Changes in nasal mucosal innervation in horses with grass sickness

REASONS FOR PERFORMING STUDY: Equine grass sickness is a dysautonomia characterised by widespread destruction of autonomic ganglia, resulting in the clinical signs of dysphagia, constipation, profuse sweating, tachycardia, rhinitis sicca and high mortality rate. Rhinitis sicca is a common finding in horses with the chronic form and we have postulated that alterations in autonomic innervation of the nasal mucosa might underlie this clinical presentation. OBJECTIVES: In this study, the expression and distribution of nerve fibres immunoreactive for calcitonin gene-related peptide (CGRP), substance P (SP), the general neuronal marker protein gene-product 9.5 (PGP 9.5; ubiquitin) and the intermediate neurofilaments (PAN-N; neurorfilaments L, M and H) in the nasal mucosa of normal horses (n = 10) and horses with EGS (n = 18; acute n = 8, subacute n = 3, chronic n = 7) was assessed. METHODS: Innervation density and distribution was investigated in the different groups using standard immunohistochemical techniques. RESULTS: Significant differences were noted when comparing the density and distribution of nerve fibres immunoreactive for PGP 95 and PAN-N, with PGP 95 consistently giving better staining in all groups and at all sites in the nasal mucosa. An apparent increase in the density of innervation was noted for acute vs. normal cases. A significant reduction in the density of innervation was noted only with PAN-N when comparing normal horses and acute cases with the chronic group (P < 0.05). CGRP and SP immunoreactive nerve fibres were typically most abundant in the epithelial and subepithelial layers, but the quality of staining and nerve fibre density was greater for SP, achieving significant difference in several comparisons. The density of innervation for SP was significantly reduced in the chronic group compared to the normal and acute groups (P < 0.01). A significant decrease was noted for CGRP only for the acute and chronic groups (P<0.05). CONCLUSIONS: These results demonstrate a reduction in the expression of the sensory neuropeptides in nasal mucosal innervation as a consequence of equine dysautonomia, and may underlie the clinical presentation of rhinitis sicca noted with this disease. POTENTIAL RELEVANCE: Nasal biopsy may be of use in antemortem diagnosis of grass sickness and identification of mucosal denervation; and might also be useful in the treatment of rhinitis in EGS cases.     

Alveolar mucosal approach to the canine nasal cavity

A nine-year-old, intact female Afghan hound was presented for evaluation of an intermittent, mucopurulent, unilateral nasal discharge with a three-year duration. Radiographs showed the ipsilateral canine tooth within the rostral nasal cavity. The tooth was removed through an alveolar mucosal rhinotomy. There has been no recurrence of the nasal discharge or complications associated with the surgical procedure during the 20-month follow-up period.     

3-Methylindole-induced pulmonary injury in lambs. A comparison with acute respiratory distress syndrome (ARDS)

Five to six month-old lambs of two different Norwegian breeds (Dala and Rygja) were dosed with 3-methyl indole (3MI) and DL-tryptophan (DL-T), and examined clinically, hematologically and post mortem. Results were compared with the findings in Acute Respiratory Distress Syndrome (ARDS), a condition of acute dyspnea occurring in 5-6 month-old lambs (of Dala and Spael breeds) after they have been moved from mountain pastures onto lush aftermath in September. The results show that such young Norwegian lambs are susceptible to 3MI-dosing, and that the symptoms and lesions were similar to those described earlier in connection with 3MI-dosing in cattle and adult sheep. The results clearly demonstrate that 3-MI-induced pulmonary injury in lambs differs from ARDS as far as both symptoms, hematology and pathology are concerned.     

Chemical modulation of 3-methylindole toxicosis in mice: effect on bronchiolar and olfactory mucosal injury

C57BL/6N mice were treated to induce tolerance, to modulate the mixed function oxidase system or to deplete glutathione (GSH) before injection with 400 mg 3-methylindole (3MI)/kg. Effect of pretreatment was determined by histologic comparison of pulmonary and nasal lesions 24 hours after 3MI. beta-Naphthoflavone and 3MI pretreatment significantly decreased 3MI-induced bronchiolar epithelial damage in male and female mice, while phenobarbital protection was significant only in female mice. Only beta-naphthoflavone decreased nasal olfactory epithelial damage. Pretreatment with piperonyl butoxide, SKF 525-A, or alpha-naphthoflavone had no significant effect on development of lesions. Diethylmaleate pretreatment significantly increased mortality and bronchiolar damage in both sexes. Significant differences between male and female mice were not detected in any group. The results suggest that pretreatment with low doses of 3MI or induction of cytochrome P-448 or P-450 protects against 3MI toxicosis while GSH depletion increases mortality and pulmonary lesions.     

Isolation and characterization of equine nasal mucosal CD172a+ cells

The nasal mucosa surface is continuously confronted with a broad variety of environmental antigens, ranging from harmless agents to potentially harmful pathogens. This area is under rigorous control of professional antigen presenting cells (APCs), such as dendritic cells (DCs) and macrophages. Mucosal APCs play a crucial role in inducing primary immune responses and the establishment of an immunological memory. In the present study, a detailed characterization of CD172a⁺ cells, containing the APCs residing in the equine nasal mucosa was performed for the first time. CD172a⁺ cells were isolated from collagenase-treated equine nasal mucosa fragments by MACS. Expression of surface markers was determined by flow cytometry and functional analysis was done by measuring the uptake of FITC conjugated ovalbumin (FITC-OVA). Cell surface phenotype of the isolated cells was as follows: 90% CD172a⁺, 30% CD1c⁺, 46% CD83⁺, 42% CD206⁺ and 28% MHC II⁺. This clearly differs from the phenotype of blood-derived monocytes: 96% CD172a⁺, 4% CD1c⁺, 11% CD83⁺, 9% CD206⁺, 72% MHC II⁺ and blood monocyte derived DCs: 99% CD172a⁺, 13% CD1c⁺, 30% CD83⁺, 51% CD206⁺ and 93% MHC II⁺. The CD172a⁺ nasal mucosal cells were functionally able to endocytose FITC-OVA but to a lesser degree than monocyte-derived DCs. Together, these results demonstrate that the isolated CD172a⁺ nasal mucosal cells resemble immature DCs in the nasal area.     

Immunoglobulin isotypes in sera and nasal mucosal secretions and their neonatal transfer and distribution in horses

OBJECTIVE: To determine concentrations of IgA and IgG subclasses in serum, colostrum, milk, and nasal wash samples of adult horses and foals. ANIMALS: Seven 2-year-old Welsh ponies, 27 adult mixed-breed horses, and 5 Quarter Horse mares and their foals. PROCEDURE: Serum was obtained from ponies and adult horses. Colostrum and milk were obtained from mares and serum and nasal wash samples from their foals immediately after parturition and on days 1, 7, 14, 28, 42, and 63. Nasal wash samples were also obtained from 23 adult horses. Concentrations of immunoglobulins were determined by use of inhibition ELISA. To determine transfer of maternal isotypes to foals, concentrations in colostrum and milk were compared with those in foal serum. Serum half-lives of isotypes in foals were also determined. RESULTS: IgGb was the most abundant isotype in serum and colostrum from adult horses, whereas IgA was the predominant isotype in milk. The major isotype in nasal secretions of adult horses and foals > or = 28 days old was IgA, but IgGa and IgGb were the major isotypes in nasal secretions of foals < or = 14 days old. Serum half lives of IgGa, IgGb, IgG(T), and IgA in foals were 176, 32, 21, and 3.4 days, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: The early immunoglobulin repertoire of neonatal foals comprised IgGa, IgG(T), and IgA; endogenous synthesis of IgGb could not be detected until 63 days after birth. The restricted repertoire of immunoglobulins in foals may influence humoral immune responses to vaccination.     

Taenia taeniaeformis larval product induces gastric mucosal hyperplasia in SCID mice

The effects of intraperitoneal implantation of Taenia taeniaeformis larvae and inoculation of in vitro larval products on gastric mucosa of SCID mice were investigated in this study. Mice surgically implanted with T. taeniaeformis larvae developed slight and moderate gastric hyperplasia. When in vitro cultured T. taeniaeformis larval excretory-secretory (TtLES) products containing 1 mg of protein were injected daily into mice, they caused gastropathy after 5-7 days. Mice injected daily with 0.5 mg of TtLES products also showed slight gastric hyperplasia after day 14 and 28. The gastropathy was characterized by reduction of both parietal and zymogenic cell number and increased number of alcian blue-periodic acid Schiff (AB-PAS)-positive cells and by two-fold extension of proliferative zone of gastric units. Larval implantation demonstrated a more potent effect in inducing gastropathy than did in vitro larval culture products. Significant decrease in number of parietal cells with concomitant increase of proliferative zone and AB-PAS-positive cell number indicated their important roles in inducing the hyperplastic lesion. Similarities with other gastropathies indicated that there is a common fundamental regulatory mechanism involved, and that the host response may not be specific to parasites. Present study validated the induction of gastric mucosal hyperplasia by larval ES products of T. taeniaeformis. This proved the hypothesis of previous studies suggesting the role of larvae-derived products in inducing gastric mucosal hyperplasia in T. taeniaeformis-infected rats.     

Experimental oral and nasal transmission of rabies virus in mice.

Weanling female white Swiss mice were exposed to challenge virus standard rabies virus and street virus isolates from various domestic and wild animals. Virus was given free choice as suspension or as infected mouse brain by stomach tube, by single injection of suspension into the oral cavity of unanesthetized mice, by repeated injection into the oral cavity of anesthetized mice and by single application to the external nares of anesthetized mice. Challenge virus standard virus in mouse brain suspension and a suspension of skunk salivary glands infected with street virus (titers greater than or equal to 10(6)MICLD50/0.03 ml) consistently produced high rates of infection in mice exposed intranasally, low to high rates of infection in mice exposed by forced feeding and other artificial methods of oral exposure and very low rates of infection when given free choice. Street virus isolates passaged intracerebrally in mice had titers less than or equal to 10(4.5) MICLD50/0.03 ml and rarely caused rabies in mice exposed orally or nasally by any method. The results indicate that with the isolates used, virus of high titer (greater than or equal to 10(6)MICLD50/0.03 ml) is required to consistently produce infection in mice by the nasal route and that the mucosa of the nasal cavity probably is the chief route of infection even after oral administration.     

3-methylindole induces transient olfactory mucosal injury in ponies

Response to 3-methylindole (3MI) varies among species. Mice recover from 3MI-induced bronchiolar epithelial injury but sustain persistent olfactory mucosal injury with scarring and epithelial metaplasia. In contrast, 3MI induces obliterative bronchiolitis in horses and ponies, but olfactory mucosal injury has not been reported. To evaluate the effect of 3MI on equine olfactory mucosa, ponies were dosed orally with 100 mg 3MI/kg (n = 9) or corn oil vehicle (n = 6). All ponies treated with 3MI developed obliterative bronchiolitis with mild olfactory injury. By 3 days after 3MI dosing, olfactory epithelium appeared disorganized with decreased and uneven surface height and scalloping of the basement membrane zone. Epithelial cells of Bowman's glands were hypertrophic. Proliferation of olfactory epithelium and Bowman's glands was supported by an increased mitotic index and positive immunohistochemical staining for proliferating cell nuclear antigen as compared with controls. The activity of 11beta-hydroxysteroid dehydrogenase, an olfactory mucosal cytosolic enzyme localized to sustentacular and Bowman's glandular epithelial cells, was concurrently decreased. By 9 days postdosing, olfactory mucosal lesions had lessened. Results indicate that 3MI transiently injures equine olfactory mucosa without the extensive necrosis, scarring, or metaplasia seen in murine olfactory mucosa or in equine bronchiolar epithelium.     

淫羊藿苷对辐射损伤小鼠保护作用研究

目的：探讨淫羊藿苷（ICA）对辐射损伤小鼠的保护作用。方法：将50只昆明小鼠按体重随机分为5组,3个剂量组分别按20、80、160 mg/kg体质量给小鼠灌服淫羊藿苷水溶液,正常对照组和照射对照组给予同体积生理盐水,1次/d,连续灌胃16 d。第6天照射对照组和3个剂量组小鼠用5.0 Gy60Co-γ射线进行单次全身照射。照射后第10天检测小鼠的血液学指标、脏器系数及MDA含量、SOD和GSH-Px活性。结果：与照射对照组比较,高剂量组的白细胞和GSH-Px活性显著增加（P〈0.05）,3个剂量组的血小板显著升高（P〈0.05）,SOD含量显著降低（P〈0.05）。结论：淫羊藿苷对小鼠辐射造成的损伤具有保护作用。     

Expression of vasoactive intestinal peptide, calcitonin gene-related peptide, substance P, and intermediate neurofilaments in nasal mucosal nerve fibers of horses without nasal disease

OBJECTIVE: To determine the distribution of nerve fibers containing calcitonin gene-related peptide (CGRP), substance P (SP), vasoactive intestinal peptide (VIP), and intermediate neurofilaments in nasal mucosa of horses. ANIMALS: 6 horses without evidence of nasal disease. PROCEDURE: Full-thickness nasal tissue specimens were obtained from the rostral portion of the nasal septum at necropsy, and fluorescence immunohistochemistry was performed to assess mucosal distribution of nerve fibers. RESULTS: Nerve fibers with CGRP-like immunoreactivity (CGRP-Li) formed a dense subepithelial network, and a large number of fibers were found coursing between epithelial cells. Fibers with CGRP-Li were also associated with blood vessels and mucous glands. Fibers with SP-like immunoreactivity (SP-Li) had a similar distribution and density. In contrast, there were few fibers with VIP-like immunoreactivity. Fibers containing intermediate neurofilaments were prominent and appeared as large nerve fiber bundles mainly adjacent to the nasal septum but also close to mucous glands and within the lamina propria. Intermediate neurofilaments were also identified in single nerve fibers at all sites, but the density of fibers with intermediate neurofilaments did not match that of fibers with CGRP- or SP-Li. CONCLUSIONS: The density and distribution of nerve fibers containing SP- or CGRP-Li in nasal mucosa of horses was similar to that reported for other species. However, expression of VIP in nerve fibers was low. Antibodies against intermediate neurofilaments identified many nerve fibers in nasal mucosa of horses but did not appear to identify small diameter fibers expressing SP or VIP.     

Brain viral persistence and myelin damage in nude mice.

Multiple foci of secondary demyelination were observed in the cerebellum of nude mice given two inoculations of avirulent Semliki Forest virus compared to single lesions seen in similarly treated Swiss A2G mice. The increased number of demyelinative plaques were attributed to brain viral persistence for 35 days in the nude mice with correspondingly low serum antibody titres. No neurological signs were observed in any of the mice.     

Maximum volume of nasal administration using a mucosal atomization device without aspiration in Japanese White rabbits

Recently, a mucosal atomization device (MAD) has been applied in veterinary medicine. In the present study, the maximum volume of nasal atomization without aspiration using MAD was examined in eight healthy female Japanese White (JW) rabbits. Each rabbit had their head and neck examined by computed tomography before and after nasal atomization with four different doses (0.15, 0.3, 0.45, and 0.6 ml per nostril) of diluted contrast medium (1:2 mixture of iohexol and saline). This was done under general anesthesia by an intramuscular administration of alfaxalone 2.5 mg/kg, medetomidine 40 μg/kg, and butorphanol 0.4 mg/kg, with a 7-day washout period between each treatment. The diluted contrast medium was distributed in the nasal cavity, external nares, and/or oral cavity in all rabbits receiving each treatment. The intranasal distribution volumes of the contrast medium were 287 (250–333) mm³ [median (interquartile range)] for 0.15 ml, 433 (243–555) mm³ for 0.3 ml, 552 (356–797) mm³ for 0.45 ml, and 529 (356–722) mm³ for 0.6 ml of treatment. The intranasal distribution volume for 0.15 ml treatment tended to be lower than that for 0.6 ml treatment (P=0.083). The contrast medium was deposited in the trachea in one rabbit (12.5%) and four rabbits (50%) receiving treatments of 0.45 and 0.6 ml per nostril, respectively. The maximum volume of nasal atomization without aspiration into the trachea was 0.3 ml per nostril for the JW rabbits.     

Lectin histochemistry for sugars on the mucosal surface of the uterus in pregnant mice

Expression patterns of sugars on the mucosal surface of the uterus in pregnant mice were investigated by using 21 kinds of lectins. In the uterine mucosa, the GlcNAc group tended to express a positive reaction before pregnant day 10, but the glucose/mannose group generally expressed a positive reaction after pregnant day 10. On the other hand, the fucose group expressed a negative reaction during all periods in pregnancy. These findings were almost the same on both the mesometrial side and anti-mesometrial side of the uterus. These differences of sugar expression probably reflect the functional change of the mucosa during pregnancy and the alteration of sugar expression may give a chance for pathogens to infect in the uterus with limited periods.     

Hermaphroditism in 3 chimeric mice

Hermaphroditism was diagnosed in three, 6-month-old, male, chimeric mice generated by microinjection of 129/Ola XY recombinant embryonic stem cells into unsexed C57BL/6 blastocysts. Grossly, mice Nos. 1 and 2 had perigenital masses and hydrometra. All mice had unilateral ovaries and cystic endometrial hyperplasia. Mice Nos. 1 and 3 also had contralateral testes and epididymides. Histologically, mice Nos. 1 and 3 were true hermaphrodites with unilateral ovotestes, while mouse No. 2 was a pseudohermaphrodite with ovarian tissue only. The presence of a uterus with cystic endometrial hyperplasia in these mice resembles XY pseudohermaphroditism in miniature schnauzers. The mice were determined to be 95 to 100% chimeric via haircoat color; however, the presence of both male and female sex organs in these phenotypically male mice suggests otherwise. Published reports note incidences for sex chimeras and hermaphroditism in genetically engineered mice of 50% and 20%, respectively. Hermaphroditism is expected to increase as the numbers of chimeric mice rise with technical advances in genetic engineering.     

Extensive nasal septum resection in horses using a 3-wire method

OBJECTIVE: To describe a surgical technique for, and report outcome after, extensive nasal septum resection in horses with congenital or acquired septal diseases. STUDY DESIGN: Retrospective study. ANIMALS: Horses (5) with nasal septal deformity. METHODS: Under general anesthesia, obstetrical wires were preplaced around the ventral, caudal, and dorsal aspects of the nasal septum by combinations of manual guidance (ventral and caudal wires) and use of a trephine hole on the dorsal mid-line (dorsal and caudal wires). The rostral part of the septum was incised with a scalpel and the ventral, dorsal, and caudal septal incisions were made with the preplaced wires. After septum removal, the nasal cavity packed with a tampon and a temporary tracheotomy was performed. RESULTS: The diseased nasal septum was completely removed in all horses. It was possible to direct the caudal cut at a sufficient angle so that the remaining stump was in a wide part of the nasal passage, where it would have minimal effect on airflow. Four horses healed, and returned to intended use but continued to make a slight respiratory noise during exercise. At 13 months after surgery, 1 horse with a concurrent wry nose was retired from race training after 2 additional surgeries failed to relieve exercise intolerance at high speed. CONCLUSIONS: The 3 wire-cut method of nasal septum resection was technically easy to perform and safe, allowed return to function in most horses, and produced an excellent cosmetic appearance. CLINICAL RELEVANCE: This technique for nasal septum resection allows removal of a large portion of the nasal septum and provides a safe alternative to conventional techniques of nasal septum resection in the horse.     

Dietary nucleotides protect thymocyte DNA from damage induced by cyclophosphamide in mice

The effects of dietary nucleotides on thymocyte DNA damages induced by cyclophosphamide (CP) in mice were examined. First, phase I experiment was conducted to determine the optimal timing of detecting thymocyte DNA damages induced by CP (150 mg/kg body weight) in mice. Thymocyte DNA damages was determined at 6, 12, 18, 24 h by single-cell gel electrophosphoresis assay (comet assay) after intraperitoneal injection of CP. The levels of DNA damage at 6, 12, 18, 24 h were all significantly higher than that of the control group (p < 0.01). The highest level of DNA damage appeared at 18 h and then decreased at 24 h. Therefore, 18 h was selected to determine DNA damages induced by CP in subsequent experiments. In phase II experiment, 30 male KunMing mice were divided into three treatments: negative control (NC), positive control (PC) and nucleotides group (NG). Mice in NC and PC were fed nucleotide-free diet, and mice in NG were fed nucleotide-supplemented diet (supplemented with 0.25% nucleotides, a mixture containing equal amounts of AMP, CMP, GMP and UMP). Mice in PC and NG groups were injected with CP (150 mg/kg body weight) at 21 days. DNA damage in thymocytes was evaluated at 18 h after CP treatment. The results indicate that dietary nucleotides do not affect the weights of the thymus and the spleen, or their organ indices (p > 0.05), but significantly decrease the percentage of comet cells and comet tail sizes (p < 0.01). This study demonstrates that dietary nucleotides could reduce the level of thymocyte DNA damage induced by CP in mice.     

对氨基苯胂酸对小鼠肝脏损伤作用试验

对氨基苯胂酸(arsanilic acid),又称阿散酸,是一种广泛用于畜禽饲料的添加剂,对畜禽的生长发育、生产性能等有明显的促进作用,并可防治畜禽的某些疾病.本实验室在之前的体外研究发现:对氨基苯胂酸能抑制体外培养的大鼠原代肝细胞的增殖,并能诱导肝细胞凋亡,其诱发肝细胞凋亡的初步机理是抑制细胞抗氧化酶的活性,并促进凋亡蛋白Bax的表达升高,对氨基苯胂酸诱导肝细胞凋亡的最小浓度范围在50～500μmol/L之间.     

绞股蓝总皂甙对衰老小鼠DNA损伤的影响

目的:本试验旨在探讨绞股蓝总皂甙(GPS)对D-半乳糖亚急性衰老模型小鼠DNA损伤的影响。方法:将200只健康昆明系小鼠随机分为5组,每组40只(雌雄各半)。一组为空白对照组,每天每只灌胃和颈背部皮下注射生理盐水0.1ml;二组为D-半乳糖衰老模型组,按120mg/kg每天每只颈背部皮下注射D-半乳糖(用时用生理盐水配成5%的溶液)0.1ml,同时灌胃生理盐水0.1ml;三、四、五组为绞股蓝总皂甙(GPS)组,每天每只灌胃GPS溶液(GPS的剂量分别为180mg/kg、120mg/kg、60mg/kg,用时用生理盐水配成溶液)0.1ml,同时按120mg/kg颈背部皮下注射D-半乳糖(用时用生理盐水配成5%的溶液)0.1ml。分别在动物用药期满2、4、6、8周时处死动物采样,每次采集大脑、肝脏组织,检测大脑、肝脏细胞DNA的损伤。结果:GPS三个剂量组小鼠大脑和肝脏细胞的拖尾率与模型组相比,均有所下降,其中大脑细胞的拖尾率在30%～45%之间,且差异显著(P<0.01或P<0.05);DNA损伤程度也有所减轻,除了低剂量组外,高、中剂量组主要以Ⅱ级损伤为主。结论:GPS可以减轻模型组小鼠大脑和肝脏组织细胞DNA的损伤程度。     

Evaluation of rhinoscopy and rhinoscopy-assisted mucosal biopsy in diagnosis of nasal disease in dogs: 119 cases (1985-1989).

The case records of 149 dogs examined from 1985 to 1989 with clinical signs of nasal disease were reviewed. Gross rhinoscopy was performed in 119 dogs, and rhinoscopy-assisted pinch biopsy was performed in 109. Rhinoscopy was performed by use of a 2.7-mm rigid fiberoptic endoscope. Mucosal biopsy specimens were obtained with rhinoscopic guidance by use of a 2 x 3-mm biopsy forcep. Gross, cytologic, and histologic findings are summarized. Ninety-four of 119 cases could be evaluated on the basis of diagnostic and follow-up criteria established by the authors. The diagnostic success rate of gross rhinoscopy with rhinoscopy-assisted biopsy was 83% (78 of 94 evaluated cases). Protracted hemorrhage was a complication in 2 of 109 cases in which rhinoscopy-assisted biopsy was performed. It was concluded that rhinoscopy with rhinoscopy-assisted biopsy contributes important diagnostic information in dogs with nasal disease without the relative invasiveness, expense, and risk of surgery.