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The source of insulin released from insulin-releasing cell clusters (IRCCs) differentiated from embryonic stem cells remains unclear. Rajagopal et al. have suggested that IRCCs do not synthesize but secrete insulin that had been absorbed from media during the multistep protocol. We report here further data relevant to this controversy. No radioisotopic labeling of insulin was observed when IRCCs were incubated in a medium containing 35S-cysteine. Less than 1% of the extra-cellular stoichiometric C-peptide equivalent to insulin was secreted during glucose stimulation. However, intracellular immunostaining and immunogold labeling were both positive for C-peptide. Finally, a mass balance calculation showed that simple equilibration of IRCCs by Fickian diffusion from media accounted for at most 4% of secreted insulin. These findings and further analysis of the results of others suggest that the mechanism of insulin secretion by IRCCs is a combination of sequestration and de novo synthesis.

Effect of membrane composition and structure on solute removal and biocompatibility in hemodialysis. Significant changes in extracorporeal membranes have occurred over the past five decades in which hemodialysis (HD) has been available as a therapy for both acute renal failure (ARF) and end-stage renal disease (ESRD). For cellulosic membranes, these changes have included a reduction in thickness, hydroxyl group substitution, and an increase in pore size. These modifications have resulted in enhanced efficiency of small solute removal, a broader spectrum of overall solute removal, and an attenuation of complement activation in comparison to the thick, unsubstituted cellulosic membranes of low permeability used in the early days of HD therapy. Synthetic membranes, originally developed specifically for use in high-flux HD and hemofiltration, have also evolved during this same time period. In fact, the initially clear distinction between low-flux regenerated cellulosic and high-flux synthetic membranes has become blurred, as membrane formulators have developed products designed to appeal to enthusiasts for both membrane formats. The purpose of this review is to characterize both the solute removal and biocompatibility characteristics of dialysis membranes according to their composition (that is, polymeric makeup) and structure. In this regard, the manner in which membrane biocompatibility interacts with flux is highlighted.

Islet-like cell clusters (ILCCs) were derived from murine embryonic stem cells using a slightly modified version of the protocol originally described by Lumelsky et al. in 2001. Analysis with enzyme-linked immunosorbent assays (ELISAs) that distinguish human from murine insulin demonstrated that insulin released from these ILCCs, upon initial in vitro glucose challenge, was of non-murine origin and in fact corresponded to the species of insulin, human or bovine, that had been added to the culture media used to derive ILCCs. This finding convincingly supports the hypothesis that ILCCs are not synthesizing insulin de novo, but rather simply regurgitating insulin taken up during tissue culture. In further experiments, ILCCs were derived in media in which insulin had been replaced by IGF-I with which it shares a common signaling pathway. These ILCCs failed to release any detectable insulin. In contrast, ILCCs produced by various protocols stained positive (dithizone and immunoselective antibodies) for intracellular insulin and, in some cases, C-peptide. Despite the presence of at least some level of de novo, synthesized insulin in ILCCs, the majority of insulin released by ILCCs was sequestered from the exogenous medium.

Background Thrice-weekly haemofiltration affords excellent outcome when it is used to treat chronic renal failure patients. Daily haemofiltration (DHF) has recently been proposed as a more intensive therapy option, but the total ultrafiltration or exchange volume (replacement volume plus net ultrafiltration volume) requirements for adequate solute clearances during this novel therapy are unknown. Methods We calculated theoretical solute kinetic profiles during six times per week DHF for comparison with those during thrice-weekly haemodialysis using a high-flux dialyser (HFHD) or during continuous ambulatory peritoneal dialysis (CAPD). HFHD and CAPD were chosen for comparison because K/DOQI guidelines have defined adequate treatment doses for these therapies. Steady-state concentrations were calculated using a two-compartment model of an anuric patient with 35 l of total body water for five solutes: urea, creatinine, vitamin B(12), inulin and beta(2)-microglobulin. Solute distribution volumes and generation rates were taken from the literature, and excess fluid (1 l/day) was assumed to accumulate in and be removed from the extracellular fluid compartment. Theoretical predictions of solute clearance were compared for a 15-l exchange volume/session during DHF, urea Kt/V of 3.6/week during HFHD and urea Kt/V of 2.0/week during CAPD as solute-specific values of the equivalent renal clearance (EKR) and standard Kt/V (stdKt/V) recently defined by Gotch. Additional comparisons of solute clearances were performed between DHF and other daily therapies including six times per week short daily haemodialysis (SDHD) and six times per week nocturnal haemodialysis (NHD). Results The calculated results predict that: (i) urea clearance during DHF with an exchange volume of 90 l/week (6x15 l) is equivalent to those during HFHD and CAPD based on urea stdKt/V; and (ii) middle molecule clearances during DHF exceed those achieved during HFHD and CAPD based on either EKR or stdKt/V. As expected, DHF therapy was inferior regarding the clearance of urea and other small solutes to SDHD and NHD; however, DHF therapy was superior to SDHD regarding the clearance of larger middle molecules, approaching the clearances achieved by NHD. Conclusions We predict that an exchange volume of approximately 40% of total body water (15/35 l=43%) per session will provide adequate clearance of small solutes and substantial clearance of middle molecules during six times per week DHF therapy. These theoretical predictions require clinical validation.

This study relates to the diffusive transport characterization of hollow fibre membranes used in implantable bio-hybrid organs and other immunoisolatory devices. Techniques were developed to accurately determine the mass transfer coefficients for diffusing species in the 10(2)-10(5) MW range, validated and then used to study one membrane type known to effectively immunoisolate both allografts and xenografts in vivo. Low-molecular-weight diffusing markers included glucose, vitamin B12 and cytochrome C; higher-molecular-weight molecules were bovine serum albumin, immunoglobulin G, apoferritin and a range of fluorescein-tagged dextrans. Overall and fractional mass transfer coefficients through the hollow fibres were determined using a resistance-in-series model for transport. A flowing dialysis-type apparatus was used for the small-molecular-weight diffusants, whereas a static diffusion chamber was used for large-molecular-weight markers. For diffusion measurements of small-molecular-weight solutes, convective artefacts were minimized and the effect of boundary layers on both sides of the membrane were accounted for in the model. In measuring diffusion coefficients of large-molecular-weight species, boundary layer effects were shown to be negligible. Results showed that for small-molecular-weight species (13,000 MW) the diffusion coefficient in the membrane was reduced relative to diffusion in water by two to four times. The diffusion rate of large-molecular-weight species was hindered by several thousand-fold over their rate of diffusion in water.

Membrane technology is being increasingly applied to the collection of plasma by plasmapheresis, a procedure in which plasma is separated from whole blood, and the cellular components reinfused to the donor. Although still somewhat controversial, there is a general understanding of the physics and fluid mechanics involved in the transport phenomena associated with the microporous filtration of whole blood. The filtration process is governed by concentration polarization, with the red blood cell being the polarizing species. Shear-enhanced cellular diffusion has been proposed to explain how such a large cell can depolarize to allow the observed separation rates. The filtration rate per unit area is proportional to the first power of the shear rate and therefore, can remain approximately constant if the shear rate is increased to accommodate a decrease in surface area. Three automated membrane-based plasmapheresis systems are currently in use, primarily at fixed-site locations. They all collect 500-600 cc of plasma within 30-50 min. Two use hollow fiber membrane filter modules, while the third uses a rotating cylinder which relies on Couette rather than Poiseuille flow to generate the required shear rates at the membrane surface. Two portable systems have been developed; one centrifuge-based system requires house current, the other membrane system uses gravity, augmented by a battery power source. Both of the latter systems, due to their size and low weight are well suited for use in an environment where the equipment is moved on a frequent basis.