@article{sai_nakanishi_scofield_tokarz_linder_cohen_ninomiya-tsuji_2023, title={Aberrantly activated TAK1 links neuroinflammation and neuronal loss in Alzheimer?s disease mouse models}, volume={136}, ISSN={["1477-9137"]}, url={http://dx.doi.org/10.1242/jcs.260102}, DOI={10.1242/jcs.260102}, abstractNote={ABSTRACT Neuroinflammation is causally associated with Alzheimer's disease (AD) pathology. Reactive glia cells secrete various neurotoxic factors that impair neuronal homeostasis eventually leading to neuronal loss. Although the glial activation mechanism in AD has been relatively well studied, how it perturbs intraneuronal signaling, which ultimately leads to neuronal cell death, remains poorly understood. Here, we report that compound stimulation with the neurotoxic factors TNF and glutamate aberrantly activates neuronal TAK1 (also known as MAP3K7), which promotes the pathogenesis of AD in mouse models. Glutamate-induced Ca2+ influx shifts TNF signaling to hyper-activate TAK1 enzymatic activity through Ca2+/calmodulin-dependent protein kinase II, which leads to necroptotic cellular damage. Genetic ablation and pharmacological inhibition of TAK1 ameliorated AD-associated neuronal loss and cognitive impairment in the AD model mice. Our findings provide a molecular mechanism linking cytokines, Ca2+ signaling and neuronal necroptosis in AD.}, number={6}, journal={JOURNAL OF CELL SCIENCE}, publisher={The Company of Biologists}, author={Sai, Kazuhito and Nakanishi, Aoi and Scofield, Kimberly M. and Tokarz, Debra A. and Linder, Keith E. and Cohen, Todd J. and Ninomiya-Tsuji, Jun}, year={2023}, month={Mar} }
 @article{krane_shockley_malarkey_miller_miller_tokarz_jensen_janardhan_breen_mariani_2022, title={Inter-pathologist agreement on diagnosis, classification and grading of canine glioma}, volume={7}, ISSN={["1476-5829"]}, url={https://doi.org/10.1111/vco.12853}, DOI={10.1111/vco.12853}, abstractNote={Histopathological evaluation of tumours is a subjective process, but studies of inter-pathologist agreement are uncommon in veterinary medicine. The Comparative Brain Tumour Consortium (CBTC) recently published diagnostic criteria for canine gliomas. Our objective was to assess the degree of inter-pathologist agreement on intracranial canine gliomas, utilising the CBTC diagnostic criteria in a cohort of eighty-five samples from dogs with an archival diagnosis of intracranial glioma. Five pathologists independently reviewed H&E and immunohistochemistry sections and provided a diagnosis and grade. Percentage agreement and kappa statistics were calculated to measure inter-pathologist agreement between pairs and amongst the entire group. A consensus diagnosis of glioma subtype and grade was achieved for 71/85 (84%) cases. For these cases, percentage agreement on combined diagnosis (subtype and grade), subtype only and grade only were 66%, 80% and 82%, respectively. Kappa statistics for the same were 0.466, 0.542 and 0.516, respectively. Kappa statistics for oligodendroglioma, astrocytoma and undefined glioma were 0.585, 0.566 and 0.280 and were 0.516 for both low-grade and high-grade tumours. Kappa statistics amongst pairs of pathologists for combined diagnosis varied from 0.352 to 0.839. 8 % of archival oligodendrogliomas and 61% of archival astrocytomas were reclassified as another entity after review. Inter-pathologist agreement utilising CBTC guidelines for canine glioma was moderate overall but varied from fair to almost perfect between pairs of pathologists. Agreement was similar for oligodendrogliomas and astrocytomas but lower for undefined gliomas. These results are similar to pathologist agreement in human glioma studies and with other tumour entities in veterinary medicine.}, journal={VETERINARY AND COMPARATIVE ONCOLOGY}, author={Krane, Gregory A. and Shockley, Keith R. and Malarkey, David E. and Miller, Andrew D. and Miller, C. Ryan and Tokarz, Debra A. and Jensen, Heather L. and Janardhan, Kyathanahalli S. and Breen, Matthew and Mariani, Christopher L.}, year={2022}, month={Jul} }
 @article{krane_carly a. o'dea_malarkey_miller_miller_tokarz_jensen_janardhan_shockley_flagler_et al._2021, title={Immunohistochemical evaluation of immune cell infiltration in canine gliomas}, volume={7}, ISSN={["1544-2217"]}, DOI={10.1177/03009858211023946}, abstractNote={Evasion of the immune response is an integral part of the pathogenesis of glioma. In humans, important mechanisms of immune evasion include recruitment of regulatory T cells (Tregs) and polarization of macrophages toward an M2 phenotype. Canine glioma has a robust immune cell infiltrate that has not been extensively characterized. The purpose of this study was to determine the distribution of immune cells infiltrating spontaneous intracranial canine gliomas. Seventy-three formalin-fixed, paraffin-embedded tumor samples were evaluated using immunohistochemistry for CD3, forkhead box 3 (FOXP3), CD20, Iba1, calprotectin (Mac387), CD163, and indoleamine 2,3-dioxygenase (IDO). Immune cell infiltration was present in all tumors. Low-grade and high-grade gliomas significantly differed in the numbers of FoxP3+ cells, Mac387+ cells, and CD163+ cells (}, journal={VETERINARY PATHOLOGY}, author={Krane, Gregory A. and Carly A. O'Dea and Malarkey, David E. and Miller, Andrew D. and Miller, C. Ryan and Tokarz, Debra A. and Jensen, Heather L. and Janardhan, Kyathanahalli S. and Shockley, Keith R. and Flagler, Norris and et al.}, year={2021}, month={Jul} }
 @article{bomba_sheets_valdivia_khagi_ruterbories_mariani_borst_tokarz_hingtgen_2021, title={Personalized-induced neural stem cell therapy: Generation, transplant, and safety in a large animal model}, volume={6}, ISSN={["2380-6761"]}, DOI={10.1002/btm2.10171}, abstractNote={In this study, we take an important step toward clinical translation by generating the first canine-induced neural stem cells (iNSCs). We explore key aspects of scale-up, persistence, and safety of personalized iNSC therapy in autologous canine surgery models. iNSCs are a promising new approach to treat aggressive cancers of the brain, including the deadly glioblastoma. Created by direct transdifferentiation of fibroblasts, iNSCs are known to migrate through the brain, track down invasive cancer foci, and deliver anticancer payloads that significantly reduce tumor burden and extend survival of tumor-bearing mice. Here, skin biopsies were collected from canines and converted into the first personalized canine iNSCs engineered to carry TNFα-related apoptosis-inducing ligand (TRAIL) and thymidine kinase (TK), as well as magnetic resonance imaging (MRI) contrast agents for in vivo tracking. Time-lapse analysis showed canine iNSCs efficiently migrate to human tumor cells, and cell viability assays showed both TRAIL and TK monotherapy markedly reduced tumor growth. Using intraoperative navigation and two delivery methods to closely mimic human therapy, canines received autologous iNSCs either within postsurgical cavities in a biocompatible matrix or via a catheter placed in the lateral ventricle. Both strategies were well tolerated, and serial MRI showed hypointense regions at the implant sites that remained stable through 86 days postimplant. Serial fluid sample testing following iNSC delivery showed the bimodal personalized therapy was well tolerated, with no iNSC-induced abnormal tissue pathology. Overall, this study lays an important foundation as this promising personalized cell therapy advances toward human patient testing.}, number={1}, journal={BIOENGINEERING & TRANSLATIONAL MEDICINE}, author={Bomba, Hunter N. and Sheets, Kevin T. and Valdivia, Alain and Khagi, Simon and Ruterbories, Laura and Mariani, Christopher L. and Borst, Luke B. and Tokarz, Debra A. and Hingtgen, Shawn D.}, year={2021}, month={Jan} }
 @article{reece_varikuti_kilburg-basnyat_dunigan-russell_hodge_luo_madenspacher_thomas_tokarz_tighe_et al._2021, title={Scavenger Receptor BI Attenuates IL-17A-Dependent Neutrophilic Inflammation in Asthma}, volume={64}, ISSN={["1535-4989"]}, DOI={10.1165/rcmb.2020-0007OC}, abstractNote={Asthma is a common respiratory disease currently affecting more than 300 million worldwide and is characterized by airway inflammation, hyperreactivity, and remodeling. It is a heterogeneous disease consisting of corticosteroid-sensitive T-helper cell type 2-driven eosinophilic and corticosteroid-resistant, T-helper cell type 17-driven neutrophilic phenotypes. One pathway recently described to regulate asthma pathogenesis is cholesterol trafficking. Scavenger receptors, in particular SR-BI (scavenger receptor class B type I), are known to direct cellular cholesterol uptake and efflux. We recently defined SR-BI functions in pulmonary host defense; however, the function of SR-BI in asthma pathogenesis is unknown. To elucidate the role of SR-BI in allergic asthma, SR-BI-sufficient (SR-BI}, number={6}, journal={AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR BIOLOGY}, author={Reece, Sky W. and Varikuti, Sanjay and Kilburg-Basnyat, Brita and Dunigan-Russell, Katelyn and Hodge, Myles X. and Luo, Bin and Madenspacher, Jennifer H. and Thomas, Seddon Y. and Tokarz, Debra A. and Tighe, Robert M. and et al.}, year={2021}, month={Jun}, pages={698–708} }
 @article{yaeger_reece_kilburg-basnyat_hodge_pal_dunigan-russell_luo_you_bonner_spangenburg_et al._2021, title={Sex Differences in Pulmonary Eicosanoids and Specialized Pro-Resolving Mediators in Response to Ozone Exposure}, volume={183}, ISSN={["1096-0929"]}, DOI={10.1093/toxsci/kfab081}, abstractNote={Abstract Ozone (O3) is a criteria air pollutant known to increase the morbidity and mortality of cardiopulmonary diseases. This occurs through a pulmonary inflammatory response characterized by increased recruitment of immune cells into the airspace, pro-inflammatory cytokines, and pro-inflammatory lipid mediators. Recent evidence has demonstrated sex-dependent differences in the O3-induced pulmonary inflammatory response. However, it is unknown if this dimorphic response is evident in pulmonary lipid mediator metabolism. We hypothesized that there are sex-dependent differences in lipid mediator production following acute O3 exposure. Male and female C57BL/6J mice were exposed to 1 part per million O3 for 3 h and were necropsied at 6 or 24 h following exposure. Lung lavage was collected for cell differential and total protein analysis, and lung tissue was collected for mRNA analysis, metabololipidomics, and immunohistochemistry. Compared with males, O3-exposed female mice had increases in airspace neutrophilia, neutrophil chemokine mRNA, pro-inflammatory eicosanoids such as prostaglandin E2, and specialized pro-resolving mediators (SPMs), such as resolvin D5 in lung tissue. Likewise, precursor fatty acids (arachidonic and docosahexaenoic acid; DHA) were increased in female lung tissue following O3 exposure compared with males. Experiments with ovariectomized females revealed that loss of ovarian hormones exacerbates pulmonary inflammation and injury. However, eicosanoid and SPM production were not altered by ovariectomy despite depleted pulmonary DHA concentrations. Taken together, these data indicate that O3 drives an increased pulmonary inflammatory and bioactive lipid mediator response in females. Furthermore, ovariectomy increases susceptibility to O3-induced pulmonary inflammation and injury, as well as decreases pulmonary DHA concentrations.}, number={1}, journal={TOXICOLOGICAL SCIENCES}, author={Yaeger, Michael J. and Reece, Sky W. and Kilburg-Basnyat, Brita and Hodge, Miles X. and Pal, Anandita and Dunigan-Russell, Katelyn and Luo, Bin and You, Dorothy J. and Bonner, James C. and Spangenburg, Espen E. and et al.}, year={2021}, month={Sep}, pages={170–183} }
 @article{mohan_neequaye_malur_soliman_mcpeek_leffler_ogburn_tokarz_knudson_gharib_et al._2020, title={Matrix Metalloproteinase-12 Is Required for Granuloma Progression}, volume={11}, ISSN={["1664-3224"]}, DOI={10.3389/fimmu.2020.553949}, abstractNote={Background: Sarcoidosis is a chronic inflammatory disease of unknown cause characterized by granuloma formation. Mechanisms for chronic persistence of granulomas are unknown. Matrix Metalloproteinase-12 (MMP12) degrades extracellular matrix elastin and enables infiltration of immune cells responsible for inflammation and granuloma formation. Previous studies report increased MMP12 in sarcoidosis patients and association between MMP12 expression and disease severity. We also observed elevated MMP12 in our multiwall carbon nanotube (MWCNT) murine model of granulomatous inflammation. Here we hypothesized that MMP12 is important to acute and late phases of granuloma pathogenesis. To test this hypothesis, we analyzed granulomatous and inflammatory responses of mmp12 KO mice at 10 (acute) and 60 days (late) after MWCNT instillation. Methods: C57BL/6 (wildtype) and mmp12 KO mice underwent oropharyngeal instillation of MWCNT. Lungs were harvested at 3, 10, 20, and 60 days post instillation for evaluation of MMP12 expression and granulomatous changes. Bronchoalveolar lavage (BAL) cells were analyzed 60 days after MWCNT instillation for expression of mediators thought to play a role in sarcoid granulomatosis: peroxisome proliferator-activated receptor-gamma (PPARγ), interferon-gamma (IFNγ), and CCL2 (MCP-1). Results: Pulmonary granuloma appearance at 10 days after MWCNT instillation showed no differences between wildtype and mmp12 KO mice. In contrast, by 60 days after MWCNT instillation, mmp12 KO mice revealed markedly attenuated granuloma formation together with elevated PPARγ and reduced IFNγ expression in BAL cells compared to wildtype. Unexpectedly, mmp12 KO mice further demonstrated increased alveolar macrophages with increased CCL2 at 60 days. Conclusions: The striking reduction of granuloma formation at day 60 in mmp12 KO mice suggests that MMP12 is required to maintain chronic granuloma pathophysiology. The increased PPARγ and decreased IFNγ findings suggest that these mediators also may be involved since previous studies have shown that PPARγ suppresses IFNγ and PPARγ deficiency amplifies granuloma formation. Interestingly, a role of MMP 12 in granuloma resolution is also suggested by increases in both macrophage influx and CCL2. Overall, our results strongly implicate MMP12 as a key factor in granuloma persistence and may represent an attractive therapeutic target in pulmonary sarcoidosis}, journal={FRONTIERS IN IMMUNOLOGY}, author={Mohan, Arjun and Neequaye, Nicole and Malur, Anagha and Soliman, Eman and McPeek, Matthew and Leffler, Nancy and Ogburn, David and Tokarz, Debra A. and Knudson, Warren and Gharib, Sina A. and et al.}, year={2020}, month={Sep} }
 @article{mcpeek_malur_tokarz_lertpiriyapong_gowdy_murray_wingard_fessler_barna_thomassen_2019, title={Alveolar Macrophage ABCG1 Deficiency Promotes Pulmonary Granulomatous Inflammation}, volume={61}, ISSN={["1535-4989"]}, DOI={10.1165/rcmb.2018-0365OC}, abstractNote={Pulmonary granuloma formation is a complex and poorly understood response to inhaled pathogens and particulate matter. To explore the mechanisms of pulmonary granuloma formation and maintenance, our laboratory has developed a multiwall carbon nanotube (MWCNT)-induced murine model of chronic granulomatous inflammation. We have demonstrated that the MWCNT model closely mimics pulmonary sarcoidosis pathophysiology, including the deficiency of alveolar macrophage ATP-binding cassette (ABC) lipid transporters ABCA1 and ABCG1. We hypothesized that deficiency of alveolar macrophage ABCA1 and ABCG1 would promote pulmonary granuloma formation and inflammation. To test this hypothesis, the effects of MWCNT instillation were evaluated in ABCA1, ABCG1, and ABCA1/ABCG1 myeloid-specific knockout (KO) mice. Histological examination revealed significantly larger pulmonary granulomas in ABCG1-KO and ABCA1/ABCG1 double-KO animals when compared with wild-type animals. Evaluation of BAL cells indicated increased expression of CCL2 and osteopontin, genes shown to be involved in the formation and maintenance of pulmonary granulomas. Single deficiency of alveolar macrophage ABCA1 did not affect MWCNT-induced granuloma formation or proinflammatory gene expression. These observations indicate that the deficiency of alveolar macrophage ABCG1 promotes pulmonary granulomatous inflammation and that this is augmented by additional deletion of ABCA1.}, number={3}, journal={AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR BIOLOGY}, author={McPeek, Matthew and Malur, Anagha and Tokarz, Debra A. and Lertpiriyapong, Kvin and Gowdy, Kymberly M. and Murray, Gina and Wingard, Christopher J. and Fessler, Michael B. and Barna, Barbara P. and Thomassen, Mary Jane}, year={2019}, month={Sep}, pages={332–340} }
 @article{schreeg_evans_allen_lewis_luckring_evola_richard_piner_thompson_adin_et al._2019, title={Cardiac Leiomyosarcoma in a Cat Presenting for Bilateral Renal Neoplasia}, volume={168}, ISSN={["1532-3129"]}, DOI={10.1016/j.jcpa.2019.02.005}, abstractNote={A 10-year-old neutered female domestic longhair cat was presented to a tertiary care veterinary hospital for evaluation of a right renal mass that was identified incidentally on abdominal radiographs and classified further as a sarcoma based on fine needle aspiration cytology. Further diagnostic workup, including ultrasound and cytology, identified a sarcoma in the left kidney. After approximately 1 month of conservative medical management, the clinical condition deteriorated and the cat was humanely destroyed. Post-mortem examination confirmed bilateral renal masses with multifocal infarction and extensive necrosis, and further identified a large mass at the apex of the heart as well as multiple pulmonary nodules. Microscopical examination of the masses identified a population of poorly-differentiated neoplastic spindle cells, consistent with sarcoma. Immunohistochemically, the neoplastic cells expressed smooth muscle actin and muscle-specific actin, but were negative for myoglobin and factor VIII. Phosphotungstic acid–haematoxylin staining was unable to identify cross-striations in the neoplastic cells. Based on these results and the pattern of lesion distribution, the cat was diagnosed with cardiac leiomyosarcoma with pulmonary and bilateral renal metastasis.}, journal={JOURNAL OF COMPARATIVE PATHOLOGY}, author={Schreeg, M. E. and Evans, B. J. and Allen, J. and Lewis, M. C. and Luckring, E. and Evola, M. and Richard, D. K. and Piner, K. and Thompson, E. M. and Adin, D. B. and et al.}, year={2019}, month={Apr}, pages={19–24} }
 @article{friedenberg_vansteenkiste_yost_treeful_meurs_tokarz_olby_2018, title={A de novo mutation in the EXT2 gene associated with osteochondromatosis in a litter of American Staffordshire Terriers}, volume={32}, ISSN={0891-6640}, url={http://dx.doi.org/10.1111/jvim.15073}, DOI={10.1111/jvim.15073}, abstractNote={Background We aimed to identify mutations associated with osteochondromatosis in a litter of American Staffordshire Terrier puppies. Hypothesis We hypothesized that the associated mutation would be located in a gene that causes osteochondromatosis in humans. Animals A litter of 9 American Staffordshire puppies, their sire and dam, 3 of 4 grandparents, 26 healthy unrelated American Staffordshire Terriers, and 154 dogs of 27 different breeds. Methods Whole genome sequencing was performed on the proband, and variants were compared against polymorphisms derived from 154 additional dogs across 27 breeds, as well as single nucleotide polymorphism database 146. One variant was selected for follow‐up sequencing. Parentage and genetic mosaicism were evaluated across the litter. Results We found 56,301 genetic variants unique to the proband. Eleven variants were located in or near the gene exostosin 2 (EXT2), which is strongly associated with osteochondromatosis in humans. One heterozygous variant (c.969C > A) is predicted to result in a stop codon in exon 5 of the gene. Sanger sequencing identified the identical mutation in all affected offspring. The mutation was absent in the unaffected offspring, both parents, all available grandparents, and 26 healthy unrelated American Staffordshire Terriers. Conclusions and Clinical Importance These findings represent the first reported mutation associated with osteochondromatosis in dogs. Because this mutation arose de novo, the identical mutation is unlikely to be the cause of osteochondromatosis in other dogs. However, de novo mutations in EXT2 are common in humans with osteochondromatosis, and by extension, it is possible that dogs with osteochondromatosis could be identified by sequencing the entire EXT2 gene.}, number={3}, journal={Journal of Veterinary Internal Medicine}, publisher={Wiley}, author={Friedenberg, Steven G. and Vansteenkiste, Daniella and Yost, Oriana and Treeful, Amy E. and Meurs, Kathryn M. and Tokarz, Debra A. and Olby, Natasha J.}, year={2018}, month={Feb}, pages={986–992} }
 @article{messenger_hall_jima_house_tam_tokarz_smart_2018, title={C/EBPβ deletion in oncogenic Ras skin tumors is a synthetic lethal event}, volume={9}, ISSN={2041-4889}, url={http://dx.doi.org/10.1038/S41419-018-1103-Y}, DOI={10.1038/s41419-018-1103-y}, abstractNote={Abstract Therapeutic targeting of specific genetic changes in cancer has proven to be an effective therapy and the concept of synthetic lethality has emerged. CCAAT/enhancer-binding protein-β (C/EBPβ), a basic leucine zipper transcription factor, has important roles in cellular processes including differentiation, inflammation, survival, and energy metabolism. Using a genetically engineered mouse model, we report that the deletion C/EBPβ in pre-existing oncogenic Ha-Ras mouse skin tumors in vivo resulted in rapid tumor regression. Regressing tumors exhibited elevated levels of apoptosis and p53 protein/activity, while adjacent C/EBPβ-deleted skin did not. These results indicate that the deletion of C/EBPβ de-represses p53 in oncogenic Ras tumors but not in normal wild-type Ras keratinocytes, and that C/EBPβ is essential for survival of oncogenic Ras tumors. Co-deletion of C/EBPβ and p53 in oncogenic Ras tumors showed p53 is required for tumor regression and elevated apoptosis. In tumors, loss of a pathway that confers adaptability to a stress phenotype of cancer/tumorigenesis, such as DNA damage, could result in selective tumor cell killing. Our results show that oncogenic Ras tumors display a significant DNA damage/replicative stress phenotype and these tumors have acquired a dependence on C/EBPβ for their survival. RNAseq data analysis of regressing tumors deleted of C/EBPβ indicates a novel interface between p53, type-1 interferon response, and death receptor pathways, which function in concert to produce activation of extrinsic apoptosis pathways. In summary, the deletion of C/EBPβ in oncogenic Ras skin tumors is a synthetic lethal event, making it a promising target for future potential anticancer therapies.}, number={11}, journal={Cell Death & Disease}, publisher={Springer Science and Business Media LLC}, author={Messenger, Zachary J. and Hall, Jonathan R. and Jima, Dereje D. and House, John S. and Tam, Hann W. and Tokarz, Debra A. and Smart, Robert C.}, year={2018}, month={Oct} }
 @article{mcpeek_malur_tokarz_murray_barna_thomassen_2018, title={PPAR-gamma pathways attenuate pulmonary granuloma formation in a carbon nanotube induced murine model of sarcoidosis}, volume={503}, ISSN={["1090-2104"]}, DOI={10.1016/j.bbrc.2018.06.061}, abstractNote={Peroxisome proliferator activated receptor gamma (PPARγ), a ligand activated nuclear transcription factor, is constitutively expressed in alveolar macrophages of healthy individuals. PPARγ deficiencies have been noted in several lung diseases including the alveolar macrophages of pulmonary sarcoidosis patients. We have previously described a murine model of multiwall carbon nanotubes (MWCNT) induced pulmonary granulomatous inflammation which bears striking similarities to pulmonary sarcoidosis, including the deficiency of alveolar macrophage PPARγ. Further studies demonstrate alveolar macrophage PPARγ deficiency exacerbates MWCNT-induced pulmonary granulomas. Based on these observations we hypothesized that activation of PPARγ via administration of the PPARγ-specific ligand rosiglitazone would limit MWCNT-induced granuloma formation and promote PPARγ-dependent pathways. Results presented here show that rosiglitazone significantly limits the frequency and severity of MWCNT-induced pulmonary granulomas. Furthermore, rosiglitazone attenuates alveolar macrophage NF-κB activity and downregulates the expression of the pro-inflammatory mediators, CCL2 and osteopontin. PPARγ activation via rosiglitazone also prevents the MWCNT-induced deficiency of PPARγ-regulated ATP-binding cassette lipid transporter-G1 (ABCG1) expression. ABCG1 is crucial to pulmonary lipid homeostasis. ABCG1 deficiency results in lipid accumulation which promotes pro-inflammatory macrophage activation. Our results indicate that restoration of homeostatic ABCG1 levels by rosiglitazone correlates with both reduced pulmonary lipid accumulation, and decreased alveolar macrophage activation. These data confirm and further support our previous observations that PPARγ pathways are critical in regulating MWCNT-induced pulmonary granulomatous inflammation.}, number={2}, journal={BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS}, author={McPeek, Matthew and Malur, Anagha and Tokarz, Debra A. and Murray, Gina and Barna, Barbara P. and Thomassen, Mary Jane}, year={2018}, month={Sep}, pages={684–690} }
 @article{duke_thompson_ihrie_taylor-just_ash_shipkowski_hall_tokarz_cesta_hubbs_et al._2018, title={Role of p53 in the chronic pulmonary immune response to tangled or rod-like multi-walled carbon nanotubes}, volume={12}, ISSN={["1743-5404"]}, DOI={10.1080/17435390.2018.1502830}, abstractNote={The fiber-like shape of multi-walled carbon nanotubes (MWCNTs) is reminiscent of asbestos, suggesting they pose similar health hazards when inhaled, including pulmonary fibrosis and mesothelioma. Mice deficient in the tumor suppressor p53 are susceptible to carcinogenesis. However, the chronic pathologic effect of MWCNTs delivered to the lungs of p53 heterozygous (p53+/-) mice has not been investigated. We hypothesized that p53+/- mice would be susceptible to lung tumor development after exposure to either tangled (t-) or rod-like (r-) MWCNTs. Wild-type (p53+/+) or p53+/- mice were exposed to MWCNTs (1 mg/kg) via oropharyngeal aspiration weekly over four consecutive weeks and evaluated for cellular and pathologic outcomes 11-months post-initial exposure. No lung or pleural tumors were observed in p53+/+ or p53+/- mice exposed to either t- or rMWCNTs. In comparison to tMWCNTs, the rMWCNTs induced the formation of larger granulomas, a greater number of lymphoid aggregates and greater epithelial cell hyperplasia in terminal bronchioles in both p53+/- and p53+/+ mice. A constitutively larger area of CD45R+/CD3+ lymphoid tissue was observed in p53+/- mice compared to p53+/+ mice. Importantly, p53+/- mice had larger granulomas induced by rMWCNTs as compared to p53+/+ mice. These findings indicate that a combination of p53 deficiency and physicochemical characteristics including nanotube geometry are factors in susceptibility to MWCNT-induced lymphoid infiltration and granuloma formation.}, number={9}, journal={NANOTOXICOLOGY}, author={Duke, Katherine S. and Thompson, Elizabeth A. and Ihrie, Mark D. and Taylor-Just, Alexia J. and Ash, Elizabeth A. and Shipkowski, Kelly A. and Hall, Jonathan R. and Tokarz, Debra A. and Cesta, Mark F. and Hubbs, Ann F. and et al.}, year={2018}, month={Oct}, pages={975–991} }
 @article{tokarz_heffelfinger_jima_gerlach_shah_rodriguez-nunez_kortum_fletcher_nordone_law_et al._2017, title={Disruption of Trim9 function abrogates macrophage motility in vivo}, volume={102}, ISSN={0741-5400 1938-3673}, url={http://dx.doi.org/10.1189/jlb.1A0816-371R}, DOI={10.1189/jlb.1a0816-371r}, abstractNote={Abstract The vertebrate immune response comprises multiple molecular and cellular components that interface to provide defense against pathogens. Because of the dynamic complexity of the immune system and its interdependent innate and adaptive functionality, an understanding of the whole-organism response to pathogen exposure remains unresolved. Zebrafish larvae provide a unique model for overcoming this obstacle, because larvae are protected against pathogens while lacking a functional adaptive immune system during the first few weeks of life. Zebrafish larvae were exposed to immune agonists for various lengths of time, and a microarray transcriptome analysis was executed. This strategy identified known immune response genes, as well as genes with unknown immune function, including the E3 ubiquitin ligase tripartite motif-9 (Trim9). Although trim9 expression was originally described as “brain specific,” its expression has been reported in stimulated human Mϕs. In this study, we found elevated levels of trim9 transcripts in vivo in zebrafish Mϕs after immune stimulation. Trim9 has been implicated in axonal migration, and we therefore investigated the impact of Trim9 disruption on Mϕ motility and found that Mϕ chemotaxis and cellular architecture are subsequently impaired in vivo. These results demonstrate that Trim9 mediates cellular movement and migration in Mϕs as well as neurons.}, number={6}, journal={Journal of Leukocyte Biology}, publisher={Wiley}, author={Tokarz, Debra A. and Heffelfinger, Amy K. and Jima, Dereje D. and Gerlach, Jamie and Shah, Radhika N. and Rodriguez-Nunez, Ivan and Kortum, Amanda N. and Fletcher, Ashley A. and Nordone, Shila K. and Law, J. McHugh and et al.}, year={2017}, month={Oct}, pages={1371–1380} }
 @article{chernyavskaya_mudbhary_zhang_tokarz_jacob_gopinath_sun_wang_magnani_madakashira_et al._2017, title={Loss of DNA methylation in zebrafish embryos activates retrotransposons to trigger antiviral signaling}, volume={144}, ISSN={0950-1991 1477-9129}, url={http://dx.doi.org/10.1242/dev.147629}, DOI={10.1242/dev.147629}, abstractNote={Complex cytoplasmic nucleotide-sensing mechanisms can recognize foreign DNA based on a lack of methylation and initiate an immune response to clear the infection. Zebrafish embryos with global DNA hypomethylation caused by mutations in the}, number={16}, journal={Development}, publisher={The Company of Biologists}, author={Chernyavskaya, Yelena and Mudbhary, Raksha and Zhang, Chi and Tokarz, Debra and Jacob, Vinitha and Gopinath, Smita and Sun, Xiaochen and Wang, Shuang and Magnani, Elena and Madakashira, Bhavani P. and et al.}, year={2017}, month={Jul}, pages={2925–2939} }
 @article{allen_gracieux_talib_tokarz_hensley_cores_vandergriff_tang_de andrade_dinh_et al._2016, title={Angiopellosis as an Alternative Mechanism of Cell Extravasation}, volume={35}, ISSN={1066-5099}, url={http://dx.doi.org/10.1002/stem.2451}, DOI={10.1002/stem.2451}, abstractNote={Stem cells possess the ability to home in and travel to damaged tissue when injected intravenously. For the cells to exert their therapeutic effect, they must cross the blood vessel wall and enter the surrounding tissues. The mechanism of extravasation injected stem cells employ for exit has yet to be characterized. Using intravital microscopy and a transgenic zebrafish line Tg(fli1a:egpf) with GFP-expressing vasculature, we documented the detailed extravasation processes in vivo for injected stem cells in comparison to white blood cells (WBCs). While WBCs left the blood vessels by the standard diapedesis process, injected cardiac and mesenchymal stem cells underwent a distinct method of extravasation that was markedly different from diapedesis. Here, the vascular wall undergoes an extensive remodeling to allow the cell to exit the lumen, while the injected cell remains distinctively passive in activity. We termed this process Angio-pello-sis, which represents an alternative mechanism of cell extravasation to the prevailing theory of diapedesis. Stem Cells 2017;35:170-180 Video Highlight: https://youtu.be/i5EI-ZvhBps.}, number={1}, journal={STEM CELLS}, publisher={Wiley}, author={Allen, Tyler A. and Gracieux, David and Talib, Maliha and Tokarz, Debra A. and Hensley, M. Taylor and Cores, Jhon and Vandergriff, Adam and Tang, Junnan and de Andrade, James B.M. and Dinh, Phuong-Uyen and et al.}, year={2016}, month={Jul}, pages={170–180} }
 @article{kol_christopher_skorupski_tokarz_vernau_2012, title={B-cell lymphoma with plasmacytoid differentiation, atypical cytoplasmic inclusions, and secondary leukemia in a dog}, volume={42}, ISSN={0275-6382}, url={http://dx.doi.org/10.1111/vcp.12003}, DOI={10.1111/vcp.12003}, abstractNote={Abstract A 7‐year‐old male castrated J ack R ussell T errier was presented to the oncology service at the U niversity of C alifornia– D avis V eterinary M edical T eaching H ospital for evaluation of suspected lymphoma. The dog had several enlarged lymph nodes and moderate lymphocytosis. Aspirates of an enlarged inguinal lymph node contained a bimorphic population of large immature lymphocytes and smaller cells with plasmacytoid features. Both cell types often contained a single large cytoplasmic inclusion that varied from clear to pale pink to sky blue. Cytologic changes were interpreted as most consistent with lymphoid neoplasia. Based on the predominantly mature cell morphology and some morphologic heterogeneity, the peripheral lymphocytosis was interpreted as most likely reactive in nature. However, the immunophenotype of the cells ( CD 20+, CD 21+, CD 79a+, MUM ‐1+, and MHCII +) and clonality assays showed that tissue and blood lymphocytes were neoplastic B cells with clonal identity despite their different morphologic appearances. The cytoplasmic inclusions were positive with periodic acid‐Schiff and were immunoreactive for IgM and IgG. By transmission electron microscopy, inclusions consisted of aberrant rough endoplasmic reticulum; a few small Russell bodies were also noted. A final diagnosis of high‐grade B‐cell lymphoma with plasmacytoid differentiation, atypical cytoplasmic inclusions, and secondary leukemia was made. Chemotherapy was initiated, but the dog was euthanized due to severe and uncontrolled seizures 9 months after the initial diagnosis. This case extends the morphologic repertoire of canine plasmacytoid neoplasms and emphasizes their continuum with multicentric lymphoma. This case also demonstrates the need for advanced diagnostic techniques in establishing blood involvement in lymphoma in some instances.}, number={1}, journal={Veterinary Clinical Pathology}, publisher={Wiley}, author={Kol, A. and Christopher, M.M. and Skorupski, K.A. and Tokarz, D. and Vernau, W.}, year={2012}, month={Dec}, pages={40–46} }
 @article{yamout_tokarz_magdesian_le jeune_2012, title={Intrathoracic pulsion diverticulum in a horse}, volume={53}, journal={Canadian Veterinary Journal}, author={Yamout, S. and Tokarz, D. and Magdesian, K.G. and le Jeune, S.S.}, year={2012}, pages={408–411} }
 @article{tokarz_poppenga_kaae_filigenzi_lowenstine_pesavento_2011, title={Amanitin Toxicosis in Two Cats with Acute Hepatic and Renal Failure}, volume={49}, ISSN={0300-9858 1544-2217}, url={http://dx.doi.org/10.1177/0300985811429307}, DOI={10.1177/0300985811429307}, abstractNote={Amanitin is a toxic cyclopeptide present in several species of poisonous mushrooms. Amanitin toxicosis was diagnosed in 2 cats from separate premises. Both cats initially had lethargy and vomiting, and they rapidly developed depression and neurological signs over 24-48 hours. Marked elevation of alanine aminotransferase was the primary finding, with subsequent serum chemistry values compatible with hepatic and renal failure. Histopathological findings consisted of submassive to massive acute hepatic necrosis, renal proximal tubular epithelial necrosis, and foci of necrosis and inflammation in the gastrointestinal tract. Amanitin exposure was confirmed postmortem by detection of α-amanitin in the kidney by liquid chromatography-mass spectrometry. A similar clinical course and pathological changes are reported in human and canine amanitin intoxication; however, gastrointestinal lesions are not typically described.}, number={6}, journal={Veterinary Pathology}, publisher={SAGE Publications}, author={Tokarz, D. and Poppenga, R. and Kaae, J. and Filigenzi, M. and Lowenstine, L. J. and Pesavento, P.}, year={2011}, month={Dec}, pages={1032–1035} }
 @article{sykes_marks_mapes_schultz_pollard_tokarz_pesavento_lindsay_foley_2010, title={Salmon Poisoning Disease in Dogs: 29 Cases}, volume={24}, ISSN={0891-6640 1939-1676}, url={http://dx.doi.org/10.1111/j.1939-1676.2010.0493.x}, DOI={10.1111/j.1939-1676.2010.0493.x}, abstractNote={Background: Salmon poisoning disease (SPD) is a trematode-borne disease of dogs caused by Neorickettsia helminthoeca. Objectives: To determine risk factors and spatial epidemiology of SPD in dogs from northern California; to describe the clinicopathologic, microbiologic, and imaging findings of SPD in these dogs; and to evaluate treatments and outcomes for SPD. Animals: Twenty-nine dogs with SPD based on the finding of trematode ova in the feces, or organisms consistent with N. helminthoeca in specimens submitted for microscopic examination. Methods: Information regarding signalment, fish exposure, clinical signs, diagnostic evaluation, treatments, and outcomes was obtained for each dog. Archived lymph node aspirates and histopathology specimens were subjected to polymerase chain reaction (PCR) testing for Neorickettsia spp. Results: Labrador Retrievers and intact male dogs were overrepresented. Exposure locations were often distant from the dogs' residence. Some dogs had neurologic signs, including twitching and seizures. Dogs lacking peripheral lymphadenomegaly had abdominal lymphadenomegaly on ultrasound examination. A combination of centrifugation fecal flotation and sedimentation had greatest sensitivity for finding fluke ova. N. helminthoeca DNA was amplified by PCR from 4/10 dogs. Penicillins, cephalosporins, and chloramphenicol did not appear to be effective treatments. Mortality rate was 4/29 (14%). Conclusions and Clinical Importance: SPD should be suspected in dogs with inappetence, gastrointestinal, or neurologic signs, with or without fever or peripheral lymphadenomegaly in the appropriate geographical setting. Diagnosis is facilitated by a combination of fecal sedimentation and centrifugal flotation, abdominal ultrasonography, and PCR-based assays on lymphoid tissue. The treatment of choice is tetracycline antimicrobials.}, number={3}, journal={Journal of Veterinary Internal Medicine}, publisher={Wiley}, author={Sykes, J.E. and Marks, S.L. and Mapes, S. and Schultz, R.M. and Pollard, R.E. and Tokarz, D. and Pesavento, P.P. and Lindsay, L.L. and Foley, J.E.}, year={2010}, month={Mar}, pages={504–513} }
 @article{rainier_bui_mark_thomas_tokarz_ming_delaney_richardson_albers_matsunami_et al._2008, title={Neuropathy Target Esterase Gene Mutations Cause Motor Neuron Disease}, volume={82}, ISSN={0002-9297}, url={http://dx.doi.org/10.1016/j.ajhg.2007.12.018}, DOI={10.1016/j.ajhg.2007.12.018}, abstractNote={The possibility that organophosphorus (OP) compounds contribute to motor neuron disease (MND) is supported by association of paraoxonase 1 polymorphisms with amyotrophic lateral sclerosis (ALS) and the occurrence of MND in OP compound-induced delayed neuropathy (OPIDN), in which neuropathy target esterase (NTE) is inhibited by organophosphorylation. We evaluated a consanguineous kindred and a genetically unrelated nonconsanguineous kindred in which affected subjects exhibited progressive spastic paraplegia and distal muscle wasting. Affected subjects resembled those with OPIDN and those with Troyer Syndrome due to SPG20/spartin gene mutation (excluded by genetic linkage and SPG20/spartin sequence analysis). Genome-wide analysis suggested linkage to a 22 cM homozygous locus (D19S565 to D19S884, maximum multipoint LOD score 3.28) on chromosome 19p13 to which NTE had been mapped (GenBank AJ004832 ). NTE was a candidate because of its role in OPIDN and the similarity of our patients to those with OPIDN. Affected subjects in the consanguineous kindred were homozygous for disease-specific NTE mutation c.3034A→G that disrupted an interspecies conserved residue (M1012V) in NTE's catalytic domain. Affected subjects in the nonconsanguineous family were compound heterozygotes: one allele had c.2669G→A mutation, which disrupts an interspecies conserved residue in NTE's catalytic domain (R890H), and the other allele had an insertion (c.2946_2947insCAGC) causing frameshift and protein truncation (p.S982fs1019). Disease-specific, nonconserved NTE mutations in unrelated MND patients indicates NTE's importance in maintaining axonal integrity, raises the possibility that NTE pathway disturbances contribute to other MNDs including ALS, and supports the role of NTE abnormalities in axonopathy produced by neuropathic OP compounds. The possibility that organophosphorus (OP) compounds contribute to motor neuron disease (MND) is supported by association of paraoxonase 1 polymorphisms with amyotrophic lateral sclerosis (ALS) and the occurrence of MND in OP compound-induced delayed neuropathy (OPIDN), in which neuropathy target esterase (NTE) is inhibited by organophosphorylation. We evaluated a consanguineous kindred and a genetically unrelated nonconsanguineous kindred in which affected subjects exhibited progressive spastic paraplegia and distal muscle wasting. Affected subjects resembled those with OPIDN and those with Troyer Syndrome due to SPG20/spartin gene mutation (excluded by genetic linkage and SPG20/spartin sequence analysis). Genome-wide analysis suggested linkage to a 22 cM homozygous locus (D19S565 to D19S884, maximum multipoint LOD score 3.28) on chromosome 19p13 to which NTE had been mapped (GenBank AJ004832 ). NTE was a candidate because of its role in OPIDN and the similarity of our patients to those with OPIDN. Affected subjects in the consanguineous kindred were homozygous for disease-specific NTE mutation c.3034A→G that disrupted an interspecies conserved residue (M1012V) in NTE's catalytic domain. Affected subjects in the nonconsanguineous family were compound heterozygotes: one allele had c.2669G→A mutation, which disrupts an interspecies conserved residue in NTE's catalytic domain (R890H), and the other allele had an insertion (c.2946_2947insCAGC) causing frameshift and protein truncation (p.S982fs1019). Disease-specific, nonconserved NTE mutations in unrelated MND patients indicates NTE's importance in maintaining axonal integrity, raises the possibility that NTE pathway disturbances contribute to other MNDs including ALS, and supports the role of NTE abnormalities in axonopathy produced by neuropathic OP compounds. Exposure to neurotoxic organophosphorous (OP) compounds of sufficient magnitude and duration causes neurodegeneration in humans, livestock, and laboratory animals.1Abou-Donia M.B. Organophosphorus ester-induced delayed neurotoxicity.Annu. Rev. Pharmacol. Toxicol. 1981; 21: 511-548Crossref PubMed Scopus (350) Google Scholar OP compound exposures occur in industrial or agricultural environments and as the consequences of terrorism, accidents, suicide attempts, or chemical warfare. Recent attention has focused on OP neurotoxicity after the use of Sarin in the 1995 Tokyo subway terrorist incident2Lotti M. Becker C.E. Aminoff M.J. Organophosphate polyneuropathy; pathogenesis and prevention.Neurology. 1984; 34: 658-662Crossref PubMed Google Scholar and because of concerns about persistent neurological effects after exposures to neurotoxic OP agents in the Gulf War. During the Prohibition era in the United States, consumption of Jamaica ginger extract (“Ginger Jake”) adulterated with triorthocresyl phosphate (TOCP) led to OP compound-induced delayed neuropathy (OPIDN) in an estimated 50,000 Americans.3Woolf A.D. Ginger Jake and the blues: A tragic song of poisoning.Vet. Hum. Toxicol. 1995; 37: 252-254PubMed Google Scholar, 4Morgan J.P. Penovich P. Jamaica ginger paralysis. Forty-seven-year follow-up.Arch. Neurol. 1978; 35: 530-532Crossref PubMed Scopus (97) Google Scholar, 5Morgan J.P. Tulloss T.C. The Jake Walk Blues. A toxicologic tragedy mirrored in American popular music.Ann. Intern. Med. 1976; 85: 804-808Crossref PubMed Scopus (31) Google Scholar Additional outbreaks of OPIDN causing paralysis of tens of thousands of individuals have occurred in Morocco, Fiji, and India due to consumption of cooking oil contaminated with lubricating oil (containing triorthocresyl phosphate).6Smith H.V. Spalding J.M. Outbreak of paralysis in Morocco due to ortho-cresyl phosphate poisoning.Lancet. 1959; 2: 1019-1021Abstract PubMed Scopus (106) Google Scholar, 7Taylor P. Anticholinesterase agents. Goodman and Gilman's Pharmacologic Basis of Therapeutics.Ninth Edition. McGraw Hill, New York1996Google Scholar, 8Sorokin M. Orthocresyl phosphate neuropathy: Report of an outbreak in Fiji.Med. J. Aust. 1969; 1: 506-508PubMed Google Scholar, 9Srivastava A.K. Das M. Khanna S.K. An outbreak of tricresyl phosphate poisoning in Calcutta, India.Food Chem. Toxicol. 1990; 28: 303-304Crossref PubMed Scopus (12) Google Scholar, 10Sarkar J.K. Outbreaks of paralystic disease in West Bengal due to tricresyl phosphate poisoning.J. Indian Med. Assoc. 1974; 63: 359-361PubMed Google Scholar, 11Mehta R.S. Dixit I.P. Khakharia S.J. Toxic neuropathy in Raipur due to triorthocresylphosphate (TOCP).J. Assoc. Physicians India. 1975; 23: 133-138PubMed Google Scholar Neurologic syndromes following OP toxicity are highly variable and depend on such factors as the specific OP compound, dose and duration of exposure, species, age, and various physiologic factors at the time of OP exposure. Whereas acute OP toxicity usually produces acute cholinergic crisis,7Taylor P. Anticholinesterase agents. Goodman and Gilman's Pharmacologic Basis of Therapeutics.Ninth Edition. McGraw Hill, New York1996Google Scholar acute cholinergic crises are often absent in OPIDN. In contrast, OPIDN often begins with sensory impairment, ataxia, weakness, muscle fasciculation, and hyporeflexia and may progress to complete flaccid paralysis followed by progressive spastic paraplegia.4Morgan J.P. Penovich P. Jamaica ginger paralysis. Forty-seven-year follow-up.Arch. Neurol. 1978; 35: 530-532Crossref PubMed Scopus (97) Google Scholar, 7Taylor P. Anticholinesterase agents. Goodman and Gilman's Pharmacologic Basis of Therapeutics.Ninth Edition. McGraw Hill, New York1996Google Scholar, 12Inoue N. Fujishiro K. Mori K. Matsuoka M. Triorthocresyl phosphate poisoning - A review of human cases.J. UOEH. 1988; 10: 433-442PubMed Google Scholar Neuropathologic analyses of OPIDN have shown distal degeneration of the longest central- and peripheral-nervous-system axons.1Abou-Donia M.B. Organophosphorus ester-induced delayed neurotoxicity.Annu. Rev. Pharmacol. Toxicol. 1981; 21: 511-548Crossref PubMed Scopus (350) Google Scholar, 12Inoue N. Fujishiro K. Mori K. Matsuoka M. Triorthocresyl phosphate poisoning - A review of human cases.J. UOEH. 1988; 10: 433-442PubMed Google Scholar Distal axon degeneration is also the primary neuropathologic feature of hereditary spastic paraplegia (HSP) (reviewed in13Fink J.K. Hereditary spastic paraplegia.in: Rimoin D. Connor J.M. Pyeritz R.E. Korf B.R. Emery and Rimoin's Principles and Practice of Medical Genetics. 5 ed. Churchill Livingstone Elsevier, Philadelphia2007: 2771-2801Google Scholar). Some complicated forms of HSP,13Fink J.K. Hereditary spastic paraplegia.in: Rimoin D. Connor J.M. Pyeritz R.E. Korf B.R. Emery and Rimoin's Principles and Practice of Medical Genetics. 5 ed. Churchill Livingstone Elsevier, Philadelphia2007: 2771-2801Google Scholar such as Troyer syndrome (MIM 275900), also exhibit lower-motor neuron involvement.14Patel H. Cross H. Proukakis C. Hershberger R. Bork P. Ciccarelli F.D. Patton M.A. McKusick V.A. Crosby A.H. SPG20 is mutated in Troyer syndrome, an hereditary spastic paraplegia.Nat. Genet. 2002; 31: 347-348Crossref PubMed Scopus (198) Google Scholar OPIDN pathogenesis involves neuropathy target esterase (NTE), a neuronal membrane protein, either through direct OP-induced inhibition of NTE or through generation of OP-NTE neurotoxic complexes (“aged NTE”). NTE's functions are incompletely understood. NTE is capable of hydrolyzing several intrinsic membrane lipids and thus may be a factor in determining the composition of neuronal membranes.15van Tienhoven M. Atkins J. Li Y. Glynn P. Human neuropathy target esterase catalyzes hydrolysis of membrane lipids.J. Biol. Chem. 2002; 277: 20942-20948Crossref PubMed Scopus (138) Google Scholar NTE's catalytic domain for esterase activity has been mapped to a 489 amino acid region between residues 727 and 1216.16Forshaw P.J. Atkins J. Ray D.E. Glynn P. The catalytic domain of human neuropathy target esterase mediates an organophosphate-sensitive ionic conductance across liposome membranes.J. Neurochem. 2001; 79: 400-406Crossref PubMed Scopus (12) Google Scholar Participation of NTE in a cell-signaling pathway controlling interactions between neurons and accessory glial cells in the developing nervous system has also been proposed.17Glynn P. Neuropathy target esterase.Biochem. J. 1999; 344: 625-631Crossref PubMed Scopus (107) Google Scholar, 18Glynn P. Neural development and neurodegeneration: Two faces of neuropathy target esterase.Prog. Neurobiol. 2000; 61: 61-74Crossref PubMed Scopus (107) Google Scholar We studied a consanguineous family of Ashkenazi Jewish ancestry (Figure 1A) and a nonconsanguineous family of northern European ancestry (Figure 1B) in which affected subjects developed childhood onset of insidiously progressive lower-extremity spastic weakness and progressive wasting of distal upper- and lower-extremity muscles. Electrophysiologic studies were consistent with a motor axonopathy affecting upper and lower extremities. Magnetic resonance imaging demonstrated spinal cord atrophy, particularly in the thoracic region. The affected phenotype in each family conformed both to OPIDN and to “Troyer syndrome,” an autosomal-recessive form of HSP associated with distal muscle wasting.14Patel H. Cross H. Proukakis C. Hershberger R. Bork P. Ciccarelli F.D. Patton M.A. McKusick V.A. Crosby A.H. SPG20 is mutated in Troyer syndrome, an hereditary spastic paraplegia.Nat. Genet. 2002; 31: 347-348Crossref PubMed Scopus (198) Google Scholar, 19Cross H.E. McKusick V.A. The Troyer syndrome. A recessive form of spastic paraplegia with distal muscle wasting.Arch. Neurol. 1967; 16: 473-485Crossref PubMed Scopus (93) Google Scholar Troyer syndrome patients variably exhibit additional neurologic and systemic abnormalities (delayed milestone acquisition, skeletal abnormalities, and cerebellar, extrapyramidal, and cognitive impairment). These features were not observed in our patients. The University of Michigan Institutional Review Board approved this study. We analyzed the SPG20/spartin gene (MIM 607111) coding sequence, mutations in which cause Troyer syndrome (SPG20 HSP),14Patel H. Cross H. Proukakis C. Hershberger R. Bork P. Ciccarelli F.D. Patton M.A. McKusick V.A. Crosby A.H. SPG20 is mutated in Troyer syndrome, an hereditary spastic paraplegia.Nat. Genet. 2002; 31: 347-348Crossref PubMed Scopus (198) Google Scholar, 19Cross H.E. McKusick V.A. The Troyer syndrome. A recessive form of spastic paraplegia with distal muscle wasting.Arch. Neurol. 1967; 16: 473-485Crossref PubMed Scopus (93) Google Scholar and found no mutations in the index (consanguineous) family (data not shown), and we excluded the SPG2014Patel H. Cross H. Proukakis C. Hershberger R. Bork P. Ciccarelli F.D. Patton M.A. McKusick V.A. Crosby A.H. SPG20 is mutated in Troyer syndrome, an hereditary spastic paraplegia.Nat. Genet. 2002; 31: 347-348Crossref PubMed Scopus (198) Google Scholar locus in the consanguineous family by genetic-linkage analysis (Table 1). Two-point linkage analyses for exclusion mapping were performed with the MLINK subroutine of the LINKAGE program20Lathrop G.M. Lalouel J. Julient C. Ott J. Multipoint linkage analysis in humans: Detection of linkage and estimation of recombination.Am. J. Hum. Genet. 1985; 37: 482-498PubMed Google Scholar with an autosomal-recessive model of disease inheritance and a disease allele frequency of 0.001. We assigned a genetic penetrance of 0.90 for LOD-score calculations.Table 1Genetic-Linkage Analysis Excludes Chromosome 13q12.3MarkerLOD θ = 00.010.020.030.040.05D13S1841−3.4−1.23−0.94−0.77−0.66−0.57D13S1842−2.19−1.19−0.92−0.76−0.64−0.56D13S1851−3.4−1.23−0.94−0.77−0.66−0.57D13S1843−3.4−1.23−0.94−0.77−0.66−0.57D13S2190.080.070.070.060.060.06D13S1844−2.19−1.19−0.92−0.76−0.64−0.56Genetic-linkage analysis excludes SPG20 “Troyer syndrome” locus on chromosome 13q12.3. Open table in a new tab Genetic-linkage analysis excludes SPG20 “Troyer syndrome” locus on chromosome 13q12.3. We then initiated genome-wide linkage analysis (Figure 2) in the larger, consanguineous family with ten subjects (three living affected subjects, seven living unaffected subjects, no spouses of descendants) and in the second smaller, nonconsanguineous family. Marrying-in spouses were asymptomatic and had no evidence of similar neurologic disorders in their families. Maximum-likelihood estimates of marker allele frequencies were estimated from all 15 subjects in both families. The average number of alleles was 4 (range = 3–7), and the average observed heterozygosity of these markers in the families was 0.64 (range = 0.32–0.84). We analyzed 400 polymorphic microsatellite markers spaced ∼10 cM apart (ABI MD-10 Linkage Mapping Set), of which 164 markers heterozygous for at least one affected individual excluded 41% of the genome. Six of these markers were homozygous for all affected individuals in the consanguineous family. We analyzed adjacent markers for each homozygous marker. Only markers adjacent to D19S209 yielded an extended linked haplotype that spanned 22 cM between D19S565 and D19S884 on chromosome 19p13 (Figure 1A). In the smaller, nonconsanguineous family, these same markers also resulted in haplotype sharing in affected subjects consistent with genetic linkage of this region (Figure 1B). The positive chromosome 19 region was analyzed with multipoint methods to incorporate information from all 27 informative markers. The GeneHunter program21Kruglyak L. Daly M.J. Reeve-Daly M.P. Lander E.S. Parametric and nonparametric linkage analysis: A unified multipoint approach.Am. J. Hum. Genet. 1996; 58: 1347-1363PubMed Google Scholar was used to obtain exact multipoint LOD scores and multipoint nonparametric linkage (NPL) scores. The NPL score is based on allele sharing identical by descent (IBD) and does not assume a transmission model. The NPL analysis allows an assessment of the robustness of the result to model misspecification. Because of the size and complexity of the largest pedigree, it had to be trimmed for analysis in GeneHunter. Therefore, analyses were also performed with SimWalk2.22Sobel E. Lange K. Descent graphs in pedigree analysis: Applications to haplotyping, location scores, and marker sharing statistics.Am. J. Hum. Genet. 1996; 58: 1323-1337PubMed Google Scholar, 23Sobel E. Sengul H. Weeks D.E. Multipoint estimation of identity-by-descent probablilities at arbitrary positions among marker loci on general pedigrees.Hum. Hered. 2001; 52: 121-131Crossref PubMed Scopus (145) Google Scholar, 24Sobel E. Papp J.C. Jange K. Detection and integration of genotyping errors in statistical genetics.Am. J. Hum. Genet. 2002; 70: 496-508Abstract Full Text Full Text PDF PubMed Scopus (282) Google Scholar SimWalk2 uses Markov chain Monte Carlo (MCMC) and simulated annealing methods to perform multipoint analyses, allowing computation on large, complex pedigrees. Although the method is not exact, all pedigree members could be included in the analysis. We performed parametric LOD score and NPL analyses in SimWalk2. For each analysis, we set the “parallel runs” flag to generate two simulated annealing runs of the data. The runs produced a maximum score at the same location with the same scores within rounding error. p values given by SimWalk2 were calculated with 10,000 simulations. For nonparametric analyses, p values were used to generate the reported scores (score = −log10 [p value]). Multipoint LOD score and nonparametric analyses of the two families clearly mirrored the shared haplotype region (Figure 2). With all methods (model-based parametric analysis or nonparametric analysis with either GeneHunter or SimWalk2) used, the maximum LOD score occurred near marker D19S869. Maximum multipoint parametric LOD scores were 2.58 with GeneHunter and 3.82 with SimWalk2. The maximum NPL scores were 3.30 with GeneHunter and 3.36 with SimWalk2. The p value associated with the GeneHunter statistics is 0.002; the simulation-based p value for the SimWalk2 statistics is 0.0004. The index family contributed most to these scores (3.28 and 3.09 for the parametric and NPL SimWalk2 analyses, respectively, and 2.07 and 3.07 for the GeneHunter parametric and NPL analyses, respectively). It is important to note that the second small, nonconsanguineous family supported this peak, with scores close to the maximum expected for an affected-sibling-pair family. To further characterize the region on chromosome 19 and to fully characterize all shared chromosomal regions, we carried out high-density SNP typing with the Affymetrix 250K NspI chips on all family members for whom DNA was available: 12 subjects in the consanguineous family (including all three affected individuals and two additional unaffected subjects ascertained after the initial microsatellite linkage analysis had been performed) and five subjects in the nonconsanguineous family. We analyzed the SNP data by looking for regions of autozygosity in the entire genome with the Find Autozygous Regions program in the GeneSpring GT analysis package (Agilent). The results of this analysis are shown in Figure 3; they clearly show a single major autozygosity region on chromosome 19 in the three affected individuals. This 4.7 megabase region is defined by the map location from 2784431 to 7541689 bp (NCBI build 35). This region corresponds to the same region identified by the microsatellite markers. The region includes the interesting candidate gene neuropathy target esterase (NTE) (GenBank AJ004832 ), whose location on chromosome 19 is from 7505075—7532647 bp (UCSC Genome Browser on NCBI build 35). The NTE gene was an obvious candidate because of its role in OPIDN and the similarity of symptoms seen in our patients to those reported for OPIDN. Analysis of NTE's coding sequence in the index family showed that each affected subject was homozygous for (and each obligate carrier heterozygous for) substitution of guanine for adenine at NTE cDNA 3034. This mutation was absent in 105 control subjects (data not shown) and caused substitution of valine for methionine at amino acid position 1012 (M1012V). This mutation disrupts an interspecies conserved residue within NTE's catalytic domain (Figure 4C).25Lush M.J. Li Y. Read D.J. Willis A.C. Glynn P. Neuropathy target esterase and a homologous Drosophila neurodegeneration-associated mutant protein contain a novel domain conserved from bacteria to man.Biochem. J. 1998; 332: 1-4Crossref PubMed Scopus (122) Google Scholar Mouse, Drosophila, and C. elegans species all contain M at residue 1012. Polyphen analysis of M1012V mutation gave a PSIC profile-difference score of 2.590 predicting that the mutation would be damaging. On the other hand, SIFT analysis predicted that M1012V substitution would be tolerated. We then analyzed the NTE coding sequence in the second, unrelated, nonconsanguineous family (Figure 1B). Analysis of NTE's coding sequence in affected subjects in this second family showed that they were compound heterozygotes for two NTE mutations. Like the mutation in the index family, these mutations also occurred within NTE's catalytic domain. One allele had a 2669G→A mutation corresponding to an R890H substitution in NTE's catalytic domain. Human and mouse contain R at residue 890, whereas Drosophila and C.elegans contain K at residue 890. Polyphen analysis (PSIC profile-difference score = 0.184) and SIFT analysis predicted that R890H would be a tolerated substitution. The other allele had a four base pair insertion (NTE mRNA position 2946) that caused a frameshift and protein truncation after residue 1019. The truncated protein is predicted to be missing the last 235 residues of NTE's catalytic domain (which extends from amino acid position 727 to position 1216). These mutations were present separately in each carrier parent and absent in 105 control subjects and the unaffected sibling. In summary, we identified homozygous and compound heterozygous NTE mutations in subjects from two unrelated families with autosomal-recessive progressive spastic paraplegia associated with distal upper- and lower-extremity wasting. Because of its clinical similarity with SPG20 HSP (Troyer syndrome), we reserved for this disease the next-available HSP locus designation (SPG39) and refer to this disorder as NTE-related Motor Neuron Disorder (NTE-MND). These NTE mutations are disease-specific and considered pathogenic for several reasons. First, they were present in affected subjects in two unrelated kindreds and absent in control subjects. Second, they affect amino acid residues within NTE's catalytic domain. Third, NTE plays a central role in OPIDN, an upper- and lower-motor neuron disorder whose symptoms bear a striking resemblance to those exhibited by our NTE-MND patients. Two mechanisms for NTE involvement in OPIDN have been proposed. The first mechanism proposes that neurotoxicity is the consequence of OP-induced inhibition of NTE. The second mechanism proposes that OP-NTE complexes are toxic. These proposed mechanisms are not mutually exclusive because OP interaction with NTE could both be toxic as well as interfere with NTE activity. The fact that identified NTE mutations in NTE-MND subjects disturb NTE's catalytic domain suggests that they could result in altered NTE activity in vivo. This suggests that modified NTE activity alone could be sufficient to cause corticospinal tract and peripheral motor axonopathy. Nonetheless, it is also possible that the NTE mutations identified result in neurotoxic or “aged” NTE. Observations that NTE mutations are associated with progressive upper- and lower-motor neuron disease indicate the importance of NTE in maintaining the integrity of corticospinal tract and peripheral motor axons. The finding that NTE mutations underlie corticospinal tract and peripheral motor axon degeneration raises the possibility that other NTE polymorphisms (or genetic variation in factors that regulate or interact with NTE) could contribute to other motor neuron disorders including amyotrophic lateral sclerosis (ALS) and primary lateral sclerosis (PLS). We have shown that two NTE mutations are the likely cause of autosomal-recessive motor neuron disease that closely resembles OPIDN. These NTE mutations were sufficient to cause the disorder even in the absence of apparent exposure to neurotoxic OP compounds. It will be important to determine whether other polymorphisms in NTE and/or proteins with which it interacts influence the susceptibility to OP-induced neurologic disease. Our findings, together with the recently identified association of PON1 polymorphisms with ALS,26Slowik A. Tomik B. Wolkow P.P. Partyka D. Turaj W. Malecki M.T. Pera J. Dziedzic T. Szczudlik A. Figlewicz D.A. Paraoxonase gene polymorphisms and sporadic ALS.Neurology. 2006; 67: 766-770Crossref PubMed Scopus (72) Google Scholar further support the possibility that neurotoxic OP compounds contribute to motor neuron disease. This research is supported by grants from the University of Michigan Institute of Gerontology (to S.R.); and the Veterans Affairs Merit Review, the National Institutes of Health (National Institute of Neurological Disorders and Stroke [NINDS] R01-NS053917 and R01-NS045163), the Spastic Paraplegia Foundation, and the National Organization for Rare Disorders (to J.K.F.). Those funding this study did not play any role in design and conduct of the study, in the collection, analysis, and interpretation of the data, or in the preparation, review, or approval of the manuscript. We are grateful for the Elderly Subjects Program of the University of Michigan Institute of Gerontology through which control subjects were ascertained, the technical assistance of Shalini Guduri, the expert secretarial assistance of Lynette Girbach, and the participation of research subjects and their families without whom this investigation would not be possible. Download .xls (.1 MB) Help with xls files Document S1. Table of Genescan Data The URL for online data listed herein is as follows:Online Mendelian Inheritance in Man, http://www.ncbi.nlm.nih.gov/Omim/.}, number={3}, journal={The American Journal of Human Genetics}, publisher={Elsevier BV}, author={Rainier, Shirley and Bui, Melanie and Mark, Erin and Thomas, Donald and Tokarz, Debra and Ming, Lei and Delaney, Colin and Richardson, Rudy J. and Albers, James W. and Matsunami, Nori and et al.}, year={2008}, month={Mar}, pages={780–785} }
 @article{schrenzel_maalouf_gaffney_tokarz_keener_mcclure_griffey_mcaloose_rideout_2005, title={Molecular characterization of isosporoid coccidia (Isospora and Atoxoplasma spp.) in passerine birds.}, volume={91}, ISSN={0022-3395 1937-2345}, url={http://dx.doi.org/10.1645/ge-3310}, DOI={10.1645/ge-3310}, abstractNote={Prevalence and disease caused by isosporoid coccidia in passerine birds are well recognized, but confusion about the life cycles of the parasites has led to taxonomic inconsistencies. In this study, we characterized segments of the chromosomal small and large-subunit ribosomal RNA (rRNA) genes of coccidial parasites from 23 species of passerine birds, as well as heat shock protein 70, apicoplast rRNA, and chromosomal 5.8s rRNA genes from a subgroup of these animals, and we correlated genetic data with morphologic findings for different parasite developmental stages, host phylogeny, and overall taxonomic relations within the phylum Apicomplexa. Our findings indicate that isosporoid coccidia of passerine birds are monophyletic but exhibit substantial diversity, with most avian species having one or several unique parasite lineages that underwent synchronous speciation with their hosts, interrupted by sporadic episodes of lateral transmission across species and families. Molecular analyses support a homoxenous life cycle, with sexual forms occurring chiefly in the intestines and asexual merozoites present systemically. Rarely, extraintestinal sexual stages can occur. The passerine coccidia are genetically most closely related to species of Eimeria rather than Isospora. We suggest that these parasites, whether identified from blood merozoite stages or fecal oocysts, be provisionally grouped as a homogeneous clade of individual species in a single taxon and formally named when reliable criteria allowing reclassification of related genera in the suborder Eimeriina are clarified.}, number={3}, journal={Journal of Parasitology}, publisher={American Society of Parasitologists}, author={Schrenzel, Mark D. and Maalouf, Gabriel A. and Gaffney, Patricia M. and Tokarz, Debra and Keener, Laura L. and McClure, Diane and Griffey, Stephen and McAloose, D. and Rideout, Bruce A.}, year={2005}, month={Jun}, pages={635–647} }
 @article{rainier_thomas_tokarz_ming_bui_plein_zhao_lemons_albin_delaney_et al._2004, title={Myofibrillogenesis Regulator 1 Gene Mutations Cause Paroxysmal Dystonic Choreoathetosis}, volume={61}, ISSN={0003-9942}, url={http://dx.doi.org/10.1001/archneur.61.7.1025}, DOI={10.1001/archneur.61.7.1025}, abstractNote={Paroxysmal dystonic choreoathetosis (PDC) is characterized by attacks of involuntary movements that occur spontaneously while at rest and following caffeine or alcohol consumption. Previously, we and others identified a locus for autosomal dominant PDC on chromosome 2q33-2q35.To identify the PDC gene.Analysis of PDC positional candidate genes by exon sequencing and reverse transcription-polymerase chain reaction.Outpatient clinical and molecular genetic laboratory at a university hospital. Patients Affected (n = 12) and unaffected (n = 26) subjects from 2 unrelated families with PDC and 105 unrelated control subjects.We identified missense mutations in the myofibrillogenesis regulator gene (MR-1) in affected subjects in 2 unrelated PDC kindreds. These mutations were absent in control subjects and caused substitutions of valine for alanine at amino acid positions 7 and 9. The substitutions disturb interspecies conserved residues and are predicted to alter the MR-1 gene's amino-terminal alpha helix. The MR-1 exon containing these mutations (exon 1) was expressed only in the brain, a finding that explains the brain-specific symptoms of subjects with these mutations.Although MR-1 gene function is unknown, the precedence of ion channel disturbance in other episodic neurologic disorders suggests that the pathophysiologic features of PDC also involve abnormal ion localization. The discovery that MR-1 mutations underlie PDC provides opportunities to explore this condition's pathophysiologic characteristics and may provide insight into the causes of other paroxysmal neurologic disorders as well as the neurophysiologic mechanisms of alcohol and caffeine, which frequently precipitate PDC attacks.}, number={7}, journal={Archives of Neurology}, publisher={American Medical Association (AMA)}, author={Rainier, S. and Thomas, D. and Tokarz, D. and Ming, L. and Bui, M. and Plein, E. and Zhao, X. and Lemons, R. and Albin, R. and Delaney, C. and et al.}, year={2004}, month={Jul}, pages={1025–1029} }
 @article{rainier_chai_tokarz_nicholls_fink_2003, title={NIPA1 Gene Mutations Cause Autosomal Dominant Hereditary Spastic Paraplegia (SPG6)}, volume={73}, ISSN={0002-9297}, url={http://dx.doi.org/10.1086/378817}, DOI={10.1086/378817}, abstractNote={The hereditary spastic paraplegias (HSPs) are genetically heterogeneous disorders characterized by progressive lower-extremity weakness and spasticity. The molecular pathogenesis is poorly understood. We report discovery of a dominant negative mutation in the NIPA1 gene in a kindred with autosomal dominant HSP (ADHSP), linked to chromosome 15q11-q13 (SPG6 locus); and precisely the same mutation in an unrelated kindred with ADHSP that was too small for meaningful linkage analysis. NIPA1 is highly expressed in neuronal tissues and encodes a putative membrane transporter or receptor. Identification of the NIPA1 function and ligand will aid an understanding of axonal neurodegeneration in HSP and may have important therapeutic implications. The hereditary spastic paraplegias (HSPs) are genetically heterogeneous disorders characterized by progressive lower-extremity weakness and spasticity. The molecular pathogenesis is poorly understood. We report discovery of a dominant negative mutation in the NIPA1 gene in a kindred with autosomal dominant HSP (ADHSP), linked to chromosome 15q11-q13 (SPG6 locus); and precisely the same mutation in an unrelated kindred with ADHSP that was too small for meaningful linkage analysis. NIPA1 is highly expressed in neuronal tissues and encodes a putative membrane transporter or receptor. Identification of the NIPA1 function and ligand will aid an understanding of axonal neurodegeneration in HSP and may have important therapeutic implications. Ten loci for autosomal dominant hereditary spastic paraplegia (ADHSP) have been mapped, and four ADHSP genes have been identified: SPG4/spastin, SPG3A/atlastin, SPG13/chaperonin 60, and SPG10/KIF5A (Hazan et al. Hazan et al., 1999Hazan J Fonknechten N Mavel D Paternotte C Samson D Artiguenave F Davoine C-S Cruaud C Durr A Wincker P Brottier P Cattolico L Barbe V Burgunder JM Prudhomme JF Brice A Fontaine B Heilig B Weissenbach J Spastin, a new AAA protein, is altered in the most frequent form of autosomal dominant spastic paraplegia.Nat Genet. 1999; 23: 296-303Crossref PubMed Scopus (528) Google Scholar; Zhao et al. Zhao et al., 2001Zhao X Alvarado D Rainier S Lemons R Hedera P Weber C Tukel T Apak M Heiman-Patterson T Ming L Bui M Fink JK Mutations in a novel GTPase cause autosomal dominant hereditary spastic paraplegia.Nat Genet. 2001; 29: 326-331Crossref PubMed Scopus (294) Google Scholar; Hansen et al. Hansen et al., 2002Hansen JJ Durr A Cournu-Rebeix I Georgopoulous C Ang D Nielsen MN Davoine C-S Brice A Fontaine B Gregersen N Bross P Hereditary spastic paraplegia SPG13 is associated with a mutation in the gene encoding the mitochondrial chaperonin Hsp60.Am J Hum Genet. 2002; 70: 1328-1332Abstract Full Text Full Text PDF PubMed Scopus (294) Google Scholar; Reid et al. Reid et al., 2002Reid E Kloos M Ashley-Koch A Hughes L Bevan S Svenson IK Graham FL Gaskell PC Dearlove A Pericak-Vance MA Rubinsztein DC Marchuk DA A kinesin heavy chain (KIF5A) mutation in hereditary spastic paraplegia (SPG10).Am J Hum Genet. 2002; 71: 1189-1194Abstract Full Text Full Text PDF PubMed Scopus (418) Google Scholar). (For reviews of hereditary spastic paraplegia [HSP] [“Strümpell-Lorrain syndrome,” MIM 182600], see Fink and Hedera Fink and Hedera, 1999Fink JK Hedera P Hereditary spastic paraplegia: genetic heterogeneity and genotype-phenotype correlation.Semin Neurol. 1999; 19: 301-310Crossref PubMed Scopus (54) Google Scholar; Fink Fink, 2001Fink JK Hereditary spastic paraplegia.in: Rimoin DPRCJ Emery KB Emery & Rimoin's principles and practice of medical genetics. Harcourt Publishers Limited UK, London2001: 3124-3145Google Scholar, Fink, 2002Fink JK Hereditary spastic paraplegia: the pace quickens.Ann Neurol. 2002; 51: 669-672Crossref PubMed Scopus (31) Google Scholar; Reid Reid, 2003Reid E Science in motion: common molecular pathological themes emerge in the hereditary spastic paraplegias.J Med Genet. 2003; 40: 81-86Crossref PubMed Scopus (135) Google Scholar.) Despite these advances, the molecular pathophysiology of the ADHSPs is largely unknown. Elsewhere, we identified a locus for uncomplicated ADHSP in chromosome 15q (SPG6) (Fink et al. Fink et al., 1995aFink JK Sharp G Lange B Wu C-TB Haley T Otterud B Peacock M Leppert M Autosomal dominant hereditary spastic paraparesis, type I: clinical and genetic analysis of a large North American family.Neurology. 1995a; 45: 325-331Crossref PubMed Scopus (31) Google Scholar;Fink et al., 1995bFink JK Wu C-TB Jones SM Sharp GB Lange BM Lesicki A Reinglass T Varvil T Otterud B Leppert M Autosomal dominant familial spastic paraplegia: tight linkage to chromosome 15q.Am J Hum Genet. 1995b; 56: 188-192Crossref PubMed Scopus (36) Google Scholar). We now report identification of disease-specific mutations in a novel gene, NIPA1 (fig. 1a) in the original SPG6-linked kindred (Fink et al. Fink et al., 1995aFink JK Sharp G Lange B Wu C-TB Haley T Otterud B Peacock M Leppert M Autosomal dominant hereditary spastic paraparesis, type I: clinical and genetic analysis of a large North American family.Neurology. 1995a; 45: 325-331Crossref PubMed Scopus (31) Google Scholar) and in an unrelated kindred with ADHSP (fig. 1b). The SPG6 locus extends 6.1 cM between D15S128 and the centromere (Rainier et al. Rainier et al., 2000Rainier S Bui M Jones SM Fink JK Chromosome 15q linked autosomal dominant hereditary spastic paraplegia: new mapping information and candidate gene analysis.Am J Hum Genet. 2000; : 391Google Scholar) (fig. 2a), an interval often involved in deletions that result in Prader-Willi syndrome (PWS) or Angelman syndrome (AS). PWS and AS are characterized by genetic imprinting, a phenomenon in which gene expression and the phenotype depend on the sex of the transmitting parent (Nicholls and Knepper Nicholls and Knepper, 2001Nicholls RD Knepper JL Genome organization, function, and imprinting in Prader-Willi and Angelman syndromes.Ann Rev Genomics Hum Genet. 2001; 2: 153-175Crossref PubMed Scopus (506) Google Scholar). We analyzed a large kindred—ADHSP-ARK1 (fig. 1b), in which ADHSP was linked to the SPG6 locus—and found no evidence of genetic imprinting (Fink et al. Fink et al., 1995aFink JK Sharp G Lange B Wu C-TB Haley T Otterud B Peacock M Leppert M Autosomal dominant hereditary spastic paraparesis, type I: clinical and genetic analysis of a large North American family.Neurology. 1995a; 45: 325-331Crossref PubMed Scopus (31) Google Scholar). Therefore, we analyzed as SPG6 candidates the four unique, nonimprinted, and highly evolutionarily conserved genes mapped proximal to the imprinted domain and within the pericentromeric region of chromosome 15q (Chai et al. Chai et al., 2003Chai J-H Locke DP Greally JM Knoll JHM Ohta T Dunai J Yavor A Eichler EE Nicholls RD Identification of four highly conserved genes between breakpoint hotspots BP1 and BP2 of the Prader-Willi/Angelman syndromes deletion region that have undergone evolutionary transposition mediated by flanking duplicons.Am J Hum Genet. 2003; 73 (in this issue): 898-925Abstract Full Text Full Text PDF PubMed Scopus (162) Google Scholar [in this issue]). These candidate genes included nonimprinted in Prader-Willi/Angelman loci 1 (NIPA1) (NCBI accession number BK001020) and 2 (NIPA2) (NCBI accession number BK001120) (Chai et al. Chai et al., 2003Chai J-H Locke DP Greally JM Knoll JHM Ohta T Dunai J Yavor A Eichler EE Nicholls RD Identification of four highly conserved genes between breakpoint hotspots BP1 and BP2 of the Prader-Willi/Angelman syndromes deletion region that have undergone evolutionary transposition mediated by flanking duplicons.Am J Hum Genet. 2003; 73 (in this issue): 898-925Abstract Full Text Full Text PDF PubMed Scopus (162) Google Scholar [in this issue]), GCP5 (NCBI accession number AF272884) (Murphy et al. Murphy et al., 2001Murphy SM Preble AM Patel UK O’Connell KL Dias DP Moritz M Agard D Stults JT Stearns T GCP5 and GCP6: two new members of the human gamma-tubulin complex.Mol Biol Cell. 2001; 12: 3340-3352Crossref PubMed Scopus (145) Google Scholar), and CYFIP1 (NCBI accession number NM_014608) (Koybayashi et al. Koybayashi et al., 1998Koybayashi K Kuroda S Fukata M p140Sra-1 (specifically Rac1-associated protein) is a novel specific target for Rac1 small GTPase.J Biol Chem. 1998; 273: 291-295Crossref PubMed Scopus (179) Google Scholar; Schenk et al. Schenk et al., 2001Schenk A Bardoni B Moro A A highly conserved protein family interacting with the fragile X mental retardation protein (FMRP) and displaying selective interactions with FMRP-related proteins FXR1P and FXR2P.Proc Natl Acad Sci USA. 2001; 98: 8844-8849Crossref PubMed Scopus (276) Google Scholar). We identified a nucleotide substitution at position 159 of the NIPA1 cDNA (159C→G; fig. 1a), which resulted in an amino acid substitution at position 45 of the NIPA1 protein (T45R) in each affected subject (n=28) in ADHSP-ARK1 (fig. 1b). In contrast, each unaffected subject (n=17) had only C at this position (fig. 1b), which agrees with the known human genomic sequence (NCBI accession number NT_024668). We examined 105 control subjects (ascertained through the Elderly Subjects Program of the University of Michigan Institute of Gerontology). Each control subject had only C at position 159 of the NIPA1 cDNA (data not shown). We analyzed the coding sequence of the other three nonimprinted genes (GCP5, CYFIP1, and NIPA2) in two affected members of ADHSP-ARK1 and identified no disease-specific mutations (data not shown). We then analyzed the NIPA1 coding sequence in affected probands from 62 kindreds with ADHSP, 6 kindreds with probable autosomal recessive HSP, and 13 subjects with all the signs and symptoms of HSP but no family history (“apparently sporadic” spastic paraplegia). Affected subjects in one unrelated kindred (ADHSP-IRQ1) (fig. 1b) had precisely the same NIPA1 mutation (159C→G) (fig. 1b) as affected subjects in the ADHSP-ARK1 kindred. Unaffected subjects from ADHSP-IRQ1 kindred showed only the normal nucleotide (159C). Whereas the ADHSP-ARK1 kindred was linked to the SPG6 locus (Fink et al. Fink et al., 1995bFink JK Wu C-TB Jones SM Sharp GB Lange BM Lesicki A Reinglass T Varvil T Otterud B Leppert M Autosomal dominant familial spastic paraplegia: tight linkage to chromosome 15q.Am J Hum Genet. 1995b; 56: 188-192Crossref PubMed Scopus (36) Google Scholar), the ADHSP-IRQ1 kindred was too small for meaningful linkage analysis. Clinical features of the ADHSP-ARK1 affected individuals are typical of uncomplicated HSP of late-teenage to early-adult symptom onset (Fink et al. Fink et al., 1995aFink JK Sharp G Lange B Wu C-TB Haley T Otterud B Peacock M Leppert M Autosomal dominant hereditary spastic paraparesis, type I: clinical and genetic analysis of a large North American family.Neurology. 1995a; 45: 325-331Crossref PubMed Scopus (31) Google Scholar). Clinical features of ADHSP-IRQ1 were similar: onset of insidiously progressive spastic weakness in both legs that began in late teen-age years and was associated with urinary urgency and mild vibratory sensation impairment in the toes. Kindreds ADHSP-ARK1 and ADHSP-IRQ1 are of Irish and Iraqi ancestry, respectively. Analysis of haplotypes for polymorphic markers linked to this locus (D15S541, D15S542, D15S646, D15S817, and D15S1021) showed no evidence of haplotype sharing between ADHSP-ARK1 and ADHSP-IRQ1 kindreds (data not shown). This indicates that these two kindreds with ADHSP are not closely related and suggests that the same NIPA1 mutation arose independently in each of them. Disease-specific NIPA1 mutations in ADHSP-ARK1 and ADHSP-IRQ1 occur in NIPA1 exon 1 and are predicted to change threonine to arginine at amino acid position 45 (T45R) (fig. 2b). This amino acid is conserved in mouse, chicken, and fish (zebrafish and Fugu) (Chai et al. Chai et al., 2003Chai J-H Locke DP Greally JM Knoll JHM Ohta T Dunai J Yavor A Eichler EE Nicholls RD Identification of four highly conserved genes between breakpoint hotspots BP1 and BP2 of the Prader-Willi/Angelman syndromes deletion region that have undergone evolutionary transposition mediated by flanking duplicons.Am J Hum Genet. 2003; 73 (in this issue): 898-925Abstract Full Text Full Text PDF PubMed Scopus (162) Google Scholar [in this issue]; R.D.N., unpublished data) and is predicted to occur at the end of the first of nine transmembrane domains in the NIPA1 polypeptide (fig. 2b). NIPA1 does not contain an AAA domain (like that present in spastin [Hazan et al. Hazan et al., 1999Hazan J Fonknechten N Mavel D Paternotte C Samson D Artiguenave F Davoine C-S Cruaud C Durr A Wincker P Brottier P Cattolico L Barbe V Burgunder JM Prudhomme JF Brice A Fontaine B Heilig B Weissenbach J Spastin, a new AAA protein, is altered in the most frequent form of autosomal dominant spastic paraplegia.Nat Genet. 1999; 23: 296-303Crossref PubMed Scopus (528) Google Scholar]) or a GTPase domain (like that present in atlastin [Zhao et al. Zhao et al., 2001Zhao X Alvarado D Rainier S Lemons R Hedera P Weber C Tukel T Apak M Heiman-Patterson T Ming L Bui M Fink JK Mutations in a novel GTPase cause autosomal dominant hereditary spastic paraplegia.Nat Genet. 2001; 29: 326-331Crossref PubMed Scopus (294) Google Scholar]) and does not bear other homology to genes that cause other forms of HSP. Although the NIPA1 function is not known, the clear prediction based on hydrophobicity analysis of an integral membrane-associated protein (Chai et al. Chai et al., 2003Chai J-H Locke DP Greally JM Knoll JHM Ohta T Dunai J Yavor A Eichler EE Nicholls RD Identification of four highly conserved genes between breakpoint hotspots BP1 and BP2 of the Prader-Willi/Angelman syndromes deletion region that have undergone evolutionary transposition mediated by flanking duplicons.Am J Hum Genet. 2003; 73 (in this issue): 898-925Abstract Full Text Full Text PDF PubMed Scopus (162) Google Scholar [in this issue]) suggests that NIPA1 functions as a receptor or transporter. Many individuals with PWS or AS have chromosome 15q class I deletions that include NIPA1 (fig. 2a) (Chai et al. Chai et al., 2003Chai J-H Locke DP Greally JM Knoll JHM Ohta T Dunai J Yavor A Eichler EE Nicholls RD Identification of four highly conserved genes between breakpoint hotspots BP1 and BP2 of the Prader-Willi/Angelman syndromes deletion region that have undergone evolutionary transposition mediated by flanking duplicons.Am J Hum Genet. 2003; 73 (in this issue): 898-925Abstract Full Text Full Text PDF PubMed Scopus (162) Google Scholar [in this issue]). The fact that such individuals do not exhibit progressive spastic paraplegia indicates that NIPA1 haploinsufficiency does not cause progressive spastic paraplegia. Therefore, we suggest that the NIPA1 T45R missense mutation identified in these kindreds with ADHSP is pathogenic through a dominant negative, gain-of-function mechanism. NIPA1 mRNA is expressed constitutively at low levels with 2.2-kb and 7.5-kb transcripts in all human tissues but shows significant enrichment in the brain (fig. 2c). The latter expression pattern is found throughout the CNS, whereas spinal cord shows equal expression of the two NIPA1 mRNAs (fig. 2c). The alternative mRNA isoforms arise from alternative polyadenylation within NIPA1 exon 5, and equivalent expression patterns are found for mouse (Chai et al. Chai et al., 2003Chai J-H Locke DP Greally JM Knoll JHM Ohta T Dunai J Yavor A Eichler EE Nicholls RD Identification of four highly conserved genes between breakpoint hotspots BP1 and BP2 of the Prader-Willi/Angelman syndromes deletion region that have undergone evolutionary transposition mediated by flanking duplicons.Am J Hum Genet. 2003; 73 (in this issue): 898-925Abstract Full Text Full Text PDF PubMed Scopus (162) Google Scholar [in this issue]). Our observations of the same NIPA1 gene mutation (159 C→G; T45R) in two unrelated kindreds with ADHSP that disrupts an interspecies conserved amino acid and was absent in control subjects (n=105) strongly support the pathogenic significance of this mutation. Discovery of NIPA1 mutations as the cause of SPG6-linked HSP improves the ability to diagnose HSP and to provide genetic counseling. The presence of a dominant negative NIPA1 mutation in a putative membrane receptor or transporter suggests that SPG6 arises from either altered signal transduction or small-molecule transport through a membrane. We suggest that this property makes NIPA1 and its unknown ligand an attractive target for therapeutic intervention in SPG6 and, perhaps, in other spastic paraplegias. Identification of the NIPA1 cellular and subcellular localization, function, and ligand will therefore aid an understanding of axonal neurodegeneration in HSP and may have important therapeutic implications. This research is supported by grants from the Veterans Affairs Merit Review (to J.K.F.), the Muscular Dystrophy Association (to R.D.N.), and the National Institutes of Health (NINDS R01NS33645, R01NS36177, and R01NS38713 to J.K.F. and HD31491 and ES10631 to R.D.N.). We gratefully acknowledge Dr. Gregory B. Sharp and Dr. Bernadette M. Lange (University of Arkansas) for ascertaining and evaluating ADHSP-ARK; the technical assistance of Colin Delaney and Donald Thomas; the expert secretarial assistance of Lynette Girbach; and the participation of subjects with HSP and their families, without whom our investigations of HSP would not be possible.}, number={4}, journal={The American Journal of Human Genetics}, publisher={Elsevier BV}, author={Rainier, Shirley and Chai, Jing-Hua and Tokarz, Debra and Nicholls, Robert D. and Fink, John K.}, year={2003}, month={Oct}, pages={967–971} }