Publications by authors named "Franco Guscetti"

33 Publications

Histological and immunohistochemical investigation of canine prostate carcinoma with identification of common intraductal carcinoma component.

Vet Comp Oncol 2021 May 7. Epub 2021 May 7.

BioDiscovery Institute, University of Nottingham, Nottingham, UK.

A limited number of species, including men and dogs, spontaneously develop prostate cancer (PC). The histological and molecular relevance of canine PC as a model for the disease in men remains controversial. To address this challenge, this study aimed to assess the histomorphology and expression of basal cell, urothelial and neuroendocrine markers [p63, high molecular weight cytokeratin (HMWCK), Uroplakin 3 (UPIII), neuron-specific enolase (NSE)] in canine PC (n = 41). Based on histomorphology, 10/41 (24%), 21/41 (51%) and 9/41 (22%) were classified as adenocarcinoma (AC), urothelial carcinoma (UC), and mixed carcinoma, respectively. Tumour inflammation was common, frequently severe [20/41 (49%)], and associated with neutering (p < .02) and urothelial differentiation (p < .02). Most (36/40, 90%) cancers contained only rare cells with basal cell marker expression or were negative. The expression of UPIII was absent or weak in the majority (33/38, 87%) of tumours, with moderate to strong staining in the remaining cases. NSE expression in PC was rare and limited to 2/14 (14%) cases. Tumour extension into benign ducts and glands was a common finding with presence in 17/39 (44%) of carcinomas with and without urothelial differentiation. In conclusion, we confirm that canine PC is characterized by absent or weak expression of basal cell and urothelial markers. Although rare, NSE expression, potentially indicating neuroendocrine differentiation, is reported for the first time in canine PCa. Intraductal carcinoma of the prostate with concurrent invasive PCa (IDCP-inv) is a frequent, not previously described, finding in dogs with PC.
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http://dx.doi.org/10.1111/vco.12704DOI Listing
May 2021

Identification of disease-promoting stromal components by comparative proteomic and transcriptomic profiling of canine mammary tumors using laser-capture microdissected FFPE tissue.

Neoplasia 2021 04 29;23(4):400-412. Epub 2021 Mar 29.

Institute of Veterinary Pharmacology and Toxicology, Vetsuisse Faculty, University of Zurich, Zurich, Switzerland. Electronic address:

Cancer-associated stroma (CAS) profoundly influences progression of tumors including mammary carcinoma (mCA). Canine simple mCA represent relevant models of human mCA, notably also with respect to CAS. While transcriptomic changes in CAS of mCA are well described, it remains unclear to what extent these translate to the protein level. Therefore, we sought to gain insight into the proteomic changes in CAS and compare them with transcriptomic changes in the same tissue. To this end, we analyzed CAS and matched normal stroma using laser-capture microdissection (LCM) and LC-MS/MS in a cohort of 14 formalin-fixed paraffin embedded (FFPE) canine mCAs that we had previously characterized using LCM-RNAseq. Our results reveal clear differences in protein abundance between CAS and normal stroma, which are characterized by changes in the extracellular matrix, the cytoskeleton, and cytokines such as TNF. The proteomics- and RNAseq-based analyses of LCM-FFPE show a substantial degree of correlation, especially for the most deregulated targets and a comparable activation of pathways. Finally, we validate transcriptomic upregulation of LTBP2, IGFBP2, COL6A5, POSTN, FN1, COL4A1, COL12A1, PLOD2, COL4A2, and IGFBP7 in CAS on the protein level and demonstrate their adverse prognostic value for human breast cancer. Given the relevance of canine mCA as a model for the human disease, our analysis substantiates these targets as disease-promoting stromal components with implications for breast cancer in both species.
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http://dx.doi.org/10.1016/j.neo.2021.03.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8042244PMC
April 2021

Molecular homology between canine spontaneous oral squamous cell carcinomas and human head-and-neck squamous cell carcinomas reveals disease drivers and therapeutic vulnerabilities.

Neoplasia 2020 12 2;22(12):778-788. Epub 2020 Nov 2.

Institute of Veterinary Pharmacology and Toxicology, Vetsuisse Faculty, University of Zürich, Zürich, Switzerland. Electronic address:

Spontaneously occurring canine oral squamous cell carcinomas (COSCC) are viewed as a useful model for human head and neck squamous cell carcinomas (HNSCC). To date however, the molecular basis of COSCC remains poorly understood. To identify changes pertinent to cancer cells in COSCC, we specifically analyzed tumor cells and matched normal epithelium from clinical formalin-fixed paraffin-embedded specimens using laser-capture-microdissection coupled with RNA-sequencing (RNAseq). Our results identify strong contributions of epithelial-to-mesenchymal transition (EMT), classical tumor-promoting (such as E2F, KRAS, MYC, mTORC1, and TGFB1 signaling) and immune-related pathways in the tumor epithelium of COSCC. Comparative analyses of COSCC with 43 paired tumor/normal HNSCC from The Cancer Genome Atlas revealed a high homology in transcriptional reprogramming, and identified processes associated with cell cycle progression, immune processes, and loss of cellular differentiation as likely central drivers of the disease. Similar to HNSCC, our analyses suggested a ZEB2-driven partial EMT in COSCC and identified selective upregulation of KRT14 and KRT17 in COSCC. Beyond homology in transcriptional signatures, we also found therapeutic vulnerabilities strongly conserved between the species: these included increased expression of PD-L1 and CTLA-4, coinciding with EMT and revealing the potential for immune checkpoint therapies, and overexpression of CDK4/6 that sensitized COSCC to treatment with palbociclib. In summary, our data significantly extend the current knowledge of molecular aberrations in COSCC and underline the potential of spontaneous COSCC as a model for HNSCC to interrogate therapeutic vulnerabilities and support translation of novel therapies from bench to bedside.
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http://dx.doi.org/10.1016/j.neo.2020.10.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7642746PMC
December 2020

Chlamydia suis is associated with intestinal NF-κB activation in experimentally infected gnotobiotic piglets.

Pathog Dis 2020 08;78(6)

Department of Pathobiology, Institute of Veterinary Pathology, Vetsuisse Faculty, University of Zurich, Winterthurerstrasse 268, CH-8057 Zurich, Switzerland.

Chlamydia suis intestinal infection of single-animal experimental groups of gnotobiotic newborn piglets was previously reported to cause severe, temporary small intestinal epithelium damage. We investigated archived intestinal samples for pro-inflammatory nuclear factor kappa B (NF-κB) activation, Interleukin (IL)-6 and IL-8 production and immune cell influx. Samples were collected 2, 4 and 7 days post-inoculation with C. suis strain S45/6 or mock inoculum (control). Increased nuclear localization of epithelial NF-κB, representative of activation, in the jejunum and ileum of C. suis-infected animals, compared to uninfected controls, began by 2 days post-infection (dpi) and persisted through 7 dpi. Infected animals showed increased production of IL-8, peaking at 2 dpi, compared to controls. Infection-mediated CD45-positive immune cell influx into the jejunal lamina propria peaked at 7 dpi, when epithelial damage was largely resolved. Activation of NF-κB appears to be a key early event in the innate response of the unprimed porcine immune system challenged with C. suis. This results in an acute phase, coinciding with the most severe clinical symptoms, diarrhea and weight loss. Immune cells recruited shortly after infection remain present in the lamina propria during the recovery phase, which is characterized by reduced chlamydial shedding and restored intestinal epithelium integrity.
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http://dx.doi.org/10.1093/femspd/ftaa040DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8140907PMC
August 2020

Cross species application of quantitative neuropathology assays developed for clinical Alzheimer's disease samples.

Pathobiol Aging Age Relat Dis 2019 3;9(1):1657768. Epub 2019 Sep 3.

Department of Pathology, University of Washington, Seattle, WA, USA.

A major obstacle for preclinical testing of Alzheimer's disease (AD) therapies is the availability of translationally relevant AD models. Critical for the validation of such models is the application of the same approaches and techniques used for the neuropathological characterization of AD. Deposition of amyloid-β 42 (Aβ42) plaques and neurofibrillary tangles containing phospho-Tau (pTau) are the pathognomonic features of AD. In the neuropathologic evaluation of AD, immunohistochemistry (IHC) is the current standard method for detection of Aβ42 and pTau. Although IHC is indispensable for determining the distribution of AD pathology, it is of rather limited use for assessment of the quantity of AD pathology. We have recently developed Luminex-based assays for the quantitative assessment of Aβ42 and pTau in AD brains. These assays are based on the same antibodies that are used for the IHC-based diagnosis of AD neuropathologic change. Here we report the application and extension of such quantitative AD neuropathology assays to commonly used genetically engineered AD models and to animals that develop AD neuropathologic change as they age naturally. We believe that identifying AD models that have Aβ42 or pTau levels comparable to those observed in AD will greatly improve the ability to develop AD therapies. : Alzheimer's disease (AD); amyloid β 42 (Aβ42); phospho-Tau (pTau); immunohistochemistry (IHC).
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http://dx.doi.org/10.1080/20010001.2019.1657768DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6735310PMC
September 2019

A review of canine B cell clonality assays and primer set optimization using large-scale repertoire data.

Vet Immunol Immunopathol 2019 Mar 25;209:45-52. Epub 2019 Jan 25.

Department of Pathobiology, Ontario Veterinary College, University of Guelph, Guelph, Ontario, Canada. Electronic address:

Several molecular clonality assays have been developed to assess canine B cell proliferations. These assays were based on different sequence data, utilized different assay designs and employed different testing strategies. This has resulted in a complex body of literature and complicates evidence-based selection of primer sets. In addition, further refinement of primer sets is difficult because it is unknown how well current primer sets cover the expressed sequence repertoire. The objectives of this study were 1) to provide an overview of published IGH clonality assays that highlights key differences in assay design and testing strategy and 2) to propose a novel method for optimizing primer sets that leverages large-scale sequencing data. A review of previously published assays highlighted confounding factors that hamper a direct comparison of performance metrics between studies. These findings illustrate the need for a multi-institutional effort to harmonize veterinary clonality testing. A novel in silico analysis of primer sequences using a large dataset of expressed sequences identified shortfalls of existing primer sets and was used to guide primer optimization. Three optimized primer sets were tested and yielded qualitative sensitivity values between 80-90%. The qualitative sensitivity ranged from 1% to over 50% and was dependent on the size of the neoplastic clone and the sample DNA used. These findings illustrate that inclusion of high-throughput sequencing data for primer design can be a useful tool to guide primer design and optimization. This strategy could be applied to other antigen receptor loci or species to further improve veterinary clonality assays.
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http://dx.doi.org/10.1016/j.vetimm.2019.01.002DOI Listing
March 2019

Graft-versus-host disease, but not graft-versus-leukemia immunity, is mediated by GM-CSF-licensed myeloid cells.

Sci Transl Med 2018 11;10(469)

Institute of Experimental Immunology,University of Zurich, CH-8057 Zurich, Switzerland.

Allogeneic hematopoietic cell transplantation (allo-HCT) not only is an effective treatment for several hematologic malignancies but can also result in potentially life-threatening graft-versus-host disease (GvHD). GvHD is caused by T cells within the allograft attacking nonmalignant host tissues; however, these same T cells mediate the therapeutic graft-versus-leukemia (GvL) response. Thus, there is an urgent need to understand how to mechanistically uncouple GvL from GvHD. Using preclinical models of full and partial MHC-mismatched HCT, we here show that the granulocyte-macrophage colony-stimulating factor (GM-CSF) produced by allogeneic T cells distinguishes between the two processes. GM-CSF drives GvHD pathology by licensing donor-derived phagocytes to produce inflammatory mediators such as interleukin-1β and reactive oxygen species. In contrast, GM-CSF did not affect allogeneic T cells or their capacity to eliminate leukemic cells, retaining undiminished GvL responses. Last, tissue biopsies and peripheral blood mononuclear cells from patients with grade IV GvHD showed an elevation of GM-CSF-producing T cells, suggesting that GM-CSF neutralization has translational potential in allo-HCT.
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http://dx.doi.org/10.1126/scitranslmed.aat8410DOI Listing
November 2018

Bovine Fetal Placenta During Pregnancy and the Postpartum Period.

Vet Pathol 2019 Mar 24;56(2):248-258. Epub 2018 Oct 24.

1 Institute of Veterinary Pathology, University of Zurich, Vetsuisse Faculty, Zurich, Switzerland.

Bovine abortion is a worldwide problem, but despite extensive histopathologic and molecular investigations, the cause of abortion remains unclear in about 70% of cases. Cellular debris is a commonly observed histopathologic finding in the fetal placenta and is often interpreted as necrosis. In this study, the nature of this cellular debris was characterized, and histologic changes in the normal fetal placenta during pregnancy and after delivery were assessed. In addition, the presence of the most common abortifacient pathogens in Switzerland ( Chlamydiaceae, Coxiella burnetii, Neospora caninum) was tested by polymerase chain reaction. We collected 51 placentomes and 235 cotyledons from 41 and from 50 cows, respectively. In total, cellular debris was present in 48 of 51 (94%) placentomes and in 225 of 235 (96%) cotyledons, inflammation occurred in 1 of 51 (2%) placentomes and in 46 of 235 (20%) cotyledons, vasculitis was seen in 1 of 51 (2%) placentomes and 46 of 235 (20%) cotyledons, and 18 of 51 (35%) placentomes and 181 of 235 (77%) cotyledons had mineralization. The amount of cellular debris correlated with areas of positive signals for cleaved caspase 3 and lamin A. Therefore, this finding was interpreted as an apoptotic process. In total, 10 of 50 cotyledons (20%) were positive for C. burnetii DNA, most likely representing subclinical infections. The results of our study indicate that histologic features in the fetal placenta such as cellular debris, inflammation, vasculitis, and mineralization must be considered physiological processes during pregnancy and after delivery. Therefore, their presence in placentae of aborted fetuses must be interpreted with caution and might not be necessarily linked to an infectious cause of abortion.
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http://dx.doi.org/10.1177/0300985818806453DOI Listing
March 2019

Cutaneous Tumors in Swiss Dogs: Retrospective Data From the Swiss Canine Cancer Registry, 2008-2013.

Vet Pathol 2018 11 21;55(6):809-820. Epub 2018 Aug 21.

4 Institute of Animal Pathology, Department of Infectious Diseases and Pathobiology, Vetsuisse Faculty, University of Bern, Bern, Switzerland.

Data collected in animal cancer registries comprise extensive and valuable information, even more so when evaluated in context with precise population data. The authors evaluated 11 740 canine skin tumors collected in the Swiss Canine Cancer Registry from 2008-2013, considering data on breed, sex, age, and anatomic locations. Their incidence rate (IR) per 100 000 dogs/year in the Swiss dog population was calculated based on data from the official and mandatory Swiss dog registration database ANIS. The most common tumor types were mast cell tumors (16.35%; IR, 60.3), lipomas (12.47%; IR, 46.0), hair follicle tumors (12.34%; IR, 45.5), histiocytomas (12.10%; IR, 44.6), soft tissue sarcomas (10.86%; IR, 40.1), and melanocytic tumors (8.63%; IR, 31.8) with >1000 tumors per type. The average IR of all tumor types across the 227 registered breeds was 372.2. The highest tumor incidence was found in the Giant Schnauzer (IR, 1616.3), the Standard Schnauzer (IR, 1545.4), the Magyar Vizsla (IR, 1534.6), the Rhodesian Ridgeback (IR, 1445.0), the Nova Scotia Duck Tolling Retriever (IR, 1351.7), and the Boxer (IR, 1350.0). Mixed-breed dogs (IR, 979.4) had an increased IR compared to the average of all breeds. Previously reported breed predispositions for most tumor types were confirmed. Nevertheless, the data also showed an increased IR for mast cell tumors and melanocytic tumors in the Nova Scotia Duck Tolling Retriever and for histiocytomas in the Flat Coated Retriever. The results from this study can be taken into consideration when selecting purebred dogs for breeding to improve a breed's health.
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http://dx.doi.org/10.1177/0300985818789466DOI Listing
November 2018

Dynamic In Vivo Profiling of DNA Damage and Repair after Radiotherapy Using Canine Patients as a Model.

Int J Mol Sci 2017 Jun 1;18(6). Epub 2017 Jun 1.

Division of Radiation Oncology, Vetsuisse Faculty University of Zurich, CH-8057 Zurich, Switzerland.

Time resolved data of DNA damage and repair after radiotherapy elucidates the relation between damage, repair, and cell survival. While well characterized in vitro, little is known about the time-course of DNA damage response in tumors sampled from individual patients. Kinetics of DNA damage after radiotherapy was assessed in eight dogs using repeated in vivo samples of tumor and co-irradiated normal tissue analyzed with comet assay and phosphorylated H2AX (γH2AX) immunohistochemistry. In vivo results were then compared (in silico) with a dynamic mathematical model for DNA damage formation and repair. Maximum %DNA in tail was observed at 15-60 min after irradiation, with a rapid decrease. Time-courses of γH2AX-foci paralleled these findings with a small time delay and were not influenced by covariates. The evolutionary parameter search based on %DNA in tail revealed a good fit of the DNA repair model to in vivo data for pooled sarcoma time-courses, but fits for individual sarcoma time-courses suffer from the heterogeneous nature of the in vivo data. It was possible to follow dynamics of comet tail intensity and γH2AX-foci during a course of radiation using a minimally invasive approach. DNA repair can be quantitatively investigated as time-courses of individual patients by integrating this resulting data into a dynamic mathematical model.
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http://dx.doi.org/10.3390/ijms18061176DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5485999PMC
June 2017

Retrospective clinical study on outcome in cats with nasal planum squamous cell carcinoma treated with an accelerated radiation protocol.

BMC Vet Res 2017 Apr 4;13(1):86. Epub 2017 Apr 4.

Division of Radiation Oncology, Vetsuisse-Faculty, University of Zurich, CH-8057, Zurich, Switzerland.

Background: Cutaneous squamous cell carcinoma of the nasal planum in cats is a common indication for antitumor treatment such as external beam radiation therapy. Curative-intent radiation therapy has been described as a valuable treatment option, resulting in long and stable tumor control in these patients. The aim of the current study was to evaluate outcome and toxicity, as well as possible prognostic factors using an accelerated hypofractionated radiation therapy protocol. Cats with squamous cell carcinoma of the nasal planum treated with an accelerated radiation protocol (10 × 4.8 Gy, over one week) were retrospectively evaluated. Tumor- and treatment-associated variables were evaluated in respect to local control and survival.

Results: Forty-four cats met the inclusion criteria for this study. All cats showed complete response to therapy. Median disease-free interval (DFI) for all cases was 916 days (95% CI: 456-1377). One- and two-year DFIs were 71% (95% CI: 56-86%) and 60% (95% CI: 43-77%). Of the tested variables, only tumor volume showed a tendency to influence DFI, with larger tumors having a 5.4 times greater risk of recurrence than the smaller ones (HR 1.33 (95% CI: 0.99-1.79), p = 0.054). Median overall survival (OS) was 902 days (95% CI: 862-942). One- and 2-year OSs were 79.3% (95% CI: 67.3-91.3) and 58.4% (95% CI: 42.8-74). Of the tested variables, again, only tumor volume influenced OS with larger tumors having a 6.3 times greater risk of dying than the smaller ones (HR 1.36 (95% CI: 1.07-1.73), p = 0.010). The acute and late toxicity profile was low and hence clinically acceptable.

Conclusions: Curative-intent radiation therapy with an accelerated fractionation schedule can be considered a safe, cosmetically superior treatment option for cutaneous squamous cell carcinomas of the nasal planum in cats, resulting in long and stable tumor control.
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http://dx.doi.org/10.1186/s12917-017-1018-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5381142PMC
April 2017

Hypoxia-Related Marker GLUT-1, CAIX, Proliferative Index and Microvessel Density in Canine Oral Malignant Neoplasia.

PLoS One 2016 23;11(2):e0149993. Epub 2016 Feb 23.

Division of Radiation Oncology, Vetsuisse Faculty University of Zurich, Zurich, Switzerland.

For various types of tumor therapy, it is suggested that co-targeting of tumor microenvironment, mainly tumor vasculature, mediates tumor response mechanisms. Immunohistochemistry for glucose transporter-1 (GLUT-1), carbonic anhydrase-IX (CAIX), Ki-67, and von Willebrand factor VIII for microvessel density (MVD) were performed on formalin-fixed paraffin-embedded samples of canine oral malignant neoplasms. Polarographic oxygen measurements (median pO2) and perfusion data via contrast-enhanced power Doppler ultrasound (median vascularity, median blood volume) provided additional information. Ninety-two samples were analyzed: sarcomas (n = 32), carcinomas (n = 30), and malignant melanomas (n = 30). Polarographic oxygen and perfusion data was available in 22.8% (sarcomas n = 9, carcinomas n = 7, melanomas n = 5), and 27.1% (sarcomas n = 10, carcinomas n = 8, melanomas n = 7) of cases, respectively. GLUT-1 expression was detected in 46.7% of all samples, and was generally weak. CAIX expression was found in 34.8% of all samples. Median Ki-67 score and MVD count was 19% and 17, respectively. The evaluation of the GLUT-1 score and continuous data showed significantly lower GLUT-1 levels in sarcomas (mean 5.1%, SD 6.2) versus carcinomas and melanomas (mean 16.5%/ 19.0%, SD 17.3/ 20.9, p = 0.001). The expression of CAIX correlated mildly positively with GLUT-1 (p = 0.018, rho = 0.250) as well as with Ki-67 (p = 0.014, rho = 0.295). MVD showed a significantly lower level in melanomas (mean 12.6, SD 7.7) versus sarcomas and carcinomas (mean 21.8/ 26.9, SD 13.0/20.4, p = 0.001). Median vascularity and blood volume were significantly lower in sarcomas (mean 10.4%, SD 11.0, and mean 6.3%, SD 6.5, respectively) versus carcinomas (mean 39.2%, SD 16.4 and mean 33.0%, SD 25.6, respectively) and melanomas (mean 36.0%, SD 18.3, and 31.5%, SD 24.5). Between the 3 histological groups, there was neither a significant difference in the GLUT-1 and CAIX score and continuous data, nor the Ki67 score, or polarographic oxygen measurements. GLUT-1 continuous data and Ki-67 (p<0.001, rho = 0.403), as well as Ki-67 and MVD (p = 0.029, rho = 0.228) correlated positively and a mild correlation was found between vascularity and GLUT-1 (p = 0.043, rho = 0.408). GLUT-1, CAIX, proliferative index and MVD levels were established as microenvironmental descriptors with the purpose of creating a baseline in order to follow changes seen in the tumor microenvironment after hypofractionated radiation with high doses.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0149993PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4764341PMC
July 2016

Endocrine control of canine mammary neoplasms: serum reproductive hormone levels and tissue expression of steroid hormone, prolactin and growth hormone receptors.

BMC Vet Res 2015 Sep 15;11:235. Epub 2015 Sep 15.

Institute of Veterinary Anatomy, Vetsuisse Faculty University of Zurich, Winterthurerstrasse 260, Zurich, 8057, Switzerland.

Background: Neoplasms of the mammary gland are among the most common diseases in female domestic dogs (Canis familiaris). It is assumed that reproductive hormones influence tumorigenesis in this species, although the precise role of the endocrine milieu and reproductive state is subject to continuing discussion. In line with this, a recent systematic review of available data on the development of mammary neoplasms revealed weak evidence for risk reduction after neutering and an effect of age at neutering. Investigation of several hormone receptors has revealed decreased expression of estrogen receptor-alpha (ERα, ESR1), progesterone (P4) receptor (PGR), prolactin (PRL) receptor (PRLR) and growth hormone receptor (GHR) associated with neoplastic differentiation of mammary tissues. In other studies, increased levels of estrogens, progesterone and prolactin were found in serum and/or tissue homogenates of dogs with malignant neoplasms. However, the association between these entities within one animal population was never previously examined. Therefore, this study investigated the association between circulating serum concentrations of estradiol-17β, progesterone and prolactin, and gene expression of ERα (ESR1), ERβ (ESR2), PGR, PRLR, PRL and GHR, with respect to reproductive state (spayed vs. intact) and cycle stage (anestrus vs. diestrus). Additionally, the expression of E-cadherin (CDH-1) was evaluated as a possible indicator of metastatic potential.

Results: For all receptors, the lowest gene expression was found in malignant tumors compared to normal tissues of affected dogs. Steroid levels were not influenced by their corresponding receptor expression in mammary neoplasms, but increased PRL levels were negatively associated with low PRLR gene expression in malignant tumors. The expression of CDH-1 was influenced by tumor malignancy and cycle stage, i.e., the highest gene expression was found in benign mammary tumors in diestrous dogs compared to normal and malignant mammary tissues of anestrous and spayed dogs.

Conclusions: Herein, it has been confirmed that transformation towards malignant neoplasms is associated with significant reduction of gene expression of particular hormone receptors. Only PRLR in malignant tumors seems to be influenced by circulating PRL levels. In dogs, CDH-1 can be used as a prognostic factor; its expression, however, in benign tumors is influenced by cycle stage.
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http://dx.doi.org/10.1186/s12917-015-0546-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4570623PMC
September 2015

Penetration depth of corneal cross-linking with riboflavin and UV-A (CXL) in horses and rabbits.

Vet Ophthalmol 2016 Jul 27;19(4):275-84. Epub 2015 Jul 27.

Division of Ophthalmology, Equine Department, Vetsuisse Faculty, University of Zurich, Winterthurerstrasse, 260, 8057, Zurich, Switzerland.

Objective: CXL penetration depth is an important variable influencing clinical treatment effect and safety. The purposes of this study were to determine the penetration depth of CXL in rabbit and equine corneas in epithelium-on and epithelium-off procedures and to assess an ex vivo fluorescent biomarker staining assay for objective assessment of CXL penetration depth.

Procedures: CXL treatment was performed according to a standardized protocol on 21 and 17 rabbit eyes and on 12 and 10 equine eyes with and without debridement, respectively. Control corneas were treated similarly, but not exposed to CXL. Hemicorneas were stained with either phalloidin and DAPI to visualize intracellular F-actin and nuclei, or with hematoxylin and eosin. Loss of actin staining was measured and compared between groups.

Results: Epithelium-off CXL caused a median actin cytoskeleton loss with a demarcation at 274 μm in rabbits and 173 μm in horses. In non-CXL-treated controls, we observed a median actin cytoskeleton loss with a demarcation at 134 μm in rabbits and 149 μm in horses. No effect was detected in the epithelium-on procedure.

Conclusions: CXL penetration depth, as determined by a novel ex vivo fluorescent assay, shows clear differences between species. A distinct effect was observed following epithelium-off CXL treatment in the anterior stroma of rabbits, but no different effect was observed in horses in comparison with nontreated controls. Different protocols need to be established to effectively treat equine patients with infectious corneal disease.
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http://dx.doi.org/10.1111/vop.12301DOI Listing
July 2016

Classification of primary hepatic tumours in the cat.

Vet J 2014 Nov 15;202(2):255-66. Epub 2014 Jul 15.

Department of Clinical Sciences of Companion Animals, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands. Electronic address:

Hepatic tumours in dogs have recently been re-classified to follow a revised human classification system that takes account of identified hepatic progenitor cells. This study investigated the presence and relative frequency of morphological types of feline primary hepatic neoplasms and aimed to determine whether a similar new classification scheme could be applied in cats. Feline primary liver tumours (n = 61) were examined histologically and with a series of immunohistochemical markers. Six cases of nodular hyperplasia and 21 tumours of hepatocellular origin were diagnosed. The latter were subdivided into hepatocellular tumours that were well differentiated and had no evidence of metastases (n = 18) and tumours that showed poorly differentiated areas with marked cellular and nuclear pleomorphism and had intrahepatic and, or, distant metastases (n = 3). These malignant feline hepatocellular tumours maintained their hepatocellular characteristics (HepPar-1, MRP2, pCEA positive) and were negative, or only <5% positive, for K19. Twenty-five cholangiocellular tumours were diagnosed and all had intrahepatic and, or, distant metastases. Eight NSE positive small cell carcinomas (carcinoids) were diagnosed and subdivided into small cell carcinomas with HPC characteristics (K19 positive) and neuroendocrine carcinomas (K19 negative). In addition, one squamous cell carcinoma originating from the distal part of the choledochal duct was recognised. Feline primary hepatic neoplasms can be sub-divided into benign and malignant hepatocellular tumours, cholangiocellular carcinomas, small cell carcinomas with HPC characteristics, neuroendocrine carcinomas and squamous cell carcinomas. The marked species difference justifies a specific classification for feline primary hepatic neoplasms.
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http://dx.doi.org/10.1016/j.tvjl.2014.07.002DOI Listing
November 2014

First meeting of the European canine lymphoma group. Workshop: state of the art and comparative aspects in canine lymphoma. CH-Lugano, 22 June 2013.

Hematol Oncol 2014 Jun 5;32(2):68-71. Epub 2013 Nov 5.

Dipartimento di Scienze Veterinarie e Sanità Pubblica, Università degli Studi di Milano, Italy.

This satellite meeting to the 12th International Conference on Malignant Lymphoma was conceived to bring together European researchers focused on canine lymphoma to explore several facets of this promising model of human disease. A series of invited lectures showed striking similarities between the two diseases namely in topics related to pathogenesis, diagnosis and classification and therapy. In particular, the potential value of the model was shown at the level of the NF-kB/p65 pathway, the Bcl-2 family of proteins, Ki67 and the S-phase fraction, as well as the MMPs, VEGF and PDGF. The utility of the growing body of well-characterized canine cell lines was stressed. The value of cytology and flow cytometry as tools for diagnosis, disease progression monitoring and prognosis were emphasized, whereas the failure so far of the standard immunohistochemical panel to differentiate between germinal centre and non-germinal centre diffuse large B-cell lymphomas subtypes in dogs was discussed. Further contributions included the report of encouraging results from a chemo-immunotherapy trial administered to dogs with diffuse large B-cell lymphoma, an overview on the use of radiation therapy for canine lymphoma and the role of surgery in splenic lymphoma. Altogether, the success of this meeting, attended by more than 160 participants, documents the rising interest for the spontaneous canine lymphoma model.
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http://dx.doi.org/10.1002/hon.2115DOI Listing
June 2014

Classification of primary hepatic tumours in the dog.

Vet J 2013 Sep 5;197(3):596-606. Epub 2013 Sep 5.

Department of Clinical Sciences of Companion Animals, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands.

Many advances have been made in the characterisation of primary liver tumours in humans, in particular relating to the identification and role of hepatic progenitor cells, resulting in a new classification. The aim of the present study was to investigate the presence and relative frequency of morphological types of canine primary hepatic neoplasms and to determine whether a classification similar to the human scheme can be applied to these canine neoplasms. Canine primary liver tumours (n=106) were examined histologically and with the immunohistochemical markers keratin 19, HepPar-1, epithelial membrane antigen/mucin-1, CD10, neuron-specific enolase and chromogranin-A. Eleven nodular hyperplasias and 82 tumours of hepatocellular origin were diagnosed. The latter were subdivided in hepatocellular tumours with 0-5% positivity for K19 (n=62), which were well differentiated and had no evidence of metastasis, tumours with >5% positivity for K19 (n=17), which were poorly differentiated and had intrahepatic and/or distant metastasis, and a scirrhous subgroup (n=3) with an intermediate position with regard to K19 staining and malignancy. Ten cholangiocellular tumours (nine cholangiocellular carcinomas and one cholangiolocarcinoma) were diagnosed and all had intrahepatic and/or distant metastases. Three neuroendocrine carcinomas were also diagnosed. Histopathological and immunohistochemical examination of canine primary hepatic neoplasms can differentiate hepatocellular, cholangiocellular and neuroendocrine tumours, in accordance with the most recent human classification system.
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http://dx.doi.org/10.1016/j.tvjl.2013.05.027DOI Listing
September 2013

Immunohistochemical expression of Bax and Bak in canine non-neoplastic tissues.

Vet J 2013 Oct 26;198(1):131-40. Epub 2013 Aug 26.

Institute of Veterinary Pathology, Vetsuisse Faculty, University of Zurich, Winterthurerstrasse 268, 8057 Zurich, Switzerland.

Apoptosis is critical for embryonic development, maintenance of tissue homeostasis and protection against malignant transformation. The Bcl-2 family of proteins plays a key role in intrinsic apoptosis by controlling the integrity of the outer mitochondrial membrane, and the multidomain pro-apoptotic Bcl-2 family members Bax and Bak are essential components of this pathway. The aim of this study was to provide data on the expression of these proteins in normal canine tissues. Two antibodies against Bax recognising different conformations of the protein and one antibody against Bak were validated by immunohistochemistry and immunoblotting using canine recombinant proteins and keratinocytes treated with ultraviolet light. The antibodies were used immunohistochemically to label a wide panel of histologically normal tissues assembled on tissue microarrays. In addition, a subset of the tissues was evaluated by Western blot analysis. Immunohistochemical and Western blot analyses revealed that both Bax and Bak are widely expressed in non-neoplastic tissues from adult dogs. Immunohistochemistry showed almost exclusively cytoplasmic labelling and prominent labelling of epithelial cells. In lymph nodes, immunohistochemical labelling was diffuse for both proteins and showed enhanced intensities in the mantle zones for Bax and the germinal centres for Bak. Strong reactivity for the active conformation of Bax was detected only in enterocytes and Leydig cells and in scattered lymphocytes. These data indicate widespread expression of Bax and Bak in normal canine tissues. Knowledge of the expression of Bax and Bak in normal tissues is a prerequisite in assessing the role of these proteins in canine neoplastic disease.
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http://dx.doi.org/10.1016/j.tvjl.2013.07.029DOI Listing
October 2013

Thoracic computed tomography, angiographic computed tomography, and pathology findings in six cats experimentally infected with Aelurostrongylus abstrusus.

Vet Radiol Ultrasound 2013 Sep-Oct;54(5):459-69. Epub 2013 May 29.

Section of Diagnostic Imaging, Vetsuisse Faculty, University of Zurich, Winterthurerstrasse 260 8057, Zurich, Switzerland.

Aelurostrongylus abstrusus infection is common in endemic areas and may cause severe respiratory clinical signs. Computed tomography (CT) is an important tool to diagnose pulmonary disease, because it allows detection of small lesions and discrimination of superimposed structures. The purpose of this study was to characterize by CT and angiographic CT the pulmonary lesions in six cats before, and 48 and 81 days after inoculation with 100 or 800 A. abstrusus infective larvae. Histological examination of the accessory lung lobe was performed to determine the microscopic, pathomorphologic correlate of the CT findings. The predominant CT lesion consisted of multiple nodules of varying size distributed throughout the lungs, severity depending on infectious dose. The histological correlate of the nodular lesions was multifocal dense granulomatous to mixed inflammatory cell infiltrates, including eosinophils distributed in the parenchyma and obliterating the alveoli. Marked, multifocal, dose-dependent thickening of the bronchi and adjacent interstitial changes blurred the margins of the outer serosal surface of the bronchi and vessels. Histologically, this was due to peribronchial mixed cell inflammation. During the course of infection some of the nodular and peribronchial changes were replaced by areas of ground-glass opacity. In addition to providing detailed depiction of pulmonary lesions resulting from an infectious cause and clearly defining lesions with respect to time and severity of infection, CT allowed quantitative assessment of bronchial thickness and lymph node size during the course of disease. Findings indicated that CT characteristics of this disease are consistent with pathologic findings.
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http://dx.doi.org/10.1111/vru.12044DOI Listing
April 2014

Expression of prolactin receptors in normal canine mammary tissue, canine mammary adenomas and mammary adenocarcinomas.

BMC Vet Res 2012 May 30;8:72. Epub 2012 May 30.

Clinic for Reproductive Medicine, Unit of Small Animal Reproduction, University of Zurich, Zurich, Switzerland.

Background: Mammary tumors represent the most common neoplastic disease in female dogs. Recently, the promoting role of prolactin (PRL) in the development of human breast carcinoma has been shown. Possible proliferative, anti-apoptotic, migratory and angiogenic effects of PRL on human mammary cancer cells in vitro and in vivo were suggested. The effects of PRL are mediated by its receptor, and alterations in receptor expression are likely to play a role in tumor development. Currently, not much data is available about prolactin receptor (PRLR) expression in canine mammary tumors. To set the basis for investigations on the role of PRL in mammary tumorigenesis in this species, prolactin receptor expression was evaluated by semi-quantitative real time PCR and immunohistochemistry on 10 formalin-fixed, paraffin-embedded samples each of canine non-neoplastic mammary tissue, mammary adenomas and adenocarcinomas.

Results: The highest PRLR expression levels were found in normal mammary tissue, while adenomas, and to an even higher degree adenocarcinomas, showed a significant decrease in prolactin receptor expression. Compared to normal tissue, PRLR mRNA was reduced 2.4 fold (p = 0.0261) in adenomas and 4.8 fold (p = 0.008) in adenocarcinomas. PRLR mRNA expression was significantly lower in malignant than in benign lesions (p = 0.0165). Immunohistochemistry demonstrated PRLR expression in all three tissue types with signals mostly limited to epithelial cells.

Conclusions: Malignant transformation of mammary tissue was associated with a decline in prolactin receptor expression. Further studies are warranted to address the functional significance of this finding.
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http://dx.doi.org/10.1186/1746-6148-8-72DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3488549PMC
May 2012

Luteal and placental function in the bitch: spatio-temporal changes in prolactin receptor (PRLr) expression at dioestrus, pregnancy and normal and induced parturition.

Reprod Biol Endocrinol 2011 Aug 3;9:109. Epub 2011 Aug 3.

Institute of Veterinary Anatomy, University of Zurich, Zurich, Switzerland.

Background: Endocrine mechanisms governing canine reproductive function remain still obscure. Progesterone (P4) of luteal origin is required for maintenance of pregnancy. Corpora lutea (CL) are gonadotrop-independent during the first third of dioestrus; afterwards prolactin (PRL) is the primary luteotropic factor. Interestingly, the increasing PRL levels are accompanied by decreasing P4 concentrations, thus luteal regression/luteolysis occurs in spite of an increased availability of gonadotropic support. PRL acts through its receptor (PRLr), the expression of which has not yet been thoroughly investigated at the molecular and cellular level in the dog.

Methods: The expression of PRLr was assessed in CL of non-pregnant dogs during the course of dioestrus (days 5, 15, 25, 35, 45, 65 post ovulation; p.o.) as well as in CL, the utero/placental compartments (Ut/Pl) and interplacental free polar zones (interplacental sites) from pregnant dogs during the pre-implantation, post-implantation and mid-gestation period of pregnancy and during the normal and antigestagen-induced luteolysis. Expression of PRLr was tested by Real Time PCR, immunohistochemistry and in situ hybridization.

Results: In non-pregnant CL the PRLr expression was significantly upregulated at day 15 p.o. and decreased significantly afterwards, towards the end of dioestrus. CL of pregnancy showed elevated PRLr expression until mid gestation while prepartal downregulation was observed. Interestingly, placental but not interplacental expression of PRLr was strongly time-related; a significant upregulation was observed towards mid-gestation. Within the CL PRLr was localized to the luteal cells; in the Ut/Pl it was localized to the fetal trophoblast and epithelial cells of glandular chambers. Moreover, in mid-pregnant animals treated with an antigestagen, both the luteal and placental, but not the uterine PRLr were significantly downregulated.

Conclusions: The data presented suggest that the luteal provision of P4 in both pregnant and non-pregnant dogs may be regulated at the PRLr level. Furthermore, a role of PRL not only in maintaining the canine CL function but also in regulating the placental function is strongly suggested. A possible functional interrelationship between luteal P4 and placental and luteal PRLr expression also with respect to the prepartal luteolysis is implied.
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http://dx.doi.org/10.1186/1477-7827-9-109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3171325PMC
August 2011

The canine hair cycle - a guide for the assessment of morphological and immunohistochemical criteria.

Vet Dermatol 2011 Oct 14;22(5):383-95. Epub 2011 Mar 14.

Institute of Animal Pathology, Vetsuisse Faculty, University of Bern, Länggassstrasse 122, CH-3012 Bern, Switzerland.

The hair follicle has a lifelong capacity to cycle through recurrent phases of controlled growth (anagen), regression (catagen) and quiescence (telogen), each associated with specific morphological changes. A comprehensive classification scheme is available for mice to distinguish the cycle stages anagen I-VI, catagen I-VIII and telogen. For dogs, such a classification system does not exist, although alopecia associated with hair cycle arrest is common. We applied analogous morphological criteria and various staining techniques to subdivide the canine hair cycle stages to the same extent as has been done in mice. Of all the staining techniques applied, haematoxylin and eosin stain, Sacpic, Masson Fontana and immunohistochemistry for vimentin and laminin proved to be most useful. To evaluate the applicability of our criteria, we investigated skin biopsies from healthy beagle dogs (n=20; biopsies from shoulder and thigh) kept in controlled conditions. From each biopsy, at least 50 hair follicles were assessed. Statistical analysis revealed that 30% of the follicles were in anagen (12% early and 18% late), 8% in catagen (2% early, 5% late and 1% not determinable) and 27% in telogen. Thirty-five per cent of hair follicles could not be assigned to a specific cycle stage because not all follicles within one biopsy were oriented perfectly. In conclusion, this guide will not only be helpful for the investigation of alopecic disorders and possibly their pathogenesis, but may also serve as a basis for research projects in which the comparison of hair cycle stages is essential, e.g. comparative analysis of gene expression patterns.
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http://dx.doi.org/10.1111/j.1365-3164.2011.00963.xDOI Listing
October 2011

Hyperglycaemia but not hyperlipidaemia decreases serum amylase and increases neutrophils in the exocrine pancreas of cats.

Res Vet Sci 2010 Aug 4;89(1):20-6. Epub 2010 Feb 4.

Clinic for Small Animal Internal Medicine, Vetsuisse Faculty, University of Zürich, Winterthurerstrasse 260, 8057 Zürich, Switzerland.

The goal of the study was to determine whether hyperglycaemia or hyperlipidaemia causes pancreatitis in cats and to assess the effect of excess serum glucose and lipids on amylase and lipase activity. Ten-day hyperglycaemic and hyperlipidaemic clamps were carried out in five and six healthy cats, respectively. Ten healthy cats received saline and served as controls. The activity of amylase was below the normal range in 4 of 5 hyperglycaemic cats by day 10. The activity of lipase did not vary in any of the cats. Samples of exocrine pancreas were normal on histological examination, but the number of tissue neutrophils was increased in hyperglycaemic cats (P<0.05). In a retrospective study 14 of 40 (35%) cats with naturally occurring diabetes mellitus had amylase activities below the reference range at the time of admission. Amylase activities normalised within 1 week of insulin therapy and subsequent glycaemic control. Lipase activity was increased in 26 of 40 (65%) diabetic cats and remained elevated despite glycaemic control. In conclusion, hyperglycaemia, but not hyperlipidaemia, increases pancreatic neutrophils in cats. However, because the histological morphology of the exocrine pancreas was normal, hyperglycaemia may play only a minor role in the pathogenesis of pancreatitis. Low amylase activities in diabetic cats may reflect an imbalance in glucose metabolism rather than pancreatitis.
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http://dx.doi.org/10.1016/j.rvsc.2010.01.006DOI Listing
August 2010

Assessment of six different collagenase-based methods to isolate feline pancreatic islets.

Res Vet Sci 2009 Dec 21;87(3):367-72. Epub 2009 Apr 21.

Clinic for Small Animal Internal Medicine, Vetsuisse Faculty, University of Zürich, Winterthurerstr. 260, 8057 Zürich, Switzerland.

Isolation of pancreatic islets is necessary to study the molecular mechanisms underlying beta-cell demise in diabetic cats. Six collagenase-based methods of isolation were compared in 10 cat pancreata, including single and double course of collagenase, followed or not by Ficoll centrifugation or accutase, and collagenase plus accutase. Morphometric analysis was performed to measure the relative area of islet and exocrine tissue. Islet specific mRNA transcripts were quantified in isolates by real-time PCR. The single and double course of collagenase digestion was successful in each cat and provided similar islet-to-exocrine tissue ratio. Quantities of insulin mRNA did not differ between the two methods. However, on histological examination either method yielded only approximately 2% of pure islets. The other methods provided disrupted islets or insufficient samples in 1-7 cats. Although pancreas digestion with single and double course of collagenase was superior, further studies are needed to improve islet isolation in cats.
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http://dx.doi.org/10.1016/j.rvsc.2009.03.017DOI Listing
December 2009

Quantitative real-time PCR detection of insulin signalling-related genes in pancreatic islets isolated from healthy cats.

Vet J 2010 Mar 31;183(3):287-93. Epub 2008 Dec 31.

Clinic for Small Animal Internal Medicine, Vetsuisse Faculty, University of Zürich, Winterthurerstr. 268, 8057 Zürich, Switzerland.

The cat has recently been proposed as a valuable model for type 2 diabetes mellitus (T2DM), because feline diabetes shares several similarities with the disease in humans. Impaired beta-cell function, decreased beta-cell mass, insulin resistance that is often related to obesity, and pancreatic amyloid deposition, are among these common features. In this study, and to further develop the cat as a model of T2DM, feline pancreatic islets were isolated and real-time PCR quantification of mRNA transcripts of genes central to beta-cell function and survival established. In particular, mRNA quantification systems were determined for insulin, the insulin enhancer pancreatic duodenal homeobox-1 (PDX-1), the insulin suppressor CCAAT/enhancer binding protein-beta (C/EBPbeta), glucose transporter isoform 2 (GLUT2), Fas receptor, the caspase-8 inhibitor FLIP (FLICE [caspase-8]-inhibitory protein) and two chemokines, interleukin (IL)-8 and monocyte chemoattractant protein-1 (MCP-1). Pancreatic islets were isolated by collagenase digestion from healthy cat donors. Partial feline mRNA sequences were determined for PDX-1, C/EBPbeta, GLUT2 and FLIP using primers identified from conserved regions of human, dog and rat mRNA. These novel and the previously available sequences (insulin, Fas receptor, IL-8 and MCP-1) were used to design feline-specific primers suitable for real-time PCR in isolated pancreatic islets. The adopted protocol of collagenase digestion yielded pancreatic islets that were frequently surrounded by acinar cells. Quantification of mRNA transcripts was simple and reproducible in healthy cats. Characterisation of genes related to insulin signalling in cats will prove useful to better understand the pathogenesis of feline diabetes and possibly of human T2DM.
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http://dx.doi.org/10.1016/j.tvjl.2008.11.012DOI Listing
March 2010

Aberrant chlamydial developmental forms in the gastrointestinal tract of pigs spontaneously and experimentally infected with Chlamydia suis.

Vet Microbiol 2009 Mar 13;135(1-2):147-56. Epub 2008 Sep 13.

Institute of Veterinary Pathology, University of Zurich, Vetsuisse Faculty, Winterthurerstrasse 268, CH-8057 Zurich, Switzerland.

The phenomenon of persistence is well known from in vitro studies, where it is associated with the production of aberrant bodies, but its occurrence in vivo is less well documented. The objective of this study was to search for aberrant bodies in intestinal tissues from pigs, describe their ultrastructure, and investigate the suitability of immunohistochemical staining for chlamydial heat shock protein 60 (cHSP60) to detect such forms. Intestinal tissues derived from pigs naturally and experimentally infected with Chlamydia (C.) suis were examined by immunohistochemistry, transmission electron microscopy and immunogold electron microscopy. The chlamydial species involved in the natural infection were determined using an Array Tube Microarray to C. suis and Chlamydophila abortus. Ultrastructurally, aberrant bodies were detected in the gut of both naturally and experimentally infected pigs. Immunogold electron microscopy showed that the aberrant bodies were labeled less strongly than the normal forms by antibodies against LPS and cHSP60 respectively. It was concluded that aberrant bodies occur in vivo in pigs and that the gnotobiotic pig model might be suitable for the study of chlamydial persistence in vivo. The antibody against cHSP60 does not appear to be suitable to specifically detect such forms.
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http://dx.doi.org/10.1016/j.vetmic.2008.09.035DOI Listing
March 2009

Experimental enteric infection of gnotobiotic piglets with Chlamydia suis strain S45.

Vet Microbiol 2009 Mar 13;135(1-2):157-68. Epub 2008 Sep 13.

Institute of Veterinary Pathology, University of Zurich, Vetsuisse Faculty, Winterthurerstr. 268, CH-8057 Zurich, Switzerland.

Enteric chlamydial infections of pigs with Chlamydia (C.) suis are frequent and often subclinical. The enteric pathogenicity of C. suis strain S45 was investigated in gnotobiotic piglets. Piglets from three litters (n=31) were inoculated with egg-grown chlamydiae at 2-3 days of age (n=17) or used as controls (n=14). They were observed for clinical signs, killed and necropsied sequentially at 2-13 days postinoculation (DPI). Feces were collected daily and investigated with an ELISA for chlamydial antigen. At necropsy, specimens were collected for histopathology and for immunohistochemical, PCR-based, and serological (complement fixation test, ELISA) detection of chlamydiae. Chlamydial replication and associated symptoms and lesions were observed from 2 to 13 DPI and were particularly pronounced within the first week PI. Clinical symptoms consisted of moderate-to-severe diarrhea, slight and transient anorexia, weakness and body weight loss. Immunohistochemistry and ELISA revealed that chlamydial replication was particularly marked at 2-4 DPI and primarily located in the small intestinal villus enterocytes. Further sites of replication included large intestinal enterocytes, the lamina propria and Tunica submucosa, and the mesenteric lymphnodes. Histopathological changes included moderate-to-severe villus atrophy with flattened enterocytes and focal villus tip erosions, and moderate mucosal inflammatory cell infiltrates and lymphangitis in the small intestine. PCR of spleen tissue and blood was mostly negative for chlamydiae, indicating that they did not substantially disseminate into the host up to 13 DPI. All sera were negative for anti-chlamydial antibodies. In conclusion, C. suis strain S45 elicited significant enteric disease and lesions in gnotobiotic piglets indicating its pathogenic potential for swine.
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http://dx.doi.org/10.1016/j.vetmic.2008.09.038DOI Listing
March 2009

Validation of tissue microarrays for immunohistochemical analyses of canine lymphomas.

J Vet Diagn Invest 2007 Nov;19(6):652-9

Institut für Veterinärpathologie, Vetsuisse-Fakultät, Universität Zürich, Winterthurerstrasse 268, CH-8057 Zürich, Switzerland.

In most validation studies of tissue microarrays (TMAs), a fixed number of cores with a given diameter are analyzed to determine the degree of accuracy by which the TMA represents the whole section. The statistical model described in the present study predicts this property for various combinations of 2 core sizes (0.6 mm and 1.2 mm) and different core numbers. The model was based on artificial TMA core biopsies generated from Ki-67 and active caspase-3 immunostains of 40 canine lymphoma samples. Positivity was scored on a continuous scale, and a large number of cells were analyzed with the help of semiautomated cell counting. Despite considerable differences in range and distribution of Ki-67 and active caspase-3 positivity values, the model predictions showed a high degree of agreement for both markers. Comparison of 0.6 mm and 1.2 mm cores indicated that the use of small cores necessitates inclusion of a larger number of samples but requires counting a markedly smaller number of cells. Suitability of TMAs to determine the immunophenotype of the whole section was assessed using 2 different combinations of core sizes and numbers. Both displayed a high degree of concordance with the whole section (kappa(0.6) = 0.79; kappa(1.2) = 0.91). The present study provides a basis for the use of TMAs in future high-throughput immunohistochemical investigations of selected markers in canine lymphomas. The statistical model presented can be used to determine an optimal TMA design depending on a desired accuracy.
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http://dx.doi.org/10.1177/104063870701900606DOI Listing
November 2007

Simultaneous application of the vascular endothelial growth factor (VEGF) receptor inhibitor PTK787/ZK 222584 and ionizing radiation does not further reduce the growth of canine oral melanoma xenografts in nude mice.

Vet J 2007 May 20;173(3):564-70. Epub 2007 Apr 20.

Section of Diagnostic Imaging and Radio-Oncology, Vetsuisse Faculty, University of Zürich, Switzerland.

PTK787/ZK 222584 is an inhibitor of the vascular endothelial growth factor (VEGF) receptor tyrosine kinases. In this study, the effectiveness of PTK787/ZK 222584 and radiation on canine oral melanoma xenografts was assessed. Xenografts were treated with radiotherapy (4x6Gy), or with PTK787/ZK 222584, or with a combination of both. Treatment efficacy was assessed using a tumour growth delay assay, serial power Doppler and pO(2) measurements during and after therapy. There was a significant growth delay for the group treated with radiation alone and for the combined treatment group. However, tumour growth delay was similar in both groups. Tumours were hypoxic before irradiation and no significant re-oxygenation occurred during therapy. In all tumours, vascularity and perfusion were significantly lower at the end of the study but no significant differences in perfusion, vascularity and oxygenation status were observed between and within treatment groups. The combination of PTK787/ZK 222584 and radiotherapy did not perform better than radiotherapy alone in this model.
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http://dx.doi.org/10.1016/j.tvjl.2007.03.001DOI Listing
May 2007