Miracle-dz/UniFAR-FTUs huggingface dataset:
Detection and measurement of stress hormones with mass spectrometry

The purpose of this thesis was to develop and validate a liquid chromatography mass spectrometry (LC-MS) method to measure salivary cortisol and testosterone. Furthermore, to use this method to analyse saliva samples collected during an exercise study measuring the hormonal response to acute exercise. The ongoing contribution to knowledge provided by this research is the application of a developed and validated LC-MS method to analysis of cortisol from pre and post exercise saliva samples, and evaluation of the cortisol and testosterone response to three different training sessions; including an interval session (INT), tempo run (TEMP) and aerobic circuit training (CIR). An LC-MS method was developed to measure cortisol and testosterone. The mass spectrometer used was a triple quadrupole BioQ (Waters/Micromass, UK) coupled with a 1200 series HPLC machine (Agilent Technologies, UK). Optimisation of various parameters was undertaken including: cone voltage and capillary voltage, followed by optimisation of liquid chromatography (LC) parameters such as; mobile phase gradient and flow rate. Saliva samples were collected at rest, and hormones measured with the new LC-MS method and ELISA to validate the developed method. Finally, heart rate and salivary and plasma cortisol and testosterone response to acute exercise were tested, to compare the response to different training protocols in runners. Following this, the MS method underwent further validation against ELISA. Results revealed optimum MS parameters were: cone voltage 25 V; capillary voltage 3.5 kV and LM and HM resolution 8 Da. For the LC method, mobile phase flow rate was optimised at 0.1 mL/min, with a gradient profile ranging from 50 to 95% methanol (MeOH), and a run time of 15 minutes. Sample preparation was also considered (solid phase extraction); and wash phase of 10% MeOH, 10 times sample concentration and reconstitution with 80% MeOH were optimised to improve analyte recovery and detection. Comparison of cortisol measured at rest with LC-MS and ELISA (n=22) revealed a correlation between methods (r=0.83, P<0.001). The exercise study showed the INT elicited a higher peak heart rate (172±11) than CIR (148±10) or TEMP trials (163±10). INT also produced a higher RPE (15 ±2) than CIR (13±1) and TEMP (14±2) trials. Salivary cortisol increased at all time points post exercise in INT and remained higher than rest (1.29 ± 0.51 ng/mL) at 60 minutes recovery (3.13 ±1.59 ng/mL). Salivary testosterone increased pre to post exercise in CIR (119.7±39.5 pg/mL to 150.1 ±51.5 pg/mL), TEMP (142.4±76.5 pg/mL to 185.0±49.1 pg/mL) and INT (117.5±23.5 pg/mL to 176.3±46.0 pg/mL) trials. There was also a correlation between cortisol measured in plasma and saliva (r=0.813, p=0.0001). Additionally, comparison of salivary cortisol measurements obtained with mass spectrometry and ELISA (n=85) showed a correlation (r=0.934, p=00001). In conclusion, validation revealed cortisol could be quantfied accurately and precisely with LC-MS. Additionally, INT elicited a higher mean RPE and cortisol response than TEMP or CIR.
Viability and Antioxidant Effects of Traditional Cooling Rice Powder (bedak sejuk) Made from Oryza sativa ssp. Indica and Oryza sativa ssp. japonica on UVB-Induced B164A5 Melanoma Cells

Background: Traditional cooling rice powder (bedak sejuk) is a fermented rice-based cosmetic that is applied topically on one’s skin, as an overnight facial mask. According to user testimonies, bedak sejuk beautifies and whitens skin, whereby these benefits could be utilised as a potential melanoma chemopreventive agent. Objective: Hence, this study aimed to determine the effects of bedak sejuk made from Oryza sativa ssp. indica (Indica) and Oryza sativa ssp. japonica (Japonica) on UVB-induced B164A5 melanoma cells, and also identify the antioxidant capacities of both types of bedak sejuk. Methods: The optimum dose of Indica and Japonica bedak sejuk to treat the cells was determined via the MTT assay. Then, the antioxidant capacities of both types of bedak sejuk were determined using the FRAP assay. Results: From the MTT assay, it was found that Indica and Japonica bedak sejuk showed no cytotoxic effects towards the cells. Hence, no IC50 can be obtained and two of the higher doses, 50 and 100 g/L were chosen for treatment. In the FRAP assay, Indica bedak sejuk at 50 and 100 g/L showed FRAP values of 0.003 ± 0.001 μg AA (ascorbic acid)/g of bedak sejuk and 0.004 ± 0.0003 μg AA/g of bedak sejuk. Whereas Japonica bedak sejuk at 50 g/L had the same FRAP value as Indica bedak sejuk at 100 g/L. As for Japonica bedak sejuk at 100 g/L, it showed the highest antioxidant capacity with the FRAP value of 0.01 ± 0.0007 μg AA/g of bedak sejuk which was statistically significant (p < 0.05) when compared to other tested concentrations. Conclusion: In conclusion, Japonica bedak sejuk has a higher antioxidant capacity compared to Indica bedak sejuk despite both being not cytotoxic towards the cells. Regardless, further investigations need to be done before bedak sejuk could be developed as potential melanoma chemoprevention agents.
Extensive characterization of the composition and functional activities of five preparations of human platelet lysates for dedicated clinical uses

Abstract Human platelet lysates (HPLs), rich in various growth factors and cell growth-promoting molecules, encompass a new range of blood products that are being used for regenerative medicine, cell therapies, and tissue engineering. Well-characterized dedicated preparations, tailor-made to best fit specific therapeutic applications, are needed for optimal clinical efficacy and safety. Here, five types of HPL were prepared from the same platelet concentrates and extensively characterized to determine and compare their proteins, growth factors, cytokines, biochemical profiles, thrombin-generating capacities, thrombin-associated proteolytic activities, phospholipid-associated procoagulant potential, contents of extracellular vesicles expressing phosphatidylserine and tissue factor, and antioxidative properties. Our results revealed that all five HPL preparations contained detectable supraphysiological levels, in the ca. 0.1 ~ 350-ng/ml range, of all growth factors assessed, except insulin-like growth factor-1 detected only in HPL containing plasma. There were significant differences observed among these HPLs in total protein content, fibrinogen, complement components C3 and C4, albumin, and immunoglobulin G, and, most importantly, in their functional coagulant and procoagulant activities and antioxidative capacities. Our data revealed that the biochemical and functional properties of HPL preparations greatly vary depending upon their mode of production, with potential impacts on the safety and efficacy for certain clinical indications. Modes of preparation of HPLs should be carefully designed, and the product properties carefully evaluated based on the intended therapeutic use to ensure optimal clinical outcomes.
Novel inhibitory brainstem neurons with selective projections to spinal lamina I reduce both pain and itch

Sensory information is transmitted from peripheral nerves, through the spinal cord, and up to the brain (“bottom up” pathway). Some of this information may be modulated by “top‐down” projections from the brain to the spinal cord. Discovering endogenous mechanisms for reducing pain and itch holds enormous potential for developing new treatments. However, neurons mediating the top‐down inhibition of pain are not well understood, nor has any such pathway been identified for itch sensation. Here, we identify a novel population of GABAergic neurons in the ventral brainstem, distinguished by prodynorphin expression, which we named LJA5. LJA5 neurons provide the only known inhibitory projection specifically to lamina I of the spinal cord, which contains sensory neurons that transmit pain and itch information up to the brain. Chemogenetically activating LJA5 neurons in male mice reduces capsaicin‐induced pain and histamine‐induced itch. Identifying this new pathway opens new treatment opportunities for chronic, refractory pain, and pruritis.
Ustilago maydis effector Jsi1 interacts with Topless corepressor, hijacking plant jasmonate/ethylene signaling

Summary Ustilago maydis is the causal agent of maize smut disease. During the colonization process, the fungus secretes effector proteins that suppress immune responses and redirect the host metabolism in favor of the pathogen. As effectors play a critical role during plant colonization, their identification and functional characterization are essential to understanding biotrophy and disease. Using biochemical, molecular, and transcriptomic techniques, we performed a functional characterization of the U. maydis effector Jasmonate/Ethylene signaling inducer 1 (Jsi1). Jsi1 interacts with several members of the plant corepressor family Topless/Topless related (TPL/TPR). Jsi1 expression in Zea mays and Arabidopsis thaliana leads to transcriptional induction of the ethylene response factor (ERF) branch of the jasmonate/ethylene (JA/ET) signaling pathway. In A. thaliana, activation of the ERF branch leads to biotrophic susceptibility. Jsi1 likely activates the ERF branch via an EAR (ET‐responsive element binding‐factor‐associated amphiphilic repression) motif, which resembles EAR motifs from plant ERF transcription factors, that interacts with TPL/TPR proteins. EAR‐motif‐containing effector candidates were identified from different fungal species, including Magnaporthe oryzae, Sporisorium scitamineum, and Sporisorium reilianum. Interaction between plant TPL proteins and these effector candidates from biotrophic and hemibiotrophic fungi indicates the convergent evolution of effectors modulating the TPL/TPR corepressor hub.
Compartmentalized evolution of hepatitis B virus contributes differently to the prognosis of hepatocellular carcinoma.

Serum hepatitis B virus (HBV) mutations can predict hepatocellular carcinoma (HCC) occurrence. We aimed to clarify if HBV evolves synchronously in the sera, adjacent liver, and tumors and predict HCC prognosis equally. A total of 203 HBV-positive HCC patients with radical hepatectomy in Shanghai, China during 2011-2015 were enrolled in this prospective study. Quasispecies complexity (QC) in HBV core promoter region was assessed using clone-based sequencing. We performed RNA-sequencing on tumors and paired adjacent tissues of another 15 HCC patients and analyzed it with 3 public datasets containing 127 samples. HBV QC was positively correlated to APOBEC3s' expression level (r=0.28, p<0.001), higher in the adjacent tissues than in the tumors (p=6.50e-3), and higher in early tumors than in advanced tumors (p=0.039). The evolutionary distance between the sera-derived HBV strains and the tumor-derived ones was significantly longer than that between the sera-derived ones and the adjacent tissue-derived ones (p<0.001). Multivariate Cox regression analyses indicated that high HBV QC in the sera predicted an unfavorable overall survival (OS) (p=0.002) and recurrence-free survival (RFS) (p=0.004) in HCC; whereas it in the tumors predicted a favorable RFS (p<0.001), respectively. APOBECs-related HBV mutations including G1764A were more frequent in the sera than in the adjacent tissues. High-frequent A1762T/G1764A in the sera predicted an unfavorable RFS (p<0.001); whereas it in the tumors predicted a favorable RFS (p=0.035). In conclusion, HBV evolves more advanced in the sera than in the tumors. HBV QC and A1762T/G1764A in the sera and tumors have contrary prognostic effects in HCC.
Tissue Expression Of LPHN3 in Breast Cancer: An Immunohistochemistry Method

Objective: Breast cancer is one of the most important public health problems among women worldwide. It is a major cause of morbidity especially among women in developing countries including Thailand. The purpose of this study was to study the expression of LPHN3 protein in normal breast tissue compared to breast cancer tissue. Methods: We had studied the expression of LPHN3 in 65 breast tissues using an immunohistochemistry method. The association between LPHN3 expression and breast cancer metastasis to nearby axillary lymph nodes was also examined. Results: Among the 65 breast cancer and normal breast tissues examined, LPHN3 expression with an immunohistochemistry index (IHC index) greater than 4 was more frequently found in breast cancer tissues than in normal breast tissues (P-value = 0.001, OR (95% CI) = 7.04 (2.16-23)). Moreover, a high expression of LPHN3 (IHC index > 4) was more frequently found in breast cancer tissues with negative axillary lymph nodes than in those with positive ones (P-value = 0.038, OR (95% CI) = 0.25 (0.07-0.96)). LPHN3 protein might be a new metastasis suppressor gene in breast cancer and a marker for breast cancer metastasis prevention. Conclusions: The present study indicated that a decrease of LPHN3 protein expression in breast cancer tissue might be a marker indicating the aggressiveness of breast cancer. These results also suggested that a decrease of LPHN3 expression could be functionally involved in breast cancer progression and metastasis.
Exploring nature’s bounty: identification of Withania somnifera as a promising source of therapeutic agents against COVID-19 by virtual screening and in silico evaluation

Abstract Coronaviruses are etiological agents of extreme human and animal infection resulting in abnormalities primarily in the respiratory tract. Presently, there is no defined COVID-19 intervention and clinical trials of prospective therapeutic agents are still in the nascent stage. Withania somnifera (L.) Dunal (WS), is an important medicinal plant in Ayurveda. The present study aimed to evaluate the antiviral potential of selected WS phytoconstituents against the novel SARS-CoV-2 target proteins and human ACE2 receptor using in silico methods. Most of the phytoconstituents displayed good absorption and transport kinetics and were also found to display no associated mutagenic or adverse effect(s). Molecular docking analyses revealed that most of the WS phytoconstituents exhibited potent binding to human ACE2 receptor, SAR-CoV and SARS-CoV-2 spike glycoproteins as well as the two main SARS-CoV-2 proteases. Most of the phytoconstituents were predicted to undergo Phase-I metabolism prior to excretion. All phytoconstituents had favorable bioactivity scores with respect to various receptor proteins and target enzymes. SAR analysis revealed that the number of oxygen atoms in the withanolide backbone and structural rearrangements were crucial for effective binding. Molecular simulation analyses of SARS-CoV-2 spike protein and papain-like protease with Withanolides A and B, respectively, displayed a stability profile at 300 K and constant RMSDs of protein side chains and Cα atoms throughout the simulation run time. In a nutshell, WS phytoconstituents warrant further investigations in vitro and in vivo to unravel their molecular mechanism(s) and modes of action for their future development as novel antiviral agents against COVID-19. GRAPHICAL REPRESENTATION OF PROPOSED STUDY Communicated by Ramaswamy H. Sarma
Clinical Utility of BRAF, NRAS, and TERT Promoter Mutation in Preoperative Thyroid Fine-Needle Aspiration Biopsy: A Diagnostic Study From Dharmais Cancer Hospital

Objective: Molecular testing of thyroid nodules becomes important for improving the accuracy of fine-needle aspiration biopsy (FNAB). This study aimed to investigate the diagnostic utility of BRAF, NRAS, and TERT promoter mutation in thyroid nodules at Dharmais Cancer Hospital. Methods: We performed a prospective diagnostic study involving 50 patients with thyroid nodules who needed surgery between September 2013 and August 2014. Mutational hotspots in BRAF exon 15, NRAS exon 3, and TERT promoter region were analyzed by Sanger sequencing from FNAB specimens. Cytology and molecular data were compared to histopathology results. Results: Of the 50 cases included in the analysis, 39 cases (78%) were thyroid malignancies. Mutations of BRAF, NRAS, and TERT promoter were detected in 31% (12/39), 18% (7/39), and 13% (5/39) cases, respectively. BRAF and NRAS mutations were found mutually exclusive, while all of TERT promoter mutation was found coexistent either with BRAF (40%) or NRAS (60%). The combination of FNAB cytology and molecular testing resulted in 69% sensitivity, 100% specificity, 100% positive predictive value, 48% negative predictive value, and 76% accuracy. Conclusion: Molecular testing of BRAF, NRAS, and TERT mutations improve the sensitivity of thyroid FNAB and is beneficial for more definitive treatment in selective cases. However, the NPV is relatively low to avoid the need for diagnostic surgery. Therefore, further studies to identify more sensitive methods and more comprehensive molecular markers in the diagnosis of thyroid nodules are needed.
Usability Assessment of the Rehabilitation Lower-limb Orthopedic Assistive Device by Service Members and Veterans With Lower Limb Loss.

INTRODUCTION Telehealth is an increasingly common approach to improve healthcare delivery, especially within the Veterans Health Administration and Department of Defense (DoD). Telehealth has diminished many challenges to direct access for clinical follow-up; however, the use of mobile telehealth for specialty rehabilitative care is emerging and is referred to as telerehabilitation. As early adopters of telehealth, the Veterans Affairs and DoD have supported collaborated efforts for programs designed to increase the access and quality of rehabilitative care while improving the functional ability of our service members (SMs) and veterans with lower limb amputation (LLA). The DoD and Veterans Health Administration collaborated on a Mobile Device Outcomes-based Rehabilitation Program (MDORP) to help injured SMs and veterans with LLA. The MDORP project utilized a mobile health system called the Rehabilitative Lower Limb Orthopedic Accommodating Device (ReLOAD) to assess walking quality. The ReLOAD system includes real-time auditory biofeedback to notify the user of their most prominent gait deviation and then recommends exercises that address specific balance and strength impairments. The purpose of this study was to describe the responses to a postintervention survey evaluating the feasibility and usability of ReLOAD completed by SMs and veterans with LLA who used the system for 5 months. MATERIALS AND METHODS A link to an anonymous usability survey was emailed to all participants who completed MDORP. The survey was modeled after the System Usability Scale, with agreeableness to items rated on a 5-point Likert-style questionnaire in addition to open feedback. Data visualization of Likert-style questionnaires was conducted using ggplot2 and reshape2 statistical packages and was analyzed using R. We obtained institutional review board approval through both Miami Veterans Affairs Healthcare System and Walter Reed National Military Medical Center. RESULTS The majority of participants reported that they would use the system again for home rehabilitation (65%) and that auditory biofeedback helped them walk better (59%). Participants also suggested that future work should include a greater variety of exercise options and the use of smart phones for the ReLOAD application in addition to the iPad tablet. CONCLUSIONS The participants provided positive and constructive feedback that will enhance the value and usability of telerehabilitation interventions like the ReLOAD system for future users.
Real-Time Shadow Detection From Live Outdoor Videos for Augmented Reality

Simulating shadow interactions between real and virtual objects is important for augmented reality (AR), in which accurately and efficiently detecting real shadows from live videos is a crucial step. Most of the existing methods are capable of processing only scenes captured under a fixed viewpoint. In contrast, this article proposes a new framework for shadow detection in live outdoor videos captured under moving viewpoints. The framework splits each frame into a tracked region, which is the region tracked from the previous video frame through optical flow analysis, and an emerging region, which is newly introduced into the scene due to the moving viewpoint. The framework subsequently extracts features based on the intensity profiles surrounding the boundaries of candidate shadow regions. These features are then utilized to both correct erroneous shadow boundaries for the tracked region and to detect shadow boundaries for the emerging region by a Bayesian learning module. To remove spurious shadows, spatial layout constraints are further considered for emerging regions. The experimental results demonstrate that the proposed framework outperforms the state-of-the-art shadow tracking and detection algorithms on a variety of challenging cases in real time, including shadows on backgrounds with complex textures, nonplanar shadows, fast-moving shadows with changing typologies, and shadows cast by nonrigid objects. The quantitative experiments show that our method outperforms the best existing method, achieving a 33.3% increase in the average <inline-formula><tex-math notation="LaTeX">$F_{measure}$</tex-math><alternatives><mml:math><mml:msub><mml:mi>F</mml:mi><mml:mrow><mml:mi>m</mml:mi><mml:mi>e</mml:mi><mml:mi>a</mml:mi><mml:mi>s</mml:mi><mml:mi>u</mml:mi><mml:mi>r</mml:mi><mml:mi>e</mml:mi></mml:mrow></mml:msub></mml:math><inline-graphic xlink:href="xing-ieq1-3041100.gif"/></alternatives></inline-formula> on a self-collected database. Coupled with an image-based shadow-casting method, the proposed framework generates realistic shadow interaction results. This capability will be particularly beneficial for supporting AR applications.
Estimating Population Abundance of Burying Beetles Using Photo-Identification and Mark-Recapture Methods

Abstract Successful conservation and management of protected wildlife populations require reliable population abundance data. Traditional capture-mark-recapture methods can be costly, time-consuming, and invasive. Photographic mark-recapture (PMR) is a cost-effective, minimally invasive way to study population dynamics in species with distinct markings or color patterns. We tested the feasibility and the application of PMR using the software Hotspotter to identify Nicrophorus spp. from digital images of naturally occurring spot patterns on their elytra. We conducted a laboratory study evaluating the identification success of Hotspotter on Nicrophorus americanus (Olivier, 1790) and Nicrophorus orbicollis (Say, 1825) before implementation of a mark-recapture study in situ. We compared the performance of Hotspotter using both ‘high-quality' and ‘low-quality’ photographs. For high-quality photographs, Hotspotter had a false rejection rate of 2.7–3.0% for laboratory-reared individuals and 3.9% for wild-caught individuals. For low-quality photographs, the false rejection rate was much higher, 48.8–53.3% for laboratory-reared individuals and 28.3% for wild-caught individuals. We subsequently analyzed encounter histories of wild-caught individuals with closed population models in Program MARK to estimate population abundance. In our study, we demonstrated the utility of using PMR in estimating population abundance for Nicrophorus spp. based on elytral spot patterns.