Origen e historia de la Toxicología / Origin and history of Toxicology

Se presentan en orden cronológico acontecimientos relacionados con la Toxicología ocurridos a nivel mundial y en Cuba. Se realizó una revisión bibliográfica sobre los orígenes e historia de la Toxicología, la cual surgió desde inicios de la humanidad. Se destaca el actuar de reconocidos médicos de la antigüedad, así como acciones realizadas por famosos envenenadores; todo esto recogido en los papiros, la literatura mitológica, religiosa, médica y universal. También se mencionan diversos tóxicos descubiertos o utilizados a lo largo de la historia y su clasificación de acuerdo con su origen animal, vegetal o mineral. Además se evidencia la vinculación que existe entre esta ciencia y la Medicina Legal. La Toxicología quedó constituida como ciencia independiente en la década del cincuenta del pasado siglo.

This paper chronologically described the events in toxicology that took place worldwide and in Cuba. A literature review was made about the origin and history of toxicology, which emerged since the very beginning of mankind. It reflected the actions taken by well known doctors from ancient times, as well as the work of renowned poisoners. Such information was retrieved from papyri and from mythological, religious, medical and universal literature. A number of toxics discovered or used throughout history were mentioned, including their classification according to their origin: animal, vegetable or mineral. Additionally, the linkage between this science and forensic medicine was stressed. Toxicology became an independent science in the 50's of the last century.

The basement membrane of the isolated rat colonic mucosa. A light, electron and atomic force microscopy study.

Basement membranes (BM) are structures of the extracellular matrix (ECM), which are involved in epithelial barriers, but also play an important role in processes such as cell adhesion, cell growth and tissue healing. The aim of this study was to investigate possible effects of cell removal on the structure of the BM of the colonic mucosa. The superficial epithelium was removed with EDTA and the samples were then mechanically fixed for immunohistochemistry, TEM, SEM and AFM. For SEM and AFM, some samples were also prepared according to the OTO method. BM marker proteins were detected after cell removal by immunohistochemistry, indicating that BM remains. However, a lamina lucida (LL) was no longer visible in TEM, it disappeared and the BM became slightly thinner. The surface topography of the BM is characterized by the presence of globules, fenestrations and pore-like structures, which were visualized with SEM and AFM. Noteworthy is the visualization for the first time with AFM of a 3D network of fine fibers and filaments ("cords"), which very much resembled that described with TEM by Inoue (1994). An unresolved question is whether the pore-like structures observed in this study, especially with SEM, actually correspond to the pores of the BM whose existence has been demonstrated functionally. In conclusion, the structural patterns and changes described could be considered as a reference to evaluate the effects of other decellularization protocols on BMs, such as those used in tissue engineering.

The surface topography of the choroid plexus. Environmental, low and high vacuum scanning electron microscopy.

Environmental scanning electron microscopy (ESEM) allows the examination of hydrated and dried specimens without a conductive metal coating which could be advantageous in the imaging of biological and medical objects. The aim of this study was to assess the performance and benefits of wet-mode and low vacuum ESEM in comparison to high vacuum scanning electron microscopy (SEM) using the choroid plexus of chicken embryos as a model, an organ of the brain involved in the formation of cerebrospinal fluid in vertebrates. Specimens were fixed with or without heavy metals and examined directly or after critical point drying with or without metal coating. For wet mode ESEM freshly excised specimens without any pre-treatment were also examined. Conventional high vacuum SEM revealed the characteristic morphology of the choroid plexus cells at a high resolution and served as reference. With low vacuum ESEM of dried but uncoated samples the structure appeared well preserved but charging was a problem. It could be reduced by a short beam dwell time and averaging of images or by using the backscattered electron detector instead of the gaseous secondary electron detector. However, resolution was lower than with conventional SEM. Wet mode imaging was only possible with tissue that had been stabilized by fixation. Not all surface details (e.g. microvilli) could be visualized and other structures, like the cilia, were deformed. In summary, ESEM is an additional option for the imaging of bio-medical samples but it is problematic with regard to resolution and sample stability during imaging.

Experimental femtosecond laser-assisted nanosurgery of anterior lens capsule.

PURPOSE: To investigate femtosecond laser-assisted nanosurgery of the anterior lens capsule in a prospective in vitro study. METHODS: Eight anterior lens capsules obtained during conventional phaco surgery were irradiated with a nonamplified 80-MHz near-infrared 800-nm titanium:sapphire femtosecond laser. Line intratissue laser cuts were examined by femtosecond multiphoton laser scanning microscopy (MLSM) and transmission electron microscopy (TEM). RESULTS: Speed parameters of the laser beam, laser ablation time, and pulse power determined the width of the lesions, which ranged from 220±40 nm (SD) to 1.49±0.15 µm. Both MLSM and TEM revealed minimal collateral alterations in the tissue surrounding the laser cuts. CONCLUSIONS: Nonamplified near-infrared femtosecond laser pulses at low pulse energies may be a promising strategy for precise noncontact nanosurgery of the anterior lens capsule with minimal collateral damage to surrounding tissue. High-resolution MLSM offers 3-dimensional, noninvasive, nondestructive imaging at submicrometer resolution within seconds before and after ablation.

The Bionas technology for anticancer drug screening.

BACKGROUND: The Bionas 2500(®) analyzing system is an advanced label-free technology using a cell-based multi-sensor array, which is commercially available. Data on metabolism, respiration, adhesion, cell proliferation and cell death rates, as well as ligand-receptor interactions (multi-parametric) can be acquired and statistically evaluated. Noteworthy is the possibility of analyzing later after effects and/or recovery after drug treatment. In addition, Bionas supports all conceivable drug application modes (one, two or more drugs). Specimens for drug screening with Bionas consist of human permanent cell lines, primary cell cultures as well as 'tumor slices' obtained from biopsies and surgery material. OBJECTIVE/METHODS: Examples of measurements are presented and discussed. CONCLUSION: Although drug throughput is modest, the high quality of the information allows in-depth evaluation.

In vitro femtosecond laser-assisted nanosurgery of porcine posterior capsule.

PURPOSE: To investigate femtosecond laser-assisted nanosurgery of the posterior capsule in a prospective in vitro animal study. SETTING: Department of Ophthalmology and Eye Hospital, University of Saarland, Homburg, and Fraunhofer Institute for Biomedical Engineering, St. Ingbert, Germany. METHODS: The posterior capsules of 12 porcine eyes were irradiated with a nonamplified 90 MHz near-infrared 750 nm titanium:sapphire femtosecond laser. Intratissue and superficial laser cuts of laser-ablated (5 capsules) and control (1 capsule) specimens were examined by femtosecond multiphoton laser scanning microscopy (MLSM) and transmission electron microscopy (TEM). RESULTS: Laser exposure time and pulse power determined the width of the lesions, which ranged from 0.69 microm+/-0.19 (SD) to 2.81+/-0.5 microm. Both MLSM and TEM revealed minimal collateral alterations in the tissue surrounding the laser cuts. CONCLUSIONS: Nonamplified near-infrared femtosecond laser pulses at low pulse energies may be a promising strategy for precise lamellar noncontact nanosurgery of the posterior capsule, with minimal structural collateral damage to surrounding tissue. High-resolution MLSM offers 3-dimensional, noninvasive, nondestructive imaging at submicrometer resolution within seconds before and after ablation.

Posttraumatic GABA(A)-mediated [Ca2+]i increase is essential for the induction of brain-derived neurotrophic factor-dependent survival of mature central neurons.

A shift of GABA(A)-mediated responses from hyperpolarizing to depolarizing after neuronal injury leads to GABA(A)-mediated increase in [Ca2+](i). In addition, central neurons become dependent on BDNF for survival. Whether these two mechanisms are causally interrelated is an open question. Here, we show in lesioned CA3 hippocampal neurons in vitro and in axotomized corticospinal neurons in vivo that posttraumatic downregulation of the neuron-specific K-Cl cotransporter KCC2 leads to intracellular chloride accumulation by the Na-K-2Cl cotransporter NKCC1, resulting in GABA-induced [Ca2+](i) transients. This mechanism is required by a population of neurons to survive in a BDNF-dependent manner after injury, because blocking GABA(A)-depolarization with the NKCC1 inhibitor bumetanide prevents the loss of neurons on BDNF withdrawal. The resurgence of KCC2 expression during recovery coincides with loss of BDNF dependency for survival. This is likely mediated through BDNF itself, because injured neurons reverse their response to this neurotrophin by switching the BDNF-induced downregulation of KCC2 to upregulation.

Resistance of mitochondrial p53 to dominant inhibition.

BACKGROUND: Mutation of a tumor suppressor allele leaves the second as backup. Not necessarily so with p53. This homo-tetrameric transcription factor can become contaminated with mutant p53 through hetero-tetramerization. In addition, it can be out-competed by the binding to p53 DNA recognition motifs of transactivation-incompetent isoforms (DeltaN and DeltaTA-isoforms) of the p53/p63/p73 family of proteins. Countermeasures against such dominant-negative or dominant-inhibitory action might include the evolutionary gain of novel, transactivation-independent tumor suppressor functions by the wild-type monomer. RESULTS: Here we have studied, mostly in human HCT116 colon adenocarcinoma cells with an intact p53 pathway, the effects of dominant-inhibitory p53 mutants and of Deltaex2/3p73, a tumor-associated DeltaTA-competitor of wild-type p53, on the nuclear transactivation-dependent and extra-nuclear transactivation-independent functions of wild-type p53. We report that mutant p53 and Deltaex2/3p73, expressed from a single gene copy per cell, interfere with the stress-induced expression of p53-responsive genes but leave the extra-nuclear apoptosis by mitochondrial p53 largely unaffected, although both wild-type and mutant p53 associate with the mitochondria. In accord with these observations, we present evidence that in contrast to nuclear p53 the vast majority of mitochondrial p53, be it wild-type or mutant, is consisting of monomeric protein. CONCLUSION: The extra-nuclear p53-dependent apoptosis may constitute a fail-safe mechanism against dominant inhibition.

Femtosecond laser-assisted retinal imaging and ablation: experimental pilot study.

PURPOSE: To investigate retinal imaging and ablation using femtosecond laser pulses. MATERIALS AND METHODS: Two non-amplified near-infrared femtosecond lasers were used to irradiate porcine retinal specimens in vitro. The lasers were used for tissue removal as well as multiphoton laser scanning microscopy. RESULTS: Ablation of the nerve fiber layer was performed at pulse energies of 1.0 nJ to 3.9 nJ. Control laser scanning images were acquired within seconds after irradiation. Specimens were additionally investigated with electron microscopy. CONCLUSIONS: Non-amplified femtosecond lasers may allow precise surgery controlled by fast high-resolution imaging of the target.

In vitro noncontact intravascular femtosecond laser surgery in models of branch retinal vein occlusion.

PURPOSE: To investigate intravenous femtosecond laser surgery in models of branch retinal vein occlusion. MATERIALS AND METHODS: Non-amplified near infrared femtosecond laser was used to ablate polyamide sutures and human hairs inserted into the vascular lumina of porcine retinal veins in vitro. Specimens were subjected to multiphoton laser scanning microscopy and electron microscopy. RESULTS: Regular laser cuts within sutures and hairs were detected with laser microscopy and electron microscopy. Neither laser microscopy nor histology revealed collateral damage of the vascular wall. CONCLUSIONS: Non-amplified femtosecond lasers may allow precise atraumatic non-contact intravenous retinal surgery controlled by high-resolution imaging of the target.

A new method to assess drug sensitivity on breast tumor acute slices preparation.

A method for assessing tumor drug sensitivity is described that is based on preparation of tissue slices and use of silicon chips equipped with electrochemical sensors (multisensor array). The tumor slices (200-300 microM thick) are prepared after surgery and incubated in a medium for recovery after slicing. The advantage, compared to other preparations, is that the original three-dimensional structure is retained. Multisensor arrays measure: (a) pericellular acidification (anaerobic metabolism) and (b) oxygen consumption (respiration). The innovative aspect is that such measurements can be made online, as opposed to using a large battery of endpoint tests on cell vitality and proliferation. Electron microscopy of slices serves to determine cell density and structure and induction of apoptosis/necrosis. Slices of more than 200 breast tumors were used. Metabolic activity was inhibited by sodium fluoride, which reduces glycolysis, and potassium cyanide, which inhibits respiration. These changes are thus reflected in the curves of acidification and oxygen consumption. In other experiments the cytostatic Taxol, an anticytoskeletal agent, was used showing dose and time-dependent effects on acidification and oxygen consumption. In conclusion, the method presented here, is able to provide information on drug sensitivity of a tumor, which aids in designing individualized therapy and is used for drug screening.

Long-term catheters for apheresis and dialysis with surface treatment with infection resistance and low thrombogenicity.

Infection, thrombosis, and stenosis are among the most frequent complications associated with blood-contacting catheters. These problems are usually related to surface properties of the base catheter material. Surface treatment processes, such as ion implantation and ion beam assisted deposition (IBAD) and microdomain structured surfaces, can be used to mitigate such complications. This study evaluated silver coated and implanted large bore catheters used for extracorporeal detoxification. In a 186 patient prospective study, 225 large bore catheters were inserted into the internal jugular or subclavian veins. Eighty-five surface-treated catheters (Spi-Argent, Spire Corporation, Bedford, MA, USA) and 28 catheters with surface treatment (Spi-Silicone, Spire Corporation) were inserted in 90 patients. One hundred and twelve untreated catheters placed in 96 patients served as controls, After removal, the catheters were cultured for bacterial colonization using standard microbiologic assays. They also were examined using a scanning electron microscope (SEM). Bacterial colonization was observed in 8% of the treated catheter compared with 46.4% of untreated catheters. The SEM investigations showed all treated catheters to possess low thrombogenicity. Catheters with microdomain structured surfaces showed same results in preliminary observation. The surface treatments of the large bore catheters can be used to improve thrombus and infection resistance of blood contacting catheters.

Dural cell culture. A new approach to study duraplasty.

Fistulas of the cerebrospinal fluid are often repaired by insertion of grafts of various kinds. However, current knowledge of wound healing after graft insertion is limited, and only a few animal studies are available. The objective of this study is to test whether an in vitro model is suited to analyze cellular healing aspects after duraplasty and to assess dura substitutes in such conditions in regard to their surface attractiveness for cellular migration from the dura margins. Harvested dura pieces from minipigs were perforated to mimic central dura lesions, placed on various coated surfaces (collagen, laminin, poly-L-lysine) or grafts, and investigated in a cell culture for cellular closure of the perforation. Cellular migration from the dura into the central perforation was noted on collagen-coated surfaces and when defects were filled with collagen gels, but there was no cell growth on surfaces with poly-L-lysine or laminin coating. Immunocytochemistry identified the migrating cells mainly as fibroblasts with some intermingled epithelial cells. Scanning electron microscopy proved cellular closure of defects after dura placement on allogenic non-crosslinked collagen transplants. Less cellular migration was observed on poly-P-dioxanon sheets, while no cells migrated into the central dura perforation after placement on a cartilage substitute. Cell counting indicated enhanced cellular closure of the dura opening after introduction of insulin or fibroblast growth factor (sign test for both: 0.031). Our study succeeded in establishing a cell culture model for duraplasty and indicated cellular migration from the dura borders at the site of the defect during the wound healing process. The cell culture model presented in this report shows that collagen grafts are best suited for duraplasty. In accordance with the immunocytological finding of fibroblast migration from the dura borders additional application of fibroblast-stimulating growth factors accelerated cellular defect closure.