Skin inflammation exacerbates food allergy symptoms in epicutaneously sensitized mice.

BACKGROUND: Cutaneous exposure to food antigen through impaired skin barrier has been shown to induce epicutaneous sensitization, thereby causing IgE-mediated food allergies. OBJECTIVE: We examined whether skin barrier impairment following epicutaneous sensitization exacerbates food allergies. METHODS: BALB/c mice were epicutaneously sensitized by repeated application of ovalbumin (OVA) to MC903-pretreated ear skin for 48 hours weekly and then intragastrically challenged with OVA. After the first oral challenge, the skin barrier was disrupted with topical application of MC903 or by tape-stripping. Mice were monitored for changes in body temperature and the occurrence of diarrhea after undergoing the second oral challenge. Serum levels of mouse mast cell protease-1 (mmcp1) and OVA-specific IgE, IgG1, IgG2a antibodies and OVA-specific IgA levels in intestinal lavage fluid were measured by ELISA. Tissue accumulation of eosinophils was determined histologically. RESULTS: Epicutaneously sensitized mice developed anaphylaxis after intragastric challenge, as evidenced by diarrhea, decreased body temperature, and increased serum mmcp1 levels. Skin barrier disruption by MC903 treatment or tape-stripping exacerbated allergic reactions induced by oral challenge. MC903 treatment increased serum baseline and postchallenge mmcp1 levels. Topical pretreatment with dexamethasone alleviated allergic reactions that were exacerbated by MC903 treatment. CONCLUSION: Even after eliminating exposure to the antigen, inflammation from skin barrier disruption can exacerbate the severity of food allergy symptoms. Serum baseline mmcp1 levels might be an effective marker for predicting the severity of antigen-induced allergic symptoms.

Prognostic significance of cyclin E overexpression in resected non-small cell lung cancer.

Cyclin E plays a pivotal role in the regulation of G1-S transition and relates to malignant transformation of the cells. However, the clinical significance of cyclin E expression in patients with non-small cell lung cancer remains unknown. We examined the expression of cyclin E in 242 resected non-small cell lung cancer in pathological stages I-IIIa and analyzed its relation to clinicopathological factors. Cylin E overexpressions were observed frequently in deeply invasive tumors. Multivariate analysis revealed that complete resection, pathological stage, and cyclin E expression were independent prognostic indicators. When cyclin E and proliferating cell nuclear antigen are combined, the cases negative for both had a significantly better prognosis than the other cases. We concluded that cyclin E overexpression relates to deeply invasive tumors and is correlated with poor prognosis. New therapeutic options may be provided by combination of cyclin E expression and proliferating cell nuclear antigen overexpression.

Presence of ras oncogene mutations and human papillomavirus DNA in human prostate carcinomas.

The frequency of H-ras, K-ras, and N-ras mutations and the presence of high-risk human papillomavirus (HPV 16, 18, and 33) DNA were studied in 75 paraffin-embedded specimens obtained from 68 Japanese patients with a variety of prostate carcinomas by using polymerase chain reaction and DNA hybridization with sequence-specific oligonucleotides. Ten specimens each of normal and benign hyperplastic prostatic tissues from the same number of patients were also examined for this analysis. Of 68 carcinoma cases, ras gene mutations were present in 16 cases (24%) and HPV DNAs in 28 cases (41%). Eleven mutations were detected in codon 61 of H-ras, 4 in codon 12 of N-ras, and 2 in codon 61 of K-ras. HPV 16, 18, and 33 DNAs were found in 11, 17, and 5 cases, respectively. Eight of the 16 cases with ras mutation also harbored HPV DNAs. The frequency of ras mutations and the HPV infection increased in patients with advanced stages of the tumor and with the higher Gleason score. There was the predominant presence of H-ras codon 61.2 (CAG-->CTG) mutation and HPV 18 DNA in prostatic carcinomas metastasizing to the bone. None of the normal or benign hyperplastic prostatic specimens contained either ras mutation or HPV DNA. Our results suggest that ras gene mutations and HPV infections are relatively frequent, at least in prostate carcinoma of Japanese patients. These two factors appear to be related to the progression of the tumor. Moreover, H-ras codon 61.2 mutation and HPV 18 infection may have some predictive roles for bone metastasis in prostate carcinoma.

Expression of CD44 variant exon 6 in stage I non-small cell lung carcinoma as a prognostic factor.

Specific CD44 isoforms are the surface adhesion molecules that have been shown to be associated with metastasis. In the present study, the role of the expression of standard and variant CD44 isoform (CD44s and CD44v6) as prognostic indicators in pathological stage I non-small cell lung carcinoma was investigated immunohistologically using monoclonal antibodies. The results showed that the expression of CD44v6 correlates with adverse prognosis in stage I non-small cell lung carcinoma but not CD44s. The 5-year survival rate of the patients with CD44v6-positive tumors was 50%, which was significantly lower than that of CD44v6-negative patients (88%; P = 0.001). The incidence of recurrent distant metastasis in the CD44v6-positive patients (45%; 9 of 20 patients) was significantly higher than that in the CD44v6-negative patients (20%; 10 of 49 patients; P = 0.038), suggesting the involvement of CD44v6 in hematogeneous metastasis of lung carcinoma. Although the incidence of the expression of CD44v6 in squamous cell carcinoma was significantly higher than that in adenocarcinoma, histological type was not a significant prognostic factor. No significant correlation was found between the expression of CD44v6 and lymphatic or vascular vessel invasion, although lymphatic vessel invasion was found to be an independent prognostic factor in the multivariate analysis. To investigate the relationship between the expression of CD44v6 and proliferative activity of tumor cells, the expression of proliferating cell nuclear antigen (PCNA) was examined. The expression of CD44v6 was more frequently observed in PCNA-positive patients than in PCNA-negative patients (P = 0.019), but the expression of PCNA was not a statistically significant prognostic indicator. The results of multivariate analysis by the Cox proportional hazards model showed that CD44v6 was an independent prognostic indicator. We concluded that the expression of CD44v6 is a useful prognostic factor in stage I non-small cell lung carcinoma.

Spontaneous loss-of-function mutations of the 8-oxoguanine DNA glycosylase gene in mice and exploration of the possible implication of the gene in senescence.

8-Oxoguanine is one of the major premutagenic oxidative base legions in vivo and is suspected to play a crucial role in various pathophysiological processes, such as cancer and aging. Mammalian 8-oxoguanine DNA glycosylase (OGG1) is thought to play a major role in the removal of 8-oxoguanine adducts in vivo. We have identified several inbred mouse strains with a spontaneous mutation, OGG1-R336H or double mutations, OGG1-R304W/R336H. R304W mutation caused a complete loss of OGG1 activity, while the R336H mutation led to disruption of nuclear localization of the enzyme although the activity remained normal. Among the double mutants was SAMP1, which exhibits accelerated senescence and short lifespan. We assessed the possible implication of the mutant OGG1 and 8-oxoguanine in aging utilizing SAMP1 mice. SAMP1 retained 1.5- to 1.9-fold increase in 8-oxoguanine level of hepatic nuclear DNA as compared with normal mice, until at least 12 months of age. A genetic association study, however, indicated that the mutant Ogg1 gene per se is not responsible for the accelerated senescence and short lifespan of SAMP1. Mutant OGG1 may be associated with pathologic conditions in other mouse strains.

Development of congenic strains of mice carrying amyloidogenic apolipoprotein A-II (Apoa2c). Apoa2c reduces the plasma level and the size of high density lipoprotein.

A congenic strain of mice with amyloidogenic apolipoprotein A-II (Apoa2c) on the genetic background of the amyloidosis-resistant SAM-R/1 strain was produced by 12 generations of backcrossing. Genome mapping using endogenous murine leukemia proviral markers was done in the congenic strain, termed R1.P1-Apoa2c. We confirmed that only a small region surrounding the apoA-II gene on chromosome 1 was transferred from the genome of the donor SAM-P/1 strain. The level and particle size of plasma high density lipoprotein were decreased in R1.P1-Apoa2c mice compared to those in the progenitor SAM-R/1 mice. The function of apoA-II can be studied using this strain of mice.

Immune abnormality in relation to nonimmune diseases in SAM mice.

A series of related strains of senescence-accelerated mouse (SAM) shows strain-unique age-related diseases, such as amyloidosis, deficit in learning and memory, osteoporosis, and brain atrophy, while many of these disease-prone mouse (SAMP) strains have impaired immune activity as young adults, and have a short life span, probably not due directly to the diseases. Because the mean life span was prolonged and the time of the disease onset was delayed by a low-calorie dietary condition or a specific pathogen-free environment, both of which ameliorate the impaired immune activity, the enhancement of immune activity may help decrease the deteriorative process of aging, to that seen in ordinary strains of mice. Studies using the SAMP model may help elucidate the role of immunity in the aging process. Herein, we review the cellular and genetic basis of the immune abnormality in SAMP mice, then discuss the relationship between immune abnormality and development of the age-related disease, senile amyloidosis, findings obtained on SAMP hybrid mice and congenic mice for disease-related genes. Activation of the gene(s) for senile amyloid per se shortened the life span, and the early development of the immune dysfunction primarily seems to be both genetically and physiologically independent of amyloidosis, although the disease may be indirectly modified in the aged with depressed immune activity.

Differentiation induction in non-lymphocytic leukemia cells upon treatment with mycophenolate mofetil.

Inosine 5'-monophosphate (IMP) dehydrogenase catalyzes the rate-limiting reaction of guanine nucleotide biosynthesis and has been implicated in the reaction of cell growth and differentiation. We investigated the ability of mycophenolate mofetil, a prodrug of mycophenolic acid, to induce differentiation in HL-60 and U937 leukemic cells as well as in fresh leukemia cells from patients with non-lymphocytic leukemia. Treatment with mycophenolate mofetil reduced the intracellular guanosine 5'-triphosphate (GTP) levels and induced morphologic and functional differentiation in HL-60 and U937 cells dose-dependently. HL-60 and U937 cells developed macrophage-like cytoplasm as well as the expression of CD11b and CD14 antigens and the ability to oxidize nitroblue tetrazorium (NBT). These changes became evident when the intracellular GTP levels decreased to approximately 20-30% of the untreated control level and were abrogated by the addition of guanosine. In the fresh leukemic cells, differentiation induction was shown in the cells derived from seven of 13 patients. The fresh leukemia cells responding to mycophenolate mofetil revealed significant higher positivity to CD11b, CD14, and NBT before treatment and significantly reduced intracellular GTP levels after treatment compared to the non-responding cells. These findings suggest that mycophenolate mofetil induces differentiation in HL-60 and U937 cells and some fresh leukemia cells with moderate tendency to maturation, by causing a decrease in the intracellular GTP levels. Mycophenolate mofetil could be a promising differentiation inducer in vivo.

Modification of beta 2-microglobulin with D-glucose or 3-deoxyglucosone inhibits A beta 2M amyloid fibril extension in vitro.

beta 2-microglobulin (beta 2M) is a major constituent of amyloid fibrils (fA beta 2M) deposited in patients with A beta 2M amyloidosis. Recently, advanced glycation end products (AGE) of beta 2M and fA beta 2M have been suggested to play an important role in the pathogenesis of A beta 2M amyloidosis. We first characterized the states of AGE modification of fA beta 2M. Western blot analysis with a monoclonal anti-AGE antibody showed that purified fA beta 2M was naturally modified with AGE. Immunohistochemical studies of amyloid-deposited tissue have revealed a patchy distribution of the AGE-modified area in the amyloid deposits. Then we modified beta 2-m either with D-glucose or with 3-deoxyglucosone (3-DG) and investigated the effect of these modification on fA beta 2M extension in vitro, using the recently established first-order kinetic model of fA beta 2M extension in vitro. Western blot analysis and enzyme linked immunosorbent assay with a monoclonal anti-AGE antibody showed that these sugar-modified beta 2M contained AGE. During the incubation of fA beta 2M with native beta 2-m at 37 degrees C, the fluorescence of thioflavin T increased without a lag phase and proceeded to equilibrium. On the contrary, only a slight increase in fluorescence was observed during the incubation of fA beta 2M with sugar-modified beta 2M. Moreover, sugar-modified beta 2M exhibited a dose-dependent inhibitory effect on the extension reaction of fA beta 2M with native beta 2M. These results may suggest that in some in vivo situations, the modification of beta 2-m with AGE could play an inhibitory role for the formation of fA beta 2M.

Extension of A beta2M amyloid fibrils with recombinant human beta2-microglobulin.

In order to elucidate the pathogenesis of A beta2M amyloidosis, we established an experimental system to study the mechanism of amyloid fibril formation or degradation in vitro. We compared the kinetics of A beta2M amyloid fibril (fA beta2M) extension with native beta2microglobulin (n-beta2M) purified from the urine of a patient suffering from renal insufficiency, with that with recombinant beta2M (r-beta2M) in vitro. n-Beta2M and r-beta2M were incubated with fA beta2M purified from synovial tissues excised from A beta2M amyloidosis patients. The fA beta2M extension reaction could be explained by a first-order kinetic model in both beta2Ms. The extension reaction was greatly dependent on the pH of the reaction mixture and maximum around pH 2.5-3.0 in both beta2Ms. The fA beta2M extended with both beta2Ms assumed the similar helical filament structure, although the fibrils extended with r-beta2M were slightly wider than those extended with n-beta2M and the former fibrils assumed a helical structure more clearly as compared to the latter. In order to obtain pure, unmodified fA beta2M, we next extended fA beta2M repeatedly by the algorithmic protocol with r-beta2M. As the generation of the extended fibrils proceeded, the initial rate of the extension reaction increased The ultrastructure of fibrils was completely preserved throughout the repeated extension steps. Sodium dodecyl sulfate polyacrylamide gel electrophoresis and immunoblotting revealed that fA beta2M extended repeatedly with r-beta2M were composed solely of r-beta2M. The use of these r-beta2M and fA beta2M will be advantageous to assess the effects of several amyloid-associated molecules in the formation or degradation of fA beta2M in vitro.

Accelerated senile amyloidosis induced by amyloidogenic Apoa-II gene shortens the life span of mice but does not accelerate the rate of senescence.

The SAMP1 strain is a mouse model for accelerated senescence and severe senile amyloidosis. We studied the effects of the amyloidogenic apolipoprotein A-II gene (Apoa2c) on senile amyloidosis and the life span and progress of senescence of congenic mice (R1.P1-Apoa2c) which have Apoa2c of the SAMPI strain on the genome of the normally aging SAMR1 strain. Age-associated and severe amyloid deposits were detected in R1.P1-Apoa2c, as well as a 20% shorter life span than that of SAMR1. The scores of senescence increased more rapidly with age in R1.P1-Apoa2c than that of SAMR1, and the Gompertz function showed a bigger Y intercept but the same slope of regression line. These results suggest that severe senile amyloidosis induced by the Apoa2c gene shortens the life span of mice but does not accelerate the rate of senescence.

Apolipoprotein E and alpha 1-antichymotrypsin in dialysis-related amyloidosis.

Dialysis-related amyloidosis as represented by carpal tunnel syndrome is a serious complication of long-term dialysis treatment of patients with chronic renal failure. beta 2-microglobulin has been identified as a structural component of the amyloid deposits, but other factors also are associated with amyloid formation. We recently demonstrated the presence of apolipoprotein E and alpha 1-antichymotrypsin in the amyloid deposits. We therefore analyzed how polymorphic variants of both genes were related to the onset of amyloidosis. Among the apolipoprotein E genotypes, allele epsilon 2 represented a protective factor that delayed the onset of disease. In contrast, polymorphic alpha 1-antichymotrypsin alleles had no effect on the onset of amyloidosis. Thus, the apolipoprotein E epsilon 2 allele can be added to the list of factors that determine the onset of dialysis-related amyloidosis, which include patient age at initiation of dialysis therapy, dialysis duration, and the dialysis membrane used.

Hepatic tumor rupture following effectual treatment with irinotecan in a patient with highly refractory malignant lymphoma.

We describe a patient with highly refractory malignant lymphoma who died of hepatic tumor rupture following treatment with irinotecan (CPT-11). This 60-year-old man with non-Hodgkin's lymphoma (diffuse large B-cell lymphoma) demonstrated disease recurrence in the liver and the vertebrae following high-dose chemotherapy and autologous hematopoietic stem cell transfusion. He was treated with CPT-11 at a dose of one third of the conventional dose used for non-Hodgkin's lymphoma in Japan. The tumor in the liver markedly decreased in size but then ruptured. Although pathologic hepatic tumor rupture is a rare complication in patients with malignant lymphoma of the liver, this case demonstrates that hepatic tumor rupture may occur in refractory malignant lymphomas that reveal extensive degradation by this new, effective salvage therapy.

Torsade de pointes associated with hypokalemia after anthracycline treatment in a patient with acute lymphocytic leukemia.

Severe dose-dependent anthracycline cardiotoxicity is reported to cause myocardial damage resulting in congestive heart failure. However, torsade de pointes, a life-threatening arrhythmia caused by chronic anthracycline cardiotoxicity, has not been reported previously. A 16-year-old girl who developed torsade de pointes after 6 months of chemotherapy for acute lymphocytic leukemia (French-American-British classification L2) is described. When the patient was readmitted to the hospital because of syncope, peripheral blood and bone marrow analysis indicated a relapse. In addition, the patient was hypokalemic. Twenty-four-hour ambulatory electrocardiographic monitoring demonstrated QT prolongation and an episode of torsade de pointes. The electrocardiographic changes and arrhythmia improved after correction of the hypokalemia. An inverse correlation between leukocyte count and hypokalemia was observed. The patient died from pulmonary hemorrhage. Autopsy examination demonstrated myocardial degeneration consistent with damage induced by antineoplastic antibiotics. The cumulative dose of anthracycline and anthraquinone was less than the conventional dose limit associated with chronic cardiotoxicity, even for children who are more sensitive to anthracyclines. Torsade de pointes can occur in the setting of chronic anthracycline cardiotoxicity. Therefore, children or young adults who are more sensitive to anthracycline need careful observation that includes electrolyte monitoring, especially for potassium.

MR of pituitary metastasis in a patient with diabetes insipidus.

We present a case of acute-onset diabetes insipidus in a 69-year-old man who had been treated for lung cancer. T1-weighted MR images showed a thickened pituitary stalk and absence of the normal high intensity of the posterior pituitary lobe. Dynamic imaging demonstrated poor enhancement in the posterior lobe, whereas the anterior lobe was strongly enhanced. Autopsy revealed that metastatic tumor from lung cancer had infiltrated the posterior lobe as well as the pituitary stalk.

Expression of E-cadherin and lymph node metastasis in resected non-small-cell lung cancer.

This study was conducted to clarify the relationship between E-cadherin expression on tumor and lymph node metastasis as well as its prognostic roles in resected non-small-cell lung cancer. Two hundred forty-nine patients, who underwent surgical resection (stage I-IIIA), were examined. Paraffin-embedded sections of the primary tumors in all cases and of the metastatic lymph nodes in stage IIIA disease were stained with a monoclonal antibody against E-cadherin. Decreased expression of E-cadherin correlated with pathologic stage, tumor size, lymph node metastasis, and histological grade. The 5-year survival rate of E-cadherin-negative patients with stage IIIA disease was significantly lower than that of E-cadherin-positive patients. Multivariate analysis in stage IIIA disease indicated that E-cadherin was an independent prognostic factor. In the patients with clinical N0 tumors, the frequency of pathological N2 tumors was significantly higher in cases where the primary tumor was recognized as E-cadherin expression negative than in cases where the primary tumor was recognized as positive. Decreased E-cadherin expression showed correlation with presence of lymph node metastasis in resected non-small-cell lung cancer and with the prognosis of patients with stage IIIA disease.

Molecular remission induced by fractionated dose-escalating donor leukocyte infusion without graft-versus-host disease in a patient with chronic myelogenous leukemia relapsed after allogeneic bone marrow transplantation.

A 39-year-old male with CML who relapsed 5 years and 8 months after allogeneic bone marrow transplantation achieved complete molecular remission following fractionated dose-escalating donor leukocyte infusions. Acute or chronic graft-versus-host disease (GVHD) did not occur and the patient remained asymptomatic throughout treatment. Since no prophylaxis against GVHD was administered, this case indicated that the graft-versus-leukemia effect is entirely separate from GVHD in certain conditions.

Acceleration of chromosome aberrations in senescence-accelerated strains of mice.

Age-related changes in the frequency of chromosome aberrations were examined using bone marrow cells of senescence-accelerated strains of mice (SAM). An accelerated senescence-prone strain, SAM-P/1, showed a striking increase in the frequency of chromosome aberrations, from age 3 to 8 months, whereas an accelerated senescence-resistant strain, SAM-R/1, at the same ages showed only a slight increase. Both these strains were derived from the same ancestral strain (AKR/J). The rate of increase of chromosome aberration frequency paralleled the advancement of senescence in both strains. These observations suggest that there are genetic factors which closely relate to chromosomal instability and acceleration of the senescence processes.

The persistence of high uptake of serum albumin in the olfactory bulbs of mice throughout their adult lives.

Brain to plasma concentration ratios of i.v. administered human serum albumin (HSA) in the olfactory bulb, frontal cortex and cerebellum were evaluated in DDD mice of different ages. We measured the brain uptake of serum albumin excluding intravascular content by using a double isotope technique and examined the time course of the brain uptake to evaluate the brain uptake at different time intervals. In young adult mice, the value was significantly higher in the olfactory bulb than in other brain regions 3-24 h after (125)I-HSA injection. It was about 2.3 times higher in the olfactory bulb than in the cerebellum (P < 0.01). The high concentration ratios in the olfactory bulb were observed in all 4-22-month-old mice. Moreover, the ratio in the olfactory bulb 24 h after (125)I-HSA injection was higher in 22-month-old mice than in younger animals. The high uptake of serum albumin in the olfactory bulb suggests that intravascular macromolecules can be transported into the olfactory bulb more easily than in other brain regions with tight endothelium, and the persistence of high uptake during adult life may be associated with age-related morphological changes in the olfactory bulb.

Age-related changes in barrier function in mouse brain I. Accelerated age-related increase of brain transfer of serum albumin in accelerated senescence prone SAM-P/8 mice with deficits in learning and memory.

The time course of brain accumulation of radiolabelled human serum albumin ((125)I-HSA) injected intravenously and the transfer of (125)I-HSA from blood to brain were evaluated in DDD mice using a double isotope technique. The brain accumulation of (125)I-HSA at 3 and 9 h but not at 24 h postinjection and the brain transfer rates were significantly higher in 22-month-old DDD mice than in 4-month-old ones. The brain transfer rates of (125)I-HSA were measured also in senescence accelerated prone mice (SAM-P/8) with age-related deficits in learning and memory, and in senescence accelerated resistant mice (SAM-R/I) without these deficits. The brain transfer rates were significantly higher in 13-month-old SAM-P/8 and 22-month-old SAM-R/1 than in 3-month-old mice of the same strains, respectively. The mean brain transfer rates in five regions observed in 22-month-old DDD mice, 22-month-old SAM-R/1 and 13-month-old SAM-P/8 increased by 31%, 41% and 51% compared with corresponding values in 3- or 4-month-old mice of the same strains. DDD mice and SAM-R/1 mice with normal characteristics of aging showed similar age-related significant changes in brain transfer rates. Age-related increase in the brain transfer rate was manifested at the youngest age in SAM-P/8 among the three strains examined. These findings show that the transfer of human serum albumin into the mouse brain increases with aging and suggest that the barrier function in the mouse brain against macromolecules changes with aging.