Histomorphometric estimation of age in paraffin-embedded ribs: a feasibility study.

Estimation of age at death from human bones in legal medicine or in anthropology and archaeology is hampered by controversial results from the various macroscopic and histological techniques. This study attempted an estimation of age at death by histomorphometric analysis, from the fourth left rib adjacent to the costochondral joint in 80 forensic cases. Use of the picrosirius dye provided a reliable staining of the decalcified paraffin-embedded ribs. The total bone cortical area, the major and minor diameter as well as the area of the Haversian canals, the osteon areas of intact and remodelled secondary osteons, the area of non-Haversian canals were evaluated by means of image analysis, and derived parameters were calculated on both the internal and external sides of the rib. Most of the variables exhibited consistency between three different observers. Noteworthy, morphometric measurements in the internal cortex of the rib showed less variability than in the external cortex. Finally, discriminant statistical analysis from the 80 cases in this study indicated that the osteon population density was virtually sufficient to significantly discriminate between three groups of age: 20-39 (adulthood), 40-59 (middle age) and a group superior to 60. A subsequent blind evaluation of ten new subjects satisfactorily classified seven subjects out of ten within the three age groups. These results make feasible a larger study aimed at characterization of the practical relationships between bone tissue histomorphometry in ribs and chronological age in forensic cases.

The minipig in dermatotoxicology: methods and challenges.

In view of more morphological and physiological similarities between human and porcine skin than for other laboratory animal species, the minipig is a preferred model to evaluate the safety profile of dermally applied xenobiotics. Different methods of dermal administration and examples of non-invasive and invasive investigations during the in-life phase of the studies are described. Routine and special post-mortem examinations in dermal studies are presented to assess responses to the topical treatment of minipig skin. Challenges in dermal minipig studies are discussed with respect to animal welfare and husbandry, test formulations, application methods and different types of investigations. One of the most significant issues for dermal minipig studies is the extensive measures required to prevent cross-contamination of blood and tissue samples taken to monitor local and systemic exposure to the test item.

The presence of drug in control samples during toxicokinetic investigations--a Novartis perspective.

During a submission procedure, the validity of a few dietary toxicity studies was questioned because low levels of the drug were detected among control toxicokinetic samples. Although several lines of reasoning suggested that these findings arose from ex vivo contamination, the Regulatory Authority stated that it was not possible to establish a no-effect-level in any of the studies and so the submission was withdrawn. In response, Novartis conducted a thorough review and modification of the procedures involved in the collection and analysis of toxicokinetic samples to minimize such contamination in future studies. Ongoing monitoring of contamination in toxicology studies has subsequently revealed that although it was not possible to completely eliminate the problem, the new procedures together with an increasing awareness of the issue have considerably reduced the incidence of contamination. The process of contamination and its control was also modeled in a feeding study in mice. This provided good evidence that the detection of drug in control samples in the previous studies originated from external sources and not from in vivo exposure.

LAV694, a new antiproliferative agent showing improved skin tolerability vs. clinical standards for the treatment of actinic keratosis.

The skin tolerability of the tubulin polymerisation inhibitor LAV694 was compared to that of 5% 5-fluorouracil (5-FU) and 0.5% podophyllotoxin in vitro using a human reconstructed epidermis (HRE), and in vivo using minipigs. Topical treatment of HRE for 1 or 3 days with a 0.2, 0.6 or 1% LAV694 cream or the placebo showed no signs of irritation in terms of morphology, cell viability (lactate dehydrogenase leakage) or interleukin-8 mRNA expression and release. 5-FU increased interleukin-8 production and induced morphological signs of irritation. The substances were also applied under occlusion to the back of two minipigs, twice daily, for 9 days to allow intraindividual comparison of skin effects and tolerability. Skin reactions were monitored by visual scoring, chromometry, pro-inflammatory activity, cell cycle and apoptosis by RT-PCR, laser scanning cytometry and histopathological examination of biopsies. Application of podophyllotoxin and 5-FU had to be stopped on days 4 and 8, respectively, due to severe skin lesions. LAV694 (1%) induced only moderate skin reddening after 9 days. 5-FU and podophyllotoxin, but not LAV694, increased mRNA expression of pro-inflammatory cytokines. LAV694 arrested keratinocytes in the M phase of the cell cycle and apoptosis was detected histologically in the basal layer. LAV694 increased the expression of pro-apoptotic genes in both experimental models. In conclusion, LAV694 selectively induced apoptosis, rather than necrosis, of growth-arrested keratinocytes, thus avoiding the occurrence of extensive inflammation. This resulted in an improved skin tolerability in comparison with 5-FU and podophyllotoxin.