Blastocyst formation, embryo transfer and breed comparison in the first reported large scale cloning of camels.

Cloning, through somatic cell nuclear transfer (SCNT), has the potential for a large expansion of genetically favorable traits in a population in a relatively short term. In the present study we aimed to produce multiple cloned camels from racing, show and dairy exemplars. We compared several parameters including oocyte source, donor cell and breed differences, transfer methods, embryo formation and pregnancy rates and maintenance following SCNT. We successfully achieved 47 pregnancies, 28 births and 19 cloned offspring who are at present healthy and have developed normally. Here we report cloned camels from surgical embryo transfer and correlate blastocyst formation rates with the ability to achieve pregnancies. We found no difference in the parameters affecting production of clones by camel breed, and show clear differences on oocyte source in cloning outcomes. Taken together we demonstrate that large scale cloning of camels is possible and that further improvements can be achieved.

Electrophysiological and behavioral activity of secondary metabolites in the confused flour beetle, Tribolium confusum.

Several previous studies have addressed pheromone communication in various flour beetles (Coleoptera: Tenebrionidae), including the confused flour beetle, Tribolium confusum (du Val). Different stereoisomers of 4,8-dimethyldecanal (DMD) were reported as the only components of an aggregation pheromone, but the behavioral activity of DMD is low. In the present study, additional previously reported secondary metabolites (benzoquinones and hydrocarbons) were tested for electrophysiological activity (EAG) with both sexes of T. confusum. Two benzoquinones and three monoenic hydrocarbons elicited significant EAG activity from both male and female antennae. There was an elevated male EAG response (vs. the females) to two out of the three hydrocarbons and for both quinones. The EAG-active compounds were subsequently investigated for behavioral activity in a walking bioassay. Benzoquinones are considered toxic and have been assigned a function as alarm substances in flour beetles, but we found that methyl-1, 4-benzoquinone in intermediate concentrations was attractive to both male and female beetles and could therefore act as an aggregation pheromone component. Males were also attracted to ethyl-1,4-benzoquinone. The corresponding hydroquinones, presumed precursors of the benzoquinones, did not elicit any electrophysiological response and were not tested for behavioral activity. The unsaturated hydrocarbons (1-tetradecene, 1-pentadecene, and 1-hexadecene) elicited significant EAG responses from both male and female antennae and were also attractive in the behavioral assay. Our results show that several beetle-produced compounds, in addition to 4,8-dimethyldecanal, may be part of a complex pheromone system in flour beetles and play a role in mediating aggregation in T. confusum.

Electrophysiological and behavioral responses to chocolate volatiles in both sexes of the pyralid moths Ephestia cautella and Plodia interpunctella.

Volatiles from chocolate mediate upwind flight behavior in Ephestia cautella and Plodia interpunctella. We used gas chromatography with electroantennographic detection and found 12 active compounds derived from three different chocolate types, i.e., plain, nut-containing, and rum-flavored. Eight of the compounds were identified with mass spectrometry, and the activity of three compounds, ethyl vanillin, nonanal, and phenylacetaldehyde (PAA), was subsequently confirmed in both electrophysiological and behavioral assays. In the electroantennogram experiment, PAA and nonanal were consistently eliciting responses in both species and sexes. Ethyl vanillin was active in males of both species, and also in P. interpunctella females. E. cautella females showed no antennal activity in response to ethyl vanillin. All three volatiles were attractive to E. cautella males and P. interpunctella females in a flight tunnel. E. cautella females were significantly attracted only to ethyl vanillin. P. interpunctella males were attracted to PAA. Ethyl vanillin is a novel insect attractant, whereas both nonanal and phenylacetaldehyde mediate behavior in many insect species. A final experiment revealed that a blend of the three volatiles was required to induce landing in the flight tunnel bioassay, and that the landing rate was dependent on dose. The three-component blend attracted both sexes of P. interpunctella and females of E. cautella, whereas E. cautella males were not attracted.

Combination-therapy with bedtime nph insulin and sulphonylureas gives similar glycaemic control but lower weight gain than insulin twice daily in patients with type 2 diabetes.

BACKGROUND: To study the effect on body weight and glycaemic control of two insulin treatment regimens in patients with Type 2 diabetes and moderate failure to oral hypoglycaemic agents. METHODS: Sixteen patients treated with oral hypoglycaemic agents (6 men and 10 women) were included in this open-label, randomized, parallel group study. Their age was 62 +/- 2 (mean +/- SEM) years (range 44-79 years), body weight 71.3 +/- 2.9 kg, body mass index (BMI) 24.6 +/- 0.8 kg/m(2). The patients were switched to insulin treatment with bedtime NPH insulin combined with daytime sulphonylurea (combination group) or twice daily injections of a premixed combination of regular human and NPH insulin (insulin twice daily group) with measurements as given below before and after 12 and 24 weeks of treatment. RESULTS: HbA(1c) was lowered from 8.3 +/- 0.3% to 7.0 +/- 0.2% in the insulin twice daily group (p<0.05) and from 8.3 +/- 0.3% to 6.8 +/- 0.5% in the combination group (p<0.03; ns between treatment groups). Body weight increased from 71.7 +/- 4.0 kg to 77.6 +/- 4.4 kg in the insulin twice daily group (p<0.001) and from 70.8 +/- 4.6 kg to 72.7 +/- 5.1 kg in the combination group (ns; p<0.02 between groups). The dose of insulin at 24 weeks in the insulin twice daily group was 45.8 +/- 4.2 U and 29.4 +/- 5.4 U in the combination group (p=0.03). Combination treatment reduced fasting and stimulated C-peptide levels. CONCLUSIONS: Both treatments improved glycaemic control to the same extent but the combination of bedtime NPH insulin and daytime sulphonylurea gave a very small increase of body weight over a 6 months period. We conclude that combination therapy is an attractive alternative when starting insulin treatment in patients with Type 2 diabetes as this is a critical period for weight gain in such patients.

The use of human ultralente is limited by great intraindividual variability in overnight plasma insulin profiles.

UNLABELLED: Our objective was to investigate the usefulness of human ultralente insulin as basal substitution overnight in patients with Type 1 diabetes treated with multiple insulin injection therapy by evaluating the free insulin and glucose profiles, the day-to-day variability and the impact of the time of injection. METHODS: Ten patients with Type 1 diabetes and with good metabolic control (mean HbAlc 6.0%), treated with regular human insulin before breakfast, lunch and dinner and human ultralente (Ultratard) before dinner or at bedtime, were studied. Plasma profiles of blood glucose and free insulin were measured on three occasions from 16.00 h until noon the next day. On two of these occasions Ultratard was injected before dinner and once it was injected at bedtime in randomized order. RESULTS: Injection of regular insulin before dinner resulted in a high insulin peak during the evening but no insulin peak was found that could be attributed to ultralente. The plasma concentration of free insulin at 03.00 h was 11.0+/-1.9 mU/L and it slowly decreased to 6.4+/-1.4 at 12.00 h after administration of ultralente at 17.00 h. There were no differences in the mean plasma insulin profiles compared to the other occasion when insulin was given at 17.00 h or at 22.00 h. On the other hand, the intra-individual day-to-day variability of mean insulin concentration during the night was considerable, often exceeding 50%. No differences were noted in the mean blood glucose profiles between the three occasions. CONCLUSION: Human ultralente insulin gives an insulin profile suitable for overnight substitution, but the great day-to-day variability limits its usefulness. It can be injected before dinner or at bedtime without any change in the insulin profile during the night.

No pharmacokinetic effect of retaining the infusion site up to four days during continuous subcutaneous insulin infusion therapy.

This study was designed to investigate whether the indwelling time of the needle during continuous subcutaneous insulin infusion (CSII) could contribute to one of the main problems with subcutaneous insulin administration, that is the day-to-day intra-patient variation in absorption. The effect on insulin kinetics after a bolus dose was investigated at different time intervals up to 4 days after insertion of an indwelling catheter in situ for CSII therapy. Nine insulin-dependent diabetic patients treated with pump (Nordisk Infuser; Velosulin 100 U ml-1) took part in the study. The evening before day 1 of the investigation a needle connected to the pump with a polyethylene catheter was inserted subcutaneously on the abdominal wall where it remained throughout the investigation. There were no significant differences in blood glucose or plasma free insulin profiles between days 1, 3, and 5. The intra-patient coefficient of variation of free insulin was 38% at basal and 19% at post-bolus state. We conclude that during CSII it is acceptable from a pharmacokinetic point of view to retain the infusion site for up to 4 days.

Male predominance of type 1 (insulin-dependent) diabetes mellitus in young adults: results from a 5-year prospective nationwide study of the 15-34-year age group in Sweden.

The incidence of diabetes mellitus in Sweden in the 15-34 year age group was prospectively studied on a nationwide basis, beginning 1 January 1983. A total of 1,214 male and 720 female cases of newly-diagnosed (excluding gestational) diabetes were reported over a 5-year period. This corresponds to an incidence of 20.5 per 100,000/year in male subjects and 12.7 per 100,000/year in female subjects. Most cases were classified as Type 1 (insulin-dependent) diabetes, with an incidence of 15.9 in males and 8.6 in females. The incidence of Type 1 diabetes decreased gradually with age, while the incidence of Type 2 (non-insulin-dependent) diabetes increased. A male predominance was found in all age groups, with a male-to-female ratio of 1.8:1 for Type 1 diabetes and 1.3:1 for Type 2 diabetes. Maximum blood glucose concentration at diagnosis was significantly higher in males than in females in both Type 1 and Type 2 diabetic subjects. In contrast, the percent desirable weight was significantly higher in females, both in Type 1 and Type 2 diabetic subjects. The difference in diabetes incidence therefore cannot be attributed to any methodological error. The present finding of a marked male predominance after puberty in Type 1 diabetes in an ethnically quite homogeneous population supports the hypothesis that environmental risk factors and life-style are important for the development of the disease.

Pre- and postprandial hyperinsulinemia during insulin pump treatment; role of the subcutaneous bolus and basal infusion.

The impact of the bolus dose and the basal infusion during insulin pump treatment on pre- and post-prandial hyperinsulinemia was investigated in 8 insulin-dependent diabetic patients. Frequent determinations of free insulin in blood and simultaneous determination of glucose consumption with euglycaemic clamp technique were made after a bolus of 5 U with or without basal infusion and after 10 U with basal infusion. The maximum free insulin level was reached after 75, 65 and 105 min respectively (median; NS), whereas 50% of the maximum was reached already after 15, 10, and 12.5 min (NS). The fall in free insulin after the peak was slower in all protocols compared to the breakfast induced peak in healthy subjects and it was slower with than without basal infusion (p less than 0.03). Glucose consumption followed the free insulin profiles. We conclude, that administration of a subcutaneous bolus with an insulin pump gives a dose-related free insulin peak that is unphysiological in comparison with the normal meal-related insulin secretion. Preprandial hyperinsulinemia, found after a prescribed interval of 30 min between injection and meal, as well as postprandial hyperinsulinemia, increased with a greater bolus indicating that adjustments of timing of meals and snacks may be necessary when the dose is changed. Furthermore, the insulin substitution with external pumps would probably be more physiological if the basal supply was interrupted after a bolus.

Free insulin profiles during intensive treatment with biosynthetic human insulin.

24 h profiles of free insulin and glucose were determined in insulin-dependent diabetic patients on intensive insulin regimens with biosynthetic human insulin, either as continuous subcutaneous insulin infusion (CSII) with mealtime bolus doses (n = 6), or intensified conventional insulin therapy (ICIT) with preprandial injections of regular insulin and intermediate-acting insulin at bedtime (n = 6). The free insulin profiles were similar to the normal profiles but there were some important differences: CSII gave hyperinsulinaemia at daytime compared to normal people (p less than 0.05) and also to ICIT (p less than 0.005); ICIT but not CSII gave hyperinsulinaemia at midnight (p less than 0.05) whereas fasting free insulin was too-low to keep blood glucose normal; the insulin peaks after the bolus doses were retarded with a maximum after 30-90 min and a return to the basal level after 5-7 h (ICIT) or 8-9 h (CSII). The height of the insulin peaks were of similar magnitude at all meals and did not differ significantly between ICIT, CSII, and normal people. Time-to-peak was dependent on the injection level. We conclude, that intensive regimens with biosynthetic hum insulin do not give normoinsulinaemia but insulin profiles that resemble physiology. Biosynthetic human NPH insulin may be rather short-acting for overnight glucose control. The interval that should be recommended between preprandial insulin dose and meal may vary depending on the preinjection insulin level.

Overnight metabolic control with bedtime injection of intermediate-acting insulin or continuous subcutaneous insulin infusion.

Ten insulin-dependent diabetic patients were investigated from 2100 to 0700 h during treatment with either a bedtime injection (BI) of intermediate-acting insulin or continuous subcutaneous insulin infusion (CSII) at a constant basal rate. In the evening, blood glucose was slightly higher during treatment with BI than with CSII, whereas the metabolic control in the morning was equal on both regimens with a fasting blood glucose of 5.7 mM (4.2-7.1) (median and interquartile ranges) on BI and 5.4 mM (4.6-5.8) on CSII (NS). No rise in morning blood glucose was seen, but serum beta-hydroxybutyrate tended to rise (NS). There was a significant hyperinsulinemia at midnight during BI compared with CSII with a serum free insulin of 14.5 (11.7-16.0) vs. 9.6 (7.2-11.2) mU/L (P less than .05), respectively, and the area under the curve during the middle of the night (midnight to 0400 h) was greater with BI than CSII (P less than .02). A greater fall in blood glucose was seen with BI than with CSII during this period (P less than .02). Differences in blood glucose and serum free-insulin profiles between those using NPH or lente insulin at bedtime were registered. We conclude that, although the same metabolic control in the morning was achievable with CSII at a constant basal rate and BI, CSII is superior for overnight metabolic control due to less-pronounced hyperinsulinemia during the night and a steady-state level of free insulin in the morning.

Miscibility of human semisynthetic regular and lente insulin and human biosynthetic regular and NPH insulin.

The miscibility of human semisynthetic regular and lente insulins (Actrapid human/Monotard human) and human biosynthetic regular and NPH insulins (Humulin regular/Humulin NPH) was studied in vitro and in 16 insulin-dependent diabetic patients. In vitro a decrease of regular insulin was found immediately after mixture with semisynthetic lente insulin in proportions 1:4, 1:3, 1:2, 1:1, and 2:1. This was not found after mixture with human biosynthetic regular and NPH insulins. Free insulin and blood glucose were determined over 8 h after the morning injection of regular/lente (n = 9) or regular/NPH (n = 7) insulins in proportions used by the patients (approximately 1:2) mixed in one syringe or in separate syringes. Mixing the regular/lente preparations in one syringe immediately before injection resulted in a significant loss of the early rise in free insulin (P less than .01), with a tendency to a higher free insulin in the afternoon. A corresponding change in the blood glucose profile was seen. No changes were observed in free insulin or blood glucose after mixing regular and NPH preparations compared with separate injections. We conclude that mixing human semisynthetic regular and lente insulins (Actrapid human/Monotard human) instantly results in a decrease of regular insulin, probably due to formation of a longer-acting preparation, whereas no such changes occur with human biosynthetic regular and NPH insulins (Humulin regular/Humulin NPH).

Free and total insulin as determined after precipitation with polyethylene glycol: analytical characteristics and effects of sample handling and storage.

We evaluated results of radioimmunoassays of free and total insulin after precipitation of endogenous antibodies with polyethylene glycol (PEG), and we investigated the influence of collection time, temperature, and storage in heparin- cr EDTA-treated plasma or serum on results for free insulin. Analytical recovery of free insulin was 99.3%, of total insulin 96.4%. For free insulin, assay precision (CV) was 4.0-13.0% (intra-assay) and 7.8-10.7% (inter-assay); for total insulin, 3.6-9.5% and 6.6-11.7%, respectively. Free insulin decreased in plasma (p less than 0.05) and serum (p less than 0.01) at room temperature after 3 h and in promptly analyzed serum (p less than 0.01). Storage of samples at -20 degrees C increased the concentration of free insulin in plasma (p less than 0.025) and serum (p less than 0.005), whereas the free insulin content of supernates after PEG precipitation was stable, except for a slight decrease in serum samples (p less than 0.02). We conclude that, for radioimmunoassay of free and total insulin, plasma should be used, treated with PEG without delay; supernates then are analytically stable for as long as 26 weeks at -20 degrees C.

Subcutaneous insulin substitution in insulin-dependent diabetes mellitus. Pharmacokinetic and pharmacodynamic studies.

Determination of free and total insulin with radioimmunoassay, after precipitation of endogenous insulin antibodies with polyethylene glycol, was evaluated. Insulin substitution in insulin-dependent diabetic patients was investigated, embracing the 24 h free insulin and glucose profiles with different regimens, the miscibility of insulin preparations, the overnight metabolic control, and bolus doses of different size with infusion pumps. In the free and total insulin assay precipitation of immunoglobulins with polyethylene glycol was almost complete and the recovery was high. Compared to immediately precipitated and assayed plasma samples at 37 degrees C, free insulin slightly decreased in immediately processed serum (20 degrees C), and also in plasma after 3 h at 20 degrees C. In stored (-20 degrees C) unprecipitated plasma samples free insulin increased after 4 weeks and also in serum samples after 26 weeks, whereas stored PEG-supernates were stable. In healthy controls a low basal insulin was found, increasing about tenfold postprandially. No morning rise in free insulin or glucose was found. The 24 h free insulin profile was strikingly unphysiological with 1 or 2 dose regimens; there was preprandial and nocturnal hyperinsulinaemia but absence of meal-related free insulin peaks. A considerable glucose rise was found after breakfast. Intensive regimens with conventional injections or infusion pumps, gave 24 h free insulin profiles that were similar to the physiological. However, the prandial peaks were retarded; and hyperinsulinaemia was shown with infusion pumps during daytime. An immediate loss of regular insulin was demonstrated after mixture with semisynthetic human lente insulin in vitro and in vivo, but not after mixture with biosynthetic human NPH insulin. The morning glucose control was similar with a bedtime injection of intermediate-acting insulin or continuous subcutaneous insulin infusion, but less hyperinsulinaemia overnight was found with the infusion pump at a constant rate. No early morning glucose rise was demonstrated. Dose-related free insulin profiles were shown after bolus doses with an infusion pump, although they were retarded compared to the physiological postprandial response. The postprandial hyperinsulinaemia was aggravated by continuous subcutaneous insulin infusion. Glucose consumption during euglycaemic clamp corresponded to the free insulin profiles, indicating that free insulin represents the biologically active hormone.

Unilateral reconstruction of the iliac artery. A comparative evaluation.

Unilateral reconstruction of the iliac artery because of obliterating atherosclerotic disease was performed on 122 patients (group A) in the period 1971 through 1979. A bifurcation dacron bypass was used for iliac artery reconstruction in 52 patients (group B) during the same period. Of the group A patients, 24% required eventual contralateral iliac reconstruction, and in 16% redoing of the ipsilateral operation became necessary. In group B the need for redoing was 25%. The 30-day mortality was 1.6% in group A and 7.8% in group B. The patency rates were similar up to 5 years postoperatively. The authors conclude that there is a place for unilateral iliac artery reconstruction, especially in high-risk cases or for patients with short life expectation who require relief of the vascular obstruction but for whom bilateral operation is not immediately mandatory.

Free insulin profiles in insulin-dependent diabetics treated with one or two insulin injections per day.

Twenty-four hour profiles of free insulin and blood glucose were determined in 12 healthy controls and 10 insulin-dependent diabetics treated with insulin regimens based on intermediate-acting insulin injected subcutaneously once or twice a day. The diabetics were ambulatory and in a good glycemic control, i.e. without hyperglycemic symptoms or frequent hypoglycemias and with HbA1 less than 9% (reference value 5.9-7.8%). Body weight was normal and median age (32 years) was the same in both groups. Free insulin was determined after polyethylene glycol precipitation of antibody-bound insulin. The controls had a low basal insulin level (median fasting value 3.9 mU/l) and postprandial peaks with a maximum within 30-60 min. There was no rise in plasma free insulin or blood glucose in the early morning hours. The free insulin profiles in the diabetics were highly unphysiological with hyperinsulinemia between the meals and during the night. The highest plasma free insulin value during the 24 hours was reached before lunch (approximately 5-fold compared to normals, p less than 0.01). Postprandially the free insulin concentrations did not reach the peak levels of the normals. After breakfast, blood glucose rose considerably in the diabetics (p less than 0.02 compared to normals) while the rise after lunch and dinner was not higher than in the healthy controls. The difficulties in glycemic control in the diabetic group, i.e. a blood glucose rise after breakfast and hypoglycemias in some patients, could largely be explained by the unphysiological insulin profiles.

Late mortality and morbidity after arterial reconstruction: the influence of arteriosclerosis in popliteal artery trifurcation.

Mortality and the incidence of myocardial infarction and cerebrovascular lesion have been retrospectively analyzed in patients who have survived 30 days after lower limb vascular reconstruction. These events have been related to the extent of arteriosclerotic lesions in the trifurcation of the popliteal artery (trifurcational disease, TFD) in 368 patients treated consecutively. One hundred fifty-five patients underwent aortoiliac reconstruction, 229 had femorodistal bypass, and 16 underwent both procedures. The median follow-up period was 4.4 years. The mortality rate in male and female patients was increased compared with the expected mortality. This increase was found both in patients with and without TFD. In patients having TFD the mortality was 2.0 times higher than in patients who did not have TFD (p less than 0.001). The difference in the mortality rate persisted after correction for the influence of age, sex, and diabetes mellitus. The higher mortality rate in patients who had TFD was most marked early in the postoperative period and was mainly caused by myocardial infarction. The incidence of myocardial infarction and cerebrovascular events was significantly increased in patients who had TFD compared with those who did not, particularly in the early postoperative period.

Usefulness of the on-line data base CANCERLIT: an evaluation study based on consecutive searches in CANCERLIT and MEDLINE for oncologists.

Two on-line data bases of the National Library of Medicine, CANCERLIT and MEDLINE, were evaluated in a user-oriented study for their usefulness for oncologists in cancer research and in clinical practice. The CANCERLIT data base was preferred for 12 of 16 literature searches requested by oncologists, and in 4 instances no preference was indicated. The implications of the study are discussed with regard to data base content and user preferences.

Arteriosclerosis in popliteal artery trifurcation as a predictor for myocardial infarction after arterial reconstructive operation.

The frequency of myocardial infarction and mortality within 30 days after lower limb vascular reconstruction, in relation to the extent of atherosclerotic lesions in the trifurcation of the popliteal artery, have been retrospectively analyzed in 158 consecutive patients reconstructed in the aortoiliac region and 239 consecutive patients undergoing femorodistal bypass. Among the patients without trifurcational disease (TFD) none had myocardial infarction develop postoperatively, as compared with four of the 50 patients with TFD in the aortoiliac series (p less than 0.05) and 18 of the 174 patients with TFD in the femorodistal series (p less than 0.05). In nine instances, the cause of death was myocardial infarction. The strong correlation between postoperative myocardial infarction and the presence of TFD, may be due to a direct correlation between coronary artery disease and TFD. The finding is of practical importance in the selection of treatment for patients with circulatory disorders of the lower limbs. The finding facilitates the preoperative identification of patients liable to have myocardial infarction develop. Indications for operation can be made more stringent and optimal intraoperative and postoperative monitoring can be instituted.