The incorporation of water-soluble gel matrix into bile acid-based microcapsules for the delivery of viable ß-cells of the pancreas, in diabetes treatment: biocompatibility and functionality studies.

In recent studies, we microencapsulated pancreatic β-cells using sodium alginate (SA) and poly-L-ornithine (PLO) and the bile acid, ursodeoxycholic acid (UDCA), and tested the morphology and cell viability post-microencapsulation. Cell viability was low probably due to limited strength of the microcapsules. This study aimed to assess a β-cell delivery system which consists of UDCA-based microcapsules incorporated with water-soluble gel matrix. The polyelectrolytes, water-soluble gel (WSG), polystyrenic sulphate (PSS), PLO and polyallylamine (PAA) at ratios 4:1:1:2.5 with or without 4 % UDCA, were incorporated into our microcapsules, and cell viability, metabolic profile, cell functionality, insulin production, levels of inflammation, microcapsule morphology, cellular distribution, UDCA partitioning, biocompatibility, thermal and chemical stabilities and the microencapsulation efficiency were examined. The incorporation of UDCA with PSS, PAA and WSG enhanced cell viability per microcapsule (p < 0.05), cellular metabolic profile (p < 0.01) and insulin production (p < 0.01); reduced the inflammatory release TNF-α (p < 0.01), INF-gamma (p < 0.01) and interleukin-6 (IL-6) (p < 0.01); and ceased the production of IL-1β. UDCA, PSS, PAA and WSG addition did not change the microencapsulation efficiency and resulted in biocompatible microcapsules. Our designed microcapsules showed good morphology and desirable insulin production, cell functionality and reduced inflammatory profile suggesting potential applications in diabetes

Primary Bile Acid Chenodeoxycholic Acid-Based Microcapsules to Examine ß-cell Survival and the Inflammatory Response

In past studies using hydrogel-polyelectrolyte matrix and different bile acid excipients, we microencapsulated pancreatic ß-cells using various methods, and the microcapsules were mechanically stable, displayed good morphological characteristics with good physico-chemical compatibility but had limited cell viability and poor cell survival. Using bile acids, cell survival increased but overall remained limited. Thus, this study aimed to test different microencapsulating methods and examine the effects of the primary hydrophobic bile acid, chenodeoxycholic acid (CDCA), on ß-cell microcapsules, in terms of morphology and cell function. Using the polymer sodium alginate (SA) and the co-polymer poly-l-ornithine (PLO), in 10:1 ratio, two microcapsules were made, one without CDCA and one with CDCA. During the microencapsulation process, polymer flow rate and culture media flow rate were screened (0.1–1.5 mL/min) for most uniform microcapsule. Pancreatic ß-cells (NIT-1) were microencapsulated and tested for morphology, formulation physico-chemical compatibility, stability, surface topography and chemical composition. Encapsulated cell viability, metabolism, respiration, bioenergetics, biological activity and the inflammatory profile were also measured. A polymer flow rate of 0.8 mL/min accompanied by 0.6 mL/min media flow rate were found to produce the most uniform microcapsules using 10:1 formulation ratio. The microcapsules showed poor cell viability which was improved significantly after CDCA incorporation. CDCA also enhanced insulin secretion (p &lt; 0.01), metabolism, respiration and bioenergetics (p &lt; 0.01) and significantly reduced the inflammatory response. These benefits were attained without compromising microcapsule size or stability. A polymer flow rate of 0.8 mL/min and a media flow rate of 0.6 mL/min produced good microcapsules when using SA and PLO in 10:1 ratio, and the incorporation of the primary bile acid, chenodeoxycholic acid, enhanced microcapsule stability and significantly increased cell survival and reduced inflammation which suggests potential applications in ß-cell microencapsulation and transplantation

Increasing creatine kinase activity protects against hypoxia / reoxygenation injury but not against anthracycline toxicity in vitro

The creatine kinase (CK) phosphagen system is fundamental to cellular energy homeostasis. Cardiomyocytes express three CK isoforms, namely the mitochondrial sarcomeric CKMT2 and the cytoplasmic CKM and CKB. We hypothesized that augmenting CK in vitro would preserve cell viability and function and sought to determine efficacy of the various isoforms. The open reading frame of each isoform was cloned into pcDNA3.1, followed by transfection and stable selection in human embryonic kidney cells (HEK293). CKMT2- CKM- and CKB-HEK293 cells had increased protein and total CK activity compared to non-transfected cells. Overexpressing any of the three CK isoforms reduced cell death in response to 18h hypoxia at 1% O2 followed by 2h re-oxygenation as assayed using propidium iodide: by 33% in CKMT2, 47% in CKM and 58% in CKB compared to non-transfected cells (P<0.05). Loading cells with creatine did not modify cell survival. Transient expression of CK isoforms in HL-1 cardiac cells elevated isoenzyme activity, but only CKMT2 over-expression protected against hypoxia (0.1% for 24h) and reoxygenation demonstrating 25% less cell death compared to non-transfected control (P<0.01). The same cells were not protected from doxorubicin toxicity (250nM for 48h), in contrast to the positive control. These findings support increased CK activity as protection against ischaemia-reperfusion injury, in particular, protection via CKMT2 in a cardiac-relevant cell line, which merits further investigation in vivo