Typing of Treponema pallidum in a Brazilian sample and follow-up of treatment using molecular assays.

Syphilis remains a significant public health concern, with serological assays being the primary method for diagnosis. However, molecular techniques have proven to be reliable tools for the diagnosis and understanding of the transmission dynamics of Treponema pallidum infection. This study aimed to evaluate the efficacy of syphilis treatment using molecular assays, perform Enhanced Centers for Disease Control and Prevention (ECDC) typing, and analyze resistance (macrolide and doxycycline) in the T. pallidum isolate. PCR assay amplified treponemal DNA only from the lesion sample, whereas qPCR was able to amplify DNA in both lesion and blood samples before treatment. Throughout the treatment follow-up, qPCR effectively did not identify treponemal DNA in the blood for up to one to two weeks after treatment. ECDC typing revealed the genotype 14 e/g in the Brazilian T. pallidum isolate, and the presence of the A2058G mutation in 23 S rRNA gene, indicating macrolide resistance. Although, the G1058C mutation in 16 S rRNA gene was not detected. Notably, qPCR demonstrated its potential for diagnosing T. pallidum in blood samples, even when the treponemal DNA levels were low, enabling more accurate and sensitive diagnosis and guiding better syphilis therapy. In addition, to the best of our knowledge, this study represents the first identification of subtype 14 e/g and azithromycin resistance in a Brazilian T. pallidum isolate.

Soil greenhouse gas emissions from dead and natural mangrove forests in Southeastern Brazil.

Mangroves forests may be important sinks of carbon in coastal areas but upon their death, these forests may become net sources of carbon dioxide (CO2) and methane (CH4) to the atmosphere. Here we assessed the spatial and temporal variability in soil CO2 and CH4 fluxes from dead mangrove forests and paired intact sites in SE-Brazil. Our findings demonstrated that during warmer and drier conditions, CO2 soil flux was 183 % higher in live mangrove forests when compared to the dead mangrove forests. Soil CH4 emissions in live forests were > 1.4-fold higher than the global mangrove average. During the wet season, soil GHG emissions dropped significantly at all sites. During warmer conditions, mangroves were net sources of GHG, with a potential warming effect (GWP100) of 32.9 ± 10.2 (±SE) Mg CO2e ha-1 y-1. Overall, we found that dead mangroves did not release great amounts of GHG after three years of forest loss.

Molecular characterization of Treponema pallidum isolates from Brazil.

Syphilis remains a public health concern in Brazil, and the data on the characterization and resistance of Treponema pallidum in Brazil is limited. The present study aimed to detect Treponema DNA in the lesions and blood samples obtained from individuals diagnosed with syphilis. The Brazilian isolates were submitted to the Enhanced Centers for Disease Control and Prevention (ECDC) scheme and also analyzed for resistance gene. Treponemal DNA from 18 lesions and 18 blood specimens were submitted for amplification using Polymerase Chain Reaction (PCR) and Polymerase Chain Reaction in Real Time (RT-PCR). Eight samples from lesions and eight from blood were positive in the RT-PCR analysis. Eight lesions and three blood samples were positive using PCR. Two samples exhibited azithromycin resistance. The Brazilian isolate types 14d/g, 14 d/c, 15d/c, and 15d/e were identified using the ECDC scheme. The three subtypes 14d/c, 15d/c, and 15d/e have been identified in Brazil for the first time.

Immune Response to an Inactivated Vaccine of SARS-CoV-2 (CoronaVac) in an Indigenous Brazilian Population: A Cohort Study.

INTRODUCTION: Although the adaptive immune responses to the CoronaVac vaccine are known, their dynamics in indigenous communities remain unclear. In this study, we assessed the humoral and cellular immune responses to CoronaVac (Sinovac Biotech Life Sciences, 2021 NCT05225285, Beijing, China), in immunized Brazilian indigenous individuals. METHODS: We conducted a prospective cohort study on indigenous Brazilian people between February 2021 and June 2021. Analyses of immune responses were carried out before (T1) and after a vaccination schedule was completed (T2). Demographic data were collected using a questionnaire. RESULTS: We initially included 328 patients; among them, 120 (36.6%) had no SARS-CoV-2 antibodies. Peripheral blood mononuclear cells (PBMCs) were collected from 106 patients during follow-up visits, of which 91 samples were analyzed by immunophenotyping assay to detect SARS-CoV-2-specific memory T-cell response. Post-vaccination, the levels of memory B-cells and Natural Killer T-lymphocytes increased. Bororó village residents, females, and Terena ethnic group members had higher levels of anti-spike IgG antibodies post-vaccination, whereas alcohol and tobacco users had lower concentrations. CONCLUSIONS: To our best knowledge, this was the first comprehensive assessment of antibody and T-cell responses against CoronaVac vaccination in indigenous patients. Our findings showed that antibody response and T-cell immunity against SARS-CoV-2 were present in most patients following the vaccination schedule.

The inclusion of Amazon mangroves in Brazil's REDD+ program.

The Legal Amazon of Brazil holds vast mangrove forests, but a lack of awareness of their value has prevented their inclusion into results-based payments established by the United Nations Framework Convention on Climate Change. Based on an inventory from over 190 forest plots in Amazon mangroves, we estimate total ecosystem carbon stocks of 468 ± 67 Megagrams (Mg) ha-1; which are significantly higher than Brazilian upland biomes currently included into national carbon offset financing. Conversion of mangroves results in potential emissions of 1228 Mg CO2e ha-1, which are 3-fold higher than land use emissions from conversion of the Amazon rainforest. Our work provides the foundation for the inclusion of mangroves in Brazil's intended Nationally Determined Contribution, and here we show that halting mangrove deforestation in the Legal Amazon would generate avoided emissions of 0.9 ± 0.3 Teragrams (Tg) CO2e yr-1; which is equivalent to the annual carbon accumulation in 82,400 ha of secondary forests.

Overlay databank unlocks data-driven analyses of biomolecules for all.

Tools based on artificial intelligence (AI) are currently revolutionising many fields, yet their applications are often limited by the lack of suitable training data in programmatically accessible format. Here we propose an effective solution to make data scattered in various locations and formats accessible for data-driven and machine learning applications using the overlay databank format. To demonstrate the practical relevance of such approach, we present the NMRlipids Databank-a community-driven, open-for-all database featuring programmatic access to quality-evaluated atom-resolution molecular dynamics simulations of cellular membranes. Cellular membrane lipid composition is implicated in diseases and controls major biological functions, but membranes are difficult to study experimentally due to their intrinsic disorder and complex phase behaviour. While MD simulations have been useful in understanding membrane systems, they require significant computational resources and often suffer from inaccuracies in model parameters. Here, we demonstrate how programmable interface for flexible implementation of data-driven and machine learning applications, and rapid access to simulation data through a graphical user interface, unlock possibilities beyond current MD simulation and experimental studies to understand cellular membranes. The proposed overlay databank concept can be further applied to other biomolecules, as well as in other fields where similar barriers hinder the AI revolution.

Efficient delivery of carotenoids to adipocytes with albumin.

Carotenoids are very effectively delivered by albumin to adipocytes. The uptake of carotenoids to the cells occurs in the form of self-aggregates that localize in the vicinity of the adipocyte membrane, as shown by high spatial resolution Raman spectroscopy. The binding of carotenoids to albumin and the mechanism of their transport were elucidated with the help of chiroptical spectroscopies, in tandem with molecular docking and molecular dynamics simulations. In particular, apart from the recognized high affinity pocket of albumin that binds a carotenoid monomer in domain I, we have identified a hydrophobic periphery area in domain IIIB that loosely bounds the self-aggregated carotenoid in aqueous media and enables its easy detachment in hydrophobic environments. This explains the effectiveness of albumins as nanocarriers of carotenoids to adipocytes in vitro.

Centrality measures in psychological networks: A simulation study on identifying effective treatment targets.

The network theory of psychopathology suggests that symptoms in a disorder form a network and that identifying central symptoms within this network might be important for an effective and personalized treatment. However, recent evidence has been inconclusive. We analyzed contemporaneous idiographic networks of depression and anxiety symptoms. Two approaches were compared: a cascade-based attack where symptoms were deactivated in decreasing centrality order, and a normal attack where symptoms were deactivated based on original centrality estimates. Results showed that centrality measures significantly affected the attack's magnitude, particularly the number of components and average path length in both normal and cascade attacks. Degree centrality consistently had the highest impact on the network properties. This study emphasizes the importance of considering centrality measures when identifying treatment targets in psychological networks. Further research is needed to better understand the causal relationships and predictive capabilities of centrality measures in personalized treatments for mental disorders.

Good Syndrome in a Young Woman: An Unusual Presentation.

Good Syndrome is a rare disease that comprises the presence of a thymoma, immunodeficiency, and recurrent opportunistic infections. We report the case of a young woman who was diagnosed with Good Syndrome, who had a long-term history of recurrent infections, often due to atypical agents, and who also had a previous history of immunodeficiency and a B1 thymoma invading the large vessels, lung, and pericardium (Masaoka stage IV). She underwent surgical resection of the mediastinal mass, requiring vena cava superior reconstruction due to the extent of invasion, followed by adjuvant radiotherapy and immunoglobulin G supplementation. Despite relative stability in the subsequent years, without serious infections, after three years she had a thymoma recurrence requiring a new therapeutic approach. This case highlights the importance of a thorough investigation of the underlying causes of recurrent infections, which may be the result of an immunodeficiency secondary to malignancy. In young patients, early diagnosis is crucial to avoid disease progression and to reduce mortality rates. To achieve such outcomes, a multidisciplinary team and a comprehensive therapeutic strategy are necessary.

Cross-Modal Sensory Boosting to Improve High-Frequency Hearing Loss: Device Development and Validation.

Background: High-frequency hearing loss is one of the most common problems in the aging population and with those who have a history of exposure to loud noises. This type of hearing loss can be frustrating and disabling, making it difficult to understand speech communication and interact effectively with the world. Objective: This study aimed to examine the impact of spatially unique haptic vibrations representing high-frequency phonemes on the self-perceived ability to understand conversations in everyday situations. Methods: To address high-frequency hearing loss, a multi-motor wristband was developed that uses machine learning to listen for specific high-frequency phonemes. The wristband vibrates in spatially unique locations to represent which phoneme was present in real time. A total of 16 participants with high-frequency hearing loss were recruited and asked to wear the wristband for 6 weeks. The degree of disability associated with hearing loss was measured weekly using the Abbreviated Profile of Hearing Aid Benefit (APHAB). Results: By the end of the 6-week study, the average APHAB benefit score across all participants reached 12.39 points, from a baseline of 40.32 to a final score of 27.93 (SD 13.11; N=16; P=.002, 2-tailed dependent t test). Those without hearing aids showed a 10.78-point larger improvement in average APHAB benefit score at 6 weeks than those with hearing aids (t14=2.14; P=.10, 2-tailed independent t test). The average benefit score across all participants for ease of communication was 15.44 (SD 13.88; N=16; P<.001, 2-tailed dependent t test). The average benefit score across all participants for background noise was 10.88 (SD 17.54; N=16; P=.03, 2-tailed dependent t test). The average benefit score across all participants for reverberation was 10.84 (SD 16.95; N=16; P=.02, 2-tailed dependent t test). Conclusions: These findings show that vibrotactile sensory substitution delivered by a wristband that produces spatially distinguishable vibrations in correspondence with high-frequency phonemes helps individuals with high-frequency hearing loss improve their perceived understanding of verbal communication. Vibrotactile feedback provides benefits whether or not a person wears hearing aids, albeit in slightly different ways. Finally, individuals with the greatest perceived difficulty understanding speech experienced the greatest amount of perceived benefit from vibrotactile feedback.

Are volatile methylsiloxanes in downcycled tire microplastics? Levels and human exposure estimation in synthetic turf football fields.

Downcycled rubber, derived from end-of-life tires (ELTs), is frequently applied as crumb rubber (CR) as infill of synthetic turf in sports facilities. This practice has been questioned in recent years as numerous studies have reported the presence of potentially hazardous chemicals in this material. CR particles fall into the category of microplastics (MPs), making them possible vectors for emerging micropollutants. A preliminary study where volatile methylsiloxanes (VMSs) were found in CR originated the hypothesis that VMSs are present in this material worldwide. Consequently, the present work evaluates for the first time the levels and trends of seven VMSs in CR from synthetic turf football fields, while attempting to identify the main sources and impacts of these chemicals. A total of 135 CR samples and 12 other of alternative materials were analyzed, employing an ultrasound-assisted dispersive solid-phase extraction followed by gas chromatography-mass spectrometry (GC-MS), and the presence of VMSs was confirmed in all samples, in total concentrations ranging from 1.60 to 5089 ng.g-1. The levels were higher in commercial CR (before field application), a reflection of the use of VMS-containing additives in tire production and/or the degradation of silicone polymers employed in vehicles. The VMSs generally decreased over time on the turf, as expected given their volatile nature and the wearing of the material. Finally, the human exposure doses to VMSs in CR (by dermal absorption and ingestion) for people in contact with synthetic turf in football fields were negligible (maximum total exposure of 20.5 ng.kgBW-1.year-1) in comparison with the European Chemicals Agency (ECHA) reference doses: 1.35 × 109 ng.kgBW-1.year-1 for D4 and 1.83 × 109 ng.kgBW-1.year-1 for D5. Nevertheless, more knowledge on exposure through inhalation and the combined effects of all substances is necessary to provide further corroboration. This work proved the presence of VMSs in CR from ELTs, another family of chemical of concern to take into account when studying MPs as vectors of other contaminants.

Current views on in vivo models for breast cancer research and related drug development.

INTRODUCTION: Animal models play a crucial role in breast cancer research, in particular mice and rats, who develop mammary tumors that closely resemble their human counterparts. These models allow the study of mechanisms behind breast carcinogenesis, as well as the efficacy and safety of new, and potentially more effective and advantageous therapeutic approaches. Understanding the advantages and disadvantages of each model is crucial to select the most appropriate one for the research purpose. AREA COVERED: This review provides a concise overview of the animal models available for breast cancer research, discussing the advantages and disadvantages of each one for searching new and more effective approaches to treatments for this type of cancer. EXPERT OPINION: Rodent models provide valuable information on the genetic alterations of the disease, the tumor microenvironment, and allow the evaluation of the efficacy of chemotherapeutic agents. However, in vivo models have limitations, and one of them is the fact that they do not fully mimic human diseases. Choosing the most suitable model for the study purpose is crucial for the development of new therapeutic agents that provide better care for breast cancer patients.

Seroprevalence Of SARS-COV-2 infection in asymptomatic indigenous from the largest Brazilian periurban area.

This study assessed the seroprevalence of SARS-CoV-2 in 496 asymptomatic individuals from Mato Grosso do Sul, located in Dourados, the largest periurban indigenous area in Brazil, from January 25 to February 4, 2021. The volunteers participated before receiving their first dose of the CoronaVac inactivated vaccine. For screening, blood samples were collected and analyzed using SARS-CoV-2 rapid tests and the enzyme-linked immunosorbent assay (ELISA). We observed varying trends in total anti-SARS-CoV-2 antibodies across different variables. Seropositivity among the participants tested was 63.70% (316/496) using the rapid test and 52.82% (262/496) were positive using the ELISA method. The majority of participants identified with the Guarani-Kaiowá ethnic group, with 66.15% (217/328), and other ethnic groups with 58.84% (193/328). The median age of the subjects was 30.5 years, with 79.57% (261/328) being femaleThis research showed the elevated seroprevalence of SARS-CoV-2 antibodies in asymptomatic Brazilians. The findings indicate a high seropositivity rate among the asymptomatic indigenous population of Midwest Brazil. This underscores the overlooked status of these communities and underscores the need for targeted national initiatives that emphasize the protection of vulnerable ethnic groups in the fight against COVID-19.

Dermis resident macrophages orchestrate localized ILC2 eosinophil circuitries to promote non-healing cutaneous leishmaniasis.

Tissue-resident macrophages are critical for tissue homeostasis and repair. We previously showed that dermis-resident macrophages produce CCL24 which mediates their interaction with IL-4+ eosinophils, required to maintain their M2-like properties in the TH1 environment of the Leishmania major infected skin. Here, we show that thymic stromal lymphopoietin (TSLP) and IL-5+ type 2 innate lymphoid cells are also required to maintain dermis-resident macrophages and promote infection. Single cell RNA sequencing reveals the dermis-resident macrophages as the sole source of TSLP and CCL24. Generation of Ccl24-cre mice permits specific labeling of dermis-resident macrophages and interstitial macrophages from other organs. Selective ablation of TSLP in dermis-resident macrophages reduces the numbers of IL-5+ type 2 innate lymphoid cells, eosinophils and dermis-resident macrophages, and ameliorates infection. Our findings demonstrate that dermis-resident macrophages are self-maintained as a replicative niche for L. major by orchestrating localized type 2 circuitries with type 2 innate lymphoid cells and eosinophils.

HOP1 and HAP2 are conserved components of the meiosis-related machinery required for successful mating in Leishmania.

Whole genome analysis of Leishmania hybrids generated experimentally in sand flies supports a meiotic mechanism of genetic exchange, with Mendelian segregation of the nuclear genome. Here, we perform functional analyses through the generation of double drug-resistant hybrids in vitro and in vivo (during sand fly infections) to assess the importance of conserved meiosis-related genes in recombination and plasmogamy. We report that HOP1 and a HAP2-paralog (HAP2-2) are essential components of the Leishmania meiosis machinery and cell-to-cell fusion mechanism, respectively, since deletion of either gene in one or both parents significantly reduces or completely abrogates mating competence. These findings significantly advance our understanding of sexual reproduction in Leishmania, with likely relevance to other trypanosomatids, by formally demonstrating the involvement of a meiotic protein homolog and a distinct fusogen that mediates non-canonical, bilateral fusion in the hybridizing cells.

Long-term impacts on estuarine benthic assemblages (2015-2020) after a mine tailing spill in SE Brazil.

The Rio Doce estuary was critically impacted in 2015 by the world's largest mining tailing spill, with still unclear long-term effects on the aquatic biota. Here we present a long-term (2015 to 2020) assessment of estuarine benthic assemblages, where we demonstrate that despite a decline in the absolute concentrations of potentially toxic elements; sediment contamination is still above pre-impact conditions. The presence of these contaminants is likely responsible for a continued low habitat quality for the benthic fauna, characterized by a reduction of 96 % of the macroinvertebrate density and persistent change in the benthic assemblage composition. Our study supports previous work indicating the long-term nature of pollution impacts in estuaries, and demonstrate that although water quality levels were quickly adequate under regulatory terms, they largely lack significance to the overall ecosystem health assessment, as they are not related to the recovery of bottom- dwelling assemblages in estuarine ecosystems.

Time-domain proton-detected local-field NMR for molecular structure determination in complex lipid membranes.

Proton-detected local-field (PDLF) NMR spectroscopy, using magic-angle spinning and dipolar recoupling, is presently the most powerful experimental technique for obtaining atomistic structural information from small molecules undergoing anisotropic motion. Common examples include peptides, drugs, or lipids in model membranes and molecules that form liquid crystals. The measurements on complex systems are however compromised by the larger number of transients required. Retaining sufficient spectral quality in the direct dimension requires that the indirect time-domain modulation becomes too short for yielding dipolar splittings in the frequency domain. In such cases, the dipolar couplings can be obtained by fitting the experimental data; however ideal models often fail to fit PDLF data properly due to effects of radiofrequency field (RF) spatial inhomogeneity. Here, we demonstrate that by accounting for RF spatial inhomogeneity in the modeling of R-symmetry-based PDLF NMR experiments, the fitting accuracy is improved, facilitating the analysis of the experimental data. In comparison to the analysis of dipolar splittings without any fitting procedure, the accurate modeling of PDLF measurements makes possible three important improvements: the use of shorter experiments that enable the investigation of samples with a higher level of complexity, the measurement of C-H bond order parameters with smaller magnitudes |SCH| and of smaller variations of |SCH| caused by perturbations of the system, and the determination of |SCH| values with small differences from distinct sites having the same chemical shift. The increase in fitting accuracy is demonstrated by comparison with 2H NMR quadrupolar echo experiments on mixtures of deuterated and non-deuterated dimyristoylphosphatidylcholine (DMPC) and with 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (POPE) membranes. Accurate modeling of PDLF NMR experiments is highly useful for investigating complex membrane systems. This is exemplified by application of the proposed fitting procedure for the characterization of membranes composed of a brain lipid extract with many distinct lipid types.

Prevalence of HIV-1 infection and associated characteristics in a Brazilian indigenous population: a cross-sectional study.

Background: Despite significant progress in the areas of prevention, diagnosis, and treatment, HIV continues to result in a substantial number of fatalities on a global scale each year. Gaining insights from epidemiological data can prove instrumental in the development of health promotion strategies, particularly within vulnerable populations, such as indigenous groups. Consequently, our study aimed to investigate the prevalence of HIV infection within the indigenous population residing in the second-largest region of Brazil. Additionally, we sought to explore the subtypes of HIV-1 and detect any drug-resistance mutations present within this population. Methods: In this cross-sectional study, we aimed to evaluate the prevalence of HIV-1 infection and explore its associated characteristics within the indigenous population residing in the villages of Jaguapiru and Bororó, located in the Dourados area of Mato Grosso do Sul (MS), Brazil. Blood samples were collected for rapid HIV screening, serological tests, nucleic acid amplification, and HIV subtyping. Additionally, the HIV-1 viral load and CD4+ T lymphocyte count of the people living with HIV (PLHIV) were assessed at the time of recruitment and 24 weeks later. Findings: Out of the 2190 invited individuals, 1927 (88%) were included in this study. The average age of the participants was 34.2 (±13.8) years, with a majority of 74% being female. Moreover, 68.44% of the participants identified themselves as belonging to the Guarani-Kaiowa ethnic group. HIV seroprevalence was 0.93% (18/1927), and 73.22% (1411/1927) were unaware of their serological status. The prevalence of HIV-1 was higher in single indigenous people [10/617 (1.62%)], who received government benefits [14/1021 (1.37%)], had less than five years of formal education [11/685 (1.61%)], had sexual intercourse with users of injectable drugs [2/21 (9.52%)], with history of sexually transmitted infections (STIs) [10/62 (16.2%)] and incarceration [3/62 (4.84%)]. Of 18 positive samples, 44.4% (8/18) were successfully amplified, and HIV-1 subtype C was prevalent. Furthermore, we identified HIV-1 drug resistance mutations in four patients, specifically from the classes of Protease Inhibitor, Nucleoside Reverse Transcriptase Inhibitor, and Non-Nucleoside Reverse Transcriptase Inhibitor. Notably, three of these patients exhibited a high viral load even after 24 weeks of undergoing antiretroviral therapy. Out of the 18 PLHIV, 66.66% (12/18) had a viral load below 1000 copies/mL, while 50% (9/18) had a CD4+ T lymphocytes count greater than 350 cells/mL after 24 weeks of treatment. Interpretation: Despite the concerted efforts to control HIV infection, the prevalence observed in the indigenous population under study surpassed that reported in other Brazilian indigenous groups. This disparity highlights the disproportionate impact of the disease on this particular group. The detection of drug-resistance mutations further emphasizes the critical need to expand diagnostic coverage, closely monitor treatment strategies, and maintain ongoing molecular surveillance. These measures are imperative for enhancing HIV management within this vulnerable population. Funding: This study was partially funded by Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Fundação de Apoio ao Desenvolvimento do Ensino, Ciência e Tecnologia do Estado de Mato Grosso do Sul (FUNDECT), Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES), Secretaria do Estado de Saúde (SES) of Governo do Estado de Mato Grosso do Sul, and Universidade Federal da Grande Dourados (UFGD).

Effects of Cathepsin S Inhibition in the Age-Related Dry Eye Phenotype.

Purpose: Aged C57BL/6J (B6) mice have increased levels of cathepsin S, and aged cathepsin S (Ctss-/-) knockout mice are resistant to age-related dry eye. This study investigated the effects of cathepsin S inhibition on age-related dry eye disease. Methods: Female B6 mice aged 15.5 to 17 months were randomized to receive a medicated diet formulated by mixing the RO5461111 cathepsin S inhibitor or a standard diet for at least 12 weeks. Cornea mechanosensitivity was measured with a Cochet-Bonnet esthesiometer. Ocular draining lymph nodes and lacrimal glands (LGs) were excised and prepared for histology or assayed by flow cytometry to quantify infiltrating immune cells. The inflammatory foci (>50 cells) were counted under a 10× microscope lens and quantified using the focus score. Goblet cell density was investigated in periodic acid-Schiff stained sections. Ctss-/- mice were compared to age-matched wild-type mice. Results: Aged mice subjected to cathepsin S inhibition or Ctss-/- mice showed improved conjunctival goblet cell density and cornea mechanosensitivity. There was no change in total LG focus score in the diet or Ctss-/- mice, but there was a lower frequency of CD4+IFN-γ+ cell infiltration in the LGs. Furthermore, aged Ctss-/- LGs had an increase in T central memory, higher numbers of CD19+B220-, and fewer CD19+B220+ cells than wild-type LGs. Conclusions: Our results indicate that therapies aimed at decreasing cathepsin S can ameliorate age-related dry eye disease with a highly beneficial impact on the ocular surface. Further studies are needed to investigate the role of cathepsin S during aging.

Rocking motion in solid proteins studied by the 15N proton-decoupled R1ρ relaxometry.

Recently it has been revealed that proteins in solid samples undergo slow overall rocking. The parameters of this motion depend on intermolecular interactions. Therefore, the characterization of the rocking motion enables one to investigate protein-protein interactions. NMR R1ρ relaxometry is the most suitable tool to study slow molecular motions. However, the time scale of the rocking motion is on the edge of the dynamics window of the standard R1ρ experiment, precluding the R1ρ data analysis from being precise and reliable. In this work, we apply a modified R1ρ relaxation method to characterize the slow motion in solids with much higher precision and reliability. The modification is the simultaneous use of a strong 1H-CW pulse and a weak/moderate 15N spin-lock pulse. We demonstrate theoretically and experimentally that under this condition, R1ρ decays have a significantly better signal-to-noise ratio and a much shorter "dead time" caused by the initial oscillations compared to the conventional R1ρ experiment. Moreover, the proton-decoupled R1ρ's can be measured at a much smaller difference between the spin-lock and MAS frequencies; thus, much slower molecular motions can be sampled. The proton decoupling during the 15N spin-lock pulse also suppresses the interfering coherent spin-spin relaxation pathway at low spin-lock fields, which overlaps the Bloch-McConnell (chemical exchange) range of R1ρ dispersions. The proton-decoupled and standard R1ρ experiments were used to study the rocking motion of 15N,2H-enriched protein GB1 in two solid forms, microcrystals and lyophilized amorphous powder. The most striking finding is that the correlation function of this motion consists of two components with very different correlation times, 2-20 µs and a few hundred µs. The rocking motion parameters in microcrystals and powder are quite different, revealing the distinct nature of inter-protein interactions in these two samples.