Publications by authors named "Saila Holopainen"

9 Publications

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A missense variant in IFT122 associated with a canine model of retinitis pigmentosa.

Hum Genet 2021 Nov 19;140(11):1569-1579. Epub 2021 Feb 19.

Department of Veterinary Biosciences, University of Helsinki, Helsinki, Finland.

Retinitis pigmentosa (RP) is a blinding eye disease affecting nearly two million people worldwide. Dogs are affected with a similar illness termed progressive retinal atrophy (PRA). Lapponian herders (LHs) are affected with several types of inherited retinal dystrophies, and variants in PRCD and BEST1 genes have been associated with generalized PRA and canine multifocal retinopathy 3 (cmr3), respectively. However, all retinal dystrophy cases in LHs are not explained by these variants, indicating additional genetic causes of disease in the breed. We collected DNA samples from 10 PRA affected LHs, with known PRCD and BEST1 variants excluded, and 34 unaffected LHs. A genome-wide association study identified a locus on CFA20 (p = 2.4 × 10, p = 0.035), and subsequent whole-genome sequencing of an affected LH revealed a missense variant, c.3176G>A, in the intraflagellar transport 122 (IFT122) gene. The variant was also found in Finnish Lapphunds, in which its clinical relevancy needs to be studied further. The variant interrupts a highly conserved residue, p.(R1059H), in IFT122 and likely impairs its function. Variants in IFT122 have not been associated with retinal degeneration in mammals, but the loss of ift122 in zebrafish larvae impaired opsin transport and resulted in progressive photoreceptor degeneration. Our study establishes a new spontaneous dog model to study the role of IFT122 in RP biology, while the affected breed will benefit from a genetic test for a recessive condition.
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http://dx.doi.org/10.1007/s00439-021-02266-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8519925PMC
November 2021

Matrix metalloproteinase-2, -7, and -9 activities in dogs with idiopathic pulmonary fibrosis compared to healthy dogs and dogs with other respiratory diseases.

J Vet Intern Med 2021 Jan 4;35(1):462-471. Epub 2020 Dec 4.

Faculty of Veterinary Medicine, Department of Equine and Small Animal Medicine, University of Helsinki, Helsinki, Finland.

Background: Canine idiopathic pulmonary fibrosis (CIPF) is a chronic, interstitial lung disease that mainly affects West Highland white terriers (WHWTs) and is characterized by excessive deposition of extracellular matrix (ECM) in the lung. Matrix metalloproteinases (MMPs) participate in remodeling of ECM.

Objectives: To compare metalloproteinase-2, -7 and -9 activities in blood or bronchoalveolar lavage fluid (BALF) samples or both of CIPF WHWTs with healthy WHWTs, healthy dogs of other breeds, and dogs with other lung diseases and determine if these MMPs could be used as diagnostic and prognostic markers for CIPF.

Animals: Forty-four CIPF WHWTs, 24 dogs with chronic bronchitis (CB), 17 with eosinophilic bronchopneumopathy (EBP), 10 with bacterial pneumonia, 39 healthy WHWTs, and 35 healthy dogs of other breeds.

Methods: Cross-sectional observational study. Pro-MMP and active MMP activities were analyzed by zymography.

Results: In serum, significantly higher (P < .01) pro-MMP-7 activities were observed in CIPF WHWTs compared to healthy dogs of other breeds, dogs with CB and dogs with EBP. In BALF of CIPF WHWTs, both pro-MMP-9 and pro-MMP-2 activities were significantly higher (P < .01) compared to healthy WHWTs, but these differences were not detected in plasma. The CIPF WHWTs had significantly higher (P < .05) activities of pro-MMP-9 compared to dogs with CB and of pro-MMP-2 compared to dogs with CB and EBP. No statistically significant prognostic factors were observed in CIPF WHWTs.

Conclusions And Clinical Relevance: Serum MMP-7 and BALF MMP-2 and -9 potentially may be useful diagnostic markers but not prognostic markers for CIPF.
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http://dx.doi.org/10.1111/jvim.15970DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7848316PMC
January 2021

A putative silencer variant in a spontaneous canine model of retinitis pigmentosa.

PLoS Genet 2020 03 9;16(3):e1008659. Epub 2020 Mar 9.

Department of Veterinary Biosciences, University of Helsinki, Helsinki, Finland.

Retinitis pigmentosa (RP) is the leading cause of blindness with nearly two million people affected worldwide. Many genes have been implicated in RP, yet in 30-80% of the RP patients the genetic cause remains unknown. A similar phenotype, progressive retinal atrophy (PRA), affects many dog breeds including the Miniature Schnauzer. We performed clinical, genetic and functional experiments to identify the genetic cause of PRA in the breed. The age of onset and pattern of disease progression suggested that at least two forms of PRA, types 1 and 2 respectively, affect the breed, which was confirmed by genome-wide association study that implicated two distinct genomic loci in chromosomes 15 and X, respectively. Whole-genome sequencing revealed a fully segregating recessive regulatory variant in type 1 PRA. The associated variant has a very recent origin based on haplotype analysis and lies within a regulatory site with the predicted binding site of HAND1::TCF3 transcription factor complex. Luciferase assays suggested that mutated regulatory sequence increases expression. Case-control retinal expression comparison of six best HAND1::TCF3 target genes were analyzed with quantitative reverse-transcriptase PCR assay and indicated overexpression of EDN2 and COL9A2 in the affected retina. Defects in both EDN2 and COL9A2 have been previously associated with retinal degeneration. In summary, our study describes two genetically different forms of PRA and identifies a fully penetrant variant in type 1 form with a possible regulatory effect. This would be among the first reports of a regulatory variant in retinal degeneration in any species, and establishes a new spontaneous dog model to improve our understanding of retinal biology and gene regulation while the affected breed will benefit from a reliable genetic testing.
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http://dx.doi.org/10.1371/journal.pgen.1008659DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7082071PMC
March 2020

Genetic dissection of canine hip dysplasia phenotypes and osteoarthritis reveals three novel loci.

BMC Genomics 2019 Dec 27;20(1):1027. Epub 2019 Dec 27.

Department of Veterinary Biosciences, University of Helsinki, P.O. Box 66 (Mustialankatu 1), FI-00014, Helsinki, Finland.

Background: Hip dysplasia and osteoarthritis continue to be prevalent problems in veterinary and human medicine. Canine hip dysplasia is particularly problematic as it massively affects several large-sized breeds and can cause a severe impairment of the quality of life. In Finland, the complex condition is categorized to five classes from normal to severe dysplasia, but the categorization includes several sub-traits: congruity of the joint, Norberg angle, subluxation degree of the joint, shape and depth of the acetabulum, and osteoarthritis. Hip dysplasia and osteoarthritis have been proposed to have separate genetic etiologies.

Results: Using Fédération Cynologique Internationale -standardized ventrodorsal radiographs, German shepherds were rigorously phenotyped for osteoarthritis, and for joint incongruity by Norberg angle and femoral head center position in relation to dorsal acetabular edge. The affected dogs were categorized into mild, moderate and severe dysplastic phenotypes using official hip scores. Three different genome-wide significant loci were uncovered. The strongest candidate genes for hip joint incongruity were noggin (NOG), a bone and joint developmental gene on chromosome 9, and nanos C2HC-type zinc finger 1 (NANOS1), a regulator of matrix metalloproteinase 14 (MMP14) on chromosome 28. Osteoarthritis mapped to a long intergenic region on chromosome 1, between genes encoding for NADPH oxidase 3 (NOX3), an intriguing candidate for articular cartilage degradation, and AT-rich interactive domain 1B (ARID1B) that has been previously linked to joint laxity.

Conclusions: Our findings highlight the complexity of canine hip dysplasia phenotypes. In particular, the results of this study point to the potential involvement of specific and partially distinct loci and genes or pathways in the development of incongruity, mild dysplasia, moderate-to-severe dysplasia and osteoarthritis of canine hip joints. Further studies should unravel the unique and common mechanisms for the various sub-traits.
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http://dx.doi.org/10.1186/s12864-019-6422-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6935090PMC
December 2019

Novel protective and risk loci in hip dysplasia in German Shepherds.

PLoS Genet 2019 07 19;15(7):e1008197. Epub 2019 Jul 19.

Department of Veterinary Biosciences, University of Helsinki, Helsinki, Finland.

Canine hip dysplasia is a common, non-congenital, complex and hereditary disorder. It can inflict severe pain via secondary osteoarthritis and lead to euthanasia. An analogous disorder exists in humans. The genetic background of hip dysplasia in both species has remained ambiguous despite rigorous studies. We aimed to investigate the genetic causes of this disorder in one of the high-risk breeds, the German Shepherd. We performed genetic analyses with carefully phenotyped case-control cohorts comprising 525 German Shepherds. In our genome-wide association studies we identified four suggestive loci on chromosomes 1 and 9. Targeted resequencing of the two loci on chromosome 9 from 24 affected and 24 control German Shepherds revealed deletions of variable sizes in a putative enhancer element of the NOG gene. NOG encodes for noggin, a well-described bone morphogenetic protein inhibitor affecting multiple developmental processes, including joint development. The deletion was associated with the healthy controls and mildly dysplastic dogs suggesting a protective role against canine hip dysplasia. Two enhancer variants displayed a decreased activity in a dual luciferase reporter assay. Our study identifies novel loci and candidate genes for canine hip dysplasia, with potential regulatory variants in the NOG gene. Further research is warranted to elucidate how the identified variants affect the expression of noggin in canine hips, and what the potential effects of the other identified loci are.
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http://dx.doi.org/10.1371/journal.pgen.1008197DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6668854PMC
July 2019

Thoracic high resolution CT using the modified VetMousetrap™ device is a feasible method for diagnosing canine idiopathic pulmonary fibrosis in awake West Highland White Terriers.

Vet Radiol Ultrasound 2019 Sep 7;60(5):525-532. Epub 2019 Jun 7.

Department of Equine and Small Animal Medicine, Faculty of Veterinary Medicine, University of Helsinki, Helsinki, Finland.

Canine idiopathic pulmonary fibrosis is a chronic, progressive interstitial lung disease particularly prevalent in West Highland White Terriers. In the present prospective pilot study, we evaluated the feasibility of modified VetMousetrap™ device in high resolution CT to detect idiopathic pulmonary fibrosis in West Highland White Terriers. Twelve awake West Highland White Terriers with canine idiopathic pulmonary fibrosis and 24 clinically healthy West Highland White Terriers were scanned using a helical dual slice scanner utilizing VetMousetrap™ device without or with minimal chemical restraint with butorphanol. Three evaluators blindly assessed the images for image quality and the presence of canine idiopathic pulmonary fibrosis related imaging findings such as ground glass opacity and reticular opacities. Additionally, the attenuation of the lung was quantified with ImageJ software using histogram analysis of density over the lung fields. Computed tomography was successfully completed and motion artifact ranked in statistical analysis barely noticeable to mild in all dogs. The agreement between imaging findings and clinical status was very good with overall κ value 0.91 and percentage of agreement of 94%. There was also very good intraobserver (κ = 0.79-0.91) and interobserver agreement (κ = 0.94). Moderate to severe ground glass opacity was present in all affected dogs. In the ImageJ analysis, a significant difference in lung attenuation between the study groups was observed. We conclude that modified VetMousetrap™ device is applicable in diagnosing canine idiopathic pulmonary fibrosis in awake West Highland White Terriers avoiding anesthetic risk in these often severely hypoxic patients.
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http://dx.doi.org/10.1111/vru.12779DOI Listing
September 2019

Reflux aspiration in lungs of dogs with respiratory disease and in healthy West Highland White Terriers.

J Vet Intern Med 2018 Nov 12;32(6):2074-2081. Epub 2018 Oct 12.

Department of Equine and Small Animal Medicine, Faculty of Veterinary Medicine, University of Helsinki, Finland.

Background: Gastroesophageal reflux and microaspiration (MA) of gastric juice are associated with various human respiratory diseases but not in dogs.

Objective: To detect the presence of bile acids in bronchoalveolar lavage fluid (BALF) of dogs with various respiratory diseases.

Animals: Twenty-seven West Highland White Terriers (WHWTs) with canine idiopathic pulmonary fibrosis (CIPF), 11 dogs with bacterial pneumonia (BP), 13 with chronic bronchitis (CB), 9 with eosinophilic bronchopneumopathy (EBP), 19 with laryngeal dysfunction (LD), 8 Irish Wolfhounds (IWHs) with previous BPs, 13 healthy WHWTs, all privately owned dogs, and 6 healthy research colony Beagles METHODS: Prospective cross-sectional observational study with convenience sampling of dogs. Bile acids were measured by mass spectrometry in BALF samples. Total bile acid (TBA) concentration was calculated as a sum of 17 different bile acids.

Results: Concentrations of TBA were above the limit of quantification in 78% of CIPF, 45% of BP, 62% of CB, 44% of EBP, 68% of LD, and 13% of IWH dogs. In healthy dogs, bile acids were detected less commonly in Beagles (0/6) than in healthy WHWTs (10/13). Concentrations of TBA were significantly higher in CIPF (median 0.013 μM, range not quantifiable [n.q.]-0.14 μM, P < .001), healthy WHWTs (0.0052 μM, n.q.-1.2 μM, P = .003), LD (0.010 μM, n.q.-2.3 μM, P = .015), and CB (0.0078 μM, n.q.-0.073 μM, P = .018) groups compared to Beagles (0 μM, n.q.).

Conclusion And Clinical Importance: These results suggest that MA occurs in various respiratory diseases of dogs and also in healthy WHWTs.
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http://dx.doi.org/10.1111/jvim.15321DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6271311PMC
November 2018

ANLN truncation causes a familial fatal acute respiratory distress syndrome in Dalmatian dogs.

PLoS Genet 2017 02 21;13(2):e1006625. Epub 2017 Feb 21.

Department of Veterinary Biosciences, University of Helsinki, Helsinki, Finland.

Acute respiratory distress syndrome (ARDS) is the leading cause of death in critical care medicine. The syndrome is typified by an exaggerated inflammatory response within the lungs. ARDS has been reported in many species, including dogs. We have previously reported a fatal familial juvenile respiratory disease accompanied by occasional unilateral renal aplasia and hydrocephalus, in Dalmatian dogs. The condition with a suggested recessive mode of inheritance resembles acute exacerbation of usual interstitial pneumonia in man. We combined SNP-based homozygosity mapping of two ARDS-affected Dalmatian dogs and whole genome sequencing of one affected dog to identify a case-specific homozygous nonsense variant, c.31C>T; p.R11* in the ANLN gene. Subsequent analysis of the variant in a total cohort of 188 Dalmatians, including seven cases, indicated complete segregation of the variant with the disease and confirmed an autosomal recessive mode of inheritance. Low carrier frequency of 1.7% was observed in a population cohort. The early nonsense variant results in a nearly complete truncation of the ANLN protein and immunohistochemical analysis of the affected lung tissue demonstrated the lack of the membranous and cytoplasmic staining of ANLN protein in the metaplastic bronchial epithelium. The ANLN gene encodes an anillin actin binding protein with a suggested regulatory role in the integrity of intercellular junctions. Our study suggests that defective ANLN results in abnormal cellular organization of the bronchiolar epithelium, which in turn predisposes to acute respiratory distress. ANLN has been previously linked to a dominant focal segmental glomerulosclerosis in human without pulmonary defects. However, the lack of similar renal manifestations in the affected Dalmatians suggest a novel ANLN-related pulmonary function and disease association.
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http://dx.doi.org/10.1371/journal.pgen.1006625DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5340406PMC
February 2017

Assessment of CCL2 and CXCL8 chemokines in serum, bronchoalveolar lavage fluid and lung tissue samples from dogs affected with canine idiopathic pulmonary fibrosis.

Vet J 2015 Oct 4;206(1):75-82. Epub 2015 Jun 4.

Faculty of Veterinary Medicine, University of Liège, Bd de Colonster, Liège, Belgium.

Canine idiopathic pulmonary fibrosis (CIPF) is a progressive disease of the lung parenchyma that is more prevalent in dogs of the West Highland white terrier (WHWT) breed. Since the chemokines (C-C motif) ligand 2 (CCL2) and (C-X-C motif) ligand 8 (CXCL8) have been implicated in pulmonary fibrosis in humans, the aim of the present study was to investigate whether these same chemokines are involved in the pathogenesis of CIPF. CCL2 and CXCL8 concentrations were measured by ELISA in serum and bronchoalveolar lavage fluid (BALF) from healthy dogs and WHWTs affected with CIPF. Expression of the genes encoding CCL2 and CXCL8 and their respective receptors, namely (C-C motif) receptor 2 (CCR2) and (C-X-C motif) receptor 2 (CXCR2), was compared in unaffected lung tissue and biopsies from dogs affected with CIPF by quantitative PCR and localisation of CCL2 and CXCL8 proteins were determined by immunohistochemistry. Significantly greater CCL2 and CXCL8 concentrations were found in the BALF from WHWTs affected with CIPF, compared with healthy dogs. Significantly greater serum concentrations of CCL2, but not CXCL8, were found in CIPF-affected dogs compared with healthy WHWTs. No differences in relative gene expression for CCL2, CXCL8, CCR2 or CXCR2 were observed when comparing lung biopsies from control dogs and those affected with CIPF. In affected lung tissues, immunolabelling for CCL2 and CXCL8 was observed in bronchial airway epithelial cells in dogs affected with CIPF. The study findings suggest that both CCL2 and CXCL8 are involved in the pathogenesis of CIPF. Further studies are required to determine whether these chemokines might have a clinical use as biomarkers of fibrosis or as targets for therapeutic intervention.
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http://dx.doi.org/10.1016/j.tvjl.2015.06.001DOI Listing
October 2015
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