Animal studies in spinal cord injury: a systematic review of methylprednisolone.

The objective of this study was to examine whether animal studies can reliably be used to determine the usefulness of methylprednisolone (MP) and other treatments for acute spinal cord injury (SCI) in humans. This was achieved by performing a systematic review of animal studies on the effects of MP administration on the functional outcome of acute SCI. Data were extracted from the published articles relating to: outcome; MP dosing regimen; species/strain; number of animals; methodological quality; type of injury induction; use of anaesthesia; functional scale used; and duration of follow-up. Subgroup analyses were performed, based on species or strain, injury method, MP dosing regimen, functional outcome measured, and methodological quality. Sixty-two studies were included, which involved a wide variety of animal species and strains. Overall, beneficial effects of MP administration were obtained in 34% of the studies, no effects in 58%, and mixed results in 8%. The results were inconsistent both among and within species, even when attempts were made to detect any patterns in the results through subgroup analyses. The results of this study demonstrate the barriers to the accurate prediction from animal studies of the effectiveness of MP in the treatment of acute SCI in humans. This underscores the need for the development and implementation of validated testing methods.

Detection of PhIP in grilled chicken entrées at popular chain restaurants throughout California.

Heterocyclic amines (HCAs), compounds formed when meat is cooked at high temperatures particularly through pan frying, grilling, or barbequing, pose a potential carcinogenic risk to the public. It is unclear whether there is any level at which consumption of HCAs can be considered safe. Efforts to measure these compounds mainly include cooking studies under laboratory conditions and some measurement of home-cooked foods, but analysis of commercially cooked foods has been minimal. Attempts to estimate exposure of the public to these compounds must take into consideration dining outside the home, which could result in significant exposure for some individuals. We surveyed at least 9 locations each of 7 popular chain restaurants (McDonald's, Burger King, Chick-fil-A, Chili's, TGI Friday's, Outback Steakhouse, and Applebee's) in California, collecting one or two entrees from each location. Entrees were analyzed for 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) using high-performance liquid chromatography tandem mass spectrometry. All 100 samples contained PhIP. Concentrations were variable within and between entrees and ranged from 0.08 to 43.2 ng/g. When factoring in the weight of the entrees, absolute levels of PhIP reached over 1,000 ng for some entrees. Potential strategies for reducing exposure include the avoidance of meats cooked using methods that are known to form PhIP.

Animal models in spinal cord injury: a review.

Multiple neuroprotective agents have shown benefit for the treatment of acute spinal cord injury (SCI) in animal studies. However, clinical trials have, thus far, been uniformly disappointing. This review explores reasons for discrepancies between promising animal studies and disappointing clinical trials and potential barriers to extrapolation of research results from animals to humans. The three major barriers disclosed are: differences in injury type between laboratory-induced SCI and clinical SCI, difficulties in interpreting functional outcome in animals, and inter-species and interstrain differences in pathophysiology of SCI. These barriers can impair the effectiveness of animal models of SCI to predict human outcomes. While some of these barriers can be overcome, others are inherent to the animal models.

Development of a novel ELISA for human insulin using monoclonal antibodies produced in serum-free cell culture medium.

OBJECTIVES: Development of an ELISA for human insulin that utilizes monoclonal antibodies (mAbs) produced in serum-free medium. DESIGN AND METHODS: Insulin mAbs were produced in vitro by hybridomas in serum-free medium. A two-step ELISA was developed to replace bovine insulin (standard) and bovine serum albumin (assay buffer) with non-animal reagents. RESULTS: The sensitivity of the insulin ELISA was 0.73 uU/mL with a dynamic range of 2-200 uU/mL. No cross-reactivity with either human C-peptide or human proinsulin was observed. The intra- and inter-assay CVs were <7%. The mean recovery of insulin added to plasma samples was between 102.2% and 105.7%. The mean linearity of dilution was between 93% and 110% of undiluted plasma samples. The animal serum-free (ASF) insulin ELISA showed no marked degradation of any kit component when stored at 37 degrees C for up to 7 days. Significantly higher fasting insulin levels were observed in overweight or obese subjects (n=12) compared to lean subjects (n=10, p<0.05). Feeding markedly increased fasting insulin levels in both lean (p<0.02) and overweight or obese (p<0.005) subjects. Excellent correlation was observed between insulin levels measured by ASF insulin ELISA and another CE marked insulin ELISA (y=1.06x-0.44, r=1.00, n=44). CONCLUSIONS: This novel insulin ELISA provides precision and reliability equal to methods currently used in clinical research and serves as a guide for the development of other serum-free immunoassays.

Serum-free hybridoma culture: ethical, scientific and safety considerations.

Despite considerable progress in the development of cell culture techniques, including the development of the serum- and protein-free media that now routinely support hybridoma and mammalian cell growth, fetal bovine serum (FBS) supplemented media are still commonly used: a practice that raises ethical, scientific and safety concerns. The use of FBS in hybridoma culture media is examined here, with regards to the development and production of monoclonal antibodies (mAbs), and it is our recommendation that researchers adopt serum-free cell culture methods to reduce animal use in this area.

Toxicological and metabolic investigations of the safety of N-alpha-lauroyl-L-arginine ethyl ester monohydrochloride (LAE).

Studies were conducted to assess the safety of N-alpha-lauroyl-L-arginine ethyl ester monohydrochloride, (LAE), a novel food preservative, or Mirenat-N (a 25% solution of LAE in propylene glycol). Short term studies demonstrated low acute toxicity. LAE was shown to have mild dermal irritation effects but neither LAE nor Mirenat-N are skin sensitizers. LAE was demonstrated to be a severe eye irritant. In two 13-week feeding studies in rats, systemic NOAELs were established for LAE at 15,000 ppm and for Mirenat-N at 50,000 ppm. There were no signs of neurotoxicity with LAE after 13-weeks at dietary levels as high as 50,000 ppm. Embryo-fetal studies with LAE in rats and rabbits showed no developmental effects at oral gavage doses up to 2000 and 1000 mg/kg/day for rats and rabbits, respectively. NOAELs for systemic maternal effects (reduced food intake and body weights in rabbits) were 2000 mg/kg/day for rats and 300 mg/kg/day for rabbits. In a battery of 5 in vitro genotoxicity tests with LAE or Mirenat-N, neither material was observed to have genotoxic (clastogenic or mutagenic) activity. Metabolism studies with LAE show that it is rapidly metabolized to the amino acid arginine by hydrolysis of the ethyl ester and lauroyl amide functions. The arginine subsequently enters the naturally occurring urea cycle where it is further metabolized to ornithine and urea and eventually to CO(2) through normal mammalian biochemical pathways. The other product of LAE cleavage is lauric acid, which is a human dietary component found in many plant sources, and as such, would enter into normal fatty acid metabolism.