Hyperinsulinemic yellow KK mice and norepinephrine turnover

To clarify whether hyperinsulinemia accelerates sympathetic nervous system (SNS) activity, norepinephrine (NE) turnover, a reliable indicator of SNS activity, was measured in the interscapular brown adipose tissue (IBAT) and heart of hyperinsulinemic yellow KK and normoinsulinemic C57BL control mice at 12 weeks of age. The yellow KK mice were more obese and had higher levels of plasma glucose (about 2.3 times) and of plasma insulin (about 24 times) than did the control mice. In IBAT, the rate of NE turnover following blockade of NE synthesis with alpha-methyl-p-tyrosine (alpha-MPT) was significantly slower in yellow KK mice than in C57BL mice, although in heart, no significant difference between both groups was observed in NE turnover. These results suggest that hyperinsulinemia dose not always increase NE turnover, and furthermore that the reduced NE turnover in IBAT of yellow KK mice may be one of the important factors in the development of obesity of this animal, as it is recognized that brown adipose tissue is a main effector of diet-induced thermogenesis and its defect or absence would predispose to obesity.     

Diabetic nephropathy in KK and KK-Ay mice.

KK mice and KK-Ay mice were examined for age related changes in blood and urinary biophysiological parameters. Blood hemoglobin A1c levels were significantly higher in KK-Ay and KK mice as compared to non-diabetic ddY mice. In both diabetic mice, especially KK-Ay mice, plasma insulin levels markedly increased at 2 to 4 months of age, and the urinary glucose and microalbumin levels and albumin-to-creatinine ratios increased dependent on age. Plasma thrombomodulin levels significantly increased at 2 to 4 months of age in both KK and KK-Ay mice. Mild enlargement of mesangial matrix and segmental proliferative glomerular nephritis were revealed in KK and KK-Ay mice, respectively, at 4 months of age. KK-Ay mice with insulin resistance and high urine mAlb level might be useful as models for the early stage of diabetic nephropathy.     

STED nanoscopy of KK114‐stained pathogenic bacteria

Super‐resolution microscopy techniques can provide answers to still pending questions on prokaryotic organisms but are yet to be used at their full potential for this purpose. To address this, we evaluate the ability of the rhodamine‐like KK114 dye to label various types of bacteria, to enable imaging of fine structural details with stimulated emission depletion microscopy (STED). We assessed fluorescent labeling with KK114 for eleven Gram‐positive and Gram‐negative bacterial species and observed that this contrast agent binds to their cell membranes. Significant differences in the labeling outputs were noticed across the tested bacterial species, but importantly, KK114‐staining allowed the observation of subtle nanometric cell details in some cases. For example, a helix pattern resembling a cytoskeleton arrangement was detected in Bacillus subtilis. Furthermore, we found that KK114 easily penetrates the membrane of bacterial microorganism that lost their viability, which can be useful to discriminate between living and dead cells.     

Biotin supplementation improves glucose and insulin tolerances in genetically diabetic KK mice

Because biotin treatment may lower blood glucose in insulin-dependent diabetes, we chose to study such an effect in non-insulin dependent diabetes. Twenty-six diabetic KK mice, moderately hyperglycemic and insulin resistant, were treated for 10 weeks: 9 animals with 2 mg of biotin/Kg, 8 with 4 mg of biotin/Kg, and 9 with saline (controls). Blood glucose levels, oral glucose tolerance, insulin response to oral glucose, and blood glucose decrease in response to insulin were quantitated. Compared to controls, biotin treatment lowered post-prandial glucose levels, and improved tolerance to glucose and insulin resistance. Serum immunoreactive insulin levels in biotin-treated mice were like the controls.     

影响Ⅱ型糖尿病动物模型KK／Upj—A^y／J小鼠血糖的因素

目的探讨营养,年龄及性别等因子对Ⅱ型糖尿病动物模型-KK／Upj-A^y／J小鼠血糖的影响。方法尾部取血,用血糖仪测定实验组和对照组动物的血糖并进行实验分析。结果实验组血糖值明显高于对照组;实验组发病动物的比例也明显高于对照组动物;对照组8月龄小鼠血糖值低于糖尿病的发病标准;实验组年龄对动物的血糖影响较小;实验组和对照组性别基本不影响动物的血糖。结论（1）在同样携带A^y黄色肥胖糖尿病基因的遗传作用的条件下,饲料中的营养成分是影响小鼠血糖的主要因素,（2）动物的年龄是影响血糖的另一主要因素。（3）性别对动物血糖的影响不大。     

Chromium attenuates high-fat diet-induced nonalcoholic fatty liver disease in KK/HlJ mice

Nonalcoholic fatty liver disease (NAFLD) is a chronic liver disease associated with insulin resistance, oxidative stress, and inflammation. Evidence indicates that chromium has a role in the regulation of glucose and lipid metabolism and may improve insulin sensitivity. In this study, we report that chromium supplementation has a beneficial effect against NAFLD. We found that KK/HlJ mice developed obesity and progressed to NAFLD after feeding with high-fat diet for 8 weeks. High-fat-fed KK/HlJ mice showed hepatocyte injury and hepatic triglyceride accumulation, which was accompanied by insulin resistance, oxidative stress, and inflammation. Chromium supplementation prevented progression of NAFLD and the beneficial effects were accompanied by reduction of hepatic triglyceride accumulation, elevation of hepatic lipid catabolic enzyme, improvement of glucose and lipid metabolism, suppression of inflammation as well as resolution of oxidative stress, probably through enhancement of insulin signaling. Our findings suggest that chromium could serve as a hepatoprotective agent against NAFLD.     

Production of cellulases and hemicellulases by Aspergillus niger KK2 from lignocellulosic biomass

To investigate the production of cellulases and hemicellulases from Aspergillus niger KK2, solid state fermentation (SSF) was performed by using different ratios of rice straw and wheat bran. When A. niger KK2 was grown on rice straw alone as a solid support in SSF, the maximum FPase activity was 19.5 IU g(-1) in 4 days. Also, CMCase (129 IU g(-1)), beta-glucosidase (100 IU g(-1)), xylanase (5070 IU g(-1)) and beta-xylosidase (193 IU g(-1)) activities were concurrently obtained after 5-6 days of fermentation. The higher enzyme activities produced by A. niger KK2 is a significant advantage from the viewpoint of practical saccharification reaction. Cellulases and hemicellulases produced by A. niger KK2 might be applied to pulp and paper industry, feed industry and chemical industry.     

PAH utilization by Pseudomonas rhodesiae KK1 isolated from a former manufactured-gas plant site

Pseudomonas rhodesiae KK1 was isolated from a former manufactured-gas plant site, due to its ability to grow rapidly in a mixture of polycyclic aromatic hydrocarbons (PAHs). Radiorespirometric analysis revealed that strain KK1 was found to be able to mineralize anthracene, naphthalene and phenanthrene. Notably, phenanthrene-grown cells were able to mineralize anthracene much more rapidly than naphthalene-grown cells. Comparative analysis of amino acid sequences from 17 randomly selected dioxygenases capable of hydroxylating unactivated aromatic nuclei indicated that the enzymes for catabolism of PAHs, such as naphthalene and phenanthrene, might exist redundantly in strain KK1. Northern hybridization for cells grown on naphthalene or phenanthrene, using the putative naphthalene or phenanthrene dioxygenase gene fragment as a probe, suggested that the enzyme for naphthalene catabolism might share some homology in deduced amino acid sequences with phenanthrene dioxygenases. Also, it was found that three lipids (17:0 cyclo, 18:1 omega7c, 19:0 cyclo) increased in response to both naphthalene and phenanthrene, while the shift of other lipids varied from substrate to substrate.     

Pancreatic somatostatin contents in spontaneously diabetic KK and non-obese diabetic (NOD) mice

Alterations in the somatostatin (SRIF)-, insulin- and glucagon-containing cells were examined in two strains of spontaneously diabetic mice, KK and newly inbred non-obese diabetic (NOD) mice, using radioimmunoassay and immunohistochemical methods. The total pancreatic content and concentration of SRIF was decreased in male KK mice compared to their male controls aged 12-18 weeks. These results were consistent with the immunohistochemical findings. Pancreatic glucagon concentration and number of glucagon-containing cells were also decreased in KK mice, but pancreatic insulin concentrations were increased in KK mice. On the other hand, NOD mice aged 12-38 weeks within 15 days after onset of diabetes had increased concentrations of pancreatic SRIF. The pancreatic islets in NOD mice were decreased both in number and in size and were characterized by lymphocyte infiltration. SRIF-containing cells occupied the major part of the endocrine cells of the islets. Insulin-containing cells significantly decreased in number, but the number of glucagon-containing cells was fairly well preserved. These results and previous work concerning obob and dbdb mice indicate a parallel relationship between pancreatic SRIF and glucagon. The pancreatic glucagon thus as well as the pancreatic insulin may be an important determinant of pancreatic SRIF concentration in these diabetic animals.     

Gender-influenced obesity QTLs identified in a cross involving the KK type II diabetes-prone mouse strain

The inheritance of adiposity and related traits has been investigated in the obese, diabetes-prone KK/HlLt (KK) and the lean, normoglycemic C57BL/6J (B6) mouse strains, their F₁ hybrids, and a large intercross generation. Adiposity index (AI) was defined as the sum of four fat depot weights divided by body weight. Both male and female KK mice were obese, but AI values averaged twofold higher in females than in males. In contrast, B6 females were slightly more lean than males. A genome-wide search revealed several qualitative trait loci (QTLs) affecting AI. The proximal region of Chromosome (Chr) 9 has a large effect on AI, with a much stronger effect in females (lod = 6.3) than in males (lod = 2.7). The data for females fit a model in which a dominant allele from KK increases AI by 30%, with the lod score peak falling between markers D9Mit66 and D9Mit328. This QTL has large effects on inguinal and mesenteric fat pad weights, with smaller effects on gonadal and retroperitoneal fat pads. The region of Chr 9 containing this QTL has extensive homology to human Chr 11q. An X-linked QTL affecting AI was evident in males (lod = 3.77), but not females (lod = 0.7). Exclusion of mesenteric fat from male AI resulted in an increased lod score (lod = 5.0) at 8 cM distal to DXMit166. A suggestive AI QTL (lod = 4.2), differentially affecting males, was localized to Chr 18 near the glucocorticoid receptor locus. A region of Chr 7 had a strong effect on body weight (lod = 6.9), a significant effect on inguinal fat% (lod = 4.4), and a suggestive effect on AI in females (lod = 4.1). Plasma leptin levels were associated with genotypes on Chr 9 (lod = 5.9) and Chr 7 (lod = 4.2). A region of Chr 1 had a suggestive effect on fasted blood glucose (lod = 3.6). Received: 23 March 1999 / Accepted: 2 June 1999     

Changes in relationship between blood glucose level and plasma 1,5-anhydroglucitol level in KK mice

We compared the relationship of the blood glucose level to the plasma 1,5-anhydroglucitol (1,5AG) level between KK mice with abnormal glucose metabolism and ICR mice as controls. Although the plasma 1,5AG level did not show any significant correlation with the blood glucose level in the controls, it tended to logarithmically decrease with the rise in the blood glucose level in KK mice. Thus it is possible that the plasma 1,5AG level is specifically related to the abnormal glucose metabolism in this model of diabetes mellitus and that its routine examination in diabetic patients may help delineate the metabolic derangement in the disease.     

Effect of chronic metformin treatment of hepatic and muscle glycogen metabolism in KK mice.

Noninsulin-dependent diabetic KK mice, aged 90-100 days, with hyperinsulinemia and insulin resistance were treated with either metformin (N = 13) or water (control, N = 10) orally at a concentration of 50 mg/kg twice daily for 28 weeks. Age-matched nondiabetic Swiss Webster (SW) mice were also similarly treated. Liver and skeletal muscle glycogen synthase and phosphorylase enzymes were determined in all groups of mice. Both enzymes were significantly lower in control KK than in control SW mice. Metformin did not influence either of these enzymes in nondiabetic SW mice. However, it significantly increased the active form of glycogen synthase (a form) in both the liver and muscle of KK mice. Metformin also increased the active form of phosphorylase (a form) in the liver but not in the muscle of these mice. Hepatic glycogen content was similar in both control and metformin-treated KK mice. However, the muscle glycogen content was significantly higher in metformin-treated than in control KK mice. These data suggest that metformin preferentially stimulates glycogen synthesis in skeletal muscle, and this seems to be responsible for the observed improvement in fasting glucose and glucose response to an oral glucose load in KK mice.     

Genetics of obesity in KK mouse and effects of A y allele on quantitative regulation

KK mouse is known as a polygenic model for non-insulin-dependent diabetes mellitus with moderate obesity. To identify the quantitative trait loci (QTLs) responsible for the body weight in KK, linkage analysis with 97 microsatellite markers was carried out into 192 F₂ progeny, comprising 93 mice with a/a genotype at agouti locus and 99 mice with A ^y /a genotype, of a cross between C57BL/6J female and KK-A^y (A^y congenic) male, thereby the influence of A ^y allele on the quantitative regulation of body weight was also examined. In F₂ a/a mice, we identified a QTL on Chromosome (Chr) 4, and two loci with suggestive linkage on Chrs 15 and 18. In F₂ A ^y /a mice, a QTL was identified on Chr 6, and two loci with suggestive linkage were identified on Chrs 4 and 16. That the QTL on Chr 4 was held in common between F₂ a/a and F₂ A ^y /a progenies implies that this locus may be a primary component regulating body weight in KK and KK-A^y. These results suggest that the body weight in KK is controlled by multiple genes, and the different combination of loci is involved in the presence of A ^y allele. The QTL on Chr 6 seemed to determine the body weight by controlling fat deposition, because the linkage was identified on body weight and adiposity, and is suggested to be a component involved in the metabolic pathway in obesity caused by the A ^y allele. Received: 16 December 1997 / Accepted: 16 March 1998     

Quantitative trait loci that regulate plasma lipid concentration in hereditary obese KK and KK-Ay mice

To identify quantitative trait loci (QTLs) responsible for regulating plasma lipid concentration associated with obesity, linkage analysis was carried out on the 190 F2 progeny of a cross between C57BL/6J female and KK-Ay (Ay allele at the agouti locus congenic) male. In F2 a/a (agouti locus genotype) mice, two QTLs were identified on chromosome 1 and a QTL on chromosome 3 for total-cholesterol. A QTL for HDL-cholesterol was identified on chromosome 1 and a QTL for NEFA on chromosome 9. In F2 Ay/a mice, two QTLs for HDL-cholesterol were found on chromosome 1. Loci for other lipids with suggestive linkage were also identified. In both F2 mice, one QTL on chromosome 1 for total- and HDL-cholesterol was mapped near D1Mit150, in the vicinity of the apolipoprotein A-II (Apoa2) locus. Seven nucleotide substitutions out of 309 nucleotide apolipoprotein A-II cDNA sequences were identified between KK and C57BL/6J. The Ay allele may be an indication of the plasma lipid levels, but its influence was less apparent than in the case of weight control. The loci for lipids were not on identical chromosomes with those previously identified for obesity, suggesting that hyperlipidemia in KK does not coincidentally occur with obesity.