Behavioral and physiological responses to intraperitoneal injection of zymosan in chicks.

Zymosan is a cell wall component of the yeast Saccharomyces cerevisiae and produces severe inflammatory responses in mammals. When zymosan is peripherally injected in mammals, it induces several behavioral and physiological changes including anorexia and hyperthermia. However, to our knowledge, behavioral and physiological responses to zymosan have not yet been clarified in birds. Therefore, the purpose of the present study was to determine if intraperitoneal injection of zymosan affects food intake, voluntary activity, cloacal temperature, plasma corticosterone (CORT) and glucose concentrations, and splenic gene expression of cytokines in chicks (Gallus gallus). Intraperitoneal injection of zymosan (2.5 mg) significantly decreased food intake, voluntary activity, and plasma glucose concentration, and increased plasma CORT concentration. The injection of 0.5 mg zymosan significantly increased cloacal temperature, while 2.5 mg zymosan had a tendency to increase it. Finally, 2.5 mg zymosan significantly increased the splenic gene expression of interleukin-1ß (IL-1ß), IL-6, IL-8, interferon-γ, and tumor necrosis factor-like cytokine 1A. The present results suggest that zymosan would be one of components which induces nonspecific symptoms including anorexia, hypoactivity, hyperthermia, and stress responses, under fungus infection in chicks.

Hypertension and hepatitis C virus infection are strong risk factors for developing late renal dysfunction after living donor liver transplantation: significance of renal biopsy.

BACKGROUND: Late renal dysfunction (LRD) after liver transplantation develops due to several factors such as viral hepatitis, calcineurin inhibitor, diabetes mellitus, and hypertension. The aim of our study was to clarify the risk factors for LRD after living donor liver plantation (LDLT) by using simple criteria for LRD and paying special attention to the significance of renal biopsy. PATIENTS AND METHODS: Among the 98 recipients undergoing LDLT between March 2002 and June 2008, there were 77 patients who survived more than 1 year and had been followed at our clinic. LRD was simply defined as a postoperative serum creatinine level of 1.5/L or more at any point in time after 1 year from undergoing LDLT. The perioperative risk factors for developing LRD after LDLT were analyzed by uni- and multivariate analyses, and regardless of serum creatinine level, a renal biopsy was indicated when the patient developed clinical symptoms. RESULTS: Comparing the risk factors between 22 patients with LRD and 55 without LRD, univariate analysis revealed recipient's age, generation, hypertension, hepatitis C virus (HCV) antibody-positive, pretransplantation serum creatinine level, and graft-to-recipient weight ratio to be significant risk factors. By multivariate analysis, HCV and hypertension were selected as independent risk factors. Renal biopsy was indicated in the 4 patients with proteinuria, all of whom were positive for HCV. However, by histologic and/or electron micrographic analyses, only 1 patient was diagnosed with HCV-related membranous proliferative nephritis, 1 with diabetic nephropathy, and 2 with drug (tacrolimus) -induced renal dysfunction. CONCLUSION: Although HCV and hypertension were determined to be independent risk factors for LRD after LDLT, a renal biopsy should be performed when clinical symptoms develop regardless of creatinine levels to provide appropriate treatment.

Dynamic contrast-enhanced MRI of Implanted VX2 tumors in rabbit muscle: comparison of Gd-DTPA and NMS60.

We studied the dynamics of injected contrast enhancement in implanted VX2 tumors in rabbit thigh muscle. We compared two contrast agents Gd-DTPA and NMS60, a novel gadolinium containing trimer of molecular weight 2.1 kd. T1-weighted spin echo images were acquired preinjection and at 5-60 min after i.v. injection of 0.1 mmol/kg of agent. Dynamic T1-weighted SPGR images (1.9 s/image) were acquired during the bolus injection. Male NZW rabbits (n = 13) were implanted with approximately 2 x 10(6) VX2 tumor cells and grew tumors of 28+/-27 mL over 12 to 21 days. NMS60 showed significantly greater peak enhancement in muscle, tumor rim, and core compared to DTPA in both T1-weighted and SPGR images. NMS60 also showed delayed peak enhancement in the dynamic scans (compared to Gd-DTPA) and significantly reduced leakage rate constant into the extravascular space for tumor rim (K21 = 5.1 min(-1) vs. 11.5 min(-1) based on a 2 compartment kinetic model). The intermediate weight contrast agent NMS60 offers greater tumor enhancement than Gd-DTPA and may offer improved regional differentiation on the basis of vascular permeability in tumors.

Brain-derived neurotropic factor prevents superoxide anion-induced death of PC12h cells stably expressing TrkB receptor via modulation of reactive oxygen species.

In our previous report (Satoh et al., 1999. Regulation of reactive oxygen species by nerve growth factor but not by Bcl-2 as a novel mechanism of protection of PC12 cells from superoxide anion-induced death. J. Biochem. 125, 952-959), we reported that nerve growth factor (NGF) protected PC12 cells from superoxide anion (O2-)-induced cell death through a novel regulation of reactive oxygen species (ROS) which increased O2- and decreased hydrogen peroxide (H2O2), indicating that decreasing conversion from O2- to H2O2 is a critical process for the protection by NGF. In the present study, we performed a comparative study on protective mechanisms between NGF and brain-derived neurotrophic factor (BDNF) using TrkB-expressing PC12h cells. When compared with NGF, BDNF induced a weaker but significant protective effect on the cells from O2- induced death. BDNF did not seem to change the total amount of ROS in the cells treated with xanthine and xanthine oxidase. On the other hand, BDNF increased O2- and decreased H2O2- levels in the same cells, although not so strongly as NGF. These results suggest that decreasing conversion from O2- to H2O2 is also critical for the protection by BDNF, which is considered to play a central role in survival and differentiation of CNS neurons.

Brain-derived neurotrophic factor stimulates interactions of Shp2 with phosphatidylinositol 3-kinase and Grb2 in cultured cerebral cortical neurons.

Shp2, a protein tyrosine phosphatase possessing SH2 domains, is utilized in the intracellular signaling of various growth factors. Shp2 is highly expressed in the CNS. Brain-derived neurotrophic factor (BDNF), a member of the neurotrophin family, which also shows high levels of expression in the CNS, exerts neurotrophic and neuromodulatory effects in CNS neurons. We examined how BDNF utilizes Shp2 in its signaling pathway in cultured cerebral cortical neurons. We found that BDNF stimulated coprecipitation of several tyrosine-phosphorylated proteins with anti-Shp2 antibody and that Grb2 and phosphatidylinositol 3-kinase (PI3-K) were coprecipitated with anti-Shp2 antibody in response to BDNF. In addition, both anti-Grb2 and anti-PI3-K antibodies coprecipitated Shp2 in response to BDNF. The BDNF-stimulated coprecipitation of the tyrosine-phosphorylated proteins, Grb2, and PI3-K with anti-Shp2 antibody was completely inhibited by K252a, an inhibitor of TrkB receptor tyrosine kinase. This BDNF-stimulated Shp2 signaling was markedly sustained as well as BDNF-induced phosphorylation of TrkB and mitogen-activated protein kinases. In PC12 cells stably expressing TrkB, both BDNF and nerve growth factor stimulated Shp2 signaling similarly to that by BDNF in cultured cortical neurons. These results indicated that Shp2 shows cross-talk with various signaling molecules including Grb2 and PI3-K in BDNF-induced signaling and that Shp2 may be involved in the regulation of various actions of BDNF in CNS neurons.

MS-430, a synthetic pyrimidine derivative, influences the intracellular signal transduction pathway leading to neuronal differentiation of PC12h cells.

Although NGF (nerve growth factor) induces neuronal differentiation of PC12 cells, EGF (epidermal growth factor) acts as a mitogen for these cells. We have studied the effects of a synthetic pyrimidine derivative, MS-430, on the NGF and EGF actions on PC12h cells. We found that MS-430 accelerated NGF-induced neurite extension of PC12h cells and that, in the presence of MS-430, PC12h cells extended neurites in response to EGF. Next, we investigated the tyrosine phosphorylation of NGF receptor TrkA and the EGF receptor (EGFR) as well as mitogen-activated protein kinase (MAPK), which is a key protein in the intracellular signal transduction pathway. It was found that MS-430 prolonged the EGF-induced phosphorylation of EGFR and MAPK compared to that without MS-430. MS-430 also prolonged the NGF-induced phosphorylation of MAPK, but the phosphorylation of TrkA induced by NGF was not affected by MS-430. These results suggest that MS-430 influences the intracellular signal transduction pathway which causes the neuronal differentiation of PC12h cells.

Comparison of survival-promoting effects of brain-derived neurotrophic factor and neurotrophin-3 on PC12h cells stably expressing TrkB receptor.

We obtained two PC12h cell lines, PC-pAB1 and PC-pAB2, stably expressing TrkB receptor and investigated the effects of BDNF and NT-3 in these cell lines. The cells differentiated into neuron-like cells in response to BDNF as well as NGF, neurite extension being more rapid in the former case. These TrkB-expressing cells also extended neurites in response to NT-3, which is a nonpreferred ligand of TrkB. Next, we examined the survival-promoting effects of NGF, BDNF, and NT-3 under apoptotic conditions of oxygen toxicity in naive cells and NGF deprivation in differentiated cells. In both cases, BDNF prevented cell death similarly to NGF. NT-3 prevented cell death induced by oxygen toxicity in naive cells, but not that induced by NGF deprivation in differentiated cells. NT-3 induced the tyrosine phosphorylation of TrkB in naive cells, but not in differentiated cells. These results indicate that NT-3 has survival-promoting effects on naive TrkB-expressing PC12h cells, but not on differentiated cells because of its inability to induce the tyrosine phosphorylation of TrkB.

Mapping rat megalin: the second cluster of ligand binding repeats contains a 46-amino acid pathogenic epitope involved in the formation of immune deposits in Heymann nephritis.

Megalin (gp330), an epithelial endocytic receptor, is a major target antigen of Heymann nephritis (HN), an autoimmune disease in rats. To elucidate the mechanisms of HN, we have mapped a pathogenic epitope in megalin that binds anti-megalin antibodies. We focused our attention on four clusters of cysteine-rich, low density lipoprotein receptor (LDLR) ligand binding repeats in the extracellular domain of megalin because they represent putative ligand binding regions and therefore would be expected to be exposed in vivo and to be able to bind circulating antibodies. Rat megalin cDNA fragments I through IV encoding the first through fourth clusters of ligand-binding repeats, respectively, were expressed in a baculovirus system. All four expression products were detected by immunoblotting with two antisera capable of inducing passive HN (pHN). When antibodies eluted from glomeruli of rats with pHN were used for immunoblotting, only the expression product encoded by fragment II was detected. This indicates that the second cluster of LDLR ligand binding repeats is directly involved in binding anti-megalin antibodies and in the induction of pHN. To narrow the major epitope in this domain, fragment II was used to prepare proteins sequentially truncated from the C- and N-terminal ends by in vitro translation. Analysis of the truncated translation products by immunoprecipitation with anti-megalin IgG revealed that the fifth ligand-binding repeat (amino acids 1160-1205) contains the major epitope recognized. This suggests that a 46-amino acid sequence in the second cluster of LDLR ligand binding repeats contains a major pathogenic epitope that plays a key role in pHN. Identification of this epitope will facilitate studies on the pathogenesis of HN.

Cloning and characterization of mycovirus double-stranded RNA from the plant pathogenic fungus, Fusarium solani f. sp. robiniae.

A mycovirus (named FusoV) from the phytopathogenic fungus, Fusarium solani f. sp. robiniae SUF704, has two kinds of double-stranded (ds) RNA genomes, designated M1 and M2. The cDNAs were constructed from FusoV genomic dsRNAs. The sequences of M1 and M2 cDNAs comprised 1645 and 1445bp, respectively. Sequence analysis showed that each dsRNA had a single long open reading frame (ORF) on only one of the strands. M1 ORF encodes a 519-amino acid residue polypeptide with a predicted molecular mass of 60 kDa. RNA-dependent RNA polymerase-conserved motifs were identified in the predicted amino acid sequence, and the polymerase synthesized dsRNA in vitro. The M2 ORF encodes a polypeptide of 413 amino acid residues with a predicted molecular mass of 44 kDa. The predicted amino acid sequence contained the sequence corresponding to those found in the purified 44-kDa capsid protein of FusoV.

Sustained trilineage recovery and disappearance of abnormal chromosome clone in a patient with myelodysplastic syndrome following combination therapy with cytokines (granulocyte colony-stimulating factor and erythropoietin) and high-dose methylprednisolone.

We report a case of hypoplastic myelodyplastic syndrome (MDS) (refractory anemia (RA)) in which sustained trilineage haematological response and persistent disappearance of an abnormal chromosome clone were achieved after treatment with combination therapy of cytokines (granulocyte colony-stimulating factor (G-CSF) and erythropoietin (Epo)) and methylprednisolone (mPSL) pulse dose. The patient's haematological recovery was rapid and maintained even after cessation of the therapy. In addition, the predominant chromosome clone 13q- in bone marrow cells disappeared in the fourth week. The patient's improved bone marrow haemopoiesis and disappearance of the abnormal chromosome has continued to the present, 13 months after treatment. The occurrence of both trilineage response and abnormal chromosome disappearance in MDS patients treated with cytokine(s) or steroids is rare. Combination therapy might therefore be advantageous in MDS.

Usefulness of histopathologic examination of thick scales in the diagnosis of X-linked dominant chondrodysplasia punctata (Happle).

Most patients with X-linked dominant chondrodysplasia punctata (X-DCDP) have whorled, thick, or spiky adherent hyperkeratosis on their whole body. Histopathologically, there are large keratotic plugs of hair follicles where calcium deposits. This is the characteristic finding in this disease. We performed histopathologic examination of skin specimens from two newborn babies with X-DCDP. The desquamated thick scales from the hyperkeratotic lesions had calcium in the center of the keratotic plugs. Histopathologic examination of thick scales is easy and helpful in diagnosing X-DCDP.

The relationships of onset and exacerbation of pustulosis palmaris et plantaris to smoking and focal infections.

Clinical data, including focal infection and habitual cigarette smoking, were obtained from 203 male patients with pustulosis palmaris et plantaris (PPP) (age: 43.3 +/- 13.4) and 266 female patients (age: 44.0 +/- 13.7) for the 20 years from 1975 through 1994 to evaluate the relationship between the onset or severity of PPP and smoking. Seasonal incidences of onset were also studied. The incidence of onset of PPP symptoms was highest in June, when it is the most humid in Japan, and lowest in December. The most common infectious disease associated with PPP was tonsillitis. The percentages of heavy smoking (more than 20 cigarettes per day) were 74.7% and 32.9% for male and female patients, while those in the normal control population in Japan were 37.2% and 9.8% for males and females. These results suggest that heavy smoking, tonsillitis, and seasonal factors such as high humidity and high temperature may be related to the onset and exacerbation of PPP.

A unique case of renovascular hypertension caused by combined renal artery disease.

We present a unique case of renovascular hypertension due to combined renal artery disease in a 22-year-old woman. Renal angiography revealed renal artery stenosis with poststenotic dilatation and an aneurysm due to fibromuscular dysplasia in the left kidney, and a congenital arteriovenous fistula in the right kidney. The results of a captopril test and plasma renin sampling demonstrated that the renin-angiotensin-aldosterone system was stimulated in both kidneys, accounting for the hypertension in this patient. Almost all cases of renovascular hypertension are due to only one underlying renal artery disease. This is the first case of renovascular hypertension associated not only with renal artery stenosis and an aneurysm due to fibromuscular dysplasia, but also with a congenital arteriovenous fistula.

Sjögren's syndrome with pleural effusion.

Sjögren's syndrome (Sjs) can cause many organic changes, but is rarely accompanied by pleuritis. We report here a 62-year-old patient with subclinical Sjs who developed unilateral pleuritis with moderate effusion. He was diagnosed to have subclinical Sjs based on the positivity of anti SS-A/SS-B antibodies and the biopsy findings of minor salivary glands which revealed lymphocyte infiltration around the duct. In the pleural effusion, both increased lymphocytes and anti SS-A/SS-B antibodies were observed. He showed no signs of infection nor malignancy. There was no direct evidence that he had other collagen diseases which cause pleuritis. We conclude that the pleuritis was caused by Sjs. In patients with Sjs, activated polyclonal B lymphocytes and autoantibodies are considered to cause systemic tissue damage. This case indicates that these factors can cause pleuritis in Sjs patients.

Radioimmunoassay of follistatin: application for in vitro fertilization procedures.

A sensitive radioimmunoassay (RIA) for follistatin was developed by using antisera raised in rabbits against purified porcine ovarian follistatin. The displacement curves generated with human follicular fluid and serum were parallel with the standard curve of porcine follistatin. Using this RIA, we have measured human serum follistatin immunoreactivity in six women undergoing ovarian hyperstimulation for in vitro fertilization (IVF). After treatment with a GnRH agonist and gonadotropin, serum estradiol and follistatin levels were increased, and there was a direct positive correlation between the elevated levels of estradiol and follistatin (r = 0.93, p less than 0.01). Thus, the level of circulating follistatin may likely reflect follicular maturation and be applicable in IVF procedures to determine the optimal timing for oocyte retrieval.

[Echocardiographic assessment of mitral annular calcification in hypertrophic cardiomyopathy with asymmetric septal hypertrophy and healthy elderly cases].

The prevalence and degree of mitral annular calcification (MAC) in elderly cases of hypertrophic cardiomyopathy with asymmetric septal hypertrophy (ASHCM) were studied in comparison with healthy elderly and chronic hypertensive elderly with concentric left ventricular hypertrophy (HT + LVH) by echocardiography. In seven elderly patients with ASHCM (mean age 77.3 +/- 8.1 years) severe MAC was observed in all cases (1 male and 6 females) and systolic anterior motion of the mitral valve (SAM) was present in 3 cases, incomplete in 2 and absent in 2 cases. Of 57 healthy elderly with a mean age of 78.8 +/- 8.8 years, 8 subjects (1 male and 7 females, 14%) showed MAC. The prevalence of MAC was 0% in the sixties, 14% in the seventies, 16% in the eighties and 29% in the nineties. MAC was severe in four subjects above eighty years old. In 3 cases with HT + LVH with a mean age of 72.3 +/- 4.7 years no MAC was observed and SAM was present in one, partially observed in one and absent in one case, respectively. Serum calcium (Ca) and phosphorus (P), reported to be related with the formation of MAC, and fasting blood sugar level (FBS) were evaluated. In healthy elderly with MAC, serum Ca was 8.78 +/- 0.46 mg/dl, P 3.55 +/- 0.37 mg/dl and FBS 97.75 +/- 29.01 mg/dl, P 3.55 +/- 0.37 mg/dl and FBS 97.75 +/- 29.01 mg/dl.(ABSTRACT TRUNCATED AT 250 WORDS)

[Isolation and properties of myoglobin from murine skeletal muscle].

Myoglobin (Mb) was isolated from skeletal muscle of JCL-ICR mice by heat denaturation-gel filtration combined with ion exchange chromatography or chromatofocusing by which isoelectric point of the main component was estimated as 7.63 +/- 0.09 (20 degrees C). The Mb was homogeneous by gel electrophoretic and ultracentrifugal analysis. The molecular weight by sedimentation equilibrium was 1.80 X 10(4) and essentially identical with the values by the iron analysis (1.82 X 10(4)) and amino acid composition (1.78 X 10(4)) in which one residue of cysteine was found per molecule. The spectroscopic properties of deoxy-, oxy-, carboxy- and ferri-derivatives of the protein were determined in ultraviolet, Soret and visible regions. The pK' of acid-alkaline transition of the ferri-form was estimated as 8.57 +/- 0.30 (30 degrees C) from the pH-dependent spectral changes. The oxygen equilibrium studies revealed complete absence of such allosteric properties as heme-heme interaction, anion effect and Bohr effect. Oxygen tension for the half-oxygenation (P50) was 0.69 +/- 0.06 Torr (20 degrees C) and its temperature-dependent change gave the delta H degrees of -14.1 kcal/mole.

Fetal ascites. A report of 3 autopsy cases.

Three rare autopsy cases of fetal ascites were presented and the etiology of each case was described. Case 1 was a male neonate, delivered by cesarean section at 32 weeks' gestation, and died of respiratory failure. The abdomen was remarkably distended with 1020 ml of ascites. The etiology of Case 1 remained unknown even after macroscopic and microscopic examinations. We considered this as "idiopathic" fetal ascites. Case 2 was a female neonate, delivered at 31 weeks' gestation, with marked abdominal distension and cyanosis. Autopsy revealed 435 ml of ascites, and she was considered to have had "polysplenia syndrome" with cardiovascular malformations. Intrauterine heart failure due to cardiac anomalies was thought to be the cause of this ascites. In case 3 embryotomy was carried out under the diagnosis of fetal ascites by ultrasound examination at 22 weeks' gestation. An urachal cyst connected to the dilated urinary bladder and deficiency of musculature of the abdominal wall composed of loose connective tissue with calcification were observed. The abdominal wall was ruptured and 1,960 ml of ascites was measured. Polycystic kidney with renal dysplasia was also found. Case 3 showed "Prune-Berry syndrome" and fetal ascites may have arisen from these anomalies.