Preparation and characterization of a colloidal gold-insulin complex with binding and biological activities identical to native insulin.

We studied the binding and biological activities of gold-insulin complexes to develop a complex with properties identical to native insulin. Stabilizing amounts of insulin absorbed to 5-, 10-, or 15-nm gold particles resulted in complexes with 40-327 insulin molecules per gold particle and 4-111 times the biological activity of unlabeled insulin, based on the molar concentration of gold complex. These data suggested that these complexes behaved as multivalent ligands. Gold-insulin complexes were prepared with 5% of the stabilizing insulin concentration and were stabilized with bovine serum albumin. This resulted in a complex with 5-7 insulin molecules per 10-nm gold particle, which stimulated glucose oxidation in rat adipocytes and competed with [125I]-insulin for binding to the insulin receptor identically to unlabeled insulin on an equimolar basis. The organization and distribution of insulin receptors occupied by this monovalent-behaving gold-insulin complex were virtually identical to previous observations using monomeric ferritin-insulin. Since multivalent ligands may affect receptor binding, re-distribution, and intracellular processing, the use of electron-dense probes that resemble the unlabeled ligand in biological and binding properties is appropriate when studying receptor dynamics of in vivo or in vitro biological systems. The gold-insulin complex developed in this study should serve this function.

Neodymium-YAG laser. A new palliative tool in the treatment of colorectal cancer.

Twenty-four patients with colorectal carcinoma and one patient with a recurrent, transitional-cell, vesical tumor invading the rectum but not amenable to surgical resection underwent neodymium-YAG laser therapy. There were 16 men and nine women with an average age of 75.6 years (range, 35 to 93 years). There were 14, 6, 1, 1, and 3 tumors found in the rectum, rectosigmoid colon, transverse colon, descending colon, and ascending colon, respectively. Symptoms consisted of bleeding or partial obstruction. Twelve patients were high-risk surgical candidates, 11 patients had widespread disease, and two patients had another primary tumor not controlled by a previous surgical procedure. The tumor was eradicated in two patients. There were two complications, namely, a perforation and a rectovaginal fistula. Thirteen patients died from their disease. The remaining patients have been kept free from obstruction and bleeding.

Nd-YAG laser applications in surgical endoscopy: a single center comprehensive experience.

A retrospective review of endoscopic procedures using the Nd-YAG laser was carried out for patients treated between September 1984 and November 1986. Two hundred twenty two procedures were performed on 104 patients (58M:46W) during this 26 month period. The study encompasses a unique period of time in this center, as it includes the initial use of the endoscopic laser technique, the learning curves associated with its application, and finally, the time when its use became routine. Treatment of tumors was the indication in 47 per cent of the cases. Gastrointestinal bleeding of benign cause was the indication in nearly half of the patients; arteriovenous malformations were the most common causes of bleeding. Laser photocoagulation of hemorrhoids was performed in 11 cases. Upper endoscopy was the route in 33 per cent of procedures. Anoscopy, flexible sigmoidoscopy, and colonoscopy were the routes in the remainder. The time spent on the procedure and the energy delivered were found to be variables of the type of lesion and the endoscopic route. Nd-YAG laser endoscopy can be used safely and effectively for a variety of bleeding and obstructive gastrointestinal conditions. Appropriate certification for granting of privileges to laser endoscopists is of utmost importance.

Effect of tunicamycin on 3-hydroxy-3-methylglutaryl coenzyme A reductase in C-6 glial cells.

The effects of tunicamycin on 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase activity and cholesterol biosynthesis have been studied in cultured C-6 glial cells. Depending on culture conditions, exposure to tunicamycin caused either a marked inhibition of induction of HMG-CoA reductase activity or, under steady state conditions, a marked reduction in enzymatic activity. Incorporation of [14 C]acetate into sterols was affected similarly. After a 24-h exposure, a 50% reduction in reductase activity was observed with a concentration of 0.05 micrograms/ml, and a maximal, 65-70% reduction occurred with 0.10 micrograms/ml of the drug. The effect of tunicamycin on reductase activity and on sterol synthesis was apparent 4 h after addition of the drug and nearly maximal after 6 h. The relative specificity of the effect of tunicamycin was indicated by the finding of no change in the activities of NADPH-cytochrome c reductase, acetyl-CoA carboxylase, or fatty acid synthetase, in incorporation of [3H]leucine into total protein, or in the rate of increase in cellular protein and phospholipid at concentrations of tunicamycin that caused the marked effect on HMG-CoA reductase. The reversibility of the effect of tunicamycin was shown by observing total recovery of reductase activity within 24 h after removal of the drug following a 24-h exposure. That the effect of tunicamycin on reductase is related to the drug's effect on glycoprotein synthesis was shown in two ways. First, the range of concentrations over which tunicamycin led to the decrease in reductase activity was essentially identical with the range over which the drug led to a decrease in incorporation of [3H]mannose into protein. Second, incubation of C-6 cells with N-acetylglucosamine simultaneously with tunicamycin was accompanied by prevention of the drug's effect on both HMG-CoA reductase and glycoprotein synthesis. These data suggest that glycoprotein synthesis is necessary for the expression of HMG-CoA reductase activity and, thereby, cholesterol synthesis in glial cells. Moreover, a link between glycoprotein and cholesterol biosynthesis could play a role in the mediation of certain maturational events in cells of neural origin.

Insulin and alpha 2-macroglobulin-methylamine undergo endocytosis by different mechanisms in rat adipocytes: I. Comparison of cell surface events.

This ultrastructural study compared the endocytosis of a peptide hormone, ferritin-labeled insulin (Fm-I) or gold-labeled insulin (Au-I), and a non-hormonal ligand, gold-labeled alpha-2-macroglobulin-methylamine (Au-alpha 2MGMA), by rat adipocytes. Quantitative analysis of the cell surface showed that coated pits occupied 0.4% of the adipocyte surface. This was one fifth to one tenth of that which has been reported on fibroblasts and hepatocytes, cell types in which receptor-mediated endocytosis has been extensively studied. In contrast, uncoated micropinocytotic invaginations were quite numerous and occupied 13.1% of the adipocyte cell surface. The frequency of micropinocytotic invaginations, 13.8 per micron 2 of plasma membrane, was 7-12 times greater than has been reported on fibroblasts. Therefore, the ultrastructure of the endocytic apparatus on rat adipocytes was different from more commonly studied cell types. At 4 degrees C, Au-alpha 2MGMA concentrated within coated pits to a density that was 52 times greater than that on the uncoated plasma membrane. Au-alpha 2MGMA was excluded from micropinocytotic invaginations by more than 93%; this exclusion was unrelated to the size of the Au-alpha 2MGMA particle. In contrast, at 4 degrees C, Fm-I did not concentrate within coated pits and occupied micropinocytotic invaginations in a random manner. At 37 degrees C, coated pits accounted for all of the endocytosis of Au-alpha 2MGMA, proving that these structures were functional despite their atypically low density. In contrast, greater than 99% of the endocytosis of Fm-I or Au-I occurred through micropinocytotic invaginations. These results demonstrated for the first time by a comparative, quantitative, ultrastructural method that insulin and Au-alpha 2MGMA undergo endocytosis by dissimilar mechanisms on rat adipocytes. Dissimilarities in the endocytosis of insulin and Au-alpha 2MGMA may be related to the different biological roles of these two molecules.

Insulin and alpha 2-macroglobulin-methylamine undergo endocytosis by different mechanisms in rat adipocytes: II. Comparison of intracellular events.

A previous ultrastructural study showed that gold-labeled insulin (Au-I) and the non-hormonal ligand gold-labeled alpha-2-macroglobulin-methylamine (Au-alpha 2MGMA) underwent endocytosis by dissimilar cell surface structures on rat adipocytes. The present ultrastructural study compared the intracellular routes taken by these two ligands in adipocytes. Intracellular Au-alpha 2MGMA was initially found within apparent coated vesicles but Au-I was not, consistent with the previous demonstration that Au-alpha 2MGMA underwent endocytosis by coated pits whereas Au-I was internalized by uncoated micropinocytotic invaginations. Early in the endocytic pathway, the two ligands were segregated within separate small vesicles and tubulovesicles. Au-alpha 2MGMA was concentrated in a small number of these structures whereas Au-I was sparsely distributed among a relatively large number. Subsequently, the two endocytic pathways converged as the ligands intermingled within pale multivesicular bodies and lysosome-like structures. Au-I was less efficiently transferred to lysosomes than Au-alpha 2MGMA since a greater proportion of intracellular Au-I remained associated with small vesicles and tubulovesicles. This study indicates that early intracellular events in the endocytic pathways of insulin and alpha 2MGMA are distinct. These findings are discussed in light of the fundamentally dissimilar biological roles of these two molecules and the possible involvement of the endocytic pathway in the insulin signaling mechanism.

Cholesterol biosynthesis and its regulation in dissociated cell cultures of fetal rat brain: developmental changes and the role of 3-hydroxy-3-methylglutaryl coenzyme A reductase.

Primary cultures of cells dissociated from fetal rat brain were utilized to define the developmental changes in cholesterol biosynthesis and the role of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase in the regulation of these changes. Cerebral hemispheres of fetal rats of 15-16 days of gestation were dissociated mechanically into single cells and grown in the surface-adhering system. Cholesterol biosynthesis, studied as the rate of incorporation of [14C]acetate into digitonin-precipitable sterols, was shown to exhibit two distinct increases in synthetic rates, a prominent increase after 6 days in culture and a smaller one after 14 days in culture. Parallel measurements of HMG-CoA reductase activity also demonstrated two discrete increases in enzymatic activity, and the quantitative and temporal aspects of these increases were virtually identical to those for cholesterol synthesis. These data indicate that cholesterol biosynthesis undergoes prominent alterations with maturation and suggest that these alterations are mediated by changes in HMG-CoA reductase activity. The timing of the initial prominent peak in both cholesterol biosynthesis and HMG-CoA reductase activity at 6 days was found to be the same as the timing of the peak in DNA synthesis, determined as the rate of incorporation of [3H]thymidine into DNA. The second, smaller peak in reductase activity and sterol biosynthesis at 14 days occurred at the time of the most rapid rise in activity of the oligodendroglial enzyme, 2':3'-cyclic nucleotide 3'-phosphohydrolase (CNP). These latter observations suggest an intimate relationship of the sterol biosynthetic pathway with cellular proliferation and with oligodendroglial differentiation in developing mammalian brain.

Effects of gastric bubble implant on weight change with and without compliance with a behavior modification program.

The purpose of this prospective study was to evaluate the effect of compliance versus noncompliance with a behavior modification program (BMP) for 35 patients who underwent implantation of 51 Garren-Edwards gastric bubbles (GEGB). Criteria for morbid obesity was met by every patient. Two patient groups were formed by self-selection. Compliant patients (group I) attended 75% or more of the BMP. Group II consisted of noncompliant patients, and was divided further into partially complaint and totally noncompliant. A significant difference was found in both loss of weight and loss of body mass index (BMI) between the complaint and noncompliant groups, with no significant difference in weight loss between the partially and totally noncompliant groups. Patients' compliant or noncompliant behavior had a significant effect on changes in weight (p = 0.007) and body mass index (p = 0.005). Sixteen patients had two consecutive bubble implants. The rate of compliance was significantly higher during the first implant program, with a significantly greater loss in both weight and BMI for compliant patients. Follow-up data for 28 of 35 patients revealed a continued, albeit small, average weight decrease. In summary, weight loss does not result from the gastric bubble alone. Our study demonstrates that the key factor is compliance with a behavior modification program.

Age as a risk factor in colonoscopy: fact versus fiction.

Age has been assumed to be an independent risk factor in patients undergoing colonoscopy. However, this has never been specifically studied. We evaluated 302 patients 65 yr old or older, to determine the incidence of complications. We found that the risk is comparable to that of the heterogeneous patient population previously reported, except for a slightly increased incidence of delayed bleeding subsequent to polypectomy. In addition, the presence of cardiac, pulmonary, liver, or kidney disease was not found to be associated with an increased incidence of complications. Neither was there an increased risk of complications in patients undergoing colonoscopy, who had previously undergone abdominal or pelvic surgery. In summary, older patients experience the same risks as younger patients, except for post-polypectomy delayed bleeding, which may be related to atherosclerosis of the blood vessels, and appears to be more common in elderly patients.

Duodenal obstruction: a complication of percutaneous endoscopic gastrostomy tube migration.

Percutaneous endoscopic gastrostomy (PEG) is now a well-established technique for providing long-term enteral feeding. The procedure is being increasingly utilized and, although generally well tolerated, it may be associated with complications. We report two patients in whom a replacement Foley balloon tip catheter migrated into the small bowel and caused obstruction. The diagnosis, management, and preventive measures of tube migration are discussed.

Dysphagia as the presenting symptom of recurrent breast carcinoma.

Esophageal obstruction secondary to recurrent breast carcinoma is not widely recognized. Frequently, the esophageal narrowing is attributed to a benign process, resulting in delays of diagnosis and treatment. Such a case prompted this report and review of the literature.

Pancreatic carcinoma is associated with delayed gastric emptying.

Fifteen patients with histologically confirmed pancreatic carcinoma, without evidence of gastroduodenal invasion or obstruction, were prospectively studied to determine the frequency of gastric emptying disorders as determined by a solid-phase gastric emptying study. Nine of these (60%) had gastric emptying curves more than two standard deviations below normal mean values. The majority of patients did not have symptoms of gastric stasis. Nausea and/or vomiting was present in 33% of patients with abnormal gastric emptying and in none of those with normal emptying. Abdominal and/or back pain was present in 8/9 with delayed gastric emptying and in 3/6 with normal emptying. Disordered gastric emptying did not correlate with tumor stage, histology, location, or hyperbilirubinemia. Delayed solid-food gastric emptying may be responsible for the nonspecific abdominal complaints that occur during the course of pancreatic carcinoma, although more frequently, gastroparesis exists on a subclinical level.

Relation of cellular phospholipid composition to oligodendroglial differentiation in C-6 glial cells.

The relation of the polar head group composition of cellular phospholipids to a biochemical expression of oligodendroglial differentiation was studied in cultured C-6 glial cells. Induction of the oligodendroglial enzyme, 2',3'-cyclic nucleotide 3'-phosphohydrolase (CNP), was determined after alteration of the polar head group composition of phospholipids by exposure of the cells to choline analogues, especially N,N'-dimethylethanolamine. To accomplish the phospholipid alteration, cells were grown in the presence of the analogue in medium free of exogenous lipid, i.e., first for 24 h in 10% delipidated serum and then for 48 h in serum-free medium. The 48-h exposure to serum-free medium resulted in untreated C-6 cells in a several fold increase in CNP activity, but in cells treated with 2.5 mM N,N'-dimethylethanolamine, total inhibition of this induction was observed. A graded, concentration-dependent inhibitory effect of the analogue on the induction of CNP was defined. The effect of the analogue was relatively specific, e.g., the activity of another plasma membrane enzyme of C-6 cells, (Na+ + K+)-activated ATPase, was not affected. Morever, there was no evidence of a toxic effect of the analogue; thus, total protein synthesis and cell growth were not altered, and the induction of CNP in serum-free medium recurred after removal of the analogue. N,N'-Dimethylethanolamine was shown to be incorporated into cellular phospholipids, primarily at the expense of phosphatidylcholine. The data define an important role for the polar head group composition of membrane phospholipids in oligodendroglial differentiation in this model system.

Common tetrasaccharide epitope NeuAc alpha 2-->3Gal beta 1-->3(Neu-Ac alpha 2-->6)GalNAc, presented by different carrier glycosylceramides or O-linked peptides, is recognized by different antibodies and ligands having distinct specificities.

A novel globo-series disialoganglioside, disialosyl galactosyl globoside (Structure 1 below), defined by new monoclonal antibody (mAb) RM2, was isolated and characterized as having terminal structure identical to that of ganglio-series ganglioside GD1 alpha (Structure 2) and a common mucin-type epitope (Structure 3) widely distributed in glycoproteins such as glycophorin A. While these three structures share a common nonreducing tetrasaccharide terminus, mAb RM2 showed strong specific reactivity only with Structure 1, not with Structures 2 or 3. Another mAb, QSH2, reacted strongly with Structure 3 but did not cross-react with Structures 1 or 2. Conformational molecular models based on minimum energy hard sphere exoanomeric calculations suggest that Structure 1 presents a unique surface topology distinct from that of Structures 2 or 3. Our findings suggest the novel concept that reactivity of a common carbohydrate epitope with different antibodies or ligands is highly dependent on the type of carrier glycosylceramide or carrier O-linked peptide. [formula: see text]

Forceful balloon dilation: an outpatient procedure for achalasia.

The initial treatment of choice in patients with achalasia is balloon dilation. Heretofore, this procedure was performed on an in-hospital basis resulting in high patient cost. This study evaluated the safety and efficacy of pneumatic dilation as an outpatient procedure. Sixty-one procedures were performed on 50 patients at two centers. An overall treatment success rate of 95% (47 of 50 patients) was achieved. Two patients had elective surgical treatment and a third underwent surgery for perforation secondary to dilation. A total of three patients complained of post-procedure chest pain within 4 hours and were hospitalized. Two had perforations; one required surgical repair. The third patient had resolution of symptoms. We conclude that performing balloon dilation as an outpatient procedure is safe, efficacious, and cost effective.

The effects of morbid obesity and the Garren-Edwards gastric bubble on solid phase gastric emptying.

Rapid solid phase gastric emptying (SPGE) resulting in decreased satiety is postulated to be a contributing factor to obesity. Twenty-six morbidly obese patients with a weight range of 189-523 lb were entered into the Garren-Edwards gastric bubble program. SPGE studies were performed pre-bubble implantation and repeated within 2 wk after bubble implant. There was no significant difference between study patients pre-bubble insertion emptying time and the normal population curve. Emptying time was found to be significantly increased, post-bubble implant, compared with pre-bubble implant over five time periods of testing. The Garren-Edwards gastric bubble has been used as a method for inducing early satiety for the morbidly obese patient. It has been postulated that delay of SPGE would be associated with satiety. Our patients experienced satiety despite their more rapid SPGE. Thus, the satiety which was experienced by our subjects was not on the basis of delayed SPGE, as previously thought.

Oxygen desaturation and changes in breathing pattern in patients undergoing colonoscopy and gastroscopy.

The respiratory effect of diagnostic colonoscopy and upper endoscopy were studied in 32 elderly patients. Twenty-two underwent colonoscopy and 10 upper endoscopy. In the group undergoing upper endoscopy, 4 of 10 patients experienced a decrease in oxygen desaturation greater than or equal to 4% during the medication period; an additional 2 patients desaturated during the procedure. In the group undergoing colonoscopy, 12 of 22 patients experienced oxygen desaturation during the medication period; 3 other patients desaturated during the procedure. Mean SaO2 for each group was lowest (p less than 0.05) during the medication period. Central apneas occurred in 13 of the patients undergoing colonoscopy during the medication period; however, only 8 of these patients with apneas experienced desaturation greater than or equal to 4% and the periods of desaturation did not correlate with the periods of apneas. Oxygen desaturation greater than or equal to 4% occurs frequently during both upper endoscopy and colonoscopy in this elderly population. This is related to the effects of sedation; the procedure itself worsened the desaturation in only 16% of the patients. Furthermore, the desaturation did not correlate with changes in the breathing patterns of the patients. Low-flow oxygen and/or close monitoring of patients during and subsequent to administration of medication is advised.