Engineered CRISPRa System for Precise and Multiplex Gene Regulation Using a Hybrid dCas12a Variant and Hairpin-Spacer crRNAs.

Clustered regularly interspaced short palindromic repeats (CRISPR)-Cas12a nucleases have emerged as a promising alternative to CRISPR-Cas9 in gene editing and expression regulation. However, the adoption of Cas12a has been hindered due to general off-target activities and limited efficiency. Here, we utilized a hybrid engineered Cas12a variant and hairpin-spacer crRNAs (h-CAP) to enhance the specificity and efficiency of the CRISPR-Cas12a system. Leveraging the h-CAP strategy, we demonstrate both single-base-specific and multiplex gene expression regulation in human cells.

Single-Cell Study Unveils Lead Lifespan in Blood Cell Populations Follows a Universal Lognormal Distribution with Individual Skewness.

Lead is a widespread environmental hazard that can adversely affect multiple biological functions. Blood cells are the initial targets that face lead exposure. However, a systematic assessment of lead dynamics in blood cells at single-cell resolution is still absent. Herein, C57BL/6 mice were fed with lead-contaminated food. Peripheral blood was harvested at different days. Extracted red blood cells and leukocytes were stained with 19 metal-conjugated antibodies and analyzed by mass cytometry. We quantified the time-lapse lead levels in 12 major blood cell subpopulations and established the distribution of lead heterogeneity. Our results show that the lead levels in all major blood cell subtypes follow lognormal distributions but with distinctively individual skewness. The lognormal distribution suggests a multiplicative accumulation of lead with stochastic turnover of cells, which allows us to estimate the lead lifespan of different blood cell populations by calculating the distribution skewness. These findings suggest that lead accumulation by single blood cells follows a stochastic multiplicative process.

Multiomics blueprint of PANoptosis in deciphering immune characteristics and prognosis stratification of glioma patients.

BACKGROUND: As the most prevalent primary brain tumor in adults, glioma accounts for the majority of all central nervous system malignant tumors. The concept of PANoptosis is a relatively new, underlining the interconnection and synergy among three distinct pathways: pyroptosis, apoptosis and necroptosis. METHODS: We performed single-cell annotations of glioma cells and determined crucial signaling pathways through cell chat analysis. Using least absolute shrinkage and selection operator (LASSO) and Cox analyses, we identified a gene set with prognostic values. Our model was validated using independent external cohort. In addition, we employed single-sample gene set enrichment analysis and xCell analyses to describe the detailed profile of infiltrated immune cells and depicted the gene mutation landscape in the two groups. RESULTS: We identified seven distinct cell clusters in glioma samples, including oligodendrocyte precursor cells (OPCs), myeloid cells, tumor cells, oligodendrocytes, astrocytes, vascular cells and neuronal cells. We found that myeloid cells showed the highest PANoptosis activity. An intense mutual cell communication pattern between the tumor cells and OPCs and oligodendrocytes was observed. Differentially expressed genes between the high-PANoptosis and low-PANoptosis cell groups were obtained, which were enriched to actin cytoskeleton, cell adhesion molecules and gamma R-mediated phagocytosis pathways. We determined a set of five genes of prognostic significance: SAA1, SLPI, DCX, S100A8 and TNR. The prognostic differences between the two groups in the internal and external sets were found to be statistically significant. We found a marked correlation between S100A8 and activated dendritic cell, macrophage, mast cell, myeloid derived suppressor cell and Treg infiltration. Moreover, we have observed a significant increase of PTEN mutation in the high risk (HR) group of glioma patients. CONCLUSIONS: In the present study, we have constructed a prognostic model that is based on the PANoptosis, and we have demonstrated its significant efficacy in stratifying patients with glioma. This innovative prognostic model offers novel insights into precision immune treatments that could be used to combat this disease and improve patient outcomes, thereby providing a new avenue for personalized treatment options.

L-arginine metabolism inhibits arthritis and inflammatory bone loss.

OBJECTIVES: To investigate the effect of the L-arginine metabolism on arthritis and inflammation-mediated bone loss. METHODS: L-arginine was applied to three arthritis models (collagen-induced arthritis, serum-induced arthritis and human TNF transgenic mice). Inflammation was assessed clinically and histologically, while bone changes were quantified by µCT and histomorphometry. In vitro, effects of L-arginine on osteoclast differentiation were analysed by RNA-seq and mass spectrometry (MS). Seahorse, Single Cell ENergetIc metabolism by profilIng Translation inHibition and transmission electron microscopy were used for detecting metabolic changes in osteoclasts. Moreover, arginine-associated metabolites were measured in the serum of rheumatoid arthritis (RA) and pre-RA patients. RESULTS: L-arginine inhibited arthritis and bone loss in all three models and directly blocked TNFα-induced murine and human osteoclastogenesis. RNA-seq and MS analyses indicated that L-arginine switched glycolysis to oxidative phosphorylation in inflammatory osteoclasts leading to increased ATP production, purine metabolism and elevated inosine and hypoxanthine levels. Adenosine deaminase inhibitors blocking inosine and hypoxanthine production abolished the inhibition of L-arginine on osteoclastogenesis in vitro and in vivo. Altered arginine levels were also found in RA and pre-RA patients. CONCLUSION: Our study demonstrated that L-arginine ameliorates arthritis and bone erosion through metabolic reprogramming and perturbation of purine metabolism in osteoclasts.

A comparison of hemodynamic measurement methods during orthotopic liver transplantation: evaluating agreement and trending ability of PiCCO versus pulmonary artery catheter techniques.

BACKGROUND: Significant hemodynamic changes occur during liver transplantation, emphasizing the importance of precious and continuous monitoring of cardiac output, cardiac index, and other parameters. Although the monitoring of cardiac output by pulse indicator continuous cardiac output (PiCCO) was statistically homogeneous compared to the clinical gold standard pulmonary artery catheterization (PAC) in previous studies of liver transplantation, there are fewer statistical methods for the assessment of its conclusions, and a lack of comparisons of other hemodynamic parameters (e.g., SVRI, systemic vascular resistance index). Some studies have also concluded that the agreement between PiCCO and PAC is not good enough. Overall, there are no uniform conclusions regarding the agreement between PiCCO and PAC in previous studies. This study evaluates the agreement and trending ability of relevant hemodynamic parameters obtained with PiCCO compared to the clinical gold standard PAC from multiple perspectives, employing various statistical methods. METHODS: Fifty-two liver transplantation patients were included. Cardiac output (CO), cardiac index (CI), SVRI and stroke volume index (SVI) values were monitored at eight time points using both PiCCO and PAC. The results were analyzed by Bland-Altman analysis, Passing-bablok regression, intra-class correlation coefficient (ICC), 4-quadrant plot, polar plot, and trend interchangeability method (TIM). RESULTS: The Bland-Altman analysis revealed high percentage errors for PiCCO: 54.06% for CO, 52.70% for CI, 62.18% for SVRI, and 51.97% for SVI, indicating poor accuracy. While Passing-Bablok plots showed favorable agreement for SVRI overall and during various phases, the agreement for other parameters was less satisfactory. The ICC results confirmed good overall agreement between the two devices across most parameters, except for SVRI during the new liver phase, which showed poor agreement. Additionally, four-quadrant and polar plot analyses indicated that all agreement rate values fell below the clinically acceptable threshold of over 90%, and all angular deviation values exceeded ± 5°, demonstrating that PiCCO is unable to meet the acceptable trends. Using the TIM, the interchangeability rates were found to be quite low: 20% for CO and CI, 16% for SVRI, and 13% for SVI. CONCLUSIONS: Our study revealed notable disparities in absolute values of CO, CI, SVRI and SVI between PiCCO and PAC in intraoperative liver transplant settings, notably during the neohepatic phase where errors were particularly pronounced. Consequently, these findings highlight the need for careful consideration of PiCCO's advantages and disadvantages in liver transplantation scenarios, including its multiple parameters (such as the encompassing extravascular lung water index), against its limited correlation with PAC.

Effect of oxide interactions on chromium speciation transformation during simulated municipal solid waste incineration.

Chromium released during municipal solid waste incineration (MSWI) is toxic and carcinogenic. The removal of chromium from simulated MSWI flue gas by four sorbents (CaO, bamboo charcoal (BC), powdered activated carbon (PAC), and Al2O3) and the effects of four oxides (SiO2, Al2O3, Fe2O3, and CaO) on chromium speciation transformation were investigated. The results showed that the removal rates of total Cr by the four sorbents were Al2O3 < CaO < PAC < BC, while the removal rates of Cr(VI) by the four sorbents were Al2O3 < PAC < BC < CaO. CaO had a strong oxidizing effect on Cr(III), while BC and PAC had a better-reducing effect on Cr(VI). SiO2 was better for the reduction of Na2CrO4 and K2CrO4 above 1000°C due to its strong acidity, and the addition of CaO significantly inhibited the reduction of Cr(VI). MgCrO4 decomposed above 700°C to form MgCr2O4, and the reaction between MgCrO4 and oxides also existed in the form of a more stable trivalent spinel. Furthermore, when investigating the effect of oxides on the oxidation of Cr(III) in CrCl3, it was discovered that CaO promoted the conversion of Cr(III) to Cr(VI), while the presence of chlorine caused chromium to exist in the form of Cr(V), and increasing the content of CaO and extending the heating time facilitated the oxidation of Cr(III). In addition, silicate, aluminate, and ferrite were generated after the addition of SiO2, Al2O3, and Fe2O3, which reduced the alkalinity of CaO and had an important role in inhibiting the oxidation of Cr(III). The acidic oxides can not only promote the reduction of Cr(VI) but also have an inhibitory effect on the oxidation of Cr(III) ascribed to alkali metals/alkaline earth metals, and the proportion of acidic oxides can be increased moderately to reduce the generation of harmful substances in the hazardous solid waste heat treatment.

A randomized-controlled trial of ischemia-free liver transplantation for end-stage liver disease.

BACKGROUND & AIMS: Ischemia-reperfusion injury (IRI) has thus far been considered as an inevitable component of organ transplantation, compromising outcomes, and limiting organ availability. Ischemia-free organ transplantation is a novel approach designed to avoid IRI, with the potential to improve outcomes. METHODS: In this randomized-controlled clinical trial, recipients of livers from donors after brain death were randomly assigned to receive either an ischemia-free or a 'conventional' transplant. The primary endpoint was the incidence of early allograft dysfunction. Secondary endpoints included complications related to graft IRI. RESULTS: Out of 68 randomized patients, 65 underwent transplants and were included in the analysis. 32 patients received ischemia-free liver transplantation (IFLT), and 33 received conventional liver transplantation (CLT). Early allograft dysfunction occurred in two recipients (6%) randomized to IFLT and in eight (24%) randomized to CLT (difference -18%; 95% CI -35% to -1%; p = 0.044). Post-reperfusion syndrome occurred in three recipients (9%) randomized to IFLT and in 21 (64%) randomized to CLT (difference -54%; 95% CI -74% to -35%; p <0.001). Non-anastomotic biliary strictures diagnosed with protocol magnetic resonance cholangiopancreatography at 12 months were observed in two recipients (8%) randomized to IFLT and in nine (36%) randomized to CLT (difference, -28%; 95% CI -50% to -7%; p = 0.014). The comprehensive complication index at 1 year after transplantation was 30.48 (95% CI 23.25-37.71) in the IFLT group vs. 42.14 (95% CI 35.01-49.26) in the CLT group (difference -11.66; 95% CI -21.81 to -1.51; p = 0.025). CONCLUSIONS: Among patients with end-stage liver disease, IFLT significantly reduced complications related to IRI compared to a conventional approach. CLINICAL TRIAL REGISTRATION: chictr.org. ChiCTR1900021158. IMPACT AND IMPLICATIONS: Ischemia-reperfusion injury has thus far been considered as an inevitable event in organ transplantation, compromising outcomes and limiting organ availability. Ischemia-free liver transplantation is a novel approach of transplanting donor livers without interruption of blood supply. We showed that in patients with end-stage liver disease, ischemia-free liver transplantation, compared with a conventional approach, led to reduced complications related to ischemia-reperfusion injury in this randomized trial. This new approach is expected to change the current practice in organ transplantation, improving transplant outcomes, increasing organ utilization, while providing a clinical model to delineate the impact of organ injury on alloimmunity.

SCANCell reveals diverse inter-cluster interaction patterns in systemic lupus erythematosus across the disease spectrum.

MOTIVATION: High-dimensional mass cytometry (CyTOF), which provides both cellular signatures and inter-cluster interactions like the antagonism between immune activation and suppression, and the pro-inflammatory synergy, sheds light on the cellular and molecular basis of disease pathogenesis. However, revealing the aberrance of inter-cluster communication networks in CyTOF datasets remains a significant challenge. RESULTS: Here, we developed Sample Classification and direct Association Network among Cell clusters (SCANCell) that quantifies the direct association (DA) network of cell clusters. SCANCell was applied to profile inter-cluster interaction patterns of a well-recruited systemic lupus erythematosus (SLE) cohort, including 8 healthy controls, 10 active SLE patients (APs) and 8 remission SLE patients (RPs). SCANCell identified decreased inter-cluster interactions of CD8+ T cells in APs compared with RPs, and enhanced DA of CD8+ T cells after stimulation with immunostimulatory cytokine interleukin-2 in vitro. These discoveries prove that SCANCell can uncover pathology- and drug stimulation-associated inter-cluster interactions, which potentially benefits understanding of pathogenesis and novel therapeutic strategies. AVAILABILITY AND IMPLEMENTATION: The main processing scripts of SCNACell are available at https://github.com/Lxc417/SCANCell. Other codes for the following data statistics are available from the corresponding author upon request. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

AOSD endotypes based on immune cell profiles: patient stratification with hierarchical clustering analysis.

OBJECTIVES: This study aimed to analyse the immune cell profiles of adult-onset Still's disease (AOSD) and to stratify disease-associated endotypes. METHODS: We included 95 cases of treatment-naïve patients with AOSD and 66 cases of healthy controls. Patients with AOSD were classified via an unbiased hierarchical cluster analysis based on circulating immune cells. Their clinical and laboratory characteristics, treatment management, systemic scores and outcomes were then analysed. RESULTS: The proportions of neutrophils and CD8+ T cells were significantly higher while monocytes and natural killer and CD4+ T cells were decreased in patients with AOSD (all P < 0.005). Unbiased hierarchical cluster analysis classified 95 AOSD into three endotype-based groups: group 1 had the highest percentage of neutrophils (neu-dominant group), group 2 had the highest percentage of monocytes (mono-dominant group) and group 3 had the highest percentage of CD8+ T cells (CD8-dominant group). Patients in group 3 had the highest systemic score at diagnosis and were more likely to have pulmonary infiltrates, pericarditis, splenomegaly and poorer treatment responses. A correlation study revealed that the CD4 to CD8 ratio was negatively correlated with the systemic score and positively correlated with treatment response in patients with AOSD (P = 0.001 and P = 0.0091). During the 24.6 (15.2) months of follow-up, the highest total number of disease flares occurred in group 3 (P < 0.0001). CONCLUSION: Hierarchical cluster analysis of peripheral immune cells classified AOSD into three disease-related endotypes. Group 3 showed higher systemic score and poorer treatment responses.

Batoclimab as an add-on therapy in neuromyelitis optica spectrum disorder patients with acute attacks.

BACKGROUND AND PURPOSE: Neuromyelitis optica spectrum disorder (NMOSD) is a severe neurological inflammatory disease mainly caused by pathogenic aquaporin-4 antibodies (AQP4-IgG). The safety and efficacy of the neonatal Fc receptor antagonist batoclimab addition to conventional intravenous methylprednisolone pulse (IVMP) therapy in patients with NMOSD acute attacks was assessed. METHODS: In an open-label, dose-escalation phase 1b study, NMOSD patients with acute myelitis and/or optic neuritis received four doses of weekly subcutaneous injections of either 340 mg or 680 mg batoclimab with concurrent IVMP and were followed up for 27 weeks. The primary end-points were safety and tolerability. Secondary end-points included pharmacodynamics and efficacy, with key efficacy assessment at week 4. RESULTS: In total nine NMOSD patients were enrolled, including two and seven in the 340 and 680 mg groups. Five patients had acute myelitis, while the remaining four had unilateral optic neuritis. Batoclimab add-on therapy had an overall good safety profile without serious adverse events. In the 680 mg group, mean immunoglobulin G (IgG) reached its maximum reduction at the last dose (day 22). In the meantime, AQP4-IgG was undetectable in six of seven subjects whose baseline AQP4-IgG titers ranged from 1:32 to 1:320. Expanded Disability Status Scale score was reduced by 1.3 ± 0.4 at week 4 (2.7 ± 1.3) compared with baseline (4.0 ± 1.0). CONCLUSIONS: Batoclimab add-on therapy to IVMP is safe and tolerated in patients with NMOSD. Preliminary evidence suggests a beneficial neurological effect. A randomized controlled trial would be needed to prove the efficacy.

Cloning and functional analysis of the root-knot nematode resistance gene NtRk1 in tobacco.

Tobacco (Nicotiana tabacum L.) is an economically important crop worldwide. Root-knot nematodes (RKNs) are responsible for yield losses in tobacco and other crops, such as tomato, potato, peanut, and soybean. Therefore, screening for resistance genes that can prevent RKN infestation and the associated damage is crucial. However, there is no report of cloning tobacco RKN resistance genes to date. Here, we cloned the tobacco RKN resistance gene NtRk1 from the resistant variety TI706, using rapid amplification of cDNA ends. NtRk1 has high homology with other RKN resistance genes (CaMi in pepper, Mi-1.1 and Mi-1.2 in tomato). Under normal conditions, NtRk1 was barely expressed in the roots; however, following RKN infection, its expression level rapidly increased. Overexpression of NtRk1 in the susceptible cultivar "Changbohuang" enhanced its resistance to Meloidogyne incognita, while RNA interference of NtRk1 in the resistant cultivar K326 resulted in its susceptibility to M. incognita. Moreover, compared with resistant variety K326, we found the salicylic acid and jasmonic acid contents of RNAi plants decreased after inoculation with M. incognita, and confirmed that the function of NtRk1 is related to these phytohormones. These findings indicate that NtRk1 is an RKN resistance gene, which is abundantly expressed in response to RKN infection and may enhance host defense responses by elevating salicylic acid and jasmonic acid levels.

A prospective cohort study on decreased serum apelin-13 levels after human aneurysmal subarachnoid hemorrhage: associations with severity and prognosis.

Apelin-13 may have neuroprotective effects. We aimed to determine whether serum apelin-13 could serve as a potential biomarker for severity, delayed cerebral ischemia (DCI), and prognosis after human aneurysmal subarachnoid hemorrhage (aSAH). In this prospective, observational, cohort, single-center study of 139 patients with aSAH and 139 healthy individuals, serum apelin-13 levels were determined. The indicators of stroke severity were the Hunt-Hess scale and the modified Fisher grading scale. The prognostic parameters were DCI and 6-month worse prognosis (Extended Glasgow Outcome Scale scores of 1-4). Using binary logistic regression analysis, the relationship between serum apelin-13 levels and prognosis was reported as odds ratios (ORs) with 95% confidence intervals (CIs). Under the receiver operating characteristic curve, prognostic abilities were shown as areas under the curve (AUCs) with 95% CIs. Serum apelin-13 levels were substantially lower in patients than in controls (median, 28.8 versus 48.6 ng/ml; P < 0.001), in patients with DCI than in non-DCI patients (median, 14.9 versus 31.6 ng/ml; P < 0.001), and in patients with worse prognosis than in those with good prognosis (median, 16.3 versus 33.7 ng/ml; P < 0.001). Serum apelin-13 levels were independently correlated with Hunt-Hess scores (beta, -6.836; 95% CI, -8.963-4.708; VIF, 2.219; P = 0.001) and modified Fisher scores (beta, -3.350; 95% CI, -6.151-0.549; VIF, 1.562; P = 0.019). Serum apelin-13 levels were an independent predictor of DCI (OR, 0.951; 95% CI, 0.914-0.990; P = 0.022) and worse prognosis (OR, 0.954; 95% CI, 0.916-0.993; P = 0.013). Serum apelin-13 levels significantly differentiated DCI and poor prognosis, with AUCs of 0.753 (95% CI, 0.656-0.850) and 0.791 (95% CI, 0.713-0.868) respectively. Using the Youden method, serum apelin-13 levels < 19.3 ng/ml distinguished the risk of DCI with 64.7% sensitivity and 77.1% specificity, and serum apelin-13 levels < 30.2 ng/ml discriminated the development of worse prognosis with 89.1% sensitivity and 63.4% specificity. Serum apelin-13 levels combined with Hunt-Hess scores and modified Fisher scores displayed a significantly higher AUC than any one of them for prognostic prediction (all P < 0.05). Decreased serum apelin-13 levels, which are strongly correlated with disease severity, independently predicted poor outcomes following aSAH, substantializing serum apelin-13 as a useful prognostic biomarker of aSAH.

Distributional learning of musical pitch despite tone deafness in individuals with congenital amusia.

Congenital amusia is an innate and lifelong deficit of music processing. This study investigated whether adult listeners with amusia were still able to learn pitch-related musical chords based on stimulus frequency of statistical distribution, i.e., via distributional learning. Following a pretest-training-posttest design, 18 amusics and 19 typical, musically intact listeners were assigned to bimodal and unimodal conditions that differed in distribution of the stimuli. Participants' task was to discriminate chord minimal pairs, which were transposed to a novel microtonal scale. Accuracy rates for each test session were collected and compared between the two groups using generalized mixed-effects models. Results showed that amusics were less accurate than typical listeners at all comparisons, thus corroborating previous findings. Importantly, amusics-like typical listeners-demonstrated perceptual gains from pretest to posttest in the bimodal condition (but not the unimodal condition). The findings reveal that amusics' distributional learning of music remains largely preserved despite their deficient music processing. Implications of the results for statistical learning and intervention programs to mitigate amusia are discussed.

Serosal involvement in adult-onset Still's disease: A multicentre and retrospective study.

OBJECTIVES: This study evaluated the characteristics of serosal involvement in adult-onset Still's disease (AOSD). METHODS: Patients meeting the Yamaguchi classification criteria were classified into AOSD with and without serosal involvement according to their manifestations and sonography/radiography. Clinical data was retrospectively reviewed from 102 patients with AOSD in two centres. RESULTS: Forty-two patients (41.2%) had serosal involvement. The frequencies of pulmonary infiltrate and impaired liver function were significantly higher in patients with serosal involvement (p = .002 and p = .007, respectively), who also had a higher modified systemic score (p = .009). In addition, the percentages of CD3+ T cells (p < .001) and, especially, the CD8+ T cells (p = .004) were significantly increased in the peripheral blood of AOSD patients with serosal involvement. Notably, patients with serosal involvement were more likely to develop macrophage activation syndrome (p = .047) and a chronic pattern (p = .016) during the follow-up. CONCLUSIONS: Patients with serosal involvement demonstrated the more severe disease activity and different immune phenotypes; these patients were more likely to develop macrophage activation syndrome, and they may require more aggressive treatment at an early time to control their systemic inflammation.

Genomic Scar Score: A robust model predicting homologous recombination deficiency based on genomic instability.

OBJECTIVE: To develop a novel machine learning-based algorithm called the Genomic Scar Score (GSS) for predicting homologous recombination deficiency (HRD) events. DESIGN: Method development study. SETTING: AmoyDx Medical Laboratory and Jiangsu Cancer Hospital. POPULATION OR SAMPLE: A cohort of individuals with ovarian or breast cancer (n = 377) were collected from the AmoyDx Medical Laboratory. Another cohort of patients with ovarian cancer treated with PARP inhibitors (n = 58) was enrolled in the Jiangsu Cancer Hospital. METHODS: We used linear support vector machines to build a Genomic Scar (GS) model to predict HRD events, and Kaplan-Meier analyses were performed by comparing the progression-free survival (PFS) of patients in different groups using a two-sided log-rank test. MAIN OUTCOME MEASURES: The performance of the GS model and the result of clinical validation. RESULTS: The GS model displayed more than 97.0% sensitivity to detect BRCA-deficient events, and the GS model identified patients that could benefit from poly(ADP-ribose) polymerase inhibitors (PARPi), as the GS score (GSS)-positive group had a longer progression-free survival (PFS) (9.4 versus 4.4 months; hazard ratio [HR] = 0.54, P < 0.001) than the GSS-negative group after PARPi treatment. Meanwhile, the GSS showed high concordance among different NGS panels, which implied the robustness of the GS model. CONCLUSIONS: The GS was a robust model to predict HRD and had broad clinical applications in predicting which patients will respond favourably to PARPi treatment.

TNF-α inhibitor tanfanercept (HBM9036) improves signs and symptoms of dry eye in a phase 2 trial in the controlled adverse environment in China.

PURPOSE: This study evaluated the clinical safety and efficacy of tanfanercept (HBM9036) ophthalmic solution as a novel treatment for dry eye disease (DED) in a controlled adverse environment (CAE) study conducted in China. METHODS: In a single-center, double-masked, randomized, placebo-controlled study, 100 patients received 0.25% tanfanercept, or placebo, twice daily for eight weeks. A mobile international CAE® DE Model was used for patient selection with a standardized challenge endpoint. Primary efficacy endpoint was fluorescein inferior corneal staining score (ICSS) pre- to post-CAE challenge from baseline. Secondary endpoints included Schirmer's Tear Test, Tear-Film Break-Up Time, Ocular Discomfort Score, Ora Calibra® Ocular Discomfort and 4-Symptom Questionnaire, total corneal staining score (TCSS), and drop comfort. Signs and symptoms were assessed both pre- and post-CAE to evaluate the efficacy of tanfanercept on both environmental and CAE endpoints. RESULTS: The tanfanercept treatment group showed improvement in ICSS pre- to post-CAE change from baseline scores when compared to placebo (- 0.61 ± 0.11 and - 0.54 ± 0.11, respectively; mean difference = 0.07, p = 0.65). TCSS pre-post-CAE change from baseline scores was also in favor of active when compared to placebo (- 1.03 ± 0.21 and - 0.67 ± 0.21, respectively; mean difference = 0.37, p = 0.23). Schirmer's score improvement was demonstrated in favor of active (1.87 ± 0.62 mm) as compared to placebo (1.28 ± 0.62 mm; mean difference = 0.59 mm, p = 0.50). Change from baseline in mean Tear-Film Break-up Time favored active treatment over placebo (mean difference = 1.21 s, p = 0.45). Notably, the tanfanercept showed more obvious benefits for each DED sign in a subgroup of subjects ≥ 35 years of age. Tanfanercept was well tolerated with no serious adverse events occurring during the study. CONCLUSION: Tanfanercept demonstrated improvements in favor of active as compared to placebo in the signs of DED, being safe and well tolerated. These data support further evaluation of tanfanercept for the treatment of DED in China. TRIAL REGISTRATION: This study was retrospectively registered at ClinicalTrials.gov (NCT04092907) on September 17, 2019.

RIPK1 downregulation enhances neutrophil extracellular traps in psoriasis.

Introduction: Psoriasis is an immune-mediated systemic disease. Neutrophils are enriched in psoriasis lesions and can form neutrophil extracellular traps (NETs) to activate keratinocytes. Receptor-interacting protein kinase RIPK1 and RIPK3 are involved in necroptosis and NET formation. Aim: To elucidate whether RIPK1 regulates circulating neutrophils to form NETs and inflammation in psoriasis. Material and methods: Blood samples of psoriasis patients (n = 20) and healthy controls (n = 20) were detected by flow cytometry. The expression level of RIPK1/3 in isolated circulating neutrophils from psoriasis patients (n = 17) and healthy controls (n = 17) was examined by quantitative real-time PCR. SYTOX Green dye and PicoGreen reagent were used to detect NET formation and DNA release in neutrophils under the stimulation of phorbol 12-myristate 13-acetate (PMA) and necrostain-1 (Nec-1). Correlation analysis was performed between RIPK1/3 expression and Psoriasis Area Severity Index (PASI), neutrophil-to-lymphocyte ratio (NLR). Results: RIPK1 and RIPK3 expression in protein levels were decreased in monocytes and neutrophils from peripheral blood of psoriasis patients. In isolated psoriasis neutrophils, RIPK1 and Caspase8 mRNA were downregulated while RIPK3 and MLKL mRNA were elevated, leading to the necroptosis pathway. In addition, RIPK1-inhitor-necrostatin-1 (Nec-1) enhanced NETosis in psoriasis neutrophils in vitro. More importantly, there is a negative correlation between RIPK1 and psoriasis disease severity. Conclusions: Our data demonstrated that downregulated RIPK1 expression in psoriasis neutrophils may enhance NET generation. RIPK1 may be identified as a novel therapeutic target in psoriasis.

Efficacy and safety of a selective URAT1 inhibitor SHR4640 in Chinese subjects with hyperuricaemia: a randomized controlled phase II study.

OBJECTIVE: To evaluate the efficacy and safety of SHR4640, a highly selective urate transporter 1 inhibitor, in Chinese subjects with hyperuricaemia. METHODS: This was a randomized double-blind dose-ranging phase II study. Subjects whose serum uric acid (sUA) levels were ≥480 µmol/l with gout, ≥480 µmol/l without gout but with comorbidities, or ≥540 µmol/l were enrolled. Subjects were randomly assigned (1:1:1:1:1) to receive once daily 2.5 mg, 5 mg, 10 mg of SHR4640, 50 mg of benzbromarone or placebo, respectively. The primary end point was the proportion of subjects who achieved target sUA level of ≤360 µmol/l at week 5. RESULTS: 99.5% of subjects (n = 197) were male and 95.9% of subjects had gout history. The proportions of subjects who achieved target sUA at week 5 were 32.5%, 72.5% and 61.5% in the 5 mg, 10 mg SHR4640 and benzbromarone groups, respectively, significantly higher than the placebo group (0%; P < 0.05 for 5 mg and 10 mg SHR4640 group). The sUA was reduced by 32.7%, 46.8% and 41.8% at week 5 with 5 mg, 10 mg SHR4640 and benzbromarone, respectively, vs placebo (5.9%; P < 0.001 for each comparison). The incidences of gout flares requiring intervention were similar among all groups. Occurrences of treatment-emergent adverse events (TEAEs) were comparable across all groups, and serious TEAEs were not reported. CONCLUSIONS: The present study indicated a superior sUA-lowering effect and well tolerated safety profile after 5-week treatment with once-daily 5 mg/10 mg of SHR4640 as compared with placebo in Chinese subjects with hyperuricaemia. TRIAL REGISTRATION: ClinicalTrials.gov number, NCT03185793.

The effects of native prosodic system and segmental context on Cantonese tone perception by Mandarin and Japanese listeners.

This study explores the effects of native prosodic system and segmental context on the perception of Cantonese tones by Mandarin and Japanese listeners. In Experiment 1, 13 Mandarin and 13 Japanese subjects took part in a two-alternative forced-choice discrimination test of Cantonese tones in different segmental contexts (familiar vs unfamiliar). In Experiment 2, 20 Mandarin listeners participated in a perceptual assimilation task that examined the cross-language perceptual similarity between Mandarin and Cantonese tones. Results showed that Mandarin listeners were comparable to Japanese counterparts in discriminability, but the former attended more to pitch contour differences while the latter were more sensitive to pitch height. Moreover, the effect of segmental context was significant exclusively in the Mandarin group, whereas the Japanese group performed stably across syllables in discriminating Cantonese tones. It seemed that unfamiliar context rendered lower perceptual similarity, which further hindered corresponding discrimination by the Mandarin group. In addition, segmental effects were mainly observed in the assimilation patterns of category goodness or uncategorized-categorized. These findings suggested that non-native tone perception could be modulated by listeners' native prosodic structures in a finer way.

Determination of Flavonoids Compounds of Three Species and Different Harvesting Periods in Crataegi folium Based on LC-MS/MS.

Crataegi folium have been used as medicinal and food materials worldwide due to its pharmacological activities. Although the leaves of Crataegus songorica (CS), Crataegus altaica (CA) and Crataegus kansuensis (CK) have rich resources in Xinjiang, China, they can not provide insights into edible and medicinal aspects. Few reports are available on the qualitative and quantitative analysis of flavonoids compounds of their leaves. Therefore, it is necessary to develop efficient methods to determine qualitative and quantitative flavonoids compounds in leaves of CS, CA and CK. In the study, 28 unique compounds were identified in CS versus CK by qualitative analysis. The validated quantitative method was employed to determine the content of eight flavonoids of the leaves of CS, CA and CK within 6 min. The total content of eight flavonoids was 7.8-15.1 mg/g, 0.1-9.1 mg/g and 4.8-10.7 mg/g in the leaves of CS, CA and CK respectively. Besides, the best harvesting periods of the three species were from 17th to 26th September for CS, from 30th September to 15th October for CA and CK. The validated and time-saving method was successfully implemented for the analysis of the content of eight flavonoids compounds in CS, CA and CK for the first time.