Biomedical subjects

K Nakakuki

Publications and source records attributed to K Nakakuki.

At least 19 recordsLinked to original sources

PubMed1998

Apolipoprotein E and antioxidants have different mechanisms of inhibiting Alzheimer's beta-amyloid fibril formation in vitro.

We compared the mechanisms of apolipoprotein E- (apoE-) and antioxidant- (AO-) mediated inhibition of beta-amyloid fibril (fA beta) formation in vitro, based on a nucleation-dependent polymerization model using fluorescence spectroscopy with thioflavin T. We first applied a kinetic plot to transform a sigmoidal time-course curve of fA beta formation from freshly prepared amyloid beta-peptides (A beta) into a straight line. Mathematical treatment of this plot demonstrated that the above-described sigmoidal curve is a logistic curve and provided us with a kinetic parameter t(1/2), the time when the rate of fA beta formation is maximum. t(1/2) of beta-amyloids (A beta) (1-42) and (1-40) were 18.7 +/- 1.7 min and 6.3 +/- 0.2 h, respectively (mean +/- SD, n = 3) and were independent of the initial A beta concentration examined. Although apoE extended t(1/2) of both A betas in a dose-dependent manner, AO did not. On the other hand, the final amount of fA beta formed was decreased by both apoE and AO dose-dependently. We then analyzed the effect of apoE and AO on the extension reaction of fA beta, based on a first-order kinetic model. Although apoE extended the time to proceed to equilibrium in a dose-dependent manner, AO did not. On the other hand, both apoE and AO dose-dependently decreased the final amount of fA beta formed. These results indicate that apoE and AO inhibit fA beta formation in vitro by different mechanisms and suggest the existence of multiple pharmacological targets for the prevention of fA beta formation.

Amyloid beta-Peptides↗

PubMed1998

Differences in the extent of inflammation caused by intratracheal exposure to three ultrafine metals: role of free radicals.

Nickel and cobalt, which belong to the same elemental group, are known to cause interstitial lung disease and bronchial asthma. The ability of these metals to injure lung cells and cause inflammation is likely to be important in their pathogenicity but comparative studies are rare. Additionally, ultrafine (uf) forms of these metals are used increasingly and there is little available information on their toxicity. Thus the inflammatory response following intratracheal instillation of ultrafine particles of Co, Ni, and TiO2 was compared. Physiological saline (PS) was used as a vehicle control and DQ12 quartz as a positive control. Male Wistar rats were intratracheally instilled with the 3 particle types at a dose of 1 mg suspended in physiological saline. At 1, 3, 7, 15, and 30 d after the injection, lung weight and the cellular and biochemical changes in bronchoalveolar lavage fluid (BALF) were determined. By all of the indices, Uf-Ni appeared to be the most injurious to the lung, causing severe and sustained inflammation, cytotoxicity and increased epithelial permeability. The next most toxic material was DQ12 quartz, with Uf-Co being closely similar in ability to cause inflammation. Uf-TiO2 was more active than the saline control in all of the indices, but was the least toxic of the particles studied. The present study reveals that three ultrafine particles of the same diameter are dramatically different in their ability to cause inflammation. The three ultrafines were compared as to their ability to cause free-radical damage to supercoiled plasmid DNA, and the result of free-radical activity was found to be Uf-TiO2 << Uf-Co = Uf-Ni. Difference in free-radical-generation activity therefore could underlie the difference in inflammation of these three ultrafine particle types.

Animals↗

PubMed1998

Metastatic tumor thrombus attached to a pacemaker electrode.

We present a patient with a large tumor embolus attached to a pacemaker electrode leading to multiple pulmonary emboli. At postmortem examination, this thrombus was composed of clusters of well-differentiated squamous cell carcinoma intermingled with fibrin. Tumor involvement was not evident in the myocardium, endocardium, or epicardium. The primary tumor was discovered in the lower intrathoracic esophagus. Tumor microemboli from the esophageal primary lesions may have accumulated around the pacemaker electrode due to turbulent flow in this region, producing a large and friable tumor embolus.

Aged↗

PubMed1997

Concentration-dependent inhibitory effects of apolipoprotein E on Alzheimer's beta-amyloid fibril formation in vitro.

Recently, many research groups have examined the effect of apolipoprotein E (apoE) on beta-amyloid fibril (betaAf) formation in vitro. However, their data were somewhat controversial and no exact kinetic assessment of the role of apoE has thus far been available. We examined the effect of human apoE on betaAf formation in vitro, starting with various concentrations of freshly prepared beta-amyloid(1-40) (beta1-40) and using fluorescence spectroscopy with thioflavine T. When 50 microM of beta1-40 was incubated with a 1:1000 to 1:100 molar ratio of apoE, a dose-dependent inhibitory effect of apoE was observed. Both the nucleation and extension phases of betaAf formation in vitro were inhibited by apoE. On the other hand, when 300 microM of beta1-40 was incubated with a 1:100 molar ratio of apoE, the inhibitory effect of apoE was completely abolished. We then focused our study on the kinetics of the inhibitory effect of apoE on the extension phase of betaAf formation in vitro, utilizing the recently established first-order kinetic model of betaAf extension in vitro [Naiki, H., & Nakakuki, K. (1996) Lab. Invest. 74, 374-383]. The mathematical treatment of the data suggests that apoE inhibits the extension of betaAf in vitro, by making a complex with beta1-40, thus eliminating free beta1-40 from the reaction mixture. The equilibrium association constant with beta1-40 was practically the same among the three major recombinant apoE isoforms. These results indicate that the effects of apoE on betaAf formation in vitro is differential and could settle some of the controversy about beta-amyloid-apoE interaction in vitro.

Alzheimer Disease↗

PubMed1997

Citrate uptake by isolated rat renal brush border membrane vesicles in cadmium-intoxicated rats.

Epidemiological Studies have reported that the formation of renal stones is often observed in workers exposed to cadmium (Cd). Citrate is known to be a protective factor against renal stone formation. We previously reported that direct exposure to high-level cadmium impaired the citrate uptake by the brush border membrane vesicles (BBMV), which may mean that the formation of renal stones from exposure to Cd could be prevented. In the present study, to clarify the mechanism of Cd-induced renal stone formation, the characteristics of BBMV-induced citrate uptake were determined in Cd-intoxicated rats. Cd intoxication was induced by daily subcutaneous injections of CdCl2 at a dose of 2 mg Cd/kg body wt per day for 17 to 20 days. The BBMV were prepared by the divalent cation precipitation method. Citrate uptake was measured by the Millipore rapid membrane filtration technique. The citrate uptake per mg protein was not significantly different between the two groups. The volume of BBMV was significantly reduced (50%) in Cd-intoxicated rats compared with that of control rats. Percentages of the equilibrium value were also significantly inhibited in Cd-intoxicated rats.

Animals↗

PubMed1997

Direct effect of inorganic mercury on citrate uptake by isolated rat renal brush border membrane vesicles.

Occupational exposure to mercury has long been associated with renal proximal injury and an increased incidence of proteinuria, as has such exposure to cadmium. Renal citrate excretion is very important with respect to acid-base balance since the metabolism of citrate generates three bicarbonate ions. In this study, we exposed isolated rat renal brush border membrane vesicles (BBMV) to mercury (Hg2+) and examined their citrate uptake characteristics. BBMV were prepared by the divalent cation precipitation method. Citrate uptake was measured by the Millipore rapid membrane filtration method. The preincubation of BBMV with 0.5 and 2 mM HgCl2 for 1 min significantly inhibited citrate uptake compared with that of BBMV without Hg preincubation. The analysis of the time course of citrate uptake during a 30-min preincubation of BBMV with 0.1 mM Hg2+ also revealed a significant reduction in the uptake compared with that of the control BBMV without preincubation. These findings indicate that the preincubation of BBMV with mercury results in a time- and concentration-dependent inhibition of citrate uptake.

Animals↗

PubMed1997

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.

Amyloidosis↗

PubMed1996

Temperature-sensitive parainfluenza type 1 vaccine virus directly accesses the central nervous system by infecting olfactory neurons.

Immunohistochemical investigation showed that intranasal inoculation of mice with a temperature-sensitive (ts) mutant of parainfluenza type 1 vaccine virus resulted in infection of some olfactory neurons as well as respiratory epithelial cells. It also disclosed the presence of viral antigens in glomeruli of the olfactory bulb but not in the secondary neurons (mitral and tufted cells). Polymerase chain reaction demonstrated the persistence of virus-specific nucleic acids in the olfactory bulb. These observations lead to the conclusion that parainfluenza virus, even with a ts phenotype, gains access to the central nervous system by infecting olfactory neurons.

Administration, Intranasal↗

PubMed1996

First-order kinetic model of Alzheimer's beta-amyloid fibril extension in vitro.

Recently, several studies have proposed models describing the mechanisms of Alzheimer's beta-amyloid fibril formation in vitro. However, these models are somewhat controversial and no exact kinetic analyses measuring the polymerization velocity as an indicator of the reaction, have thus far been available. We first formed beta-amyloid fibrils from a synthetic peptide, beta-amyloid(1-40), and determined the optimum conditions for quantitative fluorometry of these beta-amyloid fibrils with thioflavine T. Optimum fluorescence measurements of beta-amyloid fibrils were obtained at the excitation and emission wavelengths of 446 and 490 nm, respectively, with the reaction mixture containing 5 microM thioflavine T and 50 mM of glycine-NaOH buffer, pH 8.5. We then focused our study on the extension phase of beta-amyloid fibril formation in vitro. When beta-amyloid fibrils were incubated with monomeric beta-amyloid(1-40) in conditions where de novo seed formation does not occur, the extension of beta-amyloid fibrils was observed with electron microscopy. Quantitative fluorometry revealed that: (a) extension of amyloid fibrils proceeded by a pseudo-first-order exponential increase as measured by the fluorescence of thioflavine T; (b) the rate of extension was maximum around pH 7.5, and was dependent on the incubation temperature. Between 20 and 37 degrees C, good linearity was observed between the common logarithm of the initial rate and the reciprocal of the absolute temperature; (c) the rate of polymerization was found to be proportional to the product of beta-amyloid fibrils number concentration and the beta-amyloid(1-40) concentration; (d) the net rate of extension was the sum of the rates of polymerization and depolymerization. These results show that beta-amyloid fibril formation can be explained by a first-order kinetic model: i.e., the extension of beta-amyloid fibrils proceeds via the consecutive association of beta-amyloid(1-40) onto the ends of existing fibrils.

Alzheimer Disease↗

PubMed1996

Infection of human papillomavirus (HPV) and p53 over-expression in human female genital tract carcinoma.

Inactivation of p53 gene products either by mutation or by complex formation with E6 oncoprotein encoded by high risk HPV appears to be a common event in cervical carcinogenesis. This study was designed to clarify this association in 41 primary cervical, 15 endometrial, 3 ovarian and one rectal carcinomas. Polymerase chain reaction analysis revealed presence of high risk HPV in 36 (88%) cervical, 5 (33%) endometrial and none of ovarian and rectal carcinomas. HPV 16 was found in 14 cervical carcinomas, HPV 18 in 19 cervical and 2 endometrial carcinomas and HPV 33 in 28 cervical and 5 endometrial carcinomas. Expression of tumor suppressor protein p53 by using polyclonal antibody CM-1, was detected in 28 (68%) cervical, 7 (47%) endometrial, 2 (66%) ovarian and one (100%) rectal carcinoma. Twenty six cervical and 3 endometrial carcinoma cases were positive for both high risk HPV and p53. We conclude that beside cervical carcinoma HPV infection is not uncommon in endometrial carcinoma and in our experimental design there is no inverse correlation between HPV infection and p53 over-expression in a variety of the tumors analysed in the present study.

Adult↗

PubMed1995

Viremia induced by influenza virus.

A mouse model of influenza A/PR/8 virus infection was adopted to investigate the blood and various tissues of intranasally infected mice for the presence of viral RNA by using the nested polymerase chain reaction. The nucleoprotein gene was detected in the red blood cell fraction from 1 to 5 days post-inoculation, while it was found in the lung and brain up to 14 days and in the liver, spleen, kidney, heart, and skeletal muscle up to 7 days. The virus-specific messenger RNA was transiently found in these organs. When mice received the uv-inactivated virus, viremia did not occur. The prior transfer of the hyperimmune serum prevented pneumonia but not bronchitis, and viremia was totally abolished. These results suggest: (1) viremia occurs during the acute phase of infection, (2) the virus is present in various organs and there the virus gene is transiently expressed, and (3) the virus enters the blood stream possibly through capillaries of the infected alveolar wall. Viremia may influence the pathogenesis of influenza.

Animals↗

PubMed1995

In vivo induction of apoptosis by influenza virus.

Intranasal exposure of mice to influenza virus led to cell death in bronchial/bronchiolar epithelial cells, alveolar cells and lymphoid cells. These cells displayed fragmentation of nuclei and chromatin condensation. Nick end-labelling of DNA in situ confirmed that such apoptotic cells had fragmented DNA. These results suggest that the influenza virus induces apoptosis in vivo.

Animals↗

PubMed1995

Parainfluenza virus type 1 infects olfactory neurons and establishes long-term persistence in the nerve tissue.

A mouse model of Sendai virus infection was adopted to examine the in vivo neurovirulence of parainfluenza viruses. A nested polymerase chain reaction detected the Sendai virus nucleoprotein gene in the olfactory bulbs of intranasally infected mice for at least 168 days post-infection (p.i.) and virus-specific messenger RNAs for 28 days p.i. Viral proteins were histochemically detected in some olfactory neurons for 7 days p.i. They were also found in glomeruli of the olfactory bulbs but not in the mitral cells and the tufted cells. No virus was detected in the whole brain not including the olfactory bulbs. When mice were inoculated with UV-inactivated virus, the viral RNA was present in the olfactory bulbs for a short period of 14 days, with no demonstrable viraemia. These results demonstrate that the parainfluenza virus directly accesses the central nervous system via olfactory neurons and establishes long-term persistence in the nerve tissue.

Animals↗

PubMed1995

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.

Aged↗

PubMed1995

Efficacy of a temperature-sensitive Sendai virus vaccine in hamsters.

A temperature-sensitive HVJ-pB strain of parainfluenza type 1 (Sendai) virus was obtained from a persistently virus-infected cell culture. Intranasal inoculation of Syrian hamsters with the HVJ-pB temperature-sensitive mutant resulted in an abortive infection but induced a specific antibody response against Sendai virus without appreciable lesions in the respiratory tract. Prior exposure to the temperature-sensitive mutant protected hamsters from subsequent challenge with virulent wild-type virus. The efficacy of protection became apparent at 5 days after vaccination and lasted for at least 720 days. For practical use of the HVJ-pB vaccine, a single intranasal administration to newborn babies aged over 15 days is recommended.

Administration, Intranasal↗

PubMed1993

Over-expression of p53 protein in human laryngeal carcinoma.

We examined the expression of tumor-suppressor protein p53 in a variety of laryngeal carcinomas from 43 patients (25 primary, 13 metastatic and 5 recurrent cases), 13 normal laryngeal tissues and 7 benign laryngeal nodule biopsy specimens, using the polyclonal antibody CM-1. Previously we have reported a high incidence of ras mutations (51%) and human papillomavirus (HPV) infection (37%) in these laryngeal carcinomas. p53 protein was detected by immunohistochemistry in 65% of laryngeal carcinomas (60% of primary, 69% of metastatic and 80% of recurrent cases). No correlation was found between p53 over-expression and histological grading of the tumors. None of the specimens from normal larynx and laryngeal nodules revealed any detectable level of this protein. Furthermore, 11 (69%) of 16 HPV-positive cases and 17 (77%) of 22 cases with ras mutation showed variable grades of p53 expression. Twelve (71%) of 17 laryngeal carcinomas in current study having both p53 over-expression and ras mutation were moderately or poorly differentiated. Likewise, positivity for these 2 parameters was significantly increased in metastatic tumors (9 of 13 cases, 69%) as compared with primary and recurrent tumors (8 of 30 cases, 27%) (p < 0.01). Our results suggest that multiple factors are involved in this malignancy, and that the simultaneous over-expression of p53 and the presence of ras mutation may be related to the progression stage of laryngeal carcinoma.

Adult↗

PubMed1993

ras gene mutations and HPV infection are common in human laryngeal carcinoma.

To evaluate the role of ras activation and human papillomavirus (HPV) infection in laryngeal carcinoma, we analyzed tumor DNA from 43 cases, including 25 primary laryngeal tumors, 12 lymph-node and one skin metastases, and 5 recurrent laryngeal carcinomas. Thirteen normal laryngeal tissues and 7 benign laryngeal nodule biopsy specimens along with normal tissue surrounding laryngeal carcinoma in 2 cases were also included. The polymerase-chain-reaction technique was used to amplify DNA fragments containing codon 12 and 61 of H-, K- and N-ras, also HPV 16, 18 and 33 DNA, subsequently hybridized with sequence-specific oligonucleotides. DNA samples from 22 patients with laryngeal carcinoma revealed ras mutations (18 in N-ras codon 12, 6 in H-ras codon 61, and 3 in K-ras codon 61). Likewise, HPV DNA was found in 16 cases (HPV 16, 18 and 33 in 3 cases, 14 cases and 1 case respectively). ras mutations were significantly higher in metastatic tumors (10 of 13 cases) than in primary (11 of 25 cases) and recurrent laryngeal carcinomas (1 of 5 cases). HPV DNA was detected in 60% of recurrent, 44% of primary and 15% of metastatic tumors. Only 2 of the 13 normal laryngeal tissues and 1 out of 7 laryngeal nodule specimens were found to contain HPV DNA. These results suggest that ras activation, especially in N-ras codon 12.1 (GGT-->AGT) and HPV infection are 2 important factors in (multistage) laryngeal carcinogenesis. The ras mutation may be associated with metastatic ability of the tumor.

Adult↗

PubMed1993

Genetic analysis of murine senile amyloidosis.

BACKGROUND: Recent studies have suggested that not only the genotypes of apolipoprotein A-II, the precursor protein of murine senile amyloid fibrils, but also other genetic factors may contribute to the pathogenesis of murine senile amyloidosis. EXPERIMENTAL DESIGN: We investigated the mode of inheritance of murine senile amyloidosis, using 12-month-old and 14-month-old F1, F2 hybrids and backcrosses between SAM-P/1 and SAM-R/1. In SAM-P/1, the senescence process is accelerated and senile amyloidosis is evident, whereas in SAM-R/1, there is a normal aging process and no evidence of senile amyloidosis. In SAM-P/1 and SAM-R/1, the genotypes of apolipoprotein A-II are Gln/Gln and Pro/Pro, respectively, identified by restriction fragment length polymorphism of the apolipoprotein A-II gene for the restriction enzyme Cfr13I. RESULTS: Among hybrids and backcrosses, no senile amyloidosis was observed histopathologically, in Pro/Pro-type strains. Mild senile amyloidosis sparing the liver and spleen was observed in a significant percentage of Pro/Gln-type strains. Practical senile amyloidosis involving the liver and spleen was observed in all of the Gln/Gln-type strains. Quantitative fluorometric analysis with thioflavine T (Naiki H, Higuchi K, Matsushima K, Shimada A, Chen W-H, Hosokawa M, et al. Lab Invest 1990;62:768-73) revealed that the degree of murine senile amyloid fibril deposition was significantly decreased in the Gln/Gln-type hybrid and backcross strains, as compared with findings in SAM-P/1 and the degree of manifestation of accelerated senescence was significantly lower in the Gln/Gln-type hybrid and backcross strains than in the SAM-P/1. CONCLUSIONS: Murine senile amyloidosis is linked to the molecular type of apolipoprotein A-II (i.e., Gln-type apolipoprotein A-II), and is transmitted as an autosomal dominant manner with incomplete penetrance. The severity of murine senile amyloidosis is far more advanced in Gln/Gln-type strains than in Pro/Gln-type strains. Other genetic factors that determine the manifestation of accelerated senescence, may significantly contribute to the degree of murine senile amyloidosis.

Aging↗