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Biomedical subjects

T M Chang

Publications and source records attributed to T M Chang.

At least 127 records · Page 7Linked to original sources
PubMed1993

Polydisperse dextran as a diffusing test solute to study the membrane permeability of alginate polylysine microcapsules.

Applications of alginate polylysine (APL) microcapsules in cell culture engineering and hybrid artificial organs require strict control of membrane permeability. For example, hybrid artificial organs must permit the diffusion of smaller molecules including peptides and proteins and the exclusion of leucocytes and immunoglobulins (MW > 150,000). Single molecular weight solutes such as proteins have been used to study the molecular weight cut-off of the membrane. A new approach that uses a heterogenous mixture of dextrans of different molecular weights: -MW 10,000-500,000 as a test solute in diffusion experiments is described. Intra/extra capsular changes in concentration and molecular weight distribution of the heterogenous dextran mixture are monitored and determined by high performance gel chromatography. MW can then be related to diffusional Stokes radius, a more useful parameter when comparing permeability of cell membrane to different test solutes.

Alginates↗
PubMed1993

Cross-linked hemoglobin-superoxide dismutase-catalase scavenges oxygen-derived free radicals and prevents methemoglobin formation and iron release.

In this study, we prepared PolyHb-SOD-catalase (intermolecularly cross-linked hemoglobin, superoxide dismutase (SOD), and catalase). We found that PolyHb-SOD-catalase is effective in scavenging oxygen-derived free radicals. In the xanthine/xanthine oxidase system, the initial rate of cytochrome c reduction was 2.13 +/- 0.26 nmoles cyt. c/min for PolyHb alone. PolyHb- SOD-catalase reduced this to 0.56 +/- 0.08 nmoles cyt. c/min because of its ability to eliminate superoxide (O2-). Addition of PolyHb to 200 microM of hydrogen peroxide (H2O2), changed the H2O2 level slightly to 192 +/- 0.4 microM. Addition of PolyHb-SOD-catalase, on the other hand, lower the level to 41 +/- 0.3 microM. Results also show that both effects were dependent on the concentration of SOD-catalase cross-linked with hemoglobin. Oxidative challenge with H2O2 resulted in minimal changes in the absorbance spectra of PolyHb-SOD-catalase. With PolyHb, there were spectral changes reflecting the formation of methemoglobin and heme degradation. Furthermore, the amount of iron released, after incubation with 250 microM H2O2, was 6.8 +/- 1.8 micrograms/dl for PolyHb-SOD-catalase and 76.6 +/- 1.0 micrograms/dl for PolyHb. These results show that cross-linked SOD-catalase prevents oxidative reactions involving the hemoglobin component of PolyHb-SOD-catalase.

Catalase↗
PubMed1993

Genetically engineered E. coli cells containing K. aerogenes gene, microencapsulated in artificial cells for urea and ammonia removal.

Microencapsulated genetically engineered E. coli cells can efficiently remove urea without any increase in the ammonia levels in the medium. A 100 mg. alginate encapsulated bacteria rapidly reduces urea in a 100 ml. solution. The original urea concentration 100.00 +/- 1.00 mg./dl. fell to 1.55 +/- 0.13 mg./dl. in 30 minutes. There was no increase in the ammonia in the reaction medium. Extrapolated results shows that urea depletion capacity of encapsulated bacteria is sufficient to remove urea during kidney failure. Using single pool model, 40 gm. of encapsulated genetically engineered E. coli can lower urea (100 mg./dl.) in 40 litres of the body water to 1.60 mg./dl. within 30 minutes. Also, 40.00 gm. bacteria can lower ammonia (758.00 microM/l), in 40 litres of body water, to 90.42 microM/l in 20 minutes. Further studies will be required for multi-compartmental models in the physiological conditions.

Ammonia↗
PubMed1993

Use of finger-prick human blood samples as a more convenient way for in-vitro screening of modified hemoglobin blood substitutes for complement activation: a preliminary report.

Safety test of modified hemoglobin in animals does not always reflect safety in human. We have earlier reported an in-vitro preclinical screening test based on in-vitro complement activation of human plasma In this test, modified hemoglobin is added to human plasma in a test tube. Complement activation is followed by the C3a levels. Since this directly measures the effect of modified hemoglobin on human plasma, this could be an important bridge before clinical use in patients. The use of plasma is suitable for research, development and industrial applications. However, there are many extra steps involved and may not be convenient for population or patient screening on a large scale bases. The present study shows that it is possible to use small sample of whole blood obtained directly from finger pricks and use immediately for analysing complement activation using an ELISA enzyme immunoassay method.

Blood Specimen Collection↗
PubMed1993

Mass isolation of highly purified canine islets using an automated method and Histopaque Ficoll gradients.

Eighteen pancreata from adult mongrel dogs were used for the study of islet isolation. The pancreas was distended with collagenase in Hanks' solution. The automated screen method and Histopaque Ficoll gradients were used to isolate and purify the canine islets. In vitro, the viability of isolated islets was assessed by both histology and perifusion studies. In vivo, the islet function was evaluated by using a nude mice xenograft model. Fair to good isolation and purification was found in 12 experiments. Before and after purification, the isolated islet count was 4767.1 +/- 560.1 and 3637.7 +/- 333.4 islet equivalence (I.E.)/gm pancreatic tissue. The purity was above 90%. Aldehyde Fuchsin stain disclosed islets with copious beta granules. The stimulation index of islets responding to 16.7 mM glucose plus 1 mM 3-isobutyl-1-methylxanthine (IBMX) versus 1.67 mM glucose was 12.93 +/- 4.75. Normoglycemia was restored and maintained for up to 2 weeks in 7 of 10 and up to 3 weeks in 5 of 10 diabetic nude mice transplanted with canine islets. In conclusion, the automated screen method and Histopaque Ficoll gradients afford a good yield of highly purified canine islets, and functional viability was verified both in vitro and in vivo. This will be an ideal model for isolation of human islets.

Animals↗
PubMed1993

Secretion of erythropoietin from microencapsulated rat kidney cells: preliminary results.

Rat kidney epithelial cells were microencapsulated within alginate-poly(L)lysine-alginate membrane. The microencapsulated cells were incubated using a culture media containing cobalt and another without cobalt. The viability was measured by trypan blue exclusion test. Secretion of erythropoietin (EPO) was measured by radioimmunoassay (RIA). Viability of free cells was 53%. The viability of microencapsulated cells increased to 72% after 12 days of incubation and remained at this level. Samples of the culture media were collected every 2 days for RIA. Samples within the microcapsules were collected by breaking the microcapsules open. RIA of these samples showed the following for the media containing cobalt. Between day 16 and day 32 the concentrations of EPO were 5.3 mU/ml inside and 18.3 mU/ml outside the microcapsule. The medium from the same number of free cells contained 21.2 mU/ml of EPO. Culture media without cobalt collected during the same period contained 1.8 mU/ml inside and 9.9 mU/ml outside the microcapsules. The free cell culture with this media during the same period contained 8.3 mU/ml.

Alginates↗
PubMed1992

Artificial cells: 35 years.

The first artificial cells were prepared 35 years ago. They contain biologically active materials. They are now being used in medicine and biotechnology. Artificial cells containing adsorbents are already a routine form of treatment in hemoperfusion. This includes treatment for acute poisoning, high blood aluminum and iron, kidney failure, some types of acute liver failure, and other conditions. Artificial cells are being tested for use as red blood cell substitutes. Artificial cells containing cell culture are being tested in animals for the treatment of diabetes, liver failure, and others. Artificial cells containing enzymes are being tested for treatment in hereditary enzyme deficiency diseases and other diseases. Artificial cells containing complex enzyme system can convert wastes like urea and ammonia into useful amino acids. In biotechnology, artificial cells are being used for the production of monoclonal antibodies, interferons, and other biotechnological products. They are also being investigated for use in other applications in biotechnology, chemical engineering, and medicine.

Animals↗
PubMed1992

Artificial liver support based on artificial cells with emphasis on encapsulated hepatocytes.

Artificial liver support requires more than a detoxification system. We have investigated additional approaches. Microencapsulated hepatocytes increased the survival time of fulminant hepatic failure (FHF) rats. They also lowered the bilirubin in Gunn rats. Xenograft of microencapsulated rat hepatocytes into mice are immunoisolated. The viability of hepatocyte increased from 62 to 100% after 29 days. This is because of accumulation of a hepatic stimulatory factor (> 100,000 D) secreted by the hepatocytes in the artificial cells. A novel two-step method of cell encapsulation greatly improved immunoisolation and biocompatibility. Other metabolic approaches included a multienzyme system for conversion of ammonia to essential amino acid, and removal of bilirubin.

Animals↗
PubMed1992

In vitro study of multicellular hepatocyte spheroids formed in microcapsules.

To make highly differentiated encapsulated hepatocytes, we formed hepatocyte spheroids in microcapsules in cultures. Hepatocytes isolated from rats were encapsulated in alginate-poly-L-lysine artificial cells and cultured under different medium conditions. When high molecular weight poly-L-lysine was used to make the capsule membrane, the hepatocytes aggregated and formed spheroids inside microcapsules within 48 hs. We studied the viability and function of encapsulated hepatocyte spheroids; free hepatocyte spheroids; nonaggregated encapsulated hepatocytes; and free hepatocyte monolayers. Cell viability and protein-producing ability were monitored for up to 30 days. Hepatocyte spheroids in the encapsulated or free form remained viable and continued to secrete proteins throughout the 30 days of observation. The viability and function of nonaggregated hepatocytes in the encapsulated or free form declined rapidly. These results suggest that the tridimensional structure of the spheroids may be important in maintaining the viability and function of encapsulated hepatocytes.

Animals↗
PubMed1992

Identification of a transcriptional enhancer important for enteroendocrine and pancreatic islet cell-specific expression of the secretin gene.

It is well established that the gene encoding the hormone secretin is expressed in a specific enteroendocrine cell, the S cell. We now show that the secretin gene is transiently expressed in insulin-producing B cells of the developing pancreatic islets in addition to the intestine. Furthermore, secretin is produced by most established islet cell lines. In order to identify and characterize the regulatory elements within the secretin gene that control tissue-specific expression, we have introduced secretin reporter gene constructions into the secretin-producing HIT and STC-1 cell lines as well as the nonexpressing INR1-G9 glucagonoma line. Analysis of deletion mutants revealed that sequences between 174 and 53 bp upstream from the transcriptional start site are required for maximal expression in secretin-producing cells. This positive element functioned independently of position and orientation. Further deletions into the enhancer resulted in a stepwise loss of transcriptional activity, suggesting the presence of several discrete control elements. The sequence CAGCTG within the secretin enhancer closely resembles that of the core of the B-cell-specific enhancer in the insulin gene. Point mutations introduced into this putative element led to greater than 85% reduction in transcriptional activity. Gel mobility shift assays suggested that a factor in B cells closely related or identical to proteins that bind to the insulin enhancer interacts with the CAGCTG motif in the secretin gene.

Animals↗
PubMed1992

Release of cholecystokinin and secretin by sodium oleate in dogs: molecular form and bioactivity.

The release of cholecystokinin (CCK) and secretin into both circulation and duodenal lumen, after intraduodenal perfusion with sodium oleate or oral ingestion of fat, was studied in anesthetized and conscious dogs, respectively. Intraduodenal infusion with sodium oleate (4 mmol.kg.-1.h-1, pH 9.5) in anesthetized dogs with diversion of bile and pancreatic juice stimulated the release of both CCK and secretin not only into the circulation but also into the duodenal lumen. The concentration of CCK and secretin in the luminal perfusate increased from 0.2 +/- 0.1 to 2.1 +/- 0.4 nM and 0.34 +/- 0.16 to 2.59 +/- 0.63 nM, respectively. Intraduodenal infusion of NaHCO3 solution at pH 9.5 did not result in release of either hormone. Luminal release of both hormones was also observed by intraduodenal infusion of sodium oleate in the dogs without diversion of bile and pancreatic juice, albeit at lower concentrations than those released in the dogs with diversion. Analysis of the molecular form of luminal secretin by gel filtration, ion-exchange chromatography, and high-performance liquid chromatography showed only a single form of secretin with molecular size, hydrophobicity, and charge similar to those of natural porcine secretin. In contrast, multiple forms of CCK were released into both circulation and duodenal lumen with CCK-58 as the predominant form. In conscious dogs, CCK-58 was also found to be the predominant form of CCK released into the circulation after oral ingestion of fat.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Neurohormonal mechanism of pancreatic exocrine secretion stimulated by sodium oleate and L-tryptophan in dogs.

In the present investigation, we have studied the effect of atropine on the pancreatic secretion stimulated by intraduodenal administration of either sodium oleate or exogenous cholecystokinin (CCK). In four dogs prepared with gastric and Thomas duodenal cannulas, pancreatic juice was collected for measurement of volume, bicarbonate, and protein output, and peripheral venous blood samples were obtained for radioimmunoassay of both secretin and CCK. Volume, bicarbonate, and protein output of the pancreatic juice increased significantly in response to sodium oleate (1-4 mmol/h) in a dose-dependent manner. The increase in pancreatic secretion paralleled the increments in both plasma CCK and secretin. Atropine given intravenously suppressed completely both pancreatic secretion and release of CCK stimulated by sodium oleate, whereas the release of secretin was not affected. Pancreatic secretion was significantly increased in a dose-dependent manner by exogenous CCK octapeptide (CCK-8) at 16, 32, and 64 micrograms (14, 28, and 56 pmol).kg-1.h-1. Atropine inhibited protein output only partially, but it did not influence bicarbonate output. In five additional dogs, the effect of atropine on L-tryptophan-stimulated pancreatic secretion was studied. Interestingly, atropine failed to influence the CCK release and pancreatic secretion of volume and bicarbonate, except for protein secretion, which was significantly inhibited. It was shown previously that atropine inhibited significantly the pancreatic secretion of bicarbonate stimulated by secretin in physiological doses. Thus we conclude that the inhibition by atropine of the pancreatic exocrine secretion stimulated by sodium oleate is mediated by both suppression of CCK release and inhibition of action of secretin on the exocrine pancreas.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Artificial cells in immobilization biotechnology.

Artificial cells contain biologically active materials. Artificial cells containing adsorbents have been a routine form of treatment in hemoperfusion for patients. This includes acute poisoning, high blood aluminum and iron, and supplement to dialysis in kidney failure. Artificial cells are being tested for use as red blood cell substitutes. Artificial cells encapsulated cell culture are being tested in animals for the treatment of diabetes and liver failure. A novel 2 step method has prevented xenograft rejection. Artificial cells containing enzymes are being studied for treatment in hereditary enzyme deficiency diseases and other diseases. Recent demonstration of extensive enterorecirculation of amino acids in the intestine has allowed its oral administration to deplete specific amino acids. Artificial cells containing complex enzyme system convert wastes like urea and ammonia into essential amino acids. Artificial cell is being used for the production of monoclonal antibodies, interferons and other biotechnological products. It is also being investigated for drug delivery, and for use in other applications in biotechnology, chemical engineering and medicine.

Biocompatible Materials↗
PubMed1992

Purification and characterization of liposomes encapsulating hemoglobin as potential blood substitutes.

In view of the desirability to increase the survival time of the liposome-based artificial red blood cells in vivo, the variables influencing optimum hemoglobin capture and preservation for the bovine hemoglobin-loaded liposomes (LEHb) are investigated. In order to predict the in vivo response, the necessary experiments for the in vitro system characterization have been carried out. The liposomes are prepared by the Reverse Phase Evaporation technique and then purified using a Sepharose 4B column. The purified LEHbs display a unimodal size distribution in the submicron range with a volume average diameter of 0.115 microns and a particle count of 1.25* 10(15) per ml of suspension. Analysis of the lipi/Hb content of the liposomes reveals that the variations in the ratio of Hb encapsulated to lipid entrapped (Hb/L)f as a function of the initial Hb concentration ([Hb]o) is insignificant compared to the net augmentation of (Hb/L)f as a function of the increasing initial lipid to Hb loading ([L]o). Meanwhile high [Hb]o s are necessary for the preservation of oxyhemoglobin.

Animals↗
PubMed1992

Blood substitutes based on modified hemoglobin prepared by encapsulation or crosslinking: an overview.

Modified hemoglobin consists of (1) encapsulated hemoglobin and (2) crosslinked hemoglobin (polyhemoglobin, intramolecularly cross-linked hemoglobin and conjugated hemoglobin). There have been new advances in all types of modified hemoglobins. Modified hemoglobins are effective in hemorrhagic shock. However, it is important to define hemorrhagic shock models and experimental designs. Important progress has been made in research on vasoactivities, organ perfusion, organ preservation, biodistribution, hematology, complement activation immunology and other areas. A preclinical screening test may bridge the gap between animal safety studies and injection into human. Potential new sources of hemoglobin included bovine hemoglobin, recombinant human hemoglobin and synthetic heme.

Animals↗