Fully automated methods for the determination of hydrochlorothiazide in human plasma and urine

LC assays utilizing fully automated sample preparation procedures on Zymark PyTechnology™ Robot and BenchMate™ Workstation for the quantification of hydrochlorothiazide (HCTZ) in human plasma and urine have been developed. After aliquoting plasma and urine samples, and adding internal standard (IS) manually, the robot executed buffer and organic solvent addition, liquid—liquid extraction, solvent evaporation and on-line LC injection steps for plasma samples, whereas, BenchMate™ performed buffer and organic solvent addition, liquid—liquid and solid-phase extractions, and on-line LC injection steps for urine samples. Chromatographic separations were carried out on Beckman Octyl Ultrasphere column using the mobile phase composed of 12% (v/v) acetonitrile and 88% of either an ion-pairing reagent (plasma) or 0.1% trifluoroacetic acid (urine). The eluent from the column was monitored with UV detector (271 nm). Peak heights for HCTZ and IS were automatically processed using a PE-Nelson ACCESS*CHROM laboratory automation system. The assays have been validated in the concentration range of 2–100 ng ml⁻¹ in plasma and 0. 1–20 μg ml⁻¹ in urine. Both plasma and urine assays have the sensitivity and specificity necessary to determine plasma and urine concentrations of HCTZ from low dose (6.25/12.5 mg) administration of HCTZ to human subjects in the presence or absence of losartan.     

Fermentable simple sugars in self-administered medications as aetiologic agents in rampant caries. Case report

Sucrose is recognized as the most cariogenic carbohydrate. However, other simple sugars may act as substrates for cariogenic micro-organisms. The clinical features of an adult patient suffering from rampant caries are described. It was considered that the predominant aetiologic factor was prolonged intake of mineral supplements which contained the disaccharide lactose. The principal caries risk factors included drug-induced xerostomia and prolonged contact time with the substrate in a retentive form. Chemical analysis of the products revealed the presence of lactose together with starch or glucose. A strategy for preventive management prior to restorative therapy based on chlorhexidine and fluoride-containing products was followed. Possible contributions of sugar-containing medications to caries risk status are discussed.     

Expression of beta-catenin, COX-2 and iNOS in colorectal cancer: relevance of COX-2 adn iNOS inhibitors for treatment in Malaysia

BACKGROUND: Promising new pharmacological agents and gene therapy targeting cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) could modulate treatment of colorectal cancer in the future. The aim of this study was to elucidate the expression fo beta-catenin and teh presence of COX-2 and iNOS in colorectal cancer specimens in Malaysia. This is a useful prelude to future studies investigating interventions directed towards COX-2 adn iNOS. METHODS: A cross-section study using retrospective data over a 2-year period (1999-2000) involved 101 archival, formalin-fixed, paraffin-embedded tissue samples of colorectal cancers that were surgically resected in a tertiary referral. RESULTS: COX-2 production was detected in adjacent normal tissue in 34 sample (33.7%) and in tumour tissue in 60 samples (59.4%). More tumours expressed iNOS (82/101, 81.2%) than COX-2. No iNOS expression was detected in adjacent normal tissue. Intense beta-catenin immunoreactivity at the cell-to-cell border. Poorly differentiated tumours had significantly lower total beta-catenin (p = 0.009) and COX-2 scores (p = 0.031). No significant relationships were established between pathological stage and beta-catenin, COX-2 and iNOS scores. CONCLUSIONS: the accumulation of beta-catenin does not seem to be sufficient to activate pathways that lead to increased COX-2 and iNOS expression. A high proportion of colorectal cancers were found to express COX-2 and a significant number produced iNOS, suggesting that their inhibitors may be potentially useful as chemotherapeutic agents in the management of colorectal cancer.     

Combined spinal-epidural technique for vaginal hysterectomy in a patient with Machado-Joseph disease

OBJECTIVE: Machado-Joseph disease is a form of progressive spino-cerebellar ataxia with both bulbar and peripheral neurological manifestations. To date, anesthesia for patients affected by this disease has not been described. General anesthesia may be problematic because of the risk of pulmonary aspiration and hypoxia. We describe our experience with the successful use of combined spinal-epidural in a patient with Machado-Joseph Disease (MJD).CASE REPORT: A 38-year-old woman with MJD complicated by significant bulbar and peripheral neuropathy presented for an elective vaginal hysterectomy. She had no other medical history of note. After informed consent, subarachnoid block was performed by combined spinal-epidural anesthesia at the L2-3 lumbar intervertebral space with hyperbaric bupivacaine 12 mg, morphine 100 microg, and fentanyl 10 microg. Surgery proceeded uneventfully, with excellent postoperative analgesia. There was full recovery of preinduction neurologic function by the sixth postoperative hour. CONCLUSIONS: Central neuraxial anesthesia is an option for patients with MJD presenting for lower abdominal and lower extremity operations. Combined spinal-epidural anesthesia confers hemodynamic stability yet allows for augmentation of intraoperative anesthesia and postoperative analgesia.     

Intentional cryopreservation of epididymal spermatozoa from percutaneous aspiration for dissociated intracytoplasmic sperm injection cycles

BACKGROUND: To investigate the possibility of cryopreservation of spermatozoa obtained from percutaneous epididymal sperm aspiration (PESA) in patients with obstructive azoospermia and the feasibility of intentional dissociation of PESA and intracytoplasmic sperm injection (ICSI) cycles. METHODS: Fifty-six patients with obstructive azoospermia underwent diagnostic PESA before ovarian stimulation. If spermatozoa were found, they were frozen for subsequent ICSI. The outcome was compared with 17 fresh PESA/ICSI cycles. RESULTS: Among the 56 patients, diagnostic PESA obtained spermatozoa in 51 patients. The mean motility of the spermatozoa decreased from 15.2% to 4.2% after freezing and thawing. These patients underwent 96 frozen PESA/ICSI cycles. The rates of fertilization, implantation and clinical pregnancy for frozen-thawed spermatozoa (71.6, 14.0 and 40.6%, respectively) were similar to those for fresh spermatozoa (69.2, 13.2 and 41.2%, respectively). CONCLUSIONS: Sufficient numbers of spermatozoa can be obtained for cryopreservation through PESA and the spermatozoa work well after thawing. The strategy of performing diagnostic PESA before ovarian stimulation and freezing the recovered spermatozoa for subsequent ICSI is feasible for patients with obstructive azoospermia.     

Prenatal three-dimensional ultrasound diagnosis of a camptomelic dysplasia

We describe a case of camptomelic dysplasia identified prenatally with the assistance of three-dimensional ultrasonography. The typical skeletal dysplasia of camptomelic dysplasia – including anterior bowing of the tibia, with skin dimpling over a convex surface at the point of maximal deformity, and talipes equinovarus – was successfully identified using the techniques of surface-rendering, multiplanar displays and rotated volume data. Three-dimensional ultrasonography allows the diagnosis of camptomelic dysplasia , which has a poor prognosis as it is accompanied by respiratory insufficiency and spinal deformities, to be made without delay and provides information supplementary to that provided by two-dimensional ultrasound for early diagnosis of skeletal dysplasia.     

Spatial and temporal coordination of expression of immune response genes during Pseudomonas infection of horseshoe crab, Carcinoscorpius rotundicauda

Knowledge on how genes are turned on/off during infection and immunity is lacking. Here, we report the co-regulation of diverse clusters of functionally related immune response genes in a horseshoe crab, Carcinoscorpius rotundicauda. Expressed sequence tag (EST) clusters for frontline immune defense, cell signalling, apoptosis and stress response genes were expressed or repressed spatio-temporally during the acute phase of Pseudomonas infection. An infection time course monitored by virtual Northern evaluation indicates upregulation of genes in blood cells (amebocytes) at 3-h postinfection, whereas most of the hepatopancreas genes remained down regulated over 72 h of infection. Thus, the two tissues orchestrate a coordinated and timely response to infection. The hepatopancreas probably immuno-modulates the expression of other genes and serves as a reservoir for later response, if/when chronic infection ensues. On the other hand, being the first to encounter pathogens, we reasoned that amebocytes would respond acutely to infection. Besides acute transactivation of the immune genes, the amebocytes maintained morphological integrity, indicating their ability to synthesise and store/secrete the immune proteins and effectors to sustain the frontline innate immune defense, while simultaneously elicit complement-mediated phagocytosis of the invading pathogen. Our results show that the immune response against Pseudomonas infection is spatially and temporally coordinated.     

Biocompatibility of human osteosarcoma cells to root end filling materials

Ideal root end filling materials should have good physical and chemical properties, and the most important is that the material should be biocompatible with periradicular tissue. The biocompatibility of three root end filling materials, mineral trioxide aggregate, calcium hydroxide-based cement, and eugenol-based cement, were investigated in vitro by culturing extracts of these materials with human osteogenic sarcoma cells (U2OS). Extracts of each of the materials were made after incubation of the materials for 1 day and 1 week with complete McCoy's medium. The extracts were serially diluted and then incubated with U2OS cells for 24 and 48 h. Cell survival rates were assessed by means of a viability assay for mitochondrial dehydrogenase activity. Differences in mean cell survival rates were statistically assessed using one-way ANOVA. Results showed that the survival rates of U2OS cells were largest with mineral trioxide aggregate, followed by calcium hydroxide-based cement and eugenol-based cement at 24- and 48-h exposures using the 1-day and 1-week extracts. The duration of root end filling material extraction time and treatment time showed variable influence on the survival rates. The results suggest that mineral trioxide aggregate is more biocompatible than the other root end filling materials and is suitable for use in the clinical setting.