Skin wound healing in obese and lean male adolescent rats submitted to pre-weaning litter size manipulation.

We investigated whether early postnatal over-nutrition affects normal course of skin wound healing. To induce over-nutrition the litter size was adjusted on the first day after birth to four pups/nest (small litters). In parallel, as a control, normal nests of 10 pups/nest (normal litters) were used. For the wound healing experiment 30 male Sprague-Dawley rats, 15 from normal nests and 15 from small nests, were used. Two parallel full-thickness skin incisions and two full-thickness excisions were performed on the back of each rat. Samples for histological examination (excisions) and wound tensile strength measurement (incisions) were collected on days 2, 6, and 14 after surgery. Our study demonstrates that rats from the small nests had enhanced plasma levels of insulin and enhanced body weight/fat parameters. Furthermore, in small nests, rats that expressed the above-mentioned symptoms displayed slight improvement of epidermis regeneration, accelerated demarcation line formation, and increased wound tensile strength. From this point of view the small nest model used in the present experiment is helpful for exploration whether these acquired changes might be considered as a sufficient essential factor involved in the regulation of metabolic homeostasis and wound repair in juvenile obese male rats. Nevertheless, further studies need to be performed to verify the present findings also on other animal models and humans and to describe the exact underlying mechanism.

Multiphoton Microscopy and Mass Spectrometry for Revealing Metabolic Heterogeneity of Hepatocytes in vivo.

The aim of the investigation was to study the possibility of revealing the heterogeneity of normal liver hepatocytes in terms of metabolic status using the modern methods of multiphoton microscopy and mass spectrometry. Materials and Methods: Heterogeneity of hepatocytes in terms of total metabolic activity was assessed using multiphoton microscopy based on the autofluorescence intensity of intracellular cofactors NAD(P)H and FAD. Hepatocyte heterogeneity in terms of intensity of intracellular metabolic processes was determined using the fluorescence lifetime imaging (FLIM) method based on the data about fluorescence lifetime contributions of various forms of NAD(P)H. The method of time-of-flight secondary ion mass spectrometry (TоF-SIMS) was used to study the lipid and amino acid composition of hepatocytes. Results: It has been revealed using multiphoton microscopy that hepatocytes are heterogeneous in terms of general metabolic activity. Using FLIM, it was established that the heterogeneity degree was high in terms of intensity of oxidative phosphorylation, glycolysis, and synthetic processes (lipogenesis, nucleic acid synthesis, and the pentose phosphate pathway). The TоF-SIMS method revealed the presence of hepatocyte heterogeneity in terms of amino acid and lipid composition, which points to various intensities of synthetic processes in individual hepatocytes. Moreover, differences in the content of PO3 ions were revealed. The results of ToF-SIMS study correlate with the data obtained by multiphoton microscopy and FLIM, confirming the revealed heterogeneity of hepatocytes in terms of general metabolic activity and intensity of intercellular metabolic processes. Conclusion: The latest methods of fluorescence bioimaging and mass spectrometry proved to be effective in revealing hepatocyte heterogeneity in terms of metabolic status. The presence of heterogeneity should be taken into account in studying the liver tissue under various conditions with the application of fluorescence bioimaging methods.

Intradermal running suture versus 3M™ Vetbond™ tissue adhesive for wound closure in rodents: a biomechanical and histological study.

BACKGROUND: Although tissue adhesives and traditional sutures were compared in numerous studies during the early stages of healing, it has to be clarified, from the histological and biomechanical point of view, how the differences develop during the later phases. METHODS: Twenty-four male Sprague-Dawley rats were used and divided into 2 groups: the IRS (intradermal running suture) and G (glue) groups. Two parallel full-thickness skin incisions were made on the back of each rat. Wounds in the IRS group were closed by an IRS, whereas wounds in the G group were closed using tissue adhesive (n-butylcyanoacrylate). Rats were sacrificed 7 and 22 days after surgery. RESULTS: Similar wound tensile strengths of glued and sutured wounds were measured on days 7 (IRS = 10.3 ± 1.7 g/mm(2) vs. G = 12.9 ± 4.0 g/mm(2), p = 0.9612) and 22 (IRS = 95.6 ± 15.7 g/mm(2) vs. G = 85.6 ± 16.4 g/mm(2), p = 0.2502) after surgery. Histology revealed a significantly increased amount of granulation tissue formation in glued wounds on day 7. The difference in granulation tissue formation was reduced until day 22. CONCLUSIONS: Tissue adhesive based on n-butylcyanoacrylate presents a fair alternative to traditional suture wound closure techniques.

Parametrization of Pedestrian Injuries and its Utilisation in Proving Traffic Accidents Course Using Injury Signatures and Contact Signatures.

BACKGROUND: The paper points out the present limited possibility of using the verbal description of injuries for the needs of experts from the field of road transportation as relevant criminalistics traces, as well as the options of the FORTIS system that creates a new area for a deeper interdisciplinary approach in the field of expert evidence. Further a description of how to create injury signatures and contact signatures and the possibilities of their evaluation and mutual comparison based on the proven individual attributes are described. OBJECTIVE: To evaluate pedestrian injuries by the new proper FORTIS system and to show FORTIS valuability in the assessment of mechanical violence and mechanism of injuries. MATERIAL AND METHODS: Cases of traffic injuries including photodocumentation, graphic schemes, medical files and autopsy protocols processed by the new FORTIS forensic system. RESULTS: A collision between a pedestrian and a vehicle represents a matrix of physical violence having an effect on the pedestrian´s body and a matrix of the pedestrian´s body´s response to this violence. The analysis of individual cases shows the valuability of the FORTIS system. CONCLUSION: It is apparent that for the needs of traffic accidents analysts the FORTIS system has more options for being used in forensic medicine, as it covers not only a field of evaluation of traffic injuries but also all kinds of injuries and accidents (rail accidents, air accidents, violent crimes, etc.).

Effect of prolonged psychoemotional stress and melatonin on experimental mammary carcinogenesis in female rats.

The aim of the present study was to determine whether prolonged stress repeated immobilization in boxes during the period of 18 weeks (IMS) influenced development and progression of N-methyl-N-nitrosourea (NMU)-induced mammary tumors in female Sprague-Dawley rats and whether long-term MEL application affected changes caused by stress. NMU was applied intraperitoneally in two doses each of 50 mg/kg b.w. between 40-50 postnatal days. Melatonin (MEL) was administered in drinking water in a concentration of 4 microg/ml (daily from 3 p.m to 8 a.m), application was initiated 3 days prior to first NMU dose and lasted until the end of the experiment. Immobilization (2 h/day) began on the fifth day after second carcinogen application, animals were immobilized three times a week. Repeated immobilization of rats during 18 weeks decreased tumor frequency per group and per animal by 30% and tumor volume gain by 16% as opposed to control (NMU) animals. Combination of repeated immobilization and a long-term MEL application lowered incidence by 13% when compared to control, prolonged latency by 13%, decreased tumor frequency per group (by 44%) and per animal (by 35%). Tumor volume gain increased by 35% but their cumulative volume prominently decreased by 74% as opposed to control. Tumor volume was the most markedly influenced by MEL, induced tumors developed more rapidly tumor volume gain increased by 61%. However, their cumulative volume markedly decreased by 75% when compared to immobilized group drinking water. Prolonged stress inhibited development and progression of NMU-induced mammary gland tumors in female rats and this effect was enhanced by long-term melatonin administration.

Expression of MDR proteins in breast cancer and its correlation with some clinical and pathological parameters.

The aim of this work was to determine the expression of the multidrug resistance (MDR) proteins, namely MDR1 (P-glycoprotein), MRP1 (multidrug resistance-related protein) and LRP (lung resistance-related protein), in 87 samples of breast carcinoma. Detection of these proteins was provided by using indirect enzymatic immunohistochemistry. Our findings were compared with the other clinical and pathological parameters: expression of Her2/neu, estrogen receptor status (ER), progesteron receptor status (PR), histological grade and regional lymph node status. For statistical analysis, non-parametric two sided Mann-Whitney-U test was used. Majority of breast carcinoma specimens show positivity for these proteins. The MDR1 and MRP1 signal was found in the cytoplasm of cancer cells. The expression of LRP was detected in the cytoplasm close to the nuclear membrane. The samples were positive for MDR1 protein in 57%, for MRP1 in 84% and for LRP in 79%. Comparing our results with other clinical and pathological parameters, negative correlation between ER, PR and MDR1 expressions and histological grading status was found. No associations were observed between the MRP1 and LRP proteins and histological grading, as well as between the expression of three MDR proteins and the other clinically relevant parameters. In conclusion, high frequency of expression of MDR proteins in breast carcinoma cells suggests, that these proteins might be an important factor of drug resistance in breast carcinoma. Nevertheless, the negative correlation between the histological grade of malignancy of tumor and the expression of ER, PR and MDR1 indicates possible influence of progressive tumor cell de-differentiation. However, this finding has to be confirmed in additional evaluations.

Immunohistochemical detection of MRP1 protein in normal and hyperplastic human thyroid gland.

In this study we describe the localization of MRP1 in normal and hyperplastic thyroid tissues. To demostrate this protein we have used the enzyme imunohistochemical method with monoclonal antibody MRPm6. We have found MRP1 to be present in follicular epithelial cells of normal thyroid tissue and higher expression in the same cells of hyperplastic thyroid tissue. The brown colour of the diaminobenzidine reaction product allowed us to define the distribution of stain clearly. (Fig. 4, Ref. 16.)

Immunohistochemical detection of LRP protein in the normal human lung.

In this study, we have determined the LRP (lung resistance-related protein) by immunohistochemical method. LRP belongs to proteins which cause the multidrug resistance (MDR). It has been found in various normal human tissues, where it plays a protective role against toxic compounds. Multidrug-resistant cells distribute the cytotoxic drug into the perinuclear region and then redistribute it back into the cytoplasm. It is just a hypothesis today that LRP can mediate drug resistance by regulating both the cytoplasmic redistribution and the nucleocytoplasmic transport of drugs. In order to detect LRP we have used the paraffin-embedded sections of the normal human lung tissue. LRP was predominantly located in two regions: 1) in bronchial epithelial cells and 2) in alveolar macrophages. Positive cells were coloured brown and showed strong reactivity. Negative control included the omitting of primary antibody and replacing it by buffer solution. Bronchial epithelial cells and alveolar macrophages stayed uncoloured, i.e. unreactive. (Fig. 5, Ref. 17.).

[Death in an unusual body position]. / Smrt' v neobvyklej polohe tela.

Death in head-down position is rare and uncommon. The mechanism of death in such cases is rather complicated. The authors present a case of unknown man who was found dead in a wood, hanging in head-down position fixed at the branchfork of a tree by his right foot. The authors analyse the possible mechanisms of death in so-called postural asphyxia.

Photoinduced antitumour effect of hypericin can be enhanced by fractionated dosing.

The in vivo antitumour activity of the natural photosensitizer hypericin was evaluated. C3H/DiSn mice were inoculated with fibrosarcoma G5:1:13 cells. When the tumour reached a volume of 40-80mm3 the mice were intraperitoneally injected with hypericin, either in a single dose (5mg/kg; 1 or 6h before laser irradiation) or two fractionated doses (2.5 mg/kg; 6 and 1 h before irradiation with laser light; 532 nm, 70mW/cm2, 168 J/cm2). All tumours in control groups treated with hypericin alone as well as those irradiated with laser light alone had similar growth rates and none of these tumours regressed spontaneously. Complete remission of tumour in photodynamic therapy (PDT)-treated groups was similar (14-17% single dose vs. 33% fractionated dose), but the fractionated schedule of hypericin dosing was found to be more efficient than the single dose, measured by survival assay (p < 0.05). Our experimental model showed that fractionated administration of hypericin can produce a better therapeutic response than single administration.