GRAS1 non-coding RNA protects against DNA damage and cell death by binding and stabilizing NKAP.

Non-coding RNA (ncRNA) gene products are involved in diverse biological processes including splicing, epigenetic regulation, gene expression, proliferation, and metabolism. The biological mechanisms by which ncRNAs contribute to cell survival remain poorly understood. We found that the Growth Regulator Antisense 1 (GRAS1) long non-coding RNA (lncRNA) transcript promotes growth in multiple human cell types by protecting against DNA damage. Knockdown of GRAS1 induced DNA damage and cell death, along with significant expression changes in DNA damage response, intrinsic apoptotic signaling, and cellular response to environmental stimulus genes. Extensive DNA damage occurred after GRAS1 knockdown, with numerous double strand breaks occurring in each cell. The number of cells undergoing apoptosis and with fragmented nuclei increased significantly after GRAS1 knockdown. We used RNA antisense purification and mass spectrometry (RAP-MS) to identify the NF-κB activating protein (NKAP) as a direct protein interaction partner of GRAS1 lncRNA. NKAP protein was degraded after GRAS1 knockdown, in a proteasome-dependent manner. Overexpression of GRAS1 or NKAP mitigated the DNA damage effects of GRAS1 knockdown. In summary, GRAS1 and NKAP directly interact to protect against DNA damage and cell death in multiple human cell lines.

Adult Organophosphate and Carbamate Insecticide Exposure and Sperm Concentration: A Systematic Review and Meta-Analysis of the Epidemiological Evidence.

BACKGROUND: Evidence of the negative impacts of contemporary use insecticides on sperm concentration has increased over the last few decades; however, meta-analyses on this topic are rare. OBJECTIVES: This investigation assessed the qualitative and quantitative strength of epidemiological evidence regarding adult exposure to two classes of contemporary use insecticides-organophosphates (OPs) and N-methyl carbamates (NMCs)-and sperm concentration using robust and reproducible systematic review and meta-analysis methods. METHODS: Three scientific databases (PubMed, Scopus, and Web of Science), two U.S. government databases (NIOSHTIC-2 and Science.gov), and five nongovernmental organization websites were searched for relevant primary epidemiological studies published in any language through 11 August 2022. Risk of bias and strength of evidence were evaluated according to Navigation Guide systematic review methodology. Bias-adjusted standardized mean difference effect sizes were calculated and pooled using a three-level, multivariate random-effect meta-analysis model with cluster-robust variance estimation. RESULTS: Across 20 studies, 21 study populations, 42 effect sizes, and 1,774 adult men, the pooled bias-adjusted standardized mean difference in sperm concentration between adult men more- and less-exposed to OP and NMC insecticides was -0.30 (95% CI: -0.49, -0.10; PSatt<0.01). Sensitivity and subgroup analyses explored statistical heterogeneity and validated the model robustness. Although the pooled effect estimate was modified by risk of bias, insecticide class, exposure setting, and recruitment setting, it remained negative in direction across all meta-analyses. The body of evidence was rated to be of moderate quality, with sufficient evidence of an association between higher adult OP and NMC insecticide exposure and lower sperm concentration. DISCUSSION: This comprehensive investigation found sufficient evidence of an association between higher OP and NMC insecticide exposure and lower sperm concentration in adults. Although additional cohort studies can be beneficial to fill data gaps, the strength of evidence warrants reducing exposure to OP and NMC insecticides now to prevent continued male reproductive harm. https://doi.org/10.1289/EHP12678.

Endoscopic sexing in turtles and tortoises: 467 cases (2007-2017).

BACKGROUND: This multi-institutional retrospective study evaluated the feasibility and safety of endoscopic sex identification in 467 turtles and tortoises, representing 10 species. METHODS: Medical records of turtles and tortoises that underwent endoscopic sex identification at the University of Georgia, New England Aquarium and Turtle Conservancy were reviewed for presurgical management, anaesthesia, endoscopic equipment and surgical techniques, endoscopic results and complications. RESULTS: The majority of animals weighed less than 200 g, were fasted and anaesthetised using an injectable combination of ketamine, dexmedetomidine and morphine or hydromorphone, supplemented by local lidocaine at the prefemoral site. Anaesthetic reversal using atipamezole alone or in combination with naloxone was routine. For uncomplicated procedures, mean total anaesthesia, surgery and recovery times were 22, 4 and 18 minutes, respectively. All animals were placed in lateral recumbency for a prefemoral endoscopic approach to the coelom using a rigid telescope and sterile fluid infusion to visualise the gonads. Sex identification was definitive in 99.4% (n = 464) of the animals. Iatrogenic bladder perforation was the most common complication (n = 5), which necessitated extended anaesthesia and surgical time for repair. Only a single anaesthetic-related death was reported, which was associated with human error and drug overdose. CONCLUSIONS: This is the first large-scale study to retrospectively evaluate endoscopic sex identification in multiple chelonian species. Results suggest that endoscopic sexing is a safe, accurate and practical means for sex identification in turtles and tortoises, and represents a valuable tool in their reproductive management.

Evaluation of the frailty characteristics and clinical outcomes according to the new frailty-based outcome prediction model (Myeloma Risk Profile-MRP) in a UK real-world cohort of elderly newly diagnosed Myeloma patients.

The management of myeloma in the elderly is shifting its focus towards reducing the risk of under-treating fit patients and the risk of over-treating frail patients. Frailty assessment is required in this patient group in order to individualise treatment decisions. In addition to the proven prognostic values of the International Myeloma Working Group (IMWG) frailty score and the revised Myeloma Co-morbidity Index (R-MCI), a new easy-to-use frailty-based risk profile score (high-risk (i.e. frail), medium risk (i.e. intermediate-fitness) and low-risk (i.e. fit)) named Myeloma Risk Profile (MRP) was shown to be predictive of survival in the clinical trial setting. In this retrospective real-world study, we set out to evaluate the frailty characteristics and clinical outcomes according to the different MRP scoring algorithm categories (frail vs. intermediate vs fit), in a high risk cohort of elderly newly diagnosed myeloma patients treated with the fixed-duration triplet therapy VCD (bortezomib with cyclophosphamide and dexamethasone). Clinical outcomes included: reason for treatment discontinuation, overall response rate (ORR), overall survival (OS), progression-free survival (PFS), and adverse events (AEs). Out of 100 patients, 62 were frail, 27 were intermediate and 11 were fit, according to MRP scores. To enable meaningful comparisons between comparable numbers, subgroups analyses for ORR, OS, PFS, and AEs focused on frail (n = 62) versus intermediate or fit (n = 38) patients. The proportion of patients in each subgroup who were able to complete the planned course of treatment was (frail: 43.5% vs. intermediate or fit: 55.3%). A higher proportion in the frail subgroup discontinued therapy due to progressive disease (19.4% vs. 2.6%). Discontinuation due to toxicity was comparable across subgroups (14.5% vs. 15.8%), ORR in the total cohort was 75%, and this was comparable between subgroups (frail: 74.2% vs. intermediate or fit: 76.3%). There was a trend for a shorter median OS in the frail subgroup but without a statistical significance: (frail vs. intermediate or fit): (46 months vs. not reached, HR: 1.94, 95% CI 0.89-4.2, p = 0.094). There was no difference in median PFS between subgroups: (frail vs. intermediate or fit): (11.8 vs. 9.9 months, HR: 0.99, 95% CI: 0.61-1.61, P = 0.982). This cohort demonstrated a higher incidence rate of AEs in frail patients compared to those in the intermediate or fit group: patients with at least one any grade toxicity (85.5% vs. 71.1%), patients with at least one ≥G3 AE (37.1% vs. 21.1%). In conclusion, our study is to the first to evaluate clinical outcomes according to MRP in a high risk real-world cohort of patients treated exclusively with the proteasome inhibitor-based VCD therapy. Our study demonstrated a trend for worse OS in addition to worse AE outcomes in the frail group, but no difference in PFS with this fixed-duration therapy. MRP is an easy-to-use tool in clinical practice; its prognostic value was validated in the real-world in a large cohort of patients from the Danish Registry. Further evaluation of MRP in the real-world when continuous therapies are used, can further support the generalisability of its prognostic value in elderly myeloma patients.

Prognostic value of epicardial adipose tissue volume in combination with coronary plaque and flow assessment for the prediction of major adverse cardiac events.

PURPOSE: The purpose of this study was to determine whether EAT volume in combination with coronary CT angiography (CCTA)-derived plaque quantification and CT-derived fractional flow reserve (CT-FFR) has prognostic implication with major adverse cardiac events (MACE). METHODS: Patients (n = 117, 58 ± 10 years, 61% male) who had previously undergone invasive coronary angiography (ICA) and CCTA were retrospectively analyzed. Follow-up was performed to record MACE. EAT volume and plaque measures were derived from non-contrast and contrast-enhanced CT images using a semi-automatic software approach, while CT-FFR was calculated using a machine-learning algorithm. The diagnostic performance to identify MACE was evaluated using univariable and multivariable Cox proportional hazards analysis and concordance (C)-indices. RESULTS: During a median follow-up period of 40.4 months, 19 events were registered. EAT volume, CCTA ≥ 50% stenosis, and CT-FFR were significantly different in patients developing MACE (all p < 0.05). The following parameters were predictors of MACE in adjusted multivariable Cox regression analysis (hazard ratio [HR]): EAT volume (HR 2.21, p = 0.023), indexed EAT volume (HR 2.03, p = 0.035), and CCTA ≥ 50% (HR 1.05, p = 0.048). A model including Morise score, CCTA ≥ 50% stenosis, and EAT volume showed significantly improved C-index to Morise score alone (AUC 0.83 vs. 0.66, p = 0.004). CONCLUSIONS: Facing limitations in conventional cardiovascular risk scoring models, this observational study demonstrates that the prediction performance of our proposed method achieves a significant improvement in prognostic ability, especially when compared to models such as Morise score alone or its combination with CCTA and CT-FFR.

Additive value of epicardial adipose tissue quantification to coronary CT angiography-derived plaque characterization and CT fractional flow reserve for the prediction of lesion-specific ischemia.

OBJECTIVES: Epicardial adipose tissue (EAT) from coronary CT angiography (CCTA) is strongly associated with coronary artery disease (CAD). We investigated the additive value of EAT volume to coronary plaque quantification and CT-derived fractional flow reserve (CT-FFR) to predict lesion-specific ischemia. METHODS: Patients (n = 128, 60.6 ± 10.5 years, 61% male) with suspected CAD who had undergone invasive coronary angiography (ICA) and CCTA were retrospectively analyzed. EAT volume and plaque measures were derived from CCTA using a semi-automatic software approach, while CT-FFR was calculated using a machine learning algorithm. The predictive value and discriminatory power of EAT volume, plaque measures, and CT-FFR to identify ischemic CAD were assessed using invasive FFR as the reference standard. RESULTS: Fifty-five of 152 lesions showed ischemic CAD by invasive FFR. EAT volume, CCTA ≥ 50% stenosis, and CT-FFR were significantly different in lesions with and without hemodynamic significance (all p < 0.05). Multivariate analysis revealed predictive value for lesion-specific ischemia of these parameters: EAT volume (OR 2.93, p = 0.021), CCTA ≥ 50% (OR 4.56, p = 0.002), and CT-FFR (OR 6.74, p < 0.001). ROC analysis demonstrated incremental discriminatory value with the addition of EAT volume to plaque measures alone (AUC 0.84 vs. 0.62, p < 0.05). CT-FFR (AUC 0.89) showed slightly superior performance over EAT volume with plaque measures (AUC 0.84), however without significant difference (p > 0.05). CONCLUSIONS: EAT volume is significantly associated with ischemic CAD. The combination of EAT volume with plaque quantification demonstrates a predictive value for lesion-specific ischemia similar to that of CT-FFR. Thus, EAT may aid in the identification of hemodynamically significant coronary stenosis. KEY POINTS: • CT-derived EAT volume quantification demonstrates high discriminatory power to identify lesion-specific ischemia. • EAT volume shows incremental diagnostic performance over CCTA-derived plaque measures in detecting lesion-specific ischemia. • A combination of plaque measures with EAT volume provides a similar discriminatory value for detecting lesion-specific ischemia compared to CT-FFR.

News Frames and News Exposure Predicting Flu Vaccination Uptake: Evidence from U.S. Newspapers, 2011-2018 Using Computational Methods.

Flu remains a prevalent public health issue and the flu vaccine uptake remains low. This study examined the behavioral consequences of first- and second-level agenda setting by testing the effects of exposure to flu-related news coverage and different frames on flu vaccine uptake, while controlling for confounding sociodemographic factors, utilizing computational methods. Findings suggested that the media effects on children and adult influenza vaccine uptake differed. The frame of influenza caused deaths/illnesses stories was the only media frame that positively predicted flu vaccine uptake among children. Four frames, including influenza caused deaths/illnesses stories, flu prevention tips, things to know about flu vaccine, and public accountability positively predicted flu vaccine uptake among adults. Exposure rate negatively predicted flu vaccine uptake among adults. A few sociodemographic factors impacted flu vaccine uptake. Results also revealed the most and least exposed frames and state exposure rates to flu-related news stories.

Quiescent-Interval Slice-Selective MRA Accurately Estimates Intravascular Stent Dimensions Prior to Intervention in Patients With Peripheral Artery Disease.

BACKGROUND: Quiescent-interval slice-selective (QISS) magnetic resonance angiography (MRA) is a non-contrast alternative for the pre-procedural assessment of patients with peripheral artery disease (PAD). However, the feasibility of pre-procedural stent size estimation using QISS MRA would merit investigation. PURPOSE: To evaluate the feasibility of QISS MRA for pre-procedural stent size estimation in PAD patients compared to computed tomography angiography (CTA). STUDY TYPE: Retrospective. SUBJECTS: Thirty-three PAD patients (68 ± 9 years, 18 men, 15 women). FIELD STRENGTH/SEQUENCE: Two-dimensional balanced steady-state free precession QISS MRA at 1.5 T and 3 T. ASSESSMENT: All patients received QISS MRA and CTA of the lower extremity run-off followed by interventional digital subtraction angiography (DSA). Stenotic lesion length and diameter were quantified (AMF and AVS with 3 and 13 years of experience in cardiovascular imaging, respectively) to estimate the dimensions of the stent necessary to restore blood flow in the treated arteries. Measured dimensions were adjusted to the closest stent size available. STATISTICAL TESTS: The Friedman test with subsequent pairwise Wilcoxon signed-rank test was used to compare the estimated stent dimensions between QISS MRA, CTA, and the physical stent size used for intervention. Intra-class correlation (ICC) analysis was performed to assess inter-reader agreement. Significant differences were considered at P < 0.05. RESULTS: No significant difference was observed between estimated stent diameter by QISS MRA or CTA compared to physical stent diameter (8.9 ± 2.9 mm, 8.8 ± 3.0 mm, and 8.8 ± 3.8 mm, respectively; χ2  = 1.45, P = 0.483). There was a significant underestimation of stent length for both QISS MRA and CTA, compared to physical stent length (45.8 ± 27.8 mm, 46.4 ± 29.3 mm, and 50.4 ± 34.0 mm, respectively; χ2  = 11.96) which could be corrected when measurements were adjusted to the next available stent length (χ2  = 2.38, P = 0.303). Inter-reader assessment showed good to excellent agreement between the readers (all ICC ≥0.81). DATA CONCLUSION: QISS MRA represents a reliable method for pre-procedural lesion assessment and stent diameter and length estimation in PAD patients. LEVEL OF EVIDENCE: 3 TECHNICAL EFFICACY: Stage 2.

Serologic Responses following a Single Dose of SARS-Cov-2 Vaccination in Allogeneic Stem Cell Transplantation Recipients.

Immunocompromised individuals were not included in formal trials of SARS-CoV-2 mRNA vaccines. Subsequent studies in patients with hematologic malignancies and solid organ transplantation recipients suggest inferior responses to vaccination. We determined antibody responses to a single dose of vaccines in one of the most vulnerable patient groups, allogeneic hematopoietic cell transplantation (allo-HCT) recipients. Pfizer-BioNTech (PB) or AstraZeneca (AZ) SARS-CoV-2 vaccines were administered at least 3 months post-transplantation to 55 adult allo-HCT recipients. We found that older age and concurrent use of immunosuppressive medications were significantly associated with lack of antibody response to vaccination. Only 21% of patients on systemic immunosuppression mounted a response, compared with 58% of patients not on immunosuppression (P = .006). We also show that responses to the AZ vaccine may be superior to responses to the PB vaccine in this cohort. These findings highlight the need for novel immunogenic vaccine formulations and schedules in these highest-risk patients, as well as continued public healthy safety measures to protect the most vulnerable members of our society.

Efficacy and tolerability of VCD chemotherapy in a UK real-world dataset of elderly transplant-ineligible newly diagnosed myeloma patients.

OBJECTIVE: There are limited data on the efficacy and tolerability of VCD chemotherapy in transplant-non-eligible (TNE) newly diagnosed myeloma (NDMM) patients. In this retrospective study, we set out to evaluate this triplet combination in this setting across Thames Valley Cancer Network (UK). METHODS: The primary end point was overall response rate (ORR). Secondary outcomes included event-free survival (EFS), overall survival (OS) and adverse events (AEs). RESULTS: In a total cohort of 158 patients, ORR for total cohort was 72.1%. Median EFS was 10.5 months, and for subgroups by age (<75:11.7 vs ≥75:10.3 months, P = .124), by Charlson Co-morbidity Index (CCI) (<5:11.1 vs ≥5:8.2 months, P = .345). The 4-month landmark analysis showed the following median EFS results: by cumulative bortezomib dose (≥26 mg/m2 : 9.0 months vs <26 mg/m2 : 6.4, P = .13), by cumulative cyclophosphamide dose (≥7000 mg: 9.2 vs <7000 mg: 7.0 months, P = .02) and by cumulative dexamethasone dose (>600 mg: 7.8 vs ≤600 mg: 8.3 months, P = .665). Median OS was 46.9 months. The incidence rate of AE was as follows: any grade (76.8%), ≥G3 (27.1%), ≥G3 haematological AEs (7.9%), any grade infections (31.1%) and ≥G3 infections (11.9%). CONCLUSION: This study demonstrated a good ORR achieved from fixed duration VCD, which was reasonably well tolerated. This was followed by modest median EFS. We envisage that the latter may be improved in this patient group with the use of a higher cumulative bortezomib dose (≥26 mg/m2 ) which showed a trend for improved EFS although without statistical significance (P = .13), and with the use of a higher cumulative cyclophosphamide doses (≥7000 mg, P = .02), subject to tolerability and close monitoring.

Draft Genome Sequences of Four Bacterial Species as Part of an Experiential Microbiology Project at SUNY Geneseo.

We report four draft genome sequences related to the genera Bacillus and Escherichia, recovered from surfaces associated with human interaction, and Sediminibacterium, recovered from an aquatic environment. This study was part of an undergraduate microbial bioinformatics course at the State University of New York at Geneseo.

Pronounced Therapeutic Benefit of a Single Bidirectional AAV Vector Administered Systemically in Sandhoff Mice.

The GM2 gangliosidoses, Tay-Sachs disease (TSD) and Sandhoff disease (SD), are fatal lysosomal storage disorders caused by mutations in the HEXA and HEXB genes, respectively. These mutations cause dysfunction of the lysosomal enzyme ß-N-acetylhexosaminidase A (HexA) and accumulation of GM2 ganglioside (GM2) with ensuing neurodegeneration, and death by 5 years of age. Until recently, the most successful therapy was achieved by intracranial co-delivery of monocistronic adeno-associated viral (AAV) vectors encoding Hex alpha and beta-subunits in animal models of SD. The blood-brain barrier crossing properties of AAV9 enables systemic gene therapy; however, the requirement of co-delivery of two monocistronic AAV vectors to overexpress the heterodimeric HexA protein has prevented the use of this approach. To address this need, we developed multiple AAV constructs encoding simultaneously HEXA and HEXB using AAV9 and AAV-PHP.B and tested their therapeutic efficacy in 4- to 6-week-old SD mice after systemic administration. Survival and biochemical outcomes revealed superiority of the AAV vector design using a bidirectional CBA promoter with equivalent dose-dependent outcomes for both capsids. AAV-treated mice performed normally in tests of motor function, CNS GM2 ganglioside levels were significantly reduced, and survival increased by >4-fold with some animals surviving past 2 years of age.

7T MRI Predicts Amelioration of Neurodegeneration in the Brain after AAV Gene Therapy.

GM1 gangliosidosis (GM1) is a fatal neurodegenerative lysosomal storage disease that occurs most commonly in young children, with no effective treatment available. Long-term follow-up of GM1 cats treated by bilateral thalamic and deep cerebellar nuclei (DCN) injection of adeno-associated virus (AAV)-mediated gene therapy has increased lifespan to 8 years of age, compared with an untreated lifespan of ~8 months. Due to risks associated with cerebellar injection in humans, the lateral ventricle was tested as a replacement route to deliver an AAVrh8 vector expressing feline ß-galactosidase (ß-gal), the defective enzyme in GM1. Treatment via the thalamus and lateral ventricle corrected storage, myelination, astrogliosis, and neuronal morphology in areas where ß-gal was effectively delivered. Oligodendrocyte number increased, but only in areas where myelination was corrected. Reduced AAV and ß-gal distribution were noted in the cerebellum with subsequent increases in storage, demyelination, astrogliosis, and neuronal degeneration. These postmortem findings were correlated with endpoint MRI and magnetic resonance spectroscopy (MRS). Compared with the moderate dose with which most cats were treated, a higher AAV dose produced superior survival, currently 6.5 years. Thus, MRI and MRS can predict therapeutic efficacy of AAV gene therapy and non-invasively monitor cellular events within the GM1 brain.

The Easier the Better? Comparing the Readability and Engagement of Online Pro- and Anti-Vaccination Articles.

Online anti-vaccine articles contribute to the anti-vaccine movement, which leads to recent outbreaks of vaccine-preventable diseases. Previous studies indicate that anti-vaccine articles are easy to read and understand, which may increase their abilities to engage viewers. The present study aims to examine if readability levels are related to engagement. Using combination of terms to search for vaccine articles in Google in May 2017, this study examined 541 pro-vaccine online articles with a total of 508,571 words and 382 anti-vaccine articles with a total of 843,805 words. Almost all vaccine articles exceeded the American average reading comprehension level. No significant difference in readability was found between pro- and anti-vaccine articles. Pro-vaccine articles that could only be understood by college graduates were less engaging than those with lower readability levels. No significant relationship between anti-vaccine articles' readability and engagement was discovered. Different vaccine topics had different readability and engagement levels, which implied that certain combinations of themes and readability levels could enhance the health messages' persuasion effect. Recommendations for designing effective health messages are provided.

Developmental phosphoproteomics identifies the kinase CK2 as a driver of Hedgehog signaling and a therapeutic target in medulloblastoma.

A major limitation of targeted cancer therapy is the rapid emergence of drug resistance, which often arises through mutations at or downstream of the drug target or through intrinsic resistance of subpopulations of tumor cells. Medulloblastoma (MB), the most common pediatric brain tumor, is no exception, and MBs that are driven by sonic hedgehog (SHH) signaling are particularly aggressive and drug-resistant. To find new drug targets and therapeutics for MB that may be less susceptible to common resistance mechanisms, we used a developmental phosphoproteomics approach in murine granule neuron precursors (GNPs), the developmental cell of origin of MB. The protein kinase CK2 emerged as a driver of hundreds of phosphorylation events during the proliferative, MB-like stage of GNP growth, including the phosphorylation of three of the eight proteins commonly amplified in MB. CK2 was critical to the stabilization and activity of the transcription factor GLI2, a late downstream effector in SHH signaling. CK2 inhibitors decreased the viability of primary SHH-type MB patient cells in culture and blocked the growth of murine MB tumors that were resistant to currently available Hh inhibitors, thereby extending the survival of tumor-bearing mice. Because of structural interactions, one CK2 inhibitor (CX-4945) inhibited both wild-type and mutant CK2, indicating that this drug may avoid at least one common mode of acquired resistance. These findings suggest that CK2 inhibitors may be effective for treating patients with MB and show how phosphoproteomics may be used to gain insight into developmental biology and pathology.

Acquired Protective Immunity in Atlantic Salmon Salmo salar against the Myxozoan Kudoa thyrsites Involves Induction of MHIIß+ CD83+ Antigen-Presenting Cells.

The histozoic myxozoan parasite Kudoa thyrsites causes postmortem myoliquefaction and is responsible for economic losses to salmon aquaculture in the Pacific Northwest. Despite its importance, little is known about the host-parasite relationship, including the host response to infection. The present work sought to characterize the immune response in Atlantic salmon during infection, recovery, and reexposure to K. thyrsites After exposure to infective seawater, infected and uninfected smolts were sampled three times over 4,275 degree-days. Histological analysis revealed infection severity decreased over time in exposed fish, while in controls there was no evidence of infection. Following a secondary exposure of all fish, severity of infection in the controls was similar to that measured in exposed fish at the first sampling time but was significantly reduced in reexposed fish, suggesting the acquisition of protective immunity. Using immunohistochemistry, we detected a population of MHIIß+ cells in infected muscle that followed a pattern of abundance concordant with parasite prevalence. Infiltration of these cells into infected myocytes preceded destruction of the plasmodium and dissemination of myxospores. Dual labeling indicated a majority of these cells were CD83+/MHIIß+ Using reverse transcription-quantitative PCR, we detected significant induction of cellular effectors, including macrophage/dendritic cells (mhii/cd83/mcsf), B cells (igm/igt), and cytotoxic T cells (cd8/nkl), in the musculature of infected fish. These data support a role for cellular effectors such as antigen-presenting cells (monocyte/macrophage and dendritic cells) along with B and T cells in the acquired protective immune response of Atlantic salmon against K. thyrsites.

Adeno-Associated Virus Gene Therapy in a Sheep Model of Tay-Sachs Disease.

Tay-Sachs disease (TSD) is a fatal neurodegenerative disorder caused by a deficiency of the enzyme hexosaminidase A (HexA). TSD also occurs in sheep, the only experimental model of TSD that has clinical signs of disease. The natural history of sheep TSD was characterized using serial neurological evaluations, 7 Tesla magnetic resonance imaging, echocardiograms, electrodiagnostics, and cerebrospinal fluid biomarkers. Intracranial gene therapy was also tested using AAVrh8 monocistronic vectors encoding the α-subunit of Hex (TSD α) or a mixture of two vectors encoding both the α and ß subunits separately (TSD α + ß) injected at high (1.3 × 1013 vector genomes) or low (4.2 × 1012 vector genomes) dose. Delay of symptom onset and/or reduction of acquired symptoms were noted in all adeno-associated virus-treated sheep. Postmortem evaluation showed superior HexA and vector genome distribution in the brain of TSD α + ß sheep compared to TSD α sheep, but spinal cord distribution was low in all groups. Isozyme analysis showed superior HexA formation after treatment with both vectors (TSD α + ß), and ganglioside clearance was most widespread in the TSD α + ß high-dose sheep. Microglial activation and proliferation in TSD sheep-most prominent in the cerebrum-were attenuated after gene therapy. This report demonstrates therapeutic efficacy for TSD in the sheep brain, which is on the same order of magnitude as a child's brain.