Super-resolution imaging reveals the mechanism of endosomal acidification inhibitors against SARS-CoV-2 infection.

In this study, super-resolution structured illumination microscope (SIM) was used to analyze molecular mechanism of endocytic acidification inhibitors in the SARS-CoV-2 pandemic, such as Chloroquine (CQ), Hydroxychloroquine (HCQ) and Bafilomycin A1 (BafA1). We fluorescently labeled the SARS-CoV-2 RBD and its receptor ACE2 protein with small molecule dyes. Utilizing SIM imaging, the real-time impact of inhibitors (BafA1, CQ, HCQ, Dynasore) on the RBD-ACE2 endocytotic process was dynamically tracked in living cells. Initially, the protein activity of RBD and ACE2 was ensured after being labeled. And then our findings revealed that these inhibitors could inhibit the internalization and degradation of RBD-ACE2 to varying degrees. Among them, 100 nM BafA1 exhibited the most satisfactory endocytotic inhibition (~63.9%) and protein degradation inhibition (~97.7%). And it could inhibit the fusion between endocytic vesicles in the living cells. Additionally, Dynasore, a widely recognized dynein inhibitor, also demonstrated cell acidification inhibition effects. Together, these inhibitors collectively hinder SARS-CoV-2 infection by inhibiting both the viral internalization and RNA release. The comprehensive evaluation of pharmacological mechanisms through super-resolution fluorescence imaging has laid a crucial theoretical foundation for the development of potential drugs to treat COVID-19.

Safety and efficacy of zandelisib plus zanubrutinib in previously treated follicular and mantle cell lymphomas.

The combination of the phosphatidylinositol 3-kinase delta (PI3Kδ) inhibitor zandelisib with the Bruton's tyrosine kinase (BTK) inhibitor zanubrutinib was hypothesized to be synergistic and prevent resistance to single-agent therapy. This phase 1 study (NCT02914938) included a dose-finding stage in patients with relapsed/refractory (R/R) B-cell malignancies (n = 20) and disease-specific expansion cohorts in follicular lymphoma (FL; n = 31) or mantle cell lymphoma (MCL; n = 19). The recommended phase 2 dose was zandelisib 60 mg on Days 1-7 plus zanubrutinib 80 mg twice daily continuously in 28-day cycle. In the total population, the most common adverse events (AEs; all grades/grade 3-4) were neutropenia (35%/24%), diarrhoea (33%/2%), thrombocytopenia (32%/8%), anaemia (27%/8%), increased creatinine (25%/0%), contusion (21%/0%), fatigue (21%/2%), nausea (21%/2%) and increased aspartate aminotransferase (24%/6%). Three patients discontinued due to AEs. The overall response rate was 87% (complete response [CR] = 33%) for FL and 74% (CR = 47%) for MCL. The median duration of response and progression-free survival (PFS) were not reached in either group. The estimated 1-year PFS was 72.3% (95% confidence interval [CI], 51.9-85.1) for FL and 56.3% (95% CI, 28.9-76.7) for MCL (median follow-up: 16.5 and 10.9 months respectively). Zandelisib plus zanubrutinib was associated with high response rates and no increased toxicity compared to either agent alone.

TPD52 as a Potential Prognostic Biomarker and its Correlation with Immune Infiltrates in Uterine Corpus Endometrial Carcinoma: Bioinformatic Analysis and Experimental Verification.

BACKGROUND: Aberrant expression of tumor protein D52 (TPD52) is associated with some tumors. The role of TPD52 in uterine corpus endometrial carcinoma (UCEC) remains uncertain. OBJECTIVE: We aimed to investigate the involvement of TPD52 in the pathogenesis of UCEC. METHODS: We employed bioinformatics analysis and experimental validation in our study. RESULTS: Our findings indicated that elevated TPD52 expression in UCEC was significantly associated with various clinical factors, including clinical stage, race, weight, body mass index (BMI), histological type, histological grade, surgical approach, and age (p < 0.01). Furthermore, high TPD52 expression was a predictor of poorer overall survival (OS), progress-free survival (PFS), and disease-specific survival (DSS) (p = 0.011, p = 0.006, and p = 0.003, respectively). TPD52 exhibited a significant correlation with DSS (HR: 2.500; 95% CI: 1.153-5.419; p = 0.02). TPD52 was involved in GPCR ligand binding and formation of the cornified envelope in UCEC. Moreover, TPD52 expression was found to be associated with immune infiltration, immune checkpoints, tumor mutation burden (TMB)/ microsatellite instability (MSI), and mRNA stemness indices (mRNAsi). The somatic mutation rate of TPD52 in UCEC was 1.9%. A ceRNA network of AC011447.7/miR-1-3p/TPD52 was constructed. There was excessive TPD52 protein expression. The upregulation of TPD52 expression in UCEC cell lines was found to be statistically significant. CONCLUSION: TPD52 is upregulated in UCEC and may be a useful patent for prognostic biomarkers of UCEC, which may have important value for clinical treatment and supervision of UCEC patients.

Objective Neurological Testing Methods Used to Follow Up Vestibular Neuritis Depending on Different Factors.

Purpose: We analyze the impact of different factors on clinical performance and prognosis in vestibular neuritis (VN) and explore indicators that could accurately reflect changes in patients' symptoms at different stages. Methods: We observed patients with VN during the acute and recovery phases. Clinical symptoms, vertigo-related scales, neurological examination, vestibular function tests (caloric test, video head impulse test (vHIT), vestibular evoked myogenic potential (VEMP)), and the history of disease (underlying diseases, glucocorticoid therapy) were recorded at onset and at 4 and 12 weeks after onset in VN patients. Multiple linear regression analysis was used to identify vestibular function tests that had a linear regression relationship with the subjective quantitative results. Results: At 4 weeks after onset, the group without underlying disease had better improvement in EEV, gain, and UW than the group with underlying disease (P < 0.05). There was a significant difference in the change in DHI, EEV, gain of the affected horizontal semicircular canal in the vHIT and unilateral weakness (UW) between the glucocorticoid treatment group and the no glucocorticoid treatment group (P < 0.05), and glucocorticoid treatment group was better. The change value in the gain of horizontal canals in the vHIT was mainly positively and linearly correlated with the EEV scores (P<0.001). Possible dynamic correlation between vHIT results and vestibular symptoms. Conclusion: The absence of underlying disease and the receipt of glucocorticoid therapy significantly contributed to the improvement of objective vestibular function tests in the short term, while the improvement of subjective vertigo may correlate with the different objective measures and questionnaire. We believe that the improvement of the affected horizontal semicircular canal gain value in the vHIT can be used as a reference indicator of the degree of improvement of vestibular symptoms with superior vestibular neuritis.

Simulated microgravity-induced oxidative stress and loss of osteogenic potential of osteoblasts can be prevented by protection of primary cilia.

Oxidative stress has been considered to be closely related to spaceflight-induced bone loss; however, mechanism is elusive and there are no effective countermeasures. Using cultured rat calvarial osteoblasts exposed to microgravity simulated by a random positioning machine, this study addressed the hypotheses that microgravity-induced shortening of primary cilia leads to oxidative stress and that primary cilium protection prevents oxidative stress and osteogenesis loss. Microgravity was found to induce oxidative stress (as represented by increased levels of reactive oxygen species (ROS) and malondialdehyde production, and decreased activities of antioxidant enzymes), which was perfectly replicated in osteoblasts growing in NG with abrogated primary cilia (created by transfection of an interfering RNA), suggesting the possibility that shortening of primary cilia leads to oxidative stress. Oxidative stress was accompanied by mitochondrial dysfunction (represented by increased mitochondrial ROS and decreased mitochondrial membrane potential) and intracellular Ca2+ overload, and the latter was found to be caused by increased activity of Ca2+ channel transient receptor potential vanilloid 4 (TRPV4), as also evidenced by TRPV4 agonist GSK1016790A-elicited Ca2+ influx. Supplementation of HC-067047, a specific antagonist of TRPV4, attenuated microgravity-induced mitochondrial dysfunction, oxidative stress, and osteogenesis loss. Although TRPV4 was found localized in primary cilia and expressed at low levels in NG, microgravity-induced shortening of primary cilia led to increased TRPV4 levels and Ca2+ influx. When primary cilia were protected by miR-129-3p overexpression or supplementation with a natural flavonoid moslosooflavone, microgravity-induced increased TRPV4 expression, mitochondrial dysfunction, oxidative stress, and osteogenesis loss were all prevented. Our data revealed a new mechanism that primary cilia function as a controller for TRPV4 expression. Microgravity-induced injury on primary cilia leads to increased expression and overactive channel of TRPV4, causing intracellular Ca2+ overload and oxidative stress, and primary cilium protection could be an effective countermeasure against microgravity-induced oxidative stress and loss of osteogenic potential of osteoblasts.

NOTCH Pathway Genes in Ovarian Cancer: Clinical Significance and Associations with Immune Cell Infiltration.

BACKGROUND: Activation of the NOTCH signaling pathway is associated with tumorigenesis. The aim of this study was to investigate NOTCH pathway gene functions and regulatory mechanisms in ovarian cancer (OC). METHODS: We conducted a bioinformatics analysis of publicly available datasets in order to identify potential NOTCH-related mechanisms, associated genes, biological pathways, and their relation to immune function. RESULTS: Significant differential expression of the NOTCH pathway genes DLL1, DLL3, DLL4, HES1, HEY1, JAG1, NOTCH2, NOTCH3, and NOTCH4 was observed between OC samples and normal controls. Low expression of DLL4 and of NOTCH4 in OC patients was associated with International Federation of Gynecology and Obstetrics (FIGO) stage (p <0.001 and p = 0.036, respectively), while high expression of NOTCH3 was associated with race (p = 0.039) and age (p = 0.044). JAG2 and NOTCH1 expression were significantly associated with progression-free interval (PFI) (p = 0.011 and p = 0.039, respectively). DLL1 (Hazard Ratio (HR): 2.096; 95% CI: 1.522-2.886, p < 0.001) and NOTCH1 (HR: 0.711; 95% CI: 0.514-0.983, p = 0.039) expression were independently associated with PFI in multivariate analysis. DLL1, DLL3, JAG1, JAG2, NOTCH3 and NOTCH4 expression could significantly differentiate OC from non-cancer samples. Genes associated with the NOTCH pathway were mainly enriched in five signaling pathways: the NOTCH signaling pathway, breast cancer, endocrine resistance, Th1 and Th2 cell differentiation, and oxidative phosphorylation. The expression of NOTCH pathway genes was significantly associated with immune cell infiltration. CONCLUSIONS: NOTCH pathway genes appear to play an important role in the progression of OC by regulating immune cells, endocrine resistance, Th1 and Th2 cell differentiation, and oxidative phosphorylation. JAG2 and NOTCH1 are potential biomarkers and therapeutic targets for the treatment of OC.

SIM imaging resolves endocytosis of SARS-CoV-2 spike RBD in living cells.

It is urgent to understand the infection mechanism of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) for the prevention and treatment of COVID-19. The infection of SARS-CoV-2 starts when the receptor-binding domain (RBD) of viral spike protein binds to angiotensin-converting enzyme 2 (ACE2) of the host cell, but the endocytosis details after this binding are not clear. Here, RBD and ACE2 were genetically coded and labeled with organic dyes to track RBD endocytosis in living cells. The photostable dyes enable long-term structured illumination microscopy (SIM) imaging and to quantify RBD-ACE2 binding (RAB) by the intensity ratio of RBD/ACE2 fluorescence. We resolved RAB endocytosis in living cells, including RBD-ACE2 recognition, cofactor-regulated membrane internalization, RAB-bearing vesicle formation and transport, RAB degradation, and downregulation of ACE2. The RAB was found to activate the RBD internalization. After vesicles were transported and matured within cells, RAB was finally degraded after being taken up by lysosomes. This strategy is a promising tool to understand the infection mechanism of SARS-CoV-2.

Simulation analysis of carbon peak path in China from a multi-scenario perspective: evidence from random forest and back propagation neural network models.

China faces tough challenges in the process of low-carbon transformation. To determine whether China can achieve its new 2030 carbon peaking and carbon intensity reduction commitments, accurate prediction of China's CO2 emissions is vital. In this paper, the random forest (RF) model was used to screen 26 carbon emission influencing factors, and seven indicators were selected as key variables for prediction. Subsequently, a three-layer back propagation (BP) neural network was constructed to forecast China's CO2 emissions and intensity from 2020 to 2040 under the 13th Five-Year Plan, 14th Five-Year Plan, energy optimization, technology breakthrough, and dual control scenarios. The results showed that energy structure factors have the most significant impact on China's CO2 emissions, followed by technology level, and economic development factors are no longer the main drivers. Under the 14th Five-Year Plan scenario, China can achieve its carbon peaking on time, reaching 10,434.082 Mt CO2 emissions in 2030. Although the new commitment to intensity reduction (over 65%) under this scenario cannot be achieved, the 14th Five-Year Plan can bring about 73.359 and 539.710 Mt of CO2 reduction in 2030 and 2040 respectively, compared to the 13th Five-Year Plan. Under the technology breakthrough and dual control scenarios, China will meet its new commitments ahead of schedule, with the dual control scenario being the optimal pathway for CO2 emissions to peak at 9860.08 Mt in 2025. It is necessary for Chinese policy makers to adjust their current strategic planning, such as accelerating the transformation of energy structure and increasing investment in R&D to achieve breakthroughs in green technologies.

TPD52 is a Potential Prognostic Biomarker and Correlated with Immune Infiltration: A Pan-cancer Analysis.

BACKGROUND: Some tumors have a poor prognosis regarding TPD52 (tumor protein D52). This study aims to explore TPD52's role in the cancer process from a pan-cancer perspective. METHODS: A pan-cancer analysis was conducted to investigate how TPD52 may be involved in cancer as well as its association with prognosis. RESULTS: A variety of human cancers express TPD52 abnormally and correlate with clinical stage. There was a significant association between low expression of TPD52 and poor survival in BRCA, KIRP, LAML, LIHC, UCEC, and UVM. TPD52 alterations were most frequently amplified in pan-cancer. The co-occurrence of 10 genes alterations was found in the TPD52 altered group. There was a significant association between TPD52 expression and MSI in four cancer types and TMB in twelve cancer types. There was a significant correlation between TPD52 expression and immunerelated cell infiltration. A significant correlation was found between TPD52 expression in many tumor types and 8 immune checkpoint genes. There were signaling pathways involved in pan-cancer caused by TPD52, including endocytosis, Fc gamma Rmediated phagocytosis, and so on. TPD52 may be involved in chemotherapy and chemoresistance. CONCLUSION: The TPD52 gene may be important for human cancer treatment.

Diagnosis Biomarkers of Cholangiocarcinoma in Human Bile: An Evidence-Based Study.

Cholangiocarcinoma (CCA) is a multifactorial malignant tumor of the biliary tract, and the incidence of CCA is increasing in recent years. At present, the diagnosis of CCA mainly depends on imaging and invasive examination, with limited specificity and sensitivity and late detection. The early diagnosis of CCA always faces the dilemma of lacking specific diagnostic biomarkers. Non-invasive methods to assess the degree of CAA have been developed throughout the last decades. Among the many specimens looking for CCA biomarkers, bile has gotten a lot of attention lately. This paper mainly summarizes the recent developments in the current research on the diagnostic biomarkers for CCA in human bile at the levels of the gene, protein, metabolite, extracellular vesicles and volatile organic compounds.

Comprehensive surgical treatment for obstructive rectal endometriosis: a case report and review of the literature.

BACKGROUND: Intestinal obstruction caused by endometriosis maybe easily misdiagnosed as a tumor or other occupying disease in emergency condition. How to deal with it depending on the clarity of the preoperative diagnosis and the experience of the surgeon. CASE PRESENTATION: A 47-year-old woman, admitted to our emergency service with abdominal pain and distension for 5 days, anal stop exhausting and defecating for 3 days. Based on imaging and laboratory examination, we made a preoperative diagnosis of rectal endometriosis probably. After 7 days of colon decompression with a intestinal obstruction catheter, an operation of laparoscopic partial rectal and sigmoid resection without protective stoma and total hysterectomy was performed successfully. The patient obtained a smooth postoperative course and doing well after 12-weeks follow up. CONCLUSIONS: Obstruction caused by rectal endometriosis is very rare and easily overlooked by surgeon and gynecologist. Appropriate preoperative diagnosis and preoperative management can reduce the trauma and incidence of complications.

[A novel phenylisoquinoline alkaloid with cardioprotective effect from Aconiti Lateralis Radix Praeparata].

Macroporous resin chromatography, silica gel column chromatography, preparative thin layer chromatography, and semi-preparative high performance liquid chromatography were performed to isolate two compounds from the acid extract of the lateral roots of Aconitum carmichaelii: a new 9-phenylisoquinoline alkaloid(1) and a known pavine alkaloid(2). Their structures were elucidated by spectroscopy. The absolute configuration of compound 1 was identified by electronic circular dichroism(ECD) and it was determined to be(aS)-7,8-dimethoxy-9-(2-carboxy-4,5-dimethoxyphenyl)-3,4-dihydroisoquinoline-1(2H)-one(1). The cardioprotective effects of 1 and 2 against doxorubicin-induced toxicity in H9 c2 cells were evaluated. Both of the isoquinoline alkaloids showed cardioprotective activity.

In Situ Real-Time Nanoscale Resolution of Structural Evolution and Dynamics of Fluorescent Self-Assemblies by Super-Resolution Imaging.

The visualization of self-assembled structure and dynamics at the molecular level has become a powerful method to understand structure-function relationships of self-assembly. Herein, we in situ real-time imaged the dynamic process of benzyl-naphthalimide dyes at the nanoscale and inspected their internal structure with minimum 2.8 nm localization accuracy through single-molecule localization microscopy (SMLM) imaging. We monitored the growth process of three different assemblies in situ, which possessed highly heterogeneous dynamics with different shapes and growth rates. Furthermore, diverse growth rates were also found at different sites in the same assembly. These results highlight the application of super-resolution microscopy techniques for real-time visualization of internal assembled structure and dynamics in situ.

MoS2-based nanocomposites for cancer diagnosis and therapy.

Molybdenum is a trace dietary element necessary for the survival of humans. Some molybdenum-bearing enzymes are involved in key metabolic activities in the human body (such as xanthine oxidase, aldehyde oxidase and sulfite oxidase). Many molybdenum-based compounds have been widely used in biomedical research. Especially, MoS2-nanomaterials have attracted more attention in cancer diagnosis and treatment recently because of their unique physical and chemical properties. MoS2 can adsorb various biomolecules and drug molecules via covalent or non-covalent interactions because it is easy to modify and possess a high specific surface area, improving its tumor targeting and colloidal stability, as well as accuracy and sensitivity for detecting specific biomarkers. At the same time, in the near-infrared (NIR) window, MoS2 has excellent optical absorption and prominent photothermal conversion efficiency, which can achieve NIR-based phototherapy and NIR-responsive controlled drug-release. Significantly, the modified MoS2-nanocomposite can specifically respond to the tumor microenvironment, leading to drug accumulation in the tumor site increased, reducing its side effects on non-cancerous tissues, and improved therapeutic effect. In this review, we introduced the latest developments of MoS2-nanocomposites in cancer diagnosis and therapy, mainly focusing on biosensors, bioimaging, chemotherapy, phototherapy, microwave hyperthermia, and combination therapy. Furthermore, we also discuss the current challenges and prospects of MoS2-nanocomposites in cancer treatment.

The Feasibility of Antioxidants Avoiding Oxidative Damages from Reactive Oxygen Species in Cryopreservation.

Cryopreservation prolongs the storage time of cells and plays an important role in modern biology, agriculture, plant science and medicine. During cryopreservation, cells may suffer many damages, such as osmotic dehydration, large ice puncture and oxidative damages from reactive oxygen species (ROS). Classic cryoprotectants (CPAs) are failing to dispose of ROS, while antioxidants can turn ROS into harmless materials and regulate oxidative stress. The combination of antioxidants and CPAs can improve the efficiency of cryopreservation while negative results may occur by misuse of antioxidants. This paper discussed the feasibility of antioxidants in cryopreservation.

A substantia innominata-midbrain circuit controls a general aggressive response.

Although aggressive behaviors are universal and essential for survival, "uncontrollable" and abnormal aggressive behaviors in animals or humans may have severe adverse consequences or social costs. Neural circuits regulating specific forms of aggression under defined conditions have been described, but how brain circuits govern a general aggressive response remains unknown. Here, we found that posterior substantia innominata (pSI) neurons responded to several aggression-provoking cues with the graded activity of differential dynamics, predicting the aggressive state and the topography of aggression in mice. Activation of pSI neurons projecting to the periaqueductal gray (PAG) increased aggressive arousal and robustly initiated/promoted all the types of aggressive behavior examined in an activity-level-dependent manner. Inactivation of the pSI circuit largely blocked diverse aggressive behaviors but not mating. By encoding a general aggressive response, the pSI-PAG circuit universally drives multiple aggressive behaviors and may provide a potential target for alleviating human pathological aggression.

An assembly-regulated SNAP-tag fluorogenic probe for long-term super-resolution imaging of mitochondrial dynamics.

Super-resolution fluorescence microscopy has emerged as a powerful tool for studying mitochondrial dynamics in living cells. However, the lack of photostable and chemstable probe makes long-term super-resolution imaging of mitochondria still a challenging work. Herein, we reported a 4-azetidinyl-naphthliamide derived SNAP-tag probe AN-BG exhibiting excellent fluorogenicity and photostability for long-term super-resolution imaging of mitochondrial dynamics. The azetidinyl group and naphthalimide fluorophore are in a flat conformation which can effectively suppress twisted intramolecular charge transfer and then effectively improve the brightness and photostability. This planarized molecular structure is conducive to the formation of fluorescence-quenched J-aggregates, and the protein labeling process will depolymerize the probes and restore fluorescence. Fluorescent labeling mitochondrial inner membrane proteins via SNAP tags overcomes the shortcomings that variations in mitochondrial inner membrane potential will release probes attached to mitochondria by electrostatic interactions. Therefore, AN-BG realized the stable labeling of mitochondria and the long-term imaging of mitochondrial dynamics under super-resolution microscopy.

Cardioprotective effects and concentration-response relationship of aminoalcohol-diterpenoid alkaloids from Aconitum carmichaelii.

Fuzi, a well-known traditional Chinese medicine developed from the lateral roots of Aconitum carmichaelii Debx., has been widely used for the treatment of heart failure. In order to search for active compounds from Fuzi, a phytochemical study was performed, which resulted in the isolation of 14 aminoalcohol-diterpenoid alkaloids, including one new compound (1). Their cardioprotective effects against doxorubicin-induced toxicity in H9c2 cells were evaluated. All of the alkaloids showed cardioprotective effects in a nonmonotonic concentration-response manner, with the maximum protection rates ranging from 17.96 ± 2.93% to 98.31 ± 0.35%. Compound 5 exhibited the most potent cardioprotective activity. Taking the maximum protection rate as an indicator, the preliminary structure-activity relationship analysis indicated that the substitutions of C-1, C-13, C-15, C-16, and N and the configurations of OMe-6 and OH-15 are important structural features for the cardioprotective activities of the aminoalcohol-diterpenoid alkaloids.

A Cell Membrane Fluorogenic Probe for Gram-Positive Bacteria Imaging and Real-Time Tracking of Bacterial Viability.

Bacterial infections are a global healthcare problem, resulting in serious clinical morbidities and mortality. Real-time monitoring of live bacteria by fluorescent imaging technology has potential in diagnosis of bacterial infections, elucidating antimicrobial agents' mode of action, assessing drug toxicity, and examining bacterial antimicrobial resistance. In this work, a naphthalimide-derived fluorescent probe ZTRS-BP was developed for wash-free Gram-positive bacteria imaging. The probe aggregated in aqueous solutions and exhibited aggregation-caused fluorescence quenching (ACQ). The interaction with Gram-positive bacteria cell walls would selectively disaggregate the probe and the liberated probes were dispersed on the outside of the bacteria cell walls to achieve surface fluorescence imaging. There were no such interactions with Gram-negative bacteria, which indicates that selective binding and imaging of Gram-positive bacteria was achieved. The binding of zinc ions by ZTRS-BP can enhance the fluorescent signals on the bacterial surface by inhibiting the process of photoinduced electron transfer. ZTRS-BP-Zn(II) complex was an excellent dye to discriminate mixed Gram-positive and Gram-negative bacteria. Also, live and dead bacteria can be differentially imaged by ZTRS-BP-Zn(II). Furthermore, ZTRS-BP-Zn(II) was used for real-time monitoring bacteria viability such as B. cereus treated with antibiotic vancomycin.