Impact of single-port cholecystectomy on postoperative pain.

BACKGROUND: This study compared postoperative pain following four-port laparoscopic cholecystectomy (LC) and single-port cholecystectomy (SPC). METHOD: This prospective, quasi-randomized, single-centre trial focusing on postoperative pain included 49 patients undergoing elective surgery with either a conventional LC, or SPC using a surgical glove port. Postoperative pain was evaluated using a visual analogue scale (VAS) and postoperative analgesic use as primary outcome measures. Total duration of operation, length of hospital stay, blood test results on the day after surgery and total port cost were secondary outcome measures. RESULTS: Twenty-five LCs and 24 SPCs were undertaken. The VAS score on day 1 after surgery was significantly less in the SPC group than in the LC group: median (range) 24 (12-38) versus 45 (33-57) mm (P = 0·002). Significantly fewer patients in the SPC group required analgesia (9 of 24 versus 19 of 25 in the LC group; P = 0·007). There were no significant differences in total duration of operation, length of hospital stay, and blood test results on the day after surgery. CONCLUSION: Single-port surgery using a surgical glove port reduces postoperative pain compared with conventional LC.

A non-MHC locus essential for autoimmune type I diabetes in the Komeda Diabetes-Prone rat.

The Long-Evans Tokushima Lean (LETL) rat, characterized by rapid onset of insulin-dependent (type I) diabetes mellitus (IDDM), no sex difference in the incidence of IDDM, autoimmune destruction of pancreatic beta cells, and no significant T cell lymphopenia, is a desirable animal model for human IDDM. We have established a diabetes-prone substrain of the LETL rat, named Komeda Diabetes-Prone (KDP) rat, showing a 100% development of moderate to severe insulitis within 220 d of age. The cumulative frequency of IDDM was 70% at 120 d of age, and reached 82% within 220 d of age. Here, we performed the first genome-wide scan for non-MHC IDDM susceptibility genes in this strain. The analysis of three crosses has led to the revelation of a major IDDM susceptibility gene, termed Iddm/kdp1, on rat chromosome (Chr) 11. Homozygosity for the KDP allele at this locus is shown to be essential for the development of moderate to severe insulitis and the onset of IDDM. Comparative mapping suggests that the homologues of Iddm/ kdp1 are located on human Chr 3 and mouse Chr 16 and would therefore be different from previously reported IDDM susceptibility genes.

Development of non-insulin-dependent diabetes mellitus in the double knockout mice with disruption of insulin receptor substrate-1 and beta cell glucokinase genes. Genetic reconstitution of diabetes as a polygenic disease.

Non-insulin-dependent diabetes mellitus (NIDDM) is considered a polygenic disorder in which insulin resistance and insulin secretory defect are the major etiologic factors. Homozygous mice with insulin receptor substrate-1 (IRS-1) gene knockout showed normal glucose tolerance associated with insulin resistance and compensatory hyperinsulinemia. Heterozygous mice with beta cell glucokinase (GK) gene knockout showed impaired glucose tolerance due to decreased insulin secretion to glucose. To elucidate the interplay between insulin resistance and insulin secretory defect for the development of NIDDM, we generated double knockout mice with disruption of IRS-1 and beta cell GK genes by crossing the mice with each of the single gene knockout. The double knockout mice developed overt diabetes. Blood glucose levels 120 min after intraperitoneal glucose load (1.5 mg/g body wt) were 108 +/- 24 (wild type), 95 +/- 26 (IRS-1 knockout), 159 +/- 68 (GK knockout), and 210 +/- 38 (double knockout) mg/dl (mean +/- SD) (double versus wild type, IRS-1, or GK; P < 0.01). The double knockout mice showed fasting hyperinsulinemia and selective hyperplasia of the beta cells as the IRS-1 knockout mice (fasting insulin levels: 0.38 +/- 0.30 [double knockout], 0.35 +/- 0.27 [IRS-1 knockout] versus 0.25 +/- 0.12 [wild type] ng/ml) (proportion of areas of insulin-positive cells to the pancreas: 1.18 +/- 0.68%; P < 0.01 [double knockout], 1.20 +/- 0.93%; P < 0.05 [IRS-1 knockout] versus 0.54 +/- 0.26% [wild type]), but impaired insulin secretion to glucose (the ratio of increment of insulin to that of glucose during the first 30 min after load: 31 [double knockout] versus 163 [wild type] or 183 [IRS-1 knockout] ng insulin/mg glucose x 10(3)). In conclusion, the genetic abnormalities, each of which is nondiabetogenic by itself, cause overt diabetes if they coexist. This report provides the first genetic reconstitution of NIDDM as a polygenic disorder in mice.

Inhibition of RXR and PPARgamma ameliorates diet-induced obesity and type 2 diabetes.

PPARgamma is a ligand-activated transcription factor and functions as a heterodimer with a retinoid X receptor (RXR). Supraphysiological activation of PPARgamma by thiazolidinediones can reduce insulin resistance and hyperglycemia in type 2 diabetes, but these drugs can also cause weight gain. Quite unexpectedly, a moderate reduction of PPARgamma activity observed in heterozygous PPARgamma-deficient mice or the Pro12Ala polymorphism in human PPARgamma, has been shown to prevent insulin resistance and obesity induced by a high-fat diet. In this study, we investigated whether functional antagonism toward PPARgamma/RXR could be used to treat obesity and type 2 diabetes. We show herein that an RXR antagonist and a PPARgamma antagonist decrease triglyceride (TG) content in white adipose tissue, skeletal muscle, and liver. These inhibitors potentiated leptin's effects and increased fatty acid combustion and energy dissipation, thereby ameliorating HF diet-induced obesity and insulin resistance. Paradoxically, treatment of heterozygous PPARgamma-deficient mice with an RXR antagonist or a PPARgamma antagonist depletes white adipose tissue and markedly decreases leptin levels and energy dissipation, which increases TG content in skeletal muscle and the liver, thereby leading to the re-emergence of insulin resistance. Our data suggested that appropriate functional antagonism of PPARgamma/RXR may be a logical approach to protection against obesity and related diseases such as type 2 diabetes.

Essential role of insulin receptor substrate 1 (IRS-1) and IRS-2 in adipocyte differentiation.

To investigate the role of insulin receptor substrate 1 (IRS-1) and IRS-2, the two ubiquitously expressed IRS proteins, in adipocyte differentiation, we established embryonic fibroblast cells with four different genotypes, i.e., wild-type, IRS-1 deficient (IRS-1(-/-)), IRS-2 deficient (IRS-2(-/-)), and IRS-1 IRS-2 double deficient (IRS-1(-/-) IRS-2(-/-)), from mouse embryos of the corresponding genotypes. The abilities of IRS-1(-/-) cells and IRS-2(-/-) cells to differentiate into adipocytes are approximately 60 and 15%, respectively, lower than that of wild-type cells, at day 8 after induction and, surprisingly, IRS-1(-/-) IRS-2(-/-) cells have no ability to differentiate into adipocytes. The expression of CCAAT/enhancer binding protein alpha (C/EBPalpha) and peroxisome proliferator-activated receptor gamma (PPARgamma) is severely decreased in IRS-1(-/-) IRS-2(-/-) cells at both the mRNA and the protein level, and the mRNAs of lipoprotein lipase and adipocyte fatty acid binding protein are severely decreased in IRS-1(-/-) IRS-2(-/-) cells. Phosphatidylinositol 3-kinase (PI 3-kinase) activity that increases during adipocyte differentiation is almost completely abolished in IRS-1(-/-) IRS-2(-/-) cells. Treatment of wild-type cells with a PI 3-kinase inhibitor, LY294002, markedly decreases the expression of C/EBPalpha and PPARgamma, a result which is associated with a complete block of adipocyte differentiation. Moreover, histologic analysis of IRS-1(-/-) IRS-2(-/-) double-knockout mice 8 h after birth reveals severe reduction in white adipose tissue mass. Our results suggest that IRS-1 and IRS-2 play a crucial role in the upregulation of the C/EBPalpha and PPARgamma expression and adipocyte differentiation.

Administration of optimum sustained-insulin release PLGA microcapsules to spontaneous diabetes-prone BB/Wor//Tky rats.

To show the possibility of sustained-release insulin formulation composed of PLGA, the optimum one was administered to BioBreeding rat, a model of spontaneous type I diabetes mellitus (IDDM). Every 2 weeks subcutaneous administration made their blood glucose level depend on the insulin release and food intake. However, all of them kept alive with little change or rather a little gain in body weight. Furthermore, some of pregnant rats with intermittent treatment bore fetuses, although additional insulin therapy seemed necessary. Therefore, the formulation could become a new tool as a provider of basal insulin for IDDM patients.

Disruption of insulin receptor substrate 2 causes type 2 diabetes because of liver insulin resistance and lack of compensatory beta-cell hyperplasia.

To investigate the role of insulin receptor substrate (IRS)-2 in vivo, we generated IRS-2-deficient mice by gene targeting. Although homozygous IRS-2-deficient mice (IRS-2-/- mice) had a body weight similar to wild-type mice, they progressively developed type 2 diabetes at 10 weeks. IRS-2-/- mice showed insulin resistance and a defect in the insulin-stimulated signaling pathway in liver but not in skeletal muscle. Despite insulin resistance, the amount of beta-cells was reduced to 83% of that in wild-type mice, which was in marked contrast to the 85% increase in the amount of beta-cells in IRS-1-deficient mice (IRS-1-/- mice) to compensate for insulin resistance. Thus, IRS-2 plays a crucial role in the regulation of beta-cell mass. On the other hand, insulin secretion by the same number of cells in response to glucose measured ex vivo was significantly increased in IRS-2-/- mice compared with wild-type mice but was decreased in IRS-1-/- mice. These results suggest that IRS-1 and IRS-2 may play different roles in the regulation of beta-cell mass and the function of individual beta-cells.

Proper regulation of cyclic AMP-dependent protein kinase is required for growth, conidiation, and appressorium function in the anthracnose fungus Colletotrichum lagenarium.

Colletotrichum lagenarium, the casual agent of anthracnose of cucumber, forms specialized infection structures, called appressoria, during infection. To evaluate the role of cAMP signaling in C. lagenarium, we isolated and functionally characterized the regulatory subunit gene of the cAMP-dependent protein kinase (PKA). The RPK1 gene encoding the PKA regulatory subunit was isolated from C. lagenarium by polymerase chain reaction-based screening. rpk1 mutants, generated by gene replacement, exhibited high PKA activity during vegetative growth, whereas the wild-type strain had basal level activity. The rpk1 mutants showed significant reduction in vegetative growth and conidiation. Furthermore, the rpk1 mutants were nonpathogenic on cucumber plants, whereas they formed lesions when inoculated through wounds. A suppressor mutant showing restored growth and conidiation was isolated from a rpk1 mutant culture. The rpkl-suppressor mutant did not show high PKA activity, unlike the parental rpk1 mutant, suggesting that high PKA activity inhibits normal growth and conidiation. The suppressor mutant, however, was nonpathogenic on cucumber and failed to form lesions, even when inoculated through wounds. The rpk1 and suppressor mutants formed melanized appressoria on the host leaf surface but were unable to generate penetration hyphae. These results suggest that proper regulation of the PKA activity by the RPK1-encoded regulatory subunit is required for growth, conidiation, and appressorium function in C. lagenarium.

Hyperreninemia due to increased renal renin synthesis in BioBreeding Worcester rats.

It is well known that diabetes mellitus is often associated with hypertension. We previously reported the unresponsiveness of renin release to volume depletion with impaired renal prostaglandin E2 synthesis in rats with streptozotocin-induced diabetes. However, we have found that BioBreeding Worcester rats, spontaneously susceptible to diabetes mellitus either before or after the onset of diabetes, showed a pronounced fourfold to ninefold increase in plasma renin activity in comparison with control Wistar rats. Furthermore, these rats developed mild hypertension as high as 134 mm Hg after the age of 90 days. The hyperreninemia responded to 1-week sodium loading or restriction; the blood pressure increased during sodium loading. Oral administration of captopril (30 mg/kg) for 1 week resulted in a large blood pressure decrease (-47.1 +/- 5.9 mm Hg, n = 10) in comparison with controls (-17.0 +/- 4.7 mm Hg, n = 12). Vascular response to angiotensin II was also attenuated. Plasma angiotensin II levels were 5.7-fold higher and associated with a 1.5-fold increase of plasma aldosterone concentration compared with control rats, whereas angiotensinogen-plasma concentrations were lower than in control rats. The renal renin content determined enzymatically or histochemically was more enhanced in BioBreeding Worcester rats than in control rats, but the renal renin messenger RNA levels did not differ. These results suggest that the strain-specific hyperreninemia in BioBreeding Worcester rats might be due to posttranscriptional abnormalities of renal renin synthesis. Further work is needed to elucidate the specific mechanism or mechanisms responsible.

Induction of cell killing, mutation and umu gene expression by 6-mercaptopurine or 2-thiouracil with UVA irradiation.

When Escherichia coli cells were irradiated by UVA in the presence of 6-mercaptopurine (6-MP) or 2-thiouracil (S2Ura), two kinds of repair-deficient strains of recA- and uvrA- were killed more efficiently than the parental wild-type strain having normal repair capacities. In addition, these agents with UVA exposure greatly induced the incidence of mutations in the uvrA- strain as compared with the wild-type strain but not the recA- strain. Furthermore, the induction of expression of umuDC genes was investigated in two Salmonella typhimurium strains, TA1535 and TA1538, carrying a pSK1002 plasmid. In these systems, it is easy to measure beta-galactosidase activities for the induced activities of SOS responses. These agents with UVA exposure also induced expression of the umuDC genes. These results suggest that 6-MP and S2Ura with UVA induce DNA damage which is repairable by the excision repair mechanism.

Reciprocal changes in left ventricular collagen alpha 1 chain gene expression between types I and IV in spontaneously diabetic rats.

The characteristic features of diabetic cardiomyopathy have been reported to be increased collagen formation associated with impairment of ventricular performance, based on experimental models of diabetes. The present study was therefore designed to clarify collagen gene expression in hearts obtained from female spontaneously diabetic BioBreeding Worcester Tokyo (BB/W@Tky) rats. Cardiac hypertrophy was observed as early as 14 weeks in diabetic BB/W@Tky rats, i.e. 4 weeks after the onset of diabetes. Left ventricular gene expression of collagen alpha 1 (I) was decreased to 10.6% of the control level. In 24-week-old diabetic rats, which had more marked cardiac hypertrophy, the level of alpha 1 Type I collagen mRNA was further decreased to 5.7% of the control level, whereas collagen alpha 1 (IV) mRNA demonstrated a 3-fold increase. As a result, a ratio of collagen alpha 1 (IV) to actin mRNA was positively correlated with plasma glucose concentration. These results suggest that hyperglycemia may alter the gene expression of extracellular matrix proteins, resulting in the morphological and functional changes seen in diabetic cardiomyopathy.

Alteration of insulin and glucagon secretion from the perfused BB rat pancreas before and after the onset of diabetes.

Secretion of insulin and glucagon from perfused BB rat pancreas was examined in rats (5 weeks old; n = 6) before and 2 weeks after (13 weeks old; n = 5) the onset of diabetes mellitus. The secretion of insulin and glucagon was modified by increasing concentrations of glucose in the perfusate from 2.8 mM to 16.5 mM. In 5-week-old BB rats, glucagon secretion was higher at the low glucose concentration (2.8 mM) and insulin secretion lower at the high glucose concentration (16.5 mM) than the corresponding values in age-matched control rats. In 13-week-old BB rats, insulin secretion was undetectable irrespective of glucose concentration. Glucagon secretion was only moderately suppressed by the perfusion of high glucose (16.5 mM) and was significantly higher than that of age-matched controls. The reduction in insulin secretion observed in 5-week-old rats without insulitis indicated that pancreatic B cells of BB rats of this age are somehow injured by humoral factors rather than cell infiltration. Cell function was apparently modified indirectly by the destruction of neighboring B cells.

Alterations of insulin and glucagon secretion from the perfused pancreas before, at the onset and after the development of diabetes in male Otsuka Long-Evans Tokushima Fatty (OLETF) rats.

The Otsuka Long-Evans Tokushima Fatty (OLETF) rat is an obese non-insulin dependent diabetes mellitus (NIDDM) model of an inbred strain. In this study, basal (2.8 mM glucose) insulin and glucagon and their responses to glucose (16.7 mM) were examined at the age of 9 weeks (n = 3) before the onset of diabetes, at 23 weeks (n = 6) at the onset of diabetes, and at 48 weeks (n = 5) after the development of diabetes by pancreatic perfusion. In Long-Evans Tokushima Otsuka (LETO, control) rats, insulin responses to glucose showed a biphasic pattern at all three ages, while in OLETF rats, basal insulin concentrations were significantly increased compared to those in controls at the age of 9 and 48 weeks. Insulin responses to glucose showed no difference from controls at 9 and 23 weeks, however, at 48 weeks the response was significantly decreased. In controls, high basal glucagon concentrations showed significant decrease in response to glucose at all ages. In OLETF rats, basal glucagon concentrations showed significant decrease compared to those in control rats at 23 and 48 weeks. Glucagon response to glucose significantly decreased at 9 and 23 weeks, but at 48 weeks there was no change in concentration in response to glucose. Pancreatic insulin content was lower at 48 weeks in OLETF rats than in LETO rats, although no differences were observed at other ages. There were no significant differences in pancreatic glucagon content between the two groups at any age. Morphologically, in OLETF rats the number of pancreatic B cells were decreased and A cells migrated into the center of islets at 48 weeks. The results suggested that one of the causes of diabetes in OLETF rats is impaired insulin response to glucose.

An improved genetic linkage map of rat chromosome 20.

BACKGROUND AND PURPOSE: Rat chromosome 20 is one of special interest because it contains some diabetogenic genes, such as a major histocompatibilitiy complex (MHC)-linked genetic components and quantitative trait loci. We studied rat chromosome 20, using the backcross progeny between BB/Wor and PVG.R23 rats, and confirmed the genetic linkage map by use of another backcross panel. METHODS: Backcross panels were done between BB/Wor and PVG.R23 rats, and BN and KZC rats. Length variations of simple sequence length polymorphism markers were analyzed by use of polymerase chain reaction (PCR) analysis. Alleles of RT1-Bb and RT1-Db were analyzed by use of the PCR-restriction fragment length polymorphism method. Genetic maps of rat chromosome 20 were constructed, using the Map Manager computer program. RESULTS: Fifty-two loci were mapped on rat chromosome 20. Genetic length was 57.9 cM, with average spanning of 1.11 cM between markers. The positions of RT1-N1, Tnf, and RT1-Bb into the MHC region were separated and confirmed by results of two backcross panels in our linkage studies. CONCLUSIONS: The genetic linkage map of rat chromosome 20 was improved, and was a useful tool for genetic analysis of a diabetogenic gene(s) and for producing MHC congenic strains.

Comparative study on isolation calls emitted from hamster pups.

Waveforms of isolation calls emitted from hamster pups, which were Syrian hamsters, Djungarian hamsters, and Chinese hamsters, were compared in a basic study on improving reproduction by decrease of cannibalism, because it was reported that maternal behavior was induced by isolation calls in rodents. Isolation calls of hamster pups, isolated from their mother and receiving cold stress, were collected by Real-Time Spectrogram (RTS), and calculated to spectrograms and power spectra by SIGNAL. Isolation calls consisted of ultrasonic vocalizations (USVs) and audible vocalizations (ADVs) in each species. Waveforms of isolation calls emitted by the hamster pups, were shown to have several characteristic features. In this study, the species specificity of isolation calls was shown in hamster pups. It would seem that the species specificity originates in the differences of sensitivity to cold stress via the autonomic nerve in hamsters.

Mandible analysis of NOD and NON strains of mice.

10 inbred strains of mice including NOD and NON were identified by discriminant and canonical discriminant analysis of the mandible measurements. The number of erroneous discriminations was 0.5% (1/199) for the females and 0% for males (0/232). In canonical discriminant analysis (discriminant analysis with reduction of dimensionality), NOD and NON inbred strains were separately located on 2-dimensional planes, Z1-Z2, Z1-Z3, and Z2-Z3. These results clearly indicate that the genetic constitution of NOD and NON differs, although they have been established from a common ancestor ICR mouse. Causes of divergence between NOD and NON mice are discussed.

An ultrastructural study on the pancreatic islet B cells in diabetic herbivorous voles.

An ultrastructural study was performed on the pancreatic islet cells of normal herbivorous voles and of voles in which diet-induced diabetes had been induced by feeding a low-fibre, high-concentrate diet. Examination of the pancreatic islet cells revealed degranulation and a well-developed Golgi apparatus and rough endoplasmic reticulum in the B cells of the slightly diabetic voles showing moderate hyperglycaemia and hyperinsulinaemia, and markedly degenerative B cells with almost complete absence of secretory granules in the severely diabetic voles showing marked hyperglycaemia and hypoinsulinaemia. These results indicate that the pancreatic B cells had become hyperfunctional so that insulin secretion was increased in the slightly diabetic voles; thereafter the B cells degenerated rapidly and the voles fell into insulin deficiency.

[Clinical experience with S-6436 in patients with urinary tract infections (author's transl)].

Forty eight patients with urinary tract infections including a case of acute prostatitis were treated with S-6436 and the following results were obtained: 1. Thirty eight patients with acute cystitis resulted in 19 excellent, 15 good and 4 poor with effectiveness rate of 89.5%. 2. Ten patients with complicated urinary tract infections resulted in 1 excellent, 5 good and 4 poor with effectiveness rate of 60%. 3. Infecting organisms from 38 patients with acute cystitis were occupied by 28 strains of E. coli. The sensitivity rate of the infecting organisms to cephalexin was 96.4%. 4. In a few cases (6.25%) side effects of S-6436 were observed. 5. S-6436 will be one of the good first choice antibiotics for acute cystitis.

Chymase inhibition attenuates monocrotaline-induced sinusoidal obstruction syndrome in hamsters.

Chymase stored in mast cells activates matrix metalloproteinase (MMP)-9, which may relate to the progression of sinusoidal obstruction syndrome (SOS). We investigated the preventive effect of a chymase inhibitor, TY-51469, on monocrotaline-induced SOS in hamsters. Hamsters were orally administrated with a single dose of monocrotaline (120 mg/kg) to induce SOS. Treatment with TY-51469 (1 mg/kg per day) or placebo had started 3 days before the monocrotaline administration. Two days after the monocrotaline administration, significant increases in aspartate aminotransferase, alanine aminotransferase and total bilirubin and a significant reduction of albumin were observed in plasma, but their changes were significantly attenuated by treatment with TY-51469. The numerous hepatic necrosis areas were observed in the placebo-treated group, but the ratio of necrotic area to total area in liver had been significantly reduced by treatment with TY-51469. Both chymase activity and MMP-9 level in liver were significantly augmented in the placebo-treated group. Furthermore, tumor necrosis factor (TNF)-α level in liver was also augmented in the placebo-treated group. However, the chymase activity and levels of MMP-9 and TNF-α were significantly attenuated in the TY-51469-treated group. Until 14 days after monocrotaline administration, survival rates in the placebo- and TY-51469-treated groups were 25% and 70%, respectively, and a significant difference was observed. In conclusion, chymase inhibition by TY-51469 may prevent the accelerating of severity in monocrotaline-induced SOS in hamsters.