Heterologous expression and purification of membrane-bound pyrophosphatases.

Membrane-bound pyrophosphatases (M-PPases) are enzymes that couple the hydrolysis of inorganic pyrophosphate to pumping of protons or sodium ions. In plants and bacteria they are important for relieving stress caused by low energy levels during anoxia, drought, nutrient deficiency, cold and low light intensity. While they are completely absent in mammalians, they are key players in the survival of disease-causing protozoans making these proteins attractive pharmacological targets. In this work, we aimed at the purification of M-PPases in amounts suitable for crystallization as a first step to obtain structural information for drug design. We have tested the expression of eight integral membrane pyrophosphatases in Saccharomyces cerevisiae, six from bacterial and archaeal sources and two from protozoa. Two proteins originating from hyperthermophilic organisms were purified in dimeric and monodisperse active states. To generate M-PPases with an increased hydrophilic surface area, which potentially should facilitate formation of crystal contacts, phage T4 lysozyme was inserted into different extramembraneous loops of one of these M-PPases. Two of these fusion proteins were active and expressed at levels that would allow their purification for crystallization purposes.

Immunonutrition in elective gastrointestinal surgery patients.

BACKGROUND: Previous trials have shown that perioperative immunonutrition could protect patients from infectious complications after gastrointestinal cancer operations. The purpose of this study was to determine whether perioperative immunonutrition decreases postoperative morbidity, especially infection complications, mortality and length of hospital stay in patients undergoing major gastrointestinal tract surgery. METHODS: One hundred patients with a planned elective operation for benign or malignant gastrointestinal illness were randomized into two groups: group 1) oral supplementation for five days before and five days after surgery with 900 mL/day of a formula enriched with arginine, gamma-3-fatty acid and RNA + liquid diet ad libitum on one and two postoperative day and then solid food (immunonutrition group; n = 50) or group 2) no artificial nutrition before and after surgery, on one and two postoperative day intravenous solution of 5% glucose and electrolytes and then normal diet (conventional group; n = 50). RESULTS: The groups were comparable for all key baseline and surgical characteristics. There were nine (18%) infectious complications in both groups. Overall complication rates were 28% (n = 14) in the immunonutrition group and 24% (n = 12) in the conventional group. No significant difference between the groups was found in complication rates, mortality or length of hospital stay. CONCLUSION: Routine perioperative immunonutrition to the patients undergoing major gastrointestinal surgery is not beneficial.

Cost effectiveness of preoperative ultrasound in primary parathyroid surgery.

A prospective, randomized, blind study was undertaken to assess whether preoperative ultrasound (US) localization of the abnormal parathyroid glands is cost-effective in patients undergoing initial neck exploration for primary hyperparathyroidism (PHPT). Twenty-eight patients were randomly allocated into two groups. In Group I the results of preoperative US were reported to the surgeon before exploration, and in Group II he was not informed of the US results. All patients underwent bilateral neck exploration, performed by the same surgeon. The operating room time was recorded and the operating room costs calculated. They included the total costs of cervical US in Group I. The cure and morbidity rates in Group I were 100% and 14% and those in Group II 86% and 7%, respectively (P > 0.05). The mean operating room time of 97 +/- 15 min in Group I was significantly lower than that of 113 +/- 23 min in Group II (P < 0.05). The mean operating room costs, however, were almost the same in both groups being only 286 FIM higher in Group II (P > 0.4) because the costs of preoperative US, the least expensive of the localization studies, of 497 FIM negated any cost savings achieved by the reduced operating room time. We thus conclude that preoperative US before initial neck exploration for PHPT is not cost-effective.

Parathyroid re-exploration in patients with primary hyperparathyroidism.

In a prospective study fifteen consecutive patients underwent re-exploration for persistent or recurrent primary hyperparathyroidism. We aimed at definite preoperative localization of enlarged, abnormal parathyroid glands in all patients. Ultrasound guided fine needle aspirations for parathyroid hormone assay had the highest accuracy rate of 100%, those for cervical ultrasound and thallium-technetium, scintigraphy were similar, both 86%. Normocalcaemia was achieved in all patients, but five (33%) patients required more than one re-exploration. Permanent unilateral vocal cord injury occurred in two (13%) patients, but none had permanent hypocalcaemia. We conclude that the results of re-exploration are good but one third of the patients required more than one reoperation. Localization studies aiming at definite localization are mandatory before re-exploration. On the basis of our results we suggest a protocol for preoperative localization which takes into consideration both the accuracy rates and the costs of localization examinations.

Preoperative ultrasound in patients undergoing initial neck exploration for primary hyperparathyroidism.

92 patients with primary hyperparathyroidism were subjected prospectively to preoperative ultrasound to identify enlarged parathyroid glands and detect simultaneous thyroid lesions of surgical significance. The overall sensitivity and specificity rates were 62.7% and 95.7%, 85.2% and 96.8% for patients with a single adenoma, 44.3% and 88% for those with multiglandular disease, 56.8% and 95.6% for those with simultaneous thyroid disease and 20% and 100% for mediastinal glands. Ultrasound revealed a pathological thyroid gland in 26 patients (28%). The sensitivity and specificity rates were 96% and 100%. The thyroid lesion was considered clinically significant in 15 of the 92 patients (16.3%) and only in three patients (3.3%) was the diagnosis not established preoperatively. Operating room times and costs were significantly lower for patients with an accurate ultrasound scan than for those with an inaccurate scan (P < 0.001). We conclude that both the sensitivity of ultrasound for locating enlarged parathyroid glands and its yield of previously unknown thyroid lesions of surgical significance are too low to warrant preoperative ultrasound evaluation. Although our operating room expenses were significantly higher for patients with an inaccurate scan, the cost effectiveness of preoperative ultrasound cannot be proved until a prospective, randomised investigation has been carried out.

Morbidity and mortality after abdominal operations for cancer.

A review of 316 consecutive patients operated on for intra-abdominal cancer was undertaken to analyze the incidence of postoperative morbidity and mortality and identify contributory factors. The overall postoperative morbidity and mortality after operations performed for primary cancer were, respectively, 26% (75/288) and 7% (20/288), and those seen after operations for recurrent cancer 43% (12/28) and 29% (8/28). Factors that were statistically associated with a fatal outcome were recurrent cancer (p = 0.0005), palliative resection (p = 0.005), a serum albumin level under 35 G/L (p = 0.011) and a weight loss of more than 5 kilograms (p = 0.046); those associated with a significantly greater complication rate were a serum albumin level under 35 G/L (p = 0.000004), a weight loss of more than 5 kilograms (p = 0.00007), intra-abdominal carcinosis (p = 0.0057), and an emergency procedure (p = 0.048). Infective complications were more common among the patients who underwent resective surgery than among those operated on non-resectively. It is concluded that preoperative malnutrition is predictive of both postoperative complications and increased mortality, and that recurrent disease and palliative surgery are associated with high postoperative mortality.

Results of surgery in primary hyperparathyroidism.

In a prospective study 92 consecutive patients with biochemically proved primary hyperparathyroidism underwent initial neck exploration at Oulu University Hospital. The incidence of multiglandular disease was 34%. 23 patients (25%) underwent a simultaneous thyroidectomy. The cure rate after initial exploration was 91.3%, ectopic parathyroid glands and multiglandular disease being the most common causes of failure. Simultaneous thyroidectomy increased somewhat but not significantly the complication risk. There was a slight tendency for serum calcium concentrations to increase during the mean follow-up of 2.3 +/- 1.5 years. Four patients with persistent hypercalcaemia underwent a successful reoperation during that time. Thus the overall cure rate was 95.6%, but 5.4% of the patients required permanent medication for hypocalcaemia. We conclude that the most common causes for failed initial exploration were ectopic parathyroid glands and multiglandular disease. The incidence of multiglandular disease was unusually high in this series. Because simultaneous thyroidectomy increased somewhat the complication risk of initial neck exploration, the indications for this additional procedure should be carefully considered. The results of parathyroid surgery were good and dependent on how many patients underwent reoperation during the follow-up. A consequence of tendency for serum calcium concentrations to increase during the follow-up could be that a definite cure cannot always be attained in cases of primary hyperparathyroidism.

Effect of biliary tract procedures on duodenogastric reflux and the gastric mucosa.

The effect of alterations in the biliary tract on the gastric milieu was evaluated in gallstone disease and after cholecystectomy or biliary fenestration and compared with a control group. Endoscopic bile reflux was often found in gallstone patients (67%) and almost always after cholecystectomy (89%). Gastric bile acid concentrations were greater in the gallstone patients than in the control patients, were higher still after cholecystectomy, and were highest after biliary fenestration. The pH of the gastric fluid was more alkaline in the cholecystectomized groups. Lysolecithin concentrations were generally low and did not differ between the groups, nor was there any difference in scintigraphic reflux between the groups. Endoscopic erythematous gastritis correlated with the grade of bile reflux and was a common finding after biliary tract procedures. There were no consistent histologic findings associated with duodenogastric reflux. Helicobacter pylori colonization rates were similar in the various patient groups and were not affected by the reflux grade.

Sucralfate versus placebo in treatment of non-ulcer dyspepsia.

One hundred fifty-one patients with non-ulcer dyspepsia, defined as chronic epigastric pain without concomitant symptoms of the irritable bowel syndrome and with no evidence of any organic disease other than macroscopic or microscopic gastritis/duodenitis seen at endoscopy on entry into the trial, were randomly assigned to treatment for four weeks with sucralfate or a placebo, 1 g three times a day one-half hour before meals, according to a double-blind model. Seventy-nine patients received sucralfate and 72 patients received a placebo. According to patients' subjective assessment of their symptoms at four weeks, 61 patients (77 percent) in the sucralfate group and 40 patients (56 percent) in the placebo group had become symptom-free or showed improvement, whereas the condition of 18 (23 percent) in the former group compared with 32 (44 percent) in the latter group remained unchanged or deteriorated. The difference between the groups was significant (p less than 0.01). The best response to sucralfate treatment (84 percent or more symptom-free or improved) was achieved in patients with mild or moderate symptoms and without macroscopic or microscopic inflammation of their gastric mucosa--a typical patient with non-ulcer dyspepsia. Our results indicate that sucralfate is significantly more effective than placebo in the treatment of non-ulcer dyspepsia.