Parathyroid hormone (PTH) stimulates osteoblasts to produce the proinflammatory cytokine interleukin-6 (IL-6), causing bone resorption. In patients with primary hyperparathyroidism, elevated serum levels of IL-6 normalize after resection of parathyroid tumours. Because IL-6 is also expressed in normal parathyroids and in other endocrine cells (adrenal and islet), we hypothesized that parathyroid tumours might contribute directly to the elevated serum IL-6 levels in patients with hyperparathyroidism. Immunohistochemistry identified IL-6, PTH, and chromogranin-A (an endocrine and neuroendocrine tumour marker) in normal, adenomatous and hyperplastic parathyroids. Using immunofluorescence and confocal microscopy, IL-6 co-localized with PTH and with chromogranin-A in parathyroid cells. All cultured parathyroid tumours secreted IL-6 at levels markedly higher than optimally stimulated peripheral blood mononuclear cells. Supernates from cultured parathyroids stimulated proliferation of an IL-6-dependent cell line, and anti-IL-6 MoAb abolished this stimulatory effect. IL-6 mRNA was documented in cultured parathyroid tumours, cultured normal parathyroids, fresh operative parathyroid tumours and fresh operative normal specimens. In conclusion, these data show that parathyroid tumours and normal parathyroids contain, produce and secrete IL-6. Our findings present a novel pathway by which human parathyroids may contribute markedly to IL-6 production and elevation of serum IL-6 levels in patients with hyperparathyroidism. The physiological relevance of IL-6 production by human parathyroids remains to be determined, but IL-6 secretion by parathyroid tumours may contribute to bone loss and to other multi-system complaints observed in these patients.
Background: If alginate microcapsules are to be used clinically for therapeutic cell transplants, capsule formulations must be designed to enhance optimal biocompatibility and immune acceptance.
Methods: Microcapsules were generated using highly purified, endotoxin-free, ultra-low viscosity, high mannuronic acid alginate. The capsules differed with respect to gelling cation (50 mM barium or 100 mM calcium), alginate concentration (2.0% or 3.3%), alginate density (homogeneous or inhomogeneous), and the presence or absence poly-L-lysine (PLL) coating. Four types of empty capsules were implanted intraperitoneally (i.p.) in normal NOD mice, and their biocompatibility was evaluated after various time periods in vivo. Encapsulated adult porcine islets (APIs) were transplanted i.p. in diabetic NOD mice, and immune acceptance was evaluated by graft survival times, host cell adherence to capsule surfaces, and flow cytometric analysis of peritoneal host cells.
Results: All empty alginate capsules were biocompatible in vivo, but barium-gelled alginate capsules without PLL were clearly the most biocompatible, since 99% of these empty capsules had no host cell adherence up to 9 months in vivo. In diabetic NOD mice, APIs functioned significantly longer in barium-alginate capsules without PLL than in calcium-alginate capsules with PLL and had strikingly less host cell adherence, although large numbers of host cells (predominantly macrophages and eosinophils) infiltrated the peritoneal cavities of recipients with APIs in both types of capsules. Addition of PLL coatings to barium-alginate capsules dramatically decreased graft survival.
Conclusions: Inhomogeneous barium-gelled alginate capsules without PLL are the optimal candidates for clinical trials, based on their enhanced biocompatibility and immune acceptance in vivo.
Background: Adult porcine islets (APIs) constitute a promising alternative to human islets in treating type 1 diabetes. The intrahepatic site has been used in preclinical primate studies of API xenografts; however, an estimated two-thirds of donor islets are destroyed after intraportal infusion due to a number of factors, including the instant blood-mediated inflammatory reaction (IBMIR), immunosuppressant toxicity, and poor reestablishment of extracellular matrix connections. Intraperitoneal (ip) transplantation of non-vascularized encapsulated islets offers several advantages over intrahepatic transplantation of free islets, including avoidance of IBMIR, immunopro tection, accommodation of a larger graft volume, and reduced risk of hemorrhage. However, there exists evidence that the peritoneal site is hypoxic, which likely impedes islet function. Methods: We tested the effect of hypoxia (2%-5% oxygen or pO 2 : 15.2-38.0 mm Hg) on free and encapsulated APIs over a period of 6 days in culture. Free and encapsulated APIs under normoxia served as controls. Islet viability was evaluated with a viability/cytotoxicity assay using calcein AM and ethidium bromide on days 1, 3, and 6 of culture. Alamar blue assay was used to measure the metabolic activity on days 1 and 6. Insulin in spent medium was assayed by ELISA on days 1 and 6. Results: Viability staining indicated that free islet clusters lost their integrity and underwent severe necrosis under hypoxia; encapsulated islets remained intact, even when they began to undergo necrosis. Under hypoxia, the metabolic activity and insulin secretion (normalized to metabolic activity) of both free and encapsulated islets decreased relative to islets cultured under normoxic conditions. Conclusions: Hypoxia (2%-5% oxygen or pO 2 : 15.2-38.0 mm Hg) affects the viability, metabolic activity, and insulin secretion of both free and encapsulated APIs over a six-day culture period. Encapsulation augments islet integrity under hypoxia, but it does not prevent loss of viability, metabolic activity, or insulin secretion.
Background
Stem cells for cardiac repair have shown promise in preclinical trials, but lower than expected retention, viability, and efficacy. Encapsulation is one potential strategy to increase viable cell retention while facilitating paracrine effects.
Methods and Results
Human mesenchymal stem cells (hMSC) were encapsulated in alginate and attached to the heart with a hydrogel patch in a rat myocardial infarction (MI) model. Cells were tracked using bioluminescence (BLI) and cardiac function measured by transthoracic echocardiography (TTE) and cardiac magnetic resonance imaging (CMR). Microvasculature was quantified using von Willebrand factor staining and scar measured by Masson's Trichrome. Post‐MI ejection fraction by CMR was greatly improved in encapsulated hMSC‐treated animals (MI: 34±3%, MI+Gel: 35±3%, MI+Gel+hMSC: 39±2%, MI+Gel+encapsulated hMSC: 56±1%; n=4 per group; P<0.01). Data represent mean±SEM. By TTE, encapsulated hMSC‐treated animals had improved fractional shortening. Longitudinal BLI showed greatest hMSC retention when the cells were encapsulated (P<0.05). Scar size at 28 days was significantly reduced in encapsulated hMSC‐treated animals (MI: 12±1%, n=8; MI+Gel: 14±2%, n=7; MI+Gel+hMSC: 14±1%, n=7; MI+Gel+encapsulated hMSC: 7±1%, n=6; P<0.05). There was a large increase in microvascular density in the peri‐infarct area (MI: 121±10, n=7; MI+Gel: 153±26, n=5; MI+Gel+hMSC: 198±18, n=7; MI+Gel+encapsulated hMSC: 828±56 vessels/mm2, n=6; P<0.01).
Conclusions
Alginate encapsulation improved retention of hMSCs and facilitated paracrine effects such as increased peri‐infarct microvasculature and decreased scar. Encapsulation of MSCs improved cardiac function post‐MI and represents a new, translatable strategy for optimization of regenerative therapies for cardiovascular diseases.
Keywords: angiogenesis, cardiovascular diseases, heart failure, ischemia, myocardial infarction