Artesunate-loaded thermosensitive chitosan hydrogel promotes osteogenesis of maxillary tooth extraction through regulating T lymphocytes in type 2 diabetic rats.

BACKGROUND: Type 2 diabetes mellitus (T2DM) causes severe bone loss after tooth extraction as a hyperglycemic environment causes aberrant bone homeostasis. Artesunate (ART) is known to possess anti-inflammation and osteogenic properties. However, its osteogenesis property in alveolar bone remains unclear. This study aimed to explore the osteogenic and immunoregulatory effects of artesunate-loaded thermosensitive chitosan hydrogel (ART-loaded TCH) on maxilla tooth extraction in T2DM rats. METHODS: T2DM rats were induced by a high-fat diet and streptozotocin. Different concentrations of ART-loaded TCH were applied in tooth extraction sockets. Bone loss and the expression of osteogenic regulatory factors (OPG, ALP, RANK) were evaluated. The immunoregulatory effects of ART-loaded TCH were observed through detecting the infiltration of T lymphocytes and their cytokines. The underlying mechanisms were explored. RESULTS: Results showed that the 150 mg/ml ART-loaded TCH group significantly ameliorated maxilla bone height and bone mineral density when compared with the T2DM group (p < 0.05). It also improved the expression of OPG, ALP, and RANK. Although the alteration of CD4+ T, CD8+ T, and CD4+:CD8+ T ratio has no significant difference among groups, the release of Th1 and Th2 in the 150 mg/ml ART-loaded TCH group has been significantly regulated than in the T2DM group (p < 0.05). Besides, ART-loaded TCH treatment inhibited the expression of p38 MAPK and ERK1 in T2DM maxilla. CONCLUSIONS: Therefore, the results indicated that 150 mg/ml ART-loaded TCH could be an effective method to prevent bone loss in T2DM tooth extraction rats by modulating the immunoregulation of Th1 and Th2 and the MAPK signaling pathway.

Absolute quantitation of binding antibodies from clinical samples.

The quantitation of antibody responses is a critical requirement for the successful development of vaccines and therapeutics that often relies on the use of standardized reference materials to determine relative quantities within biological samples. The validity of comparing responses across assays using arbitrarily defined reference values is therefore limited. We developed a generalizable method known as MASCALE (Mass Spectrometry Enabled Conversion to Absolute Levels of ELISA Antibodies) for absolute quantitation of antibodies by calibrating ELISA reference sera using mass spectrometry. Levels of proteotypic peptides served as a proxy for human IgG, allowing the conversion of responses from arbitrary values to absolute amounts. Applications include comparison of binding assays at two separate laboratories and evaluation of cross-clade magnitude-breadth responses induced by an investigational HIV-1 vaccine regimen. MASCALE addresses current challenges in the interpretation of immune responses in clinical trials and expands current options available to make suitable comparisons across different settings.

Effortful and effortless training of executive functions improve brain multiple demand system activities differently: an activation likelihood estimation meta-analysis of functional neuroimaging studies.

Both effortful and effortless training have been shown to be effective in enhancing individuals' executive functions. Effortful training improves domain-specific EFs, while effortless training improves domain-general EFs. Furthermore, effortful training has significantly higher training effects on EFs than effortless training. The neural mechanism underlying these different effects remained unclear. The present study conducted meta-analysis on neuroimaging studies to explore the changes of brain activations induced by effortful and effortless training. The results showed that effortful training induced greater activation in superior frontal gyrus, while effortless training induced greater activation in middle frontal gyrus, precuneus and cuneus. The brain regions of MD system enhanced by effortful training were more associated with core cognitive functions underlying EFs, while those enhanced by effortless training were more correlated with language functions. In addition, the significant clusters induced by effortful training had more overlaps with the MD system than effortless training. These results provided us with possibility to discuss the different behavioral results brought by effortful and effortless training.

[In-situ Remediation Effect of Cadmium-polluted Agriculture Land Using Different Amendments Under Rice-wheat Rotation].

Exploring the effects of one-time amendment treatments on cadmium (Cd)-contaminated farmland soils is beneficial for providing a theoretical basis to effectively prevent Cd pollution in farmland soils and ensure the safe production of crops. Five amendments, including straw biochar, fly ash, sepiolite, white marble powder, and shale (particle size <0.2 mm, application rate 2.25 kg·m-2), were applied to the Cd-contaminated farmland soils. The soil nutrients, pH, soil available Cd, and Cd chemical forms in the soils and grain Cd concentration in the planted crops were determined to investigate the effects and persistence of one-time applications of the five amendments. The results showed that:① the application of the five amendments had little effect on soil nutrient content, but all of them could increase soil pH. Amendment treatments improved the transfer of Cd from the acid extraction fraction to residue fraction and further reduced the Cd availability in the soil. The decreasing amplitudes of straw biochar and white marble powder soil conditioner were 20.42%-22.53%, which was higher than those in the other treatments. ② The grain Cd concentrations in rice and wheat were significantly decreased under the amendment treatments with the decreasing amplitudes of 19.88%-48.77% and 5.06%-24.00%, respectively. The Cd concentrations in rice grains under the treatments of straw biochar, fly ash, and white marble powder soil conditioner were 0.195, 0.196, and 0.223 mg·kg-1, respectively, which were lower than those under the other treatments and were close to or approached the National Standard of Food Safety(GB 2762-2017)(0.2 mg·kg-1). ③ The immobilization effects on Cd in farmland soils were decreasing with time under one-time application of the amendments. The available Cd concentrations in the soil and Cd concentrations in crop grains were still lower than those in the control after three rounds of rice-wheat rotation. The straw biochar and white marble powder soil conditioner had a good and long-term effect on reducing Cd availability in soils and Cd concentrations in crop grain, making them ideal materials for safe production in Cd-contaminated soils.

[Effects of Different Amendments on Cadmium Accumulation in Rice Safety in Cadmium-Contaminated Farmland Under Two Flooding Treatments].

Cadmium (Cd) accumulation in rice has become a serious public concern; thus, it is important to find an effective approach to reducing Cd accumulation in rice grains to ensure food safety. To investigate the effects of different amendments on Cd accumulation in rice in Cd-contaminated farmland under different flooding treatments, a field experiment with three amendments (jade powder, biochar, and fly ash) and two flooding treatments (intermittent flooding and flooding throughout the whole growth period) was conducted. The results showed that:① without amendment application, the soil pH significantly increased, whereas the soil available Cd concentration decreased by 3.81%-17.27% and 2.25%-6.74% with the treatments of flooding throughout the whole growth period and intermittent flooding, respectively. Additionally, the immobilizing efficiency of the treatment of flooding throughout the whole growth period was better than that of intermittent flooding; ② under different flooding treatments, amendment application improved soil pH, resulting in a decrease in the soil available Cd concentration along with an increase in the residual Cd concentration. Under the treatment of intermittent flooding, the soil pH increased by 0.19-2.20 units, and the soil available Cd concentration and Cd concentration in rice grains decreased by 4.72%-22.68% and 2.60%-75.75%, respectively, with the application of different amendments. Under the treatment of flooding throughout the whole growth period, the application of different amendments decreased the soil available Cd concentration and Cd concentration in rice grains by 5.06%-36.63% and 13.28%-77.01%, respectively. The immobilizing efficiency in both flooding treatments was jade powder > biochar > fly ash. ③ Under different flooding treatments, the application of amendments significantly reduced the soil available Cd concentration and Cd concentration in rice grains. Among the three amendments, jade powder showed the best capacity of immobilizing efficiency with the treatment of flooding throughout the whole growth period; the soil Cd reduction rates were 36.63% and 25.16%, and the Cd concentrations in rice grains were 0.058 and 0.170 mg·kg-1 in 2019 and 2020, respectively. The Cd concentrations in rice grains were within the limitation of the National Food Hygienic Standard of China. Therefore, combining flooding throughout the whole growth period with jade powder can be considered as an ideal strategy for ensuring rice safety in Cd-contaminated farmland.

A composite amendment benefits rice (Oryza sativa L.) safety and production in cadmium-contaminated soils by unique characteristics after oxidation modification.

In-situ immobilization is an effective strategy for Cd remediation and food safety, while some modifications are necessary to improve immobilization efficiency. In this study, a composite amendment (RFW) derived from rice straw biochar (RSB), fly ash (FA), and white marble (WM) was modified by oxidization (RFW-O) and pyrolysis (RFW-P). The RFW-O showed stronger Cd2+ sorption ability than RFW and RFW-P due to larger BET surface area and more oxygen containing-functional groups. Complexation and iron exchange were the two main processes of Cd2+ sorption on RFW-O. As a result, the application of RFW-O significantly reduced Cd availability in soils by 10.11-26.24% along with increased soil pH. It was found to be optimal to apply the RFW-O at a dosage of 2.5 wt% for 15 days before transplantation. After RFW-O application, Cd concentrations in brown rice decreased by 40.49% and 41.59% for pot and field experiment, respectively, and were less than 0.2 mg kg-1. The catalase, dehydrogenase, acid phosphatase and alkaline phosphatase activities in soils increased significantly. Moreover, RFW-O showed no significant effect on rice yield and quality. The RFW-O is thereby considered to be an ideal amendment for in-situ immobilization of Cd-contaminated soils for rice safety and production in practice.

Bipolar Hemiarthroplasty should not be selected as the primary option for intertrochanteric fractures in elderly patients aged 85 years or more.

This study aimed to compare the early outcome of proximal femoral nail antirotation (PFNA) and bipolar hemiarthroplasty (BPH) in elderly intertrochanteric fractures (ITFs) patients aged 85 years or more.This is a prospective cohort study, and we analyzed 120 elderly patients aged 85 years or more presented with ITFs who underwent BPH and PFNA between January 2017 and July 2018. 84 patients treated with PFNA were set as Group A, and 36 patients treated with BPH were set as Group B. Data such as gender, age, period of follow-up, fracture classification (according to Evans-Jensen classification), preoperative ASA (American Society of Anesthesiologists) physical status, interval between injury and operation, method of anaesthesia, duration of operation time, blood loss during surgery, time of weight bearing after operation, incidence of complications 2 weeks after operation, mortality rates and Harris Hip Score 12 months after operation were recorded and compared.There are no statistically significant differences when compared general data in patients from group A and B (P > .05). Operation time in Group A is less than Group B (103.33, 40-230 min vs 122.64, 75-180 minute, P < .01). Blood loss during surgery in Group A is less than Group B (70.24, 50-100 mL vs 194.44, 100-500 mL, P < .01). Time of weight bearing after operation in Group A is longer than Group B (50.70, 7-100 days vs 6.67, 4-14 days, P < .01). Incidence of complications 2 weeks after operation in Group A is less than Group B (14.12% vs 36.11%, P < .01). Mortality rates 12 months after operation in Group A is similar with Group B (13.10% vs 19.44%, P > .05). Harris Hip Score 12 months after operation in Group A is similar with Group B (64.64,0-91 points vs 64.41, 0-90 points, P > .05).Although BPH and PFNA have similar functional outcome and mortality rates 12 months after operation, BPH has more postoperative complications in elderly patients aged 85 years or more with ITFs, Bipolar Hemiarthroplasty should not be selected as the primary option for ITFs in elderly patients aged 85 years or more.

A small-molecule fusion inhibitor of influenza virus is orally active in mice.

Recent characterization of broadly neutralizing antibodies (bnAbs) against influenza virus identified the conserved hemagglutinin (HA) stem as a target for development of universal vaccines and therapeutics. Although several stem bnAbs are being evaluated in clinical trials, antibodies are generally unsuited for oral delivery. Guided by structural knowledge of the interactions and mechanism of anti-stem bnAb CR6261, we selected and optimized small molecules that mimic the bnAb functionality. Our lead compound neutralizes influenza A group 1 viruses by inhibiting HA-mediated fusion in vitro, protects mice against lethal and sublethal influenza challenge after oral administration, and effectively neutralizes virus infection in reconstituted three-dimensional cell culture of fully differentiated human bronchial epithelial cells. Cocrystal structures with H1 and H5 HAs reveal that the lead compound recapitulates the bnAb hotspot interactions.

Universal protection against influenza infection by a multidomain antibody to influenza hemagglutinin.

Broadly neutralizing antibodies against highly variable pathogens have stimulated the design of vaccines and therapeutics. We report the use of diverse camelid single-domain antibodies to influenza virus hemagglutinin to generate multidomain antibodies with impressive breadth and potency. Multidomain antibody MD3606 protects mice against influenza A and B infection when administered intravenously or expressed locally from a recombinant adeno-associated virus vector. Crystal and single-particle electron microscopy structures of these antibodies with hemagglutinins from influenza A and B viruses reveal binding to highly conserved epitopes. Collectively, our findings demonstrate that multidomain antibodies targeting multiple epitopes exhibit enhanced virus cross-reactivity and potency. In combination with adeno-associated virus-mediated gene delivery, they may provide an effective strategy to prevent infection with influenza virus and other highly variable pathogens.

Potent peptidic fusion inhibitors of influenza virus.

Influenza therapeutics with new targets and mechanisms of action are urgently needed to combat potential pandemics, emerging viruses, and constantly mutating strains in circulation. We report here on the design and structural characterization of potent peptidic inhibitors of influenza hemagglutinin. The peptide design was based on complementarity-determining region loops of human broadly neutralizing antibodies against the hemagglutinin (FI6v3 and CR9114). The optimized peptides exhibit nanomolar affinity and neutralization against influenza A group 1 viruses, including the 2009 H1N1 pandemic and avian H5N1 strains. The peptide inhibitors bind to the highly conserved stem epitope and block the low pH-induced conformational rearrangements associated with membrane fusion. These peptidic compounds and their advantageous biological properties should accelerate the development of new small molecule- and peptide-based therapeutics against influenza virus.

Relating influenza virus membrane fusion kinetics to stoichiometry of neutralizing antibodies at the single-particle level.

The ability of antibodies binding the influenza hemagglutinin (HA) protein to neutralize viral infectivity is of key importance in the design of next-generation vaccines and for prophylactic and therapeutic use. The two antibodies CR6261 and CR8020 have recently been shown to efficiently neutralize influenza A infection by binding to and inhibiting the influenza A HA protein that is responsible for membrane fusion in the early steps of viral infection. Here, we use single-particle fluorescence microscopy to correlate the number of antibodies or antibody fragments (Fab) bound to an individual virion with the capacity of the same virus particle to undergo membrane fusion. To this end, individual, infectious virus particles bound by fluorescently labeled antibodies/Fab are visualized as they fuse to a planar, supported lipid bilayer. The fluorescence intensity arising from the virus-bound antibodies/Fab is used to determine the number of molecules attached to viral HA while a fluorescent marker in the viral membrane is used to simultaneously obtain kinetic information on the fusion process. We experimentally determine that the stoichiometry required for fusion inhibition by both antibody and Fab leaves large numbers of unbound HA epitopes on the viral surface. Kinetic measurements of the fusion process reveal that those few particles capable of fusion at high antibody/Fab coverage display significantly slower hemifusion kinetics. Overall, our results support a membrane fusion mechanism requiring the stochastic, coordinated action of multiple HA trimers and a model of fusion inhibition by stem-binding antibodies through disruption of this coordinated action.

APC/C is an essential regulator of centrosome clustering.

Centrosome amplification has been extensively associated with cancer. Cancer cells with extra centrosomes have the ability to cluster the extra centrosomes and divide in a bipolar fashion. Although a number of proteins have been shown to be involved in centrosome clustering, a mechanistic understanding of how this process is coordinated is not yet well defined. Here, to reveal regulators of centrosome clustering, we perform small interfering RNA (siRNA) screens with multiple assay readouts in a human isogenic cellular model. We find that APC/C activity is essential for centrosome clustering. We show that the motor kinesin Eg5 is a substrate of APC/C-CDH1, and that inhibition of APC/C results in stabilization of Eg5. Increased Eg5 protein levels disturb the balance of forces on the spindle and prevent centrosome clustering. This process is completely reversed after a short treatment with the Eg5 inhibitor, monastrol. These data advance our understanding of the regulation of centrosome clustering.

Mechanisms of hemagglutinin targeted influenza virus neutralization.

Human monoclonal antibodies have been identified which neutralize broad spectra of influenza A or B viruses. Here, we dissect the mechanisms by which such antibodies interfere with infectivity. We distinguish four mechanisms that link the conserved hemagglutinin (HA) epitopes of broadly neutralizing antibodies to critical processes in the viral life cycle. HA-stem binding antibodies can act intracellularly by blocking fusion between the viral and endosomal membranes and extracellularly by preventing the proteolytic activation of HA. HA-head binding antibodies prevent viral attachment and release. These insights into newly identified ways by which the human immune system can interfere with influenza virus infection may aid the development of novel universal vaccines and antivirals.

Aurora A kinase regulates mammary epithelial cell fate by determining mitotic spindle orientation in a Notch-dependent manner.

Cell fate determination in the progeny of mammary epithelial stem/progenitor cells remains poorly understood. Here, we have examined the role of the mitotic kinase Aurora A (AURKA) in regulating the balance between basal and luminal mammary lineages. We find that AURKA is highly expressed in basal stem cells and, to a lesser extent, in luminal progenitors. Wild-type AURKA expression promoted luminal cell fate, but expression of an S155R mutant reduced proliferation, promoted basal fate, and inhibited serial transplantation. The mechanism involved regulation of mitotic spindle orientation by AURKA and the positioning of daughter cells after division. Remarkably, this was NOTCH dependent, as NOTCH inhibitor blocked the effect of wild-type AURKA expression on spindle orientation and instead mimicked the effect of the S155R mutant. These findings directly link AURKA, NOTCH signaling, and mitotic spindle orientation and suggest a mechanism for regulating the balance between luminal and basal lineages in the mammary gland.

Aurora-A expressing tumour cells are deficient for homology-directed DNA double strand-break repair and sensitive to PARP inhibition.

The protein kinase Aurora-A is a major regulator of the cell cycle that orchestrates mitotic entry and is required for the assembly of a functional mitotic spindle. Overexpression of Aurora-A has been strongly linked with oncogenesis and this has led to considerable efforts at therapeutic targeting of the kinase activity of this protein. However, the exact mechanism by which Aurora-A promotes oncogenesis remains unclear. Here, we show that Aurora-A modulates the repair of DNA double-strand breaks (DSBs). Aurora-A expression inhibits RAD51 recruitment to DNA DSBs, decreases DSB repair by homologous recombination and sensitizes cancer cells to PARP inhibition. This impairment of RAD51 function requires inhibition of CHK1 by Polo-like kinase 1 (PLK1). These results identify a novel function of Aurora-A in modulating the response to DNA DSB that likely contributes to carcinogenesis and suggest a novel therapeutic approach to the treatment of cancers overexpressing this protein.

A cancer-associated aurora A mutant is mislocalized and misregulated due to loss of interaction with TPX2.

Mutations in protein kinases can drive cancer through alterations of the kinase activity or by uncoupling kinase activity from regulation. Changes to protein expression in Aurora A, a mitotic Ser/Thr kinase, are associated with the development of several human cancers, but the effects of somatic cancer-associated mutations have not been determined. In this study we show that Aurora A kinase activity is altered in different ways in three somatic cancer-associated mutations located within the catalytic domain; Aurora A(V174M) shows constitutively increased kinase activity, Aurora A(S155R) activity is decreased primarily due to misregulation, and Aurora A(S361*) activity is ablated due to loss of structural integrity. These alterations suggest vastly different mechanisms for the role of these three mutations in human cancer. We have further characterized the Aurora A(S155R) mutant protein, found that its reduced cellular activity and mislocalization are due to loss of interaction with TPX2, and deciphered the structural basis of the disruption at 2.5 A resolution. Previous studies have shown that disruption of the Aurora A/TPX2 interaction results in defective spindles that generate chromosomal abnormalities. In a panel of 40 samples from microsatellite instability-positive colon cancer patients, we found one example in which the tumor contained only Aurora A(S155R), whereas the normal tissue contained only wild-type Aurora A. We propose that the S155R mutation is an example of a somatic mutation associated with this tumor type, albeit at modest frequency, that could promote aneuploidy through the loss of regulated interactions between Aurora A and its protein partners.