STAT3 activation via interleukin 6 trans-signalling contributes to ileitis in SAMP1/Yit mice.

BACKGROUND AND AIMS: SAMP1/Yit mice spontaneously develops intestinal inflammation. Previously, we demonstrated that the signal transducer and activator of transcription (STAT)-3/suppressor of cytokine signalling (SOCS)-3 pathway is pivotal in human inflammatory bowel disease. In our studies in SAMP1/Yit mice, the aim was to investigate whether STAT3 activation contributes to ileitis and to examine the therapeutic effects of this signal blockade. METHODS: Intestinal expression of phospho-STAT3 in SAMP1/Yit mice and control AKR/J mice was examined by western blotting and immunohistochemistry. SOCS3 and interleukin 6 (IL-6) mRNA were determined by northern blotting and reverse transcription-polymerase chain reaction, respectively. We also examined the effects of intravenously injected hyper-IL-6, an IL-6/soluble IL-6 receptor fusion protein, and of soluble gp130-Fc, a specific inhibitor of soluble IL-6 receptor signalling, on STAT3 phosphorylation and disease severity in SAMP1/Yit mice. RESULTS: Phospho-STAT3 was expressed strongly during the disease course in SAMP1/Yit mice but only transiently in AKR/J mice. Phospho-STAT3 was localised to epithelial and mononuclear cells in the diseased intestine of SAMP1/Yit mice. SOCS3 as well as IL-6 mRNAs were expressed in affected intestine. Administration of hyper-IL-6 caused disease exacerbation and enhancement of STAT3 phosphorylation. In contrast, soluble gp130-Fc administration ameliorated the disease and suppressed STAT3 phosphorylation. CONCLUSION: STAT3 signalling is critical in the development of intestinal inflammation in SAMP1/Yit mice. Blockade of this signalling pathway by soluble gp130-Fc may have therapeutic effects in inflammatory bowel disease.

Establishment of a Tcrb and Trp53 genes deficient mouse strain as an animal model for spontaneous colorectal cancer.

A congenic C57BL/6JJcl-Tcrbtm1MomTrp53tm1 (Tcrb-/-:Trp53-/-) mouse lacking T-cell receptor beta chain (TCR beta) and transformation related protein 53 (p53) has been established at the N8th generation of backcrossing male Tcrb-/-:Trp53-/- mice, which had been obtained by mating a Tcrb-/- mouse with a Trp53-/- mouse, with female C57BL/6JJcl mice. In the mice deficient for the both genes, occurrence of tumor masses was observed mostly in the cecum with high frequency as examined at 3 months of age. The majority of the masses had histologic features of hyperplasia or dysplasia while occasional lesions were noted to be adenocarcinomas invading the submucosa (invasive adenocarcinoma). As examined at 4 months of age and thereafter, all mice had 4-5 colorectal tumors per animal, the lesions being located mainly in the cecum and, histopathologically, all the obvious neoplastic growths in the regions examined were invasive adenocarcinomas. The Tcrb and Trp53 genes deficient mouse strain which develops spontaneous colorectal carcinoma with fairly high frequency at early age would be useful as an animal model for colorectal cancer.

Intestinal microflora are necessary for development of spontaneous adenocarcinoma of the large intestine in T-cell receptor beta chain and p53 double-knockout mice.

This study was conducted to confirm the hypothesis that intestinal microflora are required for the development of adenocarcinoma in the colon of the TCRbeta and p53 double-knockout (TCRbeta-/- p53-/-) mouse. Germ-free TCRbeta-/- p53-/- mice were produced. At 7 weeks of age, the animals were divided into two groups (n = 10/group), and one of these groups was conventionalized. Animals of both groups were subjected to histopathological examination for adenocarcinoma of the colon at 4 months of age. There was no development of adenocarcinoma of the colon among the germ-free mice, whereas in the conventionalized group, adenocarcinomas of the ileocecum and cecum were detected in 70% of animals. These results indicate the usefulness of the TCRbeta-/- p53-/- mouse as a colon cancer animal model that develops spontaneous adenocarcinoma of the colon early in life, and suggest that intestinal microflora play a major role in the development of adenocarcinoma of the colon in this animal model.

Magnetic resonance imaging findings in primary amyloidosis-associated arthropathy.

The MRI findings of amyloid arthropathy associated with primary amyloidosis are presented here possibly for the first time in the literature. Two types of lesions are noted: (1) capsular and tendon lesions; these regions are thickened, hypointense and enhanced by gadolinium (Gd) on T1 weighted imaging (T1WI), and hyperintense on T2 weighted imaging (T2WI), and (2) periarticular and osseous lesions; these regions appear to be tumor-forming and hypointense on both T1WI and T2WI and are not enhanced by Gd. It is necessary to differentiate these findings from other diseases such as chondrosarcoma, rhabdomyosarcoma and chronic inflammatory lesions such as tuberculosis.

Takayasu's arteritis in a 69 year-old woman.

Takayasu's arteritis and temporal arteritis share many clinical and pathological features. The most discriminatory feature between the two diseases is the age at onset; the mean age at onset of the disease was reported as being 26 years for Takayasu's arteritis and 69 years for temporal arteritis. Here we report a 69-year-old woman who presented with a weak right radial artery pulse. The ethnic background and the presence of vascular insufficiency of the right upper extremity and the absence of clinical signs such as shoulder stiffness and tender scalp indicate that her diagnosis is Takayasu's arteritis. It must be emphasized that the two conditions could be differentiated based on the clinical findings even in a patient as old as 69 years old.

Collaborative work to evaluate toxicity on male reproductive organs by repeated dose studies in rats 22). Effects of 2- and 4-week administration of theobromine on the testis.

The effects of theobromine, a xanthine derivative, on the testis were compared between rats dosed for 2 and 4 weeks to determine whether a 2-week dosing period is long enough to detect toxicity. Theobromine was administered orally to male Sprague-Dawley rats at dose levels of 250 and 500 mg/kg for 2 weeks starting at the age of 6 or 8 weeks, and for 4 weeks from the age of 6 weeks. Histopathological examination of reproductive organs revealed toxic findings in the testis at 500 mg/kg after 2 weeks of dosing at both ages, and at 250 and 500 mg/kg after 4 weeks of dosing. The primary findings were degeneration/necrosis and desquamation of spermatids and spermatocytes, vacuolization of seminiferous tubules, and multinucleated giant cell formation. These findings were present mainly in stages I-VI and XII-XIV. From these results, it is concluded that the toxic effects of theobromine on the testis can be detected by repeated dosing for 2 weeks as well as for 4 weeks.

Bioluminescent monitoring of intracellular ATP during fermentation.

The luciferase gene was introduced as a probe into a cell in order to develop a bioluminescent monitoring of intracellular ATP during fermentation. Two plasmids were constructed with two types of promoters. One was pLac-Luc, which had the luciferase gene under the lac promoter to be expressed at a high level. The other was pTet-Luc, which had the luciferase gene under the tetracycline promoter to be stably expressed. A threonine-overproducing strain of Escherichia coli (No. 29-4) was transformed with each plasmid. The recombinant E. coli strains were characterized in their growth, threonine production and luciferase expression. The bioluminescence produced intracellularly from expressed luciferase was detected during fermentation -in a non-destructive manner. The bioluminescent intensity reflected both intracellular ATP and luciferase levels, and the results indicate that stable expression of a luciferase reporter is essential for monitoring intracellular ATP.

A case of autoimmune hemolytic anemia induced by IFN-beta therapy for type-C chronic hepatitis.

A 51-year old woman who had type-C chronic hepatitis developed autoimmune hemolytic anemia after 4 weeks treatment with a total 48 million interferon-beta, 6 million 8 times. Indirect Coomb's test turned from negative to positive through IFN therapy and a complication of asymptomatic primary biliary cirrhosis was confirmed by histologic findings of the biopsied liver which was performed during IFN therapy and a presence of anti-mitochondrial antibody. After stopping IFN therapy and adding prednisolone anemia improved in getting negative indirect Coombs test but remaining positive direct Coombs test.

Renin inhibitors. III. Synthesis and structure-activity relationships of transition-state inhibitors containing dihydroxyethylene isostere at the P1-P1 site.

The design, synthesis and structure-activity relationships of transition-state inhibitors containing the dihydroxyethylene isostere at the scissile site are described. The compounds with (2S,3R,4S)-4-amino-5-cyclohexyl-1-morpholino-2,3-pentanediol at the P1-P1 site are potent renin inhibitors. (2S,3R,4S)-4-[N-[(2S)-3-Ethylsulfonyl-2-(1-naphthylmethyl)propiony l]-L- norleucyl]amino-5-cyclohexyl-1-morpholino-2,3-pentanediol (2) (BW-175), which is the most potent inhibitor (IC50: 3.3 nM against human renin) in this series, poorly inhibits cathepsin D (IC50: 26000 nM) and pepsin (IC50: > 100000 nM), and thus it is specific for renin. Compound 2 contains only one amino acid and showed an oral bioavailability of 2.8% at 10 mg/kg and 9.7% at 30 mg/kg in rats. The interaction between renin and inhibitor 2 is discussed on the basis of molecular modeling studies.

[Early lesions of pancreatic islets in small-ob mice (C57BL/6J-ob/ob)].

Pancreatic islets of C57BL/6J-ob mice were microscopically examined at the ages of 5, 13, and 21 days, to clarify the time of the increase in number of A-cells, which appears to lead to diabetic symptoms in Small-ob mice (C57BL/6J-ob/ob). The findings of islet in 5-day-old C57BL/6J-ob mice were the same as those of C57BL/6J-+/+ mice. At 13 days of age, changes in islet of C57BL/6J-ob mice were classified into 3 groups. They were no significant changes (putative normal), hypertrophic B-cells with clear cytoplasm (Ob mice), and an increase of A-cells (Small-ob mice), respectively. Electron microscopically, a tendency toward dilatation of endoplasmic reticulum was found in the hypertrophic B-cells. In the islets that had an increase in number of A-cells, only a few vacuoles, which were suggestive of degranulation, were found in the cytoplasm of B-cells, and premature secretory granules were seen in the A-cells. In part of the A-cells, development of rough-surfaced endoplasmic reticulum was also seen. Mild hypertrophy of the islets and the B-cells were observed in more non-diabetic Ob mice at the age of 21 days than non-obese mice at the same age, and only 2 of 6 mice had the increase in number of A-cells. In Small-ob mice, the increase in number of A-cells was more obvious than in the diabetic Ob mice, while other islet findings were similar to those of non-obese mice in extent.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of side-chain structure on inhibition of yeast fatty-acid synthase by cerulenin analogues.

Yeast fatty-acid synthase (FAS) inhibition by cerulenin analogs with varying side-chain lengths was compared with that of cerulenin, tetrahydrocerulenin and iodoacetamide. Although inhibition by cerulenin was the highest, the analogs having (E,E)-delta 7,10 double bonds showed high inhibition. This strongly suggests that the (E,E)-delta 7,10 double bonds play an important role in the interaction of the inhibitors with the enzyme. It was suggested that the size of the hydrophobic cavity in the condensing enzyme terminates fatty-acid chain elongation by decreasing inhibition by the C18 analog. Like cerulenin itself, the shortest analog (C6) did not induce malonyl-CoA decarboxylase activity.

Renin inhibitors. II. Synthesis and structure-activity relationships of N-terminus modified inhibitors containing a homostatine analogue.

The synthesis and structure-activity relationships of N-terminus modified renin inhibitors containing the homostatin analogue, (2RS,4S,5S)-5-amino-2-ethyl-4-hydroxy-7-methyloctanoic acid, are described. The compounds having a 3-alkyl (or aryl)sulfonylpropionyl residue at the N-terminus are found to be potent inhibitors which contain two amino acids. (2RS,4S,5S)-N-Isobutyl-5-[N-[(2S)-3-ethylsulfonyl-2-(1- naphthylmethyl)propionyl]-L-norleucyl]-amino-2-ethyl-4-hydroxy-7- methyloctanamide (20) has an IC50 of 0.5 nM against human plasma renin and the oral bioavailability of 20 is 0.73% in rats. Interaction between renin and the N-terminus of 1 and 20 is discussed in molecular modeling studies.

Syntheses of cerulenin and its analogs. II. Synthesis and biological activity of dl-carbacerulenin, a carbocyclic analog of cerulenin.

2,3-Epoxy-4-hydroxy-4-((E,E)-3,6-octadienyl)cyclopentanone (dl-carbacerulenin 5) was synthesized via the epoxyketones 15a and 15b as a mimic of the active form of the antibiotics cerulenin 1, a potent inhibitor of fatty acid synthetase (FAS). The monobenzyl ethers (12 and 13), synthetic intermediates of 15, were prepared by direct benzylation of the epoxycyclopentene (7). Inhibitory activity of synthesized 5 toward yeast FAS was less than that of cerulenin by a factor of 1000.

[An autopsy case of a patient with myasthenia gravis who showed various symptoms of collagen diseases and complicated with malignant thymoma].

A 37-year-old man suffered from photosensitivity and urinary casts with serological findings of positive anti-DNA antibody, LE cells and false positive VD reaction in September of 1979. He developed general fatigue, dyspnea and diplopia with ptosis of bilateral eyelids in November of 1979, which were improved by the anti-cholinesterase drugs. In January of 1980, he had an attack of unconsciousness and his chest X-ray film showed several tumorous shadows in the anterior mediastinum and middle and lower lung fields. Treating him with chemotherapy of VEMP, the pulmonary shadows disappeared. However, he developed severe muscle weakness with an elevated CPK (430 mU/ml) and a myogenic EMG pattern along with an increased anti-acetylcholine receptor antibody (243 n Mol/l), dysphagia and eyelid-ptosis. He died in September of 1985 and his autopsy disclosed a malignant thymoma of mixed type in the anterior mediastinum and an atrophy and fibrosis with infiltration of inflammatory cells in the striated muscles.

Binding site of cerulenin in fatty acid synthetase.

An antibiotic cerulenin, (2R, 3S)-2,3-epoxy-4-oxo-7,10-trans,trans- dodecadienamide, irreversibly inhibits fatty acid synthetase from Saccharomyces cerevisiae. Three moles of cerulenin were bound to 1 mol of the enzyme with concomitant loss of its activity. Pretreatment of the enzyme with iodoacetamide reduced the amount of cerulenin bound to the enzyme. Since iodoacetamide is known to specifically bind to the cysteine residue on the condensing reaction domain, cerulenin is considered to bind to the same domain. Tryptic digestion of the [3H] cerulenin-treated enzyme gave a radioactive peptide; its amino acid composition was Asx 1, Thr 1, Ser 1, Glx 2, Pro 1, Gly 1, Ala 1, Val 1, Ile 1, and Leu 2. This composition included all the amino acids of the condensing reaction site (Thr-Pro-Val-Gly-Ala-Cys) previously reported by Kresze et al. (Eur. J. Biochem., 79, 181 [1977] except for Cys. When the enzyme was treated with [3H]cerulenin and digested successively with trypsin and carboxypeptidase P, a [3H] cerulenin-cysteine adduct was isolated as the sole product. This was identified with the adduct chemically synthesized from non-labeled cerulenin and cysteine, and its structure was elucidated by 1H-, 13C-NMR, and fast atom bombardment mass spectrometry. These results indicate that cerulenin, forming a hydroxylactam ring, reacts at its epoxide carbon (C-2 position) with the SH-group of the cysteine residue in the condensing reaction domain of yeast fatty acid synthetase.

A novel nonpeptidic, orally active renin inhibitor.

BW-175 is a newly synthesized renin inhibitor which is a nonpeptidic, norleucine analog. Its IC50 values for renin activity in human, squirrel monkey, marmoset, dog, hog, rabbit and rat plasma were 3.3, 6.6, 2.4, 42, 110, 86 and 3500 nM, respectively, and 26 microM for cathepsin D. Pepsin and angiotensin converting enzyme were hardly inhibited at 10(-4) M. BW-175 showed an oral bioavailability of 2.8% at 10 mg/kg and 9.7% at 30 mg/kg in rats. In normotensive, furosemide-treated high-renin marmosets, BW-175 (30 mg/kg p.o.) caused an intensive reduction in plasma renin activity and plasma angiotensin I formation, associated with a reduction in systolic blood pressure of 10-20 mm Hg for 2 hours.

Lactoquinomycin, a novel anticancer antibiotic. II. Physico-chemical properties and structure assignment.

Lactoquinomycin, a new antibiotic, C24H27NO8, mp 151 approximately 159 degrees C (dec), FAB-MS: m/z 458 (MH+), is a basic substance, showing UV lambda MeOHmax (epsilon) 215 (37,600), 254 (10,700) and 432 nm (4,760), and IR nu CHCl3max 1790 (gamma-lactone), 1665 and 1650 (quinone) cm-1. The structure of lactoquinomycin has been elucidated by 1H NMR and ORD in comparison with those of kalafungin.

[Clinico-pathological studies on small obese mice occurring in C57BL/6J-ob/ob mice].

Eight small obese mice, 10 to 79 weeks of age, which occurred in C57BL/6J obese mice, were examined clinico-pathologically. These mice were characterized by small-sized obesity, and showed hyperglycemia without hyperinsulinemia and glucosuria without ketonurina throughout their life span. Morphologically, neither hypertrophy nor hyperplasia were found in the islets of Langerhans. There were an increase of A cells and a decrease of B cells in their islets. In the kidney, thickening of mesangial matrix in glomeruli and deposition of glycogen in the nucleus of epithelial cells of the distal tubules were seen. These findings suggest that the small obese mice are an obese-diabetic animal of different type from the usual obese mice (C57BL/6J-ob/ob). The mechanism for development of this condition was unknown.