Predictive impact of elevated serum level of IL-18 for early renal dysfunction in type 2 diabetes: an observational follow-up study.

AIMS/HYPOTHESIS: The early identification of type 2 diabetic patients at risk of developing microalbuminuria-an independent risk factor for renal and cardiovascular diseases-is important to improve the patients' outcomes. We investigated whether serum levels of IL-18, a proinflammatory cytokine, were a predictor of early renal dysfunction. MATERIALS AND METHODS: A total of 249 Japanese type 2 diabetic patients without overt proteinuria were enrolled in an observational follow-up study (median follow-up 7 years), and their stage of diabetic nephropathy was classified and their estimated glomerular filtration rate (eGFR) was calculated annually. RESULTS: At baseline, serum levels of IL-18 were higher in subjects with microalbuminuria (n = 76) than in those with normoalbuminuria (n = 173). Elevated serum levels of IL-18 were associated with the progression of nephropathy to a higher stage in normoalbuminuric subjects (118 [interquartile range 91-159] ng/l vs 155 [interquartile range 121-205] ng/l, p = 0.003), but not in microalbuminuric subjects (154 [interquartile range 113-200] ng/l vs 160 [interquartile range 101-190] ng/l, p = 0.50). The adjusted risk for developing microalbuminuria was 3.6 (95% CI 1.2-10.4) in normoalbuminuric subjects with serum IL-18 levels above the median (>/=134.6 ng/l), and was significantly enhanced in those urinary AERs at the upper end of the normal range (7.5 mug/min

Glucocorticoid-induced diabetes mellitus: prevalence and risk factors in primary renal diseases.

BACKGROUND/AIMS: In patients with primary renal diseases the current knowledge of hyperglycemia associated with corticosteroid therapy is limited. We therefore examined the prevalence and risk factors of glucocorticoid-induced diabetes mellitus (DM) in primary renal diseases. METHODS: Patients were recruited with primary renal diseases who were started on corticosteroids between April 2002 and June 2005. In patients with DM, an impaired fasting glucose level and/or positive urinary glucose analyses before corticosteroids therapy were excluded. RESULTS: During corticosteroid therapy (initial dose: prednisolone 0.75 +/- 0.10 mg/kg/day), DM was newly diagnosed in 17 (40.5%) of 42 patients. All of the 17 patients were diagnosed as having DM by postprandial hyperglycemia at 2 h after lunch, although they had normal fasting blood glucose levels. Age (OR 1.40, 95% CI 1.06-1.84) and body mass index (OR 1.87, 95% CI 1.03-3.38) were determined as independent risk factors for glucocorticoid-induced DM. CONCLUSION: Over 40% of patients with primary renal disease developed DM during treatment with corticosteroids. A high age and high body mass index are the independent risk factors for glucocorticoid-induced DM. 24-hour urinary glucose analyses and postprandial plasma glucose are useful for detecting glucocorticoid-induced DM.

Use of hydrogen peroxide in combination with nisin, sodium lactate and citric acid for reducing transfer of bacterial pathogens from whole melon surfaces to fresh-cut pieces.

Hydrogen peroxide (2.5%) alone or hydrogen peroxide (1%) in combination with nisin (25 microg/ml), sodium lactate (1%), and citric acid (0.5%) (HPLNC) were investigated as potential sanitizers for reducing Escherichia coli O157:H7 or Listeria monocytogenes populations on whole cantaloupe and honeydew melons. Whole cantaloupes inoculated with E. coli O157:H7 and L. monocytogenes at 5.27 and 4.07 log10 CFU/cm2, respectively, and whole honeydew melons inoculated with E. coli O157:H7 and L. monocytogenes at 3.45 and 3.05 log10 CFU/cm2, respectively, were stored at 5 degrees C for 7 days. Antimicrobial washing treatments were applied to inoculated whole melons on days 0 or 7 of storage and surviving bacterial populations and the numbers transferred to fresh-cut pieces were determined. At days 0 and 7 treatment with HPLNC significantly (p<0.05) reduced the numbers of both pathogens, by 3 to 4 log CFU/cm2 on both types of whole melon. Treatment with HPLNC was significantly (p<0.05) more effective than treatment with 2.5% hydrogen peroxide. While fresh-cut pieces prepared from stored whole melons were negative for the pathogens by both direct plating and by enrichment, fresh-cut pieces from cantaloupe melons treated with 2.5% hydrogen peroxide were positive for both pathogens and pieces from honeydew melons were positive for E. coli 0157:H7. The native microflora on fresh-cut melons were also substantially reduced by HPLNC treatment of whole melons. The results suggest that HPLNC could be used to decontaminate whole melon surfaces and so improve the microbial safety and quality of fresh-cut melons.

Combined efficacy of nisin and pediocin with sodium lactate, citric acid, phytic acid, and potassium sorbate and EDTA in reducing the Listeria monocytogenes population of inoculated fresh-cut produce.

The inability of chlorine to completely inactivate human bacterial pathogens on whole and fresh-cut produce suggests a need for other antimicrobial washing treatments. Nisin (50 microg/ml) and pediocin (100 AU/ml) individually or in combination with sodium lactate (2%), potassium sorbate (0.02%), phytic acid (0.02%), and citric acid (10 mM) were tested as possible sanitizer treatments for reducing the population of Listeria monocytogenes on cabbage, broccoli, and mung bean sprouts. Cabbage, broccoli, and mung bean sprouts were inoculated with a five-strain cocktail of L. monocytogenes at 4.61, 4.34, and 4.67 log CFU/g, respectively. Inoculated produce was left at room temperature (25 degrees C) for up to 4 h before antimicrobial treatment. Washing treatments were applied to inoculated produce for 1 min, and surviving bacterial populations were determined. When tested alone, all compounds resulted in 2.20- to 4.35-log reductions of L. monocytogenes on mung bean, cabbage, and broccoli, respectively. The combination treatments nisin-phytic acid and nisin-pediocin-phytic acid caused significant (P < 0.05) reductions of L. monocytogenes on cabbage and broccoli but not on mung bean sprouts. Pediocin treatment alone or in combination with any of the organic acid tested was more effective in reducing L. monocytogenes populations than the nisin treatment alone. Although none of the combination treatments completely eliminated the pathogen on the produce, the results suggest that some of the treatments evaluated in this study can be used to improve the microbial safety of fresh-cut cabbage, broccoli, and mung bean sprouts.

Effectiveness of irradiation treatments in inactivating Listeria monocytogenes on fresh vegetables at refrigeration temperature.

Ionizing radiation can be effective in controlling the growth of food spoilage and foodborne pathogenic bacteria. This study reports on an investigation of the effectiveness of irradiation treatment to eliminate Listeria monocytogenes on laboratory-inoculated broccoli, cabbage, tomatoes, and mung bean sprouts. Irradiation of broccoli and mung bean sprouts at 1.0 kGy resulted in reductions of approximately 4.88 and 4.57 log CFU/g, respectively, of a five-strain cocktail of L. monocytogenes. Reductions of approximately 5.25 and 4.14 log CFU/g were found with cabbage and tomato, respectively, at a similar dose. The appearance, color, texture, taste, and overall acceptability did not undergo significant changes after 7 days of postirradiation storage at 4 degrees C, in comparison with control samples. Therefore, low-dose ionizing radiation treatment could be an effective method for eliminating L. monocytogenes on fresh and fresh-cut produce.

Development of a novel microbial sensor with baker's yeast cells for monitoring temperature control during cold food chain.

A novel microbial sensor containing a commercial baker's yeast with a high freeze tolerance was developed for visibly detecting inappropriate temperature control of food. When the yeast cells fermented glucose, the resulting gas production triggered the microbial sensor. The biosensor was a simple, small bag containing a solution of yeast cells, yeast extract, glucose, and glycerol sealed up with multilayer transparent film with barriers against oxygen and humidity. Fine adjustment of gas productivity in the biosensor at low temperatures was achieved by changing either or both concentrations of glucose and yeast cells. Moreover, the amount of time that food was exposed to inappropriate temperatures could be deduced by the amount of gas produced in the biosensor. The biosensor was stable without any functional loss for up to 1 week in frozen storage. The biosensor could offer a useful tool for securing food safety by maintaining low-temperature control in every stage from farm to fork, including during transportation, in the store, and at home.

Survival of Escherichia coli O157:H7, Salmonella enteritidis, Staphylococcus aureus, and Listeria monocytogenes in Kimchi.

The survival of gram-positive and gram-negative foodborne pathogens in both commercial and laboratory-prepared kimchi (a traditional fermented food widely consumed in Japan) was investigated. It was found that Escherichia coli O157:H7, Salmonella Enteritidis, Staphylococcus aureus, and Listeria monocytogenes could survive in both commercial and laboratory-prepared kimchi inoculated with these pathogens and incubated at 10 degrees C for 7 days. However, when incubation was prolonged, the S. aureus level decreased rapidly from the initial inoculum level to the minimum detectable level within 12 days, whereas Salmonella Enteritidis and L. monocytogenes took 16 days to reach similar levels in commercial kimchi. On the other hand, E. coli O157:H7 remained at high levels throughout the incubation period. For laboratory-prepared kimchi, the S. aureus level decreased rapidly from the initial inoculum level to the minimum detectable level within 12 days, and L. monocytogenes took 20 days to reach a similar level. E. coli O157:H7 and Salmonella Enteritidis remained at high levels throughout the incubation period. The results of this study suggest that the contamination of kimchi with E. coli O157:H7, Salmonella Enteritidis, S. aureus, or L. monocytogenes at any stage of production or marketing could pose a potential risk.

Menadione-catalyzed luminol chemiluminescence assay for the rapid detection of viable bacteria in foods under aerobic conditions.

A menadione-catalyzed luminol chemiluminescence assay was developed for the rapid detection and estimation of viable bacteria in foods. The principle of this assay is based on the extracellular menadione-catalyzed active oxygen spieces (O2- and H2O2) generated by the activity of NAD(P)H:menadione oxidoreductase in viable cells. This luminol chemiluminescence assay requires 10 min for the incubation of cells with menadione and then 2 s for the measurement of chemiluminescence intensity after an injection of luminol solution without the treatment of cell lysis. This method was evaluated using liquid food samples of milk, vegetable juice, green tea, and coffee spiked with Escherichia coli ATCC 25922. The study result revealed that E. coli contamination at 1 to 10 CFU/ml in these foods could be detected after incubation at 37 degrees C for 7 h in an enrichment medium; however, the green tea and coffee samples requires filtration. This method could be a useful tool for the rapid evaluation of microbial food contamination.

Chemical and irradiation treatments for killing Escherichia coli O157:H7 on alfalfa, radish, and mung bean seeds.

In this study, the effectiveness of dry-heat treatment in combination with chemical treatments (electrolyzed oxidizing [EO] water, califresh-S, 200 ppm of active chlorinated water) with and without sonication in eliminating Escherichia coli O157:H7 on laboratory-inoculated alfalfa, radish, and mung bean seeds was compared with that of dry-heat treatment in combination with irradiation treatment. The treatment of mung bean seeds with EO water in combination with sonication followed by a rinse with sterile distilled water resulted in reductions of approximately 4.0 log10 CFU of E. coli O157:H7 per g. whereas reductions of ca. 1.52 and 2.64 log10 CFU/g were obtained for radish and alfalfa seeds. The maximum reduction (3.70 log10 CFU/g) for mung bean seeds was achieved by treatment with califresh-S and chlorinated water (200 ppm) in combination with sonication and a rinse. The combination of dry heat, hot EO water treatment, and sonication was able to eliminate pathogen populations on mung bean seeds but was unable to eliminate the pathogen on radish and alfalfa seeds. Other chemical treatments used were effective in greatly reducing pathogen populations on radish and alfalfa seeds without compromising the quality of the sprouts, but these treatments did not result in the elimination of pathogens from radish and alfalfa seeds. Moreover, a combination of dry-heat and irradiation treatments was effective in eliminating E. coli O157:H7 on laboratory-inoculated alfalfa, radish, and mung bean seeds. An irradiation dose of 2.0 kGy in combination with dry heat eliminated E. coli O157:H7 completely from alfalfa and mung bean seeds, whereas a 2.5-kGy dose of irradiation was required to eliminate the pathogen completely from radish seeds. Dry heat in combination with irradiation doses of up to 2.0 kGy did not unacceptably decrease the germination percentage for alfalfa seeds or the length of alfalfa sprouts but did decrease the lengths of radish and mung bean sprouts.

Effectiveness of electrolyzed acidic water in killing Escherichia coli O157:H7, Salmonella enteritidis, and Listeria monocytogenes on the surfaces of tomatoes.

A study was conducted to evaluate the efficacy of electrolyzed acidic water, 200-ppm chlorine water, and sterile distilled water in killing Escherichia coli O157:H7, Salmonella, and Listeria monocytogenes on the surfaces of spot-inoculated tomatoes. Inoculated tomatoes were sprayed with electrolyzed acidic water, 200-ppm chlorine water, and sterile distilled water (control) and rubbed by hand for 40 s. Populations of E. coli O157:H7, Salmonella, and L. monocytogenes in the rinse water and in the peptone wash solution were determined. Treatment with 200-ppm chlorine water and electrolyzed acidic water resulted in 4.87- and 7.85-log10 reductions, respectively, in Escherichia coli O157:H7 counts and 4.69- and 7.46-log10 reductions, respectively, in Salmonella counts. Treatment with 200-ppm chlorine water and electrolyzed acidic water reduced the number of L. monocytogenes by 4.76 and 7.54 log10 CFU per tomato, respectively. This study's findings suggest that electrolyzed acidic water could be useful in controlling pathogenic microorganisms on fresh produce.

Calcinated calcium killing of Escherichia coli O157:H7, Salmonella, and Listeria monocytogenes on the surface of tomatoes.

This study was conducted to evaluate the efficacy of calcinated calcium, 200 ppm chlorine water (1% active chlorine), and sterile distilled water in killing Escherichia coli O157:H7, Salmonella, and Listeria monocytogenes on the surfaces of spot-inoculated tomatoes. Inoculated tomatoes were sprayed with calcinated calcium, chlorinated water, or sterile distilled water (control) and hand rubbed for 30 s. Populations of E coli O157:H7, Salmonella, and L. monocytogenes in the rinse water and in the residual (0.1% peptone) wash solution were determined. Treatment with 200 ppm chlorine and calcinated calcium resulted in 3.40- and 7.85-log10 reductions of E. coli O157:H7, respectively, and 2.07- and 7.36-log10 reductions of Salmonella, respectively. Treatment with 200 ppm chlorine and calcinated calcium reduced L monocytogenes numbers by 2.27 and 7.59 log10 CFU per tomato, respectively. The findings of this study suggest that calcinated calcium could be useful in controlling pathogenic microorganisms in fresh produce.

[A detection method for recombinant DNA from genetically modified maize CBH351].

A method using polymerase chain reaction (PCR) was designed for the detection of genetically modified maize CBH351, which has not authorized as safe for use in foods and feeds in Japan yet. We analyzed a recombinant DNA (r-DNA) sequence introduced into CBH351 maize and designed specific primer pairs to amplify a segment including part of the r-DNA. The PCR products obtained by using the designed primer pairs are specific for CBH351 and should prevent false positive results caused by other maizes and other main cereal crops. The r-DNA introduced into CBH351 could be detected from maize samples containing 0.05-0.1% CBH351 maize. This sensitivity is theoretically equivalent to a level of several genome copies and so this technique is a very efficient means to detect CBH351 maize.

Dinucleotide repeat polymorphism of matrix metalloproteinase-9 gene is associated with diabetic nephropathy.

BACKGROUND: Although genetic susceptibility has been proposed as an important factor for the development and progression of diabetic nephropathy, the definitive gene has not been identified. To identify the genetic marker for diabetic nephropathy, we examined the association between the (A-C)n dinucleotide repeat polymorphism upstream of the matrix metalloproteinase-9 (MMP-9) gene and diabetic nephropathy in a group of Japanese patients with type 2 diabetes. METHODS: Patients were divided into three groups based on their urinary albumin excretion rate (AER) and the stage of diabetic retinopathy as follows: uncomplicated group (U), normal albuminuria (AER <20 microg/min) without proliferative retinopathy and with the duration of diabetes more than 20 years (N = 32); microalbuminuria group (M), 20 < or = AER < 200 microg/min (N = 155); overt nephropathy group (O), AER > or = 200 microg/min (N = 63). The region containing the dinucleotide repeat upstream of MMP-9 gene was amplified by polymerase chain reaction (PCR). The amplified products were analyzed with 7% formamide/urea acrylamide gel electrophoresis. The promoter constructs of the MMP-9 gene were transfected with the CMV-beta-galactosidase construct into 293 cells using the liposome method. Twenty-four hours after transfection, cells were harvested, and luciferase and beta-galactosidase activities were measured. RESULTS: Nine alleles of the dinucleotide repeat polymorphism (17 to 25 repeats) were identified, and the frequency of each allele in diabetic subjects was not different from that in nondiabetic controls. The frequency of the allele containing 21 repeats (A21) was most abundant (42.4% in control and 45.6% in diabetic subjects), followed by the allele with 23 repeats (A23; 35.4% in control and 27.6% in diabetic subjects). The A21 allele was less frequent in M and O than U (O, 38.9%; M, 45.5%; U, 59.3%, chi2 = 7.18; P < 0.05, O vs. U), while the frequency of the alleles other than A21 was not different among each group. The calculated odds ratio for nephropathy in the noncarrier, heterozygote, or homozygote of A21 allele was 3.38, 1.97, and 0.2, respectively. Furthermore, the promoter assay for the MMP-9 gene revealed that the A21 allele had a higher promoter activity compared with other alleles. No significant correlation was observed between serum MMP-9 concentrations and the MMP-9 gene polymorphism. CONCLUSION: These results indicate that the patients with A21 allele of the MMP-9 gene may be protected from the development and progression of diabetic nephropathy. Thus, the microsatellite polymorphism upstream of the MMP-9 gene could be a useful genetic marker for diabetic nephropathy.

Aurantiamide acetate, a selective cathepsin inhibitor, produced by Aspergillus penicilloides.

Aurantiamide acetate was isolated from the fermentation broth of Aspergillus penicilloides for the first time. Aurantiamide acetate inhibited cysteine proteinases, in particular, cathepsin L (3.4.22.15) and B (3.4.22.1) with IC50 of 12 microM and 49 microM, respectively. In the adjuvant-arthritic rat model, subcutaneously administered 10 mg/kg body weight of this compound suppressed hind paw swelling.

A multiplex PCR method of detecting recombinant DNAs from five lines of genetically modified maize.

Seven lines of genetically modified (GM) maize have been authorized in Japan as foods and feeds imported from the USA. We improved a multiplex PCR method described in the previous report in order to distinguish the five lines of GM maize. Genomic DNA was extracted from GM maize with a silica spin column kit, which could reduce experimental time and improve safety in the laboratory and potentially in the environment. We sequenced recombinant DNA (r-DNA) introduced into GM maize, and re-designed new primer pairs to increase the specificity of PCR to distinguish five lines of GM maize by multiplex PCR. A primer pair for the maize intrinsic zein gene (Ze1) was also designed to confirm the presence of amplifiable maize DNA. The lengths of PCR products using these six primer pairs were different. The Ze1 and the r-DNAs from the five lines of GM maize were qualitatively detected in one tube. The specific PCR bands were distinguishable from each other on the basis of the expected length. The r-DNA could be detected from maize samples containing 0.5% of each of the five lines of GM maize. The sensitivity would be acceptable to secure the verification of non-GMO materials and to monitor the reliability of the labeling system.

Synthesis and biological evaluation of cytogenin derivatives.

To enhance the stability in vivo, new derivatives of cytogenin were synthesized, and their biological activity and stability in mice were estimated. 2-(8-Hydroxy-6-methoxy-1-oxo-1H-2-benzopyran-3-yl)propionic acid (NM-3) was found to be the most stable among them. It modified collagen-induced arthritis in mice. It also showed potent anti-angiogenic activity in a mouse dorsal air sac assay.

Enhancement of glomerular heme oxygenase-1 expression in diabetic rats.

An increase in oxidative stress in diabetic subjects is implicated to play a pivotal role in diabetic vascular complications. In response to oxidative stress, antioxidant enzymes are considered to be induced and protect cellular functions to keep in vivo homeostasis. However, it remains to be clarified whether antioxidant enzymes are induced against oxidative stress especially in renal glomeruli at an early stage of diabetes. To answer this question, we examined the gene expression of a variety of antioxidant enzymes in glomeruli isolated from streptozotocin-induced diabetic rats. The mRNA expression of antioxidant enzymes such as catalase, glutathione peroxidase, and CuZn-superoxide dismutase, was unaltered in glomeruli of diabetic rats and was comparable to control rats. In contrast, the mRNA expression of heme oxygenase-1 (HO-1) was enhanced in glomeruli of diabetic rats as compared with control rats. A treatment with insulin as well as with vitamin E (40 mg/kg body weight every other day, intra-peritoneal injection) normalized the mRNA expression of HO-1 in the glomeruli of diabetic rats. Immunohistochemical analysis revealed that the up-regulated expression of HO-1 protein was localized in glomerular cells of diabetic rats. In conclusion, these results provide the first evidence that among antioxidant enzymes HO-1 expression is preferentially increased in diabetic glomeruli.