy. Therefore, EUS may not be necessary routinely, and conventional endoscopy may be sufficient for determining the optimal therapeutic strategy, especially in relation to endoscopic resection for early gastric cancer.

Allopurinol-induced aplastic anemia in a patient with chronic kidney disease.

Aplastic anemia is a rare complication of allopurinol use. We report an unusual case of aplastic anemia associated with allopurinol therapy for hyperuricemia in a patient with chronic kidney disease. A 37-year-old female patient diagnosed with Stage III chronic kidney disease was admitted with pancytopenia. She had a history of taking allopurinol for 5 months. Her bone marrow showed extremely decreased cellularity (< 20%) and there was no malignant cell infiltration. She was free of infections, including parvovirus B19, cytomegalovirus and Epstein-Barr virus. These results suggested a diagnosis of aplastic anemia. Allopurinol was discontinued immediately and treatment with blood transfusions and prednisolone was begun. After 6 months, the bone marrow cellularity improved to approximately 70%. Recently, it was suggested that decreased activity of multidrug resistance P-glycoprotein may play a role in acquired aplastic anemia. So we measured the inhibitory effect of allopurinol and oxypurinol on P-glycoprotein activity. But neither allopurinol nor oxypurinol inhibited P-glycoprotein activity.

Involvement of Mrp2/MRP2 in the species different excretion route of benzylpenicillin between rat and human.

1. The purpose of this study was to investigate the involvement of rat Mrp2 and human MRP2 in benzylpenicillin transport using canalicular liver plasma membrane (cLPM) vesicles isolated from Sprague-Dawley or Easai hyperbilirubinemic (EHBR) rats, and MDCKII cells overexpressing MRP2. 2. The adenosine triphosphate (ATP)-dependent uptake of benzylpenicillin and oestradiol-17beta-D-glucuronide (E(2)17betaG), a representative substrate for Mrp2, into EHBR-cLPM vesicles was decreased relative to that seen with control-cLPM vesicles, which may reflect the absence of Mrp2 in the EHBR. The ATP-dependent uptake of taurocholate, which is not a substrate for Mrp2, was similar in both control and EHBR-cLPM vesicles. The concentration dependence of ATP-dependent benzylpenicillin uptake was reflected in a K(m) of 44.0 microM and a V(max) of 508.4 pmol mg(-1) min(-1). Additional inhibition studies using E(2)17betaG and methotrexate as representative substrates for Mrp2/MRP2 demonstrated the involvement of rat Mrp2, but not human MRP2, in benzylpenicillin efflux. Benzylpenicillin appears not to be a substrate for or inhibitor of other human efflux transporters such as MDR1, MRP1, MRP3, or BCRP. 3. In conclusion, rat Mrp2, but not human MRP2, plays an important role in ATP-dependent benzylpenicillin uptake in the bile canalicular membrane, which may explain why biliary excretion of benzylpenicillin is high in the rat but negligible in humans.

Decreased urinary secretion of belotecan in folic acid-induced acute renal failure rats due to down-regulation of Oat1 and Bcrp.

The effects of folic acid-induced acute renal failure on the renal excretion of belotecan were investigated in rats after intravenous administration. Both glomeruli and renal tubules were seriously damaged by folic acid-induced acute renal failure. The renal excretion clearance, CLr, of belotecan was significantly decreased by folic acid-induced acute renal failure. Furthermore, glomerular filtration rate and secretion clearance of the drug were dramatically decreased by folic acid-induced acute renal failure. In vivo renal uptake of belotecan was inhibited by p-aminohippurate, whereas renal excretion was inhibited by GF120918, but not by verapamil and bromosulphalein. This indicates that Oat1/3 and Bcrp are involved in the renal uptake and urinary excretion of belotecan, respectively. Both mRNA and protein levels of Oat1, Oat3 and Bcrp were significantly decreased in folic acid-induced acute renal failure rats. Based on the finding that belotecan is a substrate of OAT1 but not of OAT3, the decrease in CLr of belotecan in folic acid-induced acute renal failure could, therefore, mainly be attributed to the down-regulation of Oat1 and Bcrp, in addition to the decrease in glomerular filtration rate.

Dual-energy X-ray absorptiometry overestimates bone mineral density of the lumbar spine in persons with spinal cord injury.

BACKGROUND: Bone mineral density (BMD) of the lumbar spine (L-spine) has been reported to be normal or increased in persons with chronic spinal cord injury (SCI). OBJECTIVE: To determine BMD of the L-spine by dual-energy X-ray absorptiometry (DXA) and quantitative computerized tomography (qCT) in men with chronic SCI compared with able-bodied controls. DESIGN: Cross-sectional, comparative study. SETTING: Clinical research unit, Veterans Affairs Medical Center, Bronx, NY, USA and Kessler Institute of Rehabilitation, West Orange, NJ, USA. METHODS: Measurements of the L-spine were made in 20 men with SCI and compared with 15 able-bodied controls. The DXA images were acquired on a GE Lunar DPX-IQ. The qCT images of the L-spine were acquired on a Picker Q series computerized tomographic scanner. RESULTS: The mean ages for the SCI and control groups were 44+/-13 vs 42+/-9 years, and the duration of injury of the group with SCI was 14+/-11 years. There were no significant differences between the SCI and control groups for L-spine DXA BMD (1.391+/-0.210 vs 1.315+/-0.178 g/m(2)) or for L-spine DXA T-score (1.471+/-1.794 vs 0.782+/-1.481). L-spine qCT BMD was significantly lower in the SCI compared with the control group (1.296+/-0.416 vs 1.572+/-0.382 g/m(2), P=0.05); the T-score approached significance (-1.838+/-1.366 vs -0.963+/-1.227, P=0.059). Subjects with moderate degenerative joint disease (DJD) had significantly higher T-scores by DXA than those without or with mild DJD. CONCLUSION: Individuals with SCI who have moderate to severe DJD may have bone loss of the L-spine that may be underestimated by DXA, reducing awareness of the risk of fracture.

Metabolic syndrome and visceral obesity as risk factors for reflux oesophagitis: a cross-sectional case-control study of 7078 Koreans undergoing health check-ups.

BACKGROUND: Obesity has been associated with reflux oesophagitis. However, the relationship between metabolic syndrome characterised by visceral obesity and reflux oesophagitis is unclear. AIM: To investigate whether metabolic syndrome or visceral obesity is a risk factor for reflux oesophagitis. METHODS: A cross-sectional study of 7078 subjects undergoing upper endoscopy during health check-ups was conducted (3539 patients with reflux oesophagitis vs age- and sex-matched controls). We further analysed according to categories of visceral adipose tissue and subcutaneous adipose tissue area with 750 cases and age-, sex- and waist circumference-matched controls who underwent abdominal CT scan. RESULTS: The prevalence of metabolic syndrome was higher in cases than controls (26.9% vs 18.5%, p<0.001). Multivariate analysis demonstrated that metabolic syndrome is associated with reflux oesophagitis (odds ratio (OR) = 1.42; 95% confidence interval (CI), 1.26 to 1.60). Among the individual components of metabolic syndrome, waist circumference (OR = 1.47; 95% CI, 1.30 to 1.65) and triglyceride (OR = 1.20; 95% CI, 1.05 to 1.36) independently increased the risk for reflux oesophagitis. On sub-analysis, cases showed higher mean visceral adipose tissue area (cm(2)) (136.1 (SD 57.8) vs 124.0 (SD 54.7), p<0.001) and subcutaneous adipose tissue area (cm(2)) (145.9 (SD 56.8) vs 133.5 (SD 50.7), p<0.001). However, only visceral adipose tissue area was an independent risk factor for reflux oesophagitis after adjusting for multiple confounders including smoking, alcohol, body mass index (BMI) and subcutaneous adipose tissue area (OR = 1.60; 95% CI, 1.03 to 2.48, lowest quartile vs highest quartile). CONCLUSIONS: Metabolic syndrome was associated with reflux oesophagitis. Abdominal obesity, especially visceral obesity, was an important risk factor for reflux oesophagitis.

Genetic variants of organic cation transporter 2 (OCT2) significantly reduce metformin uptake in oocytes.

1. The authors sought to evaluate the contribution of organic cation transporters (OCTs) to the renal tubular transport of metformin using LLC-PK1 cells as an in vitro model for the renal proximal tubule, and to investigate the effects of three non-synonymous genetic variants of OCT2 on the transport activity of metformin in vitro using an oocyte over-expression system. 2. The basolateral-to-apical transport of metformin was significantly greater than the apical-to-basolateral transport and showed concentration dependency with the kinetic parameters: maximum transport rate (V(max)), 922 pmol min(-1) per 5 x 10(5) cells; Michaelis-Menten constant (K(m)), 393 microM; intrinsic clearance (CL(int)), 2.35 microl min(-1) per 5 x 10(5) cells; and diffusion constant (K(d)), 0.33 microl min(-1) per 5 x 10(5) cells. The basolateral-to-apical transport of metformin was inhibited by phenoxybenzamine, an inhibitor of OCTs, but not by cyclosporine A, MK571, or fumitremorgin C, which are inhibitors of P-glycoprotein, multidrug resistance proteins (MRPs), and breast cancer resistance protein (BCRP), respectively, suggesting that OCTs play a role in renal tubular secretion of metformin. 3. Metformin uptake was much greater in oocytes expressing OCT2-wild type (OCT2-WT) than OCT1-WT compared with uptake in water-injected oocytes. Uptake was significantly decreased in oocytes expressing OCT2-T199I, -T201M, and -A270S compared with that in OCT2-WT, suggesting that metformin is a better substrate for OCT2 than for OCT1 and that the amino acid-substituted variants of OCT2 cause a functional decrease in metformin uptake. 4. In conclusion, the genetic variants of OCT2 (OCT2-T199I, -T201M, and -A270S) decreased the transport activity of metformin and thus may contribute to the inter-individual variation in metformin disposition as OCT2 plays a pivotal role in renal excretion, the major disposition route of metformin.

Omeprazole may be superior to famotidine in the management of iatrogenic ulcer after endoscopic mucosal resection: a prospective randomized controlled trial.

BACKGROUND: Acid suppressing agents are widely used to treat the iatrogenic ulcers following endoscopic mucosal resection for gastric neoplasms. However, the relative merits of proton pump inhibitor or histamine(2)-receptor antagonist for endoscopic mucosal resection-induced ulcers are not known. AIM: To prospectively compare omeprazole and famotidine for the healing of endoscopic mucosal resection-induced ulcers and for bleeding control. METHODS: After endoscopic mucosal resection, patients were randomly assigned to omeprazole (20 mg/day) or to famotidine (40 mg/day) group for a 28-day treatment period. The ulcer sizes and stages, bleeding rates and ulcer-related symptoms were compared. RESULTS: A total of 100 patients were randomized equally to each group. Forty-one patients in each group were finally compared. The two groups were comparable in terms of baseline characteristics. Twenty-eight days after treatment, the two groups were not different with respect to ulcer stage (P = 0.137) or ulcer reduction ratio (P = 0.380). No difference was observed with respect to ulcer-related symptoms (P = 0.437) and no bleeding episode occurred in any of the 82 patients. In subgroup that underwent endoscopic submucosal dissection, fewer patients in the omeprazole group showed active ulcers than those in the famotidine group (P = 0.035). CONCLUSION: Our results demonstrate that omeprazole may be superior to famotidine for iatrogenic ulcers following endoscopic mucosal resection, especially for large ulcers.

Effect of growth and sodium butyrate on brush border membrane-associated hydrolases in human colorectal cancer cell lines.

The activities of brush border membrane-associated hydrolases such as alkaline phosphatase (Alkpase), aminopeptidase, dipeptidyl aminopeptidase IV (DAP-IV), sucrase, lactase, and trehalase were studied in 14 different human colorectal cancer cell lines. The effect of sodium butyrate, a known differentiating agent, and cell growth on the activities of these enzymes was also examined. All 14 cell lines exhibited brush border membrane enzyme activities, and in general, the activity of Alkpase, aminopeptidase, and DAP-IV was much higher than the disaccharidases. However, the specific enzyme activities varied among different cell lines. The induction of Alkpase activity by sodium butyrate occurred in most of the 14 cell lines (2- to 123-fold), while induction of the other enzyme activities was observed in several (1.5- to 3.5-fold). In some instances, butyrate caused a decrease in enzyme activity. There was no statistically significant correlation between the induction of Alkpase activity and that of other enzyme activities by sodium butyrate. Levels of aminopeptidase and DAP-IV activity were found to be dependent on cell density and increased 3- to 4-fold by the tenth day in most of the cell lines. Sodium butyrate altered the subcellular distribution pattern of the disaccharidases, causing a significant increase in activity associated with the soluble (cytoplasmic) fraction. Other enzymes such as Alkpase and DAP-IV continued to be predominantly associated with the membrane fraction in butyrate-treated cells. These data suggest that brush border membrane hydrolase activity and the effect of sodium butyrate may provide useful information regarding the differentiation of human colorectal cancer cells.

Antitumor effects of granulocyte-macrophage colony-stimulating factor production by melanoma cells.

The use of immunomodulating gene therapy in the treatment of malignant disease is under intensive investigation. In this study, we examined the potential of melanoma-derived granulocyte-macrophage colony-stimulating factor (GM-CSF) to inhibit melanoma progression in a murine model. The HGH18 murine melanoma cell line was transfected with the murine GM-CSF gene in a SV40 expression vector that resulted in melanoma clones that produced varying amounts of GM-CSF. Syngeneic mice inoculated s.c. with HFH18 parental melanoma cells or HFH18 cells transfected with the GM-CSF gene n the noncoding 3'-5' orientation [HFH18/GM-CSF(-) cells] develop large tumors that reach a mean tumor volume of 3300 mm3 by day 30. In contrast, animals inoculated with two melanoma clones producing high levels of GM-CSF [HFH18/GM-CSF(++) and HFH18/GM-CSF(+ + +)] either completely reject the tumor cells or develop tumors with a mean volume of only 40 mm3. In comparison, animals inoculated with a melanoma clone producing low levels of GM-CSF [HFH18/GM-CSF(+)] develop large tumors averaging 2000 mm3, thus demonstrating a dose-response effect of tumor inhibition by melanoma-derived GM-CSF. Additionally, vaccination with irradiated GM-CSF-producing melanoma cells conferred optimal immunogenicity against a subsequent challenge with HFH18 cells. Tissue sections from excised GM-CSF-producing tumor cell inoculation sites but not from HFH18 parental or HFH18/GM-CSF(-) inoculation sites demonstrate a dense inflammatory infiltrate composed of neutrophils, tissue macrophages, and numerous CD4- and CD8-positive lymphocytes but few melanoma cells. Large numbers of dendritic cells and cells expressing the B7-2 costimulatory molecule are detected only within HFH18/GM-CSF(+ + +) melanoma inoculation sites. Our results lend further support to clinical trials of GM-CSF gene therapy in the treatment of advanced malignant melanoma, possibly by the recruitment of dendritic antigen-presenting cells.

Peroxiredoxin II promotes hepatic tumorigenesis through cooperation with Ras/Forkhead box M1 signaling pathway.

The current study was carried out to define the involvement of Peroxiredoxin (Prx) II in progression of hepatocellular carcinoma (HCC) and the underlying molecular mechanism(s). Expression and function of Prx II in HCC was determined using H-ras(G12V)-transformed HCC cells (H-ras(G12V)-HCC cells) and the tumor livers from H-ras(G12V)-transgenic (Tg) mice and HCC patients. Prx II was upregulated in H-ras(G12V)-HCC cells and H-ras(G12V)-Tg mouse tumor livers, the expression pattern of which highly similar to that of forkhead Box M1 (FoxM1). Moreover, either knockdown of FoxM1 or site-directed mutagenesis of FoxM1-binding site of Prx II promoter significantly reduced Prx II levels in H-ras(G12V)-HCC cells, indicating FoxM1 as a direct transcription factor of Prx II in HCC. Interestingly, the null mutation of Prx II markedly decreased the number and size of tumors in H-ras(G12V)-Tg livers. Consistent with this, knockdown of Prx II in H-ras(G12V)-HCC cells reduced the expression of cyclin D1, cell proliferation, anchorage-independent growth and tumor formation in athymic nude mice, whereas overexpression of Prx II increased or aggravated the tumor phenotypes. Importantly, the expression of Prx II was correlated with that of FoxM1 in HCC patients. The activation of extracellular signal-related kinase (ERK) pathway and the expression of FoxM1 and cyclin D1 were highly dependent on Prx II in H-ras(G12V)-HCC cells and H-ras(G12V)-Tg livers. Prx II is FoxM1-dependently-expressed antioxidant in HCC and function as an enhancer of Ras(G12V) oncogenic potential in hepatic tumorigenesis through activation of ERK/FoxM1/cyclin D1 cascade.

Distribution and biosynthesis of aminopeptidase N and dipeptidyl aminopeptidase IV in rat small intestine.

The regional, cellular and subcellular distribution patterns of aminopeptidase N and dipeptidyl aminopeptidase IV were examined in rat small intestine. Aminopeptidase N of brush border membrane had maximal activity in the upper and middle intestine, while dipeptidyl aminopeptidase IV had a more uniform distribution profile with relatively high activity in the ileum. Along the villus and crypt cell gradient, the activity of both enzymes was maximally expressed in the mid-villus cells. However there was substantial dipeptidyl aminopeptidase IV activity in the crypt cells. Both enzymes were primarily associated with brush border membranes in all segments, however, in the proximal intestine, a significant amount of dipeptidyl aminopeptidase IV activity was associated with the cytosol fraction. The cytosol and brush border membrane forms of dipeptidyl aminopeptidase IV were immunologically identical and had the same electrophoretic mobility on disc gels. In contrast, the soluble and brush border membrane-bound forms of aminopeptidase N were immunologically distinct. When the total amount of aminopeptidase N and dipeptidyl aminopeptidase IV was determined by competitive radioimmunoassay, there were no regional or cellular differences in specific activity (enzyme activity/mg of enzyme protein) of either enzyme in brush border membrane and homogenate. The specific activity of both enzymes in a purified Golgi membrane fraction as measured by radioimmunoassay was about half that of the brush border membrane fraction. These results suggest that (1) aminopeptidase N and dipeptidyl aminopeptidase IV have different regional, cellular and subcellular distribution patterns; (2) there are enzymatically inactive forms of both enzymes present in a constant proportion to active molecules and that (3) a two-fold activation of precursor enzyme forms occurs during transfer from the Golgi membranes to the brush border membranes.