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Biomedical subjects

C K Colton

Publications and source records attributed to C K Colton.

At least 91 records · Page 5Linked to original sources
PubMed1972

Ultrafiltration of lipoproteins through a synthetic membrane. Implications for the filtration theory of atherogenesis.

To investigate the interaction of lipoproteins with semipermeable membranes, solutions of low density lipoproteins (LDL), very low density lipoproteins (VLDL), mixtures of the two, and diluted, normal, and hyperlipidemic serum were ultrafiltered through a synthetic membrane (500 A nominal pore diameter) using a stirred laboratory ultrafiltration cell. The pressure dependence of ultrafiltrate flux showed that a concentrated layer of lipoproteins was built up at the membrane surface (concentration polarization) and that VLDL was more subject to polarization than LDL. This phenomenon controlled the observed lipoprotein transport behavior. Whereas true membrane rejection (the fraction of the solute on the membrane surface which does not pass through the membrane) was greater than 0.95 for both LDL and VLDL, observed solute rejection varied from nearly 0 to 1.0, depending upon experimental conditions. If concentration polarization occurs in the arterial system, these results suggest that lipoprotein transport into arterial wall may be influenced not only by arterial blood pressure and the properties of the arterial wall, but also by local hemodynamic conditions and by the relative as well as absolute magnitudes of LDL and VLDL concentration.

Arteriosclerosis↗
PubMedDate not supplied

Microcinematographic studies of flow patterns in the excised rabbit aorta and its major branches.

Arterial fluid mechanics may play a role as a localizing factor for early atherosclerosis. Flow patterns in natural rabbit aortas rendered transparent were studied using a microcinematographic visualization technique. The aortic arch exhibited a single cell of clockwise-rotating helical secondary flow along the ventral and inner walls. Flow separation occurred proximal to the two arch branches with flow reversal proximal to the brachiocephalic artery. Sinusoidal flow rendered the helical motion more pronounced in systole, while the reverse flow zone periodically expanded and contracted. Steady flow in the abdominal aorta revealed streamlines which follow slow looping trajectories lateral to ostia before tracing helical paths into the branches. Flow separation was present along the dorsal wall of the aorta opposite the superior mesenteric artery. With the exception of the left renal artery, steady flow wall shear stresses were higher distal to ostia than proximal. Spatial gradients of wall shear stress were larger around branches than elsewhere. Similar to observed flow patterns, sites of enhanced macromolecular permeability, as observed previously in the normal rabbit aorta, follow a clockwise helical pattern in the arch and exhibit a distribution around ostia that correlates to some degree with regions of elevated shear stress gradients.

Animals↗
PubMedDate not supplied

Elution conditions and degradation mechanisms in long-term immunoadsorbent use.

The limited life of immunoadsorbents used for the large-scale purification of biological macromolecules poses a significant limitation to the more widespread application of this technology. In this study, the binding activity of a monoclonal antibody (MAb) to bovine serum albumin (BSA) was measured as a function of pH, ionic strength, and varying concentrations of KSCN, ethylene glycol, or DMSO. Low pH (2.5) and 3 M KSCN each reduced the antibody binding constant below 6 x 10(5) L/mol, meeting criteria derived from a simple chromatographic model for identifying effective eluents. A panel of six MAb to BSA was exposed repeatedly to adsorption conditions and the two eluents. Four MAb lost less than 50% of their initial binding capacity over 100 cycles. The other two lost 75% of their initial capacity. One MAb was stable when exposed to low pH but lost binding capacity with KSCN. In all cases, the equilibrium constant was unchanged. The loss of capacity was also shown to be a strong function of antibody loading: at 14.5 mg/mL, 98% of the initial binding capacity of one MAb was lost within 40 cycles, versus 75% loss at 1 mg/mL. Antibody leakage and nonspecific adsorption of contaminants were not responsible for significant loss of antibody activity over time.

Animals↗
PubMedDate not supplied

A microperifusion system with environmental control for studying insulin secretion by pancreatic tissue.

A continuous flow reactor (perifusion system) was fabricated and tested for measuring the kinetics of insulin secretion from isolated pancreatic islets of Langerhans in response to step changes in the glucose concentration and oxygen partial pressure in the perfusate flowing around the islets. The system was capable of making rapid changes in perfusate glucose concentration and pO2, had rapid dynamic response for measuring the change in insulin secretion rate as a result of these changes in perfusate, and was suitable for studying very small volumes of tissue. Initial experiments with this system demonstrated that (1) the response of isolated rat islets to glucose stimulation was very fast, with the first phase peak occurring in as little as about 10 s, (2) bulk perfusate oxygen partial pressure levels of 30 mmHg or less reduced the second-phase insulin secretion rate in graded fashion, (3) the reduction in secretion rate began within 1 min following an oxygen partial pressure decrease, and (4) the reduction in secretion rate was reversible, with a burst of insulin secretion occurring during the first minute after partial pressure restoration.

Animals↗
PubMedDate not supplied

Effect of oxygen on isolated pancreatic tissue.

One approach to insulin replacement therapy is transplantation of islets of Langerhans immunoisolated from host tissue by a semipermeable membrane. In this state, islets depend on diffusion of nutrients and wastes to and from the beta-cell to provide a suitable environment for survival and secretion. A perifusion system was constructed to test glucose-stimulated (100-300 mg/100 ml) insulin secretion from whole islets, or small (5-10 cell) aggregates, under controlled pO2. First phase insulin secretion from adult rat islets was unaffected by hypoxic levels of pO2, but second phase secretion was rapidly reduced at pO2 levels below 60 mmHg in the bulk media. Secretion from single-cell aggregates was unaffected until pO2 levels dropped to 12 mmHg, at which point secretion progressively decreased with falling pO2. A theoretical reaction/diffusion model was developed to correlate intraislet pO2 with reduced insulin secretion. Oxygen limited secretion was reversible, and not a result of decreased cell viability, as ascertained by both long-term static culture and trypan blue staining. Insulin secretion is more sensitive to hypoxia than is cell viability, in part because O2 uptake increases with glucose stimulation. These results indicate that O2 may be the limiting factor in the ability of immunoisolated islets to respond to blood glucose changes. We conclude that maintenance of a sufficiently high islet pO2 for maximal insulin secretion may be an important issue for graft design and implant site selection.

Culture Techniques↗
PubMedDate not supplied

Transport of 125I-albumin across normal and deendothelialized rabbit thoracic aorta in vivo.

Transmural concentration profiles of 125I-albumin in vivo were measured across the normal and balloon catheter-deendothelialized rabbit descending thoracic aorta as a function of time following intravenous injection. A tracer was injected 5 or 60 minutes after deendothelialization, and the animals were sacrificed after circulation times of 10, 30 or 60 minutes. The aorta was immediately excised and frozen flat between glass slides. Samples were serially sectioned parallel to the intimal surface in a refrigerated microtome, washed with trichloroacetic acid (TCA), and counted. Relative tissue concentration profiles of TCA-precipitable radioactivity from the media of control animals showed entry from both luminal and adventitial sides, as previously found with conscious normal rabbits, but spatial gradients at both luminal and medial-adventitial borders were less steep. Relative concentration levels in ballooned animals were 10- to 40-fold higher than in controls, and the profiles were flatter. Uptake rates at equivalent circulation times were greater in experiments initiated 60 minutes, as compared with 5 minutes, after deendothelialization, suggesting that progressive medial edema may have occurred following balloon injury. These results show that the intact endothelium is the dominant mass transfer resistance for 125I-albumin transport across the aortic wall. The data also suggest that the incomplete monolayer of platelets adherent to the subendothelium after balloon deendothelialization is not a substantial resistance to transport, as compared to that of the media, and that convection plays a more important role than diffusion for 125I-albumin transport across the deendothelialized aortic wall.

Animals↗
PubMedDate not supplied

Local variation in arterial wall permeability to low density lipoprotein in normal rabbit aorta.

Normal rabbits were injected intravenously with horseradish peroxidase (HRP) and 125I-labeled human low density lipoprotein (LDL), and the aortas were perfusion-fixed. Subsequent visualization of HRP in the aortas was produced by reaction of the tissue with diaminobenzidine and hydrogen peroxide. The luminal surface of the aortas showed many small punctate foci of brown reaction product to the HRP, which represented penetration of the HRP into the vessel wall. The foci were scattered over the luminal surface, and most of the focal areas were less than 1 mm in diameter. The concentration of LDL was up to 47 times greater in these focal areas than in surrounding noncolored regions not showing increased permeability to HRP. Small circumscribed foci of heightened permeability to LDL may predispose to the local accumulation of lipid and ultimately to the formation of atherosclerotic plaques.

Animals↗
PubMedDate not supplied

Improved vascularization of planar membrane diffusion devices following continuous infusion of vascular endothelial growth factor.

Improving blood vessel formation around an immunobarrier device should improve the survival of the encapsulated tissue. In the present study we investigated the formation of new blood vessels around a planar membrane diffusion device (the Baxter Theracyte System) undergoing a continuous infusion of vascular endothelial growth factor through the membranes and into the surrounding tissue. Each device (20 microl) had both an inner immunoisolation membrane and an outer vascularizing membrane. Human recombinant vascular endothelial growth factor-165 was infused at 100 ng/day (low dose: n = 6) and 500 ng/day (high dose: n = 7) for 10 days into devices implanted s.c. in Sprague-Dawley rats; noninfused devices transplanted for an identical period were used as controls (n = 5). Two days following the termination of VEGF infusion, devices were loaded with 20 microl of Lispro insulin (1 U/kg) and the kinetics of insulin release from the lumen of the device was assessed. Devices were then explanted and the number of blood vessels (capillary and noncapillary) was quantified using morphometry. High-dose vascular endothelial growth factor infusion resulted in two- to threefold more blood vessels around the device than that obtained with the noninfused devices and devices infused with low-dose vascular endothelial growth factor. This increase in the number of blood vessels was accompanied by a modest increase in insulin diffusion from the device in the high-dose vascular endothelial growth factor infusion group. We conclude that vascular endothelial growth factor can be used to improve blood vessel formation adjacent to planar membrane diffusion devices.

Animals↗
PubMedDate not supplied

Implantable biohybrid artificial organs.

Biohybrid artificial organs encompass all devices which substitute for an organ or tissue function and incorporate both synthetic materials and living cells. This review concerns implantable immunoisolation devices in which the tissue is protected from immune rejection by enclosure within a semipermeable membrane. Two critical areas are discussed in detail: (i) Device design and performance as it relates to maintenance of cell viability and function. Attention is focussed on oxygen supply limitation and how it is affected by tissue density and the development of materials that induce neovascularization at the host tissue-membrane interface; and (ii) Protection from immune rejection. Our current knowledge of the mechanisms that may be operative in immune rejection in the presence of a semipermeable membrane barrier is limited. Nonetheless, recent studies shed light on the role played by membrane properties in preventing immune rejection, and many studies demonstrate substantial progress towards clinically useful implantable immunoisolation devices.

Animals↗
PubMedDate not supplied

Number and volume of islets transplanted in immunobarrier devices.

Immunobarrier devices may prevent immune destruction of transplanted islets, but there are concerns about survival within such devices. Islets were transplanted in diffusion chambers that employed two laminated polytetrafluoroethylene membranes held together with titanium rings. Five hundred syngeneic mouse islets placed in devices were transplanted into the epididymal fat pads of streptozotocin (STZ) diabetic mice (B6AF1). After 2 wk the devices were removed. Sections were made parallel to the membrane surface. Eight to 13 systematically selected sections of each device were analyzed by planimetry to determine the area of the device space and of the islets within that space. From these data we estimated total volume of the device, volume of islets, and number of islets in a device. The data were segregated into two groups: group I (blood glucose less than 100 mg/dL 2 wk after implantation), and group II (over 150 mg/dL). The volume (mean +/- SE) of devices implanted for 2 wk was 2.1 +/- 0.4 microL in group I and 2.2 +/- 0.2 microL in group II. The islet volume and number within devices were 0.30 +/- 0.06 and 0.17 +/- 0.01 microL, or 340 +/- 50 and 230 +/- 20 islets in group I and group II, respectively. The volume of fibrous tissue in devices was about 0.50 microL. About 10% of the islet tissue had central necrosis. The beta cell volume in a membrane device needed for cure is comparable to that required with islets under the kidney capsule (0.25-0.80 microL). The mass of islets contained within membrane devices needed to cure diabetes is equivalent to that of a graft in an optimal transplant site such as under the kidney capsule.

Animals↗
PubMedDate not supplied

Use of a physiologic pharmacokinetic model of glucose homeostasis for assessment of performance requirements for improved insulin therapies.

Methods are presented for assessing insulin therapies using a physiologic pharmacokinetic model of glucose homeostasis in man. The model is composed of simultaneous differential equations that represent physiologic compartments and spaces in which glucose and insulin are distributed and undergo metabolic reactions. The model is used to simulate clinical experiments in which blood glucose concentration is controlled by artificial device therapies. Predictions of the theoretical model for responses of normal and diabetic individuals to standard intravenous and oral glucose tolerance tests are compared to clinical data. Reasonable agreement is obtained between predictions of the computer simulations and clinical data for normal individuals. The responses of a diabetic person to oral glucose tolerance tests are simulated by removal of the pancreas from the glucose homeostasis model and introduction of insulin into the model by a prescribed therapy. Model simulations reaffirm expectations concerning the poor blood glucose control attainable by intramuscular insulin injection. Simulations of blood glucose regulation by an artificial pancreas using closed-loop feedback control for controlling insulin delivery rate reveal hyperinsulinemia that results in a net shift in the deposition of a glucose load from liver to peripheral tissues. Simulations of this system in which the time delay for glucose measurement is varied from 1.5 to 30 min show that increases in sensor delay result in progressive loss in glucose regulation, exacerbation of hyperinsulinemia, and increased insulin requirements.

Blood Glucose↗
PubMedDate not supplied

Measurement of glucose concentration in the presence of coreactants with a platinum electrode.

An implantable glucose sensor is being developed that is based on the use of a high-area platinum electrode. The sensor is operated in a controlled potential mode, in which the potential of the platinum working electrode versus an unpolarized reference electrode is periodically varied according to a preselected voltage-time regimen. The change in potential is accompanied by a flow of current between the platinum working electrode and a counter-electrode. This current serves to periodically rejuvenate the electrode surface and to provide a signal that is proportional to the glucose concentration. A method for analyzing this signal has been developed, the compensated net charge (CNC) method, that involves integration of the current over one complete potential cycle. This method significantly improves both the sensitivity and selectivity for determining glucose in the presence of the normal physiologic coreactants. For a solution containing glucose, amino acids, and urea, a change in glucose concentration from 50 to 150 mg/dl gives about a 50% change in the net charge. A change in the urea concentration from 20 to 40 mg/dl has no effect on the net charge. A change in the amino acids concentration from 35 to 65 mg/dl has little effect on the net charge above a glucose concentration of about 100 mg/dl. Negligible effects on the net charge have also been found for creatinine (0-1.5 mg/dl) and uric acid (0-4 mg/dl).

Amino Acids↗
PubMedDate not supplied

In vitro and in vivo testing of an electrocatalytic glucose sensor.

A prerequisite for the development of an implantable artificial pancreas is the availability of a stable, long-life glucose sensor. Platinum (Pt) catalyzed electrodes have been demonstrated in vitro to show high sensitivity to glucose and long cycle life but are more sensitive to co-reactants compared with enzymatic methods. The authors developed a special data processing method (compensated net charge ratio, or CNCR) in which the measured electrode response is very sensitive to glucose, completely insensitive to urea, and only moderately sensitive to amino acids. Other endogenous and exogenous co-reactants show only minor interferences. The CNCR method involves the determination of the ratio of net oxidation charge to total charge during one complete cycle of a cyclic voltammogram. Prototype electrodes tested in vitro in spiked plasma have shown typical sensitivities of greater than 2 x 10(-4) CNCR units per 1 mg/dl change in glucose concentration, with linear response up to 400 mg/dl. For in vivo testing, a modified 5 F vascular catheter with membrane covered surface mounted electrodes was used at a vena cava site in swine. Several sensor designs were tested in vivo, with sensitivities of 1-5 x 10(-4) CNCR units (mg/dl).

Animals↗