The cyclophilins (Cyps) are family members of proteins that exhibit peptidylprolyl cis-trans isomerase (PPIase, EC 5.2.1.8) activity and bind the immunosuppressive agent cyclosprin A (CsA) in varying degrees. During the process of random sequencing of a cDNA library made from Giardia intestinalis WB strain, the cyclophilin gene (gicyp 1) was isolated. An open reading frame of gicyp 1 gene was 576 nucleotides, which corresponded to a translation product of 176 amino acids (Gicyp 1). The identity with other Cyps was about 58-71%. The 13 residues that constituted the CsA binding site of human cyclophilin were also detected in the amino acid sequence of Gicyp 1, including tryptophan residue essential for the drug binding. The single copy of the gicyp 1 gene was detected in the G. intestinalis chromosome by southern hybridization analysis. Recombinant Gicyp 1 protein clearly accelerated the rate of cis-- Amino Acid Sequence ; Animals ; Cloning, Molecular ; Cyclophilins/antagonists & inhibitors/chemistry/genetics/*isolation&purification ; Cyclosporine/metabolism/pharmacology ; Giardia lamblia/*chemistry/genetics ; Human ; Immunosuppressive Agents ; Molecular Sequence Data ; Protein Binding ; Protozoan Proteins ; Recombinant Proteins

In order to evaluate the possible roles of secretory proteases in the pathogenesis of amoebic keratitis, we purified and characterized a serine protease secreted by Acanthamoeba lugdunensis KA/E2, isolated from a Korean keratitis patient. The ammonium sulfate-precipitated culture supernatant of the isolate was purified by sequential chromatography on CM-Sepharose, Sephacryl S-200, and mono Q-anion exchange column. The purified 33 kDa protease had a pH optimum of 8.5 and a temperature optimum of 55 degrees C. Phenylmethylsulfonylfluoride and 4- (2- Aminoethyl) -benzenesulfonyl-fluoride, both serine protease specific inhibitors, inhibited almost completely the activity of the 33 kDa protease whereas other classes of inhibitors did not affect its activity. The 33 kDa enzyme degraded various extracellular matrix proteins and serum proteins. Our results strongly suggest that the 33 kDa serine protease secreted from this keratopathogenic Acanthamoeba play important roles in the pathogenesis of amoebic keratitis, such as in corneal tissue invasion, immune evasion and nutrient uptake.
Acanthamoeba/*enzymology/isolation & purification/pathogenicity ; Acanthamoeba Keratitis/*parasitology ; Animals ; Cornea/parasitology ; Humans ; Hydrogen-Ion Concentration ; Korea ; Serine Endopeptidases/chemistry/*isolation & purification/*metabolism ; Substrate Specificity ; Temperature ; Virulence Factors

We investigated the sero-prevalence of toxocariasis among healthy Korean adults in 1999. A total of 314 sera from normal inhabitants in Whachon-gun, Gangwondo, Korea was examined for specific antibody levels against excretory-secretory products of second stage larvae of Toxocara (TES). The presence of cross-reactions with other helminthiases such as cysticercosis, paragonimiasis, sparganosis or clonorchiasis was also checked by specific IgG ELISA. Sera showing positive reaction against TES were also tested by IgG immunoblot and by IgE ELISA. Out of 314 subjects, 16 was found to be positive by TES IgG ELISA and immunoblot, among whom 12 were also positive by TES IgE ELISA. Among the 16 seropositive samples, two sera showed positive reaction against Paragonimus and sparganum antigen, respectively. These results inferred that cross-reactions were negligible between toxocariasis and other helminthiases. Toxocariasis seroprevalence among Korean rural adults was detected to be approximately 5%.
Adolescent ; Aged ; Aged, 80 and over ; Animals ; Antibodies, Helminth/blood ; Cross Reactions ; Enzyme-Linked Immunosorbent Assay ; Female ; Human ; Korea/epidemiology ; Larva Migrans, Visceral/*epidemiology/immunology ; Male ; Middle Aged ; Seroepidemiologic Studies ; Support, Non-U.S. Gov't ; Toxocara/immunology

To investigate the risk factors and clinical characteristics of postrenal transplant diabetes mellitus (PTDM), we reviewed the records of 177 renal allograft recipients in Maryknoll Hospiatal whose allografts had functioned longer than 6 months. Nineteen patients (10.7%) developed PTDM at 5.0+/-7.8 (1-52) months; 9 (47%) of these within 1 month. PTDM patients were older than nondiabetic renal transplants (42+/-2 vs 37+/-1 years, P<0.05). Body mass index tended to be higher in PTDM (23.5+/-1.0 vs 21.8+/-0.3kg/m2, P=0.09). Number of acute rejections (0.6+/-0.2 vs 0.5+/-0.1) and serum creatinine at 1 year after transplantation (1.2+/-0.8 vs 1.3+/-0.3mg/dL) were not different. Fasting (103.6+/-10.4 vs 84.4+/-1.6mg/dL, P<0.05) and postprandial (189.2+/-24.8 vs 118.6+/-2.3 mg/dL, P<0.01) blood sugars, measured before transplantation, were higher in PTDM. CsA blood level at 1 month posttransplantation was higher in PTDM (350+/-34 vs 279+/-8ng/mL, P<0.05). Fasting serum insulin was significantly higher (28.2+/-12.2 vs 7.3+/-2.0 microunit/dL, P<0.05) and serum C-peptide tended to be higher in PTDM patients compared with euglycemic renal recipients (6.3+/-1.6 vs 3.8+/-0.9ng/dL, P=0.08). All the PTDM patients were treated by either insulin or oral agent; 15 of 19 required no treatment after 4.7+/-6.9 months. In conclusion, prevalence of PTDM was 10.7%. PTDM patients were older. Body mass index was tended to be higher. Fasting and postprandial blood sugars, measured before transplantation, were higher in PTDM. Faslting serum insulin was higher and C-peptide tended to be higher in diabetics. These results suggested that increased insulin resistance plays a major role in the pathogenesis of PTDM.
Allografts ; Blood Glucose ; Body Mass Index ; C-Peptide ; Creatinine ; Cyclosporine ; Diabetes Mellitus* ; Fasting ; Humans ; Insulin ; Insulin Resistance ; Prevalence ; Risk Factors*

The taxonomy of Acanthamoeba spp., an amphizoic amoeba which causes granulomatous amoebic encephalitis and chronic amoebic keratitis, has been revised many times. The taxonomic validity of some species has yet to be assessed. In this paper, we analyzed the morphological characteristics, nuclear 18s rDNA and mitochondrial 16s rDNA sequences and the Mt DNA RFLP of the type strains of four Acanthamoeba species, which had been previously designated as A. divionensis, A. parasidionensis, A. mauritaniensis, and A. rhysodes. The four isolates revealed characteristic group II morphology. They exhibited 18S rDNA sequence differences of 0.2-1.1% with each other, but more than 2% difference from the other compared reference strains. Four isolates formed a different clade from that of A. castellanii Castellani and the other strains in morphological group II on the phylogenetic tree. In light of these results, A. paradivionensis, A. divionensis, and A. mauritaniensis should be regarded as synonyms for A. rhysodes.
Acanthamoeba/*classification/*genetics ; Animals ; DNA, Mitochondrial/genetics ; DNA, Ribosomal/genetics ; Phylogeny ; Polymorphism, Restriction Fragment Length ; RNA, Ribosomal, 18S/genetics

PURPOSE: The present study investigated the psychosocial health of disease-free breast cancer survivors who receive health examinations compared to matched non-cancer controls in a community setting. MATERIALS AND METHODS: We used baseline data from the Health Examinee cohort, which is composed of subjects participating in health. The disease-free breast cancer survivors were defined as those who were ≥ 2 years from initial diagnosis of breast cancer who had completed treatment. Females without a history of cancer were randomly selected at 1:4 ratio by 5-year age groups, education, and household income as a comparison group. We analyzed results from the Psychosocial Well-being Index-Short Form (PWI-SF) as a psychosocial health measurement. RESULTS: A total of 347 survivors of breast cancer and 1,388 matched controls were included. Total scores on the PWI-SF were lower in breast cancer survivors than matched non-cancer controls (p=0.006), suggesting a lower level of psychosocial stress in breast cancer survivors. In comparison to the control group, prevalence of drinking, smoking and obesity were lower, while exercising for ≥ 150 min/wk was higher in breast cancer survivors (p < 0.05). These findings suggest that breast cancer survivors have better health behaviors than their noncancer controls. After adjusting for other sociodemographic variables, breast cancer survivors were 36% less likely to be included in the stress group (odds ratio, 0.64; 95% confidence interval, 0.42 to 0.98). CONCLUSION: The disease-free breast cancer survivors resuming daily life demonstrated better psychosocial health status compared to matched non-cancer controls.
Breast Neoplasms* ; Breast* ; Cohort Studies ; Diagnosis ; Drinking ; Education ; Family Characteristics ; Female ; Health Behavior ; Humans ; Obesity ; Prevalence ; Smoke ; Smoking ; Survivors*

PURPOSE: This study was performed to examine the histopathologic changes of muscles and the expression patterns of ubiquitin and N-CAM (neural cell adhesion molecule) in accordance with cerebral palsy patient's spasticity. MATERIALS AND METHODS: We studied thirteen specimens from seven patients with spastic cerebral palsy, five patients suspected to have neuromuscular diseases, and one normal person. We performed the routine histologic procedures, the reverse transcriptional polymerase chain reaction (RT-PCR), and immunostaining. RESULTS: There were no disease-specific abnormalities related with the degree of spasticity on histopathologic evaluation. However, in the cerebral palsy patients, the degree of spasticity seems to have positive correlations with the expression of ubiquitin gene and negative correlations with the expression of N-CAM gene. On the other hand, in the immunostaining procedures, the reactions to ubiquitin protein were all negative and reactions to N-CAM protein were strongly positive only in two hereditary motor sensory neuropathy patients. CONCLUSION: The results of our study seem to be caused by multiple mechanisms. If more studies about the changes after the transcription of ubiquitin and N-CAM genes are performed, these results can be applied to the research and treatment of cerebral palsy on molecular biologic aspects.
Cell Adhesion ; Cerebral Palsy* ; Hand ; Humans ; Muscle Spasticity* ; Muscles* ; Neural Cell Adhesion Molecules* ; Neuromuscular Diseases ; Polymerase Chain Reaction ; Ubiquitin*

PURPOSE: Using proteomic analysis, this study was performed to see the characteristics of proteins expression in the muscles of spastic cerebral palsy patients. MATERIALS AND METHODS: We studied twelve specimens from six patients with spastic cerebral palsy, three patients with myelomeningocele, and three normal people who underwent orthopaedic surgeries due to trauma. We studied the extracted proteins showing differences in the two-dimensional electrophoresis, and the prominent thirteen proteins were re-evaluated by proteomics and the reverse transcriptional polymerase chain reaction, which was to clarify the relationship between gene and protein expression. RESULTS: Among fifteen proteins, six proteins were found to be higher in normal people, and nine were found to be higher in the groups of patients by spot histogram. The results of proteomic analysis with MALDI-TOF for fifteen proteins showed that the expression of DJ-1 was related to cerebral palsy. CONCLUSION: This study shows that strong expression of DJ-1 is related to spasticity and cerebral palsy. We showed for the first time the possibility of any relationship between spastic condition and DJ-1 expression.
Cerebral Palsy ; Electrophoresis ; Humans ; Meningomyelocele ; Muscle Spasticity ; Muscles ; Polymerase Chain Reaction ; Proteins ; Proteomics

We have cloned a cDNA encoding a cysteine proteinase of the Acanthamoeba healyi OC-3A strain isolated from the brain of a granulomatous amoebic encephalitis patient. A DNA probe for an A. healyi cDNA library screening was amplified by PCR using degenerate oligonucleotide primers designed on the basis of conserved amino acids franking the active sites of cysteine and asparagine residues that are conserved in the eukaryotic cysteine proteinases. Cysteine proteinase gene of A. healyi (AhCP1) was composed of 330 amino acids with signal sequence, a proposed pro-domain and a predicted active site made up of the catalytic residues, Cys(25), His(159), and Asn(175). Deduced amino acid sequence analysis indicated that AhCP1 belongs to ERFNIN subfamily of C1 peptidases. By Northern blot analysis, no direct correlation was observed between AhCP1 mRNA expression and virulence of Acanthamoeba, but the gene was expressed at higher level in amoebae isolated from soil than those from clinical samples. These findings raise the possibility that Ahcp1 protein may play a role in protein metabolism and digestion of phagocytosed bacteria or host tissue debris rather than in invasion of amoebae into host tissue.
Acanthamoeba/*enzymology/genetics/pathogenicity ; Amebiasis/parasitology ; Amino Acid Sequence ; Animals ; Base Sequence ; Cathepsins/*genetics ; DNA, Protozoan/chemistry/genetics/*isolation & purification ; Encephalitis/parasitology ; Gene Expression ; Genes, Protozoan ; Humans ; Molecular Sequence Data ; Polymerase Chain Reaction ; Protozoan Proteins/chemistry/genetics/physiology ; Sequence Alignment ; Virulence

PURPOSE: Pathologic changes in the growth plate remain unknown in Legg-Calve-Perthes (LCP) disease. Spontaneously hypertensive rats have proven to be a good model for studying LCP disease. This study investigated the histopathologic changes and the expression of vascular endothelial growth factor in the growth plate of spontaneously hypertensive rats (SHR). MATERIALS AND METHODS: Sixty SHR rats were divided into two groups: those showing osteonecrosis (SHR+n group: 32), and those showing normal ossification (SHR-n group: 28). Thirty Wister Kyoto rats served as a control. For histomorphological measurement, the length of each zone of the growth plate was measured. Cell kinetics was measured by 5-bromo-2'-deoxyuridin (BrdU) immunohistochemistry and transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL) assays. Vascular endothelial growth factor (VEGF) immunohistochemistry was used to identify of expression of VEGF. RESULTS: The lengths of growth plates of the SHR+n group were significantly shorter in the initial growth period than those of the other groups. The lowest proliferative rate and the highest apoptosis rate were observed in the SHR+n group at the initial growth period. The expression of VEGF in the growth plate of the SHR group was lower than the control group, and it was lower in the SHR+n group than in the SHR-n group. CONCLUSION: The growth plate of the SHR+n group was found to be affected by disease process of ischemic necrosis of the femoral head, and this might explain the relative overgrowth of the greater trochanter in the later stages of LCP disease.
Animals ; Apoptosis ; Femur Head/metabolism/*pathology ; Femur Head Necrosis/metabolism/pathology ; Growth Plate/*cytology/metabolism ; Osteogenesis/physiology ; Rats ; Rats, Inbred SHR ; Rats, Sprague-Dawley ; Vascular Endothelial Growth Factor A/metabolism