Gene expression profile in rat renal isografts from brain dead donors.

BACKGROUND: Brain death (BD) and the subsequent ischemia/reperfusion (I/R) injury have cardinal implications for the pathogenesis of kidney transplantation (Tx). However, the precise mechanistic pathway of BD and the subsequent I/R injury are unknown. In this study, we performed genome-wide analysis for differential gene expression in kidney isografts from BD donors. Their gene expressions were compared with those from living sources. METHODS: Kidneys from BD rats were engrafted and their gene expressions were compared with those from living controls. Donors were intubated, and mechanically ventilated for 6 hours. Grafts were harvested 6 hours after BD, and 1 hour after engraftment. The expression profile of approximately 20,500 genes was analyzed. RESULTS: Gene expression of chemokines (Scya2 and Gro1), cytokines (IL-1 and -6) and adhesion molecules (E- and P-selectin and ICAM-1) were upregulated in the BD kidneys and 1 hour after engraftment. An antiapoptotic gene (Birc2), IkappaB-zeta, and protective gene (HO-1) were also upregulated. Other upregulated genes included oncogenes (lipocalin2, Bcl3, and CCAAT/enhancer binding protein delta), Calgranulin B, DEXRAS1, insulin-like growth factor binding protein-1, inhibin beta-B-subunit gene, IgG Fc receptor, and FK 506 binding protein 5. We also observed downregulation of the genes Amphiphsin, Jagged 1, Pace 4, Slc15a2, Kcnn2, and gap junction membrane channel protein alpha5 only in kidneys from BD donors. CONCLUSIONS: This is the first demonstration of global gene expression analysis using the rat brain-death isograft model. These results provide new insights for the detection of novel target genes for treatment and prognosis of grafts from brain-dead and extended marginal donors.

A pivotal role for DNase I-sensitive regions 3b and/or 4 in the induction of somatic hypermutation of IgH genes.

Chimeric mice were prepared from embryonic stem cells transfected with IgH genes as transgenes and RAG-2-deficient blastocysts for the purpose of identifying the cis-acting elements responsible for the induction of somatic hypermutation. Among the three transgene constructs used, the V(H) promoter, the rearranged V(H)-D-J(H), an intron enhancer/matrix attachment region, and human Cmu were common to all, but the 3'-untranslated region in each construct was different. After immunization of mice with a T cell-dependent Ag, the distribution and frequency of hypermutation in transgenes were analyzed. The transgene lacking the 3' untranslated region showed a marginal degree of hypermutation. Addition of the 3' enhancer resulted in a slight increase in the number of mutations. However, the transgene containing DNase I-sensitive regions 3b and 4 in addition to the 3' enhancer showed more than a 10-fold increase in hypermutation, reaching levels comparable to those observed in endogenous V(H)186.2 genes of C57BL/6 mice.

Immunohistochemical localization of taurine in various tissues of the mouse.

The localization of taurine was investigated in several tissues of the mouse. Immunohistochemical methods using a polyclonal antibody for taurine derived from rabbits was used in these studies. This method was used since it is a simple procedure and the results are clear and reliable. Tissues were fixed with paraformaldehyde, embedded in paraffin and treated in a microwave oven before using an avidin-biotin-complex method (ABC method). Control staining was accomplished by employing absorption staining using various amino acids: taurine, arginine, cysteine, hypotaurine and others. For purposes of comparison, radioautography (RAG) with 3H-taurine was performed to confirm the reliability of the immunohistochemical staining compared with the localization of the 3H-taurine incorporation in endothelial cells of the blood vessels of several tissues. In this investigation, immunoreactivity was broadly observed in many tissues: Purkinje cells of the cerebellum, glia cells of brain tissue, cardiac muscle cells, matrices of the bone, mucus granules of goblet cells of the intestines, and brown adipose cells of the fetus. Although the meaning of this widespread localization of taurine can not be explained completely, we surmise that taurine may have a different function in each of the tissues. In addition, taurine reactivity was observed in cell nuclei which was evidence of the presence of taurine in the nuclei.

An autopsy case of mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes (MELAS) with a point mutation of mitochondrial DNA.

A 14-year-old boy with mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes (MELAS) is reported. He had suffered blepharoptosis and cataracts prior to the stroke-like episodes, and was thus reported in 1984 as having Kearns-Shy (Sayre) syndrome. After his death, an A-to-G mutation of the mitochondrial DNA (mtDNA) at bp 3243 was identified in cardiac muscle and the liver. Neuropathologically, multiple old and recent necrotic foci were observed in the gray and white matter of the cerebrum and cerebellum. These lesions were occasionally observed in areas outside of the distribution of major blood vessels of the brain. In the recent necrotic foci, neural loss and sponginess were observed while some neurons were preserved intact. The latter finding has not been described in MELAS and suggests that metabolic degeneration had occurred in the neurons of this patient. This is the first report of a confirmed 3243 mutation of the mtDNA in an autopsied MELAS case.

Isolation and amino acid sequence of a molt-inhibiting hormone from the American crayfish, Procambarus clarkii.

A molt-inhibiting hormone (Prc-MIH) was isolated from the sinus glands of the American crayfish, Procambarus clarkii, and its amino acid sequence was determined. It comprised 75 amino acid residues and had an amidated carboxyl terminus. The amino acid sequence was much more similar to MIH of the shore crab (Cam-MIH) than to MIH of the American lobster (Hoa-MIH).

Amino acid sequence of a peptide with molt-inhibiting activity from the kuruma prawn Penaeus japonicus.

Six major peptides (Pej-SGP-I-VI) that belong to the CHH family have been isolated from the sinus gland extracts of the kuruma prawn Penaeus japonicus. By in vitro assay using the Y-organ of the crayfish Procambarus clarkii, Pej-SGP-IV was found to be active in inhibiting ecdysteroid synthesis. We determined the complete amino acid sequence. Pej-SGP-IV consists of 77 amino acid residues, with both free aimno- and carboxyl-termini. The sequence of Pej-SGP-IV shows considerable similarity to that of MIH of the shore crab Carcinus maenas and less similarity to Pej-SGP-III, whose sequence has been previously determined.

Observation on incorporation of 3H-taurine in mouse skeletal muscle cells by light and electron microscopic radioautography.

Taurine, one of the sulfur-containing amino acids, exist abundantly in the skeletal muscle tissues. The physiological function and ultrastructural cellular localization of taurine in the skeletal muscle cells are not clear. In this report, two experiments by radioautography were made to elucidate intracellular localization of this amino acid in the skeletal muscle cells. First, muscle tissue pieces obtained from normal and muscular dystrophy mice were cultured in a medium containing 3H-taurine and radioautographed. The number of silver grains appeared on muscle cells increased depending on the duration of the incubation time. Secondly, 3H-taurine was injected intraperitonealy in normal and muscular dystrophy mice. Then muscle specimens were fixed by two fixative procedures, one by a chemical fixation with glutaraldehyde and osmium tetroxide and another by cryo-fixation. Silver grains appeared over muscle cells prepared with both procedures. Silver grains were localized on myofilaments, sarcoplasmic membranes, sarcoplasmic reticulum, mitochondria, endothelial cells of blood capillaries and the cells of perineural sheath, but did not appear on Golgi apparatus, nuclei of muscle cells or adipose cells by any procedures. No difference in localization of silver grains was observed between normal and muscular dystrophy mice.

[Establishment and characterization of a new human germ cell tumor strain (NOC-Ts) in culture].

We have derived and characterized a new cell line from a teratoma with a embryonal carcinoma and seminoma. The medium used for the cell culture was Eagle's MEM synthetic culture medium (Gibco Inc.) supplemented with 10% new calf serum (Tissue plus, Mitsubishi Kasei Co.). Subcultures were performed on 3 to 4 days basis at 1:2 split by the use of 0.25% trypsin solution. The morphology of obtained cells was a epithelial-like shape and the cell grew in a monolayered sheet with about 30-32 hrs of population doubling time. The model chromosome number of this cell line was 73 including one large submetacentric marker chromosome. The temperature sensitivity and the tumorigenicity of this cell were tested in this experiment.

Subclinical pseudohypoparathyroidism type II: evidence for failure of physiologic adjustment in calcium metabolism during pregnancy.

Patients with latent disorders of hormone response mechanism are rarely found. This paper reports a case of subclinical pseudohypoparathyroidism type II in which physiological adjustment of calcium (Ca) metabolism became insufficient only in the second half of pregnancy. A 34-year-old woman examined for a slight bruise on the head was incidentally found to have marked intracranial calcification and a full set of false teeth. From her history of past pregnancy, it was revealed that she suffered from symptoms of hypocalcemia during late gestation (serum total Ca level, 4.8-6.4 mg/dl), which disappeared spontaneously after delivery. When the woman was not pregnant, although only the total Ca level was slightly below the normal range (7.7-8.4 mg/dl), the serum ionized Ca, phosphorus (P), magnesium, 1,25-dihydroxycholecalciferol and 24,25-hydroxycholecalciferol levels, plasma parathyroid hormone (PTH) level and urinary excretion of Ca were all normal without treatment. Intravenous infusion of 30 mg/kg EDTA-2Na resulted in marked elevation of plasma PTH associated with significant reduction of serum ionized Ca. In contrast, although her urinary excretion of phosphorous per hour was within the normal range in the basal state, she showed no proportional change in urinary phosphorous excretion with increase in urine cyclic AMP induced by administration of PTH(1-34). From these findings, she was diagnosed as having an incomplete form of pseudohypoparathyroidism Type II. This abnormality seems to be rare, but we consider that the present observations provide important information for preventive care of pregnant women and fetuses during gestation.

Facioscapulohumeral dystrophy associated with sensorineural hearing loss, tortuosity of retinal arterioles, and an early onset and rapid progression of respiratory failure.

Two sibling cases of facioscapulohumeral dystrophy (FSHD) are described. One was characterized by sensorineural hearing loss, marked tortuosity of retinal arterioles, an early onset and progression of severe restrictive-type pulmonary dysfunction, and cor pulmonale. The other had a mild course of FSHD without involvement of any other organ than muscles at the time of diagnosis. Recently, a new nosological entity of FSHD, with sensorineural hearing loss and tortuosity of retinal arterioles, was advocated. Our cases, especially the first case, seem to belong to this newly recognized entity of FSHD. Moreover, it is noteworthy that our first case exhibited rapid aggravation of severe restrictive-type respiratory failure and cor pulmonale, leading to death, which was never seen in any other reported cases.

Two cases of NADH-coenzyme Q reductase deficiency: relationship to MELAS syndrome.

Muscle biopsy specimens from two patients with MELAS syndrome (mitochondrial myopathy, encephalopathy, lactic acidosis, and strokelike episodes) were studied biochemically. 14CO2 production rates from (1-14C)pyruvate, (U-14C)malate, and (1-14C)2-ketoglutarate were all decreased in intact mitochondria in both patients. Rotenone-sensitive NADH cytochrome c reductase activities were decreased to 8% (patient 1) and 6% (patient 2) of control values; succinate cytochrome c reductase and cytochrome c oxidase values were within normal limits. These results indicate that both patients have a defect of NADH-CoQ reductase of the respiratory chain and that MELAS can be brought about by a defect of NADH-CoQ reductase.

Neurologic deterioration with progressive CT changes in a child with Kearns-Shy syndrome.

A case of the rare juvenile form of Kearns-Shy syndrome with progressive external ophthalmoplegia and lid ptosis, carditis, skeletal muscle weakness, seizures, mental subnormality, short stature, EEG abnormality and deafness is presented. Electromyography revealed a myopathic pattern. Histochemical studies on quadriceps biopsy specimens showed atrophy of type II fibers and "ragged-red fibers." On electron microscopy these muscle cells were seen to contain an increased amount of glycogen particles and abnormal mitochondria were increased in number and size. It is of interest that abrupt deterioration of neurological findings such as seizures, mental subnormality, speech disturbance and deafness was present in our case. Computed tomographic scanning showed progressive changes of cerebral atrophy, low density of cerebral white matter and basal ganglia calcification, which were well associated with the clinical deterioration. A review of the literature also indicated that some patients with this syndrome showed abrupt neurological deterioration in childhood. Involvement of the central nervous system in this syndrome has to be considered as the cause of sudden deterioration and death in childhood.

Role of thoracotomy in pulmonary metastases from gestational choriocarcinoma.

Choriocarcinoma responds well to chemotherapy, and its growth can be monitored by measuring human chorionic gonadotropin (HCG) as a tumor marker. Thoracotomy was carried out in six patients with the disease because their pulmonary nodules persisted after repeated courses of chemotherapy over a 4 month period. They were followed up for a long period. These observations suggest that the most important role of thoracotomy is for diagnostic purposes; it is used for therapeutic purposes only when the pulmonary lesion is the only remaining source of increased HCG excretion. Much attention must be paid to tertiary hematogeneous metastases caused by resection of pulmonary foci that are unresponsive to chemotherapy.