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  • Critser, John K.  (6)
  • 1995-1999  (6)
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  • 1995-1999  (6)
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  • 1
    Online Resource
    Online Resource
    The Effects of Microencapsulation on Pancreatic Islet Osmotically Induced Volumetric Response
    SAGE Publications ; 1999
    In:  Cell Transplantation Vol. 8, No. 6 ( 1999-11), p. 699-708
    Details
    In: Cell Transplantation, SAGE Publications, Vol. 8, No. 6 ( 1999-11), p. 699-708
    Abstract: Microencapsulation of pancreatic islets has been proposed as a means to prevent allograft rejection and to protect islets during cryopreservation. The aim of this study was to investigate: 1) the effects of the cryoprotectants (CPAs) dimethyl sulfoxide (DMSO) and ethylene glycol (EG) on the volume of Ca 2+ alginate microcapsules, and 2) the effects of microencapsulation on the volumetric response of human and canine pancreatic islets during CPA equilibration. Stock sodium alginate with a high mannuronic acid content (HM) or a high guluronic acid content (HG) was used to generate empty capsules (mean diameter 200 μm) with an electrostatic generator. The capsules were held in place by a holding pipette system and videotaped during the addition of 2 or 3 M CPA at 22°C. Islets (isolated from human cadaveric donors and mongrel dogs and then cultured overnight at 37°C) were encapsulated in alginate (HM), loaded into a microperfusion chamber, and the change in islet volume was videotaped after exposure to the same CPAs and concentrations. These were compared to the volume responses of nonencapsulated islets. Images were analyzed using a computerized image analysis system and the data were analyzed using ANOVA. HG microcapsules showed a significant (p 〈 0.05) increase in volume following exposure to EG but not to DMSO. HM microcapsule volume did not change significantly following exposure to either EG or DMSO and was therefore chosen as the substrate for islet encapsulation. Free, nonencapsulated canine and human islets responded to the osmotic challenge of the 2 M DMSO by shrinking to 70.00 ± 1.04% (mean ± SEM) and 70.11 ± 1.05%, and in 2 M EG to 72.89 ± 1.93% and 69.33 ± 1.38%, respectively, of the isotonic volume before returning to the original cell volume. Exposure to 3 M DMSO or EG resulted in a further dehydration to 65.89 ± 0.91.% and 67.67 ± 1.91% for canine and 62.22 ± 0.66.% or 65.89 ± 1.30% for human islets. Minimum volumes were reached within 30–40 s after exposure to the cryoprotectant. Encapsulated human islets reached 86.88 ± 1.47% of their original volume in 2 M and 80.33 ±0.89% in 3 M DMSO, and 87.33 ± 1.86% in 2 M and 82.80 ± 1.57% in 3 M EG. This volume change was significantly less (p 〈 0.01) than that observed in corresponding free islets. Encapsulated canine islets reached 83.67 ± 2.13% of their original volume in 2 M and 78.22 ± 0.95% in 3 M DMSO, and 85.44 ± 1.92% in 2 M and 78.11 ± 2.01% in 3 M EG. As with human islets, this was significantly different than free islets (p 〈 0.01). These minimal volumes were reached within 30–50 s. These results demonstrate that there are cryoprotectant and alginate-specific interactions and that microencapsulation modulates the degree of osmotically induced shrinkage of islets. The development or modification of existing cryopreservation protocols to improve postcryopreservation recovery or function must account for these factors.
    Type of Medium: Online Resource
    ISSN: 0963-6897 , 1555-3892
    URL: Article
    DOI: 10.1177/096368979900800615
    Language: English
    Publisher: SAGE Publications
    Publication Date: 1999
    detail.hit.zdb_id: 2020466-8
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  • 2
    Online Resource
    Online Resource
    Osmotic Characteristics of Isolated Human and Canine Pancreatic Islets
    Elsevier BV ; 1997
    In:  Cryobiology Vol. 35, No. 2 ( 1997-09), p. 106-113
    Details
    In: Cryobiology, Elsevier BV, Vol. 35, No. 2 ( 1997-09), p. 106-113
    Type of Medium: Online Resource
    ISSN: 0011-2240
    URL: Article
    DOI: 10.1006/cryo.1997.2029
    Language: English
    Publisher: Elsevier BV
    Publication Date: 1997
    detail.hit.zdb_id: 1463192-1
    detail.hit.zdb_id: 80098-3
    SSG: 12
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  • 3
    Online Resource
    Online Resource
    Water and Cryoprotectant Permeability Characteristics of Isolated Human and Canine Pancreatic Islets
    SAGE Publications ; 1999
    In:  Cell Transplantation Vol. 8, No. 5 ( 1999-09), p. 549-559
    Details
    In: Cell Transplantation, SAGE Publications, Vol. 8, No. 5 ( 1999-09), p. 549-559
    Abstract: Cryopreservation allows accumulation of the necessary islet transplantable mass as well as adequate time for tissue typing and infectious disease screening. Cryopreservation protocols may be optimized by modeling the osmotically induced volume excursions that occur during the addition and removal of cryoprotective agents (CPAs). To that end, three transport parameters were measured at 22°C in canine and human islets isolated by collagenase digestion and euroficoll purification: (i) the apparent hydraulic conductivity (L p ), (ii) the permeability coefficient of the CPA (P s ), and (iii) the associated reflection coefficient (σ). The parameters were determined by volumetric analysis of islets upon abrupt exposure to 1, 2, and 3 M dimethyl sulfoxide (DMSO), ethylene glycol (EG), glycerol (GLY), and propylene glycol (PG). The parameters were calculated using the Kedem-Katchalsky theory to describe islet volume excursion kinetics (assuming islets to be single equivalent osmotic units with the same volume and surface area of the actual islet) and a three-parameter curve fit was performed using the Marquardt-Levenberg method. It was determined that the permeability characteristics of pancreatic islets are species specific, and based upon the measured parameters, the highest P s values for canine islets were observed following exposure to 2 M EG, and the highest P s values for human islets were observed following exposure to 2 M PG. The permeability parameters were analyzed adjusting for islet radius using ANCOVA procedures to acquire least square means. For canine islets exposed to 2 M EG these values were determined to be 0.936 μm/min/atm, 2.47 μm/s, and 0.90 (for L p , P s , and ϕ, respectively) and for human islets exposed to 2 M PG the values were determined to be 1.56 μm/min/atm, 3.48 μm/s, and 0.85 (for L p , P s , and σ, respectively). These parameters were used in a model to calculate osmotically induced islet volumetric response upon addition/dilution of the optimum CPAs, taking into consideration critical volume excursion limits at which irreversible damage occurs.
    Type of Medium: Online Resource
    ISSN: 0963-6897 , 1555-3892
    URL: Article
    DOI: 10.1177/096368979900800510
    Language: English
    Publisher: SAGE Publications
    Publication Date: 1999
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  • 4
    Online Resource
    Online Resource
    Osmotic Tolerance Limits of Canine Pancreatic Islets
    SAGE Publications ; 1999
    In:  Cell Transplantation Vol. 8, No. 3 ( 1999-05), p. 277-284
    Details
    In: Cell Transplantation, SAGE Publications, Vol. 8, No. 3 ( 1999-05), p. 277-284
    Abstract: Future improvements in the recovery and function of pancreatic islets following cryopreservation will require a more precise quantification of the stresses that occur at each stage of the cryopreservation protocol. Changes in solution osmolality during the addition and dilution of cryoprotectants and during freezing and thawing induce changes in islet volume that may exceed tolerable limits. The aim of this study was to determine the range of solution osmolalities that results in significant changes in islet function. Islets were isolated from canine pancreases by collagenase digestion and Euro-Ficoll purification. Following 12-h culture at 37°C, islets were counted and dispensed into multiwell plate inserts. Islet function was assessed in each well immediately before and 24 h following a 10-min osmotic challenge with hypo- or hyperosmotic solutions of PBS (0, 75, 150, 300, 600, 1200, or 2300 mOsm/kg) at 22°C. Canine islets reached their osmotic equilibrium within 10 min. Duplicate wells were used for each osmolality treatment for each of six donors (n = 12). No significant differences in basal or glucose-stimulated insulin secretion were found between wells prior to the osmotic challenge (3.35 ± 0.45 and 20.98 ± 3.36 μIU/IE/h, respectively). Following the osmotic challenge and 24-h in vitro tissue culture, a significant increase in basal secretion was observed for islets exposed to 0 and 75 mOsm/kg solutions and a significant decrease for islets exposed to 2300 mOsm/kg solution. Islets exposed to 0 and 2300 mOsm/kg solutions showed significant decreases in the stimulated insulin secretion when compared to controls. Solution osmolalities of 150–1200 mOsm/kg appear to be tolerated by canine islets with no significant deviations in insulin secretion. The corresponding tolerable volume range was 152.6 ± 6.8% to 60 ± 5.1% of the isotonic islet volume. The minimum critical volume was used in a theoretical analysis of the islet volumes that would result from equilibrium freezing in dimethyl sulfoxide (DMSO). The calculations show that 1.5 mol/l DMSO is sufficient to prevent damage to islets due to excessive shrinkage. Further refinement of cryoprotectant addition and dilution protocols, and cooling and warming protocols for canine islets, are now possible.
    Type of Medium: Online Resource
    ISSN: 0963-6897 , 1555-3892
    URL: Article
    DOI: 10.1177/096368979900800308
    Language: English
    Publisher: SAGE Publications
    Publication Date: 1999
    detail.hit.zdb_id: 2020466-8
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  • 5
    Online Resource
    Online Resource
    The Determination of Membrane Permeability Coefficients of Canine Pancreatic Islet Cells and Their Application to Islet Cryopreservation
    Elsevier BV ; 1997
    In:  Cryobiology Vol. 35, No. 1 ( 1997-08), p. 1-13
    Details
    In: Cryobiology, Elsevier BV, Vol. 35, No. 1 ( 1997-08), p. 1-13
    Type of Medium: Online Resource
    ISSN: 0011-2240
    URL: Article
    DOI: 10.1006/cryo.1997.2018
    Language: English
    Publisher: Elsevier BV
    Publication Date: 1997
    detail.hit.zdb_id: 1463192-1
    detail.hit.zdb_id: 80098-3
    SSG: 12
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  • 6
    Online Resource
    Online Resource
    Hypoosmotic Exposure of Canine Pancreatic Digest as a Means to Purify Islet Tissue
    SAGE Publications ; 1997
    In:  Cell Transplantation Vol. 6, No. 4 ( 1997-07), p. 423-428
    Details
    In: Cell Transplantation, SAGE Publications, Vol. 6, No. 4 ( 1997-07), p. 423-428
    Abstract: The development of more effective means to separate pancreatic islets from the unwanted exocrine tissue would greatly advance the field of clinical islet allotransplantation in the treatment of insulin-dependent diabetes mellitus. Recent experiments with hamster islets have demonstrated a selective destruction of dissociated single exocrine cells when exposed to hypotonic conditions. It was the aim of this study to extend these observations to the canine model with collagenase dissociated pancreatic tissue and to evaluate the treatment's effect on islet function. Pancreases from five mongrel dogs were digested using an automated protocol of intraductal delivery of collagenase, and gentle dissociation. Duplicate samples of pancreatic digest were removed for insulin and amylase determination prior to and immediately following exposure to 50 mOsm/kg salt solution for a period of 30, 60, or 300 s before returning the digest to isoosmotic conditions. The remaining digest was cultured for a period of 48 h at 37°C before the tissue was recombined, washed, and a third sample removed for insulin and amylase. In vitro viability was then assessed using a static incubation assay with insulin content measured using a double-antibody radioimmunoassay, and amylase was determined using a colorimetric assay system. No difference in the insulin or amylase levels between the experimental groups was observed immediately following the hypotonic exposure; however, a significant decrease in the amylase content was observed following the 48-h culture period in digest that had been hypoosmotically exposed for 60 or 300 s compared with the pretreatment group (2.83 ± 0.41 IU amylase/mg pancreas vs. 1.29 ± 0.21 and 0.83 ± 0.12, mean ± SEM, p 〈 0.05). Insulin content was also significantly reduced in the 300-s exposure group compared with nontreated controls (3.2 ± 0.6 mU insulin/mg pancreas vs. 2.0 ± 0.2). The insulin/amylase ratio (I/A), a measure of islet and exocrine content, was 1.1 ± 0.13 following pancreas dissociation and 1.34 ± 0.21 for control tissue cultured for 48 h. The I/A ratio increased following hypoosmotic exposure to 1.50 ± 0.31 for tissue exposed for 30 s, 1.77 ± 0.19 for 60-s exposure, and 2.54 ± 0.13 for tissue exposed for 300 s (p 〈 0.05, vs. pretreatment group). In vitro insulin secretion was equivalent with the exception of the tissue exposed for 300 s, which had an increased basal level of insulin resulting in a significantly decreased stimulation index (3.8 ± 0.5 vs. 8.1 ± 1.2 for the purified islet control group, p 〈 0.05). These results suggest that a brief hypotonic exposure to pancreatic digest can alter the insulin/amylase ratio; however, there is a functional impairment on subsequent islet function after a period of in vitro tissue culture.
    Type of Medium: Online Resource
    ISSN: 0963-6897 , 1555-3892
    URL: Article
    DOI: 10.1177/096368979700600409
    Language: English
    Publisher: SAGE Publications
    Publication Date: 1997
    detail.hit.zdb_id: 2020466-8
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