In previous experiments, we found that the threonine requirement of neonatal piglets fed parenterally was 40% of that when fed intragastrically; we hypothesized that much of the oral supply of threonine is being used for mucin production. To investigate this hypothesis, intragastrically fed 2-day-old piglets were fed one of three treatments for 8 days: 1) a threonine-adequate diet (IG-A; 0.6 g threonine.kg(-1).day(-1) fed intragastrically); 2) a threonine-deficient diet (IG-D; 0.1 g threonine.kg(-1).day(-1) fed intragastrically); or 3) a threonine-deficient diet with adequate threonine delivered parenterally (IV-A; 0.5 g threonine.kg(-1).day(-1) fed parenterally plus 0.1 g threonine.kg(-1).day(-1) fed intragastrically). IG-D piglets experienced higher nitrogen excretion, higher plasma urea, and lower plasma threonine concentrations versus both of the other groups (P < 0.05), indicating profound threonine deficiency. Mucosal mass and total crude mucin content were lower in the colons of IG-D pigs (P < 0.05). Histopathological analysis showed lower numbers of acidic mucin-producing goblet cells in the duodenum and ileum of IG-D pigs. In IG-D pigs, acidic mucin subtypes were lower in the small intestine but higher in the colon, which corresponded with persistent diarrhea. The parenteral supply of threonine was adequate to maintain most outcome parameters, although IV-A pigs did have smaller colonic goblet cells with more acidic mucins compared with IG-A pigs. Overall, our results suggest that adequate dietary threonine was critical in the production of mucus and that a parenteral threonine supply can ameliorate most of the symptoms of oral threonine deficiency.

Evidence is accumulating that the amino acid requirements for neonates receiving total parenteral nutrition (TPN) are significantly different than those for oral feeding and need to be determined. The parenteral threonine requirement was determined in 3-d-old male Yorkshire piglets (n = 25) by examining the effect of varying dietary threonine intakes [0.05-0.6 g/(kg.d)] on phenylalanine oxidation. The diet included adequate energy, total amino acids and phenylalanine, with excess tyrosine. Phenylalanine kinetics were determined from a primed, continuous intravenous infusion of L-[1-14C]phenylalanine. Phenylalanine oxidation, estimated from the rate of 14CO2 released in expired air during isotope infusion, decreased (P < 0.05) as threonine intake increased from 0.05 to 0.15 g/(kg.d) and was low and constant for threonine intakes >0.15 g/(kg.d). Using breakpoint analysis with 95% confidence interval (CI), mean requirement and safe level of parenteral threonine intake were estimated to be 0.19 and 0.21 g/(kg. d), respectively (equivalent to 13 and 14 mg/g amino acids, respectively). To compare these data with those of orally fed controls, we then repeated the experiment by infusing identical diets intragastrically to piglets (n = 25); the varying dietary threonine intakes were 0.1-1.2 g/(kg.d). Employing identical kinetics and analyses, the mean requirement and safe level of oral threonine intake were estimated to be 0.42 and 0.51 g/(kg.d), respectively (equivalent to 28 and 34 mg/g amino acids, respectively). These data demonstrate that the threonine requirement of neonates during TPN is approximately 45% of the mean oral requirement.

Biomechanical models for biological tissues such as articular cartilage generally contain an ideal, dilute solution assumption. In this article, a biomechanical triphasic model of cartilage is described that includes nondilute treatment of concentrated solutions such as those applied in vitrification of biological tissues. The chemical potential equations of the triphasic model are modified and the transport equations are adjusted for the volume fraction and frictional coefficients of the solutes that are not negligible in such solutions. Four transport parameters, i.e., water permeability, solute permeability, diffusion coefficient of solute in solvent within the cartilage, and the cartilage stiffness modulus, are defined as four degrees of freedom for the model. Water and solute transport in cartilage were simulated using the model and predictions of average concentration increase and cartilage weight were fit to experimental data to obtain the values of the four transport parameters. As far as we know, this is the first study to formulate the solvent and solute transport equations of nondilute solutions in the cartilage matrix. It is shown that the values obtained for the transport parameters are within the ranges reported in the available literature, which confirms the proposed model approach.

Osteochondral defects can degenerate into osteoarthritis and currently there are no good treatment alternatives available to most Orthopaedic surgeons. Osteochondral allografting can restore damaged joint surfaces but its clinical use is limited by poor access to high quality tissue. Vitrification of osteochondral tissue would allow the banking of this tissue but requires high concentrations of cryoprotective agents. This study was designed to ascertain dimethyl sulfoxide (DMSO) toxicity kinetics to chondrocytes in situ after exposure to DMSO at different temperatures recorded as a function of time. Porcine osteochondral dowels were exposed to 1, 3, 5, and 6M DMSO at 4, 22, and 37 degrees C for 0.5 min to 120 min. Chondrocyte recovery was determined by membrane integrity (Syto 13 and ethidium bromide) and mitochondrial (WST-1) assays. Results demonstrated that cell recovery was concentration, temperature and time dependent. At higher concentrations and temperatures, significant cell loss occurred within minutes. A rate constant calculated for chondrocyte death was dependent on temperature. 1 M DMSO appeared relatively non-toxic. This experiment established a method to examine systematically toxicity parameters for chondrocytes in situ and this data can be used to tailor vitrification protocols by limiting exposure temperature and time or lowering DMSO concentrations below toxic levels recorded.

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