Effects of maternal care and selection for low mortality on tyrosine hydroxylase concentrations and cell soma size in hippocampus and nidopallium caudolaterale in adult laying hen.

Feather pecking and cannibalism in farm-kept laying hens are damaging behaviors both in terms of animal welfare and economic loss, and a major challenge in modern poultry farming. Both rearing with a foster hen and genetic selection have been demonstrated to reduce feather pecking in laying hens. We examined the effects of rearing with a foster hen, genetic selection for low mortality from cannibalism, and interactions between both, using cellular morphology and levels of the rate-limiting enzyme in dopamine production, tyrosine hydroxylase, in the hippocampus and nidopallium caudolaterale (NCL) as a potential measure for laying hen welfare. Hens from the second generation of a sib-selection scheme line derived from a pure-bred White Leghorn line, selected for low mortality and for production characteristics, or their control line (CL) selected only for production characteristics, were housed with or without a foster Silky hen for the first 7 wk of life. Aside from the presence or absence of a foster Silky hen during the first 7 wk of life, housing conditions were identical for all hens. The hens were then sacrificed and brains were removed at 52 wk of age. Brains were sectioned and stained using a Nissl staining to reveal cell soma morphology, or using immunocytochemistry for tyrosine hydroxlase. A greater degree of lateralization in the hippocampus was observed in hens reared without a foster hen, as measured by absolute difference in cell soma size between hemispheres (P<0.05). The low mortality line showed decreased concentrations of tyrosine hydroxylase in the NCL compared with the CL (P<0.005). Our results indicate that morphological changes in brain induced in very early life can be detected in adult hens, and that genetic selection against mortality due to cannibalism impacts tyrosine hydroxylase in the NCL of laying hens. These observations strengthen the notion that brain measures may be useful as potential readouts for animal welfare.

The standardization-generalization dilemma: a way out.

Recently, a debate has emerged on the use and necessity of standardization in experimental testing using animal subjects. The difficulties encountered when trying to reconcile standardization and generalization largely underlie this debate. The more specific the testing procedures are, the less one can generalize to more naturalistic situations, including to human clinical populations. If the goal of a study is to generalize to a larger population, there may be a higher risk attached to false-positive than false-negative results; thus the balance sways toward generalization. Heterogenization of housing conditions and of genetic makeup of experimental animals has been suggested as a possible method to increase the generalizability of results. It is important to remain cognizant, however, of situations in which false negatives can be counterproductive or even dangerous, such as when the goal is to elucidate a physiological mechanism, when expected effect sizes are small, in toxicological studies and in drug safety testing. In such cases, experiments based on standardization may provide more useful information. We pose that it is essential that the goal of the specific experiment conducted is clearly defined and that the decision to balance between standardization and generalization must be made based on the specific needs to meet the intended goal. In this light, we discuss a multi-tiered approach to animal experimentation, in which standardization and generalizability are each given precedence during different phases of a project, depending upon the goal of the experiment.

Cognitive performance in neurokinin 3 receptor knockout mice.

RATIONALE: The neurokinin 3 (NK(3)) receptor is a novel target under investigation for improvement of the symptoms of schizophrenia due to its ability to modulate dopaminergic signaling. However, research on effects of NK(3) antagonism with animal models has been hindered because of species differences in the receptor between humans, rats, and mice. OBJECTIVES: The aim of the present study is to further knowledge on the role of NK(3) in cognitive functioning by testing the effect of knockout of the NK(3) receptor on tests of working memory, spatial memory, and operant responding. MATERIALS AND METHODS: NK(3) knockout mice generated on a C57Bl/6 background were tested in delayed matching to position (DMTP), spontaneous alternation, Morris water maze, and active avoidance tasks. RESULTS: NK(3) knockout mice showed better performance in the DMTP task, though not delay dependently, which points to an effect on operant performance but not on working memory. No differences were seen between the groups in spontaneous alternation, another indication that working memory is not affected in NK(3) knockouts. There was no impairment in knockout mice in Morris water maze training, and the mice also showed faster response latency in the active avoidance task during training. CONCLUSIONS: Collectively, these results support a role for the NK(3) receptor in performance of operant tasks and in spatial learning but not in working memory.

Metabotropic glutamate receptor modulation, translational methods, and biomarkers: relationships with anxiety.

RATIONALE: The increasing awareness of the need to align clinical and preclinical research to facilitate rapid development of new drug therapies is reflected in the recent introduction of the term "translational medicine". This review examines the implications of translational medicine for psychiatric disorders, focusing on metabotropic glutamate (mGlu) receptor biology in anxiety disorders and on anxiety-related biomarkers. OBJECTIVES: This review aims to (1) examine recent progress in translational medicine, emphasizing the role that translational research has played in understanding of the potential of mGlu receptor agonists and antagonists as anxiolytics, (2) identify lacunas where animal and human research have yet to be connected, and (3) suggest areas where translational research can be further developed. RESULTS: Current data show that animal and human mGlu(5) binding can be directly compared in experiments using the PET ligand (11)C-ABP688. Testing of the mGlu(2/3) receptor agonist LY354740 in the fear-potentiated startle paradigm allows direct functional comparisons between animals and humans. LY354740 has been tested in panic models, but in different models in rats and humans, hindering efforts at translation. Other potentially translatable methods, such as stress-induced hyperthermia and HPA-axis measures, either have been underexploited or are associated with technical difficulties. New techniques such as quantitative trait loci (QTL) analysis may be useful for generating novel biomarkers of anxiety. CONCLUSIONS: Translational medicine approaches can be valuable to the development of anxiolytics, but the amount of cross-fertilization between clinical and pre-clinical departments will need to be expanded to realize the full potential of these approaches.

Expression of amphetamine sensitization is associated with recruitment of a reactive neuronal population in the nucleus accumbens core.

RATIONALE: Repeated exposure to psychostimulant drugs causes a long-lasting increase in the psychomotor and reinforcing effects of these drugs and an array of neuroadaptations. One such alteration is a hypersensitivity of striatal activity such that a low dose of amphetamine in sensitized animals produces dorsal striatal activation patterns similar to acute treatment with a high dose of amphetamine. OBJECTIVES: To extend previous findings of striatal hypersensitivity with behavioral observations and with cellular activity in the nucleus accumbens and prefrontal cortex in sensitized animals. MATERIALS AND METHODS: Rats treated acutely with 0, 1, 2.5, or 5 mg/kg i.p. amphetamine and sensitized rats challenged with 1 mg/kg i.p. amphetamine were scored for stereotypy, rearing, and grooming, and locomotor activity recorded. c-fos positive nuclei were quantified in the nucleus accumbens and prefrontal cortex after expression of sensitization with 1 mg/kg i.p. amphetamine. RESULTS: Intense stereotypy was seen in animals treated acutely with 5 mg/kg amphetamine, but not in the sensitized group treated with 1 mg/kg amphetamine. The c-fos response to amphetamine in the accumbens core was augmented in amphetamine-pretreated animals with a shift in the distribution of optical density, while no effect of sensitization was seen in the nucleus accumbens shell or prefrontal cortex. CONCLUSIONS: A lack of stereotypy in the sensitized group indicates a dissociation of behavioral responses to amphetamine and striatal immediate-early gene activation patterns. The increase in c-fos positive nuclei and shift in the distribution of optical density observed in the nucleus accumbens core suggests recruitment of a new population of neurons during expression of sensitization.

Effects of aripiprazole/OPC-14597 on motor activity, pharmacological models of psychosis, and brain activity in rats.

Aripiprazole (OPC-14597) is an antipsychotic with a unique pharmacology as a dopamine D2 receptor partial agonist, which has been demonstrated to reduce symptoms of schizophrenia. To further profile this compound in preclinical models, we examined aripiprazole-induced activity changes as measured by pharmacological magnetic resonance imaging (MRI) and characterized the drug in several rodent models of motor behaviors and of psychosis. Continuous arterial spin labeling MRI measuring blood perfusion (as an indirect measure of activity) reveals that aripiprazole dose-dependently decreased brain activity in the entorhinal piriform cortex, perirhinal cortex, nucleus accumbens shell, and basolateral amygdala. While no deficits were observed in the rotarod test for motor coordination in the simpler (8 RPM) version, in the more challenging condition (16 RPM) doses of 10 and 30mg/kg i.p. produced deficits. Catalepsy was seen only at the highest dose tested (30mg/kg i.p.) and only at the 3 and 6h time points, not at the 1h time point. In pharmacological models of psychosis, 1-30mg/kg aripiprazole i.p. effectively reduced locomotor activity induced by dopamine agonists (amphetamine and apomorphine), NMDA antagonists (MK-801 and phencyclidine (PCP)), and a serotonin agonist (2,5-dimethoxy-4-iodoamphetamine (DOI)). However, aripiprazole reversed prepulse inhibition deficits induced by amphetamine, but not by any of the other agents tested. Aripiprazole alters brain activity in regions relevant to schizophrenia, and furthermore, has a pharmacological profile that differs for the two psychosis models tested and does not match the typical or atypical psychotics. Thus, D2 partial agonists may constitute a new group of antipsychotics.

Augmented reinforcer value and accelerated habit formation after repeated amphetamine treatment.

Various processes might explain the progression from casual to compulsive drug use underlying the development of drug addiction. Two of these, accelerated stimulus-response (S-R) habit learning and augmented assignment of motivational value to reinforcers, could be mediated via neuroadaptations associated with long-lasting sensitization to psychostimulant drugs, i.e. augmented dopaminergic neurotransmission in the striatum. Here, we tested the hypothesis that both processes, which are often regarded as mutually exclusive alternatives, are present in amphetamine-sensitized rats. Amphetamine-sensitized rats showed increased responding for food under a random ratio schedule of reinforcement, indicating increased incentive motivational value of food. In addition, satiety-specific devaluation experiments under a random interval schedule of reinforcement showed that amphetamine-sensitized animals exhibit accelerated development of S-R habits. These data show that both habit formation and motivational value of reinforcers are augmented in amphetamine-sensitized rats, and suggest that the task demands determine which behavioral alteration is most prominently expressed.

In situ photoimmunotherapy: a tumour-directed treatment for melanoma.

We report a new immunological treatment for advanced cutaneous melanoma which combines laser stimulation with topical application of a toll-like receptor agonist. This treatment, in situ photoimmunotherapy (ISPI), provides an alternative to traditional therapies for melanoma patients with cutaneous metastases. A 6-week cycle of ISPI is carried out on cutaneous metastases located in a designated 20 x 20 cm treatment area: 2 weeks of pretreatment with twice-daily topical applications of imiquimod (5% cream under plastic occlusion), with a laser treatment session at week 2 and again at week 4. Topical imiquimod is continued for the entire 6-week cycle. Two patients with late-stage melanoma were treated with ISPI. Patient 1 had the primary tumour and local metastases on the left arm, as well as metastatic tumours in the lungs [American Joint Committee on Cancer (AJCC) stage IV]. Patient 2 had a head and neck melanoma with multiple local metastases (AJCC stage IIIC), which had failed repeated attempts at surgical resection and high-dose radiation therapy. Patient 1 is now free of all clinically detectable tumours (including the lung metastases) >20 months after the first treatment cycle. Patient 2 has been free of any clinical evidence of the tumour for over 6 months. These two cases demonstrate that ISPI can clear local tumour and trigger beneficial systemic responses, with a side-effect profile that compares favourably with other treatments for advanced melanoma.

C-fos activation patterns in rat prefrontal cortex during acquisition of a cued classical conditioning task.

The prefrontal cortex (PFC) is known to be involved in associative learning; however, its specific role in acquisition of cued classical conditioning has not yet been determined. Furthermore, the role of regional differences within the PFC in the acquisition of cued conditioning is not well described. These issues were addressed by exposing rats to either one or four sessions of a cued classical conditioning task, and subsequently examining c-fos immunoreactivity in various areas of the PFC. Differences in patterns of c-fos immunopositive nuclei were found when comparing the PFC areas examined. No significant differences were found between rats presented with a temporally contingent conditioned stimulus (CS) light and food (paired groups) and those presented with the same stimuli temporally non-contingently (unpaired groups). In lateral and orbital PFC, both the paired and unpaired groups showed more c-fos immunopositive nuclei than control groups exposed only to the behavioral setup (context exposed groups), and all groups showed a drop in c-fos immunopositive nuclei from session 1 to session 4. In dorsal medial PFC, no differences were seen between the paired, unpaired and context exposed groups. These groups did, however, differ from naive animals, an effect that was not seen in the ventral medial PFC. The results of this study do not support a role for the PFC in the acquisition of a cued classical conditioning task. The differences seen between paired, unpaired and context exposed groups in orbital and lateral PFC could be due to contextual conditioning or reward-related effects.

Ciprofloxacin microprecipitates and macroprecipitates in the human corneal epithelium.

In 4 corneal transplantation patients treated preoperatively with ciprofloxacin ophthalmic drops, microprecipitates associated with damaged corneal epithelium were noted in 2 patients. Another patient developed a large macroprecipitate in a corneal ulcer. All specimens were examined by electron microscopy and high-pressure liquid chromatography. The crystalline precipitates were pure ciprofloxacin. The macroprecipitate demonstrated a large zone of inhibition on agar plates seeded with a susceptible organism at 24 and 48 hours. It was bioactive and bioavailable in vitro.

Antitumor immunity induced by laser immunotherapy and its adoptive transfer.

The ideal cancer treatment modality should not only cause tumor regression and eradication but also induce a systemic antitumor immunity, which is essential for control of metastatic tumors and for long-term tumor resistance. Laser immunotherapy using a laser, a laser-absorbing dye, and an immunoadjuvant has induced such long-term immunity in treatment of a mammary metastatic tumor. The successfully treated rats established total resistance to multiple subsequent tumor challenges. To further study the mechanisms of the antitumor immunity induced by this novel treatment modality, passive adoptive transfer was performed using splenocytes as immune cells. The spleen cells that were harvested from successfully treated tumor-bearing rats provided 100% immunity in the naive recipients. The passively protected first cohort rats were immune to tumor challenge with an increased tumor dose; their splenocytes also prevented the establishment of tumor in the second cohort of naive recipient rats. This immunity transfer was accomplished without the usually required T-cell suppression in recipients.

Histological comparison of corneal ablation with Er:YAG laser, Nd:YAG optical parametric oscillator, and excimer laser.

PURPOSE: To use histological techniques to assess and compare the ablation depth, local damage, and surface quality of corneal ablations by a Q-switched Er:YAG laser, an optical parametric oscillator laser at 2.94 microm, a long pulse Er:YAG laser, and a 193-nm excimer laser. METHODS: Human cadaver eyes and in vivo cat eyes were treated with a 6.0-mm diameter, 30-microm-deep phototherapeutic keratectomy ablation and a 6.0-mm diameter, -5.00-D photorefractive keratectomy ablation. Human cadaver eyes were also treated with a 5.0-mm diameter, -5.00-D laser in situ keratomileusis (LASIK) ablation. Fluences and pulse widths used were 200 mJ/cm2 and 70 ns for the Q-switched Er:YAG, 150 mJ/cm2 and 7 ns for the optical parametric oscillator laser (OPO), 500 mJ/cm2 and 50 microseconds for the long pulse Er:YAG, and 160 mj/cm2 and 20 ns for the excimer laser. In the ablation rate study, 12 porcine eyes were ablated by the OPO laser with a range of layers and at different fluences ranging from 60 to 150 mJ/cm2, all using a 1.5-mm spot on the eye. The ablation depth of these acute ablations was evaluated by light microscopy examination. RESULTS: In the acute damage study, light microscopy showed a thin surface layer in all samples with minimal thermal damage except on the long pulse Er:YAG corneas. Transmission electron microscopy revealed less than 0.3-microm surface damage for all specimens of both the optical parametric oscillator and the excimer laser samples with no evidence of collagen shrinkage. Transmission electron microscopy showed damage layers of 0.5 to 3 microm for Q-switched Er:YAG and 3 to 10 microm for long pulse Er:YAG. Scanning electron microscopy showed smooth surfaces in all eyes, although the excimer was the roughest. In the porcine eye study, ablations were produced in both PTK and PRK modes with the ablation rate per layer increasing with the fluence. At 120 mJ/cm2, the average ablation rate was 1.9 microm per layer. CONCLUSIONS: The histology from the short pulse mid-infrared optical parametric oscillator laser at 2.94 microm was comparable to the 193-nm excimer with a smooth, damage-free, ablation zone when performing PRK and LASIK.

Healing after photorefractive keratectomy in cat eyes with a scanning mid-infrared Nd:YAG pumped optical parametric oscillator laser.

PURPOSE: To evaluate the healing characteristics of cat corneas treated with a new scanning mid-infrared laser system. METHODS: Six adult cats were treated with 6-mm diameter photorefractive keratectomy (PRK) corrections. One eye in each animal was untreated as a control and the other was treated with either a -3.00 or -6.00 diopter ablation. The laser was a new Nd:YAG pumped optical parametric oscillator laser at 2.94 microm with a new scanning delivery system. The pulse width was 7 nanoseconds, the repetition rate was 10 Hz, the size of the laser spot on the eye was 1.0 mm, and the fluence was 150 mJ/cm2. Healing of the cat corneas was followed for 4 months. Slit-lamp and corneal topography evaluations were done at each follow-up examination. Histology was performed at the end of the study. RESULTS: The corneal epithelium healed within 1 week. There was no stromal haze in any eye after the epithelium healed. After the first 2 weeks, slit-lamp examination could not identify which eye was treated. Corneal topography showed corneal flattening. Light microscopy at 4 months revealed normal epithelium and increased keratocyte density in the anterior third of the cornea. Electron microscopy showed discontinuities in the basement membrane and hemidesmosomes. The deep stroma and endothelium were normal. CONCLUSIONS: Cat corneas treated with the new optical parametric oscillator laser healed normally with no adverse effects. Increased keratocyte activity in the anterior stroma was the only noticeable response besides the flattening shown by topography.

Tumour cell damage and leucocyte infiltration after laser immunotherapy treatment.

Immune response after laser-photosensitiser application could be crucial in treatment of cancers, because without it there could be no systemic, long-term tumour control. Laser immunotherapy, a novel method for treatment of metastatic tumours, uses a near-infrared laser, a laser-absorbing dye indocyanine green, and an immunoadjuvant glycated chitosan. This modality has shown an induced antitumour immune response in treatment of rat mammary metastatic tumours. The influence of this new method on the cellular structure of the tumours and on the infiltrating immune cells was studied using optical and electron microscopes. The tumour samples were examined before and immediately after the treatment for acute effects, which appeared mainly photothermal. Two weeks after treatment, significant infiltrating lymphocytes and plasma cells were found around the surviving tumour cells. These morphological findings suggest that both cell-mediated and humoral immune responses could be responsible for the observed tumour eradication and induced long-term tumour resistance.

Measurement of x-ray attenuation coefficients of aqueous solutions of indocyanine green and glycated chitosan.

We report our experimental results of measurements of x-ray attenuation coefficients of aqueous solutions of a light absorbing dye, indocyanine green, and an immunoadjuvant, glycated chitosan. In the treatment of metastatic tumors in rats using a novel laser immunotherapy these solutions were administered in situ. The x-ray attenuation data of the solutions are essential to development of an x-ray digital imaging system for monitoring the administration of the solution, as well as for the distribution and the diffusion of the solution in tumors and in surrounding tissue. The composition of the solutions, the measurement system configuration, and the technique used to determine the attenuation coefficients are described. The experimental results show that glycated chitosan has a higher attenuation coefficient compared to indocyanine green and water. Our experimental data proved that, even at low concentrations, the x-ray attenuation through these aqueous solutions could be differentiated. Therefore, a digital x-ray imaging technique can be used effectively in monitoring and controlling the intratumor diffusion and distributions of these solutions.

Long-term tumor resistance induced by laser photo-immunotherapy.

An ideal treatment modality for metastasizing tumors should eradicate the primary tumor and elicit a systemic, tumor-selective response leading to elimination of metastases and long-term tumor resistance. Also, it should be induced by local treatment at the primary site, to limit adverse systemic effects. A new method for treating metastatic tumors which utilizes a combination of a near-infrared laser, a photosensitizer and an immunoadjuvant has been developed. It involves intra-tumor injection of the sensitizer/adjuvant solution, followed by local non-invasive laser irradiation. It has produced regression and total eradication of treated primary tumors and untreated metastases at remote sites against mammary tumors in rats. Successfully treated tumor-bearing rats showed total tumor resistance to subsequent tumor rechallenge. Our histochemical results showed that sera from cured tumor-bearing rats contained antibodies that bound strongly to the plasma membrane of both living and preserved tumor cells. Western blot analysis of tumor cell proteins using sera from successfully treated rats as the source of primary antibodies also showed distinct bands, indicating induction of tumor-selective antibodies. Our findings indicate that a systemic, long-term effect on metastatic tumors can be induced by local application of laser photo-immunotherapy.

Anatomy of mammalian conjunctival lymphoepithelium.

Ocular surface immune mechanisms are subservient to the fine function of the eye. A clear cornea with a smooth, well-lubricated facade is prerequisite to lucid vision. Hence, corneal inflammation and post-inflammatory scarring are intolerable, and the cornea contains a minimum of lymphoid elements. Although conjunctival dysfunction and consequent tear film deficiency can malign the corneal surface, conjunctival inflammation is tolerated to a considerable degree. In contrast to the human cornea, human conjunctiva contains an abundance of lymphoid tissue. Certain aspects of human conjunctival immunology elicit little debate. Langerhans cells are abundant in conjunctival epithelium. Isolated CD8+ suppressor/cytotoxic T cells predominate in conjunctival epithelium, while T cells in the substantia propria distribute equally between CD4+ T helper cells and CD8+ cells. Yet the presence of plasma cells in human conjunctiva, the expression of secretory component by human conjunctival epithelium, and the function of human conjunctival lymphoid follicles are in dispute. Confusion may derive in part from the use of inappropriate animal models; rodent conjunctiva does not appear to be a worthy facsimile for human conjunctiva. Discrepancies between different human studies likely result from variance in subject age, biopsy site and extent, histologic or histochemical technique, and perhaps the degree of inflammation present at the time of biopsy. Careful immunohistochemical and in situ molecular assays on well-defined loci within the conjunctiva of comparable human subjects may resolve such questions in the future. Organized mucosa-associated lymphoid tissue is rigorously defined as mucosal lymphoid follicles with an ultrastructurally distinct overlying lymphoepithelium. Based on available evidence, the epithelium overlying mammalian conjunctival lymphoid follicles does not contain distinct M cells. Whether zonal differences in morphology reflect real differences in the capacity to sample tear film antigens for presentation to the mucosal immune system remains to be established.

Comparative anatomy of mammalian conjunctival lymphoid tissue: a putative mucosal immune site.

Organized mucosa-associated lymphoid tissue (O-MALT) is defined by mucosal lymphoid follicles with unique overlying lymphoepithelia, and classically appears in tissues with a simple columnar epithelium. Within follicle-associated epithelium, goblet cells are characteristically absent, replaced by ultrastructurally distinct antigen-absorptive cells, termed M cells (or microfold cells) for the appearance of their apical cell membranes. To determine if mammalian conjunctiva, with its stratified squamous epithelium, can be considered as a site of O-MALT, we compared the light and electron microscopic anatomy of conjunctiva from fourteen species of non-human adult mammals, and the conjunctiva of human adults harvested at autopsy. Lymphoid follicles in the conjunctiva were demonstrated in all mammals studied except for mice and rats. In those mammals with conjunctival lymphoid follicles, the follicle-associated conjunctival epithelium was notable for an absence of goblet cells. Transmission electron microscopy demonstrated an intimate association of lymphocytes with surface epithelial cells, but epithelial cell morphology was uniform overlying the follicle, and other ultrastructural features of M cells were absent. Therefore, conjunctival lymphoid follicle-associated stratified squamous epithelium demonstrates some but not all features of O-MALT lymphoepithelia. Further studies are necessary to determine what role conjunctival lymphoid tissue may play in mucosal immunity.

Laser-photosensitizer assisted immunotherapy: a novel modality for cancer treatment.

Photosensitizer-enhanced laser treatment, where dyes are activated in situ by lasers of appropriate wavelengths, provides highly selective tissue destruction, both spatially and temporally, through photophysical reactions. Although laser-sensitizer treatment for cancer can achieve a controlled local tumor cell destruction on a large scale, total tumor eradication may not be accomplished because of the incomplete local tumor killing or the presence of tumor metastases, or both. The long-term control of cancer depends on the host immune surveillance and defense systems in which both cell-mediated and humoral responses are critical. In this study we report a novel minimally invasive cancer treatment combining the laser photophysical effects with the photobiological effects. Irradiation of a rat mammary tumor by an 805 nm diode laser, after an intratumor administration of a specific photosensitizer, indocyanine green in a glycated chitosan gel, caused immediate photothermal destruction of neoplastic cells. Concomitantly this treatment stimulated the immunological defense system against residual and metastatic tumor cells. Increases in survival rate and in the eradication of tumor burden, both primary and metastatic, were observed after this treatment. Furthermore, the resistance of successfully treated rats to tumor rechallenge demonstrated a long-lasting systemic effect of the treatment. These findings indicate that our treatment has triggered a specific humoral immune response in the tumor-bearing rats.

Effect of resin/fluoride and holmium:YAG laser irradiation on the resistance to the formation of caries-like lesions.

PURPOSE: This study compared the effects of a combination of a topical application of resin/NaF mixture and holmium-doped yttrium aluminum garnet (HO:YAG) laser energy with untreated control sites for their resistance to acid destruction/mechanical challenges of root surfaces adjacent to restorative margins. MATERIALS AND METHODS: Forty-eight extracted human teeth were prepared for restorations with coronal margins in enamel and the apical margins on root surfaces. Four types of restorative materials were used. One half of the restorative/root surface margins were treated with an application of nonfilled resin/fluoride mixture and holmium-doped yttrium aluminum garnet laser irradiation; the remaining portion served as the untreated control for each tooth. Acid (10% formic) and mechanical (air sandblaster) challenges were used to test the effects of these treated sites to their controls and to other types of restorative/root surface margins. The depth of root surface loss within 0 to 2 mm apical from the restorative margins was measured and evaluated after 24, 48, and 72 hours of acid exposures. RESULTS: The measurements of tooth loss for the 24- and 48-hour acid-exposure cycles were subjected to statistical analysis (ANOVA) using a paired t test for the variables. The variables, depth of tooth surface loss for the control sites was compared with the depth of loss of the treated sites on opposing sides of each tooth. The depth of tooth surface loss for the control sites had an average mean of 0.35 (0.25) mm for the 24-hour acid exposures, with the opposing treated sides having a mean of 0.053 (0.05) mm (significant at the 0.05 level). The 48-hour acid-exposure results showed significant increased tooth loss for the control sites, but minimum additional loss on the treated sites. The control sites showed an average mean of 1.16 (0.24) mm and a mean of 0.095 (0.103) mm for the treated areas, significant at the 0.05 level. No measurements were made for the 72-hour cycle groups of composite and amalgam alloy restorations because of the loss of most of the filling materials in the control sites. The control groups of crowns showed an average mean loss of 2.06 (0.37) mm with only 0.20 (0.19) mm surface loss for the treated sites. The paired t test for variables indicated no significant differences of tooth surface loss between various types of restorative margins used in this study. CONCLUSIONS: HO:YAG laser energy irradiation after application of resin/NaF to restorative margins and adjacent areas showed a significant increased resistance to acid/mechanical destruction on cementum-dentin root surfaces. The integrity of the restorative/dentin margins were maintained after extended exposures to formic acid and mechanical challenges.