Peritoneal pH during laparoscopy is dependent on ambient gas environment: helium and nitrous oxide do not cause peritoneal acidosis.

BACKGROUND: Little is know about the effects of different insufflation gases on peritoneal pH during laparoscopy. However, these changes may influence the intracellular signalling system, resulting in altered cell growth or adhesiveness. The aim of this study was to determine the effects of carbon dioxide (CO(2)), nitrous oxide (N(2)O), and helium (He) on parietal and visceral peritoneal pH. The effect of different intraabdominal pressures on parietal and visceral peritoneal pH was also examined. METHODS: We conducted both an ambient gas study and a pressure study. For the ambient gas study, 20 pigs were divided into the following four groups: (a) CO(2), (b) He, (c) N(2)O, and (d) abdominal wall lift (Lift) laparoscopy. Parietal and visceral peritoneal pH were measured at 15 min intervals for 180 min. For the pressure study, 15 pigs were divided into the following three groups: (a) CO(2), (b) He, (c) N(2)O laparoscopy. Baseline values were established for parietal and visceral peritoneal pH. Intraabdominal pressure was then increased stepwise at 1-mmHg intervals to 15 mmHg. After pressure was maintained for 15 min at each setting, parietal and visceral peritoneal pH were measured. RESULTS: Ambient gas environment was the major determinant of parietal peritoneal pH. Carbon dioxide caused parietal peritoneal acidosis. Helium, N(2)O, and Lift caused alkalotic parietal peritoneal pH. Intraabdominal pressure had a minor effect on parietal peritoneal pH. At higher intraabdominal pressure (12-15 vs 5-8 mmHg), CO(2) caused a slight decrease in parietal peritoneal pH, whereas N(2)O and He caused a slight increase in parietal peritoneal pH. Visceral peritoneal pH remained relatively unaffected during all studies. CONCLUSIONS: Parietal peritoneal pH during laparoscopy was highly dependent on the ambient gas environment. The effect of intraabdominal pressure on parietal peritoneal pH was of minor significance. Carbon dioxide caused a slight worsening of parietal peritoneal acidosis at higher intraabdominal pressure, whereas, N(2)O, He, and Lift did not cause parietal peritoneal acidosis.

Optimizing bacterial expression of catalytically active human cytochromes P450: comparison of CYP2C8 and CYP2C9.

1. Methods for the co-expression in Escherichia coli of human cytochrome P450 (CYP) 2C8 and CYP2C9 with NADPH-cytochrome P450 reductase (OxR) to produce a catalytically active system were compared. 2. Approaches assessed were expression of a CYP:OxR fusion construct, bicistronic plasmids, simultaneous transformation with CYP and OxR plasmids, and separate expression of CYP and OxR with reconstitution of activity by mixing the bacterial membranes. Two N-terminal modifications (Delta3-20 and 17alpha-leader) of the individual P450s were additionally investigated. 3. Each approach gave efficient expression of CYP2C8 and CYP2C9, but the bicistronic constructs under the expression conditions used gave low OxR expression and low catalytic activity. CYP expression was higher with the Delta3-20 construct for CYP2C9 and with the 17alpha-presequence construct for CYP2C8. 4. Using torsemide as substrate, all methods gave catalytically active systems with K(m) values similar to human liver microsomes. Mixing bacterial membranes containing separately expressed CYP and OxR reconstituted a catalytically active system with the Delta3-20 construct for CYP2C9 but not for CYP2C8, and with neither of the 17alpha- presequence constructs. OxR co-expressed with CYP in the same membrane interacted with CYP to reconstitute activity more effectively than addition of exogenous OxR membranes. 5. Expression construct and OxR co-expression strategy should be individualized for CYP isoforms.

Carbon dioxide pneumoperitoneum causes severe peritoneal acidosis, unaltered by heating, humidification, or bicarbonate in a porcine model.

BACKGROUND: Carbon dioxide (CO(2)) is the most common gas used for insufflation in laparoscopy, but its effects on peritoneal physiology are poorly understood. This study looks at the changes in peritoneal and bowel serosal pH during CO(2) pneumoperitoneum, and whether heating and humidification with or without bicarbonate alters the outcomes. METHODS: Twenty-one pigs divided into four groups as follows: (1) standard (STD) laparoscopy (n = 5); (2) heated and humidified (HH) laparoscopy (n = 6); (3) heated and humidified with bicarbonate (HHBI) laparoscopy (n = 5); and (4) laparotomy (n = 5). Peritoneal pH, bowel serosal pH, and arterial blood gas (ABG) were obtained at 15-min intervals for 3 h. RESULTS: Severe peritoneal acidosis (pH range 6.59-6.74) was observed in all laparoscopy groups, and this was unaltered by heating and humidification or the addition of bicarbonate. Bowel serosal acidosis was observed in all laparoscopy groups with onset of pneumoperitoneum, but it recovered after 45 minutes. No significant changes in peritoneal or bowel serosal pH were observed in the laparotomy group. CONCLUSION: CO(2) pneumoperitoneum resulted in severe peritoneal acidosis that was unaltered by heating and humidification with or without bicarbonate. Alteration in peritoneal pH may conceivably be responsible for providing an environment favorable for tumor-cell implantation during laparoscopy.

Polymorphic hydroxylation of perhexiline in vitro.

AIMS: The aims of this study were to examine the in vitro enzyme kinetics and CYP isoform selectivity of perhexiline monohydroxylation using human liver microsomes. METHODS: Conversion of rac-perhexiline to monohydroxyperhexiline by human liver microsomes was assessed using a high-performance liquid chromatography assay with precolumn derivatization to measure the formation rate of the product. Isoform selective inhibitors were used to define the CYP isoform profile of perhexiline monohydroxylation. RESULTS: The rate of perhexiline monohydroxylation with microsomes from 20 livers varied 50-fold. The activity in 18 phenotypic perhexiline extensive metabolizer (PEM) livers varied about five-fold. The apparent Km was 3.3 +/- 1.5 micro m, the Vmax was 9.1 +/- 3.1 pmol min-1 mg-1 microsomal protein and the in vitro intrinsic clearance (Vmax/Km) was 2.9 +/- 0.5 micro l min-1 mg-1 microsomal protein in the extensive metabolizer livers. The corresponding values in the poor metabolizer livers were: apparent Km 124 +/- 141 micro m; Vmax 1.4 +/- 0.6 pmol min-1 mg-1 microsomal protein; and intrinsic clearance 0.026 micro l min-1 mg-1 microsomal protein. Quinidine almost completely inhibited perhexiline monohydroxylation activity, but inhibitors selective for other CYP isoforms had little effect. CONCLUSIONS: Perhexiline monohydroxylation is almost exclusively catalysed by CYP2D6 with activities being about 100-fold lower in CYP2D6 poor metabolizers than in extensive metabolizers. The in vitro data predict the in vivo saturable metabolism and pharmacogenetics of perhexiline.

Gastric outlet obstruction secondary to pancreatic cancer: surgical vs endoscopic palliation.

BACKGROUND: Gastric outlet obstruction in patients with pancreatic cancer has a grim prognosis. Open surgical bypass is associated with high morbidity, whereas endoscopic duodenal stenting appears to provide better palliation. METHODS: We reviewed the medical records of patients with gastric outlet obstruction secondary to pancreatic carcinoma who were admitted to our clinic between 1 October 1988, and 30 September 1998. The data included stage of disease, American Society of Anesthesiologists (ASA) class, surgical interventions, complications, and survival. RESULTS: A total of 250 patients with pancreatic cancer were identified. Twenty-five of them (10%) had gastric outlet obstruction. Of these 25, 17 were treated with gastrojejunostomy, six had duodenal stenting (Wallstent), and two were resectable. There was no significant difference between the gastrojejunostomy group and the duodenal stenting group in ASA class or stage of disease. For the gastrojejunostomy group, median survival was 64 days (range, 15-167) and postoperative stay in hospital was 15 days (range, 8-39). For the duodenal stenting group, median survival was 110.5 days (range, 42-212) and postoperative stay was 4 days (range, 2-6). Ten patients (58.8%) in the gastrojejunostomy group had delayed gastric emptying. All of the patients in the duodenal stenting group were able to tolerate a soft diet the day after stent placement. Thirty-day mortality in the gastrojejunostomy group was 17.64%; in the duodenal stenting group, it was 0. CONCLUSION: In pancreatic carcinoma patients with gastric outlet obstruction, duodenal stenting results in an earlier discharge from hospital and possibly improved survival.

A cost-effectiveness approach to drug subsidy and pricing in Australia.

The Australian government offers its citizens subsidies on a select list of pharmaceuticals. For a drug to qualify for inclusion on this list, its manufacturer must demonstrate that the drug is both clinically effective and cost-effective. In part, this measure, along with others, was introduced to improve clinical and economic outcomes. Although this evidence-based system has provided transparency and consistency in decision making about which drugs will be covered, it may not have contained the rate of increase in drug costs.

Co-prescribing of SSRIs and TCAs in Australia: how often does it occur and who is doing it?

AIMS: To determine the frequency with which the selective serotonin re-uptake inhibitor (SSRI) antidepressants are used as add-on therapy to the tricyclic antidepressants (TCA) rather than as replacement therapy. METHODS: The data analysed were profiles of prescription records by date of supply to the patient. From within the national administrative dispensing claims database, the subset eligible for social security entitlements was identified as individuals by means of their coded permanent identification numbers (PINs). Following the initial supply of an SSRI in January 1996, the subsequent 6 months dispensing of SSRI and TCA antidepressants to these individuals was examined. The main outcome measure was the proportion of individuals to whom SSRIs and TCAs were dispensed concurrently, as an indirect measure of coprescription. In instances where a patient was receiving prescriptions for SSRIs and TCAs that had been written by the one doctor only, the major specialty of the doctor was investigated. RESULTS: 55 271 PINs were dispensed 63 865 SSRI prescriptions in January 1996 which represented over half (52%) of the total community SSRI prescriptions dispensed in that month. The number of these patients meeting the criteria for coprescription of SSRIs and TCAs over the next 6 months was 2773 (5%). The coprescribing instances were highest in Queensland and the prescribers most frequently involved had psychiatry major specialty codes. CONCLUSIONS: Among SSRI users there is a cohort of patients who, within the same time frame, are receiving supplies of a TCA, the nonselective drug that the SSRIs were designed to replace. This is indirect evidence of probable coprescription. Such combination use is of uncertain clinical and cost effectiveness, and carries additional risks. The SSRIs were included on the subsidy list in Australia on the basis of reasonable cost effectiveness as monotherapy compared with the TCAs. Our data imply that for some patients, antidepressant prescribing is inconsistent with the basis on which government subsidy was approved.

Acute direct adenoviral vector cytotoxicity and chronic, but not acute, inflammatory responses correlate with decreased vector-mediated transgene expression in the brain.

The potential utility of adenoviruses for the treatment of chronic neurological disease is controversial due to reports of vector-associated toxicity, inflammation, and transient transgene expression. To focus upon the mechanism by which transgene expression is lost, we injected increasing doses [1 x 10(6) to 1 x 10(9) infectious units (iu)] of a first-generation adenovirus vector expressing beta-galactosidase into the brains of immune-competent adult rats. Transgene expression was evaluated simultaneously with acute neuronal and glial cell cytotoxicity, and acute and chronic inflammation using immunohistochemistry, at 3 and 30 days post-vector administration. Our results show a clear threshold effect of viral dose upon the amount of transgene expression persisting by 30 days after vector administration. Below 10(8) iu, transgene expression remained stable over the 30-day period. Following infection of more than 10(8) iu, the extent of transgene expression at 30 days was inversely correlated with increasing viral dose. The severity of acute inflammation increased proportionally with increasing vector dose from 10(6) to 10(9) infectious units. In contrast, acute vector-mediated cytotoxicity and chronic inflammation were observed only above the threshold level of vector dose. Above 10(8) iu both the extent of the acute toxicity and the severity of the chronic inflammation were inversely correlated with transgene expression at 30 days. Thus, our data suggest that both an acute loss of cells through direct vector-mediated toxicity and the elicitation of chronic inflammation (but not acute inflammation) may account for the decline in transduction persistence at high vector doses.

Impact of the Minimum Pricing Policy and introduction of brand (generic) substitution into the Pharmaceutical Benefits Scheme in Australia.

PURPOSE: To describe the effects of introducing the Minimum Pricing Policy (MPP) and generic (brand) substitution in 1990 and 1994 respectively on the dispensing of Pharmaceutical Benefits Scheme (PBS) prescriptions both at the aggregate and individual patient level. METHODS: The relative proportion of prescriptions with a brand premium and those at benchmark was examined 4 years after introduction of the MPP and again 5 years later after generic substitution by pharmacists was permitted. To determine the impact of a price signal at the individual level, case studies involving a patient tracking methodology were conducted on two drugs (fluoxetine and ranitidine) that received a brand premium. RESULTS: From a zero base when the MPP was introduced in 1990, there were 5.4 million prescriptions (17%) dispensed for benchmark products 4 years later in 1994. At this stage generic (brand) substitution by pharmacists was then permitted and the market share of benchmark brands increased to 45% (25.2 million) by 1999. In the patient tracking studies, a significantly lower proportion of patients was still taking the premium brand of fluoxetine 3 months after the introduction of a price signal compared with patients taking paroxetine which did not have a generic competitor. This was also the case for the premium brand of ranitidine when compared to famotidine. The size of the price signal also had a marked effect on dispensing behaviour with the drug with the larger premium (fluoxetine) showing a significantly greater switch away from the premium brand to the benchmark product. CONCLUSIONS: The introduction in 1990 of the Minimum Pricing Policy without allowing generic substitution had a relatively small impact on the selection of medicines within the Pharmaceutical Benefits Scheme. However the effect of generic substitution at the pharmacist level, which was introduced in December 1994, resulted in a marked increase in the percentage of eligible PBS items dispensed at benchmark. Case studies showed a larger premium resulted in a greater shift of patients from drugs with a brand premium to the benchmark alternative.

Nonspecific binding of drugs to human liver microsomes.

AIMS: To characterize the nonspecific binding to human liver microsomes of drugs with varying physicochemical characteristics, and to develop a model for the effect of nonspecific binding on the in vitro kinetics of drug metabolism enzymes. METHODS: The extent of nonspecific binding to human liver microsomes of the acidic drugs caffeine, naproxen, tolbutamide and phenytoin, and of the basic drugs amiodarone, amitriptyline and nortriptyline was investigated. These drugs were chosen for study on the basis of their lipophilicity, charge, and extent of ionization at pH 7.4. The fraction of drug unbound in the microsomal mixture, fu(mic), was determined by equilibrium dialysis against 0.1 M phosphate buffer, pH 7.4. The data were fitted to a standard saturable binding model defined by the binding affinity KD, and the maximum binding capacity Bmax. The derived binding parameters, KD and Bmax, were used to simulate the effects of saturable nonspecific binding on in vitro enzyme kinetics. RESULTS: The acidic drugs caffeine, tolbutamide and naproxen did not bind appreciably to the microsomal membrane. Phenytoin, a lipophilic weak acid which is mainly unionized at pH 7. 4, was bound to a small extent (fu(mic) = 0.88) and the binding did not depend on drug concentration over the range used. The three weak bases amiodarone, amitriptyline and nortriptyline all bound extensively to the microsomal membrane. The binding was saturable for nortriptyline and amitriptyline. Bmax and KD values for nortriptyline at 1 mg ml-1 microsomal protein were 382 +/- 54 microM and 147 +/- 44 microM, respectively, and for amitriptyline were 375 +/- 23 microM and 178 +/- 33 microM, respectively. Bmax, but not KD, varied approximately proportionately with the microsome concentration. When KD is much less than the Km for a reaction, the apparent Km based on total drug can be corrected by multiplying by fu(mic). When the substrate concentration used in a kinetic study is similar to or greater than the KD (Km >/= KD), simulations predict complex effects on the reaction kinetics. When expressed in terms of total drug concentrations, sigmoidal reaction velocity vs substrate concentration plots and curved Eadie Hofstee plots are predicted. CONCLUSIONS: Nonspecific drug binding in microsomal incubation mixtures can be qualitatively predicted from the physicochemical characteristics of the drug substrate. The binding of lipophilic weak bases is saturable and can be described by a standard binding model. If the substrate concentrations used for in vitro kinetic studies are in the saturable binding range, complex effects are predicted on the reaction kinetics when expressed in terms of total (added) drug concentration. Sigmoidal reaction curves result which are similar to the Hill plots seen with cooperative substrate binding.