Improved Functional Survival of Human Islets of Langerhans in Three-Dimensional Matrix Culture

1994 ◽  
Vol 3 (5) ◽  
pp. 427-435 ◽  
Author(s):  
Mathias D. Brendel ◽  
Shen Shen Kong ◽  
Rodolfo Alejandro ◽  
Daniel H. Mintz
Keyword(s):  
Culture Media ◽  
Three Dimensional ◽  
Culture Method ◽  
Threshold Level ◽  
Insulin Dependent Diabetes Mellitus ◽  
Absolute Number ◽  
Human Islets ◽  
Dependent Diabetes Mellitus

The current study evaluates functional survival of human islets maintained in tissue culture for up to 4 wk in suspension media (CMRL-1066 with supplements) and contrasts these results with immobilizing three-dimensional matrices (agarose or alginate). The absolute number and volume of islets retrieved from agarose is significantly higher after two and four wk of culture compared to conventional free-floating media. In vitro function of islets, assessed by insulin/DNA content, insulin secretion into the culture media over 24 h and glucose-theophylline stimulated insulin release in a dynamic perifusion system, was not significantly different between free-floating and matrix preserved islets. In vivo islet function was evaluated by the effectiveness for reversal of insulin-dependent diabetes mellitus by transplantation of the islets under the kidney capsule of nude mice. Although adequate insulin responses to glucose were seen after culture in conventional or matrix media, only agarose embedded islets were consistently able to induce normoglycemia in diabetic recipients after 14 days of culture. Additional transplantation experiments defined the threshold level required to reverse diabetes to be between 1,000 and 1,500 agarose preserved islets. Our data suggest improved engraftment of human islets after agarose culture. This culture method may be of benefit for the accumulation of functionally competent human islets, thus facilitating the implementation of clinical protocols that utilize freshly isolated islets from multiple donors without the need for cryopreservation.

scholarly journals Genetic Predisposition of the Interleukin-6 Response to Inflammation: Implications for a Variety of Major Diseases?

2004 ◽  
Vol 50 (11) ◽  
pp. 2136-2140 ◽  
Author(s):  
Marie Bennermo ◽  
Claes Held ◽  
Sten Stemme ◽  
Carl-Göran Ericsson ◽  
Angela Silveira ◽  
...  
Keyword(s):  
Interleukin 6 ◽  
Promoter Region ◽  
Insulin Dependent Diabetes Mellitus ◽  
Juvenile Chronic Arthritis ◽  
In Vivo Studies ◽  
Dependent Diabetes Mellitus ◽  
Tnf Α

Abstract Background: A single-nucleotide polymorphism (SNP) in the promoter region of the interleukin-6 (IL-6) gene at position −174 (G>C) has been reported to be associated with a variety of major diseases, such as Alzheimer disease, atherosclerosis, and cardiovascular disease, cancer, non-insulin-dependent diabetes mellitus, osteoporosis, sepsis, and systemic-onset juvenile chronic arthritis. However, authors of previous in vitro and in vivo studies have reported conflicting results regarding the functionality of this polymorphism. We therefore aimed to clarify the role of the −174 SNP for the induction of IL-6 in vivo. Methods: We vaccinated 20 and 18 healthy individuals homozygous for the −174 C and G alleles, respectively, with 1 mL of Salmonella typhii vaccine. IL-1β, IL-6, and tumor necrosis factor-α (TNF-α) were measured in the blood at baseline and up to 24 h after vaccination. Results: Individuals with the G genotype had significantly higher plasma IL-6 values at 6, 8, and 10 h after vaccination than did individuals with the C genotype (P <0.005). There were no differences between the two genotypes regarding serum concentrations of IL-1β and TNF-α before or after vaccination. Conclusions: The −174 G>C SNP in the promoter region of the IL-6 gene is functional in vivo with an increased inflammatory response associated with the G allele. Considering the central role of IL-6 in a variety of major diseases, the present finding might be of major relevance.

Hyperglycemic athymic nude mice: factors affecting in vitro insulin secretion

1992 ◽  
Vol 263 (6) ◽  
pp. E1131-E1133
Author(s):  
A. Zeidler ◽  
P. Edwards ◽  
J. Goldman ◽  
S. Kort ◽  
W. P. Meehan ◽  
...  
Keyword(s):  
Animal Model ◽  
Insulin Secretion ◽  
Insulin Dependent Diabetes Mellitus ◽  
Dependent Diabetes Mellitus ◽  
Factors Affecting ◽  
In Vitro Insulin Secretion ◽  
Insulin Secretion In Vitro ◽  
And Control

The strain of athymic nude male mice (ANM) developed at the University of Southern California (USC) exhibits spontaneous hyperglycemia and relative hypoinsulinemia in vivo. To investigate factors that influence insulin secretion in this animal model of non-insulin-dependent diabetes mellitus, we utilized the isolated perfused mouse pancreas of the ANM-USC and control BALB/c mice. We compared in vitro glucose-induced insulin secretion in ANM-USC and control mice, inhibition of secretion by somatostatin, and variability of insulin secretion over the two-year period it took to complete these experiments. Glucose-induced insulin secretion from the isolated pancreas was biphasic in both ANM-USC and controls. Insulin secretion was quantitatively equal to or greater than control mice, depending on the phase of secretion analyzed and the source of the control mice. In contrast to pancreases of control mice, insulin secretion from ANM-USC pancreases was relatively resistant to inhibition of insulin secretion by somatostatin. Variability in insulin secretion over the two years in which these experiments were performed was greater from pancreases of control than that observed from pancreases of the ANM-USC. The hyperglycemic ANM-USC mouse does not demonstrate diminished insulin secretion in vitro yet is relatively hypoinsulinemic in vivo. Thus circulating factors other than somatostatin might contribute to the insulinopenic stage in this animal model.

99 DIFFERENTIAL GENE REGULATION OF STEROIDOGENIC TRANSCRIPTS AND ESTRADIOL PRODUCTION FOLLOWING IN VITRO PIG EMBRYO ELONGATION IN ALGINATE HYDROGEL THREE-DIMENSIONAL MATRIX

2012 ◽  
Vol 24 (1) ◽  
pp. 162
Author(s):  
J. R. Miles ◽  
C. N. Sargus ◽  
S. A. Plautz ◽  
J. L. Vallet ◽  
A. K. Pannier
Keyword(s):  
Equal Opportunity ◽  
Culture Media ◽  
Morphological Changes ◽  
Three Dimensional ◽  
Differential Gene Regulation ◽  
Alginate Hydrogels ◽  
The Media ◽  
And Control

Between Day 10 and 12 of gestation, the pig embryo elongates from a sphere to a long thin, filament. During this time, the embryo increases the production of oestrogen via an increase in steroidogenic transcripts, which is critical for maternal recognition of pregnancy. To date, attempts to elongate porcine embryos in vitro have been unsuccessful. Therefore, the objective of this study was to utilise alginate hydrogels to establish a culture system that promotes in vitro embryo elongation with a corresponding increase in steroidogenic transcripts and oestradiol production. In 3 replicate collections, White crossbred gilts (n = 15) were bred at Day 0 of the oestrous cycle. At Day 9 of gestation, reproductive tracts were collected and flushed with RPMI-1640 containing antibiotics. Embryos were recovered, grouped according to size and washed with RPMI-1640 containing antibiotics and 10% fetal bovine serum (FBS). Embryos were randomly assigned to be encapsulated using a double encapsulation technique (0.7% sodium alginate and 1.5% calcium chloride solution) or used as controls. Encapsulated and control embryos were cultured for 96 h in CO2 -pretreated RPMI-1640 containing antibiotics and 10% FBS at 38°C, 5% CO2 in air and 100% humidity. Every 24 h, the embryos were imaged and half of the media was replaced. The removed media was stored at –20°C and used to assess oestradiol levels by radioimmunoassay. At the end of culture, a subset of encapsulated and control embryos were snap frozen and used to assess the expression level of steroidogenic transcripts (STAR, CYP11 and CYP19) using quantitative PCR. All data were analysed using general linear model (GLM) procedures for ANOVA. Cell survival, assessed by blastocyst fragmentation and confirmed by live/dead staining in representative embryos, was greater (P = 0.01) for encapsulated embryos (60.1 ± 4.8%) compared with controls (33.3 ± 4.8%). Of encapsulated embryos, 27% had some morphological change (minor flattening and tubal formation) and 14% had significant morphological changes (considerable flattening and tubal formation elongating through the gel), consistent with in vivo embryo elongation. In contrast, the control embryos had no morphological changes observed and remained spherical during culture. The expression levels of STAR, CYP11 and CYP19 were significantly (P < 0.05) greater in encapsulated embryos compared with control embryos. Furthermore, a significant (P < 0.01) time-dependent increase in oestradiol levels in the culture media of encapsulated embryos was identified compared with controls and culture media alone. These results illustrate that cultured pig embryos encapsulated in alginate hydrogels undergo limited morphological changes with increased expression of steroidogenic transcripts and oestrogen production. †USDA is an equal opportunity provider and employer.

scholarly journals Multifunctionalized hydrogels foster hNSC maturation in 3D cultures and neural regeneration in spinal cord injuries

2019 ◽  
Vol 116 (15) ◽  
pp. 7483-7492 ◽  
Author(s):  
Amanda Marchini ◽  
Andrea Raspa ◽  
Raffaele Pugliese ◽  
Marina Abd El Malek ◽  
Valentina Pastori ◽  
...  
Keyword(s):  
Spinal Cord ◽  
Spinal Cord Injuries ◽  
Culture Media ◽  
Three Dimensional ◽  
Cholinergic Neurons ◽  
Good Manufacturing Practice ◽  
Neural Regeneration ◽  
Neuronal Markers

Three-dimensional cell cultures are leading the way to the fabrication of tissue-like constructs useful to developmental biology and pharmaceutical screenings. However, their reproducibility and translational potential have been limited by biomaterial and culture media compositions, as well as cellular sources. We developed a construct comprising synthetic multifunctionalized hydrogels, serum-free media, and densely seeded good manufacturing practice protocol-grade human neural stem cells (hNSC). We tracked hNSC proliferation, differentiation, and maturation into GABAergic, glutamatergic, and cholinergic neurons, showing entangled electrically active neural networks. The neuroregenerative potential of the “engineered tissue” was assessed in spinal cord injuries, where hNSC-derived progenitors and predifferentiated hNSC progeny, embedded in multifunctionalized hydrogels, were implanted. All implants decreased astrogliosis and lowered the immune response, but scaffolds with predifferentiated hNSCs showed higher percentages of neuronal markers, better hNSC engraftment, and improved behavioral recovery. Our hNSC-construct enables the formation of 3D functional neuronal networks in vitro, allowing novel strategies for hNSC therapies in vivo.

Metabolic effects of vanadyl sulfate in humans with non—insulin-dependent diabetes mellitus: In vivo and in vitro studies

Metabolism ◽  
2000 ◽  
Vol 49 (3) ◽  
pp. 400-410 ◽  
Author(s):  
Allison B. Goldfine ◽  
Mary-Elizabeth Patti ◽  
Lubna Zuberi ◽  
Barry J. Goldstein ◽  
Raeann LeBlanc ◽  
...  
Keyword(s):  
Diabetes Mellitus ◽  
Insulin Dependent Diabetes Mellitus ◽  
In Vitro Studies ◽  
Insulin Dependent Diabetes ◽  
Vanadyl Sulfate ◽  
Metabolic Effects ◽  
Dependent Diabetes Mellitus ◽  
Insulin Dependent

scholarly journals ETHYLCELLULOSE FLOATING MICROSPHERES OF ANTIDIABETIC AGENT: IN VITRO AND IN VIVO EVALUATION

2016 ◽  
Vol 9 (1) ◽  
pp. 44 ◽  
Author(s):  
Seema Kohli ◽  
Megha Sharma ◽  
Abhisek Pal
Keyword(s):  
Average Particle Size ◽  
Insulin Dependent Diabetes Mellitus ◽  
Treated Group ◽  
Antidiabetic Activity ◽  
Dependent Diabetes Mellitus ◽  
Average Particle ◽  
In Vivo Evaluation ◽  
Gastro Retentive

Objective: To develop and evaluate floating type gastro-retentive dosage form, appropriate for controlled release of repaglinide (RG) having a narrow therapeutic window.Methods: Repaglinide loaded microspheres (MS) using biological macromolecule ethylcellulose (EC) was prepared by a solvent diffusion-evaporation technique using polyvinyl alcohol (PVA) emulsifier. Compatibility of drug and polymer was studied by Fourier-transform infrared spectroscopy (FTIR). During formulation, various process optimisation parameters studied were stirring speed, the concentration of drug, polymer and emulsifier. Characterization and in vitro evaluation was performed. In vivo antidiabetic activity was performed on alloxan induced diabetic rats followed by histopathological screening.Results: The average particle size was in the range of 174-243 µm. Yield, entrapment and buoyancy of microspheres were 68.4­­-79.8, 58.6-73.1 and 71.8-84.1% respectively. 65.1% release of drug from optimised formulation was obtained which follows first-order kinetics (r2 = 0.989). Optimised formulation treated group shows significant (p<0.01) decrease in glucose level of blood as compared to pure drug treated group during the later hours of study with satisfactory results of histology.Conclusion: The investigation revealed the promising potential of gastro retentive microspheres for delivering RG for the treatment of non-insulin dependent diabetes mellitus (NIDDM).

Repeated intraperitoneal injections of interleukin 1β induce glucose intolerance in normal rats

1991 ◽  
Vol 124 (5) ◽  
pp. 470-478 ◽  
Author(s):  
Lise Wogensen ◽  
Jesper Reimers ◽  
Thomas Mandrup-Poulsen ◽  
Jørn Nerup
Keyword(s):  
Blood Glucose ◽  
Glucose Tolerance ◽  
Single Injection ◽  
Interleukin 1 ◽  
Insulin Dependent Diabetes Mellitus ◽  
Dependent Diabetes Mellitus ◽  
Mild Depression ◽  
Wistar Kyoto

Abstract. Previous in vitro findings suggest the involvement of interleukin 1 (IL-1) in the pathogenesis of insulin-dependent diabetes mellitus. The aims of the present study were to investigate the effects of single or repeated ip injections of recombinant IL-1β on blood glucose and glucose tolerance in vivo. Normal Wistar Kyoto rats were injected ip with a single injection of 4 μg/kg of the mature form of recombinant IL-1β (amino acids 117-269) or once daily on 5 consecutive days. Control rats were given vehicle and were fed ad libitum or pair-fed together with the rIL-1β treated rats. An ip glucose tolerance test (0.2 g D-glucose/100 g) was performed 2 h after injection of rIL-1β. A single injection of rIL-1β caused a mild depression in blood glucose and an improved glucose tolerance. Multiple injections of rIL-1 β induced a diminished weight gain, a 24-28% reduction in food intake, a lasting mild depression of blood glucose (7 days) and a transiently impaired glucose tolerance on day 5. We conclude that systemic IL-1 should be considered an important regulator of glucose homeostasis in vivo.

scholarly journals In vitro 3D Spheroid Culture Developed on the Parafilm Surface Using HEK-293 Cells

2020 ◽  
Vol 3 (1) ◽  
pp. 220-227
Author(s):  
Erdal Eroğlu
Keyword(s):  
Three Dimensional ◽  
Culture Method ◽  
In Vivo Studies ◽  
Preclinical Research ◽  
Hek 293 ◽  
Hek 293 Cells ◽  
293 Cells ◽  
Hanging Drop Culture

Preclinical research to predict the effects of drugs and chemicals on humans is commonly carried out either by cell culture studies in vitro condition or on animals in vivo condition. While drug studies tested on cells cultured as a monolayer in plastic flasks are incompatible with realistic results, falsifying findings can also be achieved from in vivo studies performed on different species. In recent years, research on drug tests using spheroid cultures formed by growing cells in three-dimensional (3D) in vitro has attracted great interest. 3D spheroid structures are formed by growing the cells in a drop suspended on superhydrophobic surfaces. In this study, HEK-293 cells were investigated on parafilm surfaces displaying superhydrophobic properties by growing in 2 &amp;micro;l volume using hanging drop culture method in terms of spheroid formation. Light microscopy images from spheroid structures were taken on different incubation days and the area of spheroids was measured using the ImageJ program. Our study holds important findings for a chip platform that can be developed for use in vitro drug tests.

Insulin and glucagon release of human islets in vitro: effects of chronic exposure to glucagon

1997 ◽  
Vol 152 (2) ◽  
pp. 239-243 ◽  
Author(s):  
F Bertuzzi ◽  
C Berra ◽  
C Socci ◽  
A M Davalli ◽  
G Pozza ◽  
...  
Keyword(s):  
Chronic Exposure ◽  
Insulin Response ◽  
Insulin Dependent Diabetes Mellitus ◽  
Human Islets ◽  
Dependent Diabetes Mellitus ◽  
Glucagon Release ◽  
Insulin Dependent ◽  
Insulin Response To Glucose ◽  
In Vitro Effects

Abstract Hyperglucagonemia is commonly found in insulin-dependent as well as in non-insulin-dependent diabetes mellitus, and is likely to be caused by absolute or relative insulin deficiency. The aim of the present study was to evaluate whether a chronic glucagon exposure (1·0 μm for 4 h) modifies the insulin response to acute stimuli with glucagon (1·0 μm), arginine (10·0 mm) and glucose (16·7 mm), or the glucagon response to arginine and glucose, in human islets. Chronic exposure to glucagon did not affect the insulin response to glucose and arginine, but inhibited its response to glucagon (44·6 ± 9·3 vs 168·6 ± 52·3 pg/islet per 20 min, P<0·05); the latter effect was not observed when exposure to glucagon was discontinuous (2·0 μm glucagon alternated with control medium for 30 min periods). The chronic exposure to glucagon also reduced the glucagon response to arginine (−4·9 ± 5·7 vs 19·9 ± 7·9 pg/islet per 20 min, P<0·05) without affecting the inhibition of glucagon release exerted by glucose. These data indicate that chronic exposure to glucagon desensitizes pancreatic α and β cells in response to selected stimuli. Journal of Endocrinology (1997) 152, 239–243

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