Search

Spine disorders are the leading cause of disability worldwide. To promote social inclusion, it is essential to ensure that people can participate in their societies by improving their ability, opportunities, and dignity, through access to high-quality, evidence-based, and affordable spine services for all.To achieve this goal, SPINE20 recommends six actions.- SPINE20 recommends that G20 countries deliver evidence-based education to the community health workers and primary care clinicians to promote best practice for spine health, especially in underserved communities.- SPINE20 recommends that G20 countries deliver evidence-based, high-quality, cost-effective spine care interventions that are accessible, affordable and beneficial to patients.- SPINE20 recommends that G20 countries invest in Health Policy and System Research (HPSR) to generate evidence to develop and implement policies aimed at integrating rehabilitation in primary care to improve spine health.- SPINE20 recommends that G20 countries support ongoing research initiatives on digital technologies including artificial intelligence, regulate digital technologies, and promote evidence-based, ethical digital solutions in all aspects of spine care, to enrich patient care with high value and quality.- SPINE20 recommends that G20 countries prioritize social inclusion by promoting equitable access to comprehensive spine care through collaborations with healthcare providers, policymakers, and community organizations.- SPINE20 recommends that G20 countries prioritize spine health to improve the well-being and productivity of their populations. Government health systems are expected to create a healthier, more productive, and equitable society for all through collaborative efforts and sustained investment in evidence-based care and promotion of spine health., Competing Interests: Declaration of Conflicting InterestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Objective: Conventional total knee arthroplasty (TKA) rehabilitation has little impact on sedentary lifestyles that have negative long-term health consequences. The purpose of this trial was to determine the effect of telehealth-based physical activity behavior change intervention on physical activity and functional outcomes following TKA., Methods: This study was a 2-arm, parallel randomized controlled superiority trial at a regional Veterans Affairs medical center. The participants were 92 US military veterans (mean age = 65.7 [SD =7.8] y) undergoing unilateral TKA. The Physical Activity Behavior Change (PABC) intervention included telehealth-based self-management training (10 30-minute sessions) delivered over 12 weeks. The control intervention included telehealth-based health education sessions that matched PABC frequency and duration. Both groups participated in standardized conventional outpatient rehabilitation. Physical activity, measured as average daily step count, was the primary outcome. Secondary outcomes were Life-Space Assessment questionnaire, 30-Second Chair-Stand test, Timed "Up & Go" Test, 6-Minute Walk Test, Western Ontario and McMaster Universities Osteoarthritis Index, and Veterans RAND 12-Item Health Survey. The Self Efficacy for Exercise scale and daily time spent in different postures (sitting/lying, standing, stepping) were exploratory variables. Outcomes were measured at baseline (before surgery), mid-intervention (8 weeks after surgery), after the intervention (14 weeks after surgery; primary endpoint), and follow-up (38 weeks after surgery)., Results: The PABC group had an estimated 931 (95% CI = 42-1819) more daily steps than the control group from baseline to 14 weeks, though a between-group effect was not sustained at 38 weeks. There were no group differences in secondary outcomes. Participants included only military veterans using Veterans Health Administration services. The intervention targeted self-management and did not include peer support., Conclusion: The PABC intervention improved physical activity for veterans recovering from unilateral TKA at 14 weeks after surgery, though the effect was not sustained at 38 weeks. Physical function improved with rehabilitation but was not different between groups, indicating that physical function was not a primary driver of physical activity behavior., Impact: Conventional TKA rehabilitation has a negligible effect on free-living physical activity, which is relevant to long-term health outcomes. This trial identified telehealth physical activity self-management as effective in addressing activity behaviors, separate from conventional rehabilitation strategies., (© The Author(s) 2024. Published by Oxford University Press on behalf of the American Physical Therapy Association. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Translational control mechanisms modulate microbial latency of eukaryotic pathogens, enabling them to evade immunity and drug treatments. The protozoan parasite Toxoplasma gondii persists in hosts by differentiating from proliferative tachyzoites to latent bradyzoites, which are housed inside tissue cysts. Transcriptional changes facilitating bradyzoite conversion are mediated by a Myb domain transcription factor called BFD1, whose mRNA is present in tachyzoites but not translated into protein until stress is applied to induce differentiation. We addressed the mechanisms by which translational control drives BFD1 synthesis in response to stress-induced parasite differentiation. Using biochemical and molecular approaches, we show that the 5'-leader of BFD1 mRNA is sufficient for preferential translation upon stress. The translational control of BFD1 mRNA is maintained when ribosome assembly near its 5'-cap is impaired by insertion of a 5'-proximal stem-loop and upon knockdown of the Toxoplasma cap-binding protein, eIF4E1. Moreover, we show that a trans -acting RNA-binding protein called BFD2/ROCY1 is necessary for cap-independent translation of BFD1 through its binding to the 5'-leader. Translation of BFD2 mRNA is also suggested to be preferentially induced under stress, but by a cap-dependent mechanism. These results show that translational control and differentiation in Toxoplasma proceed through cap-independent mechanisms in addition to canonical cap-dependent translation. Our identification of cap-independent translation in protozoa underscores the antiquity of this mode of gene regulation in cellular evolution and its central role in stress-induced life-cycle events.

The protozoan parasite Toxoplasma gondii causes serious opportunistic disease due to its ability to persist in patients as latent tissue cysts. The molecular mechanisms coordinating conversion between proliferative parasites (tachyzoites) and latent cysts (bradyzoites) are not fully understood. We previously showed that phosphorylation of eIF2α accompanies bradyzoite formation, suggesting that this clinically relevant process involves regulation of mRNA translation. In this study, we investigated the composition and role of eIF4F multi-subunit complexes in translational control. Using CLIPseq, we find that the cap-binding subunit, eIF4E1, localizes to the 5'-end of all tachyzoite mRNAs, many of which show evidence of stemming from heterogeneous transcriptional start sites. We further show that eIF4E1 operates as the predominant cap-binding protein in two distinct eIF4F complexes. Using genetic and pharmacological approaches, we found that eIF4E1 deficiency triggers efficient spontaneous formation of bradyzoites without stress induction. Consistent with this result, we also show that stress-induced bradyzoites exhibit reduced eIF4E1 expression. Overall, our findings establish a novel role for eIF4F in translational control required for parasite latency and microbial persistence., Importance: Toxoplasma gondii is an opportunistic pathogen important to global human and animal health. There are currently no chemotherapies targeting the encysted form of the parasite. Consequently, a better understanding of the mechanisms controlling encystation is required. Here we show that the mRNA cap-binding protein, eIF4E1, regulates the encystation process. Encysted parasites reduce eIF4E1 levels, and depletion of eIF4E1 decreases the translation of ribosome-associated machinery and drives Toxoplasma encystation. Together, these data reveal a new layer of mRNA translational control that regulates parasite encystation and latency., Competing Interests: The authors declare no conflict of interest.

The parasite Toxoplasma gondii persists in its hosts by converting from replicating tachyzoites to latent bradyzoites housed in tissue cysts. The molecular mechanisms that mediate T. gondii differentiation remain poorly understood. Through a mutagenesis screen, we identified translation initiation factor eIF1.2 as a critical factor for T. gondii differentiation. A F97L mutation in eIF1.2 or the genetic ablation of eIF1.2 (∆eif1.2) markedly impeded bradyzoite cyst formation in vitro and in vivo. We demonstrated, at single-molecule level, that the eIF1.2 F97L mutation impacts the scanning process of the ribosome preinitiation complex on a model mRNA. RNA sequencing and ribosome profiling experiments unveiled that ∆eif1.2 parasites are defective in upregulating bradyzoite induction factors BFD1 and BFD2 during stress-induced differentiation. Forced expression of BFD1 or BFD2 significantly restored differentiation in ∆eif1.2 parasites. Together, our findings suggest that eIF1.2 functions by regulating the translation of key differentiation factors necessary to establish chronic toxoplasmosis., (© 2024. The Author(s).)

Toxoplasma gondii is an obligate intracellular parasite in the phylum Apicomplexa that causes toxoplasmosis in humans and animals worldwide. Despite its prevalence, there is currently no effective vaccine or treatment for chronic infection. Although there are therapies against the acute stage, prolonged use is toxic and poorly tolerated. This study aims to explore the potential of repurposing topotecan and 10-hydroxycamptothecin (HCPT) as drugs producing double strand breaks (DSBs) in T. gondii. DSBs are mainly repaired by Homologous Recombination Repair (HRR) and Non-Homologous End Joining (NHEJ). Two T. gondii strains, RHΔHXGPRT and RHΔKU80, were used to compare the drug's effects on parasites. RHΔHXGPRT parasites may use both HRR and NHEJ pathways but RHΔKU80 lacks the KU80 protein needed for NHEJ, leaving only the HRR pathway. Here we demonstrate that topotecan and HCPT, both topoisomerase I venoms, affected parasite replication in a concentration-dependent manner. Moreover, variations in fluorescence intensity measurements for the H2A.X phosphorylation mark (γH2A.X), an indicator of DNA damage, were observed in intracellular parasites under drug treatment conditions. Interestingly, intracellular replicative parasites without drug treatment show a strong positive staining for γH2A.X, suggesting inherent DNA damage. Extracellular (non-replicating) parasites did not exhibit γH2A.X staining, indicating that the basal level of DNA damage is likely to be associated with replicative stress. A high rate of DNA replication stress possibly prompted the evolution of an efficient repair machinery in the parasite, making it an attractive target. Our findings show that topoisomerase 1 venoms are effective antiparasitics blocking T. gondii replication., Competing Interests: Declaration of competing interest All authors declare that there are no conflicts of interest. There is no financial or non-financial assistance provided by a third party for the reported work. There is no financial interest or relationship related to the subject matter., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Identification of regulators of Toxoplasma gondii bradyzoite development and cyst formation is the most direct way to address the importance of parasite development in long-term persistence and reactivation of this parasite. Here we show that a T. gondii gene (named Regulator of Cystogenesis 1; ROCY1) is sufficient for T. gondii bradyzoite formation in vitro and in vivo. ROCY1 encodes an RNA binding protein that has a preference for 3' regulatory regions of hundreds of T. gondii transcripts, and its RNA-binding domains are required to mediate bradyzoite development. Female mice infected with ΔROCY1 parasites have reduced (>90%) cyst burden. While viable parasites can be cultivated from brain tissue for up to 6 months post-infection, chronic brain-resident ΔROCY1 parasites have reduced oral infectivity compared to wild type. Despite clear defects in bradyzoite formation and oral infectivity, ΔROCY1 parasites were able to reactivate with similar timing and magnitude as wild type parasites for up to 5 months post-infection. Therefore while ROCY1 is a critical regulator of the bradyzoite developmental pathway, it is not required for parasite reactivation, raising new questions about the persisting life stage responsible for causing recrudescent disease., (© 2023. Springer Nature Limited.)

Through regulation of DNA packaging, histone proteins are fundamental to a wide array of biological processes. A variety of post-translational modifications (PTMs), including acetylation, constitute a proposed histone code that is interpreted by "reader" proteins to modulate chromatin structure. Canonical histones can be replaced with variant versions that add an additional layer of regulatory complexity. The protozoan parasite Toxoplasma gondii is unique among eukaryotes in possessing a novel variant of H2B designated H2B.Z. The combination of PTMs and the use of histone variants are important for gene regulation in T. gondii, offering new targets for drug development. In this work, T. gondii parasites were generated in which the 5 N-terminal acetylatable lysines in H2B.Z were mutated to either alanine (c-Myc-A) or arginine (c-Myc-R). The c-Myc-A mutant displayed no phenotype over than a mild defect in its ability to kill mice. The c-Myc-R mutant presented an impaired ability to grow and an increase in differentiation to latent bradyzoites. The c-Myc-R mutant was also more sensitive to DNA damage, displayed no virulence in mice, and provided protective immunity against future infection. While nucleosome composition was unaltered, key genes were abnormally expressed during in vitro bradyzoite differentiation. Our results show that regulation of the N-terminal positive charge patch of H2B.Z is important for these processes. We also show that acetylated N-terminal H2B.Z interacts with some unique proteins compared to its unacetylated counterpart; the acetylated peptide pulled down proteins associated with chromosome maintenance/segregation and cell cycle, suggesting a link between H2B.Z acetylation status and mitosis., Competing Interests: Declaration of competing interest The authors declare that there are no conflict of interest., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Background: Older adult Veterans are at high risk for adverse health outcomes following hospitalization. Since physical function is one of the largest potentially modifiable risk factors for adverse health outcomes, our purpose was to determine if progressive, high-intensity resistance training in home health physical therapy (PT) improves physical function in Veterans more than standardized home health PT and to determine if the high-intensity program was comparably safe, defined as having a similar number of adverse events., Methods: We enrolled Veterans and their spouses during an acute hospitalization who were recommended to receive home health care on discharge because of physical deconditioning. We excluded individuals who had contraindications to high-intensity resistance training. A total of 150 participants were randomized 1:1 to either (1) a progressive, high-intensity (PHIT) PT intervention or (2) a standardized PT intervention (comparison group). All participants in both groups were assigned to receive 12 visits (3 visits/week over 30 days) in their home. The primary outcome was gait speed at 60 days. Secondary outcomes included adverse events (rehospitalizations, emergency department visits, falls and deaths after 30 and 60-days), gait speed, Modified Physical Performance Test, Timed Up-and-Go, Short Physical Performance Battery, muscle strength, Life-Space Mobility assessment, Veterans RAND 12-item Health Survey, Saint Louis University Mental Status exam, and step counts at 30, 60, 90, 180 days post-randomization., Results: There were no differences between groups in gait speed at 60 days, and no significant differences in adverse events between groups at either time point. Similarly, physical performance measures and patient reported outcomes were not different at any time point. Notably, participants in both groups experienced increases in gait speed that met or exceeded established clinically important thresholds., Conclusions: Among older adult Veterans with hospital-associated deconditioning and multimorbidity, high-intensity home health PT was safe and effective in improving physical function, but not found to be more effective than a standardized PT program., (© 2023 The American Geriatrics Society. This article has been contributed to by U.S. Government employees and their work is in the public domain in the USA.)

Toxoplasma gondii is a widespread protozoan parasite that has a significant impact on human and veterinary health. The parasite undergoes a complex life cycle involving multiple hosts and developmental stages. How Toxoplasma transitions between life cycle stages is poorly understood yet central to controlling transmission. Of particular neglect are the factors that contribute to its sexual development, which takes place exclusively in feline intestines. While epigenetic repressors have been shown to play an important role in silencing the spurious gene expression of sexually committed parasites, the specific factors that recruit this generalized machinery to the appropriate genes remain largely unexplored. Here, we establish that a member of the AP2 transcription factor family, AP2XII-2, is targeted to genomic loci associated with sexually committed parasites along with epigenetic regulators of transcriptional silencing, HDAC3 and MORC. Despite its widespread association with gene promoters, AP2XII-2 is required for the silencing of relatively few genes. Using the CUT&Tag (cleavage under targets and tagmentation) methodology, we identify two major genes associated with sexual development downstream of AP2XII-2 control, AP2X-10 and the amino acid hydroxylase AAH1. Our findings show that AP2XII-2 is a key contributor to the gene regulatory pathways modulating Toxoplasma sexual development. IMPORTANCE Toxoplasma gondii is a parasite that undergoes its sexual stage exclusively in feline intestines, making cats a major source of transmission. A better understanding of the proteins controlling the parasite's life cycle stage transitions is needed for the development of new therapies aimed at treating toxoplasmosis and the transmission of the infection. Genes that regulate the sexual stages need to be turned on and off at the appropriate times, activities that are mediated by specific transcription factors that recruit general machinery to silence or activate gene expression. In this study, we identify a transcription factor called AP2XII-2 as being important for the repression of a subset of sexual stage genes, including a sexual stage-specific AP2 factor (AP2X-10) and a protein (AAH1) required to construct the infectious oocysts expelled from infected cats.

Purpose: Globally, spine disorders are the leading cause of disability, affecting more than half a billion individuals. However, less than 50% of G20 countries specifically identify spine health within their public policy priorities. Therefore, it is crucial to raise awareness among policy makers of the disabling effect of spine disorders and their impact on the economic welfare of G20 nations. In 2019, SPINE20 was established as the leading advocacy group to bring global attention to spine disorders., Methods: Recommendations were developed through two Delphi methods with international and multi-professional panels., Results: In 2022, seven recommendations were delivered to the leaders of G20 countries, urging them to: Develop action plans to provide universal access to evidence-based spine care that incorporates the needs of minorities and vulnerable populations. Invest in the development of sustainable human resource capacity, through multisectoral and inter-professional competency-based education and training to promote evidence-based approaches to spine care, and to build an appropriate healthcare working environment that optimizes the delivery of safe health services. Develop policies using the best available evidence to properly manage spine disorders and to prolong functional healthy life expectancy in the era of an aging population. Create a competent workforce and improve the healthcare infrastructure/facilities including equipment to provide evidence-based inter-professional rehabilitation services to patients with spinal cord injury throughout their continuum of care. Build collaborative and innovative translational research capacity within national, regional, and global healthcare systems for state-of-the-art and cost-effective spine care across the healthcare continuum ensuring equality, diversity, and inclusion of all stakeholders. Develop international consensus statements on patient outcomes and how they can be used to define and develop pathways for value-based care. Recognize that intervening on determinants of health including physical activity, nutrition, physical and psychosocial workplace environment, and smoking-free lifestyle can reduce the burden of spine disabilities and improve the health status and wellness of the population. At the third SPINE20 summit 2022 which took place in Bali, Indonesia, in August 2022, 17 associations endorsed its recommendations., Conclusion: SPINE20 advocacy efforts focus on developing public policy recommendations to improve the health, welfare, and wellness of all who suffer from spinal pain and disability. We propose that focusing on facilitating access to systems that prioritize value-based care delivered by a competent healthcare workforce will reduce disability and improve the productivity of the G20 nations., (© 2022. The Author(s).)

Entamoeba histolytica is a protozoan parasite that causes amoebic dysentery and liver abscess. This pathogen possesses a two-stage life cycle consisting of an environmentally stable cyst and a pathogenic amoeboid trophozoite. Since infection is acquired by ingestion of cysts from contaminated food and water, this parasite is prevalent in underdeveloped countries. A reptilian pathogen, Entamoeba invadens, which can encyst in culture, has long served as a surrogate to study stage conversion. In the host, Entamoeba species must manage stress, including nutrient deprivation and host immune pressure. In many systems, the stress response is characterized by downregulation of translation, which is initiated by the phosphorylation of eukaryotic initiation factor-2 alpha (eIF2α). In mammalian cells, this phosphorylation is carried out by a family of eIF2α kinases. A canonical eIF2α translational control system exists in Entamoeba species; however, no eIF2α kinases have been characterized. In this study, we identified two eIF2α kinases in E. invadens , EiIF2K-A and EiIF2K-B. Their identity as eIF2α kinases was validated using a heterologous yeast system. We used an RNA interference (RNAi) trigger-mediated silencing system to reduce expression of EiIF2K-A, which also reduced expression of EiIF2K-B. Parasites with decreased kinase expression exhibited decreased phosphorylation of eIF2α and increased sensitivity to oxidative stress. Diminished kinase expression also correlated with an increased rate of encystation, a decreased rate of excystation, and an increase in several virulence functions, erythrophagocytosis and adhesion to host cells. Taken together, these data suggest that EiIF2K-A and EiIF2K-B are authentic eIF2α kinases that may regulate the Entamoeba stress response. IMPORTANCE Entamoeba histolytica is a human pathogen that causes dysentery and affects millions of people worldwide. This parasite possesses a two-stage life cycle: an environmentally stable cyst and the pathogenic trophozoite. Cysts are ingested from contaminated food and water; thus, this parasite in prevalent in underdeveloped countries. Current therapies commonly cause adverse side effects; therefore, new treatments are needed. In the host, Entamoeba experiences stress brought on, in part, by the host immune system. Understanding stage conversion and the stress response of this pathogen may lead to new drug therapies. Using the model organism E. invadens , we identified two kinases similar to those involved in stress and stage conversion in other systems. We determined that these kinases may regulate the oxidative stress response, stage conversion, and virulence. This work is significant, as it will inform future studies on the life cycle and pathogenicity of Entamoeba species.

Purpose: The focus of SPINE20 is to develop evidence-based policy recommendations for the G20 countries to work with governments to reduce the burden of spine disease, and disability., Methods: On September 17-18, 2021, SPINE20 held its annual meeting in Rome, Italy. Prior to the meeting, the SPINE20 created six proposed recommendations. These recommendations were uploaded to the SPINE20 website 10 days before the meeting and opened to the public for comments. The recommendations were discussed at the meeting allowing the participants to object and provide comments., Results: In total, 27 societies endorsed the following recommendations. SPINE20 calls upon the G20 countries: (1) to expand telehealth for the access to spine care, especially in light of the current situation with COVID-19. (2) To adopt value-based interprofessional spine care as an approach to improve patient outcomes and reduce disability. (3) To facilitate access and invest in the development of a competent rehabilitation workforce to reduce the burden of disability related to spine disorders. (4) To adopt a strategy to promote daily physical activity and exercises among the elderly population to maintain an active and independent life with a healthy spine, particularly after COVID-19 pandemic. (5) To engage in capacity building with emerging countries and underserved communities for the benefit of spine patients. (6) To promote strategies to transfer evidence-based advances into patient benefit through effective implementation processes., Conclusions: SPINE20's initiatives will make governments and decision makers aware of efforts to reduce needless suffering from disabling spine pain through education that can be instituted across the globe., (© 2022. The Author(s).)

Background: Toxoplasma gondii is a protozoan parasite that differentiates from acute tachyzoite stages to latent bradyzoite forms in response to environmental cues that modify the epigenome. We studied the distribution of the histone variants CenH3, H3.3, H2A.X, H2A.Z and H2B.Z, by genome-wide chromatin immunoprecipitation to understand the role of variant histones in developmental transitions of T. gondii parasites., Results: H3.3 and H2A.X were detected in telomere and telomere associated sequences, whereas H3.3, H2A.X and CenH3 were enriched in centromeres. Histones H2A.Z and H2B.Z colocalize with the transcriptional activation mark H3K4me3 in promoter regions surrounding the nucleosome-free region upstream of the transcription start site. The H2B.Z/H2A.Z histone pair also localizes to the gene bodies of genes that are silent but poised for activation, including bradyzoite stage-specific genes. The majority of H2A.X and H2A.Z/H2B.Z loci do not overlap, consistent with variant histones demarcating specific functional regions of chromatin. The extent of enrichment of H2A.Z/H2B.Z (and H3.3 and H2A.X) within the entire gene (5'UTR and gene body) reflects the timing of gene expression during the cell cycle, suggesting that dynamic turnover of H2B.Z/H2A.Z occurs during the tachyzoite cell cycle. Thus, the distribution of the variant histone H2A.Z/H2B.Z dimer defines active and developmentally silenced regions of the T. gondii epigenome including genes that are poised for expression., Conclusions: Histone variants mark functional regions of parasite genomes with the dynamic placement of the H2A.Z/H2B.Z dimer implicated as an evolutionarily conserved regulator of parasite and eukaryotic differentiation., (© 2022. The Author(s).)

Abstract: In response to increasing dependence on US Medical Licensing Examination Step 1 scores for resident selection, and to improve medical student well-being and education, the Federation of State Medical Boards and the National Board of Medical Examiners announced that no sooner than 2022, Step 1 score reporting would change from numeric to pass/fail. To gauge the impact that this change will have on the residency application process in physical medicine and rehabilitation, an anonymous survey was electronically distributed to program directors of all accredited physical medicine and rehabilitation residency programs in the United States. A response rate of 49% was obtained. In total, 13% of program directors agreed that the scoring change was a good idea. Eighty percent of program directors felt that the change would increase emphasis on Step 2 clinical knowledge in the application process and that it will be more difficult to objectively compare applicants. Among free-text responses to the survey, 70% disapproved, 10% approved, and the remaining 20% were neutral to the change. The results of this survey show that many physical medicine and rehabilitation program directors are concerned about the implications of the Step 1 score reporting change. These data can serve as a helpful guide to program directors and students preparing for a pass/fail Step 1., Competing Interests: Financial disclosure statements have been obtained, and no conflicts of interest have been reported by the authors or by any individuals in control of the content of this article., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)

Background Context: High quality evidence is difficult to generate, leaving substantial knowledge gaps in the treatment of spinal conditions. Appropriate use criteria (AUC) are a means of determining appropriate recommendations when high quality evidence is lacking., Purpose: Define appropriate use criteria (AUC) of cervical fusion for treatment of degenerative conditions of the cervical spine., Study Design/setting: Appropriate use criteria for cervical fusion were developed using the RAND/UCLA appropriateness methodology. Following development of clinical guidelines and scenario writing, a one-day workshop was held with a multidisciplinary group of 14 raters, all considered thought leaders in their respective fields, to determine final ratings for cervical fusion appropriateness for various clinical situations., Outcome Measures: Final rating for cervical fusion recommendation as either "Appropriate," "Uncertain" or "Rarely Appropriate" based on the median final rating among the raters., Methods: Inclusion criteria for scenarios included patients aged 18 to 80 with degenerative conditions of the cervical spine. Key modifiers were defined and combined to develop a matrix of clinical scenarios. The median score among the raters was used to determine the final rating for each scenario. The final rating was compared between modifier levels. Spearman's rank correlation between each modifier and the final rating was determined. A multivariable ordinal regression model was fit to determine the adjusted odds of an "Appropriate" final rating while adjusting for radiographic diagnosis, number of levels and symptom type. Three decision trees were developed using decision tree classification models and variable importance for each tree was computed., Results: Of the 263 scenarios, 47 (17.9 %) were rated as rarely appropriate, 66 (25%) as uncertain and 150 (57%) were rated as appropriate. Symptom type was the modifier most strongly correlated with the final rating (adjusted ρ2 = 0.58, p<.01). A multivariable ordinal regression adjusting for symptom type, diagnosis, and number of levels and showed high discriminative ability (C statistic = 0.90) and the adjusted odds ratio (aOR) of receiving a final rating of "Appropriate" was highest for myelopathy (aOR, 7.1) and radiculopathy (aOR, 4.8). Three decision tree models showed that symptom type and radiographic diagnosis had the highest variable importance., Conclusions: Appropriate use criteria for cervical fusion in the setting of cervical degenerative disorders were developed. Symptom type was most strongly correlated with final rating. Myelopathy or radiculopathy were most strongly associated with an "Appropriate" rating, while axial pain without stenosis was most associated with "Rarely Appropriate.", (Copyright © 2021 Elsevier Inc. All rights reserved.)

Toxoplasma gondii is a protozoan parasite that causes opportunistic infection in immunocompromised individuals. The parasite forms latent tissue cysts that are refractory to current treatments and give rise to life-threatening reactivated infection following immune suppression. Previously, we showed that guanabenz sharply reduces brain cyst count in BALB/c mice harboring latent toxoplasmosis; however, whether cyst count would change once drug treatment stopped was not addressed. In the present study, we observed a rebound in brain cysts following the discontinuation of guanabenz or a guanabenz-pyrimethamine combination therapy. The re-expansion of brain cysts was not accompanied by symptoms of acute toxoplasmosis. We also tested whether the rebound in cyst counts could be ameliorated by administering pyrimethamine during or after guanabenz treatment., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Purpose: The Global Burden of Diseases (GBD) Studies have estimated that low back pain is one of the costliest ailments worldwide. Subsequent to GBD publications, leadership of the four largest global spine societies agreed to form SPINE20. This article introduces the concept of SPINE20, the recommendations, and the future of this global advocacy group linked to G20 annual summits., Methods: The founders of SPINE20 advocacy group coordinated with G20 Saudi Arabia to conduct the SPINE20 summit in 2020. The summit was intended to promote evidence-based recommendations to use the most reliable information from high-level research. Eight areas of importance to mitigate spine disorders were identified through a voting process of the participating societies. Twelve recommendations were discussed and vetted., Results: The areas of immediate concern were "Aging spine," "Future of spine care," "Spinal cord injuries," "Children and adolescent spine," "Spine-related disability," "Spine Educational Standards," "Patient safety," and "Burden on economy." Twelve recommendations were created and endorsed by 31/33 spine societies and 2 journals globally during a vetted process through the SPINE20.org website and during the virtual inaugural meeting November 10-11, 2020 held from the G20 platform., Conclusions: This is the first time that international spine societies have joined to support actions to mitigate the burden of spine disorders across the globe. SPINE20 seeks to change awareness and treatment of spine pain by supporting local projects that implement value-based practices with healthcare policies that are culturally sensitive based on scientific evidence., (© 2021. The Author(s).)

To curtail the rising tide of antiscience threatening the health and progress of society, scientists are increasingly engaging with the public. Here, we describe our approach to write accessible books based on personal stories as a means to help spread scientific literacy to those who normally do not read science., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Toxoplasma gondii is an obligate intracellular parasite that can cause serious opportunistic disease in the immunocompromised or through congenital infection. To progress through its life cycle, Toxoplasma relies on multiple layers of gene regulation that includes an array of transcription and epigenetic factors. Over the last decade, the modification of mRNA has emerged as another important layer of gene regulation called epitranscriptomics. Here, we report that epitranscriptomics machinery exists in Toxoplasma, namely the methylation of adenosines (m6A) in mRNA transcripts. We identified novel components of the m6A methyltransferase complex and determined the distribution of m6A marks within the parasite transcriptome. m6A mapping revealed the modification to be preferentially located near the 3'-boundary of mRNAs. Knockdown of the m6A writer components METTL3 and WTAP resulted in diminished m6A marks and a complete arrest of parasite replication. Furthermore, we examined the two proteins in Toxoplasma that possess YTH domains, which bind m6A marks, and showed them to be integral members of the cleavage and polyadenylation machinery that catalyzes the 3'-end processing of pre-mRNAs. Loss of METTL3, WTAP, or YTH1 led to a defect in transcript 3'-end formation. Together, these findings establish that the m6A epitranscriptome is essential for parasite viability by contributing to the processing of mRNA 3'-ends., Competing Interests: The authors have declared that no competing interests exist.

The intracellular parasite Toxoplasma gondii infects nucleated cells in virtually all warm-blooded vertebrates, including one-third of the human population. While immunocompetent hosts do not typically show symptoms of acute infection, parasites are retained in latent tissue cysts that can be reactivated upon immune suppression, potentially damaging key organ systems. Toxoplasma has a multistage life cycle that is intimately linked to environmental stresses and host signals. As this protozoan pathogen is transmitted between multiple hosts and tissues, it evaluates these external signals to appropriately differentiate into distinct life cycle stages, such as the transition from its replicative stage (tachyzoite) to the latent stage (bradyzoite) that persists as tissue cysts. Additionally, in the gut of its definitive host, felines, Toxoplasma converts into gametocytes that produce infectious oocysts (sporozoites) that are expelled into the environment. In this review, we highlight recent advances that have illuminated the interfaces between Toxoplasma and host and how these interactions control parasite stage conversion. Mechanisms underlying these stage transitions are important targets for therapeutic intervention aimed at thwarting parasite transmission and pathogenesis., (© 2020 John Wiley & Sons Ltd.)

Toxoplasma gondii is an obligate intracellular parasite that persists in its vertebrate hosts in the form of dormant tissue cysts, which facilitate transmission through predation. The parasite must strike a balance that allows it to disseminate throughout its host without killing it, which requires the ability to properly counter host cell defenses. For example, oxidative stress encountered by Toxoplasma is suggested to impair parasite replication and dissemination. However, the strategies by which Toxoplasma mitigates oxidative stress are not yet clear. Among eukaryotes, environmental stresses induce the integrated stress response via phosphorylation of a translation initiation factor, eukaryotic initiation factor 2 (eIF2). Here, we show that the Toxoplasma eIF2 kinase TgIF2K-B is activated in response to oxidative stress and affords protection. Knockout of the TgIF2K-B gene, Δ tgif2k-b , disrupted parasite responses to oxidative stresses and enhanced replication, diminishing the ability of the parasite to differentiate into tissue cysts. In addition, parasites lacking TgIF2K-B exhibited resistance to activated macrophages and showed greater virulence in an in vivo model of infection. Our results establish that TgIF2K-B is essential for Toxoplasma responses to oxidative stress, which are important for the parasite's ability to establish persistent infection in its host. IMPORTANCE Toxoplasma gondii is a single-celled parasite that infects nucleated cells of warm-blooded vertebrates, including one-third of the human population. The parasites are not cleared by the immune response and persist in the host by converting into a latent tissue cyst form. Development of tissue cysts can be triggered by cellular stresses, which activate a family of TgIF2 kinases to phosphorylate the eukaryotic translation initiation factor TgIF2α. Here, we establish that the TgIF2 kinase TgIF2K-B is activated by oxidative stress and is critical for maintaining oxidative balance in the parasite. Depletion of TgIF2K-B alters gene expression, leading to accelerated growth and a diminished ability to convert into tissue cysts. This study establishes that TgIF2K-B is essential for the parasite's oxidative stress response and its ability to persist in the host as a latent infection., (Copyright © 2021 Augusto et al.)

Background: Lower-limb amputation (LLA) due to non-traumatic vascular etiology is linked to extremely low physical activity and high disability., Objective: To test the feasibility of a biobehavioral intervention designed to promote physical activity., Design: A randomized, single-blind feasibility trial with a crossover design., Setting: Veterans Administration Medical Center., Participants: Military veterans (age: 65.7 [7.8] years; mean [standard deviation]) with nontraumatic lower-limb amputation (LLA), randomized to two groups: GROUP1 (n = 16) and GROUP2 (n = 15). Both groups had similar baseline amputation characteristics (level of amputation and time since amputation)., Interventions: Twelve weekly, 30-minute telehealth sessions of physical activity behavior-change intervention, with GROUP1 participating in weeks 1-12 and GROUP2 in weeks 13-24. GROUP1 noncontact phase in weeks 13-24 and GROUP2 attention control telehealth phase in weeks 1-12., Main Outcome Measures: Feasibility (participant retention, dose goal attainment, intervention acceptability [Intrinsic Motivation Inventory [IMI] Interest and Enjoyment scale], safety) and signal of efficacy (free-living physical activity [accelerometer-based average daily step count], Late Life Function and Disability Index - Disability Scale [LLFDI-DS])., Results: Participant retention rate was high (90%), with three participants lost to follow-up during the intervention period. Dose goal attainment was low, with only 10% of participants achieving an a priori walking dose goal. Intervention was rated as acceptable, with mean IMI Interest and Enjoyment score (5.8) statistically higher than the null value of 5.0 (P = .002). There were no between-group differences in adverse event rates (falls: P = .19, lower extremity wounds: P = .60). There was no signal of efficacy for change in average daily step count (d = -0.15) or LLFDI-DS (d = -0.22 and 0.17 for frequency and limitations scales, respectively)., Conclusions: Telehealth delivered biobehavioral intervention resulted in acceptable participant retention, low dose goal attainment, high participant acceptability, and low safety risk, while having no signal of efficacy (physical activity, disability) for people with nontraumatic LLA., (© 2020 American Academy of Physical Medicine and Rehabilitation. This article has been contributed to by US Government employees and their work is in the public domain in the USA.)

Toxoplasma gondii is a protozoan parasite that causes lifelong chronic infection that can reactivate in immunocompromised individuals. Upon infection, the replicative stage (tachyzoite) converts into a latent tissue cyst stage (bradyzoite). Like other apicomplexans, T. gondii possesses an extensive lineage of proteins called ApiAP2s that contain DNA-binding domains first characterized in plants. The function of most ApiAP2s is unknown. We previously found that AP2IX-4 is a cell cycle-regulated ApiAP2 expressed only in dividing parasites as a putative transcriptional repressor. In this study, we purified proteins interacting with AP2IX-4, finding it to be a component of the recently characterized microrchidia (MORC) transcriptional repressor complex. We further analyzed AP2XII-2, another cell cycle-regulated factor that associates with AP2IX-4. We monitored parallel expression of AP2IX-4 and AP2XII-2 proteins in tachyzoites, detecting peak expression during S/M phase. Unlike AP2IX-4, which is dispensable in tachyzoites, loss of AP2XII-2 resulted in a slowed tachyzoite growth due to a delay in S-phase progression. We also found that AP2XII-2 depletion increased the frequency of bradyzoite differentiation in vitro These results suggest that multiple AP2 factors collaborate to ensure proper cell cycle progression and tissue cyst formation in T. gondii IMPORTANCE Toxoplasma gondii is a single-celled parasite that persists in its host by converting into a latent cyst stage. This work describes a new transcriptional factor called AP2XII-2 that plays a role in properly maintaining the growth rate of replicating parasites, which contributes to signals required for development into its dormant stage. Without AP2XII-2, Toxoplasma parasites experience a delay in their cell cycle that increases the frequency of latent cyst formation. In addition, we found that AP2XII-2 operates in a multisubunit complex with other AP2 factors and chromatin remodeling machinery that represses gene expression. These findings add to our understanding of how Toxoplasma parasites balance replication and dormancy, revealing novel points of potential therapeutic intervention to disrupt this clinically relevant process., (Copyright © 2020 Srivastava et al.)

Toxoplasma gondii , an obligate intracellular parasite that can cause life-threatening acute disease, differentiates into a quiescent cyst stage to establish lifelong chronic infections in animal hosts, including humans. This tissue cyst reservoir, which can reactivate into an acute infection, is currently refractory to clinically available therapeutics. Recently, we and others have discovered drugs capable of significantly reducing the brain cyst burden in latently infected mice, but not to undetectable levels. In this study, we examined the use of novel combination therapies possessing multiple mechanisms of action in mouse models of latent toxoplasmosis. Our drug regimens included combinations of pyrimethamine, clindamycin, guanabenz, and endochin-like quinolones (ELQs) and were administered to two different mouse strains in an attempt to eradicate brain tissue cysts. We observed mouse strain-dependent effects with these drug treatments: pyrimethamine-guanabenz showed synergistic efficacy in C57BL/6 mice yet did not improve upon guanabenz monotherapy in BALB/c mice. Contrary to promising in vitro results demonstrating toxicity to bradyzoites, we observed an antagonistic effect between guanabenz and ELQ-334 in vivo While we were unable to completely eliminate the brain cyst burden, we found that a combination treatment with ELQ-334 and pyrimethamine impressively reduced the brain cyst burden by 95% in C57BL/6 mice, which approached the limit of detection. These analyses highlight the importance of evaluating anti-infective drugs in multiple mouse strains and will help inform further preclinical studies of cocktail therapies designed to treat chronic toxoplasmosis., (Copyright © 2020 American Society for Microbiology.)

Toxoplasma gondii is an intracellular parasite that reconfigures its host cell to promote pathogenesis. One consequence of Toxoplasma parasitism is increased migratory activity of host cells, which facilitates dissemination. Here, we show that Toxoplasma triggers the unfolded protein response (UPR) in host cells through calcium release from the endoplasmic reticulum (ER). We further identify a novel role for the host ER stress sensor protein IRE1 in Toxoplasma pathogenesis. Upon infection, Toxoplasma activates IRE1, engaging its noncanonical role in actin remodeling through the binding of filamin A. By inducing cytoskeletal remodeling via IRE1 oligomerization in host cells, Toxoplasma enhances host cell migration in vitro and dissemination of the parasite to host organs in vivo Our study has identified novel mechanisms used by Toxoplasma to induce dissemination of infected cells, providing new insights into strategies for treatment of toxoplasmosis. IMPORTANCE Cells that are infected with the parasite Toxoplasma gondii exhibit heightened migratory activity, which facilitates dissemination of the infection throughout the body. In this report, we identify a new mechanism used by Toxoplasma to hijack its host cell and increase its mobility. We further show that the ability of Toxoplasma to increase host cell migration involves not the enzymatic activity of IRE1 but rather IRE1 engagement with actin cytoskeletal remodeling. Depletion of IRE1 from infected host cells reduces their migration in vitro and significantly hinders dissemination of Toxoplasma in vivo Our findings reveal a new mechanism underlying host-pathogen interactions, demonstrating how host cells are co-opted to spread a persistent infection around the body., (Copyright © 2020 Augusto et al.)

Toxoplasma gondii is the causative agent of toxoplasmosis in animals and humans. This infection is transmitted to humans through oocysts released in the feces of the felines into the environment or by ingestion of undercooked meat. This implies that toxoplasmosis is a zoonotic disease and T. gondii is a foodborne pathogen. In addition, chronic toxoplasmosis in goats and sheep is the cause of recurrent abortions with economic losses in the sector. It is also a health problem in pets such as cats and dogs. Although there are therapies against this infection in its acute stage, they are not able to permanently eliminate the parasite and sometimes they are not well tolerated. To develop better, safer drugs, we need to elucidate key aspects of the biology of T. gondii . In this review, we will discuss the importance of the homologous recombination repair (HRR) pathway in the parasite's lytic cycle and how components of these processes can be potential molecular targets for new drug development programs. In that sense, the effect of different DNA damage agents or HHR inhibitors on the growth and replication of T. gondii will be described. Multitarget drugs that were either associated with other targets or were part of general screenings are included in the list, providing a thorough revision of the drugs that can be tested in other scenarios., (Copyright © 2020 Angel, Vanagas, Ruiz, Cristaldi, Saldarriaga Cartagena and Sullivan.)

Toxoplasma gondii is a protozoan parasite of great importance in human and veterinary health. The frontline treatment of antifolates suffers a variety of drawbacks, including toxicity and allergic reactions, underscoring the need to identify novel drug targets for new therapeutics to be developed. We previously showed that the Toxoplasma lysine acetyltransferase (KAT) GCN5b is an essential chromatin remodeling enzyme in the parasite linked to the regulation of gene expression. We have previously established that the KAT domain is a liability that can be targeted in the parasite by compounds like garcinol; here, we investigate the potential of the bromodomain as a targetable element of GCN5b. Bromodomains bind acetylated lysine residues on histones, which helps stabilize the KAT complex at gene promoters. Using an inducible dominant-negative strategy, we found that the GCN5b bromodomain is critical for Toxoplasma viability. We also found that the GCN5-family bromodomain inhibitor, L-Moses, interferes with the ability of the GCN5b bromodomain to associate with acetylated histone residues using an in vitro binding assay. Moreover, L-Moses displays potent activity against Toxoplasma tachyzoites in vitro, which can be overcome if parasites are engineered to over-express GCN5b. Collectively, our data support the GCN5b bromodomain as an attractive target for the development of new therapeutics., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Brain-eating amoebae cause devastating infections in the central nervous system of humans, resulting in a mortality rate of 95%. There are limited effective therapeutic options available clinically for treating granulomatous amoebic encephalitis and primary amoebic meningoencephalitis caused by Acanthamoeba castellanii ( A. castellanii ) and Naegleria fowleri ( N. fowleri ), respectively. Here, we report for the first time that guanabenz conjugated to gold and silver nanoparticles has significant antiamoebic activity against both A. castellanii and N. fowleri . Gold and silver conjugated guanabenz nanoparticles were synthesized by the one-phase reduction method and were characterized by ultraviolet-visible spectrophotometry and atomic force microscopy. Both metals were facilely stabilized by the coating of guanabenz, which was examined by surface plasmon resonance determination. The average size of gold nanoconjugated guanabenz was found to be 60 nm, whereas silver nanoparticles were produced in a larger size distribution with the average diameter of around 100 nm. Guanabenz and its noble metal nanoconjugates exhibited potent antiamoebic effects in the range of 2.5 to 100 μM against both amoebae. Nanoparticle conjugation enhanced the antiamoebic effects of guanabenz, as more potent activity was observed at a lower effective concentration (2.5 and 5 μM) compared to the drug alone. Moreover, encystation and excystation assays revealed that guanabenz inhibits the interconversion between the trophozoite and cyst forms of A. castellanii . Cysticdal effects against N. fowleri were also observed. Notably, pretreatment of A. castellanii with guanabenz and its nanoconjugates exhibited a significant reduction in the host cell cytopathogenicity from 65% to 38% and 2% in case of gold and silver nanoconjugates, respectively. Moreover, the cytotoxic evaluation of guanabenz and its nanoconjugates revealed negligible cytotoxicity against human cells. Guanabenz is already approved for hypertension and crosses the blood-brain barrier; the results of our current study suggest that guanabenz and its conjugated gold and silver nanoparticles can be repurposed as a potential drug for treating brain-eating amoebic infections.

Entamoeba histolytica is an intestinal parasite infecting over 50 million people worldwide and is the causative agent of amebic dysentery and amoebic liver abscess. In the human host, E. histolytica experiences stress brought on by nutrient deprivation and the host immune response. To be a successful parasite, E. histolytica must counter the stress; therefore, understanding the stress response may uncover new drug targets. In many systems, the stress response includes down-regulation of protein translation, which is regulated by phosphorylation of eukaryotic initiation factor (eIF-2α). Previous work has demonstrated that phosphorylation of the E. histolytica eIF-2α (EheIF-2α) increases significantly when exposed to long-term serum starvation, oxidative stress, and long-term heat shock. However, the effects of reagents that are known to induce nitrosative or endoplasmic reticulum (ER) stresses, on EheIF-2α have yet to be evaluated. Nitrosative stress is part of the host's immune response and ER stress can be caused by several physiological or pathological factors. We treated E. histolytica cells with various reagents known to induce nitrosative stress (DPTA-NONOate and SNP) or ER stress (BFA and DTT). We examined the morphology of the ER, tracked phosphorylation of EheIF-2α, and assessed protein translation in control and stressed cells. While all four stress-inducing reagents caused a global reduction in protein translation, only DTT was capable of also inducing changes in the morphology of the ER (consistent with ER stress) and phosphorylation of EheIF-2α. This suggests that DTT authentically induces ER stress in E. histolytica and that this stress is managed by the eIF-2α-based system. This was supported by the observation that cells expressing a non-phosphorylatable version of eIF-2α were also highly sensitive to DTT-stress. Since protein translation decreased in the absence of phosphorylation of eIF-2α (after treatment with DPTA-NONOate, SNP or BFA), the data also indicate that there are alternative protein-translational control pathways in E. histolytica. Overall, our study further illuminates the stress response to nitrosative stress and ER stress in E. histolytica., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Toxoplasma gondii is a protozoan parasite that has a tremendous impact on human health and livestock. High seroprevalence among humans and other animals is facilitated by the conversion of rapidly proliferating tachyzoites into latent bradyzoites that are housed in tissue cysts, which allow transmission through predation. Epigenetic mechanisms contribute to the regulation of gene expression events that are crucial in both tachyzoites as well as their development into bradyzoites. Acetylation of histones is one of the critical histone modifications that is linked to active gene transcription. Unlike most early-branching eukaryotes, Toxoplasma possesses two GCN5 homologues, one of which, GCN5b, is essential for parasite viability. Surprisingly, GCN5b does not associate with most of the well-conserved proteins found in the GCN5 complexes of other eukaryotes. Of particular note is that GCN5b interacts with multiple putative transcription factors that have plant-like DNA-binding domains denoted as AP2. To understand the function of GCN5b and its role(s) in epigenetic gene regulation of stage switching, we performed co-immunoprecipitation of GCN5b under normal and bradyzoite induction conditions. We report the greatest resolution of the GCN5b complex to date under these various culture conditions. Moreover, reciprocal co-IPs were performed with distinct GCN5b-interacting AP2 factors (AP2IX-7 and AP2XII-4) to delineate the interactomes of each putative transcription factor. Our findings suggest that GCN5b is associated with at least two distinct complexes that are characterized by two different pairs of AP2 factors, and implicate up to four AP2 proteins to be involved with GCN5b-mediated gene regulation., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Background Context: The need for advanced imaging before spinal intervention is an area of ongoing debate. Many studies have demonstrated the accuracy of magnetic resonance imaging (MRI) results in evaluating structural pathology in the lumbar spine, but few have addressed how frequently MRI findings change clinical management. A randomized controlled trial showed that viewing MRI results did not impact outcomes in patients with radiculopathy undergoing epidural steroid injection (ESI). The results suggested ESIs that correlated with both imaging and clinical findings experienced slightly more benefit than the blinded cohort, although statistically insignificantly., Purpose: Three related studies were conducted to (1) increase understanding of the opinions of interventional spine physicians regarding the utility of viewing imaging before injection and (2) evaluate the impact of viewing MRI results on injection planning (retrospective and prospective analyses)., Study Design: Survey, prospective, and retrospective analysis., Patient Sample: Patients presenting to a university-based spine center for initial evaluation of back or leg pain who were candidates for spinal intervention., Outcome Measures: Self-reported measures from a clinical practice questionnaire distributed to interventional spine physicians to determine rates and rationale for utilization of MRI before spine injection, physiologic measures including MRI results, functional measures including physician decision-making regarding type and location of injection performed., Methods: This study was funded by the University of Colorado Health and Welfare Trust. A survey was sent to interventional spine physicians to assess their utilization of MRI results before spine procedures. A retrospective analysis of patients who were candidates for ESI was conducted to evaluate how initial injection plan compared with the postviewing of MRI results on injection performed. In a prospective analysis, injection plans pre- and post-MRI were compared among patients presenting for initial evaluation of low back or leg pain., Results: Survey responses showed that specialists order MRI studies to correlate with physical exam (91%) and to detect the presence of synovial cysts (68%), whereas tumor/infection (93%) was most likely to cause a change in their approach. In the retrospective review, the physician's planned approach before viewing the MRI was concordant with the actual procedure 49% of the time. A different type of procedure was performed in 15% of planned injections. In such cases, the initial treatment plan was altered (ie, same procedure at a different or additional level or side) in 35% of planned injections. In the prospective data collection, 43% of injections were different from the initial physician decision. The most common reasons for altering the injection was different level affected (36%), facet pathology (22%), and different nerve root affected (16%)., Conclusions: In clinical practice, MRI before injection frequently changes management decisions in the planning and delivery of lumbar spine injections., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Toxoplasma gondii is a prevalent protozoan parasite that can infect any nucleated cell but cannot replicate outside of its host cell. Toxoplasma is auxotrophic for several nutrients including arginine, tryptophan, and purines, which it must acquire from its host cell. The demands of parasite replication rapidly deplete the host cell of these essential nutrients, yet Toxoplasma successfully manages to proliferate until it lyses the host cell. In eukaryotic cells, nutrient starvation can induce the integrated stress response (ISR) through phosphorylation of an essential translation factor eIF2. Phosphorylation of eIF2 lowers global protein synthesis coincident with preferential translation of gene transcripts involved in stress adaptation, such as that encoding the transcription factor ATF4 (CREB2), which activates genes that modulate amino acid metabolism and uptake. Here, we discovered that the ISR is induced in host cells infected with Toxoplasma. Our results show that as Toxoplasma depletes host cell arginine, the host cell phosphorylates eIF2 via protein kinase GCN2 (EIF2AK4), leading to induced ATF4. Increased ATF4 then enhances expression of the cationic amino acid transporter CAT1 (SLC7A1), resulting in increased uptake of arginine in Toxoplasma-infected cells. Deletion of host GCN2, or its downstream effectors ATF4 and CAT1, lowers arginine levels in the host, impairing proliferation of the parasite. Our findings establish that Toxoplasma usurps the host cell ISR to help secure nutrients that it needs for parasite replication., Competing Interests: The authors have declared that no competing interests exist.

Toxoplasma gondii is a ubiquitous obligate intracellular parasite that infects the nucleated cells of warm-blooded animals. From within the parasitophorous vacuole in which they reside, Toxoplasma tachyzoites secrete an arsenal of effector proteins that can reprogram host gene expression to facilitate parasite survival and replication. Gaining a better understanding of how host gene expression is altered upon infection is central for understanding parasite strategies for host invasion and for developing new parasite therapies. Here, we applied ribosome profiling coupled with mRNA measurements to concurrently study gene expression in the parasite and in host human foreskin fibroblasts. By examining the parasite transcriptome and translatome, we identified potential upstream open reading frames that may permit the stress-induced preferential translation of parasite mRNAs. We also determined that tachyzoites reduce host death-associated pathways and increase survival, proliferation, and motility in both quiescent and proliferative host cell models of infection. Additionally, proliferative cells alter their gene expression in ways that are consistent with massive transcriptional rewiring, while quiescent cells were best characterized by reentry into the cell cycle. We also identified a translational control regimen consistent with mechanistic target of rapamycin (mTOR) activation in quiescent cells and, to a lesser degree, in proliferative cells. This study illustrates the utility of the method for dissection of gene expression programs simultaneously in the parasite and host. IMPORTANCE Toxoplasma gondii is a single-celled parasite that has infected up to one-third of the world's population. Significant overhauls in gene expression in both the parasite and the host cell accompany parasite invasion, and a better understanding of these changes may lead to the development of new therapeutic agents. In this study, we employed ribosome profiling to determine the changes that occur at the levels of transcription and translation in both the parasite and the infected host cell at the same time. We discovered features of Toxoplasma mRNAs that suggest a means for controlling parasite gene expression under stressful conditions. We also show that differences in host gene expression occur depending on whether they are confluent or not. Our findings demonstrate the feasibility of using ribosomal profiling to interrogate the host-parasite dynamic under a variety of conditions., (Copyright © 2019 Holmes et al.)

Background: Protein arginine methylation is a prevalent post-translational modification. The protein arginine methyltransferase family (PRMT) is involved in many cellular processes in eukaryotes, including transcriptional regulation, epigenetic regulation, RNA metabolism, and DNA damage repair. Toxoplasma gondii, an opportunistic protozoan parasite, encodes five conserved PRMTs. PRMT5 is thought to be responsible for substantial PRMT activity in T. gondii; however, it has not yet been characterized., Methods: We tagged the 3' end of the endogenous TgPRMT5 genomic locus with sequence encoding a 3X hemagglutinin (HA) epitope. IFA and WB were performed to check the expression and subcellular localization of TgPRMT5 in tachyzoites and bradyzoites. In vitro methylation assays were performed to determine whether endogenous TgPRMT5 has arginine methyltransferase activity., Results: IFA and WB results showed that T. gondii PRMT5 (TgPRMT5) was localized in the cytoplasm in the tachyzoite stage; however, it shifts largely to the nuclear compartment in the bradyzoite stage. The in vitro methylation showed that TgPRMT5 has authentic type II PRMT activity and forms monomethylarginines and symmetric dimethylarginines., Conclusions: We determined the expression and cellular localization of TgPRMT5 in tachyzoites and bradyzoites and confirmed its type II PRMT activity. We demonstrated the major changes in expression and cellular localization of TgPRMT5 during the tachyzoite and bradyzoite stages in T. gondii. Our findings suggest that TgPRMT5 protein may be involved in tachyzoite-bradyzoite transformation.

Toxoplasma gondii is an intracellular parasite that has infected one-third of humans. The infection is permanent because the replicative form (tachyzoite) converts into a latent tissue cyst form (bradyzoite) that evades host immunity and is impervious to current drugs. The continued presence of these parasitic cysts hinders treatment and leads to chronic infection that has been linked to behavioral changes in rodents and neurological disease in humans. How these behavioral changes occur, and whether they are due to parasite manipulation or the host response to infection, remains an outstanding question. We previously showed that guanabenz possesses antiparasitic activity; here, we show that guanabenz reproducibly lowers brain cyst burden up to 80% in chronically infected male and female BALB/cJ mice when given intraperitoneally but not when administered by gavage or in food. Regardless of the administration route, guanabenz reverses Toxoplasma -induced hyperactivity in latently infected mice. In contrast, guanabenz increases cyst burden when given to chronically infected C57BL/6J mice yet still reverses Toxoplasma -induced hyperactivity. Examination of the brains from chronically infected BALB/cJ and C57BL/6J mice shows that guanabenz decreases inflammation and perivascular cuffing in each strain. Our study establishes a robust model for cyst reduction in BALB/cJ mice and shows for the first time that it is possible to reverse a key behavioral change associated with latent toxoplasmosis. The rescue from parasite-induced hyperactivity correlates with a decrease in neuroinflammation rather than reduced cyst counts, suggesting that some behavioral changes arise from host responses to infection. IMPORTANCE Toxoplasma gondii is a common parasite of animals, including up to one-third of humans. The single-celled parasite persists within hosts for the duration of their life as tissue cysts, giving rise to chronic infection. Latent toxoplasmosis is correlated with neurological dysfunction in humans and results in dramatic behavioral changes in rodents. When infected, mice and rats adapt behaviors that make them more likely to be devoured by cats, the only host that supports the sexual stage of the parasite. In this study, we establish a new mouse model of tissue cyst depletion using a drug called guanabenz and show that it is possible to reverse a key behavior change back to normal in infected animals. We also show that the mechanism appears to have nothing to do with parasite cyst burden but rather the degree of neuroinflammation produced by chronic infection., (Copyright © 2019 Martynowicz et al.)

Toxoplasma gondii is an apicomplexan protozoan parasite with a complex life cycle composed of multiple stages that infect mammals and birds. Tachyzoites rapidly replicate within host cells to produce acute infection during which the parasite disseminates to tissues and organs. Highly replicative cells are subject to Double Strand Breaks (DSBs) by replication fork collapse and ATM, a member of the phosphatidylinositol 3-kinase (PI3K) family, is a key factor that initiates DNA repair and activates cell cycle checkpoints. Here we demonstrate that the treatment of intracellular tachyzoites with the PI3K inhibitor caffeine or ATM kinase-inhibitor KU-55933 affects parasite replication rate in a dose-dependent manner. KU-55933 affects intracellular tachyzoite growth and induces G1-phase arrest. Addition of KU-55933 to extracellular tachyzoites also leads to a significant reduction of tachyzoite replication upon infection of host cells. ATM kinase phosphorylates H2A.X (γH2AX) to promote DSB damage repair. The level of γH2AX increases in tachyzoites treated with camptothecin (CPT), a drug that generates fork collapse, but this increase was not observed when co-administered with KU-55933. These findings support that KU-55933 is affecting the Toxoplasma ATM-like kinase (TgATM). The combination of KU-55933 and other DNA damaging agents such as methyl methane sulfonate (MMS) and CPT produce a synergic effect, suggesting that TgATM kinase inhibition sensitizes the parasite to damaged DNA. By contrast, hydroxyurea (HU) did not further inhibit tachyzoite replication when combined with KU-55933.

Dynamin-related proteins (Drps) are involved in diverse processes such as organelle division and vesicle trafficking. The intracellular parasite Toxoplasma gondii possesses three distinct Drps. TgDrpC, whose function remains unresolved, is unusual in that it lacks a conserved GTPase Effector Domain, which is typically required for function. Here, we show that TgDrpC localizes to cytoplasmic puncta; however, in dividing parasites, TgDrpC redistributes to the growing edge of the daughter cells. By conditional knockdown, we determined that loss of TgDrpC stalls division and leads to rapid deterioration of multiple organelles and the IMC. We also show that TgDrpC interacts with proteins that exhibit homology to those involved in vesicle transport, including members of the adaptor complex 2. Two of these proteins, a homolog of the adaptor protein 2 (AP-2) complex subunit alpha-1 and a homolog of the ezrin-radixin-moesin (ERM) family proteins, localize to puncta and associate with the daughter cells. Consistent with the association with vesicle transport proteins, re-distribution of TgDrpC to the IMC during division is dependent on post-Golgi trafficking. Together, these results support that TgDrpC contributes to vesicle trafficking and is critical for stability of parasite organelles and division., (© 2018 John Wiley & Sons Ltd.)

Background: Physical activity remains low and nearly unchanged from preoperative levels following total knee arthroplasty (TKA), and this is thought to underlie long-term functional limitations, secondary health problems, and higher health care costs after TKA., Objective: Our objective is to determine whether a telehealth-based intervention could improve physical activity and functional outcomes after TKA., Design: The design is a 2-arm, parallel, assessor-blinded, randomized controlled trial with baseline, midintervention, postintervention, and 6-month follow-up assessments., Setting: The setting is one academic medical center and one Veterans Affairs health care system., Participants: One hundred US military veterans (aged 50-85 years) scheduled for unilateral TKA will participate in this study., Intervention: The telehealth-based intervention to change physical activity behavior will be delivered through 10 sessions each of 30 minutes over a 12-week period. Participants will be provided with a wearable physical activity monitor to receive feedback on step count and guide goal-setting. Control participants will receive telehealth-based education on nonbehavioral aspects of health for the same frequency and duration as the intervention group. For both groups, telehealth sessions will occur concurrently with standardized outpatient rehabilitation., Measurements: The primary outcome will be change in physical activity, assessed as daily step counts measured using an accelerometer-based sensor. Secondary outcomes will be measured using the Life-Space Assessment questionnaire and change in physical function (30-Second Chair-Stand Test, Timed "Up & Go" Test, Six-Minute Walk Test, Western Ontario and McMaster Universities Osteoarthritis Index, and Veterans RAND 12-Item Health Survey)., Limitations: Participant and interventionist blinding is not possible., Conclusions: This trial will assess the efficacy of a novel behavior-change intervention to improve physical activity and physical function in patients after TKA. Effective physical activity behavior change could provide clinicians with a technique to augment current practice and resolve poor physical activity outcomes, long-term health problems, and high costs following TKA.

Toxoplasma gondii is an obligate intracellular parasite that has infected one-third of the population. Upon infection of warm-blooded vertebrates, the replicating form of the parasite (tachyzoite) converts into a latent form (bradyzoite) present in tissue cysts. During immune deficiency, bradyzoites can reconvert into tachyzoites and cause life-threatening toxoplasmosis. We previously reported that translational control through phosphorylation of the α subunit of T. gondii eukaryotic initiation factor 2 (eIF2α) (TgIF2α) is a critical component of the parasite stress response. Diverse stresses can induce the conversion of tachyzoites to bradyzoites, including those disrupting the parasite's endoplasmic reticulum (ER) (ER stress). Toxoplasma possesses four eIF2α kinases, one of which (TgIF2K-A) localizes to the parasite ER analogously to protein kinase R-like endoplasmic reticulum kinase (PERK), the eIF2α kinase that responds to ER stress in mammalian cells. Here, we investigated the effects of a PERK inhibitor (PERKi) on Toxoplasma Our results show that the PERKi GSK2606414 blocks the enzymatic activity of TgIF2K-A and reduces TgIF2α phosphorylation specifically in response to ER stress. PERKi also significantly impeded multiple steps of the tachyzoite lytic cycle and sharply lowered the frequency of bradyzoite differentiation in vitro Pretreatment of host cells with PERKi prior to infection did not affect parasite infectivity, and PERKi still impaired parasite replication in host cells lacking PERK. In mice, PERKi conferred modest protection from a lethal dose of Toxoplasma Our findings represent the first pharmacological evidence supporting TgIF2K-A as an attractive new target for the treatment of toxoplasmosis., (Copyright © 2018 American Society for Microbiology.)

The metabolic state of a cell is influenced by cell-extrinsic factors, including nutrient availability and growth factor signaling. Here, we present extracellular matrix (ECM) remodeling as another fundamental node of cell-extrinsic metabolic regulation. Unbiased analysis of glycolytic drivers identified the hyaluronan-mediated motility receptor as being among the most highly correlated with glycolysis in cancer. Confirming a mechanistic link between the ECM component hyaluronan and metabolism, treatment of cells and xenografts with hyaluronidase triggers a robust increase in glycolysis. This is largely achieved through rapid receptor tyrosine kinase-mediated induction of the mRNA decay factor ZFP36, which targets TXNIP transcripts for degradation. Because TXNIP promotes internalization of the glucose transporter GLUT1, its acute decline enriches GLUT1 at the plasma membrane. Functionally, induction of glycolysis by hyaluronidase is required for concomitant acceleration of cell migration. This interconnection between ECM remodeling and metabolism is exhibited in dynamic tissue states, including tumorigenesis and embryogenesis., (Copyright © 2018 Elsevier Inc. All rights reserved.)