Rearrangement patterns of the canine TCRγ locus in a distinct group of T cell lymphomas

The T cell antigen receptor chains are assembled through a rearrangement process that combines variable (V), diversity (D) and joining (J) region genes. Recently, the entire canine T cell receptor γ (TRG) locus was described. It is arranged in 8 cassettes with up to 3 V genes, 2 J genes and 1 C gene each. However, no data is available beyond the level of sequence analysis. The objective of this study was to identify rearranged genes of the canine TRG locus through experimental analysis and to assess gene usage and patterns of rearrangement in a series of canine T cell lymphomas. Rearranged genes were identified through computational analysis of recombination signal sequences (RSSs), a gene's potential to generate a polyclonal smear, and through sequencing of clonal rearrangements in a series of T cell lymphomas. Out of a total of 32 Vγ and Jγ genes, 21 genes were found to rearrange, 8 genes were considered not rearranged and 3 genes were suspected to rearrange but their status could not be determined definitely. Rearrangements of the canine TRG locus were assessed in a group of canine T cell lymphomas as well as 3 neoplastic T cell lines. An average of 4.6 rearrangements per lymphoma was found suggesting that canine T cells routinely rearrange multiple cassettes per allele. The most commonly rearranged Vγ genes belonged to subgroups Vγ2, Vγ3, and Vγ7. Genes in cassettes 2 and 3 preferentially rearranged within their respective cassettes, while Vγ genes in cassette 7 rearranged to a Jγ gene in cassette 8. There was a strong preference for Vγ2 genes to rearrange to a 3' Jγ gene and for Vγ3 and Vγ7 genes to rearrange to a 5' Jγ gene. This rearrangement pattern coincided with the conservation of the spacer sequence between V and J gene subgroups rather than the topologic location of genes. These data show that highly divergent spacer sequences allow for equally efficient recombination and suggest that spacer sequences can mediate compatibility between V and J genes.     

Characterization of the diffuse mucosal associated lymphoid tissue of feline small intestine

Characterization of the feline intestinal mucosal associated lymphoid tissue (MALT) will facilitate investigation of intestinal disease in the cat and promote the cat as an animal model for a range of human diseases which involve the intestinal lymphoid tissue. This includes inflammatory bowel disease, viral and non-viral associated intestinal lymphomas and immunodeficiency associated syndromes. Morphologic and phenotypic characterization of the normal small intestinal diffuse MALT in 22 SPF cats was performed using flow cytometry and cytology on isolated intestinal leukocytes from the intra-epithelial and lamina proprial compartments, as well as immunohistology on tissues from the feline duodenum, jejunum and ileum. The intra-epithelial compartment (IEC) was dominated by lymphocytes (>85%) which frequently contained intracytoplasmic granules. The most striking findings in the IEC were the elevated percentages of CD8 alpha+ lymphocytes (40%), presumed to express CD8 alpha alpha chains, and CD4-/CD8- (double negative) lymphocytes (44%), and the consistent presence of a minor subpopulation of CD3+/CD11d+ IELs (6%). Small percentages of CD4+ lymphocytes (10%) were observed such that the IEL CD4:CD8 ratio (0.25) was low. The LPC also contained a majority of T cells and few plasma cells. However, this compartment had reduced percentages of CD8 alpha+ lymphocytes (28%) and increased percentages of CD4+ lymphocytes (27%) relative to the IEC. However, the LPL CD4:CD8 ratio (1.0) remained low compared with the ratio in peripheral blood. In feline MALT, MHC class II expression was lower than in other peripheral lymphoid compartments. The results of this study provide important reference values for future investigations involving feline intestinal lymphocytes and demonstrates that the leukocyte distribution and phenotypic characteristics of the feline diffuse MALT appear largely similar to the murine, rat and human counterparts.     

Canine anal sac adenocarcinomas: clinical presentation and response to therapy

A retrospective study of 43 dogs with anal sac adenocarcinoma (ASAC) was performed to characterize the clinical presentation and response to treatment. Clinical signs at presentation varied considerably, with signs related either to sublumbar nodal metastasis (tenesmus or constipation) or hypercalcemia (polyuria-polydipsia and anorexia) being the most frequent findings. At the time of presentation, 23 (53%) dogs had hypercalcemia and 34 (79%) had metastases, with the regional lymph nodes (31 dogs, 72%) being the most common site of metastasis. A variety of chemotherapeutic agents were administered, with partial remission (PR) recorded in 4 of 13 (31%) dogs treated with cisplatin and in 1 of 3 (33%) dogs treated with carboplatin. The median survival for all dogs was 6 months (range, 2 days-41 months). There was no statistical association between the presence of hypercalcemia and survival, although the power of the study to detect an increase in survival of 3 months was low (.33). We conclude that platinum chemotherapy has antitumor activity in canine apocrine gland carcinoma and that further study of these agents is warranted.     

Audiograms estimated from brainstem tone-evoked potentials in dogs from 10 days to 1.5 months of age

The objective of this study was to build audiograms from thresholds of brainstem tone-evoked potentials in dogs and to evaluate age-related change of the audiogram in puppies. Results were obtained from 9 Beagle puppies 10-47 days of age. Vertex to mastoid brainstem auditory-evoked potentials in response to 5.1-millisecond Hanning-gated sine waves with frequencies octave-spaced from 0.5 to 32 kHz were recorded. Three dogs were examined at 10, 13, 19, 25, and 45 days. Four other dogs were examined at 16 days. Data from 7 dogs between 42 and 47 days of age were pooled to obtain audiogram reference values in 1.5-month-old puppies. The best auditory threshold lowered from above 60 dB sound pressure level (SPL) to values close to 0 dB SPL between 13 and 25 days of age and then stabilized. The audible frequency range widened, including 32 kHz in all tested dogs from the 19th day. In the 7 1.5-month-old puppies, the mean auditory threshold decreased by 11 dB per octave from 0.5 to 2 kHz. The auditory threshold was lowest and held the same value from 2 to 8 kHz. The mean auditory threshold increased by 20 dB per octave from 8 to 32 kHz. Near threshold, click-evoked potentials test only a small part of the audible frequency range in dogs. Use of tone-evoked potentials may become a powerful tool in investigating dogs with possible partial hearing loss, including during the auditory system maturation period.     

Indium-111 labeled vitamin B12 imaging of a ciliary adenoma with concurrent grade 2 soft tissue sarcoma of the leg in a Labrador Retriever

An 11-year-old, male castrated, Labrador Retriever was evaluated for the presence of rapidly growing concurrent leg and intraocular masses. Metastasis was noted in the chest at time of initial presentation. Indium-111 labeled vitamin B12 imaging was performed, and there was significant uptake by both primary tumors and the lung metastases. Enucleation and amputation were performed for palliative relief. The leg mass was a grade 2 soft tissue sarcoma and the ocular mass a ciliary adenoma. The dog remained symptom-free for approximately 10 weeks before developing signs of respiratory distress. He was euthanized 12 weeks after initial presentation, and there was diffuse infiltration of the lungs with metastatic sarcoma. Indium-111 labeled vitamin B12 imaging identified a ciliary adenoma in this case and may provide a useful differentiation technique for evaluation of intraocular and retrobulbar masses if it can be demonstrated that there is differential uptake between inflammatory or infectious conditions and neoplasia.     

Effects of additional starch or fat in late-gestating high nonstarch polysaccharide diets on litter performance and glucose tolerance in sows

The effects of feeding additional starch or fat from d 85 of gestation until parturition on litter performance and on glucose tolerance in sows that were fed a diet with a high level of fermentable nonstarch polysaccharides (NSP) were studied. The day after breeding, 141 multiparous sows were assigned to the experiment. At d 85 of gestation, sows were assigned to the treatments. Sows were fed 3.4 kg/d (as-fed basis) of a high-NSP diet or the same quantity of the high-NSP diet and an additional 360 g of starch (from wheat starch) daily, or the same quantity of the high-NSP diet and an additional 164 g of fat (from soybean oil) daily. During lactation, all sows were given free access to the same lactation diet. Approximately 1 wk before the expected time of parturition, an oral glucose tolerance test was performed in 38 randomly chosen sows by feeding pelleted glucose (3 g/kg BW0.75). Blood samples for glucose analyses were taken at -10, 10, 20, 30, 40, 50, 60, 70, 80, 90, 105, and 120 min after glucose was fed. The supply of additional dietary starch or fat did not increase piglet birth weight or total litter weight at birth. Sows that were fed the high-NSP diet had more (P = 0.097) live-born piglets and fewer (P = 0.084) stillborn piglets than did sows that were fed additional fat, whereas sows that were fed additional starch were intermediate for these variables. Piglet mortality after birth was not affected by dietary treatment. Body weight and backfat gains in the last month of gestation were higher for sows fed additional starch or fat than for sows fed the high-NSP diet (P < 0.001 and P = 0.017, respectively). Feed intake in lactation was greatest by sows fed the high-NSP diet, least by sows fed additional starch at the end of gestation, and intermediate by sows fed additional fat (P = 0.099). The differences in lactation feed intake did not result in differences in BW and backfat losses during lactation. Sows that were fed additional fat had the greatest glucose area under the curve (P = 0.044), indicating that these sows were less tolerant to glucose. In conclusion, feeding additional energy (starch or fat) in late-gestating sows that are fed a high-NSP diet did not increase litter weight at birth or piglet survival, but did increase maternal gain. Feeding sows additional energy from fat might induce glucose intolerance, whereas feeding sows additional energy from starch did not induce glucose intolerance.     

Characterization of a feline homologue of the alphaE integrin subunit (CD103) reveals high specificity for intra-epithelial lymphocytes

The characteristics of a feline homologue of the alphaE integrin (CD103), defined by two murine monoclonal antibodies, Fe7.1B8 (IgG1) and Fe7.2D8 (IgG1), are described. These antibodies recognized 75% of intra-epithelial (range 59-88%) and 40% of lamina proprial (range 28-46%) T cells of the intestinal mucosal tissue of the small intestine in contrast with approximately 2% of peripheral blood lymphocytes. Both antibodies immunoprecipitated a 180 kDa protein from biotinylated feline intra-epithelial mucosal leukocytes consistent with the alphaE integrin subunit in conjunction with a 120 kDa protein consistent with the beta7 subunit. The nucleotide sequence of feline alphaE integrin, generated from molecular cloning of the feline alphaE encoding cDNA, is also reported. This feline molecule shares 72% sequence homology with human and 69% homology with murine and rat counterparts. Homology includes the presence of an X (extra) domain, that appears unique to alphaE molecules as described for human, rat and mouse, as well as areas of homology common to other alpha integrins. Of note is a typical I (inserted) domain, the presence of seven repeat regions, and highly conserved sequences in the cytoplasmic tail. Transfection studies demonstrated that both antibodies recognized an extracellular component which encompassed the X and I domains of the cloned alphaE integrin subunit. These studies demonstrate that the pattern of tissue distribution, biochemical characteristics, and cDNA sequence of the feline alphaE integrin subunit are largely similar to that described for other species.     

Photosynthetic capacity of red spruce during winter

We measured the photosynthetic capacity (P(max)) of plantation-grown red spruce (Picea rubens Sarg.) during two winter seasons (1993-94 and 1994-95) and monitored field photosynthesis of these trees during one winter (1993-94). We also measured P(max) for mature montane trees from January through May 1995. Changes in P(max) and field photosynthesis closely paralleled seasonal changes in outdoor air temperature. However, during thaw periods, field photosynthesis was closely correlated with multiple-day temperature regimes, whereas P(max) was closely correlated with single-day fluctuations in temperature. There was a strong association between short-term changes in ambient temperature and P(max) during the extended thaw of January 1995. Significant increases in P(max) occurred within two days of the start of this thaw. Repeated measurements of cut shoots kept indoors indicated that temperature-induced increases in P(max) can occur within 3 h. Although significant correlations between P(max) and stomatal conductance (g(s)) or intracellular CO(2) concentration (C(i)) raised the possibility that increases in P(max) resulted from increases in stomatal aperture, fluctuations in g(s) or C(i) explained little of the overall variation in P(max). Following both natural and simulated thaws, P(max) increased considerably but plateaued at only 37% of the mean photosynthetic rate reported for red spruce during the growing season. Thus, even though shoots were provided with near-optimal environmental conditions, and despite thaw-induced changes in physiology, significant limitations to winter photosynthesis remained.