Genetic induction of tumorigenesis in swine.

The transition from basic to clinical cancer research for a number of experimental therapeutics is hampered by the lack of a genetically malleable, large animal model. To this end, we genetically engineered primary porcine cells to be tumorigenic by expression of proteins known to perturb pathways commonly corrupted in human cancer. Akin to human cells, these porcine cells were quite resistant to transformation, requiring multiple genetic changes. Moreover, the transformed porcine cells produced tumors when returned to the isogenic host animal. The ability to now rapidly and reproducibly genetically induce tumors of sizes similar to those treated clinically in a large mammal similar to humans in many respects will provide a robust cancer model for preclinical studies dependent on generating large tumors.

Genetic control of susceptibility to experimental Lyme arthritis is polygenic and exhibits consistent linkage to multiple loci on chromosome 5 in four independent mouse crosses.

C3H/He mice infected with Borrelia burgdorferi develop severe arthritis and are high antibody responders, while infected C57BL/6 and BALB/c mice develop mild arthritis and less robust humoral responses. Genetic analysis using composite interval mapping (CIM) on reciprocal backcross populations derived from C3H/HeN and C57BL/6N or C3H/HeJ and BALB/cAnN mice identified 12 new quantitative trait loci (QTL) linked to 10 murine Lyme disease phenotypes. These QTL reside on chromosomes 1, 2, 4, 6, 7, 9, 10, 12, 14, 15, 16, and 17. A reanalysis of an F(2) intercross between C57BL/6N and C3H/HeN mice using CIM identified two new QTL on chromosomes 4 and 15 and confirmed the location of seven previously identified loci. Two or more experimental crosses independently verified six QTL controlling phenotypes after B. burgdorferi infection. Additionally, Bb2 on chromosome 5 was reproduced in four experimental populations and was linked to the candidate locus Cora1. Evidence of four distinct QTL residing within the 30-cM region of chromosome 5 encompassing the previously mapped Bb2 and Bb3 loci was shown by CIM. Interestingly, some alleles contributing to susceptibility to Lyme arthritis were derived from C57BL/6N and BALB/cAnN mice, showing that disease-resistant strains harbor susceptibility alleles.

Temporal and spatial evaluation of lesion reparative responses following superthreshold exposure of rat lung to pulsed ultrasound.

This study characterized the reparative responses in rat lung. Forty-five adult female rats were exposed at two sites over the left lung to 3.1-MHz superthreshold pulsed ultrasound. The repair of lung lesions was evaluated from 0 through 44 days postexposure. Macroscopic lesions at 0 days postexposure were large bright red ellipses of hemorrhage. By 1 and 3 days postexposure, lesions were the same size and dark red to red-black, but, by 3 days postexposure, lesions had a raised surface appearance. From 5 to 10 days postexposure, lesions grew smaller in size, progressed from red-gray to yellow-brown, and retained a raised surface appearance. From 13 through 44 days postexposure, lesions gradually decreased in size, had a faint yellow-brown discoloration, and gradually lost the raised surface appearance. By 37 and 44 days postexposure, lung returned to near normal morphology, but had small areas of light yellow-brown discoloration in the areas where lung was exposed. Microscopic lesions at 0 and 1 days postexposure were areas of acute alveolar hemorrhage. By 3 days postexposure, lesions had loss of alveolar erythrocytes and the formation of hemoglobin crystals. From 5 through 44 days postexposure, iron in degraded erythrocytes was processed to hemosiderin and was negligible in quantity at 44 days postexposure. The proliferation of resident cells (likely alveolar epithelial cells, fibroblasts and endothelial cells) and the infiltration of inflammatory cells in lesions declined in intensity as the lesions aged and was minimal by 44 days postexposure. Under the superthreshold exposure conditions described, lesions induced by ultrasound do not seem to have long-term residual effects in lung.

Quantifying B-mode images of in vivo rat mammary tumors by the frequency dependence of backscatter.

OBJECTIVE: To evaluate the frequency dependence of ultrasonic backscatter for its ability to differentiate between neoplastic and healthy tissue. METHODS: Standard B-mode images were created of 5 rats with spontaneous mammary tumors, and regions of interest in the lesion and surrounding tissue were parameterized by the slope of the backscatter amplitude versus frequency. RESULTS: In 4 of the 5 rats, the averaged backscatter slope of the regions of interest in the tumor was significantly (P < .05) different from that of the surrounding tissue, and the fifth case had a moderate difference (P = .20). The consistency of the averaged slope values (1.2-1.8 dB/MHz) across all but 1 of the mammary tumors was encouraging for the prospect of identifying a tissue type by its backscatter slope. CONCLUSIONS: This work suggests that characterization and diagnosis of tissue types may be possible by using ultrasonographic images quantified by the frequency dependence of backscatter.

Superthreshold behavior and threshold estimation of ultrasound-induced lung hemorrhage in adult mice and rats.

Threshold estimates and superthreshold behaviors for ultrasound-induced lung hemorrhage were investigated as a function of species (adult mice and rats) and ultrasound frequency (2.8 and 5.6 MHz). A total of 151 6-to-7-week-old female ICR mice and 160 10-to-11-week-old female Sprague-Dawley rats were randomly divided into two ultrasonic frequency groups, and further randomly divided into seven or eight ultrasonic peak rarefactional pressure groups. Each group consisted of about 10 animals. Animals were exposed to pulsed ultrasound at either 2.8-MHz center frequency (1-kHz PRF, 1.42-microsecond pulse duration) or 5.6-MHz center frequency (1-kHz PRF, 1.17-microsecond pulse duration) for a duration of 10 seconds. The in situ (at the pleural surface) peak rarefactional pressure levels ranged between 2.5 and 10.5 MPa for mice and between 2.3 and 11.3 MPa for rats. The mechanical index (MI) ranged between 1.4 and 6.3 at 2.8 MHz for mice and between 1.1 and 3.1 at 5.6 MHz for rats. The lesion surface area and depth were measured for each animal as well as the percentage of animals with lesions per group. The characteristics of the lesions produced in mice and rats were similar to those described in previous studies by our research group and others, suggesting a common pathogenesis in the initiation and propagation of the lesions at the gross and microscopic levels. The percentage of animals with lesions showed no statistical differences between species or between ultrasound frequencies. These findings suggest that mice and rats are similar in sensitivity to ultrasound-induced lung damage and that the occurrence of lung damage is independent of frequency. Lesion depth and surface area also showed no statistically significant differences between ultrasound frequencies for mice and rats. However, there was a significant difference between species for lesion area and a suggestive difference between species for lesion depth. The superthreshold behavior of lesion area and depth showed that rat lung had more damage than mouse lung, and the threshold estimates showed a weak, or lack of, frequency dependency, suggesting that the MI is not consistent with the observed findings.

Attenuation coefficient estimates of mouse and rat chest wall.

Attenuation coefficients of intercostal tissues were estimated from chest walls removed postmortem (pm) from 41 6-to-7-week-old female ICR mice and 27 10-to-11-week-old female Sprague-Dawley rats. These values were determined from measurements through the intercostal tissues, from the surface of the skin to the parietal pleura. Mouse chest walls were sealed in plastic wrap and stored at 4 degrees C until evaluated, and rat chest walls were sealed in Glad-Lock Zipper sandwich bags, and stored at -15 degrees C. When evaluated, chest wall storage time ranged between 1 and 2 days pm for mice and between 41 and 110 days pm for rats. All chest walls were allowed to equilibrate to 22 degrees C in a water bath prior to evaluation. For both mouse and rat intercostal tissues, the estimated frequency normalized attenuation coefficient was 1.1 dB/cm-MHz. In order to determine if there was an effect of storage time on estimates of attenuation coefficient, an independent experiment was conducted. The intercostal tissues from six mouse chest walls were evaluated at three time points (1, 22, and 144 days pm), and from six rat chest walls were evaluated at four time points (1, 22, 50, and 125 days pm). There was no difference in the estimated intercostal tissue attenuation coefficient as a function of time postmortem.

Superthreshold behavior of ultrasound-induced lung hemorrhage in adult mice and rats: role of pulse repetition frequency and exposure duration.

Superthreshold behavior for ultrasound-induced lung hemorrhage was investigated in adult mice and rats at an ultrasound center frequency of 2.8 MHz to assess the role of pulse repetition frequency and exposure duration. One hundred fifty, 6-7-week-old female ICR mice and 150 10-11-week-old female Sprague-Dawley rats were each divided into 15 exposure groups (10 animals per group) for a 3 x 5 factorial design (3 exposure durations of 5, 10, and 20 s and 5 pulse repetition frequencies of 25, 50, 100, 250, and 500 Hz). The in situ (at the pleural surface) peak rarefactional pressure of 12.3 MPa and the pulse duration of 1.42 micros were the same for all ultrasonically-exposed animals. In addition, 15 sham exposed mice and 15 sham exposed rats were included into both studies. In each study of 165 animals, the exposure conditions were randomized. The lesion depth and surface area were measured for each animal, as well as the percentage of animals with lesions per group. The characteristics of the lesions produced in mice and rats were similar to those described in studies by our research group and others, suggesting a common pathogenesis for the initiation and propagation of the lesions at the gross and microscopic levels. The proportion of lesions in both species was related statistically to pulse repetition frequency (PRF) and exposure duration (ED), with the exception that PRF in rats was not quite significant; the PRF x ED interaction (number of pulses) for lesion production was not significant for either species. The PRF, but not ED, significantly affected lesion depth in both species; the PRF x ED interaction for depth was not significant for either species. Both PRF and ED significantly affected lesion surface area in mice, while neither affected area in rats; the PRF x ED interaction for surface area was not significant for either species.

Cardiopulmonary function in rats with lung hemorrhage induced by pulsed ultrasound exposure.

OBJECTIVE: To assess cardiopulmonary function in rats exposed to pulsed ultrasound using superthreshold exposure conditions known to produce significant lung hemorrhage. METHODS: In 1 group of 9 anesthetized Sprague-Dawley rats, 5 foci of ultrasound-induced hemorrhage were produced in the left lung of each rat. In a second group of 6 rats, 5 foci of ultrasound-induced hemorrhage were produced in the left and right lungs of each rat. Each lesion was induced using superthreshold pulsed ultrasound exposure conditions (3.1-MHz center frequency, 1.7-kHz pulse repetition frequency, 1.3-micro-second pulse duration, 60-second exposure duration, 39-MPa in situ peak compressional pressure, and 17-MPa in situ peak rarefactional pressure). After exposure, the lungs were fixed in formalin and assessed histologically. The total lesion volume was calculated for each lesion in each lung lobe. Measurements of cardiopulmonary function included assessment of pulsatile arterial pressure, heart rate, end-tidal carbon dioxide, respiratory rate, and arterial blood gases (PCO2 and PO2). Functional data were quantified before (baseline) and 30 minutes after exposure to ultrasound. RESULTS: In the 9 rats that had lesions in only the left lung, the mean (SEM) lesion volume was 97 (13) mm3 and represented about 3.4% of the total lung volume. In the 6 rats that had lesions in both the left and right lungs, the left, right, and total mean lesion volumes, respectively, were 102 (16), 114 (11), and 216 (18) mm3 and represented about 3.7%, 4.2%, and 7.9% of the total lung volume. There were no statistically significant differences in cardiopulmonary measurements between baseline values and values obtained after exposure to ultrasound in the 9 rats exposed on the left lung only. The 6 rats exposed bilaterally had statistically significant differences in arterial pressure (134 +/- 4 versus 113 +/- 9 mm Hg; P= .047) and arterial PO2 (70 +/- 5 versus 58 +/- 4 mm Hg; P = .024) between baseline values and values obtained after exposure to ultrasound. CONCLUSIONS: The severity of ultrasound-induced lesions produced in 1 lung did not affect measurements of cardiopulmonary function because of the functional respiratory reserve in the unexposed lung. However, when both the left and right lungs had ultrasound-induced lesions, the functional respiratory reserve was decreased to a point at which rats were unable to maintain systemic arterial pressure or resting levels of arterial PO2.

Superthreshold behavior and threshold estimates of ultrasound-induced lung hemorrhage in adult rats: role of beamwidth.

It is well documented that ultrasound-induced lung hemorrhage can occur in mice, rats, rabbits, pigs, and monkeys. The objective of this study was to assess the role of the ultrasound beamwidth (beam diameter incident on the lung surface) on lesion threshold and size. A total of 144 rats were randomly exposed to pulsed ultrasound at three exposure levels and four beamwidths (12 rats per group). The three in situ peak rarefactional pressures were about 5, 7.5, and 10 MPa. The four 19-mm-diameter focused transducers had measured pulse-echo -6-dB focal beamwidths of 470 microm (2.8 MHz; f/1), 930 microm (2.8 MHz; f/2), 310 microm (5.6 MHz; f/1), and 510 microm (5.6 MHz; f/2). Exposure durations were 10 s, pulse repetition frequencies were 1 kHz, and pulse durations were 1.3 micros (2.8 MHz; f/1), 1.2 micros (2.8 MHz; f/2), 0.8 micros (5.6 MHz; f/1) and 1.1 micros (5.6 MHz; f/2). The lesion surface area and depth were measured for each rat as well as the percentage of rats with lesions per group. Logistic regression analysis and Gaussian-Tobit analysis methods were used to analyze the data. The effects of in situ peak rarefactional pressure and beamwidth were highly significant, but ultrasonic frequency was not significant. In addition, the estimated interaction between in situ peak rarefactional pressure and beamwidth was positive and highly significant. The ultrasound beamwidth incident on the lung surface was shown to strongly affect the percentage and size of ultrasound-induced lung hemorrhage lesions. Even though ultrasonic frequency was an experimental variable, it was not shown to affect the lesion percentage or size.

Meningeal osteosarcoma in a dog.

A meningeal osteosarcoma was diagnosed in a dog displaying neurologic signs compatible with a space-occupying cerebellar lesion. Gross lesions, restricted to the brain, consisted of a solitary, compressive mass attached to the dura mater overlying the left cerebellum. The mass was composed of single and multinucleated, atypical polygonal cells that lined or rested within lacuna surrounded by eosinophilic, mineralized matrix. The matrical component stained dark green-yellow to blue with Movat's pentachrome stain, deep blue to red with Heidenhain aniline blue stain, and brown-black with Von Kossa stain. Results of these stains were interpreted as tumor osteoid. Foci of dural mineralization and osseous metaplasia were present at the point of tumor attachment. The microscopic observations were interpreted as an osteosarcoma of extraskeletal origin. To our knowledge, these findings represent the first documented case of a meningeal osteosarcoma in a domestic animal species.

Inflammatory response to intramuscular implantation of polyacrylonitrile ultrafiltration probes in sheep.

Polyacrylonitrile is used in the manufacture of dialysis membranes. These membranes are fundamental to the functioning of implantable probes for microdialysis and ultrafiltration sampling of tissue fluids. Although in vivo experimentation using polyacrylonitrile has been reported to cause little inflammatory response when implanted subcutaneously, such information is not available for intramuscular implantation in sheep. The procaine and benzathine salts of penicillin are formulated for intramuscular injection. These salts of penicillin or the formulation excipients may cause inflammatory reactions. Use of polyacrylonitrile probes to draw samples from sites at which these formulations have been injected may be compromised by inflammation or direct interaction between formulation excipients and the dialysis membrane. The aim of this project was to describe tissue responses to intramuscular implantation of polyacrylonitrile in the presence and absence of either procaine or procaine plus benzathine salts of penicillin G. Each of 20 normal sheep was implanted with two ultrafiltration probes, one at the site of an injection of procaine or benzathine plus procaine penicillin G. Similar injections were also made at remote intramuscular sites. After 8, 9, and 11 days of the experiment, sheep were killed and the injection and implantation site muscle were excised and prepared for histopathological examination. The implantation of the probe alone caused greater inflammatory response than the injection of procaine or procaine plus benzathine penicillin G at remote intramuscular sites. The histopathological lesions were greatest where the implantation site was coupled with the injection of either formulation of penicillin G. Polyacrylonitrile may not be a suitable dialysis membrane material for intramuscular implantation in sheep.

Ultrasound-induced lung hemorrhage is not caused by inertial cavitation.

In animal experiments, the pathogenesis of lung hemorrhage due to exposure to clinical diagnostic levels of ultrasound has been attributed to an inertial cavitation mechanism. The purpose of this article is to report the results of two experiments that directly contradict the hypothesis that ultrasound-induced lung hemorrhage is caused by inertial cavitation. Elevated hydrostatic pressure was used to suppress the involvement of inertial cavitation. In experiment one, 160 adult mice were equally divided into two hydrostatic pressure groups (0.1 or 1.1 MPa), and were randomly exposed to pulsed ultrasound (2.8-MHz center frequency, 1-kHz PRF, 1.42-micros pulse duration, 10-s exposure duration). For the two hydrostatic pressure groups (80 mice each), 8 in situ peak rarefactional pressure levels were used that ranged between 2.82 and 11.8 MPa (10 mice/group). No effect of hydrostatic pressure on the probability of hemorrhage was observed. These data lead to the conclusion that lung hemorrhage is not caused by inertial cavitation. Also, the higher hydrostatic pressure enhanced rather than inhibited the impact of ultrasonic pressure on the severity (hemorrhage area, depth, and volume) of lesions. These counterintuitive findings were confirmed in a second experiment using a 2 x 5 factorial design that consisted of two ultrasonic pressure levels and five hydrostatic pressure levels (100 mice, 10 mice/group). If inertial cavitation were the mechanism responsible for lung hemorrhage, then elevated hydrostatic pressures should have resulted in less rather than more tissue damage at each ultrasonic pressure level. This further supports the conclusion that the pathogenesis of ultrasound-induced lung hemorrhage is not caused by inertial cavitation.

Identification of genetic loci controlling the characteristics and severity of brain and spinal cord lesions in experimental allergic encephalomyelitis.

Experimental allergic encephalomyelitis (EAE) is the principal genetically determined animal model for multiple sclerosis (MS), the major inflammatory disease of the central nervous system (CNS). Although genetics clearly play a role in susceptibility to MS, attempts to identify the underlying genes have been disappointing. Considerable variation exists between MS patients with regard to the severity of clinical signs, mechanism of demyelination, and location of CNS lesions, confounding the interpretation of genetic data. A mouse-human synteny mapping approach may allow the identification of candidate susceptibility loci for MS based on the location of EAE susceptibility loci. To date, 16 regions of the mouse genome have been identified that control susceptibility or clinical signs of EAE. In this work, we examined the genetic control of histopathological lesions of EAE in an F2 intercross population generated from the EAE susceptible SJL/J and EAE resistant B10.S/DvTe mouse strains. Composite interval mapping was used to identify 10 quantitative trait loci (QTL), including seven newly identified loci controlling the distribution and severity of CNS lesions associated with murine EAE. QTL on chromosome 10 control lesions in the brain, whereas QTL on chromosomes 3, 7, and 12 control lesions in the spinal cord. Furthermore, sexually dimorphic QTL on chromosomes 2, 9, and 11 control CNS lesions in females, whereas QTL on chromosomes 10, 11, 12, 16, and 19 control lesions in males. Our results suggest that the severity and location of CNS lesions in EAE are genetically controlled, and that the genetic component controlling the character and severity of the lesions can be influenced by sex.

Genetic analysis of the influence of pertussis toxin on experimental allergic encephalomyelitis susceptibility: an environmental agent can override genetic checkpoints.

Pertussis toxin (PTX) is a potent ancillary adjuvant used to elicit several different autoimmune diseases, including experimental allergic encephalomyelitis (EAE). To delineate the genetics of PTX effect in EAE, we mapped EAE-modifying (eae-m) loci in cohorts of backcross mice immunized with and without PTX. In this study, we analyzed the genetic basis of EAE susceptibility and severity and the intermediate phenotypes of mononuclear cell infiltration, suppuration, and demyelination. In animals immunized with PTX, one major locus, eae9, controls disease susceptibility and severity. Eae9 also regulates the extent of mononuclear cell infiltration of the spinal cord in male mice. Without PTX, five eae-m loci were noted, including three new loci in intervals on chromosomes 8 (eae14), 10 (eae17), and 18 (eae18). Taken together, these results suggest that eae9 controls the effects of PTX in EAE susceptibility, and is capable of overriding the other genetic checkpoints in the pathogenesis of this disease.

Mice deficient in interleukin-1beta converting enzyme resist anorexia induced by central lipopolysaccharide.

Interleukin-1beta (IL-1beta) is expressed in the mouse brain after intracerebroventricular injection of lipopolysaccharide (LPS) and is thought to be responsible for many of the behavioral and neuroendocrine changes that occur during inflammation. In this study we show that LPS in the brain also induces expression of interleukin-1beta converting enzyme (ICE) and that ICE is important for the characteristic anorectic response of mice to intracerebroventricular LPS. Specifically, mice that were deficient in ICE (ICE(-/-)) resisted the anorexia caused by intracerebroventricular injection of LPS but were sensitive to the anorectic properties of recombinant IL-1beta. The typical anorectic response seen in wild-type (WT) mice after LPS was restored in ICE(-/-) mice by intracerebroventricular administration of the ICE analog cathepsin G. Conversely, anorexia induced by intracerebroventricular injection of LPS in WT mice was blocked by prior intracerebroventricular injection of the ICE antagonist YVAD. CMK. Furthermore, in situ hybridization immunohistochemistry revealed intense expression of ICE mRNA in the hippocampus and dorsomedial hypothalamus of WT mice after intracerebroventricular injection of LPS. Thus ICE mRNA is expressed in brain after intracerebroventricular injection of LPS and is important for induction of anorexia, presumably because it generates mature IL-1beta. These results suggest that preventing generation of mature IL-1beta can inhibit anorexia induced by LPS in the brain and, therefore, reveal ICE as a potential target for regulating food intake during brain inflammation.

Dual role of interleukin-10 in murine Lyme disease: regulation of arthritis severity and host defense.

In the murine model of Lyme disease, C3H/He mice exhibit severe arthritis while C57BL/6N mice exhibit mild lesions when infected with Borrelia burgdorferi. Joint tissues from these two strains of mice harbor similar concentrations of B. burgdorferi, suggesting that the difference in disease severity reflects differences in the magnitude of the inflammatory response to B. burgdorferi lipoproteins. Stimulation of bone marrow macrophages from C3H/HeN mice with the B. burgdorferi lipoprotein OspA resulted in higher-level production of the inflammatory mediators tumor necrosis factor alpha, nitric oxide, and interleukin-6 (IL-6) than that of macrophages from C57BL/6N mice. In contrast, macrophages from C57BL/6N mice consistently produced larger amounts of the anti-inflammatory cytokine IL-10 than did C3H/HeN macrophages. Addition of recombinant IL-10 suppressed the production of inflammatory mediators by macrophages from both strains. IL-10 was found to modulate B. burgdorferi-induced inflammation in vivo, since C57BL/6J mice deficient in IL-10 (IL-10-/-) developed more severe arthritis than wild-type C57BL/6J mice. The increase in arthritis severity was associated with a 10-fold decrease in the number of B. burgdorferi organisms present in ankle tissues from IL-10-/- mice. These findings suggest that in C57BL/6 mice, IL-10-dependent regulation of arthritis severity occurs at the expense of effective control of bacterial numbers.

Sequence polymorphisms in the chemokines Scya1 (TCA-3), Scya2 (monocyte chemoattractant protein (MCP)-1), and Scya12 (MCP-5) are candidates for eae7, a locus controlling susceptibility to monophasic remitting/nonrelapsing experimental allergic encephalomyelitis.

Experimental allergic encephalomyelitis (EAE), the principal animal model of multiple sclerosis, is genetically controlled. To date, 13 disease-modifying loci have been identified in the mouse by whole genome scanning using an F2 intercross between EAE-susceptible SJL/J and EAE-resistant B10.S/DvTe mice. Two quantitative trait loci (QTL), eae6 and eae7, on chromosome 11 were identified by classical marker-specific linkage analysis and interval mapping. Both QTL were reported to be associated with severity and duration of clinical signs. eae7 was subsequently shown to be a unique locus controlling the development of monophasic remitting/nonrelapsing EAE. In this study, composite interval mapping resolved eae6 into two linked QTL: eae6a at 0-13 cM is associated with disease severity, and eae6b at 19-28 cM associated with the duration of clinical signs. Additionally, composite interval mapping significantly refined the locations of eae6a, eae6b, and eae7, thereby facilitating systematic candidate gene screening by cDNA sequencing of SJL/J and B10.S/DvTe alleles. Sequence polymorphisms were not seen in Lif and IL12 beta, candidate genes for eae6a and eae6b, respectively. Similarly, cDNA sequence polymorphisms in Nos2, Scya3, Scya4, Scya5, Scya6, Scya7, Scya9, Scya10, and Scya11 were excluded as candidates for eae7. However, multiple sequence polymorphisms resulting in significant amino acid substitutions were identified in Scya1 (TCA-3), Scya2 (monocyte chemoattractant protein (MCP)-1), and Scya12 (MCP-5). Given the role of chemokines in EAE, these sequence polymorphisms are promising candidates for eae7, a locus associated with severity of clinical signs and susceptibility to the shorter, less severe monophasic remitting/nonrelapsing form of disease.

Expression and localization of p80 and p68 interleukin-1 receptor proteins in the brain of adult mice.

The biological effects of interleukin-1 (IL-1) are mediated by two distinct receptors, the p80 type I IL-1 and p68 type II IL-1 receptor proteins (IL-1RI and IL-1RII, respectively), both of which have been recently co-localized to the growth hormone synthesizing cells of the adenohypophysis. Previous studies have shown that IL-1 can bind to specific structures in the central nervous system, but the distribution of IL-1RI and IL-1RII proteins in the adult mouse brain has not been reported. Here we have used immunohistochemistry to study the expression, distribution and cellular localization of both isoforms of the IL-1 receptor proteins in the adult mouse brain. Using a combination of processing techniques (AMeX fixation and cryosectioning), we have immunolabeled brain sections for each isoform of the IL-1R. Both isoforms are expressed in the CNS, particularly in neuronal soma of the granular layer of the dentate gyrus and pyramidal cells of fields CA1-CA4 of Ammon's horn of the hippocampus, in epithelial cells of the choroid plexus and ependymal layer, and in neuronal soma of Purkinje cells of the cerebellum. The IL-1RII isoform, but not IL-1RI, is expressed in specific neuronal soma and proximal cell processes of neurons of the paraventricular gray matter of the hypothalamus. These immunohistochemical data directly demonstrate the neuronal expression of both IL-1R proteins in situ. The distribution and cellular localization of IL-1R proteins in the CNS provide a molecular basis for understanding reciprocal interactions between the immune system and the brain.

Genetic analysis of disease subtypes and sexual dimorphisms in mouse experimental allergic encephalomyelitis (EAE): relapsing/remitting and monophasic remitting/nonrelapsing EAE are immunogenetically distinct.

Experimental allergic encephalomyelitis (EAE) is the principal animal model of multiple sclerosis (MS), the major inflammatory disease of the central nervous system. Murine EAE is generally either an acute monophasic or relapsing disease. Because the clinical spectrum of MS is more diverse, the limited range of disease subtypes observed in EAE has raised concern regarding its relevance as a model for MS. During the generation of a large F2 mapping population between the EAE-susceptible SJL/J and EAE-resistant B10.S/DvTe inbred lines, we identified four distinct subtypes of murine EAE resembling clinical subtypes seen in MS. We observed acute progressive, chronic/nonremitting, remitting/relapsing, and monophasic remitting/nonrelapsing EAE. An additional subtype, benign EAE, was identified after histologic examination revealed that some mice had inflammatory infiltrates of the central nervous system, but did not show clinical signs of EAE. Genome exclusion mapping was performed to identify the loci controlling susceptibility to each disease subtype. We report three novel EAE-modifying loci on chromosomes 16, 7, and 13 (eae11-13, respectively). Additionally, unique loci with gender-specific effects govern susceptibility to remitting/relapsing (eae12) and monophasic remitting/nonrelapsing (eae7 and 13) EAE.

Identification of quantitative trait loci governing arthritis severity and humoral responses in the murine model of Lyme disease.

A spectrum of disease severity has been observed in patients with Lyme disease, with approximately 60% of untreated individuals developing arthritis. The murine model of Lyme disease has provided strong evidence that the genetic composition of the host influences the severity of arthritis following infection with Borrelia burgdorferi: infected C3H mice develop severe arthritis while infected C57BL/6N mice develop mild arthritis. Regions of the mouse genome controlling arthritis severity and humoral responses during B. burgdorferi infection were identified in the F2 intercross generation of C3H/HeNCr and C57BL/6NCr mice. Rear ankle swelling measurements identified quantitative trait loci (QTL) on chromosomes 4 and 5, while histopathological scoring identified QTL on a unique region of chromosome 5 and on chromosome 11. The identification of QTL unique for ankle swelling or histopathological severity suggests that processes under distinct genetic control are responsible for these two manifestations of Lyme arthritis. Additional QTL that control the levels of circulating Igs induced by B. burgdorferi infection were identified on chromosomes 6, 9, 11, 12, and 17. Interestingly, the magnitude of the humoral response was not correlated with the severity of arthritis in infected F2 mice. This work defines several genetic loci that regulate either the severity of arthritis or the magnitude of humoral responses to B. burgdorferi infection in mice, with implications toward understanding the host-pathogen interactions involved in disease development.