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PubMed1991

Direct polymerase chain reaction test for detection of Helicobacter pylori in humans and animals.

We designed a polymerase chain reaction (PCR) for amplifying the Helicobacter pylori gene encoding 16S rRNA. Primers for the specific detection of H. pylori were designed for areas of the 16S rRNA gene in which there is the least sequence homology between H. pylori and its closest relatives. The specificity of detection was confirmed by ensuring that the primers did not amplify DNA extracts from the campylobacters H. cinaedi, H. mustelae, and Wolinella succinogenes, which are the closest relatives of H. pylori, as determined by 16S rRNA sequencing. Serial dilution experiments revealed the detection of as little as 0.1 pg of DNA by PCR and 0.01 pg by nested PCR. H. pylori DNA was detected successfully in clinical paraffin-embedded and fresh gastric biopsy specimens from patients positive for the bacterium and also in fecal suspensions seeded with the organism. The DNA from the nonculturable coccoid form of H. pylori was also identified by the primers. Universal primers designed for highly conserved areas on the 16S rRNA gene enabled large amplification products to be produced for direct sequencing analysis. Gastric bacteria resembling H. pylori have been isolated from animals. DNA of these animal gastric bacteria amplified with H. pylori-specific primers yielded PCR products identical to those from human isolates of H. pylori, as confirmed by the use of a 20-base radiolabelled probe complementary to an internal sequence flanked by the H. pylori-specific primers. The results of PCR amplification and partial 16S rRNA gene sequence analysis strongly support the contention that the gastric organisms previously recovered from a pig, a baboon, and rhesus monkeys are H. pylori.

Animals↗
PubMed1990

Evaluation of occupational asthma from airborne egg protein exposure in multiple settings.

We previously reported IgE-mediated occupational asthma among workers exposed to airborne egg protein at a plant that produces liquid and dried powdered egg products. To demonstrate that our original observations are generalizable to other facilities that process eggs, and to estimate the prevalence of IgE-mediated occupational asthma among egg-exposed workers, we conducted surveys at two additional plants. We administered a questionnaire to 188 employees to identify workers with symptoms suggestive of occupational asthma. We further evaluated 88 workers with and without symptoms by a clinical examination by a physician blinded to results of other tests, serial peak expiratory flow rate (PEFR) determinations every three hours while awake for one week, and skin prick tests and serum specific IgE levels to extracts of factory egg products, commercial egg test reagents, and egg white protein fractions. Fourteen workers had work-related asthmalike symptoms by questionnaire, a physician diagnosis of occupational asthma, and evidence of IgE-mediated sensitization to one or more egg proteins. Workers exposed exclusively to liquid egg aerosol, as well as workers exposed primarily to dried airborne egg protein, developed occupational asthma. This study replicated our original observations and demonstrated that workers in all areas of liquid and powdered egg production are at risk of developing occupational asthma from exposure to airborne egg proteins.

Air Pollutants, Occupational↗
PubMed2001

Expression and immunolocalization of AQP6 in intercalated cells of the rat kidney collecting duct.

The expression and localization of AQP6 were examined in rat kidneys. In the kidney compartments, the expression was more intense in the outer medulla than in the cortex or inner medulla, and was negative in the glomerulus. During development, the AQP6 mRNA expression in the kidney was not detected in the fetus, but was recognized at birth, increased gradually by 4 weeks of age, and was unchanged thereafter. In situ hybridization demonstrated significant signals for AQP6 mRNA along the outer and inner medullary collecting ducts. Since the localization of the AQP6 mRNA-expressing cells was comparable to that of immunoreactive H+ ATPase-bearing cells in the collecting duct, they were identified as intercalated cells. No AQP6 mRNA signals were recognizable in other cells in the kidneys, including glomerular cells. No glomerular expression of AQP6 mRNA was confirmed by RT-PCR using total RNA extracted from the glomeruli. Immunohistochemistry using an antibody raised against recombinant rat AQP6 protein could localize the immunoreactivity in a population of collecting duct cells. Serial section observations indicated that the AQP6-immunoreactive cells corresponded to H+ ATPase bearing intercalated cells.

Animals↗
PubMed2000

Quantification of enterovirus RNA in sludge samples using single tube real-time RT-PCR.

We have developed a quantitative RT-PCR method that can be used to determine the amount of enterovirus RNA in urban sludge samples. This method combines Taq-Man technology with the ABI Prism 7700 real-time sequence detection system. We optimized a one-step RT-PCR that uses a dual-labeled fluorogenic probe to quantify the 5' noncoding region of enteroviruses. For accurate quantification of the number of copies, a Mahoney type 1 poliovirus RNA standard was designed and produced using genetic engineering. This fragment, quantified using the Ribogreen method, was used in serial dilutions as an external standard. The method had a 7-log dynamic range (5 to 2 x 10(7)). PCR inhibitors were removed by extracting viral RNA (after virus concentration) using the RNeasy mini kit with added polyvinylpyrrolidone (PVP) and running the amplification reaction with a mixture containing PVP and T4 gene 32 protein. This real-time quantification of enterovirus RNA allows large numbers of samples to be screened. Its sensitivity, simplicity and reproducibility render it suitable as a screening method with which to characterize enteroviruses, the presence of infectious particles being subsequently confirmed by cell culture.

Enterovirus↗
PubMed1975

Myocardial uptake of ouabain in intact dog and man.

The time corse of ouabain extraction from the coronary circulation was determined after single i.v. doses (3-H-labeled in the dogs). Plasma ouabain concentration ([O]) was measured in serial, paried arterial (A) and coronary sinus (CS) blood samples by radioimmunoassay in the patients and by 3-H-counting in the dogs. In both species, the difference in [O] between A and CS (A - CS [O]) narrowed rapidly during the initial 1 to 3 minutes and then at a slower rate in an exponential fashion the next 4 to 12 minutes, with a mean half-life of 2.6 plus or minus 0.4 (S.D.) minutes in the patient (n equals 6) and 1.4 plus or minus 0.5 minutes in the canine (n equals 10) studies. In four dogs, the A - CS concentration difference of sucrose (an extracellular fluid marker) was gone within 60 seconds after an i.v. bolus. In the canine studies, the exponential phase of ouabain removal from the coronary circulation accounted for 49 plus or minus 8% of that directly measured in left ventricular samples obtained shortly after ouabain uptake was complete (17.0 plus or minus 4.5 minutes after the i.v. dose). Left ventricular [O] in these samples was 11.6-fold higher than in simultaneously obtained plasma. Thus, the initial rapid efflux of ouabain, like sucrose, from the coronary circulation would appear toreflect movement into interstitial fluid, whereas the subsequent exponential removal of ouabain, occurring after equilibrium between plasma and intersitital fluid, reflects binding to myocardial cells.

Angina Pectoris↗
PubMed2002

Prenatal diagnosis of alpha-thalassemia-1 (SEA type) by chorionic villus sampling.

OBJECTIVE: To describe the experience of prenatal diagnosis for Hb Bart's disease, by chorionic villus sampling (CVS) with DNA analysis. DESIGN: Descriptive study SETTINGS: Department of Obstetrics and Gynecology, Faculty of Medicine, Chiang Mai University. SUBJECTS: Sixteen high risk pregnancies at risk of Hb Bart's disease who were eligible for CVS criteria between 1 January, 1999 and May 31, 2000. MATERIAL AND METHOD: Fetal villi were obtained by either transcervical (TC) or transabdominal (TA) CVS route to extract DNA and detect for alpha-thal-1 gene deletion (SEA type) with modified Chang's method. The CVS results were confirmed by either serial ultrasound or cordocentesis or diagnosis after pregnancy termination. MAIN OUTCOME MEASURES: The efficacy, safety and pregnancy outcomes. RESULTS: CVS was successfully done in all of 16 cases (5 with TC and 11 with TA), The mean gestational age was 13.25 +/- 2.9 weeks. The procedure time for TA was shorter than that of TC (4.64 +/- 5.4 vs 10.4 +/- 11.3 min). The CVS result showed as follows: 3 normal fetuses, 7 alpha-thal-1 carriers, 4 fetal Hb Bart's, 1 misdiagnosis and 1 failure to diagnosis due to technical error. The sensitivity and specificity were 100 per cent (4/4) and 90.91 per cent (10/11), respectively. One case of Hb Bart's misdiagnosis and one failure case were later confirmed for alpha-thal-1 trait and alpha-thal-1/ Hb E trait by cordocentesis, respectively. The pregnancy outcomes included 11 livebirths, 4 terminated cases and 1 fetal loss of continuing pregnancies. No serious complications occurred. CONCLUSION: This preliminary experience suggests that CVS is an effective method for early prenatal diagnosis of fetal Hb Bart's.

Chorionic Villi Sampling↗
PubMed1986

[Humoral immune responses to stages of Brugia pahangi in experimentally infected rats].

In the present experiment, Sprague-Dawley rats were inoculated with Brugia pahangi, a lymphatic dwelling filarial parasite. Each rat received 100 third stage larvae (L3) of B. pahangi freshly harvested from Aedes togoi. Rats in the control group were injected with saline. After inoculation, tail vein blood was collected from each rat at 2-week intervals. The immune responses of the infected rats to B. pahangi were measured by the enzyme-linked immunosorbent assay (ELISA). Four types of antigen were prepared from different stages of B. pahangi. The antigen extracts were prepared from microfilaria, L3, and male and female adult worms by homogenization, eight times of freezing and thawing, and centrifugation at 11,000 X g for 45 min at 4 degrees C. Aliquots of supernatant extracts were stored at -20 degrees C. These antigens were adjusted to the same concentration of their protein contents and employed in the ELISA test. Serial serum dilutions were made to determine the concentrations of stage-specific antibodies in each serum sample collected during the infection course. Microfilaria specific antibodies appeared in the blood of infected rats at the 6th week of infection. Peak titer was observed at the 10th week. Subsequently, a gradual decrease was seen in the following month. Serum antibody titers against L3 and adult antigens rised since the 4th week and reached to the peak at 14th week. The exist of common antigens shared by the mature adults and infective larvae were detected by the ELISA and conformed by disc gel electrophoresis.

Animals↗
PubMedDate not supplied

Investigations on the presence of papova virus in certain forms of human cancer. Note 2. Brain tumors.

Two serially transmissible tumors (metastatic lymphosarcomas) were induced in the hamster by inoculation of human brain tumor material--tissue homogenate from a malignized fibroma or DNA extracted from a meningioma. SV40 V antigen was made evident by indirect immunofluorescence in 4 out of 18 human tumors, while SV40 T antigen could only be demonstrated in the tumors experimentally induced in hamsters. Antibodies to SV40 T antigen were detected in 12 out of 60 patients with different tumors, in 1 out of 40 blood donors and in 7 out of 18 hamsters carrying experimentally induced tumors.

Adult↗
PubMed1981

Pharmacokinetics of dyphylline elimination by uremic patients.

The elimination kinetics of dyphylline were investigated in four uremic patients receiving chronic hemodialysis treatment. Dyphylline (1000 mg) was administered to each patient orally 2 hr before hemodialysis. Serial arterial and venous blood samples as well as outflowing dialysate samples were collected and analyzed by high-pressure liquid chromatography for dyphylline. The extraction efficiency of the hollow-fiber dialyzers averaged 44.6%. The mean dialysis clearance was 108.7 ml/min which compares favorably to the 57 ml/min metabolic clearance of the drug. A mean of 278.6 mg or 28.34% of the administered dose was recovered in the dialysate during the 4-hr dialysis interval. Half-lives for dyphylline during dialysis ranged from 3.43 to 7.62 hr. The off-dialysis half-lives for two of the four patients studied on a separate occasion were 10.6 and 13.2 hr. The overall clearance calculation projects an average half-life reduction of approximately 65% during hemodialysis. We conclude that dyphylline is dialyzable. Hemodialysis patients receiving dyphylline treatment may require a dosage regimen alteration. In addition, hemodialysis may be useful for the treatment of dyphylline overdose because of its effectiveness in accelerating the removal of the drug.

Adult↗
PubMed1994

Evaluation of cerebral metabolism and quantitative electroencephalography after hypothermic circulatory arrest and low-flow cardiopulmonary bypass at different temperatures.

Although widely used for repair of complex cardiovascular pathologic conditions, long intervals of hypothermic circulatory arrest and low flow cardiopulmonary bypass may both result in cerebral injury. This study examines cerebral hemodynamics, metabolism, and electrical activity to evaluate the risks of cerebral injury after 60 minutes of hypothermic circulatory arrest at 8 degrees C, 13 degrees C, and 18 degrees C, compared with 60 minutes of low flow cardiopulmonary bypass at 18 degrees C. Thirty-two puppies were randomly assigned to one of four experimental groups and centrally cooled to the appropriate temperature. Serial evaluations of quantitative electroencephalography, radioactive microsphere determinations of cerebral blood flow, calculations of cerebral oxygen consumption, cerebral glucose consumption, cerebral vascular resistance, cerebral oxygen extraction, systemic oxygen metabolism, and systemic vascular resistance were done. Measurements were obtained at baseline (37 degrees C), at the end of cooling, at 30 degrees C during rewarming, and at 2, 4, and 8 hours after hypothermic circulatory arrest or low flow cardiopulmonary bypass. At the end of cooling, cerebral vascular resistance remained at baseline levels in all groups, but systemic vascular resistance was increased in all groups. Cerebral oxygen consumption became progressively lower as temperature was reduced: it was only 5% of baseline at 8 degrees C; 20% at 13 degrees C; and 34% and 39% at 18 degrees C. Quantitative electroencephalography was silent in the 8 degrees C and 13 degrees C groups, but significant slow wave activity was present at 18 degrees C. Systemic vascular resistance and cerebral oxygen consumption returned to baseline values in all groups by 2 hours after hypothermic circulatory arrest or low flow cardiopulmonary bypass, but cerebral vascular resistance remained elevated at 2 and 4 hours, not returning to baseline until 8 hours after hypothermic circulatory arrest or low flow cardiopulmonary bypass. All but two of the long-term survivors (27 of 32) appeared neurologically normal; after hypothermic circulatory arrest at 8 degrees and 18 degrees C two animals had an unsteady gait. Comparison of quantitative electroencephalography before operation and 6 days after operation showed a significant increase in slow wave activity (delta activity) after hypothermic circulatory arrest and low flow cardiopulmonary bypass at 18 degrees C, a change that suggests possible cerebral injury. Although undetected after operation by simple behavioral and neurologic assessment, significant differences in cerebral metabolism, vasomotor responses, and quantitative electroencephalography do exist during and after hypothermic circulatory arrest and low flow cardiopulmonary bypass at various temperatures and may be implicated in the occurrence of cerebral injury.(ABSTRACT TRUNCATED AT 400 WORDS)

Analysis of Variance↗
PubMedDate not supplied

Vacuum extraction delivery complicated with acute subdural hematoma and cerebral infarction: report of one case.

We present a term, 2950 gm female neonate who was delivered by vacuum extraction and subsequently developed convulsions 23 hours after birth. Acute subduralhematoma was rapidly diagnosed by computed tomographic brain scan and successfully removed. The patient had a smooth postoperative recovery but cerebral infarction was found during follow up imaging studies. Magnetic resonance angiography revealed poor visualization of bilateral posterior cerebral arteries. After a follow-up period of 4 months, the infant had normal neurological development except for mild microcephalus. We report the occurrence of cerebral infarction in an infant following evacuation of a subdural hematoma. Onset of the late complication suggests the importance of serial neuroradiological and clinical evaluations.

Cerebral Infarction↗
PubMed2003

Garlic extract and two diallyl sulphides inhibit methicillin-resistant Staphylococcus aureus infection in BALB/cA mice.

OBJECTIVES: The inhibitory effect of garlic extract, diallyl sulphide and diallyl disulphide against methicillin-resistant Staphylococcus aureus (MRSA) infection in BALB/cA mice was studied. The influence of these agents upon the levels of fibronectin, interleukin-6 and lipid oxidation in MRSA-infected mice was examined. METHODS: Garlic extract at 100% and 50%; diallyl sulphide (DAS) at 10% and 5%; diallyl disulphide (DADS) at 1% and 0.5% were used in this study. Sixteen clinical MRSA isolates obtained from infected patients were used in this study (n=16). Mice were infected by injecting 200 microL MRSA-PBS solution, which contained 10(7) cfu, via the tail vein. At 16 h post-infection (p.i.), garlic extract, DAS or DADS at 200 microL was administrated orally. At 24 h p.i., mice were killed and blood, liver, kidney and spleen of each mouse were collected. Plasma and the filtrate from each organ and serial dilutions were used to determine colony count. Plasma fibronectin level was determined by rabbit anti-rat fibronectin antibody and quantified by ELISA. Interleukin-6 levels were determined by commercial kit. Lipid oxidation was determined by measuring malondialdehyde levels. RESULTS: The oral administration of these agents significantly decreased the viability of MRSA, in plasma, liver, kidney and spleen (P<0.05). MRSA infection significantly increased fibronectin and interleukin-6 levels in plasma of MRSA-infected mice (P<0.05); however, the oral administration of garlic extract and two diallyl sulphides significantly reduced both fibronectin and interleukin-6 levels (P<0.05). MRSA infection also significantly enhanced lipid oxidation in plasma and three organs (P<0.05). The treatments of garlic extract and two diallyl sulphides significantly decreased the malondialdehyde level and showed antioxidant protection (P<0.05). CONCLUSIONS: These data strongly supported the conclusion that garlic extract, diallyl sulphide and diallyl disulphide possessed multiple protective functions against MRSA infection, in which diallyl sulphide and diallyl disulphide could be considered as novel therapeutic agents for the treatment of MRSA infection.

Allyl Compounds↗
PubMed2000

Capillary electrophoretic determination of sanguinarine and chelerythrine in plant extracts and pharmaceutical preparations.

Capillary electrophoresis was employed to determine the principal quaternary benzo[c]phenanthridine alkaloids, sanguinarine and chelerythrine, in two plant extracts and one oral hygiene product. Phosphate-Tris buffer of pH 2.5 was used as a background electrolyte, limits of detection were 3 micromol/l(-1) (sanguinarine) and 2.4 micromol,l(-1) (chelerythrine) using UV detection at 270 nm. The method, which correlated well with HPLC, is suitable for serial determination of sanguinarine and chelerythrine in plant products and pharmaceuticals.

Alkaloids↗
PubMed1984

Nuclear estrogen receptor and nonhistone chromosomal proteins in hormonal independency of murine breast cancers.

The capability of nuclear binding of cytosol estrogen receptors (ERc) was studied in GR mouse mammary tumors during their alteration of hormonal dependency through serial transplantations. Nuclei from GR mouse mammary tumors were incubated with uterine cytosol receptor complexes labeled with 125I-estradiol, and the amount of receptor binding in the 0.4 M KCl nuclear extracts was determined. The originally ERc-positive-hormone-dependent (type I) tumors were capable of nuclear receptor binding, while this function was markedly reduced in the evolved hormone-independent (type II) tumors, although the ERc content in the latter was still positive. The originally hormone-independent (ERc-negative, type III) tumors, however, retained the nuclear binding capability. It appears that the hormonal independency in type III tumors is due to a lack of ER, while in type II tumors it may be attributed to the loss of nuclear binding capability for receptor complexes. Nonhistone chromosomal proteins (NHCP) were analyzed by the 2-dimensional gel electrophoretic technique. A Mr 31,000 NHCP was present in 11 of 12 type I, and four of four type III tumors. Following serial transplantation of the type I tumors, this NHCP was either markedly diminished or not observed in all 14 type II tumors examined. Although it coincides with the capability of nuclear receptor binding, the biological function of this NHCP is still undefined and warrants further investigation.

Animals↗
PubMed1978

[Studies on the antimicrobial effect of natural and synthetic humic acids (author's transl)].

Preparations of humic acids extracted from different soils by various methods and model humus substances obtained synthetically by oxidation of hydroquinone and pyrocatechin are tested for growth inhibition of representative strains of human pathogenic microorganisms using a micro serial dilution technique. Within the concentration range of less than or equal to 2500 micrograms/ml 57 of 81 natural and also the two synthetic humic acids show antimicrobial activity with differing spectra. These substances inhibit St. epidermidis, St. aureus, Str. pyogenes, S. typhimurium, Prot. vulgaris, Ent. cloacae, Ps. aeruginosa and C. albicans, but not Str. faecalis and E. coli. The degree of activity or the sensitivity of test organisms, respectively, amounts to 2500--1250 micrograms/ml predominantly, partially 625--312 micrograms/ml and can reach values of up to 39 micrograms/ml with synthetic hydroquinone humic acid. The spectrum and degree of activity vary according to the origin and extraction mode of the natural humic acids. The in vitro evidence of efficiacy against human pathogenic microorganisms gives a rational basis of therapeutic use of substances of humic acid type in infectious conditions.

Anti-Bacterial Agents↗
PubMed1980

[Sensitization of lymphocyte to tumor extract detected by macrophage migration inhibition test(MIT) in patients with cervical cancer (author's transl)].

In vitro cellular hypersensitivity to the tumor-associated antigens (TAA) in 30 patients with cervical cancer (12 cases with stage 1b and 18 cases with stage 11) was studied by the method of macrophage migration inhibition test (MIT). Tumor soluble extracts as antigen were obtained by sonication or 3M-KCL method. The results of MIF production was presented as a migration index (MI) by the following formula. (Formula: see text), The result of MI were as follows: 79.25 +/- 21.69 in patients with stage 1b, 79.50 +/- 14.65 with stage 11. A positive MI case (MIF positive) which was less than 75, was found in 8 cases (66.7%) with stage 1b and 5 cases (27.8%) with stage 11. Relationship between the result of MI and skin reactions were as follows: 75.83 +/- 17.30 in PPD positive (18 cases), 84.75 +/- 16.96 in PPD negative (12 cases); 78.70 +/- 18.24 in DNCB positive (20 cases), 84.75 +/- 16.96 in DNCB negative (10 cases); 77.93 +/- 15.79 in PHA positive (27 cases) and 97.33 +/- 24.79 in PHA negative (3 cases). The serial assay of MI were performed in 10 cases and MIF production disappeared within 6 weeks after curative operation in all cases. From these findings, the cellular immune response to autologous tumor extracts was found in this assay system.

Antigens, Neoplasm↗
PubMed2002

Complete tooth extraction: a case report.

Advanced Trauma and Life Support protocol is used in trauma situations to identify life-threatening injuries, and after those are addressed, it provides a systematic approach to identify minor injuries. A 17-year-old male, who was involved in a motor vehicle accident, was treated for an open toe fracture. He also had an "avulsion injury" to his left hand and was missing a tooth. On follow-up two days later, he was found to have a tooth in his left hand after he presented with cellulitis of a hand wound. He was appropriately treated with foreign body removal, incision and drainage, and antibiotics. This case is not only presented for being unique but also to emphasize the importance of the secondary survey and serial examinations instructed by Advanced Trauma and Life Support.

Adolescent↗
PubMed1987

Propagation of differentiating normal human tracheobronchial epithelial cells in serum-free medium.

Serial-passage cultures of normal human tracheobronchial (TB) epithelial cells that exhibit functional differentiation have been established in serum-free medium supplemented with bovine pituitary extract (25 micrograms/ml), insulin (5 micrograms/ml), hydrocortisone (0.5 micrograms/ml), EGF (5 ng/ml), 10(-6)M each of ethanolamine and phosphoethanolamine, and antibiotics. The cells proliferated in this medium with a population doubling time of approximately 80 hours. Further, the passaged cultures retained differentiated morphology as evidenced by secretion of glycoproteins, binding of concanavalin A lectin, and presence of alcian blue and periodic acid Schiff-positive material in their cytoplasm. Ultrastructural observations further supported the functional epithelial nature of the cultures. Most cells exhibited characteristic microvilli on cell surfaces and showed junctional complexes between them. The cytoplasm contained a large number of perinuclear secretory vesicles, a characteristic feature of the differentiated cells. These cultures provide an excellent model to study factors that regulate synthesis and secretion of glycoproteins in normal human TB cells.

Adult↗

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