High Throughput Screening for SARS-CoV-2 Helicase Inhibitors.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for nearly 7 million deaths worldwide since its outbreak in late 2019. Even with the rapid development and production of vaccines and intensive research, there is still a huge need for specific anti-viral drugs that address the rapidly arising new variants. To address this concern, the National Institute of Allergy and Infectious Diseases (NIAID) established nine Antiviral Drug Discovery (AViDD) Centers, tasked with exploring approaches to target pathogens with pandemic potential, including SARS-CoV-2. In this study, we sought inhibitors of SARS-CoV2 non-structural protein 13 (nsP13) as potential antivirals, first developing a HTS-compatible assay to measure SARS-CoV2 nsP13 helicase activity. Here we present our effort in implementing the assay in a 1,536 well-plate format and in identifying nsP13 inhibitor hit compounds from a ∼650,000 compound library. The primary screen was robust (average Z' = 0.86 ± 0.05) and resulted in 7,009 primary hits. 1,763 of these compounds upon repeated retests were further confirmed, showing consistent inhibition. Following in-silico analysis, an additional orthogonal assay and titration assays, we identified 674 compounds with IC50 <10 µM. We confirmed activity of independent compound batches from de novo powders while also incorporating multiple counterscreen assays. Our study highlights the potential of this assay for use on HTS platforms to discover novel compounds inhibiting SARS-CoV2 nsP13, which merit further development as an effective SARS-CoV2 antiviral.

ALKALINE PHOSPHATASE AND LIVER FIBROSIS AT DIAGNOSIS ARE ASSOCIATED WITH DEEP RESPONSE TO URSODEOXYCHOLIC ACID IN PRIMARY BILIARY CHOLANGITIS.

OBJECTIVE: Primary biliary cholangitis is a chronic and progressive autoimmune liver disease, whose prognosis can be improved by normalizing alkaline phosphatase and bilirubin. While ursodeoxycholic acid (UDCA) is first line standard of care, approximately 40% of patients exhibit incomplete response. We aimed to identify prognostic markers for deep response to UDCA therapy at presentation. PATIENT AND METHODS: Data from the Brazilian Cholestasis Study Group cohort were analyzed retrospectively. Patients were assessed for deep response, defined as normal alkaline phosphatase and bilirubin, after 1 year of UDCA treatment. Additionally, the performance of the UDCA response score in predicting deep response was evaluated. RESULTS: A total of 297 patients were analyzed, with 57.2% achieving an adequate response according to the Toronto criteria, while 22.9% reached deep response. Cirrhosis (OR 0.460; 95% CI 0.225-0.942; p=0.034) and elevated baseline alkaline phosphatase levels (OR 0.629; 95% CI 0.513-0.770; p<0.001) were associated with reduced odds of deep response. The UDCA response score exhibited moderate discrimination power (AUROC=0.769) but lacked calibration. CONCLUSIONS: Baseline ALP and liver fibrosis emerge as the most important prognostic factors to predict normalization of alkaline phosphatase and bilirubin after UDCA. The UDCA response score was inadequate for predicting deep response in the Brazilian PBC population.

Frequencies of chromosomal aberrations and related biochemical parameters in welders.

Stainless steel welders are exposed to heavy filler metals. We evaluated the concentration of these metals in whole blood and urine, and the relevant biochemical parameters in relation to the total chromosomal aberrations (CAs), chromatid-type (CTA-type, CTAs) and chromosome-type (CSA-type, CSAs), in 117 welders and control individuals. Statistically higher concentrations of the total Cr, Ni and Mn were observed in whole blood and urine of welders, and the concentrations were higher in welders who smoked. On the contrary, concentrations of urinary heavy metals Cr and Mn adjusted for creatinine were significantly higher in the control groups. A statistically higher frequency of total CAs was observed in the whole group of welders, and also in the non-smoking welders, as compared to controls. The frequency of total CAs significantly correlated with the concentration of Cr, Ni and Mn in whole blood (R=0.61, P˂0.0001, R=0.33, P˂0.0001 and R=0.66, P˂0.0001, respectively), with urinary concentrations of Ni and Mn (R=0.27, P=0.003 and R=0.28, P=0.003, respectively) and with urinary concentrations of Cr, Ni and Mn adjusted for creatinine (R=0.22, P=0.029, R=0.26, P=0.005 and R=0.20, P=0.030, respectively). Likewise, the frequency of CTA-types significantly correlated with the concentration of Cr and Mn in whole blood (R=0.31, P=0.0007 and R=0.34, P=0.0002). The frequency of CSA-types significantly correlated with concentrations of Cr, Ni and Mn in whole blood (R=0.43, P˂0.0001, R=0.38, P˂0.0001 and R=0.46, P˂0.0001, respectively). The statistically higher values of serum creatinine and total bilirubin were detected in all welders, as well as in smokers when compared to the corresponding controls. The exposure to heavy metals in welders increased the frequencies of CAs and altered the balance between urinary excretion of heavy metals and their possible accumulation.

Postmortem biochemistry of GFAP, NSE and S100B in cerebrospinal fluid and in vitreous humor for estimation of postmortem interval: a pilot study.

Postmortem interval (PMI) is a challenging issue in forensic practice. Although postmortem biomarkers of traumatic brain injury (TBI) are recognised as an emerging resource for PMI estimation, their role remains controversial. This study aims to evaluate postmortem concentrations of three TBI biomarkers (GFAP, NSE and S100B) in two matrices (cerebrospinal fluid and vitreous humor), in order to find out if these markers could be adopted in PMI estimation. Thirty-five deceased individuals with known PMI who underwent forensic autopsy at the University of Parma were examined. Matrices were collected during autopsy, then biomarker concentrations were determined through the enzyme-linked immunosorbent assay. Statistical significance of the data in relation to PMI was studied. The correlation of biomarkers with PMI, examined with samples divided into six groups according to the number of days elapsed since death, was not statistically significant, although S100B in cerebrospinal fluid showed an increasing trend in cases from 1 to 5 days of PMI. Comparison between cases with 1 day of PMI and those with 2 or more days of PMI showed a statistically significant correlation for GFAP and NSE in cerebrospinal fluid. GFAP and NSE in cerebrospinal fluid represent appropriate biomarkers in PMI estimation to distinguish cases with one day of PMI from those with two or more days of PMI. The current study was limited by the scarcity of the cohort and the narrow spectrum of cases. Further research is needed to confirm these observations.

Current advances in the development of bioluminescent probes toward spatiotemporal trans-scale imaging.

Bioluminescence imaging has recently attracted great attention as a highly sensitive and non-invasive analytical method. However, weak signal and low chemical stability of the luciferin are conventional drawbacks of bioluminescence imaging. In this review article, we describe the recent progress on the development and applications of bioluminescent probes for overcoming the aforementioned limitations, thereby enabling spatiotemporal trans-scale imaging. The detailed molecular design for manipulation of their luminescent properties and functions enabled a variety of applications, including in vivo deep tissue imaging, long-term imaging, and chemical sensor.

Effects of dietary stevia extract supplementation on growth performance, serum biochemical indices and intestinal health of Yellow-Feathered Broilers.

Stevia, a perennial shrub from the genus Stevia in the Asteraceae family, contains active ingredients like chlorogenic acid and shows promise as a natural feed additive. Despite this potential, there is limited research on the impact of stevia extract specifically on yellow-feather broilers. The study aimed to evaluate the effects of dietary stevia extract with varying concentrations of chlorogenic acid on the growth performance, serum biochemical indices, and intestinal health of yellow-feathered broilers. A total of 425 one-day-old female yellow-feathered broilers were randomly allocated into five treatment groups with five replicates of 17 broilers each, and the feeding trial lasted 63 days. The groups included a control and those supplemented with stevia extract at concentrations of 100 mg/kg, 200 mg/kg, 300 mg/kg, and 400 mg/kg. Results showed that adding 100 mg/kg of stevia extract to the basal diet significantly increased the daily weight gain (ADG) of the broilers, while reducing the average daily feed intake (ADFI) and feed conversion ratio (F/G). However, supplementation with stevia extract at concentrations up to 300 mg/kg led to decreased final weight and ADG. Conversely, dietary supplementation with 100-200 mg/kg of stevia extract improved serum antioxidant capacity and reduced serum total cholesterol levels compared to the control group. Additionally, the cecum n-butyric acid level was significantly higher in the 200 mg/kg stevia extract group than in the control group. In conclusion, supplementing yellow-feathered broilers' diets with stevia extract can enhance growth performance, antioxidant and immune capacity, and intestinal health. The optimal concentration of stevia extract for these benefits is between 100 mg/kg and 200 mg/kg.

Effectiveness and biochemical impact of ozone gas and silica nanoparticles on Culex pipiens (Diptera: Culicidae).

Culex pipiens (Diptera: Culicidae) is a vector of many serious human diseases, and its control by the heavy use of chemical insecticides has led to the evolution of insecticide resistance and high environmental risks. Many safe alternatives, such as ozone gas (O3) and silica nanoparticles (silica NPs) can reduce these risks. Therefore, O3 and silica NPs were applied to 3rd larval instars of Cx. pipiens at different concentrations (100, 200, and 400 ppm) for different exposure times (1, 2, 3, and 5 min for O3 and 24, 48, and 72 h for silica NPs). The activity of some vital antioxidant enzymes as well as scanning electron microscopy of the body surface were also investigated. A positive correlation was observed between larval mortality % and the tested concentrations of O3 and silica NPs. O3 was more effective than silica NPs, it resulted in 92% mortality at 400 ppm for a short exposure time (5 min). O3-exposed larvae exhibited a significant increase in glutathione peroxidase, glutathione S-transferase, and catalase activities as well as the total antioxidant capacity. Scanning electron microscopy showing disruptive effects on the body surface morphology of ozone and silica NPs treated larvae. These results provide evidence that O3 and silica NPs have the potential for use as alternative vector control tools against Cx. pipiens.

Promoter role of putrescine for molecular and biochemical processes under drought stress in barley.

Drought, which adversely affects plant growth and continuity of life and reduces product yield and quality, is one of the most common abiotic stresses at the globally. One of the polyamines that regulates plant development and reacts to abiotic stressors, including drought stress, is Putrescine (Put). This study compared the physiological and molecular effects of applying exogenous Put (10 µM) to barley (Hordeum vulgare cv. Burakbey) under drought stress (- 6.30 mPa PEG 6000). The 21-day drought stress imposed on the barley plant had a strong negative effect on plant metabolism in all experimental groups. Exogenous Put treatment under drought stress had a reformative effect on the cell cycle (transitions from G0-G1 to S and from S to G2-M), total protein content (almost 100%), endogenous polyamine content, malondialdehyde (MDA) (70%), and ascorbate peroxidase (APX) (62%) levels compared to the drought stress plants. Superoxide dismutase (SOD) (12%) and catalase (CAT) (32%) enzyme levels in the same group increased further after exogenous Put application, forming a response to drought stress. Consequently, it was discovered that the administration of exogenous Put in barley raises endogenous polyamine levels and then improves drought tolerance due to increased antioxidant capability, cell division stimulation, and total protein content.

Duddingtonia flagrans and its crude proteolytic extract: concomitant action in sheep coprocultures.

The presence of infective larvae (L3) of gastrointestinal nematode (GIN) parasites in pastures directly contributes to the constant recurrence of infections in ruminant herds. This study aimed to evaluate the nematophagous fungus Duddingtonia flagrans (AC001) (proteolytic crude extract and/or conidia) in the in vitro control of GIN L3 in coprocultures. To produce the proteolytic crude extract, a suspension (107 conidia/mL) of D. flagrans was inoculated into a liquid medium. After 6 days, the medium was filtered, centrifuged, and its proteolytic activity was measured. For the experimental assay, fecal samples were collected directly from the rectal ampulla of naturally infected sheep, and egg counts per gram of feces (EPG) were performed. Coprocultures were prepared using 10 g of fecal material with the groups defined as follows: control group G1 (1.0 mL of denatured proteolytic crude extract); treated group G2 (1.0 mL of active proteolytic crude extract); treated group G3 (1.0 mL of active proteolytic crude extract + 1.0 mL of AC001 conidia). The coprocultures were maintained at room temperature (25ºC), for 7 days, and then the L3 larvae were recovered. The results demonstrated that AC001 successfully produced protease (56.34 U/mL). The treatments with active proteolytic crude extract (G2) and active proteolytic crude extract + AC001 conidia (G3) were significantly different (p < 0.01) from the control group with denatured proteolytic crude extract (G1). AC001 and its proteolytic crude extract acted concomitantly on helminths directly in the fecal environment, suggesting potential future applications in the field.

Utility of 64Cu-Sarcophagine-Bombesin PET/CT in Men with Biochemically Recurrent Prostate Cancer and Negative or Equivocal Findings on 68Ga-PSMA-11 PET/CT.

Despite a high detection rate of 68Ga-prostate-specific membrane antigen (PSMA) PET/CT in biochemical recurrence (BCR) of prostate cancer, a significant proportion of men have negative 68Ga-PSMA-11 PET/CT results. Gastrin-releasing peptide receptor, targeted by the copper-chelated bombesin analog 64Cu-sarcophagine-bombesin (SAR-BBN) PET/CT, is also overexpressed in prostate cancer. In this prospective imaging study, we investigate the detection rate of 64Cu-SAR-BBN PET/CT in patients with BCR and negative or equivocal 68Ga-PSMA-11 PET/CT results. Methods: Men with confirmed adenocarcinoma of the prostate, prior definitive therapy, and BCR (defined as a prostate-specific antigen [PSA] level > 0.2 ng/mL) with negative or equivocal 68Ga-PSMA-11 PET/CT results within 3 mo were eligible for enrollment. 64Cu-SAR-BBN PET/CT scans were acquired at 1 and 3 h after administration of 200 MBq of 64Cu-SAR-BBN, with further delayed imaging undertaken optionally at 24 h. PSA (ng/mL) was determined at baseline. All PET (PSMA and bombesin) scans were assessed visually. Images were read with masking of the clinical results by 2 experienced nuclear medicine specialists, with a third reader in cases of discordance. Accuracy was defined using a standard of truth that included biopsy confirmation, confirmatory imaging, or response to targeted treatment. Results: Twenty-five patients were enrolled. Prior definitive therapy was radical prostatectomy (n = 24, 96%) or radiotherapy (n = 1, 4%). The median time since definitive therapy was 7 y (interquartile range [IQR], 4-11 y), and the Gleason score was 7 or less (n = 15, 60%), 8 (n = 3, 12%), or 9 (n = 7, 28%). The median PSA was 0.69 ng/mL (IQR, 0.28-2.45 ng/mL). Baseline PSMA PET scans were negative in 19 patients (76%) and equivocal in 6 (24%). 64Cu-SAR-BBN PET-avid disease was identified in 44% (11/25): 12% (3/25) with local recurrence, 20% (5/25) with pelvic node metastases, and 12% (3/25) with distant metastases. The κ-score between readers was 0.49 (95% CI, 0.16-0.82). Patients were followed up for a median of 10 mo (IQR, 9-12 mo). Bombesin PET/CT results were true-positive in 5 of 25 patients (20%), false-positive in 2 of 25 (8%), false-negative in 7 of 25 (28%), and unverified in 11 of 25 (44%). Conclusion: 64Cu-SAR-BBN PET/CT demonstrated sites of disease recurrence in 44% of BCR cases with negative or equivocal 68Ga-PSMA-11 PET/CT results. Further evaluation to confirm diagnostic benefit is warranted.

Suitability of raw and heat-treated Amaranthus spinosus in broiler diets: Effects on growth performance, meat antioxidant capacity, haemato-biochemical parameters, intestinal histomorphometry, and cecal volatile fatty acid profile.

This study aimed to examine the effects of incorporating amaranth (Amaranthus spinosus, either raw or heat-treated) into broiler diets on growth performance, meat antioxidant capacity, haemato-biochemical parameters, intestinal histomorphometry, and cecal volatile fatty acid profile. A total of 210 male Ross 308 broiler chicks were allocated to five dietary treatments in a completely randomized design, with each treatment comprising six replicates of seven birds each. The control group received a diet based on maize and soybean meal, while the remaining dietary groups were formulated to be isonitrogenous and isocaloric to the control, with exact levels of 10% and 20% raw or heat-treated amaranth in the diet. Body weight and feed intake were monitored on days 0, 10, 24, and 39 of the study. On day 39, two birds per replicate were randomly selected for blood sampling, followed by slaughtering for further parameter examination. Incorporating A. spinosus up to 20% in broiler diets had no adverse effect on body weight gain compared to the control. However, higher levels of amaranth led to a negative impact on the feed conversion ratio, attributed to increased feed intake. Furthermore, amaranth supplementation did not negatively influence carcass yield or various organ weights, except for the gizzard, which was heavier in the amaranth-fed groups. Notably, amaranth supplementation reduced abdominal fat, enhanced meat antioxidant status, and had no detrimental effects on blood biochemical or hematological indices. Additionally, amaranth feeding resulted in decreased blood triglyceride levels but had no effect on cholesterol levels. While heat treatment of amaranth did not significantly alter the performance of broiler chickens, it enhanced the beneficial effects of amaranth feeding on the histomorphological features of the duodenum and ileum, and increased blood IgG levels. The cecal volatile fatty acid profile remained largely unaffected by amaranth inclusion, although heat-treated amaranth led to increased levels of branched-chain fatty acids and valerate. Overall, the findings suggest A. spinosus as a promising alternative feed ingredient for broilers when included at 10% of the diet. However, further research is needed to investigate the effect of various amaranth species, processing methods and enzyme supplementation on poultry nutrition to expand its inclusion rate.

Correlated changes in stress resistance and biochemical parameters in response to long-term protein restriction in Drosophila melanogaster.

Studies in fruit flies, Drosophila melanogaster, have observed considerable variation in the effect of dietary protein restriction (PR) on various fitness traits. In addition, not only are there inconsistent results relating lifespan to stress resistance, but also the long-term effects of PR are unexplored. We study PR implementation across generations (long term) hypothesizing that it will be beneficial for fitness traits, stress resistance and storage reserves due to nutritional plasticity transferred by parents to offspring in earlier Drosophila studies. By imposing two concentrations of PR diets (50% and 70% of control protein) from the pre-adult and adult (age 1 day) stages of the flies, we assessed the stage-specific and long-term effect of the imposed PR. All long-term PR flies showed increased resistance against the tested stressors (starvation, desiccation, H2O2-induced oxidative stress). In addition, we also found long-term PR-induced increased stress resistance across generations. The PR flies also possessed higher protein and triglyceride (TG) content, reduced glucose and unaffected glycogen levels. We also assayed the effect of returning the PR flies to control (AL) food for a single generation and assessed their biochemical parameters to witness the transient PR effect. It was seen that TG content upon reversal was similar to AL flies except for PRI70 males; however, the glucose levels of PR males increased, while they were consistently lower in females. Taken altogether, our study suggests that long-term PR implementation contributes to increased stress resistance and was found to influence storage reserves in D. melanogaster.

Undetectable PSA predicts outcome after salvage radiotherapy for biochemical recurrence following radical prostatectomy.

BACKGROUND AND PURPOSE: Salvage radiotherapy (SRT) is a curative treatment option in patients with biochemical recurrence after radical prostatectomy (RP). Undetectable prostate-specific antigen (PSA) < 0.1 ng/mL following SRT predicts biochemical progression-free survival (BPFS). The aim of this large retrospective study was to evaluate whether this effect persists in an extended follow-up of >5 years. MATERIALS AND METHODS: A total of 678 patients treated with SRT for biochemical recurrence after RP were included. Exclusion criteria were lymph node or distant metastases, pre-SRT PSA > 3 ng/mL, and receipt of androgen deprivation therapy (ADT) between RP and SRT. All patients received a median dose of 70.2 (range 59.4-72.0) Gy to the prostatic fossa. The log-rank test (Kaplan-Meier) and Cox regression analysis were used to evaluate the impact of disease- and treatment-related parameters on BPFS, metastasis-free survival (MFS), and overall survival (OS). RESULTS: Median follow-up after SRT was 5.6 (range 0.1-14.5) years. The 5-year BPFS was 77.8 % in patients with a PSA nadir < 0.1 ng/mL (undetectable) and 16.3 % in the remaining cohort (p < 0.001). Five-year MFS was 95.3 % with undetectable PSA versus 84.0 % with detectable PSA (p < 0.001), and 5-year OS values were 97.5 % and 92.7 % with undetectable versus detectable PSA, respectively (p = 0.04). In multivariate analysis, undetectable PSA was the strongest predictor of BPFS (HR = 0.122; 95 %CI: 0.080-0.187; p < 0.001) and MFS (HR = 0.262; 95 %CI: 0.136-0.594; p < 0.001), but was not significant for OS (HR = 0.615; 95 %CI: 0.298-1.269; p = 0.189). CONCLUSION: PSA < 0.1 ng/mL following SRT without ADT is a significant predictor of BPFS and MFS. The results suggest that it might be feasible to withhold ADT in selected patients if they have undetectable PSA after SRT. Prospective studies are warranted to confirm these findings.

Improved biochemical and neurodevelopmental profiles with high-dose hydroxocobalamin therapy in cobalamin C defect.

Cobalamin C (Cbl-C) defect causes methylmalonic acidemia, homocystinuria, intellectual disability and visual impairment, despite treatment adherence. While international guidelines recommend parenteral hydroxocobalamin (OH-Cbl) as effective treatment, dose adjustments remain unclear. We assessed OH-Cbl therapy impact on biochemical, neurocognitive and visual outcomes in early-onset Cbl-C patients treated with different OH-Cbl doses over 3 years. Group A (n = 5), diagnosed via newborn screening (NBS), received high-dose OH-Cbl (median 0.55 mg/kg/day); Group B1 (n = 3), NBS-diagnosed, received low-dose OH-Cbl (median 0.09 mg/kg/day); Group B2 (n = 12), diagnosed on clinical bases, received low-dose OH-Cbl (median 0.06 mg/kg/day). Biochemical analyses revealed better values of homocysteine, methionine and methylmalonic acid in Group A compared to Group B1 (p < 0.01, p < 0.05 and p < 0.01, respectively) and B2 (p < 0.001, p < 0.01 and p < 0.001, respectively). Neurodevelopmental assessment showed better outcome in Group A compared to low-dose treated Groups B1 and B2, especially in Developmental Quotient, Hearing and Speech and Performance subscales without significant differences between Group B2 and Group B1. Maculopathy was detected in 100%, 66% and 83% of patients in the three groups, respectively. This study showed that "high-dose" OH-Cbl treatment in NBS-diagnosed children with severe early-onset Cbl-C defect led to a significant improvement in the metabolic profile and in neurocognitive outcome, compared to age-matched patients treated with a "low-dose" regimen. Effects on maculopathy seem unaffected by OH-Cbl dosage. Our findings, although observed in a limited number of patients, may contribute to improve the long-term outcome of Cbl-C patients.

Interobserver and intraobserver agreement in PET/CT with [18F]DCFPyL according to TNM molecular and PSMA-RADS 2.0 criteria.

PURPOSE: The aim of this study was to determine the agreement between three observers with different levels of experience using the PSMA-RADS 2.0 criteria and the miTNM system for the interpretation of PET-PSMA with [18F]DCFPyL in males with prostate cancer. MATERIALS AND METHODS: PET-PSMA images from 114 prostate cancer patients were blindly reported twice by three different observers at intervals of 8 weeks. The evaluations were performed according to the molecular imaging TNM (miTNM) and PSMA-RADS 2.0 criteria. We used Fleiss' Kappa to analyse inter and intraobserver agreements. RESULTS: Moderate overall agreement was obtained in the assessment of the PET-PSMA results (Fleiss'k = 0.53; 95% CI 0.45-0.62; p < 0.001), with significant agreement in the miT, miN and miM reports. There was a substantial level of agreement in the reporting of prostatic disease and lymphatic involvement (Fleiss'k = 0.66 and 0.65), being lower than that observed in the reporting of metastatic disease (Fleiss'k = 0.86), especially in the M0 group (Fleiss'k = 0.99). Upon re-evaluation of the images, observer 1 had moderate overall agreement for miT (Fleiss'k = 0.51) and substantial agreement for miN and miM (Fleiss'k 0.75 and 0.63, respectively). CONCLUSIONS: The use of a structured scoring system such as PSMA-RADS 2.0, as well as the miTNM classification system in the interpretation of PET-PSMA images in prostate cancer patients, provides a highly reproducible report format. High levels of interobserver and intraobserver agreement are found, especially when ruling out disease, which supports its use in routine clinical practice.

Monitoring of parasite Orobanche cumana using Vis-NIR hyperspectral imaging combining with physio-biochemical parameters on host crop Helianthus annuus.

KEY MESSAGE: This study provided a non-destructive detection method with Vis-NIR hyperspectral imaging combining with physio-biochemical parameters in Helianthus annuus in response to Orobanche cumana infection that took insights into the monitoring of sunflower weed. Sunflower broomrape (Orobanche cumana Wallr.) is an obligate weed that attaches to the host roots of sunflower (Helianthus annuus L.) leading to a significant reduction in yield worldwide. The emergence of O. cumana shoots after its underground life-cycle causes irreversible damage to the crop. In this study, a fast visual, non-invasive and precise method for monitoring changes in spectral characteristics using visible and near-infrared (Vis-NIR) hyperspectral imaging (HSI) was developed. By combining the bands sensitive to antioxidant enzymes (SOD, GR), non-antioxidant enzymes (GSH, GSH + GSSG), MDA, ROS (O2-, OH-), PAL, and PPO activities obtained from the host leaves, we sought to establish an accurate means of assessing these changes and conducted imaging acquisition using hyperspectral cameras from both infested and non-infested sunflower cultivars, followed by physio-biochemical parameters measurement as well as analyzed the expression of defense related genes. Extreme learning machine (ELM) and convolutional neural network (CNN) models using 3-band images were built to classify infected or non-infected plants in three sunflower cultivars, achieving accuracies of 95.83% and 95.83% for the discrimination of infestation as well as 97.92% and 95.83% of varieties, respectively, indicating the potential of multi-spectral imaging systems for early detection of O. cumana in weed management.

Transcriptional, biochemical, and histochemical response of resistant and susceptible cultivars of Brassica juncea against Albugo candida infection.

White rust disease caused by a biotrophic oomycete Albugo candida is one of the most serious impediments in realizing the production potential of Brassica juncea. Due to the obligate nature of the pathogen, R-gene-based resistance is unstable as the newer virulent races emerge quickly. For this, a deep understanding of the molecular basis of resistance is essential for developing durable resistant varieties. In this study, we selected one susceptible cultivar, 'Pusa Jaikisan' and its single R gene based resistant NIL, 'Pusa Jaikisan WRR as the source of understanding the defense mechanism in B. juncea against A. candida. Comparative histochemical analysis at 12 dpi showed higher callose deposition in the resistant cultivar than in the susceptible which hints towards its possible role in defense mechanism. Based on the biochemical markers observation, total protein was found to have a negative correlation with the resistance. The antioxidant enzymes (POX, CAT, and SOD) and non-enzymatic ROS scavenging compounds such as polyphenols and proline showed a positive correlation with the white rust resistance. Polyphenol Oxidase (PPO) total chlorophyll and total carotenoids were also found to be more abundant in the 'Pusa Jaikisan WRR'. Based on the heat map analysis, PAL was identified to be the comparatively most induced enzyme involved in the defense mechanism. The polyphenol oxidase, total chlorophyll and total carotenoids were also found to show higher activity in the 'Pusa Jaikisan WRR'. Furthermore, to study the defense response of 'Pusa Jaikisan WRR' compared to 'Pusa Jaikisan' against A. candida infection, the gene expression analyses of salicylic acid (SA)-marker PR protein genes (PR1 and PR2) and jasmonic acid (JA)-marker PR protein genes (PR3 and PR12) were done by qRT-PCR. Based on the results, PR2 emerged as the best possible gene for defense against A. candida followed by PR1. PR3 and PR12 also showed positive correlation with the disease resistance which may be due to the JA pathway acting complementary to the SA pathway in case of B. juncea-A. candida interaction. This provides evidence for the JA-SA hormonal crosstalk to be synergistic in case of the white rust resistance.

SERPINH1 modulates apoptosis by inhibiting P62 ubiquitination degradation to promote bone metastasis of prostate cancer.

Prostate cancer (PCa) is one of the most prevalent urogenital malignancies. Bone metastasis from PCa reduces patient survival rates significantly. There currently exists no effective treatment for bone metastatic PCa, and the underlying mechanisms remain unclear. This study performed transcriptomic screening on PCa bone metastasis specimens and intersection analysis in public databases and identified SERPINH1 as a potential target for treatment. SERPINH1 was found to be upregulated in PCa bone metastases and with poor prognosis, high Gleason score, and advanced metastatic status. SERPINH1 induced PCa cells' bone metastasis in vivo, promoted their proliferation, and mitigated apoptosis. Mechanistically, SERPINH1 bound to P62, reducing TRIM21-mediated K63-linked ubiquitination degradation of P62 and promoting proliferation and resistance to apoptosis of PCa. This study suggests the regulation of ubiquitination degradation of P62 by SERPINH1 that promotes PCa bone metastasis and can be considered as a potential target for treatment of bone metastatic PCa.

Short-term impact of vitamin K2 supplementation on biochemical parameters and lipoprotein fractions.

This study explored the short-term effects of vitamin K2 (VK2) supplementation on biochemical parameters (vitamin D, vitamin E, vitamin A, alkaline phosphatase, calcium, phosphorus (P), magnesium, metallothionein, triglycerides, cholesterol, high-density lipoprotein (HDL), low-density lipoprotein (LDL), and lipoprotein fractions (albumin, HDL, very low-density lipoprotein (VLDL), LDL, and chylomicrons). A short-term experiment (24 h, six probands) was performed to track changes in VK2 levels after a single-dose intake (360 µg/day). Liquid chromatography-tandem mass spectrometry was used to monitor vitamin K levels (menaquinone-4 (MK-4), menaquinone-7 (MK-7), and vitamin K1 [VK1]) with a limit of detection of 1.9 pg/mL for VK1 and 3.8 pg/mL for the two forms of VK2. Results showed that MK-7 levels significantly increased within 2-6 h post-administration and then gradually declined. MK-4 levels were initially low, showing a slight increase, whereas VK1 levels rose initially and then decreased. Biochemical analyses indicated no significant changes in sodium, chloride, potassium, calcium, magnesium, albumin, or total protein levels. A transient increase in P was observed, peaking at 12 h before returning to baseline. Agarose gel electrophoresis of lipoprotein fractions revealed distinct chylomicron bands and variations in VLDL and HDL mobility, influenced by dietary lipids and VK2 supplementation. These findings suggest effective absorption and metabolism of MK-7 with potential implications for bone metabolism and cardiovascular health.

Hepatoprotective activity of Mentha rotundifolia aqueous extract against hepatocellular damage induced by CCL4 in rats.

This study aims to assess hepatoprotective properties of M. rotundifolia. Decoction was used to prepare the aqueous extract. The preliminary cytotoxicity evaluated against Caco 2 and RAW 264 cells demonstrate no cytotoxic effect. The preventive impact of the extract against liver damage was evaluated by examining blood levels of AST, ALT, ALP, total proteins, and histological alterations in liver tissues. Thirty albino rats were separated into five groups: the first served as normal group, the second was injected by olive oil (3 ml/kg), and the third was injected by CCL4 (3 ml/kg). However, groups IV and V received daily doses of 250 and 500 mg extract/kg bw, respectively before CCL4 injection. The results showed that the administration of the extract led to a marked improvement in plasma biochemical markers and a reduction in symptoms of CCL4-induced liver damage. The extract exhibits hepatoprotective activity, which may be attributed to its phytochemical components.